Methodist Hospital Research Institute

NIH Research Projects · FY 2025 · 2024-08

Contact PD/PI: MYLONAKIS, ELEFTHERIOS PROJECT SUMMARY Despite the availability of anti-retroviral therapy, oral candidiasis caused by the overgrowth of Candida spp. is the most frequent opportunistic infection among individuals living with HIV. The widespread use of azoles to treat oral candidiasis in individuals living with HIV often results in treatment failure. In addition to antifungal resistance, the current therapeutic options for oral candidiasis are limited by side effects the challenges posed by biofilm drug recalcitrance. Moreover, Candida auris has recently emerged as a multi-drug–resistant fungal pathogen and is classified as 1 of the 5 pathogens in the highest category—Urgent Threats—in the Antibiotic Resistance Threats in the United States. Polyphenols are secondary plant metabolites that have protective properties for human health, including antioxidant, anti-inflammatory, anticancer, and antimicrobial activity. We found that caffeic acid phenethyl ester (CAPE) and ellagic acid (EA) are both active against Candida spp., including C. auris resistant strains. The minimum inhibitory concentration for EA ranged from 0.125 to 0.25 µg/mL and, at subinhibitory concentrations, EA inhibited phospholipase production by C. auris. CAPE inhibited fungal filamentation and biofilm formation. Both compounds were active in the invertebrate model hosts Caenorhabditis elegans and Galleria mellonella. We further demonstrated that neither CAPE nor EA is toxic to human erythrocytes. Importantly, in a mouse model of oral candidiasis, CAPE significantly increased the expression of the murine antifungal defensin β- defensin 3 and reduced pseudomembranous lesions, invasion of hyphae on epithelium surfaces, tissue damage, and inflammatory infiltrates. The purpose of this proposal is to advance the development of these natural phenolic compounds for the management of oral candidiasis. We have published “proof of concept” studies that gellan can be impregnated with CAPE, and that the hydrogel is active in a mouse model of oral candidiasis. We will further advance these goals with the following Aims: 1. To optimize hydrogels loaded with EA and/or CAPE for maximal release of compounds and test their antifungal activity and cytotoxicity in vitro. 2. To test the optimized hydrogels loaded with EA and/or CAPE in an established mouse model of oral candidiasis. 3. To test the antifungal immune response to hydrogels loaded with EA and/or CAPE in the oral mucosa of volunteers living with HIV. Our overall hypothesis is that the combination of CAPE and EA could provide an ideal natural treatment against oral candidiasis with direct antifungal activity and activity against resistant strains (including C. auris), efficacy against filaments and biofilms, and immunomodulatory activity through the production of antimicrobial peptides and chemokines. In this context, the proposed studies could represent a pivotal stage in the development and translation of these natural compounds as clinical antifungal agents. References Cited Page 1

NIH Research Projects · FY 2025 · 2024-08

ABSTRACT Tuberculosis continues to kill about 1 million people including children each year with 8 million new cases. Regrettably, coinfection with HIV-1 has aggravated the problem because HIV-1 depletes CD4 T cells, which are the main defense mechanism against tuberculosis. For the reason, BCG, which is a live attenuated vaccine against tuberculosis, cannot be given to people living with HIV. We recently discovered that Sirtuin type of protein and histone deacetylates play a major role in regulating the growth of both M. tuberculosis and HIV-1 in human macrophages and drug targeting Sirtuins could control both pathogens in cell culture models and in humanized mice. Therefore, we propose to investigate Sirtuin-dependent intervention to understand how TB and HIV-1 infections induce these enzymes and how we can develop novel immunochemotherapy for confections. Specifically: Aim-1: We will analyze the impact of early Mtb and HIV infection on Sirtuin gene and protein induction in macrophages and identify their epigenetic targets. We will examine the hypothesis that Mtb-induced Sirtuin-2 can augment HIV-1 replication, whereas HIV-1 induced Sirtuin-2, in turn enhances Mtb growth in MФs thereby aggravating coinfections. We will then develop an immunochemotherapy (ICT) using a combination of Sirtuin drugs, TB and HIV targeting drugs to kill and eradicate both HIV-1 and M. tuberculosis in macrophages. Aim-2: We will investigate the effect of Sirtuin drugs on early and late stages of TB-HIV-1 coinfection using humanized mice and determine whether ICT can eradicate confections. Aim-3: Tuberculosis granulomas can restrict growth of M. tuberculosis pathogen, although prior infection with HIV-1 can deplete CD4 T cells and affect granuloma formation. We will use a TB-HIV coinfection model of humanized mice to determine if Sirtuin-drug therapy can activate macrophages of granulomas and increase host-defense against tuberculosis. Our overall goal is to develop innovative immune defense-based intervention methods to eradicate TB-HIV coinfections.

NIH Research Projects · FY 2025 · 2024-07

Pancreatic cancer (PanCan) is notorious for its resistance to therapy including immunotherapy, which is largely contributed by a suppressive tumor microenvironment. Targeting molecular pathways integral to PanCan resistance to current therapy remains an unmet need. Notch signaling regulates PanCan growth, metastasis, and the tumor microenvironment. While other studies have shown that Notch signaling in macrophage and myeloid derived suppressor cells plays an immune-suppressive role in breast cancer and lung cancer, for example, our proposal will study previously unrecognized role of JAG1 in the regulation of the metabolic fitness of conventional dendritic cells (cDCs) that underlies the poor anti-tumor responses to immunotherapy. The overarching objective of this application is to define if targeting Jagged1 (JAG1), a Notch ligand, whose expression correlates with a worse prognosis and inversely correlates with BATF3, essential for type I cDC (cDC1) development, could reverse the immunotherapy resistance in tumors with poor tumor-infiltrating cDC1 and T cells. cDC1 cells present tumor antigens to tumor-killing T cells and thus play a critical role in supporting T cell anti- tumor immunity in immunotherapy. However, dendritic cells are present in low numbers and often display functional suppression in pancreatic cancer patients. Our recent work revealed that JAG1 functions as an inhibitory Notch ligand for cDC1 development as well as cDC metabolic fitness. We also found that ablating JAG1 induced marked tumor regression and prolonged tumor-bearing mice long-term survival by increasing and reviving tumor-infiltrating DCs and Notch-activated tumor-killing T cells. We will test our central hypothesis that JAG1 promotes PanCan therapy resistance by suppressing cDC1 metabolic fitness and CD8 T cell anti-tumor response through three interrelated aims. In aim 1, we will test if anti-JAG1 blocking antibody could sensitize resistant tumors to immunotherapy. In aim 2, we will investigate the mechanism by which JAG1 impairs cDC metabolic fitness essential for anti-tumor activation. Aim 3 will focus on the mechanism of anti-tumor T cell reinvigoration mediated by JAG1 ablation/blockade. We will use a combination of the autochthonous KPC mouse model, immunophenotypically defined KPC orthotopic tumors, human PanCan organoids/myeloid cell coculture, and human PanCan specimens. We will employ cutting-edge mass cytometry to investigate the interplay between tumor cells and the surrounding immune cells and the tumor stroma. We will integrate the immunophenotyping and the spatial transcriptomics of rare immune cell populations with clinical and histopathological information to correlate molecular findings with patient outcomes. This work is innovative as we illuminate a novel function of JAG1 in the maintenance of PanCan therapy resistance and explore its potential as a novel immunomodulatory treatment for this deadly disease.

