KITAJKA KLARA (HU)
OZSVARI BELA (HU)
NAGY LAJOS ISTVAN (HU)
FABIAN GABRIELLA (HU)
REPASI JOZSEF (HU)
SZABO ANDRAS (HU)
KERTESZ MARIUSZ (HU)
UBICHEM KUTATO KORLATOLT FELELOSSEGU TARSASAG (HU)
PUSKAS LASZLO (HU)
KITAJKA KLARA (HU)
OZSVARI BELA (HU)
NAGY LAJOS ISTVAN (HU)
FABIAN GABRIELLA (HU)
REPASI JOZSEF (HU)
SZABO ANDRAS (HU)
KERTESZ MARIUSZ (HU)
WO2007126609A1 | 2007-11-08 | |||
WO2003092617A2 | 2003-11-13 |
US3954741A | 1976-05-04 | |||
US4085139A | 1978-04-18 |
MOORE, A., WARING, C.P.: "Electrophysiological and endocrinological evidence that F-series prostaglandins function as priming pheromones in mature male Atlantic salmon (Salmo salar) PARR", J. EXP. BIOL., vol. 199, 1996, pages 2307 - 2316
BALAPURE, A.K., REXROAD, C.E., KAWADA, K. ET AL.: "Structural requirements for prostaglandin analogous interaction with the ovine corpus luteum prostaglandin F2a receptor", BIOCHEM PHARMACOL, vol. 38, 1989, pages 2375 - 2381
KARIM, S.M.M., ADAIKAN, P.G., KOTTEGODA, S.R.: "Prostaglandins and human respiratory tract smooth muscle: Structure activity relationship", ADV PROSTAGLANDIN THROMBOXANE RES., vol. 7, 1980, pages 969 - 980
"Prostaglandins and Related Substances: A Practical Approach", 1987, IRL PRESS, pages: 13 - 16
COLGAN, J. CLIN. INVEST., vol. 102, 1998, pages 1161
HARBIGE LS, YEATMAN N, AMOR S, CRAWFORD MA, BR J NUTR., vol. 74, no. 5, November 1995 (1995-11-01), pages 701 - 15
BIOCHEM SOC TRANS., vol. 25, no. 2, May 1997 (1997-05-01), pages 3425
SCHER JU, PILLINGER MH, J INVESTIG MED., vol. 57, no. 6, August 2009 (2009-08-01), pages 703 - 8
CALDER PC, BIOCHIMIE., vol. 91, no. 6, June 2009 (2009-06-01), pages 791 - 5
YACOUBIAN S, YANG R, ANNU. REV. PATHOL., vol. 3, 2008, pages 279 - 312
SAILAJA DEVI MM, DAS UN, PROSTAGLANDINS LEUKOT ESSENT FATTY ACIDS, vol. 74, no. 1, January 2006 (2006-01-01), pages 39 - 60
YUKIHIKO SUGIMOTO, SHUH NARUMIYA J, THE JOURNAL OF BIOLOGICAL CHEMISTRY, vol. 282, no. 16, 20 April 2007 (2007-04-20), pages 11613 - 11617
PROSTAGLANDIN E RECEPTORS, CLIN INVEST., vol. 109, no. 7, April 2002 (2002-04-01), pages 883 - 939
MINGHETTI L. J, NEUROPATHOL EXP. NEUROL., vol. 63, no. 9, September 2004 (2004-09-01), pages 901 - 910
GARDNEROVA M, BLANQUE R, GARDNER CR, CURR DRUG TARGETS., vol. 1, no. 4, December 2000 (2000-12-01), pages 327 - 364
LISTING J, STRANGFELD A, KARY S ET AL.: "Infections in patients with rheumatoid arthritis treated with biologic agents", ARTHRITIS RHEUM., vol. 52, no. 11, 2005, pages 3403 - 3412
MAREHBIAN J, ARRIGHI HM, HASS S, TIAN H, SANDBORN WJ.: "Adverse events associated with common therapy regimens for moderate- to-severe Crohn's disease", AM. J. GASTROENTEROL., vol. 104, no. 10, October 2009 (2009-10-01), pages 2524 - 2533
FISHER JR., C. J., AGOSTI, J. M., OPAL, S. M., LOWRY, S. F., BALK, R. A., SADOFF, J. C., ABRAHAM, E., SCHEIN, R. M., BENJAMIN, E.: "Treatment of septic shock with the tumor necrosis factor receptor :Fc fusion protein. The Soluble TNF Receptor Sepsis Study Group", N. ENGL. J. MED., vol. 334, 1996, pages 1697 - 1702
WINTER, C.A., RISLEY, E.A., NUSS, G.W.: "Carrageenan-induced edema in hind paw of the rat as an assay for antiinflammatory drugs", PROCEEDINGS OF THE SOCIETY FOR EXPERIMENTAL BIOLOGY AND MEDICINE, vol. 111, 1962, pages 544 - 547, XP008066655
Claims Use of the compounds of the general formula wherein R represents a hydrogen atom, hydroxyl group, amino group or NH-R' group, wherein R' represents an alkyl group, alkyhydroxy group, optionally substituted heteroaryl group, and the pharmaceutically acceptable salts of the compounds of the general formula (I) for reduction of inflammation. se of the compounds of the general formula wherein R represents a hydrogen atom, hydroxyl group, amino group or NH-R' group, wherein R' represents a Ci-C6 alkyl group, Ci-C6 alkyhydroxy group, Ci-C6 heteroaryl group containing 1 - 3 heteroatoms wherein the heteroatom is sulphur, nitrogen or oxygen atom and which heteroaryl group is optionally substituted with a C1-C5 alkyl group or C1-C5 cycloalkyl group or Ci-C6 heteroaryl group where the heteroatom is nitrogen atom, and the pharmaceutically acceptable salts of the compounds of the general formula (I) for reduction of inflammation. 3. Use of the compounds of the general formula (I), wherein R represent a hydrogen atom, hydroxyl group, amino group or NH-R' group, wherein R' represents a C1-C4 alkyl group, methylhydroxy, ethylhydroxy or branched or unbranched butylhydroxy group, C1-C6 heteroaryl group containing 1-3 heteroatoms, wherein the heteroatom is sulphur, nitrogen or oxygen atom selected from the following groups: thiazole, oxazole, thiadiazole and these heteroaryl groups are optionally substituted with a methyl group or cyclopropyl group or pyridinyl group, and the pharmaceutically acceptable salts of the compounds of the general formula (I) for reduction of inflammation. 4. Use according to any of claims 1 to 3 wherein the compound are selected from the following compounds: 7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l yl ] cyclopentyl ] heptanoic acid; 7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l yl] cyclopentyl] heptanamide; 7- [ (1R, 2R, 3R, 5S) -3 , 5-dihydroxy-2- [ (IE, 3S ) -3-hydroxyoct-l-en-l- yl] cyclopentyl] -N- ( 2-hydroxyethyl ) heptanamide; 7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l- yl] cyclopentyl] -N- ( l-hydroxy-2-methylpropan-2-yl ) heptanamide; 7- [ (1R, 2R, 3R, 5S) -3 , 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l- yl ] cyclopentyl ] -N- (1, 3-thiazol-2-yl) heptanamide ; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l- yl] cyclopentyl] -N- ( 5-metihyl-l , 2-oxazol-3-yl ) heptanamide; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S ) -3-hydroxyoct-l-en-l- yl] cyclopentyl] -N- ( 3-methyl-l, 2-thiazol-5-yl ) heptanamide; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l- yl] cyclopentyl] -N- (1,3, 4-thiadiazol-2-yl ) heptanamide; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S ) -3-hydroxyoct-l-en-l- yl] cyclopentyl] -N- [5- (pyridin-4-yl ) -1, 3, 4-thiadiazol-2- yl] heptanamide; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l- yl] cyclopentyl] -N- (3-methyl-l, 2-oxazol-5-yl ) heptanamide; N- ( 5-cyclopropyl-l , 3, 4-thiadiazol-2-yl ) -7- [ (2R, 3R, 5S) -3,5- dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l-yl ] cyclopentyl] heptanamide; and their pharmaceutically acceptable salts. 5. An anti-inflammatory drug composition containing compounds of the general formula (I) and/or its pharmaceutically acceptable salt and inert pharmaceutical carriers and/or excipients . 6. A process for preparation of an anti-inflammatory drug composition characterized in that compounds of the general formula (I) and/or its pharmaceutically acceptable salt and inert pharmaceutical carriers and/or excipients are mixed and brought to a galenic form. 7. A process for preparation of an anti-inflammatory drug composition containing compounds of the general formula (I) and/or its pharmaceutically acceptable salt and inert pharmaceutical carriers and/or excipients. 8. A process for reduction of inflammation characterized in that a pharmaceutically efficient amount of the compounds of the general formula (I) and/or its pharmaceutically acceptable salt is administered to the patients in need of treatment. 