NZ252855A

NZ252855A - Polypeptide having terminal substituted by an alpha-substituted acetic acid, the substituent being a xanthene, thioxanthene or dibenzocycloheptene derivative

Info

Publication number: NZ252855A
Application number: NZ252855A
Authority: NZ
Inventors: Wayne Livingston Cody; Annette Marian Doherty; John Gordon Topliss
Original assignee: Warner Lambert Co
Priority date: 1992-04-22
Filing date: 1993-04-16
Publication date: 1996-11-26
Also published as: JPH07505890A; NO944013L; AU678357B2; KR950701344A; FI944905A0; CA2133090A1; EP0647236A1; NO944013D0; FI944905A; HUT68862A; CZ256994A3; WO1993021219A1; HU9403017D0; SK128794A3; AU4290493A

Description

<div class="application article clearfix" id="description"> New Zealand Paient Spedficaiion for Paient Number £52855 New Zealand No. International No. 252855 PCT/US93/03658 TO BE ENTERED AFTER ACCEPTANCE AND PUBUCATION Priority dates: International fifing date: Classification: COIKI/Ob' Afc>/ K3t>/0£> PubDcacion date: 2 6 No'v »96 Journal No.: lu.n COMPLETE SPECMCATION Title of invention: Endothelin antagonists II. Name, address and nationality of ' applicant(s) as in international application form: WARNER-LAMBERT COMPANY, 201 Tabor Road, Morris Plains, xt at.* tr\-mn a.. atrtrn mrt * NEW ZEALAND PATENTS ACT 1953 New Jersey 07950 US (FOLLOWED BY PAGE 1 A) -1- £5 2 8^ ENDOTHELIN ANTAGONISTS II BACKGROUND OF THE INVENTION The present invention relates to novel antagonists of endothelin useful as pharmaceutical agents, to methods for their production, and to pharmaceutical compositions which include these conqpounds and a pharmaceutically acceptable carrier. More particularly, the novel compounds of the present invention are antagonists of endothelin useful in treating elevated levels of endothelin, acute and chronic renal failure, hypertension, myocardial infarction, metabolic, endocrinological and neurological disorders, congestive heart failure, endotoxic shock, subarachnoid hemorrhage, arrhythmias, asthma, preeclampsia, atherosclerotic disorders including Raynaud's disease, restenosis, angina, cancer, pulmonary hypertension, ischemic disease, gastric mucosal damage, hemorrhagic shock, ischemic -bowel disease, and diabetes. Endothelin-1 (ET-1), a potent vasoconstrictor, is a 21 amino acid bicyclic peptide that was first isolated from cultured porcine aortic endothelial cells. Endothelin-1, is one of a family of structurally similar bicyclic peptides which include; ET-2, ET-3, vasoactive intestinal contractor (VTC), and the sarafotoxins (SRTXs). The unique bicyclic structure and corresponding arrangement of the disulfide bridges of ET-1, which are the same for the endothelins, VIC, and the sarafotoxins, has led to significant speculation as to the importance of the resulting induced secondary structure to receptor binding and functional activity. ET-1 analogues with incorrect disulfide pairings exhibit at least 100-fold less vasoconstrictor activity. The flexible OFFICE] 1 ?■" ."IN 1MB I O 93/21219 PCT/US93/03658 -2- C-terminal hexapeptide of ET-1 has been shown to be important for binding to the ET receptor and functional activity in selected tissues. Additionally, the C-tenninal amino acid (Trp-21) has a 5 critical role in binding and vasoconstrictor activity, since ET[l-20] exhibits approximately 1000-fold less functional activity. Endothelin is involved in many human disease states. 10 Several in vivo studies with ET antibodies have been reported in disease models. Left coronary artery ligation and reperfusion to induce myocardial infarction in the rat heart, caused a four- to sevenfold increase in endogenous endothelin levels. 15 Administration of ET antibody was reported to reduce the size of the infarction in a dose-dependent manner (Watanabe, T., et al, "Endothelin in Myocardial Infarction," Nature (Lond.) 344:114 (1990)). Thus, ET may be involved in the pathogenesis of congestive 20 heart failure and myocardial ischemia (Margulies, K.B., et al, "Increased Endothelin in Experimental Heart Failure," Circulation 82:2226 (1990)). Studies by Kon and colleagues using anti-ET 25 antibodies in an ischemic kidney model, to deactivate endogenous ET, indicated the peptide's involvement in acute renal ischemic injury (Kon, V., et al, "Glomerular Actions of Endothelin In Vivo," J. Clin. Invest. 83:1762 (1989)). In isolated kidneys, 30 preexposed to specific antiendothelin antibody and then challenged with cyclosporine, the renal perfusate flow and glomerular filtration rate increased, while renal resistance decreased as compared with isolated kidneys preexposed to a nonimmunized rabbit serum. 35 The effectiveness and specificity of the anti-ET antibody were confirmed by its capacity to prevent renal deterioration caused by a single bolus dose £vO 93/21219 PCT/US93/03658 -3- (150 pmol) of synthetic ET, but not by infusion of angiotensin II, norepinephrine, or the thromboxane A2 mimetic U-46619 in isolated kidneys (Perico, N., et al, "Endothelin Mediates the Renal Vasoconstriction 5 Induced by Cyclosporine in the Rat," J. Am. Soc. Neohrol. 1:76 (1990)). Others have reported inhibition of ET-1 or ET-2-induced vasoconstriction in rat isolated thoracic aorta using a monoclonal antibody to ET-1 (Koshi, T., 10 et al, "Inhibition of Endothelin (ET)-1 and ET-2- Induced Vasoconstriction by Anti-ET-1 Monoclonal Antibody," Chem. Pharm. Bull.. 39:1295 (1991)). Combined administration of ET-1 and ET-1 antibody to rabbits showed significant inhibition of the blood 15 pressure (BP) and renal blood flow responses (Miyamori, I., et al, Systemic and Regional Effects of Endothelin in Rabbits: Effects of Endothelin Antibody," Clin. Exp. Pharmacol. Phvsiol.. 17:691 (1990)). 20 Other investigators have reported that infusion of ET-specific antibodies into spontaneously hypertensive rats (SHR) decreased mean arterial pressure (MAP), and increased glomerular filtration rate and renal blood flow. In the control study with 25 normotensive Wistar-Kyoto rats (WKY) there were no significant changes in these parameters (Ohno, A. Effects of Endothelin-Specific Antibodies and Endothelin in Spontaneously Hypertensive Rats," J. Tokvo Women's Med. Coll.. 61:951 (1991)). 30 In addition, elevated levels of endothelin have been reported in several disease states (see Table I below). Burnett and co-workers recently demonstrated that exogenous infusion of ET (2.5 ng/kg/mL) to 35 anesthetized dogs, producing a doubling of the circulating concentration, did have biological actions (Lerman, A., et al, "Endothelin has Biological Actions 93/21219 PCT/US93/03658 -4- at Pathophysiological Concentrations," Circulation 83:1808 (1991)). Thus heart rate and cardiac output decreased in association with increased renal and systemic vascular resistances and antinatriuresis. 5 These studies support a role for endothelin in the regulation of cardiovascular, renal, and endocrine function. In the anesthetized dog with congestive heart failure, a significant two- to threefold elevation of 10 circulating ET levels has been reported (Cavero, P.6., et al, "Endothelin in Experimental Congestive Heart Failure in the Anesthetized Dog," AmT J. Phvsiol. 259:F312 (1990)), and studies in humans have shown similar increases (Rodeheffer, R.J., et al, 15 "Circulating Plasma Endothelin Correlates With the Severity of Congestive Heart Failure in Humans," Am. J. Hypertension 4:9A (1991)). When ET was chronically infused into male rats, to determine whether a long-term increase in circulating ET levels 20 would cause a sustciined elevation in mean arterial blood pressure, significant, sustained, and dose-dependent increases in mean arterial BP were observed. Similar results were observed with ET-3 although larger doses were required (Mortenson, L.H., et al, 25 "Chronic Hypertension Produced by Infusion of Endothelin in Rats," Hypertension. 15:729 (1990)). The distribution of the two cloned receptor subtypes, termed ETA and ETB, have been studied extensively (Arai, H., et al, Nature 348:730 (1990), 30 Sakurai, T., et al, Nature 348:732 (1990)). The ETA, or vascular smooth muscle receptor, is widely distributed in cardiovascular tissues and in certain regions of the brain (Lin, H.Y., et al, Proc. Natl. Acad. Sci. 88:3185 (1991)). The ETB receptor, 35 originally cloned from rat lung, has been found in rat cerebellum and in endothelial cells, although it is not known if the ETB receptors are the same from these O 93/21219 PCT/US93/03658 -5- sources. The human ET receptor subtypes have been cloned and expressed (Sakamoto, A., et al, Biochem. Biophvs. Res. Chem. 178:656 (1991), Hosoda, K., et al, FEBS Lett. 287:23 (1991)}. The ETA receptor clearly 5 mediates vasoconstriction and there have been a few reports implicating the ETB receptor in the initial vasodilatory response to ET (Takayanagi, P.., et al, FEBS Lett. 282:103 (1991)). However, recent data has shown that the ETB receptor can also mediate 10 vasoconstriction in some tissue beds (Panek, R.L., et al, Biochem. Biophvs. Res. Commun. 183(2):566 (1992)). Comparison of the receptor affinities of the ETs and SRTXs in rats and atria (ETA) or cerebellum and 15 hippocampus (ETB), indicate that SRTX-c is a selective ETB ligand (Williams, D.L., et al, Biochem. Biophvs. Res. Commun.. 175:556 (1991)). A recent study showed that selective ETB agonists caused only vasodilation in the rat aortic ring, possibly through the release 20 of EDRF from the endothelium (ibid). Thus, reported selective ETB agonists, for example, the linear smalog ET[1,3,11,15-Ala] and truncated analogs ET[6-21, 1,3,11,15-Ala], ET[8-21,ll,15-Ala], and N~Acetyl-ET[10-2l,ll,15-Ala] caused vasorelaxation in 25 isolated, endothelium-intact porcine pulmonary arteries (Saeki, T., et al, Biochem. Biophvs. Res. Commun. 179:286 (1991)). However, some ET analogs are potent vasoconstrictors in the rabbit pulmonary artery, a tissue that appears to possess an ETB 30 y, nonselective type of receptor (ibid). Plasma endothelin-l levels were dramatically increased in a patient with malignant hemangioendothelioma (K. Nakagawa et al, Nippon Hifuka Gakkai Zasshi. 1990, 100, 1453-1456). 35 The ET receptor antagonist BQ-123 has been shown to block ET-1 induced bronchoconstriction and tracheal smooth muscle contraction in allergic sheep providing 93/21219 PCT/US93/03658 -6- evidence for expected efficacy in bronchopulmonary diseases such as asthma (Noguchi, et al, Am. Rev. Respir. Pis.. 1992, 145 (4 Part 2), A858) . Circulating endothelin levels are elevated in 5 women with preeclampsia and correlate closely with serum uric acid levels and measures of renal dysfunction. These observations indicate a role for ET in renal constriction in preeclampsia (Clark B.A., et al. Am. J. Obstet. Gvnecol.. 1992, 166, 962-968). 10 Plasma immunoreactive endothelin-1 concentrations are elevated in patients with sepsis and correlate with the degree of illness and depression of cardiac output (Pittett J., et al, Ann Sura.. 1991, 213(3), 262) . 15 In addition the ET-l antagonist BQ-123 has been evaluated in a mouse model of endotoxic shock. This STa antagonist significantly increased the survival rate in this model (Toshiaki M., et al, 20.12.90. EP 0 436 189 Al). 20 Endothelin is a potent agonist in the liver eliciting both sustained vasoconstriction of the hepatic vasculature and a significant increase in hepatic glucose output (Gandhi C.B., et al, Journal of Biological Chemistry. 