Yale University

NIH Research Projects · FY 2025 · 2025-08

Malaria remains a significant global infectious disease burden. Common prevention strategies include vector control measures and long-acting chemoprevention for the most at-risk populations. Recently, World Health Organization guidelines for use of preventative antimalarials have expanded, both for demographic groups and regions where chemoprevention is acceptable. Data suggest mosquitoes refeed frequently in their lifespan. More population-wide drug use thus raises questions about mosquito antimalarial ingestion via human bloodmeals, and whether this could affect mosquitoes and/or parasites developing within them – a largely unexplored topic. Plausibly, antimalarials could affect parasite development and drug resistance selection within the mosquito, with critical drug resistance transmission implications. I have developed an experimental model for mosquito antimalarial exposure and have found detectable drug levels in mosquito circulatory fluid (hemolymph) several days post drug exposure, highlighting this phenomenon as an important and innovative area of study that I now propose to expand into P. falciparum-infected mosquitoes. Here, I will investigate the effect of antimalarials on parasite development, strain complexity, and drug resistance propagation within the vector. My overarching hypothesis is that antimalarials taken by humans can exert selective pressure on parasites progressing through vector stages. Broadly, my aims include assessing the impact of commonly deployed long-acting antimalarials on 1) basic mosquito fitness and vector-stage parasite development and 2) parasite genomic strain complexity and drug-resistant allele selection within mosquitoes. Combining use of controlled lab parasite and mosquito strains, as well as previously collected field samples from Burkina Faso and Senegal, will allow comprehensive analysis of mosquito antimalarial uptake and its consequences. My work will help inform mass treatment strategies and drivers of resistance, with research contributing to future work on malaria transmission-blocking strategies. I am a physician-scientist with a background in infectious disease bench research and global health. My career goal is to become an independently funded project leader researching infectious diseases affecting resource- limited settings, focusing on malaria. The proposed training will build expertise necessary to carry out the proposed project, including skills in bioinformatics and genomic analysis, malaria vector-stage pathogenesis and entomology research, and leadership and professional skills to run clinical studies and foster a successful academic career. My mentors support me with their expertise in malaria pharmacology, genomic epidemiology, and vector biology. I have a unique access to world-class lab resources at Yale and previously collected field samples from Burkina Faso and Senegal which will allow me to answer my research questions. My PhD background in blood-stage malaria pathogenesis is strong, but further training in vector and genomics research will further shape me into a well-rounded malariologist with an important niche and potential for R01 success.

NIH Research Projects · FY 2026 · 2025-08

PROJECT SUMMARY Uninsured children, many of whom are children in immigrant families—who have at least one parent born outside the US and make up over 25% of the US child population—may experience more health problems and may have limited health care access compared with children enrolled in health insurance. In 2019, when 6% of US children were uninsured overall, 28% of immigrant children were uninsured. Outreach and support that enable children to enroll in health insurance, along with as multilevel outreach efforts that promote enrollment in health services for which families are eligible, are urgently needed to increase access to health care and improve health outcomes for children. Dr. Julia Rosenberg is an Assistant Professor of Pediatrics at Yale University whose career goal is to become an independent investigator who applies community-engaged principles to evaluate and inform multi-level health interventions to improve health for children. Alongside her multidisciplinary mentorship team, Dr. Rosenberg has devised a career development plan for this proposed K23 award that will promote her successful transition to independence through formal coursework and mentorship in: (1) repeated cross-sectional analysis, longitudinal quantitative analysis, and causal inference; (2) economic evaluations; (3) mixed methods research; (4) grant and scientific writing; and (5) ethical research principles in community-based participatory research. The objectives of the proposed study that will accompany the career development plan are to evaluate the extent to which new health insurance eligibility opportunities are associated with changes in uninsurance and in utilization of preventive health care and to employ community-based participatory research methodology to identify multilevel barriers to and facilitators of healthcare-seeking behavior for families. In Aim 1, we will use repeated cross-sectional data from the National Survey of Children’s Health to determine the extent to which state-level health insurance eligibility opportunities are associated with changes in health insurance enrollment and utilization of preventive health care. In Aim 2, we will use electronic health record data to evaluate difference-in-differences of uninsurance, utilization of preventive health care, and health care costs for children within vs. outside the limited age range of expanded eligibility in Connecticut (≤12 years old vs. >12 years old). In Aim 3, we will apply the NIMHD research and socioecological frameworks to assess multilevel (individual, interpersonal, community, and societal) barriers to and facilitators of changes in healthcare-seeking behavior and health services use for families by conducting qualitative assessments of public hearing testimonies and completing interviews with key informants from the community via purposive sampling. Results from this evaluation can inform future support and outreach efforts to improve health for children and families and will establish a foundation for future multi-state, longitudinal evaluations of long-term health and economic outcomes related to health insurance eligibility expansion.

