Novel arylsulfonamides possessing sub-picomolar HIV protease activities and potent anti-HIV activity against wild-type and drug-resistant viral strains

Article abstract of DOI:10.1016/j.bmcl.2003.12.008

A novel series of P1′ chain-extended arylsufonamides was synthesiszed and evaluated for wild-type HIV protease inhibitory activity and in vitro antiviral activity against wild type virus and two protease inhibitor-resistant mutant viruses. All of the comp

Full text of DOI:10.1016/j.bmcl.2003.12.008

Bioorganic & Medicinal Chemistry Letters 14 (2004) 959–963
Novel arylsulfonamides possessing sub-picomolar HIV protease
activities and potent anti-HIV activity against wild-type and
drug-resistant viral strains
John F. Miller,* Eric S. Furfine,y Mary H. Hanlon, Richard J. Hazen, John A. Ray,
Laurence Robinson,{ Vicente Samano and AndrewSpaltenstein
GlaxoSmithKline, 5 Moore Drive, Research Triangle Park, NC 27709, USA
Received 8 August 2003; revised 24 November 2003; accepted 2 December 2003
Abstract—A novel series of P10 chain-extended arylsufonamides was synthesiszed and evaluated for wild-type HIV protease
inhibitory activity and in vitro antiviral activity against wild type virus and two protease inhibitor-resistant mutant viruses. All of
the compounds showed dramatic increases in enzyme activity as compared to the currently marketed HIV protease inhibitors
amprenavir, indinavir, and nelfinavir. In addition, significant improvements in antiviral potencies against wild type and the two
mutant viruses were also realized.
# 2003 Elsevier Ltd. All rights reserved.
The introduction of protease inhibitors into the HIV
therapeutic arsenal in the mid-1990’s dramatically
improved the treatment of HIV disease. Therapies that
combine HIV protease inhibitors (PIs) with agents
that target other aspects of the viral replication cycle
(such as nucleoside reverse transcriptase inhibitors)
yield dramatic and sustained reductions in viral load in
HIV positive patients.1 Such combination therapies,
known as highly active antiretroviral therapy or
‘HAART’, have transformed HIV into a much more
manageable disease and have therefore been adopted as
standard clinical practice. In spite of their success, the
currently marketed HIV-PIs are not without problems.
Included among their shortcomings are significant pill
burdens, gastrointestinal side effects, and long term
metabolic disturbances.2 Of particular concern is the
Figure 1.
emergence of strains of HIV that are resistant to the cur-
rent generation of PIs.3 As a result of these limitations,
a number of pharmaceutical companies are currently
engaged in a search for newer generation HIV-PIs with
improved side effect profiles and increased potency
against both wild-type and resistant viral strains.
Amprenavir is a potent, well tolerated HIV-PI that was
introduced in 1999.4 Recent efforts in our laboratories
have been focused on identifying an improved, next
generation HIV-PI by systematically modifying the side
chains attached to the amprenavir arylsulfonamide
scaffold. Toward this end, we have explored a number
of modifications of the P2, P10, and P20 positions
(Fig. 1). These efforts led to the identification of the
* Corresponding author. Tel.: +1-919-483-2750; fax: +1-919-483-
6053; e-mail: john.6.miller@gsk.com
y
Current address: Regeneron Pharmaceuticals Inc., 777 Old SawMill
River Road, Tarrytown, NY 10591, USA.
{
Current address: c/o Margaret Tisdale, GlaxoSmithKline Medicines
R&D Center, Gunnels Wood Road, Stevenage, Hertfordshire SG1
2NY, UK.
