Boston Children'S Hospital

NIH Research Projects · FY 2025 · 2022-09

Endometriosis-associated pain is an important driver of opioid use in women. Endometriosis is and estrogen sensitive, angiogenesis dependent disease that affects ~10% of women of childbearing age and is found in half of women with chronic pelvic pain. Endometriosis increases the likelihood of chronic opioid use, opioid dependence/abuse, and opioid overdose. Endometriosis is currently treated with NSAIDS, hormonal therapy targeting estrogen production, and surgery, but these options are not durably effective for ~30% of patients. Thus, new targets are needed. CMG2 is an integrin-like extracellular matrix receptor that we have shown regulates angiogenesis. It is also overexpressed in endometriosis tissue vs. normal endometrium. In mouse models, published work and our preliminary data show that treatment with CMG2 antagonists that we discovered (PGG, PGM) decreases lesion incidence, growth, and endometriosis-associated pain. These observations suggest that CMG2 may be a useful target for the treatment of endometriosis-associated pain. However, key gaps in our understanding of CMG2 biology would slow any development program with CMG2 as a target. The first key gap is that the cell type whose targeting resulted in decreased pain upon CMG2 antagonist treatment is unclear. Second, the molecular mechanism underlying CMG2 signaling in endometriosis is not known. Finally, both the maximum efficacy and, importantly, safety of CMG2 targeting is not known. We propose to fill these gaps by using cell-type-specific CMG2 knockout mice to identify cell types where CMG2 expression supports endometriosis lesion growth and pain. Because CMG2 lacks any catalytic domains, we hypothesize that CMG2 signals via differential protein interaction. Then, expression of CMG2 in appropriate cells in human lesions will be confirmed. Next, we will use proximity proteomics to identify downstream molecules that differentially interact with CMG2 ±inhibitor. Candidate mediators of CMG2 signaling will then be confirmed using CRISPR knockout and cell-based assays of CMG2 function. Finally, we will evaluate safety and efficacy of CMG2 targeting in a mouse model using a well-characterized, high- specificity CMG2 inhibitor. Throughout ex vivo and in vivo assays, RNAseq and scRNAseq will be used to identify transcriptional signatures of CMG2 blockade, both to validate in vitro assays and to generate potential pharmacodynamic markers of CMG2 inhibitors. Completion of the proposed work will validate CMG2 as a target for the treatment of endometriosis- associated pain. It will also provide key insights into endometriosis pathophysiology that will enable the generation of novel therapeutics and may identify additional targets for treatment of the disease. Development of such drugs will improve the lives of many women and decrease the use of opiods to treat this disease, thereby improving many lives.

NIH Research Projects · FY 2025 · 2022-09

Project Summary The goal of this proposal is to dissect the mechanisms of self-DNA detection by the enzyme cyclic GMP-AMP synthase (cGAS), and to determine the relative contribution of its diverse signaling activities to inflammation in the microenvironment of implantable murine tumors. cGAS operates in virtually all cell types as a DNA sensory protein, which synthesizes the second messenger cyclic GMP-AMP (cGAMP) upon binding DNA. This second messenger stimulates interferon (IFN) and inflammatory activities via the downstream protein STING. Because cGAS detects the sugar-phosphate backbone of DNA, a major question relates to how this enzyme is regulated to ensure self-nonself discrimination. This question relates to much fundamental biology and the answer will impact the increasing number of clinical endeavors that target the cGAS-STING pathway. The cGAS-STING pathway is oft-discussed in the context of antitumor immunity, but the activities of this pathway that are beneficial (or not) to immunity remain unclear. For example, inflammatory activities induced by cGAS-STING in the tumor microenvironment (TME) have been reported to induce protective inflammatory and cytolytic CD8+ T cells. But cGAS-STING signaling events have also been reported to promote tumor growth and disease progression. A central idea that drives our work is that the ubiquitous presence of the cGAS-STING pathway in most cell types, along with its diverse signaling effectors (cGAMP, IFNs, cytokines), can create a complex TME prone to unpredictable outcomes (e.g. disease resolution or progression). In order to understand each activity of this pathway, new experimental tools are needed to disentangle its effector functions. Herein, we describe new synthetic biology-based genetic circuits that can dissect the effector functions of cGAS-STING, within cancer cells specifically. Notably, these systems led to the discovery of specifies-specific differences in the ability of primate and murine cGAS proteins to detect self-DNA. This finding raises questions of the suitability of mice and certain primates as accurate preclinical models for cGAS-STING function, and provide a mandate to define the mechanisms and consequences of cGAS-mediated self-DNA activities. The work in this proposal is based on the hypothesis that the cGAMP, IFNs and cytokines induced by the cGAS-STING pathway play differential roles in tissue inflammation and immunity, and that understanding the role of each of these activities, within specific cell types, requires a detailed characterization of the mechanisms of self (and nonself) DNA detection. To address this hypothesis, we propose to determine how distinct intra-tumoral cGAS activities influence protective T cell immunity (Aim 1). In Aim 2, we propose to determine mechanisms of self-DNA reactivity by human cGAS through comparative analysis of the human, mouse, chimpanzee and orangutan proteins, each of which display distinct means of self-DNA reactivity.

NIH Research Projects · FY 2026 · 2022-09

PROJECT SUMMARY/ABSTRACT Rare diseases are individually rare yet are collectively common and affect millions of patients and their families in the United States. To date, most efforts for clinical decision support systems (CDSS) for rare diseases are aimed at deriving diagnoses from genomic data. Although those tools can be critical for genetic specialists, it is essential for primary care providers to identify suspected patients and make appropriate referrals at an early stage to get their genomic testing. In this study, we propose a SMART-on-FHIR based Rare Disease Detection and Escalation Support (RESCUE) CDSS. It will use a centralized informatics approach to identify suspected rare disease patients from clinical data warehouse (CDW) and send alerts to physicians with escalation support including phenotype summarization, genetic/genomic test requisition and research opportunity discovery. We will leverage the cutting-edge natural language processing technics to unlock the clinical features in the clinical narratives and convert them to the GA4GH-based standard for optimized genetic disease information exchange. To overcome the challenge of insufficient knowledge and limited sample size for a single rare disease, we propose a hybrid approach to address this challenge. Expert-curated and knowledgebase-derived phenotype- based queries will be issued to identify “silver-standard” cases first, and then a neural network-based analytical model will be trained to further identify potential undiagnosed rare disease patients. We will investigate the efficiency and accuracy of this analytical model by conducting chart-reviews using the clinical data warehouse at the Columbia University Irving Medical Center (CUIMC). To ensure its generalizability, we will further validate the model using Children’s Hospital of Philadelphia as the secondary site. The proposed RESCUE CDSS will apply this analytical model to identify suspected rare disease patients and notify physician by triggering a provider-facing SMARTapp during the patient encounter. By engaging patients (and their families), primary care physicians, genetic specialists, researchers, key policy makers and ELSI experts, our stakeholder-centered participatory approach will ensure that the design of this SMARTapp is interoperable, comprehensible, and actionable. We will collaborate with Epic IT teams to deploy RESCUE and evaluate its usability in ambulatory pediatrics clinics at CUIMC. To optimize the performance and to increase interoperability with various EHR systems, the CUIMC validated software will be further deployed and validated at CHOP. We will further extend RESCUE to enable an enhanced escalation support including genetic/genomic testing support and patient- research matching, and retrospectively validate the extended module. Our transferable software and tools developed in this study will be shared through multiple outlets including CTSAs, the eMERGE network, and the Observational Health Data Sciences and Informatics (OHDSI) community.

