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WO2000052034A2 - Inhibiteurs d'activite de serine protease, methodes et compositions de traitement d'infections virales - Google Patents

Inhibiteurs d'activite de serine protease, methodes et compositions de traitement d'infections virales Download PDF

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Publication number
WO2000052034A2
WO2000052034A2 PCT/US2000/005558 US0005558W WO0052034A2 WO 2000052034 A2 WO2000052034 A2 WO 2000052034A2 US 0005558 W US0005558 W US 0005558W WO 0052034 A2 WO0052034 A2 WO 0052034A2
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Prior art keywords
carbonyl
methylpropyl
oxadiazolyl
aat
benzyloxycarbonyl
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PCT/US2000/005558
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English (en)
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WO2000052034A3 (fr
Inventor
Leland Shapiro
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The Trustees Of University Technology Corporation
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Priority to AU37191/00A priority Critical patent/AU3719100A/en
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Publication of WO2000052034A3 publication Critical patent/WO2000052034A3/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/55—Protease inhibitors
    • A61K38/57—Protease inhibitors from animals; from humans
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
    • A61K31/401—Proline; Derivatives thereof, e.g. captopril
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7042—Compounds having saccharide radicals and heterocyclic rings
    • A61K31/7052—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides
    • A61K31/706—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom
    • A61K31/7064—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines
    • A61K31/7068—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines having oxo groups directly attached to the pyrimidine ring, e.g. cytidine, cytidylic acid
    • A61K31/7072—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines having oxo groups directly attached to the pyrimidine ring, e.g. cytidine, cytidylic acid having two oxo groups directly attached to the pyrimidine ring, e.g. uridine, uridylic acid, thymidine, zidovudine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals

Definitions

  • the present invention relates to enzyme inhibitors and their respective ligands. More particularly, the present invention relates to substances exhibiting inhibitory activity toward retroviral replication and spread, which are facilitated or mediated by serine protease activity.
  • Serine proteases serve an important role in human physiology by mediating the activation of vital functions. In addition to their normal physiological function, serine proteases have been implicated in a number of pathological conditions in humans . Serine proteases are characterized by a catalytic triad consisting of aspartic acid, histidine and serine (Asp-His-Ser) at the active site. The naturally occurring serine protease inhibitors are usually, but not always, polypeptides and proteins which have been classified into families primarily on the basis of the disulfide bonding pattern and the sequence homology of the reactive site.
  • Serine protease inhibitors have been found in microbes, in the tissues and fluids of plants, animals, insects and other organisms. Protease inhibitor activities were first discovered in human plasma by Fermi and Pernossi in 1894. At least nine separate, well-characterized proteins are now identified, which share the ability to inhibit the activity of various proteases .
  • alpha- 1 -proteinase inhibitor Several of the inhibitors have been grouped together, namely alpha- 1 -proteinase inhibitor, antithrombin III, antichymotrypsin, C 1 -inhibitor, eglin, and alpha-2-antiplasmin, which are directed against various serine proteases, i.e., leukocyte elastase, thrombin, cathepsin G, chymotrypsin, plasminogen activators, and plasmin. These are referred to as the alpha- 1 -proteinase inhibitor class.
  • the protein alpha-2-macroglobulin inhibits members of all four catalytic classes: serine, cysteine, aspartic, and metalloproteases.
  • alpha- 1 -proteinase inhibitor also known as ⁇ ,-antitrypsin or AAT
  • inter-alpha-trypsin inhibitor inhibit only serine proteases
  • alpha- 1 -cysteine protease inhibitor inhibits cysteine proteases
  • alpha- 1-anticollagenase inhibits collagenolytic enzymes of the metalloenzyme class.
  • AAT is a glycoprotein of MW 51,000 with 394 amino acids and 3 oligosaccharide side chains. Human AAT was named anti-trypsin because of its initially discovered ability to inactivate pancreatic trypsin. Human NAT is a single polypeptide chain with no internal disulfide bonds and only a single cysteine residue normally intermolecularly disulfide-linked to either cysteine or glutathione . The reactive site at position 358 of AAT contains a methionine residue, which is labile to oxidation upon exposure to tobacco smoke or other oxidizing pollutants.
  • AAT Such oxidation may reduce the biological activity of AAT; therefore substitution of another amino acid at that position, 1 e alanme, valine, glycine, phenylalanine, arginine or lysine, produces a form of AAT which is more stable AAT can be represented by the following formula MPSSNSWGILLLAGLCCLNPNSLAEDPQGDAAQKTDTSHHDQDHPTFNKITPNLAEFAFSLYRQL ASTNIFFSPVSIATAFAMLSLGTKADTHDEILEGLNFNLTEIPEAQIHEGFQELLRTLNQPDSQLQL TTGNGLFLSEGLKLNDKFLEDVKKLYHSEAFTVNFGDTEEAKKQLNDYNEKGTQGKINDLNKEL
  • AAT human antitrypsin
  • ATIII antithrombm
  • ACT antichymotrypsin
  • C 1 -inhibitor C 1 -inhibitor
  • tP A-inhibitor mouse AT, mouse contrapsin, barley protein Z, and ovalbumm
  • ACT antichymotrypsin
  • tP tP A-inhibitor
  • mouse AT mouse contrapsin
  • barley protein Z and ovalbumm
  • Alpha- 1 -antitrypsin known to be an acute phase protem in humans, is augmented in autoimmune diseases such as systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), mixed connective tissue disease (MCTD), Sjogren syndrome, sclero
  • enzymes such as trypsin, chymotrypsin, cathepsin G, plasmin, thrombin, tissue kalhkrem, and factor Xa can also serve as substrates
  • SEC se ⁇ in-enzyme complex
  • elastase is a protease which causes degradation and fragmentation of elastic fibers as a result of its proteolytic activity on rubber-like elastin
  • Other connective tissue proteins such as type I, III, and IN collagens, the protein portion of proteogly
  • NAT is one of few naturally-occurring mammalian serine protease inhibitors clinically approved for the therapy of protease imbalance Therapeutic AAT became commercially available m the mid
  • PROLASTIN® is a trademark for a purified variant of AAT, is currently sold by Bayer Company (U S Pat No 5,610,285 Lebing et al , March 11, 1997) Recombinant unmodified and mutant variants of AAT produced by genetic engineering methods from transformed cells are also known (U S Pat No 4,711, 848), methods of delivery are also known, e g , AAT gene therapy/delivery (U S Pat No 5,399,346 to French Anderson et al )
  • HIV Human immunodeficiency virus
  • protease activity required for the cleavage of gag-pol precursor proteins This enzymatic activity is similar to activity of renin - aspartyl protease produced by the kidney
  • the close relationship between renin and HIN encoded protease led to an accelerated generation of specific HIN-1 protease inhibitors as effective agents in treatment of AIDS (Scha ⁇ e, et al , "Proteases and their inhibitors today and tomorrow", Biochimie, 73(1) 121-6 (1991)
  • HIN protease have been developed as a consequence and used successfully in AIDS patients
