University Of Washington

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT Clinical genetic testing, including exome sequencing, has diagnosed many rare and novel genetic disorders. These diagnoses have helped guide treatment, suggest novel or precision therapies, and provide long-term prognostic information for individuals and their families. However, at least 50% of individuals with a suspected genetic disorder remain undiagnosed after a complete clinical evaluation, which can take years to complete. There are two major technical reasons for this low solve rate. First, structural variants such as large deletions, insertions, inversions, and repeat expansions account for a significant fraction of unsolved cases, but they are challenging to analyze using traditional short-read sequencing technologies. Second, many variants lie in recently duplicated genomic regions that are difficult to evaluate using short-read sequencing technologies. Recently, it has become economically feasible to pursue whole-genome long-read sequencing (LRS) of human genomes with sufficient coverage to analyze these complex genomic regions. This presents an opportunity to systematically evaluate the incremental diagnostic rate of LRS over current testing approaches and apply LRS technology to a large cohort of individuals with unsolved genetic disorders. Moreover, LRS is unique in that as a single test it has the potential to replace nearly all existing clinical genetic testing (microarray, exome, methylation studies), making it possible to perform a complete clinical genetic evaluation in days to weeks using a single dataset. The overall goal of this project is to establish a framework for the clinical adoption of long-read DNA and RNA sequencing by systematic evaluation of individuals with suspected or unsolved genetic disorders. To achieve this goal, LRS will be performed on 50 critically ill neonates and their parents who had nondiagnostic trio short-read whole-genome sequencing (WGS) and 25 trios from the same cohort who had diagnostic WGS (Aim 1). This will help establish the incremental diagnostic rate of LRS over short-read WGS. To evaluate the technical advantages of LRS over traditional clinical testing, I will sequence 50 individuals with a suspected genetic disorder who remain unsolved after at least clinical exome sequencing (Aim 2). In up to 20 persons that remain undiagnosed after LRS, long-read RNA sequencing will be used to identify high-priority regions that influence gene expression or splicing for further analysis. I will then evaluate the sensitivity of LRS as a single genetic test by sequencing 50 individuals undergoing a typical outpatient clinical evaluation (Aim 3). Demonstrating that LRS can be used as a single data source will help streamline the current stepwise approach to clinical testing, leading to an increase in the diagnostic rate and an overall cost savings to the health system through reduced testing and clinic visits. Overall, the aims outlined in this proposal will benefit patients and families by increasing the rate of genetic diagnosis, decreasing the time to diagnosis, and identifying novel disease-causing mechanisms, which will lead to a better understanding of the causes of human disease.

NIH Research Projects · FY 2025 · 2022-09

The long-term objective of this K01 Mentored Research Scientist Development Award is to support Dr. Tessa Frohe, in building an independent research career focused on adapting, designing, and implementing efficacious telehealth alcohol interventions among communities with lived and living experience of substance use and/or homelessness. To date, Dr. Frohe’s research has focused on risk factors associated with pain and substance use. During her postdoctoral training, she has begun working within a community-based and qualitative methods framework to examine alcohol use specifically among people experiencing homelessness and AUD; however, Dr. Frohe seeks to expand her training from basic alcohol research to implementing and testing technology-based interventions informed by community stakeholders and user-centered design to improve treatment delivery among this population. This long-term objective will be achieved through a five-year training and research plan involving a carefully selected mentor team, targeted coursework and hands-on training experiences. The proposed research aims to adapt an in-person harm reduction intervention (HaRT-A) into a telehealth platform (eHaRT-A) and test it among individuals with lived experience of homelessness and AUD. This project will occur in two phases: Phase 1 will entail codevelopment with iterative usability testing of the telehealth platform with a community advisory board (e.g., Housing First residents) to leverage stakeholder ideas to inform and build the eHaRT-A prototype (Aim 1) and to make design changes that will improve the feasibility and acceptability of the eHaRT-A platform (Aim 2). In Phase 2, a randomized controlled trial will be conducted to test the efficacy of eHaRT-A compared to “supportive services as usual” in improving alcohol-related outcomes (i.e., peak alcohol use, alcohol-related harm, AUD symptoms, and positive urinary ethyl glucuronide tests) and health related quality of life over time (Aim 3). This proposal aligns with national (NIH; NOT-AA-20-011) health initiatives to integrate technology-based interventions among permanent supportive housing communities with pre-existing substance use treatments. If successful, this study will develop a clinically relevant intervention that is more easily transportable within many different community settings because it will be developed for, by, and with the community it aims to serve. The training plan for this application will focus on intervention development and testing, innovative methods to enhance technology-based intervention implementation, and advanced statistics. Mentors (Drs. Clifasefi, Collins, Comtois, Hsieh) and collaborators (Drs. Larimer, Hallgren) are committed to Dr. Frohe’s training and each will provide unique expertise to her proposed research and training plan. Support from this award will be essential to Dr. Frohe’s development as an independent scientist who can contribute to alcohol research by codeveloping, adapting, and testing successful technology-based interventions to ameliorate alcohol-related harm and improve individuals’ quality of life. The University of Washington is well suited to provide a stellar training experience and will promote NIH’s mission to develop early investigators who can be competitive for long-term research funding.

NIH Research Projects · FY 2025 · 2022-09

Engaging the most vulnerable people with HIV in treatment is central to efforts to end the HIV epidemic. Low- barrier care (LBC) is an evidence-based multicomponent intervention that includes walk-in access to medical care, incentives for clinic visits and viral suppression, intensive support to address social determinants of health, and multisectoral service coordination. The Max Clinic in Seattle is a flagship LBC clinic that has substantially improved viral suppression among people with complex barriers to care, including unstable housing, substance use, and mental health disorders. Expanding LBC is central to the Ending the HIV Epidemic (EHE) plan in King County, Washington. The overall goals of this 5-year proposal are to study LBC expansion in King County, identify factors that facilitate LBC implementation, and prepare for the intervention’s scale-up in other EHE jurisdictions. The team will employ a multifaceted implementation strategy to establish and improve two new LBC clinics in addition to the Max Clinic. We will use the Reach, Effectiveness, Adoption, Implementation & Maintenance (RE-AIM) framework and structured tools to assess intervention adaptation and scalability. For Aim 1, we will evaluate the reach and effectiveness of LBC expansion in King County using a population-based observational open cohort design. For Aim 2, we will describe the implementation and maintenance of LBC in King County, including intervention adaptations, organizational strategies to overcome barriers, and costs. Using the Framework for Reporting Adaptations and Modifications-Enhanced (FRAME), we will conduct focus group discussions with key stakeholders in the LBC clinics and the health department at baseline and conduct brief re- assessments in six-month intervals over four years to describe intervention adaptations and elucidate implementation and maintenance strategies. We will quantify the costs of LBC expansion from the health department and healthcare organization perspectives using micro-costing methods. For Aim 3, we will assess the scalability of the LBC intervention and identify a consortium of health department and clinic leaders to scale- up LBC in cities and states prioritized for the first phase of EHE funding throughout the U.S. We will partner with NASTAD (the National Association of State & Territorial AIDS Directors) to identify health department HIV program leaders supportive of LBC implementation, and in collaboration with them, recruit leaders of Ryan White-funded HIV Clinics or Community Health Center clinics. We will conduct in in-depth interviews with both the health department and clinic leaders using the Intervention Scalability Assessment Tool (ISAT) for applied assessment of LBC implementation considerations in each local context. Expected Outcome: Our results will guide optimization of LBC implementation, inform intervention adaptations, and lay the groundwork for strategic scale-up of LBC in other parts of the country, which will in turn, advance progress toward achieving EHE goals.

