Non-peptide αvβ3 antagonists. Part 3: Identification of potent RGD mimetics incorporating novel β-amino acids as aspartic acid replacements

Article abstract of DOI:10.1016/S0960-894X(01)00666-7

Potent non-peptidic α_vβ₃ antagonists have been prepared incorporating various β-amino acids as aspartic acid mimetics. Modification of the β-alanine 3-substituents alters the potency and physicochemical properties of these receptor a

Full text of DOI:10.1016/S0960-894X(01)00666-7

Bioorganic & Medicinal Chemistry Letters 12 (2002) 31–34
Non-Peptide ꢀvꢁ3 Antagonists. Part 3: Identification of Potent
RGD Mimetics Incorporating Novel ꢁ-Amino Acids as Aspartic
Acid Replacements
Paul J. Coleman,a,* Karen M. Brashear,a Cecilia A. Hunt,a William F. Hoffman,a
John H. Hutchinson,a Michael J. Breslin,a Carol A. McVean,a Ben C. Askew,a
George D. Hartman,a Sevgi B. Rodan,b Gideon A. Rodan,b Chih-Tai Leu,b
Thomayant Prueksaritanont,c Carmen Fernandez-Metzler,c Bennett Ma,c
Laura A. Libby,c Kara M. Merkle,c Gary L. Stump,d Audrey A. Wallace,d
Joseph J. Lynch,d Robert Lynchd and Mark E. Duggana
aDepartment of Medicinal Chemistry, Merck Research Laboratories, West Point, PA 19486, USA
bDepartment of Bone Biology and Osteoporosis Research, Merck Research Laboratories, West Point, PA 19486, USA
cDepartment of Drug Metabolism, Merck Research Laboratories, West Point, PA 19486, USA
dDepartment of Pharmacology, Merck Research Laboratories, West Point, PA 19486, USA
Received 18 June 2001; accepted 2 October 2001
Abstract—Potent non-peptidic avb3 antagonists have been prepared incorporating various b-amino acids as aspartic acid mimetics.
Modification of the b-alanine 3-substituents alters the potency and physicochemical properties of these receptor antagonists and in
some cases provides orally bioavailable avb3 inhibitors. # 2001 Elsevier Science Ltd. All rights reserved.
Osteoporosis is a chronic skeletal disease characterized
by both decreasingbone mass and increased bone fra-
gility.1 In healthy adults, bone mass is maintained by a
homeostatic balance between the processes of bone
resorption and new bone formation. In post-menopau-
sal women, increases in osteoclast-mediated bone
resorption lead to net reductions in bone mass and an
increased incidence of fracture. The mechanism of bone
resorption involves the attachment and migration of
osteoclast cells to the bone surface in a process that is
mediated by the highly expressed transmembrane integrin
avb3.2 This adhesion event presumably involves integrin
bindingto an RGD (arg-gly-asp) tripeptide sequence
found in extracellular matrix proteins on the bone sur-
face. Once osteoclasts attach to the bone matrix, a cas-
cade of intracellular signalling events leads to the
formation of a sealed acidified environment and secretion
of matrix degrading proteinases at the osteoclast-bone
interface.3 Antagonism of the RGD binding domain of
avb3 inhibits bone resorption and offers the potential of
a selective, non-hormonal therapy for osteoporosis.4
In a previous communication, we disclosed a novel ser-
ies of constrained RGD mimetics as avb3 antagonists.5
These integrin antagonists exemplified by 1 incorpo-
rated three key structural features: (1) a tetrahydro-
[1,8]napthyridine as a conformationally-locked guani-
dine replacement with reduced basicity, (2) a stereo-
differentiated pyrrolidinone central constraint, and (3) a
3-substituted 3(S)-pyridinyl b-amino acid as an aspartic
acid replacement. In spite of sub-nanomolar binding
affinity of 1 to human avb3, poor oral pharmacokinetics
emerged as a limiting feature for this compound. We
envisioned replacingthe 3-pyridinyl substituent in
1
with more lipophilic aryl moieties to further enhance
potency in this series and to confer improved bioavail-
ability. In this communication, we disclose novel, potent
C-terminal aspartic acid replacements6 as part of our
continuingeffort to discover non-peptidic avb3 antago-
nists that may be administered orally in man for the
treatment and prevention of osteoporosis.
*Correspondingauthor. Tel.: +1-215-652-4618; fax: +1-215-652-
7310; e-mail: paul_coleman@merck.com
0960-894X/02/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00666-7

