University Of Pennsylvania

NIH Research Projects · FY 2026 · 2022-05

Glutaminolysis, the cellular catabolism of glutamine, is an important metabolic pathway for aggressive and treatment-resistant cancers, including many triple-negative breast cancers (TNBCs). It is well accepted that glutamate produced from glutamine by mitochondrial glutaminase (GLS) fuels the TAC cycle, which provides energy and precursors for biosynthesis. Emerging data have revealed a less recognized but important contribution of glutaminolysis in mediating oxidative stress introduced internally by active growth of aggressive cancer cells and externally by treatments including chemotherapy and immunotherapy. Targeting inhibitors of GLS to block glutaminolysis is a therapeutic strategy that has been tested in clinical trials of breast and other cancers with acceptable toxicity, but limited efficacy, owing in good part to a lack of clinical markers to guide patient selection and assess target impact. Preliminary data from our lab have shown that dual targeting of GLS and the plasma membrane glutamate transporter, xCT (SLC7A11), resulted in dramatic sensitization of resistant TNBC to chemotherapy. We propose three aims based upon an overall theme to develop a kinetic framework for non-metabolized amino acid analog PET tracers to measure cellular pool sizes as an indicator of catabolism and cellular transport. Specifically, we will (1) validate quantitative markers for cellular glutamine pool size from dynamic [18F]fluciclovine PET; (2) develop and validate markers for cytosolic glutamate pool size and transport using 4-(3-[18F]fluoropropyl)-L- glutamic acid ([18F]FSPG) PET, and (3) determine the utility of combined [18F]fluciclovine and [18F]FSPG PET for predicting and measuring response to dual-targeted treatment designed to sensitize TNBC to chemotherapy. As part of this work, we will address mechanistic questions regarding cytosolic glutamate transport from mitochondrial pools and to/from extracellular fluid to guide the interpretation of PET tracer kinetics. We will also test approaches to target TNBC metabolic vulnerabilities, specifically the dependence glutamine metabolism and glutamate transport, guided by the PET methods we develop and validate in our pre-clinical TNBC models. The proposed work will lead to a deeper understanding of the mutual engagement between glutaminolysis and redox homeostasis of cancer cells and will yield quantitative imaging methodologies ready to translate to the clinic.

NIH Research Projects · FY 2025 · 2022-05

PROJECT SUMMARY The prevalence of cirrhosis and end-stage liver disease (ESLD) in the U.S. has nearly doubled over the past two decades. Liver transplant (LT) is the only life-saving option for ESLD and is projected to increase 23% by 2040. LT requires considerable healthcare resources, costing the U.S. health system $26.7 billion annually. It is increasingly performed among older adults; from 2006 to 2018, adults 65 and older were the most rapidly growing age group on transplant waitlists. By 2019, 70% of LTRs were adults 50 and older. With advances in immunosuppression (IS) and surgical techniques, average 5-year post-LT survival now exceeds 70%. However, maintaining optimal, long-term liver graft function and overall health are contingent upon proper adherence to prescribed treatment. This can be difficult, as liver transplant recipients (LTRs) contend with high regimen complexity, taking on average 11 new medications that have side effects and require frequent dosage changes. Many LTRs have lingering cognitive impairments due to pre-transplant hepatic encephalopathy, or other psychosocial, financial, or cultural factors that all can affect the ability to adhere to treatment. As a result, a third of LTRs demonstrate inadequate adherence to IS regimens. Poor adherence is a leading cause of liver graft rejection, liver failure, poorly controlled medical comorbidities and subsequent mortality. Similarly, inadequate medication knowledge and unintentional misuse are associated with post-transplant hospitalizations. Despite the negative and costly consequences of inadequate adherence, very few prospective interventions have been developed and tested to optimize medication-taking behaviors and subsequent outcomes post-LT. However, there are unique considerations within the context of LT that can be leveraged by interventions: 1) LTRs remain indefinitely connected to transplant centers for follow-up with a range of available resources; nurse coordinators, pharmacists, psychological and social services; 2) to be eligible for LT, patients must have involved caregivers to provide post-LT support (yet no interventions to date have involved caregivers). Our primary aim is to test the effectiveness of a technology-enabled intervention (TEST trial) to improve treatment adherence and optimize patient-centered and clinical outcomes among a cohort of 360 de novo LTRs at 3 diverse transplant programs in Philadelphia, PA, Chicago, IL, and Miami, FL. Our intervention leverages existing transplant center resources, caregiver support, and widely available patient-centered mobile health tools to remotely monitor LTRs' adherence behaviors, `phenotype' adherence concerns, and tailor transplant center responses to those concerns. We additionally plan to assess intervention fidelity, enabling us to evaluate patient, caregiver, provider and health-system barriers to optimal implementation to enhance scalability. Finally, we will determine the costs and cost-effectiveness of the intervention from a transplant center and healthcare sector perspective. The TEST trial will enable the timely identification of the presence and nature of LTR adherence barriers and mobilize transplant centers and caregivers to deploy tailored solutions by leveraging existing resources to optimize health.

NIH Research Projects · FY 2026 · 2022-05

ABSTRACT Colorectal cancer (CRC), the second leading cause of death in older adults in 2019, was diagnosed in 145,600 patients and was responsible for 51,020 deaths. In the absence of metastatic disease, surgery is the standard of care for more than 90% of CRC patients. Insight from existing literature and our preliminary studies suggest that the most essential surgical disparities in CRC are related to surgical risk and strong hospital- associated differences in mortality and morbidity. Significant variation in CRC surgical outcomes exists across hospitals (e.g. mortality rates 0.6%-14.7%) with known disparities adversely affecting black patients. Black patients have lower surgical utilization rates, worse surgical outcomes, and lower survival rates compared to White patients. Black patients are more likely to use lower quality, lower volume hospitals for surgery, even when a higher quality choice can be found closer to home. Significantly worse outcomes are also observed in rural settings. Access to high quality hospitals is a critical barrier to optimal surgical care. Data to drive hospital selection is limited. Our preliminary studies demonstrate that most Black patients (86%) have a higher quality hospital located within close proximity of their home and data driven referrals have the potential to reduce disparities by >30% and improve outcomes. Existing risk stratification tools to assist in the hospital selection process lack the requisite combination of factors to facilitate rational decision-making including: 1) disease specificity, 2) attention to complex patient-provider interactions, 3) information on hospital quality, and 4) comparative statistics. Our preliminary data suggest that accurate risk prediction can be performed that meet these criteria. In the proposed study, we will refine the personalized prediction models, scale them to the national level, and develop the tools to make statistical comparisons possible. As disparities drive unnecessary health care expenditures, we demonstrate the gains in Societal Welfare of data driven referrals using counterfactual simulation. Further, we will use scenario testing to simulate the effects of data driven referrals on the willingness of referring providers to trade-off convenience and reputation for enhanced quality. This information is critical to drive policy reform to advance surgical care. Our goal is to reduce disparities by referring older, black CRC patients to higher quality hospitals by 1) developing personalized risk models to differentiate across hospitals (or surgeons), 2) providing evidence to inform policies designed to incentivize data driven referrals, and 3) setting strategies to promote data driven referrals for CRC. This pioneering work will provide 1) new methods of risk stratification, 2) an estimate of the Societal Welfare benefits of data driven referrals for policy makers when designing new policies to minimize surgical disparities and 3) new knowledge on physician preferences to inform interventions to promote adoption of data driven referrals. This work will serve as a template for subsequent efforts to extend data driven referrals across all surgically treated solid organ malignancies.

