University Of Pennsylvania

NIH Research Projects · FY 2026 · 2026-05

Project Summary Bones patients decreased worldwide, improve and repair. are normally able to heal robustly after injury. However, this capacity can go awry in 5-10% of in the form f delayed or nonunion fractures, causing substantial morbidity, loss of productivity, and quality of life. Old age is a major risk factor. With the rising prevalence of geriatric fractures there is an urgent need to understand the bone repair mechanism and identify new therapies to fracture healing. Bone regeneration is an intricate and complex process involving various cell types growth factors. The periosteum houses mesenchymal progenitor cells that play a central r ole in skeletal Impaired activity of periosteal mesenchymal progenitors contributes to delayed healing in aging. o We recently performed an unbiased and comprehensive single-cell RNA sequencing (scRNA-seq) analysis of intact and fracture-injured periosteum in young adult mice and identified a novel mesenchymal progenitor population - termed proliferative progenitor cells (PPCs)- which rapidly and robustly expands post-fracture. PPCs exhibited myofibroblast-like features and occupied a transitional state along the differentiation trajectory from primitive mesenchymal progenitor cells (MPCs) to terminally differentiated osteoblasts and chondrocytes. Interestingly, PPCs highly expressed Activin A, a member of TGFβ superfamily of growth factors. Analyses of both mouse and human periosteum revealed a marked post-fracture increase of Activin A, which was significantly blunted in aged mice compared to young ones. In culture, Activin A stimulated the proliferation of periosteal mesenchymal progenitors and promoted their differentiation into myofibroblasts, chondrocytes, and osteoblasts. Using two bone injury models (fracture and drill hole), we obtained complementary evidence that bone repair is delayed by systemic administration of a neutralizing antibody against Activin A and accelerated by local delivery of Activin A. Our central hypothesis is that Activin A expressed by a myofibroblast-like mesenchymal subpopulation, the PPCs, at the early stage of bone healing is essential for regulating periosteal mesenchymal progenitors to stimulate bone regeneration. Our objectives are to define the roles of this effector in bone repair and develop approaches targeting its mode of action to treat delayed or nonunion fracture healing, particularly in aging. Our aims are to: 1) elucidate the cellular mechanism of Activin A action on bone healing by examining the repair process in mice with Activin A deficiency in various cell types, including PPCs; 2) delineate the molecular mechanism of Activin A action on bone healing by studying Activin A signaling components in fracture callus, primary periosteal mesenchymal progenitors, and aged fractures; 3) target Activin A pathway for accelerating bone repair by designing a new hydrogel drug delivery system. Our proposal. proof-of-concept multidisciplinary team has worked together to generate the exciting preliminary data for this If successful, this proposal will reveal the critical role of Activin A in bone regeneration and provide evidence for targeting this novel pathway in fracture therapy development.

NIH Research Projects · FY 2026 · 2026-05

Project Summary/Abstract: We request funding through the NIH S10 Shared Instrumentation Grant program to acquire a LUMICKS Avidion Cell Avidity Analyzer, a transformative high-throughput platform designed to quantitatively measure cell-cell interaction avidity. Currently, our institution faces significant limitations in systematically assessing cellular avidity, a critical parameter predictive of efficacy in adoptive cellular immunotherapies such as chimeric antigen receptor (CAR) T-cell therapies. The proposed instrument addresses this gap, enabling simultaneous, high-throughput quantification of avidity across hundreds of experimental conditions, overcoming bottlenecks of existing low- throughput methods. The Avidion analyzer uniquely integrates advanced centrifugal force-based technology, multicolor fluorescence imaging, automated liquid handling, and robotic cartridge management. This combination provides unprecedented sensitivity, throughput, and specificity in measuring interactions between engineered immune cells and their targets. Our major and minor users propose projects leveraging this advanced capability to significantly accelerate discovery and clinical translation. Major user projects include: (1) Systematic CRISPR-based screening of adhesion and costimulatory molecules influencing immune synapse formation and CAR T functionality; (2) High-throughput quantification of anti-CD79 CAR T-cell avidity to predict in vivo efficacy in lymphoma models; (3) Development of peptide-centric CAR-T cells targeting KRAS G12V and other neoantigens in solid tumors; (4) Identification of resistance mechanisms in BCMA- targeted immunotherapies in multiple myeloma through avidity profiling; and (5) Characterization of T- cell avidity impairments caused by lipid-rich tumor microenvironments in ovarian cancer ascites. Minor users will utilize the instrument to optimize novel CAR constructs, synthetic adhesion molecules, and T- cell receptor-based therapies, addressing critical scientific and clinical questions across hematologic malignancies and solid tumors. Acquisition of the LUMICKS Avidion analyzer will significantly enhance NIH- funded translational research efforts, driving innovative therapies from bench to bedside, ultimately benefiting public health through improved patient outcomes.

NIH Research Projects · FY 2026 · 2026-05

Project Summary Dialysis is an important life-sustaining therapy for kidney failure in the critical care setting, and a crucial method for volume management for patients with either severe acute kidney failure or chronic kidney failure. While most patients will initiate continuous kidney replacement therapy (CKRT) in the intensive care unit, many patients who continue to require dialysis throughout the hospitalization will transition to intermittent hemodialysis (iHD). The transition from continuous to intermittent dialysis is a hemodynamically vulnerable period due to the combination of increased thermal and hemodynamic stress attributed to iHD and exposure to faster rates of fluid removal (ultrafiltration) as a result of the shortened dialysis time. Moreover, irrespective of volume status at the time of transition, even minimal rates of fluid removal to maintain net even fluid balance may cause circulatory compromise due to the lingering effects of critical illness, which impact “ultrafiltration tolerance” and may persist for weeks after initial resolution of shock. Consequently, hemodynamic instability and intradialytic events during iHD in the week after transition from CKRT are common and result in recurrent fluid overload, a return to CKRT, and deleterious effects on recovery of kidney function or loss of residual kidney function. Thus, successful transition from CKRT to iHD depends on not only hemodynamic stability at the time of CKRT cessation, but also the anticipated volume removal needs and assessment of “ultrafiltration tolerance” to determine if maintenance of volume and hemodynamic stability on iHD is likely. However, “ultrafiltration tolerance” is difficult to predict. There is currently a lack of an established approach to guide the optimal timing of the transition and the prescription of iHD following CKRT, leading to large variability in practice patterns and unclear criteria for assessing readiness to transition dialysis modalities. This proposal combines (1) an analysis of a large electronic health record database using machine learning algorithms and longitudinal modeling methods to characterize hemodynamic patterns of “ultrafiltration tolerance” and develop and validate a clinical risk score tool to guide decisions on transitioning from CKRT to iHD (Aim 1); and (2) a prospective observational study with novel integrated hemodynamic monitoring technology to elucidate the role of plasma refill rate (the ability to reconstitute the intravascular space) as a mechanism of “ultrafiltration tolerance” (Aim 2). The proposed studies provide an innovative perspective for CKRT delivery, with a focus on detailed phenotyping of patient and dialysis factors, rather than the volume or rate of fluid removal, as determinants of fluid removal tolerability. Results from these studies will provide both practical and scalable risk prediction models that can be tested in other electronic databases, clinical risk scores that can be incorporated for clinical use, and potentially intradialytic monitoring tools with clearly defined metrics to guide decisions about fluid removal during transitions from continuous to intermittent dialysis.

