University Of Alabama At Birmingham

NIH Research Projects · FY 2025 · 2024-03

Project Summary: Amphetamines (AMPHs) are a class of psychostimulants that includes compounds commonly used for the treatment of neuropsychiatric disorders (e.g. attention deficit disorders) as well as drugs of abuse. The abuse liability of AMPHs stems from their capacity to elevate synaptic levels of dopamine (DA) and activate natural reward pathways in the brain. AMPH does this, at least in part, via reversal of the dopamine transporter (DAT), a membrane protein physiologically responsible for import of DA into presynaptic neurons. Reversal of DAT leads to mobilization of cytoplasmic DA through non-vesicular DA release (DA efflux). The psychomotor stimulant properties of AMPH, as well as induction of reward, are dependent upon this efflux. In fact, inhibition of DA efflux reduces both the ability of AMPH to increase locomotion as well as AMPH preference. A growing amount of data shows reciprocal communication between the microbiome and the brain. Fusobacterium nucleatum (F. nucleatum) is a member of the human microbiome that enhances behavioral responses to AMPH by secreting butyrate, inhibiting histone deacetylase complex activity, and increasing expression of the dopamine transporter. Interestingly, not only does F. nucleatum enhance host responses to AMPH, AMPH increases abundance of F. nucleatum in both rodents, and humans, suggesting a feedforward paradigm that reinforces both imbalances in the microbiome (dysbiosis) as well as amphetamine use disorders. How AMPH enhances the abundance of F. nucleatum is not known. F. nucleatum is a gram-negative anaerobic rod that uses biofilm formation with aerobic or facultative partners to generate a protective barrier that stabilizes F. nucleatum colonies in the host. Our data demonstrates that AMPH enhances biofilm formation between F. nucleatum and the facultative bacteria Streptococcus mutans (S. mutans). Of note, F. nucleatum and S. mutans are both bacterial species that are highly associated with dental caries and gingivitis, periodontal diseases commonly comorbid with AMPH use disorders. Interestingly, biofilm formation in S. mutans is dependent upon glucosyl transferases (Gtfs), enzymes that aid in assembly and organization of the extracellular matrix of biofilms, and AMPH enhances expression of these Gtfs. We hypothesize that Gtfs, expressed by S. mutans, are required for formation of F. nucleatum:S. mutans mixed species biofilms, increases in abundance of F. nucleatum, and enhancement of host responses to AMPH. We test this hypothesis in the following aims: S.A. #1: Determine how Gtf activity regulates AMPH-enhanced biofilm formation, between F. nucleatum and S. mutans, and host colonization. S.A. #2: Define how Gtf activity alters AMPH-induced behavioral responses.

NIH Research Projects · FY 2026 · 2024-03

Post-translational modifications play a crucial role in cell proliferation, differentiation, and tissue development. SUMOylation is a post-translational modification that alters stability, activity, protein interactions and cellular distribution of target proteins. The sequential process of SUMOylation requires three enzymes. The small ubiquitin-related modifiers (Sumo) are initially activated by the E1 enzyme and transferred from E1 to Ubc9, the only E2 conjugating enzyme. Finally, the E3 ligases transfer Sumo from E2 to the target protein. Abnormal SUMOylation has been associated with cancers, neurodegenerative and skeletal disorders, including rheumatoid arthritis and Paget’s disease of bone. SUMOylation occurs during embryonic development but the functional role of SUMOylation during skeletogenesis remains unknown. The development of skeletal tissue in mice is first noted at embryonic day 11.5 and increases through embryonic and postnatal life. We found that the level of Runx2 increased by 13-fold from E12 to birth. A similar progressive expression was noted for Sp7 gene. Sumo1 expression was progressively decreased from E12 to newborn stage. The vital substrate Sumo2, however showed a constitutive expression throughout embryonic skeletal development. These data indicate the availability of Sumo substrate during skeletogenesis. The evolutionarily conserved Ube2i gene encodes the only conjugating enzyme in vertebrates, Ubc9, that is required for SUMOylation. Our data shows that the levels of Ubc9 mRNA remain stable during embryonic skeletogenesis. Our mRNA sequencing confirmed the presence of all Sumo substrates, E1, E2 and critical E3 ligases in chondrocyte and during endochondral ossification. Based on these data, we hypothesize that Ubc9 is required for differentiation of chondrocytes during embryonic and post-natal skeletogenesis. In this study we aim to elucidate the role of Ubc9 mediated SUMOylation during endochondral ossification. In Aim 1 we will uncover the requirement of Ubc9 mediated SUMOylation during endochondral ossification. We will first establish the expression pattern of the of essential components of the SUMOylation pathway in the growth plate chondrocyte by RNA and protein analysis. To uncover how SUMOylation regulates chondrocyte differentiation and endochondral ossification, the Ubc9 gene will be deleted in resting chondrocytes using Col2a-Cre transgenic mice. In Aim 2, will investigate the role of SUMOylation in hypertrophic chondrocyte mediated cartilage turnover and bone formation by conditionally deleting Ubc9 in hypertrophic chondrocytes using Col10-Cre mice. Completion of this proposed study will contribute to the field by providing novel mouse models for studying SUMOylation during endochondral ossification. Additionally, elucidating the role of Ubc9 during chondrogenesis will provide critical knowledge about the role of SUMOylation during skeletal cell differentiation, which could lead to new therapeutics for many skeletal disorders.

NIH Research Projects · FY 2026 · 2024-03

PROJECT SUMMARY/ABSTRACT The overall goal of this 5-year K23 proposal is to support Camille Schneider Worthington, PhD, RD to become an independent clinical investigator with a research program focused on behavioral interventions for promoting maternal-child cardiometabolic health. The postpartum period is a critical time for intervening to improve maternal-child health. Existing behavioral postpartum weight management interventions have had modest success, particularly among women with overweight/obesity (Ow/Ob; body mass index ≥ 25 kg/m2). Timing of energy intake may contribute to weight gain and worsening metabolic health postpartum, but no studies have examined the effects of altering the timing of energy intake to manage postpartum weight and its metabolic consequences. Time-restricted eating, which is a form of intermittent fasting that involves restricting the daily ingestive period to ≤10 hours, has received considerable attention for promoting weight loss. Early time-restricted eating (eTRE) appears to be particularly beneficial as it is a form of time-restricted eating that involves shifting the majority of caloric intake earlier in the day to align with circadian rhythms in metabolism (i.e., chrononutrition). eTRE is a simple, focused dietary approach which may confer benefits behaviorally (e.g., improved adherence) and physiologically (e.g., improved insulin sensitivity) that are especially relevant postpartum. Yet, an eTRE intervention for improving cardiometabolic health postpartum has not been studied. To begin addressing this gap, Dr. Worthington received an institutional grant and is currently collecting dietary data and qualitative feedback from postpartum women to inform the development of a postpartum-adapted eTRE intervention (completion date: 6/2023). The proposed K23 will then be used to test the feasibility of the eTRE intervention among postpartum women with Ow/Ob. The proposed 2-arm pilot randomized trial (N=60) is rigorously designed to accomplish the following Specific Aims: Aim 1: Determine the feasibility and acceptability of a 12-week eTRE intervention among postpartum women with Ow/Ob. Aim 2: Examine the preliminary effects of the eTRE intervention on weight, body composition, and insulin sensitivity. Aim 3: Investigate the preliminary effects of the eTRE intervention on energy intake, appetite, and fatigue. Through this proposal, Dr. Worthington will accomplish the following training objectives: 1) Develop advanced knowledge and experience in behavioral intervention trial design, implementation, and adaptation; 2) Develop skills and experience in chrononutrition and underlying circadian and physiological mechanisms; 3) Develop advanced skills and hands-on experience in qualitative research methods; 4) Professional development and progress to research independence. This proposal represents a 5-year comprehensive mentoring, training, and research plan to transition the candidate to a successful independent clinical investigator. By the end of the award period, Dr. Worthington will have contributed novel data in the growing field of chrononutrition and maternal-child health and have submitted a competitive R01 to evaluate the clinical and cost-effectiveness of the refined postpartum eTRE intervention.

