Brigham And Women'S Hospital

NIH Research Projects · FY 2026 · 2024-01

Project Summary/Abstract Non-Hodgkin lymphoma (NHL) and multiple myeloma (MM) are cancers arising from mature lymphocytes that collectively pose a substantial public health, personal and financial burden. Present knowledge of their etiology is inadequate to inform prevention strategies; data from prospective studies, and for specific histologic subtypes, are particularly limited. Inflammation, heightened immune activation and growth factor dysregulation contribute to their pathogenesis, as indicated, for example, by well-known associations of NHL risk with severe immune compromise and with pre-diagnosis plasma immune marker profiles that indicate inflammation and B- cell activation, as well as by consistent findings of an increased risk of MM and some NHL subtypes in obese individuals and in persons with autoimmune disease. Relevant to the present application, evidence from both model organism and human occupational studies support the hypothesis that environmental pollutants with carcinogenic and/or immunotoxic effects—such as dioxins and hazardous air pollutants (HAPs)—may be risk factors for MM and/or NHL. However, those studies have many limitations, and most do not assess general population risk from low-dose, passive and possibly chronic exposure to those pollutants or to combinations of multiple pollutants. Geographic information system (GIS)-based modeling of these exposures offers a valid and well accepted tool for examining environmental risk factors for MM, NHL and common histologic subtypes of NHL and could yield novel insights into their etiopathogenesis. The prospective studies proposed in this application will leverage strong investigator expertise and an active collaboration and will include eight large and diverse cohorts, including geocoded residential histories spanning many decades and a projected sample size of ~5,386 incident MM and ~20,354 incident NHL cases. Cox proportional hazard models will be used to examine GIS-modeled passive exposure to dioxins and HAPs (individual and mixtures of pollutants, the latter using quantile G-computation and Bayesian approaches) with known or plausible carcinogenic or immunotoxic properties in relation to risk of MM, NHL, major histologic subtypes and clinically relevant molecular subtypes of diffuse large B-cell lymphoma (DLBCL), an aggressive histologic type of NHL with unknown etiologies. The analyses will incorporate already-harmonized individual-level time-varying risk factor data to assess and control confounding by lifestyle, occupation, demographic and medical history- related risk factors. Additional analyses will assess timing of susceptibility and heterogeneity of observed associations by NHL histologic subtype, calendar period, sex, age, race/ethnicity, body mass index, smoking and socio-demographic and neighborhood contextual factors. The proposed studies have strong potential to yield novel insights on the contribution of passive, low-level exposure to environmental factors to the etiology of MM and NHL, including on risk to individual histologic subtypes of NHL and molecular subtypes of DLBCL, and thus to provide urgently needed identification of novel risk factors for these understudied cancers.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY/ABSTRACT Autism spectrum and related neurodevelopmental disorders are thought to arise from widespread mild synaptic dysfunction that leads to an altered trajectory of brain network development (Johnson et al., 2015). In Rett syndrome (RTT), a debilitating neurodevelopmental disorder, loss-of-function mutations in MECP2 on the X chromosome lead to a devastating loss of language, motor and visual function at the behavioral level. Yet no therapy exists to stop or reverse the cognitive decline. Previous work in RTT mouse models reveal synaptic- and network-level defects that precede symptom onset. Mecp2-deficiency alters the timing of excitatory synaptic maturation in excitatory and inhibitory neurons in the developing cortex (Mierau et al, 2016). Microelectrode array (MEA) recordings and calcium imaging of neuronal activity in cultured murine cortical neurons reveal delays in the development of functional connectivity in the Mecp2-deficient cortical networks including smaller network size, weaker strength of connectivity and impairments in the development of network topology including features that predict local and global efficiency of the networks (Dunn et al, unpublished). These findings reveal cellular- scale deficits in information processing that likely underlie the cognitive impairment in RTT and could be targeted with novel therapies to rescue cognitive function. In my proposed K02 and subsequent R01 research, we will translate this approach to a human in vitro cellular model of RTT. In Aim 1, air-liquid interface cortical organoids (ALI-COs) will be generated from RTT patient-derived induced pluripotent stem cells (iPSCs) and allogenic controls to first identify network-level defects in the development of cortical networks. The ALI-CO model shows further cortical maturation than previous organoid models and avoids the necrotic core (Giandomenico et al., 2019). In Aim 2, we will target excitatory synaptic maturation in a specific type of inhibitory neuron in the MeCP2- deficient ALI-COs using novel cell-type specific modulators we identified (Mierau et al, unpublished). Our goal is to prevent early synaptic defects and ensure the formation of network activity and topology necessary to support cellular-scale information processing. This strategy could be used in our future R01 research to prevent the cognitive decline in RTT. In Aim 3, we will develop novel tools for detecting and modulating network dynamics in the MeCP2-deficient ALI-COs as a platform for testing strategies to rescue network function in RTT after the cognitive decline has occurred. Re-expressing Mecp2 in adult Mecp2-deficient mice rescues many behavioral and synaptic features, raising the hope that neuronal function can be rescued in RTT even after the cognitive decline. We will next test dynamic electric, optogenetic, and pharmacologic approaches for modulating cognitive function. This will support the subsequent R01 aims in Year 4 and 5 of the K02 award to test whether modulating network function in real-time can rescue network function in RTT ALI-COs. This strategy would facilitate screening of novel therapies to unlock function in symptomatic RTT patients and could be applied in the future to develop new therapies for autism spectrum and other cognitive disorders.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY / ABSTRACT Alzheimer’s disease (AD), the most common cause of dementia, is accompanied by progressive neurodegeneration and stereotypic deposition of misfolded amyloid β- (Aβ) and tau proteins. However, the basis for cellular dysfunction in AD is not well understood, underscoring an important need to broaden the biologic scope of AD research. Technological advances, based on single-cell whole-genome sequencing (scWGS) on frozen postmortem human brain tissue, now allow examination of previously inaccessible aspects of the genome of individual cells in the brain. Recent research from our group has found that neurons in individuals with AD show greater somatic single-nucleotide variants (sSNV) in their genomes than in individuals without AD. Using mutational signature analysis, our study also found that the specific causes of somatic mutations differed in AD, implicating oxidative stress and signal suggesting other cytopathologic events. This study aims to understand neuronal somatic mutation in AD at a deeper level, to probe how disease progression and protein misfolding impact the burden and signatures of sSNV. The first aim will examine neurons from individuals with different pathologic stages of AD. This will give us a better understanding of when somatic mutations develop in disease, and whether certain mutagenic events occur with distinct timing in the development of AD. The second aim will study how intracellular tau pathology relates to somatic mutation, utilizing a novel method to separate individual nuclei according to tau tangle formation. In applying this method to specific brain regions at specific pathologic stages at the forefront of AD progression, we will be able to determine if somatic mutation is tied to tau pathology at the level of individual cells. In the final aim, we will test whether Aβ trigger somatic mutation, using a human iPSC-derived neuronal cellular model and incubation with human brain-derived Aβ oligomers. With complementary approaches of novel scWGS technology, single-nucleus sorting, and specific human tissues and and cellular systems, we will examine the mechanistic role of neuronal somatic mutations in AD. This examination of the interplay between disease progression, protein misfolding, and the landscape of somatic mutations will be an important step in understanding the molecular causes of AD. Moreover, the capability of somatic mutational signature analysis to reveal previously unrecognized disease mechanisms lends additional promise toward development of better treatments and cures.

