Northwestern University

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY Family formation plans (e.g., whether/when to have children) vary by micro-level factors like sociodemographics (age, race/ethnicity) and individual- and couple-level influences like finances and job security. These micro-level influences do not fully account for variations in reproductive life plans, which are also influenced by macro-level factors like state and local laws and structural inequities. Sexual and gender minority (SGM) people's health and wellbeing may be particularly affected by these macro-level factors. Our research and that of others demonstrate that structural stigma (macro-social conditions like anti-LGBT legislation that negatively impact wellbeing) worsens SGM people's mental and physical health. Given current high levels of structural stigma in the US, there is an urgent need to understand the potential health and wellbeing impacts to halt the widening of already gaping health disparities. One aspect of health and wellbeing potentially impacted by anti-LGBT legislation and other forms of structural stigma is having children. Almost 40% of SGM people are interested in parenting but may face unique barriers in doing so. Being unable to freely plan when, whether, and how to have children excludes SGM people from a health-promoting life stage, which may drive some SGM-related health disparities (e.g., alcohol use, cardiovascular health, depression). The proposed mixed-methods study will prospectively examine multi-level impacts on SGM couples' family formation plans. Aim 1. Quantitatively test multi-level influences on family formation plans through a large-scale survey of SGM couples. We hypothesize that couples living in states with higher levels of structural stigma will be less likely to plan to have children and more likely to report that state-level policies influence their decision- making. These associations will be moderated by the racial/ethnic and gender composition of the couple, as well as by SES. Aim 2. Qualitatively describe multi-level factors influencing SGM couples' family formation planning. From the Aim 1 sample, we will recruit couples from states with high and low structural stigma for in- depth dyadic interviews (N=120). We will describe multi-level influences on family formation planning. Aim 3. Given the dynamic sociopolitical landscape for marginalized populations, we will quantify changes in impacts of multi-level factors on family formation plans and well-being over time. We will follow the Aim 2 subsample with dyadic pulse surveys (i.e., brief/regular surveys across three years) and a final in-depth dyadic interview. Using qualitative trajectory methods and prospective analyses of dyadic pulse surveys, we will test our hypotheses that changes (or lack thereof) in couples' contexts (e.g., moving to another state, legislation changes) will influence decision-making, mental health, relationship quality, and family formation. This project will produce the first investigation of how multi-level factors prospectively influence SGM couples' family formation plans. This work will have a positive impact by revealing for the first time the multi-level needs of LGBTQ couples as they form families, thereby informing interventions to reduce entrenched inequities.

NIH Research Projects · FY 2025 · 2024-09

Project Abstract Whereas the contribution of tumor microenvironment to the profound immune suppression of glioblastoma (GBM) is clear, tumor-cell intrinsic mechanisms that regulate resistance to CD8 T cell mediated killing are less studied. We performed an in vivo CRISPR screen to identify glioma intrinsic kinases that contribute to evasion of tumor cells from CD8 T cell recognition. The screen revealed checkpoint kinase 2 (Chek2) to be the most important kinase contributing to escape from CD8 T-cell recognition. Genetic depletion or pharmacological inhibition of Chek2 with blood-brain-barrier permeable drugs that are in the clinical trials, in combination with PD-1/PD-L1 blockade, led to survival benefit in preclinical glioma models. Furthermore, our experiment using ex vivo gliomas showed that glioma tumor cell intrinsic Chek2 interacts with Y-Box Binding Protein 1 and 3 (YBX1&3). The YBX1&3 are DNA/RNA binding proteins implicated in transcriptional regulation of immune-modulatory genes. However, the mechanism underlying the immunosuppressive function of Chek2 in gliomas is unknown. Therefore, in this proposal, we propose to determine the Chek2-YBX1&3 interaction mediated transcriptional changes that modulate CD8 T cell response using chromatin immunoprecipitation sequencing and CD8 T cell -mediated tumor cell killing assay (Aim 1). As an independent mechanism, we will investigate the contribution of Chek2 inhibition/depletion dependent STING pathway activation in enhancing CD8 T cell response by inhibiting STING pathway and studying its impact on CD8 T cell proliferation using OT- 1 CD8 T cell assay (Aim 2). As a potential novel therapeutic approach, we will test if targeting YBX1 enhances response to PD-1 blockade by determining the effect of this treatment strategy on the survival of preclinical glioma models (Aim 3). Together, this project will elucidate the mechanism underlying the immunosuppressive function of Chek2 and will provide a novel therapeutic strategy combining YBX1 inhibition, Chek2 inhibition and PD-1 blockade for testing in GBM patients.

NIH Research Projects · FY 2024 · 2024-09

Cardiovascular disease has emerged as a significant contributor to cognitive impairment, operating both independently and as a catalyst for the progression of Alzheimer’s disease (AD) from low-grade to overt dementia. Amongst all vascular diseases, atherosclerosis stands out because of its prevalence as a common comorbidity in AD patients. Atherosclerosis impairs vascular brain function at multiple levels. First, by directly impacting the circle of Willis and carotid arteries, atherosclerosis leads to narrowing of the vascular lumen, reducing blood flow for the supported regions. Second, rupture of atherosclerotic plaques often leads to thrombosis, resulting in either vessel occlusion or thromboembolisms. Depending on the size of the embolus, it may cause lesions that range from “silent” infarcts or microinfarcts to large cerebral infarcts with overt clinical symptoms. Third, independent of plaques, its associated hyperlipidemia impairs endothelial and smooth muscle cell function in both systemic and brain vessels. Our investigations into the brain vasculature of mice at the onset of atherosclerosis revealed distinctive vulnerabilities and increased vascular pathology in those with AD amyloid pathology. At the cellular level, perivascular macrophages exhibited prominent alterations in density, distribution, and molecular profile. We also observed dysfunctional permeability, specifically affecting the cortex and hippocampus. Deficiencies in vascular contractility and glymphatic flow were more pronounced in mouse models of AD compared to similarly aged mice without AD pathology. Overall, the findings underscore the impact of atherosclerosis disease on the brain’s microvasculature and provide the underlying impetus to focus on the small vessel disease in response to metabolic lipid deregulation. Our underlying hypothesis posits that hyperlipidemia contributes to vascular dysfunction in both additive and synergistic manners to existing AD pathology. To delve deeper, we propose a comprehensive analysis encompassing structural, molecular, and functional aspects of the vasculature in the context of AD, utilizing two disease models of AD amyloidosis. Crucially, we will evaluate the secretome of endothelial, smooth muscle, pericytes and perivascular macrophages, capturing both intracellular and serum- released proteins as potential cell-specific biomarkers for disease progression. Furthermore, we aim to investigate whether correcting hyperlipidemia, either pharmacologically or by modifying the diet and reversing low-density lipoprotein receptor levels, can restore observed alterations. Correlating these interventions with AD severity will provide important insights into potential therapeutic avenues. In tandem, we plan to develop an integrated computational network that overlays multiple datasets in a spatiotemporal manner, establishing connections with AD pathology. This holistic approach seeks to deepen our understanding of the intricate relationships between vascular health and AD, aiming to inform targeted interventions and therapeutic strategies.