NIH Research Projects · FY 2025 · 2022-09

Solid tumors progress towards metastasis as their cells diversify in space and genotype. Unfortunately, by the time nearly all solid tumors are discoverable, the dynamics governing this diversification are obscured. To overcome this limitation, I will develop methodology that enables both reconstruction of key diversification dynamics throughout a tumor’s past and robustly predicts which current tumor cellular lineages, if unmitigated, will permit tumor progression towards poor clinical outcomes; i.e., a tumor ‘time machine’. In this proposal, I describe how I will construct such a time machine from phylodynamic methodologies, which synthesize phylogenetic and population dynamic models and were originally developed for the study of viral evolution and geographic spread. To make such methods suitable for the analysis of tumor sequencing data, I will extend these models to incorporate multiple attributes of tumor growth dynamics. I will apply these new tools at scale to analyze cancer evolutionary processes, and develop novel diagnostic tools to yield clinically actionable insights into tumor progression. Crucially, application of these methods relies on cancer single cell DNA sequencing data, which has recently gained prominence and reveals a much more detailed view of cancer evolution than previous tumor sequencing technologies. I hypothesize that these data will yield still greater insight when subjected to the rigorous, quantitative examinations enabled by the cancer-calibrated phylodynamic methodology I will develop. In applying these new tools, I will derive precisely-calibrated clinical timelines, including estimates of tumor initiation and the age of important clonal subpopulations, which will help us understand personalized tumor progression. I will disentangle heterogeneous cancer growth rates from within a single tumor, enabling both the prediction of the clinically most important variants and a new approach for discovering driver mutations. These phylodynamic methods will also illuminate how and when cancer cells disperse within a primary tumor as a function of genotype and/or environment, enabling us to chart the development of metastatic lineages. In addition to answering fundamental questions about the driving forces shaping tumorigenesis and cancer progression, this proposal aims to provide a powerful new set of methodologies that will drive a paradigm shift in the analysis of tumor single cell sequencing data in diverse basic science, clinical and translational cancer settings.

NIH Research Projects · FY 2024 · 2022-09

Engaging the most vulnerable people with HIV in treatment is central to efforts to end the HIV epidemic. Low- barrier care (LBC) is an evidence-based multicomponent intervention that includes walk-in access to medical care, incentives for clinic visits and viral suppression, intensive support to address social determinants of health, and multisectoral service coordination. The Max Clinic in Seattle is a flagship LBC clinic that has substantially improved viral suppression among people with complex barriers to care, including unstable housing, substance use, and mental health disorders. Expanding LBC is central to the Ending the HIV Epidemic (EHE) plan in King County, Washington. The overall goals of this 5-year proposal are to study LBC expansion in King County, identify factors that facilitate LBC implementation, and prepare for the intervention’s scale-up in other EHE jurisdictions. The team will employ a multifaceted implementation strategy to establish and improve two new LBC clinics in addition to the Max Clinic. We will use the Reach, Effectiveness, Adoption, Implementation & Maintenance (RE-AIM) framework and structured tools to assess intervention adaptation and scalability. For Aim 1, we will evaluate the reach and effectiveness of LBC expansion in King County using a population-based observational open cohort design. For Aim 2, we will describe the implementation and maintenance of LBC in King County, including intervention adaptations, organizational strategies to overcome barriers, and costs. Using the Framework for Reporting Adaptations and Modifications-Enhanced (FRAME), we will conduct focus group discussions with key stakeholders in the LBC clinics and the health department at baseline and conduct brief re- assessments in six-month intervals over four years to describe intervention adaptations and elucidate implementation and maintenance strategies. We will quantify the costs of LBC expansion from the health department and healthcare organization perspectives using micro-costing methods. For Aim 3, we will assess the scalability of the LBC intervention and identify a consortium of health department and clinic leaders to scale- up LBC in cities and states prioritized for the first phase of EHE funding throughout the U.S. We will partner with NASTAD (the National Association of State & Territorial AIDS Directors) to identify health department HIV program leaders supportive of LBC implementation, and in collaboration with them, recruit leaders of Ryan White-funded HIV Clinics or Community Health Center clinics. We will conduct in in-depth interviews with both the health department and clinic leaders using the Intervention Scalability Assessment Tool (ISAT) for applied assessment of LBC implementation considerations in each local context. Expected Outcome: Our results will guide optimization of LBC implementation, inform intervention adaptations, and lay the groundwork for strategic scale-up of LBC in other parts of the country, which will in turn, advance progress toward achieving EHE goals.

NIH Research Projects · FY 2025 · 2022-09

ABSTRACT Elucidating the underlying genetic causes of the awesome phenotypic diversity observed in natural populations is a major challenge in biology. Despite the importance of understanding the genetic basis of complex traits, we currently lack complete knowledge of the relevant genetic components. Genetic variation affecting the structure and copy number of chromosomal segments is an underappreciated component of the genetic architecture. In this proposal, we seek to obtain a species-wide view of structural variation (SV) and copy number variation (CNV) and how these contribute to the phenotypic landscape of natural populations. In order to accomplish this, we will take advantage of the genetic workhorse Saccharomyces cerevisiae, for which we have recently completed whole genome resequencing of 1,011 natural isolates, plus accompanying large scale phenotyping efforts. CNVs are frequent in our strain collection and have outsize phenotypic affects in the association tests we have so far completed. We propose to: Aim 1. Apply long read sequencing to better characterize the structural variation present in these strains and generate high quality de novo assemblies. In Aim 2, we will collect quantitative strain phenotypes over a large panel of environmental conditions, with a particular emphasis on conditions that we and others have found to be particularly sensitive to CNVs. We will also include molecular traits such as mRNA and protein abundance. We will then perform association testing with the variants discovered in Aim 1 in order to determine over an entire species the degree to which SVs and CNVs contribute to trait variation. Since association methods perform poorly for rare genetic variants, we will in Aim 3 use a diallel panel where controlled crosses allow us to better survey all variation present. Finally, in Aim 4 we will complement our work on naturally occurring variation by building libraries of engineered variants. We will create large dosage series pools of segmental amplifications and deletions to better understand their consequences across multiple genetic backgrounds. Our work will result in a more complete understanding of the complexities of the genotype-phenotype connection with respect to this important but understudied class of genetic variants.

