Design, Synthesis, and Evaluation of a Pyrrolinone-based Matrix Metalloprotease Inhibitor

Article abstract of DOI:10.1021/ol000254p

formula presented A pyrrolinone-based hydroxamate matrix metalloprotease inhibitor, (-)-1, has been designed and synthesized. Enzymatic assay revealed that (-)-1 inhibited three of the ten matrix metalloprotease enzymes examined and as such represents a n

Full text of DOI:10.1021/ol000254p

ORGANIC
LETTERS
2000
Vol. 2, No. 24
3809-3812
Design, Synthesis, and Evaluation of a
Pyrrolinone-Based Matrix
Metalloprotease Inhibitor
,†
Amos B. Smith, III,* Thomas Nittoli,† Paul A. Sprengeler,† James J.-W. Duan,‡
,†
Rui-Qin Liu,‡ and Ralph F. Hirschmann*
Department of Chemistry, UniVersity of PennsylVania, Philadelphia, PennsylVania
19104, and DuPont Pharmaceuticals Company, Route 141 and Henry Clay Road,
Wilmington, Delaware 19880-0500
smithab@sas.upenn.edu
Received August 31, 2000
ABSTRACT
A pyrrolinone-based hydroxamate matrix metalloprotease inhibitor, (−)-1, has been designed and synthesized. Enzymatic assay revealed that
(−)-1 inhibited three of the ten matrix metalloprotease enzymes examined and as such represents a new, potentially important lead structure.
Research in our laboratory has focused on the development
of novel nonpeptidyl scaffolds that mimic â-turns, â-sheets/
strands, and helices, the three principle types of secondary
structure found in peptides and proteins.1 In the area of
â-sheets/strands we designed the 3,5-linked (nitrogen dis-
placed) pyrrolinone scaffold that directly substitutes on a per
residue basis for R-amino acids (except proline and glycine).2-7
Importantly, this structural motif derived from the D-amino
acids maintains both the spacial orientation of the amino acid
side chains and the capacity to form intermolecular hydrogen
bonds with the receptor or enzyme.3
The advantage of nonpeptidyl peptidomimetics, in general,
is their ability to resist degradation by proteases and to
possess additional favorable pharmacokinetic properties as
a result of reduced solvation.4 These critical features were
demonstrated in our research on HIV-1 protease, wherein
we designed a pyrrolinone-based inhibitor more potent in
† University of Pennsylvania
‡ DuPont Pharmaceuticals
(3) Smith, A. B., III; Keenan, T. P.; Holcomb, R. C.; Sprengeler, P. A.;
Guzman, M. C.; Wood, J. L.; Carroll, P. J.; Hirschmann, R. J. Am. Chem.
Soc. 1992, 114, 10672.
(4) Hirschmann, R.; Smith, A. B., III; Sprengeler, P. A. New PerspectiVes
in Drug Design; Dean, P. M., Jolles, G., Newton, C. G., Eds.; Academic
Press: New York, 1995; p 1.
(5) Smith, A. B., III; Akaishi, R.; Jones, D. R.; Keenan, T. P.; Guzman,
M. C.; Holcomb, R. C.; Sprengeler, P. A.; Wood, J. L.; Hirschmann, R.;
Holloway, M. K. Biopolymers 1995, 37, 29.
(6) (a) Smith, A. B., III; Benowitz, A. B.; Guzman, M. C.; Sprengeler,
P. A.; Hirschmann, R.; Schweiger, E. J.; Bolin, D. R.; Nagy, Z.; Campbell,
R. M.; Cox, D. C.; Olson, G. L. J. Am. Chem. Soc. 1998, 120, 12704. (b)
Smith, A. B. III; Benowitz, A. B.; Sprengeler, P. A.; Barbosa, J.; Guzman,
M. C.; Hirschmann, R.; Schweiger, E. J.; Bolin, D. R.; Nagy, Z.; Campbell,
R. M.; Cox, D. C.; Olson, G. L. J. Am. Chem. Soc. 1999, 121, 9286.
(7) Smith, A. B., III; Wang, W.; Sprengeler, P. A.; Hirschmann, R. J.
Am. Chem. Soc., submitted for publication.
(1) (a) Hirschmann, R. Proceedings of the 2nd Japan Symposium on
Peptide Chemistry. Pept. Chem. 1992, 466. (b) Hirschmann, R. Joseph
Rudinger Award Lecture. Peptides 1996, 3.
(2) (a) Smith, A. B., III; Hirschmann, R.; Pasternak, A.; Akaishi, R.;
Guzman, M. C.; Jones, D. R.; Keenan, T. P.; Sprengeler, P. A.; Darke, P.
L.; Emini, E. A.; Holloway, M. K.; Schleif, W. A. J. Med. Chem. 1994,
37, 215. (b) Smith, A. B., III; Hirschmann, R.; Pasternak, A.; Guzman, M.
C.; Yokoyama, A.; Sprengeler, P. A.; Darke, P. L.; Emini, E. A.; Schleif,
W. A. J. Am. Chem. Soc. 1995, 117, 11113. (c) Smith, A. B., III;
Hirschmann, R.; Pasternak, A.; Yao, W.; Sprengeler, P. A.; Holloway, M.
K.; Kuo, L. C.; Chen, Z.; Darke, P. L.; Schleif, W. A. J. Med. Chem. 1997,
40, 2440. (d) Smith, A. B., III; Guzman, M. C.; Sprengeler, P. A.; Keenan,
T. P.; Holcomb, R. C.; Wood, J. L.; Carroll, P. J.; Hirschmann, R. J. Am.
Chem. Soc. 1994, 116, 9947. (e) Smith, A. B., III; Holcomb, R. C.; Guzman,
M. C.; Keenan, T. P.; Sprengeler, P. A.; Hirschmann, R. Tetrahedron Lett.
1993, 34, 63.
10.1021/ol000254p CCC: $19.00 © 2000 American Chemical Society
Published on Web 11/05/2000