NIH Research Projects · FY 2026 · 2026-05

Summary Leukocyte motility is critical for immunology, inflammation, and hemostasis. Immune cells exchange molecular information by direct contact, enabled by motility within secondary lymphoid organs. In inflammation, neutrophils crawl into sites of infection after adhering to blood vessel walls. Hematopoietic stem and progenitor cells (HSPCs) migrate into to bone marrow after adhesion to the blood vessel wall under flow. In work largely funded by NIGMS, the Hammer laboratory has worked to understand the chemo- mechanics of leukocyte migration for over two decades. We have used traction force microscopy (TFM), in which we measure the forces exerted by cells during motility by monitoring the defection of fiduciary beads embedded within an elastic polyacrylamide gel. We have used this method to measure the traction forces of neutrophils and macrophages, as well as many other cell types. With the technique of TFM in hand, we are now positioned for significant breakthrough in our molecular understanding of traction stresses during leukocyte motility, owing to the development of methods to delete or alter intracellular components within a cell. Of specific interest is the fascinating phenomenon of upstream migration, in which leukocytes migrate against the direction of flow on surfaces presenting intercellular adhesion molecule-1 (ICAM-1). Using CRISPR-Cas9, we now are able do a directed screen of molecules that have been implicated in cell migration, and specifically, upstream migration, to understand precisely how these molecules contribute to the generation of traction forces in leukocytes. We will use two cultured cells lines – KG1a cells (a model HSPCs) and HL-60 cells (a model neutrophil) – that allow us to compare the role of different intracellular effector molecules in cell motility to establish universal mechanisms. We will conduct a directed screen of a limited but important set of effector molecules which have been implicated in upstream migration and leukocyte motility, such as cytoskeletal regulators and Rho-GTPases. Our elucidation of traction stresses will be complemented by immuno-fluorescent staining of the spatial distribution of adhesion receptors and actin cytoskeleton to provide information about cell organization. This MIRA is organized in three projects. In Project 1, we will perform traction mapping of KG1a cells during upstream migration. We will then screen candidate controllers of upstream migration using CRISPR-Cas9 and then measure how deletion affects both directional motility and traction stresses. In Project 2, we will use CRSIPR-Cas9 to screen a family of motility modulators and study their effect on HL-60 cell chemokinesis and chemotaxis. In Project 3, we will use TFM to measure the traction stresses of HL-60 cells during upstream migration and when upstream migration is reversed through CRISPR-Cas9 deletion. Then, we will measure the correlation between upstream migration of HL-60 cells and trans-endothelial migration on HUVEC monolayers.

NIH Research Projects · FY 2026 · 2026-05

7. PROJECT SUMMARY/ABSTRACT Globally, approximately 300 million live with chronic HBV (CHB) and 40 million people live with HIV. Due to shared transmission routes, approximately 10% of people with HIV (PWH) also live with HBV (PHBV). Importantly, HIV/HBV coinfection is associated with increased mortality. Like HIV, while effective HBV treatments are available, there is no cure for HBV. Hepatitis B surface antigen (HBsAg) loss, the definition of HBV functional cure, decreases incidence of hepatocellular carcinoma (HCC) and end-stage liver disease (ESLD), yet is not easily attained with current therapy. As such, the development of novel HBV therapies, aiming for HBV functional cure, is rapidly advancing. HBV functional cure research has become a scientific priority of the NIH, industry, academia, and the community with over 50 clinical trials and compounds in development. Interestingly, HBV functional cure occurs more often in PWH/PHBV, making this an ideal cohort in which to study HBV curative therapies. Despite the pace and volume of HBV cure research, there is little knowledge about patient and provider perspectives on key HBV clinical trial approaches – unlike in HIV cure research, where over a decade of experience has informed priorities. One of the key clinical trial–related questions is whether and how to implement HBV treatment discontinuations, which will be necessary to study the efficacy of curative and finite HBV novel therapies. This will pose specific challenges in PWH/PHBV, such as the heightened likelihood of rebound hepatitis and the ongoing need for HIV antiretroviral treatment (ART). There is a notable lack of data regarding both patient and provider knowledge and priorities concerning this pivotal research phase, including questions of whether HBV treatment discontinuations are desired, the duration of discontinuation, or how to manage it. Similarly, there are limited data on trial design elements such as type of novel therapies, timing of therapy initiation, and willingness to participate in HBV cure trials. These knowledge gaps remain key hurdles to advancing effective, safe, and acceptable HBV cure strategies. Data from this application will be crucial for the development of HBV clinical trials attractive to PWH/PHBV and PHBV. Our proposed work will inform outreach, recruitment, consent, and retention of participants in future HBV clinical trials. To this end, our Specific Aims are as follows: Aim 1 will explore priorities of PWH/PHBV, PHBV and providers around key aspects of HBV clinical trials including HBV treatment discontinuations, choice of novel HBV agents, and trial participation using in-depth interviews. Aim 2 will quantify priorities of PWH/PHBV and PHBV around these same trial elements through a national U.S. survey. Aim 3 will develop ethical considerations and obtain feedback on provider- and patient- facing materials in collaboration with key stakeholders. Together, our three aims will clarify therapeutic advancement priorities and guide the ethical design of future innovative HBV clinical research with significant implications for population health. This work is directly relevant to the health of all Americans, as it addresses chronic disease management, co-morbidities, and the reduction of long-term burdens on the healthcare system.

