Yale University

NIH Research Projects · FY 2025 · 2022-09

We propose to evaluate the effects of pandemic eviction prevention policies on individual- and area-level death rates, leveraging variation in policies over time and between locations to gain new insight into the relationship between eviction and mortality. Stagnant wages and rising rents over the last several decades have resulted in unprecedented levels of housing cost burden in the U.S. and, in turn, forced millions of renters to face the threat of eviction each year. The pandemic worsened this crisis, resulting in catastrophic job and wage loss. To prevent a surge in evictions, federal, state, and local policymakers established a range of eviction prevention policies, most notably eviction moratoria and emergency rental assistance (ERA). These policies varied across the country in how and when they were implemented, creating large exogenous variation in eviction risk. We exploit that variation to evaluate the effects of eviction on mortality and inequalities in mortality. In so doing, we provide insight into the potential for eviction prevention policies to advance health. Our project has three aims: 1) We examine the effect of eviction filings on all-cause adult mortality overall and by age, race, and sex using a unique linkage of individual eviction filing records and Census Bureau microdata. We also estimate the number of lives saved by eviction prevention policies and project mortality gains had more protective measures been comprehensively implemented. 2) We analyze the effects of eviction prevention policies on county-level mortality using a difference-and-differences framework. We analyze how the staggered lifting of eviction moratoria (2020) and disbursal of ERA (2021-22) affected all-cause, cause- specific, and age-specific adult mortality 3) We explore the lived experience of eviction prevention policies using in-depth interview data collected with renters across two distinct policy landscapes. Our project combines the strengths of multiple approaches. Our unique linkage of individual-level eviction-filing and mortality records allows us to produce precise estimates of deaths averted due to eviction prevention policies. We also leverage natural experiments to examine the causal impact of these policies on county-level measures of mortality, capturing potential spillover effects. And our qualitative data help to clarify mechanisms underlying these impacts.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Adolescents with type 1 diabetes struggle more than any other age group to meet recommended glycemic targets. Continuous glucose monitoring devices, which enable real-time determination and communication of blood glucose levels, identification of trends, alerts, and alarms, and remote communication and monitoring, have improved metabolic and lifestyle outcomes and have markedly transformed the clinical management of diabetes in youth. However, these technological improvements have not benefited all populations similarly: lower rates of uptake and continued use of these devices in Black and Latinx young people have exacerbated pre- existing racial disparities in diabetes care and outcomes. Beyond socioeconomic issues such as access, developmentally and culturally specific factors relating to burden, diabetes distress, and self-efficacy are modifiable targets for specific and innovative interventions. Structured peer mentoring programs have been successfully employed in other chronic illness models to improve attitudes and behaviors around disease management. Using a stakeholder engagement approach, we will develop, optimize, and evaluate a peer- support intervention, Glu-COACH, to improve the uptake and maintenance of CGM use in Black and Latinx adolescents with type 1 diabetes. The input from a stakeholder panel representing patients, parents, and clinicians will inform each phase of the study. In Phase 1, we will collaboratively develop Glu-COACH and finalize study protocols with our stakeholders. In Phase 2, we will conduct a randomized controlled pilot study (n=50) comparing Glu-COACH to an enhanced standard care control group, convening quarterly research progress meetings with our stakeholders for ongoing collaboration. We hypothesize that the support, validation, identification, vicarious experience, and role modeling provided by trained peer mentors of the same cultural identity will significantly improve our primary aim of CGM use in Glu-COACH participants compared to the control group. We will also examine the effects of the Glu-COACH intervention on glycemic outcomes, such as time in range, mean glucose, and A1c, and behavioral outcomes, such as diabetes-related self-efficacy and distress, as well as benefits and burdens related to CGM use. Finally, we will critically examine the feasibility, acceptability, and implementation of the Glu-COACH intervention. This intervention will be upscaled and evaluated in a subsequent multi-site randomized controlled trial if proven effective.

NIH Research Projects · FY 2025 · 2022-09

Social determinants, including those related to food and housing, have a powerful impact on health, but there is uncertainty on how to best leverage policy to improve individuals’ social circumstances and related health outcomes. Presently, 11% of Americans have food insecurity, including 35% of families with incomes below the federal poverty line, and rates of food insecurity have increased dramatically in recent years. A growing literature finds associations between food security and mental health, positing a bi-directional relationship, wherein food insecurity increases stress and anxiety leading to poorer mental health, and serious mental illness impairs consistent access to food. Improving food security may be critical to improving mental health in low-income populations, but opportunities to examine the causal impact of such an intervention are rare. The Supplemental Nutrition Assistance Program (SNAP) has been shown to be effective in reducing, but not eliminating, food insecurity for low-income households. Many families on SNAP have residual food insecurity due to early exhaustion of SNAP benefits. Many policy experts believe that an increased investment in SNAP would both enhance the capacity of recipients to purchase healthy food and smooth the consumption of their benefits, potentially improving health outcomes. But this hypothesis has not been rigorously tested to date. During the pandemic, the federal government took unprecedented steps to increase SNAP benefits to low-income families. In January 2021, SNAP household budgets were increased by 15%, leading to an increase in federal SNAP benefits of about $40 per month on average. We leverage this natural experiment to examine whether increased food security from the SNAP changes reduced mental illness during the pandemic. We will thereby elucidate the causal relationship between food security and mental health, and their links to healthcare utilization. Finally, we will assess changes in the rate of benefit spending to inform the broader design of SNAP policy to optimize health outcomes. Our specific Aims are to a) assess whether increased SNAP budget allotments was associated with changes in indicators of mental health, b) examine the effect of an increase in SNAP budget allotment on the use of, and adherence to, recommended care services, and rates of adverse outcomes and health care costs among low-income individuals with previously diagnosed conditions, and c) estimate whether an increase in SNAP budget allotments impacted patterns of SNAP monthly benefit use. To answer these first-order policy questions, we use data from a novel, linked panel dataset that includes administrative Medicaid claims, administrative claims from the SNAP program, and data on the weekly use of SNAP benefits for beneficiaries. Our team is well-positioned to examine the understudied link between food insecurity and mental health and to assess the related public health, clinical, and policy implications.

