Boston Children'S Hospital

NIH Research Projects · FY 2026 · 2024-04

PROJECT SUMMARY The following proposal outlines a 5-year career training plan that will prepare Dr. Alexander Holtz to be an independent physician-scientist and leader in the field of induced pluripotent stem cell (iPSC) biology and cellular therapies for pulmonary vascular disorders. Pulmonary vascular diseases, such as pulmonary hypertension (PH), are devastating illnesses associated with high morbidity and mortality with frustratingly limited treatment options. Endothelial cell dysfunction is a core mechanistic driver of these disorders, especially in cases where gene mutations impact endothelial cell biology (FOXF1, BMPR2, etc.). Dr. Holtz’s long-term vision is to develop autologous endothelial replacement therapies where patient-derived iPSCs are generated, undergo gene-correction ex vivo, and then are differentiated to endothelial cells (iEndos) to provide a limitless supply of healthy donor endothelial cells for transplantation without the need for lifelong immunosuppression. Dr. Holtz presents his initial discoveries that patterning of iEndos with BMP9 shifts cells towards a ‘lung-like’ molecular profile, including induction of the lung endothelial cell marker TMEM100, and enables durable engraftment of transplanted iEndos into the mouse lung microvasculature. He also shows that this BMP9- mediated patterning process requires active Notch signaling. Using this novel system, Dr. Holtz will test the hypothesis that BMP9 and Notch signaling cooperatively induce TMEM100 expression to produce functional, engraftable cells for treatment of monogenic pulmonary vascular disorders. Specifically, he will 1) assess the differentiation capacity, longevity, and progenitor function of engrafted iEndos in the lung microvasculature; 2) utilize a ‘competitive lung reconstitution assay’ to delineate the functional role of BMP9- and Notch-mediated induction of TMEM100 to facilitate iEndo engraftment and to test the translational potential of iEndos derived from gene-corrected PH patient-specific hiPSCs (BMPR2, FOXF1); and 3) test the efficacy of endothelial replacement therapies in immunocompetent hosts using a mouse model of FOXF1-mediated pulmonary vascular disease. This work will provide a fundamental advancement towards developing endothelial replacement therapies for a broad range of congenital and acquired pulmonary vascular diseases. Dr. Holtz has 90% protected time from Boston Children’s Hospital Division of Genetics and Genomics to accomplish these aims under the guidance of Dr. Darrell Kotton at the Center for Regenerative Medicine at Boston University/Boston Medical Center. He has assembled a remarkable team of advisors with diverse expertise to assist in his career development and scientific research. Dr. Holtz details a comprehensive training plan that takes advantage of his unique cross-institutional collaboration that includes mentored research, didactic coursework, attendance and presentation at national meetings, preparation of manuscripts, and acquiring additional grant support culminating in an R01. Dr. Holtz has the commitment of both institutions to accomplish these goals and transition to an independent physician-scientist position by the end of the award.

NIH Research Projects · FY 2025 · 2024-04

PROJECT SUMMARY/ABSTRACT Humans have evolved an expanded and elaborated brain capable of higher-order cognition. However, the sequence variants and resulting neural specializations that distinguish humans from other mammals are commonly dysregulated in diseases such as autism spectrum disorder (ASD). This suggests that human-specific non-coding variants are enriched for neural functions and may underlie genetic and phenotypic disease vulnerabilities. Thus, there is a critical need to identify and characterize the non-coding variants that underlie human neural specializations. However, identifying the causal variants that contribute to human neural specializations is a daunting challenge that has been likened to searching for needles in a haystack. In this proposal, Dr. Janet Song will improve prioritization of human-specific variants for further functional analysis using two complementary approaches (Aims 1 and 2) and determine whether prioritized variants regulate nearby gene expression in a high-throughput manner (Aim 3). Dr. Song will use the human-chimpanzee tetraploid system to link regulatory regions that are differentially accessible between species to nearby differential genes in neural progenitor cells and excitatory neurons, two cell types that are profoundly changed in humans and are commonly dysregulated in neurological diseases (Aim 1 – K99 phase). As a complementary approach, Dr. Song will identify constrained human-specific insertions and assess their contribution to ASD risk (Aim 2 – K99/R00 phase). Dr. Song will then evaluate the effects of human-specific variants on nearby gene expression in neural cell types using CRISPR inhibition screens (Aim 3 – K99/R00 phase). This K99/R00 proposal will support Dr. Janet Song in her pursuit of the genetic basis of human neural specializations and allow her to acquire new skills in comparative and functional genomics that will open up innovative approaches to explore this problem. This proposal will be initiated during the mentored period in Dr. Christopher Walsh’s lab at Boston Children’s Hospital / Harvard Medical School and continue in Dr. Song’s own lab upon securing an independent position. In addition to providing immediate insights into the genetic basis of human neural specializations, the proposed research will lay the foundation for Dr. Song’s independent research program. It will provide a framework for future studies in additional cell types or paradigms, pinpoint high-priority loci for single-locus studies, and identify a corpus of human-evolved elements that may contribute to genetic risk for neurodevelopmental and neuropsychiatric diseases. Long-term, Dr. Song’s independent research program will dissect how sequences that changed in humans relative to other mammals result in human-specific neural phenotypes, and ultimately, contribute to neurodevelopmental and neuropsychiatric diseases.

NIH Research Projects · FY 2026 · 2024-04

Platelets are critical in maintaining hemostasis and platelet counts are tightly regulated in healthy individuals. In several pathologies, the disruption of thrombopoiesis leads to abnormal platelet counts which significantly impact clotting and/or bleeding risks. Thrombopoietin (TPO) signaling through the myeloproliferative leukemia virus (MPL) receptor is the only known driver of megakaryocyte (MK) differentiation and maturation from hematopoietic stem cells (HSCs). However, therapies targeting the TPO/MPL axis do not work for all patients, leading to an unmet need for identifying new targets to modulate platelet counts. We have found that membrane polyunsaturated fatty acids (PUFAs) in MK progenitors regulate MK development and platelet production, representing a novel target for modulating megakaryopoiesis and platelet count. Our main hypothesis is that MKs and their progenitor cells utilize the CD36 receptor to take up PUFAs from exogenous sources, and accumulation of membrane PUFAs is essential for maintaining normal MK development and platelet count. This hypothesis will be tested in three specific aims using novel techniques such as click chemistry, cellular barcoding and phospho-flow cytometry, and the newly published Fatty Acid Library for Comprehensive ONtologies (FALCON) platform paired with human and murine in vitro models, in vivo murine models, and human iPSC-derived organoids. Aim 1 will determine the cell-type-specific enzyme requirement for PUFA accumulation during megakaryopoiesis. We will use in vivo and in vitro models to determine which enzymes play a role in PUFA accumulation in MKs and whether they are viable targets to manipulate MK development. Aim 2 will reveal if the CD36 receptor preferentially takes up PUFAs at the expense of saturated fatty acids. We will use CD36-/- mice and a novel in vivo click-chemistry technique to provide the first in vivo evidence of selective PUFA uptake. We will determine if CD36 within MKs and MK progenitors preferentially promotes the internalization of fatty acids with a specific saturation status and whether this process is impacted by dietary fatty acid composition. Aim 3 will examine the mechanisms by which PUFA uptake influences megakaryopoiesis. This inquiry will be addressed using 1) a targeted approach focusing on MPL signaling via interaction with membrane lipids and 2) a hypothesis-generating approach using the FALCON platform to generate new leads into potential TPO-independent MK differentiation pathways impacted by fatty acids. Successful completion of the proposed experiments will extend the discovery of the role of membrane PUFAs in MK development and platelet production to reveal a detailed and actionable roadmap of the mechanisms by which both human and murine MKs and progenitors accumulate and utilize PUFAs to drive platelet production, potentially leading to therapeutic and dietary interventions to modulate platelet production for patients with diverse thrombotic and platelet disorders.

