HK1262206A1

HK1262206A1 - Use of mgbg for manufacturing a medicament for the treatment or prevention of progressive ms and its progression

Info

Publication number: HK1262206A1
Application number: HK19122133.2A
Authority: HK
Inventors: J·布利策; J·麦基尔恩
Original assignee: 帕萨罗杰卡有限公司
Priority date: 2013-01-08
Filing date: 2019-04-11
Publication date: 2020-01-10

Description

Use of MGBG in the preparation of a medicament for the treatment or prevention of progressive MS and its progression

The present application is a divisional application filed on application date 2014, No. 01, No. 08, international application number PCT/US2014/010714, national application number 201480007129.3, entitled "methods and compositions for treating demyelinating diseases".

This application claims benefit of priority from U.S. provisional application No. 61/750,336 filed on 8.1.2013 and application No. 61/823,276 filed on 14.5.2013, the disclosures of which are incorporated by reference in their entirety as if written herein.

Disclosed herein are novel oral pharmaceutical compositions of SAMDC inhibitors, polyamine analogs, and polyamine biosynthesis inhibitors, and their use in treating conditions including demyelinating diseases, autoimmune disorders affecting the nervous system, and other neurodegenerative conditions.

MGBG (methylglyoxal bis (amidinohydrazone); mitoguazone hydrazone) is a competitive polyamine inhibitor of S-adenosylmethionine decarboxylase (SAMDC, AMD-I) which catalyzes the synthesis of spermidine, a polyamine. Amino acid-derived polyamines have long been associated with cell growth and cancer, and specific oncogenes and tumor suppressor genes regulate polyamine metabolism. Inhibition of polyamine synthesis has been shown to be generally ineffective as an anti-cancer strategy in clinical trials, but is an effective cancer chemoprevention strategy in preclinical studies. Despite its new mechanism of action and promising preclinical data, the initial clinical trial of MGBG ceased in the mid-60's of the 20 th century due to severe toxicity (especially to self-renewing normal tissues such as bone marrow and intestinal tract, especially severe mucositis), both cases being dose and regimen dependent.

In any event, studies on MGBG continue. Many studies have examined the potential use in combination with other chemotherapeutic agents and innovative dosing regimens, designed to minimize side effects and doses when possible. Other studies have focused on elucidating the mode of action of MGBG in vivo. There are also studies investigating the activity of MGBG in non-cancer diseases.

Up to now MGBG was limited to intravenous use, perhaps as a result of negative clinical studies in these early studies. In fact, this presents numerous problems with the treatment of many diseases, especially chronic or recurrent pathologies. Administration via IV injection or infusion must be done by a medical professional in a hospital setting. This not only causes inconvenience and increased costs to the subject, but also exposes him or her to nosocomial infections and diseases, the latter due to venipuncture and visit to a hospital or clinic per se. In immunocompromised individuals, the individual undergoesTreatment with inhibitors of the immune system is indicated and in the elderly this is a related problem. Thus, a subject suffering from a long-term chronic condition, such as an autoimmune or hyperproliferative disorder, or a physician treating the subject, may find the cost, inconvenience, and risk of the treatment more important than any potential therapeutic benefit that a drug may provide. In addition, intravenous infusion drugs have a high C as opposed to more slowly absorbed oral drugs_{Maximum of}And T_{Maximum of}The adverse conditions of (1).

In contrast, oral formulations of MGBG would present several benefits. First, oral formulations, such as simple pills or tablets, can be taken outside of a hospital environment, thereby increasing the likelihood of ease of use and compliance by the subject. This allows the subject to avoid the risk of infection with IV administration and hospital visits. This is of particular benefit when early treatment can prevent the development of disease complications. Long-term low dose administration of MGBG is almost impossible in IV formulations. In addition, oral delivery typically avoids the high concentration peaks and rapid clearance associated with IV bolus doses. A further advantage of oral medicaments is the ability to formulate MGBG as a combined composition with one or more other therapeutic agents.

Brief Description of Drawings

Figure 1 shows the mean clinical scores of subjects in the first 28-day murine model of Experimental Autoimmune Encephalomyelitis (EAE) (a chronic progressive model of MS) induced by vaccination with Myelin Oligodendrocyte Glycoprotein (MOG) peptide, where test subjects were administered vehicle, 30mpk MGBG (twice daily) or 3mpk fingolimod (once daily), and compared to mock-immunized subjects.

Figure 2 shows the mean percent weight change (relative to study start) for subjects in the first 28-day MOG EAE model, where test subjects were administered vehicle, 30mpk MGBG (twice daily) or 3mpk fingolimod (once daily), and compared to mock-immunized subjects.

Figure 3 shows a reduction in spinal cord inflammation as measured by the number of inflammatory foci in a spinal cord histopathological sample from a 28-day MOG EAE model, in which test subjects were administered vehicle, 30mpk MGBG (twice daily) or 3mpk fingolimod (once daily), and compared to mock-immunized subjects.

Figure 4 shows the reduction of spinal cord demyelination in histopathological samples from the 28-day MOG EAE model, where test subjects were administered vehicle, 30mpk MGBG (twice daily) or 3mpk fingolimod (once daily), and compared to mock-immunized subjects.

Figure 5 shows a reduction in apoptosis in spinal cord histopathology samples from a 28-day MOG EAE model, in which test subjects were administered vehicle, 30mpk MGBG (twice daily) or 3mpk fingolimod (once daily), and compared to mock-immunized subjects.

Figure 6 shows the mean clinical scores of subjects in the second 28-day murine MOG EAE model, where test subjects were administered vehicle, 30mpk MGBG (twice daily) or 1mpk fingolimod (once daily), and compared to mock-immunized subjects; MGBG inhibited EAE equally well as 1mpk fingolimod.

Figure 7 shows the mean percent weight change (relative to study start) for subjects in the second 28-day MOG EAE model, where test subjects were administered vehicle, 30mpk MGBG (twice daily) or 1mpk fingolimod (once daily), and compared to mock-immunized subjects.

Figure 8 shows that MGBG, but not fingolimod, reduces CNS a) CD11c + dendritic cells associated with antigen presentation and B) IL12+ dendritic cells associated with promoting Th1 responses.

Figure 9 shows that MGBG but not fingolimod reduces a) Th1 and B) M1 macrophages.

Figure 10 shows fingolimod preferentially a) inhibits Th17 and B) increases M2 macrophages; therefore, MGBG and fingolimod have distinct but therapeutically complementary activities in MS.

Figure 11 shows that although plasma levels of MGBG were barely detectable by 28 hours after a single dose into rats, MGBG levels in spleen and liver remained detectably higher even 48 hours after dosing. This is consistent with the selective uptake mechanism of MGBG.

Figure 12 shows the daily mean clinical score of subjects in the third 28-day murine MOG EAE model, where fingolimod was administered at 0.1mpk (once daily); MGBG (twice daily) administered at 30mpk inhibited EAE equally well as 0.1mpk fingolimod, and the combination of MGBG and fingolimod completely inhibited the development of EAE throughout the course of the study (none of the animals suffered from disease under combination therapy).

Figure 13 shows the cumulative mean clinical scores of subjects in the third EAE study, further illustrating the significance of the above results. MGBG at 30mpk (twice daily) and fingolimod at 0.1mpk (once daily) were equally effective in preventing EAE development, and the combination of the two completely inhibited EAE development.

Figure 14 shows the weight change of the subject (relative to the start of the study) in the EAE model on the third 28 days, confirming the results in figure 13.

Figure 15 shows reduction of spinal cord inflammation as measured by the number of inflammatory foci in spinal cord histopathology samples from the third EAE model. Furthermore, MGBG at 30mpk (twice daily) and fingolimod at 0.1mpk (once daily) were equally effective in reducing the number of inflammatory foci, and the combination of both apparently completely inhibited their development.

Figure 16 shows a reduction in apoptosis in spinal cord histopathological samples from the third EAE model. Furthermore, 30mpk of MGBG (twice daily) and 0.1mpk of fingolimod (once daily) were equally effective in reducing apoptosis, and the combination of both clearly completely inhibited apoptosis.

Fig. 17 shows a reduction in spinal cord demyelination in histopathological samples from the third EAE model. Furthermore, 30mpk of MGBG (twice daily) and 0.1mpk of fingolimod (once daily) were equally effective in reducing demyelination, and the combination of the two clearly completely prevented demyelination.

Summary of The Invention

Accordingly, provided herein is a method of treating or preventing a demyelinating disease or symptoms thereof, comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the demyelinating disease is selected from multiple sclerosis, optic neuritis, idiopathic inflammatory demyelinating diseases, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, transverse myelitis, Barlow's concentric sclerosis, pernicious anemia, mid-pontine myelination, tabes spinalis, neuromyelitis optica (NMO), Progressive Multifocal Leukoencephalopathy (PML), anti-MAG (myelin-associated glycoprotein) neuropathy, hereditary motor and sensory neuropathy (peroneal muscular atrophy disease), tendonomatosis (cereebronenious xanthomatosis), and leukodystrophy, including adrenoleukodystrophy, adrenomyeloneuropathy, metachromatic leukodystrophy, globuloid cellular leukodystrophy (krabbe's disease), canavan disease, effaceous leukosis, alexander disease, refsum disease, and pelizaeus-merzbach disease.

In certain embodiments, the demyelinating disease is multiple sclerosis.

In certain embodiments, the methods further comprise administering an agent selected from interferon β -1a, interferon β -1b, glatiramer acetate, mitoxantrone, natalizumab, fingolimod, laquinimod, dimethyl fumarate, and teriflunomide.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

Also provided herein are embodiments wherein each embodiment in paragraphs [025] - [036] above is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

Also provided herein is a method of treating a symptom of an autoimmune disease affecting the nervous system comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the symptom is selected from CNS inflammation, demyelination, and paralysis.

In certain embodiments, the autoimmune disease affecting the nervous system is selected from multiple sclerosis, polymyalgia, myasthenia gravis, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, transverse myelitis, Barlow's concentric sclerosis, pernicious anemia, acute disseminated encephalomyelitis (ADME), Amyotrophic Lateral Sclerosis (ALS), autoimmune peripheral neuropathy, lupus erythematosus, psoriatic arthritis, rheumatoid arthritis, osteoarthritis, and rheumatic fever.

In certain embodiments, the autoimmune disease that affects a neurological disease is multiple sclerosis.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

Also provided herein are embodiments wherein each embodiment in paragraphs [038] - [050] above is combined with one or more other non-contradictory embodiments such that a resulting embodiment includes two or more of the recited elements and/or limitations.

Also provided herein is a method of treating or preventing a demyelinating disease comprising the simultaneous administration of a SAMDC inhibitor and an agent selected from interferon β -1a, interferon β -1b, glatiramer acetate, mitoxantrone, natalizumab, fingolimod, dimethyl fumarate, and teriflunomide.

In certain embodiments, the other agent is fingolimod.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the demyelinating disease is selected from multiple sclerosis, guillain-barre syndrome, chronic inflammatory demyelinating polyneuropathy, transverse myelitis, baro-concentric sclerosis, pernicious anemia, mid-pontine myelinolysis, tabes spinosus, neuromyelitis optica (NMO), Progressive Multifocal Leukoencephalopathy (PML), anti-MAG (myelin associated glycoprotein) neuropathy, hereditary motor and sensory neuropathy (peroneal atrophy disease), tendonoxanthomatosis, and leukodystrophy including adrenoleukodystrophy, adrenomyeloneuropathy, metachromatic leukodystrophy, globoid cellular leukodystrophy (krabbe disease), canavan disease, evaporative leukosis, alexander disease, refsum disease, and pelizaeus-merzbach disease.

In certain embodiments, the demyelinating disease is multiple sclerosis.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

Also provided herein are embodiments in which each embodiment in paragraphs [052] - [062] above is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

Also provided herein is a pharmaceutical composition comprising a SAMDC inhibitor and another agent selected from interferon β -1a, interferon β -1b, glatiramer acetate, mitoxantrone, natalizumab, fingolimod, and teriflunomide, and a pharmaceutically acceptable carrier.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the other agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

In certain embodiments, a pharmaceutical formulation comprising a SAMDC inhibitor and fingolimod is formulated for oral administration.

In certain embodiments, the pharmaceutical formulation produces therapeutically effective systemic plasma MGBG levels when orally administered to a subject.

In certain embodiments, the pharmaceutical formulation is formulated for once daily administration.

In certain embodiments, the pharmaceutical formulation comprises 0.5mg fingolimod per dosage unit.

In certain embodiments, the pharmaceutical formulation comprises less than 0.5mg fingolimod per dosage unit.

In certain embodiments, the pharmaceutical formulation comprises 0.25mg fingolimod per dosage unit.

In certain embodiments, the pharmaceutical formulation is formulated for administration twice daily.

In certain embodiments, the pharmaceutical formulation comprises less than 0.25mg fingolimod per dosage unit.

In certain embodiments, the pharmaceutical formulation comprises about 0.125mg fingolimod per dosage unit.

Also provided herein are embodiments wherein each embodiment in paragraphs [064] - [081] above is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

Also provided is a method of preventing relapse or progression of a demyelinating disease in a patient, or reducing the severity of relapsed symptoms, comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, there is provided a method of preventing relapse or progression of a demyelinating disease in a patient, or reducing the severity of relapsing symptoms, comprising administering a therapeutically effective amount of MGBG.

In certain embodiments, the demyelinating disease is selected from multiple sclerosis, optic neuritis, idiopathic inflammatory demyelinating disease, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, transverse myelitis, Barlow's concentric sclerosis, pernicious anemia, mid-pontine myelination, tabes dorsalis, neuromyelitis optica (NMO), Progressive Multifocal Leukoencephalopathy (PML), anti-MAG (myelin-associated glycoprotein) neuropathy, hereditary motor and sensory neuropathy (peroneal muscle wasting disease), tendonomatosis, and leukodystrophy, including adrenoleukodystrophy, adrenomyeloneuropathy, metachromatic leukodystrophy, globuloid cellular leukodystrophy (krabbe's disease), canavan disease, effaceous leukosis, alexander disease, refsum disease, and pelizaeus-merzbach disease.

In certain embodiments, the demyelinating disease is multiple sclerosis.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the SAMDC inhibitor is not a T cell modulator.

In certain embodiments, the administration occurs with a reduced incidence of at least one side effect selected from the group consisting of: cytopenia, nephrotoxicity, hepatotoxicity, cardiotoxicity, teratogenicity, impaired lung function, macular edema, peripheral neuropathy, severe skin reactions, increased risk of infection (including latent bacteria and viruses), innate immune damage, adaptive immune damage, and flushing.

In certain embodiments, the demyelinating disease is multiple sclerosis.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

In certain embodiments, the administration occurs with a decrease in the incidence of at least one side effect selected from the group consisting of: cytopenia, renal toxicity, hepatotoxicity, cardiotoxicity and teratogenicity.

In certain embodiments, the cytopenia is selected from the group consisting of lymphopenia and neutropenia.

In certain embodiments, the administration occurs with a reduction in the incidence of at least two side effects selected from the group consisting of: cytopenia, renal toxicity, hepatotoxicity, cardiotoxicity and teratogenicity.

In certain embodiments, administration occurs with a reduced incidence of cytopenia, nephrotoxicity and hepatotoxicity.

In certain embodiments, the administration occurs additionally with a reduced incidence of cardiotoxicity and teratogenicity.

Also provided herein are embodiments wherein each embodiment in the above paragraphs [083] - [0105] is combined with one or more other non-contradictory embodiments such that a resulting embodiment comprises two or more of the recited elements and/or limitations.

Also provided is a method of treating progressive multiple sclerosis in a patient comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the progressive multiple sclerosis is primary progressive.

In certain embodiments, the progressive multiple sclerosis is secondary progressive.

In certain embodiments, the progressive multiple sclerosis is progressive relapsing.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

In certain embodiments, the treatment prevents relapse or progression of MS.

In certain embodiments, the SAMDC inhibitor is not a T cell modulator.

In certain embodiments, the demyelinating disease is multiple sclerosis.

Also provided herein are embodiments wherein each embodiment in the above paragraphs [0107] - [0130] is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

Also provided is a method of blocking antigen presentation on cells of a patient having a demyelinating disease, wherein the antigen is derived from a mimetic or similar antigen in the myelin sheath, comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, the cells are of myeloid lineage.

In certain embodiments, the cell is pro-inflammatory.

In certain embodiments, the cell is selected from a dendritic cell, a macrophage, and a B cell.

In certain embodiments, the cell is a dendritic cell.

In certain embodiments, the cell is a M1 macrophage.

In certain embodiments, the demyelinating disease is multiple sclerosis.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

In certain embodiments, the SAMDC inhibitor is not a T cell modulator.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the demyelinating disease is multiple sclerosis.

Also provided herein are embodiments wherein each embodiment in the above paragraphs [0132] - [0157] is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

Also provided is a method of inhibiting infiltration of antigen presenting cells into the central nervous system of a patient having a demyelinating disease, comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, the cell is pro-inflammatory.

In certain embodiments, the cell is a dendritic cell.

In certain embodiments, the cell is a M1 macrophage.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

The method of any of claims 76-80, wherein the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

In certain embodiments, the demyelinating disease is multiple sclerosis.

Also provided herein are embodiments wherein each embodiment in the above paragraphs [0159] - [0181] is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

Also provided is a method of preventing or reducing the severity of an autoimmune response in an initial stage of a patient having a demyelinating disease comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, the administering additionally prevents or reduces an autoimmune response at an expansion stage in a patient having a demyelinating disease.

In certain embodiments, the demyelinating disease is multiple sclerosis.

In certain embodiments, the SAMDC inhibitor is not a T cell modulator.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

In certain embodiments, the demyelinating disease is multiple sclerosis.

Also provided herein are embodiments wherein each embodiment in the above paragraphs [0183] - [0204] is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

Also provided is a method of treating or preventing demyelination in a subject in need thereof, comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the demyelinating disease is multiple sclerosis.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

Also provided herein are embodiments wherein each embodiment in the above paragraphs [0206] - [0217] is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

Also provided is a method of reducing the severity of apoptosis in a neural tissue of a subject in need thereof, comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the demyelinating disease is multiple sclerosis.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

Also provided herein are embodiments wherein each embodiment in paragraphs [0219] - [0230] above is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

Also provided is a method of preventing or slowing the development of an inflammatory lesion in a neural tissue of a subject in need thereof, comprising administering a therapeutically effective amount of a SAMDC inhibitor.

In certain embodiments, the SAMDC inhibitor is selectively taken into the cell.

In certain embodiments, the SAMDC inhibitor is MGBG.

In certain embodiments, the administration of MGBG is oral.

In certain embodiments, MGBG is administered at 20 mg/day to 400 mg/day.

In certain embodiments, the demyelinating disease is multiple sclerosis.

In certain embodiments, the agent is fingolimod.

In certain embodiments, fingolimod is administered at 0.5mg per day.

In certain embodiments, fingolimod is administered at less than 0.5mg per day.

In certain embodiments, fingolimod is administered at 0.25mg per day.

Also provided herein are embodiments wherein each embodiment in paragraphs [0232] - [0243] above is combined with one or more other non-contradictory embodiments such that the resulting embodiments include two or more of the recited elements and/or limitations.

The present disclosure includes not only the methods disclosed above, but also:

the corresponding use of the above compounds in the treatment of diseases and in the specific diseases discussed; and

the corresponding use of the above compounds in the manufacture of a medicament for the treatment of the diseases in question.

To avoid repetition, the embodiments are not explicitly rewritten herein. However, these embodiments should be understood as being included as if written as such. For example, the present disclosure provides:

use of a therapeutically effective amount of a SAMDC inhibitor for the treatment or prevention of a demyelinating disease;

use of a therapeutically effective amount of a SAMDC inhibitor for the treatment of symptoms of an autoimmune disease involving the nervous system;

use of a therapeutically effective amount of a SAMDC inhibitor and an agent selected from interferon β -1a, interferon β -1b, glatiramer acetate, mitoxantrone, natalizumab, fingolimod, dimethyl fumarate, and teriflunomide in the treatment or prevention of a demyelinating disease;

use of a therapeutically effective amount of a SAMDC inhibitor for preventing the recurrence or progression of a demyelinating disease or reducing the severity of recurring symptoms of a demyelinating disease;

use of a therapeutically effective amount of a SAMDC inhibitor for treating progressive multiple sclerosis in a patient;

use of a therapeutically effective amount of a SAMDC inhibitor for blocking antigen presentation on cells of a patient having a demyelinating disease, wherein the antigen is derived from a mimetic or similar antigen in the myelin sheath;

use of a therapeutically effective amount of a SAMDC inhibitor for inhibiting infiltration of antigen presenting cells into the central nervous system of a patient suffering from a demyelinating disease;

a method of preventing or reducing the severity of an initial stage autoimmune response in a patient with a demyelinating disease comprising administering a therapeutically effective amount of a SAMDC inhibitor.

Use of a therapeutically effective amount of a SAMDC inhibitor for treating or preventing demyelination in a subject in need thereof;

use of a therapeutically effective amount of a SAMDC inhibitor for reducing the severity of apoptosis in a neural tissue of a subject in need thereof; and/or

Use of a therapeutically effective amount of a SAMDC inhibitor for preventing or slowing the development of an inflammatory lesion in a neural tissue of a subject in need thereof;

and all the dependent embodiments listed in paragraphs [025] - [0243] above and in the claims below. The present disclosure also provides:

use of a SAMDC inhibitor in the manufacture of a medicament for the treatment or prevention of a demyelinating disease;

use of a SAMDC inhibitor for the manufacture of a medicament for the treatment of symptoms of an autoimmune disease affecting the nervous system;

use of a SAMDC inhibitor and an agent selected from interferon β -1a, interferon β -1b, glatiramer acetate, mitoxantrone, natalizumab, fingolimod, dimethyl fumarate, and teriflunomide in the manufacture of a medicament for the treatment or prevention of a demyelinating disease;

use of an inhibitor of SAMDC in the manufacture of a medicament for preventing the recurrence or progression of a demyelinating disease or reducing the severity of recurring symptoms of a demyelinating disease;

use of a SAMDC inhibitor in the manufacture of a medicament for treating progressive multiple sclerosis in a patient;

use of a SAMDC inhibitor in the manufacture of a medicament for blocking antigen presentation on cells of a patient having a demyelinating disease, wherein the antigen is derived from a mimetic or similar antigen in the myelin sheath;

use of a SAMDC inhibitor in the manufacture of a medicament for inhibiting infiltration of antigen presenting cells into the central nervous system of a patient suffering from a demyelinating disease;

a method of manufacturing an agent for preventing or reducing the severity of an autoimmune response in an initial stage of a patient with a demyelinating disease comprising administering a SAMDC inhibitor.

Use of a SAMDC inhibitor in the manufacture of a medicament for treating or preventing demyelination in a subject in need thereof;

use of a SAMDC inhibitor in the manufacture of a medicament for reducing the severity of apoptosis in a neural tissue of a subject in need thereof; and/or

Use of a SAMDC inhibitor in the manufacture of a medicament for preventing or slowing the development of an inflammatory lesion in a neural tissue of a subject in need thereof;

and all the dependent embodiments listed in paragraph [00]24- [0242] above and in the claims below.

Also disclosed herein are oral pharmaceutical formulations of MGBG and other SAMDC polyamine analogs, polyamine biosynthesis inhibitors, and polyamine inhibitors. Also disclosed are methods for treating diseases comprising administering MGBG and other SAMDC polyamine analogs, polyamine biosynthesis inhibitors, and polyamine inhibitors.

Also provided herein are methods for treating pain comprising administering MGBG and other SAMDC polyamine analogs, polyamine biosynthesis inhibitors, and polyamine inhibitors.

Also provided herein is a pharmaceutical composition for oral delivery comprising a polyamine analog or a polyamine biosynthesis inhibitor and at least one pharmaceutically acceptable oral excipient.

Also provided herein is an oral pharmaceutical composition comprising a polyamine analog or polyamine biosynthesis inhibitor and at least one oral pharmaceutically acceptable excipient that results in therapeutically effective systemic plasma polyamine analog or polyamine biosynthesis inhibitor levels when orally administered to a subject.

Also provided herein is an oral pharmaceutical composition comprising a polyamine analog or polyamine biosynthesis inhibitor and at least one oral pharmaceutically acceptable excipient that when orally administered to a subject results in a therapeutically effective systemic plasma polyamine analog or polyamine biosynthesis inhibitor level for treating pain.

In certain embodiments, the polyamine analog or polyamine biosynthesis inhibitor is a compound disclosed herein.

In certain embodiments, polyamine analogs or polyamine biosynthesis inhibitors are known in the art.

In certain embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma polyamine analog or polyamine biosynthesis inhibitor levels for a period of at least 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 30, 36, or 48 hours. In other embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma polyamine analog or polyamine biosynthesis inhibitor levels for a period of at least 6 hours. In other embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma polyamine analog or polyamine biosynthesis inhibitor levels for a period of at least 12 hours. In other embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma polyamine analog or polyamine biosynthesis inhibitor levels for a period of at least 18 hours. In other embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma polyamine analog or polyamine biosynthesis inhibitor levels for a period of at least 24 hours.

In certain embodiments, the pharmaceutical composition produces a polyamine analog or polyamine biosynthesis inhibitor plasma level that has a peak plasma concentration of at least 25, 50, 55, 60, 65, 75, 80, 85, 90, or 95% for at least 4 hours. In certain embodiments, the pharmaceutical composition produces a polyamine analog or polyamine biosynthesis inhibitor plasma level of at least 75% of the peak plasma concentration for at least 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, or 24 hours. In certain embodiments, the pharmaceutical composition produces a plasma level of the polyamine analog or polyamine biosynthesis inhibitor of at least 75% of the peak plasma concentration for at least 4 hours. In certain embodiments, the pharmaceutical composition produces a plasma level of the polyamine analog or polyamine biosynthesis inhibitor of at least 75% of the peak plasma concentration for at least 6 hours. In certain embodiments, the pharmaceutical composition produces a plasma level of the polyamine analog or polyamine biosynthesis inhibitor of at least 75% of the peak plasma concentration for at least 8 hours. In certain embodiments, the pharmaceutical composition produces a polyamine analog or polyamine biosynthesis inhibitor plasma level of at least 50% of the peak plasma concentration for at least 8 hours. In certain embodiments, the pharmaceutical composition produces a polyamine analog or polyamine biosynthesis inhibitor plasma level of at least 50% of the peak plasma concentration for at least 12 hours. In certain embodiments, the pharmaceutical composition produces a polyamine analog or polyamine biosynthesis inhibitor plasma level of at least 50% of the peak plasma concentration for at least 18 hours. In certain embodiments, the pharmaceutical composition produces a plasma level of the polyamine analog or polyamine biosynthesis inhibitor of at least 25% of the peak plasma concentration for at least 18 hours. In other embodiments, the peak plasma concentration is a therapeutically effective concentration. In other embodiments, the percentage of peak plasma concentration is therapeutically effective over a given period of time.

In certain embodiments, a pharmaceutical composition comprising a polyamine analog or a polyamine biosynthesis inhibitor has an oral bioavailability of at least 10, 20, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 45, 50, 55, or 60%. In other embodiments, the pharmaceutical composition has an oral bioavailability of at least 10%, 20%, 25%, 30%, 35%, 40%, or 45%. In other embodiments, the pharmaceutical composition has an oral bioavailability of at least 30%, at least 35%, at least 40%, or at least 45%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 20%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 30%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 35%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 40%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 45%. In certain embodiments, the pharmaceutical composition has an oral bioavailability that results in therapeutically effective plasma polyamine analog or polyamine biosynthesis inhibitor levels in a subject administered once daily for a period of at least 24 hours. In certain embodiments, the pharmaceutical composition has an oral bioavailability that results in therapeutically effective plasma polyamine analog or polyamine biosynthesis inhibitor levels in a subject administered twice daily for a period of at least 24 hours. In certain embodiments, the pharmaceutical composition has an oral bioavailability that results in therapeutically effective plasma polyamine analog or polyamine biosynthesis inhibitor levels for a period of at least 24 hours in a subject administered three times per day.

