Ut Southwestern Medical Center

NIH Research Projects · FY 2025 · 2024-01

Although brain oscillations are a promising target for neuromodulation for cognitive disease, we have a poor understanding of the molecular mechanisms that lead to oscillatory activity. Motivated by this gap in knowledge, the Lega-Konopka collaboration has developed groundbreaking techniques to study the links between gene expression and memory-enhanced brain oscillations. By integrating gene expression data and memory-relevant iEEG signatures of the same human epilepsy patients, the collaboration discovered significant associations between memory-associated oscillations and specific gene expression patterns in the temporal pole. This work identified SMAD3 as a highly promising candidate for future study. We found strong links between SMAD3 gene expression and slow theta oscillations, which are uniquely important for human cognitive processes. We also found evidence that SMAD3 binds the regulatory elements of dozens of other genes connected with slow theta oscillations. Many of these putative SMAD3 gene targets are implicated in neuropsychiatric disorders characterized by cognitive dysfunction. Taken together, this evidence suggests that SMAD3 may coordinate the transcription of many genes that affect brain activity during memory processing. However, the brain-specific transcriptional targets of SMAD3 and the precise impact of SMAD3-mediated gene networks on human neural activity have yet to be definitively identified. The few studies on the functional role of SMAD3 in the brain showed that Smad3 knockout mice have impaired neurogenesis and long-term potentiation. Although animal models generate valuable insight into brain function, the complicated nature of human cognitive processes and related oscillations decreases the translational value of approaches using animals. To overcome these problems, I will use human organotypic slice culture (OSC) to define the function of SMAD3 in the regulation of memory-related gene networks and neural activity in the human brain. In Aim 1, I will enhance SMAD3 activity in human OSC then use snRNA-seq and snATAC-seq to identify SMAD3 gene targets. In Aim 2, I will demonstrate the ability to silence SMAD3 expression in human OSC using lentiviral shRNA constructs. I will then use high-density microelectrode array recordings to understand how the loss of SMAD3 impacts activity at the single neuron and network levels. By increasing our understanding of the mechanisms underlying memory-relevant brain activity, the completion of this project will lead us toward new therapies for cognitive disorders. This would be impossible without the stellar support of Drs. Lega and Konopka, who have expertly guided my development as an aspiring physician-scientist. The training plan in place for this fellowship period will allow me to learn and develop innovative skills for studying human brain tissue in vitro with a combination of electrophysiology, molecular biology, and genomic approaches. Benefitting from the unique combination of expertise from my mentors and the resources provided by UTSW, this training will allow me to grow as a rigorous, passionate physician-scientist with the tools to develop novel methods to study and treat the neurological disorders of my patients.

NIH Research Projects · FY 2026 · 2024-01

Plasma HDL levels are inversely associated with coronary, cerebral and peripheral arterial disease (PAD), and also type 2 diabetes mellitus (T2DM). However, how HDL influences these conditions remains poorly understood. We previously showed that HDL attenuates vascular inflammation and promotes neovascularization via scavenger receptor class B type I (SR-BI) and its adaptor protein PDZK1 in endothelial cells (EC). We recently identified Breakpoint Cluster Region (BCR) protein as a novel PDZK1 interacting protein in human EC. In other contexts BCR has known functions modulating Rac1 and RhoA, and we discovered it to be a novel kinase for Akt kinase activated by HDL in EC. Our studies in BCR null mice then revealed for the first time that BCR is required for HDL-related atheroprotection, HDL-induced endothelial repair and angiogenesis, and normal glucose homeostasis. The Overall Goal of the present project is to determine HOW BCR actions in EC contribute to HDL-related atheroprotection and promotion of neovascularization and normal glucose homeostasis. Three Aims are proposed in cultured EC and mice. Aim 1 will determine how endothelial BCR impacts atherosclerosis. We recently discovered in culture that BCR is necessary for HDL attenuation of both the monocyte-EC adhesion and the Rac1-dependent EC LDL transport that converge to drive atherogenesis. Using cultured EC, floxed BCR mice, and nanoparticle-based EC cDNA delivery to reconstitute EC wild-type or mutant BCR expression in vivo, we will test the hypothesis that HDL-related atheroprotection is mediated by EC BCR and its capacity to function as a kinase or inhibitor of Rac1. In cultured EC we will also query how HDL subspecies with varying Apo-A1 and Apo-A2 content and size impact BCR-dependent atheroprotective processes. Aim 2 will determine how EC BCR impacts neovascularization, which is critical to PAD pathogenesis and resolution. We recently showed that EC migration prompted by HDL is BCR-dependent. Using cultured EC, floxed BCR mice, reconstitution of EC BCR in vivo, and mouse models of EC repair, angiogenesis and hindlimb ischemia, we will test the hypothesis that EC BCR function as a kinase or an activator of RhoA underlies HDL-induced neovascularization. In culture, how HDL subspecies impact EC neovascularization mechanisms will also be examined. Aim 3 will determine how EC BCR impacts glucose control. Knowing that HDL has antidiabetic action and that EC insulin transport to skeletal muscle drives processes underlying 80-90% of total body glucose disposal, we have discovered that HDL stimulates EC insulin transport via BCR. Using cultured EC, floxed BCR mice, reconstitution of EC BCR and glucose control phenotyping in vivo, we will test the hypothesis that EC BCR function as an Akt kinase kinase underlies HDL promotion of normal glucose homeostasis. In culture, how HDL subspecies impact EC insulin transport will also be studied. The proposed work providing new insights into how HDL affords cardiometabolic protection has the potential to add clarity to why HDL has varying impact on individuals, and to reveal new therapeutic targets to leverage against cardiovascular and metabolic disease.

NIH Research Projects · FY 2026 · 2024-01

ABSTRACT A key feature in intestinal immunity is the dynamic intestinal barrier, which separates the host from resident and pathogenic microbiota through a mucus gel impregnated with antimicrobial peptides. The mechanisms underlying the maintenance and function of this intestinal barrier are not completely understood. Using a mouse forward genetic screen for defects of intestinal homeostasis, we have found a mutation in Tvp23b, which conferred susceptibility to both chemically induced and infectious colitis. Golgi apparatus membrane protein TVP23 homolog B (TVP23B) is a transmembrane protein conserved from yeast to humans. In the intestine, the protein is localized to the epithelium and its deficiency in the hematopoietic extrinsic compartment was essential to the colitis phenotype. We found that TVP23B controls the homeostasis of Paneth cells and function of goblet cells in vivo, leading to a decrease in antimicrobial peptides as well as a more penetrable mucus layer. As a result, Tvp23b-/- mice displayed decreased barrier function and a loss of host-microbe separation. TVP23B- deficient colonocytes have a loss of core-3 O-glycosylation of colonic proteins, which is the major O-glycosylation present on gel forming mucins. TVP23B binds with another Golgi protein, YIPF6, which is similarly critical for intestinal homeostasis. The Golgi proteomes of YIPF6 and TVP23B-deficient colonocytes have a common deficiency of several critical glycosylation enzymes, including those necessary for core-3 glycosylation of mucins. TVP23B is necessary for the formation of the sterile mucin layer of the intestine and its absence disturbs the balance of host and microbe in vivo. In this proposal, we will examine the components that mediate the cellular and molecular dysfunction in TVP23B deficiency and the resulting pathology in three Specific Aims: (1) Determine the role of TVP23B in maintaining barrier function during infectious and chronic colitis (2) To examine the role of TVP23B in intestinal secretory cell differentiation and intestinal regeneration. (3) To understand role of TVP23B on Golgi enzyme trafficking. The aims in this proposal will help elucidate the mechanisms underlying the mechanisms by which epithelial cells regulate host-microbe interactions.

