2-[(Aminopropyl)amino] Ethanethiol-Mediated Reductions in 60 Co g-Ray and Fission-Spectrum Neutron-Induced Chromosome Damage in V79 Cells

Schwartz, Jeffrey L.; Giovanazzi, Susan M.; Karrison, Theodore; Jones, Carol; Grdina, David J.

doi:10.2307/3577187

Cited by 37 publications

(13 citation statements)

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“…Because of the nature of the mutagenic agent used in our original mutagenesis study (fission-spectrum neutrons), it was likely that a deletion mutation had occurred (23,24). The genetic lesion causing the NEUT2 trait has been shown here by Southern blot analysis to be a deletion mutation in the gene encoding lO-formyl-THF DH.…”

Section: Protein Sequencing Partial Purification Ofproteins Msn13mentioning

confidence: 83%

Identification of a heritable deficiency of the folate-dependent enzyme 10-formyltetrahydrofolate dehydrogenase in mice.

Champion

Cook

Tollaksen

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

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During the analysis of liver protein expression in the offspring of male mice Irradiated with fissionspectrum neutrons, one ofprin displayed a heritable 50% decrease in the abundance of two proteins. Homozygous mice lacing detectable quantities of these proteins were obtained through breeding. Characterization of this protein deficiency has identified these liver proteins as forms of the enzyme 10-formyltetrahydrofolate dehydrogenase (10- Although the physical properties and substrate and product binding characteristics of 10-formyl-THF DH have been described, the physiological role of this enzyme in mammalian metabolism has not been directly addressed. Certain characteristics of mice lacking 10-formyl-THF DH indicate that disruption of the structural gene has significant physiological consequences. While homozygous mice appear completely normal despite this enzyme deficiency, breeding pairs of homozygotes show a dramatically decreased reproductive efficiency (6). Analysis of pool sizes of hepatic folate derivatives revealed shifts in the steady-state abundance of some forms of THF in mice lacking 10-formyl-THF DH. EXPERVIMENTAL PROCEDURESLiver Protein Sample Preparation. Liver protein samples were prepared as described (7) and were either frozen at -700C and later ground in liquid nitrogen or homogenized in 8 volumes of a solution containing 9 M urea, 2% 2-mercaptoethanol, 2% ampholytes (Pharmacia LKB, pH 9-11), and 4% Nonidet P-40. The homogenates were centrifuged at 435,000 x g for 5 min in a Beckman TL100 ultracentrifuge. The resulting supernatants containing denatured liver proteins were collected and stored frozen at -700C until analysis by two-dimensional electrophoresis (2DE).Preparation of Clarified Mouse Liver Cytosol. Mouse liver was minced with scissors and homogenized in 5 volumes (4-5 ml) of 0.25 M sucrose buffered with 10 mM Hepes using a motor-driven pestle at 3000 rpm for 30 strokes on ice. Aliquots (1 ml) of homogenate were then centrifuged for 15 min at room temperature in a Beckman TL100 ultracentrifuge. The supernatants were collected, combined, and dialyzed at 40C against 0.02 M phosphate buffer (pH 7.1), which was changed three times at hourly intervals.2DE. Aliquots (10 pIl) of mouse liver samples were separated in the first dimension by isoelectric focusing in 10-inch rod gels (8). A mixture of 25% Biolyte (Bio-Rad) pH 3-10/ 25% Servalyte pH 3-10/50%6 Biolyte pH 5-7 was used to generate the pH gradient. The second-dimension separation was performed as described by O'Farrell (9) on 10-17% linear gradient slab gels prepared as described (10). Gels were fixed

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Section: Protein Sequencing Partial Purification Ofproteins Msn13mentioning

confidence: 83%

Identification of a heritable deficiency of the folate-dependent enzyme 10-formyltetrahydrofolate dehydrogenase in mice.

