Washington University

NIH Research Projects · FY 2024 · 2017-08

Project Summary/Abstract This multi-PI basic science project aims to transform understanding of G-protein function in physiology and disease, and provide broadly applicable routes for developing therapeutics to treat currently untreatable diseases caused by mutant constitutively active G protein α-subunits. It does so by developing knowledge required to design and synthesize, at scales required for preclinical and eventual clinical studies, a family of bioavailable inhibitors, each of which selectively targets closely related groups of heterotrimeric G proteins. The translational/clinical potential of this approach is based on recent studies indicating that a bioavailable inhibitor that targets G protein α-subunits of the Gq/11 class is therapeutically effective in mouse models of uveal melanoma, an untreatable disease that is driven by mutant constitutively active Gq/11. Similar approaches could have broad impact, because many other untreatable diseases are driven by various types of mutant constitutively active G protein α-subunits, including hormone-secreting pituitary tumors, mucosal melanoma, choroidal hemangiomas, hepatic small-vessel neoplasms, ~10-15% of all cancers, Sturge-Weber syndrome, autosomal dominant hypoparathyroidism, and certain forms of hyper- and hypocalcemia. Moreover, this approach could be used to modulate G-protein activity in diseases where GPCR-targeted drugs are ineffective due to receptor redundancy or to G proteins that cause dose-limiting side effects such as respiratory suppression or drug tolerance. Bioavailable inhibitors that directly target specific subclasses of G proteins would be extremely valuable for basic science. They would provide simple, fast, cheap and reliable chemical probes with which to identify novel functions of G proteins in normal physiology and in animal models of disease, in contrast to conventional knockout or knockdown strategies, which are slow and expensive, and can suffer from compensatory or off- target effects. The foundation of this project is a pair of nearly identical, bioavailable, cyclic depsipeptide natural products that potently and selectively inhibit the Gq/11 subfamily of G protein α-subunits. The Specific Aims of this project will address four crucial challenges: 1) limited availability of these inhibitors; 2) lack of inhibitor derivatives that could be targeted to disease tissues for chronic therapy; 3) limited understanding of the inhibitory mechanism, which has precluded design of inhibitors that target other subtypes of G proteins; and 4) absence of inhibitors that selectively target G protein subtypes other than Gq/11. These Aims will be pursued by using innovative synthetic organic chemistry, computational-based docking and inhibitor design, single- molecule FRET, NMR, and biochemical and cell-based assays of G protein function.

NIH Research Projects · FY 2026 · 2017-07

Project Summary Noise exposure damages synaptic connections between cochlear inner hair cells and innervating auditory nerves. Data from mammalian models and humans indicate that loss of some inner hair cell synapses can be permanent, leading to the slow degeneration of detached auditory nerves. Yet recent research also supports that the mammalian cochlea possesses the intrinsic capacity for hair cell synaptic repair following noise damage. Defining the cellular mechanisms of synapse repair following traumatic noise is a critical step toward identifying therapeutic targets to promote repair of hair cell synaptic contacts and prevent loss of auditory nerves. The overall goal of this proposal is to understand the molecular basis of morphological and functional hair cell organ repair and recovery following noise-induced damage. Current gaps in our understanding of how hair cell synapses repair following traumatic noise are in large part due to our inability to define the cellular processes that promote synaptic repair in mammalian model systems. This project will circumvent these issues by investigating mechecaniclly induced hair-cell synapse loss and subsequent repair in the zebrafish lateral line— a mechanosensory organ which is made up of clusters of innervated hair cells. Zebrafish lateral-line hair cells are comparable to mammalian hair cells at the molecular and cellular level, including a shared mechanism of hair cell synapse loss and de-innervation following traumatic overstimulation. Yet lateral line hair cells rapidly and unambiguously repair lost synaptic connections within hours following stimulus-induced damage. Aim 1 of our proposal will test the hypothesis that hair cell activity governs synaptic repair, while Aim 2 will define the contribution of inflammation to synaptic recovery and reinnervation. The results of each of our Aims will provide information on how hair cell synaptic connections are restored following traumatic overstimulation and will help identify strategies to promote endogenous repair in noise exposed cochlea, thereby preventing subsequent auditory nerve degeneration and hearing loss.

NIH Research Projects · FY 2026 · 2017-07

PROJECT SUMMARY Human pluripotent stem cells (hPSCs) are a promising renewable source of differentiated insulin-producing islets for diabetes cell replacement therapy. Within the islets of diabetic patients, beta cells are dead or dys- functional, causing loss of blood glucose control. There is no cure for diabetes, and current treatments are insufficient in controlling the disease for many patients. Transplantation of insulin-secreting cells could be an effective treatment for diabetes, and a small number of patients have been implanted with cadaveric donor islets, remaining insulin independent for years. Unfortunately, many factors limit this approach, particularly the scarcity and variability of isolated human islets, with patients often requiring islets from multiple donors to achieve normal blood sugar. The lack of mature replacement tissues is a critical barrier to cellular therapy. We have previously published a strategy for the scalable generation of hPSC-derived islets (SC-islets) in vitro that are capable of secreting insulin and restoring normoglycemia in diabetic mice. However, a current challenge with these in vitro-derived cells is the incomplete and uncontrolled differentiation to fully mature SC-islets that are equal to primary cells. Our goal is to understand how modulating the microenvironment affects hPSC cell fate decisions to and the subsequent maturation of SC-islets and to use this to innovate SC-islet generation strategies for use in cellular therapy. To this end, we will study patterning of definitive endoderm and other intermediate cell types produced by microenvironmental perturbation. Subsequent generation of the SC-islets will be investigated, including characterizing in vitro and in vivo function and marker expression. The outcome of this proposal will have a positive impact by filling gaps in our understanding of what controls SC-islet cell fate specification to individual cell types, as well as how these cells mature. Successful completion of our studies will inform strategies to improve SC-islet differentiation protocols for diabetes cell replacement therapy and disease modeling.

NIH Research Projects · FY 2025 · 2017-07

PROJECT SUMMARY In my first K24 award, I mentored a total of 32 mentees, working with them on 87 papers in the last six years and shepherding 19 successful NIH K-series career development awards (with 6 as primary mentor). In the next five years, I seek to renew my K24 award to maintain protected time for mentoring and advance three new and exciting perspectives in mentoring. First, is to evolve from an “apprenticeship” model of mentoring to a “co-pilot” model based on a therapeutic alliance (from the 2020 National Academies of Sciences report) using an “Appreciative Inquiry” approach — a strengths-based approach well-known in organizational psychology but new to mentoring as well as set the pathway for their eventual transition to mentors themselves. To do so, I will seek to use a standard approach for soliciting how mentees see their successes and strengths as a basis for a shared objective in the mentoring relationship. Second, I have come to look beyond the dyadic relationship to see mentoring as creating a professional ecosystem in which a mentee can thrive. I therefore propose to foster peer-to-peer and peer-to-other mentor activities, including an annual retreat to solidify peer relationships between mentees. Third, I see success in science more explicitly as translational impact rather than solely publications and grants, and will use the K24 award to support training for mentees on the Translational Science Benefits Model — a novel framework for valuing scientific outputs — and communications skills. For my research enterprise, I propose to use the K24 renewal to make modest but critical extensions to my existing research program advancing person- centered public health. Person-centered in public health context does not imply that every individual’s preference can be accommodated but rather takes three feasible forms. First, personalization implies that public health services can detect and respond to effects on individuals. To carry that forward, I propose learning more about adaptive behavioral interventions to strengthen engagement in HIV care. Second, personalization also implies that the interpersonal domains of public health services are important, and patients — even in a public health setting — when treated with respect, have better engagement and outcomes. Finally, person-centered services in the context of today’s HIV epidemic implies the growing integration of non- communicable diseases in HIV programs — a key domain of person-centered services and an area of policy and scientific interest. The new research proposed in this K24 represent incremental scientific steps nested within a larger, well-funded research infrastructure. These analyses will yield preliminary data and hypothesis, generating findings that have value to new mentees and can yield accessible preliminary data for additional proposals. At the end of this next cycle, I hope to have nurtured a crop of leading-edge, implementation- oriented researchers focused on addressing the HIV/AIDS pandemic, both here in the US and globally. All of my work is embedded within my institutional leadership roles and is supported by institutional leadership.

NIH Research Projects · FY 2026 · 2017-07

PROJECT SUMMARY/ABSTRACT Role of glial circadian clock dysfunction in the pathogenesis of Alzheimer’s Disease Chronic disruptions of the circadian system, manifesting as sleep disturbances, day-night confusion, and “sundowning”, are well-described and debilitating symptoms of Alzheimer’s Disease (AD). While circadian disruption has long been considered a consequence of the degenerative process in AD, accumulating human and mouse data suggest that circadian rhythm abnormalities may begin before overt cognitive symptoms, and could play an important contributory role in AD pathogenesis. Circadian rhythms are generated in cells by specific clock genes, which are expressed in neurons and glia throughout the brain and control 24-hour oscillations in transcription. These cellular clocks are synchronized to the external environment by the central clock in the suprachiasmatic nucleus in the brain. Cellular circadian clocks are particularly robust in glial cells, regulating cellular activation and inflammatory responses in both astrocytes and microglia. We have found that the circadian clock protein BMAL1 regulates astrocyte activation, neuroinflammation, and amyloid plaque deposition in mice. We have also found that amyloid plaques cause large-scale alterations in circadian transcriptional rhythms in astrocytes. Thus, we will address the bidirectional relationship between circadian clock disruption and AD-related pathology in mouse models of AD, focusing on how the central and cellular clocks regulate astrocyte responses to protein aggregation. Using novel methods to interrogate cell type- specific transcription in vivo, we will compare the effects of central vs. cellular clock disruption on circadian function in astrocytes, both in healthy brain and in a model of AD. We then evaluate the effects of central and cellular clock disruption on pathology caused by Aβ and tau, and determine specific clock-regulated pathways in astrocytes that control protein degradation and inflammation. By understanding the bidirectional relationship between circadian rhythms and astrocyte function, we hope to identify novel therapeutic targets to prevent protein aggregation and inflammation in Alzheimer’s Disease.

NIH Research Projects · FY 2026 · 2017-06

I. Institutional Career Development Core (KL2), 7. Project Summary/Abstract: The goal of the Washington University (WU) KL2 Career Development Program is to provide team-oriented, competency-based, multidisciplinary mentored research training, didactic coursework, and professional development for junior faculty. Our Clinical Research Training Center (CRTC) provides educational resources and infrastructure for the WU Institute of Clinical and Translational Science (ICTS) and partner institutions. We are requesting 9 KL2 slots to support junior faculty for 2-3 years. To enhance our successful KL2, we will develop additional innovative coursework, training and career development programs across the full spectrum of T1-T4 research, emphasizing informatics and data science, and implementation for impact. To address the regional and national CTSA goals, we propose the following specific aims: 1: Enhance the CRTC and KL2 infrastructure and programs to provide personalized, competency based, rigorous, high-impact clinical/translational research training and skills. We will develop and provide new curricula and training programs emphasizing implementation science for impact. We will enhance our hybrid and online courses to improve access and flexibility for trainees and share materials with ICTS partners and other CTSA hubs. (CTSA Goal: Translational Workforce Development) 2: Provide high-quality, informatics training. We will enhance our informatics and data science training(workshops, seminars, courses, certificates, and degrees), and integrate new informatics methods, tools and skills into our courses and training programs to meet individual scholar needs. (CTSA Goal: Informatics) 3: Broaden the range of research disciplines represented among investigators leading high-impact, multidisciplinary clinical and translational research teams. We will train scholars, faculty and mentors from multiple different disciplines. To expand high-impact research in a broad range of communities and populations across the lifespan, we will integrate new team-science, mentoring and leadership development into our courses and career development programs (CTSA Goals: Integrating Populations across the Lifespan) 4: Increase collaborations between the faculty, mentors, and scholars in the KL2 program and stakeholders, ICTS program functions, partners, and CTSA hubs. We will create new resources, share best practices, and advance interdisciplinary scientific teams with a broad range of skillsets to foster patient and community engaged research and training at local, regional, and national levels. (CTSA Goal: Collaboration and Engagement) 5: Demonstrate the impact of the KL2 program. We will assess the short- and long-term performance of scholars, determine the efficacy of training methods, and use this data to improve training, trainee and faculty satisfaction, mentoring, leadership and outcomes. (CTSA Goal: Methods and Processes)

NIH Research Projects · FY 2026 · 2017-06

Overall Component, 7. Project Summary/Abstract: The Institute of Clinical and Translational Sciences aligns with the CTSA goals of developing innovative solutions to improve the efficiency, quality, and impact of translational science. We will pursue five aims: Aim 1: Advance interdisciplinary programs to develop, promote, mentor, and retain highly qualified faculty, trainees, and staff who translate scientific discoveries into action, thereby becoming agents for change in their institutions and communities. We will enhance our existing research education and training programs, expand our workforce, and promote team science approaches to conduct and translate high-impact research that has the potential for changing the health of our community. (Translational Workforce Development). Aim 2: Facilitate research designed for implementation by engaging communities and stakeholders in multidisciplinary collaborative teams at all stages of the translational research process, thereby demonstrating the benefit and impact of translational science. We will improve community health by collaborating with patient advocates, developing new partnerships, advancing the science of collaboration, and capitalizing on strengths in implementation science (Collaboration and Engagement). Aim 3: Integrate research across individual lifespans and apply translational science in multiple populations to improve individual and community health through meaningful research collaborations and sustainable partnerships. We will address health needs across the lifespan, identify challenges that affect various populations, catalyze the formation of transdisciplinary teams, and evaluate the outcomes of our work (Integration). Aim 4: Drive innovation, quality, and efficiency in translational research to enhance collaborations and catalyze the implementation of discovery science. We will solve key logistical roadblocks in translational science and implement innovative methods to engage stakeholders and research participants, thereby targeting different populations, life stages, and health states (Methods and Processes). Aim 5: Apply innovative informatics and biostatistics solutions to improve quality and efficiency at every stage of translational research and create an ecosystem that integrates various types of data and facilitates the interoperability, use, and reuse of digital assets. We will ensure the interoperability of current and archived data sets, implement new informatics technologies to enhance investigator productivity, and promote the formation of interdisciplinary research teams (Informatics). Impact. Successful completion of these five aims will transform translational research, accelerate the dissemination of new innovations across our entire population, and strengthen our workforce.

