Wistar Institute

NIH Research Projects · FY 2026 · 2026-06

PROJECT SUMMARY Antiretroviral therapy (ART) results in suppression of Human Immunodeficiency Virus (HIV) replication, increase in CD4+ T cell count, and partial restoration of immune responses. Yet despite suppressive ART, both transcriptionally silent and transcriptionally active [i.e. cell-associated RNA (CA-RNA)] HIV viruses remain in various CD4+ T cell subsets in persons living with HIV (PLWH) and are proposed to contribute to residual immune activation, lower immune reconstitution and development of comorbidities. Much attention has been given in our field to the clinical significance of continued HIV expression after ART, but we still lack plasma host biomarkers that could illustrate the impact of CA-RNA or could link CA-RNA to future risk for comorbidities. Part of the limitation to address this gap has been that studies attempting to find a correlation between HIV reservoir levels and plasma markers of immune activation have included only a limited number host cellular or plasma variables, have not started with a pre-defined cohort with known reservoir levels, have not linked variables to predefined prognostic biomarkers, and/or have yielded inconsistent results (see background). This R21 now addresses these limitations by studying a cohort with known HIV reservoir levels and by introducing recent advances in Somalogic-based technology inclusive of (a) measuring a large set (at least 7000) of plasma proteins and (b) determining links of predefined host biomarker "signatures" to risk scores for comorbidity outcomes as shown by our preliminary data and other studies. This R21 proposal is possible as a result of the collection through the BEAT-HIV Martin Collaboratory program of plasma and peripheral blood mononuclear cells (PBMC) from 94 ART suppressed PLWH with known distribution of HIV proviral DNA measured by the Intact Provirus Assay (IPDA) which in turn is associated with CA-RNA. Specifically, we hypothesize that in PLWH on suppressive ART the levels of cell-associated HIV RNA will be associated with a) plasma host proteomic biomarkers, and b) pre-defined clinical comorbidity risk scores for cardiovascular, liver/kidney, and metabolic disease following age adjustment. We will test this hypothesis by the following specific aims: (1) identify host plasma biomarkers among 7000 protein measures that will best predict PLWH on ART with higher CA-RNA independently of age, and (2) determine if plasma clinical prognostic scores for comorbidity risks of cardiovascular, liver/kidney, and metabolic disease are higher in PLWH on ART with high CA-RNA when adjusted for age. Completion of this R21 proposal will provide foundational data to further evaluate the usage of biomarker changes in future RO1s describing strategies targeting transcriptionally active reservoirs on ART. The long-term impact of this proposal is its potential to advance cure-directed strategies targeting HIV expressing cells (i.e., immune-based, gag- pol/CARD8 activating, etc.) and to make possible early determinations in biomarker changes of significance to prognosis risk profiles upon a reduction of persistent HIV reservoirs.

NIH Research Projects · FY 2026 · 2026-06

Project Summary Objective: The overall goals of this application are understanding the molecular basis of RNA Polymerase II (RNAPII) pausing and pause-release and determining how disease-associated mutations in the PP2A phosphatase reverse the pausing balance to promote transcriptional elongation. Proposed research: The mammalian RNAPII transcription cycle is regulated by cyclin-dependent kinases (CDKs) that target the catalytic core of RNAPII and many of its co-factors. We recently identified Int-PP2A, a module of the RNAPII-associated Integrator complex that recruits the PP2A phosphatase to active genes to oppose CDK9 activity. We demonstrated that Int-PP2A targets Ser-2 residues at the C terminal domain (CTD) of RNAPII as well as DSIF/Spt5. While our work revealed that phosphatases are recruited co-transcriptionally in mammalian cells to promote pausing, it remains unknown how Int-PP2A’s activity is established at the proximal promoter and how it is modulated. Here we propose to: a) determine how Int-PP2A establishes paused RNAPII via the INTS3 subunit; b) determine the role of PP2A carboxy-methylation in transcriptional regulation; c) determine the effect of mutant PP2A on transcription in models of neuronal differentiation.

NIH Research Projects · FY 2026 · 2026-05

PROJECT SUMMARY Glioblastoma (GBM), the most aggressive and lethal form of brain cancer in adults, is highly resistant to promising immunotherapies, largely due to a myeloid cell-driven immunosuppressive microenvironment (TME). The emerging key contribution of neutrophils to this immunosuppressive milieu that restricts anti- cancer immunity, therefore promoting tumor growth and resistance to immunotherapies, has sparked interest in therapeutically targeting these cells in brain tumors. Despite recent advances in understanding the phenotypic and functional heterogeneity of neutrophils in tumor beds, there are no effective approaches to specifically target immunosuppressive subsets. Thus, the goal of this study is to dissect targetable tumor- driven mechanisms sustaining pro-tumoral and immunosuppressive neutrophils in the TME; as this knowledge is the key to developing impactful, new, and selective neutrophil-targeting therapies to unleash anti-tumor immunity and sensitize resistant tumors, like GBM, to different forms of immunotherapy. Our preliminary data show that the GBM TME specifically reprogrammed a subset of immunosuppressive neutrophils into cells with prolonged lifespans, providing abnormally high levels of long-lasting immunosuppressive cells in tumor beds. Immunosuppressive neutrophils showed boosted glucose metabolism, which correlated with high resistance to ferroptosis and high levels of histone lactylation (Kla). Since neutrophils are generally considered short-life cells, it becomes important to understand if and how local tumoral cues are linked to adaptive changes enabling survival of specific neutrophil subsets. Based on our preliminary key observations, we hypothesize that actionable histone lactylation-driven resistance to ferroptosis selectively maintains the pool of immunosuppressive neutrophils in hypoxic niches of tumors. We will test this hypothesis through the following aims: Aim1. To define metabolic cues that sustain immunosuppressive neutrophils in tumor beds; Aim2. To dissect the glucose catabolism-driven epigenetic mechanism determining resistance to cell death of immunosuppressive neutrophils; Aim3. To establish the therapeutic potential of targeting suppressive neutrophils. Thus, the proposed studies will elucidate tumor- induced metabolic and epigenetic determinants that sustain the pool of immunosuppressive neutrophils in tumor beds. Our work will exert a profound effect in the field by providing a mechanistic rationale to specifically target a major immunosuppressive, tumor-promoting component of the GBM TME, therefore contributing to the refinement and expansion of therapeutic approaches to overcome immunosuppression and enhance the efficacy of immunotherapy.

