WO2025210089A1 - A combination of vasoactive intestinal peptide (vip) receptor agonist and a mucolytic agent against inflammatory diseases of the respiratory tract - Google Patents

Abstract

The present invention relates to therapeutically effective combinations of vasoactive intestinal peptide (VIP) receptor agonist and a mucolytic agent. Furthermore, the invention relates to pharmaceutical compositions for use in the treatment or prevention of an inflammatory respiratory disease. Preferably, the VIP receptor agonist in said combination is aviptadil and the mucolytic agent is acetylcysteine (NAC).

Description

TITLE

A combination of vasoactive intestinal peptide (VIP) receptor agonist and a mucolytic agent against inflammatory diseases of the respiratory tract

DESCRIPTION

Technical Field

The present invention relates to combinations of vasoactive intestinal peptide (VIP) receptor agonist and a mucolytic agent, and pharmaceutical compositions for use in the treatment or prevention of inflammatory respiratory diseases.

State of the Art

The treatment of inflammatory diseases of the respiratory tract is primarily based on the administration of steroids to reduce inflammation. Steroids have been proven to be effective against IgE-mediated diseases such as allergic asthma. However, steroids show no significant effects in many diseases such as some forms of chronic obstructive pulmonary disease (COPD) and post/long-COVID-19 syndrome.

COPD is a major and increasing global health problem and is the fourth most common cause of death in the developed countries. The condition is associated with progressive breathlessness with slow decline in lung function. COPD will account for over six million deaths per year and is predicted to increase from the sixth to the third leading cause of death worldwide. The disease is associated with oxidative stress, inflammation and accelerated decline in lung function, while patient’s antioxidant defense gradually weakens. Patients with COPD experience a poorer quality of life, suffer from comorbidities associated with cardiovascular, muscular (skeletal) and neurological problems. These comorbidities are associated with increased oxidative stress.

The inflammatory process can alter the bronchi, bronchioles, sinuses and pulmonary parenchyma, leading to progressive restriction of airflow, resulting in emphysema and chronic bronchitis. The pathogenesis of emphysema includes destruction of alveolar septa, increased air space, and loss of elastic recoil due to hyperinflammation and oxidative stress. Chronic bronchitis involves the overproduction and hypersecretion of mucus by goblet cells, thereby reducing airflow.

Three interrelated complexes contribute to the pathophysiology of COPD. First, there is simple, i.e. non-obstructive chronic bronchitis, which is characterized by a hypertrophy of the bronchial mucosa with increased mucus secretion and is also associated with a dysfunction of the ciliated epithelium. Second, there is chronic obstructive bronchitis, which leads to an expiratory collapse of the bronchi due to mucosal atrophy and hyperresponsiveness with destruction of the bronchial structure. Third, there is pulmonary emphysema, which is characterized by an imbalance of endogenous tissue-damaging substances (including elastases and matrix metalloproteins) and protective substances (especially antitrypsin).

Antioxidant agents, such as thiol compounds/donors and analogs (GSH and mucolytic drugs, such as N-acetyl-L-cysteine (NAC), nacystelyn (NAL), erdosteine and ergothioneine have been reported to scavenge/detoxify free radicals/oxidants, increase intracellular thiol levels, control NF-KB activation, and consequently inhibit inflammatory gene expression. Other compounds include dietary antioxidants, superoxide dismutase (SOD) mimetics, and synthetic redox modulating agents. Several strategies either to increase the expression/activity of the antioxidant enzymes or to mimic their function by enzyme mimetics have been developed. These antioxidant molecules have been suggested to have the potential to be used as pharmacological therapeutic agents in the management and treatment of COPD. Chronic inflammation contributes to the maintenance of the disease process, which is initiated by noxious agents of various kinds. Recent findings suggest that there are differences in the composition of the products of cytokine gene expression in the inflammatory infiltrate, particularly in contrast to bronchial asthma. At the cellular level, COPD is characterized by an increase in T lymphocytes, neutrophil granulocytes and macrophages. In particular, the number of CD8-positive lymphocytes is increased, which is directly linked to the deterioration in lung function. This can result in a systemic disease, where other body components such as heart, muscles, blood vessels and bones are affected.

VIP (vasoactive intestinal peptide) is a peptide hormone that has a variety of effects in the body, particularly in the gastrointestinal tract, the nervous system and the circulatory system. VIP acts by binding to a specific membrane receptor. VIP is a 28 amino acid polypeptide that was first isolated from the intestines. Later, VIP was identified in the central and peripheral nervous system, and has since been recognized as a neuropeptide widely distributed, which acts as a neurotransmitter or neuromodulator in many organs and tissues, including heart, lung, thyroid, kidney, immune system, urinary tract and genitals. As neurotransmitter, VIP acts through activating the PACAP subfamily of class B1 G protein- coupled receptors, i.e. VIP receptor 1 (VPR1) and VIP receptor 2 (VPR2). The International Nonproprietary Name (INN) for VIP having 28 amino acids is “aviptadil”. The biological effects of aviptadil are mediated by G protein-coupled receptors, VPAC1 , VPAC2 and the PAC1 receptor.

EP 3 583 933 A1 describes kits for use in treating Chronic Beryllium Disease by administering specifically designed aerosols of aviptadil.

WO 2020/225246 A1 describes the use of VIP in the treatment of drug-induced pneumonitis. In one embodiment, a liquid pharmaceutical composition is aerosolized for administration.

WO 2021/152119 A1 describes anti-inflammatory peptides such as aviptadil for use in the treatment or prophylaxis of inflammatory pulmonary diseases by inhalation administration. An aerosol comprising solid particles or liquid droplets containing the anti-inflammatory peptide is administered to a patient of the inflammatory pulmonary disease, especially ARDS, for patients suffering from an infection by a Coronavirus, such as SARS-CoV-2.

US 2023/0381280 A1 describes the use of aviptadil in the treatment of a lung disease in a patient in need thereof, wherein the lung disease is Covid-19 lung disease caused by SARS- CoV-2 virus.

US 2024/0181014 A1 describes the use of aviptadil either alone or in combination with Alpha Lipoic Acid for the treatment of a post-viral infection syndrome such as SARS-CoV-2 infection.