NIH Research Projects · FY 2025 · 2025-08

Abstract The survival of living organisms depends on the timely and accurate duplication of chromosomal DNA. Defects in DNA replication jeopardize genome integrity and organismal viability and are linked to numerous diseases ranging from developmental syndromes to cancer. Research in my laboratory is broadly focused on elucidating the mechanisms that control the onset of DNA replication. In eukaryotes, DNA replication begins with the binding of initiator proteins, the origin recognition complex (ORC), to DNA at replication origins. ORC subsequently recruits and deposits the Mcm2-7 helicase motor onto DNA as an MCM double hexamer to ‘license’ these origins for DNA replication. Much of our knowledge about origin licensing and DNA replication initiation is derived from studies using budding yeast as a model system, but how these steps are accomplished in metazoan systems remains ill-defined mechanistically. In the past, we have employed biochemical, biophysical, and structural approaches to uncover new, fundamental principles for how eukaryotic ORC operates to license replication origins. For example, we have determined high-resolution structures of ORC-containing assemblies in various functional states to define how ORC binds DNA and cofactors, and how these activities are regulated. More recently, we have reconstituted human origin licensing in vitro, which revealed that human Mcm2-7 can be loaded onto DNA through multiple pathways, suggesting that not all human origins may be licensed through the same mechanism. Despite these advancements, numerous questions persist with respect to the mechanisms that regulate origin recognition and origin licensing in multicellular eukaryotes, which will provide the basis for our research program in the next 5 years. Building on our strengths in biochemical reconstitution and in structural biology, we plan to resolve how metazoan MCM loading intermediates mature into MCM double hexamers through the various licensing pathways, how these pathways have evolved across multicellular eukaryotes, and how origin licensing occurs in different types of chromatin contexts. In the long-term, our studies will help explain disease mechanisms and aid in the development of new treatment modalities for diseases linked to dysregulated DNA replication initiation such as cancer.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY/ABSTRACT Aneurysmal subarachnoid hemorrhage (aSAH) is a devastating type of hemorrhagic stroke with high rates of morbidity and mortality, and after initial stabilization, the mainstay of care is supportive. Early elevation of pro- inflammatory cytokines in the central nervous system is associated with adverse outcomes after hemorrhage. Whether time-to-resolution of this pro-inflammatory phase is a determinant of outcomes remains uncertain. The goal of this research project and training plan is to provide Dr. Magid-Bernstein, an Assistant Professor of Neurology at Yale University and a Physician-Scientist with expertise in immunology and neurocritical care with the critical skills to become an independent investigator at the juncture of neuroimmunology and acute brain injury. Her career development aims are: (1) to learn advanced statistical methods to analyze neuroimaging, (2) to obtain expertise in advanced statistical techniques including analysis of large datasets and computational transcriptomic analysis, (3) to gain skills in managing clinical research cohorts and clinical trial design, and (4) to develop the leadership and academic skills to lead a research program. To achieve these goals, the PI has assembled a mentorship team of experts in translational research in inflammation and stroke, neuroimaging, and advanced biostatistical methods including transcriptomics. Our preliminary data show that pro-inflammatory cytokines including interleukin-6, interleukin-8, C-C motif chemokine ligand-2, and vascular endothelial growth factor are elevated in the cerebrospinal fluid (CSF) of patients with aSAH within the first 3 days after ictus. The proposed study will investigate the duration of the pro- inflammatory phase as a potentially modifiable determinant of outcome after hemorrhage. Our central hypothesis is that failure to transition from a pro-inflammatory state to a reparative and anti- inflammatory CSF profile in patients with aSAH will be associated with poor neurologic outcome, persistent hydrocephalus, and inflammation on magnetic resonance imaging (MRI). We will test this hypothesis by collecting serial CSF samples from 200 aSAH patients and measuring cytokines and numbers of immunologic cells within the CSF to characterize the trajectory of inflammation over time and its relationship to outcome (Aim 1). We will investigate the relationship between this immune profile, persistent post-hemorrhagic hydrocephalus on computed tomography (CT) images, and on vessel wall MRI (Aim 2). Finally, in an exploratory aim, we will investigate the relationship between the above inflammatory CSF profiles, hydrocephalus on CT imaging, inflammation on vessel wall MRI, and outcome (Aim 3). The proposed research is significant because no disease modifying interventions exist for critically ill patients with aSAH, and identification of CSF and imaging markers associated with poor outcome will enable development of targeted interventions to modify the course of this devastating disease. This project is innovative because it will identify multimodal markers of neuroinflammation after aSAH, paving the way for novel intervention.

NIH Research Projects · FY 2025 · 2025-08

7. Project Summary/Abstract Funding is requested for the purchase of the current state-of-the-art Orbitrap Astral mass spectrometer system. The Orbitrap Astral is the newest hybrid (quadrupole/Orbitrap/Astral) MS platform in its class with a novel Astral mass analyzer capable of fragment ion scanning above 200 Hz at a resolution of 80,000 at m/z 524, a leading-edge Orbitrap mass analyzer which enables scan speeds of up to 40 Hz at resolution up to 480,000 at 200 m/z, and sub-part-per-million mass accuracy. The ability to parallelize precursor/fragment ion detection in the two mass analyzers along with the ultrafast scan rates offered by this cutting-edge instrument enable >5-fold more MS/MS scans/second as compared to the best mass spectrometers currently housed in the Keck MS & Proteomics Resource, enabling incredibly deep coverage from single-shot proteomics experiments. The Orbitrap Astral MS will be coupled to a Vanquish Neo UHPLC system that has excellent peak separation efficiency and reproducibility, and the proposed MS system will have all the necessary hardware and software needed for rapid identification, profiling, and quantitative proteomics workflows. Based on the manufacturer’s data, we expect this instrument will be able to identify over 9,000 proteins in a 15-minute LC gradient or 12,000 proteins in a 60-minute gradient for complex mammalian extracts. This unprecedented depth of coverage over short gradients would enormously improve the Resource’s sample throughput for biomarker discovery, helping to minimize the current 3+ week sample backlog and ensuring that the Resource has the state-of-the-art mass spectrometry capabilities to accommodate additional investigators who would benefit from these advanced technologies. Addition of the Orbitrap Astral will also substantially increase sensitivity for the identification of low-level proteins and their post translational modifications and allow the Resource to push the envelope of single-cell proteomics. The advances incorporated into this instrument make it the ideal platform to support the LFQ, DIA, and PTM workflows that have become highly-requested services by the 142+ Yale and non-Yale investigators submitting >7,668 samples to the MS & Proteomics Resource in FY23 and FY24 to date. This application is supported by 28 investigators encompassing 8+ different disciplinary research fields and 40+ different projects, and the proposed eight Major users of the Orbitrap Astral MS include members of four NIH and National Centers at Yale (the Yale/NIDA Neuroproteomics Center, the Yale Cancer Center, the Yale Alzheimer’s Disease Research Center, and the Interstitial Lung Disease Center of Excellence). The strengths of this proposal are manifold and include continued very strong institutional support not only in the initial committed investment (with >$227K committed towards service maintenance costs), but also in long-term assurances to cover any deficit that may occur for the Keck MS & Proteomics Resource; the sustained, balanced operational budget of the Resource; and the continued usage of and productive collaboration with the Resource by hundreds of NIH-funded investigators annually.

NIH Research Projects · FY 2025 · 2025-08

Abstract The multi-layered cell envelope of Mycobacterium tuberculosis (Mtb) is a critical barrier to antibiotics and an important target for drug development. Disrupting its assembly increases bacterial susceptibility to existing antibiotics, creating opportunities for synergistic therapies. This proposal aims to elucidate the molecular mechanisms underlying envelope assembly, focusing on key membrane proteins, whose structures we have determined. Aim 1 investigates the cytoplasmic synthesis and transport of mycolic acids, essential fatty acids in the mycomembrane, by investigating the spatio-temporal localization of Pks13 and its interaction with fatty acid synthases. Aim 2 defines the physiological role of the essential efflux pump EfpA, hypothesizing it functions as a lipid transporter, and explores its inhibition as a therapeutic strategy. Aim 3 characterizes proteins that coordinate mycomembrane biosynthesis, particularly PgfA, to understand how this process is regulated for balanced cell growth. These studies will provide crucial insights into M. tuberculosis envelope construction, paving the way for novel, synergistic TB therapies that enhance the efficacy of existing treatments.

