University Of California, San Francisco

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT Implantable deep brain stimulation (DBS) is a second-line surgical neuromodulation for Parkinson's disease (PD) that can provide significant relief of motor symptoms when medications become less effective, however there are currently no reliable predictors of therapeutic efficacy. While the gold standard suggests that a patient will benefit from DBS if their motor symptoms respond to PD medications with at least 30% improvement, the pre- dictive accuracy of this criteria is variable across studies, and has been disproportionately evaluated in the con- text of only one of two common brain targets for PD. A lack of reliable prognostic criteria to predict overall out- comes with DBS, including risk for cognitive side-effects in balance with motor symptom improvement, has led to variable patient outcomes. Some not considered candidates by the gold standard have been reported to re- spond well to DBS, while others have experienced limited benefit despite strong candidacy and well positioned electrodes. With over 4000 DBS surgeries performed in the US for PD each year, there is an increasing demand for better prognostic tools and streamlined approaches to inform optimal candidate and brain target selection. We aim to address this unmet need by leveraging advanced MRI techniques for improved prediction of patient outcomes after one year of DBS. Previous studies have shown that measures of brain connectivity derived from functional MRI (fMRI) and diffusion tensor imaging (DTI), can be used to predict motor symptom response to DBS. Brain iron accumulation in the basal ganglia, a marker of PD severity derived from susceptibility contrast on T2* MRI, has also shown promise for predicting DBS motor outcomes. However, practical implementation of the results from previous studies in the pre-operative setting is limited by the use of normative connectomes, post-operative electrode coordinates, and less sensitive susceptibility techniques for prediction, along with out- come data from only one of two brain targets for PD. To overcome these limitations, we will use patient-specific pre-operative MRI data to predict outcomes for both PD targets. Specifically, we propose a novel multivariate approach that incorporates fMRI and DTI with quantitative susceptibility mapping (QSM), a superior susceptibility technique to T2* MRI, to enhance prediction accuracy. By using complimentary features of disease burden that are highly relevant to DBS effects on brain connectivity and individual basal ganglia structures, we expect that our approach will improve upon the current gold standard. In 100 patients with PD undergoing DBS, we aim to: 1) evaluate the impact of 3T MRI on clinical prediction of motor outcomes, 2) identify MR and clinical features most relevant for predicting overall versus individual motor and cognitive outcomes, and 3) investigate additional variance in patient outcomes explained by post-operative targeting accuracy. The results will provide a framework in which DBS outcomes can be reliably predicted at the patient and symptom level to inform candidate and target selection, and even therapeutic settings. In this way, we can ensure that resources are geared toward patients most likely to benefit from DBS.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT An R25 PREP Grant would provide the core funding for a new postbaccalaureate program at UCSF. This program will build on our highly successful pilot program, PROPEL, which will end this year. The goal of the PREP program will be to provide trainees from historically underrepresented backgrounds with the research experience and enhanced scientific training needed to be competitive for top-tier biomedical science PhD and MD/PhD programs. To achieve this goal, we have developed a research education plan that includes defined Aims and measurable outcomes. Aim 1 is to provide PREP scholars with the opportunity to obtain substantive experience conducting rigorous biomedical research in a full-time mentored research position. A substantive experience conducting original research in a laboratory is one of the strongest predictors of success in graduate school. However, research internships are few and far between at many universities, and we find that insufficient research experience is one of the most common reasons why trainees from historically underrepresented groups are not competitive for our graduate programs. Aim 1 addresses this gap in experience by placing trainees into UCSF labs, where they obtain first-hand experience conducting discovery-based research in a supportive, mentored environment. Progress on their research as well as toward their scientific and career development goals are defined in a Mentoring Plan and monitored by their research mentor and in biannual committee meetings. Aim 2 is to promote the development of scientific knowledge and communication skills. A broad foundation of knowledge about modern biomedical science is important to prepare scholars for graduate level research in this multidisciplinary era, and to help them make informed decisions about what type of PhD program they want to go into. To develop this knowledge, PREP scholars attend lectures, and participate in educational activities such as a literature review workshop and biostatistics and programming classes. The scholars’ scientific development is assessed by the instructors, and the program content is evaluated and updated regularly. Aim 3 is to promote career development skills and a sense of community that reinforce scientific identity. A major barrier for many applicants to graduate school is a lack of sufficient experience with professional work environments as well as a lack of knowledge about the specific expectations of a graduate program in the biological sciences. To address this need, our scholars complete a series of workshops, mentored activities, and self-directed exercises that help to develop this crucial knowledge and professional skill set. The scholar’s career development is monitored through regular meetings with their mentors and with structured evaluations, and tangible products from these activities, such as an individual development plan and draft personal statement. These are used to reinforce the scholars’ scientific identity and prepare graduate school applications. Collectively, we expect that the experiences in the PREP Program will make our scholars more competitive for graduate school and will substantially advance their careers.

NIH Research Projects · FY 2025 · 2022-09

Project Summary The goal of this project is to comprehensively characterize the role of polymorphic natural killer (NK) cell receptors in multiple sclerosis (MS). Substantial data implicate NK cells in MS pathogenesis, but their precise function in mediating risk and disease course is poorly understood. Given that a large and diverse set of highly variable receptors govern NK cell activity, their characterization in MS promises to yield important insights into the role of NK cells in disease. Thus, we will characterize the nature and extent of the association of genomic variation of NK receptors across a set of diverse, established and deeply phenotyped MS cohorts. This goal will be achieved via our novel high-throughput, high-resolution next-generation sequencing (NGS) methodology. Using state-of-art technologies, we will contextualize this genomic variation by considering NK cell phenotype in disease. Finally, we will explore the functional implications of the observed NK receptor variation, allowing a mechanistic explanation of the impact of NK receptor expression to more fully understand the role of these highly variable receptors in MS. Together, the leukocyte receptor complex (LRC) on chromosome 19 and the natural killer complex (NKC) on chromosome 12 encode more than 90 distinct NK cell receptors, including the extremely polymorphic KIR (killer-cell immunoglobulin-like receptor) and LILR (leukocyte immunoglobulin-like receptors). In the work described here, we approach these complex systems across several key modalities, examining genomic, phenotypic and functional variation of NK receptors in MS to provide the first comprehensive depiction of the role of NK receptors in disease. Our preliminary work in a cohort of European ancestry identified a significant association of KIR variation with risk for developing MS, as well as with clinical and MRI features of disease. In Specific Aim 1, we will extend these findings to diverse patient populations and characterize KIR variation in MS across ancestries, in an additional cohort with longitudinal clinical and MRI data, and by investigating genomic variation of all known polymorphic NK receptors and their corresponding ligands. In Specific Aim 2, we will more fully resolve the role of these receptors in disease course by examining temporal expression patterns of NK cell receptors in individuals with MS through longitudinal CyTOF analysis. Finally, in Specific Aim 3, we will contextualize these results by harnessing CRISPR technology to characterize the impact of NK cell receptor expression level on NK cell function. This work promises to fill critical gaps in our understanding of the role of NK cells in MS. Because NK cell activity is tightly governed by the highly variable receptors that we investigate in this proposal, this work will provide important insight into the regulatory mechanisms underlying the influence of NK cells in MS susceptibility and disease course. The results will bear both on our understanding of disease pathogenesis and future efforts to develop NK cell-based therapeutic options.

