Genomic approach to study cell death induced by Grifola frondosa (D-fraction) in MCF-7 breast cancer cells.

BALOGH, GABRIELA; POSTEMSKY, PABLO; MAILO, DANIEL; CURVETTO, NÉSTOR

Denver, Colorado

Congreso; 100th American Association of Cancer Research Annual Meeting; 2009

American Association of Cancer Research

&amp;lt;!-- /* Style Definitions */ p.MsoNormal, li.MsoNormal, div.MsoNormal {mso-style-parent:""; margin:0cm; margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Times New Roman"; mso-fareast-font-family:"Times New Roman";} @page Section1 {size:612.0pt 792.0pt; margin:70.85pt 3.0cm 70.85pt 3.0cm; mso-header-margin:36.0pt; mso-footer-margin:36.0pt; mso-paper-source:0;} div.Section1 {page:Section1;} --&amp;gt; Grifola frondosa also known as Maitake is one of the most researched medicinal mushrooms. Maitake D-fraction, is a polysaccharide fraction extracted from this mushroom, which has been reported to exert anti-tumor activity in tumor-bearing mice by enhancing the immune system through activation of macrophages, T cells, and natural killer cells. This anti-tumoral effect was reported in ten patients with cancer who were not currently taking any chemotherapeutic drugs. Moreover in another clinical study, 165 patients with various types of advanced cancer were given Maitake D-fraction alone or associated with chemotherapy. Cancer regression or significant symptoms improvement was observed in 58%, 69% and 62% of liver, breast and lung cancer patients respectively. This fraction was able to increase the immune cell activities in combination with chemotherapy in comparison with chemotherapy alone. In the present study, the death mechanims of tumor MCF-7 breast cancer cells treated with Maitake D-fraction (Grifon Maitake D-Fraction® N.J., U.S.A) was evaluated, using both a protocol to follow cytotoxicity response and a protocol for cDNA microarrays genomic analysis. MCF-7 tumor cells were treated in the absence and in the presence of increasing doses of D-fraction in cell culture medium (0.55mg/15ml, 1.375mg/15ml, 2.75mg/15ml and 5.5mg/15ml).The experiment was performed by triplicate. After 24 hours, cell death was determined using Toluidine Blue stain and the alive cells stained were counted under light microscope. The results shown that D-fraction induces cell death (75%) in the tumor MCF-7 cells in culture; achieving the maximum effect at 5.5mg/15ml (p<0.01). The cell death was also observed and recorded by time lapse microscopy every 10 minutes. The genomic signature induced by D-fraction on MCF-7 cells was evaluated using cDNA microarray technology containing 10,000 human genes. The analysis revealed that Maitake D-fraction induces or modify the expression level of 4,321 genes in the tumor cells respect to the controls. These genes are related with several biological functions, i.e. some of them are related to apoptosis, cell-death, arrest of cell cycle, negative regulation of cell cycle progression and gene transcription. It were detected 29 genes related to apoptosis, such as BCL2-antagonist/killer 1 (BAK), nerve growth factor receptor (TNFR superfamily, member 16), tumor necrosis factor receptor (superfamily, member 19) and tumor necrosis factor receptor (superfamily, member 21). It can be concluded that Maitake D-fraction induces cell-death mechanisms in MCF-7 breast tumor cells by involving the activaction of those genes. This work was supported by the National Research Council (CONICET) and the Universidad Nacional del Sur from Bahia BlancaCity (Argentina).