ING l 1245 PCT

Agents Useful in Treating Inflammatory Bowel Disease

Field of the Invention

The invention relates to the use of modulators, in particular inhibitors of human neutrophil elastase (HNE) for the prevention, treatment or amelioration of Inflammatory Bowel Disease (IBD), in particular Ulcerative Colitis and Crohn's disease.

Background of the Invention

Inflammatory Bowel Disease (IBD ^" )

Inflammatory Bowel Disease (IBD) is a generic term that refers to chronic, non-specific disorders with unknown etiology (so-called idiopathic forms of

IBD). The two most common, but distinct forms are Ulcerative Colitis (UC) and

Crohn's disease, both being severe inflammatory diseases. The incidence of IBD varies between different geographic areas, with the highest rates in countries such as the United States, the United Kingdom, Norway and Sweden. For example, the number of people in the United States suffering from conditions of Ulcerative Colitis and Crohn's disease is one million, most of them diagnosed before age 30. IBD is a chronic condition that may flare many times during life. Non-idiopathic forms of

IBD, which mimic the pathology of Ulcerative Colitis and Crohn's disease, are caused by infections of the small intestine and the colon. Such non-idiopathic forms of IBD may be caused, e.g., by bacterial infection, viral infection, chlamydial infection, amebian infection, antibiotics, or ischemia.

The basic distinctions between Ulcerative Colitis and Crohn's disease are the location and the severity of the symptoms. In Ulcerative Colitis, sites of inflammation are colon and rectum, whereas inflammation associated with Crohn's disease is detectable anywhere from the mouth to the rectum, most commonly in the small and/or large intestines. Both diseases include extraintestinal inflammation of the skin, eyes, or joints (for a review see Harrison's Principles of Internal Medicine, 15 ^th Edition, Editors Braunwald, Fauci, Kasper, Hauser, Longo and Jameson; MacGraw-Hill Medical Publishing Division, 2001).

Depending on the severity of the disease, one or more of the following symptoms are associated with Inflammatory Bowel Diseases: inflammation of small intestine, colon and rectum, diarrhea, blood in the faeces, abdominal pain, abdominal cramping, crypt abscesses, rectal bleeding, and ulceration.

Facultative symptoms occasionally associated with IBD are fever; arthritis, in particular arthritis in the knees, ankles, wrists (in approx. 15% of UC patients); sacroiliitis of the lower spine (in approx. 10% of UC patients); ankylosing spondylitis; skin lesions, in particular pyodeπna gangrenosum and erythema nodosum (in approx. 14% of UC patients); uveitis (in approx. 2% of IBD patients; episcleritis (in approx. 4% of IBD patients); low bone density (in approx. 41% of IBD patients); liver disease, in particular sclerosing cholangitis and bile duct carcinoma; and colorectal cancer (in approx. 10% of UC patients).

Up to the present day, existing therapies cannot cure IBD. The primary goal of present drug therapy is the reduction of intestinal inflammation. Treatment is often accompanied by antibiotics (e.g., metronidazole). Currently, there are no drugs approved by the Food and Drug Administration (FDA) to treat symptoms of mild Crohn's disease. In mild to strong cases, 5-acetylsalicylic acid (5-ASA) (e.g., sulfasalazine, olsalazine) and / or corticosteroids (e.g., prednisolone, betamethasone) are administered orally. In severe cases, intravenous administration of immunosuppressors (e.g., azathioprine, cyclosporine) or surgery is necessary. Many

of the above listed drugs exhibit strong side effects and/or cannot be used in long- term therapy of IBD.

Colitis Models

A well-accepted mouse colitis model is the dextrane sulfate sodium

(DSS) model, representing an experimental model for human IBD (Mahler et al., 1998). As described by Mahler, almost all mice developed loose faeces or diarrhea during feeding of DSS. Gross evidence of blood was frequently detected in the faeces. Most mice gradually recovered clinically while receiving only water until necropsy. Histological examination revealed various degrees of typhlitis (inflammation of the cecum) and colitis in mice of all strains. Characteristic findings included mucosal ulceration, infiltration with mononuclear and polymorphonuclear leukocytes in the lamina propria and submucosa, crypt distortion, hyperplastic epithelium, and ulcers healing by reepithelialization.

Targeted mutations in a variety of mouse genes produce colitis. For example, mice homozygous for a disrupted interleukin-10 gene supported the hypothesis that a dysregulated immune response to enteric flora can trigger IBD. The severity of the colitis depends on the inbred strain background in which the disrupted gene is placed (Kuhn et al., 1993).

Recently, Hirota et al. (2004a) described a novel colitis model using

Syrian hamsters. Colitis was induced by intraco Ionic administration of acetic acid, manifested by symptoms like crypt abscess, neutrophil infiltration, hemorrhage, ulceration, erosion and edema. The duration of inflammation in this model is rather short compared to the inflammation duration in a DSS mouse colitis model.

Human Neutrophil Elastase (HNE)

IBD symptoms are accompanied by strong tissue inflammation. Neutrophils are known to be involved in triggering the immune response to the inflammation process and to release HNE during the immune response. HNE belongs to the chymotrypsin family of serine proteases, which are able to solubilize fibrous elastin. Elastin is an important matrix protein that has major structural function in lungs, arteries, skin, and ligaments. Release of HNE during inflammation is related to degradation of connective tissue and increase of vascular permeability, causing tissue damage and organ failure.

It has been postulated that HNE is involved in numerous human diseases, including emphysema, chronic obstructive pulmonary disease (COPD), cystic fibrosis, acute respiratory distress syndrome (ARDS), ischemic-reperfusion injury, rheumatoid arthritis, and severe congenital neutropenia (Moraes et al., 2003; Horwitz and Benson, 2003; Scaglioni and Pandolfϊ, 2003).

In addition, human neutrophil elastase is known for years as a marker for IBD. Gouni-Berthold et al. (1999) analyzed plasma elastase levels in patients suffering from Crohn's disease and Ulcerative Colitis. Based on the sensitivity and specificity of plasma elastase detection it was concluded that HNE is a reliable marker for IBD activity, being more suitable than other parameters of inflammation, like CRP or ESR. However, treatment of IBD by targeting human neutrophil elastase with inhibitors of HNE has, to our knowledge, not been suggested in the prior art.

Inhibitors of Human Neutrophil Elastase (HNEI

In recent years, HNE has been a target of extensive research and several attempts were made to develop inhibitors with potential function in the treatment of respiratory diseases, such as acute respiratory distress syndrome

(ARDS), cystic fibrosis, chronic obstructive pulmonary disease (COPD), acute and chronic bronchitis, and pulmonary emphysema. A number of HNE inhibitors have

entered the preclinical and clinical phases for the treatment of these medical indications, for example ONO-5046, ONO-6018, ZD-8321, ZD-0982, and EPI-hNE4. These HNE inhibitors and their use in the above-mentioned indications have been described in several publications, patents and patent applications, as described below.

WO 02/20774 and US 6,552,082 inter alia disclose the HNE inhibitor

ONO-5046 for use in the treatment of acute pulmonary disorders. ONO-5046 has been launched in Japan under the Trademark Elaspol® for the treatment of acute lung injury associated with systemic inflammatory response syndrome (SIRS). In view of its low bio-availability, Elaspol must be administered intravenously.

WO 02/20516 inter alia discloses the HNE inhibitor ONO-6818 for the treatment of arthritis, periodontal disease, nephritis, dermatitis, psoriasis, cystic fibrosis, chronic bronchitis, atherosclerosis, Alzheimer's disease, organ transplantation, corneal ulcers, and malignant tumors. Phase II trials for COPD have been suspended in the USA and in Japan due to abnormal elevation of liver function test (LFT) values. Hirota et al. (2004a) briefly mentioned that ONO-6818 may exhibit an inhibitory effect on colonic NE activity and ulcer area in acetic acid induced colitis in Syrian hamster. However, no details or data are shown in this publication. In a subsequent publication, Hirota et al. (2004b) published experimental results pertaining to administration of ONO-6818. In these experiments, ONO-6818 was administered orally prior to the colitis induction. Since the duration of inflammation is too short in this colitis model, the therapeutic effectiveness of ONO-6818 by post- treatment after colitis induction could not be evaluated in this animal model. In any event, it remains crucial for a clinical application of the HNE inhibitors that the inhibitory effect on colitis is shown by post-treatment after colitis induction.

WO 96/23812 discloses ZD-8321 for the treatment of ARDS, rheumatoid arthritis, atherosclerosis, pulmonary emphysema and airway inflammatory diseases characterized by increased and abnonnal airway secretion, such as acute and chronic bronchitis and cystic fibrosis. ZD-8321 had been in development for the potential treatment for ARDS and bronchitis. The development

of this compound was discontinued in August 2000 after having entered phase II trials for ARDS.

HNE inhibitor ZD-0892 is disclosed in US 6,037,363 for its use in the treatment of acute and chronic bronchitis, pulmonary emphysema, reperfusion injury, ARDS, cystic fibrosis and peripheral vascular disease (PVD). Development of ZD- 0892 was discontinued during phase I clinical trials of ZD-0892 for potential treatment of COPD and PVD.

US 4,910,190 discloses a method for preparation of compounds ZD- 8321 and ZD-0892, and their use in the treatment of pulmonary emphysema, atherosclerosis, rheumatoid arthritis, and osteo arthritis. ZD-0892 is specifically disclosed as Example 20.

WO 96/20278 and WO 02/36158 disclose a non-naturally occurring protein EPI-hNE4, comprising a genetically engineered aprotinin-like Kunitz domain that binds and inhibits HNE; its production and its use in an inhalable pharmaceutical formulation for the treatment of cystic fibrosis, emphysema, ARDS, and COPD.

