Combined Silencing Of Oncogenes K-ras And AKT2 Achieves Synergistic Effects In Inhibiting Pancreatic Cancer Cell Growth In Vitro And In Vivo

Pancreatic ductal adenocarcinoma(PDAC) is exceptionally aggressive with high mortality.Since no effective therapeutic methods have been found up to now,it is urgent to find more powerful method to diagnose the disease earlier and treat it more effectively.RNAi is a strong tool to inhibit more than one gene at the same time.Therefore, in this study,we used a plasmid-based RNAi system to evaluate whether simultaneously silencing two oncogenes K-ras and AKT2,which are key players in two distinct signaling pathways-RAS/MAPK and PI3K/AKT,would have a better tumor inhibition effect than silencing a single oncogene.The experimental results both in vitro and in vivo confirmed our hypothesis.The following aspects were studied:1.Construction of recombinant plasmids,including plasmids that can silence the oncogene K-ras,plasmids that can silence the oncogene AKT2,and plasmids that can silence oncogenes K-ras and AKT2 simultaneously.2.Effect of the recombinant plasmids on the corresponding mRNA and protein levels in the Panc-1 cells.3.Effect of the recombinant plasmids on proliferation and colony formation of the Panc-1 cells.4.Effect of the recombinant plasmids on apoptosis of the Panc-1 cells.5.Effect of the recombinant plamids on downstream signaling kinases in the Panc-1 cells.6.In vivo effect of the recombinant plamids on Panc-1 cells.The results showed that: 1.RNAi inhibited oncogenes K-ras and AKT2 specifically at mRNA and protein levels in Panc-1 cells.2.Specific inhibition of oncogenes K-ras and/or AKT2 reduced cell proliferating capacity and colony formation of Panc-1 cells.3.Specific silencing of oncogenes K-ras and/or AKT2 induced apoptosis in Panc-1 cells.4.Combined inhibition of oncogenes K-ras and/or AKT2 can inhibit the phosphorylation level of p-ERK and p-GSK and reduce the protein level of c-myc protein.5.Combined inhibition of K-ras and AKT2 inhibited tumor growth in vivo more effectively than inhibition of each of the two genes aloneConclusion:Combined silencing of oncogenes K-ras and AKT2 achieved synergistic effects in inhibiting pancreatic cancer cell growth and inducing cell apoptosis in vitro and in vivo,and the possible mechanism may be due to the inhibition of the two signaling pathways of RAS/MAPK and PI3K/AKT simultaneously, inducing the antiapoptic protein BAD and promoting the proteolysis of the oncoprotein c-myc.This result may also offer potential opportunities for clinical cancer therapy.