NIH Research Projects · FY 2026 · 2024-04

Cancer immunotherapy using checkpoint blockade (CPB) has revolutionized cancer treatment, providing durable cures with acceptable toxicity in some cancer patients. However, only a few cancer types respond and even in responding tumor types, response is often limited to a minority of these cancers, which raises the question whether directly targeting T cells is sufficient and whether other aspects of immunity can be exploited to stimulate or boost antitumor immunity. Inflammatory, or immunogenic, cell death is emerging to be an important immune node that bridges innate and adaptive immunity to stimulate antitumor immunity as well as to potentiate CPB. While antineoplastic agents currently used in humans can also sometimes induce inflammatory cell death to help re-establish immune surveillance in the tumor microenvironment, these agents do not predictably induce immunogenic cell death, thus leading to variable outcomes. The hypothesis we propose in this project can fundamentally change this scenario by inducing predictable immunogenic cell death through targeting key molecules known as gasdermins (GSDMs) in particular GSDMD, which are widely expressed in cancers and can directly induce immunogenic cell death upon activation. GSDMD mediates pyroptosis and cytokine release downstream of inflammasomes, which are supramolecular complexes that activate inflammatory caspases (caspase-1/-4/-5 in humans, caspase-1/-11 in mice). Activated caspase-1 processes IL-1 family cytokines to their active forms and all inflammatory caspases cleave GSDMD to produce an N-terminal (NT) fragment that forms pores in the cell membrane to induce pyroptosis, and to release IL-1 family cytokines and other inflammatory mediators, including ATP and HMGB1. We specifically propose to identify small molecule agonists of GSDMD for direct induction of immunogenic pyroptosis in cancer cells, which may also synergize with other immune modulators as well as with CPB. GSDMD is widely expressed in cancers. While GSDMD is also expressed in some normal tissues, the need for only a small fraction of tumor cells to undergo pyroptosis to alert the immune system may make GSDMD agonism non-toxic even when administered systemically.

NIH Research Projects · FY 2026 · 2024-04

ABSTRACT The major goal of this project is to understand how glypicans protect against bacterial lung infection by coordinating a cellular defense mechanism that promotes bacterial clearance and inhibits bacterial invasion. Pathogens use all tricks available to survive in the hostile host environment. Many pathogens bind to host extracellular matrix (ECM) components and their receptors for their attachment, invasion and immune evasion, suggesting that normal functions of the ECM are exploited for pathogenesis. However, while both host cells and microbes are known to change their phenotypes and adapt to the artificial environment when cultured in vitro, the current paradigm that the host ECM promotes infection as attachment and invasion receptors is based predominately on data from in vitro systems. Furthermore, while the ability of the ECM to promote pathogenesis has received much of the attention, potential beneficial functions of ECM components in infections have been largely ignored. The abundant expression of ECM components and their receptors at sites where pathogens frequently encounter the host, such as at the cell surface and subepithelial compartments, suggests that certain ECM components may actually function in host defense. Glypicans, of which there are 6 members in mammals, comprise a major family of cell surface heparan sulfate proteoglycans, that function as receptors for morphogens, growth factors, cytokines/chemokines, and ECM components, among other heparin/heparan sulfate-binding proteins. We found in preliminary studies that glypican-4 null (Gpc4-/-) mice are significantly hypersusceptible to Staphylococcus aureus lung infection, suggesting that Gpc4 protects against S. aureus pathogenesis. S. aureus is a major human pathogen and it has a unique ability to infect virtually every tissue and organ system, including the lung. Our studies suggested that Gpc4 provides protection against S. aureus lung infection via 2 distinct mechanisms: a neutrophil- dependent and a neutrophil-independent mechanism in which Gpc4 inhibits S. aureus invasion into host cells. This proposal will establish the importance of these previously unknown mechanisms and define their molecular and cellular details in 3 Specific Aims. Aim 1 will determine how glypicans regulate early neutrophil recruitment in bacterial infection. Aim 2 will define how glypicans inhibit bacterial entry of host cells, and Aim 3 will elucidate how bacteria counteract defense mechanisms mediated by glypicans.

NIH Research Projects · FY 2026 · 2024-04

ABSTRACT Autism Spectrum Disorder (ASD) and other neurodevelopmental conditions are currently classified according to behavioral criteria. This clinical diagnostic framework does not take into account underlying pathophysiology and is often insufficient to predict outcomes. The long-term goal is to develop biomarkers that will help cluster individuals with ASD and related conditions into biologically distinct groups (“biotypes”) and provide the means to understand the underlying neural mechanisms. Developing biomarkers for use in children is particularly important so that they can be used from the time symptoms are first recognized. Objective measurement of sensory processing via electroencephalography (EEG) is a non-invasive and scientifically translatable method to measure function of neural circuits. The proposed project tests the central hypothesis that EEG measures of sensory processing predict behavioral phenotype (i.e., they are relevant to functional outcome) but also inform the reclassification of neurodevelopmental conditions by neural circuit pathophysiology to complement behavioral phenotype. This project uses EEG to measure sensory/neural circuit function in 3-4 year old children with ASD (n=150), typical development (TD; n=75), and children who do not have ASD but whose caregivers have concerns about how they process sensory information (Sensory Processing Concerns (SPC) group; n=150). Aim 1: Determine how circuit-level brain activity, measured in response to sensory stimuli, is altered in children with ASD vs. SPC vs. TD. Aim 2: Test if sensory EEG endophenotypes predict behavioral phenotype (e.g., anxiety, attention, irritability, social skills) within and across groups (ASD, TD, SPC). Aim 3: Delineate neurobiologically-informed subtypes of neural circuit activity in individuals. Rather than relying on behavioral phenotype as the gold standard, this exploratory aim takes a data-driven approach to cluster individuals in ASD (and potentially SPC) into biologically distinct subgroups (biotypes) defined by EEG measures of neural circuit activity to sensory inputs. The proposed project is conceptually innovative because it uses the paradoxical co- occurrence of sensory hyper- and hypo-reactivity within individuals as the nidus for developing biomarkers that reflect the granular neurobiological principles underlying sensory processing and ASD. The proposed project is technically innovative because many of the proposed EEG measures are novel or have never been used to study ASD. The project is significant because it moves the field from developing biomarkers that reflect behavioral understanding of ASD towards biomarkers that provide neurobiological understanding of ASD. Ultimately, this will allow development of diagnostic and stratification biomarkers that guide biologically targeted, personalized treatments in children with ASD and other neurodevelopmental conditions.

