Brigham And Women'S Hospital

NIH Research Projects · FY 2025 · 2016-08

Breaking down barriers to translational research is the key to finding new approaches to inflammatory arthritis and related diseases in adults and children. Four years ago, with P30 support, we built the Joint Biology Consortium (JBC), a shared infrastructure based at the Brigham and Women’s Hospital and Boston Children’s Hospital, to accelerate the work of an arthritis-focused Research Community now spanning 18 research centers across the United States and abroad. Based on the success of the JBC, we propose to continue and expand the fundamental design of three Cores designed around the shared needs of JBC members to enhance the efficiency of existing studies, facilitate innovation, and foster junior investigators. 1. The Administrative Core is the organizational heart of the JBC, coordinating operations and cultivating the scientific potential of the JBC research network, coordinated through the JBC Web Portal. The JBC Synergy Meeting and Visiting Professor program promote scientific interchange and the JBC Enrichment Program incorporates innovations from award-winning mentors to support JBC Young Investigators via grant aims review, mentoring, and 12 yearly JBC Microgrants of $5,000 to facilitate utilization of JBC services for pilot and feasibility studies. 2. The Human Biosamples Core provides “one-stop shopping” for adult and pediatric biospecimens essential to research in arthritis and related diseases. The HBC leverages 19 distinct sources of samples. The new JBC Recruitment Core provides targeted prospective recruitment from over 100,000 consented individuals based on phenotype and, in many cases, genotype to build a pipeline for biospecimens across the lifespan. 3. The Cellular Systems Core provides resource- and expertise-intensive tools for arthritis research. Next-generation sequencing tools include single-cell RNAseq, CITE-seq, ATAC-seq, T-scan, PhIP-seq, and spatial transcriptomics, while protein-based services include CyTOF, tissue mass cytometry, and custom Luminex, all analyzed with assistance and education from the new JBC Bioinformatics Core. Spearheaded by committed investigators and mentors Director PI/PD Dr. Peter Nigrovic and Associate Director and Co-PI Dr. Jeffrey Sparks, the JBC spurs innovation within a highly collaborative network of senior and junior investigators as an engine of translational research in adult and pediatric arthritis and related disease. *

NIH Research Projects · FY 2025 · 2016-08

ABSTRACT Obstructive Sleep Apnea (OSA) is a prevalent disorder that remains undertreated due to poor adherence with continuous positive airway pressure (CPAP). Several studies suggest that the pharyngeal airway can collapse at different sites in patients with OSA (velum, oropharyngeal lateral walls, tongue base, and/or epiglottis). Furthermore, many of the non-CPAP treatments for OSA have been shown to work better for some sites of collapse than others. The problem is, the site of collapse requires expensive or invasive procedures, such as drug-induced sleep endoscopy (DISE). This proposal seeks to solve that problem by determining the site of collapse from acoustic analysis of snoring sounds. In Aim 1, a model for predicting the site of collapse will be developed using DISE with simultaneously measured snoring sounds in 800 individuals. In Aim 2, natural sleep endoscopy will be performed in a subset of people from Aim 1 to confirm that the collapsing structure(s) produce the same sound in natural sleep as drug-induced sleep. Finally, Aim 3 will test the ability to measure snoring sounds in the home environment. Comparisons will be made between snores from natural sleep endoscopy and natural sleep at home (in the same individual). Additionally, snoring will be measured at home on separate nights to test reproducibility across nights. The studies in this grant are expected to produce an algorithm for estimating the site of pharyngeal collapse from snoring sounds. This algorithm could improve the selection of patients for non-CPAP therapies and potentially increase the number of patients effectively treated for OSA.

NIH Research Projects · FY 2025 · 2016-07

Project Summary/Abstract: Overall Component Mouse models have provided many biologic insights and remain the most popular system in which to conduct skin disease research. However. there are significant differences between the skin and immune systems of mice and humans and these differences are incompletely characterized, making it difficult to know if observations made in mice will hold true in humans. Research carried out on human cells and tissues can address this knowledge gap. Human biobanks and powerful new analytic techniques have become available that make high-quality human skin disease research accessible. The goals of this Center are to accelerate human skin disease research by providing researchers at any institution with access to human specimens and cutting edge analytic techniques and to bring new diverse investigators into the field of human skin disease research. We include 45 research projects from investigators who wish to utilize Center services; however, any researcher wishing to carry out human skin disease research is a potential member of the research community. The Center is composed of an Administrative Core and three Resource Cores. The Administrative Core manages and oversees all activities of the Center and administers Diversity and Outreach activities, including funding for diverse investigators at multiple stages of training and the biennial International Conference on Human Skin Disease. The Human Tissues Biobank Core provides access to over 113,000 highly characterized consented patients, over 1.5 million banked pathologic specimens, both searchable by diagnosis, as well as to fresh human skin, purified cell populations from human skin and immunodeficient mice grafted with human skin and blood. The Single Cell and Immunoanalysis Core provides access to single cell RNA and ATAC sequencing, flow cytometry-based single cell imaging, mass cytometry by time of flight (CyTOF) and high throughput TCR sequencing. The Next Generation Tissue Analysis & Imaging Core provides access to six color tyramide amplification based immunostaining combined with spectral imaging and automated cell analysis, NanoString RNA and DNA profiling and Digital Spatial Profiling, a state-of-the-art technique that that can profile expression of 10-1000s of protein or RNA targets in FFPE and frozen sections. In summary, the Center provides access to biobanks and cutting-edge human analytic techniques that enable translational researchers to carry out high quality human skin disease research.

NIH Research Projects · FY 2026 · 2016-06

PROJECT SUMMARY Effector and regulatory T cells targeting the central nervous system (CNS) play a pivotal role in the pathogenesis of multiple sclerosis (MS) and its model experimental autoimmune encephalomyelitis (EAE). Dendritic cells (DCs) control CNS-specific T cells, but the pathways regulating DC function are poorly characterized. Published studies suggest a role for the ligand-activated transcription factor aryl hydrocarbon receptor (AHR) in the control of DCs. During the previous grant cycle we have made the following findings: 1) Deletion of AHR expressed in classical DCs (AHRcDC) worsens EAE and exacerbates autoimmune T-cell responses, 2) Single-cell RNA-seq analyses of mouse and human classical DCs (cDCs) detected the coordinated expression of AHR with the transcription factor KLF4 and the ectoenzyme CD39, 3) Indeed, AHRcDC drives the expression of KLF4 in cDCs (KLF4cDC), a transcription factor known to limit NF-kB activation and drive anti-inflammatory gene expression in macrophages, 4) KLF4cDC deletion worsens EAE and exacerbates pathogenic T-cell responses, 5) AHRcDC also drives the expression of CD39 in cDCs (CD39cDC), which degrades pro-inflammatory extracellular ATP (eATP) and participates in the synthesis of anti-inflammatory adenosine, 5) CD39cDC deletion worsens EAE and exacerbates pathogenic T-cell responses, 6) AHR activation induces mouse and human tolerogenic cDCs, and 7) A novel probiotic engineered to produce the AHR agonist IAA (named EcNIAA) suppresses EAE in an AHRcDC-dependent manner. Based on these findings we hypothesize that AHRcDC-driven KLF4 and CD39 expression in cDCs limits CNS autoimmunity. Our specific aims are: Specific Aim 1: DETERMINE THE ROLE OF KLF4 IN THE CONTROL OF cDCS BY AHRcDC. We propose to: 1) Define KLF4-dependent and KLF4-independent transcriptional programs controlled by AHRcDC in single cell and bulk genomic studies, and 2) Establish the role of KLF4 in the control of NF-kB by AHR in cDCs. Specific Aim 2: ESTABLISH THE ROLE OF CD39 IN THE CONTROL OF T CELLS BY AHRcDC. We propose to: 1) Define the effects of AHRcDC-induced CD39cDC expression on myelin-specific T cells, 2) Determine whether the AHRcDC-CD39cDC axis promotes regulatory T cell differentiation, and 3) Establish the roles of AHRcDC, KLF4cDC and CD39cDC in the control of T cells by DCs in healthy controls and MS patients. Specific Aim 3: DEFINE THE ROLE OF AHRcDC IN EAE SUPPRESSION BY EcNIAA. This aim evaluates the therapeutic potential of activating AHRcDC with EcNIAA, a novel probiotic engineered to produce the AHR agonist IAA. We propose to: 1) Evaluate the therapeutic effects of EcNIAA in EAE, 2) Define the roles of AHRcDC and KLF4cDC on the transcriptional modulation of DCs by EcNIAA, 3) Establish the contribution of AHRcDC and CD39cDC to the control of effector and regulatory T cells by EcNIAA. IN SUMMARY, in this competitive renewal we use unique tools and clinical samples to study a novel aspect of AHRcDC as a regulator of T-cell autoimmunity, and its potential as a therapeutic target in autoimmune diseases.

