Impact of Integrase Inhibition Compared With Nonnucleoside Inhibition on HIV Reservoirs in Lymphoid Tissues.

BACKGROUND: HIV is produced in lymphoid tissues (LT) and stored on the follicular dendritic cell network in LT. When antiretroviral therapy is started, plasma viremia decays in 2 phases; the first within days of starting therapy and the second over weeks. Raltegravir (RAL), an integrase inhibitor, has been associated with only a single rapid phase of decay, and we speculated this may be due to higher intracellular concentration (IC) of RAL in LT. We have previously measured suboptimal ICs of antiretroviral therapy agents in LT, which were associated with slower decay of both vRNA+ cells and the follicular dendritic cell network pool. SETTING: Outpatient clinic at the Joint Clinical Research Center in Kampala, Uganda. METHODS: We compared the rate of decay in LT in people starting RAL with those starting efavirenz (EFV). RESULTS: There was no difference in the rate of virus decay in LT. The ratio of the ICs of RAL and EFV in lymph node to the concentration of drug that inhibits 95% of virus in blood was 1 log lower in lymph node for EFV and >3 logs lower for RAL. CONCLUSION: These data further highlight the challenges of drug delivery to LT in HIV infection and demonstrate that RAL is not superior to EFV as judged by direct measurements of the source of virus in LT.

Transplantation of CCR5∆32 Homozygous Umbilical Cord Blood in a Child With Acute Lymphoblastic Leukemia and Perinatally Acquired HIV Infection.

BACKGROUND: Allogeneic hematopoietic cell transplantation (allo-HCT) in a CCR5∆32 homozygous donor resulted in HIV cure. Understanding how allo-HCT impacts the HIV reservoir will inform cure strategies. METHODS: A 12-year-old with perinatally acquired, CCR5-tropic HIV and acute lymphoblastic leukemia underwent myeloablative conditioning and umbilical cord blood (UCB) transplantation from a CCR5∆32 homozygous donor. Peripheral blood mononuclear cells (PBMCs) and the rectum were sampled pre- and post-transplant. The brain, lung, lymph node (LN), stomach, duodenum, ileum, and colon were sampled 73 days after transplantation (day +73), when the patient died from graft-vs-host disease. Droplet digital polymerase chain reaction (ddPCR) and in situ hybridization (ISH) were used detect the HIV reservoir in tissues. CCR5 and CD3 expression in the LN was assessed using immunohistochemistry (IHC). RESULTS: HIV DNA (vDNA) was detected in PBMCs by ddPCR pretransplant but not post-transplant. vDNA was detected by ISH in the rectum at days -8 and +22, and in the LN, colon, lung, and brain day +73. vDNA was also detected in the lung by ddPCR. IHC revealed CCR5+CD3+ cells in the LN postmortem. CONCLUSIONS: HIV was detected in multiple tissues 73 days after CCR5∆32 homozygous UCB allo-HCT despite myeloablative conditioning and complete donor marrow engraftment. These results highlight the importance of analyzing tissue during HIV cure interventions and inform the choice of assay used to detect HIV in tissue reservoirs.

Simultaneous cell-to-cell transmission of human immunodeficiency virus to multiple targets through polysynapses.

Human immunodeficiency virus type 1 (HIV-1) efficiently propagates through cell-to-cell contacts, which include virological synapses (VS), filopodia, and nanotubes. Here, we quantified and characterized further these diverse modes of contact in lymphocytes. We report that viral transmission mainly occurs across VS and through "polysynapses," a rosette-like structure formed between one infected cell and multiple adjacent recipients. Polysynapses are characterized by simultaneous HIV clustering and transfer at multiple membrane regions. HIV Gag proteins often adopt a ring-like supramolecular organization at sites of intercellular contacts and colocalize with CD63 tetraspanin and raft components GM1, Thy-1, and CD59. In donor cells engaged in polysynapses, there is no preferential accumulation of Gag proteins at contact sites facing the microtubule organizing center. The LFA-1 adhesion molecule, known to facilitate viral replication, enhances formation of polysynapses. Altogether, our results reveal an underestimated mode of viral transfer through polysynapses. In HIV-infected individuals, these structures, by promoting concomitant infection of multiple targets in the vicinity of infected cells, may facilitate exponential viral growth and escape from immune responses.

Determination of simian immunodeficiency virus production by infected activated and resting cells.

OBJECTIVE: To determine the relative amount of virus produced by activated and resting CD4+ T cells. DESIGN: The total quantity of virus produced by an activated cell relative to a resting cell in vivo was estimated from 'snap-shots' of virus production by infected cells at one time point. METHODS: Bayesian statistical methods were used to determine a credible interval for the desired ratio. RESULTS: The posterior mean of the ratio of virus produced by a typical activated cell to a typical resting cell is 0.82 to 4.28, depending on the half-lives of the resting infected cells. Simian immunodeficiency virus-infected resting cells could accordingly be responsible for 70 to 93% of peak virus production in the acute stage of infection. CONCLUSIONS: Whereas in 'snap-shots' the infected resting cells apparently produce much less virus than infected activated CD4+ T cells, the coincidence of peak SIV production with predominant infection of resting cells along with longer half-lives for productively infected resting cells point to a major contribution to virus production in early infection.

Transendothelial migration of leukocytes carrying infectious HIV-1: an indicator of adverse prognosis.

OBJECTIVE: To ascertain the likelihood that perivascular leukocyte infiltrates are sites for replication and dissemination of HIV-1. DESIGN AND METHODS: We measured the ability of HIV patients' peripheral blood mononuclear leukocytes to migrate through confluent endothelial monolayers in vitro and infect phytohemagglutinin-stimulated allogeneic lymphoblasts. We also measured the ability of migratory leukocytes to transmit virus to uninfected leukocytes that have localized outside an endothelial barrier, and the subsequent ability of these newly infected cells to reverse-migrate back across the endothelial barrier - a process that might facilitate reentry of infected cells into the circulation and dissemination of the virus to distant sites. RESULTS: Leukocytes from 27 out of 63 unselected patients spontaneously carried infectious virus across endothelial barriers. On follow-up, these 27 patients were frequently observed to develop uncontrolled viremia, despite treatment, and be hospitalized for secondary infections. Migratory leukocytes transmitted HIV to both T lymphocytes and non-T cells that had previously crossed the endothelial barrier. Either cell type could subsequently reverse-migrate carrying virus back across this barrier. CONCLUSIONS: Reverse-migration of HIV-1 infected leukocytes out of perivascular reservoirs may provide a way to disseminate HIV-1 and expand the body burden of virus in some patients receiving highly active antiretroviral therapy.