WO 96/32152 and US 5,780,014 relate to an inhibitory glycoprotein which functions as an inhibitor of alphai -proteinase. In 2003, the FDA approved Zemaira™ for chronic augmentation and maintenance therapy in adults with alphai- proteinase inhibitor deficiency and clinical evidence of lung emphysema.

WO 98/27998 claims another HNE inhibitor named FK-706 and its use for the treatment of cerebral ischemic diseases. Further patent applications relate to the use of FK-706 in the prevention of skin aging (see WO 99/43352) and in the treatment of refractory injuries (see WO 01/26685). Phase II clinical trials in Western Europe and in Japan for potential treatment of lung emphysema and ARDS were discontinued in 1999.

As described above, the current therapeutic approaches for the treatment of Inflammatory Bowel Disease (IBD) are not satisfactory, not the least because the existing therapies give rise to severe side effects, thereby preventing long-term

treatment of the disease that is necessary in view of the chronic nature of the disease. Accordingly, there is an urgent need in the art for novel therapeutic strategies for the treatment of IBD, and in particular for the treatment of Ulcerative Colitis and Crohn's disease.

Summary of the Invention

The inventors of the present invention have surprisingly found that small molecule compounds known to be inhibitors of human neutrophil elastase (HNE) are suitable for preventing, treating or ameliorating a medical condition generally referred to as Inflammatory Bowel Disease (IBD).

Accordingly, in a first aspect the invention relates to the use of a small molecule human neutrophil elastase inhibitor for the preparation of a pharmaceutical composition for the prevention, treatment or amelioration of Inflammatory Bowel Disease (IBD). In one embodiment of this aspect of the present invention, the pharmaceutical composition may further comprise at least one other pharmaceutically active compound known to be effective in the treatment of IBD.

In another aspect, the invention relates to pharmaceutical compositions comprising at least one small molecule human neutrophil elastase inhibitor and further comprising at least one other pharmaceutically active compound known to be effective in the treatment of IBD.

In yet another aspect, the invention relates to a method for the prevention, treatment or amelioration of Inflammatory Bowel Disease (IBD) in a mammal, particularly in a human subject, in need thereof, said method comprising administering to said mammal or said human subject a pharmaceutical composition comprising at least one small molecule human neutrophil elastase inhibitor or a combination of said human neutrophil elastase inhibitor with at least one other HNE inhibitor, or a composition comprising at least one HNE inhibitor together with at

least one other pharmaceutically active compound known to be effective in the treatment of IBD as referred to above.

Description of the Figures

Figure 1 summarizes the therapeutic effect of ONO-5046 and prednisolone in the colon of dextran sulfate sodium (DSS) induced colitis mice. Mice were treated with DSS supplied to the drinking water for 5 days, followed by supply of normal water. DSS treatment led to colitis-like symptoms such as diarrhea and blood in the faeces. Over a period of 14 days, each group (N=IO) of the DSS-treated mice received an intraperitoneal application of either the HNE inhibitor ONO-5046 (ONO- 5046), the corticosteroid prednisolone (prednisolone; a well-known drug used for IBD treatment), or isotonic NaCl solution without a pharmaceutically active compound (NaCl control) twice a day. 5 mice without DSS treatment and without administration of the pharmaceutically active compounds served as an additional control (untreated control). After day 14 of compound administration, all mice were sacrificed and histopathological analysis of each colon was performed. Analysis comprised examination and grade scoring of colon submucosa , inflammatory infiltration, of regeneration of former ulceration, and of proliferation of lymphatic aggregates. Scoring was performed based on different grade levels from 0 to 5, according to the grade list provided in the Annex to Example 1. The higher the number, the more severe are the symptoms observed. The scoring numbers from all mice belonging to one treatment group were summed. As shown in Figure 1, ONO- 5046 significantly reduced the degree of colon submucosa inflammation (scoring number: 13), similar to prednisolone (scoring number: 14) and in contrast to NaCl control mice (scoring number: 23) without active compound treatment. DSS- untreated mice are inconspicuous (scoring number: 2). Regeneration of former ulceration by reepithelization was strongly promoted by ONO-5046 (scoring number: 3), whereas no such effect was observed in prednisolone treated mice (scoring number: 12) and in NaCl control mice (scoring number: 14). Proliferation of

lymphatic aggregates in the colon mucosa was not affected by ONO-5046 (scoring number: 18) when compared to NaCI control mice (scoring number: 20). In contrast, prednisolone significantly reduced the degree of proliferation of lymphatic aggregates (scoring number: 10). In sum, the overall therapeutic effect in the treatment of colitis is very similar, according to the total scoring numbers received from ONO-5046 (sum scoring number: 34) and prednisolone (sum scoring number: 36).

Detailed Description of the Invention

General Definitions

The pharmaceutically active compounds that are used in the present invention according to the first aspect of the invention are small molecule inhibitors of the protease human neutrophil elastase (HNE, EC_3.4.21.37). The term "inhibitor" of HNE refers to compounds that by any possible mechanism inhibit the activity of this protease, either by direct interaction with HNE, e.g., by binding to or modifying HNE or by triggering certain events that lead to an inactivity of HNE. In a preferred embodiment, the compounds of the invention inhibit HNE activity by directly interacting with, or binding to HNE.

The term "small molecule" refers to compounds having a molecular weight of no more than 2000 Dalton, or no more than 1500 Dalton. Smaller molecules having a molecular weight of no more than 1000, 500, 400, 300 or even 200 Dalton which display the above property of inhibiting HNE activity are likewise suitable in the context of the present invention. Such small molecules may include small peptides, peptide analogs or peptidomimetics provided they do not exceed the molecular weight limitation defined above. It will be understood that whenever it is referred to "inhibitors of HNE", such inhibitors belong to the class of small molecules as defined above, even if the term "inhibitor" is used without any further qualification throughout the specification and the claims of the present invention.

The mechanisms by which the inhibitors of the present invention may inhibit the activity of HNE comprise all mechanisms known by the person skilled in the art, e.g., competitive or ailosteric inhibition; posttranslational modifications, such as phosphorylation, tyrosinylation, methylation, amidation, sulfatation, acylation, myristoylation, palmitation, farnesylation, glycosylations, cleavage of a proprotein, cleavage of a signal sequence or inhibition of the reversal of such posttranslational modifications; cellular mistargeting, such as the targeting of HNE to inappropriate cellular compartments; degradation of HNE, or inhibiting or prolonging protein interaction between HNE and third molecules.

Furthermore, the inhibitors of the present invention are useful not only in the treatment of IBD, preferably Ulcerative Colitis and Crohn's Disease, but they are also active in preventing these diseases, or in ameliorating (i.e., alleviating the symptoms of) these diseases. Therefore, for the purposes of the present invention, whenever it is referred hereinbelow to the "treatment" of IBD, and in particular of Ulcerative Colitis and Crohn's Disease, this also implies "prevention" and "amelioration" of said diseases.

In the context of the present invention, the teπns "Inflammatory Bowel Disease" or "IBD" are intended to refer to medical conditions generally referred to as Inflammatory Bowel Disease (IBD), as well as Ulcerative Colitis and Crohn's disease. The terms "Inflammatory Bowel Disease" or "IBD" are meant to include idiopathic disease forms of chronic intestinal inflammation. The term "idiopathic" in connection with the above-mentioned diseases is intended to mean that the cause(s) for said diseases are not known or have not been definitively elaborated. The most common forms of idiopathic IBD are known as Ulcerative Colitis and Crohn's disease.

Furthermore, the teπns "Inflammatory Bowel Disease" or "IBD" as used in the present invention also refer to non-idiopathic disease forms of IBD. The pathologies of these disease forms mimic those of Ulcerative Colitis and Crohn's disease. They may be caused by infections, like bacterial infection, viral infection,

chlamydial infection, amebian infection, or due to side effects of antibiotics, or due to ischemia, e.g. ischemic enteritis. In the context of the present invention, it will be understood that when reference is made to Ulcerative Colitis and Crohn's disease, these terms are intended to include in their meaning non-idiopathic Intestinal Bowel Diseases that closely resemble the pathology of Ulcerative Colitis and Crohn's disease.

Finally, whenever the terms "Inflammatory Bowel Disease" or "IBD" are used in the present invention, it is contemplated that the use of these terms not only includes inflammatory bowel disease in a strict sense such as Crohn's disease and Ulcerative Colitis, but also lesions of the intestine accompanying Behcet's disease, hemorrhagic rectal ulcer, ileum pouchitis, intestinal tuberculosis, drug- induced colitis, radiation enteritis, etc.

With respect to all of the pharmaceutically active compounds of the invention mentioned herein, it is contemplated that the present invention relates to both the use of these compounds for the preparation of a pharmaceutical composition (or medicament) for the prevention, treatment or amelioration of Inflammatory Bowel Disease, and in particular of Ulcerative Colitis and Crohn's Disease, as well as to methods for preventing, treating, or ameliorating said medical conditions, characterized in that the compounds or pharmaceutical compounds comprising such compounds are administered to a patient in need thereof. Thus, all preferred embodiments of said use and said method of the invention both analogously relate to this aspect of the present invention, even if this is not always expressly mentioned in the detailed description below.

Any compounds of the present invention containing one or more asymmetric carbon atoms may occur as racemates and racemic mixtures, single enantiomers, diastereomeric mixtures and individual diastereomers. Unless otherwise indicated, all such isomeric forms of these compounds are expressly included in the present invention. Each asymmetric carbon atom may be in the R or S configuration. Suitable methods of separating racemates into the individual enantiomers, such as

chiral chromatography and others, are known to the skilled person. Some of the compounds of the invention can exist in more than one tautomeric form. The invention includes all such tautomers.

