University Of Pennsylvania

NIH Research Projects · FY 2025 · 2024-03

ABSTRACT The subgenual anterior cingulate cortex (sgACC) has dominated the literature utilizing repetitive transcranial magnetic stimulation (rTMS) to treat depression for good reason: treatment focusing on the frontal-sgACC brain circuit is effective and changes in resting fMRI connectivity with the sgACC accompany symptom improvements. However, it has not yet been proven that TMS actually engages the sgACC which would support continued focus on this circuit at the exclusion of other potential targets from the fMRI literature. With the application of interleaved TMS/fMRI, it is possible to capture brain responses caused by TMS to any brain surface site. Using resting fMRI connectivity to guide TMS targeting, we seek to establish neuromodulation of the sgACC pathway (R61, Aim 1) and the best connectivity pipeline for robustly engaging the sgACC (R61, Aim 2). Upon establishing these successful benchmarks, the R33 phase will demonstrate that the sgACC evoked response at baseline predicts depression improvement when treatment delivered to this same circuit (R33, Aim1). To cement the relationship between modulation of the frontal-sgACC circuit with treatment and depression improvement, we hypothesize an association between brain changes and clinical changes (R33, Aim 2). We hypothesize that the R33 clinical associations with evoked brain responses will be established for active but not for sham rTMS. Together, this research will elucidate basic mechanisms of rTMS treatment that will accelerate brain circuit targeted neurotherapeutics.

NIH Research Projects · FY 2026 · 2024-03

PROJECT SUMMARY Multiple lines of clinical and experimental evidence suggest that seizures in early life can be associated with long lasting cognitive and behavioral deficits. In rodent models, we have showed that early life seizures (ELS) impair normal synaptic plasticity, critical period plasticity, later life learning and social behavioral deficits. Improved understanding as to how seizure activity and hyperexcitable networks dysregulate synaptic plasticity will be required to develop new therapeutic strategies in this clinical space where no current cure exists. We will focus on dysplasticity related to alterations of the excitatory synaptic glutamate receptor and its related signaling pathways. Tracking alterations of synaptic glutamate receptors in neurons activated by ELS is a specific challenge given that they occur in the midst of the synaptic critical period, the refinement of synaptic connections and the dispersion of neurons with development, which makes it difficult to localize neurons for functional studies later in life, despite the persistence of impaired synaptic plasticity and cognitive deficits. Similarly, sampling of a neuronal population for gene and protein expression may fail to show alterations occurring in a small, critical, subset of cells. To address these issues, we have adapted a method to permanently label cells activated by ELS in mice so that we can measure synaptic responses, gene and protein expression at a single neuron level, and then differentially label them during subsequent later life seizure (LLS) events. Using our ELS models, we aim to determine whether neurons activated by ELS have persistent, life-long, alterations of glutamate receptor function associated with impaired synaptic plasticity and hyperexcitability compared to neurons from no-seizure control mice (Aim 1). We will correlate these functional changes with measurements of gene and protein expression related to glutamate receptor function compared to neurons from no-seizure control mice (Aim 2). Finally, we will determine whether neurons activated by ELS are differentially affected by a second later life seizure (LLS) in adulthood compared to control neurons in seizure free mice (Aim 3). The synapse is a convergence point for the likely many upstream derangements of network function, and therefore an ideal target of study. We hypothesize that tracking the evolution of changes over time in select neuronal populations following ELS will allow us to both “stage” the evolution of changes and identify new therapeutic targets for this comorbidity and consequence of seizures in the immature brain.

NIH Research Projects · FY 2026 · 2024-03

Project Summary Fragile X Syndrome (FXS) is caused by expansion of the CGG short tandem repeat (STR) located in the 5’ UTR of the FMR1 gene. Upon expansion to mutation-length, local DNA methylation at the FMR1 promoter leads to silenced transcription which is thought to drive the pathophysiology of FXS. However, Fmr1 knock-out mice do not reproducibly recapitulate the range of FXS clinical presentations, suggesting that FMR1 dysregulation alone cannot explain the pathophysiology of FXS. Recently, our lab uncovered Megabase-scale domains of the histone modification H3K9me3 at the FMR1 locus on chromosome X and multiple autosomes. The H3K9me3 domains encompass silenced genes encoding synaptic plasticity and epithelial integrity, which correlate with symptoms experiences by FXS patients, raising the possibility that these heterochromatin domains contribute to the pathophysiology of FXS. The objective of my proposal is to investigate the mechanisms by which the mutation- length CGG STR coordinates Megabase-scale heterochromatin domains and their inter-chromosomal contacts. My central hypothesis is that expansion of CGG STR to mutation-length is necessary and sufficient for the heterochromatinization of the FMR1 locus and a subset of autosomal domains. Upon heterochromatinization, the domains form pathologic inter-chromosomal contacts with each other in a H3K9me3-dependent manner. I have formulated my hypothesis based on my unpublished imaging data demonstrating that (1) ectopic trans interactions form between H3K9me3 domains in induced pluripotent stem cells (iPSCs) with the mutation-length CGG STR tract, (2) in single cells, FXS domains that form inter-chromosomal contacts are more enriched in H3K9me3 than those that do not, and (3) cut-back of the CGG STR can reverse H3K9me3 signal at a subset of domains. I will test my hypothesis by leveraging a newly developed protocol for STR synthesis with CRISPR/Cas9 engineering to generate iPSC clones with the same genetic background but varying STR length and sequence at the 5’ UTR of FMR1. I will measure the effect of the mutation-length CGG STR, as well as the overexpression of H3K9me3 writer and eraser enzymes, on H3K9me3, trans interactions, and transcriptional silencing using state-of-the art genomics and multimodal imaging techniques including CUT&Run, Hi-C, sequential Oligopaint FISH, and single-molecule RNA FISH. Successful completion of these experiments will demonstrate the contribution of both STR sequence and length to the multi-chromosomal, Megabase-scale heterochromatinization of the FXS genome and defined the requirement for H3K9me3 for trans interactions. My work is significant because it will expand classic models of how the mutation-length CGG STR causes FXS to include Megabase-scale heterochromatinization and ectopic inter-chromosomal contacts. Studying these mechanisms will have a broad impact on our understanding of the role for heterochromatin and miswiring of the 3D genome in a wide range of repeat expansion disorders and in cancers with repeat instability.

