Case Western Reserve University

NIH Research Projects · FY 2026 · 2025-11

PROJECT SUMMARY/ABSTRACT There are currently 50 million people suffering globally with Alzheimer’s disease (AD). 95% of the population over age 65 is concerned about their dementia risk and 80% are interested in dementia screening. There is a critical need for accessible and cost-effective biomarkers that can be used to identify those on the ADRD continuum – including the asymptomatic stages – not only in research and specialty-care centers, but in community-based and primary care settings as well. This information could dramatically improve referrals for early clinical trial enrollment, the triage process for specialty evaluation, and comprehensive care planning. The methods used must be appropriate for point-of-care, community, or at-home deployment while maintaining accuracy and predictive value. Olfactory (sense of smell) dysfunction (OD), in combination with machine learning (ML) algorithms, is a promising non-invasive biomarker for ADRD. We have previously demonstrated the reliability of the Affordable Rapid Olfactory Measurement Array (AROMA) to objectively measure OD and categorize olfactory phenotypes (patterns of correct and incorrect responses to various odorants and multiple concentrations). AROMA uses essential oils, which are complex blends of odor molecules and may be more reflective of “real world” olfaction than the single chemicals used in most other tests. This is because when scents are encountered in real life, the brain processes and recognizes the odorant combinations making up each complete scent differently from the individual component chemicals. Our research with AROMA in ADRD has shown that AROMA can distinguish cognitively unimpaired (CU), mildly cognitively impaired (MCI), and AD patients from one another. Additionally, olfactory phenotypes were detected using machine learning and differentiated between disease states. Our algorithms had 100% sensitivity, 83% specificity for correctly classifying CU versus MCI/AD. Algorithms tasked with classifying MCI versus AD had 100% sensitivity, 75% specificity. We propose longitudinal testing of CU, MCI, AD subjects (n=324 men and women > 55 years) over 3 years to assess changes in OD, functional status, and neurocognition. A group of neurologic controls will be included to ensure olfactory phenotypes are specific for ADRD. Using traditional statistics and machine learning techniques to examine the relationship of AROMA performance with ATN-biomarkers and clinical markers of disease (Aim 1); define predictive models using AROMA data to predict changes in function and frailty (Aim 2); and develop a streamlined ADRD-version of AROMA using only the scents and concentrations of highest influence (Aim 3). Our long-term goal is for point-of-care olfactory biomarker data, analyzed in real-time by ML algorithms, to be widely accessible to meaningfully inform clinical, research, and caregiver decisions.

NIH Research Projects · FY 2025 · 2025-09

Abstract Atherosclerotic cardiovascular disease continues to be a significant health concern worldwide. Improving risk stratification is particularly crucial to identify individuals at risk for near term cardiovascular incidents and detect disease earlier. Using cardiac CT scans for coronary artery calcium (CAC) scoring is a well-established risk assessment method, but the traditional Agatston calcium scoring method lacks sensitivity for near-term and early-stage disease detection. On cardiac CT images, small, low-density, “spotty” calcifications have been linked to high-risk plaque characteristics, like inflammation and stress, and signal an early stage of atherosclerosis before progressing into larger calcifications. Yet, these small calcifications are usually overlooked in standard Agatston scoring due to their low density and/or small size. The objective of the proposed research is to apply advanced computational methods like deep learning to optimize the detection of these “sub-Agatston” calcifications (Sub-Aim 1a) and derive reliable image features that characterize these sub-Agatston CACs and are predictive of cardiovascular risk (Sub-Aim 1b). To gain insights into the mechanisms of atherosclerosis initiation and progression, sub-Agatston calcification features will be causally associated with potential drivers of atherosclerosis, including socioeconomic, clinical, and image-derived-metabolic risk factors (pericoronary and epicardial adipose tissue) (Sub-Aim 2a). Lastly, the predictive value of sub-Agatston calcifications for major adverse cardiac events will be assessed in over 100,000 patients through Cox proportional hazards modeling (Sub-Aim 2b). This proposed research will yield: (1) Improved risk prediction models accounting for new (sub- Agatston) and understudied features (socioeconomic status). (2) New insights into atherosclerosis onset and progression (3) Technical innovations in machine learning and causal inference for medical imaging. (4) Translational software tools suitable for clinical risk assessment. This proposal, levering big CT data analytics, will provide crucial knowledge on the prognostic value of small, low-density calcifications detected on CT images, advance preventative cardiovascular medicine through the early detection of heart disease, and enable a new understanding of atherosclerosis initiation and progression.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY Alzheimer’s disease (AD) is a progressive neurodegenerative disease with memory loss and impaired mental function. A defining feature of AD biology is the accumulation of amyloid plaques in the brain. While plaques are a major target for late-onset AD (LOAD) therapeutics, the resulting treatments do not cure LOAD and are only moderately effective in slowing progression. The challenges of developing effective LOAD therapies after the onset of disease underscore the need to identify modifiable risk factors that can be leveraged earlier in life to avoid onset or even after onset to slow progression and promote higher quality of life for both patients and their care givers. Several non-modifiable risk factors include increased age, female sex, genetics, and family history of AD. One modifiable factor associated with protection from LOAD is physical activity where exercise is associated with lower risk of developing LOAD. Several studies suggest that physical activity modifies genetic risk for LOAD and that this relationship can be statistically described in gene x environment (GxE) association studies. Most LOAD GxE studies are candidate variant or gene focused on APOE, the strongest independent LOAD genetic risk factor. With now ~75 genome-wide associated genetic variants/genomic regions, the majority of LOAD genetic risk has yet to be tested for modification by physical activity or other possible modifiable factors. Critical gaps in the field of AD research are to understand the molecular basis of how exercise exerts a neuroprotective effect, and how the genetic factors that influence AD risk are modified by physical activity. The molecular impact of physical activity, while difficult to study in humans, is now being systematically mapped in rats as part of the Molecular Transducers of Physical Activity Consortium (MoTrPAC). MoTrPAC offers a public molecular map of the effects of induced exercise based on ‘omic data generated from 19 tissues harvested from 6-month old male and female rats that underwent up to 8 weeks of exercise training. We propose to leverage public cross-species transcriptomic MoTrPAC data to prioritize LOAD-associated genomic variants/regions for human GxE studies for LOAD and physical exercise in the diverse All of Us Research Program. We expect that upon the completion of this one-year project, we will have undertaken the most thorough examination of physical activity as a modifier of APOE risk in diverse populations as well as developed a MoTrPAC-informed pipeline to continue the work of identifying other genomic regions modified by exercise relevant to LOAD risk in other populations.

