De novo expression of α_vβ₆ integrin by myoepithelial cells in ductal carcinoma in situ may be an important marker of disease progression

Myoepithelial cells (MEC) are essential to the maintenance of normal breast function, and loss of normal MEC function is commonly associated with breast cancer. Most established invasive breast carcinomas develop through an in situ phase known as ductal carcinoma in situ (DCIS). We have identified up-regulation of β₆ integrin on MEC in a subset of DCIS. Normal MEC exhibit potent tumour suppressor function, but it is not clear whether this is compromised in DCIS. The aim of the present study is to investigate the effect of β₆ expression on myoepithelial tumour suppressor function.

Immunohistochemical analysis of DCIS of different grades with and without an invasive breast cancer was carried out to determine the β₆ status. For in vitro studies magnetic bead sorting was used to isolate a pure normal-like myoepithelial cell line from the immortalised 1089 cell line (N-1089 MEC). These were used to generate overexpressing myoepithelial cells (DCIS-modified MEC) by β₆ retroviral transduction. The lines were characterised by immuno-fluorescence and flow cytometry, and the tumour suppressor function of both lines was compared with primary normal and DCIS MEC in coculture with breast cancer cell lines.

Analysis of a series of primary DCIS tissues (n > 400) demonstrated induction of β₆ in MEC in a subset of cases, predominantly high grade. Upregulation was almost universal in DCIS associated with invasive disease. Initial characterisation of the 1089 line revealed a mixed phenotype from which pure MEC were selected on the basis of β₄ integrin. This population exhibited all characteristic myoepithelial markers. Coculture assays demonstrated that N-1089 MEC could produce the same tumour suppressor effect as primary MEC, leading to significant reduction in tumour invasion (P < 0.001) and proliferation (P < 0.001). N-1089 MEC transduced with α_vβ₆ (DCIS-1089 MEC) demonstrated enhanced binding and migration to the β₆ ligand LAP, and activation of a transforming growth factor beta reporter, indicating that the α_vβ₆ was functional. Whilst primary DCIS MEC showed loss of suppressor function (P < 0.05), DCIS-1089 MEC exhibit altered behaviour with a more migratory phenotype then the normal counterpart, but at least some of the tumour-suppressor function was maintained.

We have shown that MEC exhibit an altered phenotype in DCIS with de novo expression of α_vβ₆. We have generated normal-like cell lines that exhibit all the characteristics of primary MEC and recapitulate primary MEC tumour suppressor function. Primary DCIS MEC show loss of suppressor function whereas β₆-overexpressing MEC (which resemble DCIS-like MEC) promote or suppress breast cancer cell invasion in a cell-type-specific manner. These findings suggest that changes in MEC during DCIS may influence disease progression, and these cell lines provide a powerful model to study further the mechanisms involved.