The compounds of the invention may also be present in the form of a prodrug. Prodrugs include those compounds that, upon simple transformation, are modified to produce the compounds that are expressly disclosed in this application. Simple chemical transformations include hydrolysis, oxidation and reduction which occur enzymatically, metabolically or otherwise. Prodrugs of the compounds of the invention may have attached thereto one or more functional groups which facilitate e.g. the use of the compound as a drug in the body, e.g. by facilitating entry into cells etc. Such functional "prodrug moiety" may be cleaved from the compound by said simple chemical transformations. Prodrug moieties include phosphate groups, peptide linkers, sugars and others. When a prodrug of this invention is administered to a patient, the prodrug may be transformed into a compound as disclosed hereinbelow, thereby imparting the desired pharmacological effect.

The compounds used in the present invention as described hereinbelow can also be targeted for specific delivery to a certain cell type to be treated by conjugation of the compounds to a targeting moiety, such as antibodies, cytokines and receptor ligands that are specific to the cell to be treated.

Whenever it is referred to a "pharmaceutically acceptable salt, analog, or derivative" for the purposes of the present invention, it is referred to any pharmaceutically acceptable acid, salt or ester of a compound of the present invention. Pharmaceutically acceptable salts include those derived from pharmaceutically acceptable inorganic and organic acids and bases. Examples of suitable acids include hydrochloric, hydrobromic, sulphuric, nitric, perchloric, fumaric, maleic, phosphoric, glycolic, lactic, salicylic, succinic, oxalic, toluene-p- sulfonic, tartaric, acetic, citric, methanesulfonic, formic, benzoic, malonic, naphthalene-2-sulfonic, benzenesulfonic acids. Salts derived from appropriate bases include alkali metal (e.g., lithium, sodium, potassium), alkaline earth metal (e.g.,

magnesium, calcium), ammonium and N-(Ci- ₄ alkyl) ₄ , and organic amine salts such as morpholine, piperidine, dimethylamine or diethylamine salts. Analogs or derivatives of the compounds of the invention may have variations in the chemical structure that essentially do not change the inhibitory properties of the compounds of the present invention. Since the 3-D crystal structure of human neutrophil elastase has been determined (see, e.g., Bode et al., 1986, where the structure of HNE is described in the presence of a transition state analog) and suitable assays to determine HNE activity are available (cf. ,e.g., Yoshimura et al., 1994), it is possible for those of skill in the art to identify positions in the inhibitor molecule which may be derivatized without affecting the inhibitory properties of the compound.

As indicated above in the section entitled "Summary of the Invention", the first aspect of the invention relates to the use of small molecule inhibitors of human neutrophil elastase as described in detail below for the preparation of a pharmaceutical composition for the prevention, treatment or amelioration of Inflammatory Bowel Disease.

Inhibitors of HNE are known in the art. The major types of HNE inhibitors known in the art are electrophilic ketone inhibitors, transition-state inhibitors and enzyme-acylating agents. Thus, in a preferred embodiment of this aspect of the present invention, the HNE inhibitors to be used in the present invention may be selected from the group of electrophilic ketone inhibitors, transition-state inhibitors or enzyme-acylating agents.

Particularly preferred HNE inhibitors according to the present invention are trifluoromethyl ketone substituted peptide derivatives according to formula (Ia) or (Ib) as set out below, including their pharmaceutically acceptable salts , as described in US 4,910,190:

wherein

Rl is an alkyl group containing from 1 to 5 carbon atoms. Preferably, Rl contains from 2 to 5 carbons, and most preferably 3 carbon atoms; R2 is selected from the group consisting of:

(I) an alkyl group containing from 1 to 10 carbons, preferably from 2 to 5 carbons, and most preferably 3 carbon atoms;

(II) an alkyl group containing from 1 to 6 carbon atoms substituted by at least one member selected from the group consisting of:

(a) hydroxy;

(b) amino; (c) alkylamino containing from 1 to 6 carbons;

(d) dialkylamino wherein each alkyl group contains from 1 to 6 carbons;

(e) alkanoyl containing from 1 to 6 carbons;

(f) arylcarbonyl wherein the aryl contains from 5 to 14 carbons ;

(g) aralkanoyl containing 8 to 13 carbons; (h) amido which may be attached to the alkyl group via either a nitrogen or carbon of said amido;

(i) alkylcarbonylamino wherein the alkyl group contains from 1 to 6 carbons;

(j) alkylaminocarbonyl wherein the alkyl group contains from 1 to 6 carbons;

(k) arylcarbonylamino wherein the aryl group contains from 5 to 14 carbons;

(1) aralkylcarbonylamino wherein the aralkyl group contains from 7 to 15 carbons;

(m) arylaminocarbonyl wherein the aryl group contains from 5 to 14 carbons;

(n) aralkylaminocarbonyl wherein the aralkyl group contains from 7 to 15 carbons;

(o) carboxy; (p) aryloxycarbonyl wherein the aryl group contains from 5 to 14 carbons;

(q) aralkoxycarbonyl wherein the aralkoxy group contains from 7 to 15 carbons;

(r) alkanoyloxy containing from 1 to 6 carbons;

(s) aroyloxy wherein the aryl portion contains from 5 to 14 carbons; (t) aralkanoyloxy containing from 8 to 16 carbons;

(u) alkylsulfonamido wherein the alkyl group contains from 1 to 6 carbons;

(v) aralkylsulfonamido wherein the aralkyl group contains from 7 to 15 carbons;

(w) arylsulfonamido wherein the aryl group contains from 5 to 14 carbons; (x) acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) (1 to

15 carbons) including acylsulfonamido wherein the acyl group contains 1 to 7 carbons when it is the terminal portion of the acylsulfonamide and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro; (y) alkoxycarbonyl wherein the alkoxy group contains from 1 to 6 carbons;

(z) aralkoxycarbonylamino containing from 8 to 16 carbons (e.g., benzyloxycarbonyl amino);

(aa) aryloxycarbonylamino wherein the aryloxy group contains from 5 to 14 carbons; (bb) alkoxycarbonylamino wherein the alkyloxy group contains from 1 to 6 carbons;

(cc) aryl containing from 5 to 14 carbons (e.g., phenyl, biphenyl, naphthyl);

(dd) aryl containing from 5 to 14 carbons and substituted by 1 to 3 members selected from the group consisting of chloro, bromo, iodo, fluoro, trifluoromethyl, hydroxy, alkyl (1 to 6 carbons), alkoxy (1 to 6 carbons), alkoxycarbonyl (1 to 6 carbons), carboxy, 5-tetrazolo, and acylsufonamido

(i.e. acylaminosulfonyl and sulfonylaminocarbonyl) (1 to 15 carbons) and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro; (ee) cycloalkyl containing from 3 to 15 carbons (e.g., cyclohexyl, adamantyl, norbornyl);

(ff) alkylureido wherein the alkyl group contains from 1 to 6 carbons;

(gg) aralkylureido containing from 8 to 16 carbons;

(hh) arylureido wherein the aryl group contains from 5 to 14 carbons; and (III) an aryl group of 6 carbons, e.g. phenyl;

R3 is selected from the group consisting of:

(I) an alkyl group containing from 1 to 12 carbons, preferably from 1 to 6 carbons, and most preferably 1, 2, 3, 4, or 5 carbon atoms;

(II) an alkyl group containing from 1 to 12 carbons and from 1 to 4 hetero atoms each of which is selected independently from the group consisting of nitrogen and oxygen;

(III) an alkyl group containing from 1 to 12 carbons and, optionally, 1 to 4 hetero atoms each of which is selected independently from the group consisting of nitrogen and oxygen, and substituted on at least one of carbon or nitrogen by 1 to 3 members selected independently from the group consisting of: For carbon:

(a) hydroxy, provided that it may not be on a carbon directly bonded to A;

(b) amino, provided that it may not be on a carbon directly bonded to A; (c) alkylamino containing from 1 to 6 carbons, provided that it may not be on a carbon directly bonded to A;

(d) dialkylamino wherein each alkyl group contains from 1 to 6 carbons, provided that it may not be on a carbon directly bonded to A;

(e) alkanoyl containing from 1 to 6 carbons;

(f) arylcarbonyl wherein the aryl contains from 5 to 14 carbons; (g) aralkanoyl containing 8 to 16 carbons;

(h) amido which may be attached to the alkyl group via either a nitrogen or carbon of said amido;

(i) alkylcarbonylamino wherein the alkyl group contains from 1 to 6 carbons; (j) alkylaminocarbonyl wherein the alkyl group contains from 1 to 6 carbons; (k) arylcarbonylamino wherein the aryl group contains from 5 to 14 carbons;

(k)-(l) arylcarbonylamino wherein the aryl group contains from 5 to 14 carbons and is substituted by a member selected from carboxy, alkoxycarbonyl, where alkoxy is 1 to 3 carbons, carboxyalkylaminocarbonyl wherein the alkyl group contains 1 to 6 and preferably 1 carbon(s), 5- tetrazolo, and acylsulfonamido (i.e. acylaminosulfonyl and sulfonylaminocarbonyl) containing 1 to 15 carbons and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro; (1) aralkylcarbonylamino wherein the aralkyl group contains from 7 to 15 carbons;

(I)-(I) aralkylcarbonylamino wherein the aralkyl group contains 7 to 15 carbons and is substituted on the aryl portion by a member selected from carboxy, alkoxycarbonyl, where the alkoxy has 1 to 3 carbons, 5-tetrazolo, and acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) containing 1 to 15 carbons and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro;

