Spirocyclic nonpeptide glycoprotein IIb-IIIa antagonists. Part 2: Design of potent antagonists containing the 3-azaspiro[5.5]undec-9-yl template

Article abstract of DOI:10.1016/S0960-894X(01)00216-5

The synthesis and biological activity of novel glycoprotein IIb-IIIa anatagonists containing 3-azaspiro[5.5]undec-9-yl nucleus are described. The potent activity of these compounds as platelet aggregation inhibitors demonstrates the utility of the monoaza

Full text of DOI:10.1016/S0960-894X(01)00216-5

Bioorganic & Medicinal Chemistry Letters 11 (2001) 1293–1296
Spirocyclic Nonpeptide Glycoprotein IIb–IIIa Antagonists.
Part 2: Design of Potent Antagonists Containing the
3-Azaspiro[5.5]undec-9-yl Template
A. Pandey,a J. Seroogy,a D. Volkots,a M. S. Smyth,a J. Rose,a M. M. Mehrotra,a
J. Heath,a G. Ruhter,b T. Schottenb and R. M. Scarborougha,*
aCOR Therapeutics, Inc., Department of Medicinal Chemistry and Biology, South San Francisco, CA 94080, USA
bLilly Research Laboratories, Hamburg, Germany
Received 17 January 2001; revised 16 March 2001
Abstract—The synthesis and biological activity of novel glycoprotein IIb–IIIa anatagonists containing 3-azaspiro[5.5]undec-9-yl
nucleus are described. The potent activity of these compounds as platelet aggregation inhibitors demonstrates the utility of the
monoazaspirocyclic structure as central template for nonpeptide RGD mimics. # 2001 Elsevier Science Ltd. All rights reserved.
Efforts to discover orally active GPIIb–IIIa antagonists
with appropriate pharmaceutical properties for treating
thrombotic disease have been numerous.1 We recently
identified the 3,9-diazaspiro[5.5]undecane template as a
suitable conformationally constrained nucleus for the
preparation of potent and specific GPIIb–IIIa antago-
nists such as CT51688. (See ref 2: Part 1, preceding
paper.) Based on these results we initiated efforts to
explore the potential utility of the monoazaspirocyclic
template to identify potent GPIIb–IIIa antagonists
which would afford more flexibility in appending basic
and acidic pharmacophore elements.
ladium-catalyzed coupling reactions. A third series of
compounds was prepared with direct linkage of the
piperidine nitrogen to pyridines, and we again used Pd-
catalysis for the synthesis (Schemes 3 and 4).2
Also prepared were compounds in which ether- or
amine-containing linkages were utilized. (Scheme 4).
Finally, a series in which the basic pharmacophore
groups were attached through the carboxyl group of the
9-azaspiro[5.5]undecane-3-carboxylate template were
prepared as shown in Scheme 5. The cyano group was
converted to the corresponding amidines via hydroxyl-
amine addition followed by hydrogenolysis.
The synthesis of compounds 15–17 was accomplished
by coupling the spironucleus with various a- and b-
substituted carboxylic acid chains (Scheme 3). Synthesis
of the pyridyl-9-oxa-3-aza-spiro[5.5]undecane nucleus
was achieved by spiroannulation methodology, using 4-
formylpiperidine and methyl vinylketone under basic
conditions (Scheme 4). The carboxyl group of the 9-
azaspiro[5.5]undecane template was coupled to the basic
bipiperidyl and pyridylpiperazine groups followed by
acylation or alkylation of the piperidine ring with acid
chloride esters or alkyl bromoesters, respectively
(Scheme 5).
To begin the investigation of this nucleus, we prepared
the ethyl 9-azaspiro[5.5]undecane-3-carboxylate tem-
plate as described in the literature.3 The amide-linked
and direct-linked benzamidine moieties were incorpo-
rated as shown in Schemes 1 and 2. The direct-linked
benzamidine series (Scheme 2) was prepared using pal-
All of the synthesized spirocyclic analogues were
assayed for their ability to inhibit the aggregation of
ADP-stimulated human platelets and to block the
*Corresponding author. Tel.: +1-650-244-6822; fax: +1-650-244-
9287; e-mail: rscarborough@corr.com
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00216-5

