Abstract:
Rosiglitazone, 5-ASA or structurally analogous Compounds according to the general formula (I): or Compounds according to the general formula (Ia): For the induction of CAMP expression in tissues having PPAR-gamma receptors. Such tissues include epithelia or mucosae tissue having PPAR-gamma receptors and of particular interest is CAMP expression in the gut.

Description:
FIELD OF THE INVENTION 
       [0001]    The present invention relates to the induction of the defensins by PPAR-gamma agonists. Defensins provide immunoprotection to the skin, oral, nasal, ocular epithelia and other epithelia mucosae, including the vaginal mucosae. In particular, the invention is concerned with stimulation of defensins production by such agonists through activation of the PPAR receptors in epithelia and/or mucosae. More particularly, the invention is concerned with stimulation of enteric defensins production by such agonists by activation of the gut PPAR receptors. 
       BACKGROUND TO THE INVENTION 
       [0002]    Inflammatory bowel diseases (IBD), including Crohn&#39;s disease (CD) and ulcerative colitis (UC), are chronic recurrent diseases with remissions and exacerbations, appearing principally in young patients. Inflammation may affect all regions of the bowel and all layers of the gut mucosa, including the adjacent mesenteric adipose tissue and perianal areas in CD. These diseases are clinically characterized by prolonged and variable courses, the diversity of intestinal manifestations, and by occurrence of serious local and systemic complications. The aetiology of CD and UC remains unknown, although the pathological intestinal inflammatory response is thought to be a consequence of a breakdown of tolerance to bacterial flora in the gastrointestinal tract of genetically predisposed individuals 1 . 
         [0003]    IBD are more prominent in developed countries and particularly in Western Europe, North America, and Australia. Prevalence of CD and UC is about 1-2/1000 inhabitants. Notably in 2005, a total of 120 000 CD and 80 000 UC is estimated in France and about 2.5 million IBD patients in the United States. Without any available curative therapeutic molecules, the therapeutic management associates symptomatic therapy (analgesic, antibiotics, nutrition), anti-inflammatory and immunosuppressive agents (aminosalicylates, steroids, azathioprine, methotrexate, cyclosporine, monoclonal anti-TNF antibodies such as infliximab) and surgical treatment. Taken together, the development of new therapeutic molecules is therefore critical for the clinical management. 
         [0004]    The digestive mucosa has evolved various immune strategies to tolerate intimate contact with commensals and prevent pathogenic bacteria from spreading into host tissues. The recognition of food-borne indigenous and pathogenic microbes is an essential barrier function for the survival of insects and mammals. In particular, mammalian resistance to pathogens is mainly conferred by membrane-bound Toll-like receptors (TLRs) 1  and the recently identified family of cytosolic nucleotide-binding oligomerization domain leucine-rich repeat containing proteins (NOD-LRRs) 2 . NOD1 and NOD2 possess an N-terminal caspase recruitment domain (CARD), a central nucleotide-binding domain (NOD) and a C-terminal leucine-rich repeat domain (LRR) 2 . 
         [0005]    Considerable attention has been focused on NOD2 signalling, as mutations of this gene have been associated with Crohn&#39;s disease (CD) 3,4 . NOD2 acts as a cytosolic pattern-recognition molecule (PRM) for bacterial peptidoglycan s  by detecting a major muropeptide released and recycled during bacterial growth—the muramyl dipeptide MurNAc-L-Ala-D-isoGln (MDP) 6-8 . Following recognition of MDP, NOD2 promotes and regulates both innate and adaptive immunity via transcriptional factors and kinase activation 2 . Hence, the absence of NOD2 signalling in mice understandably confers oral susceptibility to  Listeria monocytogenes  via the regulation of certain enteric cationic antimicrobial peptides (CAMPs) 8 . Recent work has provided evidence that NOD1 confers responsiveness to peptidoglycans containing meso-diaminopimelic acid (primarily found in Gram negative bacteria) 9,10 . Similarly to the physiological role of NOD2, NOD1 is required for expression of certain β-defensins by gastric epithelial cells during  Helicobacter pylori  infection 11 . 
       Gastrointestinal Antimicrobial Peptides (CAMPs: Defensins, Cathlelicidins): 
     Implications for Inflammatory Disease 
       [0006]    Recent reports have shed light on the effector role of CAMPs in monitoring gut homeostasis and containing invading microbes, since the stem cells that replenish gut epithelium require continuous antimicrobial protection. The level of CAMP expression parallels intestinal development in metazoans, from the immaturity of local defense mechanisms during gestation to bacterial colonization of the gut after birth 12 . Of particular interest are the NF-κB-dependent and independent regulation of two types of CAMP which are prevalent in the mammalian gut, namely the defensins and cathelicidins. 
         [0007]    The α- and β-defensins are small polypeptides with spatially separated hydrophobic and positively charged residues. Six invariant cysteines form 3 specific intramolecular disulfide bonds and thus stabilize the protein in a complexly folded, triple-stranded beta-sheet configuration 12,13  Whereas the α-defensins HD-1 to HD-4 (also known as human neutrophil proteins 1 to 4) are expressed by neutrophils, HD-5 and HD-6 (also known as cryptdins in mice) are produced by specialized intestinal epithelial cells, called Paneth cells. The latter are located primarily at the base of the crypts of Lieberkühn in the small intestine and have a major role in the innate immunity of the ileal mucosa by synthesizing and releasing proteinaceous granules into the lumen following exposure to microbes and/or microbial products. These secretory granules are rich in amphipathic peptides which can cause microbial death by disrupting membrane integrityl 12    
         [0008]    Paneth cells play a crucial role in maintaining the tolerance towards commensals and in repelling pathogenic infections by producing antimicrobial peptides, such as defensin. The Wnt/Tcf/beta-catenin signaling pathway is essential in controlling intestinal homeostasis and Paneth cells differentiation. 
         [0009]    Interestingly, impaired intestinal expression of enteric alpha-defensins (namely HD-5) has been reported in CD which might contribute to changes in the luminal flora and/or generate vulnerability throughout the epithelial barrier to infection with enteropathogens 4 , such as adherent-invasive  E. coli   59  and  M. paratuberculosis   60  ( FIG. 2 ). The influence of other microbial sensors and the CAMPs on the emergence of colonic disease also needs to be clarified, since the physiological bacterial load is higher in the colon than in the small intestine. Finally, the use of mice with mutant CAMPs and a recently developed mouse model carrying the major CD-associated NOD2 mutation 61  may help determine if impaired enteric defensin function and/or natural NODs mutations are sufficient to trigger the development of intestinal inflammatory diseases. The in vivo antimicrobial function of α-defensins has been experimentally exemplified by observation of greater resistance to  Salmonella typhimurium  infection in HD-5 transgenic mice, accompanied by marked changes in the composition of the dominant flora in the gastrointestinal lumen 14 . Unlike β-defensins, the α-defensins produced by Paneth cells are mainly regulated at a post-transcriptional level by extracellular proteases 15 , including matrix metalloproteinase-7 (MMP-7, matrilysin) and trypsin in mice and humans, respectivelyl 16,17 . Hence, MMP-7 −/−  mice accumulate inactive forms of cryptdins and succumb more readily to oral infection with  S. typhimurium  than do wild-type animals 17 . Six human β-defensins (hBD-1 to hBD-6) are primarily synthesized by most epithelial cells. Mice lacking the hBD1 orthologue show increased susceptibility to  Staphylococcus aureus  infection 18 , supporting a role for this protein in innate immunity. 
         [0010]    Cathelicidins are CAMPs which are structurally and evolutionary distinct from defensins but which have a similar abundance and distribution in the gastrointestinal tract 12 . They are synthesized as large precursor peptides containing a highly conserved N-terminal domain (cathelin), linked to a C-terminal peptide with antimicrobial activity. As with defensins, cathelicidins are activated by extracellular, partial proteolysis 19 . Although several members of the family have been identified in other mammalian species, humans and mice possess a single cathelicidin gene (referred as LL37/FALL39/hCAP18 and cathelin-related anti-microbial peptide (CRAMP), respectively) 20 . Experimental evidence has indicated that mice lacking CRAMP are more susceptible to cutaneous infection by group A streptococci and urinary tract infection by invasive  Escherichia coli   21,22 . Furthermore, CRAMP-deficient macrophages failed to control replication of  S. typhimurium   23 . 
         [0011]    Mice bearing mutations in the NF-κB and MyD88 signalling pathways display increased susceptibility to  Helicobacter -induced colitis 24  and commensal-triggered colitis respectively 25 , indicating an essential role for NF-κB in gut tolerance/resistance to bacteria and a potential involvement in the regulation of CAMP production. In humans, hBD-1 expression is constitutive in the small intestine and the colon, whereas colonic synthesis of hBD-2 to -4 is strongly dependent on NF-κB activation by infectious agents in the digestive tract (such as  H. pylori ) and/or pro-inflammatory cytokines 12 . In addition to the regulatory impact of TLR signalling 26 , the NOD1 and NOD2 signalling pathways have been shown to trigger hBD-2 expression 11,27 . Furthermore, recent findings have shown that activation of the MAP kinase pathways is also required for hBD-2 and/or -3 expression 11,26 . 
