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To the Editor: While current antiretroviral therapy (ART) has proven to be effective in suppressing HIV, the eradication of the virus remains elusive in people living with HIV (PLWH). Given [...]

Clinical trials including an analytical treatment interruption (ATI) are vital for evaluating the efficacy of novel strategies for HIV remissions. We briefly describe an interactive tool for predicting viral rebound timing in ATI trials and the impact of posttreatment controller (PTC) definitions on PTC frequency estimates. A 4-week viral load threshold of 1000 cps/ml provides both high specificity and sensitivity for PTC detection. PTC frequency varies greatly based on the definition of a PTC.

Sustained, drug-free control of HIV-1 replication is naturally achieved in less than 0.5% of infected individuals (here termed 'elite controllers'), despite the presence of a replication-competent viral reservoir1. Inducing such an ability to spontaneously maintain undetectable plasma viraemia is a major objective of HIV-1 cure research, but the characteristics of proviral reservoirs in elite controllers remain to be determined. Here, using next-generation sequencing of near-full-length single HIV-1 genomes and corresponding chromosomal integration sites, we show that the proviral reservoirs of elite controllers frequently consist of oligoclonal to near-monoclonal clusters of intact proviral sequences. In contrast to individuals treated with long-term antiretroviral therapy, intact proviral sequences from elite controllers were integrated at highly distinct sites in the human genome and were preferentially located in centromeric satellite DNA or in Krüppel-associated box domain-containing zinc finger genes on chromosome 19, both of which are associated with heterochromatin features. Moreover, the integration sites of intact proviral sequences from elite controllers showed an increased distance to transcriptional start sites and accessible chromatin of the host genome and were enriched in repressive chromatin marks. These data suggest that a distinct configuration of the proviral reservoir represents a structural correlate of natural viral control, and that the quality, rather than the quantity, of viral reservoirs can be an important distinguishing feature for a functional cure of HIV-1 infection. Moreover, in one elite controller, we were unable to detect intact proviral sequences despite analysing more than 1.5 billion peripheral blood mononuclear cells, which raises the possibility that a sterilizing cure of HIV-1 infection, which has previously been observed only following allogeneic haematopoietic stem cell transplantation2,3, may be feasible in rare instances.

If strategies currently in development succeed in eradicating HIV reservoirs in peripheral blood and lymphoid tissues, residual sources of virus may remain in anatomic compartments. Paired blood and semen samples were collected from 12 individuals enrolled in a randomized, double-blind, placebo-controlled therapeutic vaccine clinical trial in people with HIV (PWH) who began antiretroviral therapy (ART) during acute or early infection (ClinicalTrials registration no. NCT01859325). After the week 56 visit (postintervention), all participants interrupted ART. At the first available time points after viral rebound, we sequenced HIV-1 env (C2-V3), gag (p24), and pol (reverse transcriptase) regions amplified from cell-free HIV RNA in blood and seminal plasma using the MiSeq Illumina platform. Comprehensive sequence and phylogenetic analyses were performed to evaluate viral population structure, compartmentalization, and viral diversity in blood and seminal plasma. Compared to that in blood, HIV RNA rebound in semen occurred significantly later (median of 66 versus 42 days post-ART interruption, P < 0.01) and reached lower levels (median 164 versus 16,090 copies/ml, P < 0.01). Three of five participants with available sequencing data presented compartmentalized viral rebound between blood and semen in one HIV coding region. Despite early ART initiation, HIV RNA molecular diversity was higher in semen than in blood in all three coding regions for most participants. Higher HIV RNA molecular diversity in the genital tract (compared to that in blood plasma) and evidence of compartmentalization illustrate the distinct evolutionary dynamics between these two compartments after ART interruption. Future research should evaluate whether the genital compartment might contribute to viral rebound in some PWH interrupting ART.IMPORTANCE To cure HIV, we likely need to target the reservoirs in all anatomic compartments. Here, we used sophisticated statistical and phylogenetic methods to analyze blood and semen samples collected from 12 persons with HIV who began antiretroviral therapy (ART) during very early HIV infection and who interrupted their ART as part of a clinical trial. First, we found that HIV RNA rebound in semen occurred significantly later and reached lower levels than in blood. Second, we found that the virus in semen was genetically different in some participants compared to that in blood. Finally, we found increased HIV RNA molecular diversity in semen compared to that in blood in almost all study participants. These data suggest that the HIV RNA populations emerging from the genital compartment after ART interruption might not be the same as those emerging from blood plasma. Future research should evaluate whether the genital compartment might contribute to viral rebound in some people with HIV (PWH) interrupting ART.