(m) arylaminocarbonyl wherein the aryl group contains from 5 to 14 carbons ; (n) aralkylaminocarbonyl wherein the aralkyl group contains from 7 to 15 carbons;

(o) carboxy;

(p) aryloxycarbonyl wherein the aryl group contains from 5 to 14 carbons;

(q) aralkoxycarbonyl wherein the aralkoxy group contains from 7 to 15 carbons;

(r) alkanoyloxy containing from 1 to 6 carbons; (s) aroyloxy wherein the aryl portion contains from 5 to 14 carbons;

(t) aralkanoyloxy containing from 8 to 16 carbons;

(u) alkylsulfonamido wherein the alkyl group contains from 1 to 6 carbons;

(u)-(l) cycloalkylsulfonamido wherein the cycloalkyl portion contains 3 to 15 carbons (e.g., cyclohexyl, adamantyl, norbornyl); (v) aralkylsulfonamido wherein the aralkyl group contains from 7 to 15 carbons;

(w) arylsulfonamido wherein the aryl group contains from 5 to 14 carbons ;

(x) acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) (1 to

15 carbons) including acylsulfonamido wherein the acyl group contains 1 to 7 carbons when it is the terminal portion of the acylsulfonamide, and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro;

(y) alkoxycarbonyl wherein the alkoxy group contains from 1 to 6 carbons;

(z) aralkoxycarbonylamino containing from 8 to 16 carbons (e.g., benzyloxycarbonylamino);

(z)-(l) aralkylaminocarbonyloxy containing 8 to 16 carbons;

(z)-(2) aryloxy wherein the aryl contains from 5 to 14 carbons ;

(z)-(3) aryloxy wherein the aryl contains from 5 to 14 carbons and is substituted by a member selected from aminocarbonyl, aminocarbonylalkyl where the alkyl has 1 to 3 carbons, alkoxycarbonyl having 2 to 4 carbons, and carboxy;

(aa) aryloxycarbonylamino wherein the aryloxy group contains from 5 to 14 carbons;

(aa)-(l) arylaminocarbonyloxy wherein the aryl group contains from 5 to 14 carbons;

(bb) alkoxycarbonylamino wherein the alkyloxy group contains from 1 to 6

carbons;

(bb)-(l) alkoxycarbonylamino wherein the alkoxy group contains 1 to 6 carbons and is optionally bonded on carbon to a carbon of an aromatic heterocyclic group as described in (gg) under R3 ; (bb)-(2) alkoxycarbonylamino wherein the alkoxy group contains 1 to 6 carbons substituted by an aliphatic heterocyclic group as described in (ff) under R3 ;

(bb)-(3) aryloxyalkylcarbonylamino wherein the aryl contains from 5 to 14 carbons, preferably 6 or 10 carbons and the alkyl has 1 to 6 carbons; (bb)-(4) alkylaminocarbonyloxy wherein the alkyl group contains 1 to 6 carbons;

(cc) aryl containing from 5 to 14 carbons carbons (e.g., phenyl, naphthyl, biphenyl); (cc)-(l) aryloxy containing from 5 to 14 carbons carbons; (dd) aryl containing from 5 to 14 carbons carbons and substituted by 1 to 3 members independently selected from the group consisting of chloro, bromo, iodo, fluoro, hydroxy, trifluoromethyl, alkyl (1 to 6 carbons), alkoxy (1 to 6 carbons), alkoxycarbonyl (2 to 6 carbons), carboxy, aminocarbonylalkyl (2 to 6 carbons), aminocarbonyl, 5-tetrazolo, and acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) (1 to 15 carbons), and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro;

(dd)-(l) aryloxy containing from 5 to 14 carbons and substituted on carbon by 1 to 3 members independently selected from the group consisting of chloro, bromo, iodo, fluoro, hydroxy, trifluoromethyl, alkyl (1 to 6 carbons), alkoxy (1 to 6 carbons), alkoxycarbonyl (2 to 6 carbons), carboxy, aminocarbonylalkyl (2 to 6 carbons), aminocarbonyl, 5-tetrazolo, acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) (1 to 15 carbons) and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo,

iodo and nitro;

(ee) cycloalkyl containing from 3 to 15 carbons (e.g., cyclohexyl, adamantyl, norbornyl);

(ee)-(l) cycloalkyloxy containing from 3 to 15 carbons; (ff) an aliphatic heterocyclic group of at least 4 atoms containing from 1 to 5 carbons and from 1 to 4 hetero atoms each of which is selected independently from the group consisting of nitrogen and oxygen (e.g., morpholine, piperazine), wherein the aliphatic heterocyclic group may optionally contain 1 or 2 double bond(s), which aliphatic heterocyclic group may be substituted at any nitrogen with an alkyl group containing from 1 to 6 carbons, an alkanoyl group containing from 1 to 6 carbons, an aryloxycarbonyl group wherein the aryl group contains 6, 10 or 12 carbon atoms, an aralkyloxycarbonyl group wherein the aralkyl group contains from 7 to 15 carbons or an alkoxycarbonyl group wherein the alkyl group contains from 1 to 6 carbons; (££)-( 1) an aliphatic heterocyclic oxy group wherein the oxy link is bonded directly to a carbon atom of the aliphatic heterocyclic group of at least 5 atoms containing from 1 to 5 carbons and from 1 to 4 hetero atoms each of which is selected independently from the group consisting of nitrogen and oxygen, (e.g., morpholine, piperazine), wherein the aliphatic heterocyclic group may optionally contain 1 or 2 double bond(s), which aliphatic heterocyclic group may be substituted at any nitrogen with an alkyl group containing from 1 to 6 carbons, an alkanoyl group containing from 1 to 6 carbons, an aryloxycarbonyl group wherein the aryl group contains from 5 to 14 carbons, an aralkyloxycarbonyl group wherein the aralkyl group contains from 7 to 15 carbons or an alkoxycarbonyl group wherein the alkyl group contains from 1 to 6 carbons;

(gg) an aromatic heterocyclic group of from 4 to 15 carbons and from 1 to 4 heteroatoms each of which is selected independently from the group consisting of sulfur, nitrogen and oxygen and which form 1 to 3 five or six- membered rings at least one of which is aromatic, and optionally, wherein up to 3 carbons of the aromatic ring(s) may be substituted with a member of the

group consisting of fluoro, chloro, bromo, iodo, trifluoromethyl, alkyl containing from 1 to 6 carbons, alkoxy containing from 1 to 6 carbons, alkanoyl containing from 2 to 6 carbons, carboxy, aminocarbonylalkyl (2 to 6 carbons) and aminocarbonyl, and provided further that any nitrogen atom may be substituted by an alkyl group containing from 1 to 6 carbon atoms;

(gg)-(l) an aromatic heterocyclic oxy group wherein the oxy link is bonded directly to a carbon of an aromatic heterocyclic group of from 5 to 15 carbons and from 1 to 4 heteroatoms each of which is selected independently from the group consisting of sulfur, nitrogen and oxygen and which form 1 to 3 five or six-membered rings at least one of which is aromatic, and optionally, wherein up to 3 carbons of the aromatic ring(s) may be substituted with a member of the group consisting of fluoro, chloro, bromo, iodo, trifluoromethyl, alkyl containing from 1 to 6 carbons, alkoxy containing from 1 to 6 carbons, alkanoyl containing from 2 to 6 carbons, carboxy, aminocarbonylalkyl (2 to 6 carbons) and aminocarbonyl, and provided further that any nitrogen atom may be substituted by an alkyl group containing from 1 to 6 carbons; (hh) alkylureido wherein the alkyl group contains from 1 to 6 carbon atoms; (hh)-(l) cycloalkylureido wherein the cycloalkyl group contains 3 to 15 carbons; (ii) aralkylureido wherein the aralkyl group contains from 7 to 15 carbons;

(jj) arylureido wherein the aryl group contains from 5 to 14 carbons; Qj)-(I) arylureido wherein the aryl group contains from 5 to 14 carbons and is substituted by 1 to 3 members selected independently from the group consisting of chloro, bromo, iodo, fluoro, hydroxy, trifluoromethyl, alkyl (1 to 6 carbons), alkoxy (1 to 6 carbons), alkoxycarbonyl (2 to 6 carbons), carboxy, aminocarbonylalkyl (2 to 6 carbons), aminocarbonyl, 5-tetrazolo, and acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) (1 to 15 carbons) including acylsulfonamido wherein the acyl group contains 1 to 7 carbons when it is the terminal portion of the acylsulfonamide and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro;

For nitrogen:

(a) alkyl of 1 to 3 carbons;

(b) alkanoyl containing from 2 to 6 carbon atoms;

(c) arylcarbonyl wherein the aryl contains from 5 to 14 carbons, preferably 6, 10 or 12 carbons;

(d) aralkanoyl containing 8 to 14 carbons;

(e) formyl;

(f) an aliphatic heterocyclic group wherein the amino link is bonded directly to a carbon of an aliphatic heterocyclic group defined in (ff) for the carbon substituents;

(g) an aromatic heterocyclic group wherein the amino link is bonded directly to a carbon of the aromatic heterocyclic group defined in (gg) for the carbon substituents;

(IV) an aryl group containing from 5 to 14 carbons, preferably containing 5, 6, 10, 12 or 14 carbons;