NIH Research Projects · FY 2025 · 2016-03

Project Summary/Abstract The proposed project is focused on the crosstalk between metabolism and cell fate during development of the paraxial mesoderm, the tissue which forms skeletal muscles and vertebrae. One striking characteristic of this tissue is its segmentation into repeated structures termed somites, a process driven by a molecular oscillator called segmentation clock [1]. Defects in paraxial mesoderm development can lead to severe malformations such as congenital scoliosis, spina bifida or caudal agenesis. The paraxial mesoderm arises from a population of progenitors located in the primitive streak and tail bud. Remarkably, these progenitor cells exhibit aerobic glycolysis and an inverted intra- vs. extracellular pH gradient, which are characteristic of the Warburg effect of cancer cells [2, 3]. In the tailbud, we demonstrated that glycolysis increases the intracellular pH to promote acetylation of β-catenin and Wnt activation, which ultimately leads to paraxial mesoderm induction [3]. As these processes are difficult to study in vivo, we have developed in vitro systems in which embryonic stem (ES) cells or induced pluripotent stem (iPS) cells can be efficiently differentiated toward the paraxial mesoderm fate recapitulating the normal features of its metabolism, signaling and even oscillations of the segmentation clock [4-7]. We will now take advantage of these in vitro systems, as well as mouse and chicken embryos, to characterize in detail the role of aerobic glycolysis in paraxial mesoderm development to see how it relates to the Warburg effect. In this application, we propose to carry out large scale multi-omics experiments (metabolomics, transcriptomics, proteomics, epigenomics) and live imaging of cellular metabolic state to characterize the impact of metabolic transitions on the regulation of gene expression, protein function and cell fate. As the physiological significance of the Warburg effect is not well understood, carefully dissecting its role in the embryo might help shed light on its role in cancer. Finally, we propose to use in vivo, ex vivo and in vitro systems recapitulating the oscillations of the segmentation clock to study the role of metabolism in the control of the oscillatory period. We will analyze the differences in metabolism regulation between mouse and human paraxial mesoderm cells, with special focus on mitochondrial respiration. The period of the oscillations diverges significantly between the two species and can be used as a proxy for developmental timing to try to understand why human development proceeds more slowly than mouse development. We expect these experiments to shed light on the molecular basis of developmental timing, which is tightly linked to longevity in mammals.

NIH Research Projects · FY 2026 · 2016-01

7. Project Summary The primary goal of this proposal is to develop an effective approach to screening for early stages of pulmonary fibrosis by assessing the diagnostic and prognostic value of clinical, environmental, genetic and genomic factors in at-risk relatives of patients with idiopathic pulmonary fibrosis (IPF). IPF, the most common and severe form of pulmonary fibrosis has a mortality rate comparable to that of many end-stage malignancies. Although IPF has historically been unresponsive to pharmacotherapy, recent studies have finally demonstrated that medical therapy can reduce the rate of decline in lung function, particularly when started early in the course of disease. In the prior grant cycle of this application we demonstrated that first-degree relatives were at high-risk to develop early stages of pulmonary fibrosis and that genetic testing helped to improve risk prediction. Based on these findings, we hypothesize that we will continue to observe a high prevalence of early pulmonary fibrosis in at-risk relatives; that we will be able to develop a clinically useful screening algorithm that combines key clinical, genetic, genomic, and environmental features for the early detection and prognostication of interstitial lung abnormalities (ILA) and/or pulmonary fibrosis in populations of diverse ethnic backgrounds; and that a subset of genes whose reduced expression predicts accelerated disease progression harbor pathogenic variants that help to drive this process. To assess these hypotheses, we propose the following Specific Aims: Aim 1) Develop an algorithm that can be used in clinical practice to identify relatives at the highest risk for pulmonary fibrosis, Aim 2) Prognosis: Define the baseline clinical, genetic, and genomic features in relatives found to have ILA that best predict their risk of disease progression, and 3) Identify novel genetic variants that contribute to pulmonary fibrosis susceptibility using an integrative genomics approach. In addition to providing a greater understanding of the role of that genetic variation plays in the development of IPF, the results from this study will motivate a clinical trial evaluating the use of screening and early therapeutic intervention in relatives at high-risk to develop IPF.

NIH Research Projects · FY 2025 · 2015-12

ABSTRACT The use of RNA technologies to specifically target genetic alterations in tumor cells has shown great potential of becoming a novel therapy modality for cancer treatment. Nevertheless, systemic delivery of RNA agents such as messenger RNA (mRNA) to tumor cells in vivo commonly faces multiple barriers, including low stability, rapid elimination by renal excretion, insufficient cellular uptake, poor endosomal escape, and transient activities. Our long-term objective is to develop robust nanoparticle (NP) platforms for effective and safe RNA delivery to solid tumors, and along with cancer target validation in vivo, to eventually transition the RNA nanomedicines into clinical development. In the last funding cycle, a lipid-polymer hybrid RNA NP system has been engineered with favorable features, such as small size, high RNA encapsulation, efficient cytosolic translocation, and relatively long blood circulation. We have also pioneered the application of these hybrid NPs for mRNA delivery to restore tumor suppressors (e.g., PTEN) in different cancer types including prostate cancer (PCa) and non-small cell lung cancer, which represents a novel approach to cancer treatment that is independent of oncogene antagonism. In our latest work, we further reveal that PTEN restoration in PTEN-null/mutated murine tumor cell lines can induce immunogenic cell death (ICD). Preliminary in vivo studies show that PTEN mRNA NP treatment triggers cytotoxic T cell responses, modulates the immunosuppressive tumor microenvironment, and improves the responses of immune checkpoint blockade therapy. In this renewal application, we propose to i) address the unique challenge of transient bioactivity in mRNA delivery by developing a new generation of hybrid mRNA NPs, and ii) apply the new hybrid mRNA NPs to explore PTEN restoration-induced ICD and evaluate the anti-tumor efficacy of PTEN restoration along with immune checkpoint blockade. Specifically, the three Aims underlying the proposal are: 1) To optimize the new generation of hybrid NPs and study the NP-mediated long duration of mRNA bioactivity with the goal of achieving prolonged PTEN expression in PCa tumors using as infrequent injections as possible; 2) To apply the optimized mRNA NPs to investigate the mechanisms underlying PTEN-mediated ICD and anti- tumor immune responses and to evaluate the therapeutic effect and safety in subcutaneously grafted, orthotopic, and transgenic models of PCa; and 3) To expand the new hybrid mRNA NPs to systemic co-delivery of PTEN mRNA and CpG oligodeoxynucleotide (a toll-like receptor-9 agonist) for stronger ICD and to test the co-delivery NPs for PCa treatment together with immune checkpoint inhibitors. We expect that successful completion of this project will lead to development of a novel synthetic mRNA nanotherapy that could benefit cancer patients with loss/mutation of PTEN. Moreover, this NP delivery strategy could be readily expanded to other tumor suppressor- encoding mRNAs for various malignancies.

