Precision cancer therapy through nanoparticle delivery of siRNA against KRAS.

Abstract

260 Background: Small interfering RNA (siRNA) has potential for highly specific gene manipulation, making it attractive for delivering precision therapy to cancer patients. However, efforts to employ siRNA therapeutically have been limited by its short half-life in circulation, low target tissue specificity, and cellular entrapment within endosomes. We utilized serum-stable, cell-penetrating, and endosomolytic peptide-based nanoparticles (NPs) to overcome these obstacles and deliver siRNA against KRAS to KRAS-mutant human and mouse pancreas and colorectal cancers. Methods: Human and mouse pancreas and colorectal cancer cell lines were tested for NP uptake in vitro utilizing fluorescent siRNAs. Uptake was assessed via fluorescent microscopy and flow cytometry (FC). Mice bearing tumors from these cells were injected IV with the same NP, and uptake was assessed with an in vivo imaging system (IVIS), and FC. Cell lines were treated with KRAS-siRNA NP and KRAS knockdown was assessed by real-time PCR. Results: Mouse and human pancreas and colorectal cancer cell lines took up NP in vitro, with signal detected within > 93% of cells at 24 hours. Tumors from these cells grown in mice were strongly fluorescent after IV injection of fluorescent NP within 2 hours, and until at least 30 hours. FC of a tumor treated with fluorescent NP showed that 86% of tumor cells expressed fluorescent signal 24 hours post-injection. IVIS revealed signal in mouse liver and kidneys, but when assessed by FC, only 17.8% and 13.5% of cells from these tissues were fluorescent, respectively. The brain, heart, lungs, spleen, and pancreas of mice receiving injections were negative. Cancer cell lines exposed to KRAS-siRNA NP for 48 hours express KRAS at levels that are 4.5 to 15.1% of untreated cells. Conclusions: Human and mouse pancreas and colorectal cancers efficiently and specifically take up NP in vitro and in vivo. Selected limitations of siRNA are overcome with this NP delivery system, and NP-packaged siRNA effectively inhibits KRAS. This platform represents a highly specific approach to targeting tumor genes of interest, which may ultimately enable selective knockdown of putative drivers of tumor progression.