New, potent P1/P2-morpholinone-based HIV-protease inhibitors

Article abstract of DOI:10.1016/j.bmcl.2006.07.014

We have developed efficient synthesis of morpholinone-based cyclic mimetics of the P1/P2 portion of the HIV-1 protease inhibitor Amprenavir. This effort led to discovery of allyl- and spiro-cyclopropyl-P2-substituted inhibitors 17 and 31, both 500 times m

Full text of DOI:10.1016/j.bmcl.2006.07.014

Bioorganic & Medicinal Chemistry Letters 16 (2006) 5226–5230
New, potent P1/P2-morpholinone-based HIV-protease inhibitors
Wieslaw M. Kazmierski,a,* Eric Furfine,a Andrew Spaltensteina and Lois L. Wrightb
aGlaxoSmithKline, MV CEDD, Five Moore Drive, Research Triangle Park, NC 27709, USA
bGlaxoSmithKline, Screening & Compound Profiling, Five Moore Drive, Research Triangle Park, NC 27709, USA
Received 5 May 2006; revised 17 June 2006; accepted 5 July 2006
Available online 10 August 2006
Abstract—We have developed efficient synthesis of morpholinone-based cyclic mimetics of the P1/P2 portion of the HIV-1 protease
inhibitor Amprenavir. This effort led to discovery of allyl- and spiro-cyclopropyl—P2-substituted inhibitors 17 and 31, both 500
times more potent than the parent inhibitor 1. These results support morpholinones as novel mimetics of the P1/P2 portion of
Amprenavir and potentially of other HIV-protease inhibitors, and thus provide a novel medicinal chemistry template for optimiza-
tion toward more potent and drug-like inhibitors.
ꢀ 2006 Elsevier Ltd. All rights reserved.
Proteolytic cleavage of gag and gag-pol gene protein
products by the virally encoded aspartyl protease
(HIV-PR) is one of the key steps in the life cycle of
HIV. Several HIV-protease inhibitors (PI), such as
Amprenavir (Fig. 1), have been developed for use in
Highly Active Antiretroviral Therapy (HAART), which
is credited for a dramatic reduction in AIDS-related
mortality and morbidity.1 Nonetheless, the high pill bur-
den generally required by the early HAART regimens is
not conducive to patient compliance, and thus may re-
sult in the emergence of viral resistance. In order to ad-
dress this issue, we have been involved in a research
program directed toward PI prodrugs with improved
solubility2,3 as well as next generation PIs with increased
potency, by exploring heterocyclic mimetics for the P1/
P2 portion of Amprenavir.4–9
embarked on a medicinal chemistry program incorpo-
rating P2 substituents in the morpholinone scaffold in
order to optimize the potency in this series.4,10
To that end, we have explored a number of approaches
toward mono-, di-, and spiro-substituted morpholinones
in order to establish the SAR in this series. Herein, we
disclose chemical routes toward analogues of 1, which
enabled the discovery of potent inhibitors, such as 17
and 31.
Our initial target was the racemic 2-methyl-5-(phenyl-
methyl)-3-morpholinone, which was obtained according
to the previously published method.10 Interestingly, we
found that the Williamson cyclization of the diastereo-
meric mixture 2 resulted in the exclusive formation of
a pure (2S,5S)-2-methyl-5-(benzyl)-morpholin-3-one 3
in 34% yield (Fig. 2).11–14 While thermodynamic equili-
bration leading to a single diastereomer cannot be ruled
out as the mechanism leading to pure 3, molecular mod-
eling also supports a kinetic argument in that cyclization
of the S,R-2 diastereomer appeared energetically much
more facile, thus enabling the formation of 2S,5S-3,
while the cyclization of the S,S-2 diastereomer appears
disfavored, precluding the formation of 2R,5S-3.14,15
In the latter approach, we have explored cyclic lactams,
cyclic sulfonamides, cyclic ureas, and cyclic sulfamates
among other heterocyclic scaffolds,4–9 and in this letter
we report our findings with the morpholinone ring sys-
tem as a putative P1/P2 scaffold. Molecular modeling
supported the design rationale toward compound 1, ob-
tained from (5S)-5-(phenylmethyl)-3-morpholinone
(A = B = H) and the Amprenavir-like P10/P20 scaffold
(R1 in Fig. 1). This prototype molecule indeed exhibited
moderate potency in the HIV-1 protease enzyme assay
(Ki = 1.0 lM).10 Encouraged by this result, we
Fragment 3 was next coupled to epoxide 4 in a fashion
similar to one previously reported,4,10 yielding HIV-PR
inhibitor 5 (Ki = 38 nM, Table 1), which was 50 times
more potent than the parent compound 1.
