University Of Washington

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY. Adolescence is a period of heightened vulnerability to psychopathology and sensitivity to peer social contexts. Due to the increasing use of online platforms, social media has been identified as a risk factor for the significant rise in adolescent mental health issues. However, our current understanding is limited by methods that do not fully capture that the link between social media and psychopathology is bidirectional, influenced by the positive or negative nature of online content, and operates over both short and long-term timeframes. This association changes over development and is highly variable across individuals and time. Understanding the underlying factors that contribute to variations in outcomes in group-level studies of the impact of social media on youth development is crucial for prevention and intervention at the individual level. Our project proposes conceptual and methodological improvements to capture interactions between social media and emerging psychopathology at different person and time scales. First, conducting person-specific research enables the identification of heterogeneity in the unique individual, bidirectional dynamics often obscured by the pooling necessary to estimate average, group-level effects. Second, ecological momentary assessment, which captures real-time dynamics of constant social media exposure, is ideal for testing the cumulative effects of small influences. Third, given the transformation of peer interactions in adolescence, and their contributing role to psychopathology, positive and negative interpersonal experiences on and off social media emerge as a promising pathway to investigate. Fourth, longitudinal models are necessary for capturing relative contributions of online versus in-person social interactions to developmental risk during adolescence. We will use a developmental cohort-accelerated longitudinal design to recruit a sample of youth in early adolescence (n=150, 13-14 years) and middle adolescence (n=150, 15-16 years). Adolescents will be tracked across four years, completing annual 30-day bursts of ecological momentary assessment of in-person and social media interactions, socioemotional vulnerability, and psychopathology. Data accumulating across cohorts will provide longitudinal coverage for each person over the span of four years, permitting analytic leverage from early to late adolescence (i.e., observations from age 13 to age 19). We aim to test (1) real-time differential susceptibility to positive/negative experiences across individuals and time, identifying developmental windows of susceptibility to social media effects, (2) pathways for risk accumulation, and (3) online vs in-person social contributions to developmental risk. We plan to directly test heterogeneity in the links between social media use and psychopathology, which will provide critical insights beyond estimating the average risk of social media use for adolescents. The long-term goal is to identify who is at the highest risk of psychopathology, when during adolescence, due to what specific social media experiences, and how these experiences bidirectionally accelerate or mitigate socioemotional risk and psychopathology.

NIH Research Projects · FY 2024 · 2023-09

PROJECT SUMMARY. Adolescence is a period of heightened vulnerability to psychopathology and sensitivity to peer social contexts. Due to the increasing use of online platforms, social media has been identified as a risk factor for the significant rise in adolescent mental health issues. However, our current understanding is limited by methods that do not fully capture that the link between social media and psychopathology is bidirectional, influenced by the positive or negative nature of online content, and operates over both short and long-term timeframes. This association changes over development and is highly variable across individuals and time. Understanding the underlying factors that contribute to variations in outcomes in group-level studies of the impact of social media on youth development is crucial for prevention and intervention at the individual level. Our project proposes conceptual and methodological improvements to capture interactions between social media and emerging psychopathology at different person and time scales. First, conducting person-specific research enables the identification of heterogeneity in the unique individual, bidirectional dynamics often obscured by the pooling necessary to estimate average, group-level effects. Second, ecological momentary assessment, which captures real-time dynamics of constant social media exposure, is ideal for testing the cumulative effects of small influences. Third, given the transformation of peer interactions in adolescence, and their contributing role to psychopathology, positive and negative interpersonal experiences on and off social media emerge as a promising pathway to investigate. Fourth, longitudinal models are necessary for capturing relative contributions of online versus in-person social interactions to developmental risk during adolescence. We will use a developmental cohort-accelerated longitudinal design to recruit a sample of youth in early adolescence (n=150, 13-14 years) and middle adolescence (n=150, 15-16 years). Adolescents will be tracked across four years, completing annual 30-day bursts of ecological momentary assessment of in-person and social media interactions, socioemotional vulnerability, and psychopathology. Data accumulating across cohorts will provide longitudinal coverage for each person over the span of four years, permitting analytic leverage from early to late adolescence (i.e., observations from age 13 to age 19). We aim to test (1) real-time differential susceptibility to positive/negative experiences across individuals and time, identifying developmental windows of susceptibility to social media effects, (2) pathways for risk accumulation, and (3) online vs in-person social contributions to developmental risk. We plan to directly test heterogeneity in the links between social media use and psychopathology, which will provide critical insights beyond estimating the average risk of social media use for adolescents. The long-term goal is to identify who is at the highest risk of psychopathology, when during adolescence, due to what specific social media experiences, and how these experiences bidirectionally accelerate or mitigate socioemotional risk and psychopathology.

NIH Research Projects · FY 2026 · 2023-09

Project Summary / Abstract Healthy diets reduce the risk of cardiovascular disease (CVD). Plant-based diets, which are comprised predominantly of plant foods and are low in animal products, are healthy dietary patterns that are associated with a lower risk of CVD. However, the precise mechanisms underlying plant-based diet-CVD associations are poorly understood. The present proposal aims to elucidate the mechanisms through which plant-based diets are associated with CVD using proteomics and epigenetics. Specific aims of the present proposal are to: 1) identify protein signatures of 4 plant-based diet indices [overall plant-based diet index (PDI), provegetarian diet, healthy plant-based diet index (hPDI), and unhealthy plant-based diet index (uPDI)], 2) evaluate the prospective associations between plant-based diet-related proteins and incident CVD, and 3) explore DNA methylation signatures of 4 plant-based diet indices. The proposed study will use the Atherosclerosis Risk in Communities (ARIC) study, a richly phenotyped community-based prospective study of middle-aged African American and European American with usual dietary intake, proteomics and DNA methylation data, and adjudicated CVD outcomes as the discovery data set. Framingham Heart Study and Multi-Ethnic Study of Atherosclerosis, well-characterized prospective cohorts with similar data, will be used for external validation. Completion of the aims will identify novel proteins, epigenetic markers, and pathways that are modifiable by plant-based diets. Such novel evidence will be the first step for developing biomarkers for early detection and identifying therapeutic targets of CVD. Hyunju Kim, PhD seeks a K01 Mentored Research Scientist Development Award to acquire essential skills, knowledge, and mentored research experience to prepare for a future career as an independent investigator with expertise in nutrition, multi-omics, and CVD. Dr. Kim has a track record of research productivity in the areas of nutritional epidemiology and nutritional metabolomics. However, Dr. Kim needs additional training in proteomics, epigenetics, and cardiovascular biology to achieve research independence. This research proposal details a five-year plan consisting of didactic coursework, hands-on research training, career enrichment programs under the primary mentorship of Dr. Casey Rebholz, PhD (expert in nutritional proteomics), and co-mentorship by Dr. Josef Coresh, MD, PhD (expert in proteomics and vascular disease), and Dr. Dan Arking, PhD (expert in epigenetics). The exceptional mentoring team in addition to external collaborators and advisors with multi-disciplinary expertise, and a highly collaborative environment at Johns Hopkins Bloomberg School of Public Health will ensure the successful transition of Dr. Kim into an independent investigator.

