Beckman Research Institute/City Of Hope

NIH Research Projects · FY 2026 · 2026-06

ABSTRACT Over 50,000 allogeneic hematopoietic cell transplantation (allo-HCT) procedures are performed annually worldwide for malignant and non-malignant diseases. While robust thymic function and T cell production are essential for a competent immune response during allo-HCT, thymic injury decreases thymic output of T cells, which increases the risk of malignancies, graft-versus-host disease (GVHD), morbidity, and all-cause mortality. Although the thymus is sensitive to allo-HCT preconditioning, it also has a remarkable capacity for repair. There is thus an unmet clinical need for strategies to boost thymic function and address lymphopenia, particularly for patients undergoing cancer therapies and for recipients of allo-HCT. We have demonstrated that thymic regeneration is critical for the renewal of immune competence following thymic injury and that we can harness pathways driving thymic regeneration to promote immune reconstitution and improve outcomes in allo-HCT. Tregs are known to actively orchestrate the repair of other organs and likely play a critical role in the recovery of thymic epithelial cells (TECs). Our preliminary preclinical studies suggest that a pathway of endogenous thymic regeneration is mediated by a recently identified population of mature, recirculating thymic regulatory T cells (Tregs). These cells migrate between the periphery and the thymus, coexist with newly generated cells during negative selection, and secrete regenerative factors such as amphiregulin (AREG) and Trefoil factor 1 (TFF1). Our overarching hypothesis is therefore that Tregs and their products are important mediators of regeneration in the thymus after allo-HCT. We propose to study AREG (Aim 1) and TFF1 (Aim 2) secreted by Tregs as mediators of T cell and thymic regeneration after allo-HCT. Specifically, we will use preclinical models to evaluate the role of AREG and TFF1 in thymic regeneration and immune reconstitution in allo-HCT with and without GVHD, and to understand their interactions with the metabolic function and signaling of TECs through the epidermal growth factor receptor (EGFR) pathway. We will also determine whether an analogous population of recirculating Tregs persists in the human thymus post-injury and analyze the clonal expansion and functional profile of these cells. Finally, we will investigate the therapeutic potential to ameliorate immune reconstitution in allo-HCT and acute GVHD by enhancing thymic regeneration through adoptive transfer of thymic Tregs and their products (Aim 3). This study builds upon our extensive experience studying allo-HCT and thymic regeneration in both preclinical models of thymic injury and human cohorts. We expect our proposed mechanistic and preclinical studies to improve our understanding of recirculating thymic Tregs and position us to translate our findings into the clinic. These clinical approaches could benefit allo-HCT recipients, as well as individuals receiving radiotherapy or chemotherapy or suffering from T cell deficiencies due to age-related lymphoid atrophy, autoimmune diseases, infectious diseases, and shock.

NIH Research Projects · FY 2026 · 2026-06

Project summary Mantle cell lymphoma (MCL) is an incurable disease with frequent relapses despite high initial remission rates. MCL primarily originates from naïve B-cells that harbor the hallmark t(11;14)(q13;q32) chromosomal translocation leading to overexpression of the cell cycle regulatory protein cyclin D1. B-cell receptor (BCR) signaling is an oncogenic driver in mantle cell lymphoma (MCL). Inhibiting the BCR-downstream Bruton’s tyrosine kinase (BTK), using the drug ibrutinib, has proven to be a successful treatment although its effectiveness is only temporary. The causes of resistance to ibrutinib are not completely understood. Patients who initially respond to ibrutinib, but later experience relapse, face a grimmer outlook, emphasizing the need to understand resistance mechanisms for better treatment options. Through analysis of gene expression from MCL patients treated with ibrutinib, we have discovered several potential mechanisms that may enable MCL to evade the selective inhibition of BTK. Among these, increased expression of the plasma-cell biomarker syndecan-1 (SDC1/CD138) and erb-b2 receptor tyrosine kinase 4 (ERBB4) are particularly prominent in ibrutinib-resistant cases. We generated isogenic MCL cell-line models of ibrutinib resistance and independently demonstrated the critical role of ERBB signaling as observed in clinical samples. Importantly, simultaneous inhibition of these newly identified drug-resistance pathways with FDA-approved drugs, afatinib (targeting ERBB) and trametinib (targeting MEK), demonstrated significant and synergistic cytotoxicity against MCL cells resistant to ibrutinib. These initial findings provided the basis for our central hypothesis that MCL tumors can overcome BTK dependence by shifting towards survival mechanisms mediated through ERBB signaling. The objectives of this proposal are to elucidate the mechanisms driving resistance to BTK inhibition and to identify potential therapeutic targets that could effectively treat refractory and relapsed MCL. We will test the hypothesis by accomplishing the following specific aims: SA1, determine the role of ERBB signaling in ibrutinib-resistant (IR) MCL; and SA2, determine the efficacy of combination therapy using approved drugs such as afatinib and trametinib to treat IR MCL in vitro and in vivo using MCL patient-derived xenografts (PDXs). In vivo studies in immunodeficient mice are necessary to assess efficacy, pharmacodynamics of target engagement, and tolerability of drug combination in MCL PDX tumors within a physiologically relevant microenvironment, which cannot be adequately modeled in vitro or ex vivo alone. Our study proposal will have a two-fold impact on cancer research and patient outcomes. By understanding how a specific cancer can find ways to escape treatment, we can change the way we approach cancer research. We will not just focus on one target; we will also watch out for other potential ways the cancer might resist treatment. This understanding of how resistance works can also benefit patients because we can predict which treatments might work best based on their genetics and the likelihood of the cancer coming back. This way, we can tailor treatments to each patient and increase their chance of beating cancer.

NIH Research Projects · FY 2026 · 2026-06

Project Summary/Abstract This proposal seeks to identify and characterize critical regulators of macrophage-mediated Programmed Cell Removal (PrCR), a vital process in tumor surveillance and elimination. While immunotherapy has achieved significant breakthroughs, substantial challenges persist, including insufficient immune cell infiltration into tumors and compromised immune responses caused by the immunosuppressive tumor microenvironment (TME). Tumor-associated macrophages (TAMs), which are among the most abundant immune cell populations within the TME, represent a promising target for immunotherapy. PrCR, a process of macrophage-mediated cancer immunosurveillance, involves the recognition and phagocytosis of target cells, playing a critical role in tumor control. However, in contrast to the well-established field of T cell immune checkpoint therapies, the identification and characterization of innate immune checkpoints regulating PrCR remain limited, highlighting a significant knowledge gap. A better understanding of these innate immune regulatory mechanisms is essential to advance the development of effective PrCR-based immunotherapies. Using a high-throughput CRISPR-based screening approach, we identified a novel stimulatory innate immune checkpoint that interacts with the lymphocyte function- associated antigen 1 (LFA-1) receptor on macrophages to induce PrCR. Suppression of the stimulatory phagocytosis checkpoint in preclinical models significantly reduced PrCR and promoted tumor growth, whereas its overexpression enhanced macrophage phagocytosis and inhibited tumor development. These findings provide a compelling rationale for further investigation into the underlying mechanisms by which stimulatory phagocytosis checkpoints mediate cancer-immune interaction and regulate PrCR. In Aim 1, we will evaluate the role of such stimulatory phagocytosis checkpoint in regulating PrCR and its impact on tumor development and the efficacy of immunotherapy using preclinical models of triple-negative breast cancer (TNBC) and microsatellite-stable colorectal cancer (MSS CRC). Aim 2 will focus on elucidating the molecular mechanisms by which the stimulatory checkpoint interacts with and activates LFA-1 in macrophages. This will involve resolving high-resolution structures of the phagocytosis checkpoint complex using cryo-electron microscopy, coupled with molecular dynamics simulations, to uncover the mechanism underlying their activation and signaling. Aim 3 will investigate how activation of the LFA-1 receptor drives phagocytic machinery and reprograms macrophages for sustained PrCR. Together, this research aims to establish a previously unrecognized stimulatory phagocytosis checkpoint and elucidate the underlying mechanisms for its activation and function in PrCR. These findings will significantly enhance our understanding of macrophage-mediated cancer immunosurveillance and create new opportunities to develop innovative cancer immunotherapies that harness the tumoricidal potential of macrophages.