NIH Research Projects · FY 2025 · 2015-07

PROJECT SUMMARY/ABSTRACT Overnutrition is central to the pathogenesis of metabolic dysfunction-associated steatotic liver disease (MASLD). This research proposal addresses the unanswered question of how molecular mechanisms that normally promote energy conservation become maladaptive and promote MASLD in response to overnutrition. The long-term goal of this research is to understand the regulatory relationships between cellular lipid molecules and metabolism. The objective of this research is to understand fundamental mechanisms for the regulation of energy homeostasis and nutrient metabolism. Our central hypothesis is that Them1 conserves energy by limiting thermogenesis in brown and beige adipose tissue through its functions both as a lipid- regulated enzyme that reduces rates of fatty acid oxidation, and as a transcriptional co-regulator. In obesity, we postulate that Them1 becomes maladaptive. In addition to limiting energy expenditure in thermogenic adipose tissue, high fat diet-induced Them1 upregulation in liver leads to steatosis and excess gluconeogenesis. The rationale for the proposed research is that the mechanisms by which Them1 limits energy expenditure, while promoting hepatic steatosis and insulin resistance, will reveal specific new targets for the management of MASLD. Guided by extensive preliminary data, the central hypothesis will be tested in three specific aims: 1) To establish the mechanisms by which Them1 regulates triglyceride metabolism; 2) To elucidate transcriptional regulation of nutrient homeostasis by Them1; and 3) To delineate lipid-mediated regulation of Them1 cellular and enzymatic activities. In Aim 1, we will utilize biochemical and biophysical approaches to test the hypothesis that dynamic formation of membrane free biomolecular condensates (puncta) and their dissolution in brown adipose tissue provides a tissue-specific physiological mechanism for rapid suppression of thermogenesis. We will define whether static puncta are responsible for Them1-mediated regulation of hepatic lipid droplets. Aim 2 will test the hypothesis that enzymatically active Them1 within the nucleus regulates gene transcription. Using mouse models and cell culture systems, we will leverage recombinant adeno-associated virus to express wild type and Them1 mutant constructs that probe the influence of cellular localization and enzymatic activity on gene transcription profiles. We will define nuclear interacting partners for Them1 in liver and brown adipose tissue that contribute to the regulation of nutrient homeostasis. Aim 3 will test the hypothesis that lipid molecules bind and regulate the cellular localization and enzymatic activity of Them1. We will determine whether bound lysophosphatidylcholines and fatty acids control the exposure and functional utility of a nuclear localization signal located within the Them1 START domain. We will elucidate the allosteric control of Them1 activity by lipid binding. Overall, this proposal will elucidate Them1-mediated metabolic regulation, which is significant because mechanisms that conserve energy in health may promote disease in response to overnutrition. These studies are expected to identify therapeutic opportunities for the management of MASLD.

NIH Research Projects · FY 2025 · 2015-04

Abstract Kaposi's sarcoma (KS) herpesvirus (KSHV) is the etiologic agent of KS, primary effusion lymphoma (PEL), and multicentric Castleman's disease (MCD). These tumors occur most commonly in individuals with AIDS or other immunocompromising conditions. Currently, there are no specific therapies for these diseases. KS is the leading AIDS malignancy, and is epidemic throughout sub Saharan Africa. KS commonly involves the oral cavity and can widely disseminate to visceral organs. Saliva is the vehicle of transmission for KSHV. Following infection, epigenetic modifications associated with transcriptional activation are deposited on the KSHV genome, leading to widespread, but brief, viral gene expression. Failure to inhibit this expression leads to lytic replication. Repressive H2AK119ub and H3K27me3 modifications subsequently accumulate to silence lytic gene promoters. H2AK119ub accrues initially, followed by H3K27me3, in contrast to the classical model in which H3K27me3 marks precede H2AK119ub. Latency is the hallmark of KSHV and gammaherpesvirus infection. KSHV latently infects cells, including tumor cells, and viral genomes persist as circular, extrachromosomal, multi-copy, episomes. To persist in proliferating cells, viral episomes must replicate, and subsequently, segregate to daughter nuclei. Tumor cell viability is dependent on latent KSHV infection. The latency-associated nuclear antigen (LANA) is one of a limited number of virus genes expressed in latency. LANA is responsible for KSHV episome maintenance and is necessary and sufficient for virus episome persistence in the absence of other viral genes. In addition to episome persistence, LANA exerts important roles in transcriptional regulation and growth control. LANA is involved in silencing the viral genome. We have discovered LANA interacts with a component of the DNA damage response (DDR), and that the DDR silences the viral genome following infection, thereby inhibiting lytic replication and allowing latency establishment. This work will use rigorous, detailed, in depth approaches to investigate the mechanistic basis of these findings. Experiments will investigate the LANA-DDR interaction and its role in viral genome silencing, and suppression of lytic replication. We will investigate the dynamics and sites of deposition of key DDR factors on the KSHV genome and LANA’s role in these events. Experiments will also investigate the role of the DDR in establishing the KSHV repressive epigenome. The silencing of the KSHV genome following infection is central to the establishment of viral latency, and this work therefore provides novel and important insight into a fundamental component of KSHV biology.