NIH Research Projects · FY 2026 · 2023-12

PROJECT SUMMARY The global burden of kidney disease is substantial and increasing, with a recent estimated global prevalence of over 700 million cases, with over 37 million in the United States. Mitochondrial dysfunction leading to metabolic dysregulation manifested by suppressed fatty acid oxidation is a key component in the development of kidney disease. Ketone metabolism is a central component of metabolic homeostasis. Ketogenesis occurs when fatty acids are oxidized and converted into ketones. While the liver is the main ketogenic organ, mitochondrial Hydroxymethylglutaryl-CoA synthase 2 (HMGCS2), the rate limiting enzyme for ketogenesis, is induced in the proximal tubule of the kidney in response to fasting. Using liver- and kidney-specific Hmgcs2 deletion mouse models, we found that renal HMGCS2 does not contribute to circulating ketones during fasting and is thus likely acting locally. Based on our preliminary data, we hypothesize that proximal tubular HMGCS2 has two independent intra-renal functions. First, we hypothesize that there is regional ketone metabolic cooperativism in the kidney whereby ketones are produced by the proximal tubule and are utilized by the distal convoluted tubule. This is based on our finding that the spatial expression of HMGCS2 in the proximal tubule follows a pattern in which HMGCS2-expressing proximal tubular cells are in close proximity to distal convoluted tubular cells that highly express the ketolytic enzyme 3-Oxoacid CoA-Transferase 1. Second, we hypothesize that there is a cell autonomous effect of proximal tubular HMGCS2. Using a mouse model capable of isolating proximal tubular- specific mitochondria with or without Hmgcs2 deletion, we found that HMGCS2 deficient mitochondria have a mitochondrial respiratory defect. Next, we discovered that after renal ischemia-reperfusion injury (IRI), kidney HMGCS2 is suppressed in both the early injury and late fibrotic phases. This is consistent with human CKD kidney biopsies which also exhibit suppressed HMGCS2 levels. Importantly, we found that mice lacking renal Hmgcs2 are more susceptible to renal IRI, developing more acute tubular damage and late fibrosis, relative to controls. Combining a conditional deletion strategy and capitalizing on our model capable of isolating kidney cell- specific mitochondria, we aim to understand the intra-renal ketone metabolic network in kidney health and disease. In this proposal, we will test the central hypothesis that proximal tubular HMGCS2 acts locally to 1) engage in de novo ketogenesis to fuel neighboring distal convoluted tubular cells, and to 2) support proximal tubular mitochondrial respiration and function. In Aim 1, we will define proximal tubular and distal convoluted tubular ketone metabolism and determine whether the distal convoluted tubule preferentially utilizes proximal tubule-derived ketones for energy. In Aim 2, we dissect the intracellular role of proximal tubular HMGCS2 in supporting mitochondrial function in fasting and ischemic kidney injury.

NIH Research Projects · FY 2025 · 2023-12

Project Summary Transcription of antiviral factors by the host cell is a fundamental aspect of innate immune responses to virus infection. We recently discovered a novel transcriptional response in humans termed the “FACT-ETS-1 Antiviral Response (FEAR)” Pathway that restricts the replication of vaccinia virus (VV), a large DNA virus belonging to the poxvirus family. Activation of this pathway requires the FACT complex, an ancient histone chaperone that is conserved from yeast to humans. The human FACT complex is comprised of two protein subunits, hSpt16 and SSRP1, that function together to regulate cellular gene transcription. Our work discovered that FEAR pathway activation requires a novel, SUMOylated form of hSpt16 that is normally found in the cytoplasm of cells but that translocates to the nucleus upon virus infection to form specialized FACT complexes that activate expression of ETS-1. The ETS-1 protein is a member of the ETS family of transcription factors that arose in multicellular animals ~600 million ago during evolution. Our previous work demonstrated that FACT- induced ETS-1 expression is required for the restriction of VV in human cells. However, the VV-encoded A51R protein functions as a FEAR pathway inhibitor by directly binding to SUMOylated hSpt16 and tethering it to cytosolic microtubules. More recently, we discovered that both FACT and ETS-1 are also required to restrict the replication of the RNA virus, vesicular stomatitis virus (VSV). Moreover, we found that the VSV-encoded matrix (M) protein promotes the specific depletion of SUMOylated, but not non-SUMOylated, hSpt16 subunits during infection. However, a mutant VSV strain encoding a single amino acid substitution in its M protein both fails to deplete SUMOylated hSpt16 subunits during infection and strongly induces ETS-1 expression. These data suggest that the FEAR pathway may also both restrict, and be antagonized by, RNA viruses. However, the human genes that are regulated by ETS-1 during infection that contribute to RNA virus restriction are unknown. Our preliminary data suggest that that genes upregulated by ETS-1 during VSV infection are largely distinct from the interferon response, a well-characterized antiviral transcriptional response that inhibits diverse viruses. Notably, we have also discovered that RNA viruses unrelated to VSV, such as paramyxoviruses and flaviviruses, also specifically promote depletion of SUMOylated hSpt16 subunits during infection. We hypothesize that VSV and these other RNA viruses encode FEAR pathway antagonists that deplete SUMOylated hSpt16 levels to prevent FEAR pathway activation and expression of ETS-1-regulated antiviral genes. Thus, our study goals are to: 1) Identify the human genes regulated by ETS-1 during RNA virus infection that are involved in virus restriction and 2) Determine if the FEAR pathway restricts other RNA viruses besides VSV and whether these viruses encode FEAR pathway inhibitors. Our long-term goal is to understand how the FEAR pathway broadly restricts DNA and RNA viruses and how virus-encoded FEAR pathway antagonists contribute to viral pathogenesis.

NIH Research Projects · FY 2026 · 2023-11

ABSTRACT: Balance of excitation and inhibition is critical for proper function of the nervous system. In many brain regions, there is feedforward and feedback inhibition from interneurons that release the inhibitory transmitter GABA to balance the release of excitatory glutamate from long-range inputs. However, in the lateral habenula (LHb) - a key component of the brain's reward system – there is little, if any, feedforward or feedback inhibition and few inhibitory neurons. Instead, our work suggests that excitatory/inhibitory balance is achieved, at least partly, by co-release of GABA with glutamate from individual inputs arising from the globus pallidus (GPi), a major input to the LHb. The activity of GPi inputs to the LHb and the activity of LHb neurons affect motivational vigor. They are also phasically excited by negative environmental feedback and phasically inhibited by positive environmental feedback, which guide learning. Rats with low motivation and negatively biased feedback sensitivity have elevated LHb activity and unbalanced GABA and glutamate signaling from the GPi to LHb. Based on these observations, we believe that GABA/glutamate balance from GPi to the LHb is important for (1) regulating tonic (basal) LHb activity and motivational vigor, and (2) bidirectional phasic signaling of negative and positive feedback in the LHb and behavioral sensitivity to negative and positive feedback. The objective of this grant is to test these hypotheses by artificially manipulating GABA/glutamate balance from GPi to LHb in mice and measuring (1) tonic LHb activity and motivation, and (2) phasic LHb responses and behavioral sensitivity to negative feedback (reward omission) and positive feedback (reward). Based on studies of GABA/glutamate co- release in the hippocampus, we further hypothesize that long-term changes in tonic and/or phasic LHb activity will induce homeostatic plasticity of GABA/glutamate co-transmission from the GPi to LHb. We believe this study is significant because it will reveal the relationship between excitatory/inhibitory balance and fundamental motivational and learning processes that are affected in many psychiatric disorders, while also investigating the function of GABA/glutamate co-transmission.

