Design, synthesis, and evaluation of a pyrrolinone - Peptide hybrid ligand for the class II MHC protein HLA-DR1 [20]

Full text of DOI:10.1021/ja982973t

12704
J. Am. Chem. Soc. 1998, 120, 12704-12705
Scheme 1
Design, Synthesis, and Evaluation of a
Pyrrolinone-Peptide Hybrid Ligand for the Class II
MHC Protein HLA-DR1
Amos B. Smith, III,* Andrew B. Benowitz, Mark C. Guzman,
Paul A. Sprengeler, and Ralph Hirschmann*
Department of Chemistry, UniVersity of PennsylVania
Philadelphia, PennsylVania 19104
Edwin J. Schweiger, David R. Bolin, Zoltan Nagy,
Robert M. Campbell, Donald C. Cox, and Gary L. Olson*,†
Department of Inflammation/Autoimmune Diseases
Hoffmann-La Roche Inc., Nutley, New Jersey 07110
ReceiVed August 19, 1998
In 1992, we reported the design and synthesis of nonpeptide
â-strand peptidomimetics based on the 3,5,5-pyrrolin-4-one
scaffold.1a These polypyrrolinones, which are stable to proteases,
adopt extended conformations both in solution and in the solid
state that mimic the antiperiplanar disposition of the carbonyls
and side chains of peptidal â-strands. More recently, we success-
fully exploited the pyrrolinone scaffold to construct potent,
bioavailable, small-molecule inhibitors of the HIV-1 protease.1b-d
In this communication, we disclose the design and synthesis of a
pyrrolinone-based peptide hybrid which proved to be a competent
ligand for the rheumatoid arthritis-associated class II major
histocompatibility complex protein HLA-DR1, thereby further
demonstrating the utility of the pyrrolinone scaffold.
The primary role of the class II major histocompatibility
complex (MHC), an extracellular polymorphic2 membrane-bound
protein widely dispersed on B lymphocytes, macrophages, and
other specialized antigen-presenting cells,3 is to present antigenic
peptides4 derived from extracellular pathogens and toxins for
† Current address: Provid Research, 10 Knightsbridge Road, Piscataway,
NJ 08854.
(1) (a) 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. (b) 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. (c) 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. (d) 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 and
references therein.
(2) Kappes, D.; Strominger, J. L. A. ReV. Biochem. 1988, 57, 991.
(3) (a) Unanue, E. R.; Allen, P. M. Science 1987, 236, 551. (b) Steinman,
R. M. Annu. ReV. Immunol. 1991, 9, 271.
(4) (a) Adorini, L.; Muller, S.; Cardinaux, F.; Lehmann, P. V.; Falcioni,
F.; Nagy, Z. A. Nature 1988, 334, 623. (b) Rudensky, A. Y.; Preston-Hurlburt,
P.; Hong, S.-C.; Barlow, A.; Janeway, C. A., Jr. Nature 1991, 353, 622. (c)
Hunt, D. F.; Hanspeter, M.; Dickinson, T. A.; Shabanowitz, J.; Cox, A. L.;
Sakaguchi, K.; Appella, E.; Grey, H. M.; Sette, A. Science 1992, 256, 1817.
(d) Chicz, R. M.; Urban, R. G. Immunol. Today 1994, 15, 155. (e) Engelhard,
V. H. Annu. ReV. Immunol. 1994, 12, 181.
(5) Todd, J. A.; Acha-Orbea, H.; Bell, J. I.; Chao, N.; Fronek, Z.; Jacob,
C. O.; McDermott, M.; Sinha, A. A.; Timmerman, L.; Steinman, L.; McDevitt,
H. O. Science 1988, 240, 1003.
(6) (a) Brown, J. H.; Jardetzky, T. S.; Gorga, J. C.; Stern, L. J.; Urban, R.
G.; Strominger, J. L.; Wiley, D. C. Nature 1993, 364, 33. (b) Stern, L. J.;
Brown, J. H.; Jardetzky, T. S.; Gorga, J. C.; Urban, R. G.; Strominger, J. L.;
Wiley, D. C. Nature 1994, 368, 215. (c) Jardetzky, T. Structure 1997, 5, 159.
(7) Hammer, J.; Valsasnini, P.; Tolba, K.; Bolin, D.; Higelin, J.; Takacs,
B.; Sinigaglia, F. Cell 1993, 74, 197.
