A non-covalent peptide-based strategy for siRNA delivery.
The development of short-interfering RNA (siRNA) has provided great hope for therapeutic targeting of specific genes responsible for pathological disorders. However, the poor cellular uptake of siRNA together with the low permeability of the cell membrane to negatively charged molecules, remain major obstacles to clinical development. So far there is no universal method for siRNA delivery as they all present several limitations. Several non-viral strategies have been proposed to improve the delivery of synthetic siRNAs in both cultured cells and in vivo. Cell-penetrating peptides (CPPs) or protein transduction domains (PTD) constitute very promising tools for non-invasive cellular import of siRNA and non-covalent CPP/PTD-based strategies have been successfully applied for ex vivo and in vivo delivery of therapeutic siRNA molecules. We recently described a new peptide-based system, CADY, for efficient delivery of siRNA in both primary and suspension cell lines. CADY is a secondary amphiphatic peptide able to form stable non-covalent complexes with siRNA and to improve their cellular uptake independently of the endosomal pathway. This chapter describes easy to handle protocols for the use of the CADY-nanoparticle technology for the delivery of siRNA into both adherent and suspension cell lines. It will also highlight different critical points in the peptide/siRNA complex preparation and transfection protocols, in order to obtain siRNA-associated interfering response at low nanomolar concentration.
Department of Molecular Biophysics and Therapeutics, Centre de Recherches de Biochimie Macromoléculaire, Montpellier, France.
Laurence
L
Crombez
Gilles
G
Divita
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002448:Cell Adhesion; D002460:Cell Line; D057846:Cell-Penetrating Peptides / Q000737:chemistry / Q000378:metabolism*; D004337:Drug Carriers / Q000737:chemistry / Q000378:metabolism*; D020868:Gene Silencing; D006801:Humans; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism*; D014162:Transfection / Q000379:methods*
D057846:Cell-Penetrating Peptides; D004337:Drug Carriers; D012333:RNA, Messenger; D034741:RNA, Small Interfering