A Novel Approach for Glioblastoma Treatment by Combining Apoptosis Inducers (TMZ, MTX, and Cytarabine) with E.V.A. (Eltanexor, Venetoclax, and A1210477) Inhibiting XPO1, Bcl-2, and Mcl-1

Adjuvant therapy for Grade 4 Glioblastoma (GB) with Temozolomide (TMZ) ultimately fails due to the development of therapeutic resistance, highlighting the need for alternative strategies. The limited efficacy of apoptosis induction in GB cells arises from the overexpression of anti-apoptotic proteins, particularly from the XPO1 and Bcl-2 families. We evaluated the effects of TMZ, Methotrexate (MTX), and Cytarabine (Ara-C)—agents known to promote apoptosis—combined with inhibitors targeting XPO1, Bcl-2, and Mcl-1 to restore apoptotic function in GB cell lines and primary glioblastoma stem-like cells (GSCs).

Dose-response curves were generated using CellTiter-Glo® and Caspase-3 activity assays, while gene and protein expression of anti-apoptotic factors were assessed through PCR and Western blot analysis. Optimal KPT-8602 drug combinations were further analyzed for their effects on the cell cycle and apoptosis via FACS, alongside toxicity assessments in healthy mouse brain slices.

Ara-C and MTX demonstrated 150- to 10,000-fold greater potency in triggering apoptosis compared to TMZ. Treatment with the inhibitors Eltanexor (targeting XPO1; E), Venetoclax (targeting Bcl-2; V), and A1210477 (targeting Mcl-1; A) resulted in compensatory upregulation of the corresponding anti-apoptotic genes. However, combining TMZ, MTX, or Ara-C with E, V, and A produced highly lethal effects in GB cells. Notably, no significant toxicity was observed in mouse brain tissue, suggesting that this drug combination is both effective in vitro and likely to exhibit minimal side effects in vivo.