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CROI 2012: HIV Persistence, Latency and Eradication
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David H. Shepp, MD
Associate Professor of Medicine
Hofstra North Shore-LIJ School of Medicine
North Shore University Hospital - Manhasset, NY
Combination antiretroviral therapy (ART) durably suppresses HIV replication to levels that allow for improvement in immune function, freedom from opportunistic illnesses and greatly enhance life-expectancy. However, it does not cure HIV infection. Many patients continue to have very low levels of HIV RNA detectable in plasma and lymphoid tissues. Essentially all patients have HIV genomes detected as integrated proviral DNA in long-lived memory CD4+ T-lymphocytes that can produce HIV after cell activation. To date, all attempts to discontinue ART no matter how complete the suppression of HIV RNA or how long the duration of therapy have resulted in viral rebound and immunologic deterioration within a matter of weeks to months. The apparent cure of a single patient ("the Berlin patient") who has remained free of HIV off ART after receiving an allogeneic hematopoietic stem cell transplant (HSCT) for treatment of lymphoma has spurred new interest in seeking an HIV cure. Some research is focused on complete eradication of HIV, while other approaches aim for a "functional" cure, meaning a state where HIV remains detectable but disease progression does not occur even in the absence of ART. An oral abstract session during CROI presented studies on HIV persistence, latency and eradication.
Eradication of HIV. The treatment of the Berlin patient was unique because the stem cell donor was a CCR5Æ32 homozygote whose cells lack a critical HIV entry co-receptor. It is not known if this was the critical factor in achieving a cure or if other factors contributed. Cillo et al. (oral abstract 154) assessed the presence of HIV in 10 patients with HIV-related lymphomas who were treated with autologous HSCT. Nine of 10 continued to have HIV RNA detected in plasma and all had detectable HIV DNA in cells. This study suggests that aggressive myeloablative chemotherapy, as is used in autologous HSCT, does not eradicate HIV. However, the specific chemotherapy regimens used in these patients differed from that used for the Berlin patents. They also did not receive total body irradiation, did not experience graft vs. host disease, as occurs with allogeneic HSCT, were reinfused with their own stem cells, which could have been contaminated with HIV-infected T-cells and not did not lack the CCR5 co-receptor. Therefore the most important aspect of the Berlin patient's treatment leading to eradication of HIV remains uncertain.
When memory CD4+ T-cells bearing latent HIV proviral DNA are activated, HIV RNA is expressed and virions are made. In patients on effective ART, new rounds of infection are prevented. However, cells with latent proviral DNA cannot be selectively activated. Past attempts to purge the latent reservoir by non-selective T-cell activation have produced unacceptable toxicity. This has led to new avenues of research seeking to induce HIV expression without T-cell activation. Cellular mechanisms controlling gene expression are incompletely understood, but histone deacetylases (HDAC) are an important class of enzymes that help maintain the binding of DNA to histones, preventing transcription. Drugs that inhibit HDAC have been shown to enhance HIV gene expression in latently infected T-cells in vitro, without inducing T-cell activation. Vorinostat (VOR), also referred to as SAHA, is an HDAC inhibitor that is approved for the treatment of cutaneous T-cell lymphoma. Margolis reported a small pilot study (n=6) of single-dose VOR in aviremic HIV+ individuals (oral abstract 157LB). HIV RNA expression was measured in cells obtained by leukapheresis before and 4 to 8 hours after the dose. HIV RNA was also measured in plasma using a sensitive single-copy assay. Levels of HIV RNA expression in resting CD4+ cells were significantly increased in all 6 patients (mean increase 4.8X) after VOR while levels of RNA in plasma were unchanged. No drug related adverse effects were seen. This is the first demonstration of in vivo disruption of latency in HIV and is encouraging. However the results are very preliminary. The investigators did not attempt to measure changes in the size of the latent pool, as changes were felt to be unlikely after single dose treatment. VOR is mutagenic in bacteria and mammalian cells and increases fetal loss and malformations in animals. It is still unclear if reduction in the latent pool or eradication can be achieved, what duration of therapy might be needed, or if the safety profile of VOR will allow this agent to be used in longer term studies.
The study of VOR suggested disruption of latency is possible, but data presented in another study suggested this alone may not be sufficient to reduce the latent pool. When memory CD4+ T-cells bearing latent HIV proviral DNA are activated, they undergo apoptosis and are eliminated from the latent pool. The fate of cells that express HIV genes after HDAC inhibitor treatment is not known. Shan et al. (oral abstract 153) isolated resting CD4+ T-cells from HIV+ individuals and treated them with VOR. These cells, which remained in the resting state, were not killed by HIV reactivation alone, remaining viable for up to 18 days in culture. When autologous CD8+ T-cells were added to cultures of latently infected CD4+ T-cells that had been treated with VOR, killing of the CD4+-infected cells occurred when the donor was an elite controller, a patient who maintains plasma HIV RNA below the limit of detection without ART. The cells from elite controllers had good spontaneous cytotoxic T-cell (CTL) activity. However, CD8+ T-cells from 7/8 donors on ART did not have good CTL activity and did not exhibit killing. In vitro stimulation of the CD8+ cells with HIV peptides improved CTL activity and killing. This study suggests that disruption of latency with agents that do not induce T-cell activation may not reduce the latent pool. Adjunctive immunization to stimulate CTL activity may be necessary to effect killing of infected cells after induction of HIV gene expression.