NIH Research Projects · FY 2025 · 2022-09

Project Summary The ultimate product of our Center will be a series of comprehensive developing human and non-human primate (NHP) brain atlases of unprecedented cellular, spatial, and anatomical resolution. In Aim 1, we will characterize transient cell populations, establish the diversity of cell types present in specific brain regions, unravel complex developmental trajectories, and reveal conserved and divergent cell-type specific features. We will jointly profile of single nucleus RNA (snRNA-seq) and accessible chromatin (snATAC-seq) using the 10X Genomics snMultiome platform. All aims will include 30 anatomically distinct regions of fresh frozen developing human, rhesus macaque, and marmoset brains at four developmental epochs: mid-gestation, neonatal, childhood, and adolescence and perform probe-based validation. In Aim 2, we will conduct spatial transcriptomic and epigenomic mapping of cell types in fresh-frozen developing human and NHP brains using DBiT spatial-RNA-seq and spatial-ATAC-seq platforms. This approach will allow us to discover spatial and temporal features, including the developmental niche, proximity of cell types to each other, and regional abundance. In Aim 3 we will create Common Coordinate Frameworks for the developing human and NHP brain using high resolution (9.4T and 7T) MRI-based developmental structural atlases and leveraging existing developing human MRI data. Our final aim will create a cross species molecular and spatial atlas of brain development in human and NHP. This integration will enable us to identify conserved and diveregent aspects of the human brain and identify the developmental stages, spatial distribution, gene regulatory elements and cell types vulnerable to neurodevelopmental and neuropsychiatric disorders. We will coordinate to ensure that our developmental atlases merge with adult human, macaque and marmoset atlases that other BICAN centers create. The data collected by our Center will be perfectly aligned with the overarching goal of the BICCN in generating a comprehensive census of brain cell types across the lifespan that integrates molecular, anatomical, functional, and cell lineage data for describing cell types in human and NHP brains. By leveraging innovations in cell capture and spatial mapping technologies, the current proposal will have broad implications for understanding the cellular origins of diseases and for highlighting patterns of selective cell type vulnerability in neurodevelopmental disorders such as autism and schizophrenia. Additionally, our plans to create developmental cellular and molecular resolution maps of marmoset and macaque will provide foundational data for establishing primate models of human disease. Finally, our atlas of conserved molecular, epigenetic, and spatial properties will support the precise monitoring, targeting, and replacement of specific cell types and the improvement of in vitro models.

NIH Research Projects · FY 2025 · 2022-09

Abstract Protein-protein interactions (PPIs) are ubiquitous in biology, and their dysregulation is closely associated with diseases, from cancer to neurodegeneration to rare genetic disorders. PPIs often form complex networks that include highly interacting ‘hub’ proteins. Methods to modulate single interactions would provide great insight into the functions of these hubs. Small-molecule probes and drug leads have focused on blocking PPIs; however, stabilizing PPIs could be just as important for drug discovery and could provide greater selectivity for chemical biology. However, there are few systematic methodologies to discover PPI stabilizers prospectively. This proposal focuses on the systematic discovery of selective small-molecule PPI stabilizers, using the hub protein 14-3-3 as a model system. 14-3-3s are seven highly homologous adaptor proteins that bind to serine and threonine sites on client proteins to alter their function and fate. Hundreds of proteins in signal transduction pathways, cell-cycle regulation, transcription regulation, and protein homeostasis are clients of 14-3-3. Given the importance of protein phosphorylation and the ubiquity of 14-3-3 as an effector of phosphorylation, it is surprisingly underappreciated. We propose that developing a tool kit of selective, cell-active stabilizers of native 14-3-3/client PPIs will stimulate biological study and may lead to new drugs. Based on the structural diversity of clients, we hypothesize that we can develop client-selective PPI stabilizers that bind to the composite 14-3- 3/client interface. These selective stabilizers should amplify the native biology of the 14-3-3/client complex. We will provide proof-of-concept for this approach through three aims: Aim 1. Screen for selective stabilizers of 14-3-3/phosphopeptide clients. We have previously discovered disulfide-bound fragments that stabilize 14-3- 3/phosphopeptide complexes. We will now screen six, structurally and biologically diverse 14-3-3/client complexes, using a native C38 residue found only on the 14-3-3 isoform. We hypothesize that client sequences with more open or flexible structures near the C38 will yield higher quality hits. Aim 2. Optimize 14-3-3/client stabilizers for cell-based activity. We have demonstrated the ability to convert disulfides to cell-active electrophilic warheads and to tune the selectivity of 14-3-3/client stabilizers. We will optimize C38-bound fragments with (or without) an electrophile with the goal of achieving target-selective PPI stabilization in cells for the 14-3-3 clients CRAF kinase, estrogen receptor  (ER), and the transcription factor FOXO1. Aim 3. Design PROTAC-based degraders of 14-3-3 clients. PROTACs are bifunctional molecules that induce proximity between a ubiquitin ligase and a target, leading to the target’s degradation. We will expand the targets accessible to PROTAC technology by using 14-3-3 as a scaffolding protein to link intrinsically disordered proteins (IDPs) to the ubiquitin ligase. We will first develop the technology for ER, where PROTACs are known, then translate our learning to IDPs, including FOXO1. Successful completion of these aims will provide approaches that are broadly applicable to the 14-3-3 network and are extendable to other native and nonnative PPIs.

NIH Research Projects · FY 2025 · 2022-09

Between 1999 and 2018 more than 446,000 people died from profoundly slow and shallow breathing after opioid overdose. This number grows at a faster rate each year and in 2019 there were nearly 50,000 overdose deaths. Opioids are perhaps the most effective analgesic, therefore, despite concern, they will persist as a staple therapy in medicine and within our society into the future. Thus, novel solutions to safely use opioids must be developed. But to discover these, we must first understand the key cellular and molecular mechanisms for opioid depression of breathing, known as opioid induced respiratory depression (OIRD). Recently we demonstrated that opioids depress breathing mostly through their action upon µ-opioid receptor (MOR) expressing neurons in the same site where the breathing rhythm is created, the preBötzinger Complex (preBötC). In this proposal, a central goal is to define the key preBötC neural type and MOR signaling pathway(s) that cause OIRD. Results from both aims will have direct clinical and therapeutic impact. Here, we hypothesize that opioids cause OIRD by blocking synaptic vesicle release from just 140 MOR+ excitatory preBötC neurons. Additionally, we expect that these same neurons, despite being less than 10% of preBötC, are specialized for creating the pace of breathing in general, analogous to the cardiac pacemaker cells. If so, these will be the first neurons identified with such an important purpose. This is the second central goal of the proposal. To test these three hypotheses, we have designed a sophisticated intersectional genetic approach to selectively delete MOR in adult mice from either excitatory or inhibitory preBötC neurons or nearby upper airway motor neurons to then compare if and how much OIRD changes. Next, we will use a sensitive in vitro system to uniformly test the necessity of each primary MOR signaling pathway in suppressing preBötC rhythmicity. And last, we will use an intersectional genetic approach to measure changes in breathing after acutely, ectopically silencing the preBötC MOR+ excitatory neurons. The identification of the cellular and molecular mechanisms of OIRD will establish a framework for future studies to develop novel pharmacological approaches to block or rescue it. Moreover, this proposal aims to determine if MOR+ excitatory neurons are pacemakers for breathing. If so, we will have identified, perhaps, some of the most vital neurons in the brain.