  • indinavir and c ⁇ xivan are aspartyl protease inhibitors, which inhibit cleavage of pre-protein of HIN by viral own protease and thereby suppress viral proliferation
  • Lezdey et al disclose the method of using AAT, Secretory Leukocyte Protease Inhibitor (SLPI), and alpha antichymotrypsin (AAC) for inhibition of proliferation of a variety of viruses that require gag-pol cleavage
  • AAT, SLPI, and AAC generally known as serine protease inhibitors, inhibit such viruses by binding to viral or cellular aspartic protease While it is unknown whether this mechanism may take place in such circumstances, several lines of evidence exist, which indicate that serine protease inhibitors may interfere with viral replication through inhibition of host's serme proteases but not HIV encoded aspartyl protease
  • serine protease inhibitors may interfere with viral replication through
  • a neutralizing epitope of HIV on external envelope glycoprotem was found to have homologous sequences to lnter-alpha-trypsin inhibitor (ITI) Human urinary trypsin inhibitor (UTI), a protein indistinguishable from ITI, as well as synthetic peptides including epitope beta inhibited syncytium formation caused by the HIV-infected CCRF-CEM and umnfected Molt-4 cells m a dose-dependent manner (0 1-1 mM)
  • SLPI secretory leukocyte protease inhibitor
  • a cathepsin G-like proteinase at the surface of U-937 cells reacting with the V3 loop of HIV-1 gpl20 was reported by Av ⁇ l et al , (Av ⁇ l, et al , "Identification of the U-937 membrane-associated proteinase interacting with the V3 loop of HIV-1 gpl 20 as cathepsin G", FEBS Lett, 345(1) 81-6) (1994)
  • At least five separate T lymphocyte-derived enzymes were identified by Harvima et al , (Harvima et al , "Separation and partial characterization of proteinases with substrate specificity for basic ammo acids from human MOLT-4 T lymphocytes identification of those inhibited by var ⁇ able-loop-V3 peptides of HIV-1 (human immunodeficiency virus- 1) envelope glycoprotem", Biochem J, 292 ( Pt 3) 711-8) (1993)
  • T lymphocyte associated elastase was reported by B ⁇ stow et al , as a protease involved in HIV infection and synthetic elastase inhibitors MAAPVCK but not FLGFL were shown to interfere with HIV infection (B ⁇ stow, et al , "Inhibition of HIV-1 by modification of a host membrane protease” , Int Immunol, 7(2) 239-49) (1995)
  • PC6A and PC6B isomers were also proposed as gpl 60 processing enzymes (Miranda et al , "Isolation of the human PC6 gene encoding the putative host protease for HIV-1 gpl 60 processing in CD4+ T lymphocytes", Proc Natl Acad Sci U S A, 93(15) 7695-700) (1996)
  • V3 loop of gp 120 was found to be homologous with trypstatin and peptides mimicking V3 region were found to inhibit HIV infection (Cox et al , "Synergistic combinations and peptides in the inhibition of human immunodeficiency virus", Adv Enzyme Regul, (31 85-97) (1991)
  • serine proteases as HIV facilitating enzymes has gradually increased and today in addition to TL2 it includes an assortment of enzymes including furin, kexin, convertase, cathepsin G, subtihsm, subtilisin-hke proteases, tryptase M, acrosin, PACE4, PC5/6-B, PCI, VEM, etc
  • HIV-1 (Ohnishi et al , "A funn-defective cell line is able to process correctly the gpl 60 of human immunodeficiency virus type 1", J Virol, 68(6) 4075-9 9 (1994), Gu et al , "Furm is important but not essential for the proteolytic maturation ofgpl60 of HIV-1", FEBS Lett, 365(1) 95-7) (1995), Inocencio et al , “Endoprotease activities other than furm and PACE4 with a role n processing of HIV-I gpl 60 glycoproteins n CHO-K1 cells", J Biol Chem, 272(2) 1344-8) (1997)
  • AAT was positive in 40% of HIV-positive patients with cryptospo ⁇ dial infections and none of 12 HIV-positive patients with non-cryptospo ⁇ dial diarrhea (Lima et al , "Mucosal injury and disruption of intestinal barrier function in HlV-mfected individuals with and without diarrhea and cryptosporidiosis in northeast Brazil", Am J Gastroenterol, 92(10) 1861-6) (1997) Serum concentrations of a tumor-associated trypsin inhibitor
  • TATI Tumor-associated trypsin inhibitor n induced and acquired immunodeficiency Studies on transplanted and HIV- infected patients
  • the incidence of abnormal AAT phenotypes was 16 3% in the homosexual group which was significantly different (p less than 0 03) than the 8 7% in the heterosexual group
  • Faecal alphal antitrypsin concentration were reflective of abnormal pancreatic function of paediat ⁇ c HIV infection (Carroccio et al , "Pancreatic dysfunction and its association with fat malabsorption in HIV infected
  • the present invention offers useful insight into therapy and pathogenesis of viral infection
  • a method of treating viral infection facilitated or mediated by a serine proteolytic (SP) activity comprising administering to a subject suffering or about to suffer from said viral infection a therapeutically effective amount of a compound having a serine protease inhibitory or se ⁇ in activity comprising o j -antitrypsin activity (ANT)
  • the viral infection may include retroviral infection such as human immunodeficiency virus (HIV) infection
  • a method of preventing or inhibiting delivery of viral nucleic acid into the nucleus of a mammalian host as well as a method of preventing or inhibiting the exit of a vi ⁇ on particle from a mammalian host harboring an agent of a viral infection is provided
  • these processes are mediated by endogenous host serine protease (SP) or SP-like activity and will be counteracted by administering a pharmacologically effective amount of a substance exhibiting mammalian o , -antitrypsin (NAT) or AAT-hke activity
  • the post-exposure prophylaxis is contemplated in order to block establishment of productive infection m a mammal exposed to HTV-contaminated fluids such as blood, saliva, semen, sweat, urine, vaginal secretion, tears, and other body fluids that may contain HIV either in cell-free form or m cell-associated from It also understood that said method is effective in preventing mother-to-child HIV transmission during pregnancy According to this method pharmac
  • AAT substantially pu ⁇ fied natural or recombinant AAT AAT and similarly active compounds may be identified by a senes of assays wherein a compound (AAT) will exhibit inhibitory activity versus control in an assay
  • AAT comprises blocking interleukin- 18 or IL-18- induced human immunodeficiency virus (HIV) production in Ul monocytic cells
  • Other assays involve blocking stimulants such as IL-6, ⁇ aCl, LPS, T ⁇ F. and other HIV stimulants known in the art
  • Other assays involve MAGI-CCR-5 cell assay and PBMC assay as described in detail in the body of the disclosure
  • FVFLM SEQUENCE ID NO 1
  • FVFAM SEQUENCE ID NO 2
  • FVALM SEQUENCE ID NO 3
  • FVFLA FVFLA
  • FLVFI SEQUENCE ID NO 5
  • FLMII SEQUENCE ID NO 6
  • FLFVL SEQUENCE ID NO 1
  • FLFW SEQUENCE ID NO 8
  • FLFLI SEQUENCE ID NO 9
  • FLFFI SEQUENCE ID NO 10
  • FLMFI SEQUENCE ID NO 11
  • FMLLI SEQUENCE ID NO 12
  • FIIMI SEQUENCE ID NO. 13
  • FLFCI SEQUENCE ID NO. 14
  • FLFAV SEQUENCE ID NO. 15
  • FVYLI SEQUENCE ID NO. 16
  • FAFLM SEQUENCE ID NO. 17
  • AVFLM SEQUENCE ID NO. 18
  • pentapeptides can be represented by a general formula (I): I-A-B-C-D-E-F-G-H-II, wherein I is Cys or absent; A is Ala, Gly, Val or absent; B is Ala, Gly, Val, Ser or absent; C is Ser, Thr or absent; D is Ser, Thr, Ans, Glu, Arg, He, Leu or absent; E is Ser, Thr, Asp or absent; F is Thr, Ser, Asn, Gin, Lys, T ⁇ or absent; G is Tyr or absent; H is Thr, Gly, Met, Met(O), Cys, Thr or Gly; and II is Cys, an amide group, substituted amide group, an ester group or absent, wherein the peptides comprise at least 4 amino acids and physiologically acceptable salts thereof.
