Protomer selectivity of type II RAF inhibitors within the RAS/RAF complex.

Area of Science:

• Oncology
• Molecular Biology
• Pharmacology

Background:

• RAF- and RAS-driven cancers represent significant therapeutic challenges.
• RAF dimer inhibitors hold promise by targeting the RAS-RAF signaling complex.
• Effective drug utility depends on simultaneous binding to both RAF protomers.

Purpose of the Study:

• To develop and validate a method for quantifying drug-target occupancy at specific RAF protomers within the active RAS-RAF complex in cellular environments.
• To assess RAF target engagement irrespective of KRAS mutation status.
• To evaluate the cellular milieu's impact on the high-affinity state of RAF dimer inhibitors.

Main Methods:

• Development of a conditional quantification method for drug-target occupancy at RAF protomers.
• Measurement of RAF target engagement in the presence and absence of mutant KRAS alleles.
• Assessment of intracellular protomer selectivity for clinical-stage type II RAF inhibitors.

Main Results:

• A novel method allows quantification of drug-target engagement at RAF protomers within the RAS-RAF complex.
• Clinical-stage type II RAF inhibitors demonstrate selectivity for ARAF protomer engagement over BRAF or CRAF.
• ARAF protomer engagement, unlike BRAF or CRAF, correlates with MAPK signaling inhibition across various mutant RAS cell lines.

Conclusions:

• ARAF plays a critical role in mutant RAS-driven signaling pathways.
• The study reveals limited ARAF protomer vulnerability for several RAF inhibitors currently in clinical trials.
• Targeting ARAF may be a key strategy for effective RAF inhibitor therapy in specific cancers.