University Of Chicago

NIH Research Projects · FY 2024 · 2020-09

Abstract This proposal is focused on the challenge of understanding and thus improving treatment for idiopathic pulmonary fibrosis, a devastating interstitial lung disease without effective treatments. Deregulation of lipid signaling and metabolism has long been studied in pulmonary fibrosis, pointing to changes in sphingolipid and ceramide pathways that may impact pathology. In recent years, increased attention has been paid to a possible role for senescent cells in the lung as a critical factor driving pulmonary fibrosis. Senescent cells express inflammatory factors, including signaling lipids, which may drive the fibrotic process. Current questions include why senescent cells accumulate in these patients and how their pro-inflammatory activity might be mitigated. An inference is that preventing formation of senescent cells, blocking their lipid signaling and/or promoting their clearance from the lung might prevent pulmonary fibrosis or block disease progression. Importantly, there may be a direct link between sphingolipid pathways and cellular senescence. Ceramides have been shown to induce senescence in otherwise proliferating cells. Our studies have implicated lipid peroxidation and its aldehyde end-products such as 4-hydroxynonenal as key mediators of accelerated senescence. Transcriptomic, proteomic and lipidomic analysis of proliferative and senescent lung cells will be used to identify key senescence factors and networks that may point to the specific lipid metabolic pathways that drive senescence and inflammatory signaling. We will then examine lipids and modulators for the ability to promote or prevent senescence. Finally, we will examine whether manipulation of lipid metabolic pathways can be used to potentiate clearance of senescent cells and thereby limit pulmonary fibrosis. With success in these studies, we anticipate identification of candidate therapeutics with potential to move to clinical trials.

NIH Research Projects · FY 2024 · 2020-09

PROJECT SUMMARY Annual vaccination remains the primary public health strategy to mitigate the burden of influenza infection, and there is evidence that repeated influenza vaccination can affect the efficacy of the vaccine. This evidence arises not only from multiple observational studies of vaccine effectiveness but also studies of immunogenicity, including small trials. Understanding what causes influenza vaccines to be more or less effective in different people and populations is critical to the rational deployment of existing vaccines and the development of universal vaccines. But the causes of altered effectiveness and immunogenicity in repeat vaccinees are intrinsically difficult to study in populations in which vaccination is universally recommended, because repeat vaccinees differ from other vaccinees and non-vaccinees in important ways. These differences leave open the possibility of residual confounding in infection and vaccination history, and thus make it difficult to identify the effects of vaccination itself. We propose a randomized, clinical trial to investigate the effects of repeat vaccination and their underlying immunological causes in an adult population with low vaccination coverage and no recommendation for influenza vaccination. Approximately 820 adults in Hong Kong will be randomized into five groups, with one group vaccinated the first year, and other groups receiving placebo (saline) injections; each year, another group will start receiving the influenza vaccine, and will be vaccinated annually until the study ends after four years. This design will allow comparison of vaccine responses and failures (infections) in the placebo, newly vaccinated, and repeatedly vaccinated participants. Additionally, it will provide longitudinal samples of immune status and influenza-specific responses over time, from which we will develop predictive models of the response to vaccination and infection, including repeat vaccination. The proposed high-dimensional immunological profiling, coupled with statistical approaches that can accommodate the complexity of the key hypotheses, should maximize insight into the effects of repeated vaccination on seasonal influenza. The models will formalize, evaluate, and extend current theory, and thus provide a quantitative basis for anticipating vaccine non- responsiveness and improving vaccination strategies. Banked specimens will enable new hypotheses to be tested in the future.

NIH Research Projects · FY 2024 · 2020-09

PROJECT ABSTRACT: Medical decisions for many chronic diseases such as diabetes are becoming more complex as the number of available therapies expand and as we learn how risk factor targets and treatments may need to be individualized based on a patient's clinical characteristics, genetic profiles, treatment preferences, and social circumstances. To strengthen training and research in medical decision making in the study of chronic diseases of older adults, Dr. Elbert Huang, Professor of Medicine at the University of Chicago, is submitting this K24 renewal proposal. The University of Chicago is an ideal environment for this program because of its well-established clinical research programs, history of interdisciplinary collaboration, and significant strengths in the social sciences. The overall goal of the program is to support young investigators who are studying medical decision making in older adults living with individual or multiple chronic diseases. Trainees of the program will have access to cohort studies of older adults (Kaiser Permanente Northern California Diabetes Registry, the National Social Life Health and Aging Project, national Medicaid and Medicare Claims), support for the construction and evaluation of simulation models of chronic diseases, guidance for developing decision support interventions, and facilitated access to the UChicago Practice-Based Research Network. In his own research in geriatric diabetes, Dr. Huang will pursue the following aims: 1) advance the study of variation in diabetes onset, complications and treatment response in observational studies; 2) advance the development of simulation models of diabetes complications; and 3) advance the development of diabetes decision support for personalizing care for older adults. In two ongoing observational studies (R01 AG060756, R01 AG063391), Dr. Huang will explore population-level variation in the onset of diabetes and its complications as well as variation in response to achieved glucose targets and treatments. As part of Aim 2, Dr. Huang is part of an investigative team led by his mentee, Dr. Neda Laiteerapong (R01 MD013420) developing a new simulation model of diabetes complications using multi-ethnic data from the Kaiser Registry. This model will be comprised of a series of individual outcome prediction models (cardiovascular events, hypoglycemia, dementia, mortality) that interwoven together. Under Aim 3, Dr. Huang will first complete evaluation of a trial of a decision support tool (My Diabetes GOAL) created during his initial K24. The tool, embedded within the electronic medical record, is designed to engage patients in personalized goal setting and disease management with the support of a nurse. He will also develop an updated stakeholder-informed diabetes decision support intervention for personalized care for older adults, addressing both goal setting and treatment selection. This K24 award will allow Dr. Huang to strengthen his mentorship program in medical decision making in chronic diseases and expand his research portfolio in new directions that will guide individualization of care for diverse populations of patients living with diabetes.

