University Of Pennsylvania

NIH Research Projects · FY 2025 · 2024-06

PROJECT SUMMARY Obstructive sleep apnea (OSA) is the most common sleep disorder and a recognized risk with an estimated worldwide prevalence of one billion people. The ensuing repeated episodes of nocturnal hypoxia and hypoxemia are at the core of the disorder’s pathogenesis, leading to upregulation of neuro inflammation and oxidative stress, which predispose OSA patients to cardiac and neurovascular disease along with impaired cognitive function and neurodegeneration. The investigators have previously examined the neurovascular-metabolic alterations in terms of the cerebral metabolic rate of oxygen (CMRO2) at rest and in response to apneic challenges during wakefulness in the form of repeated cued breath-holds mimicking the hypercapnic-hypoxic events of spontaneous apnea, by means of temporally-resolved MRI-based brain oximetry. Although this work provided new insights into chronic and acute neurometabolic consequences of the disorder, the response to coached volitional apneas likely differs from that of spontaneous apneas during sleep. Also unknown are the upper airway’s morphologic changes that occur during apneas (full airway closure) and hypopneas (partial closure) that cause the metabolic alterations. Leading up to the proposed project we have been able to monitor cerebral oxygen metabolism in healthy subjects in the scanner with concurrent electroencephalography (EEG) and designed an imaging procedure that returns the vascular-metabolic parameters and upper airway morphology during continuous scanning. We illustrate the method’s potential with model apneas induced in test subjects involving the oropharyngeal phase of swallowing, causing airway closure and the expected hypoxic-hypercapnic response and, more recently, in a patient with OSA during 90 minutes of continuous scanning at six seconds temporal resolution during sleep in the scanner. The key hypothesis underlying the proposed research is that the method can evaluate state-dependent O2 brain metabolism and airway anatomy in OSA patients during wakefulness and sleep and during spontaneous apneas and further, that the acute airway structural manifestations during apneas and hypopneas correlate with the metabolic response to apneas. The project comprises three specific aims: (1) Optimize the temporally resolved interleaved structural and metabolic MRI protocol and synchronized airway plethysmography to confirm the method’s ability to simultaneously detect the metabolic and airway structural changes during induced apneas; (2) examine the state dependence of O2 metabolism and upper airway anatomy in OSA patients differing in disease severity, with the method of aim 1 and concurrent EEG monitoring; (3) evaluate the hypothesis that the transient brain metabolic and upper airway changes during apnea can be predicted by baseline measurements during respiration in the awake state along with the subjects’ biological profile obtained from blood markers of oxidative stress and neuro inflammation. The proposed research should provide new insight into the structural and neurometabolic implications of OSA and the disorder’s biological underpinnings, and ultimately guide the development of improved treatment methods.

NIH Research Projects · FY 2025 · 2024-06

ABSTRACT Estimates of suicidal ideation and behaviors (SIBs) amongst adolescents living with HIV (ALWH) in Sub- Saharan Africa (SSA) range from 18-69%. Despite this significant burden, prevention efforts in the region are lagging due to a myriad of structural and socio-cultural barriers. This need is particularly dire in Malawi, where estimates of suicidal ideation amongst ALWH are high, mental health stigma is pervasive, suicide attempts are criminalized, and psychiatric human resources are limited. Resource appropriate interventions that allow for the identification, prevention, and management of SIBs amongst ALWH are urgently needed. The Friendship Bench (FB) is an evidence-based counseling intervention designed to be delivered by trained, lay health workers. The FB has proven highly effective for addressing common mood disorders such as depression, which is major determinant of suicidality. Further, the FB has recently been adapted for ALWH in Malawi. Enhancing FB with evidence-based suicide prevention activities, such as the Safety Planning (SP) Intervention, may be a feasible and effective opportunity to meet the needs of ALWH with SIBs. SP aims to reduce acute SIBs through the co-creation of coping strategies which can be utilized in the event of a suicidal crisis to avert acute suicidal thoughts and manage suicidal urges. As few evidence-based suicide prevention interventions have been implemented in Africa, SP is ideal for Malawi, given it has been delivered by non- psychiatric specialists in the region, and can easily be incorporated into existing services to fully address chronic SIBs. The long-term goal of our research program is to generate and implement an evidence-based model to prevent suicide in Malawi amongst ALWH through increasing the capacity of the health system to identify SIBs and provide evidence-based care as well as through enhancing the capacity of research and policy makers to enact change. Our specific aims are: 1) to enhance the FB model with SP that is adapted to meet the developmental and contextual needs of ALWH in Malawi through formative research; 2) evaluate the feasibility, acceptability, fidelity, and preliminary efficacy of the enhanced FB+SP model through a pilot randomized controlled trial; and 3) enhance capacity of mental health researchers and policy makers to advocate for legislative change through symposiums, workshops and meetings for multisectoral collaboration. The proposed aims pave the way for a R01 application to test the enhanced FB+SP intervention in a cluster randomized controlled trial and represent an important step towards preventing suicide amongst ALWH in SSA.

NIH Research Projects · FY 2026 · 2024-06

Project Summary A canonical feature of migraine is visual discomfort (i.e., “photophobia”), with particular sensitivity to flicker (time-varying modulations of light). We lack a mechanistic understanding of this symptom generally, and specifically require a framework that unites the phenomenon of discomfort with the properties of the visual environment, perception, and neural response. Such a synthesis may be offered by recent work in information- optimal representation. Computationally “efficient” representations represent the statistical structure of the environment and maximize sensory information storage. Recent research in experimental psychology has shown that these “efficient coding” models account for aspects of human sensory judgments and the properties of neural activity. Importantly, these models explain how changes in the statistics of the visual environment to lead to changes in perception. Our project is motivated by the idea that photophobia is an experience of “inefficient” information processing. Over three Aims we will apply the efficient coding framework to understand the properties of flicker exposure, perception, and neural representation in typical observers and in people with migraine and photophobia. Using personal light-logging devices, we will test the idea that people with migraine and photophobia experience a systematically different visual world. Using psychophysical and discomfort measures we will test for the effects of stimulus properties upon flicker perception, and for differences between people with migraine and headache free controls. Finally, we will examine the neural representation of visual flicker using functional MRI to test for the signature of efficient coding in distributed neural responses.

