University Of Colorado Denver

NIH Research Projects · FY 2024 · 2021-09

Summary To ensure a successful and sustained response to the COVID-19 crisis it becomes imperative that the functional implications of the considerable genotypic and phenotypic variation in natural human immunity are understood. Natural killer (NK) cells have major roles in controlling the innate and adaptive immune response to viral infections, including herpesviruses, HIV-1, influenza, and SARS. NK cells comprise a significant part of the front-line defense against pathogen invasions and are present at large numbers in lung tissues. NK cell effector functions, including cytokine release and cytotoxicity, are modulated by interactions of killer cell immunoglobulin-like receptors (KIR) with class I human leukocyte antigens (HLA) expressed on tissue cells. Across individuals, there is enormous diversity in the number and nature of viable receptor and ligand pairs and within individuals, there is a multitude of NK cell subsets distinguished by their receptors. Previous studies of epidemic diseases have identified clear relationships between this diversity and susceptibility, resistance or control of infection. Likely reflecting exposure throughout human history to multiple, diverse and geographically discrete pathogens, the HLA and KIR genes are highly variable across individuals and population groups. These genetic variations have direct impact on NK cell functions and the response to infection. Allotype-dependent interactions of KIR with HLA inform, modulate and diversify NK cells in their role of identifying and eliminating virus-infected tissue cells. Consideration of the full extent of this variation across human populations is thus critical to understanding, diagnosing and treating SARS-CoV-2 infection, and for developing and testing vaccines. The overarching hypothesis that we will investigate is that genetic variation of HLA and KIR can determine the course of immunity following SARS-CoV-2 infection, leading to severe COVID-19 in some individuals. The first Aim of our study will examine a large multi-ethnic cohort of 11,500 SARS-CoV-2 infected patients, to determine the association of HLA and KIR genetic diversity with severity of disease. The cohorts are drawn from the countries hardest hit by the pandemic, including Brazil, Italy, Spain, UK and the USA. NK cells recognize infected cells through loss of ligands for inhibitory receptors or gain of ligands for activating receptors. Many viruses are known to exploit any or all of these mechanisms to evade immune detection. The second Aim will examine the role of SARS-CoV-2 derived proteins in evading NK cell driven immune responses, and how this varies across all known HLA and KIR allotype interactions. NK cells can be activated by antibodies that are bound to virus segments on the surface of infected cells, and we have shown this activity is also dependent on HLA and KIR diversity. The final Aim will therefore examine the role of antibody-dependent elimination of SARS-CoV-2 infected cells, and the impact of KIR and HLA polymorphism on this response. Validating our approach, our preliminary findings already identified one potential therapeutic target. Our findings will thus have immediate consequence for identifying individuals most at risk for developing severe COVID-19, for developing both universal and personalized treatment, and to aid in vaccine design.

NIH Research Projects · FY 2025 · 2021-08

PROJECT SUMMARY/ABSTRACT Each year more than 700,000 patients survive an episode of acute respiratory failure (ARF) that required endotracheal intubation with mechanical ventilation. Occurring in as many as 44% of these ARF survivors, post-extubation aspiration is associated with deleterious consequences including pneumonia, percutaneous feeding tube placement, long term care facility admission, and increased hospital mortality. Nevertheless, the complications of aspiration must be weighed against the consequences of inappropriately delaying the resumption of oral feeding. Delayed resumption of oral nutrition is associated with prolonged enteral tube feeding, increased caregiver burden, patient dissatisfaction, and increased health-related costs. With the support of an NINR R21 award, our multidisciplinary and multicenter team discovered three novel findings: 1) we developed a 5-item screening decision tree algorithm that detects patients at high risk for post-extubation aspiration; 2) we identified certain unique upper airway abnormalities that are associated with post-extubation aspiration and dysphagia; and 3) the size of the endotracheal tube (ETT) is independently associated with post-extubation aspiration. This proposal will continue to utilize our robust and established multi-center research group at Colorado, Boston University, Stanford, and Yale; all of whom have long-standing dysphagia and aspiration research groups focused on ARF. We will conduct a multi-center cohort study with three complementary aims that are interrelated and use the same patients; but are not dependent upon the results of each other. Aim #1 will determine whether our 5-item decision tree algorithm is a more effective screening tool to identify patients at high risk for post-extubation aspiration compared to the three-ounce water swallow test (3-WST) and the Toronto Bedside Swallowing Screening Test (TOR-BSST). These results will establish the optimal screening test for post-extubation aspiration and identify those patients who require further invasive diagnostic testing such as a FEES examination. Aim #2 will identify unique subphenotypes of patients with post-extubation aspiration based upon FEES-related measures of upper airway structure and function. Using a novel latent class analysis, we will determine whether these subphenotypes are associated with different trajectories of recovery and identify unique patients who benefit from different personalized and targeted therapies. Aim #3 will determine the association between ultrasound determined ETT size/tracheal diameter ratio and post-extubation aspiration while accounting for other confounding variables. This aim will identify the most effective method to select the ETT size that optimizes ventilatory management and decreases post- extubation aspiration. The results of this aim will pave the wave for dramatic improvements in the intubation process for all ARF patients who require mechanical ventilation. With an innovative and protocolized approach, the results of this proposal will demonstrate novel methods to minimize post-extubation aspiration, transform the multidisciplinary care of ARF survivors, and improve patient outcomes and quality of life.

NIH Research Projects · FY 2025 · 2021-08

ABSTRACT – University of Colorado Denver With our participation in this iteration of the Collaborative Pediatric Critical Care Research Network (CPCCRN), we contributed to a series of discoveries that enhanced our understanding of disease processes and resulted in a multitude of potentially effective treatments for critically ill and injured children that require definitive evaluation in clinical trials. There continues to be tremendous need for a collaborative research network in pediatric critical care to facilitate the swift and effective translation of scientific and technologic discoveries into clinical advances. Access to appropriate patients and expertise in the execution of clinical trials must be linked to scientists and laboratories working within and across disciplines at the cutting edge of scientific advancement. At the University of Colorado Denver (UCD) and Children’s Hospital Colorado (CHCO), we have a long history of successful collaborative basic, translational, and clinical science programs focused on pediatric critical illness and injury. We will provide several unique areas of strength to the CPCCRN, including: 1) a distinctive multidisciplinary team of collaborative researchers with specific expertise in mechanistic investigations of cardiovascular and pulmonary diseases, 2) a proven and successful track record as a clinical site in CPCCRN, 3) expertise with pediatric clinical trials execution, including innovative programs to enhance positive research culture among clinical staff and participation among patients and families, 4) a 7-state catchment area with high volumes of critically ill and injured children, 5) an in-house rehabilitation program and extensive outpatient outreach program capable of providing follow-up care to nearly all our critically ill patients, and 6) a dynamic partnership with the Children’s Hospital Colorado Research Institute and Colorado Clinical Translational Science Institute to provide extensive research resources. Our ancillary site, Arkansas Children’s Hospital (ACH) provides 1) experience with pediatric clinical trials execution, including previous successful membership in CPCCRN 2) provision of care to nearly all of the critically ill children from the State of Arkansas and a 4-state catchment area, 3) representation of an Institutional Development Award (IDeA) state (historically received low levels of NIH support) and involving their unique populations (rural, Marshallese, and medically underserved communities) in the NIH mission, and 4) a rich partnership with the ACH Research Institute and University of Arkansas Clinical Translational Science Award to provide comprehensive research resources. Utilizing these assets, we will augment the contributions of CPCCRN to the care of critically ill children through rigorous clinical trial execution, innovative translational and clinical trial proposals, participation in CPCCRN leadership committees, and training the next generation of pediatric critical care researchers. The proposed trial of “Personalized Immunomodulation in Sepsis induced Multiple Organ Dysfunction Syndrome” is an incredibly innovative approach that moves beyond a “one therapy for all” paradigm to an individualized therapeutic approach that will dramatically improve outcomes for children with sepsis. We are enthusiastic and committed to participate in this project with the new 24-site CPCCRN.