NIH Research Projects · FY 2025 · 2015-04

The hypothalamic-pituitary-gonadal (HPG) axis regulates puberty initiation and reproductive function. The timing of puberty initiation is associated with risks for development of a wide range of diseases in adulthood, including obesity, diabetes, cardiovascular/cardiometabolic disorders, and cancer. Pubertal development is a complex process that is regulated by the activity of the HPG axis and is influenced by genetic, nutritional and environmental factors. The HPG axis is active in the embryonic and neonatal stages of life; it is then suppressed during childhood until reactivation at the time of puberty. Premature re-activation of the HPG axis results in central precocious puberty (CPP). The precise mechanisms that regulate GnRH secretion to constrain the HPG axis during infancy and childhood and subsequently trigger puberty initiation remain elusive. Genetic studies of patients with reproductive disorders have led to identification of genes that regulate GnRH secretion and have increased our understanding of the neuroendocrine regulation of reproductive function. We used an unbiased approach to identify loss-of-function mutations in MKRN3 in patients with CPP, linking this imprinted gene with the reproductive axis for the first time. Mutations in MKRN3 are now recognized to be the most common genetic cause of CPP. The long-term goal of this project is to elucidate the molecular, cellular, and physiologic mechanisms by which MKRN3 controls the timing of puberty onset. We hypothesize that MKRN3 acts in the hypothalamus to inhibit the reproductive axis. In the first aim of this proposal, we will study the roles and mechanisms of action of MKRN3 in male and female neural development and synaptic plasticity during puberty. In the second aim, we will examine candidate targets of MKRN3, including KISS1, NKB, IGF2BP1 and LIN28B, as well as mRNA targets, in the regulation of the reproductive axis. In the third aim, we will identify new MKRN3 targets and leverage mutations in key protein domains identified in patients with CPP to investigate the roles of different MKRN3 domains in protein function. The successful completion of these aims will help us to understand the actions of MKRN3, a novel regulator of GnRH re-activation, in the neuroendocrine control of pubertal timing. A better understanding of the role of MRKN3 may also identify novel factors involved in the neuroendocrine control of reproduction and lead to the development of new tools for the management of pubertal and reproductive disorders.

NIH Research Projects · FY 2025 · 2015-03

PROJECT SUMMARY Immune checkpoint receptors (e.g. CTLA-4, PD-1, Tim-3) are expressed on dysfunctional or “exhausted” CD8+ tumor-infiltrating lymphocytes (TILs) that exhibit defective effector functions (cytotoxicity and pro-inflammatory cytokine production) and are thus poor mediators of tumor clearance. In the last decade, immune checkpoint blockade (ICB) has achieved durable responses in many cancers, including melanoma, lung, and renal cancer. Despite this success, current estimates indicate that only 12% of all cancer patients respond to ICB. These observations underscore the remaining unmet clinical need in cancer treatment and the need to understand what constitutes effective response to ICB in order to improve response rates. Through examination of the population and single-cell RNA profiles of CD8+ TILs upon ICB, we have identified stem-like CD8+ TILs that are integral for the response to ICB. These cells are tumor antigen-specific, exhibit polyfunctional effector capacity, and increase in proportion upon various ICBs across different cancer types. Although the transcription factor TCF-1 plays an important role in the maintenance and effector function of these cells, we have found that TCF-1 expression in CD8+ T cells is not requisite for positive response to ICB in all tumor contexts. These observations underscore the relevance of stem-like CD8+ TILs for effective response to ICB and the need to better understand how TCF-1 and additional factors regulate their biology. Stem cells reside in niches where crosstalk between stem cells and other cells in the niche regulates not only their maintenance and function but also the function of niche cells. We have found that tumor-associated dendritic cells (DCs) are altered when TCF-1 is absent in mature CD8+ T cells. We thus hypothesize that 1) stem-like CD8+ TILs reside within niches in the tumor micro-environment (TME) where they interact with and modulate antigen-presenting cells; 2) this intercellular communication circuit within the niche may be required for effective priming of anti-tumor T cell responses; and 3) modulation of stem-like CD8+ TILs may positively or negatively influence this communication circuit, thus affecting the efficacy of ICB. Our overarching goal is to understand the cell-autonomous regulation of stem-like CD8+ TILs, their crosstalk with other cells in the TME, and how together they govern the activation of proficient anti- tumor CD8+ T cell responses and ICB efficacy. Accordingly, we propose the following aims: 1) Determine the role of TCF-1 in the generation of proficient anti-tumor T cell responses; 2) Test the role of novel candidate regulators of stem- and effector-like CD8+ TILs, and 3) Characterize the crosstalk between stem-like CD8+ TILs and the TME.

NIH Research Projects · FY 2025 · 2015-02

The role of fibroblasts in end organ fibrosis is well established, but insights into their roles in chronic inflammatory diseases in peripheral tissues like rheumatoid arthritis (RA) is still emerging. We identified a highly expanded inflammatory subpopulation of fibroblasts in the sublining region of RA synovial tissue. It accounts for >50% of all fibroblasts in the synovium in RA, but it is a rare population in osteoarthritis (OA). The expanded population is distinguished by high expression of CD90 (Thy1, a sublining marker) and HLA-DR, and the production of IL-6 and many chemokines. We hypothesize that these CD90+DR+IL-6+ fibroblasts are key in driving inflammation directly by secreting inflammatory factors and indirectly by recruiting and activating leukocytes to maintain chronic inflammation. When analyzing single cell RNA-seq data from the RA/SLE Accelerating Medicines Partnership (AMP) Consortium, we found that markers of lining and sublining fibroblasts in synovium were not absolute – but instead represented a gradient in gene expression in trajectory analysis. We found that this transcriptional gradient corresponds to an anatomic spatial gradient in the synovium emanating from blood vessels. Our data suggest that Notch signaling is a dominant driver of the gradient starting with fibroblasts around blood vessels and extending to sublining fibroblasts that express Notch3 receptors and Jagged (Jag)1 Notch ligands. Here, we wish to determine if Notch 3 signaling specifically on fibroblasts drives the spatial pattering and the differentiation of sublining fibroblasts. To accomplish this, in Aim 1 we use mixed cell organoids with endothelial tubules and fibroblasts to compare spatial pattering and differentiation of Notch3 deficient compared to control fibroblasts. In Aim 2, we determine the location of the CD90+DR+ inflammatory cytokine producing fibroblasts and Notch3+ fibroblasts in the synovium and determine which fibroblast population(s) most significantly associate with leukocytes (T cells, B cells and macrophages). In Aim 3 we activate synovial fibroblast lines with inflammatory cytokines that are found in RA, in the presence or absence of Notch ligands. We use flow cytometry, RNA-seq, LDA, and trajectory analyses to compare fibroblast cell states induced in vitro with those found in the synovium in RA. Then, we extend the Notch gradient concept from fibroblast differentiation to how fibroblast-derived Notch ligands activate attached T cell in organoids. Finally, in Aim 4, we determine if targeted, conditional disruption of Notch signaling in fibroblasts or targeted conditional deletion of Notch ligands in fibroblasts prevents inflammatory arthritis in mouse models. Together, these studies will advance our knowledge of how fibroblasts differentiate in RA to become drivers of inflammation and pathology in chronically inflamed synovial tissues, and how they might be targeted therapeutically in murine models.