Despite the recognized beneficial therapeutic properties of aviptadil, the stability of aviptadil and the accessibility to the target tissue still pose a problem in the prior art. Since aviptadil and the other VIP analogs are peptides, their stability in solution is limited. When administered by inhalation, the drug has difficulty reaching the target tissue if the bronchial secretion represents a barrier that the peptide has difficulty overcoming. As peptides are rapidly broken down in the body by the body's own enzymes, this greatly limits the duration of action. Furthermore, the storage of the dissolved aviptadil in the drug poses a problem, as the peptide is not sufficiently stable in solution. For example, it has been found that aviptadil is acid-sensitive in isotonic saline solutions at temperatures between 30°C and 60°C. Pharmaceutical compositions comprising aviptadil in a buffer that maintains the pH of the formulation at a pH from 5.9 to 6.1 showed improved stability compared to solutions with pH below 5 or higher than 7 (see PT1855661 E). However, there are still other factors that affect stability of aviptadil in pharmaceutical compositions, including, but not limited to the chemical reactivity of the active ingredients, possible interactions between the active and non-active ingredients, the manufacturing process, the dosage form, the container closure system, and conditions environmental encountered during shipping, storage, handling and amount of time between manufacture and use.

The maintenance of stability of the VIP receptor agonists such as aviptadil is still an unsolved task. It would therefore desirable to have improved compositions comprising VIP receptor agonists such as aviptadil or other VIPs that are therapeutically more potent over a longer period of time.

Description of the Invention

It is therefore the object of the present invention to provide improved therapeutically effective compositions containing a vasoactive intestinal peptide (VIP) receptor agonist for the treatment of inflammatory respiratory diseases, in particular inflammatory pulmonary diseases, which are more stable and thus effective than compositions containing the VIP receptor agonist alone.

The present invention solves this object by providing a therapeutically effective combination of at least one VIP receptor agonist and a stabilizing mucolytic agent.

The present invention is based on the surprising finding that a combination of VIP receptor agonist and a mucolytic agent improves the treatment or prevention of an inflammatory pulmonary disease in a synergistic manner. In more particular, it was found that the addition of a mucolytic agent stabilizes the VIP receptor agonist in a pharmaceutical composition and thereby enhances therapy efficiency. As shown herein, stabilization of the VIP receptor agonist by the mucolytic agent in the composition leads to prolonged treatment time and hence a better therapeutic outcome.

As used herein, the term “vasoactive intestinal peptide (VIP)” denotes the natural VIP peptide found in the human or animal body.

The name “aviptadil” denotes a synthetically produced analog of VIP and is named by the WHO according to INN standards. The term “VIP receptor agonist” denotes a substance which acts as an agonist at at least one of the following PACAP receptors: VPAC1 , VPAC2 and PAC1. Preferably the effect at VPAC1 and VPAC2 is beneficial for therapeutic purposes.

The term “N -acetylcysteine (NAC)” is the same as acetylcysteine (ACC) or N-acetyl-L- cysteine. The abbreviations NAC and ACC are used interchangeably.

The term “pharmaceutically acceptable” denotes a material that is not biologically, clinically or otherwise undesirable, i.e. , the material can be administered to an individual along with the relevant active compounds without causing clinically unacceptable biological effects or interacting in a deleterious manner with any of the other components of the pharmaceutical composition in which it is contained.

The term "subject" denotes any animal, including mammals. Mammals include, but are not limited to farm animals (such as, for example, horses, cows, and pigs), companion animals (such as, for example, dogs and cats), laboratory animals (such as, for example, mice, rats, and rabbits), and non-human primates. In preferred embodiments, the subject is a human. In some embodiments, the human is a patient under the care of a physician.

The “effective amount” denotes a nontoxic but sufficient amount of the compound to provide the desired result. As will be pointed out below, the exact amount required will vary from subject to subject, depending on the species, age, and general condition of the subject, the severity of the disease that is being treated, the particular compound used, its mode of administration, and the like. Thus, it is not possible to specify an exact “effective amount.” However, an appropriate effective amount can be determined by one of ordinary skill in the art using only routine experimentation.

The invention is based on the unexpected finding that a mucolytic agent contributes to the stabilization of the VIP receptor agonist in a pharmaceutical composition, thereby retaining a longer activity of the VIP receptor agonist. A better stabilization of the VIP receptor agonist peptide is crucial in many therapeutic applications. The combination of the VIP receptor agonist with a mucolytic agent such as NAC stabilizes the peptide structure and hence activity of the peptide.

In one aspect, the combination according to the present invention comprises a VIP receptor agonist, wherein the VIP receptor agonist is a natural VIP, a VIP derivative or a synthetic analog of VIP. In a preferred embodiment, the synthetic VIP is aviptadil or a functional derivative or precursor thereof.

The term “functional derivative”, as used herein, denotes any molecule that can be derived from a parent molecule, wherein both the parent molecule and the derivative have the ability to bind to at least one receptor of interest.

In some embodiments, the functional derivative described herein is a functional derivative of VIP binding to the receptor(s) VPAC1 , VPAC2, and/or PAC1, preferably to VPAC1, VPAC2, and PACT For example, functional derivatives of aviptadil can be prepared by derivatizing with one or more functional group, by amino acid deletions, insertion of one or more amino acids acids and/or by replacing one or more amino acids and/or by any other method known to the person skilled in the art (see e.g. US8329640B2, EP3311828A1 ; Campos-Salinas, J., et al., 2014, The Journal of biological chemistry, 289(21), 14583-14599). In some embodiments, the functional derivative has at least one improved property such as increased stability, increased half-life, altered ionic charge, altered hydrophobicity index, altered percentage of a-helix, or increased specificity.

The invention also covers variations, modifications, substitutions, deletions, additions of the amino acid sequence of aviptadil provided that the peptide still retains its therapeutic function. For example, such a functional derivative may have one or more amino acid substitutions, additions, deletions or chemical modifications in the amino acids sequence His- Ser-Asp-Ala-Val-Phe-Thr-Asp-Asn-Tyr-Thr-Arg-Leu-Arg-Lys-GIn-Met-Ala-Val-Lys-Lys-Tyr- Leu-Asn-Ser-lle-Leu-Asn (SEQ ID NO: 1). In some embodiments, a number of 2 to 7 amino acids are replaced, deleted or added to the basic 28 aa amino acid sequence of aviptadil.