NIH Research Projects · FY 2026 · 2026-05

PROJECT SUMMARY The main barrier to developing an efficacious HIV-1 vaccine is the requirement of eliciting broadly neutralizing antibodies (bNAbs) targeting neutralization-sensitive epitopes of the Envelope (Env) glycoprotein. A major problem is the frequent and overwhelming induction of non-neutralizing antibody responses directed to the gp41 base of Env when using soluble Env trimers as immunogens. These off-target antibodies fail to neutralize HIV- 1, as the Env base is occluded by the viral surface and not accessible for antibody recognition in HIV-1 virions. The base of soluble Env immunogens has been shown to be immunodominant in preclinical animal models and humans, where base antibodies have derailed multiple clinical trials. Despite numerous attempts to occlude the Env base using glycans, membrane-bound Env and nanoparticle scaffolds, the immunodominance of the base persists. Interestingly, our preliminary studies show that the specificity of anti-Env antibody lineages is not fixed, instead, antibody lineages can switch their epitope specificity through somatic hypermutation in response to immunization with Env. In particular, we show that V3-glycan bNAb lineages can lose track to become base antibodies and that base antibodies can be redirected to target the neutralizing V3-glycan epitope. Thus, we hypothesize that bNAb precursors can be derailed from maturing into bNAbs and that non-neutralizing antibodies can be rescued to mature into bNAbs through immunization. Here, we will investigate this phenomenon of epitope switching from on-target to off-target epitopes and vice versa through characterization of “derailed” and “rescued” antibodies. We will investigate the on-target to off- target switch to inform vaccine design strategies that prevent antibody switching towards the Env base. We will investigate the off-target to on-target switch to design vaccine approaches that recycle base responses to induce bNAbs. In Aim 1, we will investigate the mechanisms of epitope switching from the neutralizing V3-glycan epitope of Env to the base using mouse models. In Aim 2, we will investigate the mechanism of epitope switching from the base to the neutralizing V3-glycan epitope and the potential of “rescued” antibodies to mature into bNAbs in mouse models. In Aim 3, we will investigate whether other bNAb lineages and other off-target antibodies can undergo epitope switching too. We have produced nine new humanized and rhesus-ized mouse models and innovative AI-generated immunogens that we will leverage for these specific aims. Structural approaches will be used to characterize the molecular mechanisms of epitope switching. These aims will inform immunogen design strategies to keep bNAb lineages on track after initial activation and to “rescue” off-target responses. Collectively, this proposal will investigate previously overlooked challenges for HIV-vaccine design and will bring new opportunities not only for HIV vaccines but for vaccine design in general.

NIH Research Projects · FY 2026 · 2026-04

Project Summary The treatment of pancreatic ductal adenocarcinoma (PDAC) remains a significant challenge, with a 5-year survival rate of just 13%. Resistance to therapies is largely attributed to the immuno-suppressive tumor microenvironment (TME), which is characterized by a fibrotic stroma and high infiltration of immuno- suppressive cells, including tumor-associated macrophages (TAMs), that hinder effector T cell responses. Shifting the immuno-suppressive TAM phenotype toward an immune-activated state presents a promising therapeutic strategy. Previous studies, including our own, suggest that TAM phenotypes are shaped by metabolic pathways. Notably, the metabolism of branched-chain amino acids (BCAAs) has gained attention, as elevated BCAA levels have been associated with more than a twofold increased risk of developing PDAC. While prior research has focused on BCAA metabolism in tumor cells, particularly through the study of BCAA metabolism-related kinases and enzymes, the role of BCAA metabolism in TAMs and its impact on TAM phenotype and anti-tumor immunity remains unexplored. Consequently, it remains unknown whether, and how, this pathway could be therapeutically targeted. Our findings indicated that immuno-suppressive TAMs exhibit reduced BCAA oxidation and reduced incorporation of 13C-labeled BCAAs into the TCA cycle. Increasing BCAA oxidation in TAMs (via BCKDK deletion) enhanced their immuno-stimulatory phenotype, activated CD8+ T cells, and reduced PDAC growth. Conversely, BCAA supplementation induced an immuno- suppressive TAM phenotype and activated the protein kinase RNA-like endoplasmic reticulum kinase (PERK) pathway, a key arm of the unfolded protein response. PERK deletion in TAMs reversed this effect, promoting immune activation. Pharmacologically targeting BCAA metabolism—either by increasing BCAA oxidation or inhibiting PERK signaling—enhanced anti-tumor immunity and reduced PDAC growth. Lastly, in patients with solid tumors, high BCAA oxidation and low PERK signaling correlated with better responses to anti-PD1 therapy. We propose that reduced BCAA oxidation and increased BCAA signaling in TAMs drive immuno- suppression in PDAC. Aim 1 will investigate the mechanisms by which BCAA oxidation and BCAA signaling in TAMs differentially influence immune responses in PDAC, focusing on TAM-specific (i) BCAA oxidation via the TCA cycle and OXPHOS, and (ii) BCAA signaling via the PERK pathway. Aim 2 will test whether pharmacologically targeting BCAA metabolism improves chemo-immunotherapy responses in aggressive PDAC mouse models and patient-derived organoids. Successful completion of these objectives will elucidate how TAM-specific BCAA oxidation and BCAA signaling influence immuno-suppressive TAM phenotype. In the long term, this work could lead to novel therapies targeting TAM-specific metabolism for PDAC treatment.

NIH Research Projects · FY 2026 · 2026-04

PROJECT SUMMARY Most ovarian cancer (OC) patients respond to the front-line chemotherapy but succumb to chemoresistant relapse with peritoneal metastases. How chemoresistance develops in OC patients after the front-line chemotherapy remains poorly understood. Recent understanding of the tumor immune microenvironment (TIME) suggests that inflammatory mediators from myeloid cells play key roles in developing chemoresistance. This proposal seeks to understand whether and how a classical inflammatory mediator from myeloid cells, interleukin 1β (IL1β), promotes OC chemoresistance, aiming to eradicate metastatic OC by overcoming IL1β-mediated chemoresistance. Based on published and our preliminary results, we hypothesize that myeloid cell-derived IL1β promotes chemoresistance in metastatic OC by modifying TIME via activation of OC cells and fibroblasts; combining IL1β neutralization and chemotherapy eliminates chemoresistant OC. This will be tested with the following Aims. Aim 1: Test the hypothesis that the combination of chemotherapy and IL1β neutralization overcomes OC chemoresistance. Aim 2: Test the hypothesis that myeloid cell-derived IL1β drives OC chemoresistance by modifying TIME. Aim 3: Test the hypothesis that myeloid-derived IL1β promotes formation of chemoresistant spheroids.