9. Use according to any of claims 1 to 10 for the treatment of acute inflammation, encephalomyelitis, sepsis, mucositis, rheumatoid arthritis, psoriasis, allergy, Crohn's disease, sclerosis multiplex, diabetes, osteoarthritis, obesity, eye diseases, angiogenesis of cancerous cells and progressive neurodegenerative diseases such as Alzheimer' s disease or Parkinson's disease. 10. The compounds of the general formula (I), (I) wherein R represents a hydroxyl group, amino group or NH-R' group wherein R' is an alkyl group, alkylhydroxy group, heteroaryl group, and the pharmaceutically acceptable salts of the compounds of the general formula (I) with the limitation that R is not a hydrogen atom. 11. The compounds of the general formula (I), wherein R represents a hydroxyl group, amino group or NH- R' group wherein R' represents a Ci-C6 alkyl group, Ci-C6 alkylhydroxy group, Ci~C6 heteroaryl group containing 1 - 3 heteroatoms wherein the heteroatom is sulphur, nitrogen or oxygen atom and the heteroaryl group is optionally substituted with a C1-C5 alkyl group or Ci-C5 cycloalkyl group or Ci-C6 heteroaryl group wherein the heteroatom is nitrogen atom; and the pharmaceutically acceptable salts of the compounds of the general formula (I); with the limitation that R is not a hydrogen atom. 12. The compounds of the general formula (I), wherein R represents a hydroxyl group, amino group or NH-R' , wherein R' represents a C1-C4 alkyl group, methylhydroxy, ethylhydroxy or branched or unbranched butylhydroxy group, Ci-C6 heteroaryl group containing 1-3 heteroatoms wherein the heteroatom is sulphur, nitrogen or oxygen atom selected from the following groups: thiazole, oxazole, thiadiazole or these heteroaryl groups are optionally substituted with a methyl group or cyclopropyl group or pyridinyl group, and the pharmaceutically acceptable salts of the compounds of the general formula (I) with the limitation that R is not a hydrogen atom. 13. The compounds according to claim 10 selected from the following compounds: 7- ; (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl ] heptanamide; 7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl ] - N- ( 2-hydroxyethyl ) heptanamide ; 7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl ] - N- ( l-hydroxy-2-methylpropan-2-yl ) heptanamide; 7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl ] - N- (1, 3-thiazol-2-yl ) heptanamide; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ ( IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl ] - N- (5-methyl-l, 2-oxazol-3-yl ) heptanamide; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cycopentyl ] -N - (3-methyl-l, 2-thiazol-5-yl ) heptanamide; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl ] - N- (1, 3, -thiadiazol-2-yl ) heptanamide; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl ] - N- [5- (pyridin-4-yl) -1, 3, 4-thiadiazol-2- yi] heptanamide ; 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S ) -3-hydroxyoct-l-en- 1- yi: cyclopentyl] - N- (3-methyl-l, 2-oxazol-5-yl ) heptanamide; N- ( 5-cyclopropyl -1, 3, 4-thiadiazol-2-yl) -7- [ ( 2R, 3R, 5S ) -3 , 5 - dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l-yl] cyclopentyl] heptanamide ; and their pharmaceutically acceptable salts. |
reduction of inflammation
The invention pertains to the therapeutic use of compounds of the general formula (I) and of their pharmaceutically
acceptable salts and pharmaceutical compositionscontaining compounds of the general formula (I) and their
pharmaceutically acceptable salts. Furthermore, the invention pertains to novel compounds of the general formula (I) and their pharmaceutically acceptable salts.
Prostaglandin Fi a (hereinafter: PGFi a , CAS identifier: 745-62- 0) is a representative of prostaglandins, IUPAC name: 7- [ (1R, 2R, 3R, 5S) -3 , 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l- yl ] cyclopentyl heptanoic acid. Very few data, publications have been published relating to the compound in the
literature. Presumably it is a metabolite of dihomo-γ- linolenic acid forming in the course of the metabolism of cyclooxygenase (COX) . It has been demonstrated that PGFi a and PGF 2a are the main pheromones of male Atlantic salmon with a threshold concentration of about 1CT 11 M. (Moore, A., Waring, CP. Electrophysiological and endocrinological evidence that F-series prostaglandins function as priming pheromones in mature male Atlantic salmon (Salmo salar) PARR. J. Exp. Biol. 199; 2307-2316 (1996) ) .
PGFi a is bound to the ovine corpus luteum FP receptor with only 8 % of the relative potency of PGF 2a - (Balapure, A.K., Rexroad, C.E., Kawada, K., et al. Structural requirements for prostaglandin analogous interaction with the ovine corpus luteum prostaglandin F 2Q receptor .. Biochem Pharmacol 38; 2375- 2381 (1989)) . Based on the reduced activity, it can be presumed that PGFi a does not influence the physiologic function of corpum luteum at all or only to a very slight extent.
Activity of PGFi a is only the half of the in vitro activity of PGF2a in the case of a human respiratory tract smooth muscle. (Karim, S.M.M., Adaikan, P.G., Kottegoda, S.R. Prostaglandins and human respiratory tract smooth muscle: Structure activity relationship. Adv Prostaglandin Thromboxane Res. 7 969-980 (1980) ) . Based on these results, the blood pressure
influencing effect of PGFi a is probably also very slight.
There are only presumptions regarding in vivo existence of prostaglandin Fi a , few publications have been published in connection with PGFi a , its literature is poor, neither its exact pharmacological role nor its mechanism of action has been clarified. Similarly to prostaglandin F ia , 6-keto- prostaglandin F la also belongs to the family of
prostaglandins . In opposition to PGFi a , there is a great number of data known of 6-keto-prostaglandin F ia .
6-keto-prostaglandin Fi a is a physiologically active and stable metabolite of prostacyclin. It can be found in almost every mammal tissue, it has a vasodilatory effect and inhibits platelet aggregation. Based on the literature, the retarded or expanded increase of the 6-keto-prostaglandin Fi a level has caused septic shock in animals suffering from fecal
peritonithis or appendicitis (cecal ligation) . The plasma concentration of 6-keto-prostaglandin Fi a was found to be high in patients suffering from epidemic haemorrhagic fever, acute obstructive suppurative cholangitis and gynaecological
cancerous diseases. Furthermore, it showed significant
correlation with the plasma concentration of HDL (high density lipoprotein) . Its sodium salt is also used for the treatment of primary pulmonary hypertonia. (PMID 1976492, 2298410,
2379443, 2111556) . 6-keto-prostaglandin Fi a is a stable compound forming during the hydrolysis of PGI2 in mammals, it is often used for detection of PGI 2 in blood and urine ( Prostaglandins and
Related Substances: A Practical Approach ; C. Benedetto, R. G. McDonald-Gibson, S. Nigam, and T. F. Slater, Eds. ; IRL Press : Oxford, 1987, pp. 13-16) .
Recently it has been demonstrated that 6-keto-prostaglandin Fie is an effective chloride secretagogue, which is secreted by the epithelial cells of the bowel in hypoxia (Colgan et . al., J. Clin. Invest ., 102 : 1161 (1998)) .