NIH Research Projects · FY 2026 · 2022-05

ABSTRACT Optogenetics is a revolutionary technique in neuroscience. By combining light-sensitive proteins with intracranial light delivery, optogenetics offers unprecedented, cell-type specific control over neuronal activity. The technique has become the dominant approach for studying neural circuits in small animal models such as mice and flies. Unfortunately, optogenetics has so far failed to have a major impact on research using larger animals more similar to humans, such as macaque monkeys, undermining its translational potential for human patients. We conducted a world-wide Open Science initiative to identify the challenges remaining to be solved in primate optogenetics (Tremblay et al. Neuron, 2020). We identified the sheer size of the macaque monkey brain, which is 200 times bigger than the mouse brain, as well as its immune system, as the main challenges for both gene expression and light delivery. Our multidisciplinary team of investigators will overcome these obstacles by developing and optimizing three new technologies: 1) large-scale, safe delivery of ultra-sensitive opsins using gene therapy techniques; 2) chronically-implantable, ultra-thin, flexible, biocompatible LED arrays; and 3) implantable, battery-powered LED drivers for wireless control during unrestrained, naturalistic behavior. This approach will allow precise control of large volumes of the primate brain with cell-type specificity and millisecond resolution in monkeys free of physical restraint, thus permitting causal dissection of the neural circuits mediating natural behavior relevant for understanding and treating human brain disorders. This technology platform could be directly applied as a cell-type-specific optogenetic therapy for humans suffering from neurological disorders that affect specific neural populations, such as focal epilepsy.

NIH Research Projects · FY 2025 · 2022-05

Nurses' daily encounters with ethical challenges in providing appropriate care—situations that may go unresolved or are resolved unsatisfactorily—may lead to moral distress (i.e., the inability to carry out what is believed to be an ethically appropriate action because of internal or external constraints). As a result, many experience anxiety, depression, suicide risk, and other health-related problems. Little systematic research has focused on factors that reduce or exacerbate moral distress—and, critically, how nurses' moral distress affects nurse, patient, and organizational outcomes. This application builds upon our successful scientific program of empirical bioethics research and uses a mixed-methods design to address three complementary objectives, by surveying 20,652 practicing registered nurses (RNs) across four geographically diverse states (California, Pennsylvania, Maryland, and Massachusetts) and innovatively linking these survey data to publicly available patient outcome data sources. Thus, our first aim is to gather detailed data on nurses' perspectives on their workplace challenges by identifying and assessing (via quantitative and qualitative methods) individual, work environment, and ethical factors that contribute to moral distress and ethical confidence (i.e., self-confidence about making ethics-related decisions in clinical practice). Our second aim is to develop (via cluster analysis) a typology of practicing nurses, detailing similarities/differences across the personal, workplace, and ethics issues that most impact moral distress, ethical confidence, health, and wellbeing; and from this typology, recommend strategies for reducing moral distress , and create (via perceptual mapping) 3-dimensional perceptual models showing how each type of nurse conceptualizes the relationships among these factors and use these maps to assess safety culture as experienced by the different types of nurses, which will permit even more specific recommendations about moral stress reduction. Our third aim is to examine how the relationships identified in aim 1 are associated with outcomes of hospitalized patients, including patient perceptions of care, inpatient mortality, hospital-acquired conditions, and excess days in acute care within 30 days of hospital discharge as well as nurses' well-being, patient safety grade and intention to leave. The findings will provide a clearer picture of factors contributing to nurses' moral distress and to their ethical confidence and provide evidence on how moral distress and ethical confidence affect patient safety and quality outcomes. Healthcare is increasingly provided in complex organizations where resolving ambiguous ethical issues may deplete the best of clinicians. Our findings will inform development of behavioral interventions, structural/operational workplace changes, and message campaigns that reduce moral distress in existing and future cohorts of nurses and improve the safety culture in which they work.

NIH Research Projects · FY 2026 · 2022-05

Title: B cell lineage directed rational vaccine strategies based on CAP256SU Env-Ab co-evolution Project Summary/Abstract Despite decades of effort, no HIV-1 vaccine has successfully elicited protective broadly neutralizing antibodies (bnAbs) by immunization. This failure is largely due to the virus’s evolved mechanisms of immune evasion, including glycan shielding, epitope variability, conformational masking, and trimer exclusion. As a result, bnAbs must overcome substantial hurdles, such as requiring long HCDR3s and extensive somatic hypermutation— features that are rare or difficult to induce by vaccination. Among known bnAbs, those targeting the V2-apex epitope are among the earliest and most frequently elicited in natural HIV-1 infection and in SHIV-infected rhesus macaques (RMs), and they require relatively limited affinity maturation. Building on these unique features and our extensive prior work on V2-apex–directed immunogen design, we have developed CAP256.SU-GT1, a CAP256.SU HIV-1 Envelope lineage-based germline-targeting (GT), native-like trimer immunogen. In our preliminary studies, this immunogen—delivered as a soluble trimer or nanoparticle—successfully primed rare V2-apex bnAb precursors and elicited cross-neutralizing responses in outbred RMs. However, the breadth and durability of these responses remain variable. We hypothesize that optimizing the affinity of CAP256.SU GT immunogens for diverse V2-apex bnAb rare precursors will enhance B cell activation, germinal center responses, and memory/plasma cell formation—laying the foundation for a successful prime-boost vaccine strategy. Our approach integrates rational structure-guided and directed evolution-based immunogen design, novel delivery platforms, and iterative testing in the RM model to induce durable bnAb responses. Aim 1 will optimize the CAP256.SU GT trimer or nanoparticle immunogen using reverse protein engineering to enhance in vivo priming of diverse rhesus and human V2-apex bnAb precursors. Aim 2 will evaluate the priming efficacy of optimized CAP256.SU GT-immunogens in outbred RMs using conventional and novel vaccine delivery platforms to select the most effective format for priming bnAb precursors and for robust memory B cell induction. Aim 3 will develop and test CAP256 Env lineage-based and heterologous HIV-1 trimer boost strategies to expand neutralization breadth and induce durable serum V2-apex bnAb responses. A successful demonstration of enhanced and durable bnAb induction in rhesus macaques will represent a significant step toward a protective HIV-1 vaccine, with strong translational potential for human clinical evaluation.

