Metallothionein-3 has poorly characterized functions in neuroblastoma. Cisplatin-based chemotherapy is a major regimen to treat neuroblastoma, but its clinical efficacy is limited by chemoresistance. We investigated the impact of human metallothionein-3 (hMT3) up-regulation in neuroblastoma cells and the mechanisms underlying the cisplatin-resistance. We confirmed the cisplatin-metallothionein complex formation using mass spectrometry. Overexpression of hMT3 decreased the sensitivity of neuroblastoma UKF-NB-4 cells to cisplatin. We report, for the first time, cisplatin-sensitive human UKF-NB-4 cells remodelled into cisplatin-resistant cells via high and constitutive hMT3 expression in an in vivo model using chick chorioallantoic membrane assay. Comparative proteomic analysis demonstrated that several biological pathways related to apoptosis, transport, proteasome, and cellular stress were involved in cisplatin-resistance in hMT3 overexpressing UKF-NB-4 cells. Overall, our data confirmed that up-regulation of hMT3 positively correlated with increased cisplatin-chemoresistance in neuroblastoma, and a high level of hMT3 could be one of the causes of frequent tumour relapses.

• MeSH
• Drug Resistance, Neoplasm drug effects   genetics   MeSH
• Cisplatin pharmacology   MeSH
• Chick Embryo MeSH
• Humans MeSH
• Metallothionein 3 biosynthesis   genetics   MeSH
• Cell Line, Tumor MeSH
• Neoplasm Proteins biosynthesis   genetics   MeSH
• Gene Expression Regulation, Neoplastic drug effects   MeSH
• Animals MeSH
• Check Tag
• Chick Embryo MeSH
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Cisplatin MeSH
• Metallothionein 3 MeSH
• Neoplasm Proteins MeSH

The efficiency of cisplatin (CDDP) is significantly hindered by the development of resistance during the treatment course. To gain a detailed understanding of the molecular mechanisms underlying the development of cisplatin resistance, we comparatively analyzed established a CDDP-resistant neuroblastoma cell line (UKF-NB-4CDDP) and its susceptible parental cells (UKF-NB-4). We verified increased chemoresistance of UKF-NB-4CDDP cells by analyzing the viability, induction of apoptosis and clonal efficiency. To shed more light on this phenomenon, we employed custom cDNA microarray (containing 2234 probes) to perform parallel transcriptomic profiling of RNA and identified that 139 genes were significantly up-regulated due to CDDP chemoresistance. The analyses of molecular pathways indicated that the top up-regulation scoring functions were response to stress, abiotic stimulus, regulation of metabolic process, apoptotic processes, regulation of cell proliferation, DNA repair or regulation of catalytic activity, which was also evidenced by analysis of molecular functions revealing up-regulation of genes encoding several proteins with a wide-spectrum of enzymatic activities. Functional analysis using lysosomotropic agents chloroquine and bafilomycin A1 validated their potential to re-sensitize UKF-NB-4CDDP cells to CDDP. Taken together, the identification of alterations in specific genes and pathways that contribute to CDDP chemoresistance may potentially lead to a renewed interest in the development of novel rational therapeutics and prognostic biomarkers for the management of CDDP-resistant neuroblastoma.

• Keywords
• chemoresistance, cisplatin, lysosomes, microarray, neuroblastoma, transport,
• MeSH
• Clone Cells MeSH
• Drug Resistance, Neoplasm drug effects   genetics   MeSH
• Chloroquine pharmacology   MeSH
• Cisplatin pharmacology   therapeutic use   MeSH
• Gene Ontology MeSH
• Gene Regulatory Networks drug effects   MeSH
• Humans MeSH
• Lysosomes drug effects   metabolism   MeSH
• Macrolides pharmacology   MeSH
• Cell Line, Tumor MeSH
• Neuroblastoma drug therapy   genetics   pathology   MeSH
• Gene Expression Regulation, Neoplastic drug effects   MeSH
• Transcriptome drug effects   genetics   MeSH
• Cell Shape drug effects   MeSH
• Up-Regulation drug effects   genetics   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• bafilomycin A1 MeSH Browser
• Chloroquine MeSH
• Cisplatin MeSH
• Macrolides MeSH