NIH Research Projects · FY 2024 · 2022-09

Primary Immune Deficiency (PID) is a debilitating condition that affects one in 1,200 persons in the US. Growing evidence demonstrates that PID is underdiagnosed in the US and globally. Of note, prior to the implementation of population-wide newborn screening, severe combined immunodeficiency (SCID) was estimated to affect approximately 1 in 100,000 infants, with ~90% of identified cases occurring in non-Hispanic Whites3. Following the introduction of newborn screening, the observed incidence of SCID nearly doubled and cases were found to occur at similar rates across different ethnic groups5, suggesting significant disparities in diagnosis prior to universal screening. Underdiagnosis of PID is not confined to SCID, most of which cannot yet be detected through newborn screening. Delay in the treatment of PID can result in serious health consequences, including irreversible organ damage and death. There is therefore an urgent need to address gaps in the diagnosis of PID. Our long-term goal is to improve timely diagnosis and treatment of PID. To address disparities in the diagnosis of PID, we propose the following specific aims: (1) Identify patterns of diagnostic disparities in PID; (2) Identify factors contributing to diagnostic disparities in PID; and (3) pilot a targeted educational intervention aimed at increasing awareness of PID and the risk factors associated with its underdiagnosis. We will combine analyses of electronic health record (EHR) data (Aim 1) with qualitative analysis of patients’ experience and real-world perspectives from healthcare providers to understand the contributing factors of delayed PID diagnosis (Aim 2). Additionally, we will apply advanced machine learning analysis as an innovative approach to enable a more comprehensive understanding of the patterns of diagnostic delay. PID is a complex group of diseases with highly variable clinical manifestations. We anticipate that the application of machine learning methods to EHR data can facilitate identification of under-recognized patterns of diagnostic delay and will enable us to learn from large clinical datasets in a scalable manner. Integrating knowledge from these analyses, we will then develop and evaluate an educational outreach program targeting healthcare providers to raise awareness of PID and the risk factors associated with its underdiagnosis (Aim 3). The study will be conducted at 2 major healthcare systems in Massachusetts: Beth Israel Lahey Health and Boston Medical Center. This body of work represents the first systematic effort to investigate the patterns and contributing factors of diagnostic disparities in PID. The proposed educational outreach program will be the first educational initiative aimed at addressing PID diagnostic disparities in the US, and will provide a foundation towards our longer-term goal of designing and developing a regional/national educational program to improve diagnosis of PID.

NIH Research Projects · FY 2025 · 2022-09

Project summary Aging in humans is associated with a host of brain diseases, including tumors, age related neurodegeneration, and Alzheimer’s Disease (AD). In many tissues, the aging process leads to a derepression of transposable elements (TEs) that lead to inflammation and cell senescence. Several recent studies have demonstrated that multiple subfamilies of TEs are expressed at higher levels in postmortem AD brain than healthy controls as a direct result of the accumulation of mis-folded Tau proteins characteristic of AD pathology. Many of the hallmarks of AD, including neuroinflammation, heterochromatin remodeling, genomic instability, and the recently implicated T-cell infiltration, can be triggered by the activation of TEs. However, because of the unique epigenomic landscape of different cell types in the brain combined with the dependence of TE mobilization on cell division and other factors, the dynamics and consequences of TE activation are likely cell-type-specific. This study aims to characterize the activation of TEs in the brain during aging and AD and identify the functional consequences of activated TEs at the level of individual cells across multiple cell types and to develop the novel computational tools necessary to answer these questions. The first aim will establish the genomic and epigenomic changes related to TEs during normal aging. This analysis will help both to determine whether TEs are involved in the normal brain aging process and age-related neuronal decline, and to establish the healthy controls for comparison with AD. The second aim will perform similar analysis, this time focusing on AD and including the separation of neurons with and without accumulation of pathogenic Tau. The final aim will measure the transcriptome at the single-cell level for all the samples profiled in the first two aims. This sequencing will allow both the measuring gene expression related to innate and adaptive immune responses and the identification of TE derived sequences capable of triggering those responses. The experimental tools and sequencing technologies now exist to examine these questions, and this study is designed to determine how TE activation impacts normal brain aging and AD. Understanding the relationship between TEs, neuroinflammation, and AD pathology may open the door for new treatments and cures.

NIH Research Projects · FY 2025 · 2022-09

(PLEASE KEEP IN WORD, DO NOT PDF) Enter the text here that is the new abstract information for your application. This section must be no longer than 30 lines of text. The hematopoietic system has been, arguably, the best well studied tissue from a developmental and stem cell biology perspective. Historically, the study of hematopoiesis has taken advantage of flow cytometry and cell transplantation approaches as the gold standard assays to examine functional behaviors. While these studies have provided an elegant textbook view of hematopoiesis, we argue here that our knowledge of how early blood formation works is largely incomplete. This is predominantly due to the limitations of utilizing traditional assays and readouts that disrupt the normal marrow tissue archicture and neglect the anatomical localization of cells. Thus, one key biological component missing in many studies in the field is that of spatial organization. Basic questions such as when and where hematopoietic fate commitment occurs, what the exact cellular intermediates are between a stem cell and mature daughter cells, and what the identity and role of the cellular microenvironement (niche) is in the hematopoietic process remain largely unsettled. Here, we propose an interdisciplinary approach based on state-of-the-art live microscopy manipulations in combination with novel hematopoietic fate reporters to enable a comprehensive study of hematopoiesis entirely in situ. Using novel two photon-guided collection of stem cell clones directly from the bone marrow, and longitudinal live-imaging analysis of stem cell progenies, we aim to generate a spatial, temporal and molecular map of hematopoiesis at single cell resolution. Complementary proximity-based photolabelling approaches will be taken to catalogue the cellular composition of the niche in the steady-sate, diseased, and aged settings. By moving away from tissue disruptive approaches to a system that relies on observation, manipulation, and analysis of single hematopoietic cells within the bone marrow, we hope to obtain unique insights into the cellular organization of blood production.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT Mutations in the SLC13A5 gene, which encodes a plasma membrane citrate transporter, result in a newly diagnosed form of genetic epilepsy termed early infantile epileptic encephalopathy, which is characterized by multi-focal seizures in neonates. These infants subsequently develop cognitive and behavioral deficits. Human genetics has identified both commonly occurring missense and deletion mutations, but it is not known how distinct genetic mutations affect disease presentation and seizure severity. We are addressing this question by characterizing an array of SLC13A5 mutant mouse models carrying: i) ablation of its endogenous murine Slc13a5 gene (knockout), ii) the most common patient mutation, the G222R point mutation (equivalent to the human mutation G219R), and iii) the second most common patient mutation, the T230M (equivalent to the human mutation T227M). Our preliminary data demonstrates that homozygous Slc13a5 knockout mouse demonstrates abnormal epileptiform electroencephalogram (EEG) profiles, while the G222R has seizures and significantly more severe epileptiform activity. Preliminary histopathology reveals differential interneuron reduction between the knockout and G222R, with the G222R additionally showing oligodendroglial loss. These results are consistent with our interpretation that specific missense mutations may acquire dominant gain-of-function effects which exacerbate vulnerability and neuronal hyperexcitability. The knockout has shown reduced excitatory and inhibitory postsynaptic activity suggestive of reduced neurotransmitter levels. We will further characterize these mutant mouse models with the following experiments: experiment 1.1, the characterization of interictal discharges and seizures in the mutant alleles using EEG, in parallel with analysis of brain histopathology. In experiment 1.2, I will apply electrophysiological techniques to investigate whether a deficiency in intracellular citrate transport in Slc13a5 knockout and missense mutations affects neuronal function by altering glutamate and GABA concentrations. A metabolomics screen for neurotransmitters will be performed to corroborate functional data. In experiment 1.3, I will investigate the effect of these SLC13A5 mutant proteins on cellular pathologies. Combined, these experiments will investigate the molecular and cellular mechanisms that contribute to the seizure phenotype in the SLC13A5 disease, which in turn will inform therapeutic approaches. I will acquire training in electrophysiology and molecular biology to follow through with these experiments. Our research training plan and ongoing professional development will provide me with skills to transition me to the next stages of my scientific career. In the postdoctoral stage, I plan to gain experience with gene therapy strategies in experimental models of nervous system disease. I will continue to investigate cellular and molecular mechanisms underlying brain disease, and to apply these findings to optimize gene therapy approaches. Furthermore, I will continue to develop the professional skills required to become an independent primary investigator at an academic research institution.

