Washington University

NIH Research Projects · FY 2025 · 2022-09

Pain is experienced by up to 20% of the US population at any given time. Opioid pain medications are potent analgesics commonly used to treat pain symptoms, but the misuse of opioids presents a major limitation for pain management. There is increasing support for sex differences for opioid misuse among pain patients. Although, prevalence rates for prescription opioid misuse between men and women are often found to be similar, this finding contrasts with the consistently higher prevalence of abuse in men compared to women. For example, male pain patients treated with opioids are more likely to increase intake of their opioid doses, misuse prescriptions, and die from overdose. However, little is known about sex-specific trends in long-term use of prescribed opioids for pain conditions. Sexual differences appear to play an important role in opioid misuse, but preclinical data on how pain or prior history of opioid intake contributes to sex differences in opioid abuse liability are limited. Preliminary findings from our lab have recapitulated the sex-dependent trends found in clinical data using a preclinical model. Our data show that male rats self-administer higher amounts of fentanyl than females. In addition, we found that persistent inflammatory pain increases fentanyl intake in males relative to females. This suggests that inflammatory pain conditions lead to a higher misuse liability selectively in males. In this project, we propose to investigate how pain alters neuroadaptative processes during long-term opioid use in a sex-specific manner and whether these functional changes are predictive of maladaptive patterns of fentanyl intake. The mesolimbic system is a key network node that integrates pain and reward. DA transmission in the mesolimbic system, via the ventral tegmental area (VTA) to the nucleus accumbens (NAc), has long been recognized for its role in reward processing and motivated behaviors. However, it remains unclear whether sex- dependent effects of pain on opioid use are mediated by sex differences in the patterns of VTA DA activity. Our preliminary data shows that persistent inflammatory pain reduces VTA DA neuron excitability by increasing the inhibitory drive onto VTA DA neurons from the GABAergic inputs of the rostromedial tegmental nucleus (RMTg). This suggests that under conditions of pain, higher opioid doses are necessary to trigger levels of VTA DA release comparable to those produced in non-pain conditions. However, it is unknown whether similar mechanisms underlie sex differences in the patterns of opioid misuse. Based on our preliminary findings, we hypothesize that pain exacerbates RMTg-mediated inhibition of VTA DA cell activity in males, but not females following opioid self-administration. This project aims to address this gap in our understanding by investigating the sex-specific functional consequences of pain and long-term opioid use within the RMTgVTA pathway.

NIH Research Projects · FY 2024 · 2022-09

(PLEASE KEEP IN WORD, DO NOT PDF) Enter the text here that is the new abstract information for your application. This section must be no longer than 30 lines of text. Interstitial cystitis/bladder pain syndrome (IC/BPS) and chronic prostatitis/chronic pelvic pain syndrome (CP/ CPPS), are common, costly, and debilitating syndromes characterized by persistent bladder and/or pelvic pain, and urinary symptoms, such as urgency and frequency. The cause of these conditions, together referred to as urologic chronic pelvic pain syndrome (UCPPS), is unknown and their pathophysiology is poorly understood,making treatment challenging. Although several therapies have been proposed for UCPPS, none has been found to work consistently in all patients and each is either invasive or prone to significant side effects. Therefore, new therapeutic approaches are clearly needed. One such promising, but understudied, approach is dietary exclusion. This approach is supported by findings from one small single-arm trial (n=10), two case reports, our recent Multidisciplinary Approach to the Study of Chronic Pelvic Pain 1 (MAPP-1) Research Network case-crossover study, and several patient surveys indicating that a large proportion of patients believe that diet triggers their flares. However, as patient-reported triggers are numerous and wide-ranging (up to 144 candidates), identification of one common diet suitable for all patients has proven challenging. Elimination diets are also difficult to implement and test because of the large number of possible food triggers that need to be eliminated in the “elimination” phase, the large number that need to be re-introduced in the “re-introduction” phase, and the challenges of blinding elimination diets. Therefore, tools to personalize these diets are urgently needed. In pilot trials of patients with migraines, Crohn’s disease, irritable bowel syndrome, and asthma, researchers have observed significant symptom reductions following exclusion diets informed by food-specific IgG antibody concentrations. This approach is promising because it reduces the time, complexity, and burden of dietary exclusion, while simultaneously increasing its chance of success and potential for rigorous blinded evaluation. Food-specific IgG antibodies are also plausible candidates to inform UCPPS therapy because they act by biologic mechanisms implicated in UCPPS pathology (e.g. mast cell degranulation and histamine release) and in a “delayed” time frame (hours to days) consistent with patient-reported food trigger experiences. However, before we can initiate a randomized controlled trial (RCT) of food-specific IgG antibody-informed exclusion diets for UCPPS, additional empirical evidence is necessary to support this approach. For many conditions for which biomarker-informed exclusion diets have shown preliminary efficacy, researchers first demonstrated a greater likelihood of having food-specific IgG antibodies in patients than controls. We have also generated promising pilot data in a small sample of 10 UCPPS patients (100% had food-specific IgG antibodies compared to 15-33% of historical controls). In this R01, we plan to build on these promising findings by formally testing whether UCPPS patients have greater food sensitivity than controls, using existing plasma samples and clinical phenotyping data from 100 cases and 100 controls from the MAPP-1 study.

NIH Research Projects · FY 2025 · 2022-09

Summary Females may be more susceptible to Alzheimer’s disease (AD)-pathology, yet, in the early stages of AD, females perform better on verbal memory tests than males with similar levels of pathology. Nonetheless, despite this early verbal memory advantage, as disease progresses, females show faster cognitive decline. Thus, more research is needed to better characterize sex differences in AD biomarker progression, and to clarify potential mechanisms of cognitive resiliency or vulnerability to AD-pathology across the disease spectrum. To address these critical knowledge gaps, I will capitalize on our ongoing longitudinal biomarker study with the largest autosomal dominant AD kindred due to a single mutation (E280A) in the Presenilin-1 gene (PSEN1). PSEN1 mutation carriers are genetically determined to develop early-onset dementia, with mild cognitive impairment emerging at a median age of 44 and dementia at age 49. This extraordinary cohort offers the opportunity to examine sex differences in AD with few age-related confounds and methodological challenges. To this end, the candidate proposes: (1) training objectives to establish expertise in sex biology, longitudinal and multivariate modeling, and multimodal neuroimaging data, which together will further career development into an independent clinical researcher in AD; (2) a research objective to examine sex differences in the accumulation of AD-related pathology, neurodegeneration, and cognitive decline, and the potential role of steroid hormones in autosomal dominant AD; (3) a team of mentors and advisors to ensure the candidate’s success, with expertise in autosomal dominant AD (Dr. Yakeel Quiroz), biological sex differences (Dr. Jill Goldstein), sporadic AD (Dr. Reisa Sperling), multimodal neuroimaging (Dr. Chen), sex-specific differences and risk in AD (Dr. Michelle Mielke), and longitudinal and multivariate modeling (Dr. Hui Zheng). The proposed specific aims are to determine the: (1) effect of sex on AD-related pathology and neurodegeneration in PSEN1 mutation carriers; (2) effect of sex and AD biomarkers on cognitive decline in PSEN1 mutation carriers; (3) effect of steroid hormones on AD biomarker accumulation and cognitive decline in PSEN1 mutation carriers. This proposed research is innovative for investigating longitudinal sex differences in autosomal dominant AD using multimodal neuroimaging and examining the role of steroid hormones in AD biomarker abnormalities and cognitive decline. The proposed research is significant because further understanding of sex differences is crucial to inform research on prevention, early detection, design of clinical trials, and development of treatments. Overall, this project and training plan will promote the candidate’s career development by facilitating an independent program of research examining sex differences in AD and elucidating mechanisms of AD risk and resilience to inform precision interventions and treatments.

