Grants

NIAID - National Institute of Allergy and Infectious Diseases

Abstract Malaria is a leading killer of children worldwide, and existing WHO-recommended vaccines offer only modest protection that is of short duration. Malaria vaccine innovation has been impeded by our limited knowledge of the antigens and epitopes targeted by T cells and the characteristics of malaria-specific CD4 T cells that are most critical for immune protection. We will address these critical gaps by identifying hundreds of novel P. falciparum CD4 T cell epitopes and using innovative T cell assays to characterize epitope-specific T cells at the individual cell level. P. falciparum has a complex life cycle, and the human immune response differs according to the stage of parasite development. The most successful malaria vaccine strategies identified to date require vaccination with live attenuated sporozoites, which infect hepatocytes but undergo developmental arrest at the liver stage and do not establish blood-stage parasitemia. Evidence from both human and animal studies of sporozoite vaccination indicates that CD4 T cells targeting liver-stage parasites are critical for this protection, but the antigens targeted by protective T cells are almost entirely unknown. In preliminary studies, we have developed a bioinformatic strategy to identify P. falciparum antigens transcriptome-wide that are highly and selectively expressed during the liver stage of infection. In the first aim, we will use a novel assay to generate large DNA-barcoded P. falciparum peptide libraries, including geographically relevant sequence variants, and perform highly multiplexed screens to identify peptides that bind HLA-DRB1 allotypes common in East Africa. We will then test the recognition of candidate epitopes using samples from a large cohort of highly malaria- exposed Ugandan children. In the second aim, we will use the peptide:HLA binding data to build a 1000-plex DNA-barcoded multimer probe-set to label cells for single-cell sequencing, which will enable us to simultaneously discern the epitope specificity, HLA restriction, full transcriptomes, and TCRα:β sequences of P. falciparum-specific CD4 T cells. We will use this novel genomic assay platform to compare the transcriptional and functional phenotype of CD4 T cells targeting liver- vs. blood-stage epitopes in children residing in a highly malaria-endemic region of Uganda. Lastly, we will use a unique set of samples from a randomized trial of childhood chemoprevention to test the hypothesis that selective suppression of blood-stage infection by antimalarial drugs fosters the development of a broader and more functionally robust CD4 T cell response to liver-stage antigens. Together, these novel assay platforms will enable us to identify hundreds of novel P. falciparum epitopes recognized by malaria-exposed Ugandan individuals and characterize the responding CD4 T cells in unprecedented detail to uncover the functional and phenotypic features critical for T cell-mediated protection.

NCI - National Cancer Institute

The field involving antibody-dependent cellular cytotoxicity (ADCC) and complement-mediate cytotoxicity (CDC) activities mediated by therapeutic antibodies is referred to as humoral immuno-oncology (HIO). Tumor-produced factors that may have immunosuppressive effects on ADCC and CDC activities are called HIO factors. We screened several HIO factor candidates (HIO-Xs) based on their production/secretion by various tumor types. HIO-Xs can be serum biomarkers representing proteins shed by the tumor. Using a set of optimized HIO assays which we developed, two new HIO factors were found to be immunosuppressive and were named HIO-2 and HIO-3. The focus of this proposal is on the characterization of HIO-2. HIO-2 is a mucin protein produced and over-expressed in breast, lung, ovarian, endometrial, bladder, and gastrointestinal cancers. HIO-2 is present in a membrane-bound form as well as a soluble/shed form. Preliminary data suggest that soluble HIO-2 binds to CD16a/Fc gamma receptor, blocking antibody binding, and ultimately inhibiting ADCC activity of trastuzumab and pertuzumab due to reduced antibody/Fc receptor engagement. HIO-2 blockade of CD16a function creates an immunosuppressed tumor microenvironment potentially impacting therapeutic antibodies activity. In fact, elevated serum levels of HIO-2 correlate with worse progression-free survival after treatment with trastuzumab. The goal of our program is to generate and test compounds that can overcome immunosuppression mediated by HIO-2. This proposal focuses on the initial tasks of an overarching approach aimed at bringing HIO-2 antagonists into clinical testing after appropriate discovery and preclinical validation. In Year 1, we will be answering the question: Can a pharmaceutically viable HIO-2 antagonist candidate be developed? In Aim 1, effort will be allocated to defining the HIO-2/CD16a binding interface which will guide future structure-activity relationship analyses of HIO-2 antagonists. In Aim 2, an expanded natural compound library will be screened to identify additional HIO-2 antagonist candidates having improved activity profile. Lead candidates and their analogs will be tested in vitro for potential toxicity and metabolic stability. In Year 2, we will be answering the question: Can HIO-2 antagonist enhance the anti-tumor effect mediated by therapeutic antibodies in vivo? In Aim 3, the current lead HIO-2 antagonist, or any better lead(s) identified in Aim 2, will be assessed for oral availability in mice and compared to intravenous administration. The maximum tolerated dose of the lead antagonist will be assessed for the selected route (PO or i.v.). Finally, the lead HIO-2 antagonist will be tested alone and in combination with trastuzumab using a patient-derived xenograft (PDX) model of breast cancer, wherein this PDX has high levels of HER2 expression, high levels of HIO-2, and is somewhat resistant to trastuzumab anti-tumor effect. The benefit of a HIO-2 antagonist strategy extends to current and future therapeutic antibodies with immune effector function, including rituximab, daratumumab, cetuximab, and others.