NIH Research Projects · FY 2026 · 2024-01

Lung diseases exhibit differences between males and females in risk, progression, symptoms, and response to therapy; rigorous investigation is needed to define the molecular reasons for these differences. Our previous work has demonstrated that modeling Omics data using gene regulatory networks can discover key differences driving health and disease. The goal of this K24 is to merge our prior approach to modeling sex differences with mentoring researchers in data science. Methods will support modelling sex as a biologic variable using a data science framework, to promote translation of data from the NHLBI Trans-Omics for Precision Medicine(TOPMed) program to actionable clinical innovations for COPD. The goal of this Patient Oriented Research program is to mentor trainees in Omics data science and Network Medicine through a lens considering sex as a biological variable; to date this approach has been limited for lung diseases. To advance the dissemination of data analytic approaches for COPD research, we will focus on epidemiology, Omic analyses inclusive of X and Y chromosome data, and gene regulatory network methods. In addition to mentoring and career development, trainees will pursue the hypothesis that somatic mosaicism on the X chromosome (mCA X) and loss of Y (LoY) will associate with COPD phenotypes and COPD progression. To facilitate mentoring, scientific Aims include 1) Investigating whether mCA X and LoY mediate sex divergent features of COPD and 2) Defining pathway and gene regulatory network rewiring associated with mCA X and LoY, using single Omic and network-based approaches. Innovative aspects include the evaluation of sex associations with COPD; investigation of mCA X and LoY and COPD; extension of gene regulatory network approaches to include somatic mosaicism; and development of a sex-aware data science toolkit for inclusion of X and Y chromosome data. Understanding sex differences using tools to integrate Omics and the gene regulatory landscapes with mCA X and LoY will improve our understanding of COPD and support precision medicine initiatives. This paradigm shifting K24 will support the training of the next generation of pulmonary data scientists interested in Patient Oriented Research, with specific application to COPD, a leading cause of death in the United States.

NIH Research Projects · FY 2025 · 2023-12

Abstract Cardiovascular diseases (CVDs) are a group of disorders of the heart and blood vessels, and represent the leading cause of death worldwide, taking more lives than all forms of cancer combined. An estimated ~18 million people die from CVDs annually, representing 31% of all global deaths, making it a leading cause of death for both men and women. Clinical studies have revealed significant disparities between women and men in the incidence of, and mortality from, CVDs. CVDs develops 7 to 10 years later in women than in men but is still the major cause of death in women. Since 1984, overall mortality from CVDs has remained higher in women than men. For far too long, cardiovascular disease was considered a “man’s illness”, and women have been under-represented for years as subjects in all phases of cardiovascular research, from basic science to clinical application. Because men and women are biologically and physiologically distinct, often suffer differently from the same diseases, and respond differently to the drugs, there is an urgent need for improvement in the current preventive, diagnostic, and treatment strategies by considering sex-specific differences in the etiology and risk factors of CVDs. As an example, recent advances in organ-on-a-chip platforms have enabled the recapitulation of both physiological and pathological conditions of complex tissues and organs in vitro, including those of the heart. However, none of these models ever truly considered sex as an important biological variable. The goal of this bold new methods R21 project is to reveal sex differences in an engineered microfluidic cardiac hypertrophy-on-a-chip platform, populated with sex-specific cardiac cells. It is hypothesized that there exist critical differences between sexes in the development of biomechanically induced and/or biochemically induced cardiac hypertrophy in the biomimetic on-chip platform. Sex as an important biological variable in heart-on-a-chip systems has not been investigated so far and we envision our high-risk, high-reward project to lay down the basis for future in-depth investigations on this indispensable but highly underrepresented parameter.

NIH Research Projects · FY 2026 · 2023-12

Project Summary/Abstract The cytokines IL-4, IL-5, and IL-13 are thought to be key effectors of airway inflammation, but remarkably little is known about either their global function within the respiratory tract or the potential consequences of their complete inhibition by newly available biologic drugs. The overall goal of the proposed research is to define the underlying human biology of IL-4Rα and IL-5R signaling in airway inflammation, both their role in driving type 2 inflammation and their role in counterregulating other systemic immune pathways. We will determine the cell-specific impacts of these cytokines within the respiratory tract and confirm the influence of IL-4Rα and IL-5R on the chronic inflammation of patients with chronic rhinosinusitis with nasal polyposis (CRSwNP), severe asthma, and aspirin-exacerbated respiratory disease (AERD). Further, given that some drugs now available to inhibit these pathways have poorly understood side effects like new-onset joint pain, we will establish the systemic immunologic consequences of inhibiting a receptor as widely expressed as IL-4Rα and will identify novel biomarkers to predict those immune responses. In three separate but interrelated projects, we aim to: 1. Define the cellular mechanism(s) of anti-IL-4Rα-induced therapeutic benefit in AERD with a prospective study of 8 weeks of dupilumab treatment. 2. Define the systemic immunologic consequences of IL-4Rα inhibition with dupilumab and determine which immunologic changes correlate with the development of arthralgias. 3. Characterize the functional significance of IL-5Rα on upper airway nasal plasma cells and epithelial cells, from patients with healthy sinus mucosa, CRSwNP, and AERD. These goals will be achieved using patient-oriented research (POR), as each of these aims involve patient interactions and longitudinal study visits to collect and verify clinical outcomes, and through the mentorship of the next generation of patient-oriented allergic disease investigators. The proposed studies and the clinical questions to be examined offer outstanding training opportunities for junior investigators interested in clinically relevant research in allergy and asthma. Dr. Tanya Laidlaw is a recognized investigator in the causes of CRSwNP and AERD and the candidate's many ongoing POR projects and collaborations, commitment to and track record of mentorship, along with the exceptional institutional resources, provide a first-rate environment for the development of junior investigators. Critically, this award will provide Dr. Laidlaw protected time for POR in airway inflammation and allergic respiratory disease to both refine her mentoring and leadership skills and expand her expertise to include predictive biomarker identification, pathway-level multi-omics analysis, and high dimensional data analyses.