NIH Research Projects · FY 2025 · 2023-09

Neurons in adults are essentially irreplaceable and especially vulnerable to the accumulation of protein aggregates, dysfunctional mitochondria, and similarly distractive agents. The most important pathway available to neurons to limit such damage is autophagy. This pathway, initially described in the context of the mTor-regulated starvation-induced metabolic rescue pathway in yeast and mammalian cells, is initiated by the formation of an isolation membrane followed by its expansion, the engulfment of cytoplasmic content into a closed autophagosome and its fusion to the lysosomes and degradation of autophagosomal content. Beyond its importance in the starvation response, starvation and mTor-independent autophagy is increasingly recognized as an important quality control mechanism that reduces degeneration of neurons and photoreceptor cells and has implications for cancer and infectious diseases. Therefore, the distinct cellular signaling pathways that adjust the rate of autophagy to the cell’s physiology are important to understand. Because excessive autophagy is lethal to cells, the different signaling pathways inducing autophagy must be careful coordinated and calibrated. For one such pathway, the Acinus protein is as a critical regulator. The Acinus protein integrates signals from multiple pathways to modulate the function of core autophagy proteins and stimulate the induction of starvation-independent autophagy. This grant aims to understand the molecular mechanisms that regulate the levels of Acn protein and its activity. For this purpose, in Aim 1, we propose to define upstream regulators of Acinus including the phosphatases and kinases responsible for regulating its activity and explore their potential as possible drug targets. In Aim 2, we will analyze the mechanistic link between Acinus and its effector Atg1, the master regulatory kinase of the autophagy pathway. In Aim 3, we will explore physiological consequences of disrupting the Acn-Atg1 signaling module in the context of visual system.

NIH Research Projects · FY 2025 · 2023-09

Hepatocellular carcinoma (HCC) is a major cirrhosis complication producing an alarming rise in mortality. The prognosis for HCC is poor due to extremely high recurrence rate even after curative-intent surgical therapies and limited efficacy of available medical therapies. Given its refractory nature, prevention of HCC in cirrhosis patients will be the most impactful strategy to improve its poor prognosis; however, effective HCC prevention remains a major unmet need. Retrospective and pre-clinical studies have suggested that statins are a viable form of HCC chemoprevention, with a differential effect between lipophilic and hydrophilic statins. Further evidence suggests that statins may modulate HCC risk through Hedgehog and Hippo signaling pathways. However, the clinical validation of statins has been hampered by the requirement for large and lengthy clinical trials to define their clinical utility. To overcome these challenges, we have developed a serum-based HCC risk biomarker, the Prognostic Liver Secretome signature (PLSec). Of note, PLSec is therapeutically modifiable and the magnitude of PLSec modulation is associated with future HCC incidence as demonstrated by our previous and preliminary studies. In a retrospective case-control series, PLSec-based HCC risk level was lower in cirrhosis patients on statins compared to non-users. Based on these observations, PLSec is now being tested as a surrogate endpoint in HCC chemoprevention trials of atorvastatin (TORCH trial). To achieve the goal of establishing statins as viable HCC chemoprevention with PLSec as a surrogate endpoint, we have assembled a team of cirrhosis and HCC experts to analyze serum samples from three nation-wide multi-center prospective cohorts (Liver Cirrhosis Network, Southern Liver Health Study, and Mass General Brigham cohorts) and two randomized controlled trials (TORCH and LCN RESCU trials). Aim 1. Validate lower biomarker-based HCC risk level in cirrhosis patients on statins compared to non-users. We will validate our preliminary finding in prospective case-control series of cirrhosis patients form the three cohorts. We will explore patient characteristics and types of statins associated with the PLSec-based HCC risk level, along with mechanistic markers of Hedgehog/Hippo signaling. Aim 2. Determine magnitude of biomarker-based HCC risk modulation after starting or stopping statins. We will conduct target trial emulation mimicking single-arm clinical trials with statins to determine the magnitude of PLSec modulation in patients who start or stop statins from three cohorts. We will explore patient characteristics and types of statins associated with PLSec-based HCC risk modulation, along with mechanistic markers. Aim 3. Compare biomarker-based HCC risk modulation between lipophilic and hydrophilic statins. We will compare the magnitude of placebo-adjusted PLSec modulation between lipophilic (atorvastatin) and hydrophilic (rosuvastatin) statins by analyzing serum samples from two parallel randomized clinical trials. We will explore patient characteristics associated with differential PLSec modulation, along with mechanistic markers. Our strategy showcases a novel approach to substantially advance clinical translation of HCC chemoprevention therapies.

NIH Research Projects · FY 2025 · 2023-09

Project Summary/Abstract Over the past several years, concerns have escalated across the globe regarding the lack of knowledge about the long-term effects of gender-affirming hormone therapy (GAHT; puberty suppression, estrogen, testosterone) for transgender and gender diverse (TGD) youth, resulting in widespread legislation and policy recommendations restricting TGD youth’s access to this treatment. Simultaneously, research demonstrating short-term improvements in psychosocial functioning (e.g., mental health (MH) and quality of life (QoL)) among TGD youth receiving GAHT has been steadily accumulating and standards of care continue to support their use. Unfortunately, studies remain limited by short follow-up periods and small sample sizes. As the volume and diversity of youth presenting for care is growing, concerns over the stability of TGD youth’s identities and the likelihood they will regret treatment are also increasing. However, lack of research on within-group differences in TGD youth’s psychosocial trajectories while receiving GAHT leaves providers with little guidance on how to individualize care. More broadly, research is limited by poor measurement of gender dysphoria and lack of inclusion of minority stress and resilience (MSR) factors, which are particularly pronounced in the current sociopolitical climate. The functioning of parents of youth receiving GAHT has also been overlooked despite the critical role parents play in the mental health of TGD youth. The proposed project leverages the longest-running study of TGD youth receiving GAHT in the United States to address each of these critical gaps in the literature. This study is being conducted at the first and largest multidisciplinary gender-affirming care program for TGD youth in the Southwest US and has enrolled nearly 700 families since the program was founded in 2014. The aims of the project are to (1) characterize the 5- and 8-year trajectories of psychosocial functioning among TGD youth receiving GAHT (including gender dysphoria, MSR, MH, QoL) and their parents (including MSR and QoL), including how these trajectories influence each other over time, (2) identify and explore the characteristics of subgroups of individuals who share similar baseline and longitudinal experiences of gender dysphoria (youth) and MSR (youth, parents), and (3) assess the temporal relationships between MH (youth), QoL (youth and parents), and sociopolitical stress, including factors that moderate the impact of this unique form of stress. We will also explore the characteristics and trajectories of youth who experience changes in their gender and/or stop treatment. The project will be guided by a community-based participatory approach that will collaborate with TGD youth and their parents throughout the research process, including in the selection of additional measures to better capture gender dysphoria and sociopolitical stress. Given TGD youth are at heightened risk of experiencing a broad range of adverse psychosocial outcomes, identifying methods to relieve distress and promote healthy development is particularly urgent. The proposed project will provide critical empirical guidance on how to do so.