NIH Research Projects · FY 2025 · 2023-09

Project Abstract Co-use of alcohol and cannabis (using both substances, either simultaneously or concurrently) represents a significant public health concern among young adults (YAs), as co-use of these substances is common and worsens alcohol-related harms. However, YAs are unlikely to seek treatment for their substance use, and existing brief interventions (BIs) for alcohol and cannabis misuse produce modest effects. Thus, there is a critical need to explore novel treatment components to increase participant engagement and maximize effects. From a social-developmental perspective, romantic partners have the strongest social influence on YA substance use, and an underexplored area for intervention. Dyadic research suggests romantic couples mutually influence each other’s substance use, increasing risk for greater substance use involvement and more negative interpersonal consequences. Whether these associations extend to or are exacerbated by co- use remains understudied. Crucially, there is no known BI for co-use among YA couples. This innovative study seeks to fill these gaps in two phases. Phase 1a: 75 YA couples will provide 30 days of daily reports on alcohol and cannabis use, co-use, negative (e.g., conflict) and positive interpersonal consequences (e.g., intimacy), and relationship outcomes (e.g., satisfaction). Multilevel models will test whether a person’s co-use is associated with a greater likelihood of their partner co-using. Multilevel Actor Partner Independence Models will test if couples experience greater negative consequences, fewer positive consequences, and poorer relationship functioning on days when both partners report co-use than on days when only one partner reports co-use, and vice versa (consequences → next day co-use). Phase 1b: 15 participants from Phase 1 will participate in qualitative interviews to assess treatment need and inform intervention content. Data and rapid prototyping (Phase 1 c) will inform development of an online, couples- based BI aimed at decreasing co-use and promoting healthy relationship skills. Phase 2: A second cohort of co-using YA couples will be randomly assigned to the intervention (n = 15 couples) or control (n = 15 couples) and complete baseline, post-assessment, and 3-month follow-up surveys testing feasibility, acceptability, and initial support. Completing this research will support my development as an independent alcohol and cannabis researcher with a focus on developing novel interventions for high-risk YA populations. To achieve my career objectives, I require additional training in 1) social-interpersonal influences that impact etiology, maintenance, and treatment of alcohol and cannabis use among YAs in romantic relationships, 2) couples-based intervention delivery, 3) dyadic ecological momentary assessment research and dyadic data analysis, 4) user-centered design, and 5) randomized control trial (RCT) design. The project will provide pilot data for an R01 submission to NIAAA which will test efficacy of the intervention in a full-size RCT.

NIH Research Projects · FY 2025 · 2023-09

SUMMARY Breastfeeding has multiple beneficial effects on maternal and neonatal health; however, the statistics indicate that up to 96% of lactating women in the US take one or more medications while breastfeeding. Medications consumed by lactating women may be transferred into breast milk to a significant extent, resulting in unintentional infant exposure of medications and in some cases adverse health outcomes for the infants. Quantifying drug transfer into human breast milk is important for rational risk assessment balancing the toxicity risk of drug exposure to infants and the benefits of breastfeeding. However, clinical pharmacokinetic (PK) studies in the population of lactating women are challenging and logistically not possible for every drug taken by lactating women, necessitating the use of prediction methods to address this issue. One historical approach is the prediction of drug concentrations (or drug AUC) in breast milk based on maternal plasma concentration (or AUC) and the milk-to-plasma (M/P) concentration or AUC ratio. The M/P ratio itself can be predicted using both physicochemical characteristics of drugs and physiological parameters of breast milk. While this approach may predict the M/P ratios of drugs that enter the milk predominantly by passive diffusion, no methods are currently available to accurately predict milk secretion of drugs via transport mechanisms. Nonetheless, milk secretions of many drugs, xenobiotics and endogenous substances are known to be mediated by transporters expressed in mammary epithelial cells (MECs). In this application, we propose a systems pharmacology approach to predict transporter-mediated milk secretion of drugs. Our hypothesis is that the transporter-mediated drug PK in human breast milk can be predicted using in vitro experimental data combined with Physiologically Based Pharmacokinetic (PBPK) modeling and simulation (M&S). Specifically, we propose an innovative approach which utilizes human MECs and transporter-transfected cells or plasma membrane vesicles expressing individual transporters of interest (i.e. OCT1, BCRP). Using quantitative targeted proteomics, the human MECs will allow us to determine the protein abundance of these transporters in the mammary gland. The transporter-transfected cell or plasma membrane vesicle studies will allow us to determine the in vitro intrinsic transport clearance of a drug by a single transporter. Then, the in vitro intrinsic transporter-mediated clearances will be extrapolated to in vivo in the mammary gland for PBPK M&S. PBPK model predictions will be verified using the drug PK data in human breast milk obtained from a clinical study conducted with a transporter substrate. Combined, these data will allow us to predict transporter-mediated drug PK in the milk of lactating women. These studies will address a critical gap in our understanding of drug PK in human breast milk during lactation. Since our approach can be applied to other drugs that are substrates of any transporters of interest, its significance goes well beyond the drug and transporters investigated here.

NIH Research Projects · FY 2026 · 2023-09

Co-occurring neuropathologies such as Lewy bodies, hippocampal sclerosis, and microinfarcts likely influence heterogeneity in genetic studies of Alzheimer’s disease (AD). The genetic architecture of these co-occurring neuropathologies is not clear. Most AD genetic analyses use clinical AD dementia phenotypes and are limited by phenotype misclassification: many clinical AD “controls” harbor AD pathology while clinical AD “cases” have no or minimal AD pathology. Only 20% of pathology confirmed AD dementia cases have only AD pathology (“pure AD”); 80% have co-occurring neuropathologies (“mixed AD”). A complementary strategy is to perform genome-wide association studies (GWAS) specifically distinguishing between pure and mixed AD. The overarching goal of this proposal is to further scientific understanding of pure AD and mixed AD by integrating neuropathology together with advanced statistical approaches and extensive multi-omics data. Aim 1: Distinguish the genetic architecture of pure AD vs. mixed AD vs. pathology-free controls and determine how associations are driven by particular neuropathological patterns. 1a. Perform GWAS of pathology-confirmed AD controls vs. pure AD vs. mixed AD using a multinomial regression framework. We will also examine how associations are driven by particular neuropathological features furthering our understanding of underlying mechanisms. 1b. Infer possible mechanisms by integrating biological knowledge. We will perform transcriptome- wide analyses and gene-network analyses of pure AD and mixed AD, integrating human PPI data. Aim 2: Identify factors specific to pure AD or mixed AD. 2a. Use machine learning and harmonized clinical data to isolate factors associated with pure or mixed AD. Identified clinical risk factors could be used for future clinical trial stratification. 2b. Perform genetic correlation analysis and Mendelian randomization to estimate correlated genetic effects and potential causal effects between candidate risk factor traits and pure or mixed AD. Genetic correlation analysis will gain novel insights into the shared genetic basis between biobank-scale GWAS traits and pure AD and mixed AD. We will follow-up significant correlations with local genetic correlation and Mendelian randomization (MR) to identify specific genomic regions contributing to the correlation and quantify causal effects of these traits on pure AD and mixed AD. Aim 3. Characterize the cellular and molecular consequences of pure AD vs mixed AD. We will harness the data generated by the SEA-AD and the ROS-MAP studies to characterize the molecular changes occurring for our phenotypes at cell type resolution. We will perform abundance analyses (3a), differential gene expression (3b), and differential chromatin accessibility (3c) in each cell type across multiple brain regions. We will investigate loci from recent GWASs of AD dementia and candidates from Aim 1.

