The cyclin-dependent kinases (CDKs) are critical regulators of cell cycle progression, and are involved in uncontrolled cell proliferation-a hallmark of cancer. This suggests that small molecular inhibitors of CDKs might be attractive as prospective antitumor agents. To explore the relationship between the structures of substituted isoquinoline-1,3-(2H,4H)-diones and their inhibition of CDK4, 3D-QSAR studies were performed on a dataset of 48 compounds. The bioactive conformation of template compound 34 was obtained by performing molecular docking into the ATP binding site of the homology model of CDK4 and ranking by highest consensus score, which was then used to build and align the rest of the molecules in the series. The constructed comparative molecular similarity indices analysis (CoMSIA) produces significantly better results than comparative molecular field analysis (CoMFA), with r(2)(cv) = 0.707 and r(2) = 0.988. The contours analysis provides useful information about the structural requirements for substituted isoquinoline-1,3-(2H,4H)-diones for CDK4 inhibitory activity.