NIH Research Projects · FY 2026 · 2024-07

ABSTRACT Peripheral artery disease (PAD) causes severe morbidity as plaques obstruct blood flow, preventing adequate perfusion to limbs, which may result in amputation or death. There are one million interventions to treat PAD per year, emphasizing the high prevalence of this disease. The preferred treatment is percutaneous vascular interventions (PVI), where vessel plaque is penetrated and threaded (crossed) by a guidewire, followed by a balloon over the wire and/or other adjunctive devices to open the blood vessel. Unfortunately, PVI fails immediately for 20% of patients because the plaque proves impenetrable -- meaning patients are put at risk for no health benefit. Furthermore, after ballooning open there is damage to the vessel wall and vessels block again within a year for 70% of patients with below-the-knee plaques. These failures then require additional invasive interventions. The difficulties with PVI are frustrating for the clinicians performing the surgery and needlessly risky for our patients. There is a fundamental gap in knowledge in how to select patients that benefit from PVI and how different preparation devices alter vessel wall injury. We propose a two-pronged approach: 1) improve patient selection for PVI using a novel MRI-histology based anatomic scoring system that identifies patients with impenetrable plaques; and 2) improve device selection by identifying devices that reduce vessel wall injury during PVI using histopathologic analysis after PVI in a cadaveric model. Our preliminary data show that our in vivo MRI-histology method can visualize hard plaque components (both calcium and dense collagen) to decipher individual patients' plaque morphology and determine which plaques are penetrable. Furthermore, our cadaveric model indicates differences in vessel wall injury following different preparation devices using detailed ex vivo plaque analysis post-intervention. Our Specific Aims are: 1. Establish a novel MRI-histology anatomic scoring system that predicts PVI immediate technical failure to improve patient selection for PVI. We will image patients prior to their PVI with our clinical MRI-histology protocol to prospectively score individual patients’ plaques to predict successful guidewire crossing. 2. Describe the benefits of using orbital atherectomy before balloon angioplasty versus using balloon angioplasty alone for various plaque types in arteries below the knee. We will randomize amputated legs from PAD patients with calcified below-the-knee arteries to undergo plain balloon angioplasty versus atherectomy preparation device prior to angioplasty. Pre-procedure 3T MRI-histology, intravascular ultrasound, and ex vivo plaque analysis with histology, will determine vessel wall injury between intervention groups. We will establish a patient selection process for PVI and define vessel wall response with ballooning only versus vessel preparation with atherectomy through histologic analysis of cadaveric lesions. Our study will empower clinical decision-making toward the safest, most effective strategies to ultimately save patient limbs and lives.

NIH Research Projects · FY 2024 · 2024-07

Project Summary T cell-based immune therapies are a promising approach to treat cancer. Chimeric antigen receptor (CAR) is one such therapy developed by engineering a protein comprised of a tumor recognition motif, a T cell major signaling molecule, and costimulatory molecules. Engineered T cells expressing CAR (CAR-T cells) have shown remarkable clinical success in treating B cell malignancies, but their efficacy in controlling solid tumors is limited. The major hurdle to T cell treatments is overcoming their chronic stimulation and CAR-T cell exhaustion. Chronic T cell activation induces oxidative stress and accumulation of stress granules (SG). SG are subcellular cytoplasmic compartments that contain RNA-binding proteins and translationally repressed mRNAs. Significant progress has been made in understanding the components of the SG core and uncovering the therapeutic potential of SG regulators in cancer and neurodegenerative diseases. However, the functional role of the T cell SG network in T cell exhaustion and the mechanistic link between the SG core and CAR-T cell exhaustion are completely unknown. We propose to test the hypothesis that augmentation of the SG regulatory network leads to CAR-T cell exhaustion. Using our recently devised CAR construct with a fluorescence reporter to track SG assembly and maintenance in living cells, we will analyze the SG dynamics of activated and exhausted CAR-T cells when the SG core regulator, G3BP1, is downregulated. We will measure how effective the G3BP1-downregulated CAR-T cells are in controlling tumor cell growth in vitro and in vivo to determine if the G3BP1 downregulation improves CAR-T function and longevity. Next, we will target upstream metabolic stress regulators to test whether CAR-T cell function improves when metabolic stress regulators are deficient. Our innovative proposal to explore the T cell SG network, the translational regulators of metabolic stress as regulatory targets to improve CAR-T cell longevity, and the mechanistic tie between the SG network and CAR-T cell addresses a critical gap in the field. This project is expected to provide novel insights into an unexplored area of T cell biology that will not only spur a better understanding of this process but will facilitate the development of novel cancer therapeutics.

NIH Research Projects · FY 2025 · 2024-07

PROJECT SUMMARY/ABSTRACT The rise of antibiotic-resistant microorganisms globally has become a critical public health priority due to the devastating consequences that it may have to the world health and economy. Antibiotic resistance (AMR) threatens the progress of medicine in all areas, and as such, the issue has reached the highest level of government, including the Office of the US President and the United Nations. AMR is a top public health priority for both CDC and WHO and has been designated as the “silent” pandemic. Houston is the home to the largest cluster of healthcare institutions in the world, the Texas Medical Center (TMC), with more than 9,200 hospital beds and 10 million patient visits per year. The Houston area’s strong history of outstanding infectious diseases research and training includes a focus on AMR and antibiotic stewardship. The combined efforts of the recently formed Houston Methodist Center of Excellence for Infectious Diseases and the existing Gulf Coast Consortium (GCC) for Antimicrobial Resistance have resulted in an active, multidisciplinary and comprehensive research and educational program amalgamating the endeavors of AMR researchers and creating the resources, personnel, funding and fertile ground to develop an ambitious, innovative, unparalleled AMR training program. We propose to establish the AMR Training Program in the Texas Medical Center (AMR-TPT) that trains postdoctoral scholars, clinical residents/fellows and PharmD fellows from eight institutions in the TMC (Houston Methodist Research Institute, University of Texas Health Science Center at Houston, MD Anderson Cancer Center, Baylor College of Medicine, University of Texas Medical Branch, Rice University, University of Houston, and Institute of Biosciences and Technology of Texas A&M University) on advanced aspects of AMR research. We will take advantage of the strong administrative expertise of the GCC on successful T32 programs in other areas and the educational activities already in place, combined with the expertise of world-class AMR researchers. AMR-TPT seeks to leverage and create resources to train the next generation of scientists and clinician-scientists (MDs, PhDs and PharmDs) focused on tackling the pressing AMR public health crisis. The highly collaborative environment provides the perfect opportunity for trainees to acquire the skills, expertise and intellectual abilities to foster innovative research that has a strong translational component and could be developed to directly influence patient care. Along with a highly interactive AMR Foundations course and myriad career/professional development opportunities, the proposed training grant includes expertise in and a focus on i) molecular basis of resistance, ii) bacterial genomics and bioinformatics, iii) diagnostics, iv) pharmacological aspects of resistance, v) microbiome science, vi) clinical epidemiology and biostatistics of AMR and vii) antibiotic stewardship. We believe we are poised to continue developing a unique, innovative and comprehensive training program that truly provides trainees with exceptional tools and abilities and creates a strong cohort of new world-class AMR leaders and researchers.

