University Of Pennsylvania

NIH Research Projects · FY 2025 · 2025-08

Summary The goal of this proposal is to test the hypothesis that the transcriptional program mediating the process by which endothelial cells in the embryo differentiate into hematopoietic stem and progenitor cells (HSPCs) is coordinately regulated by cis-acting regulatory sequences derived from retroelements. Retroelements are repetitive DNA sequences comprising approximately 50% of mouse and human genomes. The long terminal repeats (LTRs) of the retrotransposon family of retroelements function as enhancers, promoters, and polyadenylation signals for endogenous gene expression and are highly enriched for transcription factor binding sites (TFBS). We have identified families and specific subfamilies of retrotransposons that are more highly expressed in the small population of endothelial cells in the embryo that differentiate into HSPCs (hemogenic endothelial cells) than in non-hemogenic endothelial cells, and in particular give rise to progenitors with potent lymphoid potential. Some of these subfamilies of retrotransposons contain RUNX1 motifs in their LTRs, and we hypothesize that RUNX1 regulates HSPC differentiation from hemogenic endothelial cells through these LTRs. We will use long read sequencing technology and bioinformatics approaches adapted for the purpose of analyzing retroelements to map transcriptionally active retroelements based on their expression and chromatin configuration to specific genomic coordinates and determine if they activate genes involved in HSPC formation from hemogenic endothelial cells. We will also determine if the RUNX1 transcription factor regulates the activity of a subset of these LTRs.

NIH Research Projects · FY 2025 · 2025-08

ABSTRACT: As specialized cells with limited cytoplasm, sperm have long been viewed as contributing only the paternal genome to progeny. However, in the past decade experiments in mice have revealed that sperm microRNAs (miRNAs) influence early embryogenesis and are sufficient for the paternal transmission of non-genetically inherited phenotypes. miRNAs, along with their protein cofactor Argonaute (Ago), bind messenger RNA (mRNA) targets and subsequently downregulate their expression through mRNA decay and translational repression. However, the mechanism underlying how sperm miRNAs act as inherited information remains undetermined. We hypothesize that sperm miRNAs modulate early development through the direct regulation of key mRNA targets like chromatin modifiers or signal transduction pathways, thus altering downstream gene expression programs in a manner that persists through development. Due to the difficulty of identifying targets of miRNAs in the embryo, the targets of sperm miRNAs are still currently unknown. The Fx-miRs are a cluster of miRNAs that are highly expressed in the sperm of every mammalian species analyzed. When wildtype eggs are fertilized with Fx-miR deficient sperm, resulting embryos have decreased developmental potential and demonstrate significantly altered gene expression in preimplantation embryogenesis. Despite these phenotypes, the mechanistic functions of the Fx-miRs in the early embryo, especially what mRNAs these miRNAs target and how these binding events impact overall gene expression during early development, remain unknown. I hypothesize that sperm miRNAs directly target and regulate a subset of maternally provided or zygotically expressed mRNAs postfertilization which then leads to downstream gene expression effects that modulate embryonic development. In Aim 1a, I will use ribosome profiling to evaluate the impact of individual Fx-miRs and the whole cluster on global translation to discover additional regulation by these miRNAs not captured by mRNA-seq. Here, I will use mESCs as a model for early embryonic development, as my preliminary data has demonstrated that mESCs are an efficient model to profile the molecular functions of sperm miRNAs. In Aim 1b, I will use Ago enhanced Crosslinking and Immunoprecipitation (Ago eCLIP) to identify the direct targets of the Fx-miRs on the level of individual miRNAs, also using mESCs as a model. Finally, in Aim 2, to dissect direct versus secondary effects, I will knockdown specific Fx-miR targets identified in my preliminary data via siRNA in embryos fertilized with Fx-miR deficient sperm and perform single embryo RNA-seq. Downregulation of specific Fx-miR targets and the analysis of the resulting differential gene expression will allow for the separation of direct versus downstream effects of the Fx-miRs in vivo. This project will be the first to identify the postfertilization gene regulatory functions and direct targets of an important cluster of sperm miRNAs and thus reveal mechanistically how sperm miRNAs can program embryonic development to initiate the non-genetic inheritance of offspring phenotypes.

NIH Research Projects · FY 2025 · 2025-08

Project Summary The goal of this proposal is to develop a modular chemical biology tool to probe the biology of alpha-synuclein post-translational modifications. Parkinson’s disease is a devastating neurodegenerative disease characterized by pathological aggregations that include alpha-synuclein (a-syn). a-syn is a small (15 kDa) intrinsically disordered protein with the propensity to aggregate into cytotoxic oligomers or fibrils. It is suspected to play a substantial role in the pathophysiology of Parkinson’s disease. a-syn has dozens of recognized post translational modifications (PTMs), including phosphorylation, acetylation, or arginylation. Prior work has demonstrated that some PTMs, such as arginylation or acetylation, are associated with reduced pathogenicity of a-syn fibrils. Phosphorylation, by contrast, is associated with a-syn pathology and its presence is treated as a proxy for a-syn-fibril-induced cytotoxicity in cell and animal models. However, prior research into a-syn PTMs has been hindered by the lack of tools to directly induce PTMs of fibrils. Without methods to selectively modulate the PTMs of a-syn fibrils it is difficult to make causal inferences on the native function of each PTM. Bifunctional molecules are one method of introducing such PTMs, as by binding to PTM enzymes and the protein of interest they can cause proximity-induced labeling of the protein of interest. Prior work from our laboratory has demonstrated the use of bifunctional molecules to induce ubiquitinylation and degradation of target proteins labeled with e. Coli Dihydrofolate Reductase (eDHFR), a small (18 kDa) protein tag targetable with trimethoprim (TMP) based bifunctional molecules. In this proposal, I will adapt the eDHFR protein tag, alongside TMP-based bifunctional molecules targeting a-syn fibrils, to create a modular system for the small- molecule-induced post translational modification of a-syn fibrils. By this, we will gain a modular, selective, and inducible way to induce PTMs onto alpha synuclein fibrils, empowering research into the role of a-syn PTMs. In aim 1 I will synthesize TMP-based bifunctional molecules targeting a-syn fibrils and eDHFR and I will use them to induce the arginylation or acetylation of a-syn fibrils in cells transduced with eDHFR-enzyme fusion proteins. I will then use this system to study the impact of arginylation or acetylation on the cytotoxicity of a-syn fibrils in a Parkinson’s disease model system. In aim 2 I will adapt the system to study the effects of the phosphorylation of a-syn fibrils in Parkinson’s models. I hypothesize that the direct arginylation and acetylation of a-syn fibrils, as shown in aim 1, will reduce the cytotoxicity of a-syn fibrils in Parkinson’s models. I additionally hypothesize that direct phosphorylation of a-syn fibrils, as shown in aim 2, will exacerbate the toxicity of a-syn fibrils in Parkinson’s models. The demonstration of cytoprotective PTMs of a-syn would point to future directions for Parkinson’s therapy; the demonstration of the cytotoxicity of a-syn phosphorylation would clarify the chain of causality between a-syn phosphorylation and cytotoxicity, providing valuable insight into the role of a-syn phosphorylation in Parkinson’s pathogenesis.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY The tumor immune microenvironment (TIME) is a complex system, composed of immune and cancer cells, tissues, and other biological components, that play an important role in the regulation and progression of cancer. Traditional methods like flow cytometry and scRNA-seq have frequently failed to capture the complex biology and mechanisms that drive immune cell interactions within this environment. However, advancements in spatial transcriptomics have transformed our understanding of cellular heterogeneity, enabling the identification of new cellular interactions and functional dynamics between different cell types and their microenvironments that were previously elusive. Spatial transcriptomics builds upon the principles of single-cell transcriptomics by preserving the spatial information of gene expression within tissue samples. This technology allows researchers to study gene expression in the context of tissue architecture, providing a more complete understanding of the cellular function and interactions between cells. Although this method preserves spatial molecular information, it only provides a snapshot of gene expression at a single point in time, limiting our ability to track molecular changes over time. To address this limitation, I propose developing a new method for spatiotemporal monitoring of live tissue transcriptomics, allowing us to capture how immune cells interact and respond to changes in the TIME during tumor development. In my predoctoral studies, I have engineered a nanostraw array with an electrophoresis setup for RNA extraction from live cells with high viability, and optimized a next-generation sequencing (NGS) spatial barcoding tool to preserve the spatial context of RNA molecules post-extraction, allowing for longitudinal cell tracking. For the F99 phase, I will further refine the nanostraw extraction setup and assess whether its integration with the NGS-based spatial barcoding platform can preserve extracted RNA molecules between individual cells. To validate the live spatiotemporal mapping technology's ability to monitor gene expression in the TIME over time, I will culture patient-derived tumor tissue on nanostraws and track transcriptional changes in cancer and immune cells over 2-3 weeks, while also examining spatial dynamics between these populations. This new technology will offer unprecedented insights into cellular behavior, revealing spatial distributions, cell-cell interactions, and disease processes, with transformative implications for cancer research. For the K00 phase, I will identify a combination of renowned bioengineering and immunotherapy labs to explore the evolution and rearrangement of the TIME during therapy and develop innovative tools at the intersection of synthetic and computational biology, integrating single-cell genomics with both spatial and temporal resolution. Completion of this project will successfully prepare me to launch an NIH-funded laboratory focused on designing novel sequencing platforms that bridge space and time to study TIME dynamics and tumor evolution, for therapeutic targeting and personalized medicine.

