By 1,3-bis(2-chloroethyl)-1-nitrosourea Research Articles

Oxidative cell injury in the killing of cultured hepatocytes by allyl alcohol

The killing of cultured hepatocytes by allyl alcohol depended on the metabolism of this hepatotoxin by alcohol dehydrogenase to the reactive electrophile, acrolein. An inhibitor of alcohol dehydrogenase, pyrazole, prevented both the toxicity of allyl alcohol and the rapid depletion of GSH. Treatment of the hepatocytes with a ferric iron chelator, deferoxamine, or an antioxidant, N,N'-diphenyl-p-phenylenediamine (DPPD), prevented the cell killing but not the metabolism of allyl alcohol and the resulting depletion of GSH. Inhibition of glutathione reductase by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) sensitized the hepatocytes to allyl alcohol, an effect that was not attributable to the reduction in GSH with BCNU. The cell killing with allyl alcohol was preceded by the peroxidation of cellular lipids as evidence by an accumulation of malondialdehyde in the cultures. Deferoxamine and DPPD prevented the lipid peroxidation in parallel with their protection from the cell killing. These data indicate that acrolein produces an abrupt depletion of GSH that is followed by lipid peroxidation and cell death. Such oxidative cell injury is suggested to result from the inability to detoxify endogenous hydrogen peroxide and the ensuing iron-dependent formation of a potent oxidizing species. Oxidative cell injury more consistently accounts for the hepatotoxicity of allyl alcohol than does the covalent binding of acrolein to cellular macromolecules.

Preferential alkylation by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) of guanines with guanines as neighboring bases in DNA

The base sequence of DNA has been shown to influence the kinds and amounts of alkylation of purine bases by N-methyl- N-nitrosourea [W.T. Briscoe and L-E. Cotter, Chem. Biol. Interact. 56, 321 (1985)]. In the present study, the alkylation of DNA polymers of denned sequence by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) has been investigated. The assay involved treating poly (dG)-poly(dC), poly(dG-dC)-poly(dG-dC), poly(dA-dC)-poly(dG-dT), poly(dA-dG)-poly(dC-dT), and calf thymus DNA with BCNU, followed by hydrolysis to release the modified purine bases and separation and quantitation of these by HPLC. Analysis of the results revealed that there was a 24-fold increase of 7-(β-hydroxyethyl)guanine (HOEtG) in poly(dG)-poly(dC) relative to poly(dAdG)· poly(dC-dT). There was also a 3-fold increase in HOEtG in poly(dG-dC)· poly(dG-dC), poly(dAdC)·poly(dG-dT) and calf thymus DNA relative to poly(dA-dG)· poly(dC-dT). A 2- to 4-fold increase of 7(β-aminoethyl)guanine (AmEtG) was observed in poly(dG-dC)· poly(dG-dC) relative to the other polymers tested. This study has determined that guanines in certain base sequences in polydeoxy-ribonucleotides are more susceptible to BCNU alkylation at the N-7 position than guanines in other sequences.

Inhibition of O6-alkylguanine-DNA-alkyltransferase activity potentiates cytotoxicity and induction of SCEs in human glioma cells resistant to 1,3-bis(2-chloroethyl)-1-nitrosourea.

SF-188 is a human glioma-derived cell line resistant to the cytotoxic effects of and the induction of sister chromatid exchanges (SCEs) by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). Pretreatment of SF-188 cells with N-methyl-N-nitrosourea (MNU) for 1 h increased the cytotoxicity of a 1-h treatment with BCNU 2- to 10-fold and doubled the number of SCEs; the magnitude of these effects was dependent on the dose of both agents. Treatment of SF-188 cells with MNU resulted in a dose-dependent inhibition of O6-alkylguanine-DNA-alkyltransferase (O6-AT) activity. Low doses of MNU, which did not significantly inhibit O6-AT, did not potentiate SCE induction. Higher doses of MNU inhibited O6-AT and potentiated cytotoxicity and the induction of SCEs. These results are consistent with the hypothesis that in resistant cells treated with BCNU, O6-AT repairs O6-chloroethylguanine before it can form a DNA interstrand cross-link. Inhibition of this enzyme allows for the formation of BCNU-induced DNA interstrand cross-links resulting in increases in cytotoxicity and induction of SCEs. The correlation between cytotoxicity and the induction of SCEs suggests that measurement of SCEs may be useful for determining the cellular response of normal and tumor cells to in vivo treatment with combinations of chemotherapeutic agents.

Preliminary report on the pathogenicity of Legionella pneumophila for freshwater and soil amoebae.

Legionella pneumophila, the causative organism of Legionnaires' disease, is pathogenic for free living, ubiquitous, freshwater, and soil amoebae of the genera Acanthamoeba and Naegleria. Some species support the growth of strains from serogroups 1 to 6, others only strains from certain serogroups. Initial studies with seeded material indicate that amoebal enrichment could be utilised for the isolation of legionellae from clinical specimens and natural habitats. It is suggested that a vacuole, or amoeba, full of legionellae, rather than free legionellae, could be the infective particle for man.

Alterations induced by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) in deoxyribonucleic acid synthesis in normal and tumorous tissues: Implications for optimal chemotherapy

Alterations in DNA synthesis are induced by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) in host target tissues and L1210 tumor. The timing of suppression and recovery of DNA synthesis was similar at the four doses studied (4, 8, 16 and 32 mg/kg); however, a dose-response effect was seen as the magnitude of suppression of DNA synthesis increased with increasing dosages. Of greater potential therapeutic importance, however, was that a pattern of suppression and recovery of DNA synthesis occurred in the host target tissues (bone marrow and gastrointestinal mucosa) which was different from that seen in the tumorous tissue. These differences in suppression and recovery of the normal and tumorous tissues were exploited for the prediction of dose schedules which resulted in improved survival of the tumor-bearing mice. The alterations in DNA synthesis induced by BCNU suggested an early fall in the de novo thymidine pool size. The delay in onset of DNA suppression suggests that the intact molecule is not the active principle, nor is there direct enzymatic blockade of DNA synthesis.

Immunologic response of mice bearing leukemia L1210.

The primary antibody response of mice bearing transplanted leukemias was studied using the direct plaque-forming cell (PFC) technique. Decrease of the immune response was found in mice bearing advanced isogenic or allogeneic leukemia, whereas normal or even increased PFC response was seen when the antigenic stimulus was given at an early stage of the disease. Leukemia remission induced by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) did not prevent the immunosuppression of leukemic mice, although the same treatment schedule did not interfere with antibody production in nonleukemic animals.