In certain embodiments, the pharmaceutical composition comprising a polyamine analog or a polyamine biosynthesis inhibitor has a half-life of at least 4, 6, 8, 10, 12, 14, 16, 18, 19, 20, 21, 22, 23, 24, 26, 28, 30, or 36 hours. In certain embodiments, the pharmaceutical composition has a half-life of at least 12 hours. In other embodiments, the pharmaceutical composition has a half-life of at least 18 hours. In other embodiments, the pharmaceutical composition has a half-life of at least 20 hours. In other embodiments, the pharmaceutical composition has a half-life of at least 24 hours. In certain embodiments, the pharmaceutical composition has a half-life of at least 48, 72, 96, or 120 hours.

Additionally, provided herein is a pharmaceutical composition for oral delivery comprising MGBG and at least one pharmaceutically acceptable oral excipient.

Also provided herein is an oral pharmaceutical composition comprising MGBG and at least one oral pharmaceutically acceptable excipient, which when orally administered to a subject results in a therapeutically effective systemic plasma MGBG level.

Also provided herein is an oral pharmaceutical composition comprising MGBG and at least one oral pharmaceutically acceptable excipient that when orally administered to a subject results in a therapeutically effective systemic plasma MGBG level for treating pain.

In certain embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma MGBG levels for a period of at least 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 30, 36, or 48 hours. In other embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma MGBG levels for a period of at least 6 hours. In other embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma MGBG levels for a period of at least 12 hours. In other embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma MGBG levels for a period of at least 18 hours. In other embodiments, the pharmaceutical composition produces therapeutically effective systemic plasma MGBG levels for a period of at least 24 hours.

In certain embodiments, the pharmaceutical composition produces MGBG plasma levels for at least 4 hours at a peak plasma concentration of at least 25, 50, 55, 60, 65, 75, 80, 85, 90, or 95%. In certain embodiments, the pharmaceutical composition produces MGBG plasma levels for at least 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, or 24 hours at a peak plasma concentration of at least 75%. In certain embodiments, the pharmaceutical composition produces MGBG plasma levels at a peak plasma concentration of at least 75% for at least 4 hours. In certain embodiments, the pharmaceutical composition produces MGBG plasma levels for at least 6 hours at a peak plasma concentration of at least 75%. In certain embodiments, the pharmaceutical composition produces MGBG plasma levels for at least 8 hours at a peak plasma concentration of at least 75%. In certain embodiments, the pharmaceutical composition produces MGBG plasma levels for at least 8 hours at a peak plasma concentration of at least 50%. In certain embodiments, the pharmaceutical composition produces MGBG plasma levels for at least 12 hours at a peak plasma concentration of at least 50%. In certain embodiments, the pharmaceutical composition produces MGBG plasma levels for at least 18 hours at a peak plasma concentration of at least 50%. In certain embodiments, the pharmaceutical composition produces MGBG plasma levels at a peak plasma concentration of at least 25% for at least 18 hours. In other embodiments, the peak plasma concentration is a therapeutically effective concentration. In other embodiments, the percentage of peak plasma concentration is therapeutically effective over a given period of time.

In certain embodiments, a pharmaceutical composition comprising MGBG has an oral bioavailability of at least 10, 20, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 45, 50, 55, or 60%. In other embodiments, the pharmaceutical composition has an oral bioavailability of at least 10%, 20%, 25%, 30%, 35%, 40%, or 45%. In other embodiments, the pharmaceutical composition has an oral bioavailability of at least 30%, at least 35%, at least 40%, or at least 45%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 20%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 30%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 35%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 40%. In certain embodiments, the pharmaceutical composition has an oral bioavailability of at least 45%. In certain embodiments, the pharmaceutical composition has an oral bioavailability that results in therapeutically effective plasma MGBG levels in a subject administered once daily for a period of at least 24 hours. In certain embodiments, the pharmaceutical composition has an oral bioavailability that results in therapeutically effective plasma MGBG levels in a subject administered twice daily for a period of at least 24 hours. In certain embodiments, the pharmaceutical composition has an oral bioavailability that results in therapeutically effective plasma MGBG levels in a subject administered three times daily for a period of at least 24 hours.

In certain embodiments, the pharmaceutical composition comprising MGBG has a half-life of at least 4, 6, 8, 10, 12, 14, 16, 18, 19, 20, 21, 22, 23, 24, 26, 28, 30, or 36 hours. In certain embodiments, the pharmaceutical composition has a half-life of at least 12 hours. In other embodiments, the pharmaceutical composition has a half-life of at least 18 hours. In other embodiments, the pharmaceutical composition has a half-life of at least 20 hours. In other embodiments, the pharmaceutical composition has a half-life of at least 24 hours. In certain embodiments, the pharmaceutical composition has a half-life of at least 48, 72, 96, or 120 hours.

Also provided herein is a pharmaceutical composition comprising MGBG and at least one oral, pharmaceutically acceptable excipient that, when orally administered to a subject, produces therapeutically effective systemic plasma MGBG levels that do not have significant dose limiting side effects. In certain embodiments, the side effect is of the gastrointestinal tract. In other embodiments, the gastrointestinal side effect is selected from nausea, vomiting, diarrhea, abdominal pain, oral mucositis, oral ulceration, pharyngitis, stomatitis, perforation of the gastrointestinal tract, ulceration of the gastrointestinal tract, obstruction of the gastrointestinal tract, and bleeding of the gastrointestinal tract. In other embodiments, the gastrointestinal side effect is selected from the group consisting of inhibition of gastrointestinal mucosal hyperplasia, inhibition of migration of developing epithelial luminal cells, and inhibition of differentiation of stem or progenitor cells into epithelial luminal cells. In certain embodiments, the side effect is selected from thrombocytopenia, leukopenia, phlebitis, laryngitis, cellulitis, dermatitis, and hypoglycemia.

Also provided herein is a low dose oral pharmaceutical composition for long term delivery comprising a therapeutically effective amount of MGBG and at least one pharmaceutically acceptable excipient that does not have significant gastrointestinal side effects. In certain embodiments, a low dose oral pharmaceutical composition for long term delivery (which does not have significant gastrointestinal side effects) comprising a therapeutically effective amount of MGBG and at least one pharmaceutically acceptable excipient produces therapeutically effective plasma levels of MGBG in a subject administered once daily for a period of at least 24 hours.

In certain embodiments, the pharmaceutical composition is formulated as a tablet or capsule. For example, in certain embodiments, the pharmaceutical composition comprises:

0.1-50% of a polyamine analog or a polyamine biosynthesis inhibitor;

0.1-99.9% of a filler;

0-10% of a disintegrant;

0-5% of a lubricant; and

0-5% glidant.

In certain embodiments, the pharmaceutical composition comprises:

0.1-50％MGBG；

0.1-99.9% of a filler;

0-10% of a disintegrant;

0-5% of a lubricant; and

0-5% glidant.

In other embodiments of the present invention, the substrate may be,

the filler is selected from sugar, starch, cellulose and poloxamer;

the disintegrant is selected from povidone and crospovidone;

the lubricant is magnesium stearate; and is

The glidant is silicon dioxide.

In other embodiments of the present invention, the substrate may be,

the filler is selected from lactose and microcrystalline cellulose;

the disintegrant is selected from povidone and crospovidone;

the lubricant is magnesium stearate; and is

The glidant is silicon dioxide.

In certain embodiments, the pharmaceutical composition comprises:

10-300mg of a polyamine analogue or polyamine biosynthesis inhibitor constituting 2-50% of the tablet content or capsule filler content;

0-10% of a disintegrant;

0-5% of a lubricant;

0-5% glidant; and

30-98% of filler.

In certain embodiments, the pharmaceutical composition comprises:

10-300mg MGBG, constituting 2-50% of the tablet content or capsule filler content;

0-10% of a disintegrant;

0-5% of a lubricant;

0-5% glidant; and

30-98% of filler.

In other embodiments, the pharmaceutical composition comprises

0.1-10% of a binder;

0-5% of a surfactant;

0-10% of an intergranular disintegrant; and

0-10% of an extra-granular disintegrant.

In other embodiments, the pharmaceutical composition may additionally comprise

0-10% of a binder;

0-5% of a surfactant;

0-10% of an intergranular disintegrant; and

0-10% of an extra-granular disintegrant.

In other embodiments of the present invention, the substrate may be,

the binder is selected from copovidone, hydroxypropyl cellulose, hydroxypropyl methyl cellulose and povidone;

said surfactant is selected from polyoxyethylene (20) sorbitan monooleate, poloxamer and sodium lauryl sulfate;

the intergranular disintegrant is selected from croscarmellose sodium, sodium starch gluconate, and crospovidone; and is

The extragranular disintegrant is selected from the group consisting of croscarmellose sodium, sodium starch gluconate, and crospovidone.

Also provided herein is a method of treating or delaying the onset or development of a condition in a subject in need thereof comprising administering an oral pharmaceutical composition comprising MGBG and at least one pharmaceutically acceptable excipient. In certain embodiments, the oral pharmaceutical composition is delivered in a therapeutically effective amount. In certain embodiments, the oral pharmaceutical composition has an oral bioavailability of at least 30%. In certain embodiments, the oral pharmaceutical composition does not have significant dose limiting side effects. In certain embodiments, the plasma level of MGBG is at least 75% of the peak plasma concentration for 4 or more hours. In other embodiments, the oral pharmaceutical composition, when orally administered to a subject, produces therapeutically effective systemic plasma MGBG levels for a period of at least 12 hours.

In certain embodiments, the condition is selected from proliferative disorders, inflammatory diseases, and autoimmune diseases, neurological diseases, and neurodegenerative diseases. In certain embodiments, the condition is selected from rheumatoid arthritis, osteoarthritis, multiple sclerosis, amyotrophic lateral sclerosis, HIV neuropathy, and HIV-associated dementia.

In certain embodiments, the proliferative disorder is selected from cancer, psoriasis, psoriatic arthritis, and atopic dermatitis. In certain embodiments, the neuropathy is selected from peripheral neuropathy, diabetic neuropathy, entrapment neuropathy (carpal tunnel syndrome), postherpetic neuralgia (PHN), chemotherapy-induced neuropathy, and HIV neuropathy.

In certain embodiments, the condition is selected from the group consisting of a proliferative disorder, rheumatoid arthritis, osteoarthritis, multiple sclerosis, and amyotrophic lateral sclerosis. In certain embodiments, the proliferative disorder may be selected from, for example, cancer, psoriasis, psoriatic arthritis, and atopic dermatitis.

Also provided is an oral pharmaceutical composition comprising a polyamine analog or polyamine biosynthesis inhibitor and at least one oral pharmaceutically acceptable excipient which when orally administered to a subject results in a therapeutically effective systemic plasma polyamine analog or polyamine biosynthesis inhibitor level for treating pain. Also provided is an oral pharmaceutical composition comprising MGBG and at least one oral pharmaceutically acceptable excipient, which when orally administered to a subject results in a therapeutically effective systemic plasma MGBG level for treating pain.

Also provided herein is a method of treating pain in a subject in need thereof comprising administering a polyamine analog or a polyamine biosynthesis inhibitor or a salt or protected derivative thereof. Also provided herein are methods of treating pain in a subject in need thereof comprising administering MGBG. In certain embodiments, MGBG is administered in a therapeutically effective amount. Further provided is a method of treating pain in a subject in need thereof comprising administering a therapeutically effective amount of a pharmaceutical composition comprising MGBG and at least one pharmaceutically acceptable excipient.

In certain embodiments, the pain is selected from inflammatory pain, pain due to nerve injury, chronic pain, refractory cancer pain, complex regional pain syndrome, neuropathic pain, surgical or post-surgical pain, dental pain, pain resulting from skin injury, low back pain, headache, migraine, tactile allodynia, and hyperalgesia. In certain embodiments, the pain is chronic. In other embodiments, the pain is acute. In certain embodiments, the pain is inflammatory pain.

In certain embodiments, the administration of MGBG or a pharmaceutical composition thereof is oral. In other embodiments, the administration is intravenous.

In certain embodiments, the administration is a combination of oral and intravenous. In certain embodiments, the first administration is oral and the second is IV; in other embodiments, the first is IV and the second is oral; in either case, additional oral or IV administrations may follow. In certain embodiments, the pain is surgical or post-surgical pain. For example, in certain embodiments, the preoperative administration is oral and the perioperative administration is IV; in other embodiments, the preoperative administration is IV, preoperative or IV and the postoperative administration is oral. In either case, additional oral or IV administrations may follow. In certain embodiments, the pre-operative, perioperative, and post-operative administration is IV.

Also provided herein is a method of treating a condition in a subject in need thereof comprising administering

An oral pharmaceutical composition comprising MGBG and at least one pharmaceutically acceptable excipient; and

another therapeutic agent.

In certain embodiments, MGBG is delivered in a therapeutically effective amount. In other embodiments, MGBG is delivered in a subtherapeutic amount. In certain embodiments, the additional therapeutic agent is delivered in a therapeutically effective amount. In other embodiments, the additional therapeutic agent is delivered in a subtherapeutic amount. In certain embodiments, MGBG and the other therapeutic agent are co-delivered in amounts that are individually sub-therapeutic but together therapeutically effective. In other embodiments, MGBG and the other therapeutic agent are co-delivered in amounts that are individually therapeutically effective.

Additionally, provided herein is a method of treating a condition. The method comprises administering to a subject in need of such treatment an effective amount of MGBG, a salt of MGBG, a protected derivative of MGBG, or a polyamine analog or a polyamine biosynthesis inhibitor or a salt, a protected derivative, or a stereoisomer thereof, wherein the condition is selected from the group consisting of crohn's disease, parkinson's disease, inflammatory bowel disease, Multiple Sclerosis (MS), Amyotrophic Lateral Sclerosis (ALS), hepatitis, HBV, HCV, nephritis, encephalitis, glomerulonephritis, rheumatoid arthritis, type 2 diabetes, cardiac fibrosis and angiotensin II-related hypertension, osteoporosis, IgE-producing mast cell-mediated hypersensitivity responses, peripheral sensory neuropathy associated with HIV infection or diabetes, asthma, autism, dermatomyositis, frailty, obesity, primary biliary cirrhosis, primary sclerosing cholangitis, post-radiation complications, inflammatory bowel disease, or a salt, a protected derivative, or a stereoisomer thereof, Psoriatic arthritis, sarcoidosis, scleroderma with or without pulmonary fibrosis, a kidney-related autoimmune condition, diabetic nephropathy, diabetic vascular complications, and an autoimmune condition associated with lymphatic proliferation.

Further provided herein is a method of reducing differentiation of macrophages from monocytes comprising contacting the monocytes with an effective amount of an agent that inhibits S-adenosylmethionine decarboxylase in the monocytes or inhibits polyamine biosynthesis. In certain embodiments, the agent is MGBG, or a salt or protected derivative thereof.

In one embodiment, the agent is capable of inhibiting SAMDC or any pathway containing AMD I, e.g. any entity upstream or downstream of a SAMDC-containing pathway, in particular any pathway containing SAMDC and associated with adenosine production. In another embodiment, the agent is capable of inhibiting polyamine biosynthesis or any pathway involved in polyamine biosynthesis. In general, a pathway containing SAMDC or adenosine is understood to refer to a pathway in which SAMDC or adenosine is involved (including, for example, as a substrate, catalyst, product, or byproduct).

The agent may be any of a variety of known or later-discovered agents that inhibit the activity of the enzyme S-adenosylmethionine decarboxylase, which inhibits polyamine biosynthesis, for example, in a cell. In one embodiment, the agent is a chemical agent, including but not limited to organic molecules and salts thereof, protected derivatives and stereoisomers, inorganic molecules, or various ionic or elemental entities.

Compounds for use in the methods and compositions disclosed herein include polyamine analogs and polyamine biosynthesis inhibitors, as well as salts, prodrugs, solvates, anhydrous forms, protected derivatives, structural isomers, stereoisomers, amino acid conjugates, and porphyrin conjugates thereof. Any polyamine analog is suitable for use in the method of the invention.

Exemplary polyamine analogs for use in the methods of the present invention include compounds of structural formulae 1, 2, 3, 4, 5, 6, and 7 and their corresponding stereoisomers, salts, and protected derivatives.

Formula 1 has the structure

Wherein

R₁、R₂、R₄、R₆And R₇Independently selected from hydrogen, alkyl and aryl; and is

R₃And R₅Is an alkyl group.

Formula 2 has the structure

Wherein

R₁、R₂、R₄、R₆、R₈And R₉Independently selected from hydrogen, alkyl and aryl; and is

R₃、R₅And R₇Is an alkyl group.

Formula 3 has the structure

Wherein

R₁、R₂、R₄、R₆、R₁₀And R₁₁Independently selected from hydrogen, alkyl and aryl; and is

R₃、R₅、R₇And R₉Is an alkyl group.

Formula 4 has the structure

Wherein

R₁And R₅Independently selected from methyl, ethyl, n-propyl and isopropyl;

R₂、R₃and R₄Independently selected from C₁-C₆Alkyl radical, C₂-C₆Alkenyl radical, C₃-C₆Cycloalkyl radical, C₁-C₆alkyl-C₃-C₆cycloalkyl-C₁-C₆Alkyl radical, C₃-C₁₀Aryl and C₁-C₆alkyl-C₃-C₁₀aryl-C₁-C₆An alkyl group; and is

R₆、R₇、R₈And R₉Independently selected from hydrogen, methyl and ethyl;

formula 5 has the structure

Wherein

R₁And R₆Independently selected from methyl, ethyl, n-propyl and isopropyl;

R₂、R₃、R₄and R₅Independently selected from C₁-C₆Alkyl radical, C₂-C₆Alkenyl radical, C₃-C₆Cycloalkyl radical, C₁-C₆alkyl-C₃-C₆cycloalkyl-C₁-C₆Alkyl radical, C₃-C₁₀Aryl and C₃-C₁₀aryl-C₁-C₆An alkyl group; and is

R₇、R₈、R₉、R₁₀And R₁₁Independently selected from hydrogen, methyl and ethyl.

In another embodiment, the polyamine analog is a compound of structures 2 and 3, wherein

R₃、R₅、R₇And R₉Independently is (CH)₂)_xA group;

x is an integer from 2 to 6; and is

R₄、R₆And R₈Is a hydrogen atom.

In yet another embodiment, the polyamine analog is a compound of structures 2 and 3, wherein

R₃、R₅、R₇And R₉Independently is (CH)₂)_xA group;

x is an integer from 2 to 6;

R₄、R₆and R₈Is a hydrogen atom;

R₁and R₁₀Is an alkyl group; and is

R₂And R₁₁Is a hydrogen atom.

R₃、R₅、R₇And R₉Independently is (CH)₂)_xA group;

x is an integer from 2 to 6;

R₄、R₆and R₈Is a hydrogen atom;

R₁and R₁₀Is an alkyl group;

R₂and R₁₁Is a hydrogen atom; and is

The polyamine analogs have a molecular weight of less than 500.

Other embodiments of the compounds of structure 4 include those wherein R is₆、R₇、R₈And R₉Are those of hydrogen.

In other embodiments, R₁And R₅Is ethyl.

In still other embodiments, the first and second electrodes are,

R₆、R₇、R₈and R₉Is hydrogen; and is

R₁And R₅Is ethyl.

In still other embodiments, the first and second electrodes are,

R₂and R₄Independently selected from C₁-C₆An alkyl group; and is

R₃Is selected from C₁-C₆Alkyl radical, C₂-C₆Alkenyl radical, C₃-C₆Cycloalkyl radical, C₁-C₆alkyl-C₃-C₆cycloalkyl-C₁-C₆Alkyl radical, C₃-C₁₀Aryl and C₁-C₆alkyl-C₃-C₁₀aryl-C₁-C₆An alkyl group.

Additional polyamine analogs suitable for use in the present invention include compounds of formula 6, and the corresponding stereoisomers, salts, and protected derivatives thereof:

wherein

R₄Is selected from C₂-C₆N-alkenyl radical, C₃-C₆Cycloalkyl radical, C₃-C₆Cycloalkenyl radical and C₃-C₆An aryl group;

R₃and R₅Independently selected from single bond, C₁-C₆Alkyl and C₁-C₆An alkenyl group;

R₂and R₆Independently selected from C₁-C₆Alkyl radical, C₁-C₆Alkenyl radical, C₃-C₆Cycloalkyl radical, C₃-C₆Cycloalkenyl radical and C₃-C₆An aryl group;

R₁and R₇Independently selected from hydrogen, C₁-C₆Alkyl and C₂-C₆An alkenyl group; and is

R₈、R₉、R₁₀And R₁₁Is hydrogen.

In certain embodiments of the compounds of formula 6, R₁And R₇Independently selected from C₁-C₆Alkyl and C₂-C₆An alkenyl group.

Additional polyamine analogs suitable for use in the present invention include compounds of formula 7, and the corresponding stereoisomers, salts, and protected derivatives thereof:

wherein

R₄Is selected from C₁-C₆N-alkyl and C₁-C₆A branched alkyl group;

R₃and R₅Independently selected from single bond or C₁-C₆An alkyl group;

R₂and R₆Independently selected from C₁-C₆Alkyl radical, C₁-C₆Alkenyl radical, C₃-C₆Cycloalkyl radical, C₃-C₆Cycloalkenyl or C₃-C₆An aryl group;

R₁and R₇Independently selected from H, C₁-C₆Alkyl or C₂-C₆An alkenyl group; and is

R₈、R₉、R₁₀And R₁₁Is hydrogen.

In certain embodiments of the compounds of formula 7

R₂And R₇Independently selected from C₁-C₆Alkyl or C₂-C₆An alkenyl group;

R₄is selected from C₁-C₆Saturated n-alkyl and C₁-C₆A saturated branched alkyl group; and is

R₃And R₅Independently selected from the group consisting of single bond and C₁-C₆Saturated n-alkyl.

According to another embodiment of the invention, the agent is a chemical moiety that inhibits S-adenosylmethionine decarboxylase activity, inhibits polyamine biosynthesis, and/or increases adenosine activity.

Examples of such moieties include, but are not limited to, those listed in table 1. Regardless of the form of the moieties listed in table 1, it is understood that the moieties include salts, protected derivatives, and stereoisomers thereof, if applicable.

Table 1.

In yet another embodiment, the agent is a compound selected from the group consisting of: MGBG, MDL73811, CGP48664, Berenil, pentamidine, SL47, and SL93, or a combination of two or more thereof. In yet another embodiment, the agent MGBG, SL47 or SL 93. In yet another embodiment, two or more reagents are used. The two or more agents may be used sequentially or simultaneously.

MGBG is 1, 1' [ methyldimethylenebiguanide ] dinitrilo-diguanide and is also known as methylglyoxal bis (guanylhydrazone), methyl-GAG, Me-G and mitoguazone hydrazone. As used herein, MGBG includes the free base and salts thereof. It is usually, but not necessarily, used as the dihydrochloride. MGBG may exist as any one of the following isomers, or a tautomer and/or cis/trans isomer, or a mixture of one or more thereof:

and

in certain embodiments, MGBG may exist as one of the following isomers, or a tautomer and/or cis/trans isomer thereof, or a mixture of one or more thereof:

and

in certain embodiments, the compounds have a structure selected from formulas 8a-8 c:

R₁-R₆selected from the group consisting of hydrogen, alkyl groups having 1 to 12 carbon atoms and aralkyl groups, provided that in formula (8a), R is₁And R₆Is not hydrogen;

R₇selected from the group consisting of hydrogen, alkyl groups having 1 to 12 carbon atoms, aryl groups and aralkyl groups;

m, n are each independently an integer of 3 to 6, 3 and 6 being included; and is

v, w, x, y and z are each independently an integer of 3 to 10, 3 and 10 being included.

Further disclosure can be found in WO98/10766, the disclosure of which is incorporated by reference in its entirety as if written herein, for example at pages 3-4.

In certain embodiments, the compound has the structure of formula 9 a:

E-NH-B-A-B-NH-B-A-B-NH-B-A-B-NH-B-A-B-NH-E

wherein

A is independently selected from the group consisting of: c₁-C₆Alkyl radical, C₂-C₆Alkenyl radical, C₂-C₆Alkynyl, C₃-C₆Cycloalkyl radical, C₃-C₆Aryl and C₃-C₆A cycloalkenyl group;

b is independently selected from the group consisting of: single bond, C₁-C₆Alkyl and C₂-C₆An alkenyl group; and is

E is independently selected from the group consisting of: hydrogen, C₁-C₆Alkyl radical, C₂-C₆Alkenyl radical, C₂-C₆Alkynyl, C₃-C₆Cycloalkyl radical, C₃-C₆Aryl and C₃-C₆A cycloalkenyl group;

with the proviso that at least one moiety a is selected from the group consisting of: c₂-C₆Alkenyl radical, C₂-C₆Alkynyl, C₃-C₆Cycloalkyl radical, C₃-C₆Aryl and C₃-C₆Cycloalkenyl, or at least one B moiety is selected from the group consisting of: c₂-C₆An alkenyl group; and all salts, hydrates, solvates and stereoisomers thereof.

In another embodiment, the conformationally constrained polyamine analog is selected from the group of compounds of formula 9 b:

E-NH-B-A-B-NH-B-A-B-NH-B-A-B-NH(-B-A-B-NH)x-E

wherein:

x is an integer from 2 to 16;

provided that at leastOne moiety a is selected from the group consisting of: c₂-C₆Alkenyl radical, C₂-C₆Alkynyl, C₃-C₆Cycloalkyl radical, C₃-C₆Aryl and C₃-C₆Cycloalkenyl, or at least one B moiety is selected from C₂-C₆Alkenyl groups;

and all salts, hydrates, solvates and stereoisomers thereof.

In another embodiment, x is 4, 6, 8 or 10.

In another embodiment, x is 4. In another embodiment, x is 6.

In another embodiment, x is 8.

In another embodiment, x is 10.

In another embodiment, the conformationally constrained polyamine analog is selected from the group of compounds of formula 9 c:

E-NH-B-A-B-NH-B-A-B-NH-B-A-B-NH(-B-A-B-NH)x-E

wherein:

E is independently selected from the group consisting of: c₁-C₆Alkyl radical, C₁-C₆Alkanol, C₃-C₆Cycloalkanol and C₃-C₆Hydroxyaryl, with the proviso that at least one E moiety is selected from the group consisting of: c₁-C₆Alkanol, C₃-C₆Cycloalkanol and C₃-C₆A hydroxyaryl group; and is

x is an integer from 0 to 16;

and all salts, hydrates, solvates and stereoisomers thereof.

In another embodiment, the conformationally constrained polyamine analog is selected from the group of compounds of formula 9 d:

E-NH-D-NH-B-A-B-NH-D-NH-E

wherein A is selected from the group consisting of: c₂-C₆Olefins and C₃-C₆Cycloalkyl, cycloalkenyl, and aryl;

b is independently selected from the group consisting of a single bond and C₁-C₆Alkyl and alkenyl;

d is independently selected from the group consisting of: c₁-C₆Alkyl and alkenyl, and C₃-C₆Cycloalkyl, cycloalkenyl, and aryl;

e is independently selected from hydrogen, C₁-C₆Alkyl and alkenyl; and all salts, hydrates, solvates and stereoisomers thereof.

In another embodiment, the conformationally constrained polyamine analog is a macrocyclic polyamine selected from the group consisting of formula 9 e:

wherein

A₁Each A₂(if present) and A₃Independently selected from C₁-C₈An alkyl group; each Y is independently selected from hydrogen or C₁-C₄An alkyl group;

m is selected from C₁-C₄An alkyl group;

k is 0, 1, 2 or 3; and is

R is selected from C₁-C₃₂An alkyl group;

and all salts, hydrates, solvates and stereoisomers thereof.

In another embodiment, the Y group is hydrogen or-CH₃。

In another embodiment, A₁Each A₂(if present) and A₃Independently selected from C₂-C₄An alkyl group.

In yet another embodiment, M is-CH 2-.

In another embodiment, the conformationally constrained polyamine analog is selected from the group consisting of macrocyclic polyamine analogs of formula 9 f:

wherein

A₁Each A₂(if present) and A₃Independently selected from C₁-C₈An alkyl group;

A₄is selected from C₁-C₈Alkyl or null;

x is selected from-hydrogen, -Z, -CN, -NH₂、-C(＝O)-C₁-C₈Alkyl or-NHZ, with the proviso that when A₄When not effective, X is hydrogen, -C (═ O) -C₁-C₈Alkyl or-Z;

z is selected from the group consisting of: amino protecting groups, amino end capping groups, amino acids and peptides:

each Y is independently selected from hydrogen or C₁-C₄An alkyl group;

m is selected from C₁-C₄An alkyl group;

k is 0, 1, 2 or 3; and is

R is selected from C₁-C₃₂An alkyl group;

and all salts, hydrates, solvates and stereoisomers thereof.

