University Of California, San Francisco

NIH Research Projects · FY 2025 · 2025-09

Overall Summary The University of California at San Francisco Diabetes Research Center (DRC) will function as a basic and clinical research enterprise at the forefront of type 1 diabetes (T1D) research. The DRC crosses the boundaries of Departments and Schools to support a highly interactive team investigating all forms of diabetes to advance the understanding and treatment of the disease. The DRC integrates the research activities of the UCSF Diabetes Center, the administrative home of the DRC, with a broad range of diabetes-related research throughout UCSF. The combined intellectual and research expertise of the DRC encompasses 43 diabetes investigators from UCSF, UC Berkeley, and UC Davis, focused on both basic and clinical research, supported with $22M in annual direct costs in diabetes-related grant funding, including $12M in Federal funding. In this application, we propose to continue the success of the UCSF DRC in promoting, enhancing and integrating T1D research at UCSF. That goal is realized through support for the following activities: Administrative Core provides leadership, infrastructure, administrative support, advice and oversight to the other components and members of the DRC; regularly adapts Cores and programs to fit the needs of the members; and interfaces with the NIDDK, other NIH Diabetes Centers, and the lay community. Enrichment Program organized by the Administrative Core supports DRC members and the regional diabetes research community by enhancing scientific exchange and training with seminar series, invited speakers, and an annual retreat. Biomedical Medical Research Cores Islet Core: Provides mouse and human islets and technologies for working with those islets, together with expertise, advice and training. Flow and Mass Cytometry Core: Provides equipment, assistance and training for human and animal cell analyses. Microscopy Core: Provides advanced imaging equipment, technologies, advice and training. Pilot & Feasibility Program funding small innovative projects that support new investigators, attract new investigators to diabetes and the DRC, explore new questions by established investigators, and promotes interactions and collaborations leading to new external funding support to further goals of the DRC.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY Rac1, a member of the Rho GTPase subfamily, regulates essential cellular functions including cytoskeletal arrangement, cell motility, cell cycle progression, and proliferation. Dysregulation of Rac1, such as through the oncogenic P29S mutation or the expression of the alternative splice variant Rac1b, is common in cancer and contributes to tumor progression, metastasis, and drug resistance. While Rac1 gain-of-function mutations, amplification, or increased activity of activating proteins are necessary for driving tumorigenesis, the Rac1b variant alone has also been shown to promote oncogenicity. While Rac1 and Rac1b differ by only a 19- amino acid insertion after Rac1b's Switch II pocket, this alteration changes the Switch I and II dynamics of Rac1b, resulting in changes in cellular activity and promotion of tumorigenesis. Furthermore, distinguishing the roles of these variants in cellular signaling and disease is challenging due to their sequence and structural similarity, coupled with the scarcity of selective chemical tools for effectively targeting these proteins in cellular systems. Current inhibitors of Rac1 reversibly target the conserved GEF binding pockets or compete with GDP/GTP for the conserved Switch I/II region, resulting in compounds with low potency and lack of specificity to Rac1. Given the differences in the dynamics of the Switch I and II pockets among Rac1 variants, I hypothesize that targeting these regions will lead to the development of Rac1 variant selective inhibitors. My preliminary data demonstrates that Switch II pocket inhibitors can exhibit selectivity for Rac1 isoforms, aiding in understanding their distinct roles in cancer progression. For Aim 1, I will utilize structure-based design to optimizing selective and cellular active Switch II pocket inhibitors towards Rac1/Rac1b G12C. To assess the potency and selectivity of these compounds in a cellular system, I will introduce the Rac1/Rac1b G12C mutation in immortalized cancer cell lines via CRISPR Prime editing. To create therapeutics towards oncogenic Rac1 variants, Aim 2 of this proposal focuses on developing covalent inhibitors targeting the conserved cysteine in the Switch I pocket of Rac1 GTPases. This strategy will exploit Rac1 P29S and Rac1b fast-cycling phenotype and low affinity for GTP to create inhibitors that outcompete GTP for binding the Switch I pocket. In conjunction with the Arkin lab, I have identified fragments from a disulfide tethering screen with affinity for the Switch I pocket, which will be optimized for binding Rac1 P29S and Rac1b. Since Rac1 P29S and Rac1b have different Switch I conformations and lower affinity for GTP than Rac1 WT, I hypothesize that these identified covalent Switch I pocket binders will preferentially inhibit GTP binding of these oncogenic variants over Rac1 WT. This project aims to create selective chemical tools to elucidate Rac1 isoforms’ role in oncogenesis and provide therapeutic inhibitors for selectively targeting oncogenic Rac1 variants in cancers. By continuing my training in integrating chemical genetics and cell biological techniques and leveraging the expertise and collaborative environment provided in the Shokat lab, I have the tools to successfully complete this project.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY/ABSTRACT Tumor-reactive CD8 T cells become exhausted and lose their capacity to kill cancer cells in the tumor microenvironment. Progenitor-exhausted CD8 T cells (Tpex) found primarily in tumor-draining lymph nodes retain their stem-like properties and respond to immune checkpoint blockade. However, our group’s recent work suggests that Tpex responses to ICB are disrupted in the metastatic lymph nodes of head and neck cancer patients. Mass cytometry showed that fewer Tpex are present in metLN; spatial proteomics also revealed that Tpex were less likely to co-localize with their differentiated, cytotoxic counterparts and more likely to be surrounded by immunosuppressive CD4 T and dendritic cells (DC). Furthermore, in a retrospective analysis of patients with recurrent head and neck cancer, we found that those with metLN were less likely to respond to ICB. To investigate the biology underlying the interruption of Tpex responses to ICB in metLN, we will use the metastatic 4MOSC1 (ICB responsive) and 4MOSC2 (ICB non-responsive) orthotopic mouse models of head and neck cancer. In our preliminary studies, we have identified immune cell subsets, including endogenous Tpex, in the tdLN of 4MOSC1 tumors and have established workflows for the isolation and co-culture of T cells and DC from individual tdLN in vitro. This proposal will use the 4MOSC1/2 mouse model of head and neck cancer to evaluate mechanisms by which Tpex responses to ICB are disrupted in metLN. Aim 1 will interrogate whether defects in the S1P/S1PR1 lymphocyte migration axis in metLN affect Tpex responses to ICB. Aim 2 will evaluate whether activation of Tpex in metLN confers cell-intrinsic defects that affect their capacity to differentiate, survive, and signal through their T cell receptor. Aim 3 will identify mechanisms by which metastases in tdLN affect the polarization of CD4 T cells and activation of DC proximal to Tpex. Aims 1 and 2 will use well-established in vivo and in vitro methods; Aim 3 will use spatial proteomic and transcriptomic tools. These studies will be the first to mechanistically interrogate how early tumor-reactive CD8 T cell responses are impeded in metLN. Uncovering this biology has the potential to elucidate novel strategies for potentiating immunotherapy responses in patients with advanced, metastatic disease in several cancer types. This research project and fellowship training will be conducted at a top-funded research institution, the University of California, San Francisco (UCSF), in the laboratory of Dr. Matthew Spitzer with expert mentorship from Dr. Jason Cyster and Dr. Karin Pelka. Dr. Spitzer is an expert in CD8 T cell and lymph node biology, systems immunology, and high-dimensional single cell analyses. Dr. Cyster is a world-class scientist who discovered the lymphocyte trafficking mechanism explored in Aim 1. Dr. Pelka is a cancer immunologist with expertise in the spatial biological tools to be utilized in Aim 3. Overall, this institution and team provide a rich training environment for completion of this research and the development of professional skills necessary for a career in academic research.