vitro than Crixivan, against a clinically significant mutant
strain, which proved to be orally bioavailable in dogs
(13%).2c In addition, we designed and synthesized a com-
petent pyrrolinone inhibitor of renin,2a,5 a high affinity
peptide-pyrrolinone hybrid ligand for the class II major
histocompatibility complex (MHC) protein HLA-DR1,6 and
most recently a polypyrrolinone â-turn mimic.7 The latter
exploited the D,L-alternating (i.e., heterochiral) polypyrroli-
none structural motif. Given these successes, we sought to
extend further the scope of the pyrrolinone scaffold. We
report here the design, synthesis, and enzymatic evaluation
of a pyrrolinone-based matrix metalloprotease inhibitor.
displaced nitrogens in bispyrrolinone 1, but the trajectories
of the attached hydrogens were quite similar, providing the
potential for intermolecular hydrogen bonding with the
enzyme. This low energy conformation was then docked into
the active site of the MMP-1 enzyme (Figure 2). As
Matrix metalloproteases (MMPs) comprise a family of
more than 20 zinc-containing enzymes that are involved in
the degradation of extracellular connective tissue.8 They have
been implicated in a number of inflammatory and degenera-
tive diseases including arthritis, multiple sclerosis, Guillian
Barre′ syndrome, stroke, and cancer.9 Thus, this enzyme class
represents an attractive target for the design and synthesis
of selective, small molecule inhibitors that can modulate the
severity of the underlying disease.10
The prospective MMP inhibitor 1 (Figure 1) was designed
on the basis of the Roche peptidyl inhibitor Ro-31-4724 (2),
reported to have an IC50 of 9 nM against human fibroblast
collagenase (MMP-1).11 The X-ray structure of peptidyl
inhibitor 2 cocrystallized with MMP-1 (2.2 Å resolution)12
clearly indicated that the hydroxamate in the peptidyl
inhibitor effectively coordinates to the active-site zinc(II),
and that the S1′-S3′ pockets of the enzyme are occupied
with isobutyl side chains at P1′ and P2′ and a methyl at P3′.
Energy minimization of 1 employing Macromodel 5.013 led
to a low energy model; the overlay of this model with the
Roche inhibitor (2) held in the enzyme bound conformation
(Figure 1) revealed remarkable similarities, as anticipated
from our prior work.2-7 The backbone, hydroxamate, side
chains, and carbonyls all overlaid very well, while some
differences were observed at the C-terminus. Not surpris-
ingly, the amide nitrogens of 2 did not overlay with the
Figure 2. (a) Bispyrrolinone 1 (red) docked into the active site of
human fibroblast collagenase (MMP-1) and (b) a cartoon of the
active site for clarity.
anticipated, we observed coordination of the hydroxmate to
the active site zinc(II), hydrogen bonding of both the
pyrrolinone carbonyls and NH’s with the enzyme backbone,
and S1′-S3′ occupation with the respective side chains as
observed with peptidyl inhibitor 2.
From the synthetic perspective, we envisioned construction
of 1 to exploit our base-mediated pyrrolinone ring synthetic
protocol,2-7 followed by functional group manipulation
(Scheme 1). Bispyrrolinone 9 would thus require 2 equiv of
known amino ester (-)-76 and 1 equiv of aldehyde 6.
(8) (a) Birkedal-Hansen, H. J. Oral Pathol. 1988, 17, 445. (b) Birkedal-
Hansen, H. Curr. Opin. Cell Biol. 1995, 7, 728. (c) Emonard, H.; Grimaud,
J. A. Cell. Mol. Biol. 1990, 36, 131. (d) Murphy, G.; Docherty, A. J. P.
Am. J. Respir. Cell Mol. Biol. 1992, 7, 120. (e) Baramova, E.; Foidart, J.
Cell Biol. Int. 1995, 19, 239. (f) Borkakoti, N. Prog. Biophys. Mol. Biol.
1998, 70, 73. (g) Johnson, L. L., Dyer, R., Hupe, D. J. Curr. Opin. Chem.
Biol. 1998, 2, 466. (h) Shapiro, S. D.; Senior, R. M. Am. J. Respir. Cell
Mol. Biol. 1999, 20, 1100.
(9) (a) Chandler, S.; Miller, K. M.; Clements, J. M.; Lury, J.; Corkill,
D.; Anthony, D. C. C.; Adams, S. E.; Gearing, A. J. H. J. Neuroimmunol.
1997, 72, 155. (b) Toi, M.; Ishigaki, S.; Tominaga, T. Breast Cancer Res.
Treat. 1998, 52, 113. (c) Michaelides, M. R.; Curtin, M. L. Curr. Pharm.
Des. 1999, 5, 787. (d) Lukes, A.; Mun-Bryce, S.; Lukes, M.; Rosenberg,
G. A. Mol. Neurobiol. 1999, 19, 267. (e) Kieseier, B. C.; Seifert, T.;
Giovannoni, G.; Hartung, H. Neurology 1999, 53, 20. (f) Curran, S.; Murray,
G. I. J. Pathol. 1999, 189, 300. (g) Whittaker, M.; Floyd, C. D.; Brown,
P.; Gearing, A. J. H. Chem. ReV. 1999, 99, 2735.
(10) Zask, A.; Levin, J. I.; Killar, L. M., Skotnicki, J. S. Curr. Pharm.
Des. 1996, 2, 624.
(11) Johnson, W. H.; Roberts, N. A.; Borkakoti, N. J. Enzyme Inhib.
1987, 2, 1.
(12) Borkakoti, N.; Winkler, F. K.; Williams, D. H.; D’Arcy, A.;
Broadhurst, M. J.; Brown, P. A.; Johnson, W. H.; Murray, E. J. Struct.
Biol. 1994, 1, 106.
(13) MacroModel, ver 5.0; Still, W. C.; Mohamadi, F.; Richards, N. G.
J.; Guida, W. C.; Lipton, M., Liskamp, R.; Chang, G.; Hendrickson, T.;
Degunst, F.; Hasel, W. Department of Chemistry, Columbia University:
New York, 10027.
Figure 1. Overlay of active-site conformation of Roche compound
Ro-31-4724 (2) with model of bispyrrolinone 1.
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Org. Lett., Vol. 2, No. 24, 2000