NIH Research Projects · FY 2025 · 2022-09

Abstract (Overall) The goal of the KIDney Single cell and Spatial Molecular Atlas Project (KIDSSMAP) is to create an anatomical and molecular “metaverse” of the human kidney. KIDSSMAP will accomplish this by generating multiscalar, multidimensional and multimodal maps of the adult non-diseased human kidney at a single cell resolution. This work will build on and extend the existing efforts of the investigative team through the first phase of HuBMAP. We will specifically focus on micro functional tissue units (FTUs) and structures that are along the cortico- medullary axis of the kidney. The technologies generating data include paired single nucleus chromatin accessibility and RNA expression (snRNA/ATAC-seq -cell type and state diversity), smFISH and DART-FISH (targeted high resolution spatial interrogation of 30-1000 transcripts), spatial transcriptomics (untargeted genome wide spatial mapping of dissociative technologies), CODEX (multiplexed spatial protein interrogation to bridge with RNA technologies and 3D IF technologies), 3D multiplexed immunofluorescence (3D IF- subcellular resolution to define neighborhoods in microFTUs), lightsheet fluorescence microscopy (LSFM- anatomical maps at mesoscale of key and 3D maps of neurovascular associations with FTUs) and Scattering Raman Spectroscopy (SRS-2D, 3D label free volumetric mapping at subcellular scale). The maps generated will define quantitative metrics of cell diversity, spatial defined neighborhoods across wide scales for different types of analytes interrogated by a diverse set of single call based and spatial technologies encompassing chromatin states, gene expression, protein, extracellular matrix, metabolites and lipids ranging from subcellular to volumetric meso and anatomic scale. These will be further augmented by characterizing biologically diverse (sex, age, race) samples. These tasks will be accomplished through a KIDney Organ Specific Project (KIDOSP) and a KIDney Data Analysis Center (KIDDAC), with their outputs, milestones and delivery to the HIVE centrally managed. KIDSSMAP is formed by the alliance of investigators who have extensive expertise in kidney tissue interrogation, spatial mapping and data integration. The investigators have a track record of contributing to large consortia such as HuBMAP and KPMP through fruitful and collaborative interactions. By mapping the human kidney with its diverse functional units at high resolution and large scale, KDSSMAP will generate a valuable resource for the community to understand kidney health and disease.

NIH Research Projects · FY 2025 · 2022-09

Biomedical Informatics and Data Science at Institute for Informatics (BIDS@I2) provides opportunities for undergraduate and masters’ students to explore the field of biomedical informatics and data science; to train future biomedical researchers in biomedical informatics and data science core competencies; to engage students in scholarly activities under the guidance of experienced informatics and data science faculty; and lastly, to foster their understanding of translating biomedical informatics research into practice. The objectives of the BIDS@I2 are to 1) expand students’ knowledge of theories and applications in biomedical informatics; 2) provide training and hands-on research experience in biomedical informatics and data science research; 3) assist students in enhancing and applying their knowledge and skills in translational science and dissemination of research findings; 4) encourage students to pursue graduate education and/or research careers through exposure to professional role models or mentors; and 5) develop and improve students’ skills in interdisciplinary teamwork and communication. BIDS@I2 offers a unique environment for undergraduate and graduate students to explore the breadth of biomedical informatics research. BIDS@I2 has the capacity to host approximately 20 students each year. We request funding from the NLM R25 Research Education Program to support 5 undergraduate and 4 masters students each year to cultivate a BIDS career pathway. Students’ research experiences will be enhanced through our structured training curriculum: orientation, training modules, lunch and learn research seminars, mentor lab meetings, scientific presentations, and writing, as well as through interactions and collaborations with peers, fellows, and graduate trainees from multiple disciplines, such as medicine, nursing, pharmacy, psychology, public health, biostatistics, and computer science. BIDS@I2 will promote interdisciplinary collaboration in biomedical informatics, which aims to solve problems in clinical practice, individual and population health, data science, and biomedical research through the optimal use of information. BIDS@I2 will support the following areas: 1) a supportive environment for all students; 2) various research directions to accommodate students from different academic backgrounds and skills; 3) interdisciplinary collaboration across the spectrum of biomedical informatics and data science; 4) research projects with a long-term impact on the use of information to improve clinical practice, life science, and patient-centered outcomes; and 5) prepare students for graduate school as well as future careers in STEM research.

NIH Research Projects · FY 2025 · 2022-09

The goal of this proposal is to improve genetic profiling and risk stratification for patients with myelodysplastic syndromes (MDS) using clinical whole-genome sequencing. MDS is a heterogenous group of clonal bone marrow disorders that are often fatal due to marrow failure or progression to acute myeloid leukemia (AML). Accurate prediction progression risk is therefore critical for the management of MDS patients in order to prolong survival and minimize the potential for morbidity and mortality associated with more aggressive treatments. Cytogenetic analysis of bone marrow cells from MDS patients via metaphase karyotyping is an essential component of MDS risk assessment algorithms, and is used to detect chromosomal deletions, duplications, and aneuploidies that are associated with differential clinical outcomes. Although karyotyping has been used effectively for decades, it has several disadvantages. These include low genomic resolution and high failure rates that can result in incomplete genetic risk profiles for some patients. We recently developed and validated ChromoSeq, a robust CAP/CLIA-compliant whole-genome sequencing (WGS) assay for genetic profiling of patients with myeloid malignancies. We showed that this method was 100% sensitivity for clinically relevant cytogenetic abnormalities in AML and identified additional cytogenetic events in up to 25% of patients that were not detected by standard cytogenetics. These findings included new risk-defining chromosomal abnormalities in almost 15% of patients, which resulted in better prediction of clinical outcomes. Although MDS and AML are closely related diseases that share many features, the genomic characteristics and cellular composition of MDS is distinct. In addition, the use of ChromoSeq results to form existing MDS risk groups has not been clinically validated. We hypothesize that optimization of the ChromoSeq whole-genome sequencing assay for MDS samples will improve the accuracy of genetic profiling and risk stratification of MDS patients. Here we propose to use a combination of retrospective and prospective clinical MDS samples to validate ChromoSeq for genetic profiling and risk assessment in MDS patients. We will first use retrospective MDS samples to optimize and validate our existing CAP/CLIA-compliant ChromoSeq WGS assay to improve the detection of low frequency mutations, copy number alterations (CNAs) and copy neutral loss of heterozygosity (CNLOH), which are common in MDS (Aim 1; UH2 component). We will then use a prospective MDS cohort to establish the clinical validity of ChromoSeq assay for genomic profiling and risk assessment of MDS patients. This project will expand the use of the CAP/CLIA-compliant ChromoSeq assay to MDS samples so that it may be used for future interventional clinical trials and routine clinical testing of patients with this malignancy.