0960-894X/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2003.12.008

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J. F. Miller et al. / Bioorg. Med. Chem. Lett. 14 (2004) 959–963
Scheme 1. (a) 1.1 equiv 4-nitrophenylchloroformate, pyridine, CH2Cl2, 89%; (b) t-butylamine, MgSO4, Et2O, 3 days, 60%; (c) for 5a: LDA, N-(2-
chloroethyl)dibenzylamine, THF, ꢀ20 ꢁC to 0 ꢁC, 20%; for 5b: LDA, 1-bromo-4-chlorobutane, THF, ꢀ20 ꢁC to 0 ꢁC, 95%; (d) NaN3, 1:1 DMF/
H2O, 100 ꢁC, 75%; (e) 1:1 TFA/CH2Cl2, then 1N aqueous NaOH, 86%; (f) 1.0 equiv 3, DIEA, MeCN, 77%; (g) H2 (40 psi), 5% Pd(C), MeOH,
98%; (h) 1.1 equiv 4-cyano-2,2-dimethylbutanal, NaBH(OAc)3, 9:1 THF/AcOH, 75%; (i) 1.1 equiv ArSO2Cl, 10 mol% DMAP, DIEA, THF, 81–
89%; for 10a Ar=4-nitrophenyl, for 10b Ar=3-nitrophenyl, for 10c Ar=3,4-methylenedioxyphenyl; (j) H2 (40 psi), Raney Ni, 2 M NH3 in MeOH,
79–91%; (k) 1.1 equiv methyl chloroformate, DIEA, THF, 82–98%; (l) 1.2 equiv 5a, NaBH(OAc)3, 3:1 THF/AcOH, 70%; (m) 1.1 equiv 3,4-
methylenedioxybenzenesulfonyl chloride, 10 mol% DMAP, DIEA, THF, 89%; (n) 5% HCOOH/MeOH, 10% Pd(C), 68%; (o) 1.1 equiv methyl
chloroformate, DIEA, THF, 95%; (p) 1.2 equiv 5b, NaBH(OAc)3, 4:1 THF/AcOH, 61%; (q) 1.1 equiv 3,4-methylenedioxybenzenesulfonyl chloride,
10 mol% DMAP, DIEA, THF, 88%; (r) 1.2 equiv KCN, DMSO, 60 ꢁC, 42%; (s) H2 (40 psi), Raney Ni, 2 M NH3 in MeOH, 89%; (t) 1.0 equiv
methyl chloroformate, DIEA, THF, 93%.
stereochemically defined bis-THF subunit shown in
Figure 2 as the optimal fragment at the P2 position.
This moiety, which was originally reported by Ghosh and
co-workers, yields dramatic improvements in both enzyme
inhibitory activity and in vitro antiviral potency.5 The
improved enzyme activity is attributed to two additional
hydrogen bonds between the bis-THF ring oxygens and
Asp 29 and Asp 30 backbone NH’s of the enzyme.
with bound inhibitors suggests the possibility of achiev-
Figure 2.
More recently, we have attempted further optimization
of this series by exploring alternate substitution patterns
on the P20 phenylsulfonyl group while simultaneously
elaborating the P10 moiety. Specifically, we were inter-
ested in tethering polar substituents to the P10 isobutyl
sidechain. Our rationale here was the possibility of
introducing additional enzyme–inhibitor binding inter-
actions and the potential to favorably modulate
physiochemical properties such as aqueous solubility.
Examination of HIV protease X-ray crystal structures
ing hydrogen bond interactions between a distal P10
polar group and the guanidine side-chain of Arg 8. A
viable synthetic plan would allow for the introduction
of several different P10 terminal groups as well as vari-
ation of the tether chain-length in order to optimize any
potential newinteractions. This paper describes the
synthesis of a series of P10 amino derivatives that have
the bis-THF group in the P2 position. We have addres-
sed 3 structural variables: the P20 phenylsulfonyl sub-
stitution pattern, the nature of the P10 amino capping

J. F. Miller et al. / Bioorg. Med. Chem. Lett. 14 (2004) 959–963
961
Scheme 2. (a) For 16a: 1.1 equiv acetyl chloride, DIEA, THF, 86%; for 16b: 1.1 equiv methanesulfonyl chloride, DIEA, THF, 97%; for 16c: 5.0
equiv methyl isocyanate, THF, 77%; (b) LDA, 1-bromo-3-chloropropane, ꢀ20 ꢁC to 0 ꢁC, 70%; (c) ethylene glycol, PTSA toluene, reflux, 75%; (d)
KCN, DMSO, 100 ꢁC, 18 h, 85%; (e) HCl, H2O/THF, 70 ꢁC, 4.5 h; (f) H2 (40 psi), 10% Pd(C), MeOH, 98%; (g) 1.2 equiv 18, NaBH(OAc)3, DMF/
THF/AcOH, 55%; (h) 1.10 equiv 3,4-methylenedioxybenzenesulfonyl chloride, 10 mol% DMAP, DIEA, THF, 86%; (i) 1:1 TFA/CH2Cl2, then 1 N
aqueous NaOH, 100%; (j) 1.1 equiv 3, DIEA, MeCN, 76%; (k) H2 (40 psi), Raney Ni, 2 M NH3 in MeOH, 89%; (l) 1.1 equiv methyl chloroformate,
DIEA, THF, 89%.