NIH Research Projects · FY 2025 · 2021-08

PROJECT SUMMARY & ABSTRACT Does what time of day you exercise matter for weight loss? The objective of this proposal is to determine the effect of an equivalent dose of morning versus evening aerobic exercise on change in body weight, energy intake (EI), and components of energy expenditure (EE) in adults with overweight or obesity. Nearly two-thirds of US adults who attempt to lose weight engage in exercise as a strategy for weight loss. However, weight loss from exercise alone is often substantially less than predicted based on the calories burned in exercise. This is due to compensatory changes that occur in response to exercise (e.g. increases in EI and decreases in non-exercise EE) that limit the energy deficit produced by exercise. Thus, strategies that reduce the compensatory response could enhance the weight loss efficacy of exercise. Preliminary data suggests that morning exercise may limit development of compensation and thus be superior to evening exercise for weight loss. In a secondary data analysis of a 10-month supervised exercise intervention in adults with overweight or obesity, individuals who predominantly exercised in the morning exhibited three-fold greater weight loss compared to individuals who predominantly exercised in the evening, despite equivalent exercise EE and adherence. Further, 81% of morning exercisers achieved ≥5% weight loss, compared to 36% of evening exercisers. There was also evidence of differences in compensation. Despite no intervention on diet, morning exercisers decreased EI across the intervention, while evening exercisers increased EI. Morning exercisers increased total daily energy expenditure (TDEE) proportionately to the EE of exercise, while evening exercisers demonstrated attenuated increases in TDEE. However, these results were from a retrospective analysis in which participants were categorized by the time of day in which they predominantly performed exercise sessions. A rigorously designed randomized trial is needed to confirm these novel findings. In the proposed study, adults with overweight or obesity will be randomized to 7 months of supervised aerobic exercise (2000 kcal/week) performed either in the morning or the evening, and then followed for an additional 6 months. The central hypothesis is that morning exercise will result in greater weight loss as compared to evening exercise due to attenuated development of compensatory changes in EI and EE. The study will compare the effects of morning versus evening exercise on changes in body weight and body composition (Aim 1), changes in EI and appetite (Aim 2), changes in EE, non-exercise physical activity, and sedentary time (Aim 3), and changes in meal timing and sleep (Exploratory Aim 4). The approach is rigorous and innovative as exercise will be prescribed based on EE using indirect calorimetry, free- living TDEE and EI will be measured objectively (doubly-labeled water), and 24-hr patterns of physical activity, sedentary time, meal timing, and sleep will be measured to understand pathways through which exercise timing alters energetics and weight loss. The study is significant as it could identify a practicable strategy to enhance weight loss from exercise and provide insight on how timing of exercise impacts compensatory behaviors.

NIH Research Projects · FY 2025 · 2021-08

Project Summary: This proposal addresses a significant public health problem – childhood obesity and related health disparities faced by Hispanic children from low-income families – for which interventions are lacking. This Randomized Controlled Trial (RCT) will test the effectiveness of a family-inclusive weight management program developed for low-income Hispanic populations. Interventions to prevent and treat childhood obesity have historically excluded Spanish speaking families. The intervention to be tested has been designed through in-depth collaborative research involving childhood-obesity clinician/researchers, safety-net clinics, community organizations, and the low-income and mostly Hispanic families they serve. The Healthy Living Program / La Vida Saludable (HeLP) has been designed to achieve reach, retention, and effectiveness for low-income and Hispanic families, including Spanish Speakers. HeLP incorporates the core components of evidence-based family-based-behavioral- therapy at a dose the US Preventive Services Task Force has recommended to be effective. The key novel design components of HeLP include: 1) Inclusion of the entire family. Previous interventions have focused on parent-child dyads. In HeLP, children with obesity and all siblings 2-16 years and adult caregivers receive targeted curricular components. Younger healthy weight siblings of children referred for obesity are targeted for obesity prevention, 2) Hands-on family training in meal-planning, shopping, and cooking is focused on reducing food insecurity while adapting traditional cuisine, 3) Delivery close to home through a partnership of primary care clinics and recreation centers, and 4) Delivery by trained, low-cost, culturally and linguistically concordant health educators. The goal of this research is to definitively assess using a randomized controlled design the effectiveness of the HeLP intervention for treatment and prevention of overweight and obesity vs a primary care counseling protocol active control condition—Recommended Treatment of Obesity in Primary Care (RTOP). The aims of this study are to: 1) compare the effectiveness of HeLP vs. RTOP at BMI reduction in three age groups of low- income Hispanic children with obesity: 2-6, 7-12, and 13-16 years (secondary measures include: cardiometabolic lab tests, fitness, quality of life, eating behaviors, and food security); 2) To compare the effectiveness of HeLP at obesity prevention by comparing the BMI trajectory of non-obese 2-11yr old children with BMI above median whose siblings are enrolled in HeLP vs. RTOP; 3) To study implementation of HeLP and RTOP within the RE-AIM framework, including: Reach, Adoption, Implementation (fidelity, replication costs, and cost per unit BMI change), and Maintenance. This proposal seeks to fill a major gap in understanding of how childhood obesity might be effectively treated in low-income and Hispanic families.

NIH Research Projects · FY 2025 · 2021-08

PROJECT SUMMARY Eating disorders have the highest mortality rate of all psychiatric illnesses and profound health consequences for those affected. Relative to their heterosexual peers, sexual minority women are at heightened risk of developing an eating disorder, yet little is known about the risk factors contributing to this disparity. The overarching goal of this Pathway to Independence Award is to gain advanced training and mentorship required to launch Dr. Simone's career as an independent investigator who rigorously investigates the mechanisms contributing to disparities in sexual minority populations. To achieve this goal, a multifaceted training plan including coursework, training, mentorship, and research is proposed in: (1) community-based and community- engaged research, (2) mixed-methods design and analysis, and (3) sexual minority health equity research. The knowledge derived from this training plan will prepare Dr. Simone to execute the proposed research, including three research aims and two data collection efforts. During the K99 phase, a mixed-methods case-control study will gain a broad scope of population-specific eating disorder risk factors in sexual minority women (Aim 1) through rich quantitative assessments about eating disorder symptoms, sociocultural pressures and minority stress, and video diaries that capture the response to discrimination, social pressure, and eating disorder episodes in real time. The results from Aim 1 and the community stakeholder advisory board (n=6) will inform the scope of the subsequent research. A well-known limitation of health disparity research is a lack of validated measures for use with minority samples. In line with NIH priorities, during the R00 phase, a population-specific measure of eating disorder risk factors will be developed (Aim 2) with guidance from the community advisory board and pertinent themes from Aim 1. Cognitive interviews with 30 sexual minority women will elucidate the content validity of each item. The psychometric properties of the scale will be derived from the Aim 3 sample. Finally, the temporality of associations between population-specific risk factors and eating disorder behaviors and the extent to which such relations differ by sexual identity (e.g., queer, lesbian), race, and ethnicity (Aim 3) by employing a daily process ecological momentary assessment in a case-control study. The proposed novel mixed-methods research will elucidate population-specific eating disorder risk factors in an understudied population, sexual minority women. Identification of population-specific risk factors and their associations with eating disorder symptoms in racially and ethnically diverse sexual minority women will produce guidelines for community programs and intervention efforts to rectify disparities, and the proposed measure will enhance surveillance of population-specific eating disorder risk and advance future research of such mechanisms. This award will launch the PI's independence by laying a foundation for a career in innovative multi-method research that may be applied to additional mental health outcomes wherein disparities are present and with other diverse sexual and gender minority populations.