NIH Research Projects · FY 2026 · 2022-05

PROJECT SUMMARY This study will provide novel insights on cognitive trajectories among new liver transplant recipients, and the impact of cognitive function on self-management, health behaviors, and patient outcomes. The prevalence of cirrhosis and end-stage liver disease (ESLD) in North America is estimated at up to 1,000 per 100,000 population and has nearly doubled over the past two decades with baby boomers (born 1945-1965) accounting for half of cases. From 2009 to 2016, there has been a 65% increase in cirrhosis mortality in the U.S. Decompensated cirrhosis has a 5-year survival of only 34-56% with liver transplantation (LT) as the only life- saving option. By 2033, LT demand will increase by 23% and per-patient LT-associated costs will rise from $1.4 to $2.1 million, resulting in a total of $26.7 billion in transplant-related medical expenses over a 10-year time horizon. To maximize the benefits of LT, liver transplant recipients (LTRs) must have strong self-management skills to navigate health systems, adhere to clinical monitoring, and take complex, multi-drug regimens. All of these tasks require formidable cognitive abilities for active learning and problem solving. LTRs are at higher risk for poorer cognition due to high prevalence of pre-transplant cognitive impairment (hepatic encephalopathy), multiple chronic conditions, alcohol use, and increasing age. The cognitive status of LTRs over time could affect self-management and transplant outcomes, yet these issues have not been thoroughly investigated. Our proposed cohort study (`LivCog') will longitudinally characterize cognitive performance using the NIH Toolbox among 450 new LTRs from 3 diverse transplant centers, beginning at transplant waitlisting and then at 1, 4, 12, and 24 months post-LT. We will also serially assess self-management skills, physical function, health behaviors, patient-reported and clinical outcomes. Potentially modifiable post-transplant targets will be investigated, including caregiver support, physical activity, sleep, and treatment adherence to understand causal pathways that could inform future health system responses. Our specific aims are to: 1) assess pre to post-LT cognitive trajectories and identify risk factors for persistent cognitive impairment, 2) evaluate associations between cognitive function and self-management skills, health behaviors, functional health status, and post- transplant outcomes, and 3) investigate potential mediators and moderators of associations between cognitive function and post-transplant outcomes. Our proposed LivCog study will fill critical gaps in understanding cognitive recovery and function, risk factors and consequences of cognitive impairment among LTRs. Findings will directly inform future interventions to improve post-transplant outcomes.

NIH Research Projects · FY 2025 · 2022-05

Summary/Abstract The trafficking of leukocytes from the blood stream to the sites of inflammation to find infectious agents is a hallmark of the innate immune response. Neutrophils, myeloid leukocytes of the innate immune response, are the so-called “first responders” and will rapidly traffic to the sites of inflammatory insult. Trafficking is initiated by the leukocyte adhesion cascade, a well-characterized, stepwise sequence of events in which blood borne immune cells tether, roll, firmly arrest, and migrate on the endothelium and then enter tissues to perform immune cell functions. These events all occur within blood vessels, normally post-capillary venules, where cells encounter high shear rates while interacting with and transmigrating through the endothelium. Recently, an interesting phenomenon wherein certain cells of hematopoietic origin – such as lymphocytes (T-cells and B-cells) as well as hematopoietic stem and progenitor cells (HSPCs) – will crawl upstream, against the direction of flow, after arrest on surfaces that present the ligand intercellular adhesion molecule-1 (ICAM-1). Upstream migration is mediated by the β2 integrin, αLβ2, also known as Lymphocyte Function-associated Antigen-1 (LFA-1), which binds to ICAM-1. Neutrophils express LFA-1, as well as an additional receptor for ICAM-1, Macrophage-1 Antigen (Mac-1), which is upregulated when neutrophils are activated. Our laboratory hypothesized that neutrophils could also be induced to crawl upstream on ICAM-1 if Mac-1 was disabled or blocked, allowing LFA-1/ICAM-1 interactions to dominate. Our preliminary results show that by either blocking MAC-1 (with an antibody) or by genetically manipulating Mac-1 (using CRIPSR- Cas9 deletion), neutrophils can migrate upstream on ICAM-1 surfaces, setting the premise for this application. Here, we propose to determine the critical signals which govern the upstream migration of neutrophils, how neutrophils generate force when migrating upstream, and identify the physiological implications of upstream neutrophil migration. In Aim 1 we will determine the key molecular signals involved in upstream migration in neutrophils, by first analyzing the differential signaling that occurs on ICAM-1 vs. VCAM-1 surfaces and then identify and delete the signals that operate downstream of LFA-1 and Mac-1 using CRISPR editing. Specifically, we will focus on deleting the signals downstream of Mac-1/ICAM-1 binding to determine the signals that inhibit neutrophil upstream migration. In Aim 2, we will measure and quantify the forces generated by neutrophils engineered to migrate upstream using traction force microscopy. Using the knockouts generated in aim 1, we will measure the differences in the spatial arrangement and magnitude of force generation between upstream and downstream crawling neutrophils. Finally, in Aim 3 we will determine the physiological relevance of upstream migration in neutrophils by determining the change in time needed for altered to transmigrate in vitro on endothelium and the differential trafficking of disabled neutrophils into the peritoneal cavity in a murine model of acute inflammation.

NIH Research Projects · FY 2026 · 2022-05

SUMMARY Arterial spin labeled (ASL) perfusion MRI provides noninvasive quantification of tissue blood flow in physiological units of ml/100g/min using magnetic labeling of blood water as an endogenous diffusible flow tracer, and is one of the few MRI parameters whose biological basis is known. ASL MRI has primarily been used in the brain to measure cerebral blood flow (CBF), a key physiological parameter that serves a biomarker of cerebrovascular integrity and regional brain function with a broad range of applications in basic and clinical neuroscience research and in clinical care. ASL MRI was originally conceived by our laboratory at the University of Pennsylvania, and we have been responsible for demonstrating many of its technical advances and applications in biomedical research. Although ASL MRI has been translated to clinical use, commercial ASL MRI technologies have failed to keep up with research progress. In response to the special funding mechanism: PAR-18-530, this Academic Industrial Partnership project will provide dedicated resources to further develop, maintain, and deliver state-of-the-art ASL MRI acquisition and processing technologies for clinical research on the Siemens MRI platform, which is the most widely used MRI platform in neuroscience. An Academic Industrial Partnership is needed because market forces for commercial MRI technologies have been insufficient to drive the development of state-of-the-art ASL MRI capabilities in product sequences, yet close collaboration between academia and industry are required to deliver a streamlined capability to users. The resulting technologies will be disseminated free of charge to research sites through a new code exchange platform developed by Siemens. While a major innovation will be the delivery of a free ASL MRI software package featuring state-of-the-art approaches to maximize sensitivity, spatial and temporal resolution, and robustness to artifacts to meet evolving research and clinical requirements for noninvasive quantification of regional cerebral blood flow, next-generation approaches leveraging deep machine learning and other improved computing hardware and algorithms are also proposed to achieve higher spatial and temporal resolution, faster online image reconstructions, and improved robustness to artifacts than are currently possible. The proposed alliance will leverage the interdisciplinary expertise of the investigative team to provide a reliable, reproducible, flexible and user friendly technology for quantifying a key parameter of brain health and function that also has numerous clinical applications, including the evaluation of brain tumors and other organ systems. The feasibility of the proposed work is supported by our preliminary data and track record of ASL MRI technology development and dissemination.