NIH Research Projects · FY 2025 · 2025-09

ABSTRACT Breast cancer (BC) is a heritable disease with 10 known high- and intermediate-penetrance genes and several hundred common genetic variants associated with risk to develop disease. Most of the genes known to affect BC risk, such as BRCA1, BRCA2 and PALB2, are part of the homologous recombination repair (HRR) pathway. However, there are still additional genes to be identified that increase risk, and we hypothesize that some of these additional genes are in the HRR pathway. Moreover, drugs that inhibit poly-adenosine diphosphate-ribose polymerase (PARP) and platinum-based chemotherapy are extremely effective in patients with pathogenic genetic variants that cause HRR deficiency. Therefore, identifying other genes in this pathway has important implications for genetic cancer risk assessment for prevention, early detection, and treatment. Hispanic/Latina (H/L) women and other minority populations are limited in participating in genetic studies. We recently completed a large sequencing study of breast cancer among H/L women that included over 4,000 cases and 4,000 controls. In that study, we identified a strong association between loss-of-function (LoF) variants in FANCM and estrogen-receptor (ER)-negative breast cancer, as well as with known breast cancer genes. FANCM is involved in initiating the HRR pathway. We also identified suggestive associations with other genes in HRR and breast cancer risk including ATR. Therefore, we propose to focus on genes in HRR in H/L women using a larger dataset. Our objective is to improve genetic risk assessment for breast cancer in H/L women. In Aim 1, we will discover additional variants in HRR genes associated with BC in H/L women, expanding our numbers to include ~12,000 H/L breast cancer cases and ~30,000 controls. We will use a novel deep-learning approach to classify missense variants in order to improve power for discovery. In Aim 2, we will characterize the effects of LoF variants in FANCM and other novel HRR genes on the somatic mutational landscape in tumors. Using cell line models, we will measure the effects of LoF variants on DNA repair and on treatment response to PARP inhibitors and platinum-based therapies. In addition, we will evaluate the LoF variants in FANCM for their effects in repair of stalled replication forks and protection of common fragile sites. In Aim 3, we will assess the joint association of breast cancer polygenic risk scores (PRS) and pathogenic variants in breast cancer susceptibility genes and risk of developing breast cancer. Impact: H/L women have poorer outcomes from breast cancer compared to Non-H/L White women yet are under-studied for genomics of breast cancer. This exacerbates disparities and increases mortality rates because there are insufficient data for identifying many women at the highest risk of breast cancer. Insufficient data also limit prevention efforts and the identification of women who may benefit from targeted therapy once they develop breast cancer. Our study will provide much-needed data to improve risk prediction and provide tools for clinicians to detect BC earlier. Our findings also may be important to guide therapy for women carrying HRR gene variants.

NIH Research Projects · FY 2025 · 2025-09

Project Summary/Abstract The overall goal of my K23 award research program is to adapt an existing effective anti-stigma intervention to improve communications between US pharmacy staff and people seeking PrEP. Communications from healthcare professionals that lead to experiences of stigma reduce initiation and ongoing use of pre-exposure prophylaxis (PrEP). These communications are a form of stigma, or experiences of judgement that are inextricably linked to multiple aspects of an individual’s personal characteristics. Existing anti-stigma interventions are effective but do not focus on US contexts, pharmacy settings, or acknowledge multiple overlapping experiences of stigma. During this K23 award, in Aim 1 I will qualitatively ask pharmacists, pharmacy technicians, and people seeking PrEP what their experiences and preferences are for non-judgmental communication while obtaining PrEP in a pharmacy, with special attention to experiences of multiple overlapping stigma experiences and relying on the implementation science framework Consolidated Framework for Implementation Research. In Aim 2 I will use the Assessment, Decision, Adaptation, Production, Topical experts-Integration, Training, and Testing (ADAPT-ITT) framework to systematically adapt an existing effective anti-stigma intervention to develop PRISM-P (PrEP-Related Intervention for Stigma Mitigation among Pharmacy staff). In Aim 3, I will pilot PRISM-P among pharmacy staff in Northern California independent pharmacies in Ending Human Immunodeficiency Virus (HIV) Epidemic jurisdictions, to understand whether pharmacy staff find the intervention to be feasible and acceptable, and if participants have decreased incorrect beliefs towards PrEP-seekers. I will record pharmacy staff interactions with simulated patients seeking PrEP, to describe changing pharmacy communications from baseline and post-intervention. To achieve the proposed research aims and to transition to career independence, I will require significant training and mentorship in (1) knowledge of pharmacy interventions, stigma and their applications in HIV prevention and, (2) expertise in personalized communication in healthcare, and (3) expertise in applied implementation science methods. An experienced mentor team will guide my research and training. Dr. Saberi (co-primary mentor) is a pharmacist with HIV prevention expertise. Dr. Seidman (co-primary mentor) is an obstetrician-gynecologist and reproductive infectious disease specialist, with expertise person-centered care. Dr. Bauermeister (co-mentor), has expertise in stigma theories and HIV prevention research. Dr. Chang (co-mentor) is an expert in healthcare communication research and skills training. Dr. Liu (co-mentor) is an expert in implementation science in HIV prevention trials. Dr. Steward (consultant) is an expert in implementation science theories and frameworks. Guided by this excellent team, the training and research plan in this K23 award will ultimately support an R-level proposal of a type 1 hybrid effectiveness-implementation trial to test PRISM-P’s ability to reduce stigma and assess participants’ adherence.

