Antitumor activity of a new antibiotic, kazusamycin

The antitumor activity of a new antibiotic, kazusamycin, against various murine tumors was studied by various treatment schedules. Single, intermittent, and successive injections of the antibiotic were almost equally effective against Ehrlich carcinoma, IMC carcinoma and sarcoma 180 tumor, but successive injections showed more efficacy than the other schedules against Meth A fibrosarcoma and Lewis lung carcinoma. Interestingly, there was no clear dose response for the efficacy of kazusamycin on murine tumors. When HeLa cells were exposed to kazusamycin for 72 hours in vitro, the IC50 value was about 1 ng/ml, however the cytotoxicity of the antibiotic depended on the incubation time.     

Resorthiomycin, a novel antitumor antibiotic. I. Taxonomy, isolation and biological activity

Resorthiomycin, a novel antitumor antibiotic, was isolated from the fermentation broth of a strain of Streptomyces collinus by ethyl acetate extraction, silica gel chromatography and HPLC. Resorthiomycin exhibited an in vitro cytotoxic activity against mouse leukemia L5178Y cells (IC50, 15.5 micrograms/ml) and also inhibited the clonogenic activity of a multidrug-resistant mutant of human hepatoma PLC/PRF/5 cells to a greater extent than that of the parental cells. On the other hand, this antibiotic does not possess any antibacterial or antifungal activity.     

Mechanism of action of lactoquinomycin A with special reference to the radical formation

Lactoquinomycin A (LQM-A), an antibiotic containing a quinone moiety in the molecule, inhibited biosyntheses of DNA, RNA and protein to a similar extent in doxorubicin-resistant mouse leukemia L5178Y cells at concentrations higher than 0.08 micrograms/ml. The antibiotic caused cell death in a short period of incubation and the degree of cell death correlated with that of the inhibition of macromolecular syntheses, suggesting that the inhibition of macromolecular syntheses was not a primary effect of LQM-A. LQM-A served as a good electron acceptor, when cytochrome c reductase was used as a quinone reductase. The treatment of the cells with LQM-A significantly reduced cellular NADH and ATP levels. The generation of superoxide radical by LQM-A in cell lysate was observed by reduction of nitro blue tetrazolium, and the production of hydroxyl radical was confirmed by electron spin resonance. The importance of radical formation for the cytotoxicity of LQM-A is discussed.     

Lactoquinomycin, a novel anticancer antibiotic. I. Taxonomy, isolation and biological activity

Lactoquinomycin, a novel basic antibiotic, was isolated from the culture broth of a soil streptomyces by repeated solvent extraction and adsorption column chromatography. Morphological, cultural and physiological studies revealed that the organism belongs to the species Streptomyces tanashiensis. The antibiotic was active against bacteria, particularly Gram-positive organisms, and neoplastic cells in vitro. Antibiotic-resistant cell sublines of L5178Y lymphoblastoma were more significantly inhibited by lactoquinomycin than the parental cell line. Lactoquinomycin was effective against Ehrlich carcinoma in mice.     

丹酚酸A抑制核苷转运并增强化疗药物的抗肿瘤作用丹酚酸A抑制核苷转运并增强化疗药物的抗肿瘤作用

目的观察丹酚酸A(SAA)的抑制核苷转运活性及其抗肿瘤作用。方法用3H-TdR和3H-UR转运测定法,克隆生成测定法以及小鼠移植性肉瘤180模型。结果SAA抑制艾氏腹水癌细胞的胸苷和尿苷的转运,其IC₅₀分别为18.1和17.1 μmol·L^-1。SAA能明显增强5-FU、丝裂霉素C、MTX对KB细胞、肝癌BEL-7402细胞的细胞毒性。体内试验,SAA 200 mg·kg^-1和5-FU 10 mg·kg^-1单独使用的抑瘤率分别为41%和27%;SAA和5-FU联合使用的抑瘤率为63%(CDI=0.86)。结论SAA有抑制肿瘤细胞核苷转运的活性,可增强5-氟尿嘧啶等药物的抗肿瘤作用,有可能用于肿瘤联合化疗。     

Comparative uptake, retention and action of vincristine, vinblastine and vindesine on murine leukaemic lymphoblasts sensitive and resistant to vincristine.