NIH Research Projects · FY 2025 · 2025-08

Summary The imbalanced chromosome number, or aneuploidy, is associated with various human diseases, including cancers and trisomic syndromes. The genomic imbalance in these aneuploid cells leads to copy number alterations (CNAs) for a large number of genes, affecting both proteomic abundance and lifetime. In the past five years, we have developed a set of reproducible mass spectrometry (MS)-based proteomic methods for measuring the abundance and lifetime regulations of thousands of proteins and their post-translational modifications (PTMs), such as phosphorylation. We have exploited these toolkits in analyzing several meticulously selected isogenic aneuploidy models and have discovered characteristic proteostasis and cell signaling pathways that challenge current paradigms in the field. Our future research will expand in several directions. First, we will distinguish and determine the proteomic remodeling and tolerance mechanisms in chromosome Gain and Loss type aneuploidies. We observed a notable absence of protein degradation regulation in maintaining protein complex stoichiometry in a model of Chromosome 3p hemizygous deletion, in which we detected changes in protein thermal stability. We propose to extend the study of proteostasis in Loss type aneuploidies that are clinically important but have previously been underexplored and to further profile the protein interactome reformation and perturbation through cross-linking MS and other techniques. Second, we will apply phosphoproteomics to identify the direct dosage effects of kinases and phosphatases and their downstream signaling events. A specific focus will be the functional characterization of phosphosites on Death domain-associated protein 6 (DAXX), a histone H3.3 chaperone, in the context of human trisomies. The profiled phosphorylation regulations will be summarized to expose druggable vulnerabilities associated with trisomic syndromes and cancer aneuploidies. Our third line of research will adopt a cross-species strategy to study aneuploidy and gene CNAs. We will measure homologous aneuploid chromosome-induced effects in terms of protein and phosphosite abundance and turnover and apply an evolutionary biological perspective to understanding genome dosage imbalance. Finally, more broadly, we will develop bottom-up quantitative technologies based on limited proteolysis and matrix-assisted laser desorption ionization imaging mass spectrometry to assess how the native lipid environment impacts protein conformation and function in disease states. Collectively, our proposed technology developments and biological studies over the next five years are poised to significantly advance our understanding of the physiology and biology of genome imbalance stress and aneuploidy-associated abnormalities.

NIH Research Projects · FY 2025 · 2025-08

Targeting a novel glyco-immune checkpoint for cancer immunotherapy ABSTRACT: Immunotherapies have revolutionized cancer treatment in recent decades, but most patient responses are of limited duration, and primary or secondary resistance is the norm. There is an urgent need to identify additional immune checkpoints and develop inhibitory drugs to allow immune attack of established tumors. Checkpoints typically function through cell-cell contact between ligand-receptor pairs, so identifying novel interactions is of great benefit. To this end, we constructed a protein library comprising the extracellular domains of 512 molecular species from IgSF and TNFSF families for specific protein-protein interaction screening. Using these techniques, we identified the novel interaction between IgSF members VSIG4 and Siglec-7. Of note, these proteins diverge significantly from non-primates. Our data implicate Siglec-7 as an innate immune inhibitory receptor for NK cell activation, suggesting that the Siglec-7-VSIG4 interaction functions as an immune checkpoint. Here we propose to develop in vivo models to elucidate the mechanisms of function of these novel human-specific interactions. We will utilize our humanized mouse platform, MISTRG6-A2, which affords us the opportunity to manipulate innate (and adaptive) immune cells in vivo. Given the overexpression of VSIG4 on multiple myeloma (MM) cells (especially after lenalidomide treatment) and acute myeloid leukemia inv(16) cells (AMLinv(16)), we will explore the impacts of the Siglec-7- VSIG4 checkpoint on these malignancies in vitro and in vivo. Completion of these aims will enhance our understanding of immune activation and tumor immuno-evasion, as well as pinpoint the disease(s) most relevant for future clinical development of immune checkpoint inhibitory antibodies. Intervening on novel checkpoints will open the door to new therapies with the capacity to improve patient outcomes in MM and AMLinv(16), with potential applicability across many other tumor types.

NIH Research Projects · FY 2025 · 2025-08

Abstract The complex relationship between HIV, HCV and opioid use disorder has resulted in an acceleration of several inter-related public health emergencies that include new HIV and HCV infections and unparalleled overdose deaths in the United States. Despite a number of evidence-based practices to address these inter-related epidemics, they are often not adequately scaled to meet the unique needs of the specific community. While evidence-based practices such as opioid agonist therapies, syringe services programs, pre-exposure prophylaxis and antiretroviral therapy are core elements of an HIV prevention and response toolkit, these interventions are often unavailable or under-scaled in many settings. Moreover, other evidence-based practices like overdose education and naloxone distribution and direct-acting antiviral medications for HCV may not tackle HIV related outcomes, but could influence opioid and HCV outcomes, respectively. Determining the scale of each intervention within the constraints of a specific intervention allows a multi-pronged approach to addressing overlapping conditions. Public health and policy experts need new and comprehensive tools to plan an effective strategy to address inter-related risks for diseases that are often financed through different funding streams. Our proposal aims to guide the public health and service planning to response to diverse epidemics. Specifically, this project will: 1) Develop a cost-based epidemiological planning tool to identify the optimal portfolio of EBPs to disrupt the adverse consequences of the HIV, HCV and OUD epidemics in the 50 states. The model will be implemented as an interactive, web-based dashboard to assist policy makers in planning an effective response and provide improved estimates of state-wide and local epidemic trajectories; 2) Conduct early usability and feasibility testing of the dashboard by collaborating with a team of state and local public health officials in order to optimize the future implementation of this dashboard. This portal will disseminate the epidemic model that can be used by local public health experts to test the impact of various HIV and HCV rapid responses through simulation. The proposed study may help guide a more efficient allocation of scarce resources to stem the HIV epidemic and consequently improve health outcomes among PWID in a high-risk HIV setting. As part of this exploratory R21, future directions will include assessing implementation strategies to improve use of the newly developed dashboard to guide a public health response.

