Abstract 5861: CDK4/6 therapy-induced senescence as a resistance mechanism-MiDAS touch
Authors: Gokmen-Polar Yesim , Gu Yuan , Ruan Ling , Badve Sunil S. 
Published: March 2024

Background: Cyclin-dependent kinase inhibitors (CDK4/6i) have revolutionized the therapy for estrogen receptor positive (ER+) breast cancer. Despite the response to CDK4/6i, resistance and recurrence are still a clinical reality, resulting in death due to breast cancer. G1 cell cycle arrest has been considered as the main mechanism of action of CDK4/6i in luminal breast cancer. However, the response to therapy does not correlate with the expression of CDK4/6 or key cell cycle proteins. It is also unclear whether CDK4/6i-induced senescence enhances the therapeutic response or mediates development of resistance.

Experimental Procedures: We have developed drug-tolerant persisters resistant to CDK4/6 inhibitors (CDK4/6i-DTPs) in a panel of estrogen receptor positive (ER+) cells to explore the molecular mechanisms associated with resistance. Using different cell phenotype assays (i.e., cell viability/cell cycle, senescence/senescence reversibility and wound healing) as well as senescence-associated secretory phenotype (SASP) human cytokine array of 105 markers, we characterized the phenotype and secretome of these CDK4/6i DTPs.

Summary: CDK4/6i-DTPs induced by abemaciclib, palbociclib and ribociclib (all from Selleckchem) demonstrated resistance to these drugs and dramatically slowed growth. The percentage of senescent cells reached 30-89% of the total population for all cell lines in all three drugs, whereas control parental cells exhibited around 10%. CDK4/6i DTPs also exhibited significant cell migration, and increased lysosomal phenotype compared to their parental sensitive cells. Further analysis with human cytokine array revealed that protein levels of growth differentiation factor 15 (GDF-15) are elevated in both cell lysates and conditioned-media of CDK4/6i DTPs, whereas Serpin E1 (Plasminogen Activator Inhibitor, Type I, PAI-1) was unique to secreted CDK4/6i DTPs. Interestingly, we did not observe changes in pro-inflammatory senescence-associated secretory phenotype (SASP) markers such as interleukins. Apart from being a SASP protein, GDF-15 is a mitochondrial cytokine (mitokine), suggesting a role in mitochondrial dysfunction-associated senescence (MiDAS). HMGB1, and LMNB1 are also down-regulated in CDK4/6-DTPs supporting the MiDAS phenotype. CDK4/6i-induced MiDAS appears to be transient, as CDK4/6i-DTPs reverted to the parental phenotype in drug-free culture media 20 days after the removal of CDK4/6i's. This included growth characteristics and sensitivity to CDK4/6i. This reversion of phenotype was observed in all five cell line models tested with all three drugs.

Conclusions: CDK4/6-induces a mitochondrial dysfunction-associated senescence (MiDAS). This appears to be transient in nature as withdrawal of the drug results reacquisition of parental cell phenotype and drug-sensitivity. Targeting MiDAS pathway might be critical to improve efficacy of CDK4/6 inhibitors.

Citation Format: Yesim Gokmen-Polar, Yuan Gu, Ling Ruan, Sunil S. Badve. CDK4/6 therapy-induced senescence as a resistance mechanism-MiDAS touch [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5861.