A practical synthesis of (S) 3-tert-butoxycarbonylamino-2-oxo-2,3,4,5-tetrahydro-1,5-benzodiazepine-1- acetic acid methyl ester as a conformationally restricted dipeptido-mimetic for caspase-1 (ICE) inhibitors

Article abstract of DOI:10.1016/S0960-894X(02)00107-5

A simple and versatile method for the synthesis of (S) 3-tert-butoxycarbonylamino-2-oxo-2,3,4,5-tetrahydro-1,5-benzodiazepine-1- acetic acid methyl ester (4), a dipeptide mimetic, has been developed. The regioselective functionalization of the N1 and N5 r

Full text of DOI:10.1016/S0960-894X(02)00107-5

Bioorganic & Medicinal Chemistry Letters 12 (2002) 1225–1227
A Practical Synthesis of (S) 3-tert-Butoxycarbonylamino-2-oxo-
2,3,4,5-tetrahydro-1,5-benzodiazepine-1-acetic Acid Methyl Ester
as a Conformationally Restricted Dipeptido-Mimetic for
Caspase-1 (ICE) Inhibitors
David J. Lauffer* and Michael D. Mullican
Vertex Pharmaceuticals, Inc., 130 Waverly Street, Cambridge, MA 02139-4211, USA
Received 10 January 2002; accepted 30 January 2002
Abstract—A simple and versatile method for the synthesis of (S) 3-tert-butoxycarbonylamino-2-oxo-2,3,4,5-tetrahydro-1,5-benzo-
diazepine-1-acetic acid methyl ester (4), a dipeptide mimetic, has been developed. The regioselective functionalization of the N1 and
N5 ring nitrogens and the C3 amino group is demonstrated in the synthesis ofan interleukin-1 b converting enzyme inhibitor 13.
# 2002 Elsevier Science Ltd. All rights reserved.
The development ofsmall molecule inhibitors ofcaspases
as therapeutics has been a target ofintense research.
Interleukin-1b converting enzyme (ICE, caspase-1) a
were: (1) preservation ofthe key interactions ofthe
inhibitor 1 with ICE, particularly the hydrogen bonding
pattern observed in the crystal structure ofICE/ 1,12 (2)
ease ofsynthesis ofthe optically active scaffold, and (3)
synthetic versatility ofthe scaffold, specifically ofr
access to P3 ofthe ICE inhibitor binding site via the N5
ring nitrogen ofthe benzodiazepine ring.
1
member ofthe caspase family activates interleukin-1b
(IL-1b) and IL-18, important mediators ofinflamma-
tion and infectious diseases.2 The tetra-peptide 1,3
a
potent ICE inhibitor, blocks the release ofmature IL-1 b
from human whole blood stimulated with heat-killed
Staphylococcus aureus with an IC50 of4 mM.4 Recently,
we reported that prophylactic intravenous (iv) treatment
with peptide-based irreversible ICE inhibitors in a
collagen-induced mouse model ofarthritis significantly
delayed the onset ofinflammation and reduced its
severity.5 Compound 1 was found to be poorly suited for
therapeutic uses, as is the case with many peptide derived
agents. We and others have directed our efforts towards
overcoming these shortcomings.6À11 For instance,
pyridazinodiazepine ICE inhibitors (2) show good oral
bioavailability in rodents and dogs7,11 and were reported
to be efficacious when orally dosed in a collagen-
induced arthritis model in mice.11 An ICE inhibitor
has been reported containing a benzoxazepine scaffold
(3).7 In this paper, we report the synthesis of( S) 3-tert-
butoxycarbonylamino - 2 - oxo - 2,3,4,5 - tetrahydro - 1,5 -
benzodiazepine-1-acetic acid methyl ester (4), a novel
dipeptide mimetic ofthe Val-Ala moiety in the tetra-
peptide ICE inhibitor 1. The key design criteria of 4
The syntheses ofrelated enantiomerically pure benzaze-
pinones,13 1,5-benzoxazepinones,14 and 1,5-benzothia-
zepinones15 have been reported and utilized as dipeptide
mimetics.16 However, reported syntheses ofenantio-
merically pure 3-amino-1, 5-benzodiazepinones are
*Corresponding author. Fax: +1-617-444-6920; e-mail: david_lauffer@
vpharm.com
0960-894X/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(02)00107-5

