Univ Of North Carolina Chapel Hill

NIH Research Projects · FY 2025 · 2021-04

PROJECT ABSTRACT We aim to define host factors that are associated with the more virulent human rhinoviruses (RV), RV-A and RV- C. Although RV are the most frequent cause of the common cold, they also provoke more severe manifestations, particularly in those with underlying lung disease. RV are the most frequent cause of asthma exacerbations, specifically induced by RV-A and RV-C, and only rarely RV-B. We have found that RV-A and RV-C require the host factor Stimulator of Interferon Genes (STING) for replication of their genome, while RV-B do not. Although STING is an immune adaptor critical for immune sensing in DNA viruses, its importance during RNA infections is becoming increasing recognized. In humans, STING1 exists as multiple variant alleles containing one or more single nucleotide polymorphisms. The most common STING variant is deficient in type I interferon (IFN-I) signaling, while other common variants have reduced IFN-I. As RV are restricted to only infect humans, we have found that for RV-A and RV-C, STING is a restricting host factor. While human STING robustly supports RV-A and RV-C replication, murine STING does not. The three Aims of the Project will explore the mechanism for STING host factor activity, determine how natural STING alleles influence RV-A and RV-C replication, and identify which nonstructural components of the virus interact with STING or the STING complex to promote infection. By studying the host factor role of STING during RV replication, the proposed research will define factors important for the increased virulence between RV species in this important human pathogen.

NIH Research Projects · FY 2025 · 2021-04

PROJECT SUMMARY/ABSTRACT Genome stability is determined by multiple DNA repair pathways, including both error-prone and error-free mechanisms. Mutations can be caused by inactivation of DNA repair pathways (e.g. BRCA1 defects) or by pathological activation of error-prone repair. The tumor suppressor p53 has pleiotropic effects on this balance. It physically interacts with base excision repair (BER), modulates nucleotide excision repair (NER), and regulates mismatch repair (MMR). Wild type p53 may inhibit error prone, but not error-free non-homologous end joining (NHEJ), and can modulate homologous recombination (HR). Recently, new roles for p53 have been identified, such as a role in APOBEC3B activation. Many mechanistic studies have studied the complex roles of p53 in DNA repair, but few large-scale studies of human tumors have investigated p53 and DNA repair pathway function in human tumors, and even fewer have evaluated these relationships by race. A more refined understanding of the relationships between p53 loss, DNA repair, and mutational signatures is now possible due to: (1) the advent of mutational signatures, which can provide DNA evidence of the functional effects and balance across multiple error prone and error free DNA repair pathways; and (2) recent improvements in expression profiling from formalin-fixed paraffin embedded (FFPE) samples. These advances are important for understanding breast cancer mortality disparities because they enable broad scale study in population-based resources. Our previous population-based studies have shown that p53 mutations are more common in African American breast cancer patients (60% p53 mutant vs. 35% among white breast cancer patients). Furthermore, DNA repair is critical for response to chemotherapy, both due to direct effects of DNA repair on chemotherapy resistance, and indirect effects on activation of immune responses. The current project will use an integrative approach to evaluate p53-related DNA repair pathway irregularities in human tumors, measuring both RNA expression and mutational signatures (Aim 1a). Then, p53 loss and DNA repair imbalance will be evaluated in relation to immune activation using both RNA and protein-based, spatial assays of immune markers (Aim 1b). These analyses will leverage, the Carolina Breast Cancer Study (CBCS), a study of 3000 women with breast cancer that oversampled black women (50% black women, n=1500). To better understand the germline determinants of black-white differences in DNA repair, existing genome wide SNP data will be used to identify genetic variants linked with DNA repair imbalance and immune response (Aim 2). This investigation will identify key DNA repair and immune pathways, in context of p53 mutation status and race, that interact to cause cancer progression and chemoresistance. The elucidation of these pathways is a key underlying step in identifying clinical and public health interventions to reduce mortality disparities. 1

NIH Research Projects · FY 2026 · 2021-04

Project Summary The expansion of effective interventions for HIV infection necessitates translation of these interventions to practice. Without careful scale-up, evidence-based interventions (EBIs) may not achieve their promise. Implementation science is the study of methods to promote the systematic uptake of research findings into routine practice. This D43 proposal is designed to expand HIV-related implementation science (IS) in Vietnam, a country that has been the focus of many NIH-funded intervention studies but has little IS capacity. The training program, referred to as Vietnam Implementation Science Advancement (VISA), uses a depth and diffusion model to achieve the following long-term goals: 1) to expand the capacity of Vietnamese researchers to conduct IS research and 2) to ensure that researchers, policymakers, and program leaders understand IS principles. These long-term goals will be achieved by addressing the following aims: 1) Initiate the Hanoi Medical University Implementation Science Program (HMU-ISP); 2) Develop IS knowledge and research capacity among four health science universities in Vietnam; 3) Develop IS capacity among Vietnamese policymakers and health program leaders to foster translation of HIV research into practice. We will build depth by establishing the HMU- ISP, which will serve as the home for in-person training in Aims 2 and 3. By completion of this D43, the HMU- ISP will have 9 faculty members, including the director (Dr. Giang, co-I), two VISA Senior Faculty who will receive 3-month immersion training in implementation science at UNC; two VISA PhD Fellows who will obtain their PhD in implementation science at UNC; and four VISA Faculty Trainees who will participate in our training program (Aim 2). To diffuse IS knowledge throughout Vietnam, we will train 12 VISA Faculty Trainees from four Vietnamese universities (including HMU) and 12 VISA Policy/Program Trainees, who will be selected from HIV- focused governmental and non-governmental agencies. The curriculum for the VISA Faculty and Policy/Program Trainees will focus on HIV-related IS knowledge and skills and research training. VISA Faculty Trainees will also develop an IS-focused course to expand implementation science training in their home institutions. VISA Faculty and Policy/Program Trainees will gain IS knowledge through in-person and virtual short courses and a series of webinars led by leaders in implementation science. Research training will include courses in grant writing and manuscript writing focusing on an HIV-related IS pilot project, which will be completed in the second year of training. The pilot projects will be led by an interdisciplinary team including one VISA Faculty and one VISA Policy/Program Trainee and will be supervised by a team of exceptional faculty, from both the United States and Vietnam. This innovative D43 brings together a strong team to expand HIV-related implementation science in Vietnam. By engaging both academia and public health policymakers in IS training, the potential for future translation of EBIs into practice will be maximized.

