Brigham And Women'S Hospital

NIH Research Projects · FY 2026 · 2022-02

APOE4 is the strongest genetic risk factor for late-onset Alzheimer’s disease (LOAD). The role of human APOE variants in AD has been studied extensively in the regulation of microglia and astrocytes but not in neutrophils. APOE is also expressed in neutrophils and controls their activation. Moreover, neutrophils have been shown to play a negative role in AD mice via the induction of microgliosis. Thus, a key question is whether APOE variants derived from neutrophils control immune responses driven by microglia and contribute to disease progression. Our long-term goal is to define the role of APOE signaling in regulation neutrophil-microglia interactions in neurodegeneration and determine which phenotypes and functions play a role in AD. We made the following preliminary observations: 1) Induction of APOE expression in microglia in AD and tau mice is associated with a phenotype switch from homeostatic (M0) to neurodegenerative microglia (MGnD); 2) APOE4 drives a neurodegenerative signature in neutrophils; 3) Recruited APOE4-neutrophils promote MGnD-microglia in APP/PS1 and P301S mice. Based on these findings, we hypothesize that APOE4 inflammatory neutrophils promote MGnD-microglia and accelerate neurodegeneration and cognitive decline in AD. We will address our hypothesis in the following aims: Aim 1: Define how APOE variants in neutrophils affects microglia. We propose to 1) Define the role of APOE variants in neutrophils in the regulation of neutrophil-microglia crosstalk; and 2) Determine whether replacement of APOE4 neutrophils with APOE2/3 neutrophils will restore microglial neuroprotective functions. Aim 2: Define the impact of APOE variants in microglia on neutrophil recruitment to the diseased brain. We will 1) Determine whether APOE variants modulate microglia to induce recruitment of neutrophils to the brain; and 2) Investigate the spatial distribution of microglia and neutrophils in the brain of AD and tau mouse models. Aim 3: Define the role of APOE variants in human neutrophils and their impact on human microglia in AD. We propose to 1) Characterize human neutrophils isolated from APOE e2, e3 and e4 AD carriers and whether they directly regulate the MGnD signature in iPSC-microglia; and 2) Investigate the neutrophil-microglia spatial interactions in AD brain of human APOE e2, e3 and e4 AD carriers. IN SUMMARY, targeting the APOE-neutrophil-microglia axis may provide a novel approach for therapeutic modulation of innate immunity in AD and dementia.

NIH Research Projects · FY 2025 · 2022-02

PROJECT SUMMARY Women are twice as likely as men to develop depression, and among some women, reproductive transitions trigger unique hormonal risks for reproductive-endocrine mood disorders. Changing reproductive steroid dynamics contribute female-specific endocrine risk factors in postpartum depression (PPD), premenstrual dysphoric disorder (PMDD), and perimenopausal depression (PeriDep). However, PeriDep lags far behind PPD and PMDD, each of which has FDA-approved therapies that leverage the reproductive endocrine changes underlying hormonally-linked depression. For example, the neurosteroid allopregnanolone (ALLO) in its proprietary form brexanolone has proven antidepressant efficacy for PPD. Endogenous ALLO levels decline after delivery, as women traverse menopause, and are lower in women with than without depression. Despite parallels to PPD, and despite the large population potentially affected by PeriDep—approximately 5.4 million women are perimenopausal annually—the contributions of ALLO to PeriDep have not been investigated. Key pilot data show a protective benefit of the ALLO precursor progesterone (P4) in PeriDep and that P4 correlates with advantageous neuroprotective, inflammatory, and neurophysiologic sleep profiles, all known ALLO targets. Thus, this project will use a mechanistic placebo-controlled trial to uncover the behavioral and neurobiological mechanisms through which ALLO exerts its therapeutic effects in women with PeriDep. Specifically, the project will examine key mechanistic targets underlying depression to include behavioral (Aim 1a: rumination, negative attentional bias), circuit-based (Aim 1b: functional connectivity within default mode network and between default mode and salience networks), molecular (Aim 2a: circulating and magnetic resonance spectroscopy neurotrophic and pro-inflammatory molecules), and physiological (Aim 2b: sleep EEG wake after sleep onset) outcomes. Eighty women with mild to severe PeriDep will be randomized to double-blinded placebo or ALLO administered as a 60-hour brexanolone infusion, stratified by early vs. late perimenopausal status. Analyses will test the acute (immediately post-treatment) and durable (30-days post-treatment) effects of ALLO on each of the selected mechanistic outcomes, mirroring the efficacy and biological data in human and animal models of PPD. Results will be integrated (Aim 3) to determine how each mechanistic outcome mediates ALLO’s effect on global measures of depression severity and to examine modification by depression illness course and early vs. late perimenopausal status. This innovative project pairing a mechanistic intervention with robust behavioral and neurobiological outcomes will exploit mechanistic pathways underlying the role of ALLO in PeriDep and translate findings to identify novel therapeutic targets that are specific for PeriDep.

NIH Research Projects · FY 2026 · 2022-02

Project Summary Chronic pain affects over 25 million adults in the United States and is a major cause of disability. Currently available pain treatments such as opioids are often ineffective and associated with unacceptable side effects including respiratory depression and addiction. A major goal for new pain therapeutics is to inhibit the sensory neurons which transmit pain signals (nociceptors) selectively without affecting other neurons involved in innocuous sensation or the central nervous system. However, nociceptor-specific therapeutic approaches remain in their infancy. Viral-based gene therapy offers several attractive advantages in treating refractory pain, as viruses can be engineered to deliver a wide range of molecules, can be administered locally or systemically, and have been recently approved by the Food and Drug Administration for a number of indications. However, nociceptor-specific viral tools do not presently exist in large part due to the extraordinary heterogeneity of sensory neurons that has made it difficult to identify molecular features that are unique to these cells. Recent advances in single-cell genomics have enabled us to generate a cell atlas that describes the genes that are selectively expressed in mouse and human nociceptors. This proposal aims to uncover the endogenous gene regulatory elements that mediate nociceptor-specific gene expression patterns and engineer these elements into barcoded adeno-associated viral libraries. To accomplish this, we propose the following specific aims: 1) Mapping nociceptor-specific gene regulatory elements in mouse and human and 2) Generation of a nociceptor-specific AAV toolkit. The nociceptor-specific viruses we develop in this proposal will be immediately useful to the scientific community for accessing nociceptors in wild-type mice and likely other species. In addition, because we will prioritize gene regulatory elements that are conserved between mouse and human nociceptors, we are optimistic that the viruses we screen in mice will also drive nociceptor-specific gene expression in humans. These next generation nociceptor-specific gene therapies would be ideally suited for treating certain refractory pain disorders because they can be administered locally to neuropathic sensory ganglia through standard outpatient procedures and can be engineered to drive expression of ion channels that silence nociceptor activity only in the presence of a specific drug (chemogenetics). This opioid-free approach could have significant translational applications for patients with refractory chronic pain who often have no other options for relief.