NIH Research Projects · FY 2025 · 2021-08

ABSTRACT Repeated concussions (mild Traumatic Brain Injury, mTBI), which are particularly prevalent in athletes and military personnel, can lead to long-term brain health issues including dementia, depression, and other psychiatric conditions. Recent studies suggest that mTBIs may give rise to increased risk for Alzheimer's disease (AD) or other AD-related dementias (ADRDs), but there are few conclusive studies, and no reliable blood biomarkers available as a predictive diagnostic tool. We are studying a unique cohort of NCAA Division I athletes in high impact sports to develop a reliable blood biomarker assessment and examine biological mechanisms for AD/ADRD risk after multiple mTBIs. To our knowledge, studies have not been conducted using neuron- or astrocyte-derived exosomes (NDEs vs. ADEs) to detect Tau and amyloid pathology and seeding capacity from those with sports-related brain injuries. The overall hypothesis of this project is that exosome alterations after repeated mTBIs reflect and contribute to long-term risk for AD/ADRD. The hypothesis and the aims have not changed. This supplement application is necessary to accomplish the aims set forth in the parent application. The budget of the parent grant was significantly reduced due to our move from University of Denver to University of Colorado Anschutz Medical Campus (CU). One of the many benefits of our move is that we have been gained access to additional blood samples from individuals with moderate to severe TBIs who are patients at the Intensive Care Unit of UC Health at a relatively low additional cost. When we started examining the samples obtained from the athlete mTBI cohort, we realized that we would need to include all levels of TBI severity to increase the rigor of the study. We have been added to the TBI IRB protocol that was in existence in our department (Neurosurgery) at CU. The addition of these samples will allow a more rigorous approach in the parent grant Aims. The Administrative Supplement will therefore allow the expansion of existing studies and access to valuable new sources for the biomarker study.

NIH Research Projects · FY 2025 · 2021-08

PROJECT SUMMARY/ABSTRACT This K01 Mentored Research Scientist Development Award will provide the candidate, Nicole R. Tuitt, DrPH, with advanced training and structured mentoring to facilitate her transition to research independence. The candidate's goal is to become an independent investigator with expertise in state-of-the-art approaches in the development and evaluation of culturally relevant, strengths-based multilevel interventions which aim to reduce substance use and sexual health disparities among urban American Indian and Alaska Native (AIAN) and Black youth. The proposed scope of work will fill a gap in existing literature on multilevel approaches to reduce substance use and sexual health inequities among youth of color. The inability of efforts to reduce adolescent substance use and sexual health disparities to yield sustained improvements likely stem from failure to account for mutually-reinforcing structural disadvantages and multifaceted mechanisms that include all socioecological levels that underlie adolescent health inequities. Most risk reduction strategies focus only at the individual and interpersonal levels. Developing and operationalizing a strengths-based conceptual framework grounded in the socioecological model is critical to inform a multilevel intervention to reduce substance use and sexual risk- taking among urban AIAN and Black youth. The candidate will employ a transformative, sequential mixed methods approach to accomplish the following specific aims, each training goal is matched with the aim in which the researcher will apply her new skills. AIM 1: Examine multilevel determinants of substance use and sexual risk-taking among Black and AIAN high youth (ages 14-18) in Denver Metro, CO. Training: (1) Advance expertise in theoretical and methodological foundations in social and spatial epidemiology to understand the impact of structural contexts on risk and resilience to adolescent substance use and sexual risk-taking; and (2) multilevel structural equation modeling, with geographic contexts, to explore the direct, indirect, and interactive relationships across levels. AIM 2. Engage urban AIAN and Black youth to elucidate distinct conceptual frameworks of risk and resilience to substance use and sexual risk-taking guided by findings from Aim 1; and identify the most critical levels on which to intervene. Training: (1) Develop skills in scenario-based qualitative interviewing (SBI) to engage youth to interpret quantitative data in the development of conceptual frameworks of risk and resilience. The K01 award will position Dr. Tuitt as a trained researcher with expertise on the influence of structural disadvantages on adolescent substance use and sexual risk-taking as well as multilevel approaches. Given her shift away from a focus on individual behaviors to structural factors, expertise in SBI, social and spatial epidemiology, and multilevel SEM will ensure the candidates successful transition into an independent researcher and will situate her to strengthen the field of adolescent health equity research and prevention science.

NIH Research Projects · FY 2025 · 2021-08

ABSTRACT Chronic infections like cytomegalovirus (CMV) may contribute to both cognitive and physical function decline in older adults and have been implicated in the development of Alzheimer's Disease (AD), vascular dementia, and frailty. Due to immune senescence and immune suppression, older Kidney Transplant (KT) recipients ≥50 years, are at especially high risk of early and frequent reactivations of latent CMV and may fail to reconstitute CMV-specific cell-mediated immunity (CMV-CMI), leading to high rates of CMV disease. This research will investigate for the first time the role of CMV and CMV-CMI reconstitution in cognitive and physical functional decline in KT recipients. This research is relevant to the NIA mission because it will advance the understanding of the critical mechanisms driving CMV-related aging in older adults by studying the impact of CMV on decline in cognitive and physical function, AD, and frailty. This proposal's central hypothesis is that inflammation secondary to CMV replication may explain the mechanism by which CMV may contribute to cognitive and physical function decline, AD, and progression of frailty. This hypothesis will be tested through the following specific aims: 1) To quantify the association of CMV serostatus with trajectories of cognition and physical function in an existing Functional Assessment of Renal Disease (FAIR) cohort of older KT recipients; 2) To assess the relationship of CMV-CMI reconstitution with trajectories of cognitive and physical function and frailty in a newly established Colorado cohort of CMV+ older KT recipients; 3) To investigate the relationship of inflammatory markers with trajectories of cognition and physical function among CMV+ and CMV- KT recipients at 1-year post-KT in Colorado cohort. This approach is innovative because it: 1) allows the first investigation of the relationship of CMV, with the trajectories of cognitive and physical function among older KT recipients, and 2) may identify the role of CMV-CMI and inflammation driving the relationship of CMV with cognitive and physical function decline. This project is significant because it has the potential to improve health outcomes for older KT recipients. Complimentary to the proposed research plan, a five-year mentored career development training plan has been devised that incorporates research training in geriatrics and viral immunology. The candidate is co- mentored by internationally recognized experts in gerontology, viral immunology, kidney transplantation, and epidemiology. The candidate's long-term career goal is to become an independent investigator studying immune protection in the context of chronic viral infections with geriatric outcomes.