PGFic and 6-keto-prostaglandin Fi a are compounds different in their structure, metabolism and mechanism of action. It is known that 6-keto-prostaglandin Fi a is a marker of the inflammation process, however, there are no data showing any connection between the inflammation and PGFi a .
The object of our invention is to determine the
pharmacological role of PGFi a as exactly as possible and to examine any possible use of PGFi a -nak and its analogues in medicine .
In 1995 Harbige LS et al. showed that lipids rich in orally administered GLA (gamma linolenic acid) inhibited experimental autoimmune encephalomyelitis (hereinafter: EAE) induced by myelin homogenatein Lewis rats (Prevention of experimental autoimmune encephalomyelitis in Lewis rats by a novel fungal source of gamma-linolenic acid; Harbige LS, Yeatman N, Amor S, Crawford MA. Br J Nutr. 1995 Nov; 74 (5) : 701-15. ) .
In 1997, the same research group recognized during their further experiments, that omega-6 lipids mitigate the symptoms of experimental autoimmune encephalomyelitis (hereinafter: EAE) (Biochem Soc Trans. 1997 May; 25 (2):342S; Protective mechanisms by omega-6 lipids in experimental autoimmune encephalomyelitis are associated with cytokines and
eicosanoids; Harbige LS, Layward L, Morris M, Amor S.). During their experiments it was found that the TGFi b i and PGE 2 levels increased on the effect of orally administered lipids rich in GLA (gamma linolenic acid) (e.g. borage oil) compared with the animals obtaining normal food. With reference to previous results, they stated that the dihomo-g-linolenic acid (DHGLA) and arachidonic acid (AA) levels were significantly higher in the lineal cells of the animals treated that way. However, the role of PGFi a was not examined in these experiments.
The inventors of the present invention presume that the EAE (experimental autoimmune encephalomyelitis) inhibition is not a consequence of the increased PGE 2 level. It is true that the anti-inflammatory effect of PGE 2 has been demonstrated in several cases, however, several studies describe the
inflammation-inducing effect of PGE 2 as well, so this
prostaglandin has a contradictory role (The anti-inflammatory effects of prostaglandins. Scher JU, Pillinger MH. J Investig Med. 2009 Aug; 57 ( 6) : 703-8 ) . We also presume that another active compound should be present in this process. PGE 2 forms from GLA through dihomo-g-linolenic acid (DHGLA) in a 4-step synthesis, however, probably PGFi a also forms from DHGLA so this mediator may be a key of the anti-inflammatory effect. Prostaglandin F la is a lipid mediator compound, a
representative of prostanoids. Prostanoids are compounds occurring in the whole organism with various physiological and pathological functions e.g. in the central nervous system, circulatory system, gastrointestinal tract, genitalias, endocrine system, hormonal system, during respiration and in the immune system.
Prostaglandins have great importance in the clinical practice mainly as cyclooxygenase inhibiting non-steroidal anti ¬ inflammatory drugs the effect of which is based on the
inhibition of prostaglandin synthesis among others.
There are several publications known describing the anti ¬ inflammatory effect of various prostaglandins, but there is no reference to the effect of PGFi a on inflammation in the studies. (For example: The anti-inflammatory effects of prostaglandins. Scher JU, Pillinger MH. J Investig Med. 2009 Aug; 57 ( 6) : 703-8. ; Polyunsaturated fatty acids and inflammatory processes: New twists in an old tale. Calder PC. Biochimie. 2009 Jun; 91 (6) : 791-5. ; Anti-inflammatory and proresolving lipid mediators, erhan CN, Yacoubian S, Yang R. Annu. Rev. Pathol. 2008; 3:279-312.) .
The various inflammatory bowel diseases (herinafter: IBD) are among the frequent diseases nowadays. Such diseases can be found worldwide in almost every country. There are two basic forms of the inflammatory bowel diseases. One of them is colitis ulcerosa, which attacks only the colon and the
inflammatory reaction is localized to the mucous membrane; the other is the Crohn' s disease where inflammation is present in all layers of the intestinal wall from mucosa to serosa
(transmural) . This inflammatory bowel disease was described by the American physician Burrill Crohn in 1932 and it was named after him. Distinction between the diseases can be made and the exact diagnosis is possible on the basis of clinical, endoscopic and histological characteristics.
It has been shown that oxidative stress is a key factor in the formation of intestinal epithelium injury. It is
characteristic of oxidative stress that reactive oxygen and nitrogen species (hereinafter: RONS, reactive oxygen and nitrogen species) form in the organism. Inflammation-inducing factors increase the formation of RONS of which it has been shown that those aggravate the course of IBD themselves and due to their interaction and take part in the formation of epithelial and vascular injuries in the colon. Inflammation- generating cytokines such as TNF-a, IL-6 and nitrogen monoxide (NO) generated by inducible nitric oxide synthase
(hereinafter: iNOS, inducible Nitric Oxide Synthase) can be considered as such inflammation-inducing transmitting
substances. It is known that peroxynitrite coming from the interaction of nitrogen monoxide and superoxide free radical is highly toxic whith a clear pathologic role during formation of IBD.
The inflammatory processes can be induced directly by bacteria (Gram negative or positive) causing infections or their molecular products such as the endotoxin of Escherichia coli (lipopolysaccharide-LPS) . The latter processes are called systemic inflammatory response syndrome (hereinafter: SIRS) . During the inflammatory processes the neutrophil cells and endothelial cells start to produce a considerable amount of inflammation-inducing cytokines (IL-1, IL-6, TNF-a) . The first aim of the drug treatment used for the inflammatory bowel diseases is to mitigate the symptoms of the disease, to stop the inflammatory process, to heal any complication
(abscess, fistula) and to maintain the recovered state
(remission) . The way of the drug therapy may be different according to the seriousness of the disease and the place of its appearance. There are five main groups of the drugs used in the treatment of IBD: salicylic acid derivatives, (5-ASA, SASP) , corticosteroids, immune suppressors, antibiotics, biological agents (infliximab) .
The TNBS ( 2 , 4 , 6-trinitrobenzol-szulfonsav) model reproduces well many macroscopic, tissue and immunological change
characteristic of patient suffering from Crohn's disease, so it is a widespread and accepted model in the study of IBD.
According to certain results (Effect of prostaglandins against alloxan-induced diabetes mellitus. Sailaja Devi MM, Das UN. Prostaglandins Leukot Essent Fatty Acids. 2006 Jan; 74(1) :39- 60.), PGEi, PGE 2 , PGFi a and PGF 3c< eliminate diabetes mellitus chemically induced in experimental animals and mitigate the oxidative stress occurring during diabetes mellitus. The publication refers to a possible role of PGFi « in indication of diabetes mellitus; however it does not mention any antiinflammatory effect of the compound.
With the knowledge of the above literature, we started to examine the role of PGFi a and its derivatives in various inflammatory processes.
The receptors binding the PGF 2a compounds as substrates constitute a group of prostaglandin receptors. This group of receptors includes PTGFR (prostaglandin F receptor) and prostaglandin D2 receptor (or GPR44, G Protein-Coupled
Receptor) . GPR44 directs pre-inflammation chemotaxis of eosinophil, basophil and Th 2 lymphocytes forming during the allergic reaction.
The PGF 2a -FP receptor signal can direct the function and inflammatory processes of the endothelial cell.
The inventors of the present invention presume that the compounds of the general formula (I) exert their antiinflammatory effect bound to prostaglandin receptors which prevent the effect of inflammatory mediators (e.g. PGE 2 , PGD 2 ) as an antagonist, competitive agonist or reverse antagonist. Of the prostaglandin receptors, the 4-type receptor of PGE 2
(PTGER 4 ) , PTGFR and PD 2 should be emphasised. (A comprehensive review on PTG E receptors: Yukihiko Sugimoto and Shuh Narumiya J: The Journal Of Biological Chemistry Vol. 282, NO. 16, pp. 11613-11617, April 20, 2007; Prostaglandin E Receptors, Clin Invest. 2002 Apr ; 109 ( 7 ) : 883-939 ; The prostaglandin receptor
EP4 suppresses colitis, mucosal damage and CD4 cell activation in the gut, Kabashima K, Saji T, Murata T, Nagamachi M,
Matsuoka T, Segi E, Tsuboi K, Sugimoto Y, Kobayashi T, Miyachi Y, Ichikawa A, Narumiya S.)