AIDS is a pandemic disease caused by HIV that affects 37 million people worldwide. Current antiretroviral therapy slows disease progression but does not eliminate latently infected cells, which resupply active virus, thus necessitating lifelong treatment with associated compliance, cost, and chemoexposure issues. Latency-reversing agents (LRAs) activate these cells, allowing for their potential clearance, thus presenting a strategy to eradicate the infection. Protein kinase C (PKC) modulators-including prostratin, ingenol esters, bryostatin, and their analogs-are potent LRAs in various stages of development for several clinical indications. While LRAs are promising, a major challenge associated with their clinical use is sustaining therapeutically meaningful levels of the active agent while minimizing side effects. Here we describe a strategy to address this problem based on LRA prodrugs, designed for controllable release of the active LRA after a single injection. As intended, these prodrugs exhibit comparable or superior in vitro activity relative to the parent compounds. Selected compounds induced higher in vivo expression of CD69, an activation biomarker, and, by releasing free agent over time, significantly improved tolerability when compared to the parent LRAs. More generally, selected prodrugs of PKC modulators avoid the bolus toxicities of the parent drug and exhibit greater efficacy and expanded tolerability, thereby addressing a longstanding objective for many clinical applications.

BACKGROUND:We aimed to characterize the impact of ART initiation on GALT at various sites along the gastrointestinal site. METHODOLOGY:Peripheral blood and duodenal and rectal biopsies were obtained from 12 HIV and 33 treatment-naïve HIV subjects at baseline and after 9-months ART. Tissue was digested for immunophenotyping. Inflammatory, bacterial translocation and intestinal damage markers were measured in plasma. RESULTS:Twenty-six HIV subjects completed follow-up. The lowest reconstitution of CD4 T-cells and the lowest CD4/CD8 ratio during ART compared to blood were observed in the duodenum with the rectum being either intermediate or approaching blood levels. Regulatory T-cells (Treg) were in higher proportions in the duodenum than the rectum and neither declined significantly during ART. Several correlations with biomarkers of microbial translocation were observed including increases in LTA levels, which reflects gram-positive bacterial translocation, correlated with increases in %CD4 T-cells in duodenum (Rho 0.773, P = 0.033), and with decreases duodenal Treg populations (Rho -0.40, P = 0.045). CONCLUSIONS:HIV-mediated immunological disruption is greater in the duodenum than rectum and blood before and during ART. Small intestine damage may represent a unique environment for T-cell depletion, which might be attenuated by interaction with gram positive bacteria.

BackgroundHIV posttreatment controllers are rare individuals who start antiretroviral therapy (ART), but maintain HIV suppression after treatment interruption. The frequency of posttreatment control and posttreatment interruption viral dynamics have not been well characterized.MethodsPosttreatment controllers were identified from 14 studies and defined as individuals who underwent treatment interruption with viral loads ≤400 copies/mL at two-thirds or more of time points for ≥24 weeks. Viral load and CD4+ cell dynamics were compared between posttreatment controllers and noncontrollers.ResultsOf the 67 posttreatment controllers identified, 38 initiated ART during early HIV infection. Posttreatment controllers were more frequently identified in those treated during early versus chronic infection (13% vs 4%, P < .001). In posttreatment controllers with weekly viral load monitoring, 45% had a peak posttreatment interruption viral load of ≥1000 copies/mL and 33% had a peak viral load ≥10000 copies/mL. Of posttreatment controllers, 55% maintained HIV control for 2 years, with approximately 20% maintaining control for ≥5 years.ConclusionsPosttreatment control was more commonly identified amongst early treated individuals, frequently characterized by early transient viral rebound and heterogeneous durability of HIV remission. These results may provide mechanistic insights and have implications for the design of trials aimed at achieving HIV remission.