NIH Research Projects · FY 2025 · 2025-08

Project Summary/Abstract: An estimated 400 million people worldwide have experienced persisting and sometimes debilitating symptoms for months and years after SARS-CoV-2 infection, known as Long COVID (LC). No proven therapies exist to treat LC, and the underlying mechanisms driving disease remain poorly understood. Reactivation of herpesviruses such as Epstein-Barr virus (EBV) and varicella zoster virus (VZV) has been linked to LC, but it is unclear how SARS-CoV-2 infection alters immune responses to these common viruses. Recent studies have also provided emerging evidence for persistence of SARS-CoV-2 in some individuals with LC, but key questions remain unanswered, including how viral reservoirs persist, why the immune system fails to clear virus, whether persisting virus drives ongoing immune stimulation, and how therapies to treat LC will affect immune responses to SARS-CoV-2. Efforts to identify effective treatments for LC depend on answering these questions. Our preliminary data demonstrate elevated activation of SARS-CoV-2-, EBV-, or VZV-specific T cells in 40% of LC patients, providing one of the most sensitive measures of immunopathology in LC to date. Thus, there is an urgent need to investigate virus-specific T cell activation in LC to define underlying mechanisms of disease and identify promising therapeutic targets. The proposed research will respond to this need by testing three working hypotheses: first, that SARS-CoV-2-specific CD8 T cells survey tissue reservoirs of SARS-CoV-2 and sense viral antigens, but fail to clear the virus due to impaired functionality in LC; second, that therapies suppressing SARS-CoV-2 replication will reduce T cell stimulation in LC but may not achieve a durable cure; and third, that immune stimulation by common herpesviruses is associated with disease pathology in LC. These hypotheses will be tested by leveraging custom HLA-I/peptide tetramers to identify and sort rare virus-specific CD8 T cells from hundreds of longitudinal blood and tissue samples from people with LC and people who fully recovered after SARS-CoV-2 infection. Cells will be analyzed using spectral flow cytometry and single-cell sequencing approaches to elucidate the mechanisms driving LC pathology, identify promising therapeutic targets for LC, and investigate how immune stimulation by herpesviruses shapes human disease. This research will be integrated with a comprehensive training plan to develop skills in tissue immunology, single-cell sequencing, and bioinformatics, which will be reinforced with targeted coursework and professional development activities in scientific writing, responsible conduct of research, laboratory management, and mentorship of junior personnel. The research and training plan will take full advantage of the outstanding environment at the University of Pennsylvania and the wealth of expertise in Dr. John Wherry’s lab. Completing the proposed research and training will substantially advance the human immunology field and form the foundation of my research program as I open my independent laboratory, creating a pathway to my goal of being an impactful independent researcher in human infectious disease immunology.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY The goal of this proposal is to identify the mechanisms underlying the expansion of phosphatidylinositol glycan type A (PIGA) gene-mutated hematopoietic cells in patients with paroxysmal nocturnal hemoglobinuria (PNH) disease. PIGA is a gene required for the biosynthesis of glycosylphosphatidylinositol (GPI) anchors. Patients with PNH disease develop somatic mutations in PIGA in their hematopoietic stem and progenitor cells (HSPCs). PIGA protein deficient (PIGA–) HSPCs lack all GPI-anchored proteins and clonally expand, causing life-threat- ening hemolytic anemia and thrombosis in PNH patients. Although progress has been made in treating hemolytic anemia associated with PNH, current treatments do not reverse PIGA– HSPC expansion and require life-long adherence to complement inhibitor therapy. The main challenge to developing more effective and curative ther- apies for PNH has been a gap in our understanding of the effects of PIGA loss on HSPC function and causes of clonal expansion of PIGA– HSPCs. Here, we propose to fill these gaps by defining the effects of PIGA loss on HSPC function. We recently developed a mouse model of hematopoietic-specific PIGA loss, in which we can compare the hematopoietic function of PIGA– and control HSPCs directly in a competitive environment within the same animal by taking advantage of X chromosome inactivation. Our preliminary data suggest that PIGA– hematopoietic stem cells (HSCs) produce greater numbers of mature blood cells than PIGA+ HSCs and have hyperactive Notch signaling that may contribute to the enhanced PIGA– HSPC expansion under conditions of immune bone marrow injury. We hypothesize that PIGA– HSPCs have a competitive advantage in immune- mediated bone marrow failure, mediated in part through increased Notch signaling. The objective of this proposal is to determine the effects of PIGA loss on HSPC function and identify mechanisms that underlie clonal expan- sion of PIGA– HSPCs. In Aim 1, we will determine how PIGA loss affects HSPC function, at steady state and in the context of immune-mediated bone marrow failure. Aim 2 will investigate the hypothesis that hyperactive Notch signaling stimulates PIGA– HSPC expansion in steady state conditions and during stress hematopoiesis associated with immune-mediated bone marrow injury. In Aim 3, we will use integrated single-cell protein and DNA sequencing and transcriptome profiling to determine factors that cause PIGA– clone expansion in patients. Our studies will address critical knowledge gaps in understanding the effects of PIGA loss on HSC function and reasons for competitive advantage and clonal expansion of PIGA– cells. Insights gained from these studies will guide future mechanism-based strategies to prevent and reverse PIGA– clone expansion and restore healthy hematopoiesis in immune-mediated bone marrow failure disorders to cure PNH.