NIH Research Projects · FY 2025 · 2022-09

Project Summary Gliomas are the most common brain cancer. Whereas malignant gliomas predominate in adults, low-grade gliomas (LGGs) comprise the majority of brain tumors in the pediatric population. While LGGs are not typically fatal, young children with these neoplasms commonly have long-term medical morbidities, from either the tumor itself or the neurotoxicity associated with conventional therapies. This is particularly true for individuals with the Neurofibromatosis type 1 (NF1) cancer predisposition syndrome, where nearly 20% of children will develop LGGs involving the optic pathway (optic pathway gliomas; OPGs) that can lead to vision loss. Currently, therapies for NF1-LGGs are focused on arresting the growth of the cancer cells using either genotoxic (e.g., carboplatin/vincristine) or molecularly targeted (e.g., MEK inhibitors) treatments, with variable durable effects. Importantly, 30-50% of the cells in human NF1-LGGs are non-neoplastic cells, such as neurons, lymphocytes (T cells) and monocytic cells (macrophages and microglia), which our laboratory has shown are required for both tumor formation and growth in experimental murine models of Nf1-OPG. Using these Nf1-OPG mouse strains, we have previously defined a “neuron-immune-cancer cell circuit” in which Nf1-mutant neurons activate T cells to produce cytokines that stimulate microglia to support LGG formation and continued growth. Specifically, we demonstrated that NF1-mutant human and murine neurons produce midkine, which activates T cells in vitro and in vivo to secrete Ccl4, which then acts on microglia to induce Ccl5 expression, an essential growth factor for Nf1-OPG formation and growth. Surprisingly, we found that CD8+ T cells predominate in both human and mouse NF1-LGG, where high CD8, but not CD4, levels correlate with reduced overall survival in people with LGG. Moreover, studies in our laboratory revealed that antibody-mediated CD8+ T cell depletion reduces mouse Nf1- OPG growth in vivo. Based on these findings, we hypothesize that CD8+ T cells function in a neuron- immune-cancer cell circuit as obligate modulators of LGG development and progression. To test this hypothesis, we have designed a series of experiments we have designed a series of experiments that aim to (a) define the immune composition of Nf1 optic gliomas in mice, (b) determine why CD8+ T cells are selectively recruited in these murine brain tumors, and (c) elucidate how NF1 mutation in neurons modifies T cell-microglia interactions. Collectively, these studies aim to mechanistically dissect the role of CD8+ T cells in neuron-immune- cancer cell axis regulation of LGG formation and growth, relevant to the development of future immunomodulatory therapeutic strategies. OMB No. 0925-0001/0002 (Rev. 03/2020 Approved Through 02/28/2023) Page 1 Continuation Format Page

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Emotion dysregulation is transdiagnostic, integral to most affective and disruptive behavior disorders, and is associated with impaired functioning across domains from health to academics. The study of dysregulation of negative affect, or irritability, has resulted in a better understanding of how it predicts later impairment and general psychopathology and has identified neural correlates. However, there has been less focus on dysregulation of positive affect, despite recent evidence suggesting it contributes to the development of psychopathology, particularly externalizing symptoms, and impairment across domains from general to social functioning. Dysregulated positive affect may not only confer independent risk, but is likely to have separate underlying neural correlates, as positive and negative emotional valence systems are different domains in the Research Domains Criteria. While surgency, the temperamental measure of high positive affect, has been related to increased aggression and externalizing symptoms in infants and young children, how this relates to clinical dysregulation of positive affect, or excitability, is unknown. Moreover, the relationships between surgency and excitability with psychopathology and impairment have been largely unstudied in young children. This proposal addresses these gaps in understanding by studying the overlapping and separable contributions of surgency (normative high positive affect), excitability (clinically related dysregulated positive affect), and irritability (clinically related dysregulated negative affect) to symptoms of psychopathology and impairment in school age children. Additionally, this proposal will assess the overlap and distinctions in brain-behavior relationships between dysregulation in positive and negative affect. Specifically, 100 7-10-year-old children enriched for emotion dysregulation will be assessed using a research diagnostic interview and parent and self- report measures of emotional and general functioning at baseline and after one year. At baseline, children will undergo functional MRI scans during emotion response and regulation tasks. Consistent with the NIMH Strategic plan, particularly Strategy Objective 2, to “chart mental illness trajectories to determine when, where, and how to intervene,” understanding the separable contributions of surgency, excitability, and irritability to risk trajectories and elucidating the neural correlates of such can provide meaningful targets for early identification and intervention in multiple disorders. Under the mentorship of a diverse team of experts in emotion regulation and development, developmental psychology and psychopathology, and longitudinal and statistical methodology, the training provided through this proposal will facilitate the applicant gaining expertise in fMRI methods for studying affective processing, dimensional constructs in developmental psychopathology, and longitudinal design and analysis. This training will provide the foundational components of the applicant's long-term goals of understanding the neural and behavioral development of emotion dysregulation, specifically excitability, to inform early identification and interventions for improving emotion regulation prior to significant impairment.

NIH Research Projects · FY 2025 · 2022-09

PROJECT ABSTRACT Approximately 1.7 million children under 15 years old were living with HIV in 2020; and most new HIV infections (85%) occurred in Sub-Saharan Africa (SSA). People living with HIV (PLHIV) often struggle with mental health comorbidities that lower their antiretroviral therapy (ART) adherence. However, 76% to 85% of PLHIV in SSA receive no treatment for serious mental health disorders, especially depression. Depression reduces ART adherence, which negatively impacts health and increases HIV transmission risks. Older adolescents (≥14 years) living with HIV are particularly vulnerable to these risks as caregivers withdraw or lessen their support during their transition to young adulthood. Moreover, older adolescents are also moving into larger and less accommodating adult HIV clinic settings and are at risk for dropping out of ART programs. Given that mental health services are severely under equipped in SSA, including in Uganda, and are inaccessible by many YLHIV, new solutions to increase access to mental health care and close the treatment gap are urgently needed. The overall goal of this proposed R21/33 study is to develop an mHealth intervention (Suubi-mhealth) for use among Ugandan youth (14-17 years) with comorbid HIV and depression, taking into account their unique contextual, cultural, and developmental needs. This digital therapy intervention delivered via a mobile application, will utilize the core tenets of cognitive-behavioral therapy (CBT) found to improve depression and ART adherence. The proposed study will specifically: Phase 1. R21 Aim 1: Develop and iteratively refine an intervention protocol for Suubi-Mhealth based on formative work to understand needs of youth living with HIV (YLHIV). We will conduct four focus groups with youth and two focus groups with health care providers (6-8 youth each) for feedback on intervention content and methods to increase participation and retention. R21 Aim 2: Based on results of Aim 1, explore the feasibility and acceptability subsequent refinement for the larger R33 phase. Phase of Suubi-Mhealth on a small scale (N=30), to inform 2. R33 Aim 1: Test the preliminary impact of Suubi- Mhealth versus a waitlist control group (N=200), on youth outcomes (depression, ART adherence, mental health functioning, quality of life, stigma). R33 Aim 2: Examine barriers and facilitators for integrating Suubi-Mhealth into health care settings for YLHIV. The study will be conducted in 10 health clinics in the greater Masaka region in Southern Uganda. We expect for Suubi-Mhealth to be an acceptable and feasible mHealth tool to reduce depression, improve ART adherence and overall mental health functioning among YLHIV. If the results of this pilot are promising, then the next step is an R01 to rigorously test Suubi-Mhealth in a larger trial, spanning multiple sites across Uganda.