NIH Research Projects · FY 2025 · 2023-09

Project Abstract Brain metastases (BM) are the most commonly diagnosed type of central nervous system tumor, more frequent than primary intracranial neoplasms. Progress on various therapies have accelerated over the past decade through ongoing clinical trials, and the historically poor outcomes for patients with BM have been markedly improved. Contrast-enhanced T1-weighted MRI is routinely applied to depict BM with the size of the enhanced lesions for assessing treatment response. Lesion enhancement due to the disruption of blood-brain barrier is rather nonspecific of the functions of brain tumors. The most studied MRI methodology for perfusion measurement is dynamic susceptibility contrast perfusion weighted imaging (DSC-PWI), which measures cerebral blood flow (CBF) and cerebral blood volume (CBV). CBV is the widely adopted perfusion measure as a sensitive marker of tumor vascularity. However, its clinical applicability in BM studies is hampered by its lack of absolute quantification, the contrast-leakage effect, and frequent susceptibility artifacts. Arterial spin labeling (ASL) is ideal for frequent non-invasive longitudinal monitoring of tumor vascularity. The standardized spatially selective ASL technique for CBF mapping is the pseudo-continuous ASL (PCASL) method using a single post- labeling delay, which may render underestimation of CBF due to transit time delay caused by slow arterial flow typical in elderly patients. Velocity-selective ASL (VSASL) was proposed to remove the time-delay sensitivity. Our group has implemented the first velocity-selective inversion (VSI) based VSASL with 3D segmented GRASE acquisition and demonstrated its higher sensitivity to perfusion signal over conventional ASL methods. Additionally, our group first developed VSASL based CBV mapping by removing labeling delay, which delivered much higher SNR than ASL based CBF mapping. Furthermore, our preliminary data showed that VSASL with 3D stack-of-spiral based FLASH acquisition delivered better perfusion image quality with less artifacts than using GRASE, and high temporal resolution potentially allowing adequate retrospective motion correction. The purpose of this study is: Aim 1, to conduct further technical developments for VSASL based CBF and CBV mapping protocols with accelerated acquisitions; Aim 2, to evaluate the sensitivity of the two optimized VSASL protocols to CBF and CBV changes within a month after the radiation therapy, and assess their early prediction to treatment outcomes; Aim 3, to compare the specificity of VSASL derived CBF and CBV values in the distinction of metastatic recurrence from radiation-induced effects; Aim 4, to ensure high reproducibility of the VSASL protocols between multiple scanners with different vendors and field strength. By completing the proposed aims, the advanced VSASL based CBF and CBV mapping methods are expected to demonstrate important values for monitoring treatment response in BM, will be readily available for large-scale clinical studies, and can benefit for studies of all primary and metastatic tumors in the body.

NIH Research Projects · FY 2025 · 2023-09

Project Summary/Abstract The overall goal of this project is to advance the care of pediatric patients (up to 21 years of age) during medical ground or air transport from one hospital to another within a regional network. The project will build on our previous experience with neonatal patient transport and use a five-stage innovation cycle, including: problem analysis, design, development, implementation, and evaluation to identify and address the salient issues and risks of regional neonatal transportation for which new and innovative approaches are needed. In collaboration with transport providers and other stakeholders, we will analyze current workflow processes, transport records (local and statewide databases), and facilities at referral and receiving facilities to develop a complete understanding of system issues and to define the current and ideal states. This detailed problem analysis phase will enable the integration of real-time data into a transport “digital twin” model to optimize regional consultation, triage, and transport of pediatric patients to facilities with the appropriate level of care and availability of space and staffing. With the input of clinical transport team staff and stakeholders, the project team will work with collaborators from the University of Washington Industrial and Systems Engineering Department to integrate machine learning into the Transport Monitoring and Communications (T-MAC) system to support the analysis of data feeds while on transport. The machine learning augmented T-MAC system will undergo repeated testing and revision to ensure that it can functionally and efficiently facilitate information flow between the medical control physician, referring facility and transport team. In addition, we will develop robust processes to support information flow to patients and families in the peri- transport period. To accomplish this, we will work with families to identify gaps in communication and connection to local and receiving facility resources and identify transport-specific needs. The efficacy of the T-MAC system will be evaluated in a realistic in situ simulation and clinical settings. We are confident that the lessons learned through this study will improve pediatric patient safety on medical transports and will be transferable to patient populations that undergo both short and long-range interfacility transports.

NIH Research Projects · FY 2026 · 2023-09

PROJECT SUMMARY/ABSTRACT A single mutation in a gene can lead to a disease. Many mutations, however, are benign. How do we determine which mutations are pathogenic and which are benign? There are hundreds of millions of single nucleotide variants in the human genome that have been identified by sequencing, with 5 million of these in protein-coding DNA. Despite this wealth of data, only ~2% of these variants have clinical annotations, half of which are “variants of uncertain significance”. High throughput functional screening of variants is thus needed for the millions of variants that have not been annotated. One approach for studying such large variant libraries is to use a method called deep mutational scanning, in which a library containing millions of gene variants is screened in parallel. This approach has proven useful in classifying variants of BRCA1, PTEN, and others by carrying out screens in cell culture. While this approach has the benefit of scale, it does not enable mutations to be studied in a developmental or tissue-specific context. However, human genetic diseases, including cancer, are tissue- specific. It is therefore desirable to study mutations in the context of whole organisms. We have recently shown in our lab that we can generate many Drosophila melanogaster with genomically integrated gene variants on the order of 1,000 or more variants at a cost of ~$1/variant for DNA delivery. Using this new ability to generate large numbers of transgenic D. melanogaster, we propose use a D. melanogaster seizure disease model to phenotype over 1,000 gene variants of para, a closely related ortholog to the human gene SCN1A, the most commonly implicated gene in epilepsy. Aim 1a: Generate over 1,000 D. melanogaster harboring different para variants. Aim 1b: Phenotype the variant flies in parallel using a seizure assay. Pool flies by their seizure severity, sequence them, and report findings to ClinVar to improve clinical variant annotation. Aim 2a: Increase the scale of variant libraries that can be made in D. melanogaster by characterizing more highly efficiency DNA recombinases for genomic integration of DNA into the genome. Aim 2b: Use a gene gun to increase the rate at which DNA is delivered to embryos, up to a thousand or more embryos per second. In this work, we will use our recently developed approach for generating large numbers of transgenic D. melanogaster to perform mutational scanning of over 1,000 variants of para. These results will provide supporting evidence for variant classification of the analogous variants of the human gene SCN1A, which is very closely related to para. In this work we will also increase the scale of DNA delivery and efficiency of integration into the fly genome so that larger numbers of gene variants can be screened in the future. We believe our approaches will prove very useful for assigning clinical annotations for the vast majority of variants of disease-related genes that show tissue-specific effects. We also expect the technology to be widely adopted by the developmental biology community, as it will enable rapid phenotyping of thousands of variants of developmentally important genes in a whole organism context.