NSF Awards · FY 2025 · 2025-08

The field of topology dates back at least to Poincare in the early 1900's, who was well aware of the interaction between geometric shapes ("manifolds") and dynamical systems, which evolve with time according to set rules. These insights, initially motivated by the dynamics of the solar system, became a far-reaching theory in areas of math both near and far to physics. Today mathematicians and scientists often study flows in manifolds, in which points travel in intricate orbits constrained by the overall shape of the manifold, as well as foliations, which are decompositions of a manifold into lower-dimensional slices like leaves of a layered pastry. This project aims to study such phenomena and build the understanding of the intricate connections between geometry and dynamics which occur especially in "low dimensional" settings such as surfaces and three-dimensional manifolds. Experience has shown that these low-dimensional settings, amenable to visual and geometric intuition, are also a valuable testing ground and provide inspiration for many more general phenomena within and outside of mathematics. Graduate students funded by the grant will, in addition to their research work, be trained in mathematics education and outreach, preparing them for contributions to society through higher education and other leadership roles. Additional activities of the PI and his students and collaborators will include public educational events for local school children and their families, bringing cutting-edge mathematical knowledge to the greater New Haven community. In this project, the principal investigator (PI) will explore a number of connections among foliations, flows, dynamics and hyperbolic geometry in low dimensions. One topic will be the topological dynamics of horospherical flows in infinite-volume manifolds. Building on recent successes of the PI and coauthors in the case of Z-covers of compact surfaces, the project will seek to extend results to higher rank abelian covers, higher dimensions and other generalizations. The PI also plans to revisit connections between two mature subjects in the setting of 3-manifolds: hyperbolic geometry on one hand and the topology and dynamics of taut foliations and pseudo-Anosov flows on the other. The goal here is to obtain a more robust and uniform theory relating topological features of foliations and flows to geometric features of the hyperbolic metric. In a third direction, the PI will study problems associated to Thurston's skinning map and the construction of uniform models for hyperbolic 3-manifolds. A motivating conjecture here is a uniform bound on Thurston's skinning map under certain topological hypotheses, for which the PI and coauthors have a promising line of attack. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY The largest determinant of survival and good neurologic outcomes in patients successfully resuscitated after cardiac arrest (CA) is the burden of hypoxic-ischemic brain injury (HIBI), a result of both the primary anoxic event and accrual of secondary brain injury. Mitigating secondary brain injury is critical to improving neurologic outcomes in this devastating disease. The lack of early, deployable biomarkers to phenotype CA patients has stifled advancements in therapeutics. Conventional high-field MRI is a guideline recommended component of neuroprognostication. A diffuse pattern of restricted diffusion on MRI is a reliable predictor of severe HIBI and functional dependence. Lack of access to MRI amongst critically ill CA survivors has led to underutilization, hindering the development of imaging-based biomarkers and intermediate endpoints. Low-field portable MRI costs a fraction of high-field MRI, is FDA approved, does not require magnetic shielding, and can be deployed in the emergency department and intensive care unit without disruption of vital patient monitoring. The proposed protocol will provide the scientific community with the largest systematically ascertained prospective observational cohort of low-field portable MRI in CA patients. To ensure the acquisition of clinically useful images, in agreement with conventional high-field MRI, we will evaluate the inter-rater agreement of low-field and high-field MRI for the evaluation of HIBI in 60 participants. Parallel to this, we will acquire ultra early (≤ 6 hours) and early (12-24 hours) low- field portable MRI to establish the positive predictive value and specificity of diffusion weighted imaging lesions for predicting HIBI on conventional MRI. Unlike focal ischemia, global ischemia, as seen in HIBI, progresses over days and may result in delayed apparent diffusion coefficient signal changes. Low-field portable MRI provides a unique opportunity to evaluate the time- dependent nature of HIBI on MRI. The development of early MRI-based HIBI biomarkers requires firm knowledge of the temporal and spatial variability of HIBI, including identification of early findings that persist on delayed imaging. This proposal is the first step towards developing a novel neuroimaging tool, low-field portable MRI, to understand HIBI mechanisms, therapeutic targets, and prognosis in patients resuscitated from CA.

NIH Research Projects · FY 2025 · 2025-08

Summary Preeclampsia is a hypertensive disorder of pregnancy that causes 70,000 maternal and 500,000 neonatal deaths per year globally. The only specific treatment is delivery of the baby and placenta, resulting in 3-4 times more frequent preterm birth compared to normotensive pregnancies. The underlying pathophysiology of preeclampsia is likely heterogeneous, but the culmination is maternal endothelial dysfunction causing hypertension, kidney failure, coagulopathy, and strokes. A history of preeclampsia is associated with ischemic heart disease and renal failure for the mother and long-term cardiovascular disease in the offspring. Therapies that specifically improve maternal endothelial function are urgently needed to permit safe pregnancy prolongation and improve mother and child outcomes. Plasminogen activator inhibitor 1 (PAI1) is elevated in preeclampsia, yet its pathophysiologic role is unknown. PAI1 plays a deleterious role in numerous cardiovascular diseases, and pharmaceuticals are in development. We and others have shown that PAI1 promotes endothelial dysfunction through inhibition of endothelial nitric oxide synthase and promotion of inflammatory and vasoactive genes that are upregulated in preeclampsia. PAI1’s role in hemostasis is important in pregnancy, but our preliminary findings suggest that PAI1-mediated inflammation is independent of its role in the coagulation cascade. These data suggest PAI1 contributes to the pathophysiology of preeclampsia and make it a promising therapeutic target. The objective of this proposal is to gain insights into maternal endothelial dysfunction that can be leveraged to develop novel therapeutic strategies with a specific focus on PAI1 as a drug target. In Aim 1, we test whether PAI1’s inflammatory effect can be blocked without disrupting its role in hemostasis by identifying how PAI1 alters inflammatory signaling pathways and determining the molecular interactions required. In Aim 2, we test the relevance of PAI1-mediated endothelial dysfunction in plasma from patients with preeclampsia using a PAI1 inhibitor. We use statistical models to associate clinical and biochemical characteristics with the effect of preeclamptic plasma on endothelial function. In Aim 3, we use a mouse model that recapitulates endothelial dysfunction in preeclampsia to test the role of PAI1 in mediating systemic effects including blood pressure, vascular function, cytokines, and renal damage. We test the in vivo efficacy of blocking PAI1 therapeutically. There is no treatment for preeclampsia, which leads to long-term cardiovascular disease for mother and child. This project mechanistically investigates the therapeutic potential of targeting PAI1 to improve endothelial function while also collecting new hypothesis-generating data about the effect of plasma proteins in preeclampsia on the endothelium. Our studies will drive future drug discovery to specifically target endothelial function with the goal of prolonging pregnancy and improving the health of the mother and baby.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY RNA chemical modifications are critical regulators of gene expression through regulation of RNA stability, splicing, translation, structure, and localization. Because of their important roles in several biological pathways, RNA modifications are frequently implicated in disease. N6-methyladenosine (m6A), the most prevalent internal RNA modification in eukaryotic messenger RNA (mRNA), has been extensively studied and implicated in a wide-array of human diseases, most notably cancer. Due to the established role of m6A in cancer, m6A and its associated machinery have been proposed as potential diagnostic and predictive biomarkers and therapeutic targets. Although the role of m6A in several cancer is relatively well-established, its regulatory role in glioblastoma (GBM) pathogenesis remains largely unclear. GBM, a grade IV glioma tumor, is the most common primary malignant brain tumor. GBM is a devastating disease associated with poor patient prognosis, limited therapeutic options, and frequent recurrence with acquired therapy resistance. Because m6A has been extensively demonstrated to play an important regulatory role in cancer, the goal of my proposal is to characterize molecular mechanisms by which m6A regulates GBM pathogenesis, and uncover how manipulation of m6A regulation may improve GBM treatment. The first portion of my research will examine the mechanism by which m6A post-transcriptionally regulates MGMT, a critical biomarker and predictor of chemotherapy response in GBM. To accomplish this, I will map m6A sites on MGMT mRNA, determine how disrupting m6A installation on MGMT affects MGMT expression and stability, and identify reader proteins which facilitate this effect. I will then determine the consequences of this regulatory mechanism on chemoresistance of glioma cells. In the second portion of my research, building upon preliminary RNA sequencing data, I will investigate mechanisms transcriptome-wide as to how m6A regulates the transcriptome of glioma cells in response to chronic treatment with an alkylating agent. To accomplish this, in chemoresistant glioma cells, I will (1) map m6A sites, (2) temporally sequence RNAs to measure the effect of m6A perturbation on RNA stability, and (3) sequence the interactome of canonical downstream regulators m6A-modified transcripts. Together, these complementary studies will increase our understanding of mechanisms by which m6A regulates GBM pathogenesis and reveal vulnerabilities that can be targeted with m6A-mediated therapeutics.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY Synucleinopathies are associated with increased phosphorylation of specific α-synuclein residues, such as Serine 129 (pS129) in Parkinson's Disease. Recent work has suggested that phosphorylation of α-synuclein is also relevant to its physiological function in synaptic vesicle (SV) trafficking. For example, pS129 was shown to be neuronal activity-dependent and allows α-synuclein to differentially associate with lipid membranes to ultimately regulate synaptic transmission. To better understand the mechanisms of synucleinopathies, the impact of other α-synuclein post-translational modifications (PTMs) on both pathology and physiology must be determined. We are interested in investigating the impact of phosphorylation at Threonine 81 (pT81), a modification that was discovered, by unbiased proteomics, to be enriched in the midbrain of Multiple Systems Atrophy (MSA) patients. To investigate pT81, we developed and validated a phospho-specific polyclonal antibody against pT81 α-synuclein and confirmed the presence of pT81 α-synuclein in intranuclear, neuronal inclusions in MSA patient tissue. Interestingly, a recent phosphoproteomics screen revealed pT81 to be a novel, activity-dependent phosphorylation site on α-synuclein that modulates SV trafficking. Hence, we hypothesize that pT81 PTM, similarly to pS129, plays both a role in synucleinopathy pathology and physiological SV dynamics. Our long-term goal is to study pT81 in vivo to understand its physiological relevance as well as how it may contribute to pathology in MSA. To achieve this, we will use CRISPR/Cas9 editing to create a phosphorylation-null T81A α-synuclein knock-in (KI) mouse and a phosphorylation-mimetic T81E α-synuclein KI mouse (Aim 1). We will then begin to characterize these models by assessing heterozygous and homozygous KI mice for α-synuclein expression and phosphorylation at T81 and S129 (Aim 2). Currently there are limited in vivo tools to study the causal role of α-synuclein PTMs, which are widespread across synucleinopathies and may be crucial to synaptic function. By utilizing both a phospho-null and a phospho-mimetic mouse model, we can elucidate both the importance of this PTM to synaptic transmission as well as the effects of upregulated pT81 in MSA.