NIH Research Projects · FY 2026 · 2022-05

PROJECT SUMMARY / ABSTRACT Traumatic brain injury (TBI) is common and is associated with significant morbidity and mortality. The sequelae from TBI can be long-lasting, and multiple studies have reported an increased rate of cognitive decline and higher risk of dementia among persons with TBI. However, the mechanisms linking TBI to dementia remain poorly understood, although vascular dysfunction, neuroinflammation, and aggregation of proteins have been proposed. This gap in knowledge was recently highlighted in the 2020 Lancet Commission on Dementia, which added TBI as one of twelve potentially modifiable risk factors for dementia, and in the 2019 NINDS Alzheimer’s Disease-Related Dementias (ADRD) Summit, which formally recommended further study into the role of TBI in dementia, including an emphasis on studying mechanism and developing TBI-AD/ADRD-related biomarkers. To directly address this research need, this application builds on my prior NINDS R25 award and proposes to use epidemiology, clinical studies, and vascular-related blood-based and neuroimaging biomarkers to investigate vascular injury and subsequent vascular dysfunction as mechanisms linking TBI and dementia. The central hypothesis of this proposal is that TBI is associated with cognitive decline and dementia risk in part via vasculopathy-mediated pathways which accelerate neurodegeneration for years post-TBI. This proposal will leverage existing data from 2 ongoing prospective cohort studies (Aims 1 and 2) and new data from a prospectively recruited cohort (Aim 3). The aims of this study are: 1) to determine if acute vascular injury is associated with poor short-term TBI-related cognitive outcomes in the trauma center-based Transforming Research and Clinical Knowledge in TBI (TRACK-TBI) Study, 2) to investigate if chronic vascular dysfunction mediates associations of TBI with poor long-term cognitive outcomes in the community-based Atherosclerosis Risk in Communities (ARIC) Study, and 3) to evaluate if the trajectory of post-TBI vascular dysfunction is associated with short-term cognitive outcomes in a prospectively recruited cohort nested within ongoing studies at the University of Pennsylvania Trauma Center. The overall objective of this proposal is to use multi-modal biomarkers of vasculopathy to investigate mechanisms linking TBI and neurocognitive outcomes and to identify time-periods during which future interventions may be effective at preventing TBI-related dementia. In addition to the proposed research, this project will provide me with critical gap-based training, including in the design, conduct, and implementation of clinical-epidemiologic studies and in the use of biomarkers as a method to investigate disease mechanisms linking TBI to outcomes in clinical-epidemiologic studies. This training will enable me to build an independent research program focused on vascular health in TBI with the goal of elucidating disease mechanisms and characterizing TBI outcomes using well-designed prospective clinical- epidemiologic studies of individuals with TBI. Completion of the proposed study will be facilitated by an institutional environment that prioritizes collaboration and provides exemplary research and career support.

NIH Research Projects · FY 2026 · 2022-05

Abstract Alzheimer’s Disease (AD) neuropathology is largely driven by two pathological AD proteins, tau, and ß-amyloid (Aß), both of which induce neuronal hyperexcitability and are thought to play a role in the high comorbidity between AD and epilepsy. Tau load and its regional brain distribution correlate more closely with cognitive decline than amyloid plaque deposition in both AD and epilepsy patients, making tau an attractive target for disease modification in both conditions. AD patients have an increased incidence of epilepsy compared to non-AD patients, and we recently showed that kindled seizures can exacerbate amyloid pathology, mediated by the mammalian target of rapamycin complex 1 (mTORC1) activation in an AD mouse model. To address how neuronal hyperactivity can increase AD pathology, we propose to use a tau seeding approach using two novel AD mice models to determine the effects of later seizures on the spatiotemporal accumulation of pathologic tau. We will also test the hypothesis that tau transmissibility in AD occurs through synaptic activation by adapting a method to permanently label cells activated by kindled seizures following tau seeding. This will allow us to measure the levels and spatial and temporal distribution of tau and AD pathology throughout the entire brain, as well as gene and protein expression at single neuron level. We will also utilize human brain tissue from AD patients with and without a seizure history, and controls, to validate molecular changes observed in the mouse models. Finally, given our prior observations regarding the therapeutic effects of rapamycin, and the hypothesis that tau accumulation and transmission is accelerated by seizures, we will use the two tau seeding mouse models to assess the therapeutic efficacy of post-seizure chronic treatment with the mTORC1 inhibitor rapamycin or the antiseizure drug levetiracetam in attenuating AD progression.

NIH Research Projects · FY 2025 · 2022-05

Project Summary Sinusoidal and lymphatic vessel regeneration are predicted to improve treatment of hematopoietic diseases and lymphedema, but are presently limited by an incomplete understanding of the molecular and genetic pathways that control growth of these specialized vascular beds. Prior studies by us and others have demonstrated that VEGFC is required for fetal liver hematopoiesis, but a clear mechanism for this requirement has not been identified. Our preliminary studies demonstrate that loss of VEGFC/VEGFR3 function or gain of CDH5 function confers identical defects in sinusoidal and lymphatic vascular growth. Further, our genetic studies demonstrate that partial loss of CDH5 rescues both the anemia and edema conferred by the loss sinusoidal and lymphatic vascular growth, respectively, in VEGFR3-deficient animals. We hypothesize that a reciprocal VEGFC/VEGFR3-CDH5 regulatory loop controls sinusoidal and lymphatic vascular growth. This proposal will test this hypothesis in vivo and in vitro, investigate the molecular mechanism of this co-regulatory axis, and determine whether manipulation of CDH5 can be used to stimulate sinusoidal and lymphatic regeneration in mature animals. These studies are predicted to provide fundamental new insights into sinusoidal and lymphatic vessel growth that may be leveraged to treat patients with hematopoietic and lymphatic vascular diseases.

NIH Research Projects · FY 2025 · 2022-05

ABSTRACT Immunotherapy has shown remarkable benefit for some patients with solid cancers. However, it has largely been ineffective for the treatment of pancreatic ductal adenocarcinoma (PDAC). Notably, cancers with liver metastasis often show decreased responsiveness to immunotherapy and PDAC commonly metastasizes to the liver. Our lab has previously shown that PDAC triggers liver inflammation during early disease pathogenesis that subsequently supports metastasis to the liver. However, it remains unclear how the liver might influence outcomes to immunotherapy. The liver is classically known as an immunoregulatory organ involved in regulating T cell tolerance. Preliminary data show that serum amyloid A (SAA) proteins released by the liver in the setting of PDAC development suppress T cell surveillance in mouse models of PDAC such that genetic deletion of SAA converts PDAC tumors from T cell “poor” to T cell “rich”. SAA are known to signal through toll-like receptor 2 (TLR2) and consistent with this, preliminary data also show that genetic deletion of TLR2 causes T cell infiltration into PDAC tumors even in the presence of high levels of SAA. This finding implicates the SAA/TLR2 signaling pathway as a determinant of T cell surveillance in PDAC. However, little is known about how TLR2 regulates T cells surveillance in cancer. It is the central hypothesis of this proposal that TLR2 signaling causes sequestration of tumor-specific effector T cells in the liver which then undergo apoptosis, thereby limiting productive T cell surveillance in PDAC. To test this hypothesis, in Aim One I will define cellular mechanisms by which TLR2 regulates T cell surveillance in PDAC. In Aim Two, I will determine the impact of TLR2 on tumor-specific T cell fate and the efficacy of T cell immunotherapy. Altogether, studies in this proposal will improve our understanding of the role of tumor-extrinsic features in regulating immune resistance in PDAC with the aim to identify novel strategies for improving the efficacy of immunotherapy. This project will be sponsored by an established mentor with expertise in cancer immunology and tumor biology. This sponsorship entails a strong commitment to mentorship at an institution with an exceptional environment for training in cancer immunology and tumor biology. The project also includes a rigorous graduate training plan to complement training in scientific research with didactics and training in scientific communication and teaching.