More and more experimental results prove that the inflammation of the nervous cells plays part not only in the classical diseases involving neurotis as sclerosis multiplex or
amyotrophic lateral sclerosis, but also in the pathophysiology of progressive neurodegenerative diseases as Alzheimer' s disease and Parkinson's disease. (Cyclooxygenase-2 (COX-2) in inflammatory and degenerative brain diseases. Minghetti L. J Neuropathol Exp. Neurol. 2004 Sep; 63 ( 9 ) : 901-910 ) . A considerable part of inflammatory rheumatoid diseases are polysystemic autoimmune diseases involving several organ systems. Inflammation of joints (arthritis) is characteristic of almost all of them. In many diseases arthritis and the inflammation of spine (spondylarthritis) are dominant also accompanied by autoimmune symptoms. Rheumatoid arthritis is chronic arthritis involving several joints at the same time, which is treated with steroids and non-steroid cyclooxygenase inhibitors and by biological therapy.
Rheumatoid arthritis is characterized by a high level of destructivity while seronegative arthritis is characterized by chronic inflammation (synovitis and enthesitis) . The cytokines taking a key role in these processes are the tumour necrosis factor alpha (TNF- ) , interleukin 1 (IL 1) and RANK ligand (RANKL) . TNF-a has a central role in both processes. It causes inflammation by inducing IL 1 and several other cascades (e.g. COX 2 enzyme) and triggers the destructive processes through RANKL and metalloproteinase expression. Due to the central role of TNF-a, the greatest therapeutic effect can be expected from blocking that cytokine. (The use of TNF family ligands and receptors and agents which modify their interaction as therapeutic agents. Gardnerova M, Blanque R, Gardner CR. Curr Drug Targets. 2000 Dec; 1 (4 ) : 327-364.) .
The essence of the biological therapy of arthritis is the use of the antagonists of the large molecule protein components taking role in synovitis, as a biological response; or these components are inhibited using monoclonal antibodies (antibody against TNF-a first of all) during the treatment. These physiological proteins are usually produced by gene
technology . Good recovery results, a significant anti-inflammatory effect can be achieved by the anti-TNF-a antibody therapy, however, it is very expensive, requires intravenous administration and the therapy can not be repeated many times. The same therapy is used in the treatment of Crohn's disease as well.
The bowel inflammation reducing effect of the compounds according to the invention reaches the extent of the effect of the anti-TNF-a antibody; their advantages are that those can be produced more economically at a large scale and it can be administered more easily. Repeated application of anti-TNF-a antibody increases the frequency of bacterial and viral infections as well (Listing J, Strangfeld A, Kary S, et al.: Infections in patients with rheumatoid arthritis treated with biologic agents. Arthritis Rheum. 2005; 52 ( 11 ): 3403-3412 ; and Marehbian J, Arrighi HM, Hass S, Tian H, Sandborn WJ. : Adverse events associated with common therapy regimens for moderate- to-severe Crohn's disease. Am. J. Gastroenterol. 2009 Oct.; 104 (10) : 2524-2533. ) .
In sepsis, the anti-TNF-α antibody was not effective in many cases, moreover, it aggravated the disease during clinical trials (Fisher Jr., C. J., Agosti, J. M., Opal, S. M., Lowry, S. F. , Balk, R. A., Sadoff, J. C, Abraham, E . , Schein, R. M . , Benjamin, E. (1996) Treatment of septic shock with the tumor necrosis factor receptor : Fc fusion protein. The Soluble TNF Receptor Sepsis Study Group. N. Engl. J. Med. 334,1697-1702.). From these results it can be seen that several other
inflammatory intermediates also take role in sepsis.
Furthermore, the compounds according to the invention are also effective in the treatment of arthritis of patients suffering from psoriasis and in mitigation of inflammations occurring during asthma and allergy.
The object of our invention is to determine the
pharmacological role of PGFi a as exactly as possible and to examine the possible therapeutic use of PGFi a and its analogous compounds. The further object of the invention is examine the effect of PGFi a molecule and its derivatives and to develop a new anti-inflammatory composition.
The above object is achieved with the compounds having the general formula (I),
(I) where
R represents a hydrogen atom, hydroxyl group, amino group or NH-R' group, where
R' is an alkyl group, alkylhydroxy group, optionally
substituted heteroraryl group,
and with the pharmaceutically acceptable salts of the compounds of the general formula (I) , which can be used for reduction of inflammation in therapy.
More specifically, the object of the invention is the use of compounds of the general formula (I),
where R represents a hydrogen atom, hydroxyl group, amino group or NH-R' group, where
R' represents a C1-C6 alkyl group, C1-C6 alkylhydroxy group, C1-C6 heteroaryl group containing 1-3 heteroatoms wherein the heteroatom is sulphur, nitrogen or oxygen atom and which heteroaryl group is optionally substituted with a C1-C5 alkyl group or C1-C5 cycloalkyl group or C1-C6 heteroaryl group wherein the heteroatom is a nitrogen atom,
and the pharmaceutically acceptable salts of the compounds of the general formula (I) for reduction of inflammation. A further object of the invention is the use of compounds of the general formula (I),
where R represents a hydrogen atom, hydroxyl group, amino group or NH-R' , wherein
R' represents a C1-C4 alkyl group, methylhydroxy, ethylhydroxy or branched or unbranched butylhydroxy group, C1-C6 heteroaryl group containing 1-3 heteroatoms wherein the heteroatom is sulphur, nitrogen or oxygen atom selected from the following groups: thiazole, oxazole, thiadiazole and these heteroaryl groups optionally substituted with a methyl group or
cyclopropyl group or pyridinyl group,
and the pharmaceutically acceptable salts of the compounds o the general formula (I) for reduction of inflammation.
Preferred representatives of the compounds of the general formula (I) suitable for reduction of inflammation are as follows :
7- ; (1R, 2R, 3R, 5S) -3 , 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl] heptanoic acid;
7- ; (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl] heptanamide;
7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S ) -3-hydroxyoct-l-en- 1- yi] cyclopentyl ] -N- ( 2-hydroxyethyl ) heptanamide ;
7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl] -N- ( l-hydroxy-2-methylpropan-2-yl ) heptanamide;
7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi] cyclopentyl] -N- (1, 3-thiazol-2-yl ) heptanamide;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S ) -3-hydroxyoct-l-en- 1- yi: cyclopentyl] -N- (5-methyl-l, 2-oxazol-3-yl ) heptanamide;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi: cyclopentyl ] -N- ( 3-methyl-1 , 2-thiazol-5-yl) heptanamide ;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi: cyclopentyl] -N- (1,3, 4-thiadiazol-2-yl ) heptanamide;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en- 1- yi cyclopentyl] -N- [5- (pyridin-4-yl ) -1, 3, 4-thiadiazol-2- yi heptanamide ;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S ) -3-hydroxyoct-l-en- 1- yi cyclopentyl] -N- (3-methyl-l, 2-oxazol-5-yl ) heptanamide; N- (5-cyclopropyl-l, 3, 4-thiadiazol-2-yl) -7- [ (2R, 3R, 5S) -3,5- dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l- yl] cyclopentyl] heptanamide;
and their pharmaceutically acceptable salts.
Brief description of the figures:
in Fig. 1 the effect of PGFla on the size of carrageenin- induced pedal oedema is illustrated;
in Fig. 2 the effect of the compounds according to the
invention on experimental autoimmune encephalomyelitis (EAE) is illustrated;
in Figs 3, 4 and 5 the behaviour of the compounds according to the invention in the lipopolysaccharide (LPS) (coming from Escherichia coli bacterium) induced sepsis model is shown;
Figs. 6 - 13 show the effect of the compounds according to the invention on mucositis induced by TNBS ( trinitrobenzene sulphonic acid) .
