Most cancers preserve functional retinoblastoma (Rb) and may, therefore, respond to inhibition of D-cyclin-dependent Rb kinases, CDK4 and CDK6. To date, CDK4/6 inhibitors have shown promising clinical activity in breast cancer and lymphomas, but it is not clear which additional Rb-positive cancers might benefit from these agents. No systematic survey to compare relative sensitivities across tumor types and define molecular determinants of response has been described. We report a subset of cancers highly sensitive to CDK4/6 inhibition and characterized by various genomic aberrations known to elevate D-cyclin levels and describe a recurrent CCND1 3′UTR mutation associated with increased expression in endometrial cancer. The results suggest multiple additional classes of cancer that may benefit from CDK4/6-inhibiting drugs such as abemaciclib. •A wide range of sensitivity to abemaciclib is observed among Rb+ tumor cells•CDKN2A mutant cancers show only intermediate sensitivity to CDK4/6 inhibition•D-cyclin activating features are associated with highly sensitive cells•About 5% of endometrial cancers bear a stabilizing mutation in the CCND1 3′UTR Gong et al. identify a subset of cancers highly sensitive to CDK4/6 inhibition, which are characterized by various genomic aberrations known to elevate D-cyclin levels but not by CDKN2A mutations. They also identify a recurrent CCND1 3′UTR mutation associated with increased CCND1 expression in endometrial cancer.