Champion

Cook

Tollaksen

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

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“…In addition, we observed no effect of holding cells at 4°C for up to 3 h on the formation of DSB in nonirradiated control cells. Fission spectrum neutrons from the JANUS reactor are known to be significantly more clastogenic (Schwartz et al, 1988), mutagenic , carcinogenic (Han & Elkind, 1979;Thomson et al, 1982) and lethal (Ngo et al, 1977) than 60Co y-rays and/or X-rays. Consequently, it was of interest to assess whether similar differences could be detected at the level of DNA damage and/or repair as measured by the neutral filter elution technique at pH 7.2.…”

Section: Dna Damagementioning

confidence: 99%

“…Whether the inhibition by WR1065 of the post-irradiation formation of DSB is advantageous with respect to cellular repair at these high doses is unclear at present. However, reports that WR1065 can protect against JANUS-neutron-induced chromatid aberrations (Schwartz et al, 1988) and mutagenesis at much lower radiation doses suggest that it is capable of enhancing a post-irradiation repair process(es).…”

Section: Dna Damagementioning

confidence: 99%

“…clastogenic effects of high-LET fission spectrum neutrons Schwartz et al, 1988) in cultured cells. These observations have prompted us to expand further our studies concerning the role of aminothiols in the formation and repair of radiation-induced DNA damage.…”

mentioning

confidence: 99%

“…It is well known that selected aminothiol compounds can protect against the induction of single-and double-strand breaks in the DNA of irradiated cells (LaSalle & Billen, 1964;Sawada & Okada, 1970;Billen, 1983;Grdina & Nagy, 1986;Sigdestad et al, 1987;Murray et al, 1988 clastogenic effects of high-LET fission spectrum neutrons Schwartz et al, 1988) …”

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confidence: 99%

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The effect of 2-[(aminopropyl)amino] ethanethiol on fission-neutron-induced DNA damage and repair

Grdina

Cp²,

Dale³

et al. 1989

Br J Cancer

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Summary The effect(s) of the radioprotector 2-[(aminopropyl)amino] ethanethiol (WR1065) on fissionneutron-induced DNA damage and repair in V79 Chinese hamster cells was determined by using a neutral filter elution procedure (pH7.2). When required, WR1065, at a final working concentration of 4mM, was added to the culture medium, either 30min before and during irradiation with fission spectrum neutrons (beam energy of 0.85MeV) from the JANUS research reactor, or for selected intervals of time following exposure. The frequency of neutron-induced DNA strand breaks as measured by neutral elution as a function of dose equalled that observed for 60Coy-ray-induced damage (relative biological effectiveness of one). In contrast to the protective effect exhibited by WR1065 in reducing 60Co-induced DNA damage, WR1065 was ineffective in reducing or protecting against induction of DNA strand breaks by JANUS neutrons. The kinetics of DNA double-strand rejoining were measured following neutron irradiation. In the absence of WR1065, considerable DNA degradation by cellular enzymes was observed. This process was inhibited when WR1065 was present. These results indicate that, under the conditions used, the quality (i.e. nature), rather than quantity, of DNA lesions (measured by neutral elution) formed by neutrons was significantly different from that formed by y-rays.WR 1065 is the corresponding free thiol of the wellcharacterised radioprotector S-2-(3-aminopropylamino) ethyl phosphorothioic acid designated WR2721 (Purdie, 1979). The current clinical interest in these and similar compounds stems from early reports that these agents can preferentially protect normal as compared to neoplastic tissues against both acute and late-arising radiation-and/or chemotherapyinduced injuries (Yuhas, 1979;Phillips, 1980;Glover et al., 1984;Kligerman et al., 1984). Thiol compounds such as WR2721 and cysteamine have also been reported to be effective in protecting against oncogenesis in a number of experimental rodent systems (Marquardt et al., 1974;Apffel et al., 1975;Takeuchi & Murakami, 1978;Milas et al., 1984;Grdina et al., 1985b).WR1065 and cysteamine can both protect against radiation induced mutagenesis at the hypoxanthine-guanine phosphoribosyl transferase (HGPRT) locus in mammalian cells (Grdina et al., 1985a;Corn et al., 1987). WR1065 has also been found to be effective in protecting against the induction of HGPRT mutants by cisplatin , bleomycin and nitrogen mustard (Nagy & Grdina, 1986) and the transformation of 10T1/2 cells by ionizing radiation (Hill et al., 1986).Linked to the expression of each of these deleterious endpoints are, presumably, factors involving DNA damage and repair. It is well known that selected aminothiol compounds can protect against the induction of single-and double-strand breaks in the DNA of irradiated cells (LaSalle & Billen, 1964;Sawada & Okada, 1970;Billen, 1983;Grdina & Nagy, 1986;Sigdestad et al., 1987;Murray et al., 1988 Accordingly, the US Government retains a non-exclusive, royaltyfree licence to pu...

