BACKGROUND. The histone deacetylase (HDAC) inhibitor vorinostat (VOR) can increase HIV RNA expression in vivo within resting CD4+ T cells of aviremic HIV+ individuals. However, while studies of VOR or other HDAC inhibitors have reported reversal of latency, none has demonstrated clearance of latent infection. We sought to identify the optimal dosing of VOR for effective serial reversal of HIV latency. METHODS. In a study of 16 HIV-infected, aviremic individuals, we measured resting CD4+ T cell–associated HIV RNA ex vivo and in vivo following a single exposure to VOR, and then in vivo after a pair of doses separated by 48 or 72 hours, and finally following a series of 10 doses given at 72-hour intervals. RESULTS. Serial VOR exposures separated by 72 hours most often resulted in an increase in cell-associated HIV RNA within circulating resting CD4+ T cells. VOR was well tolerated by all participants. However, despite serial reversal of latency over 1 month of VOR dosing, we did not observe a measurable decrease (>0.3 log10) in the frequency of latent infection within resting CD4+ T cells. CONCLUSIONS. These findings outline parameters for the experimental use of VOR to clear latent infection. Latency reversal can be achieved by VOR safely and repeatedly, but effective depletion of persistent HIV infection will require additional advances. In addition to improvements in latency reversal, these advances may include the sustained induction of potent antiviral immune responses capable of recognizing and clearing the rare cells in which HIV latency has been reversed. TRIAL REGISTRATION. Clinicaltrials.gov NCT01319383. FUNDING. NIH grants U01 AI095052, AI50410, and P30 CA016086 and National Center for Advancing Translational Sciences grant KL2 TR001109.
Nancie M. Archin, Jennifer L. Kirchherr, Julia A.M. Sung, Genevieve Clutton, Katherine Sholtis, Yinyan Xu, Brigitte Allard, Erin Stuelke, Angela D. Kashuba, Joann D. Kuruc, Joseph Eron, Cynthia L. Gay, Nilu Goonetilleke, David M. Margolis
HIV-1 causes a chronic, incurable disease due to its persistence in CD4+ T cells that contain replication-competent provirus, but exhibit little or no active viral gene expression and effectively resist combination antiretroviral therapy (cART). These latently infected T cells represent an extremely small proportion of all circulating CD4+ T cells but possess a remarkable long-term stability and typically persist throughout life, for reasons that are not fully understood. Here we performed massive single-genome, near-full-length next-generation sequencing of HIV-1 DNA derived from unfractionated peripheral blood mononuclear cells, ex vivo-isolated CD4+ T cells, and subsets of functionally polarized memory CD4+ T cells. This approach identified multiple sets of independent, near-full-length proviral sequences from cART-treated individuals that were completely identical, consistent with clonal expansion of CD4+ T cells harboring intact HIV-1. Intact, near-full-genome HIV-1 DNA sequences that were derived from such clonally expanded CD4+ T cells constituted 62% of all analyzed genome-intact sequences in memory CD4 T cells, were preferentially observed in Th1-polarized cells, were longitudinally detected over a duration of up to 5 years, and were fully replication- and infection-competent. Together, these data suggest that clonal proliferation of Th1-polarized CD4+ T cells encoding for intact HIV-1 represents a driving force for stabilizing the pool of latently infected CD4+ T cells.
Guinevere Q. Lee, Nina Orlova-Fink, Kevin Einkauf, Fatema Z. Chowdhury, Xiaoming Sun, Sean Harrington, Hsiao-Hsuan Kuo, Stephane Hua, Hsiao-Rong Chen, Zhengyu Ouyang, Kavidha Reddy, Krista Dong, Thumbi Ndung’u, Bruce D. Walker, Eric S. Rosenberg, Xu G. Yu, Mathias Lichterfeld
Long-acting anti-HIV products can substantively change the standard of care for patients with HIV/AIDS. To this end, hydrophobic antiretroviral drugs (ARVs) were recently developed for parenteral administration at monthly or longer intervals. While shorter-acting hydrophilic drugs can be made into nanocarrier-encased prodrugs, the nanocarrier encasement must be boosted to establish long-acting ARV depots. The mixed-lineage kinase 3 (MLK-3) inhibitor URMC-099 provides this function by affecting autophagy. Here, we have shown that URMC-099 facilitates ARV sequestration and its antiretroviral responses by promoting the nuclear translocation of the transcription factor EB (TFEB). In monocyte-derived macrophages, URMC-099 induction of autophagy led to retention of nanoparticles containing the antiretroviral protease inhibitor atazanavir. These nanoparticles were localized within macrophage autophagosomes, leading to a 4-fold enhancement of mitochondrial and cell vitality. In rodents, URMC-099 activation of autophagy led to 50-fold increases in the plasma drug concentration of the viral integrase inhibitor dolutegravir. These data paralleled URMC-099–mediated induction of autophagy and the previously reported antiretroviral responses in HIV-1–infected humanized mice. We conclude that pharmacologic induction of autophagy provides a means to extend the action of a long-acting, slow, effective release of antiretroviral therapy.