NSF Awards · FY 2024 · 2024-06

The 43rd Annual Conference of Texas Statisticians (COTS): AI, Machine Learning, and Other Related Statistical Techniques with Applications in Healthcare, scheduled for May 9-10, 2024, will be hosted at the Houston Methodist Research Institute (HMRI), situated at 6670 Bertner Ave, Houston, TX 77030, USA. Artificial intelligence (AI) and machine learning (ML) represent a transformative force across various industries, promising advancements in fields such as medical diagnosis, national security, and crime prevention. These technologies harness the power of data to generate models capable of learning, making decisions, and predicting outcomes. Over time, they refine and adapt, becoming more effective and versatile. However, the efficacy of AI and ML relies heavily on the principles and methodologies provided by statistical science. Statistical techniques underpin the construction of robust models in AI and ML, enabling the interpretation of their outputs. This synergy between AI, ML, and statistical science forms the backbone of cutting-edge advancements in data-driven decision-making. COTS, dedicated to AI, ML, and related statistical techniques serves as a crucial platform for statisticians to exchange insights and forge collaborative opportunities. Such gatherings drive innovation, pushing the boundaries of what is achievable with the integration of AI, ML, and statistical science. COTS 2024 is dedicated to advancing the frontiers of AI and ML and related statistical techniques, particularly within healthcare. Experts will explore how AI can revolutionize treatment methodologies by harnessing patient-specific data, genetic profiles, and medical histories to tailor treatment plans with unprecedented precision, optimizing efficacy while minimizing adverse effects. COTS 2024 is committed to assembling a diverse array of leading experts, each bringing unique perspectives and expertise to the table. By fostering a robust scientific forum, the conference aims to facilitate rigorous discussions on the most recent research discoveries, spanning from fundamental research to practical applications aimed at enhancing human health and well-being. Moreover, COTS 2024 recognizes the importance of nurturing collaboration and mentorship within the scientific community. Through various networking opportunities and interactive sessions, the conference seeks to bridge the gap between junior and senior researchers, fostering an environment where knowledge exchange flourishes and innovative ideas take root and support underrepresented groups and minorities. In essence, COTS 2024 is not just a conference; it's a catalyst for transformative change, where cutting-edge research converges with real-world applications to shape the future of healthcare and beyond. Moreover, in collaboration with NSF, COTS 2024 is committed to uplifting underrepresented groups and minorities, ensuring that the benefits of progress are inclusive and accessible to all. NSF’s funding for COTS 2024 has facilitated the integration of diverse perspectives and expertise, driving forward the mission of COTS 2024 to enact meaningful change in healthcare and beyond, while prioritizing the empowerment of underrepresented groups and minorities. Conference website: https://learn.houstonmethodist.org/AI-2024#group-tabs-node-course-default1 This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2026 · 2024-06

PROJECT SUMMARY Chemotherapy remains the most used systemic treatment for cancers. However, despite significant improvement in chemotherapy agents, chemoresistance remains the major problem in cancer management. Recently, we discovered that the selective inhibitors of hepatitis C virus (HCV) NS5A replication complex elbasvir (Elb) and daclatasvir (Dac), an analog of Elb, may be used to sensitize and re-sensitize solid and hematologic cancer cells to chemo-drugs. We screened 1855 FDA-approved drugs and found that Elb was among the top drugs that significantly sensitized multiple myeloma (MM) cells to carfilzomib (Cfz, proteasome inhibitor), dexamethasone (Dex, corticosteroid), and melphalan (Mel, alkylating agent). Elb and Dac could also re-sensitize Cfz- or Dex-resistant MM cells to Cfz or Dex respectively. In addition, we observed that Elb and Dac enhanced chemosensitivity and re-sensitized different types of cancers such as pancreatic ductal adenocarcinoma to 5-fluorouracil and gemcitabine (Gem), estrogen receptor‑positive breast cancer (BC) cells to tamoxifen, and triple negative BC cells to Gem. Moreover, in the presence of Elb or Dac, lower doses of chemo-drugs were required to induce similar cancer cell death compared to chemotherapy drugs alone. We discovered that Elb and Dac significantly enhanced drug retention in cancer cells by inhibiting drug efflux through ATPase phospholipid transporting 9B (ATP9B). Importantly, although tumor microenvironment (TME) components such as tumor-associated stromal cells (TASCs) and tumor-associated macrophages (TAMs) can protect cancer cells from chemotherapy-induced cell death, Elb and Dac abrogated TASC and TAM protective effects by suppressing pro-tumor lipid secretion, reprogramming them to secrete type-I-IFNs for sensitizing MM cells to Cfz, and inhibiting TAM release of deoxycytidine for overcoming Gem resistance in solid tumors. Finally, Elb and Dac significantly improved the therapeutic efficacy of chemotherapies in MM in vivo without increased toxicity to normal tissues. Therefore, we hypothesize that HCV NS5A inhibitors Elb and Dac can be developed into cancer therapeutic agents due to their ability to (re)sensitize cancer cells to chemotherapies by enhancing chemo-drug retention in tumor cells and removing TME-provided protection. To minimize the scope of this application, we will focus on human MM (hematological malignancy) and use human MM cell lines and primary MM cells from patients and MM PDX mouse models to test the hypothesis. Aim 1 will elucidate the role and mechanism of Elb and Dac in sensitizing tumor cells to chemotherapy by inhibiting chemotherapeutic drugs efflux through ATP9B and Aim 2 will elucidate the role and mechanisms of Elb and Dac in overcoming TME-mediated extrinsic resistance by inhibiting ATP9B-mediated cholesterol uptake and deoxycytidine efflux. Accomplishing these aims will provide the justification and tools for developing novel and effective strategies for targeting both cancer drug efflux and TME to improve the therapeutic efficacy of chemotherapy.

NIH Research Projects · FY 2026 · 2024-06

PROJECT SUMMARY/ABSTRACT This K23 proposal outlines a five-year research and training plan that will accelerate Dr. Max Adelman’s career as an independent physician-scientist with expertise in Candida colonization and infection in critically ill patients. Dr. Adelman is an infectious diseases and critical care clinician whose translational research focuses on severe infections among patients in the intensive care unit (ICU), a population at high risk of infectionrelated morbidity and mortality. These patients are particularly susceptible to infections with Candida spp., which are the second-leading cause of ICU-onset bloodstream infection and are associated with up to 60% mortality. Before developing infection, patients are first “colonized” with Candida, and the primary site of Candida colonization is the gut. Importantly, while gut colonization is a risk factor for Candida bloodstream infection, there is little data on the host and Candida characteristics that predispose to gut colonization itself. Additionally, Candida bloodstream infections are increasingly caused by multidrug-resistant species including C. glabrata and C. auris, but whether these species consistently colonize the gastrointestinal tracts of ICU patients has not been determined. In this proposal, Dr. Adelman will test the hypothesis that several host and pathogen-specific factors facilitate gut colonization with Candida, which in turn affects important clinical outcomes. In Specific Aim 1, he will (a) determine whether broad-spectrum antibiotics commonly used in the ICU predispose to Candida colonization, (b) evaluate the impact of colonization on clinically important outcomes using a desirability of outcomes ranking (DOOR) analysis, and (c) determine the genomic epidemiology of gut colonizing antifungal-resistant Candida spp. In Specific Aim 2, he will examine the immune pathology that facilitates Candida colonization in the ICU by expanding on his preliminary data linking defective IFN-ã production with colonization. To accomplish these goals, Dr. Adelman has designed a training plan that builds on his strong clinical research background with advanced training in data science, Candida genomics, and host-pathogen interactions. Dr. Adelman’s research will be overseen by dedicated mentors from Houston Methodist Hospital and surrounding institutions in the Texas Medical Center with complimentary expertise in translational research, microbial genomics, Candida pathogenesis, and immune control of Candida. Additionally, this project will leverage extensive data and sample collection from an NIH-funded P01 project of gut bacterial colonization in ICU patients (AI152999) led by Dr. Adelman’s primary mentor, Cesar A. Arias, MD, PhD. Overall, this integrated training and mentorship plan will support Dr. Adelman in his discovery of factors that lead to Candida colonization in critically ill patients. Through the proposed award, Dr. Adelman will develop into an independent physician-scientist with clinical infectious diseases and critical care expertise poised to improve care for patients at high risk of Candida infection.