NIH Research Projects · FY 2025 · 2024-06

PROJECT SUMMARY/ABSTRACT Most adults in the United States consume alcohol at some point in their lifetimes. Excessive drinking may elicit detrimental consequences, such as liver cirrhosis, increased cancer risk, motor vehicle accidents, and legal problems, and increase the risk for the development of alcohol use disorder (AUD). Progression to problematic drinking varies depending upon individual differences in responses to alcohol, for which dietary differences may be a contributing factor. Preliminary data from our laboratory suggest a pharmacokinetic interaction of ketones with alcohol, such that ketones paired with alcohol decreased the intoxicating and pleasurable effects of alcohol in human participants. Ketones (-hydroxybutyrate (BHB), acetoacetate, and acetone) are produced endogenously during periods of glucose restriction or administered exogenously with nutritional ketone supplements (KS). In comparison to glucose, ketone metabolism for energy production spares nicotinamide adenine dinucleotide (NAD+) utilization. NAD+ serves as a cofactor for key alcohol metabolism enzymes, namely alcohol dehydrogenase (ADH), and increased NAD+ availability following ketosis may have implications for accelerating alcohol breakdown. Here, we propose to examine the effects of KS on alcohol sensitivity and tolerance in human participants. We propose the following specific aims: Aim 1: to characterize the pharmacokinetic effects of exogenous ketones on alcohol sensitivity, alcohol tolerance, and cognitive functioning (R21 Phase), and Aim 2: to characterize the effect of exogenous ketones and alcohol on brain NAD+ concentrations (R33 phase). We propose to utilize a cross-over designs for the R21 and R33 studies, in which socially drinking participants will undergo alcohol challenge tests on separate study visits following ingestion of KS or a taste-matched placebo. In the R21 phase, we hypothesize that KS vs. placebo will lead to a faster breakdown of alcohol and reduced sensitivity to alcohol. We will compare effects of a low (10g) and high (25g) of KS on a moderate dose of alcohol targeting breath alcohol levels of 0.08% and high dose. We will pilot proton magnetic resonance spectroscopic imaging (1H-MRS) methodology at high-field strength for the quantification of brain NAD+ concentrations. The milestones for the transition to R33 are the detection of significant effects of KS vs placebo on breath and blood alcohol levels, the reliable quantification of the 1H-MRS NAD+ peak at 9.3 ppm, and the tolerability of KS and alcohol in the high-field strength 7T scanner environment. In the R33 phase, we will perform 1H-MRS and functional imaging, and we hypothesize that NAD+ levels will increase with KS vs placebo, and brain NAD+ levels will significantly decrease with alcohol intake. The present study will test the degree to which exogenous ketones affect alcohol sensitivity and tolerance and prevent neurocognitive impairments after drinking alcohol. The study outcomes may identify new approaches for accelerating alcohol clearance and preventing neurocognitive impairments after alcohol intoxication, or novel therapeutics for AUD.

NIH Research Projects · FY 2025 · 2024-06

PROJECT SUMMARY The basis of cellular immunity is the recognition by T cells of foreign peptides in association with major histocompatibility complex (MHC) proteins on the cell surface. CD8 T cells, the subset most associated with cytolysis, are conventionally believed to recognize peptides bound to the classical, hyper-polymorphic MHC class Ia molecules (HLA-A, B, and C in humans); however, a handful of studies have suggested that CD8 T cells can also recognize pathogen-derived peptides in the context of MHC-E (Qa-1 in mice, HLA-E in humans), a nearly invariant non-classical MHC-Ib molecule typically associated with the regulation of natural killer cell function. In limited cases, MHC-E-restricted CD8 T cells have been shown to exert potent antiviral effects, but extremely little is known about how common these responses are in viral infections, what kinds of effector functions they generally exhibit, how they are regulated, and what antigen presentation pathways drive them. This proposal will shed light on this enigmatic aspect of cellular immunity, taking advantage of a recently identified Qa-1-restricted CD8 T cell epitope from influenza A virus called M-SL9. This epitope drives a CD8 T cell response that is co- dominant with conventional MHC-Ia-restricted epitopes and in preliminary studies appears to protect mice from weight loss and clinical severity during flu infection. Aim 1 of this research plan will detail the cellular pathways by which this model Qa-1-restricted epitope is processed, transported, and presented in flu-infected cells. These studies will be facilitated by an M-SL9-specific CD8 T cell hybridoma line that acts as a highly sensitive reporter of M-SL9/Qa-1 presentation. Aim 2 will test the hypothesis that Qa-1-restricted CD8 T cells occur commonly in viral infections and that they protect the host from severe disease, similarly to MHC-Ia-restricted CD8 T cells. An immunopeptidomics approach will be used to discover Qa-1 epitopes in the context of influenza A virus and mouse hepatitis virus 1, a beta-coronavirus in the same family as SARS-CoV-2. An mRNA-based vaccination strategy will be used to raise CD8 T cells against identified Qa-1 epitopes in mice, followed by a challenge with the corresponding virus and monitoring of disease severity. These results will illuminate a promising but poorly understood aspect of the cellular immune response, with potential benefits for the design of new vaccines or immune therapies against infectious diseases and cancer. This proposal will promote the applicant’s career goals of initiating an independent research career with a rapid pace of productivity, laying a foundation of valuable data that will enable several viable follow-up studies, and ultimately gaining better understanding the importance of unconventional forms of antigen presentation in human health. This study will also take place in a broader context that prioritizes the applicant’s career development by emphasizing feedback from senior scientists, mentoring trainees, and engaging with current literature and innovative research on and off campus.

NIH Research Projects · FY 2026 · 2024-06

Traumatic brain injury (TBI) is recognized as a modifiable risk factor for neurodegenerative diseases, including Alzheimer’s disease and Alzheimer’s disease-related dementias (AD/ADRD). As such, there has been an explosion of interest in the link between TBI and the development of late neurodegenerative pathologies, particularly chronic traumatic encephalopathy neuropathologic change (CTE-NC). However, the intense focus on CTE-NC has come at the expense of investigation of the broader spectrum of pathologies found after all forms of TBI. Accordingly, we have adopted the conceptual framework, “TBI-related neurodegeneration” (TReND), of which CTE-NC is just one form. Notably, studies in TBI offer a unique opportunity to discover mechanisms of neurodegeneration, since the initiating event or events – ‘time zero’- are known, thereby permitting the temporal examination of the progression of pathology. Indeed, for over two decades, our group has led the meticulous description of multiple neurodegenerative pathologies in individuals with a history of repetitive mild TBI or single severe TBI. In 2019, we established a NINDS-supported U54 center without walls, CONNECT-TBI, which has become internationally recognized for its success in coordinating prospective tissue banking in TBI, across all injury severities and exposures. To date, CONNECT-TBI has gathered unrivaled clinical datasets and tissue archives, including over 1000 prospectively collected cases across participating centers. Here, as we enter our final year of U54 support for CONNECT-TBI, we propose the next phase initiative; “Transdisciplinary Research Accelerating Neuropathology Studies and Facilitating Open Research Methods in TBI” (TRANSFORM-TBI). This initiative seeks to continue and expand the CONNECT-TBI Archive, which we will leverage to explore relationships between TReND, pathologies of AD/ADRD, and their contribution to late clinical outcomes. We also propose to democratize access to this resource to accelerate the discovery of TReND pathologies and their significance. In so doing, we have established an expert, multidisciplinary team of 26 investigators across 12 sites who will identify associations between the extent and type of neuropathological changes emerging following exposure to TBI of all severities (repetitive mild to single severe) and exposures (including sports, military, intimate partner violence). These neuropathological findings will then be compared with extensive clinical datasets to assess potential clinical correlations. Specifically, we aim to, 1) Explore the pathologic heterogeneity of TReND, including the prevalence and spectrum of AD/ADRD pathologies in late survival from TBI, 2) Characterize TReND-associated glial pathology and contrast it with the glial pathology of wider AD/ADRD, 3) Examine TReND associated clinical phenotypes and their distinction from those of aging and wider, non-TBI related AD/ADRD, and 4) Establish a Digital Slide Archive and Open Research Environment (Digital SCOPE) to support innovative, investigator-led studies across the global research community.

