RESUMEN
Antiviral therapies with reduced frequencies of administration and high barriers to resistance remain a major goal. For HIV, theories have proposed that viral-deletion variants, which conditionally replicate with a basic reproductive ratio [R0] > 1 (termed "therapeutic interfering particles" or "TIPs"), could parasitize wild-type virus to constitute single-administration, escape-resistant antiviral therapies. We report the engineering of a TIP that, in rhesus macaques, reduces viremia of a highly pathogenic model of HIV by >3log10 following a single intravenous injection. Animal lifespan was significantly extended, TIPs conditionally replicated and were continually detected for >6 months, and sequencing data showed no evidence of viral escape. A single TIP injection also suppressed virus replication in humanized mice and cells from persons living with HIV. These data provide proof of concept for a potential new class of single-administration antiviral therapies.
Asunto(s)
Partículas Similares a Virus Artificiales , Eliminación de Gen , Infecciones por VIH , VIH-1 , Interferencia Viral , Replicación Viral , Animales , Humanos , Ratones , Número Básico de Reproducción , Modelos Animales de Enfermedad , Ingeniería Genética , Infecciones por VIH/terapia , Infecciones por VIH/virología , VIH-1/genética , VIH-1/fisiología , Macaca mulatta , Prueba de Estudio Conceptual , Virus de la Inmunodeficiencia de los Simios/genética , Virus de la Inmunodeficiencia de los Simios/fisiología , Viremia/terapia , Viremia/virologíaRESUMEN
Recently published near full-length KSHV genomes from a Cameroon Kaposi sarcoma case-control study showed strong evidence of viral recombination and mixed infections, but no sequence variations associated with disease. Using the same methodology, an additional 102 KSHV genomes from 76 individuals with KSHV-associated diseases have been sequenced. Diagnoses comprise all KSHV-associated diseases (KAD): Kaposi sarcoma (KS), primary effusion lymphoma (PEL), KSHV-associated large cell lymphoma (KSHV-LCL), a type of multicentric Castleman disease (KSHV-MCD), and KSHV inflammatory cytokine syndrome (KICS). Participants originated from 22 different countries, providing the opportunity to obtain new near full-length sequences of a wide diversity of KSHV genomes. These include near full-length sequence of genomes with KSHV K1 subtypes A, B, C, and F as well as subtype E, for which no full sequence was previously available. High levels of recombination were observed. Fourteen individuals (18%) showed evidence of infection with multiple KSHV variants (from two to four unique genomes). Twenty-six comparisons of sequences, obtained from various sampling sites including PBMC, tissue biopsies, oral fluids, and effusions in the same participants, identified near complete genome conservation between different biological compartments. Polymorphisms were identified in coding and non-coding regions, including indels in the K3 and K15 genes and sequence inversions here reported for the first time. One such polymorphism in KSHV ORF46, specific to the KSHV K1 subtype E2, encoded a mutation in the leucine loop extension of the uracil DNA glycosylase that results in alteration of biochemical functions of this protein. This confirms that KSHV sequence variations can have functional consequences warranting further investigation. This study represents the largest and most diverse analysis of KSHV genome sequences to date among individuals with KAD and provides important new information on global KSHV genomics.
Asunto(s)
Genoma Viral , Herpesvirus Humano 8 , Sarcoma de Kaposi , Humanos , Herpesvirus Humano 8/genética , Sarcoma de Kaposi/virología , Sarcoma de Kaposi/genética , Masculino , Femenino , Persona de Mediana Edad , Adulto , Polimorfismo Genético , Anciano , Infecciones por Herpesviridae/genética , Infecciones por Herpesviridae/virología , Etnicidad/genética , Enfermedad de Castleman/virología , Enfermedad de Castleman/genética , FilogeniaRESUMEN
An alternative to lifelong antiretroviral therapy (ART) is needed to achieve durable control of HIV-1. Here we show that adeno-associated virus (AAV)-delivery of two rhesus macaque antibodies to the SIV envelope glycoprotein (Env) with potent neutralization and antibody-dependent cellular cytotoxicity can prevent viral rebound in macaques infected with barcoded SIVmac239M after discontinuing suppressive ART. Following AAV administration, sustained antibody expression with minimal anti-drug antibody responses was achieved in all but one animal. After ART withdrawal, SIV replication rebounded within two weeks in all of the control animals but remained below the threshold of detection in plasma (<15 copies/mL) for more than a year in four of the eight animals that received AAV vectors encoding Env-specific antibodies. Viral sequences from animals with delayed rebound exhibited restricted barcode diversity and antibody escape. Thus, sustained expression of antibodies with potent antiviral activity can afford durable, ART-free containment of pathogenic SIV infection.
