A new class of bradykinin antagonists containing indanylglycine

Immunopharmacology ELSEVIER

Irnmunopharmacology 33 (1996) 174-177

A new class of bradykinin antagonists containing indanylglycine Lajos Gera, John M. Stewart * Department of Biochemistry, Universityof Colorado School of Medicine, Denver, CO 80262, USA

Keywords: Bradykinin antagonist

1. Introduction

Bradykinin (Arg-Pro-Pro-Gly-Phe-Ser-Pro-PheArg, BK) is an endogenous linear nonapeptide hormone that has a broad range of biological activities in both normal physiology and pathophysiology. It is the role of BK in pathophysiology that has made the search for its antagonists of great interest. Most actions of BK in normal tissues are mediated by B 2 receptors, that require the entire peptide chain. In inflammation, a second class of receptors, called B1, is expressed; these recognize primarily BK(1-8) (Bhoola et al., 1992). The first B 2 antagonist to receive wide application to many biological systems, NPC-349 (DArg-Arg-Pro-Hyp-Gly-Thi-Ser-DPheThi-Arg), was developed by Vavrek and Stewart (1985) a decade ago. The second generation of BK antagonists is characterized by the Hoechst Icatibant (HOE-140), in which NPC-349 was modified further

Abbreviations: Aaa, adamantane acetyl; Aca, adamantane carboxyl; Cpg, cyclopentylglycine;Cpg(R)Arg, reduced peptide bond: -CHz-NH; Dhq, 2-dehydroquinuclidine-3-carboxyl; Eac, t-aminocaproic acid; Gun, guanidyl-; HBQ, 4-hydroxybutyl-glutamine; Hig, hexahydro-Igl; Hyp, trans-4-hydroxy-L-proline; Ica, indole2-carboxylic acid; Igl, a-(2-indanyl)glycine; Nig, N-(2indanyl)glycine; Nal, [3-2-naphthylalanine; NMF, N-methylphenylalanine; Oic, octahydroindole-2-carboxylic acid; Sin, sinapinyl(3,5-dimethoxy-4-hydroxycinnamyl); Ser(S), serine Osulfate; Thi, [3-2-thienyl-Ala; Tic, 1,2,3,4-tetrahydroisoquinoline3-carboxylic acid * Corresponding author.

at positions seven (D-Tic) and eight (Oic) (Hock et al., 1991) and the Cortech Bradycor (CP-0127), a dimer that shows potency and persistence of action in vivo (Cheronis et al., 1992). Nova investigators reported that analogs of NPC-349 containing 4-npropoxy- or 4-phenylthio-proline at position seven were particularly potent (Kyle et al., 1991). We report here a series of new BK antagonists containing indane-substituted amino acid residues in various positions of BK analogs. Some of these show very high potency in vitro and long duration of action in vivo: examples are B9224, B9340, B9430 and B9330. Probably the most remarkable property of these analogs containing a-(2-indanyl)glycine is that they show potent BK antagonism on B 1 receptors, as well as the anticipated B 2 receptor activity (B9340 and B9430). We have also synthesized analogs with reduced peptide bonds, N-terminal acylation or guanylation and serine O-sulfation.

2. Materials and methods

2.1. Chemistry Boc-a-(2-indanyl)glycine (Igl) was synthesized as described in the adjoining paper (Stewart et al., 1996). The saturated derivative (Boc-Hig) was made by catalytic hydrogenation of Boc-Igl over P d / C at 80°C, 100 atm; it was an oil. N-(2-Indanyl)glycine

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L. Gera, J.M. Stewart / lmmunopharmacology 33 (1996) 174-177

Table 1 Structures and activities of indane-substituted bradykinin antagonists Number

Structure 0

B9224-2 B9430 B9330 B9340 B9464 B9456 B9616 B9588 B9546 B9626 B9728 B9756 B9688 B9706 B9686 B9694 B9724 B9568 B9480 B9432 B9564 B9348 B9626-1 B9226-2 B9436 B9124-1 B9124-2 B9238 B9720 B9340 B9224-2 B9256 B9324 B9334 B9430 B9560 B9562 B9594 B9600 B9598 B9630 B9618 B9226-2 B9228-2 B9340 B9668 B9726 B9430 B9698 B9598 B9696

Aca

Aca Aaa Dhq Sin Aca Dhq Aaa Sin

DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg Arg DArg Arg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg DArg

Sin Aca DArg DArg DArg Gun DArg Gun-Eaa)Arg DArg Gun DArg Aca

Gun

Activity 1

Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg

2

3

4

5

6

7

8

9

Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro NMF Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro Pro

Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Pro Pro Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp Hyp

Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly Gly

Thi Igl Thi Thi DThi Dlgl lgl Igl Thi Thi Igl lgl Igl Igl Igl Igl Igl Igl Igl lgl lgl Cpg Phe Phe Thi Thi Thi Thi lgl Thi Thi Thi Thi Thi lgl lgl Igl lgl Igl Igl lgl lgl Phe Phe Thi Thi Thi Igl Igl Igl Igl

Ser Ser Ser Ser Set Set Thr Lys HBQ Glu Tyr Ser Set Set Ser Ser Ser Ser Ser Set Set Ser Ser Ser Ser Set Set Set Set Set Ser Ser Ser Set Set Ser Ser Ser Ser Ser Set Set Set Set Ser Ser Set Ser Ser Ser Ser