NIH Research Projects · FY 2025 · 2022-09

Efficient and user-friendly paradigms to detect cognitive impairment, including dementia are needed in primary care. The UCSF Brain Health Assessment accurately detects cognitive impairment via an appealing tablet interface with automated scoring and EMR integration. With the first round of funding, we validated the paradigm in English and Spanish speakers, developed regression-based norms that accurately classify patients by their likelihood of impairment, identified racial and ethnic disparities and barriers to brain health care, and at UCSF primary care the paradigm increased diagnosis rates by 40%. The primary goals of the proposed work are to evaluate the effectiveness of the paradigm for improving brain health care via a large pragmatic trial, to address challenges to implementation, and to pioneer precision medicine approaches to reference group adjustments. In Aim 1, we will conduct a pragmatic cluster randomized trial in 26 Kaiser Southern California primary care clinics to determine the effectiveness of the paradigm on detection rates and other brain health outcomes. In Aim 2, we will identify and address challenges to implementation and sustainability. In Aim 3, we will further diversify our well-characterized reference group and refine our detection algorithms to adjust for social determinants of brain health (not race), which is novel and also critical for a paradigm to be appropriate for all members of the increasingly diverse U.S. older adult population. If successful, this work will produce a validated paradigm that is available to address the unmet need to detect cognitive impairment, including dementia, in large and diverse populations seen in primary care, and provide useful information about wide scale implementation of the paradigm to mitigate healthcare disparities and improve brain health care.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY The goal of this K08 application is to provide Dr. Jonathan Chou, an Instructor of Medicine at UCSF, with the skills he will need to become an independently-funded laboratory investigator. Dr. Chou proposes to elucidate the regulatory mechanisms of NECTIN4, the target of a newly approved antibody-drug conjugate (ADC) in metastatic urothelial cancer called enfortumab vedotin (EV) and identify mechanisms of resistance using PDX and metastatic biopsy samples. The proposal builds on Dr. Chou’s recent work, which showed that NECTIN4 expression is enriched in luminal subtypes of bladder cancer, and that increasing and decreasing NECTIN4 can enhance EV sensitivity or lead to resistance, respectively. Dr. Chou hypothesizes that the transcription factor PPARG, which regulates luminal bladder cancer cell identity and integrates fatty acid signaling, is a direct regulator of NECTIN4 and that transiently augmenting NECTIN4 expression in urothelial cancer cells will enhance the efficacy of NECTIN4-targeting therapies. In Aims 1 and 2, Dr. Chou will elucidate the mechanism underlying this regulatory pathway and determine whether sensitivity to NECTIN4-targeted therapies can be enhanced by directly modulating the PPARg pathway using pharmacologic approaches, biological modifiers and dietary alterations. In Aim 3, Dr. Chou will determine whether loss of PPARg or alternatively, activation of the EMT-associated TGFb pathway downregulates NECTIN4, thus leading to resistance. He will leverage EV- resistant cell lines that he has generated, patient-derived xenograft (PDX) models (established from minority patients treated at UCSF), as well as metastatic biopsy samples from UCSF patients treated on EV, to accomplish this Aim. Dr. Chou’s training and research plan includes a combination of structured coursework and workshops, one-on-one tutorials, and hands-on research experience that will all take place at UCSF, a world- renowned NCCN Cancer Center with a history of excellence in basic and translational cancer research. Dr. Chou’s training plan will complement his existing expertise to build a strong foundation in the following areas: 1) bladder cancer biology; 2) preclinical modeling of ADCs and adoptive T cell therapies; 3) cancer metabolism and drug resistance; and 4) genomics and next-generation sequencing methods and analysis. The project will be conducted under the mentorship of Dr. Felix Feng, Professor of Radiation Oncology and Associate Director for Translational Sciences, and co-mentored by Dr. Alan Ashworth, Professor of Medicine and President of the UCSF Cancer Center. He has assembled a distinguished advisory panel with complementary expertise to guide his research and career path. At the completion of this award, Dr. Chou will have the relevant didactic and research experience to become a leader in bladder cancer models, therapeutic targeting strategies and genomic approaches to investigate drug resistance, including to ADCs. If successful, this project will also provide a translational opportunity from the laboratory to the clinic, to utilize dietary modifications and thiazolidinedione drug combinations to augment responses and potentially reverse resistance to NECTIN4-targeting therapies.

NIH Research Projects · FY 2025 · 2022-09

Alzheimer’s disease (AD) and the related dementia, frontotemporal lobar degeneration (FTLD), share several key clinical and neuropathological features and have been considered as two ends of a disease spectrum. In support of this, genetic evidence shows that patients with dominant mutations in the Progranulin (GRN) gene invariably develop FTLD with TDP-43 proteinopathy, characterized by the accumulation of RNA binding protein TDP-43 in layers 2-3 of the frontal cortex. Interestingly, single nucleotide polymorphisms (SNPs) in the 3’UTR of the GRN gene reduce Progranulin (PGRN) protein levels and have been associated with increased risk of TDP- 43 proteinopathy in the limbic regions in 30-40% of AD patients. Despite these intriguing genotype-phenotype correlations, how PGRN deficiency promotes glial and neuronal pathology remains poorly understood. To investigate the mechanism of neurodegeneration in PGRN deficiency, we performed single-nuclei RNA- sequencing (snRNA-seq) in the thalamus of Grn-/- mice during the aging process and showed that, in PGRN deficiency, microglia are the first cell type to show progressive loss of homeostatic genes and acquire transcriptomic and histopathological features of a pro-inflammatory state that promotes neuronal cell death and TDP-43 proteinopathy. To connect these results with human disease, we’ve conducted a pilot study by comparing the results from snRNA-seq using postmortem tissues from the frontal cortex and thalamus of FTLD- GRN cases with those from similar brain regions in 19-month-old Grn-/- mice. This human-mouse snRNA-seq comparison revealed shared transcriptomic changes in FTLD-GRN cases and Grn-/- mice, including cellular responses in microglia (exocytosis, immune activation, and chemotaxis) and astrocytes (astrocyte-vascular coupling, cell adhesion, and synaptic organization) in both brain regions. Furthermore, our results uncovered transcriptomic changes in excitatory and inhibitory neurons in the frontal cortex and thalamus of FTLD-GRN cases, suggesting human-specific neuronal vulnerability. Together, these results broach the hypothesis that PGRN deficiency disrupts the gene regulatory network in microglia and astrocytes and alters intricate glia-neuron interactions to promote neurodegeneration in FLTD-GRN. To test this, we propose to 1) Map the transcriptome and gene regulatory network that define glial pathology and neuronal vulnerability in the frontal cortex and thalamus of FTLD-GRN; 2) characterize the mechanism of cellular resilience in neurons and glia in the visual cortex of FTLD-GRN by mapping their transcriptomes and epigenomes; and 3) delineate the functional consequences of transcriptomic and epigenetic modifications in FTLD-GRN using IPSC-based models. This project will provide critical data that fill the knowledge gaps regarding the trajectories of glial and neuronal pathology, and brain region-specific vulnerability and resilience in FTLD-GRN. Results from this project will further provide important insights and enable the discovery of biomarkers that predict disease progression.