NIH Research Projects · FY 2026 · 2022-04

Low back and neck pain, which impose major impediments to quality of life and are responsible for significant productivity loss, are often treated with expensive, sometimes ineffective, and potentially addictive drugs. Complementary therapies, especially force-based manipulations (FBMs), have the potential to address these challenges but remain poorly understood especially for spine pain. The highly multifaceted nature of FBMs complicates a complete understanding of the physiologic mechanisms involved with pain relief, requiring the integrative synthesis of neuroscience, physics, engineering, physiology, and clinical fields. To address this need, we will create a network - SPINEWORK - of researchers from all disciplines interested in exploring the potential role of FBMs in alleviating spine pain. The network will provide members with the opportunity to identify new collaborators, to learn about other disciplines, to disseminate ideas and information to their colleagues and the larger community, and to foster better interdisciplinary communication. SPINEWORK will support the network of researchers and activities administered by a set of Committees and will be organized into intersecting and evolving Working Groups, each made up of members from multiple institutions and traditional disciplines and focused on specific areas, such as Imaging or Animal Models. The Working Groups will promote interdisciplinary research through physical and virtual gatherings, white papers, journal special issues, and video content. A special Working Group on Terminology will focus on defining a common lexicon for spine pain and forced-based manipulations; this task is essential because even simple terms can have very different meaning to different communities, leading to confusion and impeding progress for collaborative teams. SPINEWORK will sponsor a pilot and facilitation grant program for interdisciplinary collaborative teams formed by its members. The grants will be focused on opportunities to seed or boost R01 proposals, activities that build intellectual infrastructure (e.g., think tanks or workshops), and opportunities for scientists from one discipline immerse in disciplines outside their core area. Annually, SPINEWORK will bring its members together for a Summit meeting to assess its activities, report findings, spawn new Working Groups, review progress, and plan for the coming year. Overall, SPINEWORK's mission is to lay the intellectual groundwork for improved treatment of low back and neck pain via FBMs by creating and fostering a multidisciplinary, scientifically and culturally diverse network of researchers from across the spectrum of approaches to understand FBMs and spine pain.

NIH Research Projects · FY 2026 · 2022-04

The pathophysiology of Alzheimer’s disease (AD) is characterized by the accumulation of Amyloid (Aβ) plaques and tau neurofibrillary tangles (NFT). While the presence of both plaques (A+) and tangles (T+) are essential to the biological definition of AD as recently codified in the ATN research classification framework, tau (T) is thought to be the primary driver of downstream neurodegeneration (N) and the resulting cognitive impairment. However, there is substantial variability in the T-N relationship – manifested in higher or lower atrophy than expected for the level of tau in a given brain region, even in carefully curated research cohorts. What does this variability represent? In this study, we explore the idea that a quantitative measure of the variability in the canonical relationship between T and N is itself a “mismatch metric” that can help characterize different underlying phenotypes and modulatory factors. We will examine this by modeling region-wise measures of T vs. N obtained from in-vivo imaging in a cohort A+ symptomatic individuals. SUVR from tau-PET imaging and cortical thickness from structural MRI will serve as regional measures of T and N respectively. We will then use data-driven clustering for phenotype discovery based on the model residuals. Region-wise model residuals capture spatial variation in the T-N relationship, conceptually extending the ATN framework from the dichotomous T/N +/- designations to a richer description that may reflect differing spatial topography of underlying co-pathologies. The concept of the T-N mismatch metric and its ability to identify underlying phenotypes will be evaluated in multiple publicly available and institutional datasets, each of which will provide a diverse collection of phenotypes. We will also perform evaluation in a dataset of ex-vivo specimens of A+ individuals. We will obtain quantitative measures of N from ex-vivo MRI as a semi-automated cortical thickness estimate, and of T using digital histopathology techniques, in multiple brain regions. Gold standard histopathology measures (e.g. TDP- 43, alpha-synuclein, non-AD tau, vascular disease) obtained in these samples will help evaluate whether T-N mismatch metric can help identify phenotypes with non-AD co-pathology. Finally, we will evaluate if the T-N mismatch metric is predictive of future cognitive decline as well as rates of longitudinal neurodegenerative changes in the brain.

NIH Research Projects · FY 2026 · 2022-04

PROJECT ABSTRACT Between 24-40% of cancer patients in the U.S. use cannabis, principally to manage pain, anxiety, and insomnia. Importantly, evidence suggests that some patients may be substituting cannabis as a strategy to reduce opioid consumption. However, cannabis’ historical classification as a Schedule I substance (i.e., a controlled substance with no known medical use) by the United States (U.S.) Food and Drug Administration has created barriers to conducting rigorous research on its role in cancer symptom management. As a result, the 2021 National Comprehensive Cancer Network’s Adult Cancer Pain guidelines state: “Data supporting the use of cannabinoids as adjuvant analgesics for treatment of cancer pain are extremely limited and the results from what data exist are somewhat conflicting”. Moreover, despite abundant evidence of racial disparities in cancer pain treatment, little is known about the role of cannabis in mitigating racial disparities in cancer pain outcomes. Thus, there is a critical need to conduct rigorously designed research to generate new knowledge of this phenomenon. Using ecological momentary assessment (EMA) methodology and a 12-month prospective cohort design, we propose a multisite study at three health systems in the northeastern U.S. We will enroll 600 (200 per site) ambulatory patients with non-skin solid malignancies who are receiving opioid therapy: 300 cannabis users (weekly use in any form in the prior month) and 300 cannabis non-users (no use in the past 3 months). Of these, 50% will be self-identified African American patients and 50% will be White. Cannabis and opioid use will be assessed via EMA (collected for 1 week/month; 84 days total) and monthly phone surveys and patient reported outcomes (PROs) will be assessed via monthly phone surveys. This study will also describe the poorly understood phenomenon of cannabis use patterns over time by elucidating dynamic within- and between-subject changes in cannabis use, PROs, and opioid use over the course of one year. The Specific Aims are to: (1) describe dynamic within- and between-subject changes in cannabis use over time including frequency, route, source (medical vs. non-medical), indication, and composition; (2) assess if cannabis use over time is associated with key PROs (pain severity and pain-related function, sleep, anxiety, and quality of life) and opioid use (subjective and objective indices) among cancer patients; (3) test if cannabis use moderates the association between race and pain severity; and (4) explore potential moderators of the relationship among cannabis use, PROs, and opioid use including cannabis frequency, source, route, and composition, and current opioid misuse measure. This timely and comprehensive study has high potential to generate new knowledge upon which clinical practice and guidelines related to cannabis use in cancer pain and symptom management may be based. The strong multidisciplinary research team brings the requisite expertise in cancer-related pain, medical cannabis, longitudinal opioid use among patients with cancer, substance misuse, and health disparities. Overall, this research can have a sustained impact on the science of cancer pain and symptom management.