NIH Research Projects · FY 2026 · 2023-12

Respiratory viral pathogens have been responsible for major pandemics over the past two centuries with extraordinary human and economic cost. While most vaccines have focused on antibody neutralization of virus, there has been growing interest in targeting CD8 T cell immunity for more durable protective immune memory. Specifically, there has been interest in vaccines that generate tissue resident memory T cells (TRM), but one issue complicating this approach is the observed lack of durability of lung TRM, whether generated by pulmonary viral infection or targeted vaccination. Unlike TRM in other tissues, which accumulate and survive for years, lung TRM have half-lives measured in days. We have discovered an alternative population of CD8 TM that can mediate lung-focused T cell memory and protection from morbidity and mortality after skin vaccination. The durability of this immunity appears to be provided by a unique population of lung tropic CD8 T cells concentrated in the intravascular space of the lung (rather than the parenchyma). Using IAV as a model viral respiratory pathogen, these CD8 T cells appear to rapidly extravasate and enter lung parenchyma within hours after infection, providing a rapid on demand protective memory response with kinetics comparable to lung TRM. Their activity does not require CD4 T cells, and elimination of these CD8 TM cells completely abrogates robust vaccine protection. Analysis of these cells by scRNAseq reveals that they have circulatory markers, but also immediate-early genes reminiscent of skin TRM generated from the same vaccination. They are transcriptionally distinct from spleen TEM and LN TCM generated by the same vaccination and appear to be robust >6 months later. Most importantly, they provide complete protection from morbidity and mortality from viral lung challenge and function independently of antibodies and B cells. Using defined genetic models, scRNA seq, ATACseq, multidimensional flow cytometry, and lung intravital microscopy, we will test key hypotheses about the mode of generation, in vivo biological behavior, and protective immunity of these cells. We have assembled expert collaborators in each of these areas to fully explore the biology of these cells and the potential of generating these cells by epidermal disruption (ed) vaccination. We believe that these TM cells are an important and unstudied component of normal T cell memory designed to provide superior and long-lasting lung immunity without occupying valuable lung parenchymal space. We believe that optimized generation of these cells by improved vaccination approaches will provide a superior way to combat respiratory viral pathogens.

NIH Research Projects · FY 2026 · 2023-12

PROJECT ABSTRACT Each year, more than a million US women experience adverse pregnancy outcomes (APOs) associated with a two-fold higher risk of future maternal CVD. These APOs include preeclampsia, gestational hypertension, gestational diabetes, preterm delivery (<37 weeks), and delivery of a small for gestational age infant. The American Heart Association and American College of Cardiology recognize APOs as risk factors for CVD. However, it is not yet known how to identify which women with a history of an APO are most at risk; nor are specific, modifiable CVD risk factor trajectories from each APO to CVD well described. To date, risk prediction exercises have only examined the utility of adding individual APOs to existing CVD risk scores. No risk prediction exercises have addressed the clinically relevant question of which women with APOs are most at risk for CVD, and no clinical decision support tool exists to direct care after an APO. We propose to develop a risk classification approach to identify maternal risk phenotypes at the time of delivery of a pregnancy complicated by an APO that are predictive of future CVD risk factors and events. Predictive factors that inform these phenotypes may include history of individual APOs or combinations of APOs, delivery complications, infant outcomes, and maternal demographic or behavioral factors available at or shortly after delivery (e.g., preterm preeclampsia with BMI <30 kg/m2). We will use machine learning to elucidate novel maternal CVD risk phenotypes in several populations: two longitudinal Nurses’ Health Study cohorts and a retrospective cohort using electronic health record data from the Duke University Health System and University of North Carolina Healthcare System. A national board of clinical experts will advise the development of a clinical decision support tool to be tested using mixed methods among panels of obstetricians, internists, and cardiologists. Building on the existing evidence connecting APOs to CVD and leveraging the knowledge that CVD prevention should begin early, this work will create a tailored tool to help clinicians guide the millions of women with an APO history to appropriate screening, prevention, and referral to reduce their elevated risk of future CVD.

NIH Research Projects · FY 2026 · 2023-12

Abstract It is now widely appreciated that uncontrolled inflammation is a unifying component in many widespread diseases, including chronic lung disease. Inhalational exposure to respirable particulate matter can be an important precipitant or exacerbant of lung inflammation. From our earlier results, the resolution of inflammation is known today as an active process. There are several new families of specialized pro-resolving mediators (SPMs) identified and characterized in acute inflammation. These protective mediators are enzymatically produced and are agonists at specific receptors transducing cell type specific functional responses critical in tissue resolution. Resolution programs of the inflammatory response are essentially uncharted scientific terrain in environmental health and medicine. Fundamental information is critically needed on the impact of environmental agents within the resolution response and whether they perturb resolution to trigger chronic inflammatory responses and susceptibility to allergen induced asthma. Here, the laboratories of Bruce Levy, Charles Serhan, Phil Demokritou, and Adam Haber collaborate and propose an innovative proposal focusing on elucidating the impact of inhaled potentially hazardous traffic emitted ultrafine particles (UFPs) on the new resolution programs that govern pro-resolving cellular and molecular mechanisms and tissue catabasis in its return to homeostasis. Failed resolution or its disruption can lead to sustained lung inflammation and enhanced susceptibility to allergen induced asthma. This proposal will test an innovative hypothesis, namely that exposure to traffic-emitted UFPs disrupts airway resolution mechanisms associated with allergic airway responses in part via changes in specialized pro-resolving mediator production and action; and that delivering SPMs using a novel nanocarrier platform can protect from UFP-initiated disruption of endogenous resolution mechanisms. To test this hypothesis, an interdisciplinary team of scientists was assembled with expertise in environmental health and particle health effect studies (Demokritou), computational biology (Haber), asthma, lung inflammation and its resolution (Levy) and SPMs, SPM receptors and resolution (Serhan) to pursue the following specific aims: Aim 1. Collect and physicochemically characterize traffic emitted UFPs in urban Boston, Aim 2. Determine the impact of traffic emitted UFPs on the resolution of house dust mite (HDM) evoked allergic airway responses, and Aim 3. Determine the cellular and molecular mechanisms for UFP disrupted resolution of inflammation and establish an SPM delivery system to activate catabasis. These innovative and timely studies will establish the impact of environmental UFPs on endogenous resolution programs in lung inflammation and lay the groundwork for new therapeutic approaches with resolution agonists that promote active resolution of inflammation in our fight against environmental insults.

NIH Research Projects · FY 2026 · 2023-12

Asthma is the most common chronic disease among children globally. Disruptions of fetal development processes are hypothesized to give rise to asthma, therefore, prenatal interventions may help reduce its short- and long-term burden. Randomized clinical trials, including our Vitamin D Antenatal Asthma Reduction Trial (VDAART), have suggested that maternal vitamin D supplementation can help prevent asthma in the offspring. However, in these trials, not all children benefited from maternal vitamin D supplementation, and not all treated mothers had sufficient vitamin D levels. This variability in vitamin D response suggests complex mechanisms underlying the potential protective effect of maternal vitamin D on childhood asthma that are currently not well-understood. Our long-term goal is to uncover the molecular mechanisms that can enable the precision prevention of childhood asthma. The objective of this application is to identify the multi-omic determinants that modulate the influence of maternal vitamin D supplementation on childhood asthma outcomes. Our central hypothesis is that the effect of maternal vitamin D intake on offspring asthma status is modified by the genotypes of and the epigenetic modifications on the major regulators of vitamin D metabolism and signaling. In Aim 1, we will determine the joint role of vitamin D binding protein (DBP) levels and vitamin D metabolism and signaling genotypes in modulating the effect of maternal vitamin D supplementation in childhood asthma outcomes. We will measure DBP levels in the VDAART cohort and use the existing VDAART genotyping data. We will use Mixed Graphical Models and Conditional Gaussian Bayesian Networks to model the contribution of DBP levels, 25-hydroxyvitamin D (25-OHD) levels and vitamin D genotypes to the effect of maternal vitamin D intake on childhood asthma, and create polygenic and polyexposure scores for individual risk prediction. In Aim 2, we will identify the longitudinal epigenetic markers that modulate the effect of maternal vitamin D on childhood asthma. We will measure DNA methylation on VDAART mothers pre- and post-vitamin D treatment and use existing cord blood methylation data. We will perform epigenome-wide association studies, integrate their results with ChIP-seq and ATAC-seq data, and perform mediation analyses to identify the longitudinal maternal epigenetic marks that potentially mediate the effect of maternal vitamin D intake in childhood asthma. In Aim 3, we will determine the pharmacogenomic drivers of individual differences in maternal vitamin D response and its effect on childhood asthma. We will integrate genomics, transcriptomics and epigenomics from VDAART to identify the vitamin D-responsive pharmacogenomic expression and methylation quantitative trait loci (PGx-eQTLs and PGx-mQTLs). We will develop a network-based statistical method to identify the colocalization of these PGx-QTLs, perform colocalization using orthogonal approaches, and identify endotypes through which maternal vitamin D targets childhood asthma. Within each of the above aims, we will leverage the strong non-white representation among VDAART participants to address health disparities in vitamin D response and asthma outcomes across the population. Our research plan is innovative in its use of highly granular multi-omic data and cutting-edge integrative methods, and the outcomes of each aim are of high translational potential.