NIH Research Projects · FY 2026 · 2025-08

PROJECT SUMMARY | Candidate: The candidate is an Instructor of Medicine (Nephrology) with 3 years of ongoing training in clinical research in acute kidney injury (AKI) and clinical trial design and conduct under the mentorship of the director of Yale’s Clinical and Translational Research Accelerator, a leading expert in clinical trials and pharmaco-epidemiology in AKI. This award will also allow her to receive mentorship from renowned experts in heart failure translational research, renal physiology and biostatistics to eventually become an independent clinical researcher in cardiorenal syndromes. Proposed Study: More than 1/3rd of patients hospitalized with acute heart failure (AHF) develop AKI, which is an independent risk factor for cardiovascular and kidney disease progression and mortality. AKI in this setting, often known as acute cardiorenal syndrome (CRS), is a cycle of venous congestion and overactive sodium reabsorption mechanisms characterized by diuretic resistance leading to prolonged patient distress and interruptions of essential HF therapy. The long-term goal of this study is to assess the efficacy of sodium-glucose cotransporter-2 inhibitors (SGLT2i) for acute CRS. SGLT2i are oral anti-hyperglycemic drugs which target a key sodium and glucose reabsorption mechanism in the kidney that have consistently slowed long-term kidney and cardiovascular disease progression in randomized clinical trials, independent of patients’ diabetes status or heart failure type. There is also pre-clinical evidence supporting their kidney tubular and endothelial protective and reparative effects in AKI. However, the efficacy and safety of SGLT2i in humans with acute CRS is unknown and AKI in this setting is a frequent reason for discontinuation. In a multicenter cohort study, we have shown that SGLT2 inhibition during AHF-associated AKI is not associated with prolonged AKI. By inhibiting an energy- demanding sodium reabsorption mechanism in the kidney’s proximal tubule, SGLT2i particularly when added to standard loop diuretic therapy, could promote energy efficient diuresis by reducing O2 demand in the kidney’s already O2-deprived environment, reduce oxidative stress and promote tubular integrity which may enhance tubular secretion of loop diuretics and their efficient delivery to their downstream target. Based on these assumptions, we hypothesize that SGLT2i improve diuretic response and promote quicker kidney recovery in patients with acute CRS which we will test as follows. In Aim 1, we will assess the impact of SGLT2i on the clinical improvement of individuals with acute CRS in a randomized placebo-controlled clinical trial of hospitalized adults with AHF-associated AKI by comparing established symptomatic and biochemical metrics of diuretic response. We will also examine safety and tolerability. In Aim 2, we will further assess the impact of SGLT2i on early biomarkers of kidney tubular injury and health. This study has the potential to improve the lives of patients with heart failure by establishing SGLT2i as a therapy for this challenging to treat syndrome and the potential to mitigate unnecessary interruptions of SGLT2i.

