Correlation of growth capacity of cells in hard agarose with successful transfection by the activated c-Ha-ras oncogene and in vivo proliferative capacity at metastatic sites.
The purpose of this study was to determine whether the degree of anchorage-independent growth of rodent or human cells in increasing concentrations of agarose correlated with successful transfection of the cells with an activated c-Ha-ras oncogene and tumorigenicity in nude mice. NIH 3T3 cells, C3H 10T1/2 fibroblasts, four clones of the murine K-1735 melanoma with different metastatic capacities and the TE85 human osteogenic sarcoma line were transfected with plasmids containing the 6.6-kilobase BamHI fragment of the mutant human c-Ha-ras gene and the neo gene, which confers resistance to neomycin (pSV2-neoEJ). Cells transfected with pSV2-neo, a plasmid containing the neo gene, served as controls. Cells from parental or transfected lines (selected by Geneticin) were plated into medium containing 0.3%, 0.6% 0.9%, or 1.2% agarose. These cells were also injected subcutaneously and intravenously into nude mice. The production of tumor cell colonies in dense agarose (greater than or equal to 0.6%) correlated with successful transfection with pSV2-neoEJ and production of experimental metastases in the lung of nude mice. We conclude that the degree of anchorage-independent growth of cells predicts successful transfection with activated c-Ha-ras oncogene and tumorigenic behavior in vivo. Thus this technique may be useful for the detection of cells transfected with transforming oncogenes.
Fidler
I J
IJ
Department of Cell Biology, University of Texas M. D. Anderson Cancer Center, Houston 77030.
Li
L M
LM
Ananthaswamy
H N
HN
Esumi
N
N
Radinsky
R
R
Price
J E
JE
D000818:Animals; D015139:Blotting, Southern; D002455:Cell Division; D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification; D011905:Genes, ras; D006801:Humans; D008546:Melanoma, Experimental / Q000473:pathology*; D051379:Mice; D008819:Mice, Nude; D009362:Neoplasm Metastasis; D012516:Osteosarcoma / Q000473:pathology*; D010957:Plasmids; D012685:Sepharose; D014162:Transfection
D004273:DNA, Neoplasm; D012685:Sepharose