(V) an aryl group containing from 5 to 14 carbons, preferably containing 5, 6, 10, 12 or 14 carbons, suitably substituted by 1 to 3 members selected independently from the group consisting of fluoro, chloro, bromo, iodo, trifluoromethyl, hydroxy, alkyl containing from 1 to 6 carbons, alkoxy containing from 1 to 6 carbons, alkoxycarbonyl containing from 2 to 6 carbons, carboxy, alkylcarbonylamino wherein the alkyl group contains 1 to 6 carbons, carboxyalkylaminocarbonyl wherein the alkyl group contains from 1 to 6 and preferably 1 carbon(s), 5-tetrazolo, and acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) containing from 1 to 15 carbons, and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro;

(Va) an aralkyl group wherein the aryl moiety contains from 5 to 14 carbons, preferably containing 5, 6, 10, 12 or 14 carbons, and the alkyl moiety contains from 1 to 6 carbons, suitably substituted by 1 to 3 members selected independently from the group consisting of fluoro, chloro, bromo, iodo,

trifluoromethyl, hydroxy, alkyl containing from 1 to 6 carbons, alkoxy containing from 1 to 6 carbons, alkoxycarbonyl containing from 2 to 6 carbons, carboxy, alkylcarbonylamino wherein the alkyl group contains 1 to 6 carbons, 5-tetrazolo, and acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) containing from 1 to 15 carbons, and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro;

(VI) a cycloalkyl group containing from 3 to 15 carbons (e.g., cyclohexyl, adamantyl, norbornyl); (VI)-(I) a cycloalkyl group containing from 3 to 15 carbons (e.g., cyclohexyl, adamantyl, norbornyl), substituted by a member selected from the group consisting of carboxy and alkoxycarbonyl wherein the alkoxy group contains 1 to 4 carbons;

(VII) an aliphatic heterocyclic group of at least 5 atoms containing from 1 to 5 carbons and from 1 to 4 hetero atoms each of which is selected independently from the group consisting of nitrogen and oxygen, (e.g., morpholine, piperazine) which may be substituted at any nitrogen with a member selected from the group consisting of an alkyl group containing from 1 to 6 carbon atoms, an alkanoyl group containing from 1 to 6 carbons, an aryloxycarbonyl group wherein the aryl group contains 6, 10 or 12 carbons, an aralkoxycarbonyl group wherein the aralkyl group contains from 7 to 15 carbons and an alkoxycarbonyl group containing from 2 to 7 carbons, provided that when A is OCO or NHCO then A must be bonded to a carbon atom of the aliphatic heterocyclic group; (VIII) an aromatic heterocyclic group of from 5 to 15 carbons and from 1 to 4 heteroatoms each of which is selected independently from the group consisting of sulfur, nitrogen and oxygen, and which form 1 to 3 five or six- membered rings at least one of which is aromatic, and optionally, wherein up to 3 carbons of the aromatic ring(s) may be substituted at any carbon atom with a member of the group consisting of fluoro, chloro, bromo, iodo, trifluoromethyl, alkyl containing from 1 to 6 carbons, alkoxy containing from

1 to 6 carbons, alkanoyl containing from 2 to 6 carbons, carboxy, carboxyalkyl containing from 2 to 7 carbons, and provided further that any nitrogen may be substituted by an alkyl group containing from 1 to 6 carbons and optionally substituted at any carbon atom with a member of the group consisting of fluoro, chloro, bromo, iodo, trifluoromethyl, alkyl containing from 1 to 6 carbons, alkoxy containing from 1 to 6 carbons, alkanoyl containing from 2 to 6 carbons, carboxy, carboxyalkyl containing from 2 to 7 carbons, provided that when A is OCO or NHCO then A must be bonded to a carbon of the aromatic heterocycle; (IX) an alkenyl group of 2 to 10 carbons, having at least one double bond;

(X) an alkenyl group of 2 to 10 carbons, having at least one double bond and substituted by a member selected from the group consisting of

(a) aryl of from 5 to 14 carbons, preferably 6 or 10 carbons;

(b) aryl of from 5 to 14 carbons, preferably 6 or 10 carbons, substituted by 1 to 3 members selected independently from the group consisting of chloro, bromo, iodo, fluoro, hydroxy, trifluoromethyl, alkyl (1 to 6 carbons), alkoxy (1 to 6 carbons), alkoxycarbonyl (2 to 6 carbons), carboxy, 5-tetrazolo, and acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) (1 to 15 carbons) and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro; and

(c) ureidocarbonyl;

(XI) a hydroxy group, provided that it is directly bonded to a carbon or sulfur atom of residue A;

R4 is selected from hydrogen or alkyl; n is 0, 1 or 2, preferably n is 1; and A is selected from the group consisting of

O O O O

Il Il Il Il

— C— > — N— C— ' — O— C— ' — S— ; I Il

H O

and, where appropriate, the pharmaceutically acceptable acid- and base- addition salts thereof.

In the HNE inhibitors as defined under formulas (Ia) and (Ib) above, Rl is preferably a lower alkyl from 1 to 6 carbon atoms, wherein the alkyl group may be straight chain or branched-chain. More preferably, the alkyl group contains 1, 2, 3, or 4 carbons. Most preferably, Rl is a 2-propyl (i.e., isopropyl) group.

R2 in formula (Ib) above is preferably a lower alkyl from 1 to 6 carbon atoms wherein the alkyl group may be straight chain or branched-chain. More preferably, the alkyl group contains 1, 2, 3, or 4 carbons. Most preferably, R2 is a 2-propyl (i.e. ₅ isopropyl) group.

For the HNE inhibitors according to formula (Ia), R3 is preferably an alkyl group substituted on the carbon atom which is attached to group A with one subtituent as defined above under (III) (a) to Qj)-I. More preferably, the substituent is an alkyl- or arylcarbonylamino group as defined above and is attached to the carbon atom that is also attached to group A. Particularly preferred compounds according to formula (Ia) are those wherein R3 is a l-(phenylcarbonylamino-propan-2-methyl-l-yl group (i.e., attached at the 1-position to group A, wherein the phenyl ring may optionally be substituted as defined above under (V) and wherein A is preferably a carbonyl group.

For the HNE inhibitors according to formula (Ib), R3 is preferably an alkyl group as defined above under (I) and (II) or an aryl group as defined above under (V) for fonnulas (Ia) and (Ib). In these embodiments, A is preferably —CO- or -OCO-, and R4 is hydrogen or methyl.

Thus, particularly preferred HNE inhibitors of the present invention have the following formulas:

wherein R5 is a straight or branched-chain alkyl or alkoxy group containing from 1 to about 6 carbons. Most preferably, R5 is methyl, ethyl, propyl, or butyl. Alternatively, R5 is most preferably a methoxy, ethoxy, propoxy, or butoxy group; and

wherein R6 is a substituent selected independently from the group consisting of fluoro, chloro, bromo, iodo, trifluoromethyl, hydroxy, alkyl containing from 1 to 6 carbons, alkoxy containing from 1 to 6 carbons, alkoxycarbonyl containing from 2 to 6 carbons, carboxy, alkylcarbonylamino wherein the alkyl group contains 1 to 6 carbons, carboxyalkylaminocarbonyl wherein the alkyl group contains from 1 to 6 and preferably 1 carbon(s), 5-tetrazolo, and acylsulfonamido (i.e., acylaminosulfonyl and sulfonylaminocarbonyl) containing from 1 to 15 carbons, and provided that when the acylsulfonamido contains an aryl the aryl may be further substituted by a member selected from fluoro, chloro, bromo, iodo and nitro.

The preferred compounds of the present invention are of the S configuration at chiral centers marked by an asterisk in formulas (Ia) and (Ib). As will be appreciated by those skilled in the art, the activity of the individual isomers is not

the same and, it is therefore preferred to utilize the more active isomer. The present invention includes both the diastereomeric mixture and the active S and R isomers.

The most preferred HNE inhibitors suitable for the uses and in the methods of the present invention include the following compounds:

N-[2-[4-(2,2-dimethylpropioiiyloxy)phenylsulfonyl-amino]b enzoyl] aminoacetic acid (also known as ONO-5046);

N-(methoxycarbonyl)-L-valyI-L-prolyl-L-valyl-trifIuoromet hane monohydrate (also referred to as ZD-8321); N-(methyl-hydroxy-4-benzyl-l-carboxyl)-L-valyl-L-prolyl-L-va lyl- trifluoromethane monohydrate (also referred to as ZD-0892); or

3(RS)-[[4-(carboxymethylaminocarbonyl)phenylcarbonyl]-L-v alyl-L- prolyl]amino-l,l,l-trifluoro-4-methyl-2-oxopentane (also referred to as FK-706),

wherein ZD-8321, ZD-0892, and FK-706 are specific examples of formula (Ia) or (Ib).

It will be appreciated that the present invention also includes pharmaceutically acceptable salts, analogs or derivatives of any of the foregoing compounds. According to the present invention, each of the aforementioned preferred HNE inhibitors may be used alone or in combination with one or more other HNE inhibitors mentioned above in the preparation of a pharmaceutical composition for treating IBD or in the IBD treatment methods according to the present invention.

In preferred embodiments of the present invention, the disease to be treated and generally referred to as Inflammatory Bowel Disease (IBD), is selected from Ulcerative Colitis or Crohn's Disease. Inflammatory Bowel Disease, including

Ulcerative Colitis and Crohn's Disease, may either be classified as idiopathic (i.e., of unknown cause) or as non-idiopathic. Non-idiopathic IBD may inter alia be caused

by a variety of infections, including bacterial infection, viral infection, chlamydial infection, amebian infection, by antibiotics, or by ischemia. Accordingly, the compounds of the present invention are suitable for treating all of the above-listed disease variants.

