Biomedical subjects

E Multon

Publications and source records attributed to E Multon.

5 recordsLinked to original sources

PubMed1991

Rapid postmortem decrease in the ectocellular acetylcholinesterase activity in rat striatum as assessed by in vivo microspectrophotometry.

The acetylcholinesterase (AChE) activity in striatum rat was determined before and shortly after death using the in vivo microspectrophotometric method. This technique allowed us to monitor the Ellman colorimetric reaction directly inside the brain using an optical probe implanted in a live animal and to determine locally the AChE activity. Whatever the cause of the animals death, we observed a drastic postmortem decrease of the AChE activity of about 35-50%, 10 min after death. We have verified that the postmortem decrease of brain temperature or pH and postmortem optical properties changes could only explain a fraction of the AChE activity fall (16%). This phenomenon seems to be related to events strictly localized at the cellular level, since local injection of cyanide at the measuring site promotes a decrease of the enzymatic activity (40%) close to the levels observed after death. The origin of this rapid postmortem fall of the AChE activity is discussed. The technical properties of the microspectrophotometric method exclude a decrease of the ectocellular pool of enzyme after death. Our results allow us to envisage the existence of an in vivo endogenous regulation of the AChE activity which disappears shortly after death.

Acetylcholinesterase↗

PubMed1989

Topoisomerase II-mediated DNA cleavage activity induced by ellipticines on the human tumor cell line N417.

Ellipticine derivatives have been shown to induce DNA strand breaks by trapping DNA-topoisomerase II (Topo II) in an intermediary covalent complex between Topo II and DNA which could be related to their cytotoxic effects. We report here that Celiptium and Detalliptinium, two ellipticine derivatives clinically used for their antitumoral properties against breast cancer, exhibit the highest in vitro activity on Topo II DNA cleavage reaction and decatenation among a series of 14 ellipticine derivatives. The in vitro cleavage site specificity in pBR 322 plasmid DNA and in a human c-myc gene inserted in a lambda phage DNA is identical for both ellipticines, but different from m-AMSA, another Topo II related antitumoral agent. Recently, it has been shown that the ellipticine derivative Celiptium presents a strong cytotoxic activity in vitro on different human tumors including small cell lung carcinoma (SCLC). However, the studies that involved Topo II as a target for ellipticine derivatives have been performed only by using animal tumor cell lines. Therefore we have studied the in vivo DNA cleavage activity of Celiptium and Detalliptinium on a human SCLC cell line, NCI N417, comparatively to that obtained with m-AMSA. The respective IC50 on cell growth are 9, 8 and 1 microM for Celiptium, Detalliptinium and m-AMSA, respectively. Using the alkaline elution technique, we have observed that Celiptium and Detalliptinium exhibit a weak cleavage activity on genomic DNA from whole cells. The ellipticines are about 50 times less potent than m-AMSA in inducing DNA strand breaks. Analysis of in vivo c-myc gene cleavage by Southern blot hybridization also demonstrates a lack of activity of the ellipticine derivatives as no gene cleavage could be detected up to 50 microM of the drug. With m-AMSA, c-myc gene cleavage is detected at a concentration of 0.2 microM, which indicates that this methodology is less sensitive in detecting DNA strand breaks than is the alkaline elution. Further studies of the drug effect on isolated nuclei by alkaline elution also show that the DNA cleavage activity of Celiptium and Detalliptinium is increased when compared to whole cells. Our data indicate that these two drugs have a weaker cytotoxic effect than m-AMSA on NCI N417 cell line, due to a limited access to the cell nucleus rather than to a lack of activity on Topo II as assessed by in vitro and isolated nuclei experiments.

Alkaloids↗

PubMed1987

In vivo and in vitro stimulation by antitumor drugs of the topoisomerase II-induced cleavage sites in c-myc proto-oncogene.

