The p110? isoform of PI3 kinase (PI3K?) has been implicated in pathological disorders such as thrombosis and cancer and a number of PI3K?-selective inhibitors have recently progressed into clinical studies. Although crystallography studies identify a binding site conformation favored by the inhibitors, no specific interaction explains the observed selectivity. Using site-directed mutagenesis we have identified a specific tyrosine residue of the binding site Y778 that dictates the ability of the PI3K? isoform to bind these inhibitors. When mutated to isoleucine, PI3K? has reduced ability to present a specific cryptic binding site into which a range of reported PI3K? inhibitors can bind, and conversely when tyrosine is introduced into the same position in PI3Ka, the same inhibitors gain potency. The results provide a cogent explanation for the selectivity profiles displayed by these PI3K inhibitors and maybe others as well.