J. M e d . Chem. 1991,34, 1258-1271
1258
New Inhibitors of Human Renin That Contain Novel Replacements at the P2Site' Annette M. Doherty,*,t James S. Kaltenbronn,t James P. Hudspeth,t Joseph T. Repine,t William H. Roark,' Ila Sircar,t Frank J. Tinney,t Cleo J. Connolly,t John C. Hodges,t Michael D. Taylor,? B r i a n L. Batley,' Michael J. Ryan,* Arnold D. Essenburg,* Stephen T. Rapundalo,* Ronald E. Weishaar,* Christine Humblet,t and Elizabeth A. Lunneyt Departments of Chemistry and Pharmacology, Parke-Davis Pharmaceutical Research Division, Warner-Lambert Company, Ann Arbor, Michigan 48105. Received J u n e 11, 1990
A series of renin inhibitors with novel modifications at the P2site has been prepared. Structure-activity relationships reveal that for a particular P, fragment the in vitro potency is highly dependent on the nature of the P,' portion in addition to the P1-P,' group. The length of the P2 side chain and choice of t-N P, substitution have been found to be important for in vitro potency although the degree of unsaturation in the P, side chain is not particularly significant. Molecular modeling studies have shown that it is possible for the P2side chain to interact unfavorably with the P,' binding site. It has been possible to control the specificity for renin over cathepsin D by correct modification at the P,' and Pl-Pl' sites. Variations at the P4site have been utilized to lower the log P values of these renin inhibitors while maintaining high potency. Compound 42, which exhibited an IC5,, of 3.70 nM, log P of 2.3, and showed high specificity for renin, was selected for further studies. It was found to be very stable under neutral, acidic, and basic conditions. In simulated intestinal juice, compound 42 had a half-life of 37 min while it was virtually unaffected by simulated gastric juice after 4 h. Compound 42 produced a significant hypotensive response upon intravenous administration to the salt-depleted normotensive cynomolgus monkey.
have reported that replacements of the Pz imidazole group Introduction with other heterocycles of varying basicity did not adRenin is an aspartic proteinase synthesized and stored in the epithelioid cells of the juxtaglomerular apparatus Abbreviations follow the recommendations of the IUPAC-IUB of the kidney. It is a highly specific proteolytic enzyme Joint Commission on Biochemical Nomenclature for amino that cleaves the polypeptide angiotensinogen to produce acids and peptides: Eur. J . Biochem. 1984, 158, 9-31. Addithe decapeptide angiotensin I., Angiotensin I, which has tional abbreviations are as follows: ACE, angiotensin conno known biological activity, is subsequently cleaved by verting enzyme; DCC, dicyclohexylcarbodiimide, DMF, diangiotensin converting enzyme (ACE) to afford the vamethylformamide; DMSO, dimethyl sulfoxide; BOC, tertbutoxycarbonyl; Z, benzyloxycarbonyl; TFA, trifluoroacetic soactive peptide angiotensin 11. The renin-angiotensin acid; TEA, triethylamine; HOBT, 1-hydroxybenzotriazole;Sta, system plays a central physiological role in the regulation (3S,4S)-4-amino-3-hydroxy-6-methylheptanoic acid; ACHPA, of blood pressure and electrolyte homeostasis. Recent (3S,4S)-4-amino-3-hydroxy-5-cyclohexylpentanoic acid; DFSta, evidence indicates that renin and angiotensinogen genes (3R,4S)-4-amino-2,2-difluoro-3-hydroxy-6-methylheptanoic and their products are expressed at many local tissue sites.3 acid; FCS, (3S,4S)-4-amino-2,2-difluoro-3R-hydroxy-5-cycloThe existence of a local renin-angiotensin system (RAS) hexylpentanoic acid; DFO, (4S)-4-amin0-2,2-difluoro-3-0~0-6methylheptanoic acid; AEM, 4-(2-aminoethyl)morpholine. is t h o u g h t to contribute to the control of vascular t ~ n e . ~ , ~ For recent reviews, see: (a) Greenlee, W. J. Med. Res. Rev. The activity of the tissue system under different conditions 1990,10,173. (b) Greenlee, W. J.Pharm. Res. 1987,4,364. (c) may influence the pharmacologic response to inhibitors of Wood, J. M.; Stanton, J. L.; Hofbauer, K. G . J.Enzyme Znhib. the RAS6 1987,1, 169. (d) Breipohl, G.; Geiger, R.; Henke, S.; Kleeman, T h e design of ACE inhibitors as useful d r u g s in the H.