Beckman Research Institute/City Of Hope

NIH Research Projects · FY 2024 · 2024-09

PROJECT SUMMARY Systemic lupus erythematosus (SLE) is a chronic autoimmune disease. 90% of patients with SLE are women between the ages of 15 and 44. SLE is characterized by a hyperactive, dysfunctional immune system, the presence of autoantibodies, and widespread inflammation in multiple organ systems, including the skin, joints, heart, lungs, kidneys, and brain. In SLE, dysregulated plasma cells, a terminally differentiated subset of B cells, produce autoantibodies which attack DNA, RNA, and self-proteins, including histones. SLE is currently managed with anti-inflammatory and immunosuppressive approaches including pan B-cell depleting regimens that alleviate symptoms and slow tissue damage. However, sustained remission of SLE remains a clinical challenge for several reasons. First, many therapies cause significant side effects as they globally affect the immune system. Second, there is a lack of targeted therapies that simultaneously eradicate autoantibody-producing short-lived plasmablasts and long-lived plasma cells (LLPCs), as well as their B-cell precursors. To eradicate all pathogenic B-cell subsets concurrently and specifically in patients with SLE, we propose to determine whether a patented, non-toxic splice-modulating oligomer (SMO), that prevents synthesis of the long isoform of the prolactin receptor (LFPRLR), is a viable approach. Increased circulating levels of the hormone, prolactin (PRL), are known to be associated with the exacerbation of symptoms of adult and pediatric SLE. Consistent with this, our preliminary findings suggest abnormally elevated expression of PRLRs in immune cells of female mice and patients with SLE. However, whether PRL and its receptors are causal in immunomodulation in SLE is less understood. Recently, we and others published that the LFPRLR specifically promotes the retention of potentially autoreactive B cells in a mouse model of SLE. We found that knockdown of the LFPRLR reduces the numbers of short-lived plasmablasts and LLPCs and their B-cell precursors in female SLE-prone mice. How the LFPRLR maintains pathogenic plasma cells and their precursor B cells in SLE remains to be delineated. This forms the focus of our current R21. We propose two Aims. In Aim 1, we will determine the effect of LFPRLR knockdown on pathogenic B-cell subsets including their terminally differentiated derivatives, plasma cells, in murine SLE. In Aim 2, we will determine the ability of LFPRLR knockdown to reduce pathologic plasma cells in human SLE. In both Aims, we will employ complimentary high-dimensional single-cell immune profiling approaches. We will correlate these findings with measurements of standard indicators of autoimmune disease pathology in SLE- prone mice and in samples from patients with SLE. Our studies will solidify knockdown of the LFPRLR as a novel, effective, non-toxic, and isoform-specific strategy to eradicate abnormal autoantibody-producing plasma cells in patients with SLE. In the long-term, these studies will propel extensive preclinical and clinical development of agents that knockdown the LFPRLR, such as those used here, to sustain disease remission in girls and women with SLE.

NIH Research Projects · FY 2024 · 2024-09

Revised Abstract / Summary Although bariatric surgery is the most effective treatment to promote sustained weight loss and metabolic improvement, they are invasive and can cause severe long-term adverse effects. There is a pressing need for safer and more affordable therapies. A greater understanding of the roles of conjugated bile acids in mediating the metabolic benefits of bariatric surgery may provide the background and direction towards developing new approaches to treat obesity and associated fatty liver diseases. Our preliminary data and the experiments proposed in this application propose to understand how conjugated bile acids regulate metabolism after bariatric surgery. After performing VSG in obese mice, we observe that: 1) VSG induced adipocyte lipolysis and enhanced fatty acid β-oxidation; VSG induces adipocyte browning, as well as induced macrophages conversion in the adipose tissues. 2) VSG leads to a strong increase in systemic levels of conjugated bile acids. 3) VSG induces expression of the hepatic enzymes for bile acid conjugation and alters the gut microbiota (GM), including bacteria with bile salt hydrolase (BSH) activity. This may contribute to the increase of conjugated bile acids and decrease of un-conjugated bile acid levels in the serum after VSG. 4) conjugated bile acids increased the expression levels of lipolysis markers in adipocytes in a bile acid receptor, sphingosine-1-phosphate receptor 2 (S1PR2)-dependent manner; and 5) VSG activated S1PR2 downstream signaling in the white adipose tissue (WAT) of mice. Based on these and other preliminary results, we hypothesize that VSG activates a gut–adipose axis which includes a conjugated bile acids mediated S1PR2 signaling pathway to induce fat loss. We propose two specific aims to test our hypothesis: 1) To determine the effects of VSG-altered conjugated bile acids and GM on adipose tissue lipolysis; 2) To identify the molecular mechanism by which conjugated bile acids promote adipocyte lipolysis. The proposed research will expand our understanding of the molecular mechanism by which conjugated bile acids improve metabolism after VSG. This work will also locate new therapeutic targets and/or probiotics for safe, non-surgical, cost-affordable treatment of metabolic diseases.

NIH Research Projects · FY 2024 · 2024-09

PROJECT SUMMARY Chimeric antigen receptor (CAR) T cell therapy is a novel form of cellular immunotherapy in which the antigen specificity of T cells is redirected using synthetic receptors. CD19-CAR T cells have achieved complete responses in up to 90% of patients with acute lymphoblastic leukemia. However, many malignancies do not possess a single, highly expressed tumor-associated antigen (TAA) such as CD19. Furthermore, CD19-negative relapses have been frequently encountered following CD19-CAR T cell therapy, suggesting that multi-antigen- targeting approaches will be needed to reduce relapse. Acute myeloid leukemia (AML) is the most common acute leukemia in adults and the majority of patients will die from their disease. We and others are evaluating CAR T cells to treat AML. However, AML exhibits heterogeneous expression of TAAs and many of these TAAs are expressed on hematopoietic progenitor cells (HPCs), increasing the risk of antigen-negative AML immune escape and bone marrow toxicity following AML-targeting CAR T cell therapy, respectively. Additionally, AML employs many active immune-suppressive strategies that may inhibit CAR T cells. To overcome these challenges, I have recently developed a novel viral co-transduction and sorting system to allow generation and purification of T cells with multiple transgenes such as multiple CARs, immune- stimulating molecules, safety switches, and secreted cytokines. Preliminary data suggest that multi-functional CAR T cells can be engineered to overcome antigen-negative leukemia escape and immune suppression mechanisms. I hypothesize that this novel sorting system can be used to engineer T cells to overcome AML TAA heterogeneity and immune suppressive strategies. Aim 1 will investigate CAR T cells simultaneously targeting a set of AML TAAs and predicted to avoid toxicity to HPCs. CAR T cells engineered to overcome AML- induced immune suppression will also be evaluated. In Aim 2 the goal is to target a set of TAAs expressed by both AML and HPCs as part of a pre-transplant CAR T cell immunotherapy strategy. During the award period, the candidate will conduct research at Memorial Sloan Kettering Cancer Center under the mentorship of Dr. Marcel van den Brink and an Advisory Committee. He will obtain the critical skills he needs to become a tenure-track physician-scientist running his own academic laboratory developing synthetic biology approaches to improve cellular therapies and successfully competing for independent NIH funding. He will cultivate a detailed and comprehensive skill set for syngeneic, xenograft, and humanized mouse models of cellular immunotherapy, build upon an existing knowledge base of molecular construct design and cellular gene modification by mastering multiplexed CRISPR/Cas9 gene disruptions and site-specific gene integration, and develop proficiency in genomic analysis to better define T cell activation and exhaustion states and to identify novel targets for gene therapy.

