The incidence and mortality rate of cancer have become a global health problem. Basic scientists and clinicians are working together to better understand cancer biology in an effort to develop effective strategies for cancer treatment. It is now well understood that tumorigenesis results from the imbalance between cell proliferation and apoptosis. Thus how to reverse this imbalance constitutes the focus of many drug discovery projects, which have led to the discovery of many phytochemicals capable of suppressing aberrant cell proliferation by correcting dysregulated signaling pathways. The present proposal thus focuses on the mechanistic characterization and structural optimization of a novel derivative of indole-3-carbinol (I3C), a chemopreventive agent from cruciferous vegetables. In the PI』s laboratory, we have synthesized a series of I3C derivatives for pharmacological screening. Among these derivatives, A9 represents an optimal agent with 100-fold increase in the cytotoxicity against several cancer cell lines. Equally important, A9 overcomes the intrinsic chemical instability of I3C in acid. This chemical instability leads to lower antitumor potency and unpredictable pharmacokinetic behaviors of I3C in vivo. Our preliminary study indicates that the high antiproliferative potency of A9 was attributable to its ability to induce cell cycle arrest and apoptosis. Thus, mechanistic characterization and continued lead optimization represent the foci of this investigation. In the first year of this three-year project, we will prove our hypothesis that A9 mediates antiproliferative activities by sharing many signaling mechanisms with I3C. In addition to Western blot analysis of the effect of A9 on various signaling targets, the PI proposed to use microarray analysis to carry out gene expression profiling of A9 in tumor suppressor genes and oncogenes. Real-time reverse transcription-PCR in conjunction with Western blot analysis will be used to verify the gene alterations. In the second year, A9 will be used as the lead compound to establish a library of A9 derivatives. This library will be tested for cytotoxicity and undergo pharmacological screening against different biomarkers. In the third year, the optimal agent generated from the drug library will be tested against the previously identified molecular targets related to cell cycle control and apoptosis. Then, microarray and the Real-time reverse transcription-PCR will provide more information about the mode of mechanism of the optimal A9 derivative. In vivo antitumor potential of these novel agents will be further tested by different xenograft tumor models in nude mice. Overall, the aforementioned structure-activity analysis and mechanistic characterization of A9 will provide us further insights regarding the clinical use of this class of novel antitumor agents.