1. The uptake and retention of vincristine (VCR), vinblastine (VBL) and vindesine (VDS) were evaluated comparatively with respect to their cytotoxic action on a murine lymphoblastic leukaemia (L5178Y). 2. The same parameters were measured on a derived subline of cells resistant to VCR (L5178Y/r) in order to determine whether the different degree of resistance to each alkaloid correlates with the amount of drug associated with the cells. 3. VCR was the most active on L5178Y cells (IC50 = 5.8 x 10(-9) M) while the activity of VBL and that of VDS were similar (IC50 4.4 x 10(-8) M and 3.5 x 10(-8) M, respectively). Nevertheless, a considerably larger amount of VBL was taken up by the cells compared to VDS, although there were no significant differences in their cytotoxic action. 4. The VCR resistant cell line also expressed resistance to VDS, whose IC50 was increased by a factor of 11.4, but not to VBL. However, the uptake and retention of the three alkaloids were similarly reduced in L5178Y/r cells regardless of the degree of resistance expressed. 5. Although a decreased drug uptake and/or retention by the cells provides an explanation for the resistance to vinca alkaloids, they do not seem to be the only factors accounting for the resistance shown by the cell line which we have isolated. 6. The results seem to indicate that part of the VBL taken up by the cells is not used to induce the cytotoxic effect, but is diverted to some cellular compartment(s) or rate controlling process(es) which are different from the target that mediates its cytotoxic action.     

In vitro screening for antitumour activity of Clinopodium vulgare L. (Lamiaceae) extracts

Aqueous extract of Clinopodium vulgare L. showed strong antitumour activity when tested in vitro on A2058 (human metastatic melanoma), HEp-2 (epidermoid carcinoma, larynx, human) and L5178Y (mouse lymphoma) cell lines-6 h after treatment disintegration of the nuclei and cell lysis started. Applied at a concentration of 80 microg/ml it reduced the cell survival to 1.0, 5.6 and 6.6%, respectively. The concentrations of aqueous extract inhibiting the growth of A2058, HEp-2 and L5178Y cells by 50% (IC50 values) were calculated to be 20, 10 and 17.8 microg/ml respectively. Two groups of active substances were detected: the first one, probably combining glycosides, influenced adhesion, while the second one caused massive cell vacuolisation. The chloroform extract, which contained ursolic acid and gentriacontan had also cytotoxic, however a little bit weaker effect. All changes observed were irreversible.     

Evaluation of ochratoxin A for mutagenicity in a battery of bacterial and mammalian cell assays

Ochratoxin A (OA), a nephrotoxic mycotoxin, was evaluated for genotoxic potential in a battery of in vitro and in vivo assays. OA was not mutagenic to Salmonella typhimurium, either with or without metabolic activation, in the plate incorporation (Ames) test at concentrations of 50-600 micrograms OA/plate or in the gradient plate assay at concentrations of 0.1-1000 micrograms OA/ml. No induction of unscheduled DNA synthesis was evident in primary cultures of rat hepatocytes exposed to concentrations of OA ranging from 0.000025 to 500 micrograms/ml. In the mouse lymphoma forward mutation assay, exposure of L5178Y TK+/- mouse lymphoma cells to OA did not increase the numbers of L5178Y TK-/- mutants. There was no significant difference between the numbers of sister-chromatid exchanges in cells from OA-treated Chinese hamsters and those in cells from the negative-control animals.     

Biological effects of acetomycin. I. Activity against tumor cells in vitro and in vivo

The antibiotic acetomycin was active in vitro against HCT-8 human colon adenocarcinoma cells (IC50, 1.5 microgram/ml) and L1210 murine leukemia cells (IC50, 2.2 micrograms/ml). Acetomycin also had marked activity in the human tumor stem cell assay, with a 33% overall response rate (less than or equal to 30% survival) against 49 primary tumors. However, acetomycin was inactive in four in vivo tumor assay systems (L1210 and P388 leukemias, B16 melanoma and the MX-1 mammary xenograft system). This lack of in vivo activity may result from metabolic inactivation of acetomycin.     

8006-I, an antibiotic from Amblyosporium spongiosum (Pers.) Hughes sensu Pirozynski. II. Biological properties

8006-I is an antibacterial antibiotic with a rather broad spectrum of activity. The minimum inhibitory concentrations for the most sensitive bacteria are in the range of one to ten micrograms/ml. Yeasts are not affected by concentrations up to 100 micrograms/ml. Some filamentous fungi like Fusarium oxysporum and Mucor miehei are inhibited at 100 micrograms/ml. In Ehrlich carcinoma ascitic cells the incorporation of uridine and leucine and to a lesser extent that of thymidine is reduced. In isolated nuclei of these cells the incorporation of UTP into RNA is inhibited. At low concentrations, the incorporation of uracil into trichloroacetic acid-precipitable material is almost completely inhibited in cells of Bacillus subtilis; at higher concentrations all macromolecular syntheses are affected. No reduction of respiration of the cells is observed. The antibiotic exhibits weak hemolytic activity and lytic activity towards bacteria. In vitro an inhibition of both DNA- and RNA polymerase from Escherichia coli is observed. Poly(U)-directed poly(Phe) synthesis is not affected.     

