Background: We have developed and optimized a feeder layer method for cultivation of normal human mammary luminal cells and found it suitable for establishing more than 150 primary cultures from individual human breast carcinomas. Here, we investigated if the malignant cells that are in situ additionally characterized by increased numbers of proto-oncogenes can be traced by the FISH method in the ex vivo-derived cultures.
Materials and methods: Paraffin sections from 9 tumors with derived cell cultures kept frozen in our cell bank were screened by FISH for cyclin D1 (CCND1) and c-erb-B2 (HER2/neu) proto-oncogene signal numbers.
Results: In 6 tumors (5 primary tumors, 1 cutaneous metastasis), increased numbers of FISH signals were found in 55-99% of cells. Then, the relevant cell cultures were FISH screened; in cell populations maintained for up to 2-6 passages in vitro the incidence of cells with increased FISH signals was found to be low (2-16%). Moreover, the cells with multiplied signals that survived more than one passage in vitro were evidently unable to divide further. However, in all 6 tumors at least a small fraction of cells displaying only two signals of CCND1 or HER2/neu genes was identified directly in invasive tumor structures in the vicinity of cells with multiple signals.
Conclusion: Our findings suggest that these invasive tumor cells displaying only two proto-oncogene signals were most probably involved in the initiation and propagation of ex vivo tumor-derived primary cell cultures.