NIH Research Projects · FY 2026 · 2026-05

PROJECT SUMMARY Rotator cuff tendinopathy is a prevalent chronic disease that is painful, debilitating, and reduces the quality of life. Physical therapy (PT) is an effective first-line treatment, but the non-response rate to PT is still high. Long- term recovery is further limited, and the reason for pain recurrence is poorly explained. It is possible that the in- clinic gains of PT fail to translate to good real-world mechanics and are overwhelmed by the pathomechanical shoulder movements in activities of daily living. Clinical and basic science both suggest shoulder overuse and motion-driven pathomechanics are causal factors of rotator cuff pathology. These biomechanical mechanisms are unconfirmed in patients partly due to the complex interplays among pathology, mechanics, and treatment, in addition to a paucity of methods to quantify the highly variable real-world motion and in-vivo tendon loading. My overall goal is to discover how real-world shoulder biomechanics interact with the clinical outcomes of PT in rotator cuff tendinopathy. This research progresses from a cross-sectional [K99] to a longitudinal cohort study [R00]. My cross-sectional Aim 1 [K99] will use 1) wearable sensors to track 7-day bilateral shoulder cumulative motion in daily living; 2) motion analysis and musculosketeal models to identify motion-driven pathomechanics specific to daily living functional tasks; and 3) machine learning to combine sensor-detected activities with motion-driven biomechanics to quantify real-world cumulative rotator cuff tendon loading. My mentored K99 will validate research tools and define how patient shoulder motion and tendon loading deviate from normal status. The longitudinal Aim 2 [R00] will link the real-world biomechanics established in Aim 1 with changes in clinical outcomes, including pain, shoulder function, and tendon structure, through a 3-month PT protocol followed by the first 6 months after PT completion. These Aims will clarify the mechanical mechanisms on how real-world shoulder motion and loading alter the course of PT clinical outcomes in rotator cuff tendinopathy. Findings will inform data-driven continuous rehabilitation paradigms that monitor and modify daily living activities to maintain long-term shoulder health. This research will also serve as the foundation of Dr. Ke Song's career development plan to acquire 2 years of mentored training [K99] and prepare for transition into independence [R00]. Through the guidance of his experienced and interdisciplinary mentoring team, Dr. Song will perform hands-on training with field-leading experts in shoulder biomechanics, physical therapy, orthopaedics, wearable sensors, data science, and biostatistics to maximize the impact of his proposed research. Dr. Song will also learn from formal courses and professional training activities to ensure his career progress, develop clinical research proficiency, improve grant writing, mentoring, and leadership skills, and secure an independent tenure-track faculty position at a clinical research-intensive institution. The scientific training, professional development, and research plans will guide Dr. Song through a pathway to independent research in translational musculoskeletal biomechanics that help patients with rotator cuff tendinopathy and related shoulder diseases improve long-term outcomes.

NIH Research Projects · FY 2026 · 2026-05

Expanding upon the parent PHLHousing+ Study (5R01NR021122-02), the overarching goal of this proposal is to test whether interventions addressing housing insecurity as a modifiable social determinant of health (SDOH) improve youth mental health outcomes and outpatient service utilization in households of low-income renters in Philadelphia. This objective is aligned with the strategic aim of the NIMH (Goal 3) to identify opportunities to implement interventions that target modifiable SDOH (see Strategy 3.3.A). The PHLHousing+ Study comprises three groups, all of whom earn below 50% area median income, have at least one child under the age of 16 years living at home, and are renters: 301 households who receive monthly direct cash payments in lieu of a rental voucher for 3.5 years(Cash group), 169 households who receive a rental voucher (Voucher group), and 711 households on the Philadelphia Housing Authority (PHA) waitlist unlikely to receive rental assistance during the entire study period. Our analytic plan combines Cash and Voucher groups into a single Intervention group. Of the 1,181 households in the study, 95.4% are headed by single women and 86.3% are Black. There are 1,965 children in the sample, ranging in age from 3 to 15 years at baseline (M= 8.66, SD= 4.70). Monthly cash payments range from $89 to $2079, with a median payment of $881; payments vary based on household income, family size, and fair market rent. All three groups are surveyed every six months for four years; the first wave of online surveys was deployed in August 2022. Existing surveys include measures of youth emotional and behavioral problems (EBP) reported by primary caregivers. Recent approval from Philadelphia’s Department of Behavioral Health and Intellectual Disability Services (DBHIDS) allows us to pair the repeated survey assessments with de-identified Medicaid claims data for youth participants. I hypothesize that Intervention group youth will demonstrate significant decline in EBP and rates of clinically significant EBP (indicated by increased rates of symptom remission) over time compared to Control group youth (Aim 1). I hypothesize that a subsample of Intervention group youth with clinically significant EBP will be significantly more likely to initiate and retain use of outpatient mental health services compared to Control group youth (Aim 2). Study findings will inform research and policymakers of broader social and health benefits of economic interventions targeting housing as a SDOH. My fellowship training at the University of Pennsylvania will leverage extensive mentorship, coursework, workshops, and seminars. With guidance from a strong mentorship team (Drs. Jaffee, Reina, Mandell, Candon, and the Penn BAC), my proposal and the accompanying training plan provide an ideal foundation for my planned career as an independently funded, leading clinical scientist studying mental health policy and evaluating the implementation of social programs that might affect mental health and service use.

NIH Research Projects · FY 2026 · 2026-05

Project Summary Chronic kidney disease (CKD) poses a major global health burden, yet therapeutic options remain limited and largely repurposed from other diseases. A key barrier to therapeutic progress is the incomplete understanding of CKD’s molecular and genetic mechanisms. Genetic factors explain 30–50% of CKD risk, but genome-wide association studies primarily identify non-coding variants, complicating functional interpretation. Most gene expression studies in CKD have focused on total gene expression, overlooking alternative splicing. Short-read RNA sequencing of 404 human kidney samples showed alternative splicing explains an additional 5% of CKD heritability. However, short-read sequencing fragments RNA, limiting full isoform characterization. Long-read sequencing of 8 kidney samples identified 67% novel isoforms and revealed significant isoform differences between CKD and control samples. Given the limitations of short read sequencing, I will employ long read sequencing to comprehensively characterize the role splicing in kidney disease. I will first employ this in bulk kidney tissue to identify novel isoforms, compare the isoform expression between CKD and controls and to compare the identified isoforms with mass spectrometry data to clarify the relationship between the spliced isoforms and protein levels. I will then perform single cell long read sequencing to generate the first isoform-resolved atlas of kidney cell populations, enabling identification of cellular states associated with disease. Differential isoform usage at single-cell resolution will clarify which kidney cell types and isoforms drive disease pathology. This project will comprehensively characterize the splicing events in the kidney and the role in kidney disease.