NIH Research Projects · FY 2026 · 2022-02

Summary: Severe treatment recalcitrant atopic dermatitis (AD) is a debilitating condition with substantial population impact. Dermatology has experienced the emergence of targeted immuno-modulating drugs (IMDs) that have unprecedented efficacy in treating AD. Their optimal use is still unknown because their safety remains insufficiently characterized. A range of serious side effects are conceivable based on the immunologic pathways although it is unlikely that they will all play out in clinical practice. Quantifying or refuting these adverse effects is critical for a clinical benefit-risk assessment and personalized treatment decisions. Existing trials have not answered these questions and are unlikely to address them in the near future. The resulting uncertainty has led to both overly restrictive but also aggressive prescribing of highly efficacious IMDs and this proposal aims to close this glaring knowledge gap. We propose a population-based prospective drug safety monitoring system leveraging existing data sources that shortens the time to insights and provides high validity findings through advanced causal inference methods. Analyses of longitudinal healthcare databases cover a source population of >78 million Americans and include commercially insured and Medicaid beneficiaries. New and urgently needed safety insights will reflect clinical practice, including populations typically excluded from RCTs, like children, women in reproductive age, patients with complex diseases, minority populations, and patients with existing risk factors. The size of the claims data source increases statistical power and the linkage to electronic health records in subsets improves clinical depth. We use causal inference methods that demonstrated high validity in pilot data and complement them with a novel data mining approach to identify unsuspected events. Analyses are done with highest transparency and reproducibility to support clinical decision making. This project’s finding on the optimal use of IMDs in clinical practice will lead to more targeted prescribing and benefit large patient groups, including populations underrepresented in RCTs: children, older adults, pregnant women, racial minorities, patients with pre-existing infections, cancers, VTE and others. This project is highly innovative as it will generate directly applicable clinical insights on the safe and targeted use of new immuno-modulating drugs (IMDs) to treat atopic dermatitis. Leveraging existing claims data sources with added EHR data it builds on novel methods for causal inference to mitigate biases arising in real- world data analyses in dermatology. The expedited evidence generation via the proposed prospective monitoring system combined with our track record in pharmacoepidemiologic analyses, this research will efficiently close knowledge gaps for optimal IMD use in many underrepresented and high-risk patients.

NIH Research Projects · FY 2026 · 2022-01

PROJECT SUMMARY/ABSTRACT Cisplatin is a well-known chemotherapy that results in a high incidence of acute kidney injury (AKI), occurring in up to one in three adults receiving a single dose. Despite efforts to find less toxic but equally effective alternatives, platin-based therapies are still incorporated in up to 40% of chemotherapy regimens, and remain first-line treatment for a number of cancers. AKI occurring after oncologic treatments such as cisplatin is associated with significant morbidity and mortality; it is therefore of paramount importance to identify effective strategies aimed at prevention of cisplatin-associated AKI (CP-AKI). In Aim 1, we will examine the effect of 4 medications (Intravenous [IV] magnesium [Mg], mannitol, metformin, and statins) on the incidence of CP-AKI. We will leverage a unique database of >45,000 adult patients treated with IV cisplatin at 4 major cancer centers, applying the principles of target trial emulation to address common biases in observational studies. In Aim 2, we will build upon animal models showing the protective effects of IV Mg on CP-AKI and our own preliminary data showing that lower serum Mg levels are independently associated with AKI in cardiac surgery patients. We will conduct a randomized clinical trial (RCT) in mesothelioma patients receiving hyperthermic intraoperative chemotherapy with cisplatin (HIOCC) (n=80) comparing IV Mg to placebo for the attenuation of CP-AKI. We will collect blood and urine samples pre- and postoperatively, comparing changes in serum creatinine and novel markers of tubular injury between the two groups. In Aim 3, we will recruit 150 high-risk patients receiving IV cisplatin to examine whether a byproduct of the nicotinamide adenine dinucleotide (NAD+) pathway, urinary quinolinate normalized to tryptophan, is elevated in CP-AKI. If so, administration of agents directed at increasing NAD+ metabolites may be a promising preventative strategy for CP-AKI in future studies. The projects proposed in this K23 application will provide the training needed to 1) become an expert in onco-nephrology; 2) apply advanced epidemiologic techniques to observational datasets; 3) develop skills in prospective patient recruitment; 4) build a biobank; 5) and acquire skills in the conduct of RCTs. The PI has guidance from mentors Dr. David Leaf, an expert in AKI, RCTs, biomarkers, and onco-nephrology, as well as Dr. Deborah Schrag, a world-renowned oncologist, outcomes researcher, and clinical trialist. The PI has assembled an advisory committee of experts in causal inference, Mg homeostasis, statistics, and translational research. The combination of a world-class mentorship team, a rich scientific environment, and proposed training will support the PI’s goal to become an independent patient-oriented researcher in onco-nephrology.

NIH Research Projects · FY 2026 · 2022-01

Project Summary/Abstract Inflammatory bowel disease (IBD), consisting of Crohn’s disease (CD) and ulcerative colitis (UC), is a chronic inflammatory condition affecting the human intestine associated with significant morbidity and limited therapeutic options. Based on the current understanding of IBD pathogenesis, mucosal inflammation seems to arise from a dysregulated crosstalk between intestinal microbes and poorly understood host immune, genetic, and environmental factors. Current biologic therapies available for IBD target specific cytokines (e.g. TNF, IL12/23) or trafficking receptors but a substantial proportion of patients either lose response to, or have disease refractory to, these therapies. There is an urgent need to improve our understanding of IBD pathogenesis to fuel the discovery of additional therapeutic targets. Based on previous investigations, interleukin-1 (IL1) and its associated cytokine family may play a critical role in IBD pathogenesis. IL10 receptor (IL10R)-deficiency, a monogenic form of CD-like IBD leading to colitis and perianal disease within the first year of life, is characterized by enhanced IL1 production in macrophages in both patients and mice. Blockade of IL1 signaling reduces the severity of colitis in IL10R-deficient mice, and several IL10R-deficient patients have been successfully treated with anti-IL1 therapy as a bridge to curative stem cell transplantation. Deep mucosal and peripheral immunophenotyping of a large cohort of IBD and non- IBD subjects using mass cytometry (CyTOF) (Mitsialis et al, Gastroenterology, 2020) has demonstrated that active IBD, especially CD, is characterized by enhanced IL1 expression in specific myeloid populations. Although anti-IL1 therapy is not currently used in the treatment of IBD, this data has led to the hypothesis that there may be subsets of IBD patients with IL1-mediated inflammation, similar to IL10R-deficient patients, whose disease may be amenable to anti-IL1 therapy. This project aims test this hypothesis by specifically interrogating IL1-related transcriptomic and proteomic signatures in IBD. The aims of this project are to (1) define IL1-associated cellular populations in human intestinal mucosa and blood using single cell analyses (CyTOF and scRNA-seq), and (2) determine how these IL1-associated populations differ in IBD, non-IBD, and IL10R-deficient subjects, thereby enabling characterization of IBD phenotypes associated with an enhanced IL1 signature. It is anticipated that IBD patients with an enhanced IL1 signature may share transcriptomic/proteomic findings with IL10R-deficient patients and be defined by a common clinical disease phenotype such as stricturing, fistulizing, or otherwise severe disease. The hope is that the results of this work will guide and set the stage for anti-IL1 therapeutics in IBD.