Antitumor activity of spergualin, a novel antitumor antibiotic

Spergualin (SGL), a novel antitumor antibiotic, exhibited strong antitumor activity against transplantable leukemias in mice: L1210, L1210(IMC), P388, P815, C1498, EL-4 and RL male 1. It also exhibited antitumor activity against M5076 fibrosarcoma, AH66 and AH66F rat hepatomas, but not against Meth-A fibrosarcoma, B16 melanoma, Lewis lung carcinoma (LL) and C26 colon adenocarcinoma. The antitumor activity of SGL was administration-schedule dependent. The strongest activity against L1210 was obtained by ip continuous infusion for 7 days or daily ip administration for 9 days. Single ip injection of SGL at 100 mg/kg to mice caused convulsion and death within 15 minutes after injection. Such acute toxicity was not observed by continuous infusion. SGL showed its strongest activity at subtoxic dose against sensitive tumors except for L1210(IMC). Mice implanted ip with L1210(IMC) were cured by treatment with SGL at 3.13 mg/kg/day for 9 days, but died from the tumor at 50 mg/kg/day X 9. The cured mice rejected a second inoculation of up to 10(6) tumor cells. The tumor cells isolated from mice after treatment with the high dose showed resistance to SGL in vivo. Mice implanted sc with L1210(IMC) were also cured by 9 daily ip administrations of SGL at 12.5 mg/kg/day, but solid tumor was observed at the implantation site until 3 days after the final injection of SGL in some cured mice. These results suggest that the therapeutic effect of SGL has a relatively high specificity for leukemias and that the immunological effect is involved in the antitumor activity.     

Mechanism of adriamycin resistance in a subline of mouse lymphoblastoma L5178Y cells

The biochemical mechanism of anthracycline resistance was studied with an adriamycin-resistant subline of mouse lymphoblastoma L5178Y cells. Both uridine and thymidine uptakes in the resistant cells were observed more resistant to adriamycin and daunorubicin than those in the parental cells. Aclacinomycin A exhibited the same degree of inhibition of nucleic acid syntheses in the sensitive cells and in the resistant cells. The resistance pattern observed by the inhibition of RNA and DNA syntheses seemed to parallel that by growth inhibition. No significant difference was demonstrated between the parental and resistant cells in the inhibition of RNA and DNA polymerase reactions with isolated nuclei. The uptake and retention of [3H]adriamycin was observed significantly less in the resistant cells than in the sensitive cells. The results suggested that the adriamycin resistance may be due to alteration of the cytoplasmic membrane and/or cytoplasm, resulting in decreased uptake and retention of the antibiotic in the resistant cells.     

Serum effect on cellular uptake of spermidine, spergualin, 15-deoxyspergualin, and their metabolites by L5178Y cells

Spergualin (SG) and 15-deoxyspergualin (DSG) were more slowly incorporated into L5178Y cells than spermidine. SG and DSG inhibited carrier-mediated transport of [3H]spermidine competitively with inhibition constants of 0.67 mM and 0.45 mM, respectively. Addition of calf serum stimulated uptake of [3H]spermidine into the cells in a serum concentration-dependent manner. The effect was not observed when horse serum was used in place of calf serum. Preincubation of spermidine in calf serum for 1 hour before addition to cells remarkably decreased cellular incorporation of tritium. Three amine oxidase inhibitors, aminoguanidine, 3-hydroxybenzyloxyamine, and semicarbazide, inhibited stimulation of uptake of [3H]spermidine by calf serum and the decrease of it by preincubation in calf serum. So we propose that cellular incorporation or binding of products generated by oxidation of spermidine by amine oxidase in calf serum was much faster than that of spermidine itself and they were unstable and transformed quickly to unincorporable or non-binding substances if cellular targets were not present. Effect of amine oxidase inhibitors on cytotoxic activity of SG and DSG were determined in low and high concentrations of calf serum. In the presence of 10% calf serum in the basal medium, cytotoxicity to L5178Y cells by SG and DSG was suppressed at high drug concentrations (above 10 micrograms/ml) and enhanced at low drug concentrations (below 2.5 micrograms/ml) by amine oxidase inhibitors. In the presence of 0.5% calf serum suppression of cytotoxicity at high drug concentrations by amine oxidase inhibitors was also observed, but enhancement at low drug concentrations was obscure. These data may suggest the existence of two kinds of cytotoxic mechanism of SG and DSG, one dependent on and one independent of amine oxidase in serum.     