NIH Research Projects · FY 2025 · 2024-06

Abstract Technological advancements have yielded novel drug targets and improved cancer detection, but transformative innovations in treatment modalities are lacking. Leveraging our team's expertise in gene therapy, breast cancer, cancer immunology, and breast cancer animal and patient-derived xenograft models, we propose to use recombinant adenovirus with a higher tumor-specific expression of herpes simplex virus-thymidine kinase (HSV-TK), which is a suicide gene that causes tumor cell death. We will combine HSV-TK with our immune checkpoint antibody (i.e. HMR-101) that will specifically target tumor cells, release tumor antigens and change the tumor microenvironment to be more favorable for immune therapy. The immunotherapy we are developing will modify cancer-promoting immune cells to become anti-cancer immune cells. Therefore, an army of anti-cancer immune cells trained to recognize cancerous cells will mobilize and circulate around the body to destroy metastases. Given the predominance of tumor-associated immunosuppressive myeloid cells in triple-negative breast cancer, local delivery of HMR-101 could effectively reprogram the tumor microenvironment to an anti-tumor immune phenotype. The viral transduction efficiency of our viral vector is very high at the tumor site with chimeric viral vector (low neutralization antibody), and it will be low spreading to other organs. This viral vector is safe and approved by the FDA and at low cost for human use. As such, our intratumoral strategy would allow for maximizing the full potency of the immunotherapeutic antibody, thus enhancing therapeutic index. We envision this technology to move forward to clinical trials, strongly supported by the success of our recently completed STOMP trial for metastatic triple-negative breast cancer or TNBC (NCT03004183, HSV-TK, radiation, and immune checkpoint therapy on metastatic TNBC). Clinically, we envision that our intratumor gene delivery of HMR-101 will reprogram the immune system, prevent patients from relapse, and prolong anti-tumor immunity, thus benefiting patients receiving immunotherapy. For triple-negative breast cancer patients who typically have dismal clinical response due to high metastatic rate and recurrence, the potential clinical impact of our intratumoral viral delivery containing novel immune checkpoint antibody is profound. Overall, the therapeutic reach of our novel therapeutic regimen is expansive, offering a truly transformative approach to cancer treatment.

NIH Research Projects · FY 2026 · 2024-05

Project Summary Adoptive cell therapy (ACT) with chimeric antigen receptor (CAR) T cells has demonstrated impressive response rates in B cell malignancies, but ACT has not mediated sustained responses in solid tumors. CD19 CAR T cell therapy has reached up to 80% response rate in the clinic; however, the main side effects are cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS), which occur in 37–83% and about 25-35% of patients, respectively. Furthermore, one of the major obstacles in ACT is the heterogeneity of targeted antigens and relapse due to antigen escape. Recently, we screened a cohort of 16 FDA-approved anti-inflammatory drugs and identified clofazimine (CLF) as the top candidate for its desired bifunctional effect for anti-CRS/ICANS and anti-antigen escape roles. Aim 1 will determine the role of CLF in reducing macrophage-derived ROS to curtail CRS/ICANS. Aim 2 will determine the role of CLF in driving dsRNA/dsDNA signals in macrophages for the eradication of tumors. We expect this study to demonstrate the ability of CLF in potentiating the anti-antigen escape capacity in ACT, curbing intractable CRS, and may also fill a desperate clinical need to improve the dismal patient survival with ICANS. This strategy of repurposing the clinically approved CLF may hold great promise to overcome a critical obstacle in realizing the full potential of ACT with CAR-T cells. This translationally relevant work could then lay the foundation for future clinical trials.

NIH Research Projects · FY 2026 · 2024-04

PROJECT SUMMARY The mTOR pathway is a master regulator of nutrient metabolism and cell proliferation in response to environmental cues. It has long been viewed as an important driver of tumorigenesis and an ideal cancer therapeutic target. Multiple mTOR inhibitors have been tested in more than 500 clinical trials, either as single agents or in combination with chemotherapy for cancer treatment. However, among nearly 200 completed trials, few show positive outcomes, challenging the utility of mTOR as an effective cancer therapeutic target. As an integrator of extracellular signals and cellular responses, the mTOR pathway is critical for cellular adaptation to environmental changes. In particular, mTOR inhibition induces embryonic diapause, a reversible dormancy state in development in response to unfavorable environment cues. We reason that, analogous to its role in embryonic diapause, mTOR inhibition may induce an adaptive diapause-like dormant state in cancers, leading to the persistence of residual tumors following chemotherapy. In line with this notion, recent studies have indeed reported markedly reduced mTOR activity in residual tumors. However, a causal relationship between mTOR inhibition and tumor persistence has not been established or investigated. Integrating multiple state-of-the-art approaches, our recent studies show that mTOR inhibition indeed induces a diapause-like persister state in tumor cells. We confirm that this is a pan-cancer phenomenon using tumors from diverse tissue origins and demonstrate that the persister state recapitulates the residual tumors in patients following chemotherapy. Importantly, our studies identify ferroptosis as a key druggable vulnerability of persisters. Built on these compelling findings, we hypothesize that mTOR is a master regulator of a diapause-like persister state in tumors by modulating survival and dormancy. The current proposal aims to investigate the mechanistic regulation of the persister state, and test therapeutic strategies to target residual tumors in preclinical models, with the ultimate goal to prevent tumor recurrence in patients.

NIH Research Projects · FY 2026 · 2024-04

IMAT R61 Abstract In addition to its well-recognized roles in neuronal computation and cardiac contractions, the voltage across the plasma membrane has been implicated as a fundamental regulator of differentiation and proliferation. The concurrent regulation of proliferation and stem cell states by membrane voltage contributes to normal tissue morphogenesis, development, and diseases such as cancer. Membrane hyperpolarization tends to promote increased differentiation and reduced proliferation while depolarization is associated with immature and proliferative stem-like cell states. Of clinical relevance, cancer cells and cancer stem-like cells exhibit depolarized membrane voltage values compared with the normal differentiated cells from which they arise. Despite the strong association between voltage and cancer cell phenotypes, voltage modulation has not been effectively exploited as a drug target against cancer, largely reflecting the technological challenges with membrane voltage measurements in physiologically relevant contexts. Current techniques to measure membrane voltage, including electrode arrays and voltage sensitive dyes, have limitations in the number of experimental observations possible, compatibility with complementary phenotypic readouts, the duration of sampling, and the inability to perform longitudinal cell analyses. These technologic challenges motivate our hypothesis that integration of genetically encoded voltage and cell cycle sensors will provide a robust and novel high-throughput platform to identify modulators of membrane voltage and cell proliferation. We propose to develop, optimize, and validate a cell-based technology platform integrating JEDI, a novel voltage indicator, and FUCCI cell-cycle sensors to enable real time high-throughput analysis of concurrent changes in voltage and cell proliferation in living cells. To build the platform, we propose to demonstrate high-throughput imaging of voltage dynamics (Aim 1) and cell cycle states (Aim 2) in glioblastoma stem cells. We next propose to deploy these technologies for high-throughput quantification of the impact of ion channel-modulating drugs on the membrane voltage and cell-cycle changes (Aim 3). This novel platform is expected, for the first time, to provide a tool for real-time live cell readouts of concurrent changes in membrane voltage and cell proliferation. Besides its implications for the treatment of cancer, we anticipate this platform to have far reaching basic science and clinical translational applications in neuroscience, stem cell and iPSC biology, cell-based therapies, and cardiovascular diseases.