The term “precursor”, as used herein, refers to any molecule(s) that can be turned into an active component by a chemical reaction. In some embodiments, the chemical reaction turning the precursor into an active component occurs before or during the administration process (e.g. in an administration device or in the aerosol). In some embodiments, the precursor described herein is a prodrug and the chemical reaction occurs in the body. In some embodiments, the chemical reaction turning the precursor into an active component is catalyzed by an enzyme of the body, preferably an enzyme expressed by cells of the respiratory tract and/or the central nervous system. In some embodiments, the precursor described herein is pre-pro aviptadil. The production of precursors of aviptadil is known to the person skilled in the art (see, e.g. Simoncsits A, et al., 1988, Eur J Biochem. December 15; 178(2):343-50.)

In a preferred embodiment, the synthetic analog of VIP is N-stearyl-[Nle17]-VIP or Ro 25- 1553.

N-stearyl-[Nle17]-VIP is a N-stearyl, Norleucin 17 VIP hybrid which inhibits 1251-VIP binding to human K-2 cells with an IC50 value of 0.01 pM. The analog contains two chemical modifications in VIP: the addition of an N-terminal long chain aliphatic acid and the substitution of the Met in position 17 with Nle. These changes confer stability, longer half-life and increased bioavailability. This analog exhibited both a 100-fold potency as compared to VIP (maximal effect manifested at 1 pM) and specificity for a VIP receptor in neuronal survival and neuroprotection against the p-amyloid peptide fragment (the Alzheimer's disease neurotoxin).

Ro 25-1553 is a cyclic peptide analog of vasoactive intestinal peptide (VIP) that was designed to overcome many of the deficiencies inherent in this natural neuropeptide. It primarily acts on the VPAC2 receptor.

Any suitable stabilizing mucolytic agent can be used in combination with the active VIP receptor agonist according to the present invention. In preferred embodiments, the mucolytic agent is any one of N-acetylcysteine (NAC), N-acystelyn (NAL), N-isobutyrylcysteine (NIC), procysteine, gluthathion ester, erdosteine, carbocysteine, fudosteine, ergothioneine. The use of NAC together with a VIP receptor agonist is preferred.

In some embodiments, the Vip receptor agonist is aviptadil comprising the amino acid sequence set forth in SEQ ID NO: 1 , and the mucolytic agent is NAC.

Both components of the combination can be contained together in one composition. In alternative embodiments, the both components are contained in separate compositions and be combined either prior to administration or administered to a subject in two distinct dosage forms. In some embodiments, the VIP receptor agonist is provided in a first dosage form and the mucolytic agent is provided as a second dosage form, wherein a dosage regimen provides for the first dosage form to be administered to a subject concurrently with or prior to the second dosage form. Preferably, the time interval for the first dosage form is between 10 and 30 minutes before administration of the second dosage form. Preferably, the combination containing the VIP receptor agonist and the stabilizing mucolytic agent is provided in liquid form or as an aerosol.

As illustrated in the examples, the combination of a mucolytic agent and VIP receptor agonist results in a stabilization of the VIP receptor agonist in the solution. This effect is complemented by the pharmacological effect of the mucolytic agent as an expectorant and free radical scavenger in the lungs.

The present invention also relates to a method for the manufacturing of a medicament comprising the combination of a vasoactive intestinal peptide (VIP) and a mucolytic agent to a pharmaceutical composition.

The addition of a mucolytic agent, such as NAC, gives the VIP receptor agonist, such as aviptadil, a greater chance of binding to its receptor and exerting its pharmacological effect. Preferably, both components are provided in a buffer that keeps the pH value within a given range and therefore stable. Examples of such buffers include, but are not limited to phosphate buffered saline (PBS) or amino acid buffer (e.g., glycine buffer). Other excipients can be added for preservation, such as EDTA or its salts. In some embodiments, NAC is provided in concentrations that sufficiently buffer the solution so that additional buffer buffers are not required.

In preferred embodiments, the formulations optionally contain excipients to improve the stability and handling of the formulation. Lactose monohydrate is used as a carrier to ensure an even distribution of the active ingredient. Microcrystalline cellulose is used as a filler to stabilize the formulation. Magnesium stearate is used as a lubricant to ensure smooth processing of the drug during manufacture.

In preferred embodiments, the combination of the present invention comprises between 0.001 and 1.0% w/v aviptadil in a buffer, wherein said buffer maintains the pH of the formulation at a pH from 5.9 to 7.5.

In preferred embodiments, the buffer is selected from the group consisting of lactate buffer, amino acid buffer, benzoic acid, oxalate, fumarate, aniline, acetate buffer, citrate buffer, glutamate buffer, phosphate buffer, succinate, pyridine , phthalate, histidine, 2- (N- morpholino) ethanesulfonic acid, maleic acid, cacodylate, carbonic acid, N- (2-acetamido) imino-diacetic acid, 4- piperazinabis- acid (ethanesulfonic acid), BIS-TRIS-propane, ethylenediamine acid 2 - [(2-amino-2-oxoethyl) amino] - ethanesulfonic acid, imidazole, 3- (N-morfin) acid - propanesulfonic acid, dietilmalonico acid, 2- [tris (hydroxymethyl) methyl] amino-ethanesulfonic acid; and N- 2-hydroxyethylpiperazine-N-2-ethanesulfonic acid.

In some embodiments, sodium phosphate buffers are used to stabilize the pH value of inhalation solutions. These buffers consist of a mixture of sodium phosphate monobases (NaH2PC>4) and sodium phosphate dibases (N32HPO4) in various concentrations to achieve the desired pH range.

In some other embodiments, acetate buffers are used for adjusting the pH value of inhalation solutions. They consist of acetic acid and its salts (sodium acetate or potassium acetate) and can provide an acidic or alkaline pH value as required.

In some other embodiments, citric acid/trisodium citrate buffers are used to buffer inhalation solutions while providing a pleasant taste. The buffers consist of a mixture of citric acid and trisodium citrate and can effectively stabilize the pH value.