NIH Research Projects · FY 2025 · 2022-09

Project Summary/Abstract AD is an age-related neurodegenerative disorder affecting 10% of the population over 65. The genetics of Late- onset AD (LOAD) is complex. Multiple common variants usually influence LOAD with smaller effect sizes. Whether and how these predicted AD risk genes contribute to AD pathogenesis and which variant affect their expression remain, for the most part, to be characterized. The overarching goal of the proposed study is to provide a comprehensive annotation of roles for AD risk genes and determine AD casual variants contribute to AD via modulating AD-risk gene expression. We will: (1) determine and benchmark the biological consequences of putative AD risk genes identified by genetic analysis using CRISPRi and single-cell RNA-seq in iPSC-derived excitatory neurons, hippocampal dentate granule cells, and microglia. We will also follow up using functional assays in cells and organoids for AD-related phenotypes, (2) characterize enhancers and AD-associated variants for AD risk genes in iPSC-derived hippocampal DG neurons and hippocampal organoids, (3) elucidate the roles of enhancers and AD variants in iPSC-derived excitatory neurons, microglia, and cerebral organoids with microglia. The proposed work will provide a comprehensive annotation of AD risk genes and demystify AD- causal variants affecting gene expression networks and cellular functions related to AD pathogenesis. The success of our proposal will be an essential step towards better risk prediction and new therapeutic targets in AD.

NIH Research Projects · FY 2025 · 2022-09

The health and well-being of older men susceptible to or living with HIV (MSLH) has been greatly shaped by adverse daily experiences (ADE). Their contemporary history and life course have been defined by exodus from difficult situations and the formation of supportive communities. The overall goal of this research is to shed light on the extent and the manner in which ADE shape older MSLH’s health. This study will assess the relationships among health, ADE, resources, and biomarkers of health and aging in older MSLH across HIV status. This research is needed because older MSLH’s health fares worse than that of the general population, their exposure ADE is higher, and they have less access to supportive resources than older people generally. Notably, 40 years after we first faced HIV, we continue to learn about the consequences of HIV, including aging with HIV. MSLH comprise the majority of older people living with HIV. Older MSLH face unique health risks and protective factors, yet they constitute a very complex population. What we know about their health comes from the general population of well-educated, and convenient samples. Hence, the need to collect data from valid samples. This is a cross-sectional study based on San Francisco Bay Area. Quantitative and Qualitative data will be collected from a sample of 600 older MSLH. We will collect ADE, individual, and biological data to test hypotheses regarding the associations among ADE, resources, and health (e.g., mental health, HIV risk, cognitive function). Our team is multidisciplinary and includes our community partner, The Elizabeth Taylor 50-Plus Network (of the San Francisco AIDS Foundation). This proposal addresses the Office of AIDS Research Strategic Plan of tackling HIV comorbidities and health. The data and findings from this study are intended to constitute the baseline for a longitudinal study.

NIH Research Projects · FY 2025 · 2022-09

1 PROJECT SUMMARY 2 Alzheimer's disease (AD) is the most common form of dementia worldwide, incurring a projected healthcare 3 burden of $1 trillion in the United States alone by 2060. It is the 7th leading cause of mortality in the United States 4 and it disproportionately impacts minoritized racial and ethnic groups. Although many pharmacologic 5 therapeutics have been tested to slow or remove the hallmark amyloid and tau aggregates, none have proved 6 effective at reversing AD cognitive decline. Thus, there remains urgency to identify factors that contribute to 7 dementia risk and novel approaches to intervene. With known correlations to conditions that include gut microbial 8 dysbiosis and exposure to sociocontextual factors that increase dementia risk, such as neighborhood 9 disadvantage, AD may be preventable or treatable at several points of individual- and community-level 10 intervention. However, the exact mechanisms by which these factors can contribute to AD are unknown. Our 11 group was first to identify gut microbiome alterations in AD, and subsequent studies in humans and mouse 12 models have correlated AD pathology with microbial components and associated increases in peripheral and 13 central inflammation. Exposure to neighborhood disadvantage is similarly related to both AD pathology/cognitive 14 decline and to inflammatory profiles. In fact, examining health outcomes linked to the gut microbiome and 15 neighborhood disadvantage more broadly reveals shared alterations in inflammation, methylation profiles, and 16 risk for cardiovascular disease. These are all comorbid risk factors for AD, begging the question as to the role of 17 neighborhood environment in altering gut microbiota composition and subsequently exacerbating AD pathology. 18 Studies of geography and household social relationships show an effect on macro- and micro-environment on 19 microbial composition, however it remains unclear whether this impact extends to mid-level neighborhood 20 environment. Thus, the proposed research rests on the central hypothesis that exposure to neighborhood 21 disadvantage is linked to a proinflammatory microbiota, thereby disrupting the intestinal barrier and aggravating 22 AD pathology. Work leading to this F99 proposal (Aims 1A, 1B) addressed metabolic and gut barrier mechanisms 23 of microbiota impact on AD. To test neighborhood effects, the proposed project features human cohort studies 24 to address the following F99 aim: 1C. Test the extent to which gut microbiome composition mediates the 25 relationship between exposure to neighborhood level disadvantage and AD pathology. In proceeding to a 26 postdoctoral fellowship, the proposed K00 research program will feature training to studying timing, exposure 27 and mechanistic effects of neighborhood disadvantage on AD pathology. The completed research is expected 28 to elucidate combined mechanistic effects of neighborhood disadvantage on AD, informing therapeutic and policy 29 interventions and developing the applicant’s training toward career goals of conducting translational 30 neuroscience research at a major academic medical center.