Divya Prakash Gnanadhas, Prasanta K. Dash, Brady Sillman, Aditya N. Bade, Zhiyi Lin, Diana L. Palandri, Nagsen Gautam, Yazen Alnouti, Harris A. Gelbard, JoEllyn McMillan, R. Lee Mosley, Benson Edagwa, Howard E. Gendelman, Santhi Gorantla
Current strategies for HIV-1 eradication require the reactivation of latent HIV-1 in resting CD4+ T cells (rCD4s). Global T cell activation is a well-characterized means of inducing HIV-1 transcription, but is considered too toxic for clinical applications. Here, we have explored a strategy that involves a combination of immune activation and the immunosuppressive mTOR inhibitor rapamycin. In purified rCD4s from HIV-1–infected individuals on antiretroviral therapy, rapamycin treatment downregulated markers of toxicity, including proinflammatory cytokine release and cellular proliferation that were induced after potent T cell activation using αCD3/αCD28 antibodies. Using an ex vivo assay for HIV-1 mRNA, we demonstrated that despite this immunomodulatory effect, rapamycin did not affect HIV-1 gene expression induced by T cell activation in these rCD4s. In contrast, treating activated rCD4s with the immunosuppressant cyclosporin, a calcineurin inhibitor, robustly inhibited HIV-1 reactivation. Importantly, rapamycin treatment did not impair cytotoxic T lymphocyte (CTL) recognition and killing of infected cells. These findings raise the possibility of using rapamycin in conjunction with T cell–activating agents in HIV-1 cure strategies.
Alyssa R. Martin, Ross A. Pollack, Adam Capoferri, Richard F. Ambinder, Christine M. Durand, Robert F. Siliciano
Chronic immune activation, immunosuppression, and T cell exhaustion are hallmarks of HIV infection, yet the mechanisms driving these processes are unclear. Chronic activation can be a driving force in immune exhaustion, and type I interferons (IFN-I) are emerging as critical components underlying ongoing activation in HIV infection. Here, we have tested the effect of blocking IFN-I signaling on T cell responses and virus replication in a murine model of chronic HIV infection. Using HIV-infected humanized mice, we demonstrated that in vivo blockade of IFN-I signaling during chronic HIV infection diminished HIV-driven immune activation, decreased T cell exhaustion marker expression, restored HIV-specific CD8 T cell function, and led to decreased viral replication. Antiretroviral therapy (ART) in combination with IFN-I blockade accelerated viral suppression, further decreased viral loads, and reduced the persistently infected HIV reservoir compared with ART treatment alone. Our data suggest that blocking IFN-I signaling in conjunction with ART treatment can restore immune function and may reduce viral reservoirs during chronic HIV infection, providing validation for IFN-I blockade as a potential therapy for HIV infection.
Anjie Zhen, Valerie Rezek, Cindy Youn, Brianna Lam, Nelson Chang, Jonathan Rick, Mayra Carrillo, Heather Martin, Saro Kasparian, Philip Syed, Nicholas Rice, David G. Brooks, Scott G. Kitchen
Despite the efficient suppression of HIV-1 replication that can be achieved with combined antiretroviral therapy (cART), low levels of type I interferon (IFN-I) signaling persist in some individuals. This sustained signaling may impede immune recovery and foster viral persistence. Here we report studies using a monoclonal antibody to block IFN-α/β receptor (IFNAR) signaling in humanized mice (hu-mice) that were persistently infected with HIV-1. We discovered that effective cART restored the number of human immune cells in HIV-1–infected hu-mice but did not rescue their immune hyperactivation and dysfunction. IFNAR blockade fully reversed HIV-1–induced immune hyperactivation and rescued anti–HIV-1 immune responses in T cells from HIV-1–infected hu-mice. Finally, we found that IFNAR blockade in the presence of cART reduced the size of HIV-1 reservoirs in lymphoid tissues and delayed HIV-1 rebound after cART cessation in the HIV-1–infected hu-mice. We conclude that low levels of IFN-I signaling contribute to HIV-1–associated immune dysfunction and foster HIV-1 persistence in cART-treated hosts. Our results suggest that blocking IFNAR may provide a potential strategy to enhance immune recovery and reduce HIV-1 reservoirs in individuals with sustained elevations in IFN-I signaling during suppressive cART.