Interpretation of the definitions and terms used in the description is given as follows:
The term „Ci- e" means a group with a branched or unbranched carbon atom chain containing 1, 2, 3, 4, 5, or 6 carbon atoms.
The term „alkyl" or, in other groups, the initial letters „alk" (e.g. alkoxy, alkanoil, alkenyl, alkinyl group) mean a branched or unbranched carbon atom chain or their
combinations. Examples of alkyl groups are methyl, ethyl, propyl, isopropyl-, butyl-, sec-butyl-, tert . -butyl-, pentyl-, hexyl-, and heptyl group or similar groups.
The term „cycloalkyl" means optionally substituted saturated cyclic hydrocarbons. Any differences are defined separately. Examples of cycloalkyl groups with 3-7 carbon atoms are cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl groups.
The „heteroaryl and saturated heterocyclic ring systems" contain one or more heteroatoms. Examples of heteroatoms are oxygen, sulphur and nitrogen atom which may occur combined in the ring substituting the carbon atoms of the ring. Examples of the heteroaryl groups are pyridinyl, quinolinyl,
isoquinolinyl, pyridazinyl, pyrimidinyl, pyrazinyl,
quinoxalinyl, furyl, benzofuryl, debenzofuryl, thienyl, benzothienyl, pyrrolyl, indolyl, pyrazolyl, indazolyl,
oxazolyl, benzoxazolyl , isoxazolyl, thiazolyl, benzothiazolyl , isothiazolyl, imidazolyl, benzimidazolyl, oxadiazolyl,
thiadiazolyl, triazolyl and tetrazolyl. Examples of the saturated heterocyclic groups are azetidinyl, pyrrolidinyl , piperidinyl, piperazinyl, morpholinyl, tetrahydrofuranyl- , imidazolinyl- , pyrrolidin-2-one, piperidin-2-one es
thiomorpholinyl .
The term ^ pharmaceutically (pharmacologically) acceptable salts" are prepared by means of pharmaceutically
(pharmacologically) acceptable, non-toxic bases or acids. If the compound according to the present invention is acidic, the suitable salt can be prepared using a pharmaceutically
acceptable, non-toxic (inorganic or organic base). Of the salts formed with bases, especially important are the salts formed with alkali metals (e.g. sodium or potassium), alkali earth metals (e.g. calcium or magnesium), and ammonia or organic amines. The latter bases may contain other
substituents (e.g. hydroxyl or amino group) which may also influence e.g. solubility or handleability of the products. Both organic and inorganic salts can be used for the
preparation of acid addition salts. Examples of suitable organic acids are hydrochloric acid, sulphuric acid and phosphoric acid. Examples of suitable monovalent organic acids are formic acid, acetic acid, trifluoro acetic acid, propionic acid, various butyric acids, valeric acids and hexanoic acids. Examples of bivalent organic acids (dicarbonic acids) are oxalic acids, malonic acid, maleic acid, fumaric acid and succinic acid. Other organic acids as hydroxy acids (e.g.
citric acid, tartaric acid) or aromatic carboxylic acids (e.g. benzoic acid, salicylic acid) and aliphatic and aromatic sulphonic acids (e.g. methane sulphonic acid, p-toluene- sulphonic acid) can also be used. The more valuable group of the acid addition salts is the group in which the acid
component itself has no therapeutic effect and no adverse influence on the effect of the active ingredient in the dose applied. These are the pharmaceutically acceptable acid addition salts. Other acid addition salts according to the present invention - which do not belong to the group of pharmaceutically acceptable salts - may be preferred in isolation or purification of the desired compounds.
A further object of the invention is an anti-inflammatory pharmaceutical composition containing compounds of the general formula (I) and/or its pharmaceutically suitable salt and inert therapeutic carriers and/or excipients.
The compounds of the general formula (I) and their
pharmaceutically acceptable salts and the pharmaceutical composition produced from them can be administered in any usual way, e.g. orally, parenterally (including subcutaneous, intramuscular and intravenous ways of administration) ,
buccally, sublingually, nasally, rectally or transdermally . Dosage units are made from the pharmaceutical compositions using routine pharmaceutical procedures.
Liquid or solid dosage forms can be made from orally active compounds of the general formula (I) and their
pharmaceutically acceptable salts. For example, syrups, suspensions, emulsions, tablets, capsules or lozenges can be produced .
Where a liquid, e.g. a suspension solution form is made from the compounds of the general formula (I) and their
pharmaceutically acceptable salts, it will contain the
compounds of the general formula (I) and their physiologically acceptable salts in a suitable liquid carrier or carriers. Aqueous solvents (e.g. water, ethanol or glycerol) or nonaqueous solvents (e.g. polyethylene glycol or any oil) can be used. The preparation may also contain suspending agents, preservatives, flavourings and colorants.
Where the solid preparation is a tablet, it can be produced using any suitable carrier usually used in pharmaceutical production. Examples of the solid carriers are lactose, suitable silicates, saccharose, talc, gelatine, agar, pectin, gum arabic, magnesium stearate and stearic acid, etc.
Optionally a coat may be applied on the tablet by means of any standard aqueous or non-aqueous technique.
Tablets can be made from the composition according to the invention by pressing or moulding optionally using one or more agents promoting absorption or adjuvants. The tablet can be made using e.g. a suitable press; the active ingredient can be pressed in the form of a powder or granules, optionally with binding agents, lubricants, inert diluting agents, surfactants or dispersing agents.
Where the solid preparation is a capsule it can be produced using any routine capsule filling method. For example, pellets may be made from the active ingredient with a standard carrier and filled into hard gelatine capsules. An alternative way is to make a dispersion or suspension from the active ingredient with a suitable pharmaceutical carrier and filled into soft gelatine capsules. Suitable pharmaceutical carriers are e.g. water-dispersable gums, cellulose, silicates or oils.
Typical forms of the parenteral compositionsare the solutions or suspensions containing the compounds of the general formula (I) and their pharmaceutically acceptable salts in sterile aqueous carriers or parenterally administrable non-aqueous carriers e.g. polyethylene glycol, polyvinyl pyrrolidone, lecithin, peanut oil or sesame oil. Alternatively, the
solution can be lyophilized and reconstituted with a suitable solvent just before administration.
Forms of the compositions according to the invention suitable for nasal administration contain the compounds of the general formula (I) and their pharmaceutically acceptable salts in the form of an aerosol, drop, gel or powder. The aerosols
according to the invention contain the compounds of the general formula (I) and their pharmaceutically acceptable salts usually in a solution or finely dispersed suspension, physiologically acceptable aqueous or non-aqueous solvent. The sterile aerosols can be in a sealed container containing a single dose or multiple doses where dosage or refilling is ensured and usually equipped with a sprayer. Alternatively, the sealed container may be suitable for dispensing units e.g. a single-dose inhaler or an aerosol dispenser equipped with a dosing valve which can be discarded after emptying the
container. Where dosing is provided by using an aerosol dispenser, a propellant e.g. compressed gas (e.g. compressed air) or organic propellant (e.g. chlorinated fluorinated hydrocarbons) is used. Dosing of the aerosol can be provided by using a spraying pump as well.
The compositions according to the invention containing
compounds of the general formula (I) can also be administered buccally or sublingually; e.g. in the form of a tablet, lozeng or pastille; those contain the active ingredient formulated with a carrier (e.g. sugar and gum arabic, gum tragacanth or gelatine, glycerol, etc.).
The compositions according to the invention containing
compounds of the general formula (I) or their pharmaceutically acceptable salts can also be administered rectally. Usually suppositories are made which contain the active ingredient in some suppository base e.g. cocoa butter or other known
carrier. The suppositories are made in the usual way by mixing the components, softening or melting the mixture and casting into a mould and cooling.
