Abstract

Despite the enormous progress that has been made over the past decades in diagnosis, treatment and prevention of many types of tumors, survival rates in pancreatic cancer still remain poor. Pancreatic cancer is one of the most malignant and chemoresistant tumors and the profound mechanism supporting these phenomena is the constitutively activated prosurvival autophagy. The antitumor 1-nitroacridine derivative C-1748 belongs to a new set of DNA-damaging agents developed in our laboratory and is currently ready for phase I clinical trials. Here, we have uncovered a new role for C-1748 as a potent inhibitor of autophagy in four tested pancreatic cancer cell lines: Panc-1, MiaPaCa-2, BxPC-3 and AsPC-1. C-1748 treatment at 0.1 – 1 μM significantly reduced the typical hallmarks of autophagy, such as conversion of soluble LC3-I protein to its lipid-bound LC3-II isoform or development of acidic vesicular organelles, and importantly shifted pancreatic cancer cells into apoptotic cell death. Co-treatment with C-1748 and wortmannin, had a synergistic effect on autophagy inhibition and potentiated C-1748-induced apoptosis. In particular, compared with exposure to C-1748 alone, the combination treatment increased time- and dose-dependent cytotoxic activity of C-1748 with the most profound effect observed in BxPC-3 cells. Moreover, along with decrease in autophagy, co-treatment with wortmannin significantly elevated dysfunction of mitochondria, as manifested by a drop in mitochondrial membrane potential, increased caspase-3 activation, PARP cleavage, DNA fragmentation into apoptotic bodies and accumulation of lipid droplets arising due to the interruption of autophagy. In all studied cell lines, the level of apoptotic cells after dual exposure with C-1748 and wortmannin increased by about 20% as compared to C-1748 alone. Surprisingly, simultaneous exposure of pancreatic cancer cells to C-1748 in combination with the late-stage autophagy inhibitor, chloroquine, neither significantly increased the cytotoxic activity of C-1748 nor ameliorated its apoptosis-inducing potential. The synergistic effect of C-1748 and wortmannin on the inhibition of autophagy and lack of such effect when C-1748 was co-administered with chlroquine, suggests that C-1748, in addition to its DNA-damaging properties may also affects early stages of autophagy.

Authors (5)