NIH Research Projects · FY 2024 · 2020-09

Project Summary/Abstract The long-term goal of this work is to reduce the incident of stroke by identifying the most vulnerable patients using MRI scans. Currently roughly 1 of every 8 patient who have had an initial stroke from intracranial atherosclerosis disease (ICAD) will suffer a second stroke within a year. Patients who are likely to fail medical management have loss of cerebrovascular reserve, poor collateral arterial blood supply, and/or plaque that is vulnerable to rupture from active macrophage infiltration. Our goal is to identify vulnerable patients to inform the selection for new medical management protocols, stenting or stent-less angioplasty. We will develop a suite of new MRI scans and evaluate them in the intended patient population, comparing to reference standard CO2 Challenge CVR, HMPAO SPECT or direct imaging of active macrophages. Significance: ICAD is one of the most common causes of stroke worldwide and carries an extremely a high risk of recurrent stroke. ICAD patients with severe stenosis (70 to 99%) are at particularly high risk for recurrent stroke in the vascular territory of the stenosis (~12 to 20% within 12 months) despite treatment with aspirin, Plavix and management of risk factors (hypertension, smoking etc). The use of new, preventative treatment including angioplasty, new anti-platelet medication would benefit if the most vulnerable patient can be identified. Our imaging biomarkers will improve risk stratification for the of stroke in a vulnerable, high risk population. Innovation: We have developed time resolve MRI scans that are targeted to risk factor of stroke in ICAD: (1) Cardiac Gated “Snapshot” images of transient changes in the cerebral vasculature in response to arterial pressure changes induced by the cardiac cycle. These changes are muted by a loss of cerebrovascular reserve a risk fact of stroke. (2) A new mathematical deconvolution algorithm based on linear time-invariant system theory to quantify perfusion supplied to a vascular bed through collateral arterial blood supply distal to a stenosis. (3) First ever high-resolution permeability of the intracranial arterial walls to identify macrophage infiltration. Scientific Rigor: The geometry of the human head and topology of the vasculature are unique, and we therefore perform all our studies in the intended patient population: humans with ICAD. To ensure scientific rigor, we will compare directly to reference standard values of CO2 cerebrovascular reserve, collateral arterial supply, and macrophage infiltration in plaques. Probability of Success: We have built a strong, multi-disciplinary team with a long track record of successful, collaborative neurovascular research. We believe this high probability of successful completion of the aims and high likelihood of clinical translation.

NIH Research Projects · FY 2024 · 2020-08

PROJECT SUMMARY/ABSTRACT This proposal outlines an integrated research and career development plan for Nathan Schoettler, MD, PhD to conduct training in the laboratory of Dr. Carole Ober and transition to an independent academic position by establishing a research program at the interface of genetics and immunology with a NIH mentored career award (K08). The PI recently completed an NIH T32 fellowship (T32 HL007605) and is trained in the fields of immunology, genetics, molecular biology and cell biology, and during the time of this K08 award, the PI will receive additional academic guidance from additional co-mentors (Dr. Dan Nicolae and Dr. Anne Sperling) and advisors (Dr. Julian Solway, and Dr. Hae Kyung Im) at the University of Chicago. The career development plan is designed to equip the PI with the necessary knowledge and skills in statistical genetics and cellular immunology for a successful transition to an independent academician, and R01 funding. The overall goal of the proposed research is to elucidate the role of genetic variation in regulating the expression of genes in specific lung cell types that play a critical role in asthma. Asthma is a complex genetic disease with high heritability, yet how genetic variation influences pathobiological mechanisms, or endotypes, in asthma has not been resolved. Studies conducted by the PI during his post-doctoral T32 phase demonstrated that childhood- onset and adult-onset asthma have both shared and distinct genetic risk loci (Lancet Resp Med 2019). The PI furthermore showed that human lung tissue resident memory T cells, a subset of T cells that do not circulate in the blood, are programmed differently and derived from a separate pool of progenitor than lung-draining lymph node memory T cell subsets (Comm Biol, in press), and they rapidly increase the expression of key asthma cytokines when activated. This K08 research proposal tests the overall hypothesis that specific asthma sub- phenotypes have shared and distinct genetic risk factors and that these risk loci mediate effects in cell-specific manners that perturb gene expression and disease risk in unique ways. Aim 1 will test the hypothesis that clinically important asthma sub-phenotypes share a set of genetic risk variants but also have additional, sub- phenotype-specific genetic risk loci. Aim 2 will test the hypothesis that a subset of asthma-risk loci will harbor variation that has unique effects on gene expression in lung tissue resident memory T cells that have not been revealed in other tissues or cell types. Aim 3 will test the hypothesis that asthma sub-phenotypes have different sets of risk loci that influence gene expression in asthma-relevant cells from lungs, specifically T cells, smooth muscle cells and epithelial cells. The goals of this research will be achieved by integrating genome-wide association studies with expression quantitative trait loci identified in airway cells with and without asthma- relevant exposures, and lead to a mechanistic understanding of how genetic risk variants influence cellular responses, ultimately revealing potential therapeutic targets. This career award will accelerate the transition for Dr. Schoettler to an independent physician-scientist and the acquisition of competitive R01 funding.

NIH Research Projects · FY 2024 · 2020-08

PROJECT SUMMARY Almost 800,000 critically ill patients require mechanical ventilation every year and three quarters of the survivors suffer from persistent disability, which poses a major public health problem as critical care becomes more widely utilized and available. Although early mobilization, which engages patients in physical activity during mechanical ventilation, is a promising evidence-based intervention that may prevent disability, less than ten percent of pa- tients ever get out of bed. This proposal aims to apply precision medicine to identify patients who are most likely to benefit from early mobilization and elucidate how it can be implemented successfully to extend the benefits of early mobilization to critical care survivors at greatest risk for long-term disability. I hypothesize that this re- source-intensive intervention can be applied with greater precision to a subset of patients most likely to bene- fit, and that implementation science strategies can be devised to successfully drive adoption of this interven- tion beyond a clinical trial setting. I will test my hypothesis in three aims: Aim 1) I will identify the optimal critical illness phenotype for implementation of early mobilization by using cutting-edge machine learning methods; Aim 2) I will determine the effect of early mobilization on long-term functional disability to incentivize adoption of this practice; Aim 3) I will determine the barriers and facilitators of implementation of early mobilization across five institutions to identify the contextual features associated with successful implementation to inform strategies that can bridge the gap between evidence base and clinical practice. My long-term goal is to mitigate the com- plications of critical illness with clinical trials using precision-based methods to identify at-risk and yet apt-to- benefit populations paired with implementation science methodologies to illuminate how to bring these interven- tions to the bedside. To accomplish this, I have assembled an exceptional interdisciplinary team of mentors (Drs. Vineet Arora, Matthew Churpek, and John Kress) and advisors (Drs. Shyam Prabhakaran, Donald Hedeker, Laura Damschroder, and Matthias Eikermann) who have a track record of NIH-funding and successful mentor- ship of post-doctoral candidates. I intend to build on my foundation as an accomplished clinical trialist and have formulated an in-depth career development plan to gain expertise in machine learning methods to identify differ- ential treatment effects (Churpek and Prabhakaran), longitudinal data analysis, (Arora and Hedeker), and imple- mentation science methods (Arora, Prabhakaran, and Damschroder) to craft strategies that bring complex mul- tidisciplinary interventions from clinical trials (Kress and Eikermann) to everyday ICU care. Completion of this proposal will train me to fill an unmet need defined by a recent National Academy of Medicine publication which indicated that identification of differential treatment effects must be paired with rigorous implementation to help transition evidence base to routine clinical care. Equipped with advanced statistical skills and implementation science approaches, I will be able to design hybrid effectiveness-implementation trials to target and implement complex multidisciplinary interventions to vulnerable populations in future R01 level applications.

