Characterisation of a cyclic peptide that binds to the RAS binding domain of phosphoinositide 3-kinase p110α
Mohamed Ismail1, Stephen R Martin2, Roger George2
1Oncogene Biology Laboratory, Francis Crick Institute, 1 Midland Road, London, NW1 1AT, UK.
Abstract:
P110α is a member of the phosphoinositide 3-kinase (PI3K) enzyme family that functions downstream of RAS. RAS proteins contribute to the activation of p110α by interacting directly with its RAS binding domain (RBD), resulting in the promotion of many cellular functions such as cell growth, proliferation and survival. Previous work from our lab has highlighted the importance of the p110α/RAS interaction in tumour initiation and growth. Here we report the discovery and characterisation of a cyclic peptide inhibitor (cyclo-CRVLIR) that interacts with the p110α-RBD and blocks its interaction with KRAS. cyclo-CRVLIR was discovered by screening a "split-intein cyclisation of peptides and proteins" (SICLOPPS) cyclic peptide library. The primary cyclic peptide hit from the screen initially showed a weak affinity for the p110α-RBD (Kd about 360 µM). However, two rounds of amino acid substitution led to cyclo-CRVLIR, with an improved affinity for p110α-RBD in the low µM (Kd 3 µM). We show that cyclo-CRVLIR binds selectively to the p110α-RBD but not to KRAS or the structurally-related RAF-RBD. Further, using biophysical, biochemical and cellular assays, we show that cyclo-CRVLIR effectively blocks the p110α/KRAS interaction in a dose dependent manner and reduces phospho-AKT levels in several oncogenic KRAS cell lines.
Insights
Researchers developed cyclo-CRVLIR, a cyclic peptide inhibitor, to block the interaction between phosphoinositide 3-kinase alpha (PI3Kα) and KRAS. This discovery offers a potential new strategy for targeting cancer growth driven by PI3Kα/RAS signaling.
Area of Science:
- Biochemistry
- Molecular Biology
- Cancer Research
Background:
- P110α, a phosphoinositide 3-kinase (PI3K), is activated by RAS proteins, promoting cell growth and survival.
- The PI3Kα/RAS interaction is crucial for tumor initiation and progression.
- Targeting this interaction presents a therapeutic opportunity in oncology.
Purpose of the Study:
- To discover and characterize a novel inhibitor of the PI3Kα/RAS interaction.
- To develop a peptide-based therapeutic agent targeting oncogenic signaling pathways.
Main Methods:
- Screening of a cyclic peptide library (SICLOPPS) to identify inhibitors of PI3Kα.
- Affinity determination using biophysical assays (e.g., Kd measurements).
- Biochemical and cellular assays to assess inhibitory activity and downstream effects on signaling pathways (e.g., phospho-AKT levels).
Main Results:
- Discovery of cyclo-CRVLIR, a cyclic peptide inhibitor targeting the PI3Kα-RAS binding domain (RBD).
- Optimized cyclo-CRVLIR exhibits low µM affinity for PI3Kα-RBD, with selective binding over KRAS and RAF-RBD.
- Cyclo-CRVLIR effectively inhibits the PI3Kα/KRAS interaction and reduces phospho-AKT levels in KRAS-driven cancer cell lines.
Conclusions:
- Cyclo-CRVLIR is a potent and selective inhibitor of the PI3Kα/KRAS interaction.
- This cyclic peptide represents a promising lead compound for developing novel cancer therapeutics.
- Targeting the PI3Kα-RBD offers a viable strategy to disrupt oncogenic RAS signaling.
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