NIH Research Projects · FY 2024 · 2023-09

Project Summary Over 20 million people living with HIV (PLHIV) are receiving antiretroviral therapy and require HIV viral load testing to identify cases of virological failure and provide actionable information to guide alternative clinical treatment. Current methods for HIV viral load measurement rely on quantitative PCR (qPCR) or digital PCR (dPCR), which are commonly restricted to highly resourced central laboratories and there is a need to decentralize HIV viral load monitoring to enable rapid, clinic-based or home self-testing viral load measurements. We have identified a novel method for implementing digital isothermal amplification that leverages the characteristic viscous reaction buffer of recombinase polymerase amplification (RPA) isothermal amplification chemistry and commercially available porous membranes. We propose to apply amplification nucleation site analysis (ANSA) for HIV-1 viral load monitoring to accurately quantify HIV-1 RNA over clinically relevant HIV-1 subtypes and viral loads. We propose two exploratory aims to demonstrate that ANSA can achieve the required viral load dynamic range and quantitative precision across HIV-1 subtypes.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY / ABSTRACT A common and deadly form of familial heart disease is dilated cardiomyopathy (DCM), which is typically characterized by adverse cellular and ventricular remodeling and systolic dysfunction. DCM is often associated with loss-of-function mutations in genes encoding sarcomeric or cytoskeletal proteins. Mechanotransmission and mechanosignaling in cardiomyocytes (CMs) rely on these protein networks, particularly in the costamere, which provides a direct mechanical link between the extracellular matrix (ECM) and the Z-disk of the sarcomere. The costamere may therefore regulate both ‘inside-out’ mechanotransmission (transmitting sarcomere-born forces out to the ECM) and ‘outside-in’ mechanosignaling (transmitting/transducing extracellular mechanical signals into the CM)—the dysfunction of either of which may be central to DCM progression. My overall hypothesis is that the costamere and cortical cytoskeleton of cardiomyocytes provide key mechanosensitive protein networks that regulate mechanical signalling pathways initiated by intracellular and extracellular forces, and that specific defects in these structures inhibits their ability to transmit and transduce mechanical forces, causing contractile dysfunction and pathological cell remodelling. Supporting this, the costamere protein Filamin C (FLNC) has recently been implicated in a variety of human cardiomyopathies, including DCM. During my F32 postdoctoral training, I used a new mouse model that exploits cardiac-specific and inducible homozygous FLNC deletion to trigger rapid DCM development. I found that a loss of FLNC causes significant reductions in the tissue- and cell- level contractility, as well as significant CM remodeling accompanied by a reduction in cortical cytoskeleton stiffness. However, whether FLNC mutations in humans with DCM cause similar defects in cortex structure and mechanics, systolic mechanotransmission, and mechanosensitive gene regulation requires further investigation. Thus, the goal of my proposed research is to integrate quantitative subcellular-level structural and biomechanical measurements with quantitative measurements of intracellular stress distributions and hypertrophic gene expression patterns in response to intra- and extra-cellular mechanical perturbations using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) expressing a patient- specific FLNC-truncating mutation. To accomplish this, I will: (1) combine X-ray diffraction imaging, atomic force microscopy, and multiscale computational modeling to test the hypothesis that a loss of FLNC disrupts ‘inside-out’ mechanotransmission of sarcomeric forces by dysregulating myofilament lattice geometry via altered cortical cytoskeleton mechanics in murine CMs, (2) apply these biophysical methods and hypotheses to a new human DCM model made from gene-edited hiPSC-CMs expressing a patient-specific FLNC-truncating variant, and (3) combine FRET-based molecular tension sensor imaging, in-vitro extracellular mechanical loading techniques, and quantitative transcriptomics to test the hypothesis that truncated FLNC dysregulates ‘outside- in’ mechanosignaling in hiPSC-CMs and promotes DCM remodeling.

NIH Research Projects · FY 2025 · 2023-09

The purpose of this Mentored Patient-Oriented Research Career Development Award is to prepare the candidate, Dr. Maya Elias, for an independent program of geriatric critical care research. More than 60% of intensive care unit (ICU) patients are adults ages 60 and older. Following hospital discharge, about 25% of ICU survivors experience cognitive impairment comparable in severity to mild Alzheimer’s disease and related dementias. Moreover, ICU survivors often experience sleep disturbances and inactivity, and almost 80% of ICU patients experience disturbances in circadian rhythm, which may affect cognitive function. The scientific premises of the proposed study are: 1) a combined sleep promotion and cognitive training intervention will have synergistic effects to mitigate the risk of cognitive impairment and development of Alzheimer’s disease and related dementias in older ICU survivors; and 2) chronotherapeutic timing of interventions (i.e., adjusting timing of interventions according to circadian rhythm profiles) may improve intervention efficacy. The proposed research and training build upon prior NIH-funded postdoctoral research conducted by Dr. Elias (F32NR018585). Training Aim 1 will develop expertise in designing and refining behavioral sleep and cognitive interventions through training in clinical trial methodologies and statistical analyses. Training Aim 2 will expand knowledge of circadian science and chronotherapy to examine circadian rhythm as a mechanistic factor underlying delirium and cognitive decline related to Alzheimer’s disease and related dementias in older ICU survivors. This Career Development Award will also support Dr. Elias to lead a clinical trial, which will evaluate feasibility of existing behavioral sleep and cognitive interventions to inform the design of a larger randomized controlled trial. Using a 2 x 2 factorial design, 100 English- or Spanish-speaking older ICU survivors will be enrolled after discharge out of the ICU and randomized to one of 4 combinations of two interventions: SLEEP [nighttime use of earplugs and eye masks] and COG [daily computerized cognitive training sessions]. Specific Aim 1 will test the separate and combined effects of SLEEP and COG, versus AC [active control], in improving cognitive function for older ICU survivors. Specific Aim 2 will examine circadian rhythm parameters of continuous body temperature to determine the optimal window for timing of the COG intervention. Specific Aim 3 will examine if the effects of each intervention [SLEEP + COG, SLEEP, COG] on cognitive function are mediated by sleep and activity, and will examine if selected biological and clinical factors moderate the intervention effects. Exploratory Aim 4 will explore the effect of each intervention on cognitive function at 1 month and incident Alzheimer’s disease and related dementias at 6 months and 12 months after hospital discharge. Results will yield crucial preliminary data for future clinical trials and will guide Dr. Elias to establish a program of research testing interventions that will improve outcomes for older ICU survivors throughout recovery from critical illness.

