The CARD8 inflammasome dictates HIV/SIV pathogenesis and disease progression.

While CD4+ T cell depletion is key to disease progression in people living with HIV and SIV-infected macaques, the mechanisms underlying this depletion remain incompletely understood, with most cell death involving uninfected cells. In contrast, SIV infection of "natural" hosts such as sooty mangabeys does not cause CD4+ depletion and AIDS despite high-level viremia. Here, we report that the CARD8 inflammasome is activated immediately after HIV entry by the viral protease encapsulated in incoming virions. Sensing of HIV protease activity by CARD8 leads to rapid pyroptosis of quiescent cells without productive infection, while T cell activation abolishes CARD8 function and increases permissiveness to infection. In humanized mice reconstituted with CARD8-deficient cells, CD4+ depletion is delayed despite high viremia. Finally, we discovered loss-of-function mutations in CARD8 from "natural hosts," which may explain the peculiarly non-pathogenic nature of these infections. Our study suggests that CARD8 drives CD4+ T cell depletion during pathogenic HIV/SIV infections.

Exploiting Mucosal Immunity for Antiviral Vaccines.

Mucosal surfaces provide a remarkably effective barrier against potentially dangerous pathogens. Therefore, enhancing mucosal immunity through vaccines-strengthening that first line of defense-holds significant promise for reducing the burden of viral diseases. The large and varied class of viral pathogens, however, continues to present thorny challenges to vaccine development. Two primary difficulties exist: Viruses exhibit a stunning diversity of strategies for evading the host immune response, and even when we understand the nature of effective immune protection against a given virus, eliciting that protection is technically challenging. Only a few mucosal vaccines have surmounted these obstacles thus far. Recent developments, however, could greatly improve vaccine design. In this review, we first sketch out our understanding of mucosal immunity and then compare the herpes simplex virus, human immunodeficiency virus, and influenza virus to illustrate the distinct challenges of developing successful vaccines and to outline potential solutions.

Broadly Neutralizing Antibodies to HIV and Their Role in Vaccine Design.

HIV employs multiple means to evade the humoral immune response, particularly the elicitation of and recognition by broadly neutralizing antibodies (bnAbs). Such antibodies can act antivirally against a wide spectrum of viruses by targeting relatively conserved regions on the surface HIV envelope trimer spike. Elicitation of and recognition by bnAbs are hindered by the arrangement of spikes on virions and the relatively difficult access to bnAb epitopes on spikes, including the proximity of variable regions and a high density of glycans. Yet, in a small proportion of HIV-infected individuals, potent bnAb responses do develop, and isolation of the corresponding monoclonal antibodies has been facilitated by identification of favorable donors with potent bnAb sera and by development of improved methods for human antibody generation. Molecular studies of recombinant Env trimers, alone and in interaction with bnAbs, are providing new insights that are fueling the development and testing of promising immunogens aimed at the elicitation of bnAbs.

The freedom to grow.

David Baltimore's numerous major discoveries, academic leadership roles, science advocacy work, and knack for training successful scientists have shaped the direction of biomedical science for more than 50 years. Cell editor Nicole Neuman caught up with David, the recipient of the 2021 Lasker∼Koshland Special Achievement Award in Medical Science, to discuss his scientific origin story, his philosophy on mentorship, how he rose to the challenge of founding one of the premier biomedical research institutes in the world, and some of his crucial moments shaping modern scientific policy. Excerpts from this conversation, edited for clarity, are presented below, and the full conversation is available with the article online.

Fc Characteristics Mediate Selective Placental Transfer of IgG in HIV-Infected Women.

The placental transfer of maternal IgG is critical for infant protection against infectious pathogens. However, factors that modulate the placental transfer of IgG remain largely undefined. HIV-infected women have impaired placental IgG transfer, presenting a unique "disruption model" to define factors that modulate placental IgG transfer. We measured the placental transfer efficiency of maternal HIV and pathogen-specific IgG in US and Malawian HIV-infected mothers and their HIV-exposed uninfected and infected infants. We examined the role of maternal HIV disease progression, infant factors, placental Fc receptor expression, IgG subclass, and glycan signatures and their association with placental IgG transfer efficiency. Maternal IgG characteristics, such as binding to placentally expressed Fc receptors FcγRIIa and FcγRIIIa, and Fc region glycan profiles were associated with placental IgG transfer efficiency. Our findings suggest that Fc region characteristics modulate the selective placental transfer of IgG, with implications for maternal vaccine design and infant health.

An evolutionary role for HIV latency in enhancing viral transmission.