NHLBI - National Heart Lung and Blood Institute

Project Summary Neutrophils are major players in the innate immune system, known for their role in fighting infections by engulfing and destroying invading pathogens, including those in the lungs. Neutrophil heterogeneity is a key component in innate immune regulation. Distinct mature neutrophil subpopulations, defined by their signature gene expression profiles, phenotypes, behaviors, and functional states, exist even prior to encountering pathogens. Understanding the unique features and origins of these functional subpopulations will enable the effective and safe manipulation of their specific functions for personalized medicine. The objective of this proposed project is to investigate the origin and characterize the functional state of the lung interstitial neutrophil (LIN) subset. Preliminary scRNA-Seq analysis revealed a tightly regulated neutrophil reprogramming trajectory during lung infection and inflammation, based on gene expression patterns. The preliminary studies identified the functional heterogeneity of neutrophils across three different lung compartments: the vasculature, the interstitium, and the alveoli, and discovered that neutrophils lodged in the interstitium for an extended period before migrating to the alveoli. Based on these findings, it was hypothesized that transcriptional reprogramming leads to the formation of a specialized LIN subset defined by its unique gene expression profile. These neutrophils are highly specialized, with significant bactericidal activity, making the lung interstitium a primary site for bacterial elimination. Accordingly, expanding the LIN population could be a viable therapeutic strategy for enhancing neutrophil-mediated host defense, while potentially detrimental in sterile inflammation-induced acute lung injury. To advance the understanding of the function, fate, pathophysiological role, and origin of LINs and explore their therapeutic potential, three specific aims will be investigated: Aim 1 will characterize the functions and fate of LINs. In addition, neutrophil- mediated bacterial killing in the interstitium will be visualized and confirmed by fluorescent imaging. Aim 2 will elucidate the pathophysiological role of LINs in lung infection and inflammation. The LIN population will be expanded by inhibiting CCR2-mediated neutrophil transepithelial migration, and its effect on host bactericidal activity and lung injury will be tested in a neutropenia-related bacterial pneumonia model and an acid aspiration-induced acute lung injury (AA-ALI) model. Aim 3 will further investigate the microenvironment- induced transcriptional reprogramming that confers specialized functions to LINs. Additional scRNA-Seq analyses will be performed to gain a deeper understanding of neutrophil reprogramming in the interstitium and to reveal potential temporal and stimulus-specific reprogramming schemes. Together, the results from this study will establish LINs as a specialized neutrophil subpopulation with unique phenotype and function, and as a novel therapeutic target for treating lung infections and inflammation.

Fish and Wildlife Service

THIS IS A NOTICE OF INTENT TO AWARD TO A SINGLE SOURCE, AND THERE IS NO FULL ANNOUNCEMENT ASSOCIATED WITH THIS NOTICE. The U.S. Fish and Wildlife Service, Southeast Region, intends to award on a sole source basis one cooperative agreement each to Louisiana State University (LSU) and the University of Southern Mississippi (USM) to support proposed work titled: Characterization of Microhabitat Association of Fish Assemblages in the Pearl and Bogue Chitto Rivers. A determination by the Government not to compete this proposed assistance based upon Departmental Manual 505DM2, paragraph 2.14 (B) (2) is solely within the discretion of the Government. Questions concerning this announcement should be directed to Nijua Heard (404) 679-4146 or by email at nijua_heard@fws.gov

NIAID - National Institute of Allergy and Infectious Diseases

Since 2022, DCR and LBR have conducted monkeypox treatment trials in the Democratic Republic of Congo. This monkeypox Characterization non-severable task order complements this prior research given DCR/LBR further analyses of the PALM007 RCT results highlighted remaining monkeypox knowledge gaps, specifically related to the risk of death or severe disease from clade I monkeypox. In PALM007, patients who presented with flat/malignant lesions (as opposed to the classically appearing and evolving lesions) had an increased risk of mortality: 7/7 (100%) of those with flat lesions died compared with 3/590 (0.5%) of the remaining study participants. Patients presenting with thousands of lesions experiencing mild/moderate disease and others with prolonged disease after presenting with few lesions. It is suspected that this is likely related to host-related risk factors, however, researchers are unable to evaluate this fully retrospectively with the samples available.

NIAID - National Institute of Allergy and Infectious Diseases

PROJECT SUMMARY/ABSTRACT: Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is a group of systemic autoimmune dis- eases characterized by organ- and life-threatening inflammation of small and medium blood vessels. Half of AAV cases are driven by ANCAs targeting neutrophil cell-surface bound proteinase 3 (PR3), which leads to patho- genic neutrophil activation. However, total PR3-ANCA plasma levels correlate poorly with disease severity and relapse risk, and limited knowledge of the variable immunopathogenic potential of distinct PR3-ANCA subsets has challenged the advancement of more accurate diagnostic and prognostic tests. Moreover, severe infection caused by current, broadly immunosuppressive treatment strategies is the leading cause of mortality in AAV. To this end, we have developed PR3-chimeric autoantigen-T cell receptor (CATCR)-T cells, a precision cellular immunotherapy which uses engineered receptors (PR3 fused to T cell receptor subunits) to specifically engage and eliminate pathogenic ANCA+ B cells through their B cell receptor (BCR). Initial CATCR-T cell designs demonstrated potent killing of ANCA+ B cells, though moderate killing of normal B cells indicated off-target bind- ing of CATCRs to other surface antigens. The overall goal of this proposal is to characterize the pathogenic features of PR3-ANCA on the monoclonal antibody level, and to increase PR3-CATCR selectivity by identifying and disrupting off-target binding partners through targeted mutagenesis of the CATCR PR3 binding moiety. In preliminary work, we have synthesized and expressed 18 novel patient-derived monoclonal PR3-ANCAs to identify shared features of pathogenic autoantibodies in AAV (Aim 1). Standard kinetic and competitive surface plasmon resonance (SPR) and ELISA assay formats will be used to measure PR3-ANCA binding affinity, epitope specificity, and their ability to disrupt PR3 binding with its neutrophil surface receptor NB1. Individual PR3-ANCA monoclonal autoantibodies will then be assayed for their ability to trigger pathogenic neutrophil activation in vitro to identify common determinants of pathogenicity. To mitigate off-target PR3-CATCR-T cells cytotoxicity (Aim 2), we will use BioID-mass spectrometry to identify non-BCR binding partners of PR3-CATCRs. Rounds of tar- geted mutagenesis informed by in silico modeling of protein-protein interactions will be validated using in vitro binding assays. Optimized PR3-CATCR-T cells using mutagenized PR3 as a binding domain will be tested for their selective killing in model human ANCA+ B cell lines and primary human B cells from patients with AAV. Overall, this proposal aims to i) advance our understanding of PR3-AAV immunopathogenesis with the goal of improving diagnostic and prognostic clinical tests, and ii) enable the development of PR3-CATCR-T cells, a novel cellular immunotherapy modality for the treatment of AAV. This research plan provides valuable training in im- munological wet lab techniques, cell culture modeling of rheumatic disease, protein engineering, and the pro- cessing and assaying of clinical samples. Findings from this proposal will be of broad interest to the immuno- therapy field and will serve as preliminary data for a future NIH K99 Career Development Award.

