Basic Biomedical Sciences Research For Early Stage Investigators (ESI)

Principal Investigator:  Crystal Wang, PhD, UC San Diego

Budget:  $269,056

Start Date: March 1, 2026        End Date: February 29, 2028

Project Abstract: Despite viral suppression, up to 50% of people with HIV (PWH) continue to suffer from neurocognitive impairment (NCI), which is a strong predictor of functional disability. Dietary interventions represent a promising therapeutic approach, with established efficacy for improving neurocognition in conditions such as Alzheimer's disease. These interventions likely exert their beneficial effects by reducing inflammation, the primary driver of elevated NCI rates in PWH. Notably, no studies have systematically investigated the specific pro- or anti-inflammatory effects of diet in PWH. The current study will derive unbiased dietary readouts from previously collected fecal samples of 331 PWH enrolled in cohort studies at the HIV Neurobehavioral Research Program to identify pro- or anti-inflammatory foods and assess their associations with NCI. We will employ cutting-edge foodomics methodology that integrates supervised machine-learning techniques to systematically identify over 6,000 food biomarkers in fecal metabolomics data. Our findings will inform the development of a targeted anti-inflammatory dietary intervention for HIV-associated NCI.

Principal Investigator:  Joseph Happer, PhD, UC San Diego

Budget:  $269,944

Start Date: March 1, 2026        End Date: February 29, 2028

Project Abstract: Substance use is known to increase the frequency of high-risk behaviors associated with HIV exposure and transmission. In California, methamphetamine (METH) use is one of the most commonly used illicit substances among people with HIV (PWH) and can lead to faster disease progression and immune system dysfunction despite viral suppression. Both METH use and HIV independently exert deleterious effects, particularly on overlapping frontostriatal regions involved with decision-making like the basal ganglia (BG), yet their combined effects are poorly understood. Our own work suggests BG activation during reward-based decision-making is altered in PWH and a history of METH dependence. Emerging evidence suggests that dopamine (DA) signaling may be a critical mechanistic link. METH primarily targets the DA system and increases DA levels, inducing neurotoxicity, possibly through neuroinflammatory mechanisms. Both viral proteins and HIV-associated inflammation appear to preferentially affect DA-rich frontostriatal regions including the BG, contributing to an overall downregulation of DAergic functioning. Preclinical studies suggest that METH-induced DA dysfunction may potentiate HIV-associated neuroinflammation, and these combined effects are associated with impaired decision-making and heightened risk-taking behavior. Studies examining DA functioning in humans in vivo are rare, though, as they require invasive procedures or specialized neuroimaging techniques. However, emerging functional magnetic resonance imaging (fMRI)-based analytical techniques offer a promising non-invasive alternative: tissue iron deposits in the BG have been identified as a proxy for DAergic neurophysiology and functioning. Therefore, the overall goal of the proposed project is to reanalyze fMRI data with this novel method to measure iron concentrations within the BG as a proxy for DAergic functioning in PWH and METH dependence, and to examine its relationship with inflammatory biomarkers and cognitive functioning. This proposal will leverage neuroimaging and associated clinical data from participants stratified by HIV status and METH dependence collected by the Translational Methamphetamine AIDS Research Center at UC San Diego. Understanding this shared vulnerability may provide new targets for intervention to mitigate neurocognitive changes and promote behavioral health in PWH who use METH.

Principal Investigator:  Francisco Zapatero Belinchón, J. David Gladstone Institutes

Budget:  $270,000

Start Date: March 1, 2026        End Date: February 29, 2028

Project Abstract: Despite antiretroviral therapy (ART), HIV persistence in cellular reservoirs remains the fundamental obstacle to viral eradication. I propose to develop STRIKE-HIV (Specific TAR-Responsive Identification and Killing for Eradication of HIV), an innovative CRISPR/Cas12a2-based technology that specifically recognizes and kills HIV RNA-positive cells through “chromatin shredding”. This work will be conducted under the mentorship of Dr. Melanie Ott with scientific support from Dr. Jennifer Doudna, Nobel laureate for her pioneering discovery of the CRISPR technology.