NIH Research Projects · FY 2024 · 2024-02

Project Summary The overall incidence of thyroid cancer in 2016 was estimated by the National Cancer Institute to be 64,300. Patients who present with poorly differentiated thyroid cancers frequently are refractory to standard treatment regimens and have a much worse prognosis. The median overall survival in this patient population is less than a year. Undifferentiated or anaplastic thyroid cancers are typically not amenable to surgery and are highly resistant to RAI and virtually all other therapies. We therefore developed a chimeric antigen receptor (CAR) T cell therapy targeting intercellular adhesion molecule-1 (ICAM-1) (labeled as AIC100) to treat this aggressive type of thyroid cancer. While a variety of cells in the body normally express low, basal levels of ICAM-1, many human cancers have upregulated levels of expression. In particular, both refractory poorly differentiated and anaplastic thyroid cancers have greatly increased expression of ICAM-1. The key aspect of our CAR T cell technology is its ability to selectively kill tumors with over-expressed ICAM-1 while sparing normal cells with basal levels of ICAM-1 expression. To assess in vivo distribution of CAR T cells in both targeted tumors as well as non-target tissues, we have introduced the somatostatin receptor 2 (SSTR2) to follow CAR T cells over time using the clinically approved radiolabeled tracer 68Galium-DOTATATE (Netspot). In the planned phase I study, the primary objective is to assess the safety and determine the recommended dose of AIC100 for phase II study in patients with relapsed/refractory poorly differentiated thyroid cancer and in patients with anaplastic thyroid cancer. The secondary aims are to evaluate the efficacy of AIC100 in patients using PET/CT and RECIST (Response Evaluation Criteria In Solid Tumors) criteria, evaluate the feasibility of CAR T cell imaging by DOTATATE, determine if overall tumor response correlates with T cell distribution in tumor sites as measured by DOTATATE and other exploratory biomarkers, and to examine pre-infusion CAR T characteristics as a predictor of clinical response. Our investigational new drug (IND) application to open phase I study of AIC100 against advanced thyroid cancers is now approved by FDA in October 2019, and patient enrollment will commence in early 2020. As the cost for CAR T manufacturing and non-standard of care clinical costs will be sponsored by our industry collaborator (AffyImmune Therapeutics, Inc.), the major goals of this grant application are to assess the kinetics of T cell distribution by PET/CT, and determine the emergence of high clonality T cells and associated cellular and gene expression signatures with respect to clinical response, toxicity, and survival; to correlate intrinsic CAR T fitness for survival and clonal expansion with clinical response; to optimize T cell manufacturing and next-generation CAR designs to improve the fitness of CAR T cells toward more effective CAR T therapies against solid cancers.

NIH Research Projects · FY 2026 · 2024-02

Project Summary Disruption of normal mitochondrial bioenergetics and oxidative phosphorylation represents an early event during oncogenesis by changing the energy metabolism of precancerous and cancerous cells. Phenethyl isothiocyanate (PEITC), a natural compound present in cruciferous vegetables, has been shown to inhibit the development of several types of cancer in animal models. PEITC has been shown to inhibit oxidative phosphorylation and to induce cancer cell apoptosis through a mitochondria-dependent mechanism and ROS formation, suggesting the role of mitochondrial bioenergetic function and redox homeostasis in oncogenesis. This provides the rationale for conjugating PEITC to a targeting agent that drives it into mitochondria to specifically study the role of mitochondrial function and ROS formation in lung cancer development and to increase the efficacy of PEITC. Preliminary studies demonstrate that mitochondria-targeted PEITC (Mito-PEITC) is a significantly more potent inhibitor of lung carcinogenesis in mice. We hypothesize that lung oncogenesis and metastasis depend on mitochondrial respiration and that Mito-PEITC is a novel, potent inhibitor of lung carcinogenesis and metastasis acting primarily through mitochondrial mechanisms. This hypothesis will be tested using three specific aims. Aim 1 will evaluate the preventive potential and mechanisms of action of Mito-PEITC in vitro. We will use Mito-PEITC and lung cancer cells with genetically modified expression of the members of the mitochondrial electron transport chain to decipher the oncogenic mechanism and evaluate the mechanisms of action of Mito-PEITC. Aim 2 will determine preventive efficacy of Mito-PEITC on lung tumor initiation and progression in A/J mice. Aim 3 will determine the in vivo mechanism and the efficacy of Mito-PEITC to inhibit lung cancer brain metastasis. We will use state-of-the-art in vitro and in vivo assays, including small animal imaging technology to monitor the growth of primary tumors (magnetic resonance imaging) and engraftment of metastatic cells. Innovative approaches for in vivo monitoring of changes in cancer cell bioenergetics and cellular oxidant production (bioluminescence imaging) will be employed. Defining mitochondrial mechanisms of lung oncogenesis and brain metastasis and developing a novel, well-tolerated efficacious agent for prevention and treatment of lung cancer will be highly significant.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY Adoptive cell transfer using tumor-specific T cells is a promising option for cancer treatment and the most powerful tumor-killing effector T cells are CD8+ type-1 cytotoxic T cells (Tc1). We have shown that IL-9- secreting CD8+ Tc9 cells mediate stronger and long-lasting antitumor effects in vivo compared to the classical IFN-γ-secreting Tc1. However, the underlying mechanisms remain unclear. Recently, we discovered that, when we rechallenged Pmel-1 Tc9- or Tc1-treated mice on the contralateral flank with gp100-knockout (KO) B16 tumor cells, these cells rapidly grew and established tumors in Tc1 but not Tc9-treated mice. Similarly, relapsed gp100– B16 cells recovered from Pmel-1 Tc1-treated mice rapidly grew and established tumors after re-implanting to Pmel-1 Tc1- but not Tc9-treated mice, suggesting that Tc9- but not Tc1-treated mice develop a host immunity against other (than the cognate) antigens expressed by relapsed tumors. Importantly, significantly larger numbers of CD4+ but not CD8+ T cells were detected in late-stage tumors, rechallenged tumors and tumor-draining lymph nodes (TDLNs) of Tc9-treated mice compared to Tc1-treated mice. These CD4+ T cells expressed high levels of IFNγ and granzyme-B and effectively killed wild-type B16, gp100– relapsed and gp100-KO B16 tumor cells. Hence, these novel findings strongly indicate that adoptively transferred Tc9 but not Tc1 cells effectively induce a host CD4+ T cell response against relapsed tumors. To determine the mechanism underlying Tc9 cell-induced host CD4+ T cell response, we performed preliminary studies to examine immune cells in tumor microenvironment (TME). We observed that dendritic cells (DCs) were enriched in Tc9-treated primary tumors, rechallenged tumors, and TDLNs. More importantly, Tc9 cells secreted a high level of IL-24, and Tc9-treated tumor supernatants contained abundant IL-24. As our preliminary studies showed that IL-24 attracted DC migration in vitro, we speculate that Tc9 cells induce a host CD4+ T cell response through IL-24-DC circuit. We hypothesize that tumor-specific Tc9 cells may be a superb T-cell subset for cancer immunotherapy due to their capacity to elicit a host CD4+ T cell response to suppress the growth of relapsed tumors. To test our hypothesis, Aim 1 will determine the role and importance of Tc9 cell- induced host CD4+ T cell responses in recognizing and eliminating relapsed tumor cells, and Aim 2 will determine the role and mechanism of IL-24-DC circuit in TME and TDLNs in eliciting tumor-specific CD4+ T cell responses and preventing the recurrence of tumors. Completing this project will uncover a novel mechanism of Tc9 cells in mediating long-lasting antitumor activity in TME by inducing host CD4+ T cells to recognize tumor- associated antigens or neoantigens via IL-24-DC circuit and suppressing or preventing tumor recurrence in Tc9-treated mice.