NIH Research Projects · FY 2025 · 2024-04

PROJECT SUMMARY/ ABSTRACT Atrial Fibrillation (AF) is an arrhythmia characterized by the aberrant, unorganized initiation and propagation of electrical impulses across the atria. The most common serious arrhythmia, AF affected an estimated 33.5 million globally in 2010, with a lifetime risk of 1 in 3 individuals older than 55 years old. A greater understanding of the mechanisms of AF is required to design more effective treatment strategies. Genome-wide association studies have linked AF with over 143 genomic loci, including the transcription factor (TF) TBX5. In our previous study, we integrated single-nucleus RNA- and ATAC- sequencing (multiomics) of control and Tbx5 KO aCMs with TBX5 chromatin occupancy in aCMs to identify direct TBX5 targets that might contribute to AF. In the process, we uncovered a novel function for TBX5 in maintaining genomic accessibility at enhancer elements, likely by recruiting chromatin modifying proteins. TBX5 interacts with chromodomain helicase DNA binding protein 4 (CHD4), a component of the nucleosome remodeling and histone deacetylase (NuRD) complex. Although conventionally viewed as a transcriptional repressor, our Preliminary Data reveal an exciting, alternative role for CHD4 as a transcriptional activator. To better understand gene expression changes central to AF, we performed multiomics on a second AF mouse model caused by Liver Kinase B1 (LKB1) inactivation in aCMs, a gene decreased in human AF patient aCMs, revealing 632 core atrial rhythm (AR) genes that are commonly downregulated in aCMs from both models. 61% of AR genes were adjacent to regions co-occupied by CHD4-TBX5, and include the Na+ channel SCN5A, regulators FGF12/13 and the gap junction channels Connexin 43 (GJA1) and Connexin 45 (GJA5). These data lead to our central hypothesis that CHD4 maintains rhythm homeostasis in healthy atria by complexing with TBX5 to maintain the accessibility of TBX5-dependent enhancers, promoting the expression of genes that are critical for normal aCM electrical function. To determine the mechanisms by which the CHD4-TBX5 complex regulates the atrial enhancer network, we will characterize the phenotypic and genomic consequences of Chd4 inactivation in aCMs, identify CHD4- interacting proteins required for aCM gene activation, and uncover other cis-activating factors that function cooperatively with CHD4-TBX5 to regulate aCM gene expression. To identify core atrial genes required for atrial rhythm, we will test the requirement of select AR genes to maintain atrial rhythm by inactivation. Genes identified as critical regulators of rhythm will then be evaluated for therapeutic efficacy in AF prevention and reversal in our established AF mouse models. This proposal is significant because it will mechanistically characterize genes required for atrial function, determine how they are regulated, and test their potential as therapies in multiple models, laying the groundwork for the development of more effective treatment strategies.

NIH Research Projects · FY 2025 · 2024-04

PROJECT SUMMARY Traumatic brain injury (TBI) often leads to epilepsy with a delay of weeks to months after the initial trauma. Immediately following and for days to weeks after TBI, increased extracellular glutamate leads to hyperexcitation of neurons, and to neuronal injury. These effects are exacerbated by post-traumatic downregulation of glutamate transporter 1 (GLT-1), the protein responsible for synaptic glutamate clearance. Long-term, post-TBI glutamate excitotoxicity leads to a progressive loss of vulnerable inhibitory interneurons and GABAergic intracortical inhibition, and contributes to a range of chronic post-TBI symptoms, that include post-traumatic epilepsy (PTE). We identified that short-term treatment with the injectable beta-lactam ceftriaxone (CRO) increases GLT-1 expression and mitigates post-TBI neuronal death and seizures in a rat acquired epilepsy model, albeit incompletely. CRO-mediated GLT-1 rescue does not persist beyond cessation of treatment, while the TBI- induced depression of the gene that encodes for GLT-1 persists up to 6 weeks after TBI. Since CRO and similar injectable antibiotics cannot be given for weeks to months without complications, a sustained non-antibiotic treatment that can be administered safely and conveniently is highly desirable. The atypical non-bactericidal beta-lactam clavulanic acid (CLAV) also increases GLT-1 expression and protects against glutamate-mediated damage in animal models. This well-tolerated, orally available drug can potentially serve as a mainstay of prolonged post-TBI GLT-1 upregulation strategies aimed at sustained neuroprotection and anti-epileptogenesis. However, essential questions pertaining to CLAV mechanism and efficacy should be addressed in the preclinical setting. Based on prior work showing that CRO modulates GLT-1 after TBI, and published data showing CLAV does the same in other animal models, we propose a set of exploratory experiments aimed to (1) determine the dose-response relationship between CLAV and GLT-1 expression following TBI, (2) test whether long-term beta- lactam treatment mitigates TBI-induced long-term GLT-1 depression, oxidative stress, and inhibitory interneuronal death, (3) test whether long-term beta-lactam treatment prevents or mitigates progressive, post- TBI decrease in cortical inhibition, seizure threshold, and PTE. Given that no intervention targeting post-TBI epileptogenesis is available, our proposed experiments will be the first step toward a novel treatment. Despite the strong implication of glutamate-mediated changes in post-traumatic epileptogenesis, there is presently no proven long-term clinical method to clear excessive glutamate from the extracellular space after injury. Our proposal combines established basic science methods with a novel application of CLAV treatment as an antiepileptogenic intervention which can be administered acutely after TBI. As CLAV is already FDA approved for human use, positive data from the proposed studies may be rapidly translated to further development of this post-TBI treatment strategy, and to human trials. Beyond TBI, enhancement of glutamate transport may also prove useful following various other forms of brain injury.

NIH Research Projects · FY 2026 · 2024-04

ABSTRACT The neural crest is a migratory stem cell population that is essential for the development of various tissues and organs during embryogenesis. Disruptions in neural crest development contribute to the pathogenesis of craniofacial defects and other congenital malformations, imposing a significant burden on individuals and society. Understanding the molecular mechanisms underlying neural crest formation is crucial for proper diagnosis of these conditions. While previous research has focused on protein-DNA interactions in activating neural crest genes, the role of chromatin conformation changes in this process remains poorly understood. Previous studies characterizing the architecture of the neural crest genome revealed that these cells display a complex enhancer-promoter interactome characterized by a reliance on long-range interactions. This proposal aims to investigate the contribution of architectural proteins, specifically CTCF and YY1, in establishing this unique chromatin organization. We hypothesize that architectural proteins interact with pioneer transcription factors to assemble the neural crest interactome and promote activation of gene regulatory circuits. We will test this hypothesis in three specific aims. Aim 1 will define the function of architectural proteins in neural crest formation by identifying the chromatin loops mediated by CTCF and YY1 through chromatin conformation capture. We will also target architectural proteins in loss-of-function studies to uncover their relative contributions to the establishment of the neural crest enhancer interactome. Aim 2 seeks to determine how tissue-specific enhancer-promoter loops are established during neural crest specification. To accomplish this, we will investigate the functional and physical interactions between architectural proteins and the pioneer transcription factors that define the neural crest lineage. Aim 3 will define how disease-linked mutations in architectural proteins affect neural crest chromatin organization. We will employ genome engineering to reproduce CTCF/YY1 mutations in embryonic stem cells and drive these cells into adopting a cranial neural crest fate. By elucidating the function of architectural proteins in neural crest development and their role in the genesis of craniofacial malformations, this research has the potential to pave the way for future therapeutic strategies aimed at preventing or treating these debilitating conditions.