In certain embodiments, a₄Are not effective.

In other embodiments, X is-Z and-Z is hydrogen.

In other embodiments, X is-Z and-Z is 4-morpholinocarbonyl.

In other embodiments, X is-Z and-Z is acetyl.

In other embodiments, X is-Z and-Z is t-Boc or Fmoc.

In other embodiments, Y is-CH 3.

In other embodiments, M is-CH₂-。

In other embodiments, k is 1.

In other embodiments, A and A₃is-CH₂CH₂CH₂-。

In other embodiments, -CH₂CH₂CH₂CH₂-。

In other embodiments, R is C₁₃H₂₇。

In still other embodiments, a will be paired₄、X、Z、Y、M、k、A₁、A₃And one or more specific limitations of R are combined together.

In other embodiments of the macrocyclic polyamine analog compound,

A₄is C₁-C₈An alkyl group;

x is-NHZ; and is

Z is selected from one of the 20 genetically encoded amino acids (alanine, cysteine, aspartic acid, glutamic acid, phenylalanine, glycine, histidine, isoleucine, lysine, methionine, asparagine, proline, glutamine, arginine, serine, threonine, valine, tryptophan, tyrosine), a peptide of formula acetyl-SKLQL-, a peptide of formula acetyl-SKLQ-I3-alanine-, or a peptide of formula acetyl-SKLQ-.

In these cases, when Z is an amino acid or a peptide, the therapeutic agent to be used is a polyamine-amino acid conjugate or a polyamine-peptide conjugate.

In one embodiment, the only conformational restriction of the polyamine analog is due to a carbon-carbon double bond (vinyl, C ═ C) in the molecule.

In another embodiment, the only conformational constraint of the polyamine analog is due to a cycloalkyl group, such as cyclopropyl, in the molecule.

Compounds include, but are not limited to:

and

further disclosure can be found in WO2007/040535, the disclosure of which is incorporated by reference in its entirety as if written herein.

Additional analogs and derivatives include those encompassed by the following formula 10 a:

R-X-polyamines

Wherein

R is selected from H or from the following groups: a linear or branched C1-50 saturated or unsaturated aliphatic group, carboxyalkyl, alkoxycarbonylalkyl or alkoxy; a C1-8 cycloaliphatic radical; a monocyclic or polycyclic aryl-substituted aliphatic group; an aliphatic-substituted monocyclic or polycyclic aromatic group; a monocyclic or polycyclic heterocyclic group; a monocyclic or polycyclic heterocyclic 25 aliphatic group; c1-10 alkyl; an arylsulfonyl group; or cyano;

x may be-CO-, -SO₂or-CH₂-, and

the "polyamine" may be any naturally occurring, such as putrescine, spermine or spermidine, or synthetically produced polyamine.

Preferably, R is at least about C5, at least about C10, at least about C11, at least about C12, at least about C13, at least about C14, at least about C15, at least about C16, at least about C17, at least about C18, at least about C19, at least about C20, or at least about C22.

The linkage between X and the polyamine may be direct, wherein there is no atom between X and the nitrogen of the amine group of the polyamine; or indirectly, where there may be one or more atoms between X and the nitrogen of the amine groups of the polyamine. The linkage between X and the polyamine can be via any amino group within the polyamine, although primary amino groups are used in preferred embodiments of the invention.

In a preferred embodiment of the invention, wherein the linkage between X and the polyamine is indirect, the incorporated atom or atoms are preferably atoms of an amino acid or a derivative thereof. In a particularly preferred embodiment of this type, the atom or atoms incorporated are the atoms of lysine, aspartic acid, glutamic acid, ornithine or 2, 4-diaminobutyric acid. Preferred compounds of this type may be represented by formula 10 b:

R-X-L-polyamines

Wherein

R is a linear or branched C10-50 saturated or unsaturated aliphatic group, carboxyalkyl, alkoxycarbonylalkyl or alkoxy; a C1-8 cycloaliphatic radical; a monocyclic or polycyclic aryl substituted or unsubstituted aliphatic group; aliphatic substituted or unsubstituted monocyclic or polycyclic aromatic groups; a monocyclic or polycyclic heterocyclic group; a monocyclic or polycyclic heterocycloaliphatic group; an arylsulfonyl group;

x is-CO-, -SO₂-or-CH₂-; and is

L is a covalent bond or a naturally occurring amino acid, ornithine, 2, 4-diaminobutyric acid, or a derivative thereof.

The analogs and derivatives of the present invention may optionally be further substituted at one or more other positions on the polyamine. They include, but are not limited to, internal nitrogen and/or internal carbon atoms. In one aspect of the invention, preferred substituents are structures that enhance the irreversible ability of a compound to bind to a polyamine-binding molecule, such as a polyamine transporter, an enzyme, or DNA, by enhancing polyamine transport inhibition, binding affinity, or otherwise. The additional substituents include aziridine groups and various other aliphatic, aromatic, mixed aliphatic-aromatic or heterocyclic polycyclic structures. Reactive moieties like aziridine that are covalently bound to a polyamine transporter or another polyamine binding molecule are also within the scope of the invention. Examples of reactive groups that react with nucleophiles to form covalent bonds include chloro-, bromo-, and iodoacetamides, sulfonyl fluorides, esters, nitrogen mustards, and the like. Such reactive moieties are useful as affinity labels in diagnostic or research work and may provide pharmacological activity in inhibiting polyamine transport or polyamine synthesis. The reactive group may be a reactive photoaffinity group, such as an azide or benzophenone group. Chemical reagents for photoaffinity labeling are well known in the art (fleming, s.a., Tetrahedron 1995, 51, 12479-.

A preferred aspect of the present invention relates to polyamine analogs or derivatives which are highly specific polyamine transport inhibitors having pharmaceutical utility as anticancer chemotherapeutic agents. One class of polyamine analogs or derivatives of the present invention that bind to the polyamine binding site of the molecule and/or inhibit polyamine transport is depicted by the following formula 10 c:

wherein

a. b and c are independently in the range of 1 to 10;

d and e are independently in the range of 0 to 30;

each X is independently a carbon (C) or sulfur (S) atom, and

R₁and R₂As described below, or R₁X (O) n-and R₂X (O) n-is each independently replaced by H; and is

Denotes a chiral carbon position;

and with the proviso that

If X is C, then n is 1;

if X is S, then n is 2; and is

If X is C, then the X (O) group may be CH₂Such that n is o.

In the above formula, R₁And R₂Independently selected from H or from the following groups: a linear or branched C1-50 saturated or unsaturated aliphatic group, carboxyalkyl, alkoxycarbonylalkyl or alkoxy; a C1-8 cycloaliphatic radical; a monocyclic or polycyclic aryl-substituted aliphatic group; an aliphatic-substituted monocyclic or polycyclic aromatic group; a monocyclic or polycyclic aromatic or saturated heterocyclic group; a monocyclic or polycyclic heterocycloaliphatic group; c1-10 alkyl; an arylsulfonyl group; or a cyano group.

Examples of heterocycles as used herein include, but are not limited to, pyrrole, furan, thiophene, imidazole, oxazole, thiazole, pyrazole, 3-pyrroline, pyrrolidine, pyridine, pyrimidine, purine, quinoline, isoquinoline, and carbazole.

All of the above depicted aliphatic, carboxyalkyl, alkoxycarbonylalkyl, alkoxy, 30 cycloaliphatic, aryl, aromatic and heterocyclic moieties can of course also be optionally substituted with 1 to 3 substituents independently selected from: halo (fluoro, chloro, bromo or iodo), lower alkyl (1-6C) and lower alkoxy (1-6C).

As used herein, carboxyalkyl refers to the substituent-R '-COOH, wherein R' is alkylene; and alkoxycarbonylalkyl means-R '-COOR where R' and R are alkylene and alkyl, respectively. In a preferred embodiment, alkyl means a saturated straight or branched chain hydrocarbon group of 1 to 6 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, n-pentyl, 2-methylpentyl, n-hexyl and the like. Alkylene is the same as alkyl except that the group is divalent. The aryl or alkylsulfonyl moiety has the formula SO₂R and the alkoxy moiety has the formula-O-R, whereinR is alkyl as defined above, or is aryl, wherein aryl is phenyl, optionally substituted with 1-3 substituents independently selected from: halo (fluoro, chloro, bromo or iodo), lower alkyl (1-6C) and lower alkoxy (1-6C).

A preferred group of compounds encompassed by the above are those wherein d is 4 and e is 0.

Another class of polyamine analogs or derivatives of the present invention that bind to the polyamine binding site of the molecule and/or inhibit polyamine transport are depicted by the following formula 10 d:

wherein

a. b and c are independently in the range of 1 to 10;

d and e are independently in the range of 0 to 30;

R₁and R₂As defined above for formula 8c, and

R₃and R₄Independently selected from organic substituents including-CH₃And as above in formula 8c for R₁And R₂As defined. This group of analogs results from the reductive amination of a free amino precursor with a ketone.

In a preferred embodiment of the invention, R₁And R₂Same and as described for formula 8 c. Position R₃And R₄Or may be the same, and all R' s₁To R₄The same may be true. In addition, the position R in formula 8d₁、R₂、R₃And R₄Each may also independently be H.

In another aspect of the invention, the proximal and/or distal amino groups relative to a polyamine (e.g., spermine) can be dialkylated to form tertiary amines.

R₁、R₂、R₃And R₄Each can be independently varied and is as defined above for formula III. R₁、R₂、R₃And R₄Each may also independently be H. a. The values of b, c, d, and e are as described above for equation 8 d. This aspect of the invention is depicted by the following formula 10 e:

in another aspect of the invention, compounds are also provided that lack a proximal or distal amino group on the acyl portion of the molecule. They are represented by formula 10 f:

wherein

Z₁Is NR₁R₃And Z is₂Is selected from-R₁、-CHR₁R₂or-CR₁R₂R₃(wherein R is₁、R₂And R₃As defined above for formula 8 c); or Z₂Is NR₂R₄And Z is₁Is selected from-R₁、-CHR₁R₂or-CR₁R₂R₃(wherein R is₁、R₂And R₃As defined above for formula 8 d). a. The values of b and c independently range from 1 to 10; d and e are independently in the range of 0 to 30. The compounds encompassed by formula V can be prepared by first coupling an amino acid derivative (modified to contain a non-amine having a Z group) to a polyamine, followed by appropriate derivatization of the amine containing the Z group. The chemistry of the reaction is known in the art and disclosed herein.

In a preferred embodiment of the inventionPosition R of all formulae listed above₁、R₂、R₃And R₄Independently selected from the following, wherein g, h, i, j and k are each independently selected from 0 to 15:

wherein E refers to the "contralateral (entgegen)" and Z refers to the "ipsilateral (zusammen)".

Compounds include, but are not limited to:

further disclosure can be found in WO2002/053519, the disclosure of which is incorporated by reference in its entirety as if written herein.

Additional analogs and derivatives include synthetic derivatives of the original polyamines wherein the carbon atoms of the original polyamine comprise the amide group, which synthetic derivatives inhibit cellular uptake of the natural polyamines by specifically binding to the cellular transporters of the natural polyamines.

In certain embodiments, the carbon at which the amide group is located is between the two internal nitrogen atoms of the original polyamine.

In certain embodiments, the synthetic derivatives comprise dimers of the original polyamine, the monomers of the dimers being linked together by a spacer side chain anchored to the amide group of each monomer.

In certain embodiments, the primary polyamine is selected from the group consisting of: putrescine, spermidine and spermine.

In certain embodiments, the primary polyamine is spermine.

In certain embodiments, the synthetic derivatives have the following general formula 11 a:

wherein R is₁And R¹ ₁Independently represents a hydrogen atom or an alkyl group having 1 to 2 carbon atoms, R₂、R¹ ₂Or R₃And R¹ ₃Independently represents a hydrogen atom or a methyl group, w and z independently represent an integer of 2 or 3, x represents an integer of 0 to n, n represents an integer of 3 to 6, the sum of x and y is equal to n, and S represents a hydrogen atom or a molecule that cannot be captured by the natural polyamine transporter.

In certain embodiments, the monomer has the following general formula 11 b:

wherein R is₁And R¹ ₁Independently represents a hydrogen atom or an alkyl group having 1 to 2 carbon atoms, R₂、R¹ ₂Or R₃And R¹ ₃Independently represents a hydrogen atom or a methyl group, w and z independently represent an integer of 2 or 3, x represents an integer of 0 to n, n represents an integer of 3 to 6, the sum of x and y is equal to n, and wherein the spacer side chain comprises a main chain containing a linear hydrocarbon of 3 to 8 atoms.

In certain embodiments, the backbone comprises sulfur, oxygen, or nitrogen.

In certain embodiments, w-2, z-2 x-o and y-3.

In certain embodiments, w-2, z-2, x-o and y-3.

In certain embodiments, w-2, z-2, x-o and y-4.

Compounds include, but are not limited to:

further disclosure can be found in WO98/17632, the disclosure of which is incorporated by reference in its entirety as if written herein.

Additional analogs and derivatives include those encompassed by the following formula 12 a:

wherein n may be 0 to 8 and the aminomethyl functionality may be ortho-, meta-or para-substituted, R is hydrogen, -CH₃、-CH₂CH₃2-aminoethyl, 3-aminopropyl, 4-aminobutyl, 5-aminopentyl, 6-aminohexyl, N-acetylhexyl, N-,7-aminoheptyl, 8-aminooctyl, N-methyl-2-aminoethyl, N-methyl-3-aminopropyl, N-methyl-4-aminobutyl, N-methyl-5-aminopentanyl, N-methyl-6-aminohexyl, N-methyl-7-aminoheptyl, n-methyl-8-aminooctyl, N-ethyl-2-aminoethyl, N-ethyl-3-aminopropyl, N-ethyl-4-aminobutyl, N-ethyl-5-aminopentyl, N-ethyl-6-aminohexyl, N-ethyl-7-aminoheptyl or N-ethyl-8-aminooctyl and R is a moiety selected from the group consisting of: hydrogen or a linear or branched C1-20 saturated or unsaturated aliphatic group; aliphatic amines instead of propylamine (when R ═ H, n ═ 1 and the aminomethyl functionality is para-substituted); an alicyclic group; a monocyclic or polycyclic aromatic group; a monocyclic or polycyclic aryl-substituted aliphatic group; an aliphatic-substituted monocyclic or polycyclic aromatic group; a monocyclic or polycyclic heterocyclic group; a monocyclic or polycyclic heterocycle-substituted aliphatic group; an aliphatic-substituted aromatic group; and halogenated forms thereof.

In certain embodiments, analogs and derivatives that may be used in accordance with the present disclosure may be further modified as depicted in formula 12 b:

wherein n may be 0 to 8, R and R₁As mentioned above, R₂Can be independently selected from hydrogen, -CH₃、-CH₂CH₃And R is₃And R₄May be the same or different and are independently selected from hydrogen or fluorine.

In certain embodiments, compounds that may be used according to the present disclosure are depicted by formula 12 c:

wherein m and n can independently be 0 to 7, but m cannot equal n, when R₁Is equal to R₂And R is₃Is equal to R₄When o may be 2 to 4, R may independently beSelected from H, -CH₃、-CH₂CH₃，R₁And R₂Can be independently selected from hydrogen, -CH₃、-CH₂CH₃And R is₃And R₄May be the same or different and are independently selected from hydrogen or fluorine.

In certain embodiments, the compound has formula 12 d:

wherein R is hydrogen, -CH₃、-CH₂CH₃M and n may independently be 0 to 7 and o

May be 2 to 4, R₂Can be independently selected from hydrogen, -CH₃、-CH₂CH₃And R is₃And R₄May be the same or different and are independently selected from hydrogen or fluorine.

In certain embodiments, the compounds of the present invention are represented by formula 12 e:

wherein R is hydrogen, -CH₃、-CH₂CH₃M may be 0 to 7, n may be 0 to 8 and o may be 2 to 4, R₂Can be independently selected from hydrogen, -CH₃、-CH₂CH₃And R is₃And R₄May be the same or different and are independently selected from hydrogen or fluorine.

Compounds include, but are not limited to:

further disclosures can be found in WO05/105729, the disclosure of which is incorporated by reference in its entirety as if written herein.

Additional analogs and derivatives include compounds of formulas 13 a-d:

wherein R is₁And R₂Independently selected from the group consisting of: -C₁-C₁₀Alkyl, -C₃-C₁₀Cycloalkyl, -C₁-C₁₀Alkylene-cycloalkyl, -C₆-C₁₀Aryl and-C₁-C₁₀Alkylene-aryl, wherein when R is₁And R₂When both are alkyl, R₁And R₂At least one of which is-C₂-C₁₀Alkyl and wherein R₁And R₂Are neither tert-butyl; and all salts, hydrates, solvates and stereoisomers thereof; and all mixtures of stereoisomers thereof, including racemic mixtures. In one embodiment, the substituents on the cyclopropyl ring are trans to each other. In another embodiment, the substituents on the cyclopropyl ring are cis to each other.

In one embodiment, when R₁And R₂When both are alkyl, R₁And R₂Is a straight chain alkyl group. In another embodiment, when R₁And R₂When both are alkyl, R₁And R₂Are all straight chain alkyl. In one embodiment, R₁And R₂is-C₁-C₁₀Alkyl and the other is-C₂-C₁₀An alkyl group. In one embodiment, R₁And R₂is-C₁-C₁₀Alkyl and the other is-C₄-C₁₀An alkyl group. In one embodiment, R₁And R₂Are all-C₄-C₁₀An alkyl group. In one embodiment, R₁And R₂is-C₆-C₁₀An alkyl group. In one embodiment, R₁And R₂Are all-C₆-C₁₀An alkyl group. In one embodiment, R₁And R₂is-C₁-C₁₀Alkyl and the other being selected from the group consisting of-C₂-C₄Straight chain alkyl and-C₄-C₁₀Alkyl groups. In another embodiment, R₁And R₂Independently selected from the group consisting of: -CH₃、-(CH₂)₃CH₃And- (CH)₂)sCH₃With the proviso that R is₁And R₂Are neither-CH₃。

In one embodiment, R₁And R₂is-C₁-C₁₀Alkyl and the other is-C₃-C₁₀Cycloalkyl, -C₁-C₁₀Alkylene-cycloalkyl, -C₆-C₁₀Aryl or-C₁-C₁₀Alkylene-aryl groups. In one embodiment, R₁And R₂is-C₁-C₁₀Alkyl and the other is-C₃-C₁₀Cycloalkyl or-C₁-C₁₀Alkylene-cycloalkyl groups. In one embodiment, R₁And R₂is-C₁-C₁₀Alkyl and the other is-C₃-C₁₀A cycloalkyl group. In one embodiment, R₁And R₂is-C₁-C₁₀Alkyl and the other is-C₆-C₁₀Aryl or-C₁-C₁₀Alkylene-aryl groups. In one embodiment, R₁And R₂is-C₁-C₁₀Alkyl and the other is-C₆-C₁₀And (4) an aryl group. In another embodiment, R₁And R₂Are all-C₆-C₁₀And (4) an aryl group. In another embodiment, R₁And R₂Are all-C₃-C₁₀A cycloalkyl group. In one embodiment, the aryl group is benzene. In one embodiment, the cycloalkyl group is adamantyl. In one embodiment, the adamantyl group is a 1-adamantyl group. In another embodiment, the adamantyl groupIs 2-adamantyl. In another embodiment, R₁And R₂Independently selected from-CH₃Phenyl and adamantyl, with the proviso that R₁And R₂Are neither-CH₃。

Compounds include, but are not limited to:

further disclosure can be found in WO2008/112251, the disclosure of which is incorporated by reference in its entirety as if written herein.

Additional analogs and derivatives include compounds of formula 14 a:

R₁-X-R₂

wherein

R₁Is H or a head group selected from the group consisting of: straight or branched C_1-10Aliphatic, cycloaliphatic, mono-or polycyclic aromatic, mono-or polycyclic aryl-substituted aliphatic, aliphatic-substituted mono-or polycyclic aromatic, mono-or polycyclic heterocyclic-substituted aliphatic, and aliphatic-substituted aromatic;

R₂is a polyamine; and is

X is CO, NHCO, NHCS or SO₂

In another embodiment of the above composition, R₂Has the formula

NH(CH₂)_nNH(CH₂)_pNH(CH₂)_qNHR₃

Wherein

n, p and q vary independently and n ═ p ═ q ═ 1 to 12; and is

R₃Is H;C_1-10an alkyl group; c_1-10An alkenyl group; c_1-10An alkynyl group; an alicyclic group; an aryl group; aryl-substituted alkyl, alkenyl or alkynyl; alkyl-, alkenyl-, or alkynyl-substituted aryl; guanidino; a heterocyclic group; heterocyclyl-substituted alkyl, alkenyl or alkynyl; and alkyl-, alkenyl-, or alkynyl-substituted heterocyclic groups.

The above composition may further comprise a linker linking X and R₂And another group Y such that the composition has formula 14 b:

R₁-X-L-Y-R₂

wherein

L is C_1-10An alkyl group; c_1-10An alkenyl group; c_1-10An alkynyl group; an alicyclic or heterocyclic group;

x is CO, SO₂NHCO or NHCS; and is

Y is CONH, SO₂NH、NHCO、NHCONH、NHCSNH、NHSO₂、SO₂O or S.

In the above composition, R₁May have the formula:

wherein

R₄、R₅、R₆、R₇And R₈Independently H, OH, halogen, NO₂、NH₂、NH(CH)_nCH₃、N((CH)_nCH₃)₂、CN、(CH)_nCH₃、O(CH)_nCH₃、S(CH2)_nCH₃、NCO(CH₂)_nCH₃、O(CF₂)_nCF₃Or CO-O (CH)_nCH₃Wherein n is 0 to 10.

Or, R₁Has the formula：

Wherein

R₄And R₅Independently H, OH, halogen, NO₂、NH₂、NH(CH)_nCH₃、N((CH)_nCH₃)₂、CN、(CH)_nCH₃、O(CH)_nCH₃、S(CH2)_nCH₃、NCO(CH₂)_nCH₃、O(CF₂)_nCF₃Or CO-O (CH)_nCH₃Wherein n is 0 to 10.

In yet another embodiment, R₁Having the formula:

wherein

r and s vary independently and r-s-0 to 6;

R₄、R₅、R₆、R₇、R₈and R₉Independently H, OH, halogen, NO₂、NH₂、NH(CH)_nCH₃、N((CH)_nCH₃)₂、CN、(CH)_nCH₃、O(CH)_nCH₃、S(CH2)_nCH₃、NCO(CH₂)_nCH₃、O(CF₂)_nCF₃Or CO-O (CH)_nCH₃Wherein n is 0 to 10;

q is CONH, SO₂NH、NHCO、NHCONH、NHCSNH、NHSO₂、SO₂O or S.

Furthermore, R₁May have the formula:

wherein

r and s vary independently and are 0 to 6;

R₄、R₅、R₆and R₇Independently H, OH, NO₂、NH₂、NH(CH)_nCH₃、N((CH)_nCH₃)₂、CN、(CH)_nCH₃、O(CH)_nCH₃、S(CH2)_nCH₃、NCO(CH₂)_nCH₃、O(CF₂)_nCF₃Or CO-O (CH)_nCH₃Wherein n is 0 to 10; and is

Q is CONH, SO₂NH、NHCO、NHCONH、NHCSNH、NHSO₂、SO₂O or S.

In the above composition, R₁Can be selected from the group consisting of naphthalene, phenanthrene, anthracene, pyrene, dibenzofuran, acridine, 2, 1, 3-benzothiadiazole, quinoline, isoquinoline, benzofuran, indole, carbazole, fluorene, 1, 3-benzodiazine, phenazine, phenoxazine, phenothiazine, adamantane, camphor, piperidine, alkylpiperazine, morpholine, cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, thiophene, furan, pyrrole, alkyl-1, 2-oxadiazole, alkylimidazole, alkyl-1H-1, 2, 3-triazole, alkyl-1H-1, 2, 3, 4-tetrazole, thiazole, oxazole, 1, 3, 4-thiadiazole, pyridyl, pyrimidine, 1, 2-diazine, 1, 4-diazine and 1, 3, 5-triazine, 4-dimethylaminoazobenzene, 3-phenyl-5-methylisoxazole, 3- (2-chlorophenyl) -5-methylisoxazole, 2- (4-chlorophenyl) -6-methyl-7-chloroquinoline, 6-chloro-imidazo [1, 3, 78 ] imidazole, 891, 4-chloro- β]Thiazole, α -methyl cinnamic acid and 2- [1, 2-dihydro-2H-1, 4-benzodioxepin-yl]And (b) a thiazole.

R1 may also be a D or L amino acid.

Also provided are the above compositions, wherein R1 has a formula selected from the group consisting of

(A)R₁₂-R₁₃-Y₁-R₁₄

(B)R₁₂-Y₁-R₁₃-Z₁-R₁₄

(C)

(D)

Wherein

R₁₂And R₁₃Independently of each other H, naphthalene, phenanthrene, anthracene, pyrene, dibenzofuran, acridine, 2, 1, 3-benzothiadiazole, quinoline, isoquinoline, benzofuran, indole, carbazole, fluorene, 1, 3-benzodiazine, phenazine, phenoxazine, phenothiazine, adamantane, camphor, piperidine, alkylpiperazines, morpholine, cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, thiophene, furan, pyrrole, alkyl-1, 2-oxadiazole, alkylimidazole, alkyl-1H-1, 2, 3-triazole, alkyl-1H-1, 2, 3, 4-tetrazole, thiazole, oxazole, 1, 3, 4-thiadiazole, pyridyl, pyrimidine, 1, 2-diazine, 1, 4-diazine and 1, 3, 5-triazine, 4-dimethylaminoazobenzene, 3-phenyl-5-methylisoxazole, 3- (2-chlorophenyl) -5-methylisoxazole, 2- (4-chlorophenyl) -6-methyl-7-chloroquinoline, 6-chloro [2, 1- β ] -chloroimidazole]Thiazole, α -methyl cinnamic acid or 2- [1, 2-dihydro-2H-1, 4-benzodioxepin-yl]A thiazole;

and further, in the above-described manner,

wherein R in the formulae (A), (B) and (D)₁₂、R₁₃Or both, optionally substituted with one or more of: OH, halogen, NO₂、NH₂、NH(CH)_nCH₃、N((CH)_nCH₃)₂、CN、(CH)_nCH₃、O(CH)_nCH₃、S(CH2)_nCH₃、NCO(CH₂)_nCH₃、O(CF₂)_nCF₃Or CO-O (CH)_nCH₃Wherein n is 0 to 10

R₁₄And R₁₅And R in the formula (C)₁₃Independently is (CH)₂)_n、(CH₂)_nCH＝CH、(CH₂)_n(CH＝CH)_mCO or (CH)₂)_nCO, wherein n-0 to 5 and m-1 to 3;

Y₁and Z₁Independently CONH, SO₂NH、NHCO、NHCONH、NHCSNH、NHSO₂、SO₂-NHSO₂、SO₂O, S or COO;

or

When R is₁When having formula (A) or (B), Y1 represents R₁₂With R and C or N atoms of₁₃A bond between C or N atoms of (a), and Z₁Represents R₁₃With R and C or N atoms of₁₄A bond between the C or N atoms of (a); or

When R is₁Having the formula (C) or (C), Y₁Represents R₁₃And a bond between C and C or N atom of (A), and Z₁Represents R₁₄A bond between the C and C or N atoms of (a); or

When R is₁When having formula (D), Y₁Represents R₁₂With R and C or N atoms of₁₄A bond between C or N atoms of (a), and Z₁Represents R₁₃With R and C or N atoms of₁₅A bond between the C or N atoms of (a).

In the above composition, R₂Preferably of the formula

NHCH(Z₁)(CH₂)_nNH(CH₂)_pNH(CH₂)_qCH(Z1)NHR₃

Wherein

n, p and q vary independently and n ═ p ═ q ═ 1 to 12; and is

R₃Is H; c_1-10An alkyl group; c_1-10An alkenyl group; c_1-10An alkynyl group; an alicyclic group; aryl-substituted alkyl, alkenyl or alkynyl; alkyl-, alkenyl-, or alkynyl-substituted aryl; guanidino or heterocyclyl; and is

Z is CH₃、CH₂CH₃Or a cyclopropyl group.