BackgroundNotwithstanding 1 documented case of HIV-1 cure following allogeneic stem cell transplantation (allo-SCT), several subsequent cases of allo-SCT in HIV-1 positive individuals have failed to cure HIV-1 infection. The aim of our study was to describe changes in the HIV reservoir in a single chronically HIV-infected patient on suppressive antiretroviral therapy who underwent allo-SCT for treatment of acute lymphoblastic leukemia.Methods and findingsWe prospectively collected peripheral blood mononuclear cells (PBMCs) by leukapheresis from a 55-year-old man with chronic HIV infection before and after allo-SCT to measure the size of the HIV-1 reservoir and characterize viral phylogeny and phenotypic changes in immune cells. At day 784 post-transplant, when HIV-1 was undetectable by multiple measures-including PCR measurements of both total and integrated HIV-1 DNA, replication-competent virus measurement by large cell input quantitative viral outgrowth assay, and in situ hybridization of colon tissue-the patient consented to an analytic treatment interruption (ATI) with frequent clinical monitoring. He remained aviremic off antiretroviral therapy until ATI day 288, when a low-level virus rebound of 60 HIV-1 copies/ml occurred, which increased to 1,640 HIV-1 copies/ml 5 days later, prompting reinitiation of ART. Rebounding plasma HIV-1 sequences were phylogenetically distinct from proviral HIV-1 DNA detected in circulating PBMCs before transplantation. The main limitations of this study are the insensitivity of reservoir measurements, and the fact that it describes a single case.Conclusionsallo-SCT led to a significant reduction in the size of the HIV-1 reservoir and a >9-month-long ART-free remission from HIV-1 replication. Phylogenetic analyses suggest that the origin of rebound virus was distinct from the viruses identified pre-transplant in the PBMCs.

Plasma, duodenal, and rectal tissue antiretroviral therapy (ART) drug concentrations, human immunodeficiency virus (HIV) RNA and HIV DNA copy numbers, and recovery of mucosal immunity were measured before and 9 months after initiation of 3 different ART regimens in 26 subjects. Plasma and tissue HIV RNA correlated at baseline and when 9-month declines were compared, suggesting that these compartments are tightly associated. Antiretroviral tissue:blood penetration ratios were above the 50% inhibitory concentration values in almost 100% of cases. There were no correlations between drug concentrations and HIV DNA/RNA. Importantly, no evidence was found for residual viral replication or deficient tissue drug penetration to account for delayed gastrointestinal-associated lymphoid tissue immune recovery.

The ability of HIV to establish a long-lived latent infection within resting CD4+ T cells leads to persistence and episodic resupply of the virus in patients treated with antiretroviral therapy (ART), thereby preventing eradication of the disease. Protein kinase C (PKC) modulators such as bryostatin 1 can activate these latently infected cells, potentially leading to their elimination by virus-mediated cytopathic effects, the host's immune response and/or therapeutic strategies targeting cells actively expressing virus. While research in this area has focused heavily on naturally-occurring PKC modulators, their study has been hampered by their limited and variable availability, and equally significantly by sub-optimal activity and in vivo tolerability. Here we show that a designed, synthetically-accessible analog of bryostatin 1 is better-tolerated in vivo when compared with the naturally-occurring product and potently induces HIV expression from latency in humanized BLT mice, a proven and important model for studying HIV persistence and pathogenesis in vivo. Importantly, this induction of virus expression causes some of the newly HIV-expressing cells to die. Thus, designed, synthetically-accessible, tunable, and efficacious bryostatin analogs can mediate both a "kick" and "kill" response in latently-infected cells and exhibit improved tolerability, therefore showing unique promise as clinical adjuvants for HIV eradication.

To the Editor: HIV remains an uncurable disease in infected individuals receiving antiretroviral therapy (ART). Despite unparalleled research efforts to find a cure, there is a growing sentiment in the [...]

Whether initiation of antiretroviral therapy (ART) regimens aimed at achieving greater concentrations within gut associated lymphoid tissue (GALT) impacts the level of mucosal immune reconstitution, inflammatory markers and the viral reservoir remains unknown. We included 12 HIV- controls and 32 ART-naïve HIV patients who were randomized to efavirenz, maraviroc or maraviroc+raltegravir, each with fixed-dose tenofovir disoproxil fumarate/emtricitabine. Rectal and duodenal biopsies were obtained at baseline and at 9 months of ART. We performed a comprehensive assay of T-cell subsets by flow cytometry, T-cell density in intestinal biopsies, plasma and tissue concentrations of antiretroviral drugs by high-performance liquid chromatography/mass spectroscopy, and plasma interleukin-6 (IL-6), lipoteichoic acid (LTA), soluble CD14 (sCD14) and zonulin-1 each measured by ELISA. Total cell-associated HIV DNA was measured in PBMC and rectal and duodenal mononuclear cells. Twenty-six HIV-infected patients completed the follow-up. In the duodenum, the quadruple regimen resulted in greater CD8+ T-cell density decline, greater normalization of mucosal CCR5+CD4+ T-cells and increase of the naïve/memory CD8+ T-cell ratio, and a greater decline of sCD14 levels and duodenal HIV DNA levels (P = 0.004 and P = 0.067, respectively), with no changes in HIV RNA in plasma or tissue. Maraviroc showed the highest drug distribution to the gut tissue, and duodenal concentrations correlated well with other T-cell markers in duodenum, i.e., the CD4/CD8 ratio, %CD4+ and %CD8+ HLA-DR+CD38+ T-cells. Maraviroc use elicited greater activation of the mucosal naïve CD8+ T-cell subset, ameliorated the distribution of the CD8+ T-cell maturational subsets and induced higher improvement of zonulin-1 levels. These data suggest that combined CCR5 and integrase inhibitor based combination therapy in ART treatment naïve patients might more effectively reconstitute duodenal immunity, decrease inflammatory markers and impact on HIV persistence by cell-dependent mechanisms, and show unique effects of MVC in duodenal immunity driven by higher drug tissue penetration and possibly by class-dependent effects.

