Differential Responses of Adrenergic- and Mesenchymal-Like Neuroblastoma Cells to Snake Venom Toxins

Neuroblastoma is a heterogeneous pediatric malignancy characterized by distinct cellular states, including adrenergic (ADRN) and mesenchymal (MES) phenotypes, which differ in their molecular profiles and therapeutic responses. Snake venom toxins have shown antitumor potential, including the ability to induce apoptotic cell death. This study investigated the differential sensitivity of ADRN- and MES-like neuroblastoma cells to purified snake venom toxins, considering their basal expression of apoptosis-related proteins. Six snake venom toxins (crotamine, BthTX-I, MjTX-II, crotoxin B subunit [CB], a metalloprotease, and BJcuL lectin) were screened in SH-SY5Y (ADRN-like) and SK-N-AS (MES-like) cells. Cellular responses were assessed using methylene blue staining, followed by the MTT assay at concentrations up to 50µg/mL. Basal expression of apoptotic and survival proteins (caspases-2, -8, -9, -10; Bcl-XL, Mcl-1, FLIP, Bid, Bak, DR4, and DR5) was investigated by Western blot. Initial screening identified four toxins for further evaluation, with BthTX-I and CB showing the most relevant differential responses. At 50 µg/mL, BthTX-I reduced methylene blue staining and MTT activity in SH-SY5Y cells to approximately 55% and 60% of control levels, respectively, while SK-N-AS cells were less affected. Conversely, CB preferentially affected SK-N-AS cells, reducing methylene blue staining to ~40% and MTT activity to 60%, whereas SH-SY5Y values remained above 80%. Western blot revealed distinct basal profiles: SK-N-AS cells exhibited stronger caspase-8 and caspase-10 signals, with detectable Bcl-XL and DR5, whereas SH-SY5Y showed prominent Bid and weak/undetectable caspase-10 and Bcl-XL. Caspase-9, caspase-2, Mcl-1, and FLIP were detected in both. These differences suggest distinct regulation of apoptotic pathways influencing cellular responses. BthTX-I and CB exhibited differential activity against ADRN- and MES-like cells displaying distinct basal protein profiles. These findings suggest that variations in apoptotic signaling contribute to toxin sensitivity. Future investigations will determine if apoptosis is the primary mechanism of cell death and evaluate effects on cell adhesion and migration.