Apalutamide Induces Dose and Time-dependent Apoptosis in C6 Glioma Cells Through Modulation of Key Apoptotic Regulators

Apalutamide is a non-steroidal androgen receptor antagonist primarily used in the treatment of castration-resistant prostate cancer. Despite its well-characterized efficacy in prostate cancer, its effects on central nervous system tumors, particularly gliomas, remain unexplored. In this study, we investigated the potential cytotoxic and apoptotic effects of apalutamide on C6 glioma cells using the MTT assay, Annexin V-FITC/PI flow cytometry, and immunofluorescent staining of apoptosis-related proteins.

C6 glioma cells were treated with increasing concentrations of apalutamide (1, 3, 10, and 30 μM) for 24, 48, and 72 hours. Cell viability was assessed via MTT assay to determine IC₅₀ values. Apoptotic cell populations were analyzed by flow cytometry using Annexin V-FITC/PI double staining. Furthermore, immunofluorescence microscopy was employed to evaluate the expression of apoptosis-related markers including cleaved caspase-3, Bax, and Bcl-2.

Our results demonstrated a dose- and time-dependent decrease in cell viability, with a calculated IC₅₀ value at 48 hours of approximately 10 μM. Immunofluorescence analysis confirmed up-regulation of cleaved caspase-3 and Bax, along with down-regulation of anti-apoptotic Bcl-2 expression, supporting the activation of the intrinsic apoptotic pathway. Flow cytometry revealed a significant increase in both early and late apoptotic populations upon apalutamide treatment compared to controls.

In conclusion, this study provides the first evidence that apalutamide exerts cytotoxic and pro-apoptotic effects on C6 glioma cells, suggesting a potential off-target utility of androgen receptor antagonists in glioma treatment strategies. Further in vivo and mechanistic studies are warranted to elucidate the underlying pathways and therapeutic relevance.