  • I is Cys or absent
  • A is Ala, Gly, Val or absent
  • B is Ala, Gly, Val, Ser or absent
  • C is Ser,
  • the peptides of interest are homologous and analogous peptides. While homologues are natural peptides with sequence homology, analogues will be peptidyl derivatives, e.g., aldehyde or ketone derivatives of such peptides . Typical examples of analogues are oxadiazole, thiadiazole and triazole peptoids . Without limiting to AAT and peptide derivatives of NAT, compounds such as oxadiazole, thiadiazole and triazole peptoids are preferred.
  • the preferred doses for administration can be anywhere in a range between about 10 ng and about 10 mg per ml of biologic fluid of treated patient.
  • the therapeutically effective amount of AAT peptides or drugs that have similar activities as AAT or peptide drug can be also measured in molar concentrations and may range between about InM and about 1 mM per ml of biologic fluid of treated patient.
  • compositions with serine protease inhibiting activity comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of at least one of the compounds of the present invention or a pharmaceutically acceptable salt or prodrug form thereof.
  • viral infections are contemplated to be treated wherein such viral infections are caused/facilitated by a deficiency in AAT levels or by a dysfunction of AAT.
  • Clinical conditions and viral infections resulting from uncontrolled serine protease activity are also within the scope of the present invention and will be treated alike.
  • a general method of treating a mammal suffering from a pathological condition that is mediated by endogenous serine protease (SP) or SP-like activity comprises administering a therapeutically effective amount of a substance exhibiting mammalian ⁇ , -antitrypsin (AAT) or AAT-like activity.
  • This pathological condition can be caused at least in part by a viral infection.
  • a compound of choice may be one that inhibits proteinase-3, cathepsin G, or elastase.
  • a novel class of chemical compounds that are capable of inhibiting and/or blocking the activity of the serine protease(s), which halts the proliferation of a virus including HIV, pharmaceutical compositions containing these compounds, novel intermediates for compounds which inhibit and block the activity of the HIV facilitating serine protease, novel methods for making such compounds, and use of the compounds as inhibitors of the HIV.
  • a method which consists of treating an individual having a physiological condition caused, in whole or part, by virus shedding.
  • a method of inhibiting virus release wherein the target of the therapy is a cell and one will contact such cell with an effective amount of a compound having AAT activity.
  • reverse transcriptase inhibitor can be selected from a group including nucleoside RT inhibitors: Retrovir (AZT/ zidovudine; Glaxo Wellcome); Combivir (Glaxo Wellcome); Epivir (3TC, lamivudine; Glaxo Wellcome); Videx (ddl/didanosine; Bristol-
  • NRTIs Non-nucleoside reverse transcriptase inhibitors
  • the pharmaceutical composition may be a peptide or a small molecule, which exhibits AAT or AAT-like activity.
  • AAT ⁇ , -antitrypsin
  • AAT-like activity is another object of this invention.