NIH Research Projects · FY 2025 · 2022-09

SUMMARY Deployment of new antimicrobials is promptly circumvented by the rapid evolution of resistance, underscoring the critical need for new strategies to stay ahead in the arms-race against bacterial pathogens. Developing a detailed understanding of the circumstances as well as genetic and mechanistic basis for which antibiotic resistance develops provides opportunities for pre-emptively subverting this process. While infections caused by organisms harboring antimicrobial resistance (AMR) genes are a major cause of antibiotic treatment failure (ATF), ATF frequently occurs when the etiological agents are not AMR by traditional susceptibility testing. It is becoming increasingly recognized that transient cell-states such as tolerance, persistence and hetero-resistance are critical drivers underlying treatment failure. However, there is a paucity of data with regards to the genetic and mechanistic basis for these cell-states as well as a lack of diagnostic-detection approaches. ATF cell-states initially exist as minority variants within a population and display a transient phenotype that tends to dissipate as the stress subsides, making them challenging to detect and consequently missed in current diagnostic assays. These enabler cell-states remain mechanistically poorly understood and seem to preferentially arise during fluctuating treatment regimens, for instance caused by a drug’s PK/PD characteristics, whereby ATF cell-states can drive the re-emergence of the (susceptible) bacterial infection after antibiotic pressure wanes. Importantly, this creates opportunities where multi-step high-level resistance mutations are given an extended opportunity to emerge. Therefore, because antibiotic resistant variants often follow closely on the heels of the occurrence of ATF cell-states, these cell-states can be viewed as enablers of antibiotic treatment failure and AMR. This proposal focuses on untangling the importance of ATF cell-states in the emergence of antibiotic resistance and treatment failure, and designs new approaches and strategies to identify, track and target them. The main team consists of 4 principal investigators that have a very successful collaboration history. Together they will work on 5 challenges distributed across 3 projects and supported by an administrative and a genomics and bioinformatics core. In Challenge: 1) the full profile of possible genetic pathways that can induce ATF cell-states is determined; 2) treatment regimens that drive the emergence of ATF-cell states are determined; 3) it is determined how ATF cell-states enable the emergence of AMR; 4) drugs and compounds are screened for, that target ATF cell-state collateral sensitivities; 5) a computational deconvolution approach is developed that predicts the presence and frequency of ATF cell-states in a complex bacterial population. Overall this proposal contains a collection of conceptually and technically innovative aspects that are geared towards understating the genetic mechanisms and evolutionary forces that sit at the root of the emergence of resistance, with the ultimate goal to design new diagnostics and antimicrobial strategies that can slow or even stop the current endless arms-race “that takes all the running we can do, to keep in the same place”.

NIH Research Projects · FY 2025 · 2022-09

Project Summary/Abstract Background. The purpose of the proposed research is to test the feasibility and safety of inhaled hydrogen gas (H2) administration as a rescue therapy during cardiac arrest requiring extracorporeal cardiopulmonary resuscitation (ECPR). Among patients with congenital heart disease (CHD) receiving ECPR, 52% either die or suffer severe neurologic impairment. Diatomic hydrogen (H2) chemically reduces toxic oxygen mediators that directly damage cellular structures, and has been shown to improve neurologic and renal function in a number of preclinical ischemic injury models. Recently, we have demonstrated that inhalation of 2.4% H2 for up to 72 hours is safe in healthy adults. However, H2 has never been applied to the ECPR population. Study design. We propose an early-phase, two-site, randomized trial of H2 in ECPR patients entitled the ‘Hydrogen FAST Trial’ (Hydrogen’s Feasibility And Safety as a Therapeutic agent). Key Inclusion criteria are (1) patients with CHD (broadly defined, including myocarditis, channelopathies, transplant rejection), (2) experiencing a cardiac arrest >5 minutes and receiving ongoing CPR, and (3) a decision made to resuscitate using ECPR. The trial will be led by MPIs John Kheir, MD and Lynn Sleeper, ScD, who have a strong collaborative history in this area and complementary expertise in critical care, translational research, and clinical trials. Furthermore, the trial design has received favorable feedback from the FDA. During the R61 Phase: (1) Regulatory (IRB and FDA) approvals will be obtained, including exception from informed consent (EFIC) requirements. This is necessary because the emergent nature of ECPR and time- sensitivity of H2 preclude traditional informed consent. Instead, non-opt-out patients will be randomized and enrolled emergently, with subsequent traditional informed consent. (2) Investigational product (IP) manufacturing, storage, and administration logistics will be established. (3) Endpoint adjudication processes and DSMB infrastructure will be established. (4) Study roll-out and clinical staff education will be completed, followed by a 3-patient vanguard phase (Phase 0) trial. During the R33 Phase, 53 patients with CHD will be randomly assigned in a 3:2 (32/21) ratio to either usual care plus 2.4% H2 gas following ECPR for 72 hours or to usual care from two sites. Primary endpoints include measures of feasibility and safety (rate of treatment-associated severe adverse events, adjudicated by an independent committee blinded to treatment group). Secondary endpoints will include functional status, brain biomarkers and cranial images obtained using a new point-of-care MRI device. Impact. The experienced team and infrastructure in place for this trial will ensure the successful completion of R31 and R466 milestones. If H2 therapy is shown to be feasible and safe, this work will provide a foundation for H2 administration in the critical care environment, and subsequent testing in cardiac arrest, stroke, myocardial infarction, and other disorders in which ischemia-reperfusion injury plays a role.