NSF Awards · FY 2025 · 2025-09

Fatigue refers to the mechanical failure of materials subjected to repeated cyclic loads. It occurs in all materials including metals and alloys, and is a significant limitation that affects all engineering structures and devices (both structural and functional). It is a common cause of failure (and accidents) in devices including computers, cars, bridges and airplanes, and thus a significant economic, societal and national defense challenge. Unfortunately many fundamental aspects of fatigue remain incompletely understood, even though decades of empirical knowledge have provided (conservative) material specific guidelines for the design of engineering components to avoid fatigue in specific materials. The advent of additive manufacturing and advanced alloys have pushed us beyond this empirical knowledge base. In particular, recent experimental observations suggest situations where additively manufactured metallic alloys have fatigue strength that greatly exceed their conventionally manufactured counterparts, and other situations where they greatly underperform. The investigators will develop a new data-driven approach predicated on the view that the spread of fatigue life can be attributed to particular details of defects and microstructure, and a fundamental understanding of this relationship can lead to new Structural Alloys for Fatigue Endurance (SAFE). The approach is to create an integrated database and knowledge map of material and processing parameters, microstructure, comprehensive mechanical characterization, post-failure analysis and computational experiments on the fatigue behavior of additively manufactured structural alloys. Innovations in methodology including efficient high throughput testing and serial sectioning combined with Electron Backscatter Diffraction (EBSD) to acquire three-dimensional images of microstructure with orientation around individual crack initiation sites, in operando synchrotron observation and accelerated approaches to simulation enable the creation of this database. New methodologies are developed to sample the knowledge map and add new experimental results and simulation until there is a significant area of knowledge where the control parameters predict the quantities of interest. This leads to a deep fundamental understanding of the exact mechanisms and features that initiate, inhibit and accelerate fatigue cracks, and subsequently to the inverse problem of designing new SAFE materials. The approach is developed with a focus on the widely used aerospace alloy Ti-6Al-4V. The team consists of experts in additive manufacturing, texture & characterization, fatigue, computational modeling and the application of machine learning to materials. 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 · 2025-09

In the U.S., 7.5 million persons ages 60 and older experience elder abuse (EA) annually. Family caregivers (CGs) are among the most likely to commit EA: one-third to one-half of CGs self-report engaging in EA towards their family member living with Alzheimer’s disease or related dementia (AD/ADRD), an at-risk population. Psychological EA is the most common type of abuse among CGs of those with AD/ADRD, making up 25%-42% of cases. The mental health harms of psychological EA are comparable physical EA: older adults who experience psychological EA are at an increased risk of anxiety, depression, and chronic disease burden. Interventions targeting CGs have gained traction as a promising way to prevent EA. Yet, effective approaches to EA prevention with CGs remain emergent. Interventions targeting risk factors alone (e.g., poor CG mental health) have been unsuccessful, and little is known about effective mechanisms of change to prevent psychological EA. Other fields of family violence have shown success by focusing on relationship dynamics to prevent abuse, yet this is a novel approach to EA. A relationship-focused approach to prevention is supported by observational studies showing relationship strain as a risk factor for EA and low-quality caregiving. Beyond mitigation of relationship strain alone, development of resourcefulness skills is proposed to prevent psychological EA by equipping CGs with skills to manage challenging caregiving scenarios that may precede psychological EA. The Knowledge and Interpersonal Skills to Develop Enhanced Relationships (KINDER) intervention was developed to prevent psychological EA and promote high quality caregiving by lowering relationship strain and developing CG resourcefulness. In a recent pre- and post-test pilot study (N=45), CGs who participated in KINDER reported reduced frequency of psychological EA (p<0.01) and higher quality of caregiving (p<0.03). Investigators also observed reduced relationship strain (p<0.01) and increased resourcefulness (p<0.01). The goal of this two-arm, randomized control trial is to determine the efficacy of KINDER at mitigating psychological EA by family CGs to persons living with AD/ADRD and to describe how the intervention may work to reduce psychological EA. The investigators hypothesize that family CGs exposed to the KINDER intervention will have improvements in psychological EA and quality of caregiving when compared to CGs exposed to an information-only control condition. Further, increased resourcefulness and reduced relationship strain will explain reductions in psychological EA and improved quality of caregiving. The investigators will also assess differences in effects according to time in the caregiving role, kin relationship, as well as CG demographic characteristics. This research responds to the too-common occurrence of psychological EA by family CGs, and may help prevent the destructive effects of psychological EA in a vulnerable and growing population of persons living with AD/ADRD.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY/ABSTRACT Adverse early-life experiences can have profoundly negative consequences for a person’s behavioral, social, and physical health throughout their lifespan, a problem with great relevance for public health and for the applicant’s career goal of training as a pediatrician-scientist. Nearly three decades after this association was uncovered, the mechanisms by which adverse experiences are biologically embedded in the nervous system during developmental sensitive periods remain largely unknown, due in part to a lack of tractable model systems. This proposal describes a novel model of experience embedding in the Drosophila melanogaster olfactory system, in which the experience of chronic early-life exposure to the common food cue ethyl butyrate (EB) is stored as instructions for glia to activate an innate immune engulfment response and aggressively phagocytose a critical EB-sensing circuit upon (1) stimulation with high levels of EB later in life or (2) injury to EB sensory neurons. The central hypothesis of this proposal is that early-life experiences are embedded in glia as an innate immune memory that primes them to vigorously respond to subsequent experiences or other stimuli, a potentially long-lasting maladaptive state. Previous studies have demonstrated that astrocytes and microglia can store molecular memories of certain stimuli in the form of epigenetic modifications or sustained activation of innate immune pathways. A preliminary screen identified a little-studied yet highly conserved acetyltransferase, histone acetyltransferase 1 (Hat1), which is dispensible for acute pruning but is required in glia for the embedding of early-life EB exposure. This proposal seeks to (1) define the duration, timing, and intensity of the experience required to induce glial priming, and whether a primed response can be evoked by heterologous stimuli, (2) characterize the functional state of primed glia, including their morphology, activity, and activation of innate immune pathways, and (3) interrogate the role of Hat1 in glial priming by engineering the hat1 locus and knocking down expression of Hat1 complex members. These aims take full advantage of the applicant’s training environment, including a research group with deep expertise in Drosophila innate immunity and glial biology, and are well-integrated with the applicant’s training goals and preparation for a career as a physician and independent investigator.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY Despite the impressive therapeutic success of chimeric antigen receptor (CAR)-T cell therapy in treating previously incurable hematological malignancies, its use is associated with significant safety concerns. The most common and potentially life-threatening adverse events (AEs) after CAR-T cell infusion are cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS). The incidence of high- grade events makes up a quarter to a half of cases of patients who experience these toxicities. In addition to risk to patient safety, management of these AEs increases health care resource utilization and financial burden. Both events are associated with a heightened immune effector state following CAR-T cell infusion, yet their underlying pathophysiology remains elusive. After infusion, the activated CAR-T cell repertoire stimulates recruited macrophages to propagate a dysregulated pro-inflammatory state which can cause vascular leakage and CRS. Progression into the central nervous system causes ICANS. IL-6 is a central mediator in this mechanism and its neutralization with tocilizumab is standard of care for treating CRS. However, CRS may recur in some tocilizumab-treated patients and, significantly, tocilizumab does not prevent progression to ICANS, which is managed with glucocorticoids. Novel discoveries into the mechanisms of CRS and ICANS may uncover treatment strategies that could more effectively manage both AEs. To this end, we are interested in studying a novel role for B-cell activating factor (BAFF) in CRS/ICANS pathophysiology. BAFF is a TNF family cytokine that drives B-cell maturation and survival and classically studied for its pro-tumorigenic effects in B-cell malignancies. We recently found that serum BAFF levels are significantly elevated with increased IL-6 relative to pre-treatment levels in patients who experienced CRS following CAR-T cell infusion. When exploring a potential role of BAFF in CRS/ICANS pathophysiology, we found that interferon gamma (IFNγ) stimulates monocytes to secrete BAFF and upregulates two receptors of BAFF. BAFF stimulation of peripheral blood mononuclear cells and, more specifically, monocytes, results in increased expression of multiple pro-inflammatory cytokines involved in CRS/ICANS pathophysiology. Conversely, neutralization of soluble BAFF with belimumab resulted in decreased expression and production of these cytokines. Based on this preliminary data, we hypothesize that IFNγ released from activated CAR-T cells stimulates recruited monocytes to increase BAFF production and expression of BAFF receptors. BAFF ligation may be an early event that stimulates production of pro-inflammatory cytokines to propagate the CRS and ICANS sequelae. Clinically, targeting BAFF may represent a promising therapeutic approach to blunt the dysregulated proinflammatory cascade at an early stage and limit the development of both AEs. Based on the pro-tumorigenic role of BAFF, this could concurrently help to limit disease burden. In summary, discoveries made in this proposal will add mechanistic insight to the most common toxicities that accompany CAR-T cell therapy and potentially lead to novel therapeutic approaches to make this life-saving therapy safer.