NIH Research Projects · FY 2026 · 2014-09

PROJECT SUMMARY Multiple sclerosis (MS) is a chronic autoimmune disease of the central nervous system (CNS). Despite advances in our understanding of MS pathophysiology, there are minimal disease-modifying treatments or preventions for innate-mediated, secondary-progressive forms of MS. Our long-term goal is to define the role of interactions between microglia and recruited monocytes at different stages of MS and determine which phenotypes and functions lead to progressive forms of MS. We made the following preliminary observations: 1) ITGB8-TGFb signaling regulates astrocyte-microglia crosstalk in neurodegeneration with Irf8 regulation; 2) Deleting microglial Irf8 enhances an MGnD phenotype associated with suppression of TGFb-IFNg upstream regulators; 3) monocytes from SPMS patients have a suppression in IRF8 and induction of neurodegenerative signature; and 4) TGFb-IFNg stimulation restores homeostatic signature in monocytes/macrophages. Based on these findings, we hypothesize that TGFb/IFNg-IRF8 axis induces beneficial microglia and monocyte phenotypes in MS. Here, we propose a multi-disciplinary approach that integrates novel tools, experimental paradigms and our complementary expertise on monocytes, microglia, astrocytes and MS and its pre-clinical models. We will address our hypothesis in the following aims: Aim 1: Investigate the role of TGFb/IFNg-IRF8 signaling in monocytes on the regulation of microglia in MS models. We will 1) Determine the effect of TGFb/IFNg-mediated IRF8 signaling in monocytes on the regulation of microglial phenotypes in EAE and 2) Determine the role of IRF8 in monocytes in the regulation of ITGB8-mediated astrocyte-microglia crosstalk, and 3) Evaluate the effects of monocytes on microglia-astrocytes crosstalk in situ by MERFISH analysis. Aim 2: Modulate microglia-astrocyte crosstalk via IRF8-ITGB8 signaling in MS models. Irf8 deletion polarizes microglia to a neurodegenerative phenotype, activating astrocytes via a Lgals3-ITGB8 negative feedback regulation. To elucidate this crosstalk, we will 1) Conditionally remove IRF8 in pan-microglia during the demyelination stage; 2) Perform fate-map and conditional deletion of Irf8 in MGnD microglia; and 3) Genetically and pharmacologically inhibit ITGB8 in astrocytes. Aim 3: Establish whether modulation of TGFb/IFNg-IRF8 signaling in human monocytes and microglia can serve as a novel therapeutic modality in MS. We will 1) Determine whether restoration of IRF8 signaling via TGFb/IFNg signaling in SPMS monocytes polarizes homeostatic microglia; 2) Elucidate how IRF8 modulation in monocytes affects microglia-astrocyte crosstalk, and whether this effect is ITGB8-dependent; 3) Investigate the role of IRF8 in human microglia and their contribution to demyelination in humanized cuprizone model. IN SUMMARY, this application will elucidate the previously unrecognized role of TGFb/IFNg-IRF8 axis in monocytes-astrocyte-microglia crosstalk in multiple sclerosis progression.

NIH Research Projects · FY 2024 · 2014-09

Project Summary: Chronic obstructive pulmonary disease (COPD) is a progressive, debilitating disease for which new, disease-modifying treatments are desperately needed. Since drug targets supported by human genetic evidence are more likely to lead to FDA-approved treatments, functional characterization of genome- wide association study (GWAS) loci is a translational research priority. Our group has played a leading role in COPD GWAS, and the largest COPD GWAS to date has identified 82 significant loci, most of which have not been functionally characterized. In the first phase of this project, we combined emphysema GWAS results with expression quantitative trait locus (eQTL) studies, using GWAS-eQTL colocalization methods to identify COPD GWAS target genes. This approach allowed us to prioritize TGFB2 and ACVR1B for functional studies in airway epithelial cells and lung fibroblasts that identified functional variants in these loci. However, >50% of COPD GWAS loci have not yet shown strong colocalization with eQTLs, due in part to inherent limitations of eQTLs which depend on gene-level expression quantifications that do not reflect effects of alternative splicing. Alternative splicing is an important functional mechanism for GWAS loci, perhaps equally as important as eQTLs. In the next phase of this project, we propose to identify novel COPD-associated genetic variants that alter splicing (sQTLs) and characterize their isoform-specific effects. In Aim 1, we will perform genome-wide discovery of splicing QTLs (sQTLs) using two RNA-seq resources with large numbers of subjects with COPD – blood RNA-seq from 4,515 subjects in the COPDGene Study and lung RNA-seq from 1,078 subjects in the Lung Tissue Research Consortium (LTRC). Using colocalization methods, we will identify novel COPD GWAS target genes whose splicing is altered by COPD-associated genetic variants. In Aim 2, we will identify differentially expressed and differentially used isoforms in COPD using estimated isoform quantifications from short read lung tissue RNA-seq in 1,078 COPD cases and controls in the LTRC. We will then generate Oxford Nanopore Technologies (ONT) long read RNA-seq for 10 COPD GWAS genes in 80 LTRC subjects with COPD and 80 controls using a targeted enrichment approach. In Aim 3, we will combine fine mapping and functional studies to identify COPD GWAS variants that alter splicing in primary lung cells. First, we will use targeted long read RNA-seq to characterize the cell-type specific isoform profiles of COPD GWAS target genes in four primary lung cell types. We will then functionally validate fine- mapped COPD GWAS variants using splicing reporter assays, and we will characterize the effects of these variants on COPD-related cellular phenotypes in airway epithelial cells selected by genotype from the Marsico Lung Institute cell bank. Our multi-disciplinary research team has the requisite expertise in COPD genetics and genomics, molecular biology, long read sequencing, splicing and RNA biology to complete this important project to identify novel COPD GWAS target genes involved in alternative splicing.

NIH Research Projects · FY 2025 · 2014-04

PROJECT SUMMARY The Alliance is an experienced multi-institutional cancer clinical trials group that has been reorganized in response to the IOM consensus report and NCI directives for cooperative group reorganization. Alliance provides a comprehensive and highly efficient clinical trials infrastructure, access to experienced collaborators across all disciplines of oncology clinical trials research, and a diverse portfolio of trials for patients with breast, gastrointestinal, genitourinary, respiratory, central nervous system, hematological malignancies, and selected rare tumors. Particular strengths of the Alliance include the long-standing tradition of collaborative research of the legacy members, including many who have participated in cooperative group research since its inception in 1955. The Mission of the Alliance for Clinical Trials in Oncology is to reduce the impact of cancer by: – Conducting high quality multidisciplinary cancer control, prevention, and treatment trials that engage a comprehensive research network; – Furthering our understanding of the biological basis of the cancer process and its treatment; from discovery, to validation, to clinical practice; – Providing a scientific and operational infrastructure for innovative clinical and translational research in the academic and community settings.

NIH Research Projects · FY 2026 · 2014-03

This is a renewal application for the Harvard Summer Research Program in Kidney Medicine (HSRPKM) which provides a hands-on experience for talented undergraduate students to establish and build upon their interest in kidney and urology research. The HSRPKM was initiated and funded in 2014. Over the last 10 years we have enrolled 118 students from 67 colleges/universities in 32 states. The average GPA of the students over the past 10 years is 3.89. Our program is built around providing students with an intensive, hypothesis-driven, mentored research experience. Each student’s project is mentored by an investigator at Brigham and Women’s Hospital, Beth Israel Deaconess Medical Center, Boston Children’s Hospital or Massachusetts General Hospital. A site director at each hospital helps place students with the appropriate mentor. We have an outstanding group of 30 Program Faculty involved in research and career mentoring, retention, and assessment. In addition to their daily research, students participate in a weekly core curriculum to introduce them to the breadth of kidney biology in health and disease. This includes: 1) an introduction to the principles of renal physiology; 2) a renal gross pathology session with autopsy specimens; 3) training for and performing a community screening for kidney disease; 4) a visit to an outpatient dialysis center to appreciate the impact of kidney disease and experience this current treatment approach for patients with kidney failure; 5) an opportunity to observe the clinical work of a nephrologist or urologist; and 6) additional sessions on patient perspectives and the contributions of research in the biotech/pharma industry to new therapeutic approaches. Students present their work locally and then nationally at the NIDDK-KUH Summer Research symposium. There are close ties with the Kidney Disease Screening and Awareness Program (KDSAP). HSRPKM alumni have started 17 KDSAP chapters (52% of the current chapters) which enable longitudinal experiences with, and increased peer awareness of, kidney health and disease. A year-round program of mentorship, career development sessions, and networking helps sustain alumni interest in Nephrology and Urology. The Program Directors have assembled an Advisory Committee to oversee the program and judge its effectiveness. They have also assembled a Mentorship and Retention Faculty group to help develop and implement recruitment and retention strategies. An Assessment Committee will continue to expand evaluative approaches to optimize the trainee experience and determine the effectiveness of the HSRPKM in achieving the NIDDK-KUH Institute’s goals.

