Synthesis and antiviral activities of the major metabolites of the HIV protease inhibitor ABT-378 (Lopinavir)

Article abstract of DOI:10.1016/S0960-894X(01)00243-8

The HIV protease inhibitor ABT-378 (Lopinavir) is metabolized rapidly and extensively by CYP-3A4 catalyzed oxidation. Three of the major metabolites identified were synthesized and their antiviral (HIV) activities determined.

Full text of DOI:10.1016/S0960-894X(01)00243-8

Bioorganic & Medicinal Chemistry Letters 11 (2001) 1351–1353
Synthesis and Antiviral Activities of the Major Metabolites of the
HIV Protease Inhibitor ABT-378 (Lopinavir)
Hing L. Sham,* David A. Betebenner, Thomas Herrin, Gondi Kumar, Ayda Saldivar,
Sudthida Vasavanonda, Akhter Molla, Dale J. Kempf, Jacob J. Plattner
and Daniel W. Norbeck
Pharmaceutical Discovery, D47B, Building AP-10, Abbott Laboratories, Abbott Park, IL 60064-6101, USA
Received 9 February 2001; accepted 14 March 2001
Abstract—The HIV protease inhibitor ABT-378 (Lopinavir) is metabolized rapidly and extensively by CYP-3A4 catalyzed oxida-
tion. Three of the major metabolites identified were synthesized and their antiviral (HIV) activities determined. # 2001 Elsevier
Science Ltd. All rights reserved.
The HIV protease inhibitor ABT-3781 (Lopinavir) is the
antiviral component of KaletraTM (approved by FDA
in September, 2000), the latest approved HIV protease
inhibitor for the treatment of human immunodeficiency
virus (HIV) infection. ABT-378 is a highly potent inhi-
bitor of HIV protease (Ki=1.3–28 pM) against wild-
type and mutant HIV protease.1,2 ABT-378 (1) was
metabolized very extensively and rapidly by liver
microsomes from mouse, rat, dog, monkey, and
humans. The predominant site of oxidative metabolism
was the cyclic urea moiety of ABT-378.3 In all five
species, the major metabolites (out of M-1–M-12) were
M-1, M-3, and M-4 (Scheme 1); M-3 and M-4 are the
epimeric C-4 hydroxy products of oxidation in the cyclic
urea moiety and M-1 is the C-4 oxo product. They were
characterized by mass and NMR spectroscopy. The
very small quantities of these metabolites isolated pre-
cluded the determination of their antiviral activities. Thus,
the synthesis of these major metabolites was undertaken to
provide materials both to confirm the reported structures
and to determine their antiviral activities.
of the key carboxylic acid 5. Coupling of known inter-
mediates 3, 4, and then with 5 would provide M-1,
which upon reduction would provide M-3/M-4.
Michael addition of the t-butyl ester of l-valine to
acrylic acid methyl ester provided 6 (94%). Hydrolysis
of the diester 6 by aqueous lithium hydroxide provided
the corresponding mono-acid, which was reacted with
disuccinimidocarbonate to give the active ester (Scheme
3). Aminolysis of the succinimidyl ester with con-
centrated ammonium hydroxide provided the amide 7
(68%, three steps). Refluxing amide 7 with carbonyldi-
imidazole in THF provided the cyclic oxo-urea 8 in 61%
yield. Trifluoroacetic acid hydrolysis of the t-butyl ester
provided the key intermediate acid 5 in 95% yield.
The synthesis of ABT-378 has been reported.4 A retro-
synthetic analysis of the synthesis is shown in Scheme 2.
The synthesis of the core diamino compound 3 has also
been reported.5,6 We envisioned that the synthesis of M-
1 and M-3/M-4 could be accomplished by the synthesis
*Corresponding author. Fax: +1-847-938-5034; e-mail: hing.l.sham@
abbott.com
Scheme 1. ABT-378 and its three major metabolites.
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00243-8

