Michele Di Mascio

Towards an HIV cure: a global scientific strategy

Given the limitations of antiretroviral therapy and recent advances in our understanding of HIV persistence during effective treatment, there is a growing recognition that a cure for HIV infection is both needed and feasible. The International AIDS Society convened a group of international experts to develop a scientific strategy for research towards an HIV cure. Several priorities for basic, translational and clinical research were identified. This Opinion article summarizes the groups recommended key goals for the international community.

A Novel Antiviral Intervention Results in More Accurate Assessment of Human Immunodeficiency Virus Type 1 Replication Dynamics and T-Cell Decay In Vivo

ABSTRACT Mathematical models provide an understanding of in vivo replication kinetics of human immunodeficiency virus type 1 (HIV-1). With a novel intervention designed for increased potency, we have more accurately deduced the half-lives of virus-producing CD4+ T cells, 0.7 day, and the generation time of HIV-1 in vivo, approximately 2 days, confirming the dynamic nature of HIV-1 replication.

Induction of prolonged survival of CD4+ T lymphocytes by intermittent IL-2 therapy in HIV-infected patients

HIV infection leads to decreases in the number of CD4 T lymphocytes and an increased risk for opportunistic infections and neoplasms. The administration of intermittent cycles of IL-2 to HIV-infected patients can lead to profound increases (often greater than 100%) in CD4 cell number and percentage. Using in vivo labeling with 2H-glucose and BrdU, we have been able to demonstrate that, although therapy with IL-2 leads to high levels of proliferation of CD4 as well as CD8 lymphocytes, it is a remarkable preferential increase in survival of CD4 cells (with half-lives that can exceed 3 years) that is critical to the sustained expansion of these cells. This increased survival was time-dependent: the median half-life, as determined by semiempirical modeling, of labeled CD4 cells in 6 patients increased from 1.7 weeks following an early IL-2 cycle to 28.7 weeks following a later cycle, while CD8 cells showed no change in the median half-life. Examination of lymphocyte subsets demonstrated that phenotypically naive (CD27+CD45RO-) as well as central memory (CD27+CD45RO+) CD4 cells were preferentially expanded, suggesting that IL-2 can help maintain cells important for host defense against new antigens as well as for long-term memory to opportunistic pathogens.

HIV infection-associated immune activation occurs by two distinct pathways that differentially affect CD4 and CD8 T cells

HIV infection is characterized by a brisk immune activation that plays an important role in the CD4 depletion and immune dysfunction of patients with AIDS. The mechanism underlying this activation is poorly understood. In the current study, we tested the hypothesis that this activation is the net product of two distinct pathways: the inflammatory response to HIV infection and the homeostatic response to CD4 T cell depletion. Using ex vivo BrdU incorporation of PBMCs from 284 patients with different stages of HIV infection, we found that CD4 proliferation was better predicted by the combination of CD4 depletion and HIV viral load (R2 = 0.375, P < 0.001) than by either parameter alone (CD4 T cell counts, R2 = 0.202, P < 0.001; HIV viremia, R2 = 0.302, P < 0.001). Interestingly, CD8 T cell proliferation could be predicted by HIV RNA levels alone (R2 = 0.334, P < 0.001) and this predictive value increased only slightly (R2 = 0.346, P < 0.001) when CD4 T cell depletion was taken into account. Consistent with the hypothesis that CD4 T cell proliferation is driven by IL-7 as a homeostatic response to CD4 T cell depletion, levels of phosphorylated STAT-5 were found to be elevated in naive subsets of CD4 and CD8 T cells from patients with HIV infection and in the central memory subset of CD4 T cells. Taken together these data demonstrate that at least two different pathways lead to immune activation of T cells in patients with HIV infection and these pathways differentially influence CD4 and CD8 T cell subsets.

Elite Suppressors Harbor Low Levels of Integrated HIV DNA and High Levels of 2-LTR Circular HIV DNA Compared to HIV+ Patients On and Off HAART

Elite suppressors (ES) are a rare population of HIV-infected individuals that are capable of naturally controlling the infection without the use of highly active anti-retroviral therapy (HAART). Patients on HAART often achieve viral control to similar (undetectable) levels. Accurate and sensitive methods to measure viral burden are needed to elucidate important differences between these two patient populations in order to better understand their mechanisms of control. Viral burden quantification in ES patients has been limited to measurements of total DNA in PBMC, and estimates of Infectious Units per Million cells (IUPM). There appears to be no significant difference in the level of total HIV DNA between cells from ES patients and patients on HAART. However, recovering infectious virus from ES patient samples is much more difficult, suggesting their reservoir size should be much smaller than that in patients on HAART. Here we find that there is a significant difference in the level of integrated HIV DNA in ES patients compared to patients on HAART, providing an explanation for the previous results. When comparing the level of total to integrated HIV DNA in these samples we find ES patients have large excesses of unintegrated HIV DNA. To determine the composition of unintegrated HIV DNA in these samples, we measured circular 2-LTR HIV DNA forms and found ES patients frequently have high levels of 2-LTR circles in PBMC. We further show that these high levels of 2-LTR circles are not the result of inefficient integration in ES cells, since HIV integrates with similar efficiency in ES and normal donor cells. Our findings suggest that measuring integration provides a better surrogate of viral burden than total HIV DNA in ES patients. Moreover, they add significantly to our understanding of the mechanisms that allow viral control and reservoir maintenance in this unique patient population.

