The Walker 256 carcinoma: a cell type inherently sensitive only to those difunctional agents that can form DNA interstrand crosslinks.

The Walker 256 rat tumour has been maintained in vivo for over 60 years and until recently was used as a primary screen for new antitumour agents. This screen was particularly useful in identifying difunctional alkylating agents as potentially useful anticancer agents and it would seem that the Walker tumour is composed of cells sensitive towards this type of agent. A cell line (WS) established from the Walker tumour retained the sensitivity of the tumour towards difunctional agents and we have examined its phenotype in comparison to a derived, resistant, cell line (WR). The response of WR cells to a range of cytotoxic agents was similar to other established cell lines whilst WS cells were much more sensitive only towards difunctional reacting agents. There were no significant differences in the binding of these agents to the DNA of WS or WR cells. All the agents towards which WS cells showed sensitivity were, without exception, capable of reacting with DNA in Walker cells and forming DNA-DNA interstrand crosslinks. WS cells were not sensitive to busulphan, BCNU, CCNU or Me-CCNU but these agents did not produce interstrand crosslinks in the DNA of either WS or WR cells. Thus WS cells are intrinsically sensitive to specific DNA damage and this is probably a DNA interstrand crosslink. Hybrid cells produced by fusion of WS with WR cells lacked the inherent sensitivity of the WS cells towards cisplatin; sensitivity was therefore a recessive characteristic. Transfection of WS cells with human DNA also gave rise to 2 cisplatin-resistant clones, although it could not be ascertained if these clones were true transfectants or revertants. The survival of these resistant clones, after treatment with cisplatin, was about the same as WR cells a finding which would be consistent with complementation by a transferred gene or reversion of a single gene defect in WS cells. In their sensitivity only to difunctional compounds and lack of an apparent DNA excision repair defect the phenotype of Walker cells strongly resembles those cells from human patients suffering from Fanconi's anaemia and also of yeast snm1 mutant cells. The mechanisms giving rise to this failure to tolerate specific DNA damage (which seems to involve the inability to recover from the initial inhibition of DNA synthesis and may involve a single defect of a gene involved in the late steps of crosslink repair), do not involve drug uptake, drug binding to DNA, cell size, cell doubling time or DNA excision repair.