  • Yet another preferred embodiment of this invention is to provide cci -antitrypsin (AAT) or a compound with AAT-hke activity for types of cancer that may or may not be virus induced but are capable of metastasizing due to SP activity
  • Such tumors may comprise fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogemc sarcoma, chordoma, angiosarcoma, endothehosarcoma, lymphangiosarcoma, lymphangioendothehosarcoma, synovioma, mesothehoma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, rhabdosarcoma, colorectal carcinoma, pancreatic cancer, breast cancer, ovarian cancer,
  • FIG 1 illustrates the effect of AAT on HTV production m PBMC as performed without pre-mcubation
  • FIG 2 illustrates the effect of AAT on HIV production in PBMC as performed with pre-incubation
  • FIG 3 illustrates the effect of AAT on HTV production in MAGI cells
  • FIG 4 illustrates the effect of FVYLI (SEQUENCE ID NO 16) peptide on HIV production in MAGI cells
  • FIG 5 illustrates the effect of AAT on HIV production rn Ul cells upon induction with IL-18
  • FIG 6 illustrates the lack of effect of Prolastin on HIV production in Ul cells upon induction with IL- 18
  • FIG 7 illustrates the effect of AAT on HIV production m Ul cells upon induction with IL-6
  • FIG 8 illustrates the effect of AAT on HTV production in Ul cells upon induction with TNF
  • FIG 9 illustrates the effect of AAT on HIV production in Ul cells upon induction with LPS
  • FIG 10 illustrates the effect of AAT on HIV production in Ul cells upon induction with NaCl
  • FIG 11 illustrates the effect of AAT-mimicking drug on HIV production in Ul cells upon induction with IL-18
  • FIG 12 illustrates the effect of AAT on viability and number of Ul cells
  • FIG 13 illustrates the p24 antigen output of HIV when grown m normal or AAT-deficient whole blood
  • FIG 14 illustrates the effect of AAT and AAT-mimickmg drug (CE 2072) in reducmg IL-18- ⁇ nduced NF- ⁇ B activation
  • the present invention provides a method of treatment that is totally opposite to the prevailing therapeutic approaches and provides enzyme antagonist instead of a trypsin-hke enzyme Throughout this application various publications and patents are referenced The disclosures of these publications and patents in their entireties are hereby inco ⁇ orated by reference into this application in order to more fully describe the state of the art to which this invention pertains
  • AAT preparations of the instant invention can be obtained by a variety of methods
  • U S Pat Nos 5,529,920 to Cole et al , and 5,665,589 Harris et al disclose hver-de ⁇ ved cell lines producing AAT U S Pat Nos 5,861,299 to Archibald et al , 5,780,009 Karatzas et al , and 5,476,995 to Clark et al , disclose methods of obtaining transgenic AAT from the milk of transgenic mammals (cows, goats, sheep, pigs, etc ) as a source of AAT
  • U S Pat Nos 4,839,283 to Kawasaki et al , 5,593,858 to Fleer et al , 5,641,670 to Treco et al , and 5,565,334 to Kufe et al disclose various recombinant methods of wild type AAT production (wild type refers to AAT which is essentially identical to one found in human plasma)
  • the peptide-based serine protease inhibitors may be prepared by any suitable synthesis method such as originally described by Mer ⁇ field, J Am Chem Soc Vol 85, p 2149 (1963) Synthetic peptides, which exhibit inhibitory activity toward serine proteases and methods for prepa ⁇ ng and using the same are disclosed for example in U S Pat Nos 4,829,052, 5,157,019 to Glover, U S Pat No 5,420,110 to Miller, U S Pat No 4,963,654 to Katunuma, and inco ⁇ orated herein by reference Those skilled in the art of biochemical synthesis will recognize that for commercial-scale quantities of peptides, such peptides can also be prepared usmg recombinant DNA techniques
  • the compounds of the present mvention are used as therapeutic agents in the treatment of a physiological (especially pathological) condition caused in whole or part, by uncontrolled serine protease activity
  • the peptides may be administered as free peptides or pharmaceutically acceptable salts thereof
  • the terms used herein conform to those found in Budava ⁇ , Susan (Editor), "The Merck Index" An
  • pharmaceutically acceptable salt refers to those acid addition salts or metal complexes of the peptides which do not sigmficantly or adversely affect the therapeutic properties (e g efficacy, toxicity, etc ) of the peptides
  • the peptides should be administered to individuals m need thereof as a pharmaceutical composition, which, in most cases, will comp ⁇ se the peptide and/or pharmaceutical salts thereof with a pharmaceutically acceptable carrier
  • pharmaceutically acceptable carrier refers to those solid and liquid carriers, which do not significantly or adversely affect the therapeutic properties of the peptides
  • the pharmaceutical compositions containing peptides of the present invention may be administered to individuals, particularly humans, either intravenously, subcutaneously, intramuscularly, mtranasally or even orally
  • the necessary dosage will vary with the particular condition bemg treated, method of administration and rate of clearance of the peptide from the body In most cases dosages between 0
  • Routes of administration include, but are not limited to, topical, transdermal, parenteral, gastrointestinal, transbronchial and transalveolar Topical administration is accomplished via a topically applied cream, lotion, emulsion, gel, suppository, pessary, tablet, sachet, spray, rmse, etc containing therapeutically effective amounts of se ⁇ ins Topical routes are vaginal, rectal, nasal, sub ngual, ocular, etc , and which are either transmucosal or transdermal means of delivery
  • Transdermal administration is accomplished by application of a cream, rinse, gel, etc capable of allowing the se ⁇ ins to penetrate the skin and enter the blood stream
  • Parenteral routes of administration include, but are not limited to, direct injection such as intravenous, intramuscular, intraperitoneal or subcutaneous injection
  • Gastrointestinal routes of administration mclude, but are not limited to, mgestion and rectal
  • Transbronchial and transalveolar routes of administration include, but are not
  • the compounds described herein and/or their derivatives may be administered as the pure chemicals, it is preferable to present the active ingredient as a pharmaceutical composition
  • the invention thus further provides the use of a pharmaceutical composition comp ⁇ sing one or more compounds and/or a pharmaceutically acceptable salt thereof, together with one or more pharmaceutically acceptable carriers therefor and, optionally, other therapeutic and or prophylactic ingredients
  • the car ⁇ er(s) must be acceptable in the sense of being compatible with the other ingredients of the composition and not deleterious to the recipient thereof
  • compositions include those suitable for oral or parenteral (including intramuscular, subcutaneous and intravenous) administration
  • the compositions may, where appropriate, be conveniently presented in discrete unit dosage forms and may be prepared by any of the methods well known m the art of pharmacy Such methods include the step of b ⁇ nging into association the active compound with liquid carriers, solid matrices, semi-sohd carriers, finely divided solid carriers or combination thereof, and then, if necessary, shaping the product into the desired delivery system
  • compositions suitable for oral administration may be presented as discrete unit dosage forms such as hard or soft gelatin capsules, cachets or tablets, each containing a predetermined amount of the active ingredient, as a powder or as granules, as a solution, a suspension or as an emulsion
  • the active ingredient may also be presented as a bolus, electuary or paste
  • Tablets and capsules for oral administration may contain conventional excipients such as binding agents, fillers, lubricants, disintegrants, or wetting agents
  • the tablets may be coated according to methods well known in the art , e g , with enteric coatings
  • Oral liquid preparations may be in the form of, for example, aqueous or oily suspension, solutions, emulsions, syrups or elixirs, or may be presented as a dry product for constitution with water or another suitable vehicle before use
  • Such liquid preparations may contain conventional additives such as suspendmg agents, emulsifying agents, non-aqueous vehicles (which may include edible oils), or preservative