NIH Research Projects · FY 2025 · 2021-08

Droughts are an important natural disaster but the health implications for elderly populations in the United States have not been extensively studied. This study seeks to provide evidence on the cardiometabolic health risks posed by drought. Droughts can contribute to groundwater over-pumping, and this may in turn lead to arsenic release from aquifer clays into drinking water sources. The impacts of drought on arsenic exposure and resultant health outcomes in human populations are potentially important and this study seeks to address that knowledge gap. We propose to conduct a retrospective cohort study using data from the San Luis Valley Diabetes Study, as well as hydrogeological measurements of water and environmental conditions in the San Luis Valley, to clarify relationships between drought, arsenic in water, arsenic exposure (as measured by urine biomarker), and cardiometabolic health outcomes including incident diabetes (DM), incident cardiovascular disease (CVD), and all-cause mortality. This work is community-engaged and includes a K12 outreach component. Aim 1 quantifies the relationship of drought to individual arsenic exposures and thence to incident CVD, incident DM, and mortality. Aim 2 generates a model for groundwater arsenic as a function of hydrological predictors using machine learning approaches, and uses this to explore relationships between drought, modeled water arsenic and the outcomes of Aim 1. Aim 3 estimates the association between cumulative arsenic exposure and incident CVD, incident DM, and mortality, using a nested case-control design and predictions over time of arsenic exposure from the Aim 2 model. Aim 4 is a simulation study projecting future CVD and DM outcomes under different scenarios of drought conditions, with and without counterfactual interventions to reduce drinking water arsenic.

NIH Research Projects · FY 2025 · 2021-08

Project Summary/Abstract The Acute Respiratory Distress Syndrome (ARDS) is a common critical illness characterized by severe hypoxemia in response to either direct (e.g. viral pneumonia) or indirect, systemic (e.g. sepsis) insults to the lung. Most patients with ARDS will develop delirium, defined by acute, fluctuating disturbances in cognition. Delirium during ARDS is strongly associated with poor outcomes including long-term disability and death. Despite this clinical significance, the mechanisms responsible for delirium in ARDS remain poorly understood. One recent scientific advance with direct relevance to delirium in ARDS is the endothelial glycocalyx, a chondroitin sulfate (CS)-rich layer that lines the vascular lumen. The glycocalyx is degraded early in lung injury, releasing large concentrations of CS into the bloodstream with resultant increases in hippocampal CS content, a brain region implicated in delirium pathogenesis. Remarkably, the presence of elevated levels of highly- sulfated CS subtypes in humans with critical illness is associated with risk of delirium. In preliminary experiments, I observed that these same highly-sulfated CS subtypes can directly potentiate the activity of α- amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs), the primary excitatory ion channel in the brain. This activation of AMPAR activity may be biologically relevant to delirium during ARDS, as I have observed concordant increases in neuronal excitability in the hippocampi in a lung injury model. The work outlined in this proposal will investigate the importance of AMPAR potentiation by CS in the pathogenesis of delirium in both direct and indirect forms of ARDS. My work seeks to 1) determine if CS is predominantly released from the pulmonary endothelial glycocalyx during lung injury and if release is heparanase-dependent, 2) determine if hippocampal-penetrating CS is responsible for increased AMPAR excitability, and 3) determine whether pulmonary-endothelium derived CS contributes to the pathophysiology of delirium in ARDS through hippocampal AMPAR potentiation. Through these investigations, I will gain expertise in modeling of direct lung injury, isolated perfused lung preparations to isolate and assess pulmonary physiology and pulmonary endothelial biology, ex vivo whole-cell electrophysiology and in vivo electroencephalography in murine hippocampi, and a murine behavioral measure relevant to human delirium. These highly-novel studies will allow me, as an early career physician-scientist, to develop a unique expertise studying the lung-brain axis in delirium, while generating essential preliminary data for future independent research awards (e.g. R01).

NIH Research Projects · FY 2025 · 2021-07

PROJECT SUMMARY/ABSTRACT There are significant gaps in our knowledge of how peripheral GFAP+ glia regulate sensory neuronal activity and their involvement in primary afferent sensitization. The lack of understanding of the roles of peripheral glial cells, mainly due to our inability of selectively perturbing signaling pathways in subpopulations of peripheral glia in vivo, presents an important obstacle in developing creative and effective strategies for investigating glial contribution in chronic diseases. The long-term goal of this project is to target GFAP+ satellite glial cells (SGCs) for gene therapies for preventing and treating peripheral sensitization. The objective of this proposal is to identify signaling pathways underlying SGC-neuron interaction in the sensory ganglia, as well as to characterize the translatome changes in sensory SGC signaling involved in visceral pain and bladder dysfunction. Strong preliminary data and past findings in sensory SGCs in the context of chronic pain led to the central hypothesis that Gq-GPCR signaling in SGCs potently decreases the excitability and activity in bladder- projecting sensory neurons and contributes to neural control of bladder functions. This hypothesis will be tested by pursuing three specific aims: 1) Test the hypothesis that sensory satellite glial Gq-GPCR signaling decreases bladder afferent sensitivity and alters micturition in physiological conditions and in inflammation- induced visceral pain and bladder overactivity model; 2) Identify the molecular link between satellite glial Gq- GPCR activation and decreased sensory neuronal excitability; and 3) Discover the changes in sensory satellite glial translatome during the onset and chronic phase of inflammation-induced pelvic pain and lower urinary tract symptoms (LUTS). Under the first aim, targeted and selective gene manipulation in peripheral GFAP+ glia will be utilized to study the role of SGCs in physiological and pathological regulation of bladder afferent excitability. In the second aim, the cellular and molecular mechanism underlying SGC-neuron interaction will be identified in sensory ganglia explants using approaches similar to those used in studying CNS glial-neuronal interactions in situ. Additional experiments will be performed in vivo to test the therapeutic potential of targeting the identified signaling pathway(s) for alleviating visceral pain and bladder overactivity in this aim. Under the third aim, TRAP technology will be employed to identify changes in sensory SGC translatome as a function of inflammation-induced bladder overactivity and pelvic painin a non-biased manner.The proposed research is significant because 1) it demonstrates a new pharmacogenetic approach for selectively activating peripheral glia in vivo, which benefits broader research community in Neuro-urology research; 2) it is expected to fill the knowledge gap on glial modulation of bladder function, a completely unexplored research field with high therapeutic potential; 3) it provides the first line of evidence for subsequent pre-clinical and clinical trials of targeting satellite glial cells and peripheral adenosine receptors in treating pelvic pain; and 4) it lays the groundwork for genetic analysis on specific subsets of peripheral glia during physiology and disease.