NIH Research Projects · FY 2026 · 2023-12

Project Summary Pulmonary hypertension (PH) is a heterogeneous condition that occurs most frequently as complicating comorbidity of very common disease in the western world, including COPD, heart failure, and blood clots in the lung. In patients with PH secondary to COPD (Group III) mortality risk is about twice as high than in patients with COPD and normal pulmonary artery pressure. Medications developed successfully for Group I disease have not shown much utility in Group III PH. Additionally, non-invasive screening and early detection remain challenging. For this reason, etiology and evolution of PH are still poorly understood and the response to potential interventions is very difficult to measure. In recent years, radiologists have been making observation of the loss of distal vasculature (pruning), increased vascular “tortuosity”, and proximal vascular dilation in patients with PH. In this career development proposal, we hypothesize that the great power and recent advances in artificial intelligence can be leveraged to identify new functional and structural markers that define the presence of PH in smokers from non-contrast CT images. These CT-based features will significantly improve our ability to define the main structural and functional effects of PH in COPD patients and assess pathological conditions, leading to a better understanding of the genetic and biological underpinnings of the disease. Aim 1 of this application involves refining our algorithm for vascular morphology assessment and applying it to investigate how the pulmonary vasculature is affected in presence of PH Group III. Five markers of presence of disease will be explored. The goal of Aim 2 is to develop an innovative approach to assess the level of pulmonary perfusion from non-contrast non cardiac gated CT images and to apply it to explore new functional markers of disease. Both in Aims 1 and 2 association with biological outcomes will be performed. Finally, in Aim 3 we will validate the biological relevance of the endpoints determined in Aims 1 and 2 by performing common and rare variant association studies. Preliminary data obtained with our published methods show promising results. We show that using artificial intelligence we can effectively segment pulmonary vessels, separate arteries and veins, and measure small vessels with an accuracy which outperforms state-of-the-art methods. Additionally, we developed an innovative technique that synthesized perfusion maps from non-contrast CT to assess perfusion defects in PH patients. Together, the research proposed in the aims of this award will take full advantage of the comprehensive dataset available through the COPDGene study. The execution of the aims in this proposal will be possible through active collaboration with Dr. Raul San Jose Estepar, Ph.D. as the mentor and an outstanding Advisory Committee including renowned leaders in the fields of medical image analysis, translational research, quantitative imaging in pulmonary vascular disease, and the genetic epidemiology of COPD.

NIH Research Projects · FY 2026 · 2023-12

Project Summary/Abstract Respiratory syncytial virus (RSV) is the major cause of bronchiolitis and pneumonia in young children and the leading cause for hospitalization of infants. A subset of RSV-infected children develops severe protracted lung inflammation that leads to respiratory failure and even death but effective therapies for RSV are lacking. Our preliminary data for this proposal implicates a role for NK cells in the pathobiology of RSV infection. We have determined that NK cells are abundant in the airways of RSV-infected children with profound hypoxemia and that NK cell cytotoxicity is impaired in pediatric RSV. NK cells are pivotal mediators of innate viral immunity and are also key cellular effectors of inflammation resolution to clear inflamed tissues of virus-infected and activated cells through immune synapse formation and lytic granule delivery to trigger targeted apoptosis. Disrupted inflammation resolution can perpetuate tissue damage and chronic inflammation; hence, NK cell resolution function is critical to prevent protracted lung inflammation. Resolution of inflammation is an active process mediated in part by specialized pro-resolving mediators (SPMs), a family of endogenous fatty acid- derived molecules that counter inflammatory signals and accelerate resolution through cell-specific actions. We recently identified that SPMs mitigate RSV-induced lung inflammation in mice, but whether RSV impacts SPM resolution pathways in humans remains unknown. In preliminary data for this proposal we identify for the first time that NK cells in human RSV express receptors for the SPMs resolvin D2, resolvin E1, and lipoxin A4. We have determined that the SPM lipoxin A4 up-regulates NK cell cytoskeletal proteins to promote lytic granule trafficking and augment cytotoxicity, distinct from steroids that disrupt NK cell cytoskeletal dynamics and impair cytotoxicity. This proposal will test our central hypothesis that RSV infection alters airway and circulating NK cell immunophenotype and disrupts cytoskeletal signaling pathways to impede NK cell resolution functions that can be restored by exogenous SPMs. To test this hypothesis in pediatric RSV+ ICU patients, we will pursue the following specific aims: Aim 1: Determine NK cell immunophenotypes and effector functions in pediatric RSV infection; Aim 2: Define mechanisms for disrupted lytic granule trafficking that governs NK cell cytotoxicity; and Aim 3: Establish circuits of defective inflammation resolution linked with pediatric RSV infection. This proposal brings an innovative approach to the study of RSV by leveraging human pediatric biospecimens to determine a novel NK cell–SPM immunoregulatory axis and utilizes cutting-edge mechanistic immunology techniques to identify and probe novel targets in RSV. This proposal has relevance to public health as an unprecedented post-pandemic surge of pediatric RSV infection is currently overwhelming hospitals and pediatric ICUs and identifying new immune targets for potential therapeutic or preventative strategies is urgently needed.