  • the compounds may also be formulated for parenteral administration (e g , by injection, for example, bolus injection or continuous infusion) and may be presented in unit dose form in ampoules, pre- filled syringes, small bolus infusion contamers or m multi-does containers with an added preservative
  • parenteral administration e g , by injection, for example, bolus injection or continuous infusion
  • the compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizmg and/or dispersing agents
  • the active ingredient may be in powder form, obtained by aseptic isolation of sterile solid or by lyophi zation from solution, for constitution with a suitable vehicle, e g , sterile, pyrogen-free water, before use
  • the compounds may be formulated as ointments, creams or lotions, or as the active ingredient of a transdermal patch Suitable transdermal delivery systems are disclosed, for example, in Fisher et al (U S Pat No 4,788,603) or Bawas et al (U S Pat Nos
  • Ointments and creams may, for example, be formulated with an aqueous or oily base with the addition of suitable thickemng and/or gelling agents
  • Lotions may be formulated with an aqueous or oily base and will in general also contain one or more emulsifying agents, stabilizing agents, dispersing agents, suspending agents, thickening agents, or coloring agents
  • the active ingredient can also be delivered via iontophoresis, e g , as disclosed in U S Pat Nos 4, 140, 122,
  • compositions suitable for topical administration in the mouth include unit dosage forms such as lozenges comprising active ingredient in a flavored base, usually sucrose and acacia or tragacanth, pastilles comprising the active ingredient in an inert base such as gelatin and glycerin or sucrose and acacia, mucoadherent gels, and mouthwashes comp ⁇ sing the active ingredient in a suitable liquid carrier
  • the delivery system can be in slow release sustained form known in the art
  • the above-described compositions can be adapted to provide sustained release of the active ingredient employed, e g , by combination thereof with certain hydrophilic polymer matrices, e g , comprising natural gels,
  • the amount of the compound, or an active salt or derivative thereof, required for use in treatment will vary not only with the particular salt selected but also with the route of administration, the nature of the condition being treated and the age and condition of the patient and will be ultimately at the discretion of the attendant physician or clinician
  • the compound is conveniently administered in unit dosage form, for example, containing 5 to 2000 mg, conveniently 10 to 1000 mg, most conveniently, 50 to 500 mg of active ingredient per unit dosage form
  • the active ingredient should be administered to achieve peak plasma concentrations of the active compound of from about 100 ng to 10 mg, preferably, about 1 microgram to 5mg most preferably, about 2 to about 4 mg per ml of plasma fluid This may be achieved, for example, by the intravenous injection of a 0 05 to 5% solution of the active ingredient, optionally in saline, or orally administered as a bolus contammg about 10-1000 mg of the active ingredient Desirable blood levels may be maintamed by continuous infusion to provide about 0 01-5 0 mg/kg/hr or by intermittent infusions containing about 0 4- 20 mg/kg of the active ⁇ ngred ⁇ ent(s) Buffers, preservatives, antioxidants and the like can be mco ⁇ orated as required
  • the desired dose may conveniently be presented m a single dose or as divided doses administered at appropriate mtervals, for example, as two, three, four or more sub-doses per day
  • the sub-dose itself may be further divided, e g , into a number of discrete loosely spaced administrations, such as multiple inhalations from an insufflator or by application of a plurality of drops into the eye
  • Alpha-1-ant ⁇ tryps ⁇ n (AAT) used in these studies is purified from the blood of healthy volunteers AAT is purified to single-band homogeneity
  • the AAT protein is diafiltered into a diluent consistmg of NaCl, sodium phosphate, pH 7 05
  • the AAT preparations are maintained at stock concentrations of 14-
  • IL human interleukin
  • Vertex Pharmaceuticals Inc (Cambridge, MA)
  • IL-6 and tumor necrosis factor (TNF) are obtained from R & D Systems, Minneapolis, MN
  • endotoxin-free NaCl and endotoxm (lipopolysacchande, LPS) is obtained from Sigma (St Louis, MO)
  • RPMI 1640 medium purchased from Mediatech (Herndon, VA) containing 2 5 mM L-glutamme, 25 mM Hepes, 100 units/ml penicillin and streptomycin (GIBCO/BRL, Rockville, MD) with 10% or 7 5% (vol/vol) heat-inactivated fetal bovine serum (FBS, GIBCO) for Ul cell and MAGI-CCR5 cell cultures, respectively
  • PBMC are cultured in R3 medium consistmg of RPMI 1640 medium (Mediatech), 20% FBS (GIBCO), 100 units/ml penicillin and streptomycin (GIBCO) and 5% (vol/vol) IL-2 (Hemagen, Waltham, MA) Ul monocytic cell assay Ul cells are obtained from the AIDS Research and Reference Reagent
  • NIH Ul cells are maintained in T-175 polystyrene flasks (Falcon, Becton Dickinson, Franklin Lakes, NJ) in medium and used when in log phase growth Cells are counted in a hemacytometer, examined for viability by Trypan blue exclusion (> 95% for all experiments) and resuspended in fresh medium at 2 X 10 6 per ml
  • Two-hundred fifty ml of cell suspension are added to wells of 24-well polystyrene tissue culture plates (Falcon), followed by the addition of medium or NAT to produce the final concentration to be tested m a volume of 450 ml
  • 50 ml of medium (control) or stimulus diluted in medium are added to wells to produce the final concentration of stimulus to be tested
  • the final culture volumes are 500 ml and contained 1 X 10 6 cells per ml
  • PBMC Peripheral Blood Mononuclear Cells
  • PBMC from HIV-1 negative healthy subjects are isolated from hepa ⁇ nized blood by Ficoll-Hypaque density-gradient centnfugation
  • concentration of PBMC in aliquots are counted using a hemacytometer (viability > 95% by trypan blue exclusion for each experiment) and PBMC are diluted at 1 X 10 6 per ml in R3 medium supplemented with additional 5% (vol/vol) IL-2 and 3 3 mg/ml phytohemagglutinin (PHA, Sigma)
  • PHA phytohemagglutinin
  • Cell suspensions are then incubated for 2 days (37 °C, 5% CO 2 ) in T-175 polystyrene tissue culture flasks (Falcon)
  • the stocks of lymphocyte-tropic HIV-1 strain A018A are titered by standard protocol and are used to infect PBMC Following the 2 days of incubation, PBMC from each donor are removed from tissue culture flasks, divided into 2
  • PBMC suspension 10 6 cells per ml
  • a separate 250 ml aliquot of PBMC suspension is added to a 1 5 ml polypropylene microfuge tube (Fisher) along with 200 ml R3 medium and 50 ml of 10% (vol/vol) Tnton- X-100 (Fisher)
  • This sample is frozen and designated time 0 Cultures in 24-well plates are incubated for 4 days, after which T ⁇ ton-X-100 (Fisher) is added (final concentration of 1% vol/vol as described above for Ul cell cultures) and plates frozen and thawed once Corresponding time 0 samples are thawed with each plate and cell lysates assayed for p24 antigen by ELISA MAGI-CCR5 cell assay
  • the MAGI (Multmuclear Activation of a Galactosidase Ind ⁇ cator)-CCR-5 cell line is a clone de ⁇ ved from the HeLa cell line that expresses high levels of CD4 It has been transfected with a single integrated copy of a galactosidase gene under control of the HIV-1 long terminal repeat Beta- galactosidase is expressed upon production of HIV-1 Tat protein following one round of HIV-1 replication within the cell
  • the MAGI-CCR-5 cell line is derived from MAGI cells into which the CCR-5 HIV-1 co- receptor gene has been inco ⁇ orated These cells constitute an assay for early infection events and can be infected with either lymphocyte-tropic or macrophage-tropic HTV-1 strains MAGI-CCR-5 cells are obtained from the AIDS Research and Reference Reagent Program, National Institute of Allergy and
  • NLH Cells are cultured m polystyrene T-175 flasks (Falcon) in medium until cells are noted to be in log growth phase Cells are then resuspended in fresh medium and ahquoted into 24- well polystyrene plates (Falcon) at 4 X 10 4 cells per well (1ml total volume) After 24 hr incubation adherent cells are 30-40% confluent and all medium is removed Two hundred ml of fresh medium is then added to each well without (negative control) or with AAT and incubated for 1 hour AAT diluent is added to a separate well at a volume equivalent to that of the highest concentration of AAT tested (control) One hundred thirty TCID 50 of HIV-1 and DEAE dextran in medium are added to each well T-cell tropic HIV-1 strain A018A is used After 2 hr incubation, medium is added to each well to adjust the final volume of each well to 500 ml.