1990, 265(29), 17432). In 25 streptozotocin-diabetic rp.ts there is an increased sensitivity to endothelin-1 (Tammesild P.J., et al, Clin. Exp. Pharmacol. Physiol.. 1992, 19(4), 261). In addition increased levels of plasma ET-1 have been observed in microalbuminuria insulin-dependent 30 diabetes mellitus patients indicating a role for ET in endocrine disorders such as diabetes (Collier A., et al, Diabetes Care. 1992, 15(8), 1038). ETA antagonist receptor blockade has been found to produce an antihypertensive effect in normal to low 35 renin models of hypertension with a time course similar to the inhibition of ET-1 pressor responses (Basil M.K., et al, J. Hypertension. 1992, WO 93/21219 PCT/US93/03658 -7- 10(Svippl 4), S49) . The endothelins have been shown to be arrhythmogenic, and to have positive chronotropic and inotropic effects, thus ET receptor blockade would be expected to be useful in arrhythmia and other 5 cardiovascular disorders (Han S.-P., et al, Life Sci.. 1990, 46, 767). The widespread localization of the endothelins and their receptors in the central nervous system and cerebrovascular circulation has been described 10 (Nikolov R.K., et al, Drugs of Today. 1992, 28(5), 303-310). Intracerebroventricular administration of ET-l in rats has been shown to evoke several behavioral effects. These factors strongly suggest a role for the ETs in neurological disorders. The 15 potent vasoconstrictor action of ETs on isolated cerebral arterioles suggests the importance of these peptides in the regulation of cerebrovascular tone. Increased ET levels have been reported in some CNS disorders, i.e., in the CSF of patients with 20 subarachnoid hemorrhage and in the plasma of women with pree'clampsia. Stimulation with ET-3 under conditions of hypoglycemia have been shown to accelerate the development of striatal damage as a result of an influx of extracellular calcium. 25 Circulating or locally produced ET has been suggested to contribute to regulation of brain fluid balance through effects on the choroid plexus and CSP production. ET-1 induced lesion development in a new model of local ischemia in the brain has been 3 0 described. Circulating and tissue endothelin immunoreactivity is increased more than twofold in patients with advanced atherosclerosis (A. Lerman, et al, New England J. Med.. 1991, 325, 997-1001). 3 5 Increased endothelin immunoreactivity has also been associated with Buerger's disease (K. Kanno, et al, J. Amer. Med. Assoc.. 1990, 264, 2868) and Raynaud's 93/21219 PCT /US93/03658 -8- phenomenon (M.R. Zamora, et al, Lancet. 1990, 336, 1144-1147). Likewise, increased endothelin concentrations were observed in hypercholesterolemic rats (T. Horio, et al, Atherosclerosis. 1991, 89, 239-5 245). An increase of circulating endothelin levels was observed in patients that underwent percutaneous transluminal coronary angioplasty (PTCA) (A. Tahara, et al, Metab. Clin. Exp.. 1991, 40, 1235-1237, K. 10 Sanjay, et al, Circulation. 1991, 84(Suppl. 4), 726). Increased plasma levels of endothelin have been measured in rats (T.J. Stelzner, et al, Am. J. Physiol.. 1992, 262, L614-L620) and individuals (T. Miyauchi, et al, Jpn. J. Pharmacol.. 1992, 58, 15 279P, D.J. Stewart, et al, Ann. Internal Medicine. 1991, 114 464-469) with pulmonary hypertension. Elevated levels of endothelin have also been measured in patients suffering from ischemic heart disease (M. Yasuda, et al, Amer. Heart J.. 1990, 119 20 801-806, S.G. Ray, et al, Br. Heart J.. 1992, 67, 383- 386) and either stable or unstable angina (J.T. Stewart, et al, Br. Heart J.. 1991, 66, 7-9). Infusion of an endothelin antibody lh prior to and lh after a 60 minute period of renal ischaemia 25 resulted in changes in renal function versus control. In addition, an increase in glomerular platelet-activating factor was attributed to endothelin (A. Lopez-Parre, et al, J. Physiology. 1991, 444, 513-522). In patients with chronic renal failure as well 30 as in patients on regular hemodialysis treatment mean plasma endothelin levels were significantly increased (F. Stockenhuber, et al, Clin. Sci. (Lond.). 1992, 82, 255-258) . In addition it has been suggested that the proliferative effect of endothelin on mesangial cells 35 may be a contributing factor in chronic renal failure (P.J. Schultz, J. Lab. Clin. Med.. 1992, 119, 448-449) . fW'O 93/21219 PCT/US93/03658 -9- Local intra-arterial administration of endothelin has been shown to induce small intestinal mucosal damage in rats in a dose-dependent manner (S. Mirua, et al. Digestion. 1991, 48, 163-172). Administration 5 of endothelin-1 in the range of 50-500 pmol/kg into the left gastric artery increased the tissue type plasminogen activator release and platelet activating formation, and induced gastric mucosal hemorrhagic change in a dose dependent manner (I. Kurose, et al, 10 Gut. 1992, 33, 868-871). Furthermore, it has been shown that an anti-ET-1 antibody reduced ethanol-induced vasoconstriction in a concentration-dependent manner (E. Masuda, et al. Am. J. Phvsiol.. 1992, 262, G785-G790). Elevated endothelin levels have been 15 observed in patients suffering from Crohn's disease and ulcerative colitis (S.H. Murch, et al, Lancet. 1992, 339, 381-384). Recently the nonpeptide endothelin antagonist RO 46-2005 has been reported to be effective in models 20 of acute renal ischemia and subarachnoid hemorrhage in rats (3rd International Conference on Endothelin. Houston, Texas, February 1993). In addition, the ETA antagonist BQ-123 has been shown to prevent early cerebral vasospasm following subarachnoid hemorrhage 25 (M. Clozel and H. Watanabe, Life Sci.. 52:825-834 (1993)). ^ O 93/21219 PCT/US93/03658 -10- TABLE I. Plasma Concentrations of ET-1 in Humans Condition Normal Control ET Plasma Levelb Reported (pg/mL) Atherosclerosis 1.4 3.2 pmol/L Surgical operation 1.5 7.3 5 Buerger's disease 1.6 4.8 Takayasu's arteritis 1.6 5.3 Cardiogenic shock 0.3 3.7 Congestive heart failure (CHF) 9.7 20.4 Mild CHF 7.1 11.1 10 Severe CHF 7.1 13.8 Dilated cardiomyopathy 1.6 7.1 Preeclampsia 10.4 pmol/L 22.6 pmol/L Pulmonary hypertension 1.45 3.5 Acute myocardial infarction 1.5 3.3 15 (several reports) 6.0 11.0 0.76 4.95 0.50 3.8 Subarachnoid hemorrhage 0.4 2.2 Crohn's Disease , . 0-24 fmol/mg 4-64 fmol/mg 20 Ulcerative colitis 0-24 fmol/mg 20-50 fmol/mg Cold pressor test 1.2 8.4 Raynaud's phenomenon 1.7 5.3 Raynaud's/hand cooling 2.8 5.0 Hemodialysis <7 10.9 25 (several reports) 1.88 4.59 Chronic renal failure 1.88 10.1 Acute renal failure 1.5 10.4 Uremia before hemodialysis 0.96 1.49 Uremia after hemodialysis 0.96 2.19 30 Essential hypertension 18.5 33.9 Sepsis syndrome 6.1 19.9 Postoperative cardiac 6.1 11.9 Inflammatory arthritides 1.5 4.2 Malignant hemangioendothelioma 4.3 16.2 35 (after removal) Rovero, P., et al, British Journal of Pharmacology 101. pages 232-236 (1990) disclosed 40 various analogs of the C-terminal hexapeptide of ET-l, none of which were reported to be antagonists of ET-l. Doherty, A. M., et al, Abstract, Second International Conference on Endothelin, Tsukuba, Japan, December 9, 1990, and the published manuscript 45 (J. Cardiovasc. Pharm. 17 (Suppl. 7), 1991, pp. 559-561) disclosed various analogs of the C-terminal hexapeptide of ET-l, none of which exhibited any functional activity. Qvo 93/21219 PCT/US93/03658 -11- Copending United States Patent Application Serial Number 07/995,480 discloses a series of novel antagonists of endothelin. However, we have surprisingly and unexpectedly 5 found that a series of C-terminal hexapeptide and related analogs of ET-1 are receptor antagonists of endothelin. Additional data for the activity of this series of peptides is found in the following references (W.L. Cody, et al, J. Med. Chem.. 1992, 35, 10 3301-3303., D.M. LaDouceur, et al, FASEB, 1992). SUMMARY OF THE INVENTION 15 Accordingly, the present invention is a compound of Formula I AA1 - AA2 - AA3 - AA4 - AA5 - AA6 I 20 wherein AA1 is R1 O R-C-C— 25 30 wherein R is hydrogen, 35 alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, aryl, heteroaryl, fluorenylmethyl, PCT/US93/03658 . -12- N-R2 2 3 , wherein R and R are each the same or l3 R different and each xs hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, heteroaryl, or fluorenylmethyl, 0 II 2 2 -C-OR , wherein R is as defined above, -OR2, wherein R2 is as defined above, 0 1 3 _ a -N-C-N-R , wherexn R and R are as defxned ^2 |^2 above, 0 'I 9 -C-C(Rs,)3, wherein Rs is F, CI, Br, or I, -CH2-OR2, wherein R2 is as defined above, O « 3 -N-C-R , l2a R wherein R2a is hydrogen or alkyl and R3 is as defined above, O « 3 -N-C-OR , wherein R2a and R3 are as defined above excluding R3 is hydrogen, or 0 II 2 -C-R , wherexn R^ xs as defxned above, R1 is hydrogen or alkyl, O 93/21219 PCT/US93/03658 -13- Z is -O- , -S(0)fif wherein m is zero or an integer of 5 1 or 2, -N-, wherein R2 is as defined above, R2 -(CH2)n-, wherein n is zero or an integer 10 of 1, 2, 3, or 4, -(CH2)n-CH=CH-(CH2)n-, wherein n is as defined above, O II 15 -C-, -CR1-, wherein R1 and R2 are as defined ,2 above, or 20 35 OR R2 -C- , l3 R wherein R2 and R3 are each the same or 25 different and each is as defined above, X and Y are the same and substituted at the same position on the aromatic ring and each may be one, two, three, or four substituents selected from the group consisting of 3 0 hydrogen, halogen, alkyl, -C02R2, wherein R2 is as defxned above, 2 2 3 -CONR , .-/herein R and R are as defxned l3 R above, 2 2 3 -NR , wherein R and R are as defined I3 . R above, or ^VO 93/21219 PCT/US93/03658 -14- 15 nitro or 10 wherein R, Z, X, and Y are as defined above; AA2 is R1 O I * II —N—C-C— | I R1 (CH2)n R4 wherein R4 is hydrogen, alkyl, 2 0 alkenyl, alkynyl, cycloalkyl, aryl. 25 -N-R3b heteroaryl, -R R2b wherein R*4"" and RJW are each the ,2b -,3b same or different and each is 30 hydrogen, alkyl, cycloalkyl, aryl, or heteroaryl, 35 -OR2b, wherein R2b is as defined above, O -LH-R3b, 40 R2b Qvo 93/21219 PCT/US93/03658 -15- wherein R2b and R3b are each the same or different and each is as defined above for R2b and R3b, II 2b 2b -C-R , wherein R is as defined above, NH -NH-C-NH-R2b wherein R2b is as defined above, or O I! 2b 15 -C-OR , wherein R is as defined above, and R1 and n are as defined above, or AA2 is absent; AA3 is 10 20 R1 O I * II —N —C-C— I I R1(CH2)n R5 25 wherein R5 is hydrogen, alkyl, aryl, heteroaryl, 30 O II 3b -C-N-R , 1.2b OK OK 35 wherein R and R are each the same or different and each is as defined above, $VO 93/21219 PCT/IJS93/03658 -16- C-R2b, wherein R2*3 is as defined above, or II 2b -C-OR , wherein R xs as defined above, and R1 and n are as defined above, or 10 AA3 is absent; AA4 and AA5 are each independently absent or each is independently R1 0 I * II —N—C—C— I I 15 R1 (CH2)n 30 40 R6 wherein Rs is hydrogen, alkyl, 2 0 alkenyl, alkynyl,. cycloalkyl aryl, or heteroaryl, and 25 R1 and n are as defined above; AA6 is R1 8 -N--C -R ' 'i 1 R1 (CH2)_ I R7 35 wherein R7 is aryl or heteroaryl, R8 is O •C-OR1, wherein R1 is as defined above, £vO 93/21219 PCT/US93/03658 -17- -OR1, wherein R1 is as defined above, 0 1 i 5 -C-N-R-1-, wherein Rx is as defined above, or -CHj-OR1, wherein R1 is as defined