Despite continuing advances in the identification of new targets for cancer therapy, many approaches show modest clinical benefits, induce a response only in a portion of the patients, and/or become inefficient after patients develop drug resistance. New approaches are therefore needed to overcome these effects. There is tremendous optimism regarding the potential of RNA-based therapies where oncogenes could be suppressed by small non-coding RNAs (specifically, siRNAs and miRNAs). Many challenges, including delivery and toxicity, are being solved however the problem of off-target effects persist. One option to avoid this issue is to combine these small RNAs at low concentrations, ensuring that genuine targets are inhibited and that unintended effects on other genes are minimal and widely distributed.
Here we select small RNAs to target simultaneously the main components of the RAS-MAPK pathway, a cell signalling cascade that governs cell survival and is often dysregulated in cancer through genetic alteration. To evaluate the small RNAs efficiency to inhibit their target expression, RT-qPCR and western-blot were used for measuring RNA and protein levels of the RAS-MAPK effectors, respectively. The RAS-MAPK pathway activity was measured through the quantification of phospho-ERK level using an automated western-blot-like system and live-cell imaging was used to assess cell survival.
We show that combination of these small RNAs at sub-nanomolar level efficiently reduces the activity of the RAS-MAPK pathway and significantly decrease cell survival in cancer cells resistant to front-line therapy. Future work aims to confirm the reduction of off-targets effects and verify the efficiency of this strategy on tumour growth in vivo.
This work will lead to the validation of multiplexing small RNAs as a potential therapeutic strategy with high specificity and less toxicity to treat drug-resistant cancer cells.