RESUMEN
Antibody-mediated depletion studies have demonstrated that CD8+ T cells are required for effective immune control of SIV. However, this approach is potentially confounded by several factors, including reactive CD4+ T cell proliferation, and provides no information on epitope specificity, a likely determinant of CD8+ T cell efficacy. We circumvented these limitations by selectively depleting CD8+ T cells specific for the Gag epitope CTPYDINQM (CM9) via the administration of immunotoxin-conjugated tetrameric complexes of CM9/Mamu-A*01. Immunotoxin administration effectively depleted circulating but not tissue-localized CM9-specific CD8+ T cells, akin to the bulk depletion pattern observed with antibodies directed against CD8. However, we found no evidence to indicate that circulating CM9-specific CD8+ T cells suppressed viral replication in Mamu-A*01+ rhesus macaques during acute or chronic progressive infection with a pathogenic strain of SIV. This observation extended to macaques with established infection during and after continuous antiretroviral therapy. In contrast, natural controller macaques experienced dramatic increases in plasma viremia after immunotoxin administration, highlighting the importance of CD8+ T cell-mediated immunity against CM9. Collectively, these data showed that CM9-specific CD8+ T cells were necessary but not sufficient for robust immune control of SIV in a nonhuman primate model and, more generally, validated an approach that could inform the design of next-generation vaccines against HIV-1.
Asunto(s)
Linfocitos T CD8-positivos , Inmunotoxinas , Macaca mulatta , Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Animales , Linfocitos T CD8-positivos/inmunología , Virus de la Inmunodeficiencia de los Simios/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Inmunotoxinas/inmunología , Inmunotoxinas/farmacología , Productos del Gen gag/inmunología , Replicación Viral/inmunología , Replicación Viral/efectos de los fármacos , Depleción Linfocítica/métodosRESUMEN
The capacity of HIV-1 to replicate during optimal antiretroviral therapy (ART) is challenging to assess directly. To gain greater sensitivity to detect evolution on ART, we used a nonhuman primate (NHP) model providing precise control over the level of pre-ART evolution and more comprehensive analyses than are possible with clinical samples. We infected 21 rhesus macaques (RMs) with the barcoded virus SIVmac239M and initiated ART early to minimize baseline genetic diversity. RMs were treated for 285-1200 days. We used several tests of molecular evolution to compare 1352 near-full-length (nFL) SIV DNA single genome sequences from PBMCs, lymph nodes, and spleen obtained near the time of ART initiation and those present after long-term ART, none of which showed significant changes to the SIV DNA population during ART in any animal. To investigate the possibility of ongoing replication in unsampled putative tissue sanctuaries during ART, we discontinued treatment in four animals and confirmed that none of the 336 nFL SIV RNA sequences obtained from rebound plasma viremia showed evidence of evolution. The rigorous nature of our analyses reinforced the emerging consensus of a lack of appreciable ongoing replication on effective ART and validates the relevance of this NHP model for cure studies.