NIH Research Projects · FY 2026 · 2017-06

Washington University has a robust research infrastructure, outstanding faculty mentors, well-developed clinical and translational research (CTR) training programs, and a large pool of excellent pre- and postdoctoral candidates. The objective of the TL1 program is to ignite an interest in becoming a CTR scientist and provide research experience, coursework, and career development opportunities to achieve this goal. We have successfully trained predoctoral MD, PhD and allied health doctoral students, as well as postdoctoral trainees interested in entrepreneurship, technology transfer, and intellectual property development. We aim to expand our program to add support for a mentored career development pathway for MD clinical fellows. We will introduce new innovative curricula in biomedical informatics, dissemination and implementation science, stakeholder collaboration, team science, ensuring proportionate access to health resources, and online initiatives. Simultaneously, we will develop new mentor training programs coordinated by the Clinical Research Training Center.

NIH Research Projects · FY 2025 · 2017-05

Substance use disorder is multifactorial with genetic and environmental contributions and this illness is a leading cause of death worldwide. Approaches from single disciplines have done much to advance science, and methodological and technological advances have established the need for increasingly multidisciplinary approaches to address complex questions about substance use disorder. Bringing this potential of scientific discovery to advance the prevention and treatment of substance use disorder requires clinician scientists who can bridge the gaps between scientific discovery and clinical care. The goal of the K12 program is to train the next generation of clinician scientists to fill these gaps and undertake this important and exciting work. To accomplish this goal, we have established three aims: Expand the workforce of clinician scientists in the prevention and treatment of substance use disorder. We will fund two scholar faculty positions per year at Washington University School of Medicine. Using a scholar-centric approach, we will design a career development plan that will meet each scholar’s needs and interests. We will provide short-term and long-term mentorship through formal mentoring relationships with local and national faculty and advisors from multiple disciplines. New opportunities to aid the scholars include wellness and resilience support, career coaching, and guidance for work/life balance. We are committed to the training and retention of scholars from multiple disciplines. Continue to develop and enhance the K12 career development opportunities and continue robust evaluation and tracking of K12 scholars, faculty, mentors, curriculum, and training opportunities. We will enhance our current program by implementing online individual career development plans that can be monitored by scholars, mentors, and Program Directors. A Research Navigator will increase support of the scholars in managing and conducting their research projects. We will monitor and evaluate the scholars, mentors, faculty and training programs using surveys, interviews, and other metrics. Establish a national network of NIDA K12 Program Directors and scholars. This national consortium will serve as an avenue of communication between Program Directors and scholars across all NIDA K12 programs, creating a community with common goals, with the intent to share resources, enhance networking, and foster the development of the next generation of researchers in substance use disorder. Successful completion of these aims will result in increased numbers of multi-disciplinary, well- trained clinician scientists in the prevention and treatment of substance use disorder and its consequences.

NIH Research Projects · FY 2025 · 2017-05

ABSTRACT To enable efficient specialization and dynamic regulation of subcellular regions, many cells have evolved local translation of mRNA - yet the fundamental principles of such translation regulation in astrocytes are unknown. Long studied in neurons, local translation of a variety of proteins is thought to be essential for the synapse- specific changes that underlay learning and memory. In the prior cycle of this award we provided evidence astrocytes also have a regulated local translation by using a variety of approaches. Here, we propose to continue this work, focusing on the following question regarding this fascinating new basic biological phenomenon: what is the regulatory grammar that determines when and which transcripts are locally translated in astrocytes? Our central model is that elements in the untranslated regions (UTRs) of transcripts are responsible for their enrichment or depletion from ribosomes in peripheral astrocyte processes (PAPs), via UTR interactions with RNA binding proteins (RBPs) and microRNAs (miRNAs). However, with hundreds of potential elements to screen, new, scalable methods are needed to systematically characterize how RNA localization and translation is regulated in astrocytes, both at baseline and in response to signaling cues. Furthermore, glial morphology only reaches full maturity in vivo, requiring in vivo functional studies. Therefore, we have developed a new method, a synaptoneurosome–massively parallel reporter assay (SN-MPRA) which allows us to assess functional effects of thousands of candidate UTR elements in vivo simultaneously. We will apply this to define the sequences that modulate RNA localization in astrocytes. Furthermore, to better understand how a subset of these elements function, we will define the role of a specific RBP, ‘quaking’ (QKI), in modulating local translation in astrocytes. Finally, to understand how sets of transcripts might be regulated in a coordinated fashion for local translation, we will examine the role of miRNA effector proteins (Ago2) along with specific miRNAs in regulating local translation in astrocytes.

NIH Research Projects · FY 2026 · 2017-04

Project Summary Nervous system functions, including vision, arises from precise patterns of synaptic communication. In neurodegenerative diseases, synapse loss can long precede cell death and predict functional impairments. We recently discovered that the cell adhesion molecule (CAM) Netrin-G ligand 2 (NGL2) is required for the maintenance of rod photoreceptor synapses throughout life and that viral delivery of NGL2 can restore synapse numbers and induce the formation of extra synapses in adult wild-type and Ngl2 knockout mice, respectively. Based on these findings, we hypothesized (1) that viral delivery of NGL2 may be able to rescue synapses in inherited retinal degenerations (IRDs), a genetically heterogenous group of diseases in which photoreceptors lose synapses and die causing visual impairments, including blindness. We also hypothesized (2) that NGL2- mediated synapse rescue may be neuroprotective and slow photoreceptor death. Our preliminary data support these hypotheses, which we further test in this proposal. In Aim 1, we will examine whether synapse loss and photoreceptor degeneration progress differently IRD models on wild-type and Ngl2 KO backgrounds. We will determine to what extent and at which time viral delivery of NGL2 can rescue synapses, protect photoreceptors, and preserve vision. Finally, we will assess whether NGL2-gene therapy generalizes across IRD models that differ in pathogenesis and disease progression. CAMs nucleate large protein complexes that control synapse formation, maintenance, and function. The molecular composition and signaling mechanisms of these complexes is mostly unknown. In Aim 2, we will analyze the composition of NGL2 complexes at photoreceptor synapses and determine the contributions of NGL2’s interaction partners to synapse maintenance, rescue, and neuroprotection. Many therapeutic approaches to neurodegeneration that succeed in mice are lost in translation to humans. Differences in the anatomy, cellular composition, transcriptome, and function of neural circuits in mice vs. humans contribute to this translational challenge. We have developed an organotypic culture system of human retinas of patients undergoing enucleation surgery and organ donors. We have established an IRD model in this system. In Aim 3, we use these advances to test the ability of NGL2 and its interaction partners to rescue synapses and protect photoreceptors in the human retina. Together our studies will determine the potential of CAM-gene therapy for the treatment of neurodegenerative disease. Our mutation-agnostic approach could be widely applicable for genetically heterogenous IRDs and may be adaptable to neurodegenerative disease in other parts of the nervous system.

