PI3Kδ is selectively inhibited by roginolisib by stabilizing the C-terminal helix kα12.

Phosphoinositide 3-kinases (PI3Ks) are major regulators of cell growth, proliferation and signalling, constituting key therapeutic targets in cancer, inflammation, and other diseases. Individual class I PI3K isoforms control key cellular functions, imposing the need to generate isoform-specific inhibition for therapeutic intervention. Roginolisib is a selective PI3Kδ inhibitor that shows promise for the treatment of cancer. Using a combination of X-ray crystallography, molecular dynamics simulations, and hydrogen-deuterium exchange mass spectrometry, we describe the mechanism driving roginolisib's potent and isoform-selective inhibition of PI3Kδ. Roginolisib uniquely stabilises the catalytic C-terminal helix kα12, locking the enzyme in an inactive conformation. This binding mode also results in more sustained inhibition of phosphatidylinositol 3,4,5-trisphosphate formation in tumour samples of CLL patients. Inhibition of PI3Ks by stabilization of helix kα12 into an inactive conformation has not, to the best of our knowledge, been described before and may provide the basis for novel, more selective and effective pharmacological strategies.