NIH Research Projects · FY 2025 · 2024-03

PROJECT SUMMARY/ABSTRACT The specific objective of this proposal is to demonstrate proof-of-concept for a novel screening approach for cervical cancer (CC) using a topically applied, disease-specific plasmid containing a secreted reporter. CC is the fourth most common cancer in women worldwide. Despite the availability of clinical screening methods and effective treatments for early stages of CC in the U.S., disproportionately high incidence and mortality due to CC is prevalent in rural Black communities. This disparity is due partly to poor follow-up resulting from socioeconomic barriers and poor access to clinical care in these communities. The long-term goal of this project is to mitigate these barriers by developing a sensitive, home-based screening approach for CC by means of a diagnostic plasmid. As a first step in achieving the long-term goal, this proposal will test the hypothesis that a topically applied, disease-specific plasmid, containing a secreted reporter protein driven by the Inhibition of Differentiation 1 (Id1) promotor, can be used to detect CC. Id1 has minimal expression in normal adult tissues but is overexpressed in CC, where expression correlates with the stage of disease. Topical application of the plasmid to the cervix would promote transfection via nonspecific endocytosis. Following transfection, CC cells that have upregulation of Id1, but not normal cervical cells, will secrete the reporter protein into the cervical cavity for non- invasive collection. The experimental approach will use CC cells, CC tissues freshly resected from human patients, and animal models to establish feasibility and specificity for CC screening with a plasmid containing the Id1 promotor and secreted reporter. Aim 1 will determine correlations between Id1 expression and reporter production in CC cell lines with varying degrees of Id1 expression and in fresh human CC tissues following transfection in vitro. Specificity of reporter production as a function of Id1 and due to the presence of CC relative to healthy cervical tissue will be evaluated. Aim 2 will establish feasibility for topical application of the plasmid in vivo using mice bearing human CC tumor xenografts. Plasmid-induced reporter production will be measured as a function of time and as a function of the number of transfected cells following topical transfection. The minimum tumor burden required for reporter-based detection in cervical fluids will be determined. Successful completion of the aims will demonstrate feasibility and specificity of the diagnostic plasmid approach in CC models. Long- term impact: This work will establish the foundation for advancing the plasmid-based approach into future stages of development using translationally relevant vectors and cervical applicator materials for plasmid delivery and reporter protein collection. Ultimately, we envision a diagnostic approach that in the long-term will enable highly specific, cost-effective, self-applied CC screening to facilitate targeted clinical follow up and treatment for individuals in high-risk settings associated with reduced access to routine gynecological care.

NIH Research Projects · FY 2025 · 2024-03

SUMMARY Non-alcoholic fatty liver disease (NAFLD) affects about 25% of the general population and is associated with other common conditions including kidney stone disease. The amount of oxalate excreted in urine is a significant risk factor for developing calcium oxalate kidney stones and may also play a role in the progression of chronic kidney disease. Previous studies have shown that about 50% of the urinary oxalate pool is derived from endogenous oxalate synthesis. Ascorbic acid (AscA) turnover and glyoxylate metabolism are important components of endogenous oxalate synthesis. A recent report demonstrated in human steatotic liver biopsies that mRNA levels of various genes regulating oxalate synthesis from glyoxylate, including alanine gloxylate aminotransferase (AGXT), are downregulated in NAFLD. Furthermore, urinary oxalate was positively associated with the severity of hepatic steatosis in overweight or obese children and adolescents with biopsy-proven NAFLD. Oxidative stress which is prevalent in those with NAFLD could promote the conversion of the anti- oxidant, AscA to oxalate. The turnover of AscA to oxalate may be heightened in NAFLD, as the progression of NAFLD is marked by increased inflammation and oxidative stress, as well as decreased plasma AscA. Thus, increased endogenous oxalate synthesis from both glyoxylate and AscA could be a major driver of kidney stone risk in those with NAFLD. This proposal will test the central hypothesis that endogenous oxalate synthesis increases with the severity of NAFLD in non-kidney stone forming adults. Leveraging our expertise in controlled dietary studies and utilization of carbon-13 precursors to study metabolism, as well as the large pool of patients with NAFLD cared for at UAB, we will test the mechanistic hypotheses that AscA turnover and the metabolism of glycolate (a glyoxylate precursor) to oxalate are increased in individuals with more severe forms of NAFLD. We expect this pilot and feasibility study will generate important preliminary data for a future multi- disciplinary/multi-institutional NIH R01 application focused on mechanisms underlying endogenous oxalate synthesis in calcium oxalate kidney stone formers with and without NAFLD.

