Mutagenesis of Single/Combined NRTI Drugs in Human Cells

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DESCRIPTION (provided by applicant): Highly active antiretroviral therapy (HAART) has been effective at reducing the transmission of HIV from mother to infant, but the combinations of drugs used for HAART in pregnant women poses a risk for neoplasia in the infants exposed in utero to nucleoside analogue reverse transcriptase inhibitors (NRTIs). The purpose of this proposal is to determine the extent of nuclear DNA damage induced by exposure to specific NRTI's, alone or in combination, in human lymphoblastoid cells (AZH-1), and to determine the variability in the mutation susceptibility phenotype in human cord blood lymphocytes. Experiments will be performed to measure (by specific RIAs) DNA incorporation of zidovudine (AZT), lamivudine (3TC), and/or stavudine (d4T), and to measure and characterize the mutagenic response at the HPRT and APRT loci of AZH-1cells and the HPRT locus of cord blood cells (using cell cloning assays) exposed in culture to clinically important antiretroviral drugs, as single agents or combinations. This work is an extension of studies previously conducted by our group to investigate the in utero mutagenicity of perinatal exposure of infants to AZT. It was demonstrated that a direct correlation exists between the level of the AZT incorporated into DNA and the levels of mutations induced at specific loci in human cells. Second, co-exposure to AZT and a second NRTI (ddI) potentiates AZT-DNA incorporation and mutation induction in vitro. Third, NRTI therapy using AZT alone or with 3TC induces significant genotoxic and mutagenic effects in infants exposed in utero, and the increases in mutagenic responses persist for at least one year after birth. Data indicate that this increase is driven by a subset of the children, suggesting that individual susceptibility factors may influence the levels of DNA damage and mutation that results from in utero prophylaxis. AZT is also a transplacental carcinogen in exposed mice and rats. The proposed work will identify the drugs/drug combinations with the lowest mutagenic potential, will initiate studies to define the basis for the increased susceptibility to mutagenesis in a subset of AZT-3TC exposed infants, and will form the foundation for quantitative risk assessments of perinatal prophylaxis using individual NRTIs or combinations of NRTIs.