NIH Research Projects · FY 2025 · 2022-09

Cervical cancer is the leading cause of cancer death in Zambia, where HIV prevalence is also high (11.3%). HIV heightens the risk of developing and dying from cervical cancer. Primary cervical cancer prevention can prevent many cases of cervical cancers. However, use of these primary prevention tools is low in low- and middle- income countries (LMICs), particularly among girls with HIV, who are most at risk of poor cancer outcomes. To ensure that ALHIV have access to evidence-based primary cervical cancer prevention tools, we propose to support strengthening of primary cervical cancer prevention integrate into routine care in adolescent HIV clinics. Adolescent HIV clinics in Zambia have regular contact with ALHIV and are trusted sources of health information for the community. Given the known challenges of providing cervical cancer prevention in LMICs, including Zambia (e.g., resources, staffing, supply chain), integrating primary cervical cancer prevention requires a multilevel approach, Ministry of Health engagement, and diversified implementation strategies. To achieve success, we will co-design a package of implementation strategies using a previously successful implementation research approach developed for cervical cancer prevention in LMICs: the Integrative Systems Praxis for Implementation Research (INSPIRE). INSPIRE is a novel, formal, and comprehensive framework to develop, implement, and evaluate implementation science efforts. Following key elements of INSPIRE, our specific aims are to: 1) Identify the unique multilevel contextual factors (barriers and facilitators) across HIV settings (rural, urban, peri-urban) that influence primary prevention; 2) Use Implementation Mapping to translate findings from Aim 1 and Ministry of Health priorities into a package of implementation strategies to integrate into HIV clinics; 3) Conduct a feasibility trial to evaluate the package of multilevel implementation strategies for primary prevention into HIV clinics. Our research team has significant expertise in HIV and cancer prevention, and strong institutional support including $325,000 over the course of the study; strong support, technical expertise, and resources from the Zambian Ministry of Health; and political will for scale-up. If successful, this implementation model could be transported to HIV clinics across Zambia and serve as a model to address cancer prevention priorities for those with HIV in other LMICs.

NIH Research Projects · FY 2025 · 2022-09

Environmental factors that increase or decrease disease risk may do so by chemically altering parts of the DNA that regulate gene expression. These modifications to the genetic code are referred to as “epigenetics.” Epigenetic changes can be acquired via a variety of exposures, including pollution and lifestyle behaviors. The benefit or harm of these changes depends on the purpose of the gene and how the change alters gene expression. Communication strategies that help people understand and use epigenetic information are needed to facilitate the translation of epigenetics research to clinical settings. Our objective is to facilitate the translation of basic epigenomics science into clinical and public health practice by developing epigenetic communication strategies that are understandable and useful to the public. The specific aims will: 1) Determine how people come to understand epigenetics information; (2) Examine how public perceptions of the benefits and risks of epigenetics research might facilitate or impede (a) understanding of epigenetics concepts and (b) acceptance of using epigenetic technologies in clinical settings; (3) identify strategies for communicating information about epigenetics in a way that increases understanding of epigenetic concepts and increases acceptance of using epigenetics in clinical settings; and (4) Evaluate the generalizability of the findings from Aims 1-3 to the U.S. population. We will use an exploratory sequential mixed methods design that includes one qualitative phase followed by two consecutive quantitative phases. The qualitative phase will include 20 focus groups (N=160 total, n=8 per group). The quantitative phases will include a nationally survey (N=1,870) and a full-factorial experiment (N=1,954). Our proposed research will overcome a critical barrier to the translation of basic epigenetics research into clinical and public health practice: limited scientific knowledge about how to communicate about epigenetics in way a that is understandable and useful to the public. Furthermore, our findings could be used as a foundation for research that communicates information about other emerging genomics technologies to the public. We will expand our impact further by developing a toolkit for disseminating our discoveries.

NIH Research Projects · FY 2025 · 2022-09

Overall Summary Given the historical outbreaks of coronaviruses, coupled with the recent emergence of SARS- CoV-2 and the destabilizing consequence of COVID-19 on global health and economy, there is an urgent and critical need to develop new vaccines capable of broad protection against existing and future Sarbecoviruses and Merbecoviruses. This P01 program project (PPG) addresses the hypothesis that a combination of evolutionarily-designed and optimized B and T cell antigens can confer broad and protective immunity against Sarbecoviruses and Merbecoviruses that currently exist or could emerge from zoonotic reservoirs. The PPG integrates the work of eleven leading laboratories with records of collaboration that have expertise in coronavirus biology, viral pathogenesis, B and T cell immunity, vaccine development, animal challenge studies, structural biology, antibody structure and function, antigen design, and evolutionary analysis of viruses. All Projects and Cores plan interactive studies with the focused goal of designing optimized B and T cell antigens for incorporation in adenoviral (ChAd) and vesicular stomatitis virus (VSV) vectors to create mucosal and systemic vaccines that protect against infection and disease caused by a range of coronaviruses of potential concern. The PPG is served by a central Animal Challenge Core that performs vaccination and infection experiments in mice and hamsters and a central Administrative Core that streamlines data management and sharing, provides computational analysis for down-selection and scientific decision-making, and facilitates communication. Our proposal and antigen design program serves as a blueprint for possible product development with ChAd vaccines, VSV-based vaccines, or even with other platforms (e.g., mRNA vaccines, nanoparticles, etc.) not directly evaluated here. By the conclusion of our PPG, we envision generating at least one and likely multiple viral- vectored vaccine platforms that induce broad spectrum immunity to multiple coronaviruses of concern including human and zoonotic Sarbecoviruses and Merbecoviruses that could emerge in the future.

NIH Research Projects · FY 2025 · 2022-09

ABSTRACT Alzheimer’s disease (AD) manifests itself differently across men and women, but the genetic and molecular factors that drive this remain largely elusive. AD is the most common cause of dementia, affecting over 5 million people in the USA alone, and till today remains essentially untreatable. Because AD has a strong genetic component, with inheritance estimates between 50-80%, studying the genetics of AD can importantly aid the discovery of novel drug targets. However, evidence from various lines of research suggests that sex differences are an integral part of AD. It is therefore crucial to study the genetics of AD in a sex-specific manner, as this will help the field gain important insights into disease pathophysiology, identify novel sex-specific risk factors relevant to personalized genetic medicine, and uncover potential new AD drug targets that may benefit both sexes. To date, surprisingly few sex-specific variants/genes have been identified, which we hypothesize is because prior studies were faced with several obstacles such as data limitations and unexplored research avenues. This project proposes to use big data together with state-of-the-art approaches in order to leverage established sex differences in AD as a means of elucidating novel AD risk genes. Aim 1 will improve on prior sex-stratified genome-wide association studies (GWAS) by using larger sample sizes and extensively harmonized data. This includes a cross-cohort phenotype harmonization and powerful models using age information to improve AD risk associations. In addition, we will, for the first time, explore the role of rare variants on AD in a sex-specific manner. Aim 2 will use parallel strategies to Aim 1, but will focus on the X chromosome, which has remained largely unexplored in the field of AD genetics. For both Aims 1 and 2, we will further validate putative associations by evaluating their sex-specific effects on gene transcript expression and protein levels in brain tissue. Similarly, associations will be validated in a sex-specific manner using AD-relevant endophenotypes (e.g. tau levels in the cerebrospinal fluid) from deeply phenotyped cohorts. Aim 3 will follow a different innovative approach to sex- specific AD gene discovery by identifying sex-specific AD-related protein changes in brain tissue and determining the genetic variants that drive them. The latter variants will then be validated by relating them to risk for AD. The independent phase of this project will focus on the use of multi-omics data to corroborate sex-specific gene associations with AD risk, as well as proteomics data to discover new AD risk genes. Central to the success of this proposal, Dr. Belloy will have the support from an established group of experts in genetics, imaging, and neurology (Dr. Michael Greicius), multi-omics data integration (Dr. Stephen Montgomery), proteomics analyses (Dr. Nicholas Seyfried), sexual dimorphism (Dr. Marcia Stefanick), and rare variant analyses (Dr. Zihuai He), providing him with the necessary skillsets to embark on a career as an independent scientist.