NIH Research Projects · FY 2025 · 2021-04

PROJECT SUMMARY / ABSTRACT: Nanoparticles (NPs) hold great promise for delivering more effective and safer cancer treatment than the small molecule drugs that are commonly used. This is based on studies reporting that these agents can potentially achieve greater exposure in solid tumors. However, these promises are largely hampered by a low and inefficient tumor uptake in which only 5-10% of NPs in the plasma are actually distributed from plasma to solid tumors. Certain tumors, such as pancreatic cancer (PaCa), have even greater inherent barriers to the tumor delivery of NPs. Thus, there is a strong need to discover methods that can significantly and safely enhance the overall delivery of NPs to tumors. Our overall hypothesis is that induction minibeam radiation therapy (MRT), a novel radiation treatment, is such a method. Whereas, conventional broad beam radiation (BRT) only moderately enhances drug delivery to tumors (0.2- to 2-fold). MRT is an experimental radiation therapy with unique spatial and dosimetric characteristics that are drastically different from conventional BRT. Solid preclinical studies have demonstrated that MRT is capable of an ultra-high therapeutic ratio. We recently discovered that MRT, in contrast to BRT, modifies tumor vasculature and increases tumor perfusion. We hypothesize that we can take advantage of the changes in tumor perfusion induced by MRT to significantly and safely enhance NP delivery to tumors compared to NPs alone or after BRT. This hypothesis is supported by our extensive results in genetically engineered mouse models (GEMMs) of breast cancer where induction MRT prior to administration of PEGylated liposomal doxorubicin (Doxil®; PLD) enhanced the delivery of PLD to tumors by an unprecedented magnitude of 6- to 10-fold and the enhancement was sustained safely with weekly treatments. In addition, MRT produced a 4-fold greater increase in the tumor delivery of PLD to GEMMs of breast cancer, which was associated with higher levels of overall and PD-L1 expressing macrophages compared to BRT. Our 2nd pilot study in PaCa GEMMs showed that MRT was able to increase the tumor exposure of PEG-liposomal irinotecan (Onivyde®, FDA approved for PaCa treatment) and its active metabolite SN38 by >4-fold compared to Onivyde alone. This grant will allow us to translate our ground breaking MRT results to PaCa where the barriers to NP delivery are extensive, surgical resection is the only curative option but only 15% of patients have resectable disease and the MRT + NP regimen would be ideal for pre-surgical neoadjuvant treatment of PaCa. This work will be performed by a multidisciplinary research team using novel models, technologies and FDA approved drugs that can be readily translated to clinical trials in 3 aims over 5 yrs: AIM 1. Evaluate induction MRT-enhanced delivery of NP anticancer drugs in GEMMs of PaCa; AIM 2. Investigate mechanistic effects of induction MRT-enhanced tumor delivery of NPs in GEMMs of PaCa; AIM 3. Evaluate induction MRT-enhanced efficacy of NP anticancer drugs in GEMMs of PaCa. This proposal aims to overcome the inherent major barriers in NP delivery to tumors, especially in PaCa, which has significant barriers to drug tumor delivery.

NIH Research Projects · FY 2025 · 2021-04

TYRO3 is a member of the TAM (TYRO3, AXL, MERTK) family of receptor tyrosine kinases. All three family members are aberrantly expressed in cancer cells, where they function to promote cell survival, mediate resistance to a variety of cytotoxic chemotherapies and molecularly-targeted agents and have additional roles in macrophages and other innate immune cells where they function to suppress anti-tumor immunity, leading to enhanced tumor growth and metastasis. These and other data implicate the TAM kinases as potential therapeutic targets in a wide variety of human tumors. Moreover, because of the oncogenic roles for TAM kinases in both tumor and immune cells, inhibitors are expected to provide anti-tumor action mediated by both direct tumor cell killing and modulation of the innate immune response. While the TAM kinases have overlapping functions, they also play unique roles in some contexts. Specifically, our preliminary data suggest that suppression of anti-tumor immunity is particularly dependent on TYRO3. Here, we propose to utilize a well-established and productive team of researchers along with computational-aided drug design and enzymatic, cell-based and pharmacodynamic assays to develop novel, potent, and selective TYRO3 inhibitors and validate their biochemical and functional activities in TYRO3- dependent tumor xenograft models and immune-competent syngeneic cancer models. TYRO3 can localize to the nucleus and inhibition of nuclear localization induced apoptosis in colon cancer cells, suggesting non- canonical oncogenic functions for TYRO3 which might not be effectively targeted by kinase inhibition alone. Thus, both traditional small molecule kinase inhibitors and proteolysis-targeting chimeric (PROTAC) degraders that selectively target TYRO3 for ubiquitination and degradation will be developed and compared. At the completion of this work, we expect to deliver a TYRO3-selective inhibitor suitable for advancement to GLP toxicity studies in multiple species, sufficient preclinical validation studies to support an IND application describing this compound, and a viable method for large-scale synthesis of the compound.

NIH Research Projects · FY 2025 · 2021-04

PROJECT SUMMARY Non-invasive functional magnetic resonance imaging (fMRI) has revolutionized our understanding of macroscopic functional brain networks. However, inherent constraints of current fMRI methodologies in humans limit our ability to probe the mechanisms underlying these networks. The overarching goal of this project is to shed light on cellular and circuit mechanisms underlying the functional organization of the default-mode network (DMN) – a large-scale brain network that is crucial for a wide range of behaviors. While the new technologies in rodents allows us to experimentally reveal causal control of DMN, rodent DMN topology has only been defined using resting-state fMRI, but not functionally in terms of activation or suppression of brain activity in response to behaviorally relevant salient stimuli. This represents a critical barrier preventing any straightforward translation between rodent and human DMN research findings. To address this, we developed a novel silent zero-echo- time (ZTE) fMRI technique, enabling awake rodent imaging and the use of an auditory oddball paradigm, wherein deviant oddball stimuli presented amongst a sequence of repetitive control stimuli can drive attention and suppress DMN. We also developed an MR-compatible, four-channel, spectrally-resolved fiber-photometry system, allowing concurrent recording of ground-truth neuronal activities during fMRI. To shed light on the circuit mechanisms governing the DMN, we proposed two complementary research Aims building on our rigorous prior research. In Aim 1, we will determine how attention to salient stimuli alters DMN activity and connectivity using the novel ZTE-photometry platform. In Aim 2, we will introduce time-locked optogenetics on defined cell types to causally manipulate the activity of anterior insula – the brain region assumed to be responsible for DMN dynamic switching in numerous fMRI causal modeling studies. Functionally dissecting the rodent DMN architecture is critical to the understanding of DMN transition mechanisms, which will enable us to causally model, and make predictions about brain states, bringing insight into the network basis of human behavior and neuropsychiatric/neurological disorders. 1