NIH Research Projects · FY 2024 · 2024-09

More than 300,000 surgical or endovascular lower extremity revascularization procedures are performed annually for Medicare beneficiaries who have lower extremity peripheral artery disease (PAD) without limb threatening ischemia, and rates of these procedures are increasing. In these patients, revascularization typically improves, but does not eliminate, PAD related walking impairment. Lower extremity revascularization combined with supervised exercise significantly improves walking performance compared to revascularization alone in people who have PAD without limb threatening ischemia. However, supervised exercise is inaccessible or burdensome for most PAD patients. We hypothesize that home-based exercise combined with lower extremity revascularization will significantly improve walking performance compared to revascularization alone in patients with PAD undergoing revascularization for disabling PAD. We further hypothesize that inorganic nitrate, a major source of nitric oxide (NO) abundant in beetroot juice, will improve walking performance after lower extremity revascularization, compared to placebo. In preclinical models, NO inhibits inflammation, neointimal hyperplasia, thrombosis, and vascular smooth muscle cell migration at sites of revascularization. NO increases angiogenesis and perfusion, repairs skeletal muscle damaged by ischemia, and stimulates mitochondrial activity. A large body of evidence, assembled over more than 25 years, supports our hypothesis that inorganic nitrate (such as that in nitrate-rich beetroot juice) will improve walking performance and other lower extremity outcomes after lower extremity revascularization. Effective, accessible, and safe therapies are needed to enhance the benefits and improve durability of lower extremity revascularization. Therefore, in a randomized clinical trial with a 2 x 2 factorial design, we will test the following two primary hypotheses in 386 patients randomized within three months of a successful lower extremity revascularization for disabling PAD: First, that home-based exercise combined with lower extremity revascularization will improve six-minute walk distance more than revascularization alone at 6-month follow-up (Primary Aim #1). Second, that nitrate-rich beetroot juice combined with lower extremity revascularization will improve six-minute walk, compared to placebo combined with revascularization at 6-month follow-up (Primary Aim #2). In Secondary Aims, we will establish whether home-based exercise, compared to control, and whether nitrate-rich beetroot juice, compared to placebo, improve the Walking Impairment Questionnaire and the PROMIS Mobility Questionnaire and prevent hemodynamic failure of the revascularized vessel. In exploratory aims, we will test the effects of the interventions on plasma nitric oxide bioavailability and on gastrocnemius muscle biopsy measures of capillary density, myofiber size, and mitochondrial activity. If our hypotheses are correct, results of this trial will have a major impact on improving mobility in the large and growing number of people undergoing revascularization for disabling PAD.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY Asian Americans are one of the fastest growing ethnic groups in the United States, yet they remain underrepresented in discussions about the utility of genetics and genomics and how this technology should or should not be implemented into clinical care. One condition, Asian Alcohol Flushing Syndrome, affects over 25% of East Asians and is caused by a genetic variant, ALDH2*2. ALDH2*2 also has links to serious medical conditions such as stroke, myocardial infarction, and Alzheimer’s Disease, and it affects individuals’ alcohol metabolism, making alcohol consumption more toxic and cancer causing. Lifestyle changes, such as reducing alcohol consumption, can reduce the risk of esophageal cancer for East Asian Americans with this variant. However, we lack information about East Asian American community members’ perceptions of genetic testing for ALDH2*2 and how to implement this testing into clinical care. The goal of the proposed research is to engage an underrepresented community in the implementation of genomic testing that has direct relevance to their health outcomes. I will accomplish this goal by evaluating the effect of ALDH2 genetic testing on the drinking behaviors of East Asian Americans and assessing clinician and patient opinions of moving this test from direct-to-consumer to clinical practice. This proposal has three specific aims. Aim One: To evaluate East Asian American community members’ perceptions of personal utility for ALDH2*2 genetic testing and preferences for screening. Aim Two: To identify barriers and facilitators to implementation of ALDH2*2 genetic testing in primary care. Aim Three: To measure the effect of direct-to- consumer ALDH2*2 genetic testing and education on health behaviors of East Asian Americans who flush when they drink alcohol. The proposed research will provide community-driven recommendations for implementing ALDH2*2 genetic testing in primary care. Dr. Young will achieve these aims by drawing on her current skills in qualitative research and genetics, as well as additional training in community-engaged research, pragmatic clinical trials, and implementation science. These research and training goals are to be carried out at Northwestern University in conjunction with a Community Advisory Board of East Asian Americans who flush when they drink alcohol and an experienced team of mentors. Dr. Young is already an accomplished scholar with a track record of high-quality research. The outlined training plan will enable Dr. Young to accomplish her long-term career goal to become an independent, interdisciplinary researcher implementing ethical and equitable genetic testing interventions for underserved communities.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY/ABSTRACT Delayed Sleep Wake Phase Disorder (DSWPD) is a common circadian rhythm sleep-wake disorder that is characterized by difficulty in initiating sleep at night and subsequent difficulty in waking at times required for work or school. Consequently, DSWPD is often associated with excessive daytime sleepiness, reduced academic and work performance, mood disturbances and reduced quality of life. Considering the significant negative impact of DSWPD on sleep quality, performance, mental and physical health and well-being, the physiological basis of the disorder surprisingly remains unclear. Classically, DSWPD is assumed to stem from a delay of central circadian timing. However, recent evidence shows that those clinically diagnosed with DSWPD fall into at least two distinct phenotypes: those with delayed circadian timing and sleep (DSWPD1) and those with typical circadian timing but delayed sleep (DSWPD2). Using pupillometry, our group has shown differences in light sensitivity between the DSWPD1 and DSWPD2 subtypes, as well as within the DSWPD1 group (high (HLS) and low sensitivity to light (LLS)), indicating that clinically diagnosed DSWPD likely represents a complex interplay between underlying physiological differences and behaviors that can affect sleep timing. However, current diagnostic criteria do not consider these distinct phenotypes, which can limit diagnostic accuracy and hinder the development of personalized treatments that target the underlying alterations in physiology and behavior of these DSWPD subtypes. The overall goal of the proposed research is to identify the physiological basis of DSWPD subtypes, and to use these findings to inform development of mechanistically based approaches in circadian medicine. This project will enroll 220 patients with DSWPD who will undergo circadian physiological and behavioral phenotyping by actigraphy, dim light melatonin sampling, pupillary light sensitivity, as well as assessment of mood, alertness and meal timing using questionnaires. In addition, we will evaluate light sensitivity in LLS DSWPD1, as measured by the suppression of melatonin, and test whether the SSRI citalopram can enhance light sensitivity. Finally, mathematical modeling will be used to integrate physiological (pupillometry) and behavioral assessments to identify DSWPD subtypes. The proposed research will form the basis for the development of light sensitivity-based tools and algorithms that integrate physiological and behavioral biomarkers which can lead to a personalized multimodal approach for the care of patients with circadian rhythm disorders.