Principal Investigator: Elizabeth Hastie, MD, UC San Diego

Budget:  $265,658

Start Date: March 1, 2026        End Date: February 29, 2028

Project Abstract: Transgender women with HIV (TWH) experience profound health disparities, including an estimated global HIV prevalence of 20%, yet remain underrepresented in HIV research. Gender-affirming hormone therapy (GAHT) improves engagement in care, quality of life, and HIV outcomes, as shown in the landmark LEGACY study (Lancet HIV, April 2025). While the importance of GAHT in HIV care has been established, its immunologic effects on HIV pathogenesis remain poorly defined, exposing a critical gap affecting an underserved population.

Principal Investigator: Johannes Schlachetzki, MD, University of California, San Diego

Budget:  $270,000

Start Date: March 1, 2026        End Date: February 29, 2028

Project Abstract: As the population of people with HIV (PWH) ages, there is a rising need to address the increasing prevalence of neurocognitive impairment (NCI), characterized by deficits in learning, memory, and executive function. Despite effective viral suppression, approximately 50% of PWH are diagnosed with NCI. Emerging evidence indicates accelerated brain aging, including premature cerebral amyloid-beta deposition observed in postmortem studies of HIV-positive individuals without traditional Alzheimer’s disease (AD) risk factors. Amyloid plaques appear at a significantly earlier age and are more widespread in the brains of aging PWH than in HIV-negative peers. With improved control of the HIV infection over the last couple of decades and decreased mortality, over 40% of PWH in the U.S. are now over 55, and understanding this intersection is critical to prevent missed treatment opportunities, economic burden, and caregiver strain.

Principal Investigator: Avik Biswas, Salk Institute for Biological Studies

Budget:  $270,000

Start Date: March 1, 2026        End Date: February 29, 2028

Project Abstract: In the absence of a functional cure, antiretroviral therapy (ART) represents the primary treatment option against HIV. However, drug resistance affects all classes of ART, and estimates suggest that levels of resistance can be as high as 50-97% in populations failing therapy. There are also emerging reports of pan-resistant HIV. Drug-resistance mutations (DRMs), although detrimental in the wild-type virus, can become strongly favored in the presence of specific patterns of mutations occurring under drug selection pressure, leading to an evolutionary “entrenchment” that traps DRMs and has significant implications for the transmission and persistence of resistance. In prior work, using a Potts machine learning (ML) fitness landscape model built on HIV sequences from the Stanford HIV database, we have accurately captured the DRM acquisition times in people living with HIV (PLWH) and shown that primary DRMs against all classes of ART drugs are entrenched in the population. However, the specific sequence of mutational events (pathways) and the molecular mechanisms leading to entrenchment are not known. I hypothesize that the fitness landscapes under drug selection pressure determine the mutational pathways while structures of the mutant proteins can provide the molecular mechanisms of entrenchment along a pathway. This proposal aims to identify the mutation patterns and the sequential order of mutational events that constitute pathways leading to the entrenchment of DRMs against the latest generation of ART drugs. Entrenchment patterns will be tested in vitro using a combination of virology and biochemistry to provide direct evidence of the evolutionary trapping effect on DRMs and probe the associated consequence on transmission and persistence of resistance. Finally, mutants along specific entrenchment pathways will be analyzed using high resolution cryogenic electron microscopy (cryo-EM) to provide the molecular mechanisms of resistance along the pathway. Overall, the project seeks to produce critical insights into the mechanisms of HIV evolution under drug selection pressure that can help design next generation therapeutic strategies targeting resistant strains and lead to the development of surveillance tools to forecast emerging drug resistance. This work will also pave the way for independent research extending beyond HIV to other pathogenic infections.