HIV latency is the chief obstacle to eradicating HIV but is widely believed to be an evolutionary accident providing no lentiviral fitness advantage. However, findings of latency being "hardwired" into HIV's gene-regulatory circuitry appear inconsistent with latency being an evolutionary accident, given HIV's rapid mutation rate. Here, we propose that latency is an evolutionary "bet-hedging" strategy whose frequency has been optimized to maximize lentiviral transmission by reducing viral extinction during mucosal infections. The model quantitatively fits the available patient data, matches observations of high-frequency latency establishment in cell culture and primates, and generates two counterintuitive but testable predictions. The first prediction is that conventional CD8-depletion experiments in SIV-infected macaques increase latent cells more than viremia. The second prediction is that strains engineered to have higher replicative fitness­via reduced latency­will exhibit lower infectivity in animal-model mucosal inoculations. Therapeutically, the theory predicts treatment approaches that may substantially enhance "activate-and-kill" HIV-cure strategies.

A hardwired HIV latency program.

Biological circuits can be controlled by two general schemes: environmental sensing or autonomous programs. For viruses such as HIV, the prevailing hypothesis is that latent infection is controlled by cellular state (i.e., environment), with latency simply an epiphenomenon of infected cells transitioning from an activated to resting state. However, we find that HIV expression persists despite the activated-to-resting cellular transition. Mathematical modeling indicates that HIV's Tat positive-feedback circuitry enables this persistence and strongly controls latency. To overcome the inherent crosstalk between viral circuitry and cellular activation and to directly test this hypothesis, we synthetically decouple viral dependence on cellular environment from viral transcription. These circuits enable control of viral transcription without cellular activation and show that Tat feedback is sufficient to regulate latency independent of cellular activation. Overall, synthetic reconstruction demonstrates that a largely autonomous, viral-encoded program underlies HIV latency­potentially explaining why cell-targeted latency-reversing agents exhibit incomplete penetrance.

The HIV capsid mimics karyopherin engagement of FG-nucleoporins.

HIV can infect non-dividing cells because the viral capsid can overcome the selective barrier of the nuclear pore complex and deliver the genome directly into the nucleus1,2. Remarkably, the intact HIV capsid is more than 1,000 times larger than the size limit prescribed by the diffusion barrier of the nuclear pore3. This barrier in the central channel of the nuclear pore is composed of intrinsically disordered nucleoporin domains enriched in phenylalanine-glycine (FG) dipeptides. Through multivalent FG interactions, cellular karyopherins and their bound cargoes solubilize in this phase to drive nucleocytoplasmic transport4. By performing an in vitro dissection of the nuclear pore complex, we show that a pocket on the surface of the HIV capsid similarly interacts with FG motifs from multiple nucleoporins and that this interaction licences capsids to penetrate FG-nucleoporin condensates. This karyopherin mimicry model addresses a key conceptual challenge for the role of the HIV capsid in nuclear entry and offers an explanation as to how an exogenous entity much larger than any known cellular cargo may be able to non-destructively breach the nuclear envelope.

Evolution and Diversity of Immune Responses during Acute HIV Infection.

Understanding the earliest immune responses following HIV infection is critical to inform future vaccines and therapeutics. Here, we review recent prospective human studies in at-risk populations that have provided insight into immune responses during acute infection, including additional relevant data from non-human primate (NHP) studies. We discuss the timing, nature, and function of the diverse immune responses induced, the onset of immune dysfunction, and the effects of early anti-retroviral therapy administration. Treatment at onset of viremia mitigates peripheral T and B cell dysfunction, limits seroconversion, and enhances cellular antiviral immunity despite persistence of infection in lymphoid tissues. We highlight pertinent areas for future investigation, and how application of high-throughput technologies, alongside targeted NHP studies, may elucidate immune response features to target in novel preventions and cures.

Tumor-necrosis factor impairs CD4(+) T cell-mediated immunological control in chronic viral infection.

Persistent viral infections are characterized by the simultaneous presence of chronic inflammation and T cell dysfunction. In prototypic models of chronicity--infection with human immunodeficiency virus (HIV) or lymphocytic choriomeningitis virus (LCMV)--we used transcriptome-based modeling to reveal that CD4(+) T cells were co-exposed not only to multiple inhibitory signals but also to tumor-necrosis factor (TNF). Blockade of TNF during chronic infection with LCMV abrogated the inhibitory gene-expression signature in CD4(+) T cells, including reduced expression of the inhibitory receptor PD-1, and reconstituted virus-specific immunity, which led to control of infection. Preventing signaling via the TNF receptor selectively in T cells sufficed to induce these effects. Targeted immunological interventions to disrupt the TNF-mediated link between chronic inflammation and T cell dysfunction might therefore lead to therapies to overcome persistent viral infection.