In some other embodiments, PBS is used as buffer solution containing an isotonic saline solution with a defined pH value. PBS is often used as a basis for the preparation of inhalation solutions as it has a physiological composition and is well tolerated by the respiratory tract.

Furthermore, amino acid buffers can be used for the production of inhalation solutions. Amino acid buffers offer an alternative way of adjusting and stabilizing the pH value of inhalation solutions. Amino acid buffers can offer advantages over conventional inorganic buffer solutions in some cases. For example, they may have lower toxicity and interact better with certain active ingredients, particularly those that are sensitive to pH or interact with proteins. However, the use of amino acid buffers in inhalation medication depends on various factors, including the specific properties of the active ingredient, the desired pH stability and respiratory tolerability. A preferred inhalation solution uses glycine buffer as amino acid buffer, which consists of the amino acid glycine and its associated buffer base.

Some preparations do not require a separate buffer system since acetylcysteine itself also acts as a buffer substance.

The buffer solutions can be selected according to the requirements of the specific inhalation medication and the properties of the active ingredient. It is important that the buffer solution stabilizes the pH, ensures the stability of the drug and at the same time is safe and well tolerated by the patient.

In preferred embodiments, aviptadil is used in available formulations at a concentration of 0.01% (w/v). Suitable formulations have between 0.001% and 0.1% (w/v) aviptadil. In a preferred embodiment 0.005% to 0.05%. It is known to the skilled person which volumes of the solution must be used for inhalation. For example, a volume of 1 ml per day of a 0.01% (w/v) solution is recommended for inhalation in patients of average body weight.

The mucolytic agent used in the combination of the present invention may trigger different effects including mucus clearance, antioxidant activity, anti-inflammation, paracetamol intoxication and protection against oxygen deficiency. Most importantly and surprisingly, the mucolytic agent stabilizes the VIP receptor agonist is solution and extends its effective time. The VIP therapy will be more effective and efficient. In addition, the mucolytic agent has an expectorant effect, which has proven to be beneficial in the combination treatment with the VIP receptor agonist, in particular aviptadil.

The therapeutically effective combination according to the invention is in particular suitable for the treatment or prevention of an inflammatory respiratory disease, preferably an inflammatory pulmonary disease. Preferably, the inflammatory respiratory disease, in particular inflammatory pulmonary disease, is any one of chronic obstructive pulmonary disease (COPD), pneumonia, pulmonary fibrosis, alveolitis, pneumonitis, sarcoidosis, chronic obstructive bronchitis, acute respiratory distress syndrome (ARDS), or pulmonary hypertension resulting from inflammatory disease, inflammatory viral respiratory disease, COVID-19 disease or COPD, checkpoint inhibitor pneumonitis (CIP), berylliosis, steroid resistant asthma, allergic rhinitis.

In preferred embodiments, the inflammatory pulmonary disease is a COPD-mediated by neutrophil cells. This is a special form of COPD that involves neutrophils and can be distinguished from other forms of COPD.

The present invention also relates to pharmaceutical compositions comprising a VIP receptor agonist and a mucolytic agent as defined herein and a pharmaceutically acceptable carrier or diluent. Preferably, such a pharmaceutical composition is used in the treatment or prevention of inflammatory pulmonary diseases mentioned above. In some embodiments, the composition is provided for inhalation, oral or intranasal administration.

In the upper airways, treatment of a subject can also be carried out by inhalation (nasal), e.g. by the application of a nasal spray. Preferred inflammatory diseases of the upper airways to be treated with a formulation according to the present invention are allergic rhinitis, chronic rhinitis, allergic sinusitis or chronic sinusitis. These indications are in particular relevant for people suffering from chronic rhinitis as those people have a higher risk of developing COPD.

If the combination of the invention is provided as an inhalation solution, the exact composition of the VIP receptor agonist and the mucolytic can vary depending on the manufacturer and the specific formulation. Typically, an inhalation solution contains a defined amount of VIP receptor agonist and a defined amount of the mucolytic agent as well as various excipients that may serve to ensure the stability of the solution, facilitate administration or improve the taste. In preferred embodiments, the combination is provided as a powder for inhalation that is highly effective.

A typical inhalation solution contains approximately 1 - 20 % (w/v) of the mucolytic agent such as NAC solved in water. Water provides the basis for the inhalation solution and is typically used for injections. To ensure the stability of the solution and to control the pH value, additional buffer substances such as sodium hydroxide or hydrochloric acid can be added. To prevent contamination of the solution and extend its shelf life, preservatives such as benzalkonium chloride or EDTA can be used. Since some mucolytic agents can have a characteristic taste that is unpleasant to some people, flavorings or sweeteners can be added to improve the taste and increase the acceptability of the solution. In some embodiments, sterile filtration of the solution could be required.

In a preferred embodiment, the doses comprise 40 to 80 pg of aviptadil, preferably between 50 and 70 pg of aviptadil, more preferably 70 pg of aviptadil. If the dose comprises about 70 pg of aviptadil, the administration dose can be around 280 pg of aviptadil per day by applying two doses of aviptadil (each 70 pg) preferably in the morning and two doses of aviptadil (each 70 pg) in the evening of each day.

The two biologically active components of the combination can also be used in a combination therapy. By using a combination therapy, a broader mechanistic coverage is achieved which benefits treatment. In one aspect, the combination targets biological pathways and mechanisms of disease processes that a single agent does not fully manage. The combined, synergistic effect of the combination of a VIP receptor agonist and a mucolytic agent is greater than the sum of their individual effects. This synergy will improve treatment outcomes, particularly in complex inflammatory lung diseases where multiple pathways are involved. A monotherapy may only target one specific component of a disease pathway. However, many inflammatory lung diseases involve multiple pathways or redundant systems that can bypass a single blockade. The therapeutic application of the combination according to the present invention can cover a wider range of these processes because distinct drugs are used with different mechanisms and modes of action.

In one aspect, the pharmaceutically composition comprises a therapeutically effective combination as defined herein and a pharmaceutically acceptable carrier or diluent for use in the prevention or treatment of an inflammatory pulmonary disease.

In a preferred embodiment, the VIP receptor agonist and the mucolytic agent in the therapeutically effective combination are provided in a single dose form or in two distinct dose forms. The one embodiment, the single dose form contains both the VIP receptor agonist and the mucolytic agent. In an alternative embodiment, one dose form contains the VIP receptor agonist and another dose form contains the mucolytic agent.

