Modified Nucleotides in Bifunctional Antisense Oligonucleotides: Exploring their Anticancer Potential

Bifunctional antisense oligonucleotides (BASOs) are splice-switching molecular tools composed of two, functional parts, that is, antisense part that hybridizes to target pre-messenger RNA fragment and regulatory part that recruits splicing factors to that localization. The aim is to verify the influence of variously modified nucleotides on alternative splicing regulation of the pyruvate kinase M1/2 gene and the anticancer potential of BASOs. The effect of modified BASOs on the phenotype of cancer cells is evaluated by real-time measurement of cell proliferation and motility. The studies reveal chemical modifications that significantly increase the therapeutic properties of BASOs as evidenced by decrease in cell proliferation rate, cancer cell death, and reduced motility. As the most promising modifications, it is considered that β-L-RNA and unlocked nucleic acid, present in BASOs, not only show splicing regulatory properties but also anticancer effects. The studies propose efficient modifications that might be used in protein-binding oligonucleotides to improve their therapeutic efficacy.