NIH Research Projects · FY 2026 · 2026-05

Project Summary/Abstract High grade serous ovarian cancer (HGSOC) is characterized by high genomic instability where roughly 50% have deficiency in homologous recombination (HR) due to mutations in BRCA1 and BRCA2. Poly (ADP-ribose) polymerase inhibition (PARPi) is the standard for maintenance therapy for HR deficient cancers such as HGSOC. Interestingly, HGSOC tumors have chronic elevation of type 1 interferon signaling. Type 1 interferon (IFN) signaling is a critical immune mediating pathway that when acutely activated by strategies such as PARPi promotes an anti-tumor immune response in murine models. However, HGSOC and other cancers with genome instability associate with poor responses or resistance to immunotherapies and have immune suppressive tumor microenvironments (TME). Clinical trials that combine PARPi and immunotherapy had low response rates. The mechanism driving this immune suppression and tumor progression is unknown. The goal of this study is to identify the immune cells and immune signaling mediators that drive an immune suppressive TME and promote BRCA deficient HGSOC tumor progression. To address the relationship between BRCA deficiency, high inflammatory signaling, and an immune suppressive TME, I have generated novel, syngeneic BRCA deficient HGSOC murine cell line models to allow analyses in immune competent hosts. My preliminary data demonstrates that abolishment of type 1 IFN signaling in BRCA2 deficient HGSOC tumors does not affect tumor progression and survival of mice, suggesting possible alternative immune mediating pathways driving tumor progression and immune suppression. Interestingly, I observe increased activity of the inflammasome, a multimeric protein complex that promotes pro-inflammatory cytokine release. Loss of caspase-1, the major inflammasome effector, in BRCA2 deficient HGSOC tumors prolonged survival of mice. Thus, I hypothesize that genomic instability and DNA damage due to BRCA deficiency activates the inflammasome in HGSOC that mediates the immune response towards an immune suppressive, pro-tumor TME. With our novel, syngeneic BRCA deficient HGSOC models, I am uniquely positioned to Aim 1: Determine the mechanism of inflammasome activation in BRCA deficient HGSOC tumors and Aim 2: Identify the immune cells and tumor cell intrinsic mediators driving BRCA deficiency associated TME inflammation and determine their roles in immune suppression in HGSOC tumors. In Aim 1, I will determine the conditions necessary for inflammasome activation: amount of DNA damage, duration of DNA damage in the cells, presence of cytoplasmic DNA or DNA damage signaling activity. In Aim 2, I will determine the immune cell mediators, by immunofluorescence, and which effectors of the inflammasome, by genetic knockout of each one, drives an immune suppressive TME and tumor progression. Additionally, I will test how loss of each inflammasome effector impacts HGSOC tumor sensitivity to PARPi and immunotherapies alone or in combination. Completion of these aims may inform predictive biomarkers and therapeutic targets to improve HGSOC responses to immunotherapies and patient outcomes.

NSF Awards · FY 2026 · 2026-05

Early embryonic development is characterized by a dramatic transition from cellular dependence on RNAs that were in the egg prior to fertilization to RNAs produced by the embryo itself. This process is known as zygotic genome activation (ZGA). The timing of ZGA is linked to changes in cell size, specifically the ratio of nuclear content (DNA) to cytoplasmic volume (also known as the nuclear-to-cytoplasmic or N/C ratio), which increases as cells replicate their DNA without growth. How cells measure this ratio and use it to control gene expression is a fundamental biological question that remains poorly understood. This project will address it using fruit fly embryos as a model system, combining cutting-edge live imaging with genetic and genomic approaches to reveal the molecular logic by which cells sense their size and activate transcription accordingly. Understanding how cells coordinate gene expression with cell size has broad implications for developmental biology and human health, as defects in this process can lead to developmental disorders and are implicated in diseases such as cancer. This collaborative project will also provide interdisciplinary training opportunities for graduate students at the interface of genomics, quantitative imaging, and computational biology, addressing workforce development in biotechnology and biomedicine. The project will also include educational outreach activities that engage K-12 students with key concepts in basic biology and genetics. This project will use Drosophila embryos to uncover the molecular mechanisms by which the N/C ratio controls the timing of zygotic genome activation. In Aim 1, the investigators will use existing RNA-seq data sets from embryos arrested at low N/C ratios to systematically identify genes that directly respond to the N/C ratio to initiate zygotic transcription. A subset of the resulting candidate genes will be selected for quantitative live imaging using the MS2/MCP transcription reporter system to determine specific transcriptional parameters (e.g., probability and timing of transcriptional activation) that respond to changes in the N/C ratio. Mathematical modeling will be used to extract quantitative transcriptional parameters from the data and determine what aspects of transcription respond to the changing N/C ratio. In Aim 2, the investigators will leverage the genome-wide set of N/C ratio-sensitive genes to identify cis-regulatory sequences and trans-acting factors that confer N/C ratio sensitivity. Bioinformatic analysis will be used to identify DNA motifs enriched in the regulatory regions of N/C ratio-sensitive genes, and systematic enhancer dissection will define the minimal sequences sufficient for N/C ratio-dependent transcription. Candidate transcription factors identified through motif enrichment and enhancer analysis will be experimentally manipulated to test whether altering their concentrations shifts the N/C ratio threshold for gene activation, both at individual loci and on a genome-wide scale. The outcomes will provide new mechanistic understanding of how cells measure their N/C ratio and coordinate transcriptional activation during a critical developmental stage. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2026 · 2026-05

Abstract: Our laboratory has spearheaded the idea of engineering cytotoxic T cells (CAR T cells) to reduce activated fibroblasts in the injured heart thus improving cardiac function in animal models of heart failure. We have also shown that we can engineer cytotoxic T cells in vivo using targeted lipid nanoparticle to delivery modified mRNA. This approach is attractive as a potential therapeutic for interstitial fibrosis such as that associated with hypertension and other types of heart disease. However, in the setting of myocardial infarction, CAR T therapy directed at activated fibroblasts may risk increasing the chance of myocardial rupture. Here, we propose to develop an alternative immune- based cell therapy for heart failure associated with myocardial infarction. T regulatory cells (Tregs) suppress inflammation and fibrosis but are not cytotoxic. We propose to generate CAR Tregs that will accumulate in fibrotic, damaged regions of the myocardium and suppress further inflammation to promote healing and improve function. We describe approaches to generate CAR Tregs targeted to activated fibroblasts expressing Fibroblast Activation Protein (FAP) both ex vivo and in vivo using targeted lipid nanoparticles to deliver modified mRNA. We also propose methods to “armor” CAR Tregs to enhance their anti-inflammatory functions.

NIH Research Projects · FY 2026 · 2026-05

Project Summary/Abstract Tissue restoration is essential to maintain skeletal muscles after injury, due to overuse, aging or disease. Repeated cycles of muscle damage and repair are associated with muscle stem cell (MuSC) dysfunction and impaired myogenesis. Telomeres protect and stability and telomeric proteins do not only the end of our chromosomes from deterioration but are main components of the stem cell progenitor cells `ignition' mechanism, which maintain tissue homeostasis and genome by repairing damage throughout life .Although the telomere protective machinery has been primarily established during carcinogenesis and aging; its importance during regeneration and particularly in muscle injuries, a tissue known for its high regenerative capacity and low propensity for carcinogenesis, is not well understood. We previously demonstrated that telomere attrition is a distinct feature of dystrophic MuSCs in both mice and patients, even at very young ages. More recently, we discovered that TRF2, a key telomere-capping protein, is dynamically regulated in skeletal muscles and has distinct functions, independent of its conventional telomeric role. We also developed genetic tools to define how this protein operates in uninjured, injured, and diseased skeletal muscles. The studies proposed here will determine the extent of previously unknown extra-telomeric functions of TRF2 in muscle stem cells (Aim 1), they will define its new role in regenerating myofibers upon acute and chronic injuries and will uncover new interacting proteins during this process (Aim 2). We expect that this project will fundamentally advance our understanding of the molecular mechanisms by which TRF2 maintains stem cell identity versus how it regulates reparative myogenesis and could effectively guide ways of promoting regeneration and function in healthy and diseased conditions.