NIH Research Projects · FY 2025 · 2025-08

Project Summary/Abstract Both a driver of climate change and an ingredient in bubbly drinks, carbon dioxide is a small, volatile gas imperceptible to humans. Many insect species, however, possess specialized receptors capable of sensing changes in carbon dioxide levels in their environment. Behavioral responses to carbon dioxide vary significantly based on species and context. For instance, bees utilize elevated carbon dioxide levels as a signal to flap their wings to ventilate their hives whereas flies avoid the gas unless searching for a food source. Additionally, a number of disease-carrying insects such as Anopheles gambiae (malaria), Aedes aegypti (yellow fever and dengue virus), and tsetse flies (sleeping sickness) use carbon dioxide as a cue for host detection. Carbon dioxide sensing is critical for many insect species, yet the underlying molecular mechanisms are not fully understood. Individual receptor subunits have been identified but their contributions to ligand recognition and channel gating remain unclear. Uncovering the mechanisms of host carbon dioxide detection would allow for the development of novel antiviral strategies against some of the world’s deadliest diseases. The proposed project aims to determine the molecular basis underlying carbon dioxide detection through three aims. In aim 1, we will determine how to express and purify carbon dioxide receptors. In aim 2, we will obtain a structure of the Aedes aegypti carbon dioxide receptor and finally in aim 3, we will compare ligand binding pockets of two species, Aedes aegypti and Drosophila melanogaster. By understanding species-specific binding pocket differences, we can develop compounds that target the receptors of disease-carrying insects with high specificity. Together, these three aims will establish the molecular mechanisms underlying carbon dioxide detection and widen strategies for combating vector-borne diseases.

NIH Research Projects · FY 2025 · 2025-08

Project Summary/Abstract Cluster-randomized trials (CRTs) offer a powerful experimental design to evaluate the impact of complex interventions in medicine and public health, and have become among the most common study designs for embedded pragmatic trials seeking to improve patient care. Unlike individual-randomized trials, CRTs randomize intact clusters of subjects to alternative conditions, resulting in correlated observations that require specialized inferential tools. To maximize the patient-centered information gleaned from CRTs, there is an increasing interest in investigating the net treatment effects defined through win estimands (net benefit, win ratio, win odds) on multiple—prioritized and non-prioritized—outcomes that capture benefit or risk. However, existing statistical methods in CRTs have been primarily developed to address a single outcome measure. Principled methods that leverage the power of win statistics to guide the analysis of multiple and composite outcomes in CRTs are sparse, and their formal development requires addressing several methodological challenges. First, CRTs present a multilevel data structure that necessitates clear definitions of estimands. However, the existing estimands in CRTs do not address pairwise comparisons with multiple outcomes. Second, while covariate adjustment is a promising technique to improve estimation precision with a single outcome, robust and efficient covariate adjustment methods when targeting win estimands in CRTs remain under-developed, leading to missed opportunities in optimizing trial efficiency. Third, there is a lack of regression tools that enable the association analyses between win parameters and covariates when multiple outcomes collected from different individuals are correlated. To overcome these challenges, we will develop a suite of novel methods using clustered win statistics by maximizing information extraction from both covariates and outcomes, and apply our new methods to several CRTs that assess electronic health record alerts interventions for patients with heart failure. In Specific Aim 1, we will leverage the semiparametric efficiency theory of U-statistics to develop model-robust and efficient covariate-adjusted win estimators in CRTs with multivariate outcomes. In Specific Aim 2, we will expand the covariate-adjusted win estimators to censored time-to-event outcomes, and capitalize on the dual role of covariates in simultaneously addressing censoring bias and improving efficiency. In Specific Aim 3, we will develop marginal win regression methods to study the associational effects of covariates on win fractions under cluster correlated data, when the outcomes are either quantitative, or censored time to event. In Specific Aim 4, we will develop open-source software packages, tutorial papers, case studies, and short courses for all proposed methods, and use the NIH Pragmatic Trials Collaboratory as a unique national channel for effective dissemination. This work promises to advance the knowledge gained from past and future CRTs with complex outcomes by substantially expanding the toolbox for addressing win estimands in the multilevel data setting.

NIH Research Projects · FY 2026 · 2025-08

PROJECT SUMMARY Placental insufficiency- caused by deficient extravillous trophoblast invasion and/or syncytiotrophoblast nutrient transport and inadequate blood flow to the placenta- is a central hallmark in numerous obstetrical diseases, including fetal growth restriction (FGR) and preeclampsia. These diseases each affect 3-8% of pregnancies worldwide and have lifelong complications for both the mother and baby. While placental insufficiency is a common pathological feature, whether this arises from intrinsic placental developmental defects or altered maternal signals, eg. during viral infection that increases FGR and preeclampsia risk, remain unknown. Findings from our lab and others have implicated maternal endometrial dysfunction in the early pathophysiology of FGR and preeclampsia. Endometrial stromal cells are the major cell type of the endometrium and they undergo decidualization spontaneously every month in preparation for pregnancy. Decidualized endometrial stromal cells (decidual cells) promote endometrial receptivity and regulate placentation and vascular remodeling at the maternal-fetal interface. These effects can be mediated by decidual secretion of extracellular vesicles (EVs) that contain specific miRNA cargos. We have shown that exposure to viral double-stranded RNA (dsRNA) impairs decidualization and alters the miRNA cargo of decidual EVs. These decidual cells and EVs negatively impacted trophoblast stem cell differentiation and angiogenesis in vitro and in vivo. Based on these findings, our central hypothesis is that uncontrolled uterine inflammation alters decidual EV miRNA cargo which impairs trophoblast stem cell differentiation to syncytiotrophoblast while increasing uterine vascular dysfunction, leading to placental insufficiency. We will take a targeted hypothesis-driven approach to examine the effects of decidual EVs, and EV-associated miR-30a-5p and miR-206, on trophoblast stem cell differentiation, syncytiotrophoblast function (Aim 1) and uterine vascular functions (Aim 2) using physiologically-relevant trophoblast models, 3D models of angiogenesis and ex vivo vascular studies. Finally, we will confirm these findings clinically by examining markers of decidual inflammation and altered EV cargo in biospecimens from patients with FGR and preeclampsia (Aim 3). Successful completion of this project will highlight the importance of maternal decidual health for pregnancy success and reveal decidual EVs as an early pathological player in the pathogenesis of FGR and preeclampsia. The long-term goal of this research program is to identify new opportunities for the development of early biomarkers and therapeutic strategies to lessen the immediate and lifelong impacts of placental insufficiency.