NIH Research Projects · FY 2026 · 2024-02

Project Summary Parkinson’s disease (PD) is a systemic, progressive neurodegenerative movement disorder and type of synucleinopathy characterized by the presence of misfolded α-synuclein (αSyn) protein aggregates (Lewy bodies). The majority of PD etiology is not well explained by inherited genetic risk variants, and conversely, is strongly tied to exposure to environmental contaminants, particularly industrial byproducts such as pesticides, metals, and organic solvents. Data from our lab indicates that exposure to environmental toxicants associated with PD risk impairs the autophagy-lysosomal pathway (ALP), causing aberrant proteostasis and cellular dysfunction that influences disease development. Furthermore, new data suggests that the PD associated genetic risk factor and kinase protein LRRK2 plays a key role in lysosomal homeostasis, which implicates the lysosome as a convergence point for genetic and environmental risk factors in PD. The chlorinated organic solvent trichloroethylene (TCE) is associated with 2-6 times elevated risk for PD, however, the mechanisms by which this occurs remain vastly understudied considering the pervasiveness of its environmental contamination. We previously showed that exposure to TCE induced endolysosomal deficits, endogenous αSyn accumulation, and dopaminergic neurodegeneration in adult rodents. We also observed that TCE elevated the kinase activity of wildtype LRRK2 in the brain. In line with this, we have new preliminary evidence that TCE exposure causes loss of lysosomes, reduced proteolysis in dopaminergic neurons and significantly worsens αSyn preformed fibril (PFF) accumulation in brain tissue of adult rats. Thus, we hypothesize that an important mechanism of TCE is disruption of lysosomal function, leading to proteostasis impairment and neurotoxicity that triggers or facilitates PD development. This proposal is designed to uncover the mechanisms of TCE-induced lysosomal impairment, aberrant proteostasis, and dopaminergic neurodegeneration associated with PD risk. To do this, we will use our novel, environmentally relevant TCE inhalation model to measure how the solvent modulates αSyn PFF aggregation within the rat brain (Aim 1), and if loss of lysosomal function in central and peripheral neurons using newly developed Thy1-RFP-GFP-LC3 transgenic mice correlates with αSyn seed amplification signal measured via RT-QuIC (Aim 2). Last, we will assess how TCE-induced LRRK2 kinase activation influences lysosomal function and if pharmacological inhibition of LRRK2 prevents lysosomal impairments and neurodegeneration induced by TCE (Aim 3). Together, these experimental aims support a fundamental new mechanism in the risk for PD and related synucleinopathies from environmental factors and advances the NIEHS strategic plan to improve human health though environmental exposure research.

NIH Research Projects · FY 2025 · 2024-02

The direct cause of free sialic acid storage disorder is the genetic mutation in the lysosomal sialic acid transporter Sialin. Although the disease has been discovered for over 20 years, no cure has been developed. The long-term goal is to help create valuable therapeutics for patients with pathogenic Sialin mutations. The overall objective of this proposal is to establish a basic understanding of the membrane transporter. The rationale is that, with detailed knowledge about the structure-function relationship of Sialin, it will be possible to design small molecules to restore the substrate translocation of dysfunctional Sialin. To accomplish the goal, three specific aims will be pursued: 1) understand the substrate translocation mechanism of Sialin; 2) probe the connection between defective Sialin and cardiovascular diseases; and 3) restore the transporter function of pathogenic Sialin mutants. Specifically, in the first aim, biochemical assays, cryo-electron microscopy, and molecular dynamics simulation will reveal the molecular mechanism. In the second aim, Sialin will be characterized in human cardiac cells and a human induced pluripotent stem cell-based platform. In the third aim, small molecules to rescue dysfunctional Sialin will be identified by high-throughput screening and confirmed by secondary and tertiary assays. The research proposal is innovative because it focuses on finding a new strategy to rescue the normal function of pathogenic Sialin mutants that are directly connected with human diseases. This proposal is significant because it is expected to reveal the basic molecular mechanisms of Sialin and provide scientific justification for further development of drugs against various pathogenic Sialin mutations.

NIH Research Projects · FY 2022 · 2024-02

Contact PD/PI: Cingolani, Gino Project Summary Protein aggregation underlies several neurodegenerative diseases and is a pathological hallmark of Amyotrophic Lateral Sclerosis (ALS). In this devastating disease, cytoplasmic inclusions containing aggregated, often hyper post-translationally modified proteins are found in degenerating motor neurons and surrounding oligodendrocytes. There is no specific pharmacological treatment to prevent protein aggregation or promote disaggregation and clearance of existing aggregates that drive the progression of neurodegeneration. This exploratory R21 proposal builds upon the recent discovery that nuclear import factors of the importin β-superfamily can exert disaggregase activity toward other proteins. Combining the power of protein biochemistry and structural biology with our know-how in nucleocytoplasmic transport, we seek to learn from importins the biological principles for disaggregase activity and use this knowledge to design more versatile protein therapeutics. We believe that the high-risk, high-reward R21 funding mechanism will fuel the creative and diligent pursuit of answers to difficult biological questions, permitting our research program to achieve significant advancements in developing novel protein therapeutics that reduce pathogenic protein aggregation. Project Summary/Abstract Page 6