NIH Research Projects · FY 2026 · 2024-03

PROJECT SUMMARY: Obesity and its comorbidities pose an ever-increasing challenge to public health despite massive investments in pharmacologic, surgical, and lifestyle-modifying therapeutic strategies. New strategies are needed to alleviate the worsening metabolic health of the national and global populations. One promising strategy is to harness the innate calorie-burning properties of brown adipose tissue (BAT), a metabolic organ specialized for the conversion of chemical energy to heat. Although active BAT is highly correlated with metabolic health in humans, its overall prevalence is low and declines with age and obesity. BAT-targeted therapeutics will thus require the generation of increased BAT mass. Therefore, it is imperative to understand the physiologic development of BAT. BAT derives from the dermomyotome (DM), a multipotent embryonic tissue that also gives rise to skeletal muscle and dermis. The goal of the current proposal is to define the embryonic progenitor cells in the DM-to-BAT lineage and identify the molecular mechanisms controlling the specification and differentiation of brown adipocytes. Preliminary work has identified candidate progenitor cell populations marked by expression of Cdh4 and Dpp4, respectively, and confirmed that these cell populations derive from the DM. The first aim of this proposal will employ adipogenic differentiation assays and lineage tracing to determine whether Cdh4+ and Dpp4+ cells develop into brown adipocytes in vitro and in vivo. In addition to their potential role as progenitor cells, Dpp4+ cells encapsulate developing BAT and express several signaling molecules, including the brown adipogenic factor BMP7, that may regulate BAT development. Preliminary studies show that GATA6, a transcription factor expressed in Dpp4+ cells, is essential for embryonic BAT development. The signaling genes most enriched in Dpp4+ cells are regulated by GATA6 in other developmental contexts, suggesting that GATA6 may promote BAT development by regulating signaling in Dpp4+ cells. Thus, the second aim of this proposal will test the hypothesis that Dpp4+ cells secrete BMP7 and other signaling factors in a GATA6-dependent manner to promote BAT development. Completion of the proposed work will elucidate the developmental trajectory and molecular mechanisms underlying BAT development, uncovering new cellular and molecular targets for BAT-directed therapeutic interventions. Importantly, the fellowship applicant conducting these studies, Ethan Fein, will obtain rigorous research training that is integrated with his clinical training as a student in the Medical Scientist Training Program (MSTP). His training goals will be accomplished through a comprehensive training plan developed by Ethan and his sponsor, Dr. Patrick Seale. Ethan will benefit from the outstanding training environment provided by the Seale lab, the MSTP, and the University of Pennsylvania as a whole. The research training plan outlined in this fellowship application will maximize Ethan's potential to develop into a successful physician-scientist.

NIH Research Projects · FY 2026 · 2024-03

PROJECT SUMMARY The goal of this proposal is to determine how the crosstalk between RNA and chromatin shapes epigenetic states in pluripotent and differentiated cells. My laboratory studies epigenetic memory, both at a mechanistic level, using biochemistry and functional genomics in mouse embryonic stem cells, and at an organismal level, using ants and flies as model systems. In the nine years since the lab opened, our mechanistic work has been recognized for our contributions to understanding the role of protein–RNA interactions in chromatin regulation and in particular the Polycomb pathway. We have developed technologies to map RNA-binding sites on specific protein complexes or the entire proteome using photocrosslinking and mass spectrometry, and used this information to dissect the functional contribution of RNA to epigenetic pathways. We also established an acute depletion system in embryonic stem cells that allow us to interrogate epigenetic dynamics during cell fate transitions. In the next five years, we will build on these studies and develop new systems and technologies to obtain a deeper understanding of the RNA foundations of chromatin regulation and epigenetic phenomena in mammalian cells. Using a directed differentiation system from embryonic stem cells to neurons combined with acute protein and RNA depletion technologies, we will determine the order of biochemical events that culminate in Polycomb-mediated silencing and the role of RNA at two critical steps: Polycomb target selection and gene repression via Polycomb body formation. Expanding on our protein–RNA studies with mass spectrometry, we will study more broadly the role of different classes of RNA and their chemical modifications in chromatin factor recruitment and design separation-of-function mutants using a newly developed high-throughput mutational screening method. Finally, we will follow up on some intriguing preliminary findings from a genome-wide knockout screen to explore a new direction, the molecular mechanism by which mobile RNAs are selected for incorporation into extracellular vesicles and transfer to recipient cells, where they might exert epigenetic functions. This work will add to our mechanistic understanding of RNA-mediated regulation of chromatin processes, which in turn will provide new opportunities to decode and engineer epigenetic states, with broad impact on research, biotechnology, and medicine.

NIH Research Projects · FY 2026 · 2024-03

Abstract. While CT remains the most highly utilized diagnostic tool in clinical practice, its technology still calls for research and development along the entire imaging chain to improve diagnostic accuracy and patient outcomes. The clinical translation of such developments must be safe, efficient, and timely; however, it remains a challenge to the community how to precisely predict performance during and after the development phase. Technology development can be aided by CT phantoms, which are specialized tools used to calibrate, test, and evaluate scanners. Most existing CT phantoms are relatively expensive and lack accurate representations of anatomy and diagnostic tasks. There is a lack of patient-based phantoms that fully represent attenuation profiles and textures seen in clinical CT acquisitions, which this proposal aims to address. As a result of its ability to create accurate and detailed physical models at a fraction of the cost of traditional methods, three-dimensional (3D) printing has become increasingly popular in medicine. CT phantom 3D printing studies include manufacturing geometrically accurate organ models, generating realistic texture samples, and generating accurate attenuation profiles. Although these approaches produce phantoms that are more similar to actual anatomical structures, several limitations remain, such as the loss of the natural look and feel of anatomical and pathological features. Recently, we proposed a 3D printing solution, called PixelPrint, that can achieve accurate organ geometry, image texture, and attenuation profiles while eliminating the complexities and limitations of previous methods. Our solution is a one-step method for translating CT or simulated images into printer instructions. As a preliminary study shows, isolated organs can be replicated so lifelike that an expert reader can't tell the difference between the CT scan and the original. Our proposal aims to develop 3D printing hardware and software that will be capable of creating patient-based phantoms with accurate spectral x-ray characteristics and a natural look and feel of anatomical and pathological structures. By completing the following aims, we aim to provide a more efficient and cost-effective method for developing and validating novel CT technology: (i) to design and construct a dedicated multi-material, quad extruder 3D printer for CT phantoms, (ii) to develop algorithms to preprocess and translate spectral CT images into instructions for 3D printing, and (iii) to evaluate the performance and reproducibility of patient-based CT phantoms. The academic and clinical CT community will benefit from a rapid and inexpensive manufacturing process. To drive dissemination of our development, we will, as part of this project, distribute dedicated patient-based phantoms to academic institutions. Our research environment and our team’s unique breadth of expertise are perfectly placed to execute this project. By utilizing the proposed phantoms, the community will be able to facilitate the translation of novel CT technologies for various diagnostic tasks into clinical practice.

NIH Research Projects · FY 2026 · 2024-03

Project Summary: Amylin signaling decreases food intake and gastric emptying in both humans and rats via activation of CNS amylin receptors (calcitonin receptor, CTR; heterodimerized with a receptor activating modified protein, RAMP1-3) making it a potential target for the development of novel pharmacotherapies to treat obesity. Despite the distributed nature of CNS amylin receptors, research on the role of amylin signaling in the control of energy balance has been largely focused on hypothalamic and hindbrain nuclei, leaving other nuclei with abundant CTR expression such as the mesopontine laterodorsal tegmental nucleus (LDTg) understudied. Recent work from our lab showed that LDTg CTR signaling reduces food intake. Additionally, in the absence of endogenous LDTg CTR signaling, rats show increased food intake and body weight gain. However, the mechanism by which CTR signaling in the LDTg modulates energy balance, as well as the downstream nuclei targeted by LDTg CTR expressing neurons (LDTgCTR) has not been explored. The LDTg is known to modulate dopamine signaling to regulate motivated behavior via direct projections to the ventral tegmental area (VTA) and our preliminary data shows that several LDTgCTR neurons send direct projections to the VTA. Consequently, we will use chemogenetics to activate LDTgCTR neurons that project to the VTA and evaluate the effect of this manipulation on feeding and motivation to obtain palatable food rewards (Aim 1). We will also use a dual AAV approach to projection-specifically knockdown CTR expression in LDTg neurons that project to the VTA and evaluate the role of these neuron’s endogenous CTR activity in the regulation of energy balance and palatable food reward seeking motivation (Aim 2). Lastly, we will use fiber photometry to monitor the activity of LDTgCTR neurons in lean and obese animals in response to food availability at different energetic states. Then, we will determine how diet-induced obesity affects the ability of LDTgCTR neurons to modulate downstream VTA dopaminergic (VTATH) neurons (Aim 3). Altogether, we hypothesize that this novel mesopontine-limbic signaling pathway will have the ability to reduce food intake and body weight without inducing symptoms of malaise, via modulation of downstream VTATH neurons. Consequently, the proposed work will functionally characterize a promising target for the development of effective obesity pharmacotherapies.