The composition according to the invention containing
compounds of the general formula (I) or their pharmaceutically acceptable salts is also suitable for transdermal
administration in the form of e.g. an ointment, gel or
plaster .
Preferably, the compositions according to the present
invention containing compounds of the general formula (I) or their pharmaceutically acceptable salts are brought to a dosage unit form (e.g. tablet, capsule or ampoule).
The composition according to the present invention containing compounds of the general formula (I) or their pharmaceutically acceptable salts is prepared with an active ingredient content of 0.1-200 mg per oral dosage unit.
The composition according to the present invention containing compounds of the general formula (I) or their pharmaceutically acceptable salts is prepared with an active ingredient content of 0.1-200 mg per oral dosage unit.
Also the object of the invention is a procedure for producing an anti-inflammatory pharmaceutical composition wherein the compounds of the general formula (I) or their pharmaceutically acceptable salts are mixed with inert pharmaceutical carriers and/or excipients and brought to a galenic form. A further object of the invention is a process for manufacturing an anti-inflammatory pharmaceutical composition containing the compounds of the general formula (I) and/or their
pharmaceutically acceptable salts and inert pharmaceutical carriers and/or excipients.
The invention also pertains to a process inducing reduction of inflammation wherein a pharmacologically efficient amount of the compounds of the general formula (I) or their
pharmaceutically acceptable salts are administered to the patient in need of treatment.
The use according to the invention also pertains to the treatment of acute inflammation, encephalomyelitis, sepsis, mucositis, rheumatoid arthritis, psoriasis, allergy, Crohn's disease, sclerosis multiplex, diabetes, osteoarthritis, obesity, eye diseases, angiogenesis of cancerous cells and progressive neurodegenerative diseases such as Alzheimer' s disease or Parkinson's disease.
Furthermore, the invention pertains to the novel compounds of the general formula (I),
wherein
R represents a hydroxyl group, amino group or NH-R' group wherein
R' is an alkyl group, alkylhydroxy group, heteroaryl group, and the pharmaceutically acceptable salts of the compounds of the general formula (I) with the limitation that R may not be a hydrogen atom.
A further object of the invention is the novel compounds of the general formula (I),
wherein R represents a hydroxyl group, amino group or NH-R' group wherein R' represents a C1-C6 alkyl group, Ci-C 5 alkylhydroxy group, Ci-C 6 heteroaryl group containing 1 - 3 heteroatoms wherein the heteroatom is sulphur, nitrogen or oxygen atom and the heteroaryl group is optionally substituted with a C1-C5 alkyl group or C1-C5 cycloalkyl group or Ci-C 6 heteroaryl group wherein the heteroatom is a nitrogen atom;
and the pharmaceutically acceptable salts of the compounds of the general formula (I);
with the limitation that R may not be a hydrogen atom.
Also the object of the invention is the novel compounds of the general formula (I)
wherein R represents a hydroxyl group, amino group or NH-R' , wherein
R' represents a C1-C4 alkyl group, methylhydroxy, ethylhydroxy or branched or unbranched butylhydroxy group, C1-C6 heteroaryl group containing 1 - 3 heteroatoms wherein the heteroatom is sulphur, nitrogen or oxygen atom selected from the following groups: thiazole, oxazole, thiadiazole or these heteroaryl groups optionally substituted with a methyl group or
cyclopropyl group or pyridinyl group,
and the pharmaceutically acceptable salts of the compounds of the general formula (I)
with the limitation that R may not be a hydrogen atom. Preferred representatives of the compounds of the general formula (I) according to the present invention are as follows:
7- t (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l -en- 1- yi] cyclopentyl] heptanamide;
7- [ (1R, 2R, 3R, 5S) -3 , 5-dihydroxy-2- [ (IE, 3S ) -3-hydroxyoct-l -en- 1- yi] cyclopentyl ] - N- (2-hydroxyethyl) heptanamide;
7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S ) -3-hydroxyoct-l -en- 1- yi] cyclopentyl ] - N- ( l-hydroxy-2-methylpropan-2-yl ) heptanamide;
7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l -en- 1- yi] cyclopentyl ] - N- (1, 3-thiazol-2-yl) heptanamide;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en -1- yi: cyclopentyl] - N- (5-methyl-l, 2-oxazol-3-yl ) heptanamide;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en -1- yi: cycopentyl ] -N - (3-methyl-l, 2-thiazol-5-yl ) heptanamide;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ ( IE, 3S) -3-hydroxyoct-l-en -1- yi: cyclopentyl ] - N- (1, 3, 4-thiadiazol-2-yl ) heptanamide;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ ( IE, 3S) -3-hydroxyoct-l-en -1- yi: cyclopentyl ] - N- [5- (pyridin-4-yl) -1, 3, 4-thiadiazol-2- yi: heptanamide ;
7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en -1- yi " cyclopentyl ] - N- (3-methyl-l , 2-oxazol-5-yl ) heptanamide;
N- ( 5-cyclopropyl -1, 3, -thiadiazol-2-yl) -7- [ (2R, 3R, 5S) -3, 5- dihydroxy-2- [ (IE, 3S) -3-hydroxyoct-l-en-l-yl ] cyclopentyl] heptanamide;
and their pharmaceutically acceptable salts.
Preparation of the compounds according to the invention is illustrated in Flow Chart 1.
Based on that, 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- ( (E) - (S) -3- hydroxyoct-l-en-yl) -cyclopentyl ] heptanoic acid was reacted in ethyl acetate with an equimolar quantity of an amine
derivative in the presence of 1.1 equivalent connecting agent 4- (4, 6-dimethoxy-l, 3, 5-triazin-2-yl ) -4-methylmorpholinium chloride (hereinafter: DMTMM) . The crude products were
purified by means of HPLC in every case. The purified products were analysed by ESI-MS.
Biological effect of the compounds according to the invention were studied though the tests included in the examples. Our results show that the compounds of the general formula (I) are strong anti-inflammatory agents in the animal model of Crohn' s disease, in the arthritis models and the sepsis model.
Our invention is described in detail through the
implementation examples below without limiting the object of our invention to them.
Examples
Preparative examples : Example 1 Synthesis of 7- [ (1R, 2R, 3R, 5S) -3 , 5-dihydroxy-2- [ (IE , 3S) -3- hydroxyoct-l-en-l-yl] cyclopentyl] heptanamide (F02)
10 mg (0.028mmol) of 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- ( (E) - (S) -3- hydroxyoct-l-en-yl ) -cyclopentyl ] heptanoic acid was dissolved in 2 ml of tetrahydrofuran saturated with ammonia and 9 mg (0.031 mmol) of DMTM was added. It was stirred overnight at room temperature. The reaction mixture was poured on 10 ml of water and extracted with 3 x 4ml of ethyl acetate. It was dried with water-free sodium sulphate and evaporated. 15 mg of colourless oil obtained was purified by means of preparative HPLC. After lyophilization 1.5 mg of white solid was obtained.
Example 2
Synthesis of 7- [ (2R, 3R, 5S) -3 , 5-dihydroxy-2- [ (IE , 3S) -3- hydroxyoct-l-en-l-yl] cyclopentyl] -N- (3-methyl-l , 2-thiazol-5- y1) he tanamide (FO7)
To 10 ml of ethyl acetate, under argon, 60 mg (0.168 mmol) of 7- [ (2R, 3R, 5S) -3, 5-dihydroxy-2- ( (E) - (S) -3-hydroxyoct-l-en-l- yl ) -cyclopentyl ] heptanoic acid, 25.3 mg of (0.168 mmol) of 5- amino-3-methylisothiazol , 0.024 ml (0.168 mmol) of triethyl amine and 52 mg (0.186 mmol) of DMTMM were added. The mixture was stirred overnight at room temperature then washed with 2x3 ml of water. The organic phase was dried with water-free sodium sulphate and evaporated. From 98 mg crude product, 18 mg of product was obtained after purification by HPLC.
