Virginia Commonwealth University

NIH Research Projects · FY 2026 · 2022-12

PROJECT SUMMARY The objective of this project is to provide mechanistic information on the function and regulation of cardiac palmitoyl-proteome (palmitoylome) in health, and how changes in the palmitoylation status of key proteins in the heart contribute to disease development. Palmitoylation is a post-translational modification, where a fatty acyl chain, most often palmitoyl chain, is covalently linked to the thiol side chain of cysteine. This increase in hydrophobicity drives protein trafficking and changes their interaction with neighboring molecules. Palmitoylation is important for 2 reasons: (1) prevalence - ~20% of human proteins are palmitoylatable, (2) reversible - providing dynamic control of protein distribution and function. Recently, we globally purified palmitoylated proteins from human, dog and rat hearts, and used proteomic approach to identify 454 proteins forming a core 'cardiac palmitoylome'. Our study defined the scope of protein palmitoylation in the heart, and broadly characterized them as 'subcellular microdomain organizers'. We further identified 11 palmitoylating (DHHC) enzymes expressed in the heart. The current proposal is built upon these recent progresses, and addresses 2 main questions. First, given the numbers of known DHHC enzymes expressed in the heart (11) and substrates (454), how is the DHHC enzyme/substrate relationship determined in cardiac myocytes? Aim 1 will test the hypothesis that DHHC enzymes, as transmembrane proteins not freely mobile in cytosol, have their 'territories' demarcated in cardiac myocytes, and proteins within their territories are potential substrates. Second, what is the role of cardiac palmitoylome in disease development when dysregulated? We will use junctophilin-2 (JPH2) as a case study. JPH2 is the major jSR/PM tether. It requires palmitoylation of its cysteine side chains to strengthen the jSR/PM junctions. Genetic variants in JPH2 have been linked to cardiomyopathies, but the mechanisms underlying their pathogenicity are not clear. Aim 2 will investigate the role of palmitoylation in determining JPH2's distribution and functions in myocytes. Aim 3 will explore the possibility that some JPH2 genetic variants compromise palmitoylation, and this deficiency in JPH2 palmitoylation contributes to their pathogenicity in cardiomyopathies. Our research team combines 6 areas of expertise: (1) multiscale detection/quantification of protein palmitoylation, (2) quantitative proteomics, (3) high-resolution imaging/analysis, (4) bioinformatics/biostatistics, (5) modulation of CICR (Ca-induced Ca release) in cardiomyocytes, and (6) profiling/quantification of post-translational modifications. This combination allows us to probe the cardiac palmitoylome from single molecules to global proteome. We will also provide fundamental information on the structure and function of JPH2, which is critical for understanding why disease-related genetic variants in JPH2 are pathogenic. 1

NIH Research Projects · FY 2026 · 2022-12

Summary Studies have revealed the exciting promise of CRIPSR/Cas genome editing to excise provirus for HIV cure. However, a major barrier to their clinical application is how to deliver it to latently infected cells effectively and specifically in vivo. The overall objective of this proposal is to close this gap by developing a new lentivirus-like particle fusogenic resicle (LVLP-R) that will specifically deliver multiplexed Cas12a ribonucleoprotein (RNP) and mRNA via MS2 coat protein (MCP) to CD4-expressing cells in vivo for excision of HIV proviruses and coreceptor CCR5. This CD4-targeted LVLP-R (LVLP-R-CD4) will deliver Cas12a protein/mRNA and multiplexed guide RNAs (gRNAs) with an increased excision efficiency and reduced off-target potential due to lack of lentiviral reverse transcription and integration. We hypothesize that LVLP-R-CD4 with Cas12a RNP/mRNA can simultaneously excise HIV proviral DNA and CCR5 in vivo. This hypothesis is supported by our preliminary data and pre-existing reports on successful RNP/mRNA LVLP delivery and multiplex genome editing in vivo. In Aim I, we will optimize our established VSVG-pseudotyped LVLP-R (LVLP-R-V) delivery of Cas12a RNP/mRNA both in vitro and in vivo for HIV proviral and CCR5 excision. In Aim II, we will develop and optimize DARPin or HIV Env-mediated LVLP-R-CD4 that efficiently delivers Cas12a RNP/mRNA to human CD4 T cells for genome editing in vitro and in vivo. In Aim III, we will evaluate HIV proviral and CCR5 excision for an HIV cure using LVLP-R-CD4 delivery of multiplexed Cas12a RNP/mRNA in HIV-infected ART-suppressed humanized-BLT mice. We focus on targeting CD4 T cells as our primary test platform because (1) our group was the first to apply CRISPR/Cas genome editing to excise HIV proviral DNA in T cell; (2) CD4 T cells are the major latent reservoir cells of HIV infection; and (3) the research team in this proposal has extensive expertise in studying lentiviral delivery and HIV infection of CD4 T cells using various approaches, including the humanized-mouse model. This high-reward proposal will explore a new LVLP-R-CD4 delivery of advantaged LbCas12a RNP/mRNA to CD4 cells and excise HIV provirus and CCR5 as a novel cure strategy. Completion of the proposed studies will offer a novel tool to deliver genome editors to CD4 cells in vivo and may provide a new gene therapy approach to HIV and other T cell-related diseases.

NIH Research Projects · FY 2026 · 2022-11

Project summary: Lyme disease is the most common tick-borne disease in the northern hemisphere and its incidence is steadily increasing. The ability of the Lyme disease spirochetes to adapt to changing environmental conditions is dependent in part on regulation mediated by the secondary messenger molecule, cyclic-di-GMP (c-di-GMP). This proposal will identify the critical functional determinants of PlzA, the sole identified c-di-GMP binding protein produced by all Lyme disease spirochete isolates. Based on the recently determined atomic structure of PlzA which revealed that PlzA belongs to the unique xPilZ domain class of c-di-GMP binding proteins and data demonstrating that PlzA has RNA chaperone activities, we will perform a comprehensive analysis to dissect structure-function relationships of both apo and holo forms of the protein. Based on comparative sequence and structural analyses, surface-exposed amino acid residues will be targeted for site-directed mutagenesis. The impact of these mutations on PlzA structure, c-di-GMP binding, RNA winding, RNA unwinding, and protein-protein interactions will be assessed in vitro. Based on these analyses, a series of B. burgdorferi transgenic strains in which wild-type plzA is replaced with plzA genes that encode PlzA proteins with altered activity will be generated. The ability of each strain to infect and transmit between ticks and mammals will be determined. These analyses will define the functional domains and biological mechanisms by which apo and holo PlzA regulate cellular processes required for the completion of the enzootic cycle.