NIH Research Projects · FY 2025 · 2021-04

Delirium is a serious cognitive disorder associated with Alzheimer’s disease and related dementias (ADRD) that affects ~2.6 million older adults yearly. It is a frequent complication of acute illness, surgery and, now, of COVID-19 infection in older adults. Recognizing the relative dearth of delirium research, the National Institute for Aging (NIA) supported the establishment of the Network for Investigation of Delirium: Unifying Scientists (NIDUS), a collaborative interdisciplinary group of 28 investigators, from 27 institutions, to advance delirium research and develop network infrastructure. This included the creation of an annual “NIDUS bootcamp” conference, to bring together the growing national- and international delirium research community for networking and education. The bootcamp aims are to advance the science of the field and to provide junior investigators with intensive mentorship, through mock NIH application reviews, clinical and research lectures, breakout sessions, and post-bootcamp networking. Bootcamp alumni are provided guidance on: 1) using the NIDUS Delirium Research Hub, Measurement resources and Bibliography, 2) submitting proposals to the NIDUS Pilot Program (13 one-year $50,000 grants awarded), NIA GEMSSTAR/CLINSTAR, the Alzheimer’s Association, and other foundations, 3) attending Mentoring webinars, 4) participating in Junior Faculty Working Groups, and 5) submitting research abstracts to the American Delirium Society (ADS) Annual Meeting. As PIs, 94 alumni have received 46 grants, of which 18 (40%) were NIH-funded, and published 265 original peer- reviewed articles. NIDUS has jumpstarted the careers of many young investigators, particularly bootcamp alumni, enabling them to launch independent programs in delirium research. The goal of this application is to support continuation of a yearly, themed Delirium Bootcamp Conference (DBC), to ensure that the progress of this active research community is sustained. The first-year theme will be the inter-relationship between delirium and ADRD. The Specific Aims are to: (1) Engage and support junior investigators in delirium research through mentorship and access to the NIDUS resources/network (2) Boost the researchers’ funding success (3) Facilitate publication of delirium research and provide ongoing mentorship, and (4) Facilitate networking among junior, mid-career, and senior researchers during and after DBC. As the pool of delirium investigators expands, there is a critical need for a conference focused on addressing cutting-edge research methods in all areas of delirium research, including the relationship with ADRD, “-Omics” research, machine learning and big data, innovations in randomized trials, animal models and mechanistic research, and clinical practice improvement. The DBC will provide an unparalleled opportunity to advance cutting-edge delirium research through interactive didactic sessions and in-depth guidance on complex and nuanced research methods essential for the highest caliber and most impactful delirium research.

NIH Research Projects · FY 2025 · 2021-04

PROJECT SUMMARY/ABSTRACT. Polypharmacy is a highly prevalent problem in older adults, particularly in those with Alzheimer’s Disease (AD) and AD-Related Dementias (AD/ADRD) and those residing in the nursing home (NH). Many older adults continue to receive medications originally prescribed for disease prevention until the end of life, despite a lack of sufficient evidence to justify their continued use in advanced age. The use of bisphosphonates for fracture prevention is one example. Although bisphosphonates are effective in reducing fractures in healthier, community-dwelling individuals, there is insufficient evidence of continued benefits that is generalizable for NH residents with AD/ADRD. There is also substantial clinical heterogeneity in this population with regards to fracture risk, mobility, and life expectancy, creating further uncertainty as to whether all NH residents with AD/ADRD benefit from bisphosphonate use. Deprescribing is a patient-centered approach to reduce or stop medications that are no longer appropriate considering goals of care, time until benefit, and life expectancy. Considering the lack of strong generalizable evidence and potential for side effects, bisphosphonates may be targeted for deprescribing in NH residents with AD/ADRD. Deprescribing may also be justifiable considering the extended period of benefit of bisphosphonates, which may last for up to 2 years after discontinuation exceeding the life expectancy of many residents with AD/ADRD. However, no studies to date have evaluated the appropriateness of deprescribing bisphosphonates as a means to reduce the burden of polypharmacy and adverse effects in this population. Large observational studies of secondary data are uniquely positioned to evaluate the benefits and harms of medication use and deprescribing in older NH residents with AD/ADRD, given the barriers to conducting randomized studies in this population. This study will evaluate determinants, clinical outcomes, and cost-effectiveness of deprescribing bisphosphonates in NH residents with AD/ADRD. In Aim 1, we will conduct a qualitative study using semi-structured interviews to identify determinants of deprescribing bisphosphonates from the perspectives of family/informal caregivers and prescribers of NH residents with AD/ADRD. In Aim 2, we will conduct an observational study of Medicare administrative data to evaluate clinical outcomes (fractures and adverse effects) associated with deprescribing bisphosphonates in a sample of older NH residents with AD/ADRD. In Aim 3, we will evaluate the cost-effectiveness of deprescribing bisphosphonates in NH residents with AD/ADRD, considering medication costs and utilization for fractures and adverse effects. This study will address a critical gap in knowledge and inform future recommendations for optimizing bisphosphonate use to prevent fractures in this vulnerable and medically complex population. This award will also provide the principal investigator with protected time to develop skills in qualitative research methods, advanced techniques to reduce confounding in observational studies, and methods for cost-effectiveness analyses.

NIH Research Projects · FY 2026 · 2021-04

Project Summary/Abstract Dopaminergic neurons (DANs) are a molecularly, anatomically and functionally heterogeneous neuron group that is essential for learning across animal phyla. In the midbrain, distinct populations of DANs are responsible for memories with different valence or stability. Thus, the dopamine system comprises parallel subsystems, each of which operates as a qualitatively distinct learning system. This raises two important questions: 1. How does the heterogeneity of DANs impact synaptic plasticity to form distinct types of memories in each subsystem? 2. How are the signals from parallel subsystems integrated to ultimately trigger a unified behavior? Answers to these questions are required to understand the logic that governs the parallel memory systems. The mushroom body (MB), the major associative learning center in the Drosophila brain, is an excellent model to tackle these questions because it comprises dopamine subsystems, each of which is clearly defined by a unique set of DANs and MB output neurons (MBONs). These individual MB compartments support distinct types of memories that vary in valence and stability, properties shared with mammalian dopamine subsystems. However, in both invertebrate and vertebrate brains, it remains an open question whether the diversity of memory properties is derived from intrinsic characteristics of DANs or from an extrinsic circuit architecture. Aim 1 will test the hypothesis that combinations of DAN cotransmitters define compartment-specific rules of synaptic plasticity and thereby determine the memory properties. By identifying novel DAN cotransmitters and their physiological and behavioral roles, the causal relationship between plasticity rules and memory properties will be tested. In Aim 2, integration mechanisms of different types of memories will be determined by identifying neurons that pool input from multiple MBONs. Synaptic integration, behavioral roles and activity changes after learning will be determined in these integrator neurons. In this project, cell-type-specific transcriptome and the comprehensive connectome data available in the field will guide our molecular and circuit interrogation by in vivo electrophysiology, calcium imaging and behavioral assays. Collectively, this project will address fundamental questions regarding the heterogeneous organization of the dopamine systems and pioneer the circuit motif that is currently inaccessible in vertebrates.

