University Of California At Davis

NIH Research Projects · FY 2025 · 2023-07

Project Summary/Abstract Inherited retinal diseases (IRDs) are a major source of blindness worldwide. These diseases are typically single-gene disorders that result in the degeneration of photoreceptor cells. Historically they have been classified based on the clinical phenotype and electrophysiological results, and then grouped into various disease entities. In the past three decades the genetic basis of many forms of inherited retinal diseases have been discovered leading to the identification of over 270 retinal disease genes. Importantly, the field of medicine and vision science has resulted in an FDA-approved viral mediated gene therapy for IRD associated with mutations in RPE65. Children with this condition are treated with AAV carrying RPE65 delivered to the macula in the subretinal space at the time of vitrectomy. Many other single gene disorders affecting the retina are currently being targeted in various clinical trials, mostly through similar strategies to deliver virus to the macula through the subretinal route. Viral mediated gene therapy administered in this fashion is limited by treatment of only the macular area. Furthermore, it requires intraocular surgery, detachment of the macula from the retinal pigmented epithelium, and carries the risk of sight-threatening surgical complications. Recent studies have demonstrated progressive macular atrophy (retinal cell death) at the site of the subretinal bleb in ~15% of patients receiving this gene therapy within the first year after treatment, suggesting subretinal gene therapy may ultimately cause more harm than good. Therapeutic administration via the intravitreal route is much less invasive, takes place in a clinical setting, and can potentially be repeated to achieve maximal therapeutic effect. However, intravitreal viral mediated gene therapy can be associated with increased inflammation. In addition, native AAVs do not penetrate the retinal layers to transduce photoreceptors efficiently. In this study, we will take advantage of non-human primates with a mutation in PDE6C, a key component of phototransduction, causing electrically silent cone photoreceptors. With a proven effective therapeutic AAV-PR1.7-PDE6C vector in hand, we will compare the efficacy of AAV delivered via the subretinal vs intravitreal route to rescue cone function. Furthermore, we will compare inflammatory reaction between these contexts, determine the active components of the immune system, and define the role of pre- existing anti-AAV antibodies in host animals. We will measure the degree to which the route of delivery and inflammation affects the physiologic rescue. This proposal will advance the field of intravitreal gene therapy for inherited retinal diseases. The aims of this proposal will determine the efficacy of intravitreal compared to subretinal gene therapy, the role of pre-existing circulating antibodies in the host that may mitigate the efficacy of treatment, and the degree to which ocular inflammation affects the visual rescue. Together, the aims of this study will advance the understanding of intravitreal gene therapy and its relationship with ocular inflammation and treatment efficacy.

NIH Research Projects · FY 2024 · 2023-07

Spatial profiling of melanocytic tumors and their microenvironment Understanding tissue structure is fundamental for biological sciences and for distinguishing benign versus malignant neoplasms, but histopathological assessment alone is inaccurate for the diagnosis of certain tumors, including a subset of melanocytic neoplasms (melanocytic nevi and melanomas), resulting in diagnostic errors and worsened patient outcomes. Therefore, novel biomarkers for the diagnosis of melanoma are needed. To identify such markers, it is imperative to better understand the interaction between melanocytes and neighboring keratinocytes, immune cells, and other components of the complex tumor microenvironment in nevi and melanoma. Nevi and early primary melanoma display intratumoral heterogeneity, often coupled with low cellularity and purity. Therefore, the tumor-microenvironment interactions could be missed by bulk approaches or single-cell sequencing of advanced/metastatic tumors only, which has been the focus of most prior studies. The goal of this research is to build high-resolution spatial maps of gene expression of the tumor and its microenvironment in morphologically preserved nevi and melanomas to identify novel diagnostic biomarkers. The hypothesis is that melanocytic tumors and their microenvironments contain subpopulations of cells with characteristic gene expression patterns that differ between nevi and melanoma. We hypothesize that some of these differentially expressed genes are spatially confined and cell-type specific and could be used as potential diagnostic markers. Our prior data demonstrated differences in CDK2 gene expression between melanocyte- rich regions of nevi and melanoma. In Aim 1, we will assess spatial expression of CDK2 by immunohistochemistry in a tumor panel of over 200 nevi versus melanoma, comparing it to proliferative markers, as well as established melanoma biomarkers, including PRAME. In Aim 2, to identify novel biomarkers, we will establish high-resolution spatial maps of gene expression of tumor and microenvironment subpopulations in nevi versus melanoma by performing a spatial whole transcriptome analysis and a high-plex single-cell imaging. Top differentially expressed genes in these subpopulations will be validated via immunohistochemistry in the tumor panel described above. This study improves current theoretical concepts by investigating tumor and microenvironment populations in nevi and early primary melanoma – the common, yet previously understudied tumor types. Furthermore, this study utilizes improved state-of-the-art approaches, including high-plex single-cell spatial gene expression profiling. This research will improve the understanding of tumor-microenvironment subpopulations in their spatially correct context, relevant for tumor biology and biomarker development, ultimately leading to improved diagnostic accuracy of melanoma and improved patient outcomes.

NIH Research Projects · FY 2024 · 2023-07

Project Summary Organisms adapt to seasonal changes in environmental conditions to survive. These adaptations rely predominantly on photoperiod (i.e., daylength), but are also influenced by temperature. Recent studies indicate that photoperiodic changes affect the neuronal composition of brain areas involved in circadian (i.e., daily) timekeeping and modulate the number of dopaminergic neurons, in a process known as neurotransmitter switching. Other studies show that the brain also undergoes profound structural changes across seasons. However, the relationship between these functional and structural changes in the brain and seasonal adaptations remains a major gap in knowledge. Moreover, whether other relevant seasonal cues, in particular temperature, contribute to these changes is not known. The overall goal of this project is to understand the nature and role of neuronal plasticity in the integration of seasonal cues to promote seasonal adaptations. My hypothesis is that seasonal adaptations are mediated by functional and structural plasticity in neurons from circadian and aminergic circuits in response to environmental cues. To test this, I propose 3 specific aims: investigate structural and functional plasticity of (1) the circadian clock neuronal network and of (2) aminergic circuits in response to seasonal cues and its impact on social and locomotor behavior, and (3) determine how the plastic changes in the circadian clock and aminergic circuits regulate brain connectivity and encode the behavioral output of these circuits. I will accomplish this project in the genetically tractable Drosophila model and will leverage a combination of versatile neurogenetics, high-resolution microscopy, and well-established behavioral analysis. Thus far in my postdoctoral career in the Chiu lab at UC Davis, I obtained training in molecular genetics and biochemistry, which I used to explore the role of circadian peptides in modulating seasonal adaptations in Drosophila. Moving forward, I will build on my current research to study the neuronal mechanisms of seasonal plasticity and behavior. During the K99 training period, I will use available tools in Drosophila to assess the functional and structural changes in the circadian clock neurons and aminergic circuits in response to seasonal cues. Moreover, I will test the functional consequences of these changes by using available genetically encoded sensors and by generating new, more sensitive, sensors to assess aminergic function in vivo under the guidance of Dr. Lin Tian. I will expand the use of these tools in the R00 stage to determine how the interaction between these two circuit systems modulate their functions and how they affect seasonal behavior concertedly. I believe that the mentorship of Drs. Chiu and Tian, together with the support provided by the K99/R00 award, will allow me to build a strong foundation that will enable my success as an independent investigator. The results of the proposed studies will elucidate the neuronal basis underlying sensory integrations in the context of seasonal adaptations, shedding light on the mechanisms behind seasonal modulation of health physiology and disorders.