NIH Research Projects · FY 2026 · 2024-03

PROJECT SUMMARY Malaria is an important global cause of morbidity and mortality. Most of this disease burden falls on children and pregnant women and is caused by infection with Plasmodium falciparum. The signs and symptoms of human malaria result from the exponential expansion of parasite biomass that occurs during asexual replication of parasites in human red blood cells. During this clinically important blood stage, P. falciparum parasites divide by schizogony – a process wherein components for several daughter cells are produced within a common cytoplasm prior to a complex and synchronized cytokinesis known as segmentation. Cytokinesis via segmentation is highly divergent from the process of cell division in human cells. The generation of the invasive daughter parasites, known as merozoites, occurs with high fidelity, ensuring that each daughter has a single nucleus and the required organelles. The fundamental molecular mechanisms that facilitate segmentation are incompletely understand, and this is a significant knowledge gap. Successful segmentation requires two parasite-specific structures, the inner membrane complex (IMC) and the basal complex. The IMC is a double lipid bilayer with associated proteins that, together with an underlying cytoskeletal network, dictates parasite shape and rigidity. The basal complex is a group of proteins at the posterior (i.e., basal) end of the IMC. This multi-protein molecular machine is essential for parasite cell division and is hypothesized to facilitate proper biogenesis of the IMC, likely to contribute to cell shape, and to mediate the final abscission step of cytokinesis. In the current proposal, we will move stepwise towards a molecular understanding of basal complex function. We have a high-confidence list of basal complex components, and our preliminary data demonstrate that individual proteins define subcompartments within the complex and dynamically join and depart the complex. In the first aim, we will utilize high-resolution live-cell time lapse microscopy to determine the precise order of assembly and disassembly of the basal complex. In the second aim, we will utilize super-resolution ultrastructure expansion microscopy to localize individual proteins into subcompartments of the basal complex and proximity labeling to identify novel components within these compartments. In the third aim, we evaluate the requirement for the remaining basal complex components for segmentation and determine the minimal core of proteins that are essential for basal complex function. In summary, the basal complex is essential for asexual replication of P. falciparum, is insufficiently understood, and is not targeted by any current therapeutics. The studies in this proposal will narrow the significant knowledge gap around the molecular mechanisms of basal complex function and may identify important targets for future antimalarials.

NIH Research Projects · FY 2026 · 2024-03

PROJECT SUMMARY This CoFAR application brings together seasoned clinical and laboratory investigators in food allergy (FA) with expertise in clinical studies and trials and translational immunologic, microbiome and metabolomic pathways. The investigators have long track records in implementing single sites and multi-site clinical trials and observational studies in allergic diseases, including FA, to the standards of NIH funded clinical research networks, in conducting NIH fundamental research on disease mechanisms in FA and in training generations of investigators in FA research. In part A we demonstrate that we have the personnel and facilities to conduct CoFAR network-wide and Clinical Research Center-specific research, and a great capacity to recruit subjects from all age groups and different racial and ethnic backgrounds. We have a highly experienced team, a very solid and rich clinical research and laboratory infrastructure, data management facility with quality control plans, and capability to upload data into the NIAID designated repositories. In part B our network-wide trial proposes to evaluate the safety and efficacy of microbiota transplantation therapy (MTT) in teens with peanut allergy. Our basic and translational studies have provided critical evidence on the role of microbiome manipulation in FA. Our cutting-edge phase I trial in adults showed very promising results. We hypothesize that MTT with and without low dose peanut oral immunotherapy will be safe in peanut-allergic teens, will increase their threshold dose of reactivity to peanut and will lead to immunological and microbiome changes. In Part C, we propose to investigate FA novel immunological biomarkers identified in our laboratories. Specifically, RORgt+ Treg cells play a critical in mediating oral tolerance while Th2 cell-like reprogrammed regulatory T (Treg) cells play an essential role in disease pathogenesis. The thymic stromal lymphopoietin receptor (TSLPR) expression is predictive of Th2 cell-like reprogramming and disease promotion. Resistin-like beta (RELMβ) is elevated in FA in inverse relationship with RORgt+ Treg cells and plays a critical role in disease pathogenesis by promoting anaphylaxis. We hypothesize that changes in Treg cell subpopulations and markers and RELMb play a critical in the outcome of natural tolerance or successful peanut desensitization. This project evaluates novel microbiome therapies and mechanisms fundamental to FA and pave the way for future intervention and biomarker driven approaches to inform future precision therapy. We address a critical need in investigating novel therapeutic approaches and immunological pathways associated with peanut allergy. We will contribute extensively to the CoFAR as a CoFAR-Clinical Research Center, with our infrastructure and expertise.

NIH Research Projects · FY 2026 · 2024-03

Summary Pyroptosis is a most “explosive” and immunogenic form of lytic cell death involving spillage of cellular contents, and has been defined as cell death mediated by the gasdermin (GSDM) family of proteins. This breakthrough understanding was brought about in 2015 when gasdermin D (GSDMD) was identified as a downstream effector of inflammasomes, which are supramolecular complexes that activate inflammatory caspases (-1, -4 and -5 in human and -1 and -11 in mouse). GSDMD gets cleaved by caspases to generate an N-terminal fragment (GSDMD-NT) and a C- terminal fragment (GSDMD-CT), and GSDMD-NT was shown to mediate pyroptosis, as well as the release of IL-1 family cytokines, which are processed by caspase-1 to the mature form. The GSDM family comprises six members in humans (GSDMA, GSDMB, GSDMC, GSDMD, GSDME/DFNA5, and DFNB59). In 2021, Ninjurin1 (NINJ1), a member of the NINJ family comprised of 2 transmembrane proteins in mammals, was shown to act downstream of GSDMD pore formation to induce the plasma membrane rupture required for full release of damage- associated molecular patterns (DAMP) such as lactate dehydrogenase (LDH). We and others found that upon cleavage by inflammatory caspases, GSDMD-NT specifically binds to acidic lipids, and exhibits strong membrane-disrupting cytotoxicity in mammalian cells by forming pores on membranes during pyroptosis and in vitro. Other GSDMs, upon cleavage by appropriate proteases, also form transmembrane pores. Here we propose to elucidate the structural mechanism of pore formation by the GSDM family, and of membrane disruption by NINJ1. Understanding how GSDMs are regulated and exert their pore forming activity, and how NINJ1 oligomerizes to induce plasma membrane rupture will not only provide new insights on pyroptosis, but also afford new therapeutic strategies for treating inflammasome-related and pyroptosis-related diseases.

NIH Research Projects · FY 2026 · 2024-03

Project Summary: Brain MRIs are widely used in children for diagnosis and treatment monitoring. A typical exam comprises multiple sequences with different contrast preparations, and often requires 20+ minutes to complete. Potential motion during such lengthy acquisitions necessitates sedation or anesthesia. However, repeated sedation or anesthesia in children increase the risk of long-term detrimental effects on cognitive development. 40% of these pediatric scans also require gadolinium (Gd) based contrast agent (GBCA) administration. It has been recently shown that Gd is retained in the brain and body, which may be particularly harmful to children since their developing brains are more susceptible to heavy metal exposure, and free lanthanides are known to be neurotoxic. Pediatric brain exams are excessively long because of inefficient use of parallel imaging technology that only provides R=2–3-fold acceleration. Acceleration in two dimensions, including the slice/partition axis, through controlled aliasing in parallel imaging (CAIPI) has enabled R=4–6-fold speed-up, and has become popular in functional/diffusion imaging using simultaneous multislice (SMS) encoding. Unfortunately, adoption of SMS in clinical sequences has been extremely slow. Acceleration via Compressed Sensing (CS) is a promising solution, but its availability as a product solution is variable among MR vendors, and it often comes at the cost of low-contrast image features. Deep learning (DL) has emerged as a powerful reconstruction and image enhancement tool. Vendors’ DL solutions include denoising and super- resolution enhancement, but these are limited to the newest software versions and host computers with GPUs. While promising a better trade-off between image quality and scan time, they are currently implemented for a small number of sequences on different vendors, and are therefore constrained by poor availability and scalability. Thus, current clinical technologies have been hampered by limited availability and faster has often meant suboptimal quality. Lastly, no vendor has DL-based solutions for Gd dose reduction for either adults or children. Given the need for repeated injections of sedatives and GBCA in children who are scanned periodically for treatment monitoring, there is an unmet need in imaging technology that makes these young populations vulnerable to severe and long-term health risks. We propose data acquisition, reconstruction and contrast enhancement strategies to address this unmet need. In Aim1, we will develop a rapid, comprehensive brain exam by combining our advanced controlled aliasing strategy, wave-CAIPI, and extend this to SMS encoding for rapid FLAIR/TSE imaging. Combining this with DL super-resolution reconstruction will enable R=9-fold acceleration with high fidelity to create a 6-min protocol. In Aim2, we will develop and validate a DL- based contrast enhancement algorithm to synthesize full dose images from 5× reduced Gd dose in pediatric exams using our rapid protocol from Aim1. To that end, our novel technologies would speed up the clinical MR exams and minimize both the amount of sedation and the injected contrast agents dose in children.