above, and 10 R1 and n are as defined above; * stereochemistry at C in AA1, AA2, AA3, AA4, or AA5 is D, L, or DL and * 15 stereochemistry at C in AA6 is L; or a pharmaceutically acceptable salt thereof. Elevated levels of endothelin have been postulated to be involved in a number of pathophysiological states including diseases 20 associated with the cardiovascular system as well as various metabolic and endocrinological disorders. As antagonists of endothelin, the compounds of Formula I are useful in the treatment of hypertension, myocardial infarction, metabolic, endocrinological and 25 neurological disorders, congestive heart failure, endotoxic shoe'::, subarachnoid hemorrhage, arrhythmias, asthma, and chronic and acute renal failure, preeclampsia, atherosclerotic disorders including Raynaud's disease, restenosis, angina, cancer, 30 pulmonary hypertension, ischemic disease, gastric mucosal damage, hemorrhagic shock, ischemic bowel disease, and diabetes. A still further embodiment of the present invention is a pharmaceutical composition for 35 administering an effective amount of a compound of Formula I in unit dosage form in the treatment methods mentioned above. Finally, the present invention is directed to methods for production of a compound of Formula I. 40 ®v O 93/21219 PCT/US93/03658 -18- DETAILED DESCRIPTION OF THE INVENTION In the compounds of Formula I, the term "alkyl" means a straight or branched hydrocarbon radical 5 having from 1 to 12 carbon atoms and includes, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, n-hexyl, n-heptyl, n-octyl, n-nonyl, n-decyl, undecyl, dodecyl, and the like. 10 The term "alkenyl" means a straight or branched unsaturated hydrocarbon radical having from 2 to 12 carbon atoms and includes, for example, ethenyl, 2-propenyl, 1-butenyl, 2-butenyl, 1-pentenyl, 2-pentenyl, 3-methyl-3-butenyl, 1-hexenyl, 2-hexenyl, 15 3-hexenyl, 3-heptenyl, l-octenyl, 1-nonenyl, l-decenyl, l-undecenyl, l-dodecenyl, and the like. The term "alkynyl" means a straight or branched triple bonded unsaturated hydrocarbon radical having from 2 to 12 carbon atoms and includes, for example, 20 ethynyl, 2-propynyl, l-butynylf 2-butynyl, 3-butynyl, 1-pentynyl, 3-pentynyl, l-hexynyl, 2-hexynyl, 3-hexynyl, 3-heptynyl, 1-octynyl, 2-octynyl, 1-nonynyl, 2-nonynyl, 3-nonynyl, 4-nonynyl, 1-decynyl, 2-decynyl, 2-undecynyl, 3-undecynyl, 3-dodecynyl, and 25 the like. The term "cycloalkyl" means a saturated hydrocarbon ring which contains from 3 to 12 carbon atoms, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, adamantyl, and the like. 30 The term "cycloalkylalkyl" means a saturated hydrocarbon ring attached to an alkyl group wherein alkyl is as defined above. The saturated hydrocarbon ring contains from 3 to 12 carbon atoms. Examples of such are cyclopropylmethyl, cyclopentylmethyl, 35 cyclohexylmethyl, adamantylmethyl and the like. The terms "alkoxy" and "thioalkoxy" are O-alkyl or S-alkyl as defined above for alkyl. fVO 93/21219 PCT/ US93/03658 10 -19- The term "aryl" means an aromatic radical which is a phenyl group, a benzyl group, a naphthyl group, a biphenyl group, a pyrenyl group, an anthracenyl group, 3,3-diphenylalanyl, 10,ll-dihydro-5H-dibenzo[a,d]-(cyclohepten-5-yl)glycyl, or a fluorenyl group and the like, unsubstituted or substituted by 1 to 4 substituents selected from alkyl as defined above, alkoxy as defined above, thioalkoxy as defined above, O II hydroxy, thiol, nitro, halogen, amino, -NH-C-alkyl 0 II wherein alkyl is as defined above, -C-O-alkyl wherein 15 0 II alkyl is as defined above, -C-alkyl wherein alkyl is as defined above, or aryl. The term "arylalkyl" means an aromatic radical 20 attached to an alkyl radical wherein aryl and alkyl are as defined above for example benzyl, fluorenylmethyl and the like. The term "heteroaryl" means a heteroaromatic radical which is 2-or 3-thienyl, 2- or 3-furanyl, 2-25 or 3-pyrrolyl, 2-, 4-, or 5-imidazolyl, 3-, 4-, or 5-pyrazolyl, 2-, 4-, or 5-thiazolyl, 3-, 4-, or 5-isothiazolyl, 2-, 4-, or 5-oxazolyl, 3-, 4-, or 5-isoxazolyl, 3- or 5-1,2,4-triazolyl, 4- or 5-1,2,3-triazolyl, tetrazolyl, 2-, 3-, or 4-pyridinyl, 30 3-, 4-, or 5-pyridazinyl, 2-pyrazinyl, 2-, 4-, or 5-pyrimidinyl, 2-, 3-, 4-, 5-, 6-, 7-, or 8-quinolinyl, 1-, 3-, 4-, 5-, 6-, 7-, or 8-isoquinolinyl, 2-, 3-, 4-, 5-, 6-, or 7-indolyl, 2-, 3-, 4-, 5-, 6-, or 7-benzo[b]thienyl, or 2-, 4-, 5-, 35 6-, or 7-benzoxazolyl* 2-, 4-, 5-, 6-, or 7-benzimidazolyl, 2-, 4-, 5-, 6-, or 7-benzothiazolyl, unsubstituted or substituted by 1 to 2 substituents selected from alkyl as defined above, aryl as defined 0vo 93/21219 PCT/US93/03658 10 -20- above, alkoxy as defined above, thioalkoxy as defined above, hydroxy, thiol, 0 II nitro, halogen, formyl, amino, -NH-C-alkyl wherein O II alkyl is as defined above, -C-O-alkyl wherein alkyl O is as defined above, -C-alkyl wherein alkyl is as defined above or phenyl. The term "heterocycloalkyl" means 2- or 3-tetrahydrothieno, 2- or 3 -tetrahydrofurano, 2- or 15 3-pyrrolidino, 2-, 4-, or 5-thiazolidino, 2-, 4-, or 5-oxazolidino, 2-, 3-, or 4-piperidino, N-morpholinyl or N-thiamorpholinyl. "Halogen" is fluorine, chlorine, bromine or iodine. 20 The following table provides a list of abbreviations and definitions thereof used in the present invention. 0vo 93/21219 PCT/US93/03658 -21-TABLE Abbreviation* Amino Acid Ala Alanine 5 Arg Arginine Asn Asparagine Asp Aspartic acid Cys Cysteine Glu Glutamic acid 10 Gin Glutamine Gly Glycine His Histidine He Isoleucine Leu Leucine 15 Lys Lysine Met Methionine Phe Phenylalanine Pro Proline Ser Serine 20 Thr Threonine Trp Tryptophan Tyr Tyrosine Val Valine 25 Abbreviation* Modified and Unusual Amino Acid Bhg 10, ll-Dihydro-5H-dibenzo [a,d] - (cyclohepten-5-yl)glycine or a-Amino-10,11-dihydro-5H-dibenzo-[a,d]cycloheptene-5-acetic acid Bip (Paraphenyl)phenylalanine If the optical activity of the amino acid is other than L(S), the amino acid or abbreviation is preceded by the appropriate configuration D(R) or DL(RS). |WO 93/21219 PCT/US93/03658 -22- 10 15 20 25 Abbreviation* Dip 3 Hyp 4 Hyp N-MePhe N-MeAsp Nva Nle Orn Abu Alg Arg (N02) Atm Cpn Chx Emg His(Dnp) HomoGlu HomoPhe Met(O) Met(02) 1-Nal 2-Nal Nia Pgl Pgy Modified and Unusual Amino Acid (cont) 3,3-Diphenylalanine 3 -Hydroxyproline 4-Hydroxyproline N- Me thylphenylalanine N-Methylaspartic acid Norvaline Norleucine Ornithine 2-Aminobutyric acid 2-Amino-4-pentenoic acid (Allylglycine) I^-nitroarginine 2-Amino-3-(2-amino-5-thiazole)propanoic acid 2-Amino-3-cyclopropanepropanoic acid (Cyclopropylalanine) Cyclohexylalanine (Hexahydrophenyl-alanine) 2-Amino-4,5(RS)-epoxy-4-pentenoic acid Nim-2,4-Dinitrophenylhistidine 2-Aminoadipic acid 2-Amino-5-phenylpentanoic acid (Homophenylalanine) Methionine sulfoxide Methionine sulfone 3 -(1'-Naphthyl)alanine 3 - (2'-Naphthyl)alanine 2-Amino-3-cyanopropanoic acid (Cyanoal anine) Phenylglycine 2-Aminopentanoic acid (Propylglycine) ^VO 93/21219 PCT/US93/03658 -23- 10 Abbreviation* Pha Pyr Tic Tza Tyr(Ot-Bu) Tyr(OMe) Tyr(OEt) Trp(For) Bheg Txg Modified and Unusual Amino Acid (cont) 2-Amino-6-(1-pyrrolo)-hexanoic acid 2-Amino-3-(3-pyridyl)-propanoic acid (3-Pyridylalanine) 1,2,3,4-Tetrahydro-3 -isoquinolinecarboxylic acid 2-Amino-3-(4-thiazolyl)-propanoic acid O-tertiary butyl-tyrosine O-Methyl-tyrosine O-Ethyl-tyrosine Nin- Formyl - tryptophan 5H-Dibenzo [a, d] cycloheptene glycine 9H-Thioxanthene glycine Abbreviation Protecting Group Ac Acetyl 15 Ada l-Adamantyl acetic acid Adoc Adamantyloxycarbonyl Bzl Benzyl MeBzl 4-Methylbenzyl Z Benzyloxycarbonyl 20 2-Br-Z ortho-Bromobenzyloxycarbonyl 2-CI-Z ortho-Chlorobenzyloxycarbonyl Bom Benzyloxymethyl Boc tertiary Butyloxycarbonyl TBS tertiary Butyldimethylsilyl 25 Dnp 2,4-Dinitrophenyl For Formyl Fmoc 9-Fluorenylmethyloxycarbonyl N02 Nitre Tos 4-Toluenesulfonyl (tosyl) O 93/21219 PCT/US93/03658 . -24- Abbreviation Protecting Group (cont) Trt Triphenylmethyl (trityl) Ada 1-Adamantyl acetic acid Bz Benzylcarbonyl tBu t-Butylcarbonyl CF3CO Trifluoroacetyl Cxi Cyclohexylacetyl Cxi (U) Cyclohexylurea Et Propionyl Pya 3-Pyridylacetyl Me(U) Methylurea Abbreviation Solvents and Reagents HOAc Acetic acid CH3CN Acetonitrile DCM Pi chlorome thane PCC N,N' -Dicyclohexylcarbodiimide PIEA N, N-Diisopropylethylamine PMF Pimethylfonnamide HC1 Hydrochloric acid HF Hydrofluoric acid HOBt 1-Hydroxybenzotriazole KOH Potassium hydroxide TFA Trifluoroacetic acid MBHA Resin Methylbenzhydrylamine resin PAM Resin 4-(Oxymethyl)-phenylacetamidomethyl resin The confounds of Formula I are capable of further forming both phannaceutically acceptable acid addition and/or base salts. All of these forms are within the scope of the present invention. £WO 93/21219 PCT/US93/03658 -25- Phannaceutically acceptable acid addition salts of the compounds of Formula I include salts derived from nontoxic inorganic acids such as hydrochloric, nitric, phosphoric, sulfuric, hydrobromic, hydriodic, 5 hydrofluoric, phosphorous, and the like, as well as the salts derived from nontoxic organic acids, such as aliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxy alkanoic acids, alkanedioic acids, aromatic acids, aliphatic 10 and aromatic sulfonic acids, etc. Such salts thus include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, mouehydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, acetate, trifluoroacetate, 15 propionate, caprylate, isobutyrate„ oxalate, malonate, succinate, suberate, sebacate, fumarate, maleate, mandelate, benzoate, chlorobenzoate, isethylbenzoate, dinitrobenzoate, ph'c-halate, benzenesulfonate, toluenesulfonate, phenylacetate, citrate, lactate, 20 maleate, tartrate, methanesulfonate, and the like. Also contemplated are salts of amino acids such as arginate and the like and gluconate, galacturonate (see, for example, Berge, S. M., et al, "Pharmaceutical Salts,n Journal of Pharmaceutical 25 Science. 6£, pp. 1-19 (1977)). The acid addition salts of said basic compounds are prepared by contacting the free base form with a sufficient amount of the desired acid to produce the salt in the conventional manner. Preferably a peptide 30 of Formula I can be converted to an acidic salt by treating with an aqueous solution of the desired acid, such that the resulting pH is less than 4. The solution can be passed through a CI8 cartridge to absorb the peptide, washed with copious amounts of 35 water, the peptide eluted with a polar organic solvent such as, for example, methanol, acetonitrile, aqueous mixtures thereof, and the like, and isolated by 0WO 93/21219 PCT/US93/03658 -26- concentrating under reduced pressure followed by lyophilization. The free base form may be regenerated by contacting the salt form with a base and isolating the free base in the conventional manner. The free 5 base forms differ from their respective salt forms somewhat in certain physical properties such as solubility in polar solvents, but otherwise the salts are equivalent to their respective free base for purposes of the present invention. 