NIH Research Projects · FY 2024 · 2023-07

Giardia lamblia is a widespread protozoan parasite of humans and animals causing significant diarrheal disease worldwide. Giardia cysts are transmitted between hosts through feces. Once ingested, cysts transform into trophozoites that colonize the duodenal epithelium – a region of varied nutrients, redox stress, and immune responses. Unknown external cues in the gut trigger trophozoites to encyst, and then cysts are passed to new hosts. Giardia is also reported to have a quasi-meiotic stage, termed diplomixis, that occurs in late encystation and is characterized by fusing and exchange of DNA between cyst nuclei. Due to a historical lack of molecular genetic tools, the pathogenesis and basic biology of Giardia is grossly understudied. Developmental transitions and environmental responses are commonly mediated by transcription factors (TFs) – modular proteins that often contain DNA binding domains (DBDs). However, only 36 TFs with DBDs are predicted in the Giardia genome as compared to about 200 predicted in similar sized eukaryotic genomes. Over 500 proteins have more general motifs such as basic leucine-zipper domains, and these can also function as TFs. Only a handful of Giardia TFs have been studied in any detail; all are associated with early to mid-encystation. Basal TFs, redox responsive TFs, late encystation/diplomixis TFs, or cell cycle related TFs have not yet been identified. Our lack of knowledge of the identities and functions of Giardia TFs has severely limited our understanding of genetic regulatory networks throughout the Giardia life cycle, particularly those associated with cell division and pathogenesis. To address these deficits, we will use an unbiased, high-throughput, genome-wide yeast one-hybrid (Y1H) screen to identify additional DNA binding proteins likely representing candidate TFs (Aim 1). In many systems, such screens are commonly used to define genome-wide protein-DNA interactions. Using robotic screening of arrayed “prey” libraries, we will screen a total of nine promoter “baits” (three constitutive loci; three redox stress- associated loci, and three encystation loci) against three “prey” cDNA libraries (constitutive; redox stress; late encystation/diplomixis) for a total of 27 (3 x 3 x 3) combinations. Both predicted DBD proteins and candidate TFs identified in the Y1H screen will be additionally prioritized by CRISPRi mediated knockdown, with phenotypic screening for defects in growth, mid-encystation and genetic marker exchange in cysts (diplomixis), and redox stress. Knockdowns of candidate TFs with strong defects will be further characterized using promoter-luciferase assays and a novel dCas-TF luciferase fusion assay to evaluate transcriptional activation or repression (Aim 2). Overall, this project will define and prioritize Giardia TFs as groundwork for future interrogation of Giardia regulatory networks throughout the life cycle.

NIH Research Projects · FY 2025 · 2023-07

PROJECT SUMMARY Metastasis is responsible for the majority of breast cancer deaths, and standard-of-care treatments fail to effectively target metastasizing cells. Circulating tumor cells (CTCs) in the bloodstream rely on the physical protection and chemical signals of platelets to survive and seed metastatic lesions. One such chemical signal is transforming growth factor beta (TGF-β), which, after secretion by platelets, has been shown to modulate CTC gene expression and behavior. TGF-β has also been shown to upregulate expression of the cell-surface glycoprotein Mucin-4 (MUC4) in various cellular contexts. MUC4 has been implicated in tumor development and maintenance and was recently observed to contribute to platelet-CTC interactions. This raises the question of whether platelet-secreted TGF-β may be upregulating CTC-MUC4, enhancing platelet-CTC interaction and generating a positive feedback loop. The hypothesis driving the proposed studies is that platelet-TGF-β upregulates CTC-MUC4, reinforcing CTC-platelet binding and enhancing metastatic cell survival. Specific Aim 1 will determine the effects of platelet-derived TGF-β on tumor cell MUC4 expression using cellular, molecular, and biochemical techniques, and assess MUC4-dependent cellular aggressiveness in vitro. Specific Aim 2 will characterize the role of MUC4 in platelet-tumor cell interactions using in vitro binding assays. Specific Aim 3 will assess the effects of platelet-TGF-β and CTC-MUC4 crosstalk in vivo using tail vein and orthotopic xenograft tumor mouse models of metastasis. Successful completion of this research will reveal a novel form of platelet- CTC crosstalk, exposing an important means by which metastasizing cells survive and illuminating a potential new therapeutic target for breast cancer metastatic prevention.

NIH Research Projects · FY 2025 · 2023-07

Project Summary The general goal of the proposed research is to define the mechanistic underpinning for the synthetic lethality network between a deficiency in homologous recombination (HR) and defects in single-strand annealing (RAD52) or POLq-mediated end-joining (TMEJ). Exciting preliminary data revealed that BRCA2 and RAD52 delay the repair of S-phase-associated DNA double-stranded breaks (DSB) by TMEJ until M-phase. This regulation avoids TMEJ-mediated chromosomal rearrangements of one-sided DSBs that are produced by replication fork breakage. Our approach combines innovative cell cycle phase resolved imaging of DNA repair proteins and DNA damage markers with mechanistic biochemical analysis using purified human proteins in reconstituted reactions. Our results will have potential translational implications for the clinical application of newly developed RAD52 and POLq inhibitors for the treatment of HR-deficient tumors with respect to application protocols, patient selection, and use of DNA damage response checkpoint inhibitors as well as the response to poly(ADP-ribose) polymerase inhibition. The Specific Aims are: 1. Define the mechanism of action of BRCA2 in DSB repair pathway control. We will test the model that the DNA binding properties of BRCA2 are critical for TMEJ inhibition. In Aim 1A, we conduct foundational studies to determine the fundamental DNA binding properties of full-length BRCA2. In Aim 1B, we will define which domains of BRCA2 are required for TMEJ inhibition in cells. This combination of cell-based and biochemical studies will define the functions and regions of BRCA2 that are required for TMEJ inhibition. 2. Define the mechanism of TMEJ inhibition by BRCA2 and RAD52. BRCA2 and RAD52 employ two different modes to inhibit the DNA polymerase activity of POLq which may affect additional reaction steps in the TMEJ process. We will reconstitute TMEJ in vitro with purified proteins to determine the mechanisms by which BRCA2 (Aim 2A) and RAD52 (Aim 2B) inhibit TMEJ. We will test inhibition of the overall TMEJ reaction and individual steps including 1) DNA binding, 2) end-alignment, and 3) DNA synthesis. Analysis of wild type and catalytic mutants of POLq will be conducted in vitro and in cells. 3. Define which HR defects are susceptible to RAD52 loss of function. It is an open question whether loss of RAD52 will lead to POLq-mediated chromosome fusions and lethality in all HR-deficient backgrounds (Aim 3A) or all BRCA2 mutants (Aim 3B). Our preliminary studies suggest a model that loading of BRCA2 is the critical step to limit TMEJ to M-phase and that HR defects past this step are not affected by RAD52 inhibition.