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A. Pandey et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1293–1296
binding of fibrinogen to purified human GPIIb–IIIa.2
Although only two analogues of the amide-linked ben-
zamidine series were prepared, both displayed weak
inhibitory activity (Table 1). Replacement of the amide
linkage with the direct-linked benzamidine (analogues 3
and 4, Table 2) led to enhancement in potency of
approximately 7-fold in the platelet aggregation assay
and a 160-fold in the fibrinogen binding assay (3 vs 2).
Compound 3 was significantly more potent in the bind-
ing assay compared to the platelet aggregation assay
and this may be due to plasma protein binding by this
analogue in the aggregation assay in PRP as has been
noted in other GPIIb–IIIa inhibitors.1,4
Our next series of analogues utilized the less basic direct
linked pyridyl group, a modification chosen to improve
both the potency and pharmacokinetic properties.
Incorporation of an a-amino substitution at the carboxyl
group in the direct linked pyridine series dramatically
Scheme 1. Synthesis of amide-linked benzamidines 1 and 2: (a) 2 N
HCl, 60 ꢀC; (b) p-cyanobenzoyl chloride, Et3N, DCM; (c) b-alanine
ethyl ester, HATU, DIEA, DMF, rt; (d) (i) NH2OHꢁ HCl, EtOH,
Et3N, rt; (ii) AcOH, Ac2O, rt; (iii) 10% Pd/C, H2, 1 atm; (e) LiOH,
THF, H2O.
Scheme 3. Synthesis of compounds 5–14: (a) 4-bromopyridine, S-
BINAP, NaOtBu, Pd2(dba)3, toluene, 90 ꢀC, 65%; (b) EDC, HOBt,
DIEA, DMF, rt, 84%; (c) 3 N HCl, 60 ꢀC, 2 h, 35%.
Scheme 4. Synthesis of pyridyl-9-oxo-3-azaspiro[5.5]undecane ana-
logues: (a) MVK, KOH, EtOH, reflux, 50%; (b) H2, Pd/C; (c) 4-bro-
mopyridine, S-BINAP, NaOtBu, Pd2(dba)3, toluene, 90 ꢀC, 65%;
(d) NaBH4, THF, 65%; (e) ethyl diazoacetate, [Rh(OAc)2]2, 64%;
(f) LiOH, THF, H2O; (g) b-alanine ethyl ester or aminopropionate,
NaCNBH3, AcOH, rt.
Scheme 2. Synthesis of compounds 3 and 4: (a) p-bromobenzonitrile,
S-BINAP, NaOtBu, Pd2(dba)3, toluene, 90 ꢀC, 65%; (b) b-alanine
ethyl ester or aminopropionate, HATU, DIEA, DMF, rt, 84%;
(c) (i) NH2OH HCl, EtOH, Et3N, rt; (ii) AcOH, Ac2O, rt; (iii) 10%
Pd/C, H2, 1 atm; (d) LiOH, THF, H2O, 63%.
.