DIgl DIgl DTic DIgl Dlgl DIgl Dlgl Dlgl DIgl Dlgl DIgl DIgl DNal Dlgl DIgl DIgl Dlgl DIgl Dlgl DTic DIgl DIgl Igl Dlgl Dlgl Hig DHig Nig Oic Dlgl Dlgl Dlgl Dlgl Dlgl DIgl Dlgl Dlgl Dlgl DIgl Dlgl Dlgl Dlgl Dlgl uIgl DIgl DIgl DIgl DIgl Dlgl DIgl DIgl

Oic Oic Nig Oic Oic Oic Oic Oic Oic Oic Oic Nal Oic Ica Oic Oic Oic Oic Cpg Oic Igl Oic Oic Oic Nig Oic Oic Oic lgl Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic Oic

Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg

UTERUS pA 2 9.2 8.5 8.3 7.9 8.2 8.0 8.0 7.8 7.3 6.5 8.1 7.5 7.6 8.0 7.6 7.4 7.1 7.8 1.0% 1.0% Biph 6.5 7.9 49.0% Biph 10.0% 6.8 210.0% 7.9 9.2 7.7 8.1 8.3 8.5 7.5 7.4 7.9 7.3 7.6 7.3 7.1 7.9 8.2 7.9 Biph 7.7 8.5 7.9 7.6 7.7

ILEUM pA 2 8.4 7.9 7.2 8.0 6.0 7.8 7.0 7.6 7.1 7.0 7.7 5.5 6.7 6.0 7.5 7.3 7.6 6.8 6.8 7.8 6.5 8.1 5.1 7.6 6.7 I(P) 6.6 7.0 Enh 8.0 8.4 7.8 7.2 7.3 7.9 7.8 7.6 7.6 7.0 7.8 7.4 7.9 7.6 7.4 8.0 7.8 7.6 7.9 7.2 7.8 7.6

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L. Gera, J.M. Stewart / Immunopharmacology 33 (1996) 174-177

Table l (continued) Number

B9448-1 B9448-2 B9480 B9718

Structure

Aca Aca

Activity

0

1

2

3

4

DArg DArg DArg DArg

Arg Arg Arg Arg

Pro Pro Pro Pro

Hyp Hyp Hyp Hyp

Gly Gly Gly Gly

5 Cpg Cpg Igl Igl

6

7

8

9

Ser Ser(S) Ser Ser

Dlgl DIgl Dlgl olgl

Oic Oic Cpg Cpg(R)

Arg Arg Arg Arg

UTERUS pA 2 8.1 6.6 7.8 6.4

ILEUM pA 2 8.3 7.9 6.8 5.4

Antagonist potencies are given as the pA 2. Agonist potencies are given as per cent of BK activity. Biph, biphasic effect (agonist, then antagonist); Enh, enhances response to BK; I(P), partial inhibition.

(Nig) was made by reductive amination of 2-indanone by glycine methyl ester. Its Boc derivative had mp 130-131°C. Peptides were synthesized by standard solid-phase methods (Stewart and Young, 1984) and purified and characterized as described in the adjoining paper (Stewart et al., 1996). N-terminal acylation of peptides on synthesis resins was by use of pre-activated acids in DMF. Free peptides were similarly acylated in DMF solution with BOP or HATU activation, using DIEA as base. N-terminal guanylation of peptide-resins was done with a 4-fold excess of N,N'bisCbz-l-guanylpyrazole (Bernatowitz and Matsueda, 1994) in DMF at room temperature or 50°C. Serine-containing peptide resins were sulfated with 40 equiv, of pyridinium acetyl sulfate in DMF/pyridine (1 : 1) for 20 h at room temperature. The reduced peptide bond for analog B9718 was introduced by reductive alkylation of Arg(Tos)-resin by Boc-cyclopentylglycine aldehyde by the procedure of Sasaki and Coy (1987).

tagonist potencies on smooth muscles are given as the pA 2 value; for comparison the pD 2 value for BK in this laboratory is 7.9 on rat uterus and 7.4 on guinea pig ileum. Most of these peptides were also antagonists on rat blood pressure, and showed long duration of action in vivo; inhibition by B9430 lasted at least 4 h. Most of the antagonists also showed protracted antagonism on the isolated smooth muscles. The superior properties of these antagonists containing Igl indicate that they constitute a new, third generation of BK antagonists.

Acknowledgements This work was aided by grants from the U.S. NIH (grant HL-26284) and from Cortech Inc. We thank Robin Reed and Paul A. Bury for assistance with chemistry, and Frances Shepperdson for assays. We thank G.R. Matseuda for a gift of N,N'-bisCbz-1guanylpyrazole.

2.2. Bioassays Assays for bradykinin antagonism at B 2 receptors were done on isolated guinea pig ileum, estrus rat uterus and rat blood pressure by standard methods (Stewart et al., 1996). B~ antagonist activity was assayed on isolated rabbit aorta and by receptor binding, and is reported in later papers in these proceedings (Burkard et al., 1996; Hanson et al., 1996).

3. Results and discussion Structures and smooth muscle activities of the BK analogs containing Igl are reported in Table 1. An-

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