HIV-1 Integrase

Division Of Basic Sciences - Nci

Investigators

Linked publications & trials

Abstract

INSTI development: The development of new, broadly effective anti-HIV drugs that have little or no toxicity is a high-priority NIH goal for HIV/AIDS research. Because integration is an essential step in the virus life cycle, IN is an important target for antiretroviral drugs. The HIV-1 integration reaction proceeds in two steps, both of which are carried out by IN. In the first step (3' processing, or 3'P), IN removes two nucleotides from the 3' ends of the linear viral DNA. In the second step, IN inserts the trimmed 3' ends of the viral DNA into host DNA; this reaction is called strand transfer, or ST. IN has only one active site which carries out both the 3'P and ST reactions. The four approved anti-IN drugs (raltegravir, elvitegratir, dolutegravir, and bictegravir) all target the ST reaction, and are, for this reason, called integrase strand transfer inhibitors, or INSTIs. Despite their relatively recent development, INSTIs are potent drugs with few side effects that are becoming increasingly important in antiretroviral therapies (ART). Cabotegravir has been formulated with rilpivirine and this combination has shown promise in long-term treatment strategies. However, as is the case with all anti-HIV drugs, INSTIs select for resistant strains of HIV. We are developing new INSTIs that are broadly effective against the known drug-resistant mutants. We have made excellent progress in developing IN inhibitors that have low nanomolar potency against the WT virus in a one-round inhibition assay, retain potency against a broad panel of resistant mutants, and show little or no toxicity in cultured cells. Our collaborator, Dmitry Lyumkis, has solved high resolution structures of HIV-1 IN (both WT and drug resistant) in complexes with both FDA approved INSTIs and with the most promising of the compounds developed and synthesized in the laboratory of another collaborator, Dr. Terry Burke. Although we continue to make and test new INSTIs, we have made sufficient progress that, with help from the NCI, pharmacokinetic testing is being done in macaques on the best of our compounds. Integration site analysis: There are millions of retroviral integration sites in the host cell genome, but for many retroviruses, including HIV, integration is far from random. We have been studying the distribution of HIV proviruses (integration sites) in both cultured cells and patients. HIV integration is known to favor the bodies of highly expressed genes. In work that was done in cultured cells, we were part of a collaboration that showed that the host protein HRP2 could replace LEDGF in directing HIV-1 integration to the bodies of highly expressed genes. We also showed that the host factor CPSF6 plays a key role in guiding the preintegration complex to regions of the genome that are gene rich and contain numerous highly expressed genes. We and others have used integration site analysis to show that there is extensive clonal expansion of HIV-infected cells in patients on ART. We also showed, in HIV-infected individuals on long-term ART, that more than 40% of the infected T cells are in clones and that, in some cases, proviruses integrated into six oncogenes can contribute to clonal expansion of infected T cells. Some of these highly expanded clones carry infectious proviruses and a small fraction of the cells in the clones can release infectious virions into the blood. Thus, expanded clones of HIV infected T cells that carry intact infectious proviruses represent a large fraction of the infected cells that comprise the reservoir that has made it impossible to cure HIV infections with the available anti-viral therapies. The reservoir, which arises early in infection, persists on ART and can rekindle an active infection if ART is discontinued. Because the number of possible sites of HIV DNA integration is very large, each infected cell, and all of its descendants, can be identified by the site where the provirus is integrated. Surprisingly, we found that the distribution of HIV proviruses, even after years of complete virological suppression, in which the majority of the infected surviving cells are removed from the originally infected cells by many divisions, closely resembles the distribution of the proviruses at the time the cells were initially infected. In our experiments, the initial distribution was represented by the distribution of the proviruses in PBMC acutely infected in vitro. In addition to a modest modification to the initial distribution by a positive selection for proviruses integrated in six known oncogenes, the distribution of proviruses is also modified by selection against cells with proviruses integrated in highly expressed genes. Thus, although proviruses in oncogenes can cause the extensive in a small fraction of the clones of infected cells, our data imply that most important factors causing clonal expansion of HIV infected T cells are antigen and/or cytokine stimulation. Our data also strongly support the idea that the integration site of a provirus has little if any effect on the expression of the integrated provirus. Retroviruses can cause cancers in animals by integrating in or near oncogenes. Although HIV-1 infection increases the risk of cancer, most of the risk is associated with HIV induced immunodeficiency which allows oncogenic viruses to proliferate (EBV, KSHV, HPV). As was described in the previous paragraph, HIV-1 proviruses integrated in six oncogenes can cause clonal expansion of infected T cells in vivo; however, the infected cells are not transformed. Until very recently, it was generally believed that HIV-1 does not cause cancer directly. However, we recently showed that HIV-1 proviruses integrated in the first introns of STAT3 and LCK can play an important role in the development of T cell lymphomas. The development of these cancers appears to be a multi-step process involving additional non-viral mutations, which could help explain why T cell lymphomas are rare in people living with HIV-1 infections.