NIH Research Projects · FY 2026 · 2024-03

PROJECT SUMMARY/ABSTRACT Deletion of a region of Chromosome 22q11.2 (22qDS) encoding over 40 protein-coding genes is the most common microdeletion syndrome (~1/2000 live births) and predisposes to multiple neurodevelopmental disorders (NDDs). Individuals with 22qDS display microcephaly and 22qDS models suggest deficits in neural stem and progenitor cell (NSPC) proliferation may be contribute, though the complement of genes and mechanisms involved are not known. Six of the genes in the 22qDS deleted region encode mitochondrial proteins and mitochondria are important regulators of neurogenesis, suggesting loss of these genes may contribute to disturbed NSPC proliferation. Preliminary data derived from high throughput behavioral screening of 22qDS orthologs indicates that two mitochondrial proteins (mrpl40 and prodha) encoded in the 22qDS deleted region regulate NSPC proliferation, brain size, and behavior in zebrafish. The goal of this proposal is to define the mechanisms through which mrpl40 and prodha govern NSPC proliferation with the hypothesis that NSPC mitochondrial dysfunction in 22qDS may represent a convergent pathologic mechanism. To this end, experiments will employ an innovative combination of in vivo zebrafish studies focused on individual 22qDS genes and cortical organoid studies that model the entire 22q11.2 deletion. Aim 1 will employ cell-type specific transgenic rescue approaches to determine in which cell types mrpl40 and prodha function to regulate brain structure and behavior. Single cell RNA sequencing in zebrafish mutants will be used to define how progenitor/post-mitotic cell populations are altered. Aim 2 will utilize in vivo imaging of transgenic reporters of redox status and cell cycle in zebrafish to define redox dynamics during neurogenesis and to determine how mrpl40 and prodha function to regulate NSPC redox status and proliferation. Aim 3 will use cortical organoid approaches to define NSPC proliferation abnormalities caused by 22q11.2 deletion in a model of human cortical development. The contribution of mrpl40 will then be assessed by analyzing mrpl40 mutant cortical organoids. This proposal fits within NINDS' Strategic Plan, to understand how genes guide healthy brain development and the basic mechanisms underlying NDDs and is expected to generate important insights into mitochondrial regulation of neurogenesis and mechanisms underlying NSPC dysfunction in 22qDS and NDDs. To complement his scientific background, Dr. Campbell will receive training in induced pluripotent stem cell models, next generation sequencing approaches, and in vivo imaging. Dr. Campbell will receive mentorship from Drs. Granato and Anderson who possess complementary expertise and are uniquely suited for this proposal. A thoughtfully selected advisory committee will provide further scientific and career mentorship. Together with the world-class resources and scientific community available at the University of Pennsylvania, the proposed scientific and training objectives will create a strong foundation to establish an independent research program focused on mitochondrial mechanisms governing brain development and underlying NDDs.

NIH Research Projects · FY 2026 · 2024-02

SUMMARY Exercise is an extremely effective lifestyle intervention that dramatically lowers the risk for cardiovascular, metabolic, neoplastic, chronic inflammatory, and neurodegenerative diseases. Despite these beneficial effects, the modern human lifestyle is highly sedentary and new approaches to understanding and improving exercise performance are urgently needed. In this proposal, we will take a new approach and investigate physical activity through the lens of the gastrointestinal tract. In preliminary work using gnotobiotic mice and microbiota depletion approaches, we have recently discovered a critical role for the intestinal microbiome in regulating exercise performance. We found that intestinal microbial colonization contributes to the exercise-induced surge in dopamine in the brain. Dopamine, in turn, is a major element of the reward and reinforcement centers of the brain that drive the engagement in physical activity. Importantly, elevating dopamine levels in the striatum of microbiota-depleted mice restores their exercise performance. This effect of the microbiota on the brain during exercise is dependent on TRPV1+ afferent sensory neurons. These findings provoke the central hypothesis that that the microbiome effect on exercise performance is mediated by neuronal gut-brain signaling which regulates the availability of dopamine in the striatum. We will employ an innovative toolbox at the interface of microbiome science, exercise physiology, and neuroscience to address three central questions: (1) Which microbial genes and which intestinal metabolites influence exercise performance? (2) Which dopamine-sensitive neurons respond to exercise and the microbiome to enhance physical activity? (3) Which sensory neurons transmit the microbiome-derived gastrointestinal signal to the brain to enhance exercise performance? Collectively, these studies will mechanistically define elements of a gut-brain pathway linking the commensal microbiota, gut-innervating sensory neurons, and striatal activity during exercise. Further, the proposed experiments will provide important pre-clinical evidence on the therapeutic potential of gastrointestinal interventions, such as diet and microbial modulation, aimed at enhancing physical activity.

NIH Research Projects · FY 2026 · 2024-02

PROJECT SUMMARY Clear cell ovarian cancer (CCOC) is one of the most challenging subtypes of ovarian cancer (OVCA) to treat, as it is resistant to standard chemotherapy and is associated with poor outcomes. CCOCs likely arise from endometriosis, supporting their unique molecular landscape with ARID1A-inactivating mutations (ARID1Amut) being the most prevalent (50%). Our proposed research capitalizes on the high prevalence of ARID1Amut in CCOC using a novel synthetic lethal approach to provide a new treatment option for patients addressing a clinically unmet need. Our preliminary data shows that ARID1A loss sensitizes CCOC tumors to combination ATR and BET family protein inhibition (ATRi-BETi). ATRi-BETi treatment is synergistic in killing CCOC cells and patient-derived xenograft (PDX) tumors, compared to monotherapies in an ARID1A-dependent manner. These results have led to a Phase IB investigator-initiated clinical trial that will be run through the NRG cooperative clinical trials group evaluating next-generation ATRi (M1774) and BETi (ZEN-3694) in recurrent CCOC. The overarching goals of the research proposed herein are to develop ARID1A as a biomarker for ATRi- BETi treatment and to identify new functionally relevant biomarkers that will facilitate patient selection for this therapy for CCOC in future clinical trials. These goals will be realized through the use of human samples collected from this clinical trial complemented by the use of CCOC PDX models and response data, which will be employed to validate ARID1Amut as a predictive biomarker of response and help identify novel biomarkers that further enhance the effectiveness of this combination. Novel predictive biomarkers will be identified using a multi- dimensional molecular profiling approach that integrates: 1) mutations and gene expression alterations that correlate with responsiveness in humans and animal models as determined by computational modeling; and 2) factors that impact the response to drug treatment at the molecular site of BETi-ATRi action, the genome, and the DNA replication fork. This approach is designed to discover mechanistically relevant biomarkers that predict response to therapy. Our overarching hypothesis is that ARID1Amut and additional molecular alterations will serve as biomarkers of response to ATRi-BETi in CCOC. In summary, these proposed studies will: 1) provide a new treatment option for CCOC addressing a clinically unmet need; 2) determine if ARID1A loss is a biomarker of response to ATRi-BETi; and 3) identify additional mechanistically relevant biomarkers of response to guide future clinical trials.