Principal Investigator:  Mohamed Bouzidi, PhD, Vitalant Research Institute

Budget:  $270,000

Start Date: February 1, 2024        End Date: January 31, 2026

Project Abstract:  CRISPR-Cas9-mediated excision of the HIV provirus is a promising cure strategy. However, little is known about the actual efficacy of this approach in people living with HIV (PLWH). Reports have shown that HIV latency, the main barrier to an HIV cure, is maintained through chromatin condensation at the integration site and methylation of the provirus, making it inaccessible to CRISPR-Cas9 complexes. Taken together, it appears unlikely that the CRISPR-Cas9 approach alone will be sufficient to achieve a cure for HIV infection. Several methylation and chromatin state modulators, such as romidepsin and decitabine, are used in cancer treatment and have been proposed as latency reversal agents (LRA). Building on our promising previous work and preliminary data, we propose to investigate how pharmacological manipulation of DNA methylation and chromatin state can improve anti-HIV gene therapy. Under the mentorship of Dr. Satish Pillai, I will use DNA methyltransferase (DNMT) inhibitors and histone deacetylase (HDAC) inhibitors to improve CRISPR-Cas9 editing of the HIV provirus. We will administer DNMT inhibitors, inducing genome-wide hypomethylation, to primary CD4+ T cells and nucleofect them with anti-HIV Cas9 ribonucleoproteins (Cas9-RNPs). We will apply targeted bisulfite sequencing and Tracking of Indels by Decomposition (TIDE) analysis to evaluate the impact of methylation on gene editing. We will transduce J-Lat cells (latently-infected Jurkat cells) with the CRISPRoff-V2.1/TeTv4 system, consisting of methylation or demethylation machinery, respectively fused with a dead Cas9, to hyper- and hypomethylate specific sites in the provirus and apply the previously described approach. Nucleofection of a panel of J-Lat clones, characterized by unique integration sites, with anti-HIV Cas9-RNPs followed by assay for transposase accessibility and sequencing (ATAC-seq) and Tracking of Indels by DEcomposition (TIDE) analysis will be used to determine the impact of chromatin state on anti-HIV gene editing. Additionally, we will administer HDAC inhibitors to latently infected primary CD4+ T cells and use the same method to confirm our observations ex vivo. We expect to observe higher editing efficiency in proviruses harboring unmethylated CpG islands, located in open chromatin regions. Ultimately, this project may lead to key enhancements in anti-HIV gene therapy.

Principal Investigator:  Wenli Mu, PhD, UC Los Angeles

Budget:  $269,998

Start Date: February 1, 2024        End Date: January 31, 2026

Project Abstract:  Hematopoietic stem and progenitor cell (HSPC) transplantation underlies a HIV cure reported in at least 3 individuals to date. HSPC-based gene therapy can support lifelong generation of functional immune progeny. Whereas peripheral T cell gene therapy often involves ex vivo non-physiologic stimulation that skews T cell phenotypes and alters function and persistence in vivo, HSPCs display lifelong self-renewal properties and produces naturally developed, functionally normal progeny. Importantly, CD4-chimeric antigen receptor (CAR)-modified HSPCs can differentiate into multiple hematopoietic lineages, including T cells, macrophages and NK cells in humanized BLT mice and nonhuman primates (NHPs), suggesting that CAR-engineered HSPCs are capable of providing greater and broader immune responses to combat HIV. Our preliminary data demonstrates that CD4-based CAR macrophages (MQs) can direct phagocytosis and cytotoxicity against HIV envelope-expressing cells in vitro. However, besides CAR-expressing T cells, HSPC-derived CAR cells in other lineages have not been fully characterized, and their functionality as well as their interaction with CAR T cells remains unknown. Our hypothesis is that CAR-expressing natural killer (NK) cells and MQs constantly produced from gene modified HSPCs can provide innate immune surveillance benefits by facilitating antigen presentation and secreting pro-inflammatory cytokine to boost both CAR T and endogenous CTL function while targeting different anatomical reservoirs. We propose by carefully selecting CAR molecule candidate domains that can enhance whole-cell phagocytosis and effector functions, we aim to optimize the function of anti-HIV CAR-MQs and CAR-NK to ultimately improve their therapeutic potential and provide critical antiviral activities in synergy with CAR-T cells. By transducing HSPCs with optimized CD4-based CAR constructs, we aim to provide a lifelong source of CAR effector cells in multiple lineages capable of suppressing/eliminating HIV-1 replication to achieve functional cure. 