Liang Cheng, Jianping Ma, Jingyun Li, Dan Li, Guangming Li, Feng Li, Qing Zhang, Haisheng Yu, Fumihiko Yasui, Chaobaihui Ye, Li-Chung Tsao, Zhiyuan Hu, Lishan Su, Liguo Zhang
A rare subset of HIV-1–infected individuals is able to maintain plasma viral load (VL) at low levels without antiretroviral treatment. Identifying the mechanisms underlying this atypical response to infection may lead to therapeutic advances for treating HIV-1. Here, we developed a proteomic analysis to compare peripheral blood cell proteomes in 20 HIV-1–infected individuals who maintained either high or low VL with the aim of identifying host factors that impact HIV-1 replication. We determined that the levels of multiple histone proteins were markedly decreased in cohorts of individuals with high VL. This reduction was correlated with lower levels of stem-loop binding protein (SLBP), which is known to control histone metabolism. Depletion of cellular SLBP increased promoter engagement with the chromatin structures of the host gene high mobility group protein A1 (
Ming Li, Lynne D. Tucker, John M. Asara, Collins K. Cheruiyot, Huafei Lu, Zhijin J. Wu, Michael C. Newstein, Mark S. Dooner, Jennifer Friedman, Michelle A. Lally, Bharat Ramratnam
In HIV-1–infected patients, increased numbers of circulating CD8+ T cells are linked to increased risk of morbidity and mortality. Here, we identified a bystander mechanism that promotes CD8 T cell activation and expansion in untreated HIV-1–infected patients. Compared with healthy controls, untreated HIV-1–infected patients have an increased population of proliferating, granzyme B+, CD8+ T cells in circulation. Vβ expression and deep sequencing of CDR3 revealed that in untreated HIV-1 infection, cycling memory CD8 T cells possess a broad T cell repertoire that reflects the repertoire of the resting population. This suggests that cycling is driven by bystander activation, rather than specific antigen exposure. Treatment of peripheral blood mononuclear cells with IL-15 induced a cycling, granzyme B+ phenotype in CD8+ T cells. Moreover, elevated IL-15 expression in the lymph nodes of untreated HIV-1–infected patients correlated with circulating CD8+ T cell counts and was normalized in these patients following antiretroviral therapy. Together, these results suggest that IL-15 drives bystander activation of CD8+ T cells, which predicts disease progression in untreated HIV-1–infected patients and suggests that elevated IL-15 may also drive CD8+ T cell expansion that is linked to increased morbidity and mortality in treated patients.
Souheil-Antoine Younes, Michael L. Freeman, Joseph C. Mudd, Carey L. Shive, Arnold Reynaldi, Soumya Panigrahi, Jacob D. Estes, Claire Deleage, Carissa Lucero, Jodi Anderson, Timothy W. Schacker, Miles P. Davenport, Joseph M. McCune, Peter W. Hunt, Sulggi A. Lee, Sergio Serrano-Villar, Robert L. Debernardo, Jeffrey M. Jacobson, David H. Canaday, Rafick-Pierre Sekaly, Benigno Rodriguez, Scott F. Sieg, Michael M. Lederman
Current antiretroviral therapy (ART) is not sufficient to completely suppress disease progression in the CNS, as indicated by the rising incidence of HIV-1–associated neurocognitive disorders (HAND) among infected individuals on ART. It is not clear why some HIV-1–infected patients develop HAND, despite effective repression of viral replication in the circulation. SIV-infected nonhuman primate models are widely used to dissect the mechanisms of viral pathogenesis in the CNS. Here, we identified 4 amino acid substitutions in the cytoplasmic tail of viral envelope glycoprotein gp41 of the neurovirulent virus SIVsm804E that enhance replication in macrophages and associate with enhanced antagonism of the host restriction factor BM stromal cell antigen 2 (BST-2). Rhesus macaques were inoculated with a variant of the parental virus SIVsmE543-3 that had been engineered to contain the 4 amino acid substitutions present in gp41 of SIVsm804E. Compared with WT virus–infected controls, animals infected with mutant virus exhibited higher viral load in cerebrospinal fluid. Together, these results are consistent with a potential role for BST-2 in the CNS microenvironment and suggest that BST-2 antagonists may serve as a possible target for countermeasures against HAND.
Kenta Matsuda, Chia-Yen Chen, Sonya Whitted, Elena Chertova, David J. Roser, Fan Wu, Ronald J. Plishka, Ilnour Ourmanov, Alicia Buckler-White, Jeffrey D. Lifson, Klaus Strebel, Vanessa M. Hirsch
The rare patients who are able to spontaneously control HIV replication in the absence of therapy show signs of a particularly efficient cellular immune response. To identify the molecular determinants that underlie this response, we characterized the T cell receptor (TCR) repertoire directed at Gag293, the most immunoprevalent CD4 epitope in the HIV-1 capsid. HIV controllers from the ANRS CODEX cohort showed a highly skewed TCR repertoire that was characterized by a predominance of
Daniela Benati, Moran Galperin, Olivier Lambotte, Stéphanie Gras, Annick Lim, Madhura Mukhopadhyay, Alexandre Nouël, Kristy-Anne Campbell, Brigitte Lemercier, Mathieu Claireaux, Samia Hendou, Pierre Lechat, Pierre de Truchis, Faroudy Boufassa, Jamie Rossjohn, Jean-François Delfraissy, Fernando Arenzana-Seisdedos, Lisa A. Chakrabarti