NIH Research Projects · FY 2025 · 2023-06

Abstract Patients with concerning breast masses require diagnosis, triage, and management. This proposal seeks to combine three recently developed technologies to provide rapid diagnostic support for patient breast tumor evaluation. Currently, this process consists of biopsy, standard histology, phenotype determination via im- munohistochemistry, and increasingly, molecular assays. In well-served areas, the challenges involve obtain- ing appropriate and representative biopsies, waiting for slide preparation, review by a pathologist, then im- munostaining (IHC), and optionally, submission for nucleic-acid based studies. We propose a simple tissue-direct-to-digital slide-free imaging solution (FIBI, fluorescence imitating brightfield imaging, UC Davis) coupled with a virtually hands-off millifluidic specimen handling device (CoreView, UW). Images, intrinsically digital, are of diagnostic quality, and can be interpreted locally by trained pathology per- sonnel, or streamed anywhere in the world for real-time evaluation. Eventually, on-site or cloud-based com- puter-assisted tools will be available. An additional goal is to achieve same-session multiplexed immunostain- ing and imaging, based on rapid-quench click chemistry approaches developed by Dr. Ko (Univ. Penn.) We will build and test versions of the combined imaging and automated sample handling of needle-derived specimens initially using excess fresh or recently fixed human breast tissue, and subsequently clinically ac- quired core-needle biopsies. Tasks involve optimization of staining, imaging, image display, reproducibility demonstration, and sample integrity assessments. Incorporation of single-step immunostaining with far-red la- bels will be included, and methods for rapid probe cycling (for multiplexing—ER, PR, HER2) will be applied and tested. Subspecialty pathology review will ensure that the FIBI images and immunostains are reliable and of diagnostic quality. The goal is to implement a context-appropriate, automated instrument that can capture his- topathology images from core-needle biopsies, while retaining their integrity for downstream processing as needed. Results will be available within minutes and allow for informed patient triage or even immediate on-site treatment, especially valuable if the patient has had to travel long distances to reach the clinic. Statement of Potential Impact: Current biopsy evaluation interposes at least 1-3 days delay, and involves consuming some or all of small tissue specimens. Immunostaining contributes additional delay. The situation is more challenging in poorly served areas, as histology laboratories and the pathologists capable of interpreting the results are rare outside major urban centers. The CoreView-FIBI approach could dramatically improve pa- tient-level care velocity, but also enterprise-level efficiency, allowing for same-day disease treatment planning. For biospecimen applications, preanalytical variables will be under complete control, the non-destructive nature of the imaging means that more tissue is available for storage and research, and imaging of the specimen prior to banking provide assurance of tissue content and abundance estimates of relevant components.

NIH Research Projects · FY 2026 · 2023-06

PROJECT SUMMARY/ABSTRACT Black Americans experience a disproportionate burden of cardiovascular disease (CVD) risk factors and Alzheimer's disease and related dementias (ADRD). Studies have identified midlife (~ ages 45-65) dyslipidemia as a risk factor for ADRD. Black Americans have more favorable lipid profiles compared to Whites or Latinos, but their incidence of dyslipidemia in midlife is higher. This seemingly paradoxical relationship between favorable midlife lipid profiles yet high incidence of midlife dyslipidemia and high risk of ADRD among Black Americans has been severely understudied. Lipids play a vital role in neurodegenerative disease, and epidemiologic and metabolomic studies have identified commonly tested lipids (total cholesterol, HDL cholesterol, LDL cholesterol, and triglycerides) as well as sphingolipids (ceramides) as predictors of cognitive aging. Prior work has largely overlooked the contributions of lipids to ADRD risk in Black Americans and few studies have examined the relationship of ceramides with cognitive aging and ADRD in this high-risk group. This project will leverage over 40 years of longitudinal data from two NIH/NIA funded cohort studies, the Kaiser Healthy Aging and Diverse Life Experiences (KHANDLE) Study and the Study of Health Aging in African Americans (STAR), to better understand the role of lipids in cognitive aging and ADRD among Black Americans. The scientific objective of this research plan is to characterize the relationship of early adulthood (~ age 30) total cholesterol, HDL cholesterol, LDL cholesterol, and triglycerides as well as the novel lipid biomarker, ceramides, with late life cognitive decline, ADRD, and MRI markers of neurodegeneration and vascular brain injury in an all-Black cohort of older adults. The proposed research seeks to: 1) define the role of early adulthood lipids in cognitive aging and ADRD; 2) examine genetic (APOE) and psychosocial (racism) factors as potential effect modifiers; and 3) determine whether the relationship of early adulthood lipid levels with late life cognitive decline and ADRD is partially mediated by midlife (~ ages 45-65) cardiometabolic disease (hypertension, diabetes, obesity, and dyslipidemia). For aging Black Americans with high prevalence of CVD risk factors and at disproportionate risk of dementia, prevention of ADRD has enormous health and economic consequences. This research will be complimented by a detailed training plan based at the University of California, Davis, with guidance from an exceptional mentorship team of nationally and internationally recognized ADRD, lipids, dementia, and cognitive aging researchers. The training will build upon the applicant's background in CVD and dementia epidemiology by incorporating specialized training in lifecourse theory and modern causal inference methods, biology of lipids, psychometric testing, and measurement and modeling of neuroimaging biomarkers. The combined research and training will prepare the applicant to successfully transition to an independent researcher of disparities in vascular contributions to ADRD.

NIH Research Projects · FY 2026 · 2023-06

PROJECT SUMMARY/ABSTRACT Evidence from human imaging, postmortem analysis, and animal models suggests that atrophy of neurons in the prefrontal cortex (PFC) plays a key role in the pathophysiology of both neuropsychiatric and neurodegenerative diseases. Structural changes—including retraction of dendritic arbors, loss of dendritic spines, and reductions in synapse density—lead to functional deficits that manifest as impaired cognition, decreased motivation, anhedonia, high anxiety, and increased impulsivity. Thus, therapeutic strategies aiming to restore PFC structure/function have broad therapeutic potential. Psychoplastogens—small molecules that promote structural and functional neuroplasticity in the PFC—produce both rapid and long-lasting therapeutic effects after a single administration. However, many psychoplastogens, including ketamine and serotonergic psychedelics, induce hallucinations, which greatly limit their therapeutic potential and clinical scalability. Fortunately, increasing evidence suggests that the hallucinogenic effects of ketamine and psychedelics may not be necessary for their therapeutic properties, and our group recently introduced the first non-hallucinogenic psychoplastogens. The advent of non-hallucinogenic psychoplastogens represents an exciting new direction for the treatment of many brain disorders, but there is an urgent need to further optimize their efficacy and safety profiles. Our primary goals are to, 1) establish robust synthetic strategies to psychoplastogenic natural products and chemical scaffolds that are amenable to medicinal chemistry, 2) develop high-throughput cellular assays for assessing psychoplastogen efficacy and safety, and 3) advance new in vivo assays uniquely suited to evaluate the long-lasting effects of psychoplastogens. Taken together, these efforts will enable structure- activity relationship (SAR) studies of key psychoplastogenic scaffolds, filling the gap in our knowledge about which structural motifs are critical for both psychoplastogenic and hallucinogenic effects. Ultimately, the work described here will enable the rational design of safer, non-hallucinogenic alternatives to psychedelics for treating a wide variety of neuropsychiatric and neurodegenerative diseases.

NIH Research Projects · FY 2026 · 2023-05

Therapeutic HIV vaccines confront an immune system whose anti-pathogen responses are established and have usually been evolving for years. T-cell responses to immunodominant epitopes have been generated and may have been maintained—or could have forced viral escape and eventual expansion of T cells with alternative, originally sub-dominant specificities. Many HIV-specific T cells have low proliferative capacity, rendering them incapable of a robust response when antiretroviral treatment is stopped. Furthermore, the established T-cell repertoire failed to control acute infection, so expansion of pre-existing T cells with their attendant functional deficiencies is unlikely to be therapeutically effective. The primary goal of therapeutic vaccination must instead be expansion of new T-cell clones with superior function, and/or the restoration of function to pre-existing memory cells. We suggest that only such qualitative improvements can provide the host with new capacity for control over the virus. A central objective of our program is to understand the extent to which pre-existing host immune and metabolic features constrain the quality of T-cell responses to therapeutic vaccination. Which immunometabolic conditions predict and perhaps foster qualitatively superior responses? What fraction of the T-cell response to vaccination is represented by new and previously undetected clonotypes, and how do the functional capacities and differentiation of the new clonotypes differ from pre-existing ones? A second major objective is to evaluate the relative ability of different vaccine regimens and metabolic interventions to expand new HIV/SIV-specific T cells with stem-like qualities. We and others have shown that HIV-specific T cells in natural HIV controllers express high levels of the memory-promoting transcription factor, TCF-1, retain proliferative capacity, and exhibit metabolic plasticity. We hypothesize that vaccine- induced cells with stem-like properties often derive from naïve T-cell clonotypes not previously expanded or chronically exposed to antigen, and that different vaccine regimens differ in their ability to recruit such cells. A third and central objective of our effort is to learn how peptide specificity, stemness, and metabolic capacities of T cells responding to vaccination are related to control over viremia during ATI. A large literature supports our premise that high T-cell quality is required for control over viremia, and more specifically that T cell memory, or “stemness”, features are associated with effective host responses. However, these connections remain relatively unexplored in the context of therapeutic vaccination, in part due to scarcity of large therapeutic-vaccine studies that have yielded an appreciable efficacy signal. We will use samples from human and non-human primate therapeutic-vaccine studies that have shown evidence for T cell-mediated virologic suppression to understand if the T-cell features previously linked to control over infection are also typical of successful immune responses to therapeutic vaccines.

