Tomoko Yoshinari

Effect of genistein on topoisomerase activity and on the growth of [Val 12]Ha-ras-transformed NIH 3T3 cells

Genistein inhibited topoisomerase II and I; it increased the enzyme-DNA complex in L1210 cells at 1 micrograms/ml, and interfered with pBR322 DNA relaxation by the enzymes. To test the role of topoisomerase in the transformation by oncogenes, the effect of genistein on the transformation of NIH 3T3 cells by transfection with [Val 12]Ha-ras was compared with that of N-alpha-tosyl-L-lysyl-chloromethyl ketone (TLCK), since genistein inhibits tyrosine kinase as well as TLCK. Genistein reduced the number of foci of the transformed cells, and suppressed selectively the growth of ras-transformed NIH 3T3 cells but not normal NIH 3T3 cells. In contrast, TLCK did not affect the transformation. It inhibited the growth of the normal cells but not the transformed cells.

Synthesis and biological activities of NB-506 analogues modified at the glucose group.

A new indolocarbazole compound, NB-506 (1), modified at the glucose group yielded a beta-D-glucopyranoside, J-107,088 (2), which showed potent anticancer activity. A beta-D-ribofuranoside, J-109,534 (3), was found to be 6 times more potent than J-107,088 at inhibiting topoisomerase I.

Synthesis and biological activities of NB-506 analogues: Effects of the positions of two hydroxyl groups at the indole rings.

In the course of a study of 6-N-amino-substituted analogues of NB-506 (1), a more potent anticancer drug, J-109,404 (2), in which the formyl group of NB-506 was replaced with a 1,3-dihydroxypropane group, was reported. A study of further modification in the positions of two hydroxyl groups at the indole rings of 2 resulted in the discovery of a 2,10-dihydroxy analogue, J-107,088 (3), which is a promising anticancer agent with a broader therapeutic window than J-109,404.

Sequence-selective DNA cleavage by a topoisomerase I poison, NB-506

An indolocarbazole compound, NB‐506, inhibits the activity of topoisomerase I by stabilizing the DNA‐topoisomerase I complex (cleavable complex). NB‐506 inhibited the re‐ligation step of topoisomerase I activity more potently than camptothecin or its derivative, topotecan. A cleavage assay using an end‐labeled fragment of DNA revealed that the pattern of cleavage induced by NB‐506 was different from that induced by camptothecin. The preferred cleavage sites of NB‐506 were found to be not only T but also A or C at the 3′‐terminus of the cleaved DNA (position −1), while the DNA cleavage sites of camptothecin always had T at position −1. At the 5′‐terminus of the cleaved DNA (position +1), NB‐506 showed a preference for G, which is a feature shared in common with camptothecin. Therefore, the difference in cleavage patterns was most likely due mainly to the preferred base at position −1. Moreover, the re‐ligation rate was significantly slower at NB‐506‐selective sites, which had C at position‐1, than at camptothecin‐selective sites or at sites cleaved by both NB‐506 and camptothecin. Our data suggest that NB‐506 is an unique topoisomerase I poison and that its potent inhibition of topoisomerase I is partly dependent on retardation of re‐ligation at sites selectively induced by NB‐506. Int. J. Cancer 75:145–150, 1998.© 1998 Wiley‐Liss, Inc.

Reversal of multidrug resistance by new dihydropyridines with lower calcium antagonistic activity.

SummaryBS compounds, a series of new dihydropyridines, successfully overcame multidrug resistance in P388/ADR cells in vitro. These agents synergistically potentiated the cytotoxicity of Adriamycin to P388/ADR cells at a concentration of 1–2 μM, whereas they showed hardly any synergistic effect in the parental cell line (P388/S) at the same concentration. They inhibited the active drug efflux in P388/ADR cells as well as the binding of [G-3H]-vinblastine to membrane vesicles from P388/ADR, which was increased in resistant P388 cells as compared with parental cells. Besides, unlike the activity of clinically used calcium antagonists, the calcium antagonistic activity associated with BS compounds was very weak: their arterial relaxation activity was <21% of that of verapamil. These data suggest that BS compounds specifically overcome multidrug resistance without the serious hypotensive side effects that accompany the use of verapamil orother calcium antagonists.

Inhibition of Topoisomerase II by a Novel Antitumor Cyclic Depsipeptide, BE‐22179

BE‐22179, a novel cyclic depsipeptide antibiotic having two 3‐hydroxyquinoline moieties, inhibited the DNA‐relaxing activity of L1210 topoisomerase II completely at 0.08 μM. This effect was far stronger than that of VP‐16. However, it did not show any marked effect on topoisomerase II‐mediated DNA cleavage. BE‐22179 was ineffective in inhibiting the DNA relaxation by topoisomerase I at concentrations up to 10 μM, but showed DNA‐intercalating ability (DNA unwinding) at 30 μM. The structure of BE‐22179 is quite novel for a topoisomerase II inhibitor. Echinomycin, a quinoxaline antibiotic structurally related to BE‐22179, interfered with DNA relaxation by topoisomerase II, though the effect was not due to inhibition of the catalytic activity of topoisomerase II but to conformational change of DNA based on its intercalation into DNA. Therefore, the potent inhibitory activity on topoisomerase II might not be a common activity of quinoxaline antibiotics, but might rather be specific to BE‐22179. BE‐22179 prevented DNA synthesis as well as RNA synthesis in L1210 cells and inhibited the growth of the cells. However, it remains unclear to what extent the topoisomerase II inhibition was responsible for the cytotoxicity of BE‐22179.

