Malignant progression of an SV40-transformed human epidermal keratinocyte cell line

Abstract

Summary Human foetal keratinocytes were transfected with a recombinant plasmid (pSV6-1) which contained an origin defective SV40 genome. The resulting transformed cell line had many properties in common with previously described SV40-transformed keratinocytes, including expression of simple epithelialtype keratins. It was non-tumourigenic in nude mice at early passages, forming small benign cysts, however, after approximately 46 in vitro passages, these transformed keratinocytes formed invasive squamous cell carcinomas in athymic nude mice. Several in vitro changes were associated with this acquisition of tumourigenicity (a) an alteration in cellular morphology, (b) development of a cytogenetically marked clone and (c) loss of cell surface fibronectin. The loss of fibronectin was also observed in vivo; cysts formed by SV6-1 Bam/HFK produced human fibronectin whereas tumours did not, although both tumours and cysts were laminin-and keratin-positive. These results indicate that the spontaneous development of secondary events in immortalised human cells may lead to the acquisition of a malignant phenotype.

Tumor Progression of Skin Carcinoma Cells in Vivo Promoted by Clonal Selection, Mutagenesis, and Autocrine Growth Regulation by Granulocyte Colony-Stimulating Factor and Granulocyte-Macrophage Colony-Stimulating Factor

“…56,57 There are a few examples in the literature describing an in vitro progression to a more malignant phenotype. 19,24 However, this in vitro progression either occurred in simian virus-40 immortalized cell lines, 19 that are characterized by an increased genetic instability, or required the action of an additional mutagen such as radiation. 24 We have found a decrease in tumor progression in vitro for the HaCaT-ras model system and these findings are in agreement with reports demonstrating an in vitro selection against tumorigenic subpopulations in breast adenocarcinomas, 58 SCCs arising in Bowen' disease, 59 and prostate carcinomas.…”

Characterisation of human thyroid epithelial cells immortalised in vitro by simian virus 40 DNA transfection

“…(Kahn et al, 1983). It is not clear why SV40-transformed human epithelial cells are usually non-tumorigenic in nude mice (an exception being late-passage SV40-transformed keratinocytes; described by Brown & Gallimore, 1987), while SV40-transformed rodent epithelial cells, such as rat hepatocytes (Woodworth et al, 1988), are often strongly tumorigenic (Freedman & Shin, 1978). The fact that some SV40-transformed human epithelial cell lines, such as the 184A1 mammary line (Clark et al, 1988) and the HBL-100 mammary line (Saint-Ruf et al, 1988), can be converted to tumorigenicity by ras oncogenes (activated Ha-ras and Ki-ras respectively) strongly suggests that the parent lines are indeed incompletely transformed.…”

“…Both of the thyroid cell lines (HTori-3 and HTori-5) subjected to chromosome analysis were aneuploid, which is to be expected after transformation by SV40. Periodic re-analysis in the future will be required to determine the stability of the genomes, but evidence from most other studies suggests that aberrations are likely to increase with progressive generations (Meisner et al, 1988;Brown & Gallimore, 1987). The apparent integration of only a single copy of SV40 plasmid in each of the established cells lines analysed corresponds with previous observations in other human cell types (Neufeld et al, 1987;Canaani et al, 1986;Murnane et al, 1985) and it seems that immortalisation is also favoured by a low copy number of integrated SV40 DNA in cells infected with wildtype SV40 (Kucherlapati et al, 1978;Hara & Kaji, 1987).…”

“…Human cells are recognised to be more resistant than rodent cells to in vitro transformation, whether spontaneous or induced by various means (DiPaolo, 1983;Sager, 1984) and epithelial cells of human origin appear to be even more resistant than mesenchymal derivatives such as fibroblasts, as illustrated by studies of SV40-induced transformation (Girardi et al, 1965;Chang, 1986). Most success has been achieved with human mammary epithelium and keratinocytes (Brown & Gallimore, 1987;Taylor-Papadimitrou et al, 1982) transformed by SV40. Transformation of human brochial epithelium with SV40 large T (Brash et al, 1987;Reddel et al, 1988), of human uroepithelial cells (Christian et al, 1987) and fetal colonic epithelial cells (Berry et al, 1988) with SV40, and human thyroid epithelium with SV40 early region by electroporation (Whitley et al, 1987) has also been reported.…”

In vitro culture of human foetal colonic epithelial cells and their transformation with origin minus SV40 DNA

“…None of these cell lines was tumorigenic in athymic nude mice, which is consistent with many previous reports of SV40 trans- formation of human epithelial cells. Generally, SV40 transformed human epithelial cells rarely escape 'crisis' and progress to become tumorigenic (Chang, 1986;Brown & Gallimore, 1987), suggesting that they are incompletely transformed and that other events are necessary for the full expression of the malignant phenotype. It is possible in those cells that do become tumorigenic that the continual rearrangement of chromosomes that begins soon after transformation with SV40 (Chang, 1986) could activate cellular mechanisms that in rare circumstances may result in a unique variant escaping crisis and progressing in vitro.…”