West Virginia University

NIH Research Projects · FY 2025 · 2016-07

Project Summary Coenzyme A (CoA) is an essential cofactor and the major acyl group carrier in mammalian cells. CoA plays a central and regulatory role in energy metabolism, as its acyl-CoA derivatives are substrates for hundreds of metabolic reactions and the posttranslational modification of histones and key metabolic enzymes. CoA- dependent processes occur in multiple subcellular compartments, and major pools of CoA are found in the mitochondria, peroxisomes and cytosol. At the whole tissue level, the concentration of CoA is tightly regulated and dynamically adjusted to changes in the metabolic state. The importance of such a tight control over CoA levels is underscored by the fact that genetic manipulations that force the concentration of CoA outside of its homeostatic range result in loss of metabolic regulation and organ function. For example, the inability to increase CoA levels during a fast blunts fatty acid oxidation and gluconeogenesis in the liver, causing fasting hypoglycemia. On the other hand, an abnormally high concentration of CoA in skeletal muscle is associated with decreased muscle mass, ATP levels and exercise performance, highlighting the importance of mechanisms that prevent the accumulation of this cofactor to toxic levels. The concentration of CoA is regulated by balancing its synthesis and degradation. The process of CoA degradation is poorly characterized. Furthermore, the mechanisms that regulate the different subcellular CoA pools are incompletely understood. During the previous funding cycle, we have characterized the biochemical and regulatory properties of two CoA-degrading enzymes, Nudt7 and Nudt19, which reside in liver and kidney peroxisomes, respectively. Our published and unpublished observations support the conclusion that these enzymes regulate peroxisomal lipid metabolism. Furthermore, deletion of Nudt19 leads to the accumulation of 3-hydroxy-3-methylglutaryl-CoA in the kidneys, suggesting a connection to cholesterol synthesis. We also identified the first mammalian CoA-degrading enzyme, Nudt8, which resides in the mitochondria, and we have recently generated Nudt8-/- mice. The existence of CoA- degrading enzymes in both peroxisomes and mitochondria suggests that these enzymes contribute to the regulation of the CoA pools within these organelles. The long-term goal of our research program is to understand the mechanisms that regulate tissue CoA levels and to harness them to manipulate the metabolic network for the treatment or prevention of metabolic disorders. To move toward this goal, we propose to 1) determine the mechanisms through which Nudt19 regulates kidney lipid metabolism and kidney function and 2) determine the role played by Nudt8 in the regulation of the mitochondrial CoA pool and mitochondrial metabolism. This research program will advance our understanding of the mechanisms that regulate the peroxisomal and mitochondrial CoA pools. Furthermore, identifying the processes regulated by each CoA-degrading enzyme will aid in the development of strategies to target and correct specific CoA-dependent pathways in metabolic disorders.

NIH Research Projects · FY 2024 · 2015-08

PROJECT SUMMARY/ABSTRACT A key role of the TME in cancer progression, treatment resistance, and as a target for therapeutic intervention is increasingly appreciated. Noninvasive in vivo EPR-based spectroscopy and imaging of tissue oxygenation (pO{2}), extracellular pH (pH{e}), redox, glutathione (GSH) and interstitial inorganic phosphate (Pi) provide unique insights into biological processes in the TME, and may serve as a tool for preclinical screening of anticancer drugs and optimizing TME-targeted therapeutic strategies. In this competitive renewal application, our new directions are built on knowledge and discoveries made during the previous funding period. There are two goals of our R01 renewal proposal. First is to further advance paramagnetic probes and magnetic resonance technology with a focus on improving probes biocompatibility, functionality, and developing multifunctional imaging modalities. Second is to utilize these advances and knowledge accumulated using spectroscopic modalities in the initial R01 project period to obtain further insights into the role of TME in cancer progression, and to test our hypothesis on the roles of interstitial inorganic phosphate in tumorigenesis. The specific aims are: (SA1) To advance molecular multifunctional EPR-based imaging technology and paramagnetic probes. The biocompatible derivatives of multifunctional trityl HOPE probe sensitive to acidity (pH{e}), oxygen (pO{2}) and phosphate (Pi) in Extracellular tissue microenvironment, and dual function nitroxide pH & redox probes will be optimized for concurrent multifunctional imaging using cutting edge imaging modalities, rapid scan EPR imaging and Overhauser-enhanced magnetic resonance imaging. (SA2) To perform molecular imaging of chemical TME as the mammary tumors progress to malignancy. Multifunctional spatially-resolved TME profiling will validate morphological and gene expression-based staging in a mouse model of breast cancer and will map tumor regions with different phenotypes. Tissue samples from the areas with the chemical TME characteristics of a malignant phenotype will be isolated and the TME study will be complemented with immunohistochemical, biochemical, and genetic tissue analysis, and with measurements of total phosphorus and phosphate (Pi) contents in blood. In summary, the developed multifunctional imaging techniques and probes will broaden the scope of preclinical EPR allowing for mapping of physiologically relevant tissue parameters in various disease models in cancers and beyond. New knowledge on stage-specific TME evolution during tumor progression is required to optimize TME-targeted anticancer therapies. It will also provide a scientific basis to evoke public awareness of high content of the phosphate-based modifiers in the processed food and the potential health risk.