NIH Research Projects · FY 2025 · 2021-07

Project Summary Intracranial electrical brain stimulation (EBS) remains a central method in the clinic as well as for research in several animal model systems. However, little is actually known about the ensembles of neurons activated by typical and clinical intracranial EBS protocols. These stimulation protocols often require a trial-and-error learning period (during and after invasive neurosurgery) to determine what stimulation parameters are effective, if any at all are effective. It remains mysterious why some stimulation patterns work in the clinic while others do not, and what underlying ensembles are activated by various stimulation patterns. It is known that focal electrical microstimulation activates nearby excitable membranes, including neural somas, dendrites, and axons. It is also known that the recruited ensemble of neurons may be locally non- homogenous and that clinical effects may rely more on axons of passage than somatic stimulation. Efforts to model EBS cannot overcome our current gaps in knowledge about the homogeneity of local propagation and brain-wide extent of activation. This ambiguity demands a more detailed understanding of local electric field propagation, particularly in the in vivo mammalian brain. Utilizing recent technological advances, we propose to fill these gaps empirically with high density electrophysiological monitoring and temporally precise fluorescent labeling methods, to quantify clinically relevant activation patterns with high spatial resolution and cell-type specificity. Here we propose an experimental study in mice, based on a biophysically realistic model of mouse cerebral cortex, of the spatial and temporal propagation of activation via focal EBS. The study will test the hypothesis that local electrical stimulation is non-isotropic and cell-type specific. We propose to measure EBS stimulation with more than 1000 electrodes arranges in three dimensions around a site of stimulation, in combination with genetic cell type identity through optotagging (Aim 1). To agnostically isolate brain-wide ensembled activated by EBS, we couple a fluorescent reporter to electrical stimulation for ex vivo whole-brain tissue clearing and light sheet imaging (Aim 2).

NIH Research Projects · FY 2025 · 2021-07

PROJECT SUMMARY/ABSTRACT Osteoporosis can be defined as the progressive loss of bone mass and strength with age, leading to increased risk of fragility fracture. Osteoporotic fracture and fracture-related traits, such as bone mineral density (BMD), are highly heritable and Genome-wide association studies (GWAS) for BMD have identified over 1100 associations for the phenotype of BMD. Further, there are many mono-allelic conditions, such as osteogenesis imperfecta, that lead to low BMD and low-trauma fractures in children. Bone is in a constant state of remodeling, with formation mediated by the osteoblast and resorption by the osteoclast and when these processes remain balanced, there is no net change in BMD. Imbalances in remodeling results in the loss of bone seen in osteoporosis, but a GWAS done for BMD cannot determine which of these physiological processes are affected by each locus. All current fracture prevention therapies focus on tipping the remodeling balance away from bone loss. There are three bone anabolic therapies approved by the FDA, but each of these has black box warnings, each can only be used for a limited time (1 to 2 years respectively) and none of them can be used in children. We have shown in previous work that bone mineralization by the osteoblast is a highly heritable, complex genetic trait and that genetic mapping for the absolute amount of mineralization possible yields information that is complementary to that identified by GWAS for BMD. However, the osteoblast is a highly regulated, complex cell that undergoes an as of yet incompletely described differentiation process, must be able to migrate to the site of bone remodeling, must be able to produce the proteinaceous extracellular matrix of bone and then must be able to execute mineralization. The goal of this application is to identify the key genes and pathways that control these aspects of osteoblastogensis and osteoblast function. In Aim 1, we will map high-resolution quantitative trait loci (QTL) for osteoblast maturation, migration and rate of mineral apposition. In Aim 2, we will use cutting edge Bayesian network analyses based on single cell RNA seq and single cell ATAC seq to define master control genes of various stages of osteoblast development. In Aim 3 we conduct functional follow up on genes found via our preliminary analyses that control the late stages of osteoblast function. We expect that this comprehensive and complementary approach to identify key genes for osteoblastic processes will provide critical insight into how bone is formed by the osteoblast. More importantly, the genes that we identify will serve as potential therapeutic targets capable of increasing bone formation in the setting of osteoporosis and in other formation disorders.

NIH Research Projects · FY 2025 · 2021-07

PROJECT SUMMARY PARP inhibitor (PARPi) use in the clinic is expanding into multiple cancer types, and consequently, PARPi resistance is a growing clinical problem. High grade serous ovarian cancer (HGSOC) tumors and cells remain an optimal model system to assess PARPi response and resistance. We have developed a panel of unique isogenic PARPi sensitive and resistance HGSOC cell lines and patient-derived xenograft (PDX) models. We published that hyperactivation of the Wnt/-catenin pathway promotes PARPi resistance. Through the current literature and our preliminary investigation, we have discovered that Wnt-mediated PARPi resistant HGSOC cells have increased expression of the immune checkpoint, PD-L1, and reduced expression of the tumor suppressor, interferon regulatory factor 1 (IRF1). Further, Wnt/-catenin signaling directly inhibits effector T cell differentiation and promotes a tumor-promoting, M2-like macrophage. We will continue to collaborate with MD2 Biosciences to investigate a first-in-class allosteric -catenin inhibitor, 1525. We hypothesize that Wnt-dependent PARPi resistance inhibits anti-tumor immunity, and combining ICB with Wnt inhibition will promote immune activation to eradicate PARPi resistant HGSOC. We are proposing to use both in vitro and in vivo models to determine the role of PARPi resistance and Wnt signaling in promoting an immune-suppressive environment. In Aim 1, we will use our unique PARPi resistant cell line models to establish -catenin regulation of PD-L1 (gene – CD274) and IRF1. In Aim 2, we will determine whether secreted factors from PARPi resistant cells attenuates T cell activation and promotes macrophage M2 differentiation. In Aim 3, we will use our novel syngeneic and humanized mouse models to assess the 1525 - catenin inhibitor combined with anti-PD-1. The proposed work has the potential for a high impact on understanding ovarian cancer biology and improving therapeutic options. We anticipate combining -catenin inhibition with an immune checkpoint blocker will overcome PARPi resistance and provide a therapeutic option for those who are no longer responding to PARP inhibitors. Thus, the proposed work's long-term goal is to develop an investigator-initiated clinical trial at the University of Colorado.

NIH Research Projects · FY 2025 · 2021-07

PROJECT SUMMARY/ABSTRACT This proposal outlines a 5-year training plan for the transition of the candidate to an independent investigator. The long-term goal of the candidate is to be a physician-scientist at an academic medical center studying the molecular basis of genetic mutations in myelodysplastic syndrome (MDS) pathogenesis, in addition to directing a molecular pathology laboratory. The training plan proposed will expand upon the candidate's clinical and scientific background and provide him access to a supportive environment that will prepare him for a career as an independent investigator. The applicant will be mentored by Dr. Matthew Walter, an expert in MDS genetics and biology and Director of the Edward P. Evans Center for MDS at Washington University (WU). An advisory committee consisting of basic science and clinical/translational MDS experts and pathologists will provide scientific and career advice. The applicant's laboratory is well equipped for these studies, and WU provides a rich environment for training and development as access to core facilities, scientific resources, and clinical samples is exceptional. The experiments outlined in this application aim to clarify the distinct roles of different U2AF1 mutants in MDS pathogenesis, as highly recurrent heterozygous mutations in splicing factor genes (e.g., U2AF1) occur in over half of MDS patients. Hotspot mutations in U2AF1 (encoding U2AF1S34 and U2AF1Q157 mutants) are associated with different clinical features, outcomes, dysregulated RNA splicing, and co-occurring gene mutations in patients with MDS. For example, truncating mutations in ASXL1 cooccur more frequently with U2AF1Q157 than with U2AF1S34 mutations in MDS patients. The candidate has observed U2af1S34F/+ and U2af1Q157R/+ conditional knock-in mice develop different hematopoietic phenotypes, including cytopenias, and have distinct gene expression patterns and aberrant RNA splicing in hematopoietic stem and progenitor cells (HSPC). This proposal aims to understand how the distinct molecular perturbations induced by U2AF1S34F and U2AF1Q157R mutants contribute to the divergent hematopoietic phenotypes observed in MDS patients by determining: (1) how activation of MYC and mTORC1 pathways leads to HSPC dysfunction and multilineage cytopenias in U2af1S34F/+ mice and whether U2AF1S34F-specific splicing alterations result in activation of these pathways, and (2) how mutant ASXL1 and U2AF1Q157 associated histone modification and RNA splicing aberrancies selectively cooperate in MDS pathogenesis. Deciphering the role of MYC and mTORC1 activation will allow these models to be used to test pathway-specific therapeutic strategies to alter cytopenias, and identification of cooperation between ASXL1 and U2AF1 mutations in vivo may reveal novel mechanisms by which RNA splicing and histone modifications synergize in MDS. In sum, the proposed studies and training environment will facilitate the candidate's success in becoming an independent investigator.