-W.; Knolle, J.; Ruppert, D.; Scholkens, B. A.; Urbach, H.; treatment of hypertension and heart failure has been reWagner, A.; Wegmann, H. Spec. Publ. R. SOC.Chem. 1988,65, 101. (e) Peach, M. J. Physiol. Rev. 1977,57(2),313. (f) Val~ i e w e d . Renin ~ however possesses much greater substrate loltman, M. B. Trends Pharmacol. Sci. 1987, 8, 69. specificity than ACE, and this property has led to intense Campbell, D. J. J . Cardiovasc. Pharmacol. 1987,10 (Suppl. 7 ) , interest in the search for effective inhibitors of renin.8 s1. Most of the p o t e n t renin inhibitors reported are based Dzau, V. J. J . Cardiovasc. Pharmacol. 1987,lO (Suppl. 7), S9. upon the natural substrate angiotensinogen sequence, inDzau, V. J. Circulation 1988, 77 (Suppl. I), 1-4. corporating nonhydrolyzable transition-state moieties at Blaine, E. H.; Schorn, T. W.; Boger, J. Hypertension 1984, 6 (Suppl. I), 1-111. the active site.+15 Thus these compounds remain partly (a) Ondetti, M. S.; Rubin, B.; Cushman, D. W. Science 1977, peptidic in nature and their therapeutic efficacy is limited 196, 441. (b) Cushman, D. W.; Cheung, H. S.; Sabo, E. F.; due to low oral activity and short duration of action. Ondetti, M. A. Biochemistry 1977, 16, 5484. (c) Kostis, J. B. Several studies of renin inhibitors in animals1620and more A m . Heart J . 1988, 116(6-1),1580. (d) Ertl, G. Eur. Heart J. recently i n h u m a n s have been reported.21~22 1988, 9, 716. (e) Ball, S. G. J . Cardiouasc. Pharmacol. 1987, In a search for novel and effective renin inhibitor^,,^ we 9 (Suppl. 3), S15. (f)Gavras, H.; Gavras, I. Hypertension 1988, I 1 (3), Suppl. 11, 37. (g) Nicholls, M. G.; Arakawa, K. J. became interested in variation at the P, site (nomenclature Cardiouasc. Pharmacol. 1987,9 (Suppl. 3), S19. (h) Zanchetti, for renin inhibitors illustrated for compound 17 in Scheme A. J . Cardiovasc. Pharmacol. 1987,9 (Suppl. 3), S2. (i) Kostis, I)24 where a histidine residue is present in the s u b s t r a t e J. B. A m . J . Hypertens. 1989, 2, 57. angiotensinogen and similarly in m a n y of t h e p o t e n t in(a) Haber, E. J . Hypertens. 1989, 7 (Suppl. 2), S81. (b) Kohibitors synthesized to date."15 It has been reported that kubu, T.; Hiwada, K. Drugs of Today 1987,23 (2),101. (c) the presence of histidine at P, is important for potency11J3 Baldwin, J. J.; Sweet, C. S. Ann. Rep. Med. Chem. 1988, 23, 59, and previous volumes in this series. as well as high s e l e c t i ~ i t y toward ~ ~ J ~ ~renin ~ ~ over related Boger, J.; Payne, L. S.; Perlow, D. S.; Lohr, N. S.; Poe, M.; aspartic proteinases. It has been suggested that the proBlaine, E. H.; Ulm, E. H.; Schorn, T. W.; LaMont, B. I.; Lin, tonated state of the imidazole group at P, is a contributor Tsau-Yen; Kawai, M.; Rich, D. H.; Veber, D. F. J. Med. Chem. to the inhibitor specificity.26 However Rosenberg et al. 1985, 28, 1779. Department of Chemistry.
* Department of Pharmacology.
Johnson, R. L. J . Med. Chem. 1984,27, 1351. Luly, J. R.; Bolis, G.; BaMaung, N.; Soderquist, J.; Dellaria, J. F.; Stein, H.; Cohen, J.; Perun, T. J.; Greer, J.; Plattner, J. J. J . Med. Chem. 1988,31, 532.