NIH Research Projects · FY 2025 · 2022-11

Project Abstract. Medicaid expansion is a critical antipoverty tool that protects individuals from health care costs and reduces food and housing insecurity, significant contributors to poor health and health disparities. Medicaid expansion increases access to primary care, a hypothesized mechanism by which this public insurance coverage for low-income individuals reduces social needs. However, social needs and access to primary care are influenced by the sociocultural and built environments as well as health care system supply. This study applies the NIMHD’s Research Framework to understand how these domains of influence differentially shape access to primary care across those with or without social needs and in turn explores a possible mechanism in which Medicaid expansion reduces food and housing insecurity, primary care use. This work will leverage a well-established state-university partnership and unique data set consisting of the experiences of newly enrolled Medicaid expansion members in Virginia. The following existing data will be linked for this study by Medicaid member identification number: 1) pre-enrollment surveys asking members about their experiences in the year prior to Medicaid expansion enrollment (n=1,255), 2) a follow-up survey asking members about experiences in the year after enrollment (n=580), 3) all encounter and pharmacy claims data for these same Virginia Medicaid members, and 4) publicly-available measures of sociocultural, built environment, and health care system domains of influence linked to members’ census tracts or counties. The proposed research is supported by Virginia Medicaid as it aligns with the agency’s commitment to advance health equity through increasing primary care and reducing social needs. This proposed work aims to: 1) Assess how sociocultural environment, built environment, and health care system domains modify the effect of food and housing insecurity in the year prior to Medicaid expansion enrollment on primary care utilization in the year after enrollment, and 2) Explore whether primary care utilization reduces food and housing insecurity after the first year of enrollment in Medicaid expansion. Ultimately this work will provide much needed evidence to policymakers about the disparities in access to primary care across sociocultural and built environment factors as well as provide insight into how Medicaid expansion is reducing social needs. Addressing social needs through health care policies and primary care system design will improve population health and is necessary to achieve health equity. This application describes an excellent research and training environment for the proposed work, supported by a collaborative mentorship team that includes both faculty members and Virginia Medicaid’s Chief Medical Officer. This proposal will provide integrated, robust clinical and research training in health disparities, health policy, and geospatial statistics to a physician-scientist dedicated to practicing family medicine and pursuing a faculty position focused on advancing health equity through health policy.

NIH Research Projects · FY 2026 · 2022-11

Chronic alcohol consumption induces transcriptional changes in genes encoding structural components and regulators of the extracellular matrix (ECM). The molecular mechanisms underlying these alterations and the functional implications of altered ECM gene expression are not well understood in the context of excessive alcohol drinking. We hypothesize that activation of the immune response-related transcription factor STAT3 in astrocytes results in transcriptional changes in genes encoding ECM components, resulting in alterations in ECM structure and function, synaptic changes, and increased ethanol intake. The proposed experiments will address this hypothesis with three specific aims. In Specific Aim 1, we will use an unbiased approach to determine changes in the cortical ECM proteome, or “matrisome” in mice that have chronically consumed ethanol in a binge-like drinking protocol (drinking in the dark) for 6 weeks. Cutting-edge proteomics and glycomics will be performed to quantitatively measure changes in cortical ECM. These results will be compared to changes in ECM-related genes in existing and new INIA transcriptome datasets and the genes analyzed for STAT3 binding motifs in their promoters. In Specific Aim 2, we will determine the role of astrocyte-expressed STAT3 in alcohol consumption by conditionally knocking out Stat3 in astrocytes using floxed Stat3 mice and the astrocyte-specific Cre line, Aldh1l1-CreERT2. Mice will be tested for binge-like ethanol consumption and dependence-induced escalation of ethanol intake. Association of activated STAT3 with gene promoters in the prefrontal cortex will also be determined after chronic binge-like drinking using chromatin immunoprecipitation with an antibody to phosphorylated STAT3, followed by whole-genome DNA sequencing (ChIP-Seq). Finally, we will determine the role of a specific ECM gene and putative STAT3 target, brevican (Bcan) in alcohol consumption, by conditionally knocking out Bcan in astrocytes as described above. We will also reduce the expression of Bcan in the prefrontal cortex of mice using viral-delivered shRNA and measure binge-like ethanol intake. Brevican is localized perisynaptically and regulates synaptic plasticity. To determine if brevican is more highly associated with synapses after chronic ethanol drinking and if synapse structure in the PFC is altered by chronic ethanol, we will perform super-resolution microscopy with brevican antibody and antibodies to excitatory and inhibitory synaptic markers in mice after chronic binge-like drinking. Within each Specific Aim, we will integrate and collaborate with INIA-Neuroimmune and INIA-Stress investigators. The completion of these Specific Aims will contribute to fundamental knowledge of the molecular and cellular mechanisms by which chronic alcohol drinking alters the ECM and potentially provide new cellular targets to reduce excessive alcohol consumption.

NIH Research Projects · FY 2024 · 2022-09

Impact of a Novel Community-Based Biobehavioral Chronic Pain Team Training Program (4PCP) on Practitioner and Patient Outcomes ABSTRACT Although primary care medical practitioners (PCPs) provide the first and sometimes only line of treatment for patients with chronic pain, surveys show they are generally not comfortable with its management. This may reflect a dual shortcoming in their educational preparation. First the average medical school spends 0 to 6 hours on chronic pain, when pain organizations like the International Association for the Study of Pain (IASP) suggest 74 hours. Second, medical training is primarily oriented towards a biopharmacologic framework. While drugs may help the nociceptive component of pain, and anxiety or depression, they offer little to address the core emotional, cognitive and behavioral roots of chronic pain. This biopharmacologic mindset, that a drug might “do it all”, may have set a stage for today's opioid crisis. Managing these deeper chronic pain issues requires a biopsychosocial framework, that views a chronic pain syndrome as a brain disorder, reinforced by years of negative cognitive, emotional and behavioral habits, requiring retraining through physical & behavioral activation, and cognitive restructuring. However, since this framework and skillset are foreign to most PCPs, they remain ill-prepared to manage the totality of a chronic pain syndrome. In the 4PCP (Primary Practice Physician Program for Chronic Pain) pilot, we demonstrated that PCPs eagerly learned and implemented this new framework into their practice, increasing confidence with chronic pain management, reducing visit times, and improving patient pain. That study did not formally address opioids, though physicians tapered opioids spontaneously. We now ask 2 new questions: (1) Will a dissemination-ready, fully codified, intensive version of 4PCP produce better outcomes, sustained for 2 years? (2) Will practitioners find new confidence and skills in managing pain as a team and taper opioids? In 27 practices (4 practitioners each, 108 total) in 4 cities (Bozeman, Cleveland, Madison, Milwaukee) we will teach the new curriculum, 3 practices at a time every 2 months over 18 months, using a stepped cluster design. Practices will co-train with a paired psychologist and PT to build a clinical team. For each practitioner, we will identify 10 patients with chronic pain, ideally on opioids (1080 total) for patient outcomes. We will measure 3 types of outcomes before and for 2 years after the training intervention: (1) patient improvement; (2) the change in practitioner satisfaction with care rendered and practitioner burnout; (3) the change in opioid prescribing in the enrolled patients based on state records.