Synthesis and biological activities of topoisomerase I inhibitors, 6-N-amino analogues of NB-506

6-N-Amino analogues of NB-506 [6-N-formylamino-12,13-dihydro-1,11-dihydroxy-13-(beta-D-glucopyranosyl) -5H-indolo[2,3-a]pyrrolo[3,4-c]carbazole-5,7(6H)-dione] (3b) were synthesized and tested with respect to topoisomerase inhibition, cytotoxicity and anticancer effects. Among them, a 1,3-dihydroxypropane analogue (J-109,404, 5t) showed more than ten times more potent anticancer activity in MKN-45 human stomach cancer cells implanted in mice than NB-506.

ED-110, a Novel Indolocarbazole, Prevents the Growth of Experimental Tumors in Mice

A new indolocarbazole compound, ED‐110, which was obtained by glucosylating a microbial product (BE‐13793C) and is a potent topoisomerase I inhibitor, showed characteristic inhibitory effects on the growth of 12 human tumor cell lines tested. The IC50 values of ED‐110 against 9 of the 12 lines ranged from 11.5 μg/ml to 0.07 μg/ml, while the remaining 3 lines were quite resistant (IC50, >100μg/ml). In in vivo experiments, i.p. treatment with ED‐110 increased the survival period by more than two‐fold in mice implanted i.p. with P388, L1210, L5178Y or EL4 murine leukemic cells. The minimum effective dose increasing the life‐span of mice bearing P388 leukemia by 25% was <2.5 mg/kg/day × 10 and the maximum tolerated dose was > 160 mg/kg/day × 10. ED‐110 was also effective against the spontaneous metastasis of mouse Meth A fibrosarcoma cells and the growth of xenografted MKN‐ 45 human stomach cancer cells as well as s.c. implanted mouse colon 26 and IMC carcinoma cells. These results indicated that ED‐110 may have potential as a new antineoplastic agent with a large chemotherapeutic index and a wide range of effective doses.

Stereo(C7)‐dependent Topoisomerase II Inhibition and Tumor Growth Suppression by a New Quinolone, BO‐2367

A new antimicrobial quinolone (—)BO‐2367, (‐)‐7‐[(1R*,2R*,6R*)‐2‐amino‐8‐azabicyclo[4.3.0.]‐non‐3‐en‐8‐yl]‐l‐cyclopropyl‐6,8‐difluoro‐l,4‐dihydro‐4‐oxo‐3‐quinorinecarboxyric acid, strongly inhibited both mammalian and bacterial topoisomerase II. The IC50 values of (—)BO‐2367 against the DNA relaxation activity of L1210 topoisomerase II and the supercoiling activities of Escherichia coli gyrase and Micrococcus luteus gyrase were 3.8, 0.5, and 1 μM, respectively. This compound enhanced double‐stranded DNA cleavage mediated by topoisomerase II not only with purified enzyme, but also with intact L1210 cells. All these activities of (—)BO‐2367 were more than 2‐fold stronger than those of VP‐16. Intriguingly, (+)BO‐2367, which has an enantiomeric substitnent at the C7 position of (‐)BO‐2367, did not affect the activity of the mammalian topoisomerase II, while it inhibited E. coll gyrase. Intraperitoneal injection of (‐)BO‐2367 at 0.08 mg/kg increased the lifespan of CDF1female mice bearing ascitic L1210 leukemia by 2.4 times, and subcutaneous injection at 1.25 mg/kg completely inhibited the growth of colon 26 carcinoma implanted subcutaneously. These results suggest that (—)BO‐2367 is a potent antitumor agent which targets topoisomerase II. These enantiomers should be a useful tool for studying drug‐topoisomerase II interactions.

Evaluation of the safety, tolerability, and pharmacokinetics of a single bolus injection of daptomycin in healthy Japanese subjects

UNLABELLED This was a phase I double-blind, randomized, placebo-controlled, 2-period, single-dose, crossover study in healthy Japanese subjects to evaluate the safety, tolerability, and pharmacokinetics of a single bolus injection of daptomycin. Twenty healthy subjects were randomized; 16 received a single intravenous (IV) administration of 6 mg/kg of daptomycin and 4 received a single intravenous administration of placebo (0.9% sodium chloride) by either bolus injection (10 s) or infusion (30 min) following an overnight fast in Periods 1 or 2. There was a minimum 5-day washout period from the administration in Period 1 to the administration in Period 2. Administration of a single bolus injection of daptomycin 6 mg/kg was generally well tolerated. The geometric mean ratios (GMRs) (90% confidence intervals [CIs]) for AUC0-∞, AUC0-24 h, Cmax, and C24 h of daptomycin in plasma following bolus injection over 10 s relative to IV infusion over 30 min were 1.01 (1.00, 1.03), 1.02 (1.00, 1.03), 1.50 (1.41, 1.60) and 1.05 (1.02, 1.08), respectively. Because no existing studies of this nature were available, Cmax following daptomycin 6 mg/kg/10 s multiple-dose bolus injections was simulated. It was estimated at 178 μg/mL (upper limit) and was expected to be equal to or less than the confirmed Cmax in Japanese or non-Japanese healthy subjects following single- or multiple-dose IV infusion over 30 min. From the results of this study, daptomycin multiple-dose bolus injections over 10 s are expected well tolerated and to have similar Cmax values as IV infusion over 30 min, thus offering potential clinical benefit. TRIAL REGISTRATION Because this was a phase I trial in healthy subjects, the trial was not registered.