Cancer heterogeneity and microenvironmental aspects within a tumor are considered key factors influencing resistance of carcinoma cells to distinct chemotherapeutical agents. We evaluated a high concentration of metformin in combination with gemcitabine on a syngeneic orthotopic mouse model using 6606PDA cells. We observed reduced tumor size and reduced cancer cell proliferation after three weeks of chemotherapy with either compound and noticed an additive effect between gemcitabine and metformin on tumor weight. Interestingly, distinct areas of the carcinoma responded differently to either compound. Metformin inhibited the proliferation of cancer cells close to the desmoplastic reaction, whereas gemcitabine inhibited the proliferation of cancer cells mainly 360–570 μm distant to the desmoplastic reaction. Indeed, co-culture of pancreatic stellate cells with 6606PDA, 7265PDA or MIA PaCa-2 cells increased gemcitabine resistance. Metformin resistance, however, was increased by high glucose concentration in the medium. Other factors such as hypoxia or the pH of the medium had no influence on gemcitabine or metformin induced inhibition of cancer cell proliferation. These data demonstrate a spatial heterogeneity in drug resistance within pancreatic adenocarcinomas and that microenvironmental aspects such as supply of glucose and the presence of pancreatic stellate cells regulate cancer cell sensitivity towards metformin or gemcitabine.

Many preclinical and clinical studies are currently evaluating metformin in combination with classical therapeutic agents as anti-cancer therapy. In this study we used three distinct pancreatic cancer cell lines and evaluated cell death by trypan blue assay and Western Blots using antibodies directed against cleaved caspase 3 and PARP. Surprisingly, we observed that 20mM metformin did not enhance, but rather inhibited gemcitabine induced cell death in murine 7265PDA, 6606PDA and 6606l cells. Microenvironmental aspects such as oxygen supply or the pH value did not influence the inhibition of cancer cell apoptosis by metformin. Glucose concentration in the medium, however, had a major effect on the impact of metformin. Medium with 0.5g/L glucose strongly increased metformin induced apoptosis and also prevented the inhibitory effect of metformin on gemcitabine induced cell apoptosis, when compared with medium containing 4.5g/L glucose. We conclude that the combination of metformin with gemcitabine has inappropriate effects for a successful treatment of pancreatic cancer. Thus, it might be more promising to use metformin in combination with other drugs that reduce the uptake or the metabolism of glucose.

BackgroundThe involvement of Wnt in carcinogenesis and progression of pancreatic cancer is currently intensely discussed. We evaluated activation of the Wnt signaling pathway by using a Wnt reporter mouse strain expressing β-galactosidase under the control of the Axin2 promotor during pancreatitis induced formation of precancerous lesions. We also evaluated activation of Wnt signaling during interaction of pancreatic cancer with the tumor stroma.ResultsActivation of Wnt signaling was observed during acinar-to-ductal metaplasia after chronic as well as acute pancreatitis. Activation of Wnt signaling was also noticed during growth of pancreatic cancer in an orthotopic syngeneic pancreas cancer model. Activation of Wnt signaling was, however, not observed in carcinoma associated fibroblasts, but was detected in few cell clusters inside the tumor. Genetic ablation of Axin2 significantly reduced body weight without having a major impact on blood glucose concentration. However, ablation of Axin2 had no influence on the observed β-galactosidase positive cell clusters or on tumor weight.ConclusionThese data demonstrate that the Wnt signaling pathway is activated during acinar-to-ductal metaplasia after injury to the pancreas. However these data do not support a major role of Wnt signaling or of Axin2 in carcinoma associated fibroblasts and tumor growth.Electronic supplementary materialThe online version of this article (doi:10.1186/s13578-016-0116-4) contains supplementary material, which is available to authorized users.

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