(8) (a) Hammer, J.; Belunis, C.; Bolin, D.; Papadopoulos, J.; Walsky, R.;
Higelin, J.; Danho, W.; Sinigaglia, F.; Nagy, Z. Proc. Natl. Acad. Sci. U.S.A.
1994, 91, 4456. (b) Bolin, D.; Campbell, R. Hoffmann-La Roche Inc.,
unpublished results.
(9) The structure assigned to each new compound is in accord with its
infrared, 500-MHz 1H NMR, and 125-MHz 13C NMR spectra, as well as
appropriate parent ion identification by high-resolution mass spectrometry.
(10) (a) Karady, S.; Amato, J. S.; Weinstock, L. M. Tetrahedron Lett. 1984,
25, 4337. (b) Seebach, D.; Fadel, A. HelV. Chim. Acta 1985, 68, 1243. (c)
Seebach, D.; Aebi, J. D.; Gander-Coquoz, M.; Naef, R. HelV. Chim. Acta
1987, 70, 1194.
inspection by CD4 T cells of the immune system. Inhibition of
this process holds considerable promise for the treatment of
numerous autoimmune diseases.5
The recent X-ray crystal analysis by Wiley and co-workers6a,b
revealed that the HLA-DR1 MHC class II protein binds the
influenza virus hemagglutinin peptide fragment PKYVKQN-
TLKLAT (HA 306-318)6b in an extended, â-strand-like confor-
mation. The presence of anchor residues at positions 1 and 9
(Scheme 1) in conjunction with a complex network of hydrogen
bond interactions between the HA 306-318 backbone and the
amino acid side chains in the DR1 MHC binding site are
postulated to account for a large portion of the binding energy.4,7
On the basis of alanine scans of HA 306-318 and related 7-mer
peptide ligands,8 we postulated that the side chains of the VKQN
sequence in HA 306-318 could be altered without significantly
affecting HLA-DR1 binding. Indeed, control peptide PKYGLLL-
TLKLAT bound to the HLA-DR1 protein with an IC50 of 176
nM, comparable to an IC50 of 89 nM for the native HA 306-318
peptide. On the basis of this information, we speculated that
bispyrrolinone 2a (Scheme 1) might serve as a viable replacement
for the amide backbone of the VKQN sequence of HA 306-318.
Molecular modeling revealed that the global low-energy confor-
mations of the bispyrrolinone segment of 1 and the corresponding
region of HA 306-318 bound to HLA-DR1 would be quite similar.
Moreover, with the exception of a hydrogen bond between Gln-9
of HLA-DR1 and the N-H at position 4 of HA 306-318, the
hydrogen bonding network was predicted to remain intact. Integral
to this design was the expectation that Fmoc derivative 2b would
permit use of Fmoc-based solid-phase peptide synthesis to
construct the hybrid ligand 1.
10.1021/ja982973t CCC: $15.00 © 1998 American Chemical Society
Published on Web 11/20/1998

Communications to the Editor
J. Am. Chem. Soc., Vol. 120, No. 48, 1998 12705
The synthesis of bispyrrolinone 2b (Scheme 1) was envisioned
to involve two iterations of our now well-established pyrrolinone
synthetic protocol,1 beginning with the construction of amino ester
3 (Scheme 2). Treatment of (+)-7,9 available from D-leucine via
Scheme 4
Scheme 2
the Seebach/Karady oxazolidinone,1,10 with OsO4 and NMO11
provided a mixture (1.3:1) of diastereomeric diols. Protection as
the acetonides followed by separation and transfer hydrogenoly-
sis12 of the less polar diastereomer (+)-8a9 gave (-)-3.9
Preparation of aldehyde 4 (Scheme 3) entailed ozonolysis of
(+)-9,9,13 followed by protection of the derived aldehyde and
transfer hydrogenolysis12 to provide amino ester (-)-5.9 Con-
densation with aldehyde (+)-6,9,14 to form the imine, followed in
turn by treatment with KHMDS (4 equiv) and acid gave aldehyde
(-)-4;9 the overall yield was 57% (five steps).