NIH Research Projects · FY 2026 · 2023-05

PROJECT SUMMARY Breast cancer is the most common cancer in women and the second leading cause of cancer death in the United States (US). Although women included in the US Census racial/ethnic categories Hispanic/Latina (H/L) and Asian American/Pacific Islander (AA/PI) have relatively low breast cancer incidence compared to non-Hispanic White (NHW) women, multiple studies have reported a higher proportion of human epidermal growth factor receptor positive (HER2+) tumors in these groups (18-30%) compared to NHWs (14-18%). Expression of HER2 is clinically significant because it determines if a patient can receive targeted treatment. HER2+ disease, independent of hormone receptor (HR) status, is also associated with poor outcome compared to the most common HR+ HER2- subtype. The use of European-centric data to predict cancer risk and prognosis in non- Europeans remains a critical barrier for equity in the implementation of precision medicine. Overall, there is a gap in knowledge regarding the genetic factorsand molecular correlates relevant to the etiology of HER2+ breast cancer in diverse populations. Supported by a) our previous work showing a consistent association between Indigenous American ancestry and HER2+ subtypes in H/L breast cancer patients, b) the higher proportion of HER2+ tumors described in AA/PI and Asian populations, c) the closer genetic distance between these populations relative to European groups, and d) promising preliminary data, we hypothesize that germline variants more common in Indigenous American and Asian genomes contribute to the higher risk of HER2 amplification/expression in breast tumors of individuals with these ancestries. To test this hypothesis, we propose to integrate and leverage our own existing studies for a total of 17,049 cases (~4,200 HER2+) and 15,409 controls for discovery, and the NCI’s Confluence Project Data for replication (~600,000 cases and controls combined). Our main goal is to discover germline genetic variants contributing to the higher incidence of HER2+ breast cancer in women of Latin American and Asian heritage. To achieve this goal, we will 1) identify germline variation associated with HER2+ breast cancer in H/L and Asian women and replicate across diverse ancestries, 2) develop, validate, and test trans-ancestry and ancestry-specific polygenic risk scores for HER2+ disease, and 3) identify genes associated with HER2+ breast cancer risk in H/L and Asian women. The results of our study will lead to the discovery of genetic factors contributing to the observed HER2 subtype-ancestry association in women of Indigenous American and Asian ancestry for improved prediction, and provide a better understanding of HER2+ tumor etiology, which could lead to improved precision prevention strategies and the development of new targeted treatments for HER2+ disease.

NIH Research Projects · FY 2025 · 2023-05

PROJECT SUMMARY/ABSTRACT Although immunotherapy, especially immune checkpoint inhibition (ICI) with PD-1/PD-L1 inhibitors, has rapidly become the fourth pillar in cancer therapy with increasing breakthrough advances, barriers still exist to its success. Given their ability to rapidly exert their cytotoxic effects on heterogeneous tumor cells with minimal adverse events, natural killer (NK) cells have emerged as promising tools to expand the benefits of cancer immunotherapy, including for patients who never start or stop responding to ICI. However, lack of consistent responses in human NK cell trials, especially for solid tumors, calls for innovative methods to successfully translate novel NK immunotherapy approaches to the clinic. Dogs with cancer are an excellent way to assess novel immunotherapies because they recapitulate fundamental clinical and genetic features of human cancers, including the development of spontaneous tumors in the setting of an intact immune system. To speed translation of NK immunotherapy approaches, the proposed project will test an innovative treatment of allogeneic NK adoptive transfer in combination with a novel caninized anti-PD-L1 antibody developed by our comparative oncology group. Using a co-clinical Phase II trial format, dogs with locally advanced melanoma will be treated with radiation therapy (RT), adoptive transfer of expanded/activated allogeneic NK cells from healthy beagle donors, and immune checkpoint blockade using our dog anti-PD-L1 antibody. As the first trial to use NK cell transfer in combination with ICI on spontaneous tumors in a clinical setting, the results of this study will provide potentially transformative insights into mechanisms of both therapies and will evaluate barriers for future first-in- human trials on solid tumors. Since NK cell activity is known to be mediated by the PD-1/PD-L1 axis with PD-L1 being a critical inhibitory NK marker, the proposed study will offer critical insight into potential mechanisms of overcoming NK dysfunction responsible for unimpressive responses with NK cell immunotherapies alone. Furthermore, RT is part of the standard of care for unresectable malignancies and has been shown to have important immunomodulatory effects, including sensitization of tumor cells to NK cytotoxicity. The Canter Lab is a leader in canine clinical trials as well as their use as tools to perform multidimensional analyses of NK cells. Similarly, the potentially high impact of this novel immuno-oncology (IO) therapy will be studied through extensive correlative studies including flow cytometry to follow the regional differences of donor and endogenous NK cells, killing assays to assess changes in cytotoxicity, and RNA sequencing to characterize differential gene expression of relevant immune populations. Although we hypothesize meaningful clinical and immunologic effects from this novel therapy, we will nevertheless gain key insights into the dog as a comparative model for future dog and human IO studies. Thus, beyond the potential for significant scientific and clinical impact, the completion of this study will provide me with cutting edge training in comparative cancer immunotherapy to prepare me for a successful career as a veterinary scientist in cancer immunology and NK immunotherapy.

NIH Research Projects · FY 2026 · 2023-04

Abstract Several clinical variables are associated with outcomes following ischemic stroke (IS). However, clinical and demographic parameters account only for a portion of the outcome variance, thus it is difficult for clinicians to reliably predict long-term IS outcome. Hence, new biomarkers are needed. Molecules in blood are also associated with IS outcome including pro-inflammatory cytokines, anti-inflammatory cytokines and others. Genetic risk factors have also been associated with IS outcome. Unfortunately, combined blood and genetic biomarkers have not improved IS outcome predictions compared to clinical parameters since age, sex and initial NIHSS is said to predict outcome with a modest c-statistic approaching 0.7. Our preliminary data show gene expression in blood after IS can predict 90-day outcome better than age, sex and NIHSS. We hypothesize that a whole-genome approach of measuring RNA, which reflects the genome × environment × lifestyle interaction, to assess inflammatory, trophic and clotting genes will improve IS outcome prediction compared to clinical features alone. Thus, we propose the following aims: Aim #1a. Perform whole-genome RNA sequencing (RNAseq) of blood on a derivation cohort of IS patients at 1 day and 3 days after IS compared to matched vascular risk factor controls (VRFC). Aim #1b. Identify the most significantly regulated genes and pathways in blood at 1d/3d after IS that correlate with outcome, as measured by three outcome scales – mRS (modified Rankin Scale), NIHSS (NIH Stroke Scale) and Barthel Index at 90 days after IS. Aim #1c. Use Network Analysis to identify key hub genes after IS associated with outcome that might be causative. Aim #1d. Use Feature Engineering and Logistic Regression and/or other Machine Learning approaches, such as Support Vector Machines (SVM) and SVM Regression, to identify the least number of genes at 1 and/or 3 days that predict the three stroke 90-day outcome measures. Aim #1e. In a separate validation cohort of IS patients perform RNAseq to obtain the expression of the biomarker genes from Aim #1d. Input these into Machine Learning algorithms to predict patient 90-day outcomes (mRS, NIHSS, Barthel Index). Aim #2a. Demonstrate that gene expression is a better 90-day outcome predictor compared to each or a combination of clinical variables, such as stroke volume, initial NIHSS, location, etiology, age, sex, glucose levels, blood pressure, atrial fibrillation, and neutrophil, monocyte, lymphocyte, and platelet counts. Aim #2b. Delineate the underlying biology of these clinical outcome contributors by identifying genes and networks that correlate with them and pinpoint which of the genes/networks also correlate with each of the three outcomes. Significance: The findings of this study will develop biomarkers of ischemic stroke outcome to help clinicians predict IS outcome and aid future clinical trials in stratifying IS patients, thus significantly increasing chances for trial success. Equally important, the findings will provide much needed potential new treatment targets and unprecedented knowledge of how the peripheral blood transcriptome contributes to outcome and improve our understanding of the biology of repair and recovery after IS in humans.