NIH Research Projects · FY 2025 · 2022-09

Project Summary New solutions are desperately needed to reduce the annual global burden of dengue. As we are currently witnessing for SARS-CoV-2, virus genomics can be harnessed to directly inform public health control measures. For most other pathogens, including dengue virus, we do not have the depth of genomic information for these uses. For dengue virus, part of the issue is its complexity: it is comprised of four genetically distinct serotypes with many defined genotypes and even more undefined variants. Furthermore, our current surveillance and research programs are not set up to fully take advantage of virus genomics. In this proposal, I will address the major barriers for modernizing dengue virus genomic programs: (1) development of a universal-dengue virus whole genome sequencing approach utilizing the framework being established globally for SARS-CoV-2; (2) formation of a collaborative network in the underserved region of the Caribbean to uncover the circulating dengue virus diversity and provide local training and sequencing support; (3) utilization of travel surveillance to provide information from beyond the network to serve as a template for other genomic surveillance systems; (4) development of a new dengue virus genetic classification system to more easily identify variants of concern/interest and to better utilize the genomic data without needing complex phylogenetic analysis; (5) creation of a webtool to provide easy access to the variant classification system, detailed regional and county-level analysis, genomic data, and protocols; and (6) detailed phylogenetic analysis to uncover key epidemiological processes, like the outbreak emergence interval and patterns of spread, to aid in future disease forecasting. Completion of the goals outlined in this proposal would significantly advance our knowledge and assessment of dengue virus diversity, evolution, and epidemiology. Researchers will be able to capitalize on these data and tools to facilitate the innovation of genomics-informed control strategies.

NIH Research Projects · FY 2025 · 2022-09

Significance: Non-small cell lung cancer (NSCLC), the leading cause of death from cancer in the U.S., is plagued by racial disparities across the spectrum of care, particularly between Black and White patients. Despite the fact that these differences in the care of Black and White patients are long-standing and well-described, fundamental knowledge gaps remain. First, the degree to which differences in NSCLC care and outcomes vary across geographic regions has not been thoroughly explored. Second, little is known about how community factors contribute to disparities in care and outcomes at the regional level. Historically, health services research has focused on the healthcare system as the primary driver of disparities, with a limited exploration of community factors such as area-level socioeconomic status and access to health care. Further, it is unclear how key stakeholders within the healthcare and community settings address and overcome barriers to providing high-quality NSCLC care. Objective: We will address these critical knowledge gaps, with the overarching goal of mitigating disparities between Black and White patients with NSCLC in diagnosis, treatment, and outcomes, by identifying modifiable health system and intermediary factors and strategies that influence quality in lung cancer care. We intend to conduct an explanatory sequential mixed methods study, combining novel retrospective, population-based studies of disparities in lung cancer care with a positive deviance analysis. Through qualitative interviews with key stakeholders in regions with high versus low quality of lung cancer care, we will reveal strategies and processes that influence quality in lung cancer care and outcomes. Our hypothesis is that the magnitude of Black-White disparities in quality varies substantially across counties, and there are specific factors, such as health system factors, that exacerbate these differences. Specific Aims: We propose an explanatory sequential mixed methods study of Medicare beneficiaries diagnosed with NSCLC from 2013 through 2017: (1) To assess variation in racial differences in NSCLC care (non-Hispanic Black vs. non- Hispanic White) across counties within SEER regions. (2) To assess foundational factors associated with these differences in NSCLC care. (3) To assess health system/intermediary determinants (e.g. hospital characteristics and competition, patient access) of these differences in NSCLC care. (4) To identify stakeholder strategies associated with ensuring high quality NSCLC care, regardless of county socioeconomic characteristics. The proposed mixed methods study will have a high impact, using a combination of innovative methods to identify modifiable foundational, socioeconomic, and social factors that contribute to lung cancer differences. Furthermore, we will elucidate strategies that stakeholders have used to decrease care disparities, identifying approaches that can be adopted more broadly to improve quality of care for patients with NSCLC.

NIH Research Projects · FY 2025 · 2022-09

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive and lethal cancer that is resistant to currently available therapies. Notably, we discovered that PDACs are exquisitely susceptible to a range of therapies directed at RNA splicing. However, it is unknown how alterations in RNA splicing drive PDAC tumorigenesis or impact therapeutic responses. Thus, identification of the role of aberrant RNA splicing in PDAC tumorigenesis could reveal novel therapeutic targets for PDAC. Our long-term goal is to identify, design, and test novel mechanistic-based targeted therapies for highly aggressive tumors such as PDAC. The main objective of this proposal is to characterize the role of RNA splicing factor mutations in PDAC pathogenesis and treatment response. Recently, we identified that the most common gain-of-function driver mutations found in 50% of PDACs (mutant KRAS and p53) synergize and cooperate through altered RNA splicing. While >90% of PDAC cases harbor mutant KRAS, only 50% co-occur with mutant p53. We performed mutual exclusivity analysis among hundreds of annotated mutations in PDACs and identified two mutant splicing factors, SF3B1 and RBM10, that co-occur with mutant KRAS but do not co-occur with mutant p53 and are mutually exclusive between each other. Additionally, our proof-of-concept studies using newly generated genetically engineered mouse models, oligo-therapy and RNA splicing inhibitors demonstrated that RNA splicing is a therapeutic target. Our central hypothesis is that persistent RNA splicing defects, downstream of SF3B1 and RBM10 mutations, are a required adaptive mechanism for KRAS-mediated tumorigenicity and represent a therapeutic target for PDAC. We aim to 1) determine the role of aberrant RNA splicing in PDAC tumorigenesis, 2) identify the function and correct RNA splicing defects in pancreatic cancer, and 3) evaluate the impact of mutant RNA splicing factors on the therapeutic efficacy of spliceosome inhibitors and chemotherapeutic agents. Our expected outcomes include identification of 1) how aberrant RNA splicing underlies major cancer-driving events, 2) novel therapeutic targets for our newly generated oligo-therapy or small molecule inhibitors, and 3) a previously overlooked mechanism for how cancer can evolve through multiple mutations converging at a common mechanistic function. We will use innovative newly generated genetically engineered mouse models co-expressing KRAS and splicing-factor mutations in the pancreas leading to autochthonous PDAC resembling patient tumors, and we will employ novel computational and genetic biotechnologies to identify and correct RNA splicing defects. These results will uncover a fundamental, yet novel non-mutational mechanism required for PDAC pathogenesis: altered RNA splicing. This will provide a strong basis for future approaches to treat PDAC, which would significantly impact personalized therapies and patient outcomes. This research directly aligns with NCI’s mission to advance scientific knowledge of drivers and targets of cancer to improve patient’s lives.

NIH Research Projects · FY 2025 · 2022-09

Consumption or rejection of food are outcomes of a hierarchical multisensory decision- making process. Food-related decisions must frequently resolve conflicts, such as whether to consume a fruit that is visually appealing but smells rotten. Such decisions require neural substrates for evaluating characteristics of food—appearance, smell, taste, touch—and comparing/contrasting such characteristics to decide whether to approach, assess, and/or consume. We study food-related decision making in Drosophila melanogaster flies and Caenorhabditis elegans nematode worms, which have simpler behavioral repertoires, are highly experimentally tractable, and have proven utility at generating biological insights of relevance to mammalian model systems and human beings. The proposed Drosophila studies provide detailed mechanistic understanding of the circuit substrates, neuromodulatory pathways, and neural encoding of sweet/bittersweet food choice in Drosophila, and a foundation for research more broadly into decision making under conflicting information. The proposed C. elegans studies provide extensive evidence of worm color detection and the underlying cellular/molecular mechanisms, revealing pathways that could underlie an evolutionarily ancient opsin-independent color detection system present in other animals including mammals.