NHLBI - National Heart Lung and Blood Institute

Project Summary: This proposal seeks to study the relationship between STK38 kinase, Wnt signaling, and heart development, aiming to unravel STK38’s molecular mechanism with therapeutic implications for the treatment of congenital heart defects. The Wnt pathway plays a critical role in developmental processes such as heart development, and dysregulation of Wnt signaling has been linked to congenital heart defects. We have recently identified a new component of the Wnt pathway, STK38, and preliminary studies in zebrafish indicate that it plays a critical role in heart development. Through Aim 1, employing advanced techniques involving co-immunoprecipitation, mass spectrometry, and cellular labeling, I aim to identify STK38’s substrates within the Wnt signaling cascade. Characterizing these substrates and assessing their function will shed light on STK38's role as a regulator of Wnt signaling. Additionally, our preliminary studies suggest that STK38 interacts with Pygopus, a core nuclear Wnt component shown to play a role in vertebrate heart development, providing further evidence for the role of STK38 in Wnt signaling and cardiogenesis. This interaction will be characterized, which will aid in our understanding of STK38’s role in Wnt signaling. In Aim 2, I leverage induced pluripotent stem cells (iPSCs) as a powerful platform for modeling the progression from stem cells to cardiomyocytes. By monitoring endogenous STK38 levels during iPSC cardiac differentiation and correlating them with known periods of Wnt pathway activity, I will elucidate STK38's temporal expression dynamics during cardiac development. Furthermore, I will utilize iPSCs to chemically induce precise temporal depletion of STK38, enabling me to dissect STK38’s influence on Wnt signaling and cardiomyocyte differentiation/proliferation during critical periods of heart development. Through these complementary but non-overlapping aims, my research proposal seeks to advance our understanding of STK38's involvement in Wnt signaling and heart development. Successful completion of the grant holds promises for identifying novel therapeutic targets and enhancing our knowledge of congenital heart defects. In addition, this proposal will serve to greatly advance my knowledge and skills, propelling me forward toward my goal of becoming a successful academic researcher.

NCI - National Cancer Institute

PROJECT SUMMARY / ABSTRACT Human endogenous retroviruses (HERVs) comprise roughly 8% of our genome. Originally considered “junk DNA”, we now know that HERVs play diverse roles relating to human health, innate immunity, and cancer. Repetitive elements have been notoriously difficult to study and quantify due to their repetitive nature and the lack of a standardized database. In this project, I will leverage a comprehensive database of HERV loci (HERVdb) created in the Blanco-Melo Lab to accurately measure the expression of HERVs and investigate their role in clear cell renal cell carcinoma (ccRCC), a common and aggressive form of kidney cancer that accounts for 80% of renal cancer cases. HERV expression in ccRCC has been linked to tumor immunogenicity and response to immunotherapy, making it a promising area of research. Our hypothesis is that HERVs are abnormally transcriptionally activated in tumor cells, where they are recognized by the immune system due to absence of expression in normal cells; HERV products (RNA or protein) can thus prime the anti-tumor immune response, resulting in a HERV specific immune response. To test this, I will pursue two main aims: Aim 1: Define the unique signature of HERVs in ccRCC (clear cell Renal Cell Carcinoma). Aim 2: Explore the role of HERV-derived tumor-associated antigens across ccRCC samples. The goal is to establish a clear baseline of HERV expression in ccRCC, identify tumor-specific HERVs, and explore relevant HERV-derived peptides that could serve as potential therapeutic targets. Ultimately, this project aims to demonstrate that HERVdb can be used to identify HERVs as targets for ccRCC treatments, and to establish a reproducible workflow for the interrogation of HERVs in other cancers.

NINDS - National Institute of Neurological Disorders and Stroke

Abstract Spatial navigation requires the coordination of neural activity across many areas in the brain to integrate sensory, motor, and cognitive information. Through its interconnections with many of these areas, the retrosplenial cortex plays an important role in navigation by representing key aspects of the environment, sustaining memories, and translating between different perspectives in space. Many of these studies report the activity of individual neurons and lesioning of the entire area to evaluate function. However, it remains unclear which molecularly defined cell types within the area contribute to its function for navigation. This lack of investigation into cell types for navigation is notable given the repeated circuit motifs and distinct roles of cell types discovered throughout cortex for other functions. Our lab has identified a specific cell type, called Sst44 cells, in the posterior parietal cortex, an adjacent cortical area also involved in navigation, where its activity corresponds to corrections of navigational errors. I will build on this work to determine the role of Sst44 cells in retrosplenial cortex in mice during goal- directed navigation. My preliminary results suggest the activity of Sst44 cells in retrosplenial cortex may respond to a different aspect of navigational errors. Whereas the posterior parietal cortex has a privileged role in actions and egocentric processing of navigational information, the retrosplenial cortex contributes to the transformation of related signals to allocentric representations. Here, I propose two approaches to systematically determine the cell type’s function. In Aim 1, I will design virtual reality navigation tasks to quantify the activity of Sst44 cells in RSC in relation to egocentric and allocentric errors. In Aim 2, I will inhibit the activity of Sst44 cells to evaluate how the loss of their function impacts navigation and representation of spatial information in other cell types. Addressing these aims will bring unique precision in our understanding of a cell type’s contribution to navigation.