Asunto(s)
Antirretrovirales , Macaca mulatta , Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Replicación Viral , Animales , Virus de la Inmunodeficiencia de los Simios/genética , Virus de la Inmunodeficiencia de los Simios/efectos de los fármacos , Virus de la Inmunodeficiencia de los Simios/fisiología , Síndrome de Inmunodeficiencia Adquirida del Simio/virología , Síndrome de Inmunodeficiencia Adquirida del Simio/tratamiento farmacológico , Replicación Viral/efectos de los fármacos , Antirretrovirales/uso terapéutico , Evolución Molecular , ARN Viral/genética , Carga Viral/efectos de los fármacos , Viremia/tratamiento farmacológico , Viremia/virología , ADN Viral/genética , MasculinoRESUMEN
Identifying immune correlates of protection is a major challenge in AIDS vaccine development. Anti-Envelope antibodies have been considered critical for protection against SIV/HIV (SHIV) acquisition. Here, we evaluated the efficacy of an SHIV vaccine against SIVmac251 challenge, where the role of antibody was excluded, as there was no cross-reactivity between SIV and SHIV envelope antibodies. After 8 low-dose intrarectal challenges with SIVmac251, 12 SHIV-vaccinated animals demonstrated efficacy, compared with 6 naive controls, suggesting protection was achieved in the absence of anti-envelope antibodies. Interestingly, CD8+ T cells (and some NK cells) were not essential for preventing viral acquisition, as none of the CD8-depleted macaques were infected by SIVmac251 challenges. Initial investigation of protective innate immunity revealed that protected animals had elevated pathways related to platelet aggregation/activation and reduced pathways related to interferon and responses to virus. Moreover, higher expression of platelet factor 4 on circulating platelet-leukocyte aggregates was associated with reduced viral acquisition. Our data highlighted the importance of innate immunity, identified mechanisms, and may provide opportunities for novel HIV vaccines or therapeutic strategy development.
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Linfocitos T CD8-positivos , Inmunidad Innata , Macaca mulatta , Vacunas contra el SIDAS , Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Animales , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/prevención & control , Virus de la Inmunodeficiencia de los Simios/inmunología , Vacunas contra el SIDAS/inmunología , Inmunidad Innata/inmunología , Linfocitos T CD8-positivos/inmunología , Anticuerpos Antivirales/inmunología , Masculino , Vacunas Atenuadas/inmunologíaRESUMEN
The rebound competent viral reservoir (RCVR)-virus that persists during antiretroviral treatment (ART) and can reignite systemic infection when treatment is stopped-is the primary barrier to eradicating HIV. We used time to initiation of ART during primary infection of rhesus macaques (RMs) after intravenous challenge with barcoded SIVmac239 as a means to elucidate the dynamics of RCVR establishment in groups of RMs by creating a multi-log range of pre-ART viral loads and then assessed viral time-to-rebound and reactivation rates resulting from the discontinuation of ART after one year. RMs started on ART on days 3, 4, 5, 6, 7, 9 or 12 post-infection showed a nearly 10-fold difference in pre-ART viral measurements for successive ART-initiation timepoints. Only 1 of 8 RMs initiating ART on days 3 and 4 rebounded after ART interruption despite measurable pre-ART plasma viremia. Rebounding plasma from the 1 rebounding RM contained only a single barcode lineage detected at day 50 post-ART. All RMs starting ART on days 5 and 6 rebounded between 14- and 50-days post-ART with 1-2 rebounding variants each. RMs starting ART on days 7, 9, and 12 had similar time-to-measurable plasma rebound kinetics despite multiple log differences in pre-ART plasma viral load (pVL), with all RMs rebounding between 7- and 16-days post-ART with 3-28 rebounding lineages. Calculated reactivation rates per pre-ART pVL were highest for RMs starting ART on days 5, 6, and 7 after which the rate of accumulation of the RCVR markedly decreased for RMs treated on days 9 and 12, consistent with multiphasic establishment and near saturation of the RCVR within 2 weeks post infection. Taken together, these data highlight the heterogeneity of the RCVR between RMs, the stochastic establishment of the very early RCVR, and the saturability of the RCVR prior to peak viral infection.
Asunto(s)
Infecciones por VIH , Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Animales , Síndrome de Inmunodeficiencia Adquirida del Simio/tratamiento farmacológico , Virus de la Inmunodeficiencia de los Simios/fisiología , Macaca mulatta , Replicación Viral , Antirretrovirales/uso terapéutico , Antirretrovirales/farmacología , Infecciones por VIH/tratamiento farmacológico , Carga ViralRESUMEN
HIV-1 persistence during ART is due to the establishment of long-lived viral reservoirs in resting immune cells. Using an NHP model of barcoded SIVmac239 intravenous infection and therapeutic dosing of anti-TGFBR1 inhibitor galunisertib (LY2157299), we confirm the latency reversal properties of in vivo TGF-ß blockade, decrease viral reservoirs and stimulate immune responses. Treatment of eight female, SIV-infected macaques on ART with four 2-weeks cycles of galunisertib leads to viral reactivation as indicated by plasma viral load and immunoPET/CT with a 64Cu-DOTA-F(ab')2-p7D3-probe. Post-galunisertib, lymph nodes, gut and PBMC exhibit lower cell-associated (CA-)SIV DNA and lower intact pro-virus (PBMC). Galunisertib does not lead to systemic increase in inflammatory cytokines. High-dimensional cytometry, bulk, and single-cell (sc)RNAseq reveal a galunisertib-driven shift toward an effector phenotype in T and NK cells characterized by a progressive downregulation in TCF1. In summary, we demonstrate that galunisertib, a clinical stage TGF-ß inhibitor, reverses SIV latency and decreases SIV reservoirs by driving T cells toward an effector phenotype, enhancing immune responses in vivo in absence of toxicity.
