Arachidonoylcyclopropylamide (ACPA) reduces viability and induces endoplasmic reticulum stress in glioma cells

Objective: The endocannabinoid system (ECB) is present throughout the human body and has been extensively studied, particularly in the nervous system. Cannabinoid 1 (CB1) ligands and receptors are primarily found in the central nervous system and its cells. Neuroglioma is an aggressive and lifethreatening cancer that accounts for approximately 50% of primary central nervous system (CNS) tumors. The treatment protocol for gliomas typically includes surgery, radiation therapy or chemotherapy. In this study, we evaluated the antiproliferative effects of Arachidonoylcyclopropylamide (ACPA), a CB1 receptor agonist, on H4 human neuroglioma cell line. Methods: In this study, we investigated the antiproliferative, pro-apoptotic, antimigratory and organelle-stress-related effects of ACPA on H4 neuroglioma cells. xCELLigence cell impedance assay (n=6) was performed on untreated H4 cells exposed to ACPA at half the maximum inhibitory concentration (IC50) dose. Annexin V/PI labeling was performed by flow cytometry on IC50 ACPA-exposed and untreated H4 cells. In addition, immunofluorescence labeling of anti-GRP78, anti-LC3B and anti-beta tubulin antibodies were analyzed to evaluate the effects on organelle-stress relationship (n=3). Results: The IC50 dose (2.1X10-7 M) of ACPA on H4 cells was determined by xCELLigence cell impedance assay and this dose reduced the proliferation capacity of H4 neuroglioma cells in a dose-time dependent manner (p < 0.05). Flow cytometry analysis showed that cells treated with IC50 ACPA had a higher late apoptotic rate (n=3). Immunofluorescence analysis showed a significant increase in GRP78 and LC3B labeling. Furthermore, a significant decrease in beta tubulin immunolabeling was observed compared to the untreated group (p < 0.05). Conclusion: ACPA induced apoptosis, suppressed proliferation and was able to induce endoplasmic reticulum stress and autophagy in neuroglioma cells. These findings highlight the potential of ACPA as a therapeutic agent for neuroglioma refractory to clinical treatment and warrant further functional validation.