NIH Research Projects · FY 2025 · 2025-08

Filoviruses (Ebola [EBOV] and Marburg [MARV]) are emerging global pathogens that cause both acute hemorrhagic disease and chronic persistent infections with long-term sequelae and death. A comprehensive understanding of the filovirus-host interactome is critical for the development of novel host-oriented countermeasures to treat or prevent these deadly viral infections. We have recently identified YAP (Yes- Associated Protein; the downstream transcriptional effector of the Hippo pathway), and filamin A and B, members of a family of actin crosslinking/stabilizing proteins, as novel host determinants of viral infectivity, egress, and spread. Functionally, we find that filamin-A is critical for mediating infectivity of authentic EBOV and MARV in multiple cell types including primary human macrophages, while in contrast, filamin-B restricts filovirus infectivity. Filamin-A forms a mechanosensory complex with β1 integrin to transduce signals from the extracellular matrix (ECM) to internal signaling pathways such as the Hippo pathway, a critical regulator of proliferation, migration, and EMT (epithelial mesenchymal transition) induction. Previously, we found that filamin-A-deficient melanoma cells are softer than wild-type cells and maintain the Hippo pathway in an ON condition in which YAP is predominantly cytoplasmic. Together with our recent data, these findings support the novel hypothesis that ECM composition and stiffness will impact EBOV/MARV infectivity, with EBOV/MARV more easily infecting cells embedded in a stiffer ECM (where YAP is transcriptionally active) than in a softer ECM (where YAP is transcriptionally inactive). In Aim 1, we will test our hypothesis that ECM stiffness is a determinant of filovirus infectivity using innovative approaches, such as atomic force microscopy (AFM), viscoelastic polyacrylamide (PAA) hydrogels, and live filovirus infection assays. In preliminary studies to identify host proteins that functionally contribute to the robust and opposing effects of filamin-A and filamin-B on EBOV/MARV infectivity, we identified 5 ECM-associated proteins, a subset of which are YAP target genes. In Aim 2, we will use innovative approaches including live filovirus infection, pseudotype transductions, CRISPR/Cas9 KO cells, and RNA Seq. to determine whether expression of these 5 candidate ECM-associated proteins modulate filovirus infectivity. Intriguingly, EBOV infection induces EMT, and we have recently identified YAP as a novel EBOV/MARV VP40 interactor and regulator of filovirus egress and spread. Based on these data, in Aim 2 we will also test the novel hypothesis that filovirus infection will reciprocally impact the cellular environment by affecting YAP localization/transcriptional-activity and expression of ECM-associated proteins within the matrisome. Findings from this exploratory proposal will provide a solid foundation for future studies designed to interrogate more mechanistically the role of the ECM and matrisome in regulating filovirus infection, the subsequent host immune response, and the potential to manipulate the ECM to inhibit filovirus infection.

NIH Research Projects · FY 2026 · 2025-08

The overarching goal of this proposal is to further delineate the mitochondrial involvement in acute and chronic poisoning from organophosphates (OP). We will investigate the mitochondrial contribution to neurological, cardiac and microcirculation sequela with a focus on novel biomarker development and therapy. An estimated 385 million cases of acute unintentional pesticide poisoning occur worldwide each year, resulting in around over 300,000 fatalities from OPs alone. Sources of OP exposure occur in farming communities, occupational settings and as agents of threats. The primary mechanism of action is increased cholinergic activation leading to excessive muscarinic symptoms such as respiratory distress and seizures. In addition, chronic OP exposure can lead to long-term neurologic and cardiac sequelae such as motor dysfunction and heart failure. While the effects of OPs on the autonomic nervous system (acetylcholine) is thought to be the primary mechanism of injury, our group has mapped out significant mitochondrial involvement at Complex I. Survivors of acute poisoning and those with chronic exposure demonstrate long-term sequela often without any overt cholinergic symptoms. Significant knowledge gaps include: (1) the degree of mitochondrial injury contributing to long-term sequelae; (2) lack of tissue-specific biomarkers to gauge severity of disease and guide therapy; (3) lack of complementary treatment strategies to mitigate cardiac and neurologic disability. This project will further delineate the mitochondrial pathways involved in OP poisoning using extracellular cell-free DNA (cfDNA) as a biomarker in a rodent model of OP poisoning, furthering the mechanistic understanding of OP poisoning and developing cfDNA as potential biomarker and tissue source using DNA methylation patterns. CfDNA is an established prognostic indicator for mortality in an array of disease states. CfDNA is released into circulation upon injury and have been used as a marker of disease severity. We also propose to investigate a select mitochondrial-based therapeutic in vivo to improve mitochondrial respiration, sustain cellular function, and limit organ injury circumventing the known Complex I effect seen in our supporting publications and preliminary data.

NSF Awards · FY 2025 · 2025-08

Mechanical metamaterial, i.e., engineering materials whose properties arise from geometry rather than chemical composition, hold great promise for applications in aerospace, biomedical engineering, robotics, and beyond. By integrating active elements, these materials can respond dynamically to environmental stimuli such as heat or light, enabling adaptive and programmable behavior. Despite their potential, the practical design of such materials remains constrained by computationally expensive and highly specialized modeling tools. This award supports fundamental research to establish a general, experimentally validated continuum modeling framework for active mechanical metamaterials. This framework will enable efficient prediction of material behavior and systematic design across a wide range of geometries and actuation mechanisms. The project advances fundamental understanding while supporting technological innovation in advanced manufacturing, adaptive devices, and reconfigurable structures. It also leverages interdisciplinary collaboration and integrated fabrication-modeling workflows to train students across educational levels and to engage the public through outreach initiatives. The research integrates theoretical, experimental, and computational approaches. A continuum modeling framework will be constructed to connect unit-cell geometry with macroscopic deformation, including both planar and three-dimensional behaviors. The models will incorporate internal variables and compatibility conditions to capture soft deformation modes and the effects of actuation. Experimental work will validate the framework through fabrication, including direct ink writing, molding, and conventional 3D printing, and mechanical testing of passive and active metamaterials, making use of digital image correlation to quantify deformation. A systematic scaling study will define the limits of continuum applicability. The final phase will implement the models into finite element simulations to enable inverse design, allowing metamaterials to be tailored for specific responses. Collectively, these efforts will yield general design principles and computational tools to accelerate the adoption of mechanical metamaterials in advanced engineering applications. 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 2025 · 2025-08