NIH Research Projects · FY 2026 · 2024-02

PROJECT SUMMARY The Penn-CHOP proposal builds on collaborations and complementary expertise in phenomics and genomics. The 22q11.2 deletion syndrome (22qDS) is associated with high risk for neuropsychiatric disorders across the lifespan. The clinical presentation and course are markedly heterogeneous, with a range of developmental neuropsychiatric disorders, including ADHD, Anxiety, ASD, and Psychosis Spectrum Disorders. Notably, presentation and course resemble idiopathic disorders. Therefore, beyond the specific genetic syndrome investigated, the proposed accelerated longitudinal design will identify convergent risk mechanisms for developmental trajectories of neuropsychiatric disorders in the broader population. We are uniquely positioned to establish developmental trajectories during a critical period, adolescence and emerging adulthood. Lacking in the literature of 22qDS is a systematic examination of environmental exposures, which play an important role in psychopathology and neurocognition in the general population. The nature and degree of medical burden have likewise not been examined in 22qDS. Taking a `genetics first' approach of ascertainment based on a known deletion will allow us to overcome barriers posed by the genetic and phenotypic complexity of idiopathic developmental neuropsychiatric disorders. We postulate that 22q11.2 deletion exerts a large main and multifactorial effects on psychopathology, with contributions from multifaceted environmental exposures and common and rare genetic variants. Dissecting these effects with dimensional measures of psychopathology and neurocognition can elucidate the combined contribution of genetic and environmental mechanisms to psychiatric conditions and build models of risk prediction. Our ability to pursue such a large-scale study capitalizes on our existing successful collaborations, complementary expertise, and institutional commitments to achieve these goals. We propose to parse dimensional measures of psychopathology, neurocognition, and environmental exposures, to elucidate the architecture of risk for neuropsychiatric disorders in 22qDS focusing on the emergence of psychosis. Prospective evaluation with dimensional measures relevant to neuropsychiatric disorders will be applied to a cohort of 300 individuals with 22qDS and their parents, establishing trios. Thus, we will examine family and environmental factors that can contribute to the heterogeneity of presentation and developmental course in 22qDS. Recruitment for longitudinal prospective phenotyping will leverage an existing large cohort with a wealth of clinical data, many of whom have already been ascertained and comprehensively characterized with a range of phenotypic measures. We will also utilize existing genetic data from the largest available case-control samples in the PGC to generate polygenic risk scores for the most common neuropsychiatric disorders evident in 22qDS and examine their relation to outcome. This project will contribute to common phenomic and genomic resources established for data sharing.

NIH Research Projects · FY 2026 · 2024-02

ABSTRACT The microcirculation plays a critical role in organ homeostasis and in disease pathogenesis. Much effort has been dedicated to developing methods to image the microcirculation, however developing quantitative methods to assess organ-specific microcirculation remains an ongoing challenge. Identifying microvascular phenotypes from existing imaging modalities would help overcome these limitations. Most vascular imaging studies focus on larger vessels (> 1mm) due to limited instrument resolution. However, these studies often collect time-course data containing dynamic information that reflects blood flow. Since the microcirculation is primarily responsible for regulating flow, blood flow data reflects microvascular function when there is no proximal stenosis. Thus, we can use time-course dynamic data from imaging studies to identify microvascular phenotypes without directly imaging the micro-vessels. Our central hypothesis is that the time course of contrast material in blood vessels and the dynamics of contrast material in tissue regions contain intravascular and tissue parameters, respectively, which reflect the status of the microcirculation. We propose to develop robust image analysis techniques to discover image-based microvascular phenotypes. We will initially focus on the coronary microcirculation, given the broad public health implications of Ischemic Heart Disease. In Aim 1, we will develop, test, and validate (a) a recently-developed Hybrid Intelligence (HI) approach to segment major vessel segments and myocardial tissue regions in clinical coronary angiograms, and (b) methods to estimate parameters of blood flow in segmented vessels and perfusion in segmented tissue regions. In Aim 2, we will determine the optimal imaging biomarkers for coronary microvascular function using two leading methods currently used to quantify coronary microvasculature. First, we will compare vessel-specific parameters and tissue-based parameters to global and regional myocardial blood flow as measured by Rubidium-82 perfusion cardiac PET. Then, we will compare our parameters against TIMI frame count measurements, an established yet laborious method to quantify coronary flow on coronary angiograms. These studies will develop a novel imaging technology to establish coronary- angiogram based microvascular phenotypes and biomarkers. These methods are also applicable to additional angiography datasets (2D projection x time) including cerebral, renal, pulmonary, and peripheral vascular angiograms, and could be extended to 4D datasets (3D imaging x time) as seen in perfusion computed tomography and magnetic resonance imaging studies. They would therefore allow for assessment of organ- specific microcirculation from existing imaging studies and allow for microvascular phenotyping to greatly improve clinical care and accelerate research in this urgently needed area.

NIH Research Projects · FY 2026 · 2024-02

Summary Reductions in β-cell mass underlie the pathogenesis of all forms of diabetes, raising the relevance of understanding the mechanisms controlling postnatal β-cell growth. Transcriptional networks regulate the development, differentiation, and expansion of β cells, operating through islet enhancers, super-enhancers and promoters forming 3-dimensional hubs. The homeodomain transcription factor and diabetes gene Pdx1 is a critical member of this network, playing roles in β-cell differentiation, proliferation, and function. Despite the importance of Pdx1 for β-cell growth, knowledge of the topological and biophysical properties of Pdx1 that regulate β-cell mass are unclear. Our preliminary data reveal that β cells exhibit altered subnuclear localization and reduced levels of Pdx1 protein as they advance through the cell cycle. Further, ectopically elevated levels of Pdx1 prevent cell cycle progression and increase β-cell death, suggesting that dynamic regulation of expression is required for effective β-cell expansion and maintenance of functional β-cell mass. We identify an intrinsically disordered protein region (IDPR) of unknown function in the Pdx1 C-terminus (aa 207-223). IDPRs, commonly found within transcription factors, lack fixed secondary structure and are amenable to flexible conformations and phase separation. IDPRs promote protein-protein interactions and transcriptional hub formation at super enhancers necessary for coordinated gene regulation. We previously identified the E3 ubiquitin ligase substrate adaptor protein SPOP as a PDX1 C-terminus protein partner (via aa224-238) that mediates ubiquitination and proteasomal degradation of PDX1. SPOP binds other IDPR partners in phase separated nuclear compartments critical for their function. Thus, we hypothesize that the IDPR and SPOP interaction domains within the Pdx1 C-terminus play critical, possibly interdependent, roles in Pdx1 protein localization, expression, and function crucial for β-cell gene expression and proliferation. This hypothesis will be tested in three Aims: (1) To investigate the dynamic subnuclear localization of PDX1 during the cell cycle and in response to glucose. (2) To determine the role of IDPR-regulated phase separation in the function and localization of PDX1. (3) To determine the role of SPOP in fine tuning PDX1 protein level and function. Our studies will determine a novel and cohesive role for unstudied structural features of the Pdx1 C-terminus and how they influence β-cell growth and glycemic control. Results of our proposed studies will inform therapeutic efforts to optimize β-cell mass expansion ex vivo for cell based therapies and in vivo to treat patients with diabetes.