Asunto(s)
Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Femenino , Animales , Factor de Crecimiento Transformador beta , Replicación Viral , Leucocitos Mononucleares , Linfocitos T CD4-Positivos , Carga ViralRESUMEN
Fc-mediated antibody effector functions, such as antibody-dependent cellular cytotoxicity (ADCC), can contribute to the containment HIV-1 replication but whether such activities are sufficient for protection is unclear. We previously identified an antibody to the variable 2 (V2) apex of the HIV-1 Env trimer (PGT145) that potently directs the lysis of SIV-infected cells by NK cells but poorly neutralizes SIV infectivity. To determine if ADCC is sufficient for protection, separate groups of six rhesus macaques were treated with PGT145 or a control antibody (DEN3) by intravenous infusion followed five days later by intrarectal challenge with SIVmac239. Despite high concentrations of PGT145 and potent ADCC activity in plasma on the day of challenge, all animals became infected and viral loads did not differ between the PGT145- and DEN3-treated animals. To determine if PGT145 can protect against a neutralization-sensitive virus, two additional groups of six macaques were treated with PGT145 and DEN3 and challenged with an SIVmac239 variant with a single amino acid change in Env (K180S) that increases PGT145 binding and renders the virus susceptible to neutralization by this antibody. Although there was no difference in virus acquisition, peak and chronic phase viral loads were significantly lower and time to peak viremia was significantly delayed in the PGT145-treated animals compared to the DEN3-treated control animals. Env changes were also selected in the PGT145-treated animals that confer resistance to both neutralization and ADCC. These results show that ADCC is not sufficient for protection by this V2-specific antibody. However, protection may be achieved by increasing the affinity of antibody binding to Env above the threshold required for neutralization.
Asunto(s)
Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Animales , Macaca mulatta , Anticuerpos Antivirales , Citotoxicidad Celular Dependiente de AnticuerposRESUMEN
The rebound-competent viral reservoir, composed of a virus that is able to persist during antiretroviral therapy (ART) and mediate reactivation of systemic viral replication and rebound viremia after ART interruption (ATI), remains the biggest obstacle to treating HIV infection. A better understanding of the cellular and tissue origins and the dynamics of viral populations that initiate rebound upon ATI could help develop therapeutic strategies for reducing the rebound-competent viral reservoir. In this study, barcoded simian immunodeficiency virus (SIV), SIVmac239M, was used to infect rhesus macaques to enable monitoring of viral barcode clonotypes contributing to virus detectable in plasma after ATI. Blood and tissues from secondary lymphoid organs (spleen, mesenteric lymph nodes, and inguinal lymph nodes) and from the colon, ileum, lung, liver, and brain were analyzed using viral barcode sequencing, intact proviral DNA assay, single-cell RNA sequencing, and combined CODEX and RNAscope in situ hybridization. Four of seven animals had viral barcodes detectable by deep sequencing of plasma at necropsy, although plasma viral RNA remained below 22 copies per milliliter. Among the tissues studied, mesenteric lymph nodes, inguinal lymph nodes, and spleen contained viral barcodes detected in plasma. CD4+ T cells were the main cell type harboring viral RNA after ATI. Furthermore, T cell zones in lymphoid tissues showed higher viral RNA abundance than B cell zones for most animals. These findings are consistent with lymphoid tissues contributing to the virus present in plasma early after ATI.