NIH Research Projects · FY 2026 · 2022-04

α-Synuclein is a small neuronal protein that is the primary component of the proteinaceous aggregates that are the hallmark of Parkinson’s disease and other synucleinopathies. Recent evidence supports the idea that structural differences between α-Synuclein aggregates, or ‘strains’, underlie different synucleinopathies. While the molecular details are not yet well-understood, it has been suggested that post-translational modifications to α-Synuclein may underlie conformational differences between ‘strains’. However, understanding how these modifications impact aggregate structure, and ultimately pathology, is extremely challenging, both given the large number of reported post-translational modifications to α-Synuclein, as well as their heterogeneous distribution in patient-derived samples. From a biochemical and biophysical perspective, many of these modifications have been addressed individually and found to have striking impacts on α-Synuclein properties, including aggregation kinetics and cellular uptake and seeding. However, there is a significant gap in our understanding of how multiple simultaneous modifications may work cooperatively to alter aggregate structure or α-Synuclein function. Our proposed research will address this deficit by taking advantage of the collective expertise of the three PIs in protein chemical synthesis, cellular and molecular biophysics, and structural biology. This will include using a novel semi-synthesis strategy – combining unnatural amino acid mutagenesis, chemoenzymatic modification, thiol-ene reactions, and native chemical ligation – to produce α-Synuclein site specifically modified both at single and multiple sites (Aim 1); determining the impact of α-Synuclein modifications on functional interactions with lipid bilayers, on the kinetics of self-association and on the structural features of the aggregates (Aim 2); and relating these structural effects to internalization of α-Synuclein by primary neurons, and subsequent seeded aggregation of endogenous α-Synuclein (Aim 3). We have selected seven different disease-associated sites on α-Synuclein that are subject to modification with diverse groups, including phosphorylation, acetylation and ubiquitination, and we will compare and contrast the individual effects of these modifications as well as their cross-talk. Through this research we expect to characterize the impact of these modifications both on α-Synuclein functional interactions as well as fibrillar structure and spread. Ultimately, the impact of this work will be in providing a thorough understanding of the molecular basis of ‘strain’ differences in synucleinoapthies and may guide the development of therapies targeted at post-translational modifications, or even open the door to new therapeutic strategies.

NIH Research Projects · FY 2025 · 2022-04

Project Summary Background: Misinformation on social media, or the sharing of false or incomplete information without intent to cause harm, is contributing to vaccine hesitancy among parents and impeding uptake of the HPV vaccine among adolescents in the United States. Innovative communication strategies that can be rapidly developed and deployed to correct evolving misinformation about HPV vaccine on social media and increase uptake of HPV vaccine are needed. Hypothesis: We hypothesize that theory-based corrective messages directly addressing the content of misinformation will improve HPV vaccine series initiation and completion as compared to general corrective messages and a non-message control group. Proposal: We will efficiently identify recently circulating misinformation on social media using natural language processing (NLP) methods, develop and test a theory-based corrective messaging tool using a validated message testing protocol, and assess the efficacy of the tool on increasing HPV vaccine series initiation and completion among adolescents in an online, block-randomized controlled trial with three arms and longitudinal follow-up. Participants (n=2500) will be parents of adolescents ages 8-11 recruited from social media and followed up over a 12-month period. Importance and Innovation: If successful, in addition to improving HPV vaccination uptake, the methods from this study have the potential to be adapted to address other forms of misinformation swiftly and efficiently in public health such as that surrounding COVID19, childhood vaccinations, or cancer prevention and treatment strategies.

NIH Research Projects · FY 2026 · 2022-04

PROJECT SUMMARY Adoptive T-cell immunotherapies and, in particular, chimeric antigen receptor T cells (CART) generated unprecedented responses in patients with highly refractory CD19+ B cell malignancies. However, only a limited number of patients treated with anti-CD19 CART will experience prolonged remission while most patients either do not respond or eventually relapse. Moreover, in the setting of solid cancers, CART cells have generally been disappointing. Increasing CART effector function against cancer would represent a vertical improvement in the field of adoptive T-cell immunotherapy. In this proposal, we aim at increasing the anti-tumor efficacy of CAR T cells by reducing the inhibition of CAR activation. Current approaches to enhance CART efficacy focus on reducing long-term T cell exhaustion by targeting the PD-1/PD-L1 axis. However, there is a lack of investigation on strategies to enhance early CAR and T cell receptor (TCR) signaling, which is critical for effective tumor killing, especially in the setting of the immunosuppressive tumor microenvironment. To this goal, we studied the scavenger receptor CD5. CD5 associates with the TCR complex and inhibits its activation through several mediators, including SHP-1, CBL, CBL-B, and GRB2. The central hypothesis of this grant is that CD5 deletion increases the anti-tumor efficacy of engineered T cells through enhancement of TCR signaling. In our preliminary work, we have found that CRISPR-Cas9 CD5 knock out in CART cells enhances their anti-tumor activity in xenograft models of B-cell leukemia (CART19) and T-cell lymphoma (CART5). Moreover, we demonstrated that, upon stimulation, CD5 knocked-out CART cells show higher phosphorylation of key CAR signaling mediators as compared to wild-type. In Aim#1, we will test the hypothesis that the deletion of CD5 in T cells broadly enhances the anti-tumor efficacy of adoptive T-cell immunotherapies, by studying CD5 knockout in the setting of CAR T cells for solid tumors. In Aim#2, we will test the hypothesis that CD5 deletion augments T-cell anti-tumor activity by disinhibiting CAR signaling by performing in vitro and in vivo mechanistic studies. We will also test the safety of this approach. This hypothesis-driven proposal is highly innovative as it proposes the study of a novel immunomodulatory target - CD5 - and uses cutting edge technologies like digital spatial profiling and single-cell RNA sequencing to investigate CD5 deleted CAR T-cell in vivo in both human and murine models. A successful outcome of the proposed investigations is expected to impact the field of cancer immunotherapy significantly, providing a rational and generalizable approach to improving T-cell immunotherapies and ultimately drive the development of a first-in-human clinical trial of CD5 knocked-out CART for refractory cancers.

NIH Research Projects · FY 2026 · 2022-04

Modified Project Summary/Abstract Section: Advances in B cell biology and molecular virology have enabled the discovery, characterization, and commercial development of several classes of broadly neutralizing antibodies (bnAbs), with applications for prevention, treatment, and cure of HIV disease are under study. Yet, virus resistance remains the central vulnerability of effective bnAb use. This application proposes to address the problem of bNAb escape by rationally selecting combination bNAb therapy that limits virus escape. We propose to apply lessons from the successful development of combination antiretrovial therapy (cART), whereby bidirectional phenotypic antagonism was exploited. Our multidisciplinary team has secured plasma virus or virus sequences from recent or ongoing prevention and treatment studies of VRC01-class CD4 binding site (CD4bs)-targeting monotherapy, as well as combination therapy with V3 glycan-targeting (Table 1). We will leverage these unique samples to map the in vivo escape pathways of virus replicating in the presence of sub-suppressive levels of these clinically relevant bNAbs (Aim 1). Using the evolving escape variants, we will identify putative complementary bNAbs with maintained or inverse antibody sensitivities from rationally designed panels of candidate bNAbs (Aim 1). We will then characterize the autologous neutralizing antibody (anAb) response in the treatment cohorts, to determine the capacity of anAbs to impede virus escape from administered bNAbs (Aim 2). Finally, we will test the most promising complementary bNAbs to restrict virus escape in cell culture, in vivo in humanized mice and a validated barcoded TF SHIV/NHP model (Aim 3). Our scientific premise is that in vivo mapping of virus escape from bNAbs, identification of complementary bNAbs, defining the role of autologous antibodies, and rigorous in vivo testing in an authentic NHP model will elucidate basic mechanisms of virus resistance to bNAbs and inform more effective use of bNAbs across the HIV prevention, treatment and cure fields. If accomplished we will (i) have defined the sensitivities of escaped viruses from clinical trials to alternate bNAbs, (ii) identified bNAbs that cannot mutually escape using the same pathway, (iii) defined the role of anAbs in bnAb escape and (iv) tested the ability of complementary bnAbs to improve therapy in an authentic NHP model.