NIH Research Projects · FY 2026 · 2015-04

Hypoxia and macrophages drive tumor aggressiveness and treatment outcome leading to worse prognosis for breast cancer patients. Contrary to acute hypoxia where tissue homeostasis is vital for development and repair, chronic hypoxia observed in solid tumors stimulate unproductive angiogenesis by excessive vascular endothelial growth factor expression leading to dysfunctional vessels that perpetuate hypoxia and acidic extracellular pH; factors that limit effective perfusion of treatment modalities like chemotherapies and radiotherapies and promote tumor aggressiveness. Macrophages are intimately involved in regulating unproductive angiogenesis thru secretion of soluble factors that support this activity. In the parent grant, we advanced electron paramagnetic resonance (EPR)-based techniques towards in vivo real-time tumor microenvironment (TME) profiling in animal cancer models. Using these approaches, we showed macrophage hypoxia-inducible transcription factors (HIF)- 1α and HIF-2α had disparate roles in regulating TME parameters like oxygen and pHe through structural and functional alterations in vessels that dictated docetaxel efficacy. We showed that HIF-1α augments expression of endothelial tyrosine kinase (TIE2) receptor on macrophages called TIE2-expressing macrophages (TEMs) previously-reported to be “pro-angiogenic”, but now better defined as “pro-hypoxic” by dysregulating vessels leading to poor perfusion. The overall objective of the renewal is to investigate macrophage location and function that perpetuates a hypoxic TME detrimental to perfusion of therapeutic modalities. To achieve this central objective, we propose these specific aims: (SA1): To optimize magnetic resonance imaging modalities for in vivo multifunctional mapping of local tumor tissue parameters. Advances in paramagnetic probes and imaging technologies such as rapid scan EPR imaging and Overhauser-enhanced MRI allow for mapping specific areas of hypoxia and acidosis and characterizing their relationship to tumor macrophage locoregional populations. (SA2): Elucidate tumor macrophage location-specific functions in regulating hypoxia and acidosis in a mouse model of breast cancer. We will sample tumor origin and regions of hypoxia in PyMT breast cancer models using image-guided biopsy to understand the bi-directional shaping of TME and macrophages that contribute to poor vessel perfusion and hypoxia. (SA3): Investigate recruited and tissue- resident macrophage populations and their respective roles in contributing to tumor hypoxia and acidosis that dictate chemotherapy effectiveness. We will track fluorescent bone marrow monocytes to tumor origin, and generate conditional macrophage-deficient breast tumor mice to systematically determine a causal role of specific macrophage populations. Summarizing, in vivo mapping of tumor hypoxia and acidosis using innovative magnetic resonance technology in mice deficient in specific macrophage populations or lacking hypoxia-regulated macrophage functions in a mouse model of breast cancer may provide new insight into a macrophage/TME axis that suppresses the efficacy of clinically-relevant anti-cancer therapies.