NIH Research Projects · FY 2024 · 2022-09

Project Summary Heart failure is the leading cause of death in the world. At the core of the pathophysiology of heart failure is the inability of the adult mammalian heart to regenerate following injury. In contrast to adults, the newborn mouse heart is capable of complete regeneration following various types of injury, providing a new inroad into possible mechanisms of cardiac regeneration and repair. Previously, we discovered that Nuclear Factor Erythroid-derived 2-related factor 1 (NFE2L1, also known as NRF1) is highly expressed in a regenerative cardiomyocyte population in neonatal mouse hearts. Although NRF1 does not directly promote cardiomyocyte proliferation, it confers strong protection to cardiomyocytes under stress conditions both in vitro and in vivo. Recent studies show that NRF1 regulates proteostasis and redox balance in multiple tissues in response to cellular stress, while its cardiac functions are largely unknown. This proposal outlines a comprehensive plan to further dissect the biological and pathological functions of NRF1 in cardiomyocytes to provide critical insight into the protective mechanism that underlies regeneration and stress adaptation in the heart. In this research plan, Aim 1 will establish how NRF1 protects neonatal cardiomyocytes. Aim 2 will demonstrate the function of NRF1 in mammalian neonatal heart regeneration. Aim 3 will uncover the role of NRF1 as a stress regulator in adult hearts following ischemic injury. In the mentored phase, Aims 1 and 2 will be carried out in the laboratory of the renowned molecular biologist Dr. Eric Olson, and will generate data and mouse knockout models for continued investigation in the independent phase. Aim 3 will be initiated during the K99 phase and continued during the independent phase, dedicated to investigating the therapeutic potential of NRF1 to treat adult ischemic heart disease. In order to gain research independence through mentored training, I will continue to develop expertise in applying transcriptome profiling, including single-nucleus RNA sequencing, to study neonatal heart regeneration (Aims 1 and 2). Such an experience will complement my prior training and constitute an important data generating platform throughout my career. The proposed research requires that I acquire additional mentoring in adult cardiac injury models and related cardiovascular phenotyping (Aim 3). Investigation of NRF1 and its downstream pathways in regulating the stress response of cardiomyocytes during this mentored training will lead to the establishment of an independent niche for my own academic career. In summary, this application will provide me with the scientific training, mentoring, and career development necessary as I transition to independence. Most importantly, the collective body of work generated through the completion of the proposed aims will make major contributions to the field of cardiovascular science.

NIH Research Projects · FY 2025 · 2022-09

Over the last 30 years, a new field––known as computational epidemiology (comp epi)––has emerged at the intersection of digital data streams (e.g., news and social media, search query, and mobility data), machine learning (e.g., nonlinear optimization, natural language processing, and agent-based modeling), and public health crises. Due to the ongoing SARS-CoV-2 pandemic, as well as other pathogens that have resurged in the United States (e.g., mumps virus), comp epi has shifted part of its focus as a field to improving public health decision-making during outbreaks and epidemics of infectious disease. In this proposal, we present four foundational challenges within the context of infectious disease research and comp epi more broadly. While the first three of these challenges are more conventionally scientific in nature, the fourth involves scientific community-building: (1) estimating the time-varying transmissibility (i.e., the effective reproduction number, REff) of a given infectious disease; (2) real-time monitoring and measurement of transmissibility-relevant behaviors (e.g., human mobility, etc.); (3) forecasting of outbreaks and epidemics as a function of individual health behaviors; and (4) recruitment of new scholars to the yet-insular field of comp epi. To address these challenges, we propose the development of (1) a meta-analytical tool for ensemble estimation of REff across multiple research groups; (2) a surveillance system to monitor transmissibility-relevant behaviors, including an inference system to produce more representative measures for human mobility; (3) a generalizable agent-based model for epidemic forecasting that features behavioral parameters, as informed by the aforementioned surveillance and inference systems; and (4) a virtual laboratory for comp epi scholars to collaborate on infectious disease research across institutional silos. By addressing the first three challenges, we hope to help clinicians and public health policymakers make data-informed decisions during infectious disease crises while simultaneously providing opportunities for other public health researchers to augment their own efforts in transmissibility estimation and epidemic forecasting by harnessing expected products from our proposed research. Meanwhile, by addressing the fourth challenge, we hope to help new scholars form meaningful collaborations both with pioneers in comp epi and with each other, while simultaneously promoting growth of the field as we move forward.