NIH Research Projects · FY 2026 · 2017-04

ABSTRACT Parkinson disease (PD) is a progressive neurodegenerative disease characterized by motor, cognitive, and psychiatric manifestations resulting from abnormal protein deposition and neurotransmitter deficits. The variability in clinical presentation and progression likely reflects different PD subtypes, which may be associated with the underlying variability in brain pathology. Although current treatments provide dramatic motor benefit in PD, they fail to alleviate some aspects of gait impairment and non-motor symptoms and may exacerbate cognitive and psychiatric features. To develop more personalized interventions to treat, forestall or prevent these features, more information regarding PD subtypes is necessary for patient selection and stratification, predicting progression, and evaluation of novel treatments. Therefore, we propose to examine the clinical and prognostic utility of PD subtypes and their stability through longitudinal behavioral assessments; determine the biological markers of PD subtypes from multimodal neuroimaging, CSF, and brain autopsy data; and translate the PD subtypes to a clinical setting.

NIH Research Projects · FY 2025 · 2017-03

ABSTRACT Breast cancer (BC) is the most common cancer diagnosed in women. Approximately 70% of BCs are estrogen receptor (ER) positive (ER+) and human epidermal growth factor receptor 2 negative (HER2-). Endocrine therapy (ET) reduces recurrence risk and improves survival for many in this group. However, despite standard of care and adjuvant ET, over 20% patients with ER+/HER2- BC experience metastatic recurrence in the years to come, and virtually all patients with metastatic disease eventually experience disease progression on ET due to intrinsic or acquired resistance mechanisms. There are currently no biomarkers that reliably identify which of these advanced breast cancer patients will benefit from ET-based approaches so that chemotherapy could be avoided or delayed. To address this unmet need, the objective of this proposal is to develop co-clinical quantitative PET/CT imaging strategies integrated genoproteomic discovery to predict response to ET in patients with ER+/HER2- metastatic breast cancer (MBC). To that end, we will interface with a recently awarded phase II multicenter Translational Breast Cancer Research Consortium (TBCRC) trial to assess the functional status of estrogen receptor in patients with ER+/HER2- MBC. The U24 will have three specific aims: in Aim 1 we will optimize animal modeling and the quantitative accuracy of PET imaging agents of response to ET in ER+/HER2- BC patient-derived tumor xenografts (PDX). In Aim 2 we will implement optimal quantitative methods to predict response to ET in ER+/HER2- and integrate with multi-scale genoproteomic data across the co-clinical trial. And in Aim 3 we will populate content from the co-clinical investigation on a web-accessible research resource and expand capabilities of co-clinical database (CCDB). In addition, high value multi-scale analytic data will be generated, including whole exome sequencing (WES), RNASeq, pathology, and CODetection by indEXing (CODEX) to characterize tumor heterogeneity. All data will be uploaded to an informatics resource available to the co-clinical community to test new algorithms and mine for novel leads integrating imaging and multi-scale analytic data to predict therapeutic response. Overall, this proposal aims to have a far-reaching and high impact on the implementation of precision medicine in identifying, stratifying, and predicting response to ET+CDK4/6i in patients with ER+/HER2- MBC, integrating quantitative imaging with genoproteomic discovery.

NIH Research Projects · FY 2026 · 2017-03

Abstract Natural killer (NK) cell tolerance to self is incompletely understood despite wide-spread acceptance of the now familiar “missing-self” hypothesis. Serving as a guiding principle for several decades, it proposed that NK cells survey tissues for ubiquitously expressed major histocompatibility complex class I (MHCI) molecules as self. Normal levels of MHCI do not allow NK cell attack but if MHCI is down-regulated in a pathologic event, NK cells attack. The applicant and his laboratory discovered the Ly49 family of receptors specific for target cell MHCI molecules that inhibit NK cell activation receptor function, providing a basis for understanding the missing-self hypothesis. However, some predictions of the missing-self hypothesis were not observed, such as hyper-reactive NK cells in MHCI-deficient hosts, rather hypo-responsive NK cells were found. This can now be explained by other findings from the applicant's laboratory that the inhibitory Ly49 receptors have a second function to license or educate NK cells to self-MHCI, such that licensed NK cells have functionally competent activation receptors. Meanwhile, other issues. For example, in prior studies, the applicant's lab showed that different MHCI alleles appear to have graduated effects on Ly49 functions, suggesting signaling strength accounts for these functions, possibly due to Ly49 affinities for self-MHCI, in part related to the rheostat model for receptor function that has not been well studied. Moreover, MHCI affects the repertoire of Ly49s that are expressed in a variegated manner with more than one Ly49 per NK cell. Data from the applicant's laboratory suggest that signaling by a self-MHCI-specific Ly49 influences expression of another Ly49 that is self-MHCI- specific, potentially providing an explanation for how MHCI alleles affect the Ly49 repertoire. Finally, it is not known how the Ly49s confer licensing, such as the possibility that the inhibitory receptors may directly signal the licensing phenotype. Herein the applicant proposes to study NK cell tolerance utilizing novel mice recently generated in his laboratory, including knockout mice lacking all Ly49 expression on conventional NK cells and knockin mice with essentially monoclonal expression of a single Ly49 on all NK cells. Therefore, the Specific Aims of this proposal are to study: 1) Ly49 affinity for self-MHCI in licensing, effector inhibition and missing-self; 2) Establishment of the Ly49 repertoire; and 3) Inhibitory Ly49 signaling. Thus, these studies will markedly enhance our understanding of NK cell tolerance.