NIH Research Projects · FY 2026 · 2024-02

PROJECT SUMMARY/ABSTRACT Unprecedented clinical successes in past 12 years with immune checkpoint blockers (ICBs) such as anti- CTLA-4 and anti-PD-1 re-instigate strong interests in recent years to study the interplay between radiotherapy (RT) and the immune system, a long-forgone observation. Recent seminal efforts from many groups show that RT, by releasing danger signals and novel neoantigens, drives dendritic cell maturation that in turn cross prime T cells to mediate anti-tumor immune responses. However, T cell-intrinsic mechanisms in RT have been elusive, despite the indispensable role of T cells in governing RT efficacy. Moreover, by turning tumors into in situ “personalized” vaccines and debulking ICB-resistant large tumors independent of direct killing by IFN-γ, RT represents an ideal therapeutic option to overcome ICB resistance associated with tumor loss of IFN-γ signaling, a major mechanism of resistance to ICB that we and others recently identified. This study is logically built upon our preliminary data showing that our RT regimen activates mTOR and its downstream target, the hypoxia-inducible factor 1α (HIF1α) in tumor-infiltrating T cells (TILs); specific deletion of HIF1α in T cells abolishes therapeutic effects of RT. Importantly, although not as potent as in WT tumors, our RT regimen is still able to induce significant suppression of tumors lacking functional IFN-γ signaling. We therefore hypothesize that the mTOR-HIF1α axis in TILs is a major mechanism underscoring the therapeutic and immunological effects of RT, which can be utilized to overcome ICB resistance. In Aim 1, we will establish T cell-intrinsic mTOR-HIF1α axis controls RT efficacy. In Aim 2, we will determine cellular and molecular mechanisms by which activated HIF1α orchestrates anti-tumor immunity elicited by RT. In Aim 3, we will rigorously test our RT regimen, either alone or in conjunction with “targeted” chemotherapies and/or ICBs, in both human and murine melanomas lacking functional IFN-γ signaling, with an overall goal to overcome this major mechanism of ICB resistance. Our studies will reveal for the first time T cell-intrinsic mechanism(s) in RT. Since our tested compounds are clinically approved, our findings will facilitate a rapid bench to bedside translation. Furthermore, our rationally designed RT and RT+ICB regimens may guide future clinical trials.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY This NIH F31 grant application presents a comprehensive four-year plan designed to facilitate the professional development of Morgan Greene, the Principal Investigator, and equip her with the necessary skills to pursue an independent career as a physician-scientist. The research project's overarching goal is to unravel the transcriptomic programs driving the development of uterine natural killer (uNK) cells (Aim 1), which are essential for successful pregnancy establishment and maintenance. Additionally, the study will explore the factors that influence uNK cell differentiation into decidual natural killer (dNK) cells, a process critical to determining the pregnancy's health. Employing advanced techniques such as transcriptome-wide association studies, single- cell RNA sequencing, incubation assays, and flow-cytometry, the project's primary objective is to identify the transcriptional programs that underlie uNK cell differentiation from peripheral blood cells into dNK cells. The ultimate objective is to apply these findings to develop clinical tools that guide treatment strategies for pathological reproduction, infertility, and miscarriage, ultimately leading to improved reproductive health outcomes. The proposed training plan for Morgan Greene is supported by her project mentor, Dr. Paige Porrett, and co- sponsored by Dr. James George, and aims to establish a strong foundation for a successful career as a physician-scientist. The project-based translational approach offers an ideal training environment, with experiences designed to enhance Morgan's development in three key areas: 1) rigorous research in reproductive immunology, encompassing literature review, data evaluation, and responsible research conduct; 2) training in genetic sequencing and bioinformatics; and 3) career and professional development, including manuscript and grant writing, scientific communication, and translation of research outcomes to clinical applications. This proposal is designed to equip the PI with the advanced immunology skills necessary for a career focused on precision medicine and reproductive health outcomes as an obstetrics and gynecology physician-scientist, and drive the development of rigorous scientific research in reproductive immunology.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY/ABSTRACT This K01 Mentored Research Scientist Career Development Award will facilitate Dr. Jones’ long-term career goal of conducting translational research focusing on the vascular contribution to cognitive aging in vulnerable populations, namely individuals living with HIV who are at risk of cardiovascular disease and neurocognitive impairment, and the benefits of exercise interventions. As age and HIV have independent and synergistic effects on the brain, it places older people living with HIV (OPWH) at an increased risk of HIV-associated neurocognitive disorder and Alzheimer’s disease and related dementias. Accelerated vascular aging, assessed via arterial stiffness, is likely linked to those neurocognitive changes and exercise may be an effective, nonpharmacological strategy to counterbalance the adverse vascular effects contributing to neurocognitive disorders. The candidate is an exercise physiologist with a strong background in research on aging and vascular health and function. The proposed research and career development plan builds directly on my prior experiences to assess vascular health and function in aging, vulnerable populations. In line with the NIA’s Strategic Directions of developing effective interventions to reduce the burden of age-related conditions, the proposed pilot/feasibility study will explore the impact of high-intensity exercise on arterial stiffness and cognitive functioning among OPWH who have neurocognitive disorders, and refine and finalize elements critical to conducting future, fully powered randomized controlled trials (RCTs). Specific Aim 1 will evaluate the feasibility (i.e., safety, adherence, retention) and preliminary impact of 12 weeks of high-intensity exercise compared with continuous moderate exercise on arterial stiffness and cognition. Specific Aim 2 will be guided by the Theoretical Domains Framework to qualitatively assess the baseline barriers to engagement in exercise and the participant experiences of the exercise intervention. This award will enhance Dr. Jones’ career development to conduct future, large-scale RCTs by 1) enhancing his proficiency in HIV and HIV-comorbidity research, 2) developing expertise in HIV-associated neurocognitive impairment and assessment, 3) advancing knowledge of exercise clinical trial development and implementation, and 4) attaining skills in qualitative research methods to understand barriers to engagement and participant experiences. Dr. Jones’ mentorship team is well-suited to facilitate the research and career development plan, with combined expertise in HIV, cognition, exercise clinical trials, and qualitative methodology.

NIH Research Projects · FY 2026 · 2024-01

Project Summary/Abstract Title: A comprehensive approach to understanding the genetic causes of urinary stone disease: combining monogenic and polygenic analyses One in eight Americans is affected by Urinary Stone Diseases (USDs), with 52% of them experiencing recurrence within 10 years. Despite their high prevalence, our understanding of the underlying mechanisms of stone formation remains incomplete. USDs are thought to result from a combination of genetics and environmental factors. Genetic studies have identified 51 monogenic causes, accounting for 11-20% of pathogenesis. The heritability of USDs is estimated to be 45-50%. We hypothesize that there are additional novel monogenic and polygenic causes, as well as gene-gene (GxG) interactions, contributing to the pathogenesis of USDs. With this in mind, we plan to systematically investigate both monogenic and polygenic genes to identify novel variants associated with USDs. To accomplish this, we will perform whole genome sequencing and whole exome sequencing analyses on patients recruited from Harvard Medical School and NIH's "All of Us" project. We will also conduct GxG analysis to analyze the relationship between the risk loci and monogenic genes and develop a polygenic risk score (PRS) for USDs. Furthermore, we will integrate the burden analysis from monogenic genes into PRS to develop a mono-polygenic risk score (MPRS) for clinical and research use. Our goal is to employ this comprehensive approach to gain a deeper understanding of USDs, facilitating future precision pharmaceutical or environmental interventions. This K08 grant proposal also outlines a five-year plan for the development of Dr. Chen-Han Wilfred Wu, MD, PhD, into an independent physician-scientist specializing in Urology and Genetics. Dr. Wu has had a unique interdisciplinary training in Urology and Genetics, and he will continue to strive forward as a uro-genetics researcher by gaining additional expertise in both monogenic and polygenic approaches, epidemiology and biostatistics, and integrating physiology and functional studies with genetics. Through a structured mentorship plan, multiple advisory committees, coursework, and conferences, Dr. Wu will gain the necessary skills to advance his career as a physician-scientist. Under the guidance of Dr. Friedhelm Hildebrandt, MD, an esteemed expert in monogenic studies, and Dr. Fredrick Schumacher, PhD, MPH, a respected statistical geneticist, Dr. Wu is firmly committed and passionately devoted to advancing science and medicine to improve patient care as a physician-scientist.