NIH Research Projects · FY 2025 · 2022-09

Project Summary/Abstract Alcohol use disorder (AUD) in mid-life is a significant, independent predictor of late-life dementia, particularly vascular cognitive impairment and dementia (VCID) that includes the common subtype cerebral amyloid angiopathy (CAA) characterized by deposits of amyloid-β (Aβ) in the cerebral vasculature. In only one decade, from 2005-2014, the rate of alcohol binging among the elderly rose 20% overall and by 50% in women only. The mechanisms underlying alcohol’s toxic effects on the central nervous system (CNS) remain incompletely understood, which limits the development of strategies targeted to reduce or prevent disease burden in heavy drinkers with and without CAA VCID. An underlying cause of alcohol’s damaging effects on the CNS is a poorly understood dysregulation of CNS fluid homeostasis. Preliminary evidence indicates that alcohol-associated abnormal fluid homeostasis manifests as enlarged cerebral ventricles and altered CSF flow dynamics. In humans, specifically, this manifests as lateral ventricular enlargement, and may reflect not only tissue loss but also itself be contributing to tissue damage and cognitive impairment. We propose that ethanol disrupts fundamentals of fluid homeostasis, by reducing CSF flow through the interconnected glymphatic and lymphatic systems, through biochemical and neuroimmune alterations thereby directly damaging the tissue. The functioning of glymphatic/lymphatic system supports fluid homeostasis and clears waste and toxins from the brain. Our proposed studies include a comprehensive series of experiments to quantitatively analyze alcohol’s effects on CSF secretion, glymphatic and lymphatic transport, and CSF/lymph ‘omics’ profiling in normal brain and with CAA VCID pathology. In Aim 1, we will determine how cEtoh interferes with CSF secretion in rats with/without VCID, using a novel MRI acquisition method to measure choroidal CSF secretion, as well as blood perfusion of the choroid plexus and cortex. We will also implement MRI sequences to measure brain morphometry in cEtoh-exposed rats with/without VCID and sample CSF, the meninges, and brain tissue to study pathology and biochemical lymphatic and inflammatory factors. In Aim 2, we will assess how cEtoh affects glymphatic-lymphatic transport, cerebral and meningeal lymphatic pathology, lymphatic waste output and cognition in rats with/without VCID. We will use dynamic contrast-enhanced MRI and computational fluid dynamics to measure CSF flow dynamics, glymphatic transport, and lymphatic waste drainage as affected by cEtoh and VCID, as well on CAA pathology and cognitive decline. In Aim 3 we will study how cEtoh compromises biochemical signaling and bio-physical changes in the CSF and lymph, including proteins that regulate fluid volume (renin-angiotensin) and proteins functioning in ion transport/signaling (Voltage dependent calcium signal, V-type proton ATPase and Carbonic Anhydrase). Molecular pathways that we identify can later be probed for therapeutic benefit.

NIH Research Projects · FY 2025 · 2022-09

Project Summary / Abstract: Cancer is a leading cause of death, but recent emergence of immunotherapies including the immune checkpoint inhibitors has offered promising new cancer treatment options. It is also evident that tumor immunity is highly complex and incompletely understood. The better understanding of tumor immune regulation may lead to identification of additional targets for cancer immunomodulatory therapy. In this study we will investigate the mechanism by which the cell surface receptor protein LRP5 regulates NK cell activities and tumor immunity. In our preliminary studies, we found that loss of LRP5 led to increased activation of NK cells independently of β-catenin stabilization despite LRP5 is involved in β-catenin regulation in other cells. Thus, we hypothesize that LRP5 regulates NK activation via previously uncharacterized mechanisms. Because genetic inactivation of LRP5 in NK cells suppresses tumor progression accompanied with enhanced NK activities in mice, this new mechanism is potentially of important clinical implications. Our preliminary results suggest that LRP5 may function as a fatty acid transporter leading to regulation of mTROC1 activity. In this application, we will carry out comprehensive in vitro and in vivo investigations to characterize this new mechanism for LRP5 and to determine the roles of this new mechanism in NK biology and tumor surveillance.

NIH Research Projects · FY 2025 · 2022-09

ABSTRACT Malaria caused by Plasmodium falciparum remains one of the leading causes of death globally of both children and pregnant women. The recent global stall in the reduction of malaria deaths has made the development of a highly effective vaccine essential. A major challenge to developing an efficacious vaccine is the extensive diversity of Plasmodium falciparum antigens. While genetic diversity plays a major role in immune evasion and is a barrier to the development of both natural and vaccine-induced protective immunity, it has been underprioritized in the evaluation of malaria vaccine candidates. This proposal will use genomic approaches to credential next generation malaria vaccine candidates. Reverse vaccinology is a method of identifying potential antigens for a vaccine that starts with the genomic sequence of an organism and uses that information to identify epitopes and antigens that might make suitable vaccine candidates. Since the genome sequence of Plasmodium falciparum was published, only four new potential candidate vaccines have entered clinical development, including PfRh5. The main objective of the proposed study is to use a reverse-vaccinology approach utilizing parasite genomic data directly from infected patients to identify and functionally interrogate the importance of diversity in these antigens. For these current and novel candidates, including PfRh5 and binding partners, we will test the role of genetic diversity on immune neutralization by creating transgenic parasites by using efficient CRISPR-Cas9 genome editing. These parasite lines will be used to assess the role of specific variants in immune evasion prior to Phase 2 clinical trials. We will use IgG from malaria-immune individuals, followed closely in long-term longitudinal cohorts, and IgG from subjects in vaccine trials to assess the degree of inhibition of replication of malaria parasites by growth inhibition assays, neutrophil respiratory burst, and opsonophagocytosis of merozoites. This approach requires the cohesion of genomic sequencing technologies to identify potential candidate antigens and naturally occurring diversity, well-characterized human longitudinal cohorts to follow evolution of infection and immunity, standardized assays to serve as in vitro correlates of immunity, structure-based approaches for vaccine design, and strong ties to both scientists and institutions in endemic countries. Our research team is uniquely positioned to combine these critical requirements to investigate the implications of parasite diversity on the development of protective immunity and vaccine efficacy, an essential factor to accelerate malaria vaccine discovery. This approach fills a critical need in the malaria vaccine development field in that it brings genetic diversity in candidate antigens to the forefront of vaccine candidate validation and credentialing. This study holds exceptional promise to discover new vaccine candidate combinations that will provide broadly neutralizing antibodies for inclusion in a globally effective vaccine, one that circumvents the parasite’s natural strategy to evade the immune system.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Alzheimer's disease (AD) disrupts brain organization, which is evident across spatial scales, from synapse loss to whole-brain connectivity, and manifests prior to or at the first sign of symptoms. Many neuroimaging modalities have contributed to our understanding of these changes, yet mechanistic insight into how dysfunction translates across scales, and species, is lacking. In part, these knowledge gaps exist because imaging modes specialize within a finite spatial milieu with access to a limited number of contrasts. To close these gaps, we propose a multimodal approach that leverages the strengths of complementary modes to deepen our understanding of changes in brain organization and inform the development of early-stage and preventative treatment strategies. Fully aligned with PAR-22-059, we propose a multimodal approach to interrogate AD-precipitated brain circuit disturbances and treatment-facilitated recovery. Our approach links excitatory and inhibitory synapse losses (positron emission tomography, PET), cell-type specific circuit-level dysfunction (wide-field calcium, WF-Ca2+, imaging) and brain-wide changes in the blood-oxygen-level-dependent (BOLD) functional magnetic resonance imaging (fMRI) signal in an established murine gene knock-in model of AD. Data will span relevant AD stages and be powered to address sex as a biological variable. Image collection will coincide with behavior testing. Our objective is to leverage the combined strength of complementary imaging modes to identify the synaptic changes that correspond to alterations in circuit and network-level dysfunction in the prodromal and early stages of AD. These data will provide mechanistic insights into the underlying causes of AD-related changes in brain functional organization that are evident in clinically accessible contrasts (PET and BOLD-fMRI). Aim 1. Map excitatory (E) and inhibitory (I) synapse losses. Using three PET tracers, we will map changes in the density of E, I and all synapses. These data will yield a better understanding of how E/I synapses are lost during AD pathogenesis and how these local changes in micro-circuits contribute to more global E/I imbalances. Aim 2. Discover the E/I circuit disruptions that underpin BOLD-fMRI network changes. WF-Ca2+ imaging affords cell-type specific measures of cortex-wide activity. By co-labeling inhibitory and excitatory neurons, we will measure E/I cortical circuit-level activity. Through simultaneous WF-Ca2+ and BOLD-fMRI, we will link cell-type specific E/I activity to brain-wide changes in BOLD-fMRI. Data will be collected from awake unanesthetized mice. Aim 3. Uncover the imaging correlates of treatment-facilitated recovery. Approaches from Aims 1 and 2 will be interleaved. Mice will be given one of two treatments to prevent synapse loss and cognitive decline. Synapse loss (PET), circuit and network function (WF-Ca2+/BOLD-fMRI) will be characterized longitudinally. The innovation of this work lies in the multimodal approach which bridges spatial scales from the synapse to the whole-brain. The significance of this proposal lies in deepening our understand how synaptic, circuit and network-level changes are interwoven and how together they shape AD pathogenesis and treatment response.

