Abstract P6-04-02: CDK6 might be a key factor for efficacy of CDK4/6 inhibitor and the hormone sensitivity following acquired resistance
Authors: Iida M , Nakamura M , Tokuda E , Niwa T , Ishida T , Hayashi S-I 
Published: February 2018

Background: CDK4/6 inhibitors have received FDA breakthrough therapy designation as 1st line treatment for advanced estrogen receptor positive breast cancer patients. However, the benefit offered by CDK4/6 inhibitors is individually different and furthermore acquired resistance to the drugs is monumental challenges. It is urgent need to search for the biomarker and understand the drug sensitivity and its alteration after acquired resistance.

Results: To identify the efficacy of CDK4/6 inhibitors, we assessed IC50 of ribociclib in several cell lines. Luminal cell lines (MCF-7, T-47D) exhibited lower ribociclib IC50 than HER-2 (SK-BR-3) and triple negative cell lines (MDA-MB-231, BT-20). Immunoblot analysis of Luminal cell lines showed extremely lower levels of CDK6 compared with others. CDK6-transfected MCF-7 by means of expression vector reduced the sensitivity equivalent to MDA-MB-231 not only to ribociclib but also to palbociclib and abemaciclib. Protein level of ERα in CDK6-transfected MCF-7 stayed unchanged and fulvestrant sensitivity was unaltered as well.

Subsequently, we detect the efficacy of ribociclib in hormone resistant cell lines. Estrogen deprivation-resistant (EDR) cells (EDR1:ER-positive, EDR2:ER-negative) and fulvestrant resistance (MFR) cells (loss of ER expression) established from MCF-7 maintained ribocilib sensitivity to the same degree with MCF-7. No marked difference in IC50 was observed between EDR1/2 and MFR, and CDK6 expressions were comparable to MCF-7. These results suggest that high level of CDK6 expression weaken the sensitivity to CDK4/6 inhibitors. The inhibitors would provide more effective benefits to tumors expressing lower level of CDK6 than the higher, independent of hormone sensitivity.

To understand the characteristics in acquired resistance, ribociclib resistant cell lines (RIBR1/2) were established from EDR1 by long-term culture with ribociclib. RIBR designed lower level of p21, p27 and ERα by immunoblot analysis. EDR1 were promoted cell growth by estrogen addition, while RIBR were not. Further, ER activity of RIBR was intensely decreased, and mRNA levels of the ER target genes, PgR and EGR3 were also decreased. Therefore, the responsiveness to tamoxifen and fulvestrant were lost. On the other hand, PI3K inhibitor and mTOR inhibitor suppressed cell growth to the same extent as EDR1, suggesting that RIBR were reduced ER dependence and remain reliant on PI3K/Akt/mTOR pathway.

Conclusion: The possibility of CDK6 as a biomarker is corroborated by the finding that low level of CDK6 expression is positively correlated with the efficacy of CDK4/6 inhibitor. Further ER dependence had decreased after acquired CDK4/6 inhibitor resistance whereas the dependence on PI3K/Akt/mTOR pathway still remain, indicating the inhibition of PI3K/Akt/mTOR pathway would be amenable to therapeutic target.

Citation Format: Iida M, Nakamura M, Tokuda E, Niwa T, Ishida T, Hayashi S-I. CDK6 might be a key factor for efficacy of CDK4/6 inhibitor and the hormone sensitivity following acquired resistance [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P6-04-02.