NSF Awards · FY 2025 · 2025-09

With the support of the Macromolecular, Supramolecular, and Nanochemistry Program in the Division of Chemistry, Dr. Genevieve Sauve of Case Western Reserve University will explore new chemical strategies to improve the stretchability and strength of semiconducting conjugated polymers (CP), necessary for making the next generation of flexible electronic devices, such as foldable displays and wearable technology. To be effective semiconductors, CP must be able to transport charges effectively. However, optimizing stretchability, strength, and charge transport in these polymers is a challenge. This project will address this by introducing special cross-links between the polymer chain ends. These cross-links will be based on dynamic metal-ligand (ML) bonds, which are known to improve the toughness of non-conjugated polymers. This research will transform the field of organic electronics by providing an easy way to obtain tough CP without compromising charge transport. The enhanced toughness will lead to mechanically stable CP films, essential for preventing cracking of flexible devices. This will impact electronic devices that provide energy, information, or light. The improved toughness will also enable new functionalities such as wearability, compact packaging, and portability. This research will train graduate, undergraduate, and high-school students to be the next generation of scientists. Participation in the American Chemical Society local section will further increase impact. Semiconducting CP have the potential to revolutionize the electronic industry by enabling lightweight and flexible electronic devices, but to reach their true potential, their mechanical properties must be tuned for a given application. Many strategies have been reported to obtain stretchable CP for biomedical applications, but the development of CP that are both stretchable and strong remains a challenge. This project’s main hypothesis is that tough CP with good charge transport properties can be obtained using dynamic ML cross-links between the ends of CP. This hypothesis will be tested by synthesizing ligand-terminated CP and investigating the effect of the ML cross-links on structural, mechanical, and electrical properties. CP tensile strength will be tuned with the ML binding strength and loading. Stretchability will be enhanced using a flexible spacer between the CP and ligand. To simultaneously optimize mechanical and electrical properties, a novel scalable method to introduce cross-links in films will be investigated. The fundamental structure-property relationship will be studied using well-defined regioregular poly(3-hexylthiophene) as a model polymer. The strategy will be expanded to high-performance donor-acceptor CP. This approach will also be applied to fabricate mechanically stable organic photovoltaic devices. This project will explore for the first time how using dynamic cross-links at the CP chain ends can tune film properties, thus advancing fundamnetal knowledge of CP end-group chemistry and its effects on properties. It has the potential to transform the field of organic electronics by providing an easy way to increase toughness of CP while retaining desirable electronic properties. 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 · 2025-09

Project Summary The proposed Diabetes Endothelial Keratoplasty Study (DEKS) Renewal will follow up from the initial findings of the DEKS, driving evidence-based decisions regarding use of corneal tissue from donors with diabetes for Descemet membrane endothelial keratoplasty (DMEK), the leading form of endothelial keratoplasty in the US. There remains the need to definitively resolve the discrepant findings regarding the use of diabetic donor tissue to guide the corneal transplant community. While we hypothesize that overall non-diabetic corneas are superior, it is unlikely that all donors with diabetes are inferior. Rather, based on preclinical and DMEK eye bank stripping data, we presume that the extremes of diabetes severity in donors drive the adverse effects on DSAEK outcomes that our group noted in the Cornea Preservation Time Study (CPTS) where diabetes in donors and recipients was not collected systematically. In the current DEKS, we seek to determine graft success varies between donor subgroups predefined by presence or absence of diabetes as well as various diabetes severity scales after 1 year of follow-up which we expect will be welcomed findings by eye banks and corneal surgeons guiding them to avoid inefficient distribution of inferior tissue for DMEK. However, the current DEKS, with follow-up limited to 1 year, does not address long term endothelial cell loss (ECL) and accompanying graft failure of diabetic vs non-diabetic donors as well as the recipient diabetes effect. Nor does it examine long term effects of diabetes severity and other predictors (genetic) on ECL and graft failure. The renewal of the DEKS would address this by a powered analysis of long-term ECL out to 5 years as a marker for graft survival in the first specific aim while exploring impact on graft failures in the second specific aim. The third specific aim in the renewal of DEKS will be continuation of the third specific aim in DEKS, exploring the relationship of severity of diabetes in the donor (as measured by eye bank-determined diabetes risk categorization scores, post-mortem HbA1c, and skin AGEs and oxidation markers), and in the recipient (as measured by diabetes risk categorization scores, and HbA1c). In addition, in the most novel aspect of this renewal, we will explore donor and recipient genetics (mutations in TCF4, other Fuchs dystrophy genes, and diabetes polygenic risk scores) on ECL and graft failure 5 years following DMEK. These long-term insights on the diabetic donor and associated graft outcomes as well as recipient diabetic status and Fuchs genetics will provide further guidance to the eye banking and corneal transplant surgeon community on the use of an increasing number of diabetics in the corneal donor pool and increasing number of diabetics in recipients also affected by Fuchs Dystrophy.

NIH Research Projects · FY 2025 · 2025-09

Existing literature indicates significant differences in outcomes among patients with HPV+ OPSCC, with some groups experiencing poorer survival rates. Our analysis of two independent OPSCC cohorts from Cleveland supports these findings; patients identified as p16+ in one group had a 64% and 69% lower risk of death compared to another p16+ patient group at the University Hospitals Seidman Cancer Center and MetroHealth System, respectively. Interestingly, time to treatment initiation, defined as the number of days between pathologic diagnosis and the start of treatment, was statistically similar between these groups in both cohorts. This suggests that differences in access to medical care are not the primary contributor to the observed survival difference. While clinical outcomes in OPSCC are multifaceted and influenced by various interconnected factors, our findings highlight the potential role of biological differences in contributing to the survival gap between these patient populations. Our Cleveland-based OPSCC study cohort consists of 562 patients, including 74 individuals from this high-risk subgroup, representing the largest such cohort studied to date with HPV status based on NGS. Our proposed work, which leverages our robust OPSCC cohort, aims to provide valuable insights into the biological factors, such as HPV status, HPV genotype, and tumor-immune microenvironment landscape, which may contribute to the observed differences in clinical outcomes.