NIH Research Projects · FY 2026 · 2023-12

PROJECT ABSTRACT Glial cells of the central nervous system (CNS), such as astrocytes and microglia, are intimately involved in tuning neural circuit activity, which is crucial in brain areas where increased activity leads aberrant behavior. The amygdala is one such limbic brain area that encodes associations between environmental stimuli and fear, and fires only if a threat is detected. The basolateral amygdala (BLA) is the amygdala input nucleus that integrates processed sensory information and initiates fear responses. However, pro-inflammatory signals induced in response to recurrent environmental threats boost BLA activity and increase fear responses. Glial cells in the amygdala are integral for inhibiting inflammatory signals that otherwise drive neural activity and fear behavior. Yet, many pathways in glial cells that tune BLA immune responses underlying fear are unknown. This lack of knowledge is largely due to an absence of methods that profile the mechanisms regulating cross-talk among glial cells at high throughput. Recently, I developed a new forward genetic screening platform to profile molecules controlling cell-cell interactions called SPEAC-seq (Systematic Perturbation of Encapsulated Associated Cells followed by SEQuencing). SPEAC-seq permits the co-encapsulation of cell pairs from differing cell classes within picoliter droplets (e.g., astrocytes and microglia), where prior to encapsulation one cell type expresses an inducible state-dependent fluorescent reporter and the other cell type is randomly perturbed with an sgRNA from a genome-wide CRISPR/Cas9 library. Cell pairs are encapsulated, co-cultured, and then sorted in droplets at high throughput. Isolation of sgRNA sequences from droplets containing an activated cell permits the association of molecular regulators with cellular states. My preliminary SPEAC-seq data uncovered amphiregulin (AREG) in microglia as a negative regulator of pro-inflammatory NF-kB signals in astrocytes by signaling through EGFR. Interestingly, microglia-astrocyte AREG-EGFR signals negatively regulated expression of the BLA-restricted transcription factor ETV1 in astrocytes. In preliminary studies, I found that ETV1 drove pro-inflammatory NF-kB signaling in astrocytes and was induced in response to chronic stress in the BLA. Chemogenetic activation of BLA astrocytes boosted helplessness behavior following chronic stress while genetic perturbation of Areg in BLA microglia or Egfr in BLA astrocytes exacerbated fear responses following chronic stress. Together, these data suggest that AREG+ microglia limit the activation of NF-kB in EGFR+ astrocytes by suppressing ETV1 and stress-induced fear behavior. In this proposal, I hypothesize that AREG+ microglia limit ETV1-driven NF-kB activation in EGFR+ BLA astrocytes to decrease stress-induced fear behavior. In Aim 1, I will test the role of AREG+ BLA microglia in limiting astrocyte ETV1 expression and NF-kB signals. In Aim 2, I will uncover the transcriptional and epigenetic networks of ETV1 and its family members in BLA astrocytes by multi-omic Perturb- seq. In Aim 3, I will determine how acute or chronic activation of ETV1+ astrocytes controls fear behavior. IN SUM, this R01 proposal will study how an anti-inflammatory cell circuit in the BLA shapes stress-induced fear.

NIH Research Projects · FY 2025 · 2023-12

Peripheral artery disease (PAD), an arterial occlusive disease that impedes blood flow to the lower extremities, can develop into critical limb ischemia (CLI), characterized by chronic ischemic rest pain with high risk for amputation and cardiovascular complications. Patients with diabetes are disproportionally afflicted by both PAD and CLI with limited medical interventions to improve limb perfusion. Angiogenesis is impaired in diabetic patients and the mechanisms controlling this process are not fully understood. MicroRNAs (miRNAs) are small, non-coding RNAs capable of repressing gene expression and are involved in a variety of pathophysiological processes with important therapeutic potential, though their role in angiogenic signaling pathways in diabetic CLI remains poorly defined. Because miRNAs exhibit high conservation across species, sequencing of miRNAs from the plasma of human subjects with diabetes and increasing severity of PAD and diabetic mice with limb ischemia was used to identify overlapping, new miRNA targets including a top candidate miR-130b. Preliminary gain and loss-of-function studies revealed that miR- 130b overexpression rapidly promoted proliferation, migration, and sprouting in endothelial cells (ECs), whereas miR-130b inhibition exerted anti-angiogenic effects. ECs exposed to high glucose downregulated miR-130b and co-transfection of miR-130b under high glucose conditions accelerated EC wound closure. Local delivery of miR-130b mimics into ischemic muscles of diabetic db/db mice following femoral artery ligation (FAL) promoted revascularization by increasing angiogenesis and markedly improved limb necrosis and amputation. Mechanistically, overlapping downregulated transcripts from RNA-seq and miRNA prediction algorithms identified that miR-130b directly targeted and repressed inhibin-b-A (INHBA), a subunit involved in the formation of activin A, and downstream Smad2 signaling. Indeed, ectopic delivery of siRNA targeting Inhba in db/db ischemic muscles following FAL improved revascularization and limb necrosis, recapitulating the phenotype of miR-130b delivery. These observations provide the foundation for the central hypothesis that the miR-130b-INHBA signaling axis may serve as a critical regulator of EC angiogenic responses for patients with PAD and diabetes at risk of developing CLI. To better understand the precise role of miR-130b in INHBA signaling and angiogenesis, we will in Aim1 delineate the molecular basis for miR-130b’s ability to regulate INHBA signaling and EC angiogenic functions. In Aim2, we will determine the effect of altered miR-130b expression in experimental critical limb ischemia in diabetic mice. Finally, in Aim3 we will assess the expression of the miR-130b-INHBA signaling axis in a unique cohort of human subjects with CLI with or without diabetes. Our studies will address a major gap in our understanding of diabetic CLI and inform how miR-130b- INHBA mediated control of EC angiogenic functions may provide new targets for therapy.

NIH Research Projects · FY 2026 · 2023-12

Although older adults account for 40% of inpatient surgical procedures and for two-thirds of all patients undergoing the 10 most common operations, their numbers are on the rise, and they suffer significantly worse outcomes than younger counterparts, critical knowledge gaps exist in how to best care for them and improve their outcomes. Surgeon-scientists are an important bridge between investigation and bedside patient care. Unfortunately, the surgeon-scientist workforce is threatened by increasing pressures to generate clinical revenue, the declining numbers of surgeons with federally funded research programs, and lack of mentorship for future generations. Research training during surgical residency has been suggested as a solution for reversing this trend and some have predicted that surgeon-scientists will become “extinct” without such opportunities. There is a critical need to train surgeons in aging research who will be able to tackle important research and clinical challenges to meet the growing need for surgery among older adults. We propose the Mentored Research Training in Aging and Surgery (MERITAS) Program at the Center for Surgery and Public Health (CSPH) at Brigham and Women’s Hospital (BWH) to train surgical residents in health services research at the intersection of surgery and aging with a particular focus on frailty, Alzheimer’s Disease and Related Dementias, Multimorbidity, and Serious Illness. The overall goal of this training program is to create a community of n=8 superbly trained surgeon-scientists to conduct studies to take on the mounting research, clinical, and policy challenges to improving care for older surgical patients. The MERITAS program has three aims: 1) To leverage existing rigorous post-doctoral training infrastructure at CSPH, BWH, the Longwood Medical Area (Harvard Medical School and the TH Chan Harvard School of Public Health) and the Boston Claude D. Pepper Older Americans Independence Center to generate resident-led, hypothesis-driven research from concept to dissemination intersecting surgery and aging under the mentorship of a multidisciplinary cadre of health services researchers in aging and surgery. 2) To increase access to rigorous training in health services research in surgery and aging for surgical residents from clinical training programs lacking resources to support such research training through targeted recruitment and mentorship. And 3) To launch a community of next-generation health services researchers in surgery and aging prepared for independent research careers through networking, longitudinal sponsorship and alumni activities. The program will be co-led by surgeon-scientists Drs. Zara Cooper and Louis Nguyen, and supported by 14 experienced mentors, with critical input from an Executive Committee and External Advisory Board comprised of researchers and leaders in surgery, geriatrics, epidemiology, health policy and other key disciplines. This program will yield a cohort of future surgeon-scientists dedicated to improving the health and well-being of older surgical patients.