NIH Research Projects · FY 2024 · 2024-09

Diabetes is now a global epidemic. More than 95% of diabetes is type 2 diabetes (T2D), a chronic disease that can cause serious complications, including heart attack, stroke, kidney failure, blindness, and lower limb amputation. Progressive deterioration in pancreatic islet β-cell function is a hallmark of T2D, however, the mechanism of β-cell loss in T2D remains elusive. Βeta cells increase insulin secretion in response to hyperglycemia. If insulin production surpasses the capacity of endoplasmic reticulum (ER) to make functional proteins, misfolded proteins will buildup, a process called the ER stress. Metabolic stress due to obesity, aging, and other lifestyle changes disrupts energy homeostasis, which triggers ER stress in β cells leading to the activation of UPRs. Chronic ER stress under metabolic stress, which is characterized by insulin resistance, turns adaptive UPR to maladaptive UPR, which is believed a driving force for β-cell mass loss in T2D. Our long-term goal is to identify the tilting point of UPR under metabolic stress and to explore its translational potential for therapeutic interventions. Xbp1s, a UPR transducer activated by ER stress, is crucial for β-cell function and survival. Consistent with the requirement for Xbp1s to protect β-cells from dysfunction under metabolic stress, Xbp1s is found elevated in β-cells in preT2D donors and prediabetic models. However, the overexpression of Xbp1s, when studied in vitro showed conflicting results: one showing Xbp1s promotes β-cell apoptosis, and the other showing Xbp1s promotes β-cell proliferation. Despite the controversy about Xbp1s overexpression in positive or negative regulation of β-cell survival, it is clear that the action of Xbp1s in β-cells is complicated and it is insufficient to elucidate the impact of Xbp1s on β-cell integrity under metabolic stress by in vitro studies solely. Meanwhile, the Xbp1 loss-of-function mouse models used in the field are deficient in both spliced and unspliced form of Xbp1. Therefore, a knowledge gap remains for the pathological role of Xbp1s in β-cell function and survival under metabolic stress. Using a novel mouse model that allows inducible ablation of spliced Xbp1 selectively in β-cells, we found a striking impact of Xbp1s on β-cell function and survival. Elucidation of the role of Xbp1s as an integrative player in adaptive remodeling of β-cells under metabolic stress will advance our understanding of the progressive deterioration in pancreatic islet β-cells in T2D and pave a way for novel, more effective therapeutic design for preventing β-cell loss in T2D progression.

NIH Research Projects · FY 2025 · 2024-09

This R50 application will support the activities of Dr. Alex Herrera as a Research Specialist (Clinician Scientist) promoting and supporting National Cancer Institute (NCI)-sponsored clinical trials network (CTN) research, both nationally and within the City of Hope Comprehensive Cancer Center (COHCCC). Dr. Herrera is a clinical and translational lymphoma investigator who leads a clinical trial program focused on developing novel therapies for the treatment of Hodgkin and non-Hodgkin lymphomas. Within the COHCCC, Dr. Herrera is the Associate Medical Director of the Clinical Trials Office and Briskin Center for Clinical Research, Chief of the Lymphoma Division within the Department of Hematology, and Co-Leader of the Lymphoma Disease Team. He is a national leader in the NCI clinical trial networks as a member of the SWOG Lymphoma Committee Working Group and the NCI Lymphoma Steering Committee and serves as the study chair/principal investigator or translational medicine chair of several clinical trials in the National Clinical Trials Network (NCTN) and Experimental Therapeutics Clinical Trials Network (ETCTN). Notably he is the study chair of the groundbreaking SWOG S1826 trial, the largest Hodgkin lymphoma trial conducted in the NCTN and the first NCTN collaboration between the Hodgkin adult and pediatric cooperative groups, whose outcome is poised to change the standard of care. He is also a national leader in the mentorship of junior clinical faculty, with a focus on reducing barriers to clinical trial enrollment. Through this R50 award, Dr. Herrera will spearhead efforts to increase COHCCC NCI-sponsored CTN research activities by (1) building specialized clinical research infrastructure to manage NCI-CTN trials and grow a multidisciplinary institutional NCTN leadership team; (2) lead expansion and integration of the COHCCC clinical research infrastructure across the regional/national enterprise to increase NCTN/ETCTN trial enrollment; (3) reduce barriers to clinical trial access for patients from COH network sites, in order to enhance enrollment to NCTN/ETCTN trials. This application will also support expansion of Dr. Herrera’s efforts toward setting the national NCI-sponsored clinical trial network’s research agenda for lymphoma, serving as the SWOG Lymphoma adolescent young adult (AYA) liaison, mentoring junior investigators designing NCTN and ETCTN trials, and conducting practice-changing lymphoma clinical research in the NCTN/ETCTN. This R50 award will allow Dr. Herrera to unify his research agenda on a theme of driving and expanding internal

NIH Research Projects · FY 2024 · 2024-09

7.0 Project Summary/Abstract Up to 70% of patients with advanced cancer experience significant levels of FOP. FOP is the most common psychosocial concern among GYN and other cancer survivors. Current FOP interventions are time, resource- intensive, and administered by psychologists, limiting access and sustainability. Further, most have focused on early-stage cancer. Advanced cancer patients are severely understudied. The “Day by Day” intervention was adapted from “Conquer Fear,” which has shown effectiveness in reducing FOP in cancer survivors treated with curative intent. The proposed intervention is the first to use a blended e-Health intervention to address FOP in patients with advanced cancer. Our preliminary pilot studies established feasibility of a 1:1 nurse-led, videoconferencing intervention in patients with stage III or IV gynecologic or lung. Based on our experience, we adapted the intervention to a group and online format. We developed didactic, patient, and additional mindfulness practice videos hosted on the website. We tailored the intervention to patients with advanced cancer, with a focus on Acceptance and Commitment therapy because it addresses common existential concerns and values- driven goals and actions. We will conduct a randomized pilot study to test the feasibility and efficacy of this blended e-Health intervention in patients with stage 3 or 4 gynecologic cancer. The intervention consists of 2 group videoconferencing sessions, 3 web-based self-study modules, and a brief check-in call with the facilitator (nurse or social worker). The intervention modules focus on the cancer experience, values and goal setting, unhelpful beliefs about worry, skills practices, managing triggers, and important conversations. The Enhanced Usual Care group will be delivered in the same format and consists of education only. Sessions focus on the stress response, clinical trial participation, identifying reliable information online, and healthy lifestyle education. Feasibility measures include enrollment rate, attrition, and attendance. Participants complete questionnaires assessing FOP, distress, anxiety, metacognitions, and mindfulness at baseline, 6, and 10 weeks and an evaluation survey at T2 or drop-out. Given the increasing number of survivors needing help with FOP and the lack of trained clinicians, a multidisciplinary approach is needed to meet the demand. Implementing brief nurse and social worker guided FOP interventions may be scalable and address barriers to accessing treatment. Studies incorporating novel delivery models such as blended models will facilitate access and sustainability. Further, including a group component can foster support and may be less resource intensive. This blended e- Health intervention may offer high applicability to underserved communities where mental health providers are scarce. Use of an e-health platform with group support may facilitate access, engagement, and sustainability.