In another embodiment, R₂Having the formula:

wherein

x is 1 to 4; y is equal to 1 to 3,

R₁₀and R₁₁Independently H, (CH)₂)nNHR₁₂Or (CH)₂)_kNH(CH₂)1NHR₁₂Wherein n-k-1-10, and R₁₂Is H or C (N ═ H) NH₂。

In the above composition, R₂Preferably selected from the group consisting of: n1-acetylspermine, N1-acetylspermidine, N8-acetylspermidine, N ' -guanidinoprospermine, cadaverine, aminopropylcadaverine, homo-spermidine, caldine (horsperamine), 7-hydroxypspermidine, thermoamine (norspermine), thermospermine, canavalinate, aminopropyl homo-spermidine, N ' -bis (3-aminopropyl) cadaverine, aminopentyl norspermidine, N4-aminopropyl norspermidine, N4-aminopropyl spermidine, thermophilic pentylamine, homothermophilic pentylamine, N4-bis (aminopropyl) norspermidine, thermopentylamine, N4-bis (aminopropyl) spermidine, thermophilic hexylamine, homohexylamine, N- (3-aminopropyl) -1, 3-propanedioxane, N ' -bis (3-aminopropyl) ethylenediamine, N, N '-bis (3-aminopropyl) -1, 4-piperazine, N' -bis (3-aminopropyl) -1, 3-piperazineOxazines, N '-bis (3-aminopropyl) -1, 3-propanedioxane, N' -bis (2-aminoethyl) -1, 3-propanedioxane, tris (3-aminopropyl) amine and tris (aminoethyl) amine.

Compounds include, but are not limited to:

further disclosures can be found in WO99/03823, the disclosure of which is incorporated by reference in its entirety as if written herein.

Yet another compound includes, but is not limited to:

additional disclosures can be found in the following documents: ackermann, JM; pegg, AE; McCloskey, DE; progress Cell Cycle Research, 2003, Vol.5, 461-; ekelund, S; nygren, P; larsson, R; biochemical Pharmacology, 2001, 61, 1183-1193; huang, Y; piedgie, A; casero Jr, RA; davidson, NE; Anti-Cancer Drugs, 2005, 16, 229-; and Marton, JL; annu. rev. pharmacol toxicol.1995, 35, 55-91; the disclosure of which is incorporated by reference in its entirety as if written herein.

The polyamine analogs depicted above can be prepared as salts and as free bases. In certain embodiments, the salt is a hydrochloride salt. In certain embodiments, a coordinating ion pair (e.g., H)⁺Cl^-) The number of (a) will be proportional to the number of amino groups in the polyamine. This coordination usually takes place at the amino group, forming for example NH₃ ⁺Cl^-A group. However, not every amino group may be coordinated. For example, if an amino group is close to an electron withdrawing group such as a carbonyl group or a sulfonyl group, it cannot retain a sufficient electron density of a complex ion. In other embodiments, the number of coordinating ions will be proportional to the number of primary and/or secondary amino groups in the polyamine.

Additional compounds that may be used in the methods and compositions described herein include: naturally occurring polyamines, polyamine analogs, polyamine biosynthesis inhibitors, and polyamine transport inhibitors found in prokaryotic and eukaryotic cells.

Naturally occurring polyamines found in prokaryotic and eukaryotic cells include, but are not limited to: putrescine, spermidine, spermine, diaminopropane, cadaverine, norspermidine, aminopropylcadaverine, spermine, norspermine, thermal spermine, aminopentyl norspermine, bis (aminopropyl) cadaverine, aminopropyl spermine, 30 canavaline tetramine, spermine, thermophilic pentylamine, aminopropyl canavalin, bis (aminopropyl) spermine, bis (aminobutyl) norspermine, aminobutyl canavaline tetramine, aminopropyl spermine, semamylamine, N5-aminobutyl spermine, thermophilic hexylamine, thermal hexylamine, homohexylamine, agmatine, and N6-methylguanidine. See, e.g., Morgan d.m.l., 1999, Molecular Biotechnology 11: 229.

polyamine analogs include, but are not limited to: BE-4444[1, 19-bis (ethylamino) -5, 10, 15-triazadecane ]; BE-3-3-3[ N1, N11-diethyl norspermine; DENSPM; 1, 11-bis (ethylamino) -4, 8-diazaundecane; a hot amine; Wamer-Parke-Davis ]; BE-3-3[ N1, N7-bis (ethyl) norspermidine ]; BE-3-4[ N1, N8-bis (ethyl) spermidine ]; BE44[ N1, N9-bis (ethyl) spermidine ]; BE-343[ N1, N12-bis (ethyl) spermine; diethyl spermine-N1-N12; DESPM ]; BE-373[ N, N' -bis (3-ethylamino) propyl) -1, 7-heptanediamine, Merrell-Dow ]; BE-4-4-4[ N1, N14-bis (ethyl) spermine; diethyl spermine-N1-N1-1 ]; BE-3-4-4-3[1, 17-bis (ethylamino) -4, 9, 14 triazadecane ]; BE-4-3-3-4[1, 17-bis (ethylamino) -5, 9, 13-triazaheptadecane ]; and 1, 12-Mez-SPM [1, 12-dimethyl spermine ] (WOO 2007/040535).

The polyamine synthesis Inhibitors include, but are not limited to, Inhibitors of ornithine decarboxylase such as DFMO, acetylenic putrescine, 1-aminooxy-3-aminopropane, degradative Enzyme, 2-butylputrescine, cadaverine, L-paracaseinidine, 5 '-deoxy-5' - [ N-methyl-N- [3 (aminooxy) ethyl ] adenosine, 5 '-deoxy-5' - [ N-methyl-N- [3- (hydrazinopropyl) amino ] diaminopropane, 1, 3-diamino-2-propanol, 2-difluoromethyl putrescine, difluorophenylethyl (4-aminopropylamidino), 2, 3-dimethylputrescine, N-dimethylputrescine, 2-ethylputrescine, (+ or-) - α fluoromethylornithine, 2-fluoromethylputrescine, 2-hexylputrescine, 2-hydrazinoornithine, ibuprofen, D-methylacetyleneamine, methylglyoxal bis (3-aminopropylamidino hydrazone), 2-methylglycinylhydrazinothiohydrazinothiohydrazinothiohydrazinothiohydrazinodine), thiohydrazinothiohydrazinothiohydrazinodine, thiohydrazinodine, thiodine.

Additional spermine analogs include N- (2-mercaptoethyl) spermine-5-carboxamide (MESC), its disulfide form, i.e., 2, 21-dithiobis (N-ethyl-spermine-5-carboxamide) (DESC), and N- [2, 2, 1-dithio (ethyl 1, 1-aminoethyl) ] spermine-5-carboxamide (DEASC). (WO98/17623)

Polyamine effectors that are small molecule inhibitors or modulators of key enzymes in the polyamine biosynthetic pathway include, but are not limited to, ODC inhibitors such as Difluoromethylornithine (DFMO), α -monofluoromethylornithine (MFMO), and methylacetylpyridinium putrescine (MAP), AdometDC inhibitors such as S- (5-deoxy-5-Adenosyloxy) Methylthioethylhydroxylamine (AMA), 5-deoxy-5- [ (2-aminooxyethyl) methylamino ] adenosine (MAOEA), and methylglyoxal bis (amidinohydrazone) (MGBG), spermidine synthase inhibitors such as S-adenosine 1, 8-diamino-3-thiooctane (AdoDATO), cyclohexylamine, and butylamine, spermine synthase inhibitors such as S-adenosine-1, 12-diamino-3-thio-9-azadodecane (AdoDATAD), and N- (N-butyl) -1, 3-diaminopropane (BDAP).

In certain embodiments, the polyamine effector is a polyamine or arginine analog bearing a functional group that confers cellular or DNA protection to the molecule or modulates a polyamine biosynthetic or catabolic pathway. Compounds having this property include, but are not limited to, amifostine, NG-hydroxy-arginine (NORA), N1, N11-bis (ethyl) norspermine (BE-3-3-3), N12-bis (ethyl) spermine (BE-3-4-3), N, N-bis [3- (ethylamino) -propyl ] -1, 7 heptanediamine (BE-3-7-3), BE-3-3-3, BE-3-4-3, BE-3-7-3, N1-ethyl-N11-propargyl 4, 8-diazamentane, and the analogues SL-11141 and SL-II050 (structures listed in one or more of U.S. Pat. No. 5,889,061, Valasinas et al, 2001, supra, WO00/66587, and WO 02/38105). Further disclosures can be found in WO03/013245, the disclosure of which is incorporated by reference in its entirety as if written herein.

As used herein, the following terms have the indicated meanings.

Cytokines can include, but are not limited to, IL-1, IL1-Ra, IL-2, IL-6, IL8, IFN γ, IP-10, IL-17, MCP-1, MMP-9, MIP-1 β, TNF- α, TGF β, CRP, OPN, and RANTES.

When numerical ranges are disclosed and use is made of "from n₁To n₂Is "or" at n₁And n₂Notation of "in between (unless otherwise specified, n₁And n₂Is a number), this notation is intended to include the number itself and ranges intermediate thereto. This range can be an integer or continuous number between and including the endpoints. For example, a range of "2 to 6 carbons" is intended to include two, three, four, five, and six carbons, as carbons are expressed in integer units. In contrast, for example, a range of "1 to 3 μ M (micromolar)" is intended to include 1 μ M, 3 μ M, and any value therebetween to any number of significant values (e.g., 1.255 μ M, 2.1 μ M, 2.9999 μ M, etc.).

The term "about" as used herein qualifies a value that it modifies, meaning that such a value is variable within a margin of error. When no specific margin of error is recited, such as the standard deviation of the mean value given in a chart or table of data, the term "about" should be understood to mean that the range including the stated value and the range including by rounding up or down to that value when significant figures are considered.

The term "significantly" as used herein is intended to mean predominantly or of an overwhelming nature such that any adverse or diminishing features reach a nonsensical level. For example, a composition that is "substantially" free of water need not absolutely contain all trace amounts of water, but will be sufficiently anhydrous that any remaining water does not affect the composition in any significant way. By way of further example, a "significant dose-limiting side effect" may be a side effect that limits the dose to a level below that required for therapeutic efficacy.

The term "disease" as used herein is intended to be generally synonymous with and interchangeable with the terms "disorder", "syndrome" and "condition" (as in medical conditions), as they all reflect an abnormal state of the body or part thereof of a human or animal that impairs normal function, usually manifested by distinguishing signs and symptoms and giving the human or animal a shortened life span or reduced quality of life.

A "proliferative disorder" may be any disorder characterized by dysregulated cell proliferation. Examples include cancer, psoriasis and atopic dermatitis.

As used herein, "hyperalgesia" means an increased sensitivity to pain, and can be considered as a type of pain or a degree of pain-related behavior.

As used herein, "progressive" multiple sclerosis refers to a form of disease that progresses over a period of time to an increasingly worsening disease state. Progressive MS includes, for example, primary progressive MS, secondary progressive MS, and MS with progressive relapse. These subtypes may or may not be characterized by a stage-wise flare-up of the disease, but each is associated with symptoms that increase over time, such as increased demyelination or pain and decreased motor capacity.

As used herein, reference to "treating" a patient is intended to include prophylaxis. Treatment may also be preliminary in nature, i.e., it may include prevention of disease. Prevention of a disease may involve complete protection from the disease, for example in the case of prevention of infection with a pathogen, or may involve prevention of disease progression. For example, prevention of a disease does not necessarily mean complete exclusion of any effect at any level associated with the disease, but may instead mean prevention of the symptoms of the disease to a clinically significant or detectable level. Prevention of a disease may also mean preventing the disease from progressing to a later stage of the disease.

The term "combination therapy" means the administration of two or more therapeutic agents to treat a therapeutic condition or condition described in this disclosure. The administration includes co-administering these therapeutic agents in a substantially simultaneous manner, such as in a single capsule with a fixed ratio of active ingredients or in multiple separate capsules for each active ingredient. In addition, the administration also includes the use of one type of therapeutic agent in a continuous manner. In either case, the treatment regimen will provide a beneficial effect of the drug combination in treating the conditions or disorders described herein.

The term "patient" is generally synonymous with the term "subject" and means an animal, including all mammals and humans, having a disease, disorder or condition that can be treated according to the methods disclosed herein. Examples of patients include humans, livestock such as cows, goats, sheep, pigs and rabbits, and companion animals such as dogs, cats, rabbits and horses. Preferably, the patient is a human.

An "effective amount" or "therapeutically effective amount" is an amount of a compound (e.g., MGBG, a polyamine analog, a polyamine biosynthesis inhibitor, or any agent) sufficient to achieve a desired effect in a treated subject. For example, this can be an amount necessary to treat a disease, disorder, condition, or adverse state (e.g., pain or inflammation) or otherwise measurably alter or alleviate a symptom, marker, or mechanism of a disease, disorder, condition, or adverse state. As just one example, an effective amount for treating pain is an amount sufficient to prevent, delay onset of, or reduce pain or one or more pain-related symptoms in a subject, as measured by methods known in the art. Similar methods of assessing response to treatment of many diseases are well known in the art. The effective amount of the compounds of the present invention may vary depending on the route of administration and the dosage form. In addition, the specific dosage can be adjusted according to the following: a condition of the disease, the age, weight, general health, sex and diet of the subject, dosage interval, route of administration, rate of excretion and combination of agents.

The term "low dose" with respect to a low dose formulation of a drug or a method of treatment specifically with a "low dose" drug means that at least one indication is a sub-therapeutic dose, or a fraction of the dose that is generally administered for at least one indication. Taking the example of a drug for the treatment of a proliferative disorder, a low dose formulation for the treatment of i.e. multiple sclerosis, may be part of the dose for the treatment of an aggressive cancer. In this way, the dose for one disease may be a sub-therapeutic amount for another disease. Alternatively, for drugs that are therapeutic at different doses and effective over a range of doses in different individuals or populations, the low dose may simply be the dose for the low identified therapeutic efficacy. Chronic diseases represent embodiments that can be treated with low dosage formulations and methods. In addition, a sub-therapeutic amount of a drug may be used in combination with one or more other drugs (per se in therapeutic or sub-therapeutic amounts) to produce a potentiated combined preparation or treatment, i.e., more effective than the intended effect of the sum of the drugs administered alone. The low dose for the treatment of an indication may be two, three, four, five, six, seven, eight, nine, ten, fifteen, twenty, thirty, forty, fifty, possibly one hundred times less than the therapeutic dose for a different indication.

The phrase "therapeutically effective" is intended to qualify the amount of active ingredient used in treating a disease or disorder or to achieve a clinical endpoint.

The term "therapeutically acceptable" refers to those compounds (or salts, prodrugs, tautomers, zwitterionic forms, etc.) that are suitable for use in contact with the tissues of a subject without excessive toxicity, irritation, and allergic response, commensurate with a reasonable benefit/risk ratio, and effective for their intended use.

The term "drug" is used interchangeably herein with "compound" and "agent".

When a compound is referred to herein as "not a T cell modulator," it means that any direct activity against T cells is negligible and/or secondary to activity attributable to another leukocyte subtype. In certain embodiments, a cell that is "not a T cell modulator" will be a cell of myeloid lineage. In certain embodiments, such cells will be dendritic cells, monocytes or macrophages.

The phrase "a reduced incidence of at least one side effect" as used herein means reduced to a significant extent. Significance can be demonstrated statistically (i.e., by non-overlapping standard deviations or appropriate confidence intervals). The reduced significance of the incidence of side effects can also be demonstrated with reference to qualitative measures, such as, for example, the ability to achieve or maintain a therapeutic dose without dose-limiting toxicity (in all patients or in a subpopulation of patients), the ability to prevent or delay disease recurrence or progression, or patient preference.

The phrase "approved for the treatment of a demyelinating disease" as used herein means approved by the drug regulatory agency (in the united states, europe or any EPO country, japan, canada or australia) for the treatment of a demyelinating disease. In any of the embodiments disclosed herein, the demyelinating disease may specifically be multiple sclerosis.

The term "SAMDC inhibitor" means an inhibitor of the enzyme S-adenosylmethionine decarboxylase. MGBG is considered one such SAMDC inhibitor, and other polyamines, polyamine analogs, and polyamine biosynthesis inhibitors may also be SAMDC inhibitors.

As used herein, "polyamines" are any group of aliphatic linear amines derived from amino acid biosynthesis; polyamines are reviewed in Marton et al (1995) ann.rev.pharm.toxicol.35: 55-91. "polyamine" generally means a naturally occurring polyamine or a polyamine that is naturally produced in eukaryotic cells. Examples of polyamines include putrescine, spermidine, spermine, and cadaverine.

As used herein, a "polyamine analog" is an organic cation that is structurally similar but not identical to a naturally occurring polyamine, such as spermine and/or spermidine and their precursors, the diamine putrescine. The polyamine analogs can be branched or unbranched, or incorporate a cyclic moiety. The polyamine can comprise primary, secondary, tertiary or quaternary amino groups. In one embodiment, all nitrogen atoms of the polyamine analog are independently secondary, tertiary, or quaternary amino groups, but are not so limited. Polyamine analogs can include imines, amidines, and guanidines in place of amine groups. The term "polyamine analog" includes stereoisomers, salts and protected derivatives of the polyamine analog.

"stereoisomers" are any optical isomers of a compound, including enantiomers and diastereomers. Unless otherwise stated, the structural formulae of the compounds are intended to include all possible stereoisomers.

"salts" or "pharmaceutically acceptable salts" are compounds formed by replacing one or more hydrogen atoms with an element or group, which consists of an anion and a cation, which are typically ionized in water; for example, salts are formed by neutralizing acids with bases. Examples of salts include, but are not limited to, halides (e.g., chloride, bromide, or iodide), nitrates, sulfates, bisulfates, phosphates, acid phosphates, isonicotinates, acetates, lactates, salicylates, citrates, tartrates, pantothenate, bitartrates, ascorbic acid, succinates, maleates, gentisates, fumarates, gluconates, glucuronates, gluconates, formates, benzoates, glutamates, ethanesulfonates, benzenesulfonates, p-toluenesulfonates, and pamoate (i.e., 1, 1' -methylene-bis- (2-hydroxy-3-naphthoate)).

"protected derivative" is used to refer to a compound protected with a protecting group. "protecting group" refers to a chemical group that exhibits the following characteristics: 1) selectively reacting with the desired functional group in good yield (preferably at least 80%, more preferably at least 90%, more preferably at least 95%, more preferably at least 99%) to obtain a protected substrate which is stable to the intended reaction requiring protection; 2) optionally removed from the protected substrate to provide the desired functional group; and 3) can be removed in good yield (preferably at least 80%, more preferably at least 90%, more preferably at least 95%, more preferably at least 99%) by reagents compatible with other functional groups present or generated in the intended reaction. Examples of suitable protecting Groups can be found in Greene et al (1991) Protective Groups in Organic Synthesis, 2 nd edition (John Wiley&Sons, inc., New York). Exemplary protecting groups for amino functional groups include, but are not limited to, mesitylenesulfonyl (MesSO)₂) Benzyloxycarbonyl (CBz), tert-butyloxycarbonyl (Boc), tert-butyldimethylsilyl (TBDIMS), 9-fluorenylmethyloxycarbonyl (Fmoc) or a suitable photolabile protecting group such as 6-nitroveratryloxycarbonyl (Nvoc).

The term "acyl", alone or in combination, as used herein, refers to a carbonyl group attached to an alkenyl group, an alkyl group, an aryl group, a cycloalkyl group, a heteroaryl group, a heterocycle, or any other moiety attached to the carbonyl group whose atom is carbon. "acetyl" means-C (O) CH₃A group. "alkylcarbonyl" or "alkanoyl" refers to an alkyl group attached to the parent molecular moiety through a carbonyl group. Examples of the group include methylcarbonyl and ethylcarbonyl. Examples of acyl groups include formyl, alkanoyl and aroyl.

The term "alkenyl" as used herein, alone or in combination, refers to a straight or branched chain hydrocarbon radical having one or more double chains and containing from 2 to 20 carbon atoms. In certain embodiments, the alkenyl group will comprise 2 to 6 carbon atoms. The term "alkenylene" refers to a carbon-carbon double bond system attached at two or more positions, such as ethenylene [ (-CH ═ CH-), (-C:: C-) ]. Examples of suitable alkenyl groups include ethenyl, propenyl, 2-methylpropenyl, 1, 4-butadienyl, and the like. Unless otherwise specified, the term "alkenyl" may include "alkenylene" groups.

The term "alkoxy" as used herein, alone or in combination, refers to an alkyl ether group, wherein the term alkyl is defined below. Examples of suitable alkyl ether groups include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, sec-butoxy, tert-butoxy and the like.

The term "alkyl" as used herein, alone or in combination, refers to straight or branched chain alkyl groups containing from 1 to 20 carbon atoms. In certain embodiments, the alkyl group will contain 1 to 10 carbon atoms. In other embodiments, the alkyl group will contain 1 to 6 carbon atoms. Alkyl groups may be optionally substituted as defined herein. Examples of alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, hexyl, octyl, nonyl, and the like. The term "alkylene" as used herein, alone or in combination, refers to a saturated aliphatic group derived from a straight or branched chain saturated hydrocarbon attached at two or more positions, such as methylene (-CH)₂-). Unless otherwise specified, the term "alkyl" may include "alkylene" groups.

The term "alkylamino", as used herein, alone or in combination, refers to an alkyl group attached to the parent molecular moiety through an amino group. Suitable alkylamino groups can be mono-or di-alkylated to form groups such as, for example, N-methylamino, N-ethylamino, N-dimethylamino, N-ethylmethylamino, and the like.

The term "alkynyl", alone or in combination, as used herein, refers to a straight or branched chain hydrocarbon group having one or more triple bonds and containing from 2 to 20 carbon atoms. In certain embodiments, the alkynyl group contains 2 to 6 carbon atoms. In other embodiments, the alkynyl group contains 2 to 4 carbon atoms. The term "alkynylene" refers to a carbon-carbon triple bond attached at two positions, such as ethynylene (-C:: C-, -C ≡ C-). Examples of alkynyl groups include ethynyl, propynyl, hydroxypropynyl, butyn-1-yl, butyn-2-yl, pentyn-1-yl, 3-methylbutyn-1-yl, hexyn-2-yl, and the like. Unless otherwise specified, the term "alkynyl" may include "alkynylene" groups.

The terms "amido" and "carbamoyl", alone or in combination, as used herein, refer to an amino group, as described below, appended to the parent molecular moiety through a carbonyl group, or vice versa. The term "C-amido", alone or in combination, as used herein, refers to the group-C (o) N (RR '), wherein R and R' are as defined herein or as defined by the specific recitation of "R" groups as indicated. The term "N-amido", alone or in combination, as used herein, refers to an rc (o) N (R ') -group, wherein R and R' are as defined herein or by the specific recitation of an "R" group as indicated. The term "acylamino" as used herein, alone or in combination, includes an acyl group attached to the parent moiety through an amino group. An example of an "amido group" is an acetamido group (CH)₃C(O)NH-)。

The term "amino" as used herein, alone or in combination, refers to-NRR ', wherein R and R' are independently selected from the group consisting of: hydrogen, alkyl, acyl, heteroalkyl, aryl, cycloalkyl, heteroaryl, and heterocycloalkyl, any of which may be optionally substituted on its own. In addition, R and R' may combine to form a heterocycloalkyl group, both of which may be optionally substituted.

The term "aryl" as used herein, alone or in combination, means a carbocyclic aromatic system containing one, two or three rings, wherein the polycyclic ring systems are fused together. The term "aryl" includes aromatic groups such as phenyl, naphthyl, anthryl and phenanthryl.

The terms "arylalkyl" or "aralkyl," alone or in combination, as used herein, refer to an aryl group attached to the parent molecular moiety through an alkyl group. The term "carboxy" or "carboxyl" as used herein refers to-c (o) OH or the corresponding "carboxylate" anion, such as carboxylate. "O-carboxy" refers to an RC (O) O-group, wherein R is as defined herein. "C-carboxy" refers to the group-C (O) OR, wherein R is as defined herein.

The term "cyano", alone or in combination, as used herein, refers to-CN.

The term "cycloalkyl" or alternatively "carbocycle" or "alicyclic", alone or in combination, as used herein, refers to a saturated or partially saturated monocyclic, bicyclic or tricyclic alkyl group in which each cyclic moiety contains from 3 to 12 carbon atom ring members and may optionally be an optionally substituted benzo-fused ring system as defined herein. In certain embodiments, the cycloalkyl group will contain from 5 to 7 carbon atoms. Examples of such cycloalkyl groups include cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyl, tetrahydronaphthyl, indanyl, octahydronaphthyl, 2, 3-dihydro-1H-indenyl, adamantyl, and the like. As used herein, "bicyclic" and "tricyclic" are intended to include fused ring systems, such as decalin, octahydronaphthalene; and polycyclic (multicenter) saturated or partially unsaturated types. The latter type of isomer is generally exemplified by bicyclo [1, 1, 1] pentane, camphor, adamantane, and bicyclo [3, 2, 1] octane.

The term "halo" or "halogen" as used herein, alone or in combination, refers to fluoro, chloro, bromo, or iodo.

The term "heteroalkyl," alone or in combination, as used herein, refers to a stable straight or branched chain or cyclic hydrocarbon radical, or combinations thereof, fully saturated containing from 1 to 3 unsaturations, consisting of the recited number of carbon atoms and from 1 to 3 heteroatoms selected from the group consisting of O, N and S, and wherein the nitrogen and sulfur atoms may optionally be oxidized and the nitrogen heteroatom may optionally be quaternized. The heteroatoms O, N and S can be placed at any internal position of the heteroalkyl group. Up to two heteroatoms may be consecutive, e.g. like-CH₂-NH-OCH₃。

The term "heteroaryl" as used herein, alone or in combination, refers to a 3 to 15 membered unsaturated heteromonocyclic or fused monocyclic, bicyclic or tricyclic ring system wherein at least one fused ring is aromatic and contains at least one atom selected from the group consisting of O, S and N. In certain embodiments, the heteroaryl group will contain from 5 to 7 carbon atoms. The term also includes fused polycyclic groups in which a heterocycle is fused to an aryl ring, in which a heteroaryl ring is fused to another heteroaryl ring, in which a heteroaryl ring is fused to a heterocycloalkyl ring, or in which a heteroaryl ring is fused to a cycloalkyl ring. Examples of heteroaryl groups include pyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazolyl, pyranyl, furanyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, thiadiazolyl, isothiazolyl, indolyl, isoindolyl, indolizinyl, benzimidazolyl, quinolinyl, isoquinolinyl, quinoxalinyl, quinazolinyl, indazolyl, benzotriazolyl, benzodioxolyl, benzopyranyl, benzoxazolyl, benzodiazolyl, benzothiadiazolyl, benzofuranyl, benzothiophenyl, chromonyl, coumarinyl, benzopyranyl, tetrahydroquinolinyl, tetrazolopyridazinyl, tetrahydroisoquinolinyl, thienopyridyl, furopyridyl, pyrrolopyridyl, and the like. Exemplary tricyclic heterocyclic groups include carbazolyl, benzindolyl, phenanthrolinyl, dibenzofuranyl, acridinyl, phenanthridinyl, xanthenyl, and the like.

