Abstract 3625: Nanodelivery platform for targeting mutant-KRAS and improving response to gemcitabine therapy

Abstract

Abstract Background: Although, surgical resection and chemotherapy are the gold standard for treating pancreatic cancer (PanCa), poor patient survival remains the problem. Despite being one of the most common oncogenes in human cancer, to date, no success has been achieved to inhibit KRAS. Targeting KRAS has been shown to synergize anti-cancer activity of gemcitabine in pancreatic cancer. Herein, we have developed a supermagnetic iron oxide (SP) nanoparticles for the sustained delivery of KRAS siRNA to the tumor and simultaneous sensitization of gemcitabine in PanCa. Methods: PanCa cells, AsPC1 and Panc-1 were used in the study. A precipitation approach was employed to develop the SP formulation. These particles were further conjugated with siKRAS (12D; most common KRAS mutation) and investigated for its anticancer efficacy alone and in combination with gemcitabine. Particles were investigated for size, physico-chemical characterization (Dynamic light scattering), hemocompatiblity (hemolysis assay) and the complexation of siKRAS (gel retardation assay). Cellular internalization and uptake of the particles were investigated using FAM labelled siRNA and Prussian blue assay. kRAS silencing was confirmed at both mRNA and protein levels using quantitative reverse-transcription PCR and Western blotting, respectively. Anti-cancer efficacy of SP-siKRAS particles alone or in combination with gemcitabine treatment was determined using in vitro functional assays for cell viability (MTT), migration (Boyden chambers), invasion (Matrigel), colonogenicity and tumor spheroid formation. Results: Our results demonstrate optimal particle size (190.137 nm) and zeta potential (18.73mV) of SP-siKRAS formulation. SP-siKRAS efficiently internalized in PanCa cells and suppressed KRAS G12D expression as well as its downstream targets, YAP and PDL-1. SP-siKRAS improved gemcitabine response as observed through enhanced inhibition of cell proliferation, clonogenicity, migration, and invasion of pancreatic cancer cells. Additionally, SP-siKRAS together with gemcitabine resulted in the activation of death related mechanisms in PanCa cells, such as Bax, bcl-2, PARP cleavage. Interestingly, SP-siKRAS inhibited the secondary tumorsphere formation in combined PanCa and cancer associated fibroblast (CAFs) cells, alone as well as in combination with gemcitabine. At day 14, analysis of secondary tumorspheres, treated with SP-siKRAS revealed diminished levels of KRAS G12D through PCR. This further provides a clinical validation demonstrating potential of SP-siKRAS particles to efficiently silence KRAS expression. SP-siKRAS also exhibited haemocompatibility, suggesting its potential of silencing KRAS without being toxic to the body. Conclusion: Therefore, SP-siKRAS provide a highly efficient and safe platform for silencing KRAS and improving gemcitabine therapy in pancreatic cancer. Citation Format: Nirnoy Dan, Sheema Khan, Saini Setua, Sonam Kumari, Pallabita Chowdhury, Kamalika Samanta, Meena Jaggi, Murali Yallappu, Subhash Chauhan. Nanodelivery platform for targeting mutant-KRAS and improving response to gemcitabine therapy [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 3625.