NIH Research Projects · FY 2026 · 2020-08

PROJECT SUMMARY/ABSTRACT The stability of cellular identities is essential for coordination and maintenance of specialized organ systems that provide organismal homeostasis. However, alterations in somatic cell fates can be necessary to repair damaged tissue or adapt to changing microenvironments. How phenotypic variation is constrained or promoted to regulate the balance between cellular stability and plasticity is largely unknown. Our laboratory recently identified ‘chromatin accessibility noise’, the fluctuations in background nucleosome dynamics, as a critical determinant of cellular plasticity. Using medullary thymic epithelial cells (mTECs) – which facilitates the selection of a self- discriminating repertoire of T cells for adaptive immunity – as a physiological model system, we found chromatin accessibility noise is amplified during mTEC maturation via repression of the tumor suppressor p53. Augmenting p53 activity stabilized chromatin barriers, inhibited ectopic transcription and limited phenotypic plasticity, ultimately leading to the escape of harmful self-reactive T cells from negative selection. The objective of this proposal is to bridge the key gaps in understanding how p53 is repressed during mTEC maturation and the modes by which p53 senses and suppresses chromatin accessibility noise. Using multi-omic tools to jointly profile the chromatin accessibility landscape and the genes that are expressed in each individual cell, we propose to address these gaps by testing the influence of p53 negative regulators induced during mTEC maturation and the target genes that are activated by p53 hyperactivity on chromatin accessibility noise and cellular plasticity. We will complement these profiles with localization studies that quantify where p53 binds across the genome in mature mTECs to delineate the direct and indirect mechanisms of p53-mediated suppression of chromatin noise. Achieving these objectives will provide foundational understanding of a molecular lever that regulates cell fate integrity versus plasticity with broad implications for mechanisms underpinning tumorigenesis, cellular reprogramming, tissue homeostasis and immunological tolerance.

NIH Research Projects · FY 2025 · 2020-07

This proposal seeks to renew funding for the University of Chicago (UC) Center for Healthy Aging Behaviors and Longitudinal InvestigationS (CHABLIS). CHABLIS’ purpose remains to promote a sustained research and infrastructure development program that leverages longitudinal data from observational and interventional studies to examine how demographic and economic factors facilitate or suppress individual healthy aging behaviors to influence outcomes for older adults over the life course. Spanning five divisions and schools within UC and a growing national network of individual and institutional affiliates, CHABLIS will bring many areas of expertise and methodologies to bear on the demography and economics of aging. CHABLIS’ central hypothesis is that by connecting people across disciplines (e.g., economics, sociology, medicine), academic units, and institutions with a focus on longitudinal studies of healthy aging behaviors, we can: 1) enhance innovation in the demography and economics of aging, and 2) cultivate the next generation of leaders in social science approaches to aging research. Informed by the NIA Health Disparities Research Framework, we pursue these aims recognizing the disproportionate distribution of health within and across populations. We include multiple health disparity populations and perspectives, including several projects on the large, and often problematic, role that the health care system plays in the aging experience and how that role may vary with socioeconomic factors. Our Program Development Core continues to focus on supporting research and career development for emerging scholars and pilot projects that span the demography and economics of aging in both medical and social science realms, but now enhances support for investigators from institutions with historically low levels of NIH funding. Our External Network Core builds on UC’s tradition of integrating social science insights into the professions by integrating research in the economics and demography of aging and clinical research. We propose both to extend our current major research areas (including our Comprehensive Care Program, pharmacogenetics, and oral health clinical studies, and connections to major NIA-funded population-based studies of aging) and to develop a complementary new research area on learning health system approaches that use real world data from clinical settings. Our External Research Resources Support Core builds on our highly successful Collaborative for Innovation on Data and Measurement (CIDMA) that seeks to identify, design, conduct, and assess innovations in data collection and measurement for use in data-focused studies of aging, and to share and continually advance those findings. With the movement of CIDMA leaders Hotz from Duke to UC and Cagney from UC to the University of Michigan’s Institute for Social Research (ISR), we transition CIDMA from its original partnership with Duke to a collaboration with Cagney and colleagues from ISR. Our Administrative and Research Support Core and Communication and Dissemination Core work across the center to ensure CHABLIS operates, communicates and disseminates its work for maximum impact.

NIH Research Projects · FY 2023 · 2020-07

In this research project, we propose to study the molecular mechanisms by which RE1 silencing transcription factor (REST) regulates cardiomyocyte cell cycle using synergistic approaches of mouse genetics, molecular and systems biology. Our ultimate goal is to identify potential therapeutic targets to combat heart disease by promoting cardiomyocyte proliferation to repair the injured adult heart. The adult heart has a limited repairing capacity, because most adult cardiomyocytes are non-dividing cells. For adult heart regeneration by pre- existing cardiomyocytes, we need to understand how cardiomyocyte proliferation is controlled, what population of cardiomyocytes maintains proliferation potential, whether developmental mechanisms of cardiomyocyte proliferation are re-activated under diseased conditions, and if they can be further enforced. Here we plan to address these questions by investigating REST functions in the mouse heart, because our recent studies have identified REST as a new intrinsic regulator of cardiomyocyte proliferation required for embryonic heart development and neonatal heart regeneration. REST represses the cell cycle inhibitor p21 to maintain cardiomyocyte cell cycling. REST is also required for the expression of several key cell cycle activators. Based on these findings, we hypothesize that upregulation of REST may improve the renewal of the injured heart by modulating the expression of cell cycle genes and regulators to promote cardiomyocyte proliferation. We will test this hypothesis in two aims. Aim 1 will determine whether forced REST expression in failing hearts promotes cardiomyocyte cell cycle and improves the disease outcome. We will specifically inactivate REST in the cardiomyocytes, with or without the p21 inactivation, as well as forced REST expression, to determine whether REST is required for adult heart repair. Aim 2 will investigate how REST controls cardiomyocyte cell cycle in normal and failing hearts by studying and comparing the REST regulatory network in the mature (MYH6+) and immature (MYH7+) cardiomyocytes. We will also study the REST regulatory network by determining the REST recruitment of different chromatin co-factors for gene activation or repression. This will be achieved by using the cell-type specific, genome wide REST chromatin immunoprecipitation sequencing (ChIP-seq) and transcriptomics (RNA-seq) for MYH6+ versus MYH7+ cardiomyocytes from normal or injured hearts. The human relevance of mouse findings will be ascertained by the functional study of key REST- regulated genes in the proliferation of cardiomyocytes derived from the human inducible pluripotent stem cells (iPSC). By completing this project, we will have determined the REST role in promoting cardiomyocyte proliferation to improve the renewal and function of failing adult hearts. We will have also learned a genetic regulatory network by which REST regulates adult cardiomyocyte cell cycle and identified new targets for improving cardiomyocyte proliferation for heart regeneration. The new information will advance our understanding of cardiac biology and provide a new direction to treat heart failure.

