An in vitro evaluation of drugs used in the Kenyan ART program

Abstract

The majority of anti-HIV drug susceptibility tests have been performed on subtype B HIV-1 strains, since these are the most prevalent in countries designing, testing, and manufacturing the current anti-HIV agents. The increasing global spread of HIV subtype highlights the need to determine the activity of anti-HIV drugs against subtypes of HIV other than subtype B. Furthermore an increasing number of individuals infected with many of the non subtype B virus strains now receive antiretroviral therapy because of rollout programs in developing countries as well as increasing migration to the developed world. The phenotypic susceptibility of two laboratory strains HIV-1JFRL and HIV-1IIIB (representing subtype B) and two clinical isolates HIV-104RTA and HIV-1025RTA (representing subtypes A and D respectively) was determined. The in vitro drug susceptibility testing of the isolates was carried out in C8166 cell line and in peripheral blood mononuclear cells (PBMCs). The study revealed that the drugs used in the Kenyan national ART program inhibited HIV-1 replication in-vitro as their inhibitory concentrations (IC50) compared well with the standard Inhibitory concentration values. The results also suggest a biochemical similarity of the reverse transcriptase (RT) and protease enzymes from these subtypes despite the divergence at the genetic level. The findings suggest that similar clinical benefits of antiviral therapy obtain in persons infected with other subtypes of HIV-1other than subtype B and that the generic drugs used in the national ART program in Kenya are as efficacious as branded drugs in inhibiting HIV replication in vitro despite the limited number of the viruses studied.

Keywords: HIV; anti retroviral therapy; susceptibility.

Figure 1

Drug susceptibility curves for AZT, ABC and EFV. LPV/r and NVP/3TC/d4T against (A) HIVI_IIB, (B) HIV 04RTA, © HIV_025RTA and HIV-1_JRFL (D). Each dilution of the drug was tested in duplicate in C8166 cell line for the three X4 viruses and in PBMCs for the JRFL (R5). Results represent the means of two independent experiments; the figure shows percent inhibition of P24 antigen production in culture supernatants. The intersection of horizontal broken lines denotes the 50% inhibitory concentration [IC 50] of each drug

Figure 2

Inhibition of cytopathic effect [CPE] by the drugs. C8166 cells were plated in 96-well plates at a concentration of 1x10⁵/ml in RPMI1640 culture medium and infected with HIV-1025RTA at a MOI of 0.01 and various concentrations of ARVs ranging from 0 to 5µg/ml added. The micro plates were incubated for 3 to 5 days and cells observed daily for CPE under inverted microscope. A and C show inhibition of CPE by the ARVs at a concentration of 5µg/ml and mock infected control respectively. At 0.04µg/ml drugs concentration B a typical CPE of giant multinucleated cells is seen. Magnification x 400