Reduced variation in the clonogenic assay obtained by standardization of the cell culture conditions prior to drug testing on human small cell lung cancer cell lines

Abstract

An advantage of established tumor cell lines compared to fresh human tumor specimens used in sensitivity assessments is the possibility of repeated experiments. Ultimately a database of sensitivity profiles on a panel of cell lines can be made and the sensitivity to new drugs compared with historical data. A prerequisite of this strategy is a minimal interexperimental variation. The sensitivity of eight human small cell lung cancer cell lines to adriamycin, daunomycin, aclacinomycin A, and mitoxantrone was tested in the clonogenic assay. A covariation in the sensitivity to the drugs emphasized the importance of simultaneous drug testing on the same batch of cells. On one cell line (NCI-N592) the interexperimental variation was further evaluated and a significant correlation was found between preexposure culture conditions, size of S-phase, and sensitivity to adriamycin, daunomycin, and mitoxantrone. Rigorous standardization of the growth conditions prior to clonogenic assay reduced the variation in the sensitivity to adriamycin from a factor of five to only 10-15%. It is concluded that simultaneous experiments on the same batch of cells in drug comparisons should be used if possible. Specification and standardization of culture conditions are necessary in the comparison of drugs tested in different experiments. Inclusion of the same reference drug in all experiments may further increase the validity of comparisons in different experiments.

In vitro evaluation of the potential of aclarubicin in the treatment of small cell carcinoma of the lung (SCCL)

“…Despite a variation with a factor of two in the S-phase distribution, the interexperimental variation in LD50 is less than 25% and not correlated to the size of the S-phase. With similar S-phase variations the sensitivity with 1-h incubation varied 250% (Jensen et al, 1989 (Hill et al, 1989). Although is relatively more sensitive to DAU, the sensitivity to ADR and DAU covariate and is almost equal in most cell lines, whereas DAU is more potent than ADR with 1-h incubation.…”

“…We have previously shown that the sensitivity of the SCCL cell lines to ADR, DAU and MITO with 1-h incubation is dependent on the size of the S-phase fraction at the time of drug exposure and that reduced variation in the clonogenic assay can be obtained by standardisation of the cell culture conditions (Jensen et al, 1989). In the present study, the inter-experimental variation after continuous incubation was evaluated on cell line NCI-N592 in three experiments with varying S-phase distribution (Table II).…”

“…Recently, a phase III trial has found ACLA to be superior to DAU in the treatment of de novo AML (Hansen et al, 1988). In a previous study on the inter-experimental variation in the clonogenic assay we observed that the sensitivity pattern to ACLA in a panel of small cell lung cancer (SCCL) cell lines was different from the sensitivity to adriamycin (ADR), DAU and mitoxantrone (MITO) after short time incubation (Jensen et al, 1989). The present investigations were initiated to further elucidate the differences between ACLA and the other analogues.…”

Differential cytotoxicity of 19 anticancer agents in wild type and etoposide resistant small cell lung cancer cell lines

“…Determination of 3H-daunorubicin accumulation This was performed as described by Skovsgaard (1978) with the modification that the SCLC cells in single cell suspensions were incubated for 15 min with DNase I (0.025% w/v) (Sigma) before being exposed to daunorubicin (Versantvoort et al, 1992 (Skovsgaard, 1978 (Jensen et al, 1989a). In our attempts to rationalize 'in vitro phase II trials in SCLC' the task was therefore to obtain more dose-response curves on one batch of cells.…”

Glycated Collagen I (GC) impairs angiogenesis in vitro—A study using an innovative chamber for cell research

“…Another applicable area for the chamber is pharmacological research. Using this method, it will be possible to monitor simultaneously effects of drugs on different cell types (13) In conclusion, our novel chamber facilitates cell research, bringing an opportunity for better standardization of experimental conditions and simultaneous observation of different experimental groups, reducing the possible effect of any systematic error. A novel chamber.…”