NSF Awards · FY 2025 · 2025-09

Cells have evolved sophisticated strategies to ensure the accurate expression of genetic information and safeguard against potentially harmful effects of defective messenger RNA (mRNA) and proteins. One such strategy recognizes mRNA translation events that terminate prematurely and targets the faulty mRNA transcript for rapid degradation. This critical quality control activity prevents accumulation of unproductive mRNAs and averts expression of incomplete, potentially toxic proteins. Despite it being a conserved and vital mechanism of gene regulation in most cells, how a cell distinguishes premature translation termination from normal termination and how that leads to accelerated degradation of the faulty mRNA remains poorly understood. This project seeks to identify and investigate key molecular interactions between the translation machinery and mRNA surveillance components necessary to target mRNA for degradation. In addition to advancing understanding of a fundamentally important biological process, this work will provide research experiences for students at the high school, undergraduate and graduate levels. Findings will be disseminated broadly among scientific communities and shared with the public to promote engagement in science. This research will interrogate the molecular interactions and events that occur between the translation and mRNA surveillance machineries to mediate recognition and rapid degradation of nonsense-containing mRNA via nonsense-mediated mRNA decay (NMD). Using the budding yeast model system, innovative genetic and biochemical approaches will be employed to define how UPF1—the central effector of the NMD machinery—impinges on prematurely terminating ribosomes to trigger accelerated decay of the mRNA, and how NMD cofactors, UPF2 and UPF3, contribute to this process. The study aims to uncover previously uncharacterized molecular events that occur downstream of NMD substrate recognition and to advance understanding of this critical RNA quality control mechanism. 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

This project is in asymptotic geometric analysis and affine convex geometry. One main emphasis of the research is on high dimensional objects and phenomena. This leads to applications in areas as physics, biology and medicine, computer science, optimization and economics and material science. Indeed, many objects appearing in these areas exhibit the property of being convex, including crystals, organs, high dimensional data clouds. How can one reconstruct or gain substantial knowledge of such convex objects when one only has partial information? This is one classical problem in convexity, called geometric tomography, and its research finds applications in medicine and biology where convex shapes occur naturally. In addition to the research activities, the PI will continue giving lectures to the scientific community, as well as to the general public and training graduate students and postdocs. Important features of the proposed project are the study of high dimensional objects and phenomena and their links with other areas of mathematics and mathematical sciences, such as probability, statistics and information theory. Of particular interest are the affine invariant functionals on convex bodies in high dimensions. Among the most important such functionals are affine surface area and p-affine surface area. Their corresponding affine isoperimetric inequalities, established by the PI and collaborators for all p, are stronger than their Euclidean counterparts and related to the famous Mahler conjecture which is still open in dimensions four and higher. It was shown by the PI that p-affine surface areas are directly related to entropies of cone measures of convex bodies which establishes a link between convex geometry and information theory. This link will be further explored, also in the context of log concave functions which are a natural extension of convex bodies in the realm of functions. Moreover, affine surface area appears naturally in questions on approximation of convex bodies by polytopes, a further main topic of study. The goal is to establish optimal dependence on all the relevant parameters involved in the approximation, like e.g., dimension, the number of vertices of the approximating polytopes. These issues will be considered in Euclidean, spherical and hyperbolic space. The PI and her collaborators extended the notions of affine surface area recently to a functional setting and to spherical and hyperbolic space. To establish the corresponding inequalities in those spaces is a further topic of study. 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 Multiple sclerosis (MS) is the most common demyelinating disease of the central nervous system and stems from autoimmune damage against the myelin sheath, which is necessary for rapid and efficient conduction of electrical impulses along neurons. Oligodendrocyte progenitor cells (OPCs) serve a vital role in regulating the progression of MS because they differentiate into myelinating oligodendrocytes to restore myelin integrity in demyelinated lesions. However, many demyelinated lesions in MS experience remyelination failure, a poorly understood phenomenon that may be triggered by neuroinflammation. Recent evidence has showed that OPCs have notably decreased differentiation to oligodendrocytes and interestingly adopt novel antigen presentation functions in settings that mimic neuroinflammation. Even so, how the potential connection between these two separate phenotypes is biologically meaningful to remyelination failure and what molecular mechanisms drive their occurrence in this disease state remain unclear. To answer these questions, I developed a robust, scalable in vitro culture system to generate pure OPCs from mouse induced pluripotent stem cells that are then treated with interferon-gamma (IFNγ), a proinflammatory cytokine that is greatly increased in MS lesions and can broadly simulate the neuroinflammatory milieu in this disease. Through bulk RNA-seq and high-throughput chemical screen studies, I have found that STAT1 activity controls both IFNγ-induced phenotypes. In this proposal, Aim 1 will assess how IFNγ alters the temporal dynamics of differentiation and immune function activation to gain greater insights into MS pathophysiology. Because I can precisely control IFNγ exposure, I seek to probe potential transcriptional and functional memory in IFNγ-treated OPCs that can be broadly relevant to relapsing-remitting MS, the most common MS subtype. I will also use special nanofiber plates for 3D culturing of oligodendrocytes and interrogation of memory in myelination behavior (i.e. fiber wrapping) by modulating IFNγ exposure over a longer duration. Next, Aim 2 will assess functional targets of STAT1 to determine mechanisms for these two phenotypes. In Ctrl- versus IFNγ-treated OPCs, I will perform CUT&RUN for H3K27ac to assess genome-wide epigenetic changes and for STAT1 to determine its global binding profile and how STAT1 can cause these changes in acetylation and also have context- dependent functions. A rigorous computational pipeline will identify key differentially regulated genes and pathways, and I will validate them in vitro via genetic knockdown studies and in human MS single- cell RNA-seq datasets and patient brain sections. The experiments outlined here will significantly bolster our knowledge of oligodendrocyte functional plasticity in remyelination failure and pave the way for new, exciting therapeutic approaches that go beyond conventional MS disease-modifying therapies aimed at immune cell depletion and instead harness the innate regenerative capacity of oligodendrocytes.