NIH Research Projects · FY 2026 · 2022-09

SUMMARY Age-related decline in vascular function is a key factor in decreased organ function, vitality, resistance to stress and increased morbidity/ mortality. Our laboratory has made long-standing contributions about how lipid mediators (sphingolipids and prostanoids) enhance vascular endothelial cells (EC) function and resilience to pathological processes. Our recent data suggest that age-dependent decline in EC protective sphingosine 1-phosphate (S1P) signaling contributes to EC dysfunction and pathology of various organs. Specifically, circulatory HDL-bound S1P signaling via EC S1P receptor-1 (S1PR1) is a key mechanism that enhances EC resilience and that therapeutic strategies designed to counter its age-dependent decline was efficacious in reducing organ pathology. This premise is further supported by unbiased studies in humans which show age-dependent decrease in ApoM, the S1P chaperone on HDL. We hypothesize that aging compromises vasculoprotective S1P pathway which enhances EC resilience, thus contributing to rapid decline in organ function. The corollary of the hypothesis is that mechanism-based therapeutic enhancement of EC S1PR1 pathway will decrease the rate of decline of organ-specific EC. To understand organ-specific vascular aging mechanisms, we profiled the transcriptome and chromatin regulatory sites globally from freshly isolated EC from normal mouse aorta, lung and retina. EC defects in these organs lead to atherosclerosis, reduced resistance to viral infections and vascular pathology in central nervous system (CNS), respectively. First, we will characterize an aging-induced aortic endothelial cell-2 (AEC2) population which has attenuated S1PR1 signal while exhibiting inflammatory and fibrotic gene signature. We will define age-associated chromatin regulators of AEC2 cells and assess the ability of S1PR1/ Gi-biased signals to counteract this pathological EC phenotype change in aged mice. Second, we will test the hypothesis that EC S1PR1 signaling in lung EC (LEC) supports resilience against viral infections. Mechanisms by which aging attenuates LEC S1PR1 signaling will be elucidated. Therapeutic strategies that mimic HDL-S1P that suppress pathological phenotypes will be tested. Third, using the retinal EC (REC) as a model of the CNS vasculature, we will examine whether S1PR1 signal can counter age-related barrier breach, transporter gene expression and astrogliosis either alone or in combination with the Wnt signaling activators. These studies are anticipated to lead to new insights by which lipid mediators contribute to vascular dysfunction and disease during aging, which could ultimately lead to novel therapeutic strategies to combat age-related organ dysfunction and decline.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT Survivors of childhood and adolescent cancer (“survivors”) face high risks for early mortality and treatment- related late effects, including subsequent breast cancer. Approximately 30% of female survivors previously treated with chest radiation will develop breast cancer before age 50, a risk similar to BRCA1 mutation carriers. The Children’s Oncology Group recommends early screening for breast cancer (in those who received radiation) starting at age 25, but <15% of survivors are adherent, many because of economic barriers. Risk- reducing medications, such as selective estrogen receptor modulators and aromatase inhibitors, could allow some survivors to avoid breast cancer entirely (vs. early detection with screening and treatment) since these drugs reduce the risk of estrogen receptor positive (ER+) tumors by 50% and are recommended for high-risk women by the U.S. Preventive Services Task Force. However, risk-reducing medications are not currently standard care for high-risk survivors which includes not only those previously treated with radiation but given emerging data, those who were exposed to high doses of anthracycline chemotherapy. The rarity of childhood and adolescent cancer and the long latency needed to capture subsequent cancers limit the feasibility of prospective prevention trials. Addressing all RFA-CA-20-027 priority areas, we propose to use simulation modeling and longitudinal observational data to inform clinical care by evaluating the clinical benefits and harms of risk-reducing medications among childhood and adolescent cancer survivors. We will build on our established collaboration with the Cancer Intervention and Surveillance Modeling Network (CISNET), the Childhood Cancer Survivor Study (CCSS) and the St. Jude Lifetime Cohort Study to: (1) refine two CISNET models to reflect current knowledge on breast cancer risk, screening and prevention among survivors; (2) provide model results in readily accessible online look-up tables summarizing benefits (e.g., avoiding breast cancer), harms (e.g., medication side effects) and costs to women and society of adding risk-reducing medication use for 5 years to screening; and (3) conduct exploratory analyses to assess the impact of risk- reducing medications and screening on outcomes by race. Our proposed research will have high potential to reshape current paradigms for survivorship care, using breast cancer risk-reducing medications as an example for understanding how preventive agents can be incorporated into current survivorship recommendations and practice. This work will also provide a framework to illuminate key elements and intervention points that can guide efforts to minimize disparities. This project is conceptually innovative and clinically important by simultaneously considering proven effective primary cancer prevention medicines together with screening recommendations. This work will be highly translational by providing data to inform clinical care guidelines and create resources that survivors and care-providers can use to guide care discussions on breast cancer prevention and early detection.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Experiments in this proposal address how sensory signals trigger effective action. We focus on light-driven modulation of the mammalian pupil, which merits study for its own sake and offers a tractable system for understanding the steps that lead from photon capture to motor output. The pupillary light response appears simple but is critical for vision; a large pupil increases photon collection to support sight in dim light, while a small pupil reduces optical aberration to sharpen visual acuity in bright light. The pupil mediates this trade-off across variations in environmental light intensity that span orders of magnitude, and does so in a manner that appears optimal. We propose to investigate how features of molecules, cells, and circuits in the retina meet the requirements of pupil control. Our overarching hypothesis is that these mechanisms are well-tuned, to the extent that their features propagate through brain circuits to manifest overtly in the pupil. To test this hypothesis, we will apply in vivo approaches to mice, analyzing retinal signals within the brain’s first relay for pupillary control while simultaneously monitoring the pupil. We will draw on our knowledge of retinal mechanisms to examine their actions in these areas, using quantitative and systematic experiments. We will examine mice that have normal visual pathways or are engineered to lack candidate mechanisms. Moreover, we will employ ex vivo methods to clarify select mechanisms, such that we can analyze their in vivo influences with greater precision. Taken together, these experiments will uncover origins of the pupillary light response, inform the question of how sensory information affects motor action, and provide insight into the steps by which mechanisms at lower levels of biological organization influence the whole animal.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Many Americans (mostly children) have a genetic disease so rare it is termed an “orphan disease” with no approved treatment and little incentive for investment in therapy given the rarity. However, it is now possible to design, develop, and deliver gene-targeted treatments that work for as few as a single patient, i.e., as truly individualized medicines. These “n-of-1” treatments began with a class of drugs called “antisense oligonucleotides” (ASOs), first demonstrated in 2018 when a customized ASO was designed to target a specific pathogenic genetic variant on behalf of a child with a fatal and otherwise untreatable genetic condition. This effort created a blueprint for treating other individuals with orphan diseases. Not surprisingly, that pilot case brought forth a multitude of hopeful families asking about their children’s eligibility for similar interventions, and at least six academic institutions have launched efforts in this space to develop additional individualized n-of-1 therapies. The development of customized investigational therapies for single or few individuals is at present expensive, both in terms of cost and time, and raises a host of ethical, legal, and social implication (ELSI) challenges, including therapeutic misconception, hope-therapeutic optimism, informed consent, experimental treatment of children unable to consent or assent, best interests of the child, and appropriate thresholds of evidence for safety and efficacy when dealing with fatal orphan diseases that lack other treatments. There is a critical need to gather expert and stakeholder input to address these considerations and provide guidance, not only for sake of those interested in individualized ASO development, but for other emerging gene-targeting therapeutic platforms that might be similarly individualized (e.g., genome editing). The goal of this study is to develop and deliver empirically-informed guidance that addresses the complex ELSI of individualized genomic medicine, and to chart a course that is fair, transparent, and responsible. In Aim 1 we will conduct qualitative interviews with ASO Site teams involved in the development of individualized therapies, Societal Issue experts (e.g., sociologists, anthropologists, economists, bioethicists), Parents of children with and without genetic conditions, Oversight experts without n-of-1 ASO experience, and representatives of foundations and patient advocacy. In Aim 2, informed by our experience and combined with domains and themes identified in Aim 1, we will combine a case-based modified Delphi process, capped by a roundtable session to develop two tiered guidance for addressing the ELSI challenges attendant to individualized therapy: 1) recommendations (“overall consensus”) and 2) points to consider (key issues below the pre-determined threshold of “overall consensus”), along with a source casebook. The two tiered guidance will inform evolving policies around the provision of individualized genomic medicine for orphan diseases. Findings and recommendations will be broadly disseminated in a half-day conference, as well as global professional meetings and in peer-reviewed journals.

NIH Research Projects · FY 2024 · 2022-08

In this study, we seek to improve assessment of critical safety and relationship skill domains by developing and validating a novel measurement battery. Sensitive and objective tools that assess these domains are extremely limited. This lack of established measurement tools poses a critical challenge to studies of the efficacy and effectiveness of important safety interventions and relationship skill protocols, especially for populations uniquely vulnerable to negative outcomes, such as adults on the autism spectrum (AA). The financial, emotional, and physical costs of failing to adequately prepare individuals to navigate relationships safely and effectively has been demonstrated across many diagnoses. Effectively measuring related behaviors—such as navigating public/private boundaries, recognizing when others are interested in a relationship, determining the trustworthiness of potential partners, judging situations for safety, using indirect communication, and taking the perspective of others—is a critical task needed to accurately characterize the challenges of AA and other adults with social difficulties and to track progress both over time and in response to adult social skills interventions. We propose a battery of performance-based tools that target these skill domains via behavioral, eye tracking, and psychophysiological responses to social information. The strength of this battery is the objectivity of the chosen tools (e.g., behavioral responses, tracking eye movements, and heart rate) and their ecological validity (i.e., mimicking everyday experiences such as using the internet, navigating dating apps, and engaging in various types of relationships). The total length of the proposed project is three years. The measurement battery will be presented at two time points (3 months apart) to a total of 200 adults (100 AA, 100 adults without autism). We have three aims: (1) To examine the feasibility of acquisition of the proposed measures for AA; (2) To assess the utility of the measurement battery for assessing social difficulties of the AA and non-ASD groups, and (3) To validate the measures for use in clinical trials by examining (3a) their short term stability, (3b) convergent validity with a measure of general social functioning, and (3c) demonstrate their utility as measures of target engagement by confirming their correlation to outcome measures of romantic and sexual functioning. Achieving these aims will provide a novel toolkit for investigators to use to measure social cognitive, behavioral, and physiological changes in response to intervention programs for AA and, ultimately, populations with similar safety and relationship skills deficits (e.g., intellectual disability, ADHD, social anxiety, schizophrenia).