NIH Research Projects · FY 2025 · 2022-09

Project Summary The goal of the proposed project is to investigate functional and structural brain networks that predict disruptive behavior in children. Disruptive behavior disorders (DBD) affect over 113 million youths worldwide and are characterized by irritability/anger, aggression, noncompliance, and/or antisocial behavior. These disorders are of great interest because they are highly predictive of delinquency, criminality, and substance abuse in later adolescence and adulthood. DBD also co-occur in over 50% of children with autism spectrum disorder (ASD). A large body of evidence links DBD with perturbations in frontoparietal circuitry that support the cognitive control of emotion (i.e., emotion regulation), particularly connections between regions of the dorsal and ventral prefrontal cortex (d/vPFC) and amygdala. However, it is unclear if dysregulation in emotion regulation circuitry can contribute to a biomarker of DBD in children with and without ASD. With a focus on the amygdala-d/vPFC circuit, this study will be the first to examine disruptions in brain-wide connectivity and structure in emotion regulation networks as a transdiagnostic biomarker of DBD and disruptive behavior problems more broadly in children. First, we will develop and test a multimodal imaging biomarker of DBD in the Adolescent Brain Cognitive Development study dataset, which contains clinical and fMRI data for 11,878 9-12 year olds in the first and second releases. Next, we will test the hypothesized disruptions in emotion regulation circuitry using a fMRI task of cognitive reappraisal in a new, transdiagnostic sample of children with disruptive behavior with and without ASD. This study will leverage cutting-edge neuroimaging analytics that resonate with several NIMH research priorities: network neuroscience or connectomics, multimodal imaging, computational modeling (machine learning), big data analytics, and the RDoC domain of cognitive control. The proposed research will push forward the development of brain-based biomarkers of disruptive behavior that could guide development of targeted interventions, refinement of existing treatments, or identify children likely to respond to a particular treatment. The proposed project will prepare Dr. Karim Ibrahim to become an independent clinical researcher with a unique niche and expertise in transdiagnostic brain biomarkers of emotion regulation in childhood-onset psychiatric disorders using connectomics, multimodal imaging, and predictive modeling approaches. To accomplish this, the proposed training will provide Dr. Karim Ibrahim with multidisciplinary training in network neuroscience/connectomics, machine learning/predictive modeling, biostatistical approaches for the analysis of large imaging datasets, and emotion regulation circuitry. The training and research are enhanced by the intellectually rigorous environment at the Yale Child Study Center and Department of Psychology. The mentorship of a multidisciplinary team with complementary expertise in fMRI, connectomics, and child psychopathology ensures achievement of the research and training aims.

NIH Research Projects · FY 2026 · 2022-09

PROJECT SUMMARY: Plasticity occurs in the primary visual cortex (V1) in response to visual associative learning, where one stimulus is paired with reward (CS+) and another is not (CS-). However, learning is not a unitary process, and distinct stages of learning might drive contrasting plastic changes in V1. Indeed, previous work has found dynamic changes in V1 pyramidal neuron activity and inputs at different times throughout learning. Experiments separating distinct learning phases behaviorally are lacking, making it difficult to fully dissect V1 plasticity changes throughout learning. To address this, we have designed a 3-phase associative learning task that separates early, stimulus non-specific learning from late, stimulus-specific learning and reversal learning. We will also manipulate stimulus value in two ways: devaluation and spout removal. Using this behavioral paradigm along with 1- and 2-photon calcium imaging of the mouse dorsal cortex, we will test the following hypotheses: 1) that visual associative learning differentially recruits V1 activity during distinct learning stages, 2) activity in V1 is rapidly plastic in response to changes in stimulus value, 3) individual V1 neurons exhibit bidirectional plasticity across learning, 4) visual associative learning changes cortico-cortical functional connectivity, and 5) V1 activity is required to maintain these cortico-cortical functional connectivity changes. Our results will provide an unprecedented level of insight into V1 plasticity throughout learning and stimulus value manipulation. We will, for the first time, address how plasticity in V1 is mirrored across the dorsal cortex, or required for cortico-cortical plasticity.