Keywords: HIV; HIV-protease inhibitor; Amprenavir; HIV-1; Aspartyl
protease inhibitor; Mimetic; Drug; Inhibitor.
*
We already reported that in the related 3,5-(P1/P2)-di-
substituted pyrrolidone scaffold,4 the 3S,5R-isomers
Corresponding author. Tel.: +1 919 483 9462; fax: +1 919 315
0430; e-mail: wieslaw.m.kazmierski@gsk.com
0960-894X/$ - see front matter ꢀ 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.07.014

W. M. Kazmierski et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5226–5230
5227
P1'
P1
OH
H
NH2
N
O
O
N
OH
O
S
O
O
N
O
O
O
N
A
S
P2'
B
P2
P2
O
O
P1
R1
Amprenavir Ki= 0.04 nM
1, A=B=H, Ki= 1 uM
Figure 1. Structure of Amprenavir and its morpholinone P1/P2 mimetic-containing analogue 1.
chromatography, provided the individual 2S,5S and
2R,5S diastereomers 9 and 10, respectively.11
Bn
NH
Br
a
O
b
c
HO
Br
Br
NH
Interestingly, the 2R,5S diastereomer 10 turned out to
be about 10 times more potent than 2S,5R isomer 9,
revealing that stereochemical preference also existed in
the morpholinone scaffold (Table 1). Consequently, we
explored other approaches to optically pure diastereo-
mers or to easily separable mixtures of diastereomers.
Thus, the alkylation of 12 with allyl iodide proceeded
with a much higher diastereoselectivity (95:5%, Fig. 4),
yielding essentially pure 2R,5S-13,4,11 and enabling fac-
ile entry into the desired diastereomer series.
O
O
O
2
3
O
O
OH
O
N
O
S
N
N
O
O
S
4
O
O
O
5
Figure 2. (a) DIEA, (S)-phenylalaninol; (b) NaH, DMF, 0 ꢁC to rt,
1 h; 34% yield over two steps; (c) NaH, DMF, 0!80 ꢁC, 1–3 h, 4, 94%.
We also examined the alkylation of advanced intermedi-
ate 14, a TBDMS-protected product of the condensa-
tion of morpholinone 1110 and epoxide 4, with allyl
iodide.10,16 Although the alkylation of 14 proceeded
with modest diastereoselectivity, the resulting mixture
of 15 (2R,5S-morpholin-3-one) and 16 (2S,5S-morphol-
in-3-one) was readily separable on silica gel, affording
inhibitors 17 and 18 after treatment of 15 and 16 with
1 M TBAF in THF. Similarly, the alkylation of 14 with
benzyl bromide also yielded a mixture of diastereomers,
affording the 2R,5S-morpholin-3-one 19, after chroma-
tographic separation of the minor 2S,5S component
(Fig. 4).11 The stereochemical outcome of these alkyla-
tions is likely controlled by A1,3 allylic strains in 12
and 14.17
Table 1. Inhibitory potencies of final products in the HIV-PR assay18
Compound
Ki [nM]
1
5
1000
38
9
490
55
10
17
18
19
21
23
24
25
27
28
29
31
35
2
270
35
30
9
81
600
11
Although we found that the alkylation of 12 could pro-
vide a single diastereomer, we decided that a more prac-
tical route toward diastereomerically pure 2R,5S
morpholin-3-one-based inhibitors (Fig. 5) would be to
utilize the easily available and scalable 2R,5S intermedi-
ate 15. Accordingly, inhibitor 21 incorporating the 2-
hydroxyethane as P2 was obtained in several steps from
alcohol 20. The conversion of 20 to mesylate 22 followed
by the treatment of the latter with lithium hexamethyldi-
silazide and deprotection with 1 M TBAF in tetra-
hydrofurane yielded novel spiro-cyclopropyl analogue
23. Furthermore, mesylate 22 was also converted to
azide 24, and the latter next reduced to 2-aminoethyl
P2-substituted inhibitor 25. Finally, 22 was converted
to nitrile 26, which served as a convenient intermediate
toward inhibitors 27–29 (Fig. 5).
