AIM Gallic acid (GA) and methyl gallate (MG) derived from grape seeds possess growth-inhibiting and apoptosisinducing activities in cancer cell lines in vitro.In the present study, we have investigated the impact of defined combinations of GA and MG (ratio 1∶ 1,2∶ 1, 3∶ 1, 4∶1 and 5∶1) on cell proliferation and apoptosis induction in cultured A.549 lung carcinoma cells and then investigated the possible molecular mechanisms underlying the observed toxic effects.METHODS Human lung adenocarcinoma A549 cells were treated with a combination of GA and MG in different dose ratio, and the changes in cell viability and apoptosis were determined by MTT assay and Annexin V-FITC staining assay, respectively.Apoptosis-related proteins were determined by a Western blot analysis.The malate dehydrogenase, voltage-dependent anion-selective channel protein 1 (VDAC1), calreticulin and brain acid soluble protein 1 were measured by two-dimensional gel electrophoresis.VDAC1 siRNA and negative control siRNA Were transfected by using lipofectamine-2000 into cell lines according to the manufacture.RESULTS The percent viability of cells decreased with GA and MG treatment in a dose dependent manner.GA and MG exhibit strong antagonistic interaction on the inhibit proliferation of A549 cells and induction of apoptosis in a dose ratio dependent manner.Western blot analysis also showed that caspase 9 expression decreased and Bcl-2 expression increased in a dose ratio dependent manner.Biomarkers were identified using by two dimension electrophoresis, four biomarkers were up-regulated proteins, namely Malate dehydrogenase, VDAC1, calreticulin and brain acid soluble protein 1, respectively.Two of these proteins, VDAC1 and calreticulin are associated with apoptosis.GA up-regulated VDACI expression in a dose dependent manner and combination of GA and MG down-regulated in a dose ratio dependent manner.Transfection with VDAC1 siRNA slightly increased the cell viability after treatment of GA.MEK and p38 inhibitors did not affect the growth inhibition of GA or MG-treated A549 cells, JNK inhibitor also did not affect the growth inhibition of MG alone treated-A549 cells, but siguificanfly intensified the growth inhibition in GA treated-A549 cells.MEK, JNK and p38 inhibitors also did not affect the growth inhibition of two different combination ratios of GA and MG-treated A549 cells.CONCLUSION GA and MG exhibit strong antagonistic interaction on the inhibit proliferation of A549 cells and induction of apoptosis in a dose ratio dependent manner.The mechanisms were likely due to regulating of VDAC1 protein expression.MAPK signaling may be was not tightly related to cell growth inhibition in GA and/or MG treated-A549 lung cancer cells.