Background: Neuroblastoma is an aggressive childhood cancer poorly responsive to therapy. Moreover, survivors experience long-term side effects from their treatment highlighting the need for effective and less toxic therapies. Catechin is a natural polyphenol with anti-cancer properties and low toxicity. Poor serum stability of Catechin limits its clinical use that we overcame by chemical functionalization with Dextran. Herein, we investigated the efficacy of Dextran-Catechin as neuroblastoma treatment.
Methods: Anticancer activity was tested in 4 independent neuroblastoma cell lines using the viability assay Alamar Blue and apoptosis was assessed using PARP cleavage. Gene expression was determined using qRT-PCR and protein expression by western blotting. Intracellular copper was measured by spectrophotometric analysis. Fluorescence-lifetime imaging microscopy was used to determine oxidative stress induction. Cellular levels of the antioxidant GSH was examined using a colorimetric assay. PET imaging with Cu64 was performed in a xenograft neuroblastoma model to monitor copper homeostasis. In vivo anticancer activity of Dextran-Catechin was assessed in human xenograft model of neuroblastoma.
Results: The neuroblastoma cell lines SH-SY5Y, IMR-32, BE(2)C and doxorubicin-resistant BE(2)C-ADR were sensitive to Dextran-Catechin (IC50 9.7 µg/ml, 17.83 µg/ml, 16 µg/ml and 18.2 µg/ml, respectively) at concentrations not toxic for non-malignant MRC-5 cells. However, Cisplatin-resistant cells, IMR-32-CisRes, were 2.5-fold resistant against Dextran-Catechin compared to the parental cells. Copper transporter 1 (CTR1) mediates cisplatin uptake and IMR-32-CisRes exhibited 50% lower expression of CTR1 and lower intracellular copper compared to the IMR-32 cells. In contrast, Dextran-Catechin sensitive neuroblastoma cells showed high copper levels. We demonstrated that Dextran-Catechin reacts with copper generating reactive oxygen species and inducing cancer cell death. PET imaging analysis of the neuroblastoma xenograft model revealed high accumulation of copper in the tumor mass. Importantly, we showed that Dextran-Catechin significantly reduced copper uptake and tumor growth in human xenograft model of neuroblastoma with no evidence of side effects.
Conclusion: Dextran-Catechin targets copper, inhibits tumour growth, and has therapeutic potential for cancers dependent on copper for their growth.