NIH Research Projects · FY 2026 · 2024-01

Project Summary/Abstract This NIH F31 application describes a four-year plan for mentored research and career development for the PI. This proposal is focused on understanding the mechanisms of 14-3-3 phosphorylation in synucleinopathies. Parkinson’s Disease (PD) and Dementia with Lewy bodies (DLB) are synucleinopathies characterized by their aggregation of alpha-synuclein (αsyn). These diseases are characterized by a mix of progressive motor, cognitive, and autonomic symptoms. Our lab studies the role 14-3-3 proteins play in these diseases. 14-3-3s are a ubiquitously expressed family of proteins representing nearly 1% of all soluble protein in the brain that mainly exert their functions through protein-protein interactions (PPIs). We have found 14-3-3 proteins, in particular the 14-3-3θ isoform, are protective against αsyn while inhibition of these proteins leads to increased toxicity. Furthermore, we found that human cortical lysates of PD and DLB patients showed increased 14-3-3θ phosphorylation at S232 compared to healthy, age-matched controls. Testing the impact of 14-3-3θ phosphorylation, we found that the non-phosphorylatable S232A mutant is protective in PD models, while the phosphomimetic S232D mutant showed no protection or accelerated toxicity. These results lead us to hypothesize S232 phosphorylation plays a role in 14-3-3θ’s loss of protective functions. The goal of this project is to understand the causes and downstream effects of S232 phosphorylation. In Aim 1 I propose to identify what kinases cause this phosphorylation. I will work with Dr. Christopher Willey, who is in charge of the UAB Kinome Core. I will use both a candidate approach and non-biased approach to identify key kinases that increase 14-3-3 phosphorylation by PD-associated toxicants, trichloroethylene (TCE) and rotenone. For an unbiased approach, I will use the PamStation Kinomics chip to identify kinases activated by rotenone and TCE. I will validate these results in-vitro utilizing kinase inhibitors and will evaluate the biological relevance of identified kinases by genetic knockdown as well as pharmacological inhibitors of the kinases. Dr. Willey will guide me on the bioinformatic techniques as well as selection of candidate kinases for further study. In Aim 2 I want to understand what the downstream consequences are for the hundreds of 14-3-3 PPIs. I will work with Dr. James Mobley, head of UAB’s Mass spectrometry and proteomics core, to perform mass spectrometry of co-immunoprecipitates from wildtype, S232A or S232D knock-in cortical brain lysates to understand how protein networks binding to 14-3-3θ change in response to 14-3-3θ phosphorylation. The experiments in addition to the input from my mentor, Dr. Talene Yacoubian as well as my committee will help me meet my goals of increasing understanding of neurodegeneration, develop lab and bioinformatic skills, communication skills, teaching and mentorship skills, and clinical skills necessary for future patient care. All these will help me become an excellent physician-scientist focused on neurodegenerative disorders.

NIH Research Projects · FY 2025 · 2023-12

ABSTRACT During tumor progression, loss of cell:cell adhesion and detachment from the extracellular matrix induces dynamic reorganization of the cytoskeleton, formation of invadopodia by the tumor cells, and activation of epithelial-to-mesenchymal transition. This is enabled by tuning molecular programs in order to adapt to loss of attachment. Merlin, encoded by the NF2 gene, is a member of the ezrin-radixin-moesin protein family that links membrane proteins to the cortical actin cytoskeleton and regulates adhesion, migration, cell-cell contact, proliferation, and signal transduction. Merlin is critically involved in contact-dependent inhibition of growth. We found that Merlin protein levels are reduced in the majority of cases of infiltrating ductal carcinoma and metastatic breast cancer tissues. Decreased Merlin protein expression was also seen with advanced nodal involvement in breast cancer. In order to ascribe clinical relevance, we re-capitulated the loss of Merlin in breast cancer cells. Merlin deficiency elicited a markedly invasive phenotype, morphologically and programmatically evident as epithelial-to-mesenchymal transition. In order to overcome the challenge of embryonic lethality of a total Nf2-knockout, we generated a unique mammary-specific Nf2-knockout mouse mammary tumor model. These mice show remarkably accelerated development of tumors. In this R21 application, our objective is to target potential liabilities in Merlin-deficient mammary tumors to mitigate their metastasis-conducive tumor portfolio. Using multiple Merlin-deficient mammary tumor systems and tumor- derived organoids, we will investigate (i) targeting aberrantly activated Hh signaling in Merlin-deficient tumors, and (ii) the potential to target deregulated tryptophan metabolism in Merlin-deficient tumors. Our approaches are non-overlapping and complementary. Outcomes from our work will present two unprecedented strategies to target possible liabilities in Merlin-deficient breast cancer; these findings can be expanded and/or translated to other tumor types that harbor Merlin deficiency.

NIH Research Projects · FY 2024 · 2023-12

Antiretroviral therapy (ART) can reduce viremia to undetectable levels in people living with HIV (PLWH) . However, replication-competent virus persists in peripheral blood and tissues that is capable of reestablishing the infection upon antiretroviral therapy interruption (ATI). One of our long-term goals is to aid the development of effective HIV cure strategies by gaining a better understanding of the host physiological and metabolic processes by which cellular reservoirs are established and maintained, and those of viral reactivation. Regardless of how HIV infection is acquired, the gastrointestinal (GI) tract is a major site of HIV replication. Although the majority of immune cells in the body, including CD4+ T cells, reside in the GI tract, little is known about the HIV reservoir that is established in this tissue. Analyses of ART-treated patients have demonstrated higher HIV-DNA levels in GI tract cells compared to blood cells, suggestive of a larger HIV reservoir. Examination of HIV transcripts in cells from the blood and rectum of ART-treated PLWH also indicates that the GI tract may be enriched in latently-infected cells and suggests that HIV latency is maintained by different mechanisms in the GI tract and/or that a deeper s tate of latency may be maintained. Based on these observations, our overarching hypothesis is that HIV establishes a latent infection in the GI tract that contributes to virus rebound during ATI. T o our knowledge there is virtually no information addressing the role of the microbiome in establishing or maintaining the latent reservoir in the GI tract. Currently, a large amount of indirect evidence suggests that the GI microbiome is involved in HIV persistence. The GI microbiome in PLWH promotes inflammation and immune activation in the GI tract, which may influence HIV reservoir size. Furthermore, in vitro studies show that microbiota and microbial metabolites can influence HIV transcription. Therefore, we further hypothesize that the intestinal microbiome contributes to the establishment and persistence of the HIV reservoir and thereby impacts the contribution of the GI tract to virus rebound. A role for the microbiome in HIV remission would be fundamentally important to HIV cure development. Previously, we and others demonstrated that HIV establishes a latent infection in ART-suppressed humanized mice that upon discontinuation of ART results in robust virus rebound. Importantly, we recently demonstrated reproducible induction of HIV in resting CD4+ T cells in multiple tissues of ART-suppressed humanized mice using two different latency reversal approaches. Our objective is to analyze the HIV reservoir in the GI tract using BLT humanized mice, a well- characterized model of HIV latency, persistence and reactivation. We will 1) analyze HIV reservoir formation in the GI tract, 2) evaluate the contribution of the GI tract to viral rebound, and 3) assess HIV induction in the Gl tract by latency reversing agents. We will also use an innovative germ-free BLT mouse model to determine how the presence of the GI microbiome contributes to HIV persistence and rebound.