NIH Research Projects · FY 2025 · 2022-09

Autism spectrum disorder (ASD) refers to a group of neurodevelopmental conditions characterized by impairments in social skills, verbal and nonverbal communication, and repetitive behaviors. ASD is highly prevalent, affecting up to 1 in 44 children in the United States and has been studied for decades, yet the underlying neural mechanisms that lead to the disorder are poorly understood. No biological tests to diagnose ASD earlier nor treatments targeting the core symptoms of ASD are available. Identification and functional analyses of high-confidence ASD risk genes are beginning to uncover convergent pathways by which genes with diverse functions lead to ASD. De novo mutations in Chromodomain Helicase DNA Binding Protein 8 (CHD8) are among the most strongly associated with ASD. CHD8 encodes a chromatin modifier that affects cell cycle progression through its role in gene expression regulation. Cell cycle control affects the timing of neural progenitor cell (NPC) proliferation and differentiation into neurons, which lays the foundation for proper neurodevelopment. Although numerous studies to date have improved our insight into CHD8 function, there are still critical gaps in our understanding of how CHD8 mutation alters cell cycle progression and neural proliferation, when these alterations take place, and the molecular mechanisms underlying these changes. To address these gaps, I will use a zebrafish line with a loss-of-function mutation in chd8. Zebrafish provide key advantages for studying early neurodevelopment processes in ways not feasible with mouse or in vitro models. Neural proliferation and migration take place from ~14 to 96 hours post-fertilization (hpf), providing easy access to embryonic timepoints that are difficult to study in mice. This vertebrate system will allow us to directly study fundamental neurodevelopmental processes at cellular, transcriptional, and circuit levels at critical time points, which is essential for understanding the function of CHD8 in shaping the developing brain. Based on my preliminary data, I hypothesize that CHD8 mutation causes decreased expression of proteins involved in translation initiation and cell cycle phase transitions, resulting in altered cell cycle timing and neural proliferation and differentiation, which may ultimately affect synaptogenesis and circuit formation. Aim 1 will investigate how and when mutations in CHD8 affect cell cycle timing and the balance of NPC proliferation and differentiation across early vertebrate neurodevelopment in vivo. Aim 2 will investigate the mechanisms underlying changes in cell cycle progression by investigating the impact of CHD8 mutations on protein synthesis. Together, my research will use rigorous multi-level approaches to elucidate the role of CHD8 in neurodevelopmental cell cycle regulation and protein synthesis, which will advance our understanding of ASD neurobiology. In addition to learning technical approaches in a powerful model system that will provide me with rigorous scientific training, my training plan affords me numerous growth opportunities with the support of superb mentors to pursue my long-term goal of becoming an independent physician-scientist.

NIH Research Projects · FY 2025 · 2022-09

Project Summary Investigating the neural bases of social behavior is critical for our understanding of complex, high-level cognition. Despite recent efforts to examine complex social cognition on a neural level, there are still many questions left unanswered. One prominent question is whether internal states typically associated with motivation and arousal can also be found in social domains. Internal states are indexed by continuous neural activity patterns that fluctuate over time and reliably predict specific behaviors. So far, internal states predictive of social behavior, known as social states, have only been shown in flies and mice. These results are foundational for the study of social states but cannot speak to complex social behaviors found in other species. Nonhuman primates exhibit some of the richest social behaviors that are likely guided by social states. Preliminary data in our lab shows that specific neural states predict social gaze during dyadic social interactions and expressing social preferences in macaques. This early evidence supports future endeavors in characterizing primate social states, including those described in this proposal. Internal states are often impacted by external environmental factors. For social states, this may include incoming social information from the environment such as visual or auditory communication from conspecifics. Such states may also be systematically affected by prosocial and antagonistic communication. However, whether social states are systematically influenced by social environmental information, and are therefore vulnerable to manipulation, is unknown. It is also unknown what regions of the brain may be implicated in social states. A prime candidate region for investigating this question is the basolateral amygdala (BLA), an area known for its broad involvement in social cognition and internal states. This proposal introduces the overarching hypothesis that social behaviors are driven by internal states and are vulnerable to changes by external social stimuli. In Aim 1 I will investigate whether visual and auditory stimuli carrying specific social information (‘social stimulus conditioning’) impacts specific behaviors involving social attention and perception. Potential impacts by social stimulus conditioning will be evaluated across two behavioral paradigms and provide knowledge on whether social states are generalized. In Aim 2, I will expand this research to include neural investigations of social states including possible involvement of the BLA. Neural states predicting specific social behaviors will be characterized through machine learning approaches. This will be the first study to compare social stimulus conditioning and social state induction in nonhuman primates. Overall, this proposal will address novel questions of primate social states and further our understanding of neural mechanisms underlying complex social behavior.

NIH Research Projects · FY 2025 · 2022-09

Problem: There is urgent need to reach women involved in criminal justice (WICJ) for lifesaving, evidence-based PrEP and medications for OUD (MOUD), using innovative healthcare delivery models that surmount existing social and structural barriers to engagement. Purpose: We use our newly validated decision aid and eHealth to remotely deliver integrated PrEP and MOUD to community-based WICJ with OUD in New Haven, Connecticut (CT) and Birmingham, Alabama (AL). Methods: We will enroll 250 PrEP-eligible WICJ with OUD to undergo randomization to: a) the “Athena” strategy, which includes the decision aid + eHealth for remote integrated PrEP/MOUD with a provider using outputs from the decision aid; or b) decision aid-only with referral to community-based PrEP/MOUD. Randomization will be stratified by site; past 6-month use of any stimulants; and baseline receipt of MOUD. Follow-up study assessments occur at months 1, 3, and 6. To understand implementation, we will conduct population modeling and engage with relevant stakeholders through focus groups using nominal group technique and in-depth individual interviews. Aims: The Aims of the project are: 1) To compare the Athena strategy to decision aid-only in terms of patient-level engagement in the PrEP and OUD care continua, considering key site differences; and 2) To assess scale-up potential of the Athena strategy in terms of modelled long-term outcomes and how stakeholders interact with eHealth for integrated PrEP/MOUD in WICJ in two diverse epidemiological and implementation contexts (CT and AL), using standardized definitions of implementation outcomes. Significance: The proposed research directly addresses the key objectives of RFA- DA-22-040 and priorities of the NIH Office of AIDS Research by testing innovative interventions to reduce stigma and improve the PrEP and OUD care continua in the key priority population of WICJ with OUD. Importance is high due to the focus on WICJ with OUD who have high HIV risk and in whom PrEP delivery strategies are limited. Innovation is high through use of a stigma- reducing decision aid and eHealth to overcome documented barriers to PrEP uptake, and a design that measures clinical alongside implementation outcomes in two diverse settings. Feasibility is high due to access to large numbers of WICJ with OUD in both contexts and a research team with considerable content expertise, collaborations, and supportive research infrastructure. Public health significance is high because delivering PrEP to key populations is central to EHE and assessing implementation factors to guide scale-up in two diverse contexts will help guide other settings where HIV prevention is needed.

