Structural and biochemical studies revealed how p27 interacts with and activates CDK4.

  • Major Finding: Structural and biochemical studies revealed how p27 interacts with and activates CDK4.

  • Concept: p27 binding prevented inhibition of CDK4 by palbociclib, a CDK4/6 inhibitor used in the clinic.

  • Impact: Palbociclib may not work as previously envisioned, and p27′s role in CDK4 regulation is complex.


Some CDK4/6 inhibitors are approved for the treatment of HER2-negative, ER-positive breast cancer and are being investigated for the treatment of other cancers in clinical trials; however, questions remain about CDK4 regulation. Guiley and colleagues investigated CDK4 regulation by the intrinsically disordered proteins p21 and p27, which have seemingly contradictory roles in CDK4 regulation: Some evidence shows that they are CDK inhibitors, but other evidence indicates that they are required for CDK4–cyclin D assembly and activity in vivo. Crystal structures of the p21–CDK4–cyclin D1 and p27–CDK4–cyclin D1 trimer complexes revealed that p21′s and p27′s inhibitory functions may arise from their blockage of a substrate-docking site on cyclin D and the fact that interactions with either one's D2 domains shifts some amino-acid residues that form part of CDK4′s ATP-binding pocket. Interestingly, though, comparison of these trimer structures with the structure of the previously published CDK4–cyclin D dimer complex revealed that binding of p21 or p27 caused rotation of the N-lobe of CDK4 relative to the C-lobe, releasing the activation segment of the protein from the active site, and caused a rearrangement that poised the catalytic lysine residue for activity. Thus, while still inhibited, the trimer conformations may represent intermediates along the pathway to activation. Biochemical studies showed that tyrosine phosphorylation of p27, but not p21, led to increased RB phosphorylation by CDK4, and a structure of the phospho-p27–CDK4–cyclin D complex indicated that this effect was due to relief from inhibition by p27′s D2 domain, which binds CDK4′s N-lobe. Notably, the CDK4/6 inhibitor palbociclib could not bind p27–CDK4–cyclin D trimers and poorly inhibited these trimers both in vitro and in a breast cancer cell line. Instead, palbociclib primarily bound CDK4/6 monomers in cells, and treatment appeared to indirectly cause CDK2 inhibition, indicating that its mechanism may be different than what is generally thought. In summary, this study provides mechanistic insight into a process important in cancer and sheds light on a potentially misunderstood drug mechanism.

Guiley KZ, Stevenson JW, Lou K, Barkovich KJ, Kumarasamy V, Wijeratne TU, et al. p27 allosterically activates cyclin-dependent kinase 4 and antagonizes palbociclib inhibition. Science 2019;366. pii: eaaw2106.

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