NIH Research Projects · FY 2025 · 2025-09

Project Summary/Abstract Interstitial lung diseases (ILD) are a group of chronic lung diseases characterized by progressive fibrosis and loss of lung function over time. Prior research has appropriately focused on respiratory function and attenuation of disease progression; however, the myriad impacts of ILD on health beyond the respiratory system are less well studied. Reflecting these insights, the American Thoracic Society has emphasized the need to incorporate patient-centered outcomes (PCOs) in ILD research. Patients with ILD suffer from lower quality of life, increased functional limitation, and increased frailty, but the extrapulmonary drivers of the impacts are not well defined. My early findings have identified body composition as a novel risk factor for poor outcomes in advanced ILD. Obesity – a pathologic state of elevated fat mass – and sarcopenia – a pathologic state of low muscle mass and function, have been associated with worse functional status, reduced quality of life, and increased disability. As age increases, for a given body mass index (BMI), people experience an increase in adiposity and decrease in muscle mass and function. At their extremes, this parallel change is termed sarcopenic obesity, which appears to have multiplicative impacts on health. These aging-related changes in body composition, and the limitation of BMI to identify them, are relevant in ILD which increases in prevalence with age. The study of body composition in ILD is nascent, and limited evidence suggests both sarcopenia and obesity are associated with reduced quality of life and exercise capacity, akin to the associations better established in chronic obstructive pulmonary disease. Secondary cross-sectional analyses of two prospective cohort studies (R01 HL134851 [completed]; U01 HL163242 [ongoing]) are proposed to define the association between body composition and PCOs in adults with advanced ILD undergoing lung transplantation evaluation. My central hypotheses are that abnormalities in body composition will be associated with worse PCOs, that the magnitudes of association will be largest in participants with sarcopenic obesity, and that these associations will not be apparent when using BMI as a primary exposure variable. A projected 872 participants with baseline pre-transplantation visits will meet inclusion and exclusion criteria and will be categorized by bioelectrical impedance as non-obese/non-sarcopenic, obese by fat mass, sarcopenic, and sarcopenic obese. The outcomes of interest will be frailty by the Short Physical Performance Battery and Fried Frailty Phenotype, exercise capacity by six-minute walk distance, disability by the Lung- Transplant Valued Life Activities Scale, and general and respiratory-specific health-related quality of life by the Short Form 12 and Airways Questionnaire 20-Revised, respectively. This proposal is significant by exploring a novel and potentially intervenable risk factor for poor PCOs in a population already vulnerable to significant morbidity, while also supporting my growth as an early-stage investigator through strong cross-disciplinary mentorship alongside didactic and experiential training.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY Pancreatic islet transplantation is a therapeutic option for type 1 diabetes (T1D), providing glycemic control and eliminating the need for insulin injections. It offers the advantage of a less invasive, catheter-assisted procedure compared to another surgical intervention, whole pancreas transplantation, which carries high surgical complication risks. However, the long-term success of islet transplantation is limited, with the insulin independence rate dropping to ~10% of patients after five years. This is in contrast to whole pancreas transplantation, which achieves higher insulin independence rates of >70% at five years post-transplant. The key difference lies in the loss of the islet microenvironment during isolation for islet transplantation; disrupting vasculature and extracellular matrix support leads to significant graft attrition. This underscores the need for creating an islet-supportive microenvironment. Fabricating an islet organoid in vitro-with interactions between islets and supporting cells prior to transplantation-is a promising solution. However, fabricating a clinically scalable organoid has been challenging due to the insufficient oxygen supply to the organoid core. Our longterm goal is to develop clinical-scale transplantable islet organoids that incorporate an islet-supportive microenvironment to improve islet engraftment efficiency in T1 D patients. In this application, we propose a practical approach to deliver oxygen to the diffusion-limited organoid core. We engineered oxygen-transporting microcapillary mesh made from clinically proven parylene material. Oxygen is delivered through the ultra-thin 25-μm microcapillary mesh via passive diffusion from the surrounding environment. This achieves a uniform and self-sustaining oxygenation throughout the organoid. We hypothesize that establishing an islet-supportive microenvironment within an islet organoid, prior to transplantation, will improve post-transplant engraftment. We will test this through the following Specific Aims: Aim 1: Fabricate scalable human islet organoids using oxygen transport from culture media in a dynamic culture system. Aim 2: Fabricate scalable human islet organoids using oxygen transport from ambient air in a static culture system. We will fabricate human islet organoids of 5 mm in thickness that far exceed the general oxygen diffusion limit of ~50-100 μm. The organoids will be integrated with multiple layers of our oxygen-transporting mesh to deliver continuous oxygen to the core. We will use human cadaveric and stem cell-derived islets as potential insulin-producing cell sources and transplant these organoids under the skin in diabetic immunodeficient animals to assess the therapeutic benefits compared to conventional islet transplantation. These aims share the same concept-delivering oxygen into the organoid core through the oxygen-transporting mesh-but employ distinct oxygen sources with mesh modifications. Successful completion will demonstrate a scalable solution in large organoid fabrication. This research aligns with the mission of the NIDDK by integrating cross-cutting science in biology, physiology, and engineering using a unique device to address a critical gap in T1D treatment, with implications for multiple organoid-based therapies.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY/ABSTRACT Endometriosis is a disorder of ectopic endometrium-like tissue that affects 10% of reproductive-age people with a uterus. Despite its prevalence, it is difficult to diagnose, taking an average of 4-11 years for definitive diagnosis, and has a high rate of recurrence after hormonal and surgical therapies. The humoral immune response in endometriosis has the potential to shed insight into disease pathophysiology and diagnostic biomarkers. However, our understanding of the role antibodies play in endometriosis is largely limited to the knowledge that up to 50% of patients have anti-endometrial autoantibodies, and that a subset of infertility patients with endometriosis have autoantibodies against laminin-111 that contribute to early miscarriage. In addition, there is evidence that altered extracellular matrix (ECM) protein expression and remodeling plays a pathogenic role in endometriosis. Preliminary data suggest that patients with endometriosis have an enrichment of autoantibodies that target the ECM compared to patients with other uterine disorders. The long-term goal of this work is to identify autoantibodies that are specific to endometriosis that will serve as non-invasive diagnostics, and to examine the functional impact of ECM-targeting autoantibodies. This will be accomplished in Aim 1 by characterizing the specific reactivity of autoantibodies to ECM proteins expressed in endometriotic lesions and interrogating their functional consequences in cellular invasion assays using endometriotic spheroids. In Aim 2, unbiased serological antibody profiling and machine learning will be leveraged to identify a minimal set of endometriosis-associated autoantibody targets. This will provide a concrete list of biomarkers that can be leveraged as a non-surgical diagnostic and will be validated using an independent cohort. These experiments will accelerate the development of methods to definitively diagnose, prevent, and ultimately treat endometriosis. Under this fellowship, the applicant will be supported by extraordinary clinical and research training in the University of California San Francisco’s MD/PhD program. A mentorship team consisting of experts in the reproductive sciences, immunology, and computational methods will support the applicant in achieving her goal of becoming a physician-scientist gynecologist who studies the intersection of reproductive health disorders and dysregulated immunity.