CXCR5(+) follicular cytotoxic T cells control viral infection in B cell follicles.

During unresolved infections, some viruses escape immunological control and establish a persistant reservoir in certain cell types, such as human immunodeficiency virus (HIV), which persists in follicular helper T cells (TFH cells), and Epstein-Barr virus (EBV), which persists in B cells. Here we identified a specialized group of cytotoxic T cells (TC cells) that expressed the chemokine receptor CXCR5, selectively entered B cell follicles and eradicated infected TFH cells and B cells. The differentiation of these cells, which we have called 'follicular cytotoxic T cells' (TFC cells), required the transcription factors Bcl6, E2A and TCF-1 but was inhibited by the transcriptional regulators Blimp1, Id2 and Id3. Blimp1 and E2A directly regulated Cxcr5 expression and, together with Bcl6 and TCF-1, formed a transcriptional circuit that guided TFC cell development. The identification of TFC cells has far-reaching implications for the development of strategies to control infections that target B cells and TFH cells and to treat B cell-derived malignancies.

Vaccine-Induced Protection from Homologous Tier 2 SHIV Challenge in Nonhuman Primates Depends on Serum-Neutralizing Antibody Titers.

Passive administration of HIV neutralizing antibodies (nAbs) can protect macaques from hard-to-neutralize (tier 2) chimeric simian-human immunodeficiency virus (SHIV) challenge. However, conditions for nAb-mediated protection after vaccination have not been established. Here, we selected groups of 6 rhesus macaques with either high or low serum nAb titers from a total of 78 animals immunized with recombinant native-like (SOSIP) Env trimers. Repeat intrarectal challenge with homologous tier 2 SHIVBG505 led to rapid infection in unimmunized and low-titer animals. High-titer animals, however, demonstrated protection that was gradually lost as nAb titers waned over time. An autologous serum ID50 nAb titer of ∼1:500 afforded more than 90% protection from medium-dose SHIV infection. In contrast, antibody-dependent cellular cytotoxicity and T cell activity did not correlate with protection. Therefore, Env protein-based vaccination strategies can protect against hard-to-neutralize SHIV challenge in rhesus macaques by inducing tier 2 nAbs, provided appropriate neutralizing titers can be reached and maintained.

Isolation and Structure of an Antibody that Fully Neutralizes Isolate SIVmac239 Reveals Functional Similarity of SIV and HIV Glycan Shields.

HIV- and SIV-envelope (Env) trimers are both extensively glycosylated, and antibodies identified to date have been unable to fully neutralize SIVmac239. Here, we report the isolation, structure, and glycan interactions of antibody ITS90.03, a monoclonal antibody that completely neutralized the highly neutralization-resistant isolate, SIVmac239. The co-crystal structure of a fully glycosylated SIVmac239-gp120 core in complex with rhesus CD4 and the antigen-binding fragment of ITS90.03 at 2.5-Å resolution revealed that ITS90 recognized an epitope comprised of 45% glycan. SIV-gp120 core, rhesus CD4, and their complex could each be aligned structurally to their human counterparts. The structure revealed that glycans masked most of the SIV Env protein surface, with ITS90 targeting a glycan hole, which is occupied in ∼83% of SIV strains by glycan N238. Overall, the SIV glycan shield appears to functionally resemble its HIV counterpart in coverage of spike, shielding from antibody, and modulation of receptor accessibility.

Pathogenesis of HIV-Related Lung Disease: Immunity, Infection, and Inflammation.

Despite anti-retroviral therapy (ART), human immunodeficiency virus-1 (HIV)-related pulmonary disease continues to be a major cause of morbidity and mortality for people living with HIV (PLWH). The spectrum of lung diseases has changed from acute opportunistic infections resulting in death to chronic lung diseases for those with access to ART. Chronic immune activation and suppression can result in impairment of innate immunity and progressive loss of T cell and B cell functionality with aberrant cytokine and chemokine responses systemically as well as in the lung. HIV can be detected in the lungs of PLWH and has profound effects on cellular immune functions. In addition, HIV-related lung injury and disease can occur secondary to a number of mechanisms including altered pulmonary and systemic inflammatory pathways, viral persistence in the lung, oxidative stress with additive effects of smoke exposure, microbial translocation, and alterations in the lung and gut microbiome. Although ART has had profound effects on systemic viral suppression in HIV, the impact of ART on lung immunology still needs to be fully elucidated. Understanding of the mechanisms by which HIV-related lung diseases continue to occur is critical to the development of new preventive and therapeutic strategies to improve lung health in PLWH.

