ALBERT

Description: Background: MCM7 is one of the subunits of the MCM2-7 complex that are essential for DNA replication licensing and control of cell cycle progression. It also participates in mRNA transcription and DNA damage regulation. It is found that MCM7 was highly expressed in many cancer cells including Leukemia. Thus, lentivirus-mediated siRNA targeting MCM7 was used to suppress its endogenous expression in K562 cells and develop a novel therapeutic strategy for leukemia. Method: Lentivirus-mediated siRNA targeting MCM7 were designed and infected K562 cells. Down-regulation of MCM7 mRNA and protein expression in K562 cells were determined by Real-time PCR and Western blot analysis. The proliferation of the transfected K562 cells was assessed by CCK8 assay. Cell cycle progression and apoptosis were calculated by flow cytometry. The related protein levels of the transfected K562 cells were detected by Western blot. The stably transfected K562 cells were injected into nude mice. The transplanted tumors were measured and weighed, then Western blot and Immunocytochemistry were used to evaluated the expression of the proteins for mechanism. Results: The mRNA and protein expression of MCM7 in the stably transfected K562 cells decreased with the percentage of 92.3±1.88 and 91.18%±0.93 respectively. Compared with control and negative control groups, the proliferative rate of K562 cells was significantly reduced after LV-MCM7-RNAi infection. The cell cycle progression of K562 cells was blocked, with a massive increase in the percentage of cells in the G0/G1 phase and decrease in S phase. The apoptosis rate of the transfected K562 cells was significantly higher. PLK1 and pPLK1 protein were down-regulated in response to MCM7 inhibition; P53 and bax protein were up-regulated simultaneously. In vivo, the tumor size and weight dramatically decreased in the MCM7-RNAi group than in negative control group. PLK1, pPLK1 proteins in transplanted tumors were down- regulated and P53,bax were up-regulated correspondingly. Conclusions: Lentivirus-mediated MCM7 shRNA has effectively down regulated the expression of MCM7 gene on mRNA and protein levels. Moreover, the silence of MCM7 has led to the significant decrease in proliferation, blocking the cell cycle progression of K562 cells in vitro. Simultaneity, the apoptosis of the leukemia cell was enhanced. Both of the functions above cause the inhibition of the tumor growth in nude mice. Overall, these results suggest that the proliferation and cell cycle of K562 cells could be regulated by silencing MCM7 gene which is a promising gene therapeutic method to treat gastric cancer. Disclosures No relevant conflicts of interest to declare.