Preferred representatives of the compounds of the general formula (I) are listed in Table 1. The compounds were prepared in a way similar to the procedure described in Example 2 using the corresponding amine compound. The first column ("Symbol") includes the names of the
compounds .
The compounds are referred in the pharmacological examples based on this nomenclature. The compound with the symbol FOl refers to the compound PGFi a (7- [ (1R, 2R, 3R, 5S) -3, 5-dihydroxy- 2- [ (IE, 3S) -3-hydroxyoct-l-en-l-yl] cyclopentyl] heptanoic acid) known according to the state of the art.
Table 1: Some preferred compounds according to the invention
The following instrument was used for the MS measurements: Instrument: Perkin-Elmer Sciex Instruments API 2000 Triple Quadropole LC/MS/MS Mass spectrometer
Source: Turbo IonSpray
Source temperature: 300.0°C
Polarity: positive and negative
Pharmacological experiments : Example 3
Effect on acute inflammation
Fifty bred mice were randomly distributed into five groups. The groups containing ten mice each are as follows: control group (obtained only the carrier), F01 group (PGFi a 0.2 mg/kg) , F01 group (PGF ia 0.5 mg/kg) , dexamethasone group
(reference active substance, 0.7 mg/kg) and PGE 2 group (0.5 mg/kg). The same quantity (0.1 ml) of physiological salt solution was administered to the mice through the saphenous vein. Inflammation was induced at the same time by means of a subplantar injection prepared from 30 μΐ 2 % suspension of carrageenan ( Sigma-Aldrich) with 0.9% physiological salt solution administered in the right hind paw aponeurosis of the mouse. (Winter, C.A., Risley, E.A., Nuss, G.W., 1962.
Carrageenan-induced edema in hind paw of the rat as an assay for antiinflammatory drugs. Proceedings of the Society for Experimental Biology and Medicine 111, 544-547.).
The leg volume was measured before treatment and six hours after treatment. Leg oedema was induced by the carrageenan injection in the case of every animal.
As it is shown in Fig. 1, the size of leg oedema induced carrageenan was reduced by PGFi a in a way depending on th concentration. Intravenous administration of 0.5 mg/kg PGFi„ produced the same reduction of inflammation as did
dexamethasone in 0.7mg/kg concentration. The prostaglandin PGE 2 had no anti-inflammatory effect.
Example 4
Effect on Experimental Autoimmun Encephalomyelitis (EAE)
Induction and measurement of EAE was conducted as follows:
C57B1/6 bred mice were immunized with 200 g of MOG35-55 peptide (prepared by the Department of Medicinal Chemistry, University of Szeged) by means of 2 mg of Freund' s complete adjuvant substituted with heat-killed H37RA mycobacterium tuberculosis bacterium (SIGMA) . On day 0 and day 2 mice were treated with 400 ng and 200 ng of pertussis toxin (List
Biological Laboratories, Campbell, CA) intravenously and the clinical levels of the forming disease were recorded (level 1: weak tail, level 2: weakness in hind legs, level 3: paralysis in hind legs, level 4: total paralysis in hind legs, level 5: total paralysis in hind legs and paralysis of bottom or incontinency, level 6: dying or death) . Feed and water were placed on the bottom of the mouse cage. The moribund mice were killed by euthanasia. All the animal experiments were
conducted following the national ethical guidelines, with the knowledge of the ethical rules.
Treatment of mice was started on day 7. 100 ul volume (98 % physiological salt solution, 2 % acetone) was administered by intravenous injection once a day for 5 days. The control mice obtained only the carrier buffer, the F01 group obtained buffer containing PGFi a with a dose of 0.3 mg/kg. 15 animals were treated in both groups. It can be seen from Fig. 2 that the disease was much milder in the animals treated with PGFi a compared with the control group. Moreover, only 3 animals were symptom-free in the control group on day 14 while there were 12 symptom-free animals in the group treated with PGFic,. Example 5
Lipoploysaccharide (LPS) (from Escherichia coli bacterium) induced sepsis model in the case of treatment with the
compound F3
8 week Balb/c female mice were used in the experiment. The average body weight of the animals was 25 g. Three groups were used in the experiment: Group 1: control group obtaining only the carrier (98% physiological salt solution, 2% acetone); Group 2: group obtaining 250 pg/kg PGFla; Group 3: group obtaining 250 pg/kg F03 compound. Dosing was repeated three times .
One hour before the treatment each animal obtained LPS (LPS Escherichia coli 0111:B4; Sigma L3012-10mg; 058K4006)
dissolved in 100 μΐ of physiological salt solution, once, intraperitoneally . LPS was used in a concentration of 3 mg/kg (LPS 1.72 mg/ml) . After the treatment with LPS the animals were randomized. 10 animals were in each of the control and the treated groups.
Each animal obtained 250 pg/kg F01. 1, 4 and 8 hours after the treatment with LPS the treated groups obtained 100 μΐ PGFi a or F03 intravenously through the saphenous vein.
For the intravenous treatment, the tails of the mice were hanged in handwarm water for the sake of expansion of the saphenous vein. Survival of mice was determined as 96 hours after the LPS injection. It can be seen in Fig.3 that both Group 2 treated with PGFi a and Group 3 are protected in LPS-induced sepsis. It was well reflected in the survival of the animals. Example 6
Lipoploysaccharide (LPS) (from Escherichia coli bacterium) induced sepsis model in the case of treatment with the
compounds F05, F07 and F08 8 week Balb/c female mice were used in the experiment. The average body weight of the animals was 25 g. The groups used in the experiment: Group 1: control group obtaining only the carrier (98% physiological salt solution, 2% acetone); Group 2: group obtaining 250 g/kg PGFla; Group 4: group obtaining 250 μg/kg F05 compound; Group 5: group obtaining 250 g/kg F07 compound; Group 6: group obtaining 250 μg/kg F08 compound;
Dosing was repeated three times.
One hour before the treatment each animal obtained LPS (LPS Escherichia coli 0111.B4; Sigma L3012-10mg; 058K4006)
dissolved in 100 μΐ of physiological salt solution, once, intraperitoneally . LPS was used in a concentration of 5 mg/kg. After the treatment with LPS the animals were randomized. 10 animals were in each of the control and the treated groups.
Each animal obtained 250 g/kg FOl. 1, 4 and 8 hours after the treatment with LPS, the treated groups obtained 100 μΐ of
PGFia or the compounds F05, F07 and F08 respectively, through the saphenous vein.
For the intravenous treatment, the tails of the mice were hanged in handwarm water for the sake of expansion of the saphenous vein. Survival of mice was determined as 96 hours after the LPS injection. It can be seen in Fig.4 that each of the Group 2 treated with PGFic and the Groups 4, 5 and 6 are protected in LPS-induced sepsis. It was well reflected in the survival of the animals.
Example 7
Lipoploysaccharide (LPS) (from Escherichia coli bacterium) induced sepsis model in the case of treatment with the
compounds F06, F09, F10 and Fll
8 week Balb/c female mice were used in the experiment. The average body weight of the animals was 25 g. The groups used in the experiment: Group 1: control group obtaining only the carrier (98% physiological salt solution, 2% acetone); Group 2: group obtaining 250 g/kg PGFla; Group 7: group obtaining 250 μg/kg F06 compound; Group 8: group obtaining 250 g/kg F09 compound; Group 9: group obtaining 250 g/kg F10 compound;
Group 10: group obtaining 250 g/kg Fll compound. Dosing was repeated three times.
One hour before the treatment each animal obtained LPS (LPS Escherichia coli 0111.B4; Sigma L3012-10mg; 058K4006)
dissolved in 100 μΐ of physiological salt solution, once, intraperitoneally . LPS was used in a concentration of 2 mg/kg. After the treatment with LPS the animals were randomized. 10 animals were in each of the control and the treated groups.
Each animal obtained 250 μg/kg F01. 1, 4 and 8 hours after the treatment with LPS the treated groups obtained 100 μΐ of PGFi a or the compounds F06, F09, F10 and Fll, through the saphenous vein .