NSF Awards · FY 2025 · 2025-08

Measurements of neutral hydrogen in the Universe with radio telescopes can inform our understanding of dark energy, a mysterious component in our Universe causing its expansion to accelerate today. To make these measurements, astronomers must understand the radio telescopes very precisely. In a previous grant, researchers from Yale University and West Virginia University, in collaboration with Canadian astronomers, developed a radio calibrator source using a new, fast chip that can be flown on a drone with signal to noise good enough for this precise calibration. Previously, the researchers focused mainly on development and testing in the lab. They will now upgrade the source, expand its capabilities, and focus on using it to calibrate radio telescopes by deploying it on the telescopes and drones. They will also develop a more robust lab version of a radio receiver outreach lab, used primarily for high school teachers and students. New 21cm interferometers targeting measurements of dark energy, reionization, and the dark ages require precise calibration, particularly of the instrument beam and gain, to remove bright foregrounds and extract the cosmological signal of interest. Currently, the incoherent (power only) calibrators developed to address this challenge limit the dynamic range of the measurement and also have no direct sensitivity to instrument phase. This research team has developed, tested, and validated a digital calibration source that addresses these critical gaps to make full-sky, high signal-to-noise measurements of the instrument beam. The researchers propose to use newly available versions of a Xilinx RFSoC board to update this source for wider bandwidth, improved stability, develop the ability to use multiple such sources simultaneously, deploy these sources on drones to calibrate new telescope arrays particularly well suited to drone beam mapping, and explore gain stabilization with this source. They also propose to leverage efforts already underway in an NSF-funded radio instrumentation outreach program (DSPIRA program) to continue developing education-oriented radio receivers, which can be used in STEM programs at WVU and Yale. Graduate students supported by this ATI grant will use the DSPIRA receivers as a prototype to make a more robust lab version appropriate for outreach activities, based on their experiences building a week-long outreach module for the Pathways to Science program at Yale. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NSF Awards · FY 2025 · 2025-08

Algorithms, cloud computing, and data define the infrastructure of the modern digital economy. The prices users pay for computation, software services, and information must reflect the true value to society of these goods. This is necessary in a market economy both because such prices help foster innovation and because they lead to efficient use of resources. This research analyzes three linked questions: (i) how to price the tokens that govern access to large language models; (ii) how to design cloud-computing contracts that reward sustained but flexible demand; and (iii) how platforms can share and aggregate data while respecting users’ private information. By developing rigorous economic models and translating the results into actionable pricing rules, the project advances the efficient allocation of digital resources, informs regulatory and antitrust debates, and supports workforce development through graduate training and open educational materials. The investigators build and solve mechanism-design models that capture the multidimensional nature of modern digital services. The first component studies a monopolistic provider of a specific kind of AI service (large language models) that sells finite-input, finite-output, and function tokens. This project uses a Cobb–Douglas production framework to derives cost-based nonlinear pricing plans. The team also characterizes welfare outcomes, and identifies empirically testable pricing ratios. The second component models sequential cloud-compute contracts in which users commit to future demand but retain real-time flexibility. This component shows that two-part and budget contracts implement the revenue-maximizing allocation subject to incentive and participation constraints. The third component analyzes data markets in which platforms, advertisers, and sellers trade information about consumer types. The component it compares distribution-platform and advertising-platform regimes, establishes profit and welfare bounds, and derives algorithms for privacy-preserving data sharing. Extensions examine competition among multiple providers, dynamic ticket pricing for compute resources, and heterogeneous buyer populations. Results are derived analytically and, where closed forms are infeasible, with numerical examples based. Outputs include scholarly articles, policy briefs, and open-source code for tariff computation and welfare analysis, enabling researchers and practitioners to apply the findings across sectors that rely on artificial intelligence, scalable computing, and data-driven decision making. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY/ABSTRACT This proposal comprises a five-year research and career development program for Dr. Diana Athonvarangkul to achieve independence as an investigator in disorders of bone and mineral metabolism. Dr. Athonvarangkul is a physician-scientist who completed her training in an NIH-sponsored Medical Scientist Training Program. The proposed research and career development activities will occur at Yale University. To facilitate her goal of leading her own independent research group, she has developed a training program based on high-yield didactics and professional development opportunities with support from a dedicated mentoring committee comprising scientists with a broad range of expertise relevant to the proposal and an impressive record of mentorship. She has laid out a clear timeline for these activities with timing of subsequent publications and acquisition of funding. The combination of research and career development plans described in this proposal will allow the candidate to mature into a successful and independent physician-scientist. Osteoporosis affects 10 million Americans and leads to 2 million broken bones each year. While a great deal is known about the molecular pathways by which osteoclasts remove bone, relatively little is known about how the most abundant cell type in bone, the osteocyte, remodels bone. Osteocytes remove bone from their surrounding matrix in a process known as osteocytic osteolysis. Our laboratory established that during lactation, osteocytic osteolysis is the physiological response to increased maternal calcium demand. The focus of this proposal is to elucidate mechanisms regulating osteocytic osteolysis using a lactation model, with an emphasis on how the osteoclasts and osteocytes coordinate their resorbing activities. Aim 1 will interrogate whether osteoclastic bone resorption influences a phenotypic switch in which osteocytes upregulate osteoclast-like gene programs and enlarge their lacunae and canalicular network. Aim 2 will follow up on previous observations to determine whether the lactational hormone, parathyroid hormone-related peptide (PTHrP), stimulates production of Receptor Activator of NFB Ligand (RANKL) to activate osteocytic osteolysis. Aim 3 will evaluate whether transforming growth factor beta (TGF) is a coupling factor released by osteoclast activity that regulates osteocytic osteolysis by interacting with PTHrP/PTH1R signaling. Aims 1 and 2 of this proposal will allow Dr. Athonvarangkul to gain experience and proficiency in the approaches needed to study bone biology. Aim 3 will facilitate the discovery of new molecule(s) and communication pathways among bone cells. As an independent investigator, Dr. Athonvarangkul will use the skills acquired in this proposal to determine how communication between osteoclasts and osteocytes is disrupted in diseases with excess bone loss. The experiments presented in this proposal will provide a new mechanistic understanding of intercellular communication within the bone environment during reproduction and may identify new therapeutic targets for the treatment of osteoporosis.

NSF Awards · FY 2025 · 2025-08

Representation theory is concerned with the study of linear symmetries. These symmetries form algebras that often arise as quantizations or more general deformations of functions on classical spaces -- this is a mathematical counterpart of passing from Classical to Quantum Mechanics or Field theory in Physics. Individual representations form categories, and understanding these categories leads to understanding the representations themselves. Tools to study categories include understanding their own symmetries and showing that categories of different origin are, in fact, equivalent. The focus of this project is to study categories of representations of quantizations, the symmetries of these categories, and their equivalences. The project also involves training young mathematicians and writing books and survey articles to benefit undergraduate and graduate students. In more detail, the project consists of three parts. The first part proposes the study of categories of highest weight modules over affine quantum algebras. These categories should be viewed as categorical analogs of the polynomial representations of double affine Hecke algebras. They depend on parameters, and the task is to establish derived equivalences between the categories corresponding to different parameters and to relate their t-structures. The second part of the project seeks to relate several categorical versions of the elliptic Hall algebra, a close relative of the double affine Hecke algebra, proving derived equivalences between these versions. The third part seeks a conceptual geometric understanding of unitary representations of complex semisimple Lie groups. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2025-08