32
P. J. Coleman et al. / Bioorg. Med. Chem. Lett. 12 (2002) 31–34
administration to dogs precluded further evaluation of
this analogue in in vivo models of bone resorption. We
suspected that the hydrophilic character of 1 (measured
logP=ꢀ0.61) adversely affected oral absorption.
Therefore, we focused our efforts on incorporation of
more lipophilic, non-basic 3-aryl groups in this series
with the goals of preserving sub-nanomolar potency
while obtainingan acceptable pharmacokinetic profile.
The synthesis of pyrrolidinone avb3 receptor antagonists
described in this paper involved the union of 3-aryl b-
aminoesters with carboxylic acid 6 (Scheme 1). The
resultant esters were saponified to yield the final analo-
gues (8–22) described in Table 1.7 Enantioselective
synthesis of the 3-aryl b-aminoesters was generally
In evaluatingclosely-related fibrineong receptor
antagonists, we and others12 had previously identified
the 3-quinolinyl b-amino acid as a potent aspartic acid
mimetic. The 3-quinolinyl analogue 10 exhibited a 10-
fold gain in SPAV3 potency versus the 3-pyridinyl ana-
logue 1. This highly potent antagonist demonstrated
additional lipophilicity (measured logP 0.73) over the 3-
pyridyl derivative.
8
achieved by utilizingthe Davies method for hetero-
conjugate addition of (R)-(+)-N-benzyl-a-methylben-
zylamine to the desired 3-aryl acrylates (Scheme 1).
Dependingon the commercial availability of starting
materials, the 3-aryl acrylates were prepared by either
Wittigolefination with the desired aryl aldehyde ( 2!4)
or via a Heck cross-couplingwith the aryl halide ( 3!4).
Syntheses of the 3-pyridinyl, 5-benzodioxole, 3-quino-
line, and dihydrobenzofuranyl b-amino esters are
described in the literature.9 Carboxylic acid 6 was
transformed to receptor antagonists 7 by standard
carbodiimide couplingand ester hydrolysis.
We next investigated the utility of a 3-benzodioxole b-
amino acid as an aspartic acid replacement5b in these
integrin antagonists. Incorporation of a benzodioxolyl
3-substituent in this lactam-constrained integrin
antagonist series resulted in potent avb3 antagonist 11
(Table 1). The benzodioxole moiety has been reported
to induce and inhibit human P-450 isozymes via forma-
tion of a metallocarbene complex between the dioxy-
methylene and heme functions.13 Indeed, the ethyl ester
prodrugof 11 was a potent time-dependent inhibitor or
human CYP3A although the acid 11 was not. Never-
theless, because of these metabolic concerns, an effort to
identify potent, structurally related bicyclic systems was
undertaken. Receptor antagonists (12–18) that contain
benzodioxole-like 3-substituents are shown in Table 1.
Blockingof the metabolic activation site of the benzo-
dioxole by difluoro-substitution to give 12 resulted in a
loss of potency. Both the 5- and 6-[2,3]-dihy-
drobenzofurans were prepared and evaluated in our
primary bindingassays. We were gratified to discover
that the 6-dihydrobenzofuran 14 had superior potency
relative to the benzodioxole and lacked P-450 inhibi-
tion.9d The 5-dihydrobenzofuran isomer 13 had much
lower receptor affinity. Benzofuran 14 suggests the need
for heteroatom substitution in the meta position of the
3-aryl ring. The critical importance of the furanyl oxy-
gen atom in analogue 14 was illustrated by evaluation of
the carbocyclic pseudo-isostere 15. Replacement of the
The compounds synthesized in this study were eval-
uated for their ability to inhibit the in vitro bindingof a
high affinity radioligand to human avb3 immobilized on
scintillation proximity beads (SPAV3).10 Selectivity for
avb3 versus the fibrinogen receptor (aIIbb3) was deter-
mined by measuringinhibition of the rate of ADP-sti-
mulated platelet aggregation in human plasma.11 This
series of avb3 antagonists has uniformly low affinity for
the fibrinogen receptor (IC50 >1 mM).
The 3-aryl substituent is an important feature for
enhancingreceptor potency and modulatingphysical
properties in this series. For example, the 3-phenyl ana-
logue 9 binds with nearly 60-fold greater affinity relative
to the unsubstituted derivative 8 (Table 1). Although
the 3-pyridinyl substituent in 1 provides an additional
10-fold gain in potency versus the 3-phenyl analogue 9,
the poor pharmacokinetic profile of 1 followingoral
Scheme 1. Preparation of avb3 receptor antagonists.