NIH Research Projects · FY 2026 · 2023-12

Inflammation negatively affects Tregs stability and suppressive function, impairs the induction of solid organ transplant tolerance and enhances acute and chronic rejection, leading to worse recipients’ long-term outcomes. Insights into key inflammatory pathways that suppress Tregs during allo-immune activation can revolutionize our therapeutic approach to promoting stable endogenous Tregs. Our work strongly suggests that an inflammatory cytokine, macrophage migration inhibitory factor (MIF), secreted by myeloid cells and alloreactive T cells during rejection and inflammation binds CD74 (MIF receptor) on the surface of activated Tregs to dampen its function and homeostasis with opposite effect on alloreactive T cells. We show that inhibiting this novel pathway either through monoclonal antibodies or genetic deletion promotes immune regulation and induce exhaustion of alloreactive T cells leading to indefinite heart allograft survival in a murine fully mismatched model. MIF is a potent pro-inflammatory cytokine produced and released by immune cells, particularly macrophages, and plays an integral role in innate immunity. However, its role in adaptive immunity is not well known and will be investigated. Our data show that graft infiltrating Tregs and Teff upregulate their expression of MIF receptor CD74. However, graft infiltrating myeloid cells and Teff but not Tregs secrete MIF. MIF then binds to CD74 on Tregs surface to suppress and on Teff to promote their function. Our single cell transcriptomic analysis, TCR sequencing, flow cytometry and functional assays showed that CD74 deficient graft infiltrating Tregs reduced Interferon regulatory factor 1 (IRF1) and shifted toward a more stable Treg phenotype that exhibits superior suppressive ability in vitro and in vivo. IRF1 is a negative regulator of Foxp3 transcriptional activity that contributes significantly to Treg instability in the setting of inflammation. Interestingly, CD74 signaling is exerted through its intracellular-domain (ICD), shown to bind to transcription factors to modulate gene expression. Recent work identified IRF1 as one of the binding TFs for CD74-ICD13. In this application we will test the hypothesis that a novel inflammatory pathway in which MIF produced by myeloid cells and Teff suppresses Treg function through IRF1 mediated CD74 signaling (SA1), while promoting the Teff function (SA2). We will also develop a clinically relevant strategy to dampen the MIF mediated inflammatory process in the allograft using anti-CD74 monoclonal antibody generated in our lab (SA3). This anti-mouse CD74 is equivalent to the FDA approved human anti CD74 (milatuzumab). Hence, this work has high translation potentials.

NIH Research Projects · FY 2026 · 2023-11

PROJECT SUMMARY Atopy, Asthma and food allergies are highly prevalent inflammatory disorders that are increasing at an epidemic rate. Progression of allergies from one system to other, ultimately involving multiple organs, called “atopic march” is not uncommon in infancy. Aberrant type 2 immune responses to allergens have long been appreciated as the major driver of allergic reactions, with symptoms of very severe bronchospasms in asthma to vomiting and diarrhea observed in food allergies. Type II cytokines (IL-4 and IL-13) are both the drivers and effector molecules that drive allergic reactions. It is believed that IL-4 and IL-13 cytokines work on innate and adaptive immune system to induce and perpetuate type II inflammatory allergic reactions. We undertook a scRNAseq analysis to identify the cell types where IL-13R is expressed and we identified a cluster of enteric sensory neurons and multiple clusters of DRG and nodose ganglion neurons that express IL-13-specific receptors. This raises the issue of what is the role of IL-13R on the sensory neurons in the lung and gut in regulating allergic inflammation. Our preliminary data show that enteric neuron-specific deletion of IL-13R (Il13ra1) failed to control the worm burden and fecundity of the intestinal helminth parasite Heligmosomoides Polygyrus (Hp). Furthermore, Hp-infected Il13ra1 conditional knock-out (cKO) mice displayed severe impairments in the upregulation of Nmu and Calcb neuropeptide expression, suggesting neuronal IL-4/IL-13-receptor signaling contributes to the amplification of immunomodulatory neuropeptide expression during intestinal helminth infection. Initial scRNAseq of the immune and non-immune cells of the intestines of the Hp- infected IL-13R conditional knock-out mice suggest epithelial cell reprogramming and reduction is the number of ILC2. Based on these preliminary data, we hypothesize that cytokine-neuropeptide feedback loops between immune cells and neurons regulate type 2 inflammation and the development of type II allergic inflammation. We propose two specific aims to address this hypothesis: 1) Determine the role of type -2 cytokine receptor, specifically IL-13R signaling in neurons for the development of type 2 inflammation in the lung and gut; 2) How do the cytokine receptors on enteric neurons affect barrier integrity and host defense? Together these two aims will begin to address the functional role of type II cytokine receptors (specifically IL-13R) in regulating neuronal activation, neuropeptide production, and development of type II inflammation. The project will also focus on characterizing novel molecular interactions between cytokines, neurons and the immune system to regulate the differentiation and function of the innate and adaptive immune system, with an ultimate aim to find translatable therapeutic targets for chronic allergic diseases.

NIH Research Projects · FY 2026 · 2023-11

Abstract: Many gut-infecting bacterial pathogens undergo growth expansion in the gut lumen as part of the virulence, and as supported by their underlying metabolism. Commensal metabolism in the gut provides an opportunity to impede these processes by depriving pathogens of growth- promoting nutrients. Using the amino acid and carbohydrate-fermenting pathogen Clostridioides difficile, we will evaluate how redox metabolism of target carbon sources by the pathogen and commensals can enhance or limit its ability to colonize and infect the gut. Analyses will leverage innovative approaches to track nutrient flow among pathogen, commensal, and host tissues. Studies will further perturb host environments using carbon sources that are fermented by the pathogen, protective commensals, or both, to assess their role in disease pathogenesis and progression. Findings will inform small molecule and bacteriotherapeutic interventions in disease prevention and treatment.