NIH Research Projects · FY 2026 · 2023-04

Project Summary This career development award will provide the applicant (Alyssa Weakley, PhD) with the necessary training, knowledge, and experience to transition to an independent research career. Dr. Weakley’s overall career goal is to develop and validate accessible and innovative technology-based interventions that improve everyday functioning in individuals with Alzheimer’s disease and related dementias (ADRD), caregiving needs of informal family caregivers, and the relationship/bond of the care dyad. Dr. Weakley will build toward this goal with training and mentorship in (1) methods for identifying stakeholder (i.e., caregiver, care receiver) needs regarding technology use, adoption, and adherence; (2) developing inclusive (in terms of race/ethnicity) and accessible (in terms of technology familiarity) caregiver interventions; (3) randomized controlled trials (RCTs) of behavioral interventions; and (4) multidisciplinary team leadership and technology-based data management. Training aided by expert mentorship in these four areas will provide Dr. Weakley with the knowledge and skills to complete three research aims, the focus of the proposed research project. These aims include (1) identifying approaches to help cognitively impaired care receivers and distance caregivers engage with technology via stakeholder focus groups; (2) iteratively adapt and refine, through stakeholder co-design and participatory research methods, an intervention that facilitates use and integrates a remote caregiving web-based tool (Interactive-Care (I-CARE)) into everyday life; and (3) implementing a pilot RCT examining the feasibility and acceptability of the I-Care target intervention relative to basic I-Care training alone. The proposed research is significant because it couples innovative remote caregiver technology with stakeholder engagement to develop and test a novel intervention designed to improve outcomes for care receivers, caregivers, and their dyadic relationship. No research, to the applicant’s knowledge, has utilized a co-design approach for technology training that encapsulates remote caregiver and care receiver needs. Results will serve as pilot data for an R01 study that further investigates the most robust, scalable, and accessible approaches to improving functional outcomes in older adults with ADRD and caregivers. To complete her research and training aims, Dr. Weakley will leverage existing resources with expertise at three centers at University of California Davis: National Institute on Aging funded Alzheimer’s Disease Research Center, the Healthy Aging in a Digital World initiative, and the Family Caregiving Institute. These resources, along with the mentoring team’s strong commitment to Dr. Weakley’s research program, will provide Dr. Weakley with an enriched environment that will undoubtedly help her achieve her career development goals.

NIH Research Projects · FY 2026 · 2023-04

PROJECT SUMMARY Racial/ethnic minorities are at a higher risk for age-related cognitive dysfunction compared to non-Hispanic (NH) Whites. The mechanisms underlying this disparity are not well understood. Exposure to air pollution may be one important driver of these disparities. Recent epidemiologic studies suggest that increased exposure to air pollution increases risk of Alzheimer’s disease and related dementias (ADRD). Compared to NH Whites, racial/ethnic minorities are more likely to live in neighborhoods with higher levels of air pollution. No prior research, however, has quantified the contribution of air pollution as a driver of racial/ethnic disparities in ADRD. The scientific objective of this research is to evaluate air pollution as a potential driver of racial/ethnic disparities in ADRD and to understand the potential implications of air quality regulations on ADRD risk. We will focus this research on disparities between Mexican American and NH White racial/ethnic groups. This project uses an innovative approach to combine data from two existing cohort studies: The Sacramento Area Latino Study on Aging and the Ginkgo Evaluation of Memory Study. We will harmonize outcome and exposure data from the two cohorts and examine whether the effect of air pollution on ADRD risk varies across racial/ethnic groups. We will also examine whether air pollution drives disparities in ADRD by examining racial/ethnic differences in air pollution exposure, individual- and neighborhood-level susceptibility to air pollution. Finally, we will measure the potential impacts of air quality regulations on ADRD risk, using air quality regulations related to seaports as an example. The proposed research will leverage advanced causal inference methods including econometric quasi-experimental study designs to carry out the proposed research. This research plan is complemented by a mentored training plan that builds upon the candidate’s background in environmental epidemiology and includes coursework, structured mentoring, and experiential learning in the following areas: 1) racial/ethnic disparities in ADRD, 2) environmental health disparities, 3) harmonization of cognitive data, 4) air quality regulations, and 5) application of advanced social epidemiology and causal inference methods. Together, the proposed research and training plan will prepare the candidate for a successful independent research career dedicated to identifying environmental determinants of healthy aging, especially in racial/ethnic minority populations.

NIH Research Projects · FY 2026 · 2023-04

Imaging of brain oxygen extraction fraction in vascular contributions to dementia Vascular pathology is increasingly recognized as a major contributor to cognitive impairment and dementia, and arises from many pathophysiological mechanisms including endothelial damage of cerebral vessels, hypoxia, and blood-brain barrier breakdown. Vascular abnormalities have been identified as white matter hyperintensities (WMHs) on structural magnetic resonance imaging (MRI) scans, but the heterogeneity of and mechanisms underlying WMH evolution remain unclear. This lack of fundamental understanding of WMHs to predict cognitive impact is in large part due to limited imaging technologies to directly assess brain pathophysiology in clinical settings. In this project, we develop and optimize a new MRI technique to assess a critical parameter of brain vascular health, oxygen extraction fraction (OEF), in elderly patients with WMHs indicating the presence of cerebrovascular disease. In ischemic brain disorders, vulnerable tissue around an infarct compensates for reduced blood flow by increasing OEF, such that pathologically high OEF is an important indicator of a “penumbra” of at-risk tissue. However, MRI tools to measure OEF are limited by low signal-to-noise ratio and biological confounds on the MRI signal, and have not been fully tested in elderly patients at risk of cognitive impairment. We aim to address these limitations by using a novel, clinically feasible MRI method to map OEF in brain tissues. This quantitative BOLD (blood oxygenation level dependent) technique adopts a unique MRI acquisition in synergy with a biophysical model of microvessels in each voxel to quantify OEF. Using quantitative BOLD MRI in patients with vascular contributions to dementia (VCID), we will 1) characterize OEF abnormalities in WMHs and surrounding penumbra for different brain locations and their relationship to cognition; (2) associate longitudinal OEF changes with microvascular MRI markers of white matter injury to establish an ischemic pathophysiological mechanism of WMH evolution; and (3) test the hypothesis that OEF changes in white matter have long-range effects on functional connectivity within brain networks that support episodic memory and executive function. Successful completion of this project will provide critical biological knowledge about oxygenation in WMHs and link multiple vascular biomarkers in a mechanism for WMH progression over time. The novel OEF MRI approach also shifts the paradigm in imaging of VCID toward quantitative, physiologically- specific measures of vascular risk. Ultimately, non-invasive OEF imaging may detect early microvascular changes that drive neuronal injury in cognitive impairment and dementia, and serve future longitudinal studies of interventions to prevent cognitive decline due to vascular disease.