NIAID - National Institute of Allergy and Infectious Diseases

PROJECT SUMMARY Bacteroides fragilis is a gram-negative intestinal commensal that comprises up to 1% of total gut bacteria. While often considered harmless, this microbe remains a leading cause of anaerobic sepsis and abdominopelvic infections. The rise of antimicrobial resistance (AMR) in B. fragilis clinical isolates has complicated treatment of these infections. In particular, a distinct clade of B. fragilis strains (clade II) that represent 10% of B. fragilis from human clinical isolates were found to harbor cfiA, a genomically encoded metallo-β-lactamase. Carbapenem resistance in these strains is concerning not only due to the lack of available treatment options but also the risk of transmission to other microbes. Recent studies have also identified differences in average nucleotide identity between clade II and clade I—the remaining 90% of B. fragilis. This suggests that these clades might actually represent two different species, and thus, biology of clade II cannot simply be assumed from clade I. As all B. fragilis niche acquisition studies to date have utilized clade I strains, the mechanisms by which a clade II strain comes to dominate and persist within the colonic niche remain unknown. Previous microbiome studies in adults have revealed predominance of a single B. fragilis strain within an individual microbiota. Interestingly, no studies have identified a single host to harbor both clade I and clade II simultaneously, suggesting that intraspecific competition between clade I and clade II may influence niche establishment and strain dominance. Further studies are thus necessary to identify factors unique to clade II biology, as long-term colonization by these strains may engender a reservoir for carbapenem resistance within the host. Recent work has identified Bcf1 as a novel secreted toxin that is utilized by clade II in the context of interbacterial competition against clade I. This method of antagonism is dependent on an interaction with Bat1—a confirmed functional target of Bcf1 and a putative TonB-dependent transporter. Orthologs of Bcf1 can be found throughout Bacteroides spp., suggesting that this may be a novel family of toxins. However, the mechanism of Bcf1 activity remains unknown and all functional studies to date have only been conducted in vitro. The proposed studies will provide mechanistic insight into Bcf1 function (Aim 1) and its role in interbacterial competition in vivo (Aim 2). Aim 1 will test the hypothesis that Bcf1 binds to Bat1 and blocks translocation of an unidentified nutrient through Bat1 via a form of nutritional competition. Aim 2 will leverage various animal models to test the hypothesis that Bcf-mediated competition allows clade II to dominate its niche and achieve long-term colonization. These studies will enable us to consider functional links between interbacterial competition and high-level drug resistance from an ecologic perspective. Understanding the inherent differences in intraspecies biology and strain-level competition dynamics may inform the development of microbiome-targeted therapies to prevent colonization of pathogenic B. fragilis strains or translate to future consideration of AMR risk during clinical treatment of B. fragilis infections.

NICHD - Eunice Kennedy Shriver National Institute of Child Health and Human Development

PROJECT SUMMARY/ABSTRACT Motivation: Sensitive periods are temporal windows of elevated neuroplasticity, facilitating adaptation to and learning from early environments across a range of domains. In non-human animal models, excitation/inhibition (E/I) balance, or the expression of the primary excitatory (glutamate) to inhibitory (GABA) neurotransmitters, governs the degree of neuroplasticity throughout the sensitive period. Until recent methodological advances, measuring E/I balance minimally invasively in the human brain was infeasible, and thus our understanding of the role of E/I balance in human sensitive period neurobiology is extremely limited. Normative individual and clinical phenotypic differences exist in learning outcomes supported by sensitive periods. Research in non- human animal models suggests a range of clinical outcomes may result from differences in sensitive period neuroplasticity. Therefore, the proposed research will have a broader impact by first furthering understanding of normative sensitive period mechanisms, so future work can understand clinical deviations in these mechanisms. Here, we focus on an early visual cortical sensitive period because it is well-characterized and translatable across species. The proposed project investigates if E/I balance indexes sensitive period neuroplasticity in human infant visual cortical development as it does in non-human animal models. Methods: In this accelerated longitudinal study, infants will participate in visits at 2-to-4 months, 7-to-9 months, and 10- to-12 months. At all visits, infants will participate in a visual-evoked potential (VEP) electroencephalography (EEG) task as a readout of visual neurodevelopmental maturation throughout the sensitive period. Also at all visits, the MEGA-PRESS magnetic resonance spectroscopy (MRS) sequence will be collected from infants during natural sleep to extract concentrations of glutamate and GABA in a visual cortical seed. To measure visually-mediated cognitive outcomes, infants will participate in the cognitive scale of the Bayley Scales of Infant Development at the last visit. Aims: Aim 1 of the proposed project will characterize age-related changes in the MRS-derived E/I balance over the sensitive period using Generalized Additive Models for Location, Scale, and Shape. Aim 2 will relate individual differences in the MRS-derived E/I balance to the VEP readout of visual cortex function over the sensitive period using longitudinal multilevel modeling. Aim 3 will test if individual differences in the MRS- derived E/I balance from 2 to 9 months predicts emerging visually-mediated cognition at 10-to-12 months, at the end of the sensitive period, using longitudinal multilevel modeling. Training: To support the applicant’s goal of being an independent developmental neuroscientist using multimodal infant neuroimaging and to complete the proposed project, the applicant will develop expertise in 1) infant MRS, 2) advanced longitudinal modeling, and will 3) accelerate her professional development. The applicant is well-poised to complete this project and advance towards her career goals with a strong mentorship team, detailed training plan, and position at an R1 research institution.

NIAAA - National Institute on Alcohol Abuse and Alcoholism

PROJECT SUMMARY The ability to adaptively and appropriately respond to changes in the environment is a crucial behavior. It is cognitive flexibility that enables this adaptive responding. An inability to adaptively shift behavior (cognitive inflexibility) despite negative outcomes is observed in various psychiatric disorders and is a key diagnostic criterion of alcohol use disorder (AUD). The long-term objective of this work is to elucidate how chronic alcohol and stress promote AUD through cognitive inflexibility. In humans, AUD is accompanied by aberrant frontal cortex function alongside deficits in cognitive flexibility. A significant body of work, including that from our lab, has demonstrated impaired cognitive flexibility in animal models following chronic alcohol which can be exacerbated by stress exposure. However, the frontal cortical neuroadaptations that mediate these alcohol and stress induced cognitive impairments remain unresolved. This is a critical gap given that stress is a key risk factor in perseverative relapse for individuals with AUD. My central hypothesis is that functional network changes across the dorsomedial frontal cortex underlie cognitive flexibility impairments following repeated alcohol and stress exposure. Moreover, I hypothesize that changes in modulatory norepinephrine inputs to this region are critical contributors to these flexibility deficits. To test these hypotheses, I will investigate the mechanisms underlying the negative impact of chronic alcohol and stress on cognitive flexibility using a novel attentional set shifting task. In Aim 1 I will use in vivo calcium imaging to investigate alcohol- and stress- associated changes in dorsomedial frontal cortex dynamics underlying behavioral impairment. In Aim 2, I will determine whether chronic alcohol and stress functionally disrupts noradrenergic regulation of the dorsomedial frontal cortex using in situ hybridization readouts and pharmacological interventions to recover cognition. These proposed studies will provide ample training in cutting-edge techniques and facilitate my professional development in the alcohol research field. At the same time, this work will advance our understanding of the impact of long-term alcohol and stress exposure on cortical function and identify potential mechanisms for therapeutic interventions.