NIH Research Projects · FY 2026 · 2026-05

Project Summary We seek NIH support for the acquisition of an Asymmetric Flow Field-Flow Fractionation system coupled with Multi-Angle Light Scattering (AF4-MALS) to be housed in the Johnson Foundation Structural Biology and Biophysics Core (JFBSB Core) at the University of Pennsylvania. This platform will provide critical capabilities for the separation and label-free analysis of macromolecules and nanoparticles in solution, including lipid nanoparticles (LNPs), viral vectors (e.g., AAVs), protein-nucleic acid complexes, and phase- separated assemblies. The requested commercial instrument integrates two powerful technologies: 1. field- flow fractionation (AF4) for size-based, non-destructive separation of complex or fragile species and 2. multi- angle light scattering (MALS) for the direct measurement of molar mass, radius of gyration, hydrodynamic radius, and particle size distribution without reliance on calibration standards. Additional detectors, including differential refractive index (dRI), dynamic light scattering (QELS), and ultraviolet-visible absorbance, allow the system to rigorously quantify particle concentration, heterogeneity, and conjugation state in real time. These capabilities are essential for fully understanding the structure-function relationships of therapeutic macromolecular formulations and advancing gene delivery technologies. The proposed system will support the work of 10 NIH-funded projects in structural biology and nanomedicine. Projects will include structural optimization of LNP formulations for nucleic acid delivery, analysis of biologically relevant higher-order protein assemblies and aggregates, and separation of nucleoprotein complexes. This technology complements and enhances existing SEC-MALS, SAXS, and AUC platforms at Penn, enabling orthogonal workflows across the campus research landscape. No equivalent system currently exists at the University of Pennsylvania. The requested AF4-MALS system from Wyatt Technology offers unmatched integration with ASTRA software for advanced analysis, U.S.-based support, trade-in options for legacy systems, and the lowest risk of import-related tariff costs among evaluated vendors. The JFBSB Core, with a strong track record of S10 stewardship, will ensure broad access, expert support, and long-term sustainability. The requested instrumentation will have immediate and wide-ranging impact on federally funded research programs across Penn and its affiliated institutions.

NIH Research Projects · FY 2026 · 2026-05

PROJECT SUMMARY Standard of care treatment for pancreatic ductal adenocarcinoma (PDAC) provides limited benefits to patients, highlighting the need for alternative therapies. Given that >90% of PDAC is driven by KRAS mutations, novel KRAS/RAS inhibitors, including pan-RAS inhibitors (RASi), hold the potential to improve patient survival. Despite objective clinical responses observed in early RASi trials, relapses are nearly universal. My preliminary studies indicate that relapse arises from a subset of cells that can enter a drug-tolerant persister cell state, characterized by negligible growth, which eventually adapt bona fide resistance mechanisms to resume proliferation. However, the molecular mechanisms governing these cell states and the drivers of cell state transitions remain poorly understood. Moreover, RASi studies performed in our genetically homogenous PDAC models, genomic analyses of human PDAC treated with allele-specific KRAS inhibitors, and prior studies of the persister cell state in other cancers underscore the importance of studying non-genetic resistance mechanisms, including epigenetic mechanisms. Traditional resistance studies compare pre- and post-treatment, late-stage disease, failing to capture the cell state trajectories of individual cells on the path to resistance. Studying these dynamic cell state transitions may uncover vulnerabilities in the transient persister cell state or even pre-existing states that dictate survival. Importantly, my preliminary lineage tracing experiments suggest that some lineages are intrinsically primed to resist RASi. Thus, this proposal tests the hypothesis that pre-existing epigenetic states promote persistence (survival) when cells are exposed to RASi, permitting the subsequent emergence of drug resistance and clinical relapse. To test this, I will pair DNA-barcoding with single-cell technologies to trace PDAC through RASi treatment in vitro to gain a time-resolved understanding of clonal dynamics and phenotypes that mediate resistance. In Aim 1, I will assess the potential of targeting pre-existing cell state heterogeneity in PDAC by quantifying the degree to which the state of a cell prior to RASi treatment influences cell fate. In Aim 2, I will study the gene expression and chromatin accessibility profiles of resistant PDAC lineages over time, before and after RASi, to elucidate the patterns and drivers of epigenetic plasticity. Ultimately, this work will provide critical insights into the mechanisms enabling PDAC progression to resistance, which will inform the development of combination therapies to prevent RASi relapse and effect a cure.

NIH Research Projects · FY 2026 · 2026-05

Project Summary Periodontal disease, caused by bacterial infection and inflammation of the gums and supporting bone, affects nearly 50% of U.S. adults over the age of 30, with approximately 9% suffering from severe periodontitis. The hallmark of this disease is the progressive destruction of alveolar bone, ultimately leading to tooth loss and significant oral disability. Periodontitis is particularly prevalent among older adults and is associated with increased risk for systemic conditions such as atherosclerosis, rheumatoid arthritis, and diabetes mellitus. The disease arises from polymicrobial dysbiosis that disrupts the balanced oral microbiota, alongside impaired host innate immune responses that contribute to tissue destruction. While the roles of microbial communities and immune dysfunction in periodontitis are well established, the specific gingival cell types involved in protective responses remain poorly understood. Recent studies have identified taste-like chemosensory cells—solitary chemosensory cells (SCCs) and tuft cells—in various mucosal tissues as key sentinels that detect microbial signals and initiate innate immune responses. Building on this, our preliminary work has identified SCCs and macrophages (MΦ) in the mouse gingiva as components of the epithelial barrier that protect against bacterial invasion. Both cell types express bitter taste receptors and associated signaling components, respond to bitter and bacterial stimuli, and activate innate immune pathways that mitigate periodontal inflammation and bone loss. This proposal aims to expand upon these findings by: • Aim 1: Investigating how taste signaling in gingival macrophages contributes to both the pathogenesis and protection mechanisms in periodontal disease. • Aim 2: Identifying the protective mechanisms by which gingival SCCs and MΦ reduce periodontal bone loss in response to bitter compound stimulation. Overall, this study represents a significant advancement in our understanding of periodontal disease pathophysiology. Activating SCCs and MΦ through bitter compounds offers a novel, non-invasive therapeutic strategy. Furthermore, this approach holds promise for clinical translation through advanced delivery systems such as nanoparticles and hydrogels.

