Abstract PD2-04: Different mechanism of CDK4/6 inhibitor resistance between ribociclib and abemaciclib
Authors: Iida Masafumi , Toyosawa Daichi , Nakamura Misato , Tokuda Emi , Niwa Toshifumi , Yoshida Ryuichi , Ishida Takanori , Hayashi Shin-ichi 
Published: March 2020

Abstract
Background: CDK4/6 inhibitors—palbociclib, ribociclib, and abemaciclib—result in prolonged progression free survival for advanced estrogen receptor positive breast cancer patients. However, the optimum CDK4/6 inhibitor for each patient remains unclear until the emergence of side effects, which are the key differentiators between the two treatments. Furthermore, an optimum treatment after acquisition of resistance is poorly understood. Therefore, we elucidated the difference in CDK4/6 inhibitor resistance mechanisms using ribociclib- and abemaciclib-resistant cell lines in vitro.Methods: We established hormone-resistant cell lines. Estrogen deprivation-resistant (EDR) cell lines (EDR1:ER-positive, EDR2:ER-negative) and fulvestrant-resistant (MFR) cell lines were established from the MCF-7 cell line. We established ribociclib-resistant cell lines (RIBR) and abemaciclib-resistant cell lines (ABER) from EDR1 by long-term culture (7 months for RIBR and 12 months for ABER) with ribociclib and abemaciclib, respectively. We also established CDK6 overexpressed cell lines (MCF7-C6) from MCF-7 cell line using the stably transfected CDK6 expression vector.Results: First, we identified the efficacies of ribociclib and abemaciclib in several cell lines. Luminal cell lines (MCF-7 and T-47D) exhibited high sensitivity to ribociclib and abemaciclib compared to non-luminal cell lines (MDA-MB-231, BT-20, and SKBR3). The hormone-resistant cell lines (EDR1, EDR2, and MFR) exhibited similar sensitivity to ribociclib and abemaciclib as that of MCF-7; this was independent of hormone resistant mechanism.Subsequently, we compared RIBR and ABER, which are the models of acquired ribociclib- and abemaciclib-resistant cell lines, respectively. We confirmed that RIBR and ABER decreased ribociclib and abemaciclib sensitivity, respectively. RIBR showed low sensitivity to abemaciclib. Similarly, ABER exhibited low sensitivity to ribociclib. Immunoblot analysis showed that compared to EDR1, RIBR and ABER had extremely low levels of total Retinoblastoma (RB). Interestingly, compared to EDR1, CDK6 levels were upregulated in ABER and remained unaltered in RIBR. On the other hand MCF7-C6 reduced sensitivity not only abemaciclib but also ribociclib compared to MCF-7 cell line. Furthermore, compared to EDR1, RIBR exhibited lower levels of p21 and p27 compared to EDR1 (P21 levels were extremely reduced, but p27 levels were slightly reduced). Additionally, ABER showed markedly lower levels of p27; however, the p21 level was slightly increased compared to EDR1. PI3K inhibitor and mTOR inhibitor suppressed cell growth in RIBR and ABER to the same extent as EDR1, indicating that resistance to CDK4/6 inhibitors was still dependent on the PI3K/Akt/mTOR pathway. Chemotherapeutic drugs were effective both in RIBR and ABER.Discussion & Conclusion: There are various mechanisms of CDK4/6 inhibitor resistance, such as loss of RB, CDK6 amplification, and high CDK2 activity. However, the mechanism of resistance between ribociclib and abemaciclib might be different because the CDK6 level was different, although RB level was decreased in both RIBR and ABER. Furthermore, both p21 and p27 are considered as the internal inhibitors of CDK2, although more factors in RIBR and ABER were different. CDK4/6 inhibitor resistant cell lines acquired cross resistance to another CDK4/6 inhibitor. Although the mechanism of resistance between ribociclib and abemaciclib might be different, use of drugs targeting the PI3K/Akt/mTOR pathway and chemotherapeutic drugs would be an effective strategy following the development of resistance. Our findings suggest that the mechanism of resistance differs depending on the type of CDK4/6 inhibitors used, and provide some insights into the treatment strategies after acquisition of CDK4/6 inhibitor resistance.
Citation Format: Masafumi Iida, Daichi Toyosawa, Misato Nakamura, Emi Tokuda, Toshifumi Niwa, Ryuichi Yoshida, Takanori Ishida, Shin-ichi Hayashi. Different mechanism of CDK4/6 inhibitor resistance between ribociclib and abemaciclib [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr PD2-04.