NIH Research Projects · FY 2025 · 2025-08

The K12 proposed by the Clinical and Translational Research Collaborative of Northern Ohio is an innovative, flexible program to prepare a highly skilled cohort in clinical and translational (C/T) research to address the lack of optimal management strategies for hundreds of diseases and conditions and for unique approaches across the entire population, including special and vulnerable populations of all ages. The K12 seeks support for an educational curriculum designed to meet the individual needs of emerging investigators across all disciplines, with an emphasis on addressing differences in health outcomes, expanding participation across disciplines, and improving alignment of candidate backgrounds with C/T research objectives. This program is designed to adapt to the individual needs of each scholar and to address the unmet educational and career development needs of clinical research scholars in the rapidly evolving field of C/T science, with a focus on examining variations in health outcomes across population groups. Unmet needs will be addressed by educating leaders in multidisciplinary clinical and translational research; introducing clinical research education earlier in the life cycle of scholars from varied academic disciplines (nursing, bioinformatics, social work, pharmacology, etc.); tailoring programs to the preferences, special interests, research plans, strengths, and weaknesses of each scholar with appropriate modifications; and setting the standards and developing innovative approaches to C/T career development. The program builds on lessons learned from prior experience in operating a highly successful KL2 program, proven strategies from published national studies on mentorship, collaboration with other CTSA Hub KL2 sites, new and innovative programs to re-enforce the strengths of scholars in a broad and deep spectrum of capabilities, and a robust education and career development program that utilizes all resources within the CTSC and synergizes with the post-doctoral T32 and the UM1's Workforce Development Module. The K12 Career Development program aims to 1) further enrich and expand our integrated CTSC- wide innovative and individually tailored K12 program, 2) prepare the next generation of investigators with the multidisciplinary skills required to lead cutting-edge C/T research and meet the opportunities and challenges of medicine in the 21st Century, and 3) build the multidisciplinary workforce of the future. This innovative K12 program focuses on addressing each scholar's individual gaps in knowledge and skill. The program will benefit from a cross-institutional, cross-disciplinary C/T research career development program, which, in turn, derived from the successful introduction of the Roadmap K12 multidisciplinary C/T research program launched in 2004. As the program evolved, best practices were kept from earlier iterations, scholar feedback was incorporated, available institutional resources carefully considered, and attention paid to desired outcomes (what was accomplished and who was reached). Results (short, intermediate and long term) and overall impact of the entire program will be measured and shared through publications and presentations, with specific measures to determine the impact on health outcomes.

NIH Research Projects · FY 2025 · 2025-08

Project Description Selective autophagy plays an important role in maintaining cellular homeostasis in various organelles. Since proteins in the secretory pathway and membrane proteins must fold into their native structures in the endoplasmic reticulum (ER) before their anterograde trafficking, selective autophagy of the ER, or ER-phagy (reticulophagy), has been recently recognized as an important protein quality control pathway of aggregation-prone proteins in the ER. A key development of ER-phagy is the identification of six specific transmembrane ER-phagy receptors, including CCPG1, the focus of this research. However, there is a significant gap about the role of many of these ER-phagy receptors in physiological and pathophysiological conditions, especially in the central nervous system (CNS). The CNS must maintain a delicate balance between inhibition and excitation. The inhibitory signal is mainly dictated by gamma-aminobutyric acid type A (GABAA) receptors. Proteostasis maintenance of GABAA receptors is critical since their proteostasis defects lead to neurological diseases, such as epilepsy and developmental delay, resulting from variations in genes encoding these neuroreceptors. We and other groups have demonstrated that one major disease-causing mechanism is due to the protein misfolding of these variants in the ER and thus their reduced trafficking to the cell surface for function. Preliminary data demonstrated that GABAA receptor variants that aggregate in the ER are targeted to the lysosome for degradation; furthermore, among the known six membrane ER-phagy receptors, only CCPG1 interacts with the variants to promote clearance. However, the role of CCPG1 in the CNS is unknown yet. Therefore, based on preliminary data, we hypothesized that CCPG1 directs aggregation-prone GABAA receptor variants in the ER to the ER-phagy pathway, and loss of function of CCPG1 leads to protein aggregation and proteostasis deficiency in neurons. We proposed to test the hypothesis with the following specific aims by combining approaches in cell biology and electrophysiology. Here, in Specific Aim 1, we will elucidate CCPG1-mediated ER-phagy pathway for pathogenic neuroreceptor variants. In Specific Aim 2, we will determine the effect of CCPG1 on proteostasis maintenance of neurons.

NIH Research Projects · FY 2025 · 2025-08

Abstract / Project Summary The brain's extracellular matrix (ECM) is a three-dimensional milieu that plays a crucial role in synaptic function and plasticity during development in addition to its regulatory role in axon regeneration and remyelination from injury. Despite its significance, our understanding of how the content of the major brain ECM components, glycosaminoglycans (GAGs) and GAG-modified proteins (proteoglycans), are regulated is limited. Mucopolysaccharidosis VII (MPS VII) is a multi-organelle lysosomal storage disorder caused by the deficiency of β-glucuronidase (GUSB), resulting in neurodevelopmental delay and broad neurological impairment. Our preliminary studies using an MPS VII mouse model that is globally deficient in β-glucuronidase enzyme show prominent intracellular WFA-stained aggregates observed selectively in CD68+ microglia. These results suggest a critical role of microglia-mediated proteoglycan/GAG catabolism in CNS development. These findings lead us to hypothesize that the loss of GUSB function in microglia is a key driver of MPS VII neuropathology. However, there are no existing genetic tools to specifically target and ablate β-glucuronidase activity in microglia. Additionally, there are no existing biomarkers to assay for non-degraded GAGs accumulating intracellularly from the loss of β-glucuronidase activity. The objective of this proposal is to develop new genetic and biochemical tools to test for the importance of microglia-mediated GAG catabolism in the CNS and its dysfunction as a root cause for MPS VII neuropathology. In Aim 1 we will develop and validate new genetic tools to conditionally ablate β- glucuronidase mediated GAG catabolism within microglia and other cells of neural lineage. In Aim 2 we will measure for non-degraded intracellular GAGs that selectively accumulate in MPS brain tissue. Combining these newly generated genetic reagents with the biochemical approach to measure non-degraded intracellular GAGs will allow us to discern and assess microglial mediated GAG catabolism and their dysfunction in MPS VII neuropathology.

NIH Research Projects · FY 2025 · 2025-08

Our goal is to develop and optimize novel neurostimulation strategies to restore intuitive and informative somatosensation to people with tetraplegia. Sensory feedback is critical for regulating dexterous grasp during activities of daily living and for emotional connection with loved ones. Neuroprostheses to restore voluntary control of reaching and grasping to users with paralysis will be more likely to achieve clinically-meaningful benefits in independence and quality of life if sensory feedback is integrated into these systems. The central hypothesis is that restoring sensation with intracortical microstimulation (ICMS) of primary somatosensory cortex and/or peripheral nerve stimulation (PNS) will improve performance of brain-controlled grasp tasks. The secondary hypothesis is that paired ICMS and PNS will promote neuroplasticity and enable users to recover some of the below-injury sensory function lost due to spinal cord injury. Three participants with chronic tetraplegia enrolled in an existing clinical trial who have already received chronically-implanted cortical arrays and nerve cuff electrodes will participate in the proposed study. The study has three specific aims: 1) Determine the impact of spatiotemporal ICMS paradigms on perception and function. Novel ICMS encoding schemes that prioritize different neural coding properties and information content of the transmitted touch stimuli will be developed and compared. The perceptual response to each ICMS paradigm will be evaluated with classical psychophysical techniques. Decoders that map cortical activity to desired hand/arm movement commands will be developed and used to perform grasp tasks in virtual reality. The functional utility of each ICMS paradigm will be evaluated and compared. 2) Determine the impact of spatiotemporal PNS paradigms on perception and function. Novel PNS encoding schemes will be developed and their impacts on perception and brain-controlled task performance will be compared. The effect of a PNS training paradigm on below-injury sensory function will also be evaluated. 3) Determine the impact of hybrid (ICMS+PNS) neurostimulation on perception and function. Novel hybrid stimulation paradigms will be developed through systematic investigations of percept integration resulting from manipulations of encoding scheme, electrode contacts, and inter-stimulus timing. Hybrid stimulation will be compared to ICMS-only and PNS-only paradigms in both perceptual characteristics and virtual task performance gains. The effect of a hybrid neurostimulation training paradigm on recovery of below-injury sensory function will also be evaluated. The proposed work will improve our understanding of the perception of neurostimulation applied throughout the sensory nervous system, the benefits of restored sensation on the functional utility of brain-controlled neuroprostheses, and the role of touch information content in these outcomes. This study will also determine the feasibility of a future therapeutic intervention using neurostimulation for below-injury sensory recovery. Study findings have implications for the future development and translation of neuroprostheses to restore sensorimotor function to people with tetraplegia.