group, and the length of the P10 tether (Fig. 2).
In addition, we present enzyme inhibition data for wild
type HIV protease and in vitro antiviral data for
wild type virus and two drug-resistant mutant viruses.
The synthesis of the 6-carbon methyl carbamate 20 was
achieved using an alternate approach in which the bis-
THF group was introduced at a later stage in the route
(Scheme 2). The requisite aldehyde building block 18
was prepared using a straightforward 4-step sequence.
Aldimine 4 was subjected to LDA alkylation with 1-
bromo-3-chloropropane followed by treatment with
ethylene glycol under acidic conditions to afford acetal
17. Intermediate 17 was converted to cyanoaldehyde 18
by reaction with KCN at elevated temperature and then
cleavage of the acetal protecting group. The BOC-pro-
tected azide 7 was subjected to catalytic reduction of the
azide group followed by reductive alkylation with alde-
hyde 18. The resulting secondary amine was sulfony-
lated with 3,4-methylenedioxybenzenesulfonyl chloride
and then treated with TFA to give amine 19. Acylation
of 19 with PNP carbonate 3 followed by Raney nickel
reduction of the cyano group and acylation with methyl
chloroformate afforded compound 20.
Syntheses of compounds containing 4, 5, and 7 carbons
in the P10 chain all proceed through primary amine
intermediate 9 (Scheme 1). The route begins with com-
mercially available epoxide 6 which was converted to
amino azide 8 by ring opening with NaN3 followed by
removal of the BOC protecting group. The bis-THF
subunit was introduced by acylation with the PNP-car-
bonate derivative of optically pure alcohol 2.5b Com-
pound 8 was subjected to catalytic reduction to afford
intermediate 9. Reductive alkylation of 9 with 4-cyano-
2,2-dimethylbutanal6 followed by sulfonylation with the
appropriate sulfonyl chloride and then Raney nickel
reduction afforded primary amines 10a–c. Acylation
of 10a–c with methyl chloroformate gave carbamate
derivatives 11a–c.
Table 1 shows enzym0e activities and in vitro antiviral
amino arylsulfonamides. Also
The preparation of 4 and 7 carbon derivatives 13 and 15
required aldehyde building blocks 5a and 5b which were
prepared by alkylation of aldimine 4 with the appro-
priate electrophile. Conversion of 9 to dibenzyl pro-
tected amine 12 was achieved by reductive alkylation
with aldehyde 5a followed by sulfonylation with 3,4-
methylenedioxybenzenesulfonyl chloride.7 Deprotection
of 12 by catalytic transfer hydrogenation gave the pri-
mary amine which was then acylated with methyl
chloroformate to afford 13. Reductive alkylation of 9
with chloroaldehyde 5b followed by sulfonylation gave
a primary chloride which was reacted with KCN to
afford cyano derivative 14. Raney nickel reduction fol-
lowed by acylation with methyl chloroformate gave 15.