NIH Research Projects · FY 2025 · 2022-09

Older Americans are not all likely to receive high quality end-of-life (EOL) care due to the substantial variations in care quality as well as in goal-concordant care, adequate pain treatment, and palliative care access across organizations and geographic regions. Despite decades of research documenting variation in EOL care quality, there has been little progress in achieving uniform improvement. This is in part due to a failure to recognize organizational and systems-level factors (e.g., education, employment, and healthcare systems) as a root causes of variation in care quality. There is a critical gap in our understanding of how organizational and systems-level factors the life course influences the quality of serious illness care for older adults near the end of life. The objective of this project is to understand and address those factors as they relate to the systems level the quality of care provided near the end of life. I use a research approach that centers mutual learning and collaborative planning with local patients. This project’s career development plan will provide Dr. Elizabeth Dzeng with skills and knowledge in participatory research methods, implementation sciences, clinical trials, leadership in local-academic partnerships, geriatrics, and life course perspectives. A Beeson award will enable her to become an independent investigator in understanding and addressing the role of systems-level factors in EOL care using participatory methods and support her long-term goal of becoming an international research leader in geriatric palliative care. This study will be conducted at the San Francisco Bay Area to enable understanding of the influence of local and organizational histories, policies, and regions on healthcare. Dr. Dzeng has already been establishing local connections and developing her Patient Advisory Board. The first aim of this proposal seeks understand older adults’ experiences with systems-level factors and how these are perceived to influence care near the end of life. The project’s second aim is to examine how individual and organization-level factors influence the quality of care provided near the end of life and assess what enables or prevents improvements within healthcare organizations. The third aim is to refine and test an intervention that connects hospital leaders and local representatives to tailor a co-ordinated care approach aimed at improving the experience of older adults near the end of life. This study is highly innovative because it would be the first study to examine and address organizational and systems-level factors in geriatric palliative care using life course perspectives and participatory research methods. It uses novel interview and participatory methods to examine the multi-level factors that contribute to variation in EOL care. This study is significant because it will develop understanding at a deeper level of end-of-life care varies for patients with different life courses and address this variation through a multi-disciplinary, multi-component intervention. This study will inform a future R01 application for a multi-center hybrid effectiveness-implementation pragmatic trial of an organizational-level protocol intervention to mitigate variation EOL care quality.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Women in South Asia often return to their natal homes in different communities during pregnancy, for childbirth, and postpartum. Our team conducted the first robust measurement of this phenomenon in two states of India (coining the term “temporary childbirth migration (TCM)”) and found that about one-third of women returned to the natal home point during the perinatal period. Given India’s reliance on community-based maternal and child health care delivery, TCM may disrupt care continuity, negatively impacting health care access and health outcomes. Conversely, given the lower status of women in their marital homes, TCM could potentially improve health outcomes for migrants, leaving those “left behind” at higher health risk. A greater understanding of TCM, its impact, and associated mechanisms are critical inputs to inform policy and practice to improve maternal and neonatal health in India. However, despite anecdotal evidence of TCM’s widespread prevalence across diverse Indian contexts and its potential impact on health, it is not well defined or quantified in the literature. The objective of this study is to characterize TCM in India, describe its impact on perinatal outcomes, and explore the health care continuity and social support as potential mediators of these relationships. First, we will collect cross- sectional data from roughly 6000 women at eight Health and Demographic Surveillance Sites throughout India to define and characterize temporary childbirth migration, including magnitude, timing, drivers, and sociodemographic and geographic heterogeneity. Triadic qualitative interviews with household members in 2 sites will shed additional light on selection and drivers of TCM. Then, we will select three sites based on geographical distribution and differing levels of TCM for a longitudinal study. We will recruit 1000 women in each site (3000 total) who are early in pregnancy and follow them until one year postpartum, some of whom will migrate and some not. We will collect monthly telephone surveys and five longer in-depth surveys from women about migration patterns, health care use, maternal and child health outcomes, social support, and socio- demographics. We will ask women to take photographs on their mobile phones (provided to those without phones) of health record data to have higher quality maternal and infant outcome data. We will estimate the impact of TCM on preterm birth and identify women most at risk. We will then examine disruptions in the continuum of perinatal care and social support as potential mediators between temporary childbirth migration and health. These findings will help us understand factors that may be contributing to poor maternal and child health outcomes in India, and potentially other parts of South Asia and other regions. A better understanding of the impact of TCM can help programs and policies better support women to obtain the care and support they need in pregnancy and postpartum.

NIH Research Projects · FY 2025 · 2022-09

THE CANCER CELL MAP INITIATIVE v2.0 OVERALL SUMMARY The Cancer Genome Atlas and sister projects have now sequenced over 20,000 tumor genomes, providing a catalog of gene mutations, copy number variants and other genetic alterations associated with cancer. These data have made it clear that every cancer is a distinct genetic disease, with tumors that look physiologically similar often driven by patterns of gene mutations that are strikingly different. Due to this molecular heterogeneity, it is typically unclear what are the key driver mutations or dependencies in a given cancer and how these influence pathogenesis and response to therapy. One key observation for interpreting tumor genomes is that the many rare tumor mutations can be shown to converge on common molecular networks. Based on this premise we created the Cancer Cell Map Initiative (CCMI), whose mission is to create comprehensive maps of cancer molecular networks and to use these maps in intelligent systems for personalized therapy. In 2017, the CCMI was funded as an NCI U54 Research Center for Cancer Systems Biology, integrating expertise in network mapping, bioinformatic analysis and cancer research from leading academic laboratories at two University of California campuses (UCSF and UCSD). We have since generated comprehensive networks of protein interactions in breast and head-and-neck tumor cells and, from these data, identified several hundred protein complexes under selective mutational pressure in cancer (NeST v1.0). We have piloted deep learning systems (DCell, DrugCell and TCRP) that can use this protein network information to translate a patient’s tumor mutation profile to a predicted drug response, including FDA-approved and exploratory agents. We have implemented a rich portfolio of training opportunities and, leveraging UC institutional support, expanded the CCMI consortium to include more than a dozen faculty at UC and, most recently, Stanford. In the next five years, the CCMI will seek to: (1) Generate comprehensive protein interaction networks centered on key cancer driver genes in lung squamous cells (in healthy and diseased states) as well as the PIK3CA and TP53 pathways, which are central to many tumor types; (2) Systematically extend the CCMI collection of cancer protein interaction data with protein immunofluorescent imaging and cryo-electron microscopy to formulate multi-scale cancer cell maps; (3) Dissect the functional logic of these networks and maps by systematic genetic screening experiments in the same tumor types and pathways, using a panel of scalable cell proliferation, phenotype and pathway readouts; (4) Significantly advance and harden our DrugCell interpretable deep learning system for cancer precision medicine; (5) Train the current and next generation of scientists in network biology and its applications to cancer research; and (6) Continue to build a cadre of leading investigators to expand CCMI into a global coordinated partnership.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Our general strategy is to take advantage of novel tools and methodologies that we have developed during our first two CTD^2 funding periods– more specifically pioneering and applying CRISPR based technologies to aid the discovery and characterization of novel cancer targets and their modulators– using innovative high throughput technologies. Our end goal is to uncover optimal combinations of targets with the potential to eliminate all cancer cells, despite their clonal heterogeneity and environmental context. This requires us to better understand tumor biogenesis, namely the combinations of genes that drive oncogenesis, and tumor heterogeneity which complicates effective therapeutic treatment. In this proposal we build upon exciting systems allowing us to quantitate genotypic and phenotypic cell heterogeneity in cell culture and in vivo. The overall goal is to identify synthetic gene combinations necessary for clinical resistance and related to inter- and intra-tumor heterogeneity. We hypothesize that altered cell states such as inflammatory phenotypes and lineage plasticity fuels therapy tolerance and resistance. We apply single-cell approaches and cutting-edge lineage tracing tools to investigate the genesis of pathogenic cellular state changes and use genetic screening, computational and pharmacologic approaches, and clinically relevant in vitro and in vivo tumor models to identify mechanistically calibrated, specific therapeutic vulnerabilities. These approaches will be applied to two cancer, lung and breast adenocarcinoma. Tumor biogenesis and evolution is a challenging area of research, largely due to the complexity of cell types and behaviors and the combinations of genes that drive cancer types and subtypes is poorly understood. We have developed next generation GEMMs to interrogate gene combinations that promote cancer. In this aim, mouse models will be generated that contain combinations of genetic perturbations of the top 30 TCGA recurrent mutations. These studies will associate the combination of perturbagens with specific cell states, despite their clonal heterogeneity and cell state and lay a solid foundation for identifying which combinations of recurrent genes respond to which therapy, thus helping to stratify patients. This part of the research program focuses on lung cancer as it synergizes with other components of the proposal. We apply an evolved lineage tracing technology with single cell RNA-seq readout that lets us follow tumor evolution with unprecedented resolution. These studies will help us understand how tumor plasticity enables cancers to evade therapeutic challenges. And importantly, how the loss of tumor suppressor genes or gene combinations, alters the preferred evolutionary paths a single transformed cell takes to reach aggressive and metastatic states.

