Resistant to Nitrogen Mustards but Collaterally Sensitive to Cellular and Chromosome Morphology in a Rat Cell Line Carbamoylation of Glutathione Reductase and Changes in

A Walker 256 rat carcinoma cell line (WR) with acquired resistance to nitrogen mustards has been found to lack crossresistance to nitrosoureas. Although total cellular glutathione pools were similar in the parent (WS) and resistant cell lines (WS, 2.5 x 10~6; WR, 2.0 x 10~6 mol/mg protein), glutathione reductase activity was 3.98 in WR compared to 8.67 nmol reduced nicotinamide adenine dinucleotide phosphate oxidized per /¿g protein per min in WS cells. Treatment of cells with a carbamoylating nitrosourea, /V,A/'-bis(frans-4-hydroxycyclohexyl)-A/'-nitrosourea, produced a dose-dependent inhibition of glutathione reducÃ-aseand depletion of thiols in both cell lines. The drug caused no direct DMA strand breakage, but a differ ential mitotic spindle-chromosome stain showed that spindle formation was inhibited in WR cells at A/,A/'-bis(fAans-4-hydroxycyclohexyl)-/V'-nitrosourea concentrations of greater than 50 ^MIn WS cells, mitotic figures were still visible at 100 MM-Chromo somal damage was expressed in both cell lines at concentrations of 25 UM. The number and extent of these aberrations were greater in WR than WS. Observed karyotypic abnormalities included polyploidy, chromosome decondensation, and endoreduplication. In interphase cells, transmission electron microscopy showed that the most prevalent drug-induced lesions included (a) disappearance of plasma membrane filopodia, (b) appearance of membrane blebbing, and (c) development of irregular crescentshaped nuclei. These morphological and cytogenetic changes correlate with cytotoxic responses of these cell lines to N,N'bis(frans-4-hydroxycyclohexyl)-A/'-nitrosourea and would be consistent with drug-induced inhibition of glutathione reducÃ-ase.