NIH Research Projects · FY 2026 · 2022-09

PROJECT SUMMARY & ABSTRACT The glycocode, or collection of holistic features arising from specific combinations of glycan and protein biomolecules, is a currency used at the cell surface to exchange information and direct biological processes. The unique molecular surfaces created by specific glycan-protein combinations are a critical axis of information, yet they remain poorly understood due to numerous analytical challenges associated with studying their complexity. Emerging evidence from immuno-oncology, virus pathobiology, and beyond underscore that this blind spot can no longer be afforded. This proposal generates a suite of new technologies to capture the combinatorial patterns of the glycocode using innovations in mass spectrometry (MS) and chemical glycoproteomics. Together these will enable systems-scale interrogation of the human glycocode. Aim 1 introduces a new platform for cell surface glycoproteomics that retains glycan-protein patterns rather than relying on traditional protein-centric methods that ignore the glycan component. By synthesizing novel chemical probes that append an easily enrichable phosphonate handle to cell surface glycoproteins through live cell labeling, we provide a chemical glycoproteomics platform for “catch-and-release” enrichment of intact glycoconjugates that provides access to combinatorial glycocode features. Aim 2 accompanies our chemical glycoproteomics approach with advances in the MS methods used to identify and quantify glycocode constituents (i.e., glycopeptides, glycoproteins, and glycans). Through the use of real-time analyses to enable adaptive instrument control, we improve the sensitivity and throughput of glycopeptide characterization. To complement bottom-up glycopeptide methods, we also establish a novel top-down approach to fingerprint combinatorial glycocode modifications on intact glycoproteins using ion-ion gas-phase chemistry to extract glycoform information from typically challenging denatured species. In Aim 3, we leverage these tools to define a human glycocode atlas across 12 different cell types to connect heterogeneous glycocode expression with specialized cellular functions. Furthermore, we extend this atlas to investigate dynamic glycocode reprogramming during epithelial-mesenchymal transition to discover cell-type specific signatures of migratory phenotypes that can be targeted in diagnostic and therapeutic strategies. In all, this proposal represents a significant advance in chemical glycoproteomics and MS methods, generates a long-needed resource for cell surface biology, and provides a robust foundation for me to build an independent research career focused on glycocode regulation in human health and disease. Key to accomplishing these aims will be training in chemical synthesis to make novel chemical glycoproteomic reagents during the K99 phase, which will equip me with the skills necessary to execute my vision for chemical glycoproteomics to investigate glycocode. Work proposed here will be fundamental in my transition to independence and will establish me as a leader in the promising field of systems glycobiology.

NIH Research Projects · FY 2024 · 2022-09

PROJECT SUMMARY/ABSTRACT Patients with Alzheimer’s disease and related dementias (ADRD) often experience high-intensity, potentially burdensome care near the end-of-life that may not align with their values and goals. Patients with ADRD and their families often face complex decisions about the most appropriate intensity of care. Goals-of- care discussions with clinicians are associated with improved patient and family outcomes and reduced intensity of care at the end of life. Although goals-of-care discussions can facilitate delivery of goal-concordant care, they currently occur infrequently or too late in the illness. Electronic health records (EHR) provide a key opportunity to identify patients who would benefit from these discussions and to promote these discussions, yet few prior interventions have used the EHR for this purpose. To address this gap, we propose an NIH Stage III hybrid efficacy-effectiveness trial of a communication-priming intervention, called Jumpstart, to increase the occurrence and quality of goals-of-care discussions with patients with ADRD and their family caregivers in the outpatient setting. We will use EHR data to identify patients and facilitate implementation of the intervention. In Aim 1, we will conduct a NIH Stage III hybrid efficacy-effectiveness trial of the adapted Jumpstart intervention compared to usual care to assess effectiveness for promoting occurrence and improving quality of goals-of care discussions and improving patient- and family-reported outcomes. The primary outcome is EHR documentation of a goals-of-care discussion by 3 months after enrollment. We will enroll 1200 patients with ADRD and their family caregivers. Aim 2 examines the mechanisms of action of the intervention on important patient- and family-centered outcomes including patients’ palliative care needs and the intervention’s effect on healthcare utilization, and monitors for unintended consequences of the intervention in terms of psychological distress and one-year all-cause mortality. In Aim 3, we will conduct a mixed-methods evaluation to explore further intervention improvements and identify barriers and facilitators to intervention implementation. Our team is well-positioned to accomplish these aims. We have experience conducting large, randomized trials with patient- and family-centered outcomes, biostatistics, qualitative and mixed methods analyses, ADRD research and policy, and implementation science methods. We have used informatics integrated with the EHR to identify patients with ADRD and measure quality of care similar to the methods proposed here, including measurement of the primary outcome. This study will provide important data for patients with ADRD, their family caregivers, clinicians, and healthcare systems on the efficacy of the Jumpstart intervention to improve communication and facilitate goal-concordant care and will provide insights for implementation of similar EHR-based interventions designed to improve communication and patient outcomes.

NIH Research Projects · FY 2024 · 2022-09

PROJECT SUMMARY/ABSTRACT The vascular endothelial growth factors (VEGFs) direct key signaling processes in obesity and at least 70 other diseases. However, focus on this signaling node alone has not achieved the promise of predictable angiogenic control. Current models are incomplete as other growth factors, besides VEGF, contribute to vascular disease progression, presenting a complexity that cannot be predictably regulated by targeting one node in this system. Therefore, there is a continuing need to account for the complexity of additional, multi-component signaling networks, a goal that can be achieved via data-driven, computational systems biology in close concert with experimental analysis of signaling and functional response. Toward this goal, we aim to examine a novel paradigm of network regulation called cross-family signaling, in which members from one growth factor family [e.g., platelet derived growth factors (PDGFs)] bind to and signal through members of another family (e.g., VEGFRs). We hypothesize that systematic examination of protein structure and downstream signaling within the cross-family paradigm via simulation, ligand-engineering, network quantification, and computational modeling can uncover novel mechanisms to control angiogenesis. We will test this hypothesis through three aims, sensitively quantifying receptor activation rates and functional responses of cross-family binding (e.g., proliferation, migration, and barrier function); predicting and measuring the structural properties of cross-family binding via molecular simulations and directed evolution; and developing validated deterministic models (mass-action kinetic modeling) of cross- family signaling and applying them to study and control the dynamics of cross-family signaling in human cell systems, in silico. We are primed to lead this new research because we are among the first to pursue this important theoretical paradigm, and we lead this cause to understand cell signaling via structure/function–based computational modeling. This work will catalyze a shift in perspective and innovation in the areas of cell signaling, systems biology, and predictive design of obesity- focused therapies.