OD - NIH Office of the Director

ABSTRACT Older persons with HIV (PWH) face a higher risk, greater burden, and faster progression of age-related comorbidities, including cardiovascular diseases, metabolic disorders, renal dysfunction, obstructive lung disease, liver cirrhosis, and cancers, compared to those without HIV. These comorbidities lead to polypharmacy, adverse outcomes, and premature death. Understanding the cascade of comorbidity events and the natural history of these conditions among PWH is essential to guide secondary prevention and early interventions. The MACS/WIHS Combined Cohort Study (MWCCS) has significantly advanced HIV care by providing insights into HIV transmission, risk factors, and AIDS and non-AIDS outcomes among PWH using an interval cohort design. We propose to enrich this data source by establishing an electronic health record registry nested within the MWCCS (Aim 1) to study comorbidity burden and cascade among people with and without HIV. Furthermore, we will leverage existing cohort-wide biobehavioral data and the newly linked EHR data to develop predictive models of the comorbidity cascade among people with and without HIV (Aim 2). The proposed R21 work directly responds to PAR-24-092 by studying the aging and comorbidities of HIV using multidisciplinary approaches. Advanced clinical informatics tools, statistical learning models, and leveraging existing MWCCS infrastructure and biobehavioral measurements will shed light on aging and comorbidities in PWH. It will identify high-risk individuals prone to high comorbidity burden, thus paving the way to eventually be able to provide guidance on clinical decision making and care. Notably, this transformative initiative will serve as a pilot platform, leveraging MWCCS infrastructure, real-time EHR data, and existing cohort biomarkers. The data pipeline developed through this project will provide a scalable foundation for expansion to other MWCCS study sites in the near future, significantly broadening the scope and impact of ongoing research in HIV, aging, and comorbidity.

NIDDK - National Institute of Diabetes and Digestive and Kidney Diseases

Project Summary/Abstract: Inflammatory bowel disease (IBD) is a chronic inflammatory process initiated in the intestinal mucosa, typically onset in young adults. IBD causes severe morbidity and emergent complications often necessitating surgery, with a high proportion of IBD patients relapsing or being unresponsive to anti-TNF therapy. Despite the urgent clinical need for improved IBD treatments, pathophysiology of IBD remains incompletely understood, thought to result from complex genetic and environmental factors. Abnormalities in adaptive and innate immunity, cellular and ER stress, increased mucosal barrier permeability, dysbiosis, and genetic variants in autophagy and immune signaling components may contribute to increasingly pro-inflammatory gut immune dynamics, ultimately spreading to extra-intestinal manifestations in almost half of patients. Adaptive immunity, especially the Th17 T-cell subset converting to a pro-inflammatory phenotype, has been widely accepted as an important proponent of inflammation in IBD, with specific microbial strains and viral integration implicated as possible drivers. However, the antigen presentation landscape of T-cell epitopes has never been systematically characterized. In this research effort, we will profile antigen presentation by experimental immunopeptidomics, then computationally prioritize these HLA-presented peptides for immunogenicity, also examining molecular mimicry with self-antigens, to more deeply understand IBD pathophysiology (Aim 1). Adaptive immune responses can be triggered by ER stress, a biochemically complex phenomenon whose far-reaching role in IBD contributes to numerous pathologic features. Since evidence shows ER stress alters antigen presentation, we will attempt to draw a mechanistic link in the context of IBD, resolving specific antigen presentation changes induced by ER stress (Aim 2). Finally, we will attempt to validate immunogenicity of peptide epitopes by detecting antigen-specific T-cell receptors. Combining the abundant IBD resources in our biobanks and IBD Center with the expertise of an experienced, interdisciplinary team, we will endeavor to answer these important questions about adaptive immunity in IBD, while creating a high-quality dataset that lays the groundwork for future studies.

NIAID - National Institute of Allergy and Infectious Diseases

Project Summary Systemic lupus erythematosus (SLE) is a complex autoimmune disease driven by the breakdown of B cell tolerance and the production of autoreactive antibodies. We recently identified the presence of autoreactive B cells in spleens from healthy individuals and observed a 20-fold increase in spleens derived from SLE patients. This proposal will investigate a critical gap in understanding how autoreactive B cells transition from a tolerogenic state to a pathogenic one through: (i) analysis of dysregulated transcriptional pathways in spheromer-sorted autoreactive B cells derived from SLE patients, (ii) investigation of human-specific TLR7 mutants in mediating autoreactive B cell activation using immune organoids, and (iii) characterization of proinflammatory monocytes triggered by bacterial peptidoglycans in SLE. We will test the hypothesis that autoreactive B cells in SLE have distinct transcriptomic profiles and increased somatic hypermutation frequencies compared to healthy donors. We further hypothesize that amplified TLR7 signaling due to gene mutations and proinflammatory monocytes activated by bacterial peptidoglycans are pathognomonic for distinct SLE disease subtypes and characteristics. Aim 1 will characterize the transcriptomic and B cell receptor (BCR) profiles of autoreactive B cells from lupus patients. Aim 2 will investigate the autoimmune response in immune organoids expressing the TLR7Y264H mutant. Aim 3 will examine the ability of proinflammatory monocytes from SLE and healthy monocytes primed by peptidoglycan to trigger autoreactive B cell activation. The success of the proposed studies will elucidate the mechanisms by which autoreactive B cells become pathogenic and cause SLE, fundamentally transforming our understanding of SLE and potentially leading to novel therapeutic strategies.

