Abstract 30P
Background
Gene silencing by small interfering RNA (siRNA) is a promising therapeutic approach for a wide range of disorders, including cancer. siRNAs against BCR-ABL can be a supportive or alternative measure to traditional chronic myeloid leukemia (CML) tyrosine kinase inhibitors (TKIs) therapies, especially given frequently-noted clinical TKI resistance. The main challenge for such approaches remains the development of effective RNAi systems for intracellular delivery. Cell-penetrating peptides (CPPs) are oligopeptides which have the ability to deliver various cargo molecules. They represent a promising approach of achieving siRNA internalization. Despite the active research mechanisms through which CPPs are internalized remain unclear. It has already been shown that CPPs and their cargoes can be taken up by cells via single or multiple endocytic pathways. Precise siRNA delivery mechanism seems to depend on experimental conditions. In present study we investigated cellular uptake efficiency, internalization mechanism and gene-silencing efficiency of a non-covalent nanocomplex consisting of CPP peptide EB1 and siRNA directed against the BCR-ABL oncogene in the K562 human CML cell line.
Methods
The transfection efficiency of the investigated peptide EB1-BCR-ABL siRNA complexes was measured by flow cytometry analysis. In order to study the complex’s internalization mechanisms, transfection was carried out at 4 °C and was analyzed by flow cytometry and confocal microscopy. RNAi modulatory effects were investigated on both mRNA and protein levels of the BCR-ABL using RT-PCR and Western blotting.
Results
We have shown that the transfection efficiency of the investigated complexes was greater than 90%. It was demonstrated that complexes effectively deliver siRNA into K562 cells by endocytic mechanisms. Thereby, transfection of K562 cells by the peptide complexes significantly reduced the levels of BCR-ABL mRNA, with a maximum at 72 h post treatment. Similar decreases in BCR-ABL protein were detected.
Conclusions
Obtained data indicates that CCP-based delivery of siRNAs enables an effective antisense suppression of certain oncogene and represents a promising new class of therapies for CML.
Clinical trial identification
Editorial acknowledgement
Legal entity responsible for the study
The authors.
Funding
Has not received any funding.
Disclosure
All authors have declared no conflicts of interest.