Stimulation of topoisomerase II cleavage activity by antitumor drugs with or without DNA intercalative ability has been tested in vivo on the c-myc proto-oncogene. Two human tumor cell lines (N417 and HL60 cells) were treated with mAMSA, OH-9-ellipticine, VM26, and BD-40 (an aza-ellipticine analog), and DNA breaks were mapped in the gene locus by Southern blot hybridization with c-myc probes. Most of the major cleavage sites induced in vivo by drugs in the 5' end of c-myc were also observed in vitro using purified topoisomerase II and a c-myc gene DNA insert. Moreover, they closely mapped to some DNAse I hypersensitive sites, the presence of which reflects gene activity. DNA from drug treated cells probed with a human beta 1 globin pseudogene, and the c-mos proto-oncogene did not reveal topoisomerase II cleavage bands, suggesting that topoisomerase II-mediated drug activity may correlate with gene activity.

Amsacrine↗

PubMed1986

The in vitro involvement of topoisomerase II in the activity of aza-ellipticine analogues is not correlated with drug activity on isolated nuclei.

Aza-ellipticines are DNA intercalative ellipticine analogues with antitumor activity that induce protein-linked DNA breaks in NIH 3T3 cells in culture. The effects of two aza-ellipticine congeners (BD-40 and BR-76) on the activity of purified Calf Thymus type II topoisomerase were studied using pUC13 DNA as substrate. DNA cleavage was stimulated by both molecules at those doses required for inducing lethal effects in cells (DE5O). This effect was reversed by high salt treatment, indicating that it was actually mediated by Topo II. Mapping of cleavage sites on linearized and 3' end-labelled pUC13 DNA showed that ellipticine and aza-ellipticines stimulated the same sites, which differed from those stimulated by m-AMSA. Decatenating activity of Topo II on Trypanosoma cruzi kDNA was both inhibited by ellipticine and BD-40 at concentrations much higher than DE50 concentrations. Activity of aza-ellipticines was also investigated on isolated nuclei. Unlike ellipticine which promoted DNA-breaking activity, BD-40 and BR-76 were repeatedly inactive. Prior treatment of DNA by Proteinase K did not reveal hidden breaks which are formed in intact cells treated with BD-40 (Vilarem et al., 1984, Nucleic Ac. Res. 12, 8653). Concordant with these data, BD-40 did not impair DNA-synthetic activity in isolated nuclei, while Ellipticine largely decreased it. These results indicate that lesions induced in DNA by Aza-ellipticines are mediated by Topo II. The absence of effect of these drugs on isolated nuclei compared to that of Ellipticine may be due to some specific features of the association between Topo II and Aza-ellipticines or reflect a bioactivation step as a prerequisite for in vivo activity.

Alkaloids↗

PubMed1986

In vivo stimulation by antitumor drugs of the topoisomerase II induced cleavage sites in c-myc protooncogene.

Several antitumor drugs including DNA intercalative and non intercalative agents induce in vitro and in vivo double-stranded DNA breaks by stabilization of a topoisomerase II-DNA complex. In order to locate cleavage sites in an actively transcribed oncogene, N417 cells, originating from a human small cell lung carcinoma and containing 45-50 copies of c-myc oncogene, were treated with mAMSA, 9 hydroxyellipticine and VM 26. The presence of DNA lesions in c-myc was investigated by Southern blot hybridization with a human c-myc probe. In addition to normal bands, DNA patterns of drug treated-cells revealed the presence of new bands most likely corresponding to topoisomerase II-mediated cleavage as these bands were not found in untreated control DNA and in DNA treated with oAMSA, a biologically inactive stereoisomer of mAMSA. Major cleavage sites induced by drugs in the N417 cell c-myc locus were located in the 5' end of the c-myc exon 1 closely to some DNAse I hypersensitive sites which are assumed to reflect an activity of the gene. Therefore our data suggest that TopoII-mediated drug activity correlates with gene activity.

Aminoacridines↗