Baylor College Of Medicine

NIH Research Projects · FY 2025 · 2025-07

ABSTRACT Multiple Sclerosis (MS) is one of the most common autoimmune diseases affecting the central nervous system, impacting approximately 2.5 million people worldwide. It is characterized by immune cell-driven inflammatory demyelination and axonal degeneration, leading to a spectrum of motor, sensory, and cognitive deficits. A key pathological hallmark of MS is increased blood-brain barrier (BBB) permeability. The BBB, composed of multiple cell types, including astrocytes, is critical for maintaining CNS homeostasis. Astrocytic dysfunction has been implicated in BBB impairment across several CNS disorders. Recent evidence suggests that Slc4a4, an astrocyte-enriched gene, plays an essential role in regulating BBB integrity during brain injury. However, its involvement in MS pathology and progression remains unexplored. To address this, we combined a novel astrocyte-specific Slc4a4 conditional knockout (Slc4a4-cKO) mouse model with the experimental autoimmune encephalomyelitis (EAE) model, a well-established MS model that recapitulates key clinical and pathological features of the disease. Our preliminary data reveal that Slc4a4-cKO mice exhibit more severe EAE clinical symptoms, including heightened blood-protein leakage, reduced expression of endothelial tight junction markers, and increased demyelination. Furthermore, bulk RNA sequencing of Slc4a4-cKO mice post-EAE induction highlights an enrichment of genes associated with endothelial cell dysfunction, although the underlying mechanisms remain unclear. To bridge this knowledge gap, I propose three aims to investigate the role of astrocytic Slc4a4 in MS progression. First, I will characterize Slc4a4 expression in human MS tissue and assess EAE pathology across different disease stages in Slc4a4-cKO mice. Next, I will explore whether overexpression of astrocytic Slc4a4 confers therapeutic benefits in EAE. Finally, I will examine the role of Edn1, a candidate gene upregulated in Slc4a4-cKO mice, along with its associated receptors, in human MS and during EAE in Slc4a4-cKO mice. Additionally, I will test whether pharmacological or genetic inhibition of the Edn1 pathway can rescue the exacerbated EAE phenotypes in Slc4a4-cKO mice. Altogether, these studies aim to uncover the Slc4a4 pathway as a potential mechanism underlying astrocyte-driven BBB dysfunction and MS pathogenesis, potentially identifying new therapeutic targets for this disease.

NIH Research Projects · FY 2026 · 2025-07

PROJECT SUMMARY/ABSRACT Forward genetic screens through chemical mutagenesis in Drosophila have made significant contributions to fundamental discoveries that span multiple biological disciplines and provided novel biological insights that are directly relevant to diverse human diseases. However, due to the difficulty to map causative mutations and its labor-intensiveness, many researchers cannot take advantage of this approach. A collection of fly strains that are obtained from a well-designed chemical mutagenesis screen that simultaneously explores numerous phenotypes can serve as excellent resource for the broader research community since a number of secondary screens could be conducted on the same collection and individual mutant lines can be used to probe diverse biological processes that are evolutionarily conserved. However, such resource does not exist in the Drosophila community and most fly strains isolated from previously conducted chemical mutagenesis screens were left unmapped and subsequently discarded. We performed a large scale F3 clonal EMS-mutagenesis screen on the Drosophila X-chromosome and have been maintaining 1,385 recessive lethal mutant strains. So far, we have molecularly mapped the lethality-causing mutations of 614 lines to 165 genes. This collection was built on a healthy isogenic y w FRT19A X-chromosome using low concentration of EMS and have been selected for mutants that show interesting morphological and/or neurophysiological phenotypes in mosaic animals. Importantly, the careful design of the screen allowed us to enrich for essential genes that are evolutionarily conserved (~90% of mapped genes are conserved in human) and have direct relevance to human diseases (~70% of mapped genes have human homologs that cause genetic diseases in human). Mapped mutations from this collection have been used in >80 manuscripts by many laboratories that study a variety of biological contexts and have also led to discovery of novel human disease genes and mechanisms. Here, we will sequence and attempt to demonstrate the causality of the remaining 771 unmapped lines that have been maintained using a new streamlined sequencing and bioinformatic analysis pipeline followed by rigorous genetic analysis. Upon completion, this project will provide Drosophila researchers with a collection of >1,000 fly strains which are carefully phenotyped that have been generated on a well-defined genetic background and mapped to >300 conserved genes. This resource can be used to perform numerous secondary screens and subsequent deep investigations by numerous laboratories in the US and around the world. Missense mutations identified from this effort can further provide novel structure-function information on conserved proteins. The rich phenotypic information of clonal phenotypes will allow us to establish a comprehensive database that could be mined by Drosophila biologists as well as experts in other disciplines such as clinical geneticists interested in novel disease gene discovery in humans.

NIH Research Projects · FY 2025 · 2025-07

Outcomes have stagnated for patients with locally advanced human papillomavirus (HPV)-negative head & neck squamous cell carcinoma (HNSCC) over the past few decades. Recent studies have shown that ∼15% of all HPV-negative HNSCC patients have altered KEAP1/NFE2L2(NRF2) pathway, which strongly correlates with radioresistance and poor outcomes. Now we know that many factors, such as genetic lesions, pH, oxygenation, and interactions with other cells in the tumor microenvironment, play a critical role in reprogramming cancer cell metabolism. No one has studied Keap1’s radioresistance role in HNSCC using a mouse model that closely recapitulates human disease. Consequently, we developed a novel genetically engineered mouse model (GEMM) that lacks Keap1 (or overexpress Nrf2 downstream targets) for studying HNSCC, allowing us to control the temporal and spatial induction of primary tumors. Using these in vivo models, we showed that even a partial loss of Keap1 resulted in (1) sustained hyperactivation of the Keap1/Nrf2 signaling pathway, (2) increased infiltration of myeloid cells, particularly macrophages & granulocytes, and (3) significant radioresistance of primary HNSCC tumors. Furthermore, we discovered that partial loss of Keap1 increased the radiosensitization of poly ADP ribose polymerase (PARP) inhibitors in vitro. This application aims to understand the role of the tumor immune microenvironment (TIME) in radioresistance and whether PARP inhibitors can overcome this. Aim 1 – Does NRF2 signaling reshape TIME in HNSCC tumors post-RT? We will utilize HNSCC cell lysates and supernatants at baseline and post-RT to quantify the production and secretion of inflammatory mediators, as well as their influence on various leukocyte chemotaxis. This will be achieved using Ibidi µ-slide chambers filled with cell supernatants to create a chemical gradient. Finally, we will quantify the changes in the TIME of primary tumors at baseline and following RT using a pre-validated panel of fluorescently labeled flow cytometry antibody markers for leukocyte subsets. Aim 2 – Investigate whether manipulating the DDR pathway in NRF2 overexpressing tumors could create favorable TIME. We will transiently knockdown or overexpress Nrf2 in HNSCC cell lines, treat them with RT +/- PARPi, and collect cell lysates/supernatants. We will assess (1) DNA damage, (2) DDR, (3) activation of the cGAS-STING pathway, and (4) the release of damage-associated molecular patterns (DAMPs). Finally, we will collect primary tumors following RT +/- PARPi treatment and perform RNA-seq to compute single-sample gene set enrichment analysis scores for different leukocyte subsets. The future research plans – The data and resources generated from this R03 project will provide the necessary framework for an expanded R01 grant application and ultimately lead toward improving HPV-negative HNSCC patient outcomes.