NIH Research Projects · FY 2025 · 2023-09

Summary Cholesterol is an essential lipid that plays an important role in the maintenance of membrane rigidity and permeability and is essential for the growth and viability of mammalian cells. Cholesterol also functions as a precursor in the biosynthesis of steroid hormones, bile acids, vitamin D, and is a Hedgehog signaling transducer. In this project, we will employ cell biological and structural approaches to study the membrane proteins involved in cholesterol signaling (Hedgehog signaling pathway), biosynthesis (cholesterol synthetic enzymes), and storage (cholesterol esterification enzymes). 1) Dysregulation of Hedgehog (HH) signaling, which is required for proper embryonic development and adult tissue homeostasis, leads to tumorigenesis. Over the past five years, we have determined the structures of human PTCH1 alone, human PTCH1-HH complexes, SMO-Gi complexes in distinct states, and DISP1 alone and in complex with HH. These structures, along with our functional studies, provide molecular insights into HH signal transduction. In this project, we will continue to work on this pathway. Specifically, we will focus on the trafficking and signal regulation of the HH ligand via the HH-PTCH1 axis. 2) Eighteen enzymes convert acetyl-CoA into lanosterol, the first sterol-like intermediate in a series of reactions that synthesize cholesterol in the endoplasmic reticulum (ER). One key reaction is mediated by HMGCR (3- Hydroxy-3-methylglutaryl coenzyme A reductase), which catalyzes the conversion of acetyl-CoA to mevalonic acid. HMGCR is a target of the cholesterol-lowering drugs statins due to its role as the rate-limiting enzyme in cholesterol synthesis. UBIAD1 stabilizes HMGCR and prevents its degradation, while the binding of the E3 ligase gp78 and Insig trigger the degradation of HMGCR. Recently, we determined the cryo-EM structures of HMGCR bound to UBIAD1. The successful completion of this project will aid our investigation into how HMGCR is ubiquitinated by gp78 via Insig. In addition, we will study how the membrane-embedded cholesterol synthetases carry out the cholesterol biosynthesis. 3) In the ER, the ACAT enzymes (ACAT-1 and ACAT-2) catalyze the transfer of long-chain fatty acyl groups to cholesterol, generating cholesterol esters that are integrated into lipoproteins for secretion or storage in lipid droplets. This reaction results in a low cholesterol concentration in the ER and is crucial for maintaining intracellular cholesterol homeostasis. ACAT-1 is ubiquitously expressed in tissues, whereas ACAT-2 expression is restricted to the liver and intestine. Pharmacological inhibition of ACAT- 2 reduces blood cholesterol levels and atherosclerosis. Using structural methods, we plan to investigate mechanisms for lipid-mediated regulation of ACATs to gain insight into novel modes of inhibition and use this information to develop small molecules that specifically inhibit ACAT-2 and lower plasma LDL cholesterol.

NIH Research Projects · FY 2025 · 2023-09

Summary/Abstract Leukemia stem cells (LSCs) promote therapeutic resistance and poor clinical outcomes in acute myeloid leukemia (AML). Central to the function of LSCs is a capacity for aberrant self-renewal, but the mechanisms underlying this activity are not well understood. The long-term goal is to identify these mechanisms to develop new therapies that can eradicate LSCs to improve clinical outcomes. The overall objectives in this application are to (i) determine if LSCs from specific genetic subtypes of AML are dependent on regulation of protein synthesis, (ii) determine whether LSCs in high-risk hematopoietic stem cell (HSC)-like AMLs are more dependent on regulated protein synthesis, and (iii) test a novel therapeutic strategy inhibiting protein synthesis in LSCs. The central hypothesis is that LSCs aberrantly self-renew by adopting from normal HSCs a dependence on tightly regulated protein synthesis. The rationale for this project is based on the finding that the cell surface marker CD99 is selectively overexpressed on LSCs and serves to regulate protein synthesis to promote LSC function. This offers a strong scientific framework by which new strategies to deplete LSCs can be developed. The central hypothesis will be tested by pursuing three specific aims: 1) Determining the role of regulated protein synthesis in promoting LSC function; 2) Determining if the cell-of-origin of AML influences the dependence of LSCs on regulated protein synthesis; and 3) Determining if inhibition of protein synthesis can deplete LSCs in high-risk AML. In the first aim, genetically engineered mice will be used to generate models of AML lacking CD99, to test if this leads to dysregulated protein synthesis that impairs LSC self-renewal. LSCs from these models will be evaluated to determine if they require low protein synthesis rates to prevent induction of tumor suppressors, the unfolded protein response, and the integrated stress response. Ribosome profiling will be performed to identify key LSC regulators selectively translated in the context of regulated protein synthesis. In the second aim, we will generate a mouse model of HSC-like AML which mimics high-risk human AML. We will assess if LSCs in HSC- like AML exhibit heightened sensitivity to dysregulated protein synthesis. These studies will be complemented with an evaluation of protein synthesis in HSC-like human LSCs to determine if they also require maintenance of low levels of protein synthesis. The third aim will test if the combination of a ribosome biogenesis-inhibitor with a BCL2-inhibitor currently used to treat AML can eradicate LSCs in high-risk HSC-like AML. The proposal is innovative, in the applicant’s opinion, because it aims to leverage a novel LSC-specific cell surface marker to establish a new paradigm for understanding mechanisms underlying LSC self-renewal. The proposed research is significant because it is expected to provide a strong scientific justification for the development of therapies inhibiting protein synthesis to overcome therapeutic resistance in patients with high-risk AML. Ultimately, the knowledge gained from these studies may offer insights into the mechanisms that promote the function of cancer stem cells in general, opening up opportunities for the development of new strategies to treat cancer.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Nearly all provider groups in Medicare’s five-year Oncology Care Model alternative payment program expressed a goal to reduce hospital use by cancer patients, but very few achieved this. Identifying potentially avoidable hospital care for cancer patients using diagnosis codes is difficult: depending on the definition used, 20-60% of hospital visits may be avoidable. The leading diagnosis code-based definition is the chemotherapy outpatient quality measure (OP-35), which collects emergency department (ED) and inpatient admissions with ~300 discharge diagnosis codes into 10 avoidable conditions. Unlike similar measures of avoidable hospital care for general patients, OP-35 has not yet been clinically validated. While OP-35 allows payers to compare groups of providers, two issues limit its usefulness to cancer providers: First, clinicians might agree that some OP-35 conditions (e.g. nausea/vomiting) are treatable in an outpatient or urgent care setting, but that others, such as hematemesis (bloody vomiting), would be difficult to evaluate outside of a hospital. Second, OP-35 reports only a percentage of hospital visits to each provider group, obscuring what exactly is driving avoidable hospital use. Based on preliminary work, we propose to develop a classification of actionable scenarios leading to hospital care (e.g. patient required non-emergent procedure; patient did not call for triage help beforehand) so that cancer providers can better understand how to reduce this frequent, disruptive, and costly aspect of treatment. We will assemble an integrated dataset from tumor registry, electronic medical record (EMR), and regional health information exchange data, for a diverse sample representing a range of cancers across all insurance types, including the uninsured. This dataset will identifiably link >75% of all hospital visits in a 100- mile radius of Dallas, TX, to the EMR of three large health systems in the region. Our aims are: Aim 1: Clinically validate diagnosis code-based measures of avoidable hospital care (including OP-35) with clinician EMR review; re-categorize hospital visits into actionable scenarios; and specify a new measure for oncology urgent care-treatable conditions. H1: Most OP-35 defined avoidable will not be avoidable based on clinician review. H2: Actionable categories of clinical scenarios will be identifiable in the EMR, and can be further specified by a measure that identifies conditions treatable in an urgent care setting. Aim 2: Prospectively validate our actionable categories and new oncology urgent care-treatable conditions measure with patients and ED clinicians using post-discharge interviews. H1: Patients and ED clinicians will largely agree with our categorizations, with some refinements. Aim 3: Conduct a national survey of cancer provider groups to assess the feasibility and applicability of our new definitions for avoidable hospital care, in the context of their acute care management capabilities. H1: A broad range of cancer providers will find our definitions feasible and useful. Findings from our study will advance quality measurement and data-driven care improvement, and will be especially useful to participants in Medicare’s upcoming Enhancing Oncology Model payment program.