NIH Research Projects · FY 2026 · 2022-08

Chronic pain experienced by children has the potential to persist into adulthood and drive drug addiction, mental health problems and suicidal behavior. The level of pain reported by children who experience headache is immense and likely poorly estimated based on lowered ability of children to articulate symptoms, potential malingering or underreporting of pain symptoms, and extensive variability in physical growth and brain development. Headache is a frequently reported and poorly understood primary and secondary disorder in pediatric subjects. The diverse presentation of headache underscores the potential for distinct mechanisms of headache presentation, thus placing emphasis on tailored treatment options. There is clear need to better define childhood headache in terms of the clinical presentation and underlying pain physiology. Our hypothesis is that each clinical headache disorder can be defined by unique biobehavioral characteristics. In the proposed research program, we (Aim 1) evaluate the clinical and behavioral elements of each headache disorder and pain modulation, (Aim 2) the biobehavioral signature of treatment refractory headache, and (Aim 3) develop machine learning classifiers to understand the features that differentiate headache subtype and treatment resistance. This study is likely to yield highly relevant information that will contribute towards identifying and treating headache disorders in children, identifying unique characteristics of headache and pain processing, and (3) outline biobehavioral targets for different headache disorders. Data from this investigation is likely to contribute greatly towards the treatment of pediatric pain disorders.

NIH Research Projects · FY 2024 · 2022-08

ABSTRACT Syndromic disorders account for a large proportion of the rare genetic disorders that impact the pediatric population, including many conditions with structural birth defects and other congenital anomalies. These conditions are a disproportionate cause of morbidity and mortality. Accurate molecular diagnosis is important for medical management, family planning, and engagement in research studies. Next generation sequencing has driven the pace of discovery of novel genetic syndromes, yet there are marked inconsistencies in the level of evidence for gene-disease relationships (GDRs) that complicate the application of genetic testing. We have formed the Syndromic Disorders Gene Curation Expert Panel (SD-GCEP), an international group of disease experts, gene curation framework experts, and biocurators representing 23 institutions across 5 continents to thoroughly curate the evidence supporting the relationship of a gene in causing a disease, and to quantify the strength of that evidence using the framework developed by the Clinical Genome Resource (ClinGen). The SD- GCEP consists of representatives from major stakeholders including Online Mendelian Inheritance in Man (OMIM), Monarch Initiative’s Mondo disease ontology, Genomics England PanelApp, PanelApp Australia, Centers for Mendelian Genomics, diagnostic laboratories (Ambry, Illumina, Invitae), practicing clinical geneticists, genetic counselors, and rare disease and model organism researchers. In Aim 1, we will perform 68 GDR precurations, curations and recurations representing the most commonly tested syndromic disorders not within the purview of other GCEPs, which generally focus on a specific organ or pathway. In Aim 2, we will perform 89 precurations, curations, and recurations for syndromic GDRs identified through clinical exome and genome sequencing performed by diagnostic laboratories with personnel involved in the SD-GCEP. In Aim 3A, we will perform 24 precurations, curations, and recurations for newly discovered syndromic GDRs from the Centers for Mendelian Genomics, as these are of high interest to diagnostic laboratories to determine when genes should be added to panels and to clinicians to guide clinical diagnosis and management. The SD-GCEP is highly experienced and collaborative. In Aim 3B, we will continue to accept requests from other GCEPs to curate 15 GDRs of interest that are beyond the scope of the other GCEP due to the syndromic nature of the condition. In total, we will perform 196 precurations, curations, and recurations of GDRs for syndromic disorders over three years. All curations will be performed using the ClinGen Gene Curation Interface and publicly shared though the ClinGen knowledgebase to improve genetic testing and diagnosis for syndromic disorders and to highlight where further research is needed.

NIH Research Projects · FY 2026 · 2022-08

PROJECT SUMMARY The fragile population of children with medical complexity (children with congenital or acquired multisystem disease, neurologic impairment and/or dependence on medical technology) accounts for up to one third of all pediatric medical expenditure. Systematic investigation of the contribution of nutrition to chronic disease burden in this population has not been done and is urgently needed, as it has vast potential to optimize clinical outcomes such as feeding intolerance, improve quality of life and reduce healthcare cost. Dr. Bridget Hron is a pediatric gastroenterologist and nutrition specialist with advanced training in the nutritional regulation of metabolism and inflammation who is perfectly suited to address this critical unmet need. Her career goal is to direct a clinical research program focused on discovering evidenced-based, state of the art nutritional interventions that transform the clinical care of the most vulnerable pediatric population. In Dr. Hron's recent work, she has shown that blenderized diets, which are pureed high quality table foods such as fruits, vegetables, meat and legumes administered via gastrostomy tube have a beneficial effect on health care utilization, reducing admissions for pulmonary reasons by 53%. Over the course of this career development award, Dr. Hron will study the effect of blenderized feeds varying in viscosity on gastrointestinal symptoms and on esophageal and gastric function. The innovative aggregate randomized, controlled N-of-1 study design outlined in this proposal allows each individual participant to determine diet of maximal efficacy, but also allows for estimation of population effect of the interventions. The exceptional mentorship and original scientific training proposed in this career development award are critical for Dr. Hron's academic development. Dr. Rachel Rosen, primary mentor, is an expert in aerodigestive medicine, and Dr. Christopher Schmid, co-mentor, is an expert in biostatistics with a focus on N-of-1 trials – both have outstanding backgrounds in clinical research with deep commitment to mentorship. She will be supported by her Scholarship Advisory Committee consisting of Drs. Wayne Lencer, Christopher Duggan, Jay Berry and Sharon Donovan, who lend content-area expertise. Her formal training will include a Master of Science: Concentration in Food Science at University of Illinois Urbana-Champaign, advanced courses in statistical analysis at the Harvard T.H. Chan School of Public Health (HSPH), personal instruction on mediators of esophageal and gastric motility by Dr. Rosen, and professional development courses through HSPH and Harvard Catalyst. Her training and research activities will be conducted in the unparalleled academic environments of Boston Children's Hospital and Harvard Medical School, which are firmly committed to Dr. Hron's successful transition to an independent research career.