The preparation of 5-carbon derivatives with 3 alternate
amino capping groups from amine 10c is shown at the
top of Scheme 2.
data for 10 newP1
shown are analogous data for the currently marketed
PIs amprenavir, indinavir, and nelfinavir for compar-
ison. The wild-type enzyme inhibition data is based on a
radioligand binding competition assay.8 The antiviral
IC50s for wild-type HIV virus (HXB2)9 and two multi-
PI resistant viruses (EP13 and D545701)10 were
determined in an MT4 cell line.11 The most obvious
observation is that all of the compounds are dramati-
cally more potent in the enzyme assay than any of the 3
currently marketed PIs shown in the table. The inhibitor
binding constants range from 100 to 4000 times lower
than the Ki for amprenavir. With the exception of 10c,
all of the compounds showsignificant improvements in
antiviral potency against the wild type and both PI-
resistant viruses. Another general observation is that the
enzyme SAR does not necessarily parallel the antiviral

962
J. F. Miller et al. / Bioorg. Med. Chem. Lett. 14 (2004) 959–963
SAR. This is likely due to variations in physiochemical
properties that influence the efficiency of cell penetra-
tion. This effect is particularly noticeable for primary
amine 10c which is over 900 times as potent as ampre-
navir against the enzyme but shows a 3-fold drop in
wild-type antiviral potency.
chainlength on both enzyme and antiviral activity. With
an enzyme Ki of 14 fM, compound 20 is over 4000 times
as potent as amprenavir. It also shows very impressive
antiviral activities against wild-type and both PI-resis-
tant viruses. These data are certainly consistent with a
newenzyme–inhibitor binding interaction hwich is
optimal for the n=3 chainlength of compound 20.
In addition, compound 20 was subjected to pharma-
cokinetic analysis in the rat and showed an oral
bioavailability of 20% and a half-life of 1.3 h.
A comparison of compounds 11a, 11b, and 11c indicates
a modest increase in enzyme potency for the para and
meta amino derivatives as compared to the methylene-
dioxy compound. On the other hand, 11c is significantly
more potent in the wild-type antiviral assay. The effect
of the P10 amino capping group can be inferred by
comparing compounds 11c, 16a, 16b, and 16c. Acet-
amide 16a, urea 16b, and sulfonamide 16c have similar
enzyme activities and are collectively 10-fold more
potent than methyl carbamate 11c. However, 11c shows
generally improved antiviral activity versus the other
three derivatives. A comparison of compounds 11c, 13,
20, and 15 indicates a pronounced effect of the P10
In summary, we have discovered a novel series of aryl-
sulfonamide HIV-PIs that showremarkable enzyme
inhibitory activity and very potent in vitro antiviral
activity against wild-type virus and two multi-PI resis-
tant viruses0. Structural studies aimed at characterizing
any newP1 enzyme–inhibitor interactions are currently
in progress. In addition, further SAR work in the aryl-
sulfonamide series is presently being pursued. The results
of these efforts will be reported in future accounts.
Table 1. Wild-type enzyme inhibitory activity and in vitro antiviral activity for P10 amino arylsulfonamides
Entry
n
Ar
R
H
WT Enzyme Ki (pM)
HXB2 IC50 (nM)
EP13 IC50 (nM)
D545701 IC50 (nM)
10c
11a
2
2
0.061
0.26
370
31
540
46
730
95
11b
11c
13
2
2
1
3
4
2
2
2
0.21
18
36
38
100
66
0.52
4.7
1.4
1.6
5.5
23
0.16
22
40
20
0.014
0.12
15
9.2
65
15
46
16a
16b
16c
0.054
0.048
0.058
30
120
200
150
38
110
47
33
Amprenavir
Indinavir
Nelfinavir
—
—
—
—
—
—
—
—
—
57a
70a
170a
130
50
320
440
330
450
>1000
440
>1000
a Determined using an activity assay based on a fluorescently labeled peptide substrate.12

J. F. Miller et al. / Bioorg. Med. Chem. Lett. 14 (2004) 959–963
963
References and notes
A. K.; Kincaid, J. F.; Walters, D. E.; Chen, Y.; Chaudhuri,
N. C.; Thompson, W. J.; Culberson, C.; Fitzgerald,
P. M. D.; Lee, H. Y.; McKee, S. P.; Munson, P. M.;
Duong, T. T.; Darke, P. L.; Zugay, J. A.; Schleif, W. A.;
Axel, M. G.; Lin, J.; Huff, J. R. J. Med. Chem. 1996, 39,
3278.