NIH Research Projects · FY 2025 · 2022-09

Hydrocephalus (HC) is a common and debilitating neurological condition affecting up to 1 in 500 individuals. Ventriculoperitoneal shunting (VPS) has been the standard treatment for >60 years but shunts remain highly problematic, with an unacceptably high malfunction rate and often lead to a lifetime of complex neurosurgical care. Endoscopic third ventriculostomy (ETV) is an alternative to shunting but has limited efficacy in infants. Adding choroid plexus cauterization (CPC) to ETV seems to be a viable alternative to shunting infants. Despite conflicting outcomes, ETV-CPC is currently being offered routinely and globally, and with limited understanding of the risks or benefits of this procedure on brain development. The lack of knowledge about both the surgical and physiological consequences of ETV-CPC constitutes a significant barrier to broad acceptance of this procedure. For example, the impact of ablating the choroid plexus, a major homeostatic organ which regulates neurogenesis and neurodevelopment, has yet to be determined. Experimental studies are needed to determine the role of ETV and ETV-CPC in clinical practice, but the lack of appropriate, validated large animal models in which to test these techniques on brain physiology and, critically, on brain development, has never been studied. Our hypothesis-driven proposal builds on our foundational work that developed a large, clinically-relevant animal model of HC in juvenile domestic pigs and demonstrated feasibility of ETV+CPC, and VPS, thus continuing the goals of NIH PA-180-623, “Tools to Enhance the Study of Prenatal and Pediatric Hydrocephalus”. Three Specific Aims will test our Central Hypothesis that, compared with VPS, ETV+CPC impairs brain development through disruption of VZ/SVZ and PVWM precursor biology and CSF homeostasis with pro-inflammatory signaling: Aim 1 - compare the cytological effects of VPS and ETV+CPC on VZ/SVZ and PVWM; Aim 2 - determine the effect of VPS and ETV+CPC on choroid plexus and CSF profile; Aim 3 - compare neurobehavioral and cognitive outcomes in HC animals treated with VPS and ETV+CPC. Supporting these aims are published and preliminary data showing cytopathology in the subventricular zone (SVZ) and periventricular white matter (PVWM) following treatment with VPS and ETV+CPC, ventricular zone (VZ) disruption, SVZ/PVWM cell death, glial activation, and heterotopia, PVWM glial activation and reduced oligodendrocyte progenitors, flow cytometry and single cell RNA sequencing differences in microglia and precursor lineages, CSF cellular and protein alterations, and reduced cognitive assessments. Analyses will be performed at 30- and 90-days post-treatment. Pigs were chosen for studies for their close homology to human neuroanatomy and physiology and the ability to use standard clinical neurosurgical techniques. Successful completion of the proposed studies will fill a critical void in HC research through rigorous testing of emerging surgical procedures and injury mechanisms, which is essential to identifying best neurosurgical practices for the clinical management of HC.

NIH Research Projects · FY 2025 · 2022-09

Project Summary/ Abstract Though synapses are formed between neurons, these structures are contacted with, ensheathed, and regulated by astrocytes. Period of neuronal synaptic connectivity and that of astrocyte maturation overlaps in developing brain. Neuronal signals instruct astrocyte differentiation and morphological maturation whereas astrocytes provide metabolic and trophic factors to support neuronal survival and growth. However, molecular mechanisms and signals that regulate neuron-astrocyte interactions and their role in neuronal circuit assembly and functions are largely unknown. We and others have previously shown that astrocytes modulate specific neural circuit formation, function and plasticity by several secreted proteins including hevin (SPARCL1), thrombospondins (TSPs), glypicans and norrin. While hevin is needed for assembly and plasticity of VGlut2+ (vesicular glutamate transporter 2) thalamocortical connections, TSPs facilitate VGlut1+ synapse formation. Intriguingly, expression of these synaptogenic proteins is developmentally regulated and are also altered in brain pathologies. Although a significant amount of research has been done to identify the neuronal receptors and mechanism of synapse formation by astrocyte-secreted synaptogenic factors SPARCL1 and TSPs, we do not know the signals and mechanisms that regulate their expression in astrocytes. We have recently found that neuronal contact stimulates expression of SPARCL1 and TSP4 via Sphingosine- 1-Phosphate (S1P)-S1P Receptor 1 (S1PR1). We also found that S1PR1 is primarily expressed by astrocytes and is localized to the fine astrocytic processes near and around the synapses and drives astrocyte morphological complexity and morphogenesis. Although, S1P-S1PR signaling is a drug target for many neurological disorders, its fundamental role in neuron-glia interactions and neuronal circuit assembly is not known. Our proposed studies will provide novel insight into the neuron-astrocyte bidirectional communication through S1P-S1PR1 axis in establishing synaptic connectivity and functions. Our detailed mechanistic studies will identify new signaling pathway downstream of S1P-S1PR1 axis in regulating calcium dynamics, glutamate sensing and expression of SPARCL1 and TSP4 in astrocytes. These studies will also advance our knowledge of how neurons regulate astrocyte development, morphogenesis and function. Moreover, these studies will decipher the mechanistic link between levels of S1P and the expression of SPARCL1 and TSP4 and clarify on the fundamental role of S1P/S1PR1 axis in the developing and diseased brain. This proposal thus is poised to provide novel mechanisms of targeting S1P/S1PR1 axis in alleviating neuropathologies.

NIH Research Projects · FY 2024 · 2022-09

Project Summary/ Abstract Alcohol Use Disorder (AUD) is a complex disease influenced by both genetic variability and multiple environmental factors. In the population, anxiety and stress are thought to be important drivers of alcohol consumption, with ethanol’s anxiolytic properties suggested as contributing to progressive increases in alcohol consumption and relapse. While there are foundations for genetic underpinnings of both AUD and anxiety disorders, there is little evidence to define and support the genetic variability in the mechanisms driving ethanol’s efficacy as an anxiolytic. The Miles laboratory has published a prior study, using recombinant inbred BXD mouse strains, demonstrating a robust quantitative trait locus (QTL) underlying acute ethanol anxiolysis [3]. From this QTL, Ninein (Nin) was implicated as a strong candidate quantitative trait gene driving variation in the anxiolytic- like response to ethanol in the light-dark transition model of anxiety [7]. Ninein is a microtubule associated protein (MAP) located in the centrosome and cytoplasm that anchors the minus ends of microtubules and has been implicated in axonal growth and branching [6] Ninein has been shown to interact with Gsk3b, a known ethanol- responsive gene implicated in synaptic plasticity and neurotransmitter trafficking that the Miles laboratory has shown to modulate ethanol consumption [28]. Alternative splicing variants and exon-specific associated changes in microtubule dynamics in neurons implicate that variability in Nin genomic sequence could drive genetic variation in behavioral responses to anxiety and ethanol. This project will examine the role of Nin and Nin transcript variant expression in basal anxiety, ethanol-induced anxiolytic-like activity and ethanol consumption using a regulated Nin knockout model, brain region selective viral vector gene delivery and CRISPR/Cas9 exon- skiping analysis. These objectives will be approached via the following specific aims: 1) Characterize role of region-specific Nin on ethanol-related behaviors in C57BL/6J (B6) mice using a tamoxifen induced knockout. 2) Characterize the role of alternative splicing of Ninein on ethanol and anxiety-related behaviors of C57BL/6J (B6) mice, using CRISPR/Cas9 viral vectors in exon-specific deletions.

NIH Research Projects · FY 2025 · 2022-09

Pathways for Authentic Teaching of Health Sciences (PATHS) is a five year project designed to build health literacy knowledge and skills. PATHS will provide teachers and students in rural, suburban, and urban Virginia communities with career awareness for the various educational trajectories and opportunities provided by health professions (Medical Lab Sciences, PT and OT, Radiation Science, and Dental Hygiene). PATHS will engage 60 secondary teachers (grades 7 through 12) over four years in a model program that provides curriculum and pedagogical support (professional development) reflecting best practices in science. Our aim is to support teachers and ultimately students in learning about the skills and content of health professions, the impact of social determinants of health (SDH), and to develop their ability to work with large data sets through medical tools related to precision health. The curriculum tools will be web-based modularized units to address five areas of health professions, SDH, and precision health and will provide content-rich and inquiry-based exposure to careers and skills necessary for health professions that are applicable to science, health/physical education, and career and technology education classrooms. Teachers will participate in two summer institutes with support during school-year follow-up sessions and virtual coaching. The project team along with the Virginia Area Health Education Centers (AHEC) will support teachers and their students with sharing their learning at Community Health Fairs, STEM Fairs, and other student research conferences.

