Summary
In this study, we describe the karyotypic changes associated with the spontaneous acquisition of tumorigenicity in an immortalized tumor bronchial cell line. Neoplastic transformation of the NL20 human bronchial epithelial cell line occurred after 3 yr in culture, and was associated with loss of chromosome 18 together with acquisition of multiple copies of 9q21.2→34. The nontumorigenic NL20 cell line had been established by transfection of human bronchial epithelial cells with the SV40 T antigen, and had retained a relatively stable karyotype after the first 32 passages in vitro. However, when cells from p184 were inoculated into nude mice, a transplantable tumor was obtained that was derived from a minor clone present in this otherwise stable line. Subsequent passaging of the NL20 cells in vitro did not yield further tumors, and the minor clone from which the tumorigenic NL20T cell line derived was no longer evident in NL20 cells by Passage 205. Furthermore, the original tumorigenic NL20T cells lost the neoplastic phenotype after 25 passages in vitro and reverted to the nontumorigenic karyotype observed at p189. In contrast to the loss of the tumorigenic phenotype and karyotype, which occurred with in vitro passaging of the original tumor, when the NL20T cells were passaged in other nude mice, they continued to give rise to tumors with sevenfold amplifications of 9q sequences and loss of chromosome 18, and cells from the secondary tumors (NL20T-A cells) have maintained a stable karyotype and remain tumorigenic even after 64 passages in vitro. A mixture of 10% tumorigenic NL20T-A and 90% nontumorigenic NL20 cells formed tumors in athymic nude mice when cultured in vitro on fibronectin, but not on plastic; cytogenetic analysis demonstrated that the tumors and cell cultures were composed of tumorigenic NL20T-A cells, whereas cytogenetic analysis of cells cultured on plastic were identical to the nontumorigenic NL20 cells. These data support the hypothesis that neoplastic transformation in our original cell line arose from in vivo selection of a small mutant clone, which had arisen in culture and was subsequently selected in vivo but was lost with in vitro culture.
Similar content being viewed by others
References
Band, V.; Zachowski, D.; Kulesa, V., et al. Human papilloma virus DNAs immortalize normal human mammary epithelial cells and reduce their growth factor requirement. Proc. Natl. Acad. Sci. USA 87:463–467; 1990.
Bookland, E. A.; Swaminathan, S.; Oyasu, R., et al. Tumorigenic transformation and neoplastic progression of human uroepithelial cells after exposure in vitro to 4-aminobiphenyl or its metabolites. Cancer Res. 52:1606–1614; 1992.
Brown, K.; Gallimore, P. Malignant progression of an SV40-transformed human epidermic keratinocyte cell line. Br. J. Cancer 56:545–554; 1987.
Chang, S. In vitro transformation of human epithelial cells. Biochim. Biophys. Acta 823:161–194; 1986.
Chang, S.; Keen, J.; Lane, E., et al. Establishment and characterization of SV40-transformed human breast epithelial cell lines. Cancer Res. 42:2040–2053; 1982.
Christian, B. J.; Loretz, L. J.; Oberley, T. D., et al. Characterization of human uroepithelial cells immortalized in vitro by simian virus 40. Cancer Res. 47:6066–6073; 1987.
Han, K.; Lee, W.; Harris, C. P., et al. Quantifying chromosome changes and lineage involvement in myelodysplastic syndrome (MDS) using fluorescent in situ hybridization (FISH). Leukemia 8:81–86; 1994.
Kaighn, M. E.; Reddel, R. R.; Lechner, J. E., et al. Transformation of human neonatal prostate epithelial cells by strontium phosphate transfection with a plasmid containing SV40 early region genes. Cancer Res. 49:3050–3056; 1989.
Klein-Szanto, A.; Iizasa, T.; Momiki, S., et al. A tobacco-specific N-nitrosamine or cigarette smoke condensate causes neoplastic transformation of xenotransplanted human bronchial epithelial cells. Proc. Natl. Acad. Sci. USA 89:6693–6697; 1992.