NIH Research Projects · FY 2025 · 2025-07

SUMMARY Prognosis for relapsed acute myeloid leukemia (AML) is dismal (50% relapse rate, 5-year overall survival less than 28%), underscoring the critical need for more effective, less toxic therapies. The most frequently mutated gene in AML is nucleophosmin (NPM1). Mutations in NPM1 (NPM1c+) are leukemia-generating events and often confer a more favorable prognosis (40% survival). Thus, NPM1c+ AML is defined as a distinct leukemia entity. However, the advantage associated with NPM1c+ is lost in older patients, in patients with co-occurring mutations, and in relapsed patients undergoing salvage therapy. Hence, the prognosis of this common type of AML is still very poor compared to other cancers. Therefore, more interventions that target NPM1c+ AML are critically needed. This goal is hindered because the mechanisms underlying NPM1c+-induced leukemogenesis and leukemia maintenance are unclear. In this proposal, the applicant will develop in vivo murine models to investigate the role of caspase-2 as an essential gene in NPM1c+ AML. This premise is based on preliminary and published data from the applicant's laboratory that NPM1c+ activates caspase-2, and that caspase-2 loss leads to cell cycle arrest, terminal differentiation, and downregulation of signaling pathways that regulate stem cell pluripotency of NPM1c+—but not NPM1wt— AML cells. The central hypothesis of this application is that NPM1-c+-induced caspase-2 activation promotes AML self-renewal to drive and propagate leukemia. The specific aims are to: 1) Determine the impact of loss of caspase-2 on self-renewal of NPM1c+ myeloid cells, and 2) Determine how loss of caspase-2 impacts NPM1c+-associated myeloproliferative disease (MPD) and AML. Under the first aim, established mouse models for inducible NPM1c+ will be crossed to caspase-2-deficient mice to evaluate the role of caspase-2 in the pre-leukemic state and stem cell expression. Under the second aim, an inducible Npm1c+ mouse co-expressing a Dnmt3a mutation will be crossed to caspase-2 knockout mice to determine the effect of caspase-2 loss on MPD and AML development and changes to chromatin accessibility associated with NPM1c+/Dnmt3a mutant expression. These experiments will provide the essential murine models and preliminary data to fully investigate caspase-2 as a critical downstream effector of NPM1c+ in AML. This work will allow the applicant to pursue her long-term goal to advance opportunities for AML treatment, and provide critical insight into a novel role for caspase-2 as a key regulator of cancer cell pluripotency.

NIH Research Projects · FY 2026 · 2025-07

Project Summary Elevated intraocular pressure (IOP) is the only modifiable risk factor of glaucoma, and all approved drugs for glaucoma aim to lower IOP but with limited efficacy to prevent retinal ganglion cell (RGC) loss. Mounting evidence suggests that both ocular hypertension (OHT) and vascular abnormalities coordinately contribute to the pathogenesis, whereas molecular mechanisms remain elusive. In the neurovascular unit (NVU), endothelial cells (ECs) and RGCs don’t have direct physical contact but are part of a complex interaction network connected via pericytes and glial cells. The daunting challenge to confirm the role of NVU dysfunction in glaucoma pathogenesis is the lack of EC-specific ligands with minimal binding to RGCs to demonstrate modulation of RGC function and survival through the NVU. To address the challenge, we recently developed an innovative technology of ligandomics to globally map cell-binding ligands with simultaneous binding activity quantification. This technology has led to the discovery of disease-restricted angiogenic factors and development of first-in- class disease-targeted anti-angiogenic therapies by our group. The objective of this project is to unequivocally confirm the contribution of the NVU to glaucoma pathogenesis and identify EC ligands with potential for neuroprotective therapy. Our central hypothesis is that OHT-restricted EC-specific ligands with undetectable binding to RGCs can mitigate or exacerbate RGC dysfunction and loss indirectly via the NVU in glaucoma. In Aim 1, we will globally map all ligands binding to OHT and healthy retinal ECs, RGCs, pericytes and glial cells with simultaneous binding activity quantification by ligandomics. Quantitative comparison of entire ligandomes will systematically identify OHT-selective EC-specific ligands with minimal binding to RGCs and glial cells. In Aim 2, OHT-selective EC-specific ligands will be independently validated their binding to NVU cells to confirm their disease binding selectivity and cell-binding specificity. In Aim 3, OHT-selective EC-specific ligands will be functionally validated for their capacity to mitigate or exacerbate RGC dysfunction and loss in glaucoma eyes. Validated EC-specific ligands capable of modulating RGC function and survival are promising targets to develop novel neuroprotective therapy with minimal side effects on healthy NVU cells. To our knowledge, ligandomics is the only omics technology to systematically identify disease-selective cell-specific ligands and is broadly applicable to all types of cells and neurological diseases with potential for transformative impact on neuroscience research and neuroprotective therapy.

NIH Research Projects · FY 2026 · 2025-07

This project investigates the role of Carbohydrate Responsive Element Binding Protein (ChREBP), a key transcription factor in metabolic regulation and disease, particularly focusing on its isoforms and their effects in the liver. ChREBP has been implicated in metabolic-associated steatohepatitis (MASLD), dyslipidemia, and cardiometabolic diseases, yet the specific functions of its isoforms remain controversial and poorly understood, in part due to conflicting reports published in a variety of gain- and loss-of-function ChREBP mouse models. In this project we aim to resolve the contradictory roles of ChREBP isoforms in liver metabolism, thereby enhancing our understanding of metabolic disease mechanisms. This will serve our long- term objectives of providing foundational principles of liver metabolism that may ultimately lead to improved diagnostic and therapeutic strategies for metabolic disorders including dyslipidemia, diabetes, and MASLD. The study employs genetic interventions (knockouts and overexpression) in mouse models and molecular phenotyping to dissect the metabolic pathways and molecular physiology influenced by ChREBP isoforms. Techniques include GalNAc-siRNA-mediated knockdown, ADV-mediated overexpression, and AAV-TBG- Crispr modifications in mouse models. This approach will enable a detailed analysis of the physiological impacts of ChREBP under various metabolic contexts, offering insights into its dual roles in metabolic diseases. In Aim 1, we aim to define the role of hepatic ChREBP-beta in a model where ChREBP-alpha has been ablated, testing if modest increases in its expression are sufficient to adversely affect liver and systemic metabolism. In Aim 2, we will explore dose-dependent effects of ChREBP overexpression on liver function and systemic metabolism, focusing on glucose homeostasis and mitochondrial dysfunction. Aim 3 investigates the activation of ChREBP by specific carbohydrate metabolites under different dietary conditions, employing Crispr strategies to modify key enzymes in metabolite pathways that may provide metabolites that activate ChREBP.