NIH Research Projects · FY 2025 · 2023-09

Alcohol-associated liver disease (ALD) is a major public health problem and the most common cause of death from cirrhosis in the United States. Despite the high burden of disease, major gaps remain in understanding the natural history of ALD in contemporary U.S. populations. Most data on ALD progression come from European cohorts, while prognostic studies have largely focused on either the risk of developing ALD among drinkers or short-term prognosis in severe ALD. Few studies have comprehensively examined long-term outcomes and the clinical, metabolic, and biological factors that influence prognosis once ALD is established. Although genetic variation (e.g., PNPLA3) has been implicated in risk for developing ALD, less is known about how genetics influence disease progression and outcomes after diagnosis. Similarly, the contributions of patient-level clinical features such as metabolic syndrome, alcohol use patterns, and comorbidities to heterogeneity in ALD progression are incompletely defined. The central hypothesis of this proposal is that a combination of clinical, behavioral, and biological factors underlie the variability in natural history and outcomes among patients with ALD. To test this hypothesis, I will leverage a well-characterized U.S. cohort to pursue the following specific aims: 1) Define the role of clinical and behavioral factors in ALD severity and prognosis; 2) Examine the association of genetic factors with ALD progression. 3) Derive a multilevel risk stratification model to improve prognostication in ALD. This research will establish a comprehensive natural history framework for ALD in the U.S., identify predictors of progression, and support the development of risk stratification tools that can directly inform clinical care and clinical trial design. The PI is a clinical researcher and hepatologist at UT Southwestern with a long-term vision of improving care for patients with ALD through rigorous clinical and translational research. The proposed training plan is integrated with the research aims and builds on his existing expertise in clinical research, while providing advanced training in quantitative analysis, genetics, cohort building, survey methods, and machine learning for risk prediction. He has assembled an exceptionally talented interdisciplinary team of mentors with complementary expertise: Dr. Mack Mitchell, an experienced researcher and ALD content expert; Dr. Amit Singal, a world-renowned health services researcher; Dr. Helen Hobbs, an international expert in genetics and liver disease; Dr. King, an expert in alcohol use disorder; Dr. Zhang, an expert in quantitative analyses; Dr. Kozlitina, an expert in genetic statistics; and Dr. Sandikçi, an expert in machine learning and risk prediction. The proposed studies have significant public health impact as they will fill critical gaps in our understanding of the natural history and prognosis of ALD. This award and training plan will provide the PI with the protected time, advanced skills, and mentorship necessary to develop into an independent investigator focused on improving outcomes for patients with ALD.

NIH Research Projects · FY 2025 · 2023-09

This proposal seeks to determine how TDP-43 protein aggregates dysregulate P-body function in neurons and subsequently produce neurotoxicity. Cytoplasmic aggregation of TDP-43 has been reported in nearly every age-dependent neurodegenerative disease, including in >40% of frontotemporal dementia (FTD), in the hippocampal neurons of Alzheimer’s disease (AD) patients, in >90% of ALS. It also defines a recently recognized AD-like dementia in the oldest elderly, an AD-like syndrome named Limbic-predominant Age- related TDP-43 Encephalopathy (LATE). We have identified that TDP-43 cytoplasmic aggregates regulate the liquid-liquid phase separation (LLPS) of RNA processing bodies (P-bodies) in neuron-like cells and postmortem spinal cord motor neurons in ALS patients. P-bodies are cytoplasmic membraneless ribonucleoprotein (RNP) granules composed of RNAs and protein complexes involved in translational repression and mRNA decay. Neurons carry a high number of P-bodies in the soma. We hypothesize that TDP-43 aggregation causes neuronal toxicity by disrupting the morphology and function of P-bodies. Our proposal is highly innovative because how TDP-43 proteinopathy regulates the LLPS of other membraneless organelles has not been reported in vivo. We propose to use cutting-edge imaging, proteomic, and sequencing approaches to determine the protein and RNA composition of neuronal P-bodies and how it changes in response to TDP-43 aggregation in vivo. We will first determine how TDP-43 cytoplasmic aggregates initiate P-body disassembly and then determine the RNA metabolism change in neurons carrying TDP-43 aggregates or defective P-bodies. Lastly and importantly, we will determine whether P-body proteins and RNA can serve as pathological markers for AD-related dementia, such as LATE.