NIH Research Projects · FY 2025 · 2024-06

Project Summary Macrophages are ubiquitous innate immune cells that display tissue-specific function and phenotype. Microenvironmental factors derived from a tissue niche can regulate the development of tissue specific macrophages (TSM). The molecular basis of the specialization of TSM remains poorly understood. Perivascular macrophages (pvMAC) are TSMs that can regulate vascular permeability and immune responses to blood-borne pathogens. Whether and how factors unique to the vascular or perivascular microenvironment regulate pvMACs is currently unknown. This proposal will address these key knowledge gaps. Preliminary work in our laboratory showed that pvMACs in murine fibrosarcoma tumors promote angiogenesis and immunosuppression. We discovered that pvMACs have high intracellular iron and express the GPCR endothelin receptor-B (EdnrB). Macrophage-specific deletion of EdnrB led to reduced vascular density in tumors. I also found that heme induces EdnrB expression in macrophages by inhibiting the transcription factor Bach1. Outside of tumors, I have found that pvMACs express EdnrB in lung and adipose tissue. However, we do not know whether EdnrB expression is a common feature of all pvMACs and whether heme regulates EdnrB expression in these cells. The expression of EdnrB in pvMACs close to the source of the endothelin ligand (endothelial cells) and heme (circulating erythrocytes) leads us to hypothesize that increased heme availability in the perivascular niche drives EdnrB expression in pvMACs through degradation of Bach1 and EdnrB signaling in pvMACs regulates vascular function. In Aim 1, I will assess the iron content and transcriptional profile of pvMACs in adipose tissue through single cell RNA sequencing. In Aim 2, I will identify the function of EdnrB signaling in adipose tissue pvMACs using macrophage-specific EdnrB KO mouse and reductionist ex vivo approaches. In Aim 3, I will determine the role of Bach1 in pvMAC development and function using a Bach1 KO mouse and a model of intravascular hemolysis. Better understanding of the biology of pvMACs is essential given the central importance of the vascular system. This work has far-reaching implications for many pathological conditions, such as hemolysis and ischemia. Endothelin signaling can be targeted by existing small molecule inhibitors. Hence, this work also has the potential to be translated into appropriate clinical contexts.

NSF Awards · FY 2024 · 2024-06

This Research Advanced by Interdisciplinary Science and Engineering (RAISE) award is made in response to Dear Colleague Letter 23-109, as part of the NSF-wide Clean Energy Technology initiative. Globally, Rare Earth Elements (REEs) continue to be among the most important materials for clean energy technologies; in particular Nd, Pr, Dy, and Tb, continue to be critical for the electrified economy. REE recycling from end-of-life devices holds tremendous and growing promise, as magnets within them are rich in highly desired REEs and do not contain undesired REEs. A major unmet challenge is the need for a green, distributed method to efficiently extract and selectively separate desired REE cations from mixtures generated by dissolution of the magnets. The project will study rare-earth element (REE)-Selective Protein-hydrogels to Enable Cation Trapping (REESPECT) for scalable, distributed, continuous REE recovery processes to meet the rapid growth in REE demand to support the green economy. The REESPECT platform is based on hydrogels formed from engineered alpha-synuclein fibrils that contain selective REE binding loops to capture and separate REE from aqueous streams. The hydrogels will be formed into beads used in packed beds to allow high capacity and selective capture of REEs over non-REE cations, as well as discrimination among specific REEs. In addition to the potentially transformative impact on REE recovery and providing educational experiences for doctoral students, the project’s outcomes will be leveraged for STEM outreach and undergraduate educational initiatives. For example, demonstrations for Philly Materials Day and NanoDay will be developed. High school students will work with PhD students in the summer outreach program Penn LENS. Undergraduate researchers will be recruited from Penn and from other universities, particularly the University of Puerto Rico, for summer programs such as NSF REU. Graduate students will volunteer for Penn GEMS, a STEM outreach program for middle school students, among others. The team will develop as an undergraduate analytical chemistry lab component based on this research and will develop projects for the CBE engineering design course offered to students interested in technical solutions to problems of broad societal significance. REESPECTs (Rare Earth Element-Selective Protein-hydrogels to Enable Cation Trapping) are hierarchically assembled hydrogel materials based amyloidogenic proteins in which selective rare earth element (REE) binding loops, termed lanthanide binding tags (LBTs), are presented with exceptionally high density throughout the 3-dimensional hydrogel volume. REESPECT hydrogel beads will be fabricated and fundamentally characterized for binding capacity and transport. REESPECT beads will be exploited in proof-of-concept microfluidics separation processes that can be scaled and are suitable for distributed processing. The REESPECT platform is modular; LBTs for selective separation of particular REE identified by machine learning (ML) approaches will be identified and incorporated in REESPECT for recovery and purification of highly desired REES, including Nd, Pr, Dy, and Tb. This research is highly cross disciplinary and relies on the synergistic interactions at the interface of computational protein design; amyloid protein biophysics and genetic engineering; (bio)inorganic characterization of components and structures for selective cation-binding/triggered release; development of microfluidics/confocal fluorescence microscopy assays, and proof-of-concept separations. The project will address the following four intellectual questions: (1) Is the REEESPECT platform truly modular, allowing incorporation of ML-designed LBTs with tailored REE selectivity (Aim 1)? (2) What are optimal designs for LBT binding structures to confer selectivity and effective separation between REEs, as guided by coordination chemistry and advanced ML methods (Aim 1)? (3) How do the densities of LBT presentation and hydrogel crosslinking alter REE binding along amyloid fibers (Aim 1) and binding capacity and transport under flow within REESPECT beads (Aim 2)? (4) What are optimal conditions for REE recovery and separation under flow of REE-rich streams in the presence of competing non-REE cations (Aim 3)? The REESPECT platform is designed to provide a green, all-aqueous, scalable, distributed, continuous process with the potential to transform REE recovery. 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.

NSF Awards · FY 2024 · 2024-06

It is difficult to control gene expression in mammalian cells. Accomplishing this is key to developing effective cell therapies. Small molecules can exert that control. Unfortunately, small molecules are often toxic, and tend to have a limited, on-off capability. The aim of this project is to develop an improved small molecule system for mammalian cells. This system will be tunable instead of on/off. Highly sensitive, it will interact with multiple genes to direct more complex behaviors. Undergraduates from underrepresented groups and high school teachers will learn techniques of synthetic biology and CRISPR technology. Proteolysis Targeting Chimeric (PROTAC)-Chemically Induced Dimerization (CID) systems will be developed as sensitive, multiplex, tunable, and safe toolkits to control gene expression in mammalian cells. Combining PROTAC-CID with genome editors will enable precise genomic modification. This will greatly increase the accuracy and safety of using genome editors for basic and therapeutic applications. The specific research goals are to 1) establish a novel PROTAC-CID inducible gene expression system; 2) engineer multiplex inducible gene regulations; 3) design high-induction and low-basal level PROTAC-CID systems for Cre recombinase expression, and 4) develop PROTAC-CID inducible CRISPR base editors. Successful completion should result in novel PROTAC-CID inducible gene expression platforms for multiplex gene regulations in living eukaryotic cells. These systems will provide deep insights into the approaches to achieve high-induction and low basal-level gene expressions. Finally, applying PROTAC-CID systems to CRISPR base editors will permit inducible genomic DNA modification at a defined time to reduce the off-target effect and increase the accuracy and safety in genome editing. 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 · 2024-06