NIH Research Projects · FY 2025 · 2022-04

Spatially Responsive Mass Vaccination Campaigns for Urban Rabies SUMMARY Epidemics of vaccine-preventable zoonotic diseases are ongoing in major urban centers across Latin America and worldwide. Mass vaccination campaigns can prevent and control epidemics of infectious diseases among humans and animals. The realities of mass vaccination efforts, however, often fall short of their promise. In place of high and even vaccination coverage, many campaigns leave spatial 'pockets' of under-vaccinated individuals. Pathogens, taking advantage of these under-vaccinated areas, can persist, diversify, and re-emerge. Modern computational approaches can mitigate geographic inequities through the careful placement of vaccination sites, increasing demand for vaccination. However, increased demand creates long queues for service, an additional barrier to receiving vaccination. The main hypothesis of our study is that participation in mass vaccination campaigns can be significantly increased through spatially responsive vaccination strategies. These strategies have the potential to maximize coverage, reduce spatial heterogeneity, minimize waiting time at vaccination sites, and increase equity in access to immunization in hard-to-reach populations. We test our methods in the context of an ongoing canine rabies epidemic in the city of Arequipa, Peru. In the first aim we compare the coverage and spatial equity of a new vaccination strategy, incorporating spatial optimization and queueing theory, to current practice through a stepped-wedge cluster randomized trial. In the second aim we develop and test data-driven methods to reach underserved populations through mop-up “precision vaccination” campaigns. In the third aim we assess the acceptability, scalability, and transferability of spatially responsive vaccination strategies for local, regional, and national stakeholders. Our work will lead to new principles and practices that bridge operational research and zoonotic disease epidemiology to better guide vaccination programs. It will allow for the design of effective elimination strategies that will take into account access to healthcare and spatial behavior of vaccinators.

NIH Research Projects · FY 2025 · 2022-04

PROJECT SUMMARY/ABSTRACT The goal of this proposal is to improve treatment selection and survival in women with advanced ovarian cancer. Ovarian cancer is the leading cause of deaths from gynecologic cancers in the U.S. Most women present with advanced disease, with distant spread at the time of diagnosis. Even so, more than 1 in 10 of such women will survive >10 years after an initial diagnosis, usually with periodic recurrences. The possibility of long-term survival underscores the paramount importance of each treatment decision. Determining the best treatment strategy for an individual patient is difficult. For women with newly diagnosed, advanced ovarian cancer, both surgery and chemotherapy are recommended. However, many women fail to complete both, rates of death from surgery are high, and chemotherapy delivery is often limited by toxicities. New therapies that are designed to target cells at the molecular level (“PARP inhibitors”) have recently been approved for use, but their benefits vary based on the presence of specific tumor mutations, and their costs exceed $150,000/year. Carefully tailored decisions about the sequence of surgery and chemotherapy, types of chemotherapy, and the way chemotherapy is delivered, could improve long-term outcomes and reduce costs. To address this problem, we will build a simulation modeling framework that projects the outcomes of women treated for advanced ovarian cancer, and use it to identify personalized treatment approaches. We have built a preliminary model that projects outcomes for women with newly diagnosed, advanced ovarian cancer. We will extend our model to include detailed patient and tumor characteristics – age, comorbidities, stage, and mutation status – that influence survival, as well as new therapies and toxicities (Aim 1.1). We will also simulate the treatment of recurrent cancer (Aim 1.2). Using our new modeling framework, we will identify tailored treatment approaches that optimize survival (Aim 2.1), minimize treatment toxicities (Aim 2.2), and are cost-effective (Aim 3). Finally, we will identify future studies that are likely to have the greatest impact in improving treatment decisions (Aim 4). To ensure that our findings are accessible to patients, physicians, and policymakers, we will create an online, interactive version of our modeling framework that can project outcomes, quantify trade-offs, and support decision-making in real time. The proposed research will result in: 1) personalized treatment recommendations; 2) real-time guidance for decision-making; 3) the capacity to rapidly weigh benefits of new therapies with long-term risks and costs; and 4) prioritization of future research. The knowledge gained will provide new opportunities to improve treatment selection and survival in ovarian cancer.

