Description:
Technology: Researchers at MUSC and NYU Langone Medical Center have identified a synthetic protein (R15) that selectively binds to the nucleotide-free state of the RAS proto-oncogene (apoRAS) and is the first experimental tool to selectively inhibit the signaling and oncogenic activity of selected oncogenic RAS mutants (Figure 1). The synthetic protein is based on Monobody technology. Monobodies are single-domain synthetic protein scaffolds that achieve affinity and selectivity like antibodies, but are refractory to the reducing environment of cells and thus can be utilized as genetically encoded reagents. Their research has shown R15 to specifically inhibit the HRAS(Q61L), KRAS(Q61L), NRAS(Q61L), KRAS(G13D), and KRAS(A146T) RAS oncogenic mutants.
Figure 1 (above): HEK cells were transfected with CFP or the indicated CFP-tagged Mb along with MYC-tagged ERK and further co-transfected with various RAS mutants. As controls for off-target effects, oncogenic MEK(DD) and BRAF(V600E) were also examined. MYC-ERK was then immunoprecipitated and probed for pERK or total ERK. R15m10 selectively inhibits ERK activation by Q61L, G13D and G12D mutants but not G12V, MEK(DD) or BRAF(V600E) indicating selectivity toward mutants with fast exchange rates. Figure 2 (below): Experiments were also performed with different RAS mutants that are present at high frequency in human tumors, and R15m10 was again found to selectively inhibits ERK activation in G12D, Q61H, Q61R and G13D RAS mutants.
Additionally, the MUSC researches have shown that the R15 Monobody can target RAS(G12D) which is an extremely common RAS mutation found in pancreatic cancers. Additional work will be conducted to further characterize the molecular mechanisms underlying the specificity of R15 for RAS proteins, determine any off-target effects of R15 in cells and design strategies to effectively deliver R15 into cells and tissues.
Overview: About 30% of cancers are driven by mutations in RAS genes, including 95% of pancreatic cancers and 45% of colorectal cancers [1]. Pancreatic cancer is estimated to affect 57,000 Americans this year, and this number is a staggering 145,000 for rectal and colon cancers [2,3]. In the US alone, $87.8 billion was spent in 2014 on cancer-related health care [4]. So far, developing ways to block RAS protein function has been unsuccessful, except for a single mutation (G12C). These mutant RAS proteins have been difficult to target because they are defective in intrinsic enzyme activity and accumulate in the GTP-bound state. ApoRAS is regarded as a highly transient state of RAS that cannot be directly targeted with a therapeutic reagent, but MUSC and NYU researchers have shown that certain mutants can be trapped in this state and inhibited by R15. Further advancement of this discovery will lead to novel methods and compositions for the detection, diagnosis, prevention and treatment of diseases or disorders including cancer.
Applications:
Diagnosis – marker for diagnosis/detection, assessing disease severity, and monitoring effectiveness of treatment
Cancer Therapeutics - RAS mutant targeting to suppress cancer proliferation
Drug Delivery - anti-RAS localizing for targeted drug
Advantages:
- inhibits signaling and oncogenicity of currently undruggable RAS mutants
- R15 can be a tool in isolation of a novel anti-RAS drug targeting apoRAS
Key Words: monobody, oncogenic signaling, RAS, nucleotide-free RAS (apoRAS), cancer
Inventors: John P. O’Bryan, Shohei Koide, Akiko Koide
Patent Status: PCT/US2020/038363 published
MUSC-FRD Technology ID: P19125
Available for Licensing!