0022-2623/91/1834-1258$02.50/0 0 1991 American Chemical Society
Human Renin Containing Replacements a t the P2Site
Journal of Medicinal Chemistry, 1991, Vol. 34, No. 4 1259
Table I. Lysine Derivatives
compd 1
monkey plasma
R=H R = C(=S)NHCH, R = C(=O)NHCH3 R = C(=S)NHPh R = P(=O)(OPh), R = C(=NCN)SCH,
3 490 23 44 138 48% at 41 144
23 24 25
R = C(=O)NH, R = Tos R = COCH,
89 48% at 36
26 27 28
R = C(=S)NHCH, R = C(=NH)NHNOp R = C(=NCN)SCH,
29
R=Z R = C(=S)NHCH3 R = C(=NH)NHNO,
0
17 18 19
20 21 22
human plasma
log Po molecular formulab see ref 30 RNMA-His-Sta-NHCH2CH(CH3)CH&H3 85 61 BNMA-Lys(R)-Sta-NHCH2CH-(CH3)CH2CH3
structure
M
M
2 580 13 29 58 740 19 87
5.25 5.71 5.63
50 3000 6.2
5.31 6.70 4.38
120 45
3.91 4.09 4.67
6.60
7.88 5.51 7.21
BNMA-Lys(R)-Sta- AEM
30 31
125 40 90
C
BNMA-Lys(R)-Sta-Leu-NHCH,Ph 8 000 2 900 99 60 49 15
7.69 6.70
5.51
R’-L~s(C(=S)NHCH~)-ACHPA-AEM 32 33
R’ = BNMA Ph
29 27
C
c
5.44 2.64
36
20
1.68
693 350 395
108 c c
2.43 1.68 1.95
c
6.63
c c
4.83
14
R’= +oJco-
qo
34
II
R’ =
cn-c-
37
R = C(=S)NHCHB R = C(=NCN)SCH, R = C(=NH)NHNO,
38
BNMA-Lys(C(=S)NHCHB)-ACHPA-NHCH*- 47
35 36
CH(CHJCHZCH3 39
BNMA-Lys(C(=S)NHCH,)-Sta-N(CH,)(OCH,) 10560
40 41
BNMA-Lys(C(=S)NHCH,)-DFSta- AEM
Me,NS02-Phe-Lys(C(=S)NHCH,)-ACHPA-AEM 15
42
O~SO,-Phe-L*(C(=s)NHCH,)-ACHPI\
43
OANSO,--Phe-Lys(Z)--DFD-AEM
W
44
OANSOphe
45
OANSOphe-Hk
w
-Lys(C( =S)NHCHd -FCS
-ACHPA
-AEM
-AEM
-
AEM
350
5.50
3.2
C
2.41 2.26
3.3
C
2.32
2.3
c
2.93
5.0
5.4
2.25
W “log P values measured at pH 7.4. bAnalysis for C, H, N were within f40% except where noted. ‘Not measured. d T ~ isomers. o ‘N: calcd, 14.87; found 13.81. f S : calcd, 4.20; found 4.72.
versely affect the potency of a series of statine-derived renin inhibitomn Moreover, potent renin inhibitors with
a @-aspartylresidue at the P2site have recently been reported.28
(12) Thaisrivongs, S.; Pals, D. T.; Kroll, L. T.; Turner, S. R.; Han, F.-S. J . M e d . Chem. 1987, 30, 976.
(13) Hui, K. Y.; Carlson, W. D.; Bernatowicz, M. S.; Haber, E. J. Med. Chem. 1987,30, 1287.
Doherty et al.
1260 Journal of Medicinal Chemistry, 1991, Vol. 34, No. 4
Scheme I OH
’I BOC -NHCHCHCH2C02H
H2N
---
I v --
OH
- ’I
BOC-NHCHCHCH2CONHCH2CH( C H 3 I C H 2 C H j b
t-
DCC.
t-
HOBT
2
1. H C 1 , CH2C12 E-Z-L~S-S~~-NHCH~CH(CH~)CH~CH~
b
2.
(N-a-BOC-N-E-Z)
3. T F A .
- L y s i ne,
DCC,
HOBT
3
CH2Cl2
( q i C H C O 2 H DCC,
H2.