NIH Research Projects · FY 2025 · 2023-09

The research activities proposed in this application address a pressing need in American Indian (AI) communities to better understand multi-level (i.e., community- , household- , and individual- level) factors in the physical and socio-cultural environment that impact diet and cardiovascular diseases. The premise of the proposed project was informed by: (1) our previous work in the Strong Heart Study (SHS) and the Strong Heart Family Study (SHFS)—the largest multi-tribal cohort studies of risk factors for cardiovascular disease in AIs in the USA—that highlighted the exceedingly high burden of obesity and poor diet quality among AIs; and (2) findings from our Healthy Food, Healthy Families Feasibility Study—a small pilot study in a SHS/SHFS community that assessed the major barriers and facilitators to eating healthy. Although the SHS/SHFS has followed more than 4,000 AIs from 12 rural communities for the past 34 years, all data collected as part of the study has focused on individual-level biological and behavioral factors associated with cardiovascular diseases; no data has been collected on multi-level components of the physical or sociocultural food environments in this cohort. As such, we have incomplete knowledge about the influence of these domains on diet and cardiovascular health in these rural AI communities. To address these gaps, we will leverage existing data from the SHS/SHFS, and add comprehensive assessments of multi-level factors related to the physical and socio-cultural environment (factors selected based on previous pilot work) that may impact diet and cardiovascular health to the upcoming SHS/SHFS exam in 12 rural AI communities. For this project, we will assess associations of community-level (e.g., food outlet availability, food cost, norms around healthy eating), household-level (e.g., food insecurity, food outlet accessibility; food preferences), and individual-level (e.g., perceived nutrition environment) aspects of the physical and socio- cultural environment with diet (reported intake of fruits and vegetables; sugar-sweetened beverages) and cardiovascular risk factors (BMI; fasting glucose) among SHS/SHFS participants from 12 tribes. We will also assess the influence of participation in nutrition assistance programs (i.e., supplemental nutrition assistance program (SNAP) or the Food Distribution Program on Indian Reservations) on diet and cardiovascular risk factors (i.e., reported intake of fruits and vegetables; sugar-sweetened beverages; BMI; fasting glucose). Exploratory outcomes include: HDL cholesterol, triglycerides, systolic blood pressure, incident diabetes and incident cardiovascular diseases. Greater efforts are needed to understand the most salient factors that impact diet and cardiovascular health in rural Tribal communities. This ancillary study will leverage existing resources from the SHS/SHFS, while adding a comprehensive dietary assessment, measures of food security, and assessments of nutrition economic and neighborhood environments to the study.

NIH Research Projects · FY 2024 · 2023-09

PROJECT SUMMARY Although air-purifying respirators (APRs) protect against the spread of harmful particulate matter from nat- ural disasters like wildfires or pandemics like COVID-19, industry-standard APRs can cause contact dermatitis, discomfort, and bruising after prolonged wear; obstruction of visual communication, significant environmental waste, and, in many types of commonly used masks, provide insufficient or unknown filtration. When the appro- priate fit of an N95 filtering facepiece respirator (N95 FFR) is pursued, long-term wear can cause painful bruising of the skin, making them unsuitable for long-term occupational use. This NIH R21 project proposes the develop- ment of a new type of personal respiratory protection device called the Smart, Individualized, Near-face, Ex- tended Wear (SINEW) Mask. The proposed design uses electrostatic filters to overcome many of the barriers of traditional masks while providing a high degree of protection. Using an N95 FFR significantly increases breathing resistance which is problematic for workers performing physically demanding tasks. To avoid increasing breath- ing resistance, the innovative design removes particles from airflow, while the mask does not come in contact with the skin of the wearer’s face. Instead, it uses a comfortable headband to keep the mask in place. The filtering action takes place along the entire perimeter of the mask, 3 to 10 mm from the face surface. Particles and droplets present in the air are removed with electrostatic forces. While this principle is common for industrial electrostatic precipitators (ESP) and is in HVAC equipment, a wearable filter for the face using electrostatic precipitation is a novel approach, enabled only by recent miniaturization of power electronics and advances in material science, specifically in catalytic coatings. Resistance to airflow is negligible because of the fundamental feature of the design – the gap between the mask and the face lets air through freely while removing the harmful particles in both the inhalation and the exhalation cycles. This feature is very important for exercise and for comfortable breathing by workers with health conditions (e.g., asthma) that would prevent them from using a traditional FFR. Other advantages of the design include a clear face shield, which allows people to read the lips and facial expressions of the wearer. The SINEW Mask is sterilizable and reusable, so it does not contribute to the escalating global waste problem of disposable masks. The recent COVID-19 pandemic demonstrated that uncomfortable masks lead to reduced usage. SINEW Masks can improve the long-term health of firefighters, healthcare workers, and any worker who needs comfortable, sterilizable, reusable protection against harmful particulate matter.

NIH Research Projects · FY 2024 · 2023-09

ABSTRACT Numerous promising investigational therapies for the treatment of epilepsy have been identified using well- established animal models of seizure and epilepsy for over 80 years. In this time, the maximal electroshock test in mice and rats, the subcutaneous pentylenetetrazol test in mice and rats, and more recently the 6 Hz assay in mice, have all been utilized as primary models of electrically or chemically evoked seizures in neurologically intact rodents. In addition, rodent kindling models, in which network hyperexcitability has developed, have been used to identify new and highly impactful agents, such as levetiracetam. This screening approach has successfully identified several marketed drugs to manage the symptomatic seizures associated with epilepsy. Despite the numerous antiseizure drugs (ASDs) on the market today, nearly 30% of patients with epilepsy are resistant to these currently available medications. Further, no treatment has yet been identified to slow or prevent the development of epilepsy altogether. To address this unmet medical need, more effective and better tolerated treatments are still desperately needed by the patient with epilepsy. Our group has recently uncovered the antiseizure efficacy of several repurposed therapeutic agents that engage untapped molecular targets associated with normal nervous system development and tissue homeostasis in adulthood. Further, we have demonstrated in a mouse model of evoked chronic seizures that repeated administration of repurposed agents is well-tolerated and can possibly disrupt the formation of a hyperexcitable neuronal network, suggesting a possible disease modifying effect. This study will therefore extend the antiseizure efficacy profile of these two promising investigational compounds for the treatment of symptomatic seizures to establish a pharmacokinetic and pharmacodynamic relationship, as well as assess their potential to prevent epilepsy altogether in well-established rodent seizure and epilepsy models. We will use a clinically relevant rat model of acquired epilepsy that provides a valid platform on which to evaluate the disease-modifying potential of two promising candidate compounds. This study aims to further characterize the therapeutic potential of two promising compounds for the treatment of acute seizures, as well as define the extent to which treatment modifies the onset of spontaneous recurrent seizures, i.e., demonstrate whether epilepsy is delayed or prevented. Further, this study will determine whether the use of novel repurposed agents with promising preliminary efficacy profiles demonstrate the potential to be first-in-class therapies for the treatment, and possibly prevention, of epilepsy.