In another aspect of the invention, the HNE inhibitor or combinations of different HNE inhibitors according to the present invention may be used for the preparation of pharmaceutical compositions, wherein the pharmaceutical compositions further comprise one or more other pharmaceutically active compound(s) with known effectiveness in the treatment of Inflammatory Bowel Disease, including Ulcerative Colitis and Crohn's Disease. The inventors have found that compositions comprising at least one HNE inhibitor and at least one other known compound commonly employed for treating IBD may even increase the effectiveness of treating the medical conditions according to the present invention. Accordingly, such pharmaceutical compositions are particularly preferred in all aspects of the present invention.

More preferably, the known pharmaceutically active compounds effective for IBD treatment that may be present in the pharmaceutical compositions according to the present invention are selected from the group of a corticosteroid, an 5-ASA agent, budesonid, a TNF-alpha antagonist (e.g., adalimumab (Humira®), infliximab (Remicade®), or etanercept (Enbrel®)), an integrin4 antagonist (e. g., SB- 683698, or natalizumab (Tysabri®)), an integrin4 receptor antibody (e. g. , MLN-02), and an interleukin antibody (e. g., MRA directed against IL-6, ABX-IL8 directed against IL-8, or J695 directed against IL-12).

Particularly preferred corticosteroids to be used in the pharmaceutical compositions of the invention include, but are not limited to dexamethasone, betamethasone, triamcinolone, triamcinolone acetonide, fluocinonide, fluocinolone acetonide, prednisolone, methylprednisolone, cortisone acetate, hydrocortisone, and beclometasone propionate, or pharmaceutically acceptable salts, analogs or derivatives thereof. Particularly preferred 5-ASA agents include, but are not limited

to sulfasalazine, olsalazine, balsalazine, mesalamine (such as the compounds known under the tradenames Asacol ^® and Pentasa , respectively), and mesalazine (e.g., Claversal ^® ).

Pharmaceutical compositions comprising the following combinations of an HNE inhibitor with a further pharmaceutically active compound effective in treating IBD are most particularly preferred for the uses and methods according to the present invention:

1.) N-[2-[4-(2,2-dimethylpropionyloxy) phenylsulfonyl-amino] benzoyl] aminoacetic acid (i.e., ONO-5046) and prednisolone;

2.) N-[2-[4-(2,2-dimethylpropionyloxy) phenylsulfonyl-amino] benzoyl] aminoacetic acid (i.e., ONO-5046) and sulfasalazine;

3.) N-[2-[4-(2,2-dimethylpropionyloxy) phenylsulfonyl-amino] benzoyl] aminoacetic acid (i.e., ONO-5046) and budesonid;

4.) N-[2-[4-(2,2-dimethylpropionyloxy) phenylsulfonyl-amino] benzoyl] aminoacetic acid (i.e., ONO-5046) and a TNF-alpha antagonist;

5.) N-[2-[4-(2,2-dimethylpropionyloxy) phenylsulfonyl-amino] benzoyl] aminoacetic acid (i.e., ONO-5046) and an integrin4 antagonist;

6.) N-[2-[4-(2,2-dimethyIpropionyloxy) phenylsulfonyl-amino] benzoyl] aminoacetic acid (i.e., ONO-5046) and an integrin4 receptor antibody;

7.) N-[2-[4-(2,2-dimethylpropionyloxy) phenylsulfonyl-amino] benzoyl] aminoacetic acid (i.e., ONO-5046) and an interleukin antibody;

8.) A trifluoromethyl ketone substituted peptide derivative of formula

(Ia) or (Ib) and prednisolone;

9.) A trifluoromethyl ketone substituted peptide derivative of formula

(Ia) or (Ib) and sulfasalazine;

10.) A trifluoromethyl ketone substituted peptide derivative of formula

(Ia) or (Ib) and budesonid;

11.) A trifluoromethyl ketone substituted peptide derivative of formula

(Ia) or (Ib) and a TNF-alpha antagonist;

12.) A trifluoromethyl ketone substituted peptide derivative of formula (Ia) or (Ib) and an integrin4 antagonist;

13.) A trifluoromethyl ketone substituted peptide derivative of formula

(Ia) or (Ib) and an integrin4 receptor antibody;

14.) A trifluoromethyl ketone substituted peptide derivative of formula

(Ia) or (Ib) and an interleukin antibody;

15.) N-(methoxycarbonyl)-L-valyl-L-prolyl-L-valyl-trifluoromethan e

(ZD-8321) and prednisolone;

16.) N-(methoxycarbonyl)-L-valyl-L-prolyl-L-valyl-trifluoromethan e

(ZD-8321) and sulfasalazine;

17.) N-(methoxycarbonyl)-L-valyl-L-prolyl-L-valyl-trifluoromethan e (ZD-8321) and budesonid;

18.) N-(methoxycarbonyl)-L-valyl-L-prolyl-L-valyl-trifluoromethan e

(ZD-8321) and a TNF-alpha antagonist;

19.) N-(methoxycarbonyl)-L-valyl-L-prolyl-L-valyl-trifluoromethan e

(ZD-8321) and an integrin4 antagonist;

20.) N-Cmethoxycarbony^-L-valyl-L-prolyl-L-valyl-trifluoromethane

(ZD-8321) and an integrin4 receptor antibody;

21.) N-(methoxycarbonyl)-L-valyl-L-prolyl-L-valyl-trifluoromethan e

(ZD-8321) and an interleukin antibody;

22.) N-(methyl-hydroxy-4-benzyl-l-carboxyl)-L-valyl-L-prolyl-L- valyl-trifluorometliane monohydrate (ZD-0892) and prednisolone;

23.) N-(methyl-hydroxy-4-benzyl- 1 -carboxyl)-L-valyl-L-prolyl-L- valyl-trifluoromethane monohydrate (ZD-0892) and sulfasalazine;

24.) N-(methyl-hydroxy-4-benzyl- 1 -carboxyl)-L-valyl-L-prolyl-L- valyl-trifluoromethane monohydrate (ZD-0892) and budesonid;

25.) N-(methyl-hydroxy-4-benzyl- 1 -carboxyO-L-valyl-L-prolyl-L- valyl-trifluoromethane monohydrate (ZD-0892) and a TNF-alpha antagoiiist;

26.) N-(methyl-hydroxy-4-benzyl- 1 -carboxyty-L-valyl-L-prolyl-L- valyl-trifluoromethane monohydrate (ZD-0892) and an integrin4 antagonist;

27.) N-(methyl-hydroxy-4-benzyl- 1 -carboxyty-L-valyl-L-prolyl-L- valyl-trifluoromethane monohydrate (ZD-0892) and an integrin4 receptor antibody;

28.) N-(methyl-hydroxy-4-benzyl- 1 -carboxyl)-L-valyl-L-prolyl-L~ valyl-trifluoromethane monohydrate (ZD-0892) and an interleukin antibody;

29.) 3(RS)-[[4-(carboxymethylaminocarbonyl) phenylcarbonyl]-L- valyl-L-prolyl]amino- 1,1,1 -trifluoro-4-methyl-2-oxopentane (FK- 706) and prednisolone;

30.) 3(RS)-[[4-(carboxymethylaminocarbonyl) phenylcarbony I]-L- valyl-L-prolyI]amino- 1,1, l-trifIuoro-4-methyl-2-oxopentane (FK- 706) and sulfasalazine;

31.) 3(RS)-[[4-(carboxymethylaminocarbonyl) phenylcarbony I]-L- valyl-L-prolyl]amino-l,l,l-trifluoro-4-methyl-2-oxopentane (FK-

706) and budesonid;

32.) 3(RS)-[[4-(carboxymethylaminocarbonyl) phenylcarbonyl]-L- valyl-L-prolyl]amino- 1,1,1 -trifluoro-4-methyl-2-oxopentane (FK- 706) and a TNF-alpha antagonist;

33.) 3(RS)-[[4-(carboxymethylaminocarbonyl) phenylcarbonyl]-L- valyl-L-prolyl]amino- 1,1,1 -trifluoro-4-methyl-2-oxopentane (FK- 706) and an integrin4 antagonist;

34.) 3(RS)-[[4-(carboxymethylaminocarbonyl) phenylcarbonyl]-L- valyI-L-prolyI]amino- 1 , 1 , l-trifluoro-4-methyl-2-oxopentane (FK- 706) and an integrin4 receptor antibody;

35.) 3(RS)-[[4-(carboxymethylaminocarbonyl) phenylcarbonyl]-L- valyl-L-prolyl]amino- 1 , 1 , l-trifluoro-4-methyl-2-oxopentane (FK- 706) and an interleukin antibody.

In another aspect, the pharmaceutical compositions as defined herein are also provided by the present invention. These pharmaceutical compositions include all the specific embodiments as defined hereinabove, including compositions comprising one HNE inhibitor, compositions comprising a combination of at least two different HNE inhibitors, particularly selected from the group of ONO-5046, ZD-

8321, ZD-0892, and FK-706, or compositions comprising at least one HNE inhibitor and at least one other pharmaceutically active compound known to be effective in treating IBD. Particularly preferred compositions of the present invention include the specific combinations as listed above (see combinations numbered 1 to 35).

Optionally, the pharmaceutical compositions of the present invention may further comprise pharmaceutically acceptable carriers, diluents and/or excipients, and the like.

In any event, it will be understood that any of these pharmaceutical compositions as defined above may be administered to a human patient in the IBD treatment methods according to the present invention.