NIH Research Projects · FY 2024 · 2020-07

PROJECT SUMMARY Germline cancer genetic testing has become a standard evidence-based practice, with risk reduction and cancer screening guidelines for genetic carriers. Yet, many at-risk patients do not have access to genetic services, leaving many genetic carriers unidentified. Given increasing precision medicine applications, and a limited and geographically restricted workforce of genetic providers, innovative delivery models for genetic services that are responsive to the needs of geographically and sociodemographically diverse patient populations in their local health care systems are needed. Suboptimal access to genetic services is an acute problem for childhood cancer survivors, who have high rates of subsequent malignant neoplasms (SMN). Studies indicate that >10% of survivors carry a pathogenic or likely pathogenic germline mutation in cancer susceptibility genes (eg. TP53, BRCA ½). In order to identify high-risk survivors for early surveillance and intervention, NCCN and Children's Oncology Group Guidelines recommend that survivors with a personal and/or family history of cancer be referred for genetic services. However, <15% of these survivors have access to genetic counseling services and both survivors and their Primary Care Providers (PCPs) are largely unaware of their health risks and thus, adherence to high-risk surveillance guidelines is low. Our studies in adult patients with a personal or family history of cancer suggest that remote telegenetic services (by phone or videoconferencing) may increase access to genetic services and identification of genetic mutation carriers. Yet, our current studies have examined these models partnering on-site with community oncology practices, limiting scalability. This is of particular importance for childhood cancer survivors, who are widely distributed nationally and >85% are receiving their care with PCPs. Thus, our premise is that our adapted in-home, collaborative PCP model of remote telegenetic services can provide a scalable model that will result in increased uptake of evidence-based recommendations for cancer genetic services in survivors. We propose a 3-arm randomized Hybrid 1 effectiveness and implementation study in the Childhood Cancer Survivor Study (CCSS) to evaluate the effectiveness of our in-home, collaborative PCP model of remote telegenetic services to increase uptake of cancer genetic testing in childhood cancer survivors compared to usual care (Aim 1), to evaluate the effectiveness of videoconferencing to provide greater increase in knowledge, and decrease in distress and depression as compared to remote phone services (Aim 2a), the moderators of patient outcomes (Aim 2b), and a cost evaluation of the three study arms (Aim 2c). Also, we will conduct a multi-stakeholder mixed-methods implementation evaluation to understand patient, provider and system factors associated with uptake of our remote telegenetic services model, facilitators and barriers to uptake, and recommendations for future adaptation and sustainability (Aim 3). We expect our findings will provide critical data for the basis for further dissemination of these services among cancer survivors and other populations in need of genetic services.

NIH Research Projects · FY 2024 · 2020-07

PROJECT SUMMARY. Mycobacterium tuberculosis (Mtb), the causative agent of the disease tuberculosis (TB), is estimated to infect one-fourth of the world's population, resulting in approximately 1.6 million deaths each year. The emergence of multidrug- and extensively drug-resistant Mtb strains and the variable efficacy of the currently used vaccine, M. bovis Bacille Calmette Guerin (BCG), are barriers to the global control of TB. Thus, there is a critical need to better understand the mechanisms of TB immunopathogenesis, as such mechanisms can be targeted to improve host control of Mtb infection. The tubercle granuloma is long been considered a hallmark of TB. Our published data suggest that the presence of inducible bronchus-associated lymphoid tissue (iBALT)-containing granulomas is indicative of protective granulomas that mediate Mtb control during TB latency. In contrast, infiltrating myeloid derived suppressor cells (MDSCs) as well as neutrophils producing proinflammatory molecules are characteristic of non-protective granulomas during pulmonary TB. MDSCs are induced during pulmonary TB in humans, nonhuman primates (NHPs) and mice and suppress protective T cell responses. Our new data show a protective role for the proinflammatory cytokine, Interleukin (IL)-17 in dampening lung MDSC accumulation and limiting T cell suppression in the lung during TB. Additionally, we show that the MDSC-derived proinflammatory proteins, S100A8/A9 heterodimers are induced upon Mtb infection in humans, NHPs and mice. Furthermore, S100A8/A9-expressing myeloid cells accumulate within the tubercle granuloma and amplify lung MDSC accumulation to mediate Mtb susceptibility. In the current proposal, using mouse and NHP models of TB, we will elucidate the mechanism(s) which regulate and promote MDSC accumulation during TB, and characterize whether MDSCs and their pathways can be targeted as host-directed therapeutics (HDTs) for TB. In Specific Aim 1, using gene deficient and conditional gene deficient mouse models we will determine the IL-17-dependent pathways that limit MDSC accumulation during TB. In Specific Aim 2, we will evaluate the role of S100A8/A9 proteins in driving MDSC accumulation and susceptibility to TB, and also determine whether blocking S100A8/A9 signaling will limit TB relapse. Finally, in Specific Aim 3 we will evaluate if MDSC depletion can prevent TB progression in nonhuman primates (NHPs). At the completion of the aims proposed here, we will have considerably expanded our understanding of the Mtb-specific signaling pathways and factors that positively (S100A8/A9 pathways) and negatively (IL-17 dependent pathways) regulate MDSC accumulation during TB. Additionally, our translational studies in NHPs will enable the use of HDTs to limit MDSCs during TB.

NIH Research Projects · FY 2024 · 2020-07

Checkpoint blockade antibodies targeting PD-1 have demonstrated improved survival in metastatic head and neck squamous cell carcinomas (HNSCC) patients by reactivating effector T cells that have infiltrated the tumor microenvironment. However, PD-1 blockade still has low overall response rates approximating 18%, suggesting that the different treatment outcomes are due to intrinsic differences in the patients' diseases, such as tumor microenvironments. Recently, we have developed a new class of radioenhancers, nanoscale metal- organic frameworks (nMOFs), that can alter the immune microenvironment. Constructed via coordination between hafnium-oxo clusters and porphyrin-like molecules, nMOFs generate both hydroxyl radicals and singlet oxygen in a process termed radiotherapy-radiodynamic therapy (RT-RDT). The objective in this application is to define the mechanisms by which RT-RDT and nMOF-enabled immunotherapy alter the immune microenvironment in order to sensitize HNSCCs to checkpoint blockade. Our central hypothesis is that nMOFs can deliver the CpG oligodeoxynucleotides and synergize with RT-RDT-induced antigen release and Type I IFN expression, which stimulates CD8+ and CD4+ T cell proliferation and infiltration into HNSCCs to regress both irradiated and non-irradiated tumors treated with PD-1/PD-L1 blockade. The goal for this proposed research is to identify a novel therapy and define the mechanisms by which it alters the immune microenvironment to sensitize HNSCCs and possibly other cancers to current clinical immunotherapies. This project will use innovative molecularly tunable nMOFs having unprecedented radioenhancement via the unique RT-RDT mechanism. This proposal is significant because it addresses an unmet need of treating radioresistant and metastatic HNSCCs both directly via RT-RDT and by acting as an immunostimulant to enhance the efficacy of existing checkpoint inhibitors. This proposal will test the central hypothesis by pursuing four specific aims: (1) define the cellular mechanisms of innate immune activation after RT-RDT; (2) determine how RT- RDT affects the tumor microenvironment in squamous cell cancers; (3) evaluate the contributions of different immune components on the efficacy of RT-RDT and immunotherapy combinations; and (4) determine effective therapies for HNSCCs resistant to PD-1/PD-L1 blockade. Aim 1 will treat cells and ex vivo stimulated or cultured immune components with nMOFs and radiation to determine how RT-RDT initiates STING and Type I interferon signaling in the tumor microenvironment. Aim 2 will determine how the tumor microenvironment and extracellular matrix are affected by nMOF-mediated RT-RDT. Aim 3 will evaluate the contribution of different immune components to the anticancer efficacy of nMOFs. Aim 4 will use primary oral tumor models that are resistant to PD-1/PD-L1 blockade as a model to identify novel immunotherapy combinations that synergize with RT-RDT. Ultimately, this project will afford new therapeutic strategies using clinically relevant nanomedicines to enhance both the radiation therapy and immunological rejection of HNSCCs.