Project Summary In this R21, we propose a detailed study of Covid-19’s impact on the mortality of single-year birth cohorts of males and females in France and the United States, with additional analyses by race for the United States. The findings from this R21 will complement existing studies of the mortality impact of Covid-19 that have almost exclusively adopted a period approach using measures such as period life expectancy. Our proposed analyses will combine important developments in the formal demography of cohort survivorship by Guillot and colleagues with a causal identification strategy employing a novel cohort discontinuity design developed by Wu, Mark, and colleagues to identify the causal effect of the Great Recession on U.S. fertility. Our analyses will provide credible causal estimates of: (1) the effect of Covid-19 on all-cause mortality for single-year birth cohorts of males and females in France and the United States, and for Black and White Americans in the United States; and (2) the degree to which the Covid-19 mortality shock has offset or reversed pre-pandemic progress in cohort survival. Our analyses will rely throughout on secondary data from French and U.S. vital registers that provide detailed, high quality data on all-cause mortality for single-year birth cohorts of males and females. Findings will break new ground by providing credible causal evidence relevant to ongoing mortality research and debates on racial disparities, health systems, pension schemes, and population dynamics.

NIH Research Projects · FY 2026 · 2024-05

PROJECT SUMMARY/ABSTRACT The overall goal of the present application is to define dysbiosis and identify microbial changes that are predictive of an ecological disruption that ultimately leads to periodontal destruction and to provide mechanistic insights into why this occurs. The major strength of our approach is our existing biobank derived from U01- DE021127 “Biomarkers of Periodontal Disease Progression (BPDP) [R. Teles (PI), F. Teles (Co-I) et al.]. Periodontitis is the most common cause of tooth loss among US adults. Further, it increases the risk for systemic conditions. Proper patient management is critical to minimize tooth loss, allow resource allocation and limit the potential systemic sequelae of these infections. By exploring our biobank, we have a unique opportunity to capitalize on existing longitudinal clinical data as well as subgingival plaque and gingival crevicular fluid samples from periodontitis patients who experienced disease progression over a 12-month period of monitoring, in the absence of treatment. This approach will reduce costs of conducting prospective, longitudinal trials. Results from our study will provide mechanistic insight into periodontal destruction based on human clinical data. Such data does not currently exist. Aim 1 will determine longitudinal dysbiotic microbial changes that culminate in periodontitis progression versus microbial signatures that reflect periodontal stability. Aim 2 will utilize computational approaches and predictive models to gain mechanistic insight into clinically relevant dysbiotic changes. Strengths of this proposal include: 1) readily available, curated and validated samples coupled with periodontal data from hundreds of periodontitis patients; 2) utilization of pairs of samples with and without periodontitis progression, collected from the same subject, 3) Analysis of microbial, immunological and metabolomic signatures from the same periodontal site, 4) a multidisciplinary study team that includes a PI involved in the original study (U01-DE021127); and experts in microbiology, immunology, host microbial interactions and computational biology tools, 5) utilization of state-of-the-art platforms, including whole genomic sequencing, metabolomics and immunoassasys; 6) application of bioinformatics tools, multi-omics integration and predictive models that have been established in other fields to provide mechanistic insight in bacterial and host changes that lead to dysbiosis. This project supports the NIDCR’s commitment to facilitate the translation of science into clinical practice. Our contribution to the field will be the generation of models for mechanistic validation that will affect strategies for the prevention and treatment of periodontitis based on an understanding of dysbiosis in the periodontium that is lacking.

NIH Research Projects · FY 2026 · 2024-05

Principal Investigator: Robertson, Erle S. Program summary Approximately 20% of all human cancers are associated with viruses that function as biological cofactors driving these cancers. Some of these viruses may have a direct role in mediating these cancers as in the case of HIV-related cancers, which includes Kaposi’s sarcoma, pleural effusion lymphomas and lymphoproliferative disease. There is also an increase in the number of HPV- related patients for example in the immunocompromised patients who are on HAART therapy and in head and neck squamous cell carcinomas. The Tumor Virology Training Program at the University of Pennsylvania serves as the central forum for facilitating interactions among investigators involved in cancer-related viral research on the Penn campus which provides a more directed training for trainees. Program members have expertise in EBV, KSHV, HPV, HCV, HIV and other retroviruses. The biomedical community at Penn would like to continue the momentum of this training program in Tumor Virology for training predoctoral and postdoctoral students. There are 21 trainers in this program all of whom are committed to training pre and postdoctoral fellows for biomedical research careers. The faculty have well-funded NIH programs supported by different NIH funding mechanisms, and other Government and private foundation funding. The training program seeks to continue support of 2 predoctoral and 4 postdoctoral students, annually, for each of the 5 years of funding. The number of trainees in labs of the trainers of this program has been consistently increasing and we anticipate this trend continuing in the coming years. Thus, we would like to have available slots for the continued increase in the number of predoctoral and postdoctoral candidates available to the program. Viral related cancers are expected to increase as the technology for identifying these agents improves. We continue to provide an atmosphere of collaboration between clinical and basic scientists for our trainees who will have the opportunity to formulate ideas which will lead to basic and translational studies initiating and maintaining a cohesive group in tumor virology. The increased incidence of viral associated cancers, the commitment of the trainers and the institution with a well-organized training program will continue to provide an outstanding training environment for pre- and postdoctoral candidates in tumor virology.

NIH Research Projects · FY 2026 · 2024-05

Project Summary Over one million people live in U.S. nursing homes, two-thirds of whom have Alzheimer’s disease and related dementias (AD/ADRD) and one-quarter of whom are dually eligible for Medicaid and Medicare. Dual-eligible residents with AD/ADRD often receive low-quality nursing home care at high costs, due in part to the conflicting financial incentives created by Medicaid and Medicare. Nursing homes rely on generous Medicare payments for short skilled nursing home stays. At the same time, nursing homes care predominantly for Medicaid-funded long-stay residents, stays for which Medicaid payment often does not cover the costs. This creates conflicting incentives for nursing homes and fragmented care for dual-eligible residents. This includes high rates of potentially avoidable hospitalizations as nursing homes shift the high costs of caring for a sicker resident to a Medicare-paid hospitalization. It also causes nursing home residents to cycle between the nursing home and hospital, increasing the use of Medicare-covered skilled nursing home stays on return to the nursing home, which may be unnecessary and costly to Medicare. One potential solution to these conflicting incentives is for states to increase Medicaid per-diem rates, which would decrease the Medicaid-Medicare payment differential. This would better align incentives for nursing homes to provide higher-acuity care for residents rather than transferring them to the hospital. It would also eliminate the incentive to provide Medicare-funded skilled care due only to the higher profitability of these stays, rather than the potential clinical benefit to residents. Another potential solution is to use globally capitated payments for nursing home stays, which hold nursing homes financially accountable for spending. A prominent example of this is Medicare-established institutional special needs plans (I-SNPs) for dual-eligible nursing home residents. I-SNPs are specialized Medicare Advantage plans that bear risk for all Medicare-covered spending for nursing home residents and as a result, may incentivize nursing homes to invest in capabilities to better manage high-acuity residents in the nursing home rather than in the hospital, allowing nursing homes to higher-intensity skilled care without a preceding hospitalization. Despite the promise of these approaches and urgent need to decrease fragmentation, little is known about how to reform nursing home payment to accomplish this. This is particularly important for individuals with AD/ADRD given their large numbers in nursing homes, high likelihood of being dually enrolled in Medicare and Medicaid, the high costs of care, and poor outcomes from unnecessary and burdensome transitions of care. Our overall objective is to examine ways to align the historically misaligned incentives created by the Medicare and Medicaid programs in nursing homes and, in doing so, substantively improve care for residents with AD/ADRD. These results will provide critical and rigorous evidence on how to integrate Medicaid- and Medicare-funded care for dual-eligible nursing home residents with AD/ADRD and improve outcomes. These results will inform current policy debates and future directions of ongoing integration efforts.