NIH Research Projects · FY 2024 · 2022-09

Auditory verbal hallucinations (AVH) are among the most distressing symptoms in psychosis, and up to 30% of patients exhibit little to no response to current treatments. This is especially concerning given that the presence of hallucinations alone increases risk of suicide in patients with psychosis. One potential route toward development of new treatments for AVH is based on new evidence that many people with AVH never develop the need to seek treatment, despite the fact that AVH in treatment-seeking and non-treatment-seeking individuals tend to be similar in terms of low-level acoustic qualities such as loudness, duration, and location of voices. One particularly promising predictor comes from the fact that non-treatment-seeking voice-hearing populations consistently endorse a higher degree of control over their experiences than their treatmentseeking counterparts. Perhaps most strikingly, some individuals report an ability to fully control the onset and offset of their voices, which may make the experience of living with these voices significantly less disruptive and distressing. Moreover, new evidence indicates that voluntary inhibition of AVH may be developed in both treatment-seeking and non-treatment-seeking voice-hearers. Understanding the mechanisms specifically driving voluntary control over AVH could lead to new insights into potential treatment strategies to bolster these abilities. We and others have proposed hypotheses for how voluntary control of AVH might arise from cognitive inhibition, alterations in perceptual inference, or interactions between these processes. We propose to identify the mechanisms underlying inhibitory control of hallucinations. We will recruit 102 clinical and non-clinical voice-hearers with a range of control abilities for participation in a set of behavioral, imaging, and electrophysiological tasks designed to identify how voluntary control over voice-hearing occurs. Controlling for key confounds, we will relate our proposed measures to control abilities as measured on the newly-validated Yale Control Over Perceptual Experiences (COPE) Scale. We will also follow a subset of 60 of these participants over two years to identify predictors and correlates of changes in control abilities. To control for presence of hallucinations and psychosis, respectively, we will also recruit 51 matched healthy controls and 51 participants with psychosis but no voice-hearing to participate in the cross-sectional study. Principally, we hypothesize that exertion of control over AVH will result in: 1) activation of separable AVH- and control-related brain networks, the interaction between which will relate to abilities to exert control over AVH; 2) specific alterations in perception as demonstrated by the Conditioned Hallucinations task and EEG measures, corresponding to a decreased precision of perceptual priors, increased perceptual belief updating, or both; and 3) dual-task interference on the Think/No-Think task, an assay of cognitive inhibitory skills. Our goal is to translate the insights gained to new, mechanistically-informed treatments to enhance control over AVH.

NIH Research Projects · FY 2025 · 2022-09

Auditory verbal hallucinations (AVH) are among the most distressing symptoms in psychosis, and up to 30% of patients exhibit little to no response to current treatments. This is especially concerning given that the presence of hallucinations alone increases risk of suicide in patients with psychosis. One potential route toward development of new treatments for AVH is based on new evidence that many people with AVH never develop the need to seek treatment, despite the fact that AVH in treatment-seeking and non-treatment-seeking individuals tend to be similar in terms of low-level acoustic qualities such as loudness, duration, and location of voices. One particularly promising predictor comes from the fact that non-treatment-seeking voice-hearing populations consistently endorse a higher degree of control over their experiences than their treatmentseeking counterparts. Perhaps most strikingly, some individuals report an ability to fully control the onset and offset of their voices, which may make the experience of living with these voices significantly less disruptive and distressing. Moreover, new evidence indicates that voluntary inhibition of AVH may be developed in both treatment-seeking and non-treatment-seeking voice-hearers. Understanding the mechanisms specifically driving voluntary control over AVH could lead to new insights into potential treatment strategies to bolster these abilities. We and others have proposed hypotheses for how voluntary control of AVH might arise from cognitive inhibition, alterations in perceptual inference, or interactions between these processes. We propose to identify the mechanisms underlying inhibitory control of hallucinations. We will recruit 102 clinical and non-clinical voice-hearers with a range of control abilities for participation in a set of behavioral, imaging, and electrophysiological tasks designed to identify how voluntary control over voice-hearing occurs. Controlling for key confounds, we will relate our proposed measures to control abilities as measured on the newly-validated Yale Control Over Perceptual Experiences (COPE) Scale. We will also follow a subset of 60 of these participants over two years to identify predictors and correlates of changes in control abilities. To control for presence of hallucinations and psychosis, respectively, we will also recruit 51 matched healthy controls and 51 participants with psychosis but no voice-hearing to participate in the cross-sectional study. Principally, we hypothesize that exertion of control over AVH will result in: 1) activation of separable AVH- and control-related brain networks, the interaction between which will relate to abilities to exert control over AVH; 2) specific alterations in perception as demonstrated by the Conditioned Hallucinations task and EEG measures, corresponding to a decreased precision of perceptual priors, increased perceptual belief updating, or both; and 3) dual-task interference on the Think/No-Think task, an assay of cognitive inhibitory skills. Our goal is to translate the insights gained to new, mechanistically-informed treatments to enhance control over AVH.

NIH Research Projects · FY 2025 · 2022-09

Abstract Acute kidney injury (AKI) is a syndromic term encompassing a wide range of insults and pathogenic responses that lead to a rapid reduction in glomerular filtration. Although we have made significant progress in our understanding of kidney injury in animal models, far less attention has been focused on the pathogenesis and treatment of the diverse types of human AKI. The largest barrier in achieving this knowledge is the limited number of kidney biopsies performed for AKI and the small amount of tissue obtained from renal biopsy. The Kidney Precision Medicine Project is tackling this complex issue by recruiting altruistic patients with AKI who are willing to have kidney biopsies in order to advance our knowledge of human kidney disease. These biopsies are being interrogated using multiple complementary technologies. We propose to use Imaging Mass Cytometry to help provide a highly detailed, quantitative cellular map of nearly all cells present in sections from those kidney biopsies, including their differentiation state and activation of injury and repair pathways. Combined with the cell sequencing, metabolomic, and proteomic data generated under KPMP guidance, this will provide a substantial increase in our understanding of human AKI and CKD. Imaging mass cytometry (IMC) uses a high-resolution laser combined with a mass cytometer to detect the presence, location and amount of up to 42 different heavy metal conjugated antibodies hybridized to a tissue section. We have successfully used IMC with a panel of 22 heavy metal conjugated validated antibodies to identify resident kidney cell populations, infiltrating cell populations, and cell activation and injury states using archival FFPE human kidney tissue, and developed a machine learning technique termed Kidney MAPPS to rapidly and accurately identify, quantify and localize ~92% of all cells in those biopsies. We now propose to increase that validated antibody panel to >30 antibodies that will allow identification of >95% of cells and improve cell injury and activation state assessment, and to optimize the IMC and Kidney-MAPPS analysis pipeline to perform 2D and 3D quantitative assessment of cell location, cell-cell interactions and cellular responses in human AKI and CKD biopsy tissues (SA1). We will standardize a defined work-flow protocol coupled with rigorous quality control assessment steps at key points (SA2), and then apply this IMC work-flow to kidney samples provided by KPMP and integrate with the KPMP Central Hub and consortium members to develop accurate protocols for mapping the scRNAseq/snRNAseq data, proteomics data and metabolomics data onto the appropriate cells and locations using Kidney-MAPPS (SA3).