NIH Research Projects · FY 2025 · 2025-07

Enter the text here that is the new abstract information for your application. This section must be no longer than 30 lines of text. The Houston Area Molecular Biophysics Program (HAMBP) is a research training program for molecular biophysics PhD students in the Houston-Galveston area. It builds on our program in place since 1989. Funding for 11 students is sought to allow continued excellence and innovation. The program is led by the Program Director, Co-Director and a steering committee with representatives from five graduate schools in the area,0(Baylor College of Medicine, Rice University, University of Texas MD Anderson Cancer Center/University of Texas Health Science Center-Houston, University of Houston, and University of Texas Medical Branch at Galveston). The program provides didactic and seminar courses, three annual research conferences with trainee presentations, monthly trainee meetings, career/professional development seminars and workshops, attendance at national meetings, and annual presentation and review of trainee research progress. All students are required to participate in training in the responsible conduct of research and workshops in rigor and reproducibility. Mentors include 41 faculty members at six institutions, affiliated with 17 different departments and 14 graduate programs. These faculty have exemplary training and research records, as well as strong research funding. They have trained 158 predocs and 229 postdocs over the past 10 years and currently have 112 predocs (56 TG-eligible) and 62 postdocs currently in their labs. Trainees join HAMBP after one year of study following selection of one of our faculty mentors as their major thesis advisor. Trainees are selected in a highly competitive process from students whose projects in mentors' laboratories provide training in molecular biophysics. Trainee selection has the goal of supporting the most promising students. Strengths include x-ray crystallography, macromolecular, cryo-electron microscopy, a wide range of spectroscopic and microscopic techniques, single molecule methods, computational biophysics, membrane biophysics, protein folding, nucleic acid structure and ultrastructure, thermodynamics, kinetics and mechanistic enzymology. Supported students generally publish at least 3 papers from their thesis research, including many in high impact journals, and go on to successful, research-related careers in academia, industry, government agencies, and private organizations. Location in the Texas Medical Center and the collaborations among the participating institutions facilitate cutting-edge biophysical research relevant to human health. Our program provides quantitative and interdisciplinary skills and prepares our trainees to be leaders in highly productive careers related to research advancing biomedical science.

NIH Research Projects · FY 2026 · 2025-06

PROJECT SUMMARY Spreading depolarization (SD) is a wave of massive, near-complete cellular depolarization that slowly propagates across brain tissue. Recent decades of clinical studies have convincingly demonstrated that this prolonged depolarization wave occurs under certain conditions in the human brain and can contribute to deleterious acute and chronic neurological deficits. SD is distantly related to seizures, and their co-occurrence has been observed in numerous clinical and experimental settings. However, the majority of the findings are based on experimentally evoked SD, often in anesthetized animals. It remains an open question whether SD spontaneously emerges and contributes to neurological comorbidities in epilepsy patients. In order to address the neurological significance of spontaneous SD, we recently developed a chronic DC-band EEG recording and demonstrated that SD events indeed occur spontaneously in awake genetic and acquired epilepsy mouse models, each exhibiting distinct generation patterns and interactions with seizures unique to the respective models. With advanced multimodal electrophysiological and state-of-the-art behavioral analysis tools, we are now investigating both the neurological consequences and underlying regulatory mechanisms of these pathophysiological depolarizations in hyperexcitable neuronal circuitries. This project will investigate the contributions of SD to epilepsy comorbidities and disease progression in Dravet syndrome, a major example of developmental epileptic encephalopathy (DEE), using a well-established Scn1a deficient mouse model (Scn1a+/R1407X). Leveraging our expertise in acute and chronic electrophysiological monitoring in awake juvenile mice, we will thoroughly characterize the pathological significance of SD events in the developing juvenile Scn1a+/R1407X mice, in which the risk of premature mortality is high, neurobehavioral abnormalities begin to emerge, and spontaneous cortical and subcortical SD are present. Specifically, we will investigate 1) the correlation between SD and cardiorespiratory instability (AIM1), 2) acute and chronic neurobehavioral deficits associated with SD (AIM2), and 3) the contribution of hippocampal SD during early-life hyperthermic seizure to subsequent circuit dysfunctions (AIM3). These studies will investigate the understudied SD pathology in the well-established DS mouse model. These studies will examine the pathological significance of understudied SD phenomenon in the one of the one of the most studied mouse model of DEEs. The experimental methods established in this study will be directly applied to other epilepsy mouse models in our laboratory, further elucidating the general and disease-specific role of SD. The results obtained in this study may also provide insights into other SD-related pathologies such as stroke, traumatic brain injury, and migraine with aura.

NIH Research Projects · FY 2025 · 2025-06

Food insecurity is a major social determinant of health that impacts nearly 20% of youth with type 1 diabetes (T1D). Food insecurity contributes to a number of negative T1D health, dietary, and psychosocial outcomes among youth and their families. Food prescription (Food Rx) programs, which are partnerships between community-based food access programs and healthcare settings to provide healthful food to individuals with chronic health conditions, have strong potential to improve health, dietary, and psychosocial outcomes of youth with T1D. Indeed, Food Rx combined with a healthy eating intervention delivered by Community Health Workers (CHW) has been associated with improved glycemic outcomes among adults with type 2 diabetes. Yet, Food Rx combined with a healthy eating intervention has not been evaluated among youth with T1D and their families from food insecure households. The purpose of this study is to assess the feasibility and preliminary outcomes of a healthy eating program that consists of Food Rx combined with an evidence-based healthy eating family behavioral intervention refined for delivery by CHWs for families of youth with T1D (ages 10-15 years) from food insecure households. The program will be implemented through a partnership between the Texas Children's Hospital's (TCH) Diabetes Care Center and The Houston Food Bank's (HFB) Food Rx Program. The study will be conducted in two phases. Phase 1 is a formative phase where investigators will engage stakeholders consisting of T1D healthcare providers, primary care providers, community health center providers, the HFB Food Rx Program staff, and families of youth with T1D to refine the program for families of youth with T1D from food insecure households who receive care in various healthcare settings such as specialty diabetes clinics, primary care, and community health centers. In Phase 2, the healthy eating program (Food Rx + healthy eating family behavioral intervention) will be evaluated in a randomized pilot trial where families will be randomized 1:1 (intervention: standard diabetes care). Stakeholders will continue engagement with the study team in Phase 2 to finalize the study protocol, problem-solve challenges, and disseminate findings. This study will provide critical preliminary data about the feasibility and preliminary outcomes of a refined healthy eating program (Food Rx + healthy eating family behavioral intervention) for youth with T1D to prepare for a more extensive, fully-powered study.