ABSTRACT This proposal provides salary support for a highly productive Research Specialist, Dr. Charly Good, working in the lab of a well-established Unit Director, Dr. Shelley Berger. This proposed work is an integral part of Dr. Berger's NCI funded cancer research program, which focuses on understanding how wildtype (WT) and gainof- function (GOF) mutant p53 sculpt the genome for gene regulation. p53 is a transcriptional activator that directly binds DNA to induce tumor suppressor gene expression. Dr. Berger's group has made seminal discoveries in how p53 functions in cancer, including the groundbreaking discovery that select hotspot mutations in p53 (GOF p53) confers oncogenic abilities through activation of key epigenetic pathways. Understanding how GOF p53 regulates its target genes is a major question that, after exhaustive study, still remains unanswered. Enhancer clusters (ECs) are groups of enhancers that cluster together in 3D space to drive high levels of gene expression. Dr. Good is currently addressing the following questions. 1) what proteins interact with GOF p53 to drive cancer growth? 2) does GOF p53 reorganize ECs to drive high levels of gene expression? 3) what transcription factor(s) does GOF p53 collaborate with to rewire ECs? 4) what target genes does GOF p53 regulate via rewiring of ECs? P53 also regulates genes by bringing them close to nuclear speckles (NS) to boost expression. Dr. Good is mentoring trainees in the lab to 5) identify which GOF p53 target genes associate with NS and test the function of this regulation and 6) identify how p53 associates with NS by identifying the proteins that interact with the praline rich domain of p53. Dr. Good will be responsible for: (1) generating the high-throughput HiChlP, RNA-seq, and Poree datasets in cancer cells with WT and GOF p53 mutations; (2) analyzing the sequencing data to identify how normal WT p53 functions at ECs and importantly, how GOF p53 rewires ECs to promote cancer growth; (3) performing molecular biology experiments and cancer assays to test the functional consequences of disrupting critical ECs and nuclear speckle association using CRISPR gene editing approaches. This work will be viewed with a translational lens, whereby all genomics data will be analyzed in a way that offers clinically relevant insights to improve upon patient therapy. Dr. Good is also involved in the ongoing T cell work in the Unit Director's lab, publishing several first-author papers on T cell exhaustion in cancer and mentoring trainees in this area. Dr. Good has worked with Dr. Berger since 2017. Together, they have published numerous high-quality manuscripts and have advanced our understanding of cancer biology. Dr. Good is a key resource in Dr. Berger's laboratory, where she directly mentors graduate students and postdocs and teaches both wet and dry lab techniques to trainees. The Research Specialist award would be a tremendous opportunity for Dr. Good and would support her career development as a staff scientist. She will be instrumental in advancing the robust cancer research program in Dr. Berger's laboratory and will help to train the next generation of scientists.

NIH Research Projects · FY 2025 · 2025-08

Project Summary/Abstract While imatinib remains the gold standard for systemic therapy in gastrointestinal stromal tumors (GISTs), the vast majority eventually progress while on treatment and some patients present with resistant tumors. While novel immunotherapies have shown potential in preclinical models, they have yet to demonstrate significant clinical efficacy alone or in combination with tyrosine kinase inhibition. Tumor associated macrophages (TAMs) have been shown to be essential in antitumoral immunity in GIST, and there is evidence that their activation can synergize with imatinib therapy. Dectin-1 is a pattern recognition receptor (PRR) present on TAMs originally described for its ability to stimulate an immune response to fungal pathogens. Activation of dectin-1 on TAMs and dendritic cells (DCs) has been shown to stimulate antitumoral immunity in some preclinical models of solid tumors but there are discrepant results. We found that dectin-1 is highly expressed on TAMs and DCs in a genetically engineered mouse model of GIST, as well as TAMs from human GISTs, regardless of imatinib treatment status. Dectin-1 appears to be largely restricted to M1-like TAMs and DCs, as opposed to tumor cells or other immune cell types. As such, we hypothesize that dectin-1 activation will cause GIST regression via TAMs and DCs and this effect will be augmented by imatinib therapy. Since dectin-1 likely works via immune cells, it will be therapeutic in imatinib-resistant GIST. We will study to role of dectin-1 in the immune response to GIST by performing flow cytometry, RNA seq, and in vitro functional experiments on TAMs and DCs derived from mice treated with vehicle or d-zymosan, a specific dectin-1 agonist. Furthermore, we will isolate TAMs and DCs from mouse and human GISTs and culture them with d-zymosan to determine its direct effects. To determine the anti-tumor efficacy of dectin-1 activation in our mouse model, we will assess tumor size, histology, and KIT signaling after d-zymosan administration. The combination of dectin-1 activation and tyrosine kinase inhibition will be tested in imatinib-sensitive and imatinib-resistant mouse models. The mechanism of dectin-1 activation will be elucidated by depleting TAMs or DCs. The work will take place in the well- established lab of Dr. DeMatteo, a world leader in GIST research, and exemplary surgeon-scientist at the University of Pennsylvania. The fellowship will include immersion in a collaborative and dynamic lab environment, regular seminars, attendance at national conferences, and mentoring of medical and undergraduate students.

NIH Research Projects · FY 2025 · 2025-08

Project Summary Heart failure with preserved ejection fraction (HFpEF) is a complex and heterogeneous clinical syndrome accounting for over 50% of patients with heart failure. Major HFpEF risk factors include age, hypertension, diabetes, kidney disease and atherosclerosis. Prior studies have identified distinct phenogroups within the overall HFpEF population which are defined based on different predisposing co-morbidities. However, the links between these phenogroups and underlying pathophysiologic processes are poorly defined with limited tissue-based evidence. Recognizing that obesity and type-2 diabetes are pro-inflammatory, and that oxidative stress produces microvascular injury, I hypothesize that obesity and diabetes promote pathological structural and functional microvasculature remodeling that contribute to the development of HFpEF. I will test this hypothesis with a combination of observational and interventional studies using an animal model of HFpEF that is well-matched to the task. The ZSF1-obese rat is a genetic model of HFpEF with two leptin receptor mutations resulting in an obese, diabetic and hypertensive phenotype impacting multiple organ systems. Control ZSF-1 lean rats have a single leptin receptor mutation and develop hypertension without obesity or diabetes. I previously validated the accelerated development of HFpEF in ZSF1-obese vs. ZSF1- lean rats. In Aim 1, I will compare the temporal progression of microvascular structural and molecular changes in ZSF1-obese and lean rat myocardium using advanced morphological characterizations and spatial transcriptomic and proteomic techniques that resolve to cell type-specific expression patterns. In parallel, I will perform the same assays on banked myocardial specimens from a cohort of patients with features of HFpEF and relevant controls. In Aim 2, I will connect these tissue-based findings from the obese and lean rats with in vivo assessments of coronary microvascular function using micro single-photon emission computed tomography (µSPCT). In Aim 3, I will employ treatment with a GLP-1 receptor to test whether mitigation of obesity and diabetes can prevent and/or treat microvascular remodeling in the ZSF1- obese rat. Beyond the scientific merits of the proposed research, conducting these studies will broaden my understanding of cardiovascular pathophysiology and help develop my proficiency with several versatile and powerful techniques. My primary mentor, Dr. Ken Margulies, is a senior physician-scientist with expertise in myocardial pathophysiology and translational heart failure research using preclinical models and human myocardium. I have an advisory committee with expertise in each new topic area or technique (microvasculature, quantitative image analysis, spatial transcriptomics and proteomics, bioinformatics), and they are committed to my training. I have the full support of my institution where the environment favors dynamic and collaborative interactions among translational researchers and clinical experts. Together, the research, training, mentors and institutional environment assures my successful transition to independence.