NIH Research Projects · FY 2025 · 2025-09

SUMMARY/ABSTRACT We first discovered that the rate of sudden cardiac death (SCD), the most feared manifestation of cardiovascular disease, is substantially higher in people with HIV (PWH). We established the POstmortem Systematic InvesTigation of Sudden Cardiac Death (POST SCD) study, a prospective medical examiner-based cohort using autopsy to refine presumed SCDs to true cardiac causes, with banked tissue and data on PWH and uninfected control cases. ~80% of HIV+ SCDs were on ART, thus this a one-of-a-kind resource to study the direct myocardial tissue effects of treated HIV. HIV POST SCD showed that the increased SCD risk in PWH is attributable to higher levels of myocardial fibrosis, a known substrate for fatal arrhythmias, and our updated incidence analysis confirms a significant, 2-fold higher rate of arrhythmic death in PWH (IRR 2.001, 95% CI 1.02- 3.93, p=0.044). Yet, the mechanisms by which HIV leads to myocardial fibrosis or other tissue processes to cause non-CAD SCD are poorly understood. We have generated expression, immunohistochemical, and viral persistence data in myocardial tissue sampled at the time of SCD from 20 PWH, matched to >40 HIV- control SCDs that demonstrate: (1) hearts from PWH, most on ART, exhibit higher immune activation and cardiac ion channel dysregulation; (2) PWH on ART have downregulated expression of acute heart failure (HF) genes, suggesting that SCD in PWH is less related to HF and may be due to a distinct arrhythmogenic substrate or other inflammatory process; and, (3) the level of myocardial immune upregulation in PWH on ART is comparable to the highly inflammatory myocardial state triggered by traumatic injury. Via regional digital spatial profiling, we demonstrate the first direct tissue evidence of HIV RNA in myocardial-resident macrophages (MΦ) and that HIV may preferentially affect the epicardium, a known frequent source of VT/VF triggers, in the ART-suppressed heart, suggesting a regional myocardial specificity to the effects of HIV. Our central hypothesis is that HIV- induced MΦ activation in myocardium of PWH on ART, with particular augmentation in the arrhythmogenic epicardium, leads to chronic cardiac inflammation, interstitial fibrosis, ion channel dysregulation, and disruption of normal electrical coupling. Together, these process exert a direct myocardial tissue effect beyond the vascular space, to ultimately result in arrhythmic substrate that underlies increased SCD risk due to fatal arrhythmias in PWH. Our aims are to test the following hypotheses: 1A) myocardium from PWH on ART with SCD vs. HIV- SCDs and trauma controls have a higher burden of HIV-infected or infiltrating/inflammatory MΦ, which in turn correlates with levels of myocardial inflammation, fibrosis, and electrical remodeling; 1B) myocardium from PWH on ART with SCD vs. HIV- SCDs and trauma controls demonstrate chronic immune activation and tissue remodeling specific and distinct to the highly arrhythmogenic and inflammatory epicardial layer, including its adipocytes; and 2) chronic immune activation promotes fibrosis and decreased electrical coupling in specialized conduction tissues from PWH on ART with SCD vs. HIV- SCDs and trauma controls.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY This mentored Ruth L. Kirschstein National Research Service Award will provide the trainee, a PhD student in epidemiology at UCSF, with the training necessary to become a researcher in cancer health disparities. Her training goals for this fellowship are to gain expertise in translational research, geospatial data analysis, and causal inference methods under the guidance of a team of expert mentors. This fellowship will provide the applicant with the skills, content knowledge, and practical experience to launch an independent research career in social epidemiology, focusing on translational research on cancer health disparities. Ovarian cancer is a significant cause of cancer-related deaths among women in the United States, with heterogeneous survival rates across racial and ethnic groups. Considerable health disparities in ovarian cancer outcomes are observed by race and ethnicity, socioeconomic status, and other factors, and are driven by multilevel causes from structural and historical societal processes to risk behaviors and biologic processes. Access to cancer specialists, including spatial accessibility, is essential for improved ovarian cancer outcomes. Addressing limitations of previous research, the proposed study will utilize novel comprehensive measures of spatial medical accessibility and causal inference framework to evaluate the interplay of structural, social, and spatial factors in ovarian cancer survival disparities in California. Building on prior work of the trainee, this F31 uses the data from the statewide population-based California Cancer Registry and publicly available spatial and contextual datasets to: (Aim 1) identify structural and social factors (residential economic and racial and ethnic segregation, racial and ethnic composition, neighborhood socioeconomic status, rurality, and transit access) associated with spatial accessibility of gynecologic oncologists in California; (Aim 2) quantify the extent to which the disparities in ovarian cancer survival in California, that are driven by structural and social factors, are mediated by spatial accessibility of gynecologic oncologists. This evidence will inform policies to increase equitable access to ovarian cancer care and narrow disparities. Knowledge gained from this research will advance the goals of the NCI’s National Cancer Plan to eliminate inequities, deliver optimal care, and maximize data utility. The major strengths of this proposal are 1) the use of population-based California Cancer Registry data with which the applicant has experience working and which represents a region with a diverse landscape, local economies, and racial and ethnic composition; 2) the use of the state-of-the-art geospatial method for the development of the individual-level spatial medical accessibility measure; 3) the application of multilevel health disparities framework and rigorous causal inference methods for studying modifiable contributors of ovarian cancer disparities. The interdisciplinary training environment and the expert mentorship team will provide the applicant with an excellent training opportunity to develop methodological and content expertise for a future career as an independent researcher of cancer health disparities.

NIH Research Projects · FY 2025 · 2025-09

Abstract Adverse patient safety event rates remain stubbornly high in hospitals. For children, adverse events are 1.5-2 times more common than in adult inpatients (40.0 vs. 25.1 harms/100 admissions). Family members and patients are often close observers of care, focusing on only one patient and one set of diagnoses. Text messaging and mobile phone applications technologies offer the opportunity to gather patients’ and families’ safety reports in real-time, addressing limitations of prior work. The PI and team co-designed a mobile phone- based approach, the Family Input for Quality and Safety (FIQS) with families and clinicians. They tested it across 3 local hospitals and 9 units, successfully engaging family members, patients, staff, and safety and quality leadership, and leading to safety improvement projects. The objectives of this proposal are to address key outstanding dissemination and implementation questions, focusing on minimizing sign-up burden, supporting ongoing participant engagement, and enhancing related safety efforts, informed by the CFIR framework. Aim 1: In a cluster-randomized study (n=5645, 15 units), assess the effect of two sign-up strategies (text only, text with in-person orientation) on reporting rates, and interactions by race, ethnicity, & language. Hypothesis: Rates of enrollment and safety-oriented reporting will differ between strategies. Aim 2: In a 1:1 randomized trial, test the effect of two engagement strategies on patient and family safety reporting rates, and interactions by race, ethnicity, & language (n=3386). Group 1 participants: Reports will be shared with units without identifiers (unless requested). Group 2 participants: All reports are shared with units with identifiers. Hypothesis: Reporting rates will be higher for those whose reports are not identified, who access the website, and for whom there is service recovery; results may vary by race, ethnicity, or language. Aim 3: Using mixed methods, evaluate barriers and facilitators to the successful integration of FIQS data into safety efforts. Over an 18-month implementation period, in 21 units, assess inner drivers of uptake & outcomes. Quantitative outcomes: # of units integrating FIQS report reviews into existing safety workflows; numbers of FIQS reports reviewed, # of system-level interventions undertaken and completed. Qualitative data: characterization of interventions made in response to FIQS reports; barriers and facilitators of successful integration of FIQS reports into safety. The proposed research is innovative in its paradigm-shifting conceptual model of 1) its use of mobile phone technology for real time safety reporting, with highly feasible patient-level randomization, and 2) text-message based opt-out approach, to engage patients and caregivers to share safety observations, moving prior evidence into real-world implementation. The contribution of the research will be to answer key dissemination and implementation questions about using patient-facing digital technology to improve patient safety, in partnership with unit leaders and families. These contributions will be significant because they are key to implement and evaluate a potential new approach to improving inpatient safety.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY Meningiomas comprise 40% of primary intracranial tumors and approximately 1% of humans will develop a meningioma in their lifetime. Meningioma treatments are largely restricted to surgery and radiotherapy (RT), and systemic therapies remain ineffective or experimental. The World Health Organization (WHO) has historically graded meningiomas according to histological features. According to WHO criteria, many grade 1 meningiomas can be effectively treated with surgery or RT, but many WHO grade 2 or grade 3 meningiomas are resistant to treatment and cause significant neurological morbidity and mortality. Approximately 30% of WHO grade 1 meningiomas develop recurrences that cannot be predicted from histological features, and some WHO grade 2 or grade 3 meningiomas are unexpectedly well controlled with surgery and RT. These data indicate that improvements in meningioma risk stratification are needed, but limited understanding of meningioma biology and the misconception that all meningiomas are “benign” has encumbered medical advances for patients. Postoperative RT improves local control of meningiomas, but the benefits of meningioma radiation must be weighed against long-term toxicities, which can include neurocognitive deficits and secondary cancers. Most meningioma patients survive 5 years or more after diagnosis and are therefore at risk of long-term side effects of ionizing radiation on the normal brain, including white matter change, microvascular damage, and cognitive deficit. In recognition of the controversies surrounding meningioma risk stratification and treatment, clinical trials in North America and in Europe currently randomize patients with newly diagnosed WHO grade 2 meningiomas to postoperative surveillance or postoperative RT after gross total resection. Thus, there are unmet needs for improved risk stratification and prediction of postoperative RT responses for meningioma patients. To address this, we performed molecular profiling on 1856 frozen or formalin-fixed paraffin-embedded meningiomas from 12 institutions across 3 continents to develop a predictive 34-gene expression biomarker that outperforms all other risk stratification systems and identifies tumors that benefit from postoperative RT. The clinical and analytical validity of this biomarker were established in external cohorts and archival samples from NRG/RTOG 0539, the only successful prospective study of RT for meningiomas in North America. Despite these advances, all our meningioma gene expression profiling was performed in research laboratories, and our biomarker calculations largely relied on a hybridization and barcode-based platform for transcript quantification that is not widely available in the clinic. Our central hypothesis is that gene expression profiling will enable biomarker detection in clinical meningioma samples in a CLIA/CAP-certified setting. To test this, we will deploy two platforms for transcript quantification that rely on hybridization and barcoding, or on RNA sequencing, to test archival (Aim 1) or prospective (Aim 2) meningiomas. Successful completion of this proposal sets the stage for biomarker-stratified clinical trials that are currently under development for meningiomas through NRG Oncology.