BackgroundEarly initiation of combination antiretroviral therapy (cART) to human immunodeficiency virus type 1 (HIV-1)-infected infants controls HIV-1 replication and reduces mortality.MethodsPlasma viremia (lower limit of detection,

Background A better understanding of the dynamics of human immunodeficiency virus (HIV) reservoirs in CD4+ T cells of people with HIV (PWH) receiving antiretroviral therapy (ART) is crucial for developing therapies to eradicate the virus. Methods We conducted a study involving 28 aviremic PWH receiving ART with high and low levels of HIV DNA. We analyzed immunologic and virologic parameters and their association with the HIV reservoir size. Results The frequency of CD4+ T cells carrying HIV DNA was associated with higher pre-ART plasma viremia, lower pre-ART CD4+ T-cell counts, and lower pre-ART CD4/CD8 ratios. During ART, the High group maintained elevated levels of intact HIV proviral DNA, cell-associated HIV RNA, and inducible virion-associated HIV RNA. HIV sequence analysis showed no evidence for preferential accumulation of defective proviruses nor higher frequencies of clonal expansion in the High versus Low group. Phenotypic and functional T-cell analyses did not show enhanced immune-mediated virologic control in the Low versus High group. Of considerable interest, pre-ART innate immunity was significantly higher in the Low versus High group. Conclusions Our data suggest that innate immunity at the time of ART initiation may play an important role in modulating the dynamics and persistence of viral reservoirs in PWH. [ABSTRACT FROM AUTHOR]

A number of highly potent and broadly neutralizing antibodies (bNAbs) against the human immunodeficiency virus (HIV) have recently been shown to prevent transmission of the virus, suppress viral replication, and delay plasma viral rebound following discontinuation of antiretroviral therapy in animal models and infected humans. However, the degree and extent to which such bNAbs interact with primary lymphocytes have not been fully delineated. Here, we show that certain glycan-dependent bNAbs, such as PGT121 and PGT151, bind to B, activated T, and natural killer (NK) cells of HIV-infected and -uninfected individuals. Binding of these bNAbs, particularly PGT121 and PGT151, to activated [CD4.sup.+] and [CD8.sup.+] T cells was mediated by complex-type glycans and was abrogated by enzymatic inhibition of N-linked glycosylation. In addition, a short-term incubation of PGT151 and primary NK cells led to degranulation and cellular death. Our data suggest that the propensity of certain bNAbs to bind uninfected/bystander cells has the potential for unexpected outcomes in passive-transfer studies and underscore the importance of antibody screening against primary lymphocytes., Introduction Recent advances in antibody cloning technologies have led to the isolation of a number of highly potent and broadly neutralizing antibodies (bNAbs) against the human immunodeficiency virus (HIV). Some [...]