NIH Research Projects · FY 2025 · 2024-02

The proposed NIDCR Dentist Scientist Pathway to Independence Award (K99/R00) will provide me advanced research and academic training to become an independent dentist-scientist at the interface of oral microbiology, biophysics, and spatial omics with impact in Early Childhood Caries (ECC). ECC is a major public health problem characterized by high microbial carriage forming intractable plaque-biofilms on teeth exposed to sugar-laden dietary habits. The disease leads to rampant tooth-decay, is costly to treat and can cause systemic complications in children. Previous studies and a recent multi-omics analysis of dental plaque from two large community-based cohorts of pre-school children (>400) have identified Selenomonas sputigena (Ss), a motile bacterium, to be strongly associated with ECC. This finding was further validated in an in vivo caries model whereby Ss exacerbated the severity of carious lesions when co-infected with Streptococcus mutans (Sm). However, the role of Ss and its motility on biofilm formation, interspecies interaction with Sm, and cariogenic functions are unknown. To address this, I will focus on the overall hypothesis that the motile Ss colonizes tooth surface and interacts with Sm to mediate biofilm spatial structuring and community functions that promote emergent caries- causing properties of supragingival biofilms through three Aims: (1) Characterize Ss motility on surface colonization and biofilm initiation; (2) Determine the dynamics of biofilm assembly, spatial transcriptomics and disease-associated functions; (3) Investigate Ss-mediated colonization, interspecies spatial structuring/omics and biofilm virulence in vivo. The outcome will elucidate the role of Ss and its motility in supragingival biofilm formation and Ss contributions to the etiopathogenesis of ECC. During the K99 mentored phase, I will conduct research in the lab of my primary mentor/co-mentor, while developing key expertise in 3 areas: 1) advanced skills in biophysical methods to study motile bacteria; 2) acquiring knowledge and technical skills in single-cell RNA sequencing and spatial transcriptomics; 3) improving scientific communication and grant writing skills. In addition, I will incorporate mentoring and laboratory managing skills as well as networking. In transition to independent R00 phase, I will implement and complete the spatial transcriptomics studies to understand how Ss influence biofilm interspecies interactions and determine its pathogenic role in dental caries in vivo. The data will provide ample opportunities for further mechanistic studies and targeted strategies for ECC. In addition, it will provide a platform to study other motile oral bacteria in health and disease, which remain understudied. Collectively, the proposal will broaden my vision and skills by capitalizing on highly experienced mentor and co- mentor with an interdisciplinary advisory committee with complementary expertise in biophysics, spatial multi- omics, oral microbiome, and clinical research. I will gain essential knowledge, skills, and experience to build my own research program with the goal of receiving an R01 prior to the end of award, and successfully guide me to pursue independent dentist-scientist career in oral microbiology and cariology with multidisciplinary vision.

NIH Research Projects · FY 2025 · 2024-02

ABSTRACT Obstructive sleep apnea is an extremely common condition that contributes independently to multiple adverse outcomes, including cardiovascular disease, diabetes, and neurodegeneration. It is a major public health problem given its high prevalence. OSA is heterogeneous, with established differences in symptoms and outcomes in individuals with comparable disease severity based on traditional metrics. We were the first to show that there are different symptom subtypes of OSA using unsupervised clustering analysis, including groups defined by disturbed sleep with symptoms of insomnia, minimal symptoms, and marked excessive sleepiness. The existence of these subtypes has since been replicated around the world in multiple population-based and clinical cohorts. We have also shown that there are differences in outcomes between the these subtypes, particularly for cardiovascular events and mortality, with increased risk in the excessively sleepy subtype. However, there are still many unanswered questions. To begin to address these unanswered questions, we propose to use data from the STAGES study, which recruited patients undergoing sleep studies across six sleep centers in the United States and Canada. Participants had in-depth phenotyping that included: detailed questionnaires about their sleep and health, depression and anxiety scores, actigraphy to assess sleep duration over a 2-week period, psychomotor vigilance test to assess sleepiness objectively, and a comprehensive battery of tests to assess cognition. In this project, we will first examine if there are differences in objectively-measured sleepiness among the symptom subtypes, i.e., are individuals who complain of sleepiness also objectively sleepy? We will also assess what role chronic insufficient sleep (based on actigraphy) and poor sleep quality (based on a new measure of sleep depth—the Odds Ratio Product) play in determination of symptom subtypes. Next, we will expand knowledge regarding the clinical relevance of these subtypes by evaluating differences in neurocognitive outcomes. We hypothesize that the excessively sleepy group will have worse cognition than the other groups, after controlling for key covariates. Finally, we hypothesize that objective data on sleepiness, sleep duration, and sleep quality can be used to better understand additional heterogeneity among subjects with OSA. Specifically, we will perform unsupervised clustering based on both subjective symptoms and objective measures of sleepiness and sleep behavior (e.g., sleep duration and quality). We predict that we will identify new subgroups of subjects and will assess whether there are larger differences in neurocognitive outcomes among these subtypes when compared to subtypes defined only by subjective symptoms. The rich phenotype data in STAGES allow us to address all of these important questions. Overall, this project is aimed at further defining the heterogeneity of obstructive sleep apnea, an essential step in development of more personalized approaches to diagnosis and management. Future directions for this study will include follow-up analyses in other cohorts to validate the results and assess impact on other health outcomes.

NIH Research Projects · FY 2026 · 2024-02

The objective of this project is to develop methodology for energy-based background estimation that can be applied to clinical data and produce accurate quantitative PET images over challenging imaging situations such as low collected counts, high multiple scatter, and prompt gamma contamination when imaging non-standard PET isotopes. The goal is to enhance the accuracy of PET imaging in situations where current state-of-art scatter estimation techniques are limited in accuracy or perform poorly. In this proposal, we develop a data driven scatter estimation methodology that makes full use of the annihilation photon energy information present to estimate scatter. This method is also extended to provide correction for bias arising from prompt gammas present in data collected form some non-standard PET isotopes. We implement, optimize, and evaluate this algorithm on measured data from a clinical PET scanner for standard and non-standard isotopes, and subsequently apply the methodology to organ- specific scanners (brain and breast). The proposed work will be accomplished through the following specific aims: (i) optimization and evaluation of the EB method for scatter estimation, (ii) application of the EB methodology to dedicated brain and breast PET scanner geometries, and (iii) extension of the EB methodology to correct for prompt gamma contamination present in data acquired from non-standard PET isotopes. In addition to its advantages over existing scatter estimation methodology in situations with low collected counts and/or data with higher level of multiple scatter, the proposed technique is expected to be faster, does not require knowledge of activity distribution outside the imaging field-of-view, and does not require a transmission or CT image. Successful demonstration of this technique will significantly impact routine oncologic imaging where heavy patients with increased scatter, reduced counts and limited imaging field-of-view will be susceptible to reduced quantitative accuracy. In addition, this technique can also expand the application of quantitative PET/CT in new oncology imaging areas such as treatment monitoring with low-dose repeat PET scans, imaging with new biomarkers that use low positron yield radionuclides (e.g. 124I, 86Y, etc.), or acquiring data at high count-rates (as in cardiac imaging or imaging with 124I or 86Y). Beyond oncology, it will also provide improved quantitation in cardiac studies (82Rb, 13NH3, or 11C-actetate). Since, the proposed scatter estimation method does not require a CT image it may have an application in PET/MR imaging as well as clinical studies with some patient motion – both situations where the CT image is either not available or is compromised leading to errors in the traditional way of estimating scatter.