NIH Research Projects · FY 2025 · 2025-08

Tobacco use remains the leading causes of preventable death and disease in the U.S. Little cigars and cigarillos (LCCs) are not used in the same way as other tobacco or nicotine products due to their unique attributes (e.g., breadth of flavor, low cost per unit), yet remain understudied. As a result, LCCs are often used along with other tobacco products as part of a multiple tobacco product (MTP) use pattern. MTP is a growing concern for young adult populations. Furthermore, co-use of cannabis is similarly prevalent among young adults who use LCCs, which may make it more difficult for young adults to stop using tobacco. To date, the measurement and conceptualization of MTP and cannabis co-use is not well defined particularly among those who use LCCs. The overall goal of this proposal is to build the candidate’s research expertise on MTP and cannabis co-use. This award will support the candidate’s career development as a sociobehavioral epidemiologist. The career development plan includes mentoring and training in 1) addiction science of tobacco and cannabis use, 2) mixed methods, 3) ecological momentary assessment, and 4) grant writing and academic leadership. Aim 1 of the proposed research study includes a longitudinal secondary data analysis of a representative national sample of young adults. This data will be used to 1) evaluate tobacco and cannabis consumption profiles, and 2) examine the role of LCCs in transitions to other tobacco and cannabis product use as well as in developing symptoms of tobacco use disorder. Aim 2 is a phased mixed methods study that will test the feasibility and acceptability of conducting an ecological momentary assessment (EMA) among individuals who use LCCs and other tobacco and cannabis products. Phase 1 will include cognitive interviews to pretest data collection tools to be used in the subsequent study phase. Phase 2 will be a pilot EMA including a baseline and follow-up surveys, momentary assessments, and daily experience diaries. Phase 3 will include in-depth qualitative interviews to understand the patterns of tobacco and cannabis use and their impact on the symptoms of tobacco use disorder among those who use LCCs. Through successful execution of the mentoring and research aims, this research will lay the foundation for a scaled study that captures dynamic patterns of substance use and their relationship to symptoms of tobacco use disorder to identify novel, tailored strategies that disrupt these dynamic patterns to promote successful smoking cessation among young adults.

NIH Research Projects · FY 2025 · 2025-08

PROJECT ABSTRACT This NIGMS MIRA will support an Early Stage Investigator and help bridge the widening gap between cutting-edge research and improving patient care in the field of hematology-immunology-infectious diseases, through the investigation of heme dysregulation of the innate and adaptive immune system in patients with sepsis or chronic hemolytic diseases ± red blood cell (RBC) transfusion as a function of circulating heme availability. Sepsis is an invasive infectious disease associated with dysregulated systemic inflammation and more than 4 million deaths worldwide per annum. Specific targeted therapies are lacking. Likewise, in chronic hemolytic diseases such as Sickle Cell Disease (SCD) and Thalassemia, > 50,000 patients have died over the past twenty years. Many patients (~50% in sepsis and >80% in chronic hemolytic diseases) receive a RBC transfusion as supportive therapy or have ongoing ‘low levels’ of intravascular hemolysis. Recent evidence from our laboratory indicate that this hemolysis may further disrupt a patient’s immune system and lead to worse outcomes. This disruption may be causally linked to release of free heme from the underlying hemolysis. We have recently shown that repetitive heme exposure may tolerize the innate and adaptive immune response rendering the host immunosuppressed, which may explain why these patients are at high risk for infection. This proposal will explore a causal relationship in human samples between dysregulated plasma heme metabolism in states of hemolysis (both acute and chronic) and after RBC transfusion that lead to altered host immunity (potentially via TLR4 binding or co-stimulation), further exacerbate inflammation, and worsen outcomes. Program 1 will establish new understandings on relevant heme binding capacity and affinity during critical illness and homeostasis. This overarching goal will define the effects of heme trafficking in plasma from RBC transfusion in sepsis in comparison to chronic hemolytic diseases (hemoglobinopathies) by further refining how binding protein-affinity for heme are affected by conditions of stress (sepsis, transfusion) compared to homeostatic conditions. Second, we hope to further characterize how heme circulates in plasma with differing binding molecules and whether this binding affect direct activation or suppression of canonical innate and adaptive host immunologic pathways (Program 2). Finally, our program will develop new strategies in humans (ex vivo) to scavenge circulating heme or block relevant molecular heme-immune pathways to mitigate negative immune alterations. Collectively, the PI, as an Early-Stage Investigator under this NIGMS MIRA, will close major gaps in knowledge in heme metabolism and its impact on immune activation, providing key insights into acute and chronic effects resulting from exposure to circulating heme in sepsis, SCD, critical illness, or after RBC transfusion. The mechanistic understandings gained from these studies will help develop novel therapeutics to minimize the morbidity and mortality of at-risk patients.

NIH Research Projects · FY 2026 · 2025-08

ABSTRACT Pediatric sepsis induced multiple organ dysfunction syndrome (siMODS) affects ~60% of critically ill children, with sepsis and has a very high mortality. These patients may develop persistent impaired host protective immunity leading to potentially chronic critical illness marked by a failure to completely eradicate the initial invading pathogens, recurrent secondary nosocomial infections, increased hospital length of stay, and late mortality. Key gaps in knowledge exist in understanding why some children develop these persistent defects in host protective immunity. New translational approaches have been hindered by a lack of useful timely technology deployed from bedside to bench and back for precisely delivered therapies. Key challenges hindering the development and deployment of immunomodulatory therapies in restoring immune function and improving long-term outcomes in siMODS include: (1) defining specific functional immune endotypes, refining the heterogeneity over time in immunosuppressive or hyperinflammatory phenotypes and the unique biological and physiological mechanisms that define these groups; and (2) identifying immunomodulatory molecules that effectively restore normal immunity against these endotypes. This proposal aims to close these critical knowledge gaps, increase collection of data on both innate and adaptive markers of immune alteration in MODS, and pave the way while improving the design of targeted clinical trials of immune stimulation or suppression. Our group has studied over the last 10 years and determined real-time immune function in adults with sepsis by stimulating their whole blood (WB) and measuring ex vivo production of IFNγ and TNFα using enzyme-linked immunospot (ELISpot). The results remarkably correlated with key clinical metrics that reflect the patient’s immune competence. Here, we plan to test these interesting results in children. Our scientific premise is that immune profiling of pediatric siMODS patients using temporal functional immune analyses combined with transcriptomic analyses will: (1) identify functional immune endotypes that reflect protective immune status in siMODS children; (2) uncover the mechanistic pathways to help design targeted therapies to restore protective immune status; (3) identify heterogeneity in ex vivo response to immune therapies among afflicted endotypes; and (4) separate the endotypes with the highest risk and poor clinical outcomes, which may require personalized therapies. To achieve these goals, we build upon existing collaborations across four children’s hospitals via a prospective, observational trial of 382 siMODS and critically ill nonseptic MODS patients This study fills a critical gap in knowledge by assessing and clarifying the heterogeneity and dynamic functional immune state in siMODS and establishing endotypes to predict relevant outcomes. The assessment of ex vivo response to mechanistically derived candidate immunoadjuvant therapies will allow for immediate and precisely designed clinical trials to evaluate potentially life-saving therapies to accurately endotyped patients along with the elucidation of their molecular drivers that will illuminate a new landscape for effectively positioning existing and new therapies.