NIH Research Projects · FY 2026 · 2026-04

Project Summary Nuclear processes such as transcription, replication, and repair disrupt the double stranded nature of DNA to result in the formation of non-canonical structures, the most prevalent being R-loops and G-quadruplexes (G4s). R-loops are three-stranded nucleic acid structures, which occur co-transcriptionally and comprise an RNA-DNA hybrid and a displaced single strand of DNA. The displaced ssDNA can form G4s if it contains spaced stretches of guanine nucleotides. G4 formation can facilitate the formation and stabilization of R-loops. Both R-loops and G4s have important roles in normal cellular function but are also deregulated in several developmental disorders and cancers. Our long-term goal is to elucidate how R-loops and G4 levels are managed in normal cells, how they regulate gene expression and genome organization, and how their misregulation contributes to various diseases. We discovered an enrichment for zinc finger domain containing proteins at R-loops and characterized one such factor, ADNP, to show that ADNP can resolve R-loops in vitro and that its loss results in accumulation of R-loops in vivo. We also identified a role for R-loops and G4s in genome organization by showing that these structures promote CTCF binding genome-wide, providing a new model for how transcription-induced R-loops can alter genome folding and contribute to lineage-specific gene expression programs during development. Through our work on another R-loop regulator, ATRX, we showed that while ATRX cannot resolve R-loops in vitro, its RNA binding activity can prevent R-loops from forming in vitro. In the next award cycle, we propose to build on these studies to: 1) identify a broader role for zinc finger proteins in genome organization through their interactions with R-loops and G4s; 2) determine the extent and mechanisms of ATRX-mediated R-loop regulation genome-wide and the impact on the localization of epigenetic regulators; and 3) elucidate the contribution of R- loops and G4s on the spatial organization of the genome.

NIH Research Projects · FY 2026 · 2026-04

PROJECT SUMMARY 1. Overview of research in the laboratory There are currently two focuses of my lab: One focus is on mechanisms controlling the development of macrophages after differentiating from monocytes in the steady state and disease settings. This is an important but poorly understood biological process. Most studies in the past have been focusing on the differentiation process from monocytes to macrophages, not much on their continued development afterwards. As described in the application, we propose a novel concept that in order to develop into professional phagocytes that handle apoptotic cells efficiently and “silently”, the Tim4- macrophages need to be “trained and tested” continuously to become qualified to develop into Tim4+ macrophages. Furthermore, we aim to reveal novel epigenetic and metabolic regulators for this developmental process. The other focus of my lab is to leverage myeloid cell activation to treat metastatic ovarian cancer. The first manuscript of my lab on this topic was recently published in the Journal of Experimental Medicine and has received top 2-3% attention scores. 2. Goals for the next five years The goals include the following: (1) to test how efferocytosis and IL4 signaling synergistically promote the development of Tim4+ macrophages, (2) to profile the epigenetic and metabolic changes of macrophages during their development into Tim4+ macrophages, and (3) to identify novel epigenetic and metabolic regulators for the development of Tim4+ macrophages. 3. Overall vision of the research program This research program is designed to address the fundamental question: “What factors dictate the development Tim4+ macrophages from Tim4- macrophages after they finish differentiation from monocytes?”. This question is of paramount significance for general medicine because of the broad presence and key role of Tim4+ macrophages in virtually all tissues as well as them being the best efferocytes, i.e., phagocytosing apoptotic cells silently and efficiently, in the body. Dysregulation of Tim4+ macrophages could affect a wide spectrum of diseases, from autoimmune diseases to severe infections, to cancer, and to tissue injury. Based on published results from others and promising data fully generated in my own laboratory at The Wistar Institute in the past year, we propose a new model for the development of Tim4+ macrophages: “continuous “training and testing” of silent and efficient efferocytosis by macrophages eventually produces Tim4+ macrophages”. Completion of this research program will provide convincing evidence for this new model and reveal novel epigenetic and metabolic regulators for this process. This will open a new direction for modulating macrophage heterogeneity. Considering its significance in almost all diseases, future work will focus on manipulating the newly identified regulators to modulate macrophage subsets for better disease outcomes.

NIH Research Projects · FY 2026 · 2026-03

Project Summary The long-term goal of our lab is to understand the fundamental mechanisms by which RNA structures and RNA modifications can help cells distinguish between "self" and "non-self" molecules. One of these potent non-self molecules is double-stranded RNA (dsRNA), a particular structure of RNA that is not present at high levels in unstressed cells, but can be seen as a hallmark of viral infection. Unfortunately, transcription of endogenous repetitive elements within our own genomes can lead to the formation of dsRNA and activation of RNA sensors and inappropriate destructive cell signaling pathways even in the absence of infection. As such, the balance between sensing pathogenic dsRNA and tolerating self dsRNA must be finely tuned by negative regulators of RNA sensing. In addition, RNA sensing is also compartmentalized, with sensors in the cytoplasm kept away from the nucleus where RNA transcription takes place. Important for human health, loss of this balance in either direction is harmful, with negative regulator loss-of-function leading to inflammatory disease or autoimmunity while gain-of-function has been associated with cancer. Our research has shown that human viruses can actively prevent the formation of dsRNA by regulation of RNA splicing and removal of complementary introns. However, perturbation of viral splicing efficiency leads to production of nuclear dsRNA that is sensed by cytoplasmic sensors that subsequently move to the nucleus. This tool provides a unique opportunity to assess how cells respond to dsRNA derived from the nuclear compartment, and how tolerance of this non-self molecule can be broken. This proposal will explore the basic biological principles governing the interactions between nuclear dsRNA, RNA processing, and cytoplasmic RNA sensors. We will ask fundamental questions about how RNA structures, including intramolecular hairpins and intermolecular two-stranded helices, influence RNA sensor activation and the interplay between redundant sensing pathways. Additionally, we will investigate whether nuclear dsRNA carries specific chemical modifications that modulate its structure, recognition by sensors, or sensor activity. Finally, we will determine RNA binding proteins that specifically bind or regulate nuclear dsRNA using quantitative proteomics. The results will shed light on how healthy cells prevent inappropriate activation of RNA sensing responses to self-derived RNA. We will also learn how stressful situations, such as viral infection, can rewire existing signaling pathways to allow nucleic acid surveillance of the nucleus. The completion of this project will ultimately lead to a greater mechanistic understanding of nucleic acid sensing in cellular health and homeostasis.