NIH Research Projects · FY 2025 · 2022-09

ABSTRACT Two million Americans are hospitalized for sepsis each year, and 1 in 3 die. Those that survive, however, are not cured. Neurocognitive disorders occur in up to 50%, and cognitive decline continues for up to 8 years. Sepsis hospitalizations account for a higher proportion of unplanned readmissions than those for myocardial infarction, heart failure, and COPD. Five-year mortality for sepsis survivors exceeds that for heart failure and stroke. The mechanisms underlying this persistent loss of health remain to be defined. We hypothesize that early during sepsis the mitochondrion is restructured as an adaptive mechanism to protect the cell against any future environmental stress, such as recurrent sepsis. These structural changes impart lasting alterations to the mitochondrial calcium (Ca2+) homeostasis and metabolism necessary to support a cellular phenotype, which for a multicellular organism are poorly tolerated and underlie a persistent loss of health. Our lab has spent nearly two decades studying sepsis to elucidate the Ca2+-dependent mechanisms that regulate mitochondrial biology to balance Ca2+ homeostasis and ATP generation and preserve cellular health. We have shown that early after sepsis, mitochondrial depolarization generates a Ca2+ signal. Members of the family of Ca2+ /calmodulin-dependent protein kinases (CaMK) transduce these Ca2+ signals and work in tandem to mediate adaptive changes in mitochondrial fission, mitophagy, and oxidative metabolism to lessen cellular damage. More recently, we observed that sepsis restructures the mitochondrial calcium uniporter (MCU) complex, imposing long-lasting changes to mitochondrial and cellular Ca2+ homeostasis and metabolism that perturb cellular and tissue function across the entire organism. We propose that as a ‘learned’ response to sepsis, the cell restructures the MCU complex to counter the potential for Ca2+ overload with future insult; this imparts long-lasting alterations in Ca2+ homeostasis, oxidative metabolism, and tissue phenotype. Using models of lower-respiratory tract and intraperitoneal infection and correlative human samples, we propose the following aims: Aim 1. To study in mice and humans how a restructured MCU complex alters Ca2+ homeostasis and oxidative metabolism and thereby, the phenotype of each tissue comprising the organism. Aim 2. To define the mechanisms of mitophagy and protein degradation through the lysosome and proteasome as underlying causes of the persistent loss of MICU1 expression in murine models of sepsis and in human sepsis survivors. This new experimental work will provide foundational knowledge as to how the mechanisms governing mitochondrial Ca2+ and metabolism are restructured during sepsis to underlie a persistent loss of cellular phenotype that leads to a progressive loss of health and shortened survival.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Mutations in β-cardiac myosin cause cardiomyopathies, which are leading causes of sudden cardiac death. There are currently no cures for cardiomyopathies. Drugs targeting β-cardiac myosin would transform treatment of cardiomyopathies and heart failure, the leading cause of hospitalizations in the United States. Efforts to develop new therapeutics have been hampered by an incomplete understanding of how mutations in β-cardiac myosin cause cardiomyopathy and of how existing compounds targeting myosins achieve specificity. Recent studies have found that several hypertrophic cardiomyopathy (HCM) variants release myosin motors from an auto-inhibited state where two myosin motors fold back on each other. Unfortunately, only a small subset of cardiomyopathy variants has been functionally characterized, even though insight into a variant’s molecular effects is crucial for determining pathogenicity and predicting drug response. Drugs targeting β-cardiac myosin need to achieve a high degree of specificity to avoid disrupting critical functions carried out by other myosin proteins. Though the field has focused on sequence differences at compound binding sites, one myosin inhibitor discovered in high throughput screens is highly specific even though sensitive and insensitive myosin isoforms share identical sequences at its binding site. This suggests structural dynamics at the binding site are an important determinant of specificity. We hypothesize that hidden structural states during equilibrium fluctuations of myosin motors determine functional effects of mutations and the specificity of compounds targeting myosins. We propose using molecular dynamics simulations to elucidate the effects of a large number of β-cardiac mutations and to compare pocket openings between myosin isoforms. We will leverage our unique combination of enormous computational power and cutting-edge machine learning tools for analyzing protein allostery. Aim 1 is to identify β-cardiac myosin mutations that alter the balance of active and auto-inhibited states. Specifically, we will investigate the effects of HCM variants without functional data, dilated cardiomyopathy variants, and “variants of unknown significance.” Aim 1 directly addresses the NIH’s goals of investigating new pathobiological mechanisms and leveraging data science to open new frontiers in biomedical research. Aim 2 is to predict drug specificity based on differences in conformational ensembles of myosin isoforms. In particular, we will compare pocket opening probabilities at the binding sites of three clinically relevant compounds before designing and experimentally testing novel compounds that bind specific pockets in β- cardiac myosin. Aim 2 directly addresses the NIH’s goals of developing novel therapeutic strategies. Together, these results will inform our understanding of how mutations in β-cardiac myosin cause cardiomyopathies and advance drug discovery for cardiac disease.

