Probing RAS Function Using Monobody and NanoBiT Technologies.

Area of Science:

• Oncology
• Molecular Biology
• Drug Discovery

Background:

• Missense mutations in RAS oncogenes (HRAS, KRAS, NRAS) are implicated in ~20% of human cancers, presenting a significant therapeutic target.
• Recent successes with KRAS^G12C inhibitors and ongoing trials for KRAS^G12D inhibitors have challenged the notion of RAS being undruggable.
• However, numerous RAS mutants driving cancer remain without effective therapeutic strategies, necessitating further research into RAS biology and novel drug development.

Purpose of the Study:

• To develop novel protein-based inhibitors targeting specific RAS isoforms and mutants using monobody (Mb) technology.
• To create tools for analyzing RAS protein-protein interactions and screening for RAS-binding small molecules.
• To identify new therapeutic vulnerabilities for the development of anti-RAS drugs.

Main Methods:

• Utilized monobody (Mb) technology to engineer specific protein inhibitors for RAS oncogenes.
• Combined monobodies with NanoLuc Binary Technology (NanoBiT) for live-cell assays.
• Employed high-throughput screening assays to analyze RAS protein-protein interactions and identify small molecule binders.

Main Results:

• Demonstrated the successful application of monobodies for targeting specific RAS mutants.
• Established a novel NanoBiT-based system for live-cell analysis of RAS interactions.
• Developed a high-throughput screening platform for identifying potential anti-RAS therapeutics.

Conclusions:

• Monobody technology offers a promising avenue for developing targeted anti-RAS therapeutics against challenging cancer mutations.
• The combined Mb and NanoBiT approach provides a powerful tool for dissecting RAS biology and accelerating drug discovery.
• This work contributes to overcoming the limitations of current RAS-targeted therapies and addresses the need for drugs against previously undruggable RAS mutants.