The terms "heterocycloalkyl" and, interchangeably, "heterocycle" as used herein, alone or in combination, each refer to a saturated, partially unsaturated, or fully unsaturated monocyclic, bicyclic, or tricyclic heterocyclic group containing at least one heteroatom as a ring member, wherein each said heteroatom may be independently selected from the group consisting of nitrogen, oxygen, and sulfur. In certain embodiments, the heterocycloalkyl group will contain from 1 to 4 heteroatoms as ring members. In other embodiments, the heterocycloalkyl group will contain 1 to 2 heteroatoms as ring members. In certain embodiments, the heterocycloalkyl group will contain from 3 to 8 ring members in each ring. In other embodiments, the heterocycloalkyl group will contain from 3 to 7 ring members in each ring. In still other embodiments, the heterocycloalkyl group will contain from 5 to 6 ring members in each ring. "heterocycloalkyl" and "heterocycle" are intended to include sulfones, sulfoxides, N-oxides of tertiary nitrogen ring members, and carbocyclic fused and benzo fused ring systems; in addition, both terms also include systems in which a heterocyclic ring is fused to an aryl or another heterocyclic group as defined herein. Examples of heterocyclic groups include aziridinyl, azetidinyl, 1, 3 benzodioxolyl, isoindolyl, isoquinolinyl, cinnolinyl, benzodioxinyl, dihydro [1, 3] oxazolo [4, 5-b ] pyridyl, benzothiazolyl, indolinyl, dihydropyridyl, 1, 3-dioxanyl, 1, 4-dioxanyl, 1, 3-dioxolanyl, isoindolyl, morpholinyl, piperazinyl, pyrrolidyl, tetrahydropyridinyl, piperidinyl, thiomorpholinyl and the like. Unless expressly prohibited, heterocyclic groups may be optionally substituted.

The term "lower", alone or in combination, as used herein, is intended to mean, without additional specific definition, containing from 1 to and including 6 carbon atoms.

The term "sulfonyl", alone or in combination, as used herein, refers to-S (O)₂-。

Any definition herein may be used in combination with any other definition to describe a composite structural group. By convention, any trailing element defined is the one that is connected to the parent moiety. For example, a complex group alkylamido would represent an alkyl group attached to the parent molecule via an amido group, and the term alkoxyalkyl would represent an alkoxy group attached to the parent molecule via an alkyl group.

When a group is defined as "null", that means lacking the group.

The term "optionally substituted" means that the preceding group may be substituted or unsubstituted. When substituted, the substituents of the "optionally substituted" group may include, but are not limited to, independently selected from the following groups or groups, alone or in combinationSubstituents for the groups specified in the formula: lower alkyl, lower alkenyl, lower alkynyl, lower alkanoyl, lower heteroalkyl, lower heterocycloalkyl, lower haloalkyl, lower haloalkenyl, lower haloalkynyl, lower perhaloalkyl, lower perhaloalkoxy, lower cycloalkyl, phenyl, aryl, aryloxy, lower alkoxy, lower haloalkoxy, oxo, lower acyloxy, carbonyl, carboxyl, lower alkylcarbonyl, lower carboxyl ester, lower carbonylamino, cyano, hydrogen, halogen, hydroxyl, amino, lower alkylamino, arylamino, amido, nitro, thiol, lower alkylthio, lower haloalkylthio, lower perhaloalkylthio, arylthio, sulfonate, sulfonic acid, trisubstituted silyl, N₃、SH、SCH₃、C(O)CH₃、CO₂CH₃、CO₂H. Pyridyl, thiophene, furyl, lower carbamates and lower ureas. Two substituents may be linked together to form a fused five-, six-or seven-membered carbocyclic or heterocyclic ring consisting of zero to three heteroatoms, for example to form a methylenedioxy or ethylenedioxy group. The optionally substituted group may be unsubstituted (e.g., -CH)₂CH₃) Completely substituted (e.g., -CF)₂CF₃) Are monosubstituted (e.g., -CH)₂CH₂F) Or substituted at any level between complete or mono-substitution (e.g. -CH)₂CF₃). When a substituent is recited without strict limitation on substitution, both substituted and unsubstituted forms are included. When a substituent is suitable as "substituted", the form of substitution is specifically contemplated. In addition, different groups of optional substituents for a particular moiety may be defined as desired; in these cases, optional substitution will be as defined, often immediately following the phrase "optionally substituted".

Unless otherwise defined, the term R or the term R', appearing alone and without a numerical designation, refers to a moiety selected from the group consisting of: hydrogen, alkyl, cycloalkyl, heteroalkyl, aryl, heteroaryl, and heterocycloalkyl, any of which may be optionally substituted. The R and R' groups should be understood as being optionally substituted as herein described. Irrespective of the R groupWhether or not there is a numerical designation, each R group, including R, R' and RⁿWhere n ═ n (1, 2, 3,. n), each substituent and each term are to be understood as being independent of all others with respect to the choice from the group. When any variable, substituent or term (e.g., aryl, heterocycle, R, etc.) occurs more than one time in a formula or generic structure, its definition in each case is independent of its definition in all other cases. One skilled in the art will further recognize that certain groups may be attached to the parent molecule or may occupy positions in the series of elements from both ends. Thus, by way of example only, asymmetric groups such as-C (O) N (R) -may be attached to the parent moiety at carbon or nitrogen.

Asymmetric centers are present in the compounds disclosed herein. These centers are designated by the symbols "R" or "S", depending on the configuration of the substituents around the chiral carbon atom. It is to be understood that the present invention encompasses all stereochemically isomeric forms, including diastereoisomeric, enantiomeric and epimeric forms, as well as the d-and l-isomers, and mixtures thereof. Individual stereoisomers of compounds may be prepared synthetically from commercially available starting materials containing chiral centers or by preparing mixtures of enantiomeric products, followed by separation, e.g., conversion to a mixture of diastereomers, and then separation or recrystallization, chromatographic techniques, direct separation of enantiomers on chiral chromatographic columns, or any other suitable method known in the art. Starting compounds of a particular stereochemistry are either commercially available or can be prepared and resolved by techniques known in the art. In addition, the compounds disclosed herein may exist as geometric isomers. The present invention includes all cis (cis), trans (trans), cis (syn), trans (anti), contralateral (E) and ipsilateral (Z) isomers and suitable mixtures thereof. In addition, the compounds may exist as tautomers; all tautomers are provided by the present invention. In addition, the compounds disclosed herein can exist in unsolvated as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like. In general, the solvated forms are considered equivalent to unsolvated forms.

The term "bond" refers to a covalent bond between two atoms or between two moieties when the atoms joined by the bond are considered part of a larger substructure. Unless otherwise specified, a bond may be a single bond, a double bond, or a triple bond. The dashed line between two atoms in a molecular diagram indicates that another bond may or may not be present at that position.

The term "prodrug" refers to a compound that becomes more active in vivo. Certain compounds disclosed herein may also exist as prodrugs, such as Hydrolysis in Drug and prodrag Metabolism: chemistry, Biochemistry, and Enzymology (Testa, Bernard and Mayer, Joachim M.Wiley-VHCA, Zurich, Switzerland 2003). Prodrugs of the compounds described herein are structurally modified forms of the compounds that readily undergo chemical changes under physiological conditions to provide the compounds. Alternatively, prodrugs can be converted to compounds by chemical or biochemical means in an ex vivo environment. For example, a prodrug may be slowly converted to a compound when placed in a transdermal patch reservoir with a suitable enzyme or chemical agent. Prodrugs are often useful because, in some cases, they may be easier to administer than the compound or the parent drug. They may be bioavailable, for example by oral administration, whereas the parent drug is not. The prodrug may also have improved solubility in pharmaceutical compositions over the parent drug. A wide variety of prodrug derivatives are known in the art, such as those that rely on hydrolytic cleavage or oxidative activation of the prodrug. An example of a prodrug would be, but is not limited to, a compound that is administered as an ester ("prodrug"), but is then metabolically hydrolyzed to a carboxylic acid, the active entity. Additional examples include peptidyl derivatives of the compounds.

The compounds disclosed herein may exist as therapeutically acceptable salts. The invention includes the compounds listed above in salt form, including acid addition salts. Suitable salts include those formed with organic and inorganic acids. The acid addition salts will generally be pharmaceutically acceptable. However, non-pharmaceutically acceptable salts may have utility in the preparation and purification of the compounds in question. Base addition salts may also be formed and are pharmaceutically acceptable. For a more complete discussion of salt preparation and selection, see Pharmaceutical Salts: properties, section, and Use (Stahl, P.Heinrich.Wiley-VCHA, Zurich, Switzerla nd, 2002).

The term "therapeutically acceptable salt" as used herein means a salt or zwitterionic form of a compound disclosed herein, which is water or oil soluble or dispersible and therapeutically acceptable as defined herein. Salts may be prepared during the final isolation and purification of the compounds or separately by reacting the appropriate compound in free base form with a suitable acid. Representative acid addition salts include acetate, adipate, alginate, L-ascorbate, aspartate, benzoate, benzenesulfonate (bezenesulfonate), bisulfate, butyrate, camphorate, camphorsulfonate, citrate, digluconate, formate, fumarate, gentisate, glutarate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate (isethionate), lactate, maleate, malonate, DL-mandelate, trimesylate, methanesulfonate, naphthylene sulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate, pectate, persulfate, 3-phenylpropionate, Phosphonates, picrates, pivalates, propionates, pyroglutamate, succinates, sulfonates, tartrates, L-tartrates, trichloroacetates, trifluoroacetates, phosphates, glutamates, bicarbonates, p-toluenesulfonate (p-toluenesulfonate), and undecanoates. Further, the bases in the compounds disclosed herein may be quaternized with: methyl, ethyl, propyl and butyl chlorides, bromides and iodides; dimethyl, diethyl, dibutyl and diamyl sulfates; decyl, lauryl, myristyl and sterol chlorides, bromides and iodides; and benzyl and phenethyl bromides. Examples of acids which may be used to form therapeutically acceptable addition salts include inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid and phosphoric acid; and organic acids such as oxalic acid, maleic acid, succinic acid, and citric acid. Salts may also be formed by the complexation of compounds with alkali or alkaline earth ions. Thus, the present invention includes sodium, potassium, magnesium, and calcium salts, and the like, of the compounds disclosed herein.

Base addition salts can be prepared during the final isolation and purification of the compounds by reaction of the carboxyl group with a suitable base such as the hydroxide, carbonate or bicarbonate of a metal cation or with ammonia or an organic primary, secondary or tertiary amine. Cations of therapeutically acceptable salts include lithium, sodium, potassium, calcium, magnesium, and aluminum, as well as non-toxic quaternary amine cations, such as ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine, pyridine, N-dimethylaniline, N-methylpiperidine, N-methylmorpholine, dicyclohexylamine, procaine, dibenzylamine, N-dibenzylphenethylamine, 1-ephedra amine, and N, N' -dibenzylethylenediamine. Other representative organic amines suitable for forming base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine, and piperazine.

While the compounds disclosed herein may be administered as raw chemicals, they may also be delivered as pharmaceutical formulations. Accordingly, provided herein are pharmaceutical formulations comprising one or more of certain compounds disclosed herein, or one or more pharmaceutically acceptable salts, esters, prodrugs, amides, or solvates thereof, and one or more pharmaceutically acceptable carriers thereof and optionally one or more other therapeutic agent ingredients. The carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. The appropriate formulation depends on the chosen route of administration. Any well-known techniques, carriers and excipients may be used as appropriate and as understood in the art; for example in Remington's pharmaceutical sciences. The pharmaceutical compositions disclosed herein may be manufactured in any manner known in the art, for example, by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping (entropping) or compressing processes.

The agent-polyamine analogs, polyamine biosynthesis inhibitors, polyamine transport inhibitors, or agents that inhibit SAMDC-can also be administered in combination with one or more entities. In one embodiment, the entity is a therapeutic entity, including but not limited to an antiviral or antiretroviral agent, a steroid or other anti-inflammatory agent. In another embodiment, the entity is a pharmaceutically acceptable carrier.

The optimal dose, frequency of administration, and duration of treatment of the agent in a subject can vary from subject to subject, depending on the disease to be treated or the clinical endpoint to be achieved (e.g., inhibition of macrophage infiltration into tissue, or reduction of pain), the condition of the subject, the age, weight, response to treatment, and the nature of the therapeutic entity. Determination of the optimal dosage and duration of treatment is within the ability of those skilled in the art. The optimal dosage and duration of treatment can be optimally determined by monitoring the subject's response during the course of treatment. In some instances, higher doses of administration may allow for less frequent administration, and lower doses may require more frequent administration in order to achieve a clinically significant improvement in the subject's condition. The agents of the invention may be administered as a single dose or in multiple doses.

Generally, a therapeutically effective dose of an agent according to the methods of the invention will be from about 10 to about 1100mg/m²One or more doses of (a). Lower dosage regimens include 10-200, 10-100, 10-50 and 20-200mg/m²The dosage of (a). The higher dosage schemes include 200-400, 250-500, 400-600, 500-800, 600-1000 and 800-1100mg/m². In one embodiment, the dosage regimen is at 200-400mg/m²Within the range. In another embodiment, the dosage regimen is at 250-500mg/m²Within the range. In yet another embodiment, the dosage regimen is at 600-1000mg/m²Within the range. In some embodiments, the agent is administered daily, weekly, every other week, or monthly. In one embodiment, at 200-²Dosage regimen within the range of weekly administrationIt is used once. In another embodiment, at 250-²Dosage regimens within the range are administered once every other week.

The dose may be constant throughout the treatment period, or it may increase or decrease during the course of treatment. In one embodiment, the agent is administered once per week and from 200mg/m²Started and increased to 300mg/m in the second and third weeks, respectively²And 400mg/m². In another embodiment, the agent is administered once every other week and remains constant throughout the duration of treatment at 250mg/m²And (4) application. The dose of agent may be administered for at least one week, at least two weeks, at least three weeks, at least four weeks, at least 6 weeks, or even at least 8 weeks. Adjusting the dosage of the agent within these ranges for a particular subject is within the ability of the general clinician.

The agent may be administered via any conventional route commonly used for the administration of pharmaceutical agents, including, but not limited to, oral, parenteral (including subcutaneous, intradermal, intramuscular, intravenous, intraarticular, and intramedullary), intraperitoneal, transmucosal (including nasal), transdermal, rectal, and topical (including dermal, buccal, sublingual, and intraocular) routes. Intravenous delivery may be via bolus injection or via infusion; infusion can be accomplished in a period of time from less than 1 minute to several hours to continuous. In certain embodiments, the course of treatment will involve administration by a combination of routes.

For example, the agent may be administered via a combination of intravenous and oral routes for the treatment of pain or another condition. In one embodiment, the "loading" dose may be administered intravenously to bring the drug concentration to the desired therapeutic level, followed by administration of one or more maintenance doses via the oral route to keep it there. In yet another embodiment, a combination of oral and intravenous delivery may be used to reduce pain in a surgical patient. The agents may be delivered preoperatively, perioperatively, and postoperatively by a combination of intravenous and oral routes. In one embodiment, the patient may be administered the drug orally or may self-administer the drug prior to surgery, via IV infusion during and immediately after surgery, and may thereafter be administered orally or may self-administer the drug after surgery. In another embodiment, the patient may be administered the drug intravenously prior to surgery, via IV infusion during and immediately after surgery, and may thereafter be administered orally or may self-administer the drug after surgery.

The agent may be administered as a pharmaceutical composition in a variety of forms including, but not limited to, liquids, powders, suspensions, tablets, pills, capsules, sprays, and aerosols. The pharmaceutical composition may include various pharmaceutically acceptable additives including, but not limited to, carriers, excipients, binders, stabilizers, antimicrobials, antioxidants, diluents, and/or carriers (supports). Examples of suitable excipients and carriers are described, for example, in "Remington's pharmaceutical sciences," Mack pub. In some embodiments, the agent may be administered via IV infusion in an aqueous sugar solution. The agent may also be combined with another substance that facilitates delivery of the agent. For example, the agent may be associated with a liposome. Liposomes can in turn be conjugated to a target substance such as an IgGFc receptor.

Formulations of the compounds disclosed herein suitable for oral administration may exist as follows: discrete units such as capsules, cachets or tablets, each containing a predetermined amount of active ingredient; powder or granules; solutions or suspensions in aqueous or non-aqueous liquids; or an oil-in-water liquid emulsion or a water-in-oil liquid emulsion. The active ingredient may also be presented as a bolus, electuary or paste.

Pharmaceutical preparations which can be used orally include tablets, push-fit capsules made of gelatin, and soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. Tablets may be prepared by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder, inert diluent or lubricant, surfactant or dispersant. Molded tablets may be prepared by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein. All formulations for oral administration should be in dosages suitable for such administration. Push-fit capsules can contain the active ingredient in admixture with fillers such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. Additionally, stabilizers may be added. Dragee cores are provided with suitable coatings. For this purpose, concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinylpyrrolidone, carbopol gel, polyethylene glycol and/or titanium dioxide, lacquer solutions and suitable organic solvents or solvent mixtures. Dyes or pigments can be added to the tablets or dragee coatings for identifying or characterizing different combinations of active compound doses.

The compounds may be formulated for parenteral administration by injection, for example by bolus injection or continuous infusion. Formulations for injection may be presented in unit dosage form, e.g., in ampoules or in multi-dose containers, with an added preservative. The compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents. The formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampoules and vials, and may be stored in powder form or in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example, saline or sterile pyrogen-free water, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.

Formulations for parenteral administration include aqueous and non-aqueous (oily) sterile injection solutions of the active compound which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; aqueous and non-aqueous sterile suspensions of suspending agents and thickening agents may be included. Suitable lipophilic solvents or vehicles include fatty oils, such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol or dextran. Optionally, the suspension may also contain suitable stabilizers or agents that increase the solubility of the compound to allow for the preparation of highly concentrated solutions.

In addition to the formulations described previously, the compounds may also be formulated as depot preparations. The long acting formulation may be administered by implantation (e.g. subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, the compounds may be formulated with suitable polymeric or hydrophobic materials (e.g., as an emulsion in an acceptable oil) or ion exchange resins or as sparingly soluble derivatives, e.g., as a sparingly soluble salt.

For buccal or sublingual administration, the compositions may take the form of tablets, lozenges, pastilles or gels formulated in a conventional manner. The composition may comprise the active ingredient in a flavouring base such as sucrose and acacia or tragacanth.

The compounds may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter, polyethylene glycols or other glycerides.

Certain compounds disclosed herein can be administered locally, that is, by non-systemic administration. This includes applying the compounds disclosed herein externally to the epidermis or buccal space and instilling the compounds into the ear, eye and nose so that the compounds do not significantly enter the blood stream. In contrast, systemic administration refers to oral, intravenous, intraperitoneal, and intramuscular administration.

Formulations suitable for topical administration include liquid or semi-liquid formulations suitable for penetration through the skin to the site of inflammation, such as gels, liniments, lotions, creams, ointments or pastes, and drops suitable for application to the eye, ear or nose. Active ingredients for topical application may constitute, for example, 0.001 to 10% w/w (by weight) of the formulation. In certain embodiments, the active ingredient may constitute as much as 10% w/w. In other embodiments, it may constitute less than 5% w/w. In certain embodiments, the active ingredient may constitute from 2% w/w to 5% w/w. In other embodiments, it may constitute from 0.1% to 1% w/w of the formulation.

For administration by inhalation, the compound may be conveniently delivered by an insufflator, spray pressurizing pack, or other convenient means of delivering an aerosol propellant. The pressurized pack may contain a suitable propellant, such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered quantity. Alternatively, for administration by inhalation or insufflation, the compounds according to the invention may take the form of a dry powder composition, for example a powder mix of the compound with a suitable powder base such as lactose or starch. The powder compositions may be presented in unit dosage forms, such as capsules, cartridges, gelatin or blister packs, from which the powder may be administered by means of an inhaler or insufflator.

Exemplary unit dose formulations are those containing an effective dose, or an appropriate number of parts of active ingredient, as described below.

Fillers useful in the compositions herein include all those now known and in use, as well as those to be developed in the future. Examples of fillers or diluents include, but are not limited to, lactose, mannitol, xylitol, glucose, sucrose, sorbitol, compressible sugar, microcrystalline cellulose (MCC), powdered cellulose, corn starch, pregelatinized starch, dextrates, dextran, dextrin, glucose, maltodextrin, calcium carbonate, dibasic calcium phosphate, tribasic calcium phosphate, calcium sulfate, magnesium carbonate, magnesium oxide, poloxamers such as polyethylene oxide and hydroxypropyl methylcellulose. The filler may have complex solvent molecules, as in the case where the lactose used is lactose monohydrate. The filler may also be a special fillerFor use, e.g. in fillers(available from JRSPharma).Is a proprietary, optionally high density silicified microcrystalline cellulose consisting of 98% microcrystalline cellulose and 2% colloidal silicon dioxide. Silicidation of microcrystalline cellulose is achieved by patented methods, resulting in an intimate bond between the colloidal silicon dioxide and the microcrystalline cellulose. ProSolv appears in different grades based on particle size and is a white or almost white, fine or granular powder, practically insoluble in water, acetone, ethanol, toluene and dilute acids and in 50g/l sodium hydroxide solution.

Disintegrants for use in the compositions herein include all those now known and used as well as those that will be developed in the future. Examples of disintegrants include, but are not limited to, sodium starch glycolate, sodium carboxymethylcellulose, calcium carboxymethylcellulose, croscarmellose sodium, povidone, crospovidone (polyvinylpolypyrrolidone), methylcellulose, microcrystalline cellulose, powdered cellulose, low substituted hydroxypropyl cellulose, starch, pregelatinized starch, and sodium alginate.

Lubricants useful in the compositions herein include all those now known and in use, as well as those to be developed in the future. Examples of lubricants include, but are not limited to, calcium stearate, glyceryl monostearate, glyceryl palmitostearate, hydrogenated vegetable oil, light mineral oil, magnesium stearate, mineral oil, polyethylene glycol, sodium benzoate, sodium lauryl sulfate, sodium stearyl fumarate, stearic acid, talc, and zinc stearate.

Glidants used in the compositions herein include all those now known and in use and those to be developed in the future. Examples of glidants include, but are not limited to, silicon dioxide (SiO)₂) Talc corn starch and poloxamers. Poloxamer (orAvailable from BASF Corporation) is an a-B-a block copolymer in which block a is a hydrophilic polyethylene glycol homopolymer and block B is a hydrophobic polypropylene glycol homopolymer.

Tablet binders useful in the compositions herein include all those now known and in use and those that will be developed in the future. Examples of tablet binders include, but are not limited to, acacia, alginic acid, carbomer, sodium carboxymethylcellulose, dextrin, ethylcellulose, gelatin, guar gum, hydrogenated vegetable oil, hydroxyethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, copovidone, methylcellulose, liquid glucose, maltodextrin, polymethacrylates, povidone, pregelatinized starch, sodium alginate, starch, sucrose, tragacanth, and zein.

Examples of surfactants include, but are not limited to, fatty acids and alkyl sulfonates; commercial surfactants, e.g. benzethonium chloride: (1622, available from Lonza, inc., fairlaw, n.j.); DOCUSTATE(available from Mallinckrodt spec.chem., st.louis, MO); polyoxyethylene sorbitan acid ester (C)Available from ICI Americas inc, Wilmington, DE;p-20, available from Lipochem Inc., Patterson NJ;POE-0 available from Abitec Corp, Janesville, Wis.), polyoxyethylene (20) sorbitan monooleate (TWEEN)Available from ICI Americas inc, Wilmington, DE); and natural surfactants such as sodium taurocholate, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, lecithin, and other phospholipids and mono-and diglycerides. The substance may advantageously be used to increase the rate of dissolution by promoting wetting, thereby increasing the maximum dissolved concentration, and also to inhibit crystallization or precipitation of the drug by interacting with the dissolved drug via a mechanism such as complexation, forming inclusion complexes, forming micelles, or adsorbing to the surface of the solid drug.

Pharmaceutical complexing and solubilizing agents for use in the compositions herein include all those now known and in use and those which will be developed in the future. Examples of pharmaceutical complexing or solubilizing agents include, but are not limited to, polyethylene glycol, caffeine, xanthene, gentisic acid, and cyclodextrin.

It may also be beneficial to add a pH adjusting agent, such as an acid, base, or buffer, to retard or increase the rate of dissolution of the composition, or alternatively to help improve the chemical stability of the composition. Suitable pH adjusting agents for use in the compositions herein include all those now known and in use and those to be developed in the future.

It will be appreciated that in addition to the ingredients particularly mentioned above, the formulations provided herein may contain other agents conventional in the art having regard to the type of formulation in question. The appropriate formulation depends on the chosen route of administration. Any well-known techniques, carriers and excipients may be used as appropriate and as understood in the art; such as in Remington, described above. The pharmaceutical compositions may be manufactured in a manner that is itself known, e.g., by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping or compressing processes.

The compounds may be administered orally or via injection at a dose of 0.1 to 500mg/kg per day. The dosage for adults will generally range from 5mg to 2 g/day. Tablets or other presentation forms provided in discrete units may conveniently contain an amount of one or more compounds which is effective at that dose or in multiple doses thereof, for example a unit containing from 5mg to 500mg, typically from about 10mg to 200 mg.

The precise amount of the compound administered to the subject will be the responsibility of the attending physician. The specific dosage level for any particular subject will depend upon a variety of factors including: the activity of the particular compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration, rate of excretion, drug combination, the precise condition being treated, and the severity of the indication or condition being treated. Likewise, the route of administration may vary depending on the condition and its severity. The frequency of administration can also be selected or adjusted based on factors including those above, as well as the formulation of the compound being delivered. Administration may occur, for example: once daily, twice daily, three or four times daily, every other day, weekly, biweekly or monthly; or in a cycle comprising a sustained administration period followed by a non-administration period; or as desired.

In certain instances, it may be appropriate to administer at least one compound described herein (or a pharmaceutically acceptable salt, ester, or prodrug thereof) in combination with another therapeutic agent. By way of example only, if one side effect experienced by a subject upon receiving a compound herein is hypertension, then administration of an antihypertensive agent in combination with the initial therapeutic agent may be appropriate. Alternatively, by way of example only, the therapeutic efficacy of one of the compounds described herein may be enhanced by administration of an adjuvant (i.e., an adjuvant alone may have only minimal therapeutic benefit, but when combined with another therapeutic agent, the overall therapeutic benefit to the subject is enhanced). Or, by way of example only, the benefit experienced by the subject may be increased by administering one compound described herein and another therapeutic agent (which also includes a therapeutic regimen) that also has therapeutic benefit. By way of example only, in the treatment of a neuropathy involving administration of one of the compounds described herein, increased therapeutic benefit may be achieved by also providing the subject with another therapeutic agent for the neuropathy. In any case, regardless of the disease, disorder, or condition being treated, the overall benefit experienced by the subject may be merely an additive of the two therapeutic agents or the subject may experience a synergistic benefit.

In certain embodiments, the other therapeutic agent is an agent for treating multiple sclerosis, such as interferon β -1a, interferon β -1b, glatiramer acetate, mitoxantrone, natalizumab, fingolimod, dimethyl fumarate (tecfidera), and teriflunomide, as well as other interferons and immunomodulatory, immunosuppressive, or anti-inflammatory drugs.

In other embodiments, the other therapeutic agent is a TNF inhibitor. The TNF inhibitor may be: monoclonal antibodies, such as, for example, infliximab (Remicade), adalimumab (Humira), certolizumab (Cimzia), or grilimumab (Simponi); circulating receptor fusion proteins, such as etanercept (Enbrel); or small molecules such as pentoxifylline or bupropion (Zyban, Wellbutrin).

In other embodiments, the other therapeutic agent is a disease-modifying antirheumatic drug (DMARD). Examples of DMARDs include azathioprine, cyclosporine (cyclosporin a), D-penicillamine, gold salts, hydroxychloroquine, leflunomide, Methotrexate (MTX), minocycline, sulfasalazine (SSZ), and cyclophosphamide.

In other embodiments, the other therapeutic agent is methotrexate.

Other agents used in combination include interleukin 1(IL-1) blockers such as anakinra (Kineret), T cell costimulatory blockers such as aberrapu (Orencia), interleukin 6(IL-6) blockers such as tollizumab (anti-IL-6 receptor antibody; RoActemra, Actemra), monoclonal antibodies against B cells such as rituximab (Rituxan), and other biologicals (e.g., Ocrelizumab, Aframumab, Grimumab, and Setuzumab).

In other embodiments, the other therapeutic agent is a glucocorticoid or a nonsteroidal anti-inflammatory drug (NSAID). NSAIDs include propionic acid derivatives such as ibuprofen, naproxen, fenoprofen, ketoprofen, flurbiprofen, and oxaprozin; acetic acid derivatives such as indomethacin, sulindac, etodolac, and diclofenac; enolic acid (oxicam) derivatives such as piroxicam and meloxicam; fenamic acid derivatives such as mefenamic acid and meclofenamic acid; selective COX-2 inhibitors (Coxibs) such as celecoxib (celecoxib), rofecoxib, valdecoxib, parecoxib, lumiracoxib, and etoricoxib.