NIH Research Projects · FY 2026 · 2026-02

Project Summary/Abstract While the advent of antiretroviral therapy (ART) has improved the lives of many people with HIV (PWH), HIV infection remains persistent for the lifetime of most individuals. ART interruption leads to viral rebound and renewed disease progression, thus highlighting the existence of the latent HIV which exists even when viral RNA is undetectable in plasma during ART. To date, there is no broadly accessible cure for HIV. This requires most PWH to stay virally suppressed with a constant ART regimen. As such, there remains an urgent need to find functional cures of HIV and alleviate the need for a lifetime of costly ART and its associated long-term side effects. However, the complex nature of the HIV reservoir has impeded our collective efforts to understand the reservoir for elimination strategies. Viral transcription decreases over time on ART and can be nonexistent in many HIV+ cells, thus leading to evasion of immunosurveillance. The leading concept in the field for eliminating the reservoir is to reverse latency using pharmacological agents so that viral antigen is produced to render HIV+ cells permissive for elimination by an interventional therapy. However, current attempts at latency reversal are neither efficient nor complete as only a fraction of HIV+ cells are reactivated. This suggests that transcriptional regulation of the various proviruses (i.e. the integrated viral DNA) is highly heterogeneous. Evidence further suggests that proviral epigenetics, defined as the regulation of transcription at the chromatin level independent of sequence, is critical in understanding how the latent HIV reservoir is established, regulated, and disrupted. Some proviruses are situated in more condensed (heterochromatic) DNA without transcription, while others are in more accessible (euchromatin) DNA that is more conducive to transcription. This spectrum of reactivation potential means that reservoir elimination will first require a single-cell investigation to precisely define these proviral epigenetic states and assess their contribution to latency reversal. Therefore, the primary objective for this project is to define the ex vivo proviral epigenetic landscape at single-cell resolution and to develop a pioneering strategy to selectively reverse latency. The underlying central hypothesis is that HIV+ cells with heterochromatin-associated proviruses are more resistant to reservoir disruption and latency reversal. The central hypothesis will be tested through two specific aims: 1) define the proviral epigenetic landscape and its susceptibility to reservoir disruption from ART-treated PWH; and 2) determine the efficacy of mRNA-LNP delivery of HIV-specific transcription activator-like effectors (TALEs) for latency reversal. This proposal is highly innovative because it represents a major shift in how the field studies the HIV reservoir by using new single-cell methodology to circumvent the challenges inherent to studying the HIV reservoir. The outcomes will uniquely define the identities of the latent HIV reservoir and will offer additional opportunities to target and eliminate the HIV reservoir.

NIH Research Projects · FY 2025 · 2025-07

SUMMARY We have demonstrated multiple lines of evidence supporting the role for autologous neutralizing antibodies (anAbs) to restrict virus rebound. The positive relationship between anAb suppression of virus reactivation ex vivo and time to rebound in vivo indicates that anAbs are a strong host determinant in the suppression of the HIV viral reservoir. This offers a unique opportunity to develop an individualized HIV cure strategy targeting the anAb-resistant component of the HIV reservoir that is not controlled by the host upon reactivation. Taking advantage of the full complement of immune responses (i.e., a combination of innate, humoral and cell-mediated components), this proposal seeks to develop an individually tailored strategy that will bring together a) neutralizing antibody responses enhanced via mRNA-lipid nanoparticle (LNP) technology, b) anAb-resistant HIV Env HLA-E restricted adaptive NK cells or creation of multi-specific molecules targeting the HIV loaded HLA-E MHC complex, and c) engineered cell-mediated effector strategies, consisting of multivalent CAR T cells and CD64-transduced NK cells with a membrane-bound combinations of broadly neutralizing antibodies optimized against the anAb-resistant reservoir of each participant. Our preliminary data indicates that each of the strategies proposed has potential for antiviral control, supporting their inclusion in the combined approach proposed. Our central hypothesis is that characterization of the anAb-resistant HIV reservoir will permit development of a targeted, personalized combination strategy of antiviral innate, humoral and cell-mediated responses, which together will elicit sustained viral control and/or viral eradication. To test this hypothesis, we will first conduct comprehensive reservoir screens in persons with HIV suppressed on ART after early or late ART initiation (to evaluate strategies against distinct levels of reservoir diversity) to identify sequences of anAb-resistant reservoir viruses which will be used to generate individual tailored anAbs (induced by mRNA-LNP-based vaccination) and, concurrently, to identify the best combination of existing broadly neutralizing antibodies (bnAbs) that can achieve virus control. Second, we will build personalized cell-mediated responses to clear infected cells by (i) HLA-E responsive Env peptides for adaptive NK cell generation, (ii) phage display targeting for bi/tri-specific HLA-E- Env peptide complex engagers, and (iii) individually defined bnAb combinations loaded on to CD64-NK or expressed as tri-bnAb constructs on CAR T cells. To facilitate future clinical deployment of this combined immune strategy, we will develop personalized virus reactivation strategies, wherein individual’s CD4+ cells bearing integrated HIV will be characterized to design personalized LRA approaches optimized to maximize HIV reactivation. Together with significant institutional and industry commitment, our proposal brings together collaborative groups from Accelevir, Merck, Acuitas Therapeutics, BioNtech, BlueWhale Bio, CytoImmune Therapeutics, George Washington Univ., Univ. of Nebraska, Ragon Institute, Massachusetts Institute of Technology, Duke Univ., Philadelphia FIGHT Johns Hopkins Univ., Univ. of Pennsylvania, and Wistar Institute.

NIH Research Projects · FY 2025 · 2025-07

Summary Novel vaccine strategies is needed to develop a safe, effective, and broadly accessible HIV vaccine to end the HIV pandemic. This proposal is built upon several innovative advancements. First, learned from natural immunity observed in a group of HIV resistant Kenyan female sex workers, we found 12 protease cleavage sites (PCS) in HIV are critical for virus maturation due to their essential function in tightly controlled cleavage of Gag, Gag-Pol, and Nef precursor proteins. Sequences surrounding the HIV PCS are immunogenic and highly conserved across global HIV subtypes, thereby a vaccine based on 12 PCS immunogens will be globally accessible and likely to prevent virus escape mutations. Further, our recent studies demonstrated that a PCS vaccine delivered by recombinant vesicular stomatitis virus (rVSV) vector and nanoformulation protected more than 80% of vaccinated female Mauritian cynomolgus macaques (MCMs) against SIVmac251 intravaginal challenges, which is associated with vaccine elicited potent PCS-specific CD8 T cell responses. However, in previous studies, 12 PCS immunogens were individually expressed in the rVSV. To improve vaccine immunogen presentation and facilitate the vaccine preparation, 12 PCS immunogens should be expressed in a single cassette, i.e., multi-epitope PCS (MEPCS). i.e., multi-epitope PCS (MEPCS). To that end, we have developed a cold-chain friendly and long-term stable MEPCS-mRNA-LNP vaccine. We found that SIV MEPCS-mRNA-LNP vaccine induced a potent PCS-specific CD8 T-cell immunity without inducing generalized inflammation and CD4 T-cell activation. Based on our strong preliminary date, we hypothesize that MEPCS vaccines would protect rhesus macaques against SIVmac251 rectal transmission. The reason we will use a rhesus macaque (RM)-SIV rectal challenge model is that the effectiveness of MEPCS vaccine needs to be validated in the most rigorous rhesus macaques and anal sex is the most common HIV transmission in the United States of America. The primary objective of this study is to evaluate the effectiveness of cold- chain friendly and long-term stable MEPCS-mRNA-LNP vaccine in comparison with rVSV-MEPCS vaccines in protecting RMs against SIVmac251 intrarectal challenge and to better understand the immune correlated protection. in this proposed study we will comparatively evaluate the effectiveness of MEPCS-mRNA-LNP with rVSV-MEPCS vaccines against SIVmac251 intrarectal challenge in RMs. The proposed study has the great potential to develop a new HIV/SIV vaccine to prevent HIV/SIV rectal transmission. The novel MEPCS immunogens plus the cold-chain friendly and long-term stable mRNA- LNP formulation makes this proposed study highly innovative and significant. .