         [0012]    It has been shown that three mutations in the NOD2 gene (namely R702W, G908R and 1007fs) lead to a predisposition to CD 3,4 . Genotype-phenotype correlations have established that NOD2 mutants are predominantly linked to ileal CD 42 . Both common and rare mutations have been associated with impaired MDP-induced NF-κB activation 5,7  and cytokine production in peripheral blood monocytes 7,43-45 . Lala and collaborators recently reported that NOD2 is highly expressed in Paneth cells 46,47 , a finding which might account for the association between NOD2 mutations and the development of ileal inflammatory lesions 42 . In agreement with a protective effect of NOD2 in the ileum, Nod2-knockout mice displayed (i) enhanced susceptibility to oral infection (but not systemic infection) with the Gram positive facultative intracellular bacterium  L. monocytogenes  and (ii) markedly decreased expression of a subgroup of cryptdin genes 8 . 
         [0013]    Importantly, decreased production of HD-5 and HD-6 has been found in surgical resection specimens and biopsies from ileal CD patients 14,48,49 ; reportedly, the CD-associated NOD2 mutations contributed to this impairment. On the other hand, individuals with Crohn&#39;s colitis displayed normal α-defensin levels but have a much reduced copy number for the β-defensin gene hBD-2, resulting in impaired expression in the colon 50 . As with NOD2 mutations, complex intronic polymorphism of the NOD1 gene has been associated with the pathogenesis of IBD 51 . In addition, Nod1-deficient mice display increased susceptibility to  H. pylori  infection 52  and decreased expression of certain β-defensins 11 . 
         [0014]    Reduced expression of defensins in ileal CD might contribute to changes in the luminal flora, thus generating vulnerability throughout the epithelial barrier to infection with CD-associated pathogens such as adherent-invasive  E. coli   59  and  M. paratuberculosis   60    
         [0015]    In parallel, NF-κB-independent signalling pathways might control CAMP production by regulating epithelium cell renewal, differentiation and/or lineage commitment. Interestingly, impaired Wingless (Wnt) signalling is associated with a complete lack of proliferative cells in the foetal small intestinal epithelium 28 , suggesting that this pathway has an essential role in maintaining the proliferative/undifferentiated status of intestinal epithelial cells. The absence of the ephrinB3 gene (which is downregulated by the Wnt signalling pathway) was seen to result in abnormal Paneth cell lineage commitment 29 . Moreover, the Wnt signalling pathway might monitor defensin gene expression (via the transcription factor 4, Tcf4) in cells derived from Paneth cells, since cryptdins were not detected in the small intestine of embryonic Tcf4−/− mice or that of adults lacking the Wnt receptor Frizzled-5 30 . Conversely, cryptdin genes are overexpressed in mice showing mutational activation of the Wnt signalling pathway 30,31 . 
         [0016]    A site-directed mutational analysis of α-defensin promoters revealed an essential, regulatory role for TCF binding sites 30 . Taken as a whole, these findings indicate that activation of the Wnt signalling is required for the production of Paneth cell-derived CAMPs. Hence, Paneth cell determinants (such as Mtgr1 and Gfi1) should be considered as potential candidates for susceptibility to chronic inflammatory disorders 32,33 . 
         [0017]    Finally, and given the crucial role of certain nuclear receptors in immunity, bacterial-induced inflammation and cell proliferation/maturation, it has been suggested that these receptors may have a potential role in gastrointestinal innate immunity by regulating CAMP biogenesis. Interestingly, a glucocorticoid receptor agonist (dexamethasone) has been shown to enhance hBD-2 expression, although the mechanism remains to be determined 34 . More recently, cathelicidin- and hBD2-encoding genes were identified as targets of the vitamin D receptor (VDR) 35 , a nuclear receptor required for resistance to  M. bovis  infection 36 . Treatment of monocytes with a synthetic VDR ligand led to dose-dependent up-regulation of cathelicidin gene transcription, which exerts a direct antimicrobial effect on  M. tuberculosis   37 . In agreement with these findings, individuals with decreased endogenous VDR ligand levels display increased susceptibility to  M. tuberculosis  infection 37 . 
         [0018]    To circumvent the CAMPs&#39; microbicidal activity, microorganisms (generally pathogens) have developed a range of strategies which are reminiscent of those involved in antibiotic resistance 39 . One way to achieve inactivation is to produce proteases, which degrade CAMPs; however, in the case of defensins, the intramolecular disulphide bridges render the peptides relatively resistant to enzymatic proteolysis. Another stratagem consists in reducing the net cationic charge of the bacterial envelope, in order to lower its affinity for CAMPs; this is achieved by incorporating positively-charged groups into the teichoic acid polymers (D-alanine) and in the lipid A (aminoarabinose) in the bacterial cell wall. Other bacterial approaches to CAMP resistance include preventing the host effectors from accessing their target via extracellular capture by secretory proteins and actively pumping the peptides across the cytoplasmic membrane 39 . However, despite these various protective weapons (which are not mutually exclusive), microorganisms will probably still be inhibited by CAMPs if the host is capable of releasing the latter in high amounts into the intestinal lumen, as in the case of defensins. In such a situation, down-regulation of CAMP-encoding genes at the transcriptional level by bacterial components (as reported in patients with shigellosis 40  and in mice orally infected with  S. typhimurium   41 ) may be a very sophisticated counter-mechanism ( FIG. 2 ). 
         [0019]    Impaired microbial sensing and aggression by specific enteric microbes may affect CAMPs function in the gut and result in the development of chronic inflammatory disorders, such as inflammatory bowel disease (IBD). These findings shed new light on the pathogenesis of CD, which is classically viewed as resulting from an abnormal T-cell activation by microbial immungens. 
         [0020]    In addition to the antimicrobial activity of CAMPs, pleitotropic functions have been attributed to defensins and cathelicidins (Table 1) 53 . Both CAMPs have the ability to chemoattract immunocytes involved in innate immunity (neutrophils and monocytes/macrophages), adaptive immunity (dendritic cells and T lymphocytes) and allergic/inflammatory reactions (mast cells). Furthermore, hBD2 might activate the TLR4-dependent signalling pathway in dendritic cells 54 . 
         [0021]    On the other hand, Hancock et al. recently reported that LL-37 may dampen TLR-dependent activation in human monocytes 55  and may promote maturation of dendritic cells, resulting in Th1 polarization of T cells 56 . Taken as a whole, these findings indicate that CAMP interactions can initiate and control the inflammatory response by linking innate and acquired immunity (Table 1). Finally, CAMPs such as LL-37 have the ability to promote angiogenesis, as demonstrated by decreased vascularization during skin wound repair in CRAMP-deficient mice 57 . Since Paneth cell biology influences intestinal angiogenesis through the recognition of commensals 58 , these findings provide insight into the pathogenesis of IBD. Enteric CAMPs have a number of essential and emerging roles in both innate and adaptive immunity of the gastrointestinal tract by modulating microbial resistance, angiogenesis, chemotaxis and the activation/maturation of the humoral response (Table 1). In particular, release of CAMPs into the lumen is thought to protect the mitotically-active crypt cells (which renew the epithelial cell monolayer) from colonization by pathogenic microbes. The use of transgenic animals yield a better understanding of the physiological role and regulation of these effectors. NOD1/2 have been shown to exert bactericidal activity by modulating the epithelial production of defensins—suggesting a possible mechanism whereby PRMs Pattern Recognition Molecules might protect the host from the development of CD ( FIG. 2 ). 
       Epithelial Antimicrobial Peptides: Implications for Immune System Stimulation 
       [0022]    Several recent studies have implicated antimicrobial peptides, including defensins, in protective roles in skin, oral, nasal, ocular epithelia and other epithelia mucosae, including vaginal mucosae. All mucosae share a common embryogenetic origin, since all originate from the same embroyonic layer and might be expected to exhibit similar biochemical responses, including protective defensin expression. 
         [0023]    In 2003, Dinulos et al. examined the antimicrobial activity of keratinocyte expression of β-defensin-2 in cutaneous immune defenses. β-defensin-2 expression was found to be induced by  Staphylococcus aureus, Streptococcus epidermidis, Escherichia coli , and  Pseudomonas aeruginosa , whereas  Streptococcus pyogenes  was found to be a poor 3-defensin-2 inducer. The study indicated that the ability to induce β-defensin-2 expression in combination with it antimicrobial effects may contribute to the rarity of skin infections with gram-negative bacterial organism whereas the lack of stimulation afforded by  Streptococcus pyogenes  may point towards its ability to evade immune system defences and cause skin disease. 67    
         [0024]    In the past year, Huang et al. have carried out antimicrobial assays to investigate the antimicrobial activity of a number of human ocular surface expressed antimicrobial peptides, including β-defensins 1-3, against microbes such as  Pseudomonas aerginosa  (PA),  Staphylococcus aureus  (SA) and  Staphylococcus epidermidis  (SE) in the presence of NaCl or tears. β-defensin-3 was shown to have potent activity against both SA and SE, whereas β-defensin-2 had moderate activity and β-defensin-1 showed no activity against these strains. Activity was attenuated by NaCl and tears completely inhibited the activity of β-defensin-1 and 2, but did not affect the β-defensin-3 activity. The study validates the role of some defensins as an in vivo antimicrobial. 68    
         [0025]    Towards late 2007, Vanhinsbergh showed that reduced levels of certain immunomodulatory gene expression, particularly toll-like receptors (TRLs) and defensins, are associated with allergy development, for example allergic and nonallergic rhinitis development. 69    