In any case, the multiple therapeutic agents (at least one of which is a compound disclosed herein) can be administered in any order or even simultaneously. If simultaneous, multiple therapeutic agents may be provided in a single unified form or in multiple forms (by way of example only, as a single pill or as two separate pills). One of the therapeutic agents may be given in multiple doses, or both may be given in multiple doses. The time between doses of the multiple therapeutic agents may be of any duration ranging from a few minutes to four weeks, if not simultaneously.

Thus, in another aspect, certain embodiments provide a method for treating a condition in a human or animal subject in need of such treatment, comprising administering to the subject a compound disclosed herein in an amount effective to reduce or prevent the condition in the subject, optionally in combination with at least one additional agent for treating the condition known in the art. Specific diseases to be treated with the compounds, compositions, and methods disclosed herein, alone or in combination, include, but are not limited to: pain; neuropathy; inflammation and related disorders; arthritis; a metabolic inflammatory disorder; respiratory disorders; autoimmune disorders; neurological disorders; and proliferative disorders including cancer and non-cancerous diseases.

The compounds disclosed herein are useful for treating patients suffering from pain, including neuropathic and/or neuropathic pain, and inflammatory pain. Pain indications include, but are not limited to, the treatment or prevention of surgical or post-operative pain including amputation, post-cardiac surgery, dental pain/extraction of teeth for various surgical procedures, pain caused by cancer, muscle pain, breast pain, pain caused by skin injury, lower back pain, headaches of various etiologies including migraine, menstrual cramps, and the like. The compounds are also useful in the treatment of pain related conditions such as tactile allodynia and hyperalgesia. Pain may be somatotropic (nociceptive or neuropathic), acute and/or chronic.

Peripheral neuropathies that can be treated with the compounds disclosed herein include mononeuropathy, mononeuropathies, and polyneuropathy, including axonal and demyelinating neuropathies. Including sensory and motor neuropathy. Neuropathy or neuropathic pain can be associated with many peripheral neuropathies of different etiologies, including but not limited to:

trauma-induced neuropathy, including neuropathy caused by physical injury (e.g., blunt trauma, abrasion, or burn) or disease state, physical damage to the brain, physical damage to the spinal cord, or stroke associated with brain injury; neurological disorders associated with neurodegeneration; and post-operative neuropathy and neuropathic pain (e.g., from tumor resection, mastectomy, etc.)

Infectious and viral neuropathies, including neuropathies induced by leprosy, lyme disease, herpes virus (and more specifically by herpes zoster virus, which can cause neuralgia on herpes zoster), human immunodeficiency virus (HIV, which can cause HIV neuropathy), or papilloma virus, or any other pathogen;

toxin-induced neuropathy (including but not limited to neuropathy induced by alcoholism, vitamin B6 intoxication, hexacarbon intoxication, amiodarone, chloramphenicol, disulfiram, isoniazid, gold, lithium, metronidazole, misonidazole, nitrofurantoin);

drug-induced neuropathy, including therapeutic agent-drug-induced neuropathy, particularly a) chemotherapy-induced neuropathy caused by anticancer agents such as taxol, docetaxel, cisplatin, nocodazole, vincristine, vindesine, and vinblastine, and b) antiviral neuropathy caused by antiviral agents such as ddI, DDC, d4T, foscarnitin, dapsone, metronidazole, and isoniazid;

neuropathy induced by vitamin deficiency, including neuropathy caused by vitamin B12 deficiency, vitamin B6 deficiency, and vitamin E deficiency;

hereditary neuropathy (including but not limited to ataxia, familial amyloid polyneuropathy, high density lipoprotein deficiency disease, fabry disease);

diabetic neuropathy and neuropathy caused by metabolic disorders, such as renal insufficiency and hypothyroidism;

neuropathy secondary to tumor infiltration,

autoimmune neuropathies, including those caused by Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, monoclonal gammopathy and polyneuropathy of unknown significance, and multiple sclerosis;

other neuropathic and neuropathic pain syndromes, including inflammation-induced nerve damage, neurodegeneration, posttraumatic neuralgia, central neuropathic pain syndromes such as phantom limb pain, complex regional pain syndromes (including but not limited to reflex sympathetic dystrophy, causalgia), pain associated with neoplasia, vasculitis/vasculopathy and sciatica; and

idiopathic neuropathy.

In certain embodiments, neuropathic pain may alternatively manifest as allodynia, hyperalgesic pain, thermal hyperalgesia, or phantom pain. In another embodiment, the neuropathy may instead result in loss of pain sensitivity. Additional sub-categories of neuropathic Pain are discussed in Dworkin, Clin J Pain (2002) Vol.18 (6), pages 343-9.

The compounds disclosed herein are also useful for treating or preventing opioid tolerance in patients in need of extended opioid analgesics, and benzodiazepine tolerance in patients taking benzodiazepines, as well as other addictive behaviors, such as nicotine addiction, alcoholism, and eating disorders. In addition, the compounds disclosed herein are useful for treating or preventing drug withdrawal symptoms, for example, treating or preventing withdrawal symptoms of opiate, alcohol, or tobacco addiction.

The compounds disclosed herein may also be used to treat or prevent respiratory diseases or conditions, including: asthma conditions including allergen-induced asthma, exercise-induced asthma, pollution-induced asthma, cold-induced asthma, and virus-induced asthma; chronic obstructive pulmonary diseases including chronic bronchitis with normal airflow, chronic bronchitis with airway obstruction (chronic obstructive bronchitis), emphysema, asthmatic bronchitis, and bullous disease; and other pulmonary diseases involving inflammation, including bronchiectasis, cystic fibrosis, hypersensitivity pneumonitis, farmer's lung, acute respiratory distress syndrome, pneumonia, aspiration or inhalation injury, pulmonary fatty embolism, acidotic inflammation of the lung, acute pulmonary edema, acute mountain sickness, acute pulmonary hypertension, persistent pulmonary hypertension in newborn, perinatal inhalation syndrome, hyaline membrane disease, acute pulmonary thromboembolism, heparin-protamine reaction, sepsis, asthma persistence, hypoxia, hyperoxic lung injury, and injury induced by inhalation of certain harmful mediators, including smoking, leading to complications thereof, such as lung cancer.

The compounds disclosed herein may also be used to treat or prevent inflammation and inflammatory conditions. Inflammatory conditions include, but are not limited to: arthritis, including subtypes and related conditions, such as rheumatoid arthritis, spondyloarthropathies, gouty arthritis, osteoarthritis, systemic lupus erythematosus, juvenile arthritis, acute rheumatoid arthritis, enteropathic arthritis, neuropathic arthritis, psoriatic arthritis, and suppurative arthritis; osteoporosis, tendonitis, bursitis and other related bone and joint disorders; gastrointestinal conditions such as reflux esophagitis, diarrhea, inflammatory bowel disease, crohn's disease, gastritis, irritable bowel disease, ulcerative colitis, acute and chronic pancreatic inflammation; pneumonia, such as that associated with viral infections and cystic fibrosis; skin-related conditions such as psoriasis, eczema, burns, sunburn, dermatitis (such as contact dermatitis, allergic dermatitis, and allergic dermatitis), and urticaria; pancreatitis, hepatitis, pruritus and leukoderma. In addition, the compounds of the present invention are also suitable for use in organ transplant patients, either alone or in combination with conventional immunomodulators.

The compounds disclosed herein may also be used to treat or prevent autoimmune disorders. Autoimmune disorders include crohn's disease, ulcerative colitis, dermatitis, dermatomyositis, type 1 diabetes, goodpasture's syndrome, graves ' disease, guillain-barre syndrome (GBS), autoimmune encephalomyelitis, hashimoto's disease, idiopathic thrombocytopenic purpura, lupus erythematosus, mixed connective tissue disease, Multiple Sclerosis (MS), myasthenia gravis, narcolepsy, pemphigus vulgaris, pernicious anemia, psoriasis, psoriatic arthritis, polymyositis, primary biliary cirrhosis, rheumatoid arthritis, sjogren's syndrome, scleroderma, temporal arteritis (also known as "giant cell arteritis"), vasculitis, and necrotizing granulomatosis. The compounds disclosed herein can modulate TH-17 (interleukin 17 producing T-helper cells) cells or IL-17 levels.

Autoimmune disorders can involve the nervous system. Examples of autoimmune disorders affecting the nervous system include polymyalgia, myasthenia gravis, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, transverse myelitis, Barlow's concentric sclerosis, pernicious anemia, acute disseminated encephalomyelitis (ADME), Amyotrophic Lateral Sclerosis (ALS), autoimmune peripheral neuropathy, lupus erythematosus, psoriatic arthritis, rheumatoid arthritis, osteoarthritis, and rheumatic fever.

The compounds disclosed herein may also be used to treat or prevent demyelinating diseases, including Multiple Sclerosis (MS), optic neuritis, idiopathic inflammatory demyelinating diseases, guillain-barre syndrome (and subtypes), chronic inflammatory demyelinating polyneuropathy, transverse myelitis, baroconcentric sclerosis, pernicious anemia, pontine myelinolysis, tabes, neuromyelitis optica (NMO), Progressive Multifocal Leukoencephalopathy (PML), anti-MAG (myelin associated glycoprotein) neuropathy, hereditary motor and sensory neuropathy (peroneal atrophy disease), tendonoxanthomatosis, and leukodystrophy, including adrenoleukodystrophy, adrenomyeloneuropathy, metachromatic leukodystrophy, globuloid cellular leukodystrophy (krabbe disease), canavan disease, evaporative leukopathy, alexander disease, amateur disease, multiple sclerosis, guillain-barre syndrome, central myelina, progressive multifocal leukodystrophy, progressive leukodystrophy, and multiple sclerosis, Refsum disease and pelizaeus-merzbach disease.

The compounds disclosed herein are also useful for treating or preventing certain diseases and disorders of the nervous system. Central nervous system disorders in which nitric oxide inhibition is useful include cortical dementias including alzheimer's disease, central nervous system injury resulting from stroke, ischemia including cerebral ischemia (focal ischemia, thrombotic stroke and general ischemia, e.g. secondary to cardiac arrest), and trauma. Neurodegenerative disorders in which nitric oxide inhibition is useful include neurodegeneration or nerve necrosis in disorders such as hypoxia, hypoglycemia, epilepsy, and in the following cases: central Nervous System (CNS) trauma (e.g., spinal cord and craniocerebral injuries), hyperbaric oxygen-induced convulsions and toxicity, dementias such as alzheimer's disease and AIDS-related dementia, cachexia, west-arm-and-hall chorea, huntington's chorea, parkinson's disease, Amyotrophic Lateral Sclerosis (ALS), korsakoff's psychosis, cognitive disorders related to cerebrovascular disorders, hypersensitivity, sleep disorders, schizophrenia, depression or other symptoms associated with premenstrual syndrome (PMS), and anxiety.

The compounds disclosed herein are also useful for treating or preventing metabolic disorders typically associated with amplified inflammatory signaling, such as insulin resistance, diabetes (type I or type II), metabolic syndrome, non-alcoholic steatohepatitis, atherosclerosis, cardiovascular disease, congestive heart failure, myocarditis, atherosclerosis, and aortic aneurysm.

The compounds disclosed herein are also useful for treating or preventing organ and tissue damage associated with severe burns, sepsis, trauma and hemorrhage or resuscitation-induced hypotension, but also for treating or preventing diseases: such as vascular disease, migraine, periarteritis nodosa, thyroiditis, aplastic anemia, hodgkin's disease, scleroderma, rheumatic fever, type I diabetes, neuromuscular junction diseases including myasthenia gravis, leukoencephalopathy including multiple sclerosis, sarcoidosis, nephritis, nephrotic syndrome, behcet's syndrome, polymyositis, gingivitis, periodontal disease, swelling that occurs after injury, ischemia including myocardial ischemia, cardiovascular ischemia, and ischemia secondary to cardiac arrest, and the like.

The compounds disclosed herein may also be used for the treatment or prevention of (highly) proliferative diseases, in particular cancer, alone or in standard-of-care combinations, in particular those agents which target tumor growth by relocating aberrant apoptotic mechanisms in malignant cells. Hematologic and non-hematologic malignancies that can be treated or prevented include, but are not limited to, multiple myeloma, acute and chronic leukemias, including Acute Lymphocytic Leukemia (ALL), Chronic Lymphocytic Leukemia (CLL) and chronic myelogenous leukemia (CLL), lymphomas, including hodgkin's lymphoma and non-hodgkin's lymphoma (low, intermediate and high), and solid and malignant tumors of the brain, head and neck, breast, lung, reproductive tract, upper gastrointestinal tract, pancreas, liver, kidney, bladder, prostate and colorectal. The compounds and methods of the invention are also useful for treating fibrosis, such as that which occurs with radiation therapy. The compounds and methods of the invention are useful for treating subjects having adenomatous polyps, including subjects having Familial Adenomatous Polyposis (FAP). In addition, the compounds and methods of the invention are useful for preventing polyp formation in patients at risk for FAP. Non-cancerous proliferative disorders additionally include psoriasis, eczema and dermatitis.

The compounds disclosed herein are also useful for treating or preventing polycystic kidney disease and other diseases of renal dysfunction.

The compounds disclosed herein are also useful for treating or preventing ophthalmic diseases such as glaucoma, retinal ganglionic degeneration, ocular ischemia, corneal neovascularization, optic neuritis, retinitis, retinopathies such as glaucomatous retinopathy and/or diabetic retinopathy, uveitis, ocular photophobia, dry eye, sjogren's syndrome, seasonal and chronic allergic conjunctivitis, and inflammation and pain associated with chronic ocular disorders and acute damage to ocular tissue. The compounds are also useful in the treatment of post-operative inflammation or pain from ophthalmic surgery, such as cataract surgery and refractive surgery.

The compounds of the invention may also be used in co-therapy, partially or completely replacing other conventional anti-inflammatory therapies, such as steroids, NSAIDs, COX-2 selective inhibitors, 5-lipoxygenase inhibitors, LTB₄Antagonists and LTA₄Together with a hydrolase inhibitor. The compounds of the present invention may also be useful in the prevention of tissue damage when combined therapeutically with an antibacterial or antiviral agent.

Exemplary embodiments of the present process are provided in the following examples. The following examples are set forth to illustrate the methods of the present invention and to assist those of ordinary skill in the art in using the methods, and are not to be construed as limiting the scope of the invention.

MGBG oral Activity assay

The following standard abbreviations are used to indicate relevant pharmacokinetic parameters.

AUC to the area under the curve of last measurable concentration plus the last measurable concentration (at t)_{Finally, the}C of_{Finally, the}) AUC to concentration extrapolation within an infinite range: AUC_INFobs＝AUC_{0-t last}+C_{Finally, the}λ z (where λ z is the first order rate constant associated with the terminal (log-linear) portion of the curve)

AUC_0-12Area under the curve between dosing time and 12h time point

AUC_0-24Area under the curve between dosing time and 24h time point

Fraction available for F (bioavailability):

F＝[AUC_{is administered orally}]Dosage of_iv/[AUC_iv]Dosage of_{Is administered orally}

Cl_obsObserved clearance

V_SSobsSteady state volume of distribution

V_dVolume of distribution (often used orally)

Cl/F_obsApparent total body clearance as a function of bioavailability

t_1/2Ultimate half-life (HL)_λz)

C_{Maximum of}Maximum observed concentration

T_{Maximum of}C_{Maximum of}Time of appearance

Single dose of rhesus monkey

Two groups of male rhesus monkeys (three per group) were fasted overnight before being administered the test item MGBG either as a single bolus intravenous dose of 1mg/kg (group 1) or as a single oral gavage dose of 10mg/kg (group 2). Dose formulation analysis it was tested for groups 1 and 2, respectively, whether the administered dose solution was within 14% of the target concentration of 1 and 10 mg/kg.

Blood samples from femoral vein/artery (approximately 1.0mL) were collected into tubes containing lithium heparin, and plasma MGBG concentrations were measured in all intravenously dosed animals before dosing and after dosing at approximately T ═ 0.083(5min), 0.25(15min), 0.5(30min), 1, 2, 4, 8, and 24 hours. Blood samples for plasma MGBG concentration measurements were collected from all orally dosed animals before dosing, at approximately 1, 2, 4, 8, 12, 24, and 36 hours after dosing. Food intake was also stopped during the first four hours of blood sample collection.

After sample collection was completed at each interval, the samples were centrifuged under refrigerated conditions. The resulting plasma was separated and stored frozen at about-70 ℃ until analysis.

PK analysis was performed on a single plasma concentration-time curve of MGBG using the WinNonlin non-compartmental method (linear trapezoidal rule for AUC calculation). The nominal dose value and the sampling time are used for the calculation. For analytical purposes, all MGBG plasma concentration measurements reported as BQL (< 2.51ng/mL) were set equal to zero. Following IV and PO administration of MGBG, plasma PK profile parameters were calculated using WinNonlin default selection criteria for λ Z selection.

At the time point after IV and PO administration of MGBG, evidence of systemic plasma MGBG exposure was observed in all collected plasma. Hemolysis was noted in one animal in group 1 at a single time point, which can have a negative impact on MGBG plasma concentration analysis in this animal. Thus, model-dependent two-compartment analysis is used to calculate bioavailability.

Single dose in dogs

Two groups of male beagle dogs weighing 9.0-10.7kg and aged 8-30 months (three dogs each) were fasted overnight before the test item MGBG was administered either as a single bolus intravenous dose of 1mg/kg (group 1) or as a single oral gavage dose of 10mg/kg (group 2). Dose formulation analysis it was tested for groups 1 and 2, respectively, whether the administered dose solution was within 17% of the target concentration of 1 and 10 mg/kg.

Blood samples (about 2.0mL) were collected in order to measure plasma MGBG concentrations in all intravenously dosed animals before dosing and at about T ═ 0.083(5min), 0.25(15min), 0.5(30min), 1, 2, 4, 8, and 24 hours after dosing. Similar procedures were used for orally administered animals, except that T ═ 1, 2, 4, 8, 12, 24, and 36 hours after administration were collected. After sample collection was completed at each interval, the samples were centrifuged under refrigerated conditions. The resulting plasma was separated and stored frozen at about-70 ℃ until analysis.

Analysis was performed by LC/MS and the plasma PK profile parameters were calculated for selection of λ Z using the last five plasma concentrations of IV (1-24h) and PO (4-36h) administration. These results should be interpreted carefully due to variability between animals and limited end-stage data.

No clinically abnormal results were found after intravenous or oral administration. Systemic exposure was observed at all time points.

Single dose in rats

18 Sprague Dawley male rats (Charles river) weighing 217-263g and aged 8-9 weeks were administered the test article MGBG, either as a single bolus intravenous dose of 1mg/kg (group 1) or as a single oral gavage dose of 10 mg/kg. One group of three animals was passed through CO after final blood collection at each point of 2, 4, 12, 24, 36 and 48 hours post-dose₂Sacrificed by inhalation anesthesia. Dose formulation analysis checks whether the dose solution administered is within a target concentration of 17% of 10 mg/kg.

Analysis was performed by LC/MS/MS. Pharmacokinetic analysis was performed on the mean MGBG plasma concentration versus time data curve using a non-compartmental method (linear trapezoidal rule for AUC calculation). The WinNonlin sparse sampling tool is used for PK calculations. All samples reported as BLQ (below the limit of quantitation, 2.50ng/mL in plasma) were converted to 0.00ng/mL for analysis. Dose formulation analysis showed that the formulation was within 15% of the target dose concentration of 10 mg/kg.

No abnormal clinical results were found after administration. A single PO administration of 10mg/kg MGBG produced evidence of measurable MGBG levels in plasma over a 12 hour time point; except that point, some samples began measuring BLQ.

In addition, in rats dosed with a single oral administration of 10mg/kg of MGBG as above, three rats were sacrificed immediately after plasma sampling (i.e., 2, 4, 12, 24, 36 and 48 hours after dosing) for each group, and spleen and liver tissues were collected and flasks were frozen. As shown in FIG. 11, a single oral administration of 10mg/kg of MGBG resulted in a greater overall exposure (as by C) of MGBG to liver (120-fold and 160-fold, respectively) and spleen (4.0-fold and 9.3-fold, respectively) tissues versus plasma_{Maximum of}And AUC_{All are}Assessed). This is selective to MGBGThe uptake mechanism is consistent. Without wishing to be bound by theory, other SAMDC inhibitors that are selectively taken up by cells may be suitable for use in the methods and compositions disclosed herein like MGBG.

Single dose in mice

Twenty-four male DBA/1 mice weighing 19.5-24.7g and aged 7-9 weeks were administered the test article MGBG via the lateral tail vein in a single bolus intravenous dose of 1mg/kg (group 1, n-12) or a single oral gavage dose of 10mg/kg (group 2, n-12). Each dose group consisted of 4 cohorts of 3 animals each. Group 1 samples were taken 5, 15 and 30 minutes after dosing; and samples were taken 1, 2, 4, 8 and 24 hours after dosing. Group 2 was sampled at 1, 2, 4, 8, 12, 24 and 36 hours after dosing. Starting from the first time point, a new group is sampled at each successive time point until the 1 hour (set 1) or 12 hour (set 2) time point. The sampling sequence in the cohort is repeated for subsequent time points (some cohorts may only draw blood once). The second bleeding for each group was terminal. Via CO after final blood collection₂Animals were sacrificed by inhalation anesthesia.

After sample collection was completed at each interval, the samples were centrifuged under refrigerated conditions. The resulting plasma was separated and stored frozen at about-70 ℃ until analysis. Analysis was performed by LC/MS/MS. Pharmacokinetic analysis was performed on the mean MGBG plasma concentration versus time data curve using a non-compartmental method (linear trapezoidal rule for AUC calculation). The WinNonlin sparse sampling tool is used for PK calculations. Dose formulation analysis showed IV and PO formulations within 15% of the target concentration.

No abnormal clinical results were found after administration. At the time point after IV and PO administration of MGBG, evidence of systemic plasma MGBG exposure was observed in all collected plasma.

The results of the above measurement are shown in tables 2 and 3 below. Values reported are mean values between treatment groups, not including standard deviation.

TABLE 2

TABLE 3

In Table 2 above, the double asterisks indicate that the reported rat AUC is the AUC from time zero to the time of the last plasma concentration measurement_{All are}. Each of these values should account for the fact that the end measurements are subject to different extrapolation methods.

Multidose rat pharmacokinetics and tolerability studies

The objective of this study was to determine the Pharmacokinetic (PK) properties and tolerability of MGBG in rats. In addition, recovery from any toxic effects was assessed after a seven day non-dosing period. Tolerance is evidenced in animals treated with the test article by weight changes and lack of adverse clinical observations similar to those of the control group.

Three male Sprague Dawley (r) (222.7-252.0 g) aged 7-9 weeks and weighed 222.7-9.0 g per group were administered by oral (PO) gavageIGS, Charles River), twice daily at 10, 20 or 30 mg/kg/dose (20, 40 or 60 mg/kg/day), for 7 consecutive days. A seven day washout period follows. Approximately 200 μ L of whole blood was collected from the tail veins of all animals and bled at six (day 1), seven (day 7), or one (day 9 to 15) time points in groups 5, 6, and 7, respectively. Whole blood samples were collected in heparin lithium microtainers and processed into plasma by centrifugation. The plasma was frozen at-70 ℃. Pharmacokinetic analysis was performed on individual animal MGBG plasma concentration versus time data using WinNonlin (linear trapezoidal rule for AUC calculation). The nominal dose value and the sampling time are used for the calculation. At time zero for study day 7The reported MGBG concentration values were used to calculate AUC. On study day 1, no distribution parameters were reported, as the end-stage data were insufficient to characterize these parameters. Plasma PK profile parameters were calculated for plasma concentrations obtained after the second administered dose (T ═ 12-192h) using WinNonlin default selection criteria for the selection of λ Z, half-life, AUC, following PO administration of MGBG on study day 7_INFobsAnd Cl/F_obsThe rate constant of cancellation based on; inter-animal variability was noted.

Plasma samples collected from test article treated animals on days 1 and 7 were subjected to bioanalysis and confirmed systemic exposure to the test article at all time points. Within the dose range evaluated, T_{Maximum of}The values were dose dependent and ranged from 3.33 to 14.0h, and showed that absorption was slightly delayed on study day 7 compared to study day 1. Systemic exposure (e.g. by C) _{Maximum of}And AUC_{All are}Assessed) increases with increasing dose, and the increase in both parameters is slightly less than the dose at each evaluation interval. Similarly, for the 20, 40 and 60mg/kg/day dose groups, respectively, two PO administrations per day of MGBG compared to the AUC at day 1 of the study_{All are}The average values were 3.77-fold, 4.03-fold and 3.68-fold increased. On study day 7, Cl/F was observed_obsAnd evidence of dose-dependent distribution of elimination half-life, due to Cl/F_obsAnd the mean parameter value for elimination of half-life increased and decreased with increasing dose level, respectively.

Differences in some blood parameters (lower reticulocyte counts and percentages) and some serum chemistry parameters (e.g., osmolality and electrolyte changes consistent with slight dehydration) were noted between the control group and the 60mg/kg/day dose group. However, these changes are not considered disadvantageous because they do not meet other signs of frank toxicity (frank toxicity) and the changes in serum chemistry prove to be reversible. No gross or subtopic lesions were observed at the end of the dosing period in animals treated with the test article, and no gross lesions were observed at the end of the recovery period in animals treated with the test article.

Based on the results of this exploratory study, there was no observable level of side effects (NOAEL) for two consecutive 7 days of MGBG administration by PO gavage to male Sprague Dawley rats twice daily (60 mg/kg/day).

TABLE 4

TABLE 5

Abnormal growth rate and predicted human effect

The multi-species allometric growth ratios based on the pharmacokinetic parameters disclosed in tables 2 and 3 were used to calculate the predicted pharmacokinetic parameters of the human according to methods known in the art. See, e.g., Ings RM, "Interspecific calling and complications in drug depletion and toxicokinetics," Xenobiotica, 1990 Nov; 20(11): 1201-31 and Khor, SP et al, "Dihydropyrimidine dehydrogenisation and 5-fluoroouracil pharmacokinetics: allometric scaling of animal data, pharmacokinetics and toxicosylamides of 5-fluoroauracil in humans, "cancer chemither Pharmacol (1997)39 (3): 833-38. The expected values are given in tables 6 and 7 below.

TABLE 6

TABLE 7

In the murine carrageenan-induced paw edema and hyperalgesia model, the highest effective dose of MGBG was 30mg/kg PO BID (60 mg/kg/day total). Based on this dosing regimen in mice, at least two methods can be used to estimate equivalent dosing in humans.

The first method was based on Body Surface Area (BSA) normalization (described in Reagen-Shaw et al (2007) FASEBJ.22, 659-661), as the authors noted that BSA correlates well between species for various biological parameters including basal metabolic rate, blood volume, caloric expenditure, plasma protein levels, and renal function. Using this method, a 60mg/kg/day dose in mice would translate to about 4.9 mg/kg/day in humans.

A second method for converting an effective 60mg/kg/day dose in mice to an equivalent dose in humans is more directly based on the rate of allogenic growth. Data from the MGBG pharmacokinetic study consisted of a 10mg/kg oral dose in mice, modeled in a simulated manner to determine the theoretical AUCINF value for a dosing regimen of 30mg/kg PO BID, which was 9050h ng/mL. Second, the predicted human clearance values as determined by the single and multiple species allometric growth ratios are used to estimate the dose likely to produce an exposure in humans (AUCINF) similar to 60mg/kg/day in mice. Using the single species growth rate and a range of predicted human clearance values, the human equivalent dose will range from 1.73 mg/kg/day to 4.51 mg/kg/day. Using the multi-species allometric growth ratio, the predicted human equivalent dose was about 4.2 mg/kg/day.

In the murine carrageenan model, the efficacy of MGBG at lower doses, including 3mg/kg PO BID and 10mg/kg PO BID, was also observed, which was scaled to a human dose of about 0.49 mg/kg/day and about 1.6 mg/kg/day.

The average body weight of a normal male is often assumed to be 70 kg. Thus, the daily dose based on the above predictions can be estimated to be in the range of about 25 mg/day to about 350 mg/day.