  • Cultures are incubated for 48 hr, which allows infection of the MAGI- CCR-5 cells.
  • Medium is aspirated and the cells fixed for 5.0 min at room temperature by adding 1.0 ml of a 1% formaldehyde/0.2 % glutaraldehyde solution in phosphate buffered saline (PBS). Fixing solution is then aspirated and cells washed with PBS. This is followed by addition of galactosidase staining solution. Fifty min of incubation is followed by a blinded optical count of pigmented cells under a microscope.
  • PBS phosphate buffered saline
  • AAT inhibits production of HIV-1 in Ul cell cultures.
  • the Ul cell line is derived from human monocytic U937 cells into which 2 copies of HIV-1 provirus are inco ⁇ orated into host genome. Exposing Ul cells to pro-inflammatory cytokines such as IL- 18, IL-1, IL-6 and TNF, phorbol esters or hyperosmolarity results in the induction of HIV-1 as assessed by p24 antigen. Stimulation of Ul cells with 0.5 nM IL-18 induced large amounts of p24 antigen after 48 hr of incubation in 3 separate experiments.
  • the percent reductions observed compared to stimulation with IL-18 alone are 65 ⁇ 1.8, 93 ⁇ 3.0 and 98 ⁇ 1%, respectively.
  • the mean p24 antigen measured in Ul cells cultured in medium alone (control) is 1,207 ⁇ 361 pg/ml (Fig. 7).
  • Stimulation with 100 ng/ml IL-6 results in a 3.6-fold increase in p24 antigen production, to 4,337 ⁇ 2,006 pg/ml.
  • Stimulation with IL- 6 in the presence of AAT results in dose-dependent inhibition of p24 production compared to that measured in the absence of AAT.
  • the measured P24 antigen values are 6,228 ⁇ 2,129, 3,992 ⁇ 1,987, 3,850 ⁇ 1,943, 2,597 ⁇ 1,253, 2,155 ⁇ 1,085, 1,838 ⁇ 881 and 1,213 ⁇ 658 pg/ml, respectively.
  • the corresponding mean percent reductions for NAT additions of 3, 4 and 5 mg/ml are 80, 88 and 100%, respectively.
  • T ⁇ F-induced p24 antigen reduced T ⁇ F-induced p24 antigen to 16,405 ⁇ 8,449, 16,863 ⁇ 7,718, 15,328 ⁇ 7,129, 12,566 ⁇ 4,981, 9,341 ⁇ 2,730, 9,091 ⁇ 3,436 and 6,868 ⁇ 2,737, respectively.
  • the mean percent reductions in T ⁇ F-induced p24 antigen observed in the presence of 3, 4, and 5 mg/ml AAT are 56, 60, and 73%, respectively.
  • LPS is a cell wall component of gram-negative bacteria with several pro-inflammatory activities.
  • Ul cells cultured in the presence of 500 ng/ml LPS for 48 hrs contained 1,427 ⁇ 39 pg/ml p24 antigen, as shown in Figure 9. This represents a mean 3 -fold increase compared to p24 produced in control (medium alone) cultures, where 476 ⁇ 76 pg/ml p24 antigen was measured.
  • the mean p24 antigen measured in ⁇ aCl-stimulated and control cultures are 7,511 ⁇ 707 and 295 ⁇ 29 pg/ml, respectively.
  • AAT resulted in mean p24 levels of 11,054 ⁇ 3,231, 7,363 ⁇ 485, 5,657 ⁇ 48, 2,83 8 ⁇ 466, 1,919 ⁇ 594, 425 ⁇ 32 and 266 ⁇ 26pg/ml, respectively
  • ANT added at 3 0, 4 0 and 5 0 mg/ml the corresponding percent inhibitions are 76, 98 3 and 100% (Fig 10)
  • AAT inhibits p24 antigen production m HIV-1 -infected PBMC
  • AAT inhibits early infection-associated events in MAGI-CCR5 cells exposed to HIV-1
  • the MAGI-CCR-5 cell assay evaluates early events in the HTV-1 infection process These events include cell-surface binding and internalization, uncoatmg, reverse transcription and translation, protein processing and Tat activity Binding of the tat protein to a reporter construct withm the MAGI-CCR-5 cells enables quantification of these early HIV-1 events
  • MAGI-CCR-5 cells are infected with A018A strain of HIV-1 as described supra
  • a mean positive cell count of 2 3 is obtained
  • an increase in mean positive cell count is observed, to 72 ⁇ 13 (31-fold increase, P ⁇ 0 001)
  • MAGI-CCR-5 cells exposed to HIV-1 and cultured with added AAT demonstrate significant and dose- dependent inhibition of positive cell counts
  • Addition of 0 1, 1 0, 2 0, 3 0, 4 0 and 5 0 mg/ml ANT resulted in mean positive cell counts of 74 ⁇ 13, 75 ⁇ 17, 56 ⁇
  • MAGI-CCR-5 cell early infection events Compared to cultures containing HIV-1 alone, significant inhibition of MAGI-CCR-5 cell early infection events is significant for AAT concentrations of 2.0, 3.0, 4.0 and 5.0 mg/ml. These values correspond to 23, 41, 66 and 76% inhibition.
  • AAT concentrations 2.0, 3.0, 4.0 and 5.0 mg/ml. These values correspond to 23, 41, 66 and 76% inhibition.
  • MAGI-CCR-5 cells are exposed to virus and a diluent volume equivalent to that of AAT solution added to 5.0 mg/ml cultures. Cultures containing diluent produced a positive cell count of 72 ⁇ 16, which is not significantly different from cultures containing HTV-1 alone (+HTV), as shown on the horizontal axis.
  • Example 3 Failure of commercial AAT preparation (Prolastin) to inhibit HIV Prolastin used as a control preparation of AAT in the experimental setting that is similar to those described above.