HIV reservoirs as obstacles and opportunities for an HIV cure.

The persistence of HIV reservoirs remains a formidable obstacle to achieving sustained virologic remission in HIV-infected individuals after antiretroviral therapy (ART) is discontinued, even if plasma viremia has been successfully suppressed for prolonged periods of time. Numerous approaches aimed at eradicating the virus, as well as maintaining its prolonged suppression in the absence of ART, have had little success. A better understanding of the pathophysiologic nature of HIV reservoirs and the impact of various interventions on their persistence is essential for the development of successful therapeutic strategies against HIV or the long-term control of infection. Here, we discuss the persistent HIV reservoir as a barrier to cure as well as the current therapeutic strategies aimed at eliminating or controlling the virus in the absence of ART.

Adjunctive Dexamethasone for Tuberculous Meningitis in HIV-Positive Adults.

BACKGROUND: Adjunctive glucocorticoids are widely used to treat human immunodeficiency virus (HIV)-associated tuberculous meningitis despite limited data supporting their safety and efficacy. METHODS: We conducted a double-blind, randomized, placebo-controlled trial involving HIV-positive adults (≥18 years of age) with tuberculous meningitis in Vietnam and Indonesia. Participants were randomly assigned to receive a 6-to-8-week tapering course of either dexamethasone or placebo in addition to 12 months of antituberculosis chemotherapy. The primary end point was death from any cause during the 12 months after randomization. RESULTS: A total of 520 adults were randomly assigned to receive either dexamethasone (263 participants) or placebo (257 participants). The median age was 36 years; 255 of 520 participants (49.0%) had never received antiretroviral therapy, and 251 of 484 participants (51.9%) with available data had a baseline CD4 count of 50 cells per cubic millimeter or less. Six participants withdrew from the trial, and five were lost to follow-up. During the 12 months of follow-up, death occurred in 116 of 263 participants (44.1%) in the dexamethasone group and in 126 of 257 participants (49.0%) in the placebo group (hazard ratio, 0.85; 95% confidence interval, 0.66 to 1.10; P = 0.22). Prespecified analyses did not reveal a subgroup that clearly benefited from dexamethasone. The incidence of secondary end-point events, including cases of immune reconstitution inflammatory syndrome during the first 6 months, was similar in the two trial groups. The numbers of participants with at least one serious adverse event were similar in the dexamethasone group (192 of 263 participants [73.0%]) and the placebo group (194 of 257 participants [75.5%]) (P = 0.52). CONCLUSIONS: Among HIV-positive adults with tuberculous meningitis, adjunctive dexamethasone, as compared with placebo, did not confer a benefit with respect to survival or any secondary end point. (Funded by the Wellcome Trust; ACT HIV ClinicalTrials.gov number, NCT03092817.).

Massively multiplexed nucleic acid detection with Cas13.

The great majority of globally circulating pathogens go undetected, undermining patient care and hindering outbreak preparedness and response. To enable routine surveillance and comprehensive diagnostic applications, there is a need for detection technologies that can scale to test many samples1-3 while simultaneously testing for many pathogens4-6. Here, we develop Combinatorial Arrayed Reactions for Multiplexed Evaluation of Nucleic acids (CARMEN), a platform for scalable, multiplexed pathogen detection. In the CARMEN platform, nanolitre droplets containing CRISPR-based nucleic acid detection reagents7 self-organize in a microwell array8 to pair with droplets of amplified samples, testing each sample against each CRISPR RNA (crRNA) in replicate. The combination of CARMEN and Cas13 detection (CARMEN-Cas13) enables robust testing of more than 4,500 crRNA-target pairs on a single array. Using CARMEN-Cas13, we developed a multiplexed assay that simultaneously differentiates all 169 human-associated viruses with at least 10 published genome sequences and rapidly incorporated an additional crRNA to detect the causative agent of the 2020 COVID-19 pandemic. CARMEN-Cas13 further enables comprehensive subtyping of influenza A strains and multiplexed identification of dozens of HIV drug-resistance mutations. The intrinsic multiplexing and throughput capabilities of CARMEN make it practical to scale, as miniaturization decreases reagent cost per test by more than 300-fold. Scalable, highly multiplexed CRISPR-based nucleic acid detection shifts diagnostic and surveillance efforts from targeted testing of high-priority samples to comprehensive testing of large sample sets, greatly benefiting patients and public health9-11.

An Mtb-Human Protein-Protein Interaction Map Identifies a Switch between Host Antiviral and Antibacterial Responses.

