No selection of nucleoside reverse transcriptase inhibitor resistance associated mutations by acyclovir suppressive therapy in herpes simplex virus-2/HIV-1 dually infected persons

Abstract

Genital herpes simplex virus type 2 (HSV-2) shedding has been associated with increased concentrations of HIV-1 in the genital tract and plasma [1,2]. Recent data from randomized controlled trials suggest that daily HSV-2 suppressive therapy with acyclovir (ACV) or valacyclovir over periods of up to 3 months may reduce plasma and genital HIV RNA load by 0.3–0.5 log10 copies/ml among HIV/HSV-2 coinfected individuals not receiving antiretroviral therapy (ART) [3–7]. However, a recent trial of suppressive therapy [ACV 400 mg twice daily (b.i.d.)] delivered to the HIV/HSV-2 seropositive partner in HIV discordant couples found no decrease in the rate of HIV transmission to the HIV negative partner [8]. ACV is an acyclic guanosine analog that inhibits HSV DNA polymerase after conversion to its triphosphate form. As the first phosphorylation is synthesized selectively by HSV thymidine kinase [9], ACV is mainly active in HSV-infected cells and was therefore thought to have no or poor antiretroviral activity. The postulated mechanism of impact on HIV is that herpes suppressive therapy is able to prevent HIV replication by disrupting direct HSV-2/HIV-1 interaction and, indirectly, by reducing the systemic and local immune activation induced by HSV-2 [10]. However, recent in-vitro experiments have suggested that ACV may directly inhibit HIV replication and induce the selection of mutations in the HIV reverse transcriptase known to confer resistance to nucleoside reverse transcriptase inhibitors (NRTIs) [11,12]. ACV has been shown to select the mutation V75I, associated with the reverse transcriptase multidrug resistance complex, and also, to a lesser extent, T69N and M184I, which typically precedes the emergence of the M184V mutation that confers a high level of resistance to lamivudine/emtricitabine. These results suggest that the use of ACV may select HIV variants harboring mutations associated with resistance to NRTI. If confirmed in vivo, this may impact the choice and efficacy of antiretroviral regimens among HIV patients who have previously been treated with ACV, and would suggest that the use of ACV in ART-naïve HIV-positive patients should be carefully considered. To determine the impact of ACV suppressive therapy on the selection of mutations in the HIV reverse transcriptase gene in vivo, we analyzed plasma samples from dually HIV/HSV-2-infected women receiving ACV suppressive therapy who were enrolled in a randomized placebo-controlled trial of ACV 400 mg b.i.d. for HSV suppression in Tanzania [13,14]. The detection of mutations was performed by HIV reverse transcriptase gene sequence analysis on plasma samples selected from women with a reported high-ACV adherence (defined as taking >90% tablets by tablet counts) and HIV RNA load more than 3 log10 copies/ml after 12 months of ACV suppressive therapy, and who were not on ART. Among 21 selected samples, none of the mutations V75I, T69N and M184I were detected. No other mutations known to confer resistance to NRTI were detected in the reverse transcriptase gene. In four samples, mutations at the codon 69 (69A n = 2, 69P n = 1, T69S n = 1) were detected. T69A and T69P are not known to be associated with NRTI resistance, but T69S in combination with other mutations may be associated with tenofovir resistance. This mutation may have been selected owing to ACV exposure or owing to natural polymorphism. These results suggest that b.i.d. HSV-2 suppressive therapy with a standard dose of 400 mg of ACV for 12 months did not induce the selection of NRTI resistance mutations in HIV-infected women not receiving ART. Because the threshold of resistant variant detection with sequence analysis is above 20%, we cannot completely rule out the presence of minor populations containing NRTI resistance mutations. However, we selected samples from women with high-ACV adherence to guarantee the highest selective pressure of ACV, and restricted the laboratory analyses to those who had been on ACV therapy for 12 months in order to be able to detect late emergence of ACV-associated NRTI resistance mutations. Thus, the selection of mutations in the reverse transcriptase gene by ACV exposure seems unlikely in our population. Considering the bioavailability of oral ACV, plasma concentrations after 400 mg b.i.d. should not exceed 1 μmol/l, five-fold lower than the half maximal inhibitory concentration for HIV [12] and such concentrations might not be sufficient to achieve antiretroviral activity. Our results do not confirm the in-vitro data obtained studies using high-dose ACV [12], but corroborate results recently presented on patients receiving ACV or valacyclovir for periods of 8–24 months [15]. These reassuring results for the use of 400 mg b.i.d. ACV in HSV-2/HIV coinfected patients deserve further confirmation especially if higher dosage of ACV is used in HIV-infected individuals.