Determination of virus burst size in vivo using a single-cycle SIV in rhesus macaques

A single-cycle simian immunodeficiency virus (scSIV) that undergoes only one round of infection and replication was constructed to calculate the total number of virons produced by an SIV-infected cell in vivo. Four Mamu-A*01 rhesus macaques were inoculated on two occasions 11 weeks apart with the scSIV by ex vivo infection and i.v. reinfusion of autologous cells. After each inoculation, plasma viral loads peaked between 1 and 2.5 days and then declined exponentially in one or two phases to below detection limits within 2 weeks. Although higher levels of SIV-specific cytotoxic T lymphocytes and modest increases in antibody responses were observed for each animal after the second inoculation, decay rates of the infected cells were only minimally affected. Analyzing the viral load data with a mathematical model, the in vivo viral burst size averaged 4.0 × 104 and 5.5 × 104 virions per cell for the first and second inoculations, respectively, with no significant difference between the two inoculations. This estimate, in conjunction with our prior understanding of other quantitative viral and cellular parameters during SIV and HIV infection, provides critical insights into the dynamic process of viral production and its interplay with the infected host in vivo.

Determining the Relative Efficacy of Highly Active Antiretroviral Therapy

Despite the clinical benefits of combination antiviral therapy, whether maximal antiviral potency has been achieved with current drug combinations remains unclear. We studied the first phase of decay of human immunodeficiency virus type 1 (HIV-1) RNA in plasma, one early indicator of antiviral activity, after the administration of a novel combination of lopinavir/ritonavir, efavirenz, tenofovir disoproxil fumarate, and lamivudine and compared it with that observed in matched cohorts treated with alternative combination regimens. On the basis of these comparisons, we conclude that the relative potency of highly active antiretroviral therapy may be augmented by as much as 25%-30%. However, it is important to emphasize that further study is warranted to explore whether these early measurements of relative efficacy provide long-term virologic and clinical benefits. Nevertheless, we believe that optimal treatment regimens for HIV-1 have yet to be identified and that continued research to achieve this goal is warranted.

Viral Blip Dynamics during Highly Active Antiretroviral Therapy

ABSTRACT Although intermittent episodes of low-level viremia are often observed in well-suppressed highly active antiretroviral therapy (HAART)-treated patients, the timing and amplitude of viral blips have never been examined in detail. We analyze here the dynamics of viral blips, i.e., plasma VL measurements of >50 copies/ml, in 123 HAART-treated patients monitored for a mean of 2.6 years (range, 5 months to 5.3 years). The mean (± the standard deviation) blip frequency was 0.09 ± 0.11/sample, with about one-third of patients showing no viral blips. The mean viral blip amplitude was 158 ± 132 human immunodeficiency virus type 1 (HIV-1) RNA copies/ml. Analysis of the blip frequency and amplitude distributions suggest that two blips less than 22 days apart have a significant chance of being part of the same episode of viremia. The data are consistent with a hypothetical model in which each episode of viremia consists of a phase of VL rise, followed by two-phase exponential decay. Thus, the term “viral blip” may be a misnomer, since viral replication appears to be occurring over an extended period. Neither the frequency nor the amplitude of viral blips increases with longer periods of observation, but the frequency is inversely correlated with the CD4+-T-cell count at the start of therapy, suggesting that host-specific factors but not treatment fatigue are determinants of blip frequency.

In a Subset of Subjects on Highly Active Antiretroviral Therapy, Human Immunodeficiency Virus Type 1 RNA in Plasma Decays from 50 to <5 Copies per Milliliter, with a Half-Life of 6 Months

ABSTRACT Three of five virally suppressed human immunodeficiency virus type I (HIV-1)-infected patients treated with highly active antiretroviral therapy and followed intensively with a supersensitive reverse transcriptase PCR assay with a lower limit of quantitation of 5 copies/ml showed statistically significant viral load decays below 50 copies/ml, with half-lives of 5 to 8 months and a mean of 6 months. This range of half-lives is consistent with the estimated half-life of the latent HIV-1 reservoir in the peripheral blood. Those patients without decay of viral load in plasma may have significant cryptic HIV-1 residual replication.

Concurrent Measures Of Total And Integrated HIV DNA Monitor Reservoirs And Ongoing Replication In Eradication Trials

Objectives:Interest in targeting HIV reservoirs is fueling trials that may decrease reservoir size and/or induce viral replication. Therefore, we aimed to develop strategies to sensitively measure changes in these parameters in patients on and off antiretroviral therapy (ART). Achieving these goals may help evaluate the effects of future clinical trials. Design:To determine the relationship between measurements of total and integrated HIV DNA and their role as markers of reservoir size and ongoing replication, these parameters were measured during the first year of ART, during long-term effective ART, and during a clinical trial aimed at targeting reservoirs. Methods:Total and integrated HIV DNA were measured in patient samples using quantitative PCR techniques. CD4+T cell counts and plasma viremia were also monitored. Results:Unintegrated HIV DNA became undetectable during the first year of ART. Total and integrated HIV DNA levels were generally equal in well controlled patients on ART, and low-level plasma viremia correlated best with integration measures. Finally, patients who controlled plasma viremia (<400 copies/ml) during interferon-&agr; monotherapy exhibited a decrease in the level of integrated but not total HIV DNA and a rise in the ratio of total to integrated HIV DNA over time. Conclusion:Our findings suggest that appearance of unintegrated HIV DNA reflects residual HIV expression and de-novo reverse transcription, providing insight into the mechanism by which interferon-&agr; reduces the HIV reservoir. We conclude that concurrent measurements of total and integrated HIV DNA provide information regarding reservoir size and ongoing replication in trials targeting HIV.