NIH Research Projects · FY 2026 · 2022-01

Abstract Cancer mortality rates in the US declined by 26% between 1991 and 2015, but the decline has not been equal across all populations. Approximately 19% of the US population resides in rural areas; for over three decades, this population has experienced increasingly inferior outcomes when compared with their urban counterparts. The underlying cause(s) of the widening rural/urban gap in cancer outcomes is hypothesized to be multifactorial, with socioeconomic status (SES) both at the individual-level and geographic area-level likely playing a significant role. Gaps in evidence include the use of multiple “rural” definitions, a lack of evidence on associated geographic factors, and limited evidence based on multi-level approaches to understand the complex nature of rural disparities. Therefore, the overall objective of this study is to conduct a comprehensive examination of the underlying causes of rural/urban disparities in mortality among individuals diagnosed with cancer. In the first aim, the Surveillance, Epidemiology, and End Results (SEER) population-level data will be used to examine mortality among individuals diagnosed with cancer across three definitions of rurality, providing researchers and policy makers with the magnitude of differences by each definition. In the second aim, SEER will be linked via county- indicators to 16 databases (US Census Bureau, Area Deprivation Index, Bureau of Labor Statistics, County Health Rankings and Roadmaps, AMA Healthcare Workforce Mapper, BRFSS, Social Vulnerability Index, Health Information National Trends Survey, etc.). These linkages will allow us to estimate the contribution of specific area- level factors (e.g., area-level SES, access to high-quality care) on rural/urban mortality differences using effect decomposition methodology. In the third aim, the Reasons for Geographic and Racial Differences in Stroke (REGARDS) cohort (U01 NS041588) will be linked to state level cancer registry data using the Virtual Pooled Registry Cancer Linkage System (VPR-CLS). REGARDS includes longitudinally collected data for 30,239 participants (44% blacks), oversampled from stroke belt/buckle states (56% of participants from NC, SC, GA, TN, AL, MS, AK, LA) with large rural populations experiencing the highest mortality. REGARDS collects information at the individual level on sociodemographics, health literacy, and distance to healthcare, transportation, and risky health behaviors. Multi-level modeling and mediation modeling approaches will allow for the examination of the contribution of individual-level characteristics and the area-level characteristics simultaneously. Results will provide estimates of how much of the mortality disparity is explained by differences in urban and rural geographic characteristics overall as well estimates that describe the potential impact of hypothetical interventions on specific mediating factors. Findings will provide the critical evidence needed to inform policy and intervention development aimed at addressing the systemic disparities in mortality experienced by rural patients with cancer.

NIH Research Projects · FY 2025 · 2022-01

Project Summary/Abstract Acute graft-versus-host disease (aGVHD) is a major limiting complication of allogeneic stem cell transplantation for the treatment of leukemia/lymphoma and certain solid tumors. aGVHD occurs when T lymphocytes from donor bone marrow attack cells within the recipient that are critical to the function vital organs, namely skin, liver and gut. During the past three decades of research, Dr. Murphy’s laboratory has found that in skin and squamous mucosae, the first sites of effector-target cell interaction provoked by differences in minor histocompatibility antigens, involve T cells bearing Vb-specific markers that home selectively to epithelial rete ridges as well as the epithelial stem cell-rich bulge regions of murine hair follicles. This association results in death by apoptosis of adjacent epithelial cells that express the stem cell marker, cytokeratin 15. Systemic therapeutic inhibition of these effector limbs is riddled with problems that include collateral impact on immune reconstitution and blunting of graft-versus-tumor effects. However, we now know through discoveries made in collaboration with co-PIs of this proposal, Drs. Markus and Natasha Frank, that the skin and squamous mucosae harbor intrinsic cellular pathways capable of potent immunosuppression. Specifically, we have discovered a dermal dendritic cell that has stem cell characteristics, expresses the multidrug resistance transporter ABCB5, and also expresses PD-1. This cell has the ability to thwart T cell responses, including alloreactivity, and therefore holds significant but as yet unexplored promise as a skin- intrinsic pathway that may be leveraged to inhibit the tissue injury in aGVHD. In this proposal, Dr. Murphy and the Drs. Frank have combined forces to address the long-term objective of harnessing the skin’s intrinsic immunosuppressive capabilities to thwart aGVHD. The proposal’s specific objective is to determine the impact of PD-1-expressing immunosuppressive dermal mesenchymal stem cells on the earliest and most specific effector-target cell interactions responsible for aGVHD. To this end, we have proposed three specific aims: 1) to correlate spatiotemporal multiplex immune profiling with all aspects of disease morbidity and recovery in aGVHD; 2) to dissect ABCB5 dermal stem cell function in aGVHD and define PD-1/PD-L1-centered strategies for skin-intrinsic immunosuppression; and 3) to apply ABCB5+ dermal stem cell transplant strategies to influence aGVHD severity using spatiotemporal multiplex immune profiling as effector-target bioassay. The approaches involve state-of-the-art multiplex immunophenotyping of effector-target cell interactions that sensitively and specifically typify tissue injury in aGVHD, and leverage ABCB5 gene knockout and lineage tracing models in context of PD-L1 Ig administration and ABCB5+ stem cell engraftment. The rationale is that upon successful completion of this work, we will identify novel therapeutic approaches to the treatment and prevention of aGVHD skin and squamous mucosal pathology through manipulation of skin- intrinsic immunosuppressive pathways.

NIH Research Projects · FY 2025 · 2021-12

Coronary heart disease remains the leading cause of deaths in the US. Despite the success of lipid lowering, residual risk remains. Recent clinical studies support the role of inflammation in CVD, a longstanding topic of basic research. Group 2 innate lymphoid cells (ILC2) are innate lymphocytes that play essential role in obesity by promoting adipocyte beiging and recruiting beige cells to the white adipose tissue, thereby limiting obesity development. Limited information is available regarding ILC2 function in CVD, although transplantation of bone-marrow from ILC2-deficient mice promoted atherogenesis. Eosinophils (EOS) accumulate in blood or at the site of inflammation after allergic sensitization or parasite infection. Blood EOS count and EOS cationic protein (ECP) levels associate positively with major CV risk factors and CVD prevalence and mortality. Yet other studies reported reduced blood EOS count and ECP levels in patients with major adverse cardiac events and heart failure. Therefore, the role of ILC2 and EOS in human CVD remains unsettled. Our preliminary studies demonstrated ILC2 accumulation in mouse heart after myocardial infarction (MI). ILC2 deficiency (ILC2KO) or diphtheria toxin A (DTA)-induced ILC2 depletion (ILC2DTA) exacerbated cardiac dysfunction, myocardial cell death and fibrosis post-MI. Further study showed that ILC2 deficiency blunted blood IL5, an essential type-2 cytokine from ILC2 that controls the expansion and migration of EOS, dendritic cells (DC), and regulatory T cells (Treg) to the bloodstream or the site of inflammation. FACS analysis revealed deficiency of splenic, blood, or heart EOS and DC in ILC2KO mice post-MI. Administration of mouse recombinant IL5 reversed EOS loss in blood, providing a mechanistic explanation of ILC2 activity in alleviating cardiac dysfunction post-MI and suggesting a concurrent cardioprotective role of EOS and ILC2 in post-MI heart. Consistent with this hypothesis, exacerbated post-MI cardiac dysfunctions in ILC2KO mice got improved after mice receiving adoptive transfer of ILC2 or EOS from wild-type (WT) mice, but not ILC2 from IL5-/- mice. We recently reported high blood EOS count in patients with acute MI and EOS accumulation in mouse heart post- MI. EOS-deficient ∆dblGATA mice (EOSKO) demonstrated exacerbated cardiac dysfunction and myocardial fibrosis, all which can be corrected by repopulation of donor EOS from WT mice or by giving mice recombinant murine EOS cationic protein mEar1 (human ECP ortholog). Similar to the EOSKO mice, DTA-induced depletion of EOS in iPHIL mice (EOSDTA) also worsened the cardiac function post-MI. Based on these observations, we hypothesize that ILC2 protect heart from post-MI cardiac dysfunction. One mechanism is to release type-2 cytokines, such as IL5 to promote the development and migration of bone-marrow EOS to the bloodstream and heart where EOS exert a similar reparative role to that of ILC2 in mitigating ischemic cardiac injury. We propose two Aims to examine whether and how ILC2 protect heart from MI-induced cardiac dysfunction and to explore the EOS-mediated reparative mechanisms in response to myocardial ischemic injury.

NIH Research Projects · FY 2025 · 2021-12

PROJECT SUMMARY Inflammation is crucial for the host defense response, but unregulated inflammation is a key element of atherothrombosis and links plaque initiation to subsequent growth and acute rupture, leading to clinical cardiovascular (CVD) events. Clinical trials of inflammation inhibition have significantly reduced CVD events, confirmed the inflammation-CVD hypothesis, and stimulated interest in targeting inflammation specifically for CVD prevention. Such therapies could have widespread impact, since residual inflammatory risk in clinical practice is common and undertreated. Importantly, recent evidence demonstrates that the resolution of inflammation is not a passive process but occurs in an active coordinated process involving chemical mediators called specialized pro-resolving mediators (SPMs) that include resolvins, maresins, lipoxins, protectins (neuroprotectins), and aspirin-triggered pro-resolving mediators, each with characteristic biological actions. Yet a major gap in knowledge is which specific SPMs or sets of SPMs may be cardioprotective in human populations. Motivated by our exciting preliminary findings that plasma levels of certain SPMs are related to CVD risk and inflammatory traits, and in response to NHLBI NOT-ES-20-018: Promoting Fundamental and Applied Research in Inflammation Resolution to identify the role of SPMs in CVD, we propose to test the hypothesis that specific SPMs are early indicators of CVD protection from inflammation and are associated with its resolution over time. This cost-efficient proposal leverages resources from three well-phenotyped and genotyped prospective studies. We will examine an extensive panel of circulating plasma SPMs and proinflammatory mediators using a high-throughput mass spectrometry assay in relation to incident CVD (total 2339 cases) including repeated measures over time in a subset. Our primary aims are to: 1) Evaluate associations of SPMs with future CVD events in primary and secondary prevention populations enriched with chronic inflammation, and assess effect modification by randomized aspirin therapy vs placebo since aspirin has important pro-resolving effects on SPM biosynthesis; 2) Examine associations of circulating SPMs with CVD risk factors and downstream biomarkers and cytokines of systemic inflammation; and perform genome-wide association study of circulating SPMs, to better understand biological pathways for mechanistic insights into SPM functions; and 3) Assess temporal changes in SPMs over time in relation to concomitant changes in levels of downstream proinflammatory biomarkers and cytokines, and examine modulation of SPMs with randomized low-dose methotrexate vs placebo. We will examine functional actions of the relevant CVD-associated SPMs using ex vivo leukocyte assays for inflammatory gene expression and macrophage pro-resolving function. Results from this proposal will identify and validate resolution mediators and pathways that may play a pivotal role in cardioprotection and could have important public health significance since residual inflammatory risk is common and remains undertreated with current therapeutics.

NIH Research Projects · FY 2026 · 2021-11

Kaposi's sarcoma (KS) herpesvirus (KSHV) is the causative agent of KS and primary effusion lymphoma (PEL), and is tightly linked with multicentric Castleman's disease (MCD). These tumors occur most commonly in immunocompromised individuals, especially those with AIDS. There are no specific therapies for these malignancies. KS is the leading AIDS malignancy, and is epidemic in sub Saharan Africa. KS commonly involves the oral cavity and can disseminate to visceral organs. Saliva is the vehicle of transmission for KSHV. Latency is the hallmark of KSHV and gammaherpesvirus infection. KSHV latently infects cells, including tumor cells, and viral genomes persist as extrachromosomal, circularized, multi-copy, episomes. To persist in proliferating cells, viral episomes must replicate, and following mitosis, segregate to daughter cell nuclei. Tumor cell viability is dependent on latent KSHV infection. The latency-associated nuclear antigen (LANA) is one of several viral genes expressed in latency. LANA mediates KSHV episome maintenance, and is necessary and sufficient for episome persistence in the absence of other viral genes. In addition to episome persistence, LANA exerts important roles in transcriptional regulation. Epigenetic histone H3 lysine 4 (H3K4) tri-methylation (H3K4me3) marks are associated with actively transcribed genes and are deposited by histone methyltransferase (HMT) complexes. There are six HMTs (MLL1-4, Set1A/B) in mammals responsible for catalyzing methylation of histone H3 at K4 through a SET domain. LANA is highly enriched at H3K4me3 peaks at both viral and host chromatin, yet the mechanism of LANA recruitment to and its function at these sites remains unclear. We have discovered novel LANA interactions with HMT components and that LANA regulates specific HMT activity. Further, we find this HMT activity is critical for virus latency establishment. This work will use rigorous, detailed, in depth approaches to investigate the mechanistic basis of these findings. Experiments will investigate the role of the HMT activity in LANA mediated episome persistence. We will investigate the role of the LANA-HMT interaction in LANA and HMT chromatin targeting of virus and host, and its effects on H3K4me3 deposition and gene expression. Experiments will also investigate the mechanism of LANA’s regulation of HMT activity and its role in virus latency. LANA and HMT activity are critical for latency, and this work therefore provides novel and important insight into a fundamental component of KSHV biology.