NIH Research Projects · FY 2026 · 2023-12

Project Summary Rejection remains a major hurdle to long-term transplant survival, in which Teff cells are prominently involved. We surmise that besides signals from the TCR, costimulatory and cytokine receptors, targeting the “T cell epigenome” downstream of T cell activation may represent novel therapeutic opportunities in the induction of transplant tolerance. Now we have new preliminary data showing that the chromatin in Teff cells are highly compartmentalized, in that “super enhancers” at accessible chromatin regions appear to control a Teff cell fate by recruiting a chromatin reader called BRD4. We then developed a Brd4-floxed mice and showed that conditional deletion of Brd4 in T cells completely abrogated the lethal autoimmune phenotype in Scurfy mice and produced long-term allograft survival in Brd4f/fCd4-Cre mice. Thus, dissecting mechanistically how BRD4 epigenetically regulates Teff cells and transplant survival is the central goal of this proposal. Our working hypothesis is that BRD4 occupancy at the accessible chromatin regions locks active chromatin modules in an accessible state or triggers reorganization of “super enhancers” into transcriptionally hyper-active “hotspots” to drive a Teff cell fate. We proposed 3 Aims in this proposal to test this hypothesis. Aim 1 is to determine whether BRD4 locks “active” chromatin modules in an accessible state, allowing the formation of 3D chromatin configurations to specify a Teff cell fate, and Aim 2 is to examine whether BRD4 enables stable SE “hotspots”, allowing Teff cells to establish essential features of Teff profiles. Aim 3 is to test whether therapeutically targeting BRD4 enables long-term allograft survival in a heart transplant model. We believe that the proposed studies will uncover new insights and open new therapeutic opportunities in transplantation. The animal models, tools, and cutting-edge technologies we have developed in the lab put us in a unique position to carry out the proposed studies.

NIH Research Projects · FY 2025 · 2023-09

Abstract: (30 lines) Vancomycin is one of the most commonly used antimicrobial drugs in inpatient settings. National guidelines recommend Bayesian models to monitor the therapeutic drug concentration of vancomycin, especially for methicillin-resistant Staphylococcus aureus (MRSA), to minimize drug toxicity while maintaining its efficacy. Existing Bayesian models, despite being claimed as patient-specific pharmacokinetic (PK) models, use simple patient features and are studied in limited patient populations for the population-based PK parameters (the Bayesian prior). Increasingly available real-world electronic health records (EHR) provide a wide range of patient-specific data, including data on vancomycin dosage and serum levels. However, the limited flexibility of the Bayesian model structure prohibits the full use of these rich data. Deep-learning models, such as recurrent neural network (RNN), are particularly attractive for PK of vancomycin in EHR, compared to Bayesian models and other traditional machine learning models, because deep-learning models enable more flexible patient- specific inputs and possess a higher latent capacity. Thus, they deliver better predictions for a diverse population. Our deep-learning model for vancomycin (PK-RNN-V) outperforms publicly available Bayesian models but can be improved on various aspects. In Aim 1, we will improve PK-RNN-V model architectures and add more patient-specific data and a finer timestep. We will construct two-compartment PK-RNN models to increase predictive power in patients with unsteady states. We will augment PK-RNN-V with Med-BERT to improve the embedding of categorical data. We will also develop multi-track ordinary differential equations models for simultaneous prediction of serum creatinine and vancomycin levels. In Aim 2, we will use EHR from different sources to validate our PK-RNN-V model and improve the data-extraction flow and pre-processing to harmonize data from healthcare systems. We will use EHR from Houston Methodist Hospital and Memorial Hermann Hospital System/The University of Texas Health Science Center in Houston, TX, the University of Arizona in Phoenix, AZ, and the publicly available MIMIC-IV database (Boston, MA). These databases contain data from more than 121,007 patients who received at least one dose of intravenous vancomycin. In Aim 3, we will add dosing recommendations based on PK-RNN-V model predictions as a feature and validate our model in specific subgroups with challenging vancomycin PK to predict PK levels. This project will deliver substantial model improvements, leading directly to the optimization of vancomycin use in hospitals, increased in patient safety by minimizing adverse events, and reduced healthcare costs, which align with NIH’s research mission.

NIH Research Projects · FY 2025 · 2023-09

Abstract There is growing interest in the use of electrical stimulation to promote the recovery of sensorimotor and autonomic function after neural injury. Previous research from our lab and others has demonstrated that stimulation of spinal lumbar segments activates central pattern generators, which, in turn, facilitates standing and walking. However, while there is extensive animal, pre-clinical, and clinical data examining the impact of lumbar stimulation, studies that apply neuromodulation to the cervical spinal cord for upper limb are very limited. Here, we propose that fundamental blind spots exist in the field of neuromodulation for upper limb, including where to stimulate anatomically, how to dose and, especially, mechanisms of action. The Horner lab has developed a clinically relevant rat model of cervical spinal cord injury and engineered a self-contained epidural stimulation device that can be deployed in freely behaving rats to stimulate sensorimotor circuitry from multiple surfaces of the spinal cord. Hence, we are well positioned to test critically important hypotheses on the role of cervical stimulation in the restoration of upper limb function. We present exciting preliminary data demonstrating that epidural stimulation of the cervical spinal cord improves forelimb function. The rationale for the proposed research is that the site of epidural stimulation provides unique access to motor circuitry. We hypothesize that ventral positioning of electrodes (VSS) will provide access to stimulate motor circuitry at the site of lesion that are inaccessible from the more common dorsal approach (DSS). Further, we propose that VSS will produce novel mechanisms of function plasticity that can amplify recovery when combined with DSS. To test this hypothesis, we propose the following aims: Aim 1: Determine acute molecular and physiological mechanisms of VSS when applied to subacute cervical spinal cord injury. Aim 2: Establish the functional impact of site of stimulation and rehabilitative training on recovery from early chronic cervical spinal cord injury. Aim 3: Establish the synergistic effects of combined VSS and DSS after cervical spinal cord injury. These studies will explore an exciting new approach to promote neural recovery of the upper limb, an area of research that has had limited investigation, but remains a primary concern for the patient. Our approach will rigorously establish the physiological and functional effects of the site of stimulation on the molecular and physiological mechanisms of upper limb plasticity.