BNMA- Lys ( 2 ) - Sta -NHCHzCH( CH3 ) C H ~ C H J
HOBT
A
Pd/C b
BNMA- Lys - S t a -NHCHzCH( CH3 )CH2CH3
MeOH
5
Table 11. Chain Leneth Variation ICw, nM monkey Dlasma human Dlasma
lop P BNMA-NHCH(R”)CO-Sta-NHCHzCH(CH3)CH2CH3
comDd
structure
46 47
R” = CH2CHZNHCOCH3 R” = CH2CH2NHC(=S)NHCH3
48
R” = (C”&- J - N
4150 2490 390
>io4 M 9350 540
39 R” = (CH2)6NHC(=S)NHCH3 20 R” = (CHZ)SNHC(=NH)NHNOz 44 R” = (CHz)6NHC(=S)NHCH3 “log P values measured at pH 7.4. *Analysis for C, H, N were within *0.4%
49 50 51
3.6/4.4c 5.515.T 7.09
molecular formulab
c43H56N405 C43H57N504S.0.35CHC13 C46H58N404.0.5H20
40 5.41 C~~H~~NSO~S.O.~CH~CI~ 26 5.10 C45H6lN706.0.3CH2C12 48 5.64 C47H6sN504S.0.33C4H~O2 except where noted. C T wisomers. ~
Our study into the effect of variation a t the Pz site indicates that replacement of histidine with modified lysine
derivatives maintains high potency, but often reduces selectivity for renin. However, suitable modifications, par-
(14) Luly, J. R.; Yi, N.; Soderquist, J.; Stein, H.; Cohen, J.; Perun, T. J.; Pattner, J. J. J. Med. Chem. 1987, 30, 1609. (15) Allen, M. C.; Fuhrer, W.; Tuck, B.; Wade, R.; Wood, J. M. J . Med. Chem. 1989, 32, 1652. (16) Wood, J. M.; Criscione, L.; de Gasparo, M.; Buhlmayer, P.; Rueger, H.; Stanton, J. M.; Jupp, R. A.; Kay, J. J. Cardiouas. Pharmacol. 1989, 14, 221. (17) Wood, J. M.; Jobber, R. A.; Baum, H.-P.; de Gasparo, M.; Nussberger, J. J . Hypertension 1989, 7, 615. (18) Greenfield, J. C.; Cook, K. J.; O’Leary, I. A. Drug. Metab. Dispos. 1989, 17 (51, 518.
(19) Morishima, H.; Koibe, Y.; Nakano, M.; Atsuumi, S.; Tanaka, S.; Funabashi, H.; Hashimoto, J.; Sawasaki, Y.; Mino, N.; Nakano, M.; Matsushima, K.; Nakamichi, K.; Yano, M. Biochem. Biophys. Res. Commun. 1989, 159 (3), 999. (20) Verburg, K. M.; Kleinert, H. D.; Kadam, J. R. C.; Chekal, M. A.; Mento, P. F.; Wilkes, B. M. Hypertension 1989, 13, 262. (21) Delabays, A.; Nussberger, J.; Porchet, M.; Waeber, B.; Hoyos, P.; Boger, R.; Glassman, H.; Kleinert, H. D.; Luther, R.; Brunner, H. R. Hypertension 1989, 13, 941. (22) De Gasparo, M.; Cumin, F.; Nussberger, J.; Guyenne, T. T.; Wood, J. M.; Menard, J. Br. J . Clin, Pharmac. 1989, 27, 587.
