Abstract 3564: MicroRNA-145 gene therapy for pancreatic cancer using exosome as carrier source derived from cancer cells

Abstract

Abstract Background-Pancreatic cancer (PanCa) is the third deadliest cancer in United States with a poor survival rate. Despite extensive research efforts, there is no substantial progress in cancer therapeutics due to barrier against intracellular drug delivery. Our group has previously identified that microRNA-145 (miR-145) is downregulated in PanCa, the restoration of which inhibits tumor growth and enhances gemcitabine sensitivity. However, lack of an effective tumor-specific delivery system remains an unmet clinical challenge for successful translation of microRNAs. Herein, we have utilized exosomes (extra cellular vesicles) isolated from pancreatic cancer cells as a vehicle for delivering miR-145 in PanCa cells. Exosomes, unlike other vectors for gene delivery, are non-immunogenic in nature and can protect the RNA/gene of interest from digestion making them a more efficient vehicle for gene delivery. Method- Exosomes were isolated from pancreatic cancer cells HPAF-II, AsPC-1 by the principle of precipitation using exosome isolation reagents. Physico-chemical characterization (DLS, TEM), presence of exosomal marker (CD63; immunoblotting), protein concentration (Bradford assay) and cellular internalization (confocal microscopy) of the exosomes were performed. miR-145 transfection was performed in pancreatic cancer cells using exofectamine. miR-145 restitution and anti-cancer efficacy was investigated using in vitro functional assays for cell viability (MTT), migration (Boyden chambers), invasion (Matrigel), colonogenicity and tumor spheroid formation. The effect of exosome mediated miR-145 restitution was investigated using Western blotting and PCR analysis. Results- Exosomes show effective size and zeta potential (AsPC-1:164 nm -9.4mV respectively; HPAF-II: 157 nm -9.2 mV), which is ideal for drug delivery purposes. The purification of exosomes was confirmed by analyzing the expression of exosomal markers, such as CD63. Immunofluorescence for CD63 expression confirmed the efficient delivery of exosomes in PanCa cells. miR-145 loaded exosome treatment efficiently delivered miR-145 into the cancer cells and inhibited the tumorigenic features such as, proliferation, migration, invasion and colonogenicity of PanCa cells. The restoration of lost miR-145 levels using exosome mediated delivery was confirmed in PanCa cells using PCR analysis. MicroRNA-145 fold expression was found to be high at both pH, 7.4 and 6.8, which further indicates relevance for their utilization in the delivery of therapeutic modalities. Conclusion- Our observations offer importance of the utilization of exosomes for delivery of therapeutic modalities and developing personalized medicine in PanCa. Citation Format: Kamalika Samanta, Sheema Khan, Saini Setua, Sonam Kumari, Nirnoy Dan, Murali Mohan Yallapu, Meena Jaggi, Subhash Chandra Chauhan. MicroRNA-145 gene therapy for pancreatic cancer using exosome as carrier source derived from cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3564.