Pleasingly, hydrolysis of the dimethylacetal with TsOH at
40 °C led to the corresponding aldehyde in nearly quantitative
yield; a second pyrrolinone ring construction, again using
amino ester (-)-76 led to bispyrrolinone (-)-9. The ef-
ficiency of our iterative pyrrolinone construction protocol
was clearly demonstrated by the 77% overall yield of (-)-9
from (+)-6.
Scheme 1
Scheme 3
Hydrolysis of the dimethylacetal in (-)-9 was next
achieved with 1 N HCl at 40 °C. Unfortunately, oxidation
of the derived aldehyde to the corresponding carboxylic acid
by employing a variety of different conditions (e.g., Jones,
sodium chlorite, PCC, etc.) proceeded only in low yield.
Careful examination of the product mixture revealed that the
pyrrolinone rings were not stable to the oxidation conditions.
To circumvent this problem, (-)-9 was protected as the bis
Cbz derivative (+)-10 (Scheme 4); although this operation
led to a less reactive pyrrolinone ring, the acetal proved
resilient to hydrolysis. We therefore decided to remove the
SEM group first. Treatment of (+)-10 with TsOH and
methanol at 40 °C furnished alcohol (+)-11. A two-step
oxidation with Dess-Martin periodinane15 and then with
sodium chlorite produced the C-terminal acid in 81% yield.
Esterification followed by removal of the acetal (TsOH in
wet THF at 40 °C) furnished the intermediate bispyrrolinone
aldehyde; immediate oxidation with sodium chlorite led to
(+)-13. We speculate that the steric congestion in (+)-10,
emanating from the SEM and Cbz groups, inhibits acetal
hydrolysis. Completion of the synthesis was achieved via
coupling (+)-13 with O-benzyl hydroxylamine (EDCI and
HOBt), followed by hydrogenolysis with Pd/BaSO4;16 the
overall yield of (-)-1 for the two steps was 51%. Since the
carboxylic acid functionality is also known to coordinate to
the active-site zinc(II) in matrix metalloproteases,9g (+)-13
was subjected to hydrogenolysis to furnish acid (-)-14
(Scheme 5).
Synthesis of aldehyde 6 (Scheme 2) began with Evans
alkylation of (S)-propionyloxazolidinone (+)-3 with prenyl
bromide to furnish oxazolidinone (+)-4 in 64% yield (>98%
ee). Reduction with lithium borohydride in wet THF,14
followed by protection of the hydroxyl with 2-(trimethylsi-
lyl)ethoxymethyl chloride (SEM-Cl) led to the SEM ether,
which was subjected to ozonolysis to furnish aldehyde (+)-
6; the overall yield from (+)-3 was 39%.
Scheme 2
To construct monopyrrolinone (+)-8, amino ester (-)-76
was condensed with aldehyde (+)-6 (Scheme 3); dehydration,
followed by base-promoted pyrrolinone ring formation
(KHMDS) then furnished (+)-8 in 93% yield (two steps).
(15) (a) Dess, D. B.; Martin, J. C. J. Org. Chem. 1983, 48, 4155. (b)
Dess, D. B.; Martin, J. C. J. Am. Chem. Soc. 1991, 113, 7277. (c) Ireland,
R. E.; Liu, L. J. Org. Chem. 1993, 58, 2899.
(14) Penning; T. D.; Djuric, S. W.; Haack, R. A.; Kalish, V. J.; Miyashiro,
J. M.; Rowell, B. W.; Yu, S. S. Synth. Comm. 1990, 20, 307.
(16) Nikam, S. S.; Kornberg, B. E., Johnson, D. R.; Doherty, A. M.
Tetrahedron Lett. 1995, 36, 197-200.
Org. Lett., Vol. 2, No. 24, 2000
3811