NIH Research Projects · FY 2026 · 2025-06

PROJECT SUMMARY AND ABSTRACT Stem cells are essential for tissue homeostasis following environmental challenges. During aging however, these stem cells lose this ability to maintain tissue integrity. Dysregulation in the proliferation and the differentiation of stem cells are implicated in several pathologies such as cancer, hematological disorders, and gastrointestinal disease. Intestinal stem cells (ISCs) reside in the digestive tract epithelium and their balance between active and quiescent states is disrupted during aging. Several signaling pathways that are dysregulated in aging ISCs have been characterized, however, the mechanism by which ISCs transit between active and quiescent states during an organism’s lifetime is relatively unknown. Here I propose using Drosophila melanogaster to model the regulation of quiescent and active states with their Drosophila ISC (dISC) population given the excellent orthology with mISCs. Drosophila have the advantage of being inexpensive, reproducing rapidly, short-lived, and having plentiful genetic resources available. There are many datasets available to characterize dISCs and their changes during aging. These resources include single cell sequencing datasets from young and old fly guts developed in the lab: the Fly Cell Atlas (2022 Science) and the Aging Fly Cell Atlas (2022 Science). Using a bioinformatics analysis-guided genetic screen approach I identified an uncharacterized transcription factor required for proper regulation in dISCs, Chronophage (Cph, “time-eater”). This gene is an ortholog of two essential genes in humans: BCL11A and BCL11B, which also have specific expression in mISCs from curation of mammalian scRNA-seq datasets. Preliminary evidence from the screen showed an defect for ISCs to proliferate and differentiate when Cph was lost. Additionally, this gene’s expression is reduced in aged dISCs, suggesting a potential role in gut degeneration during aging. The role of Cph has only been defined in neural stem cells and muscle cell progenitors, showing the critical function of Cph for proliferation and differentiation in these cell types. Therefore, I hypothesize Cph is required for the homeostasis and proper regulation of dISCs between active and quiescent states, and is indispensable for gut maintenance during aging. For aim 1, I will characterize the cellular and molecular mechanism this occurs by generating a new reporter line and driving cell-specific KD of Cph, characterizing potential regulators of Cph, and then apply a novel single cell multiome sequencing approach to identify genes that Cph regulates and how dysfunction impacts the microenvironment. In aim 2, I will study the age-associated decrease of Cph and explore whether and how its decline contributes to gut degeneration. This work will have significant implications for how aging impacts ISC function and how stem cells regulate the transition between active and quiescent states.

NIH Research Projects · FY 2025 · 2025-06

Project Summary / Abstract The brain continuously regulates its internal states in order to efficiently and flexibly interact with the environment. Brain states vary at a global scale due to changes in arousal, whereas local changes are typically observed during selective attention. Although arousal and attention operate at different spatial scales, they have similar physiological signatures. However, the major limitation of our understanding of the impact of brain states on neuronal circuits is the fact that arousal and attention have mostly been studied separately and in different species. Indeed, despite clear similarities between neural correlates of arousal in the mouse and attention in the primate, research on brain states in the two species has proceeded largely in parallel, with very few direct collaborations between primate and rodent labs. Therefore, whether and how the mechanisms regulating brain states on different spatial scales intersect to jointly influence neuronal function and behavior is unknown. Furthermore, with few exceptions, attention and arousal have been typically examined in one brain area at a time, hence precluding our understanding of the coordination of brain states at global and local spatial scales, their neural mechanisms, and their influence on behavioral performance. Here, we propose to develop several technical capabilities that will allow us to study multiple features of brain state changes across quiescence, arousal, and attention in mice and monkeys. We will develop a workflow for performing large-scale electrophysiological and optical recordings from multiple cortical areas relevant for visual behavior, establish a complementary behavioral paradigm in both species, and validate an optogenetic approach to silencing feedback. These capabilities will allow us to test three central hypotheses in a subsequent R01: (1) brain state changes in the mouse are a subset of changes in the primate, with similar underlying mechanisms but different spatial scales (Aim 1 ); (2) selective attention is built on top of mechanisms that mediate fast changes in global arousal (Aim 2), and (3) feedback from higher areas onto specific neural cell types mediates selective attention and is gated by global neuromodulation (Aim 3). These experiments a will generate a potentially transformative multi-modal dataset for understanding the mechanisms of attention and arousal at single cell resolution, which will also advance our understanding of dysfunctional brain states and their impact on neurological disorders.

NIH Research Projects · FY 2026 · 2025-06

PROJECT SUMMARY/ABSTRACT Neural crest cells (NCCs) comprise a multipotent population of cells that migrate throughout the vertebrate embryo and differentiate into a wide variety of cell types and tissues such as the enteric, autonomic, and peripheral nervous system; tissues of the adrenal gland, thyroid gland, heart, and eye; melanocytes of the skin; and craniofacial bone and cartilage. Therefore, disruptions in NCC specification, migration, and/or differentiation can affect many aspects of embryonic development leading to an array of debilitating congenital malformations collectively called neurocristopathies. Thus, our long-term goal is to define the cellular and molecular mechanisms governing NCC development leading to new therapeutic targets. In this proposal we aim to define the syndrome combined methylmalonic acidemia and homocystinuria, cblC-type (cblC) – an inborn error of cobalamin (vitamin B12) metabolism (IECM) – as also a neurocristopathy impacting mammalian development. CblC is caused by mutation of the gene MMACHC, which encodes an enzyme essential for intracellular trafficking and metabolism of cobalamin into its active coenzyme states. Without active cobalamin, dependent metabolic pathways cannot function properly and consequently, severe neurodevelopmental defects, anemia, retinopathy, and other congenital disorders result. However, the specific contributions of each metabolic perturbation to the tissue-specific pathophysiology of cblC is largely unknown and there are no effective treatments for affected infants and children. In our preliminary studies, we discovered that Mmachc may play a role in neural crest development and thereby redefine cblC as a neurocristopathy. Therefore, this proposal aims to use novel mouse models of IECM to identify which specific cobalamin-dependent pathway, when disrupted, leads to neural crest developmental defects. We will also investigate whether modulation of the maternal environment can improve the phenotypic presentation of cblC defects in offspring. In total, these finding will shed much needed light on the pathophysiology of cblC and likely inform patient treatment.

