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Antimicrobial Agents and Chemotherapy, July 2000, p. 1783-1788, Vol. 44, No. 7
Department of Infectious Diseases and
Microbiology, Graduate School of Public Health, University of
Pittsburgh,1 Department of Medicine,
University of Pittsburgh School of Medicine,4
and Veterans Affairs Medical Center,5
Pittsburgh, Pennsylvania; Center for Drug Discovery and
Department of Pharmaceutical and Biochemical Sciences, University
of Georgia, Athens, Georgia2; and
Department of Pediatrics, Emory University School of
Medicine, and the Georgia Veterans Affairs Research Center on AIDS
and HIV Infection, Veterans Affair Medical Center, Decatur,
Georgia3
Received 28 October 1999/Returned for modification 22 February
2000/Accepted 20 March 2000
Human immunodeficiency virus type 1 (HIV-1) isolates resistant to
(
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
In Vitro Selection of Mutations in the Human
Immunodeficiency Virus Type 1 Reverse Transcriptase That Decrease
Susceptibility to (
)-
-D-Dioxolane-Guanosine and
Suppress Resistance to 3'-Azido-3'-Deoxythymidine
)-
-D-dioxolane-guanosine (DXG), a potent and
selective nucleoside analog HIV-1 reverse transcriptase (RT) inhibitor, were selected by serial passage of HIV-1LAI in increasing
drug concentrations (maximum concentration, 30 µM). Two independent selection experiments were performed. Viral isolates for which the DXG
median effective concentrations (EC50s) increased 7.3- and
12.2-fold were isolated after 13 and 14 passages, respectively. Cloning
and DNA sequencing of the RT region from the first resistant isolate
identified a K65R mutation (AAA to AGA) in 10 of 10 clones. The role of
this mutation in DXG resistance was confirmed by site-specific mutagenesis of HIV-1LAI. The K65R mutation also conferred
greater than threefold cross-resistance to 2',3'-dideoxycytidine,
2',3'-dideoxyinosine, 2',3'-dideoxy-3'-thiacytidine,
9-(2-phosphonylmethoxyethyl)adenine, 2-amino-6-chloropurine dioxolane,
dioxolanyl-5-fluorocytosine, and diaminopurine dioxolane but had only
marginal effects on 3'-azido-3'-deoxthymidine (AZT) susceptibility.
However, when introduced into a genetic background for AZT resistance
(D67N, K70R, T215Y, T219Q), the K65R mutation reversed the AZT
resistance. DNA sequencing of RT clones derived from the second
resistant isolate identified the L74V mutation, previously reported to
cause ddI resistance. The L74V mutation also decreased the AZT
resistance when the mutation was introduced into a genetic background
for AZT resistance (D67N, K70R, T215Y, T219Q) but to a lesser degree
than the K65R mutation did. These findings indicate that DXG and
certain 2',3'-dideoxy compounds (e.g., ddI) can select for the same
resistance mutations and thus may not be optimal for use in
combination. However, the combination of AZT with DXG or its orally
bioavailable prodrug (
)-
-D-2,6-diaminopurine-dioxolane
should be explored because of the suppressive effects of the K65R and
L74V mutations on AZT resistance.
*
Corresponding author. Mailing address: 603 Parran Hall,
Graduate of School of Public Health, University of Pittsburgh, 130 DeSoto St., Pittsburgh, PA 15261. Phone: (412) 624-8512. Fax: (412)
383-7982. E-mail: mellors{at}msx.dept-med.pitt.edu.
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