NIH Research Projects · FY 2026 · 2024-05

Project Summary Human hippocampus is among the most complex tissue types with a wide range of cells organized in spatially precisely defined regions. Alzheimer’s disease (AD) is one of the diseases strongly affected the hippocampal region, which underlies the core feature of the disease-memory impairment and remains refractory to therapy, due in part to the complexity of gene regulatory network coupled with cell-type-specific mechanisms of AD progression. What types or subtypes of cells are affected by this process and their spatial heterogeneity in the tissue context as well as how these cells impact the tissue environments remain poorly understood, which precludes the development of strategies to target these cells to improve healthspan/lifespan or harnessing these cells to promote tissue remodeling and repair, highlighting a pressing need for tools to map cells and the surround microenvironments in these tissues and generate biomarkers to define spatial and phenotypic heterogeneity in AD. This grant aims to (1) develop a first-of-its-kind technology for spatial co-profiling of epigenome, transcriptome, and a panel of proteins in the same tissue section, and (2) deploy the high-resolution, high-content and high-throughput spatial multi-omics technologies to construct comprehensive maps of cellular states and the associated environments in human hippocampus from AD patients and health donors. The expected outcomes and the major contributions of our project include: (1) Fundamental knowledge on diverse cell types and their molecular basis of selective vulnerability (and conversely the resilience of other cell types) in the context of tissue organization in the AD human hippocampus, allowing us to develop a high-spatial-resolution cell census which will in turn allow us to identify molecular signature patterns, gene regulatory networks, and biological processes potentially mediating cell type specific differences in the AD, and (2) Offer the possibility of testing new therapeutic approaches for AD that are not targeted by currently approved treatments. The resulting data will lead to better understanding of the relationship between tissue organization, function, and gene regulatory networks in AD.

NIH Research Projects · FY 2025 · 2024-05

Project Summary/Abstract Clostridioides difficile is the leading cause of hospital-acquired gastrointestinal infections in the United States. Pathogenesis is mediated by secreted toxins that cause the death of intestinal epithelial cells (IECs) and breakdown of the intestinal barrier. C. difficile infection is primarily treated with antibiotics; however, the high rate of recurrent infections necessitates alternative treatment options. Our lab has found that orally administering a toll-like receptor 7 (TLR7) agonist, R848, protects mice against severe disease following lethal C. difficile challenge. R848 treatment induces the expression of inflammatory cytokines including interferons. Interestingly, while protection was not dependent on type I or type II interferon signaling, we found that mice deficient for type III interferon (IFN-λ) signaling are not protected following R848 treatment. While IFN-λ is best known for its role in antiviral immunity, recent works by others has shown that IFN-λ stimulates intestinal stem cell proliferation to reduce the severity of experimental colitis. This proposal seeks to determine the mechanism by which IFN-λ contributes to R848-mediated protection against C. difficile infection. The overarching hypothesis for this project is that IFN-λ directly acts on IECs to stimulate stem cell proliferation and thereby repairing the damage to the intestinal barrier from C. difficile toxin. Aim 1 will identify the critical cell type that responds to IFN-λ for R848-mediated protection against C. difficile infection using bone marrow chimeric mice and cell- lineage specific knockout mice. Aim 2 will test the hypothesis that IFN-λ promotes intestinal stem cell function and stimulates proliferation by utilizing Lgr5EGFP reporter mice and intestinal organoid culture models. This project will elucidate the mechanism behind a novel role for IFN-λ in the immune response against C. difficile and may inform future development of therapeutics against this major public health threat.

NIH Research Projects · FY 2026 · 2024-05

FOXL1 positive telocytes in intestinal development and homeostasis Abstract The intestinal stem cell niche plays an essential role in enabling the self-renewal of the columnar gut epithelium, a process that occurs continuously throughout life. Over the past five years we have established that subepithelial fibroblasts resembling telocytes that express the DNA-binding transcription factor Foxl1 are a critical component of the intestinal stem cell niche. Removal of Wnt signals emanating from Foxl1+ cells or elimination of these specialized cells themselves causes cessation of epithelial proliferation, crypt failure, and rapid death of mutant mice. In addition, Foxl1+ cells demarcate critical signaling centers during intestinal development, and Foxl1 itself is required for villus development in fetal life. Based on these exciting findings, we have built several crucial tools to further explore the function of Foxl1+ telocytes and fetal telocyte precursors in three independent but interrelated specific aims. In specific aim 1, we will ablate critical components of the planar cell polarity pathway in Foxl1+ telocyte precursors to determine their contribution to villus formation during the epithelial transition in midgestation, based on our findings from single cell RNAseq analysis that these factors are highly enriched in the Foxl1+ telocyte progenitor populations during villus formation, and the fact that villification is abnormal in Foxl1 null embryos. In specific aim 2, we will test the hypothesis that the closely linked Foxl1 and Foxf1 genes function cooperatively in telocytes or their progenitors to control both gut development and the function of the adult stem cell niche. We will employ an innovative ‘double floxed’ mutant allele to ablate both genes simultaneously in the Foxl1+ lineage and analyze fetal gut development and adult tissue homeostasis using histology, immuno-staining, and molecular assays. Finally, based on our discovery that GLP-2R, the receptor for the intestinal mitogen GLP- 2, and its downstream signal IGF1 are highly enriched in Foxl1+ telocytes, we will test the hypothesis that telocytes are the GLP-2 target cell using inducible conditional gene ablation of both Glp2r and Igf1 and evaluate GLP-2 action in multiple paradigms. Together, these linked yet independent aims will dramatically increase our understanding of intestinal telocyte biology, a cell type we have shown to be a critical component of the intestinal stem cell niche.