A pharmaceutical composition according to the invention may be formulated with any known pharmaceutically acceptable carrier or diluent as well as any other known adjuvants and excipients in accordance with conventional techniques. The pharmaceutically acceptable carriers, diluents, adjuvants and excipients should be suitable for the chosen inductor of the present invention and the chosen mode of administration. A pharmaceutical composition of the present invention may also include diluents, fillers, salts, buffers, detergents (e. g., a nonionic detergent), stabilizers (e. g., sugars or protein-free amino acids), preservatives, tissue fixatives, solubilizers, and/or other materials suitable for inclusion in a pharmaceutical composition.

The pharmaceutical compositions of the present invention can be formulated by methods known to those skilled in the art. For example, such pharmaceutical compositions can be used parenterally, as injections which are sterile solutions or suspensions including the compositions along with water or another pharmaceutically acceptable liquid. For example, such compositions may be formulated as unit doses that meet the requirements for the preparation of pharmaceuticals by appropriately combining the compositions with pharmaceutically acceptable carriers, diluents, adjuvants or excipients, specifically with sterile water, physiological saline, a vegetable oil, emulsifier, suspension, surfactant, stabilizer, flavoring agent, excipient, vehicle, preservative, binder or such. In such preparations, the amount of active ingredient is adjusted. Preferred embodiments are pharmaceuticals that are adapted for administration by inhalation, oral or intranasal administration.

The application of the therapeutically effective combination of the invention by inhalation of an aerosol using an inhalation device is preferred. Inhalers or nebulizers can be used that are designed in accordance with the guidelines on pharmaceutical quality of inhalation and nasal products of the European Medicines Agency. It is important that all requirements regarding product safety and hygiene requirements are met.

In some embodiments, the aerosol of particles or droplets (dry or liquid) has a diameter of 0.5 to 10.0 pm, although diameters between 2.0 and 6.0 pm are preferred. The aerosol consists of liquid or dry droplets or particles and the carrier gas, which may preferably be an inert gas such as helium or N2. Compressed air can also be used, but care must be taken to ensure that no undesirable oxidation occurs due to the presence of oxygen.

The particle size for inhalation of an aerosol can be of critical importance in determining the site of deposition in the respiratory tract. The optimum size for particles to settle in the airways is between 1 and 5 pm mass median aerodynamic diameter (MMAD), preferably between 2 and 5 pm MMAD. Larger particles settle out in the upper airways, whereas smaller particles remain suspended and are therefore exhaled. In some embodiments, drug particles of about 1 pm MMAD are delivered, e.g. by using drugs formulated in nonflammable hydrofluoroalkane (HFA) aerosol propellant gases.

In a preferred embodiment, the nebulizers or inhalers comprise a dosing device, whereby a specific aerosol volume is provided. The dose preferably comprises 40 to 70 pg, more preferably about 70 pg of aviptadil. With this dose it is possible to administer about 280 pg of aviptadil per day, e.g. by applying two doses of 70 pg of aviptadil preferably in the morning and two doses of about 70 pg of aviptadil in the evening of each day. In some embodiments, the dose is around 50 pg of aviptadil.

The two pharmaceutically active components of the combination of the present invention, i.e. the VIP receptor agonist and the mucolytic agent can be combined in a single composition for use in the treatment or prevention of an inflammatory pulmonary disease. In alternative embodiments, the VIP receptor agonist and the mucolytic agent are provided separately in two distinct compositions. In a first aspect, the composition containing the VIP receptor agonist and the composition containing the mucolytic agent can be combined prior to administration to a subject in need thereof. In a second aspect, the composition comprising the VIP receptor agonist and the composition comprising the mucolytic agent are administered as a single dose or repeated dose to a subject in need thereof. In a third aspect, the composition comprising the VIP receptor agonist and the composition comprising the mucolytic agent are administered to a subject in need thereof within a desired time interval. For example, the composition comprising the VIP receptor agonist is given to a subject in need thereof at a first time point, while the composition comprising the mucolytic agent is given to a subject in need thereof at a second time point within a given time interval. Preferably, the time interval for administering a single or repeated dose of the first composition comprising the VIP receptor agonist and administering a single or repeated dose of the second composition comprising the mucolytic agent is at least 2 days, preferably at least 4 days. The administration of the first and/or second composition can be repeated several times. In addition, the number of administrations, the composition and concentration of the compositions comprising the VIP receptor agonist and/or mucolytic agent can vary. For example, the individual doses given to a subject may vary and depend, inter alia, from gender, age, constitution and state of illness.

In some embodiments, the VIP receptor agonist is in the same dosage form as the mucolytic agent such that the VIP receptor agonist is administered concurrently with the mucolytic agent. In a preferred embodiment, the dosage form is a single fluid or inhalation solution. In some embodiments, the dosage form including the VIP receptor agonist and the mucolytic agent is prepared by combining a composition including the VIP receptor agonist and a composition including the mucolytic agent. In some embodiments, in the pharmaceutical composition of any one of embodiments disclosed herein the VIP receptor agonist is in a separate dosage form from the mucolytic agent such that the VIP receptor agonist can be administered before or after the mucolytic agent. A delayed administration of the mucolytic agent relative to the VIP receptor agonist is preferred.

As shown herein, combining a VIP receptor agonist with a mucolytic agent such as NAC leads to a higher stability of the VIP receptor agonist in solution. This could be explained in a first aspect by the fact that the mucolytic has an antioxidant effect against the oxidation of the VIP receptor agonist. The mucolytic agent is a strong reducing agent and scavenges reactive oxygen species (ROS) leading to oxidation of the peptide. This prevents or slows down oxidative degradation processes and maintains the biological activity of the VIP receptor agonist over a longer period of time In a second aspect, the mucolytic agent protects against disulfide bridge formation and aggregation. Peptides such as aviptadil contain sulphur-containing amino acids (e.g. cysteine), which can be prone to unwanted disulphide bond formation and aggregation. Mucolytic agents such as acetylcysteine act as a reducing agent and stabilize free thiol groups, preventing the formation of faulty disulfide bonds. This minimizes aggregation and maintains the solubility of the peptide.