NIH Research Projects · FY 2026 · 2025-04

Project Summary/Abstract The ultimate goal of this proposal is to address the fundamental gap in knowledge on the mechanisms driving resistance to immune checkpoint blockade (ICB), particularly in melanoma patients who exhibit downregulation or deletion of p16 (~40% of cases). The research plan focuses on elucidating the mechanistic role of p16 loss in regulating zinc (Zn) homeostasis through the Zn transporter SLC39A9. The goal is to explore whether manipu- lating Zn levels enhance the efficacy of current ICB in p16low melanomas. Drawing on a compelling in vivo CRISPR knockout screen, our data reveal that the knockdown of SLC39A9 sensitizes p16low melanomas to anti- PD1 ICB. Additionally, increased SLC39A9 expression correlates with unfavorable outcomes in melanoma pa- tients undergoing ICB. Notably, loss of p16 expression leads to an increase in plasma membrane SLC39A9, resulting in the sequestration of Zn from the microenvironment and creating a Zn-depleted tumor microenviron- ment. We propose two major consequences of this reprogramed Zn homeostasis in p16low tumors that contribute to impaired ICB response. Firstly, increased intracellular Zn in p16low cancer cells decreases cGAS-STING sig- naling and downregulates CXCL10, a T cell chemokine. Secondly, limited access to microenvironmental Zn compromises CD8 T cell effector function, given its crucial role in early TCR activation. This research plan is structured around two overarching scientific aims: 1) Interrogate the mechanism underlying Zn-dependent atten- uation of cGAS-STING signaling as a consequence of SLC39A9 and 2) Determine the contribution of low TME Zn on CD8 T cell recruitment and differentiation and ICB response. Successful completion of these aims will not only yield new insights into the mechanistic role of p16 in Zn metabolism but will also position the modulation of Zn homeostasis as a promising strategy to enhance therapeutic outcomes for melanoma patients with low p16 expression. Given the prevalence of p16 alterations in approximately 50% of all human cancers, the implications of these studies extend far beyond melanoma, offering a foundation for identifying metabolic vulnerabilities and informing future cancer therapeutic strategies on a broader scale.

NIH Research Projects · FY 2026 · 2025-01

PROJECT SUMMARY ABSTRACT High-grade serous ovarian carcinoma (HGSOC) is the most prevalent histosubtype of epithelial ovarian cancer. Therapy for HGSOC relies on DNA damaging agents, with a high percentage of cancers having de novo or acquired chemoresistance due in part to homologous recombination (HR) proficiency. Our preliminary data has uncovered a potential metabolic mechanism contributing to HR proficiency that could be used for novel targeted therapies for HGSOC. This proposal tests the overarching hypothesis that acetylcarnitine increases site specific histone acetylation post-translational modifications that promote HR-mediated DNA damage repair and allow resistance to standard-of-care DNA damaging agents in HGSOC. Manipulation of acetylcarnitine will therefore sensitize HGSOCs to DNA damaging agents. This project will: 1) quantitatively and mechanistically map the acetylcarnitine-dependent histone acetylation axis and its contribution to HR-mediated DNA repair; and 2) interrogate whether interventions that suppress intracellular acetylcarnitine sensitize HGSOCs to standard-of- care DNA damaging agents. Targeting acetylcarnitine metabolism for sensitization to DNA damaging agents is a novel strategy. Ultimately, this research will help develop metabolic therapeutic strategies against chemoresistance that occurs in many cancer patients.

NIH Research Projects · FY 2025 · 2024-08

PROJECT SUMMARY The loss of endogenous tumor suppressors is an obligatory step in tumor onset and progression, but the underlying mechanisms are elusive and the breadth of their targets mostly unknown. We have now discovered that Parkin, a mitochondria-associated E3 ubiquitin ligase biallelically altered in early-onset Parkinson’s Disease, functions as a novel, dual mode tumor suppressor. This involves inhibition of intrinsic tumor traits of cell motility and metabolism but also reprogramming of an antitumor immune microenvironment. In this pathway, Parkin recruitment to mitochondria “primes” immunogenic cell death, controls the release of Damage- Associated Molecular Pattern (DAMP), and activates kinase cascades for transcription of interferon genes and inflammatory cytokines, independently of mitophagy. The result is enhanced intratumoral infiltration of CD8+ T cells, reduced myeloid immunosuppression, and inhibition of primary and metastatic tumor growth. Because Parkin is epigenetically silenced in all human cancers, dual mode tumor suppression is a universal barrier against malignancy. Therefore, the hypothesis that Parkin tumor suppression bridges mitochondrial activation of interferon signaling to immune reprogramming of the microenvironment can be formulated and will constitute the focus of the present application. Three complementary and multidisciplinary specific aims will elucidate the role of Parkin innate immunity in tumor suppression. In the first specific aim, we will characterize how Parkin “primes” immunogenic cell death, elucidate the role of ER stress and autophagy in mitochondrial necroptosis, and map the cellular and biochemical requirements of DAMP release. The second specific aim will dissect the signaling requirements of Parkin activation of innate immunity with respect to cytosolic cGAS-STING and inflammasome activation, mitochondrial RIG-I-MAVS sensing and assembly of a STAT1-regulated ISGF3 transcriptional complex for T cell activation and dendritic cell function. In the third specific aim, we will test the impact of Parkin innate immunity in preclinical orthotopic and transgenic models of primary and metastatic tumor growth, in vivo, modulation of antitumor vaccination strategies, and differential sensitivity to conventional and immune therapies. The application builds on expansive preliminary data, a novel concept of dual mode Parkin tumor suppression, and the redirection of mitochondrial innate immunity, previously known in viral infections, for antitumor responses. The results may be practice changing. As proposed here, the elucidation of Parkin innate immunity will identify new strategies to restore an antitumor immune microenvironment, dampen myeloid immunosuppression and enable broader, more durable patient responses to conventional and immune therapy, including in late-stage disease.