NIH Research Projects · FY 2025 · 2017-01

Abstract. Long-term survival of patients with glioblastomas (GBM) are associated with two competing priorities: 1) gross total resection and 2) preservation of the patient’s function. Stereotactic navigation, in which reconstructed magnetic resonance images (MRI) of the brain are used for real-time intraoperative anatomic guidance, has become an essential tool for tumor resection. Further, there are emerging insights that glioma- specific perturbations of the functional organization of the brain impact the patient’s survival. However, the current barrier is that there is no FDA approved navigation system that enables the surgeon to visualize the functional architecture of the brain and the impact a tumor has on the brain’s network organization to inform prognosis. Resting state functional MRI (rs-fMRI) has emerged as a powerful tool for mapping clinically relevant brain networks and defining critical glioma-neuronal interactions. rs-fMRI is highly efficient, task independent, and multiple resting state networks (RSNs) can be mapped simultaneously. With this in mind, the long-term goal of our research is to improve treatment, survival, and quality of life for patients with brain tumors by improving the identification of eloquent cortex and providing actionable metrics for survival prognosis to best tailor a patient’s care. In our first Academic Industry Partnership between Washington University and Medtronic we were extremely productive in creating an integrated brain-mapping navigation technology using rs-fMRI. Specifically, we created a robust image acquisition/analysis pipeline that includes pre-processing of raw data, quality control analytics, and clinical validation demonstrating superior performance over task-based fMRI. We have also been leaders in deriving prognostic radiomic biomarkers from rs-fMRI. In this continuation, we will build on these successes. The overall objective is to create advanced rs-fMRI machine learning (ML) tools to more efficiently and accurately define functional cortex and provide preoperative prognostic metrics of survival as a comprehensive surgical/care navigation system. We have the expertise, infrastructure, and data, to advance rs- fMRI to be a powerful tool for neurosurgical decision support. The proposal entails three specific aims: 1) Advance an ML algorithm to enable more accurate and data efficient rs-fMRI brain-mapping software, 2) Create an rs-fMRI ML algorithm to preoperatively predict survival in glioblastoma (GBM) patients, and 3) Validate impact of mapping and prognostic algorithms on clinical decision making in prospective feasibility clinical trial. The expected outcome of this work will be an integrated imaging/surgical navigation technology using rs-fMRI for clinical decision support with defined performance, clinical validation, and a regulatory path for FDA clearance. Thus, this proposal is innovative because 1) the software will map networks with substantially shorter image acquisition times, thus enabling more widespread adoption and 2) provide critical pre-operative survival insights to inform surgical decisions. This work is significant because it will disseminate technology that fundamentally enhances more tailored approaches to improving patient outcomes and quality of life.

NIH Research Projects · FY 2026 · 2017-01

Abstract The emergence of drug resistant human immunodeficiency virus type-1 (HIV-1) variants and the lack of an effective vaccine require the development of novel anti-retroviral drugs. The catalytic activity of HIV-1 integrase (IN) has been successfully targeted by several highly effective and well tolerated IN strand transfer inhibitors (INSTIs). However, despite high barriers with the second-generation INSTIs, mutations conferring resistance to multiple INSTIs have been reported in clinical settings. Thus, targeting IN through an alternative mechanism can complement the existing therapeutic strategies and substantially increase the barrier to emergence of drug resistant HIV-1 variants upon INSTI treatment. IN has long been known to have an enigmatic non-catalytic function in the HIV-1 life cycle. Certain mutations in IN, collectively referred to as class II mutations, are reportedly pleiotropic and result in defects in viral particle assembly, maturation and reverse transcription. In a paradigm-shifting study, we have discovered that HIV-1 IN binds to the viral RNA genome (gRNA) in virions and that this interaction is critically important for accurate virion morphogenesis. Inhibition of IN-gRNA interactions through allosteric integrase inhibitors (ALLINIs) or class II IN substitutions results in the formation of aberrant “eccentric” particles with the gRNA is mislocalized between the empty capsid (CA) lattice and the viral envelope. Furthermore, we have shown that IN tetramerization is critical for RNA-binding and that a number of class II IN substitutions located throughout IN inhibit RNA binding through modulation of IN tetramerization. Finally, work from our lab demonstrated that premature degradation of the gRNA and its physical separation from the reverse transcriptase enzyme in virions underlies the reverse transcription defects of eccentric particles in target cells. Importantly, this untimely gRNA degradation is not due to inhibition of IN-gRNA interactions per se, but rather due to loss of protection with the CA lattice, as a similar outcome was observed upon CA destabilization. Together, these studies cemented the role of IN-gRNA interactions in virion maturation and demonstrated the critical importance of the CA lattice in protection of viral nucleic acids in target cells. Based on these novel findings and extensive preliminary data, we propose to elucidate the nature and rules of HIV-1 IN-gRNA interactions, how IN binding to the gRNA mediates proper assembly of the HIV-1 capsid lattice and how infected cells sense and respond to aberrant particles generated upon inhibition of IN-gRNA interactions and destabilization of the CA lattice. These studies will fill a critical gap in our understanding of the critical noncatalytic function of HIV-1 IN in particle maturation and the consequences of inhibiting these interactions. Together, this project will not only enhance our basic knowledge of HIV-1 replication but also aid in the development of novel antivirals that can complement INSTI-based therapies in clinical settings.

NIH Research Projects · FY 2026 · 2017-01

SUMMARY The goal of this project is to understand how two virulence factors from Mycobacterium tuberculosis (Mtb), CpsA and phthiocerol dimycocerosate (PDIM), impair immunity by undermining both classical and non-classical autophagy. Mtb is the causative agent of tuberculosis (TB), the leading cause of death worldwide from a bacterial infection. The main cellular niche for Mtb is macrophages and neutrophils, the very immune cells that are meant to clear infection. How Mtb survives the innate immune response to establish infection is not well understood. In the previous project period, we discovered that an exported protein, CpsA, is critically important for Mtb virulence. We showed that CpsA inhibits phagosomal recruitment of the NADPH oxidase. The NADPH oxidase makes reactive oxygen species (ROS), an important mediator of the innate immune response. In addition to its direct antimicrobial activity, ROS is required for a lysosomal trafficking pathway called LC3-associated phagocytosis (LAP), a non-classical form of autophagy. Thus, by inhibiting ROS, CpsA also inhibits LAP. We showed both in macrophages and mice that CpsA protects Mtb from the NADPH oxidase and LAP. Interestingly, CpsA physically interacts NDP52 and TAX1BP1, autophagy adaptors that function in a form of classical autophagy (xenophagy), suggesting that CpsA may also impair xenophagy. Moreover, we found that the Mtb virulence lipid, PDIM, also inhibits the NADPH oxidase. Previous studies proposed an array of roles for PDIM and showed that it protects Mtb from a poorly defined innate killing mechanism. Our data suggest that an unappreciated virulence property of PDIM is blocking the NADPH oxidase and LAP. Thus, we hypothesize that CpsA inhibits xenophagy and works in concert with PDIM to inhibit the NADPH oxidase and LAP. Further, we propose that the infectious dose of Mtb depends upon its ability to evade these innate defenses in myeloid cells that are recruited to the lungs during initial infection. Here, we will define how CpsA inhibits the NADPH oxidase and investigate whether it also impairs xenophagy by blocking NDP52 and TAX1BP1 function. We will determine whether PDIM also impairs recruitment of the NADPH oxidase to mycobacterial phagosomes and evaluate the contribution that PDIM plays towards subverting the NADPH oxidase and LAP in vivo. To determine whether CpsA promotes the establishment of infection, we will use an ultra-low dose infection model in mice. Using conditional knockout (cKO) mice, we will determine in which cells CpsA functions to inhibit the NADPH oxidase and LAP during acute and chronic infection. Our findings will provide mechanistic insight into how two key virulence factors in Mtb collaborate to undermine immunity. Our studies will reveal a cell type-specific virulence strategy of the bacilli, delineate the cell types that participate in LAP in vivo, and define host-pathogen interactions that govern the establishment of infection. Our studies will provide detailed mechanistic insight into the immune evasion strategies of one of the world's most formidable pathogens. By elucidating the mechanism of action of two crucial virulence factors in Mtb, we will advance innovative approaches to prevent and treat TB.