NIH Research Projects · FY 2026 · 2023-04

ABSTRACT Metabolic stressors and pathologic conditions such as diabetes can lead to pronounced physiologic alterations in protein regulation, resulting in muscle atrophy, which is highly associated with morbidity. However, the mechanisms underlying this association remain elusive. Understanding the mechanisms of maintaining muscle protein content is crucial to improving insulin resistance and overall metabolic health. Adipose tissue is an endocrine organ that responds to acute and chronic energetically challenging conditions by secreting proteins, referred to as adipokines. Evidence suggests that signaling mediated by adipokines plays a crucial role in organ crosstalk and systemic energy metabolism. Because of their direct signaling action, they can be developed as therapeutic agents for metabolic diseases. While considerable effort has been made to target the adipose tissue as a component of insulin resistance, the involvement of adipokines in muscle physiology is understudied. My research training plan will leverage endocrinology, muscle biology, bioinformatics, and newly generated genetic mouse models as a toolkit to investigate the function and mechanisms of a previously understudied hormone in promoting muscle growth with potentially fewer side effects. This proposal tests the central hypothesis that Isthmin-1 (Ism1) mediates adipose-muscle crosstalk that regulates muscle proteostasis. The following aims are proposed: Aim 1: To determine whether Ism1 mediates muscle growth through adipose-muscle crosstalk. Aim 2: To determine whether Ism1 alters balance between muscle protein synthesis and degradation. Aim 3: Using phosphoproteomics to discover novel pathways that regulate muscle proteostasis. If successful, this research will establish a novel adipose-muscle pathway in muscle growth with potential future therapeutic implications, since the Ism1 hormone itself can be used as a direct pharmacologic treatment for atrophy and disorders for which exercise is not an option. This project will also provide training for Dr. Zhao’s long-term goal as an independent investigator to dissect the hormonal crosstalk between metabolic organs to improve metabolic diseases. The candidate Dr. Zhao has extensive and suitable prior training in metabolic physiology, with 13 publications including 5 as first- author since 2016. The Career Development Plan is tailored to enable Dr. Zhao to gain new experimental skills and concepts in muscle biology, biochemistry, and endocrine signaling from expert physiologists, endocrinologists, and biochemists in the mentoring team. The environment at Stanford University is unparalleled for collaborative and innovative research and career development training. In summary, the strong mentoring environment and training plan are anticipated to fully prepare Dr. Zhao to launch her independent career. The proposed studies promise to offer mechanistic insights into adipose-muscle crosstalk to identify therapeutic targets that can counter muscle protein loss and associated metabolic disease.

NIH Research Projects · FY 2026 · 2023-04

PROJECT SUMMARY/ABSTRACT The objective of the proposed research is to engineer a targeted biological nanoparticle platform with high intracranial delivery and glial cell targeting for broad applicability in drug delivery and imaging. A great deal of work has already been accomplished elucidating the ability of certain extracellular vesicles (EVs) to cross endothelial barriers, especially the blood-brain barrier (BBB). Other work has established that EVs exhibit excellent tropism towards particular tissues and cell types. The focus of this proposal is to understand the mechanisms by which certain EV subpopulations accomplish these feats, and to engineer them into a hybrid liposome-EV drug delivery platform. Given the plethora of recent research into EV structure and function, it is well known that they exhibit considerable compositional heterogeneity. But fundamental questions still exist as to how EV prescribed functions differ across these subpopulations. It is likely that off-target effects and inefficiencies in capturing native EV functions with engineered mimetics are due to their substantial heterogeneity. Our first hypothesis is that homogenization of EVs towards a narrow size range with uniform biomolecular content will result in a more potent and controllable drug delivery platform that maintains native EV function yet reduces off-target toxicity. Our second hypothesis is that fusion of homogenized EVs and liposomes with various functions (i.e., efficient BBB permeation through receptor mediated transcytosis) will deliver an engineered product combining desired functions. We plan on addressing these hypotheses through rigorous engineering to homogenize EVs (Aim 1) alongside biochemical assays to detangle the mechanisms important for EV intracranial delivery. We will utilize EVs isolated from gliatropic “experts”, namely a vast library of glioblastoma (GBM) patient derived primary cell lines, brain-metastasizing breast cancer cells, and other glial and neuronal cells like astrocytes and neurons. Key molecular players important for intracranial delivery identified from those studies will feedback into synthesis of engineered EVs (eEVs) via subsequent fusion with carrier EVs (Aim 2). For the engineered eEV product, we will also incorporate synthetic liposomes decorated with known ligands to trigger receptor mediated transcytosis through the BBB endothelial layer. To provide the greatest opportunity to measure efficiency of functional intracranial delivery, we plan to load formulated, labeled, and homogenized eEVs with a chemotherapeutic payload and determine drug-release profile, biodistribution, and efficacy in healthy mice with intact BBBs and then an orthotopic GBM model (Aim 3). The proposed work is important because it seeks to eliminate the highly confounding factor of particle-to-particle variability plaguing effective application of EVs as potent drug-delivery vehicles. Success in homogenizing eEVs will result in an increased understanding of their biological function and assist in their application to combat a wide variety of neurological disorders where current drug delivery approaches are thwarted by low intracranial delivery.

NIH Research Projects · FY 2026 · 2023-04

Project Summary The largest class of genetic alterations that cause disease are single point mutations. Most of these disease-causing errors can be remedied if a specific adenosine is changed to guanosine on the RNA transcript. This proposal aims to repurpose an RNA editing enzyme and direct it to selectively edit targeted adenosines in mRNA to treat genetic disorders. The RNA editing enzyme Adenosine Deaminase acting on RNA (ADAR) can convert adenosine to inosine (A-to-I) by catalyzing a deamination reaction on the nucleobase. Inosine is read as guanosine by the cellular translation machinery providing the ability to alter codons in mRNA. This proposal will focus on selectively editing disease-causing nonsense mutations, by directing ADAR to edit the adenosine in the stop codon thus allowing the transcript to continue translation, producing functional full- length protein. Because ADARs selectively edit adenosines in regions of dsRNA, disease-causing nonsense mutations can be selectively targeted by furnishing an appropriate guide oligonucleotide to create a dsRNA substrate. Canonical Watson-Crick complementarity of duplex RNA does not produce efficient substrates for ADARs, making it challenging to design effective guide oligonucleotides to target specific nonsense mutations. A high-throughput assay is proposed to search all sequence space of guide RNA oligonucleotides to identify lead sequences displaying high editing efficiency of the targeted nonsense transcript using endogenous ADARs. These lead sequences can be further optimized by structure-guided rational design methods. The lab has significant experience in determining ADAR-RNA structures to atomic resolution and leveraging this knowledge to improve editing efficiency. Both X-ray crystallography and Cryo-EM techniques are proposed for ADAR1 or ADAR2 complexed with dsRNA of the lead sequence bound to its targeted mRNA segment. These structures will provide the basis for rational design adjustments to develop nucleotide analogs to incorporate into guide oligonucleotides that can fashion structural features for improved editing and increased metabolic stability. This method of site-directed RNA editing (SDRE) to treat genetic disorders offers many advantages over current editing tools, which often require addition of sizable proteins (e.g., CRISPR/Cas). When fully developed, this method would permit the simple administration shorter oligonucleotides allowing the cell’s endogenous ADARs to recode the nonsense mutation to treat many genetic disorders.

