A Human Breast Cell Model of Preinvasive to Invasive Transition

Abstract

A crucial step in human breast cancer progression is the acquisition of invasiveness. There is a distinct lack of human cell culture models to study the transition from preinvasive to invasive phenotype as it may occur ''spontaneously'' in vivo. To delineate molecular alterations important for this transition, we isolated human breast epithelial cell lines that showed partial loss of tissue polarity in three-dimensional reconstituted basement membrane cultures. These cells remained noninvasive; however, unlike their nonmalignant counterparts, they exhibited a high propensity to acquire invasiveness through basement membrane in culture. The genomic aberrations and gene expression profiles of the cells in this model showed a high degree of similarity to primary breast tumor profiles. The xenograft tumors formed by the cell lines in three different microenvironments in nude mice displayed metaplastic phenotypes, including squamous and basal characteristics, with invasive cells exhibiting features of higher-grade tumors. To find functionally significant changes in transition from preinvasive to invasive phenotype, we performed attribute profile clustering analysis on the list of genes differentially expressed between preinvasive and invasive cells. We found integral membrane proteins, transcription factors, kinases, transport molecules, and chemokines to be highly represented. In addition, expression of matrix metalloproteinases MMP9, MMP13, MMP15, and MMP17 was up-regulated in the invasive cells. Using small interfering RNA-based approaches, we found these MMPs to be required for the invasive phenotype. This model provides a new tool for dissection of mechanisms by which preinvasive breast cells could acquire invasiveness in a metaplastic context. [Cancer Res 2008;68(5):1378-87]

Characterization of the Dimerization Interface of Membrane Type 4 (MT4)-Matrix Metalloproteinase

“…Therefore, we wished to determine whether MT4-MMP also generates species consistent with dimerization. Previous studies demonstrated expression of MT4-MMP protein in cell lines and eosinophils (10,19,21,31). However, these studies could not be used as a reference, because the full spectrum of MT4-MMP species at reducing and nonreducing conditions was not shown (10,19) or protein expression was not displayed by immunoblotting (21,31).…”

“…Chabottaux et al (18) and Hotary et al (20) failed to demonstrate an invasion promoting activity of recombinant MT4-MMP expressed in human breast cancer MDA-MB-231 and monkey kidney COS cells, respectively. In contrast, Rizki et al (21) and Hegedus et al (31) showed that down-regulation of MT4-MMP by siRNA in human breast cancer T4-2 and MDA-MB-231 cells, respectively, inhibited Matrigel invasion. Likewise, Huang et al (19) found that invasion through Matrigel, but not cell migration, of the tongue squamous cell carcinoma SAS cell line was enhanced by MT4-MMP expression.…”

“…In addition, MT4-MMP did not cleave the ␣1-proteinase inhibitor (data not shown), a known substrate of MT6-MMP (6,38). Previous studies indicated a role for MT4-MMP in supporting in vitro cell invasion of Matrigel (19,21,31). Therefore, we examined whether MT4-MMP (wild type and mutants) supported the migration and invasion of MDCK cells using established in vitro assays.…”

14-3-3σ stabilizes a complex of soluble actin and intermediate filament to enable breast tumor invasion

“…HMT-3522 S1, S2, S3C, and T4-2 were maintained in tissue culture monolayers or in 3D (Matrigel) cultures as described previously (4). The MCF10 series was kindly provided by Fred Miller (Karmanos Cancer Institute, Detroit, MI) and cultured as recommended (8).…”

“…5 and 6), encompassing nonmalignant (S1), premalignant (S2), preinvasive (S3), and tumorigenic (T4-2) cell populations, and have found this model recapitulates a number of features of BLBC progression. Xenograft tumors formed from T4-2, a cell line that became malignant without activation of exogenous oncogenes and after prolong cultivation in culture (5,6), display many cardinal features of human BLBCs, including high histological grade; positivity for epidermal growth factor receptor (EGFR) and basal cytokeratins K5/K14; negativity for HER2, estrogen receptor (ER), and progesterone receptor (PR); a high Ki-67-proliferative index; and aggressive invasion into surrounding tissues (4). Another isogenic cell culture model of breast cancer progression, the MCF10 series, is thought also to recapitulate features of basal-like malignant progression albeit with exogenous oncogene addition (7)(8)(9).…”