NIH Research Projects · FY 2025 · 2022-09

Project Summary/Abstract This proposal describes a 5-year plan to achieve the candidate’s goal of becoming an independent physician- scientist studying interactions between the airway epithelium and the type 2 immune system. With a comprehensive plan of coursework and hands-on training, guided by a skilled scientific advisory and mentoring team, and situated within the exceptional intellectual environment at UCSF, Dr. Kotas will utilize established, specialized tools and techniques from her mentor’s lab and become newly proficient in primary airway epithelial culture, translational studies using human tissues, and state-of-the-art transcriptional techniques. By the end of this project, she will possess a unique toolset that will differentiate her from her mentor and will have sufficient data to launch an independent research group while contributing new knowledge to the scientific community. The investigative purpose of this proposal is to elucidate how tuft cells—specialized epithelial cells with poorly understood function—influence the behavior of nearby epithelial and immune cells during chronic type 2 airway inflammation. As chronic allergic inflammation is the underlying cause of morbidity and mortality in millions of patients with diseases such as nasal polyps and type 2 high asthma, an improved understanding of the cells and molecules that contribute to these pathologies is needed to design new therapies. In preliminary data, Dr. Kotas finds that tuft cells in the chronically allergic upper airway environment adopt a distinct “allergic” phenotype characterized by increased production of prostaglandin E2 (PGE2). This is concurrent with a transcriptional signature of PGE2 on the neighboring epithelium, while in vitro, PGE2 stimulates epithelial fluid secretion. Transcriptional signatures of allergic tuft cells and PGE2 activation are also observed in the bronchus in type 2 high asthmatics, suggesting similar pathology throughout the allergically-inflamed respiratory tract. These findings urge further examination of tuft cells and tuft cell-derived PGE2 in allergic airway disease. In this proposal, Dr. Kotas will build upon her preliminary data to probe the effects of tuft cells and PGE2 on airway homeostasis. Aim 1 will use mouse cells in vitro and whole animal modeling in vivo to examine tuft cell- dependent alterations mucociliary movement and airway surface liquid composition. Aim 2 will examine the effects of tuft cells and PGE2 on the type 2 immune system in mouse models of allergic airway disease. And Aim 3 will return focus to human subjects and determine the phenotype and activation state of tuft cells in the lower airway in type 2 high asthma. Together, the proposed experiments will improve our fundamental understanding of how tuft cells contribute to allergic airway disease, and may identify new molecular and cellular targets for future therapy.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Survival outcomes for patients with lower grade gliomas continue to improve as diagnosis and treatment evolve. However, damage caused by tumor growth and by the consequences of treatment often leads to significantly impaired cognitive function. Our previous work has demonstrated that radiation therapy reduced ratios of N-acetyl-aspartate (a neuronal biomarker) levels to creatine derive from proton-1 MR spectroscopic imaging within the normal-appearing white matter. This steady state metabolic imaging also provides other metabolic parameters to differentiate tumor cells from normal brain, detect the presentence of IDH mutation and predict survival in lower grade glioma. In addition, stable and treatment-free lower grade glioma had impaired cognition and quality of life, with the severity associated with the history of treatment and the volume of T2 lesions. These results suggest that the use of multiparametric MRI could improve tumor delineation, identify patients at risk for specific deficits and provide an opportunity for intervention. The objective of this translational proposal is to utilize a novel multimodality MR protocol, which integrates dynamic and steady state MR metabolic imaging with diffusion, perfusion, and resting-state functional MRI to provide quantitative metrics on dynamic and steady-state metabolism, white matter integrity, blood volume, and functional networks, to evaluate cognitive functioning and quality of life in patients with lower grade astrocytoma. We will take advantage of our unique experience in proton-1 MR spectroscopic imaging, which has been implemented into routine clinical examinations, and hyperpolarized carbon-13 pyruvate imaging, where we performed the first-in-human [2-13C]pyruvate study to image real-time glycolysis and oxidative metabolism simultaneously, to assess tumor burden, cognitive functioning, and quality of life. Once the multimodality MR protocol has been established in Aim 1, we will evaluate the normal and abnormal brain changes during radiation therapy and then correlate these changes to impairments in cognitive functioning and quality of life in Aim 2. Aim 3 will examine signatures associated with recurrent tumors and evaluate the impact of tumor burden on cognition and quality of life. The results of the proposed study will be critical for assessing response to treatment, developing effective treatment strategies, and improving quality of life. Ultimately, it will provide effective clinical management of patients and aid neuro-oncologists in making timely decisions.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT There are ~1.4 million adults with congenital heart disease (ACHD) in the US and their number is increasing by 40,000-50,000/year due to improving pediatric CHD care. Up to 85% of the ACHD patients fail to establish or maintain ACHD specialist care which results in poor outcomes, the most vulnerable period for these gaps being at the time of transition and transfer from pediatric to adult healthcare. Transition includes fostering of patients’ knowledge of their CHD and of self-management and self-efficacy skills needed for lifelong management of chronic disease. Transfer is the event when a patient's care is taken over by the adult healthcare team. Since chronic management of CHD interferes with daily life activities (such as job, family planning, traveling), it is difficult for many of these adults to engage in their own care. Knowledge, self-management, self- efficacy, and patient activation are important skills for patient engagement, and strategies that enhance these skills are known to reduce gaps in ACHD care. Majority of the ACHD patients are young and own smartphones. This provides a unique opportunity to use mobile app-based intervention as a relatively inexpensive and scalable solution to support ACHD patient engagement skills. But, for its success, it is critical to incorporate theory of behavior change into its design. Thus, my central hypothesis is that an automated, interactive, mobile app-based intervention refined using ‘Capability, Opportunity, Motivation-Behavioral’ theory to evaluate the determinants of behavior can enhance skills known to support ACHD patient engagement and ACHD specialist visit. To test this hypothesis, Aim 1 will shed light on the pertinent features of a mobile app to support patient engagement skills using semi-structured interviews of the ACHD patients, clinicians, researchers, and clinic staff. Aim 2 will use an iterative process with inputs from an Advisory Board of ACHD patients, clinicians, and researchers to design and revise an automated interactive user-friendly app. Aim 3 will carry out a pilot study to determine feasibility and acceptability of the mobile app to enhance ACHD patient engagement skills. These aims will create foundational knowledge for future studies to determine effectiveness of a mHealth based intervention to support ACHD patient engagement and ensure ACHD specialist visit. Candidate is an ACHD cardiologist and health services researcher at UCSF. The candidate, her mentors and her scientific advisors have developed a comprehensive career development plan that includes training in mHealth-based behavioral interventions, qualitative methods, and clinical trials. With strong institutional commitment, the candidate is well-positioned to attain research independence.