Human Renin Containing Replacements at the PzSite
Journal
of Medicinal Chemistry, 1991,
Vol. 34, No. 4 1261
Table 111. Unsaturation in the Pz Lysine Side Chain
compd
52 53
54 55a 55b
56 57 58
59 60
structure
IC,. nM monkey human plasma plasma
log P BNMA-NHCH(R”’)CO-Sta-NHCH,CH(CHq)CHeCHq
R”’ R”’ R”’ R”’ R”’
= CHzC=C-CHzNHZ = CHzCH=CHCH,NHC(=S)NHCH3
R”’ R”’ R’” R”‘ R”’
= BOC = C(=S)NHCHB = C(=NH)NHNOz = COCH3 = CHO
(E)
= CHzCH=CHCHzNHCOCH3 (2) = CH2C=CCHzNHCOCH3 (isomer A) = CH2CsCCH2NHCOCH3(isomer B)
248 26 100 4.6 41
156 41
7:13/7:37‘ 5.23 5.56 5.32 5.06
g
2.0 29
~
molecular formula*
C51H~oN406.0.33C4H8Oz~ C46H,N504S.O.O7CHC13 C45H5aN405*0.5CH30He C45HsN405.0.1CHC1, C45HsN405.0.05CHC13
BNMA-NHC(CHL!EC-CH,NHR“’)HCO-ACHPA-AEM 700 48 12 9.0 8.7 0(7NSO,--Phe--NHCHCO--ACHP-A
u
--AEM
I
C H & E C -CH,NHR”’
R”’ = BOC (isomer A) 11.0 g 3.96 C41H65N7010S R”’ = BOC (isomer B) 700 g 3.13 C4,H6,N,O701oS.O.8CHCl~ R”’ = C(=S)NHCH3 (from 61a) 3.3 g 2.90 C~~H~NSO~S~.C~H~O~.~.~H~O~O 0.5 g 2.12 C38H59N7O&C6H807*1.4HzO.O.O5CHC13 63 R”’ = COCH, (from 61a) ulogP values measured at pH 7.4. *Analysis for C, H, N, were within *0.4% except where noted. c T w isomers. ~ dC: calcd, 53.57; found 53.04. ‘N: calcd, 7.46; found 8.00. fN: calcd, 6.70; found 6.18. gNot measured.
61a 61b 62
ticularly at the P2’site in our lysine-type P2-substituted renin inhibitors restores high selectivity. Thus we have prepared a large range of novel compounds (Tables 1-111) many of which are highly potent and selective for renin.
Chemistry Table I lists compounds (17-45) with a lysine P2residue, where the t-amine is substituted with a variety of groups. These compounds were prepared by using known synthetic methods.29 Scheme I illustrates the synthesis of 17, exemplary of this group. In addition the corresponding histidine analogue 1 is included in Table I for comparison (BNMA-H~S-S~~-NHCH,CH(CH,)CH,CH,).~~ Thus BOC-statine was coupled with (S)-3-methylbutylamine (MBA),30by using the standard DCC/HOBT procedure to afford 2 which was followed by hydrochloric acid deprotection and subsequent coupling with N-aBOC-N-t-Z-lysine. Trifluoroacetic acid treatment and neutralization gave 3. Coupling with bis(1-naphthylmethy1)acetic acidm gave intermediate 4. Hydrogenolysis to remove the Z protecting group gave 5, which was treated
-
Scheme I1 1. N a H .
DMSO
BNMA-NHCH(C02Et)2
BNMA-NHCH(C02Et)Z
I
2 . CH3SOzO(CHz)5NHZ
7
(CH2)sNHZ
a 1. NaOH
BNMA- NHCHCOzH
I
D
2 . QCH3. A
(CH2)sNHZ 9
Scheme 111 N a H . DMSO BNMA-NHCH(C0zEt )z ICHzCHzNHBOC 7
-
C02Et
I BNMA-NHC
10 11
1. N a O H . MeOH 2 . OCH3. A
BNMA-NHCHC02H
h
*
3. NaOH. MeOH
NHBOC
SB.NHCH$H(CH3)CI+&Y
*
H O B T . DCC
12
Kaltenbronn, J. S.; Hudspeth, J. P.; Lunney, E. A.; Michniewicz, B. M.; Nicolaides, E. D.; Repine, J. T.;Roark, W. H.; Stier, M. A,; Tinney, F. J.; Woo, P. K. W.; Essenburg, A. D. J. Med. Chem. 1990, 33, 838. Schechter, I.; Berger, A. Biochem. Biophys. Res. Commun. 1967,27, 157. Cumin, F.; Nisato, D.; Gagnol, J.-P.; Corvol, P. Biochemistry 1987,26,7615. Maibaum, J.;Rich, D. H. J . Med. Chem. 1988,31, 625. Rosenberg, S. H.; Plattner, J. J.; Woods, K. W.; Stein, H. H.; Marcotte, P. A.; Cohen, J.; Perun, T. J. J. Med. Chem. 1987, 30,1224.
