Considering the important role that cyclin dependent kinases play in regulating the cell cycle, these proteins are attractive targets for drug development. Selective CDK4/6 inhibitors such as palbociclib, abemaciclib, and ribociclib have become mainstays for use in HR+ breast cancer. Building upon the success of CDK4/6 inhibitors, attention has recently turned to the development of selective CDK2 inhibitors. Recent clinical evidence has implicated increased CDK2 activity in resistance to CDK4/6 inhibitors. However, the broadly essential kinase CDK1 has high homology to CDK2 and is a key off-target challenge in the development of CDK2 inhibitors. We describe here our efforts in utilizing a novel molecular dynamics based strategy for the identification of selectivity via a residue conserved in both CDK2 and CDK1, despite nearly homologous ATP binding sites. This strategy led to a dramatic enrichment in compounds with >100x selectivity over CDK1 as well as tumor growth inhibition activity in efficacy studies. In the course of advancing these selective and potent compounds, we encountered unique clearance mechanisms in rodent that were not predicted from in vitro studies and did not apply to higher species. We will describe the additional characterization that was undertaken through detailed mechanistic in vivo studies and the relevance to human dose prediction. These overall learnings have implications for medicinal chemistry programs where selectivity and species-specific PK challenges are encountered.