Biomarkers of Oxidative Injury and Their Modulation in Prostatic Tissue from Patients With Prostate Cancer

摘要

Prostate cancer (PCa) continues to be the most frequently diagnosed cancer in men in the United States. The development of potent prevention strategies to diminish or eliminate this threat is in order and remains the long-term goal of this project. One possible etiologic factor in the development of PCa, is exposure to chronic oxidative stress, leading to the accumulation of potentially promutagenic oxidized DNA bases such as 8- hydroxydeoxyguanosine (8- OHdG). The detoxifying enzyme GSTPl is inactivated in nearly 100% of PCa and in prostatic intraepithelial neoplasia (PIN) lesions. In the past year, we have developed an accurate HPLC-MS-MS method for measuring intracellular reduced and oxidized glutathione. Once developed, we exposed the human PCa cell line, LNCaP (no GSTP1 protein expression) and subclones of LNCaP engineered to overexpress GSTP1, in vitro, to chronic oxidative stress inflicted by protracted, low dose irradiation (LDR) to determine if re-expression of GSTP1%1. results in 'bankruptcy' of intracellular glutathione and subsequently increases cell death. As anticipated, re-expression of GSTP1 results in an increase in oxidized glutathione following LDR but does not significantly diminish intracellular reduced glutathione levels. These data solidity our preliminary results and implicate GSTPl as a major regulator of oxidative DNA damage. In the past year, we have also verified measurement of oxidized guanine by the HPLC-MS-MS methods we developed, a more quantitative and accurate technique than older and routine HPLC-ECD methods. This new technique has allowed us to precisely measure 8-OHdG in DNA extracted from LNCaP xenografts before and after exposure to oxidative injury by LDR. These data reveal that, like our in vitro data with LNCaP subclones expressing GSTPl, LNCaP xenografts expressing GSTP1 do not accumulate significant levels of 8-OHdG following exposure of animals bearing these tumors to LDR.