NIH Research Projects · FY 2024 · 2020-07

Project Summary — Translational regulation has emerged as a key process in the evolution of hepatocellular carcinoma (HCC)1,11. Support of rapid proliferation in cancer requires enhanced protein production as well as gene-specific translational changes that facilitate reprogrammed cellular activities. Recent research has revealed that ribosome composition, including rRNA modification stoichiometry, can affect translational function in cells and bias translation of oncogenic transcripts, altering cell state6,17–20. Although 2% of all rRNA bases are modified, only two of them are N6-methyladenosine (m6A) — 28S m6A4220 and 18S m6A1832. Our lab previously characterized the 28S m6A4220 methyltransferase (ZCCHC4)21, and we have now biochemically characterized METTL5 (M5) as the 18S m6A1832 methyltransferase, stabilized by the cofactor TRMT112. The function of this site is still unknown, but structural analysis suggests roles in translation initiation and re-initiation events29,31, as it is near the mRNA channel and at the binding site of ribosome recycling factors. Furthermore, our M5 knockout HeLa cells display a markedly hypoproliferative phenotype while overexpression of M5 has been associated with hyperproliferation, including in hepatocellular carcinoma (HCC)8,9,22. We have also shown by imaging and biochemical fractionation that M5 is localized in both the nucleolus and cytosol, suggesting that the 18S m6A site may be dynamically methylated in the cytosol, even after ribosome biogenesis, to regulate and/or fine tune translational processes. The goal of this proposal is to define the functional effects of M5 on translation and investigate the mechanism by which it supports cell proliferation and cancer development. We hypothesize that M5 dynamically methylates m6A1832 in response to oncogenic cell stress with functional consequences in ribosome composition, translational function, and ribosome recycling activities that support proliferation and tumorigenesis. To investigate this hypothesis, we will first thoroughly define the impact of M5 on ribosome composition and function as follows: quantifying M5 and m6A1832 under normal and stress conditions by LC- MS/MS and HPLC; characterizing the effect of M5 on translation through nascent protein synthesis assays, ribosome profiling, and translation reporter assays; and monitoring M5-related changes in stoichiometry of ribosomal proteins and translation-related factors. Then, we will investigate the role and mechanism of M5 in HCC proliferation and tumorigenesis by examining the effects of M5 on ribosome binding and translational activities of ribosome recycling factors, and by evaluating the role of M5 in proliferation of HCC cell lines and tumorigenesis in HCC xenograft mouse models with respect to ribosome recycling processes. Successful completion of this proposal will unveil the function of 18S m6A1832 in translation, clarify the link between M5 and HCC prognosis, potentially guide new translation-based therapy development for HCC, and contribute to our understanding of how dynamic regulation of rRNA modifications can affect the proteome and cellular state in support of cancer development.

NIH Research Projects · FY 2025 · 2020-07

PROJECT ABSTRACT The developing mammary gland (MG) is vulnerable to environmental and lifestyle risk factors that increase breast cancer (BC) burden in later adulthood. Therefore, optimizing BC prevention and care requires a lifespan approach to identify specific early life risk factors, to understand these risk factors’ underlying molecular mechanisms in promoting cancer risk, and to design appropriate interventions that reduce BC in adulthood. Using a Sprague-Dawley rat model of human BC, we have established a dynamic and successful transdisciplinary collaboration among a breast cancer biologist, an endocrinologist, and a biopsychologist to understand how adverse early life exposures lead to increased mammary cancer risk in adulthood. We find that glucocorticoid (GC) reactivity to everyday stressors is heighted by social isolation in puberty and young adulthood and is associated with increased adult mammary cancer burden. Moreover, heightened GC reactivity during puberty impairs ductal development and increases mammary stem cell populations, two characteristics that have been linked to increased mammary cancer. We now propose to determine how heightened GC reactivity disrupts MG development and increases cancer burden by examining the underlying molecular mechanisms connecting glucocorticoid receptor (GR) activation with MG developmental defects (Aim 1). In Aim 2 we will introduce both pharmacological- and social environmental-interventions in early adulthood to reverse heightened stress reactivity. We predict these interventions will restore normal MG ductal differentiation and thereby decrease later cancer risk. In Aim 3, we will examine how heightened GC reactivity during puberty inappropriately preserves mammary stem cell (MaSC) populations that are known to increase later cancer risk. We will also investigate the association between circulating steroid hormone levels, in conjunction with their localized production within the MG microenvironment, and ductal maturation and MaSC biology. Completion of these studies will uncover novel stress-mediated molecular and cellular mechanisms of disrupted MG development linked to subsequent mammary cancer and determine whether these stress- mediated events are reversible with early adulthood interventions.

NIH Research Projects · FY 2024 · 2020-07

The long-term goal of the University of Chicago Neuroscience Early Stage Scientist Training Program (NESSTP) is to diversify the Neuroscience research workforce. In order to increase underrepresented Scholars' readiness and success in the neuroscience research workforce, we propose to implement interventions at critical transition stages along the academic pathways. To accomplish this goal, we will take advantage of the unified campus and single faculty at the University of Chicago which hosts undergraduate, graduate and postdoc training in both basic science and medical areas. This will allow us to offer enhanced training for scholars at all three training levels, and also to emphasize cross-level mentoring in order to facilitate young scientists' development as leaders and trainers in Neuroscience. To enhance the undergraduate Neuroscience pipeline, we will provide research experiences, aid in articulating career goals, and facilitate graduate school preparation for our underrepresented (UR) students pursuing Neuroscience majors in the College. To support UR graduate students' and postdocs' academic and career pathways, we will provide professional skill development training, career exploration and networking opportunities to aid academic career success. Finally, we will foster a continuity of Mentorship across all career stages by introducing good mentoring practices for the trainees and their faculty mentors, pairing NESSTP trainees across stages to engage in near-peer mentoring, and establishing a pan-Neuroscience Mentoring Committee to establish good mentoring practices and promote a diverse and inclusive Neuroscience training community at the University of Chicago.