5800
49
2
23
(with trans rather than cis relationship of P1 and P2 sub-
stituents) were more potent in the HIV-PR enzyme as-
say.4,5 To explore such stereochemical preference in
the morpholinone series described herein, in addition
to Williamson chemistry yielding the 2S,5S (e.g.,
2S,5S-3) diastereomer, we also needed to develop
routes toward 2R,5S diastereomers. To this end, we
explored the alkylation of p-methoxybenzyl-protected
morpholinone 6 (Fig. 3) and of Boc-protected morpholi-
none 12 (Fig. 4). The alkylation of 6 with m-CN-benzyl
bromide proceeded with a modest diastereoselectivity of
3:1, 2R,5S/2S,5S. The resulting mixture 7 was then
deprotected and coupled with sulfate 8, and after
A more than 100-fold increase in potency against the
HIV-1 PR18 was observed in the case of spiro-cyclopro-
pyl inhibitor 23 versus reference compound 1 (Table 1).

5228
W. M. Kazmierski et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5226–5230
Bn
Bn
Bn
O
OH
b, c
a
O
O
O
N
N
N
m-CN-Bn
N
MOB
S
A
MOB
O
O O
O
O
A
6
7
N
N
8
2S, 5S
2R, 5S
9
O
O
S
O
O
O
N
S
O O
10
Figure 3. (a) LHMDS, ꢀ78 ꢁC; m-CN-benzyl bromide; 74%; (b) CAN in MeCN/water, 25%; (c) P4-phosphazine, 1 equiv, ꢀ20 ꢁC, THF, cyclic
sulfate 8, 1 N aq sulfuric acid, 93% (cumulative for both diastereomers d 9 and 10).
Bn
Bn
e
a, b
O
Bn
O
O
N
N
87%
N
Boc
Boc
O
O
O
12
11
13
g, h
TBDMS
TBDMS
Bn
Bn
a, b
55%
O
O
O
O
O
O
N
N
N
N
S
S
O
O
O
O
O
O
14
15(2R,5S) and 16 (2S,5S)
i, d
36%
a, c, d
18 (2S,5S)
Bn
Bn
OH
OH
O
O
O
O
N
N
N
N
S
S
Bn
O
O
O
O
O
O
17 (2R, 5S)
19
Figure 4. (a) LHMDS, ꢀ78 ꢁC; (b) allyl iodide, 1.5 h at ꢀ78 ꢁC; (c) BnBr; (d) 1 M TBAF in THF and chromatography, 85–95%; (e) Boc2O, DMAP,
CH3CN, 96%; (g) NaH, 4, DMF, 80 ꢁC; (h) TBDMS-triflate, 79%, two steps; (i) silica gel chromatography.
This prompted synthetic explorations of spiro-cyclic
morpholinones such as 30,12,13 which afforded potent
inhibitor 31 (Fig. 6, Ki = 1.7 nM).19
found 500-fold, while analogues 23 and 27 100-fold
more potent in comparison to reference compound
1.10 Consistent with the knowledge of the inhibitor
binding site in the HIV protease, we also found out
that polar P2 residues, such as a primary amine in
28, substantially reduced the inhibitory potency. In
contrast, the isosteric inhibitor 21 (OH in place of
NH2) maintained relatively high inhibitory potency
(Table 1). We thus conclude that 2R,5S-disubstituted
morpholin-3-ones, especially those with small and
non-polar P2-substituents, can be suitable mimetics
of the P1/P2 portion of Amprenavir21 and conceivably
of other HIV-1 protease inhibitors. These findings
support the rationale for additional explorations in
the morpholinone scaffold with the goal of further
improving the potency and potentially fine-tuning
drug-like properties in this class of inhibitors. This
work will be reported in due course.