NIH Research Projects · FY 2025 · 2025-09

1 PROJECT SUMMARY 2 Organoids derived from stem cells are valuable tools that have potential to unlock cellular-based regenerative 3 medicine applications for otherwise incurable illness. Severe liver dysfunction, which affects millions of people 4 and is the 10th leading cause of death in the United States per year, can only be treated with a liver transplant, 5 but a critical shortage of organ donors means many patients die while on the transplant waiting list. Creating 6 functional and transplantable bioengineered liver tissue that can treat severe liver dysfunction would address a 7 critical unmet need in modern-day medicine. Achieving closed system operation and effective cryopreservation 8 in tissue culture are critical requirements for producing clinical grade bioengineered tissue that meets 9 regulatory requirements. With the goal of producing highly functional bioengineered liver tissue for cell-based 10 regenerative medicine applications, we developed a custom bioreactor to biomanufacturer induced pluripotent 11 stem cell (IPSC)-derived liver organoids in the simulated microgravity of low shear rotational suspension 12 culture. Our custom bioreactor, Tissue Orb, supports isochoric supercooling cryopreservation and is the 13 world’s first isochoric supercooling chamber that can support sterile tissue culture. Isochoric supercooling uses 14 constant volume to maintain aqueous solutions in a liquid state at sub-zero temperatures without the harmful 15 formation of ice. Importantly, the constant volume confinement has been shown to impart stability to the 16 supercooled condition, which decreases the need of toxic cryoprotectants that are required in other 17 cryopreservation methods. Furthermore, isochoric supercooling is simple to implement, involving a rigid 18 container with no moving parts, and was recently demonstrated to be a promising cryopreservation method 19 that can be easily translated to clinical or research settings to preserve biological material. Our lab is the first to 20 apply isochoric cryopreservation to liver organoids. We are now combining this technology with simulated 21 microgravity tissue culture in efforts to produce high quality liver organoids for regenerative medicine therapies. 22 The objective of this proposal is to improve isochoric supercooling protocols for liver organoids and complete 23 the design of the Tissue Orb to allow tissue culture and cryopreservation to occur in a single, closed system 24 and is a critical step towards achieving scalability and repeatability that is needed for high quality, clinical grade 25 products. We will expand on our prior work through a biological aim which looks to improve the protocol for 26 isochoric supercooling of liver organoids (Aim 1) and an engineering aim which looks to complete the design 27 for the heat exchange system to allow integrated isochoric supercooling in the Tissue Orb (Aim 2). The 28 proposed aims will result in information and technology that can lead to substantial improvements in our ability 29 to produce and preserve liver organoids, which, in turn, can drastically increase the availability of high-quality 30 bioengineered liver tissues to researchers and healthcare facilities around the world.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY/ABSTRACT During pre-implantation mammalian development, cells must make their first critical fate choice to specify the trophectoderm (which forms the extra-embryonic placenta) or inner cell mass (which develops into the embryo proper). To achieve this lineage commitment, the embryo coordinates the major mechanical changes of compaction and polarization with critical signaling programs that drive cell fate. The role of Yap mechanotransduction in guiding trophectoderm fate beginning at the 8-cell stage has been well-studied. However, trophectoderm development is not driven by Yap signaling alone, but also requires activation of the Notch pathway. While other developmental contexts provide insight into how Notch could interact with mechanics and Yap to guide cell fate decisions, the role of Notch in mouse pre-implantation lineage determination remains unknown. This proposal will elucidate how Notch integrates with Yap and mechanics to guide trophectoderm specification and pre-implantation embryogenesis. I will investigate the central hypothesis that Notch lateral inhibition starting at the 2-cell stage primes a subset of cells toward the trophectoderm lineage; during polarization, Notch signaling could stabilize the apical domain to reinforce Yap signaling and maintain trophectoderm identity through blastocyst morphogenesis. Historically, live cell dynamics of pre-implantation embryo development have been challenging to study due to phototoxicity. However, recent advancements in light-sheet microscopy, biosensors, and optogenetics allow us to visualize and perturb signaling dynamics with unprecedented spatiotemporal resolution. I will harness this high-resolution imaging system to determine (1) how Notch signaling relates to embryo mechanics and (2) how Notch and Yap signaling dynamics specify and maintain trophectoderm fate. For the completion of these aims, I will gain training in advanced microscopy, quantitative developmental biology, scientific communication, teaching, and mentorship. The expertise from my sponsor, Dr. Orion Weiner, an expert in signaling dynamics and cellular mechanics, and support from the F31 NRSA will ensure that I receive the mentorship and resources necessary to complete my proposed research and training plan. My work will reveal how the embryo integrates different modes of signaling with mechanics to guide cell fate decisions. A better understanding how pre-implantation development is regulated will help advance Assisted Reproductive Technologies and ameliorate infertility.

NIH Research Projects · FY 2026 · 2025-09

The proposed K01 Award will provide the candidate, Dr. Lauren Suchman, with 1 training and skills to achieve her long-term goal of becoming an independent investigator who enhances access to quality sexual and reproductive health (SRH) care across the lifespan. Dr. Suchman is an Assistant Professor at the University of California San Francisco (UCSF) with expertise in women’s health, SRH, and HIV with a strong foundation in qualitative methods. The tailored training and research programs described in this K01 application will enhance Dr. Suchman’s ability to conduct independent research in line with NIH’s new high-priority area outlined in NOT-OD-24-119: Research Opportunities Centering the Health of Women Across the HIV Research Continuum. She requires additional training to: 1) develop topical expertise in HIV and menopause with a focus on symptom management and its impact on health outcomes; 2) develop proficiency in clinical research and statistical methods; 3) obtain experience in participatory intervention design; and 4) develop professional skills for career advancement. She has assembled a mentorship team of experts in HIV and menopause, the epidemiology of brain health outcomes in aging women with HIV (WWH), biostatistical methods, and participatory intervention design. These mentors will support Dr. Suchman’s transition to independence through didactic coursework, one-on-one meetings and tutorials, access to adjacent research teams and working groups, and completion of an independent research project. Dr. Suchman’s program of research centers on the midlife health of WWH in the U.S. with a particular focus on the San Francisco Bay Area. Menopause is a critical inflection point in the health of midlife cisgender women, which disproportionately impacts WWH. The menopausal transition and severity of associated symptoms has also been associated with suboptimal engagement in HIV care and antiretroviral treatment. However, WWH are less likely to be offered efficacious treatments for menopausal symptoms and less likely to accept these treatments if offered compared to women without HIV. To address these gaps, the study for this K01 aims to understand how menopausal phase and symptom severity affect HIV outcomes in midlife WWH and determine how therapeutic management of menopausal symptoms can best be implemented in an HIV clinic. Study outcomes will serve as proof of concept for an R01 proposal to develop and empirically test a multi-level intervention for both providers and patients. Research and training will occur at UCSF and will leverage the Multicenter AIDS Cohort Study (MACS)/Women's Interagency HIV Study (WIHS) Combined Cohort Study (MWCCS) and the UCSF Women’s HIV Program. Research findings and skills obtained through this K01 award will facilitate Dr. Suchman’s transition to become an independent investigator who develops interventions aimed at improving SRH care for WWH as they age.