NIH Research Projects · FY 2024 · 2024-08

PROJECT SUMMARY/ABSTRACT The Society for Integrative Oncology (SIO) 21st International Conference, "Full Circle Translational Integrative Oncology: Bedside to Bench and Back - The transformative power of clinical observation to ignite scientific discovery and optimize patient outcomes,” aims to encourage scientific and clinical collaboration across global and inter-professional boundaries to advance knowledge, therapies, and care delivery that will ultimately improve the quality of life and clinical outcomes for individuals and families living with cancer around the world. This year’s conference – SIO2024 – will focus on disseminating important and practical findings from integrative oncology research that have the potential to transform cancer care. This three-day meeting will be held in Orange County, CA from October 25 to October 27, 2024, and hosted by City of Hope (COH). SIO2024 will feature three keynote lectures that highlight the conference theme: 1) an overview of the newest immune-oncology approaches with a focus on CAR T-cell therapy; 2) natural product drug discovery for anticancer therapy, and 3) the intersection of spirituality and integrative oncology. The conference will include three moderated plenary panels that focus on the current state of the science for: 1) natural products and immunotherapy; 2) nutrition and metabolomics; and 3) applying artificial intelligence/machine learning to integrative oncology. The conference will also include seven educational sessions and workshops, five concurrent oral abstract sessions, three lunch session focusing on nutrition, mind-body medicine, and diversity & equity in integrative oncology, as well as patient advocacy and trainee/new investigator tracks We will solicit at least 75 oral and up to 150 poster presentations of original research from members of the national and international research community. The conference’s specific aims are to: 1) create an international forum in which of researchers, clinicians, trainees, and patient advocates with an emphasis on diversity including women and minorities can learn about and discuss the current evidence for the interplay of integrative oncology research and clinical application to enhance their understanding of the advantages and challenges of this field; 2) foster critical dialogue and collaboration opportunities among researchers, health professionals, and patient advocates to strengthen the translational potential and clinical relevance of future research in integrative oncology; and 3) support the training of early investigators and patient advocates to interpret, disseminate, and develop evidence-based integrative oncology research. Ultimately, we hope that SIO2024 will improve the outcomes of cancer care in patient communities around the world.

NIH Research Projects · FY 2024 · 2024-07

PROJECT SUMMARY The immediate objective of our proposal is to increase immune density within solid malignancies to rectify defects in anti-tumor immunity and maximize response rates to immune checkpoint blockade (ICB) therapy. An important hallmark of cancer is the ability to suppress or exclude innate and adaptive antitumor immune subsets, resulting in a tumor microenvironment less conducive to ICB treatment. Studies using combinatorial strategies have concluded that increased CD8 T cell density during immune checkpoint inhibition significantly enhances anti-tumor efficacy in non-immunogenic “cold” tumors. To increase CD8 T cell infiltration, we have developed a tumor-targeting bacterial platform engineered to secrete chemokines and cytokines, which we refer to as biological beacons or “bio-beacons,” and are meant to recruit and activate select subsets of immune cells. We have completed proof-of-principle experiments to show that bio-beacons, composed of tumor- colonizing E.coli vectors secreting murine C-X-C motif chemokine ligand 9 (CXCL9) and interleukin-15 (IL-15), mediate migration and expansion of murine lymphocytes. While cytotoxic CD8 T cells are thought to mediate the majority of tumor cell killing during immune checkpoint inhibition, it is becoming increasingly clear that overall efficacy of the anti-tumor response depends on a broader spectrum of cell types present within the tumor microenvironment that provide support, such as CD4 T cells, dendritic cells (DCs) and natural killer (NK) cells. Therefore, we propose to generate additional bio-beacons secreting murine CCL3 and CCL4, which are anticipated to recruit a broader spectrum of immune cells including CD4, NK, DCs, and monocytes. We hypothesize that bio-beacons will increase sensitivity of solid tumors to immunotherapy, specifically ICB treatment, by increasing intratumoral density of tumor-reactive immune subsets. We propose to elucidate changes in intratumoral immune composition following bio-beacon treatment and their potential synergy with immune checkpoint blockade therapy as a prelude for clinical development and translation. A series of studies under two Specific Aims will be conducted to: i) develop and evaluate novel bio-beacons intended to recruit supportive immune subsets; ii) determine the changes in intratumoral immune composition mediated by single or combinations of bio-beacons in various solid tumor models; and iii) determine bio-beacon combinations that significantly enhance efficacy of ICB therapy in tumor-bearing mice. These studies will delineate the intratumoral frequency and functional phenotype of immune subsets following bio-beacon administration, which can be used to uncover potential mechanisms contributing to any observed improvements in ICB therapy. If successful, we predict that bio-beacons can be used to increase the frequency of tumor-reactive immune subsets in a variety of solid tumor types and, thus, increase sensitivity to ICB treatment. In future endeavors, we imagine leveraging the ability of bio-beacons to recruit and expand T lymphocytes as a strategy to restrict the cytolytic activity of cell-based therapies, such as chimeric antigen receptor (CAR) T cell therapy, to solid tumors.

NIH Research Projects · FY 2025 · 2024-07

Project Summary More than 10% of the global adult population currently lives with diabetes mellitus (DM) and are experiencing or at higher risk for various diabetic vasculopathies such as peripheral artery disease (PAD). Despite available treatment, many patients with DM continue to develop diabetic vasculopathy, highlighting the urgent need for more effective therapies. Mounting research has shown that the dysfunction of endothelial cells (ECs), known as endotheliopathy, plays a crucial role in the onset of DM and its vasculopathies. However, there is still a gap in knowledge regarding the epigenetic regulators that drive endotheliopathy in DM and cause persistent vascular damage, and whether these regulators can be targeted for treatment. Synergizing efforts from our funded studies on long non-coding RNA (lncRNA)-regulated EC function (R01HL145170) and diabetic epigenetic memory (R01HL106089), my group has uncovered important roles of non-coding RNAs, including microRNAs and lncRNAs, in endotheliopathy, vascular inflammation, and impaired vascular regeneration in DM and diabetic PAD. Furthermore, we identified chromatin-associated lncRNAs as key mediators of metabolic memory, which may explain the persistent vascular damage observed in patients with DM, even after restoration of normoglycemia. We have also broken new ground to unravel the role of the N6-methyladenosine (m6A)-modified epitranscriptome in DM-induced endotheliopathy and to harness nanoparticles and modified RNA to develop therapeutics that target the dysfunctional EC epigenome. In this R35 application, our overall vision is that elucidating the RNA-mediated epigenetic and epitranscriptional mechanisms underlying DM-associated endotheliopathy in depth will enable us to develop novel therapeutics to ameliorate diabetic vasculopathy. Our proposed studies will bring together expertise in RNA, chromatin, and endothelial biology; gene therapy; and nanomedicine to address this pressing medical issue. We will achieve our goals through three independent yet inter-connected projects to: 1) identify the roles and mechanisms of action of novel lncRNA regulators in driving endotheliopathy and diabetic vasculopathy; 2) determine the role of m6A RNA modification in diabetic endotheliopathy and vasculopathy; and 3) evaluate the efficacy of innovative nanoparticle-mediated epigenetic therapeutics in ameliorating diabetic vasculopathy, with a focus on PAD, a painful disease with limited treatment options. The unique R35 Emerging Investigator Award mechanism will allow me to further my group’s research on RNA-mediated endotheliopathy by testing new hypotheses using innovative technologies. Aligned with the NHLBI mission, our proposed studies will advance our understanding of diabetic vasculopathy and have the potential to facilitate the development of much-needed new therapies for patients with diabetic vasculopathy.