NIH Research Projects · FY 2025 · 2025-08

We have only a basic understanding of how variation in human T cell responses elicited after Mycobacterium tuberculosis (Mtb) exposure and infection are linked to clinical outcomes. Emerging data in animal models and humans demonstrate that direct T cell recognition of Mtb-infected macrophages, the niche cell for Mtb infection, is central to a protective response. The premise of this proposal is that insight into which antigens are processed and presented by Mtb-infected macrophages, together with the composition, distribution, and kinetics of the T cell repertoire that target these antigens is vital to developing vaccines that prevent active tuberculosis (TB). In Uganda, our original study of household contacts (HHCs) of active pulmonary TB cases (since ’02) not only captured co-prevalent and incident TB but also new Mtb infection characterized by IFNg release assay (IGRA). Our current, follow-up TB HHC study focused on enrolling asymptomatic IGRA-negative contacts to identify persons undergoing IGRA conversion, i.e. “converters” (CVTR). HHCs with baseline and stable positive IGRAs in follow-up (i.e. LTBI), and “nonconverters” (NCVTRs), who remained IGRA negative, were also enrolled. Evaluating T cells and MF from peripheral blood and broncho-alveolar lavage (BAL) samples from the CVTRs form the basis of our proposed studies focusing on longitudinal immune responses. The experimental approach for this proposal builds on two complementary, ongoing lines of investigation. First, to quantify Mtb-derived peptide presentation in the context of diverse HLA alleles in Mtb-infected cells, we developed mass spectrometry approaches. Despite MHC-II allelic diversity, we found that peptides derived from CFP10, an Esx1 protein which is absent in the BCG vaccine, were presented by all MHC-II alleles analyzed while other antigens were found only in the context of certain MHC-II alleles. Second, using single-cell RNA sequencing (scRNAseq) with T antigen cell receptor (TCR) mapping, we found that Mtb-specific CD4+ T cell recognition of infected macrophages is variable and depends both on macrophage phenotype and T cell antigen specificity. Leveraging our team’s combined expertise, we will define the antigens presented by Mtb-infected macrophages and compare Mtb-specific T cell responses to these antigens at early vs. late time points after IGRA conversion, in peripheral blood and in the lungs. In Aim 1, we will use immunopeptidomics to identify Mtb antigens presented on Mtb-infected human macrophages. We will then enumerate the circulating CD4+ T cells specific for these antigens after Mtb exposure and IGRA conversion among CVTRs. In Aim 2, we will quantify direct T cell recognition of Mtb-infected macrophages, using scRNAseq to determine the TCR repertoire, antigen specificity, and function of T cells from BAL and peripheral blood, longitudinally, after IGRA conversion. In Aim 3 we will compare the capacities to control bacterial growth among human CD4+ T cells that target antigens presented by Mtb-infected monocyte-derived and alveolar macrophages vs. other immunodominant Mtb antigens. The antigens, TCRs, and T cell functions gained from this project will directly inform TB vaccine design.

NIH Research Projects · FY 2025 · 2025-07

Project Summary N-methyl-D-aspartate receptors (NMDARs) are gated by the primary excitatory neurotransmitter, glutamate. They play essential roles in neuronal formation, synaptic maturation, as well as various central nervous system functions, such as learning and memory. NMDARs are heterotetrameric assemblies, typically composed of two GluN1 subunits and two GluN2 subunits. The GRIN genes that encode the GluN subunits of NMDARs are highly intolerant to genetic variation which indicates mutations are more likely to result in disease states. Recently, whole exome sequencing has identified a large number of mutations to the GRIN genes that result in a loss of receptor function and reduced expression on the cell surface. The subunit composition of NMDARs is vital for their regional and functional expression on the cell surface, but receptors must first conform to the native structures within the endoplasmic reticulum (ER) before being trafficked to the plasma membrane. Despite the importance the ER has in establishing the correct folding of proteins, much remains to be understood regarding the essential steps in NMDAR folding, oligomerization, and anterograde trafficking. The overarching goal of this study is to elucidate the proteostasis network regulating NMDA receptor folding, assembly, trafficking, and degradation. We further seek to determine how disease-associated variants (DAVs) perturb these molecular mechanisms. Utilizing numerous biochemical and molecular techniques, Aim 1 will test the hypothesis that DAVs within the GluN2B subunit are unable to assemble into functional receptors, failing ER quality control checkpoints, resulting in their accumulation within the ER and subsequent activation of the unfolded protein response. Aim 2 will test the hypothesis that that the clearance and degradation of the DAV GluN2B subunits occurs through the lysosome via two distinct pathways: macroautophagy and selective ER- phagy. NMDARs are anticipated to undergo degradation via autophagy under physiological and pathological conditions. However, in the case of DAVs, we believe selective ER-phagy is also activated to remove aberrant GluN2B subunits from the crowded ER lumen. Additionally, Aim 2 will investigate the role of the LC3B interacting region (LIR) motif present in the highly conserved C-terminal domain of the GluN2B subunit. The proposed study will also characterize the effects of these DAVs in the context of cortical neurons derived from human induced pluripotent stem cells. Results from these studies will provide great insight into the molecular mechanisms and cellular pathways responsible for maintaining the homeostasis of NMDARs, perhaps even defining subunit-specific mechanisms. Indeed, these results hold promise to identify novel therapeutic targets for treatment of disorders in which NMDARs are dysregulated, including GRIN diseases, schizophrenia, autism spectrum disorder, and Alzheimer’s disease.