NIH Research Projects · FY 2025 · 2023-09

Resistance to standard of care (SOC) temozolomide (TMZ) and radiation treatment (XRT) limits survival of the most common adult brain cancer, glioblastoma (GBM), to 12-18 months. Despite promising results in other cancers, robust immunosuppression in GBM has also limited the responses to immune checkpoint inhibitors (ICIs). Reciprocal interactions in the GBM tumor microenvironment (TME) between mesenchymal changes and immunosuppression enhance resistance to chemoradiation and immunotherapy, respectively. Therefore, a compelling therapeutic strategy for GBM is to concurrently reprogram the mesenchymal and immune suppressive TMEs and potentiate ICI and SOC therapy. The NR4A1 and NR4A2 orphan nuclear receptors are compelling targets to achieve this goal. We synthesized a series of novel bis-indole–derived ligands (CDIMs) with potent dual antagonism of NR4A1 and NR4A2 and negligible in vivo toxicity, of which six demonstrated nM range KDs for both receptors. Genetic based NR4A 1 and 2 loss of function strongly activates anti-cancer immune responses by reversing T-cell exhaustion which underlies poor ICI responses in GBM. In addition, NR4A1/2 promote EMT in other cancers which contributes to chemoradiation resistance. Finally, high levels of NR4A1/2 expression in GBM are strongly associated with decreased patient survival after conventional treatments. Therefore, we propose to test the hypothesis that CDIM inhibition of NR4A1/2 reprograms the GBM TME and potentiates ICI and TMZ/XRT responses. Further NR4A1/2 upregulates PD-L1 and TWISt1, key regulators of GBM immune suppression and mesenchymal phenotypes, respectively. Preliminary studies demonstrated that CDIMs inhibit PD-L1 and TWIST1 expression, reverse immune suppressive myeloid and T cell phenotypes and prolong survival of experimental GBMs. We will test our hypothesis by i) identifying lead dual NR4A1/2 CDIM compounds based on inhibition of malignant and mesenchymal GBM cell properties (proliferation, self-renewal, invasion) and reversal of dysfunctional T cell phenotypes in vivo (Aim 1), ii) establishing the functional impact of NR4A1/2 CDIMs to reprogram the immune suppressive and mesenchymal TME (Aim 2) and iii) quantifying the effects of NR4A1/2 CDIMs to potentiate ICI and SOC responses in mouse GBM syngeneic models and define the role of PD-L1 and TWIST1 in their mechanisms of action (Aim 3). Quantification of potency, pharmacokinetics and preclinical anti-tumor activity alone and in combination with to standard of care TMZ-XRT and ICIs is expected to identify candidate CDIMs for consideration in future clinical trials. Through application of powerful bioinformatic tools that model the cancer TME through integration of scRNAseq, cyTOF and IMC data, these studies are expected to shed new light on novel mechanisms by which mesenchymal and immune suppressive TMEs interact to coordinately drive treatment resistance in GBM through NR4A1/2. To achieve our goals, we assembled a unique multidisciplinary team with expertise in medicinal chemistry, neuro-oncology, GBM biology and preclinical modeling, and immuno-oncology.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY Recently, we made a novel and exciting discovery that tumors or the tumor microenvironment (TME) cause T- cell dysfunction and death by inducing ferroptosis in T cells. We analyzed the sc-RNA-seq data of tumor- infiltrating T cells from melanoma patients and discovered that tumor-infiltrating CD8+ T cells had significantly increased expressions of genes associated with lipid peroxidation and ferroptosis compared to blood CD8+ T cells from healthy individuals. More importantly, our unpublished studies further revealed that among different CD8+ T cell subpopulations, effector memory (TEM) and terminally differentiated effector (TTE) CD8+ T cells are more sensitive to tumor-induced ferroptosis. We examined tumor-infiltrating CD8+ T cells from the bone marrow (BM; tumor bed) of patients with multiple myeloma. By separating the T cells into naïve, TTE or TEM cells based on their expression of CCR7 and CD45RA, we found that TEM and TTE CD8+ T cells expressed higher levels of lipid peroxidation- and ferroptosis-associated genes and were more sensitive to tumor-induced ferroptosis compared to naïve CD8+ T cells although they expressed similar levels of CD36. Similarly, in mouse melanoma and MM models, increased ferroptosis mainly occurred in tumor-infiltrating CD8+ TEM and TTE cells but not in naïve CD8+ T cells from mice with large tumor burdens compared to those with small tumor burdens. Our ex vivo studies confirmed that CD8+ TEM and TTE cells were more sensitive to tumor- or FA- induced ferroptosis than naïve T cells and their production of cytotoxic cytokines such as IFNγ and TNFα was inhibited. To elucidate the underlying mechanisms, RNA-seq was used and showed that CD8+ TEM and TTE cells expressed a significantly lower level of 2,4-dienoyl-CoA reductase 1 (DECR1), a rate-limiting enzyme for polyunsaturated fatty acid (PUFA) β-oxidation, compared to naïve CD8+ T cells. Knockdown (KD) of DECR1 in naïve CD8+ T cells resulted in an increased PUFA expression and peroxisomal dysfunction and sensitized them to tumor- or FA-induced ferroptosis than control T cells. Based on these novel findings, we hypothesize that CD8+ TEM and TTE cells, due to uptake of more FAs and reduced expression of DECR1 and PUFA oxidation, are sensitive to tumor/TME-induced ferroptosis, and inhibiting TEM and TTE cell ferroptosis may effectively enhance the therapeutic efficacy of immunotherapy in cancer patients. Aim 1 will determine the mechanism underlying tumor- or FA-induced ferroptosis in CD8+ TEM and TTE cells in TME. Aim 2 will elucidate the role and mechanisms of tumor and TME accumulation of FAs and induction of lipid peroxidation in CD8+ TEM and TTE T cells and Aim 3 will inhibit CD8+ TEM and TTE cell ferroptosis to enhance the efficacy of cancer immunotherapies. Accomplishing these aims will provide us with in-depth understanding of the mechanisms underlying how tumors and the TME induce T cell lipid peroxidation and how ferroptosis mediates T cell metabolic malfunction and death. Understanding these mechanisms is extremely important and will greatly assist us in developing novel therapeutic approaches to target T cell lipid metabolism and TME to significantly improve the efficacy of cancer immunotherapy.

NIH Research Projects · FY 2024 · 2023-09

PROJECT SUMMARY / ABSTRACT Alzheimer’s disease (AD) is a progressive neurodegenerative disease that results in cognitive decline. The neuroinflammatory events that are associated with the pathology of AD exacerbate neurodegeneration. Elderly people with dementia or AD have a higher risk of falling because their cognitive function is affected. They lose their orientation and are very prone to breaking their hips, other bones, or even more severe falls that can cause brain trauma. Although they are treated in the hospitals with subsequent rehabilitation, their weakened immune state typically affects their recovery phases. We need to develop new treatments for AD patients who have had a traumatic brain injury (TBI). This vulnerable population could be particularly amenable to precision-microbiota therapy. Although not fully understood, the brain-gut-microbiota axis plays a major role in the onset and severity of many neurological diseases. We plan to develop precision-microbiota therapy to protect against neurodegeneration and functional behavior in AD mice after TBI. The gut microbiome is emerging as an essential neuromodulator of brain-gut axis signaling. It can significantly impact brain inflammation and outcome after CNS trauma and alter anxiety- and depression-like behaviors. This research proposal is based on the scientific premise that gut bacteria can impact behavior and that brain injury can disrupt diversity in a healthy gut microbiome. The overarching hypothesis is that the gut microbiota dysfunction caused by brain trauma contributes directly to neuroinflammation and neurodegeneration responses in AD mice, and the microbiota modulation will delay the progression of AD neuropathology. Our preliminary data indicate how the microbiota of AD mice affected adversely to TBI outcomes and how probiotics help brain recovery in C57B6/J young mice. This proposal investigates the mechanistic linkage between gut microbiota and AD progression following TBI and explores potential intervention strategies. We will test this premise with the following three Specific Aims: we 1) will determine if the gut microbiome plays a role in accelerating AD pathology and cognitive decline induced by TBI, and 2) will investigate the impact of fecal transplants on AD pathology and cognitive decline after TBI, and 3) will assess the ability of multi-strain probiotics to reduce AD pathology after TBI. We expect to identify a particularly vulnerable patient population by defining which inflammatory responses associated with brain trauma accelerate AD pathogenesis and are regulated by brain-gut-microbiome signals. Our approach is significant because we will establish a “translatable” foundation for the potential of therapeutic approaches for AD after TBI by using the gut as a non-CNS target of precision-microbiota therapy.