         [0026]    Finally, Chung et al have identified specific signaling routes that pathogens and commensals take in stimulating antimicrobial peptides such as defensins and cathelicidins in skin, oral mucosal and other epithelia, and identify these routes with development of new therapeutic agents for periodontal diseases. 70    
         [0027]    Such work reinforces the protective role of defensins in the human immune system and highlights the importance of increasing immune response to pathogens by providing routes to stimulation of defensin production in the body. 
       OTHER BACKGROUND ART 
       [0028]    U.S. Pat. No. 6,326,364 describes 5-aminosalicylate compounds such as 5-ASA as having selective antimicrobial effects in vitro. Examples show inhibitory effects against a series of  Clostridium  bacterial cultures on agar plates in aerobic and anaerobic conditions. No effect was observed against colonies of  Lactobacillus, Enterococcus  or  Bacteroides.    
       PPAR-gamma Roles 
       [0029]    Recently, the nuclear receptor peroxisome proliferator-activated receptor gamma (PPAR-gamma) was identified as a target of anti-inflammatory drugs used in the treatment of IBD, indicating a mechanism by which they mediate in vivo anti-inflammatory effect in the gut 2 , However, presumably due to the common mucosae embryogenetic origin, PPAR receptor expression has been demonstrated in various mucosal zones, other than the gut. Recent studies have suggested that PPAR-gamma may contribute to chronic inflammation of the nasal muscosa in perennial allergic rhinitis 70 . 
         [0030]    PPAR-gamma is an essential nuclear receptor controlling intestinal homeostasis by interacting with beta-catenin and T cell transcription factor (Tcf-4). Tcf-4 is an essential transcription factor in determining intestinal cell fate and in regulating the expression of natural antibiotics by Paneth cells 3 . 
         [0031]    5-aminosalycilate (5-ASA) is an anti-inflammatory drug (mesalazine) widely used in the treatment of inflammatory bowel diseases, but the mechanism underlying its intestinal effects remained poorly understood. Recently, the nuclear receptor peroxisome proliferator-activated receptor gamma (PPAR-gamma as identified as a target of 5-ASA, indicating a mechanism by which 5-ASA mediate in vivo its anti-inflammatory effect in the gut 2 . Hence given the antimicrobial properties of 5-ASA, such compounds and its derivatives might modulate the expression of antimicrobial genes through PPAR-Y activation. Rosiglitazone also effects activation of the peroxisome proliferator-activated receptors (PPARs), specifically PPAR-gamma, it has an antiinflammatory effect. 
       DEFINITIONS 
       [0032]    To facilitate understanding of the invention, a number of terms are defined below. 
         [0033]    As used herein, the term “antimicrobial peptides” refers to either α- or β-defensins (e.g. HD-5 for human defensin 5), or small polypeptides with spatially separated hydrophobic and positively charged residues. 
         [0034]    As used herein, the term “activates defensins”, when used in reference to any molecule that activates defensins, refers to a molecule (i.e., a PPAR-gamma agonist) that induces the gene expression of α- or β-defensins. 
         [0035]    As used herein, the term “primer” refers to a synthetic oligonucleotide, which is capable of acting as a point of initiation of synthesis when placed under conditions in which synthesis of a primer extension product which is complementary to a nucleic acid strand is induced. 
         [0036]    As used herein, the term “polymerase chain reaction” (hereinafter “PCR”) is a method for amplifying a segment of a target sequence in a mixture of genomic DNA without cloning or purification. The two primers are complementary to their respective strands of the double stranded target sequence. 
         [0037]    As used herein, the term “structural analogues” relate to compounds whose molecular structure act as mimics of 5-ASA in respect of their binding ability to the PPAR-gamma receptor. In particular, those compounds whose structures allow for analogous types of hydrogen bonding, and electrostatic interactions at the PPAR-gamma receptor. 
         [0038]    As used herein, the term “Caco-2 cells” refers to human Caucasian colon adenocarcinoma cells. 
         [0039]    As used herein, the term “rosiglitazone” refers to a highly selective and potent chemical agonist for PPAR-gamma. As used herein, the term “epithelia” refers to body tissues composed of layers of cells that cover organ surfaces such as surface of the skin and inner lining of digestive tract. 
         [0040]    As used herein, the term “muscosae” refers to the mucous membranes are linings of mostly endodermal origin, covered in epithelium, which are involved in absorption and secretion. Muscosae line various body cavities that are exposed to the external environment and internal organs. They are found continuous with skin at the nostrils, the lips, the ears, the genital area, and the anus. 
       OBJECT OF THE INVENTION 
       [0041]    It is an object of the present invention to provide PPAR-gamma agonists which can stimulate PPAR-gamma receptors to induce enteric defensin expression in the gut. 
         [0042]    It is a further object of the present invention to provide PPAR-gamma agonists which can stimulate PPAR-gamma receptors to induce defensin expression in epithelia or other tissue where PPAR-gamma receptors are found, in particular the skin, oral, nasal, ocular epithelia and other epithelia mucosae, including the vaginal mucosae. 
         [0043]    It is an object of the present invention to provide PPAR-gamma agonists such as 5-ASA, rosiglitazone, derivatives thereof and a series of structural analogues thereof which comprise a series of compounds found to be active on the PPAR receptor for use as stimulants to induce enteric CAMP expression in the gut, particularly defensin expression. 
         [0044]    It is a further object of the present invention to provide PPAR-gamma agonists which can stimulate PPAR-gamma receptors to induce defensin expression in the epithelia or other tissue where PPAR-gamma receptors are found, and in particular the skin, oral, nasal, ocular epithelia and other epithelia mucosae, including the vaginal mucosae. 
         [0045]    It is another object of the present invention to provide compounds that are capable of killing microbes by stimulating CAMP expression. CAMPs may be defensin and/or cathelicidin. Microbes are agents that are capable of killing bacterial, viruses, fungi and other infectious agents. 
         [0046]    It is a further object still to provide compounds that enhance the body&#39;s defense mechanisms through expression of CAMP. CAMP may be defensin and/or cathelicidin. 
         [0047]    It is yet further object of the present invention to provide compounds for use in the intervention of gastrointestinal tract conditions such as Crohn&#39;s disease, ulcerative colitis, intestinal bowel syndrome and acute diverticulitis. 
         [0048]    It is a further object of the invention to provide an intervention for the prevention of conditions such as acute diverticulitis in patients affected by colonic diverticulosis, indeterminate colitis and infectious colitis. 
         [0049]    It is yet further object of the present invention to provide compounds for use in the intervention of skin inflammatory conditions and infections such as impetigo, erysipela, dermatitis, folliculitis, acne and vulgaris. 
         [0050]    It is yet further object of the present invention to provide compounds for use in the intervention of muscoal inflammatory conditions and infections such as those affecting ocular, oral, nasal or vaginal mucosae, in particular conditions such as ocular inflammation and infections, periodontal disease, allergic and non allergic rhinitis and bacterial vaginosis. 
         [0051]    It is a further object of the invention to provide compounds for use in preparation of a medicament for the treatment and prevention of such diseases. 
         [0052]    It is a further object of the present invention to provide a method to allow the development of novel therapeutic strategies based on regulating CAMP expression in the gastrointestinal tract of susceptible individuals. Compounds having anti-inflammatory, antibiotic and/or, antimicrobial effects may be identified through stimulation of CAMP expression, particularly defensin expression. 
         [0053]    It is an object of the invention to provide modulators of CAMP expression. 
       SUMMARY OF THE INVENTION 
       [0054]    According to the present invention, there is provided a method which uses a range of chemical entities which act on PPAR-gamma, to induce CAMP production. Such compounds can be used in the induction of CAMP expression in tissues having PPAR-gamma receptors, such as epithelia and/or mucosae. In particular, the compounds can be used to induce enteric CAMP expression in the gut. Defensins are examples of CAMPs. 
         [0055]    According to the present invention, there are provided PPAR-gamma agonists such as 5-ASA, rosiglitazone, derivatives and a series of structural analogues which comprise a series of compounds found to be active on the PPAR-gamma receptor, for use as stimulants to induce CAMP expression in tissue, particularly defensin expression. 
         [0056]    According to the present invention, there are provided PPAR-gamma agonists such as 5-ASA, rosiglitazone, derivatives and a series of structural analogues which comprise a series of compounds found to be active on the PPAR-gamma receptor, for use as stimulants to induce CAMP expression in epithelia and/or mucosae having PPAR-gamma receptors, particularly defensin expression. 
         [0057]    According to the present invention, there are provided PPAR-gamma agonists such as 5-ASA, rosiglitazone, derivatives and a series of structural analogues which comprise a series of compounds found to be active on the PPAR-gamma receptor, for use as stimulants to induce enteric CAMP expression in the gut, particularly defensin expression. 
         [0058]    The compounds described herein can be defined according to the general formula (I): 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    in which
 