NIH Research Projects · FY 2025 · 2022-09

Intracranial atherosclerotic disease (ICAD) can lead to ischemic stroke and there is increasing evidence that ICAD, even in the absence of stenosis, is associated with embolic stroke of undetermined source (ESUS). Vessel wall magnetic resonance imaging (MRI) of the intracranial vasculature is increasingly in demand to diagnose such ESUS patients so that appropriate treatment can be administered. Multi-contrast intracranial vessel wall (IVW) MRI with efficient analysis is currently recommended by the American Society of Neuroradiology to diagnose various vessel wall pathologies including ICAD. While this urgent need for IVW has stimulated availability of 3D spin echo sequences on major scanner platforms (VISTA on Philips, SPACE on Siemens and CUBE on GE), there are no efficient and effective methods for clinicians to analyze the multi-contrast IVW MRI sequences that can be obtained on modern clinical MRI scanners. Quantitative measurements of the vessel wall across scanner platforms are also required to enable multi-center studies for ICAD assessment in ESUS. Variability in sequence implementation affects multi-center studies on multiple scanner platforms and must be overcome to enable robust IVW measurements. Therefore, we propose to develop an automated IVW analysis pipeline for multi-contrast multi-platform application using a domain adaptive and deep learning approach. We have pioneered multiple semiautomatic approaches (3D-registration, artery tracing, artery labeling, multi-planar reformatting, vessel wall segmentation, multi-contrast feature identification) towards vessel wall quantification. Leveraging this expertise, we will develop a novel artificial intelligence (AI) empowered multiplanar viewing for artery characterization (AI-MOCHA) pipeline as follows: In Aim 1 we will construct the MOCHA pipeline and train AI-MOCHA using transfer learning from labeled IVW images and test AI-MOCHA against radiologist labeled IVW from ICAD patients. We will also test whether AI-MOCHA improves the inter-scan and inter-reader reproducibility of IVW image analysis. In Aim 2 we will develop domain adaptation to overcome scanner-platform differences in IVW images and develop a Domain Adaptive AI-MOCHA. We will then show that domain adaptive AI-MOCHA improves the inter-scan and inter-reader reproducibility of IVW image analysis over AI-MOCHA. In Aim 3, we will test the hypothesis that non-stenotic ICAD is more frequently detected by AI-MOCHA in the vascular territory of ESUS than in other territories using Domain Adaptive AI-MOCHA. To achieve this, we will scan 65 ESUS subjects each on the Philips, Siemens and GE 3T scanner platforms in a multi-center setting (three different hospitals) and demonstrate the utility of domain adaptive AI-MOCHA for robust and efficient IVW analysis. In doing so, we will not only establish the importance of non-stenotic ICAD in ESUS but also develop a clinically applicable IVW analysis pipeline for monitoring ICAD progression that will assist in optimizing medical therapies in individual patients. Further, the pipeline will provide a rapid, reliable tool for identifying patients who do not respond to conventional therapies for clinical trial participation.

NIH Research Projects · FY 2025 · 2022-09

Project Summary/Abstract The sensitive period for phonetic learning, which occurs between 6~12 months of age, is demonstrably one of the earliest milestones for language acquisition, as infants’ speech processing during this period can reliably predict later language skills. Most recently, my work has shown for the first time that neural processing of speech during this period can predict not only individual grammar skills at 6 years, but also the risk of developing speech and language disorders in typically developing infants. This result suggests an important clinical relevance of the sensitive period and demonstrates a crucial need to further understand the underlying neural mechanisms of the sensitive period, particularly whether it may already be altered in infants at high-risk of developing language disorders, such as the developmental language disorder (DLD). Elucidating the neural mechanism would not only allow for the expansion of theories to encompass atypical language development, but more importantly, help identify early markers for language disorders and aid in the development of targeted early interventions. Specifically, the sensitive period is characterized by a divergence in speech processing, whereby infants’ sensitivity for native speech contrasts improves while sensitivity for nonnative speech contrasts declines. It is theorized to be a specialization process for the native language, where less native language specialization is associated with slower language growth. However, behavioral and functional neuroimaging studies have only investigated typically developing infants (Low-Risk), but not infants with a family history of communication disorders (High-Risk). It is possible that differences already emerge during this period that separate High-Risk infants and these differences may underlie later language difficulties in the High-Risk population. The current Katz ESI proposal aims to shift the focus and specifically examine High-Risk infants with a family history of DLD and compare them with Low-Risk infants at ~6 (start), ~12 (end) and ~14 (delayed end of sensitive period) months of ages, with the central theoretical hypothesis that High-Risk infants would demonstrate a protracted sensitive period, or a lagged native language specialization than Low-Risk infants. Particularly, we examine both higher and lower levels of neural processes for speech, as well as in the link between the neural processes. The higher-level processes related to speech discrimination is indexed by the mismatch response (MMR) while lower-level sensory encoding of speech is indexed by the complex auditory brainstem response (cABR). Both measures will be obtained simultaneously with Magnetoencephalography (MEG). Together, the proposed study will help uncover neural mechanisms underlying atypical language development and perfectly aligns with the mission of NIDCD to conduct research in disordered processes of speech and language.

NIH Research Projects · FY 2024 · 2022-09

ABSTRACT Preventing HIV transmission with enhanced Pre-exposure Prophylaxis (PrEP) access, particularly among marginalized populations with disproportionate HIV incidence (e.g., minoritized racial/ethnic and sexual orientation and gender groups), is a crucial step in ending the HIV epidemic. Though unhealthy alcohol use is a central modifiable risk factor for HIV incidence and over-represented among marginalized populations, few HIV prevention interventions exist that synergistically address both unhealthy alcohol use and PrEP. Research is needed to understand PrEP uptake among individuals with unhealthy alcohol use and to develop scalable patient-centered interventions that synergistically address unhealthy alcohol use and HIV. The Veterans Health administration (VA) is a leader in provision of evidence-based care for unhealthy alcohol use, yet substantial gaps in PrEP implementation for Veterans with unhealthy alcohol use exist—our preliminary research suggests major gaps in PrEP knowledge, use, and reach, and Veterans with unhealthy alcohol use. Evidence-based interventions/treatments are available for unhealthy alcohol use, but alcohol use is historically under targeted in HIV prevention interventions. Our team of interdisciplinary experts in HIV and addiction medicine, the intersection of alcohol use and HIV, implementation science, health disparities research, and community-partnered research will use sequential mixed methods guided by socioecological theory, the Consolidated Framework for Implementation Science, and the Discover/Design/Build/Test (DDBT) process to understand the impact of unhealthy alcohol use on the PrEP care continuum and then refine an existing decision aid to increase PrEP initiation in primary care at the point of alcohol-related care in the VA. Quantitative work will include ~1.9 million individuals, including adequate numbers of minoritized patients (e.g., ~115,000 individuals with minoritized sexual orientation and ~11,000 individuals with minoritized gender identity) and will assess variation in patterns of PrEP initiation and persistence across VA facilities and patient subgroups (Aim 1). Findings will be used to inform purposive sampling for qualitative work to refine (Aim 2) and then pilot test an existing decision aid for developed previously with NIAAA support (Aim 3). Our study is patient-centered and innovative in characterizing PrEP care overall and within diverse subgroups of patients with and without unhealthy alcohol use, leveraging a novel natural language processing (NLP)--driven algorithm for identifying sexual minority groups, and tailoring an existing decision aid to facilitate shared decision-making for co-occurring HIV prevention and alcohol use. Study activities will be conducted with iterative input from a community advisory board. The study is highly responsive to NIAAA priorities and has potential for high impact as it will lay foundation for integrating a patient- centered and multi-targeted novel decision aid for HIV prevention in routine primary care settings that may have potential to increase equity in care.