Asunto(s)
Infecciones por VIH , Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Animales , Virus de la Inmunodeficiencia de los Simios/genética , Macaca mulatta , Infecciones por VIH/tratamiento farmacológico , Antirretrovirales/uso terapéutico , Antirretrovirales/farmacología , Tejido Linfoide , Replicación Viral , ARN Viral , Carga Viral , Linfocitos T CD4-PositivosRESUMEN
HIV rapidly rebounds after interruption of antiretroviral therapy (ART). HIV-specific CD8+ T cells may act to prevent early events in viral reactivation. However, the presence of viral immune escape mutations may limit the effect of CD8+ T cells on viral rebound. Here, we studied the impact of CD8 immune pressure on post-treatment rebound of barcoded SIVmac293M in 14 Mamu-A*01 positive rhesus macaques that initiated ART on day 14, and subsequently underwent two analytic treatment interruptions (ATIs). Rebound following the first ATI (seven months after ART initiation) was dominated by virus that retained the wild-type sequence at the Mamu-A*01 restricted Tat-SL8 epitope. By the end of the two-month treatment interruption, the replicating virus was predominantly escaped at the Tat-SL8 epitope. Animals reinitiated ART for 3 months prior to a second treatment interruption. Time-to-rebound and viral reactivation rate were significantly slower during the second treatment interruption compared to the first. Tat-SL8 escape mutants dominated early rebound during the second treatment interruption, despite the dominance of wild-type virus in the proviral reservoir. Furthermore, the escape mutations detected early in the second treatment interruption were well predicted by those replicating at the end of the first, indicating that escape mutant virus in the second interruption originated from the latent reservoir as opposed to evolving de novo post rebound. SL8-specific CD8+ T cell levels in blood prior to the second interruption were marginally, but significantly, higher (median 0.73% vs 0.60%, p = 0.016). CD8+ T cell depletion approximately 95 days after the second treatment interruption led to the reappearance of wild-type virus. This work suggests that CD8+ T cells can actively suppress the rebound of wild-type virus, leading to the dominance of escape mutant virus after treatment interruption.
Asunto(s)
Infecciones por VIH , Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Animales , Macaca mulatta , Replicación Viral/fisiología , Linfocitos T CD8-positivos , Epítopos , Carga Viral , Antirretrovirales/uso terapéutico , Antirretrovirales/farmacologíaRESUMEN
HIV-1 persistence during ART is due to the establishment of long-lived viral reservoirs in resting immune cells. Using an NHP model of barcoded SIVmac239 intravenous infection and therapeutic dosing of the anti-TGFBR1 inhibitor galunisertib (LY2157299), we confirmed the latency reversal properties of in vivo TGF-ß blockade, decreased viral reservoirs and stimulated immune responses. Eight SIV-infected macaques on suppressive ART were treated with 4 2-week cycles of galunisertib. ART was discontinued 3 weeks after the last dose, and macaques euthanized 6 weeks after ART-interruption(ATI). One macaque did not rebound, while the remaining rebounded between week 2 and 6 post-ATI. Galunisertib led to viral reactivation as indicated by plasma viral load and immunoPET/CT with the 64Cu-DOTA-F(ab')2-p7D3-probe. Half to 1 Log decrease in cell-associated (CA-)SIV DNA was detected in lymph nodes, gut and PBMC, while intact pro-virus in PBMC decreased by 3-fold. No systemic increase in inflammatory cytokines was observed. High-dimensions cytometry, bulk and single-cell RNAseq revealed a shift toward an effector phenotype in T and NK cells. In summary, we demonstrated that galunisertib, a clinical stage TGFß inhibitor, reverses SIV latency and decreases SIV reservoirs by driving T cells toward an effector phenotype, enhancing immune responses in vivo in absence of toxicity.
RESUMEN
One approach to 'functional cure' of HIV infection is to induce durable control of HIV replication after the interruption of antiretroviral therapy (ART). However, the major factors that determine the viral 'setpoint' level after treatment interruption are not well understood. Here we combine data on ART interruption following SIV infection for 124 total animals from 10 independent studies across 3 institutional cohorts to understand the dynamics and predictors of post-treatment viral control. We find that the timing of treatment initiation is an important determinant of both the peak and early setpoint viral levels after treatment interruption. During the first 3 weeks of infection, every day of delay in treatment initiation is associated with a 0.22 log10 copies/ml decrease in post-rebound peak and setpoint viral levels. However, delay in initiation of ART beyond 3 weeks of infection is associated with higher post-rebound setpoint viral levels. For animals treated beyond 3 weeks post-infection, viral load at ART initiation was the primary predictor of post-rebound setpoint viral levels. Potential alternative predictors of post-rebound setpoint viral loads including cell-associated DNA or RNA, time from treatment interruption to rebound, and pre-interruption CD8+ T cell responses were also examined in the studies where these data were available. This analysis suggests that optimal timing of treatment initiation may be an important determinant of post-treatment control of HIV.