NIH Research Projects · FY 2025 · 2023-09

Abstract The remodeling of the cervix to prepare for birth begins early in pregnancy and is orchestrated by a precisely timed change in the structure and mechanical function of the cervical extracellular matrix (ECM). Understanding the molecular pathways that alter the ECM in pregnancy while maintaining a synthesis and degradation equilibrium is necessary to discern cervical function and how it mechanically protects the fetus. More importantly, it is critical to determine when and how these cervical ECM synthesis and degradation processes are perturbed, which compromise cervical mechanical function and creates a risk for preterm birth. Our groups prior work provides evidence 1) of rapid collagen turnover rates in both nonpregnant and pregnant cervix 2) dysfunctional cervical extracellular matrix leads to disrupted cervical biomechanical function in novel mouse models 3) cervical mechanical properties evolve in distinct stages in concert with shifts in ECM structure and 4) mouse models of cervical dysfunction are a useful tool to evaluate cervical mechanics. Building on these findings, we aim to better define molecular pathways of collagen degradation and elastic fiber elastogenesis that ensure ECM homeostasis in physiologic remodeling. Secondly, we aim to understand the complexity of mechanical parameters derived from collagen and elastic fibers at multiple length scales. Collectively, the proposed studies, based on compelling preliminary data, can expand our understanding of basic mechanisms in cervical biology and mechanobiology and expand the possibilities for clinical intervention in PTB.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY Large gaps in current strategies for risk stratification and surveillance contribute to frequent late-stage detection and poor outcomes in patients with hepatocellular carcinoma (HCC). Our Translational Liver Cancer (TLC) Research Center has made important scientific contributions that directly addressed HCC risk stratification and surveillance in patients with cirrhosis. Specifically, we conducted a series of phase II and phase III biomarker studies in patients with cirrhosis to validate 1) the first blood-based biomarker for risk stratification (PLSec-AFP), 2) abbreviated MRI for HCC surveillance, and 3) two biomarker panels, GALAD and Doylestown Plus for HCC surveillance. We also conducted modeling studies to evaluate how these data can be incorporated into clinical practice, evaluating the cost-effectiveness of a precision surveillance strategy in these patients. For our TLC renewal, we leverage our infrastructure and operational expertise to similarly develop an optimized, evidence-based approach to early HCC detection in patients with indeterminate liver nodules (ILNs). Our preliminary data demonstrate that patients with ILNs have an annual HCC risk of 6-10%/year, more than double that of those with cirrhosis without ILNs; however, they experience wide variation in HCC risk and surveillance strategies – with some patients who develop HCC failing to undergo surveillance in the year prior to diagnosis and some patients undergoing repeated CT/MRI-based surveillance despite never developing HCC. Our work highlights the need for accurate risk stratification and surveillance strategies in patients with ILNs to optimize the overall value of early HCC detection programs – gaps that are directly addressed by our proposal. We will leverage our Early Detection Research Network (EDRN)-funded Clinical Validation Center for HCC to efficiently recruit a large cohort of patients with ILNs and (1) validate the effectiveness of a novel biomarker- based risk stratification model, (2) evaluate the effectiveness of surveillance abbreviated MRI and contrast enhanced ultrasound for detecting early-stage HCC, and (3) compare the cost effectiveness of surveillance strategies including a precision screening model in patients with ILNs. Our proposal aligns with the principles of precision medicine and would maximize benefits (via early tumor detection) and minimize harms (via false positive results) for each patient, thereby optimizing the patient- centeredness, cost effectiveness, and overall value of HCC surveillance in patients with ILNs. In addition to our patient cohorts and platform of unique biomarker and imaging data, our TLC research center will contribute methodological expertise in HCC early detection, biomarker validation, and HCC imaging to trans-network projects. Overall, our proposal will transform our approach to early HCC detection in patients with ILNs by validating evidence-based and cost-effective strategies to optimize HCC risk stratification and surveillance.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT An estimated 2-8 million people in the United States struggle with primary muscle tension dysphonia (pMTD)— a functional voice disorder that adversely impacts daily living, occupational productivity, and quality of life and results in $2 billion in lost annual productivity costs due to absenteeism. Although pMTD leads to the same emotional, social, financial, and occupational hardships as organic, structural, and neurological voice disorders, its pathophysiology is poorly understood. These gaps make it difficult to understand what causes symptom com- plaints of vocal fatigue, vocal tract discomfort, odynophonia, and difficulties projecting or speaking for prolonged periods in patients pMTD—especially in the context of a lack of overt structural or neurological laryngeal abnor- malities. Although muscle tension and hyperfunction in the laryngeal system is the most proposed mechanism underlying symptoms, it is largely theoretical and based on anecdotal observations. Recent studies from the PI’s lab using well-vetted and validated physiological metrics to quantify tension and hyperfunction in the (para)laryngeal muscles found no group differences between patients with pMTD and healthy controls. However, patients with pMTD consistently reported significantly higher vocal effort, vocal fa- tigue, and vocal tract discomfort with voice use across these studies and self-reported significantly different sensory experiences on measures of interoceptive awareness. These findings suggest altered sensations with voice use in patients with pMTD may be an alternative mechanism underlying symptomology to simply muscle tension or hyperfunctional motor output. We test the central hypothesis that the somatosensory system plays a role in pMTD signs and symptoms across two aims. In Aim 1, we compare laryngeal sensation to pMTD symptom severity in patients with pMTD and controls using laryngeal aesthesiometer sensory testing. In Aim 2, we compare paralaryngeal and widespread musculoskeletal sensation to pMTD symptom severity in patients with pMTD and controls using mechanical pressure and dynamic temporal summation quantitative sensory testing (QST) assays. Outcomes of this proposal will elucidate the role of localized and central sensory mechanisms in pMTD and will lead to improved diagnostics and management for this debilitating voice disorder.

NIH Research Projects · FY 2024 · 2023-09

Abstract/Project Summary Impairment of microcirculatory blood flow has been implicated as a pivotal pathophysiologic event in acutely critically ill patients. As the brain is particularly sensitive to insufficient perfusion, cognitive impairment, particularly in the elderly, can prolong the recovery period and significantly impact the ability to live independently. Resuscitation or perfusion management in critically ill patients is primarily guided by macrocirculatory assessment (i.e., arterial blood pressure (ABP)) with little consideration of the microcirculation, as microcirculatory parameters are technically difficult to assess. Such practice operates under the assumption that resuscitation aimed at correcting macro-hemodynamic variables is also effective in correcting microcirculatory perfusion and oxygen delivery to the brain tissue, a relationship termed hemodynamic coherence. However, recent studies in pathophysiologic states (e.g., sepsis, shock) strongly suggest that microvascular perfusion is not restored despite the optimization of macrocirculatory parameters. Thus, there is an urgent need to learn what governs cerebral microcirculatory flow, dissect conditions that compromise hemodynamic coherence and define effective microcirculation resuscitation targets. Further, while research tools exist, there is no commonly accepted technique which can measure microcirculation through the intact skull. Therefore, it is essential to identify more accessible microcirculatory beds (e.g., sublingual, retinal) which may serve as surrogates for cerebral microcirculatory flow. Characterizing surrogate microcirculations as flow-biomarkers is a critical step towards practical clinical implementation of microcirculatory targets for resuscitation. Our long-term goal is to develop effective strategies to enhance microcirculatory perfusion and effective oxygen delivery to the brain under critical clinical conditions that require resuscitation. The primary goals of this proposal are to a) investigate the role of the microcirculation in cerebral hemodynamic coherence in the gyrencephalic brain and b) characterize effective cerebral microcirculation proxies that can be developed as non-invasive surrogate biomarkers for bedside clinical management. We will manipulate cardiovascular physiology reflecting common intraoperative scenarios (i.e., hemorrhagic hypotension) and hypothesize that current resuscitation strategies, in particular the use of high concentrations of vasopressors, do not restore microvascular function but lead to long- lasting cerebral microvascular constriction and result in brain injury. In Aim 1 we will assess cerebral hemodynamic coherence and oxygen delivery during induced hemorrhagic hypotension and resuscitation using multimodal microvascular imaging techniques. In Aim 2 we will determine the relationship between a) sublingual and b) retinal and cerebral microcirculatory dynamics and investigate their potential as surrogate biomarkers for bedside cerebral microcirculation perfusion management. Successful completion of this project will delineate resuscitation strategies that maintain microcirculatory perfusion and characterize surrogate microcirculations that can be monitored non-invasively and thereby help minimize brain injury.

NIH Research Projects · FY 2025 · 2023-09

Project summary Postural orthostatic tachycardia syndrome (POTS) is a chronic disorder causing disabling symptoms and substantial loss of productivity. Not all POTS patients are the same, and the causes and significance of different subtypes are not known. For example, some patients may have abnormal regulation of their immune system while others may have problems with the response of their muscles to activity. Exercise training has been shown to be a beneficial treatment for POTS, but about 25% of patients are unable to complete the exercise program. Our research study will examine the different types of POTS by systematically and comprehensively studying a large diverse group of patients. This will include careful assessment of symptoms, blood tests, and heart structure as well as a detailed analysis of the muscle response to exercise. This study will be the first to define the full picture of different POTS subtypes and the relationship between these subtypes. Our studies will allow clinicians to better evaluate and individualize treatment and will guide future POTS research.

