115 Endosomolytic Peptide-Mediated Delivery of Therapeutic siRNAs into Oral Cancer Cells

Thursday, March 22, 2012: 8 a.m. - 9:30 a.m.
Presentation Type: Oral Session
M. SOKOLOSKY, and A. JAKYMIW, Dept. of Craniofacial Biology, Medical University of South Carolina, Charleston, SC
An obstacle for targeted small interfering RNA (siRNA)-mediated therapies has been the retention of siRNA cargo in endosomes and their subsequent degradation in lysosomes, resulting in inefficient gene silencing. Objective: Examine the feasibility of utilizing a peptide with endosome-disruptive properties to noncovalently deliver siRNAs into oral cancer cells and induce silencing of the targeted oncogene, CIP2A. Methods: An endosome-disruptive peptide, 599, was designed and chemically synthesized consisting of a derivative sequence of the amino-terminal sequence of influenza virus hemagglutinin and a stretch of densely packed cationic amino acid residues which would enable siRNAs to be delivered into cells via electrostatic interactions. Agarose gel shift assays were then used to examine the ability of the 599 peptide to interact efficiently with siRNAs. Subsequently, indirect immunofluorescent and qRT-PCR analyses were performed to measure the ability of the 599 peptide to internalize siRNAs and induce gene silencing of CIP2A in CAL 27 oral cancer cells. Results: The 599 peptide was demonstrated to bind siRNA molecules at a peptide to siRNA molar ratio of 20:1, but not 1:1. Upon examining the cellular localization of this peptide premixed with fluorescently-labeled siRNAs in oral cancer cells, the 599 peptide was found to deliver the siRNA molecules into the cytoplasm of cells with increasing peptide concentrations after either a 2 or 48 hour incubation. Conversely, control peptides containing either the endosome-disruptive or cationic residue portions alone had no apparent effect on siRNA delivery. Moreover, 599-mediated siRNA delivery promoted silencing of CIP2A in oral cancer cells after a 48 hour incubation. Conclusion: Together, these data suggest that peptides with endosome-disruptive properties in combination with dense cationic residues can effectively deliver siRNAs into cells, induce the silencing of their target genes, and potentially offer new therapeutic avenues in combating oral cancer.
This abstract is based on research that was funded entirely or partially by an outside source: NIDCR, R00DE018191 (AJ); Bankhead-Coley Florida Cancer Research Program, 08BN-02 (AJ); NCRR, P20RR017696

Keywords: Carcinogenesis, Delivery systems, RNAi and Therapeutics