Melphalan sensitivity as a function of progressive metastatic growth in two subpopulations of a mouse mammary tumour

Abstract

Summary In order to examine in detail the sensitivity to chemotherapy of tumour cells at various organ sites and at various stages of metastasis, we have used a series of cell lines, all selected from sister subpopulations derived from a single mouse mammary tumour, which can be distinguished and quantitated from normal cells and from each other through growth in selective medium. For the studies described here, we used two lines, 4T07 and 66FAR, which will form colonies in vitro in medium containing 60 iM 6-thioguanine Material and methods MiceMale BALB/c mice 8 to 12 weeks old were produced in our animal colony, which was established by caesarean derivation of a litter of mice from BALB/cfC3H parents obtained from the Cancer Research Laboratory, Berkeley, CA. Tumour cell lines used in the experiments described here grow and respond to chemotherapy equally well in male and female mice.Tumour cell lines Tumour subpopulation lines 66 and 410.4 were isolated from a single spontaneously arising mammary tumour from a BALB/cfC3H mouse (Miller et al., 1983;Dexter et al., 1978). Line 66FAR is a diaminopurine (and fluoradenine) -resistant variant of line 66 obtained by stepwise selection in increasing concentrations of diaminopurine, followed by 5-fluoroadenine. Line 4T07 is a thioguanine-, ouabain-resistant variant of line 410.4 (Miller et al., 1987). Doubling times for these cultures in vitro were approximately 15 h, 18 h, and 13 h for lines 4T07, 66FAR, and 66, respectively (n = 3). Plating efficiencies averaged 64% (n = 8 cultures per cell line) and were not significantly different for the three lines.Cell culture Cells were grown in monolayer in DME-10, DME supplemented with 2.5% foetal bovine serum, 7.5% calf serum, 1 mM mixed non-essential amino acids, and 2 mM L-glutamine. Cultures were split and recultured twice a week, using 0.25% trypsin plus 0.05% EDTA in Hanks buffered salt solution to remove cells from the tissue culture flasks. Experimental metastasisCells from culture were suspended with trypsin/EDTA, rinsed once, and suspended in DME-10. Cell suspensions (usually 2.5 x 106 ml-') were injected intravenously via the lateral tail vein at 0.2 ml/mouse.

Immunotoxicologic effects of cyclosporine on tumor progression in models of squamous cell carcinoma and B-cell lymphoma in C3H mice

“…Studies on the effects of suspect "immunotoxicants on immune surveillance with experimental and spontaneous metastases with transplanted tumor cells have been an important part of immunotoxicity testing for over 20 years (Luster et al, 1988;McCay, 1995;Ng et al, 2010). However, a number of immunosuppressive agents can decrease metastases, e.g., actinomycin D and daunorubicin (Marks et al, 1977), everolimus (Khariwala et al, 2006), 5-flurouracil (Suzuki et al, 1995, and melphalan (Miller et al, 1993), and the results with some agents can be contradictory, e.g., cyclophosphamide can either inhibit or promote metastases (Hill and Stanley, 1977;Strzadala et al, 1985). Further, similar to our findings with CSA, other immunosuppressive agents have been reported to inhibit the growth of primary tumors (e.g., azathioprine [Evans, 1977], busulfan [Tarnowski et al, 1966], everolimus [Khariwala et al, 2006], and serolimus [Boffa et al, 2004]).…”

Azelaic Acid Has Sensitizing Effect in the Chemotherapeutic Treatment of Several Melanoma Cell Lines

“…Our results appear to point to a possible connection between the degree of differentiation (origin, morpholom, ultrastructure, and Dopa oxidase activity) and the mean resistance of each cell line. Other studies have pointed to the relationship between resistance to treatment and the degree of evolution of the lesion from which the cell line was established (Jang and Hill, 1991;Lu et al, 1993;Miller et al, 1993). The control cultures from the B16F10 line showed morphological and ultrastructural characteristics similar to those of well-differentiated melanomas, unlike the SK-MEL-28 and SK-MEL-1 controls, which showed characteristics similar to those of more aggressive and evolved human melanomas (Vicente-Ortega et al, 1995).…”

Development of a novel bi-specific monoclonal antibody approach for tumour targeting

“…The 410.4 cell line has a high malignant potential, metastasizes spontaneously with high frequency (> 80%) to the lungs of mice injected subcutaneously or intravenously, and shows lack of immunogenicity (Miller et al, 1993). The 410.4 cell line was transfected with 3.9 kb full-length MUC1 cDNA, kindly provided by Dr M Hollingsworth (Batra et al, 1991), under the control of the β-actin promoter, in the mammalian expression vector pHb-Apr-1-neo by the calcium phosphate precipitation method (M Hudson et al, manuscript in preparation).…”