NIH Research Projects · FY 2025 · 2025-06

Project Summary The Intellectual and Developmental Disabilities Research Center at Baylor College of Medicine (BCM IDDRC) has been instrumental in advancing basic science, translational, and clinical endeavors to improve the lives of individuals with intellectual and developmental disabilities (IDD). Beyond discoveries, our Center has mentored several generations of scientists and physicians engaged in research and treatment of individuals with IDD. BCM IDDRC’s mission is to identify as many causes of IDD as possible, understand their pathogenesis, and develop approaches to diagnosis and therapy. In this request, we propose to continue our mission for the next year, accelerating the research of our Investigators, and advancing development of therapies for IDD. We will continue to focus on these aims: 1) To continue providing Core facilities and services to advance translational IDD research. Six cores of our center will continue providing our current services while also adding new services. These cores provide innovative, high-quality, and cost-effective research services for BCM IDDRC investigators to study molecules (Molecular and Expression Analysis Core), cells and tissues (Cell and Tissue Pathogenesis Core), circuits (Circuit Analysis and Modulation Core), and whole organisms (Preclinical and Clinical Outcomes Core). The Clinical Translational Core will continue providing services specific for clinical translation. The Administration Core (AC) will continue to coordinate overall Center operations along with stakeholder engagement, communication and education; 2) To continue promoting and enhancing collaborative efforts and dissemination activities with a comprehensive engagement, communication, and education plan. The AC will continue to promote local, national, and international interactions and implement best practices for community partnerships, dissemination of research findings, and enhance the training of next-generation IDD researchers; 3) To continue our multidisciplinary signature research project and expand on our previous clinical trial readiness achievements. The emergence of genomic therapies, coupled with exciting discoveries and preclinical studies from the BCM IDDRC, have provided us with great opportunities to develop therapies for IDDs. Because many IDD-causing genes are dosage sensitive there is a serious challenge which requires development of robust biological markers that are meaningful for individuals rather than the population. Building upon our success over the past 5 years, the Signature Project will continue to develop multidimensional biomarkers for target engagement, safety, and efficacy in dosage-dependent IDDs. For the next year, our center will continue to support ~ 88 investigators and over 50 research projects. As we have done for more than 30 years, the BCM IDDRC will continue to foster an environment that welcomes and supports additional investigators and emphasizes training. In the coming years, Center investigators, their collaborators, and trainees will be poised to transform exciting discoveries into safe therapeutics that will improve the quality of life and well-being of individuals with IDD.

NIH Research Projects · FY 2026 · 2025-06

Project Summary Hepatocellular carcinoma (HCC) is the third-leading cause of cancer-related death worldwide, with the fastest- growing rate. Nonalcoholic fatty liver disease (NAFLD) affects approximately 25% of the world’s population and is becoming a leading cause of HCC. The progression of NAFLD to HCC, including nonalcoholic steatohepatitis (NASH), liver fibrosis, and cirrhosis, is well-recognized, but the pathogenesis of NAFLD- associated HCC is not well understood. Circadian rhythms allow mammals to anticipate daily environmental changes and maintain physiological homeostasis. Epidemiologic studies indicate that circadian misalignment contributes to the development of many cancers. Moreover, the concept of chronotherapy is attracting more attention to improving drug efficacy when drugs are provided at the optimized time of the day. We and others unexpectedly uncovered that although core clock genes still retain robust rhythmic expression, the rhythms of thousands of genes are altered in NAFLD, suggesting critical roles of noncanonical clock regulators. In preliminary studies, we performed the first Chromatin Interaction Analysis with Paired-End-Tag sequencing (ChIA-PET) experiments in healthy and NAFLD livers and identified multiple noncanonical clock transcription factors (TFs), including Estrogen Related Receptor Gamma (ESRRγ), which are enriched in the enhancer- promoter circadian looping anchors. Indeed, the rhythmic expression of ESRRγ is enhanced in NAFLD, and ESRRγ is associated with nearly half of these NAFLD-induced E-P circadian loops. Moreover, hepatocyte- specific overexpression of ESRRγ slows, but hepatocyte-specific knockout of ESRRγ accelerates NAFLD- related HCC progression. This proposal will test the hypothesis that ESRRγ, as a noncanonical clock gene, is a key regulator mediating circadian E-P interactions in NAFLD and is critical for the progression of NAFLD to HCC. In functional Aim 1, We will use adeno-associated virus expression systems to introduce ESRRγ expression in constitutive, in-phase, and anti-phase rhythmic patterns in Esrrγ-knockout livers from multiple preclinical NAFLD–HCC mouse models to define how ESRRγ rhythmicity affects the progression of NAFLD and HCC. In mechanistic Aim 2, we will determine how noncanonical clock transcription factors regulate circadian enhancer-promoter looping and gene expression. In translational Aim 3, we will determine whether ESRRγ agonists affect NAFLD-to-HCC progression in a time-dependent manner by administering them at the peak and trough of ESRRγ expression in newly generated ESRRγ reporter mouse models and liver slice culture tissues from people with NAFLD. Completion of this work will provide novel insights into how a noncanonical clock TF, ESRRγ, remodels the 3D circadian chromatin architecture and gene expression during the NAFLD-to-HCC progression, laying the intellectual groundwork for future chronotherapy strategies that maximize efficacies by considering the timing of drug administration.

NIH Research Projects · FY 2025 · 2025-05

Summary This NIH grant proposal seeks funding to acquire a cutting-edge Femtonics Atlas microscope at Baylor College of Medicine (BCM), with a specific focus on its applications for neuroscience research. The Atlas will enable rapid, high-resolution imaging of activity-dependent fluorescence in vitro and in vivo using calcium, voltage, or neurotransmitter reporters. The Atlas combines two-photon excitation with acousto- optical deflectors (AOD) scanning, allowing fast recordings of fluorescence from arbitrary locations or shapes in a 3D volume without moving parts. Over the past two decades, traditional two-photon laser-scanning microscopy (2PLSM) has significantly advanced our understanding of neuronal function and structure due to its ability to image activity deep in scattering tissue. However, standard 2PLSM is limited by single-plane scanning, which is often not compatible with the geometry of target structures such as dendrites or astrocytes. When combined with methods for fast Z focusing, 2PLSM can scan multiple planes, but at the expense of reduced temporal resolution. The Femtonics Atlas overcomes these limitations by rapidly pointing the laser beam to any point within a 3D volume, enabling high-speed, random-access imaging that is crucial for recording fast neuronal activity with the latest GCaMP8 calcium sensors and with genetically-encoded voltage indicators (GEVIs) like JEDI-2P (developed at the St-Pierre lab here at BCM). The proposal highlights the unique advantages of the Atlas to support a diverse set of NIH-funded projects within the neuroscience department at BCM, with potential applications in other departments as well. These projects will benefit enormously from the ability to track activity in complex 3D structures of neurons and glia, to image sparsely-labeled cells with high temporal resolution, and to deliver precise two-photon optogenetic stimulation in parallel with imaging. With a group of nearly a dozen enthusiastic neuroscientists looking forward to applying the Femtonics Atlas’s capabilities to their research, bringing this microscope to Houston has the potential to significantly advance our understanding of brain function and disease.