NIH Research Projects · FY 2025 · 2025-07

PROJECT SUMMARY Membrane proteins play a critical role in platelet function by relaying extracellular signals outside in. Changing the quantity or quality of these proteins can alter platelet physiology and impact health. The native membrane environment provides additional cues that regulate the protein structure and mechanism of action. Cryogenic electron microscopy (cryo-EM) has revolutionized structural biology by solving high-resolution structures of membrane proteins and large macromolecules from homogeneous samples. A recent breakthrough in data processing, the "Build and Retrieve" (BaR) methodology, performs in silico purification from large heterogenous cryo-EM datasets, making it feasible to obtain protein structures from crude preparations in their native environments. Using Cryo-Focused Ion Beam (cryo-FIB) on whole cell sample preparation provides opportunities to directly visualize macro-complex in situ. How the native environment regulates platelet function at the near-atomic level will be uncovered by combining these technical advances. The preliminary studies using cryo-EM coupled with BaR focused on proteins that are ~200kDa. It solved the first unmodified integrin αIIbβ3 structure in inactivated and intermediate states at 2.76Å and 2.49Å with metal ions, N-linked glycans, and a ligand preserved. A third structure identified a novel dimer conformation of αIIbβ3 at 2.61Å, which potentially indicates an unknown αIIbβ3 self-regulatory mechanism in the native environment. The scientific goal of this application is to solve membrane protein structures from resting and activated platelets from healthy donors without artifacts due to purification processes. Aim 1 (K99 phase) will focus on extending the preliminary αIIbβ3 data to determine the in-situ dynamics of integrin activation. Aim 2 (R00 phase) will focus on building the structural atlas of platelet membrane proteins using the combination of single-particle cryo-EM, in-situ cryo-EM, and cryo-ET with the BaR method. This aim will start with a specific focus on mitochondrial supercomplex and ATP synthase. Ultimately, solving the structures of membrane proteins in their native environment will reveal conformations and interactions that govern protein function. Obtaining these insights will pave the way for significant advancements in disease orientation, disease diagnostics, and therapeutic interventions in patients with platelet disorders. The career goal in this application is to set the trajectory for the candidate’s independent research program to conduct highly innovative research to bridge the hematology and structural biology fields. The candidate will acquire experimental training for in-situ cryo-EM studies and cryo-ET during the K99 phase. Working with the mentoring team, this application outlines an intensive training plan to foster the further growth of the core competencies (communication, critical thinking, professionalism, creativity, teamwork) for transitioning the candidate into a well-equipped independent investigator in hemostasis and thrombosis field with a unique combination of research skillsets and a highly innovative and scientifically promising research platform.

NIH Research Projects · FY 2025 · 2025-07

PROJECT ABSTRACT Heart failure with reduced ejection fraction (HFrEF) is a condition of decreased left ventricular contractility affecting 32 million people worldwide, causing low exercise tolerance and eventually death. For nearly 40 years, standard HFrEF care has primarily managed symptoms. Myosin activation is an emerging therapeutic strategy that directly addresses the contractile deficiency. Unlike failed inotropic therapies targeting non-specific adrenergic and calcium signaling, myosin activators bind directly to cardiac myosin to increase contraction. Currently, danicamtiv, a next-generation myosin activator, is in clinical trials. My experiments and initial clinical results indicate that danicamtiv improves upon the shortcomings of the first-generation activator, omecamtiv mecarbil, which impaired diastolic function. Thus, myosin activation is poised to advance care for HFrEF patients. Despite these promising results, the long-term effects of chronic myosin activation on the heart remain unknown. This knowledge gap is significant for HFrEF patients who might use myosin activators for life. My preliminary experiments show that 3 days of myosin activation elicits lasting cardiac effects after a 3-day washout period. These include shorter isovolumic contraction time, increased left ventricular wall thickness, decreased left ventricular diameter, and inverted ECG T-waves. This suggests that chronic myosin activation may stimulate concentric hypertrophy. These persistent effects likely involve z-disk mechanosensation. The z-disk anchors excitation-contraction coupling machinery and thin filaments in adjacent sarcomeres. It senses force through stress-sensitive ion channels and protein complexes involving muscle LIM protein (MLP). Changes in force then trigger cellular responses like hypertrophy via signaling proteins docked at the z-disk. Since the molecular effects of myosin activation resembles some hypertrophic cardiomyopathy (HCM) myosin mutations, they may share similar cardiac compensation mechanisms and long-term effects. While hypertrophy in HCM is highly pathological, it may benefit HFrEF patients by counteracting left ventricular dilation and atrophy in end-stage HFrEF. However, HFrEF pathophysiology may limit the heart's adaptation to chronic myosin activation due to dysregulation of z-disk mechanosensation, excitation-contraction coupling, and metabolism. These lead to my central hypothesis that chronic myosin activation elicits a hypertrophic-like cardiac response via z-disk mechanosensation, but HFrEF pathophysiology will blunt this effect. Aim 1 will test if chronic myosin activation with danicamtiv elicits a hypertrophic-like cardiac response. Aim 2 will utilize mice lacking MLP that develop progressive HFrEF to test if disrupted z-disk mechanosensing and HFrEF pathophysiology blunt the response to chronic myosin activation by danicamtiv. Successful completion of these experiments may identify a new mechanism by which myosin activation improves HFrEF and inform how to best manage myosin activation therapy in patients to maximize its effectiveness.

NIH Research Projects · FY 2025 · 2025-07

PROJECT SUMMARY Glioblastoma (GBM) is the most common and aggressive adult brain tumor with a median survival time of 15 months. Medical imaging is crucial in delineating GBM tumor extent for surgical and radiotherapy planning, with magnetic resonance imaging (MRI) being the gold standard due to its excellent soft tissue contrast. Unfortunately, even MRI is known to underestimate the full extent of GBM tumors, with stereotactic biopsies revealing microscopic tumor infiltration beyond visible enhancing tissue borders and into adjacent edematous regions. Although recurrence overwhelmingly occurs in this edema-rich, peritumoral region, there is no consensus regarding inclusion of peritumor within the clinical target volume. This is because no existing non-invasive methods can successfully discriminate GBM infiltration from non-malignant peritumoral edema. To address this, there have been extensive efforts to develop MRI-based artificial intelligence prediction models for GBM infiltration. Despite promising results, these infiltration models often have poor reproducibility and generalizability, which is due to MRI’s high sensitivity to differences in acquisition protocol, scanner hardware, and processing methods. As a result, GBM recurrence is inevitable and currently incurable. Magnetic resonance fingerprinting (MRF) could potentially solve this problem. MRF is a quantitative MRI acquisition framework for rapid and robust multiparametric mapping of intrinsic tissue properties. MRF maps reflect quantitative measures of physical tissue characteristics, which leads MRF to have superior sensitivity and reproducibility compared to conventional MRI. These factors make MRF an ideal imaging technique for accurate, reproducible, and generalizable radiomic modeling of GBM infiltration. The overall objective of this project is thus to develop and validate magnetic resonance fingerprinting (MRF) artificial intelligence (AI) models for prediction of infiltrated GBM peritumor. This project has two specific aims. In Aim 1, defining and discriminative MRF image features of infiltrated peritumor will be identified to train an MRF radiomic model for pre-operative GBM infiltration prediction. We will implement two technical improvements for infiltration modeling, namely transfer learning via data-driven infiltration risk priors and semi-supervised learning using unlabeled peritumor. The developed MRF radiomic model will then be applied in Aim 2 for longitudinal assessment of radiotherapy (RT) changes. GBM patients undergoing RT will be scanned with MRF and MRI across four time points. At each time point, the developed infiltration model will be applied to generate whole tumor infiltration maps and evaluate peritumoral infiltration load. Longitudinal MRF and MRI image differences between RT-treated and non-treated peritumor will be used to optimize the infiltration model developed in Aim 1 to predict future recurrence regions. This project’s success will demonstrate the feasibility of quantitative MRF radiomics for GBM infiltration prediction, paving the groundwork for targeted GBM therapy to reduce recurrence and extend survival. The technical framework developed in this project will also be highly translatable to other studies leveraging quantitative MRI image analytics to improve brain tumor care.