Enhanced gene silencing by the application of multiple specific small interfering RNAs

Small interfering RNA duplexes (siRNA) induce gene silencing in various eukaryotic cells, although usually in an incomplete manner. Using chemically synthesized siRNAs targeting the HIV-1 co-receptor CXCR4 or the apoptosis-inducing Fas-ligand (FasL), co-transfection of cells with two or more siRNA duplexes targeting different sites on the same mRNA resulted in an enhanced gene silencing compared with each single siRNA. This was shown in the down-regulation of protein and mRNA expression, and functionally in the inhibition of CXCR4-mediated HIV infection and of FasL-mediated cell apoptosis. Transfection efficiency determined for the FasL-specific siRNAs was dose-dependent and varied among the siRNAs tested, but was not the main reason for the enhanced gene silencing.