Pharmaceutical Formulations and Modes of Administration

The compounds of the present invention, in particular the human neutrophil elastase (HNE) inhibitors according to formula (Ia) or (Ib), such as ZD- 8321, ZD-0892 and FK-706, or ONO-5046, optionally in combination with other pharmaceutically active compounds known to be effective in treating IBD, may be administered to a subject, e.g., a mammal, such as a human patient, in need thereof, in a variety of forms adapted to the chosen route of administration, e.g., orally, rectally or parenterally, by intravenous, intramuscular, topical, transdermal or subcutaneous routes.

The compounds of the present invention may be administered systemically, e.g., orally, in combination with a pharmaceutically acceptable vehicle such as an inert diluent or an assimilable edible carrier. They may, e.g., be enclosed in hard or soft shell gelatin capsules, may be compressed into tablets, or may be incorporated directly with the food of the patient's diet. For oral therapeutic administration, the compounds of the invention may also be combined with one or more excipients and used, e.g., in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like. Such compositions and preparations should contain at least 0.1% of the compounds of the present invention. The percentage of the compounds of the present invention in the compositions and preparations may, of course, be varied and may conveniently be between about 2% to about 80% of the weight of a given unit dosage form. The amount of compounds of the present invention in such therapeutically useful

compositions is such that an effective dosage level will be obtained and may easily be determined by those of ordinary skill in the art.

The tablets, troches, pills, capsules, and the like may also comprise the following: binders such as gum tragacanth, acacia, corn starch or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid and the like; a lubricant such as magnesium stearate; and/or a sweetening agent such as sucrose, fructose, lactose or aspartame or a flavoring agent such as peppermint, oil of wintergreen, or cherry flavoring. When the unit dosage form is a capsule, it may contain, in addition to materials of the above type, e.g., a liquid carrier, such as a vegetable oil or a polyethylene glycol. Various other materials may be present as coatings or to otherwise modify the physical form of the solid unit dosage form. For instance, tablets, pills, or capsules may be coated with, e.g., gelatin, wax, shellac or sugar and the like. A syrup or elixir may comprise the compounds of the invention, sucrose or fructose as a sweetening agent, methyl and propylparabens as preservatives, a dye and flavoring such as cherry or orange flavor. Of course, any material used in preparing any unit dosage form should be pharmaceutically acceptable and substantially non-toxic in the amounts employed. In addition, the compounds of the present invention may be incorporated into sustained-release preparations and devices.

In view of the nature of the disease to be treated, it will be appreciated that the preferred route of administration for the compounds of the present invention must fulfill two criteria. First, delivery of the compound or compounds should be convenient for the patient to be treated yet at the same time offer a highly selective, localized release of the active compounds at the desired location in the body. For the treatment of IBD, the compounds of the present invention are thus preferably administered orally, and are released specifically into the gastrointestinal tract of the patient suffering from said medical condition. Preferably, the drug is released into the intestine and/or colon of the human subject to be treated.

Although oral delivery has become a widely accepted route of administration of therapeutic drugs, the gastrointestinal tract presents some arduous barriers to drug delivery. In order to achieve successful delivery to, e.g., the colon, the active compounds of the present invention need to be protected from absorption and /or the environment of the upper gastrointestinal tract and then be released into the proximal colon, which is considered the optimum site for colon-targeted delivery of drugs. The various strategies for targeting orally administered compounds to the colon include exploitation of carriers that are degraded specifically by colonic bacteria, formulation of timed release systems, coating with pH-sensitive polymers, bioadhesive systems, osmotic controlled drug delivery systems and covalent linkage of a drug with a carrier. Microbially degradable polymers, especially azo-crosslinked polymers have also been investigated for use in targeting of drugs to colon. Certain plant polysaccharides such as amylose, inulin, pectin and guar gum are not attacked by gastrointestinal enzymes, thereby offering ways for the formulation of colon targeted delivery systems for the compounds of the present invention. Moreover, the concept of using pH as a trigger to release a drug in the colon is based on the pH conditions that vary continuously within different regions of the gastrointestinal tract. Time-dependent drug delivery systems are based on the principle to prevent release of drug until 3-4 h after leaving the stomach. Additional drug delivery systems suitable for colon delivery include redox sensitive polymers.

The formation of prodrugs of the compounds of the invention involves covalent linkage between the compound and the carrier. Preferred prodrugs of the compounds of the present invention include azo bond prodrugs in view of the presence of azo reductase enzymes in the colon and glycosidic prodrugs which are cleaved by glycosidase activity of the colonic microflora. Release of drugs from azo polymer coated dosage forms is supposed to take place after reduction and thus cleavage of the azo bonds by the azoreductase enzymes present in the colonic microflora. Natural polysaccharides may also be used as tools to deliver the compounds specifically to the colon. These polysaccharides remain intact in the physiological environment of the stomach and small intestine but once the dosage form enters into colon, it is acted upon by polysaccharidases that degrade the

polysaccharide and release the compound into the colon. The polysaccharides should be protected while entering the stomach and small intestine to prevent swelling of such dosage forms. Protection may be achieved either by chemical cross-linking or by addition of a protective coat.

In a preferred embodiment of this aspect of the present invention, the compounds or compositions of the invention may be formulated into a polymer delivery system and administered orally. The linkage between the polymer and the drug can alter the properties of the therapeutic so that it has a higher affinity for specific sites, such as cells or tissues. This helps to reduce potential toxic side effects occurring during the course of a treatment and protects the drug from the various attacks from enzymes and other molecules of the human body. The delivery system is designed such that the polymer begins to break down via diffusion or surface erosion once it enters the target side and slowly releases the drug. Such a delivery system is described for example in EP 0 759 303 and US 6,368,629. In these patents, a colon- specific drug release system, which comprises a drug coated with an pH-sensitive inorganic acid-soluble polymer material (first layer next to the drug), a saccharide which rapidly generates an organic acid by the action of enterobacteria in the lower part of the gastrointestinal tract, and another enteric coating polymer material (outer layer) is described (see also in Chu, 2003).

Alternatively, the compounds and compositions of the present invention may be delivered by a so-called oral controlled-release (OCR) delivery system. OCR drugs are less susceptible to gastric degradation and generally exhibit better bioavailability and bioabsorption than their immediate-release counterparts.

As already indicated hereinabove, another aspect of the present invention relates to a method of preventing, treating, or ameliorating Inflammatory

Bowel Disease as defined above in a human subject, said method comprising administering to said human subject a pharmaceutical composition as described and claimed herein. Also encompassed by the present invention is the use of the

compounds described and claimed herein in said methods and for the preparation of a pharmaceutical composition intended for preventing, treating, or ameliorating IBD.

Preferred target cells for the above-described methods or uses are cells of the gastrointestinal tract of the human subject to be treated, which particularly includes cells of the small intestine, large intestine, and colon, and wherein such cells are located within loci of inflammation.

It will be appreciated that all pharmaceutical compositions and formulations described herein may principally be used in the therapeutic methods described and claimed herein. Particularly preferred formulations for delivery of the compounds of the present invention to the target cells of a human subject to be treated include the oral controlled-release delivery formulations as described hereinabove.

The compounds of the invention may also be administered intravenously or intraperitoneally by infusion or injection. Solutions of the compounds of the invention or its salts may be prepared in water, optionally mixed with a nontoxic surfactant. Dispersions may also be prepared, e.g., in glycerol, liquid polyethylene glycols, triacetin, and mixtures thereof and in oils. Under ordinary conditions of storage and use, these preparations may comprise a preservative to prevent the growth of microorganisms.

The pharmaceutical dosage forms suitable for injection or infusion may include sterile aqueous solutions or dispersions or sterile (e.g. lyophilized) powders comprising the compounds of the invention, which are adapted for the extemporaneous preparation of sterile injectable or infusible solutions or dispersions, optionally encapsulated in liposomes.

In all cases, the ultimate dosage form must be sterile, fluid and stable under the conditions of manufacture, preparation, and storage. The liquid carrier or vehicle may be a solvent or liquid dispersion medium comprising, for example, water, ethanol, a polyol (for example, glycerol, propylene glycol, liquid polyethylene glycols, and the like), vegetable oils, nontoxic glycerol esters, and suitable mixtures

thereof. The proper fluidity may be maintained, for example, by the formation of liposomes, by the maintenance of the required particle size in the case of dispersions or by the use of surfactants. The prevention of the action of microorganisms may be brought about by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars, buffers or sodium chloride. Prolonged absorption of injectable compositions comprising the compounds of the present invention may be brought about by the use of agents delaying absorption, for example, aluminum monostearate and gelatin.

Sterile injectable solutions are prepared by incorporating the compounds of the present invention in the required amount in the appropriate solvent with various of the other ingredients enumerated above, as required, followed by filter sterilization. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum drying and the freeze drying techniques, which yield a powder of the compounds of the invention plus any additional desired ingredient present in the previously sterile-filtered solutions.

For topical administration, in case the compounds of the present invention are liquids, they may be applied in pure form. However, it will generally be desirable to administer them to the skin as compositions or formulations, in combination with a dermatologically acceptable carrier, which may be a solid or a liquid.

Useful solid carriers include finely divided solids such as talc, clay, microcrystalline cellulose, silica, alumina and the like. Useful liquid carriers include water, hydroxyalkyls or glycols or water-alcohol/glycol blends, in which the compounds of the invention can be dissolved or dispersed at effective levels, optionally with the aid of non-toxic surfactants. Adjuvants such as fragrances and additional antimicrobial agents can be added to optimize the properties for a given use. The resultant liquid compositions may be applied from absorbent pads, or may

be used to impregnate bandages and other dressings, or may be sprayed onto the affected area using pump-type or aerosol sprayers.