NIH Research Projects · FY 2026 · 2020-06

ABSTRACT Chicago’s community areas demonstrate wide variation in environmental quality, disease risk, and outcomes, indicating a critical need for environmental health research across the city. In 2017, the “ChicAgo Center for Health and EnvironmenT (CACHET)” was established at UChicago – the first NIEHS P30 Environmental Health Core Center in the Chicago area. As the CACHET membership and projects have grown, so have prospective training opportunities. As a key CACHET goal is to improve environmental health-related outcomes for residents of Chicago and beyond, it offers the ideal context for encouraging undergraduates to pursue careers in environmental health. Multidisciplinary environmental health research training is offered by clinician, laboratory, and population scientists to understand, evaluate, and ultimately improve environmental health-related outcomes. Research areas of emphasis include air, water, and soil, biomarkers of exposure and effect, and molecular and cellular processes of toxicity. CACHET offers a robust structure to support training for undergraduates, with extensive research and mentoring opportunities by CACHET investigators and ongoing research opportunities. Since 2021, we have leveraged CACHET, its EHS faculty pool, and its research portfolio to support educational activities that encourage students to pursue further studies or careers in EHS research through the Chicago UP on the Environmental Health Sciences Program (UP on EHS). To accomplish this goal, we: 1) provide 2-year mentored research experiences and career/skill development to undergraduate students (rising juniors from institutions across Chicago) to inspire and prepare them for further study in the environmental health sciences; 2) provide opportunities for dissemination in Chicago communities as well as in academic publications or presentations; and 3) create a strong and long-lasting network of support for students throughout their career journeys. For each student, we assemble a dedicated mentor team, provide hands-on research training, and tailor support and projects to trainee needs and interests. We also communicate with program alumni and offer personalized assistance and encouragement. In the first 4+ years of funding, this has translated into very positive outcomes for our trainees, illustrating our dedication to preparing students for graduate school or careers in research. We have proven that our leadership team has the scientific, educational, and research programs necessary to ensure the continued success of Chicago UP on EHS. We plan to continue engaging cohorts of undergraduates in this program to build upon the strong NIEHS P30 research/researcher infrastructure.

NIH Research Projects · FY 2025 · 2020-05

ABSTRACT The pathogenesis of type 1 diabetes (T1D) involves complex intercellular interactions in the pancreatic microenvironment, where endocrine, exocrine, and immune cells drive early inflammatory responses. Macrophages are among the earliest immune cells to infiltrate the islets, initiating a dialog with β cells that activates adaptive immune responses, promoting autoimmunity and β-cell loss. Our recent work identified polyamines, small organic molecules involved in cellular growth and stress responses, and hypusine, a polyamine-derived modification unique to the translation factor eIF5A, as critical regulators of mRNA translation and inflammation in β cells. eIF5A requires hypusination for its activity and influences specific mRNA translation linked to inflammatory pathways. As polyamine and hypusine biosyntheses can be modulated by diet or small molecules, they offer real-world therapeutic targets to modify T1D progression. We hypothesize that the polyamine biosynthetic pathway in early T1D orchestrates post-transcriptional inflammatory responses in β cells. In the following Aims, this renewal application will leverage new findings, innovative animal models, and cutting-edge technologies to test this overarching hypothesis: Aim 1: Expose the pivotal role of β-cell ornithine decarboxylase (ODC) in the onset of T1D. Aim 2: Decipher how hypusine modification shapes β-cell inflammation and drives T1D progression. Aim 3: Reveal how β-cell polyamine metabolism modifies the inflammatory response in early T1D. Whereas T1D is an autoimmune disease, therapies exclusively targeting the immune system have seen variable success. Recent clinical successes using drugs that block inflammation and stress pathways more broadly suggest a need to revise therapeutic approaches to T1D. Collectively, this study will significantly advance our knowledge of how polyamine biosynthesis and hypusine modification scale inflammatory responses, β-cell function, and survival in early T1D. Through these aims, we will uncover regulatory pathways that could serve as novel therapeutic targets to preserve β-cell mass and function, addressing critical gaps in current strategies for T1D prevention.

NIH Research Projects · FY 2026 · 2020-05

Predicting the effects of noncoding sequence on gene regulation and complex traits remains one of the most important problems in genomics. Many expression quantitative trait loci do not overlap enhancers or promoters. Our previous work showed that about one third of all expression quantitative trait loci likely function after transcription initiation. Further, we have implicated a deep contribution of pre-mRNA splicing to human variation. Assays beyond RNA-seq that report on the pathway of RNA splicing and in a manner independent of decay are sorely lacking, significantly compromising our ability to account for how, and which, genetic variants affect RNA splicing. We have been developing genome-wide assays that explicitly target pre-mRNA splicing. By one assay, we uncovered an under-appreciated mechanism contributing to gene expression and human phenotypic variation: unproductive splicing, which results in transcripts subject to cytoplasmic nonsense- mediated decay. By another assay, we have defined the timing of splicing genome-wide and in doing so discovered an unexpected degree of non-canonical splicing that competes with canonical splicing. In this grant, we propose novel assays to explicitly and efficiently measure canonical and non-canonical splicing. We will develop analytical methods to measure inter-individual variation in these processes and use this variation to study the mechanisms and impact of these regulatory phenotypes. At the end of this project, we will have developed innovative genomic assays that will illuminate novel RNA splicing biology and will have tested hypotheses regarding the function and regulatory mechanisms of RNA splicing. Our work will help improve our understanding of functional gene regulatory variation.

NIH Research Projects · FY 2026 · 2020-05

Abstract The long term objectives of this project are to explain the basis for the complex genotype-phenotype relationships for a growing number of severe calcium channel gene disorders and develop therapies based on these new insights. To make these advances we will capitalize on our new discovery that at least three of these calcium channel genes are bicistronic i.e. they encode two distinct proteins, the calcium channel proteins and a newly discovered transcription factor. We have discovered that the transcription factor is translated by internal translation by a process resembling an internal ribosomal entry site (IRES). We hypothesize that mutations in these Ca2+ channel genes may have a diversity of outcomes, affecting, in distinct cases, neuronal firing, calcium signaling, regulation of the expression of the transcription factor or directly altering the function of the transcription factor. In this study we will systematically explore the function and biological action of these three novel transcription factors, how their expression is regulated by the IRES, how normal cellular physiology governs their translocation to and from the nucleus, and how different mutations affecting channel gating, IRES function and transcription factor cause impaired neuronal development and or viability in these different disorders. We will study this using recombinant calcium channels expressed in primary neurons and human reprogrammed neurons from normal and patient sources, and in transgenic mice expressing well characterized mutations. We will study gene binding and expression using next generation approaches, nuclear translocation using epitope and fluorescent tags with physiological stimuli, IRES function using dual luciferase reporters and immunoblotting, neuronal development using immunofluorescent microscopy and corrective therapy using antisense oligos, miRNA and AAV viral vectors expressing transcription factors.