In third aspect, the mucolytic agent provides pH stabilization to prevent hydrolysis. VIP receptor agonists such as aviptadil are particularly sensitive to pH fluctuations, as it hydrolyzes easily in acidic or strongly alkaline environments. As shown herein, NAC buffered to pH 7.2 provides a stable, slightly neutral environment in which the peptide is less susceptible to hydrolytic degradation. A stable pH value also prevents deamidation reactions that could impair the effectiveness of the peptide.

In a fourth aspect, the mucolytic agents support chelation with metal ions to inhibit catalytic degradation mechanisms. In physiological saline solution, traces of ions (e.g. iron or copper) can catalyze oxidative reactions that break down the VIP receptor agonist (aviptadil). The mucolytic agent such as NAC can bind these metal ions by chelation and thus inhibit catalytic degradation mechanisms.

In fifth aspect, the mucolytic agent provides increased solubility and bioavailability since it can improve the homogeneity of the solution due to its mucolytic and interaction-reducing properties. As a result, the VIP receptor agonist remains evenly distributed, which ensures its effectiveness over longer storage periods.

As shown herein, the combination of aviptadil and NAC is superior to the administration of aviptadil alone. NAC provides an antioxidant effect against oxidative degradation, protects against disulfide bridge formation and aggregation, and stabilizes the pH to prevent hydrolysis. This is important since aviptadil is particularly sensitive to pH fluctuations, as it hydrolyzes easily in acidic or strongly alkaline environments. NAC buffered to pH 7.2 provides a stable, slightly neutral environment in which the peptide is less susceptible to hydrolytic degradation. Furthermore, a stable pH value also prevents deamidation reactions that could impair the effectiveness of the peptide. Finally, NAC binds to metal ions (e.g. iron or copper) by chelation and thus inhibit catalytic degradation mechanisms. The therapeutic success of the inventive combination can be evaluated, for instance, by using the CAT score. The COPD Assessment Test (CAT) is a recognized scoring system for COPD patients, which provides a simple method for assessing the impact of COPD on the patient's health.

The present invention is further illustrated in the following examples. By no means shall the present invention be limited to these specific examples. The invention also covers combinations and modifications of single features or entire embodiments.

Brief of the Fi

Figure 1 shows the CAT score of a patient treated with a combination comprising aviptadil and NAC by inhalation.

Figure 2 shows the increased stability of a solution containing both aviptadil and NAC in comparison to a solution containing aviptadil only.

Best Ways to Carry Out the Invention

EXAMPLES

Example 1 - CAT score analyses using aviptadil and NAC combination

Formulation (per dose):

* Aviptadil: 50 pg

* Acetylcysteine (NAC) 50 mg

* Lactose monohydrate: 50 mg

* Microcrystalline cellulose: 20 mg

* Magnesium stearate: 1 mg

The formulation enables targeted administration of the active ingredient into the airways of a subject to be treated in order to achieve a maximum effect on the airways.

Aviptadil, the main ingredient of the drug, is used in an amount of 50 pg per dose. The combination is produced in the form of a powder for inhalation that is highly pure and effective. For application, patients use a special inhalation device (M-Neb® dose+ mesh nebulizer MN-300/8) to take the correct dose of the powder. The dosage and frequency of patient’s inhalation are determined according to the severity of the illness and the individual needs of the individuals to be treated. It is recommended that a specialist doctor or a specialist in the field of inhalation medication determines the exact dosage and use of the medication.

The results of the combined application of aviptadil and NAC are summarized in Figure 1. The treatment with aviptadil leads to a significant improvement of the CAT score in the patient. After 6 months no further improvement was observed. However, and most importantly, when aviptadil plus NAC as stabilizing and mucolytic agent was administered, a further significant improvement in CAT score could be observed.

Conclusion:

The addition of NAC improves the treatment of COPD patients applying VIP receptor agonist inhalation. Targeted administration into the airways achieves an optimized effect to slow down the progression of the inflammatory respiratory disease and relieve patients' symptoms.

Example 2: Administration of inhalation formulation comprising aviptadil and NAC

Composition containting aviptadil and NAC:

0.9 % (w/v) NaCI

0.017 % (w/v) aviptadil

10 % (w/v) acetylcysteine (NAC) pH 7 to 8 (in aqueous solution)

Several patients suffering from inflammatory lung diseases were treated with an aerosol and an inhaler. A M-neb® dose+ mesh nebulizer MN-300/8 was used to accurately measure the particle size diameter. Diameters between 2.0 and 6.0 pm were selected.

When aviptadil was administered in combination with ACC to patients who suffer from COPD, an anti-inflammatory effect could be observed. This was particularly pronounced in patients whose COPD was characterized by high levels of neutrophil granulocytes and who exhibited symptoms that largely corresponded to the clinical picture of chronic obstructive bronchitis. Furthermore, it was noticeable in COPD patients and in patients who suffer from post/long Covid-19 disease (ICD-10: U09.9!) that the clinical picture led to a pulmonary hypertension, which was significantly improved by the combination therapy resulting in an overall higher fitness and improved resilience of the patients. Although this effect could also be observed in patients who suffer from COPD with pronounced neutrophil granulocyte infiltration in the lungs when aviptadil was administered alone (control, without ACC), the effect was significantly more pronounced when NAC was present in the composition.

In patients suffering from chronic rhinitis, the inhalation of Aviptadil in combination with NAC led to a significant improvement in symptoms. The obstruction of the nose and sinuses was significantly reduced by a 4-week therapy.

Example 3 - Improvement of COPD using a combination of aviptadil and NAC

Initial situation of the patient:

• Patient 1: 61 years old

• Diagnosis: COPD with moderate impairment (Gold 2)

(CAT score: 24 points)

• Symptoms: o Occasional shortness of breath on exertion o Moderate, productive cough o Restless sleep due to night-time coughing o Fluctuating energy levels, sometimes tiredness during the day

Phase 1 : 6 weeks

Aim: Improvement of sleep quality, reduction of breathlessness and stabilization of symptoms.