10 Phannaceutically acceptable base addition salts are formed with metals or amines, such as alkali and alkaline earth metals or organic amines. Examples of metals used as cations are sodium, potassium, magnesium, calcium,. and the like. Examples of 15 suitable amines are N,N'-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, dicyclohexylamine, ethylenediamine, N-methylglucamine, and procaine (see, for example, Berge, S. M., et al., "Pharmaceutical Salts," Journal of Pharmaceutical 20 Science. ££, pp. 1-19 (1977)). The base addition salts of said acidic compounds are prepared by contacting the free acid form with a sufficient amount of the desired base to produce the salt in the conventional manner. Preferably, a 25 peptide of Formula I can be converted to a base salt by treating with em aqueous solution of the desired base, such that the resulting pH is greater than 9. The solution can be passed through a C18 cartridge to absorb the peptide, washed with copious amounts of 30 water, the peptide eluted with a polar organic solvent such as, for example, methanol, acetonitrile, aqueous mixtures thereof, and the like, and isolated by concentrating under reduced pressure followed by lyophilization. The free acid form may be regenerated 35 by contacting the salt form with an acid and isolating the free acid in the conventional manner. The free acid forms differ from their respective salt forms O 93/21219 PCT/US93/03658 -27- somewhat in certain physical properties such as solubility in polar solvents, but otherwise the salts are equivalent to their respective free acid for purposes of the present invention. 5 Certain of the compounds of the present invention can exist in unsolvated forms as well as solvated forms, including hydrated forms. In general, the solvated forms, including hydrated forms, are equivalent to unsolvated forms and are intended to be 10 encompassed within the scope of the present invention. Certain of the compounds of the present invention possess one or more chiral centers and each center may exist in the R(D) or S(L) configuration. The present invention includes all enantiomeric and epimeric forms 15 as well as the appropriate mixtures thereof. A preferred compound of Formula I is one wherein AA1 is 20 X—■— II I —5i—Y 'Z' wherein R is -N-R2, ,R3 25 wherein R2 and R3 are each the same or different and each is hydrogen, alkyl, alkenyl, 3 0 alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, 35 heteroaryl, or fluorenylmethyl, 93/21219 PCT/US93/03658 •28- -N-C-N-R3, wherein R2 and R3 are as 5 R2 R2 defined above, 0 I' 9 -C-C(R )3, wherein R3 is F, CI, Br, or I, 10 o II 3 3 . • -NH-C-R , wherein R is as defined 15 above, or -NH-C-OR3, wherein R3 is as defined above excluding R3 is hydrogen, Z is -O-, 20 "S(0)fif wherein m is zero or an integer of 1 or 2, -N-, wherein R2 is as defined above, 25 R2 - (CH2)jj, wherein n is zero or an integer of 1, 2, 3, or 4, -(CH_) -CH-CH-(CH_) -, wherein n is as a ix m zjl defined above, 30 -C-, -CH-, wherein R1 is hydrogen or alkyl, 35 OR1 R2 I -C-, <3 40 RJ wherein R2 and R3 are each the same or different and each is as defined above and ,WO 93/21219 PCT/US93/03658 -29- X and Y are the same and substituted at the same position on the aromatic ring and each substituent is selected from the group consisting of 5 hydrogen, halogen, or alkyl; AA* is Q II —NH—CH—C— 10 I (CH2)n R4 wherein R4 is hydrogen, 15 alkyl, alkenyl, alkynyl, cycloalkyl, aryl, 2 0 heteroaryl, -N-R3b, R2b wherein R2b and R3b are each 25 the same or different and each is hydrogen, alkyl, cycloalkyl, 30 aryl, or heteroaryl, -OR2b, wherein R2b is as defined above, .0 35 £-N-R3b, R2b 0VO 93/21219 PCT/US93/03658 -30- wherein R2b and R3b are each the same or different and each is as defined above for R2b and R3b, 0 1 2b -C-R , wherein R2b is as defined above, NH -NH-C-NH-R2b, wherein R2b is as defined above, or O II 2b 15 -C-OR , wherein R is as defined above, and n is as defined above or AA2 is absent; O 10 AA3 is II 2 0 —NH—CH—C<— I (CH2)n ,5 25 wherein R5 is aryl, heteroaryl, 0 30 -C-N-R3b, i2b wherein R and R are each the same or different and each is as 35 defined above, 0 1 2b -5K -C-R , wherein R^° is as defined above, or WO 93/21219 PCT/US93/03658 10 -31- 0 II 2b 5K -C-OR , wherein R ° is as defined above, and n is as defined above, or AA3 is absent; AA4 and AA5 are each independently absent or each is O II —NH—CH—C— 1 (CH2)n R6 wherein R6 is hydrogen 15 alkyl, alkenyl, alkynyl, cycloalkyl, aryl, or 2 0 heteroaryl, and n is as defined above; AAS is -NH-*CH-C0,H I 25 (CH2) n I R7 wherein R7 is aryl or 30 heteroaryl, and n is as defined above, or 0 . 1 -NH- CH-C--N-R1 35 || wherein R7, R1, and n are as ^<TH2^n R defined above, R7 kWO 93/21219 PCT/US93/03658 10 -32- * 1 2 3 4 5 stereochemistry at CH m AA , AA , AA , AA , or AA is D,L, or DL, and * stereochemistry at CH in AA6 is L; or a phannaceutically acceptable salt thereof. A more preferred compound of Formula I is one wherein AA1 is 2 •• 15 wherexn R is -N-R , i3 RJ 2 3 wherein R and R are each the same or different and each is 20 hydrogen, alkyl, aryl, or fluorenylmethyl, 0 25 II -N-C-N-R3, wherein R2 and R3 are p2 jj2 as defined above, O 30 || -C-C(R9)3,wherein R9 is F, CI, Br, or I, or O II 10 in . 35 -NH-C-0R , wherein R1U is hydrogen, alkyl, aryl, or arylalkyl, excluding R10 is hydrogen, Z is -0-, 40 -S-, -NH- , gMVO 93/21219 PCT/US93/03658 -33- -{CH2)n, wherein n is zero or an integer of l, 2, 3, or 4, or - (CH2)n-CH=CH- (CH^jj, 5 wherein n is zero or an integer of l or 2 and X and Y are each the same and substituted at the same position on the aromatic ring ax. i ®ach substituent is selected from 10 the group consisting of hydrogen, halogen, or alkyl; AA2 is 15 O II -NH-CH-C- , 20 «fH2>n R4 wherein R4 is hydrogen, alkyl, aryl, 25 heteroaryl, -N-R3b , i2b wherein R2b and R3b are 30 each the same or different and each is hydrogen or alkyl, 0 » 3b 35 -C-N-R ' Ub R wherein R2b and R3b are each the same or 40 different and each is hydrogen or alkyl, AVG 93/21219 PCT/US93/03658 -34-NH -NH-C-NH-R2b wherein R2b is as defined above, 5 or O -C-OR213, wherein R2b is as defined above, and 10 n is an integer of 1, 2, 3, or 4 or AA2 is absent; AA3 is O II 15 -NH-CH-C-, I (^H2)a R5 wherein R5 is 20 aryl, heteroaryl, O -C-NHR3b, wherein R3b is 25 hydrogen or alkyl, 0 -C-R2b, wherein R2b is 3 0 hydrogen or alkyl, or O -C-OR2b, wherein R2b is 3 5 hydrogen or alkyl, and n is an integer of 1, 2, 3, or 4; AA4 and AA5 are each independently 0 40 •NH-CH-C- Wn 45 R6 C 03/21219 PCT/ US93/03658 -35- wherein Re. is hydrogen, alkyl, cycloalkyl, or aryl, and n is an integer of 1, 2, 3, or 4; AA6 is * -NH-CH- CCUH I 10 (CH2)n l7 R wherein R7 is aryl or heteroaryl, and n is zero or an integer of 1, 2, 3, 15 or 4, or 0 . II -NH-CH-C--N-R1 | | wherein R7, R1, and n are as 20 ^?H2^n R defined above, R7 * 1 2 3 4 5 stereochemistry at CH in AA , AA. , AA , AA , or AA is 25 D, L, or DL and * stereochemistry at CH in AA6 is L; or a pharmaceutically acceptable salt thereof. Particularly valuable are: 30 L-Bhg-Leu-Asp-lie-Ile-Trp; D-Bhg-Leu-Asp-Ile-Ile-Trp; Ac-L-Bhg-Leu-Asp-Ile-Ile-Trp; Ac-D-Bhg-Leu-Asp-lie-Ile-Trp; Ac-D-Bhg-Orn-Asp-lie-Ile-Trp; 35 Ac-D-Bhg-Lys-Asp-Ile-Ile-Trp; Ac-D-Bhg-Asp-Asp-lie-Ile-Trp; Ac-D-Bhg-Glu-Asp-lie-Ile-Trp; Ac-D-Bhg-Phe-Asp-Ile-Ile-Trp; Ac-D-Bhg-Arg-Asp-Ile-Ile-Trp; 40 Ac-D-Bhg-Asp-Ile-Ile-Trp; Pmoc-D-Bhg-Leu-Asp-lie-Ile-Trp; Fmoc-D-Bhg-Orn-Asp-lie-Ile-Trp; -36- Fmoc-D-Bhg-Lys-Asp-Ile-Ile-Trp; Fmoc-D-Bhg-Asp-Asp-lie-IIe-Trp; Fmoc-D-Bhg-Glu-Asp-lie-lie-Trp; Fmoc-D-Bhg-Phe-Asp-lie-lie-Trp; Fmoc-D-Bhg-Arg-Asp-lie-Ile-Trp; Fmoc-D-Bhg-Asp-lie-lie-Trp; Ac-D-Bhg-Leu-Phe-lie-Ile-Trp; Ac-D-Bhg-Leu-Asn-lie-Ile-Trp; Ac-D-Bhg-Leu-Glu-Ile-Ile-Trp; Ac-D-Bhg-Leu-Gin-lie-lie-Trp; Ac-D-Bhg-Leu-Tyr-lie-Ile-Trp; Ac-D-Bhg-Leu-1-Nal-lie-lie-Trp; Ac-D-Bhg-Leu-2-Nal-Ile-Ile-Trp; Ac-D-Bhg-Leu-Trp-lie-Ile-Trp; Ac-D-Bhg-Leu-Asp-Val-IIe-Trp; Ac-D-Bhg-Leu-Asp-Ile-Val-Trp; Ac-D-Bhg-Leu-Asp-Chx-Ile-Trp; Ac-D-Bhg-Leu-Asp-lie-Chx-Trp; Ac - D - Bhg-Arg-Asp - lie - Chx-Trp; Ac-D-Bhg-Lys-Asp-lie-Chx-Trp; Ac-D-Bhg-Orn-Asp-lie-Chx-Trp; Ac-D-Bhg-Asp-Asp-He-Chx-Trp ; Ac-D-Bhg-Glu-Asp-lie-Chx-Trp; Fmoc-D-Bhg-Leu-Phe-lie-lie-Trp; Fmoc-D-Bhg-Leu-Asn-lie-lie-Trp; Fmoc-D-Bhg-Leu-Glu-lie-Ile-Trp; Fmoc-D-Bhg-Leu-Gln-Ile-Ile-Trp; Fmoc-D-Bhg-Leu-Tyr-lie-lie-Trp; Fmoc-D-Bhg-Leu-Asp-Val-Ile-Trp; Fmoc-D-Bhg-Leu-Asp-Ile-Val-Trp; Fmoc - D - Bhg - Leu - Asp - Chx-II e - Trp; Fmoc -D - Bhg - Arg - Asp- Chx- lie - Trp; Fmoc - D - Bhg - Ly s - Asp - Chx -11 e - Trp; Fmoc-D-Bhg-Orn-Asp-Chx-lie-Trp ; Fmoc - D - Bhg - Asp - Asp - Chx -11 e - Trp; Fmoc-D-Bhg-Glu-Asp-Chx-lie-Trp; Fmoc-D-Bhg-Leu-Asp-lie-Chx-Trp; -37- Firio c - D - Bhg - Arg - Asp -lie- Chx - Trp ; Fmoc-D-Bhg-Lys-Asp-lie-Chx-Trp; Fmoc-D-Bhg-Orn-Asp-lie-Chx-Trp; Fmoc - D - Bhg - Asp - Asp - II e - Chx - Trp; Fmoc-D-Bhg-Glu-Asp-lie-Chx-Trp; Ac-D-Bheg-Leu-Asp-Ile-Ile-Trp; Ac-D-Bheg-Orn-Asp-lie-Ile-Trp; Ac-D-Bheg-Lys-Asp-lie-lie-Trp; Ac-D-Bheg-Asp-Asp-lie-lie-Trp; Ac-D-Bheg-Glu-Asp-lie-lie-Trp; Ac-D-Bheg-Phe-Asp-Ile-Ile-Trp; Ac-D-Bheg-Arg-Asp-lie-lie-Trp; Ac-D-Bheg-Asp lie-Ile-Trp; Fmoc-D-Bheg-Leu-Asp-lie-lie-Trp Fmoc-D-Bheg-Orn-Asp-lie-Ile-Trp Fmoc-D-Bheg-Lys-Asp-lie-lie-Trp Fmoc-D-Bheg-Asp-Asp-lie-IIe-Trp Fmoc-D-Bheg-Glu-Asp-lie-lie-Trp Fmoc-D-Bheg-Phe-Asp-lie-IIe-Trp Fmoc-D-Bheg-Arg-Asp-lie-lie-Trp Fmoc-D-Bheg-Asp-lie-lie-Trp; Ac-D-Bheg-Leu-Phe-lie-Ile-Trp; Ac-D-Bheg-Leu-Asn-Ile-Ile-Trp; Ac-D-Bheg-Leu-Glu-Ile-Ile-Trp; Ac-D-Bheg-Leu-Gln-Ile-Ile-Trp; Ac-D-Bheg-Leu-Tyr-lie-Ile-Trp; Ac-D-Bheg-Leu-l-Nal-lie-Ile-Trp Ac-D-Bheg-Leu-2-Nal-lie-Ile-Trp Ac-D-Bheg-Leu-Trp-lie-Ile-Trp; Ac-D-Bheg-Leu-Asp-Val-lie-Trp; Ac-D-Bheg-Leu-Asp-Ile-Val-Trp; Ac-D-Bheg-Leu-Asp-Chx-I1e-Trp; Ac-D-Bheg-Leu-Asp-lie-Chx-Trp; Ac-D-Bheg-Arg-Asp-lie-Chx-Trp; Ac-D-Bheg-Lys-Asp-lie-Chx-Trp; Ac-D-Bheg-Orn-Asp-Ile-Chx-Trp; Ac-D-Bheg-Asp-Asp-I1e-Chx-Trp; |WO 93/21219 PCT/US93/03658 -38- Ac-D-Bheg-Glu-Asp-Ile-Chx-Trp; Fmoc-D-Bheg-Leu-Phe-IIe-I1e-Trp; Fmoc-D-Bheg-Leu-Asn-lie-Ile-Trp; Fmoc-D-Bheg-Leu-Glu-lie-lie-Trp ; 5 Fmoc-D-Bheg-Leu-Gln-lie-Ile-Trp; Fmoc-D-Bheg-Leu-Tyr-lie-lie-Trp; Fmoc-D-Bheg-Leu-Asp-Val-Ile-Trp; Fmo c - D - Bheg - Leu - Asp - II e - Val - Trp; Fmoc-D-Bheg-Leu-Asp-Chx-Ile-Trp; 10 Fmoc-D-Bheg-Arg-Asp-Chx- Ile-Trp; Fmo c - D - Bheg - Ly s - Asp - Chx -11 e - Trp; Fmoc-D-Bheg-Om-Asp-Chx- Ile-Trp; Fmoc - D - Bheg - Asp-Asp - Chx- lie - Trp; Fmoc-D-Bheg-Glu-Asp-Chx-Ile-Trp; 15 Fmoc-D-Bheg-Leu-Asp-Ile-CJix-Trp; Fmoc-D-Bheg-Arg-Asp-lie-Chx-Trp; Fmoc-D-Bheg-Lys-Asp-IIe-Chx-Trp; Fmo c - D - Bheg - Orn - Asp - II e - Chx - Trp; Fmoc - D - Bheg - Asp-Asp - lie - Chx-Trp; 20 Fmoc-D-Bheg-Glu-Asp-lie-Chx-Trp; Ac-D-Txg-Leu-Asp-lie-lie-Trp; Ac-D-Txg-Orn-Asp-lie-lie-Trp; Ac-D-Txg-Lys-Asp-lie-lie-Trp; Ac-D-Txg-Asp-Asp-lie-lie-Trp; 25 Ac-D-Txg-Glu-Asp-Ile-Ile-Trp; Ac-D-Txg- Phe -As£>-lie-lie-Trp; Ac-D-Txg-Arg-Asp-Ile-Ile-Trp; Ac-D-Txg-Asp-lie-Ile-Trp; Fmoc-D-Txg-Leu-Asp-lie-lie-Trp; 30 Fmoc-D-Txg-Orn-Asp-lie-lie-Trp; Fmoc-D-Txg-Lys-Asp-lie-lie-Trp; Fmoc-D-Txg-Asp-Asp-lie-lie-Trp; Fmoc-D-Txg-Glu-Asp-lie-Ile-Trp;. Fmoc-D-Txg-Phe-Asp-lie-lie-Trp; 35 Fmoc-D-Txg-Arg-Asp-lie-Ile-Trp; Fmoc-D-Txg-Asp-lie-lie-Trp; Ac-D-Txg-Leu-Phe-lie-lie-Trp; 93/21219 5 10 15 20 25 30 PCT/US93/036S8 -39- Ac-D-Txg-Leu-Asn-lie-lie-Trp; Ac-D-Txg-Leu-Glu-lie-lie-Trp; Ac-D-Txg-Leu-Gin-lie-lie-Trp; Ac-D-Txg-Leu-Tyr-Ile-Ile-Trp; Ac-D-Txg-Leu-1-Nal-Ile-Ile-Trp; Ac-D-Txg-Leu-2-Nal-lie-lie-Trp; Ac-D-Txg-Leu-Trp-Ile-Ile-Trp; Ac-D-Txg-Leu-Asp-Val-Ile-Trp; Ac-D-Txg-Leu-Asp-Ile-Val-Trp; Ac-D-Txg-Leu-Asp-Chx-lie-Trp; Ac-D-Txg-Leu-Asp-lie-Chx-Trp; Ac-D-Txg-Arg-Asp-lie-Chx-Trp; Ac-D-Txg-Lys-Asp-Ile-Chx-Trp; Ac-D-Txg-Orn-Asp-lie-Chx-Trp; Ac-D-Txg-Asp-Asp-lie-Chx-Trp; Ac-D-Txg-Glu-Asp-lie-Chx-Trp; Fmoc-D-Txg-Leu-Phe-lie-lie-Trp; Fmoc-D-Txg-Leu-Asn-lie-lie-Trp; Fmoc-D-Txg-Leu-Glu-lie-lie-Trp; Fmoc-D-Txg-Leu-Gin-Ile-Ile-Trp; Fmoc-D-Txg-Leu-Tyr-lie-lie-Trp; Fmoc-D-Txg-Leu-Asp-Val-lie-Trp; Fmoc-D-Txg-Leu-Asp-lie-Val-Trp; Fmoc-D-Txg-Leu-Asp-Chx-lie-Trp ; Fmoc-D-Txg-Arg-Asp-Chx-lie-Trp ; Fmoc-D-Txg-Lys-Asp-Chx-lie-Trp ; Fmoc-D-Txg-Orn-Asp-Chx-lie-Trp; Fmoc-D-Txg-Asp-Asp-Chx-lie-Trp; Fmoc-D-Txg-Glu-Asp-Chx-lie-Trp; Fmoc-D-Txg-Leu-Asp-lie-Chx-Trp; Fmoc-D-Txg-Arg-Asp-lie-Chx-Trp; Fmoc - D - Txg - Lys - Asp - lie - Chx - Trp ; Fmoc-D-Txg-Orn-Asp-lie-Chx-Trp ; Fmoc-D-Txg-Asp-Asp-lie-Chx-Trp ; Fmoc-D - Txg - Glu-Asp - lie - Chx-Trp; Et-D-Bhg-Leu-Asp-lie-lie-Trp; Bz-D-Bhg-Leu-Asp-Ile-Ile-Trp; O 93/21219 PCT/US93/03658 -40- Pya-D-Bhg-Leu-Asp-Ile-Ile-Trp; Cxi-D-Bhg-Leu-Asp-Ile-Ile-Trp; Ada-D-Bhg-Leu-Asp-lie-Ile-Trp; Cxi(U)-D-Bhg-Leu-Asp-lie-Ile-Trp; 5 Me (U)-D-Bhg-Leu-Asp-lie-Ile-Trp; tBu-D-Bhg-Leu-Asp-lie-Ile-Trp; CF3CO-D-Bhg-Leu-Asp-lie-Ile-Trp; Et-D-Bheg-Leu-Asp-lie-Ile-Trp; Bz-D-Bheg-Leu-Asp-lie-Ile-Trp; 10 Pya-D-Bheg-Leu-Asp-lie-Ile-Trp; Cxi-D-Bheg-Leu-Asp-lie-Ile-Trp; Ada-D-Bheg-Leu-Asp-lie-lie-Trp; Cxi(U)-D-Bheg-Leu-Asp-lie-Ile-Trp; Me(U)-D-Bheg-Leu-Asp-lie-lie-Trp; 15 tBu-D-Bheg-Leu-Asp-Ile-Ile-Trp; CF3CO-D-Bheg-Leu-Asp-lie-Ile-Trp; Ac-D-Bhg-Leu-Asp-Phe-Ile-Trp; Ac-D-Bhg-Orn-Asp-Phe-Ile-Trp; Ac-D-Bhg-Lys-Asp-Phe-Ile-Trp; 20 Ac-D-Bhg-Asp-Asp-Phe-Ile-Trp; Ac-D-Bhg-Glu-Asp-Phe-Ile-Trp; Ac-D-Bhg-Phe-Asp-Phe-Ile-Trp; Ac-D-Bhg-Arg-Asp-Phe-lie-Trp; Ac-D-Bheg-Leu-Asp-Phe-Ile-Trp; 25 Ac-D-Bheg-Orn-Asp-Phe-Ile-Trp; Ac-D-Bheg-Lys-Asp-Phe-Ile-Trp; Ac-D-Bheg-Asp-Asp-Phe-Ile-Trp; Ac-D-Bheg-Glu-Asp-Phe-Ile-Trp; Ac-D-Bheg-Phe-Asp-Phe-Ile-Trp; and 30 Ac-D-Bheg-Arg-Asp-Phe-Ile-Trp; or a phannaceutically acceptable acid or base addition salt thereof. The compounds of Formula I are valuable antagonists of endothelin. The t sts employed 35 indicate that compounds of Formula I possess endothelin antagonist activity. Thus, the compounds of Formula I were tested for their ability to inhibit kWO 93/21219 PCT/US93/03658 -41- [125I] -ET-l ([125I] -Endothelin-1) binding in a receptor assay according to the following procedures: ENDOTHELIN RECEPTOR BINDING ASSAY-A (ERBA-A) 5 INTACT CELL BINDING OP [125I] -ET-1 Materials and Terms Used; Cells The cells used were rabbit renal artery vascular 10 smooth muscle cells grown in a 48-well dish (1 cm2) (confluent cells). Growth Media The growth media was Dulbecco's Modified 15 Eagles/Ham's F12 which contained 10% fetal bovine serum and antibiotics (penicillin/streptomycin/ fungizone). Assay Buffer 20 The assay buffer was a medium 199 containing Hanks salts and 25 mM Hepes buffer (Gibco 380-2350AJ), supplemented with penicillin/streptomycin/fungizone (0.5%) and bovine serum albumin (l mg/mL). 25 [125IJ -ET-1 Amersham radioiodinated endothelin-1 [125I] -ET-1 was used at final concentration of 20,000 cpm/0.25 mL (25 pM). 30 Protocol First, add 0.5 mL warm assay buffer (described above) to the aspirated growth media and preincubate for 2 to 3 hours in a 37°C water bath (do not put back in the 5% carbon dioxide). Second, remove the assay 35 buffers, place the dish on ice, and add 150 fil> of cold assay buffer described above to each well. Third, add 50 mL each of cold [125I]-ET-1 and competing ligand to €T" O 93/21219 PCT/US93/03658 -42- the solution (at the same time if possible). Next, place dish in a 37°C water bath for about 2 hours and gently agitate the dish every 15 minutes. Discard the radioactive incubation mixture in the sink and wash 5 wells 3 times with 1 mL of cold phosphate buffered saline. Last, add 250 mL of 0.25 molar sodium hydroxide, agitate for 1 hour on a rotator, and then transfer the sodium hydroxide extract to gamma counting tubes and count the radioactivity. 10 ENDOTHELIN RECEPTOR BINDING ASSAY-B (ERBA-B) [125I] -ET-1 BINDING IN RAT CEREBELLAR MEMBRANES Materials and Terms Used: 15 Tissue Buffer The tissue is made up of 20 znM tris (hydroxy-methyl)aminomethane hydrochloride (Trizma) buffer, 2 mM ethylenediaminetetra acetate, 100 pM phenylmethylsulfonyl fluoride. 20 Tissue Preparation First, thaw one aliquot of frozen rat cerebellar membranes (2 mg protein in 0.5 mL) . Next, add 0.5 mL membrane aliquot to 4.5 mL cold tissue buffer, 25 polytron at 7,500 revolutions per minute for 10 seconds. Finally, dilute tissue suspension 1/100 (0.1 mL suspension + 9.9 mL tissue buffer), polytron again, and place ice. 30 Dilution Buffer Medium 199 with Hank's salts plus 25 mM Hepes + l mg/mL bovine serum albumin. 35 [12SI] -ET-1 Amersham [125I]-ET-1 (aliquots of 2 x 106 cpm per 100 mL aliquot of [125I]-ET-1 with 5.2 mL dilution ^VO 93/21219 PCT/US93/03658 -43- buffer, place on ice until use (final concentration will be 20,000 cpm per tube, or 25 pM). Protocol 5 Add 50 fih each of cold [12S]-ET-1 and competing ligand to tubes on ice. Mix in 150 pL of tissue to each tube, vortex briefly, then tap to force all liquids to bottom (total assay volume = 250 /*L) . Then place the tubes in a 37°C water bath for 2 hours. 10 Add 2.5 mL cold wash buffer (50 mM Trizma buffer) to each tube, filter, and then wash tube with additional 2.5 mL wash buffer and add to filter. Finally, wash filters with an additional 2.5 mL of cold wash buffer. 15 Count filters for radioactivity in gamma counter. IK VITRO INHIBITION OF ET-1 STIMULATED ARACHIDONIC ACID RELEASE (AAR) IN CULTURED RABBIT VASCULAR SMOOTH 20 MUSCLE CELLS BY COMPOUNDS OF FORMULA I Antagonist activity is measured by the ability of added compounds to reduce endothelin-stimulated arachidonic acid release in cultured vascular smooth 25 muscle cells as arachidonic acid release (AAR). [3H] Arachidonic Acid Loading Media (LM) is DME/F12 + 0.5% FCS x 0.25 mCi/mL [3H] arachidonic acid (Amersham). Confluent monolayers of cultured rabbit renal artery vascular smooth muscle cells were 30 incubated in 0.5 mL of the LM over 18 hours, at 37°C, in 5% C02. The LM was aspirated and the cells were washed once with the assay buffer (Hank's BSS + 10 iriM HEPES + fatty acid-free BSA (1 mg/mL)), and incubated for 5 minutes with 1 mL of the prewarmed assay buffer. 35 This solution was aspirated, followed by an additional l mL of prewarmed assay buffer, and further incubated for another 5 minutes. A final 5-minute incubation #vo 93/21219 PCT/US93/03658 -44- was carried out in a similar manner. The same procedure was repeated with the inclusion o£ 10 fiL of the test compound (l nM to 1 #iM) and 10 ph ET-1 (0.3 nM) and the incubation was extended for 5 30 minutes. This solution was then collected, 10 fih of scintillation cocktail was added, and the amount of [3H] arachidonic acid was determined in a liquid scintillation counter. 10 IN VITRO ANTAGONISM OF ET-1 STIMULATED VASOCONSTRICTION IN TEE RABBIT FEMORAL ARTERY (ETA) AND SARAFOTOXIN 6c STIMULATED VASOCONSTRICTION IN TEE RABBIT PULMONARY ARTERY (ETb) 15 Male New Zealand rabbits were killed by cervical dislocation and exsanguination. Femoral and pulmonary arteries were isolated, cleaned of connective tissue, and cut into 4-mm rings. The endothelium was denuded 20 by placing the rings over hypodermic tubing (32 guage for femoral rings and 28 guage for pulmonary rings. Small Parts, Inc, Miami, Florida) and gently rolling them. Denuded rings were mounted in 20 mL organ baths containing Krebs-bicarbonate buffer (composition in 25 mM: NaCl, 118.2; NaHC03, 24.8; KC1, 4.6; MgS04 7-H20, 1.2; KH2P04, 1.2; CaCl2-2H20; Ca-Na2 EDTA, 0.026; dextrose, 10.0), that was maintained at 37°C and gassed continuously with 5% C02 in oxygen (pH 7.4). Resting tension was adjusted to 3.0 g for 30 femoral and 4.0 g pulmonary arteries; the rings were left for 90 minutes to equilibrate. Vascular rings were tested for lack of functional endothelium (i.e., lack of an endothelium-dependent relaxation response to carbachol (1.0 ftM) in norepinephrine (0.03 /iM) 35 contracted rings. Agonist peptides, ET-l (femoral), and S6c (pulmonary), were cumulatively added at 10-minute intervals. The ET antagonists were added O 93/21219 PCT/US93/03658 -45- 30 minutes prior to adding the agonist and pA 2 values were calculated (Table I). The data in Table I below show the endothelin receptor binding and antagonist activity of 5 representative compounds of Formula I. • • • • TABLE I. Biological Activity of Compounds of Formula I Example Number Compound ERBA-A IC50 (fOi) ERBA-B ICso IflM) aar IC50 (/iM) Femoral pAj Pulmonary 1 Ac-D-Bhg-Leu-Asp-lie-Ile-Trp 0.0026 0.019 0.0049 2 D-Bhg-Leu-Asp-lie-Ile-Trp 0.45 2.1 0.10 3 L-Bhg-Leu-Asp-lie-Ile-Trp 2.5 3.0 2.36 4 Ac-L-Bhg-Leu-Asp-Ile-Ile-Trp 0.56 h 0 0.56 5 Ac-D-Txg-Leu - Asp-11e-11e-Trp 0.018 co h o 6 Ac-D-Bheg-Leu-Asp-Ile-Ile-Trp 0.005 0.019 0.003 6.8 7.4 7 Ac-D-Bhg-Orn-Asp-Ile-Ile-Trp 0.022 1.5 0.037 6.5 8 Ac-D-Bhg-Glu-Aap-lle-Ile-Trp 0.0055 0.022 0.007 6.5 7.0 9 Ac-D-Bhg-Leu-Asp-Ile-Ile-Trp, 2Na+ 0.004 0.015 0.0049 6.9 7.1 ^VO 93/21219 PCT/US93/03658 -47- General Method for Preparing Compounds of Formula I The compounds of Formula I may be prepared by solid phase peptide synthesis on a peptide synthesizer, for example, an Applied Biosystems 430A 5 peptide synthesizer using activated esters or anhydrides of N-alpha-Boc protected amino acids, on PAM or MBHA resins. Additionally, the compounds of Formula I may also be prepared by conventional solution peptide synthesis. Amino acid side chains 10 are protected as follows: Bzl(Asp, Glu, Ser), 2-C1-Z(Lys), 2-Br-Z(Tyr), Bom(His), For(Trp), and MeBzl(Cys). Each peptide resin (1.0 g) is cleaved with 9 mL of HF and l mL of anisole or p-cresol as a scavenger (60 minutes, 0°C). The peptide resin is 15 washed with cyclohexane, extracted with 30% aqueous HOAc, followed by glacial HOAc, concentrated under reduced pressure, and lyophilized. (A peptide containing For (Trp) is dissolved in 0°C,. the pH is adjusted to 12.5 with IN KOH (2 minutes), neutralized 20 with glacial HOAc, desalted on C18 (as described below), and lyophilized. The crude peptide is purified by preparative reversed phase high performance liquid chromatography (RP-HPLC) on a C18 column (2.2 x 25.0 cm, 15.0 mL/min) with a linear 25 gradient of 0.1% TFA in water to 0.1% TFA in acetonitrile and lyophilized. The homogeneity and composition of the resulting peptide is verified by RP-HPLC, capillary electrophoresis, thin layer chromatography (TLC), proton nuclear magnetic 30 resonance spectrometry (NMR), and fast atom bombardment mass spectrometry (FAB-MS). The compounds of the present invention can be prepared and administered in a wide variety of oral and parenteral dosage forms. Thus, the compounds of 35 the present invention can be administered by injection, that is, intravenously, intramuscularly, intracutaneously, siibcutaneously, intraduodenally, or £wO 93/21219 PCT/US93/03658 -48- intraperitoneally. Also, the compounds of the present invention can be administered by inhalation, for example, intranasally. Additionally, the compounds of the present invention can be administered 5 transdermally. It will be obvious to those skilled in the art that the following dosage forms may comprise as the active component, either a compound of Formula I or a corresponding phannaceutically acceptable salt of a compound of Formula I. 10 For preparing pharmaceutical compositions from the compounds of the present invention, phannaceutically acceptable carriers can be either solid or liquid. Solid form preparations include powders,, tablets, pills, capsules, cachets, 15 suppositories, and dispersible granules. A solid carrier can be one or more substances which may also act as diluents, flavoring agents, binders, preservatives, tablet disintegrating agents, or an encapsulating material. 