Scheme 3
Construction of the TLKLAT peptide and incorporation of
bispyrrolinone (-)-2b proceeded smoothly on Wang resin
employing Fmoc-based solid-phase peptide synthesis. Removal
of the N-terminal Fmoc group, addition of the remaining amino
acids (PKY), and liberation of the pyrrolinone-peptide hybrid
from the resin with concomitant protecting group removal
completed the synthesis of 1.18
A second iteration of the pyrrolinone protocol employing (-)-3
and (-)-4, followed by protection of the pyrrolinone nitrogens
with the Alloc moiety, furnished (+)-109 (Scheme 4). Removal
of the acetonide and conversion of the primary alcohol in turn to
the tosylate, bromide, and azide provided (+)-12.9 Reduction
(Ph3P)15 followed by in situ protection of the resulting amine using
FmocOSu then gave (+)-13.9 A two-step oxidation with the
Dess-Martin periodinane16 and Jones reagent, following removal
of the SEM protecting group, provided keto-acid (+)-14.9
Removal of the Alloc protecting groups17 completed the synthesis
of bispyrrolinone (-)-2b.9
Affinity binding experiments19 revealed that pyrrolinone-
peptide hybrid 1 was indeed a competent ligand for HLA-DR1,
having an IC50 of 137 nM, compared with 89 nM for the HA
306-318 peptide and 176 nM for the control peptide. The similar
binding affinity of 1 relative to HA 306-318 is intriguing given
that the N-terminal pyrrolinone, due to displacement of the
backbone N-H, is unable to form a hydrogen bond with Gln-9 of
the class II MHC HLA-DR1 protein in contrast to HA 306-318.
The similar binding affinity is also of interest when one considers
that the carbonyl at position 2 of 1, required only for synthetic
purposes, is not expected to be as effective a hydrogen bond
acceptor as the backbone amide at position 2 of HA 306-318.
Notwithstanding these considerations, the similar binding affinities
of 1 and the native and control peptides suggest that additional
compensating interactions may be formed. We hope to address
this issue through X-ray crystallographic studies of the hybrid
ligand bound to the class II MHC protein HLA-DR1.
Acknowledgment. Financial support was provided by Hoffmann
LaRoche, Inc., and the National Institutes of Health (Institute of General
Medical Sciences) through Grant GM-41821. We also thank Dr. Charles
Belunis, Diana Gaizband, Dr. Jeanmarie Guenot, and Raymond C.
Makofske for their contributions to this program.
Supporting Information Available: Spectroscopic and analytical data
for 1, 2b, 3-6, 8a, 8b, and 10-14 and selected experimental procedures
(17 pages, print/PDF). See any current masthead page for ordering
information and Web access instructions.
(11) VanRheenan, V.; Kelley, R. C.; Cha, D. Y. Tetrahedron Lett. 1976,
17, 1973.
(12) Jackson, A. E.; Johnstone, R. A. W. Synthesis 1976, 685.
(13) Prepared from D-leucine in a manner analogous to that for (+)-7.
(14) Prepared from (R)-5-methyl-2-(2-methylpropyl)-4-hexen-1-ol via treat-
ment with SEMCl and i-Pr2NEt (87% yield) followed by ozonolysis and
treatment with Ph3P (74% yield).
(15) Vaultier, M.; Knouzi, N.; Carrie´, R. Tetrahedron Lett. 1983, 24, 1983.
(16) (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.
(17) (a) Guibe´, F.; Dangles, O.; Balavoine, G. Tetrahedron Lett. 1986, 27,
2365. (b) Dangles, O.; Guibe´, F.; Balavoine, G.; Lavielle, S.; Marquet, A. J.
Org. Chem. 1987, 52, 4984.
(18) The assigned structure was in accord with its amino acid analysis as
well as appropriate parent ion identification by mass spectrometry.
(19) The binding affinity of the pyrrolinone-peptide hybrid 1 to HLA-
DR1 was assessed using a scintillation proximity assay (SPA) protocol in
which an 125I-peptide was competitively displaced from HLA-DR1 (isolated
from LG2 cells, purified, and coupled to SPA beads via an LB3.1 antibody).
See: Ito, K.; Bian, H.-J.; Molina, M.; Han, J.; Magram, J.; Saar, E.; Belunis,
C.; Bolin, D. R.; Arceo, R.; Campbell, R.; Falcioni, F.; Vidovic, D.; Hammer,
J.; Nagy, Z. A. J. Exp. Med. 1996, 183, 2635.
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