NIH Research Projects · FY 2024 · 2022-09

PROJECT SUMMARY Bipolar disorder (BP) is a severe multifactorial neuropsychiatric disorder that imposes a significant burden on public health. The most recent large-scale genetic study of BP identified 64 associated genetic loci, providing initial insights in BP pathogenesis. Yet, genetic discovery in BP lags behind other key psychiatric disorders. The reported genetic loci only capture a small proportion of the total BP genetic liability, with many more variants across the common and rare allele frequency spectrum remaining to be discovered. In addition, the previous studied samples were of European ancestry, leaving population specific BP variants uncovered and uncertainty in how the BP genetic findings generalize to other populations, exacerbating health disparities, and these studies rarely employed “deep” phenotyping or assessed relevant environmental risk factors. This proposal brings together an international collaboration of leading investigators from the U.S., Taiwan, South Korea, Singapore, India, and Pakistan to form the Asian Bipolar Genetics Network (A-BIG-NET) and carry out a large-scale genetic study of BP in East and South Asia. A-BIG-NET will generate a BP genetic resource of 27,500 cases and 16,000 controls with rich phenotypic information, measures of key environmental stressors and genetic data from 4x low-pass whole genome sequencing (4xWGS). This will complement a schizophrenia genetics resource of 22,778 cases and 35,362 controls of Asian ancestry previously assembled by leaders of this network that will be available for cross-disorder comparisons. Studying BP genetics in Asia is important to the world and the U.S., as Asia constitutes 57% of the world population, and Asian American comprises 6.6% of the U.S. population (21.4 million). The five countries in A-BIG-NET cover 47% of all Asian populations. The specific aims of the proposal are to: 1) recruit and deeply phenotype 17,500 BP cases, with a focus on BP-I to maximize homogeneity, and 14,000 controls from four Asian countries; 2) carry out 4xWGS on all recruited samples plus 10,000 BP-I cases and 2,000 controls collected by a previous study using similar procedures in Pakistan; and 3) carry out a range of analyses to discover new genetic associations with BP-I across the allelic spectrum in East and South Asian populations, examine the comparative genetic architecture of BP-I across major world populations and with other major neuropsychiatric disorders, and perform a novel statistical fine-mapping analysis that leverages the multi-ancestry genomic diversity and pleiotropy across psychiatric disorders to identify putative causal variants. Aim 3 will also explore the genetic “validity” of various BP-I subtypes and fit models with joint genetic and environmental risk factors. This proposal will dramatically increase the worldwide diversity of genetics data on BP, an important step to accelerate gene discovery in this disorder and advance global mental health equity.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Bipolar disorder (BP) is a severe multifactorial neuropsychiatric disorder that imposes a significant burden on public health. The most recent large-scale genetic study of BP identified 64 associated genetic loci, providing initial insights in BP pathogenesis. Yet, genetic discovery in BP lags behind other key psychiatric disorders. The reported genetic loci only capture a small proportion of the total BP genetic liability, with many more variants across the common and rare allele frequency spectrum remaining to be discovered. In addition, the previous studied samples were of European ancestry, leaving population specific BP variants uncovered and uncertainty in how the BP genetic findings generalize to other populations, exacerbating health disparities, and these studies rarely employed “deep” phenotyping or assessed relevant environmental risk factors. This proposal brings together an international collaboration of leading investigators from the U.S., Taiwan, South Korea, Singapore, India, and Pakistan to form the Asian Bipolar Genetics Network (A-BIG-NET) and carry out a large-scale genetic study of BP in East and South Asia. A-BIG-NET will generate a BP genetic resource of 27,500 cases and 16,000 controls with rich phenotypic information, measures of key environmental stressors and genetic data from 4x low-pass whole genome sequencing (4xWGS). This will complement a schizophrenia genetics resource of 22,778 cases and 35,362 controls of Asian ancestry previously assembled by leaders of this network that will be available for cross-disorder comparisons. Studying BP genetics in Asia is important to the world and the U.S., as Asia constitutes 57% of the world population, and Asian American comprises 6.6% of the U.S. population (21.4 million). The five countries in A-BIG-NET cover 47% of all Asian populations. The specific aims of the proposal are to: 1) recruit and deeply phenotype 17,500 BP cases, with a focus on BP-I to maximize homogeneity, and 14,000 controls from four Asian countries; 2) carry out 4xWGS on all recruited samples plus 10,000 BP-I cases and 2,000 controls collected by a previous study using similar procedures in Pakistan; and 3) carry out a range of analyses to discover new genetic associations with BP-I across the allelic spectrum in East and South Asian populations, examine the comparative genetic architecture of BP-I across major world populations and with other major neuropsychiatric disorders, and perform a novel statistical fine-mapping analysis that leverages the multi-ancestry genomic diversity and pleiotropy across psychiatric disorders to identify putative causal variants. Aim 3 will also explore the genetic “validity” of various BP-I subtypes and fit models with joint genetic and environmental risk factors. This proposal will dramatically increase the worldwide diversity of genetics data on BP, an important step to accelerate gene discovery in this disorder and advance global mental health equity.

NIH Research Projects · FY 2026 · 2022-09

Abstract Myotonic dystrophy type-1 (DM1) is the most common form of muscular dystrophy in adults. The genetic basis is an expansion of CTG repeats in the non-coding region of DMPK, the gene encoding DM protein kinase. Individuals with myotonic dystrophy develop progressive muscle weakness, early cataracts, cardiac arrhythmias, and other symptoms. The disease mechanism involves a deleterious gain-of-function by the mutant DMPK mRNA, a process first described in DM1, known as RNA toxicity. RNA binding proteins become trapped on repetitive RNA, causing loss of splicing regulatory functions. Splicing changes contribute to DM1 symptoms and also may serve as biomarkers of disease severity. The discovery that DM1 is instigated by toxicity of one RNA species and characterized by misregulated splicing of other RNAs has furnished good therapeutic targets and candidate biomarkers. Several therapeutic approaches are under development and two are in early phase clinical trials. However, the design and conduct of clinical trials is limited by disease heterogeneity, scarcity of natural history data, and the lack of proven clinical endpoints or biomarkers of drug impact. We have begun a natural history study to define clinical endpoints, biomarkers, and patient characteristics for clinical trials. This study, END-DM1, has enrolled 277 participants but early progress was hampered by the COVID-19 pandemic. The current renewal application seeks to complete the study of clinical outcome assessments (Aim 1) and biomarkers (Aim 2) in DM1. We will complete enrollment of 700 adults with DM1 at 16 sites of the Myotonic Dystrophy Clinical Research Network with return visits at 12 and 24 months. Based on preliminary data, we selected a concise set of clinical measures showing acceptable reliability and responsivity to disease progression. The proposed study is designed to establish minimal clinically important differences for different measures in this population, identify baseline characteristics that predict future progression, and provide a rational basis for stratification, selection of sample size, or enrichment in future trials. Aim 2 will build on our previous efforts to develop RNA splicing biomarkers of DM1 severity and therapeutic response. This Aim is focused on tissue biomarkers that provide direct evidence of target engagement in skeletal muscle. We will assess a panel of DM1-affected splice events using a novel method that involves targeted high-throughput sequencing. Completion of this study is the logical next step to lay the groundwork for effective clinical trials in DM1, and keep pace with the rapidly expanding preclinical efforts to develop an effective drug treatment.