NIH Research Projects · FY 2025 · 2024-09

Lung transplantation is increasingly used to treat an expanding list of end-stage lung diseases, resulting in an over >50% increase in the number of lung transplant procedures in the US in the last decade. Unfortunately, survival following lung transplant is the worst amongst solid organs with only 80% and 50% of patients alive at 1 and 5 years, respectively. Primary graft dysfunction (PGD) affects over 50% of recipients within 24 hours of transplantation and has emerged as the most important risk factor for both short-term mortality and long-term graft loss from chronic lung allograft dysfunction (CLAD). The investigators in this PPG have made important contributions to a growing molecular understanding of the complex interplay between immune and lung parenchymal cells that underlie PGD and CLAD. In Project 1, we dissect molecular mechanisms underlying our discovery that lung restricted alloantibodies (LRA), present in over 30% of lung transplant recipients, are associated with PGD. In Project 2, we dissect physiologic and molecular mechanisms underlying prevalent abnormalities in esophageal function that lead to acid aspiration and worsen CLAD severity. In Project 3, we will credential profibrotic MoAMs as causal drivers of lung fibrosis in murine models and in patients with CLAD. In Project 4, ESI, we test whether mitochondrial dysfunction in the alveolar epithelium predisposes lung transplant recipients to pathologic activation of the integrated stress response (ISR) that precludes lung repair. Together our published and preliminary data support our overarching hypothesis that acute neutrophil-mediated lung injury mechanisms such as PGD and acid aspiration drive CLAD progression by promoting the development of LRA, recruiting profibrotic MoAM, and inducing epithelial cell mitochondrial damage causing an ISR-mediated barrier to epithelial repair. We will test this hypothesis in 4 interrelated and synergistic projects. Project 1. To determine whether LRA interact with donor derived NCM and recipient CM to worsen PGD via the activation of complement dependent and independent pathways. Project 2. To determine whether physiologic abnormalities in the esophagus after lung transplant cause gastric aspiration that induces LRA and CLAD progression via neutrophils and MoAM. Project 3. To determine whether CSF1 drives the recruitment and retention of profibrotic MoAM and whether their detection in bronchoalveolar lavage fluid can serve as a biomarker for CLAD. Project 4. To determine whether mitochondrial dysfunction in the airway and alveolar epithelium results in pathologic activation of the ISR that precludes normal epithelial repair to promote CLAD. The Human and Clinical Phenotyping Core (Core B), Mouse and Cell Phenotyping Core (Core C) and Administrative Core (Core A) will support these synergistic projects using murine models and clinical and molecular data collected from lung transplant recipients to identify actionable pathways that can be therapeutically targeted to improve lung transplant outcomes.

NIH Research Projects · FY 2025 · 2024-09

More than 300,000 surgical or endovascular lower extremity revascularization procedures are performed annually for Medicare beneficiaries who have lower extremity peripheral artery disease (PAD) without limb threatening ischemia, and rates of these procedures are increasing. In these patients, revascularization typically improves, but does not eliminate, PAD related walking impairment. Lower extremity revascularization combined with supervised exercise significantly improves walking performance compared to revascularization alone in people who have PAD without limb threatening ischemia. However, supervised exercise is inaccessible or burdensome for most PAD patients. We hypothesize that home-based exercise combined with lower extremity revascularization will significantly improve walking performance compared to revascularization alone in patients with PAD undergoing revascularization for disabling PAD. We further hypothesize that inorganic nitrate, a major source of nitric oxide (NO) abundant in beetroot juice, will improve walking performance after lower extremity revascularization, compared to placebo. In preclinical models, NO inhibits inflammation, neointimal hyperplasia, thrombosis, and vascular smooth muscle cell migration at sites of revascularization. NO increases angiogenesis and perfusion, repairs skeletal muscle damaged by ischemia, and stimulates mitochondrial activity. A large body of evidence, assembled over more than 25 years, supports our hypothesis that inorganic nitrate (such as that in nitrate-rich beetroot juice) will improve walking performance and other lower extremity outcomes after lower extremity revascularization. Effective, accessible, and safe therapies are needed to enhance the benefits and improve durability of lower extremity revascularization. Therefore, in a randomized clinical trial with a 2 x 2 factorial design, we will test the following two primary hypotheses in 386 patients randomized within three months of a successful lower extremity revascularization for disabling PAD: First, that home-based exercise combined with lower extremity revascularization will improve six-minute walk distance more than revascularization alone at 6-month follow-up (Primary Aim #1). Second, that nitrate-rich beetroot juice combined with lower extremity revascularization will improve six-minute walk, compared to placebo combined with revascularization at 6-month follow-up (Primary Aim #2). In Secondary Aims, we will establish whether home-based exercise, compared to control, and whether nitrate-rich beetroot juice, compared to placebo, improve the Walking Impairment Questionnaire and the PROMIS Mobility Questionnaire and prevent hemodynamic failure of the revascularized vessel. In exploratory aims, we will test the effects of the interventions on plasma nitric oxide bioavailability and on gastrocnemius muscle biopsy measures of capillary density, myofiber size, and mitochondrial activity. If our hypotheses are correct, results of this trial will have a major impact on improving mobility in the large and growing number of people undergoing revascularization for disabling PAD.

NIH Research Projects · FY 2025 · 2024-09

Project Summary: Dr. Akhil Chawla plays a pivotal role in the NCI clinical trials program at Northwestern University Robert H. Lurie Comprehensive Cancer Center (RHLCCC). As a Surgical Oncologist and clinical trial investigator, Dr. Chawla has demonstrated a long-term commitment to the support of NCI sponsored studies within at Northwestern and nationally through his service and leadership within the Alliance for Clinical Trials in Oncology (Alliance) and the NCI. Dr. Chawla leads the largest pancreatic cancer trial in North America, the Alliance A021806 Trial, for which he serves as a Principal Investigator and Co-Chair. Dr. Chawla has led protocol development, dissemination, implementation, and quality control for this practice-defining trial. As a leader in the NCI cooperative group space, Dr. Chawla has significant ambition towards the development of novel and practice changing trials. With his strong dedication to team science, Dr. Chawla has leveraged collaboration with experts in his disease space to refine and develop trial concepts he leads. As Dr. Chawla has shown unique success within NCI sponsored trial development at such an early point in his career, he has quickly become a leader at Northwestern. Dr. Chawla continues to be instrumental in RHLCCC NCI clinical trial activities, both as an investigator and supporter, including active involvement in protocol assessment, scientific review, and infrastructural expansion of the clinical trials program. With RHLCCC’s LAPS Grant designation, the NCI- designated cancer program has grown significantly with the expansion of NCI clinical trials into community sites as part of the Northwestern Medicine health system. Dr. Chawla’s clinical and investigative footprint spans from Northwestern University’s academic health campus in Chicago to community cancer sites throughout Chicagoland. He has leveraged these efforts to become a pioneer within RHLCCC and the health system as a known advocate for the dissemination of NCI sponsored clinical trials to geographically remote and underserved populations within the Northeastern Illinois. Dr. Chawla focuses on three major goals at RHLCCC: 1) lead integration of RHLCCC NCI clinical trials infrastructure across the Northwestern Medicine health system; 2) increase access and enrollment of racially and geographically diverse populations to NCI clinical trials; and 3) mentor and support junior faculty to participate in NCI-sponsored trial activity at RHLCCC and in NCI trial networks. Dr. Chawla is uniquely positioned at this point in his career to become a dedicated lifelong supporter and leader of NCI sponsored clinical trials within his institution and throughout the country.