NIH Research Projects · FY 2026 · 2023-12

SUMMARY Urinary oxalate excretion is a key risk factor in the formation of calcium oxalate kidney stones, the most common type of kidney stone, a disease that affects 9% of the US population. Therapeutic development for prevention of calcium oxalate kidney stone disease is hampered by our incomplete understanding of the factors leading to the increased urinary oxalate pool in many patients with calcium oxalate kidney stones. Environmental and lifestyle factors as well as dietary and metabolic, or endogenous, factors may all play a role, but the interplay between them has clouded the interpretation of the oxalate focused studies that have been done in kidney stone formers. Our preliminary data and published studies suggest there is both an increased synthesis of oxalate and a greater gastrointestinal absorption of oxalate in calcium oxalate kidney stone formers. These warrant confirmation and the underlying mechanisms need to be defined. The frequency of colonization with Oxalobacter formigenes, a gut commensal oxalate degrading bacterium, is substantially lower in calcium oxalate kidney stone formers and its absence has been linked with increased kidney stone risk. Colonization with Oxalobacter formigenes leads to reduced urinary oxalate excretion in healthy volunteers. Whether colonization with this organism is possible in calcium oxalate kidney stone formers and leads to reduced urinary oxalate excretion warrants study. Our hypothesis is that increased endogenous oxalate synthesis and increased gastrointestinal absorption of oxalate via greater transport capacity and higher bioavailablity due to decreased microbial oxalate degradation in the gut lead to a greater influx of oxalate and increased urinary oxalate excretion. This proposal aims to 1/ demonstrate that endogenous oxalate synthesis is increased in idiopathic calcium oxalate kidney stone formers using 13C-isotope oxalate infusion to quantify oxalate production and determine the contribution of different oxalate precursors by oral dosing with 13C-glycolate and 13C-ascorbic acid; 2/ confirm that gastrointestinal oxalate absorption is higher in idiopathic calcium oxalate kidney stone formers and define the driving mechanisms by using a combination of low and high oxalate diets and 13C-oxalate oral dosing coupled with sugar functional intestinal permeability tests that will identify gut segments and type of transport involved; 3/ demonstrate that the colonization of Oxalobacter formigenes in idiopathic calcium oxalate kidney stone formers is possible, sustainable, and results in a reduction of urinary oxalate excretion. We expect this proposal to identify mechanisms playing a role in the increased urinary oxalate pool in idiopathic calcium oxalate kidney stones patients; develop tools to investigate these mechanisms; provide a potential treatment for lowering urinary oxalate excretion by colonization with Oxalobacter formigenes. This in turn may lead to the development of targeted therapies to be tested in clinical trials for treating those afflicted with calcium oxalate kidney stones.

NIH Research Projects · FY 2026 · 2023-12

Parkinson’s disease and other movement disorders are by far the most common indications for deep brain stimulation (DBS) worldwide. Despite intense and ongoing investigation, there are no established biomarkers to identify optimal stimulation locations and parameters with increasingly complex directional and adaptive DBS devices. Here we will collaborate with our industry partner Boston Scientific to pioneer use of their novel 16- contact directional Argyle lead, informed by their emerging stimulation and recording architecture, NEXT (Neuromodulation Experimental Testbed). We will use advanced experimental and analytic methods to generate detailed spatiotemporal maps of stimulus-evoked electrophysiology in the subthalamic nucleus versus globus pallidus interna in patients with Parkinson’s disease, both awake and under general anesthesia. We will then create predictive models from these spatial maps to guide directional DBS activation and clinical programming. Our overall goal is to pair our novel analytic methods with emerging device technologies to develop predictive biomarkers to guide directional DBS therapy for Parkinson’s disease and other complex neurocognitive disorders. Our vision, and that of our industry collaborator, is for next-generation DBS devices to integrate these technologies into fully implanted devices that can reveal crucial interactions between DBS and human brain circuits to guide clinical decision-making at the point of care during both surgical targeting and clinical programming.

NIH Research Projects · FY 2026 · 2023-12

Even though schizophrenia spectrum disorders (SSD) are generally considered to be neurodevelopmental in origin, emerging evidence supports the idea that aberrant brain aging may also occur. Because this concept has only recently reemerged, data on characteristic phenotypical and brain age-related trajectories in SSD and their underlying mechanisms remain sparse. These critical knowledge gaps constitute major obstacles in identifying actionable treatment targets for aberrant aging in SSD. Our proposal has two overarching goals: (1) address existing knowledge gaps in age-related changes in SSD and (2) identify actionable targets for novel interventions designed to ameliorate, or even prevent, aberrant aging in SSD. A considerable body of work in geroscience suggests that normal aging phenomena are present at multiple levels. Specifically, age-related declines in brain structures and cognitive function are well established. Recent findings also suggest that energy metabolism (bioenergetics) in the brain, which is critical for brain plasticity and cognition, is negatively affected by aging. Emerging data further supports the idea that these phenomena are intricately connected and point towards bioenergetic aging as a potentially driving force behind structural brain aging and cognitive aging. These principles of aging in the general population provide the conceptual framework of our project. We propose to leverage state-of-the-art multimodal neuroimaging techniques and tailored cognitive tasks to systematically evaluate age-related changes in brain structure, function, and bioenergetics in the dorsal anterior cingulate cortex (dACC), which is considered a hallmark brain region implicated in the pathophysiology of SSD and plays a critical role in nonsocial and social cognition. Over a period of 5 years, we will enroll 120 individuals with SSD and 120 demographically matched controls. We will quantify (1) cortical thickness deviations in the dACC using novel normative reference models, (2) social cognition and nonsocial cognition with a tailored behavioral battery and task fMRI, and (3) in vivo spectroscopic indices of bioenergetics in the dorsal ACC (dACC) using ultra-high-field MR spectroscopy (MRS). We will determine if these measures of patients differ from those of healthy volunteers (Aim 1) and if the effect of age on these measures differs between patients and controls (Aim 2). We will also explore whether age trajectories converge across measures in an exploratory aim. The findings of this project could offer mechanistic insights into aging process in SSD by focusing on multiple units of analysis and provide actionable treatment targets for aberrant aging in SSD.

NIH Research Projects · FY 2025 · 2023-12

PROJECT SUMMARY/ABSTRACT Crohn’s disease (CD), which affects over 500,000 individuals with CD in the US, is a chronic and uniquely challenging condition that requires lifelong medications, major surgeries, and health promotion. Physical activity (PA) is an essential component of many chronic disease and surgical treatment programs, but the potential benefits of PA in CD are poorly understood as there has been a paucity of research in CD. Identifying factors that influence PA participation and the extent and intensity regular PA has a therapeutic effect (i.e., inflammatory biomarkers, mucosal healing) on disease activity in CD is important for developing effective PA interventions that can be translated to practice. Therefore, the primary objective of this research proposal is to examine the complex behavioral and biological correlates of PA in CD. This project seeks to compare PA rates between adults with CD and healthy controls (Aim 1) and examine the psychosocial and clinical influences on self-report and device-measured PA (Aims 2 and 3) in CD. The central hypothesis is that adults with CD who report lower levels of self-efficacy engage in less device-measured PA and more sedentary behavior than the general population, and that the relationship between PA and disease outcomes varies by both volume and pattern of PA behavior and the aspect of disease activity measured. The public health significance of this study lies in its potential to develop an improved, theoretically sound understanding of PA behavior in CD and the relationship between PA and disease activity. These findings could potentially result in better designed, more effective PA programs for adults with CD that ultimately have the potential to reduce healthcare costs associated with adverse postoperative outcomes. My ultimate career goal is to lead a collaborative research team that bridges the gap between evidence-based PA interventions and the translation of this evidence into clinical and community practice in the context of CD. To better prepare myself for a career as an independent investigator, I am seeking training in 4 major areas: 1) rigorous research in the CD field, including familiarity with CD clinical patient care; 2) multidisciplinary collaboration with clinicians and scientists; 3) translational research, including clinical trial design, statistical methods, and interpretation of results for clinical application and future intervention development; and 4) professional development, including principles of scientific integrity and responsible conduct of research. Training in these areas is integral to completing the research aims and providing me with a solid foundation for a successful career as an independent researcher.