1352
H. L. Sham et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1351–1353
Scheme 2. Retrosynthetic scheme of ABT-378 and the key inter-
mediates 3, 4, and 5 necessary for the synthesis of M-1 and M-3/M-4.
Scheme 4. Synthesis of M-1 and M-3/M-4.
Table 1. Inhibition of HIV protease and antiviral activities in MT-4
cells
a
50
Compounds
Ki (HIV protease), pMEC
(mM)
1 (ABT-378)
M-1
M-3/M-4
1.3
0.7
1.2
0.104
1.413
2.31
aIn the presence of 50% human serum.
Acknowledgements
We thank Dr. Stephen L. Gwaltney, II for his assistance
in the preparation of this manuscript.
References and Notes
Scheme 3. Synthesis of acid 5.
1. Sham, H. L.; Kempf, D. J.; Molla, A.; Marsh, K. C.;
Kumar, G. N.; Chen, C. M.; Kati, W.; Stewart, K.; Lal, R.;
Hsu, A.; Betebenner, D. A.; Korneyeva, M.; Vasavanonda,
S.; McDonald, E.; Saldivar, A.; Wideburg, N.; Chen, X.;
Niu, P.; Park, O.; Jayanti, V.; Grabowski, B.; Granneman,
G. R.; Sun, E.; Japour, A. J.; Leonard, J. M.; Plattner, J. J.;
Norbeck, D. W. Antimicrob. Agents Chemother. 1998, 42,
3218.
2. Molla, A.; Vasavanonda, S.; Kumar, G. N.; Sham, H. L.;
Johnson, M.; Grabowski, B.; Denissen, J. F.; Kohlbrenner,
W.; Plattner, J. J.; Leonard, J. M.; Norbeck, D. W.; Kempf,
D. J. Virology 1998, 250, 255.
3. Kumar, G. N.; Jayanti, V.; Lee, R. D.; Whittern, D. N.;
Uchic, J.; Thomas, S.; Johnson, P.; Grabowski, B.; Sham,
H. L.; Betebenner, D. A.; Kempf, D. J.; Denissen, J. F. Drug
Metab. Dispos. 1999, 27, 86.
4. Stoner, E. J.; Cooper, A. J.; Dickman, D. A.; Kolacz-
kowski, L.; Lallaman, J. E.; Liu, J. H.; Oliver-Shaffer, P. A.;
Patel, K. M.; Paterson, J. B.; Plata, D. J.; Riley, D. A.; Sham,
H. L.; Stengel, P. J.; Tien, J. H. J. J. Org. Process Res. Dev.
2000, 4, 264.
Coupling of the known intermediate 3 from the syn-
thesis of ABT-378,4 with acid 4 using a standard peptide
coupling procedure (EDAC, HOBt),7 followed by
hydrogenolysis provided amine 9 (85%). Coupling of
amine 9 with acid 5 gave M-1 in 82% yield (Scheme 4).
Reduction of M-1 with sodium borohydride in methyl-
ene chloride provided an inseparable epimeric mixture
(ꢀ1:1) of M-3/M-4 in 61% yield.8 The NMR, mass
spectroscopy, and HPLC retention times of the syn-
thetic materials were identical to that isolated from
metabolism studies. The biological activities of M-1, M-3/
M-4 in comparison with ABT-378 are shown in Table 1.
In summary, a concise synthesis of the major metabo-
lites of the potent second generation HIV protease
inhibitor ABT-378 (Lopinavir) was developed. It
allowed the confirmation of the structures of the meta-
bolites and the determination of the biological proper-
ties of these metabolites.

H. L. Sham et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1351–1353
1353
5. Stuk, T. L.; Haight, A. R.; Scarpetti, D.; Allen, M. S.;
Menzia, J. A.; Robbins, T. A.; Parekh, S. I.; Langridge, D. C.;
Tien, J. H. J.; Pariza, R. J.; Kerdesky, F. A. J. Org. Chem.
1994, 59, 4040.
6. Haight, A. R.; Stuk, T. L.; Allen, M. S.; Bhagavatula, L.;
Fitzgerald, M.; Hannick, S. M.; Kerdesky, F. A.; Menzia,
J. A.; Parekh, S. I.; Robbins, T. A.; Scarpetti, D.; Tien, J. H. J.
J. Org. Process Res. Dev. 1999, 3, 94.
7. Ho, G. J.; Emerson, K. M.; Mathre, D. J.; Shuman, R. F.;
Grabowski, E. J. (special issue). J. Org. Chem. 1995, 60, 3569
8. All compounds reported have mass, NMR, and analytical
data consistent with the structures assigned.