NIH Research Projects · FY 2025 · 2021-04

PROBING ALLOSTERY IN METHYL-LYSINE READER DOMAINS Abstract: Chromatin is the complex of histone proteins, RNA, and DNA that dynamically packages the genome within each eukaryotic cell. While cell lineage specific transcription factors clearly play a dominant role in the control of gene expression, the regulation of chromatin accessibility via post-translational modifications (PTM) of histones is of great current interest as the opportunities for pharmacological intervention in the action of the associated proteins are significantly better than in the direct perturbation of transcription factors by small molecules. The molecular details of chromatin regulation are just beginning to be understood and chemical biology is poised to play a central role in advancing scientific knowledge and assessing therapeutic opportunities in this field. Specifically, cell penetrant, high-quality chemical probes that modulate the regulation of chromatin state are of great significance. The advantages of a small molecule driven approach to exploring chromatin biology are numerous: temporal resolution; mechanistic flexibility; ease of delivery in cells and potentially, in vivo; and significantly, a chemical probe may provide an immediate transition to a drug discovery effort, possibly cutting years off the time between target selection and therapeutic intervention. This impact of a chemical probe results from simultaneously addressing target ‘validation risk’ (the likelihood that pharmacologic modulation of the target will have a favorable outcome in a disease) and ‘technical risk’ (the likelihood that a tolerable molecule that modulates the target can be discovered). While probes often lack some features required in drugs, their discovery diminishes many target validation and technical risks and creates a cascade of assays, structural and mechanistic information that is enabling to subsequent efforts focused on drugs. While high-quality probes are challenging to develop, they are achievable within the resources available to academic programs, and their creation is a fantastic training experience. To maximize the impact of our probes, we intend to continue our approach of sharing them without creation of intellectual property. We have pioneered a target-class probe discovery strategy within the large family of methyl-lysine (Kme) readers. We have been productive in this area and built momentum for future studies focused on the allosteric interactions between Kme reader domains, nucleotide binding domains, and the catalytic domains that regulate chromatin function. During this effort, we have established a network of talented collaborators that complement our strengths in chemical biology, medicinal chemistry, in vitro assay development, and biophysics; with strengths in molecular, structural and chromatin biology. Allosteric interactions in chromatin regulatory complexes are critically important phenomena that create unique opportunities for pharmacologic intervention. We will focus our future endeavors on this exciting frontier in the Kme reader family.

NIH Research Projects · FY 2024 · 2021-04

Program Director/Principal Investigator (Last, First, Middle) Pawlinski, Rafal and Key, Nigel Abstract Venous thromboembolism (VTE) is the third most common form of cardiovascular disease after myocardial infarction and ischemic stroke, with approximately 900,000 cases annually in the United States. VTE may present clinically as deep vein thrombosis (DVT) and/or pulmonary embolism (PE). However, it is unknown why some patients present with symptomatic DVT, while others present with PE. In recent years, it has been established that individuals with sickle cell disease (SCD), as well as carriers for SCD -- who are said to have sickle cell trait -- are at increased risk of VTE. Interestingly and unusually, in both sickle cell disease and trait, a skewed distribution in the proportions of patients with DVT and PE (in favor of PE) is observed. We have termed this phenomenon “the sickle cell paradox”. SCD is due to an inherited mutation in hemoglobin that is expressed exclusively in red blood cells (RBCs). Therefore, in this proposal, we postulate that a greater understanding of the mechanism of VTE in SCD, as well as an explanation for the sickle cell paradox, will be explained by a detailed study of the role of sickle RBCs in VTE. To address this question, we will utilize animal models of SCD with experimentally induced venous thrombi, while concurrently studying the qualitative aspects of blood clots formed from the blood of patients with SCD ex vivo. Towards this goal, we will address the following specific aims: in Aim 1, we will determine the effect of RBCs on venous thromboembolism in a mouse model of SCD. We expect that partial RBC exchange will result in smaller, more stable DVTs and a reduced incidence of PE. In Aim 2, we will investigate how sickle RBCs enhance thrombin generation in SCD. In the third Aim, we will determine the cellular and molecular mechanisms that attenuate sensitivity of sickle clots to fibrinolysis. We anticipate this effect is mediated both by the inherited defect in SCD RBCs, but also the fact that platelets in patients with SCD are over-activated. Finally, in Aim 4, we will determine whether reduction of platelet numbers reduces VTE and restores normal susceptibility to fibrinolysis in sickle mice and SCD patients undergoing chronic RBC exchange. With an annual worldwide SCD birth rate of more than 300,000 (and an estimated 250 million individuals with sickle trait), a greater mechanistic understanding of VTE in sickle cell disorders is a high priority. OMB No. 0925-0001/0002 (Rev. 03/16 Approved Through 10/31/2018) Page Continuation Format Page

NIH Research Projects · FY 2025 · 2021-04

Epidemiologic identification and mechanistic investigation of early life environmental risk factors for eosinophilic esophagitis ABSTRACT Eosinophilic esophagitis (EoE) is a recently recognized immune-mediated disease defined by abnormal infiltration of eosinophils into the esophageal mucosa, leading to failure to thrive, abdominal pain, vomiting, and heartburn in children, and progressing to esophageal stenosis and food impaction in adults. Though initially thought to be rare, the incidence and prevalence are rising dramatically, and over the past decade EoE has rapidly become a major cause of upper gastrointestinal morbidity. Despite increases in the understanding of the condition, it is currently not possible to determine why individual patients develop EoE. This is frustrating for patients and practitioners alike. EoE is considered to be an immune/allergen-mediated disease, and epidemiologic studies support a primarily environmental etiology. However, environmental risk factors have not been extensively studied in EoE, and prior studies, including by our own group, are limited by a crude assessment of exposures, recall bias, inability to assess fetal biomarkers, and lack of mechanistic understanding. Our goal is to address this knowledge gap by using an innovative method to precisely measure early life exposures in deciduous (primary, or “baby”) teeth that may be implicated in EoE development. Of particular interest are early life antibiotic exposure and duration and intensity of breastfeeding (which can be derived from barium levels in teeth). Increased antibiotic exposure and decreased breastfeeding have been linked to risk of atopic diseases. Measuring selected environmental exposures in teeth has never been applied to EoE, but we have documented the feasibility of this approach. This assessment, together with the use of novel cellular and molecular techniques for elucidating the mechanisms underlying the effects of these early life exposures, has the potential to greatly enhance our understanding of the pathogenesis of EoE. Our hypothesis is that the risk of EoE related to early life exposures is primarily due to an impaired esophageal epithelial barrier, and that genetic susceptibility will interact with the exposures to modify risk. The specific aims are to 1) determine the association between early life antibiotic exposure and EoE; 2) determine whether breastfeeding is associated with EoE, and evaluate whether the susceptibility genotype for CAPN14 modifies the association between breastfeeding and EoE; and 3) determine the functional significance and mechanisms of early life exposures on esophageal epithelial architecture and barrier function. To achieve these aims, we will conduct a case-control study to characterize temporal exposures, and in parallel will perform mechanistic analyses. This innovative, hypothesis-driven, and rigorously designed study will lead to robust and unbiased results. It will be conducted by a multidisciplinary team with recognized expertise in EoE, epidemiology, clinical/translational/lab research, and exposure science. The results will have a major impact on the understanding of EoE etiology and by potentially identifying opportunities for disease prevention which could lead to the development of new treatment options.

