Apoptosis is responsible for the cytotoxic effects of anti-GD2 ganglioside antibodies and aurora A kinase inhibitors on human neuroblastoma cells

Małgorzata Durbas; Hanna Rokita; Irena Horwacik; Aneta Wiśniewska; Iwona Nowak

doi:10.18388/abp.2020_6344

Małgorzata Durbas Laboratory of Molecular Genetics and Virology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Kraków, Poland
Hanna Rokita Laboratory of Molecular Genetics and Virology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Kraków, Poland
Irena Horwacik Laboratory of Molecular Genetics and Virology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Kraków, Poland
Aneta Wiśniewska Laboratory of Molecular Genetics and Virology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Kraków, Poland
Iwona Nowak Laboratory of Molecular Genetics and Virology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Kraków, Poland

DOI: https://doi.org/10.18388/abp.2020_6344

Abstract

In recent years, immunotherapy has been identified as an effective treatment method for high-risk neuroblastoma. A previous study demonstrated that an anti-GD2 ganglioside (GD2) mouse 14G2a monoclonal antibody (mAb) combined with a small molecule, i.e., an aurora A kinase inhibitor (MK-5108), significantly increased cytotoxicity against human neuroblastoma cells, as compared to monotherapy. This study aimed to demonstrate the mechanism of neuroblastoma cell death in vitro following the addition of an anti-GD2 human-mouse chimeric ch14.18/CHO mAb (presently used in clinics) and two aurora A inhibitors (MK-5108 and MK-8745). The effects of the aforementioned agents on neuroblastoma cells were determined by measuring the level of ATP, the level of apoptotic and necroptotic markers, and the activity of caspase 3/7. The results revealed that the ch14.18/CHO mAb decreased cellular ATP levels in the IMR-32 and CHP-134 neuroblastoma cell lines, similarly to the 14G2a mAb. Regarding ch14.18/CHO mAb treated IMR-32 cells, the observed cytotoxic effect was concomitant with induced caspase 3 cleavage, which indicated the induction of apoptosis in IMR-32 cells, but not in CHP-134 cells. Furthermore, the MK-5108 inhibitor induced apoptosis, as indicated by the increased cleavage of caspase 3 and increased activity of caspase 3/7. However, the presence of necroptosis was ruled out in MK-5108-treated IMR-32 and CHP-134 cells. In summary, the effects of the combination of ch14.18/CHO mAb and aurora A kinase inhibitors (MK-5108 and MK-8745) were shown to enhance apoptosis in IMR-32 cells compared to when used individually.