1226
D. J. Lauffer, M. D. Mullican / Bioorg. Med. Chem. Lett. 12 (2002) 1225–1227
impractical and lengthy and require a late stage resolu-
tion.17 Herein, we describe a short, high yielding, and
versatile synthesis 4 from BOC-(l)-2,3-diaminopro-
pionic acid (Scheme 1). The utility of 4 as a versatile
dipeptidomimetic is demonstrated in the synthesis of 13,
a potent ICE inhibitor (Scheme 2).
BOC-(R)-2,3-diaminopropionic acid. The Ki’s of 13 and
its C(3) amino group diastereomer are 90 and 850 nM,
respectively.22
In conclusion, we have described an efficient four-step
synthesis of( S) 3-tert-butoxycarbonylamino-2-oxo-2,3,
4,5-tetrahydro-1,5-benzodiazepine-1-acetic acid methyl
ester (4), from BOC-(l)-2,3-diaminopropionic acid (5)
in >50% overall yield. We have demonstrated the
regioselective functionalization of N(1), N(5), and the
C(3) nitrogens and utilized this dipeptide mimetic in 13,
a potent ICE inhibitor. The advantage ofscaffold 4 over
previously reported scaffolds is that it allows for the
broad exploration ofthe P3, thus providing an oppor-
tunity to improve potency and physical properties of
compounds in this class.
The aromatic nucleophilic substitution reaction of
1-fluoro-2-nitrobenzene (6) and BOC-(l)-2,3-diamino-
proprionic acid (5) in the presence ofNaHCO 3 in DMF
afforded product 7.18 Reduction ofthe nitro group by
catalytic hydrogenolysis followed by water-soluble car-
bodiimide mediated cyclization gave 9 in good yield.19
Regioselective alkylation ofthe lactam nitrogen in the
presence ofbis(trimethylsilyl)amide afforded the desired
intermediate 4.20
Deprotection ofthe N-tert-butoxycarbonyl group was
achieved via anhydrous HCl to afford the amine
hydrochloride salt that was coupled with benzoic acid to
afford the N-benzoyl derivative 10. Acylation with
3-phenylpropionyl chloride followed by base hydrolysis
ofthe methyl ester gave the acetic acid derivative 11.
Reaction ofsemicarbazone 12 with the acid 11 followed
by deprotection21 afforded the desired aldehyde 13
(Scheme 2). Proton NMR ofthe penultimate hydrazone
indicated the presence ofonly one diastereomer and
confirmed by the synthesis ofthe R isomer of 4 from
Acknowledgements
We thank Mark A. Murcko, Guy W. Bemis, and Keith
P. Wilson for their collaboration in the design of 13,
Roger D. Tung, M. Woods Wannamaker, and Brian E.
Ledford for helpful comments on the manuscript, and
David J. Livingston for helpful discussions.
References and Notes
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Scheme 1. Reagents and conditions: (i) NaHCO3, DMF, 70 ꢀC, 83%;
(ii) 10% Pd/C, H2 (1 atm.); (iii) EDC, DMF, 5–20 ꢀC, 71% for steps
(ii) and (iii); (iv) LHMDS (1.1 equiv), BrCH2CO2CH3, THF, 90%.
9. Semple, G.; Ashworth, D. M.; Baker, G. R.; Batt, A. R.;
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namaker, M. W.; Ledford, B.; Murdoch, R.; Matharu, S.;
Kay, D.; Batchelor, M.; Jones, S. D.; Porritt, J.; Elbaum, D.;
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Jones, P.; Laitinen, L.; Ruuth, E.; Moos, D.; Kurzel, G. U.;
Scheme 2. Reagents and conditions: (v) HCl (g), EtOAc; (vi) benzoic
acid, HOBT, EDC, 65% for steps (v) and (vi); (vii) 3-phenylpropionyl
chloride, Et3N, CH2Cl2, 83%; (viii) 2 N NaOH, quant; (ix) HOBT,
EDC; (x) (a) 25% TFA, CH2Cl2; (b) MeOH/AcOH/37% aq formal-
dehyde (5:1:1), 20 ꢀC.

