Topoisomerase I-mediated antiproliferative activity of enantiomerically pure fluorinated homocamptothecins.

Homocamptothecin (hCPT) is an E-ring modified camptothecin (CPT) analogue bearing a methylene spacer between the alcohol and carboxyl functions of the CPT lactone. Combining pronounced inhibitory activity of topoisomerase I (Topo I) with enhanced plasma stability, hCPT constitutes an attractive template for the elaboration of new anticancer agents. Fluorinated hCPT analogues, prepared in enantiomerically pure form, were assayed by their stimulation of Topo I-mediated DNA cleavage. Translation into cytotoxicity against tumor cells was evaluated on HT29 human colon adenocarcinoma and on the multidrug resistant lung and bladder tumor cell lines, A549 and T24r. Good correlation is observed between the ability of the drugs to stimulate Topo I-mediated DNA cleavage and the respective 50% inhibitory concentrations (IC(50) values) of the HT29, A549, and T24r cell growth. Fluorine substitution in the A-ring of hCPT was found to have a pronounced influence on biological activity, providing several compounds which are up to 100-fold more potent than CPT in terms of IC(50). Among these, 10,11-difluoro-hCPT has been selected for further development.

Involvement of platelet-activating factor (PAF) in endotoxin- or ischaemia-induced intestinal hyperpermeability in the rat.

We have investigated the influence of BN 50727, a PAF antagonist, and allopurinol, a free radical scavenger, on the damaging effects of ischaemia-reperfusion and endotoxin in the small intestinal mucosa. Using a rat experimental model, we determined the alterations in intestinal permeability and mucosal levels of PAF and lysoPAF following ischaemia or intravenous administration of endotoxin. Both of these treatments increased intestinal permeability and enhanced PAF levels in the mucosa. Preventive oral or intraduodenal administration of BN 50727 reduced both of these effects, by decreasing mucosal PAF formation, probably as a result of neutrophil infiltration and activation reduction. Pretreatment of the rats with allopurinol also resulted in similar protection except that the free radical scavenger was unable to inhibit the increase in PAF levels after ischaemia, suggesting that oxidative reagents are implicated in this pathology to a much greater extent than PAF.

Antithrombotic activity of BN 50341, a structurally new compound with anticalcic and PAF-antagonistic properties.

BN 50341, a new benzazepine derivative, which has been shown to possess a mild anticalcic activity decreased (oral or i.v. administration) and also totally reversed (local superfusion) the in vivo electrically induced thrombus formation in rat or guinea-pig artery. These effects of BN 50341 were correlated with an inhibition of PAF-acether- and thrombin-induced platelet activation since it decreased free cytoplasmic calcium mobilization measured on cells loaded with the fluorescent probe Quin 2. Taking into account the beneficial effects of BN 50341 on vascular spasm as well as on the early stage of the process of white thrombus formation, this drug could be of therapeutic interest.

Changes in cytosolic free calcium induced by platelet-activating factor in rabbit platelets: specific inhibition by BN 52021 and structurally related compounds.

Increases in cytosolic free Ca2+ concentration induced by PAF-acether in rabbit washed platelets were recorded by using the fluorescent Ca2+ indicator Quin 2. In the presence of 1 mM external Ca2+, PAF-acether 2 X 10(-9) M has been found to increase the intracellular level of free calcium from a basal level of 135.0 +/- 26.9 nM to 2.0 +/- 0.7 microM. Pretreatment of platelets with the PAF-acether receptor antagonists BN 52020, BN 52021 or BN 52022 inhibited dose-dependently the PAF-acether-induced fluorescence signal. This activity was specific for PAF-acether, as shown with BN 52021, the most active derivative, which at 3 X 10(-6) M totally abolished PAF-acether effect without modifying thrombin - or calcium ionophore-induced signal. Kadsurenone, another PAF-acether receptor antagonist exhibited similar efficiency as BN 52021 against PAF-acether stimulation. Studied on PAF-induced aggregation of washed rabbit platelets: BN 52020, BN 52021 and BN 52022 exhibited the same potencies for inhibition as on the Quin 2 signal induced by PAF-acether; their activities were BN 52021 greater than BN 52020 greater than BN 52022. The results confirm that these derivatives, which are structurally closely related act at receptor level. The difference in their efficiencies seem to prove that the binding on its receptor site needs a highly defined chemical structure. They could be useful tools for structure-activity relationship studies in order to elucidate the conformation of the PAF-acether binding site.