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The characterization of cell line crl-2335 as a Basal-like breast carcinoma model.

ABSTRACT: Basal-like breast cancer has been reported to be the most aggressive and deadly carcinoma sub-type. Patients diagnosed with this subtype have a less than 50% five-year survival. In addition, many studies have reported that this sub-type is more prevalent in specific ethnic groups and is believed to be a key factor that drives certain ethnic disparities in mortality. In order to effectively study this sub-type and determine unique gene expression and biochemical pathways which sustain this cancer's growth, we sought to identify human breast cancer cell lines that represent a model for the basal-like subtype. Here, we report our findings which indicate the African American cell line CRL-2335 is a true representative of basal-like breast carcinoma.

SUBMITTER: Hill Neves LA

PROVIDER: S-EPMC3091405 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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The characterization of cell line crl-2335 as a Basal-like breast carcinoma model.

Hill Neves Lori A LA Ingram Lishann L Davis Melissa B MB

Breast cancer : basic and clinical research 20110427

Basal-like breast cancer has been reported to be the most aggressive and deadly carcinoma sub-type. Patients diagnosed with this subtype have a less than 50% five-year survival. In addition, many studies have reported that this sub-type is more prevalent in specific ethnic groups and is believed to be a key factor that drives certain ethnic disparities in mortality. In order to effectively study this sub-type and determine unique gene expression and biochemical pathways which sustain this cancer ...[more]

PMID: 21603256

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Project description:The phenotypic diversity of breast carcinoma may be explained by the existence of a sub-population of breast cancer cells, endowed with stem cell-like properties and gene expression profiles, able to differentiate along different pathways. A stem cell-like population of CD44(+)CD24(-/low) breast cancer cells was originally identified using cells from metastatic pleural effusions of breast carcinoma patients. We have previously reported that upon in vitro culture as mammospheres under stem cell-like conditions, human MA-11 breast carcinoma cells acquired increased tumorigenicity and lost CD24 expression compared with the parental cell line. We now report that upon passage of MA-11 mammospheres into serum-supplemented cultures, CD24 expression was restored; the rapid increase in CD24 expression was consistent with up-regulation of the antigen, and not with in vitro selection of CD24(+) cells. In tumors derived from subcutaneous injection of MA-11 mammospheres in athymic nude mice, 76.1+/-9.7% of cells expressed CD24, vs. 0.5+/-1% in MA-11 cells dissociated from mammospheres before injection. The tumorigenicity of sorted CD44(+)CD24(-) and CD44(+)CD24(high) MA-11 cells was equal. Single cell-sorted CD24(-) and CD24(high) MA-11 gave rise in vitro to cell populations with heterogeneous CD24 expression. Also, subcutaneous tumors derived from sorted CD24(-) sub-populations and single-cell clones had levels of CD24 expression similar to the unsorted cells. To investigate whether the high expression of CD24 contributed to the tumorigenic potential of MA-11 cells, we silenced CD24 by shRNA. CD24 silencing (95%) resulted in no difference in tumorigenicity upon s.c. injection in athymic nude mice compared with mock-transduced MA-11 cells. Since CD24 silencing was maintained in vivo, our data suggest that the level of expression of CD24 is associated with but does not contribute to tumorigenicity. We then compared the molecular profile of the mammospheres with the adherent cell fraction. Gene expression profiling revealed that the increased tumorigenicity of MA-11 mammospheres was associated with changes in 10 signal transduction pathways, including MAP kinase, Notch and Wnt, and increased expression of aldehyde dehydrogenase, a cancer-initiating cell-associated marker. Our data demonstrate that (i) the level of CD24 expression is neither a stable feature of mammosphere-forming cells nor confers tumorigenic potential to MA-11 cells; (ii) cancer-initiating cell-enriched MA-11 mammospheres have activated specific signal transduction pathways, potential targets for anti-breast cancer therapy.

Project description:The human breast is composed of terminal duct lobular units (TDLUs) that are surrounded by stroma. In the TDLUs, basement membrane separates the stroma from the epithelial compartment, which is divided into an inner layer of luminal epithelial cells and an outer layer of myoepithelial cells. Stem cells and progenitor cells also reside within the epithelium and drive a continuous cycle of gland remodelling that occurs throughout the reproductive period. D492 is an epithelial cell line originally isolated from the stem cell population of the breast and generates both luminal and myoepithelial cells in culture. When D492 cells are embedded into 3D reconstituted basement membrane matrix (3D-rBM) they form branching colonies mimicking the TDLUs of the breast, thereby providing a well-suited in vitro model for studies on branching morphogenesis and breast development. Peroxidasin (PXDN) is a heme-containing peroxidase that crosslinks collagen IV with the formation of sulfilimine bonds. Previous studies indicate that PXDN plays an integral role in basement membrane stabilisation by crosslinking collagen IV and as such contributes to epithelial integrity. Although PXDN has been linked to fibrosis and cancer in some organs there is limited information on its role in development, including in the breast. In this study, we demonstrate expression of PXDN in breast epithelium and stroma and apply the D492 cell line to investigate the role of PXDN in cell differentiation and branching morphogenesis in the human breast. Overexpression of PXDN induced basal phenotype in D492 cells, loss of plasticity and inhibition of epithelial-to-mesenchymal transition as is displayed by complete inhibition of branching morphogenesis in 3D culture. This is supported by results from RNA-sequencing which show significant enrichment in genes involved in epithelial differentiation along with significant negative enrichment of EMT factors. Taken together, we provide evidence for a novel role of PXDN in breast epithelial differentiation and mammary gland development.

Dataset Information

The characterization of cell line crl-2335 as a Basal-like breast carcinoma model.

Publications

The characterization of cell line crl-2335 as a Basal-like breast carcinoma model.

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