NIH Research Projects · FY 2026 · 2026-05

SUMMARY Heart failure is a leading cause of death worldwide, and the leading cause of hospital admissions in patients over 65 in the US. The role of metabolism in cardiac pathology has long been of interest, but no therapies that target cardiac metabolism are known. Recently, Na+/glucose transporter 2 inhibitors (SGLT2i), originally designed as therapies for diabetes, have taken HF management by storm, demonstrating 20-30% reductions in heart failure hospitalizations in large phase III trials, even in patients without diabetes. How SGLT2i confer these benefits remains a mystery. In extensive preliminary data, we now show that SGLT2i directly activate pantothenate kinase (PANK), the first and rate-limiting enzymatic step in the synthesis of co-enzyme A (CoA) from pantothenate (vitamin B5). CoA is an obligate intracellular metabolite critical for most oxidative processes, especially in highly oxidative tissues like the heart. We show in our preliminary data SGLT2i directly promote cardiac CoA synthesis and oxidative metabolism, and promote cardiomyocyte contractility. Together our data lead us to hypothesize that: SGLT2i confer benefit in heart failure by activating PANK and boosting cardiac CoA metabolism. We will test this hypothesis in depth by: Aim 1: Understand the consequences of SGLT2i on cardiac CoA metabolism and fuel use. Aim 2: Test in vivo the role of cardiac PANK in the heart failure protection conferred by SGLT2i. Aim 3: Identify the molecular mechanism of activation of PANK by SGLT2i. Aim 4: Begin a discovery campaign of novel small molecules to target PANK but not SGLT2. These highly focused studies will elucidate how SGLT2i target PANK, and in turn protect from heart failure. Precise structural and physiological understanding of how SGLT2i target PANK will enable design of small molecules that improve on SGLT2i efficacy and reduce the side effects of SGLT2i. Success in these studies will thus potentially open the door for the generation of entirely new classes of drugs to treat heart failure.

NIH Research Projects · FY 2026 · 2026-05

PROJECT SUMMARY Acute myeloid leukemia (AML) is a genetically and cellularly heterogenous disease characterized by the expansion of hematopoietic cells across a range of cell states from stem-like cells to differentiated myeloid cells. The most mutated genes in AML are DNMT3A, NPM1 and the receptor tyrosine kinase FLT3. Despite early clinical responses, most patients relapse, and FLT3-mutant clones are not always eradicated. Our lab has developed genetically engineered mouse models of acute myeloid leukemia that are capable of activating mutations in Flt3 with Dre-recombinase, and then genetically reverting them with Cre-recombinase. We have used these models to benchmark Flt3 oncogene-addiction against best-in-class small molecule kinase inhibitors of FLT3, observing difference in disease remission and relapse. These studies have refined our interest on identifying which cells along the hematopoietic hierarchy are capable of driving relapse and which molecular pathways underlie their survival following chemical/genetic inhibition of FLT3. The major goal of this proposal is to understand the cellular mechanisms that maintain FLT3-mutant clone persistence during targeted therapy. Our preliminary data indicate that Flt3-inhibtion results in a profound differentiation response and induction of Type I Interferon signaling. We will complete integrative studies with human specimen and our innovative multi- recombinase mouse models of leukemia to derive clinically meaningful insights from mechanistic observations in model systems. In aim 1 we will determine which cells are capable of propagating leukemic disease and resolve cellular reservoirs of leukemic stem cell activity. We will perform these studies using genetically engineered mouse models, serial transplantation of purified cell populations, and functional cell ablation studies. We hypothesize that FLT3-inhibtion induced differentiation generates mature cells that are capable of reacquiring stem-like properties and drive relapse. These studies will resolve which cells are necessary to eliminate to prevent leukemic recurrence and provide a focusing lens for improving targeted therapy and relapse detection. In aim 2 we will determine the role of Type I Interferon signaling in differentiation and relapse using gain/loss of function systems. We will evaluate the therapeutic potential of interferon treatment in conjunction with FLT3 kinase inhibition. Finally, we will assess the clonal diversity of leukemic cell states using lentiviral barcoding and single cell RNA sequencing to evaluate which cells can induce an Interferon response, and what their long-term fate is following treatment. We hypothesize that Interferon signaling is necessary to potentiate FLT3-inhibitor driven differentiation, and that combined treatment will extend survival. We anticipate that these studies will more broadly inform the intersection between inflammation and differentiation in AML therapy.

NIH Research Projects · FY 2026 · 2026-05

Project Summary The staging model for human type 1 diabetes (T1D), proposed by Eisenbarth nearly 40 years ago, and reaffirmed recently, posits that disease progression for individuals with genetic susceptibility depends on a triggering event. The nature of this triggering event remains a crucial open question in T1D. Evidence shows an IFN response in pancreatic islets during early T1D stages, potentially triggered by viral infections. However, recent studies have not found definitive proof of a viral cause. Instead, the IFN response could also originate from non-viral sources. MDA5, a cytosolic dsRNA sensor, triggers an IFN response when detecting virus-like long dsRNA. Human genetic evidence shows that Gain-of-Function (GoF) mutations of MDA5 cause enhanced IFN activation, while Loss-of-Function (LoF) mutations protect against multiple autoimmune diseases, including T1D. ADAR1, an enzyme expressed in virtually every cell in our body, edits endogenous dsRNAs by converting adenosine to inosine to prevent MDA5 activation on “self” dsRNAs. Mutations in ADAR1 cause severe autoinflammatory diseases, suggesting its importance in regulating immune response. Study showed that loss of ADAR1 in mouse β cells triggers a significant IFN response, leading to inflammation and β cell destruction, mimicking early-stage T1D in. Our recent work, based on human genetic data, made a remarkable discovery of the role of RNA editing in T1D. We demonstrated that insufficient RNA editing caused by risk variants of T1D is linked to elevated IFN immune response observed in pancreas and immune cells of T1D patients. Together, human and mouse genetic data strongly suggest poorly edited endogenous dsRNAs as a trigger of T1D. Using human genetics, genomics, and molecular biology approaches, we propose to investigate the functional connection between RNA editing and T1D in humans. First, we will investigate RNA editing in T1D and non-T1D islet in a cell-type-specific manner. Second, we aim to determine the key islet cell types whose inadequate RNA editing triggers MDA5-dependet IFN response. Taken together, our proposed studies will identify a new pathway underlying T1D and novel therapeutic targets for delaying T1D onset.