NIH Research Projects · FY 2026 · 2025-05

Profound accumulation of macrophages (MΦs) and persistent inflammation are prominent features in as- cending thoracic aortic aneurysms and dissections (ATAAD), and the underlying mechanisms are poorly under- stood. Recent studies suggest that innate immune cells, including monocytes/MΦs, can differentiate into a pro- inflammatory phenotype and conserve this memory after exposure to stimuli, a process known as trained im- munity, thereby enhancing the subsequent inflammatory response. In our preliminary studies, scRNA-seq analysis revealed several MΦ subpopulations. Importantly, aortic MΦs in sporadic ATAAD patients and in an- giotensin (Ang II)-induced ATAAD mice exhibited pro-inflammatory status with enhanced differentiation to pro- inflammatory MΦs and suppressed differentiation to phagocytic/anti-inflammatory MΦs. Intriguingly, this pro- inflammatory status can be observed in monocytes/MΦs isolated from bone marrow of mice, and the memory can stably last for 5 days in ex vivo culture. Furthermore, scATAC-seq suggested that the training of the mono- cytes/MΦs occurred at the epigenetic level, and several TFs, including IRF3 and surprisingly IRF4 (known as an anti-inflammatory lineage-determining TF), may drive the induction of pro-inflammatory differentiation. In cul- tured cells, DNA from damaged aortic cells induced monocytes/MΦs toward a pro-inflammatory phenotype through cGAMP-STING-IRF3/IRF4 signaling. Finally, Sting-deficient mice showed preservation of phago- cytic/anti-inflammatory MΦs and reduction of pro-inflammatory MΦs. The objective of this project is to under- stand the training of monocytes/MΦs to different functional phenotypes in the aortic wall, and to identify key pathways driving monocytes/MΦs to a pro-inflammatory direction in ATAAD. In Aim 1, we will determine the sites of the immunity training, the epigenetic signatures of the trained monocytes/MΦs, and the duration of the memory in ATAAD by performing integrated omics analyses (scRNA-seq, scATAC-seq, and spatial RNA-seq) of monocytes/MΦs in bone marrow and the aortic wall in mice and will confirm the epigenetic signatures of the trained monocytes/MΦs in circulation and the aortic wall in ATAAD patients. In Aim 2, we will determine the causal role of the STING-IRF3/IRF4 circuitry in training monocytes/MΦs during ATAAD development by per- forming omics and functional analyses of aortic tissues and bone marrow monocytes from MΦ-Sting-/-, MΦ-Irf3- /-, MΦ-Irf4-/-, and WT mice, and dissect the link between dsDNA/cGAMP-STING-IRF3/IRF4, chromatin remodel- ing, gene expression, and MΦ phenotypes in cultured monocytes/MΦs. In Aim 3, we will determine the role of MΦ-specific STING-IRF3/IRF4 in ATAAD development in mice. The impact of this work will be an improved understanding of the molecular mechanisms that cause persistent aortic inflammation and degeneration, provid- ing a new direction for treatment strategies for preventing ATAAD and its fatal sequelae. Elucidating the immunity training of monocytes/MΦs is likely to have broad implications for many other cardiovascular diseases in which persistent inflammation plays a critical role in tissue destruction and disease progression.

NIH Research Projects · FY 2026 · 2025-05

Project Summary Inflammation, the initial response to injury, plays a pivotal role in wound-healing. In addition to a defense mechanism, inflammation is intertwined with tissue repair, orchestrating a delicate balance between necessary defensive actions and the promotion of tissue regeneration [ref]. However, dysregulation of this inflammatory phase can lead to impaired healing, a critical focus for effective wound management strategies. Therefore, understanding and modulating this inflammatory response is crucial to enhancing wound healing, particularly in chronic wounds where chronic inflammation impairs the healing process. Chronic wound is a debilitating condition affecting the quality of life of approximately 8.5 million people, with an annual expenditure of more than $50 billion in the US. Unfortunately, the number of patients and costs will increase further with risk factors such as diabetes, vascular disease, and aging. Although several strategies to manage chronic wounds are available, the need for effective chronic wound treatment persists. The overarching goal of my research is to develop and optimize bioengineered polymers with tunable properties to modulate inflammatory responses in the tissue microenvironment, thereby enhancing tissue repair and regeneration across a spectrum of wound types. The rationale of this research is to systematically investigate how biopolymers' chemical and mechanical properties influence inflammatory response and tissue healing. By understanding these relationships, the project aims to engineer biopolymers with optimized surface charges, elasticity, and porosity, that can either mitigate excessive inflammation in chronic wounds or enhance necessary inflammatory responses in normal wound healing. This project will integrate immunology with polymer surface chemistry and employ 3D fabrication strategies to modify the surface charges of biopolymers and optimize elasticity and porosity, respectively. Hence, we aim to develop biopolymers contributing to controlled and effective healing processes. Over the next five years, we will delve into investigating how the intrinsic properties of biopolymers can be engineered to actively influence, control, and normalize the wound microenvironment by material characterizations, in vitro and in vivo studies using well-established mouse models to recapitulate skin excision wounds and diabetic wounds. In the first project, we aim to uncover the mechanisms by which the surface charges of biopolymers affect local inflammation. In the second project, we aim to investigate the tailoring of physical-mechanical properties to enhance the therapeutic efficacy of biopolymers in wound healing applications. If successful, such understanding is vital for advancing the design and application of biopolymers in a way that is not only biocompatible but also contributes positively to the local tissue environment and overall patient health.

NIH Research Projects · FY 2026 · 2025-05

Platelet (PLT) transfusions are necessary to prevent and treat bleeding. However, children and neonates have worse outcomes with platelet transfusions. Despite being lifesaving, PLT transfusions may lead to transfusion-related complications, such as transfusion related acute lung injury, transfusion associated circulatory overload, allergic reactions, and febrile non- hemolytic transfusion reactions. These complications are largely due to the contaminants that accumulate in the PLT suspending medium. Common methods of PLT processing include centrifugation and leukoreduction filters. However, each of these modalities have significant drawbacks, including premature PLT activation, creation of inflammatory microparticles, and inefficiency to remove all contaminants (whether large or small). Our objective is to develop and validate a simple-to-use, disposable device capable of removing all contaminants while preserving the highest-functioning PLTs. Our novel microfluidic technology, controlled incremental filtration (CIF), separates blood cells by size using devices with compact footprints and minimal void volumes, while minimizing cell activation and ensuring practical flow rates. Our latest CIF device prototypes remove ≥60% of supernatant and recover 98% of large, highly functional PLT (~90% overall PLT recovery), while minimally increasing PLT activation. The purpose of this study is to test our hypothesis that purifying PLTs with high-throughput CIF- based microfluidics is feasible, effective, and safe. We will test this hypothesis through three Specific Aims. In Aim 1, we will optimize the efficiency of the CIF devices and scale them up to be able to run at clinically relevant flow rates using human PLT units obtained from a regional blood blank. In Aim 2 we will evaluate the effect of CIF purification on the hemostatic function of PLTs in vitro (using a large battery of tests, including flow cytometry, light transmission aggregometry, and flow adhesion assays (type I collagen, VWF, and thrombin) and in vivo by transfusing purified human PLT into thrombocytopenic NOD/SCID mice and evaluating thrombus formation time using photochemical and ferric chloride injury models. In Aim 3, we will evaluate the safety and efficacy of PLTs purified by CIF in a premature neonatal pig model, including piglets with necrotizing enterocolitis (NEC). We hypothesize that transfusion of CIF- processed pig PLT into premature neonatal pigs will be safer (less inflammation) and more efficacious (less bleeding) than conventional PLT processing methods in piglets with and without NEC. This project establishes the essential foundation for finalizing the device design and conducting pivotal human trials, paving the way for safer PLT transfusions in all patients.