Thaisrivongs, S.; Mao, B.; Pals, D. T.; Turner, S. R.; Kroll, L. T. J . Med. Chem. 1990,33,1337. (a) Konig, W.; Geiger, R. Chem. Ber. 1970,103,788. (b) Bodanszky, M. Principles of Peptide Synthesis; Springer-Verlag: Berlin, 1984, p 158. (c) Bodansky, M. Peptide Chemistry-A Practical Textbook;Springer-Verlag: Berlin, 1988. (a) Kokubu, T.; Hiwada, K.; Nagae, A.; Murakami, E.; Morisawa, Y.; Yabe, Y.; Koike, H.; Iijima, Y. Hypertension 1986, 8 (Suppl. II), 1. (b) Morisawa, Y.; Yabe, Y.; Kataoka, M.; Iijima, Y.; Kokubu, T.; Hiwada, K.; EP 186,977, 1986.
BNMA-NHCHCO-Sta-NHCHZCH(CH3)CHzCH,
h
13
NHBOC
with methyl isothiocyanate in dichloromethane affording renin inhibitor 17. The syntheses of renin inhibitors 40,43, and 44 in Table I began by coupling of N-a-BOC-difluorostatine (DFSta),3l with 4-(Zaminoethyl)morpholine (AEM), deprotection and subsequent coupling with N-a-BOC-N-t-Z-lysine. Acidic deprotection was followed by standard coupling procedures with either bis(1-naphthylmethy1)aceticacid (BNMA) or (31) (a) Thaisrivongs, S.; Pals, D. T.; Kati, W. M.; Turner, S. R.; Thomasco, L. M.; Watt, W. J.Med. Chem. 1986,29,2080. (b) Gelb, M. H.; Svaren, J. P.; Abeles, R. H. Biochemistry 1985, 24,1813. (c) Fearon, K.; Spaltenstein, A.; Hopkins, P. B.; Gelb, M. H. J . Med. Chem. 1987,30, 1617.
Doherty et al.
1262 Journal of Medicinal Chemistry, 1991, Vol. 34, No. 4 Table IV. Selectivitv Data ~~
compd 17 26 32 38
~
ICm, nm (and/or % inhibition at lo+ M) cathepsin D renin structure (bovine) (monkey plasma) BNMA-Lys(C(=S)NHCH3)-Sta-NHCH2CH(CH3)CHzCH3 781 23 BNMA-Lys(C(=S)NHCH,)-Sta-AEM 5 403 125 BNMA-Lys(C(=S)NHCHJ-ACHPA-AEM 9% at lo+ M 29 BNMA-L~s(C(=S)NHCH~)-ACHPA-NHCH&H(CH~)CH~CH~ 3 200 47 33 150 3.4
ratio C/R 34 43 68 9 750
BNMA-NHCH(R')-CO-Sta-NHCHzCH(CH3)CH2CH3 55a
R' = (CH2),NHC(=NH)NHNO2 R1 = CHzCH=CHCH2NHC(=S)NHCH3 R1 = CH2C=C-CH2NHCOCH3
56 58 59
R = C(=S)NHCH3 R = C(=NH)NHNO2 R = COCH3
50 52
1471 1922 78
20 41 4.6
74 47 17
BNMA-NHCH(CH&=C--CH,NHR)CO-ACHPA-AEM 81 000 0.7% at lo4 M 0.3% at 10" M
(morpholinosulfonyl)phenylalanine.32Compound 43 was obtained by Swern oxidation of the precursor difluorostatine compound. Renin inhibitors 32,33,34,41, and 42 were synthesized by using standard chemistry from intermediate Lys(Z)ACHPA-AEM,by coupling with the requisite P3-P4 group, hydrogenolysis, and acylation. The inhibitors in Table 11,46-51 illustrate the variation in chain length at the P, position. The pyrrole analogue 48 (Table 11) was prepared from aminoacid 6.33734 Me0 G
O
M
e
BOC-NHCHC02H b
a-BOC-ORN
HOAc
(
'
CH2 ) 3 - N
3
48 11 9
1688 >103 >io3
Table V. Physical Data for Renin-Inhibiting Compounds compd log P solubility,* mg/mL 17 5.25 26 3.91 30 6.70 35 2.43 41 2.41 42 2.26 a pH 7.4. * pH 7.4 phosphate buffer.