NIH Research Projects · FY 2025 · 2024-02

Candidate. Christopher Chesley, MD, MSCE, is a pulmonary and critical care physician and health disparities researcher who is committed to eliminating disparities related to acute respiratory failure (ARF) and sepsis. To promote development toward research independence, he seeks guided training to facilitate mastery of advanced causal effect methodology, qualitative study design, and system-wide intervention development. Research context. Disparate clinical outcomes are well documented for minoritized patients with ARF and sepsis, but interventions mitigating them are lacking. In part, this is due to a limited understanding of the salient mechanisms most influential to disparities. Among them, harms mediated through socioeconomic disadvantage (defined as access to poor quality socioeconomic resources) and care characteristics related to hospitals that serve the largest proportions of minority patients (often described as minority serving hospitals, or MSHs) are particularly likely to play causal roles. Thus, this proposal aims to clarify the mechanistic roles of these underexplored determinants of health disparities in the context of ARF and sepsis. Specific aims. 1) Characterize the mechanistic role of neighborhood socioeconomic disadvantage on clinical outcome disparities among critically ill patients; 2) Describe clinical processes that influence length of stay disparities for critically ill patients hospitalized at MSHs; and 3) Identify perceived barriers and facilitators of implementing an intervention to reduce hospital readmissions among critically ill patients at MSHs. Research plan. Dr. Chesley will examine relationships between minority identity and clinical outcomes of 200,000 patients across 16 U.S. hospitals. After geocoding four clinically impactful forms of neighborhood-level socioeconomic disadvantage, he will use mediation analyses to elucidate causal roles for these measures. Then, adapting research frameworks from human factors and ergonomics, he will describe the clinical processes that most strongly influence hospital length of stay disparities using semi-structured interviews and direct observation of clinical activities. Lastly, he will conduct semi-structured interviews to identify barriers and facilitators to implementing a systems intervention recommended by the Centers of Medicare and Medicaid Services. Career development plan. Working closely with his mentors and advisors, Dr. Chesley will 1) enhance his mastery of quantitative methods, with emphasis on causal effect methodology and geospatial mapping; 2) develop comprehensive skills in qualitative and mixed-methods study design; and 3) become proficient in developing and testing health systems-level, disparities-mitigating interventions. Environment: The University of Pennsylvania offers an ideal environment to pursue this training, with well-established mentors, a home department heavily dedicated to Dr. Chesley’s success, and several multi-disciplinary research centers with long track records of producing successful, independent investigators.

NIH Research Projects · FY 2026 · 2024-02

ABSTRACT The clinical benefits of cancer immunotherapies, including adoptive cell transfer (ACT) of chimeric antigen receptor (CAR) T cells, are limited when used against solid tumors. The immuno-suppressive tumor microenvironment (TME) is enriched in cellular components (regulatory T cells, myeloid derived suppressor cells, tumor-associated macrophages, etc) and acellular factors (hypoxia, deficit of nutrients, acidosis, adenosine, etc) that decrease viability and tumoricidal activities of anti-tumor native CD8+ cytotoxic T lymphocytes (CTL) and of therapeutic CAR T cells. Therapeutic neutralization of these factors and components is challenging because of their diversity and redundancy. Instead, we aim to identify and thwart the key mechanisms by which the TME-derived factors and conditions undermine viability and the anti-tumor activities of CAR T cells. We will focus on TME-triggered downregulation of type I interferon (IFN1) receptor IFNAR1, which normally supports viability and activity of native CTLs and CAR T cells. Our preliminary data show that MAPK Activated Protein Kinase 2 (MK2) and mono-ADP-ribosyl transferase PARP11 cooperate to downregulate IFNAR1 on intratumoral CAR-bearing T cells, leading to their inactivation and rapid cell death. IFNAR1 loss leads to downregulation of IFN1-inducible cholesterol 25-hydroxylase (CH25H). CH25H acts to limit the effector trogocytosis between malignant cells and specific CAR T cells. In the absence of sufficient levels of CH25H and its product 25-hydroxycholesterol (25HC), this trogocytosis undermines the activities of CAR T cells and exposes them to fratricidal killing. These and other exciting preliminary results suggest an overarching hypothesis that targeting TME-driven PARP11/MK2/CH25H-dependent mechanisms that regulate the viability and activity of CAR T cells should enhance their anti-tumor activities and increase the efficacy of CAR T ACT. To test this hypothesis, we will determine (i) the roles of TME-induced PARP11 in inactivation of CAR T cells, (ii) the importance of MK2 activity in suppression of intratumoral CAR T cells and (iii) the contribution of downregulation of CH25H in CAR T cells inactivation and decreased efficacy of CAR T ACT. Completion of these studies should gain insight into immunosuppression of intratumoral CAR T cells and help to develop novel CAR constructs and CAR T pre-treatments as well as combinatorial approaches to increase the efficacy of CAR T ACT.

NIH Research Projects · FY 2026 · 2024-02

PROJECT SUMMARY Cellular response and adaptation to external stress is a fundamental aspect of normal tissue homeostasis. In cancer, these mechanisms of cellular adaptation become detrimental as they allow cancer cells to survive through therapy, thus contributing to therapy resistance. The emergence of therapy resistance in cancer is an example of a phenomenon shaped by both selection and adaptation: While most cells die upon treatment (selection), a rare subpopulation survives, and a smaller subset even adapts to proliferate in drug medium. Thus, when studying resistance in cancer, a sample of cells at any time during treatment contains a mixture of cells with different fates: growth, senescence and death. Our preliminary data has revealed a complex and fascinating interplay between cellular mechanisms for survival and those for adaptation, with some mechanisms shared between stressors and others unique to specific stressors. Due to the pervasive role of cellular stress-response mechanisms in tissue homeostasis and disease, an understanding of how (and which) cells survive and adapt under different stresses is fundamental both for the understanding of basic tissue biology as well as the development of disease treatments. Currently, there is a lack of methods for single-cell fate tracking in these complex systems where cell fate is driven by survival and adaptation. One promising experimental approach is lineage barcoding, which uses unique DNA sequences to label and track individual cells over time. However, there is yet a lack of both experimental and computational frameworks that integrate lineage tracing with the latest multimodal single cell and spatial sequencing technologies. Further, the pervasive adoption of single-cell sequencing methods in the study of human disease underscores the importance of computational approaches for trajectory reconstruction and cell fate prediction that is applicable to a clinical setting, without the need for genetic barcoding. In this project, we develop new genetic experimental systems for integration of lineage barcoding with spatial barcoding and with single-cell multiome RNA and ATAC sequencing; in parallel, we develop a general computational framework for the prediction of cell fates in the absence of genetic lineage tracing. The new experimental systems serve as new protocols for the scientific community as well as a powerful validation framework for the computational methods development. The new computational methods for lineage reconstruction and fate prediction using multiomic RNA and ATAC sequencing data will be released as open-source software, addressing critical limitations in current methods. These methods will be applied to the study of cancer cells treated with a panel of different drugs and stresses. This human biological system is rich with lineage dynamics and multiple cellular fates as each cancer cell has the possibility of growth, senescence, or cell death. Thus, our work will reveal fundamental biology about the complex relationship between cancer cell heterogeneity and the cellular fate of drug resistance.