NIH Research Projects · FY 2024 · 2021-09

ABSTRACT Adverse events (AE) during care transitions range from 19-28% and may lead to readmissions, representing an ongoing threat to patient safety. Early identification and escalation of patient-reported symptoms to inpatient and ambulatory clinicians is critical, especially for patients with multiple chronic conditions (MCC). Clinically integrated digital health apps have the potential to more accurately predict post-discharge AEs and improve communication for patients, their caregivers, and the care team. Such tools can provide individualized risk assessments of AEs by systematically collecting relevant patient-reported outcomes (PROs) and leveraging standardized application programming interfaces (API) to combine them with electronic health record (EHR) data. While patient-reported outcomes (PROs) are increasingly used in ambulatory settings, their use for real- time symptom monitoring and escalation during transitions from the hospital is novel and potentially transformative–by both empowering patients to better understand their individualized risks of post-discharge AEs, and improving monitoring while transitioning out of the hospital. Our proposed intervention is grounded in evidence-based frameworks for care transitions, and scaling and spread of digital health tools. To inform our intervention, we propose developing and validating a predictive model of post-discharge AEs for hospitalized MCC patients using relevant PRO questionnaires and electronic health record (EHR) derived variables. We will then combine, adapt, extend, and refine our previously developed EHR-integrated hospital and ambulatory-focused digital health infrastructure to support MCC patients in real-time symptom monitoring using PROs when transitioning out of the hospital. Our intervention uses interoperable, data exchange standards and APIs to seamlessly integrate with existing vendor patient portal offerings, thereby addressing critical gaps and supporting the complete continuum of care. Our multidisciplinary team uses principles of user-centered design and agile software development to rapidly identify, design, develop, refine, and implement requirements from patients and clinicians. Our team will rigorously evaluate this intervention in a large-scale randomized controlled trial in which we compare our real-time symptom monitoring intervention to usual care for patients with MCCs transitioning out of the hospital. Finally, we will conduct a robust mixed methods evaluation to generate new knowledge and best practices for disseminating, implementing, and using this interoperable intervention at similar institutions with different EHR vendors.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY While an infectious etiology has been proposed to be involved in the initiation and progression of Alzheimer’s disease (AD), more evidence is needed to support the roles of specific infectious agents and to identify mechanisms by which they act. Given the chronic nature of AD and based on our discoveries in animal models of AD, we hypothesize that a chronic and sub-acute infection with specific Bacteroides strains contributes to AD by affecting Ab and tau phosphorylation, aggregation, and clearance, and contributing to neuronal toxicity. In support of our hypothesis, we were the first group to demonstrate that Bacteroides not only correlated with Ab levels in the brain, but actually increased amyloid plaques when administered to animal models of AD. Strikingly, Bacteroides is elevated in AD, correlates with CSF levels of Ab42 and phospho-tau, and is associated with markers of gut inflammation in AD patients, suggesting a direct relevance to human disease. In our preliminary data, we found that B. fragilis affected cortical expression of genes involved in APP and tau phosphorylation that promote their aggregation, as well as genes related to memory and neuronal death. We also found that Bacteroides downregulated microglial genes important for the clearance of Ab. Depletion of Bacteroides with metronidazole decreased amyloid plaques, increased cortical expression of insulin degrading enzyme, which can degrade Ab, and affected genes involved in multiple protein degradation pathways. We found that Bacteroides could impair macrophage Ab phagocytosis in vitro and in vivo. Importantly, we found that transfer of human gut microbiota from AD patients into mice affected similar microglia genes involved in protein degradation and clearance, as well as pathways related to neuronal cell death, suggesting that the AD gut microbiota may harbor infectious agents that contribute to the disease process. Because Bacteroides species are highly prevalent in humans and exhibit a high degree of functional variation among strains, it is not likely that all Bacteroides contribute to AD. In this proposal, we will modify Koch’s postulates to detect Bacteroides strains associated with AD vs. healthy controls and non-AD dementia controls, isolate these strains in pure culture, and use in vitro assays to optimize strain selection for in vivo studies. We will then transfer AD-derived strains to WT mice and animal models of AD, measure AD pathology, and re-isolate our AD-derived strains to determine whether they play a causal role. To investigate potential mechanisms, we will also investigate whether AD strains can affect APP and tau aggregation and clearance, neuronal toxicity, or disrupt transcriptional networks in microglia and neurons. By investigating the molecular and functional diversity in Bacteroides in AD, our studies have the potential to identify novel pathogenicity factors in Bacteroides that could be used both for the diagnosis and treatment of AD.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY Rheumatoid arthritis (RA) is a systemic autoimmune disease affecting nearly 1% of adults causing a painful, destructive inflammatory arthritis with serious long-term consequences including chronic pain, disability, accumulation of morbidities, and excess mortality. Patients with RA are susceptible to developing interstitial lung disease (ILD), a devastating complication with high morbidity and mortality. Rheumatoid factor (RF) and anti-cyclic citrullinated peptide (ant-CCP) autoantibodies are elevated in the serum of two-thirds of patients with RA. Seropositive RA patients are more likely to develop RA-ILD. Previous studies suggest that mucosal surfaces of the lung may be an initiating site for RA, after smoking or exposure to other inhalants, where RF, anti-CCP, and other autoantibodies may be formed years before joint symptoms develop. Aberrant post- translational modifications (PTM) to proteins may serve as neoantigens forming local inflammation in the lungs and production of autoantibodies related to PTM. This site of lung injury may later manifest clinically as RA-ILD and articular inflammation may impact risk for subclinical RA-ILD through systemic inflammation. Therefore, RA-related autoantibodies, articular inflammation, and RA-ILD may be linked throughout the disease course of RA. These investigations will study whether RA-related autoantibodies and articular inflammation predict subclinical and clinically-apparent RA-ILD. In the first aim, we will perform a nested case-control study using a derivation dataset in the Brigham RA Sequential Study (BRASS) and a replication dataset in the Partners Biobank. BRASS and the Partners Biobank are patient registries for research. We have identified RA-ILD cases as well as controls with RA but no ILD in these research registries and propose to measure clinical and PTM RA- related autoantibodies. In the second aim, we will perform a multi-site prospective observational study of patients with new-onset RA who will undergo serial measures of articular inflammation and chest high- resolution computed tomography imaging to evaluate whether the burden of articular inflammation in early RA reflects risk for subclinical RA-ILD. In the third aim, we will analyze whether smokers in COPDGene with elevation of RA-related autoantibodies without articular RA are more likely to have subclinical ILD. COPDGene is a large US nationwide observational study that has already been phenotyped for presence or absence of ILA on chest computed tomography imaging. Dr. Jeffrey Sparks (the PI) is an Assistant Professor of Medicine at Brigham and Women’s Hospital and Harvard Medical School. He is an early-stage investigator previously funded by NIAMS through K23 and R03 awards to investigate the role of the lung in RA etiology and outcomes. These proposed studies will interrogate the overarching hypothesis that RA autoimmunity, articular inflammation, and ILD are intrinsically linked across the disease course of RA (pre-RA, early RA, established RA). These studies have high potential for impactful results that will elucidate the pathogenesis of both RA and RA-ILD and may ultimately provide the evidence basis for RA-ILD screening and prevention strategies for this devastating complication.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY Many authorities recommend the Mediterranean diet (MedDiet) for the prevention of cardiometabolic disease. These dietary recommendations are based on population averages and may not be best suited for a given individual. Preliminary data from our group and others support that a specific dietary intervention may have highly variable effects in different individuals due to the individual composition of the gut microbiome. Furthermore, we recently reported that autologous fecal microbiota transplantation (aFMT) derived from the time of maximal weight reduction enhanced the effects of MedDiet on maintaining cardiometabolic health in an RCT. This background supports our central hypothesis that the gut microbiome can modify the effects of MedDiet on cardiometabolic disease risk. However, no studies have utilized longitudinally collected data from RCTs to test this hypothesis. Most diet-microbiome studies are limited by the use of 16S rRNA gene sequencing yielding only very general taxonomic profiling, thus omitting strain-specific diet-related biochemical functions of microbes. To gain more advanced mechanistic insights, combining shotgun metagenomics and metatranscriptomics and metabolomics in an integrated framework presents a unique opportunity to probe both the composition and functionality of gut microbial communities. This proposed project will leverage two long-term dietary RCTs, the recently completed 18-month DIRECT-PLUS trial of 294 participants and the ongoing 3-year MIND trial of 604 participants, to examine whether individual gut microbial features modify the effects of MedDiet interventions on cardiometabolic risk and body adiposity (Aim 1) and identify metabolites in feces and metabolites in plasma of gut microbial origin that explain inter-individual differences in post-intervention changes in cardiometabolic risk and adiposity (Aim 2) in the DIRECT-PLUS trial. Findings from Aims 1 and 2 will be tested for replication in the MIND trial. In Aim 3, we will investigate long-lasting, post-intervention effects of combined MedDiet and aFMT, and characterize gut microbial changes during and after the interventions in an RCT of 90 participants. Our proposal addresses a major research priority, precision nutrition, emphasized in the 2020–2030 Strategic Plan for NIH Nutrition Research and is directly responsive to PAR-19-377: “Omics-guided Biobehavioral Interventions for Improved Health Outcomes: A Step Forward in Translation” as one of the first efforts to prospectively assess the functional role of gut microbiome in explaining inter-individual heterogeneity in response to dietary interventions. Built on existing biospecimen and data collected in two well-conducted long-term RCTs and state- of-the-art multi'omics technologies, our proposal is a highly cost-efficient opportunity to generate translatable, personalized dietary interventions grounded in reproducible biological mechanisms and contribute to the paradigm shift towards precision nutrition for improved cardiometabolic health.