NIH Research Projects · FY 2024 · 2022-09

Abstract Decades of research have revealed that the composition of the human gut microbiome directly impacts human health, yet we still do not understand the mechanisms underlying formation and maintenance of this complex community. Counterintuitively, increasing evidence suggests that competitive interactions play a role in maintaining community stability. While genes encoding components of diverse systems involved in interbacterial antagonism are widespread in human gut metagenomes, the specific antagonistic interactions occuring in situ remain elusive. One of the pathways mediating interbacterial antagonism is the Esx secretion system, which is conserved in two prominent gut phyla, Firmicutes and Actinobacteria. While this pathway has been shown to mediate contact-dependent interbacterial antagonism by a limited number of Gram-positive species in vitro, many questions remain about its physiological function. Antibacterial toxins secreted by the Esx machinery are present throughout human gut metagenomes, suggesting that Esx-mediated antagonism could occur in this community. These findings led me to propose to identify the targets of Esx-mediated interbacterial antagonism in a model natural community, the murine gut microbiome. To accomplish this goal, I will take advantage of the fact that bacteria express cognate immunity genes to prevent self-intoxication by secreted toxins. In Aim 1 of my proposed studies, I will use bioinformatics and in vitro functional assays to identify Esx toxin and immunity genes encoded in a model murine gut metagenome derived from wild-caught mice. My preliminary findings indicate at least two antibacterial Esx toxins are encoded in this natural gut microbiome. In Aim 2, I describe an in situ conjugation strategy to deliver the immunity genes identified in Aim 1 to all members of the murine gut community. I hypothesize that expression of immunity genes in bacteria targeted by specific Esx toxins will increase their abundance in a community. Therefore, by identifying changes to community composition that depend on immunity genes, I can determine the targets of Esx-mediated antagonism in situ. This unique approach to examine interbacterial antagonism under physiological conditions will significantly improve our understanding of the physiological function of the Esx system in gut bacterial species. It will also provide the first direct characterization of antagonism between gut microbiome constituents. The methods I propose to develop may additionally later be applied broadly to the characterization of diverse antagonistic pathways present in gut bacterial species, an essential next step needed to define the mechanisms shaping gut microbiome composition and ultimately human health. By working in the lab of Dr. Joseph Mougous at the University of Washington and with my experienced collaborators, I will be able to learn diverse techniques, practice effective mentoring strategies, and hone my scientific communication skills. This high-quality training will enable me to succeed as an independent researcher studying microbe-microbe interactions.

NIH Research Projects · FY 2025 · 2022-09

ABSTRACT Life-threatening traumatic exposures requiring presentation to acute care medical settings are endemic in the US in the era of the COVID-19 pandemic, firearm proliferation, and extreme weather events, and constitute both a substantial source of individual suffering and a significant public health burden. Each year in the US, over 30 million individuals present to acute care medical settings after injury, and approximately 2.5 million individuals are so severely injured that they require inpatient hospital admissions. The overarching goal of the Trauma Survivors Outcomes and Support (TSOS) R01 investigation is to advance the sustainable delivery of high quality trauma center mental health screening, intervention and referral procedures for diverse injury survivors. Over the past two decades, the TSOS study team that includes research scientists, trauma surgical policymakers, patients, and frontline clinicians has established a track record of using evidence derived from NIH pragmatic trials to directly target American College of Surgeons Committee on Trauma (College) regulatory policy. The TSOS R01 investigation will refine and test optimal stepped care intervention strategies for diverse injury survivors presenting to acute care medical settings with PTSD and associated comorbidity. This single trauma center site pragmatic trial investigation will individually randomize 424 patients (212 intervention and 212 control) to a brief stepped care intervention versus College required screening and referral control conditions. The stepped care intervention consists of proactive care management, as well as medications and psychotherapy elements targeting PTSD and comorbidity. Blinded follow-up interviews at 3-, 6-, and 12-months post-injury will assess the symptoms of PTSD and related comorbidity for all patients. The emergency department health information exchange will be used to capture population-level automated emergency department/inpatient utilization data for the intent-to-treat sample. The R01 aims to test the primary hypotheses that intervention patients will demonstrate significant reductions in PTSD symptoms and emergency department/inpatient utilization when compared to control patients. The investigation will also explore mediators and moderators of intervention treatment effects that directly address actionable national trauma center quality improvements. A mixed method Rapid Assessment Procedure-Informed Clinical Ethnography implementation process assessment will facilitate the integration of study results into national College policy requirements, guidelines, and verification criteria. A national trauma center survey will elucidate the progression of PTSD and comorbidity screening, intervention and referral for all US level I and II trauma centers. An end-of-study College policy summit will harness pragmatic trial data to inform the capacity for US trauma centers to implement high quality acute care medical mental health services for diverse patient populations.

NIH Research Projects · FY 2025 · 2022-09

Project Summary Cellular mechanisms mediating the transition from a state of general anesthesia to an awake state are not understood. Patients emerge from anesthesia passively without the use of mechanistically targeted interventions, creating an unpredictable clinical outcome marked by behavioral phenomena like emergence agitation and delirium. Many anesthetics act either directly or indirectly to change excitatory/inhibitory balance in the brain, highlighting the importance of understanding inhibitory networks in emergence. My research program focuses on the effects of general anesthesia on inhibitory plasticity during the transition from anesthetized to awake state in mice. My prior work examines neuroligin-2, a central organizer of the inhibitory synapse, using cell-type and circuit specific manipulations. Neuroligin-2 is a cell adhesion protein that acts as a scaffold to regulate general inhibitory synaptic function and is recently implicated as an independent regulator of intracellular signaling and disease. I demonstrated that neuroligin-2 manipulation modulates agitation and related behaviors in mouse models. My established expertise in stress-induced inhibitory synaptic plasticity provides a strong foundation for the following three complementary research areas investigating general anesthesia emergence. (1) Inhibitory cell plasticity in emergence from anesthesia: Research area 1 will focus on effects of general anesthesia emergence on inhibitory cell plasticity, starting with an investigation of key inhibitory postsynaptic genes, like neuroligin-2, using targeted knockdown and electrophysiology studies. Single cell sequencing transcriptomic investigations will characterize all cell types and synaptic constituents modified by emergence. (2) Brain-wide cell type-specific circuit activity in emergence: Research area 2 will investigate whole brain circuit changes induced by general anesthesia emergence using activity mapping and light sheet microscopy at single cell resolution, along with recordings of neuronal activity using genetically encoded optical sensors expressed in anesthesia-regulated circuits. (3) Preclinical models for emergence delirium across the lifespan: Research area 3 will develop and validate preclinical rodent models of emergence delirium using machine learning approaches, in order to study vulnerability to anesthesia-induced delirium across the lifespan. Together, these three projects form an overarching research program to understand mechanisms of emergence from the inhibitory cellular to circuit to behavioral levels of analysis, providing a holistic view of emergence. We must understand the mechanisms of anesthetic emergence across all levels in order to design pharmaceutical interventions to bring about safer and predictable emergence.