NIH Research Projects · FY 2025 · 2025-08

ABSTRACT Vellus hairs are short and fine hairs that cover the human body and are essential for facilitating sweating, the major mechanism for human thermoregulatory cooling. Vellus hairs are formed during human gestation, in response to a shift in the growth cycle of human fetal hair follicles. During early fetal development, hair follicles first build large lanugo hairs, which are subsequently shed before birth. Starting around 20 weeks of gestation, fetal hair follicles undergo a reduction in the duration of their active growth phase (anagen), leading to the formation of smaller vellus hairs that predominate human skin post-partum. Determining the mechanisms that regulate the lanugo to vellus transition is critical in understanding the basis for this important thermoregulatory trait. A hallmark of the period that marks the onset of the lanugo to vellus transition in human fetal skin is the upregulated expression of a transcription factor, Engrailed1 (EN1). During this timeframe, EN1 expression is upregulated in multiple epidermal populations of the human fetal hair follicle, most notably in the hair matrix, the progenitor population from which the hair shaft derives. EN1 also becomes upregulated in the dermal papilla, the mesenchymal niche that regulates the hair cycle and hair production. In light of the concordance of EN1 upregulation and the onset of the lanugo to vellus transition, our lab turned to a humanized transgenic mouse model that allows for tunable En1 expression in the epidermis of the hair follicle in a manner consistent with human fetal skin. This model demonstrated that mice with ectopic En1 expression in the epidermis have shorter and thinner hairs due to a truncated growth phase (anagen) of the hair cycle, recapitulating key hair size diminution events observed during the lanugo to vellus transition. Considering these data, the hypothesis of this proposal is that En1 regulates hair size by altering the function of the hair matrix and the dermal papilla, two key populations involved in hair production. Accordingly, the aims of this proposal are to define the cellular and transcriptional mediators by which En1 functions in the two major skin compartments, epidermis (Aim 1) and dermal papilla (Aim 2), to alter hair follicle cycling. The proposed experiments will uncover the first mechanism underlying human vellus hair formation. Beyond their implications for understanding human-specific biology, the findings derived from this proposal will be critical for developing an integrative approach to comprehensive human skin regeneration.

NIH Research Projects · FY 2025 · 2025-08

Project Summary Amyotrophic lateral sclerosis (ALS) and frontotemporal degeneration (FTD) represent two extremes of an ALS- FTD spectrum, frequently characterized by TDP-43 pathological inclusions and shared genetic risk factors. We have defined normative criteria for characterizing cognitive impairment in ALS and demonstrated that 22% of ALS are cognitively impaired and, in autopsy-confirmed TDP-43 cases, there is an overall >4 hazard ratio for individuals with ALS or FTD onset to develop features of both ALS-FTD. However, prognosis is highly variable across individuals in the ALS-FTD spectrum and risk factors for developing co-occurring motor neuron dysfunction and cognitive impairment are largely unknown, even within C9orf72 repeat expansion families. The overall hypothesis of this proposal is that genetic modifiers of the FTD-ALS spectrum will identify factors predictive of ALS or FTD onset and prognosis for developing ALS-FTD. The discovery of genetic modifiers that contribute to ALS-FTD phenotypic heterogeneity will improve prognostication and clinical trial stratification while enhancing our mechanistic understanding of these highly variable and devastating disorders. This proposal will focus on two strategies for identifying genetic modifiers of binary (ALS vs FTD) and quantitative (neuropsychological and motor) traits. We will leverage the highly integrated Penn Comprehensive ALS Clinic and Penn Frontotemporal Degeneration Center, which have common procedures for neuropsychological and motor phenotyping as well as DNA and biofluid banking. First, we aim to identify polygenic modifiers of cognitive and motor features of ALS-FTD spectrum. We will generate traditional and novel module-based polygenic risk scores (PRS) of common variants associated with “disease-specific” ALS or FTD risk or associated with “domain- specific” heritable traits including cognitive and motor function. We will relate each PRS to cross-sectional and longitudinal neuropsychological and motor/functional scales. We will additionally perform Mendelian Randomization to identify causal associations between heritable traits and disease-specific outcomes. We hypothesize common genetic variation reflected by PRS will modify onset type and quantitative trait measures of cognition and motor impairment. Second, we aim to identify structural variants through long-read sequencing (LRS) of individuals and families that modify cognitive and motor features of the ALS-FTD spectrum. We will use LRS to identify structural variants (SVs) and use rare-variant aggregated and burden tests to identify SVs and genes associated with onset type and quantitative trait measures of cognition and motor. We will additionally use variants component linkage analysis in our well-characterized three-generation pedigrees to identify phenotypic modifiers. We hypothesize both strategies will uncover novel sources of common and rare genetic variation that contribute to heterogeneity in the ALS-FTD spectrum.

NIH Research Projects · FY 2025 · 2025-08

ABSTRACT Acute myeloid leukemia (AML) is the most common and fatal form of adult leukemia, with less than a quarter of patients surviving past 5 years. Outcomes are especially poor in older patients, who are not candidates for traditional “7+3” chemotherapy. They are also often refractory to or relapse on newer therapies such as BCL-2 inhibitor Venetoclax and hypomethylating agent Azacitidine. Patient xenograft and mouse models have shown that the bone marrow (BM) microenvironment plays an important role in chemoresistance and relapse. The recent emergence of spatially-resolved proteomic and transcriptomic technologies opens possibilities to examine microenvironmental spatial relationships and cellular interactions at high resolution. Examination of the AML BM microenvironment with these technologies, particularly in the context of chemotherapy, has yet to be comprehensively explored. Previous work in our lab has included using CO-detection by InDEXing (CODEX), a multiplexed immunofluorescence spatial profiling technology, to map the biogeography of the normal BM environment, as well as a few NPM1 mutant chemoresistant AML samples. We will extend this effort to a cohort of 30 AML patients to examine spatial patterns of the BM microenvironment to reveal its role in AML chemoresistance and relapse. In addition, our lab has previously developed CytoCommunity, a method for detection of tissue cellular neighborhoods, or spatial regions of distinct and homogeneous cell type composition. These neighborhoods may indicate cell types organizing to form functional niches. Several other tools have also been developed to perform neighborhood detection. However, none yet can integrate neighborhood analyses across multiple samples to assess changes over time or in response to treatment. We will develop methods for tracing neighborhood changes over time and across conditions and apply them to our spatial analysis of AML. We will examine how AML cancer cells and other elements of the microenvironment spatially reassort in response to various treatments through analysis of neighborhood evolution and other spatial statistical methods. This will include subclone-level spatial analysis to identify how intrinsic (genetic) mechanisms of resistance correlate and interact with extrinsic ones. Finally, we will use CODEX-generated cell type labels to train a computer vision model to automatically annotate cell types on co-registered H&E images. This will facilitate automatic cell type labeling on the vast supply of H&E images without corresponding CODEX data, which will allow us to expand our spatial analysis to a much broader cohort. Overall, our project aims to generate biological insights into the microenvironment’s role in AML chemoresistance and relapse which will facilitate the improvement of treatment strategies and outcomes. In addition, we will develop neighborhood evolution and automatic cell type annotation methods that will enhance future research in spatial analysis of the microenvironment in leukemia and cancer more broadly.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY Assisted Reproductive Technologies (ART) are non-coital methods of achieving a pregnancy, and most often involve the manipulation of gametes or embryos. Importantly, the use of ART is increasing as they become more accessible and more couples experience infertility or postpone childbearing. In vitro fertilization (IVF) is one of the most common ART used in clinics today, in which eggs are combined with sperm and cultured until the blastocyst stage before transfer to a recipient female. Although beneficial and safe, IVF procedures have been associated with suboptimal outcomes for mother and baby including pre-eclampsia, fetal and placental growth abnormalities, an increased rate of rare imprinting disorders, and dysregulated postnatal metabolism. Indeed, human IVF children experience elevated body weight, glucose intolerance, and insulin resistance beginning in adolescence. These outcomes may result from IVF procedures, especially embryo culture, which exposes a developing embryo to altered environmental conditions not normally experienced in vivo during a time in which the epigenome must be reprogrammed for proper development. One important environmental factor during embryonic development is oxygen (O2), which can influence gene expression, metabolism, and the activity of important epigenetic enzymes. Today, the lowest O2 tension used during embryo culture is 5%, despite evidence that sections of the mammalian female reproductive tract have O2 levels as low as 2%. Thus, decreased O2 tension during embryo culture may better mimic the in vivo environment, leading to improved pre- and postnatal outcomes in the offspring. Consistently, preliminary studies from my thesis lab have demonstrated that culture at 2% O2 improves blastocyst morphology, molecular signatures in embryonic liver, and placental outcomes compared to culture at 5% O2. Given that culture at 2% O2 improves some offspring outcomes, I hypothesize that the molecular profiles of embryos cultured at 2% O2 will more closely resemble naturally-conceived embryos (Naturals). Additionally, I hypothesize adult IVF offspring after 2% O2 culture (IVF 2%) will display less severe metabolic outcomes compared to 5% O2 culture (IVF 5%) IVF offspring. To test these important hypotheses, I will use our validated IVF mouse model, which recapitulates many of the phenotypes observed after IVF in humans. In Aim 1, I will explore how culture under difference oxygen tensions (2 vs. 5%) impacts gene expression, DNA methylation, and histone posttranslational modifications in blastocysts. In Aim 2 I will determine if metabolic outcomes in adult IVF offspring are improved after culture at 2% O2, and whether this improvement translates to the transcriptome and methylome of the liver. This project will determine how embryo culture at 2% O2 impacts both preimplantation development and adult metabolic health outcomes. Overall, this research will contribute foundational insights to our understanding of the role of O2 in early development, with key implications for clinical IVF practices.

