Given the important role of protein kinases in pathways affecting cellular growth and invasion, we have analyzed 340 serine/threonine kinases for genetic mutations in colorectal cancers. Mutations in eight genes were identified, including three members of the phosphatidylinositol-3-kinase (PI3K) pathway; the alterations in the latter genes each occurred in different tumors and did not overlap with mutations in PIK3CA or other non-serine-threonine kinase (STK) members of the PI3K pathway, suggesting that mutations in any of these genes had equivalent tumorigenic effects. These data demonstrate that the PI3K pathway is a major target for mutational activation in colorectal cancers and provide new opportunities for therapeutic intervention.