NIH Research Projects · FY 2026 · 2024-05

PROJECT SUMMARY Innervation plays a key role in muscle development, function, and regeneration. Therefore, tissue engineering strategies to develop biomimetic skeletal muscle to study developmental mechanisms as well as for regenerative medicine applications should consider the implications of neural inputs during the biofabrication process. Moreover, tissue regeneration following severe musculoskeletal injuries like volumetric muscle loss (VML) is hindered by a lack of appropriate motor innervations, which diminishes regenerative capacity and often results in minimal functional recovery. Hence, there is an unmet clinical need for an intervention strategy that augments reinnervation and promotes a pro-regenerative environment following severe musculoskeletal trauma. Over the last several years, our group has shown that pre-innervated tissue engineered muscle scaffolds can effectively enhance muscle regeneration, revascularization, and functional recovery following VML. Building on these findings, the goal of the current program is to advance the first three-dimensional tissue engineered motor units (3D-TEMUs) comprising dense bundles of centimeter-scale myofiber fascicles innervated by preformed axonal networks projecting from discrete pools of motor and/or sensory neurons. The 3D-TEMUs will be validated as an in vitro testbed to study the role of innervation in facilitating muscle development as well as a composite soft tissue to augment functional regeneration following implantation in a rodent model of VML. Notably, 3D-TEMUs will be generated using human induced pluripotent stem cell (iPSC) derived myocytes and motor/sensory neurons to perform in-depth characterization using a clinically-relevant and potentially translatable biomass. These efforts will be carried out across 3 Specific Aims. First, human 3D-TEMUs will be optimized to recapitulate the native myofascicular architecture of skeletal muscle tissue in order to provide a biofidelic testbed to better understand neuromuscular development and sensorimotor function in the context of myofiber formation and maturation as well as to refine tissue engineering strategies for augmenting muscle regeneration (Aim 1). Next, 3D-TEMU neurons will be transduced to contain optogenetically controlled (i.e., light-activated) motor inputs that allow for spatiotemporal control of muscle-fiber contraction to demonstrate refinement of motor units over time and to measure sensorimotor function (Aim 2). Then, 3D-TEMUs will be implanted in a rat model of VML to optimize acute graft cell survival (Aim 3A) followed by chronic studies assessing the ability of 3D-TEMUs in facilitating bulk muscle replacement, reinnervation, revascularization, and overall functional restoration (Aim 3B). Successful execution of these studies will significantly advance the development of 3D-TEMUs as both an in vitro testbed to study mechanisms of neuromuscular development as well as an implantable composite soft tissue to enable functional regeneration following currently intractable VML. With further advancement and clinical translation, 3D-TEMUs could potentially change the surgical paradigm for muscle repair and fundamentally alter clinical expectations for functional recovery following major musculoskeletal trauma.

NIH Research Projects · FY 2026 · 2024-05

PROJECT SUMMARY The 1.2 million people in the United States affected by hematologic malignancy (HM) each year are at high risk of infection-related complications and mortality. As a result, HM patients are exposed to prolonged antibiotic therapy, and are at increased risk of harm from multidrug resistant organisms and antibiotic- associated adverse events. Optimizing selection of antibiotics in this population is a critical antibiotic stewardship (AS) goal. A major barrier to achieving this goal is inaccurate antibiotic allergy labels. Although self-reported beta-lactam (BL) allergy is found in up to 20% of all inpatients, 90% of patients with a reported penicillin (PCN) allergy are able to tolerate PCN. Inaccurate allergy labels may result in patients receiving sub-optimal antibiotic therapy with less effective, overly broad, and/or more toxic agents. In HM patients, BLs are preferred for many infectious complications. BL allergy is common in HM patients (>25%) and those with BL allergy have significantly greater hospital LOS, 30-day mortality, and antibiotic exposure. Studies of antibiotic allergy delabeling interventions combining history, skin testing and oral challenge in general hospitalized patients have shown that up to 85-95% of patients with reported BL allergy can be safely delabeled. Patients who are successfully delabeled have increased appropriate antibiotic use and improved clinical outcomes. Despite their increased risk of antibiotic-associated harm, cancer patients with BL allergy have not been prioritized in research on delabeling interventions. Although recent studies (including our own) provide promising early evidence for the feasibility of BL delabeling in cancer populations, the impact of a BL allergy delabeling intervention on clinical outcomes in the HM population is unstudied. Our long-term goal is to optimize antibiotic use and improve clinical outcomes in high-risk immunocompromised patients. The overall objective in this application is to test the impact of a pharmacist-led BL allergy delabeling intervention (Allergy Delabeling in Antibiotic Stewardship – “RENEW”) on clinical outcomes and antibiotic use in hospitalized patients with HM while concurrently assessing social and behavioral factors that shape implementation. The Specific Aims of this study are: Aim 1: Assess the Impact of the RENEW Intervention: To assess the impact of a BL allergy delabeling intervention on antibiotic use and clinical outcomes in HM patients using a cohort study design. Aim 2: Identify Barriers and Facilitators to Implementation of the RENEW Intervention: To identify barriers and facilitators to the implementation of the RENEW intervention via a concurrent mixed-methods process evaluation that will elicit the perceptions of patients and clinicians toward the intervention. This study will generate robust evidence to support broad dissemination of our findings to the HM population, inform subsequent interventions targeting high-risk immunocompromised patients, and optimize implementation of allergy delabeling in all hospitalized patients.

NIH Research Projects · FY 2026 · 2024-05

Intrahepatic cholangiocarcinoma (iCCA) is the second most common primary liver malignancy in the United States and worldwide. iCCA has a dismal prognosis, with just a 20% 5-year survival rate, and the incidence of this condition has been increasing over the last few decades. Many factors contribute to the poor prognosis of iCCA, including issues with current diagnostic capabilities and limited treatment options. There has been significant progress in the last decade in understanding the molecular mechanisms underlying iCCA, including identifying isocitrate dehydrogenase (IDH) gain of function as one of the most common mutations specific to iCCA, with R132 being the most common. This mutation results in alpha-ketoglutarate (α-KG) being metabolized to 2-Hydroxyglutarate (2HG), an oncometabolite and a surrogate marker of the mutation. This mutation provides the potential for the development of precision medicine strategies for IDH mutant iCCA. Many new therapeutics for IDH mutant iCCA using small molecule inhibitors of mutant IDH have been developed, with the ClarIDHy trial showing promising phase III results. However, despite these advances, accurate diagnosis of iCCA, particularly diagnosing subtypes, remains challenging, slowing the implementation of appropriate treatment strategies. Current noninvasive diagnostic approaches for iCCA, such as Contrast-Enhanced Ultrasound (CEUS), Magnetic Resonance Imaging (CE-MRI), and Computed Tomography (CT), have varied reports of sensitivity18–20. Importantly, these techniques lack high specificity in the 30% of patients who present with cirrhosis, risking a misdiagnosis of hepatocellular carcinoma (HCC). Furthermore, these imaging techniques cannot determine the specific subtype of iCCA without invasive procedures. Biopsies, the gold standard for confirming iCCA diagnosis, have a long turnaround time and may be unsuitable for molecular analysis in up to 20% of cases. Due to these limitations, there is a clinical need for minimally invasive and fast imaging tools capable of detecting iCCA with IDH mutation and differentiating iCCA from HCC. To address this need, we propose the use of hyperpolarized MRI as a novel diagnostic approach for identifying mIDH-producing iCCA. By leveraging the enhanced sensitivity and spectral characteristics of hyperpolarized carbon-13 (13C) labeled α-KG and its metabolites, we aim to study the dynamics of mIDH's enzymatic behavior in vitro and in vivo. We will optimize the imaging parameters for this technique and build towards accurate noninvasive identification of IDH-mutant iCCA. Aim 1: Optimize an existing pulse sequence to maximize sensitivity to 2HG through spectral editing. Aim 2: Characterize the dynamics and sensitivity of hyperpolarized 1-13C diethyl Alpha-ketoglutarate metabolism in mIDH intrahepatic Cholangiocarcinoma (iCCA) cells. Aim 3: Noninvasively identify IDH-mutated Cholangiocarcinoma using Hyperpolarized diethyl 1-13C Alpha-ketoglutarate in patient-derived-xenograft mouse models.