A somewhat different synthetic strategy was developed
toward the spiro-tetrahydropyran-based morpholinone
33,20 obtained in 45% yield by alkylation of intermediate
6
10,11 with bis-O-iodoethane. The only major byproduct
of this reaction, the cross-linked derivative 32, could be
easily separated and a subsequent deprotection of 33
and coupling with sulfate 8 yielded potent inhibitor 35
(Table 1).
In summary, we developed efficient chemistry toward
substituted and stereochemically defined analogues of
1. In particular, advanced intermediate 15 enabled
facile and convergent syntheses of several 2R,5S-mor-
pholin-3-ones (Table 1). Analogues 17 and 31 were

W. M. Kazmierski et al. / Bioorg. Med. Chem. Lett. 16 (2006) 5226–5230
5229
TBDMS
Bn
Bn
O
OH
R1
O
O
21
HO
f
b, c
N
15
N
R1
O
O
HO
a
O
d
20
Bn
23
Bn
TBDMS
OH
Bn
O
OH
R1
i, f
N
O
O
N
R1
N
O
g
R
R1
O
Ms-O
O
22
24
R=N3
25
R=NH2
e
Bn
R
TBDMS
f
Bn
OH
R1
O
O
O
27 CN
28 CH2NH2
29 CONH2
26
g, f
h, f
N
N
R1
R
O
N
O
Figure 5. (a) LHMDS, ꢀ78 ꢁC, 90%; (b) OsO4, NaClO4, 86%; (c) NaBH4 in MeOH (titration), 93%; (d) MsCl, DIEA, 73%; (e) NaCN, DMSO, rt,
overnight; 81%; (f) 1 M TBAF in THF, 85–95%; (g) H2/Ra-Ni, 56–67%; (h) urea hydrogen peroxide, 2 h, rt, pH = 10, 82%; (i) NaN3, DMF, 80 ꢁC,
91%.
Bn
Bn
NH
c
O
O
OH
O
N
N
S
O
O O
O
30
31
Bn
R2
+
O
Bn
a
O
6
O
N
N
MOB
O
O
O
c
b
34
R2=H
35
R2=R1
33
R2=MOB
2
32
Figure 6. (a) LHMDS, ꢀ78 ꢁC, (I–CH2–CH2–)2O, 45% of 33; (b) CAN, overnight, 33%; (c) P4-phosphazine, 1 equiv at ꢀ20 ꢁC, THF, cyclic sulfate 8
then 1 N aq sulfuric acid, 31–45%.
7. Kazmierski, W. M.; Furfine, E.; Gray-Nunez, Y.; Spal-
tenstein, A.; Wright, L. L. Bioorg. Med. Chem. Lett. 2004,
14, 5685.
8. Kazmierski, W.; Andrews, W.; Baker, C.; Bock, B.;
Cleary, D.; Furfine, E.; Gray-Nunez, Y.; Hale, M.; Hazen,
R.; Salituro, F.; Schairer, W.; Spaltenstein, A.; Tung, R.;
Wright, L. Book of Abstracts, 219th ACS National
Meeting, San Francisco, CA, March 26–30, 2000,
Abstract MEDI-126, 2000.
9. Kazmierski, W.; Furfine, E.; Spaltenstein, A.; Wright, L.
In Peptides 2002, Proceedings of the European Peptide
Symposium, 27th, Sorrento, Italy, August 31–September
6, 2002; Benedetti, E., Pedone, C., Eds.; pp 972–973.
10. Salituro, F. G.; Baker, C. T.; Court, J. J.; Deininger, D.
D.; Kim, E. E.; Li, B.; Novak, P. M.; Rao, B. G.;
Pazhanisamy, S.; Porter, M. D.; Schairer, W. C.; Tung, R.
D. Bioorg. Med. Chem. Lett. 1998, 8, 3637.
References and notes
1. Moyle, G. J. Expert Opin. Invest. Drugs 1998, 7, 1313.
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3. Kazmierski, W. M.; Bevans, P.; Furfine, E.; Porter, D.;
Spaltenstein, A.; Yang, H. 223rd ACS National Meeting,
Orlando, FL, United States, April 7–11, 2002, Abstract
MEDI-160, 2002.
