PEDIATRIC ADOLESCENT VIRUS ELIMINATION (PAVE) MARTIN DELANEY COLLABORATORY (UM1 AI164566)

The immediate establishment of the latent HIV-1 reservoir in resting memory CD4+ T cells precludes HIV-1 cure, compelling ART for a lifetime in children. The mission of the PAVE Collaboratory is to use cutting-edge science to establish a deep and broad understanding of the immunopathogenesis of pediatric HIV-1 reservoirs, across the age spectrum, and to demonstrate preclinical safety and efficacy of novel therapeutics to eradicate reservoirs and control rebound that will pave the way for future interventional human studies toward a lifetime of sustained HIV-1 control off ART. We hypothesize that the unique features of the infant immune system at the time of reservoir establishment impact the characteristics of long-term virus persistence, susceptibility to immune- mediated clearance, and reactivation that are distinct from adult infections, warranting in-depth investigation to inform cure therapeutics suitable for children. We will test this hypothesis and execute the PAVE Scientific Agenda through accomplishment of the following Specific Aims: 1. Define the establishment and evolution of the HIV latent reservoir in perinatal infection. 2. Enhance pediatric immunity and broadly neutralizing antibody (bNAb) delivery to achieve post-treatment control of HIV-1 off ART. 3. Deploy immune-targeted strategies to eliminate virus reservoirs. 4. Optimize virologic, immunologic, and imaging methods to assess efficacy of HIV-1/S(H)IV cure interventions. 5. Foster community engagement in pediatric HIV cure research. The PAVE program is multidisciplinary, multicultural, and iterative with a nimble structure encompassed by four highly synergistic Research Foci and a domestic and international community program that will rapidly incorporate new scientific directions and feedback from our stakeholders. The PAVE leadership team spans diverse scientific expertise and exhibits additional diversity in terms of gender, academic rank, and country of origin. Each of the Research Foci also includes junior faculty co-Investigators to facilitate their career development within the HIV-1 research space. Through the collective efforts of our scientific leadership, Executive Committee, Scientific Advisory Board, investigators, industry partners, network collaborations, and domestic and international community program, PAVE anticipates meeting the following overall milestones of: 1) understanding early life immunity and early antiretroviral treatment on the composition and stability of the latent reservoir, including in naïve T cells, and potential for HIV-1 remission; 2) eliminating of these reservoirs in pre-clinical studies of immune-targeted strategies; 3) defining the role of myeloid cells in HIV-1 persistence and rebound, including in the CNS; 4) establishing novel approaches to enhance pediatric immunity through active and passive immunization; 5) developing cutting-edge approaches to quantify and monitor proviral reservoirs to measure clinical trial efficacy, and 6) promoting active community engagement in pediatric HIV-1 cure research. These milestones will help achieve the vision of sustained ART-free control of HIV-1 replication in pediatric populations.

IMMUNE DETERMINANTS OF PEDIATRIC HIV/SIV RESERVOIR ESTABLISHMENT AND MAINTENANCE (P01 HD112217)

New perinatal HIV infections continue, at a rate of about one every 3-4 minutes, 400 every day, and 3000 every week. For children living with HIV (CLWH), antiretroviral therapy (ART) greatly reduces mortality and morbidity; however, viral rebound quickly ensues if ART is stopped. HIV persistence in a latent reservoir in CD4+ T cells that can give rise to rebound viremia is the major barrier to a cure. However, there remains a significant knowledge gap regarding how the establishment and maintenance of HIV reservoirs are regulated by the neonatal and childhood immune system. The overall objective of this Program project application is to generate a comprehensive understanding of the complex host-pathogen interactions critical for HIV reservoir seeding and persistence such that novel cure strategies truly targeted for the unique immune environment of CLWH can be created. Our central hypothesis is that the host immune environment during the perinatal period and throughout childhood and adolescence, influenced by host/microbial metabolites and thymic output, determines the efficacy of HIV-specific immunity and the nature of reservoir establishment and maintenance. To test this hypothesis, we have compiled an outstanding team of experienced scientists and early-stage investigators (ESIs) from the Departments of Pediatrics and Pathology at Emory School of Medicine, with collaborators from the NIH, Kirby Institute, and the Université de Montréal. The research proposed builds upon the strong existing collaborations that generated the rigorous foundational research in support of each Project and develops links between new collaborators. Program oversight, communication plans, and mentoring of ESIs will be coordinated through the Administrative Core. We propose multiomic experiments in Projects 1-3 with data centralization in the Bioinformatics Core. The overall Specific Aims of our Program are: 1) To investigate the impact of IL-10, TGF-β, and thymic output on immune function and reservoir establishment and maintenance in perinatal HIV/SIV infection; 2) To determine the role of CD8+ T cells in reservoir establishment and maintenance in perinatal HIV/SIV infection; 3) To generate integrative models that define the host immune determinants of HIV/SIV reservoir establishment and maintenance and predict the size and nature of the HIV reservoir in CLWH. This deep dive into pediatric immunology will allow for insights to emerge that would not be possible if each Project were performed in isolation. A better understanding of the vulnerability of HIV reservoirs to innate and adaptive immune pressure will drive informed approaches to a cure for CLWH. The research proposed builds on our team’s expertise, the rich resources available at Emory School of Medicine, Emory National Primate Research Center, Emory Center for AIDS Research, and generous access to biospecimens from two cohorts of CLWH. We are confident that this Program will lead to important discoveries regarding immune regulation of the pediatric HIV reservoir and immune dysfunction. It is our mission to turn these discoveries into clinical trial protocols to advance research towards a cure for children with HIV.