  • this preparation fails to display anti-HIV activity at doses that are comparable to the composition of the invention (Fig. 6).
  • the lack of the activity cannot be explained by low levels of active AAT since Prolastin contains only about 8% of inactive form of total antitrypsin (Lomas DA, Elliott PR, Carrell RW.
  • Commercial plasma alphal -antitrypsin (Prolastin) contains a conformationally inactive, latent component. Eur Respir J 1997 Mar;10(3):672-5). The biological significance of this observation is unknown. However, this means that not every AAT composition is inherently antivirally active, which may explain why prior to this invention others failed to discover the anti-HTV activity of AAT.
  • Figure 4 shows representative results obtained with a carboxyterminal peptide FVYLI (SEQUENCE ID NO. 16) that is derived but not necessarily identical to a respective C-terminal pentapeptide from AAT.
  • FVFLM SEQUENCE ID NO. 1
  • FVFAM SEQUENCE ID NO. 2
  • FVALM SEQUENCE ID NO. 3
  • FVFLA SEQUENCE ID NO. 4
  • FLVFI SEQUENCE ID NO. 5
  • FLMII SEQUENCE ID NO. 6
  • FLFVL SEQUENCE ID NO. 7
  • Example 5 Anti-HIV effect of drugs having AAT activity
  • a series of drugs that may mimic AAT activity are tested for anti-HIV activity
  • These man-made drugs are made according to methods described in WO 98/24806, which discloses substituted oxadiazole, thiadiazole and triazole as serine protease inhibitors
  • U S Pat No 5,874,585 discloses substituted heterocychc compounds useful as inhibitors of serine proteases
  • U S Pat No 5,869,455 discloses N-substituted derivatives
  • U S Pat No 5,861,380 discloses protease lnhibitors-keto and di- keto contammg rmg systems
  • U S Pat No 5,807,829 discloses serine protease inhibitor— t ⁇ peptoid analogues
  • U S Pat No 5,801,148 discloses serine protease mhibitors-prolme analogues
  • U S Pat No 5,618,792 discloses substituted heteroc
  • Example 7 Antiviral Activity of Man-Made Small Molecules Without limiting to AAT and peptide derivatives of AAT, the compounds like oxadiazole, thiadiazole and triazole peptoids are preferred as they also show an equivalent antiviral activity in a mouse model as described m above
  • Anti-HIV effective doses are in a range from about 1 ⁇ g/kg to approximately 100 mg/kg
  • Specific examples of such oxadiazole, thiadiazole and triazole peptoids are molecules such as Benzyloxycarbonyl-L-valyl-N-[l-(2-(3-methylbenzyl)-l,3,4-oxad ⁇ azolyl]carbonyl)-2-
  • phenylenedialkanoate esters which are also effective in the mouse model.
  • phenylenedialkanoate esters include but are not limited to: 2,2'-(l,4-phenylene)dibutyric acid; tert-butyl-3-chloro-pivaloate; dimethyl -
  • 5,216,022 teaches other small molecules useful for the practice of this invention, including: Benzyloxycarbonyl-L-valyl-N-[l-(2-[5-(3-methylbenzyl)-l,3,4-oxadiazolyl]carbonyl)- 2-(S)-methylpropyl]-L-prolinamide (also known as CE-2072), Benzyloxycarbonyl-L-valyl-N-[ 1 -(2-(3- methylbenzyl)-l,3,4-oxadiazolyl]carbonyl)-2-(S)-methylpropyl]-L-prolinamide; Benzyloxycarbonyl-L- valyl-N-[l-(2-(5-(methyl)-l,3,4-oxadiazoly]carbonyl)- 2-(S)-methylpropyl]-L-prolinamide; Benzyloxycarbonyl)-L-valyl-N-[l-(2-(5-(
  • the Examples recited hereinabove show that compounds exhibiting AAT activity such as AAT, peptides derived analogous or homologous to C-terminal end of AAT, and man-made synthetic molecules mimicking AAT action, display he ⁇ es virus-suppressive effects in vitro and in vivo
  • AAT and AAT-related molecules displaying AAT activity are tested for possible utility as a combmation therapy with established anti-HIV drugs
  • nucleoside reverse transcriptase (RT) inhibitors such as Retrovir (AZT/ zidovudine, Glaxo Wellcome), Epivir (3TC, lamivudine, Glaxo Wellcome), Videx (ddl/didanosine, Bristol-Myers Squibb), Hivid (ddC/zalcitabine,
  • NRTIs non-nucleoside reverse transcriptase inhibitors
  • Viramune nevirapine, Roxane Laboratories
  • Resc ⁇ ptor delavirdme, Pharmacia & Upjohn
  • Sustiva efavirenz, DMP-266, DuPont Merck
  • Preveon adefovir dipivoxil, bis- POM PMEA, Gilead
  • aspartyl protease inhibitors Pieris subtilis
  • results obtained in HIV-1 -infected PBMC demonstrate several characteristics of AAT inhibition
  • Experiments are performed in PBMC from three donors infected in the absence or presence of AAT during infection
  • the presence of AAT du ⁇ ng infection did not affect p24 antigen production following removal of AAT and 4 days of culture in medium alone Therefore, any effects of AAT at the time of infection are reversible
  • AAT effects during the infection period are established by the enhancement of AAT effect when added to PBMC following infection and cultured for 4 days Enhancement of 4 day AAT effect is manifested by a larger maximal suppression and by suppression at lower AAT concentrations
  • Expe ⁇ ments performed in MAGI-CCR-5 cells indicate inhibitory effects of AAT and related compounds on early infection-associated events
  • the observed dose-dependent effect is maximal at 5 mg/ml AAT, where 76% inhibition is observed compared to control (HIV-1 added m the absence of AAT) Therefore, AAT inhibits HIV-1 events prior to integration into the host-cell genome (cell-surface receptor binding, mternahzation, integration, uncoating, reverse transcription, translation and protem processing and tat activation)
  • AAT peptides derived analogous or homologous to C-terminal end of AAT, and representative man-made synthetic molecules mimicking AAT action, display HIV-1 -suppressive effects operative during both early (PBMC and MAGI-CCR-5 cell results) and late (Ul cell results) events associated with HTV-1 infection
  • PBMC early (PBMC and MAGI-CCR-5 cell results)
  • Ul cell results late (Ul cell results) events associated with HTV-1 infection
  • the synergy appears to exist between known AIDS drugs belonging to RT and PI classes and compositions of this mvention, which belong to unrelated class of inhibitors, l e , se ⁇ ins

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Abstract

L'invention concerne une nouvelle méthode de traitement et de prévention d'une infection virale. L'invention concerne, en particulier, une méthode destinée à combattre une infection virale favorisée par une activité de sérine protéolytique (SP), consistant à administrer à un sujet souffrant ou susceptible de souffrir d'une infection virale une quantité thérapeutiquement efficace d'un composé présentant une activité d'inhibition de sérine protéase ou serpin. Parmi les composés se trouvent l'antitrypsine α1 (AAT), des dérivés peptidiques de l'extrémité carboxyterminale de l'AAT, et des composés synthétiques artificiels imitant l'action de ces composés. Parmi les infections virales préférées se trouvent les infections rétrovirales telles que l'infection du virus de l'immunodéficience humaine (VIH).