NIH Research Projects · FY 2024 · 2023-09

Project Summary My work aims to uncover non-genetic mechanisms that drive cancer cell plasticity. I focus specifically on Ewing Sarcoma, a pediatric cancer driven by a single oncogenic fusion, making it prototypical for cancers whose disease progression likely depends on non-genetic adaptations. During my postdoc thus far, I have developed two complimentary models to study heterogeneity of cell states in Ewing Sarcoma: (1) a quantitative high- resolution imaging assay that uses computer-vision based classification of single cell states within Ewing Sarcoma xenografts in zebrafish, and (2) a bimodal distribution of cell signaling states characterized by differential expression and organization of the scaffolding protein Caveolin-1. While the precise role of Caveolin- 1 in cancer remains controversial, recent works suggest that mechanical cues trigger changes in its localization and activity, implicating Caveolin-1 as a potential integrator of environmental cues and cell signaling. However, the mechanism of response and the ensuing signaling cascades remain to be understood, especially in the context of cancer. Therefore, I will leverage the unique experimental frameworks I have established to test whether Cav-1 acts as a plasticity factor promoting tumor cell adaptation in Ewing Sarcoma. This work will reveal mechanisms of fast cellular adaptation to diverse microenvironmental cues which will provide unique insight into the drivers of metastasis and drug resistance. I am eager to build upon the foundations I have established during my postdoc thus far to discover previously unapproachable mechanisms of cell adaptation. As the proposed work requires cross-disciplinary expertise, my continued development in several areas will be instrumental to my progress. Dr. Danuser and the Danuser lab will provide the ideal environment to develop advanced microscopy techniques and skills in computational analysis of 3D data. Dr. Amatruda will provide guidance and support in the use of zebrafish disease models and relevance to pediatric cancer. Dr. Lamaze and Dr. Cobb will provide scientific insight in caveolar biology, MAPK cell signaling, and cancer biology. Combined with the stellar training environment and resources available at UT Southwestern, this provides the ideal environment to carry out this work. The training I will receive will enable me to lead an independent laboratory that studies cancer cell plasticity in a variety of experimental models, with specific focus on imaging-based approaches and physiologically relevant environments.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Hypertension remains a leading contributor to cardiovascular disease-related morbidity and mortality in the United States. Older (i.e., ≥60 y) women are one group more likely to develop hypertension with inadequate blood pressure (BP) control, despite optimal drug regimens. Older women are also at a greater risk for the development of Alzheimer’s disease and related dementias which are exacerbated by hypertension. Accordingly, the prevalence of hypertension and ineffectiveness of drug treatments alone signal the need for non-pharmacological approaches to supplement standard care for BP control in this population. One such approach to achieve this is “heat therapy.” Recent studies have revealed the promising benefits of heat therapy on vascular health, autonomic activity, and cardiac function, which may ultimately reduce BP and the risk for cardiovascular diseases. These physiological adapta- tions may also translate to improvements in cognitive function through improved cerebrovascular func- tion and health. However, no information exists regarding the efficacy of heat therapy to reduce BP, induce meaningful neural-cardiovascular adaptations, and improve cognition in older women. The overall goals of this proposal will be to 1) identify changes in autonomic BP regulation and 2) assess alterations in cerebrovascular and cognitive function in older women with mild hyper- tension following 8 weeks of at-home heat therapy. Specific Aim 1 will investigate chronic lower leg heat therapy's impact on BP control and neural-cardiovascular function in older, hypertensive women using intervention and control groups. The intervention group will perform 8 weeks of at-home lower leg heat therapy using hot water immersion up to the knee (42°C, 45 min/session, 4 sessions/week). The control group will complete the same sessions with thermoneutral water (35°C). We will assess 24-hour ambulatory BP, sympathetic vascular transduction, cardiac function, and heat shock protein expression before and after the interventional period. Specific Aim 2 will measure cerebral hemody- namics and cognitive function responses to chronic lower leg heat therapy in older, hypertensive women. We will use transcranial Doppler ultrasound to assess cerebrovascular function and autoreg- ulation before and after 8 weeks of lower leg heat therapy. Further, we will use the NIH Toolbox Cog- nitive Battery to assess different cognitive function domains. To maximize the clinical relevance of this project and my scientific training, I have assembled a strong interdisciplinary research team comprised of Qi Fu, MD, PhD, Rong Zhang, PhD, Steven Romero, PhD, Heidi Rossetti, PhD, and Satyam Sarma, MD. This team will help me complete the proposed project and master several technical skills (e.g., microneurography, echocardiography, biomolecular assays), while improving my ability to obtain extra- mural research funding (e.g., NIH K99/R00) and become a successful independent clinical researcher.

NIH Research Projects · FY 2024 · 2023-09

Summary Hepatocellular carcinoma (HCC) is the leading cause of death in patients with cirrhosis, and the fastest rising cancer mortality in the U.S. Due to the limited efficacy of existing therapies for established HCC tumors, prognosis for patients remains poor, with five-year survival under 15%. Thus, HCC chemoprevention in cirrhosis is likely the most impactful strategy to improve survival. However, despite the candidate chemopreventive agents suggested in experimental studies, it remains an unmet need due to logistical difficulty in conducting clinical trials that require large sample size and long follow-up time. To overcome the challenge, we identified Prognostic Liver Secretome signature (PLSec) to quantitatively monitor therapeutic modulation of HCC risk level in cirrhosis patients, and predict reduction of future incident HCC. PLSec has been used as a surrogate endpoint in our ongoing and planned HCC chemoprevention clinical trials. Experimental studies in rodent models by us and others suggested that epigallocatechin gallate (EGCG), a green tea catechin, prevents HCC development without any adverse events. Our ex vivo organotypic culture of precision-cut liver slice (PCLS) from cirrhosis patients revealed suppression of high-risk signature by EGCG, supporting its clinical relevance. Based on these promising findings, the goal of our proposal is to test our hypothesis that EGCG treatment safely suppresses PLSec in patients with cirrhosis. Aim 1. Evaluate safety and efficacy of EGCG in cirrhosis patients (phase II double-blinded placebo-controlled clinical trial). We will evaluate 24-week EGCG treatment or placebo in 60 patients (1:1 randomization) with early-stage cirrhosis enriched for elevated HCC risk by a clinical variable-based score (FIB-4 index) and PLSec. Participants will be monitored monthly for adverse events. Serum samples will be obtained before, during, and at the end of treatment. Primary endpoint: reduction of risk level as measured by PLSec (delta-PLSec). Secondary endpoints: safety profile, change in quality of life. Exploratory endpoints: change in on-treatment PLSec, immunohistochemistry of HCC-risk-related markers for participants consented for liver biopsy, and incident HCC. Aim 2. Identify factors associated with response to EGCG in cirrhosis patients. We will evaluate pre-treatment PLSec and clinico-histological variables; on-treatment PLSec modulation and plasma concentration of EGCG and its metabolites for their association with the primary endpoint. We will also assess modulation of the FIB-4 index and liver stiffness measurement by acoustic elastography as potential alternative clinical endpoints to monitor effect of EGCG, We expect to establish novel HCC chemoprevention with a dietary supplement for subsequent pivotal phase III clinical trial toward clinical translation of this approach, which will contribute to a transformative improvement in the outcome of patients with HCC by enabling individual- risk-based, molecular-targeted, and safe chemoprevention of this deadly cancer.