For the intravenous treatment, the tails of the mice were hanged in handwarm water for the sake of expansion of the saphenous vein. Survival of mice was determined as 96 hours after the LPS injection.
It can be seen in Fig.5 that each of the Group 2 treated with PGFi c and the Groups 7, 8, 9 and 10 are protected in LPS- induced sepsis. It was well reflected in the survival of the animals .
Example 8
Effects on TNBS (trinitrobenzene sulphonic acid) induced mucositis in the case of administration of the compound FOl
In the model of inflammatory bowel disease TNBS (2,4,6- trinitrobenzene sulphonic acid) induced mucositis was
inhibited by PGF ia (compound FOl) administered once,
intraperitoneally, in a dose of 300 μς/]^.
In the experiment, 10 mg TNBS was dissolved in 0.5 % CMC (carboxymethyl cellulose) .
In addition to reduction of the extent of mucous lesions, efficiency was indicated by a decrease in the levels of TNF-a and MPO (mieloperoxidase) enzyme activities. Significant decrease was caused by PGFi a in each of the three cases.
It is illustrated by Figs. 6-11. P values were indicated with asterisks above the columns of the diagrams. P value is the percentage of total inflamed area in relation to the area not inflamed. P values were calculated referred to the group treated with TNBS and were indicated in the figure as follows
* P < 0.05,
**P < 0.01, ***P < 0.001
Results ± S.E.M.; n = 7-12.
Example 9
Effects on TNBS (trinitrobenzene sulphonic acid) induced mucositis in the case of administration of the compounds F05, F07 and F08
In the model of the inflammatory bowel disease, TNBS (2,4,6- trinitrobenene sulphonic acid) induced mucositis was inhibited by the compounds F05, F07, F08 administered intravenously during the TNBS induction.
In the experiment, 10 mg of TNBS was dissolved in 0.5 % CMC ( carboxymethyl cellulose) and administration was usually performed once or twice during the experiments.
Eight groups took part in the experiments during which the following compounds were administered:
Group 1 : absolute control, that is untreated group;
Group 2 : the animals obtained 10 mg of TNBS, once, orally; Group 3 : the animals obtained 1 mg of the compound F05
TNBS +F07 treated group 0.33 mg/kg/day i.v. treatment
(0.5 mg/kg/i.v.; 0.4 ml 0.9% NaCl/rat) The treatment was performed twice a day, the specific dose was administered divided in two parts;
Group 4 : the animals obtained 2 mg of the compound F05
TNBS +F07 treated group 0.66 mg/kg/day i.v. treatment
(1.0 mg/kg/i.v.; 0.4 ml 0.9% NaCl/rat) The treatment was performed twice a day, the specific dose was administered divided in two parts;
Group 5 : the animals obtained 1 mg of the compound F07 TNBS +F07 treated group 0.33 mg/kg/day i.v. treatment (0.5 mg/kg/i.v. ; 0.4 ml 0.9% NaCl/rat) The treatment was performed twice a day; the specific dose was administered divided in two parts;
Group 6: the animals obtained 1 mg of the compound F08
TNBS +F07 treated group 0.33 mg/kg/day i.v. treatment
(0.5 mg/kg/i.v.; 0.4 ml 0.9% NaCl/rat) The treatment was performed twice a day; the specific dose was administered divided into two parts;
Group 7 : the animals obtained infliximab
TNBS + Infliximab 3.0 mg/kg, 0.4 ml/day i.v.;
Group 8 : the animals obtained sulphazalazin
SASP 50 CMC = group treated with TNBS + sulphazalazin 50 mg/kg/day (25 mg/kg/ p.o. b.i.d.; 0.5 ml 0.5% carboxymethyl cellulose 0.9% NaCl /rat) .
Rats were treated for 3 days, the first treatment occurred 1 hour after administration of TNBS. On day 4 the animals were over-anesthetized and the small intestine was cut out. The small intestine was cut into four parallel sections lengthwise and mieloperoxidase, TNF-a levels and the extension of
inflammation were determined.
Out of the compounds of the general formula (I), the effect of the compounds F05, F07, F08 on the extension of TNBS-induced mucous lesions is shown in Fig. 12. Efficiency manifested in the decrease of the extension of the mucous lesions.
In Fig. 12, P values were indicated with asterisks above the columns of the diagrams. P value is the percentage of total inflamed area in relation to the area not inflamed. P values were calculated referred to the group treated with TNBS and were indicated in the figure as follows: * P < 0.05,
**P < 0.01,
***P < 0.001
Results ± S.E.M.; n = 8-12.
We note that the initial number of animals decreased in certain groups due to the death caused by the inflammatory bowel disease. Example 10
Effects on TNBS (trinitrobenzene sulphonic acid) induced mucositis in the case of administration of the compound F07 in various concentrations In the model of inflammatory bowel disease, TNBS (2,4,6- trinitrobenzene sulphonic acid) induced mucositis was
inhibited by the compound F07 administered once during the TNBS induction intravenously, in a dose of 0.33mg/kg then 1 mg/kg .
In the experiment, 10 mg of TNBS was dissolved in 0.5 % CMC ( carboxymethyl cellulose) .
Out of the compounds with the general formula (I), the effect of the compound F07 on the extension of TNBS-induced mucous lesions in the case of administration of various
concentrations of the compound is shown in Fig. 13.
Example 11
Preparation of pharmaceutical compositions a) Tablets : 0.01-50% active substance of the general formula (I), 15-50% lactose, 15-50% potato starch, 5-15% polyvinylpyrrolidone, 1- 5% talc, 0.01-3% magnesium stearate, 1-3% colloid silicon dioxide and 2-7% ultraamylopectine are mixed, granulated by wet granulation and tablets are pressed. b) Dragees, film-coated tablets:
Tablets prepared according to the previous example are coated with a layer containing an entero- or gastrosolvent film or sugar and talc. The dragees are glazed with a mixture of beeswax and carnaubawax. c) Capsules :
0.01-50% active substance of the general formula (I), 1-5% sodium lauryl sulphate, 15-50% starch, 15-50% lactose, 1-3% colloid silicon dioxide and 0.01-3% magnesium stearate are mixed well; the mixture is pressed through a sieve and filled into hard gelatine capsules. d) Suspensions :
Ingredients: 0.01-15% active substance of the general formula (I), 0.1-2% sodium hydroxide, 0.1-3% citric acid, 0.05-0.2% nipagin (sodium methyl 4-hydroxybenzoate) , 0.005-0.02%
nipasol, 0.01-0.5% carbopol (polyacrylic acid), 0.1-5% 96% ethanol, 0.1-1% flavouring, 20-70% sorbitol (70 % aqueous solution) and 30-50% distilled water.
To a solution of nipagin and citric acid in 20 ml of distilled water, carbopol is added in small portions under intense stirring, the solution obtained is set aside and allowed to stand for 10-12 hours. Then the sodium hydroxide in 1 ml of distilled water, the aqueous solution of sorbitol and the raspberry flavouring in ethanol are added under intense stirring. The active substance is added to this carrier in small portions and suspended by means of an immersed homogenizer. The suspension is made up to volume with
distilled water and the suspension syrup is brought to its final distribution by means of a colloid mill. e) Suppositories :
0.01-15% active substance of the general formula (I) and 1-20% lactose are mixed well and the mixture obtained is added to 50-95% fat melted and cooled to 35°C, suitable for suppository making (e.g. Witepsol 4); the mixture obtained is cast into a cooled mould. f) Lyophilized powder ampoule preparations :
A 5 % mannitol or lactose solution is made using bidistilled water for injection and the solution is filtered sterile. A 0.01-5% sterile solution of the active substance of the general formula (I) is prepared in the same way. The solutions are mixed under aseptic conditions and filled into ampoules in 1 ml doses; the content of the ampoules is lyophilized and the ampoules are sealed under nitrogen. The content of the
ampoules is dissolved in sterile water or sterile
physiological salt solution (0.9 % common salt solution) just before being administered.