N-Methylformamide (NMF), a cell-differentiating agent, was assessed for its antitumor activity against a fibrosarcoma (FSA), a hepatocarcinoma (HCA-I) and a mammary carcinoma (MCA-K), syngeneic to C3Hf/Kam mice. Tumors were grown as solitary tumors in the leg or as artificial or spontaneous micrometastases in the lung. NMF, at a dose of 300 mg/kg, was administered i.p. daily for 6 to 18 days. NMF slowed the growth of FSA and HCA-I tumors and totally inhibited the growth of the MCA-K tumor. However, the effect was transient; tumors resumed their pretreatment growth rate upon cessation of the treatment. Histologically, MCA-K tumors treated with NMF (300 mg/kg daily for six days) underwent considerable cell depopulation and reduction in mitotic activity. The number of artificial metastases, as well as the incidence and the number of spontaneous metastases, were markedly reduced by NMF. This resulted in a prolongation of the survival of mice that had artificial metastases of MCA-K tumor. The in vitro clonogenicity of MCA-K, but not of FSA or HCA-I cells, was reduced. However, in vivo reduction of MCA-K cell clonogenicity was minimal, if any. Thus, NMF is effective in restricting the growth of both solitary tumors and metastases, but the degree of response is highly dependent on tumor type.

Summary The effects of the differentiation-inducing polar solvent N-methylformamide (NMF) on artificially induced and spontaneous metastases from a murine hepatocarcinoma (HCA-1) in C3Hf/Kam mice were investigated. Exposure of HCA-1 cells in vitro for 6 days to 1.0% or 1.25% NMF resulted in an increase in the number of lung nodules formed in mice when these cells were injected into their tail veins. This in vitro NMF exposure increased cell volume and induced only a slight amount of cytotoxicity. Administration of NMF to mice I day before i.v. tumour cell inoculation resulted in a dose-dependent increase in the number of lung nodules formed, beginning at an NMF dose of 600mgkg-1. NMF caused a similar magnitude of metastasis enhancement in immunosuppressed mice. However, when the maximum dose tested (1,800 mg kg-') was administered as 6 daily fractions of 300 mg kg-each, no increase in artificial metastases was detected. Administration of NMF to mice one day after i.v. tumour cell injection resulted in a dosedependent decrease in the number of lung nodules. In mice bearing 5-6mm HCA-1 leg tumours, treatment with 6 daily fractions of NMF (300mg kg-each) significantly reduced the number of spontaneous pulmonary metastases, yet had very little effect on the growth of the primary tumour. These data suggest that, in a clinically relevant treatment setting, NMF can reduce metastasis formation.

During studies to establish why pretreatment of 9L cells with a-difluoromethylornithine (DFMO) enhanced cell killing induced by 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU) (1) and reduced cell killing induced by cis-platinum (2), we reported, in 1982, an enhancement of BCNU-induced sister chromatid exchanges (SCE's) and a reduction in cis-platinum-induced SCE frequency with DFMO pretreatment (3 killing by DFMO pretreatment had proved certain, however, and the discrepancy was puzzling in view of repeated confirmation of the correlation between reduced cell killing and SCE frequency with cis-platinum (3).New experimental results reported by others (5) show that when DFMO remains in the cell culture medium during the period of BCNU treatment, the procedure enhances the number of BCNU-induced SCE's in 9L cells. When the DFMOcontaining medium is removed and the cells are rinsed before BCNU is added, however, the additional rinsing procedure greatly diminishes or eliminates the enhancement. We believe that our replicate experiments (4) deviated from our original protocol (3) by the introduction of an additional rinsing procedure, thereby accounting for the discrepant results. There is now evidence that slides containing DFMO-treated cells degrade over time (6). Such a degradation over the 5 years between the original scoring of the slides and the rescoring attempts could have contributed to our inability to confirm the original findings.

Summary The ability of the sister chromatid exchange (SCE) assay to detect heterogeneity in intrinsic radiation sensitivity was investigated. In order to identify tumour cell subpopulations, frequency histograms of cis-diamminedichloroplatinum (II) (cPt)-induced SCEs were generated and compared to those from cultures that had been irradiated 96h before drug treatment. The results suggested that subpopulations with different radiosensitivities were present in nine of 18 human primary tumour cell cultures evaluated. When the effects of prior irradiation on the subsequent X-ray survival response and on cPt-induced SCE frequency histograms were compared, a good correlation was obtained between the two assays regarding the prediction of heterogeneity in radioresponse. These results suggest that primary cultures can contain both radiationsensitive and radiation-resistant cells, and thus heterogeneity in intrinsic radiosensitivity may exist in human solid tumours.

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