NIH Research Projects · FY 2025 · 2024-03

Project Summary Hypoplastic left heart syndrome (HLHS) is a fatal form of congenital heart disease characterized by the underdevelopment of left-sided structures, including the left ventricle (LV), mitral and aortic valves, and ascending aorta. These three structural deficiencies result in diminished circulation that invariably leads to morbidity without surgical intervention. It is currently thought that HLHS is an oligogenic disease of co- occurring structural defects caused by different genetic lesions. In some cases, however, one structural defect might be primary and the others secondary. As HLHS is clinically heterogeneous, there are likely multiple paths to disease pathogenesis caused by diverse, and often unknown, genetic etiologies. To begin studying the affected genes in the context of heart development, I mined published databases for de novo and rare inherited variants in HLHS probands for follow-up in zebrafish. I initially selected RBFOX2, a highly conserved RNA binding protein (RBP), based on its strong statistical link to HLHS and lack of study in cardiac development. I published that rbfox2 mutant zebrafish display all three HLHS-like structural heart defects that arise secondary to compromised pump function (Huang et al. Nat Commun 2022). However, unlike patients that are heterozygous for RBFOX2, heterozygous zebrafish and mice are healthy. Therefore, no appropriate model exists to investigate how halving the dose of RBFOX2 affects CM biology. Moreover, many variants statistically linked to HLHS remain unvalidated or unstudied in whole animal models. To address these knowledge gaps, I engineered and characterized RBFOX2 het and null human (h) iPSC-CMs. Both genetic cohorts show dose-dependent reductions in cell size, cell adhesion, calcium handling, and oxygen consumption rates. Myofibril alignment is also perturbed. Based on this phenotypic characterization and integration of several sequencing datasets, I propose to test the hypothesis that RBFOX2 directly regulates CM cell adhesion and growth by controlling transcript stability and splicing of an ECM-cytoskeletal-myofibril gene network in Aim 1. To identify additional HLHS-linked variants that cause cardiac phenotypes in zebrafish, I conducted a pilot CRISPR screen and recovered an ortholog of FOXC2, which encodes a transcription factor expressed in the endocardium and myocardium. Preliminary phenotyping revealed HLHS-like cardiac phenotypes in foxc2 global knock-outs that also fail to activate a Notch reporter in the endocardium. In Aim 2, I propose to test the hypothesis that endocardial-specific FOXC2-deficiency leads to primary valve and secondary myocardial defects caused by defective Notch signaling. Overall, I anticipate learning that both myocardial- and endocardial-intrinsic defects recapitulate specific aspects of HLHS. More broadly, implicating human variants as causal for HLHS pathogenesis and deciphering their mechanism of action in whole animal and human iPSC models will ultimately lead to better predictors of heart failure following surgical intervention and new therapeutic inroads for patient-specific clinical management of the disease.

NIH Research Projects · FY 2026 · 2024-03

ABSTRACT Congenital malformations are a major public health challenge. These conditions are often linked to maternal metabolic dysfunctions like diabetes and obesity. Yet, the molecular mechanisms that couple metabolism to the genetic programs that control embryonic development remain poorly understood. Neural crest cells are a type of embryonic stem cell that is particularly sensitive to metabolic perturbations and has been directly linked to multiple developmental abnormalities. Neural crest development is orchestrated by a complex gene regulatory network that endows these cells with their unique properties, like stemness, multipotency, and the ability to migrate. Our group has previously shown that proper deployment of this regulatory network depends on the initiation and maintenance of a metabolic state of increased glycolytic flux. We recently observed that this state of enhanced glycolysis contributes to the regulation of gene expression through a mechanism that involves a newly described epigenetic mark called histone lactylation. By examining the deposition of this mark, we identified cis-regulatory regions in the genome that respond to changes in the glycolytic state of neural crest cells. Notably, these putative metabolism-responsive enhancers (MREs) are located in the loci of neural crest genes that are upregulated upon metabolic reprogramming. Based on this preliminary data, we hypothesize that specialized cis-regulatory elements allow gene regulatory networks to respond to changes in cellular metabolism. We will test this hypothesis in three specific aims. First, we will characterize the patterns of genomic deposition of specific lactylation marks and test if these patterns change upon manipulation of metabolic state and lactate levels. We will define how these manipulations affect the organization of the epigenomic landscape and gene expression patterns. Second, we will examine how histone lactylation is deposited in the genome of neural crest cells. We will use a combination of genomics and functional assays to test the hypothesis that YAP/TEAD and SOX9 promote lactylation by cooperating with lactylation writers. Third, we will test if MREs respond to changes in glycolytic flux by performing STARR-seq in neural crest cells subjected to metabolic manipulation. Finally, we will use genome engineering to delete MREs in neural crest cells and test their requirement for transcription responses to metabolic reprogramming. These experiments will define how metabolic state affects the epigenomic landscape and modulates the gene regulatory networks that control embryogenic development.

NIH Research Projects · FY 2026 · 2024-03

PROJECT SUMMARY: Hemizygous microdeletions encompassing TBX1 cause 22q11.2 deletion syndrome (22q11.2DS), the most common deletion syndrome in humans. Among a broad spectrum of clinical features that often include craniofacial dysmorphia, congenital heart defects (CHDs) of the outflow tract (OFT), and aortic arch, which is derived from embryonic vessels termed pharyngeal arch arteries (PAAs), are the main causes of mortality during childhood. Importantly, specific craniofacial muscles, the OFT, and PAAs all derive from the pharyngeal apparatus, a transient series of arches and pouches that form by segmentation on the lateral surface of the head. Because segmentation is severely compromised in Tbx1-deficient animals, their craniofacial and cardiovascular defects were considered secondary. Using zebrafish as a model, my laboratory reported that tbx1 mutant embryos recapitulate several aspects of 22q11.2DS, including craniofacial muscle, OFT, and PAA deficiencies. We also demonstrated that these end structures descend from nkx2.5+ progenitors initially specified in the anterior lateral plate mesoderm (ALPM) before becoming sequestered in the cores of the pharyngeal arches. Unexpectedly, we discovered that tbx1 mutants fail to specify the nkx2.5+ pharyngeal lineage in the ALPM that gives rise to the missing derivatives. Because this phenotype precedes pharyngeal segmentation, defects in progenitor specification are likely causal for the deficiencies observed in tbx1 mutants. The aberrant fates of these progenitors, which maintain hand2 and gata5 expression in the ALPM, are unknown. Moreover, the molecular mechanisms by which Tbx1 directs specification of the pharyngeal lineage in the ALPM have not been elucidated in any model system. Here, we have the unique opportunity to uncover the molecular mechanisms by which Tbx1 directs specification of the pharyngeal progenitor cell lineage because we have defined the relevant developmental time window for analysis, identified markers of the preserved (gata5+) and missing (nkx2.5+) progenitor populations, implemented single-cell RNA and ATAC sequencing that allows for unprecedented resolution of changes in cellular populations and chromatin accessibility, and generated state-of-the-art genetic tools for tracking the aberrant fates of tbx1-expressing cells in tbx1 mutants and for identifying Tbx1 target loci that confer the pharyngeal progenitor cell fate. Overall, we propose to use the attributes of the zebrafish system, including its genetic tractability, unmatched embryonic accessibility, and imaging capabilities, to reveal new mechanistic insights that will provide the most comprehensive view of how Tbx1 is necessary and potentially sufficient for instructing the pharyngeal progenitor cell lineage during vertebrate development.

