TAG HIV Basic Science, Vaccines, and Cure Project Blog

Latency-Reversing and Immunomodulatory Activities of HDAC Inhibitors

Results from the second clinical trial to test the HDAC inhibitor vorinostat as a latency-reversing agent, first presented at CROI 2013, have now been published in the open access journal PLoS Pathogens. The study design differed from the first trial, involving a longer 14-day dosing period. The results are broadly consistent, with a significant increase in cell-associated HIV RNA documented, but no change in multiple measures of the HIV reservoir or HIV RNA as assessed by the standard viral load test. Side effects experienced by participants were grade 1 or 2 (mild or moderate in severity) with lethargy, diarrhea and thrombocytopenia being most common; all resolved after the drug was stopped. The researchers noted long-term changes in expression of multiple cellular genes and caution that this could present a safety concern, stating: “the prolonged changes in host gene expression require careful long term follow up.”

The paper adds to the evidence that HDAC inhibitors can activate at least some latent HIV, but do not shrink the HIV reservoir in most cases, suggesting additional interventions are needed to kill latently infected cells that are induced to produce virus. In terms of the latency-reversing potency of HDAC inhibitors, the results presented to date are consistent with in vitro studies indicating a hierarchy of vorinostat > panobinostat > romidepsin, with the latter mediating the greatest effect based on the detection of HIV RNA by standard viral load testing (these data were presented at the AIDS 2014 conference by Dr. Ole Schmeltz Søgaard and the presentation is available on YouTube). The safety concerns raised by the persistent alterations in host gene expression will require ongoing assessment in all the clinical trials that are being conducted.   

HDAC inhibitors also have effects on the immune system, and this aspect is evaluated in study published recently in Blood. The research did not involve HIV-positive people but rather individuals undergoing hematopoietic cell transplantation who were receiving vorinostat to prevent graft-versus-host disease. The authors found increases in the numbers of immunosuppressive regulatory CD4 T cells (Tregs) and greater suppression by these cells on a per-cell basis, along with reduced inflammatory responses by peripheral blood mononuclear cells.

Similar anti-inflammatory effects were reported in an analysis of HIV-positive participants in a clinical trial of panobinostat that was presented at AIDS 2014; the presentation was covered by Jules Levin of NATAP and the abstract and slides are posted to the NATAP website. There are reasons to believe that the anti-inflammatory activity of HDAC inhibitors could be of some benefit in HIV, but Tregs have also been linked to lower HIV-specific immunity, which could potentially be a negative (particularly if HIV-specific immunity needs to be boosted to eliminate latently infected cells). There is already some evidence that HDAC inhibitors can suppress the killing of HIV-infected cells by virus-specific CD8 T cells in laboratory assays (this paper was published in PLoS Pathogens in August).

Complicating matters further, at the recent Strategies for an HIV Cure conference Jonathan Karn gave a presentation suggesting HDAC inhibitors may enhance natural killer (NK) cell responses by causing downregulation of MHC class I on target cells and upregulating NK cell receptors, possibly consistent with some preliminary evidence presented at the same conference indicating NK cell responses were associated with declines in HIV DNA levels in a few participants in the panobinostat trial.

Additional research will be needed to investigate how the various possible consequences of HDAC inhibition balance out in HIV-positive people, and to assess the extent to which these effects persist after dosing is stopped.

PLoS Pathog. 2014 Nov 13;10(10):e1004473. doi: 10.1371/journal.ppat.1004473. eCollection 2014.

Activation of HIV Transcription with Short-Course Vorinostat in HIV-Infected Patients on Suppressive Antiretroviral Therapy.

Elliott JH, Wightman F, Solomon A, Ghneim K, Ahlers J, Cameron MJ, Smith MZ, Spelman T, McMahon J, Velayudham P, Brown G, Roney J, Watson J, Prince MH, Hoy JF, Chomont N, Fromentin R, Procopio FA, Zeidan J, Palmer S, Odevall L, Johnstone RW, Martin BP, Sinclair E, Deeks SG, Hazuda DJ, Cameron PU, Sékaly RP, Lewin SR.