NSF Awards · FY 2024 · 2024-09

This project aims to meet the national need to develop justice-oriented STEM teachers certified to teach computer science. Program participants will first earn a baccalaureate degree in a STEM major, then pursue teacher certification through Northwestern University's Master of Science in Education (MSED) program. Students will complete the requirements for an Illinois elementary or secondary teaching license in a STEM area, with a subsequent endorsement in computer science. Sixteen selected participants (two cohorts of eight each) will receive funding as Noyce Scholars for one year as MSED students. In addition to financial support, students will benefit from academic, networking, and social support through coursework to complete the MSED degree at Northwestern and two years of post-graduate teacher induction. The MSED program emphasizes social justice, culturally sustaining pedagogies, and context-specific teacher preparation. This project at Northwestern University leverages partnerships with Evanston/Skokie Community Consolidated School District 65, the McCormick School of Engineering and Applied Sciences, and the School of Education and Social Policy. Several goals will guide the project. First, the project team will recruit, prepare, and support sixteen STEM majors with baccalaureate degrees to certify them as K-12 teachers with a computer science endorsement. A second goal is to provide a locally immersive and context-specific approach to teacher preparation that enhances individuals' agency and critical analysis of social systems as the foundation of transformative education. The third goal is to produce teachers who understand computational thinking and how it contributes to learning within and across disciplines. Investigations through comprehensive mixed methods formative and summative project evaluations will provide assessment and feedback and generate new knowledge for best practices. Project investigators will disseminate challenges, successes, and findings through social media, peer-reviewed publications, and presentations at regional and national conferences. This Track 1: Scholarships and Stipends project is supported through the Robert Noyce Teacher Scholarship Program (Noyce) and is partially supported by funds from Micron Foundation. The Noyce program supports talented STEM undergraduate majors and professionals to become effective K-12 STEM teachers and experienced, exemplary K-12 teachers to become STEM master teachers in high-need school districts. It also supports research on the effectiveness and retention of K-12 STEM teachers in high-need school districts. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NSF Awards · FY 2024 · 2024-09

With the support of the Environmental Chemical Sciences (ECS) Program within the Division of Chemistry of the National Science Foundation (NSF), Professors Franz Geiger and Regan Thomson of Northwestern University and their students are investigating aspects of the fundamental chemical connections between plant-produced molecules and the Earth’s atmosphere. The unmistakable smell of a pine forest is produced by volatile chemical compounds made by trees. These chemicals, known as terpenes, are oxidized and transformed in the atmosphere, ultimately leading to the formation of particles and, in turn, clouds, which impact the global environment through processes such as rainfall and the reflection of sunlight. This project will explore the role that a particular chemical property of terpenes known as chirality, or "handedness" plays in the atmosphere. This project will investigate whether chirality affects the cloud-formation ability of the oxidation products. The project will provide interdisciplinary training to graduate students. In addition, educational outreach activities will target incarcerated students. The significance of chirality is well established in areas such as medicine where right or left-handed drugs often possess completely different medicinal properties. Chirality's role in atmospheric cloud formation remains largely unexplored. The research project will focus on the lab-based organic synthesis of chiral atmospherically relevant plant-derived terpenes in order to provide access to otherwise unavailable compounds for subsequent physical measurements. These measurements will probe key properties like surface tension, interfacial structure, and liquid-liquid phase separation (LLPS) to illuminate possible spontaneous resolution processes and higher-order hierarchical organization to discover new relationships between molecular-level organic structure and macroscopic atmospheric processes. Ultimately, the findings of the project will inform and update ongoing scientific efforts to better model and understand the Earth’s complex climate system. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2024-09

Project Summary Around 262,000 cysteine residues are intricately distributed throughout the human proteome. These cysteines play pivotal roles in influencing protein folding, stability, enzymatic functions, and the response to oxidative stress. Chemical probes designed to target cysteines are critical tools for investigating protein functionality and advancing drug development. Chemical proteomics, particularly cysteine-directed activity-based protein profiling (ABPP), has emerged as a fundamental technique for studying proteinaceous cysteines. ABPP employs a wide- spectrum cysteine-reactive probe to effectively pinpoint cysteine sites across a variety of proteins, enabling comprehensive functional exploration and facilitating ligand discovery. Traditionally, extracellular proteinaceous cysteines have been largely neglected in the field of chemical proteomics. It was also believed that these cysteines mainly formed disulfide bonds to stabilize protein structures. However, recent research has challenged this notion by revealing the dynamic nature of extracellular redox environments, which can significantly influence proteinaceous cysteine reactivity in the extracellular space. Consequently, a substantial knowledge gap remains in identifying extracellular proteins containing unmodified, reactive cysteines in specific biological contexts. To address this gap, this research program aims to establish an innovative chemical proteomic platform tailored for mapping extracellular proteinaceous cysteines across a wide range of biological scenarios. These scenarios include T cell activation, GGT5 genetic knockout, and various chemical treatments. Subsequently, we will investigate the potential druggability of these extracellular proteinaceous cysteines. Finally, utilizing the insights gained from targeting druggable extracellular proteinaceous cysteines, we aim to manipulate their functionality in pursuit of two key outcomes: inducing cell-cell interactions and promoting targeted protein degradation.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY/ABSTRACT We propose an innovative T32 postdoctoral training program, Enhancing TRanslational science to Accelerate Impact of New innovations (E-TRAIN) T32, for the Northwestern University Clinical and Translational Sciences (NUCATS) Institute. This 2-year program will catalyze the careers of postdoctoral scientists, expanding their domain expertise through mentored research, and develop foundational skills in cross-disciplinary team science to accelerate development, validation, and dissemination of new health innovations. Trainees will learn and apply study design principles for translational research that is generalizable across health conditions impacting the health of all Americans, aligned with the National Center for Advancing Translational Science goal of “bringing more treatments to all people more quickly.” E- TRAIN will emphasize reducing clinical and translational science (CTS) barriers in mentoring on strategies to advance biomedical innovations from discovery to implementation to long-term sustainability. We will train appointees to drive rapid, scalable, and sustainable translation from laboratory to clinical and community settings. Interdisciplinary co-mentoring and coaching will engage faculty with different scientific viewpoints from across Northwestern University and its clinical affiliates, providing an unparalleled experience in transdisciplinary team science. We will emphasize the development of effective communication skills to engage and inform academic, industry, and community audiences through interactive, experiential learning formats. Trainees will thus build foundational skills for future leadership in academia, industry, and communities, enriching the CTS workforce. Key E-TRAIN program elements are a) trainee-centric, holistic career development tools and supports; b) a novel co-mentoring approach that integrates implementation science at all stages of translational research; and c) multi-modal seminars and activities to attain essential knowledge, skills, and abilities across the translational science spectrum. A strong emphasis on work-life integration, wellness, and the development of reflective practices promotes resilience and retention in translational research. The program leaders have strong records in education and mentoring as well as complementary skill sets spanning pre-clinical development to implementation science.