NIH Research Projects · FY 2026 · 2021-07

PROJECT SUMMARY Among the devastating illnesses impacting rural America, few exact the physical, social, psychological, and economic toll of lung cancer. Lung cancer is the leading cause of cancer death in rural America, and the levy is particularly acute in Kentucky – a state that not only leads the nation in lung cancer incidence and mortality but is a global epicenter of lung cancer. Despite the prevailing nihilism regarding lung cancer care, lung cancer survivors are living longer. Innovations in prevention, early detection, and treatment, have created substantial optimism and opportunities for long-term lung cancer survivorship. These dramatic changes in the lung cancer care landscape have invigorated the need for quality lung cancer survivorship interventions. The overarching goal of the Kentucky LEADS Collaborative Lung Cancer Survivorship Care program is to reduce the burden of lung cancer by offering an innovative survivorship care approach that improves lung cancer quality of life, overcomes lung cancer stigma, and helps survivors engage with care. Using a novel precision survivorship approach and developed in collaboration with rural community stakeholders, the program's foundation incorporates principles of patient-centered care, shared decision making, and motivational interviewing to build survivor engagement. A large acceptability and feasibility trial conducted in collaboration with nine lung cancer care facilities in Kentucky with lung cancer survivors (N=140) demonstrated the acceptability of the intervention among survivors, caregivers, and lung cancer care clinicians. The study also revealed the feasibility of conducting the proposed study methods in rural cancer care facilities. The proposed project continues this program of research by conducting a two-group parallel randomized clinical trial comparing the impact of the Kentucky LEADS Collaborative Lung Cancer Survivorship Care program (KLCLCSC) among lung cancer survivors (N=300) against an enhanced usual care condition (bibliotherapy+assessment) on quality of life outcomes. Rural-residing lung cancer survivors will be recruited from ten oncology care facilities throughout Kentucky. The project's first aim compares the efficacy of the interventions with regard to lung cancer quality of life among survivors as measured by the FACT-L and other rigorous assessments of patient engagement, symptom burden, psychosocial well-being, and behavior change. The project's second aim evaluates the moderating impact of including caregivers as intervention partners on survivor quality of life outcomes. A third aim evaluates the cost-effectiveness of the KLCLCSC intervention in comparison to the enhanced usual care condition. Based on highly encouraging pilot data collected in collaboration with oncology care programs in Kentucky, this research holds credible potential to establish a new paradigm for addressing the challenges associated with lung cancer and for delivering quality survivorship care to rural-residing, economically distressed lung cancer survivors.

NIH Research Projects · FY 2025 · 2021-07

Enter the text here that is the new abstract information for your application. This section must be no longer than 30 lines of text. This is a new application of the University of Colorado T32 postdoctoral training program in anesthesiology research. The broad objective of this training program is to provide training to MD postdoctoral fellows in clinical and basic science related to anesthesiology and perioperative medicine. We propose the training of one fellow per year who will spend 2 years cumulative time in research in one of the participating training faculty’s laboratories or clinical research setting. We propose to recruit 2 fellows the first year to create a cohort and the opportunity for peer mentoring. In year 3 we will again recruit 2 fellows and recruit our final fellow in year 4, for a total of 5 fellows over the 5 year duration of the grant. The training faculty are from several departments within the University of Colorado System, including Anesthesiology, Division of Cardiology, Cell & Molecular Biology, Emergency Medicine, Psychology and Neuroscience at CU Boulder. Research opportunities are offered by 21 NIH-funded faculty mentors, 9 of whom are in the department of Anesthesiology, with strong records of training postdoctoral fellows. The research opportunities are organized into three major research areas relevant to anesthesiology 1) Neuroscience, cell injury & repair and pain 2) Clinical and Translational Drug abuse and 3) Trauma, coagulation and cardiac physiology. We have developed a didactic program that will provide training in research methods, presentation skills (written and oral), grant writing and responsible conduct of research. In addition to our outstanding group of faculty mentors, we have an external scientific advisory committee and an executive committee that will assist the Program Directors in the selection of candidates and oversight of the training program. The specific aims/goals of the training program are: 1) Recruit outstanding anesthesiology postdoctoral fellows pulling from a local and national applicant pool. 2) To develop an outstanding training program using didactic and experiential training to provide scientific training and the practical skills needed to successfully compete for national funding and develop and independent research career in anesthesiology. 3) Assemble a group of faculty mentors that are not only highly successful scientists, but excellent mentors that help create a supportive mentoring environment that keeps postdoctoral fellows engaged, enthusiastic about research and moving forward towards their career goals of becoming leading academic anesthesiologists.

NIH Research Projects · FY 2025 · 2021-07

PROJECT SUMMARY Despite advances in the treatment of acute myeloid leukemia (AML), only 20–30% of patients achieve long-term disease-free survival (DFS) and treatment options for relapsed AML are extremely limited. The recurrence of AML has been attributed to leukemic stem cells (LSCs) and efforts are now focused on targeting this drug resistant population of cells in order to “cure” AML. Our studies measuring energy metabolism in primary human AML specimens, using reactive oxygen species (ROS) as an indicator of metabolic activity, revealed that LSCs preferentially reside in a ROS-low state. Furthermore, high levels of activated 5' AMP-activated protein kinase (AMPK), a central regulator of metabolic pathways, were detected in the LSCs and that knockdown of AMPK resulted in increased ROS levels and concomitant loss of LSCs. Based on these findings, we propose that AMPK inhibition will leverage LSCs out of the ROS-low state decreasing their viability which may be sufficient for LSC elimination or may sensitize them to conventional therapy. There are few potent and selective AMPK inhibitors; however, the multi-kinase inhibitor sunitinib has been reported as a potent inhibitor of AMPK kinase activity. Therefore, the central goal of our research is to develop potent and selective oxindole-based AMPK-targeted agents and examine the effect of AMPK inhibition or degradation in AML models. We have developed an initial series of oxindoles and although we identified potent AMPK inhibitors from this initial series, we believe further AMPK selectivity and inhibitory potency is possible. We will use computational-based modeling to guide the development of inhibitors and evaluate their AMPK inhibitory activity using in vitro kinase assays. Then, inhibition of cellular AMPK will be determined by measuring the phosphorylation of the AMPK substrate acetyl-CoA carboxylase (ACC) by ELISA in MOLM13 and MOLM14 cells, and select inhibitors will be submitted for kinome profiling (Aim 1). AMPK inhibitors that retain an aminoalkyl side-chain extending out of the ATP-binding site will be coupled to a proteolysis targeting chimera (PROTAC) degrader and their ability to degrade cellular AMPK will be evaluated. The oxindole-based AMPK inhibitors or degraders that have a terminal dimethylamino group that interacts with the DFG motif of AMPK will be modified to incorporate a nitroimidazole hypoxia-activated prodrug moiety that are designed to introduce a tier of LSC selectivity (Aim 2). The effect of AMPK inhibitors and PROTAC degraders on cell viability, metabolism and ROS levels as single agents will be determined in MOLM13 and MOLM14 cells and in primary AML cells, LSCs, and normal hematopoietic stem cells (HSCs). Then, the effect of AMPK inhibitors or degraders in combination with venetoclax will be determined in MOLM13 and MOLM14 cells and in primary AML cells. Finally, the effect of our AMPK inhibitors or degraders as single agents and in combination with venetoclax will be evaluated using primary AML specimens transplanted into advanced in vivo tumor xenograft models (Aim 3). The objective of these studies are to develop a range of chemical tools to evaluate the role of AMPK in maintaining LSC viability and the therapeutic potential of targeting AMPK in AML.