NIH Research Projects · FY 2026 · 2026-05

Project Summary This project investigates how the native pancreatic microenvironment shapes human islet cell function, in health and type 1 diabetes (T1D). While most human islet studies rely on isolated islets, this approach disrupts key tissue interactions, potentially obscuring physiological and disease-relevant responses. We will leverage human pancreatic tissue slices, which preserve native cellular architecture, to directly compare hormone secretion and calcium signaling between slices and isolated islets from the same donors. Our first aim will quantify the impact of removing islets from their microenvironment by comparing responses to endocrine stimuli in paired samples from both T1D and non-diabetic donors. The second aim will evaluate how non-endocrine cells (e.g., acinar, ductal, vascular, neuronal) influence islet cell activity by stimulating these populations in situ and measuring endocrine cell responses. Together, these aims will generate critical insights into the physiological relevance of widely used model systems and identify key paracrine signals that regulate islet function. This study fills a major gap in our understanding of how the human islet environment contributes to hormone regulation and diabetes pathophysiology. It introduces a novel matched-comparison approach that integrates functional and imaging data across models and donor types, offering an innovative platform to advance islet biology. Findings from this work will inform the development of more physiologically relevant human islet models and support the design of targeted therapies for diabetes, directly contributing to public health by enhancing our ability to study and treat this disease.

NIH Research Projects · FY 2026 · 2026-05

PROJECT SUMMARY/ABSTRACT Type 1 diabetes (T1D) results from autoimmune destruction of the pancreatic β cells, leading to insulin deficiency and chronic hyperglycemia. Accumulating evidence suggests that β cell stress during disease progression may play an active role in amplifying this autoimmune response through the generation of non-canonical or immunogenic proteins. We and others have shown that proinflammatory cytokines (IFN-α and IFN + IL-1) activate downstream signaling pathways within the β cell and create a feedforward cycle of inflammation and endoplasmic reticulum (ER) stress. Moreover, a growing body of evidence indicates that chronic ER and oxidative stress lead to the activation of the integrated stress response (ISR), a cytoprotective mechanism that maintains cellular protein homeostasis in response to environmental and cellular stress signals. However, chronic activation of ISR may lead to dysregulation in global RNA translation and the generation of immunogenic peptides, particularly neoantigens. These, when presented within the HLA complex, can initiate autoimmunity against β cells in early T1D. Our long-term goal is to identify how distinct cytokine signatures impact or influence  cell stress pathways during different stages of T1D progression. Thus, we hypothesize that proinflammatory cytokine-mediated activation of ISR in early T1D amplifies β-cell dysfunction and immunogenicity via changes in RNA translation and increased neoantigen production and presentation. We aim to test this hypothesis by: 1) defining how proinflammatory cytokines individually and in combination selectively modulate β cell mRNA translation and production of known neoantigens and map non-canonical RNA translation (ncORF) or untranslated open reading frames (uORF) or frameshift translation to the non-canonical proteins that are actively translated during inflammatory stress in human islets and 2) 3D spatial evaluation of disease specific RNA and protein translation in intact pancreatic tissue sections of autoantibody positive (AAb+), type 1 diabetes (<1 year of duration) and age, BMI and sex-matched non-diabetic control organ donors. This research is innovative because it employs innovative multi-omics approaches to map the distinct and global effects of selected cytokines and temporally monitor their impact on islet cell RNA translation with high precision. The proposed work will significantly advance our understanding of how β cells are altered in their function, stress pathways and antigenic profile during the early stages of T1D. Results from these studies have the potential to inform the development of biomarkers and therapies aimed at preventing T1D and protecting the functional β cell mass.

NIH Research Projects · FY 2026 · 2026-05

PROJECT SUMMARY Patients diagnosed with glioblastoma (GBM) have a median overall survival of less than two years even after receiving multimodal therapies. Multiple factors account for treatment resistance including: 1) Inability of therapies to cross the blood-brain barrier to reach invading cells; 2) GBM’s molecular heterogeneity and overlapping escape mechanisms that overcome targeted therapies; 3) Evasive mechanisms that render GBMs resistant to immunotherapy. Therefore, there is an unmet need for GBM treatment approaches that address multiple resistance mechanisms. The receptor for advanced glycation end products (RAGE) is a member of the immunoglobulin superfamily, which was discovered as a transmembrane receptor for the products of nonenzymatic glycation and oxidation of proteins. RAGE is expressed by glioma cells and is activated by ligands present in the GBM tumor microenvironment (TME). Activation of RAGE stimulates multiple signaling pathways that promote GBM progression. Recently, we demonstrated that genetic ablation of intracellular RAGE in gliomas inhibited multiple oncogenic pathways that not only regulate glioma growth and invasion, but also improve the efficacy of immunotherapies by inhibiting galactin-3 production and promoting an immunologically “permissive” TME. Based on these observations, we propose to target RAGE as a multifaceted therapy for GBM. Our central hypothesis is that RAGE inactivation will suppress oncogenic pathways that are important for GBM growth and invasion and enhance responses to immunotherapy. Three aims are proposed to test this hypothesis. Aim 1 will optimize a multifunctional anti-RAGE oligo-based strategy to target GBM. As an alternative approach to blocking RAGE extracellular receptor with small molecules, we propose to inhibit its signaling by optimizing the design of anti-RAGE antisense oligos. The “scavenger” receptor property of RAGE will be exploited to develop multifunctional oligos that are rapidly internalized and selectively silence RAGE signaling in GBM models. Aim 2 will test the in vivo efficacy and determine the mechanism of RAGE ablation in enhancing the anti-tumor immune responses in syngeneic murine GBM models. Findings from this Aim will uncover novel strategies that could enhance immunotherapy efficacy in these resistant tumors. Finally, Aim 3 will assess the combinational effects of anti-RAGE oligos with immunotherapy. In this Aim, we will also perform the pre-clinical studies to optimize the dosing regimen of galactin-3 inhibitors for future GBM clinical trials. To perform this work, we have assembled a multidisciplinary and highly collaborative team with expertise in neuroimmunology, neuro-oncology, and molecular biology. The success of these studies, supported by compelling preliminary data, is expected to lead to the development of novel and critically needed GBM therapies.