NIH Research Projects · FY 2025 · 2024-02

Project Summary/Abstract Pancreatic cancer (PC) is one of the most lethal forms of cancer in the United States (5-year survival rate below 10%). PC patients are usually diagnosed with a non-resectable disease (80-85%) and have a dismal prognosis with a survival period of only 3-6 months after the diagnosis. Nucleoside analogs (e.g., gemcitabine and fluorouracil) and platinum-based antineoplastics (e.g., oxaliplatin) are commonly used as chemotherapy for PC. When administered as free drugs, gemcitabine (Gem), fluorouracil (5-FU) and oxaliplatin (Oxa) display significant off-target toxicity causing life-threatening adverse effects in many patients. There is an urgent need to develop new therapeutic strategies consisting of delivery vehicles able to target PC cells, limit Gem, 5-FU and Oxa off-target toxicity and improve the overall anticancer response. We have recently patented a platform that uses an unbiased and quantitative screening algorithm for the discovery and validation of cancer-specific surface antigens. Our recently published results show that Intercellular Adhesion Molecule 1 (ICAM1), a transmembrane glycoprotein of the immunoglobulin superfamily, is aberrantly overexpressed in PC and can serve as a PC-specific target. Our results suggest that developing a novel ICAM1-based precision nanomedicine can be successfully utilized to treat PC patients. We have recently developed a magnetic extrusion technique to synthesize endosome-derived vesicles called exosome mimetics (EMs) in a highly efficient and reproducible manner. Our EMs share the same biological origin, morphology, nanosize and composition with exosomes, a class of natural cell-secreted extracellular vesicles. Our EM synthesis outperforms conventionally used exosome isolation and loading protocols in terms of particle yield, batch-to-batch consistency, reproducibility and loading efficiency. In this proposal,we leverage our expertise in cancer-specific antigen discovery and validation, bio- nanomaterial engineering, and exosome biology to test the novel hypothesis that EMs expressing ICAM1 nanobody and loaded with Gem, 5-FU and Oxa can be used as a novel delivery vehicle for PC therapy. We will synthesize EMs engineered with ICAM1 nanobodies that recognize and kill ICAM1-expressing PC cells and load them with Gem, 5-FU and Oxa. ICAM1-EM will exhibit increased tumor specificity and reduce Gem, 5FU and Oxa off-target delivery and toxicity. These innovative studies have the potential to lead to the development of novel EM-based therapies that can improve the efficacy of current cancer drug delivery. With key experimental tools, in vivo models and extensive experience in place, we will address the following Specific Aims: 1. To engineer ICAM1-targeted exosome mimetics (ICAM1-EMs) using magnetic extrusion method 2. To determine the efficacy of loaded ICAM1-Chemo-EMs in inhibiting PC growth and progression 3. To determine the pharmacokinetics (PK) and biodistribution of loaded ICAM1-Chemo-EMs

NIH Research Projects · FY 2025 · 2024-02

PROJECT SUMMARY Perinatal white matter injury (WMI) is a common sequela of neonatal brain injury. One of the leading causes of neonatal brain injury and morbidity in premature infants is post-hemorrhagic hydrocephalus (PHH). PHH is triggered by germinal matrix and/or intraventricular hemorrhage (IVH) that results in accumulation of cerebrospinal fluid (CSF) in the brain ventricles. This accumulation of CSF can cause compression of surrounding brain tissue and injury to the developing white matter. Specifically, in PHH the degree of ventricular dilation is correlated with severity of WMI as measured by decreased myelination and increased axonal injury, cellularity, and cytoplasmic vacuolation. Myelination in the central nervous system (CNS) has been previously identified as dependent upon Autotaxin (ATX), which can be sourced via the CSF. The choroid plexus (ChP) is the tissue which controls the amount and composition of the CSF and therefore helps instruct the developing nervous system. Here, we examine expression and availability of ChP-secreted ATX influencing white matter development and myelination of the mouse CNS and hypothesize this may be an important signaling axis which could be therapeutically harnessed to treat WMI associated with PHH. The Lehtinen Lab's single nucleus sequencing data demonstrates ATX expression across all epithelial cell clusters, thereby suggesting the epithelial cells may secrete ATX into the CSF. In preliminary studies we confirm that the ChP secretes ATX and show that ATX abundance in the ChP and CSF increase throughout development, with a particularly noticeable upregulation in the critical postnatal window associated with myelination of the CNS in mouse. In Aim 1, I will determine the effect of ChP-specific ATX modulation on myelination using advanced in vivo mouse manipulation techniques to selectively delete or overexpress ATX in the ChP. In Aim 2, I will further explore role for ATX in the pathophysiological mechanisms of WMI using the lab's translationally relevant mouse model of neonatal PHH following IVH. The results of these studies will inform our understanding of the mechanisms of hemorrhage induced WMI and will help guide rational intervention strategies for treatment, thereby improving quality of life for patients and caregivers. The proposed research will take place at Boston Children's Hospital under the guidance of Dr. Maria Lehtinen, an expert in the field of choroid plexus and CSF biology, and will provide training in ChP in vivo manipulation techniques, glial biology, exposure to clinical experience, and in the design of translationally relevant preclinical studies. We will routinely consult with collaborators and mentors who are experts on both white matter development and the neurologic sequelae associated with PHH of prematurity. The results from this proposal will result in first-authored publications and a large body of support for a K99/R00 application, with the goal of launching an independent academic career.

