Molecular Mechanisms of a Novel Aliphatic Hydroxamate, WMJ-S-001, in Suppressing Colorectal Cancer Progression---The Causal Role of AMPK-p53 Modification-miRNAs Cascade

Cancer is an increasingly prevalent health problem and remains a major cause of mortality around the world. The metastatic spread of tumor cells is associated with resistance to conventional therapy and is the leading cause of death for cancer patients. Tumor cells produce a range of growth factors and/or cytokines that promote tumor metastasis by directly or indirectly stimulating angiogenesis and lymphangiogenesis, which is one of the major routes for tumor invasion and metastasis. Hydroxamate and its derivatives have attracted considerable attention, due to their pharmacological properties including anti-tumor activity. In an effort to develop novel inhibitors to suppress angiogenesis, lymphangiogenesis and tumor growth, we recently selected WMJ-S-001, a novel aliphatic hydroxamate, and characterized its anti-angiogenic, anti-lymphangiogenic and anti-tumor activities. We found that WMJ-S-001 may target VEGFR2 signaling to inhibit vascular endothelial growth factor (VEGF)-induced proliferation, migration and endothelial tube formation in human umbilical vascular endothelial cells (HUVECs). WMJ-S-001 also attenuated VEGF-induced microvessel sprouting from aortic rings ex vivo and suppressed tumor-associated angiogenesis in vivo. However, the molecular mechanism underlying WMJ-S-001-induced cancer cell death remains to be established. In addition, the post-translational modifications of p53 family proteins (p53, p63 and p73) enable them to transactivate p53-responsive genes, and induce cell cycle arrest or apoptosis. A number of microRNAs (miRNAs) have been shown to function as transcriptional targets of p53 in promoting apoptosis. The question of p53/p63/p73 regulating the miRNAs has not been addressed in depth so far. Whether p53 family proteins and miRNAs are involved in WMJ-S-001’s actions in cancer cells remains unclear. In our preliminary studies, we found that WMJ-S-001 decreased cell viability and induced apoptosis in HCT116 colorectal cancer cells. WMJ-S-001 caused p53 phosphorylation and acetylation in HCT116 cells. WMJ-S-001 caused AMPK phosphorylation and AMPK dominant negative mutant (DN) suppressed WMJ-S-001-induced p53 phosphorylation. In addition, WMJ-S-001 increased the binding of p53, p63 and p73 to the promoter region of miRNA34a. WMJ-S-001 also increased miR34a promoter luciferase activity. This project aims to establish the causal role of p53 protein family and miRNAs in WMJ-S-001-induced colorectal cancer cell death. The central hypothesis of this project is that WMJ-S-001 activation of AMPK affects the post-translational modifications of p53 family proteins, which leads to p53 downstream targets including miRNA34a expression and subsequent colorectal cancer cell death. WMJ-S-001 may be a lead compound and understanding the anti-tumor mechanisms of WMJ-S-001 will aid in future development of selective-targeting strategies for WMJ-S-001 or its derivatives in the treatment of cancer.