NIH Research Projects · FY 2025 · 2024-07

PROJECT SUMMARY Prostate cancer (PCa) is the most common cancer among men. Radiation therapy is an integral part of the standard of care for the treatment of PCa. Local recurrences of PCa after radiation therapy usually originate from the primary tumor site. A dose escalation to the clinically significant tumor (csT) foci in intermediate- and high-risk patients is reported to result in significantly improved biochemical disease-free survival (bDFS) without increasing radiation toxicity. Contouring of the intraprostatic tumor is currently based on multiparametric (mp-) magnetic resonance imaging (MRI) and the interpretation of the MRI results are based on Prostate Imaging Reporting & Data System (PI-RADS). However, relatively large variability in performance of mp-MRI, including that of PI-RADS, continue to pose barriers to identify clinically significant prostate tumor as treatment target and improve patient outcome. In this project, we propose to address the issues in both the MRI technique and scoring system aspects and develop a tool that can automatically delineate intraprostatic csT and be easily transferred to clinical use. To achieve this goal, in Aim 1, we will build a quantitative MRI (QMRI) toolset including magnetic resonance fingerprinting (MRF), arterial spin labeling (ASL), and diffusion- weighted MRI (DW-MRI). This toolset provides 3-dimensional high-resolution quantitative maps directly associated with tumor physiology and pathology features. In Aim 2, based on the QMRI output, we will develop an automated PCa tumor probability model and segmentation method that can automatically differentiate areas with clinically significant prostate cancer from other prostatic tissues. Aim 3 will be a validation study. We will validate the performance of our newly developed method against biopsy results and compare it with the performance of physician's contour of csT foci based on mp-MRI and PI-RADS as the current standard of care (SOC). We expect that the results of this project will provide a completely new set of QMRI tools and tumor segmentation method that improves current SOC mp-MRI and PI-RADS. This will allow us to accurately identify areas with clinically significant PCa in the intact prostate for boost irradiation, and achieve maximal local control in prostate cancer radiation therapy without increasing radiation toxicity.

NIH Research Projects · FY 2026 · 2024-07

Project Summary Heart failure is a leading cause of morbidity and mortality worldwide. Hypertension is one of the most important risk factors of heart failure. Despite paramount interests and urgent clinical needs, our understanding of the mechanisms of heart failure development remains limited. To accommodate the elevated demand of cardiac contractility under high blood pressure, the heart mounts an acute reaction through cardiomyocyte hypertrophic growth. This once adaptive response may decompensate and progress into heart failure. The long- term goal of this project is to identify novel mechanisms of the pathological transition from adaptive cardiac hypertrophy to heart failure and explore therapeutic interventions. With an unbiased phosphoproteomic analysis, Rps6kb1 was identified as one of the most prominent kinases in cardiomyocyte hypertrophic growth. Rps6kb1 is a classical downstream target of mTOR, however, its role in hypertensive heart disease remains incompletely understood. Moreover, Rps6kb1, with its complex activation mechanism, may integrate signals from multiple upstream pathways, above and beyond the mTOR signaling. Preliminary studies showed that Rps6kb1 deletion in the heart suppresses adaptive cardiac hypertrophic growth and consequently, cardiomyopathy and heart failure are accelerated under pressure overload. Based on these findings, a central hypothesis is formulated that Rps6kb1 is an essential player in adaptive hypertrophic growth by integrating signals from multiple upstream pathways. Additional preliminary data showed that Rps6kb1 is directly phosphorylated by ERK, independent of the mTOR signaling. A novel threonine site of phosphorylation by ERK was later identified by mass spectrometry. More importantly, ERK-mediated phosphorylation of Rps6kb1 is required for full Rps6kb1 activation. Further pilot tests demonstrated that Rps6kb1 is subjected to another post-translational modification, K63 polyubiquitination. Lysine 85 was identified as the critical site for this modification through multiple mutagenesis assays, and Lys85Ala mutation strongly diminished both ubiquitination and activation of Rps6kb1. In this grant application, the role of ERK-mediated phosphorylation of Rps6kb1 will be further characterized, and its implications in adaptive hypertrophic growth and heart failure will be illustrated using both gain- and loss-of-function mouse models. In addition, the relevance and significance of Lys85 ubiquitination in hypertensive heart disease will be delineated by identification of the responsible E3 ligase and evaluation of the novel Lys85Ala knock-in mouse model. Moreover, the interplay between this novel phosphorylation and this new ubiquitination in cardiac hypertrophy will be interrogated. Primary cardiomyocyte culture will be employed to complement in vivo animal models at the mechanistic level. Elucidation of the role of Rps6kb1 as an integrative player in pathological cardiac remodeling will advance our understanding of hypertensive heart disease and heart failure and pave a way for novel, more effective therapeutic design.

NIH Research Projects · FY 2024 · 2024-07

The goal of this application is to obtain support for the American Chemical Society Division of Chemical Toxicology (TOXI) at the 268th National Meeting, which will be held August 18-22, 2024, in Denver, CO. The mission of the Division of Chemical Toxicology is to improve human health and public welfare by promoting the understanding of chemical mechanisms that govern disease processes and the toxicity of drugs, environmental agents, and endogenous chemicals. Our goals include providing a diverse forum for communicating research in the field of chemical toxicology, encouraging research in chemical mechanisms of toxicity, and facilitating connections between academia, industry, and policy in scientific areas of mutual interest. We are also committed to developing the leadership and professional development skills of scientists from all levels and backgrounds. Thus, the requested funds will be used to support travel awards and expenses for graduate students, postdoctoral scholars, and junior faculty. The overall TOXI program is organized around the theme of “Toxicological Predictions, Markers, and Outcomes Affecting Human Health” and complements the National Meeting theme of Elevating Chemistry. The program includes five thematic symposia, which will feature invited oral presentations by a diverse group of established and emerging investigators with a range of perspectives on each topic. The five thematic symposia are: (1) “Application of Augmented Artificial Intelligence in Toxicology Metabolism Prediction”, co-promoted by the Division of Chemical Information (CINF) and Medicinal Chemistry (MEDI), will explore the role of augmented AI in predicting toxic metabolites; (2) “The Role of Investigative Toxicology in Drug Discovery & Development”, co-promoted with MEDI, will provide insight into the role of investigative toxicology in a drug discovery setting; (3) “Revealing Toxicological Mechanisms of Small Molecules using Chemical Biology”, co-promoted with the Divisions of Biology (BIOL) and Organic Chemistry (ORGN), focusing on the innovative and creative use of chemical biology and functional omics strategies that reveal the mechanisms by which small molecules affect toxicity or alter biological activity; (4) “Advances in Forensic Toxicology”, co-promoted by the Society for Forensic Toxicology, focusing on the application of toxicological methods, studies, and knowledge to situations with medico-legal consequences; (5) “Women in Toxicology”, co- promoted with Chemical Research in Toxicology, will highlight the work of women in toxicology and explore how they are making strides to improve sustainable living throughout the world. The scientific program is clearly aligned with the mission of NIEHS.

