A short-term in vitro test for tumour sensitivity to adriamycin based on flow cytometric DNA analysis

Engelholm, Svend A.; Spang‐Thomsen, Mogens; Vindeløv, Lars L.

doi:10.1038/bjc.1983.79

Cited by 18 publications

(12 citation statements)

References 10 publications

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“…Chemotherapy: The drug-used in the experiments was me1 phal an(A1 keran, Wellcome). The doses used were 10 -5, lop2, and 1Op1rng/ml I ; 1.0 ml of Eagles minimum essential medium enriched with 20% fetal bovine serum, antibiotics, and different doses of' melphalan was added to 1.0 ml of cell suspension to produce the nine different final concentrations of melphalan and a cell concentration of 10" cellsiml plus controls (5).…”

Section: Incubationmentioning

confidence: 99%

“…Furthermore, the method does not allow one to test how representative the growing cells are. In search of a faster technique to detect chemosensitivity, a method based on measurements of drug-induced cell cycle perturbations was examined in experimental tumors (5). This in vitro flow cytometry assay was found to be able to detect sensitivity to adriamycin in Ehrlich ascites tumors.…”

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confidence: 99%

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Chemosensitivity of human small cell carcinoma of the lung detected by flow cytometric DNA analysis of drug‐induced cell cycle perturbations in vitro

Engelholm

Spang‐Thomsen

Vindeløv³

et al. 1986

Cytometry

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A method based on detection of drug-induced cell cycle perturbation by flow cytometric DNA analysis has previously been described in Ehrlich ascites tumors as a way to estimate chemosensitivity. The method is extended to test human small-cell carcinoma of the lung. Three tumors with different sensitivities to melphalan in nude mice were used. Tumors were disaggregated by a combined mechanical and enzymatic method and thereafter have incubated with different doses of melphalan. After incubation the cells were plated in vitro on agar, and drug induced cell cycle changes were monitored by flow cytometric DNA analysis. Melphalan produced a dose-related S phase accumulation in the two sensitive tumors, whereas no changes in the cell cycle distribution were found in the resis- In vitro tests, especially the human tumor cloning assay (12,13), have been increasingly used to optimize treatment of malignant diseases andor to test new and more potent non-cross-resistant drugs. However, the human tumor cloning assay is time-consuming and requires 2-3 wk before conclusive results are available. Furthermore, the method does not allow one to test how representative the growing cells are. In search of a faster technique to detect chemosensitivity, a method based on measurements of drug-induced cell cycle perturbations was examined in experimental tumors (5). This in vitro flow cytometry assay was found to be able to detect sensitivity to adriamycin in Ehrlich ascites tumors.The object of the present study was to determine whether the in vitro cell cycle perturbation assay could be extended to test the chemosensitivity of human solid tumors. This demands a valid procedure to disaggregate the solid tumors into single cell suspensions representative of the original tumors. Three heterotransplanted human small-cell carcinomas of the lung (SCCL) were selected for this study because of their different sensitivities to melphalan (9). The tumors were disaggregated by a combined mechanical and enzymatic method (7).Subsequently the cells were incubated in vitro with different doses of melphalan; flow cytometric DNA analysis was used to monitor drug-induced cell cycle pertubations.The results showed that melphalan induced dose-related S phase accumulations in two sensitive tumors, whereas no cell cycle changes were found in a resistant tumor. Furthermore, the size of the S phase accumulation in vitro reflected the previously found in vivo chemosensitivity of the tumors. Thus, the results suggest that the flow cytometry method applied on disaggregated tumor cells may be extended to sensitivity testing of human tumors in general, including screening for new agents. MATERIALS AND METHODS Tumors

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Section: Incubationmentioning

confidence: 99%

mentioning

confidence: 99%

Chemosensitivity of human small cell carcinoma of the lung detected by flow cytometric DNA analysis of drug‐induced cell cycle perturbations in vitro

Engelholm

Spang‐Thomsen

Vindeløv³

et al. 1986

Cytometry

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“…Four of them were small cell carcinomas (SCCL) of the lung (Engelholm et al, 1983, two tumours were breast carcinomas (Brunner & Visfeldt, 1982;Brunner et al, 1983), and one was a malignant melanoma (SpangThomsen et al, 1980 Reinhold (1965). The tumour samples were initially prepared as described above.…”