NIH Research Projects · FY 2026 · 2023-09

Project Summary The nervous system was traditionally thought to act independently of an organism’s immune response and be an “immune privileged site”. Increasing scientific evidence has shown that the nervous system is not immune privileged but instead has a unique immune response that is critical for maintaining homeostasis and critical for central nervous system (CNS) function. Much of the work in the field of neuroinflammation has focused on the function of microglia, but little is known about the role of neurons in modulating neuroinflammatory responses in the CNS. Five years ago, we discovered that the neuronal protein, alpha-synuclein(asyn), was critical in protecting neurons from viral infection. We have extended these data to show that ayn modulates type 1 interferon (T1IFN) signaling. Asyn is known as a cause of Parkinson’s disease (PD) and is known to be dysregulated in neurodegenerative diseases, traumatic brain injury, and other diverse CNS diseases. Despite the importance of asyn in CNS disease states, the functional role of asyn expression is not well understood. We have discovered that asyn expression is necessary to support expression of specific interferon stimulated genes (ISGs) in the brain during T1IFN signaling, independent of microglia activation. Using induced pluripotent stem cells (iPSC) and CRISPR-mediated SNCA deletion to create asyn KO human dopaminergic neurons, we found that viral growth in neurons is inhibited in the presence of asyn expression and that viral- induced ISGs such as IFIT1, OAS1, and TRIM25 exbibit decreased expression in asyn KO neurons. We next found that asyn KO neurons exhibit a broad loss of ISG expression following treatment with poly I:C or type 1 interferon (2) treatment due to loss of asyn-dependent STAT2 activation and asyn nuclear localization. Taken together, our data show for the first time that asyn functions to support interferon responses in neurons. The goal of this proposal is to determine the specific mechanism of asyn-dependent innate immune responses in neurons. We hypothesize that asyn is a novel neuron-intrinsic regulator of the CNS innate immune response. We will test our hypothesis in three aims. Aim 1 will use asyn KO and WT human neurons to define the specific interactions between asyn, interferon signaling, and vesicle transport in neurons. Aim 2 will define the role of neuron-intrinsic asyn production on the innate T-cell response in the brain using an inducible, nestin-Cre-lox knockout of the asyn gene (Snca) in mice. Aim 3 will evaluate PD-specific and species specific changes in asyn that may influence its native function in neurons. Taken together, the proposed studies will significantly advance our understanding of neuron-intrinsic control of the innate immune response in the CNS and provide novel insight into the underlying immunopathogenesis that contributes to diverse human diseases of the CNS.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY Sensory hypersensitivity is a common symptom in autism and Fragile X Syndrome (FXS) and is thought to be a result of cortical circuit dysregulation. EEG studies in humans with FXS and the FXS mouse model, the Fmr1 KO, reveal cortical circuit hyperexcitability and synchrony deficits such as enhanced resting state power in the gamma band and reduced sensory-driven synchrony. In acute slices, this hyperexcitability can be observed as prolonged persistent activity states, called UP states, and increased gamma band power during UP states. I hypothesize that circuit mechanisms that mediate hyperexcitability and prolonged UP states in the neocortex may contribute to EEG phenotypes in FXS. Using positive and negative allosteric modulators (PAMs/NAMs) specific for GluN2C/D subunits of NMDA receptors, I have revealed an upregulation of GluN2C/D function in the Fmr1 KO cortex that contributes to circuit hyperexcitability. Specifically, GluN2C/D PAMs increase UP state duration and gamma power during the UP states, while NAMs rescue UP state duration. Remarkably, these interventions only affected the Fmr1 KO, not their wildtype (WT) littermates suggesting that GluN2C/D function is upregulated in the Fmr1 KO and leads to cortical circuit dysfunction. Typically, GluN2C/D subunits are expressed in cortical inhibitory neurons and astrocytes. Since my results are not consistent with effects on inhibitory neurons, GluN2C/D subunits may be misexpressed in excitatory neurons or upregulated in astrocytes in the Fmr1 KO. I hypothesize that GluN2C/D expression and/or function is increased in excitatory neurons and/or astrocytes in Fmr1 KO mice and this contributes to hyperexcitability and altered synchrony of cortical circuits. The goal of the proposed project is to test this hypothesis and determine expressional and functional changes in Glun2C/D that may contribute to cortical hyperexcitability and synchrony following three Specific Aims. Aim 1. To determine change in cortical protein and RNA expression levels as well as cell specific expression of GluN2C/D subunits in Fmr1 KO cortex. Aim 2. To determine cell specific functional contribution of GluN2C/D subunits to NMDA-mediated currents in WT and Fmr1 KO cortex. Aim 3. To determine the contribution of GluN2C/D NMDARs to in vivo sensory driven circuit excitability and altered synchrony using multi-electrode array EEG and measurement of audiogenic seizures in the Fmr1 KO mouse in collaboration with Dr. Devin Binder at UC Riverside. These experiments will not only provide insights into the molecular and cellular basis of GluN2C/D contribution to cortical circuit dysfunction, but also examine GluN2C/D subunits as a potential target for therapeutic development using translational biomarkers.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Hispanic adolescents in the United States are disproportionately affected by obesity compared to their non- Hispanic White adolescent counterparts. A recommended means of addressing adolescent obesity is through participation in family-based multicomponent behavioral interventions (targeting improvements in family functioning as a mechanism of change). Although family-based interventions (FBIs) exist, effects on adolescent obesity-related outcomes, especially among Hispanic adolescents, are small or insignificant. Limited effects may be due to gaps in the existing research: 1) few FBIs have incorporated obesity-related socioecological factors at multiple levels as tailoring variables to improve intervention effects, 2) FBIs often overlook fathers and other household members (such as grandparents in multi-generational households) that may also play a role on adolescents’ obesity risk behaviors, and 3) FBIs often assess family functioning retrospectively without consideration of day-to-day family dynamics, which may also influence obesity risk behaviors. Thus, I propose the following aims to address existing gaps: 1) conduct secondary data analysis using five waves of data from the Adolescent Brain Cognitive Development study and the Hispanic adolescent subsample (n=2411, Mage=9.5 at baseline) to examine the direct and moderating effects of factors at each level of the socioecological model on the longitudinal trajectories of obesity risk behaviors/obesity status, 2) pilot an EMA protocol with Hispanic adolescents (9-to-17 years) and household caregivers (n=20 family units) across 7 days to assess feasibility and acceptability, identify barriers and facilitators to completion of daily assessments by family units, and make needed modifications to the EMA protocol, and 3) implement a finalized EMA protocol over a 6-month period, using measurement burst design, and assess the effects of momentary changes in family functioning behaviors and associations with daily physical activity, sleep, and nutrition behaviors (n=50 family units, adolescents 9-to-17 years). The purpose of this training application is to gain mentoring and training in the following four areas: 1) intensive longitudinal data analysis, 2) recruitment, retention, and implementation, 3) innovative assessment design, and 4) professional development. Under the mentorship of a team of interdisciplinary researchers, experts in either obesity prevention, longitudinal data analysis, ecological momentary assessment, and/or minority health, Dr. Fernandez will successfully complete the plan of research at the University of Texas Health Science Center at Houston, School of Public Health and attain preliminary data to inform the development of a NIH-R01 proposal.