Our long-range goal is to understand the intracellular pharmacology of antiretroviral drugs used against HIV. The objective in this application is to investigate the in vivo effect of nbavirin on the phosphorylation process of ZDV in HIV-HCV co-infected patients. The central hypothesis to be tested is that REBETRON (ribavirin + interferon alpha) will antagonize in vivo with ZDV, diminishing the formation of ZDV intracellular metabolites (ZDV-MP and ZDV-TP), by increasing thymidine triphosphate (dTTP) intracellular concentrations which inhibits thymidine kinase 1. These events may affect HIV clinical outcomes in HIV-HCV co-infected patients, since the dTTP/ZDV-TP ratio will be higher after REBETRON therapy thereby diminishing ZDV effectiveness. The rationale behind the proposed application relies on the in vitro studies showing an 80 percent reduction of ZDV-MP and ZDV-TP intracellular concentrations in different cell lines with the concurrent treatment of ribavirin. If these observations are maintained in vivo, we might be diminishing the concentrations of the active component of ZDV (ZDV-TP) to unknown levels and we will be increasing dTTP (endogenous competitor for HIV reverse transcriptase) intracellular concentrations. Thus, in the dTTP/ZDV-TP ratio increases dramatically, we might be changing from a three-drug therapy in HAART (ZDV + two other drugs including a protease inhibitor) to a two-drug therapy (two other drugs including a protease inhibitor) increasing the possibilities for the development of resistant mutations. To accomplish the objectives of this application, we will pursue two specific aims: (1) To compare the steady state area under the concentration-time-curve between 0 and 12 hours (AUC0.12) of ZDV-MP, ZDV-TP, and dTTP in peripheral blood mononuclear cells (PBMCs) and nelfinavir in plasma from HIV-HCV co-infected patients before and after REBETRON therapy with a control group of HIV infected HCV-negative patients, and (2) To describe the effect of REBETRON in HCV-RNA viral titers, liver histology, ALT, CD4+, and HIV-RNA viral titers in HIV-HCV coinfected patients after 24 weeks of treatment. After the completion of the research project, we expect to observe at least a 20 percent reduction (conservative estimate) of ZDV-MP and ZDV-TP. Furthermore, we will determine the in vivo percent increase of dTTP after REBETRON therapy. Thus, we will be in a position to determine the effect of the dTTP/ZDV-TP ratio in the progression of HIV in this patient population. Furthermore, since there are no established therapeutic ranges for intracellular concentrations of ZDV-TP, we will determine if ZDV-TP concentrations are above the estimated in vitro 50 percent inhibitory values for HIV and if the exposure (AUC about2) is sustained after REBETRON therapy. This information would provide strong support for similar clinical efficacy for both regimens (before and after REBETRON). Moreover, these data will provide valuable insight into the inter- and intra-subject variability of intracellular ZDV-MP, ZDV-TP, and dTTP. Finally, we will describe the effect of REBETRON in the progression of HCV disease (liver histology, HCV-RNA viral titers and ALT level) in HCV-HIV co-infected patients.
| Meléndez, Margarita; Rosario, Osvaldo; Zayas, Beatriz et al. (2009) HPLC-MS/MS method for the intracellular determination of ribavirin monophosphate and ribavirin triphosphate in CEM ss cells. J Pharm Biomed Anal 49:1233-40 |