NIH Research Projects · FY 2025 · 2022-08

PROJECT SUMMARY/ABSTRACT Every year, >1 million patients undergo bone repair procedures in the United States. Autologous bone grafting remains the preferred treatment for bone defects, but this practice is limited by bone availability and donor site morbidity. Alternatively, the development of therapies that exploit the osteogenic potential of bone marrow- derived mesenchymal stem cells (bm-MSCs) continues to be a priority in regenerative medicine. However, efforts remain largely empirical due to a poor understanding of the mechanisms regulating bm-MSC osteogenic activity in vivo. Our overarching goal is to elucidate the mechanisms regulating ossification and develop therapeutic strategies for bone regeneration using autologous bm-MSCs. Previously, we showed that preserving human bm-MSCs' osteogenic potential depends on sustaining proximity to endothelial cells (ECs). More recently, we have found that the type of ECs drastically affects bm-MSC fate in vivo. Specifically, vascular networks lined by human trabecular bone arteriole ECs (tba-ECs) could spontaneously induce osteogenic differentiation of bm- MSCs. In contrast, non-bone ECs could not. Our Preliminary Data suggest that the expression of KITLG drives this unique osteoinductive potential. Indeed, silencing KITLG in tba-ECs completely abrogated osteogenesis upon implantation in vivo, whereas overexpressing KITLG in non-bone ECs conferred robust osteoinductive properties. Our data also suggest that KITLG expression in tba-ECs is regulated by type I interferon (IFN) signaling, a previously unknown link. Our central hypothesis is that a constitutive IFN-KITLG mechanism drives the distinct osteoinductive properties of human tba-ECs. We also postulate that educating induced pluripotent stem cells (iPSCs) could offer a plentiful source of surrogate tba-ECs, eliminating the need for harvesting autologous bone. To test these hypotheses, we propose three specific aims. In Aim-1, we will dissect the mechanism by which human tba-ECs mediates osteogenesis via KITLG expression. We will determine which KITLG isoform (soluble vs. membrane-bound) is indispensable and dissect the role of recruited c-Kit+ hematopoietic progenitor cells (c-Kit+ HPCs) in osteogenesis. In Aim-2, we will determine the molecular mechanism that regulates KITLG expression in human tba-ECs. We will use a CRISPR/Cas9 loss‐of‐function approach to silence components of the type I IFN pathway and unravel the interactions between IFN signaling mediators and the enhancer-promoter region of the KITLG gene. In Aim-3, we will pursue strategies to educate human iPSC-derived ECs to acquire osteoinductive function, including transient activation of KITLG and IFN signaling. In summary, these studies will define the cellular and molecular mechanisms by which human tba- ECs regulate the osteogenic differentiation of bm-MSCs and, in turn, ossification. This fundamental knowledge will form the foundation for strategies to promote bone repair and regeneration.

NIH Research Projects · FY 2026 · 2022-08

Project Summary The incidence of congenital heart defects (CHDs) and cardiovascular disease (CVD) in patients with fetal alcohol spectrum disorder (FASD) are poorly characterized. Cardiovascular abnormalities may be common in FASD; however, comprehensive retrospective studies on lifetime CVD risk in adult patient cohorts have yet to be performed. Cellular and molecular mechanisms underlying FASD-mediated CHD and CVD are also largely unknown along with any biomarkers that would allow the patient population to be stratified based on CVD risk. Here we present preliminary data from our retrospective clinic cohort that demonstrate that females with FASD have an overall increase in CHD, myocardial infarction (MI) rate, and the likelihood of being diagnosed with any CVD in adulthood. Females with FASD also have significantly reduced ejection fraction relative to matched controls. These data suggest that FASD is a risk factor for CHD in newborns and CVD in adults. In a zebrafish model of embryonic alcohol exposure (EAE), we confirmed a primary defect in cardiomyocyte migration that causes subsequent functional and structural heart abnormalities, including contractility deficits and ventricular wall abnormalities that persist through adulthood. Our findings indicate that EAE zebrafish can serve as a model for lifelong cardiac function in the presence and absence of CHD. We propose three Specific Aims to test the central hypothesis that FASD patients have an increased incidence of CVD and that the zebrafish EAE model will uncover novel molecular mediators and biomarkers that explain and predict CVD risk. In Specific Aim 1, we will perform a retrospective study to determine CVD incidence in an adult FASD patient cohort, including CHD, hypertension, cardiomyopathy, MI, cerebrovascular accident, and embolism, as well as their association with other metabolic and inflammatory conditions. In Specific Aim 2, we will define molecular mechanisms underlying embryonic heart defects in a zebrafish EAE model by identifying and functionally evaluating the impact of molecular alterations in migratory myl7+ cardiomyocytes that form the cardiac cone through hypothesis-driven (PDGF pathway) and unbiased (bulk RNA-sequencing on FACS-isolated myl7+ cardiomyocytes) approaches. In Specific Aim 3, we will test the hypothesis that EAE adults with a CHD are susceptible to cardiac dysfunction and cardiomyopathy due to lasting alterations in cardiac structure, function, and molecular signature. Taken together, the proposed studies will provide fundamental insights into the cardiovascular health outcomes of patients with FASD, reveal novel molecular mediators of EtOH-induced CHDs, and identify biomarkers of adult cardiac dysfunction in EAE adults. Cardiovascular diseases may contribute significantly to morbidity and mortality in affected patients. By identifying which CVD outcomes impact FASD patients and what additional metabolic and inflammatory factors indicate risk, we will provide an opportunity for early intervention. Further, identification of molecular mediators of CHD and cardiomyopathy in a zebrafish model of EAE will allow us to expand our mechanistic understanding of the effects of PAE across the lifespan.

NIH Research Projects · FY 2025 · 2022-08

Project Summary Our long-term goal is to understand the pathologic mechanisms underlying insulin resistance-associated metabolic disease, and in particular non-alcoholic fatty liver disease (NAFLD). Although NAFLD affects 1 in 4 Americans and significantly increases the risk of liver cancer and cardiovascular disease, no FDA-approved NAFLD therapies currently exist. Although insulin resistance increases the incidence of NAFLD, the mechanisms linking these pathologies are not completely understood and warrant further study. To identify the key factors mediating insulin resistance-associated NAFLD, we screened diabetic individuals and healthy controls for differences in plasma protein concentrations and hepatic gene expression We found that ceruloplasmin (CP), a liver-secreted copper-binding protein, was increased in the liver and plasma of diabetic patients. These data are consistent with previous findings of high plasma CP concentrations in individuals with diabetes or obesity, and positive correlations of CP with the severity of diabetes, obesity, CVD risk, and mortality. In addition, our preliminary data and data from others show that hepatic CP expression is high in humans and rodents with NAFLD, while their hepatic copper levels are low11-16, consistent with the role of CP to reduce hepatic copper content. However, the functional significance of the high hepatic CP and low hepatic copper associated with insulin resistance and NAFLD is unclear. To address this deficiency, we have characterized mouse models of insulin resistance and hepatocyte-specific CP deletion. These preliminary studies have suggested a previously unrecognized role of hepatic CP and copper in linking insulin resistance and NAFLD. While a HFD decreases hepatic copper content and promotes NAFLD, liver-specific knockdown or knockout of CP (L-CP KO) increases hepatic copper content, alters the expression of genes involved in hepatic lipid metabolism, and ameliorates NAFLD in these insulin resistant mice. We hypothesize that insulin resistance-induced hepatic CP promotes NAFLD by disrupting copper homeostasis and lipid metabolism. In Aim 1, we will determine how insulin resistance induces hepatic CP gene transcription; in Aim 2, we will determine how hepatic CP regulates copper homeostasis and the development of NAFLD; and in Aim 3, we will elucidate the molecular mechanisms by which dysregulation of hepatic copper homeostasis promotes NAFLD. We expect that the completion of the proposed studies will define the role of hepatic CP and copper in metabolic regulation and the development of NAFLD in insulin resistant states, which may suggest new means of treating NAFLD.