NIH Research Projects · FY 2026 · 2025-08

Project Summary The quest for modulators of protein interactions is undergoing a renaissance with advent of powerful computa- tional methods for the prediction and de novo design of protein structures. However, translating this static struc- tural information into an accurate prediction of the thermodynamics and kinetics of dynamic protein interactions remains a challenge, limiting our ability to robustly design tight-binding modulators of protein interactions. The limitations of existing structure-based drug design approaches can be traced back to their accounting of collective hydration effects using additive approximations, such as hydropathy scales, and their tendency to adopt a static perspective, assuming a relatively rigid protein. Thus, there is a critical need for developing novel computational methods capable of accurately accounting for the collective reorganization of the hydrogen bonding network of water as well as any conformational changes that accompany binding. To address this critical knowledge gap, the proposed program seeks to develop novel hydration-based, enhanced-sampling molecular simulation techniques for characterizing the reorganization of water structure in response to conformational change and binding, and to use such a characterization for informing the thermodynamics and kinetics of protein interactions as well as designing interaction modulators. These techniques will be developed within the context of three judiciously cho- sen projects spanning a diversity of protein interactions: (i) interactions between reader proteins and methylated histone tails, (ii) interactions between galectins and carbohydrates, and (iii) interactions between the p38! MAP kinase and single-domain antibodies. For each project, close interactions with experimental collaborators will facilitate the refinement of our methods and the validation of our predictions, enabling us to uncover the molec- ular underpinnings of specificity. By developing novel computational methods to bridge the gap from protein structures to affinities, and leveraging these methods to characterize the interactions of proteins with peptides, carbohydrates, and other proteins, the proposed program thus seeks to transform our ability to predict, under- stand and modulate protein interactions.

NIH Research Projects · FY 2025 · 2025-08

Project Summary Chimeric Antigen Receptor T (CAR T) cell therapy has been highly effective in a subset of hematological malignancies. However, this efficacy has not translated to solid tumors for a variety of reasons, including limited persistence in an immunosuppressive and metabolically challenging tumor microenvironment. Unbiased techniques such as genome-scale CRISPR screens have identified several pathways to enhance therapeutic efficacy. However, these screens have mainly utilized reductionist in vitro approaches, and do not recapture a solid tumor microenvironment. I have performed a genome-wide in vivo CRISPR screen of human CAR T persistence in solid tumors. The top two hits were ATXN7L3 and USP22, components of the Spt-Ada-Gcn5 acetyltransferase (SAGA) complex deubiquitylation module (DUB). The SAGA DUB is primarily known for its role in histone H2B deubiquitination, but its role in T cell persistence and antitumor function remains to be described. I show knockout of USP22 or ATXN7L3 (USPKO/ATXKO) increased T cell memory markers and enhanced CAR T expansion and persistence in vitro. Paradoxically, USPKO/ATXKO CAR T exhibit a defect in IL- 2 secretion and fail to enhance antitumor immunity or persistence in vivo. The data indicate that USP22 and ATXN7L3 are fundamental regulators of CAR T cell function, yet the mechanisms through which they influence CAR T biology remain unclear. I hypothesize that knockout of USP22 or ATXN7L3 reprograms the CAR T transcriptome through changes in histone ubiquitination and augments CAR T persistence in an IL-2 dependent manner. In Aim 1, I propose to determine the relationship between cell-extrinsic IL-2 and USPKO/ATXKO CAR T persistence in solid tumors. I postulate that cell-extrinsic IL-2 is sufficient to rescue USPKO/ATXKO persistence and enhance antitumor activity. As USPKO/ATXKO CAR T persist in a pooled in vivo setting, I will pool wildtype CAR T cells with USPKO/ATXKO CAR T at different ratios and infuse into tumor-bearing mice to assess persistence and antitumor activity. I will also determine the effects of exogenous IL-2 supplementation on in vivo USPKO/ATXKO CAR T persistence through serial recombinant IL-2 infusions. In Aim 2, I propose to investigate the impact of USPKO/ATXKO and histone deubiquitination on CAR T transcriptional programs. I hypothesize that H2B deubiquitination through SAGA regulates gene expression programs that promote CAR T persistence. To evaluate this, I will perform bulk RNA-sequencing and histone ubiquitin CUT&RUN-sequencing on USPKO/ATXKO to determine the extent to which histone ubiquitin changes affect these programs. Together, the findings of this work will describe a novel role for the SAGA complex and histone ubiquitination in CAR T persistence and has potential for direct clinical translation for treatment of solid tumors.