Although macrophages are armed with potent antibacterial functions, Mycobacterium tuberculosis (Mtb) replicates inside these innate immune cells. Determinants of macrophage intrinsic bacterial control, and the Mtb strategies to overcome them, are poorly understood. To further study these processes, we used an affinity tag purification mass spectrometry (AP-MS) approach to identify 187 Mtb-human protein-protein interactions (PPIs) involving 34 secreted Mtb proteins. This interaction map revealed two factors involved in Mtb pathogenesis-the secreted Mtb protein, LpqN, and its binding partner, the human ubiquitin ligase CBL. We discovered that an lpqN Mtb mutant is attenuated in macrophages, but growth is restored when CBL is removed. Conversely, Cbl-/- macrophages are resistant to viral infection, indicating that CBL regulates cell-intrinsic polarization between antibacterial and antiviral immunity. Collectively, these findings illustrate the utility of this Mtb-human PPI map for developing a deeper understanding of the intricate interactions between Mtb and its host.

Overcoming barriers and stigma: new frontiers in solid organ transplantation for people with HIV.

There is a growing need for solid organ transplantation (SOT) for people living with human immunodeficiency virus (HIV). With the advent of antiretroviral therapy, people living with HIV are experiencing increased life expectancies and are, therefore, developing more comorbidities, including end-stage organ disease. In cases of advanced organ failure, SOT is often the best therapeutic option to improve quality of life and overall survival. As organ shortages persist, transplantation of organs from donors with HIV to recipients with HIV has become a potential therapeutic option. This article first reviews the current state of organ transplantation from donors without HIV to recipients with HIV (HIV D-/R+) by organ and discusses key lessons learned from these transplant trials, including those about drug-drug interactions, rejection, and opportunistic infections. It then explores transplantation from donors with HIV to recipients with HIV (HIV D+/R+), a new frontier. Finally, it investigates challenges of implementation, including public awareness and regulatory requirements, and explores future directions for SOT in people living with HIV.

SHIV-C109p5 NHP induces rapid disease progression in elderly macaques with extensive GI viral replication.

CCR5-tropic simian/human immunodeficiency viruses (SHIV) with clade C transmitted/founder envelopes represent a critical tool for the investigation of HIV experimental vaccines and microbicides in nonhuman primates, although many such isolates lead to spontaneous viral control post infection. Here, we generated a high-titer stock of pathogenic SHIV-C109p5 by serial passage in two rhesus macaques (RM) and tested its virulence in aged monkeys. The co-receptor usage was confirmed before infecting five geriatric rhesus macaques (four female and one male). Plasma viral loads were monitored by reverse transcriptase-quantitative PCR (RT-qPCR), cytokines by multiplex analysis, and biomarkers of gastrointestinal damage by enzyme-linked immunosorbent assay. Antibodies and cell-mediated responses were also measured. Viral dissemination into tissues was determined by RNAscope. Intravenous SHIV-C109p5 infection of aged RMs leads to high plasma viremia and rapid disease progression; rapid decrease in CD4+ T cells, CD4+CD8+ T cells, and plasmacytoid dendritic cells; and wasting necessitating euthanasia between 3 and 12 weeks post infection. Virus-specific cellular immune responses were detected only in the two monkeys that survived 4 weeks post infection. These were Gag-specific TNFα+CD8+, MIP1ß+CD4+, Env-specific IFN-γ+CD4+, and CD107a+ T cell responses. Four out of five monkeys had elevated intestinal fatty acid binding protein levels at the viral peak, while regenerating islet-derived protein 3α showed marked increases at later time points in the three animals surviving the longest, suggesting gut antimicrobial peptide production in response to microbial translocation post infection. Plasma levels of monocyte chemoattractant protein-1, interleukin-15, and interleukin-12/23 were also elevated. Viral replication in gut and secondary lymphoid tissues was extensive.IMPORTANCESimian/human immunodeficiency viruses (SHIV) are important reagents to study prevention of virus acquisition in nonhuman primate models of HIV infection, especially those representing transmitted/founder (T/F) viruses. However, many R5-tropic SHIV have limited fitness in vivo leading to many monkeys spontaneously controlling the virus post acute infection. Here, we report the generation of a pathogenic SHIV clade C T/F stock by in vivo passage leading to sustained viral load set points, a necessity to study pathogenicity. Unexpectedly, administration of this SHIV to elderly rhesus macaques led to extensive viral replication and fast disease progression, despite maintenance of a strict R5 tropism. Such age-dependent rapid disease progression had previously been reported for simian immunodeficiency virus but not for R5-tropic SHIV infections.