Project Summary Androgen receptor (AR) plays a fundamental role in driving the development of human prostate cancer. Most current treatments against the AR-expressing (AR+) metastatic cancer aim to inhibit AR, e.g. the standard of care with androgen deprivation therapy (ADT), or new treatment with AR inhibitors enzalutamide (Enza). Despite the recent advance, the efficacy and sustainability of anti-AR-treatments (ADT/Enza) are still limited due to treatment resistance, which leads to tumor progression and patient mortality. New understanding in the mechanism of resistance is therefore urgently needed. Tumor hypoxia occurs frequently in solid cancers including metastatic prostate cancer, and has long been considered a cause of treatment resistance. However, the exact mechanism is unclear. Recently, we have found that hypoxia confers ADT/Enza resistance via metabolic reprogramming (Geng et al, Nat. Comm. 2018). This project is to further clarify the mechanism of action centering on the cytosolic glutamate-pyruvate transaminase (GPT1). In literatures, GPT1 is known as a metabolic enzyme at the converging point of glucose and glutamine metabolic pathways, and many GPT1- downstream metabolites are oncogenic and cytoprotective to cancer cells. However, the role of GPT1 in conferring anti-AR-treatment resistance is unknown. We identified GPT1 through metabolic and transcriptomic screenings of our anti-AR-resistant cells. Our new pilot data further showed that i) GPT1 expression and activity were upregulated by ADT/Enza in hypoxia, ii) the upregulation was consistent across ADT/Enza- resistant cells, xenografts, PDX and patient samples, and iii) clinical GPT1 mRNA correlated with hypoxia, poor disease outcomes, and ADT/Enza resistance in patients. In this proposal, we will determine whether and how prostate cancer cells use GPT1 to evade ADT/Enza in hypoxia and in vivo with cell and tumor models and patient samples. We will determine the molecular function of GPT1 in conferring ADT/Enza resistance in Aim 1, understand the mechanisms in Aim 2, and confirm its clinical significance in Aim 3. Metastatic prostate cancer has the 2nd-leading cause of cancer death in American men. New mechanistic understanding and therapy strategy are unmet needs. The GPT1-based studies here may lead to new mechanistic insights, paving ways to new prognosis and treatment strategies to predict drug efficacy, monitor the onset of resistance, and prevent or reverse resistance.