NIH Research Projects · FY 2025 · 2024-06

The Beckman Research Institute of City of Hope (COH) is a world-class cancer research institution and cancer hospital with a mission of turning science into practical benefit through exquisite care, innovative research, and vital education focused on eliminating cancer and diabetes. Increases in cancer incidence and rising treatment costs highlight a need to advance cancer research through the growth of a well-trained workforce. We propose an institution-wide, 10-week research experience for undergraduates focused on cancer, aging, and metabolism, called the Cancer Aging and Metabolism research experience Program (CAMP). The CAMP R25 is committed to training future scientific leaders who have the skills to develop novel technologies and translate them into the clinic. CAMP will provide a focused, mentored research experience by matching undergraduate trainees (CAMPers) with mentor-researchers from three thematic areas: Cancer Aging, Cancer Metabolism, and Clinical Research. Each summer, CAMP will enable 25 CAMPers, selected via local and nationwide searches, to participate in research that spans the arc of cancer research, from basic to bedside. CAMPers will select from among 30 experienced COH Mentors who have a track record of mentoring students with varied interests and backgrounds. Principles of responsible conduct of research and rigor and reproducibility will be taught by faculty. Hands-on laboratory research and auxiliary activities will be enhanced by journal clubs, distinguished speaker seminars, and weekly CAMPer research presentations, as well as an end-of-summer poster session. To expand our educational impact and enhance the pipeline of future researchers, CAMP will also pilot a school-year extension program, designed to support promising local undergraduates with sustained mentorship and research engagement beyond the summer. At the end of each summer, five CAMPers will be competitively selected based on performance and interest to participate in a school-year extension, continuing research under their mentors for approximately nine months. This year-round engagement, fully supported by the R25, is designed to sustain mentorship, enhance skill development, and strengthen retention of promising students in cancer research pathways. The administrative structure of CAMP includes a highly experienced Program Director, Associate Director, and a team of faculty members serving on specialized committees in Recruitment and Admissions, Curriculum Development, Evaluation, and Mentor Training. Oversight will be provided by Internal and External Advisory Committees to ensure quality, equity, and accountability throughout the program. CAMP is grounded in City of Hope’s long-standing commitment to education and its successful track record in hosting summer programs that have launched the careers of countless trainees. This R25 proposal is uniquely positioned to fill a critical federal funding gap in undergraduate cancer research training and will help shape the next generation of researchers dedicated to improving outcomes for patients with cancer.

NIH Research Projects · FY 2025 · 2024-06

PROJECT SUMMARY: Acute myeloid leukemia (AML) is a devastating hematopoietic malignancy characterized by clonal expansion of leukemia stem cells (LSCs). Despite advancements in chemotherapy regimens and targeted therapies, most AML patients remain incurable. Thus, novel therapies targeting key leukemogenic pathways are needed. RNA modifications are essential modulators of post-transcriptional gene regulation, and their dysregulation emerges as a contributor to AML. Through an integrated analysis of genome-wide CRISPR/Cas9 screen data, we discovered that AML cells preferentially depend on METTL1, an RNA methyltransferase that mainly catalyzes 7-methylguanosine (m7G) modification on transfer RNAs (tRNAs). METTL1 is highly expressed in LSCs and AML specimens, and its high expression is associated with poor clinical outcomes. METTL1 depletion significantly suppresses AML growth, eradicates LSCs in vitro, and attenuates AML progression in an AML patient-derived xenograft (PDX) model. Importantly, those effects are largely dependent on METTL1’s m7G methyltransferase activity, offering a unique opportunity for AML therapy through the development of small- molecule inhibitors targeting its enzymatic activity. Via a high-throughput screen, we have discovered a selective and potent METTL1 inhibitor with promising anti-AML activity. Moreover, METTL1 loss significantly reduces the m7G abundance on tRNAPheGAA and its overall expression. This, in turn, leads to translation suppression of transcripts that heavily rely on tRNAPheGAA-related codons, such as tyrosine-protein kinase HCK. The decreased expression of HCK due to METTL1 depletion could further disrupt C-X-C chemokine receptor 4 (CXCR4) signaling, which is essential for LSC homeostasis. Despite these insights, we do not yet understand the exact mechanisms by which METTL1 loss results in AML suppression and LSC eradication. We hypothesize that METTL1 functions as an m7G methyltransferase to drive AML development and sustain LSC frequency, making it a potential ‘druggable’ target for treating high-risk AML. These hypotheses will be addressed in three Specific Aims. Aim 1 will further consolidate the importance of METTL1 function in AML using mouse models. In this Aim, we will utilize a number of AML models with different genetic backgrounds to rigorously determine the roles of METTL1 in AML initiation and progression. Aim 2 will understand the role of METTL1/tRNAPheGAA/HCK signaling in LSC homeostasis. This Aim will delineate the molecular mechanism through which the METTL1/m7G axis facilitates LSC homing and self-renewal ability. Aim 3 will evaluate the therapeutic potential of pharmacologically targeting METTL1 to treat AMLs.

NIH Research Projects · FY 2026 · 2024-05

Non-small cell lung cancer (NSCLC) remains the leading cause of cancer-related death in the United States. Certain populations, such as non-Hispanic Black/African American (AA) individuals, are diagnosed at younger ages and experience less favorable outcomes. AA communities are also exposed to higher levels of air pollution (AP) such as fine particulate matter (PM2.5) and nitrogen oxides (NOx). This study will investigate how long-term exposure to AP (PM2.5 and NOx) and neighborhood deprivation as measured by several place-based indicators [e.g. area deprivation index (ADI), social vulnerability index (SVI), isolation index, etc]—contribute to mutational patterns and disease progression in early-stage NSCLC in a high-risk population. We hypothesize that cumulative exposure to AP and neighborhood deprivation influences the timing and type of somatic mutations, ultimately affecting recurrence risk. We will conduct a study of 300 AA patients with stage I–II NSCLC from California, Georgia, and Detroit, integrating whole-genome sequencing (WGS) of tumor tissue with individualized environmental and geospatial exposure data. The aims of the study are to: (1) identify relationships between exposure history and mutational signatures (order and types of mutations); (2) assess how AP and neighborhood deprivation influence early recurrence (within 2 years post-surgery); and (3) compare the tumor genomics of never-smoking AA patients to the national Sherlock-Lung study, to evaluate the modifying effect of AP and neighborhood deprivation. This research will generate insights into how external exposures influence NSCLC biology and outcomes and may inform risk stratification, early detection and prevention intervention strategies tailored to individuals experiencing elevated environmental exposures to AP and neighborhood deprivation.

NIH Research Projects · FY 2026 · 2024-04

PROJECT SUMMARY Bisphenol S (BPS), bisphenol F (BPF), and diisononylphthalate (DINP) are increasingly replacing the endocrine disrupting chemicals bisphenol A (BPA) and di-2-ethylhexylphthalate (DEHP), respectively. However, BPS, BPF, and DINP have not undergone safety testing. Perinatal exposures (in utero and during lactation) to BPA and DEHP are associated with the development of non-alcoholic fatty liver disease (NAFLD). NAFLD, a common disease in children, is being diagnosed at increasingly younger ages and its prevalence is increasing in late adolescence. While BPA and DEHP have been extensively studied in this context, BPS/BPF and DINP have not. BPS exposure promoted NAFLD progression in zebrafish, and BPF serum levels are higher in NAFLD patients than in controls. BPS and BPF levels were associated with increased prevalence of obesity in children, a risk factor for NAFLD. DINP caused lipidomic disruption in neonatal mice, while phthalate exposure caused hepatic steatosis in adult mice. Thus, like BPA and DEHP, early-life BPS/BPF and DINP exposure may promote NAFLD development/progression. To date, the mechanisms by which bisphenols and phthalates, particularly as a mixture, trigger these long-term metabolic consequences are elusive. Here, we will test the hypothesis that early life exposure to a bisphenol/phthalate mixture activates RAGE signaling pathways, triggers a trained immune response in peripheral monocytes, and reprograms the epigenome/transcriptome in liver and in peripheral monocytes, thereby priming offspring for exaggerated metabolic dysfunction upon exposure to Western diet. This ViCTER consortium will address this novel hypothesis through integrated studies in mice and in human subjects.