Project Summary/Abstract Myocarditis, which is inflammation of the heart muscle mediated by immune cell infiltration and tissue damage, can lead to cardiac dysfunction or sudden death. The most common cause of myocarditis is viral infection, followed by other triggers including mRNA vaccination. Myocarditis, including after viral infection and vaccination, is more frequently seen in males, yet underlying mechanisms remain incompletely understood. A clearer understanding of male-biased myocarditis is essential for improving disease management and optimizing vaccine development. Previous work by us and others demonstrated that relative to healthy vaccinated controls, mRNA vaccine-associated myocarditis patients have elevations in circulating cytokines like IL-15, which activates T cells and innate-like bystander responses, coupled with increased peripheral activated cytotoxic T cells upregulating chemokine receptors, including CXCR3 and CCR5, implicated in heart tissue infiltration and site-specific bystander activation. In T cell bystander activation, memory T cells undergo nonspecific activation mainly via cytokines like IL-15 without TCR signaling or antigen recognition. Histopathological examination of patient heart biopsies confirmed predominant T cell infiltration. These immune responses have similarly been implicated in viral myocarditis, suggesting a potentially key role in male-biased heart inflammation. In preliminary studies, “dirty” mice with diverse prior microbial exposures and memory immune populations are shown to better model T cell bystander activation. Further, IL-15 administration in vivo, mimicking systemic triggers of pathology, induces male-biased bystander T cell responses and effects consistent with findings seen in myocarditis, which are reduced after androgen receptor blockade. Consistently, heart cytotoxic T cells were found to have elevated expression of the androgen receptor relative to various other tissues and immune subsets, with enhancing effects of androgens on heart bystander T cell activation signatures, which could explain the male bias and heart specificity. Based on this accumulating evidence, my hypothesis is that IL-15-induced bystander cytotoxic T cell responses mediate male-biased heart inflammation in an androgen-enhanced manner. To test this hypothesis, Aim 1 will define the role of IL-15-induced bystander T cells in male-biased heart inflammation using the developed dirty mouse in vivo system. Aim 2 will use androgen deprivation and supplementation to test effects on male-biased heart inflammation in vivo, including by multiplexed immunofluorescence spatial imaging. This study will advance our understanding of male-biased myocarditis and illuminate novel translational insights into sex immune differences and T cell bystander responses, both of which are widespread across numerous immune-mediated diseases. During this work, I will continue honing a unique skill set combining experimental techniques with powerful computational and bioinformatic approaches in an exceptional training and research environment toward a career advancing clinically relevant immunology as a physician-scientist.

NIH Research Projects · FY 2025 · 2025-08

Project Summary A central question in biology is how genetically identical cells generate diverse phenotypes. Cells often respond differently to the same perturbation, emphasizing the key role of internal cellular states in shaping these outcomes. Enhancing our ability to predict cellular behaviors could improve tissue regeneration, accelerate drug discovery, and better anticipate drug resistance. To address this, my lab integrates computational biology, systems biology, and cutting-edge genomic technologies to investigate how cellular diversity arises from a single genome, with applications across ovarian aging, cardiovascular disease, and hormone-dependent cancers. This proposal aims to develop a new framework for studying the causal mechanisms driving continuous phenotypic transitions in cellular states. By using PROTACs (proteolysis-targeting chimeras) as fast-acting chemical perturbations, we will induce rapid protein degradation, and apply single-cell proteogenomics to quantify how upstream biochemical changes lead to downstream gene regulatory changes. Compared to CRISPR-based genetic screens, PROTACs provide a temporally precise observation of gene expression changes before compensatory mechanisms emerge, providing clearer insights into causal relationships. This approach will enable us to build next-generation predictive models for cellular responses, advancing both fundamental biology and therapeutic development. Additionally, we are developing P3-seq, a novel single-cell technology that simultaneously measures intracellular proteins, protein-protein interactions (PPIs), and transcriptomes. PPIs are central to pathway integration and cellular signaling networks, yet no scalable single-cell method for quantifying them exists. P3- seq will provide deeper insights into how combinatorial protein interactions influence cellular outcomes—an area that remains underexplored. Over the next five years, our goal is to apply these discoveries to build predictive models that improve the ability to precisely control cellular behaviors, particularly in response to therapeutic interventions.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY Antibiotic usage disrupts intestinal homeostasis and is associated with an increased risk of infections in humans. Recent evidence suggests that non-antibiotic prescription medications can change microbiome composition, but how drug-microbiome interactions impact the risk of gastrointestinal (GI) disease is unexplored. To address this gap in knowledge, we analyzed health records, including hospitalization, drug prescription data, and pharmacy claims, for over 1 million anonymized individuals spanning 15 years to identify medications associated with increased risk of GI infection. We identified expected associations, including anti-infectives and immunosuppressive agents, that increase infection risk. Notably, we also identified unexpected associations: other prescription medications that elevate infection risk to a similar degree as antibiotics and immunosuppressants but do not fall into either of these drug classes. The central hypothesis of this proposal is that treatment with non-antibiotic prescription medications disrupts the microbiome to increase the risk of intestinal infections. We established a mouse model of Salmonella enterica subsp. Typhimurium (S. Tm) infection for two of these drugs (the cardiac drug digoxin and the anticonvulsant clonazepam), and demonstrated that drug administration increases infection risk and also alters microbiome composition. In our initial (proof-of- concept) studies, we established that the impact of digoxin on infection risk is transmissible via the microbiome. Specifically, digoxin decreases the abundance of a key group of immunomodulatory bacteria (segmented filamentous bacteria; SFB) in the mouse gut, and restoring the abundance of these bacteria rescues mice from increased infection susceptibility. Notably, clonazepam also increases infection risk, but in an SFB-independent manner. Building on this work, in Aim 1 of this proposal, I will 1) determine how digoxin alters SFB levels in the mouse gut and 2) examine whether human microbiome variability alters digoxin effects on the microbiome and S. Tm infection. I will use a similar approach in Aim 2 to examine whether clonazepam increases pathogen colonization by disrupting the microbiome. Through these aims, I will enrich my experience in mucosal immunology, analysis of large datasets, epidemiological study designs, and mouse models of infection (K99 phase). To enable this training, I have established a mentoring and advisory team consisting of experts in drug- microbiome studies, S. Tm pathogenesis, mucosal immunology, and epidemiological study designs. As I transition into independence, I will build on this unique foundation to define mechanisms of the impacts of prescription medications on the gut microbiome and infection risk (R00 phase). These studies will allow us to understand the effects of these bioactive compounds on the gut microbial communities, which are important in maintaining gut health. The underlying molecular mechanisms discovered by the completion of this study will enable us to develop strategies for mitigating GI diseases using microbiome-mediated interventions.