Thickeners such as synthetic polymers, fatty acids, fatty acid salts and esters, fatty alcohols, modified celluloses or modified mineral materials may also be employed with liquid carriers to fonn spreadable pastes, gels, ointments, soaps, and the like, for application directly to the skin of the user.

Examples of useful dermatological compositions which may be used to deliver the compounds of the present invention to the skin are known in the art; for example, see Jacquet et al. (US 4,608,392), Geria (US 4,992,478), Smith et al. (US 4,559,157) and Wortzman (US 4,820,508).

Useful dosages of the compounds of the present invention may be determined by their in vitro activity, as well as in vivo activity in animal models. Methods for the extrapolation of effective dosages in mice and other animals to humans are known to the art; see for example, US 4,938,949.

Generally, the concentration of the compounds of the present invention in a liquid composition will be from about 0.1-25 % per weight, preferably from about 0.5-10 % per weight. The concentration in a semi-solid or solid composition such as a gel or a powder will be about 0.1-5 % per weight, preferably about 0.5-2.5 % per weight.

The dosage regimen to be employed in connection with the pharmaceutical compositions, methods and uses of the invention is selected in accordance with a variety of factors including type, species, age, weight, sex and medical condition of the patient; the severity of the condition to be treated; the route of administration; the target cell or tissue, the renal and hepatic function of the patient; and the particular compound (or its salt, analog, or derivative thereof) employed. An ordinarily skilled physician or veterinarian can readily determine and prescribe the effective amount of the compound or the compounds required to prevent, ameliorate, or treat the condition.

In general, however, a suitable dose will be in the range of from about

0.1 to about 100 mg/kg body weight, preferably from about 1 to about 80 mg/kg of body weight per day [mg/kg/day], and more preferably from about 3 to about 50 mg/kg/day. Particularly preferred are doses in the range of about 3 to about 30 mg/kg/day, most preferably in the range of 5 to about 25 mg/kg/day.

The compounds or compositions of the present invention may conveniently be administered in unit dosage form; for example, containing 5 to 1000 mg, conveniently 10 to 750 mg, most conveniently, 50 to 500 mg of active ingredient per unit dosage form.

Ideally, the compounds of the present invention should be administered to achieve peak plasma concentrations of the active compound of from about 0.0005 to about 300 μM, preferably, about 0.001 to 100 μM, more preferably, about 1 to about 100 μM.

The desired dose may conveniently be presented in a single dose or as divided doses administered at appropriate intervals, for example, as two, three, four or more sub-doses per day.

An administration regimen according to the present invention includes long-term, daily treatment. By "long-term" is meant at least two weeks and preferably, several weeks, months, or years of duration. Since IBD is usually chronic in nature, it may even be necessary that the treatment be continued for the entire life of the patient, or at least be resumed in acute phases that follow phases of remission of the disease. Necessary modifications in this dosage range may be determined by one of ordinary skill in the art according to the teachings herein or the teachings described in Remington's Pharmaceutical Sciences (Martin, E. W., ed. 4), Mack Publishing Co., Easton, PA. The dosage may also be adjusted by the individual physician in the event of any complication.

The invention also provides a pharmaceutical pack or kit comprising one or more containers filled with one or more of the ingredients of the

pharmaceutical compositions of the invention as described above. Associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration, and may further include instructions for use.

The present invention also provides methods for the treatment of IBD, wherein the HNE inhibitors of the invention and the other pharmaceutically active compound(s), as listed above, with which it/they are combined in the pharmaceutical compositions according to the present invention may be either administered jointly (i.e., at the same time), or separately (i.e., sequentially), preferably according to a dosing regimen.

Thus, in one embodiment of this aspect of the present invention, dosage delivery of a HNE inhibitor of the present invention can begin 48 hours prior to delivery of the other compound, as listed above, with the preferred time being 24 hours and the most preferred time being 12, 9, 6, 5, 4, 3, 2, 1, or less than one hour prior to the delivery of the other compound. Alternatively, dosage delivery of a neutrophil elastase inhibitor of the present invention can begin 48 hours after the initiation of delivery of the other compound, with the preferred time being 24 hours and the most preferred time being 12, 9, 6, 5, 4, 3, 2, 1, or less than one hour after the delivery of the other compound .

Alternatively, the compounds may be administered concurrently (i.e., at the same time). It will be appreciated that concurrent administration may either be accomplished by formulating the at least two compounds into a single pharmaceutical formulation, or by administering multiple formulations (e.g., tablets, etc.) each containing one of the active ingredients, together in a single step.

The neutrophil elastase inhibitor(s) and/or the other pharmaceutically active compound can be independently administered by a variety of routes, including oral or rectal routes, or parenteral routes like intravenous, intramuscular, topical,

transdermal or subcutaneous routes as described in detail above, in the therapeutic methods of the present invention.

Examples

Example 1

To analyze the therapeutic effect of ONO-5046 in the colon of dextran sulfate sodium (DSS) induced colitis mice, 35 C3HeB/FeJ mice have been treated with 3,5% (wt/vol) of DSS in drinking water for 5 days, followed by supply of normal water. 5 mice of the same strain were used as "untreated control". DSS treatment led to colitis-like symptoms such as diarrhea and blood in the faeces. Over a period of 14 days after DSS treatment, each group (N=IO) of the DSS-treated mice received an intraperitoneal application of either 10 mg/kg bodyweight/day of the HNE inhibitor ONO-5046 (ONO-5046) twice a day, or of 10 mg/kg bodyweight/day of the corticosteroid prednisolone (prednisolone; a drug used for treatment of IBD in human), or of isotonic NaCl solution (NaCl control), respectively. 5 mice without DSS treatment and without compound treatment served as additional control (untreated control). After day 14 of compound administration, all mice were sacrificed and histopathological analysis of each colon was performed. Analysis comprised examination and grade scoring of colon submucosa inflammatory infiltration, of regeneration of former ulceration, and of proliferation of lymphatic aggregates. Scoring was performed based on different grade levels from 0 to 5, according to the grade list provided in the Annex to Example 1. The higher the number, the more severe are the symptoms observed. The scoring numbers from all mice belonging to one group of treatment were summed. As shown in Figure 1, ONO-5046 significantly reduced the degree of colon submucosa inflammation (scoring number: 13), similar to prednisolone (scoring number: 14) and in contrast to NaCl control mice (scoring number: 23) without compound treatment. DSS-untreated mice are inconspicuous (scoring number: 2). Regeneration of former ulceration by re- epithelization was strongly supported by ONO-5046 (scoring number: 3), whereas no

such effect was observed in prednisolone treated mice (scoring number: 12) and NaCl control mice (scoring number: 14). Proliferation of lymphatic aggregates in the colon mucosa was not affected by ONO-5046 (scoring number: 18) when compared to NaCl control mice (scoring number: 20). In contrast, prednisolone significantly reduced the degree of proliferation of lymphatic aggregates (scoring number: 10). Based on the above experimental results it may be concluded that a therapeutic treatment of IBD with ONO-5046 and prednisolone in combination seems favorable. In sum, the overall therapeutic effect in the treatment of colitis is very similar for both compounds, according to the total scoring numbers observed for ONO-5046 (sum scoring number: 34) and prednisolone (sum scoring number: 36).

Annex to Example 1.

A. Scoring of inflammatory infiltration in colon submucosa

Grade 0: infiltration of inflammatory cells in the submucosa is comparable to unchallenged and untreated control animals.

Grade 1 (minimal): single focal or diffuse areas in the submucosa (less than 5% of the section) show increased infiltration of inflammatory cells.

Grade 2 (low): few focal and diffuse areas in the submucosa (up to 25% of the section) show increased infiltration of inflammatory cells.

Grade 3 (intermediate): multifocal and diffuse areas in the submucosa (up to 50% of the section) with infiltration of inflammatory cells, edema and hyperemia.

Grade 4 (severe): multifocal and diffuse areas in the submucosa (up to 75% of the section) with infiltration of inflammatory cells, edema and hyperemia.

Grade 5 (ultimate, lethal): severe multifocal and diffuse areas in the submucosa (more than 75% of the section) with infiltration of inflammatory cells, edema and hyperemia.

B. Scoring of Regeneration

Grade 0: no focus of regeneration obvious.

Grade 1 (minimal): one or two small foci of former ulceration with residuals of granulocyte infiltration, demarcation and reepithelization.

Grade 2 (low): two large foci of former ulceration with residuals of granulocyte infiltration, demarcation and reepithelization.

Grade 3 (intermediate): three foci of former ulceration with residuals of granulocyte infiltration, demarcation and partly incomplete reepithelization.

Grade 4 (severe): three to five foci of former ulceration with residuals of granulocyte infiltration, demarcation and incomplete reepithelization.

Grade 5 (ultimate, lethal): more than five foci of former ulceration with residuals of granulocyte infiltration, demarcation and incomplete reepithelization.

C. Scoring of proliferation of lymphatic aggregates

Grade 0: one to two small aggregates of lymphocytes; not larger than mucosal thickness.

Grade 1 (minimal): two to three small enlarged aggregates of lymphocytes (minimal thicker than mucosa).

Grade 2 (low): two to three small enlarged aggregates of lymphocytes (all thicker than mucosa).

Grade 3 (intermediate): three to five prominent lymphatic aggregates with distinct bulge of the mucosa.

Grade 4 (severe): more than five prominent lymphatic aggregates with distinct bulge of the mucosa.

Grade 5 (ultimate, lethal): more than five prominent lymphatic aggregates with distinct bulge of the mucosa and signs of pathological changes (focal necrosis of the gut-associated lymphatic tissue (GALT) or the surrounding intestinal tissue).

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