NIH Research Projects · FY 2026 · 2024-02

PROJECT SUMMARY / ABSTRACT Loss of eyesight is regarded among the worst possible diseases by most Americans. Despite being rare causes of vision loss, inherited retinal diseases (IRDs) are molecularly simple single-gene defects, and advances in genetics and genomics have raised hopes for the development of gene-based treatments. For most IRDs, the natural history of disease involves progressive vision loss and successful interventions must demonstrate a clinically meaningful slowing of the natural progression. Many good metrics have been developed to measure vision loss but the continued lack of approved IRD treatments strongly suggest better outcome measures are still needed. The long-term objective of the research is to predict retinal locations maximally vulnerable to progression over the next 2 years individualized for each patient across distinct IRDs. Successful predictions of loci of vulnerability will drive reliable measurements of functional and/or anatomical changes over the duration of typical clinical trials. The focus of the current project is on two human IRDs that remain without approved treatments – autosomal dominant retinitis pigmentosa caused by RHO mutations (RHO-ADRP) and Stargardt disease caused by ABCA4 mutations (ABCA4-STGD). Average tendencies for spatio-temporal progression in both diseases are well investigated. However, the reliable and individualized prediction of the photoreceptor locations maximally vulnerable to fast progression remains a major challenge. Current literature and preliminary studies support the hypothesis that retinal cross-sectional structure at each location when considered together with the structure of its immediate neighborhood retains enough information to predict vulnerability to disease progression. The current project will involve a combination of retrospective longitudinal and prospective longitudinal studies that operate on different spatial scales and structure/function dimensions, to test the hypothesis and provide a more complete understanding of the range of photoreceptor vulnerability to disease progression in IRDs. Aim 1 will first use a unique existing data set obtained serially in RHO-ADRP patients and train an artificial intelligence (AI) model to learn input OCT features that correspond to disease progression. Trained AI will be applied to another unique existing data set obtained in ABCA4- STGD patients. With special attention to heterogenous transition zones, retinal locations maximally vulnerable will be mapped and validated against serial data. Aims 2 and 3 will use prospective serial studies in ABCA4- STGD to test predictions directly with en face imaging, ultra-wide angle OCT recordings, microperimetric evaluation of rod- and cone-specific light sensitivities, and novel adaptive-optics OCT imaging of the outer retina. The project should provide novel insight into the interaction of human photoreceptors with their diseased neighbors and allow optimum localization of visual function measurements to provide individualized and sensitive outcome measures for ABCA4-STGD clinical trials.

NIH Research Projects · FY 2026 · 2024-01

Abstract/Project Summary Candidate: Austin Kilaru, MD MSHP, is an emergency physician and health services researcher who is committed to improving outcomes for patients with acute cardiovascular illness through the implementation of evidence-based care. To achieve his career goal of becoming an independent investigator, he seeks mentored research training to strengthen skills in qualitative methods, causal inference, and implementation science. Research Context: Each year, there are 1.4 million visits to US emergency departments (EDs) for acute heart failure (AHF). Stable patients can be discharged from the ED after initial evaluation and treatment, but nearly 90% of patients are hospitalized. Evidence-based risk scores have been developed to inform hospitalization decisions for AHF, given that patients may prefer to recover at home and the costs and outcomes associated with hospitalization. However, AHF risk scores are not widely used. Moreover, they do not consider additional factors that may be important to hospitalization decisions, including access to care, capacity for self-care, and health-related social needs. Scalable approaches, like clinical decision support tools, are needed to promote AHF risk stratification and supplement risk scores with additional factors that matter to patients and clinicians. Specific Aims: 1) Identify factors that influence AHF hospitalization decisions for ED patients and physicians; 2) Compare outcomes for low-risk AHF patients who are hospitalized to those discharged from the ED; 3) Pilot and evaluate implementation of a clinical decision support tool for AHF hospitalization. Research Plan: To accomplish these aims, Dr. Kilaru will first conduct qualitative interviews with low-risk AHF patients who were either hospitalized or discharged from Penn Medicine EDs. He will also conduct interviews with ED physicians to examine decision-making factors. Then, he will use electronic health record data for a retrospective cohort study to determine the association of hospitalization with outcomes among low-risk AHF patients, seeking to further test AHF risk score effectiveness and inform implementation strategies. Finally, he will design and implement a clinical decision support tool, based in the electronic health record (EHR), among ED physicians at Penn Medicine, evaluating outcomes including acceptability, adoption, and feasibility. Career Development Plan: Working closely with his mentorship team, Dr. Kilaru will pursue didactics, seminars, and individualized instruction to complete his training goals, which are to 1) expand skills in qualitative methods to focus on patients and clinicians 2) apply causal inference techniques to analyze EHR data, and 3) gain implementation science expertise to design and test innovations in care delivery. Environment: The University of Pennsylvania and Penn Medicine offer an ideal environment to pursue this training and research. Dr. Kilaru will succeed because of the support of an experienced and dedicated mentorship team, outstanding research infrastructure, and extensive resources for professional development.

NIH Research Projects · FY 2026 · 2024-01

Summary Epilepsy affects over 70 million people globally and more than 3.6 million Americans, 1/3 of who are not controlled by medication. While new technologies like laser ablation, implantable devices, high bandwidth intracranial EEG (iEEG), and multimodal imaging have improved therapies, meaningful data sharing is lacking which limits clinical progress and translational research. There is a tremendous need to aggregate, share and collaborate on increasingly large and complex multimodal data from patients with medication-resistant epilepsy to advance these efforts, but a lack of novel mechanisms to share and explore these data in a meaningful way. Our central hypothesis is that a scalable, self-sustaining Epilepsy Data Ecosystem (EDE), which integrates multi-modal datasets and is responsive to the changing needs of the epilepsy community, will dramatically accelerate translational research and impact clinical care. The EDE will also accelerate the fields of Machine Learning, Neuroscience and Computer Science, communities that depend upon large, multimodal data from these patients for their research. Over the past 14 years our group has focused on building tools and community to accelerate translational epilepsy and neuroscience research through two major efforts: (1) through our open- source platform iEEG.org, for sharing data and computational tools in epilepsy. Seeded by a grant from NINDS from 2009-2013 and supported by a group of over 200 scientists focused on Engineering and Epilepsy Research (the ICTALS group), the platform now has almost 6,000 users and is self-sustaining, paid for by groups who use it for their research needs. It shares over 1,000 published data sets free of charge, and it has generated scores of publications. Advances to care, data standards and platform technologies as well as the significant increase in data volume require that iEEG.org be updated. (2) Led by Dr. Joost Wagenaar, The Pennsieve Platform was independently developed over the last 7 years as an open-source, cloud-based data management and sharing platform for large volume, multi-modal data in the Neurosciences, with a focus on standards, scalability, and sustainability. It is currently used as the data core for several NIH programs (e.g. SPARC and REJOIN) and is ideally suited to replace iEEG.org as the next generation repository for multimodal data in Epilepsy research. Our goals for this proposal are: (1) To migrate all data, tools, and users from iEEG.org to Pennsieve, to ensure continued impact of iEEG.org public datasets by updating the data to adhere to current standards, (2) To tailor Pennsieve to the Epilepsy Community's needs, to develop seamless mechanisms for submitting, finding, sharing, accessing, and publishing high impact Epilepsy Datasets in line with all FAIR requirements and standards in the age of big integrated data, (3) To build community and prospective data contributions to the EDE through educational workshops, seminars and web content, and (4) to leverage Pennsieve's workflows and existing model to ensure sustainability. This project leverages an established collaboration between investigators across Medicine and Engineering at Penn, and the International Epilepsy Community.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY The average smoker will attempt to quit smoking at least 30 times before abstaining for 12 months or longer. These attempts typically occur over decades of smoking, carcinogen and toxicant exposure, resulting in 480,000 deaths annually. As highlighted in the Surgeon General’s Report, helping smokers who cannot quit smoking switch to less harmful non-combustible nicotine-containing products, such as e-cigarettes, has the potential to reduce this health burden dramatically. Substituting e-cigarettes for combustible cigarettes might only be possible for persistent smokers if e-cigarettes are accessible and appealing. Harm reduction proponents have advocated for the continued availability of e-cigarette flavors to appeal to and aid cigarette smokers unable to quit with traditional methods. Yet, there are no prospective studies of the effect of flavor on initial and sustained switching from combustible to electronic cigarettes. Converging laboratory, epidemiological, and clinical research suggests that fruit-flavored e-cigarettes with nicotine may be a viable substitute for combustible cigarettes among persistent smokers. The proposed study seeks to answer two novel questions relevant to public health and the regulation of e-cigarette flavoring. First, do persistent smokers substitute fruit-flavored e- cigarettes more readily than traditional-flavored e-cigarettes (tobacco or menthol) for combustible cigarettes? Second, are fruit-flavored e-cigarettes more rewarding and reinforcing than traditional-flavored e-cigarettes, and do these effects facilitate switching? The proposed research will fill these gaps in the evidence base by randomizing 210 persistent cigarette smokers to a six-week regimen of fruit-flavored (FF: watermelon and blueberry, n=70), tobacco-flavored (TF n=70) or menthol-flavored (MF n=70) e-cigarettes in a between-subjects design. Baseline smoking rate will be established during days 1-5. After biochemically verified overnight cigarette smoking abstinence, laboratory visits on days 6 and 7 will assess flavor-associated subjective reward and the reinforcing value of flavored e-cigarettes relative to combustible cigarettes. Participants will then switch from cigarette smoking to e-cigarette use for six weeks. Participants will collect spent cigarette filters daily to assess cigarettes smoked per day (cpd) if they smoke. The primary outcome measure is the longitudinal daily count of cigarettes from baseline to the end of the six-week switch period, with cigarettes per day at a 6-month follow-up as a secondary endpoint. This study aligns with NCI priorities outlined in the Notice of Special Interest (NOSI NOT-OD-22-023) for research on “how ENDS use influences smoking (e.g., quit attempts, sustained abstinence, relapse).”