NIH Research Projects · FY 2025 · 2024-09

Cellular-level Vascular Oculomics (CVO) for Monitoring Systemic Vascular Health . Planned activities and achievable goals: Year 1: (1.) Hardware design and implementation (Dubra/Burns) will be completed in the first 6 months to yield (2.) four fully functional devices in 4 clinical sites (Stanford, Indiana, Northwestern, Mount Sinai). Stanford will develop software to (3.) achieve real time image distortion correction. All sites will share existing datasets with Dr. Garyfallidis to initialize the (4.) training of deep learning AI-based software for real time biomarker annotation. (5.) At the end of this year, the entire team will meet with the advisory committee to present progress and get scientific feedback on progress and proposed biomarkers. Year 2: (1.) Stanford will develop and share image stabilization software and share with Indiana for (2.) integration in its image acquisition software. Garyfallidis will share the first version of the AI annotation algorithm with Burns and Dubra for integration into image acquisition software. At the end of this year, all sites will have the ability to image healthy and disease subjects using similar devices and software, allowing (2.) dataset comparisons and (3.) generation of normative CVO data. (4.) At the end of this year, the entire team will meet with the advisory committee to discuss progress, provide feedback, and identify areas for improvement and overall direction. Year 3: (1.) All sites will recruit subjects with systemic disease and (2.) share feedback to (3.) optimize AI software, (4.) software interface, and selection of (5.) biomarkers of interest. In a final group meeting with the advisory committee, the team will establish (6.) publication plan and (7.) software sharing platform. Final products: (1.) Novel set of ophthalmoscopes with Integrated hardware and software capable of real time segmentation and quantification of microscopic vascular structure and flow biomarkers, that is, the CVO instrument, and (2.) modular open-source software for diverse applications to imaging of retinal biomarkers, based on CVO, but adaptable to color photographs, OCT Angiography, Doppler OCT or other devices.

NIH Research Projects · FY 2025 · 2024-09

ABSTRACT There are currently 1.36 million melanoma survivors in the US, and this number is expected to increase to 1.92 million by 2030. Most melanoma patients (92%) survive 5 or more years but face an increased risk of new primary melanoma. Although survivors are recommended to practice sun protection, sunburns remain common among this population. The few sun protection interventions that target at-risk adults have had weak or mixed effects on sun protection, despite often comprising multiple components. Mobile health (mHealth) approaches to reducing unprotected sun exposure, which are highly acceptable among melanoma survivors, hold potential for delivering actionable real-time information on sun protection and exposure but have not been designed to account for sun protection in real time, and little is known about which mHealth components optimize effects on unprotected sun exposure. Further, there is growing evidence to suggest moderate-to-vigorous physical activity (MVPA) decreases after melanoma diagnosis, potentially due to heightened concerns about spending time outdoors (thus, decreasing opportunities for outdoor MVPA). Therefore, it is essential that sun protective interventions do not unintentionally reduce MVPA, a health-enhancing behavior. However, few studies have explored these relationships. Guided by the Multiphase Optimization Strategy (MOST) framework, the purpose of this study is to refine and enhance our existing intervention tools in preparation for a fully-powered factorial experiment to determine what intervention components optimally reduce unprotected sun exposure, and determine their effects on MVPA. We build on technological and methodological foundations of our prior work, including an alignment algorithm that combines information from an ultraviolet radiation (UVR) sensor, actigraphy, and our validated end-of-day Minutes of Unprotected Sun Exposure (MUSE) self-report survey to determine time spent outdoors, MVPA, and protection-adjusted UVR dose. In Aim 1, through user-centered design, we will refine intervention tools, including a SESAME app that integrates a UVR sensor and simplified MUSE survey to quickly assess real-time sun protection. In Aim 2, we will test the feasibility and acceptability of promising intervention components and research trial procedures in preparation for a future clinical trial that uses a highly efficient experimental strategy powered to detect effects of individual components on unprotected sun exposure and accelerometer-measured MVPA. Components include: in-the-moment UVR alerts and UVR feedback, sun protection monitoring and feedback, UVR weather reports, e-coaching, and action planning. Findings will inform a fully-powered MOST factorial experiment with components that are effective and acceptable by melanoma survivors. As an exploratory aim, we will evaluate the effects of each intervention component on MVPA. Knowledge gained from this study will inform development of more effective, implementable, and scalable interventions to reduce disease burden in this population.