NIH Research Projects · FY 2026 · 2017-01

Project Summary/Abstract The goal of this proposal is to understand how the tetraspanin family member CD53 protects hematopoietic stem cells (HSCs) from inflammatory stress. While important for normal immune system function, inflammatory signaling can impair HSC function and promote the development of hematopoietic malignancies. In preliminary data, we identified CD53 as a critical regulator of HSC function in the context of inflammatory stress. CD53 is a member of the tetraspanin family of transmembrane proteins that organize multi-protein networks to regulate a wide variety of cellular processes such as proliferation, migration, and survival. While normally expressed at very low levels in HSCs, CD53 is markedly upregulated in response to multiple stressors including inflammatory cytokines, toll like receptor agonists and mobilizing agents. Using our newly-generated Cd53-/- mouse, we found that loss of CD53 causes a significant reduction in HSC repopulating ability and increased cycling in the face of inflammatory stress. RNA sequencing and proximity labeling studies suggest that CD53 promotes HSC quiescence in response to inflammation via activation of “DREAM,” a transcriptional repressor complex involving the Rb-like family members p107/Rbl1 and p130/Rbl2 that inhibits the expression of cell cycle genes in response to p53 and p21 activation. Based on this data, we hypothesize that CD53 promotes DREAM complex-mediated repression of cell cycle-related genes in HSCs in response to inflammatory stress, thereby promoting HSC quiescence and protecting HSC function. Notably, CD53 expression is markedly increased in HSCs deficient for Tet2 or Dnmt3a. Mutations in these epigenome regulators are commonly associated with age-related clonal hematopoiesis (CH), which involves inflammation-driven expansion of mutant HSCs and increased risk of leukemic transformation. We predict that CD53 may thus promote the clonal advantage of mutant HSCs in CH. Using a combination of proteomic, transcriptomic and in vivo HSC functional tools, we will: 1) Determine the role of CD53 in promoting HSC function in response to inflammatory stimuli; 2) Determine how CD53 regulates HSC cycling and DREAM complex activity; and 3) Determine whether elevated CD53 promotes the clonal advantage of mutant HSCs. We will perform overexpression studies to elucidate the effects of sustained CD53 expression on HSC function, and DREAM knockout mice will be used to determine the role of this complex in mediating the effects of CD53 on HSCs. We will determine the mechanisms by which CD53 regulates HSC cycling and DREAM activation using proximity ligation assays to characterize CD53- interacting partners. Finally, we will perform functional studies and chimeric modeling experiments using Cd53 and Dnmt3a knockout mice to determine whether CD53 promotes the clonal expansion of mutant HSCs. Together, our proposed studies will describe a novel mechanism that enables HSCs to resist inflammatory stress. Ultimately, understanding the mechanisms by which both normal and mutant HSCs resist inflammatory stress is essential to the development of therapeutic strategies to promote healthy HSC function and to prevent the expansion of mutant HSCs in CH.

NIH Research Projects · FY 2026 · 2016-12

PROJECT SUMMARY DNA replication stress is one of the mechanistic underpinnings of many genotoxic chemotherapies which cause replication fork stalling. Stalled forks frequently reverse to form four-way junction structures, as a natural coping mechanism, to allow repair of the DNA lesions ahead of the forks and avoid catastrophic fork collapse. However, fork reversal is a double-edged sword because the nascent DNA in the reversed arm is susceptible to excessive nuclease resection if not securely protected which can lead to severe genome instability. Many factors play tug- of-war at stalled forks to strengthen or weaken their stability, and their balance in the cell determines the fate of replication forks during stress and cellular outcome upon chemotherapy treatments. In this study, we investigate the molecular mechanisms of a novel replication fork regulator named profilin-1 (Pfn1). As a well-known actin- binding protein, Pfn1 plays an essential role in actin polymerization and dynamics. Paradoxically, it also has well- documented but poorly understood anticancer activities including the ability to sensitize cancer cells to chemotherapy treatments. In recently published work, we demonstrated for the first time that the anticancer effects of Pfn1 stem, at least partially, from its nuclear functions that are spatially and mechanistically distinct from its cytoplasmic function in actin regulation. We showed that nuclear Pfn1 directly interacts with ENL in the Super Elongation Complex (SEC) and inhibits the ability of SEC to drive transcriptional elongation of various cancer genes including MYC. We also presented clinical evidence that nuclear Pfn1 level is frequently decreased in cancer due to the upregulation of its nuclear exporter exportin-6 (XPO6), whose deletion increases nuclear Pfn1 level and decreases tumor growth. These findings establish the notion that Pfn1 has fundamentally important and cancer-relevant functions in the nucleus, and set the stage for further discovery of its involvement in additional nuclear processes. In this grant, we present novel evidence that nuclear Pfn1 promotes normal DNA replication but causes fork destabilization during stress and increases cellular sensitivity to replication stress- inducing chemotherapies including PARP inhibitors. We hypothesize that nuclear Pfn1 has context-dependent effects on DNA replication forks. Under normal conditions, it promotes fork progression by increasing chromatin relaxation through SNF2H. Under stressed conditions, it promotes fork reversal by stimulating SNF2H and increases fork resection by suppressing BOD1L, leading to fork destabilization. Aim 1: Determine the effect of nuclear Pfn1 on BOD1L-dependent replication fork protection. Aim 2: Define the role of Pfn1/SNF2H axis in replication fork remodeling and stability. Aim 3: Understand the chemotherapy-sensitizing ability of nuclear Pfn1. Work proposed in this grant has the potential to generate important mechanistic insights and proof-of-concept therapeutic data rationalizing further investigation of nuclear Pfn1 as a synthetic lethal target for genotoxic chemotherapies.