4. Spaltenstein, A.; Cleary, D.; Bock, B.; Kazmierski, W.;
Furfine, E.; Wright, L.; Hazen, R.; Andrews, W.; Almond,
M.; Tung, R.; Salituro, F. Bioorg. Med. Chem. Lett. 2000,
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5. Kazmierski, W. M.; Andrews, W.; Furfine, E.; Spalten-
stein, A.; Wright, L. L. Bioorg. Med. Chem. Lett. 2004, 14,
5689.
11. Stereochemical assignments were confirmed with NOE
1H NMR spectroscopy experiments.
6. Kazmierski, W. M.; Furfine, E.; Spaltenstein, A.; Wright,
L. L. Bioorg. Med. Chem. Lett. 2002, 12, 3431.

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12. Fritch, P. C.; Kazmierski, W. M. Synthesis 1999, 1, 112.
13. Fritch, P. C.; Kazmierski, W. M. In Peptidomimetics
Protocols; Kazmierski, W. M., Ed.; Humana Press:
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A. J. Med. Chem. 1988, 31, 671.
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16. Baker, C. T.; Salituro, F. G.; Court, J. J.; Deininger, D.
D.; Kim, E. E.; Li, B.; Novak, P. M.; Rao, B. G.;
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20. Compound 33 1H NMR: (CDCl3, 300 MHz) d 7.82 (m,
4H, ar), 7.18 (m, 3H, ar), 6.87 (d, J = 8.5 Hz, 2H, ar),
5.36 (d, J = 14.7 Hz, 1H, N–CH), 3.82 (m, 8H,
OCH3+N–CH+2 · O–CH2), 3.62 (app. d, Japp. = 12.1,
1H, O–CH2–CHa), 3.54 (app. d, Japp. = 12.1 Hz, 1H, O–
CH2–CHa), 3.28 (m, 1H, CH–a), 3.00 (m, 2H, CH–b),
2.42 (m, 1H, Cq–CH), 2.25 (m, 1H, Cq–CH), 1.78
(app. d, Japp. = 14 Hz, 1H, Cq–CH), 1.64 (app. d,
Japp. = 13.1 Hz, 1H, Cq–CH). Compound 33 13C NMR
(CDCl3, 75.42 MHz): 134.3 (ar), 126.3 (ar), 125.6 (ar),
123.7 (ar), 111.1 (ar), 75.5 (Cq), 59.5 (CH2), 57.4 (CH2),
53.3 (O–CH3), 52.2 (CHa), 44.3 (CH2), 34.0 (CH2), 32.0
(CH2), 27.3 (CH2). Compound 34 1H NMR (CDCl3,
300 MHz) d 7.25 (m, 5H, ar), 5.43 (s, 1H, NH), 3.78 (m,
7H, CHa+3 · O–CH2), 2.95 (dd, J = 6.0, 13.6 Hz, 1H,
CH-CH2–phenyl), 2.74 (dd, J = 8.4, 13.6 Hz, 1H, CH–
CH2–phenyl), 2.25 (m, 2H, spiro-cyclic CH2), 1.74 (m,
apparent ddd J = 2.1, 8.1, 16.0 Hz, 2H, spiro-cyclic
CH2).
17. Norman, B. H.; Kroin, J. S. Tetrahedron Lett. 1995, 36,
4151.
18. Toth, M. V.; Marshall, G. R. Int. J. Peptide Protein Res.
1990, 36, 544.
19. Inhibitor 31 was synthesized from 0.159 g of 30,12,13
4.4 mg of NaH, and 0.20 g of 4,10 yielding 111 mg of 31
(yield 31%). Compound 31 1H NMR (CDCl3, 300 MHz) d
7.52 (d, 2H), 7.30 (m, 5H), 6.95 (d, 2H), 4.05 (m, 1H), 3.87
(3H, s), 3.60 (m, 2H), 3.16 (m, 4H), 3.0 (m, 4H), 2.18 (1H,
m), 1.97 (m, 2H), 1.60 (m, 14H), 1.23 (m, 4H).
21. Kim, E. E.; Baker, C. T.; Dwyer, M. D.; Murcko, M. A.;
Rao, B. G.; Tung, R. D.; Navia, M. A. J. Am. Chem. Soc.
1995, 117, 1181.