NIH Research Projects · FY 2025 · 2013-09

Project summary This is the renewal application of a SPORE initiative from Dana-Farber/Harvard Cancer Center. We focus on gliomas of children and young adults. Our objective is to develop therapies targeted to oncogenic drivers and/or oncogene-induced vulnerabilities of these tumors. Towards these ends, basic scientists join with clinician scientists from Boston Children’s Hospital, Brigham and Women’s Hospital, Dana-Farber Cancer Institute and Massachusetts General Hospital. There are three projects: Project one targets pediatric low-grade gliomas (pLGGs). Nearly 75% of pLGGs are driven by truncation/fusion variants of the BRAF protein kinase. The project one team has made significant contributions to development of tovorafenib – a brain penetrant type 2 RAF inhibitor that is effective on all common BRAF oncoproteins. For this renewal application, the goals of project one are to reduce variability in response to RAF inhibitors such as tovorafenib and develop non-invasive “child friendly” tools to predict children most likely to require (and respond to) these drugs. Project two targets diffuse midline gliomas (DMGs) – the deadliest brain tumor of children. DMGs cannot be cured by conventional therapeutic modalities and the prevalent oncogenic drivers are undruggable. During the current funding period, the project two team has identified a synthetic lethal “addiction” to the alternate end joining pathway for DNA repair (“alt-EJ”) and identified brain-penetrant drugs that specifically target this pathway. Going forward, the goal of project two is to exploit alt-EJ repair antagonists as targeted therapeutics for DMG. Project three addresses the early-stage (lower- grade) IDH mutant gliomas (hereafter termed “IDHM”) of young adults. During the current funding period, the project team identified synthetic lethal vulnerabilities for the more aggressive (WHO grade 4) IDHM gliomas. Going forward, the goal of project three is to integrate these synthetic lethal therapeutic opportunities for grade 4 IDHM glioma into “the vorasidenib era” that has been opened by the recent clinical studies on early stage (grade 2) IDHM gliomas of young adults. The study plan employs contemporary methods in structural biology, cancer genomics, computer science and machine learning. Each project has a human endpoint, and two projects will involve clinical trials. Rigor and reproducibility of work will be fostered by cores for Pathology and for Biostatistics. An Administration core will enable and manage the multiple consortium agreements and collaborative interactions Harvard Medical School, the four participating Harvard teaching hospitals and facilitate clinical trials and imaging studies. Intellectual vigor within the program is sustained and refreshed by annual Career Enhancement Awards to young investigators and by annual Developmental Project Awards.

NIH Research Projects · FY 2025 · 2013-09

Project Summary / Abstract Some exposures in ophthalmology may not have an immediate effect, but instead a lag is necessary. For example, there is a literature on possible cataractogenic effects of corticosteroid eyedrops (CS) among uveitis patients. However, the precise impact of dose and/or duration of use are unknown. Also, the lag between CS administration and development of cataract is unknown. Another possible application is to study the effects of dietary and/or supplement use on development of AMD, where a lag effect is also likely to occur. The goal of specific aim 1 is to use latency analysis methods for ophthalmological endpoints. Latency methods have been used in pharmacoepidemiology, but to our knowledge, have never been used for ophthalmologic endpoints. The AREDS study was a landmark study in the epidemiology of AMD. A byproduct of this study was the development of the AREDS severity scale which is an ordinal scale ranging from 1 for no AMD to 9+ for advanced AMD (AAMD). The usual analysis for risk factors is a time-to-event analysis based on the Cox Proportional Hazards Model, where the event is reaching grade 9+. This is a valid, but inefficient analysis. There are many eyes (about 40%) which show changes (either an increase or decrease), but which don't develop AAMD. There are risk factors which are associated with these changes, but all such changes are treated as censored data and are considered “non-events”. In Aim 2, we propose to use an ordinal regression model for changes between successive visits which would provide a more efficient use of the data. There have been previous multi-state ordinal models proposed, but separate models are fit for each possible transition and are not integrated into an overall assessment of risk for specific covariates. This has application not only for AMD, but also for other ordinal scales used for other ophthalmologic conditions, such as diabetic retinopathy. For Aim 3, we propose to continue our work on applying correlated data methods to risk prediction for endpoints such as AUC. We will specifically compare methods for estimating AUC for small samples, extensive numbers of tied prediction scores and presence of both bilateral and unilateral subjects. In addition, we will incorporate clustered data methods for estimation of NRI, which to our knowledge, has never been done before. In Aim 4, we will continue our work on translation of clustered data methods for the eye research community including (a) correlated data methods in survival analysis, (b) analysis of longitudinal binary ocular data, and (c) sample size/power calculations based on the eye as the unit of analysis.

NIH Research Projects · FY 2025 · 2013-08

Project Summary During this past R01 funding period, we have mapped causal effects within the MHC region to specific HLA-DR binding groove amino acid sites, identified >100 rheumatoid arthritis (RA) non- MHC risk alleles across the genome, and have demonstrated that these alleles are largely within CD4+ T cell regulatory elements. If we could define the genetic mechanisms underpinning RA susceptibility, then it may be possible to define therapeutic strategies to abrogate or prevent RA. Central to this is defining the key T cells involved in mediating disease susceptibility – both in terms of their unique TCR sequence features and their pathogenic cell states. Here, we hypothesize that HLA-DR risk alleles act within the thymus to favor selection of “sentinel TCRs”, and that when autoantigens are presented to sentinel TCRs, risk alleles within T cell enhancers alter T cell specific gene regulation, which enables naïve T cells expressing sentinel TCRs to transition into a pathogenic state. We define “sentinel TCRs” as those receptors that bind to citrullinated peptides and trigger the initial autoimmune response. Risk alleles in T cell promoters and enhancers alter regulation of critical T cell genes that regulate the transition of T cells into pathogenic states. Hence, a T cell with a rare “sentinel TCR” can, under the right conditions, trigger the initial autoimmune response, drive a spreading immune response, and initiate persistent joint inflammation. But, the key pathogenic T cell states, sentinel TCRs, and the action of specific causal regulatory T cell alleles are not yet fully defined. Repertoire sequencing to define TCR sequences in blood and tissue, single cell analyses to resolve T cell states, and genetic engineering to interrogate causal T cell alleles and genes represent exciting methodologies that our lab has developed expertise in. In this proposal we seek to build support for this model. First, we will demonstrate that HLA class II HLA RA risk alleles alter TCR repertoire to harbor “sentinel TCRs”, using TCR and genotype data from 300 healthy individuals. Next, we will use polygenic RA risk models to define the key T cell states that RA risk alleles influence with single cell data on surface markers and transcripts in the same 300 individuals. Finally, we will define the molecular mechanisms of non-coding alleles using genomic editing in CD4+ T cells; to this end we will develop and apply strategies to edit primary T cells, sequence DNA to confirm the presence of the desired edit, and obtain RNA and ATAC sequencing data to understand the impact of the edit and confirm the functionality of non- coding alleles.