NIH Research Projects · FY 2025 · 2021-04

Abstract Individual differences in brain structure, including cortical morphology and the white matter connectome are associated with risk for psychiatric disorders. The first year of life is a period of rapid and dynamic structural and functional brain development and new data from our cohort suggests that a large portion of individual differences in brain structure in 10 and 12 year olds is already present in the first year or two of life. Early adolescence and puberty is the second major period of postnatal brain development, characterized by dynamic structural and functional brain maturation and reorganization, and emerging risk for psychiatric disorders, though it is not known how this period of development contributes to individual differences in brain structure and risk. The UNC Early Brain Development Study is a unique and innovative longitudinal study that has followed children, enrolled prenatally, with imaging and cognitive/behavioral assessments at birth, 1, 2, 4, 6, 8 and 10 years. 482 children from this cohort are now reaching adolescence, and we propose to follow these children at 12, 14 and 16 years of age. MRIs, including structural, diffusion tensor, and resting state functional imaging, will be performed. Cognitive and behavioral development will be assessed, with a focus on the phenotypes of executive function, attention, and anxiety, consistent with RDoC constructs important for psychiatric disorder risk. We will determine how adolescent brain development contributes to individual differences in relation to early childhood development and whether the white matter connectome is a useful early imaging biomarker. Knowledge gained in this study will improve our basic understanding of human brain development, and ultimately inform early intervention strategies that prevent or mitigate risk and illness severity. Relevance New knowledge gained in this study will provide a dramatically improved framework for understanding childhood brain development and its relationship to cognitive and behavioral outcomes in adolescence, and to risk for subsequent psychiatric disorders.

NIH Research Projects · FY 2024 · 2021-04

PROJECT ABSTRACT Adolescent Wellness Visits in Tanzania Getting adolescents in the door of a health facility is an entrenched health system problem, particularly for HIV and sexual and reproductive health (SRH) services. Adolescents in low-resource settings need a preventative health service platform applicable for all young people that promotes a culture of health-seeking behavior. In response to PAR- impact of Adolescent Wellness Visits (AWVs), a 19-274 [Dissemination & Implementation Research in Health], this R01 will evaluate the new health service platform, for reaching young adolescents with HTC and other evidence-based prevention services which are clinic-based and school-facilitated. We posit that by coupling sexual and reproductive health (SRH) and non-SRH information and services, issues of self- risk assessment, and access to services may be circumvented. AWVs could meet the SRH needs of at-risk adolescents, and have a larger public health impact for all adolescents on access to traditionally neglected and untreated non-SRH issues such as poor nutrition, vision, dental, and mental health problems at the time of delivery as well as in the future as adolescents continue with more timely service utilization. The AWV is designed to be delivered during the last year of primary school when school attendance is high and adolescents are on the cusp of puberty (mean age 13). This project is a collaboration between Duke University and Muhimbili University of Health and Allied Sciences in Dar es Salaam, Tanzania. Specific Aims are: 1) To assess the impact of the Adolescent Wellness Visit model on HTC (primary outcome for all adolescents) and contraceptive uptake (secondary outcome for sexually active adolescents) up to two years post-primary school via a cluster randomized controlled trial (24 school-clinic pairs: 12 intervention + 12 control; n=552 adolescents); 2) To evaluate factors that support or limit implementation of the AWV model and fidelity/adherence to implementation of the proposed package of evidence-based practices included in the AWV; and 3) To determine the cost-effectiveness of the AWV model for increased HIV testing, reaching PEPFAR diagnostic targets, and reductions in unmet need for contraception. This D&I proposal creates and leverages a new population level health service for adolescents that aligns with NICHD’s Research Theme #5: Improving Health during the Transition from Adolescence to Adulthood (NOT-HD-18-031), and as a nurse- delivered intervention, it also aligns with NINR’s strategic focus area—Wellness: Promoting Health and Preventing Illness (16-NR-778). A prevention-focused adolescent health service in LMICs can set the tone for a ‘culture of health and wellness’ during this critical developmental period.

NIH Research Projects · FY 2025 · 2021-03

Many subsets of men who attend clinical STD services do not receive gonorrhea testing, contributing to delayed treatment and potentially amplifying antimicrobial resistance. Poor test uptake is often related to costs associated with STD testing and minimal community engagement. We have developed a pay-it-forward intervention to enhance gonorrhea testing among men who attend clinical services. Pay-it-forward involves an individual receiving a gift (in this case, a gonorrhea test) and then be provided the opportunity to give a gift (in this case, money to support gonorrhea testing) to another person. Pay-it-forward is an example of upstream reciprocity theory suggesting that people who are helped by someone feel a “warm glow” that makes them more likely to help others. Pay-it-forward has been used to encourage people to donate something to another person. But in this study, we will test the effectiveness of two levels of implementation strategies based on upstream reciprocity to motivate gift recipients to get tested for gonorrhea. Specifically, we will compare a standard pay-it-forward implementation strategy with minimal encouragement to get tested and a community-engaged pay-it-forward strategy alongside a control arm in which men pay for their own STD test. Our pilot randomized controlled trial in two cities found that pay-it-forward substantially increased gonorrhea testing compared to the control arm (56% in the pay-it-forward arm, 18% in control arm). Our pilot data also suggests that greater community engagement (e.g., participatory activities to create testing messages) increases donations to the program, reinforcing the hypothesis that a community-engaged implementation strategy can activate upstream reciprocity. Intensified testing could also decrease antibiotic resistance. Building on our UNC research infrastructure, we propose the Pay-It-forward gONorrhEa tEsting RCT (PIONEER). The study has the following specific aims: (1) to use a three-arm cluster randomized controlled trial to compare gonorrhea testing uptake in a standard pay-it-forward strategy arm, a community-engaged pay-it-forward strategy arm, and a control arm reflecting current practice; (2) to determine mechanisms through which different pay-it-forward strategies activate upstream reciprocity to influence gonorrhea testing and donations; (3) to examine the impact of enhanced gonorrhea testing on the development of gonorrhea resistance. Based on our prior work, we hypothesize that a community-engaged pay-it-forward strategy will increase gonorrhea test uptake rates compared to the standard strategy, and that pay-it-forward in general is superior to the current practice of paid testing. Our proposal is innovative because it provides a new financing mechanism to support gonorrhea testing, tests implementation strategies to enhance the effectiveness of this mechanism, and will provide more detailed information about the relationship between increasing testing and the development of antimicrobial resistance. This proposal has significant public health implications for the delivery of sexual health services in low and middle-income countries and is aligned with the mission of NIAID.

NIH Research Projects · FY 2026 · 2021-03

The world witnessed a global pandemic caused by a second severe acute respiratory syndrome coronavirus (SARS2). In spite of the foreshadowing of such a pandemic by the emergence of SARS1 in 2002 and Middle East respiratory syndrome coronavirus (MERS) in 2012, we were ill equipped to address this scourge. Each of these early outbreaks yielded a substantial body of knowledge on the structures of coronavirus proteins. As with previous outbreaks, we are witnessing a redoubled coronavirus research effort. Structural biology continues to lead the way; however, our laboratory is now pledging a sustained commitment to elucidation of the fundamental enzymology and corresponding mechanisms of coronavirus genome replication. The SARS2 replisome has emerged as a clinically tractable target for development antiviral therapeutics. Remdesivir is a nucleotide analog prodrug, metabolized to the triphosphate in cells, and incorporated by the SARS2 replisome without excision by its proofreading exonuclease, ExoN. The mechanism of action of remdesivir is unclear, and the mechanism of escape from ExoN is unknown. This circumstance reflects the absence of a quantitative, mechanistic perspective of the SARS2 replisome. Such a perspective will be essential to elucidation of the mechanism of drug action and the mechanism of drug resistance. We have demonstrated the feasibility of elucidating the principles governing the dynamics and function of the SARS2 core replicase using state-of-the-art ensemble and single-molecule approaches. We will exploit these advances to pursue the following specific aims: study assembly and function of the SARS2 core replicase and its sub-assemblies (Aim 1); study utilization of incorrect nucleotides and nucleotide analogues by the SARS2 core replicase (Aim 2); and study the mechanism of error correction by the SARS2 exoribonuclease (Aim 3). Completion of these studies will represent the first, deep dive into the mechanistic enzymology of the coronavirus replisome.