NIH Research Projects · FY 2025 · 2025-08

Project Summary The goal of this project is to elucidate the biochemical mechanisms of SCMH1, an understudied accessory subunit of Polycomb Repressive Complex 1 (PRC1), during cellular differentiation. Polycomb group (PcG) proteins are critical epigenetic regulators that maintain transcriptional repression during development. Dysregulation of PcG proteins is associated with neurodevelopmental disorders, including Weaver syndrome, and can disrupt normal embryonic development. PRC1 plays a central role in chromatin compaction, long-range chromatin interactions known as Polycomb loops, and the formation of subnuclear clusters called Polycomb bodies. Despite the importance of PRC1, the biochemical role of SCMH1 in Polycomb-mediated repression remains poorly understood. The Drosophila homolog of SCMH1, SCM, is essential for maintaining epigenetic memory during development, but whether SCMH1 fulfills a similar function in mammals is unknown. Preliminary data from our lab reveal that SCMH1 loss leads to ectopic activation of Polycomb target genes, despite unaltered PRC1 occupancy and histone marks. We identified a conserved RNA-binding region (RBR) in SCMH1, suggesting that RNA binding may contribute to its function. Hi-C analysis shows that SCMH1 loss reduces Polycomb loops, implicating it in chromatin architecture. Given its RNA-binding capability and features associated with molecular condensation, such as intrinsically disordered regions and a SAM domain, we hypothesize that SCMH1 contributes to Polycomb body formation and gene regulation through RNA-mediated interactions and phase separation. Aim 1 investigates the role of SCMH1-RNA binding in Polycomb-mediated repression. Using a degron-tagged mouse embryonic stem cell (ESC) line, already in hand, I will degrade and rescue SCMH1 with wild-type or RNA- binding mutants and evaluate transcriptional repression, SCMH1 chromatin occupancy, and differentiation potential. Aim 2 examines the relationship between SCMH1 and Polycomb bodies and Polycomb loops. I will determine the SCMH1 domains required for Polycomb body localization and formation using mutants lacking phase- separation-associated domains. Immunofluorescence and Hi-C will assess how Polycomb body and loop dynamics depend on SCMH1, particularly during ESC differentiation into epiblast-like cells (EpiLCs). Together, these experiments will uncover SCMH1's role in Polycomb repression, nuclear condensation, and chromatin organization, advancing our understanding of epigenetic regulation and the mechanisms underlying Polycomb function during development. My diverse training background, combined with the expertise of my sponsor and the collaborative nature of my research environment, will ensure the success of this project and my development as a researcher.

NSF Awards · FY 2025 · 2025-08

Non-technical Abstract All around us are materials that appear to be solid, but that actually “creep” – they flow in a very slow and mysterious manner. The “glacial pace” of flowing ice is an example of creep. Creep is everywhere in nature but hard to pin down, because the rate and style of creep is sensitive to how the material was made, the tiny forces acting on it, and even background vibrations or noise. From a practical point of view, creep can be both good and bad. It can help smooth out weak spots in a material, a process that is used to make some metals stronger. But under the wrong conditions, it can also lead to sudden and catastrophic failures such as a landslide. Consider a pile of sand. It looks still, but the grains actually jiggle slightly with changes in air pressure and temperature. Over time, this jiggling can strengthen or weaken the sandpile, showing behavior that is surprisingly similar to other disordered solids like glass. The goal of this project is to understand what causes this creep in (disordered) materials like sand, toothpaste, or mud, using novel experimental methods and theoretical analysis. Experiments will track how particles inside these materials move under controlled forces that mimic nature, and will also measure how material strength changes as creep progresses. These data will be used to develop new computer models capable of predicting creep behavior, that will help to design new materials and prevent landslides. Finally, this project contributes to workforce development by training graduate and undergraduate students in soft condensed matter & materials research. The proposed work includes educational and outreach activities aimed at developing and engaging the local STEM community. Technical Abstract This proposal is about creep. The slow deformation of apparently solid materials that are both ubiquitous in nature and exquisitely sensitive to material preparation, local stresses, and noise. Creep may strengthen a given material as particle rearrangements ’iron out’ weak spots but may also weaken and lead to catastrophic failure when particle motion becomes more cooperative (e.g., avalanches) under some disturbances. Here, we aim to develop a universal understanding of creep in disordered materials by investigating the structure and dynamics of slow, sub-yield deformation in dense particulate systems -- and how these processes relate to bulk mechanical strength. We propose novel experimental investigations that traps creeping jammed suspensions at a 2D interface and in a 3D transparent bath, tunes particle interactions, and perturbs the samples with a spectrum of disturbances that relax or excite them. We will observe how materials deform at the particle level (i.e., sample microstructure) while simultaneously measuring the bulk rheology (e.g., creep compliance). We seek to relate sample microstructure to dynamics/rheology using the concept of excess entropy as an order parameter. This approach unites thermal and athermal systems and allows us to build a constitutive model that captures the pathologies of creep in disordered materials. Finally, the project aims to educate and train the next generation of physicists and engineers with expertise in soft-condensed matter, rheology, statistical physics, and mechanics. 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 2025 · 2025-08

This proposal will establish the first International Conference on Research in Williams Syndrome organized by the Armellino Center of Excellence for Williams Syndrome (ACE-WS). We intend to establish a biennial conference as there are no recurring scientific meetings devoted to research in WS. Williams syndrome (WS) is a rare (1 in 7500 births) neurodevelopmental syndrome caused by hemizygous deletion of ~28 genes at 7q11.23. WS is characterized by developmental delay, distinctive cognitive and behavioral phenotypes, visual processing deficits, cardiovascular defects, gastrointestinal problems, endocrine issues, and more. For a rare disease, it is particularly important to provide an inclusive venue for communicating ongoing research findings and enabling multidisciplinary collaboration and resource sharing between research groups focused on WS, as well as facilitating the introduction and retention of new investigators to the field, including both collaborators in related fields as well as trainees and junior investigators entering WS research. There has been recent progress in the field in preclinical studies addressing potential treatments and interventions to mitigate some of the health and behavior issues associated with WS, as well as some early clinical trials in progress. A major goal of the Armellino Center is to expand research efforts and support studies with a particular focus on translational science. This is a key time to establish a recurring WS scientific meeting in order to facilitate preclinical readiness studies, develop robust outcome measures for future trials, and build networks that will support multisite trials. The ACE-WS partners with the Williams Syndrome Association (WSA; a national non-profit organization supporting families impacted by Williams syndrome) and ACE-WS is a member of the WS Clinical Consortium overseen by the WSA (consisting of the 10 WS Specialty Clinics in the US, including the clinic operated by ACE-WS). This Consortium operates the Collaborative Registry for Williams Syndrome (CReWS), a registry which unites both patient/caregiver-entered data and data entered by the WS Specialty Clinics and will serve as a significant research resource for the WS research community. These partnerships ensure a wide reach and will foster ongoing collaborations developed in this and future meetings.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY Digital health interventions (DHIs) have shown significant promise in promoting HIV prevention and care among at-risk youth. Despite the great potential for behavior change that DHIs offer, the effectiveness of DHIs has varied across studies, largely due to the varying degrees of participant engagement. Therefore, identifying key engagement characteristics in DHIs is important to evaluate the efficacy of DHIs with rigor and enhance the efficacy of DHIs for at-risk youth by promoting engagement. DHI studies have utilized single indicators of engagement, such as amount, frequency, and duration, and have focused on cross- sectional relationships between engagement and outcomes. This approach has limited the ability to evaluate the multidimensional aspects of engagement and the prospective effects of engagement on outcomes over time. Moreover, the one-size-fits-all approach may not be suitable for designing DHIs for youth. Therefore, there is a need to analyze engagement across multicenter research trials dedicated to HIV prevention and care for youth in the US. Our research aims to identify key characteristics of engagement to enhance the efficacy of DHIs for at-risk youth by pooling data from Adolescent Medicine Trials Network for HIV Interventions (ATN) - UNC/Emory Center for Innovative Technology (iTech) trials. In this study, we propose using secondary data from more than 900 individuals who participated across five ATN-iTech trials (MyChoices, LYNX, TechStep, Get Connected, and P3). The study's specific aims are: 1) To harmonize archived paradata into common data elements of participants' DHI engagement across ATN trials; 2) To identify engagement typologies and examine their associations with HPC outcomes; and 3) To examine longitudinal associations between participants’ prospective engagement and desired HPC outcomes over time. This research significantly contributes to current scientific knowledge by detangle the complex relationship between engagement and DHI outcomes to optimize DHIs for youth by pooling a multicenter research network in the U.S. Our findings will help to understand the characteristics driving higher engagement, rigorously evaluate the efficacy of DHIs while considering engagement, and propose optimized strategies to promote engagement, ultimately enhancing the efficacy of DHIs for youth. These insights will provide a blueprint for researchers to rigorously evaluate DHIs and enhance their effect sizes for adolescents and young adults at risk for HIV and inform the development of an implementation science framework for engagement in digital HIV interventions, which may be used in future studies for at-risk youth.