NIH Research Projects · FY 2026 · 2024-01

Project Summary Valvular heart disease is an important health problem afflicting over 2.5% of the US population and catheter- based therapies to address it have advanced significantly in recent years. While surgical repair remains the gold standard, the reduced risk of catheter-based interventions has provided the ability to intervene earlier in the disease process and in patients too sick for surgery while avoiding the risks of cardiopulmonary bypass. A significant limitation of these procedures, however, is that they do not provide the capability to remove native tissue and previously implanted devices to tailor the anatomy to receive a new prosthetic device. For example, transcatheter valves rely on displacing the diseased valve leaflets rather than removing them. In transcatheter aortic valve replacement, the displaced leaflets can obstruct blood flow to the coronary arteries and prevent access for subsequent coronary interventions. In transcatheter mitral valve replacement, the native anterior leaflet can be forced into the left ventricular outflow tract resulting in restricted flow into the aorta. Recently, a transcatheter technique has been introduced to lacerate a leaflet along its midline from base to tip. When performed prior to transcatheter valve replacement, the new valve spreads the two halves of the old leaflet apart so that they do not interfere with blood flow. This innovative approach can increase the number of patients who qualify for these low-risk procedures. Since the procedure relies on the use of existing guidewires and catheters, however, it is technically challenging, requires multiple clinicians to perform and takes more time than valve replacement. Furthermore, the electrosurgical ablation used in these procedures is not as effective for calcified tissue which is often present in native and bioprosthetic leaflets. To address this need, we propose to develop a catheter-based technology that enables a single operator to perform precise cutting of native and bioprosthetic leaflets regardless of calcification. In Aim 1, we will develop a steerable cardioscopically-guided leaflet cutting catheter. We will develop and demonstrate the technology in the context of aortic and mitral leaflet modification using electrosurgical cutting. In Aim 2, we will create a laser-based leaflet cutting system to address the limitations of electrosurgery in calcified tissue. The technology will be evaluated through in vivo testing and comparison with existing methods. Key innovations of this research include real-time optical visualization of leaflets during cutting, the ability to cut a leaflet without forming a wire loop through it, the development of laser-based cutting to precisely lacerate calcified tissue and the characterization of emboli produced by leaflet laceration. This technology can provide a platform for the future development of more sophisticated transcatheter tissue modification and device removal procedures.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY/ ABSTRACT Despite significant efforts to limit youth from accessing and using electronic cigarettes (e-cigarettes), many adolescents and young adults are directly purchasing e-cigarettes from brick-and mortar retail stores. Studies show that pervasive e-cigarette marketing in the retail environment such as e-cigarette product displays, location/size of advertising, and price incentives and coupons, increase adolescent e-cigarette susceptibility and actual use. However, studies have not directly asked adolescents and young adults to identify appealing and influential characteristics of e-cigarette marketing in the retail environment that impact their e-cigarette purchase and use intentions. Such data will support the FDA’s aim of understanding marketing influences on youth tobacco use and will inform the development of communications to prevent e-cigarette use through a counter-marketing lesson, addressing appealing e-cigarette marketing characteristics in the retail environment. The proposed project will address three Specific Aims towards the development of a counter-marketing lesson and regulatory solutions: (1) Examine adolescents’ and young adults’ descriptions of e-cigarette marketing in the retail environment and its influence on their e-cigarette purchase and use behavior. (2) Identify the most important, appealing characteristics intrinsic to e-cigarette marketing in the retail environment influencing adolescents’ and young adults’ intentions to purchase and use e-cigarettes. (3) AIM 3. Develop and evaluate the effectiveness of an e-cigarette counter-marketing lesson combating marketing in the retail environment to reduce intent to use and actual use of e-cigarettes among adolescents. In the K99 phase, focus group discussions (Aim 1) and surveys including an embedded discrete-choice experiment (Aim 2) will identify how and which e-cigarette marketing characteristics influence adolescent and young adult e-cigarette purchase and use. Aims 1 and 2 will identify which characteristics require counter-marketing and would benefit from strengthened regulation. In the R00 phase, a randomized controlled trial will randomly assign adolescents-only to one of two conditions: 1) an online counter-marketing lesson about e-cigarette marketing in the retail environment (developed in this phase) or 2) an existing online e-cigarette overview lesson to assess influence on adolescents' intent to use and actual use of e-cigarettes (Aim 3). This Award and proposed research will enable the PI to grow expertise in impactful e-cigarette prevention programs and policy solutions to reduce adolescent tobacco use, a long-term goal. A training plan involving mentorship from multidisciplinary tobacco control experts in tobacco prevention, marketing, policy, survey design, statistical methods and counter- marketing design, and complementary didactic training will fill gaps in knowledge of tobacco regulatory science and vital research skills, allowing the PI to transition to an independent investigator. By utilizing preliminary data and leveraging new skills, the PI will submit a R01 grant that will examine a multi-component intervention to combat marketing, which will substantially reduce adolescents’ and young adults’ e-cigarette use.

NIH Research Projects · FY 2026 · 2024-01

ABSTRACT Pulmonary arterial hypertension (PAH) is a life-threatening disorder characterized by elevated lung pressure, right heart failure, and premature death. Current therapies fail to prevent disease progression due to their inability to suppress and reverse obliterative lesions resulting from vascular remodeling. Previous studies have centered on large arteriolar remodeling in vessels with a diameter >100 µm. However, the role of the capillary bed (vessels <10 µm) in remodeling remains largely ignored. Pericytes (PCs) are indispensable mural cells for maintaining capillary integrity and homeostasis. Our preliminary data showed that the loss of PCs induced excessive capillary and arteriolar remodeling and manifested hypoxia-induced pulmonary hypertension and right ventricular hypertrophy in transgenic animals. Additionally, human idiopathic PAH (IPAH) PCs showed high proliferation and motility, and human IPAH induced-pluripotent stem cells (IPAH-iPSC) derived mural cells lost PCs but gained more smooth muscle characteristics. Vascular organoids derived from IPAH iPSCs further demonstrated a reduced number of endothelial cell (EC) tubes and increased smooth muscle coverage, which recapitulated abnormal vasculature development without PCs. Single-cell RNAseq revealed upregulated expression of the Regulator of G-protein signaling 5 (RGS5) in the IPAH PC cluster, which was validated in IPAH PCs isolated from explanted lungs, hypoxic mouse PCs, and explanted IPAH lung tissues. Thus, we hypothesize that RGS5 overexpression mediates PC dysfunction (detachment and migration) and, in turn, capillary remodeling in IPAH. We proposed to 1) Characterize the impact of PC dysfunction on EC dysfunction, vascular integrity, and remodeling in PC-depleted transgenic animals; 2) determine how RGS5 signaling regulates PC detachment, proliferation/migration, and EC interaction in clinical IPAH samples; 3) Characterize the impact of RGS5 depletion on PC/EC cell-cell interactions in IPAH iPSC vascular organoids. This project will provide novel insight into PC pathobiology in capillary remodeling and establish RGS5 as a potential therapeutic target in PAH, for which the first drugs to treat this devastating disease may be found.

NIH Research Projects · FY 2026 · 2024-01

The Heart Center (HC) at Boston Children’s Hospital (BCH) has been an enthusiastic and productive Core Clinical Research Center in the Pediatric Heart Network (PHN) since its inception in 2001. Our volume is large and in calendar year 2022 included 840 open heart operations, 1470 catheterizations, 29,045 outpatient visits, 29,135 echocardiograms, 1,366 cardiac MRIs, and 646 cardiac CT scans. HC faculty participate in 55 multicenter trials and registries, direct 18 noninvasive imaging core labs, and are PIs of data coordinating centers for 14 multi-center studies. The HC employs 6 data managers, 36 research assistants and coordinators, and 9 statisticians (4 PhD, 5 MS). Within the PHN, BCH cardiologists have been PI/MPI of 10 trials and cohort studies and directed 10 imaging core labs (9 echo, 1 MRI). BCH recruitment has been in the top quartile of participating centers for 9 of 14 PHN studies that actively enrolled participants in the current grant period, and in the top half for 13 of 14 studies. We have successfully recruited subjects on nights and weekends via a call schedule, and, with a large, world-class adult congenital program, can recruit and retain participants across the lifespan. BCH has state-of-the-art administrative and scientific capabilities that can expand the scientific productivity of the PHN. These include a model research pharmacy, rich experience in FDA-regulated studies, and world-class data science and data integration across multiple registries and databases that include study outcomes and covariates. Our renowned biomedical informatics program encompasses medical decision making, diagnosis, therapeutic selection, care redesign, clinical trials design, cardiovascular machine learning, and artificial intelligence. BCH remains committed to recruiting participants representative of our patient population. The proposed research protocol concept, a step-wedge cluster randomized trial of developmental care that leverages the strength of BCH in cardiac neurodevelopment, has the potential to improve outcomes for children with congenital heart disease. In summary, BCH has exemplary credentials to be a PHN Clinical Research Center and has unwavering dedication to the success of the PHN’s ongoing and future studies. In the next PHN cycle, BCH will contribute to innovative research through the scientific expertise of its investigators, success in multi-disciplinary collaboration, commitment to eliminating child health disparities in CHD, extensive administrative resources, and personnel and processes with a proven track record for data integration across multiple registries and databases.