NIH Research Projects · FY 2025 · 2023-09

Project Summary/Abstract Malignant brain tumors are aggressive cancers that have high proliferative rates with much higher energy requirements and high mortality rates. Despite intense clinical and drug development efforts in the last two decades, there has been no improvement in survival. To support the abnormal growth commonly seen in tumors, the cancer cells have altered their metabolism compared to normal cells in healthy tissues. Most of the knowledge to date on cancer metabolism is derived from cultured cell lines. Probing metabolism in intact tumors will be critical to understand how the tumor cells grow in a patient under the complex biological tumor environment. From our pilot study involving a small number of patients, we have demonstrated that gliomas and brain metastases have the capacity to oxidize acetate in the citric acid cycle to meet their bioenergetic requirements, and glucose and acetate together contribute up to 63.0% of the total acetyl-CoA pool in these tumors. The remaining acetyl-CoA that provides carbon sources for biomolecular synthesis, must be derived from other nutrients. The following are the goals of this proposal: (1) determine if acetate and ketone body (beta hydroxybutyrate, BHB) utilization is a common property of all gliomas or specifically linked to high grade GBMs (2) examine whether acetate and BHB provide carbons for 2-hydroxyglutarate (2-HG) synthesis in IDH mutated glioma patients (3) preclinical testing of the effects of small molecule inhibitors of acetate and BHB, in freshly resected tumor tissue slices. We have Institutional Review Board (IRB) approved clinical protocol to infuse non-toxic and non-radioactive 13C-enriched acetate in patients who will be undergoing surgical removal of a brain tumor. Using Nuclear Magnetic Resonance (NMR) spectroscopy and mass spectrometry of these surgically resected tumor tissues, we will investigate the above described aims on energy metabolism of primary brain tumors. The attractiveness of this technology is that no radioactivity is involved. We anticipate that the outcome of this study will generate a detailed understanding of in vivo utilization of acetate and ketone body in brain tumor patients. This knowledge will lead to identification of key metabolic targets that may be further exploited for the development of new therapies. Additionally, it may identify novel biomarkers which may be helpful in designing non-invasive in vivo MRI methods to track acetate utilization by tumors for diagnostic purposes.

NIH Research Projects · FY 2025 · 2023-09

The overall goal of this project is to develop novel mathematic methods and toolkits to connect cell fate transition and epigenetic regulation across tissues and diseases. Cell fate transition often occurs in organ development, tissue regeneration, and pathogenesis. Dysregulation of the cell fate transition can lead to abnormal development or diseases, such as type 2 diabetes, obesity, heart failure, and Alzheimer’s disease. Quantitively decoding how cell fate changes can provide novel mechanistic insight into organogenesis and tissue regeneration, and help identify new strategies for the treatment of human diseases. However, our knowledge of cell fate transition and its regulation is only the tip of the iceberg due to the impracticality of long- term tracing of cell transcriptomes. In the past decade, numerous single-cell atlases containing millions of cells in different tissues, organs, developmental stages, and biological conditions are routinely developed by consortia such as the Human Cell Atlas (HCA). These atlases provide an opportunity for the unbiased study of cellular dynamics and the regulation mechanism. The lack of computational methods presents a major knowledge gap in the understanding of cell dynamics and the regulation leveraging by those large reference atlases. To address this knowledge gap, we proposed a new concept of “reference-based cellular dynamic inference”, which is a novel strategy to automatically annotate the cell state transition in new datasets by learning from the appropriate reference, allowing us to easily perform comparative analysis among different tissues and disease conditions. In this project, we will pursue three parallel but complementary research directions: 1) to develop the first computational methods and toolkits for generating cell dynamics atlases and analyzing cell state transition based on the appropriate reference atlases; 2) to develop novel statistical models for studying epigenetic regulation of cell fate from single-cell multiomics data; 3) to generate the first dynamic reference landscapes of cell differentiation, such as cardiogenesis, hematopoiesis, and neurogenesis, and in- house landscapes of transdifferentiation. This project will be built on the foundation of our recent studies for the development of computational approaches to uncover cell state transition from single-cell transcriptomes in both homogeneous and heterogeneous cell populations and the studies for investigating the role of epigenetic regulation on cell fate transition. The proposed studies will generate advanced computational toolkits and broadly applicable dynamic reference atlases, which are expected to reveal profound mechanisms controlling cell state transition in health and disease. In the long term, the ability to build cell dynamics reference landscapes will open a new horizon to understand the diversity of cell fate through comparative analyses across tissues and diseases and enhance regenerative medicine.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY Approximately 1 in 7 US adults have chronic kidney disease (CKD), and >800,000 patients are in full kidney failure also called end-stage kidney disease (ESKD). The optimal treatment for ESKD is living kidney donor transplantation (LKDT); however, the standard of care continues to be ongoing dialysis, which has poor clinical outcomes in comparison to LKDT. CKD continues to increase among Hispanics and low-income patients. Black and Hispanic/Latino persons are respectively 3.1- and 1.3-times more likely than White persons to develop ESKD. Many individuals are curious about becoming a living donor, but some decide not to donate for unknown reasons. This prevents transplant centers and kidney organizations from improving their kidney donation process and tailoring donor care to overcome barriers, particularly for vulnerable and underserved populations. Drop-out is especially common for potential donors who are Black and Hispanic, specifically those who face greater socioeconomic challenges to donating, and those who are not biologically related to their kidney patients. Drop- out from these groups of potential donors contributes to known disparities in LKDT. To reduce disparities in living donation and increase LKDT, we propose a multi-step study that includes: Aim 1a: Early identification of patients of all races/ethnicities at higher risk of dropping out from the donor evaluation process via secondary data analyses of multiple regional and national donor databases. A tailored risk prediction index (LD-DROP) will be developed and tailored to characterize the risk of drop-out among potential donors at the time of initial screening or contact, including subgroup analyses by race, ethnicity, and socioeconomic status; Aim 1b: Validation of the LD-DROP index by prospectively tracking a large, diverse group of individuals presenting for living kidney donor evaluation at two large and geographically diverse transplant centers; Aim 2: Individual interviews of potential living donors of various race, gender, and level of risk for drop- out will be conducted to explore their decision-making and readiness to donate, what vulnerabilities and barriers to donation they face, and what financial, psychosocial, peer mentoring, and educational interventions might be most helpful for overcoming these barriers; Aim 3: An intervention to address donation barriers will be piloted for potential donors at high drop-out risk through a national online peer support program and transplant center- based education programs for potential living donors. Project deliverables will be made freely available nationwide via partnerships with the Scientific Registry of Transplant Recipients Living Donor Collective initiative and the National Kidney Foundation.