Asunto(s)
Infecciones por VIH , Animales , Infecciones por VIH/tratamiento farmacológico , Linfocitos T CD8-positivos , ARN Viral , Carga Viral , Antirretrovirales/farmacología , Antirretrovirales/uso terapéuticoRESUMEN
Sustainable HIV remission after antiretroviral therapy (ART) withdrawal, or post-treatment control (PTC), remains a top priority for HIV treatment. We observed surprising PTC in an MHC-haplomatched cohort of MHC-M3+ SIVmac239+ Mauritian cynomolgus macaques (MCMs) initiated on ART at two weeks post-infection (wpi). None of the MCMs possessed MHC haplotypes previously associated with SIV control. For six months after ART withdrawal, we observed undetectable or transient viremia in seven of the eight MCMs, despite detecting replication competent SIV using quantitative viral outgrowth assays. In vivo depletion of CD8α+ cells induced rebound in all animals, indicating the observed PTC was mediated, at least in part, by CD8α+ cells. With intact proviral DNA assays, we found that MCMs had significantly smaller viral reservoirs two wpi than a cohort of identically infected rhesus macaques, a population that rarely develops PTC. We found a similarly small viral reservoir among six additional SIV+ MCMs in which ART was initiated at eight wpi, some of whom exhibited viral rebound. These results suggest that an unusually small viral reservoir is a hallmark among SIV+ MCMs. By evaluating immunological differences between MCMs that did and did not rebound, we identified that PTC was associated with a reduced frequency of CD4+ and CD8+ lymphocyte subsets expressing exhaustion markers. Together, these results suggest a combination of small reservoirs and immune-mediated virus suppression contribute to PTC in MCMs. Further, defining the immunologic mechanisms that engender PTC in this model may identify therapeutic targets for inducing durable HIV remission in humans.
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Infecciones por VIH , Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Humanos , Animales , Macaca mulatta , Linfocitos T CD8-positivos , Infecciones por VIH/tratamiento farmacológico , Macaca fascicularis , Carga Viral , Replicación Viral , Antirretrovirales/uso terapéutico , Antirretrovirales/farmacologíaRESUMEN
The rebound-competent viral reservoir (RCVR), comprised of virus that is able to persist during antiretroviral therapy (ART) and mediate reactivation of systemic viral replication and rebound viremia after antiretroviral therapy interruption (ATI), remains the biggest obstacle to the eradication of HIV infection. A better understanding of the cellular and tissue origins and the dynamics of viral populations that initiate rebound upon ATI could help develop targeted therapeutic strategies for reducing the RCVR. In this study, barcoded SIVmac239M was used to infect rhesus macaques to enable monitoring of viral barcode clonotypes contributing to virus detectable in plasma after ATI. Blood, lymphoid tissues (LTs, spleen, mesenteric and inguinal lymph nodes), and non-lymphoid tissues (NLTs, colon, ileum, lung, liver, and brain) were analyzed using viral barcode sequencing, intact proviral DNA assay, single-cell RNA sequencing, and combined CODEX/RNAscope/ in situ hybridization. Four of seven animals had viral barcodes detectable by deep sequencing of plasma at necropsy although plasma viral RNA remained < 22 copies/mL. Among the tissues studied, mesenteric and inguinal lymph nodes, and spleen contained viral barcodes detected in plasma, and trended to have higher cell-associated viral loads, higher intact provirus levels, and greater diversity of viral barcodes. CD4+ T cells were the main cell type harboring viral RNA (vRNA) after ATI. Further, T cell zones in LTs showed higher vRNA levels than B cell zones for most animals. These findings are consistent with LTs contributing to virus present in plasma early after ATI. One Sentence Summary: The reemerging of SIV clonotypes at early post-ATI are likely from the secondary lymphoid tissues.