NIH Research Projects · FY 2026 · 2026-05

Abstract The sense of smell, mediated by olfactory receptors (ORs), is essential for human interaction with the environment. ORs play a critical role in detecting and distinguishing a vast array of chemical compounds known as odorants. However, the mechanism by which a limited number (~400) of human ORs can react to the vast number of odorants remains unclear. Recent studies have revealed that certain drugs may potentially disrupt OR function and lead to taste or smell disorders, highlighting the need to further our understanding of OR-odorant selectivity, an area that has not been extensively studied due to lack of structural information. This project will utilize the latest resolved OR structures, to which I made significant contributions, and aims to explore the underlying allosteric mechanisms of OR function and develop innovative therapeutic strategies to address sensory disorders, focusing specifically on the olfactory receptors OR51E2 and OR1A1. This research will utilize advanced computational methods, including long-timescale molecular dynamics (MD) simulations and allosteric pathway analysis, to investigate the structural and dynamic properties of ORs. By identifying critical residues and pathways involved in ligand binding and receptor activation, this project seeks to uncover the allosteric mechanisms that regulate OR activity. Furthermore, the study will explore the impact of diverse lipid environments on OR function, using multiscale simulations to understand how external factors influence ligand binding and receptor dynamics. This comprehensive approach will provide valuable insights into the modulation of OR activity by lipids and other extracellular elements. The ultimate goal is to develop novel therapeutic strategies, including negative allosteric modulators and Proteolysis Targeting Chimeras (PROTACs), to modulate OR activity and address sensory disorders. These efforts will pave the way for new treatments for smell or other sense disorder, enhancing our understanding of OR regulation and improving sensory health. This award will support my training and development, allowing me to gain expertise in cutting-edge computational techniques and establish an independent research career focused on sensory biology and OR function at City of Hope.

NIH Research Projects · FY 2026 · 2026-05

PROJECT SUMMARY/ABSTRACT (30 lines or less) The premise for the proposed research stems from precedence in other diseases, such as cancer, cardiovascular, neurodegenerative, and most relevant to the current proposal, autoimmune diseases, in which circulating extracellular vesicles (cEVs) play a role in pathophysiology and are important biomarkers for early detection. However, little is known concerning the role of cEVs in human type 1 diabetes (T1D). Our overall hypothesis is that cEVs have the potential to be used as biomarkers for early pre-disease detection of T1D, based on their distinct molecular and functional phenotype in T1D and pre-disease stages, compared to healthy or low risk individuals. Our specific aims are: 1) To identify the distinct protein and RNA cargo unique to cEVs at different stages of T1D disease development; and 2) To investigate the effect of cEVs from subjects at different stages of disease progression on immune and beta cell phenotypes and elucidate the functional relevance of their distinct molecular cargo. To address these aims, we have assembled a team of investigators with highly relevant expertise and techniques. We propose to use longitudinal samples from The Environmental Determinants of Diabetes in the Young (TEDDY) study of children with T1D associated genetic risk to identify critical timepoints and underlying mechanisms mediating, A) the earliest stages of pathogenesis preceding AAb appearance of the first islet autoantibody, and B) the period of seroconversion from single to multipe AAb+ or remaining single AAb+ and C) the period after multiple AAb appearance with a highly variable rate of progression to hyperglycemia. We have the expertise and technical ability to isolate cEVs from plasma, perform proteomic and RNAseq analysis on EVs, and perform immune and beta cell related functional assays. Our Research Plan is to generate cEVs from donors at different stages of T1D disease progression, to identify the uniquely packaged protein and RNA cargo from these cEVs, to evaluate the effects of the cEVs on immune cell functional phenotype and islet health and elucidate the functional relevance of the distinct molecular cargo targets. These studies will yield novel mechanistic insights into early disease pathogenesis and identify potential novel biomarkers for T1D initiation and progression.

NIH Research Projects · FY 2026 · 2026-04

PROJECT SUMMARY T cell activation is a crucial step in initiating adaptive immune responses against pathogens such as viruses and bacteria. This process requires precise transcriptional regulation to ensure a rapid and robust immune response while preventing aberrant activation that could lead to autoimmunity or excessive inflammation. However, the mechanisms by which T cells achieve this balance to maintain immune homeostasis remain unclear. In recent years, protein arginine methylation has emerged as a key regulatory mechanism in T cell biology, modulating activation, differentiation, and cytokine production. Although targeting protein arginine methyltransferases (PRMTs) has shown promise in regulating immune responses, their therapeutic potential is limited by systemic adverse effects due to the essential roles of these enzymes in many fundamental cellular processes. We propose that targeting specific downstream effectors of arginine methylation may offer a more precise therapeutic strategy while minimizing adverse effects on normal tissues. To achieve this, we focus on Tudor domain-containing protein 3 (TDRD3), the primary methylarginine reader and effector that plays a key role in arginine methylation-mediated cellular processes, including transcription regulation. Our preliminary data reveal that, although TDRD3 is not required for normal development, knockout of TDRD3 in CD4+ T cells confers resistance to experimental autoimmune encephalomyelitis by impairing T cell activation, proliferation, and survival. Mechanistically, TDRD3 functions as a transcriptional coactivator for genes critical to T cell activation, including interleukin-2 (IL-2), the master regulator of T cell-mediated immune responses. Additionally, we discovered that T cell activation is associated with a robust increase in R-loops—three- stranded DNA structures composed of a DNA/RNA hybrid and a single-stranded DNA—particularly at the IL-2 gene promoter. We also identified a novel TDRD3-interacting protein complex that may be responsible for R- loop regulation during T cell activation. The goal of this project is to define how TDRD3-regulated R-loop dynamics influence T cell-mediated immune responses. Specifically, we will: 1) elucidate the molecular mechanisms by which TDRD3 acts as a coactivator for transcription activation, focusing on IL-2 as a major target; 2) investigate the role of TDRD3 and its interacting protein complex in R-loop regulation during T cell activation; and 3) evaluate the therapeutic potential of targeting TDRD3-mediated transcription regulation to modulate T cell immune responses in autoimmune and infectious disease models. Results from these experiments will uncover fundamental mechanisms of transcription regulation in T cells. The key pathways revealed in this study could serve as novel targets for treating immunological disorders.

NIH Research Projects · FY 2026 · 2026-04

PROJECT SUMMARY: Epstein-Barr virus (EBV) infection is associated with infectious mononucleosis, several autoimmune diseases, and multiple cancers. Yearly, there are ~350,000 new cases of EBV-associated cancers, and ~200,000 deaths from such cancers. Despite the health burden posed by EBV infection, there is no licensed prophylactic vaccine against primary EBV infection. Previous clinical efforts focused on only one glycoprotein (gp) vaccine target (i.e., gp350) and yielded unsatisfactory results. To improve EBV vaccine immunogenicity and protective efficacy, our efforts have focused on developing a single vaccine that combines gp350 with four additional gps that are key for viral entry and are all targets of neutralizing antibodies (nAbs): gB, gp42, and gHgL. Our studies using virus-like particle- and viral vector-based platforms have confirmed that a multivalent approach is immunogenic in multiple animal models, eliciting antibodies with superior in vitro and in vivo neutralizing activity than those elicited by gp350-monovalent vaccine controls. In the proposed project, we seek to employ circular RNA (circRNA), a novel RNA-based vaccine platform that improves upon traditional virus-like particle-, viral vector-, and RNA-based platforms, as a platform for pentavalent EBV vaccine development. Our hypothesis is that immunization with a pentavalent circRNA EBV vaccine expressing five gps involved in viral entry will elicit protective antibody-mediated humoral responses with nAb activity in vitro, ex vivo, and in vivo as tested in three EBV challenge models. We will test this hypothesis using unique yet complementary EBV challenge animal models that will allow us to comprehensively assess vaccine immunogenicity and efficacy against systemic infection acquired through non-natural (intravenous, rabbit) and natural (oral, common marmoset) routes, and against infection of human B cells (humanized mice). In Aim 1, we will first characterize the immunogenicity of the pentavalent circRNA-based EBV vaccine in immunized rabbits; then, we will characterize the in vivo protective efficacy of the vaccine against systemic EBV infection in immunized rabbits after intravenous viral challenge, as well as against human B cell infection in humanized mice after passive immunization with immune rabbit sera followed by viral challenge. In Aim 2, we will similarly assess vaccine immunogenicity and in vivo protective efficacy in common marmosets after immunization and oral EBV challenge. In both Aims, we will compare immunogenicity and efficacy of our circRNA-based vaccine candidate to that elicited by other pentavalent comparators and gp350-based monovalent controls. In summary, our short-term goal is to characterize a novel pentavalent circRNA-based EBV prophylactic vaccine and assess its immunogenicity and efficacy in three complementary animal models, paving the way for future clinical translation. Successful completion of this project will help advance our long-term goal to develop effective vaccines that prevent or limit primary EBV infection and its associated diseases.