NIH Research Projects · FY 2025 · 2022-09

Project Summary/Abstract Fundamental questions in developmental biology revolve around understanding the sequence of molecular and cellular events that regulates lineage specification and differentiation. Nowhere is this more poorly understood than in the early human embryo, which remains experimentally intractable for technical and ethical reasons past the onset of gastrulation - the point when the three major germ layers of the human body are specified within the embryonic epiblast tissue. Myself and others have previously developed stem cell-based systems that recapitulate key aspects of 3-dimensional (3D) mammalian development in vitro, paving the way to a more complete understanding of the specific steps that govern embryonic development. Here, I propose an innovative new experimental approach, using human pluripotent stem cells (hPSCs), combined with cutting-edge bioengineering technologies for a controllable, efficient and scalable modeling of human epiblast development in vitro. This model will permit studying the human germ layer differentiation trajectory and temporal dynamics in the correct 3D-conformation. I will combine state-of-the-art tools in genetics, imaging, and single-cell multi- omics in tandem with high-throughput computational analyses to define the key molecular and cellular events that regulate developmental patterning under different conditions. I will further investigate how these events are controlled by cellular metabolism and nutrient availability, which has important implications for understanding the early embryonic origin of numerous human diseases. My proposal will open up a completely new and powerful experimental paradigm to dissect the fundamental, inter-connected, principles of human developmental genetics at early embryonic stages that are otherwise inaccessible. This new knowledge will also directly inform efforts to efficiently generate mature tissues and cell types from stem cells for disease modeling and cell therapies. Ultimately, these findings will be critical for possible prevention of adverse pregnancy outcomes, offering a unique opportunity to understand the cellular and molecular mechanisms behind developmental disorders and congenital pathologies.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Background. Lesbian and bisexual women (LBW) represent one of the highest-risk groups for hazardous drinking (HD) and comorbid mental health problems (e.g., depression, anxiety) because of their exposure to identity-related stressors (i.e., stigma-related burdens) and associated stress reactions, like drinking to cope. Research has identified cognitive (e.g., expectations of rejection), affective (e.g., shame), and behavioral (e.g., avoidant coping) pathways through which identity-related stress places LBW at disproportionate risk of HD and comorbid depression/anxiety. Yet no interventions address these pathways. In fact, no HD intervention has ever been tested for efficacy with LBW. Preliminary Studies. With deep stakeholder input and NIH (R01MH109413-02S1) and foundation (Lesbian Health Fund) support, we created affirmative cognitive-behavioral therapy (ACBT). This treatment is a 10-session CBT intervention focused on improving LBW’s stress reactions by building self-affirming cognitive styles and reducing avoidant coping. In a waitlist-controlled pilot trial (n=60), ACBT showed strong promise for reducing HD and depression/anxiety by building adaptive responses to identity-related stress, making it the first intervention with preliminary efficacy for improving this population’s co-occurring behavioral and mental health challenges. Methods. We now seek to test ACBT’s efficacy and identify facilitators of scale-up of this promising intervention. Aim 1: In a 2-arm randomized controlled trial (RCT) with LBW who experience HD and comorbid depression and/or anxiety, we will test the efficacy of ACBT (n=225) against treatment-as-usual (i.e., supportive counseling) (n=225). Our primary outcome is proportion of heavy drinking days (≥4 drinks) on 30-day timeline followback. Secondary outcomes include reduction in WHO alcohol risk level and depression and anxiety. Both ACBT and treatment-as-usual will be delivered via telehealth (Zoom), for which we have found strong feasibility and acceptability. Aim 2: Assessments at baseline, 4, 8, and 12 months will determine if reductions in ACBT’s intended psychosocial mechanisms (e.g., internalized stigma, rejection sensitivity, emotion dysregulation) mediate heavy drinking reductions. To advance personalized medicine, we will also examine whether ACBT is differentially efficacious across key demographic factors and identity-related moderators. Aim 3: To prepare for implementation in frontline settings, we will conduct semi-structured interviews with directors (n=20), providers (n=20), and service users (n=20) from a network of 250 LBW-serving community centers. Applying the i-PARIHS framework will help identify facilitators that can support ACBT implementation at these centers.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Despite the success of immunotherapies across several cancers, immunogenicity in solid tumors remains a major problem, pointing to the need to re-invent immunotherapeutic approaches and technologies. Checkpoint immunotherapies do not show the same response across patients with the same cancer, and neither mutational burden, DNA damage or expression of checkpoint proteins can accurately predict immunotherapy response. In addition, the progress on cancer vaccines has been limited by the lack of cancer specific and highly immunogenic antigens, antigen-delivery technology, and biomarkers to guide treatment. Based on these limitations in current cancer immunotherapies, we need to discover unconventional and unidentified mechanisms that regulate anti-tumor activity in aggressive cancers. The Programs presented seek to reveal how defective RNA splicing and anti- bacterial memory in T-cells determine the fate of anti-tumor immunity. By studying these mechanisms, we will have evidence for the development of novel and effective immunotherapies with penetrable delivery and tumor specificity. The long-term goal is to develop checkpoint immunotherapies and cancer vaccines that overcome immune ignorance, escape, and suppression in solid tumors. The project objective is to understand the intrinsic mechanisms of gene expression and antigen recognition that either inhibit or promote T-cell activity against tumor cells and develop checkpoint immunotherapies and cancer vaccines to unleash the full cytotoxic activity of T cells. In Program 1, we will dive deeper into the transcriptome of T-cells, where our preliminary data suggests that specific RNA splicing alterations in tumor infiltrating T-cells impact their maturation and function. Here we will evaluate the function of these splicing defects and test the therapeutic potential of correcting splicing alterations via our patented oligotherapy in pre-clinical models. In Program 2, we build on our discovery of an embryonic transcription program that acts as a bacterial virulence factor and is turned on in solid tumors associated with worst patient outcome. We will determine the role of acquired anti-bacterial immunity in cancer progression and test a novel cancer vaccine based on the expression of this biomarker as an approach to re-ignite anti-tumor activity. These programs are designed to test biological mechanisms that have not been appreciated in the context of immuno-oncology as therapeutic targets and could yield novel immunotherapies targeted at patient and tumor specific features.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT Atrial fibrillation is a major cause of ischemic stroke and stroke-associated mortality worldwide. Though risk of ischemic stroke in patients with atrial fibrillation is reduced with oral anticoagulation (OAC), many are not suitable candidates for long-term OAC due to high bleeding risk. To help treat such patients, transcatheter left atrial appendage occlusion (LAAO) with the Watchman device was developed and approved as an alternative to OAC. However, efficient methods for evaluating clinical outcomes associated the Watchman device and other novel cardiovascular technologies are needed for long-term surveillance after regulatory approval. In this mentored patient-oriented career development award, Dr. Kamil Faridi will use a national registry of LAAO procedures linked to Medicare claims data to evaluate how different data sources and innovative statistical approaches can be utilized for evaluating the long-term safety and effectiveness of the Watchman device in real-world practice. In Aim 1, he will use traditional and advanced statistical methods to compare ischemic stroke and bleeding events identified with administrative claims data to site-reported events in a national registry of patients who undergo transcatheter LAAO. In Aim 2, he will use linked pharmacy claims data to evaluate registry-reported OAC use and compare outcomes between LAAO patients and medically managed patients with atrial fibrillation in real-world practice. To supplement quantitative strategies for post-marketing surveillance, in Aim 3 he will use qualitative methods to explore patient-reported perspectives and quality of life among patients referred for transcatheter LAAO. Accomplishing these research aims will lay the groundwork for future studies examining real-world surveillance of other cardiovascular therapies and other clinically important outcomes. This project will be led by Dr. Kamil Faridi, an early career investigator, cardiologist, and faculty member at the Yale School of Medicine with a track record of success in the field of cardiovascular outcomes research. During this 5-year program, Dr. Faridi will pursue additional training in machine learning, pharmacoepidemiology, comparative effectiveness and qualitative methods applied to patient-oriented research. This will be accomplished with a comprehensive career development program designed to provide Dr. Faridi with the skills needed to become a successful clinical researcher in cardiovascular medicine. His long-term career goal is to become an independent investigator with specific expertise in the areas of post- marketing surveillance and real-world data utilization for novel cardiovascular therapies, with the ultimate aim of improving patient care. Over the course of the research and training program, Dr. Faridi will be guided by an exceptional mentorship team and advisory committee consisting of established clinical investigators with expertise in cardiovascular medicine, outcomes research, and advanced biostatistical and qualitative methods. With implementation of this research project and the resources provided for training and career development, Dr. Faridi will be well prepared to transition to research independence by the end of the award period.