NHLBI - National Heart Lung and Blood Institute

PROJECT SUMMARY/ABSTRACT As the aging population continues to grow worldwide, age-related chronic heart and lung diseases have become an increasingly serious public health issue. Consequently, understanding the relationships between aging and these diseases is crucial for effective prevention and treatment strategies. However, the complexity of the aging process, which involves cellular senescence, inflammatory responses, and oxidative stress, and its differential impact on individuals, makes it difficult to identify the relationship between specific aging processes and the development of various heart and lung diseases. The recent concept of "ageotypes," which clusters individuals into similar molecular profiles throughout the aging process, offers a framework to identify molecular mechanisms of aging and connect this with chronic heart and lung diseases. However, existing studies of ageotypes have been limited by small sample sizes, short follow-up periods, and insufficient omics data, limiting their overall utility. This proposal aims to generate multi-omic ageotypes to understand the molecular mechanisms of aging that lead to chronic heart and lung diseases. The overall objectives of this proposal are to: 1) develop and validate multi-omic ageotypes using metabolomics and epigenetics data in two large cohorts with electronic medical records (EMR) data; 2) identify the molecular drivers that significantly contribute to each ageotype, particularly in relation to heart and lung diseases; 3) elucidate the relationship between ageotypes and chronic heart and lung diseases. Through personalized approaches for different ageotypes, we may not only improve the treatment of chronic heart and lung diseases but also potentially delay or prevent their onset, thereby having a profound impact on public health. Dr. Yulu Chen is a bioinformatician specializing in using multi-omic approaches to study complex disease phenotypes, particularly respiratory diseases and aging. Her overarching career goal is to become an independent researcher with the skills to enhance the biological understanding and clinical management of aging and related heart and lung diseases through integrative omics techniques. Dr. Chen's strong quantitative and methodological background, combined with the support from a diverse and world-class mentoring and advisory team from the Channing Division of Network Medicine at Brigham and Women’s Hospital and Harvard Medical School, has well-prepared her for the proposed research. The proposed career development plan builds on her previous training and outlines five training objectives to enhance her trajectory toward becoming an independent researcher: 1) expand her skills in integrative omics techniques, particularly network-based approaches for integrating multi-omic datasets; 2) enhance her knowledge and application of epigenomic data; 3) gain a deeper understanding of the phenotypes and experimental measurements of heart and lung diseases; 4) broaden her understanding of the biological mechanisms of aging; 5) improve her skills in research design, mentorship, scientific ethics, and research communication.

NIA - National Institute on Aging

Abstract Aging is a natural process resulting in the decline of tissue functionality. In the skin, aging results from degenerative changes in dermal and epidermal compartments. However, the molecular mechanisms behind skin aging are not fully understood. One of the poorly explored characteristics of the aged skin is the reduced number of anchoring fibrils, which connect the dermis to the epidermis, and the reduced level of their main protein component, type VII collagen (COL7A1), which is critical for the stability of the extracellular matrix. Interestingly, the skin of patients with recessive dystrophic epidermolysis bullosa (RDEB), a severe skin fragility condition caused by biallelic mutations in the gene COL7A1, shows clinical similarities with the skin of the elderly. This suggests that COL7A1 deficiency promotes premature aging in RDEB patients. However, it is not clear whether the pro-inflammatory background of chronic non-healing wounds is the only cause of premature skin aging in RDEB patients or functional COL7A1 plays an important, currently unknown role in protecting the skin from aging. My preliminary proteomic analysis of primary patient derived fibroblasts (COL7A1-/- PPF(s)), as well as patient specific organoid derived fibroblasts (iF(s)) generated from primary fibroblasts transitioned through induced pluripotency that were either uncorrected (COL7A1-/-) or genetically corrected using CRISPR/Cas9 (COL7A1+/-), revealed COL7A1 dependent perturbations in insulin-like binding protein 2 (IGFBP2) with COL7A1- /- PPF(s) and COL7A1-/- iF(s) showing marked deficiencies in its expression relative to genetically corrected iF(s). Therefore, I hypothesize that COL7A1 has a moonlighting role regulating fibroblast secretory profiles, and, in its deficiency, leads to accelerated cellular senescence and aberrant perturbations in fibroblast functionality, specifically through the IGF axis. Inhibition of IGFBP2 has been shown to lead to an overexpression of p21, p16, and p19, all powerful promotors of senescence and a potential mechanism by which premature acceleration senescence is induced in COL7A1 deficiency. During my fellowship, I will dissect the role of COL7A1 in senescence of skin cells. In aim 1, I will further analyze COL7A1 dependent perturbations in the proteome and secretome of COL7A1 deficient and COL7A1 corrected lines that are generated from induced pluripotent stem cells. Reprogramming into iPSCs erases aging associated marks in cells that arose from pro-inflammatory and fibrotic background and allows us to generate and characterize COL7A1 deficient and corrected cell lines without the influence of external factors that can mask the role of COL7A1 in skin aging. In aim 2, I will assess other age-associated marks in COL7A1 deficient and corrected cell lines to validate the connection between IGF signaling and COL7A1 functionality. If successful, this proposal will result in our better understanding of the role of COL7A1 in protecting the skin from aging and in developing novel therapies for RDEB and anti-aging interventions.

NHLBI - National Heart Lung and Blood Institute

Project Summary/Abstract Cigarette smoking is the main cause of occurrence of Chronic Obstructive Pulmonary disease (COPD). Studies show that cigarette smoke (CS)-mediated increase in cellular senescence causes premature ageing in the lung, thereby leading to the development of COPD. However, the exact mechanism of disease development via alterations in cellular senescence over a prolonged period in CS-induced COPD is not known. In this respect, various inter-individual variations are observed amongst regular smokers, thus indicating potential involvement of epigenetic regulations in modulating the downstream signaling. Considering this I hypothesize that the epigenetic regulation of cellular phenotypes and gene expression of cellular senescence- associated genes (p16, p21 and p53) drives disease-fate towards emphysema upon chronic CS exposure. To test this hypothesis, I propose to: (1) determine the time (1, 3, and 6 months(mo)) and age-dependent (2mo versus 18 mo) effects of CS-exposure on the lung cellular senescence and associated epigenetic regulation in vivo using SNARE-sequencing (K99 phase), and (2) study the determinants of cellular senescence and associated epigenetic mediators in primary human small airway epithelial cells (SAECs) from healthy and diseased (COPD) individuals (R00 phase). My postdoctoral work has shown that clearance of senescent cells results in reduction of CS-induced neutrophilic inflammation and reversal of alveolar wall damage in mouse model (p16-3MR). Thus, studying the CS-induced changes in the lung neutrophil and epithelial cell population is the focus of this project. Upon completion of these aims, I will be able to identify gene targets and associated chromatin sites that are altered on cigarette smoking and associate them with disease development in COPD. Use of two mouse models (C57BL/6J and p16-3MR) will help in understanding the role of p16-dependent senescence in CS-mediated alterations in the lung. Additionally, use of Multiome (SNARE-seq) technology in Aim1 will provide a chance to identify unique cell phenotypes based on differentially expressed genes and correlate them to the epigenetic signatures within the cell upon CS exposure. These results would lead to future publications, collaborations, and grant proposals, thus helping to launch my career as an independent researcher in the field of lung biology.