NIH Research Projects · FY 2024 · 2014-09

The Overall Objective of the WVU Stroke CoBRE is to decrease morbidity and mortality related to stroke through improved understanding of pathophysiological mechanism(s) of stroke. We will accomplish this overall goal by conducting high-quality basic and translational research addressing the modifiable risk factors, biomarkers, mechanism(s), preventative strategies, acute and chronic treatments and rehabilitation of stroke. This objective would be met by achieving the following three Specific Aims: Specific Aim 1. Expand the critical mass of funded investigators conducting stroke recovery research. In Phase II we propose to solidify the commitment of investigators to stroke research, in part, by providing cutting-edge stroke research core facilities and services (See Specific Aim 2), generating meaningful preliminary data in support of their grant applications through funding of five Junior Investigator projects, providing grantsmanship workshops/activities to improve their grant applications, and expanding the intensive mentoring program initiated in Phase I of the WVU Stroke CoBRE. Specific Aim 2. Strengthen innovative scientific cores that support and advance basic and translational stroke research. We propose to expand the scope and capacity, as well as the user-base of these vital cores. This will be accomplished by the introduction of new services (e.g., new stroke models; additional behavioral tests), and provide training sessions for research cores. In addition to the Experimental Stroke Core (ESC) and the Rodent Behavior Core (RBC), which are proposed for CoBRE funding, we will develop two additional cores, the Mitochondrial Functional Assessment Core (MFAC) and the Stroke Tissue Bank. Specific Aim 3. Advance the ongoing development of an independent, sustainable, multidisciplinary thematic program of research on stroke. The goal of achieving independences (of CoBRE funding) of the WVU Stroke CoBRE has begun and will continue in Phase II by the transition to a user-fee based funding of research core services, the independent funding of research grants that include funding for these core services and support from the WVU HSC. At the conclusion of Phase II of the WVU Stroke CoBRE, mentored training will have been provided to engage additional JIs in stroke-related research. We will have addressed the need for basic and translational research into the causes, acute and subacute treatments and recovery from stroke by (1) increasing the number of independently funded WVU stroke researchers, (2) expanding stroke research core services and user numbers, and (3) continuing the transition to sustain these resources through a user-based funding model for services at WVU.

NIH Research Projects · FY 2025 · 2012-08

Overall Abstract - Overall West Virginia (WV), the only state located entirely in Appalachia, has the lowest life expectancy in the nation and ranks at or near the bottom in many chronic disease categories. The WV Clinical and Translational Science Institute (WVCTSI) was created in 2012 through the initial Institutional Development Award for Clinical and Translational Research (IDeA-CTR) and successfully renewed in 2017. Over the past 9 years, WVCTSI has cultivated and nurtured a highly collaborative, statewide research infrastructure to address the health of West Virginians. Over the past 4 years, WVCTSI productivity has been superb with 1,325 publications, 951 funding applications, and 688 external funding awards totaling $159.3 MM. The WVCTSI’s overarching goal for the next funding cycle is to serve as a transformational force, leading statewide clinical and translational collaborative research that positively impacts health outcomes in WV. We will achieve this goal through the following specific aims: Specific Aim 1 - Build collaborative, productive research infrastructure across WV that substantively contributes to improving WV health outcomes, Specific Aim 2 - Train and position for success the next generation of clinical and translational scientists, and Specific Aim 3 - Actively engage multiple stakeholders. There are important cross-cutting themes around which our efforts will be focused, including rural health and the clinician scientists of tomorrow – able to frame the right questions and work effectively to find the answers. Central to attainment of our specific aims is the highly collaborative statewide organization that WVCTSI has developed over the preceding 9years and that now includes every major healthcare organization and academic medical center in WV, the state health department (WVDHHR), the National Institute for Occupational Safety and Health, and >100 primary care organizations across WV that constitute the WV Practice-Based Research Network. Of critical importance is the high level of collaboration and commitment among WVCTSI’s cores. Successful completion of our objectives will inform policy, practice, and community changes resulting in improved WV health outcomes.