NIH Research Projects · FY 2025 · 2022-04

A. Specific Aims Type 1 diabetes (T1D) is a progressive autoimmune disease which renders individuals incapable of regulating their blood glucose levels due to immune-mediated β cell destruction, resulting in loss of insulin production and many severe health complications that, if untreated, lead to death. Careful monitoring of blood glucose coupled with insulin injections have made T1D a chronic disease in which T1D individuals live ~ a decade less than their healthy counterparts. Thus, a cure for T1D is highly desirable. Cell gene therapy has proven to be an effective way to treat recalcitrant diseases such as pediatric leukemia, where chimeric antigen receptor (CAR) expressing T cells achieve 90% complete response, putting many individuals into very long remissions1, but to date no engineered T cell therapies have been attempted to cure T1D in humans. We propose to test the hypothesis that engineered T cells can enable islet transplant with minimal or no additional immunosuppression. Islet transplantation represents the best-case scenario to test the ability of engineered T cells to protect islet cells from immune attack and will likely lay the foundation by which strategies are developed to treat new onset T1D. In islet transplant, MHC matching of donor and recipient rarely occurs, generating highly expressed, unique islet-specific HLA antigens that can be targeted by CAR engineered T regulatory cells (CAR Tregs) or T cells engineered to express molecules that suppress the immune system (T suppressor cells or Tsups). Additionally, through a comprehensive screening process, we have identified two targets, fibroblast activation protein (FAP) and dipeptidyl peptidase like 6 (DPP6), that are highly expressed on α and β cells and have limited expression elsewhere that could be used to treat recently diagnosed individuals, and all transplant recipients without the desired MHC mismatch. In this proposal, we will further develop this toolbox to both develop better in vivo, preclinical models of T1D and new cell and gene therapies that will prevent, stall or reverse T1D. Within the last decade, rapid progress made in T cell-based therapies makes it possible to consider such therapies for T1D. Following long-term remission of 3 cancer patients treated by CD19-specific CARs developed by the Center for Cellular Immunotherapies at Penn under the leadership of Carl June2, the CAR T cell revolution was launched. This early success drove considerable investment, empowering many institutions and companies to develop ways to improve both the safety and efficacy of, and reduce cost to produce engineered T cells. Many of these innovations will also help enable cell and gene therapies for T1D, which is the overarching goal of this RFA. The ultimate goal of this application is to successfully treat three non- human primates (NHP) with engineered T cells after an islet transplant, which we predict will launch similar enthusiasm for T1D cell and gene therapy as the first three CD19-CAR recipients did for cancer CAR therapy. To achieve this goal, an experienced team of investigators with complementary expertise will lead this proposal. This team, a transplant surgeon who helped pioneer islet transplantation as a T1D therapy (Naji), a veterinary physician with expertise developing NHP T regulatory cell therapy models (Duran-Struuck), and a cell and gene therapist with a track record of engineering T cells and developing first-in-human clinical trials (Riley), have been working closely together via Helmsley Foundation funding to obtain the preliminary data presented in this application. This team is now is poised to immediately perform in vivo studies to test the ability of engineered T cell therapies to prevent, stall, or reverse T1D. Aim 1. Engineer T Suppressor Cells (Tsups) to Facilitate Islet Transplant in Humanized Mouse Models. While T regulatory cells are potent immune suppressors, there are other promising ways to induce tolerance that may work as stand-alone therapies or synergize with CAR Tregs to protect β cell function. We will explore using PD-L1, TGF-β1, and/or γ-aminobutiric acid (GABA) expressing cells targeted to the islet to mediate islet acceptance alone or in combination with CAR Treg approaches we developed using humanized mice. Aim 2. Optimize T cell Approaches to Promote Islet Transplant in NHPs. To date our efforts have focused on developing NHP CAR Tregs to enable islet transplant. We have optimized isolation, transduction, and expansion of NHP MHC-specific CAR T regulatory cells and are poised to start islet transplant experiments immediately. After we perform proof of principle experiments using CAR Tregs, we will prioritize future experiments based on the data obtained in Aim 1. A particularly exciting aspect of our studies is testing whether the ortho-IL-2 system developed by the Garcia Lab3 can work in non-human primates and support engineered Treg or other engineered T cell function and expansion while having no effect on endogenous effector cells. If successful, these studies will form the basis and rationale for Phase I clinical trials in humans.

NIH Research Projects · FY 2026 · 2022-04

Project Abstract Research: Asymmetric cell division is an evolutionarily conserved mechanism that affords self-renewal, differentiation and diversification of cell populations. It is unknown, however, if human T cells use this mechanism to induce distinct daughter cell fates. The proposed research will test the hypothesis that asymmetric cell division is an indispensable mechanism of human T cells to generate functionally distinct daughter cells. The experiments will use a novel method of target-induced labeling of the immunological synapse, multicolor flow cytometry, single cell RNA sequencing and metabolomic profiling to identify and ultimately modulate cellular programs that support long- and short-lived progeny induction in both native and genetically-engineered human T cells. A better understanding of cell division patterns will expand our knowledge on human T cell differentiation, uncover factors promoting niche-specific T cell persistence, and establish biology-driven principles and methods for optimization of T cell immunotherapy. Candidate: Dr. Ellebrecht earned his MD from the University of Lubeck, Germany and will complete Dermatology residency training at the University of Pennsylvania in 2021. Dr. Ellebrecht is pursuing postdoctoral training in Dr. Aimee Payne’s and Dr. Carl June’s laboratories at Penn. The 5-year career development plan includes structured coursework and training in single cell transcriptional profiling, bioinformatics, and metabolic profiling of human T cells from experienced mentors and collaborators, along with professional career development activities, with the goal of establishing an independent, NIH-funded research laboratory investigating fate induction and longevity of skin-resident and engineered T cells. Environment: The mentors, Dr. Aimee Payne and Dr. Carl June, are renowned NIH-funded Penn investigators, who provide unparalleled expertise in T cell biology, immunotherapy, metabolomics and single cell characterization of human T cells in combination with an exceptional mentoring record including prior K08 and K23 awardees. Dr. Ellebrecht’s focus on asymmetric cell division in skin resident T cells provides a clear path to independence that sets him apart from Dr. Payne’s and Dr. June’s focus on translational immunotherapy. Dr. Ellebrecht’s research and training will be supported by the Penn Dermatology Department, Center for Cellular Immunotherapy and CHOP metabolomics core, providing state-of the-art core facilities for flow cytometry, cell sorting, single cell sequencing, metabolomics, human skin xenografts and bioinformatics. Taken together, his mentors, collaborators and access to these top-notch technologies will create an ideal environment for Dr. Ellebrecht to thrive on his path towards becoming an independent physician scientist leading efforts to characterize and modulate T cell populations responsible for chronic inflammatory skin diseases.

NIH Research Projects · FY 2026 · 2022-04

The overall goal of this proposal is to dissect the molecular mechanisms of metabolic regulation of ATP-citrate lyase (ACLY) and to characterize ACLY inhibitors for cancer therapy. ACLY is the predominant source of nucleocytosolic acetyl-CoA, an essential building block for the production of fatty acids, cholesterol, isoprenoids and protein acetylation. Elevated ACLY activity is found in metabolic disorders, cardiovascular diseases and many cancers, prompting the development of several ACLY inhibitors. While many ACLY inhibitors have been developed, only bempedoic acid, which forms an active bempedoyl-CoA adduct in hepatocytes, has been approved by the FDA for therapeutic use. Risk of hepatocellular carcinoma is elevated in individuals with metabolic disorders, many of whom may be candidates for treatment with bempedoic acid; yet metabolic regulation of ACLY activity and its functional role in hepatocellular carcinoma remain poorly understood. Elevated levels of ACLY acetylation at K540, K546 and K554 and phosphorylation at S455 and S481, and retention of exon 14 encoding a region with S481, have also been correlated with cancer, thus also suggesting roles for ACLY posttranslational and posttranscriptional modification in cancer metabolism. ACLY is an ~500 kD multidomain homotetrameric enzyme that uses citrate, CoA and ATP cosubstrates to produce oxaloacetate (OAA) and acetyl-CoA. Until recently, the lack of structural information on intact human ACLY has hampered understanding of its molecular mechanism of catalysis and the structure-based development of inhibitors. The Wellen lab recently reported on various disease-associated phenotypes associated with dysregulated ACLY function; and the Marmorstein lab reported on the cryo-EM structures of ACLY in different reaction states, along with associated biochemical and biophysical studies, to elucidate the molecular basis for acetyl-CoA production by ACLY. The latter findings lead to several unresolved questions underlying the metabolic regulation of ACLY and set the stage for the structure-based development of more potent and selective ACLY inhibitors for therapeutic applications. These recent studies now position the Wellen and Marmorstein labs to work together to resolve important gaps in knowledge in metabolic regulation and inhibition of ACLY, through the following specific aims: (1) Evaluate the role of metabolic binders in ACLY activity, (2) Determine the molecular mechanism of how posttranslational modifications and exon 14 retention impact ACLY regulation, and (3) Evaluate the molecular mode of action of ACLY inhibitors. Together, these studies will reveal the molecular mechanisms for how ACLY activity and regulation is mediated by the binding of metabolites, and posttranscriptional and posttranslational modification and will lead to the rational development of ACLY drugs to treat cancer.