NIH Research Projects · FY 2025 · 2024-04

Allogeneic hematopoietic cell transplantation (Allo-HCT) is a curative therapy for patients with hematological malignancies (i.e. leukemia and lymphoma), through graft versus leukemia/lymphoma (GVL) effect mediated by alloreactive donor T cells. However, graft-versus-host disease (GVHD) caused by the same alloreactive T cells remains a major obstacle. Chronic GVHD (cGVHD), a systemic autoimmune-like syndrome, remains a major cause of morbidity and mortality in long-term survivors of Allo-HCT. During the past decade, we have used murine and humanized murine models of cGVHD and biospecimens from cGVHD patients to demonstrate that CD4+ T cells and B cells and their interactions in GVHD target tissues such as the liver, lung and skin play a critical role in the pathogenesis and persistence of the disease. The premise of this new proposal is that deeper mechanistic understanding of the interactions between non-circulating tissue- resident CD4+ memory T (Trm) and tissue-resident B (Brm) cells in GVHD target tissues will identify new therapeutic targets for prevention or treatment of cGVHD. In Aim 1, we will determine whether CD4+ Trm cells initiate formation of tertiary lymphoid structures (TLS) in GVHD target tissues, in which Ly108+ stem cell-like progenitors maintain the pool of Trm and Brm to perpetuate cGVHD pathogenesis. We will also develop novel cell-penetrating PS-Cas9-gRNA to target STAT3 in donor T cells to prevent development of pathogenic CD4+ Trm and cGVHD while preserving GVL activity, because our recent studies indicate that STAT3 is required for Trm development. In Aim 2, we will determine whether B cell helper PD-1hiPSGL1loCD4+ Trh cells interact with T-bet+ Brm cells to produce autoantibodies that augment systemic tissue damage. In Aim 3, we will determine whether non-B cell helper PSGL1hiCD4+ Trm cells interact with dendritic cells, macrophages, and fibroblasts in cGVHD target tissues to mediate damage and fibrosis in an IL-33/ST2 pathway-dependent manner. These studies have high biological and translational significance and may lead to a paradigm shift in our understanding of cGVHD pathogenesis and to development of novel approaches for preventing and treating cGVHD.

NIH Research Projects · FY 2026 · 2024-03

Allogeneic hematopoietic cell transplantation (allo-HCT) is a therapy with curative intent for a variety of malignant and non-malignant diseases. One of the major complications after allo-HCT is graft-versus-host disease (GVHD), especially of the gastrointestinal tract. Our group was one of the first to explore the role of the intestinal microbiome in allo-HCT in patients and preclinical models by combining next-generation sequencing with immunological, microbial and metabolomic approaches. We found relationships between the intestinal microbiome and GVHD, relapse, infections, engraftment, and immune reconstitution. The clinical and preclinical studies we performed in the previous funding period resulted in 19 manuscripts (including NEJM, Nature, Science, and Nature Med). We observed in large multi-center studies that higher diversity of intestinal microbiota was associated with better overall survival, reconstitution of CD4, mucosal-associated invariant and V2 T cells. Expansion of particular Enterococci was not only associated with GVHD in patients but exacerbated disease in mice. We showed that loss of Clostridia was associated with the onset of acute GVHD, whereas lower serum levels of butyrate were associated with an increased risk for chronic GVHD. In addition to their role in digestive physiology, bile acids (BAs) exert wide-ranging biological effects, including antimicrobial activity, intestinal epithelial homeostasis, and modulation of immunity. Primary BAs (PBA) are synthesized in the liver and transformed by the intestinal microbiota into a diverse pool of secondary bile acids (SBAs) with unique biochemical properties. BAs can bind to Farnesoid X Receptor (FXR), a nuclear receptor that is a critical regulator of BA homeostasis, inflammation, and GI barrier function. Our preliminary preclinical studies demonstrate that allo-HCT recipients with GVHD have: a) a decrease in unconjugated BAs and SBAs in cecal contents and plasma, b) a reduced abundance of bile salt hydrolase genes (critical for SBA transformation) in the intestinal microbiome, and c) decreased BA synthesis in the liver. In addition, we found that a) most BAs can counteract FXR activation by the PBA chenodeoxycholic acid and b) donor T cells deficient of FXR induce less GVHD. Our preliminary clinical studies corroborate the experimental results in that fecal α-diversity is associated with increased abundance of SBAs, which are in turn correlated to less acute GVHD and better overall survival. Based on studies demonstrating that BAs can inhibit Th17 polarization and enhance Treg generation and our preliminary data in mouse models and allo-HCT patients, we hypothesize that BA metabolism is an important modulator of alloreactive donor T cells and GVHD in allo-HCT recipients. All our research is performed through perpetual dialogue between investigation in mice and humans. Therefore, we propose to study the role of bile acids in GVHD after allo-HCT in preclinical models (Aim 1) and allo-HCT patients (Aim 2) and to develop strategies to improve GVHD by targeting BA metabolism (Aim 3). Our major goal is to ameliorate outcomes after allo-HCT by developing BA-based therapies.

NIH Research Projects · FY 2025 · 2024-03

Project Summary/Abstract Recent breakthroughs in tumor immunology led to the development of cancer immunotherapies which have been successfully applied to a range of human cancers. Among the immune cellularity in the tumor microenvironment, tumor-associated macrophages (TAMs) are often the most abundant immune cells and play critical roles in tumor immunosurveillance. Recent progress demonstrates that cancer cells upregulate anti-phagocytic “don’t eat me” signals as self-protective mechanisms against macrophage surveillance. When these signals are blocked, macrophages are capable of eliminating various malignant hematopoietic and solid tumor cells through cellular phagocytosis. Inducing cancer cell phagocytosis demonstrates significant anticancer effects and therapeutic potential in preclinical models and recent clinical trials. Tyrosine phosphorylation and dephosphorylation, mediated by protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs), is an essential mechanism underlying signal transduction involved in multiple critical biological functions including the regulation of immune responses. PTPs such as SHP1/SHP2 have been shown to be involved in the inhibitory signaling in macrophages and T cells for deactivating immune activities. Despite its importance, however, the underlying molecular mechanisms of macrophage-mediated cancer cell phagocytosis are still unclear, and the critical components mediating this process in addition to cell surface receptors have mainly remained unidentified. In this proposal, we are focusing on the functions and roles of PTPs in macrophage immunosurveillance. Through a CRISPR-based loss-of-function screening in macrophages, we have identified non-receptor PTP candidates that negatively regulate phagocytosis. When their expression is suppressed, the phagocytic ability of macrophages toward cancer cells is significantly enhanced. Our central hypothesis is that such PTPs function as inhibitory regulators of macrophage-mediated cancer cell phagocytosis, and thus their inhibition can promote the efficacy of macrophage phagocytosis-based cancer immunotherapy. In this proposal, we will first investigate to what extent the deficiency of inhibitory PTPs in macrophages promotes the efficacy of cancer cell phagocytosis with multiple preclinical models. Second, we will dissect the molecular mechanism of how deficiency of PTPs in macrophages augments cancer cell phagocytosis. Finally, we will examine the anticancer efficacy of suppressing PTPs in macrophages with in vivo cancer models. The completion of this study will identify and establish the roles of new regulators of cancer cell phagocytosis and inspire the development of new therapeutic strategies to enhance the efficacy of macrophage-based cancer immunotherapy.