NIH Research Projects · FY 2026 · 2024-01

SUMMARY Endothelial cell (EC) dysfunction is a key factor that promotes poor host defense, pro-thrombotic cardiovascular complications and bleeding during respiratory viral infections. The applicant’s laboratory has made long-standing contributions in vascular effects of sphingosine 1-phosphate (S1P), a circulating lipid mediator important for EC resilience. This protective pathway is preferentially activated by HDL-bound S1P, thus counteracting EC dysfunction and pathophysiology. However, the role of S1P in influenza viral infections in the context of thrombosis and bleeding complications is poorly understood. This proposal develops a novel paradigm to enhance vascular resilience while minimizing thrombotic and bleeding complications during virus-induced pulmonary injury. We have developed designer HDL-like nanoparticles that chaperone S1P, namely, ApoA1- ApoM (A1M)/S1P, for therapeutic activation of EC S1PR1 and suppression of vascular leak. A1M/S1P either alone or together with other barrier protective agents such as angiopoietin-1 (ANGPT-1) and prostacyclin (PGI2) enhanced EC barrier function in endothelial cells. In addition, the ApoA1 moiety of A1M suppressed cytokine- induced NFkB activation and inflammatory gene expression. We hypothesize that sustained activation of S1PR1/Gi signaling by HDL-S1P and cooperative interactions between EC protective pathways (ANGPT1/ Tie2 and PGI2/IP receptor) enhances pulmonary vascular recovery from respiratory viral infections without inducing prothrombotic transformation of the endothelium. Specific aim 1 will evaluate HDL-S1P activation of protective endothelial S1PR1/Gi signaling. Genetic mouse models of S1PR1 loss of function (EC knockout), gain of function (EC transgenic) and Gi-biased signaling (S5A phosphorylation-defective mutant), Apom KO or Apom TG mice will be analyzed to determine alveolar microvascular leak, integrity, thrombosis, bleeding and resolution of inflammation during influenza virus-induced pneumonitis. Single cell (sc)RNA-seq data from mice with viral pneumonitis will be deconvoluted to determine molecular mechanisms of paracrine signal networks between EC and pericytes important in microvascular resilience. The second aim will determine the mechanisms by which albumin-S1P/S1PR1 signaling in EC promotes thrombosis in the context of viral infection. We will use Tie2 agonists together with EC-targeted S1PR1 agonists to control bleeding in the setting of influenza infection. The third aim will develop combinatorial therapeutic approaches containing novel HDL-like nanoparticles and Tie2 activators to suppress EC injury and thromboinflammation during viral host defense responses. Designer HDL particles containing S1P that enhances the vascular resilience and HDL particles that carry stable prostacyclin (PGI2) analogs will be combined with Tie2 activators to suppress thrombosis and bleeding in lung on chip and mouse models. Together, this proposal aims to achieve a mechanistic understanding of EC pathophysiology during respiratory viral infections and develop novel therapeutic strategies.

NIH Research Projects · FY 2026 · 2024-01

Infantile nystagmus syndrome is an involuntary oscillatory movement of the eyes that begins in infancy and persists throughout life. It is visually and socially debilitating. The underlying pathophysiology is poorly understood, and no effective treatments exist. More than half of infantile nystagmus patients have an associated retinal or optic nerve disorder. Nystagmus is thus thought to develop secondary to poor vision, suggesting that proper development of the oculomotor system is dependent on visual input, particularly in the first two months of life. The long-term goal of this research is to understand the pathophysiologic mechanisms in nystagmus and identify potential new therapeutic targets. Extraocular muscles (EOMs) from patients with infantile nystagmus display a number of abnormalities, including decreased innervation, small neuromuscular junctions, and an increased proportion of slow myofibers. We have recently shown that these changes are also present in a mouse model of nystagmus—albino mice—and that the first changes are present as early as P10, before eye opening. The human samples also show central nucleation of muscle fibers and an increase in expression of the immature form of acetylcholine receptor on the fast myofibers. These changes suggest that there is continuous remodeling of the EOM innervation in nystagmus. Different EOM myofiber types are innervated by different subtypes of oculomotor neurons (OMNs), which in turn receive different premotor inputs. The “fast” and “slow” OMNs, which innervate fast and slow myofibers, respectively, have different roles in different types of eye movements. EOMs, OMNs, and premotor inputs are all dependent on each other for survival. This leads to the hypothesis that nystagmus results from improper development of the oculomotor circuitry. This proposal aims to test this hypothesis in a mouse model of infantile nystagmus syndrome by (1) determining whether OMN survival and subtype distribution are altered and how the developmental timing of any changes relate to changes in EOM innervation and nystagmus onset and (2) determining whether the premotor synaptic inputs to the OMNs develop abnormally in mice with nystagmus. Albino mice, which display spontaneous nystagmus, will be assessed for changes in EOM innervation and myofiber subtypes and corresponding differences in OMN survival and subtype distribution. Age of onset of nystagmus will be determined. Viral transsynaptic tracing will delineate the first-order synaptic connections onto the ocular motor nuclei. In all experiments, albino mice will be compared to wild-type littermates, obtained from heterozygous matings, to control for genetic background. Multiple developmental timepoints will be assessed to determine the point in development when changes associated with nystagmus occur; whether changes in the EOMs, OMNs, or premotor inputs occur first; and which changes precede nystagmus onset. This research will provide fundamental knowledge into the mechanisms of infantile nystagmus. Understanding where and when the abnormalities in infantile nystagmus first arise is the first step in identifying potential therapeutic targets or preventative interventions.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY/ABSTRACT Candidate: Dr. Medina Jackson-Browne, PhD is an environmental epidemiologist with primary research interests in the role of early-life environmental chemical exposures on the immune and respiratory systems in children. The goal of this proposal is to obtain training in the skills to build Dr. Jackson-Browne’s research career in pediatric clinical research. We propose several initiatives to strengthen Dr. Jackson-Browne’s career. Specifically, Dr. Jackson-Browne will gain new research skills and enhance her understanding of 1) the clinical trial process with Dr. Phipatanakul, MD, 2) train in early-childhood immune system development and respiratory physiology and 3) translate this research and training to position her for a future R01. To achieve this goal, she will obtain training in clinical trial design, database design and management, and advanced biostatistical methods. This will be accomplished with formal coursework, collaborative work, attendance at conferences, and guidance from established mentors/scientific advisors with relevant expertise. The proposed research and training plan builds the foundation for an independent research career that aims to address gaps in the understanding of timing and mechanism through which environmental chemical exposure affect respiratory health in early childhood. Research: Early-life exposure to environmental chemicals, particularly chemical’s known to bioaccumulate in humans, is of increasing concern in the development of asthma in children. Aeroallergen sensitization is a pivotal risk factor for developing persistent, progressive asthma throughout life and sensitized children have a higher prevalence of early-life wheeze and poor lung function. These potential associations are particularly relevant to populations with higher exposure to these chemicals and those at a higher risk for developing and/or dying from complications from asthma such as Black and low-income urban children. This proposal details a five-year plan to provide Dr. Jackson-Browne with the training and expertise to address gaps in the understanding of the timing through which perfluoroalkyl substance (PFAS) exposures may affect asthma development in children by leveraging infrastructure of an on-going pediatric clinical trial of 2-3-year-old infants at high risk of developing asthma (PI, Dr. Wanda Phipatanakul, MD). Environment: Dr. Jackson-Browne will be mentored by Dr. Phipatanakul, an expert in epidemiology, clinical trials, and clinical investigation in asthma and allergic diseases. She has assembled an extraordinary team of advisors, including Drs. Jessie Buckley, Stephanie Lovinsky-Desir and Suzanne Dahlberg who have committed their time, resources, and expertise to facilitate Dr. Jackson-Browne’s career development and successful completion of the proposed project. The academic environment created Boston Children’s Hospital and their affiliates provides an ideal training environment specific to her goals of achieving research independence.