NIH Research Projects · FY 2025 · 2024-09

Goal: This R34 study will pilot the Family Navigator Plus (FN+) program for foster parents to increase access to mental health (MH) services for children in foster care aged 6-17. Background: Youth in care experience disparities in mental health symptoms and access to treatment. Compared to the general child population, children in foster care are 4-5 times more likely to attempt suicide, and about 3 times more likely to have experienced trauma or have a diagnosed depressive or anxiety disorder. About half of children in foster care exhibit clinically significant MH needs, but less than a quarter of those children receive MH services. Those who do – experience delays in seeking treatment. Significance: This project will pilot the FN+ program to promote treatment engagement by building foster parent capacity to 1) recognize their child’s specific MH needs and address any preliminary barriers to treatment; 2) work with the child’s caseworker to engage the child in mental health services; 3) manage the child’s difficult behaviors; 4) remove remaining barriers to treatment access and use digital mental health tools during long waits for treatment, and 5) support treatment goals at home. Innovation: The proposed project is innovative in five ways. First, we plan to conduct the first pilot of a family navigator program in one of the highest-risk populations for developing psychiatric disorders: children in foster care aged 6 – 17 years. Second, we will pilot the use of a trained Community Health Worker in the family navigator role as a sustainable and cost- effective method to promote early MH treatment access. Third, we will provide rapid feedback regarding MH assessment results to promote foster parent understanding of child functioning and early treatment engagement. Fourth, we will use telehealth to facilitate family navigator access in areas with less access to support services. Fifth, we will help parents to find digital mental health tools to use during long waits for treatment. Design: Human-centered design and an open trial will inform a subsequent small randomized controlled clinical pilot to test the feasibility of the study protocol in preparation for a larger randomized controlled trial (RCT). Population: Foster parents of youth in care aged 6 to 17-years. Outcomes: All aspects of the study protocol (e.g., condition allocation, treatment and control condition procedures, data collection, etc.) will be operationalized in preparation for the subsequent RCT. The primary outcome is increased foster parent activation in the foster child’s mental health care. Secondary outcomes include reduced barriers to treatment access and perceived change in child behavior.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY/ABSTRACT Ovarian cancer is a diverse group of malignancies that can vary greatly in molecular biology, etiology, and presentation of symptoms. While it accounts for 2.5% of all cancers among women, it results in roughly 5% of cancer-related deaths due to its high fatality rate. This is because 75% of patients are diagnosed with advanced disease, largely attributed to its relatively late presentation of symptoms and a lack of reliable detection and monitoring strategies. Extracellular vesicles (EVs) are released by all cells, including ovarian cancer, and their cargo reflects their cells of origin. They have shown immense potential as stable biomarkers, however their low abundance compared to EVs from healthy cells and a lack of sufficiently sensitive characterization tools has limited their clinical translation. Surface-enhanced Raman spectroscopy (SERS) is sensitive enough to biochemically fingerprint even single EVs, and the information-rich spectra produced in EV SERS can be fed into machine learning (ML) algorithms to classify them based on their latent spectral features. Despite early progress in EV SERS, the highly heterogeneous nature of EVs indicates that their separation into distinct subpopulations prior to SERS analysis may help improve disease diagnosis, classification, and monitoring. Microfluidic devices are uniquely capable of separating EV s into subpopulations of interest while simultaneously enabling SERS spectral acquisition in a single device. In the proposed research, during the mentored phase, ML-enabled inverse design will be combined with high resolution nanofabrication techniques to improve the signal strength and spectral quality attainable from EV SERS. Preliminary data indicates that the SERS enhancement is highly dependent on the substrate’s nanoscale geometry, which is particularly important for EVs compared to conventional chemical analysis. Towards the end of the mentored phase, once the improved substrates have been thoroughly tested using bioreactor-produced EVs, they will be incorporated into two distinct microfluidic devices and tested throughout the independent phase using both cell culture and patient EV samples. One device will capture different subpopulations of EVs directly onto the microfluidic SERS substrates based on specific surface- antigens for multiplexed characterization, while the other will separate EVs precisely by size and flow them over the microfluidic SERS substrates to produce EV SERS barcodes. In parallel, ML algorithms tailored specifically to these platforms will also be developed to process and classify the acquired spectra. This proposal is multidisciplinary, utilizing advanced ML and micro-nanofabrication techniques as well as EV production, isolation, and characterization strategies. These experiments are innovative and significant because they will develop ML inverse design architectures specifically for EVs as well as microfluidic EV SERS for characterizing and classifying ovarian cancer.

NIH Research Projects · FY 2025 · 2024-09

Project Summary Despite over a century of research on the causative agent of syphilis Treponema pallidum, many questions associated with T. pallidum biology, and by extension syphilis pathology, remain unanswered due to the previous lack of an in vitro culture system. Syphilis, a quickly growing global public health concern, is a sexually transmitted, multiphasic disease with varied and devasting symptoms that can be fatal. As T. pallidum lacks many classical virulence factors (i.e., lipopolysaccharide, toxins, secretion systems) and has a paucity of outer membrane proteins, making it a stealth pathogen, it is unclear how T. pallidum causes syphilis. A growing body of work has established released peptidoglycan (PG) monomers (i.e., muropeptides) act as toxins and immune modulators. Many diderm bacteria have a highly conserved PG recycling pathway to transport muropeptides released during normal growth into the cytoplasm for reuse. When this pathway is disrupted, highly immunogenic muropeptides are released into the environment. Through bioinformatic analysis I discovered T. pallidum lacks this recycling pathway. Modifications to PG composition and architecture affect immune recognition and response, and our lab has identified unusual PG modifications in related pathogenic spirochetes, Borrelia burgdorferi (Lyme disease) and Treponema denticola (periodontal disease). Additionally, the release of unique muropeptides from B. burgdorferi during infection plays a causal role in the development of Lyme arthritis. Thus, this proposal aims to test the hypothesis that T. pallidum naturally secretes unusual muropeptides that drive syphilis pathogenesis. To test this hypothesis, I will first determine the biochemical features of T. pallidum PG as well as the fate and identity of released muropeptides via liquid chromatography tandem mass spectrometry (LCMS) and hNOD2 reporter assays. Second, I will elucidate the role of released PG in T. pallidum pathogenesis by creating a novel CRISPR platform to target T. pallidum lytic transglycosylases and testing the ability of CRISPR mutants to induce inflammation, induce chancre formation, and establish infection in vivo with a rabbit model. Collectively this proposal will increase our understanding of T. pallidum biology and syphilis pathology as well as create a new tool for the syphilis research field.

NIH Research Projects · FY 2025 · 2024-09

This Center strategically addresses a key developmental driver of the youth mental health crisis: Currently, 1 in 5 children have an identified mental health problem as early as age 3. This Mental Health, Earlier ALACRITY Implementation Science Research Center addresses the grand challenge of achieving population-level impact of early childhood mental health prevention efforts within routine pediatric care, a promising setting for achieving population impact. The Center is Northwestern based, including longstanding partnerships with Lurie Children’s Hospital, community health centers within Nemours Children’s Health and AllianceChicago’s practice-based research network, and University of Utah Health. The Center’s connective thread is a novel implementation framework, human-centered design methods, and a novel Roll-Out Implementation Optimization trial design, which systematically engages partners in strategy optimization. Center Aims: (1) Develop novel caregiver-focused, system-, and clinician-level strategies to advance implementation of developmentally- and ecologically-grounded early mental health screening and interventions in pediatric primary care; (2) Address the unique needs of varied communities and families via engagement in tailored evidence-based prevention services; and (3) Create a national resource for implementation of childhood mental health innovation in pediatric primary care, including fostering a cadre of scientific and workforce leaders and experts, and an open access toolkit. Center Signature Project (Peds-BRITE) and four Research Projects (RPs) innovate, implement, and integrated evidence-based interventions and strategies, with RPs testing novel approaches and/or levels of inquiry designed to address recognized, but understudied, barriers; Pediatricians Building Resilience through early Identification for Toddler Well-Being (Peds-BRITE): This cluster randomized trial implements an evidence-based toddler mental health risk decision tool with coordinated care to the Family Check-Up Online to support parenting and toddler self-regulation; Talking Early About Mental Health for Access (TEAM4Access): Deploys digital case-based training to improve clinician confidence and family-centered communication about toddler mental health risk for families from varied backgrounds and communities; Supporting Parent Action for Resilient Kids: A Single-Session Intervention to Boost Caregiver Adoption of Early Childhood Mental Health Services (SPARK): Adapts an evidence-based adjunctive digital single-session intervention to increase families’ mental health services use; Researching Ethics and Assessment of Unintended Consequences in Healthcare for Mental Health Earlier (REACH4MHE): Explores unintended consequences of early mental health prevention via an integrated bioethical-implementation framework. Community & Scientific Engagement & Administrative- and Methods Incubation Cores provide vision, coordination, methodologic and career development supports to advance implementation of early mental health promotion across a range of pediatric primary care systems and families.