NIH Research Projects · FY 2026 · 2021-03

ABSTRACT Primary glomerular diseases often occur in patients with prexisting diabetes mellitus, a common and systemic illness which increases the risk for infection, cardiovascular and progressive chronic kidney disease. The diagnosis, pathology, treatment, complications and outcomes of primary glomerular diseases may be modified by the risks associated with diabetes, however, the literature is greatly lacking in this area. Such information is crucial to optimizing beneficial therapies and minimizing complications and cost in the care of glomerular disease patients. CureGN-Diabetes is a prospective, longitudinal observational study of patients with one of the four most common primary glomerular diseases: IgA nephropathy (IgAN), focal segmental glomerulosclerosis (FSGS), minimal change disease (MCD), and membranous nephropathy (MN). It is an ancillary study to the parent CureGN study which is currently in its second cycle of NIH/NIDDK funding and has enrolled 2400 patients with IgAN, FSGS, MCD and MN, but excluded patients with diabetes prevalent at the time of glomerular disease diagnosis. The exclusion of diabetes had an unexpected and disproportionate effect on recruitment among minorities and sites in the southeastern U.S., as these populations suffer among the highest rates of diabetes. CureGN-Diabetes will fill this gap in the CureGN study and will also leverage control data from the TRIDENT Study, a longitudinal cohort of patients with biopsy proven diabetic glomerulosclerosis. Harmonization of data from CureGN, TRIDENT and CureGN-Diabetes, will provide an opportunity to study the effects of primary glomerular disease alone, diabetic glomerulosclerosis alone and the combined effects of primary glomerular disease AND diabetes. The aims of CureGN-GN Diabetes include: 1) To recruit and follow a multiethnic cohort of 300 adult patients with diabetes mellitus and biopsy documented IgAN, FSGS, MN, and MCD; 2) To assess the impact of diabetes on presenting features (clinical and histopathologic) and long term outcomes in patients with IgAN, FSGS, MN, and MCD; AND 3) To identify individual and clusters of morphologic lesions which carry diagnostic and prognostic value. A strength of this study is the significant overlap between investigators and clinical sites involved in the CureGN and TRIDENT studies, which will expedite the efficiency and productivity emanating from this work. Furthermore, the CureGN-Diabetes Study will add to the available biorepositories of CureGN and TRIDENT, enriching the resources for future biomarker and mechanistic studies of primary glomerular disease versus diabetes-mediated pathways

NIH Research Projects · FY 2025 · 2021-03

ABSTRACT Proper control of stem cell division is critical for tissue morphogenesis and homeostasis. When dysregulated, it can lead to hypoplasia and stem cell exhaustion on the one hand, or tissue overgrowth and cancer on the other. But mitosis is more than simple proliferation, as cell division can be controlled not only in time but also in space. Oriented cell divisions (OCDs) are an example of the latter, and for stem and progenitor cells, choices between division axes can dictate cell fate outcomes and impact tissue architecture. In stratified epithelia such as the epidermis, basal progenitors divide either within the plane of the epithelium, or perpendicular to it. Evidence suggests that planar divisions are generally self-renewing symmetric cell divisions (SCDs) while perpendicular divisions are differentiative asymmetric cell divisions (ACDs). Previous work from our lab has shown that ACDs are directed by a complex of polarity and spindle orientation proteins—converging on the critical scaffolding protein LGN (Gpsm2)—which localize asymmetrically at the apical cell cortex. More recently, we have found that the paralog AGS3 (Gpsm1) seems to oppose LGN, and functions in promoting SCDs through an unknown mechanism. In addition, we recently made the surprising discovery that division orientation is not fixed during metaphase, as previously thought, but can be further refined during late stages of mitosis. In this process, which we term “telophase correction,” roughly one-third of basal cells enter anaphase at oblique angles, but then reorient to either planar or perpendicular. We have learned that cell-cell adhesions—specifically, the mechanosensing components of the adherens junction—are important for telophase correction to occur, and can operate independently of LGN. This demonstrates that in addition to intrinsic cues such as the LGN complex, extrinsic factors such as the local tissue microenvironment influence the final division axis. Despite what we and others have learned about the molecular control of ACDs, major knowledge gaps exist in understanding how oriented divisions shape tissue architecture both during normal development and in congenital skin diseases such as epidermolysis bullosa and ectodermal dysplasia. Specifically, the objectives of this proposal are to develop a better understanding of 1) what regulates SCDs and how the choice between SCD/ACD is made (SA1), 2) how cell-cell adhesion, cell-matrix, and local cell density impact division orientation and fate decisions (SA2). To achieve these goals, we will leverage a combination of innovative approaches, centered on our rapid, high-throughput technique—lentiviral ultrasound-guided gene inactivation and gene expression (LUGGIGE)— which we will utilize to generate mouse models of both gene loss and of specific mutations found in human diseases. Combined with ex vivo imaging of skin explants and in vivo proteomic approaches to characterize the LGN and AGS3 interactomes using TurboID, these comprehensive studies will provide insights into the cell- intrinsic and extrinsic cues that determine division orientation, and how they operate during normal epidermal growth and in blistering and dysplastic skin diseases.

NIH Research Projects · FY 2025 · 2021-03

Project Summary Lung and triple-negative breast cancers (TNBCs) are leading causes of cancer-related deaths in the U.S. This high mortality rate is largely due to their propensity to rapidly progress and metastasize. While targeted therapies for lung adenocarcinoma have improved overall survival, similar advances in lung squamous carcinoma (LUSC) and TNBC have been stagnant. However, for both cancer types, immune checkpoint blockade and/or angiogenesis inhibitors improves disease control. Thus, a more complete understanding of how vascular/immune niches within the tumor microenvironment (TME) promotes LUSC and TNBC will allow us to build upon these advances. Our teams have recently demonstrated that LUSC and TNBC promote tumor growth and metastases through a convergence on fibrin remodeling (Nature Communications, 2018; J Clinical Investigation, 2019) and activation of endothelial cell wound-healing programs (Oncogene, 2019). Fibrin(ogen) that escapes leaky tumor endothelial cells (TECs) acts as a scaffold for tumor cell motility and creates a provisional matrix for tumor progression. Using highly integrated bioinformatics and novel LUSC models, we recently found that CCL2- mediated recruitment of Factor XIIIA (FXIIIA)-expressing inflammatory monocytes (IMs) promotes fibrin cross- linking, metastases and poor survival in LUSC. We also found that TEC heterogeneity (TECH) directs fibrin accumulation through a TGFβ/miR-30c/PAI-1 signaling axis - TECs with high levels of the fibrinolysis inhibitor, PAI-1, increase perivascular fibrin networks that support sprouting angiogenesis and tumor progression. We have also uncovered a secondary connection between fibrin remodeling and the RNA-binding protein Quaking (QKI) which is enriched in TECs and drives tumor angiogenesis. Silencing QKI in TECs inhibits sprouting angiogenesis and metastases, but micro-vessel density (MVD) paradoxically increases; which we posit is due to CXCL12-mediated sequestering of CXCR4+ IMs that initiate fibrin cross-linking. Based on these collective new insights from our groups, we hypothesize that (i) inhibition of TEC QKI initially blocks tumor angiogenesis and metastasis, however, a CXCL12-mediated retention of FXIIIA+ IMs promotes fibrin remodeling and rebound angiogenesis. Additionally, we propose (ii) that TECH drives the formation of aberrant and persistent perivascular fibrin scaffolds in LUSC and TNBC via a spectrum of TGFβ/miR-30c/PAI-1 expression. The objective of this proposal is to elucidate how fibrin remodeling and tumor progression depend on the heterotypic relationships between TECs and IMs, and the heterogeneity amongst TECs within the TME.