NIH Research Projects · FY 2025 · 2013-08

Project Summary Basic and translational research in hematology has been at the cutting edge of recent advances in our understanding of the molecular pathophysiology of disease for decades. This T32 grant, for which we request renewal, was submitted 5 years ago to replace the former Harvard Medical School Training Program in Molecular Hematology, which had a distinguished 25-year track record in training graduate students and postdoctoral fellows in the study of blood and its disorders, with many graduates of the program going on to highly successful academic careers. The past 30 years have seen many changes in the landscape of hematology-oncology in the Harvard Medical area, with the merging of the Hematology program at the Brigham and Women’s Hospital with the oncology programs at the Massachusetts General Hospital and the Dana-Farber to form a combined Hematology-Oncology fellowship. This has had many positive effects on the opportunities for clinical and research training, complemented by a proliferation of training programs. At the same time, an explosion of new technologies has ushered in an era in biomedical research that offers exciting opportunities for major breakthroughs in our understanding of human disease, offering hope for new clinical paradigms that can transform the treatment of many hematologic disorders. However, the loss of a dedicated Hematology fellowship has made us acutely aware of the importance of nurturing physician scientists dedicated to the study of hematology. Indeed, the disappearance of free-standing hematology fellowships nationwide and the shrinking numbers of Hematology trainees signals an urgent need to encourage future academic hematologists. This program places high priority on training physician-scientists focused on hematology, and we have been gratified to see an increasing number of clinical fellows who choose to do research as trainees on this grant. To date, we have accommodated all the MD and MD/PhD fellows with a research focus on Hematology, although we have a selection process in place should we have to limit availability because of an increase in interested fellows. This process has already been used in the first two years of the grant to select PhD fellows to fill available slots, although all subsequent slots have been taken by physician scientists. The primary site of training is the preceptor’s laboratory, but each trainee is expected to participate in relevant seminars and courses within the HMS community. Each trainee also assembles a training committee that monitors research progress with annual formal presentations. We strongly encourage the investigation of benign hematologic disorders in the areas of red cell disorders, iron metabolism, hemostasis and thrombosis, and neutrophil disorders, as well as hematologic malignancies such as leukemia, myelodysplasia, and myeloproliferative neoplasms. This strict focus on Hematology distinguishes this grant from other Harvard-related training grants, fulfilling a need that is not answered by any other training grant within adult medicine. We request renewal of this application with its 8 current training positions.

NIH Research Projects · FY 2025 · 2013-02

OVERALL PROJECT SUMMARY Glioblastoma (GBM) and recurrent GBM (rGBM) is arguably one of the most fatal of cancers: it remains impervious to treatment with the current standard of care (SOC) therapies and numerous clinical trials (including immunotherapy) have failed to meet end points for FDA approval. This makes GBM/rGBM an unmet medical need. The unresolved problem we are trying to solve is that the rGBM tumor microenvironment (TME) is highly immunosuppressive. To revert this immunosuppression, we and others have been utilizing direct intra- tumoral administration of oncolytic viruses (OVs), based on herpes simplex virus type 1 (HSV1). During the current funding cycle (2018-2023), the Principal Investigators of this P01 have contributed several discoveries both in preclinical models of GBM and in a 50 subject phase 1 clinical trial of recurrent GBM (rGBM). Major findings relate to how human subject rGBMs change in response to oHSV (Project 2/ Chiocca), how Notch signaling of myeloid cells in the GBM TME impedes oHSV function (Project 3/Kaur) and how to further immune- stimulate oHSVs by “arming” with immune-stimulatory genes (Project 1/Glorioso and Project 4/Caligiuri). Lessons learned informs the overall hypothesis for this P01: oHSVs induce a rapid and persistently inflamed rGBM microenvironment, correlating with efficacy in human subjects and informing further therapeutic exploitation by arming oHSVs with additional immunostimulatory genes. To test this overall hypothesis, the four Projects will address the following overall aims: Overall Aim 1 – Validate the findings that improved survival for human subjects treated with oHSV correlates with rGBM and peripheral blood biologic transcriptomic and proteomic signatures (Projects 2, 3 and 4); and Overall Aim 2- Engineer and preclinically test the “next-generation” oHSVs “armed” with selected immune-activating genes, focusing on clinical translation for a new phase 1 trial (Projects 1, 3, and 4) To support the efforts of these 4 Projects, we will continue to utilize the services of the oHSV Production Core (Core 1/ Goins), of the GBM Biorepository Core (Core 2/Ligon) and of the Biostatistics and Bioinformatics Core (Core 3/Mo & Zhang). The proposed timeline for the successful completion of the overall aims is 5 years. The principal investigators of this P01 espouse the principle of “from the lab to the clinic and back to the lab”, where the lessons from our treated patients can be applied to build more efficacious therapies for this fatal cancer. The potential impact of this research is that we are proposing to not only test preclinically and clinically potentially impactful oncolytic immunotherapies for an incurable cancer, but we have shown commitment to analyze the biologic and immunologic effects in treated human subjects, so that we can iteratively learn possible pitfalls and devise solutions for patients with GBM.

NIH Research Projects · FY 2024 · 2012-08

Endothelial cell (EC) activation and dysfunction increases with age and is linked to a variety of chronic vascular inflammatory disease states including atherosclerosis—the major cause of morbidity and mortality in Western Societies. Vascular senescence induced by the DNA damage response (DDR) promotes chronic inflammation in atherosclerotic lesions. Senescence-associated proinflammatory cytokines and proatherogenic risk factors (acquired or inherited) such as hyperlipidemia activate key signaling pathways that increase expression of adhesion molecules, chemokines on several cell types, including the vascular endothelium. Therefore, suppressing the senescence-associated inflammatory response in the vascular endothelium may provide a novel therapeutic approach to limit atherosclerosis. MicroRNAs (miRNAs) are small, single-stranded, evolutionary conserved non-coding RNAs that suppress the expression of target genes at the post-transcriptional level and participates in a variety of pathophysiological processes including the regulation of inflammatory responses. Our group provided the initial link implicating miR-181b in suppressing endothelial cell inflammation. During the last grant period, we identified miR-181b as a nodal regulator of endothelial cell quiescence through its regulatory effects on two major signaling pathways – NF-κB and AKT/eNOS. Consequently, endogenous miR-181b was found to function as a key determinant of the inflammatory response in vivo, findings that correlate with human inflammatory states including established coronary artery disease. We now identify endothelial miR-181b as a critical determinant of systemic vascular inflammation and atherosclerosis by controlling vascular senescence and the DNA damage response. Furthermore, we demonstrate that the adenosine-A3AR signaling pathway, a translationally relevant target that suppresses endothelial activation, functions in a miR-181b-dependent manner. These observations provide the foundation for the central hypothesis that endothelial miR-181b, via inhibitory effects on the DNA damage response, regulates senescence-associated vascular inflammation and atherosclerosis. To better understand the precise role of miR-181b in regulating vascular senescence, inflammation, and atherosclerosis, we propose 3 aims. In Aim1, we will determine the molecular basis for miR- 181b to regulate the DNA damage response and vascular senescence in response to diverse stimuli. In Aim2, we will explore the effect of altering miR-181b expression on senescence-associated secretory phenotype and atherosclerotic progression and regression in young and aged mice. In Aim3, we will determine whether the anti-senescent effects of adenosine in the vascular endothelium depend on miR-181b. This multi-disciplinary team in the fields of non-coding RNA biology, molecular imaging, nanomedicine, bioinformatics, and atherosclerosis research will establish an unprecedented molecular view of this miRNA in lesions that can inform a new frontier in the regulation of vascular senescence and atherosclerosis.

