DNA triplex-mediated inhibition of MET leads to cell death and tumor regression in hepatoma

摘要

Mesenchymal epithelial transition factor (MET) is one of the critical cell signaling molecules whose aberrant expression is reported in several human cancers. The aim of the study is to investigate the antigene and antiproliferative effect of short triplex forming oligonucleotides, TFO-1 (part of the positive regulatory element) and TFO-2 (away from the transcription start site) on MET expression. HepG2 cells transfected only with TFO-1 (but not with TFO-2 and non-specific TFO) significantly decreased MET levels, which is accompanied by decrease in antiapoptotic proteins and increase in pro-apoptotic proteins. Phosphoproteome-array analysis of 46 intracellular kinases revealed hypophosphorylation of about 15 kinases including ERK, AKT, Src and MEK, suggesting the growth inhibitory effect of TFO-1. Further, the efficacy of TFO-1 was tested on diethylnitrosamine-induced liver tumors in wistar rats. T2-weighted magnetic resonance imaging showed decrease in liver tumor volume up to 90% after treatment with TFO-1. Decreased MET expression and elevated apoptotic activity further indicate that TFO-1 targeted to c-met leads to cell death and tumor regression in hepatoma. Formation of stable DNA triplex between TFO-1 and targeted gene sequence was confirmed by circular dichroic spectroscopy and gel retardation assay. Therefore, it can be concluded that DNA triplex-based therapeutic approaches hold promise in the treatment of malignancies associated with MET overexpression.