NIH Research Projects · FY 2026 · 2023-11

Project Summary Delayed-type drug hypersensitivity reactions (dtDHR) are a major yet underappreciated public health problem, occurring commonly in skin-limited form, morbilliform drug eruption (MDE), and causing significant morbidity and mortality in severe systemic forms, Stevens-Johnson syndrome/toxic epidermal necrolysis (SJS/TEN) and drug reaction with eosinophilia and systemic symptoms (DRESS). dtDHR are significantly understudied. Consequently, understanding of immunopathogenesis has been limited, and subsequent to that, so too clinical care. Two major clinical limitations are the lack of a reliable laboratory test to identify culprit drug and an inability to accurately screen patients to prevent dtDHR. Our recent work in identifying T cell subsets mediating disease employed novel technologies to overcome research barriers in dtDHR and in-so-doing made several critical observations that, along with supporting publications by others, leads directly to this proposal. Herein, we propose the overarching, potentially transformative hypothesis that clonal repertoire analysis of skin could serve as a readout to identify culprit drug in dtDHR and to screen patients prior to drug administration to prevent dtDHR. Importantly, while this proposal has major implications for clinical care, it is purposefully designed to interrogate clonal repertoire and mechanisms of pathogenic T cell activation both broadly and mechanistically through innovative translational research as a critical next step. To achieve its goals, this grant builds on ample preliminary data and uniquely available patient populations along with the highly complementary expertise of its investigators to ensure feasibility and scientific rigor through three related but independent aims. First, it aims to interrogate clonal repertoire during active dtDHR using (i) T cell receptor (TCR) sequencing (seq) + single cell RNA seq on prospectively collected skin and blood and (ii) high-throughput TCR beta (HT-TCRb) sequencing on retrospectively obtained skin samples, from SJS/TEN, DRESS, MDE and healthy controls. Sample size allows breadth and depth across patient HLA alleles and culprit drugs. Second, it aims to investigate the temporality in vivo and reactivity ex vivo of pathogenic T cell clones. Specifically, using HT-TCRb sequencing, it tests whether drug- reactive T cells persist in skin and blood after disease resolution and/or pre-exist in skin prior to drug exposure. Study populations include patient cohorts and drug-tolerant controls. Third, it aims to identify the antigen specificity of pathogenic T cells in dtDHR by assaying in vivo the immunopeptidome with subsequent ex vivo mechanistic experimentation. In sum, this work has potential to profoundly impact the fields of dtDHR and more broadly adaptive immunity by generating incredible insight into in vivo mechanisms of antigen presentation and T cell activation, and concurrently clinical care by advancing toward development of much- needed diagnostic tests.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY Traumatic injuries caused more than 200,000 deaths and the loss of 2 Million potential life years in the US in 2019. Black, Hispanic, rural, and low-income populations bear a disproportionate burden of injury incidence and mortality. There is mounting evidence that proximity to and capacity of trauma centers are substantial drivers of these disparities, but current evidence does not identify or quantify specific system-level interventions that might reduce health disparities. We propose the following aims to identify strategies to mitigate disparities in trauma outcomes: Aim 1: Examine approaches to trauma system planning in states with and without disparities in trauma outcomes. We will conduct a series of in-depth case studies with a purposive sample of state trauma systems (n=11) identified based on injury mortality trends from 2000-2020. Using document review and semi-structured interviews with stakeholders responsible for trauma system planning in each state (e.g., medical directors, nurse managers), we will examine the overall approach to trauma system planning, and specifically probe for planning decisions intended to address disparities. Aim 2: Identify modifiable trauma system characteristics associated with injury disparities and decompose the structural pathways causing injury disparities. We will conduct a national survey of state trauma systems to identify existing trauma system resources (e.g., infrastructure, personnel), then link survey responses with outcomes data from the Centers for Disease Control and the Agency for Healthcare Research and Quality. After identifying trauma system characteristics associated with injury outcomes among disparities populations, we will use mediation pathway models to decompose the effects of trauma system resource allocation as a determinant of injury disparities. Aim 3: Identify and prioritize strategies to mitigate injury disparities through trauma system planning and resource allocation. We will conduct a Delphi consensus panel with national stakeholders, in partnership with the Coalition for National Trauma Research (CNTR). We will ask panelists to evaluate and prioritize recommendations trauma system interventions to mitigate disparities, drawing from the results of Aims 1 and 2, and from the literature. Impact of proposed work: Our proposed work will facilitate efforts to reduce disparities in injury outcomes by contextualizing current approaches to trauma systems planning, identifying specific trauma care resources that mediate injury disparities, and identifying actionable strategies to address disparities at the trauma system level. Our partnerships with national trauma leaders will support dissemination and implementation throughout the US.

NIH Research Projects · FY 2025 · 2023-09

PROJECT ABSTRACT Sugar-sweetened beverage (SSB) consumption in early childhood has profound implications on future health risks into adulthood. Family-based strategies have demonstrated effectiveness in reducing chronic health risk among children, yet few family-based interventions have specifically targeted healthy beverage consumption. This study advances a long-term goal of developing scalable, multi-level interventions to promote intergenerational health among indigenous families. Using community-based participatory methods and an indigenous conceptual framework – the Navajo Wellness Model – we will conduct a cluster randomized trial to study the effectiveness of Water is K’é, an innovative multi-level intervention designed to promote healthy beverage consumption through strengthened connections with Navajo culture, health information and access to reliable drinking water. Early child education sites will be randomly assigned to immediate or delayed intervention. Children enrolled in the programs, their primary caregiver and other family members will be enrolled and followed for 12 months to assess beverage consumption, health outcomes, as well as family cohesion and cultural continuity. Using a mixed methods design, we will explore how intergenerational family dynamics influence the way in which Water is K’é shapes family health and wellbeing. Finally, we will build greater research capacity in among Navajo partners through Community Advisory Group guidance and leadership, as well as mentorship of tribal college students interested in community research.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY / ABSTRACT Nuclear hormone receptors (NRs) are among the most widely targeted proteins in malignancy as they drive a number of common cancers. Hormones directly bind to NRs, leading to downstream transcriptional signaling. Some of the most common cancers (breast and prostate) are a result of aberrant NR signaling (estrogen and androgen receptor, respectively), and are treated using NR-targeting drugs like selective estrogen receptor modulators (SERMs) and degraders (SERDs). Other NRs, including glucocorticoid and thyroid receptors, are targeted in other diseases. Still others are orphan receptors or have yet to be therapeutically targeted. My research focuses on an important yet unanswered aspect of NR biology. It is known that hormone binding to NRs leads to transcriptional activation, followed by NR protein degradation. NR degradation regulates its transcriptional activity, but the mechanism of NR degradation and its effects on NR signaling remains not well characterized and is a central question in the field of NR biology. Understanding this phenomenon is clinically important: widely prescribed drugs for breast and prostate cancer work via binding and degrading specific NRs. I have recently discovered that, after hormone binding to NRs, the HECT E3 ubiquitin ligase UBR5 binds to and degrades NRs. Importantly, I found that multiple NRs are degraded by UBR5, delineating a common degradation mechanism of at least a large subset of NRs. We discovered that UBR5 regulates NRs following ligand activation by binding to the same site as nuclear co-activators (NCOA), which facilitate NR induced transcription, resulting in direct competition on chromatin. UBR5 therefore is not only a regulator of NR protein stability, but of transcription. However, we and others have observed that NRs can undergo ligand induced degradation without ligand induced activation, implying that NRs discriminate between NCOA and UBR5 binding; characterization and control of these interactions would lead to a specific induction of NR activation or degradation (or both), leading to significant therapeutic control over physiologic or disease processes reliant on hormone signaling. My goal is to characterize UBR5's role as a transcriptional regulator by direct degradation on chromatin and by doing so, enable an understanding of the pathways of nuclear hormone activation and degradation. If successful, this may enable the rational design of hormone derivatives that can decouple NR activation and degradation, allowing for control over hormone receptor signaling. The research proposed here also will yield basic insight into chromatin and transcriptional regulation, and ubiquitin E3 ligase biology. To achieve this long- term goal, we must first develop: a characterization of the biochemical interactions between UBR5 and its NR substrates (Aim 1), a better understanding of UBR5's regulatory capabilities over NR transcriptional activities (Aim 2), and a broader characterization of small molecule hormone derivatives that lead to NR activation, degradation, or both (Aim 3). These aims form parallel lines of investigation that seeks to establish E3 ligases as transcriptional regulators and create a platform for rational drug design of hormone derivatives.

