From intracellular antibody fragments to small molecule inhibitors of mutant KRAS
Session type: Proffered paper sessions
Theme: Diagnosis and therapy
Mutation in RAS proteins is among the most frequent in human cancer. We have previously selected an intracellular antibody single domain fragment that binds to mutant RAS and blocks RAS-effector interaction-dependent tumour initiation and overt tumour growth in mouse models.
The antibody fragment binds to GTP-bound RAS with high pM affinity and we have used this property to isolate compounds from a fragment library using a competitive SPR method that places the compounds in the region of the binding site of the antibody fragment and crucially depends on the high Kd of the antibody interaction with RAS.
Using a combination of X-ray crystallography and medicinal chemistry, we have obtained compounds that bind adjacent to the KRAS switch I region with nM affinity. These compounds interfere with RAS-RAF, RAS-RALGDS and RAS-PI3K protein-protein interactions in cell-based BRET2 assays as well as inhibiting the downstream phosphorylation of AKT and ERK that results from endogenous RAS signalling. The chemical evolution and interaction characteristics of the KRAS-binding compounds will be described and their biochemical and cell-based properties presented.
Our aim was to develop methods to employ intracellular antibody fragments as lead macromolecules for small compound selection. We have used this approach to identify compounds binding to mutant RAS with the potential to inhibit RAS-effector interactions.
We have validated our approach using an antibody fragment as a screening tool for the identification of small molecule inhibitors of RAS-effector interaction. This approach has yielded compounds that are currently being developed as potential RAS-effector inhibitors in our medicinal chemistry programme.