NIH Research Projects · FY 2026 · 2026-03

Project Summary: Colorectal cancer (CRC) is potentially preventable, but still ranks 3rd in diagnosis and 2nd in mortality worldwide: why? CRC screening trials based on fecal immunochemical testing (FIT) showed survival benefits through a stage-shifting effect, reducing stage IV diagnoses but not preventing CRC incidence. FIT sensitivity declines for early-stage CRC and precursor lesions (the advanced adenomas, AAs), emphasizing its impact on mortality rather than incidence. Endoscopy-first approaches have a higher sensitivity, but the contentious results of the NordICC trial demonstrated the numerous challenges of a colonoscopy-first approach, including invasiveness, costs, and patient compliance. In fact, only 54% of the US screening-eligible population has received a colonoscopy in the past 10 years. However, non-invasive tests promote participation. So far, biomarker studies have demonstrated great sensitivity to CRC but not to AAs and early-stage CRC. These investigations operated under the assumption that the same analyte would effectively detect both CRC and AA. However, 1) if both CRC and AAs release the same analyte into the blood, AAs do so in considerably smaller quantities; and 2) adenomas and CRC exist at opposite ends of the adenoma-carcinoma sequence; therefore, they may not yet release the same analytes. Therefore, in this proposal, we advocate for the development of an innovative liquid biopsy, a test that, built separately for AAs and CRC, can assess both AAs and CRC from a single blood draw to optimize patient compliance and resource allocation and enable timely AA detection for cancer prevention. MicroRNAs (miRNAs) are small non-coding RNAs that regulate genes implicated in every human cancer, including CRC and AAs, and may thus be ideal biomarkers. Indeed, circulating cell-free miRNAs (cf-miRNAs) have been shown to have diagnostic potential. Furthermore, the recent discovery that cancer cells actively excrete miRNAs in small extracellular vesicles called exosomes (exo miRNAs) has revolutionized the field, as tumor-derived exosomal cargo enables the identification of cancer-specific molecular markers. To date, our studies have been among only very few to directly compare the clinical significance of cf-miRNAs vs. exo- miRNAs, and our team is unique in proposing a combination of cf- and exo-miRNAs as potentially superior biomarkers. To address the limitations of cf-miRNA biomarkers and to interrogate the potential clinical significance of exo-mRNAs, we completed a systematic and comprehensive biomarker discovery and validation effort in patients with CRC, AAs, and low-risk adenomas. These successes collectively demonstrate our ability to establish a transcriptomic signature for the early detection and prevention of CRC. In this proposal, we will complete three milestone-based specific aims: 1) Refinement and large-scale validation of our assay to establish its performance in a cohort of patients with CRC and AAs. 2) Validate our assay in a cohort of individuals younger than 50 years of age. 3) Evaluate the ability of our transcriptomic signature to detect CRC and AAs at their earliest stages in pre-diagnosis serum specimens and determine lead time before disease presentation.

NIH Research Projects · FY 2026 · 2026-03

Abstract Acute myeloid leukemia (AML) is one of the most common and fatal forms of hematological malignancies caused by gene mutations and genomic rearrangements. The cure rates for AML patients have not significantly improved for decades. The molecular mechanisms underlying the pathogenesis of AML are not fully understood. By analysis of publicly available genomic data using a new machine learning approach, RNA Binding Motif Protein 33 (RBM33), an RNA binding protein, is identified as an essential gene in AML. However, the biological function of RBM33 is unknown yet. Our preliminary studies provide the first compelling evidence suggesting a novel function of RBM33 in regulating m6A RNA demethylation. More importantly, we showed that RBM33 knockdown significantly inhibited growth and survival of human and mouse leukemia cells. At a molecular level, we have identified a potential downstream target of RBM33 in leukemia cells. ALKBH5 is known as an m6A mRNA demethylase (Eraser), which removes m6A methylated groups from RNA. To date, it remains unknown whether another member of RNA binding proteins is required for ensuring recruitment of ALKBH5 to its mRNA targets. We have recently demonstrated that ALKBH5 has a critical role in AML development and maintenance. We hypothesize that RBM33 plays a critical role in the pathogenesis of AML by regulating ALKBH5-mediated m6A demethylation. To test this hypothesis, we will pursue three specific aims. In Aim1, we will determine a novel role of RBM33 in regulation of dynamic m6A RNA methylation in leukemia cells. In Aim2, we will investigate the role of Rbm33 in AML development and maintenance. In Aim 3, we will determine the downstream pathway that mediates the function of RBM33 in leukemogenesis in AML. We will employ both mouse genetic models as well as human patient-derived mouse models to elucidate the role of RBM33 in normal hematopoiesis and leukemogenesis in vivo, and will combine transcriptome and epitranscriptome analysis to identify the key downstream targets and associated downstream pathways that mediate the role of RBM33 in leukemogenesis. Our studies will uncover a novel role of RBM33 in m6A RNA modification, and define the importance and underlying mechanisms of RBM33 in AML development and maintenance as well as LSC/LIC self-renewal. Thus, the success of our project will significantly advance our understanding of the complex mechanisms underlying the m6A modification-mediated gene regulation in leukemia cells and the critical role of m6A RNA demethylation in leukemogenesis.