R 1  and R 2 , which may be identical or different, are selected from the group comprising —H or a linear or branched alkyl group having from 1 to 6 carbon atoms or together form an aromatic or aliphatic ring with 5 or 6 atoms;
 
Y and Z, which may be identical or different, are selected from the group comprising —H, —OH, —COOH, —OR 3 , —CH(OR 3 )COOH, in which R 3  is selected from H, phenyl, benzyl, —CF 3  or —CF 2 CF 3 , vinyl, allyl and a linear or branched alkyl group having from 1 to 6 carbon atoms;
 
or
 
compounds according to the general formula (Ia):
 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    in which
 
R 1  and R 2 , which may be identical or different, are selected from the group comprising H, —C n H 2n-1 , where n=1-6, a linear or branched alkyl group having from 1 to 6 carbon atoms, or together form an aromatic or aliphatic ring with 5 or 6 atoms;
 
R 3  is selected from —CO—CH, —NHOH, —OH, —OR 6  in which R 6  is a linear or branched alkyl group having from 1 to 6 carbon atoms;
 
R 4  is selected from H, a linear or branched alkyl group having from 1 to 6 carbon atoms, phenyl, benzyl, —CF 3  or —CF 2 CF 3 , vinyl or allyl; R 5 , R 7 , R 8  are hydrogen atoms;
 
or
 
R 3  and R 4 , R 4  and R 5 , or R 7  and R 8  together form a ring, fused to the benzene, aromatic or aliphatic ring with 5 or 6 atoms comprising from 1 to 2 heteroatoms selected independently from the group comprising N, O.
 
         [0059]    The compounds can be used in the induction of CAMP expression in tissues having PPAR-gamma receptors, such as epithelia and/or mucosae. In particular, the compounds can be used to induce enteric CAMP expression in the gut. 
         [0060]    In one aspect, according to the present invention, compounds which can be used in such methods include compounds comprising the general formula (I) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    in which
 
R 1  and R 2 , which may be identical or different, are selected from the group comprising —H or a linear or branched alkyl group having from 1 to 6 carbon atoms or together form an aromatic or aliphatic ring with 5 or 6 atoms;
 
Y and Z, which may be identical or different, are selected from the group comprising —H, —OH, —COOH, —OR 3 , —CH(OR 3 )COOH, in which R 3  is selected from H, phenyl, benzyl, —CF 3  or —CF 2 CF 3 , vinyl, allyl and a linear or branched alkyl group having from 1 to 6 carbon atoms.
 
         [0061]    In another aspect, the present invention also relates to use of a subgroup of compounds, of general formula (I*) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    in which
 
R 1  and R 2 , which may be identical or different, are selected from the group comprising —H or a linear or branched alkyl group having from 1 to 6 carbon atoms
 
Y and Z, which may be identical or different, are selected from the group comprising —H, —OH, —COOH, —OR 3 , —CH(OR 3 )COOH, in which R 3  is selected from —H and a linear or branched alkyl group having from 1 to 6 carbon atoms.
 
         [0062]    In some embodiments of the invention, Z and Y are different. In some embodiments of the invention, at least one of Y or Z terminates in —COOH. Therefore, in some embodiments of the invention, Y or Z (and in some embodiments at least one of Y or Z, and in some embodiments, only one of Y or Z) is —COOH. In some embodiments of the invention, Y or Z (and in some embodiments at least one of Y or Z, and in some embodiments, only one of Y or Z) is —CH(OR 3 )COOH. 
         [0063]    In another aspect still, the present invention also relates to use of compounds in the methods of the invention, according to both formula (I) and (I*), except wherein Y and Z, which may be identical or different, are selected from the group comprising —H, —COON, —OR 3 , —CH(OR 3 )COOH. 
         [0064]    In some embodiments of the invention, when Y is —H and Z is —CH(OH)COOH, the group NR 1 R 2  may be connected at the 3′ position. 
         [0065]    In other embodiments of the invention, when Z is —OCH 3  and Y is —COOH, the group NR 1 R 2  may be connected at the 4′ position. 
         [0066]    In some embodiments of the invention, when Y is —H and Z is —CH(OCH 3 )COOH, the group NR 1 R 2  may be connected at the 4′ position. 
         [0067]    In particular, the aforementioned linear or branched alkyl group having from 1 to 6 carbon atoms can be selected from —CH 3 , —CH 2 CH 3 , —CH(CH 3 ) 2 , —CH 2 CH 2 CH 3 , —C n H 2n-1 , where n=1-6. 
         [0068]    The compounds of formula (I) and (I*) can be selected from the group comprising:
   3-(3′-aminophenyl)2-hydroxypropanoic acid (compound 20)   2-(4-aminophenyl)2-methoxyacetic acid (compound 23)   2-(3-aminophenyl)2-ethoxyacetic acid (compound 32)   2-(4-aminophenyl)2-ethoxyacetic acid (compound 33)   3-(4′-aminophenyl)2-methoxypropionic acid (compound 34) “R34”   3-(4′-aminophenyl)2-ethoxypropionic acid (compound 39)   3-(3′-aminophenyl)2-ethoxypropionic acid (compound 40).   
 
         [0076]    The above compound names can also be written in standard chemical nomenclature as follows (which nomenclature will be used throughout the text):
   (±)-2-hydroxy-3-(3′-aminophenyl) propionic acid (compound 20)   (±)-2-methoxy-2-(4′-aminophenyl)acetic acid (compound 23)   (±)-2-ethoxy-2-(3′-aminophenyl)acetic acid (compound 32)   (±)-2-ethoxy-2-(4′-aminophenyl)acetic acid (compound 33)   (±)-2-methoxy-3-(4′-aminophenyl)propionic acid (compound 34) “R34” (racemic form)   (±)-2-ethoxy-3-(4′-aminophenyl)propionic acid (compound 39)   (±)-2-ethoxy-3-(3′-aminophenyl)propionic acid (compound 40).   
 
