Effective antiviral chemotherapy for patients with chronic HIV-1 infection will likely require a better understanding of the mechanisms involved in breakthrough of viral replication seen during drug therapy. Breakthrough of HIV-1 replication is defined as the emergence of detectable virus in the presence of drug. Viral breakthrough occurs in the face of seemingly effective treatment both in vitro and in vivo. Our hypothesis is that breakthrough of HIV-1 replication may result from: emergence of drug-resistant virus variants, loss of cellular response to antiviral agent(s), significant virus load overwhelming available drug, and/or inability of drug to affect viral reservoirs. The goal of this research effort is to investigate the mechanism(s) of breakthrough of HIV-1 replication both in vitro and in vivo. The mechanisms of breakthrough of HIV-1 replication in vitro will be explored by determining if drug-resistant virus emerges, if cells become refractory to zidovudine (AZT) therapy, and if significant virus load overwhelms available drug after multiple rounds of replication. This research will evaluate peripheral blood mononuclear cells derived from drug-treated HIV-1-infected patients for the presence of drug-resistant virus using CD8+ lymphocyte depletion and a mutation-specific polymerase chain reaction (PCR) assay. The cellular anabolism of AZT after prolonged monotherapy in vivo will also be investigated. The phenotype and genotype of drug-resistant virus that emerge during single agent and combined therapy in vivo will be identified. Alternative drug strategies will be evaluated against drug-sensitive and drug-resistant viruses in vitro, including 1- and 2-drug regimens that act at similar or different sites in the HIV-1 replicative cycle. To investigate the mechanism of inhibition of AZT-resistant virus by 2-drug regimens in vitro when AZT is included in the treatment regimen, the AZT-resistant viral isolate will be characterized molecularly to determine wich proportion of AZT-sensitive and AZT-resistant HIV-1 exists in the virus population. Overall, this research effort should identify strategies to achieve optimal virus suppression in HIV-1 infected individuals.