Project Summary/Abstract Sarcomas are a heterogeneous group of malignancies arising from mesoderm-derived tissues such as muscle, fat, and connective tissue. They are diagnosed in nearly 20,000 persons in the United States each year, and approximately 40% of patients die of either loco-regional recurrence or distant metastasis. Sarcomas and other solid tumors typically thrive in hypoxic and nutrient-poor conditions to proliferate and metastasize. To survive in such environments, sarcomas hijack two adaptive mechanisms: (1) activation of hypoxia inducible factor 1α (HIF-1α), which enhances the transcription of over 150 genes mediating tumor metabolism, angiogenesis, and metastasis and (2) utilization of aerobic glycolysis (a.k.a. the Warburg effect), which creates energy by means of glycolysis rather than oxidative phosphorylation. HIF-1α appears to be particularly critical for a subset of tumor cells which we will refer to as “sarcoma stem-like cells” or SSCs, characterized by their ability to self renew and differentiate. In preliminary studies, we have found that SSCs reside preferentially in hypoxic regions of tumors, exhibit elevated levels of HIF-1α, and are likely to promote chemotherapy resistance and metastasis. The reverse reaction of glycolysis is gluconeogenesis, where fructose-1, 6-bisphosphatase (FBP) acts as a rate-limiting enzyme. We also recently determined that FBP2 is consistently downregulated in 8 human sarcoma subtypes compared to normal human mesoderm-derived tissues. The long-term objective of this proposal is to expand the use of agents targeting HIF-1α and FBP2 in patients with sarcomas to reduce recurrence, distant metastasis, and chemotherapy resistance. Consequently, this proposal is designed to test the hypothesis that HIF-1α and FBP2 play critical and inter- related roles in regulating sarcomagenesis, metabolism, metastasis, and chemotherapy resistance. To test this hypothesis, this research proposal will (1) define the role of FBP2 in sarcoma metabolism, progression, and metastasis, and (2) determine the role of HIF-1α in SSC metastasis and chemotherapy resistance. The methods of this proposal include analysis of autochthonous and xenograft mouse models of sarcomas, analysis of sarcoma cell lines in vitro, metabolic studies, and correlative studies of tumor samples from sarcoma patients.