         [0084]    In particular, the compounds used in the methods of the present invention can be enantiomers of the following racemic mixtures:
   (R,S)-2-hydroxy-2-(3-aminophenyl)acetic acid (compound 10)   (R,S)-2-hydroxy-2-(4-aminophenyl)acetic acid (compound 11)   (R,S)-2-hydroxy-3-(4′-aminophenyl)propionic acid (compound 21)   (R,S)-2-methoxy-2-(3′-aminophenyl)acetic acid (compound 22)   (R,S)-2-methoxy-3-(3′-aminophenyl)propionic acid (compound 35)   (R,S)-2-methoxy-3-(3-aminophenyl)propionic acid (compound 34) “R34” (racemic form)   
 
         [0091]    Enantiomers of R34:
   (+) 2-S-methoxy-3-(3-aminophenyl)propionic acid (compound 34) “34-E1” or “E-1”.   (−) 2-R-methoxy-3-(3-aminophenyl)propionic acid (compound 34) “34-E2” or “E-2”   
 
         [0094]    Racemic mixtures of the compounds may also be used in the methods described herein. Examples of racemic mixtures include but are not limited to
   (±)-2-hydroxy-2-(3′-aminophenyl)acetic acid (compound 10)   (±)-2-hydroxy-2-(4′-aminophenyl)acetic acid (compound 11)   (±)-2-hydroxy-3-(4′-aminophenyl)propionic acid (compound 21)   (±)-2-methoxy-2-(3′-aminophenyl)acetic acid (compound 22)   (±)-2-methoxy-3-(3′-aminophenyl)propionic acid (compound 35)   (±)-2-methoxy-3-(4′-aminophenyl)propionic acid (compound 34) “R34” (racemic form).   
 
         [0101]    Compositions containing an excess of one enantiomer over another, for any of the stereoisomeric compounds described herein, may also be used in the methods described herein. 
         [0102]    According to one embodiment, the compounds which may be used in the method of the present invention include those, where R 3  of the compounds of formula (I) may be H according to the following formula (II) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , X and Y are defined above. 
         [0103]    According to another embodiment, the compounds which may be used include those where R 3  of the compounds of formula (I) can be —CH 3  according to the following formula (III) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , X and Y are defined above. 
         [0104]    According to yet another embodiment, the compounds which may be used include those where R 3  of the compounds of formula (I) can be —CH 2 CH 3  according to the following formula (IV) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , X and Y are defined above. 
         [0105]    According to another embodiment still, the compounds which may be used include those where, R 3  of the compounds of formula (I) can be —CH 2 CH 3  according to the following formula (V) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , X and Y are defined above. 
         [0106]    According to another embodiment still, the compounds which may be used include those where, R 3  of the compounds of formula (I) can be —CH 3  according to the following formula (VI) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , X and Y are defined above. 
         [0107]    According to the invention, one enantiomer of (R,S)-2-methoxy-3-(3-aminophenyl)propionic acid having formula (VI), namely, (−) 2-R-methoxy-3-(3-aminophenyl)propionic acid (compound 34) “34-E2” or “E-2”, has been found to be particularly effective in gut defensin expression induction ( FIGS. 8 &amp; 9 ). According to the present invention some enantiomers may provide superior CAMP expression than their stereoisomers. In other embodiments, some racemic mixtures may provide superior CAMP expression than their individual stereoisomers. 
         [0108]    According to another embodiment still, the compounds which may be used include those where, R 3  of the compounds of formula (I) can be —CH 3  according to the following formula (VII) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , X and Y are defined above. 
         [0109]    According to another embodiment still, the compounds which may be used include those where, R 3  of the compounds of formula (I) can be —CH 2 CH 3  according to the following formula (VIII) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , X and Y are defined above. 
         [0110]    According to another embodiment still, the compounds which may be used include those where, R 3  of the compounds of formula (I) can be —CH 2 CH 3  according to the following formula (IX) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , X and Y are defined above. 
         [0111]    According to another embodiment, the compounds which may be used include those where, R 3  of the compounds of formula (I) can be —CH 3  according to the following formula (X) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , X and Y are defined above. 
         [0112]    Preferably, the compounds of formula (Ia) which may be use in the methods of the invention can be selected from the group comprising:
   (±)-2-hydroxy-3-(3′-aminophenyl)propionic acid (compound 20)   (±)-2-methoxy-2-(4′-aminophenyl)acetic acid (compound 23)   (±)-2-ethoxy-2-(3′-aminophenyl)acetic acid (compound 32)   (±)-2-ethoxy-2-(4′-aminophenyl)acetic acid (compound 33)   (±)-2-methoxy-3-(4′-aminophenyl)propionic acid (compound 34) “R34”   (±) 2-S-methoxy-3-(3-aminophenyl)propionic acid (compound 34) “34-E1” or “E-1”   (−) 2-R-methoxy-3-(3-aminophenyl)propionic acid (compound 34) “34-E2” or “E-2”   (±)-2-ethoxy-3-(4′-aminophenyl)propionic acid (compound 39)   (±)-2-ethoxy-3-(3′-aminophenyl)propionic acid (compound 40), whose formulas are shown earlier.   
 
         [0122]    According to the present invention, compounds of general formula (Ia) can be used in the methods of the invention described herein: 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    in which
 
R 1  and R 2 , which may be identical or different, are selected from the group comprising H, —C n H 2n-1 , where n=1-6, a linear or branched alkyl group having from 1 to 6 carbon atoms, or together form an aromatic or aliphatic ring with 5 or 6 atoms;
 
R 3  is selected from —CO—CH, —NHOH, —OH, —OR 6  in which R 6  is a linear or branched alkyl group having from 1 to 6 carbon atoms;
 
R 4  is selected from H, a linear or branched alkyl group having from 1 to 6 carbon atoms, phenyl, benzyl, —CF 3  or —CF 2 CF 3 , vinyl or allyl; R 5 , R 7 , R 8  are hydrogen atoms;
 
or
 
R 3  and R 4 , R 4  and R 5 , or R 7  and R 8  together form a ring, fused to the benzene, aromatic or aliphatic ring with 5 or 6 atoms comprising from 1 to 2 heteroatoms selected independently from the group comprising N, O.
 
         [0123]    The invention also relates to the use in the method of the present invention of the specific subgroup of compounds of general formula (Ia*) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    in which
 
R 1  and R 2 , which may be identical or different, are selected from the group comprising H, —CO—CH 3 , —C n H 2n-1 , where n=1-6, a linear or branched alkyl group having from 1 to 6 carbon atoms, or together form an aromatic or aliphatic ring with 5 or 6 atoms;
 
R 3  is selected from —NHOH, —OH, —OR 6  in which R 6  is a linear or branched alkyl group having from 1 to 6 carbon atoms;
 
R 4  is selected from —H, a linear or branched alkyl group having from 1 to 6 carbon atoms; R 5 , R 7 , R 8  are hydrogen atoms;
 
or
 
R 3  and R 4 , R 4  and R 5 , or R 7  and R 8  together form a ring, fused to the benzene, aromatic or aliphatic ring with 5 or 6 atoms comprising from 1 to 2 heteroatoms selected independently from the group comprising N, O.
 