NIH Research Projects · FY 2025 · 2021-07

PROJECT SUMMARY Night shift work is known to increase risk for cardiovascular disease, cancer, and metabolic syndrome, but an adverse effect that has received little attention is the disruption of bone metabolism. Animal and human data suggest sleep restriction and circadian disruption, which are inherent in night shift work, are novel, potentially modifiable risk factors for low bone mineral density (BMD) and increased fracture risk. Humans exposed to several weeks of cumulative sleep restriction and concurrent circadian disruption induced by a recurring 28 hour/day protocol had significantly decreased bone formation, with no change or an increase in bone resorption. These changes in bone metabolism, if persistent, would be predicted to increase fracture risk by limiting the development of peak BMD in young adults and/or accelerating bone loss later in life. In fact, the Nurses’ Health Study identified an increased risk of fracture in postmenopausal women who reported 20+ years of night shift work compared to those who never worked the night shift. The acute and chronic skeletal effects of typical night shift schedules in humans, underlying mechanisms, and bone’s ability to recover or adapt are unknown. This application will fill these knowledge gaps by using simulated acute and real-world chronic night shift work to evaluate its effects on bone. The scientific objectives are to determine the effects of night shift work on bone metabolism, density, microarchitecture and strength, and investigate a plausible underlying mechanism (e.g., increased sympathetic tone) by which night shift work impairs bone metabolism to promote optimal bone strength and healthy aging. The specific aims are to: 1. Expose healthy adults to normal sleep or simulated night shift work to (a) Determine if a typical night shift work schedule acutely uncouples bone turnover markers; (b) Investigate increased sympathetic tone as a mechanism for the disruption in bone metabolism; and (c) Evaluate whether resumption of a normal sleep/wake pattern reverses bone turnover marker uncoupling. 2. Characterize changes in bone turnover markers, BMD, bone microarchitecture and strength by evaluating a cohort of hospital nurses in their first year of night compared to day shift work. This interdisciplinary, collaborative research will enhance the health of individuals. It will generate human data to establish night shift work as a novel, potentially modifiable risk factor for impaired bone health and inform mechanisms by which it alters bone metabolism in women and men. This knowledge will provide an opportunity to intervene to prevent low bone mass, osteoporosis and fractures, including the loss of functional independence and mortality they cause. Furthermore, this line of research offers new treatment options for bone health. This research will ultimately inform clinical recommendations for night shift workers and introduce a paradigm shift in the prevention, evaluation and treatment of osteoporosis.

NIH Research Projects · FY 2025 · 2021-07

Project Summary/Abstract In the U.S., 8.37 million adults over age 65 will experience a hospital stay during the next year, which often has serious and long-lasting consequences including profound deterioration in physical function. Following a hospital stay, around 1.35 million patients with deconditioning require rehabilitation in a skilled nursing facility (SNF) each year to address the deleterious musculoskeletal and functional deficits from deconditioning. Unfortunately, current rehabilitation paradigms in SNFs do not adequately restore physical function, which directly contributes to poor community discharge rates. Strikingly, only 52% of all patients admitted to SNFs are discharged to a community setting (e.g., home), which suggests a paradigm shift is required to optimize rehabilitation within SNFs. Currently, usual care rehabilitation in SNFs consists of low-intensity rehabilitation interventions, which are physiologically inadequate to induce meaningful changes in skeletal muscle strength and physical function. To address these pitfalls, a high-intensity resistance rehabilitation paradigm has been shown to improve outcomes including better physical function, increased community discharge rates, and cost-effective reductions in length of stay. The proposed pragmatic study seeks to apply this rehabilitation paradigm to multiple SNFs to further evaluate the effectiveness of high-intensity resistance rehabilitation (Aim 1), while evaluating processes, mechanisms, and determinants of successful implementation (Aim 2). We propose a cluster randomized pragmatic trial design in which a high-intensity intervention (15 SNFs) will be compared to usual care (15 SNFs). Effectiveness in terms of physical function will be determined via change in patient gait speed (primary outcome) from admission to discharge. Implementation strategies will be evaluated by reach (proportion of patients treated with the intervention), adoption (proportion of therapists appropriately adopting the intervention), implementation (fidelity assessments), and maintenance (long-term fidelity assessments) of the intervention. This study will provide the first large-scale evaluation of high- intensity rehabilitation for patients admitted to SNFs following hospitalization. Additionally, through systematic comparison and in-depth analysis of implementation across a variety of SNFs, this study will provide critical insight regarding barriers and facilitators of implementation. Overall findings from this study have the potential to 1) shift SNF rehabilitation care paradigms; 2) optimize patient outcomes and independence and 3) critically inform future work aimed at wide-scale implementation of high-intensity rehabilitation across post-acute settings.

NIH Research Projects · FY 2025 · 2021-07

Abstract Human craniofacial development is a complex process and frequently goes awry to cause a major class of birth defects, orofacial clefting, which affects approximately 1 in 700 live births. Proper facial development in mouse and human requires three sets of paired facial prominences coming together by growth, morphogenesis, and fusion. Embryonic facial development is strikingly similar in human and mouse, making the mouse the best available model system for human. Previous studies have shown that the expression of many thousands of genes changes across tissue layer, age, and/or prominence, as well as cell population during early mouse facial development. However, we still only have a rudimentary understanding of how these changes are regulated by the interaction of transcriptional modulators in the developing face. To understand how genes are transcriptionally regulated during facial development, this research seeks to construct transcriptional regulatory networks in a temporospatial manner by in silico analysis of publicly available multi- omic datasets. Aim 1 will focus on the identification and verification of transcriptional regulatory networks operating in facial mesenchyme with a focus on super-enhancers. Aim 2 will adopt a similar approach to study the ectoderm which acts as a vital signaling center for the mesenchyme. Finally, in Aim 3 I will apply knowledge from Aims 1 and 2 to build transcriptional regulatory networks at the single cell level. These aims will take advantage of available RNA-seq, ATAC-seq, histone marker ChIP-seq, transcription factor ChIP-seq, bulk and single cell RNA-seq data from wild-type or mutant mice, as well as facial enhancer expression databases. Accomplishment of these studies will predict how genes are transcriptionally regulated in a temporospatial manner during facial development and discover sets of core transcription factors and super- enhancers controlling facial development. These transcriptional regulatory networks will be relevant to the genetic and molecular underpinnings of human orofacial clefting, and will provide clear testable predictions about transcription factor function and the consequences of aberrant expression. Performance and accomplishment of these Aims will also act as a major component of my career development plan, in which my goal is to obtain and independent tenure-track faculty position and serve as a mentor to the next generation of scientists. A major aspect of my career development plan is to build on my growing strength in bioinformatics by learning more advanced techniques in this specialty alongside new computational based approaches, such as machine learning. In this respect, my Aims and career development plan are aligned with a Notice of Special Interest (NOSI) of NIDCR in Supporting Dental, Oral, and Craniofacial Research Using Bioinformatic, Computational, and Data Science Approaches (NOT-DE-20-006) for which this application is targeted. I have recruited a mentorship team with specialties in craniofacial biology, bioinformatics, machine learning, and career development to help me achieve these goals.