NIH Research Projects · FY 2025 · 2022-09

Unanticipated adverse drug reactions remain a major cause of post marketing withdrawals of drugs and of restricted access to new medications. Adverse drug reactions are often caused by reactive metabolites that form covalent adducts with proteins, but the identity and extent of protein adducts formed is often difficult to predict and characterize. Despite decades of research, current methods are unable to produce a comprehensive and quantitative catalog of xenobiotic protein adducts. This hinders progress in optimizing safety of novel medications and limits our ability to define mechanisms of clinically observed adverse drug reactions. To bridge this gap, we have developed innovative proteomic methods for discovery, characterization and quantification of protein adducts in simple and complex biological matrices. The goal of this proposal is to establish these methods for rapid, reliable and quantitative identification and characterization of drug-protein adducts, and uniquely customize these methods for human adductomics research impacting drug safety assessment. We will focus on covalent protein modifications resulting from metabolic oxidative activation of drugs. We will use a set of model compounds that are known to cause adverse events in patients and form reactive metabolites that likely result in protein adducts. In our aim 1 we will test the hypothesis that the modification masses and chemical characteristics of protein adducts formed by reactive metabolites in recombinant enzyme systems predict adduct formation in more complex systems such as liver microsomes and S9 fractions. Through this work we will optimize our proteomics methods for complex human liver preparations from individual donors. In aim 2 we will test the hypothesis that adduct formation varies quantitatively between individuals and due to differences in metabolic activity and individual genotype. In this aim we will establish quantitative adductomics for individual donors and define the basis for inter-individual variability in drug-protein adduct formation. In aim 3 we will test the hypothesis that human hepatocytes exposed to reactive metabolites generated in situ secrete proteins that have been adducted by the reactive intermediates. In this aim we will establish the in vitro relationship between hepatocyte adductomic burden and secretion of adducted proteins as biomarkers. When completed, the proposed studies will be transformative in integrating novel suite of cutting-edge tools and high-dimensional proteomics data to characterize the deep adductomic profiles of key drugs resulting in adverse drug reactions. The methods developed will enable the assessment and quantification of a broad range of adducts across the human proteome. The results will generate unprecedented insight into mechanisms of enzyme inactivation, liver adductomes formed after exposure to reactive metabolites and quantitative relationships between adduct formation and metabolic activity in the liver. The results may ultimately lead to novel drug discovery approaches that mitigate risk of adverse drug reactions resulting from adduct formation and to development of targeted therapeutic interventions designed to treat adverse drug reactions.

NIH Research Projects · FY 2024 · 2022-09

ABSTRACT Preclinical studies indicate that the neurobiology of obesity may be related to diet-induced cellular inflammatory responses in hypothalamic areas that regulate body weight. In rodent models of diet-induced obesity, overfeeding by high-fat diet rapidly induces inflammation and gliosis in the arcuate nucleus of the hypothalamus. Moreover, these glial cell inflammatory responses are both necessary and sufficient to induce hyperphagia and weight gain, suggesting that gliosis plays a mechanistic role in the ability of diet to induce obesity in rodent models. Using magnetic resonance imaging (MRI), the investigators discovered the first evidence of hypothalamic gliosis in humans with obesity. These findings have since been replicated in adults and children with obesity as well as in type 2 diabetes and insulin resistance. However, the current evidence in humans is observational, and the lack of controlled clinical experiments limits our understanding of whether findings of hypothalamic gliosis are diet-induced in humans. The overall goal of the proposed research is to determine the feasibility, promise, and safety of an experimental model in humans to test the effects of hypercaloric, obesogenic diets on hypothalamic gliosis as measured by MRI. The proposed mechanistic clinical trial is in response to PA-20-160 (Small R01s for Clinical Trials Targeting Diseases within the Mission of NIDDK). Using a randomized clinical trial design and a eucaloric control group, the proposal aims to compare the effect of two overfeeding regimens that vary in caloric load (but not macronutrient composition) on hypothalamic gliosis as measured by MRI. A second goal is to determine whether a standardized hypocaloric diet following the overfeeding regimen is a feasible strategy to mitigate any observed weight gain and/or hypothalamic gliosis due to overfeeding. Finally, the feasibility, acceptability, and long-term weight effects of study participation will be assessed. Forty-two participants with overweight will undergo serial MRI scans before, during, and at completion of 7 days of overfeeding or eucaloric diet as well as before, during, and at completion of a 7-day hypocaloric or eucaloric diet period. Diet composition during overfeeding will be based on preclinical studies of gliosis and utilize a robust stimulus that is high in dietary fat, saturated fat, and refined carbohydrates and most would consider obesogenic. Effect sizes, feasibility, and safety data will contribute to the design of future mechanistic clinical trials. Additional future directions include parsing which diet elements (e.g., caloric excess, saturated fatty acids) elicit radiologic evidence of inflammation and gliosis in the mediobasal hypothalamus. The proposed study represents a necessary first step in testing the hypothesis that a hypercaloric, obesogenic diet induces inflammation and structural changes in body-weight-regulating areas of the brain in humans—changes that promote the development of obesity and metabolic disease.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT The presence of lipid-rich deposits underneath the retinal pigment epithelium (RPE) is a pathologic feature of early age-related macular degeneration (AMD). Drusen development has been associated with RPE lipid metabolism, redox biology and extracellular matrix (ECM) degradation. Mutations in a gene affecting ECM degradation, tissue inhibitor of metalloproteinase 3 (TIMP3), results in a rare inherited retinal degeneration with similar clinical features to AMD, called Sorsby Fundus Dystrophy (SFD). The mechanism by which abnormal ECM turnover influences lipid metabolism and RPE redox resulting in the formation of sub-RPE deposits remains unknown. The goal of this proposal is to test the hypothesis that ECM degradation overloads the RPE with ECM-derived metabolites, resulting in the reprogramming of RPE towards lipid synthesis and mitochondrial oxidative metabolism. This in turn results in the deposition of excess lipids and reduced antioxidative capacity of the RPE. The proposed specific aims are: Aim 1. Determine the influence of ECM degradation on lipid metabolism. Our preliminary results show that increased ECM degradation in SFD RPE activates lipid synthesis and oxidation of branch-chain amino acids (BCAAs). BCAAs are ketogenic and abundant in the ECM. The goal of Aim 1 is to test the hypothesis that ECM degradation of protein-rich components reprograms RPE metabolism towards enhanced BCAA oxidation for lipid synthesis and lipid deposition. We will use quantitative proteomics, quantitative metabolomics, metabolic flux analysis, perifusion assays, CRISPR/Cas9 gene-editing, and single cell nuclear RNA-Seq of patient-derived iPSC RPE to comprehensively investigate the metabolic pathways in ECM remodeling and lipid deposition. Aim 2. Determine the influence of ECM degradation on redox metabolism. Our preliminary data show that both NADPH and glutathione are depleted in SFD RPE, and ECM-derived metabolites interfere with NADPH and glutathione metabolism. The goal of Aim 2 is to test the hypothesis that increased ECM turnover results in impaired NADPH and glutathione metabolism. We will quantify the metabolic flux of ECM degradation, determine the roles of ECM-related metabolites in NADPH production and glutathione synthesis, and restore cellular redox with different approaches to enhance antioxidative capacity. The proposed research will define the biochemical impacts of ECM turnover on RPE metabolism, including changes in lipid metabolism and oxidative stress, and identify the relationship between nutrient metabolism, protein synthesis and degradation, and redox biology in normal and disease-relevant RPE.