NIH Research Projects · FY 2025 · 2022-09

SUMMARY Substance use disorder afflicts nearly one in seven people in the US, placing a heavy burden on healthcare resources, the economy, and individual quality of life. Currently, there is no effective medication available for cocaine use disorder (CUD), most likely because the neurobiology underlying the disease is complex and not completely understood. Dopaminergic neurotransmission in the mesolimbic circuit has been shown to play a critical role in CUD. Cocaine inhibits the activity of the dopamine transporter (DAT) to produce its behavioral effects. Regulation of the DAT, a principal regulator of dopamine signaling, occurs primarily through post- translational modifications, such as phosphorylation, which are triggered by presynaptic auto and hetero- receptor-linked signaling cascades. These modifications alter the interaction of cocaine with the DAT, leading to changes in the stimulating and rewarding effects of cocaine. However, to date, post-translational modifications have not been a focus of investigation in the CUD field. Our studies show that phosphorylation of the Threonine-53 residue of DAT, or phospho-T53-DAT (PT53-DAT), plays a pivotal role in regulating cocaine- directed behaviors. In this proposal, we explore this novel finding using cutting-edge techniques, including a knock-in mouse model with a phosphorylation-defective DAT-Thr53Ala mutant (DAT-A53), a viral-mediated brain region-specific blockade of DAT-T53 phosphorylation, and fiber photometry-based measurements of dopamine in freely moving mice using the fluorescent sensor, dLight. With these new models and tailored biochemical, neurochemical, and behavioral studies, the current proposal aims to fill a critical gap in our understanding of the central role of DAT phosphorylation in normal dopamine neurotransmission and in the presence of cocaine, including in live animals for the first time. As a key mechanism, we have discovered that the kappa-opioid receptor (KOR) regulates DAT via PT53-DAT. Our studies also show that KOR activation increases DAT activity through PT53-DAT, and when T53 is substituted with A53, the aversive effects of a KOR agonist are attenuated. Here, we will expand and test our overarching hypothesis “Cocaine induces addiction- like behaviors when the KOR phosphorylates DAT at T53 and alters DA dynamics”. Aim 1 will investigate the effect of T53 phosphorylation on KOR-mediated DAT upregulation, trafficking and protein-protein interactions. Aim 1 will also examine if these PT53-DAT-dependent effects are sex-, and brain-region specific. Aim 2 will examine the impact of PT53-DAT on extracellular DA dynamics and DA release and clearance modulation by KOR and cocaine in vivo and in brain slices. Aim 3 will determine whether PT53-DAT plays a role in KOR- and cocaine- associated behaviors. Outcomes from the proposed studies will provide novel insights into the mechanisms of CUD, open new horizons for examining phosphorylation of DAT-T53 as an underlying mechanism of dynorphin actions in DA-signaling and cocaine addiction and expose new molecular therapeutic intervention points.

NIH Research Projects · FY 2025 · 2022-09

Hepatic decompensation (overt ascites, encephalopathy and variceal hemorrhage) and mortality in patients with nonalcoholic steatohepatitis (NASH) increases exponentially with the development of cirrhosis. Prevention of such liver-associated clinical events (LACE) is therefore a major goal of therapeutics. This requires identification of those at risk and targeting them with effective therapeutics. Current approaches to identify this population include the use of fibrosis markers such as FIB4, liver-stiffness measurement (LSM), assessment of hepatic venous pressure gradient and/or varices (endoscopy) and traditional measures of liver function. These are unfortunately limited by modest accuracy of some models, retrospective, single center nature of most data-sets published and lack of information on the impact of changes in these parameters over time with respect to dynamic changes in risk profile. This proposal innovates by novel application and integration of systemic, hepatic perfusion and function, and portal hypertension related parameters to holistically model the risk of LACE and generate a probability score that is sensitive to change. We will specifically focus on MRI-based measures of systemic metabolic dysfunction (metabo-phenotype), gadoxetate uptake and clearance (integrated measure of hepatic perfusion and clearance function) and spleen stiffness measurement (SSM) as a surrogate for portal hypertension. We further propose the novel hypothesis that holistic models including these measures and capturing the multi-factorial origin of hepatic decompensation events are superior to conventional tools used in routine practice to define the probability of LACE in patients with compensated cirrhosis due to NASH. The hypothesis will be tested in a longitudinal cohort study of compensated cirrhosis due to NASH. The specific measures of the metabo-phenotype will include visceral adipose tissue volume, fat-free muscle volume, muscle fat infiltration. Gadoxetate uptake and clearance will be measured in a 10-minute exam. The SSM probe was recently approved and will be deployed on Fibroscan 630 the flagship instrument for transient elastography. Metabo-phenotyping requires implementing an imaging protocol, based on a routine clinical imaging sequence, onto standard clinical scanners which adds less than 10 minutes of scan-time while gadoxetate is routinely used for liver-imaging. Sub-aim 1 will model baseline parameters, alone and in combination, to define outcome risk within 1-2 years. Sub-aim 2 will evaluate dynamic changes in values of the test measures over time and relate them to changes in the risk of outcomes. Both regression and machine learned approaches will be taken to generate probability scores of the outcomes. Sensitivity analyses will be performed to test the robustness of the models. The patient populations needed and all of the methods/expertise are available. The rationale of this proposal is that it will provide a method for risk-identification in those with NASH-cirrhosis which can be incorporated in to clinical care. This will have a strong positive impact by improving individual patient care, drug development and optimizing care delivery by identifying the population that will benefit most from therapeutics.

NIH Research Projects · FY 2026 · 2022-09

PROJECT SUMMARY/ABSTRACT Cognitive and behavioral inflexibility are key features of many severe psychiatric disorders. One particularly dangerous prototype of behavioral rigidity is evident in the relentless pursuit of weight loss in anorexia nervosa (AN), a disorder with alarmingly high morbidity and mortality rates. Prior efforts to intervene upon psychological inflexibility in AN have been ineffective. This may be due to inflexibility encompassing a broad set of cognitive and behavioral processes that can result from multiple distinct decision dysfunctions that require different interventions. Decision Neuroscience, which integrates translational and computational neuroscience with economics, offers a promising new paradigm for enhancing the precision of mechanistic science for psychiatric disorders. This approach posits that mental illnesses result from unique miscalculations performed by neurally- separable decision systems that can be probed with neuroeconomic paradigms. This approach holds promise for identifying precise treatment targets suited for different forms of psychiatric illness. Our pilot data derived from a Decision Neuroscience approach using a novel neuroeconomic paradigm (the Web-Surf task) have provided several insights into unique decision-making processes in AN. First, on this task, individuals with AN showed a distinct decision dysfunction suggesting overreliance on a rule-based decision system, reflected through rapid decision-making focused on maximizing long-term goals. This approach can lead to behavior that is inflexible and insensitive to current context. Second, this decision system appeared to be supported by hyperactivity within the frontoparietal cognitive control circuit (e.g., dorsolateral and ventrolateral prefrontal cortex), which is involved in long-term rule-following. Third, these biobehavioral patterns were linked to BMI and short-term weight-control behavior in AN. This frontoparietal-driven rule-based decision process differs from other decision-making patterns (e.g., fronto-dorsal striatal habit learning) that have been implicated in promoting behavioral rigidity in other psychiatric disorders, such as OCD. To extend this research to establish whether decision patterns distinguish AN from non-clinical and related clinical groups, we will recruit: a) AN; b) healthy comparison (HC); and (c) OCD (clinical comparison) groups. OCD represents a similar phenotype to AN hypothesized to be promoted through different decision processes. During fMRI, subjects will complete the Web-Surf task to identify neuroeconomic decision patterns (e.g., rule- based, reward-based, or habitual decision-making) in each group; these will be validated against other decision tasks. Participants will then provide data on symptoms via interviews and ecological momentary assessment at baseline and 12-month follow-up to examine the influence of distinct decision processes on AN and OCD symptoms. This study will evaluate the specificity with which AN is maintained by over-reliance on a frontoparietal rule-based decision system. This will permit first steps towards informing precision neuroscience- based psychiatric treatments informed by decision mechanisms, rather than solely by phenotypic presentation.