NIH Research Projects · FY 2026 · 2020-05

PROJECT SUMMARY Molecular simulations complement experiments by revealing the microscopic dynamics underlying biological mechanisms and the forces promoting those dynamics. However, most biological processes involve timescales much longer than the time step of numerical integration. While there are many methods for bridging this separation of timescales to obtain equilibrium averages, further advances are needed to estimate dynamical statistics robustly. During the current award period, we developed basis-expansion and neural-network methods for computing dynamical statistics from the trajectories of short, unbiased molecular dynamics simula- tions. These methods have the advantage that the simulation initial conditions can be distributed arbitrarily, and we developed methods that target regions of configuration space to allocate computational resources efficiently. We tested our methods on established benchmarks and, in collaboration with experimentalists, used them to learn mechanisms of insulin dimer dissociation and phenol release, protein fold switching, and voltage sensing. We now seek to expand our studies of voltage sensing and to launch studies of mechanosensing. Specifically, we will study voltage-gated sodium channels (Navs), which initiate action potentials in nerves and muscles. Navs are targets of therapies for epilepsy, paralysis, chronic pain, and cardiac arrhythmia. Advances in cryoelectron microscopy are yielding many new structures of Navs, enabling simulations. Navs have a conserved tetrameric structure in which each monomer consists of a four-helix voltage-sensing domain (VSD) coupled to a pore domain. We will investigate how specific sequence differences give rise to differences in kinetics of VSD activation, how VSD activation is coupled to opening of the pore, and conduction. In parallel, we will investigate the dynamics of proteins that convert mechanical forces from sound waves and head movements to electrical signals in the inner ear. Protocadherin-cadherin complexes link the tips of protrusions (stereocilia) on hair cells in the inner ear, and stretching these complexes opens ion channels. These dynamics are essential for hearing and balance. We will study the mechanical response of tip-link complexes and the transport of antibiotics through the associated channel, which can lead to cell death (cochleotoxicity) and hearing loss. For both voltage- and mechanosensing, our methods will enable us to go beyond existing simulation studies to characterize kinetics under physiological conditions (voltages and stretching forces/speeds). Beyond providing fundamental insights into systems of direct relevance to human health, these demanding applications will drive development of the next generation of methods. We will marry our short-trajectory methods with graph neural network architectures that we introduced recently to compute dynamical statistics and learn mechanisms with minimal reliance on manually chosen system descriptors. We will accelerate our computational workflow by incorporating generative models within mathematical frameworks with well-defined physical limits to ensure that we obtain accurate statistics.

NIH Research Projects · FY 2026 · 2019-12

ABSTRACT Staphylococcus aureus infection is a frequent cause of sepsis in humans, a disease associated with high mortality and without specific intervention. The development of immune therapeutics, vaccines and monoclonal antibodies against S. aureus, has been a major goal since the discovery of this pathogen. These approaches have focused on antigen selection, targeting key secreted toxins or bacterial surface molecules such as capsular polysaccharides; the former aiming to harness neutralizing antibodies, the latter, complement-fixing antibodies. However, many clinical isolates of S. aureus do not produce capsular polysaccharides, and several failed clinical trials have taught us that toxin-neutralization is not sufficient to ameliorate disease (NCT02940626, NCT01589185, NCT03816956). Here, we propose to pursue monoclonal antibodies, 3F6 and 2A12, that target the surface proteins, SpA and ClfA, respectively. Why these antigens? SpA is conserved and extremely abundant on the bacterial cell surface where it binds the constant region (Fcg) of IgG thereby blocking the effector functions of antibodies. ClfA is the agglutinating factor of S. aureus, it promotes the shielding of bacteria in fibrin cables and their physical escape from phagocytes. Our earlier work demonstrated that the constant region of humanized 3F6-hIgG1 can be optimized with Fcg amino acid substitutions to escape inhibition by SpA. Further, Fc g interactions with complement component 1q (C1q) or Fc g-receptors to promote complement- dependent or antibody-dependent cell-mediated phagocytosis, respectively, can be controlled by engineering Fcg glycans at Asn 297. Here, we will compare antibodies for their ability to reduce bacterial replication in freshly drawn blood and in blood stream models of infection and re-infection in adult and neonatal mice. We will ask if S. aureus clearance by 3F6 antibodies is augmented when agglutination is simultaneously inhibited by anti-ClfA antibodies (2A12) and whether phagocytic uptake is more efficient when antibodies engage C1q or Fc g-receptors. We will also generate 3F6 and 2A12 IgM variants as multimeric IgM is the most potent activator of C1q. Using this collection of antibodies, we propose to identify the cells that promote the clearance of antibody-bound bacteria and the pathways activated by these immune complexes. A future goal of this research is to exploit monoclonal antibodies to promote both the clearance and uptake of this pathogen by antigen presenting cells to improve immune outcomes and curb the problem of recurring infections.

NIH Research Projects · FY 2025 · 2019-09

Characterizing the Molecular Basis of Supergene Mimicry in Butterflies Project Summary Why is life on Earth so diverse? How do these diverse forms arise, both over evolutionary time and during development? What are the functional molecular and genetic changes that underlie the fantastic organismal diversity we see in the natural world? My research focuses on these big questions related to the origin of biodiversity, but I do so through the lens of mechanism. Butterflies, and butterfly wing patterns in particular, offer an excellent system to unlock the functional molecular and genetic mechanisms responsible for organismal evolution because of their natural diversity, a long history of research that puts this diversity in a much-needed ecological context, and our success at establishing genomic and genome editing tools for butterflies. My research team and I study diversity using multiple approaches, including molecular and population genetics, genomics, developmental biology techniques, functional genomics, genome editing, neurobiology, behavioral experiments, and computation. The ultimate goal of this work is very fundamental: I strive to characterize the functional mechanisms responsible for historical evolutionary processes while also uncovering basic principles of biological patterning, development, and behavior. Much of our research is focused on the phenomenon of sex-limited polymorphism, which is widespread in animals but is not well understood in any organism. “Supergene” mimicry in the swallowtail butterfly Papilio polytes stands out as a particularly striking example of sex-limited polymorphism and one that is amenable to functional characterization. While much theoretical work has explored the evolutionary dynamics of supergene mimicry, we are just beginning to unpack its molecular and developmental basis. Previously we determined that the gene doublesex controls the mimicry switch in P. polytes and we have studied the origin and evolution of mimicry, behavioral aspects of mimicry, natural selection in nature, and we have developed tools and methods for CRISPR and multiple functional genomics assays. Using these tools, we have begun to characterize the cis-regulatory architecture of doublesex, as well as the gene regulatory networks that are modified by doublesex to produce novel mimetic wing patterns. Over the next five years, we will investigate the functional basis of supergene mimicry in P. polytes by integrating genomics, functional genetics, molecular and developmental biology, providing the single most comprehensive investigation of its kind. Furthermore, we will greatly expand the scope of our analyses by branching out on the butterfly phylogeny to functionally characterize parallel evolution of supergene mimicry in other butterflies. Our work will greatly expand the known role of the sexual differentiation pathway and generate general insights into fundamental evolutionary genetic processes of convergence, cis-regulatory evolution, and gene co-option.