Principal Investigator:  Jennifer Dan, MD PhD, UC San Diego

Budget:  $270,000

Start Date: February 1, 2024        End Date: January 31, 2026

Project Abstract:  One of the foremost barriers to HIV cure is the latent viral reservoir. Despite antiretroviral therapy (ART), HIV lies dormant within tissues and various immune cells. Once ART is stopped, HIV re-emerges from these tissues and immune cells. Specifically, HIV lies latent within T follicular helper (Tfh) cells. A Tfh cell is a specialized CD4+ T cell whose main function is to instruct nearby B cells within germinal centers (GC). This results in B cell differentiation into memory B cells and plasma cells and the development of high affinity antibodies, critical processes of acquired immunity. However, HIV-infected GC Tfh cells have diminished Tfh function compared to uninfected Tfh cells, permitting persistence. We have developed the Activation Induced Marker (AIM) assay to identify antigen-specific CD4+ T cells and GC Tfh cells in secondary lymphoid tissue. In addition to the traditional Tfh cells and regulatory Tfh cell, we were the first to describe the phenomenon of a “killer” Tfh cell. These “killer” Tfh cells are similar to cytolytic CD4+ T cells in circulation. HIV-specific cytolytic CD4+ T cells are capable of killing HIV-infected CD4+T cells and controlling infection. Higher frequencies of HIV-specific cytolytic CD4+ T cells have been observed in long term non-progressors and elite controllers. We hypothesize that “killer” Tfh cells within lymphoid tissues are akin to cytolytic CD4+T cells in circulation with the ability to obliterate the latent viral reservoir. The NIH-funded Last Gift Study is ongoing in San Diego and provides a unique opportunity to understand the HIV reservoir across the human body. Last Gift participants, upon their death, undergo a rapid research autopsy within 6 hours, which allows collection of fresh tissue with optimal cellular viability and protein and nuclei acids integrity. The rapid autopsy team collects tissues extensively throughout the entire body including entire lymph nodes, which can be used for this pilot project to probe the latent HIV reservoir. Under the mentorship of Dr. Davey Smith, we will assess HIV-specific Tfh cells in fresh lymphoid tissues collected from 10 Last Gift participants. Our long-term goal is to show that HIV-specific “killer Tfh” cells exist within HIV lymph nodes and can be harnessed to kill HIV-infected GC Tfh cells, thereby providing a means to eliminate the latent HIV reservoir.