NIH Research Projects · FY 2026 · 2025-05

Project Summary/Abstract CRISPR/Cas9 genome editing is rapidly developing into therapeutic applications. Therefore, it has become increasingly important that tools for pre-clinical testing of novel reagents provide the maximum benefit for researchers. One critical aspect of the use of CRISPR/Cas9 is the delivery of the molecular components of the editing machinery to the target tissue. Delivery reagents, such as viral vectors, lipid nanoparticles, conjugated ribonucleoproteins, and others, need to be tested for high efficiency editing of target tissues. At the same time, the reagents must also be evaluated for editing in non-targeted tissues and organs, to avoid unexpected biological consequences. Therefore, animal reporter models that detect genome editing events, are valuable tools for researchers developing new genome editing strategies. Current animal reporters, primarily mice, rely on the expression of a fluorescent or bioluminescent molecule in response to a genome editing event. One widely used model is the “Ai9” mouse. In this transgenic strain, a “double-hit” strategy is employed, where simultaneous CRISPR/Cas9 nuclease DNA cutting events, followed by Non-Homologous End Joining (NHEJ), excises an upstream stop cassette, allowing expression of a tdTomato fluorescent protein. This model is particularly useful, since the tdTomato protein produces robust fluorescence, allowing detection of very rare events. However, the model also has a pitfall. Many editing events, especially NHEJ that generates indels instead of excision, do not result in expression of tdTomato. Therefore, the actual number of editing events is undercounted. Therefore, we propose an alternative “single-hit” strategy to detect CRISPR/Cas9 editing in the Ai9 mouse. The Cre recombinase protein will also excise the stop cassette in the Ai9 mouse. However, when the Cre protein is fused to a derivative of the human estrogen receptor ligand binding domain called ERT2, it is retained in the cytoplasm, and remains inactive. We will design and test guide RNAs for CRISPR/Cas9 editing that will target this ERT2 domain. The resulting editing of the ERT2 is anticipated to disable the retention of the Cre recombinase in the cytoplasm, allowing the protein to translocate to the nuclease and excise the stop cassette. We predict that this strategy will be significantly more sensitive for the detection of editing events. In addition, this method will be adaptable to other Cre-inducible systems with alternative fluorophores and bioluminescent proteins as reporters. Finally, we expect that this strategy can be used with other CRISPR/Cas9-derived activities, notably base editing.

NIH Research Projects · FY 2026 · 2025-04

PROJECT SUMMARY Genetic alterations triggering oncogene activation can produce neoantigens, rendering transformed cells vulnerable to immune assault. Under heightened immune pressure these cells must balance growth with immune evasion, offering a potential therapeutic avenue for early malignant cell targeting. However, the identification and study of distinct alterations contributing to immune evasion independently of tumor growth is hindered by the absence of immunocompetent models capable of separating oncogene-driven tumorigenesis from the associated immune responses. To overcome this barrier, we developed RECON, a novel immune tolerization strategy to convert transgenic proteins that may be recognized as “non-self” into self-antigens. RECON enables the use of foreign elements in the context of an intact immune system via ubiquitous expression of these elements, at the same time utilizing a dual-transcriptional/translational control system to suppress endogenous transgene levels and activity (e.g. luminescence, oncogenic transformation). Utilizing RECON's dual transgene tolerizing-suppressing effects on immunogenic oncogenes, we hypothesize that we can uncouple an oncogenes pathogenic potential from its immunogenic properties, enabling for dissection of the distinct genetic and phenotypic alterations that govern immune evasion independently of tumorigenesis during the earliest stages of cancer pathogenesis. Understanding these molecular events will shed light on how cells expressing non-native elements achieve enhanced immune evasion, thereby advancing our comprehension of cancer progression and potentially exposing new therapeutic strategies for early disease targeting. RECON presents a unique opportunity to explore oncogene-driven tumorigenesis within experimentally modified immune environments. It stands to generate fundamental insight into the trade-offs between tumorigenicity and immunogenicity during tumor progression and evolution. Overall, the RECON model represents a promising advancement in scientific research, offering a versatile solution that addresses existing limitations while extending the utility of diverse transgenic elements to various model organisms with intact immune repertoires.

NIH Research Projects · FY 2026 · 2025-04

Endometriosis, affecting approximately 10% of women of reproductive age, poses significant challenges due to its association with increased infertility and pregnancy complications. Dysregulated iron homeostasis and oxidative stress have been identified as significant contributors to these adverse reproductive outcomes. Iron, essential for critical cellular functions such as energy production and DNA synthesis, can lead to oxidative stress when present in excess, producing reactive oxygen species (ROS) that damage cellular structures. SLC40A1 (ferroportin), a key protein responsible for exporting iron from cells, plays a vital role in maintaining systemic and cellular iron levels. In endometriosis, abnormalities in SLC40A1 expression or function may disrupt iron balance within the endometrial tissue, exacerbating oxidative stress through reactive oxygen species (ROS) production via Fenton reaction. This oxidative stress can impair endometrial stromal cell decidualization, a process crucial for embryo implantation, thereby contributing to compromised early pregnancy outcomes. Preliminary findings indicate reduced SLC40A1 expression in endometriosis patients, suggesting a link to elevated local iron levels and increased oxidative stress within the endometrium, which may disrupt normal decidualization processes essential for successful pregnancy. To further investigate these mechanisms, menstrual effluent derived stromal cells and organoid models will be employed to characterize how SLC40A1 influences iron dynamics, oxidative stress markers, and lipid peroxidation during decidualization. Complementing these studies, genetic manipulation of SLC40A1 in mouse models aims to provide deeper insights into its impact on reproductive outcomes, specifically its role in protecting against decidual ferroptosis—a form of cell death associated with excessive iron and lipid peroxidation. Additionally, the transcriptional regulation of SLC40A1 by FOXO1 and GATA2, critical transcription factors involved in oxidative stress response and decidualization, will be explored through genome wide (CUT&RUN) analyses and luciferase reporter assays. These experiments aim to elucidate the direct interactions between FOXO1/GATA2 and the regulatory regions of the SLC40A1 gene, revealing how these factors influence SLC40A1 expression and modulate cellular iron levels within the endometrium. Ultimately, uncovering iron dynamic and oxidative stress networks in endometriosis could uncover novel therapeutic targets for improving reproductive health outcomes. By integrating clinical samples with mechanistic insights from cellular and animal models, this research aims to develop targeted interventions addressing endometriosis-associated infertility and pregnancy complications. The F32 grant will not only provide the necessary financial support for my research but also offer valuable career development opportunities. This includes professional training, mentorship, and the chance to build a network with leading experts in the field.