NIH Research Projects · FY 2025 · 2016-09

The DiPersio Unit strives to optimize immunotherapy, including allogeneic hematopoietic stem cell transplantation (alloHSCT), for treating hematological malignancies. HSCT is the only curative therapy for many hematological malignancies and some non-malignant diseases such as hemoglobinopathies, autoimmune diseases, and inherited disorders of metabolism. Key obstacles to the success of HSCT include collecting sufficient numbers of hematopoietic stem/progenitor cells (HSPCs) to proceed to transplant, control of graft- versus-host disease (GvHD), and treating disease recurrence both before and especially after HSCT. Dr. DiPersio has focused over the last 25 years on overcoming these obstacles to HSCT through a bench-to-bedside and back again research approach. I have been fortunate to spend my entire 20-year post-graduate research career working with Dr. DiPersio. During this time, I contributed to 37 of Dr. DiPersio’s peer-reviewed manuscripts, performed pre-clinical studies for five projects that led to first-in-human clinical trials, completed correlative studies for 21 different clinical trials involving over 550 patients, and assisted in the training of 12 post-docs/fellows and nine technicians. Dr. DiPersio’s research program over the next several years will use our strengths in preclinical modeling, cancer genomics and the design and execution of early phase clinical trials to (1) develop novel methods for HSPC mobilization and GvHD treatment; (2) define the genetic and epigenetic changes that contribute to AML relapse after alloHSCT; and (3) perform clinical trials testing bispecific antibody or chimeric antigen receptor T cell (CART) therapies to treat hematological malignancies before or after HSCT. Successful HSCT requires the infusion of an adequate number of HSPCs that are capable of homing to the bone marrow and regenerating hematopoiesis in a timely fashion. We have developed novel polyethylene glycol (PEG)-conjugated small molecule inhibitors of the integrin very late antigen 4 (VLA-4) and demonstrated that they synergistically mobilize HSPCs in mice and non-human primates when combined with plerixafor, a CXCR4 inhibitor. In research program 1, I am testing the efficacy of long-acting versions of our PEGylated VLA-4 inhibitors to (1) mobilize murine HSPCs when combined with BL-8040, a long-acting CXCR4 inhibitor and (2) treat GvHD after alloHSCT when given alone or in combination with baricitinib, a JAK1/JAK2 inhibitor. In research program 2, I am defining minor histocompatibility antigens (mHAs) in mice and man and examining if relapse after alloHSCT is mediated in part via downregulation or loss of immunogenic mHAs. Since 30%-50% of post- alloHSCT AML relapses exhibit MHC Class II downregulation, I am examining the mechanisms of MHCII downregulation in AML and developing approaches to re-induce MHCII on these immunologically cloaked tumors. In research program 3, I am completing correlative studies for trials evaluating the efficacy of Flotetuzumab, a CD123´CD3 bispecific antibody, in patients with AML who relapse after chemotherapy or alloHSCT and testing if CART therapy can be enhanced via administration of long-acting human IL-7 (NT-I7).

NIH Research Projects · FY 2026 · 2016-09

PROJECT SUMMARY The mechanisms that regulate skeletal muscle topography and perfectly align muscle morphology with the overall body plan remain poorly understood. During embryonic development, muscle precursors known as myotubes undergo a dramatic morphogenesis in which the myotube leading edges elongate, navigate to tendons, and then choose pre-determined sites for muscle attachment. Myotube guidance refers to the combined cellular processes of leading edge navigation and targeting decisions that connect muscles with the correct tendons. We have used myogenesis in the Drosophila embryo as an entry point to identify the cellular and molecular mechanisms of myotube guidance. Using forward genetic screens and genomics-based reverse genetics, we identified multiple navigational signals that direct myotube leading edge migration, and uncovered transcription factors that direct muscle morphogenesis. Our live imaging approaches revealed that myotubes actively choose the correct muscle attachment site through a putative contact-dependent mechanism with tendon cells. We hypothesize that the integrated actions of short-range navigational signals, morphogenetic gene regulatory networks, and contact-dependent cell recognition programs direct myotube guidance to ensure muscle topography perfectly complements the body plan. To achieve a comprehensive understanding of myotube guidance, we will investigate the interplay between navigation, cell recognition, and gene regulatory modules. We propose (1) to investigate how multiple navigational signals co-regulate the cytoskeleton to direct myotube leading edge migration, (2) to use functional genomics to understand how a morphogenetic gene regulatory network modulates responses to navigational signals and directs contact-dependent cell recognition, and (3) to uncover the heterophilic protein- protein interactions between myotubes and tendon cells that establish a myotendinuos code. We expect the foundational work proposed in this study will be a necessary first step toward understanding how navigational, cell recognition, and gene regulatory modules cooperate to direct myogenesis in more complex systems and human disease.

NIH Research Projects · FY 2024 · 2016-09

Project Summary/Abstract: The broad, long term objective is to identify host-protective mechanisms that counter pathogen-initiated lung inflammation and injury. Many pathogens secrete proteases to cause direct damage to the lung, but bacteria, fungi and viruses can also co-opt host proteases to increase pathogenicity and promote lung injury. We have studied Pseudomonas aeruginosa (PA) as a model of infection-induced injury to probe how a pathogen-encoded protease called Pseudomonas elastase LasB, a metalloprotease released into the extracellular milieu by the PA type II secretion system, induces lung tissue damage and secondary trigger of host-derived serine proteases such as neutrophil elastase (NE). We previously identified thrombospondin-1 (TSP-1), a matricellular protein secreted by a variety of cells following injury that disarms both pathogen-encoded LasB and host protease NE, to limit lung injury and inflammation. Key questions that have arisen from this work is how an unregulated proteolytic environment drives excessive inflammatory response and dysregulated repair following injury, and what are the host factors that calibrate this response in the lung. Our preliminary findings suggest that a feed-forward neutrophilic inflammatory response occurs in the proteolytic environment of PA infection through N-terminal processing of IL-36γ that is exaggerated in the absence of TSP-1. Moreover, platelet TSP-1 appears protective against PA-induced lung injury, but the precise mechanism related to TSP-1's role at the alveolar-capillary interface remains unknown. Furthermore, we show that PA elastase activity in clinical strains confer excessive inflammation and injury in mice and is associated with worse clinical outcomes when compared with non-elastase producers. Based upon these findings, we propose the following aims utilizing genetically deficient mice, cell-specific conditional knockout systems, and PA clinical respiratory isolates obtained from the ICU to (1) elucidate the mechanisms by which TSP-1 counters the hyperinflammatory response mediated by proteolytic processing of the pro-inflammatory cytokine IL-36γ; (2) examine the contribution of TSP-1 and platelet TSP-1 in protection against alveolar barrier disruption and stabilization of the early provisional matrix following lung injury; (3) determine whether PA ICU respiratory isolates with elastolytic properties drive unwarranted inflammation and persistent tissue injury in the susceptible host. A better understanding of host biology during severe respiratory infection could prove useful in the rational design of targeted therapeutics against pathogen-derived proteases and deregulated host inflammation as adjuncts to current antimicrobial agents and supportive pulmonary and critical care management.