Course of the first 6 weeks:

Week 2: o Initial improvements in sleep: fewer night-time awakenings o Slight reduction in coughing o CAT score: 22 (-2 points)

Week 4: o Shortness of breath on exertion continues to decrease o Patient feels more alert during the day o Cough still reduced o CAT score: 20 (-4 points, noticeable improvement)

Week 6: o Sleep quality significantly stabilized o Energy levels continue to improve o Shortness of breath barely noticeable in everyday life o CAT score: 19 (-5 points, from moderate to lower limit of impairment)

Phase 2: 4 weeks of maintenance therapy with avi further

Aim: Maintaining the achieved improvement without experiencing any deterioration.

Week 10: o Stable condition o No further deterioration o Sleep remains restful o CAT score remains at 19

Phase 3: Addition of NAC for additional 4 weeks

Aim: Improvement of mucus clearance and reduction of coughing.

Week 12: o Mucus becomes thinner, coughing is even rarer o Less coughing in the morning o CAT score: 18 (-6 points in total)

Week 14: o Almost no more productive coughing o Even more energy during the day o CAT score: 17 (-7 points, significant improvement in the lower range of impairment)

Conclusions:

The administration of aviptadil alone resulted in improved sleep quality, reduced breathlessness and stabilized COPD (-5 CAT points in 6 weeks). This condition remained stable for over 4 weeks. With the addition of NAC after 10 weeks, there was an additional improvement, particularly through mucus relief and cough reduction (-2 additional CAT points). In summary, the CAT score could be reduced from 24 to 17, which means a clear reduction in COPD impairment using a combination therapy.

Example 4 - Improvement of COPD with aviptadil and NAC in patient 2

Initial situation of the patient:

• Patient 2: (65 years)

• Smoking history: 70 packyears, continues to smoke despite COPD

• Diagnosis: Severe COPD-Gold 3 (CAT score 29 points)

• Previous therapy: o Inhaled corticosteroids (ICS) o Long-acting bronchodilators (LABA/LAMA) o Improvement through this therapy by only 3 points (CAT score: 26)

• Symptoms: o Severe shortness of breath even with the slightest exertion o Hard, productive cough with thick mucus o Frequent exacerbations and infections o Exhaustion, low physical resilience

New therapy: inhalation with acetylcysteine (NAC) and aviptadil

Aim: Further improvement of symptoms despite continued smoking.

Expected effects (6 weeks therapy with NAC + aviptadil):

Week 2: o Mucus becomes thinner, easier to cough up o Less coughing in the morning o First improvement in shortness of breath o CAT score: 24 (-2 points)

Week 4: o Noticeably better lung function due to bronchodilatation o Exercise tolerance increases (shorter recovery after exertion) o Reduction of exacerbations o CAT score: 22 (-4 points) Week 6: o Significant reduction in breathlessness at rest o Less fatigue, more energy o No further deterioration despite continued smoking o CAT score: 19 (-7 points)

Overall improvement: From severe COPD (CAT 29) to moderate-severe impairment (CAT 19), despite continued smoking habit. Aviptadil and NAG together led to an improved quality of life, particularly through better sleep, less coughing and increased daily energy.

Conclusions:

Despite continued smoking, the combination of aviptadil and NAC led to a significant reduction in the CAT score of 7 points. Aviptadil reduced inflammation and improved bronchial dilation. In addition, the presence of the mucolytic agent NAC made it easier for the patients to cough up phlegm and reduced exacerbations. In summary, the therapy was therefore more successful than the previous standard therapy using ICS + LABA/LAMA alone.

Example 5 - Improvement of COPD disease by aviptadil and NAC combination therapy

Initial situation of the patient:

• Patient 3: 67 years

• Diagnosis: Severe COPD-Gold 3 (CAT score: 28 points)

• Symptoms: o Severe cough with thick mucus o Shortness of breath even with light exertion o Frequent coughing fits at night o Low energy and fatigue

Phase 1 : 6 weeks

Week 2: o Reduction in breathlessness (less tightness in the chest) o Slight improvement in energy (less fatigue) o CAT score: 24 (-4 points) Week 4: o Fewer coughing fits, especially at night o Improved oxygen uptake — > more resilience o CAT score: 21 (-7 points, clinically relevant)

Week 6: o Significantly less shortness of breath o General quality of life improves, activity increases o CAT score: 18 (-10 points, significant improvement)

Phase 2: Combination with NAC (N-acetylcysteine) for a further 6 weeks

Week 8: o Mucus becomes thinner, easier to cough up o Fewer coughing stimuli, quieter nights o CAT score: 15 (-13 points)

Week 12: o Even less coughing and fewer exacerbations o Less shortness of breath, more movement possible o Better general well-being o CAT score: 13 (-15 points, from severe to moderate COPD)

Conclusions:

Inhalation with aviptadil led to a significant reduction in symptoms (-10 CAT points in 6 weeks). The additional inhalation of NAC further increased the effect (-5 more points), particularly by loosening mucus. Overall, the CAT score fell from 28 to 13, which means an improvement from severe to moderate COPD. In summary, the combination therapy with aviptadil and NAC noticeably improves breathlessness, cough, energy level and quality of life.

Example 6 - Stability of aviptadil is improved when combined with NAC:

The addition of 1 % - 10 % NAC buffered to pH 7.2 improves the stability of the 200 pg/ml aviptadil for inhalation in physiological saline solution, so that the shelf life can be extended from 6 months at +4 °C to 24 months. The addition of NAC to aviptadil leads to an increased solubility and bioavailability. Furthermore, the addition of NAC improved the homogeneity of the solution due to its mucolytic and interaction-reducing properties. As a result, aviptadil in the solution remained evenly distributed, which ensured its effectiveness over longer storage periods.

Test of stability of compositions containing aviptadil (control) and aviptadil + NAC:

Stability of the aviptadil solution with and without NAC when stored at 2 to 8° C. Determination was done by HPLC. Both solutions were adjusted to physiological pH.

Figure 2 summarizes the results of the stability measurements of aviptadil alone as well as aviptadil plus NAC over the tested period of time. After 12 and 24 months, an increased stability could be observed when aviptadil is combined with NAC in comparison to aviptadil alone.

Conclusion:

The addition of NAC to a solution containing aviptadil contributes significantly to extend the stability of aviptadil up to 24 months at the tested temperature of +4 °C.