NIH Research Projects · FY 2026 · 2026-05

Substance use disorders (SUDs) are characterized by long-lasting neuroadaptations, including changes in gene expression and the epigenome. The relevance of the epigenome in SUDs has been shown using overexpression or inhibition of epigenome modifying enzymes. However, such manipulations cannot distinguish between direct and indirect action of epigenetic modifications at a target gene. To address this, we apply locus-specific epigenetic editing with CRISPR/dCas9 to establish the direct causal relevance of a specific epigenetic modification to SUDs. We and others find that the epigenetic enzyme, SET domain containing 2 (Setd2), regulates cocaine-induced mouse behavior, gene expression, and alternative splicing. Setd2 specifically writes H3K36me3, a histone modification associated with active transcription and alternative splicing, which is altered in mouse brain regions by cocaine exposure. We and others have identified a role for Setd2/H3K36me3 in alternative splicing which mirrors cocaine-induced splicing events and is associated with cocaine-related behaviors in mice. We find that overexpression of Set2 in mouse brain reward regions increases H3K36me3 and drives mouse cocaine reward behavior. Together, these data provide strong evidence for Setd2/H3K36me3 in mediating the behavioral response to cocaine in mice through epigenetic and alternative splicing events. The overall goal of this proposal is to establish that Setd2 inhibition attenuates cocaine self-administration through cell-type specific regulation of alternative splicing and gene expression. First, we will determine both sufficiency and necessity of Setd2 to drive cocaine behavior, using pathway-specific Setd2 overexpression and small-molecule Setd2 inhibition (Setd2i), respectively. We hypothesize that NAc Setd2 inhibition will reduce cocaine self-administration. Second, we will define the pathway-specific effects of Setd2 on behavior, H3K36me3 and gene expression by cell-type specific epigenomic profiling and targeted epigenetic editing. Third, we will interrogate mechanisms by which Setd2 regulates alternative splicing by identifying interacting RNA-binding proteins (RBPs). Finally, this proposal will investigate non-enzymatic roles of a histone modifying enzyme in addiction. Recent studies and our preliminary data suggest non-enzymatic function of Setd2 through protein binding domains. We will distinguish Setd2 and H3K36me3 using combinatorial Setd2 overexpression, Setd2i, and epigenetic editing.

NIH Research Projects · FY 2026 · 2026-05

Alcohol Use Disorder (AUD) affects multiple body organ systems, but in the brain, it is a circuit disease spanning multiple regions and influenced by environmental and hereditary factors. The neuronal, glial, and microglial components of these circuits respond to ethanol (EtOH) challenge in myriad ways, including changes in gene transcription. EtOH-induced changes in behavior are mediated in part by RNA transcription and alterations in chromatin structure, which regulate gene accessibility for transcription. The process of RNA transcription requires chromatin to be in an open conformation with less nucleosome packing, allowing the transcription regulatory enzymes to function. Dynamic changes in chromatin structure occur over an organism's lifetime, permitting behavioral adaptation to ever-changing environmental stimuli, including those induced by pharmacological agents such as EtOH. This grant focuses on identifying the genomic consequences of alcohol abuse in identified EtOH circuit-associated cells within their natural brain microenvironment to better understand AUD. We propose to assess the cellular multi-genomics of chronic EtOH responsiveness by simultaneously quantitating the open- chromatin landscape and cytoplasmic transcriptome of in situ tissue-localized single neurons, astrocytes, and microglia from EtOH-challenged mice. To assess these ‘omics outcomes in cells within their natural brain environment, we will examine single cells in fixed brain tissue where neuronal/cellular associations are chemically fixed and immobilized. To experimentally accomplish this, we developed light-activated oligonucleotide probes that can be directed to the nucleus and cytoplasm of fixed cells. Upon light activation, the probe acts as a primer for in situ copying of nuclear single-stranded DNA (open chromatin) and/or cytoplasmic RNA (transcriptome) into barcoded complementary DNA that can then be sequenced. We will use these approaches to molecularly characterize spatially localized immunocytochemically identified single cells resident in brain tissue sections from behaviorally and physiologically characterized mice that have been chronically challenged with EtOH using a self-administration and withdrawal paradigm. While the current studies will utilize the experimentally tractable mouse model system, the development and optimization of these approaches will facilitate the omics analysis of human brain fixed tissue samples from well-characterized subjects with AUD available from brain banks. We anticipate that these multi-modal studies, coupling organismal behavior and physiology with omics responses in spatially localized single cells, will detail co-regulatory pathways that define the cellular mechanisms underlying AUD, thereby providing novel testable therapeutic targets.

NIH Research Projects · FY 2026 · 2026-05

Summary The overarching goals of this proposal are to understand how the transcription factor RUNX1 restrains an inflammatory epigenetic program in granulocyte-monocyte progenitors (GMPs) that dampens inflammatory cytokine production by neutrophils. Monoallelic mutations in RUNX1 cause familial platelet disorder with predisposition for myeloid malignancy (FPDMM). FPDMM patients have an increased incidence of clonal hematopoiesis, and the vast majority patients have inflammatory and immune disorders. We hypothesize that increased inflammation in patients contributes to accelerated clonal hematopoiesis and elevated risk of leukemia. We discovered that RUNX1 loss in GMPs results in the overproduction of inflammatory cytokines and chemokines by neutrophils in response to the toll like receptor 4 (TLR4) ligand lipopolysaccharide. We believe this is caused by the elevation of tonic type I interferon signaling in GMPs which sensitizes other innate immune signaling pathways in downstream myeloid cells. Mechanistically, we will determine how RUNX1 regulates type I interferon signaling in GMPs, and whether elevated tonic type I interferon signaling is responsible for increased retroelement accessibility observed in RUNX1 mutant GMPs and neutrophils. We will also determine whether elevated inflammation in RUNX1 mutant mice augments innate immune training. To obtain human relevance for the studies in mouse models, we will determine whether GMPs and neutrophils in FPDMM patients have an epigenomic signature of neutrophil activation, if there are differences in the proportions of neutrophil subpopulations, and if neutrophils hyper-respond to activating signals.

NIH Research Projects · FY 2026 · 2026-05

Project Summary The goal of this project is to carry out in vivo studies to understand how type-1 and type-2 diabetes alter oral and skin wound healing. Mechanistic studies are proposed based on exciting Prel Data that blocking permissive histone-3 methylation completely rescues the negative impact of diabetes on gingival and skin healing. These experiments will utilize a highly specific inhibitor, MM-102, that is effective in nanomolar concentrations. A secondary goal is to determine whether the cellular and molecular impact differs in oral and dermal wounds. The proposed experiments will utilize a number of advanced approaches including Chromium single cell RNAseq and Xenium spatial transcriptomics from 10x Genomics. They will also utilize cell sorting followed by CUT&RUN H3K4me3 analysis combined with bulk RNAseq. The different approaches will be integrated through bioinformatic analysis that will facilitate an unbiased examination of diabetes-impaired wound healing combined with a hypothesis-driven mechanistic investigation. In addition, we will carry out translational studies in a large mini-pig animal model to address the therapeutic use of the inhibitor. The experiments are significant since epigenetic changes that regulate specific genes play a key role in "glycemic memory" that alters cell behavior and are grounded in strong Prel Data. A better understanding of this specific epigenetic change and how it regulates gene expression could greatly improve our understanding of diabetic complications and lead to the identification of new therapeutic approaches to diabetic wound healing. Aim 1 will focus on a mechanistic murine studies involving the inhibitor MM-102. Aim 2 encompasses translational studies to determine whether the MM-102 inhibitor improves wound healing in a large, well-established T2D mini-pig animal model. The investigative team includes experts in diabetic wound healing, oral wound healing, skin wound healing, bioinformatics, spatial transcriptomics and murine and porcine diabetic wound healing models.