NIH Research Projects · FY 2026 · 2024-02

Project Summary The goal of this project is to determine the molecular mechanisms by which pre-mRNA splicing is regulated by protein arginine methylation, one of the most abundant post-translational modifications on the RNA binding proteins, and to define the impact of this regulation in synapse development and brain function. The human genome encodes nine protein arginine methyltransferases (PRMT1–9), which catalyze three types of arginine methylation: monomethylation (MMA), asymmetric dimethylation (ADMA), and symmetric dimethylation (SDMA). A few years ago, we characterized the newest member of the PRMT family–PRMT9 and identified the splicing factor SF3B2 as the primary substrate of PRMT9, linking its function to pre-mRNA splicing. However, the biological function of PRMT9 and the molecular mechanism by which PRMT9-catalyzed SF3B2 arginine methylation regulates pre-mRNA splicing are still unknown. Recently, mutation of PRMT9 was identified in autosomal recessive intellectual disability (ID) from a large whole genome sequencing study of 136 consanguineous families, underscoring the significance of understanding the biological function and regulation of this newest PRMT. In our preliminary studies, we determined that the ID-associated PRMT9 mutation abolishes its arginine methyltransferase activity on SF3B2, and the mutant protein is unstable and subject to heavy ubiquitination. By establishing a novel Prmt9 conditional knockout mouse model, we revealed that PRMT9 loss causes abnormal synapse development and impairs mouse learning and memory. Mechanistically, we discovered a critical protein-RNA interaction between the arginine 508 (R508) of SF3B2, the site that is exclusively methylated by PRMT9, and the pre-mRNA anchoring site, a cis-regulatory element located upstream of the branch point sequence (BPS). Additionally, we uncovered two molecular pathways that regulate PRMT9 expression at the mRNA and protein levels. Built upon these exciting discoveries, our central hypothesis is that PRMT9-mediated arginine methylation regulates SF3B2–anchoring site interaction and U2 snRNP recruitment, a process that is critical for pre-mRNA splicing and brain development. This hypothesis will be tested in three specific aims: 1) determine how PRMT9-mediated SF3B2 arginine methylation regulates pre-mRNA splicing; 2) define the molecular pathways that regulate PRMT9 expression; and 3) investigate how PRMT9 loss-of-function impacts synapse development and circuit connectivity. Successful completion of these proposed studies will reveal a novel and fundamental molecular network underlying the regulation of RNA splicing in brain development. The key pathways revealed in this study could be harnessed as targets for treating neurodevelopmental disorders.

NIH Research Projects · FY 2025 · 2024-02

PROJECT SUMMARY / ABSTRACT Triple negative breast cancer (TNBC) is a highly aggressive cancer that has a poor prognosis and limited treatment due to the high propensity for metastatic progression and absence of specific targeted treatments. Although the immune checkpoint inhibition represents a breakthrough in the treatment of diverse cancers, the response rate to PD-1 or PD-L1 antibody remains only at about 4.8%-26% in TNBC patients. A daunting challenge for improving efficacy of immunotherapy such as PD1 antibody therapy is how to turn suppressed CD8+ T cells to activated cells after the checkpoint-imposed brake is removed. Our lab and others have shown that persistent STAT3 activation is critical for suppression of antitumor immunity. Our recent studies demonstrated that PD-1-ligation on CD8+ T cells activates STAT3 to upregulate CPT1B/fatty acid oxidation (FAO) while inhibiting glycolysis/IFNγ and CD8+ T cells antitumor effector. However, the compromised antitumor effects of CD8 TEFF cells are heavily impacted by suppressive immune cells including CD5+ B cells. We reported that cancer-promoting role of B cells depends on CD5-induced STAT3 activation. In addition to CD5+ B cells, in our pilot study performed under 1R21CA241283, we found that CD5 and PD-1 are co-expressed highly on mouse breast cancer T cells, which increased STAT3 activity in tumor infiltrating CD8+ T cells to suppress T cell anti- tumor function through regulating FAO/glycolysis. Furthermore, we demonstrated that targeting CD5 on T and B cells enhanced the anti-tumor effect of PD-1 blockade in PyMT mouse model of breast cancer. However, how to turn up glycolysis and stimulate IFNγ expression in TEFF cells to improve the efficacy of antitumor immune therapy in breast cancer patients remains elusive. Guided by our preliminary data, we hypothesize that (i) CD5/PD-1- induced STAT3 activation in T cells inhibits the antitumor immune responses of CD8+ T cells and counteracts PD-1 antibody therapy by promoting FAO and inhibiting glycolysis in breast cancer patients. (ii) effectively inhibiting STAT3/FAO in tumor-associated T cells, which can be achieved by blocking CD5, will “fuel” tumor infiltrating CD8+ T cells to meaningfully improve the antitumor efficacy of PD-1 blockade. We will accomplish our overall objective by pursuing the following specific aims: Aim 1: To assess whether CD5/PD-1-STAT3 axis- regulated FAO and glycolysis metabolism in T cells from patient breast tumor samples is associated with CD8+ TEFF cell exhaustion and failure to benefit from PD-1-directed therapy. Aim 2: To assess the antitumor effect and mechanisms through which CD5 and PD-1 antibodies alters T cell metabolism to enhance CD8 TEFF activity in patient-derived tumor organoids (PDTOs) and mice bearing patient-derived xenografts (PDXs) with autologous T cells or human immune system. Together, these studies will provide new mechanistic insights and key preclinical evidence of therapeutic resistance to PD-1 blockade, and will identify novel therapeutic strategy for overcoming resistance to PD-1 blockade and extend the benefits of PD-1 blockade therapy for TNBC patients.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY In the US, 42.4% of the adults are classified as obese. Obesity associated metabolic diseases, including type 2 diabetes (T2D), nonalcoholic fatty liver disease (NAFLD) and its progressive stage nonalcoholic steatohepatitis (NASH), severely impact human quality of life and mortality. Current anti-obese and diabetic drugs are often associated with different side effects, and currently there are no Food and Drug Administration (FDA) approved drugs for NAFLD/NASH. The long-term goal is to meet this critical unmet need for innovative novel pharmacological therapies with improved safety profiles and durable treatment effects for obesity and its complications. Bile acids (BAs) play important roles in regulating lipid, glucose, and energy metabolism. CYP8B1, is exclusively expressed in the liver and it functions specifically to control the ratio of 12α- hydroxylated (OH)/non-12α-OH BAs in the-BA pool. CYP8B1 deficiency leads to reduction 12α-OH)/non-12α- OH BAs ratio, resulting in metabolically beneficial effects including weight loss, ameliorated fatty liver and inflammation, with insulin sensitivity, in mice and humans. The central hypothesis is that CYP8B1-specific inhibitors may provide a unique pharmacological approach to treat obesity and related metabolic diseases. The rationale for the proposed research is that no CYP8B1-specific inhibitors exist and the lack of high-throughput screening (HTS) methods hinders the identification of CYP8B1-specific inhibitors. The objective of this grant application is to develop strategies for CYP8B1 inhibition by identifying potent CYP8B1-specific inhibitors to pharmacologically alleviate metabolic disorders in mouse disease models. We have established a high- throughput screening (HTS) platform to identify and optimize CYP8B1-specific inhibitors with potent in vivo efficacy in metabolic disease models toward drug development. The HTS was implemented for a ~45,000- compound library that led to the identification of potent, specific CYP8B1 inhibitors with biochemical and functional validation. Most importantly, we found that a lead CYP8B1 inhibitor compound that displayed strong efficacy in inhibiting CYP8B1 in vivo. The novel HTS-assay and the identified candidate CYP8B1-inhibitors will be used to test two Specific Aims: 1) identification and characterization of CYP8B1-specific inhibitors; and 2) Determination of CYP8B1-specific inhibitor therapeutic effects in metabolic disease models. The completion of this grant proposal will help us generate first-in-class candidates for drug development. The results will positively impact efforts to improve obesity, T2D and NAFLD/NASH clinical outcomes.