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Transmitted/founder (TF) simian-human immunodeficiency viruses (SHIVs) express HIV-1 envelopes modified at position 375 to efficiently infect rhesus macaques while preserving authentic HIV-1 Env biology. SHIV.C.CH505 is an extensively characterized virus encoding the TF HIV-1 Env CH505 mutated at position 375 shown to recapitulate key features of HIV-1 immunobiology, including CCR5-tropism, a tier 2 neutralization profile, reproducible early viral kinetics, and authentic immune responses. SHIV.C.CH505 is used frequently in nonhuman primate studies of HIV, but viral loads after months of infection are variable and typically lower than those in people living with HIV. We hypothesized that additional mutations besides Δ375 might further enhance virus fitness without compromising essential components of CH505 Env biology. From sequence analysis of SHIV.C.CH505-infected macaques across multiple experiments, we identified a signature of envelope mutations associated with higher viremia. We then used short-term in vivo mutational selection and competition to identify a minimally adapted SHIV.C.CH505 with just five amino acid changes that substantially improve virus replication fitness in macaques. Next, we validated the performance of the adapted SHIV in vitro and in vivo and identified the mechanistic contributions of selected mutations. In vitro, the adapted SHIV shows improved virus entry, enhanced replication on primary rhesus cells, and preserved neutralization profiles. In vivo, the minimally adapted virus rapidly outcompetes the parental SHIV with an estimated growth advantage of 0.14 days-1 and persists through suppressive antiretroviral therapy to rebound at treatment interruption. Here, we report the successful generation of a well-characterized, minimally adapted virus, termed SHIV.C.CH505.v2, with enhanced replication fitness and preserved native Env properties that can serve as a new reagent for NHP studies of HIV-1 transmission, pathogenesis, and cure.
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Infecciones por VIH , Seropositividad para VIH , VIH-1 , Síndrome de Inmunodeficiencia Adquirida del Simio , Virus de la Inmunodeficiencia de los Simios , Animales , Humanos , Macaca mulatta/metabolismo , Productos del Gen env del Virus de la Inmunodeficiencia Humana , Replicación Viral/fisiologíaRESUMEN
[This corrects the article DOI: 10.1371/journal.pone.0092830.].
RESUMEN
Reservoirs of HIV maintained in anatomic compartments during antiretroviral therapy prevent HIV eradication. However, mechanisms driving their persistence and interventions to control them remain elusive. Here we report the presence of an inducible HIV reservoir within antigen-specific CD4+T cells in the central nervous system of a 59-year-old male with progressive multifocal leukoencephalopathy immune reconstitution inflammatory syndrome (PML-IRIS). HIV production during PML-IRIS was suppressed by modulating inflammation with corticosteroids; selection of HIV drug resistance caused subsequent breakthrough viremia. Therefore, inflammation can influence the composition, distribution and induction of HIV reservoirs, warranting it as a key consideration for developing effective HIV remission strategies.
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Infecciones por VIH , Síndrome Inflamatorio de Reconstitución Inmune , Leucoencefalopatía Multifocal Progresiva , Masculino , Humanos , Persona de Mediana Edad , Síndrome Inflamatorio de Reconstitución Inmune/tratamiento farmacológico , Síndrome Inflamatorio de Reconstitución Inmune/etiología , Leucoencefalopatía Multifocal Progresiva/tratamiento farmacológico , Leucoencefalopatía Multifocal Progresiva/etiología , Encéfalo , Sistema Nervioso CentralRESUMEN
Since the first HIV-cured person was reported in 2009, a strong interest in developing highly sensitive HIV and SIV reservoir assays has emerged. In particular, the question arose about the comparative value of state-of-the-art assays to measure and characterize the HIV reservoir, and how these assays can be applied to accurately detect changes in the reservoir during efforts to develop a cure for HIV infection. Second, it is important to consider the impact on the outcome of clinical trials if these relatively new HIV reservoir assays are incorporated into clinical trial endpoints and/or used for clinical decision-making. To understand the advantages and limitations and the regulatory implications of HIV reservoir assays, the National Institute of Allergy and Infectious Diseases (NIAID) sponsored and convened a meeting on September 16, 2022, to discuss the state of knowledge concerning these questions and best practices for selecting HIV reservoir assays for a particular research question or clinical trial protocol.
RESUMEN
[This corrects the article DOI: 10.1371/journal.ppat.1009565.].