Recent studies have pointed out the preventive role of beetroot extracts against cancers and their cytotoxic activity on cancer cells. Among many different natural compounds, these extracts contained betanin and its stereoisomer isobetanin, which belongs to the betalain group of highly bioavailable antioxidants. However, a precise identification of the molecules responsible for this tumor-inhibitory effect was still required. We isolated a betanin/isobetanin concentrate from fresh beetroots, corresponding to the highest purified betanin extract used for studying anticancer activities of these molecules. The cytotoxicity of this betanin-enriched extract was then characterized on cancer and normal cells and we highlighted the death signalling pathways involved. Betanin/isobetanin concentrate significantly decreased cancer cell proliferation and viability. Particularly in MCF-7-treated cells, the expressions of apoptosis-related proteins (Bad, TRAILR4, FAS, p53) were strongly increased and the mitochondrial membrane potential was altered, demonstrating the involvement of both intrinsic and extrinsic apoptotic pathways. Autophagosome vesicles in MCF-7-treated cells were observed, also suggesting autophagic cell death upon betanin/isobetanin treatment. Importantly, the betanin-enriched extract had no obvious effect towards normal cell lines. Our data bring new insight to consider the betanin/isobetanin mix as therapeutic anticancer compound, alone or in combination with classical chemotherapeutic drugs, especially in functional p53 tumors.

p53-related genes, p73 and p63, encode 2 classes of proteins, TAp73/p63 and DN-p73/p63. TA-p73/p63 demonstrate p53-like properties including gene transactivation and cell death promotion, whereas DN-p73/p63 lack these p53-like functions. Although p53-deficient cancer cells are often less responsive to chemotherapy, they are not completely drug resistant, suggesting that other apoptotic pathways are at work. Here, we compared for the first time to our knowledge p73 and p63 activation in various breast cancer (BC) cell lines after Adriamycin (ADR) treatment, an agent considered as mandatory in breast cancer chemotherapy. Our study was carried out using 1 p53-proficient BC cell line (MCF7 cells) and 3 BC cell lines deficient in p53 response (MCF7/ADR IGR , MDA-MB157 and T47D) after ADR-induced genotoxic stress. We report that in cells with no p53 response after ADR treatment, TAp73, but not TAp63 or DN-p73/p63, may replace p53 in triggering not only apoptosis but also cell cycle arrest or DNA repair effectors such as p21, GADD45, 14-3-3r and p53R2. We also demonstrate that TAp73 siRNA inhibits the accumulation of TAp73 in response to ADR treatment in MDA-MB157 cells and confers protection against ADR. ADR-induced downregulation of the DNp73 isoform in the T47D cell line with nonfunctional mutant p53 further supports anti-apoptotic function of the isoform antagonistic to both p53 and TA-p73/p63. Exogenous TAp73 and DNp73 overexpression in p53-response-deficient cell lines further confirms these results. cDNA microarray techniques demonstrated that the cellular response induced by p73 during ADR treatment could involve specific genes. ' 2005 Wiley-Liss, Inc.