NIH Research Projects · FY 2026 · 2026-02

Summary Type 1 diabetes (T1D) affects ~2 million people in the US, including children. T1D is a major burden to society in terms of health costs, loss of productivity; it reduces quality of life and expectancy and causes loss of life due to acute and chronic complications. Its incidence is rising worldwide. T1D is mediated by an autoimmune process that destroys insulin-secreting pancreatic beta cells, leading to lifelong insulin-deficiency and hyperglycemia. No cure or fully effective prevention is available, but the key requirements for diabetes reversal are apparent: (1) preservation of residual beta cell mass; (2) promotion of beta cell regeneration; and (3) induction of immune tolerance. The discovery that inhibitors of the dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A) enhance human β-cell regeneration empowers exploring regimens that simultaneously address autoimmunity, β-cell dysfunction and, for the first time, β-cell loss. We will advance this concept in nonobese diabetic (NOD) mice and immunodeficient mouse models, supported by preliminary data showing that: 1) DYRK1A inhibitors together with the GLP-1 receptor agonist (GLP1RA) exenatide synergistically increase human β-cell proliferation and expand β-cell mass in vivo in human islet grafts transplanted into immunodeficient mice; 2) this combination affords protection from cytokine-induced human β-cell death and decreases immunogenicity in vitro; and 3), it rapidly reverses diabetes in newly diagnosed NOD mice pre-treated with anti- CD3 antibody. However, efficacy of short course immune therapies wanes over time and there is a need for safe, chronic treatments that promote immune regulation without inducing immunosuppression. A promising agent is IL-2: at low dose, it selectively promotes regulatory T cells (Tregs) and was safe and effective in treating autoimmunity in experimental models and in clinical trials for several immune-mediated disorders. Our central hypothesis is that DYRK1A inhibition with GLP1R activation in combination with effective, long-term immunomodulation induces stable T1D remission in NOD mice. We propose that this therapy improves β- cell health and number, decreases immunogenicity, and is synergistic with immunomodulation that includes low dose IL-2 therapy to stimulate Treg cells. Specific Aim 1 will advance an expanded combination therapy with immunomodulators (anti-CD3 and IL-2/CD25 fusion protein), a novel and safer DYRK1A inhibitor, and clinically used GLP1Ras, for T1D reversal. Specific Aim 2 will establish the immunological basis by which Treg-targeting by low dose IL-2/CD25 together with immune modulation by anti-CD3 and β-cell enhancement by DYRK1A inhibition/GLP1R activation leads to T1D reversal. Specific Aim 3 will determine the mechanisms impacted by this combination therapy on the human β-cell and human immune cells. This highly translational work will characterize the therapeutic potential of the combination of DYRK1A inhibitors, GLP1RAs and immunomodulators to induce T1D remission, and the cellular/molecular mechanisms involved in the beneficial effects of this treatment, with relevance to both recent onset and established T1D.

NIH Research Projects · FY 2026 · 2026-02

Aberrant inflammation drives the development of autoimmune diseases and cancer. The incomplete understanding of molecular mechanisms controlling inflammatory activation and resolution remains a significant obstacle to developing effective treatments for inflammatory disorders. Therefore, the main objective of this proposal is to identify and characterize novel molecular pathways that regulate inflammatory responses. MicroRNAs (miRNAs) have emerged as essential regulators that fine-tune immune homeostasis through both activating and inhibitory functions. Among these, miR-146a serves as a paradigmatic example of an immunomodulatory miRNA that functions primarily as a negative feedback regulator of NFκB signaling. The indispensable role of miR-146a in immune homeostasis is demonstrated by miR-146a-deficient mice, which develop systemic autoimmunity, hematological malignancies, and bone marrow failure. Similarly, altered miR-146a expression in humans is associated with inflammatory diseases and cancer. Despite its clinical significance, the mechanisms controlling miR-146a biogenesis remain poorly understood. Our preliminary studies revealed an unexpected post-transcriptional mechanism regulating miR-146a production. We discovered that active protein synthesis is specifically required for miR-146a maturation, as translation inhibition arrests miR-146a processing at the precursor stage while other miRNAs are unaffected. Through an unbiased biochemical approach, we identified the tRNA synthetase EPRS1 as a specific pre-miR- 146a binding protein that recognizes its conserved apical loop. EPRS1 depletion significantly impairs miR-146a processing without affecting other miRNAs, establishing a novel regulatory axis linking protein synthesis machinery to inflammatory control. Our central hypothesis is that miR-146a biogenesis is controlled post- transcriptionally by EPRS1 and additional, yet-to-be-identified, translation-dependent factors. In Aim 1, we will define how EPRS1 regulates miR-146a biogenesis by characterizing their molecular interaction and identifying short-lived proteins required for EPRS1-dependent processing. In Aim 2, we will leverage CRISPR screen methodology to identify additional regulators of miR-146a biogenesis and determine their functional relationship to EPRS1. The proposed research is significant because it will reveal previously unappreciated mechanisms controlling miR-146a production and potentially explain its dysregulation in inflammatory and hematological diseases. Understanding these regulatory mechanisms may enable the development of innovative therapeutic strategies to restore miR-146a levels in patients, circumventing the limitations of direct miRNA therapeutics.

NIH Research Projects · FY 2025 · 2026-01

Abstract Myelodysplastic syndromes (MDS) are a group of diverse malignant hematological disorders that originate from hematopoietic stem cells (HSCs). Increased levels of reactive oxygen species (ROS) and DNA damage are commonly detected in hematopoietic cells from MDS patients. An elevated level of ROS, generated from either endogenous or exogenous sources including oncogene activation, leads to loss of quiescence and self-renewal of HSCs. ROS-induced DNA damage speeds up the aging process of stem cells and contributes to the mutagenesis associated with cancer development. m6A RNA methylation plays a significant role in multiple biological processes by introducing another layer of post-transcriptional regulation of gene expression within cells. The goal of this project is to elucidate the significant role of ALKBH5-mediated epigenetic regulation in the maintenance of genomic stability in hematopoietic stem/progenitor cell (HSPCs) during oxidative stress, and how deregulation of ALKBH5 contributes to promotion of leukemic transformation of HSPCs in the initiation and development of MDS. We found that ROS significantly increased global m6A RNA methylation in human cell lines, and that the elevation of m6A mRNA methylation is required for rapidly repairing ROS-induced DNA lesions and preventing cell death. Interestingly, we found that ALKBH5, the m6A RNA demethylase, is responsible for ROS-induced elevation of m6A mRNA methylation. ROS induced post- translational modification of ALKBH5, and inhibited the demethylase activity of ALKBH5. We showed that forced expression of ALKBH5 inhibited ROS-induced m6A mRNA methylation and significantly delayed repair of ROS-induced DNA damage. Thus, we hypothesize that aberrant expression of ALKBH5 disrupts HSPC functions by negatively influencing genome integrity and survival of HSPCs, thereby contributing to leukemic transformation of HSPCs during the initiation and development of MDS. In this proposal, we will determine 1) the role and underlying mechanism of ALKBH5 in the maintenance of genomic stability in HSPCs in response to oxidative stress; 2) the effects of ALKBH5/Alkbh5 overexpression on the maintenance of mouse and human primary HSPCs during ROS stress in vivo; and 3) whether ALKBH5/Alkbh5 is required for the maintenance of pre-leukemic stem cells (pre-LSCs) in MDS. Our study will provide new insights into novel mechanisms of MDS development and epitranscriptional regulation of gene expression in HSPCs in response to oxidative stress. Additionally, our study will provide the first set of evidence to support a significant role of ALKBH5- mediated m6A mRNA demethylation in the maintenance of normal HSPCs and pre-leukemic stem cell (pre- LSCs).