NIH Research Projects · FY 2024 · 2023-09

PROJECT ABSTRACT Colorectal cancer (CRC) is the third leading cause of cancer-related death in the US, with pronounced disparities in mortality by race and ethnicity. Disparities are particularly pronounced in Alaska Native and African American people with mortality rates being 2 to 3 times higher. These disparities cannot be explained by access to care alone, suggesting other contributing factors like molecular subtypes and cellular heterogeneity, need to be uncovered. Tumor profiling study in patients from multiple racial and ethnic groups will lead to novel biological insight into CRC, which greatly overcomes the limitation of previous studies that were conducted predominantly in non-Hispanic white people only. Tumor immune contexture, which refers to the spatial organization and density of the immune infiltrates within the tumor microenvironment, is complex and associated with cancer prognosis. Beyond bulk analysis, cutting-edge spatial single-cell analyses enable us to evaluate tumor immune contexture within different regions of tumor tissues (e.g. margin, center), which will add our knowledge of immune evasion and antitumor immunity that is critical for CRC survival. My objective is to better understand the molecular and cellular landscapes driving tumor progression in a racial- and ethnic-sensitive manner. This will provide me with hand-on, multi-disciplinary training in the new field of integrative tumor epidemiology. I will leverage data from a nested case-control study that includes 840 Alaska Native, African American, Hispanic and non-Hispanic White CRC patients. Each racial and ethnic group includes 70 lethal cases who died of CRC and 140 CRC controls who survived at least as long as the case to which they are matched for age, sex and stage. In the F99 phase, I will examine molecular signatures and develop a prognostic index across four racial and ethnic groups using RNAseq data. I will evaluate tumor immune contexture among Alaska Native patients using spatial-resolved single-cell technology (Akoya PhenoCycler). In the K00 phase, I will integrate transcriptomic and single-cell data into patient-level data to characterize tumor heterogeneity and its role in other prognostic factors (e.g. treatment, recurrence). Results from this project will capture a full spectrum of CRC transcriptional and cellular biology and their relationship to disease outcomes through a spatial lens and provide clinical-useful prediction tools for prognosis that can be tailored to racially- and ethnically-diverse patients. This will help to address longstanding racial and ethnic disparities in CRC mortality. This project provides me with training opportunities to develop expertise in 1) molecular epidemiologic working with racial and ethnic diverse populations, especially with the engagement of Alaska Native people, 2) cancer immunology, 3) statistical and computational analysis in high- dimensional data. Those experiences will greatly help my transition into a post-doctoral fellow and eventually an independent cancer researcher in the rapidly growing field of integrative tumor epidemiology.

NIH Research Projects · FY 2025 · 2023-09

Project Summary We are submitting this proposal in response to PAR-20-147 (Extracellular RNA carrier subclasses in processes relevant to Substance Use Disorders or HIV infection). The opioid epidemic in the United States has far reaching consequences for public health. In recent decades, the rate of newborns born with neonatal abstinence syndrome (NAS) has increased dramatically. However, not all neonates exposed to opioids during gestation develop NAS or require treatment. NAS is a complex and idiosyncratic condition, and it is currently not possible to predict even at birth which infants will require pharmacotherapy or be susceptible to its worst outcomes. Accessible biomarkers for fetal brain and CNS development during opioid exposure would be highly valuable for deciding which infants will need opioid therapy for NAS and making personalized risk assessments for opioid exposed infants. They could also shed light on the biological pathways that lead to the worst effects of opioid exposure during gestation. Extracellular vesicles (EVs) derived from the Fetal brain and central Nervous system (FNEV) can cross the fetal/placental barrier and can be isolated from maternal blood. They represent a powerful accessible tool to provide insight on fetal neurodevelopment and damage. However, EVs are highly heterogeneous, with a diverse set of surface proteins and intra-vesicular cargo, such as RNA. Current approaches to the isolation and study of EVs lack the necessary sensitivity and precision to characterize EV subpopulations. To address the current limitations of EV analyses, we developed a high-throughput single-EV flow analyzer and sorter capable of detecting single dye molecules. This new technical capability allows us to exquisitely define the profiles of plasma FNEV during pregnancy. Our studies could lead to assays that define NAS risk pre-birth and bring new insights into the mechanistic pathways of fetal brain injury.

NIH Research Projects · FY 2026 · 2023-09

ABSTRACT. Stark disparities in cardiometabolic health outcomes exist by racialized identity and income level. Low-income populations experience 40-70% higher risk for cardiovascular mortality compared to higher-income populations. Black and Latinx populations experience higher rates of at-risk cardiovascular profiles compared to White populations. Diet quality is hypothesized to play a key role in these disparities. But consuming a health- promoting diet is out of reach for many Americans, and disproportionately so for low-income populations and people of color. Structural and economic factors make high-quality diets out of reach for many. Specifically, the intersecting experience of food insecurity, poverty, structural racism, precarious employment, high cost, and difficult to access healthier foods creates a discriminatory and unjust context where accessing, purchasing, and eating a health-promoting diet, particularly one high in fruits and vegetables, is unattainable. As a result, interventions and policies that reduce these structural and economic barriers have high potential to improve food security, dietary quality, and ultimately improve health. To this end, the federal government, several states, and municipalities have introduced policy interventions that provide financial incentives to promote the purchase of fruits and vegetables by low-income populations. However, several prominent design features of these programs leave many of the structural causes of food insecurity and low fruit and vegetable consumption in place. In this study, we can improve what is known about the equity and impact of fruit and vegetable incentive programs through the study of Seattle's Fresh Bucks Program. This program has been designed with several features that have the potential to overcome the limitations of previous programs. Methodologically, we propose a mixed methods study that leverages randomized assignment for assessing causal impact, includes participatory approaches and a sequential explanatory mixed methods design to assess equity in implementation, and a quasi-experimental design to utilize electronic health records of program participants and a matched comparison population to estimate associations with long-term (2-4 years) program enrollment and health outcomes in the following Aims: 1) Determine the impact of Fresh Bucks on fruit and vegetable intake and food insecurity, leveraging randomized program assignment for causal impacts. 2) Assess Fresh Bucks implementation equity through redemption rates and experience. 3) Assess the impact of long-term access to Fresh Bucks on cardiometabolic health, including blood pressure and BMI, in a cohort of recipients and family members with matched electronic health records. Decreasing food insecurity and improving diet quality among low-income and minoritized populations is of primary importance for policymakers concerned about reducing systemic barriers to health. Upon completion of this work, we will have substantially advanced what is known about the dietary, health and implementation equity impacts of fruit and vegetable incentive programs and the potential for these policies to reduce systemic barriers to health.

NIH Research Projects · FY 2025 · 2023-09

ABSTRACT Numerous promising investigational therapies for the treatment of epilepsy have been identified using well- established animal models of seizure and epilepsy for over 80 years. In this time, the maximal electroshock test in mice and rats, the subcutaneous pentylenetetrazol test in mice and rats, and more recently the 6 Hz assay in mice, have all been utilized as primary models of electrically or chemically evoked seizures in neurologically intact rodents. In addition, rodent kindling models, in which network hyperexcitability has developed, have been used to identify new and highly impactful agents, such as levetiracetam. This screening approach has successfully identified several marketed drugs to manage the symptomatic seizures associated with epilepsy. Despite the numerous antiseizure drugs (ASDs) on the market today, nearly 30% of patients with epilepsy are resistant to these currently available medications. Further, no treatment has yet been identified to slow or prevent the development of epilepsy altogether. To address this unmet medical need, more effective and better tolerated treatments are still desperately needed by the patient with epilepsy. Our group has recently uncovered the antiseizure efficacy of several repurposed therapeutic agents that engage untapped molecular targets associated with normal nervous system development and tissue homeostasis in adulthood. Further, we have demonstrated in a mouse model of evoked chronic seizures that repeated administration of repurposed agents is well-tolerated and can possibly disrupt the formation of a hyperexcitable neuronal network, suggesting a possible disease modifying effect. This study will therefore extend the antiseizure efficacy profile of these two promising investigational compounds for the treatment of symptomatic seizures to establish a pharmacokinetic and pharmacodynamic relationship, as well as assess their potential to prevent epilepsy altogether in well-established rodent seizure and epilepsy models. We will use a clinically relevant rat model of acquired epilepsy that provides a valid platform on which to evaluate the disease-modifying potential of two promising candidate compounds. This study aims to further characterize the therapeutic potential of two promising compounds for the treatment of acute seizures, as well as define the extent to which treatment modifies the onset of spontaneous recurrent seizures, i.e., demonstrate whether epilepsy is delayed or prevented. Further, this study will determine whether the use of novel repurposed agents with promising preliminary efficacy profiles demonstrate the potential to be first-in-class therapies for the treatment, and possibly prevention, of epilepsy.