NIH Research Projects · FY 2025 · 2022-09

Project Summary. The Washington University T32 postdoctoral training grant “Translational Imaging in Radiopharmaceutical Sciences” (abbreviated as “TIRS”) is a multidisciplinary initiative to train the next generation of PET molecular imaging scientists to design, develop, and translate PET radiotracers for noninvasive detection of pathophysiology mediating development of neurodegenerative diseases and ADRDs. This training program will be spearheaded by Mallinckrodt Institute of Radiology (MIR), an institution known worldwide as a frontier for translational PET research, and a preferred first stop for radiopharmaceutical scientists for learning applied nuclear imaging research. To accomplish this novel training objective, the T32 TIRS program brings leading scientists from radiological sciences division of the MIR, the Knight Alzheimer’s Disease Research Center (Knight ADRC), Molecular Imaging Center, Institute of Clinical and Translational Sciences, Cyclotron-GMP facility and PET-Radiotracer Translational and Resource Center (P41 program through NIBIB) and faculty members as mentors from 12 departments to accomplish aims of proposed training for investigation of new or existing biomarkers mediating AD and ADRDs with overlapping pathophysiology, assist in noninvasive early detection of diseases, facilitating drug development and allow stratification of treatment paradigms for managing patient care. Of note, the T32 TIRS mentors include chemists, biochemists, biomedical engineers, immunologists, neurologists, psychiatrists, radiochemists, and radiologists. The postdoctoral trainees will be drawn through various participating departments at the medical school, and Danforth campus, including other universities in metropolitan St. Louis area, and nearby academic institutions in the Midwest. We are proposing to train 3 postdoctoral trainees each year, wherein each trainee will be enrolled for 2 years of training, over 5 years of the T32 program. All trainees will be trained through mechanism of dual- mentorship (one in basic science and other physician-scientist to provide multidisciplinary training) and all trainees will be trained in responsible conduct of research including instructions in methods of experimental rigor for accomplishing reproducible science. Of note, scientists training at the interface of neuroscience and radiopharmaceutical sciences are also rapidly declining nationwide in general, and there is a clear shortage of radiopharmaceutical scientists across the board in the US workforce. Therefore, our T32 postdoctoral training program TIRS would be expectedly to substantially meet this critical shortage of radiopharmaceutical scientists trained at the interface of neuroscience and radiopharmaceutical sciences. Although our current T32 TIRS programmatic mission is directed for trainees at ADRDs, we envision that trained molecular PET imaging research scientists could also serve other departments and fields including the medical oncology, therefore, extending significance and scope of TIRS training program well-beyond the field of neuroscience. Overall, we (the T32 TIRS program director and co-directors) are confident that postdoctoral scientists trained through this program will be transitioning into faculty positions nationwide and will be expected to serve the field of diagnostic nuclear medicine in neuroscience for several decades.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT Type 1 diabetes (T1D) is one of the most common chronic childhood diseases and has a rapidly rising incidence and prevalence. Management of blood glucose is challenging and often leads to frequent and rapid swings between normal, high, and low glucose. This glucose variability has recently been identified as an independent risk factor for diabetic complications. One of the least studied and understood complications of T1D is altered cognition and its relationship to glucose variability. As the brain uses a substantial amount of the body’s glucose to support its functions, especially during dynamic periods of brain development, there are strong biological reasons for the potential vulnerability of youth with T1D. Several studies have shown lower cognitive scores in youth with T1D compared to their peers without T1D, and shown that poorer overall glycemic control as indicated by severe glycemic events (e.g., diabetic ketoacidosis) and chronic hyperglycemia, are associated with lower cognitive scores both acutely and long-term. However, these studies we conducted in a controlled, optimized laboratory settings at a single time point. Little is still known about how typical glucose fluctuations that occur daily in real-life settings affect cognitive function in the moment, particularly dynamic cognitive skills that are sensitive to environmental or physical conditions of the individual (e.g. sleep deprivation) and thus fluctuate throughout the day (e.g., working memory, processing speed). Determining the relationship between glycemic fluctuations and dynamic cognitive function is critically important in youth given that dynamic cognitive skills are fundamental for learning, integrating, and using novel information in their daily lives. Understanding how these skills are affected in daily life, how they relate to glycemic fluctuations, and any contributing factors, could motivate improvements in academic accommodations and T1D treatment recommendations. Importantly, this work could also be expanded to better understand the impact of glycemic variability on brain health throughout the lifespan in T1D and could be translated to other forms of diabetes (e.g., type 2 diabetes, gestational diabetes). The primary goal of this study is to determine if real-time, real-life glycemic variability predicts fluctuations in dynamic cognitive function in youth with T1D. A secondary goal is to determine whether dynamic cognitive function in youth with T1D differs from youth without T1D, independent of significant glycemic extremes. To address these questions, I will apply a unique and innovative combination of continuous health and cognitive data collection methods, multivariate time-series analyses, and machine learning models. This work, combined with a tailored career development plan, will form the basis of my future work as an independent investigator, which will expand to address how diabetes-related health behaviors (e.g., food choice) impact glucose fluctuations, and subsequent brain health (i.e., cognitive function), using novel mobile health (mHealth) technologies across different types of diabetes.

NIH Research Projects · FY 2025 · 2022-09

Project Summary: Human genetic studies have linked autism and related neurodevelopmental disorders (NDD) to disruption of genes encoding epigenetic factors. While mutations in these genes can affect more than one pathway that leads to ASD, identification of a common molecular pathway across genetic causes of ASD can provide insight into broadly applicable therapies. Recent evidence from our laboratory suggests that a neuronal-specific form of DNA methylation is a shared epigenetic modification that is disrupted in ASD-associated neurodevelopmental disease. Though DNA methylation is classically considered to only occur in mammalian cells in the CpG context, neurons are uniquely enriched for methylation in non-CpG contexts established by DNMT3A. This non-CpG methylation primarily occurs at CA dinucleotides (mCA) and is critical for proper neuronal development and function. Though mCA plays an important role in regulating neuronal gene expression, it is not known how the mCA landscape is faithfully established across the neuronal genome, and whether additional NDDs involve disruption of mCA. Recent studies outside the nervous system have suggested that histone modifications may play a central role in directing DNMT3A-mediated DNA methylation across the genome, but it is not known to how these histone modifications influence DNMT3A and mCA throughout the neuronal genome. Intriguingly, mutations in H3K36 histone methyltransferases have been recently identified in ASD gene studies. Additionally, recent studies conducted outside the nervous system have suggested that H3K36 methylation recruits DNMT3A and regulates its activity. In this proposal, I will determine how disruption of H3K36 methylation-mediated interactions with DNMT3A disturbs critical patterns of mCA across the neuronal genome to drive brain dysfunction. In Aim 1, I will disrupt NDD-relevant H3K36 methyltransferases and measure how DNMT3A recruitment and mCA are affected. This will define the mechanisms underlying histone and DNA modification dynamics on regulating neuronal transcription and building our basic understanding of how disruption of mCA drives disease. In Aim 2, I will explore how loss of H3K36 dimethylase NSD1 causes dysregulation of neuronal genes via shared neuronal chromatin pathology observed across heterogenous clinical syndromes of ASD. I will use genomic approaches to examine how altered DNA methylation as a result of NSD1 loss affects enhancer activity to drive transcriptional dysregulation in nervous system dysfunction and disease. This analysis will begin to identify key downstream chromatin associated factors across a common molecular pathway involved in multiple genetic causes of ASD to provide insight into functional cellular outcomes and broadly applicable therapies.

