Experimental Model for human proliferative Breast Disease

Recently, we established a new and unique model of early human breast cancer progression. This model, which is called “MCFIOAT”, consists of preneoplastic human breast epithelial cells that are able to grow in immune-deficient mice where they undergo a sequence of progressive histological changes culminating in cases of frank neoplasia in about 25% of the animals (Miller et al., 1993; Dawson et al., 1996).

MCFlOAneoT is a cell line derived from a normal human breast epithehal hne, which spontaneously immortalized (Soule et al., 1990) and was then transfected with mutated H-ras (Basolo et al., 1991). MCFlOAneoT is a heterogeneous mixture containing a subpopulation that is a multipotent human breast epithelial stem cell that can be selected in vivo by its ability to form persistent lesions in nude/beige mice (Miller et al., 1993). Cell lines established from lesions after serial transplantation into nude/beige mice (MCF10AT# where # represents transplant generation) show more severe atypia at earlier times post-transplantation than does MCFlOAneoT (Dawson et al., 1996). Although ER can not be detected (by Abbot antibody) on cells in xenograft lesions, Dr. P.V.M. Shekhar has shown that the ras- transfected MCFIOAT cells (but not MCFIOA) do express functional ER (Manuscript submitted).

The reproducible establishment of representative stages in early breast cancer progression from the MCF1 OAT model offers an unprecedented opportunity to analyze critical events in human breast carcinogenesis. Normal ducts form initially but become proliferative and sporadically progress to carcinoma in some cases. The ducts formed in xenografts are bilayered, being composed of both myoepithelial and luminal epithelial layers. These normal structures may persist for months in immune-deficient mice.

The MCFIOAT model clearly is a precursor cell line that forms all intermediate stages of prohferative breast disease. Results with our model strengthen the perception of progression from hyperplasia to atypical hyperplasia to invasive breast carcinoma.

We confirmed the invasive phenotype in the MCFIOAT lesions classified as invasive carcinoma by demonstrating the loss of basement membrane ufilizing a silver stain method (Tait et al., 1996a). Unlike the MCFlOAneoT (i.e., ras- transfected cells prior to in vivo selection), all variants derived from xenografts formed intermediate lesions that resemble the lesions seen in human PBD including (PDWA), atypical hyperplasia, and DCIS (Dawson et al., 1996).

It is a widely held point of view that mutations in differentiated epithelial cells are of little importance due to limited growth potential of such cells and that cancer results only from mutations in stem cells. Indeed, we have found that MCFIOAT lines contain stem cells able to differentiate in xenografts into myoepithelial as well as luminal epithelial cells (Tait et al., 1996b). Although ducts formed by the stem cell appear to be normal in most respects, cytokeratin differentiation is abnormal.

MCFIOAT cells express both cytokeratin 14 and cytokeratin 18 (Pauley et al., 1996), a pattern consistent with a stem cell able to generate both cytokeratin 14-expressing myoepithelial cells and cytokeratin 18-expressing luminal cells (Taylor -Papadimitriou et al., 1989). However, neither myoepithelial nor luminal epithelial cells in the xenograft lesions consistentiy express either cytokeratin 14 or 18 (not shown).

MCFIOAT stem cells are unique, differing from previously described human breast and rat mammary stem cells in their capacity to produce organized ductular structures with myoepithelium properly oriented between a basement membrane and the luminal epithelium in situ in xenografts. Although our failure to consistentiy detect cytokeratins 14 and 18 in situ may be due to technical difficulties with the specific antibodies used, the antibodies did stain the appropriate cell layers in control, formalin-fixed, paraffin-embedded human breast tissue. The failure of MCFIOAT cells to differentiate fully in xenografts may be critical to their preneoplastic nature.

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