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Enhanced apoptosis-inducing function of MDA-7/IL-24 RGD mutant via the increased adhesion to tumor cells.

Melanoma differentiation-associated gene-7 (mda-7)/interleukin-24 (IL-24) has shown potent tumor cell apoptosis inducing capacity in multiple cancers. However, the apoptosis induction capacity of mda-7/IL-24 was low and directly correlated with the adhesion to tumor cells.Cell adhesion molecule integrin α(v)β(3) expressed on the surface of several types of solid tumor cells, and they bind to arginine-glycine-aspartic acid (RGD) which enhanced the adhesion to tumor cells. This rout was exploited to construct a tumor-targeting gene RGD-IL-24 which can express RGD-MDA-7/IL-24 protein that includes the cell adhesive sequence (164)Arg-(165)Gly-(166)Asp (A Glycine residue was inserted into the recombinant MDA-7/IL-24 between Arg164 and Asp165 to form a RGD motif). We successfully got the MDA-7/IL-24 mutant by overlapping polymerase chain reaction (PCR) and evaluated its therapeutic efficacy for tumor cell lines MCF-7, HeLa, HepG2, and normal human lung fibroblast (NHLF) line. And we found that the expression of pCDNA3.1/RGD-IL-24 was same to the expression of pCDNA3.1/IL-24. The RGD-IL-24 enhanced the apoptosis-inducing function in tumor cells, but not in normal cells. In tumor cell lines, the apoptosis-inducing activities of RGD-IL-24 was significantly higher than IL-24 detecting by MTT assay, Annexin V, and Hoechst 33258 analysis. Further, pCDNA3.1/RGD-IL-24 showed a significant increase in the ratio of pro-apoptotic (bax) to anti-apoptotic (bcl-2) proteins in tumor cell lines, but not in NHLF cell line. Together, these results suggest that RGD-IL-24 can enhance the apoptosis of tumor cells and may provide a promising drug in tumor therapy.