NIH Research Projects · FY 2025 · 2023-12

Project Summary: Millions of antibody sequences from both healthy individuals and patients with a variety of immune-mediated disorders are now freely available to the research community. This has been made possible by support primarily provided by NIAID for the research projects themselves and for the databases that curate the data. To maximize the benefits of this financial support, the Institute now funds efforts to use the curated data to explore new initiatives which are beyond the scope of the original research projects. We propose to use approximately one million of these antibody sequences to test a hypothesis concerning rare antibody sequences that was first proposed over 35 years ago but which now can be examined on a large scale by means of these NIAID supported databases. The expansion of the antibody repertoire by V(D)J gene rearrangement, N-nucleotide addition, and somatic hypermutation (SHM) introduces non-germline encoded sequence changes throughout the antibody variable domain. Many of these sequences are either totally absent from, or very rare in, the human proteome. It is already clear that these “foreign” sequences are common in V regions of antibodies even from healthy human controls, so it is unlikely that the majority are immuno-pathogenic. However, there is reason to test the hypothesis that a small subset of these forbidden/rare pentapeptides will be clinically immunogenic, i.e., will be able to induce immune responses that should be absent from normal antibodies. If this can be confirmed, it will help us address at least two major challenges now facing medical immunology. First is the persistent problem of immunogenicity in new therapeutic antibodies which substantially raises the cost of development of these agents and limits their efficacy in patients. If the proposed work is successful and the immunogenic pentapeptides are identified, it could lead to the development of new algorithms that more accurately predict the immunogenicity of therapeutic antibodies prior to expensive clinical trials. Second is the need for more specific and effective therapies for autoimmune diseases such as SLE, MS, and type 1 diabetes. A large body of evidence indicates that many of the self-reactive, pathogenic autoantibodies contain idiotopes in their structures which are recognized by anti-idiotypic antibodies. Further, it appears that the presence or absence of these anti- idiotypic antibodies may in part be responsible for periods of remission or active disease. Few idiotope structures are known, but it seems reasonable that these immunogenic segments in antibody variable regions may also contain the foreign pentapeptides; this could serve as a tool for identifying idiotopes in the sequences of pathogenic self-reactive antibodies. A major outcome of the proposed studies is a bioinformatics research tool, the Penta-Scope, which will take as input the sequences of large numbers of antibodies and return the frequencies of their component pentapeptides in human and murine proteomes. This tool will be made freely available online for other investigators to use in studies of antibody structure and/or immunogenicity.

NIH Research Projects · FY 2025 · 2023-12

ABSTRACT Delirium, or acute confusional state, affects 30-40% of hospitalized older adults, with the added cost of care es- timated to be up to $7 billion. Medications are one of the most common causes of delirium and potentially one of the most preventable and most treatable, with medications thought to be the sole precipitant for delirium in up to 39% of cases. Despite the critical role of medications as a source of delirium, surprisingly few studies have investigated this topic, often focused on only a few medications and with only modest quality. In this proposal, we will address these limitations by utilizing data collected through our Virtual Acute Care for Elders (ACE) quality improvement project, which instituted delirium screening once per shift by nursing staff for all individuals over age 65 admitted to University of Alabama at Birmingham (UAB) Hospital for the past decade. This unprecedented volume of data on more than 81,000 patients will allow us to achieve the necessary sample sizes for effective ap- plication of data mining algorithms. Data mining algorithms that discover patterns of associations in data, rather than testing predetermined hypotheses, are well suited to application in large-scale algorithms to identify med- ications associated with increased odds of delirium. Combining data mining with our Virtual ACE and hospital electronic health record (EHR) data, we will be able to evaluate more than 1,000 individual medications, as well as medication combinations and medication classes, for their association with delirium. Such a comprehensive, data-driven examination of delirium due to medications will be a significant contribution to our understanding of the risk factors and management of delirium in the older adult population.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY About 100,000 people die annually in the U.S. because of opioid overdose or complications of opioid use disorder (OUD). Three medications for OUD (MOUD) are FDA-approved and regularly used to treat OUD: methadone, buprenorphine, and extended-release naltrexone (XR-NTX). However, persons who use opioids, including those prescribed MOUDs, report sleep disruption. In addition to the sleep centers of the brain, mu opioid receptors (MORs) are also expressed in the retina (including the human retina), specifically in ganglion cells that are critically important for non-image forming photoreception including circadian regulation of sleep-wake behavior. Pre-clinical studies show that activation of MORs on these intrinsically photosensitive retinal ganglion cells (ipRGCs) reduces the electrophysiological response to light, impacting critical ipRGC functions such as synchronization of sleep-wake behavior and circadian rhythms to light (photoentrainment), light-induced melatonin suppression, and the post-illumination pupillary reflex (PIPR). Together, these results suggest that activation of MORs in the ipRGCs by opioid use and/or MOUDs may impair downstream ipRGC functions. This multi-disciplinary study will examine the novel overarching hypothesis that persistent alterations in sleep/wake behavior in OUD patients undergoing treatment are mediated by impaired ipRGC function, and biomarkers of this pathway can predict recovery and relapse. Three aims will be tested in a sample of 200 participants, 150 of whom will be engaged in MOUD therapy (e.g., 50 each on methadone, buprenorphine, and XR-NTX, respectively) and 50 of whom will be non-opioid using control participants. Aim 1 will test the hypothesis that MOUD differentially impacts function of ipRGC responses. Aim 2 will examine whether MOUD differentially impacts daytime sleepiness, daily sleep-wake behavior, sleep architecture, and sleep-disordered breathing. Finally, Aim 3 will determine if ipRGC function predicts opioid relapse among MOUD groups at 1-, 3- and 6- month follow-up. Compared to non-opioid using controls or persons receiving an opioid antagonist (XR-NTX), we predict that participants who are receiving an agonist (methadone) or partial-agonist (buprenorphine) MOUD will have the most ipRGC interference, as evidenced by reduced PIPR, attenuated light-induced melatonin suppression, reduced circadian rhythmic amplitude, increased sleep latency, and increased sleep fragmentation. Importantly, we hypothesize that impaired ipRGC function will predict worse treatment outcomes as indicated by opioid use by 6-month follow-up. Finally, an exploratory aim will examine whether the MOUD groups show different relationships between opioid craving/withdrawal symptoms and sleep-wake behavior over a 10-day assessment of the participants’ daily lives within the normal environment. The results of this study will be highly significant because it would support the use of the pupillary response to light and other indicators of ipRGC function as novel biomarkers to predict the response and outcomes to MOUDs.