Koprowski, H.; Croce, C. Tumorigenicity of Simian virus 40-transformed human cells and mouse-human hybrids in nude mice. Proc. Natl. Acad. Sci. USA 74:1142–1146; 1977.
Meisner, L. F.; Wu, S.-Q.; Christian, B. J., et al. Cytogenetic instability with balanced chromosome changes in an SV40 transformed human uroepithelial cell line. Cancer Res. 48:3215–3220; 1988.
Milo, G.; Shuler, C.; Stoner, G., et al. Conversion of premalignant human cells to tumorigenic cells by methylmethane sulfonate and methylnitrosoguanidine. Cell Biol. Toxicol. 8:193–205; 1992.
Pfeifer, A. M.; Mark, G.; Malan, S. L., et al. Cooperation of c-raf-1 and c-myc protooncogenes in the neoplastic transformation of simian virus 40 large tumor antigen-immortalized human bronchial epithelial cells. Proc. Natl. Acad. Sci. USA 86:10075–10079; 1989.
Reddel, R.; Ke, Y.; Gerwin, B., et al. Transformation of human bronchial epithelial cells by infection with SV40 or adenovirus-12 SV40 hybrid virus, or transfection via strontium phosphate coprecipitation with a plasmid containing SV40 early region genes. Cancer Res. 48:1904–1909; 1988.
Reddel, R.; Ke, Y.; Kaighn, E., et al. Human bronchial epithelial cells neoplastically transformed by v-Ki-ras: altered response to inducers of terminal squamous differentiation. Oncogene Research 3:401–408; 1988.
Reddel, R.; Salghetti, S.; Willey, J., et al. Development of tumorigenicity in simian virus 40-immortalized human bronchial epithelial cell lines. Cancer Res. 53:985–991; 1993.
Reznikoff, C.; Loretz, L.; Christian, B., et al. Neoplastic transformation of SV40-immortalized human urinary tract epithelial cells by in vitro exposure to 3-methyl-cholanthrene. Carcinogenesis 9:1427–1436; 1988.
Rhim, J.; Fujita, J.; Arnstein, P., et al. Neoplastic conversion of human keratinocytes by adenovirus 12-SV40 virus and chemical carcinogens. Science 232:385–388; 1986.
Schiller, J. H.; Bittner, G. Loss of the tumorigenic phenotype with in vitro but not in vivo, passaging of a novel series of human bronchial epithelial cell lines: possible role of an α5/β1 integrin/fibronectin interaction. Cancer Res. 55:6215–6221; 1996.
Schiller, J. H.; Bittner, G.; Oberley, T. D., et al. Establishment and characterization of a SV40 T-antigen immortalized human bronchial epithelial cell line. In Vitro Cell. Dev. Biol. 28:461–464; 1992.
Schiller, J.; Sabatini, L.; Bittner, G., et al. Phenotypic, molecular, and genetic characterization of transformed human bronchial epithelial cell strains. Int. J. Oncol. 4:461–470; 1993.
Stoner, G. D.; Kaighn, M. E.; Reddel, R. R., et al. Establishment and characterization of SV40 T-antigen immortalized human esophageal epithelial cells. Cancer Res. 51:365–371; 1991.
Willey, J.; Broussound, A.; Sleemi, A., et al. Immortalization of normal human bronchial epithelial cells by human papillomaviruses 16 or 18. Cancer Res. 51:5370–5377; 1991.
Wu, S. Q.; Voelkerding, K. V.; Sabatini, L., et al. Extensive amplification of bcr/abl fusion genes clustered on three marker chromosomes in human leukemic cell line K-562. Leukemia 9:858–862; 1995.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Schiller, J.H., Bittner, G., Wu, SQ. et al. Karyotypic changes associated with spontaneous acquisition and loss of tumorigenicity in a human transformed bronchial epithelial cell line: Evidence for In vivo selection of transformed clones. In Vitro Cell.Dev.Biol.-Animal 34, 283–289 (1998). https://doi.org/10.1007/s11626-998-0004-2
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/s11626-998-0004-2