NIH Research Projects · FY 2025 · 2022-09

Project Summary Persistent low-grade inflammation underlies the development of chronic diseases that are prevalent in the elderly. Adipose tissue displays unique susceptibility to age-related inflammation. Even healthy individuals accumulate visceral adiposity in older age, leading to increased systemic inflammation and reduced metabolic health. A strong candidate source of inflammation in aged adipose tissue is the resident immune compartment. Within every organ is a specialized repertoire of resident immune cells that is essential for tissue homeostasis and stress adaptation. In adipose tissue, resident immune cells coordinate responses to fasting and cold, and our prior work establishes that changes in adipose-resident immune cells impairs responses to both these challenges in old mice. But the mechanisms that drive adipose-resident immune dysfunction throughout the lifespan have remained elusive. This proposal will leverage new technological approaches to address this outstanding question. Peripheral immune cells accumulate intracellular defects during aging, leading to poor immune protection after vaccination or infection. Given that tissue-resident immune cells are seeded early in life and maintained through self-renewal, we expect them to be especially sensitive to age-related regulatory pressures. Importantly, immune cells are also sensitive to environmental cues, and extracellular cytokines and nutrient availability strongly influence the recruitment and function of immune cells. Therefore, we hypothesize that both immune cell-intrinsic and -extrinsic mechanisms cause dysregulation of the aging adipose-resident immune compartment. We will use an inducible fate-mapping strategy, combined with intra-vascular labeling, to study bonafide long-lived adipose-resident immune cells. We will use a viral vector-based proximity ligation approach to discover age-dependent changes in adipose secreted proteins and test how they impact resident immune cells. By comparing old and younger obese mice, we will isolate age-specific mechanisms of inflammation and adipose tissue dysfunction. We will use these results to develop new strategies that protect the aging adipose-resident immune compartment and prevent inflammation. Our methods overcome several longstanding obstacles in aging science: (1) knowing the “age” of an immune cell, and (2) pinpointing known cell sources of age-associated cytokines/secreted factors. With these new capabilities, our work stands to advance aging science and enable discovery of new disease mechanisms affecting any organ system.

NIH Research Projects · FY 2025 · 2022-09

Project summary: Uveal melanoma (UM) is the most common intraocular cancer in the United States and accounts for about 5% of all kinds of melanomas. Approximate 50% of UM patients develop metastases, predominantly to the liver with 100% mortality. UM lacks mutations in BRAF, NRAS, NF1 and KIT common to other melanoma types. Instead, over 90% harbor somatic activating mutations in the Gaq family members GNAQ or GNA11, with the remainder carrying mutations of genes also acting in the Gaq signaling pathway such as CYSLTR2, a Gaq- coupled GPCR, phospholipases C b4 (PLCb4), a direct effector of Gaq. Therefore, UM is genetically defined by activating mutations of the Gaq pathway. Despite dramatic successes in other melanoma subtypes, immune checkpoint inhibitors and targeted therapies have failed to demonstrate clinical benefits in UM, leading to an urgent need to develop novel and effective therapeutic regimens. The CYSLTR2->Gaq->PLCb-> protein kinase C (PKC) module is a linear signaling cascade that drives the essential MAP-kinase (MAPK) signaling for UM cell proliferation, making Gaq pathway the prime target for targeted therapy for this devasting disease. Although both GNAQ/11 and PKC inhibitors are very effective to suppress UM cell proliferation/survival in vitro, targeting either GNAQ/11 or PKC alone has shown limited efficacy in UM liver metastasis. Understanding the resistance to the Gaq pathway inhibition is of paramount importance to develop new strategies that work in the specific signaling context of a constitutively activated Gaq pathway. Our preliminary data show either GNAQ/11 inhibition or PKC inhibition yields strong upregulation of the Gaq-coupled receptor EDNRB, which when encountering its ligand EDN1 from the liver environment can lead to reactivation of MAPK, thus driving resistance to Gaq pathway inhibition in UM. In this proposal, we will evaluate that blocking endothelin signaling will increase the therapeutic efficacy of targeting oncogenic Gaq signaling, directly or downstream, using newly developed genetically engineered and xenograft models of UM metastatic to the liver. While the role of endothelin signaling in melanocyte development and melanoma progression is well documented, the nature of the feedback that upregulates EDNRB expression/signaling is not understood. We will utilize a combination of candidate approaches, RNAseq and phospho-proteomics to dissect the underlying mechanisms in UM cells and in melanocytes. Our preliminary data also show that secondary mutations in GNA11 can confer resistance to the Gaq inhibition. In this proposal, we will expand the emerging landscape of the mechanism underlying the adaptive and acquired resistance to Gaq pathway inhibition and identify rational therapy combinations to improve the therapeutic efficacy for metastatic UM. Using a combined genome-wide CRISPR/Cas9 synthetic lethality screen and phospho-proteomic screen we identified a lipid synthesis pathway that is essential for the survival of cells with Gaq mutation, which we will explore as entirely novel therapeutic target for UM.

NIH Research Projects · FY 2025 · 2022-09

Project Summary The aim of this phase 0/1 clinical trial is to assess the safety and preliminary efficacy of the novel, orally available, blood brain barrier (BBB) penetrant imipridone ONC206 in pediatric patients with malignant brain tumors including diffuse midline gliomas (DMG). We show that ONC206 exerts its activity in DMGs and medulloblastoma through activation of the mitochondrial protease ClpP, a serine protease that plays a central role in mitochondrial protein quality control by degrading misfolded proteins. ClpP activation through ONC206 lead to apoptosis involving upregulation of ATF4 and C/EBP homologous protein (CHOP) and induction of DR5 and TRAIL. In our preliminary analyses, we have found that ClpP expression correlates with ONC206 response in vitro and that ONC206’s effect is completely abrogated in H3K27M models with CRISPR/Cas9 knockout. Our in vivo data demonstrate that ONC206 leads to significant survival benefit in relevant in vivo model systems. Based upon this exciting preliminary data, this will be the first study to test the safety and to determine the recommended phase-2 dose (RP2D) of ONC206 as a single agent in pediatric patients including patients with DMGs and other recurrent malignant brain tumors. Further, we will test ONC206 as single agent and in combination with radiation in newly diagnosed and with re-irradiation in relapsed pediatric DMGs. We will assess the safety, PK profile and preliminary efficacy of ONC206 in four different cohorts: (1) in newly diagnosed DMG, (2) in DMG patients who completed radiation therapy, (3) in DMG patients at time of recurrence with re-irradiation and (4) other recurrent malignant brain tumors. Using a Bayesian optimal interval (BOIN) design the maximal sample size will be 3-42 per cohort. All cohorts will be expanded so that 12 patients per cohort have been treated at the RP2D. To investigate the BBB penetration of ONC206, 40 patients will be treated at the RP2D in a phase 0 component of the study, stratified by anatomic region since penetrance might differ based on tumor location (pontine, thalamic vs. other locations). Within the confines of a phase 1 study, we will also assess progression fee and overall survival. For each patient we will collect tumor tissue, CSF and serum for correlative studies with a focus CLpP expression as a potential biomarker of response. We will also assess tumor circulating DNA as potential marker for disease for treatment response. Further, we will assess quality of life and patient reported outcomes, which is critically important but has not yet been done comprehensively in this patient population. This will be the first study to comprehensively assess ONC206 in this pediatric patient population and will lay the groundwork for future combination trials.