20 In powders, the carrier is a finely divided solid which is in a mixture with the finely divided active component. In tablets, the active component is mixed with the carrier having the necessary binding properties in 25 suitable proportions and compacted in the shape and size desired. The powders and tablets preferably contain from five or ten to about seventy percent of the active compound. Suitable carriers are magnesium carbonate, 3 0 magnesium stearate, talc, sugar, lactose, pectin., dextrin, starch, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose, a low melting wax, cocoa butter, and the like. The term "preparation" is intended to include the formulation of the active 35 compound with encapsulating material as a carrier providing a capsule in which the active component with or without other carriers, is surrounded by a carrier, 0WO 93/21219 PCT/US93/03658 -49- which is thus in association with it. Similarly, cachets and lozenges are included. Tablets, powders, capsules, pills, cachets, and lozenges can be used as solid dosage forms suitable for oral administration. 5 For preparing suppositories, a low melting wax, such as a mixture of fatty acid glycerides or cocoa butter, is first melted and the active component is dispersed homogeneously therein, as by stirring. The molten homogenous mixture is then poured into 10 convenient sized molds, allowed to cool, and thereby to solidify. Liquid form preparations include solutions, suspensions, and emulsions, for example, water or water propylene glycol solutions. For parenteral 15 injection liquid preparations can be formulated in solution in aqueous polyethylene glycol solution. Aqueous solutions suitable for oral use can be prepared by dissolving the active component in water and adding suitable colorants, flavors, stabilizing 20 and thickening agents as desired. Aqueous suspensions suitable for oral use can be made by dispersing the finely divided active component in water with viscous material, such as natural or synthetic gums, resins, methylcellulose, sodium 25 carboxymethylcellulose, and other well-known suspending agents. Also included are solid* form preparations which are intended to be converted, shortly before use, to liquid form preparations for oral administration. 30 Such liquid forms include solutions, suspensions, and emulsions. These preparations may contain, in addition to tne active component, colorants, flavors, stabilizers, buffers, artificial and natural sweeteners, dispersants, thickeners, solubilizing 35 agents, and the like. The pharmaceutical preparation is preferably in unit dosage form. In such form the preparation is 2 5 2855 -50- subdivided into unit doses containing appropriate quantities of the active component. The unit dosage form can be a packaged preparation, the package containing discrete quantities of preparation, such as packeted tablets, capsules, and powders in vials or ampoules. Also, the unit dosage form can be a capsules, tablet, cachet, or lozenge itself, or it can be the appropriate number of any of these in packaged form. The quantity of active component in a unit dose preparation may be varied or adjusted from 0.1 mg to 100 mg preferably 0.5 mg to 100 mg according to the particular application and the potency of the active component. The composition can, if desired, also co/ .ain other compatible therapeutic agents. In therapeutic use as antagonist of endothelin, the compounds of this invention may be administered at the initial dosage of about 0.01 mg to about 20 mg per kilogram daily. A daily dose range of about 0.01 mg to about 10 mg per kilogram is preferred. The dosages, however, may be varied depending upon the requirements of the patient, the severity of the condition being treated, and the compound being employed. Determination of the proper dosage for a particular situation is within the skill of the art. Generally, treatment is initiated with smaller dosages which are less than the optimum dose of the compound. Thereafter, the dosage is increased by small increments until the optimum effect under the circumstances is reached. For convenience, the total dally dosage may be divided and administered in portions during the day, if desired. The following nonlimiting examples illustrate the inventors' preferred methods for preparing the compounds of the invention. kWO 93/21219 PCT/US93/03658 -51- EXAMPLE 1 Ac-D-Bhg-Leu-Asp-lie-Ile-Trp The linear hexapeptide is prepared by standard solid phase synthetic peptide methodology utilizing a 5 Boc/benzyl strategy (Stewart, J. M. and Young, J. D., Solid Phase Peptide Synthesis. Pierce Chemical Co., Rockford, IL, 1984). All protected amino acids and reagents are obtained from commercial sources with the exception of N-a-Boc-DL-Bhg and are not further 10 purified. The protected peptide resin is prepared on an Applied Biosysterns 43OA Peptide Synthesizer, utilizing protocols supplied for a dicyclohexyl-carbodiimide-mediated coupling scheme (Standard 1.0, Version 1.40) . Starting with 0.710 g of 15 N-a-Boc-Trp-PAM resin (0.70 meq/g, 0.497 meq of Boc-Trp(For) total) the protected peptide is prepared by the stepwise coupling of the following amino acids (in order of addition): N-a-Boc-Ile-0.5H20, N-a-Boc-Ile-0.5H20, N-a-Boc-Asp(Bzl), N-a-Boc-Leu*H20, 20 and N-a-Boc-DL-Bhg. A typical cycle for the coupling of an individual amino acid residue is illustrated below (reproduced from the ABI manual): All the single couple RV cycles conform to the following pattern: 25 1) 33% TFA in DCM for 80 seconds 2) 50% TFA in DCM for 18 .5 minutes 3) Three DCM washes 4) 10% DIEA in DMF for 1 minute 5) 10% DIEA in DMF for 1 minute 30 6) Five DMF washes 7) Coupling period 8) Five DCM washes After the coupling of N-a-Boc-DL-Bhg, the Boc 35 group is removed with the end-NH2 cycle (1.012 g) . The peptide is liberated from the solid support, and the carboxylate of aspartic acid deprotected by 0YO 93/21219 PCT/US93/03658 -52- treatment with anhydrous hydrogen fluoride (9.0 mL), anisole (0.5 mL), and dimethyl sulfide (0.5 mL) (60 minutes, 0°C). After removing the hydrogen fluoride under a stream of nitrogen, the resin is 5 washed with diethyl ether (3 x 30 mL) and extracted with 20% HOAc in water (3 x 30 mL) and glacial HOAc (2 x 30 mL). The aqueous extractions are combined, concentrated under reduced pressure, and lyophilized (360 mg). The crude peptide is dissolved in 4.0 mL of 10 50% TFA/H20, filtered through a 0.4 L syringe filter, and chromatographed on a Vydac 218TP 1022 column (2.2 x 25.0 cm, 15.0 mL/min, A: 0.1% TFA/H20, B: 0.1% TFA/CH3CN, Gradient; 0% B for 10 minutes, 10% to 40% B over 120 minutes). Two individual fractions 15 are collected and combined based upon analysis by analytical HPLC. The combined fractions are concentrated separately under reduced pressure (10 mL) , diluted with H20 (50 mL) , and lyophilized (40.0 mg/ea). Separation into the two diastereomers 20 (Isomers A and B) is effected under these conditions (tR = Isomer A 15.63 min., Isomer B 16.79 min.). The late running peak fractions (Isomer B) are repurified under the same experimental conditions with a gradient of 30% to 50% B over 120 minutes at 15 mL/min to 25 afford purified product. Acetylation is carried out with 20 mg of Isomer B in 90% acetic acid followed by addition of acetic anhydride (5 mL) and stirring overnight. After evaporation and drying the product Ac-D-Bhg-Leu-Asp-Ile-Ile-Trp is 99% pure by HPLC. 30 [Vydac 218 TP 1022 column (2.2 x 25.0 cm, 15.0 mL/min. A: 0.1% TFA/CH3CN, Gradient 20% to 86% B over 22 min.)] tR = 18.66 minutes. The homogeneity and structure of the resulting peptide is confirmed by analytical HPLC. Proton Nuclear Magnetic Resonance 35 Spectroscopy (H1-NMR) and Fast Atom Bombardment Mass Spectroscopy (FAB-MS), M+Na 972.0, M+2Na+ 995.9. O 93/21219 PCT/US93/03658 10 -53- In a process analogous to Example 1 using the appropriate amino acids, the corresponding compounds of Formula I are prepared as follows: EXAMPLE 2 D-Bhg-Leu-Asp-lie-Ile-Trp: FAB-MS, M+l 907.4. EXAMPLE 3 L-Bhq-Leu-Asp-lie-Ile-Trp; FAB-MS, M+l 907.4. EXAMPLE 4 Ac-L-Bhq-Leu-Asp-Ile-Ile-Trp: FAB-MS, M+l 950.0. EXAMPLE 5 15 Ac-D-Txa-Leu-Asp-lie-Ile-Trp; FAB-MS, M+Na 977.0. EXAMPLE 6 Ac-D-Bheg-Leu-Asp-lie-Ile-Trp; FAB-MS, M+l 970.3. 20 EXAMPLE 7 Ac-D-Bhg-Om-Asp-Ile-Ile-Trp; FAB-MS. M+l 951.2. EXAMPLE 8 Ac-D-Bhg-Glu-Asp-lie-lie-Trp; FAB-MS, M+Na 988.8. 25 EXAMPLE 9 Disodium salt of Ac-D-Bhg-Leu-Asp-Ile-Ile-Trp A saturated solution of sodium bicarbonate in water is prepared, diluted with water (1:10), chilled 30 to 0°C, and 10 mL of the solution is added to approximately 50 mg of Ac-D-Bhg-Leu-Asp-lie-Ile-Trp (Example 1) with stirring. The pH of the solution is greater than 9. After 10 minutes, the solution is passed through a C18 cartridge, washed with water 35 (100 mL), and the absorbed peptide is eluted with methanol (50 mL), concentrated under reduced pressure, </div>

Claims

<div class="application article clearfix printTableText" id="claims"> •* O93/21219 PCT/US93/03658 -54- resuspended in water (50 mL), and lyophilized (three times) to give the title compound. Ac-D-Bhg-Leu-Asp-Ile-Ile-Trp. disodium salt: FAB-MS, M+l 950.4, M+Na 972.1, M+2Na 994.3. 5 EXAMPLE 10 Boc-Bhq Bhg-HCl (1.70 g, 5.43 mmol) is suspended in 150 mL of p-dioxane:H20 (2:1) at room temperature. To 10 the stirred solution is added 1.40 g (6.42 mmol) of di-tert butyldicarbonate. The pH of the solution is adjusted to >9.0 with IN NaOH and maintained at between pH 9 and 10 with aliquot additions of IN NaOH, until the pH is constant. The solution is 15 concentrated under reduced pressure t ,< approximately 75 mL, overlain with ethyl acetate (50 mL), and acidified to approximately pH 2.5 with 10% aqueous HC1. The organic layer is separated, washed successively with 10% aqueous HC1 (2 x 50 mL), brine 20 (2 x 50 mL), H20 (3 x 50 mL), and dried with MgS04. The solution is filtered, concentrated under reduced pressure, and the oil is recrystallized from ethyl acetd.ce: heptane (1.82 g) . The white solid is characterized by proton NMR, fast atom bombardment 25 mass spectrometry (M+l=368), and elemental analysis. WO 93/21219 PCT/US93/03658 -55-CLAIMS A compound of Formula I AA1-AA2 - AA3 - AA4 - AA5 - AA6 wherein AA1 is 5 wherein R is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, 10 cycloalkylalkyl, aryl, heteroaryl, fluorenylmethyl, 2 2 3 -N-R , wherein R and R are each the same or 15 | R different and each is hydrogen, alkyl, alkenyl, 20 alkynyl, ; cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, •- r ■1 O 93/21219 PCT/US93/03658 -56- 25 heteroaryl, or fluorenylmethyl, 0 II 2 2 -C-OR , wherein R is as defined above, 30 -OR2, wherein R2 is as defined above, O 'I 3 2 3 -N-C-N-R , wherein Rz and R-* are as defined 35 R* R2 above' o II 9 -C-C(R')3, wherein R' is F, CI, Br, or I, - CH2 - OR2, wherein R2 is as defined above, 40 O « 3 -N-C-R , R2a 45 wherein R2a is hydrogen or alkyl and R3 is as