NIH Research Projects · FY 2024 · 2024-09

PROJECT SUMMARY South Asian is one of the fastest growing ethnic groups in the United States (US). The striking overweight and obesity prevalence (78%) among foreign-born South Asian American adults is a public health concern, as it affects not only their own health, but transgenerational obesity risks for SA American youth. Physical inactivity among South Asian American adults partly explains their high obesity risk. Despite the potential of elevated physical inactivity and obesity risks among South Asian American children, few studies evaluated physical activity and obesity among South Asian American children at a national level. Further, cultural factors, such as gender norm, among South Asian American families may discourage South Asian female youth to participate in physical activity. However, US data are lacking to understand how parents’ and child’s cultural factors influence the child’s physical activity behaviors among South Asian American families. To address these research gaps, the goals of this R03 project are to evaluate physical activity and body composition of South Asian American youth and to elucidate their cultural factors associated with physical activity behaviors. This R03 project proposes to conduct secondary data analyses to achieve two aims. Aim 1 is to evaluate physical activity and body composition among South Asian American youth using the 2011-Mar2020 National Health and Nutrition Examination Survey (NHANES) data. This evaluation will be conducted in comparison with other Asian (i.e., Chinese, Filipino) and racial/ethnic groups (i.e., Black, Hispanic, White). We will apply an innovative machine learning analytic method to estimate physical activity levels. Aim 2 is to examine the relationships between South Asian parents’ and daughters’ cultural factors and daughters’ physical activity using the baseline data of the South Asian Active Together (SAATH) clinical trial. This cost-efficient R03 project will produce high-impact knowledge, including a national evaluation of physical activity and body composition and the cultural determinants of physical activity behaviors among South Asian American youth. The outputs of the project will lay the foundation for designing an intervention for physical activity promotion of South Asian American youth. This project will also contribute to accelerating the adoption of the innovative machine learning analytic approach in physical activity research.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY Poor sleep impacts health in many ways, so improved sleep could yield wide-ranging public-health benefits. Sleep-disordered breathing constitutes a huge public-health problem and current treatments fail for many patients. Lapses in breathing can fragment sleep, disrupt oxygen levels, and cause other problems. Contemporary treatments for sleep disorders and recommendations for sleep hygiene are valuable, but more can be done to engender health-promoting sleep - this project will introduce new strategies for real-time modulation of sleep physiology that have not been applied or tested in this context. Basic memory research has shown that memory networks in the brain are active during sleep and that we can alter this activity. Variants of this method here won't aim to improve memory, as in our past studies, but rather to impact physiology to yield specific health benefits. Using soft sounds that avoid sleep disruption, with state-of-the-art monitoring devices applied in the home (nasal respiration, in-ear EEG, and chest EKG), we aim to change what people do during sleep - their mental and physical sleep habits. The moment one falls asleep, one's sleep physiology seems automatic and largely beyond control. Yet, our twofold premise is that (1) memories and habits naturally reactivate during sleep, and (2) we can intervene to strategically modify this reactivation to achieve specific health-focused goals, transcending basic memory research, as follows. We start with obstructive sleep apnea patients without structural upper-airway abnormalities. Poor neural control of breathing during sleep likely contributes to many such cases. Patients will first undergo extensive daytime training to elicit inhalation when a cue sound is played, such that responses become automatized. After training, patients continue emitting the conditioned response at night as they fall asleep and, remarkably, they continue to do so during sleep, thus reducing apnea symptoms. Lab studies testing the methodology will pave the way for at-home studies; this therapy is designed to be easy-to-use in the home via wearable tech and sound delivery configured to avoid arousal and target periods when respiration is insufficient (i.e., closedloop stimulation adjusted based on real-time respiration, movement, and EEG recorded wirelessly). Expansion to additional applications will be guided by evidence-based methodological insights and will fuel the development of a new understanding of high-quality sleep. For example, people with or without apnea may have affect-laden sleep that is maladaptive, as when unconscious rumination pervades sleep. We thus seek to bias overnight thinking in positive directions to enhance sleep quality; sound cues during sleep will displace anxious- or depressive-memory retrieval by promoting positive-memory retrieval. Given that suboptimal sleep is associated with poorer outcomes for mental and physical health, we envision a means to optimize sleep by correcting maladaptive muscular or affective activity, The high-risk challenge is to develop noninvasive, lowcost, precision-sleep-control interventions with adaptive protocols to produce better sleep at home.

NIH Research Projects · FY 2025 · 2024-09

Chicago is home to multiple populations experiencing low access to care and worsening cancer health outcomes. We propose to establish the Advancing Cancer Control Engaged Research Through Transformative Solutions in Patient Navigation (ACCERT PN) Center to develop community-engaged interventions, measures and methods for process and outcomes assessment, and researcher and community partner capacity and engagement centered around the theme of innovative multilevel patient navigation (PN) approaches and implementation science within learning health systems to address multifaceted barriers to cancer care, including social risks and contextual factors. With the goal of mitigating worsening cancer health outcomes in patient populations experiencing barriers to accessing care in Northwestern’s catchment area communities, the ACCERT PN Center’s research program will commence in the first year with the launch of a five-year Signature Research Project and two rapid and responsive, thematically linked Community Responsive Projects. The Signature Research Project will leverage innovations in systems engineering and artificial intelligence (AI) to develop and test two multilevel PN approaches via a Hybrid Type 2 randomized effectiveness-implementation trial. Community Responsive Project 1 will develop communication strategies and training resources for navigators and healthcare providers on supportive health services important to cancer survivors, via the Truth Talking Tour. Community Responsive Project 2 will explore the use of AI social robots in a Chicago community with low access to care to enhance digital literacy and cancer health literacy to inform the Signature Research Project. Additional Community Responsive Projects will be identified in subsequent years. Projects are supported by an Administrative Core that will oversee operational and scientific activities as well as community and internal and external advisory boards, and build capacity and engagement among scholars, including early career investigators in the biomedical and behavioral sciences, and community partners to implement ACCERT PN interventions that incorporate the lived experiences of those who face worsening cancer health outcomes. A Research Methods, Measures & Data Management Core will support the development, implementation, and evaluation of projects by developing measures and methods, and assessing community-level determinants of health, community engagement, and cancer control engagement processes and outcomes. This Center brings strong scientific rigor with significant potential to enact transformative solutions to advance cancer control engaged research, as demonstrated by our focus on scaling and transforming PN toward systems level care improvement to adequately close cancer care delivery gaps; a multidisciplinary community-academic research team with exceptional track records in cancer control, PN, community engagement, implementation science, artificial intelligence, human-computer interaction, systems engineering, community mobilization, and patient advocacy; and meaningful engagement with Chicago communities and multisectoral community partners with the guiding North Star of “nothing about us without us”.

