RModBlock antisense oligonucleotides as a universal tool for precise and efficient inhibition of RNA modifications

Area of Science:

• Molecular Biology
• Epigenetics
• RNA Biology

Background:

• Understanding RNA modifications is crucial for cellular function and disease research.
• Current methods, like dCas-based systems, have limitations in applicability and performance depending on modification type and context.
• Targeted inhibition of RNA modification writers is needed for precise manipulation.

Purpose of the Study:

• To introduce and validate the RNA Modification-Blocking (RModBlock) strategy for precise inhibition of RNA modifications.
• To assess the efficacy of RModBlock ASOs across different modification types (m5C, pseudouridine, m6A).
• To evaluate the therapeutic potential of RModBlock strategy in cellular and in vivo models.

Main Methods:

• Development of chemically modified antisense oligonucleotides (ASOs) incorporating locked nucleic acid for targeted RNA modification inhibition.
• Application of RModBlock ASOs to block structural contexts required by RNA modification writers.
• Testing RModBlock efficacy on m5C, pseudouridine, and m6A modifications in human cells.
• In vivo delivery of RModBlock ASOs to mouse liver.

Main Results:

• RModBlock ASOs effectively inhibited m5C and pseudouridine formation by up to 97% by blocking their required structural contexts.
• The strategy also successfully inhibited m6A modification, demonstrating broad applicability.
• RModBlock performance was comparable or superior to existing dCas13-eraser systems.
• Inhibition of cancer-relevant modifications and successful in vivo delivery highlighted therapeutic potential.

Conclusions:

• The RModBlock strategy offers a precise, efficient, and versatile approach for manipulating RNA modifications.
• This method has broad applicability in both basic science and translational research.
• RModBlock holds promise as a therapeutic tool for diseases associated with aberrant RNA modifications.