Cyclin-dependent kinase 2 (CDK2) has been garnering considerable interest as a target to develop new cancer treatments and to ameliorate resistance to CDK4/6 inhibitors. However, a selective CDK2 inhibitor has yet to be clinically approved. With the desire to discover novel, potent, and selective CDK2 inhibitors, the phenylsulfonamide moiety of our previous lead compound 1 was bioisosterically replaced with pyrazole derivatives, affording a novel series of N,4-di(1H-pyrazol-4-yl)pyrimidin-2-amines that exhibited potent CDK2 inhibitory activity. Among them, 15 was the most potent CDK2 inhibitor (K_i = 0.005 µM) with a degree of selectivity over other CDKs tested. Meanwhile, this compound displayed sub-micromolar antiproliferative activity against a panel of 13 cancer cell lines (GI₅₀ = 0.127–0.560 μM). Mechanistic studies in ovarian cancer cells revealed that 15 reduced the phosphorylation of retinoblastoma at Thr821, arrested cells at the S and G2/M phases, and induced apoptosis. These results accentuate the potential of the N,4-di(1H-pyrazol-4-yl)pyrimidin-2-amine scaffold to be developed into potent and selective CDK2 inhibitors for the treatment of cancer.

The global burden of cancer necessitates rapid and ongoing development of effective cancer therapies. One promising approach in this context is the repurposing of existing non-cancer drugs for cancer indications. A key to this approach is selecting the cellular targets against which to identify novel repurposed drugs for pre-clinical analysis. Protein kinases are highly sought-after anticancer drug targets since dysregulation of kinases is the hallmark of cancer. To identify potential kinase-targeted drug candidates from the existing portfolio of non-cancer therapeutics, we used combined in silico and in vitro approaches, including ligand-based 3D screening followed by biochemical and cellular assessments. This strategy revealed that the anti-viral drug rilpivirine is an Aurora A kinase inhibitor. In view of previous findings implicating Aurora A kinase in abnormal cell cycle regulation, we also examined the influence of rilpivirine on the growth of T47D breast cancer cells. Herein, we detail the identification of rilpivirine as an Aurora A kinase inhibitor, its molecular basis of inhibitory activity towards this kinase, and its Aurora A-mediated anticancer mechanisms in T47D cells. Our results illustrate the value of integrated in silico and in vitro screening strategies in identifying repurposed drug candidates and provide a scientific basis for further exploring the potential anticancer properties of the anti-viral drug rilpivirine.