NSF

In the Alabama-Florida (AL-FL) Gulf Coast region, ecological disturbances such as hurricanes, wildfires, and harmful algal blooms can interact and have direct impacts on ecosystem health and services. Further, this region is the primary biodiversity hotspot of the continental United States, hosting the greatest total numbers of species and endemic species of mammals, birds, amphibians, reptiles, freshwater fish, and trees. Human impacts on landscapes and the climate system are also increasing ecological vulnerability to these disturbances, which can degrade ecosystem health, wildlife habitat, and water quality. This project focuses on the impacts of multiple interacting disturbances on ecosystems, as well as how disturbances interact across the boundaries between terrestrial and aquatic ecosystems, which have not been studied in detail before. Further, this project will provide new science to guide conservation of the biodiversity and endemic species of this region. In addition to funding new science, this project will also support the training of multiple undergraduate and graduate students, and broader stakeholder education through Auburn University’s Extension System. By using paleoenvironmental approaches, the project will resolve long-term environmental histories of disturbances (hurricanes, wildfires, and harmful algal blooms) and ecosystem diversity and state to resolve the complex relationships between disturbances and ecosystems of the AL-FL Gulf Coast. Project objectives are to: (1) Reconstruct hurricane and wildfire occurrence to identify relationships between these terrestrial disturbances; (2) Reconstruct harmful algal blooms (HABs) to characterize relationships between aquatic and terrestrial disturbances; and (3) Reconstruct terrestrial and aquatic community changes to characterize how the interactions of episodic disturbances (hurricanes, wildfire, HABs) impact community state. This multi-disturbance reconstruction approach is unique to the project and will enable novel insights into the long-term disturbance interactions and ecosystem impacts. The project will enable undergraduate and graduate students to obtain important research experience, and will facilitate public outreach and education. This project is jointly funded by the NSF Ecosystem Science Cluster and the Established Program to Stimulate Competitive Research (EPSCoR). This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIMH - National Institute of Mental Health

Project Summary/Abstract Up to seventy percent of autistic individuals report challenging auditory sensory experiences despite having normal hearing thresholds. These experiences include both increased and decreased sensitivities to sounds, as well as difficulties attending to and ignoring sounds. Furthermore, these experiences are exacerbated by stress and fatigue, indicating the role of arousal-related systems in mediating these sensitivities. Despite the prevalence of these experiences, prior research has failed to identify consistent differences in neural activity in brain regions responsible for auditory sensitivity, attention, and arousal, thus hindering the development of effective treatments and sensitive measures. Importantly, these brain regions compose a circuit that adjusts auditory sensitivity in response to the environment, but no research has investigated whether the dynamics of this circuit are different in autism. We will address this gap by measuring the integrity of the neural circuit through which arousal modulates auditory perception and vice versa. We hypothesize that auditory difficulties in autism emerge from different dynamics of the neural circuit that maintains auditory sensitivity, rather than unvarying increased or decreased activity in any of these regions in isolation. We test this hypothesis with a novel experimental approach. Participants will hear sounds of varying intensity as we co-register electroencephalography (EEG) and pupil diameter (PD). Simultaneous measurement of EEG and PD allows us to determine the extent to which individual brain responses reflect the influence of exogenous factors, such as sound intensity, relative to endogenous factors such as arousal. We quantify the strength of these associations using novel regression models developed for the analyses of time-varying relationships at the level of single trials. These neurophysiological measures will be integrated with clinical assessments of auditory sensitivity to understand relationships among circuit dynamics and auditory experience. This research will generate important knowledge elucidating the mechanisms of auditory sensitivity in autism. This knowledge will facilitate progress in the development of sensory-specific biomarkers and interventions, which the field currently lacks. Finally, unlike prior research that has focused on mean differences between groups, we aim to identify an explanatory mechanism for variability within individuals.

NIGMS - National Institute of General Medical Sciences

ABSTRACT Symbiotic relationships between animals (including humans) and bacteria are ubiquitous in the natural world, and cultivating a healthy microbial community, or microbiota, is critical for the health of hosts. Our lab explores the fundamental principles of host-microbe symbiosis to learn how these relationships are formed and maintained. We focus on flagella, tail-like structures that are commonly used by bacteria to navigate their environments. Many symbiotic bacteria use flagellar motility to colonize their hosts, and the proteins that govern flagellar development and activity are critical for beneficial and pathogenic host-microbe associations. Many species from the Vibrio genus of bacteria form symbiotic relationships with animal hosts, ranging from invertebrate mutualists like Vibrio fischeri to human pathogens like Vibrio cholerae. In symbiotic Vibrios, flagellar motility is critical for the establishment of the host-microbe relationship, and mutations in the flagellar apparatus reduce both their general motility and their ability to colonize their hosts. Although flagellar motility is well-described in some bacteria in vitro, the genetic requirements for flagellar function and specific types of motility are often restricted to artificial conditions that do not reflect real world conditions. Further, we and others have found that in vitro characterizations of flagellar motility in simple environments do not generalize to the motility behaviors that bacteria use inside the bodies of their host organisms. Over the next five years, we will use the model symbiosis between the Hawaiian bobtail squid and Vibrio fischeri to characterize the role of flagellin proteins, which make up the flagellar filament, in symbiotic motility. We will determine the role of individual flagellins and flagellin diversity in motility, then will measure how flagellins shape motility in complex, host-like environments. Finally, we will assay the role flagellins play in symbiotic colonization. This work will uncover new information about the structure and function of bacterial flagella, and will further characterize how bacteria use motility to colonize and impact their hosts.