Shan (oral abstract 153) et al published paper in Immunity March 9, 2012
Stimulation of HIV-1-Specific Cytolytic T Lymphocytes Facilitates Elimination of Latent Viral Reservoir after Virus Reactivation
New HIV Eradication Research paper Published by Siliciano et al: Johns Hopkins breakthrough may end AIDS patients' need for lifelong drugs below - (03/11/12)
For reasons of potency, safety and tolerability, known agents that interfere with HIV latency may not be ideal. New agents will likely be needed. Xing et al. (oral abstract 156) described a novel screening assay to identify compounds with activity. Two classes of novel compounds with drug-like properties have been identified. These compounds do not induce cytokine secretion or T-cell activation. Their mechanism of action has not yet be determined.
Initiation of ART in the very early stages of HIV infection may reduce the pool of cells that harbor latent HIV. Buzon et al. (oral abstract 151) measured HIV DNA periodically in patients on long-term ART started during acute HIV (n=9) or chronic HIV (n=26) and in elite controllers (n=37). They also assessed the functional reservoir using an in-vitro reactivation assay. Even very early initiation of ART did not prevent the establishment of a latent reservoir. However, patients treated during acute HIV had levels of total and integrated HIV DNA and in vitro HIV reactivation levels similar to elite controllers and significantly lower than in those treated during chronic infection. A second study presented by Perelson et al (oral abstract 152) used data from 27 patients treated during acute HIV and mathematical modeling to identify correlations between viral load and T-cell dynamics and size of the functional latent reservoir. Their results lend further support to use of early ART initiation to limit the size of the latent reservoir. Because the latent pool is smaller in such patients, those initiating ART during acute HIV may be the best candidates for pilot studies attempting HIV eradication.
Functional Cure. At CROI 2010, much excitement was generated by the initial reports of adoptive transfer of CCR5 modified autologous T-cells as therapy for HIV. At this year's meeting, Tebas et al. (oral abstract 155) gave an update on this pilot study. CD4+ T-cell harvested by leukopheresis from 21 HIV-infected patients well controlled on ART were infected ex-vivo with an replication defective adenovirus vector that delivers a zinc-finger nuclease capable of interrupting the CCR5 gene. The cells were then expanded and re-infused. Rapid and sustained increases in CD4+ T-cells were observed after the infusion. Patients with higher CD4+ T-cell counts at entry had larger increases. The increases were not due to the expansion of genetically modified cells, which remained detectable at levels that only averaged around 2%. Instead, the stimulation of multiple cytokines appeared to be responsible for the expansion of unmodified T-cells. Six patients had a 12 week ART treatment interruption. All had viral load rebound, followed by modest declines. One of 6 patients had HIV RNA levels decline transiently to undetectable just prior to resuming treatment. That patient was a CCR5Æ32 deletion heterozygote and had a much higher proportion of abnormal CCR5 alleles than other patients. There were no controls, such as patients treated with infusion of unmodified T-cells or no infusion, so it is hard to know if the observed patterns were attributable to the CCR5-modified cells. The goal of this treatment is a functional cure by establishing a chimeric state with both HIV-infected and genetically modified, HIV-resistant CD4+ T-cells. These resistant cells could theoretically form the basis of an effective HIV-specific immune response that would control HIV infection in the unmodified cells, but data on the evolution of HIV-specific immune responses was not presented.
More effective immune responses against HIV have been associated with lower HIV RNA levels, lower levels of global immune activation, maintenance of normal CD4+ T-cell counts and non-progression to HIV disease. The required components of an effective immune response are not fully understood, but viral control by CD8+ T-lymphocytes has been frequently been identified. Such responses are usually low or absent in patients with progressive HIV. Ineffective CD8+ T-cells express the surface marker PD-1, a down-regulator of immune responses and a sign of T-cell exhaustion. Velu et al. (oral abstract 158LB) explored the therapeutic potential of PD-1 blockade by administering serial infusion of an anti-PD-1 monoclonal antibody in the macaque/SIV model of AIDS. There was enhancement of CD8+ T-cell anti-HIV activity in some treated animals, with corresponding declines in viral load, that was not observed in any untreated controls. However the number of animals studied was extremely small and statistical analyses were not performed, making any conclusions about the activity of anti-PD-1 antibody therapy premature.
Summary. David Ho and others projected that HIV could be eradicated after seven or so years of ART, as the first estimations of the half life of the latent pool suggested it would eventually decay. This was a common theory in the early HAART era. There were studies done at NIAID and elsewhere which failed to detect HIV DNA in PBMCs in a few patients after years of ART, so treatment was stopped, but all rebounded. It took quite a few years before the widely held belief was that ART alone would never do it. After early hopes that prolonged ART alone could eventually eradicate HIV were not realized, a cure for HIV seemed a very remote possibility. Prospects for either eradication or functional cure remain far off, but now a more identifiable pathway toward these goals can be discerned. A drug that is capable of interrupting HIV latency without T-cell activation has been identified, as have tools to identify new compounds with improved activity and safety. The potential need for additional treatments to eliminate infected cells after disruption of latency, such as immunization, have been identified. Patients treated with ART during very early acute HIV infection have the lowest burden of latently infected cells and may be the best candidates for early eradication studies. The cure of the Berlin patient suggests genetic modification of CCR5 could lead to a cure in other patients, although the contribution of other elements of allogeneic transplantation still requires clarification. Experience with adoptive transfer of autologous CCR5 gene modified T-cell is still very early and has not yet demonstrated the features of a functional cure. Novel approaches like blocking PD-1 to improve HIV-specific CD8+ T-cell function are intriguing but require much more study.
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