NIH Research Projects · FY 2025 · 2022-09

Abstract Immune privilege is a term applied to organs such as the eye that have a unique relationship with the immune response. These sites prohibit the spread of inflammation since even minor episodes can threaten vision. The breakdown of immune privilege is thought to have serious consequences for the eye; however, in spite its powerful influence on inflammation we know little about how immune privilege influences retinal diseases. Müller cells are the major glial cell of the retina that maintain structural integrity. They react in virtually every eye disease but are strikingly resistant damage; importantly they can suppress inflammation. Based on this they should be key participants in immune privilege, but this is not understood. The biological process of autophagy (literally self-eating) is a recycling system that destroys inflammation-inducing cellular debris to prevent tissue damage. Because immune privilege, Müller cells, and autophagy all have anti- inflammatory properties, we will test the novel hypothesis that Müller glial cells utilize the autophagy pathway to support the anti-inflammatory nature of the eye (i.e. immune privilege). We will do this by examining intraocular inflammation in 2 well-characterized disease models, endotoxin induced uveitis (EIU) and experimental autoimmune uveitis (EAU), utilizing mice with autophagy-deficient retinal Müller cells. Since EIU is mediated by the innate arm of the immune system and EAU is an antigen specific T cell mediated disease we will get thorough understanding as to how Müller cell autophagy influences different types of immune mediated diseases of the eye. In Aim 1 we will assess the intraocular inflammatory response in these models by comparing control mice with mice that have had the essential autophagy genes Atg5 and Fip200 deleted specifically in Müller glial cells. We will evaluate inflammatory infiltrates, cytokine production, and retinal integrity in both models. Preliminary data suggests that inflammation and retinal damage are enhanced in the presence of autophagy deficient Müller cells in both models. In Aim 2 we will we will define the molecular basis of these enhanced inflammatory response to determine the contribution of autophagy to immunoregulatory properties of Müller cells in the eye. Studies will include scRNA-seq analysis, examination of the blood retinal barrier, analysis of the T-cell response in EAU, and morphological analysis. In Aim 3 we will test the idea that autophagy supports phagocytosis in Müller cells promoting their anti-inflammatory properties using the process of LC3-associated phagocytosis (or LAP). We will test whether LAP recovers a portion of the vitamin A for the Müller visual cycle. By elucidating the role of Müller cells and autophagy in immune privilege we will better understand their influence on ocular inflammatory responses. Thus, rather than just treating inflammation, we could consider upregulating immune privilege alone, or in combination with other treatments, to target retinal disease.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY An effective epidermal permeability barrier (EPB) protects the skin from dehydration, inflammation, premature aging, environmental exposure, and infection. Epidermal barrier dysfunction is an important feature of atopic dermatitis, as well as numerous skin diseases including psoriasis, acne, and rosacea. A fundamental and holistic understanding of mechanisms regulating homeostatic barrier function is essential to effectively prevent and manage barrier abnormalities. The EPB function resides in the skin epidermis, which is home to diverse microbial communities. The microbiome is recognized as a functional unit of the skin barrier. The skin ecosystem is continuously challenged by the external exposome that includes ultraviolet radiation (UVR), air pollutants and allergens. Critical for the barrier defense and homeostasis are xenobiotic sensors that recognize external signals and help identify beneficial (e.g., commensal microbes) from harmful (e.g., pollutants, pathogens) xenobiotics to regulate barrier defenses. Recently, I have demonstrated that commensal microbes regulate epidermal differentiation and barrier permeability of the skin by activating xenobiotic sensor, the aryl hydrocarbon receptor (AHR). However, the mechanisms by which commensal microbes regulate EPB through AHR under homeostasis, and in presence of environmental insults such as UVR are unexplored. The central hypothesis of this proposal is that tuning of epithelial responses by modulating AHR-commensal interactions can alter barrier permeability. This project utilizes ‘multi-omics’ approaches by integrating transcriptomics, metagenomics, and metabolomics to understand host-microbiota interactions in skin barrier repair. In Aim 1, I will identify microbial signals from a synthetic commensal community that can activate AHR. These studies will lead to identification of microbial ligands that can be used to target AHR in barrier diseases. In Aim 2, I will test contributions of commensal microbiome in protecting against UV-induced barrier damage and use multiomics approaches to characterize microbiome-host-UV interactome in the context of AHR signaling. These studies will provide a framework to generate therapies that leverage understanding of environmental-host-microbiome interactions. During the K99 phase, I will be trained in metabolomics to identify microbial metabolites. I will receive advanced training in bioinformatics and systems biology approaches that focus on integrating multiple omics datasets. The outstanding training environment at the University of Pennsylvania coupled with the excellent advisory committee I have assembled, will greatly facilitate my research during the mentored phase as well as launch my career with the skills necessary for understanding the role of the microbiome-host- environment interactome in regulating skin barrier repair.

NIH Research Projects · FY 2025 · 2022-09

PROJECT ABSTRACT The number of adolescents living with HIV (ALHIV) in Uganda is over 170,000 and growing. Ugandan ALHIV are a priority due to social and economic challenges that make them highly vulnerable to HIV infection and sub-optimal access and adherence to antiretroviral therapy (ART). Less than 50% of ALHIV in Uganda are ART-adherent, leading to low rates of viral suppression and high attrition from HIV care. In response to calls for expanding differentiated care approaches for ALHIV and new forms of combination HIV interventions, we seek to intervene on social and economic challenges that exacerbate the risk for viral load non-suppression. Addressing these hardships can improve ALHIV’s livelihoods and give them the knowledge and resources to sustainably manage HIV. We have shown the effectiveness of Suubi (Hope) in four NIH-funded RCTs in Uganda. Suubi is an evidence-based and theory-informed combination intervention with four components: 1) Financial Literacy Training; 2) Incentivized Matched Youth Savings Accounts with income-generating activities; 3) a manualized intervention for ART adherence and stigma reduction; and 4) Engagement with HIV treatment-experienced role models who share lived experiences of HIV. Suubi has shown robust effects on viral suppression and ART adherence, mental health and psychosocial outcomes, and family financial stability. Yet, it is unknown if each component in Suubi had a positive effect, how the components interacted, or if fewer components could have produced equivalent effects. Hence, we propose a factorial experiment to unpack and optimize Suubi to enhance scale up in health systems using the multi-phase optimization strategy (MOST). We define our “optimization objective” as the most cost-effective combination of intervention components considering three real-world constraints: 1) efficiency, 2) affordability, and 3) scalability. We then evaluate the intervention component effect sizes and balance these data against real-world information and costing data to empirically arrive at optimization. The study aims are: Aim 1. Conduct a factorial experiment (optimization trial) to test the main effects of each of the four Suubi intervention components and combinations of components (interactions) on viral suppression (primary outcome); Aim 2. Test mediators and explore moderators that explain and modify the relationship between each of the four Suubi intervention component and viral suppression; and Aim 3. Compare the cost and cost-effectiveness of each of the four Suubi intervention components and every combination of components. We will use a 24 factorial experiment with 16 conditions representing all combinations of the 4 components. Health clinics (N=48) will be randomized to 16 conditions (12 ALHIV per clinic), yielding main effects and interaction effects for the 4 components on sustained viral suppression (defined as an undetectable viral load at 12-, 24- and 36-month follow-up assessments). An optimized intervention built within real-world constraints in SSA for a high-priority group is an innovative and promising way to advance intervention science for HIV care globally.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT Spinal cord injury (SCI) is a life-altering event that leads to long-lasting motor impairment. Currently, there is no cure for paralysis. Electrical spinal cord stimulation (SCS) combined with exercise training can restore posture control, stepping, and voluntary walking in humans with SCI. However, the neurorecovery mechanisms induced by electrical neuromodulation of the spinal cord are poorly understood. This project will generate evidence-based knowledge of changes in short-term excitability and long-term plasticity of the neural circuits that may mediate SCS-induced improvements in motor function. Participants with SCI and control subjects will perform 30-min leg training sessions with a non-invasive body-machine interface controlling a computer cursor, and perform game- like activities using voluntary movements and/or non-invasive transcutaneous SCS. We will quantify changes in corticospinal, reticulospinal, and spinal neural excitability will be quantified by comparing motor-evoked potentials elicited by transcranial magnetic stimulation, the StartReact response, and the F-wave responses respectively before and after training. We will determine (1) short-term changes in neural excitability that are independently enabled by SCS and activity-based training, (2) whether task-specific training used commonly used in rehabilitation enhances short-term changes in neural excitability, and (3) long-term changes in neural plasticity mediated by SCS combined with activity-based training in individuals with chronic SCI. A clear understanding of SCS-enhanced neural mechanisms and how they promote neural plasticity through residual corticospinal, reticulospinal, and spinal connections will promote the development of personalized therapies that directly target the specific excitability and plasticity states of these circuits to promote and enhance functional recovery in individuals with SCI. Throughout the award period, I will obtain new skills and expertise in conducting clinical studies as the lead investigator. In addition, I will gain further training in neurophysiological evaluations of motor and sensory function, evaluation of cortical and spinal cord plasticity, spinal cord stimulation, career development, and R01-level grant writing. To accomplish the proposed research and training, I have assembled a multi-disciplinary team of world class mentors who are committed to my success. This training will build on my previous experience in clinical and translational research as a trainee and ultimately provide me with the knowledge and skillset to establish an independent research program and transition into an independent R01- funded investigator leading global progress in understanding and exploiting neuroplasticity after SCI.