carboxylic acid (-)-14, on the other hand, failed to inhibit
the 10 proteases assayed, a result presumably of the shorter
overall chain length compared to (-)-1 and/or the known
reduced affinity of the carboxylate to zinc(II) compared to
the hydroxamate functionality.9g,17
Scheme 4
In summary, we have designed and synthesized a pyrroli-
none-based inhibitor for a series of matrix metalloproteases.
The synthesis reveals that unprotected pyrrolinone rings can
be susceptible to oxidation but that acylation alleviates this
propensity. Although only modest activity was observed, we
believe that (-)-1 represents an important new lead struc-
ture.18 In addition, these results further demonstrate that
excellent peptide mimicry can be achieved with the pyrroli-
none scaffold. Studies to increase the potency of the
pyrrolinone-based MMP inhibitors by increasing the size and
hydrophobicity of the P1′ side chain, exploiting our recently
disclosed solid-support polypyrrolinone synthetic protocol
for focused library construction,19 are currently underway
in our laboratory.
Acknowledgment. Financial support was provided by the
National Institutes of Health (National Institute of Allergy
and Infectious Diseases) through grant AI-42010. We also
thank Dr. G. Furst and Mr. J. Dykins, Directors of the
University of Pennsylvania Spectroscopic Facilities, for
assistance in obtaining NMR spectra and high-resolution
mass spectra, respectively.
Although bispyrrolinone (-)-1 was designed for the
MMP-1 isozyme, inhibitory activity was observed only for
gelatinase (MMP-2), matrilysin (MMP-7), and the membrane
type 2 matrix metalloprotease (MMP-15) with Ki values of
2.9, 6.4, and 6.8 µM, respectively. The bispyrrolinone
Supporting Information Available: Spectroscopic and
analytical data as well as experimental procedures for all
intermediates. This material is available free of charge via
the Internet at http://pubs.acs.org.
OL000254P
(17) Castelhano, A. L.; Billedeau, R.; Dewdney, N.; Donnelly, S.; Horne,
S.; Kurz, L. J.; Liak, T. J.; Martin, R.; Uppington, R.; Yuan, Z.; Krantz, A.
Bioorg. Med. Chem. Lett. 1995, 5, 1415.
Scheme 5
(18) We speculate that although all of the anticipated hydrogen bonds
in the modeled active-site of MMP-1 with (-)-1 were within 3 Å in length,
optimal geometries were not achieved, thereby reducing their strength.
Optimal hydrogen bonding occurs when the donor, hydrogen, and acceptor
atoms are colinear. Streyer, L. Biochemistry, 3rd ed.; W. H. Freeman: New
York, 1988; p 8.
(19) Smith, A. B., III; Liu, H.; Okumura, H.; Favor, D. A.; Hirschmann,
R. F. Org. Lett. 2000, 2, 2041.
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