PCT/US2000/005558 1999-03-05 2000-03-03 Inhibiteurs d'activite de serine protease, methodes et compositions de traitement d'infections virales WO2000052034A2 (fr)

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Cited By (11)

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WO2002058638A3 (fr) * 2001-01-26 2002-09-26 Gen Hospital Corp Medicaments a base de serpine pour le traitement de l'infection par vih et utilisation de ceux-ci
US6849605B1 (en) 1999-03-05 2005-02-01 The Trustees Of University Technology Corporation Inhibitors of serine protease activity, methods and compositions for treatment of viral infections
WO2007079312A3 (fr) * 2005-12-02 2008-02-21 Univ Colorado Compositions et procédés de traitement d’affections pathologiques médiées par l'actine
US7704958B1 (en) 1999-03-05 2010-04-27 Bio Holding, Inc. Methods and compositions for inhibiting apoptosis using serine protease inhibitors
WO2012035034A1 (fr) 2010-09-14 2012-03-22 F. Hoffmann-La Roche Ag Polypeptide de fusion à serpine-doigt de zinc
US8563693B2 (en) 2001-01-26 2013-10-22 Acceleration Biopharmaceuticals, Inc. Method of treating human immunodeficiency virus infection in a mammal comprising administering heparin-activated antithrombin III
US8633305B2 (en) 2003-08-26 2014-01-21 Leland Shapiro Compositions of, and methods for, alpha-1 anti trypsin Fc fusion molecules
JP2015531769A (ja) * 2012-08-23 2015-11-05 アリオス バイオファーマ インク． パラミクソウイルス感染症を治療するための化合物
US9938353B2 (en) 2011-06-24 2018-04-10 The Regents Of The University Of Colorado, A Body Corporate Compositions, methods and uses for alpha-1 antitrypsin fusion molecules
US10478508B2 (en) 2012-01-10 2019-11-19 The Regents Of The University Of Colorado, A Body Corporate Compositions, methods and uses for alpha-1 antitrypsin fusion molecules
CN113227057A (zh) * 2018-12-14 2021-08-06 Z 因子有限公司 化合物及其用于治疗α1-抗胰蛋白酶缺乏症的用途

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US5376633A (en) * 1992-09-30 1994-12-27 Lezdey; John Method for deactivating viruses in blood component containers
TR199901681T2 (xx) * 1996-12-06 2000-03-21 Cortech, Inc. Serin proteas inhibit�rleri.
AU7127298A (en) * 1997-04-14 1998-11-11 Emory University Serine protease inhibitors

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US7704958B1 (en) 1999-03-05 2010-04-27 Bio Holding, Inc. Methods and compositions for inhibiting apoptosis using serine protease inhibitors
US6849605B1 (en) 1999-03-05 2005-02-01 The Trustees Of University Technology Corporation Inhibitors of serine protease activity, methods and compositions for treatment of viral infections
US8071551B2 (en) 1999-03-05 2011-12-06 BioHolding, Inc. Methods and compositions for treating diabetes
US8563693B2 (en) 2001-01-26 2013-10-22 Acceleration Biopharmaceuticals, Inc. Method of treating human immunodeficiency virus infection in a mammal comprising administering heparin-activated antithrombin III
US7510828B2 (en) 2001-01-26 2009-03-31 Ralf Geiben Lynn Method of decreasing the infectivity of HIV in a biological sample through the administration of S-antithrombin
WO2002058638A3 (fr) * 2001-01-26 2002-09-26 Gen Hospital Corp Medicaments a base de serpine pour le traitement de l'infection par vih et utilisation de ceux-ci
US10913790B2 (en) 2003-08-26 2021-02-09 The Regents Of The University Of Colorado, A Body Corporate Compositions of, and methods for, alpha-1 anti trypsin Fc fusion molecules
US8633305B2 (en) 2003-08-26 2014-01-21 Leland Shapiro Compositions of, and methods for, alpha-1 anti trypsin Fc fusion molecules
US9499606B2 (en) 2003-08-26 2016-11-22 The Regents Of The University Of Colorado, A Body Corporate Compositions of, and methods for, alpha-1 anti trypsin Fc fusion molecules
US9695229B2 (en) 2003-08-26 2017-07-04 The Regents Of The University Of Colorado, A Body Corporate Compositions of, and methods for, alpha-1 antitrypsin Fc fusion molecules
WO2007079312A3 (fr) * 2005-12-02 2008-02-21 Univ Colorado Compositions et procédés de traitement d’affections pathologiques médiées par l'actine
WO2012035034A1 (fr) 2010-09-14 2012-03-22 F. Hoffmann-La Roche Ag Polypeptide de fusion à serpine-doigt de zinc
CN103108884A (zh) * 2010-09-14 2013-05-15 弗·哈夫曼-拉罗切有限公司 Serpin-finger融合多肽
US9403896B2 (en) 2010-09-14 2016-08-02 Hoffmann-La Roche Inc. Serpin-finger fusion polypeptide
US12030958B2 (en) 2011-06-24 2024-07-09 The Regents Of The University Of Colorado Compositions and methods of use of alpha-1 antitrypsin fusion polypeptides
US9938353B2 (en) 2011-06-24 2018-04-10 The Regents Of The University Of Colorado, A Body Corporate Compositions, methods and uses for alpha-1 antitrypsin fusion molecules
US10478508B2 (en) 2012-01-10 2019-11-19 The Regents Of The University Of Colorado, A Body Corporate Compositions, methods and uses for alpha-1 antitrypsin fusion molecules
US11014935B2 (en) 2012-08-23 2021-05-25 Janssen Biopharma, Inc. Compounds for the treatment of paramyxovirus viral infections
JP2015531769A (ja) * 2012-08-23 2015-11-05 アリオス バイオファーマ インク． パラミクソウイルス感染症を治療するための化合物
CN113227057A (zh) * 2018-12-14 2021-08-06 Z 因子有限公司 化合物及其用于治疗α1-抗胰蛋白酶缺乏症的用途
US12221422B2 (en) 2018-12-14 2025-02-11 Centessa Pharmaceuticals (Uk) Limited Compounds and their use for the treatment of alpha1-antitrypsin deficiency

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