BN80927

Comparison study of lactone stability between BN80927 and SN38. High-performance liquid chromatography monitoring of the compounds in human plasma at 37°C over a 180-min period (A) for BN80927 in human (•) or mouse (▾) plasma and for SN38 in human (○) or mouse (▿) plasma. BN80927 lactone stability in human plasma over a 24-h period (B). Best-fit curves of the experimental points were determined by least square regression.

Effect of increasing concentrations of BN80927 on plasmid DNA relaxation by topoisomerase I (Topo I) or Topo II; comparison with SN38 or with etoposide (VP-16). Native supercoiled pUC19-DNA (0.3 μg) was incubated with Topo I (gel A) or Topo II (gel B) in the presence of BN80927 (Lanes 12–15), or SN38 (Lanes 7–10, gel A) or VP-16 (Lanes 7–10, gel B) at 10, 50, 100, and 200 μm. N,N-dimethylacetamide (DMA; solvent diluted in the same way as the drugs) is used as negative control (Lanes 2–5). The first Lane of each group (1, 6, and 11) correspond to the highest concentration of drug or solvent incubated in the presence of DNA but without enzyme, to see a possible direct effect of the drug on DNA. DNA samples were separated by electrophoresis on agarose gels. Gels were then stained in ethidium bromide and photographed under UV light. Nck+Rel, nicked DNA + fully relaxed DNA; Top., topoisomers; Sc, supercoiled DNA.

Detection of DNA-topoisomerase I (DNA-Topo I) or DNA-Topo II cleavable complexes in BN80927-treated HT29 cells; comparison with SN38 or with etoposide (VP-16). Cells treated with BN80927 were compared with cells treated with the solvent alone (CONTROL), SN38, or VP-16, at indicated concentrations. After 1-h incubation, cells were lysed and fractionated using a CsCl gradient. For each fraction, DNA content was detected by fluorometry (data not shown), and the presence of Topo I or Topo II was revealed on a dot-blot using antibodies specifically directed against Topo I (blot A) or Topo II (blot B). In the frames on the right side of the blots, DNA-topoisomerase complexes (DNA-containing fractions).

DNA-protein complex (DPC) stabilization by BN80927 in HT29 cells. DNA and proteins were respectively radiolabeled with 0.6 μCi/ml [³H]thymidine and 0.1 μCi/ml [¹⁴C]leucine for 18 h. Cells were then exposed to drugs [BN80927 (▾) or SN38 (▵] at various concentration and time, at 37°C. KCl-SDS coprecipitation of proteins, followed by determination of radioactivity, allowed a quantification of DPCs. Data are expressed by the ratio of ³H-DNA to ¹⁴C-protein. Statistical analyses were performed by the Student t test (dose-response experiments) or by a two-factor ANOVA without replication (reversion experiments). For dose-response experiments (A), HT29 cells were incubated for 1 h with concentrations of drugs ranging from 0 to 50 μm; *, P < 0.05; **, P < 0.005. For reversion experiments (B), HT29 cells were exposed to 0.5 μm BN80927 or 1 μm SN38 for 1 h at 37°C; *, P < 0.1. Drugs were then removed, and the cells were incubated in fresh medium for 0–240 min.

Antiproliferative effects of BN80927 on human tumor resting cells in vitro; comparison with SN38 and camptothecin (CPT). Synchronized G₀-G₁ HT29 cells were exposed to the compounds for 16 h. Cell survival was determined 72 h after drug and stressor removal.

Evolution of tumor growth in non-androgen-dependant human prostate adenocarcinoma PC-3 and DU145 xenograft models. BN80927 schedule dependency in PC3 (A) and DU145 (B) models. Four different schedules (numbered list a–c) of orally administered BN80927 were as follows: (a) 5 mg/kg, qd×14, 5 mg/kg every day for 14 consecutive days; (b) 5 mg/kg, bid×14, 5 mg/kg twice a day for 14 consecutive days; (c) 40 mg/kg, qwk×3, 40 mg/kg once a week for 3 weeks; and (d) 8 mg/kg, 4/3/4/3/4, 8 mg/kg once a day for 4 days followed by a rest period of 3 days (a 4/3 cycle) with the cycle repeated twice more for a total of 12 administrations over the course of 3 weeks. Black arrows, the start of treatments.