NIH Research Projects · FY 2026 · 2025-09

PROJECT SUMMARY/ABSTRACT This proposal emerges from the discovery that the D1 family of dopamine receptors, a class of G protein coupled receptor (GPCR), long considered to only signal from the plasma membrane, also signal from the Golgi membranes. This subcellular and compartmentalized signaling challenges some of the basic paradigms of signaling regulation. D1 dopamine receptors are the main GPCR in the midbrain and regulate functions such as locomotion, cognition, attention and impulse control. Several pathological conditions including Parkinson's disease, schizophrenia and addiction are due to dysregulation of this signaling pathway. Many drugs of abuse promote dopaminergic transmission within the midbrain by increasing the release of dopamine and activating D1 dopamine receptors, D1DRs. This increased activity of midbrain dopaminergic neurons results in increased locomotor activity and motivational behaviors such as craving and drug-seeking. In all of these studies, it has been assumed that functions of D1DRs are mostly limited to the plasma membrane. Our data challenge this assumption. We show that these receptors signal from both the plasma membrane and the Golgi in the midbrain neurons relevant to locomotor and motivational effects regulated by dopamine. We have found that signaling from each compartment has distinct effects on the molecular and cellular consequences of D1DR activation. Importantly, we have found that D1DR signaling from the Golgi plays a critical role in the molecular mechanisms associated with dopaminergic signaling events related to addictive behaviors. The overall goals of this proposal are to elucidate the molecular, cellular and physiological consequences of D1DR signaling from the PM and the Golgi compartments in primary medium spiny neurons, MSNs. We have developed or adapted state-of-the-art tools such as a light-controlled nanobody recruitment system and photoactivatable bacterial adenylyl cyclase to selectively modulate D1DR signaling at a given subcellular location. We will combine these tools with molecular and cellular readouts of neuronal activity as well as a high throughput phosphoproteomics approaches to identify the consequences of signaling from each compartment. We will then apply these tools in zebrafish, an established animal model to study the significance of D1DR compartmentalized signaling in regulating dopaminergic-mediated behavior. This project brings together conceptually and technically innovative approaches in cells and in intact animals as well as high throughput methods to identify downstream targets and potentially new therapeutic targets for countering the effects of mis-regulation of dopaminergic signaling.

NIH Research Projects · FY 2025 · 2025-09

Project Summary/Abstract This project explores the critical and underappreciated role of reverse electron transport (RET) in lung aging and idiopathic pulmonary fibrosis (IPF), a complex biological phenomenon marked by a progressive decline in lung function and increased susceptibility to chronic diseases. IPF, in particular, presents a relentless clinical challenge with limited therapeutic options and a median survival of only 2-5 years post-diagnosis. Emerging evidence implicates mitochondrial dysfunction, especially RET, in the advancement of this condition. RET, characterized by an atypical backward flow of electrons through the mitochondrial electron transport chain, leads to a surge in reactive oxygen species (ROS) production and NAD+/NADH ratio imbalance. This dysfunction is believed to be exacerbated by aging-related mitochondrial DNA damage, heightening oxidative stress and metabolic disturbances associated with IPF pathogenesis. Our investigation, supported by promising preliminary data utilizing the RET inhibitor CPT-2008, aims to elucidate RET's impact on pro-fibrotic gene expression in lung fibroblasts and its role in lung collagen accumulation and fibrosis. The first objective is a detailed gene expression analysis to understand the specific influence of RET lung fibroblasts, considering variables like aging and lung injury. The second objective assesses the therapeutic potential of RET inhibition in ameliorating lung fibrosis, employing both young and aged mouse models. This part of the study extends to evaluating RET inhibition's effects on human IPF lung tissue, thereby linking animal model findings to potential clinical applications. This research is poised to significantly enhance our understanding of the mechanisms underlying lung aging and IPF. By shedding light on the previously unexplored role of RET in these processes, the project aims to unveil novel pathways and therapeutic targets, offering foundational advancement in knowledge about the role of RET and a transformative perspective in the management and treatment of IPF, a condition of major concern in the aging population.

NIH Research Projects · FY 2025 · 2025-09

The long-term objectives of this project are to elucidate the mechanisms underlying human endocrine lineage commitment and harness this knowledge to improve strategies for beta cell replacement in diabetes treatment. Diabetes is one of the fastest growing health emergencies worldwide, affecting hundreds of millions of individuals. Type 1 diabetes (T1D) is a disease of the endocrine pancreas characterized by lack of glucose homeostasis due to immune-mediated destruction of insulin-producing beta cells. Beta cell replacement therapy holds great promise for eliminating the need for exogenous insulin delivery and effectively curing the disease. Understanding the mechanisms underlying endocrine cell fate and function will be crucial for continued progress towards realizing the goals of both cell replacement therapy and beta cell regeneration. Although several protocols have been devised to generate insulin-secreting beta-like cells from human pluripotent stem cells (hPSCs), these protocols suffer from the production of non- endocrine cell types and a failure to match the transcriptional profiles and glucose responsiveness of native adult human islets. This may be because current protocols are based on knowledge of rodent development and may therefore be missing key regulatory pathways and lineage steps unique to human development. Indeed, multiple studies have identified discrepancies between mouse and human pancreatic islets, including structural, transcriptomic, and metabolic differences. Therefore, gaining a deeper understanding of human endocrine development is crucial for continued progress towards generating in vitro-derived beta cells that recapitulate endogenous function. Our laboratory has recently made a set of fundamental discoveries regarding endocrine progenitors (EPs) in human. We utilized single-cell multi-omics to generate a comprehensive atlas of developing pancreas, identified four novel EP states unique to human, and revealed divergent lineage relationships in human tissue versus mouse. We have identified a novel cell population, marked by the transcription factor FEV, that represents a fate-restricted EP population that gives rise to human beta cells. Our work now provides a critical new lens through which to examine human endocrine lineage commitment, and to apply this knowledge to improve beta cell differentiation from hPSCs. Our goal is to elucidate the mechanisms underlying human endocrine lineage commitment and harness this knowledge to improve strategies for beta cell replacement. The experiments outlined in this proposal begin with a focus on characterizing the spatial and temporal appearance of the novel EP populations that we have identified in developing human pancreas. In addition, studies will be undertaken to investigate the function of the gene FEV in the development of human beta cells using genome editing of hPSC-derived endocrine cells. Lastly, we will use a combination of strategies to isolate distinct populations of differentiating hPSC-derived beta-like cells and their progenitors to test whether off-target cells are detrimental and to refine which sub-population leads to optimal diabetic rescue.