NIH Research Projects · FY 2025 · 2025-12

PROJECT SUMMARY. Antibody and antibody fusion protein therapies targeting adaptive immune cells have shown variable inter-subject levels of success at halting autoimmune destruction of the insulin-producing pancreatic β-cells in individuals recently diagnosed with Type 1 Diabetes (T1D). Responsiveness to many antibody therapies vary in other autoimmune diseases and cancer according to Fc gamma receptor (FcγR) variants impacting a patient’s capacity for Fc-mediated mechanisms of action (Fc-MoA) such as antibody dependent cellular cytotoxicity (ADCC) and antibody dependent cellular phagocytosis (ADCP). Indeed, pharmacodynamics of low-dose α-thymocyte globulin (LD-ATG) and clinical efficacy of rituximab (α-CD20) have been demonstrated to be affected by FcγR variants in the context of transplantation or rheumatoid arthritis and lymphoma, respectively. However, whether responsiveness to these therapies in individuals with T1D are similarly affected by FcγR-associated variants remains unknown. Therefore, I hypothesize that FcγR-associated variants may influence efficacy of therapeutic antibodies with FcγR-binding capabilities in recent-onset T1D via regulation of ADCC and ADCP. Indeed, I have identified a significant association between an FCGR2B expression QTL (eQTL) and quantitative metabolic response (QR) in the LD-ATG trial (TN19). The objectives of my proposed studies are to build on this finding by identifying genetic variants in FcγR loci that associate with therapeutic efficacy in additional completed T1D immunotherapy trials and to evaluate the consequences of modified FcγR expression and/or function on Fc-MoA. In Aim 1, I will perform quantitative trait locus (QTL) analysis of FcγR-associated single nucleotide polymorphisms (SNPs) and copy number variants (CNVs) versus treatment efficacy and target cell depletion in TrialNet (TN) and Immune Tolerance Network (ITN) studies of native Fc-containing antibody therapeutics in recent-onset T1D. I will analyze associations between genotype microarray, clinical endpoints, and flow cytometry data from the rituximab (TN05) and alefacept (Inducing Remission in T1D With Alefacept [T1DAL], ITN) trials. In Aim 2, I will determine impacts of FcγR-associated variants on cell type-specific FcγR protein expression and regulation of Fc-MoA of therapeutic antibodies for T1D. Here, I will characterize human whole blood samples by flow cytometry to evaluate associations between cell type-specific FcγR protein expression and previously defined whole blood eQTLs, with a focus on the LD- ATG response-associated FCGR2B variant. I will also employ rapid and fluorometric ADCC (RFADCC) assays to measure LD-ATG- and rituximab-mediated ADCC and ADCP induction by genotype-selected natural killer (NK) cell and monocyte effectors and T cell or B cell targets, respectively. I expect my findings will support a precision medicine approach to identify individuals with or at-risk for T1D who are likely to respond positively to native Fc-containing immunotherapies, as well as to guide dose optimization strategies for non-responders.

NIH Research Projects · FY 2025 · 2025-11

PROJECT SUMMARY 2025 Rachmiel Levine-Arthur Riggs Diabetes Research Symposium November 14-17, 2025, Westin Pasadena, Pasadena, CA Despite the significant knowledge obtained and the progress made in the treatment of diabetes over the last 50 years since the discovery of insulin, translation of this understanding to the clinic, and implementation of acceptable standards of care for diabetics has been suboptimal. In view of the rapidly growing worldwide diabetes epidemic - the disease affected 22.3 million people in the USA and 371 million people worldwide in 2012, and is expected to double by 2030 if current trends hold - it is imperative to enhance current interactions among investigators. This is to foster new collaborations, and pool knowledge and resources so that the cellular and molecular mechanisms responsible for the disease and its complications can be determined, and novel therapeutic strategies developed that will effectively prevent, delay, and even cure diabetes. The 2025 Rachmiel Levine-Arthur Riggs Diabetes Research Symposium, to be held from November 14-17, 2025 at The Westin in Pasadena, California, will continue to meet the growing demand to keep researchers, clinicians and trainees abreast of the latest developments in diabetes- and endocrine-related research. The Symposium is organized by the City of Hope’s Arthur Riggs Diabetes and Metabolism Research Institute (AR-DMRI) and is expected to attract over 250 attendees from both the U.S. and abroad with diverse academic backgrounds including, endocrinologists, diabetologists, islet biologists, stem cell and gene transfer scientists, transplant scientists, immunologists, cell biologists, young investigators in all these areas, and health care professionals who manage individuals with diabetes. The four-day meeting will offer presentations from over 60 experts in the field of type 1 diabetes and islet cell biology. The meeting will consist of introductory lectures and plenary sessions that will each conclude with a panel discussion. In addition, the meeting will offer a debate session, oral presentations from junior investigators and trainees, and a poster session for both junior and established investigators to present new and exciting data. The Symposium provides an important venue for investigators to present their data to an audience of national and international experts, helping to foster the career growth of junior investigators.

NIH Research Projects · FY 2025 · 2025-09

Project Summary Current treatments are still limited in their ability to safely mitigate obesity and its complications. Obesity develops due to an imbalance between nutrient intake and energy expenditure; therefore, there is still a need to better understand the mechanisms governing nutrition and energy balance, with the long-term goal of developing novel strategies to treat obesity and associated metabolic diseases. My objective in this proposal is to identify and characterize an important new mechanism of intestine– white adipose tissue (WAT) communication via intestinal mTOR. I hypothesize that intestinal mTOR crosstalk with WAT browning regulates glucose homeostasis and energy metabolism by modulating the anti-microbial peptide (AMP)-controlled composition of gut microbiota (GM). My hypothesis is based on the following results from our preliminary results: 1) Significantly increased mTOR phosphorylation in the duodenum of patients with obesity and T2D; 2) an enhanced WAT browning and glucose homeostasis in intestinal epithelium-specific mTOR knockout (mTOR-IKO) mice compared to littermate controls (mTORfl/fl); 3) significantly altered gut microbiota composition and gut microbiome-derived metabolites in mTOR-IKO mice; 4) enhanced WAT browning and glucose homeostasis in WT mice after fecal microbiota transplantation (FMT) with feces from mTOR-IKO mice; 5) intestinal mTOR-mediated expression of intestinal anti-microbial peptides (AMPs), including REG3 and RELMb, which directly targets bacteria. I propose two aims to test my hypothesis. In Aim 1, I will continue to determine the role of gut microbiota and gut microbiome-derived metabolites in mediating the effects of intestinal mTOR on WAT browning and metabolic homeostasis. I will perform FMT from mTORfl/fl and mTOR-IKO mice to germ-free mice; in addition, I will identify the candidate metabolites linked to WAT browning phenotype; then I will screen culture collections to identify responsible bacterial strains which can produce the candidate metabolites; Moreover, I will perform the functional validation for the responsible bacterial stains using GF mice. In Aim 2, I will identify the mechanism by which mTOR modulates the gut microbiota profile. My preliminary studies support the hypothesis that intestinal mTOR controls gut microbiota composition through AMPs expression. I will determine gut epithelial differentiation in mTOR-IKO mice compared to mTORfl/fl mice. In addition, I will elucidate the molecular mechanism by which intestinal mTOR regulates the expression of Reg3r, RELBb and other AMPs.