1. (a) Palella, F. J.; Delaney, K. M.; Moorman, A. C.;
Loveless, M. O.; Fuhrer, J.; Satten, G. A.; Aschman, D. J.;
Holmberg, S. D. N. Engl. J. Med. 1998, 338, 853. (b)
Hammer, S. M.; Squires, K. E.; Hughes, M. D.; Grimes,
J. M.; Demeter, L. M.; Currier, J. S.; Eron, J. J.; Feinberg,
J. E.; Balfour, H. H.; Deyton, L. R.; Chodakewitz,
J. A.; Fischl, M. A.; Phair, J. P.; Pedneault, L.; Nguyen,
B. Y.; Cook, J. C. N. Engl. J. Med. 1997, 337, 725. (c)
Collier, A. C.; Coombs, R. W.; Schoenfeld, D. A.;
Bassett, R. L.; Timpone, J.; Baruch, A.; Jones, M.; Facey,
K.; Whitacre, C.; McAuliffe, V. J.; Friedman, H. M.;
Merigan, T. C.; Reichman, R. C.; Hooper, C.; Corey, L.
N. Engl. J. Med. 1996, 334, 1011.
2. (a) Miller, K. D.; Jones, E.; Yanovski, J. A.; Shankar, R.;
Feuerstein, I.; Falloon, J. Lancet 1998, 351, 871. (b)
Behrens, G.; Schmidt, H.; Meyer, D.; Stoll, M.; Schmidt,
R. E. Lancet 1998, 351, 1958. (c) Carr, A.; Samaras, K.;
Chisholm, D. J.; Cooper, D. A. Lancet 1998, 351, 1881.
3. (a) Boden, D.; Markowitz, M. Antimicrob. Agents
Chemother. 1998, 42, 2775. (b) Tisdale, M.; Myers, R. E.;
Maschera, B.; Parry, N. R.; Oliver, N. M.; Blair, E. D.
Antimicrob. Agents Chemother. 1995, 39, 1704.
6. Kuehne, M. U.S. Patent 4,841,045, 1989.
7. Tao, E. V. P.; Miller, W. D. U.S. Patent 5,387,681, 1995.
8. Hanlon, M. H.; Porter, D. J. T.; Furfine, E. S.; Carter, L.;
Danger, D.; Shu, A.; Kaldor, I.; Miller, J. F.; Samano,
V. A.; Spaltenstein, A.; Turner, E. Biochemistry, manuscript
in review.
9. The wild-type virus HXB2 was derived from the pHXB2-
D molecular clone. For further details, see: Fischer, A. G.;
Collati, E.; Ratner, L.; Gallo, R. C.; Wong-Stall, F.
Nature (London) 1985, 316, 262.
10. EP13 and D545701 viruses are multi-protease inhibitor
resistant viruses that contain the following mutations
relative to the consensus sequence of wild type virus. EP13:
protease- M46I, L63P, A71V, V82F/L, I84V; no reverse
transcriptase mutations. D545701: protease- L10I, L19Q,
K20R, E35D, M36I, S37N, M46I, I50V, I54V, I62V, L63P,
A71V, V82A, L90M; reverse transcriptase- E28K, K32E,
V35I, T39S/T, E40D/V/Y/F, M41M/L, K43E, Y181Y/C.
11. Daluge, S. M.; Purifoy, D. J. M.; Savina, P. M.; St. Clair,
M. H.; Parry, N.R.; Dev, I. K.; Novak, P.; Ayers, K. M.;
Reardon, J. E. Antimicrob. Agents Chemother. 1994, 38,
1590.
4. (a) Noble, S.; Goa, K. L. Drugs 2000, 60, 1383. (b) Fung,
H. B.; Kirschenbaum, H. L.; Hameed, R. Clinical
Therapeutics 2000, 22, 549.
5. (a) Ghosh, A. K.; Thompson, W. J.; Fitzgerald, P. M. D.;
Culberson, C.; Axel, M. G.; McKee, S. P.; Huff, J. R.;
Anderson, P. S. J. Med. Chem. 1994, 37, 2506. (b) Ghosh,
12. Toth, M. V.; Marshall, G. R. Int. J. Pep. Protein Res.
1990, 36, 544.