NIH Research Projects · FY 2024 · 2022-09

PROJECT SUMMARY/ABSTRACT Intracellular pH (pHi) dynamics are a critical regulator of cell fate changes. Our lab and others demonstrated that an increase in pHi is necessary for the differentiation and lineage specification of multiple cell types, including mouse embryonic stem cells, Drosophila ovarian follicle stem cells, mouse intestinal stem cells, and chick paraxial mesoderm. Our lab has also shown how pHi dynamics regulate cell adhesion and migration behaviors by modifying the protonation state of pH-sensing proteins, including -catenin, cofilin, talin, and focal adhesion kinase. However, the mechanisms underlying pHi regulation of cell fate changes remain unresolved. Furthermore, our understanding of pHi dynamics during embryonic development in vivo is limited, largely due to lack of appropriate models. To address this, I generated a novel system for interrogating the functional significance of pHi dynamics during embryonic development in vivo using zebrafish, a genetically tractable organism suited for in vivo live cell imaging of embryos. Using this new model, I will determine the role of pHi dynamics in the development of cranial neural crest (NC), a highly conserved vertebrate embryonic cell population that gives rise to diverse cell types, including chondrocytes, osteocytes, and odontoblasts. Many cell behaviors involved in cranial NC development are regulated by pHi dynamics in other cell types, including lineage specification, cell migration, and epithelial to mesenchymal transition. Thus, cranial NC represents an ideal model for addressing the gaps in our understanding of pHi dynamics during embryonic development in vivo. By following early NC cells through the differentiation of cranial lineages, I will test the central hypothesis that pHi dynamics regulate cranial NC development, at the stage of delamination, migration, or lineage specification. In Aim 1, I will resolve spatial and temporal pHi dynamics during zebrafish cranial NC development by in vivo live cell imaging. My preliminary data indicate a higher pHi in migratory compared with premigratory cranial NC cells. In Aim 2, I will experimentally perturb pHi in zebrafish NC cells through pharmacologic and genetic modulation of plasma membrane ion transporters and determine the effect on cranial NC cell behaviors and transcription, thus establishing the functional significance of pHi dynamics during zebrafish cranial NC development. My findings have promise to reveal new insight on the cellular and molecular factors controlling craniofacial development, with important implications for human congenital diseases and tissue repair.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY / ABSTRACT The goal of this application is to train Dr. Brian Shy, a physician scientist at the University of California San Francisco, with the skills necessary to become an independently-funded investigator studying, developing, and manufacturing engineered cellular therapies. This research proposal will evaluate CRISPR-based non-viral methods for targeted genome, epigenome, and transcriptome engineering in primary human T cells. This will be applied to understand DNA repair pathway choice and DNA toxicity responses in order to improve site-specific “knock-in” strategies with large therapeutic constructs such as chimeric antigen receptors (CARs), develop safe and efficient methods for multiplexed T cell modifications, and establish Good Manufacturing Practice (GMP) compatible processes for clinical implementation of this diverse toolset. This research is accompanied by a career development and training plan that will build on Dr. Shy's clinical training in Transfusion Medicine and Cellular Therapy, and his prior expertise in the pre-clinical development and manufacturing of experimental cellular therapies. Training will focus on 1) implementation of novel cellular engineering tools, 2) next generation sequencing (NGS) technologies and analysis of large datasets, 3) GMP manufacturing and pre-clinical development, 4) clinical trial design and standard-of-care cellular therapies, and 5) laboratory management, grant writing, presentation, and research strategy skills. This career development and training plan will include a combination of formal coursework, mentored practical training; conference, meeting, and workshop attendance; and guidance from an exceptional team of co-mentors, advisors, and collaborators. Altogether, this award will help prepare Dr. Shy for an independent research career focused on developing improved cellular therapies for treatment of cancer and other human diseases.

NIH Research Projects · FY 2026 · 2022-09

PROJECT SUMMARY / ABSTRACT Up to 10% of older adults experience durable postoperative cognitive decline at 1 year or beyond after major noncardiac surgery, which is associated with up to 50% increased risk for Alzheimer’s disease and Alzheimer’s disease related dementias (AD/ADRD). However, most older adults will be cognitively unharmed by, and will enjoy longer life and/or improved function because of, surgery. Very little information about anticipated cognitive outcome is available to guide older adults considering elective surgery, because prior research has not incorporated longitudinal pre-surgical trajectories, focused on clinically relevant cognitive outcomes, or been designed to facilitate individualized predictions. Although it has been called “ethically imperative” that physicians discuss adverse postoperative cognitive outcomes as part of surgical informed consent, they lack accurate, individualized information to share with patients. In this study, we propose to use large, population- representative, longitudinal data sources to develop and validate clinical prediction models for long-term cognitive outcomes, including AD/ADRD, after elective major surgery in older adults. We will (AIM 1) use the Health and Retirement Study (HRS), linked to Medicare billing data, to develop clinical prediction models based on preoperatively-known factors to predict (a) memory, (b) risk of AD/ADRD, and (c) ability to independently manage one’s finances and medication, two cognitively-intensive instrumental activities of daily living (IADLs), at two years after surgery. Methodologically, we will balance model complexity with clinical practicality and use methods to account for selective survival. We will (AIM 2) augment the clinical prediction models with complications or adverse events that occur after surgery, e.g., delirium, hospital readmission, and new moderate-severe pain. Findings from this Aim will enhance understanding of “best case, worst case” cognitive outcomes (memory, AD/ADRD, and cognitive-functional) to include in preoperative cognitive risk discussion. These findings will also support future hypotheses about key mechanisms that lead to adverse outcomes, guiding priorities for post-surgical management to improve long-term cognitive outcomes. Finally, we will (AIM 3) externally validate the HRS models using the National Health and Aging Trends study, again linked to Medicare data, to assess generalizability of the models predicting memory, AD/ADRD, and IADL independence after surgery. After validation, these models can be used clinically to predict 2-year cognitive outcomes, including AD/ADRD risk, so that older adults have accurate, personalized information on the likely cognitive outcomes with or without surgery. This information will enable inclusion of long-term cognitive outcomes in surgical shared decision-making, a transformative advance for older surgical patients.