NIH Research Projects · FY 2024 · 2024-09

A wide variety of environmental stressors, from pesticides to common anticancer chemotherapies, result in protein damage. This damage underlies diseases such as cancer and neurodegeneration, and when caused by prescription drugs, leads to toxic side effects limiting usage. To cope with these proteotoxic stresses, cells express molecular chaperones, co-chaperones, and a variety of additional support proteins – collectively referred to as the proteostasis network (PN) – that are integral for the proper synthesis, folding, stability and degradation of the cellular proteome. Thus, understanding the regulation and function of the PN is critically important in understanding its role in the interface between disease susceptibility and exposure to environmental stressors. We previously developed a broadly applicable framework and online resource (FIREWORKS) to interrogate biological networks using unbiased genetic screens. Using our FIREWORKS framework, we mapped the constituents of and relationships between stress response pathways in human cells coping with growth-related molecular stresses revealing a network of 146 genes with vital functions spanning diverse stress contexts. Here, we propose to expand this resource to create an unbiased and global map of the response to environmental proteotoxic stress. We will integrate our cutting-edge chemical-genetic screening platform with our FIREWORKS computational pipeline to identify components, function, and regulatory nodes critical for the cellular response to environmental stressors. In parallel, we will identify transcriptional changes that occur immediately and throughout longer periods of environmental stress exposure, to provide a framework for understanding cellular adaptation to these stressors. Collectively, this map will serve as a major resource for the research community, provide a more complete understanding of cellular environmental stress response programs, and lead to insights into the large variability in the risk of adverse events that arise from exposure to these insults.

NIH Research Projects · FY 2025 · 2024-09

Project Summary Increased evidence started to show that many germline mutations predispose individuals to myeloid neoplasms, including myelodysplastic syndromes (MDS). One of these mutations occurs on DEAD-box helicase 41 (DDX41) gene. More than 80 distinct DDX41 variants have been reported, making DDX41 one of the most frequently mutated MDS predisposition genes. Germline DDX41 mutations lead to its loss of function, whereas somatic mutations often produce hypomorphic changes. Our understanding of the roles of DDX41 in the hematopoietic system and how DDX41 mutations predispose myeloid neoplasms is still in its infancy. It is essential to dissect the molecular and cellular mechanisms of DDX41’s functions, which is critical to developing targeted therapy for DDX41 mutated MDS. G-quadruplexes (G4) are noncanonical secondary nucleic acid structures formed in guanine-rich sequences. DDX family proteins have been shown to resolve DNA G4 structures. We found in our preliminary study that loss of DDX41 led to aberrant upregulation of G4 in the hematopoietic cells, especially the erythroid lineage. Aberrant G4 accumulation due to DDX41 deficiency severely disrupted erythropoiesis with fewer effects in other lineages in vitro and ex vivo. Importantly, we confirmed these findings in vivo using lineage- specific Ddx41 knockout (KO) mice, which showed that the lethality of hematopoietic-specific Ddx41 KO mice is likely due to the defects in erythropoiesis. Our preliminary mechanistic studies indicate that DDX41 directly binds to and dissolves G4. Ddx41 deficiency-mediated G4 upregulation compromised erythroid genomic integrity and ribosomal biogenesis, which upregulates p53 and activates the cGAS-STING pathway. Our additional in vivo preliminary experiments reveal that the knockout of cGas, but not p53, rescued the lethality of hematopoietic- specific Ddx41 KO mice. Based on these data, we hypothesize that DDX41 loss of function in MDS causes aberrant G4 accumulation, which leads to genomic instability and cGAS-mediated cell death predominantly in erythroid cells. To test our hypothesis, we propose three specific aims. Aim 1 will focus on the study of the functions of DDX41 in various lineages of hematopoietic cells by generating lineage-specific Ddx41 knockout mice. In Aim 2, we will investigate the functions of DDX41 in G4 homeostasis in erythropoiesis. Aim 3 will focus on the mechanism of DDX41 deficiency-induced ineffective erythropoiesis and MDS pathogenesis. Successful completion of our proposed research will provide novel insights into DDX41’s functional role in normal and abnormal hematopoiesis, which will be impactful for the development of novel therapies for DDX41 mutation- related hematologic diseases.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY Mustard gas causes severe epithelial and deep tissue injury resulting in blisters, pain, delayed wound healing, and scarring. The complex mechanism of tissue damage following nitrogen mustard (NM) and sulfur mustard (SM) exposure involves an initial phase of rapid damage. This phase is characterized by acute damage to epidermal keratinocytes and to the basement membrane attaching the top layers of the skin with the underlying dermis. This initiates a cascade of reactions including the release of inflammatory cytokines that signal for the migration of immune cells into the skin from the circulation. The activity of tissue-infiltrating immune cells constitutes a secondary ‘hit’ to the initial, chemical-induced burn. This culminates in prolonged skin inflammation leading to protracted healing and scarring. In a clinical trial, we acquired skin tissue from human subjects experimentally exposed to NM. From those studies we collected ‘omics’ data, which we have utilized to guide experimental models and translationally relevant milestones in the development of therapeutic strategies. In preliminary studies, we hypothesized that the skin’s response to NM would be blunted via the topical application of a persistent, stable radical scavenger in the form of a synthetic mimetic of human eumelanin. That is, by mimicking the natural function of melanin as a skin protective antioxidant, we would set the wound site on a path towards reparative healing. Indeed, in studies in mice and in human skin explants, we observed that topically applied synthetic melanin particles (SMPs) significantly increase the rate of repair. Therefore, in the UG3 phase, we will leverage our experience with mustard and our abundant access to fresh human skin tissue, to evaluate a select group of SMPs for their ability to mitigate the initial injury and to accelerate wound healing. The first selection criterion will be the ability of a given SMP to prevent sub-epidermal separation (blistering) in human skin explants. The second criterion is mitigation of keratinocyte necrosis. The third is a demonstration of inhibition of key biomarkers of epidermal damage. These criteria, tested in human skin explants, will lead to the advancement of six candidates for in vivo evaluation in our established NM mouse model. Here, the first criterion is a demonstration of reduced wound area size (early-stage) and rapid re-epithelialization (late-stage). Next, a successful SMP will lead to reduced scar formation and inhibition of hallmark skin damaging factors. This algorithm will lead to endpoint-driven prioritization and selection of two SMPs; a lead and back-up candidate to bring forward to the next phase. In the UH3 phase, we will evaluate the two SMPs in a NM porcine model. This model is adapted from FDA-enabling studies that includes a demonstration of in vivo 21-day wound reduction coupled with assessment using a 15-point histology grading instrument. Studies in this phase will involve optimization of the drug vehicle, dose and dosing schedule. This will position the lead candidate for advancement to the final evaluation of the same model with SM exposure in collaboration with a contract research organization (CRO). The result will be the identification of a lead Target Product Profile and a dataset from small and large animals that are aligned with our regulatory strategy for clinical translation of the SMP.