NIH Research Projects · FY 2025 · 2025-09

Project Summary (30 lines): Together with pancreatic beta cells, alpha cells maintain circulating glucose levels within the normal range so that glucose homeostasis can be achieved. In type 1 diabetes (T1D), beta cells are destroyed by chronic autoimmune responses, resulting in severe insulin deficiency. However, the function of alpha cells is also compromised, and it contributes to both hyperglycemia and hypoglycemia. While we recognize that alpha cells are influenced by signals from neighboring beta and delta cells, the orchestration of these interactions remains poorly understood. My preliminary data supports the hypothesis that alpha cells are also functionally heterogenous, and that distinct alpha cell subpopulations may play a role in different regulatory circuits and respond to different paracrine signals. This nuanced regulation is particularly vital for understanding the mechanisms governing alpha cell behavior under both normal and diabetic conditions. Addressing these questions is therefore essential for elucidating the complexities of alpha cell regulation, ultimately paving the way for advancements in diabetes research and potential therapeutic interventions. To address my hypothesis, I will implement a multi-faceted research strategy that leverages innovative methodologies, particularly utilizing human pancreatic tissue slices, a study approach that I contributed to establish. Most studies on human islets have relied on isolated islets or dispersed cell models, which often fail to replicate the native architecture, and intercellular interactions present in pancreatic tissue, limiting our understanding the complex network of alpha cells. Utilizing human pancreatic tissue slices, my project will preserve the islets natural environment, shedding light on their behavior and interactions in ways that isolated studies cannot. I will use viral vector to express reporters for calcium and cAMP signaling and employ live-cell imaging to monitor real-time intracellular responses to various stimulations, both activating and inhibiting alpha cells. In addition, I will perform molecular profiling through single nucleus RNA sequencing and potentially spatial transcriptomics to identify specific gene expression patterns linked to each alpha cell subpopulation. This dual approach will provide a comprehensive analysis of alpha cell heterogeneity and their interactions with neighboring beta and delta cells, addressing a significant knowledge gap in diabetes research. By uncovering the nuances of alpha cell regulation and their role in glucose homeostasis, this project is relevant to support the development of targeted therapies that can improve diabetes management by restoring alpha cell function. Starting with my PhD studies I have focused on investigating human islet cells in health and diabetes, and I have developed a focus on alpha cells during my post-doctoral fellowship. The proposal developed for this K01 application is an excellent vehicle to complete my training and prepare me to undertake an independent career focus on the study of pancreatic alpha cell biology.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY/ABSTRACT Enhancing adipose tissue thermogenesis to tilt energy balance toward net expenditure has great potential to promote weight loss and treat metabolic milieus in humans. Cold exposure and β3-adrenergic receptor (AR) agonist treatment are commonly used to increase adipose tissue thermogenesis and beiging in mice. However, cold exposure-induced adipose tissue beiging predominantly recruits new beige adipocytes, while β3-AR agonists primarily promote white adipocytes to convert to beige adipocytes. Given a species difference between mice and humans in adipocyte β3-AR expression, β3-AR agonists that had remarkable efficacy in mice in promoting lipolysis and enhancing energy expenditure failed to translate into persistent weight loss and improved metabolic function in humans. So, novel approaches to promote the recruiting of new thermogenic beige adipocytes in humans are particularly interesting for developing new drugs to enhance adipose tissue thermogenesis. Our preliminary data suggest hyaluronan (HA), a polysaccharide secreted by cells, may play a crucial role in cold-induced de novo beige adipogenesis. Using several new mouse models we have generated, we showed that HA secreted within the subcutaneous white adipose tissue enhanced cold-induced adipose tissue beiging, and digestion of HA significantly reduced this process. In vitro experiments suggest HA plays a vital role in adipose progenitor cell (APC) proliferation and differentiation into beige adipocytes. Importantly, we found that aged mice had much lower HA levels in the subcutaneous white adipose tissue, which may highly correlate with the age-related decrease of white adipose tissue beiging and thermogenesis. Several critical links are still missing: A. How does cold stimulate APC to produce HA? B. Does HA directly enhance de novo beige adipogenesis in mice? C. Can we target HA to enhance thermogenesis and improve metabolic health in aged individuals? To answer these questions, we propose to determine whether cold exposure promotes APC HA production through an Adrb1-Has1 axis (Aim 1) and whether APC HA production promotes de novo beige adipogenesis and adipose tissue beiging (Aim 2). We will also determine the metabolic effects of HA-mediated APC proliferation and adipose tissue beiging in diet-induced obese and aged mice (Aim 3). Altogether, the proposed study will provide novel insights into how APC HA production mediates de novo beige adipogenesis and how it could be targeted to treat metabolic diseases.

NIH Research Projects · FY 2025 · 2025-09

Project Summary: Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive malignancy and is rapidly becoming the 2nd leading cause of cancer-related deaths. The dismal 5-year survival rate of ~10% (the lowest of any major cancer) essentially stems from the fact that PDAC is diagnosed at late stages. Achieving early detection is crucial, offering a pivotal shift in staging that holds the promise of improved survival rates. Previous attempts to develop PDAC early detection tests have suffered from flaws in their study design, including 1) failure to enroll early-stage PDAC cases, 2) assuming that a biomarker developed in metastatic settings would also detect early-stage PDAC, or 3) assuming that a tissue-derived biomarker would also function in the blood. Therefore, we will develop and validate a bloodborne test in a cohort of patients enriched for early-stage PDAC. We will use the PLCO population to evaluate the lead time to PDAC diagnosis on pre-diagnosis biospecimens. Our test will be minimally invasive, highly sensitive, and cost-effective by harnessing the potential of 5- methylcytosine (5mC) and 5-hydroxy methylcytosine (5hmC) modifications on circulating cell-free DNA. Importantly, commercially available tests based on 5mC demonstrated a high sensitivity for PDAC while a 5hmC- based test demonstrated specificity (but suboptimal sensitivity). Because 5hmC modifications occur genome- wide and are abundant, they are biologically sensitive biomarkers, while 5hmC modifications are typically gene- targeted and display >90% cancer- and stage-specificity. Therefore, a combination of the two approaches, one contributing sensitivity (5mC) and the other contributing specificity (5hmC) is desirable, innovative, and necessary to optimize the early detection of PDAC. Our goal is to develop a robust assay for detecting early- stage PDAC. Our design strategy will involve three aims. First, we will discover aberrantly methylated loci potentially diagnostic for PDAC with 5mC- and 5hmC-profiling in circulating cfDNA and matching tissues. Second, we will develop 5mC and 5hmC-based diagnostic biomarker signatures that discriminate patients with PDAC from controls using multiple advanced machine learning and statistical approaches. Here, we will train two machine learning algorithms—one for 5mC and another for 5hmC—to identify patients with PDAC based on qPCR results and then merge these two algorithms to create a detection signature for PDAC. Then, we will validate the 5mC, 5hmC, and 5mC+hmC qPCR assays in a large, cross-sectional cohort of biobanked samples from patients with PDAC vs. controls. Third, we will perform the 5mC, 5hmC, and 5mC+5hmC qPCR assays in prospectively collected samples from the PLCO study to determine how our assays may provide a positive PDAC signal much earlier than clinical diagnosis. In summary, this project aims to develop a highly sensitive, cost- effective liquid biopsy for early detection of PDAC. If successful, this proposal will involve several innovations and will have the potential to transform clinical practice.