NIH Research Projects · FY 2026 · 2024-05

Project Summary: As obligate intracellular pathogens, viruses must hijack cellular machinery to facilitate productive infection. For DNA viruses that depend on host RNA processing machinery to produce viral transcripts, common targets of manipulation include both cellular RNA-binding proteins (RBPs) and the enzymes mediating the post- translational modifications (PTMs) that govern their functions. Arginine methylation is a PTM deposited by a family of protein arginine methyltransferases (PRMTs) and involved in multiple aspects of RNA processing. While the roles of arginine methylation and PRMTs constitute an emerging field in multiple areas of biology, relatively little is known about their functions during viral infection. The objective of this project is to utilize human Adenovirus (AdV) as a model system to address the roles of arginine methylation during infection. AdV is an important human pathogen and also well recognized as a tool for investigating fundamental cellular processes. Preliminary data from the Weitzman lab demonstrate an intriguing global decrease of arginine methylation on cellular RBPs throughout AdV infection. Concurrently, arginine methylation of late region 4 (L4) 100 kDa nonstructural protein (100K) dramatically increases as infection progresses. Furthermore, 100K expression is sufficient to cause relocalization of PRMT1 from its normally nuclear subcellular compartment to the cytoplasm, an event which correlates with a 100K-dependent loss of arginine methylation on cellular RBPs. Additionally, methylation of the host RBP hnRNPA1 decreases in response to 100K expression alone. Arginine methylation of hnRNPA1 is known to regulate its splicing capacity, and AdV is well known to manipulate host splicing machinery, but knowledge of the role of hnRNPA1 during AdV infection is limited. These collective findings inform my hypothesis that 100K acts as a molecular sponge of PRMT1 activity, leading to the loss of arginine methylation of RBPs such as hnRNPA1, and thus regulating this splicing factor’s RNA-binding capacity and function to promote efficient AdV splicing. In Aim 1 I will determine the requirement of 100K for PRMT relocalization (confocal microscopy, IP-WB) and decreased hnRNPA1 methylation (isothermal calorimetry, MS- based competition experiments). In Aim 2 I will determine if 100K impacts hnRNPA1 RNA-binding (eCLIP, RNA Binding-Region Identification) and splicing abilities (qPCR-based splicing assays, molecular cloning, WB, plaque assays) to benefit viral infection. This study will be the first to describe a mechanism of viral manipulation of arginine methylation to promote infection. Results of this proposal will expand our understanding of how pathogens interfere with PTM machinery, thus informing future studies to develop appropriate therapeutics targeting arginine methylation to treat viral infections. This research will take place in the collaborative and interdisciplinary environment of the Weitzman lab and the integrated communities of both the University of Pennsylvania and the Children’s Hospital of Philadelphia. Skills gained from this training fellowship will prepare me for a career as a principal investigator investigating molecular processes governing virus-host interactions.

NIH Research Projects · FY 2025 · 2024-05

Project Summary / Abstract Collagen lays the foundation of bodily tissues, serving to strengthen, connect, and signal from the micro to the macro scale. The importance of collagen in cancer biology has been well-established: its functions in regulation of the tumor microenvironment—from increased stiffness of the extracellular matrix to dysregulation of cancer cell signaling—influence tumor proliferation and impenetrability. Key to these effects is the interaction between fibrillar collagens and the discoidin domain receptor type 2 (DDR2), a receptor tyrosine kinase implicated in multiple human cancers. Extracellular binding of fibrillar collagen to DDR2 transduces a cell signal that activates epithelial to mesenchymal transition, proliferation, and metastasis. This work seeks to develop synthetic collagen mimetic peptides (CMPs) for interrogation of the collagen-DDR2 interaction. As a permutated, triple helical polymer involved in biochemical signaling, collagen has potential for manipulation as a tool for modulating protein-protein interactions. These applications have been limited by its tripartite nature, which restricts its thermal and entropic stability. Synthetic linkage and cyclization of the three collagen strands may overcome these limitations. To this end, this work focuses on the design, synthesis, biophysical/structural characterization, and biological application of linked and cyclic CMPs targeted against DDR2 through a chemical biology approach described within two Aims. Aim 1 will evaluate the impact of strategic design strategies on the thermal and proteolytic stability of the proposed CMPs, with a goal of maximizing stability. Aim 2 will investigate the ability of the CMPs to interact with DDR2 in vitro and modulate DDR2 signaling in cellulo. Prior work has established a method for synthesis of macrocyclic CMPs by solid phase peptide synthesis with on-resin cyclization, and demonstrated that photoreaction of CMPs using diazirine-based photocrosslinking reagents can be used to prepare unimolecular collagen heterotrimers. Several linked/cyclic as well as linear DDR2-targeted CMPs have already been synthesized and have demonstrated appropriate thermal stability as well as capacity to interact with DDR2. Methods include organic synthesis, photoreaction, circular dichroism, collagenase assay, isothermal calorimetry, X-ray crystallography, and mammalian cell work coupled with Western blot. This work endeavors to develop biochemical tools for modulation of the collagen interactome—concentrating on the cancer-implicated interaction between collagen and DDR2—via an innovative chemical biology approach, laying the foundation for new discoveries surrounding the role of collagen in cancer biology with potential for applications in drug discovery. The robust, collaborative training environment at the University of Pennsylvania’s Department of Chemistry and Perelman School of Medicine’s Medical Scientist Training Program—as well as a skilled sponsorship team with expertise in organic synthesis, collagen structure and function, receptor tyrosine kinases, crystallography, and cell biology—will support completion of the proposed fellowship work and facilitate the rigorous training of an independent physician-scientist investigator.