NIH Research Projects · FY 2025 · 2022-09

Current medication options for AUD have only partial success and have a low evidence base in cirrhosis. Gut dysbiosis, which may be initiated by alcohol, contributes to cirrhosis development. Our overarching goal is to develop a potent therapeutic targeting the intestinal microbiota to alleviate the impact of AUD and AROD through the gut-liver-brain axis. We base our approach on fecal microbiota transplant (FMT), originally developed for recurrent Clostridioides difficile infection (rCDI). We have also shown in randomized, placebo- controlled clinical trials the benefit and safety of different FMT formulations in cirrhosis. Furthermore, in a recent randomized trial of actively drinking AUD cirrhotic patients, we demonstrated that FMT reduced alcohol consumption and craving, and improved cognition and psychosocial health-related quality of life (HRQOL) versus placebo. We also found similar engraftment with capsule versus enema FMT. The FDA regulates FMT as a drug and a biologic. Our group uses Good Manufacturing Practices to manufacture FMT products in both liquid and freeze-dried, encapsulated formulations. Our hypothesis is that restructuring the gut microbiota using FMT will reduce alcohol consumption compared to placebo in patients with AUD and cirrhosis. To test this, we will conduct a Phase 1b/2a double-blind, placebo-controlled, randomized clinical trial using administration of a standardized oral encapsulated FMT preparation (FMT) at baseline and day 30 in patients with AUD and cirrhosis. Aim 1: Measure the effect of FMT on alcohol consumption. The primary outcome is number of abstinent days at three months in FMT compared to placebo. We will assess daily alcohol consumption and cravings using patient-reported measures and objective urinary and plasma markers. Aim 2: Determine the impact of FMT on safety and liver dysfunction. We will monitor safety outcomes and liver function throughout the trial. Aim 3: Determine the impact of FMT on microbial compositional and function. Comparative analyses of stool microbial composition, and serum metabolomics will be performed between and within groups. Targeted metabolomics will be focused on neuroactive metabolites that are produced or modulated by microbiota, e.g., SCFA, γ-aminobutyric acid (GABA), glutamate, indolic compounds, and bile acids. Aim 4: Determine the impact of FMT on brain dysfunction and patient-reported outcomes using cognitive testing and HRQOL testing. We will evaluate HRQOL and cognition using validated instruments (Sickness Impact Profile, EncephalApp Stroop, Psychometric Hepatic Encephalopathy Score). We will enroll 80 participants with AUD cirrhosis (randomized 1:1 to FMT versus placebo) under FDA IND. The team has access to patients, microbial expertise, infrastructure and AUD trial experience to carry out the trial.

NIH Research Projects · FY 2024 · 2022-09

Project Summary/Abstract Cognitive impairment is prevalent in older adults and females carry a disproportionately greater risk for cognitive deficits like dementia and Alzheimer's disease. While estrogen deficiencies have been linked to an increased risk of cognitive decline, the mechanism for this remains unclear. Evidence suggests that women living with HIV (WLH) are more likely to experience an earlier onset of menopause, thereby reducing the circulating estrogen levels. Additionally, within the population of people living with HIV (PLWH), women are at a higher risk for developing cognitive decline which tracks with data seen in the general population of people living without HIV. Thanks to the advancement of HIV treatments, PLWH are living longer, adding to the urgency to identify mechanisms underlying the acceleration of cognitive decline. The goal of this proposal is to determine the extent to which estrogen receptor expression and function predict cognitive decline in WLH. Leveraging the framework of the Atlanta Women's Interagency HIV Study (WIHS) cohort and the Emory SCORE U54, this project will test the hypothesis that HIV serostatus interacts with menopause to alter estrogen receptor subtype expression and that changes to estrogen receptor β are predictive of inflammatory and cognitive outcomes. Aim 1 will focus on the characterization of estrogen receptor subtype protein expression in PBMCs to establish a metric for understanding the expression of the two prominent estrogen receptor subtypes (α and β) and how they differ in the context of menopause and HIV. Aim 2 will establish inflammatory and cognitive profiles for the study participants using longitudinal timepoints to illuminate changes in both inflammatory and cognitive states that can be tracked over time. Data resulting from the evaluation of estrogen receptor subtype expression in the context of menopausal status and HIV status, coupled with the cognitive data available through WIHS will enrich our understanding of the role of estrogen receptors in predicting cognitive decline, and potentially inform biomarker selection for screening women who are at higher risk of developing cognitive impairment, especially WLH. This research and training will be performed with Dr. Gretchen N. Neigh, Ph.D., at Virginia Commonwealth University. The training provided will build upon the applicant's technical, computational, and molecular skills, in addition to providing targeted professional development opportunities, and further training in hypothesis-driven project design. Ultimately, this training will prepare the applicant for a future as an independent physician-scientist, investigating factors that impact cognition with aging.

NIH Research Projects · FY 2025 · 2022-09

Since the National Partnership to Improve Dementia Care debuted in 2012 with the goal to reduce antipsychotic use in individuals with Alzheimer’s disease and/or Alzheimer’s-related dementias (AD/ADRD), CMS has scrutinized almost all associated long-stay psychoactive prescribing. While the long-stay use of these monitored psychoactive drugs has declined, prescriptions of mood-stabilizing antiepileptic drugs (AEDs) have increased. Similar to antipsychotics, mood stabilizing AEDs are used to treat the behavioral and psychological symptoms of dementia (BPSD) in the AD/ADRD population. Unlike antipsychotics and other psychoactive medications, AEDs prescribed in nursing homes are not mandatorily reported to CMS. Pilot studies suggest AED increases are concentrated entirely in AD/ADRD and other dementia patients with no diagnosis of epilepsy and purposefully prescribed for BPSD as an unmonitored alternative to antipsychotics. AEDs are not approved to treat AD/ADRD or any dementia symptoms, have weak efficacy evidence, and convey serious risk. Increasingly, the Partnership’s debut seems an inflection point where the trend toward unmonitored alternative drugs for AD/ADRD symptoms sharply increased. Furthermore, initial findings hint that the COVID pandemic represents a second critical point of inflection where the existing transition toward non-superior but unreported drugs for AD/ADRD is again rapidly accelerating. All outcomes associated with this evolving pre-scribing phenomenon remain unknown, though early data warns that harms may be increasing without benefit. We propose a retrospective analysis detailing long-stay prescribing of all psychoactives, including AEDs, for AD/ADRD and other conditions, how such prescribing is changing under the combined pressures of the pandemic and targeted reduction efforts, and the role of such prescribing in adverse health outcomes. We will accomplish this by creating a detailed dataset of quarterly cohorts of nursing home residents, including those with AD/ADRD, for the years 2009-2021 that draws on elements linked at an individual level from CMS’s Minimum Data Set (MDS), Part D, MedPAR, and Public Use files. The final linked dataset will include health, demographic, cognitive, and diagnostic variables, all psychoactive drug claims, nursing home outcomes, records of hospital and ER adverse events, and long-stay facility details, including staffing resources and COVID statistics. Finally, to help explain and validate findings, a national sampling of nursing home prescribers will be surveyed regarding their knowledge and beliefs surrounding changing approaches to the care of AD/ ADRD and associated outcomes. This study will describe current and evolving AED use in nursing homes for all indications, focusing on AD/ADRD and AD/ADRD outcomes. It will describe racial and gender disparities in the long-stay populations treated or not treated with psychoactives. It will delineate the impact of the COVID pandemic in combination with national policies on ADRD management and outcomes. Results of this study will inform policymakers, improve nursing home care and safety, and introduce new avenues for ongoing research.

