Melatonin induces autophagy in neuroblastoma by alleviating Pak2‑mediated endoplasmic reticulum stress

Neuroblastoma (NB), the most common extracranial solid tumor in children, remains challenging to treat due to limited therapeutic efficacy and poor prognosis. Emerging evidence highlights the critical roles of endoplasmic reticulum (ER) stress and autophagy in cancer progression. The present study investigated the therapeutic potential of melatonin in neuroblastoma and its underlying mechanisms. Using Neuro‑2a (N2a) cells, it demonstrated that melatonin alleviated ER stress by upregulating ER chaperones glucose‑regulated protein (GRP)78 and GRP94 and the pro‑apoptotic protein CHOP, while enhancing autophagic activity. Western blotting revealed increased LC3‑II/I ratios, elevated autophagy‑related protein 5 and Beclin1 levels, and reduced p62 expression, indicating autophagy induction. Immunofluorescence and transmission electron microscopy confirmed the dose‑dependent accumulation of autophagosomes. ER stress inhibitor 4‑phenylbutyric acid attenuated melatonin‑induced autophagy, linking ER stress relief to autophagic activation. Mechanistically, melatonin upregulated p21‑activated kinase 2 (Pak2), which suppressed mTOR phosphorylation and activated unc‑51‑like kinase 1, thereby modulating the AMP‑activated protein kinase (AMPK) pathway. Pak2 overexpression amplified melatonin's ER stress‑alleviating effects, whereas Pak2 knockdown or AMPK inhibition diminished its efficacy. These findings established that melatonin suppresses neuroblastoma growth by mitigating Pak2‑mediated ER stress to induce cytotoxic autophagy. The present study provided novel insights into melatonin as a promising therapeutic agent for neuroblastoma, warranting further exploration in preclinical models and clinical trials.