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Specific HIV Integration Sites Linked to Clonal Expansion and Persistence of Cells
Stephen H. Hughes1
1 HIV Drug Resistance Program, National Cancer Institute, Frederick, MD, United States.
The persistence of HIV-infected cells in individuals on suppressive combination antiretroviral therapy (cART) presents a major barrier for curing HIV infections.
Methods: HIV integrates its DNA into many sites in the host genome; thus integration sites can be used as markers to identify clonally expanded cells. We identified the integration sites in PBMCs and CD4+ T cells from patients and used the data to analyze the clonal expansion of infected cells. We have started to determine the sequences of viral DNAs in clonally expanded cells; these sequences are being used to study the expression of the proviruses in the expanded cells.
Results: We have identified >2500 integration sites in PBMCs and CD4+ cells of infected individuals on cART. About 40% of the integrations were in clonally expanded cells. In one patient, more than 50% of the infected cells were from a single clone; some of the expanded clones persisted for more than 10 years. There were multiple independent integrations in the same orientation as the gene in two introns of the MKL2 and BACH2 genes; many of these integrations were in clonally expanded cells. Both BACH2 and MKL2 are involved in regulating the growth of cells. DNA rearrangements involving these genes are present in human tumors. There was no evidence forintegration in one orientation, or in specific introns, in either of these genes in large libraries prepared by infecting stimulated or unstimulated PBMCs, CD34+ cells, or HeLa cells in culture. There were, in patients, multiple independent integrations in a number of other growth related genes, many of which were associated with clonal expansion of the infected cells. These data show that HIV integration at certain sites can play a critical role in the expansion and persistence of HIV infected cells. In one case, we showed that the provirus in an expanded clone was responsible for producing the majority of the virus that was present in the blood of a patient. This shows that, in this patient, immune surveillance was not sufficient to prevent clonally expanded cells from producing virions.
Conclusions: Our findings have important implications for the development and maintenance of the viral reservoir, for designing and implementing strategies to eliminate persistent HIV infection, for the use of lentiviral vectors for gene therapy in human patients, and, possibly, for the origin of some HIV-related malignancies.