The appropriate dosage will of course depend on a number of factors. Patients may weigh much more or less, or be female, elderly or young, requiring lower or higher doses. The patient may exhibit a drug metabolism profile, which may suggest lower or higher doses, such as low expression levels or active metabolic enzymes such as cytochrome P450 (CYP). This low expression or activity level may be due to a number of factors. It is known that polymorphic expression of one or more CYPs (e.g. CYP2C19 and CYP2D6, although polymorphic phenomena have been described for almost all CYPs) results in some populations being "under-represented" compared to the population at large, resulting in a "poorly metabolised" phenotype requiring lower doses. In addition, exposure to infectious agents or xenobiotics can cause inhibition of CYP expression or inhibition of the CYP present. Alternatively, the patient may be debilitating, injured or immunocompromised, all with a lower dose being suggested. Patients may take many other drugs that compete with the metabolic system for treatment, including the CYPs discussed above; this well known compound effect may require lower doses. The dosage is also dependent on the condition and its severity as discussed above. An effective dose for one disease or clinical endpoint is not necessarily the same dose as another severe chronic or otherwise critical case that may require a higher dose. However, chronic cases may also require lower doses to be administered over a longer or even indeterminate period. All of these are discussed by example to illustrate the variability of ideal dosing; it is within the ability of the skilled artisan to select the appropriate range of administration for a disease, population or individual.

In view of these factors, it is clear that human daily doses may be as low as 1 mg/day, and as high as 1 g/day. In certain embodiments, the human dose may be in the following ranges: 10 mg/day to 500 mg/day, 20 mg/day to 400 mg/day, or 25 mg/day to 350 mg/day. In other embodiments, the human dose may be in the following ranges: 120 mg/day to 350 mg/day, 150 mg/day to 350 mg/day, 200 mg/day to 350 mg/day, or 250 mg/day to 350 mg/day. In certain embodiments, the human dose can be any one of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 7075, 80, 85, 90, 95, 100, 110, 120, 125, 130, 140, 150, 160, 170, 175, 180, 190, 200, 210, 220, 225, 230, 240, 250, 260, 270, 275, 280, 290, 300, 310, 320, 325, 330, 240, or 350 mg/day.

In certain embodiments, the human dose may be any of 275, 280, 285, 290, 295, 300, 305, 310, 315, 320, 325, 330, 335, 340, 350, 355, 360, 365, 370, or 375 mg/day. In one embodiment, the dose may be 275 mg/day. In another embodiment, the dose may be 300 mg/day. In another embodiment, the dose may be 305 mg/day. In another embodiment, the dose may be 310 mg/day. In another embodiment, the dose may be 315 mg/day. In another embodiment, the dose may be 320 mg/day. In another embodiment, the dose may be 325 mg/day. In another embodiment, the dose may be 330 mg/day. In another embodiment, the dose may be 335 mg/day. In another embodiment, the dose may be 340 mg/day. In another embodiment, the dose may be 345 mg/day. In another embodiment, the dose may be 350 mg/day.

In certain embodiments, the human dose may be any of 350, 375, 400, 425, 450, 475, 500, 525, 550, or 600 mg/day. In one embodiment, the dose may be 375 mg/day. In another embodiment, the dose may be 400 mg/day. In another embodiment, the dose may be 450 mg/day. In another embodiment, the dose may be 500 mg/day.

In certain embodiments, the human dose may be any of 25, 50, 75, 100, or 125 mg/day. In one embodiment, the dose may be 375 mg/day. In another embodiment, the dose may be 25 mg/day. In another embodiment, the dose may be 50 mg/day. In another embodiment, the dose may be 75 mg/day. In another embodiment, the dose may be 100 mg/day. In another embodiment, the dose may be 125 mg/day.

In vivo carrageenan assay

Carrageenan paw test for edema and hyperalgesia

Subcutaneous injection of carrageenan into the hind foot (paw) of rats or mice induced firm inflammation and pain. The inflammatory response started 1-2 hours after carrageenan injection and continued at least five hours after vaccination. In addition, the inflamed paw of the animal is sensitive to noxious (hyperalgesia) or non-noxious (allodynia) stimuli as compared to the contralateral paw. Compounds can be evaluated for anti-hyperalgesic and anti-inflammatory activity in this model. A general increase in response threshold or time following drug administration is indicative of analgesic efficacy. A general reduction in paw swelling after drug administration suggests anti-inflammatory efficacy. It is possible that some compounds will affect the inflamed paw, but not the response to the lateral paw.

Embodiments of the carrageenan foot edema test are performed using materials, reagents and procedures substantially as described by Winter et al, (proc.soc.exp.biol.med., 111, 544 (1962)). Prophylactic and therapeutic embodiments have been developed and are known in the art. Animals were evaluated for their reactivity to noxious (paw grip, plantar test) or non-noxious (cold plate, von Frey filament) stimuli. In the following protocol, mice were used.

Animals, compounds and administrations. Healthy, young, male Swiss Webster mice were used in the study, where the weight change of the mice did not exceed ± 20% of the mean. Animals were divided into 4 groups of 40 animals each and each group was given MGBG (BID, 12 hours apart, 30mg/kg in 5mL/kg physiological saline), dexamethasone as a positive control (QD, 1mg/kg in 5mL/kg 0.5% methylcellulose) or saline vehicle (BID, 5mL/kg) by oral gavage. The fourth group served as the primary control (no carrageenan, no treatment). Treatment with MGBG was performed three days before carrageenan, 1 hour before carrageenan and 11 hours after carrageenan, respectively. Paw edema the mice were treated by subcutaneous injection of carrageenan (Sigma: lambda carrageenan) in the plantar region of the paw in the right paw in a volume of 50 ul of 1% carrageenan (w/v) in saline. The contralateral paw (left paw) received the same volume (50 μ Ι _) of saline and served as control. Mice will be anesthetized with a small dose of ketamine prior to carrageenan injection.

Edema in the paw. Immediately prior to subplantar administration of carrageenan and 2, 3, 5 and 24 hours after carrageenan, mouse paw volumes were measured using an organ fullness tester (Ugo Basile). The assessment of edema can be expressed as mean increase in paw volume relative to control.

Assessment of paw withdrawal latency. Paw withdrawal latency was determined by placing mice on an analgesia hotplate apparatus maintained at a surface temperature of 51 ℃ prior to subplantar administration of carrageenan and 0.5, 2, 3, 5 and 24 hours post-carrageenan. A 30s rest period was maintained to avoid any thermal damage to the jaws. Immediately after testing, all paws were immersed in ice cold water before returning to the cage. Paw withdrawal latency was calculated as Δ t — right paw-left paw.

Serum, plasma and tissue collection. Serum or plasma (each) was collected from eight mice per group and stored at-70 ℃ until cytokine level determination or MGBG drug level determination on day 0 prior to the first drug dose and at peak disease (5 and 24 hours post carrageenan challenge for serum, day 0 prior to the first drug dose and at the end of the study). For serum collection, whole blood samples were collected in serum separation tubes, processed by centrifugation and frozen at-70 ℃. For drug level determination, whole blood samples were collected in lithium heparin microtainers, processed into plasma by centrifugation and the plasma was frozen at-70 ℃. Additionally, the paws were collected and stored in 10% formalin for histology.

And (4) alternative schemes. In an alternative embodiment of this assay, MGBG was administered PO, BID, at 3, 10 and 30mg/kg (dexamethasone was used as positive control, saline as negative control, and treatment/carrageenan initial group, n ═ 16 per group).

And (6) obtaining the result. MGBG is effective in reducing edema and hyperalgesia in the above assays.

Carrageenan air bag model

Subcutaneous injection of air will induce the formation of a connective tissue cavity lined with cells that are similar and functionally similar to the synovial lining. This method is commonly referred to as a balloon model and is a useful animal model of inflammation that can be generated in a relatively short period of time. The balloon may be formed by injecting a volume of sterile air Subcutaneously (SQ) in the dorsal region of the neck. The model can be used to test the efficacy and potency of compounds such as polyamine analogs and polyamine biosynthesis inhibitors such as MGBG in reducing various cellular and biochemical indicators of inflammation when administered orally (PO) gavage.

Animals, compounds and administrations. Healthy male Lewis rats (Charles river Laboratories, Wilmington, Mass.) weighing between 175-200g were used. MGBG or vehicle (0.5% methylcellulose, 0.025% Tween-80) at doses between 1 and 60mg/kg was administered by oral gavage (once daily at a volume of 10 mL/kg) for 6 days. A correction factor of 1.49 was used to account for MGBG as the dihydrochloride/monohydrate form. Naproxen and dexamethasone were administered at doses of 10 and 1mg/kg, respectively, 1 day prior to carrageenan injection and in the morning of injection.

Air pockets formed in the backs of rats during the last 4 days of MGBG or vehicle administration prior to carrageenan injection. Briefly, rats were anesthetized with isoflurane during which back hair was removed and 20mL of sterile air was injected subcutaneously into the intrascapular area. The balloon was allowed to develop 4 days thereafter, and re-inflated 1 day before carrageenan injection (to maintain balloon volume).

One hour prior to carrageenan injection, animals were given a maintenance dose of drug (MGBG, vehicle, naproxen, or dexamethasone). A1% carrageenan suspension (2 mL; FMC BioPolyer, Philadelphia, Pa.) suspended in saline was injected into the capsule. 3 or 24 hours after carrageenan injection by CO₂Rats were euthanized by asphyxiation and 2.5mL PBS (Sigma Chemical, st. louis, MO) was injected directly into the bursa. The pouch was opened with surgical scissors and the pouch fluid was collected using a pipette. Collecting an aliquot forTotal cell differential counts were measured and other aliquots were used for PGE₂And (4) measuring. In the latter case, aliquots were centrifuged at 1200g for 10 minutes at 4 ℃ and supernatants were collected for PGE via ELISA₂Analysis (Cayman Chemical Company, Ann Arbor, MI).

Samples were collected from the 3 and 24 hour groups at 3 or 24 hours post carrageenan injection as previously described.

This protocol may vary according to methods known in the art. Additional tissues may be collected and/or weighed and additional sectioning, staining, and microscopic examination may be performed.

And (6) obtaining the result. MGBG was effective in this model as shown by changes relative to controls indicating a reduction in inflammation. Fundamentally, MGBG selectively inhibits the inflammatory spike production of PGE2 without affecting the basal levels of this mediator. In contrast, naproxen and dexamethasone inhibited the inflammatory and basal level production of this mediator.

In vivo murine collagen-induced arthritis

Collagen-induced arthritis model for arthritis and rheumatoid arthritis

The collagen-induced arthritis (CIA) model is considered as a suitable model for studying the activity of potential drugs in human arthritis, as many of the immunology and pathology of human arthritis are similar to human Rheumatoid Arthritis (RA), involving localized major histocompatibility, complete class II-restricted T-helper lymphocyte activation and similar tissue damage. See, e.g., Rosloniec EF et al, "Collagen-Induced Arthritis," Current Protocols in immunology, 15.5 units (1993). See also the use of Issekutz, a.c. et al, Immunology (1996) 88: 569 to CD18 and VLA-4 integrin. Features similar to this CIA model seen in RA patients include, but are not limited to: erosion of cartilage and bone at the joint margins (as seen in radiographs), proliferative synovitis, symmetrical involvement of small and medium-sized peripheral joints of the extremities rather than the medial axis bone. The following procedure was followed to assess the efficacy of MGBG in treating arthritic disease.

Animals and drug administration. Inbred male DBA/1 mice (DBA/1OlaHsd, harlan laboratories) at least 7 weeks old can be used in the following collagen-induced arthritis model. Twenty animals per compound or vehicle were assigned to the arthritis and saline groups and 4 to the control group. To induce the arthritic state, mice were anesthetized with isoflurane and given 1501 μ L of bovine type II collagen in freund's incomplete adjuvant injection (day 0 and day 21). Mice were randomized into treatment groups according to body weight on study day 7. Treatment consisted of 25mg/kg MGBG, 0.2mg/kg dexamethasone as positive control or saline as vehicle control, all given oral gavage on study day 0 and continued daily (PO, BID twice daily/12 hours apart). Twenty mice per group can be used, with serum collected from 15 animals and plasma collected from five. Four other animals served as normal (untreated, non-arthritic) controls. The survival portion of the study may be performed for 35 days.

A compound is provided. The MGBG solution may be made from hydrate dihydrochloride; other salts may be used and in any case a salt/hydrate correction factor should be implemented. Solid MGBG can be stored at room temperature, but the dosage formulation should be prepared freshly for each administration. Dexamethasone is commercially available.

Data on days 21-35, arthritis usually developed, at this time, a clinical score was given for each paw (right anterior, left anterior, right posterior, left posterior), plasma was drawn on days 0, 14 and 25 to assess pharmacokinetics, and blood was drawn on days 0 and 28 for disease analysis, edema was measured on days 18-20, 22-27 and 29-34 inflammation was assessed by infiltration and edema of inflammatory cells after euthanasia, peripheral blood drawn was collected, heparinized and frozen at-70 ℃ until cytokines such as osteopontin, TNF α, IL-1, CRP, MCP1, MIP-1 β, tes, IFN γ, TGF β, IP-10, IL-17, and mmp9 were analyzed, anterior and posterior paws and knees were collected, processed, embedded, sectioned and stained with toluidine blue for tissue analysis after 4-5 days in a calcium remover, the presence of osteoclast damage or the extent of cartilage cell loss, and cartilage cell loss were quantified.

And (5) carrying out statistical analysis. Clinical data (means for animals) for paw scores were analyzed by determining the area under the dosing curve (AUC) from day 1 to day 15. For the calculation of AUC, the daily mean score for each mouse was entered into Microsoft Excel and the area between the days of treatment and the days of termination after the onset of disease was calculated. The mean value for each group was determined and% inhibition from the arthritic control was calculated by comparing the values of the treatment to normal animals. Differences in paw scores and histological parameters (mean + SE) were analyzed for each group using the Student's t test with significance set at p 0.05. The% inhibition for the histological parameters and AUC was calculated as [ (mean disease control-mean normal) - (mean treatment-mean normal) ]/[ [ (mean disease control-mean normal) · (mean treatment-mean normal) ] -100.

MGBG is ineffective in this model as a disease-modifying antirheumatic drug (DMARD), but does affect the early stages of paw swelling/inflammation. The above protocol may be varied according to methods known in the art.

In vivo mouse MOG-EAE multiple sclerosis model

An experimental murine model of multiple sclerosis using Myelin Oligodendrocyte Glycoprotein (MOG) peptide to induce Experimental Autoimmune Encephalomyelitis (EAE) was used to determine the efficacy of MGBG in preventing and treating that disease. Due to its many similarities to MS, EAE is commonly used to study autoimmune, CNS inflammation, demyelination, cell trafficking, and tolerance-induced pathogenesis. EAE is characterized by paralysis (in some models, paralysis is remission-relapsing), CNS inflammation, and demyelination. EAE is primarily mediated by dendritic cells, myelin-specific T cells (e.g., Th1 and Th17), and M1 macrophages. B cells can also be usedPlay a role in some models of EAE. Body weight also recorded disease progression. EAE was obtained in C57BL/6 mice by MOG in CFA emulsion_35-55Or MOG_1-125Immunization was performed followed by induction with Pertussis Toxin (PTX) in PBS. The emulsion provides antigens that trigger the expansion and differentiation of MOG-specific autoimmune T cells. PTX promotes EAE development by providing additional adjuvants and promoting the entry of autoimmune T cells into the CNS.

EAE induction. In C57BL/6 mice, chronic EAE was in MOG_35-55/CFA or MOG_1-125the/CFA emulsion was developed after immunization followed by pertussis toxin injection. This model is useful for testing compounds for their potential to prevent or ameliorate EAE disease. The model can be performed with the compound administered from the time of immunization (prophylactic treatment), or with the aim of reversing the disease course and promoting recovery by administering the compound from the time of onset of EAE (therapeutic treatment). Female C57BL/6 mice, 10 to 14 weeks old at the start of the study, were used for this model. Typically, EAE develops 8-18 days after immunization. EAE development usually continues for 4 weeks (28 days) after immunization.

Stress reduces susceptibility of mice to EAE. Administration of the treatment during the disease induction period (about 0-10 days after immunization) delays the onset of the disease and reduces the severity of the disease, in addition to the effect of any compound. This is due to the stress of compound administration and the effect of vehicle on mice. More frequent administration and less tolerance to vehicle and greater impact on disease progression. Administration of therapeutic stress and vehicle had much less impact on disease progression after the clinical signs of EAE appeared.

And (4) preventive treatment. In prophylactic studies, treatment is initiated prior to onset of disease, at the time of immunization, and at the time of dispensing. Mice were assigned to treatment groups in a balanced manner to achieve groups with similar body weight distribution. A prophylactic study assessed whether treatment would affect the progression of disease before and after the first clinical sign of EAE. To compensate for the stress of treatment in prophylactic treatment studies and to achieve the target disease severity, EAE can be induced with higher doses of pertussis toxin than used in therapeutic studies. The dose of pertussis toxin is based on the expected stress resulting from administration (route, frequency and formulation of vehicle).

In prophylactic studies, the median time to onset of disease is a sensitive measure of the efficacy of a compound. Small changes in immune response can produce delayed onset of disease-inhibition of T cell activation and proliferation, antigen presentation, differentiation into Th1 and/or Th17 cells will all result in delayed onset of EAE. The delayed onset with lower maximal severity of EAE indicates the overall efficacy of the treatment compared to the negative control group.

Some studies will show delayed onset of EAE without the effect of other significant compounds. In these cases, the compound may affect the early pathways in the development of the immune response, but eventually the excess processing compensates for the loss of the blocking pathway. Another possible explanation is that the drug is unable to maintain blockade of the pathway for the duration of the study.

In other studies, EAE was delayed but mice had a higher terminal EAE score than vehicle treated mice. Typically, this is not caused by compounds that worsen EAE, but by the peak of disease, is delayed in compound-treated mice and is consistent with the recovery phase of vehicle-treated mice.

An important reading of efficacy when a compound is administered prophylactically is the reduction in maximum disease severity (mean maximum score, MMS). A reduced MMS represents an overall reduction in EAE severity.

And (6) scoring. Clinically, EAE progression is scored on a scale of 0-5 with each score representing the following clinical observations:

0: no change in function;

1: tail limping;

2: limp tail and weak hind legs;

3: one of the following:

tail lameness and complete paralysis of the hind legs; or

Tail lameness and complete paralysis of one front leg and one rear leg; or

Severe head tilt, walk along the cage edge, push against the cage walls, and rotate when the tail is picked up;

4: tail claudication with complete paralysis of the hind leg and partial paralysis of the foreleg;

5: one of the following:

-complete anterior/posterior leg paralysis; or

-spontaneous rolling in the cage; or

Death secondary to paralysis.

Progression of EAE in untreated mice. Individual mice will have a slightly different disease course. Most mice showed initial signs of EAE between 9 and 14 days after immunization. Once EAE begins, the peak of disease almost always occurs 3-4 days later. The maximum score continues for several days, then the mice partially recover. In some mice, the disease will remain at maximum severity until the end of the study. Less often, the mice will remain at peak severity for only one day, followed by the onset of recovery. The degree of recovery depends largely on the maximum severity achieved by the mouse. Most untreated or vehicle-treated mice will not fully recover, but their terminal scores will typically be 0.5 to 1.5 points lower than their maximum scores. Approximately 25% of untreated or vehicle treated mice showed worsening EAE between 24 and 28 days after immunization, with similar relapse. The spinal cords of these mice had numerous foci of inflammation at the time of EAE exacerbation (> 7 foci per section), similar to histological observations at the time of EAE onset and peak, suggesting that these are true relapses with a new wave of inflammation in the spinal cord. When mice are followed for longer periods of time, the disease severity slowly increases, resembling the chronic progressive disease course observed in human MS patients.

Changes in body weight during EAE reflect the severity of the disease. Mice often lose a small amount of body weight the day after immunization. This appears to be due to the effect of the adjuvant and pertussis toxin administered. The mice then steadily gained their body weight until onset of disease. Mice continually lost 1-2g of their body weight (5-10% body weight) on the day of EAE onset. Weight loss continues as EAE severity progresses, with loss reaching about 20% of its pre-onset body weight at peak disease. Weight loss is likely due to paralysis and decreased food intake as well as high production of proinflammatory cytokines such as TNF during the acute phase of inflammation. After the disease peaked, mice slowly gained weight even though their clinical scores did not improve. The increase in weight may be due to down-regulation of inflammation, which results in reduced levels of pro-inflammatory cytokines in the blood. Untreated or vehicle-treated mice typically have about 90% of their preimmune body weight 28 days after immunization.

Histology. Typically, histological analysis is performed at the end of the study (typically about 28 days after immunization) or at the time when the peak of disease is reached in the vehicle group (typically 14-18 days after immunization) and focuses on inflammatory foci, apoptosis and demyelination, each of which will be described below. Inflammation of the EAE generally begins in the lumbar region of the spinal cord and spreads throughout the spinal cord by the time of peak disease.

And (4) apoptosis. Apoptotic cells were identified in H & E sections and were not usually found during the first two days of disease progression. They are found during the peak and chronic phases of EAE. The average number of apoptotic cells is typically between 2 and 4 per slice. Apoptotic cells are neurons and their number is related to the stage of the disease. Apoptotic cells appear shortly after the onset of disease, so there will be many foci of inflammation at the onset of EAE, with few apoptotic cells. Subsequently, the number of apoptotic cells increases until the peak of the disease, and then remains elevated.

Inflammation is caused. At the onset of the disease, the number of inflammatory foci strongly correlates with the severity of the disease. The number of lesions increases slightly up to the peak of the disease, when 6-15 lesions/sections of inflammation are usually found throughout the spinal cord. In the chronic phase of EAE (beginning days after peak disease), many inflammatory foci resolve, typically resulting in 3-4 inflammatory foci in each spinal cord section by about 28 days after immunization.

Because the greatest number of inflammatory foci exists in the early stages of disease progression, mice with late-stage EAE episodes often have more inflammatory foci in their spinal cords than might be expected from their clinical scores if histological analysis were performed at the end of the study. For example, in a 28-day study, mice with an EAE onset 27 days after immunization and a final clinical score of 2 will likely have more foci of inflammation than mice with an EAE onset 9 days after immunization and a final score of 3.5. Similarly, mice that relapse shortly before the end of the study (relapse is defined as a 1 or greater increment in clinical score) will typically have more foci of inflammation at the end of the study than mice with stable chronic disease, even though both have the same clinical score at the end of the study.

Inflammatory foci of approximately 20 cells were counted in each H & E stained section. When the inflammatory infiltrate is composed of more than 20 cells, the estimate is made of how many foci of 20 cells are present.

Demyelination. Demyelination is usually not found during the first two days after onset of disease, but is seen at the time of peak disease (4-5 days after onset of EAE) and continues during the chronic phase of EAE. Demyelination scores vary little between the peak and 28 days after immunization and typically have an average value between 1.2 and 2.5.

Demyelination was scored in Luxol fast blue stained sections (LFB) and H & E sections. In LFB sections, white matter of the spinal cord is stained dark blue and areas of demyelination are lighter blue and associated with large vacuoles. In H & E stained sections, destruction of standard tissue with large vacuoles indicates demyelination.

The demyelination score represents an estimate of the area of demyelination for each of the following fractions:

0-No demyelination (less than 5% demyelinated area)

1-5 to 20% demyelinated area

2-20 to 40% demyelinated area

3-40 to 60% demyelinated area

4-60 to 80% demyelinated area

5-80 to 100% demyelinated area

For the Luxol fast blue stained slides, the size of the demyelinated area was estimated based on the less intense blue staining of myelin. For H & E stained sections, areas of demyelination were estimated by looking for breaks in standard tissue-pale and vacuolized to conform to edema and demyelination, and expanded axons.

And (5) carrying out statistical analysis. Unless otherwise stated, statistical analysis was performed as follows: incidence of disease compared using chi-square test; mean days of EAE onset, changes in body weight, and number of apoptotic cells compared using the 2-tailed Student's t test; median days of EAE onset compared using Wilcoxon survival test; mean Maximum Score (MMS), terminal score, and demyelination score compared using the Wilcoxon nonparametric test (LFB and H & E).

First MOG EAE scheme.

Materials and methods. Ten-week-old female C57BL/6 mice (Taconnic farm) weighing 18-23g were divided into 4 groups: mock immunization (n ═ 3) and three MOG immunization groups received 30mpk BID of MGBG (as di HCl monohydrate, correction factor 1.49), 3mpk QB fingolimod (FTY720) at the second mock vehicle dose, or 0.9% saline vehicle BID (n ═ 12 per group). MOG_35-55Peptides were prepared by applying the Hooke Kit to two sites on the back^TMMOG_35-55Emulsion component of/CFA emulsion PTX (catalog No. EK-2110(Hooke Laboratories, Lawrence MA)) (containing MOG for test article or positive control)_35-55Or PBS for negative control). One site of injection was in the upper dorsal region, about 1cm caudal to the cervical line. The second part is in the lower back region and is far from the skull of the caudal rootSide about 2 cm. The injection volume was 0.1mL per site. The pertussis toxin component of the kit (diluted with PBS to achieve 176 ng/dose for the first injection and 165 ng/dose for the second injection) was administered intraperitoneally within 2 hours of the injection of the emulsion, followed by another 24 hours after the injection of the emulsion. The volume of each injection was 0.1 mL.

Vaccinated, untreated mice were under MOG_35-55EAE developed 8-14 days after CFA immunization and chronic paralysis was maintained for the 28-day duration of the experiment. Clinical scores, body weight, and histopathology (e.g., demyelination, inflammatory infiltrates, and/or apoptosis) of the spinal cord may be measured and recorded as described above.

And (6) obtaining the result. Clinical scores given as mean score +/-mean Standard Error (SEM) are shown in figure 1. As can be seen in figure 1, vehicle treated animals developed EAE clinical signs starting on day 10 as expected, with mean clinical signs increasing until day 16 when the score outperforms 2 and remained thereafter between 2 and 3. Fingolimod was most effective at the large doses tested, yielding an average clinical score of 0 throughout the experiment, nearly indistinguishable from the levels of the mock-immunized group. MGBG also prevented the onset and progression of EAE, with first clinical signs becoming evident on day 15 and slowly increasing to a level still below 1 on day 21, and remaining below 1 for the duration of the experiment. The results indicate that MGBG is effective in preventing and treating neuropathic symptoms of MS like fingolimod in an accepted disease model.

In addition, as shown in figure 2, treatment with fingolimod or MGBG prevented weight loss in the EAE model. The results of FIG. 2, given as% change from baseline (start of study) +/-SEM, show that all groups continued to gain weight until about day 8, when about 107-113% weight was observed. Thereafter, the vehicle treated group began weight loss with a rapid decrease to 95% on day 15 and as low as about 93% on day 20, with a slight recovery to about 95% by the end of the study. In general, the sham-immunized and MGBG and fingolimod treatment groups each maintained an increase in body weight throughout the study, lingering between about 108% and about 114%. The fingolimod and vehicle groups showed a slight trend of gradual and continuous weight gain over the course of the study. Subjects treated with MGBG showed an initial maximal significant increase, approaching 115% on day 11, but showed a gradual decrease to about 108% on day 24, followed by a slight increase. These results further indicate that MGBG is effective in preventing and treating symptoms of MS like fingolimod.

Histopathology. On day 28 (end of study), all mice were sacrificed for histological analysis and perfused with PBS and the spine collected in 10% buffered formalin. For each mouse, 3 Luxol fast blue stained sections and 3H & E sections were prepared from lumbar, thoracic and cervical vertebrae and analyzed by a pathologist blinded to experimental groups and all clinical readings. The number of inflammatory foci, apoptotic cells and demyelinated regions was determined in each of the three H & E sections.

Histopathological results are shown in figures 3-5, which confirm that MGBG, like fingolimod, reduces histopathological signs of MS such as neuroinflammation, demyelination (via two stains), and apoptosis. In addition, the negative control group (data not shown) showed zero score.

Second MOG EAE scheme.

A second EAE experiment was performed essentially as disclosed above, except that the fingolimod dose was 1mpk QD and pertussis toxin was administered at 165 ng/dose for both injections.

And (6) obtaining the result. EAE was more severe in this study than in the typical study conducted according to this protocol, and more severe than in the previous study. All mice in the vehicle group developed severe EAE. Disease progression in the fingolimod treatment group was not inhibited as strongly as in the previous study or in the typical study. This is not surprising given that more severe disease is generally more difficult to suppress. Mice in the MGBG treated group developed substantially reduced disease compared to the vehicle group, showing comparable efficacy to fingolimod.

