Synthetic estrogens are used in the clinic to alleviate debilitating neurological symptoms associated with androgen deprivation therapy (ADT), an effective treatment improving survival in prostate cancer patients when administered timely in the course of carcinoma. However, this remedy to relieve the symptoms, most commonly manifested as hot flushes, causes feminizations (most prominently gynecomastia) that significantly diminishes patients' compliance because of physical and psychological discomfort. Because only estrogens can provide adequate remedy of hot flushes based on current clinical practices, there is an unmet medical need for an effective, side effect-free and, consequently, compliance-gaining intervention to alleviate these vasomotor symptoms distressing prostate cancer patients on ADT. The scientific premise of this grant application is based on it central hypotheses that treatment with 10β,17β-dihydroxyestra-1,4-dien-3-one (DHED), an innovative brain-selective bioprecursor prodrug of 17-estradiol, will ease ADT-associated hot flushes without feminizing side-effects. In the first specific aim, we will perform preclinical pharmacokinetics, distribution and bioavailability studies in orchidectomized male rats to support our particular hypotheses that DHED treatment confines the formation of 17-estradiol from the prodrug into the brain. In the second specific aim, we will assess DHED treatments through measuring effects elicited by the estrogen delivered selectively in the brain and, consequently, avoiding exposure of peripheral tissues to the hormone through multiple methodologies. One of our specific hypotheses is that oral treatment with DHED will abate orchidectomyinduced tail skin temperature elevation in a pharmacological model, and will restore diurnal changes lost after orchidectomy in a physiological paradigm, in two well-established and complementary rat models of hot flushes. Since our hypothesis is that hot flushes are due to an imbalance of neurotransmitters/neuromodulators innervating thermosensitive neurons in the preoptic area of the hypothalamus, we will monitor norepinephrine, serotonin, and neurokinin B involved in thermoregulatory responses with in vivo microdialysis to provide additional, mechanism-focused approach for assessment. As the synthesis and release of these neurotransmitters and their cognate receptors are regulated by estrogen, we hypothesize that the major action of estrogen is to re-establish the physiological balance among these neurotransmitters to stabilize neuronal activity on the thermoregulatory center; i.e., to prevent heat dissipation (hot flush). Finally, we will seek supporting evidence for the brain-specific action of DHED-derived 17-estradiol by evaluating estrogeninduced gene expression in the brain, and its lack in the breast, pituitary, prostate and prostate cancer cells. The overall goal of our aims is to support subsequent translational research focusing on DHED’s therapeutic use to remedy hot flushes and potentially other neurological symptoms in prostate cancer patients undergoing ADT to manage their malignancy.