NIH Research Projects · FY 2026 · 2026-04

PROJECT SUMMARY/ABSTRACT The University of Pennsylvania Abramson Cancer Center (ACC) has been highly engaged in the National Cancer Institute (NCI) National Clinical Trials Network (NCTN) for many years and has demonstrated an extraordinary commitment to the mission of the NCTN. The ACC has fostered an extensive cooperative group membership with many investigators serving in group or NCI committee leadership roles (44 group committee chair positions, 34 NCI committee roles, 16 study chairs, 14 study co-chairs, and nine study champions), robust accrual to NCTN clinical trials, leadership of large national trials, delivery of innovative ideas to the NCTN, and mentorship and career development of young investigators. The ACC has made a significant investment in the NCTN through the development of a centralized clinical research infrastructure focused exclusively on the conduct of NCTN trials—the NCTN@ACC. The leader of this enterprise and the PI of this LAPS grant is Dr. Jennifer Eads, Professor of Medicine in medical oncology. She has extensive experience in clinical trials, including through NCTN, for which she was named ECOG-ACRIN Investigator of the Year (2022). As Director of NCTN@ACC, Dr. Eads oversees the centralized NCTN administrative, operational, regulatory, auditing, monitoring activity, and financial aspects of the program. The NCTN@ACC is supported by five NCTN@ACC staff and three NCTN committees, all of which provide critical roles in the visionary and operational aspects of the program. Ensuring access to and representation of the communities in our catchment area in NCTN trials is a major effort across departments and the University of Pennsylvania Health System network of hospitals, led by Dr. Eads. The NCTN@ACC collaborates extensively with multiple research centers, scientific research programs, core facilities and the ACC Cancer Service Line to bring innovative ideas from these groups into the NCTN for further development of clinical trials. The NCTN@ACC is strongly committed to career development of young investigators through multiple mentoring mechanisms, including exposure to the NCTN where scientific and leadership contributions are encouraged. Specifically, NCTN@ACC aims to (1) lead an expansive NCTN clinical research program at the ACC through an efficient, centralized enterprise that enables rapid enrollment across the Penn network and the conduct of high-quality clinical research with a focus on outreach to the community; (2) drive exceptional leadership and innovation of NCTN clinical research at the national and institutional level; and (3) expand the ACC commitment to mentorship and career development of faculty and staff, as well as enrollment of populations across our catchment area including patients with rare cancers.

NIH Research Projects · FY 2026 · 2026-04

Abstract: Abnormal metabolism is characteristic of nearly all disease states, whether in pathway or extent, and often precedes changes accessible to anatomical medical imaging. Although several methods have been proposed and implemented to realize this potential for sensitivity to early disease, or to the immediate impact of a therapeutic regimen, each has a shortcoming that prevents metabolic imaging from being the general purpose diagnostic tool with broad clinical or research impact. This is equally true of solution state hyperpolarization, an MRI-based technique that enhances the signal per nucleus of selected biomolecules by >10,000x. Despite the large sensitivity gains, the technology of hyperpolarization has thus far been limited to rapid administration of high concentrations of imaging agent, limiting the accessible set of agents along with the amount that can be administered and metabolized in the short time during which the hyperpolarized state remains. In this project, we build on recent technical advances to produce a low cost, continuous flow device capable of producing highly polarized agents of clinical importance, at concentrations suitable for continuous infusion into in vitro cell culture and small animal preclinical experiments. We note that this scheme 1) maximizes metabolic signal by avoiding transporter and metabolic enzyme saturation, as well as cofactor pool depletion, while supporting greater safe total doses of the administered agent, and 2) simplifies analysis by operating under steady state conditions and allowing practical implementation of traditional MRI contrast techniques (e.g., metabolite signal saturation/inversion, diffusion encoding, etc.). We then demonstrate this utility in a series of studies in the SNU-449, comparing the metabolism of the adherent, perfused cells under a variety of conditions to similar results in induced tumors in immunocompromised mice. These studies, at `tracer' (e.g., relevant to normal physiological) concentrations will be the first of their kind using hyperpolarized agents, and are made possible using the apparatus developed in the first aim. Justified by these initial results, we anticipate that scaling up to large animal or human operation will be straightforward, and that eventual clinical adoption may be made easier by the ability to verify agent safety in steady state before infusion.

NIH Research Projects · FY 2026 · 2026-04

PROJECT SUMMARY The prevalence of Alzheimer’s disease (AD) and related dementias (ADRD) in the United States is on the order of seven million and likely to grow over the next two decades. The molecular hallmarks of ADRD are misfolded proteins (b-amyloid and tau), and although anti-amyloid antibody drugs have recently been approved for AD treatment, they have limited effectiveness, are often associated with significant side effects, and amyloid depo- sition correlates only moderately with cognitive status. This situation has spurred the search for additional mech- anisms underlying the disease. Glucose is the main substrate for ATP synthesis in the brain. A well-known feature of ADRD is disruption of the brain’s energy metabolism, which has been linked to defective processing of, or reduced access to, glucose resulting in brain hypometabolism. It is also known that ketones (b-hydroxy- butyrate and acetoacetate) formed in the liver from medium-chain length fatty acids can act as an alternate fuel for oxidative phosphorylation. Non-invasive assessment of the cerebral metabolic rate of oxygen (CMRO2) glob- ally and regionally would allow the age and neurodegeneration related brain energy gap to be quantified and validate ketogenic intervention by quantitative brain imaging. Currently, positron emission tomography (PET)- based brain oximetry relying on oxygen-15 tracers is considered the “gold-standard” imaging method for mapping cerebral energy O2 metabolism. But 15O PET is complex, costly, and not widely available. Over the past decade magnetic resonance imaging (MRI) methods have emerged for measuring CMRO2 noninvasively. Both PET and MRI derive CMRO2 from measures of venous and arterial O2 saturation to estimate oxygen extraction fraction (OEF) and, along with cerebral blood flow (CBF), CMRO2. The proposed research builds on recent developments in the applicants’ lab for 3D CMRO2 mapping via a new constrained qBOLD technique based on an extension of the Yablonskiy model for signal decay due to partially deoxygenated hemoglobin in the capillary network, in combination with quantitative susceptibility mapping. We propose to first enhance the method in terms of image acquisition efficiency and to examine its sensitivity to detect regional variations in OEF, CBF, and CMRO2 both at baseline and in response to physiologic stimuli in test subjects. Subsequently, we will investigate the growing energy gap with age and, more so, in ADRD, and evaluate the hypothesis in a small group of AD patients that ingestion of a ketone ester drink in the form of triacyl triglycerides of C-8 saturated fatty acids induces a transient increase in O2 metabolism and, possibly, acute improvement of cognition. This new MRI-based imaging tech- nology for brain oximetry can readily be integrated into standard brain imaging protocols to provide means for evaluation of the metabolic consequences of ADRD at baseline and in response to intervention.