Abstract

Human immunodeficiency virus (HIV) persistence in latently infected resting memory CD4+ T-cells is the major barrier to HIV cure. Cellular histone deacetylases (HDACs) are important in maintaining HIV latency and histone deacetylase inhibitors (HDACi) may reverse latency by activating HIV transcription from latently infected CD4+ T-cells. We performed a single arm, open label, proof-of-concept study in which vorinostat, a pan-HDACi, was administered 400 mg orally once daily for 14 days to 20 HIV-infected individuals on suppressive antiretroviral therapy (ART). The primary endpoint was change in cell associated unspliced (CA-US) HIV RNA in total CD4+ T-cells from blood at day 14. The study is registered at ClinicalTrials.gov (NCT01365065). Vorinostat was safe and well tolerated and there were no dose modifications or study drug discontinuations. CA-US HIV RNA in blood increased significantly in 18/20 patients (90%) with a median fold change from baseline to peak value of 7.4 (IQR 3.4, 9.1). CA-US RNA was significantly elevated 8 hours post drug and remained elevated 70 days after last dose. Significant early changes in expression of genes associated with chromatin remodeling and activation of HIV transcription correlated with the magnitude of increased CA-US HIV RNA. There were no statistically significant changes in plasma HIV RNA, concentration of HIV DNA, integrated DNA, inducible virus in CD4+ T-cells or markers of T-cell activation. Vorinostat induced a significant and sustained increase in HIV transcription from latency in the majority of HIV-infected patients. However, additional interventions will be needed to efficiently induce virus production and ultimately eliminate latently infected cells.

Blood DOI: http://dx.doi.org/10.1182/blood-2014-10-605238

Histone deacetylase inhibition regulates inflammation and enhances Tregs after allogeneic hematopoietic cell transplantation in humans

Sung Won Choi, Erin Gatza, Guoqing Hou, Yaping Sun, Joel Whitfield, Yeohan Song, Katherine Oravecz-Wilson, Isao Tawara, Charles A. Dinarello, and Pavan Reddy

Key Points

• HDAC inhibition reduced pro-inflammatory cytokines and increased regulatory T cell number and function after allo-HCT.
• HDAC inhibition enhanced STAT-3 acetylation and induced IDO after allo-HCT.

Abstract

We examined immunological responses in patients receiving histone deacetylase (HDAC) inhibition (vorinostat) for graft-versus-host disease (GVHD) prophylaxis following allogeneic hematopoietic cell transplant (allo-HCT). Vorinostat treatment increased histone acetylation in peripheral blood mononuclear cells (PBMC) from treated patients, confirming target HDAC inhibition. HDAC inhibition reduced pro-inflammatory cytokine levels in plasma and from PBMC, decreased ex vivo responses of PBMC to pro-inflammatory TLR-4 stimuli, but did not alter the number or response of conventional T cells (Tconv) to non-specific stimuli. However, the numbers of regulatory T cells (Tregs) were increased, which revealed greater demethylation of the Foxp3 T regulatory-specific demethylation region. Vorinostat-treated patients showed increased expression of CD45RA and CD31 on Tregs, and these Tregs demonstrated greater suppression on a per-cell basis. Consistent with preclinical findings, HDAC inhibition also increased STAT-3 acetylation and induced indoleamine-2,3-dioxygenase (IDO). Our data demonstrate that HDAC inhibition reduces inflammatory responses of PBMC but enhances Tregs after allo-HCT.

Is HIV Weakening Over Time?

A little over nine years ago I wrote a blog post with this same title, about a widely publicized paper claiming that HIV had become less virulent. Although it's grim to be in the position of pouring cold water on optimistic-sounding scenarios, that paper was based on measuring HIV’s ability to replicate using a laboratory test, and other published data raised questions as to whether the test could actually predict differences in disease progression rates. Today, it’s déjà vu all over again because there has been an explosion of very similar media stories positing that HIV is evolving into a “milder form." And once again, the study prompting the coverage relies primarily on laboratory measurements of HIV replication capacity, despite the fact that a prior publication—by several of the same authors—reports that results from this test do not predict the rate of CD4 T cell decline over time.