NIH Research Projects · FY 2025 · 2024-05

Project Summary/Abstract CD8 T cells are critical to the adaptive immune response to viral pathogens. When neutralizing antibodies are insufficient to prevent viral infection outright, virus-specific memory CD8 T cells can rapidly kill infected cells and limit the severity of infection. This has recently been identified as a potentially key mechanism by which COVID- 19 disease severity is reduced in vaccinated individuals. Memory CD8 T cells are functionally heterogeneous within and between individuals, with some cells more capable of effectively responding during viral infection. In the worst cases, dysfunctional CD8 T cell responses or ongoing responses to chronic viral infections could cause persistent and debilitating symptoms, as may be the case in Long COVID. Our understanding of the dynamics of human CD8 T cell differentiation remains incomplete, including why individuals generate functionally different responses to an identical antigen. Developing effective vaccines will require an understanding of shared molecular programs that can lead to potent effector responses across a range of individual immune landscapes and CD8 T cell identities. Two key gaps in knowledge impair our ability to fully leverage the potential of T cells in vaccines. First, little is known about the lineages that produce functionally distinct T cells in humans and what factors promote different lineages. Second, it is unclear which CD8 T cell states retain the greatest capacity to mount a potent effector response to combat infection. Therefore, there is an urgent need to define human memory CD8 T cell differentiation trajectories, how they impact future responses to viral pathogens, and how they may be dysregulated in cases where symptoms fail to resolve, such as Long COVID. We will address this need by testing three working hypotheses: first, that T cell clones differentiate down lineages with distinct functional capacities; second, that epigenetic features leave some lineages poised to mount more potent responses; and third, that the immune landscape influences CD8 T cell differentiation and functionality, with multiple classes of immune dysregulation driving Long COVID pathology. In Aims 1-2, we will use custom HLA- I/peptide tetramers to sort rare Spike-specific CD8 T cells from 5 individuals at each of a longitudinal series of memory and acute responding time points. We will then perform two cutting-edge single-cell sequencing approaches that use T cell receptor and mitochondrial variant sequences to define T cell clones and track the differentiation of these clones over time while assessing cell states by mRNA expression, protein expression, and chromatin accessibility. Aim 3 will use a systems immunology approach in healthy vaccinees and Long COVID patients to identify features of the immune landscape that correlate with optimal, sub-optimal, and dysregulated CD8 T cell responses. Collectively, these studies will evaluate multiple possible causes of Long COVID and will define the CD8 T cell differentiation lineages that produce optimal responses, providing a blueprint for improved vaccines to reduce severe disease caused by SARS-CoV-2, Influenza, and other viruses.

NIH Research Projects · FY 2026 · 2024-05

Rational drug design and understanding of how mutations cause disease are largely based on the hypothesis that a protein's sequence determines its structure, which determines its function. However, designing drugs and interpreting new variants remain difficult, suggesting there is a missing factor in this sequence-structure-function paradigm. There is good reason to believe that a sequence-ensemble-function paradigm that better accounts for the fact that proteins are not rigid bodies, but are dynamic entities that are endlessly hopping through a set of different structures (called an ensemble) would be far more powerful. However, realizing this potential has been slow because it is even harder to get an atomically-detailed picture of an entire ensemble than a single protein structure. The PI and his lab have been developing tools that combine atomically-detailed computer simulations, biophysical experiments, and machine learning to overcome this challenge. They have made significant progress on relatively small proteins with limited dynamics, enabling a deeper understanding of how mutations modulate function and the design of new drug-like molecules for controlling function. The objective of this work is to test the applicability of these tools to much larger and more complicated proteins that are of significant importance in both fundamental biology and drug design, myosin motors. Myosins are responsible for a broad range of biological functions, from muscle contraction to hearing. As a result, they are important targets for treating diseases ranging from heart failure to parasitic infections. To function, myosins must undergo a complex series of structural changes. The PI and his lab will test whether their tools for accounting for these extensive dynamics enable more accurate predictions of sequence-function relationships and the rational design of new drug-like molecules for controlling motor function. They will focus on β-cardiac myosin because of its importance in heart disease and the myosin 1 family of motors because its members have highly diverse biological functions but it remains unclear how mutations tune the motor's behavior for all these different purposes. The lab will design new myosin variants and allosteric modulators as a stringent test of their insights. Success will enable future myosin drug discovery and application of these tools to other proteins.

NIH Research Projects · FY 2026 · 2024-05

PROJECT SUMMARY Humans rely on sweating to remove excess heat from the body, making the eccrine sweat gland an indispensable component of the human thermoregulatory system and an organ essential for human survival. During development, epidermal progenitors within the eccrine placode differentiate into multiple specialized cell types that are ultimately partitioned into specialized, functional compartments. A lack of understanding of how the progressive differentiation of the eccrine anlage occurs has stalled efforts to regenerate these critical skin appendages for the repair of traumatic skin injuries, leaving affected patients with potentially life-threatening deficits in thermoregulation. We have recently identified a novel eccrine dermal population, the Engrailed 1- dependent eccrine niche (EDEN), that is required for eccrine gland formation in mice, and gives rise to a distinct dermal lineage that is persistently associated with each developing human and mouse eccrine gland. A first-of- its kind dermal niche to be described for eccrine glands, understanding how EDEN arises and is maintained, and what factors EDEN produces to instruct eccrine development are entirely unknown. Accordingly, the goals of this proposal are to define the cellular mediators required for EDEN induction, differentiation, and maintenance (Aim 1) and to identify EDEN-produced effectors required for eccrine differentiation (Aim 2). Our research strategy employs: a) functional experiments in mice using existing alleles to carry out targeted genetic perturbations of EDEN and the associated eccrine epidermal anlage and also b) candidate-based and systematic discovery of EDEN molecular effectors required for eccrine differentiation using genetically-targeted, single cell transcriptomics followed by high-throughput seqFISH+ based validation in both human and mouse eccrine- forming skin, and functional testing in vivo using genetic disruption in mice. Our findings will define the regulators, functional properties, and effectors of the eccrine niche to reveal the extrinsic drivers of eccrine developmental progression. Made possible by the unique biological and technical expertise of our team, this first-of-its kind, instructive recipe for eccrine formation will seed translational efforts to regenerate these essential appendages and identify novel genetic targets for alleviating eccrine disorders.

NIH Research Projects · FY 2025 · 2024-04

Research Training in Integration of Epidemiology and Implementation Science for Neglected Zoonotic Disease Control in Peru PROJECT SUMMARY The mission of this D43 application is to develop and establish an innovative transdisciplinary training program in implementation science and epidemiology for Neglected Zoonotic Disease (NZD) control in Peru. NZDs affect mainly the poor and have devastating effects on the affected individuals and their families and communities. Due to migration and travel, high-income countries and their health systems are also affected by NZDs. Despite NZDs impact and importance, there are no formal training programs to allow professionals to focus on these diseases in Peru. This D43 application is a collaboration between Universidad Peruana Cayetano Heredia, the leading biomedical institution in Peru; the Perelman School of Medicine at the University of Pennsylvania, a world-renowned academic health institution; and the Tulane School of Public Health and Tropical Medicine, a global powerhouse for tropical infectious disease training, with additional input from experts from other institutions in the US and overseas. It proposes to prioritize training for young scientists from disadvantaged areas in Peru, leveraging existing related infectious disease and implementation science programs, while capitalizing on the extensive network and reach of collaborating institutions and research hubs across various regions. This network includes a stellar group of faculty with diverse research projects. The Principal Investigators’ distinctive set of skills: a clinical epidemiologist, a veterinary epidemiologist, and an implementation scientist, make them a perfect “One Health” team to train scientists to tackle zoonotic diseases in Peru and Latin America. Throughout the 5-year grant period we propose to: a) train 8 Peruvian students per year (40 total) in the Diploma of Operational Research and Implementation Science; b) train 4 Peruvian students per year (20 total) in the Master’s in Control of Infectious and Tropical Diseases; c) train 2 Peruvian scientists between years 2-5 (8 total) in the Penn Summer Implementation Science Institute; and our ultimate goal, d) train 5 Peruvian students in PhD programs in Peru and the US. Focusing on coursework and hands-on experiences in implementation science and epidemiology of NZDs will provide our trainees with new tools and perspectives to improve interventions, conduct well-designed studies, and build policies and programs to reduce NZDs and, in turn, reduce inequalities in health. As former Fogarty trainees ourselves, we will apply the FIC philosophy to build local individual research capacity and move us toward the WHO goal of ending neglected tropical diseases.