A. Pandey et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1293–1296
1295
enhances the potency as seen for analogues 6–9 (Table
3). Incorporation of the 3,5-dimethylisoxazol-4-yl sul-
fonamide functionality afforded analogue 7 (CT51819),
which displayed 300-fold enhancement in activity in
platelet aggregation assay and 830-fold enhancement in
binding assay relative to the unsubstituted amine ana-
logue 5 (Table 3). These observations are consistent
with the results by a number of other groups and our
own observations.4ꢂ6 Of interest, sulfonamide 7 was
also found to be a potent inhibitor of ADP-induced
platelet aggregation of murine platelets with an IC50
value of 45 nM. Thus, this antagonist is unique relative
to many potent GPIIb–IIIa antagonists which typically
are weak inhibitors of murine GPIIb–IIIa and therefore
attractive for study in murine thrombosis models.7
The a-carbamate substituted analogue (10) resulted in a
10-fold loss in potency relative to butane-sulfonamide
analogue (9) in the PRP assay and retained similar
potency in binding assay (Table 3). There have been
extensive SARs at the position b to the carboxylic acid
terminus in several different series of GPIIb–IIIa inhi-
bitors.1,8 It is also reported that activity is greatly
Table 3. Effect of carboxylic acid segment modifications
Analogues
R
R0
ELISA
Fg/GPIIb–IIa
IC50 (mM)a
Human PRP
IC50 (mM)a
5
6
7
8
NH2
NHCBz
H
H
H
H
H
H
CH3
Ph
0.830
0.002
0.001
0.001
0.003
0.008
0.616
30
12.2
2.7
0.040
0.177
0.176
1.3
NHSO2isoxazole
NHSO2Tos
NHSO2nBut
NHCO2nBut
H
9
10
11
12
14.3
77
H
Scheme 5. Synthesis of compounds 18–23: (a) bipiperidyl, BOP,
DIEA, DMF, rt; (b) 50% TFA/DCM, 60%; (c) ClCO(CH2)n-
CO2Et, DCM, DIEA, 92%; (d) BrCH2CO2Et, K2CO3, DMF, 65 ꢀC;
(e) LiOH, THF, H2O; (f ) Pd-black, HCO2NH4, 85%; (g) pyridyl-
piperazine, BOP, DIEA, DMF; (h) Br(CH2)nCO2Et, DMF, DIEA,
65 ꢀC, 20%.
aSee Table 1.
Table 4. In vitro activity for compounds with 4-aminopyridine basic
unit
Table 1. In vitro activity for compounds 1 and 2
Compds
R
ELISA
Human PRP
Fg/GPIIb–IIa IC50 (mM)a
IC50 (mM)a
13
14
15
16
17
CONH(CH2)3CO2H
CONHCH2CH(Me)CH2CO2H
OCH2CO2H
0.319
0.332
0.25
0.16
50
12
4
1.1
0.89
4.5
Compds
R
ELISA
Fg/GPIIb–IIa
IC50 (mM)a
Human PRP
IC50 (mM)a
NHCH2CO2H
NHCH2CH2CO2H
1
2
OH
NHCH2CO2H
15
9.7
19.7
38.5
aSee Table 1.
The average error for the determinations was ꢃ15%.
aIC50 values expressed as the average of at least two determinations.
Table 5. In vitro activity for compounds with pyridyl-piperazine
basic unit
Table 2. In vitro activity for compounds with benzamidine basic unit
Compds
R
ELISA
Fg/GPIIb–IIa
IC50 (mM)a
Human PRP
IC50 (mM)a
Compds
R
ELISA
Fg/GPIIb–IIa
IC50 (mM)a
Human PRP
IC50 (mM)a
3
4
NH(CH2)2CO2H
NH(CH2)3CO2H
0.060
5
5.5
37.1
18
19
CH2CO2H
CH2CH2CO2H
0.364
3.45
0.117
7.9
aSee Table 1.
aSee Table 1.