NIH Research Projects · FY 2026 · 2021-03

ABSTRACT This project, developed in response to RFA-AG-21-008, describes core plans for data collection and dissemination of the sixth wave (Wave VI) of the National Longitudinal Study of Adolescent to Adult Health (Add Health), when cohort members will be 39-48 years of age (mean 44). Add Health is a longitudinal study of a nationally representative sample of over 20,000 adolescents who were in grades 7-12 during the 1994-95 school year and have been followed for five waves to date. Over 25 years, Add Health has collected rich demographic, social, familial, socioeconomic, behavioral, psychosocial, cognitive, and health survey data from participants and their parents; a vast array of contextual data from participants’ schools, neighborhoods, and geographies of residence; administrative data linked to participants, including birth and death certificates; and in-home physical and biological data from participants, including anthropometric measures, genetic markers, blood-based assays, and medications. Ancillary studies have added more information, including epigenetic, gene expression, and microbiome data. Thus, Add Health is exceptionally unique because it has a rich, multi- level, longitudinal array of data for a large nationally representative cohort of Americans who are entering midlife. Importantly, the overall health profile of the cohort as they make the transition to midlife is problematic across many dimensions. Moreover, health disparities by race, ethnicity, socioeconomic status, and rural- urban residence in this cohort are wide and, in some cases, widening. As such, rich longitudinal, multi-level, and nationally representative data are urgently needed to best understand the life course determinants of health trajectories and health disparities of US adults as they enter midlife. Wave VI of Add Health will fill this critical need. The overall goal of Wave VI of Add Health is to collect and disseminate the comprehensive data needed to best understand the social, economic, psychosocial, contextual, and biological determinants of health trajectories and disparities among this nationally representative cohort of Americans as they age into midlife. The project is focused around five aims: 1) Re-interviewing cohort members using predominantly web- based and in-person modes, with explicit attention to securing high response rates from all racial/ethnic and socioeconomic status groups; 2) Enriching study content in key domains that will elucidate mid- and later-life health trajectories and disparities; 3) Re-visiting cohort members who consent for an in-home health exam that includes venous blood collection and other important components of health; 4) Assaying biological specimens for important pre-disease and disease biomarkers; and 5) Cleaning, documenting, disseminating, archiving, promoting, and supporting Wave VI data for the scientific community. This project has extraordinary potential to contribute to the science of aging, health, and health disparities for decades to come, as the Add Health cohort ages into the middle adult years and beyond. Successful carryout of this project will supply essential data for thousands of researchers working on these critical issues.

NIH Research Projects · FY 2026 · 2021-03

Alzheimer’s disease (AD) causes progressive loss of memory, for which there is no cure. A growing understanding of memory impairment in AD suggests that alterations at the genetic and cellular levels contribute to circuit dysfunction. By restoring these vulnerable circuits and networks during early stages of the disease, it might be possible to reverse memory loss and slow disease progression. Recent studies have shown that GCs receive abundant subcortical inputs from the lateral supramammillary nucleus (SuM) of the hypothalamus, and the SuM inputs play a key role in modulating the strength of the EC inputs onto GCs. Importantly, we found that SuM is spared of Aβ deposition even in late-stage 5xFAD mice and exhibits intact anatomical inputs to GCs in early 5xFAD mice, thus placing SuM in an ideal position to modulate impaired EC-DG connections in AD. In addition, we found that stimulating SuM neurons or SuM-DG pathway is sufficient to increase GC activity and improve spatial memory performance during the NPR test through SuM glutamate release, suggesting that increased SuM glutamate transmission is beneficial to spatial memory performance. Furthermore, using an in vivo multi-fiber photometry recording system, we found highly correlated Ca2+ activities between SuM neurons and DG GCs during spatial memory retrieval in the NPR test, which is disrupted in early 5XFAD mice, suggesting that synchronized SuM-DG activity is critical for spatial memory retrieval. Based on these findings, we propose the following three aims to evaluate whether stimulation of SuM-DG activity can restore memory and slow disease progression. In Aim 1, we will determine whether stimulating SuM-DG activity in early AD mice restores EC-GC connections and DG activity; In Aim 2, we will determine whether stimulating SuM-DG activity restores interregional synchrony during spatial memory and improves memory performance in early AD mice; and In Aim 3, we will evaluate whether chronic stimulation of SuM neurons from the prodromal stage slows AD progression. Our proposed experiments will advance our understanding of the key neural circuits that modulate vulnerable networks during early AD pathology. Results from these studies will guide new therapeutic directions by selectively targeting these modulatory neurocircuits for treating memory loss associated with AD.

NIH Research Projects · FY 2026 · 2021-02

The NIH Helping to End Addiction Long-term Initiative, an aggressive effort to speed scientific solutions to stem the national opioid public health crisis, includes a broad range of programs and projects generating large volumes of diverse data. HEAL Initiative awards have been made with the expectation that awardees’ results and data will be shared with broader research communities and the public to promote dissemination of new knowledge and prompt further discoveries. A key principle underlying the HEAL data strategy is to make those data findable, accessible, interoperable, and reusable (FAIR) for the benefit of researchers and the public. We propose to serve as the HEAL Data Stewardship Group to provide guidance and support to HEAL investigators on readying their data to connect to the HEAL Platform, a secure data access and computing environment. To begin work, we will engage with HEAL Platform developers, HEAL investigators, and others in the research and broader communities to understand the most valuable uses for the HEAL Platform and support those “use cases” through data management efforts. We will then collaborate closely with HEAL Platform developers and HEAL investigators to augment data management efforts where needed, especially for programs not currently working with their own HEAL-funded data-coordinating efforts. Through collaboration with HEAL investigators and an exploration of the work being conducted across the initiative, we will provide long-term data strategy and sustainability guidance to the HEAL Initiative. This will facilitate data access through the HEAL Platform well into the future. Together with the HEAL Platform awardees, our efforts will ensure secure, straightforward access to HEAL results and data to maximize the impact the HEAL Initiative has on addressing opioid misuse, overdose, chronic pain, and related challenges.