Principal Investigator:  Mattia Trunfio, DTMH MD(c), UC San Diego

Budget:  $270,000

Start Date: February 1, 2024        End Date: January 31, 2026

Project Abstract:  Despite the abundance of data about HIV infection in the central nervous system (CNS), most studies focus on the brain and neglect the spinal cord. Investigations on neurocognitive complications, HIV persistence, or antiretrovirals (ARVs) penetration into the CNS have mostly used cerebrospinal fluid (CSF) and brain tissues. Our preliminary data reported higher HIV DNA levels in the spinal cord compared to brain tissues with heterogeneity in HIV reservoirs across brain areas. Compared to the brain, the spinal cord has its own peculiarities that encompass the blood-spinal cord barrier and the tissue-specific abundance and functioning of macrophages and microglia, which carry the highest burden of HIV in the CNS. Therefore, neither ARVs penetration nor the proviral dynamics in the spinal cord can be inferred from what is known for other CNS parts. Our overarching hypothesis is that the spinal cord is a distinct site of HIV persistence compared to other CNS regions (in terms of size, activity, and diversity), that it represents an independent source of viral dispersal when ARVs are interrupted, and that measures of HIV persistence and dispersal are influenced by ARVs tissue levels and inflammation. To investigate these hypotheses, we will analyze flash frozen tissues from the pons, medulla, and the cervical, thoracic, and lumbar segments of the spinal cord of 15 people with HIV (PWH) enrolled in the Last Gift program. The Last Gift program (San Diego) enrolls altruistic PWH with a life-shortening illness who participate in HIV cure research at the end-of-life, including full body donation for a rapid research autopsy. As part of this proposed project, we will 1) measure levels of HIV RNA and HIV DNA (ddPCR), and proviral diversity (full length HIV DNA single genome sequencing) in various spinal segments, 2) evaluate how these reservoir measures relate to CSF inflammation and ARVs tissue levels, 3) determine if the spinal cord is a source of HIV re-seeding in PWH who voluntarily interrupt ARVs before death. We will compare these data with similar reservoir data generated from brain, lymph tissues, blood, and CSF made available as part of the ongoing NIAID-funded program (AI169609). Our results will inform larger studies addressing HIV reservoirs in the CNS with the aim of developing strategies to effectively clear HIV infection. Mentor: Prof. Antoine Chaillon.

Principal Investigator:  Yusuke Matsui, PhD, J. David Gladstone Institutes

Budget:  $270,000

Start Date: February 1, 2024        End Date: January 31, 2026

Project Abstract:  The brain is an understudied target organ for HIV-1, and ~50% of patients on antiretroviral therapy experience neurocognitive disorders, known as HIV-associated neurocognitive disorder (HAND). Microglia are brain-resident macrophages and the target cells for HIV-1. Microglia are heterogeneous; they interact with surrounding neurons and astrocytes and alter their gene expression profiles in response to environmental cues, forming characteristic “subsets” based on transcriptome analysis. Here, we hypothesize that subset characteristics of microglia influence susceptibility to HIV-1 and the establishment and maintenance of latent infection. Determining the precise identity of the microglia reservoir is critical for our understanding of how latency is established and maintained in the brain and for the development of effective treatment aimed at HIV-1 eradication in central nervous system (CNS). We propose to test our hypothesis using brain organoids containing microglia. Brain organoids containing microglia are being generated from human induced pluripotent stem (iPS) cells and are emerging as a new tool to study brain disorders in HIV infection. Presently, microglial subsets have not been mapped in brain organoids, but transcriptome and cell surface marker analyses suggest that brain organoids reproduce the physiological brain environment. Also, iPSC-derived macrophages contained in alveolar organoids successfully reproduce in vivo subset characterization. We will generate two types of brain organoids containing microglia and classify subsets of microglia based on transcriptome expression. We will combine this technology with cutting-edge single-cell RNA sequencing, CRISPR-Cas9 knockout experiments, light-sheet microscopy, and infections with dual-fluorescent HIV clones. Our studies are significant because they address an important remaining hurdle in curing HIV: characterizing and eradicating the viral reservoir in the brain. They are innovative because they challenge the status quo by defining subset characteristics in brain organoids with microglia. After completion of our studies, we will have accomplished mechanistic understanding that will lead to new HIV-1 eradication strategies against viral reservoir of CNS by characterizing subsets of microglia that are the breeding ground for latent infection in the brain. Proposed mentor's name: Melanie Ott

