Anchorage independent colony growth of fetal hamster lung epithelial cells after treatment with diepoxybutane.

To test the reliability of a new cell transformation assay, a cloned fetal Syrian hamster lung epithelial cell line (M3E3/C3) was used. The target cells originating from the respiratory tract were treated in vitro over a concentration range of 0-10(-5) M/l with diepoxybutane, cultured during the expression period of 28 or 35 days and then transferred into soft agar. Anchorage independent colony growth in soft agar occurs only if cells are transformed. Growth and number of colonies were taken as a score of the carcinogenic potential of the test substance. Under the conditions of this cell transformation assay it was possible to detect the carcinogenic potential of diepoxybutane unequivocally.

In vitro and in vivo investigations for the development of cytostatic methylhydrazones.

In in vitro short-term (3 h) assays, the beta-chloroethyl-methyl-hydrazones B 1 and B 2 inhibit the uptake of 3H-thymidine by EAC and L 1210 leukemia cells, B 2 being 5 to 10 times more effective than B 1. The growth inhibitory effect of both compounds was also confirmed in long-term (7 days) clonal assays using agar-containing glass capillaries, B 2 again being more effective than B 1. In contrast to these differences in vitro, in vivo both substances showed remission to the same degree in EAC- and complete resistance in L 1210-bearing mice. The diverging in vitro/in vivo sensitivities were thought to result from differences in the affinity of the methylhydrazones to the tumor cells: using short exposure periods (3 h) B 1 was more inhibitory than B 2 on both EAC and L 1210 colony growth; i.e., the more hydrophilic B 2 could more easily be washed off. To further test the idea of different cell membrane affinities, the methylhydrazones ZB 1 and P 1 with increasing lipophilic properties were synthesized. In vitro, after both pulse and continuous exposure ZB 1 and P 1 showed enforced inhibitory effects on colony growth. In vivo, ZB 1 and P 1 reduced the tumor weight of EAC mice, while only P 1 increased the survival time of L 1210 mice. The results suggest that from the combination of in vitro/in vivo assays mechanistic conclusions can be derived that are valuable for further development of these cystostatics.

Cytoprotection by somatostatin of normal and malignant clonogenic cells against the in vitro cytotoxicity of bischloroethylnitrosourea (BCNU).

The various pharmacological effects of somatostatin may be explained by the hypothesis that the paracrine peptide, by "stabilizing" cell membranes, inhibits the secretion of hormones as well as protects other cells (vascular endothelium, parenchyma) from different lesions (vasculo-, organo-, cytoprotection). This hypothesis was tested in vitro, using bischloroethyl-nitrosourea (BCNU)-intoxicated stem cells of normal mouse granulopoiesis and of the L 1210 leukemia. Clonogenic mouse bone marrow and L 1210 cells were grown in agar-containing glass capillaries. Using these colony assays and a ID90 of BCNU, cyclic somatostatin influenced the BCNU-cytotoxicity neither at simultaneous nor at subsequent application. However, when given 2 h prior to BCNU, the inhibition of colony growth was almost totally abolished. This cytoprotective effect was seen with normal granulopoietic as well as with leukemic cells. The effect did not show up, if the inactive linear somatostatin was used. N-acetyl-cysteine, a SH-compound applied as a chemoprotective adjunct, did not reveal a cytoprotective effect under identical experimental conditions, either. The results were discussed in view of common efforts to reduce the toxicity of cancer chemotherapy.