ABSTRACT. Glucose homeostasis plays a critical role in multiple cellular processes, and impaired or altered glucose metabolism is associated with a wide range of pathological states. A key step in glucose metabolism is catalyzed by the glycolytic enzyme pyruvate kinase. Proliferating cells almost universally express the pyruvate kinase M2 (PKM2) isoform, which can assume either an active or inactive state. PKM2 is at the nexus of cellular metabolism, and determines whether cells metabolize glucose into ATP or use it to make more of the necessary building blocks for cell division. Multiple studies have demonstrated how dynamic changes in PKM2 expression contribute to altered glucose metabolism in different contexts. The ability to non-invasively visualize and track dynamic changes in PKM2 expression will enable improved understanding of altered glucose metabolism and the downstream mediators of glycolysis in multiple disease states. The lack of PKM2 expression within the brain and myocardium make this imaging strategy highly promising for neurological and cardiovascular applications. We have recently reported the development and human translation of [18F]DASA-23, the first clinically-relevant and specific radiopharmaceutical to detect, localize, and quantify PKM2 using positron emission tomography (PET) imaging. We have determined the biodistribution, radiation dosimetry, and brain distribution of [18F]DASA- 23 in healthy volunteers, and have explored its ability to visualize PKM2 expression in one potential application of patients with primary brain tumors. Although our results highlight the potential of imaging PKM2, [18F]DASA- 23 has several limitations that impedes widespread use, and the ability to study PKM2-mediated glycolytic reprogramming in broader applications. This includes high radiation dose to the gallbladder wall, a high degree of non-specific binding within white matter in the brain, and poor solubility in radiotracer formulation vehicle. This proposal will develop novel PKM2 radiotracers to overcome the limitations of [18F]DASA-23. Development of a safe and reliable PKM2 radiotracer will enable repeat assessment of the dynamic alterations in glucose metabolism in multiple different applications and patient populations. We will establish the synthesis and fluorine- 18 radiolabeling of two candidate small molecules with improved physicochemical properties relative to DASA- 23, pharmacological activity and specificity for PKM2, and the potential for radiolabeling. We will automate the radiosyntheses and characterize uptake and specificity in cell culture (Aim 1), determine biodistribution and radiation dosimetry (Aim 2), and assess the ability to visualize PKM2 expression in one potential application of primary brain tumors (Aim 3). Success of this proposal will develop novel radiotracers for visualizing a hallmark of metabolism. This will have important ramifications for studying altered glucose metabolism in multiple applications and could improve our collective understanding of metabolic adaptations in disease. Importantly, this technology will be adopted by a wide range of users in different pre-clinical and clinical studies.

PROJECT NARRATIVE. Metabolic processes are involved directly or indirectly in essentially everything a cell does, there still remains much to learn regarding how proliferating cell metabolism is regulated. Pyruvate kinase M2 (PKM2) catalyzes the last step in glycolysis and determines whether cells can produce enough nucleotides to proliferate; its expression has been linked to a wide range of pathologies including cancer, heart failure, neuroinflammation, and neuropathic pain. This proposal will develop novel radiotracers targeting PKM2 that will enable an innovative method of imaging a hallmark of metabolism and has potential to improve our understanding of metabolic adaptations in multiple disease states.