Abstract P3-01-02: Oxalate microcalcifications induce breast cancer tumors

Abstract One shared characteristic of breast cancer is the appearance of mammographic upon mammography examination. Microcalcifications are routinely used to detect breast cancer in its early stages, which is of key importance due to the possibility that early detection gives for the application of more conservative therapies and for a better patient outcome. Mammary microcalcifications are composed of calcium oxalate or calcium phosphate. Oxalate has been considered as an inert end product of the metabolism. However, its accumulation is thought to be toxic to living tissues because it induces some pathological circumstances. Several results support the hypothesis that Oxalates are a secretion product that is produced by many kinds of cells, including apocrine cells, kidney cells and liver cells, among others. Exposure of renal epithelial cells to oxalate leads to diverse events that include, among others, induction of immediate early genes (IEG) expression, re-initiation of DNA synthesis, cell growth, and up or down-regulation of more than 1000 genes. Not enough research has been carried out directed to investigate the impact the presence of oxalates has within the breast tumor microenvironment. Neither are the interactions between oxalates breast-epithelial cells nor the signal transduction pathways involved in it, well understood. Herein, we have obtained good evidence showing that oxalates excreted from apocrine cells induce alterations in normal breast epithelial cells, and that the exposure of breast epithelial cells to excreted oxalate in a chronic way, sets the stage for transformation from normal to fully developed breast tumors. Oxalate induces DNA synthesis and IEG overexpression in MCF7 and MDAMB231 human breast cancer cells in culture. When these cells were treated with oxalate, the IEG c-Fos and Fra-1 were over-expressed as determined by Western Blot. Additionally, in human breast cancer tissues we found a strict correlation between oxalate concentration and overexpression of c-Fos and Fra-1. Paired adjacent non-tumor tissues linked with the tumor tissues have neither high levels of oxalate nor c-Fos/Fra-1 over-expression. Two groups of female mice (n = 20 each) periodically received an injection of oxalate or of saline solution (controls) into a mammary fatpad during a month. Surprisingly, the mice that were injected with oxalate generated very aggressive tumors. c-Fos and Fra-1 were also found over-expressed in these tumors. Furthermore, the final oxalate concentrations in the mice tumors were analogous to those found in the human breast tumor tissues whereas oxalate was almost undetectable in the non-pathological control tissues. Mice breast tumors were palpable within three months in 18/20 of oxalate treated as compared to the 0/20 control animals. Eight tumors were very aggressive and showed very fast growth rates; in these tumors, it was possible to appreciate high rate of atypical mitosis upon haematoxylin/eosin staining. Five tumors were very invasive and were found multi-localized in all breast areas. These five tumors also showed an atypically high mitotic rate, big pleomorphic nuclei, and macro and multi nucleoli. We expect that if we can control oxalic production or even better, control their action, we will significantly reduce the development of human breast cancers. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P3-01-02.</jats:p

Old Players with a Newly Defined Function: Fra-1 and c-Fos Support Growth of Human Malignant Breast Tumors by Activating Membrane Biogenesis at the Cytoplasm

A shared characteristic of tumor cells is their exacerbated growth. Consequently, tumor cells demand high rates of phospholipid synthesis required for membrane biogenesis to support their growth. c-Fos, in addition to its AP-1 transcription factor activity, is the only protein known up to date that is capable of activating lipid synthesis in normal and brain tumor tissue. For this latter activity, c-Fos associates to the endoplasmic reticulum (ER) through its N-terminal domain and activates phospholipid synthesis, an event that requires it Basic Domain (BD) (aa 139–159). Fra-1, another member of the FOS family of proteins, is over-expressed in human breast cancer cells and its BD is highly homologous to that of c-Fos with two conservative substitutions in its basic amino acids. Consequently, herein we examined if Fra-1 and/or c-Fos participate in growth of breast cancer cells by activating phospholipid synthesis as found previously for c-Fos in brain tumors. We found both Fra-1 and c-Fos over-expressed in >95% of human ductal breast carcinoma biopsies examined contrasting with the very low or undetectable levels in normal tissue. Furthermore, both proteins associate to the ER and activate phospholipid synthesis in cultured MCF7 and MDA-MB231 breast cancer cells and in human breast cancer samples. Stripping tumor membranes of Fra-1 and c-Fos prior to assaying their lipid synthesis capacity in vitro results in non-activated lipid synthesis levels that are restored to their initial activated state by addition of Fra-1 and/or c-Fos to the assays. In MDA-MB231 cells primed to proliferate, blocking Fra-1 and c-Fos with neutralizing antibodies blocks lipid-synthesis activation and cells do not proliferate. Taken together, these results disclose the cytoplasmic activity of Fra-1 and c-Fos as potential targets for controlling growth of breast carcinomas by decreasing the rate of membrane biogenesis required for growth