NSF Awards · FY 2025 · 2025-08

Your mouth holds a surprising amount of information about current and future health, not just for teeth and gums, but also for serious conditions like neurological problems, heart disease, and cancer. This project will develop a new way to predict your risk for these diseases using a new handheld robotic device to carefully and quickly collect samples of plaque bacteria from hard-to-reach places in the mouth. Then, artificial intelligence will be used to analyze these samples. The system will automate plaque sample collection and detection of harmful bacteria. This information will feed into a computer model designed to predict your likelihood of developing issues like gum disease, displaying your risk on an easy-to-understand dashboard. Ultimately, because the health of the mouth is so closely linked to overall well-being, this innovative approach will pave the way for early detection and treatment for a wide range of diseases, leading to better health outcomes for all. The oral cavity holds a wealth of information related to current and future health status. This information is naturally related to oral health status, but is also increasingly related to systemic health status with implications for neurological disease, cardiovascular diseases, and cancer. The overall objective in this project is to create an approach for identifying disease risk by sampling microbial plaque (biofilm) with micro-robotic techniques and analyzing the state of the oral microbiome using artificial intelligence. This will combine a collection method and data workflow, integrating micro-robotics with bioinformatics. Sampling will be enabled by using micro-robotics for precise and rapid biofilm removal from hard-to-reach surfaces at risk for periodontal (gum) diseases. This platform will provide automated collection and real-time pathogen detection while minimizing the need for laborious sample removal and processing steps. The multimodal data generated from the device will be used to develop a temporal graph neural network model, which will assist in identifying the likelihood of periodontal disease with predicted outcomes represented in an interactive risk-assessment dashboard. Moreover, given the strong connection between the oral microbiome and systemic health, the platform will lay the groundwork for monitoring and identifying risks associated with a host of diseases, leading to early interventions and improving long-term health outcomes. 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 2025 · 2025-08

PROJECT ABSTRACT Aberrant cardiac metabolism has long been hypothesized to contribute to heart failure (HF). However, understanding of cardiac metabolism in HF remains incomplete. To fill this gap, we performed metabolomic and RNA sequencing analysis of 48 non-failing (NF) and 39 dilated cardiomyopathy (DCM) hearts. Among many differences, metabolites from one-carbon metabolism were significantly low, especially methionine and s-adenosylmethionine (SAM), a universal methyl donor of methylation. In parallel, we also observed significant decrease of creatine, and increase of phosphatidylcholine (PC) in DCM hearts, which both are methylation product of SAM. Creatine is a critical metabolite for cardiac contraction, and PC is the most abundant phospholipids in the membrane. These observations and additional preliminary data have led to the hypothesis that cardiac injury causes membrane remodeling and excessive PC synthesis that drains the SAM pool, suppressing the generation of creatine, and ultimately contributing to cardiac metabolic insufficiency. Therefore, in Aim 1, I will lay groundwork for one-carbon metabolism in the heart by quantitively define cardiac- methionine/SAM flux and dynamics in vitro and in vivo using isotope labeled methionine tracing. In Aim 2, I will test if SAM deficiency can cause or worsen heart failure using cardiac specific SAM depletion mouse model. In Aim 3, I will quantify cardiac creatine synthesis rates and estimate contribution of reduced creatine in HF caused by low SAM levels. In Aim 4, I will test if cardiac remodeling increases PC synthesis and drains the SAM pool. The support from K99/R00 pathway to independence award will provide the applicant with the necessary training in cardiac metabolism to successfully achieve goals of each aim. This training will be guided and supported by a team of world class scientists from University of Pennsylvania, Cincinnati children’s hospital and Princeton University. In conclusion, this project will lay groundwork for one-carbon metabolism in the heart and identify cause and consequences of low SAM in the heart failure, which will be a pioneering discovery that may lead to new options for diagnosis and treatment of heart failure.

NIH Research Projects · FY 2025 · 2025-08

Enter the text here that is the new abstract information for your application. Alcohol Use Disorder (AUD) is a chronic, relapsing brain disorder that accounts for 5% of deaths globally. Available treatments are not efficacious for all patients or not well tolerated, thus there is a critical need to identify novel, well-tolerated treatments to reduce alcohol craving and consumption. Individuals with AUD have low brain glucose and a high rate of acetate metabolism that persist beyond acute intoxication. Ketones (β-hydroxybutyrate [BHB] and acetoacetate) structurally resemble acetate and provide an alternative to glucose as a source of energy to the brain. We and others have found that elevating ketones with a ketogenic diet reduced alcohol consumption and signs of alcohol withdrawal in alcohol-dependent rodents, and alcohol craving and alcohol withdrawal in human patients with AUD. Because a ketogenic dietary regimen is difficult to maintain, we have begun to evaluate a ketone supplement drink (KS), which increases blood ketone levels within 30 min of administration without the need for a special dietary regimen. The main goal of this proposal is to examine the efficacy and tolerability of KS in reducing alcohol craving and consumption among individuals with AUD by (1) improving brain energetics and (2) affecting the pharmacokinetic effects of alcohol and subjective responses. Our preliminary data demonstrate that KS reduced alcohol consumption in alcohol-dependent rodents. In humans, a single dose of KS rapidly increased blood and brain BHB levels while decreasing brain glucose metabolism, and pairing KS with alcohol significantly blunted breath and blood alcohol concentrations two-fold and reduced self-reported ratings of alcohol liking and “wanting” more alcohol. Here, we propose a randomly ordered, 2-session crossover study in which 60 nontreatment seeking individuals with AUD will receive a single dose of KS and a matched placebo during two separate study visits. On each visit, participants will undergo a Proton Magnetic Resonance Spectroscopy scan edited for the detection of brain BHB, the consumption of an oral alcohol priming dose, and an alcohol bar lab choice paradigm. Consumption of KS vs placebo is expected to (1) rapidly elevate BHB metabolism, (2) result in lower breath alcohol levels and subjective alcohol liking and wanting, and in (3) lower alcohol craving and consumption. We will test the degree to which the KS-induced shift in brain energetics and -reductions in breath alcohol levels will mediate reductions in alcohol craving and consumption. Demonstrating the beneficial effects of KS on alcohol craving and consumption in human AUD would support the development of new ketone-based interventions that may significantly enhance brain health and success in recovery from AUD.