NIH Research Projects · FY 2025 · 2025-09

PROJECT ABSTRACT Cytotoxic CD8 T cells are a necessary component of the anti-cancer immune response. When CD8 T cells encounter chronic antigen stimulation, they become dysfunctional and can no longer effectively exude their cytotoxic function1-3. Our lab and others have identified a CD8 stem-like progenitor, called Tpex, that retains its ability to kill cancer cells4. It was found that upon immune checkpoint blockade (ICB) therapy, these Tpex cells differentiate, egress from the tumor-draining lymph node (tdLN), and migrate into the tumor microenvironment to kill the tumor via cell-mediated cytotoxicity4. When the tumor-draining lymph node has been metastasized (metLN), this Tpex response is abrogated, and there is not a productive anti-tumor response with ICB4. Our lab found that in the metLN, Tpex were present but were preferentially surrounded by suppressive Tregs and DCs4. Additionally, Tregs are known to play an immunosuppression role in the TME and have a negative impact on the effectiveness of many current immunotherapies5-8. How Tregs are affecting the environment in the metLN and how Treg immunosuppression impacts other immune cell types, including Tpex, is not yet known. This proposal will test the hypothesis that LN metastasis amplifies Treg function both directly and indirectly to create an immunosuppressive niche in the metLN, limiting the function of Tpex cells and impairing a proper CD8 T cell response to ICB. Aim 1 of this proposal will identify the mechanism by which Tregs are creating an immunosuppressive environment in the tumor-draining lymph node. Aim 2 will define how Tregs are impacting the Tpex functionality. Aim 3 will determine how the Treg-Tpex cell axis affects ICB response and efficacy in the tdLN. This research approach will use a variety of methods, including an ex vivo organoid model, high-dimensional flow cytometry, multiplexed ion beam imaging (MIBI), single-cell RNA sequencing, and in vitro co-culture assays. These proposed studies will be among the first to study immune cell interactions and Tpex cells using patient-derived lymph node organoids. These results could elucidate novel mechanisms of immunosuppression in the tdLN and indicate how those mechanisms impact current cancer immunotherapy. This would result in an improved understanding of Treg and Tpex function in the tdLN and how ICB impacts them in the tdLN. The use of human-derived organoids as the primary model allows this research to be greatly translationally relevant and inform our understanding of cancer immunotherapy response, specifically in patients. Furthermore, this work will better inform novel immunotherapy approaches for the treatment of patients with metastatic cancer, especially in the fields of biologics and adoptive T cells, where preventing CD8 T cell exhaustion is a major goal. This research project and fellowship training will be conducted at a top-funded research institution, the University of California, San Francisco (UCSF), in the laboratory of Dr. Matthew Spitzer. Dr. Spitzer is an investigator with expertise in systems immunology, cancer immunology, and CD8 T cell biology as well as spatial and single-cell sequencing methods. This mentor and institution will provide a high-quality training environment with ample resources for the completion of this research and development of professional skills required for a career in academic research.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY/ ABSTRACT Methamphetamine (MA) use has been increasingly recognized as a significant contributor to the worsening health outcomes of individuals with HIV (PWH), with severe comorbidities including immune dysfunction, cardiovascular disease, and pulmonary hypertension. Even in the presence of effective antiretroviral therapy (ART) that maintains viral suppression, MA use exacerbates HIV pathogenesis and complicates efforts to achieve an HIV cure. Despite the high prevalence of MA use among PWH, its direct effects on HIV cure efforts, particularly how it alters HIV reservoirs and immune function, remain poorly understood. The overarching goal of this proposal is to investigate the complex intersection of HIV, methamphetamine use, and immune dysfunction. In addition, this career mentorship award will provide protected time for Dr. Lee to expand her mentoring capacity of early-stage data science investigators in the interdisciplinary field of clinical translational HIV and substance use research. This proposal outlines a comprehensive, data-driven approach to investigate how MA exposure influences HIV reservoir transcription and host immune responses, with the goal of developing computational models that can predict the biological consequences of MA exposure, and ultimately, inform HIV cure strategies. The specific aims of this proposal will employ advanced statistical and computational methods to examine the causal relationships between MA exposure and immune dysfunction, quantify the pharmacologic effects of MA in individuals with HIV, and identify the genetic factors that mediate these effects. In Aim 1, we will apply causal inference methodologies to longitudinal data from Dr. Lee’s ongoing Effect of Methamphetamine on Residual Latent HIV Disease (EMRLHD) cohort. This analysis will focus on determining whether MA exposure causally influences host immune dysfunction, including the activation of the NLRP3 inflammasome and elevated IL-1β signaling, as well as whether it contributes to persistent HIV transcription in CD4+ T cells of individuals on ART. In Aim 2, we will use pharmacokinetic/pharmacodynamic (PK/PD) modeling to explore the dose- and host- specific effects of MA on HIV reservoir transcription and immune function. Using data from a randomized placebo-controlled trial of PWH on ART receiving oral MA, we will apply nonlinear mixed-effects PK/PD modeling to assess how varying doses of MA impact host immune responses (such as cytokine levels and gene expression) and HIV reservoir transcription. Finally, in Aim 3, we will prioritize genes that are functionally relevant to MA exposure using a transcriptome-wide association study (TWAS) approach, combined with machine learning algorithms, integrating genetic data with transcriptome data to identify genes whose expression is influenced by MA exposure. This work will address critical gaps in our understanding of how MA exposure impairs immune function and maintains HIV reservoirs, even in individuals on ART. Ultimately, this work will not only advance our understanding of MA’s role in HIV pathogenesis but also help lay the foundation for innovative, data-driven therapeutic strategies to address both HIV and substance use comorbidities in the future.

NSF Awards · FY 2025 · 2025-09

With the support of the Chemistry of Life Processes (CLP) Program in the Division of Chemistry, Professors Michael Therien and David Beratan of Duke University and William DeGrado of University of California San Francisco are studying new approaches to design materials that direct, store, and release energy. Biology has developed numerous designs that carry out these functions; chemists, however, have yet to create energy harvesting, storage, and release systems from scratch that possess the sophistication of those seen in nature. Recent advances in protein design enable chemists to construct large molecules that capture and manage the flow of positive charges, negative charges, and energy. By designing protein-based materials that migrate and collect charges and energy, unique optical, electrical, and chemical functions will be realized. The experimental procedures used in this effort provide new tools to build proteins having innovative designed functions. This pursuit allow graduate students and postdoctoral fellows to acquire specialized training in synthetic chemistry, protein design, protein biochemistry, modern computational methods, and techniques to monitor fast processes that move charge and energy. The protein design methods developed are broadly applicable and enable construction of new biologically inspired materials that carry out novel functions not seen in nature. Outreach activities of this project introduce college and pre-college students to important new technologies and teach skills important for future careers in science and engineering. Biological energy transduction relies on protein-cofactor assemblies that possess physico-chemical functionality that far exceeds that realized to date through molecular and macromolecular design and synthesis. This effort designs redox proteins that transduce energy using bound cofactors, redox-active amino acids, titratable sidechains, and buried water molecules, to orchestrate the light-triggered flow of electrons, holes, and protons, elucidating rules and principles important for driving thermodynamically reversible reactions at low overpotential and engineering vectorial control over electron and proton currents. This project takes advantage of an integrated, multi-disciplinary approach that employs: (i) design and synthesis of light-harvesting and redox-active cofactors, (ii) de novo protein design using advanced computational methods to selectively bind cofactor units in precise, organized spatial arrangements, (iii) protein expression and characterization, (iv) state-of-the-art pump-probe transient optical methods and theoretical models that interrogate photo-induced electron and proton migration reactions, and (v) spectroscopic, potentiometric, and dynamical methods, high resolution protein structure, and predictions made by theory to provide insights into how atomic-level control of cofactor environments directs energy transducing function. Information from this study elucidate fundamental principles required to understand photosynthetic energy transduction and to design proteins that possess novel electro-optic function and can transduce energy via innovative pathways. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2025-09

Project Summary/Abstract The rewiring of transcriptional circuits over evolutionary time is a major source of biological novelty. This proposal seeks to determine the detailed molecular mechanisms that underlie transcriptional rewiring using unicellular yeasts as a model system. The strategy is based on direct experimentation in many different yeast species in the Saccharomycetaceae lineage, and utilizes mutant construction, genome-wide transcriptional profiling, chromatin immunoprecipitation, phylogenetic comparisons, and ancestral protein reconstructions. Circuit comparisons among these yeasts will uncover specific examples of transcriptional rewiring, and deeper analyses will reveal the molecular mechanisms by which the wiring changes occurred. Although much rewiring is probably neutral, some of it appears adaptive: indeed, a major mechanism for evolutionary novelty involves rewiring transcriptional circuitry to allow new expression patterns of existing gene products. Thus, to truly understand the structures of transcription circuits in modern species, we need to know the mechanisms by which they rapidly evolve and how these mechanisms lead to and, thereby can account for, modern structures.