NIH Research Projects · FY 2026 · 2022-09

PROJECT SUMMARY Diabetic retinopathy (DR) is the most common cause of severe vision loss in the working-age population in the world. However, clinical studies have shown that approximately 40% DR patients do not adequately respond to anti-VEGF drugs. There is no drug treatment for those 40% DR patients. Furthermore, repeated bolus injections of these anti-VEGF agents are associated with risks of injection-associated side effects, and are a big burden to patients and the healthcare system. Fenofibrate, a peroxisome proliferator- activated receptor-α (PPARα) agonist, is the first low-cost and safe oral drug for DR with clinically proven efficacy against retinal neovascularization and diabetic macular edema in diabetic patients. However, oral fenofibrate has a short half-life and poor retinal absorption. An ocular fenofibrate delivery is needed for the treatment of DR. Intravitreal injection (IVT) of fenofibrate free drug demonstrated a short half-life of the drug in the eye, and frequent IVT injections of fenofibrate are needed to sustain the effect. We propose a new drug delivery strategy for providing sustained intraocular fenofibrate to target a non-VEGF, PPARα pathway and reducing the treatment burden from DR patients. Recently, we developed small-sized 200nm biodegradable nanoparticles containing 6% fenofibrate (Feno-NP) that demonstrated drug release for at least 8 weeks in the eye following an IVT injection, and have no detectable ocular toxicities. Our preliminary results further demonstrated that a single IVT injection of Feno-NP was effective on DR in the streptozotocin (STZ)-induced T1D rat model for 8 weeks. Here, our aim is to develop and optimize new longer-lasting large-sized fenofibrate- loaded microparticles (Feno-MP) that have higher drug loadings (≥20%) with dense PEG coating, and can safely and effectively treat DR, for ≥ 6 months with a single IVT injection. If successful, this approach would significantly improve DR patient care. In Aim 1, we will optimize and characterize reproducible large-sized Feno-MP formulations that release fenofibrate for ≥6 months, determine the maximum tolerated dose, and assess ocular pharmacokinetics (PK). The two most promising Feno-MP formulations will then be tested for dose dependent efficacy on retinal vascular leakage and inflammation in STZ-induced diabetic rats (T1D model) and db/db mice (T2D model) in Aim 2. Finally, we will carry out detailed safety and ocular PK studies of one lead Feno-MP formulation in rabbits in Aim 3 to facilitate future development and potential clinical translation.

NIH Research Projects · FY 2025 · 2022-09

Abstract Peripheral membrane proteins (PMPs) represent crucial mediators of biological and disease processes. This class of proteins exists in a water-soluble state until targeted to adhere on membranes, enabling them to perform their function. As with any membrane associated protein, PMPs are challenging to study, particularly in their membrane bound state, leaving many basic questions about function unresolved. Additionally, utilizing PMPs as drug targets is difficult since current methods are designed for water-soluble proteins and do not work well for membrane associated proteins. Thus, there is a great need for novel tools and procedures to illuminate details of PMP function and to create PMP inhibitors for use in chemical biology study and as drug leads. The goal of our research is to enable high-resolution, quantitative study of PMP interactions and allow inhibitor design for this elusive type of protein. We will initially focus our efforts on three PMPs of extraordinary biomedical interest: glutathione peroxidase 4 (GPx4) and Phox homology (PX) domains in the NADPH oxidase family (p47phox-PX and NOXO1-PX). We have initiated development of a novel membrane model, membrane- mimicking reverse micelles (mmRMs), which is based on the chemistry of cellular membranes. PMPs embed into mmRMs as they do with cellular membranes, enabling high-resolution study using NMR spectroscopy and other techniques. mmRMs have a number of advantages over current models, including greatly enhanced stability, outstanding spectroscopic properties, and an ability to house high concentrations of analyte along with the protein. We will modify our mmRM system to better reflect a variety of cellular and organelle membranes, allowing the system to be tuned according to the natural PMP environment. Our focus will then be to harness the unique properties of our mmRMs to enable unrivaled detail in study and quantification of PMP interactions with membranes and lipid substrates. To facilitate inhibitor design, we will develop a novel method that allows fragment screening of membrane-embedded PMPs, for which current methods are not suitable. Using these tools, we will initiate an inhibitor design campaign for our important PMP targets. Overall, our goal is to develop tools that will enable breakthroughs in detailed study of PMP biology as well as inhibitor and drug development for this largely untapped class of proteins.

NIH Research Projects · FY 2025 · 2022-09

Periodontal Disease (PD) continues to remain a major public health burden in the United States. Manifestation and progression of PD are multifactorial, and may vary across sex, with/without additional comorbidities, such as Type-2 Diabetes (T2D), where comorbid subjects are at an elevated risk of compromised oral health. There is an overall paucity of clinically interpretable and nationally representative cross-sectional summaries of numerous risk factors (and their complex interactions) in assessing multi-comorbidity aspects (here, PD and T2D), and precise estimation of associated causal treatments for PD in practice-based settings, factoring in sex influences. Publicly available nationwide survey databases (such as the NHANES), and large oral health databases (such as the HealthPartners®, HP) are important, but somewhat under-utilized resources for such evaluations and practical interpretations, mainly due to several unique statistical and epidemiological complexities, which are often beyond the capabilities of existing standard analytical tools and software packages. Furthermore, how to prioritize patients for oral clinic visits based on their sex determinants, and multi-comorbidity risks continues to remain unresolved. In this project, we address these challenges, and initially propose a stochastically-principled, nationally meaningful, summary risk index (Aim 1) representing cross-sectional PD association from about 11,700 adult dentate subjects, who are part of the NHANES 2009-2014 study, for the 4 target groups: (a) Males with T2D, (b) Males without T2D, (c) Females with T2D, and (d) Females, without T2D. We then refine and validate this derived index, and propose a time-varying PD index (Aim 2) for the four target subgroups, accommodating causality of periodontal treatment effects, via application to the rich, longitudinal, observational HP database of about 25,000 subjects in a practice-based setting, with further model fitting and cross-validation using the Kaiser Permanente Northwest database of about 1,17,000 subjects with similar characteristics. Next, we utilize the time-varying index to construct an optimal policy (Aim 3) for prioritizing highrisk patients for quicker clinic visits. Finally, we produce a free, interactive, web-application tool (Aim 4) via R Shiny, for estimation and computation of the personalized index and recall decisions for any future patient. Our statistically principled, comprehensive, unique index for PD integrating electronic medical records from two large HMOs will be the first of its kind to generate new knowledge in regards to assessing sex influences, in light of T2D. Furthermore, the proposed methodology is readily generalizable to other comorbidities, such as cardiovascular disease, kidney and liver disease, etc. In the longer term, pending rigorous model validation, the derived index has the potential to be integrated into popular chairside software, such as Patterson’s EagleSoft®, thereby facilitating efficient bench to bedside translation.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Hepatocellular carcinoma (HCC) is the third leading cause of cancer related deaths worldwide and its incidence is increasing due to endemic obesity. Sexual dimorphism exists in HCC incidence as women have a significantly lower risk for developing HCC than men. However, the molecular mechanisms of HCC sexual dimorphism remain unclear, hindering development of better therapies for this disease. This proposal will utilize a new model that we developed of diet-induced progression of NASH to HCC that recapitulates key physiological, metabolic, histologic and transcriptomic changes observed in the human disease to examine previously unrecognized roles of sphingosine kinase 2 (SphK2), an enzyme that regulates the balance of bioactive sphingolipid metabolites, sphingosine-1-phosphate (S1P) and ceramide, in sexual dimorphism of HCC. Similar to humans, we found that on this diet, wild-type male mice, but not females, develop HCC. Strikingly, SphK2 knockout male mice have reduced tumor incidence, whereas in females, liver cancer developed only in SphK2 knockout mice. Thus, we propose that SphK2 plays a critical role in sexual dimorphism of HCC. We will test the central hypothesis that SphK2 protects females from HCC, while promoting it in males through several mutually non-exclusive mechanisms in distinct hepatic subcellular organelles. Aim 1 will examine the role of liver SphK2 in key signaling pathways that promote HCC in males and Aim 2 will define the role of SphK2 in nuclear and mitochondrial mechanisms mediating resistance of females to HCC. Our proposal utilizes a unique animal model and state-of-the-art techniques to identify novel SphK2-regulated molecular mechanisms involved in sexual disparity in diet-induced progression of NASH to HCC, and also has translational implications for the use of SphK2 inhibitors now in clinical trials. This study will lead to better understanding of sex differences in HCC important for personalized treatment strategies for this devastating disease.

