Biomedical subjects
Publications and source records attributed to N Otake.
4'-O-Acyl derivatives of doxorubicin, daunorubicin, 13-deoxocarminomycin, 13-deoxo-10-hydroxycarminomycin, 13-deoxo-11-deoxycarminomycin were synthesized through the formation and mild acid hydrolysis of 2-oxazoline intermediates. The antitumor activity of these 4'-O-acyl derivatives against P388 leukemia was similar to or more effective than the parent anthracyclines.
Methods for determining concentrations of a new morpholino anthracycline MX2.HCl and its metabolites in biological samples using reversed-phase high-performance liquid chromatography and fluorescence detection are described. The limits of detection were less than 1 ng/ml for all compounds after extraction from 0.5 ml of plasma using C18 Sep-Pak cartridges and consecutive solvent extraction. The recoveries from rat plasma ranged from 72.0 to 89.3%. The peak-height ratio of the fluorescence intensities of these compounds versus internal standard showed a linear correlation for concentrations up to at least 500 ng/ml in the plasma (correlation coefficient r greater than 0.999). The within-day and between-day precisions of this assay were in the range 0.8-8.7% (n = 5) and 2.0-3.5% (n = 5), respectively. The concentrations of these compounds in the blood and urine can be also determined by a slight modification of the extraction procedure.
We previously reported that MX2, a new morpholino anthracycline, showed marked effects on pleiotropic drug-resistant sublines of murine P388 leukemia in vivo as well as in vitro. In this study we examine the in vitro cytotoxicity against pleiotropic drug-resistant sublines of human tumor cell lines. MX2 was effective against multidrug-resistant sublines of four human tumor cell lines; these cells, having a 4.8- to 200-fold cross-resistance to Adriamycin (ADM) showed only a 0.7- to 2.3-fold resistance to MX2 compared with the sensitive cells. To elucidate the mechanism by which MX2 overcomes multidrug resistance, the intracellular pharmacology of MX2 in human myelogenous leukemia K562 and its ADM-resistant subline (K562/ADM) was examined. Both K562 and K562/ADM cells accumulated MX2 more easily than ADM, and the intracellular accumulation of MX2 attained a steady state in both cell lines within 30 min of incubation at 37 degrees C. The amount of MX2 that accumulated in K562/ADM at a steady state was only 1.3 times lower than that in K562. However, ADM was accumulated slowly in both cell lines compared with MX2, and the intercellular concentration reached a steady state in K562/ADM after 90 min of incubation and in K562 after more than 120 min. K562/ADM cells accumulated a 3.3-fold lower concentration of ADM than K562 after 120 min of exposure. The steady-state concentration of ADM in K562/ADM was 8.3 times lower than that of MX2. In addition, greater than 70% of MX2 was retained in both cell lines after 150 min of incubation in the absence of this drug. Verapamil, a calcium antagonist, hardly augmented the cytotoxicity of MX2 against K562/ADM, and no distinct effect of this drug on both the time course and the maximal level of accumulation of MX2 was observed. Interestingly, MX2 effectively inhibited ATP/Mg2(+)-dependent [3H]vincristine binding to K562/ADM membrane preparations, indicating that MX2 could be transported outside the cell by an active efflux pump. The high intracellular accumulation and retention of MX2 in K562/ADM through the rapid influx of the drug into the cells may be one of the reasons why MX2 circumvents pleiotropic drug resistance.
The temperature-dependent and concentration-dependent 1H NMR studies as well as 13C-relaxation experiments on gilvocarcin V tetraacetate strongly suggest intermolecular stacking of the antibiotic solution.
We cloned six different DNA fragments from a curromycin producing strain, Streptomyces hygroscopicus 358AV2, which confer curromycin-resistance on a curromycin non-producing and sensitive strain, S. hygroscopicus Rgll, a protoplast regenerant of the strain 358AV2. We studied the plasmid pSHR2 carrying one of the DNA fragments. By Southern blot analysis, the cloned DNA sequence in pSHR2 was found to be deleted in the Rgll genome. From the Rgll strain, a curromycin producing revertant A-4 was obtained, indicating that the structural genes for the curromycin biosynthesis and resistance are retained in the Rgll genome. Based on the existence of A-4 and the deletion of the DNA region corresponding to the cloned DNA sequence in the Rgll genome, we conclude that the cloned DNA sequence carries a regulatory gene governing curromycin-resistance but not the resistance gene itself. The smallest DNA region in pSHR2 conferring curromycin-resistance was sequenced, and it was found that there were two small open reading frames (ORF) on each strand of the cloned DNA. In-frame fusion of ORFs to the reporter gene lacZ revealed that one ORF designated cre was indeed translated in vivo. The putative gene product deduced from the cre ORF is a basic and hydrophilic protein having a calculated molecular weight of 6 kdaltons.
3'-Deamino-3'-morpholino-13-deoxo-10-hydroxycarminomycin (MX2), a morpholino anthracycline derived from 13-deoxo-10-hydroxycarminomycin (R20X2) was 16 times less cytotoxic than R20X2 against cultured P388 leukemia cells. The reduced cytotoxicity of MX2 was not explainable by intracellular or intranuclear concentration of the drug or by its DNA-intercalating activity. Binding of MX2 and R20X2 to DNA was measured, after isolating the DNA fraction from an incubation mixture of the drugs with P388 cells or with calf thymus DNA. The amount of R20X2 bound to the DNA was obviously larger than that of MX2, and was dependent on incubation time. These data suggest that the poor binding activity of MX2 to DNA contributes to its reduced cytotoxicity.