NIH Research Projects · FY 2025 · 2023-09

The role of T cells in RA is established, but nearly all research has focused on CD4 T cells. We have shown that CD8 T cells are similarly expanded and activated in inflamed RA synovium yet do not belong to the typical granzyme (Gzm) B+ cytotoxic T lymphocyte (CTL) subset. Instead, they express high levels of GzmK whose function is not well defined. Remarkably, we find that GzmK drives a new pathway of complement activation. We show that GzmK can cleave C4 and C2 into C4b and C2a to generate an active C3 convertase (C4b2a) that cleaves C3 to C3a and C3b. We show that fibroblasts express the highest levels of complement components C2, C3 and C4 in the synovium and can secrete these proteins in response to CD8 T cell-derived TNF and IFNg. We show that GzmK can elicit formation of an active C3 covertase by cleaving complement components secreted by fibroblasts. Further, we demonstrate that GzmK generated C3a and C3b are bioactive and can drive mast cell degranulation and opsonize cells, respectively. We hypothesize that GzmK-mediated complement activation by GzmK in vivo occurs on cell surfaces as we show GzmK binds strongly to plasma membranes. Recently published research by others shows that complement activation drives inflammatory priming of murine synovial fibroblasts, mediated by mTOR activation with downstream metabolic changes and production of pro-inflammatory molecules. We propose to determine if human CD8 T cell-derived GzmK can induce inflammatory priming of human synovial fibroblasts by mediating local cellular complement activation and whether GzmK can drive complement activation and inflammatory arthritis in murine models. In Aim 1, we study GzmK binding to cell surfaces and determine its ability to assemble a C5 convertase, resulting in the generation of the anaphylatoxin C5a and a C5b-9 membrane attack complex. In Aim 2, we define the ability of GzmK to induce inflammatory priming of synovial fibroblasts via complement activation, by determining the effects of 1) recombinant and live CD8 T cell-derived GzmK on the inflammatory and metabolic activation of synovial fibroblasts. Then, using CRISPR GzmK-deleted T cells or CRISPR complement factor or complement receptor deleted fibroblasts, we confirm if these effects are mediated directly by GzmK and complement. In Aim 3, we use GzmK globally deficient mice and targeted CD8 T cell GzmK-deficient mice to demonstrate the role of GzmK in vivo in driving inflammation, complement activation, joint damage and fibroblast activation in the CIA mouse model of inflammatory arthritis. Together, these studies define a new pathway of complement activation mediated by lymphocyte- derived GzmK, and its role in inflammatory fibroblast activation and joint pathology. These studies will unravel key elements of how the pathway works and determine its role in arthritis in vivo. These findings provide new insights into the immunopathology of RA and point to GzmK and CD8 T cells as potential new therapeutic targets.

NIH Research Projects · FY 2025 · 2023-09

Project Summary/Abstract Uncertainty in exposure and outcome measurements poses substantial challenges to the identification and quantification of the causes of cancer. For example, although difficult to measure well, physical activity patterns form the basis of many etiologic hypotheses concerning cancer risk. Cancer cases identified in electronic health records (EHR) and other administrative ‘big data’ sources, such as Medicare claims data, are also subject to misclassification. This exposure and outcome uncertainty leads to considerable bias in estimated health effects, masking our ability to detect true associations, which are likely underestimated if detected at all. It is the role of measurement error and misclassification correction methods to validly and efficiently estimate the relationship between exposures and cancer outcomes. To accomplish this, a validation study is required for estimating key features of the error process. Although much has been accomplished in this domain over the years, the current aims address unsolved problems of high scientific significance that would otherwise remain unanswered without this additional work. We will drill down into the multi-faceted themes that arise in cancer research, tackling several seminal new directions of critical importance for the translation of the results of population-based research to practice and policy. These methods will include estimation of the effects of within-individual change in lifestyle behaviors on cancer risk corrected for measurement error in the change variables, utilizing complex, currently under-accessed validation studies of diet and physical activity comprised of repeated paper and online questionnaire self-reports and repeated concentration and recovery biomarkers to obtain relative risk estimates unbiased by general measurement error structures which may include correlated and biased errors, and estimating effects of exposures, including medications, other clinical treatments, and health behaviors, on cancer incidence in EHR data. The new methods will be applied to studies of the impact of within-participant change in alcohol intake on breast cancer incidence in the American Cancer Society’s CPS-II cohort and in Harvard’s Nurses’ Health Study, and to a study disentangling the impacts of diabetes and diabetes medications on colorectal cancer risk in Yale New Haven’s Epic EHRs. Dissemination is a central feature of this research. User-friendly publicly available software will accompany all new methods to be developed. The new methods will be disseminated through short courses and lectures at national and international epidemiologic and statistical conferences, and through the development of a massive online open course (MOOC). We have assembled an outstanding team of experts in measurement error methods and statistical theory, along with an exceptional team of cancer epidemiologists with much prior collaborative experience with the methods team, to guide the developments and their applications to the scientific problems at hand. With the talented junior faculty and trainees to be recruited for this project, we will solve the challenging problems that have been identified.

NIH Research Projects · FY 2025 · 2023-09

Despite compelling evidence that routine patient-reported outcome (PRO) measurement can improve outcomes and patient experiences in oncology, PRO reporting remains low among patients receiving care within community cancer care (CCC) settings. The goal of this proposed study is to increase breast cancer patient engagement in PRO reporting by establishing flexible and contextually aligned PRO implementation models in CCCs. Completing this work will set the stage to increase routine use of PROs in clinical care to improve health outcomes among patients in CCCs. In partnership with the Association of Community Cancer Centers (ACCC), we will develop models for successful implementation of PRO reporting in five CCCs in the ACCC network (Boston Medical Center, Boston, MA; Baptist Hospital, Memphis TN; Brooklyn Methodist, Brooklyn NY; University of Maryland, Baltimore, MD; Luminis Health, Lanham, MD). The HIT platform for this study will be imPROVE, an Epic-integrated PRO platform centered around the preferences, needs and values of breast cancer patients. In line with the priorities of the NCI, we will (Aim 1) identify challenges, barriers, and enablers to HIT-assisted PRO data collection among breast cancer patients receiving care in CCCs and develop PRO implementation models that address these factors; (Aim 2) conduct a phased implementation of the PRO data collection models in five CCCs using PDSA cycles to refine models and the RE-AIM framework to evaluate success; and (Aim 3) explore the potential of detecting differences in outcomes based on PRO data used in clinical care to inform future quality improvement efforts. Our expected outcomes are generalizable knowledge of the factors that trigger or deter breast cancer patient engagement, and models with training materials and tools for feasible, pragmatic, and sustainable PRO program implementation across the ACCC network. More than 80% of cancer patients nationwide receive their care at CCCs, and approximately 65% of the nation’s cancer patients are treated by a member of the ACCC network. The completion of this foundational work, with broad dissemination of lessons learned through the ACCC and beyond, will unlock the benefits of PRO reporting to a broader range of patients and advance cancer care. This work will also support a continued program of research to expand implementation and evaluation of PRO programs among other patient populations in oncology and further the use of PRO data for quality improvement.