NIH Research Projects · FY 2025 · 2024-07

Project Summary/Abstract This application focuses on advanced melanoma, a highly aggressive type of skin cancer that arises from the pigment-producing melanocytes of the body. Despite recent advances in molecularly targeted therapy and immunotherapeutic agents with impressive response rates, there remain patients who either do not respond to such therapies or who eventually relapse. The 5-year overall survival rate for metastatic melanoma is below 20%. Progressive dedifferentiation and phenotypic switching of melanoma cells under cellular stress are considered to be a major driver for both tumor progression and therapy resistance. However, the molecular mechanisms that govern this process, and their interplay with genetic lesions and the tumor microenvironment are poorly understood. TFEB is a member of the MiT/TFE family of transcription factors and master regulators of cell differentiation pathways. Our preliminary studies identified TFEB repression as a novel dependency of oncogene-driven melanoma progression. We showed that TFEB activation globally re-invigorates transcriptional differentiation of melanocytes, while its inactivation provokes phenotypic transition towards the invasive and drug-resistant states that is associated with aberrant TGF-b upregulation. Furthermore, transcriptomic and immune cell profiling of tumors from primary melanoma patients confirmed dampened TFEB expression and function that also correlates with tumors’ immune evasive microenvironment. These findings lead us to hypothesize that TFEB-mediated transcriptional reprogramming of melanoma cell differentiation states represents a key mechanism disabling melanoma progression, which also reshapes tumor immune microenvironment for enhanced anti-tumor immunity. We will test the hypothesis by (Aim 1) defining the molecular mechanisms of TFEB-mediated transcriptional reprogramming of melanoma cell plasticity and phenotype switching; and by (Aim 2) investigating in detail the in vivo impact of TFEB alteration in melanoma progression and immune evasion. Successful completion of this study will provide mechanistic insights into tumor cell-switching processes and hold promise for the development of novel therapeutic strategies to reverse this process for the prevention and elimination of tumor metastasis and recurrence.

NIH Research Projects · FY 2026 · 2024-06

PROJECT SUMMARY Given the inability of the immune response to consistently suppress HIV without antiretroviral therapy and the insufficient potency of current cure approaches, genetic engineering modalities may offer a potent alternative to intrinsic immunity. Chimeric antigen receptor (CAR) T cells have shown impressive efficacy in eliminating blood cell cancers in the clinic, and CAR T cells re-engineered to target HIV using the CD4 ectodomain represent a potent escape-resistant cellular therapy demonstrated to have enhanced cytolytic function. Our prior work has demonstrated that when engineered to express both the 4-1BB and CD28 costimulatory domains and protected from HIV infection, HIV-specific CD4 ectodomain CAR T cells can reduce acute viremia, prevent CD4+ T cell loss, and reduce viral burden in the tissues of HIV-infected humanized mice. However, the of reduction plasma viral loads was ultimately transient, suggesting that the potency of HIV-specific CAR T cells should be further optimized for clinical translation. We propose that low expression of HIV Envelope (Env) on HIV-infected cells yields targets with low antigen density. Our preliminary data demonstrates that the potency of CD4 ectodomain CARs is severely attenuated in instances of low antigen density. We reason that this necessitates the design of HIV-specific CAR T cells with enhanced antigen sensitivity. As the characteristics dictating antigen sensitivity are not yet fully described for CAR T cells, we generated a diverse panel of broadly neutralizing antibody-based CAR T cells, specifically to (1) determine the relationship between epitope accessibility, CAR avidity, and antigen sensitivity on HIV-specific CAR T cell potency and (2) identify orthogonal CAR-driven escape patterns informing optimal multi-specificity CAR products that restrict HIV escape in vivo. The ultimate goal is to generate an enhanced HIV-specific CAR T cell product with the ability to kill HIV-infected cells expressing low levels of antigen in order to improve the translational potential of a CAR T cell approach for a functional HIV cure.

NIH Research Projects · FY 2026 · 2024-06

Project Summary In EBV+ lymphomas viral protein expression is the oncogenic driver and progressive force. However, the mechanisms by which EBV infection alters B cell biology to contribute to the development of EBV+ lymphomas are not well understood. EBV infection of primary B lymphocytes results in cell activation, proliferation, and immortalization. These programs are induced by EBV mimicking the normal activation of B cells by antigens or T cells. Since LMP1 is needed for EBV to transform naïve B cells into immortalized B cells, considerable work has been attempted to pharmacologically target the LMP1-activated oncogenic pathways. However, in vivo attempts to target these pathways have failed to abolish EBV-driven oncogenesis completely. This lack of efficacy of the targeted approaches suggested to us that another, undiscovered or unappreciated, oncogenic mechanism downstream of LMP1 might be at work. This proposal aims to identify the mechanism by which EBV infection contributes to developing B cell malignancies. Our recent, published work showed that the EBV oncoprotein LMP1 activates the chromatin-associated factor PARP1 to epigenetically regulate genes that are important for sustaining cell proliferation. Expanding upon these findings, our preliminary data indicate that the reorganization of the 3D structure and function of chromatin in B cells is a novel and underappreciated mechanism targeted by EBV through LMP1 to alter gene expression and reprogramming in EBV infected cells. We discovered that: 1) expression of LMP1 alone is sufficient to alter the 3D structure of the B cell genome; 2) the EBV/LMP1-induced changes in chromatin structure are associated with PARP1 activity, and with occupancy by the chromatin organizers CTCF and YY1; 3) EBV activation of PARP1 requires ERK activation by LMP1; and 4) PARP inhibitors elicit cytotoxicity in EBV+ lymphomas in vivo. Based on these preliminary data and our previous work we hypothesize that in EBV+ B cells, LMP1 expression activates, via the ERK pathway, PARP1, promoting conformational changes in the host genome architecture that drive oncogenesis. We will also test the hypothesis that PARP1 is an attractive therapeutic target for EBV lymphomas.

