Investigation of the effect of Abemaciclib on breast cancer spheroids in a three-dimensional (3D) environment

Breast cancer is a leading cause of cancer-related mortality worldwide, with triple-negative breast cancer (TNBC) representing the most aggressive subtype due to its lack of targetable receptors and poor prognosis. Abemaciclib, a selective cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitor, has emerged as a promising therapeutic agent by blocking cell-cycle progression and limiting tumor proliferation. To better mimic tumor architecture, this study employed spheroids of the TNBC cell line MDA-MB-231 generated with the three-dimensional (3D) Petri Dish® system. The cytotoxic effects of Abemaciclib were evaluated in comparison with Doxorubicin under both two-dimensional (2D) monolayer and three dimensional (3D) spheroid culture conditions using an MTS-based cell viability assay. Complementary analyses included immunofluorescence staining of filamentous actin (F-actin) to assess cytoskeletal alterations, and flow cytometry to determine cell-cycle distribution. Within the tested concentration ranges, spheroids showed reduced sensitivity to Abemaciclib compared with monolayers, indicating that 3D culture conditions may influence drug penetration and adaptive resistance. Abemaciclib suppressed proliferation in a concentration-dependent manner by altering cell-cycle distribution, with G0/G1 accumulation at lower concentrations and G2/M accumulation at higher concentrations, accompanied by a reduced S-phase fraction.