NIH Research Projects · FY 2025 · 2021-09

We respond here to a Funding Opportunity Announcement (FOA) for multi-institutional teams to form a Glioblastoma Therapeutics Network (GTN). Basic scientists and clinical/translational investigators from three institutions in the Dana-Farber/Harvard Cancer Center (DF/HCC) have joined forces with their counterparts in the Stanford Cancer Center (SCC) to create the “Harvard/Stanford GTN”. UT Southwestern is also represented in one key collaboration. Distinctive features of this bi-coastal GTN include (i) broad and deep expertise in brain-penetrant, small molecule therapeutics and (ii) a strong presence in the emerging field of Cancer Neuroscience – a field that addresses the central role of the nervous system in glioblastoma pathogenesis. Our objective is to improve the treatment of adult glioblastomas (GBMs) and Astrocytoma, IDH- mutant, grade 4 by taking novel, effective, brain-penetrant small molecule drugs through lead optimization, to preclinical development and into early phase clinical trials. Our study plan features three projects: Project 1 targets metabolic reprogramming in Astrocytoma, IDH mutant, grade 4. Project 2 targets the constitutive, ligand-independent EGFR signaling observed in more than 50% of adult IDH wild-type GBM. Project 3 targets a recently appreciated forward-feeding gliomagenic loop between tumor cells and electrically active neurons in IDH wild-type adult glioblastomas. All three projects feature surgical window clinical trials of brain-penetrant drugs that are hitherto untested in GBM. In addition, Project 2 will develop novel allosteric inhibitors that promise to address a shortcoming of all current EGFR antagonists as GBM therapeutics – to wit, lack of a therapeutic window. Insights from clinical trials will be enhanced by a Pharmacological and Genomic Imaging Core (PGIC). This core will allow our trialists to monitor drug impact on glioblastoma cell populations using specialized single cell RNAseq protocols. Drug penetrance within tumors and drug-induced changes in key metabolites will be visualized using matrix assisted laser desorption ionization mass spectrometry imaging and non-invasive magnetic resonance spectroscopy methodologies. In addition to these clinical/translational research projects and the PGIC, the Harvard/Stanford GTN offers to host a Network Coordinating Center (NCC) for the broader GTN initiative (as described and specified by the FOA). Our proposed NCC offers essential skill sets in neuropathology, cancer genetics, clinical trials, biostatistics, and clinical trial design that will enable multiple GTN centers to work together in ways that exceed the sum of their component parts. An Administrative Core will serve as the primary contact and communication resource for the Projects, the PGIC, an Internal Advisory Board, the NCC, the GTN Steering Committee, and NCI program officials. The Harvard/Stanford GTN Principal Investigator is Tracy Batchelor, M.D. an experienced clinical trialist with much practical experience in leading large, multi- investigator/multi-institutional initiatives.

NIH Research Projects · FY 2024 · 2021-09

Project Summary Despite extraordinary advances in genome engineering, tools for precise and efficient gene correction across all cell types and desired edits remain lacking. Current programmable DNA cleavage tools, such as CRISPR-Cas9, rely on cellular DNA repair mechanisms, which are inefficient and do not function in post-mitotic cells. Thus, genome editing still needs efficient, robust tools that can make a variety of specific DNA sequence alterations. These tools could have broad applications across both basic biological discovery, allowing for new modalities of screening, and therapeutics, including engineered cell therapies. The proposed work will address these needs by combining computational discovery, biochemical characterization, and enzyme engineering to develop integrase-based tools for programmable, multiplexed insertion of large genes in diverse cell types independent of DNA repair. The discovery, characterization, and engineering of these new integrase proteins will both build upon our deep history of CRISPR enzyme discovery, as well as draw from new, high-throughput approaches to mine biological diversity. Complementary to the discovery of these new enzymes, we will combine Cas9-based genome editing with integrase engineering to develop programmable, multiplexed genome integration systems that do not depend on DNA repair mechanisms, allowing integration of large sequences in any cell type. We will explore delivery mechanisms, including viruses, electroporation, and novel lipid nanoparticle formulations to edit T cells and neurons. We will engineer aspects of the integrases, including protein engineering and site mutagenesis, to boost activity of the system and screen many insertion sites to develop design rules for the technology. Moreover, through studying orthogonal integrases sites we can develop multiplexed versions of the insertion tool to edit up to three sites in a given cell with superior efficiency over other tools. We will apply these multiplexed integrases to develop a new screening system, where tagging of multiple genes can be used for determining protein interaction partners in high throughput. Our new integrase systems will also be applied to the development of multiple-edited T-cells for improved immuno-oncology therapies. The multiple technologies resulting from these discoveries and engineering efforts will overcome the limitations of existing genome and epigenome engineering approaches and serve as a valuable resource for broader biomedical research. Programmable gene integration with CRISPR-recruited integrases will allow for more advanced genome engineering applications to be pursued in cells and in vivo, accelerating the pace of biomedical research, enabling greater exploration of basic biological processes and disease mechanisms, and promoting novel therapeutic developments.

NIH Research Projects · FY 2025 · 2021-09

Abstract The lentiviruses HIV-1 and HIV-2 cause acquired immunodeficiency syndrome (AIDS) in humans. The conventional wisdom that HIV-2 is a more benign cousin to HIV-1 was recently challenged, however, in a large study that found after 23 years of HIV-2 infection, cumulatively over 90% of participants had AIDS and 80% had died. An effective, safe, and scalable cure for HIV should account for any unique features that may characterize HIV-2 latency. During the course of its replication, HIV-2 integrates into human DNA and creates a reservoir that is a barrier to virus eradication - as is the problem for HIV-1. How similar, or different, these HIV reservoirs may be is unknown. The scientific premise of this proposal is that there are unrecognized unique features of HIV-2 biology that may impact virus eradication efforts, relative to HIV-1. As an example, the HIV-2 Vpx protein has recently been shown to counteract the Human Silencing Hub (HUSH), a highly conserved gene silencing complex known to repress HIV-1 proviruses, in ways that may impact virus latency and the reservoir landscape. HIV-2 vpx reactivates latent provirus transcription in primary human CD4+ T cells through degradation of the HUSH complex. To study HIV-2 latency, we have recruited participants with HIV-2 into our longitudinal HIV Eradication and Latency (HEAL) cohort. Our HIV-2 proviral landscape preliminary data has identified differences between the HIV-1 and HIV-2 proviral landscapes and we see phylogenetic evidence that suggests clonal expansion may maintain the HIV-2 reservoir, findings we propose to explore further in this proposal. We hypothesize that HIV-2 latency reversal differs from HIV-1 reactivation, provide data to support this, and propose experiments to investigate the mechanisms that may explain these observations. The goal of this proposal is to use HIV and host genomic information to define the landscape of HIV-2 integration and investigate the mechanisms that control HIV-2 latency and its reversal. Here we propose to leverage cutting-edge techniques to address important knowledge gaps in our understanding of HIV-2 infection biology. We hypothesize that HIV-2 differs from HIV-1 in integration landscape and reversal from latency and that Vpx plays a key role in these differences. Specific Aims of this proposal are to define the HIV-2 integration landscape in vivo, understand the role of HIV-2 vpx in virus latency, and investigate the transcriptional blocks to HIV-2 latency reversal. Our approaches test hypotheses that are central to understand HIV-2 latency and its reversal.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY Developing universal vaccines to influenza and HIV-1 is an urgent global goal. A critical challenge is that immune responses to native HIV-1 envelope (Env) and influenza hemagglutinin (HA) are dominated by non- neutralizing and highly strain-specific antibodies. Discoveries that some individuals produce broadly neutralizing antibodies (bnAbs) invigorated hope that, while not naturally dominant, broadly protective antibody responses are possible. Antibodies mature during through somatic hypermutation (SHM) and affinity-based selection in germinal centers (GCs) in competition with other antibodies that recognize different parts of the same virus. It is widely believed that a prime and boost vaccine tactic can effectively elicit bnAb precursors and strategically guide SHM trajectory can produce bnAbs. Challenges to this process are that native envelope proteins may not bind well to the bnAb precursor antibodies and may be poorly represented in the antibody repertoire. A strategic prime and boost strategy requires generation of designer viral envelope variants that bind well to bnAb ancestor antibodies acting as a primer, followed by modified variants to function as boosting immunogen(s) to shepherd bnAb maturation. This promising approach is hindered by time and effort required to identify Env or HA variants as immunogens, which traditionally require mutation library generation, in vitro static selection, cloning, expression, and validation testing. This extensive hands-on trial and error process greatly hinders the pace of progress. Here a new technology is proposed with power to explosively accelerate the pace of immunogen discovery by creatively harnessing the full spectrum of automated mutation and selection inherent in one of nature’s innovations in hyperevolution—namely the GC SHM and affinity maturation system—an automated in vivo dynamic mutation process coupled to parallel selection activity that dynamically shuttles superior binding variants back for further diversification and selection. In addition to dramatically improving binding affinity, the GC system can be engineered to generate new recognition. The objective is to create flipped GC systems in which antibody genes are replaced with viral envelope proteins— and deploy them for immunogen design. In contrast to dynamic antibody evolution to viral envelop protein in normal GCs, flipped GCs dynamically evolve viral envelop protein toward user-defined antibodies (e.g. select bnAb precursors and intermediates). The overall hypothesis is that, in the context of key modifications, the GC/affinity maturation system is sufficiently flexible to permit bioengineered viral envelope proteins to affinity mature toward user-defined bnAb precursors and intermediates. The objective will be pursued with two aims: 1) to establish parameters to engineer GCs as a platform for non-Ig protein evolution. And 2) to generate HIV-1 and influenza envelop variants from flipped GC mice. Completion of this work has potential to result in both scientific and technological breakthroughs of broad impact because it is expected to define parameters enabling the extension of the power of GC evolution beyond Ig to essentially any protein-protein interaction.