NIH Research Projects · FY 2025 · 2025-09

For an animal to successfully navigate and adapt to its environment, effective communication between neurons at specialized sites called synapses is crucial. Impairments in synaptic transmission are frequently observed in various neurological and psychiatric disorders. The postsynaptic compartment of excitatory synapses, also known as the postsynaptic density (PSD), consists of complex protein networks that allow the transduction of neurotransmitter signals into electrical and chemical signals within the neuron. The PSD undergoes continuous, activity-dependent protein remodeling that directly impacts synapse function. Thus, profiling the proteomic landscape of active synapses is a powerful approach to uncover the molecular mechanisms of activity-driven synaptic remodeling. To study this, I utilized Cal-ID, a newly developed enzyme that biotinylates proximal proteins in a calcium-dependent manner. Given that synaptic activity triggers localized increases in Ca2+ level, I employed synaptic Cal-ID as a cutting-edge technique to investigate active synapses with unprecedented spatiotemporal resolution, enabling the targeted enrichment of proteins specifically associated with synaptic activity. First, I validated the ability of synaptic Cal-ID to respond to changes in synaptic activity. Next, I used synaptic Cal-ID to perform unbiased proteomic screens in cultured neurons and mouse brains. I identified two novel candidates, Anks1a and Ubash3b, that were not previously known to be in synapses. I found that Anks1a and Ubash3b are localized to excitatory synapses, show activity-dependent synaptic localization, and play important roles in basal synaptic function. This goal of the proposal is to study Anks1a and Ubash3b by 1) determining how activity leads to synaptic recruitment of Anks1a and Ubash3b, 2) determining whether Anks1a and Ubash3b regulates synapse function through EGFR signaling, and 3) determine whether Anks1a and Ubash3b are necessary for synaptic plasticity and learning and memory. This research is significant because it will 1) bring insight into activity-dependent remodeling in synapses, 2) help identify new mechanisms that can be targeted to treat neurological diseases, and 3) validate a novel tool for analysis of active synapses, which paves new and exciting paths for the synaptic field. The applicant has proposed this work in part to further their long-term goal of establishing an independent research career to understand synapse function and activity within different cell types across development, and how disruptions in synaptic function can lead to neurodevelopmental disorders. This proposal consists of various career development and transition plans, combined with extensive training in live-imaging, super-resolution microscopy, and quantitative analysis for proteomics under the guidance of an expert mentoring team, all of whom will mentor the applicant through the transition to a tenure track academic position.

NIH Research Projects · FY 2025 · 2025-09

ABSTRACT Primary open-angle glaucoma (POAG) is a leading cause of irreversible blindness worldwide, affecting over 80 million people. It is characterized by the progressive loss of retinal ganglion cells (RGCs) and optic nerve damage. Elevated intraocular pressure (IOP) is causal risk factor of glaucoma. While current treatments focus on lowering IOP, there is a critical need for therapeutic strategies that target neuroprotection, as no existing treatments effectively prevent or slow RGC death and axonal degeneration. Current animal models for glaucoma, such as inducible IOP elevation models have limitations in replicating the gradual, spontaneous progression of human glaucoma. Moreover, inherited mouse models, such as DBA/2J, exhibit slow and variable disease progression and anterior segment inflammation. Glis1 knockout (KO) mice offer a novel genetic model to studying glaucoma. GLIS1 is a Krüppel-like transcription factor predominantly expressed in the trabecular meshwork (TM) and undetected in the retina. Importantly, GLIS1 deficiency induces progressive degeneration of TM, leading to IOP elevation. Thus, this model provides a unique system to study ocular hypertension-induced glaucomatous neurodegeneration without confounding effects of Glis1 deficiency in the retina. Moreover, Glis1 KO mice exhibit no major abnormalities outside the drainage angle specific phenotype, making them ideal for longitudinal studies. Aim 1 will characterize glaucoma-relevant phenotypes in Glis1 KO mice across two genetic backgrounds (C57BL/6J and 129S6/SvEvTac), evaluating structural and functional changes, including axonal degeneration, RGC loss, dendritic changes, and neuroinflammatory responses. Aim 2 will identify molecular pathways associated with glaucomatous neurodegeneration using single cell RNA sequencing (scRNA-seq) of RGCs. Integrating findings of the gene expression changes with human POAG genome-wide association studies (GWAS) with help prioritize candidate genes contributing to glaucoma. Our study will establish the Glis1 KO model as a valuable resource for the glaucoma research community, addressing limitations of existing models by offering a more physiologically relevant and reproducible model for investigating glaucoma pathogenesis. Insights from this experimental glaucoma model may lead to novel strategies for glaucoma treatment.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY Preterm birth affects 10% of pregnancies, with 40% of survivors developing bronchopulmonary dysplasia (BPD), a chronic lung disease marked by dysmorphic vascular growth and alveolar simplification. Despite advances in neonatal care, BPD incidence has not improved, and no therapies reverse its pathology. My long- term goal is to define critical mechanisms of lung intercellular crosstalk during alveolarization to develop therapies for BPD and other chronic lung diseases. Alveolarization is a key phase of lung development involving synchronized endothelial and alveolar epithelial cell (AT) expansion to establish the gas exchange surface. Endothelial cells (EC) influence lung development through angiogenesis and signaling to other cell types. Our data show EC-specific deletion of IKKβ, an essential NFκB regulator, impairs alveolarization by disrupting angiogenesis, reducing capillary EC and AT cell abundance, and altering cell-ECM interactions. However, mechanisms governing NFκB/IKKβ activity in specific EC subpopulations remain unclear. The central hypothesis of this proposal is that NFκB/IKKβ activity in capillary ECs promotes alveolarization through cell-autonomous mechanisms that drive angiogenesis and cell-nonautonomous mechanisms that regulate AT proliferation and alveolar niche organization. This will be tested in two aims: Aim 1: Identify cell-autonomous mechanisms downstream of NFκB/IKKβ that promote angiogenesis in specific EC subtypes. Single-nucleus RNA sequencing (snRNAseq) and CUT&RUN sequencing will define IKKβ-mediated proangiogenic gene expression in EC subpopulations. Candidate gene validation in vitro (Hmbox1, Fzd4, Glp1r and others determined from snRNAseq) will assess roles in EC proliferation and survival. Aim 2: Determine whether endothelial IKKβ promotes AT proliferation and ECM organization via cell-nonautonomous mechanisms. We will perform EdU proliferation assays, primary co-culture systems, and advanced imaging (transmission electron microscopy and confocal-generated 3D renderings from thick sections) to assess the impact of EC- IKKβ deletion on AT dynamics and alveolar niche architecture. Target genes among AT and identfied in Aim 1 will also be validated We will use the endothelial-IKKβ deletion mouse model we generated for in vitro and in vivo outcomes. These studies will provide novel insights into the role of EC subtypes and NFκB signaling in alveolarization and inform therapeutic strategies for BPD and other pulmonary diseases. Completion of this project will also equip me with the expertise to conduct rigorous, independent research on lung vascular and epithelial biology, and foster a career studying intercellular and ECM interactions within the alveolar niche.