NIH Research Projects · FY 2025 · 2021-02

ABSTRACT/PROJECT SUMMARY Background: Histoplasma is a pathogenic fungus that causes life-threatening lung infections. About 500,000 people are exposed to Histoplasma each year in the United States, and over 60% of the US population has been exposed to the fungus at some point in their life. We have shown that Histoplasma is composed of at least five different species that vary considerably in the type and magnitude of disease they cause. Broad, long-term objective: The proposed research will help us identify the genes that allow virulence to emerge and spread, as well as develop a panel of isolates that once deep-sequenced can be used by the community of medical mycologists to map any trait of interest in Histoplasma. The objective of this proposal is to discover whether the genes responsible for differences in virulence among isolates are similar across species. Specific aims: Aim 1 of the study proposes to generate genetic reference panels for three species of the human pathogen Histoplasma. We will use this resource to identify alleles involved with virulence differences within and between species. Aim 2 will genetically test the phenotypic effects (i.e., virulence in vitro and in vivo) of the genomic hypotheses produced in Aim1. Aim 3 will study the spread of alleles in clinical samples over a period of 40 years and will integrate the results from Aims 1 and 2, allowing us to determine whether any of the alleles involved in virulence have increased in frequency. Method: This haploid organism is ideal for the laboratory study of fungal pathogens, and it is well-suited for genomic analysis. We will generate genetic reference panels for three different species of Histoplasma with state-of-the-art genomic tools and genome-wide association mapping. We will use this panel to identify the genetic basis of virulence differences within isolates of the same species. Notably, we will generate an online portal to analyze GWAS data, a first in the medical mycology community. Preliminary results show that given the amount of phenotypic variance in virulence, our approach and proposed sample sizes make this project feasible. Validation of candidate virulence genes will be undertaken according to established cell culture and mouse infection assays. Our approach will generate tools and reference panels for the fungal genetics community. Health-relatedness: The disease burden caused by Histoplasma species is substantial in the United States, with a conservative estimate of at least 3.4 cases per 100,000 population. If infectious strains can transmit the ability to cause infection to less harmful strains through gene exchange, the potential future disease burden will grow as global trade, travel and climate change bring new species of the fungus into overlapping geographic regions. The proposed research will identify what loci are involved in the evolution of virulence and will study the influence of natural selection in their evolution in recent timescales. This application is in response to a recent NIH Funding Opportunity Announcement (PA-19-082) supporting research on histoplasmosis and two other endemic fungal diseases, and this program specifically encourages submission of R01 applications that will “expand understanding of speciation and impact on clinical outcome.”

NIH Research Projects · FY 2025 · 2021-02

PROJECT SUMMARY This overall goal of this mentored patient-oriented research career development award is to improve our understanding of pouchitis and other inflammatory conditions after restorative proctocolectomy with ileal pouch-anal anastomosis (IPAA). Dr. Barnes is a motivated clinical researcher at the University of North Carolina at Chapel Hill (UNC), with a specific interest in the study of inflammatory bowel diseases. Over the next five years, Dr. Barnes will work with his mentorship committee to continue his progression towards his goal of independence. His mentorship committee includes experts in gastrointestinal epidemiology and comparative effectiveness (Kappelman), patient reported outcomes [(PROs) DeWalt], and precision medicine (Kosorok). Each member has an established track record of mentoring junior faculty, consistent peer-reviewed support, and high research productivity. Dr. Barnes’s career development objectives are as follows: 1) develop a new skillset in comparative effectiveness research using real-world data; 2) obtain knowledge and experience in the development of PRO assessments; 3) acquire new skills in machine learning and precision medicine principles which will allow for the incorporation of clinical and microbiota data into new risk modeling for patients undergoing restorative proctocolectomy with IPAA, and 4) transition to independence. To achieve his career development objectives, Dr. Barnes will participate in structured coursework, conduct mentored research, and will participate in workshops through the North Carolina Translational and Clinical Sciences Institute, including the R-Writing Group. The research and training environment at UNC is well established. For fiscal year 2019, UNC ranked tenth among both private and public universities nationwide for National Institutes of Health (NIH) research funding to domestic institutions of higher education. The specific aims of this research project are 1) to evaluate the comparative effectiveness of specific biologic therapies for the treatment of chronic inflammatory conditions of the pouch after IPAA; 2) to develop PRO items for evaluating quality of life and specific symptom domains among patients after IPAA. Techniques for PRO development will include identifying symptom domains relevant to patients after IPAA using qualitative research methods, selecting candidate items for IPAA-specific PROs using existing PRO instruments where appropriate, and performing cognitive interviews with patients after IPAA to ensure content validity; and 3) to demonstrate novel methods for identifying predictors of chronic pouchitis using a machine learning approach. Expanding an existing prospective cohort, Dr. Barnes will utilize the stool microbiome, laboratory data and clinical variables to create signatures for identifying predictors of chronic pouchitis. The combination of mentorship, didactics, and research experiences offered through this K23 proposal will allow Dr. Barnes to develop a unique skillset that will aid in his ultimate transition to independence as a clinical investigator and research scientist.

NIH Research Projects · FY 2026 · 2021-02

ABSTRACT Remarkable clinical responses have been reported in B-cell malignancies by adoptive transfer of T cells redirected with a chimeric antigen receptor (CAR) specific for the CD19 antigen. However, developing CAR-Ts for the treatment of solid tumors including ovarian cancer (OC) is challenging because: (1) OC-associated antigens that are targetable by CAR-Ts are limited, generally not exclusively expressed by OC, and act as passengers, not as drivers of tumorigenesis, allowing for antigenic drift; (2) OC tumor microenvironment (TME) is highly immunosuppressive. In this proposal we aim at solving these critical issues. We have identified B7-H3 (CD276) as a suitable target for chimeric antigen receptor (CAR) T cells in OC. B7-H3 is a tumor-promoting transmembrane protein aberrantly expressed in 60% to 93% of pancreatic cancer, melanoma, leukemia, breast, prostate and OC, while limited expression is seen on normal healthy tissues. We have developed and tested B7-H3.CAR-Ts in xenogeneic and immunocompetent tumor models showing antitumor activity is several tumor models including OC and safety. Thus in Aim 1 we propose to conduct a phase I clinical study in patients with OC to assess safety and antitumor activity of autologous B7-H3.CAR-Ts inoculated intraperitoneally. An IND (IND19641) for this study has been obtained at University of North Carolina, and clinical grade reagents to manufacture B7-H3.CAR-Ts are in hands. In Aim 2 we propose to conduct a comprehensive immunologic analysis of tumor biopsies and ascites collected from patients enrolled in the study before and after treatment to assess antigen loss and immunologic perturbation of the TME in OC. In Aim 3, we propose to reprogram tumor-associated macrophages (TAMs) and myeloid derived suppressor cells (MDSC) of the OC TME to a non-immunosuppressive state by using potent and orally bioavailable TAM RTK small molecule inhibitors developed at University of North Carolina (IND128236). We will thus perform preclinical studies to evaluate whether TAM RTK signaling inhibition in macrophages and MDSC would favor the antitumor activity of B7-H3.CAR-Ts in a syngeneic model of OC. If successful, this strategy will be included into a second phase of the proposed Phase I clinical study with B7-H3.CAR-Ts.