NIH Research Projects · FY 2026 · 2024-01

Project Summary Airway tuft cells (also known as “brush” or “solitary chemosensory” cells) are rare cells found in the nose and trachea. Tuft cells are also found in the lung after injury and/or inflammation or in genetic diseases like primary ciliary dyskinesia. Tuft cells regulate local antimicrobial peptide, acetylcholine (ACh), and IL-25 secretion. IL-25 is an important driver of Th2 inflammatory responses observed in airway disease like chronic rhinosinusitis (CRS) with nasal polyps and asthma. Tuft cell ACh may activate sensory neurons and/or local mucociliary or inflammatory responses. We know little about how to target tuft cells in airway diseases, because we know little about human tuft cell function. Their rarity (≤1 in 100 cells in the nose) makes them difficult to study, though their frequency increases significantly (up to 30% of the epithelium) in nasal polyps. We know that tuft cells express a range of chemosensory G protein-coupled receptors (GPCRs) but know little about how they signal and regulate cell responses. We previously showed that activation of T2R bitter taste GPCRs stimulates Ca2+ -driven secretion of antimicrobial peptides from surrounding epithelial cells. This response is inhibited by activation of cAMP downstream of T1R sweet taste GPCRs, which sense airway surface liquid glucose or sweet bacterial D-amino acids. Other GPCRs (e.g., succinate, cholinergic receptors, adenosine receptors, etc.) exist in tuft cells, but their functions are less clear. Many studies have been done in mice, but our research revealed differences in how mouse vs human tuft cells signal. We need better methods for studying human tuft cell function to complement and extend prior mouse studies. The goal here is to identify the signaling and downstream consequences of nasal tuft cell GPCRs, which may be important therapeutic targets for respiratory infections, either to enhance antibacterial immunity or reduce nasal/lung inflammation. We will utilize a novel genetic labeling strategy to express fluorescent protein biosensors specifically in human tuft cells cultured from residual nasal surgical material. This allows optical imaging of tuft cell function within an intact epithelial monolayer differentiated at air-liquid interface. In Aim 1, we will use these methods to study how tuft cell GPCRs regulate Ca2+, cAMP, and other pathways to fine tune antimicrobial peptide secretion. In Aim 2, we will study how these pathways regulate electrical excitability of airway tuft cells and if/how membrane voltage changes contribute to tuft cell responses. In Aim 3, we will elucidate mechanisms of how tuft cells secrete/release ACh and IL-25 in the context of Th2 iairway disease. These data will clarify the regulation of intracellular signaling of human tuft cells to better understand how to target them in airway diseases like CRS and possibly asthma.

NIH Research Projects · FY 2026 · 2024-01

Project Summary Posttraumatic stress disorder (PTSD) is associated with alterations in arousal that do not respond well to evidence-based practices. Patients with PTSD tend to fall into one of two groups in extinction training (and in exposure therapy): 1) over-engagers, where arousal is too high; and 2) under-engagers, where patients are so worried about becoming upset that they distract themselves from the task (which prevents learning). In this mechanistic clinical trial, we will evaluate a strategy to augment extinction training with neuromodulation to reduce arousal and improve extinction retention. Augmenting extinction training with continuous theta burst stimulation (cTBS, a type of transcranial magnetic stimulation) delivered to the intraparietal sulcus (IPS) may lead to targeted reductions in arousal. Our team has shown that the IPS is a “connectivity hub” for arousal and that stimulating this region with TMS can reduce excessive arousal in healthy people. The goal for this R01 is to evaluate the main effects of IPS cTBS (versus sham cTBS, a between-subject comparison) and its interaction with extinction training (vs. neutral training) on arousal among patients with PTSD. We hypothesize that reducing parietal hyperexcitability will help patients with PTSD to modulate arousal during extinction training—enough arousal to ensure that they can benefit, but not too much arousal which prevents learning. These results could translate into future opportunities for novel therapeutic targets among patients with PTSD. The specific aims are as follows: Aim 1: To evaluate the optimal dose of IPS cTBS. H1: Attenuation of startle for IPS cTBS vs. sham cTBS will plateau at 1200 pulses, the anticipated optimal cumulative dose. Aim 2: To compare the main effect of IPS cTBS and its interaction with extinction training on arousal (measured by startle response). Using a two (between group: sham vs. IPS cTBS) x 2 (within group: extinction training vs. control/neutral training) randomized controlled design among patients with PTSD (N = 120), we will examine the potential benefit of cTBS and extinction training on reduction in arousal. H2a: IPS cTBS will result in greater reduction in arousal compared to sham cTBS. H2b: Participants who receive IPS cTBS will have a significantly lower difference in retention of extinction learning vs. control/neutral training (indicative of greater retention of extinction learning) compared to participants who receive sham cTBS. Secondary: We will test analogous hypotheses on subjective outcomes and on cognitive outcomes and we hypothesize similar directions of effects. Aim 3: To evaluate neural mechanisms of action of IPS cTBS + extinction training. Participants will complete a resting state fMRI scan on tests of retention of learning experimental visits to evaluate neural changes from training. H3: We will observe attenuated activation (relative to pre-test) of the IPS for participants who received IPS cTBS compared to those who received sham cTBS.