NIH Research Projects · FY 2026 · 2024-05

ABSTRACT Microbial genes in gut and diseased tissues were recently linked with progression and outcomes of different human diseases, including cancer and immune diseases. Deciphering the pathogenic roles of different microbial genes can improve diagnosis, prognostication, and treatment of human diseases. Yet, current methods for RNA and shotgun metagenomic sequencing focus on microbial species, and do not allow a systematic detection or quantification of microbial genes. Therefore, current methods for disease prognostication and biomarker discovery are unable to consider microbial genes that influence human diseases. Our overarching hypothesis is that there are unknown associations between human diseases and the microbial genes in diseased tissues and in the gut. Our long-term goal is to unravel these associations using novel computational approaches that will allow detection and quantification of microbial genes in diseases. In Aim 1 we will develop methods that harness RNA sequencing to detect microbial gene expression in diseased tissues. This will allow microbial biomarker discovery and provide a comprehensive database of the microbial genes that are expressed in various human tissues and conditions. In Aim 2 we will develop methods to quantify gut microbial gene capacity in human diseases. This will allow identification of gut microbial proteins, peptides, and domains that are important in human diseases, to ultimately yield new diagnostic and treatment strategies based on gut microbiomes. Overall, this project will provide innovative methods to allow detection and quantification of microbial genes from abundantly used sequencing technologies. We will establish user friendly software and databases, allowing new discoveries with existing sequencing platforms. We expect that the methods developed through this project will be extensively adopted by the relevant research communities, improving our understanding of the roles of microbes in human diseases and ultimately allowing the development of new disease detection and intervention strategies based on microbial genes.

NIH Research Projects · FY 2026 · 2024-04

PROJECT SUMMARY Most mammalian genes harbor multiple cleavage and polyadenylation sites, or PASs, across the gene body, resulting in mRNA isoforms with different coding sequences (CDS) and/or 3’ untranslated regions (3’UTRs). Alternative cleavage and polyadenylation (APA) is an important layer of gene regulation, affecting gene expression levels, protein diversity, and mRNA metabolism. The APA isoform expression varies across cell types and is dynamically regulated in a growing number of cell conditions, such as cell proliferation and differentiation, change of metabolic states, and cellular stress. Our lab employs interdisciplinary approaches to study APA, involving molecular biology, functional genomics, and computational biology. Our long-term goal is to understand the functional genomics of APA across species as well as molecular mechanisms and cellular consequences of gene regulation by APA in different cell contexts and pathological conditions. In the next five years, we plan to address a few key gaps in the field: First, we plan to examine regulatory rules governing intronic polyadenylation that leads to early termination of transcription. Second, we will examine the role of 3’UTR isoform regulation in cell metabolic reprogramming, such as growth and autophagy. Third, we will investigate the mechanisms and consequences of the unique APA isoform profile in secretory cells.

NIH Research Projects · FY 2026 · 2024-03

Adoptive cellular immunotherapy has revolutionized cancer treatment, with engineered T cells achieving remarkable success in hematologic malignancies. However, challenges such as relapse, limited efficacy in solid tumors, and the immunosuppressive tumor microenvironment (TME) remain. This proposal addresses critical gaps in the field by developing two innovative strategies: (1) Intrinsic regulations, through self-regulatable therapeutic T cells that dynamically adjust chimeric antigen receptor (CAR) expression in response to antigen stimulation, enabling precise control over T cell activity and minimizing toxicity; and (2) extrinsic regulations, developing novel protein chimeras that exploit endogenous cellular machinery to degrade immunosuppressive proteins in the TME, restoring anti-tumor immunity. By integrating these approaches, this research aims to transform cancer immunotherapy, offering safer and more effective treatments for solid tumors and other challenging malignancies. The proposed work has the potential to improve the safety and efficacy of CAR-T cell therapies, provide new insights into receptor dynamics and immune modulation, and align with the NCI’s mission to advance innovative cancer treatments.

NIH Research Projects · FY 2026 · 2024-03

PROJECT SUMMARY The treatment of pancreatic ductal adenocarcinoma (PDAC) remains a major hurdle with 5-year survival of 12%. PDAC resistance to chemotherapy and immunotherapy is thought to arise from the immunosuppressive tumor microenvironment (TME), characterized by a fibrotic stroma and high infiltrates of immunosuppressive cells, including tumor associated macrophages (TAMs). TAMs block anti-tumor effector T cell function and trigger exhaustion. It has been postulated that reprogramming TAMs to an immunostimulatory phenotype could improve therapy response. Recent studies suggest that metabolites originating from the gut microbiome influence phenotype of immune cells, including TAMs. However, little is understood about such metabolites, including their identity, the signaling pathways by which they alter TAM phenotype, and whether these metabolites influence cancer development. We are identifying microbial metabolites that modulate TAMs in the PDAC TME. Using unbiased, global, LC-MS/MS metabolomic screens, we found a gut microbe-derived metabolite, trimethylamine N-oxide (TMAO) that induced significant anti-tumor effects in the PDAC TME. Specifically, delivery of TMAO intraperitoneally, or by supplementing diet with the TMAO precursor choline, to PDAC-bearing mice reduced tumor growth and was associated with an immunostimulatory TAM phenotype and activated effector T cell response in the TME. The immunostimulatory macrophage phenotype was due to a direct effect of TMAO and the TMAO-conditioned macrophages could activate CD8+ T cells. Combining TMAO with immune checkpoint blockade reduced tumor burden and improved anti-tumor immune responses. These data support our hypothesis that the diet and microbiome-derived metabolites shape anti-tumor immunity and treatment response in PDAC. Aim 1 will characterize dietary and microbial sources of TMAO for anti-tumor responses in PDAC. We will characterize the (i) dietary (l-carnitine or betaine), and (ii) the microbiome (e.g., Enterococcus asini, engineered E. coliCutC/D) sources of TMAO for their anti-tumor effects. Aim 2 will test the hypothesis that TMAO induces its anti-tumor effects by potentiating the type I IFN and/or STING signaling in TAMs. We will determine (i) the requirement of type-I IFN and/or STING specifically in macrophages for the anti-tumor effects, and (ii) the impact of TMAO on the transcriptional activity of type-I IFN responsive STAT1 and/or STAT3. Aim 3 will test the efficacy of TMAO to improve treatment response in pre-clinical models of PDAC. We will evaluate the translational relevance of TMAO using (i) a genetically engineered mouse model of PDAC and patient derived PDAC organoid cultures, and (ii) a treatment strategy combining TMAO with STING agonists. Our studies will characterize the sources, mechanism of action, and translational relevance of a novel, minimally understood, high impact microbial metabolite, TMAO, in boosting anti-tumor immune responses in the PDAC TME and rendering PDAC responsive to chemo-immunotherapy. In the longer term, this work may lay the groundwork for new diet/microbiome-based treatments for PDAC.