NIH Research Projects · FY 2026 · 2016-09

Project Summary Neural circuits in the visual system must distinguish between self-generated motion and external object motion to guide survival behaviors, such as predator evasion or prey capture. Here, we will test the overarching hypothesis that a conserved retinal interneuron, the TH2 amacrine cell (TH2 AC), cancels self-motion in vision. We will investigate the mechanisms by which TH2 AC dendrites detect, integrate, and distribute motion signals across multiple retinal pathways and how these signals are transformed in the brain to guide behavior. Most amacrine cells lack axons and perform complex input-output computations in their dendrites. The nature of these computations and the underlying mechanisms remain mysterious. In Aim 1, we will combine light and serial section electron microscopy (3DEM) with two-photon imaging to map the input distribution and function on TH2 AC dendrites. We will combine experiments with compartmental computational modeling, to elucidate how synaptic inputs interact with the branching patterns and long-distance integration of TH2 AC dendrites to generate robust motion responses in two arbor compartments encoding different spatial information. Core features extracted by mammalian retinas—such as object motion—are encoded in a distributed manner across groups of retinal ganglion cell (RGC) types, whose responses vary on a common theme. How amacrine cells shape distributed encoding is unknown. In Aim 2, we will map TH2 AC outputs by 3DEM and optogenetics and combine computational population modeling with opto- and electrophysiology and cell-type-specific silencing and removal to reveal how TH2 ACs assemble bipolar cell and RGC surrounds to control gain, suppress noise, and cancel self-motion signals and shape distributed encoding of object motion across retinal pathways. How retinal features—shaped by dendritic AC computations and distributed across RGCs—are transformed in the brain to guide behavior is unclear. In Aim 3, we will trace retinal object-motion signals to the superior colliculus (SC), where TH2 AC-target RGCs converge on wide-field (WF) cells that survey large areas of visual space. By combining retrograde tracing, in vivo recordings from WF cells in awake mice, and 3D multi-camera tracking of mouse-cricket interactions, we will investigate how TH2 AC–mediated noise suppression, gain control, and self- motion cancellation help WF cells balance high convergence with single-input sensitivity. We will also determine how these mechanisms refine object-motion selectivity and figure-ground segregation to promote successful prey capture. Collectively, these studies deepen our fundamental understanding of vision, provide benchmarks for assessing visual function in disease and therapy, and could inspire new approaches to machine vision, particularly in resource-constrained settings.

NIH Research Projects · FY 2024 · 2016-09

PROJECT SUMMARY Urinary Stone Disease (USD) is an increasingly prevalent and highly recurrent condition associated with major morbidity at a rising cost to society. Thus, improved management can significantly reduce its health burden. Increasing fluid intake is recommended to all USD patients. However, knowledge gaps persist regarding the impact of fluid therapy in preventing USD recurrence including effectiveness of strategies to achieve and maintain a high urine volume, and whether such strategies reduce USD recurrence. The Prevention of Urinary Stones with Hydration (PUSH) study is a randomized clinical trial investigating the impact of increased fluid intake and increased urine output on the recurrence rate of USD in adults and children. In this study 1,642 participants will be randomized to a control or intervention arm. Participants in both arms receive a “smart water bottle”. The intervention arm involves an additional program of behavioral interventions, including financial incentives, structured problem solving, and low touch interventions designed to improve adherence to a prescribed fluid intake regimen. The primary endpoint is occurrence of a stone event during a two- year observation period. The PUSH study is in its third year, and due to multiple challenges to recruitment of study participants, follow-up of participants and data collection have not yet been completed. Additional time is needed to ensure study completion and to accomplish all study goals. Although ureteral stenting is routinely performed after urological procedures for USD to mitigate peri- operative complications, stents cause significant patient discomfort. The causal mechanisms are only partly understood. The STudy to Enhance uNderstanding of sTent-associated Symptoms (STENTS) is a prospective observational cohort study enrolling adolescents and adults undergoing ureteroscopic intervention for ureteral and/or renal stones. Participants undergo detailed symptom assessment using validated questionnaires, a psychosocial assessment, quantitative sensory testing for evaluation of pain sensitization, and detailed collection of clinical and operative data. Biospecimens (blood and urine) are being collected for future research. Recruitment to the STENTS study and follow-up of the participants are expected to be completed on time. However, additional time and resources are needed for analysis of collected study data. In Aim 1 of this application, the investigators will continue and complete participant enrollment for the PUSH study, continue biospecimen collection for the NIDDK Repository, analyze the data, and prepare and submit several planned manuscripts related to the study hypotheses. In Aim 2 of this application, the investigators will analyze the data from the STENTS studies, interpret findings, and disseminate findings through peer reviewed publications.

NIH Research Projects · FY 2026 · 2016-08

Abstract People with Parkinson disease (PD) frequently develop dementia, which is associated with neocortical deposition of alpha-synuclein (α-syn) in Lewy bodies and referred to as Lewy body dementia (LBD), an Alzheimer’s Disease Related Dementia (ADRD). In addition, neuronal loss and deposition of aggregated α-syn also occurs in multiple subcortical nuclei including substantia nigra (dopaminergic), nucleus basalis of Meynert (cholinergic), locus coeruleus (noradrenergic) and dorsal raphe nuclei (serotonergic). Accumulation of α-syn likely contributes to degeneration of cortical neurons, which may also be affected by widespread Aβ accumulation that occurs in approximately 55% of PD with dementia cases and widespread tau accumulation in fewer cases. However, the affected subcortical nuclei project rostrally to thalamic, striatal, limbic and neocortical regions, and the loss of innervation from these nuclei also may contribute to cognitive impairment in PD. We developed postmortem tissue analysis methods to quantify pathologic accumulation of α-syn, Aβ and tau, neuronal degeneration marked by loss of synaptic terminals, and loss of innervating projections from dopaminergic, serotonergic, noradrenergic and cholinergic subcortical neurons. In this project we will collect autopsies from a longitudinal study of PD participants that measures cognition, neurobehavioral function and gait. We will sample cerebellar, basal ganglia, limbic and neocortical regions from frozen brain tissue for each autopsy case and analyze the tissue with the following goals: 1) Determine the relationship between α-syn, Aβ and tau accumulation, neuronal degeneration, and loss of subcortical projections. 2) Determine whether pathologic protein accumulation, neuronal degeneration and loss of projections from subcortical nuclei relate to global cognition and specific cognitive phenotypes, including impaired attention, memory, visuospatial and executive function. Defining the pathologic substrates for cognitive impairment in PD will provide further guidance for therapeutic targets, biomarkers, and outcome measures for therapeutic trials in PD.