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Pre-clinical Selectivity Profile of the CDK4/6 Inhibitor Ribociclib Compared With That of Palbociclib and Abemaciclib

Tiedt, Ralph, Delach, Scott, Kovats, Steven, Horn, Thomas, Acker, Michael, Schacher Engstler, Barbara, Su, Fei and Caponigro, Giordano (2017) Pre-clinical Selectivity Profile of the CDK4/6 Inhibitor Ribociclib Compared With That of Palbociclib and Abemaciclib. AACR Annual meeting 2017 .


A hallmark of cancer is unchecked cell division. Retinoblastoma protein (Rb) is a human tumor suppressor that guards the entry point into S phase by binding E2F transcription factors and keeping them inactive. Many growth promoting stimuli increase the expression of D-cyclins, which bind to and activate CDK4/6 kinases. The D-cyclin bound CDK4/6 holoenzymes phosphorylate Rb resulting in the release of E2F, which in turn activates genes required for S phase entry and DNA replication. Numerous oncogenic aberrations converge at the CDK4/6-Rb pathway, providing a strong rationale for developing CDK4/6 inhibitors as cancer therapeutics.
Ribociclib (LEE011) is a selective CDK4/6 inhibitor that has received FDA Breakthrough designation and Priority Review for treatment of hormone receptor positive breast cancer in combination with letrozole and is being tested in additional clinical trials. Here, we describe the pre-clinical selectivity profile of ribociclib in biochemical and cellular assay. Ribociclib inhibits both CDK4 - cyclin D1 and CDK6 - cyclin D3 kinase activity with nanomolar IC50s in biochemical assays. To comprehensively address selectivity of ribociclib in direct comparison with two other clinical CDK4/6 inhibitors, palbociclib and abemaciclib, we made use of the KinomeScan assay platform (DiscoverX) consisting of >450 kinase active site-directed competition binding assays. Based on the binding constants (Kd) for CDK4 and CDK6, we adjusted the test concentrations in the kinase selectivity panel to account for higher potency of abemaciclib. Data indicated that both ribociclib and palbociclib have high selectivity for CDK4 (CDK6 was not covered in the panel) with very few distinct additional binding events detected. In contrast, abemaciclib is a much more promiscuous kinase inhibitor.
Next, we sought to determine the relative potencies of the three inhibitors against CDK4 vs CDK6 in cellular assays. When testing different routinely used readouts of cellular viability, we found that assays that measure metabolic activity (such as CTG) tend to underestimate the effects of CDK4/6 inhibition. Therefore, assays that either directly or indirectly assessed cell number were used in these studies. We first identified cancer cell lines primarily dependent on either CDK4 or CDK6 as judged by combined RNA expression analysis and shRNA/CRISPR-based functional assays. When determining IC50 values for these cell lines with the three CDK4/6 inhibitors, we found that both ribociclib and abemaciclib demonstrated greater activity in CDK4-relative to CDK6-dependent cells, whereas palbociclib was similarly active in both cells types.
In conclusion, the high degree of selectivity of ribociclib suggests that off-target kinase inhibition is an unlikely complication in patients. Moreover, the apparent preference for CDK4 over CDK6 could be an advantage in certain cancer types that are primarily dependent on CDK4.

Item Type: Article
Date Deposited: 11 Jul 2017 00:45
Last Modified: 11 Jul 2017 00:45


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