Effects of an antitumor agent, ascofuranone, on the macromolecular syntheses of intact cells

Ascofuranone (AF) has antitumor protective property on experimental tumors. We examined the action of AF on lymphoma L5178Y to explore the mechanism of the antitumor activity. AF completely prevented the growth of L5178Y at 25 micrograms/ml cytostatically. The compound exhibited general inhibitory effects on the macromolecular syntheses. Among them, protein synthesis was most severely inhibited by AF and to the same extent as by cycloheximide. AF, however, did not affect protein synthesis by cell-free system even at 2 mg/ml. Although AF inhibited the incorporation of [14C]acetate into total acid precipitable products only slightly, the synthetic pattern of simple lipids from [14C]acetate was significantly changed. Especially, the incorporation of [14C]acetate into squalene was almost completely blocked at 25 micrograms/ml. The incorporation of [14C]acetate into triglyceride was inhibited and that into cholesterol was enhanced. Concerning the diglycerides, the incorporation of [14C]acetate was enhanced and that of [3H]glycerol was inhibited. The incorporation of [3H]glycerol and [3H]mevalonate into the intact cell was significantly inhibited as compared with [14C]acetate. As those effects were not observed with cycloheximide, they were suggested to be characteristic of AF. AF inhibited hypotonic hemolysis. In contrast, hemolysis by deoxycholate was stimulated. Possible mechanism of the antitumor activity of AF is discussed.     

Kazusamycin B, a novel antitumor antibiotic

A novel antibiotic, kazusamycin B (C32H46O7, MW 542), was isolated from the fermentation broth of Streptomyces sp. No. 81-484 and the structure was established mainly on the basis of its physico-chemical properties. Unambiguous 13C NMR spectral analysis of kazusamycin B has been also accomplished. Kazusamycin B possesses potent cytocidal activities against L1210 (IC50 0.0018 micrograms/ml) and P388 (IC100 0.0016 micrograms/ml) leukemia cells in vitro.     

Synthesis, antiprotozoal and cytotoxic activities of new N-(3,4-dimethyl-5-isoxazolyl)-1,2-naphthoquinone-4-amino derivatives

Three derivatives of N-(3,4-dimethyl-5-isoxazolyl)-1,2-naphthoquinone-4-amino (1), a compound which exhibits significant activity against Trypanosoma cruzi and Plasmodium falciparum but with cytotoxicity toward murine L-6 cells, were synthesized with the aim of ameliorating its cytotoxicity. The in vitro antiprotozoal and cytotoxic activities of the synthesized compounds were evaluated against T. cruzi, Trypanosoma brucei rhodesiense, P. falciparum and murine L-6 cells. The hydroxymethyl (2) and the oxime (3) derivatives were active against T. cruzi, with IC50 values in a range comparable to those of 1 (IC50: 0.65 microg/ml) and benznidazole (IC50: 0.56 microg/ml) while the carboxymethyloxime (4) was inactive. Compounds 2 and 3 were cytotoxic toward L-6 cells, with IC50 values identical to that of 1 (IC50: 0.50 microg/ml). The results did not support the suggestion that 2 and 3 may be used as prodrugs of 1.     

Dynemicins, new antibiotics with the 1,5-diyn-3-ene and anthraquinone subunit. II. Antitumor activity of dynemicin A and its triacetyl derivative.

Dynemicin A showed extremely potent in vitro cytotoxicity against a variety of murine and human tumor cells. In the experimental animal tumor models implanted ip with P388, L1210 leukemias and B16 melanoma cells, dynemicin A administered ip significantly prolonged life-span of tumor-bearing mice with the wide range of activity. This antibiotic administered iv was also active against iv implanted P388 and L1210 leukemias. In the macromolecule biosynthesis of B16 melanoma cells, dynemicin A inhibited DNA synthesis specifically. The triacetyl derivative exhibited similar in vitro and in vivo antitumor activities to those of the parent antibiotic.     

In vitro leishmanicidal activity of aurones.

A series of aurones with drug-potential for Leishmania infections was identified in vitro using both a direct cytotoxicity assay against extracellular promastigotes of Leishmania donovani, L. infantum, L. enriettii, and L. major, and a test against intracellular amastigote forms of L. donovani residing within murine macrophages. The most active aurone (6-hydroxy-2-[phenylmethylene]-3(2H)-benzofuranone) had an EC50 of 0.45 microgram/ml in the extra-, and an EC50 of 1.40 micrograms/ml in the intracellular assay. Other aurones were active between 0.06-12.50 micrograms/ml and 0.04-7.81 micrograms/ml, respectively. When tested against murine bone marrow-derived macrophages as a mammalian host cell control, the compounds showed only moderate cytotoxicity (EC50 2.32 to > 25.0 micrograms/ml). This is the first report on aurones as a new class of natural products with leishmanicidal activity.