NIH Research Projects · FY 2026 · 2023-04

ABSTRACT/SUMMARY NCI clinical trials networks are essential to strengthen the cancer care of the nation by expediting the discovery of novel cancer treatments and therapeutic strategies. Central to this effort are the clinician scientists who development, implement, manage, and provide oversight to these individual trials and trial networks. While essential, these clinicians are frequently tasked with significant clinical, administrative, and teaching responsibilities that may inhibit substantiative contribution to NCI-sponsored clinical trial activity. The R50 Research Specialist /Clinician Scientist Award is designed to protect time and allow the clinician scientist to significantly contribute to NCI-sponsored clinical trials and NCI clinical trials networks through leadership in the development of national clinical trials, implementation of NCI clinical trials in their institutions, and national service to the NCI clinical trials networks. Dr. Brian Jonas, MD, PhD, is a leukemia specialist and an Associate Professor within the Department of Hematology/Oncology at UC Davis Comprehensive Cancer Center (UCDCCC). As an active and highly dedicated clinical scientist and trialist at UC Davis, he has established himself as a leader in opening clinical trials for the treatment of AML, MDS, acute lymphoblastic leukemia (ALL) and other hematologic malignancies. Moreover, he has received the coveted “Top Accrurer” Award within the cancer clinical trials unit. His clinical research is complemented by his translational studies involving AML biomarkers, chemosensitivity and therapeutic targeting of leukemia stem cells. As an asset to UCDCCC, he has served as PI on several clinical trials and as a leader in clinical trials management. This includes chairing the UCDCCC Hematologic Malignancies Disease Team Committee and the UCDCCC Data and Safety Monitoring Committee. In addition to his contributions at UC Davis, Dr. Jonas has contributed to multidisciplinary clinical and translational teams such as the Leukemia Committee and Working Group for the Southwest Oncology Group (SWOG), and the Calif ornia Cancer Consortium to shape and inf orm clinical trial portf olios at the UCDCCC, the National Clinical Trials Network (NCTN) and the Experimental Therapeutics Clinical Trials Network (ET-CTN). This includes his service as national chair of two ECTCN trials developed through the CTEP Project Team mechanism and the SWOG champion for an ECOG-ACRIN AML trial in development through the NCTN. He is also an active participant in the UM1 Consortium, the California Cancer Consortium (CCC), and a member of the CCC Heme Working Group. Overall, Dr. Jonas is a well-qualified clinician scientist candidate for the NCI R50 Research Specialist Award and has demonstrated prior and continuing leadership in the development of national clinical trials, implementation of NCI-sponsored clinical trials at UCD, and service to NCI clinical trial networks through participation in our CCC. The R50 award will facilitate continued growth of his research program and advancement of his institutional and national leadership.

NIH Research Projects · FY 2026 · 2023-04

Project Summary Alpha-synuclein (α-syn) is a neuronal protein encoded by the SNCA gene. Genetically, mutations in the SNCA gene lead to enhanced expression and aggregation of α-synuclein and cause inherited forms of Parkinson’s disease (PD). In idiopathic PD, as well as Alzheimer’s disease and related dementias (ADRD), α-syn aggregation leads to the formation of toxic α-syn fibrils that constitute the building blocks of Lewy bodies, the deviant protein deposits that accumulate and are associated with neuronal cell death. Thus, α-syn is considered a key pathological hallmark of PD. Due to our ever-extending life expectancy, the prevalence of PD is estimated to double by 2030. Age is the strongest risk factor for its development, and currently there is no cure and no therapeutic known to modify disease progression. Despite clear neuropathological consequences for α-syn accumulation in PD and ADRD there is a lack of mechanistic intracellular information regarding the molecular pathways perturbed by α-syn that lead to cell death. The goal of this application is to explore this critical gap in knowledge by examining whether α-syn alters the molecular composition of membrane contact sites. Our central hypothesis is that α-syn aberrantly remodels plasma membrane ion channels and lipids to alter endoplasmic reticulum – mitochondrial Ca2+ nanodomains leading to neurotoxicity. Our data supports the concept that PD is a nanostructural disease. To test this hypothesis, we implement an innovative multi-scale (including lipidomics, super-res imaging, genetics, and patch-clamp electrophysiology) approach to vertically integrate signaling cascades from the level of single lipids to neuronal networks, with the goal of providing fundamental knowledge that will aid in the development of novel strategies that slow or reduce neurotoxic α-syn-mediated cell death. Specific Aim 1 tests the hypothesis that α-syn remodels voltage-gated potassium and Ca2+ nanocomplexes to alter the biophysical and spatial properties of voltage-gated Ca2+ channels, leading to enhanced Ca2+ influx into neurons. Specific Aim 2 tests the hypothesis that, α-syn remodels phosphoinositide metabolizing enzymes to increase Ca2+ channel activity. Specific Aim 3 tests the hypothesis that α-syn aberrantly modifies ER and mitochondrial C a2+ signaling nanodomains leading to cytotoxicity. The proposed studies have specific relevance to the fields of neuroscience, cell biology and biophysics, but the fundamental importance of voltage-gated K+ and Ca2+ channels, as well as phosphoinositides mean it will have broad implications for medicine. Findings from this investigation will unveil crucial physiological roles for α-syn in organizing the nanoscale distribution of ion channels in health, as well as revealing novel signaling hubs that can be targeted for the development of therapeutic strategies for PD, ADRD, and synucleinopathies.

NIH Research Projects · FY 2026 · 2023-04

PROJECT SUMMARY/ABSTRACT State of the art methods for the early detection and monitoring of cancer are either invasive, time-consuming, expensive, or frequently inaccurate, which hinders the routine screening of at risk-patients to improve survival rates. The multiplexed detection of oncometabolites circulating in minimally or non-invasive biofluids, such as saliva, blood plasma, or sweat, could provide significant clinical and economic benefits. Metabolites and related circulating biomarkers are structurally unique elements with distinctive absorptive fingerprints in the infrared (IR) portion of the electromagnetic spectrum. Common approaches that provide multiplexed metabolite detection, such as mass spectrometry (MS), Raman spectroscopy, and Fourier transform infrared (FTIR) spectroscopy, are expensive and difficult to miniaturize. On the other hand, inexpensive miniaturized electrochemical techniques lack specificity, sensitivity, ease, and suffer from limited multiplexing. Portable technologies capable of rapid and accurate diagnostics of early/late-stage cancer are not readily available. To address this challenge, our multidisciplinary team proposes an innovative Neural Network Enabled Cancer Spectroscopy (NNECS) liquid biopsy platform based on plasmonic nano-micro electromechanical systems (NMEMS) to diagnose and monitor early/late-stage head neck cancer (HNC). Instead of targeting individual metabolites, we propose to process the entire IR spectrum of saliva, blood plasma, and sweat as a biomarker. Our focus is head and neck cancer (HNC), a highly metabolic disease where stratification of patients according to better diagnostic information would greatly improve outcomes. Our platform combines IR NMEMS sensors to accurately detect IR spectral fingerprints with neural network (NN) frameworks to find the appropriate combinations of spectral bands that will inform the design of highly multiplexed miniaturized biosensor. We will take a novel, interdisciplinary approach within the framework of five key components: (i) collecting and analyzing (FTIR, MS, histopathology/imaging) biofluids (saliva, sweat, blood) from a large number of early/late stage HNC patients and healthy subjects per year; (ii) developing powerful NN architectures and diagnosis tools for segregating early/late-stage HNC samples from controls, considering IR data streams from each individual biofluid as well as their potential combinations; (iii) developing a NNECS platform using arrays of plasmonic NMEMS targeting specific IR bands resolved by ML algorithms; (iv) determining NNECS early/late-stage cancer detection performance in terms of specificity, sensitivity, and accuracy; and (v) elucidating which metabolites drive the changes in the IR absorption of cancer biofluids supported by MS. The expected outcome is a miniaturized, label-free, affordable, and accurate technology able to radically improve the ability to diagnose early- stage HNC as well as the monitoring of recurrent HNC patients. Moving beyond, NNECS can be adapted for the diagnosis and monitoring of a wide range of metabolic conditions, including many types of cancer, diabetes, and heart-diseases.