         [0124]    The aforementioned linear or branched alkyl group of formula (Ia) or (Ia*) having from 1 to 6 carbon atoms can be selected from —CH 3 , —C 2 H 5 , isopropyl, propyl, CH 2n-1 , where n=1-6. 
         [0125]    In some embodiments of both formula (Ia) and (Ia*) the invention, the compounds which may be used include those where, R 7  and R 8  may form a ring. Thus R 3  and R 4  or R 4  and R 5  may together form a ring, fused to the benzene, aromatic or aliphatic ring with 5 or 6 atoms comprising from 1 to 2 heteroatoms selected independently from the group comprising N, O. 
         [0126]    In some embodiments of both formula (Ia) and (Ia*) the invention, R 7  and R 3  may form a ring except when R 4  is CH 3 . In some embodiments of both formula (Ia) and (Ia*) the invention, R 7  and R 8  may form a ring when R 4  is selected from H. In some embodiments, the invention relates to the ketolenes provided by the invention. 
         [0127]    In some embodiments the compounds which may be used include those of both formula (Ia) and (Ia*) where, R 1  and R 2  may form a ring. Thus R 1  and R 2 , which may be identical or different, may be selected from the group comprising —H, —C n H 2n-1 , a linear or branched alkyl group having from 1 to 6 carbon atoms, where n=1-6. 
         [0128]    In some embodiments the compounds which may be used include those of both formula (Ia) and (Ia*) where, R 4  may be branched. Thus R 4  may be selected from H, a linear alkyl group having from 1 to 6 carbon atoms; R 5 , R 7 , R 8  are hydrogen atoms. 
         [0129]    In some embodiments, R 4  may be branched when the amino group is at position 4′ on the phenyl ring. 
         [0130]    In some embodiments, the linear alkyl group may have only 1 carbon atom (i.e., CH 3 ). 
         [0131]    In some embodiments of both formula (Ia) and (Ia*) the invention, R 1  and R 2  are —H. In some embodiments of both formula (Ia) and (Ia*) of the invention, R 2  may not be different to 
         [0132]    In some embodiments the compounds which may be used include those of both formula (Ia) and (Ia*) where R 3  is —OH, R 4  is selected from the group consisting —H, a branched alkyl group having from 1 to 6 carbon atoms, or R 3  and R 4 , together form a ring, fused to the benzene, aromatic or ring with 5 or 6 atoms comprising from 1 to 2 heteroatoms selected independently from the group comprising N, O. In some embodiments, the branched alkyl group may be —CH(CH 3 ) 2 . In some embodiments, the branched alkyl group may be —CH(CH 3 ) 2  at the R 8  position. In some embodiments, R 3  and R 4  form a 5-membered aliphatic ring with a single O atom. 
         [0133]    In some embodiments the compounds which may be used include those of both formula (Ia) and (Ia*) where R 3  is —OH and R 4  is —H, the group NR 1 R 2  can be at the 4′ position (and should be at R 5  or R 8 ). In some embodiments, this may be particularly the case where R 1  and R 2  are —CH 3 . 
         [0134]    In some embodiments the compounds which may be used include those of both formula (Ia) and (Ia*) where, R 1  and R 2  are the same. 
         [0135]    In some embodiments the compounds which may be used include those of both formula (Ia) and (Ia*) where R 3  is —OH and R 4  is —H, the group —NR 1 R 2  may be at the R 5  (and should be at the R 8  position). In some embodiments, this may be where R 1  and R 2  are —H. 
         [0136]    In some embodiments the compounds which may be used include those of both formula (Ia) and (Ia*) where R 3  is —NHOH, R 4  is a linear or branched alkyl group having from 1 to 6 carbon atoms, (or, if formula (I) phenyl, benzyl, —CF 3  or —CF 2 CF 3 , vinyl or allyl), R 3  and R 4 /together form a ring, fused to the benzene, aromatic or aliphatic ring with 5 or 6 atoms comprising from 1 to 2 heteroatoms selected independently from the group comprising N, O. 
         [0137]    In some embodiments the compounds which may be used include those of both formula (Ia) and (Ia*) where R 3  is —NHOH, and R 4  is a linear or branched alkyl group having from 2 carbon atoms, the group —NR 1 R 2  may be at the 4′ position and can be at the R 8  position. 
         [0138]    In some embodiments the compounds which may be used include those of both formula (Ia) and (Ia*) where R 3  is —NHOH, and R 4  is —H, the group —NR 1 R 2  may be at R 8  and must be at the 4′ position. According to one embodiment, R 3  and R 4  of the compounds of formula (Ia) and (Ia*) can form a ring according to the following formula (XII) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , R 5 , R 7  and R 8  are defined above. 
         [0139]    According to another embodiment R 4  and R 5  of the compounds used according to formula (Ia) and (Ia*) can form a ring according to the following formula (XII) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , R 3 , R 7  and R 8  are defined above. 
         [0140]    According to a further embodiment R 7  and R 8  of the compounds used according to formula (Ia) and (Ia*) can form a ring according to the following formula (XIII) or (XIV) 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    while R 1 , R 2 , R 3 , R 4  and R 5  are defined above. 
         [0141]    In particular, the compounds of formula (Ia) and (Ia*) can be used in according to the methods described in the present invention and can be selected from the group comprising:
   4-amino-N-hydroxy-2-methoxybenzamide (compound 13)   5-amino-N-hydroxy-2-methoxybenzamide (compound 14)   5-amino-2,3-dihydrobenzofuran-7-carboxylic acid (compound 17)   5-amino-2-ethoxy-N-hydroxybenzamide (compound 26)   6-amino-2,2-dimethyl-4H-benzo[1,3]dioxin-4-one (compound 28)   1,2,3,4-tetrahydro-6-hydroxyquinoline-5-carboxylic acid (compound 29)   5-amino-2-isopropoxybenzoic acid (compound 31)   6-methoxy quinoline-5-carboxylic acid (compound 36)   6-methoxy-1,2,3,4-tetrahydroquinoline-5-carboxylic acid (compound 37)   5-diisopropylaminosalicylic acid (compound 38)   4-diisopropylaminosalicylic acid (compound 42).   
 
         [0153]    The present invention also provides for use of compounds wherein R 1  and R 2 , are selected from the group consisting of —H and —CH(CH 3 ) 2 . R 1  and R 2  may both be identical. In some embodiments, R 1  and R 2  may be —CH(CH 3 ) 2 . 
         [0154]    One example comprises use of the following compound (compound 38): 
         [0000]    
       
                 
         
             
             
         
       
     
         [0000]    In other embodiments of the invention, R 1  and R 2 , are both —H. 
         [0155]    The present invention also provides for use of compounds wherein R 3  is selected from the group consisting of —NHOH and —OH. In some embodiments R 3  may be —NHOH. One example comprises the following compound (compound 13): 
         [0000]    
       
                 
         
             
             
         
       
     
         [0156]    A further example comprises use of the following compound (compound 14): 
         [0000]    
       
                 
         
             
             
         
       
     
         [0157]    A further example comprises use of the following compound (compound 26): 
         [0000]    
       
                 
         
             
             
         
       
     
         [0158]    In some embodiments of the invention, R 3  may be —OH. 
         [0159]    A suitable example comprises use of the following compound (compound 17): 
         [0000]    
       
                 
         
             
             
         
       
     
         [0160]    A further example comprises use of the following compound (compound 31): 
         [0000]    
       
                 
         
             
             
         
       
     
         [0161]    In some embodiments of the invention, R 4  may be —H. In some embodiments of the invention, R 4  may be CH 3 . In some embodiments of the invention, R 4  may be, —CH 2 CH 3 . In some embodiments of the invention, R 4  may be —CH(CH 3 ) 2 . 
         [0162]    A further example comprises use of the following compound (compound 28): 
         [0000]    
       
                 
         
             
             
         
       
     
         [0163]    In some embodiments of the invention, R 3  and R 4  may together form an aliphatic ring, fused to the benzene, of 5 or 6 atoms comprising one hetero atom 0 (oxygen). The present invention also relates to methods of treatment of humans and/or mammals (including rodents, farm animals, domestic pets, mice, rats, hamsters, rabbits, dogs, cats, pigs, sheep, cows, horses). 
         [0164]    In particular, apart from the use of the specific compounds mentioned above, the following compounds can be used for the methods and applications described herein:
   5-aminosalicylo-hydroxamic acid (compound 5)   3-dimethylaminosalicylic acid (compound 6)   2-methoxy-4-aminobenzoic acid (compound 7)   2-methoxy-5-aminobenzoic acid (compound 8)   5-methylaminosalicylic acid (compound 9)   4-methylaminosalicylic acid (compound 12)   4-acetylaminosalicylic acid (compound 16)   2-ethoxy-4-aminobenzoic acid (compound 18)   2-ethoxy-5-aminobenzoic acid (compound 19)   4-dimethylaminosalicylic acid (compound 24)   2-ethoxy-4-aminobenzoylhydroxamic acid (compound 25)   6-hydroxyquinoline-5-carboxylic acid (compound 27)   2-(2-propyl)oxy-4-aminobenzoic acid (compound 30)   4-(1-piperazinyl)salicylic acid (compound 41).   
 