NIH Research Projects · FY 2025 · 2025-04

Project Summary The goal of this proposal is to build a state-of-the-art respiratory exposure system at Baylor College of Medicine. This system will address a critical gap in inhalation exposure research using rodent models. It will facilitate studies encompassing developmental to degenerative life stages, peripheral and central neurological function, respiratory control, immune functions, and cardiovascular physiology. We will offer a multi-modal inhalation exposure platform for advanced respiratory exposure research at Baylor College of Medicine. We are requesting a customized Data Sciences International (DSI) Inhalation Solution that integrates multiple options for 1) housing the mice during exposure, 2) aerosolizing the desired agent, and 3) measuring and analyzing resultant respiratory parameters during exposure for maximal flexibility across a wide variety of users and research projects. The customized DSI solution utilizes a Stackable Inhalation Tower with options to expand capacity over time without any loss of investment from the initial purchase. The Inhalation Dosing Tower uses modular levels, each level providing 7 inhalation exposure ports, and can be expanded up to 6 levels for a total of 42 possible ports. The requested equipment provides capacity for 28 ports which would allow for the exposure of 23 animals while the remaining ports are used to sample the airflow to quantify particle/aerosol concentration. The tower enables real-time respiratory measurements that when integrated with the FinePointe software allow the user to fine-tune acute exposures based on measured respiratory parameters and particulates/aerosolized components. There are three options for aerosolizing or smoking desired respiratory exposures: 1) an E-Cig/ Vape/ Tobacco Smoke (EVT) Generator, 2) a Smoke Generator, and 3) an Aerogen Nebulizer Head capable of aerosolizing a variety of desired toxins including virus and bacteria. The system will be housed in a BSL-3 safety cabinet to ensure full containment and safety. Lastly, BCM will make a substantial financial commitment in salary support and an extensive, five-year service contract to ensure consistent, optimal operational, and maximal longevity for the requested equipment. This equipment system represents a significant opportunity to advance respiratory exposure research for Baylor College of Medicine (BCM). The portfolio of research conducted at BCM represents a unique opportunity to improve experimental approaches in disease states influenced by environmental respiratory exposures including Sudden Infant Death Syndrome (SIDS) and Sudden Unexpected Infant Death (SUID), congenital respiratory disorders, Acute Respiratory Distress Syndrome (ARDS), bronchopulmonary dysplasia (BPD), asthma, pneumonia, emphysema, and BSL-2 level pathogen infections such respiratory syncytial virus (RSV).

NIH Research Projects · FY 2026 · 2025-04

PROJECT SUMMARY Amplification of the oncogene MYCN drives high-risk progressive disease, resistance to therapy, and a poor overall survival rate below 45% for high-risk neuroblastoma (NB) patients. Further, more than half of all high-risk patients will relapse, and the post-relapse survival rate is only 10%. MYCN-driven NB tumors are characterized by low mutational load, low MHC-I expression, reduced immune cell infiltration, and impaired T cell effector functions. As a result, MYCN-amplified NB is immunologically quiescent, thus limiting immunotherapy approaches. This proposal addresses a major obstacle to developing effective therapies for MYCN-amplified NB by delineating the molecular mechanisms through which MYCN drives T cell dysfunction and thus a suppressive tumor immune microenvironment (TIME). Based on our data and the recent literature, our guiding hypothesis is that MYCN reprograms T cell metabolism to drive an immune suppressive TIME. Specifically, we hypothesize that MYCN creates a lipid-rich tumor microenvironment that benefits NB growth and impairs T cell effector functions. This proposal will (1) determine how MYCN rewires T cell metabolism to drive immune suppression; and (2) elucidate how MYCN-induced de novo lipogenesis contributes to a suppressive TIME. Deciphering how MYCN contributes towards a suppressive TIME is vital for developing new and improved immunotherapy strategies for high-risk NB.

NIH Research Projects · FY 2025 · 2025-04

PROJECT SUMMARY The importance of DNA methylation in human health and disease is clear. The fundamental role of DNA methylation in normal development and disease processes, however, remains elusive. The use of gene targeting in animal models definitively demonstrated that genetic mutations at specific genes cause disease. An analogous “epigenetic gene targeting” approach is urgently needed to advance the field of epigenetics and human disease. In this regard, we developed the mouse model with an inducible CRISPR/Cas9 system for targeted manipulation of DNA methylation in vivo. Our preliminary data demonstrated that this system provides unparalleled simplicity in designing DNA methylation modifications or conducing epigenome engineering. Based on these findings, this proposal responds to PAR-21-167: Development of Animal Models and Related Biological Materials for Research. Our overall goals are to develop simple, robust, and reliable tools for targeted editing of locus- specific DNA methylation; these will have broad utility in epigenetic research. Specifically, we will: 1− Utilizing inducible dCas9-SunTagTET1 mice for precise targeted DNA demethylation. 2− Developing inducible dCas9- SunTagDNMT3A-3L mice for targeted DNA methylation. The successful completion of the proposed studies will open entirely new avenues in the field of epigenetics: investigators will be able to 1) test primary functional roles of DNA methylation in vivo, 2) understand how DNA methylation modifications work together to cause biological phenotypes, and 3) design and test targeted therapeutic approaches for epigenetic disorders. The epigenetic gene targeting approaches we propose to develop will be widely utilized and accessible to a range of investigators interested in functional epigenomics and human diseases.

NIH Research Projects · FY 2025 · 2025-04

Offspring born to mothers with nutritional stress during pregnancy have increased susceptibility to metabolic diseases including diabetes and hypertension during their adult life. Low protein (LP) diet during pregnancy in rats has been shown to cause lean Type 2 Diabetes (T2D) in adult offspring. In addition, testosterone in males and estradiol in females were also reduced in these animals. The pathophysiology of lean T2D and its associated reductions in sex steroid hormone synthesis are poorly understood, and the mechanistic insights are critical for the development of therapeutics. Therefore we will use this LP diet programmed lean T2D rat model we recently developed, to investigate the mechanisms for the reduced sex steroid hormone synthesis in males. Changes in metabolic functions of skeletal muscle and liver, and in sex steroid synthesis in gonads in this model appears to be related to mitochondrial dysfunctions. In recent studies, we found ultrastructural deformities with loss of cristae and appearance of giant mitochondria as well as changes in key genes involved in various vital mitochondrial structure and function in metabolic these tissues. Impaired quality control often results in suboptimal mitochondrial function, which affects steroidogenesis. Steroidogenesis is also regulated by characteristics of the mitochondrial interaction with the endoplasmic reticulum (ER). Mitochondrial quality and ER tethering are interdependent because the ER assists the mitochondria in fission and mitophagy. The role of the intracellular cholesterol pool and its dynamics in sex steroid synthesis are also not well understood. Pilot data from our lean T2D model suggests that mitochondrial-ER membrane contacts are reduced, and intracellular cholesterol trafficking is altered in the gonads of LP-programmed T2D animals. We hypothesize that LP diet during pregnancy causes impairment of mitochondrial quality, ER-mitochondrial association, and cholesterol homeostasis in Leydig cells of male offspring leading to lower testosterone synthesis Two specific aims are proposed. Aim 1: Determine if LP diet during pregnancy causes reductions in mitochondrial-ER membrane contacts and mitochondrial quality in Leydig cells of male offspring. Aim 2: Assess if LP diet during pregnancy impairs mitochondrial metabolism and intracellular cholesterol regulation in Leydig cells of male offspring. The studies proposed are significant because we will be the first to investigate mechanisms into the mitochondrial dysfunction associated with reduced sex steroid production in LP programmed offspring.