D. J. Lauffer, M. D. Mullican / Bioorg. Med. Chem. Lett. 12 (2002) 1225–1227
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on carbon (10% by weight) was taken into 100 mL ofmetha-
nol and stirred under hydrogen (1 atm) for 4 h. The catalyst
was removed by filtration through Celite and the solvent eva-
porated in vacuo to afford 11.95 g ofthe product as a brown
solid that was used without further purification: 1H NMR
(CD3OD) d 6.75–6.70 (3H, m), 6.65–6.58 (1H, m), 4.35–4.30
(1H, m), 3.6–3.38 (2H, m), 1.45 (9H, s).
(3S)-3-tert-Butoxycarbonylamino-2-oxo-2,3,4,5-tetrahydro-1H-
1,5-benzo-diazepine (9). 1-(3-Dimethyl-aminopropyl)-3-ethyl-
carbodiimide hydrochloride (8.45 g, 44.5 mmol) was added to
a cooled (0 ꢀC) solution of(2 S)-2-tert-butoxycarbonylamino-
3-(2-aminophenylamino)-propionic acid (8; 11.95 g, 40.5
mmol) in 100 mL ofDMF and stirred ofr 18 h. Aqueous
workup and purification by flash chromatography (SiO2, 30%
ethyl acetate/hexane eluant) gave 8 g (71%) ofthe product: 1H
NMR (CDCl3) d 7.78 (1H, s), 7.02–6.95 (1H, m), 6.88–6.82
(1H, m), 6.82–6.78 (1H, m), 6.75–6.70 (1H, m), 5.8–5.7 (1H,
d), 4.55–4.45 (1H, m), 3.95 (1H, s), 3.9–3.82 (1H, m), 3.48–
3.40 (1H, m), 1.45 (9H, s).
20. (S)-(3-tert-Butoxycarbonylamino-2-oxo-2,3,4,5-tetrahydro-
1,5 - benzodiazepine - 1 - acetic acid methyl ester (4). A 1.0 M
solution oflithium bis(trimethylsilyl)amide (3.4 mL, 3.4
mmol) in THF was added dropwise to a À78 ꢀC solution of 9
(0.94 g, 3.38 mmol) in 20 mL ofanhyd THF and stirred for 30
min. Methyl bromoacetate (0.44 mL, 4 mmol) was added
dropwise to the reaction mixture then warmed to room tem-
perature. The reaction was diluted with 100 mL ofethyl ace-
tate and washed with 0.3 N KHSO4 (50 mL), water (2Â50
mL), and brine. The combined organic layers were dried over
anhyd Na2SO4, filtered, and evaporated in vacuo to afford a
gum that was purified by flash chromatography (SiO2) eluting
with 3:7 EtOAc/Hex to afford 0.98 g (83%) desired product:
1H NMR (CD3OD) d 7.15–7.07 (2H, m), 6.98–6.94 (1H, m),
6.88–6.84 (1H, d), 5.62–5.55 (1H, d), 4.71–4.65 (1H, d), 4.65–
4.6 (1H, m), 4.33–4.27 (1H, d), 3.96–3.90 (1H, m), 3.78 (3H, s),
3.44–3.37 (1H, m), 1.4 (9H, s).
17. Slade, J.; Stanton, J. L. US Patent 4,477,464, 16 October,
1984. Claremon, D. A.; Liverton, N.; Smith, G. R.; Selnick, H.
G. US Patent 5,726,171, 10 March, 1998.
18. (2S)-2-tert-Butoxycarbonylamino-3-(2-nitrophenyl-amino)-
propionic acid (7). (2S)-2-tert-Butoxycarbonylamino-3-amino-
propionic acid (5; 10 g, 49 mmol), 2-fluoronitrobenzene (6; 5.7
mL, 54 mmol), and NaHCO3 (8.25 g, 98 mmol) were stirred in
DMF (100 mL) at 70 ꢀC under a N2 atmosphere. After 18 h,
the reaction was subjected to aqueous workup to give an orange
amorphous solid (yield 12.64 g, 83%) which was used without
1
further purification: H NMR (CD3OD) d 8.15–8.10 (1H, d),
7.54–7.48 (1H, t), 7.13–7.08 (1H, d), 6.73–6.65 (1H, t), 4.45–4.35
(1H, m), 3.9–3.8 (1H, dd), 3.65–3.55 (1H, dd), 1.45 (9H, s).
19. (2S)-2-tert-Butoxycarbonylamino-3-(2-aminophenyl-amino)
propionic acid (8). (2S)-2-tert-Butoxycarbonylamino-3-(2-
nitrophenyl-amino)propionic acid (7; 12.65 g) and palladium
21. Graybill, T. L.; Dolle, R. E.; Helaszek, C. T.; Miller, R. E.;
Ator, M. A. Int. J. Peptide Res. 1994, 44, 173.
22. Margolin, N.; Raybuck, S. A.; Wilson, K. P.; Chen, W.;
Fox, T.; Gu, Y.; Livingston, D. J. J. Biol. Chem. 1997, 272, 7223.