         [0179]    In addition to the use of the above-mentioned compounds, the present invention provides for the use of the following compounds (compound number follows prefix “2_”): 
         [0000]    
       
                 
         
             
             
         
       
       
                 
         
             
             
         
       
       
                 
         
             
             
         
       
     
         [0180]    According to the present invention there are provided compounds for use in the intervention of gastrointestinal tract conditions such as Crohn&#39;s disease, ulcerative colitis, intestinal bowel syndrome and acute diverticulitis. 
         [0181]    In one aspect of the invention, there are provided compounds for use in the prevention of conditions such as acute diverticulitis in patients affected by colonic diverticulosis, indeterminate colitis and infectious colitis. 
         [0182]    In another aspect of the invention there are provided compounds for use in the intervention of skin inflammatory conditions and infections such as impetigo, erysipela, dermatitis, folliculitis, acne and vulgaris. 
         [0183]    In another aspect of the invention there are provided compounds for use in the intervention of muscoal inflammatory conditions and infections such as those affecting ocular, oral, nasal or vaginal mucosae including those such as ocular inflammation and infections, periodontal disease, allergic and non allergic rhinitis and bacterial vaginosis. 
         [0184]    The compounds according to the present invention can be used advantageously in the medical field to stimulate PPAR-gamma to produce CAMPs. CAMPS include defensin and/or cathelicidin. Therefore another aspect of the present invention relates to a pharmaceutical composition comprising one or more compounds as defined above as active principles in combination with one or more pharmaceutically acceptable excipients or adjuvants. 
         [0185]    In another aspect, the invention provides compounds for use in preparation of a medicament for the treatment and prevention of diseases such as Crohn&#39;s disease, ulcerative colitis, intestinal bowel syndrome, acute diverticulitis and prevention of conditions such as acute diverticulitis in patients affected by colonic diverticulosis, indeterminate colitis and infectious colitis. 
         [0186]    In another aspect, the invention provides compounds for use in preparation of a medicament for the treatment and prevention of diseases involving skin inflammatory conditions and infections such as impetigo, erysipela, dermatitis, folliculitis, acne and vulgaris. 
         [0187]    In another aspect, the invention provides compounds for use in preparation of a medicament for the treatment and prevention of muscoal inflammatory conditions and infections such as those affecting ocular, oral, nasal or vaginal mucosae including those such as ocular inflammation and infections, periodontal disease, allergic and non allergic rhinitis and bacterial vaginosis. 
         [0188]    The present invention also relates to methods of treatment of humans and/or mammals (including rodents, farm animals, domestic pets, mice, rats, hamsters, rabbits, dogs, cats, pigs, sheep, cows, horses). 
         [0189]    In another aspect still, the invention provides a method to allow the development of novel therapeutic strategies based on regulating CAMP expression in the gastrointestinal tract of susceptible individuals. The invention provides for screening for compounds having potential anti-inflammatory and/or antimicrobial effects. Such compound leads may be identified through stimulation of CAMP expression, particularly defensin expression. 
         [0190]    In a particular aspect, the invention to provide modulators and up-regulators of CAMP expression, particularly defensin expression. Up-regulation or stimulation of CAMP expression, in particular defensin expression, will lead to anti-inflammatory and antimicrobial effects in the body. This is particularly the case with respect to defensin production which gives rise to antibacterial effects, where the stimulating compounds lead to defensin production as so give rise to induced antimicrobial effects, using physiological/biochemical pathways in the body. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0191]      FIG. 1 : Rosiglitazone activates HD-5 in Caco-2 cells, as determined by quantitative PCR analysis. 
           [0192]      FIG. 2 : Compound 14 activates HD-5 in Caco-2 cells, as determined by quantitative PCR analysis. 
           [0193]      FIG. 3 : Compound 40 activates HD-5 in Caco-2 cells, as determined by quantitative PCR analysis. 
           [0194]      FIG. 4 : Compound 39 activates HD-5 in Caco-2 cells, as determined by quantitative PCR analysis. 
           [0195]      FIG. 5 : Mesalazine activates HD-5 in Caco-2 cells, as determined by quantitative PCR analysis. 
           [0196]      FIG. 6 : Structure of human defensins and cathelicidin 
           [0000]    A. Human sequences of the enteric α-defensin HD-5, the 3-defensin HBD-2 and the cathelicidin hCAP-18. Gray arrows depict the cleavage site for HD-5 and cathelicidin. Patterns of the three intramolecular disulfide bonds (S—S) of the α- and β-defensins, have been notified in both the schematic and tri-dimensional structures (blue sticks).
 
B. A three-dimensional solution (h-BD2 and cathelicidin) or crystal (HD-5) structure of defensins and cathecidin is displayed. The Protein Data Bank accession numbers used for the illustration are the following: 1ZMP for HD-5, 1E4Q for h-BD2 and 2FCG for LL-37 (C-terminal fragment of hCAP-18). The beta turns are depicted in orange and the alpha-helices in red. The hydrophobicity of the molecules is displayed.
 
           [0197]      FIG. 7 : A pathophysiological model for chronic intestinal inflammation. Once microbes and/or their products are sensed by TLRs and/or NODs (left side of the figure), CAMPs are synthesized via the action of NF-κB and/or other transcription factors (cf. the main text). Following their secretion and extracellular processing, the CAMPs (i) promote tolerance and the recruitment of inflammatory cells, (ii) prevent invasion of microbial pathogens and (iii) protect against the development of chronic intestinal inflammation. Abnormal antimicrobial peptide synthesis and/or function might lead to aberrant activation of the adaptive immune system and to intestinal inflammation (right side of the figure) by microbial threats and/or impaired innate immunity (i.e. NOD2 mutations). 
           [0198]      FIG. 8 :  5 -ASA, Rosiglitazone, racemic R34 &amp; enantiomer 34-E2 induce the expression of hBD1 (human defensin-1) in Caco-2 cells. 
           [0199]      FIG. 9 : Effect of racemic R34 &amp; enantionmes 34-E1 &amp; 34-E2 on PPAR-gamma and LL37 (defensin) level at the mRNA level in healthy mice (n=5). Administration of 5-ASA (30 mM), R34 (1 mM), E1 and E2 (1 mM) by enema during 10 days in healthy mice induced colon PPARy mRNA and defensin expression. 
       
    
    