NSF

Galaxies continuously form stars over billions of years. To keep forming stars during these exceedingly long timescales, galaxies need a fresh supply of gas flowing right into their centers. Gas is the primordial building material of stars. This program will study how gas flows from outside galaxies into their inner regions, sustaining the process of star formation for billions of years. To achieve the main objectives of this program, the investigators will analyze data using several astronomical telescopes, among them the NSF-funded Very Large Array and Atacama Large Millimeter Array. This program will support the research work of a graduate student. The PI will continue her work with the Prison Education Program (PEP) at Arizona State University and support Maricopa County prison educators in developing a curriculum for juvenile and adult education. Graduate student volunteers of the PEP Astro program will travel to the prisons to deliver lectures and conduct hands-on activities. The primary objective of this program is to characterize gas flows responsible for sustaining star-forming disks in the present epoch. The investigators of this program will identify gas flows into, through, and out of galaxy disks directly and infer gas accretion in the recent past via metallicity offsets of HII regions within the disk. This program will obtain high-resolution mapping of the cold neutral gas in the HI 21cm transition to trace the content and kinematics of the interstellar medium in 14 nearby (<100 Mpc) Milky Way mass spiral galaxies. These data will be combined with optical spectroscopic measurements of HII regions probing metallicity and dust content. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIDA - National Institute on Drug Abuse

Project Summary Millions of individuals are affected with substance use disorders (SUD), posing a significant burden on these individuals, their families, and communities. There is a substantial comorbidity between SUD and HIV infection. HIV is also a risk factor for SUD because of the increased use of opioid pain medications that may lead to opioid addiction. The Single-Cell Opioid Responses in the Context of HIV (SCORCH) consortium was formed to gain insights into cellular and molecular responses in different brain regions to SUD and HIV by collecting single-cell transcriptomic and epigenomic data in affected brain regions from hundreds of human donors, as well as from animal models. It has been observed that SUD and HIV comorbidity may exacerbate cellular dysfunction beyond the effects of each condition alone, and the data generated by the SCORCH consortium provide opportunities for a comprehensive characterization of cellular states across conditions including control, HIV, OUD, and HIV+OUD. Preliminary data show substantial heterogeneity in molecular phenotypes across samples with the same exposure, e.g., HIV and SUD. Our premise is that the identifications of genetic variants mediating the effects of exposures to HIV and SUD will offer a unique angle to understand how different cell types in different brain regions respond to the exposures, and such understanding through genetic heterogeneity among individuals can lead to novel insights and clinical applications. We will apply state-of-the-art integrative methods to investigate how genetic variants affect molecular phenotypes in different cell types across brain regions with different exposure. We will accomplish this goal through three specific aims. The first aim will analyze total read counts from a transcript/isoform or peak using Bayesian methods that explicitly model shared genetic effects to borrow information across cell types and brain regions to increase statistical power. We will perform eQTL, caQTL, and isoQTL analyses. We will also leverage the multi-omic data to infer gene regulation networks and conduct grQTL analysis. The second aim will consider allele-specific analysis to complement analyses based on total counts. We will then combine allele-specific results with total read count results. To further improve statistical power, we will integrate SCORCH data with external data sets, computationally predicted effect sizes for genetic variants, and transfer known QTLs. We will develop gene expression and chromatin accessibility imputation models to facilitate genome-wide association studies. We will work with the SCORCH team to share our results with the broader scientific community. This project will be co-led by Dr. Hongyu Zhao, Dr. Mark Gerstein, and Dr. Ke Xu, who have complementary expertise covering statistical genetics/genomics, computational biology, single-cell analysis, SUD genetics, and HIV research.

NIH

Background: Veterans who lack capacity to make their own decisions and do not have a surrogate decision- maker cannot consent to discharge from VA inpatient settings. In these cases, a professional guardian must be court appointed before the Veteran can be discharged from inpatient care to other settings. The guardianship process is complex and time- and resource-intensive. As a result, medically stable, unrepresented Veterans may spend unnecessary months, or even years, in inappropriate inpatient settings awaiting guardianship. These needless delays in discharge: (1) result in inappropriate use of inpatient beds and increased risks of hospital-acquired conditions, (2) conflict with a patient’s right to care in the least restrictive setting and (3) cost VA upwards of $200 Million dollars per year, a figure that is expected to rise as the VA population ages. However, VA does not systematically document or track guardianship processes or outcomes and has no national, standardized approach to guardianship. Research is needed to gauge the scope of the problem, understand the VA guardianship process, and identify ways to improve it. Significance to VA: The US Senate Special Committee on Aging recently called for increased scrutiny of guardianship laws and outcomes, citing the lack of data nationally. According to the VA Social Work Guardianship National Committee, VA lacks national policies and processes for incapacitated Veterans who require transition from inpatient care settings to lower levels of care. The recently updated VA Directive 1004.01 highlights VA’s commitment to provide care for Veterans in the least restrictive setting possible while supporting their right to autonomous, informed participation in their health care decisions. VA must invest in infrastructure to track guardianship in Veterans and support research to enable greater understanding and meaningful improvement of guardianship processes. Innovation and Impact: The proposed study is innovative because it generates new knowledge and creates a foundation for future research on guardianship in VA. Specifically, this work focuses on a unique and underserved population of Veterans who are particularly vulnerable and often excluded from research studies. This mixed methods study is the first of its kind in VA. Our ongoing partnership with the Office of Care Management and Social Work ensures that this work will be used to inform VA policy and practice. Specific Aims: (1) Create a test dataset made up of a national sample of Veterans who have undergone the guardianship process in VA inpatient settings that includes their sociodemographic and clinical factors. (2) Understand key shareholder perspectives of current VA guardianship processes, including workflows, challenges, best practices, and opportunities for improvement. Methodology: (Aim 1): Using retrospective observational methods and an extension of methods developed by our team, we will do searches of VA electronic health record notes and administrative data, followed by manual chart reviews, to create a nationally representative test dataset of Veterans who have guardians. For each case, we will also collect information about their guardianship, health care and demographics. (Aim 2): We will conduct semi-structured interviews with VA staff and leadership who are involved in guardianship policy or practice to understand current guardianship processes, challenges, and ways to improve them at the local VAMC and national levels. We will also conduct interviews with Veterans who have guardians and their guardians to understand their experiences of the process and opportunities for improvement. Path to Translation/ Implementation: We will use the test dataset created in Aim 1 in a future IIR to validate a natural language processing approach to systematically identify Veterans who have guardians. This will facilitate research examining guardianship in VA on a national scale. Findings from Aim 2 will inform future research to develop an intervention improve the guardianship process for future study and dissemination.