NIH Research Projects · FY 2024 · 2022-08

Abstract The improvement in breast cancer survival is partially due to improvements in treatments. However, these treatments are not without mild to life-threatening side effects. For example, anthracycline chemotherapies (e.g. doxorubicin) and human epidermal growth factor receptor-2 (HER-2) targeted therapies have been shown to cause cardiotoxicity (CTX), or damage to the heart. This side effect is of great concern as survivors are more likely to die of heart disease than any other diagnosis and racial disparities exist with regard to prevalence of CTX and risk of heart-related mortality. Factors that have been shown to contribute to cardiotoxicity include Black race and the presence of comorbidities. While the cardiovascular disease literature includes the roles of psychosocial and sociocultural factors in its prevalence and the racial disparity in its manifestation and progression, these factors are unexplored in the context CTX. Additionally, cancer care delivery factors, including maintenance of CTX, deserves attention as there may be opportunities to address gaps or differences in care that may exacerbate the CTX disparity. Addressing psychosocial and sociocultural factors may provide intervention targets to mitigate CTX disparities. Guided by the Social Ecological Model, I will conduct retrospective analyses and a historical-prospective cohort study to identify factors related to the racial disparity in CTX. During the K99 mentored phase, I will abstract electronic hospital data on breast cancer survivors treated with anthracycline chemotherapy or Her-2 targeted therapies to elucidate sociodemographic, clinical, and maintenance factors that relate to onset of and racial differences in CTX (Aim 1). In Aim 2, I will conduct in-depth interviews with Black and White survivors (n=20) with and without CTX to understand psychosocial and sociocultural experiences that may contribute to CTX. During the R00 phase (Aim 3), I will use the information learned from the in-depth interviews to inform a survey that will be distributed to n=150 Black and White breast cancer survivors who were treated with adjuvant therapies. This survey, coupled with medical record data, will be used to assess relationships between psychosocial and socio-cultural factors and racial differences in CTX. In order to successfully execute these aims the candidate will receive strategic planning in cancer care delivery, cardio-oncology, and mixed methods research. These training aims will ensure the candidate's goal of successfully transitioning to independence and establishing a research program involving the development of interventions that will address racial disparities in cancer care delivery and outcomes. The expertise and guidance of the mentoring and advisory teams in addition to the wealth of resources and commitment to training provided through the institutional environment will ensure the success of the candidate's research project and transition to independence.

NIH Research Projects · FY 2026 · 2022-08

Injection drug use increases the probability of contracting HIV, and opioid use disorder (OUD) accelerates HIV- 1 infection through immune suppression and direct CNS actions. Glutamatergic excitotoxicity is a major factor in HIV-dependent CNS injury, but emerging evidence also suggests a loss of inhibitory GABAergic function in neuroHIV. The parallel loss of Cl− homeostasis and GABAergic tone will worsen excitatory outcomes since ‘disinhibition’ results in net excitation. If this is true, then interventions that protect inhibitory systems are predicted to at least partially negate ‘excitotoxic’ effects of HIV. KCC2 is the main transporter responsible for maintaining [Cl−]i homeostasis and GABAergic function in the adult CNS and is the focus of proposed studies. Although the idea of disinhibition as a driving force is embraced for other neurological disorders (e.g., autism, certain epilepsies, opioid dependence, traumatic brain/spinal cord injury), it represents a conceptual shift about mechanisms underlying synaptodendritic dysfunction in neuroHIV. The Cl− concentration inside neurons ([Cl−]i) is small and the Cl− reversal potential (ECl) is close to the resting membrane potential. Thus, minor changes in [Cl−]i can greatly affect the strength and polarity of inhibitory (e.g., GABAA) transmission. NKCC1 (Cl− uptake) and KCC2 (Cl− efflux) co-transporters are key regulators of [Cl−]i, and KCC2 expression/function is essential for adult neuron survival. Their balance can be regulated by cytokines/trophic factors (e.g., BDNF) from glia, therapeutically (CLP290), genetically, and by opioids. Importantly, we find that increasing KCC2 levels/function strongly protects against exposure to Tat, gp120, infectious HIV, and opioids in human and mouse neurons. In vivo, maintaining KCC2 phosphorylation can normalize KCC2 localization in Drd2-expressing striatal medium spiny neuron (MSN) cell membranes and reverse motor deficits due to HIV-1 Tat. Aim 1 uses in vitro models including iPSC MSNs, infective HIV, and optical electrophysiology to identify mechanisms by which HIV and/or opioids alter [Cl−]i homeostasis, dysregulate D1/D2 MSN excitability, how this triggers synaptodendritic injury, and protective strategies. Despite its fundamental importance, the role of KCC2 in excitatory/inhibitory (E-I) imbalances and altered ECl and EGABA have never been explored in the context of HIV/OUD. Aim 2 extends the studies in vivo/ex vivo. 2 transgenic models (Tat+/-, HIVTg26) that both mimic clinical pathology with considerable fidelity are crossed with transgenic mice expressing Drd1a-tdTomato (D1) and Drd2-eGFP (D2) to identify both striatal MSN populations. Acute (2 wk) and chronic (8 wk) HIV/Tat exposure times are examined; separate cohorts of mice (both sexes) receive concurrent, ramping exposure to morphine (s.c.). Comprehensive studies of [Cl−]i regulation in MSNs (gramicidin-perforated patch physiology), synaptodendritic injury, and behavior related to striatal function are performed ± the KCC2 enhancer prodrug CLP290 as an intervention strategy. Alternative [Cl−]i regulation through NKCC1, TMEM16A, and CLC-1 are also tested. GCaMP8f expression in D1/D2 MSNs in awake, behaving mice links striatal output activity to behavioral change (Inscopix miniscopes).