NIH Research Projects · FY 2024 · 2023-09

Building and implementing a predictive decision support system based on a proactive full capacity protocol to mitigate emergency overcrowding problem Project Summary Emergency departments (EDs) face a major problem of overcrowding that poses a significant patient safety risk and leads to poor healthcare service quality and high mortality rates. ED overcrowding is a patient flow problem, which can be solved by improving patient flow from arrival to admission or discharge. According to the American College of Emergency Physicians, a full capacity protocol (FCP) is a key approach for improving patient flow and consequently mitigating ED overcrowding. FCP has different levels that are triggered by different criteria, which are based on patient flow measures (PFMs). The current practice of FCP uses real-time (i.e., reactive) information to decide FCP criteria. However, when it comes to implementing FCP interventions, using real-time information is not efficient because in many cases FCP levels are activated too late when ED is already overcrowded. This project improves the reactive FCP to make it proactive through using Artificial Intelligence and predictive analytics. The PFMs will be predicted using deep learning models and then integrated with reactive FCP. A decision support system will be developed to implement the proposed proactive FCP. The overall objective of this project is to develop a framework to mitigate ED overcrowding. There are four aims (Aims 1& 2 under R21; Aims 3& 4 under R33): Aim 1: Develop deep learning models to predict different PFM values and incorporate them in a proactive FCP. Many PFM values represent the patient flow from arrival to admission. We will build multiple deep learning models to predict PFM values (e.g., numbers of boarding). Then, we will update the reactive FCP to include the predicted PFM values. Aim 2: Develop a DES model to evaluate the effectiveness of proactive FCP. Before running the proactive FCP in production, we will compare reactive and proactive FCPs on the outcomes they generate such as average length of stay (LOS), waiting time and staff satisfaction. Aim 3: Design, evaluate, and implement a decision support system (DSS) based on the proactive FCP. We will design user-centric DSS to aid clinicians and the PFCT in implementing the proactive FCP. We will use the proactive FCP criteria as input for the DSS to automate key parts of the proactive FCP interventions. Aim 4: Expand and generalize the DSS by standardizing data input and output interfaces. We will create a FHIR-based application programming interface (API) to allow site-specific configuration, model training, evaluation, and streamlining of implementation processes. Successful completion of this project delivers a state-of-the-art interoperable DSS for the implementation of a proactive FCP based on early, accurate predictions of PFM values to allow proper planning and execution of patient flow processes, thereby mitigating ED overcrowding. Our multi- disciplinary team is well positioned to successfully execute all aims.

NIH Research Projects · FY 2025 · 2023-09

Project summary: This proposal which is entitled "Defining epidemiology, diagnosis, prevention and treatment of invasive fungal infections in the United States" is an ambitious and overarching proposal which covers a host of issues in the clinical mycology arena. This proposal represents a series of independent projects that will be initiated in sequential fashion throughout the course of this 5- year award. The Mycoses Study Group (MSG) with its central unit located at UAB in the division of infectious diseases will be administering this program. The proposal has 3 broad specific aims which address some of the most important current public health issues and challenges as it relates to fungal infections. In the first aim, we will develop and maintain reliable and innovative methods of national fungal surveillance while also assessing specific risk factors for acquiring these infections in the treatment and outcomes associated with these disorders. The second aim will address a huge gap in the management of invasive mycoses, specifically improving the utility and the scope of fungal diagnostics. We will explore this through reviewing the existing utilization of fungal diagnostics but also creating a specimen repository that is linked to specific patients with the purpose of developing new diagnostics. Finally, we will explore specific public health interventions which may serve to mitigate the significant impact of fungal infections both in the general population and in specific high-risk subgroups. To accomplish these ambitious goals obviously requires significant coordination of efforts among a broad range of investigators, clinical centers, laboratories, and the CDC. The Mycoses Study Group is uniquely positioned to coordinate these efforts given his extensive history of designing coordinating and implementing both clinical and epidemiologic trials in mycology since 1978. We believe that at the completion of this award we will have moved much closer to understanding of current epidemiology of fungal infections in the United States will also have a much clearer picture of those who are at particularly high risk of developing severe complications from these infections. Moreover, we believe that we will have made significant strides towards improved deployment of existing fungal diagnostics and significant progress towards the development of new diagnostics. Finally, we will improve outcomes through increased public awareness and specific and strategic interventions which target individuals at highest risk and who may realize the greatest benefit from preventative strategies.

NIH Research Projects · FY 2025 · 2023-09

SUMMARY Gout is prevalent in the US (3.9% of the adult population) and not only directly impacts peoples’ lives but is also co-morbid with cardiometabolic disease. Gout consists of unpredictable episodes of acute inflammation or flares resulting from monocyte NLRP3 inflammasome activation by monosodium urate (MSU) crystals in people with hyperuricemia, the subsequent production of IL-1, and recruitment of large numbers of inflammatory cells into the affected joint. Epigenetic reprogramming and altered gene transcription in response to elevated levels of soluble urate are mechanisms of this enhanced cellular reactivity to the secondary stimuli, known as innate trained immunity. To better understand the factors controlling the progression from hyperuricemia to gout we have completed a very large genome-wide association study (GWAS) in gout, identifying several hundred gout-associated loci. Loci include long-noncoding RNAs (lncRNAs) that have diverse functions including regulation of gene expression, deposition of epigenetic modifications and organization of chromosome architecture. Some are immune gene-priming lncRNAs (IPLs) that direct transcriptional machinery over multiple promoters. From these loci we have identified one key causal pathway (clonal hematopoiesis of indeterminate potential (CHIP)) for which we will study the molecular genetic processes controlling its activation. The CHIP pathway is involved in control of the epigenome - DNA methylation, histone modification and metabolism of substrates. It may make people more susceptible to gout by causing the innate immune system to be hyper-responsive to MSU crystals. We hypothesize that the genetic loci identified from the gout GWAS control activity of this pathway by regulation of gene expression, including through regulation of the epigenome. In testing this hypothesis, in three Aims, we will understand the molecular control of the pathway and provide a basis for future studies in targeting this pathway for improved management of gout. In Aim 1 we will use experimental systems, including a zebrafish model of gout, to understand where regulatory regions are expressed and how the gout-associated loci influence NLRP3- inflamasome activation. Zebrafish are translucent, allowing development of innovative models, and there is 82% conservation of genes with humans and functionally equivalent macrophages and neutrophils. In Aim 2 we will compare the transcriptome and epigenome in MSU crystal-stimulated monocytes of individuals with high and low burdens of gout risk alleles in order to gain further insights into the molecular regulation of the pathways, and to identify downstream pathways. Individual genes will be knocked down in a cell line. In Aim 3 we will use our established pipeline to understand how lncRNAs (i.e. IPLs) connect transcriptional machinery to the promoters of innate immune genes at specific loci in the molecular control of activation of the gout flare. Our studies will deepen our knowledge of the mechanisms of gout flares and its genetic basis, and ultimately point to areas of research that may allow for novel treatments in gout.