TY - JOUR
T1 - Pre-steady-state kinetic studies establish entecavir 5′-triphosphate as a substrate for HIV-1 reverse transcriptase
AU - Domaoal, Robert A.
AU - McMahon, Moira
AU - Thio, Chloe L.
AU - Bailey, Christopher M.
AU - Tirado-Rives, Julian
AU - Obikhod, Aleksander
AU - Detorio, Mervi
AU - Rapp, Kimberly L.
AU - Siliciano, Robert F.
AU - Schinazi, Raymond F.
AU - Anderson, Karen S.
PY - 2008/2/29
Y1 - 2008/2/29
N2 - The novel 2′-deoxyguanosine analog Entecavir (ETV) is a potent inhibitor of hepatitis B virus (HBV) replication and is recommended for treatment in human immunodeficiency virus type 1 (HIV-1) and HBV-co-infected patients because it had been reported that ETV is HBV-specific. Recent clinical observations, however, have suggested that ETV may indeed demonstrate anti-HIV-1 activity. To investigate this question at a molecular level, kinetic studies were used to examine the interaction of 5′-triphosphate form of ETV with wild type (WT) HIV-1 reverse transcriptase (RT) and the nucleoside reverse transcriptase inhibitor-resistant mutation M184V. Using single turnover kinetic assays, we found that HIV-1 WT RT and M184VRT could use the activated ETV triphosphate metabolite as a substrate for incorporation. The mutant displayed a slower incorporation rate, a lower binding affinity, and a lower incorporation efficiency with the 5′-triphosphate form of ETV compared with WTRT, suggesting a kinetic basis for resistance. Our results are supported by cell-based assays in primary human lymphocytes that show inhibition of WT HIV-1 replication by ETV and decreased susceptibility of the HIV-1 containing the M184V mutation. This study has important therapeutic implications as it establishes ETV as an inhibitor for HIV-1 RT and illustrates the mechanism of resistance by the M184V mutant.
AB - The novel 2′-deoxyguanosine analog Entecavir (ETV) is a potent inhibitor of hepatitis B virus (HBV) replication and is recommended for treatment in human immunodeficiency virus type 1 (HIV-1) and HBV-co-infected patients because it had been reported that ETV is HBV-specific. Recent clinical observations, however, have suggested that ETV may indeed demonstrate anti-HIV-1 activity. To investigate this question at a molecular level, kinetic studies were used to examine the interaction of 5′-triphosphate form of ETV with wild type (WT) HIV-1 reverse transcriptase (RT) and the nucleoside reverse transcriptase inhibitor-resistant mutation M184V. Using single turnover kinetic assays, we found that HIV-1 WT RT and M184VRT could use the activated ETV triphosphate metabolite as a substrate for incorporation. The mutant displayed a slower incorporation rate, a lower binding affinity, and a lower incorporation efficiency with the 5′-triphosphate form of ETV compared with WTRT, suggesting a kinetic basis for resistance. Our results are supported by cell-based assays in primary human lymphocytes that show inhibition of WT HIV-1 replication by ETV and decreased susceptibility of the HIV-1 containing the M184V mutation. This study has important therapeutic implications as it establishes ETV as an inhibitor for HIV-1 RT and illustrates the mechanism of resistance by the M184V mutant.
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U2 - 10.1074/jbc.M707834200
DO - 10.1074/jbc.M707834200
M3 - Article
C2 - 17962184
AN - SCOPUS:41949114325
SN - 0021-9258
VL - 283
SP - 5452
EP - 5459
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 9
ER -