NIH Research Projects · FY 2026 · 2023-04

Summary Time-of-flight positron emission tomography is a very effective nuclear imaging modality for the diagnosis and staging of a range of pathologies such as cancer, cardiovascular diseases, or musculoskeletal disorders. Commercial TOF-PET scanners currently employ lutetium-(yttrium)-oxyorthosilicate (L(Y)SO) crystal detectors coupled to silicon photomultipliers (SiPMs) to achieve coincidence time resolutions (CTR) between 200-500 ps full width at half maximum (FWHM). High production costs of L(Y)SO crystals and their intrinsic radiation background are currently hindering the evolution and spread of very promising TOF-PET modalities such as long axial field-of-view (LA-FOV) scanners or studies involving very low doses such as cell tracking or imaging with theranostic agents. New scintillator materials with lower production cost, radiation background-free, and with TOF-level timing accuracy are needed. We propose to use thallium chloride (TlCl) as a scintillator material for TOF-PET. TlCl is a material with a simple cubic structure that allows for a relatively easy and flexible doping process. Preliminary data obtained with TlCl crystals doped with beryllium (Be) and indium (I) show a very fast scintillation component of ~10 ns that has a high potential for very accurate timing measurements. TlCl has a greater detection efficiency than LYSO or even bismuth germanate (BGO) for 511 keV gammas, is background radiation-free, and its estimated production cost is 1/3 of L(Y)SO based on its low melting point of 430C (compared to 2050C for L(Y)SO) and simple lattice structure. Moreover, unlike BGO, TlCl uniquely combines a very fast scintillation process with a high Cherenkov generation yield to further boost timing potential. We aim to prove the feasibility of using TlCl detectors for TOF-PET by combining expertise in crystal growth, simulation of light generation and detection, and benchtop characterization. First, will study the effects of Be and I as dopants in TlCl with the aim of further improve the scintillation properties observed in the preliminary data. We will also optimize the surface treatment of TlCl to maximize the light extraction toward the photodetector. Second, we will develop a simulation framework that allows us to guide the crystal development process and to understand the fundamental timing limits of TlCl. Third, we will characterize individual TlCl crystals with different choices of photodetectors to evaluate their timing and energy resolution accuracy. Results obtained with these crystals will be used to tune and validate the simulation model as well. Finally, we will evaluate the performance of TlCl detector blocks of 4x4 crystal elements. We will evaluate their timing resolution, depth-of-interaction estimation accuracy, and quality of flood histograms.

NIH Research Projects · FY 2026 · 2023-03

PROJECT SUMMARY/ABSTRACT I propose to develop and test a machine learning (ML) algorithm that uses dynamic data from pulse oximetry for critical congenital heart disease (CCHD) screening. Oxygen saturation (SpO2)-based screening is the current standard for CCHD screening; however, it fails to detect 50% of asymptomatic newborns with CCHD or nearly 900 newborns in the United States annually. Most newborns missed by SpO2 screening have defects with systemic obstruction, such as coarctation of the aorta (CoA), that do not cause hypoxemia. Pulse oximetry can also measure non-invasive measurements such as perfusion such as perfusion index (PIx), radiofemoral delay, heart rate, and other waveform characteristics. Introduction of other pulse oximetry features is expected to improve CCHD and CoA detection. My recent work revealed improved CCHD detection using ML algorithms that combined pulse oximetry features. The algorithms improved CCHD detection to at least 93%, including improved detection of CoA, while maintaining high specificity. However, the model depended on two separate measurements including simultaneously artifact free waveforms in both the right hand and a foot. Having a model with dynamic prognostication that allows for an infant’s predicted outcome to change as new data is incorporated could be better. Additionally, the amount of time to obtain two waveforms that are artifact free in a possibly moving baby needs to be understood for implementation. Therefore, I will develop and test a ML algorithm that combines pulse oximetry features and incorporates dynamic data from repeated measurements allowing a newborn’s predicted classification (CCHD vs no-CCHD) to change as new data is incorporated. I will do this in two ways. The first will utilize only inpatient measurements and will externally validate our recently developed ML algorithm. This first approach will also test a “repeat” screen for any initial “fails,” an approach that mimics the current SpO2 standard screen and is expected to keep the false positive rate below 1%. The second approach will incorporate measurements after 48 hours of age (including from the outpatient setting). Outpatient CCHD screening has not been studied. Most newborns are seen for routine follow up outpatient around the age at which CoA becomes more clinically apparent, and thus, more likely to be detected by non-invasive perfusion assessments. This study is significant because a dynamic screening model that includes perfusion data could save the lives of hundreds of newborns with CCHD that are not diagnosed by SpO2 screening annually. Additionally, it is innovative because it makes use of readily available non-invasive pulse oximetry data and will use dynamic data (inpatient and outpatient) that allows for a newborn’s prognostication to change as new data is incorporated. From this study and career plan, I will gain skills in machine learning with emphasis in dynamic approaches, and implementation science. I will use the results and skills from this proposal to then study a cluster randomized trial of our algorithm and assess implementation processes.

NIH Research Projects · FY 2025 · 2023-03

ABSTRACT Cardiovascular disease (CVD), the leading cause of death in the U.S., disproportionately impacts persons of lower socioeconomic position (SEP) -- a disparity that has been attributed to heightened exposures to both traffic- related air pollution (TRAP) and chronic social stressors. Many epidemiologic and toxicologic studies have shown that exposures to chronic stress can vastly increase susceptibility to TRAP, though growing evidence now suggests that TRAP may also strongly impact hypothalamic-pituitary-adrenal (HPA)-axis function and acute stress response, complicating the directionality and interpretation of interactions. It is critical to disentangle these two models, to develop more biologically-grounded epidemiologic model structures, and refine the design of space-time exposure metrics for both stress and pollution. Ultimately, this work will help to better identify susceptible populations, and identify effective interventions to improve health and reduce health disparities. Doing so is challenging, however, as both stress and TRAP are complex exposures with diverse multi-systemic impacts. Stress is shown to strongly impact immune, endocrine, and metabolic function, but effects are highly time-sensitive, as acute and chronic stress manifest very differently. TRAP is a highly complex mix of chemicals, each with very different physiologic impacts. In this ViCTER proposal, we establish an interdisciplinary team to quantify and compare chronic and acute stress, TRAP, and their multiple interactions, in shaping cardiovascular function. We are uniquely poised to map this unexplored terrain, as accomplished senior investigators in atmospheric science and mechanical engineering (Wexler), cardiovascular regulation and autonomic function (Chen), and exposure science and social-environmental epidemiology (Clougherty). To do so, we will use a unique TRAP delivery system with real-time concentration and chemical composition measures, a well-validated model for generating chronic and acute stress responses in rats, time-resolved measures of cardiovascular function (telemetry), and biological profiling at multiple time points for chronic and acute stress markers (e.g., cortisol, CRP, cytokines), to quantify and compare directionality in the two conceptual models described above. We hypothesize that: (1) TRAP composition (light- vs. heavy-duty vehicles) differently impact cardiovascular function; (2) Chronic stress may heighten animals’ cardiovascular response to TRAP, over the course of study; (3) TRAP may compromise animals’ cardiovascular resilience to stress challenge. This study will establish an interdisciplinary team with complementary expertise to examine complexities in the interactions among stress and pollution exposures – an issue profoundly relevant to health disparities in under-served and marginalized communities, especially for CVD, the leading cause of death. The team will work together to integrate the results of the three aims into hypotheses for subsequent R01 applications.