P. J. Coleman et al. / Bioorg. Med. Chem. Lett. 12 (2002) 31–34
33
furanyl oxygen atom in 14 with a methylene unit in 15
affords a 200-fold reduction in inhibitory potency. The
benzoxazolidone 16 had comparable potency to the
benzodioxole, while the benzothiophene 17 and ben-
zothiazole 18 were significantly less active avb3 antago-
nists. For this series of receptor antagonists, bicyclic 3-
substituents that contained a meta-oxygen atom were
more potent than the correspondingsulfur-containing
bicyclic systems.
Replacement of the 3-pyridinyl substituent with 3(3-
fluorophenyl) group provided analogue 19 (Table 1)
which had diminished potency and increased lipophili-
city. Efforts to improve the potency of 19 centered on
exploringthe potency enhancement effects of additional
substituents on the fluorophenyl ring(Table 1). Our
results with the dihydrobenzofuran 14 and 3-quinoline
suggested that we could tolerate additional substitution
at the 4-position of the aryl ring. Indeed, we discovered
that incorporation of an aryl or alkoxy substituent in
the 4-position of the 3-fluorophenyl moiety significantly
enhances potency (2- to 6-fold). In particular, analogues
that contained a 4-alkoxy-3-fluorophenyl b-amino acid
(21 and 22) were more potent than 19 while maintaining
comparable physical properties. The potency of analo-
gue 22 is comparable to our initial lead receptor
antagonists 1 and 11.
Table 1. avb3 receptor antagonists
R
Entry SPAV3 (IC50, nM)
Plaggin
ND
LogP
ND
8
230
9
4.0
8% at 10 mM
0.07
1
0.35
0.04
0.32
5.8
50% at 11 mM ꢀ0.61
From these studies, a clearer picture has emerged of
some of the structural features required in the C-termi-
nus for good receptor potency (Fig. 1). A heteroatom in
the meta-position is significantly potency enhancing
(compare 14 to 13). The observation that the 6-[2,3]-
dihydrobenzofuran 14 is 200 times more active than the
correspondingcarbocyclic isostere 15 suggests that
either a specific hydrogen bond contact or other polar
interaction may be important for high affinity binding.
Lipophilic substituents in the 4-position are also
potency enhancingpossibly interactingwith a hydro-
phobic pocket in the receptor bindingsite. This is
clearly evident in both the fluorobiphenyl analogue 21
which accommodates a phenyl group in the 4-position
and in the quinoline analogue 10.
10
11
12
13
14
15
16
17
18
19
20
21
22
50% at 776 mM
50% at 5.7 mM
3% at 10 mM
20% at 10 mM
60% at 10 mM
ND
0.73
0.26
1.44
1.1
ND
0.11
0.35
Pharmacokinetic parameters for key avb3 antagonists
are shown in Table 2. Selected compounds were dosed
orally and by intravenous injection to conscious dogs.
The benzodioxole 11 was well absorbed (F=30%) and
exhibited moderate clearance and modest half-life. In
contrast, the correspondingdihydrobenzofuran analo-
gue 14 underwent more rapid plasma clearance with
lower oral bioavailability. Oral bioavailability, plasma
clearance, and half-life were all substantially improved
29
ND
0.29
2.2
19% at 10 mM ꢀ0.49
29% at 10 mM
19% at 10 mM
6% at 10 mM
3% at 10 mM
30% at 10 mM
28% at 10 mM
1.15
0.06
0.43
3.7
1.8
0.57
0.49
0.31
>3.7
0.45
0.91
Figure 1. b-Alanine 3-aryl substituents.