New anthracycline antibiotics, 1-hydroxy-11-deoxycarminomycin II and 11-deoxycarminomycin II were produced by a blocked mutant MuW1 of Actinomadura roseoviolacea from epsilon-pyrromycinone and aklavinone, respectively. We found that the enzyme catalyzing hydroxylation at the C-11 position was not lost but was down regulated in the strain MuW1.
Feeding experiments with D-[6,6-2H2]-, D-(6R)-[6-2H1]- and D-(6S)-[6-2H1]glucose in the fermentation of Streptomyces ribosidificus, followed by field desorption MS and 2H NMR analyses of the resulting labeled ribostamycin samples, clearly demonstrated that 1) both hydrogens of the C-6 hydroxymethyl group of D-glucose are stereospecifically incorporated into the C-2 position of 2-deoxystreptamine and 2) the pro S hydrogen of the C-6 position of D-glucose is stereospecifically removed during the elaboration of neosamine C in the biosynthesis of ribostamycin. A plausible mechanism of formation of the deoxy-scyllo-inosose, an early precursor to 2-deoxystreptamine, is suggested to be analogous to the dehydroquinate synthesis in the shikimate pathway and the conversion of the C-6 hydroxymethyl group of D-glucose into the aminomethyl group of neosamine C is likely to involve a dehydrogenation step to a formyl group.
2-Phosphinomethylmalic acid (PMM) synthase catalyzes the condensation of phosphinopyruvic acid (PPA), an analog of oxalacetic acid, and acetyl-CoA to form PMM. The enzyme was purified approximately 700-fold from a cell-free extract of Streptomyces hygroscopicus SF-1293, a bialaphos producing organism, to an electrophoretically homogeneous state. The purified PMM synthase has a subunit molecular weight of 48,000 by SDS-polyacrylamide gel electrophoresis and a native molecular weight of 90,000 approximately 98,000 by gel filtration. PMM synthase was relatively unstable, showed maximum activity at pH 8.0 and 30 degrees C, and was inhibited strongly by p-chloromercuribenzoate, iodoacetamide and EDTA. Enzyme activity suppressed by EDTA was completely restored by adding Co++ or Mn++ and partially restored by addition of Ca++, Fe++ or Mg++. The specific substrates of this enzyme are PPA or oxalacetic acid in addition to acetyl-CoA. The enzyme does not catalyze the liberation of CoA from acetyl-CoA in the presence of alpha-keto acids, such as pyruvate, alpha-ketoglutarate, deamino-alpha-ketodemethylphosphinothricin or phosphonopyruvate. The condensation reaction did not take place when propionyl-CoA or butyryl-CoA was used as a substrate in place of acetyl-CoA. The Km values of the enzyme were 0.05 mM for acetyl-CoA, 0.39 mM for PPA and 0.13 mM for oxalacetate. PMM synthase is very similar to (R)-citrate synthase of Clostridium in the inhibition pattern by sulfhydryl compounds, its metal ion requirement and stereospecificity; unlike (R)-citrate synthase PMM synthase was not inhibited by oxygen.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Effort looking for new antitumor antibiotics useful for the treatment of curing cancer resulted to the discovery of a number of new compounds with newer action mechanism as well as newer structural feature. The antibiotics which have been discovered since 1984 are discussed under classifications of action mechanism and structural feature, as well. The first group, which belong to a novel class of antibiotics containing a bicyclodiynene carbon skeleton in the molecules exhibited the most strong anti-tumor activity comparing with the antitumor antibiotics so far discovered. The action mechanism of this was explained by the diradical formation of diynene-cyclization, which led to the scission of double strand DNA. Amongst, esperamicin A seems of great interest in view of the therapeutic development. Moreover, elsamicin A, a member of chatarin antibiotics, and FR-900482 compound, an antibiotic having a polycyclic alkalodal skeleton are under development for the new chemotherapeutic agents. Rhizoxin, the metabolite of Rhizopus chinensis is also a promising candidate as anticancer agent. Its action mechanism was classified as an inhibitor of mitosis by binding to the microtibline proteins. Rhizoxin A shows no cross resistance with vincristine. MX2 (KRN 8602), the morpholino derivative of 13-deoxo-10-hydroxy-carminomycin, shows anticancer activity against tumor cells resistant to P388/ADM as well as low cardial toxicity. Miscellaneous compounds whose action mechanism are unknown are described.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Arugomycin (AGM) is a new anthracycline antibiotic produced by strain No. 1098-AV2 which was identified as Streptomyces violaceochromogenes. AGM was isolated by solvent extraction, silicic acid chromatography and Sephadex LH-20 column chromatography. Acid treatment of AGM gave the chromophore, named arugorol, which was identified as 4'-epi-nogalarol, and sugar moieties. AGM inhibited the growth of Gram-positive bacteria and showed antitumor activity against sarcoma S-180 and Ehrlich ascites tumors.
The structure of arugomycin was determined by chemical degradation, and NMR and mass spectral analyses to be a new anthracycline antibiotic with arugorol (4'-epi-nogalarol) as the chromophore and two sugar chains comprising diginosyl-decilonitrosyl-2-deoxyfucose, and (4-O-fumaryl-diginosyl)-diginosyl-2-deoxyfucosyl -diginose.
Biological activities of arugomycin and its analogues obtained by chemical degradation and modification were evaluated. Differences in the sugar moieties affected their biological activities including induction of differentiation of mouse Friend erythroleukemia cells and mouse myeloid leukemia cells, antitumor activities against sarcoma S-180, Ehrlich ascites carcinoma and P388 leukemia, and cytotoxicity against murine leukemia cells. Some relationships were found between the sugar moieties and biological activities.
Explore the source record for details and available documents.