NIH Research Projects · FY 2024 · 2021-09

Project Summary/Abstract Circadian rhythm disruption is experienced by patients with Circadian Rhythm Sleep-Wake Disorders and millions of shiftworkers worldwide, which may increase their risk of developing chronic health disorders including cardiovascular disease. Treatment of circadian rhythm disruption requires appropriately-timed intervention to either shift the circadian system earlier (advance) or later (delay). A Phase Response Curve (PRC) informs when to administer the intervention, without which the disruption may either be prolonged due to inadequate phase resetting or worsened due to shifting the system in the wrong direction. Currently, the field relies on the PRC for resetting the melatonin rhythm as guide to reset the entire circadian system, despite the fact that circadian rhythms are present in many other physiological features besides melatonin. Our preliminary data show that there are robust circadian rhythms in circulating levels of total cholesterol and triglyceride in healthy young individuals and that these rhythms can also be shifted. Our pilot studies further indicate that the timing of these lipid rhythms may be more responsive to shifts in the timing of meals rather than light exposure. We have constructed preliminary PRCs of these lipid rhythms in response to a combined stimulus of light exposure and meals distributed across the 24-h day and detected robust phase advances and delays. Moreover, the shifts in these lipid rhythms are larger than those for melatonin. We do not know, however, whether light exposure or meal timing is the primary time cue for resetting these lipid rhythms. Without this knowledge, developing a comprehensive treatment for circadian rhythm disruption of lipid rhythms that likely underlies the cardiometabolic consequences of shiftwork, will remain difficult. The objective of this proposal is to construct three PRCs that systematically examine the contribution of light and meal timing on resetting lipid circadian rhythms. Young healthy adults will be randomized to three conditions: (1) bright light + 12-h meal window, (2) dim light + 12-h meal window, and (3) dim light + 6.5-h meal window (time redistricted eating), each distributed across the 24-h day. The primary outcomes include phase resetting of lipid and melatonin circadian rhythms measured under each of the three conditions, and the area-under-the curve of the lipids during the 6.5-h time restricted eating. The aims of the study are to: (1) determine if light is the primary time cue for resetting melatonin but not lipid circadian rhythms, (2) determine if meal timing is the primary time cue for resetting lipid but not melatonin circadian rhythms, and (3) evaluate the acute effects of eating across the 24-h day on circulating lipid levels. Our work will be a comprehensive evaluation of how two daily events – light exposure and meals – synchronize lipid circadian rhythms in humans. We expect our analytic paradigm to be a foundational resource that can be extended to future studies of other peripheral systems under circadian regulation in humans, and have a positive public health impact by guiding therapeutic strategies for patients with circadian disruption and the population at large, many of whom experience recurrent circadian disruption due to irregular sleep-wake schedules.

NIH Research Projects · FY 2025 · 2021-09

Project Abstract Type II diabetes mellitus affects U.S. adults at varying rates across population groups. For example, individuals who identify as Latino have an age-adjusted incidence rate of 9.7 per 1,000 persons. Among those diagnosed, glycemic control and diabetes-related complications vary widely across subgroups. These patterns in health outcomes are driven by multiple interrelated contextual living conditions and access to healthcare resources. In particular, a lack of access to consistent care and difficulty navigating the healthcare system are critical drivers of these trends. By extending care beyond the clinic, telemedicine presents an opportunity to address these challenges. Telemedicine offers patients video and telephone visits from remote settings. However, current implementation strategies have created significant differences in telemedicine access and use among Latinos. The goal of this K23 is to address this important research gap by evaluating how a tailored implementation strategy can increase telemedicine use by patients with diabetes. My central hypothesis is that by applying a human-centered design approach supported by implementation science, we can improve telemedicine use, an important part of improving diabetes care. The proposed project will test the central hypothesis with the following 3 specific aims. Aim 1 will evaluate existing electronic health record data, telemedicine platforms, and workflows to identify multilevel contributors to telemedicine use patterns among Latino patients with diabetes. Aim 2 will engage participants and health system collaborators to develop a multilevel, tailored intervention targeted at patients and clinicians to promote telemedicine use in the care of Latino patients with diabetes. Aim 3 will conduct a pilot randomized controlled trial of the multilevel, tailored intervention among patients with diabetes and their clinicians. Through an innovative application of human-centered design and implementation science, this proposal addresses overlapping health information technology and variation in diabetes care outcomes among Latino patients with diabetes. The proposed research is complemented by a rigorous training plan and a highly experienced mentorship team that will ensure my transition to independence. The training plan focuses on mixed-effect logistic regression, implementation science, human-centered design, and trial design. This grant will serve as the foundation for a future R01 application evaluating the intervention’s impact on clinical outcomes.