The new study, by Rebecca Payne and colleagues from the laboratory of Philip Goulder, was published by PNAS yesterday. Helpfully, the full text has been made freely available. Two populations of HIV-positive women are compared, from Gaborone, Botswana and Durban, South Africa. The researchers present evidence that in Botswana, where HIV has been circulating for a longer period, there has been greater virus adaptation to immune responses, leading to more escape mutations and a lower replication capacity. HIV replication capacity is compared using samples from 63 study participants in Gaborone and 16 in Durban, with these participants being closely matched for CD4 T cell counts (important because, as the paper reports, HIV replication capacity in a given individual increases as CD4 T cell counts decline). The average result of the replication capacity test was 0.72 in Gaborone and 0.81 in Durban, indicating that the virus in the former population is slightly less fit. The authors create a mathematical model based on these findings suggesting that both immune responses and the increasingly widespread use of antiretroviral therapy may be contributing to a decline in HIV virulence.

The idea that HIV replication capacity can be linked to disease progression rate derives from a documented correlation between the results of the test and both viral load and CD4 T cell count. However, these correlations are based on measurements taken at single timepoints; in other words, cross-sectional analyses. The question of whether measurement of HIV replication capacity can predict subsequent disease progression rate was addressed in a prior study, which evaluated whether there was a correlation with the rate of CD4 T cell decline over time. No such correlation was found, nor even a hint of one. The discussion section of the Journal of Virology paper containing this result states:

“Therefore, although there may be a benefit to decreased replication capacity (as supported by cross-sectional correlations with viral loads and CD4 counts), the data do not support an enduring benefit or a lasting significant impact of Gag-protease replication capacity on the rate of disease progression, at least once the chronic infection stage has been reached…the long-term clinical impact of immune-driven fitness costs requires further investigation, given the evidence for compensation and the observation that replication capacity does not correlate with the subsequent rate of CD4 decline in chronic infection.” 

It is challenging to try and reconcile this prior result with the claim that the difference in replication capacity found in the new PNAS study would equate to an additional 2.5 years in the average time it takes to progress from HIV infection to AIDS (an estimate offered by Phillip Goulder in the BBC’s coverage). Before concluding that the virulence of HIV is declining, it would be prudent to wait to see if additional studies are able to correlate the apparent differences in HIV replication capacity with differences in CD4 T cell counts and health outcomes over time. 

An additional wrinkle is that there are other studies arguing that HIV virulence is increasing over time; a meta-analysis with this finding was published in 2012 and a new analysis presented at CROI 2014 (and just published in the December issue of The Lancet HIV) reached the same conclusion. 

There might be explanations for these very disparate results, but—pending additional evidence—it seems reasonable to maintain a healthy skepticism about all of them.  

PNAS Published online before print December 1, 2014, doi: 10.1073/pnas.1413339111 

Impact of HLA-driven HIV adaptation on virulence in populations of high HIV seroprevalence

Rebecca Payne, Maximilian Muenchhoff, Jaclyn Mann, Hannah E. Roberts, Philippa Matthews, Emily Adland, Allison Hempenstall, Kuan-Hsiang Huang, Mark Brockman, Zabrina Brumme, Marc Sinclair, Toshiyuki Miura, John Frater, Myron Essex, Roger Shapiro, Bruce D. Walker, Thumbi Ndung’u, Angela R. McLean, Jonathan M. Carlson, and Philip J. R. Goulder

Abstract

It is widely believed that epidemics in new hosts diminish in virulence over time, with natural selection favoring pathogens that cause minimal disease. However, a tradeoff frequently exists between high virulence shortening host survival on the one hand but allowing faster transmission on the other. This is the case in HIV infection, where high viral loads increase transmission risk per coital act but reduce host longevity. We here investigate the impact on HIV virulence of HIV adaptation to HLA molecules that protect against disease progression, such as HLA-B*57 and HLA-B*58:01. We analyzed cohorts in Botswana and South Africa, two countries severely affected by the HIV epidemic. In Botswana, where the epidemic started earlier and adult seroprevalence has been higher, HIV adaptation to HLA including HLA-B*57/58:01 is greater compared with South Africa (P = 7 × 10−82), the protective effect of HLA-B*57/58:01 is absent (P = 0.0002), and population viral replicative capacity is lower (P = 0.03). These data suggest that viral evolution is occurring relatively rapidly, and that adaptation of HIV to the most protective HLA alleles may contribute to a lowering of viral replication capacity at the population level, and a consequent reduction in HIV virulence over time. The potential role in this process played by increasing antiretroviral therapy (ART) access is also explored. Models developed here suggest distinct benefits of ART, in addition to reducing HIV disease and transmission, in driving declines in HIV virulence over the course of the epidemic, thereby accelerating the effects of HLA-mediated viral adaptation.