NIH Research Projects · FY 2025 · 2022-09

Project Summary Progress in improving infection outcomes depends upon understanding factors that diminish antibiotic action. Recent work suggests antibiotic heteroresistance may be a key factor. Heteroresistant pathogens exhibit population-level resistance heterogeneity, meaning they contain highly resistant subpopulations, and often exhibit unstable resistance, meaning some cells become rapidly sensitive when grown without antibiotics. This proposal focuses on these two phenotypes as they are postulated to have major effects on susceptibility testing and antibiotic efficacy. Here we propose to study population-level resistance heterogeneity and unstable resistance in people with cystic fibrosis (CF) and chronic Pseudomonas aeruginosa (Pa) infections who cycle on and off inhaled tobramycin. CF offers key advantages for these studies as it is a monogenic disease with uniform manifestations, the responses of a single bacterial species to the same antibiotic can be studied in sizable cohorts, and infrastructure exists to procure primary specimens from the infected site (sputum) during antibiotic "on" and "off" periods. These advantages, and methods developed in our preliminary work enable us to test the hypotheses that Pa from chronically-infected patients exhibit marked population-level resistance heterogeneity and unstable resistance; that parameters accounting for these factors predict clinical responses to treatment; and that unstable resistance in CF Pa is due to a core set of gene amplifications and fitness-compensating point mutations. This work will generate foundational knowledge about antibiotic heteroresistance that could improve understanding of antibiotic treatment failure in CF, and suggest new precision medicine approaches to select antibiotics and improve outcomes. The findings could also guide future work in infections where patient heterogeneity, pathogen and treatment diversity, and less developed infrastructure make studies more difficult.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT Patients with Alzheimer’s disease and related dementias (ADRD) often experience high-intensity, potentially burdensome care near the end-of-life that may not align with their values and goals. Patients with ADRD and their families often face complex decisions about the most appropriate intensity of care. Goals-of- care discussions with clinicians are associated with improved patient and family outcomes and reduced intensity of care at the end of life. Although goals-of-care discussions can facilitate delivery of goal-concordant care, they currently occur infrequently or too late in the illness. Electronic health records (EHR) provide a key opportunity to identify patients who would benefit from these discussions and to promote these discussions, yet few prior interventions have used the EHR for this purpose. To address this gap, we propose an NIH Stage III hybrid efficacy-effectiveness trial of a communication-priming intervention, called Jumpstart, to increase the occurrence and quality of goals-of-care discussions with patients with ADRD and their family caregivers in the outpatient setting. We will use EHR data to identify patients and facilitate implementation of the intervention. In Aim 1, we will conduct a NIH Stage III hybrid efficacy-effectiveness trial of the adapted Jumpstart intervention compared to usual care to assess effectiveness for promoting occurrence and improving quality of goals-of care discussions and improving patient- and family-reported outcomes. The primary outcome is EHR documentation of a goals-of-care discussion by 3 months after enrollment. We will enroll 1200 patients with ADRD and their family caregivers. Aim 2 examines the mechanisms of action of the intervention on important patient- and family-centered outcomes including patients’ palliative care needs and the intervention’s effect on healthcare utilization, and monitors for unintended consequences of the intervention in terms of psychological distress and one-year all-cause mortality. In Aim 3, we will conduct a mixed-methods evaluation to explore further intervention improvements and identify barriers and facilitators to intervention implementation. Our team is well-positioned to accomplish these aims. We have experience conducting large, randomized trials with patient- and family-centered outcomes, biostatistics, qualitative and mixed methods analyses, ADRD research and policy, and implementation science methods. We have used informatics integrated with the EHR to identify patients with ADRD and measure quality of care similar to the methods proposed here, including measurement of the primary outcome. This study will provide important data for patients with ADRD, their family caregivers, clinicians, and healthcare systems on the efficacy of the Jumpstart intervention to improve communication and facilitate goal-concordant care and will provide insights for implementation of similar EHR-based interventions designed to improve communication and patient outcomes.

NIH Research Projects · FY 2024 · 2022-08

Title: Towards Single-molecule Protein Sequencing with Nanopores Abstract: This proposal concerns the development of a nascent single-molecule proteomics technology based on nanopore sequencing. In recent work by Brinkerhoff et al., we demonstrated a first-of- kind method for sequentially reading single peptide molecules with a biological nanopore, detecting single-amino acid substitutions in individual molecules and showing that extremely high fidelity may be reached through indefinitely many independent re-reads of individual proteins. This method does not require chemical labeling of amino acids, can be carried out on commercially available nanopore sequencing hardware, and retains the properties that enabled the success of nanopore DNA sequencing: low overhead cost, physical rather than chemical sensitivity to small changes in single molecules, and the flexibility to be re-engineered to target specific sequencing applications. In this proposal to fund a collaboration of the Gundlach and Dekker laboratories, both world leaders in nanopore technology development, we aim to develop this promising new technology into a fully fledged single-molecule proteomics tool. This will involve characterizing reads of a broad array of protein sequences, optimising core elements of the sequencing methodology for protein analysis, preparing natural peptides for reading, and determining the method's applicability to pressing proteomics problems such as post-translational modification and splice variant detection. Achieving these aims will position the nanopore protein reader for immediate application to proteomics problems in fundamental biology and clinical medicine.

NIH Research Projects · FY 2025 · 2022-08

PROJECT SUMMARY Throughout evolution and history humans have progressively isolated themselves from the natural cycles by building artificial habitats that shield them from the external environment. This has vastly affected when and how much humans sleep, especially since the near universalization of electric light. The key to this isolation and change in sleep patterns is the ability to manipulate artificial light and extend activity into the nighttime. A recent study from our laboratory found that the timing of sleep changes across the moon cycle, with later and shorter events of night sleep on the nights leading up to the full moon. Surprisingly, we found this lunar cycle modulation not only in rural but also in urban environments. Sleep is controlled by two simultaneous processes: the circadian clock determines the optimal times for sleep throughout the 24-h cycle while the homeostatic drive for sleep increases as we stay awake. The two processes combine to determine the natural human patterns of night sleep. This project aims to determine which of these two mechanisms that regulate sleep is affected throughout the moon cycle to delay and shorten sleep on the nights before full moon. We will combine longitudinal field recordings of sleep and sleep laboratory strategies to address the following two aims: Specific Aim 1: Assess circadian phase at opposite phases of the moon cycle in Toba/Qom communities. We will conduct longitudinal monitoring of sleep in rural and urban native Toba/Qom communities in northern Argentina. In individuals with robust lunar patterns we will assess the phase of the dim-light melatonin onset (DLMO), the gold- standard for determining circadian phase in humans, in the days leading to the full and new moons. Our hypothesis is that the phase of the circadian clock is modulated across the moon cycle, and predicts that the DLMO phase will be delayed on the days preceding the full moon relative to the pre-new moon days. Specific Aim 2: Assess circadian phase, and circadian and homeostatic regulation of sleep at opposite phases of the moon cycle in a highly urbanized community. We have already confirmed that this lunar modulation is present even in industrialized societies, where artificial light during the evening reigns over the full moon’s light. We will screen Seattle participants through longitudinal sleep monitoring and select those with the most robust lunar rhythms to participate in sleep laboratory visits twice at the opposite phases of the moon cycle. We will use polysomnographic wake and sleep recordings, as well as a constant routine protocol to study the potential lunar monthly change in circadian phase and in the homeostatic regulation of sleep after sleep deprivation. Whether the moon can affect sleep has been a matter of high controversy for decades. Our recent work demonstrated that sleep patterns vary across the moon cycle in a very predictable manner in real life conditions, in very different populations and under different environments. This project aims to shed light on the mechanisms through which this modulation occurs, which will further our understanding of the regulation of human sleep, and its impact on health and disease.