NIH Research Projects · FY 2026 · 2023-12

Myeloid neoplasms such as chronic myelogenous leukemia (CML) and myeloproliferative neoplasms (MPN) may over time transform from a chronic phase (CP) into, respectively, blast crisis (BC) or secondary acute myeloid leukemia (sAML). Both of these conditions are poorly responsive to currently available therapies and novel approaches are urgently needed. To this end, we have studied distinct mechanisms of BC/sAML transformation to discover new targets and develop corresponding therapeutic approaches that prevent and cure disease transformation. A common goal of all these studies is to eliminate leukemia stem cells (LSCs) that drive leukemia growth and BC/sAML transformation. In addition to mechanisms of BC/sAML transformation intrinsic to LSCs, we have recently gained insight into those that are extrinsic to LSCs and that involve transformation of the normal bone marrow (BM) microenvironment (or niche) into a “leukemic” one. Herein, we focus on leukemogenic mechanisms that induce T cells' loss of activity and exhaustion and that ultimately drive LSCs' escape from T cells' antileukemic immune surveillance. To this end, microRNAs (miRNAs) are small non-coding RNAs that target messenger RNAs and regulate levels of the corresponding proteins. MIR142, a highly conserved “gene”, encodes miR-142, which is involved in the development and regulation of hematopoiesis and native and adaptive immunity. In humans, MIR142 has been found mutated in lymphoma and AML, but not in CML or MPN. We recently showed that miR-142 downregulation (deficit) occurs in CD34+CD38- LSCs from BC CML patients compared with those from CP CML patients, leading us to postulate its contribution to the evolution of CP-LSCs into BC-LSCs. While investigating these findings in mouse models, we observed that compared with the Mir142+/+BCR-ABL mouse (a CP CML model), the Mir142−/−BCR-ABL mouse (a BC CML model) presented with a profound T cell lymphopenia, and a significant reduction of T cell activity. To this end, we also observed: a. reduced miR-142 level and activity in T cells from BC CML patients compared with those from CP CML patients; b. an association of miR-142 deficit with increased PD-1 expression and T cells' exhaustion; and c. correction of miR-142 deficit with a novel synthetic miR-142 mimic (CpG-M-miR-142), which restored the antileukemic activity of T cells from both human and murine BC/sAML models. Thus, we hypothesize here that miR-142 deficit is acquired by T cells during BC/sAML transformation and contributes to the phenotypic evolution of CML/MPN into BC/sAML via mechanisms that allow LSCs to escape from T cell antileukemic surveillance. To prove our hypothesis, we propose three specific aims (SA): SA#1: To determine the role of the T cells' miR-142 deficit in BC/sAML transformation. SA#2: To dissect the molecular mechanisms through which miR-142 deficit mediates T cells' loss of activity during BC/sAML transformation. SA#3: To target miR-142 deficit in T cells in order to prevent and/or cure BC/sAML transformation. Our ultimate goal is to obtain IND-enabling evidence that CpG-M- miR-142 is a novel targeting therapeutic that restores T cells' immunity, and in turn eliminates BC/sAML LSCs.

NIH Research Projects · FY 2026 · 2023-12

Project Summary Treatment outcomes for patients with acute myeloid leukemia (AML) have continued to lag behind outcomes reported for other hematological malignancies like acute lymphoblastic leukemia (ALL), in part because of the relatively slow development of immunotherapy (including immune checkpoint inhibitors [ICI]) compared with ALL. However, translation of currently available immunotherapies to AML treatment has been challenging, and novel AML-targeting drugs with immune-stimulating activity are greatly needed. The cGAS-STING signaling is a major pathway that promotes an extrinsic type I Interferon (IFN-I) response, which potently primes T cells in “immune- cold” cancers, including AML. My laboratory has been active in defining the function of protein arginine methyltransferases (PRMTs) in leukemia. Herein, our preliminary studies on the basis of primary AML cells as well as murine AML models reveal an anti-AML immune activation seen after inhibition of PRMT9, a most recently defined PRMT. Specifically, PRMT9 inhibition stimulated leukemia-intrinsic cGAS, as evidenced by cGAMP (an immunotransmitter) production, and induced a leukemia-eliminating IFN-I response in murine AML- microenvironment (ME). When combined with an ICI (PD1 inhibitor), the in-house PRMT9 inhibitor LD2 eradicated AML in animal models. Thus, we hypothesize that PRMT9 activity allows AML cells to evade immune surveillance by repressing cGAS-STING activity in the ME, and that LD2, alone or combined with an ICI (PD1 inhibitor), elicits T cell activity to eliminate AML cells. Extracellular cGAMP, which is hydrolyzed by ENPP1, and STING activation in host immune cells, are reportedly essential for immunity. Thus, in Aim 1, using two AML transplant models, we will determine whether downregulating cGAMP (via either ENPP1-overexpression or cGAS-knockout (KO) in AML cells) or STING-KO in the immune cell compartment of recipient mice would reverse PRMT9 inhibition-induced AML regression. In Aim 2, we will determine the mechanisms underlying cancer- intrinsic cGAS activation seen after PRMT9 inhibition. Specifically, we will define PRMT9 targets that when unmethylated underlie cGAS activation. In Aim 3, we will assess anti-AML activity of LD2 alone or combined with a PD1 inhibitor, using AML mouse models. Moreover, to define potential shifts in immune cell types/states after PRMT9 inhibition, we will perform single-cell transcriptome analysis on AML PDX cells from humanized mouse treated with LD2 or the combination. We are the first to identify PRMT9 as a druggable immune activation target against cancer. If successful, this work would support combining a PRMT9 inhibitor with ICIs as a therapy for AML, in which single ICI therapy has very limited effects.

NIH Research Projects · FY 2025 · 2023-09

The United States has led the world in drug discovery for over 50 years; however, the majority of scientific ideas that progress to clinical trials frequently come from Comprehensive Cancer Centers. Unfortunately, not all local universities and colleges have the resources to take their ideas forward to drug development and clinical trials. Through a partnership between University of California, Riverside (UCR) and City of Hope Comprehensive Cancer Center (CoHCCC), we aim to develop the resources, infrastructure, and training to help 1) UCR take their scientific ideas forward to develop therapeutic agents and ultimately initiate clinical trials and 2) mentor the next generation of drug development researchers and clinical trialists. Building on our successful P20 grant, in this U54 partnership, UCR and CoHCCC aim to develop the collaborations, resources, and training programs to improve drug development throughout the entire drug development pipeline. Our goal is for this program to become a focal point for UCR and CoHCCC to mentor and train cancer biologists, therapeutic and drug development scientists, and clinical trialists. Already, our P20 has fostered joint R01 grants, K01 grants, and pre-/post-doctoral fellowships. Both institutions are highly committed - CoHCCC contributed over $800K to our P20 grant and will contribute $250K/year to ensure the success of this U54 partnership. Aim 1 will strengthen UCR’s cancer research capacity and develop the resources to increase UCR/CoHCCC’s ability to jointly develop therapeutic agents. Aim 2 will increase the capacity of UCR and CoHCCC to jointly develop drugs and improve health. Aim 3 will provide the training, opportunity, and mentorship to foster the next generation of therapeutic scientists and clinical trialists.