Methods and Materials:

- Nebulizer: M-Neb® dose+ mesh nebulizer MN-300/8

- Nebulizer set (control unit, nebulizer head and mouthpiece)

- Aviptadil/NAC inhalation solution (dosage according to doctor's prescription 1.2 ml)

- Sterile water for cleaning the nebulizer head

- Clean surface for preparation and post-processing

Nebulizer Procedure:

1. Preparation:

- Wash hands thoroughly

- Place the inhaler on a stable, clean surface - Assemble the nebulizer set (control unit, nebulizer head and mouthpiece)

2. Filling the nebulizer:

- Pour the prescribed amount (1.2ml) of Aviptadil with NAC into the nebulizer chamber and close securely

3. Carrying out the inhalation:

- Switch on the inhaler

- Place the mouthpiece correctly:

Mouthpiece: Place between the lips, inhale calmly and deeply through the mouth

Caution: DO NOT exhale through the mouthpiece, but breathe past the mouthpiece

- Inhale calmly and deeply, if possible, hold the breath for a few seconds to improve absorption of the active ingredient

- Inhalation should be carried out three times a day for 10-20 minutes until there is no more visible nebulization

- It is possible to divide the inhalation with the help of a dosing finger in the nebulizer chamber (morning, noon, evening)

4. Ater inhalation:

- Clean the nebulizer head and mouthpiece according to the manufacturer's instructions and allow to dry

- Wash your hands again.

- Switch off the device, dismantle the nebulizer and allow to dry.

Claims

1. A combination of vasoactive intestinal peptide (VIP) receptor agonist and a mucolytic agent for use in the treatment or prevention of inflammatory respiratory diseases.

2. The combination according to claim 1, wherein the VIP receptor agonist is a natural VIP, VIP derivative or synthetic analog of VIP.

3. The combination according to claim 2, wherein the VIP receptor agonist is aviptadil or a functional derivative or precursor thereof.

4. The combination according to claim 2, wherein the synthetic analog of VIP is N- stearyl-[Nle17]-VIP or Ro 25-1553.

5. The combination according to any one of claims 1 to 4, wherein the mucolytic agent is any one of N-acetylcysteine (NAC), N-acystelyn (NAL), N-isobutyrylcysteine (NIC), procysteine, gluthathion ester, erdosteine, carbocysteine, fudosteine, ergothioneine.

6. The combination according to claim 5, wherein the VIP receptor agonist is aviptadil comprising the amino acid sequence set forth in SEQ ID NO: 1, and the mucolytic agent is NAC.

7. The combination according to any one of claims 1 to 6, wherein the VIP receptor agonist is provided in a first dosage form and the mucolytic agent is provided as a second dosage form, wherein a dosage regimen provides for the first dosage form to be administered to a subject concurrently with or prior to the second dosage form.

8. The combination according to any one of claims 1 to 7, wherein the combination is provided in liquid form or as an aerosol.

9. The combination according to any one of claims 1 to 8, wherein the inflammatory respiratory disease is any one of chronic obstructive pulmonary disease (COPD), pneumonia, pulmonary fibrosis, alveolitis, pneumonitis, sarcoidosis, chronic obstructive bronchitis, acute respiratory distress syndrome (ARDS), or pulmonary hypertension resulting from inflammatory disease, inflammatory viral respiratory disease, COVID-19 disease or COPD, checkpoint inhibitor pneumonitis (CIP), berylliosis, steroid resistant asthma, allergic rhinitis.

10. The combination according to claim 1, wherein the inflammatory respiratory disease is COPD mediated by neutrophil cells.

11. A pharmaceutical composition, comprising vasoactive intestinal peptide (VIP) receptor agonist and a mucolytic agent according to any one of claims 1 to 8 and a pharmaceutically acceptable carrier or diluent.

12. A pharmaceutical composition, comprising vasoactive intestinal peptide (VIP) receptor agonist and a mucolytic agent according to any one of claims 1 to 8 and a pharmaceutically acceptable carrier or diluent for use in the treatment or prevention of an inflammatory respiratory disease.

13. The pharmaceutical composition according to claim 12, wherein the inflammatory respiratory disease is any one of chronic obstructive pulmonary disease (COPD), pneumonia, pulmonary fibrosis, alveolitis, pneumonitis, sarcoidosis, chronic obstructive bronchitis, acute respiratory distress syndrome (ARDS), or pulmonary hypertension resulting from inflammatory disease, inflammatory viral respiratory disease, COVID-19 disease or COPD, checkpoint inhibitor pneumonitis (CIP), berylliosis, steroid resistant asthma, allergic rhinitis.

14. The pharmaceutical composition according to claim 12, wherein the inflammatory respiratory disease is COPD mediated by neutrophil cells.

15. The pharmaceutical composition according to any one of claims 12 to 14, wherein the composition is provided for inhalation, oral or intranasal administration.

16. A method of treatment or prevention of an inflammatory respiratory disease, in particular an inflammatory pulmonary disease, comprising the administration of a combination comprising a vasoactive intestinal peptide (VIP) receptor agonist and a mucolytic agent according to any one of claims 1 to 8 to a subject.

17. The method according to claim 16, wherein the combination is administered orally, intranasally or by inhalation.

18. The method according to according to claim 16, wherein the inflammatory respiratory disease is any one of chronic obstructive pulmonary disease (COPD), pneumonia, pulmonary fibrosis, alveolitis, pneumonitis, sarcoidosis, chronic obstructive bronchitis, acute respiratory distress syndrome (ARDS), or pulmonary hypertension resulting from inflammatory disease, inflammatory viral respiratory disease, COVID-19 disease or COPD, checkpoint inhibitor pneumonitis (CIP), berylliosis, steroid resistant asthma, allergic rhinitis.

19. The method according to claim 16, wherein the inflammatory respiratory disease is COPD mediated by neutrophil cells.

20. The method according to any one of claims 16 to 19, wherein the VIP is administered in a first dosage form and the mucolytic agent is administered as a second dosage form, wherein the first dosage form is administered to the subject concurrently with or prior to the second dosage form.

21. A method for the manufacturing of a medicament comprising the combination of a vasoactive intestinal peptide (VIP) and a mucolytic agent to a pharmaceutical composition.