Epigallocatechin-3-gallate and 6-OH-11-O-hydroxyphenanthrene limit BE(2)-C neuroblastoma cell growth and neurosphere formation in vitro

We conducted an in vitro study combining a rexinoid, 6-OH-11-O-hydroxyphenanthrene (IIF), and epigallocatechin-3-gallate (EGCG), which is the main catechin of green tea, on BE(2)-C, a neuroblastoma cell line representative of the high-risk group of patients. Neuroblastoma is the most common malignancy of childhood: high-risk patients, having N-MYC over-expression, undergo aggressive therapy and show high mortality or an increased risk of secondary malignancies. Retinoids are used in neuroblastoma therapy with incomplete success: the association of a second molecule might improve the efficacy. BE(2)-C cells were treated by EGCG and IIF, individually or in combination: cell viability, as evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay, was reduced, EGCG+IIF being the most effective treatment. Apoptosis occurred and the EGCG+IIF treatment decreased N-MYC protein expression and molecular markers of invasion (MMP-2, MMP-9 and COX-2). Zymography demonstrated nearly 50% inhibition of MMP activity. When BE(2)-C cells were grown in non-adherent conditions to enrich the tumor-initiating cell population, BE(2)-C-spheres were obtained. After 48 h and 72 h treatment, EGCG+IIF limited BE(2)-C-sphere formation and elicited cell death with a reduction of N-MYC expression. We concluded that the association of EGCG to IIF might be applied without toxic effects to overcome the incomplete success of retinoid treatments in neuroblastoma patients.