Abstract 3797: C6Ceramide as potential novel therapy for pancreatic cancer

Abstract

Abstract Pancreatic cancer is a highly aggressive chemo-resistant cancer with a five-year survival rate, emphasizing need for more effective therapies. Ceramide, an endogenous membrane lipid, regulates cell survival and induces apoptosis in cancer cell lines. Our previous studies demonstrated the ability of C6-ceramide to enhance the cytotoxicity of the chemotherapy agents Gemcitabine (Gem), Paclitaxel (Pac), and Cetuximab (Cetux) in pancreatic cancer lines, both in vitro and in vivo. The goal of this study was to further refine this combinatorial effect, as well as to test the efficacy of liposomes as potential drug delivery systems for C6-ceramide. These studies were conducted using three well-defined pancreatic cancer cell lines: PANC-1, MIA-PaCa-2, and L3.6. In vitro cytotoxicity was measured using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) cell viability assay. C6-Ceramide dose-response studies (delivered in DMSO) showed that PANC-1 was most susceptible to cytotoxicity followed by MIA and L3.6. Interestingly for all three cell lines, the potency of C6-ceramide was dependent on cell density in the well. Cell densities of 7K, 15K, 32K, and 64K cells/well were examined. C6-ceramide exhibited greatest potency at 7K and least potency at 32K cells/well. This effect of cell density was not observed to the same extent in dose-response to Gem, Pac, or Cetux alone. The second component of the study observed the effects of C6-ceramide in combination treatment with each of the three chemo agents. The results again demonstrated that cytotoxicity was dependent upon cell density when administering C6-ceramide in the combinations. In vitro studies were carried out to examine a pegylated liposome preparation for C6-ceramide delivery. These were 5:95 mole ratio 18:0 PEG2 PE:DOPC (“empty” liposomes) and 5:88:7 mole ratio 18:0 PEG2 PE:DOPC:C6-Ceramide (C6-ceramide/liposome complex). Empty liposomes had no effect on cell viability. However, in contrast to delivery in DMSO, C6-ceramide/liposomes did not demonstrate a strong cytotoxic effect, suggesting that C6-ceramide may be too tightly bound to the liposomes. The effect of the liposomal preparations on the cytotoxic response of Gem, Pac, and Cetux was examined in vitro in L3.6 cells. Increasing concentrations of empty liposomes tended to reduce the effect of the chemo agent, perhaps due to complexation of the agent. However, slightly more cytotoxicity was retained with the C6-ceramide/liposomes in presence of the chemo agent. A pilot study was carried out in SCID mice implanted with L3.6 tumors to examine the effect of C6-ceramide/liposomes on Gem therapy. Three groups were compared: Control (vehicle only), Gem alone, and Gem + C6-ceramide/liposomes. While Gem decreased tumor size and increased survival, there was no improvement with co-administration of the C6-ceramide/liposome preparation. This was consistent with the in vitro results. Other delivery systems for exogenous C6-ceramide will be tested. Citation Format: Apurva Limaye, Edith Mathiowitz, Wayne D. Bowen, Harold Wanebo. C6Ceramide as potential novel therapy for pancreatic cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3797. doi:10.1158/1538-7445.AM2015-3797