NIH Research Projects · FY 2026 · 2024-03

Project Summary The goal of this proposal is to uncover the mechanistic connection between R-loops and gene expression through effects on genome architecture to understand how R-loop deregulation can contribute to neurodevelopmental disorders. R-loops are poorly understood RNA-containing chromatin structures that accumulate in, and contribute to the etiology of, several neurodevelopmental disorders. This includes Activity-dependent neuroprotective protein (ADNP) syndrome, also known as Helsmoortel-Van Der Aa syndrome, which is a rare condition in children that exhibit signs of autism. At present, how R-loops contribute to neurological disorders like ADNP syndrome is unclear. Accumulated R loops lead to an increased DNA damage response and can also alter transcription of neighboring genes. However, the mechanistic basis for R loops-mediated changes in gene expression are unknown and the functional relevance of this process to disorders like ADNP syndrome is unexplored. We have made the surprising finding that R-loops are highly enriched at a subset of binding sites for CTCF. Preliminary data show that R-loops strengthen CTCF interactions with chromatin. We found that conditions leading to loss or gain of R-loops can decrease or increase CTCF recruitment, respectively, and affect long- range genome interactions. We recently demonstrated that the activity-dependent neuroprotective protein (ADNP), a critical protein for brain development, is a site-specific R-loop resolver. ADNP heterozygous missense or frameshift mutations cause ADNP syndrome, a severe neurodevelopmental disorder. Human induced pluripotent stem cells (hiPSCs) derived from patients with ADNP syndrome show increased R-loops and CTCF accumulation at ADNP binding sites. We find that ADNP binding sites are enriched for sequences that are recognized by the genome architectural protein YY1, which has important functions in regulating enhancer- promoter interactions especially in the neural lineage. ADNP and YY1 co-localizes at active enhancers. Our preliminary data also identified changes in DNA methylation in ADNP syndrome hiPSCs that can potentially impact CTCF and YY1 binding to cause pathogenic genome misfolding and aberrant neural gene expression in ADNP syndrome. We hypothesize that R-loops have a regulatory function, and that they target CTCF and YY1 to specific genomic sites during neurodifferentiation. We posit that they may be critical for long-range genome interactions that reinforce neural lineage specific gene expression programs. We propose to decipher the impact of R-loop deregulation on genome organization and gene expression in the neural lineage through the lens of ADNP syndrome. In Aim 1, we will evaluate R-loop mediated regulation of CTCF and YY1 localization during neurodifferentiation. In Aim 2, we will examine the epigenetic consequences of distinct ADNP syndrome mutations and their impact on genome regulatory interactions.

NIH Research Projects · FY 2025 · 2023-07

PROJECT SUMMARY The Epstein-Barr Virus (EBV) is responsible for approximately 200,000 new cancer cases each year worldwide. Among these, EBV+ Diffuse Large B-Cell Lymphoma (DLBCL) is an emergent global cancer threat in patients without overt immunosuppression, irrespective of age, and represents a growing unmet medical need. New therapeutic approaches are needed to treat EBV+ DLBCL. Only one viral-encoded protein, EBNA1, is consistently expressed in all known EBV-associated malignancies and is a validated target for inhibition of EBV- dependent transformation and carcinogenesis. Investigators at the Wistar Institute have developed VK-2019, a first-in-class EBNA1 inhibitor as a therapeutic agent, selecting it from over 2500 candidate inhibitor compounds during the hit-to-lead and lead optimization phases. VK-2019 meets or exceeds industry-accepted criteria for potency, selectivity, metabolic stability, drug suitability, drug safety, toxicology and bioavailability. We anticipated that VK-2019 would have a favorable safety profile because there are no human orthologs of EBNA1. Based on preclinical evidence and a first-in-human Phase I clinical study in patients with advanced nasopharyngeal carcinoma (NPC), VK-2019 met all safety, tolerability and pharmacokinetic endpoints with few documented adverse events (AEs) or Severe Adverse Events (SAEs). In this early study, we observed stable disease in more than a third and a significant decrease in EBV plasma levels, a known biomarker of NPC progression, in more than half of the patients, that correlated with pharmacokinetic exposure. We believe that data from this Phase I study are encouraging in terms of both on target effect and clinical benefit, and supports a follow-on proof-of-concept study in patients with EBV-positive DLBCL. The purpose of this grant is to fund a phase Ib clinical trial of daily oral administration of VK-2019 to (1) further confirm the safety profile and determine any dose-limiting toxicities (DLT) in advanced EBV+ DLBCL patient populations; (2) understand the effects of treatment on EBV-specific biomarkers, including EBV and cellular gene expression and (3) study the effects of treatment on the tumor microenvironment and immune response. The clinical trial infrastructure necessary for the conduct of this study is already in place at the Sidney Kimmel Cancer Center at Thomas Jefferson University. This clinical trial will provide critical information on the safety, tolerability, and preliminary efficacy of VK-2019 in a EBV+ DLBCL patient population. This application brings together basic and translational investigators to understand whether EBNA1 inhibitors can be a therapeutic option for latent EBV infection and cancer and examines the mechanism of action.

NIH Research Projects · FY 2026 · 2023-05

OVERALL – PROJECT SUMMARY The overall aim of this new Program Project (P01) proposal is to generate highly collaborative and integrated basic and translational research on the urgent, unmet medical need of epithelial cancers caused by Epstein-Barr Virus (EBV). EBV latent infection is causally linked to over 200,000 new cancer cases per year. EBV epithelial cancers represent over 75% of all EBV cancers with highest mortality rates and treatment failures. EBV- associated gastric carcinoma (EBVaGC) and nasopharyngeal carcinoma (NPC) have many similarities with respect to viral latency and oncogenic transformation. The mechanisms through which EBV contributes to these epithelial cancers remains elusive, and to date, there are no viral-specific therapies that are FDA approved to treat cancers infected with EBV. We have assembled a team of EBV investigators with expertise in complementary aspects of tumor virology and cancer biology, with specific areas of interest in viral genetics, epigenetics, metabolism, drug discovery, and models of EBV carcinogenesis. The Program team will collaborate in a coordinated strategy to identify key viral and cellular vulnerabilities in EBV epithelial cancers that can be targeted for therapeutic intervention. The Program will test the central hypothesis that EBV cancers arise in the context of somatic mutations in metabolic and epigenetic pathways that alter EBV latency and oncogenicity, and how this viral-host co-dependency provides therapeutic opportunities. To achieve these goals for the Program we propose three Projects and three scientific Cores to address the following: (1) Determine how EBV establishes a latent and oncogenic infection in epithelial cancer cells (2) Determine how EBV latent infection drives epigenetic and metabolic shifts including the formation of CpG island methylator phenotype (CIMP) to promote epithelial cell oncogenesis. (3) Leverage mechanistic insights to develop new therapeutic strategies to treat EBV epithelial cancers. The 3 Projects focus broadly on EBV latency and epigenome (Lieberman), PARP, NAD and DNA damage metabolism (Tempera), and DNA methylation and methionine metabolism (Gewurz). The 3 scientific Cores support bioinformatics, drug discovery, and models of EBV cancers to support each of the 3 Projects as research enhancers. Together, this team and Program will investigate key features of EBV cancer mechanisms, build new tools to study EBV cancers, and develop new therapeutic strategies that are viral-specific and precision-based medicine.