NIH Research Projects · FY 2025 · 2021-09

Project Summary Over the past decade, genome-wide association studies have discovered complex disease-associated genetic variants while at the same time whole genome sequencing studies have been identifying risk alleles for Mendelian and complex diseases. These variants have the potential to shed light on human disease mechanisms. But there are several important challenges. More than 90% of complex disease associated variants lie within non-coding regions, posing a challenge of identifying relevant cell types and cell states, target genes, and regulatory mechanisms. The important task of linking these variants to genes itself can be challenging. In addition, as our ability to identify de novo and rare mutations for complex and Mendelian diseases is rapidly expanding, defining the function of those de novo alleles, which genes and pathways they affect remains uncertain. To address these challenges, we will predict the functional impact of disease risk variants at the level of individual variants, individual genes, and pathways to elucidate disease biology. In all aims of this proposal we will utilize IGVF functional genomic data. In Aim 1, we will predict the regulatory potential of variants in disease-critical cell types/states at a single base-pair resolution. We will identify pathogenic cell-states by analyzing single cell transcriptional data sets in a disease context, and then integrate single-cell epigenetic data to define the regulatory landscape of these rare disease cell-states. These regulatory regions identified in this analysis can be used to annotate variants for potential function. Finally, to understand functionality of specific variants in regulatory regions, we quantify selective pressure using large-scale whole genome sequencing data. In Aim 2, we will predict functional impacts of genes by effectively linking variants to genes. Defining causal diseases genes is critically important since they may be important for therapeutic targeting. We develop strategies to use genetic data and functional genomic data to predict downstream genes, and evaluate these methods with a set of gold-standard casual genes from Mendelian phenotypes. In Aim 3, we focus on rare and de novo mutations with large effect sizes. Here we recognize that predicting the function of these alleles requires an understanding of the pathways they effect, models to connect rare non-coding variants to genes, and strategies to define functionality of the variants based on population genetic parameters. In Aim 4, we develop a framework to synergize with the IGVF consortium to advance consortium goals, outlining our integration plan and flexible programmatic framework. The proposal represents a collaboration between Drs. Soumya Raychaudhuri, Alkes Price, and Shamil Sunyaev, bringing analytical expertise across functional genomics, single-cell data integration, and population genetics. These investigators have a history of successful collaborations with a strong publication records integrating functional genomics data with GWAS and sequencing studies to uncover disease mechanisms.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY/ABSTRACT This translational research proposal focuses on lysine-specific demethylase 1 (LSD1), an epigenetic regulator of gene transcription. The project’s overall aim is to show that polymorphisms in LSD1 (rs587168) are involved in blood pressure regulation and the pathophysiology of renal injury in Blacks, and that excess mineralocorticoid receptor activity mediates these effects. The applicant will address this hypothesis using a database approach (Aim 1) and a physiology-directed study in Black hypertensives (Aim 2). Specific Aim 1 will determine whether Black LSD1 risk allele carriers have greater evidence of renal damage (albuminuria) than non-risk allele carriers. The applicant will perform a cross-sectional study in 180 hypertensive Blacks (90 risk and 90 non-risk LSD1 allele carriers) from the International Hypertension Pathotype Cohort (HyperPATH) to assess whether urine albumin/creatinine levels (marker of renal glomerular and tubular damage) and Kidney Injury Molecule-1 (marker of renal tubular damage) are higher in Black LSD1 risk allele carriers vs non-risk allele carriers. Specific Aim 2 is a proof-of-principle physiologic study in hypertensive Black LSD1 risk allele carriers testing the hypothesis that reductions in blood pressure will be greater with a genetically-driven anti-hypertensive approach (mineralocorticoid receptor antagonist, eplerenone) compared to a non-specific approach (amlodipine). 56 participants will be enrolled in a 12-week randomized, double-blind, active controlled, outpatient study to assess whether eplerenone (LSD1 specific treatment) proves superior in 24-hr ambulatory systolic blood pressure reduction than amlodipine (non-specific treatment). If Aim1 is positive, the applicant will also assess change in urine albumin and KIM-1 levels in the longitudinal study. Successful completion of these Aims will document whether a genetic marker, LSD1, identifies Black individuals whose blood pressure is uniquely responsive to mineralocorticoid receptor blockade--personalized, precision medicine. Further, results of this project have the potential to reduce Black-White disparities in health outcomes secondary to poor blood pressure control. The training plan includes dedicated mentorship by Gordon Williams, MD (Mentor) and Gail Adler, MD, PhD (co- Mentor), international experts in the field of cardiovascular endocrinology. In addition to Drs. Williams and Adler, the applicant will have an advisory team composed of Bernard Rosner, PhD (statistician), Joseph Bonventre, MD, PhD (nephrologist), and Herman Taylor, MD (cardiologist), each offering expertise tailored to the applicant’s needs and goals. Also, the applicant will complete formal training in clinical/translational investigation, clinical trial design, and statistics at the Harvard T.H. Chan School of Public Health. These activities will provide the applicant with the necessary tools critical for development toward her goal of becoming an independent patient- oriented investigator in the field of cardiovascular endocrinology.

NIH Research Projects · FY 2024 · 2021-09

ABSTRACT Complex human diseases including chronic obstructive pulmonary disease and many human cancers, exhibit strong differences between males and females in risk, disease progression, and response to therapy. Our previous work in normal and disease tissues from males and females has shown that while there are rarely significant differences in genetic associations or gene expression that can explain the observed sex-based differences, modeling gene regulatory processes can lead to key insights into the likely drivers of sex differences in health and disease. This suggests that both sex-based factors and epigenetic variability likely work together to help define risk and response. This observation is further supported by the fact that sex-based differences are not static but rather evolve over the course of an individual’s lifespan, during which epigenetic state and hormonal regulation of gene expression likely change in related to gender-associated factors. In this application, we propose to merge the threads of DNA methylation variation, gene regulatory modeling, and sex-based regulatory network assessments to better understand the factors that drive the observed sex and gender-related differences in human diseases. We envision three specific aims within the context of our research plan, including: Developing tools for the inclusion of sex chromosomes in network models and methods for comparing male and female networks; Refinement of network tools for setting epigenetic priors and inclusion of DNA methylation data as a primary data for studying sex differences informed by gender; and Application to diseases with sex differences, including COPD and lung cancer as primary examples. We expand network methods to develop tools to better incorporate the effects of the allosomes in the process of gene regulation. This is particularly important as we have discovered current methods for normalization of sex chromosome gene expression can influence observed gene regulatory interactions. We will refine tools to incorporate epigenetic regulation into our sex-specific models, exploring how DNA methylation may capture gender to influence gene regulatory network structure and disease risk. Understanding the sex- and gender- related differences in this regulatory landscape will help us better understand human diseases and highlight approaches to identify sex-aware therapeutic targets. This proposal is highly responsive to the goals of RFA- OD-19-029: The Intersection of Sex and Gender Influences on Health and Disease as it will investigate a systems/network based approach to investigating sex and gender influences in human disease, as well as providing publicly available tools and networks to facilitate sex and gender research.

NIH Research Projects · FY 2025 · 2021-09

Project Summary As life expectancy increases, the burden of seizures in the elderly will increase with up to half of the cases with no identifiable cause termed Late Onset Unexplained Epilepsy (LOUE). Large databases have identified patients with LOUE as having a higher risk of dementia and stroke, but studies focusing on individual patients have been limited. There is a current need to identify the factors underlying LOUE, their impact on cognition and its natural history to develop preventive and therapeutic strategies. The first goal of this project is to determine the burden of the two most common aging pathologies in a cohort of LOUE by assessing the burden of small vessel ischemic disease using MRI and blood biomarkers of Alzheimer’s disease (AD), and how they relate to cognition and neurodegeneration. The second goal is to follow the LOUE cohort over a 3-year period to determine whether the burden of the pathologies increases when someone has seizures and leads to accelerated cognitive decline and neurodegeneration. Through a junior investigator grant funded by the American Epilepsy Society, I have started recruiting a cohort of 40 subjects with LOUE for cross-sectional analyses. As part of the proposed K23, I will expand this cohort to 100 subjects and follow it longitudinally. A control group will consist of subjects from the Harvard Aging Brian Study, an NIH funded study following cognitively normal (at enrollment)older adults with clinical assessments, fluid biomarkers, and multimodal neuroimaging, of which my mentor, Dr. Gad Marshall, has been a co-investigator since its inception. Findings from this study can be applied to other diseases where seizures are a common comorbidity such as Alzheimer’s disease. My training will rely on the mentorship of Dr. Marshall (an expert in AD and aging) and Dr. Page Pennell (an expert in epilepsy), as well as an advisory committee of world leaders in the fields of AD biomarkers (Dr. Dennis Selkoe), neuroimaging (Dr. Steven Stufflebeam), neuropsychology (Dr. Rebecca Amariglio), vascular related cognitive impairment (Dr. Anand Viswanathan), and statistical analysis of longitudinal studies (Dr. Joseph Locascio). During the training period, I will gain expertise in neuroimaging analysis, the implementation of longitudinal studies in an elderly cohort, and the statistical analysis of longitudinal data. The institutional resources available through Brigham and Women’s Hospital, Massachusetts General Hospital, Harvard Medical School, and Harvard School of Public Health are world class and will support my career development in an environment that can foster high impact contributions. Upon successful completion of the project, I will be well positioned to launch a career as an independent investigator examining the interactions of seizures, neurodegeneration, cerebrovascular disease and cognition.