Chromosomal integration of genome-intact HIV-1 sequences into the host genome creates a reservoir of virally infected cells that persists throughout life, necessitating indefinite antiretroviral suppression therapy. During effective antiviral treatment, the majority of these proviruses remain transcriptionally silent, but mechanisms responsible for viral latency are insufficiently clear. Here, we used matched integration site and proviral sequencing (MIP-Seq), an experimental approach involving multiple displacement amplification of individual proviral species, followed by near- full-length HIV-1 next-generation sequencing and corresponding chromosomal integration site analysis to selectively map the chromosomal positions of intact and defective proviruses in 3 HIV-1-infected individuals undergoing long-term antiretroviral therapy. Simultaneously, chromatin accessibility and gene expression in autologous CD4* T cells were analyzed by assays for transposase-accessible chromatin using sequencing (ATAC-Seq) and RNA-Seq. We observed that in comparison to proviruses with defective sequences, intact HIV-1 proviruses were enriched for non-genic chromosomal positions and more frequently showed an opposite orientation relative to host genes. In addition, intact HIV-1 proviruses were preferentially integrated in either relative proximity to or increased distance from active transcriptional start sites and to accessible chromatin regions. These studies strongly suggest selection of intact proviruses with features of deeper viral latency during prolonged antiretroviral therapy, and may be informative for targeting the genome-intact viral reservoir., Introduction HIV-1 integration into host chromosomes, catalyzed by the viral enzyme integrase, allows the virus to persist in the human organism for the life span of the infected cell and [...]

An infant born to a woman with human immunodeficiency virus type 1 (HIV-1) infection began receiving antiretroviral therapy (ART) 30 hours after birth owing to high-risk exposure. ART was continued when detection of HIV-1 DNA and RNA on repeat testing met the standard diagnostic criteria for infection. After therapy was discontinued (when the child was 18 months of age), levels of plasma HIV-1 RNA, proviral DNA in peripheral-blood mononuclear cells, and HIV-1 antibodies, as assessed by means of clinical assays, remained undetectable in the child through 30 months of age. This case suggests that very early ART in infants may alter the establishment and long-term persistence of HIV-1 infection.

There is intense interest in developing curative interventions for HIV. How such a cure will be quantified and defined is not known. We applied a series of measurements of HIV persistence to the study of an HIV-infected adult who has exhibited evidence of cure after allogeneic hematopoietic stem cell transplant from a homozygous CCR5Δ32 donor. Samples from blood, spinal fluid, lymph node, and gut were analyzed in multiple laboratories using different approaches. No HIV DNA or RNA was detected in peripheral blood mononuclear cells (PBMC), spinal fluid, lymph node, or terminal ileum, and no replication-competent virus could be cultured from PBMCs. However, HIV RNA was detected in plasma (2 laboratories) and HIV DNA was detected in the rectum (1 laboratory) at levels considerably lower than those expected in ART-suppressed patients. It was not possible to obtain sequence data from plasma or gut, while an X4 sequence from PBMC did not match the pre-transplant sequence. HIV antibody levels were readily detectable but declined over time; T cell responses were largely absent. The occasional, low-level PCR signals raise the possibility that some HIV nucleic acid might persist, although they could also be false positives. Since HIV levels in well-treated individuals are near the limits of detection of current assays, more sensitive assays need to be developed and validated. The absence of recrudescent HIV replication and waning HIV-specific immune responses five years after withdrawal of treatment provide proof of a clinical cure.

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 group's recommended key goals for the international community.

HIV-1 infection elevates the risk of developing various cancers, including T-cell lymphoma. Whether HIV-1-encoded proteins directly contribute to oncogenesis remains unknown. We observe that approximately 1–5% of CD4+ T cells from the blood of people living with HIV-1 exhibit over-duplicated centrioles, suggesting that centrosome amplification underlies the development of HIV-1-associated cancers by driving aneuploidy. Through affinity purification, biochemical, and cellular analyses, we discover that Vpr, an accessory protein of HIV-1, hijacks the centriole duplication machinery and induces centrosome amplification and aneuploidy. Mechanistically, Vpr forms a cooperative ternary complex with an E3 ligase subunit, VprBP, and polo-like kinase 4 (Plk4). Unexpectedly, however, the complex enhances Plk4's functionality by promoting its relocalization to the procentriole assembly and induces centrosome amplification. Loss of either Vpr's C-terminal 17 residues or VprBP acidic region, the two elements required for binding to Plk4 cryptic polo-box, abrogates Vpr's capacity to induce these events. Furthermore, HIV-1 WT, but not its Vpr mutant, induces multiple centrosomes and aneuploidy in human primary CD4+ T cells. We propose that the Vpr•VprBP•Plk4 complex serves as a molecular link that connects HIV-1 infection to oncogenesis and that inhibiting the Vpr C-terminal motif may reduce the occurrence of HIV-1-associated cancers. People living with HIV-1 are at an increased risk of developing various cancers. Here, the authors suggest that HIV-1-encoded Vpr can promote oncogenesis by forming a ternary complex with VprBP and Plk4 and inducing Plk4-dependent centriole overduplication and aneuploidy. [ABSTRACT FROM AUTHOR]