Principal Investigator:  Jocelyn Kim, MD PhD, UC Los Angeles

Budget:  $270,000

Start Date: February 1, 2024        End Date: January 31, 2026

Project Abstract:  In 2020, 38 million people worldwide were living with HIV/AIDS. Antiretroviral therapy (ART) can suppress but not cure HIV infection, mainly because of the persistence of infected cells. These cells arise from the ability of HIV to integrate its viral genome into the host cell’s chromosomes and remain quiescent and elude the immune system. This reservoir of latently infected cells is the most formidable obstacle to curing HIV. One approach to eliminating latently infected cells is a “kick and kill” strategy, in which latent cells are induced to produce viral proteins by a latency reversing agent (LRA).  Our collaborator Dr. Paul Wender, an expert synthetic chemist, synthesized the first bryostatin-1 analog (SUW133) as a superior LRA compared to its parent compound. However, SUW133 reversed latent cells in vivo, but not all infected cells emerging from latency were killed.  To this end, we investigated whether natural killer (NK) cells could help kill these persistent infected cells. NK cells are immune cells that inherently control HIV infection. Due to their intrinsic anti-viral and anti-cancer function, allogeneic NK cells are already under clinical investigation to treat COVID-19 patients and have been safely and effectively used in patients with leukemia.  However, chronic HIV infection results in their loss of function, which is not completely restored by suppression of viral replication by ART.  Thus, we suspected allogeneic NK cells from healthy human donors may be an attractive source for providing a potent anti-HIV killing agent. Interestingly, we found a kick and kill strategy using the LRA SUW133 as the “kick” and human allogeneic NK cells as the “kill” yielded remarkable results compared to the LRA or NK cells alone with substantial delays in viral rebound after stopping ART in HIV-infected humanized mouse series.  Importantly, this kick and kill strategy cleared HIV infection from 40% of infected animals. This work thus far provides proof-of-concept that NK cells plus an LRA leads to unprecedented in vivo reservoir reduction, a task nearly insurmountable in the field of HIV eradication. Further research to improve this strategy is urgently needed. Here our overall objective is to engineer and improve NK cells to efficiently fight HIV infection.

Principal Investigator:  Sarah LaMere, PhD DVM, UC San Diego

Budget:  $270,000

Start Date: February 1, 2024        End Date: January 31, 2026

Project Abstract:  Despite decades of research into the mechanisms of HIV persistence, a cure remains elusive. Basic biomedical research has focused strongly on the mechanisms of viral infection and replication, leading to the development of many successful antiretroviral therapies and allowing people with HIV to live without fear of developing AIDS as long as they maintain their medication regimen. However, there are two large obstacles that have prevented the advancement of cure research: 1) poor understanding of the epigenetic regulation of the provirus and the interacting host chromatin, which impacts our ability to manipulate, excise, or silence the provirus and 2) poor characterization of the latent reservoir throughout the body due to limited access to rapid autopsy specimens, which largely impacts the success of drug delivery. Under the guidance of Dr. Davey Smith, I propose to address both of these hurdles by making use of recent advancements in single cell techniques and high-resolution chromatin characterization to investigate putative epigenetic mechanisms in tissues. These tissue reservoirs are critical, as they likely contribute to repopulation of systemic HIV upon discontinuation of ART. This project will take advantage of samples collected from the ‘Last Gift’ cohort co-created by Dr. Smith, which entails that participants provide their entire bodies after they die for a rapid research autopsy protocol. Data generated from this proposal will be integrated with data generated as part of the parent grant, which already includes cell-associated viral RNA expression, proviral load, T cell receptor sequencing, integration site sequencing, and full-length HIV genome sequencing. We hypothesize that the integrated HIV provirus impacts host chromatin conformation, requiring a 3D examination extending beyond simple epigenetic modifications. Further, we hypothesize that this impact depends heavily upon both integration site locale and cell type. Therefore, we propose to examine the impact of the provirus upon 3D chromatin conformation and associated epigenetic modifications alongside integration sites in both a CD4 T cell latency model and then in two separate tissues documented to have evidence of clonal expansion from two Last Gift participants.