NIH Research Projects · FY 2025 · 2022-09

PROJECT ABSTRACT This focused technology research and development project will deliver a new class of acoustic separation/en- richment tools for multiple biomedical research applications. Acoustic microfluidics has emerged as a key ena- bling technology in biology and medicine, providing unmatched capability for non-contact, label-free object ma- nipulation and analysis. The proposed microfluidic platform is based on a novel concept: a longitudinal standing bulk acoustic wave (LSBAW) subunit that controls micro- to nanoscale objects for functional separation and/or confinement. The patented LSBAW subunits are highly configurable, which allows arrays of repeated subunits to meet varying capacity and throughput needs, from monitoring/detection in small-volume (sub-µL) reaction chambers to high-throughput enrichment of rare species. Outcomes of this project will include purpose-built prototype systems for: (i) high-throughput enrichment/fractionation, (ii) process control at high capacity, and (iii) multiplexed analyses with real-time monitoring. To establish the versatility and utility of the LSBAW platform, different configurations will be validated in research applications of value to, for example, cancer biologists (rare cell enrichment), synthetic biochemists (antibody conjugate synthesis on ultrasound-confined reaction sub- strates), and microbiologists (monitoring/measurement of biological mechanisms in bacterial cells). The technol- ogy outcomes of this project will be relevant not only to those applications, but will be broadly applicable to any field that relies on separation, isolation, and enrichment. The project includes three Aims: Aim 1: Demonstrate scalability of LSBAW subunits for high-volume, high-throughput enrichment of rare species. Aim 2: Validate series configurations of LSBAW subunit arrays for high-capacity cell modification/labeling or custom biomolecule synthesis. Aim 3: Validate multiplexed configurations of LSBAW subunit arrays for quantification and/or detection of a target species or biological mechanism. Validation experiments will be used to rigorously assess capabilities that are relevant to specific applications. Use of standard models (e.g., microparticles as proxies for biological cells) or well-characterized biological sys- tems (e.g., commercial antibodies; standard mammalian cell lines, mixtures of cells, and microbes) will ensure consistency and reproducibility of results. In each application, success will be defined using quantitative perfor- mance criteria (e.g., throughput, capacity, specificity, sensitivity) and comparison with appropriate existing tools and methods. The team merges expertise in microfluidics, synthesis and characterization of imaging agents, microbiology, and rare cell isolation/analysis, with strong track records of technology development and deploy- ment. Completion of these aims will translate a novel acoustic microfluidics concept to a suite of powerful and broadly accessible research tools that will accelerate research in a multitude of biomedical research fields.

NIH Research Projects · FY 2025 · 2022-09

ABSTRACT Approximately 70% of breast cancers (BCs) are estrogen receptor (ER) positive (ER+) and human epidermal growth factor receptor 2 negative (HER2-). Endocrine therapy (ET) reduces recurrence risk and improves survival for many in this group. However, despite standard of care and adjuvant ET, over 20% of patients with ER+/HER2- BC experience metastatic recurrence in the years to come, and virtually all patients with metastatic disease eventually experience disease progression on ET due to intrinsic or acquired resistance mechanisms. Progression on ET, however, does not preclude continued responsiveness to alternate forms of ET, including those that combine therapies directed at ER and key signaling pathways that drive ET resistance. However, there are currently no biomarkers that can reliably identify which patients will benefit from ET-based approaches so that chemotherapy could be avoided or delayed. The PgR gene is highly regulated by ER at the RNA and protein level, and thus expression of PgR in ER+ BC would be indicative of the functional status of ER and associated predictive benefit from ET. We propose to evaluate the utility of positron emission tomography (PET) imaging with the PgR radiotracer, [18F]fluoro-furanyl-norprogesterone (FFNP) to predict response to ET-based therapies. In a recent phase II single-arm clinical trial, we demonstrated that FFNP-PET imaging, before and after a one-day estradiol (E2) challenge (ΔFFNP-PET), predicted response to ET with 100% sensitivity and 100% specificity in women with advanced ER+ BC. In this proposal, we will dissect the functional relationship between PgR and ER and its implications for ΔFFNP-PET as a predictive imaging biomarker of ER function for the full range of current and emerging ET-based approaches using patient-derived tumor xenografts (PDX) and genetically engineered models, interfacing with a clinical trial. We propose three Aims. In Aim 1, we will examine the impact of ESR1 gene mutations on ER-PgR crosstalk, PgR expression, and ΔFFNP-PET as an imaging biomarker of ER function in preclinical models. In Aim 2, we will evaluate the utility of ΔFFNP-PET in predicting response to single agent ET agents alone and in combination with targeted therapies in PDX models of ER+/HER2- BC. In Aim 3, we will interface with a clinical trial to examine the impact of tumor genomics on ΔFFNP-PET and its accuracy in predicting response to therapy in patients with metastatic ER+ HER2- breast cancer enrolled in a phase II trial of endocrine therapy in combination with the CDK4/6 inhibitor abemaciclib. Overall, this study aims to have a far-reaching and high impact on the implementation of precision medicine in identifying, stratifying, and predicting response to clinically available and novel SERDs alone and in combination with other targeted therapies in patients with advanced ER+/HER2- BC.

NIH Research Projects · FY 2026 · 2022-09

PROJECT SUMMARY/ABSTRACT Alzheimer’s disease (AD) is the most common cause of dementia, and also an age-related neurological disorder. AD not only causes severe distress for patients and caregivers, but it also becomes a major public health predicament. However, the mechanisms responsible for the pathogenesis of AD are still unclear, which is a major challenge for AD prevention and therapy. Increasing evidence suggests that dysfunctional and aging immune system may be a primary factor/inducer for the development of AD. Accumulated senescent T cells have been identified in both AD patients and in aged AD onset mice, but the causative relationship between the increased senescent T cells and AD development and progression is unknown. We recently discovered a novel suppressive mechanism that human Treg cells can induce responder naïve and effector T cell senescence. Senescent T cells exhibit active lipid metabolism and possess a unique senescence-associated secretory phenotype (SASP), producing high amounts of lipids and metabolites. Importantly, our more recent studies demonstrated that senescent T cells can promote the aggregation of amyloid precursor protein (APP), amyloid beta (Aβ) and Tau proteins in human neuronal cells. Therefore, an improved understanding of the molecular and cellular processes of senescent T cells in the pathogenesis of AD is urgently needed, which could lead to the development of novel and effective therapeutic strategies. The central hypotheses of this proposal are: 1) accumulated senescent T cells with excessive lipid metabolism promote the development and pathogenesis of AD; and 2) blockage of senescence in T cells via lipid reprogramming is a critical checkpoint to control AD pathologic processes and progression, which will provide a novel strategy for AD prevention and immunotherapy. Specific Aim 1 seeks to determine whether senescent T cells with lipid metabolism disorder are a critical driver for the pathogenesis of AD. We will dissect the causative role of the secretory lipid metabolites of senescent T cells in reprogramming functions of neuronal cells. We will also identify the molecular and metabolic signaling responsible for the functional changes in neurons induced by senescent T cells, resulting in neurodegeneration and AD development. Specific Aim 2 will propose complementary in vivo studies to identify the causative relationship between the accumulated senescent T cells and AD development and disease progression in a spontaneous senescence accelerated SAMP8 mouse model. We will then test our hypothesis and the novel concept that that reprogramming of T cell lipid metabolism to reverse T cell senescence is a novel strategy to prevent AD development and enhance efficacy for AD immunotherapy. A positive outcome of these studies should lead to novel strategies for metabolic control of T cell fate and function for AD prevention and immunotherapy.