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Scientific Reports Jul 2021Streptomyces smyrnaeus UKAQ_23, isolated from the mangrove-sediment, collected from Jubail,Saudi Arabia, exhibited substantial antimicrobial activity against...
Isolation, characterization, anti-MRSA evaluation, and in-silico multi-target anti-microbial validations of actinomycin X and actinomycin D produced by novel Streptomyces smyrnaeus UKAQ_23.
Streptomyces smyrnaeus UKAQ_23, isolated from the mangrove-sediment, collected from Jubail,Saudi Arabia, exhibited substantial antimicrobial activity against methicillin-resistant Staphylococcus aureus (MRSA), including non-MRSA Gram-positive test bacteria. The novel isolate, under laboratory-scale conditions, produced the highest yield (561.3 ± 0.3 mg/kg fermented agar) of antimicrobial compounds in modified ISP-4 agar at pH 6.5, temperature 35 °C, inoculum 5% v/w, agar 1.5% w/v, and an incubation period of 7 days. The two major compounds, K and K, were isolated from fermented medium and identified as Actinomycin X and Actinomycin D, respectively, based on their structural analysis. The antimicrobial screening showed that Actinomycin X had the highest antimicrobial activity compared to Actinomycin D, and the actinomycins-mixture (X:D, 1:1, w/w) against MRSA and non-MRSA Gram-positive test bacteria, at 5 µg/disc concentrations. The MIC of Actinomycin X ranged from 1.56-12.5 µg/ml for non-MRSA and 3.125-12.5 µg/ml for MRSA test bacteria. An in-silico molecular docking demonstrated isoleucyl tRNA synthetase as the most-favored antimicrobial protein target for both actinomycins, X and D, while the penicillin-binding protein-1a, was the least-favorable target-protein. In conclusion, Streptomyces smyrnaeus UKAQ_23 emerged as a promising source of Actinomycin X with the potential to be scaled up for industrial production, which could benefit the pharmaceutical industry.
Topics: Anti-Bacterial Agents; Computer Simulation; Culture Media; Dactinomycin; Drug Evaluation, Preclinical; Fermentation; Magnetic Resonance Spectroscopy; Mass Spectrometry; Methicillin-Resistant Staphylococcus aureus; Molecular Docking Simulation; Molecular Structure; Phylogeny; Streptomyces
PubMed: 34267232
DOI: 10.1038/s41598-021-93285-7 -
Journal of Medicinal Chemistry Dec 1975The synthesis and biological activity of three 7-substituted actinomycin D derivatives are reported. Three such derivatives, 7-nitro-, 7-amino-, and 7-hydroxyactinomycin...
The synthesis and biological activity of three 7-substituted actinomycin D derivatives are reported. Three such derivatives, 7-nitro-, 7-amino-, and 7-hydroxyactinomycin D, were synthesized via new methods which were first tested successfully with a chromophore model system. Of these, 7-nitro- and 7-aminoactinomycin D were assayed for growth inhibitory activity against mammalian cells (CCRF-CEM human lymphoblastic leukemia) in vitro and against the Ridgway osteogenic sarcoma and the L1210, P1534, and P388 murine leukemias in vivo. In these systems, the inhibitory activity of the 7-substituted analogs was comparable to actinomycin D. In two bacterial systems ( (L. casei and L. arabinosus) in vitro, on the other hand, these compounds showed inhibitory profiles which are distinctly different from actinomycin D. These studies demonstrate that substitution at the 7 position, which does not interfere with DNA binding, is capable of yielding experimental antitumor agents with significant activity against a variety of tumors.
Topics: Animals; Dactinomycin; Humans; Lactobacillus; Leukemia L1210; Leukemia, Experimental; Magnetic Resonance Spectroscopy; Mice; Mice, Inbred Strains; Osteosarcoma; Sarcoma, Experimental; Structure-Activity Relationship
PubMed: 1059773
DOI: 10.1021/jm00246a001 -
Apoptosis : An International Journal on... Jun 2022Actinomycin D (ActD) was the first anticancer antibiotic approved for the management of human cancers. However, the notorious toxicity profile limits its widespread...