NIH Research Projects · FY 2025 · 2021-07

PROJECT SUMMARY / ABSTRACT The advent of genome editing and induced pluripotent stem cell technologies now provide hope for creating therapies to treat and cure many developmental diseases. However, incomplete knowledge of the genetic regulatory mechanisms that guide cell differentiation and function stands as a significant barrier to progress. Overcoming this barrier will require a workforce with a training foundation that integrates a strong base in genetics, a deep knowledge of developmental, cellular and molecular mechanisms, and the ability to apply basic concepts to the translational potential of stem cell biology. The Genetics of Development, Disease, and Regeneration Training Program seeks to foster a broad foundation of interdisciplinary knowledge and career development that empowers a diverse population of students to apply their training to the study and treatment of developmental diseases. To achieve this goal, the GDDR Program is grounded in five training fundamentals: 1) knowledge base, 2) research skills and innovative technology, 3) hypothesis-driven research, 4) communication skills and 5) professional and career development. The GDDR Program will create a unique training environment by combining the broad and interdisciplinary research interests of our faculty with a training plan that integrates genetic principles and mechanisms into the study of development and stem cell biology. Thirty well-funded and accomplished Training Faculty drawn from 10 basic science and clinical departments create a highly collaborative and interdisciplinary environment. The breadth of our training environment will provide trainees with a diverse menu of labs for thesis training, while maintaining a supportive community and facilitating beneficial interactions between the GDDR Program and other T32-funded programs at the University of Colorado Anschutz Medical Campus. We will take full advantage of these interactions to develop courses and training activities that involve multiple programs. Thus we will broaden the exposure of our trainees to different knowledge, technologies, and career opportunities, and extend the impact of our training program to benefit a larger population of students and researchers at CU AMC. Based on the quality and size of our student pool and Training Faculty, we propose to support six trainees per year, which will permit us to be highly selective while building an interactive group of trainees. Our trainee recruitment plans are structured to attract highly talented students from diverse backgrounds, thereby contributing to a rich setting for training and accomplishment.

NIH Research Projects · FY 2025 · 2021-07

Prostate cancer is characterized by large genomic rearrangements and deletions. We show that the genes CHD1 and MAP3K7 are co-deleted in ERG translocation negative prostate cancer. To demonstrate a functional cooperativity we used a novel mouse prostate stem cell developmental model and showed that collaborative loss of CHD1 and MAP3K7 promotes an aggressive prostate cancer phenotype with altered lineage differentiation, abnormal secretory products, massive nuclear atypia, loss of E-cadherin and enrichment in neuronal and neuroendocrine markers. Profound alterations in AR expression were also observed. Using multiple human cell line models we also demonstrate that loss of CHD1 and MAP3K7 promotes castrate-resistant prostate cancer. This project will evaluate downstream targets of AR altered in tumors with loss of MAP3K7 and CHD1 using functional genomics in vitro and in animal models and assess the clinical impact of these targets on patient outcome. This work could have impact on the management of the most aggressive prostate cancer. Co-deletion of CHD1 and MAP3K7 occurs in 10-15 % of primary tumors. Relapse occurs in approximately 50% of patients with co-deletion. If these deletions occur in primary tumors and predict poor survival, men could be stratified based on MAP3K7 and CHD1 status. A functional understanding of this variant of prostate cancer could lead to novel therapeutic targeting strategies in the future.

NIH Research Projects · FY 2025 · 2021-07

Project Summary Autosomal dominant polycystic kidney disease (ADPKD) is a common inherited disorder that leads to end- stage kidney disease. To date, tolvaptan is the only approved intervention to slow kidney disease progression in patients with ADPKD. However, tolvaptan is constrained by high cost and common side effects that limit adherence and is only indicated for rapidly progressing ADPKD. Thus, alternative or concurrent interventions that may slow progression of ADPKD are of considerable clinical importance. Similar to the general population, body-mass index has been increasing in patients with ADPKD, and approximately nearly 70% of adults with ADPKD are overweight or obese. Adipocytes do not simply act as a fat reservoir, but are active endocrine organs, and thus, may be a promising clinical target for ADPKD management. Mounting evidence also suggests that a metabolic defect exists in ADPKD, which likely contributes to cystic epithelial proliferation and subsequent cyst growth. Mild-to-moderate food restriction profoundly slows cyst growth and maintains renal function in numerous rodent models of PKD via mechanisms including activation of AMP-activated kinase and suppression of mammalian target of rapamycin-S6 kinase signaling and insulin-like growth factor-1 levels. Additionally, we have shown that overweight and obesity are strong independent predictors of more rapid kidney growth, measured by total kidney volume (TKV). We recently completed a R03-funded pilot study supporting that a behavioral weight loss intervention via daily caloric restriction (DCR) in adults with ADPKD and overweight or obesity: 1) is feasible and acceptable; 2) slowed kidney growth (annual %∆ in height- adjusted TKV [htTKV]); 3) reduced abdominal adiposity; and 4) altered markers of biological pathways implicated in ADPKD progression and metabolism. However, our pilot and feasibility study was limited by a small sample size, relatively short duration, and lack of a control group. Thus, to translate these promising results of our pilot study towards clinical practice, we propose a parallel-group, randomized, controlled clinical trial in 126 adults with ADPKD and overweight or obesity to directly compare the efficacy of behavioral weight loss intervention based on DCR vs. control (standard clinical advice for ADPKD) for slowing kidney growth over a longer duration. Changes in abdominal adiposity will serve as a secondary outcome. Effects of weight loss on circulating and adipose markers of biological pathways will provide mechanistic insight. Specific Aim 1: Determine the effect of a DCR-based behavioral weight loss intervention on kidney growth (annual %∆ htTKV by MRI over 24 months) vs. control (standard clinical dietary advice for ADPKD). Specific Aim 2: Quantify changes in abdominal adiposity (visceral, subcutaneous, and total) by MRI in each group and their association with changes in htTKV and markers of biological pathways. Specific Aim 3: Measure changes in makers of biological pathways in blood and adipose tissue. Specific Aim 4: Further evaluate the safety of DCR in ADPKD vs. control, to optimize clinical translation.