Design and synthesis of 6-substituted amino-4-oxa-1-azabicyclo[3,2,0]heptan-7-one derivatives as cysteine proteases inhibitors

Article abstract of DOI:10.1016/S0960-894X(02)00765-5

A series of 6-substituted amino-4-oxa-1-azabicyclo[3,2,0]heptan-7-one compounds was designed and synthesized as a new class of inhibitors for cysteine proteases cathepsins B, L, K, and S. One compound (5S,6S)-6-(N-benzyloxycarbonyl-L-phenylalanyl) amino-4

Full text of DOI:10.1016/S0960-894X(02)00765-5

Bioorganic & Medicinal Chemistry Letters 12 (2002) 3413–3415
Design and Synthesis of 6-Substituted Amino-4-oxa-1-
azabicyclo[3,2,0]heptan-7-one Derivatives as
Cysteine Proteases Inhibitors
Nian E. Zhou,a Deqi Guo,a Jadwiga Kaleta,a Enrico Purisima,b Robert Menard,b
Ronald G. Miceticha and Rajeshwar Singha,*
aSynPhar Laboratories, currently NAEJA Pharmaceutical Inc., 4290-91A Street, Edmonton, Alberta, Canada T6E 5V2
bNational Research Council Canada, BRI, 6100 Royalmount Ave, Montreal, Quebec, Canada H4P 2R2
Received 5 July 2002; accepted 26 August 2002
Abstract—A series of 6-substituted amino-4-oxa-1-azabicyclo[3,2,0]heptan-7-one compounds was designed and synthesized as a
new class of inhibitors for cysteine proteases cathepsins B, L, K, and S. One compound (5S,6S)-6-(N-benzyloxycarbonyl-l-phenyl-
alanyl) amino-4-oxa-1-azabicyclo[3,2,0]heptan-7-one showed excellent cathepsin L and K inhibition activity with IC50 at a low
nanomolar range.
# 2002 Published by Elsevier Science Ltd.
The cysteine proteases cathepsin B, L, K, and Smay be
involved in diseases such as osteoporosis, cancer meta-
stasis, rheumatoid arthritis, and infectious diseases.1
These proteases are implicated as an important targets
for the development of inhibitors as potential ther-
apeutic agents.2 Several types of chemical functionality
have served as the central pharmacophore for cysteine
proteases inhibitors,3 such as aldehydes,4 nitriles,5
a-ketocarbonyl compounds,6 halomethyl ketones,7 acyl-
oxymethyl ketone,8 epoxide,9 vinyl sulfones,10 cyclo-
propenone,11 and cyclohexanone.12 We have reported
position,14 the Cbz-Phe fragment was selected as a sub-
stituent at the C6 position of the 4-oxa-1-azabicy-
clo[3,2,0]heptan-7-one ring. We have found that the
Cbz-Phe substitution at position 6 of 4-oxa-1-azabicy-
clo[3,2,0]heptan-7-one enhances the S2 and S3 subsite
interaction with the papain enzyme. The side chain of
Phe occupies the S2 subsite and the Cbz group interacts
with the S3 subsite. A comparison of the modeling
structure of (5S,6S)-6-(N-benzyloxycarbonyl-l-pheny-
lalanyl) amino-4-oxa-1-azabicyclo[3,2,0]heptan-7-one
(Fig. 1) as a tetrahedral intermediate complex with
papain and the X-ray structure of a substrate (Cbz-Phe-
Ala-NH-methyl) complex with papain showed good
superposition.
that
3-substituted-4-oxa-1-azabicyclo[3,2,0]heptan-7-
one derivatives exhibited good cysteine protease inhibi-
tory activity.13 Further to optimize and enhance the
activity, we studied the interaction of these inhibitors
with the papain and cathepsin B enzyme crystal struc-
tures. Molecular modeling studies suggested that the
1-N atom in 4-oxa-1-azabicyclo[3,2,0]heptan-7-one ring
can be involved in hydrogen-bonding to a protonated
Histidine in the active site of cysteine proteases (Fig. 1).
This binding may weaken the lactamic bond and acti-
vate the four member ring towards acylation of the thiol
of the cysteine residue in cysteine proteases. Since
papain has a high specificity for phenylalanine at the P2
*Corresponding author. Tel.: +1-780-462-4044; fax: +1-780-461-
0196; e-mail: rsingh@naeja.com
Figure 1. Schematic presentation of the modeling interactions of
compound 3 as a tetrahedral intermediate with papain.
0960-894X/02/$ - see front matter # 2002 Published by Elsevier Science Ltd.
PII: S0960-894X(02)00765-5

3414
N. E. Zhou et al. / Bioorg. Med. Chem. Lett. 12 (2002) 3413–3415
The synthesis of 6-substituted amino-4-oxa-1-azabicy-
clo[3,2,0]heptan-7-one compounds is shown in Scheme
1. The intermediate 1 was prepared according to the
known method15 from 6-aminopenicillanic acid. (3S)-3-
Amino-4-acetoxy-azetidin-2-one was obtained by
hydrogenation of compound 1 in the presence of Pd/C
catalyst and without purification, directly coupled with
Cbz-Phe-OH via an active ester to give compound 2.
Substitution of the acetate 2 with bromoethanol in the
presence of zinc acetate in benzene–toluene with the
azeotropic removal of water resulted in the 4-bro-
moethoxy-azetidin-2-one derivative. Cyclization of the
4-bromoethoxy-azetidin-2-one gave two optically pure
isomers (3: in 0.6% yield for two step reactions and 4: in
4% yield for two step reactions) after silica gel chroma-
tography.16 By a similar method, compounds 5 in 2.4%,
6 in 1.5% and 7 in 0.7% yield (for three-step reaction)
were also prepared. The (5S,6S) isomers of compound
5, 6 and 7 were not obtained during the purification due
to either very low formation or poor stability of such
isomers.
activity of compound 3 in comparison to compound 4
indicated the importance of the 5S stereo configuration
at C5 position. Molecular modeling suggested that in
the 5S isomer, the bridge nitrogen of oxapenam skelton
forms a hydrogen bond with His159 of papain whereas
the bridge nitrogen in the 5R isomer is too far from
His159 to form any hydrogen bonding. Replacement of
Phe with Ala (5) or phenylglycine (7) resulted in loss of
activities. Phe is generally well accepted at the P2 posi-
tion for papain type of cysteine proteases. By comparing
the activity of compound 4 and 6, the Leu residue is
more preferred for cathepsin K than Phe. These results
confirmed the molecular modeling predictions that the
C6 substitution interacts with the Ssubsite and Phe
binds at the S2 subsite.
Table 1. In vitro inhibitory activity of azetidin-2-one derivatives with
cysteine proteases
Compd
IC50 (mM)
The inhibitory activities of these compounds against
cathepsin B, L, K, and Swere determined according to
the procedure described in the literature17 by using Cbz-
Phe-Arg-AMC for cathepsin B, L and K and Cbz-Val-
Val-Arg-AMC for cathepsin S. The inhibitory activities
of the compounds are shown in Table 1. Compound 3
showed excellent cathepsin L and K inhibitory activities
with IC50 values at low nanomolar level. The better
Cath B
Cath L
Cath K
Cath S
3
4
5
6
7
12.2
1.76
>50
30
0.004
0.085
38
0.6
1.83
0.005
1.63
>2.5
0.1
n.d.a
0.077
0.43
>2.5
0.26
n.d.a
>50
an.d., not determined.
Scheme 1.

N. E. Zhou et al. / Bioorg. Med. Chem. Lett. 12 (2002) 3413–3415
3415
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