NIH Research Projects · FY 2025 · 2022-09

OVERALL SUMMARY/ABSTRACT The Yale Superfund Research and Training Program (YSRTP) is driven by regulatory and community-based concerns about emerging contaminants that affect water resources and drinking water supplies at multiple sites in the US. The YSRTP has chosen to study 1,4-dioxane (1,4-DX) because of its common occurrence in Superfund sites and drinking water supplies, and its USEPA and IARC classification as a possible human carcinogen. The carcinogenic mechanism in the liver is unknown and its interaction with co-occurring and carcinogenic water contaminants (1,1,1-trichloroethane, 1,1,1-trichloroethylene, and 1,1-dichloroethane) has never been evaluated in either animals or humans. Given that liver cancer incidence rates have more than tripled since 1980, there is an urgent need to evaluate whether emerging water contaminants, such as 1,4-DX, may be contributing to this increase. Importantly, a lack of biomarkers of exposure to or the biological effects of 1,4 -DX have hampered epidemiologic studies, risk assessment and setting standards for the contaminant. In addition, the high polarity and low biodegradability of 1,4-DX restrict the ability to remove it from aquifer systems or drinking water. Indeed, available treatment technology is both expensive and not readily applied to water supplies. The YSRTP will foster problem-based, solution-oriented research related to 1,4-DX and its co-occurring contaminants through innovative approaches to evaluate environmental occurrence and human exposure, understand the underlying basis of adverse health effects, provide cost-effective remedial mitigation solutions and ultimately set the stage for improved regulation of this emerging contaminant. The YSRTP will carry out highly interactive projects to: (a) examine the health effects and biomarkers of exposure/effect to 1,4-DX (alone and as a mixture with its co-occurring contaminants) in animal models and humans, and (b) develop systems to monitor and mitigate human exposure to 1,4-DX in water. The biomedical science research projects will elucidate the mechanism(s) associated with 1,4-DX liver toxicity and carcinogenesis in mouse and zebrafish models (project 1), and exposure assessment and early biologic responses to 1,4-DX in human populations (project 2). The environmental science and engineering research projects will create highly-sensitive and selective electrochemical sensors for on-site, real-time detection of 1,4-DX (project 3) and develop innovative advanced oxidation processes for mitigation technology (project 4). Successful completion of these innovative studies will make a significant positive health impact by more clearly defining the health risks of 1,4-DX, elucidating biomarkers of exposure, and establishing effective new ways to monitor and mitigate this important emerging contaminant. Our systems approach integrates and links the research projects with the development of effective communication and education of stakeholders, and training of future scientists to ensure the program has a far- reaching impact on how emerging contaminants are addressed both in the US and globally.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Childhood maltreatment is associated with insecure attachment, emotional dysregulation, and abnormal threat detection. Here we propose that abnormal dysregulation of the brain beta-endorphin signaling pathway plays a central role linking childhood maltreatment with insecure attachment and with long-term behavioral abnormalities. This idea is supported by work showing that dysregulation of the main beta-endorphin receptor (mu-opioid receptor, or MOR) is associated with insecure attachment, emotional dysregulation, and abnormal threat detection. To test this premise, we developed a mouse model of complex trauma, abbreviated UPS. UPS recapitulates several key features of childhood maltreatment including the presence of multiple adversities, fragmented abusive maternal care, insecure attachment, impaired maternal buffering, increased threat detection, and abnormal social exploration. We also discovered that neurons in the hypothalamus expressing the agouti- related peptide (Agrp) are rapidly activated in response to unpredictable maternal separation in infant mice. Activation of Agrp neurons triggers the emission of ultrasonic vocalizations—the equivalent of the infant cry— and solicited dam’s attention and care. Thus, Agrp neurons function as an alarm system for the infant during distress. Intertwined with Agrp neurons are proopiomelanocortin (POMC) neurons, which are the main source of beta-endorphin in the brain. We found that POMC neurons of infant mice are rapidly activated by reunion with the dam. Activation of POMC neurons suppressed, while their ablation increased, the emission of ultrasonic vocalizations in infant mice. Thus, POMC neurons function as a buffering/safety system for the infant that is triggered during interactions with the mother. Based on these and other observations detailed in the proposal, we hypothesize that complex trauma in childhood—modeled by UPS in mice—impairs the ability of the mother to activate POMC neurons leading to reduced MOR signaling in Agrp neurons. This in turn causes prolonged activation of Agrp neurons and sustained distress that further erodes maternal buffering, secure attachment, and the ability of UPS mice to socialize and assess threat later in life. Work in Aim 1 will use fiber photometry in live moving pups and slice electrophysiology to characterize the effects of UPS on POMC and Agrp neuronal activation and its impact on MOR signaling in Agrp neurons. Work in Aim 2 will determine the contribution that POMC neurons and MOR signaling make to maternal affiliation/buffering and threat detection/social behavior in adolescence. Work in Aim 3 will test the extent to which sustained activation of Agrp neurons in infants is responsible for the behavioral abnormalities seen in mice exposed to UPS. Successful completion of this work will provide new insights into the mechanisms by which complex trauma in childhood programs abnormal attachment, enhances threat detection, and impairs social behavior.