NIH Research Projects · FY 2026 · 2011-07

Summary/Abstract This Asthma and Allergic Disease Cooperative Research Center (AADCRC) grant continues to focus on the mechanistic basis of respiratory tract type 2 immunopathology (T2I), particularly in aspirin-exacerbated respiratory disease (AERD), a distinctive clinical syndrome that accounts for a disproportionate percentage of individuals with severe asthma and recurrent chronic rhinosinusitis with nasal polyposis (CRSwNP). In the current period of support, we identified novel and heterogeneous populations of mast cells (MCs) in nasal polyps, and determined that canonical MC subsets have very different effector functions. We found that transforming growth factor (TGF)-β is essential to drive the differentiation of proliferating MC progenitors (MCp) toward a tryptase+chymase- (MCT) phenotype that accumulates in the epithelial surface in aspirin tolerant (AT)-CRSwNP and especially in AERD. We determined that PGE2 drives an anti-inflammatory function of MCT that is due to their production of soluble ST2, a decoy receptor for IL-33, whereas epithelial PGE2 insufficiency in AERD permits aberrant expansion and activation of MCT that contributes to disease severity. We determined that epithelial basal cells (BCs) in nasal polyps show strong activation of mammalian target of rapamycin (mTOR), which contributes to epithelial-mesenchymal transformation (EMT), alarmin generation, and failure to repair the epithelial layer, as well as impaired PGE2 generation. Moreover, activated MCs and macrophages generate many potential drivers of epithelial mTOR signaling. Thus, feed-forward loops between structural cells and innate immune effector cells may underly the pathogenesis, persistence, and recurrence of AT- CRSwNP and AERD. A tightly interactive team of accomplished investigators with complementary skills will apply cellular, molecular, and whole animal strategies to determine the mechanistic basis for these findings, their relevance to disease pathophysiology, and their amenability to therapy. Project 1 (J. Boyce, PI) focuses on identifying the drivers of MC proliferation and activation, their relationship to disease severity, and the molecular mechanisms accounting for MC diversification by epithelial TGF-β and PGE2. Project 2 (N. Barrett, PI) focuses on the mechanisms by which mTOR signaling drives BC dysfunction, identifying the products of BCs that impact effector cell recruitment and activation, and identifying the principal drivers of mTOR originating from activated MCs and macrophages. The Projects are supported by respective Cores for Adminstration (Core A), Data Stewardship (Core B), and Patient Phenotyping and Biorepository (Core C).

NIH Research Projects · FY 2024 · 2010-08

Project Abstract The Greater Boston MSTAR Program aims to provide a short-term aging research and mentoring experience to medical students early in their career. Students will become educated and enthusiastic about the career opportunities in academic, clinical, and research geriatric medicine. The Program has operated continuously since 1989 and provided short term research training experiences to over 230 medical students. Both the Program Director and Co-Director have five years of experience in this role and have been involved with teaching and mentoring Boston MSTAR students for over a decade. The Program Director will report to the Mentor Advisory Council for program oversite, increasing the pool of high-quality mentors, and continued efforts on recruiting minority Scholars from Harvard, the University of Mississippi/Jackson Heart Study, Boston University Medical Center, the University of Massachusetts Medical School, and Brown University School of Medicine. We currently have over 40 mentors with more than 200 funded research projects whose combined direct costs for the current year total over $60 million. The heart of the Boston MSTAR Program will be the 8 to12-week mentored aging research project experience for 11 scholars: 7 from medical schools across the country, and one each from the University of Mississippi, University of Massachusetts, Boston University Medical Center, and Brown University. All students will receive training in protection of human subjects, aging research methodology, and clinical geriatrics. Students will present their research in an oral presentation and will be strongly encouraged to present their work at the annual meeting of the American Geriatrics Society. The program will track students through their medical training and career decisions. By developing a strong relationship with the students, the scholar tracking will be a continuation of the mentoring experience and a forum to increase scholar interest in aging research. The program’s past successes are a foreshadowing of its future potential. Over the past grant cycle, we have tripled the number of applicants, allowing us to select students with a genuine interest in aging research. Over two-thirds of our students present at a scientific meeting 20% publish a scientific paper from their 8-12 week mentored research experience. Long term, under the T35 mechanism, 24% of matriculated students have pursued additional research training or a career in geriatrics. As a result, the Greater Boston MSTAR Program seeks to continue the tradition of training medical students in aging research.

NIH Research Projects · FY 2025 · 2010-07

Cardiovascular (CV) disease is the leading cause of morbidity and mortality in the US and worldwide. Non-invasive imaging provides valuable information to assess differential diagnosis and guide management. However, there is a gap in academically-oriented CV imaging training programs that are able to train specialists with the knowledge base, experience, research tools, analytical and leadership skills to fully develop, apply, and evaluate the increasingly complex gamut of CV imaging possibilities. This is the third competing renewal application for this T32 program. This program offers post-doctoral research training for individuals with an MD, or combined MD/PhD degrees, who have completed clinical training in cardiology, radiology, or CV imaging who are committed to pursuing an academic career in CV imaging science. Our goal is to continue to provide outstanding opportunities for multi- disciplinary research training for clinician-scientists in CV imaging with the clinical and investigative skills to establish independent careers, mentor others, and lead their own programs. Since initial funding in 2010, we have graduated 32 post-doctoral fellows. Key indicators of the effectiveness of this program include: (1) 24/32 graduates remain involved in academic medicine and science, and 1 is completing additional clinical training; (2) 16/32 fellows who completed the program have received research funding including 13 with K awards or similar (AHA), 5 with both K and AHA awards, and 4 received ACC mentor award. 5/32 graduates have received R01 funding; (3) the grant has sponsored 188 unique publications in top peer review journals; (4) 50% of our graduates hold leadership roles in their respective imaging programs, and 38% have prominent leadership roles in professional organizations and others; (5) 40% of all 38 trainees in the program are women and 16% of Hispanic or Latino. Our mentors are primarily comprised of cardiologists and radiologists, but also include faculty from disciplines outside CV imaging including clinical and basic scientists. All the mentors have a strong track record of research collaborations. We offer research training opportunities in 9 thematic areas: (1) molecular imaging, (2) myocardial structure, function, and metabolism, (3) genetics, (4) pulmonary hypertension, (5) ischemic heart disease, (6) outcomes research, (7) women’s health, (8) cardiometabolic medicine, (9) radiomics and artificial intelligence. The proposed program supports 3 positions/year for 2 years of continued research training. With this renewal, we continue our primary goals of providing leadership in academic CV imaging and training future leaders in imaging science to improve outcomes in CV disease.