NIH Research Projects · FY 2024 · 2022-08

ABSTRACT Maximizing access and minimizing costs of delivery are key challenges for optimizing the public health impact of HIV pre-exposure prophylaxis (PrEP). Between January 2017 and December 2019, as part of Kenya's national public sector PrEP roll-out, we conducted a stepped-wedge cluster-randomized pragmatic trial to catalyze scale- up of PrEP delivery integrated in 25 public HIV clinics (The Partners Scale Up Project). We demonstrated that PrEP can be delivered in African public health facilities using existing staff: >8000 initiated PrEP (53% women) with reasonable continuation and high adherence among those returning. The study also highlighted major health system barriers including lengthy visits with multiple stops (i.e., separate rooms for triage, HIV testing, counseling, pharmacy) that burden the health system. For healthy HIV uninfected persons, long waiting at the clinic, time away from work, and costs for getting to visits challenge persons taking PrEP. Efficient delivery strategies could reduce costs, potentially improve client engagement and allow services to be available to a larger number of people. In a short-term pilot study jointly funded by NIH and PEPFAR, we tested the feasibility of one-stop PrEP provision (i.e., all PrEP services provided in a single room) at PEPFAR-supported clinics and showed that one-stop service was feasible and highly acceptable to both PrEP users and providers. One-stop significantly shortened wait time (>80%) without reducing provider-client contact time; PrEP initiation stayed stable and there was suggestion of better early continuation and on-time visit attendance, indicating that one- stop PrEP might add efficiencies to PrEP systems, without undermining quality. Building on our learning from the randomized trial and the pilot study, we propose to conduct an effectiveness-implementation cluster- randomized trial of one-stop to assess effectiveness to improve delivery efficiency and continuation on PrEP and to rigorously study health system factors. We will randomize 12 public health facilities with established PrEP programs in Western Kenya 1:1 to recruit and follow 1800 HIV-uninfected persons newly initiating PrEP to test the effectiveness of one-stop care pathway compared to usual care pathway (Aim 1). Within the large program, we will establish a randomly selected nested observational cohort of PrEP users (n=150, including clients who start but discontinue) to study longitudinal HIV prevention behavior, including reasons for discontinuation and how clients align PrEP use with HIV risk. Co-primary outcomes will be continuation and adherence quantified by tenofovir-diphosphate levels in dried blood spots. We will use the Systems Analysis and Proctor's implementation framework to evaluate implementation outcomes at the health system, facility, provider, and client-level (Aim 2). Finally, we will conduct micro costing and time and motion studies to evaluate the costs and model the budget impact and affordability of PrEP program with One-stop model (Aim 3). We have already demonstrated that PrEP can be delivered in Kenyan public facilities by existing staff; this work will extend further, aiming for greater efficiency and cost-saving, reduced provider workload, diminished client burden, and better PrEP continuation.

NIH Research Projects · FY 2025 · 2022-08

ABSTRACT Maximizing access and minimizing costs of delivery are key challenges for optimizing the public health impact of HIV pre-exposure prophylaxis (PrEP). Between January 2017 and December 2019, as part of Kenya's national public sector PrEP roll-out, we conducted a stepped-wedge cluster-randomized pragmatic trial to catalyze scale- up of PrEP delivery integrated in 25 public HIV clinics (The Partners Scale Up Project). We demonstrated that PrEP can be delivered in African public health facilities using existing staff: >8000 initiated PrEP (53% women) with reasonable continuation and high adherence among those returning. The study also highlighted major health system barriers including lengthy visits with multiple stops (i.e., separate rooms for triage, HIV testing, counseling, pharmacy) that burden the health system. For healthy HIV uninfected persons, long waiting at the clinic, time away from work, and costs for getting to visits challenge persons taking PrEP. Efficient delivery strategies could reduce costs, potentially improve client engagement and allow services to be available to a larger number of people. In a short-term pilot study jointly funded by NIH and PEPFAR, we tested the feasibility of one-stop PrEP provision (i.e., all PrEP services provided in a single room) at PEPFAR-supported clinics and showed that one-stop service was feasible and highly acceptable to both PrEP users and providers. One-stop significantly shortened wait time (>80%) without reducing provider-client contact time; PrEP initiation stayed stable and there was suggestion of better early continuation and on-time visit attendance, indicating that one- stop PrEP might add efficiencies to PrEP systems, without undermining quality. Building on our learning from the randomized trial and the pilot study, we propose to conduct an effectiveness-implementation cluster- randomized trial of one-stop to assess effectiveness to improve delivery efficiency and continuation on PrEP and to rigorously study health system factors. We will randomize 12 public health facilities with established PrEP programs in Western Kenya 1:1 to recruit and follow 1800 HIV-uninfected persons newly initiating PrEP to test the effectiveness of one-stop care pathway compared to usual care pathway (Aim 1). Within the large program, we will establish a randomly selected nested observational cohort of PrEP users (n=150, including clients who start but discontinue) to study longitudinal HIV prevention behavior, including reasons for discontinuation and how clients align PrEP use with HIV risk. Co-primary outcomes will be continuation and adherence quantified by tenofovir-diphosphate levels in dried blood spots. We will use the Systems Analysis and Proctor's implementation framework to evaluate implementation outcomes at the health system, facility, provider, and client-level (Aim 2). Finally, we will conduct micro costing and time and motion studies to evaluate the costs and model the budget impact and affordability of PrEP program with One-stop model (Aim 3). We have already demonstrated that PrEP can be delivered in Kenyan public facilities by existing staff; this work will extend further, aiming for greater efficiency and cost-saving, reduced provider workload, diminished client burden, and better PrEP continuation.

NIH Research Projects · FY 2025 · 2022-08

Motor neurons connect to muscles and comprise the major output of the nervous system. Patterns of neural activity in motor neurons cause temporally precise muscle contractions, producing coordinated and flexible behavior. These patterns are shaped by the connectivity and physiology of premotor circuits in the spinal cord that synapse onto the motor neurons. Premotor circuits combine descending motor commands with sensory feedback signals to drive motor neuron activity. How premotor networks are structured to control motor output is not well understood, due in part to an incomplete inventory of spinal cell types, and to the difficulties of recording neural activity in behaving animals. To address this gap, this project aims to use Drosophila melanogaster as a model for investigating motor control and premotor neural circuits. With an accessible and numerically compact nervous system, a large and growing suite of genetic tools, and agile, limbed locomotion, Drosophila has the potential to provide insight into fundamental problems of motor control. We introduce a task in which flies learn to generate specific amounts of force to position the femur-tibia joint in different targets. The joint is controlled by twelve neurons which can be genetically labeled for targeted neural recordings. Electrophysiological recordings will reveal how the complete population of motor neurons function together to dynamically position the leg and to sustain a given force output. These data will address long-standing hypotheses about how premotor circuits recruit subsets of motor neurons and the degree to which that control is flexible. Then, a new electron-microscopy level reconstruction of central locomotor circuits will allow identification of key premotor neurons. Electrophysiological recordings of those premotor neurons during the behavioral task will reveal their contributions to processing sensory feedback and to controlling leg force. These results will provide a foundation for understanding how descending commands interact with internal models of body state to control locomotion, a critical step toward achieving the long-term goals of designing interventions for neuromuscular disorders and algorithms for controlling engineered systems.