Nuclear beta-catenin and CD44 upregulation characterize invasive cell populations in non-aggressive MCF-7 breast cancer cells.
BACKGROUND
In breast cancer cells, the metastatic cell state is strongly correlated to epithelial-to-mesenchymal transition (EMT) and the CD44+/CD24- stem cell phenotype. However, the MCF-7 cell line, which has a luminal epithelial-like phenotype and lacks a CD44+/CD24- subpopulation, has rare cell populations with higher Matrigel invasive ability. Thus, what are the potentially important differences between invasive and non-invasive breast cancer cells, and are the differences related to EMT or CD44/CD24 expression?


METHODS
Throughout the sequential selection process using Matrigel, we obtained MCF-7-14 cells of opposite migratory and invasive capabilities from MCF-7 cells. Comparative analysis of epithelial and mesenchymal marker expression was performed between parental MCF-7, selected MCF-7-14, and aggressive mesenchymal MDA-MB-231 cells. Furthermore, using microarray expression profiles of these cells, we selected differentially expressed genes for their invasive potential, and performed pathway and network analysis to identify a set of interesting genes, which were evaluated by RT-PCR, flow cytometry or function-blocking antibody treatment.


RESULTS
MCF-7-14 cells had enhanced migratory and invasive ability compared with MCF-7 cells. Although MCF-7-14 cells, similar to MCF-7 cells, expressed E-cadherin but neither vimentin nor fibronectin, beta-catenin was expressed not only on the cell membrane but also in the nucleus. Furthermore, using gene expression profiles of MCF-7, MCF-7-14 and MDA-MB-231 cells, we demonstrated that MCF-7-14 cells have alterations in signaling pathways regulating cell migration and identified a set of genes (PIK3R1, SOCS2, BMP7, CD44 and CD24). Interestingly, MCF-7-14 and its invasive clone CL6 cells displayed increased CD44 expression and downregulated CD24 expression compared with MCF-7 cells. Anti-CD44 antibody treatment significantly decreased cell migration and invasion in both MCF-7-14 and MCF-7-14 CL6 cells as well as MDA-MB-231 cells.


CONCLUSIONS
MCF-7-14 cells are a novel model for breast cancer metastasis without requiring constitutive EMT and are categorized as a "metastable phenotype", which can be distinguished from both epithelial and mesenchymal cells. The alterations and characteristics of MCF-7-14 cells, especially nuclear beta-catenin and CD44 upregulation, may characterize invasive cell populations in breast cancer.
Uchino
Masahiro
M
Neo-Technology Development Division, Bio Matrix Research Inc., 105 Higashifukai, Nagareyama, Chiba 270-0101, Japan. uchino@biomatrix.co.jp
Kojima
Hiroko
H
Wada
Kenta
K
Imada
Mika
M
Onoda
Fumitoshi
F
Satofuka
Hiroyuki
H
Utsugi
Takahiko
T
Murakami
Yasufumi
Y
D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D051927:CD24 Antigen / Q000378:metabolism; D002448:Cell Adhesion; D002465:Cell Movement; D002467:Cell Nucleus / Q000378:metabolism*; D049109:Cell Proliferation; D004847:Epithelial Cells / Q000378:metabolism / Q000473:pathology; D005260:Female; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D020869:Gene Expression Profiling; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism*; D008648:Mesoderm / Q000378:metabolism / Q000473:pathology; D009361:Neoplasm Invasiveness; D020411:Oligonucleotide Array Sequence Analysis; D010190:Pancreatic Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000556:secondary*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D014945:Wound Healing; D023041:Xenograft Model Antitumor Assays; D051176:beta Catenin / Q000378:metabolism
D014408:Biomarkers, Tumor; D051927:CD24 Antigen; C495270:CTNNB1 protein, human; D018960:Hyaluronan Receptors; D012333:RNA, Messenger; D051176:beta Catenin; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins