1. ¹Instituto de Investigaciones Biomédicas, Alberto Sols, Consejo Superior de Investigaciones Científicas (CSIC), Universidad Autónoma de Madrid (UAM), Arturo Duperier 4, Madrid 28029, Spain
2. ²Cytogenetics Unit, Centro Nacional de Investigaciones Ocológicas (CNIO), Madrid, Spain
3. ³Department of Biology, Facultad de Ciencias, UAM, Madrid, Spain
4. ⁴Department of Pathology, Hospital Universitario de la Princesa, Facultad de Medicina, UAM, Madrid, Spain

Correspondence: M Quintanilla, E-mail: mquintanilla@iib.uam.es

Received 9 December 2004; Revised 2 June 2005; Accepted 6 June 2005; Published online 11 July 2005.

Abstract

In mouse epidermal carcinogenesis, the latest stage of malignant progression involves the transition from squamous cell carcinoma to a highly aggressive type of tumor with spindle morphology. In this work, we have isolated a minor epithelial cell subpopulation (CarC-R) contained in the highly malignant spindle carcinoma cell line CarC. CarC-R exhibited a drastic reduction in tumorigenicity when compared with CarC, but CarC-R-induced tumors were mainly sarcomatoid, although they subsequently reverted to the epithelial phenotype when tumor explants were recultured in vitro. Several single-cell clones with either stable epithelial or fibroblastic phenotypes were isolated from an explanted CarC-R tumor (CarC-RT). All these cell lines contained the same specific point mutation in H-Ras codon 61, but while CarC spindle cells had lost the normal H-Ras allele, it was retained in CarC-R- and CarC-RT-derived cell lines. Furthermore, CarC cells have inactivated p16INK4a and p19INK4a/ARF transcription, while CarC-R and CarC-RT clones expressed p19 mRNA and protein but not p16. Altogether, these results suggest that CarC-R represents a precursor stage to CarC in malignant progression. Spectral karyotyping analysis revealed that CarC-R was highly aneuploid and contained many chromosomal abnormalities. In contrast, CarC had a diploid or tetraploid modal chromosome number and contained a specific T(14;15) translocation in all of the analysed metaphases. The T(14;15) translocation was present in only a minority (1.9%) of CarC-R cells, but it was widely spread in CarC-RT and its derived cell clones, regardless of their epithelial or fibroblastic phenotype, indicating that T(14;15) segregates with malignancy.