34
P. J. Coleman et al. / Bioorg. Med. Chem. Lett. 12 (2002) 31–34
Table 2. Pharmacokinetic parameters for selected avb3 antagonists in
D. J.; Drake, F. H.; Bradbeer, J. N.; Mathur, A.; Erhard,
K. F.; Newlander, K. A.; Ross, S. T.; Salyers, K. L.; Smith,
B. R.; Miller, W. H.; Huffman, W. F.; Gowen, M. J. Pharm.
Exp. Ther. 1999, 291, 612.
dogsa
Entry
F (%)
Cl (mL/min/kg)
T1=2 h
5. (a) Meissner, R. S.; Perkins, J. J.; Duong, L. T.; Hartman,
G. D.; Hoffman, W. F.; Huff, J. R.; Ihle, N. C.; Leu, C.-T.;
Nagy, R. M.; Naylor-Olsen, A.; Rodan, G. A.; Rodan, S. B.;
Whitman, D. B.; Wesolowski, G. A.; Duggan, M. E. Bioorg.
Med. Chem. Lett. 2002, 12, 25. (b) For the first paper in this
series see: Duggan, M. E.; Duong, L. T.; Fisher, J. E.; Hamill,
T. G.; Hoffman, W. F.; Huff, J. R.; Ihle, N. C.; Leu, C.-T.;
Nagy, R. M.; Perkins, J. J.; Rodan, S. B.; Weslowski, G.;
Whitman, D. B.; Zartman, A. E.; Rodan, G. A.; Hartman,
G. D. J. Med. Chem. 2000, 43, 3736.
1
>10
1
30
6
44
12
42
34
25
33
15
20
1.0
0.5
1.2
0.7
1.5
0.8
10
11
14
19
20
aCompounds dosed orally as aqueous solutions at 1 mpk and by iv at
0.2 mpk.
6. Scarborough, R. M. Curr. Med. Chem. 1999, 6, 971.
7. For the preparation of acid 6 and representative coupling
and hydrolysis conditions, see ref 5.
for compound 19. The more potent substituted 3-fluoro-
phenyl analogues such as 20 had attenuated pharmaco-
kinetic profiles.14
8. (a) Davies, S. G.; Ichihara, O. Tetrahedron: Asymmetry
1991, 2, 183. (b) Davies, S. G.; Garrido, N. M.; Ichihara, O.;
Walters, I. A. S. J. Chem. Soc., Chem. Commun. 1993, 1153.
9. (a) 3-Pyridinyl: Rico, J. G.; Lindmark, R. J.; Rogers, T. E.;
Bovy, P. R. J. Org. Chem. 1993, 58, 7948. (b) 5-Benzodioxole:
Zablocki, J. A.; Rico, J. G.; Garland, R. B.; Roger, T. E.;
Williams, K.; Schretzman, L. A.; Rao, S. A.; Bovy, P. R.;
Tjoeng, F. S.; Lindmark, R. J.; Toth, M. V.; Zupec, M. E.;
McMakins, D. E.; Adams, S. P.; Miyano, M.; Markos, C. S.;
Milton, M. N.; Paulson, S.; Herin, M.; Jacqmin, P.; Nichol-
son, N. S.; Panzer-Knodle, S. G.; Haas, N. F.; Page, J. D.;
Szalony, J. A.; Taite, B. B.; Salyers, A. K.; King, L. W.;
Campion, J. G.; Feigen, L. P. J. Med. Chem. 1995, 38, 2378.
(c) 3-Quinoline: Duggan, M. E.; Meissner, R. S.; Perkins, J. J.
WO 99/30709. (d) Dihydrobenzofuranyl: Coleman, P. J.;
Hutchinson, J. H.; Hunt, C. A.; Lu, P.; Delaporte, E.; Rush-
more, T. Tetrahedron Lett. 2000, 41, 5803.
In conclusion, we have identified a new class of highly
potent, selective avb3 antagonists based on a non-pepti-
dic RGD motif. We have identified several novel 3-sub-
stituted b-amino acids that function in these integrin
antagonists as potent aspartic acid mimetics. Impor-
tantly, polar substituents in the meta-position of these 3-
aryl substituents greatly enhance receptor binding. b-
Amino acids that possess 3-heterobicyclics (dihy-
drobenzofuran, benzoxazolidone, and quinoline) and 3-
fluorophenyl b-substituents are noteworthy. In parti-
cular, the 3-fluorophenyl derivative 19 displayed the
best pharmacokinetic profile of the compounds studied.
Further improvements in potency have been realized in
this series by inclusion of additional 4-aryl substituents.
Relatingthese structural modifications to effects on
physicochemical properties and pharmacokinetics is
currently under study.
10. The SPAV3 assay utilizes purified human recombinant
avb3
and
4-[2-(2-aminopyridin-6-yl)ethyl]benzoyl-2-(S)-
4-125iodo phenylsulfonylamino-b-alanine. For a protocol see
ref 5b. For preparation of the radioligand see: Hamill, T. G.;
Duggan, M. E.; Perkins, J. J. J. Labelled Compd. Radiopharm.
2001, 44, 55.
11. Marguerie, G. A.; Plow, E. F.; Edgington, T. S. J. Biol.
Chem. 1979, 254, 5357.
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14. In the cases of compounds 10 and 19, pharmacokinetic
studies in bile-duct cannulated dogs indicated rapid biliary
clearance of unchanged drug (>70%) as the main route of
clearance.