NIH Research Projects · FY 2025 · 2025-09

Project Summary 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. Multiple lines of evidence have indicated that 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. Regrettably, there has been no in vivo studies about the impact of Xbp1s overexpression on β-cell survival and function. Using a novel mouse model that allows inducible overexpression of Xbp1s selectively in β-cells, we found a striking impact of Xbp1s on β-cell function and survival. Preliminary studies showed that Xbp1s acts as a regulator of β-cell heterogeneity by facilitating reversible state transitions. A novel link of Xbp1s and NeuroD1 was predicted by the multiome data which highlights a potential mechanism for Xbp1s-faciliated state transition. Moreover, the dynamic state transitions of β-cells were halted in obese mice, indicating metabolic stress could adversely affect the state transition facilitated by Xbp1s. Human islets and cell lines will be employed to complement in vivo animal models at the mechanistic level. 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 · 2025-09

PROJECT SUMMARY B-cell acute lymphoblastic leukemia (B-ALL) is the most common pediatric cancer and remains a leading cause of childhood cancer death. Over 10% of B-ALL cases have point mutations in PAX5, a gene that encodes a key transcription factor for B-cell development. PAX5 mutations are the signature lesions in two recently identified B-ALL subtypes: PAX5alt, characterized by diverse genetic alterations in PAX5, and a subtype defined by PAX5 P80R mutation. While PAX5alt patients, particularly adults, have a significantly worse prognosis compared to PAX5 P80R patients, the underlying mechanisms driving these differences remain poorly understood. Both subtypes frequently acquire additional genetic alterations, such as wild-type PAX5 allele deletion and Ras pathway mutations, which further contribute to disease progression. Despite advances in chemotherapy, many patients experience suboptimal outcomes and poor quality of life, emphasizing the need for improved therapies. Our preliminary studies show that PAX5 P80R leukemic cells have increased sensitivity to Dexamethasone (Dex) treatment, likely due to high dependence on glucose uptake and glycolytic activity. These findings suggest that metabolic alterations contribute to the differential clinical responses observed between the two PAX5-driven subtypes. To systematically dissect these mechanisms, we will use orthogonal experimental models (e.g., B- ALL cell lines, mouse leukemia models, and human B-ALL samples) alongside multi-omics platforms, such as RNA-seq, CUT&Tag, ATAC-seq, END-seq, whole-genome sequencing, and single-cell analyses in this project. We hypothesize that rather than loss of function, PAX5 mutants can exert dominant effects in B-ALL initiation through novel PAX5 binding activities and gene transcription activation, which may block normal B-cell differentiation and give rise to pre-leukemic cells; once additional genetic lesions are acquired, PAX5-mutant clones can progress to overt B-ALL. We aim to (Aim 1) identify the transcriptional and metabolic alterations induced by PAX5 mutants, particularly their different effects on Dex treatment. Additionally, we will (Aim 2) investigate the stepwise acquisition of genetic lesions that drive leukemic transformation, with a focus on the pre-leukemic cells and their transition to overt leukemia using the Pax5 P80R mouse model. Finally, we will (Aim 3) study the role of aMEGF10, a gene significantly overexpressed in PAX5 P80R B-ALL, and assess the therapeutic potential of targeting the novel aMEGF10-SYK signaling pathway. In summary, this project will provide comprehensive insights into the molecular features of PAX5-mutant B-ALL subtypes, reveal novel mechanisms of leukemogenesis and treatment responses, and identify potential therapeutic targets. These findings are expected to improve patient risk-stratification and lead to the development of more effective targeted therapies with lower toxicity for PAX5-mutant B-ALL.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY Type 2 diabetes (T2D)-associated vascular dysfunction and inflammation drives the development of several debilitating microvascular complications (MVC, e.g., retinopathy and nephropathy). Despite various treatment options, morbidity/mortality from MVC is augmented. Thus, there is a critical unmet need for in-depth examination of the underlying mechanisms that can lead to improved therapies. Increased activation of monocytes and macrophages (Monos/Macs) instigated by diabetogenic factors (i.e., high glucose, cytokines, lipids) play vital roles in inflammatory diabetic MVC. Our studies have demonstrated the roles of epigenetic mechanisms in modulating inflammatory genes/processes/functions in mouse (m) and human (h) Monos/Macs under diabetic conditions and in metabolic memory. Our long-term goal is to understand how diabetes-induced changes in key nuclear factors control specific epigenotypes and long-range chromosomal interactions to reprogram the 3D epigenome and regulate inflammatory gene expression. Our exciting new data shows that; i) T2D h- and m- Monos/Macs, as well as T2D m-kidney/adipose tissues are deficient in JARID2 (JD2), a DNA binding and chromatin remodeling protein; ii) in h-Macs, JD2 knockdown (JD2-KD) activates, while JD2 overexpression (JD2- OE) attenuates inflammatory responses; iii) RNA-seq analyses after JD2-KD revealed that JD2 target genes are associated with Macs dysfunction in MVC; iv) integrative epigenomics analysis in h-Macs revealed that diabetic conditions (and reduced JD2) dysregulate chromatin accessibility and the enhancer connectome at inflammatory gene loci and JD2 target genes; v) novel lipid nanoparticles (LNPs) are viable for JD2 delivery to Macs and target organs in vivo in T2D mice, and reduce inflammatory genes and MVC (nephropathy). The rationale is that an in- depth understanding of the anti-inflammatory roles of JD2 and its pivotal regulation of epigenome and chromatin organization can lay the foundation for novel therapeutic strategies to curb inflammation and related MVC in T2D. Our central hypothesis is that downregulation of JD2 in T2D reprograms the epigenome and the 3D chromatin architecture in Monos/Macs, and leads to the hyperinflammatory state that drives T2D-related MVC. These adverse changes are ameliorated by JD2 reconstitution. This will be tested via 3 Specific Aims: 1) Determine the functional roles of JD2 in diabetes-induced dysregulated phenotype of Monos/Macs associated with MVC; 2) Elucidate the role of JD2 in diabetes-reprogrammed epigenome and 3D chromatin architecture driving h-Monos/Macs dysfunction; 3) Determine the therapeutic potential of augmenting JD2 with LNPs in vivo to ameliorate Macs dysfunction and MVC in diabetes. This innovative study will synergize expertise from chromatin biology, epigenetics, vascular biology, Omics analysis, vascular immunology and nanomedicine to tackle the unmet need for MVC management in T2D. These results will provide unique insights into the 3D epigenomic regulation of inflammation in T2D MVCs, and pioneer mRNA-LNP therapeutics in treating MVC. This study will also positively impact efforts to improve the quality of life of patients with T2D.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY Philadelphia chromosome (Ph)-positive and Ph-like B-cell acute lymphoblastic leukemia (B-ALL) are high-risk leukemia subtypes with poor long-term survival rates. Despite advances in treatment, patients with these subtypes continue to have survival rates below 50%. Our research aims to address this urgent need by focusing on a novel hallmark gene, AL021978.1 (hereafter named A978), which is identified as the most significantly overexpressed gene in Ph/Ph-like B-ALL. Our preliminary data indicate that A978 encodes a short protein located in mitochondria, promoting leukemic cell proliferation, and contributing to resistance against tyrosine kinase inhibitors (TKIs). The long-term objective of this project is to improve treatment outcomes for patients with Ph/Ph-like B-ALL. Our specific aims are: 1) to determine the biological effects of A978 in leukemic cells; 2) to study the effects and functions of A978 in mitochondria and metabolic activities; and 3) to evaluate the therapeutic impact of pharmacologically inhibiting A978 in Ph B-ALL. To achieve these aims, we will use a combination of in vitro and in vivo models, including B-ALL cell lines and patient-derived xenograft models. We will employ a variety of techniques, such as RNA sequencing (RNA- seq), single-cell RNA-seq, BioID and mass spectrometry, and metabolomic profiling, to explore the molecular functions and biological pathways regulated by A978. Additionally, we will develop and evaluate a therapeutic strategy using a CpG-conjugated small interfering RNA to specifically silence A978 expression in leukemic cells. The significance of this project lies in its potential to uncover new mechanisms driving the proliferation and progression of high-risk B-ALL subtypes. By targeting A978, we aim to develop novel therapeutic strategies that can be combined with existing treatments to enhance their efficacy. This research is expected to largely advance our understanding of Ph/Ph-like B-ALL and contribute to the development of more effective treatments, ultimately improving survival rates for patients with these high-risk leukemias.