NIH Research Projects · FY 2026 · 2019-09

Abstract This is a competitive renewal application for a second term of this project (years 5-8). Characterizing gene regulatory differences between humans and our close primate relatives is essential for understanding the molecular basis of human-specific traits. Comparative genomics provides us with the tools to identify both species-specific and conserved regulatory features, which has provided valuable insight into the genetic architecture of adaptation in gene regulation. However, ethical and practical considerations preclude comparative studies of molecular traits in live primates, particularly apes. Frozen post-mortem tissues from non-human apes are difficult to obtain, and even when they are available, they are not optimal templates for many functional genomic assays. Thus, we are generally unable to collect samples from enough individuals to map and study gene regulatory loci in non-human apes; we are unable to study gene regulation in more than a few tissues from apes; we are unable to study the dynamics of gene regulation during development; and we are unable to study regulatory responses to evolutionarily and clinically relevant exposures. In the first term of this project, we proposed to address this challenge by establishing a comparative panel of induced pluripotent stem cells (iPSCs) from humans and chimpanzees. We have successfully done this, and we have shared these lines freely and without restriction with more than 30 labs (see letters of support), facilitating comparative functional genomic studies by investigators who would not otherwise have been able to obtain appropriate samples to conduct their research. We now request support to continue to maintain and distribute this community resource, which we will expand in two ways. First, we will generate additional chimpanzee iPSC lines using the remaining samples we collected prior to the moratorium on chimpanzee research. We expect that these lines will soon become the only population-sample resource for future studies involving chimpanzees in the USA. Second, we will use human and chimpanzee iPSCs to develop a series of dynamically differentiating organoids, which we will also share freely to allow other groups to effectively utilize the comparative iPSC panel. The unique property of these dynamically differentiating organoids is that they contain asynchronously differentiating cells. While they do not produce pure cell populations that can be meaningfully analyzed with bulk RNA-seq data, single-cell RNA-seq can be used to deconvolve the organoids into dozens of cell types and developmental stages, allowing us to explore dynamic regulatory processes that cannot be observed in adult tissues10.

NIH Research Projects · FY 2026 · 2019-07

Abstract All multicellular organisms are colonized with a diverse and complex population of microbes including bacteria, viruses, fungi, archaea, and protozoans, collectively called the microbiota. Microbiota play an integral role in modulating host health by providing necessary nutrients, by protecting against incoming bacterial pathogens and by supporting the development and maturation of the immune system. Microbiota could also impact cancer development at various stages by modulating cell proliferation and death, altering the function of the immune system, and influencing metabolism of various compounds. Murine Leukemia Virus (MuLV) is highly proficient in causing leukemia in mice from susceptible strains. Using MuLV, we found that some gut commensal bacteria promoted the development of leukemia induced by this retrovirus. The promotion of leukemia development by commensal bacteria was due to suppression of the adaptive immune response through upregulation of several negative regulators of immunity. These negative regulators included serine (or cysteine) peptidase inhibitor, clade B, member 9b (Serpinb9b) and Ring finger protein 128 (Rnf128). Serpinb9b is known to counteract killing by cytotoxic granzymes and was upregulated in leukemic cells. Rnf128 known for induction of unresponsiveness of T cells was induced uniquely in CD4 T cells. Genetic ablation of these genes conferred resistance to virally-induced leukemia even in the presence of a complex microbiota. Upregulation of Serpinb9b was mediated via the RIPK2 pathway (downstream of NOD1 and NOD2 receptors which detect various forms of peptidoglycan). As Serpinb9b and Rnf128 are associated with a poor prognosis of some spontaneous human cancers, the mechanism of bacterially-induced immunosuppression during tumor development may apply to human tumors of virally and non-viral origin. The current proposal aims at defining the precise mechanism(s) by which microbially-induced negative immune regulators contribute to leukemia progression. Specifically, we will identify the innate immune receptors bacterial products signal through to induce negative immune regulators and determine the adoptive mechanism which controls leukemia development in the absence of bacteria.

NIH Research Projects · FY 2024 · 2019-07

PROJECT SUMMARY Individuals’ adaptive immune responses are central to the epidemiology and evolution of influenza and the effectiveness of influenza vaccines. It is therefore surprising that despite nearly 70 years of study, major questions about the immune response to influenza remain unanswered. In particular, it is unclear how well natural infection protects from reinfection with the same or related types and subtypes, how vaccination affects protection against symptomatic and asymptomatic infections over time, and how protection varies with immune history, age, individual, sex, and other factors. The two main obstacles to progress have been a shortage of observations from the same individuals over time and a lack of modeling approaches that can accommodate the complex, stochastic dynamics of infection and immune response replicated across individuals. The proposed research takes advantage of an extraordinary influenza cohort and new methods for longitudinal modeling to understand how protection to influenza infections of varying severity arises, and especially how it is shaped by infection and vaccination history. The ongoing Nicaragua Pediatric Influenza Cohort Study (NPICS) has followed thousands of children since 2011 and recorded their antibody titers, infections, symptoms, and vaccination history to influenza. We will use these data to fit and evaluate a large set of stochastic, individual-level, mechanistic, dynamical models to estimate the duration of protection and its dependence on exposure history and other factors. First, we will estimate the duration of protection against reinfection with the same type or subtype and evaluate its dependence on the order of early exposures and host and viral characteristics. Next, we will measure the strength and duration of cross-protection between type and subtypes. Finally, we will compare the dynamics of protection after natural infection to those after vaccination, including repeat vaccinations. Our flexible modeling approach takes advantage of diverse data types and inference techniques while allowing precise formulation of biological hypotheses mathematically. Its recent success with similar longitudinal datasets of PCR-confirmed viral infections and influenza serology demonstrates feasibility. Preliminary results suggest a role of exposure history on heterosubtypic infection risk. This work is poised to advance basic knowledge on influenza and the development of immune memory, and it will provide a new set of dynamical modeling tools for longitudinal data. This project will thus achieve NIH MIDAS objectives by advancing the development of inference techniques and software for an important and growing type of data and by expanding knowledge of an important host-pathogen dynamic. This work also directly addresses priorities established by the NIH Strategic Plan for the development of a universal influenza vaccine, especially identifying factors associated with the severity of influenza (objective 1.2) and improving understanding of how and when exposure to influenza antigens shapes the response to infection and vaccination (objective 2.1).