NIH Research Projects · FY 2025 · 2023-09

People with HIV (PWH) carry a disproportionate burden of consequences related to substance use and HIV. The goal of this study is to better understand substance use and poor health outcomes among people with HIV (PWH) including substance use patterns, substance use treatment outcomes and HIV-related outcomes including comorbidities such as diabetes. We are particularly interested in those experiencing unstable housing, recently incarcerated, lacking social support, facing HIV-related stigma as well as other subpopulations (many of whom have high rates of substance use) as well as the impacts of use on chronic comorbidities such as diabetes. We will 1. Determine methamphetamine, cocaine and opioid-related behaviors, risk factors, and treatment outcomes to better tailor future intervention/treatment and prevention approaches. We will examine differences in drug-related behaviors including types, frequency, severity of use, mode of administration, polydrug use, overdoses, naloxone access, and substance use treatment access and outcomes. We will determine the role of demographic, behavioral, psychosocial and clinical factors, and incorporate additional biobehavioral measures based on recommendations from community partners, end-users, and critical thought partners to determine risk factors for differences in drug use patterns and substance use treatment outcomes. 2. Determine the role of methamphetamine, cocaine and opioid use in HIV-related outcomes overall and among key subpopulations such as those with unstable housing. We will examine longitudinal relationships between drug use, biobehavioral factors, and HIV care cascade steps such as engagement in care, viral suppression and health-related quality of life. We will determine the impact of drug use and other factors on age-related comorbidities, such as diabetes, frailty, myocardial infarction and chronic kidney disease. We leverage the large well-characterized cohort of PWH with comprehensive clinical data in the Centers for AIDS Research Network of Integrated Clinical Systems (CNICS) cohort. The CNICS patient reported outcomes assessment captures detailed drug use patterns, psychosocial, behavioral and other factors such as social support and housing status. CNICS geocoding and carefully adjudicated clinical outcomes facilitates including biobehavioral and other factors and evaluation of HIV-related comorbidities such as myocardial infarction. CNICS data and our team bring a breadth and depth of resources, responding to RFA-DA-23-061 by generating new knowledge about substance use impacts among PWH and the impact of clinical and other factors. This study will inform future research and assist end-users within our community partnerships in tailoring future substance use treatment programs to the unique needs associated with PWH in order to improve outcomes at the intersection of HIV care and substance use.

NIH Research Projects · FY 2025 · 2023-09

ABSTRACT ECHO Yakima Valley enriches the ECHO Program in two primary ways. 1) We will increase representation of rural and agricultural communities in the national ECHO Cohort so it may better support child health research across the range of American communities. With deep research ties to Washington's Yakima Valley, our team is poised to enroll over 1000 pregnant women into the ECHO Cohort through partnerships with their prenatal and pediatric healthcare home. Biospecimen collection will be led by a sub-team with experience establishing and directing a biorepository for over 10,000 pregnancies. 2) Our investigator team spans various career stages and specialties and will implement an interdisciplinary approach to ECHO papers, incorporating insights from environmental health science, biostatistics, epidemiology, and pediatric medicine. We will apply our expertise to advance ECHO science to describe the relationships between commonly encountered pollutants (including fine particulate matter, ozone, and nitrogen dioxide) and major chronic pediatric health concerns, such as preterm birth, low birthweight, respiratory infections, asthma development and exacerbation, obesity, blood pressure, and neurodevelopment. We propose to lead a new ECHO-wide interest group on Air Pollutants and Wildfire Smoke as well as an interest group to bring together expertise in Rural Health. We will aim to develop new ECHO manuscripts to understand child health risks related to mixtures of common air pollutants, pesticides, and drinking water contaminants. We have plans for a set of novel preconception analyses addressing contaminants of concern for drinking water wells, an underdeveloped topic for ECHO. We also propose to expand ECHO Program data resources with advanced models for estimating participant exposures to ultrafine particulate matter and wildfire smoke. We plan to lead development of brief, low-burden extensions to the ECHO Protocol to better characterize family knowledge of the Air Quality Index (AQI). These data will allow ECHO scientists to explore opportunities to improve access to this public health tool as well as evaluate its effectiveness in mitigating health effects. A proposed novel Wildfire Rapid Response Protocol will enable natural experiment analyses of wildfire related health crises nationwide. Overall, ECHO Yakima Valley will build on our team's prior successes and insights we gained as the ECHO PATHWAYS multi-cohort award center point for project coordination and team science. We will continue these efforts in ECHO Cycle 2 with a focus on solution-oriented research that may efficiently and directly inform public health policy and programs.

NIH Research Projects · FY 2025 · 2023-09

SUMMARY Von Willebrand factor (VWF) has two major roles in blood. One is to facilitate platelet adhesion and aggregation, in which a critical step is to activate the VWF A1 domain to bind with platelet protein GPIbα under flow. The other is to protect coagulation factor VIII (FVIII) from degradation, which is important for fibrin clot formation. Mutations in VWF interfering with these binding interactions can cause thrombosis or von Willebrand disease. The goal of this project is to determine molecular mechanisms governing the interactions between VWF and its binding partners GPIbα and FVIII. Several questions regarding VWF interactions with GPIbα and FVIII persist. There has not been a consistent model for how VWF A1 domain is activated to bind with GPIbα. It still remains unclear which and how conformational changes actually happen during A1 activation. To date, the interaction between VWF multimer and FVIII has been studied primarily using bulk VWF preparations. How FVIII binds to individual VWF multimers is unknown. In intravenous infusion treatment for bleeding disorders, more free FVIII in the infusion is linked to higher risk of FVIII immunogenicity, but the mechanisms remain unclear. In this project, we will combine single molecule biophysics and super resolution imaging methods to study the following aims. (1) Understand flow-induced activation of VWF A1 binding with GPIbα. (2) Dissect the VWF- FVIII binding mode and its role in protecting FVIII in blood. This study will provide direct evidence for the mechanisms of how VWF binds with GPIbα and FVIII, and provide new insights into therapeutic development to treat thrombotic or bleeding disorders.