Section: Tumoursmentioning

confidence: 99%

“…The disaggregated cells were grown on agar by a technique previously described (Engelholm et al, 1983). Aliquots of MEM culture medium containing 103, 5 x 103, and 104 cells were plated in triplicate in 35mm Petri dishes on top of a layer of hardened 0.25% agarose (Difco) medium.…”

Section: Colony Forming Efficiency (Cfe)mentioning

confidence: 99%

Disaggregation of human solid tumours by combined mechanical and enzymatic methods

Engelholm¹,

Spang‐Thomsen²,

Brünner³

et al. 1985

Br J Cancer

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Summary Two combined mechanical and enzymatic disaggregation techniques and a simple mechanical disaggregation procedure were compared. The combined procedures involved a mechanical comminution of the tumour tissue followed by incubation in trypsin. In one method, the tissue was subjected to long-term trypsinization at 40C, and in the other procedure, repeated short-term trypsinization at 37°C was applied. The results were compared in terms of the yield of viable cells, plating efficiency, the ability to produce tumours in nude mice, and DNA distribution as measured by flow cytometry. The combined techniques provided reproducible cell yields of 2-10 x 107 viable cells g-I of tissue, whereas only a small number of tumour cells was produced by the mechanical method. DNA analysis demonstrated that only the long-term trypsinization procedure resulted in a representative cell yield from all the tumours tested.

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“…In human breast cancer, very few data are available on the relation between endocrine treatment-induced tumor growth inhibition and the treatment effect on the tumor cell kinetics (33,34], Such information might prove useful in the treatment of breast cancer, since in this type of tumor, endocrine treat ment is often used in combination with cell cycle phase-specific chemotherapeutics [9], In recent years, flow cytometric DNA analysis has been applied to obtain rapid information on the cell cycle kinetics of tu mors [2,8,20,27], The method provides information on treatment-induced perturba tions in the cell cycle distribution which may be more important for an understanding of the therapeutic response than the tumor cell kinetics per se during unperturbed growth.…”

Section: Introductionmentioning

confidence: 99%

Dose-Dependent Effect of 17β-Estradiol Determined by Growth Curves and Flow Cytometric DNA Analysis of a Human Breast Carcinoma (T61) Grown in Nude Mice

Brünner

Spang‐Thomsen

Vindeløv³

et al. 1985

Pathobiology

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An estrogen and progesterone receptor-positive human breast carcinoma (T61) grown in nude mice was exposed to 1.0, 0.1, 0.01, and 0.001 mg 17β-estradiol. These doses resulted in serum peak concentrations (day 1) of estradiol ranging from 3.5 × 10^-8 to 6.9 × 10^-10M. The effect of the treatment was evaluated using growth curves and flow cytometric DNA analysis. The treatment induced a dose-dependent growth delay and dose-dependent changes in the cell cycle distribution. The cell cycle changes comprised a decrease in the G₁ phase, an accumulation of cells in the S phase, and an increasing fraction of polyploid cells. The results suggest that estradiol induces a dose-dependent cell killing effect in the T61 human breast carcinoma. The correlation between the treatment-induced growth delay and the effect on the cell cycle distribution indicates that the changes in the cell cycle are a reflection of the estradiol-induced cell destruction. Since no tumor growth stimulation could be observed even at very low serum estradiol concentrations, the T61 human breast carcinoma may represent a new aspect in the study of human breast cancer.

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A short-term in vitro test for tumour sensitivity to adriamycin based on flow cytometric DNA analysis

Cited by 18 publications

References 10 publications

Chemosensitivity of human small cell carcinoma of the lung detected by flow cytometric DNA analysis of drug‐induced cell cycle perturbations in vitro

Chemosensitivity of human small cell carcinoma of the lung detected by flow cytometric DNA analysis of drug‐induced cell cycle perturbations in vitro

Disaggregation of human solid tumours by combined mechanical and enzymatic methods

Dose-Dependent Effect of 17β-Estradiol Determined by Growth Curves and Flow Cytometric DNA Analysis of a Human Breast Carcinoma (T61) Grown in Nude Mice

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