ENHANCED LATENCY REVERSAL AND RESERVOIR CLEARANCE IN MACAQUES (R37 AI157862)

Current treatment for HIV infection requires strict adherence to daily antiretroviral therapy (ART). ART has greatly reduced mortality and morbidity but is not a cure. The main barrier to a cure is the pool of latently infected CD4+ T cells that carry integrated HIV provirus and are capable of reactivating and causing recrudescence of viremia if ART is stopped. A leading strategy to eliminate this persistent reservoir is termed “shock and kill” or “kick and kill” and consists of two interventions used with continued ART: first, a latency reversal agent (LRA) to reactivate latent virus and second, an approach to enhance clearance of infected cells. In this proposal, we will build upon our preliminary data indicating that activation of the non-canonical (nc) NF- κB signaling pathway with a mimetic of the second mitochondrial-derived activator of caspases (SMACm) results in systemic latency reversal in animal models. Using the highly relevant SIV/RM/ART model, we will answer three important questions regarding the biologic effects of SMACm treatment. First (Aim 1), we will determine the anatomic and cellular origin of the viremia produced during ART upon treatment with SMACm and its relationship with rebound viremia after ART interruption. In this study, we will utilize barcoded SIVmac239M to extensively sequence SIV RNA in sorted CD4+ T cell subsets from blood and tissues in comparison to plasma virus. We will also apply single cell transcriptomic methodologies to answer this question. Second (Aim 2), we test a novel “kick and kill” approach that combines latency reversal with SMACm plus venetoclax, a selective BCL-2 inhibitor, that has been shown to cause preferential apoptosis of HIV- expressing cells. We will determine whether this combined treatment reduces total, intact, and replication competent reservoirs in memory CD4+ T cells from multiple tissues and/or modulates viral recrudescence after ART interruption. Third (Aim 3), we will model the immunovirologic factors predictive of virus reactivation on ART with SMACm treatment and during ART interruption following therapeutic targeting of the ncNF-κB and BCL-2 pathways. This model will further reveal the potential of this cure approach and inform the design of future “kick and kill” studies. This work will allow us to elucidate the full potential of targeting the ncNF-κB pathway for latency reversal and to evaluate an innovative strategy to reduce virus persistence. We hope these results will contribute to a cure for HIV infection.

NEURODEVELOPMENT AFTER POSTNATAL ZIKA VIRUS INFECTION IN INFANT MACAQUES (R01 NS120182)

Intense research has focused on the neurologic effects of prenatal Zika virus (ZIKV) infection; however, the consequences of postnatal infection early in life are understudied. Infants exposed to ZIKV in utero but born without microcephaly can develop postnatal microcephaly, neurologic dysfunction, and neurodevelopmental ab- normalities, highlighting the potential of ZIKV to cause ongoing damage after birth. This damage is likely related to the exponential maturation of the brain that occurs during the first 2 years of life, particularly in temporal, prefrontal and parietal regions important for emotional, social and executive functions, including learning, atten- tion and memory. Our group has previously reported that postnatal ZIKV infection causes abnormalities in brain structure, function, and behavior in a pilot study of infant rhesus macaques (RMs) infected postnatally. Here, we propose to extend the scope and duration our prior pilot study and generate novel data regarding the impact of postnatal ZIKV infection on the developing brain. The Objective of this application is to bring new mechanistic insights into postnatal ZIKV infection to address the existing knowledge gap regarding outcomes and host-virus interactions. We will use our postnatal ZIKV-RM model to interrogate the neurobehavioral impact of ZIKV infection at different stages of postnatal brain develop- ment, with approaches that span from single cells to whole animal. This model of postnatal ZIKV exposure allows us to generate key data on the mechanisms by which ZIKV and/or the immune response to infection leads to cellular changes that ultimately result in aberrant postnatal development of limbic structures and behavioral def- icits later in life. We hypothesize that 1) ZIKV and/or the immune response to infection disproportionately affects limbic structures in the postnatally developing brain; 2) cellular changes in these limbic structures lead to aberrant neurodevelopment and abnormal behaviors; and 3) there may be a period of vulnerability to ZIKV during post- natal brain development. We will test our hypotheses in these Specific Aims: 1) Determine the spectrum of abnormal behavior and cognition following ZIKV infection of infant RMs at different stages of brain development; 2) Identify developmental trajectories of brain structure and function following postnatal ZIKV infection of infant RMs at different stages of brain development; and 3) Define the neurodevelopmental pathways and cell types impacted by postnatal ZIKV infection. This work will include RM infants infected with ZIKV at 1 or 6 months of age (equivalent to 4 and 24 month old humans) as well as age- and rearing-matched and viral mimic controls, that over their first 2 years of life will undergo a series of detailed assessments including validated tests of socioemotional behavior and cognition, structural and functional brain imaging, brain histology, stereology and single cell and bulk cell transcriptomics. Our results may have important public health implications for children living in ZIKV-endemic/epidemic areas as well as for travelers to these regions.