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A. Pandey et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1293–1296
Table 6. In vitro activity for compounds with bipiperidyl basic unit
compounds 7 (CT51819), 8, 9, 18, and 23 as free acids
and as their ethyl ester prodrugs were evaluated in
Sprague–Dawley rats. The compounds showed low oral
bioavailability (<7%) and rapid clearance at 1 mg/kg iv
and po dose. Highly active and specific GPIIb–IIIa
inhibitors have been discovered in the 6,6-mono-
azaspirocyclic series, which extends our previous work
with the 6,6-diazaspirocyclic template. However, the
poor absorption and pharmacokinetic properties of
these inhibitors preclude their ultimate utility as oral
GPIIb–IIIa antagonists.
Compds
R
ELISA
Fg/GPIIb–IIa
IC50 (mM)a
Human PRP
IC50 (mM)a
20
21
22
23
COCH2CO2H
CO(CH2)2CO2H
CO(CH2)3CO2H
CH2CO2H
2.40
100
100
2.5
50
50
0.250
0.289
aSee Table 1.
References and Notes
enhanced by the addition of b-substituents when
antagonists contain a b-amino acid functionality.8 This
has been ascribed to favorable interaction with a
hydrophobic binding site in GPIIb–IIIa. We have uti-
lized a similar approach to study the effect of b-sub-
stitution in our spirocyclic series. The b-phenyl
substituted analogue (12) resulted in 15-fold loss of
potency in PRP and 500-fold in the ELISA assay rela-
tive to the unsubstituted analogue 3 (Table 3). The
replacement of b-phenyl with the smaller CH3 residue
(11) enhanced the activity but overall b-substitution
resulted in decreased activity relative to unsubstituted
analogue 3 (Table 3). Analogues (15–17) with ether- or
amine-linked carboxylic acid groups did not display
enhanced potency, and this approach was not pursued
further (Table 4).
1. (a) Scarborough, R. M. J. Med. Chem. 2000, 43, 3453. (b)
Scarborough, R. M. Curr. Med. Chem. 1999, 6, 971.
2. Smyth, M. S.; Rose, J.; Mehrotra, M. M.; Heath, J.; Ruh-
ter, G.; Schotten, T.; Seroogy; J.; Volkots, D.; Pandey, A.;
Scarborough, R. M. Bioorg. Med. Chem. Lett. 2001, 11, 1289.
3. Claremont, D. A.; Liverton, N.; Baldwin, J. J. U.S. Patent
5,451,578, 1995; Chem. Abstr. 1995, 124, 145932.
4. Su, T.; Naughton, M. A. H.; Smyth, M. S.; Rose, J. W.;
Arfsten, A. E.; McCowan, J. R.; Jakubowski, J. A.; Wyss,
V. L.; Ruterbories, K. J.; Sall, D. J.; Scarborough, R. M. J.
Med. Chem. 1997, 40, 4308.
5. Egbertson, M. S.; Chang, C. T.-C.; Duggan, M. E.; Gould,
R. J.; Halczenko, W.; Hartman, G. D.; Laswell, W. L.; Lynch,
J. J.; Lynch, R. J.; Manno, P. D.; Naylor, A. M.; Prugh, J. D.;
Ramjit, D. R.; Sitko, G. R.; Smith, R. S.; Turchi, L. M.;
Zhang, G. J. J. Med. Chem. 1994, 37, 2537.
6. (a) Xue, C.-B.; Wityak, J.; Seilecki, T. M.; Pinto, D. J.;
Batt, D. G.; Cain, G. A.; Sworin, M.; Rockwell, A. L.;
Roderick, J. J.; Wang, S.; Orwat, M. J.; Frietze, W. E.; Bos-
trom, L. L.; Liu, J.; Higley, C. A.; Rankin, F. W.; Tobin,
A. E.; Emmett, G.; Lalka, G. K.; Sze, J. Y.; DiMeo, S. V.;
Mousa, S. A.; Thoolen, M. J.; Racanelli, A. L.; Hausner, E. A.;
Reilly, T. M.; DeGrado, W. F.; Wexler, R. R.; Olsen, R. E. J.
Med. Chem. 1997, 40, 2064. (b) Xue, C. B.; Mousa, S. A.
Drugs Future 1998, 23, 707. (c) Olsen, R. E.; Sielecki, T. M.;
Wityak, J.; Pinto, D. J.; Batt, D. G.; Frietze, W. E.; Liu, J.;
Tobin, E.; Orwat, M. J.; Di Meo, S. V.; Houghton, G. C.;
Lalka, G. K.; Mousa, S. A.; Racanelli, A. L.; Hausner, E. A.;
Kapil, R. P.; Rabel, S. R.; Thoolen, M. J.; Reilly, T. M.;
Anderson, P. S.; Wexler, R. R. J. Med. Chem. 1999, 42, 1178.
(d) Mousa, S. A.; Wityak, J. Cardiovas. Drug. Rev. 1998, 16,
48.
7. Law, D. A.; Nannizzi-Alaimo, L.; Cowan, K. J.; Prasad,
K. S.; Ramakrishnan, V.; Phillips, D. R. Thromb. Haemost.
1999, 82 (Suppl.), 345.
8. Zablocki, J. A.; Rico, J. G.; Garland, R. B.; Rogers, T. E.;
Williams, K.; Schretzman, L. A.; Rao, S. A.; Bovy, P. R.;
Tjoeng, D. E.; Adams, S. P.; Miyano, M.; Markos, C. S.;
Milton, M. N.; Knodle, S. G.; Haas, N. F.; Page, J. D.; Szal-
ony, J. A.; Salyers, A. K.; King, L. W.; Campion, J. G.; Fei-
gen, L. P. J. Med. Chem. 1995, 38, 2378.
In the final series prepared, the basic pharmacophore
group was attached to the carboxyl group of the 9-
azaspiro[5.5]undecane-3-carboxylate template. We ini-
tially focused on optimizing the distance between the
carboxylate and basic functionalities. In the pyridyl-
piperazine series, the extension with a methylene unit
(19) resulted in 10-fold loss of activity in PRP and
fibrinogen binding assay relative to 18 (Table 5). In the
bipiperidyl series the amide-linked carboxylic acid ana-
logues 20–22 (Table 6) were less potent relative to the
alkyl-linked carboxylic acid analogue 23. The alkyl-
linked analogue 23 displayed 10-fold enhancement in
activity in both the PRP and binding assay compared to
20.
We also examined the integrin specificity of all active
compounds in this series, and they were found to have
IC50 values >100 mM against the vitronectin receptor,
avb3. Thus, both direct-linked benzamidine- and pyridine-
containing analogues were all highly selective towards
GPIIb–IIIa. The pharmacokinetic properties of