The results are given in FIGS. 6-10. Consistent with previous studies, MGBG was effective in delaying the onset of disease and reducing the severity of EAE.

Fluorescence activated cell sorting. In addition to clinical scores, flow cytometric analysis was performed on CNS infiltration cells from 6 mice in each of the vehicle, fingolimod and MGBG treatment groups, and all three mice from the mock-immune (disease-free) group.

On day 28, brain and spinal cord tissues in the second MOG EAE study (6 out of 12 mice from each MOG-immunized group; and all 3 mice from the mock-immunized group) were pooled and infiltrated cells were isolated by Percoll gradient. After the infiltrated cells were isolated individually from each mouse, the cells were placed in a medium containing fosaprepinyl acetate (PMA, 50ng/mL), ionomycin (0.5. mu.g/mL) and brefeldin (1. mu.g/mL) and incubated for 4-5 hours. The cells were then washed and stained for flow cytometry analysis as explained below. The following analysis was performed (Becton dickinson facscan):

anti-CD 4-Cy-5/anti-IL-17A-PE/anti-IFN γ -FITC (Th1/Th17 cells)

anti-CD 4-Cy-5/anti-CD 11 c-PE/anti-CD 45.2-FITC (infiltrating dendritic cells)

anti-CD 11 b-Cy-5/anti-IL-12-PE/anti-CD 45.2-FITC (M1 macrophages)

anti-CD 11 b-Cy-5/anti-CD 206-PE/anti-IL-10-FITC (M2 macrophages)

The number of cells positive for the relevant staining and combinations of staining was calculated as the percentage of cells analyzed and the average number of cells per mouse per group.

TABLE 8 effects of CNS infiltrating cells and cytokines

Treated with MGBGMice had significantly reduced relative and absolute numbers of CNS-infiltrating dendritic cells (DC, CD45RB high, CD11 c)⁺) And a significantly reduced number of IL-12 producing cells in CNS infiltrating CD11 b-cells. The results are given in the table below, where denotes p < 0.05 and denotes p < 0.1. Tables 9 and 11 show the results with respect to the percentage of total cells, and tables 10 and 12 show the results with respect to the original number of cells (10)³) The result of (1).

TABLE 9

Watch 10

TABLE 11

TABLE 12

Third MOG EAE scheme.

The third EAE experiment was performed essentially as disclosed above, except that fingolimod was administered at 0.1mpk QD (which is well-matched to a typical human dose or 0.5 mg/day adjusted for body surface area), and another set of MGBG 30mpk combined with fingolimod 0.1mpk was added. Here again, pertussis toxin was administered at 165 ng/dose for both injections.

And (6) obtaining the result. Fingolimod and MGBG alone showed comparable efficacy in reducing disease severity and delaying disease onset at the doses tested. There were also significantly fewer inflammatory foci and significantly less demyelination in treated mice compared to vehicle-treated mice. Importantly, none of the mice in the fingolimod/MGBG combination group developed EAE, and all other clinical readings (e.g., body weight) were significantly improved (no signs of EAE) compared to the vehicle group-no detectable disease in these mice.

Overall, these results indicate that the combination of fingolimod and MGBG is highly effective in preventing the development of EAE. In addition, mice receiving combination therapy had significantly improved clinical and histological readings compared to mice receiving fingolimod or MGBG alone. This combination appears to prevent the development of disease.

Other combinations of agents can be readily tested in a protocol similar to that described above, e.g., interferon β -1a, interferon β -1b, glatiramer acetate, mitoxantrone, natalizumab, laquinimod, dimethyl fumarate (tecfidera), and teriflunomide can each be tested in combination with MGBG other SAMDC inhibitors can be similarly tested.

Fourth MOG EAE protocol.

A fourth EAE experiment was performed similarly as described above, but was designed to examine the effect of MGBG on cell populations during onset and peak of disease. In this protocol, there were 3 experimental groups (20 mice/group) and 1 mock-immunized group (8 mice). Fingolimod was given at 1mpk QD, MGBG was given at 30mpk BID, and pertussis toxin was administered at 165 ng/dose for both injections. Flow cytometric analysis was performed on CNS infiltrating cells from 16 mice in each of the vehicle, fingolimod and MGBG treatment groups and all mice from the mock-immunized group. Half of the mice analyzed from each group were collected in the vehicle group at the onset of EAE and half at the peak of EAE in the vehicle group. Mice were terminated on eight separate days, as individual mice in the vehicle group reached EAE onset and EAE peak on different days. Eight (8) mice in the vehicle group were sacrificed at each time point. For each of these mice, matched mice from each of groups 2 and 3 were sacrificed on the same day. For each two of these vehicle group mice (2), matched mice from group 4 were sacrificed. When matching mice from other groups were selected for termination, the mouse with the highest EAE score was selected. If no mice in the group had signs of EAE, mice were randomly selected.

And (6) obtaining the result. CNS infiltration of pro-inflammatory cells was reduced in MGBG treated mice, consistent with the clinical findings that MGBG reduces EAE development. Multiple proinflammatory cell populations were reduced in the CNS of MGBG-treated mice. Interestingly, the proportion and number of infiltrating dendritic cells decreased more in MGBG-treated mice than in fingolimod-treated mice, and some differences were statistically significant. These results are consistent with previous studies and indicate that MGBG selectively targets dendritic cells. In addition, the reduction in the number of IL-12 producing cells supports the view that MGBG influences the development of Th-1 type responses.

The results are given in the table below, where denotes p < 0.05 and denotes p < 0.1. Table 13 shows the results as a percentage of total cells, and Table 14 shows the results as a percentage of the original number of cells (10)³) The result of (1).

Watch 13

TABLE 14

Table 15 below illustrates the effects seen in a subset of animals that did not develop disease when the vehicle-treated animals were at peak disease. MGBG, but not fingolimod, reduces the number of dendritic cells. This suggests that MGBG (and more broadly, SAMDC inhibitors have an effect on dendritic cell antigen presentation and CNS infiltration) may be an important component in preventing or reducing the severity of EAE and MS and/or its progression and associated symptoms such as demyelination.

Watch 15

Additional in vivo models of therapeutic efficacy

The following models, presented by way of example, can be used to assess the efficacy of the compounds disclosed herein in treating a number of diseases and indications. It is within the ability of those skilled in the art to modify these models to meet the needs of the study. In addition, one skilled in the art will be familiar with additional disease models that may be employed. It is expected that MGBG, as well as other polyamine analogs and polyamine biosynthesis inhibitors and the compounds disclosed herein, will be effective in these models.

Neuropathy and neuropathic pain models

Bannit model of neuropathic pain:

peripheral mononeuropathy is produced in adult rats by loose ligation around the sciatic nerve. Post-operative behavior of these rats indicates the development of hyperalgesia, allodynia and possibly spontaneous pain (or dysesthesia). Hyperalgesic responses to noxious radiant heat are usually evident on the second day after surgery and persist for 2 months. There is also a hyperalgesic response to chemically induced pain. The presence of allodynia can be presumed to result from an nociceptive response elicited by standing on a harmless cold metal floor or by harmless mechanical stimulation (e.g., with von Frey filaments) and continued holding of the hind paw by the rat in a protected position. The presence of spontaneous pain is indicated by appetite suppression and by the frequent occurrence of pronounced spontaneous nociceptive responses. Affected hind paws are usually abnormally warm or cold in about one third of the rats. Approximately one half of the rats developed severely overgrown paws on the affected side. In a compound efficacy model, the test compound is typically delivered prior to stimulation and the vehicle serves as a control. Experiments with this animal model can be advanced to understand the neural mechanisms of neuropathic pain disorders in humans. Bennett GJ, Xie YK, 1988 "A perpheral monoeuropathy in a rate processes of a paper sensing like sugar section in man," Pain, month 4; 33(1): 87-107 (PMID: 2837713).

Chung model of neuropathic pain

Since its introduction in 1992, the Spinal Nerve Ligation (SNL) model of neuropathic pain has been widely used in various research works on the mechanisms of neuropathic pain and in screening assays for the development of novel analgesics. This model was developed by tightly ligating one (L5) or two (L5 and L6) segmental spinal nerves in rats. This procedure results in persistent behavioral signs of mechanical allodynia, thermal hyperalgesia, cold allodynia, and persistent pain. In a widely used process, many different variation patterns of SNL models have been created, either intentionally or unintentionally, by different researchers. While the factors that contribute to these changes themselves are interesting and important topics to be studied, the pain mechanisms involved in these changes may differ from the original model. Methods for generating spinal nerve ligation models that minimally induce the underlying factors that may contribute to these changes are described in detail in Chung JM, Kim HK, and Chung K, "segmented spinal neutral ligation model of neural disease pain," Methods Mol Med; 200499: 35-45 (PMID: 15131327).

Chung model in NHP

In a painful neuropathy model in primates (cynomolgus monkeys), the neuropathic state is induced by the tight junction of the L7 spinal nerve just distal to the root ganglion behind the L7 spinal nerve. Sensory testing may be done on the ventral aspect of the foot (the area including the dermatome L7). Within 1 week after surgery, primates often develop significant sensitivity to mechanical stimuli (e.g., with von Frey hair), indicating the presence of mechanical allodynia. An increase in sensitivity to mechanical stimuli is sometimes also observed on the contralateral side. A decrease in the withdrawal threshold for thermal stimuli indicates the presence of thermal hyperalgesia. The presence of various cold stimuli, such as acetone and cold water baths, suggests that cold allodynia also develops. The observed behavioral phenomena were similar to those seen in persons diagnosed with peripheral neuropathic pain. Thus, the model is useful for assessing a number of parameters associated with neuropathy and neuropathic pain conditions in humans, and for assessing the efficacy of drug candidates as treatments for the associated conditions. See, e.g., Carlton SM et al, "therapeutic models for therapeutic neural produced by spinal negative neural neutral restriction in the matrix," Pain 1994 month 2; 56(2): 155-66 (PMID: 8008406).

Tactile allodynia assessment with Von Frey filaments

The following quantitative allodynia assessment techniques can be modified to measure tactile allodynia in any of a variety of animal models of neuropathic pain. The following summary is given by way of example and refers to a model of surgical neuropathy in rats in which nociceptive behavior is induced by gentle contact with the paw. With 0.41 to 15.1g of von Frey hair, the% response at each stimulation intensity can be characterized first. A smooth log-linear relationship is generally observed. Additionally or alternatively, schemes using stimulus oscillations around the response threshold may be employed, which allow for faster, efficient measurements. The correlation coefficient between these two methods is usually high. In neuropathic rats, good intra-and inter-observer reproducibility in the top and bottom protocol was found; some variability can be seen in normal rats due to extensive testing. The fact that the threshold values in the group of rather large neuropathic rats showed insignificant variability within 20 days and 61% still met the strict neuropathy criterion after 50 days (using the survival analysis) indicates that the combination of the threshold value measurement using the top-bottom protocol and the neuropathic pain model represents an effective means for analyzing the treatment impact of neuropathic pain states. See, e.g., Chaplan SR et al, "quantitative assessment of tactle allodynia in the rat paw," J Neurosci Methods, month 7 of 1994; 53(1): 55-63 (PMID: 7990513).

Hagerivs method for assessing thermal nociception

Alternatively, methods have been described for measuring cutaneous hyperalgesia to thermal stimuli in unconstrained animals. The test protocol uses automatic detection of behavioral endpoints; repeated testing did not promote the development of the observed hyperalgesia. Carrageenan-induced inflammation leads to significantly shorter paw withdrawal latencies compared to saline-treated paws and these latency changes correspond to a reduced thermal nociceptive threshold. This sensitive thermal method detects dose-related hyperalgesia and its blockade by test compounds and allows measurement of other behavioral parameters in addition to nociceptive thresholds. See, e.g., Hargreaves K et al, "A new and positive method for measuring thermal nociception in cutaneousyphylgesia," Pain, 1 month 1988; 32(1): 77-88 (PMID: 3340425).

Inflammatory and autoimmune models

Contact dermatitis and related disorders

Contact hypersensitivity is a simple delayed-type hypersensitivity in vivo assay of cell-mediated immune function that can be used to assess potential therapeutic efficacy in many conditions with inflammatory and/or autoimmune components. The diseases include contact dermatitis, atopic dermatitis, psoriasis, allergic dermatitis and skin irritation. The compounds may optionally be administered topically in a topical formulation, or may be delivered by a non-topical (e.g., oral, intravenous, etc.) route.

Mouse model

In one procedure, exposure of skin to exogenous haptens results in a delayed-type hypersensitivity reaction, which is measured and quantified. Contact sensitivity involves an initial sensitization phase followed by an excitation phase. The priming phase occurs when a T lymphocyte encounters an antigen to which it has previously been exposed. Swelling and inflammation appear, making it an excellent model of allergic contact dermatitis in humans. Murine models also typically have the added benefit of low cost. Suitable procedures are described in detail in Gaspari AA and Katz SI, "Contact Hypersensitivity," Current Protocols in Immunology, 4.2 Unit, John Wiley & Sons, Inc. (1994). See also Grabbe S and Schwarz T, "Immunoregulationhanding involved in the experimentation of organizational contact hyper responsiveness," Immun. today 19 (1): 37-44(1998).

Pig model

The choice of animal is important in dermatological studies where it is desirable to predict human responses. For this reason, pigs and in particular miniature pigs are advantageous because of the similarity between human skin and pig skin (especially hair follicle density). See, e.g., Billskia J and Thomson DS, "organic contact modifications in the social pig. A new model for evaluating the topical anti-reflective activities of drugs and the formulations," Br J Dermatol, 7.1984; 111, supplement 27: 143 (PMID: 6743545).

Hairless guinea pig model

Allergic and irritant contact reactions were also reported in recently identified hairless guinea pigs Crl: iaf (ha) BR (a mutant from the hartley line). Irritant contact dermatitis may be induced by croton oil, 2, 4-Dinitrochlorobenzene (DNCB) or anthralin. Hairless and hairy guinea pigs developed similar responses to these chemicals. Photoallergic contact sensitization can also be induced with Tetrachlorosalicylanilide (TCSA), or with cyclophosphamide prior to sensitization with tribromophenyl salicylate (TBS). The skin changes were observed visually and under a microscope according to methods known in the art. Accordingly, hairless guinea pigs may be used as animal models for the assessment of test compounds in the treatment of immune and non-immune contact responses and related conditions. See, e.g., Miyauchi H and Horio T, "Angle animal model for contact information: the hairlessguineapip "J Dermatol.1992, month 3; 19(3): 140-5 (PMID: 1640019).

Simple skin irritation was also studied in hairless guinea pigs. In one exemplary model, test compounds were delivered in one or more topical formulations for 30min with 4 days of daily exposure. Scoring every day; evaporation assay (total epidermal water loss (TEWL)), hydration and colorimetric were measured at baseline (day 0), mid-treatment and end-of-treatment. Test compounds were administered twice daily. See, e.g., Andersen F et al, "The hairless guineea-pig as a model for treatment of clinical intervention in humans," Skin Res Technol.2006, month 2; 12(1): 60-7 (PMID: 16420540).

Mouse chimera model for psoriasis

In addition, the compounds disclosed herein can be tested in animal models of psoriasis-like diseases. The study of the causes and pathophysiological mechanisms underlying psoriatic skin lesions has been hampered by the lack of appropriate animal models for this common and unappreciated skin disease. A suitable model is, for example, that described by Nickoloff BJ et al, "Severe combined immunological specificity mouse and human regulatory skin cameras. differentiation of a newanimal model," Am J Pathol., 1995 month 3; 146(3): 580-8 (PMID: 7887440) human skin/scid mouse chimeras prepared as described. Wherein the method characterizes normal skin, pre-psoriatic skin and psoriatic plaque skin samples transplanted onto severe combined immunodeficient mice. Normal, pre-psoriatic or psoriatic plaque keratome skin samples were transplanted onto severe combined immunodeficient mice that reliably had high graft survival (> 85%) and had repeatable changes observed throughout the duration of implantation. After transplantation, normal skin remained essentially normal, whereas pre-psoriatic skin became thicker and psoriatic plaque skin maintained its characteristic plaque-like projections and scales by clinical assessment and routine light microscopy. By using a panel of antibodies and immunohistochemical analysis, the overall phenotype of human cell types (including immune cells) remaining in the transplanted skin was significantly similar to that of the skin sample prior to transplantation. In addition, the clearly recognized interface zones between human and mouse skin within the epidermal and dermal compartments can be identified by routine microscopic examination and immunostaining, with focal regions of chimerism. The numerous similarities between pre-and post-transplant human samples of normal and psoriatic skin transplanted onto severe combined immunodeficient mice make this animal model useful for assessing the efficacy of test compounds in treating psoriasis and related conditions.

Psoriasis mouse scid/scid model

Alternatively, the compounds disclosed herein may be prepared fromMP et al, "Murine psioriasis-likedisc induced by negative CD4+ T cells," Nat Med., 1997, month 2; 3(2): 183-8 (PMID: 9018237 in the scid/scid mouse model in this model, the use of minor histocompatibility mismatched naive CD4+ T lymphocytes to reconstitute scid/scid mice produced skin changes that surprisingly resembled human psoriasis in clinical, histopathological, and cytokine expression.

Asthma (asthma)

The compounds may additionally be evaluated for efficacy in treating asthma and related lung disorders. In a murine asthma model, wild-type control [ C57BL/6J, (+/+) ] and ICAM-1 (intercellular adhesion molecule-1) knockout [ C57BL/6J-ICAM-1, (/ -) ] mice were sensitized to Ovalbumin (OVA) and challenged with OVA delivered by aerosol (OVA-OVA) to induce a phenotype consistent with an asthmatic response. Bronchial responsiveness to methacholine and counts of cell number, as well as measurement of eosinophil content and cytokine levels in bronchoalveolar lavage fluid (BALF) can be measured. In addition, lymphocyte proliferation, eosinophil migration into the airway, and development of Airway Hyperresponsiveness (AHR) in allergen-sensitized and challenged mice in response to antigen can all be measured in vivo or ex vivo according to methods known in the art. See, Wolyniec WW et al, "Reduction of anti-induced air reactivity and eosinophilicity in ICAM-1-specificity micro," Am J Respir Cell Mol biol., 1998, 6 months; 18(6): 777-85 (PMID: 9618382).

Inflammatory bowel disease, Crohn's disease and ulcerative colitis

The compounds disclosed herein can also be evaluated for activity in animal models of inflammatory bowel disease, crohn's disease, and ulcerative colitis. By Scheiffele F, Fuss IJ, "introduction of TNBS colitis in mic," Curr ProtocImmunol, 8.2002; chapter 15: the protocol described in unit 15.19 (PMID: 18432874) is one of several that have been used to study the immunopathogenesis of these diseases. This model employs 2, 4, 6-trinitrobenzenesulfonic acid (TNBS), which induces severe colitis when administered intrarectally in SJL/J mice. Colitis caused by this procedure appears to be similar to the clinical and histopathological observations seen in crohn's disease. Scheifflele and Fuss discuss the key parameters required for successful induction of TNBS colitis and methods for monitoring and grading disease levels, and present a support protocol for isolation of lamina propria monocytes from the colon of mice. See also Morris GP et al, "Hapten-induced model of cyclic information and differentiation in the rat colon," Gastroenterology, 3 months 1989; 96(3): 795-803 (PMID: 2914642), which describes the original rat model of chronic colitis by intraluminal instillation of a solution containing a "barrier breaker" (e.g., 0.25ml 50% ethanol) and a hapten (e.g., TNBS, 5-30 mg). At a dose of 30mg, trinitrobenzenesulfonic acid/ethanol induced ulceration and significant thickening of the intestinal wall lasted for at least 8 weeks. Histologically, the inflammatory response involves mucosal and submucosal infiltration of polymorphonuclear leukocytes, macrophages, lymphocytes, connective tissue mast cells, and fibroblasts. Granuloma (3 weeks after induction of inflammation), giant cells of the langerhans type, segmental ulcers and inflammation. The characteristics and relatively long duration of the inflammation and ulcers induced in these models provide the opportunity to study the pathophysiology of colonic inflammatory diseases in a particularly controlled manner and to evaluate new treatments potentially applicable in human inflammatory bowel disease.

Exemplary oral pharmaceutical formulations

The following are examples of compositions that may be used to orally deliver the compounds disclosed herein in capsule form.

The compound of formula VI in solid form is passed through one or more screens to produce a consistent particle size. The excipient may also pass through a screen. An appropriate weight of compound sufficient to achieve the target dose per capsule can be measured and added to the mixing vessel or device, and then the blend mixed until homogeneous. Blending uniformity can be accomplished, for example, by sampling 3 points (top, middle, and bottom) within the vessel and testing the efficacy of each sample. A target of 95-105% of the test results (where RSD is 5%) was considered ideal; optionally, additional blending time may be allowed to achieve uniform blending. Once acceptable blend uniformity results are obtained, a measured aliquot of this stock formulation can be separated to produce lower strength. The magnesium stearate can be passed through a screen, collected, weighed, added as a lubricant to a blender, and mixed until dispersed. The final blend was weighed and blended. The capsule may then be opened and the blended material flooded into the capsule body using a spatula. The capsules in the tray may be tamped to deposit the blend in each capsule to ensure a uniform target fill weight, and then sealed by combining the filling body with a cap.

Composition examples

In the following composition examples, the target dosage may be adjusted taking into account the weight of the counter ion and/or solvate, if administered as a salt or solvated polymorph thereof. In this case, the weight of other excipients, typically fillers, is reduced. For example, in the case of the dihydrochloride monohydrate MGBG salt, a correction factor of 1.49 is used (e.g., 360mg of the salt gives 240.8mg of the free base).

Example 1A: 300mg capsule: the total fill weight of the capsules was 500mg, excluding the capsule weight. The target compound dose is 300mg per capsule

Composition (I)	Amount per capsule, mg
		MGBG	300.00
Lactose monohydrate	179.00
		Silicon dioxide	3.00
Cross-linked polyvidone	15.00
		Magnesium stearate (vegetable grade)	3.00

Example 1B: 150mg capsule: the total fill weight of the capsules was 300mg, excluding the capsule weight. The target compound dose is 150mg per capsule

Composition (I)	Amount per capsule, mg
		MGBG	150
Microcrystalline cellulose (MCC)	147
		Magnesium stearate (vegetable grade)	3

MGBG-Fingolimod combination examples: the total fill weight of the capsules is given in mg below, excluding capsule weight.

All references cited herein are incorporated by reference in their entirety as if written herein. From the above description, one skilled in the art can easily ascertain the essential characteristics of the present invention, and without departing from the spirit and scope thereof, can make various changes and modifications of the invention to adapt it to various usages and conditions. The invention disclosed herein provides embodiments wherein each of the above embodiments is combined with one or more other non-contradictory embodiments such that a resulting embodiment includes two or more of the recited elements and/or limitations.

Claims

Use of MGBG in the manufacture of a medicament for treating progressive multiple sclerosis in a patient, comprising administering a therapeutically effective amount of MGBG to a patient in need thereof.
2. The use as recited in claim 1, wherein the administration of MGBG is oral.
3. The use as claimed in claim 2, wherein MGBG is administered at 20 mg/day to 400 mg/day.
4. The use of claim 3, further comprising administering an agent selected from interferon β -1a, interferon β -1b, glatiramer acetate, mitoxantrone, natalizumab, fingolimod, laquinimod, dimethyl fumarate, and teriflunomide.
5. The use of claim 4, wherein the agent is fingolimod.
6. The use of claim 5, wherein fingolimod is administered at 0.5mg per day.
7. The use of claim 5, wherein fingolimod is administered at less than 0.5mg per day.
8. The use of claim 5, wherein fingolimod is administered at 0.25mg per day.
9. The use of claim 1, wherein the treatment prevents relapse or progression of MS.
10. The use of claim 1, wherein administration occurs with a reduced incidence of at least one side effect selected from the group consisting of: cytopenia, nephrotoxicity, hepatotoxicity, cardiotoxicity, teratogenicity, impaired lung function, macular edema, peripheral neuropathy, severe skin reactions, increased risk of infection, innate immune damage, adaptive immune damage, and flushing.
11. The use of claim 10, wherein administration occurs with a reduced incidence of at least one side effect selected from the group consisting of: cytopenia, renal toxicity, hepatotoxicity, cardiotoxicity and teratogenicity.
12. The use of claim 11, wherein the cytopenia is selected from the group consisting of lymphopenia and neutropenia.
13. The use of claim 10, wherein administration occurs with a reduced incidence of at least two side effects selected from the group consisting of: cytopenia, renal toxicity, hepatotoxicity, cardiotoxicity and teratogenicity.
14. The use of claim 13, wherein the cytopenia is selected from the group consisting of lymphopenia and neutropenia.
15. The use of claim 13, wherein the administration occurs with a reduced incidence of cytopenia, nephrotoxicity and hepatotoxicity.
16. The use of claim 15, wherein the cytopenia is selected from the group consisting of lymphopenia and neutropenia.
17. The use of claim 16, wherein the administration additionally occurs with a reduced incidence of cardiotoxicity and teratogenicity.
18. The use of claim 1, wherein the progressive multiple sclerosis is primary progressive multiple sclerosis.
19. The use of claim 1, wherein the progressive multiple sclerosis is secondary progressive multiple sclerosis.
Use of MGBG in the manufacture of a medicament for preventing the progression of multiple sclerosis in a patient, comprising administering MGBG to a patient in need thereof.
21. The use of claim 20, wherein the relapsing symptoms of multiple sclerosis are alleviated.
22. The use of claim 20, wherein the symptoms of multiple sclerosis are prevented to a clinically significant or detectable level.
23. The use of claim 20, wherein the occurrence or severity of an increased flare to multiple sclerosis progression characterized by increased demyelination or decreased motor capacity over time is prevented.
24. The use as recited in claim 20, wherein the administration of MGBG is oral.
25. The use as claimed in claim 20, wherein MGBG is administered at 20 mg/day to 400 mg/day.
26. The use of claim 25, wherein administration occurs with a reduced incidence of at least one side effect selected from the group consisting of: cytopenia, nephrotoxicity, hepatotoxicity, cardiotoxicity, teratogenicity, impaired lung function, macular edema, peripheral neuropathy, severe skin reactions, increased risk of infection, innate immune damage, adaptive immune damage, and flushing.
27. The use of claim 26, further comprising administering an agent selected from interferon β -1a, interferon β -1b, glatiramer acetate, mitoxantrone, natalizumab, fingolimod, laquinimod, dimethyl fumarate, and teriflunomide.
28. The use of claim 27, wherein the agent is fingolimod.
29. The use of claim 28, wherein fingolimod is administered at 0.5mg per day.
30. The use of claim 28, wherein fingolimod is administered at less than 0.5mg per day.
31. The use of claim 28, wherein fingolimod is administered at 0.25mg per day.
32. The use of claim 26, wherein administration occurs with a reduced incidence of at least one side effect selected from the group consisting of: cytopenia, renal toxicity, hepatotoxicity, cardiotoxicity and teratogenicity.
33. The use of claim 32, wherein the cytopenia is selected from the group consisting of lymphopenia and neutropenia.
34. The use of claim 26, wherein administration occurs with a reduced incidence of at least two side effects selected from: cytopenia, renal toxicity, hepatotoxicity, cardiotoxicity and teratogenicity.
35. The use of claim 34, wherein the cytopenia is selected from the group consisting of lymphopenia and neutropenia.
36. The use of claim 34, wherein the administration occurs with a reduced incidence of cytopenia, nephrotoxicity and hepatotoxicity.
37. The use of claim 36, wherein the cytopenia is selected from the group consisting of lymphopenia and neutropenia.
38. The use of claim 36, wherein the administration occurs additionally with a reduced incidence of cardiotoxicity and teratogenicity.
Use of MGBG in the manufacture of a medicament for preventing or reducing the severity of an autoimmune response in an initial stage of a patient suffering from multiple sclerosis, comprising administering MGBG to a patient in need thereof.
40. The use of claim 39, wherein the administration additionally prevents or reduces an amplified stage of an autoimmune response in a patient suffering from multiple sclerosis.
41. The use as recited in claim 39, wherein the administration of MGBG is oral.
42. The use as claimed in claim 41, wherein MGBG is administered at 20 mg/day to 400 mg/day.