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Structure–activity analysis of the potentiation by aminothiols of the chromosome‐damaging effect of bleomycin in G₀ human lymphocytes

Hoffmann

Buccola

Merz

et al. 2001

Environ and Mol Mutagen

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The radioprotective aminothiols 2-[(aminopropyl)amino] ethanethiol (WR-1065) and cysteamine (CSM) potentiate the induction of chromosomal damage by the radiomimetic compound bleomycin (BLM) in G0 human lymphocytes. To investigate the mechanism of potentiation, we measured the clastogenic activity of BLM in the cytokinesis-block micronucleus assay in the presence and absence of amines, thiols, and aminothiols. The hydroxy analog of WR-1065, 2-(3-aminopropylamino) ethanol (WR-OH), potentiates BLM only slightly, indicating the critical nature of the thiol group. As thiols, WR-1065 and CSM may donate electrons for the activation of Fe(+2)-BLM or for the regeneration of Fe(+2)-BLM from inactive Fe(+3)-BLM. The amines putrescine, spermidine, and spermine all potentiate BLM, but they are weaker potentiators than the aminothiols, and they are effective only at high concentrations. Their activity, like that of WR-OH, is probably a consequence of conformational alteration of DNA. Dithioerythritol (DTE) and 2-mercaptoethanol (2-ME), thiols lacking an amino group, are less effective potentiators of BLM than are the aminothiols. The thiol group of WR-1065 and CSM is therefore essential, but insufficient, for explaining the strong enhancement of BLM activity. The cationic nature of CSM and WR-1065, conferred by the amino groups, evidently concentrates the active thiol function at the site of BLM action on DNA. As expected on this basis, the diamine WR-1065 is a more effective potentiator of BLM than is the monoamine CSM, whereas cysteine and N-acetylcysteine (NAC), which lack a net positive charge, potentiate BLM only weakly. These studies suggest that potentiation of the clastogenic action of BLM by aminothiols can be explained by the combination of a thiol-mediated redox mechanism and an amine-mediated targeting of the thiol function to DNA.

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2-[(Aminopropyl)amino] Ethanethiol-Mediated Reductions in 60 Co g-Ray and Fission-Spectrum Neutron-Induced Chromosome Damage in V79 Cells

Cited by 37 publications

References 22 publications

Identification of a heritable deficiency of the folate-dependent enzyme 10-formyltetrahydrofolate dehydrogenase in mice.

Identification of a heritable deficiency of the folate-dependent enzyme 10-formyltetrahydrofolate dehydrogenase in mice.

The effect of 2-[(aminopropyl)amino] ethanethiol on fission-neutron-induced DNA damage and repair

Structure–activity analysis of the potentiation by aminothiols of the chromosome‐damaging effect of bleomycin in G₀ human lymphocytes

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2-[(Aminopropyl)amino] Ethanethiol-Mediated Reductions in 60 Co g-Ray and Fission-Spectrum Neutron-Induced Chromosome Damage in V79 Cells

Cited by 37 publications

References 22 publications

Identification of a heritable deficiency of the folate-dependent enzyme 10-formyltetrahydrofolate dehydrogenase in mice.

Identification of a heritable deficiency of the folate-dependent enzyme 10-formyltetrahydrofolate dehydrogenase in mice.

The effect of 2-[(aminopropyl)amino] ethanethiol on fission-neutron-induced DNA damage and repair

Structure–activity analysis of the potentiation by aminothiols of the chromosome‐damaging effect of bleomycin in G0 human lymphocytes

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Structure–activity analysis of the potentiation by aminothiols of the chromosome‐damaging effect of bleomycin in G₀ human lymphocytes