       [0200]    Table 1. Versatile functions of the defensins and cathelicidins. The functions of α/β-defensins and cathelicidins are listed in the Table and discussed in the main text. 
       DETAILED DESCRIPTION OF THE INVENTION 
       [0201]    Given the direct negative role of the peroxisome proliferator-activated receptor gamma (PPAR-gamma) on the Wnt/Tcf/beta-catenin signaling pathway, PPAR-gamma activation was examined to investigate defensin biogenesis. 
       Experimental Data 
       [0202]    The tested PPAR-gamma agonists, rosiglitazone (at 1 μM for 1, 3 or 6 hours) and others, activates HD-5 in Caco-2 cells, as determined by quantitative PCR analysis ( FIGS. 1-5 ). 
       Methods 
       [0203]    Cultured intestinal epithelial cell lines, namely Caco-2 (of human origin) and ICcl2 (of mice origin), were treated with the GSK-3 inhibitor LiCl (20 microM) or the phosphatase inhibitor calyculin (50 nM) followed or not by stimulation with the PPAR-gamma agonist such as rosiglitazone (1 microM). 
         [0204]    The expression of defensin and known target genes of both Wnt and PPAR-gamma signaling pathways were investigated by quantitative real-time PCR. The activation of GSK3, beta-catenin, NF-kappaB, ERK1/2, SAPK/JNK and p38 was measured by specific immunoblotting. 
         [0205]    To investigate the antimicrobial role of PPAR-gamma, the intracellular replication of the Crohn&#39;s disease associated  Escherichia coli  (LF82) was measured upon or not stimulation with rosiglitazone in the Raw macrophage cell line. 
       Results 
       [0206]    Incubation with rosiglitazone and other PPAR-gamma activators significantly increased the expression of both alpha- (HD-5 and HD-6) and beta- (Defb10) defensins by intestinal epithelial cells ( FIG. 3 ). Such antimicrobial gene expression was synergized following co-stimulation by calyculin that promotes beta-catenin degradation. Accordingly, reduced intracellular replication of LF82 through PPAR-gamma activation by rosiglitazone was observed. 
         [0207]    Conversely, the expression of a Wnt/Tcf/beta-catenin target gene, cyclin-D1, and the stability of the beta-catenin was markedly decreased upon stimulation by both calyculin and rosiglitazone. 
         [0208]    Finally, LiCl, an activator of the Wnt/TCF/beta-catenin-dependent signalling pathway, blocked the rosiglitazone-induced defensin gene expression upon co-stimulation. 
         [0209]    Incubation with test substances significantly increased the expression of both alpha- (HD-5 and HD-6) and beta- (Defb10) defensins by intestinal epithelial cells ( FIG. 3  shows effect of rosiglitazone). Accordingly, reduced intracellular replication of LF82 through PPAR-gamma activation by rosiglitazone was observed. 
       Conclusion 
       [0210]    Taken as a whole, the results indicate that PPAR-gamma activation promotes the induction of an antimicrobial gene programme by negatively regulating the formation of the Tcf/beta-catenin complex. These findings highlight the therapeutical potential of PPAR-gamma in complementing defensins deficiency in many gasterointestinal disorders such as Crohn&#39;s disease, ulcerative colitis, intestinal bowel syndrome, acute diverticulitis and for the prevention of condition such as acute diverticulitis in patients affected by colonic diverticulosis, indeterminate colitis and infectious colitis. 
         [0211]    Furthermore, these findings highlight the therapeutical potential of PPAR gamma agonists in complementing defensins deficiency in other mucosal disorders including but not limited to those such as ocular inflammation and infections, periodontal disease, allergic and non allergic rhinitis, bacterial vaginosis and skin inflammatory conditions and infections such as impetigo, erysipela, dermatitis, folliculitis, acne and vulgaris. 
         [0000]    In Vitro Studies with Racemic Compound 34 and Enantiomers 34-E1 &amp; 34-E2 Materials 
         [0212]    5-ASA was purchased at Sigma-Aldrich (St Quentin Fallavier, France). Rosiglitazone was synthesized in the laboratory according to standard procedures. The racemic compound 34 and the two enantiomers of the compound, 34-E1 and 34-E2 were provided by Giuliani SpA (Milano, Italy). Compound were re-suspended in DMEM medium (Gibco) and adjusted at pH=7 if necessary with 10N NaOH. 
         [0000]    Regulation of the Expression of hBD1 Defensin in Colonic Epithelial Cells 
       Cell Lines 
       [0213]    The colon carcinoma cell line Caco-2 (ATCC HTB-39) was routinely grown in DMEM supplemented respectively with 10% or 20% heat-FCS, and antibiotics. Cells were grown in monolayers, incubated at 37° C. in 5% CO2 and 95% relative humidity. 
         [0214]    Cell were stimulated by 5-ASA, R34, 34-E1 and 34-E2 for 24 h. Total RNA was isolated from cells using Rneasy kit (Macherey Nagel, Hoerdt, France) according to the manufacturer&#39;s instructions. RNA quantification was performed using spectrophotometry. After treatment at 37° C. for 30 min with 20-50 units of RNase-free DNase I (Roche Diagnostics Corporation, Indianapolis, Ind., USA), oligo-dT primers (Roche Diagnostics Corporation, Indianapolis, USA) were used to synthesize single-stranded cDNA. mRNAs were quantified using SYBR green Master Mix (Applera, Courtaboeuf, France) with human specific oligonucleotides for hBD1(S:5′-ATACTTCAAAAGCAATTTTCCTTTAT-3′; AS:5′-TTgTCTGAGATGGCCTCAggTggTAAC-3′) in a GeneAmp Abiprism 7000 (Applera, Courtaboeuf, France). In each assay, calibrated and no-template controls were included. Each sample was run in triplicate. SYBR green dye intensity was analyzed using the Abiprism 7000 SDS software (Applera, Courtaboeuf, France). All results were normalized to the unaffected housekeeping gene of human β-actin (S:5′-TCACCCACACTgTgCCCATCTACg-3′; AS:5′-CAgCggAACCgCTCATTgCCAATg-3′). 
         [0000]    Evaluation of β-defensin Expression in Healthy Mice 
         [0215]    5-ASA (30 mM), racemic R34 and 34-E1 &amp; 34-E2 (1 mM) were administrated by intrarectal installations for 8 days. Post-mortem, total RNA was isolated from whole mice colonic tissues using Rneasy kit (Macherey Nagel, Hoerdt, France) according to the manufacturer&#39;s instructions. RNA quantification was performed using spectrophotometry. After treatment at 37° C. for 30 min with 20-50 units of RNase-free DNase I (Roche Diagnostics Corporation, Indianapolis, Ind., USA), oligo-dT primers (Roche Diagnostics Corporation, Indianapolis, USA) were used to synthesize single-stranded cDNA. mRNAs were quantified using SYBR green Master Mix (Applera, Courtaboeuf, France) with mouse specific oligonucleotides for LL37(S:5′-gCTgATTCTTTTgACATCAgCTgTAA-3′ AS:5′-gCCAgCCgggAAATTTTCT-3′) in a GeneAmp Abiprism 7000 (Applera, Courtaboeuf, France). In each assay, calibrated and no-template controls were included. Each sample was run in triplicate. SYBR green dye intensity was analyzed using the Abiprism 7000 SDS software (Applera, Courtaboeuf, France). All results were normalized to the unaffected housekeeping gene β-actin (S:5′-gggTCAgAAggATTCCTATg-3′; AS:5′ ggTCTCAAACATgATCTggg-3′). 
       In Vivo Study 
     Regulation of Visceral Pain in Rats 
     Animals 
       [0216]    Male Sprague-Dawley rats (Charles River, l&#39;Arbresle, France) weighing 175-200 g were used in this study. Rats were maintained in laboratory conditions for 1 week before experiment. 
         [0217]    The animals were housed 5 per cage with food and water available ad libitum. All studies were performed in accordance with the proposal of the committee for Research and Ethical Issues of the International Association for the Study of Pain (Zimmermann M, Pain 1983; 16:109-110). Great care was taken, particularly with regard to housing conditions, to avoid or minimize discomfort to the animals. 
       Evaluation of Colonic Sensitivity 
       [0218]    Nociception in the animals was assessed by measuring the intracolonic pressure required to induce a behavioural response during colorectal distension (CRD) due to the inflation of a balloon introduced in the colon. This response was characterized by an elevation of the hind part of the animal body and clearly visible abdominal contraction corresponding to the severe contractions (Al Chaer, gastro 2000; Tarrerias, pain 2002; Bourdu et al., 2005). Briefly, rats were anesthetized with volatile anaesthesia (2% isoflurane), the balloon (prepared as previously described in Bourdu &amp; al, 2005) was inserted intrarectally in a minimally invasive manner to 7 cm from the anus, and the catheter was taped to the base of the tail. After 5 minutes, rats were placed in the middle of a 40×40-cm Plexiglas box and the catheter was connected to an electronic barostat apparatus (G&amp;J Electronics Inc., Toronto, Canada). Increasing pressure was continuously applied until pain behaviour was displayed or a cutoff pressure of 80 mm Hg was reached. 
       Treatment of Animals 
       [0219]    Compounds were administrated daily by intrarectal instillations. For each enema, a catheter (2-mm Fogarty catheter) was placed in the colon at 7 cm from the anus, and the animals received 500 μl of compound resuspended at optimal concentration in DMEM medium and adjusted at pH 7 by 10N NaOH if necessary for 21 days. Control animals received medium alone. Effect of the compound on visceral pain was evaluated after 2 and 3 weeks of treatment. 
       Statistics 
       [0220]    All comparisons were analyzed using the Permutation Test for two independent samples. Statistics has been calculated using the software StatXact (Cytel Inc, Cambridge, Mass., USA). Differences were considered statistically significant if the P value was &lt;0.05. 
       Conclusions 
       [0221]    The in vitro and in vivo results obtained clearly show induction of defensin expression when 5-ASA and R34, E1 and E2 are used. In particular and surprisingly E1 and E2 showed higher potency compared to 5-ASA. 
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