NIH Research Projects · FY 2025 · 2023-09

The public health crisis of overdose continues in King County, WA, with over 500 deaths in 2020, 700 in 2021, and rates remaining subsequently high. Emergency Medical Services (EMS) providers are often the first point of contact for individuals experiencing overdose and play a critical role in connecting people who use drugs (PWUD) to care. In partnership with PWUD, EMS leadership, and community stakeholders, our interdisciplinary team collected pilot data and co-designed EMS-delivered interventions aimed at reducing negative perceptions of PWUD and improving access to evidence-based care. These interventions are being implemented as the EMS Overdose Prevention Project (EMS-OPP), which includes EMS provider training in reducing negative perceptions of PWUD and trauma-informed care, a naloxone leave-behind program with fentanyl test strips, and warm hand-offs to follow-up teams for substance use treatment. King County EMS plans to iteratively adopt EMS-OPP, with a goal of 90% team participation by 2026. To evaluate the impact of EMS-OPP, we are conducting a hybrid implementation-effectiveness study using a concurrent triangulation mixed methods design. Our revised study assesses whether EMS-OPP improves overdose-related outcomes for the entire population of overdose survivors. Specifically, we aim to: Evaluate the effect of EMS-OPP on patient-level experiences (e.g., perceptions of EMS care) and health outcomes (e.g., linkage to buprenorphine treatment) through a prospective cohort study of non-fatal overdose survivors (Aim 1); Understand the experiences of overdose survivors with EMS-OPP using in-depth qualitative interviews (Aim 2); and Examine the population-level impact of EMS-OPP on outcomes such as follow-up care using an interrupted time series design with switched replication (Aim 3). This study capitalizes on a unique moment in the opioid crisis, as EMS systems expand their scope and capacity. Findings will inform future EMS interventions and data linkage strategies to support long-term surveillance of overdose outcomes. Dissemination will include community-oriented products (e.g., zines, artistic data posters), policy briefs, conference presentations, and peer-reviewed publications.

NIH Research Projects · FY 2024 · 2023-09

A major driver for immunogenicity is the presence of T-cell epitopes within a protein sequence that can activate helper T cells, resulting in neutralization of the therapeutic effect. For DMD gene therapy, the possibility of T-cell responses to pre-existing (from revertant fibers) &/or post-treatment dystrophin epitopes has recently emerged as a serious treatment issue. In the past year 5 DMD patients treated with AAV-microdystrophin (µDys) have developed T cell mediated immune responses against dystrophin. Based on immunological data, in all cases the antigenic region was highly suggested to be encoded by exons 8-11.1 Additionally, to being observed in patients deleted for this region (updates provided at MDA and ASGCT 2022 scientific conferences). A key component of our approach takes advantage of the fact that a single amino acid can anchor a novel peptide for a preferred binding state for antigen presenting cells’ via its major histocompatibility complex (MHC) and subsequent T cell receptor (TCR). It is the TCR recognition which leads to downstream signaling cascades and immune sequelae. Alternatively, conservatively modified residue(s), can disfavor antigen presentation, preventing dystrophin- specific immunity. Thus, avoidance of a specific immune response can be achieved if key epitope anchor residue(s) necessary for antigen presentation are modified to disrupt MHC recognition. Here, our proposal addresses the fundamental role of immunogenicity against dystrophin that has been found in DMD patients. This proposal integrates deimmunization protocols and structure-based protein redesign via Rosetta-software, with streamlined functional testing via our magnetic tissue array (Mantarray) platform-allowing for daily 3D DMD contractile performance assessment in-a-dish. In turn, dystrophin epitopes can be predicted & ranked according to their antigenicity by correlating their amino acid sequence content with immunological databases. Known and predicted epitopes will be targeted for redesign to reduce or eliminate putative MHC-binding peptides and subsequent T cell activation. Our deimmunization approach will build on previous efforts while focusing on exons 8-11 of dystrophin, & integrating data as it becomes available through any DMD clinical trial alliance that unfolds. The exon 8-11 region, now part of Pfizers’ updated patient exclusion criteria, encodes a portion of the actin- binding domain, extending through spectrin-like repeat 1 (note: we have already deimmunized much of exons 6- 8). Importantly, restricting deimmunization efforts to exons 8-11 of dystrophin would be beneficial for all µDys designs currently in clinical trials. In summary, we propose to deliver deimmunized µDys vectors to human DMD 3D engineered muscle tissues and measure contractile force(s) via our Mantarray system in an effort to screen for highly functional leading candidates with reduced immunogenicity. To fully validate our 3D muscle system as a predictive model we will test optimized candidates(s) via rAAV vectors driven by highly muscle specific regulatory cassettes for their capacity to ameliorate disease progression in the DMDmdx rat model of DMD. These results will provide necessary preclinical outcome metrics required to advance towards Phase 1 clinical trials.

NIH Research Projects · FY 2025 · 2023-08

PROJECT SUMMARY/ABSTRACT COVID-19 vaccination reduces the risk of SARS-CoV-2 infection and severe disease, but almost half of hospitalized breakthrough cases are in immunocompromised individuals. HIV infection is an independent risk factor for severe COVID-19, hospitalization, and mortality. Immunocompromised individuals are more likely to have prolonged SARS-CoV-2 infection and viral shedding, increasing the risk of viral transmission and allowing for rapid evolution of more virulent strains. Therefore, investigating SARS-CoV-2 pathogenesis in the context of immunosuppression is urgently needed to reveal factors driving severe COVID-19. SARS-CoV-2 lung pathogenesis is characterized by infiltration of innate and adaptive immune cells into the lung and induction of an inflammatory immune response. Similar mechanisms of inflammation and immune dysfunction during HIV infection contribute to systemic HIV pathogenesis and lung pathology. Therefore, dysregulation of immune responses during HIV infection could induce severe disease outcomes during SARS-CoV-2 coinfection. SARS- CoV-2 replication occurs in both respiratory and gastrointestinal mucosal sites and enteric symptoms are associated with COVID-19. There is a defined link between gastrointestinal microbial dysbiosis with accelerated HIV disease progression and an emerging role of lung and intestinal microbial dysbiosis with severe COVID-19. Given that the microbiome plays an important role in maintaining mucosal function and homeostasis, shifts in microbial communities due to HIV infection could contribute to inflammation and immune activation that would drive exacerbated SARS-CoV-2 pathogenesis. Here, we will test the hypothesis that increased immune dysfunction and exhaustion, inflammation, and microbial dysbiosis during HIV infection promote enhanced SARS-CoV-2 lung pathogenesis. We will leverage the pigtail macaque simian immunodeficiency virus (SIV) model of rapid HIV/AIDS and will test this hypothesis during a state of inflammatory untreated SIV infection and during primary and secondary SARS-CoV-2 viral challenges. We will evaluate the roles of SIV-induced immunosuppression and altered alveolar macrophage and neutrophil function on SARS-CoV-2 pathogenesis and pulmonary pathology. We will determine whether HIV infection causes gastrointestinal tract and lung microbial dysbiosis and its association with SARS-CoV-2 disease severity. Lastly, we will determine whether increased immune exhaustion during HIV infection promotes deficits in the generation of primary anti-viral SARS- CoV-2 responses that impairs protection from heterologous SAR-CoV-2 re-challenge. These studies will contribute to our understanding of how normal immunity drives SARS-CoV-2 lung pathophysiology and will dissect how perturbations of immune responses during HIV infection contribute to enhanced disease.