defined above, O 50 55 -N-C-OR3, J»2a R wherein R2a and R3 are as defined above excluding R3 is hydrogen, or O I' 2 -C-R , wherein R^ is as defined above, R1 is hydrogen or alkyl, Z is -0-, 60 ~S(0)A, i wherein m is zero or an integer of 1 or 2, -N-, wherein R2 is as defined above, l2' 65 R -(CH2)n-, wherein n is zero or an integer of 1, 2, 3, or 4, H'O 93/21219 PCT/US93/03658 -57- - (CH2)n-CH=CH- (CH2)n-, wherein n is as defined above, 70 O 95 -C-' -CR1-, wherein R1 and R2 are as defined 1 2 75 OR above, or R2 I -C- , l3 80 RJ wherein R2 and R3 are each the same or different and each is as defined above, X and Y are the same and substituted at the same position on the aromatic ring and each may 85 be one, two, three, or four substituents selected from the group consisting of hydrogen, halogen, alkyl, 90 -C02R2, wherein R2 is as defined above, 2 2 3 -CONR , wherein R and R are as defined R3 above, 2 .23 -NR , wherein R and R are as defined l3 R above, or nitro or wherein R, Z, X, and Y are as defined above; O 93/21219 PCT/US93/03658 ■58- AA2 is R1 0 I* II —N—C—C- L I R (CH2)n • A 100 wherein R4 is hydrogen, alkyl, alkenyl, alkynyl, 105 cycloalkyl, aryl, heteroaryl, -N-R3b, 110 R2b wherein R and R are each the same or different and each is hydrogen, alkyl, 115 cycloalkyl, aryl, or heteroaryl, -OR2b, wherein R2b is as defined above, O i2b aw wherein R and R are each 125 the same or different and each is as defined above for R2b and R3b, 0 " • 2h 9b 130 -C-R , wherein R is as defined above, -59- 25 2 8 55 NH -NH-C-NH-R2b wherein R2b is as defined 135 above, or 0 -C-0R2b, wherein R2b is as defined above, and 140 R1 and n are as defined above, or AA2 is absent; AA3 is R1 O I* II —N—C-C— I I R1 (CH2)n R5 wherein R5 is hydrogen, 145 alkyl, arylf heteroaryl, O 150 -C-N-R3b, i2b wherein R2b and R3b are each the same or 155 different and each is as O defined above, -C-R2b, wherein R2b is as 160 defined above, or O -C-OR2b, wherein R2b is as defined above, and 165 R1 and n are as defined above, or AA3 is absent; ^vo 93/21219 PCT/ US93/03658 -60- AA4 and AA5 are each independently absent or each is independently R1 O i * II —N—C-C— | i R1 (CH2)n R6 wherein R6 is hydrogen, 170 alkyl, alkenyl, alkynyl, cycloalkyl, aryl, or 175 heteroaryl, and R1 and n are as defined above; AA6 is I R1 180 -N- -C-R8 'i I R1 (CH2) n i R7 185 wherein R7 is aryl or heteroaryl, R8 is O 190 || -C-OR1, wherein R1 is as defined above, -OR1, wherein R1 is as defined above, 195 O ' 1 -C-N-R , wherein RA is as defined above, or ^vo 93/21219 PCT/US93/03658 -61- 200 -CH2-OR1, wherein R1 is as defined above, and R1 and n are as defined above; * stereochemistry at C in AA1, AA2, AA3, AA4, or AA5 is 205 D, L, or DL and * stereochemistry at C in AAe is L; or a phannaceutically acceptable salt thereof. 2. A compound according to Claim 1, in which AA1 is 2 wherein R is -N-R , i3 5 wherein R2 and R3 are each tlie same or different and each is hydrogen, alkyl, alkenyl, 10 alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, 15 heteroaryl, or fluorenylmethyl, 0 1 » • a -N-C-N-R1*, wherein R^ and RJ are as 20 jj2 jj2 defined above, O 93/21219 PCT/LJS93/03658 '3 25 or I, 0 30 40 50 55 -62- -C-C(R9)3, wherein R9 is. F, CI, Br, II 3 3 -NH-C-R , wherexn R is as defxned above, or " 3 -NH-C-OR , 3 wherein R xs as defined above excluding R3 is hydrogen, 35 Z is -O-, wherein rn is zero or an integer of l or 2, -N-, wherein R2 is as defined above, R2 - (CH2)^, wherein n is zero or an integer of 1, 2, 3, or 4, - (CH2) n"CHsCH~ n" ' wherein n is as 45 defined above, ■C-, -CH-, wherein R1 is hydrogen or alkyl, or OR1 R2 I -c-, l3 r" wherein R2 and R3 are each the same or different and each is as defined above and 60 X and Y are the same and substituted at the same position on the aromatic ring and each substituent is selected from the group consisting of hydrogen, O 93/21219 PCI7US93/03658 -63- 65 halogen, or alkyl; AA2 is O II —NH—CH—C— I (CH2)n R4 wherein R4 is hydrogen, alkyl, 7 0 alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, 75 -N-R3b, R.2b wherein R2b and R3b are each the same or different and 80 each is hydrogen, alkyl, cycloalkyl, aryl, or 8 5 heteroaryl, -OR2b, wherein R2b is as defined above. 90 -C-N-R3b,. i2b wherein R2b and R3b are each the same or different and 95 each is as defined above for R2b and R3b, 93/21219 PCT/US93/03658 -64- O || 2b -C-R , wherein R2b is. as defined 100 above, NH -NH-C-NH-R2b wherein R2b is as defined above, or 105 O II 2b 2b -C-OR , wherexn R is as defined above, and n is as defined above or 110 AA2 is absent; AA3 is O II —NH—CH—C— I (CH2)n R5 wherein R5 is aryl, heteroaryl, O || 2b -C-N-R , i2b wherein R and R are each the 120 same or different and each is as defined above, O II 2b oh -C-R , wherein R^D is as defined 125 above, or O II 2b ov. -C-OR , wherein RZD is as defined above, and 130 n is as defined above, or AA3 is absent; ^VO 93/21219 PCT/US93/03658 -65- AA4 and AA5 are each independently absent or each is 0 I! —NH—CH—C— I (CH2)n R6 wherein R® is hydrogen alkyl, 135 alkenyl, alkynyl, cycloalkyl, aryl, or heteroaryl, and 140 n is as defined above; AA6 is -NH-*CH-C0,H 1 <CHa>n R , 145 wherein R7 is aryl or heteroaryl, and n is as defined above, or 150 0 -NH-*CH-C--N-R1 | | wherein R7, R1, and n are as _cc ^?H2^n R defined above, 155 |„ R * 1 2 3 4 5 . stereochemistry at CH in AA , AA , AA , AA , or AA is D,L, or DL, and 160 * 6 stereochemistry at CH in AA is L; or a phannaceutically acceptable salt thereof. kWO 93/21219 PCT/US93/03658 -66- 3. A compound according to Claim 2, in which AA1 is O II R—CH—C— wherein R is -N-R2, i3 2 3 5 wherein R and R are each the same or different and each is hydrogen, alkyl, aryl, or 10 f luorenylmethyl, 0 1 s 2 -N-C-N-R-®, wherein R and RJ are as 15 ^2 defined above, 0 -C-C(R9)3, wherein R9 is F, CI, Br, or I, or 20 0 II 10 - ft -NH-C-OR , wherein R1U is hydrogen, alkyl, aryl, or arylalkyl, excluding R10 25 is hydrogen, Z is -0-, -S-, -NH-, -(CH2)n, wherein n is zero or an 30 integer of 1, 2, 3, or 4, or - (CH2)n-CH=CH- (CH2)ft, wherein n is zero or an integer of 1 or 2 and O 93/21219 PCT/US93/03658 -67- X and Y are each the same and substituted at 35 the same position on the aromatic ring and each substituent is selected from the group consisting of hydrogen, halogen, or 4 0 alkyl; AA2 is 45 55 60 70 O II -NH-CH-C- Wn R4 wherein R4 is hydrogen, 5 0 alkyl, aryl, heteroaryl, ,3b -N-R- J,2b wherein R2b and R3b are each the same or different and each is hydrogen or alkyl, -C-N-R3b ' R2b jjv *3 v\ 65 wherein R and R are each the same or different and each is hydrogen or alkyl, NH -NH-C-NH-R2b wherein R2b is. as defined above, or 93/21219 PCT/US93/03658 75 110 -68-0 II 2b 5b -C-OR , wherein R is as defined above, and n is an integer of 1, 2, 3, or 4 or AA2 is absent; 80 AA3 is O II -NH-CH-C- , I 85 (CH_) | 2 n R5 wherein R5 is aryl, heteroaryl, 90 O II 3b 3b . -C-NHR, wherein R is hydrogen or alkyl, 95 O -C-R2b, wherein R2b is hydrogen or alkyl, or 100 0 II 9b 9b -C-OR , wherein R is hydrogen or alkyl, and 105 n is an integer of 1, 2, 3, or 4; AA4 and AA5 are each independently O -NH-CH-C- I (fI2)n i6 wherein Rs is hydrogen, 115 alkyl, cycloalkyl, or aryl, and (0fr'O 93/21219 PCT/US93/036S8 -69- n is an integer of 1, 2, 3, or 4; AA6 is 120 -NH- CH-COoH I <CH2>n R? 125 wherein R7 is aryl or heteroaryl, and n is zero or an integer of 1, 2, 3, or 4, or 0 130 I -NH-*CH-C--N-R1 | | wherein R7, R1, and n are as ^?H2*n R defined above, 17 135 R * stereochemistry at CH in AA1, AA2, AA3, AA4, or AA5 is D, L, or DL and * 140 stereochemistry at CH in AA® is L; or a phannaceutically acceptable salt thereof. 4. A compound according to Claim 3 selected from the group consisting of: L-Bhg-Leu-Asp-lie-Ile-Trp; D-Bhg-Leu-Asp-lie-Ile-Trp; 5 Ac-L-Bhg-Leu-Asp-Ile-Ile-Trpj- Ac-D-Bhg-Leu-Asp- Ile-Ile-Trp; Ac-D-Bhg-Orn-Asp-Ile-Ile-Trp; Ac-D-Bhg-Lys-Asp-lie-lie-Trp; Ac-D-Bhg-Asp-Asp-lie-Ile-Trp; 10 Ac-D-Bhg-Glu-Asp-Ile-Ile-Trp; Ac-D-Bhg-Phe-Asp-lie-Ile-Trp; Ac-D-Bhg-Arg-Asp-lie-lie-Trp; Ac-D-Bhg-Asp-lie-Ile-Trp; Fmoc-D-Bhg-Leu-Asp-Ile-Ile-Trp; 15 Fmoc-D-Bhg-Orn-Asp-lie-Ile-Trp; 93/21219 20 25 30 35 40 45 PCT/US93/03658 -70- Fmoc-D-Bhg-Lys-Asp-lie-lie-Trp; Fmoc-D-Bhg-Asp-Asp-lie-IIe-Trp; Fmoc-D-Bhg-Glu-Asp-lie-lie-Trp; Fmoc-D-Bhg-Phe-Asp-lie-lie-Trp; Fmc-D-Bhg-Arg-Asp-Ile-Ile-Trp; Fmoc-D-Bhg-Asp-lie-lie-Trp; Ac-D-Bhg-Leu-Phe-lie-Ile-Trp; Ac-D-Bhg-Leu-Asn-lie-Ile-Trp; Ac-D-Bhg-Leu-Glu-lie-Ile-Trp; Ac-D-Bhg-Leu-Gin-lie-Ile-Trp; Ac-D-Bhg-Leu-Tyr-Ile-Ile-Trp; Ac-D-Bhg-Leu-1-Nal-lie-Ile-Trp; Ac-D-Bhg-Leu-2-Nal-lie-lie-Trp; Ac-D-Bhg-Leu-Trp-lie-Ile-Trp; Ac-D-Bhg-Leu-Asp-Val-Ile-Trp; Ac-D-Bhg-Leu-Asp-IIe-Val-Trp; Ac-D-Bhg-Leu-Asp-Chx-Ile-Trp; Ac-D-Bhg-Leu-Asp-lie-Chx-Trp; Ac-D-Bhg-Arg-Asp-lie-Chx-Trp; Ac-D-Bhg-Lys-Asp-lie-Chx-Trp; Ac-D-Bhg-Orn-Asp-lie-Chx-Trp; Ac-D-Bhg-Asp-Asp-lie-Chx-Trp; Ac-D-Bhg-Glu-Asp-lie-Chx-Trp; Fmoc-D-Bhg-Leu-Phe-lie-Ile-Trp ; Fmoc-D-Bhg-Leu-Asn-lie-Ile-Trp; Fmoc-D-Bhg-Leu-Glu-lie-Ile-Trp; Fmoc-D-Bhg-Leu-Gin-lie-Ile-Trp; Fmoc-D-Bhg-Leu-Tyr-lie-Ile-Trp; Fmoc-D-Bhg-Leu-Asp-Val-Ile-Trp; Fmo c - D - Bhg - Leu - Asp - II e - Val - Trp; Fmo c - D - Bhg - Leu - Asp - Chx - II e - Trp; Fmoc - D - Bhg - Arg-Asp-Chx-lie - Trp; Fmoc-D-Bhg-Lys - Asp-Chx-lie-Trp ; Fmoc-D-Bhg-Orn-Asp-Chx- Ile-Trp; Fmoc-D-Bhg-Asp-Asp-Chx-II. -Trp; Fmoc-D-Bhg-Glu-Asp-Chx-Ile-Trp; Fmoc - D - Bhg - Leu - Asp - lie - Chx-Trp; ^WO 93/21219 55 60 65 70 75 80 PCT/US93/03658 -71- Fmoc-D-Bhg-Arg-Asp-lie-Chx-Trp ; Fmoc - D - Bhg - Lys - Asp- lie- Chx- Trp; Fmoc - D - Bhg - Orn-Asp-lie - Chx-Trp; Fmoc - D - Bhg-Asp-Asp-lie - Chx-Trp; Fmoc - D - Bhg - Glu - Asp - lie - Chx-Trp; Ac-D-Bheg-Leu-Asp-lie-Ile-Trp; Ac-D-Bheg-Orn-Asp-Ile-Ile-Trp; Ac-D-Bheg-Lys-Asp-lie-lie-Trp; Ac-D-Bheg-Asp-Asp-lie-lie-Trp; Ac-D-Bheg-Glu-Asp-Ile-Ile-Trp; Ac-D-Bheg-Phe-Asp-Ile-Ile-Trp; Ac-D-Bheg-Arg-Asp-lie-lie-Trp; Ac-D-Bheg-Asp-lie-lie-Trp; Fmoc-D-Bheg-Leu-Asp-lie-Ile-Trp; Fmoc-D-Bheg-Orn-Asp-lie-Ile-Trp; Fmoc-D-Bheg-Lys-Asp-lie-lie-Trp; Fmoc-D-Bheg-Asp-Asp-lie-lie-Trp ; Fmoc-D-Bheg-Glu-Asp-lie-lie-Trp; Fmoc-D-Bheg-Phe-Asp-lie-lie-Trp; Fmoc-D-Bheg-Arg-Asp-lie-Ile-Trp; Fmoc-D-Bheg-Asp-lie-lie-Trp; Ac-D-Bheg-Leu-Phe-lie-Ile-Trp; Ac-D-Bheg-Leu-Asn-Ile-Ile-Trp; Ac-D-Bheg-Leu-Glu-Ile-Ile-Trp; Ac-D-Bheg-Leu-Gln-Ile-Ile-Trp; Ac-D-Bheg-Leu-Tyr-lie-Ile-Trp; Ac-D-Bheg-Leu-1-Nal-lie-Ile-Trp; Ac-D-Bheg-Leu-2-Nal-lie-Ile-Trp; Ac-D-Bheg-Leu-Trp-Ile-Ile-Trp; Ac-D-Bheg-Leu-Asp-Val-lie-Trp; Ac-D-Bheg-Leu-Asp-lie-Val-Trp; Ac-D-Bheg-Leu-Asp-Chx-lie-Trp; Ac-D-Bheg-Leu-Asp-lie-Chx-Trp; Ac-D-Bheg-Arg-Asp-lie-Chx-Trp; Ac-D-Bheg-Lys-Asp-lie-Chx-Trp; Ac - D - Bheg - Orn - Asp - lie - Chx-Trp; Ac-D-Bheg-Asp-Asp-I1e-Chx-Trp; .WO 93/21219 PCI7US93/03658 -72- 90 Ac-D-Bheg-Glu-Asp-lie-Chx-Trp; Fmoc-D-Bheg-Leu-Phe-lie-lie-Trp; Fmoc-D-Bheg-Leu-Asn-lie-lie-Trp; Fmoc-D-Bheg-Leu-Glu-lie-lie-Trp; Fmoc-D-Bheg-Leu-Gin-lie-Ile-Trp; 95 Fmoc-D-Bheg-Leu-Tyr-lie-Ile-Tr^; Fmoc-D-Bheg-Leu-Asp-Val-Ile-Trp; Fmoc-D-Bheg-Leu-Asp-lie-Val-Trp ; Fmoc-D-Bheg-Leu-Asp-Chx-lie-Trp; Fmoc-D-Bheg-Arg-Asp-Chx-lie-Trp; 100 Fmoc-D-Bheg-Lys-Asp-Chx-Ile-Trp; Fmoc-D-Bheg-Orn-Asp-Chx-Ile-Trp; Fmoc-D-Bheg-Asp-Asp-Chx-IIe-Trp; Fmoc-D-Bheg-Glu-Asp-Chx-Ile-Trp; Fmoc-D-Bheg-Leu-Asp-lie-Chx-Trp; 105 Fmoc-D-Bheg-Arg-Asp-lie-Chx-Trp; Fmoc-D-Bheg-Lys-Asp-lie-Chx-Trp; Fmo c - D - Bheg - Orn - Asp -11 e - Chx - Trp; Fmoc - D - Bheg - Asp-Asp - lie - Chx-Trp; Fmo c - D - Bheg - Glu - Asp - II e - Chx - Trp; 110 Ac-D-Txg-Leu-Asp-Ile-Ile-Trp; Ac-D-Txg-Om-Asp-Ile-Ile-Trp; Ac-D-Txg-Lys-Asp-lie-Ile-Trp; Ac-D-Txg-Asp-Asp-lie-lie-Trp; Ac-D-Txg-Glu-Asp-lie-Ile-Trp; 115 Ac-D-Txg-Phe-Asp-Ile-Ile-Trp; Ac-D-Txg-Arg-Asp-Ile-Ile-Trp; Ac-D-Txg-Asp-lie-lie-Trp; Fmoc-D-Txg-Leu-Asp-lie-lie-Trp; Fmoc-D-Txg-Orn-Asp-lie-lie-Trp; 120 Fmoc-D-Txg-Lys-Asp-Ile-Ile-Trp; Fmoc-D-Txg-Asp-Asp-lie-lie-Trp; Fmoc-D-Txg-Glu-Asp-lie-Ile-Trp; Fmoc-D-Txg-Phe-Asp-lie-lie-Trp ; Fmoc-D-Txg-Arg-Asp-lie-Ile-Trp; 125 Fmoc-D-Txg-Asp-Ile-Ile-Trp; Ac-D-Txg-Leu-Phe-lie-lie-Trp; -73- Ac-D-Txg-Leu-Asn-lie-lie-Trp; Ac-D-Txg-Leu-Glu-lie-Ile-Trp; Ac-D-Txg-Leu-Gin-lie-lie-Trp; Ac-D-Txg-Leu-Tyr-Ile-Ile-Trp; Ac-D-Txg-Leu-1-Nal-lie-lie-Trp; Ac-D-Txg-Leu-2-Nal-Ile-Ile-Trp; Ac-D-Txg-Leu-Trp-Ile-Ile-Trp; Ac-D-Txg-Leu-Asp-Val-lie-Trp; Ac-D-Txg-Leu-Asp-lie-Val-Trp; Ac-D-Txg-Leu-Asp-Chx-lie-Trp; Ac-D-Txg-Leu-Asp-lie-Chx-Trp; Ac-D-Txg-Arg-Asp-lie-Chx-Trp ; Ac-D-Txg-Lys-Asp-IIe-Chx-Trp; Ac-D-Txg-Orn-Asp-Ile-Chx-Trp; Ac-D-Txg-Asp-Asp-Ile-Chx-Trp; Ac-D-Txg-Glu-Asp-lie-Chx-Trp; Fmoc-D-Txg-Leu-Phe-lie-lie-Trp Fmoc-D-Txg-Leu-Asn-lie-lie-Trp Fmoc-D-Txg-Leu-Glu-Ile-Ile-Trp Fmoc-D-Txg-Leu-Gin-lie-lie-Trp Fmoc-D-Txg-Leu-Tyr-lie-lie-Trp Fmoc-D-Txg-Leu-Asp-Val-lie-Trp Fmoc-D-Txg-Leu-Asp-lie-Val-Trp Fmoc - D - Txg - Leu - Asp - Chx-lie - Trp Fmoc - D - Txg-Arg - Asp - Chx-lie - Trp Fmoc-D-Txg-Lys-Asp-Chx-IIe-Trp Fmoc - D - Txg - Orn-Asp - Chx-lie - Trp Fmoc - D - Txg - Asp-Asp - Chx-lie - Trp Fmoc-D-Txg-Glu-Asp-Chx- Ile-Trp Fmoc - D - Txg - Leu-Asp - lie - Chx-Trp Fmoc-D-Txg-Arg-Asp-lie-Chx-Trp Fmoc-D-Txg-Lys-Asp-lie-Chx-Trp Fmoc-D-Txg-Orn-Asp-lie-Chx-Trp Fmoc-D-Txg-Asp-Asp-lie-Chx-Trp Fmo c - D - Txg - Glu - Asp -11 e - Chx - Trp Et-D-Bhg-Leu-Asp-lie-lie-Trp; Bz-D-Bhg-Leu-Asp-lie-lie-Trp; ■74- 165 170 175 180 185 190 Pya-D-Bhg-Leu-Asp-Ile-Ile-Trp Cxi-D-Bhg-Leu-Asp-Ile-Ile-Trp Ada - D - Bhg - Leu - Asp -11 e - II e - Trp j Cxi (U) -D-Bhg-Leu-Asp-Ile-Ile-Trp; Me (U) - D - Bhg - Leu -Asp - lie - lie - Trp ; tBu-D-Bhg-Leu-Asp-Ile-Ile-Trp; CF3CO-D-Bhg-Leu-Asp-lie-lie-Trp; Et-D-Bheg-Leu-Asp-Ile-Ile-Trp; Bz-D-Bheg-Leu-Asp-lie-lie-Trp; Pya-D-Bheg-Leu-Asp-lie-lie-Trp; Cxi-D-Bheg-Leu-Asp-lie-lie-Trp ; Ada-D-Bheg-Leu-Asp-Ile-Ile-Trp; Cxi(U)-D-Bheg-Leu-Asp-lie-lie-Trp; Me(U)-D-Bheg-Leu-Asp-lie-lie-Trp ; tBu-D-Bheg-Leu-Asp-lie-lie-Trp ; CP3 CO - D - Bheg - Leu - Asp - lie -11 e - Trp ; Ac-D-Bhg-Leu-Asp-Phe-lie-Trp; Ac-D-Bhg-Orn-Asp-Phe-lie-Trp Ac-D-Bhg-Lys-Asp-Phe-Ile-Trp Ac-D-Bhg-Asp-Asp-Phe-lie-Trp Ac-D-Bhg-Glu-Asp-Phe-Ile-Trp Ac-D-Bhg-Phe-Asp-Phe-lie-Trp Ac-D-Bhg-Arg-Asp-Phe-lie-Trp; Ac-D-Bheg-Leu-Asp-Phe-Ile-Trp ; Ac-D-Bheg-Orn-Asp-Phe-lie-Trp; Ac-D-Bheg-Lys - Asp- Phe - Ile-Trp; Ac-D-Bheg-Asp-Asp-Phe - Ile-Trp; Ac-D-Bheg-Glu-Asp-Phe-lie-Trp; Ac-D - Bheg - Phe - Asp - Phe -lie- Trp; and Ac-D-Bheg-Arg- Asp-Phe-lie-Trp. 5 -75- 25 2 8 55 A pharmaceutical composition adapted for administration as an antagonist of endothelin comprising a therapeutically effective amoimt of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent, or carrier. A pharmaceutical composition adapted for administration as an antihypertensive agent comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent or carrier. A pharmaceutical composition adapted for administration as an agent for treating metabolic and endocrine disorders comprising a therapeutically effective amount of a compound according to Claim l in admixture with a phannaceutically acceptable excipient, diluent or carrier. A pharmaceutical conposition adapted for administration as an agent for treating congestive heart failure and myocardial infarction comprising a therapeutically effective amount of a compound according to Claim l in admixture with a phannaceutically acceptable excipient, diluent or carrier. A pharmaceutical composition adapted for administration as an agent for treating endotoxic shock comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent or carrier. -76- 252855 10. A pharmaceutical composition adapted for administration as an agent for treating subarachnoid hemorrhage comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent or carrier. 11. A pharmaceutical composition adapted for administration as an agent for treating arrhythmias comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent or carrier. 12. A pharmaceutical composition adapted for administration as eui agent for treating asthma comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent or carrier. 13. A pharmaceutical composition adapted for administration as an agent for treating acute and chronic renal failure comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent or carrier. 14, A pharmaceutical composition adapted for administration as an agent for treating preeclampsia conqprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannace ^t2cS/lIyN f OFFICE acceptable excipient, diluent or cairie^ '■ 0 .HIN 1996 25 2 g A pharmaceutical composition adapted for administration as an agent for treating diabetes comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent or carrier. A pharmaceutical composition adapted for administration as an agent for treating neurological disorders comprising a. therapeutically effective amount of a compound according to Claim l in admixture with a phannaceutically acceptable excipient, diluent, or carrier. A pharmaceutical composition adapted for administration as an agent for treating pulmonary hypertension comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent, or carrier. A pharmaceutical composition adapted for administration as an agent for treating ischemic disease comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent, or carrier. A pharmaceutical composition adapted for administration as an agent for protecting against gastric mucosal damage or treating ischemic bowel disease comprising a therapeutically effective amoimt of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent, or carrier. -78- 25 2 8 20. A pharmaceutical composition adapted for administration as an agent for treating atherosclerotic disorders including Raynaud's disease comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent, or carrier. 21. A pharmaceutical composition adapted for administration as an agent for treating restenosis comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent, or carrier. ) 22. A pharmaceutical composition adapted for administration as an agent for treating angina comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent, or carrier. 23. A pharmaceutical composition adapted for administration as an agent for treating cancer comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent, or carrier. 24. A pharmaceutical composition adapted for administration as an agent for treating hemorrhagic shock comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a phannaceutically acceptable excipient, diluent, or carrier. i i ' . ; -■n m ws -79- 25. A method of preparing a compound of Po: 8^5 AA1 - AA2 - AA3 - AA4 - AA5 -AA6 wherein AA1 is R1 O 15 5 wherein R is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, 10 cycloalkylalkyl, aryl, heteroaryl, fluorenylmethyl, 2 2 3 -N-R , wherein R and R are each the same or u R different and each is hydrogen, alkyl, alkenyl, 20 alkynyl, cycloalkyl, cycloalkylalkyl, aryl, arylalkyl, 25 heteroaryl, or fluorenylmethyl, 30 35 -C-OR , wherein R is as defined above, -OR2, wherein R2 is as defined above, 0 -N-C-N-R3, wherein R2 and R3 are as defined above, R2 R2 I :--r-|CEl WN 1996 -80- 252855 -C-C(R9)3, wherein R9 is F, CI, Br, or I, -CH2-OR2, wherein R2 is as defined above, 40 O II 3 -N-C-R , R2a 45 wherein R2a is hydrogen or alkyl and R3 is as defined above, O 50 55 -N-C-OR3, R2a wherein R2a and R3 are as defined above excluding R3 is hydrogen, or 0 II 2 -C-R , wherein R is as defined above, R1 is hydrogen or alkyl, Z is -0-, 60 -S(0)fi, wherein m is zero or an integer of 1 or 2, -N-, wherein R2 is as defined above, l2 65 R - (CH2) n-, wherein n is zero or an integer of 1, 2, 3, or 4, -(CH2)n-CH-CH-{CH2)n-, wherein n is as defined above, 70 0 « . -C-, -CR1-, wherein R1 and R2 are as defined 75 OR* aboV®' °r 25 2 8 5 5 -81- -C- 80 85 90 95 R3 wherein R2 and R3 are each the same or different and each is as defined above, X and Y are the same and substituted at the same position on the aromatic ring and each may be one, two, three, or four substituents selected from the group consisting of hydrogen, halogen, alkyl, -C02R2, -CONR2, l3 R3 -NR2, R3 nltro or wherein R2 is as defined above, wherein R and R are as defined above, wherein R2 and R3 are as defined above, or wherein R, Z, X, and Y are as defined above; AA2 is R1 0 I. H —N—C~*C— 1 I R1 (CH2)n • A 100 wherein R4 is hydrogen, alkyl, !office} c !'! W 19.96 120 252855 -82- alkenyl, alkynyl, 105 cycloalkyl, aryl, heteroaryl, w. i 110 R2b wherein R2b and R3b are each the Bame or different and each is hydrogen, alkyl, 115 cycloalkyl, aryl, or heteroaryl, -OR2b, wherein R2b is as defined above, O -C-N-R3b, R2b wherein R2b and R3b are each 125 the same or different and each is as defined above for O R2b and R3b, 130 -C-R2b, wherein R2b is as defined above, NH -NH-C-NH-R2b, wherein R2b is as defined 135 above, or O -C-OR2b, wherein R2b is as defined above, and 140 R1 and n are as defined above, or AA2 is absent; [N.Z P/\J pf. r 2 0 M 1995 AA3 is R1 0 L II —N — C—C— | I R1 (CH2) n 252855 -83- 145 150 155 160 •wherein R5 is hydrogen, alkyl, aryl, heteroaryl, 0 ■C-N-R' i2b 3b f 2b -C-R , wherein R2b and R3b are each the same or different and each is as defined above, wherein R2b is as defined above, or ,2b ,2b 165 -C-OR"*", wherein R is as defined above, and R1 and n are as defined above, or AA3 is absent; AA4 and AA5 are each independently absent or each is independently R1 O I* II —N—C-C— . RMCH2)n i e. 170 wherein R6 is hydrogen, alkyl, alkenyl, ^[,'^TENT \ __ ^ 0 1996 252855 -84- alkynyl, cycloalkyl, aryl, or 175 heteroaryl, and R1 and n are as defined above; AA6 is r1 180 -N- -C*-R8 >i I r7 185 wherein R7 in aryl or heteroaryl, R® is O 190 || -C-OR1, wherein r1 is as defined above, -OR1, wherein R1 is as defined above, 195 0 -C-N-R1, wherein R1 is as defined jo. above, or 200 -CH2-OR1, wherein R1 is as defined above, and R1 and n are as defined above; * stereochemistry at C in AA1, AA2, AA3, AA4, or AA5 is 205 D, L, or DL and * stereochemistry at C in AA6 is L; or a phannaceutically acceptable salt thereof comprising sequential stepwise coupling of the amino acids 210 selected from AA1, AA2, AA3, AA4, AA5, or AA6 to the preceding amino acid using conventional peptide synthesis methodology and after conventfpttaJr— O ., . 25 2 8 55 deprotection to afford a compound of Formula I and, if desired, converting a compound of Formula I to a phannaceutically acceptable salt of a compound of Formula I by conventional methodology and, if further desired, converting the obtained phannaceutically acceptable salt of a compound of Formula I to a compound of Formula I by conventional methodology. 26. A compound of Formula I as claimed in claim 1 as specifically set forth herein, 27. A pharmaceutical composition as claimed in any one of claims 5 to 24 substantially as herein described. 28. A method of preparing a compound of Formula I substantially as herein described with reference to the Examples. </div>