NIH Research Projects · FY 2026 · 2022-08

Project Summary A substantial proportion of psychiatric inpatients are readmitted within 30 days of discharge. Readmissions not only are disruptive but also cause enormous economic burden for patients and families, and are a key driver of rising healthcare costs. Reducing and predicting unplanned readmission are therefore major unmet needs of psychiatric care. Developing machine learning (ML)-based natural language processing (NLP) prediction tools using electronic health records (EHRs) is a key priority as such tools could not only be used to help target the delivery of resource-intensive interventions to those patients at greatest risk, but also reduce psychiatric health- care costs. A key aspect in building effective risk predictive models is the modeling of temporal structure in the narratives. Information about the historical and present health states and timing of events (e.g., substance use start/stop timing, recent fluctuations in suicidality or symptoms), may play a key role in predicting readmission risk. Natural language annotation (i.e., tagging text such as events, symptoms, and anchoring them on a timeline) is a key step for training ML classifiers. No psychiatry-specific resources or guidelines exist for the modeling of temporality in clinical text, and as a result no robust scalable and explainable ML predictive models incorporating temporal information have been developed. We propose to deliver a psychiatric specific temporal relation annotation scheme, build open-source tools for extracting temporal information, and develop readmission prediction models for psychiatric patients. Aim 1 is a data resource creation aim in which we create a large repository of psychiatric text for building our readmission classifier, de-identify a subset of that data to allow for sharing with the research community, and create a layer of temporal annotations for that subset. In Aim 2, we extract temporal information from the data in the repository to create temporal graphs, and apply graph neural networks to these graphs to extract features for predicting 30-day readmission risk. In Aim 3 we build and evaluate multiple versions of 30-day readmission risk classifiers, and feedback performance to Aim 2 to improve temporal modeling. We develop unsupervised clustering on top of our classifiers to discover patient sub-groups. We include practical evaluations including a comparison to human experts and an evaluation of model performance on simulated future data. The study brings together a team experienced in psychiatric phenotyping and application of EHRs, and a team active in developing cutting- edge methods in ML for natural language data. This work will serve as the foundation for future translational studies, including implementing readmission classifiers into clinical workflows and clinical trials of interventions to reduce readmission risk.

NIH Research Projects · FY 2026 · 2022-08

SUMMARY Long-term expression of broadly neutralizing antibodies (bNAbs) has the potential to suppress an established HIV-1 infection. However, current methods for maintaining high bNAb concentrations necessary for this control are inadequate. Passive infusion of bNAbs is prohibitively expensive and requires HIV-1 positive individuals to receive infusions on a weekly or monthly basis. Delivery of bNAbs by gene-therapy vectors almost invariably raises anti-drug antibodies (ADA) against expressed bNAbs, which are immunogenic due to their extensive hypermutation. Most importantly, no single set of antibodies can adequately suppress the range of viruses in the population, in large part because current antibody delivery systems fail to do what an immune system does well: adapt to a diverse and evolving pathogen. Here we describe a series of technical advances that allow us to introduce bNAb heavy- and light-chain genes into their native loci in primary B cells. These technologies enable in vivo improvement of bNAbs through affinity maturation in mice and primates, using the acquired wisdom of the humoral response to rapidly increase antibody potency, breadth, and bioavailability. They also allow us to test the core hypothesis of this proposal that B-cell delivered bNAbs can permanently suppress an established infection in the absence of anti-retroviral therapy (ART). The chief technical advance that enables these studies is the development of an efficient double-editing technique for simultaneously replacing the variable heavy and light chain segments of B cell receptors. This is made possible through use of a newly characterized Cas12a ortholog and a unique homology-directed repair template design capable of efficiently replacing nearly any endogenous BCR variable region. The net consequence is that, unlike related B-cell editing approaches, the full regulatory apparatus of the B cell is left intact, facilitating robust B-cell development and efficient affinity maturation of the B-cell receptor. The project is divided into three aims. Aim 1 will increase the breadth and potency of three well characterized bNAbs through affinity maturation in vivo. Aim 2 will extend CRISPR editing to the Fc domain, introducing a recently described set of mutations into the IgG1 Fc domain that facilitate antibody transfer across the blood- brain barrier. Finally, Aim 3 tests the ability of primary B cells expressing the bNAbs improved in Aim 1 to control a SHIV infection in rhesus macaques. A series of structured treatment interrupts will be performed to drive CAR B proliferation and generate an individualized response to virus that emerges from the reservoir. After these structured interruptions, ART will be permanently withdrawn to determine if CAR B cells alone can control an established infection.

NIH Research Projects · FY 2025 · 2022-08

Project Summary The objective of this proposal is to develop targeted therapies for venous malformations (VMs). VMs are slow- flow vascular lesions associated with disfigurement, pain, and functional impairment. Recently, systemic inhibition of the mammalian target of rapamycin (mTOR) with sirolimus has proven efficacious for treating children with complex VMs. However, systemic drug delivery is associated with side effects that limit treatment. Therefore, safe, targeted therapies that minimize systemic toxicity are required. The proposed project will develop nanoparticulate (NP) targeted drug delivery systems to achieve high local drug concentration in VMs while minimizing systemic distribution. This will be achieved by virtue of enhanced permeation and retention (EPR), a well-recognized phenomenon in cancer biology whereby leaky tumor vasculature allows for preferential uptake of nanoparticles compared to uptake in tissues with normal vasculature. Passive NP accumulation within VMs, due to EPR, will be enhanced with active targeting techniques, such as photo-targeting. The surfaces of NPs will be coated with molecules that encourage cell uptake. These molecules will be inactivated with a “caging group,” a reversibly bound molecule that is sensitive to a specific wavelength of light. Upon irradiation with light, the caging molecule will be removed from the NP. Therefore, NPs can be systemically injected and remain unbound to tissues. However, irradiation of the VM will cause “uncaging” to occur, which will activate the NPs, and allow for enhanced NP binding and drug release at the target site. To test this hypothesis, we propose three specific aims: Specific Aim 1: Formulation and characterization of NPs with prolonged dwell times in VMs. Specific Aim 2: Study of targeted NP drug delivery systems in vivo. Specific Aim 3: Study of NP drug delivery systems on therapeutic efficacy in vivo. With the guidance and mentorship of Dr. Daniel Kohane, Dr. Cullion has developed a five- year career development plan to provide the mentored research, technical skill development, and didactic training needed to achieve her goals of (1) becoming an expert in nanomedicine and drug delivery for the treatment of VMs and (2) achieving scientific independence and becoming an R01 funded physician-scientist with a career in translational research focused on nanomedicine and drug delivery for vascular anomalies.