Actinomycin D (ActD) was the first anticancer antibiotic approved for the management of human cancers. However, the notorious toxicity profile limits its widespread application in cancers, including cancers of the aerodigestive tract. Recent studies show that combining low-dose ActD with existing chemotherapies could potentially protect normal cells from the toxicity of chemotherapy drugs through p53 activation (cyclotherapy). An understanding of ActD's effect on p53 signaling is critical for the meaningful application of ActD in cyclotherapy-based combinations. This study evaluated the anti-tumor efficacy and mechanism of action of ActD in aerodigestive tract cancers. We found that ActD strongly inhibited the growth of a panel of aerodigestive tract cancer cell lines and induced efficient apoptosis, although the sensitivity varies among cell lines. The IC values of ActD spanned between 0.021 and 2.96 nM. Mechanistic studies revealed that ActD increased the expression of total and phosphorylated p53 (ser15) in a time- and dose-dependent manner. Moreover, ActD-induced apoptosis is dependent on p53 in cells expressing wild-type p53 and that ActD induced context-dependent differential expression of downstream targets p21 and PUMA without significant effects on p27. In the final analysis, this study revealed that p53-p21 is the predominant pathway activated by low-dose ActD, supporting further development of ActD in cyclotherapy.
Topics: Antibiotics, Antineoplastic; Apoptosis; Dactinomycin; Humans; Neoplasms; Tumor Suppressor Protein p53
PubMed: 35267106
DOI: 10.1007/s10495-022-01720-5 -
Journal of Bacteriology Mar 1988Three distinct classes of mutations affecting the biosynthesis of actinomycin have been established in Streptomyces chyrsomallus by crossing various...
Three distinct classes of mutations affecting the biosynthesis of actinomycin have been established in Streptomyces chyrsomallus by crossing various actinomycin-nonproducing mutants with each other by protoplast fusion. In crosses between members of different classes of mutations, actinomycin-producing recombinant progeny arose, whereas in crosses between members of the same class, no actinomycin-producing recombinants were seen. Biochemical examination of a number of mutants revealed that the expression of all actinomycin synthetases was reduced by about 1 order of magnitude in mutants belonging to class II. In mutants of class I, the specific activities of the actinomycin synthetases were comparable with those measured in their actinomycin-producing parents. Feeding experiments with 4-methyl-3-hydroxyanthranilic acid (4-MHA), the biosynthetic precursor of the chromophore moiety of actinomycin, with representative mutants of the three genetic classes revealed formation of actinomycin in minute amounts by mutants of class I. It is suggested that mutants belonging to class I are mutated at a genetic locus involved in the biosynthesis of 4-MHA. Mutants belonging to class II appear to carry mutations at a locus involved in the regulation of the expression of the actinomycin synthetases. The role of the locus in class III mutations could not be assigned. Mapping studies in S. chrysomallus based on conjugal matings revealed the chromosomal linkage of all three loci. Mutations belonging to classes I and III were closely linked. Their genetic loci could be localized in a map interval of the chromosomal linkage group which is significantly distant from the gene locus represented by mutations belonging to class II.
Topics: Dactinomycin; Genes, Bacterial; Genetic Complementation Test; Genetic Linkage; Mutation; Streptomyces
PubMed: 2449423
DOI: 10.1128/jb.170.3.1360-1368.1988 -
Journal of Bacteriology Jun 1968Streptomyces antibioticus synthesizes a mixture of actinomycins which differ at the "imino acid" site of the peptide chains. In the presence of exogenous pipecolic acid,...
Streptomyces antibioticus synthesizes a mixture of actinomycins which differ at the "imino acid" site of the peptide chains. In the presence of exogenous pipecolic acid, several new actinomycins were synthesized and 70% of the proline in the antibiotic mixture was replaced by the analogue. Three new antibiotics (designated Pip 1alpha, Pip 1beta, and Pip 2) were isolated from culture filtrates, purified, and crystallized. The molar ratio of pipecolic acid to proline was: Pip 1alpha, 1:0; Pip 1beta, 1:1; Pip 2, 2:0. These compounds inhibited the growth and cell division of gram-positive, but not gram-negative, bacteria. The relative inhibitory activity against bacteria, Escherichia coli deoxyribonucleic acid (DNA)-dependent ribonucleic acid (RNA) polymerase in vitro, and RNA synthesis in Bacillus subtilis and mouse L-929 cells was: actinomycin IV = Pip 1beta > Pip 2 > Pip 1alpha. Protein synthesis in B. subtilis was less affected, and DNA synthesis was inhibited only at higher concentrations of antibiotic tested. In L cells, DNA formation was reduced less than RNA synthesis, whereas protein synthesis was not blocked under the experimental conditions employed. Kinetic studies with B. subtilis revealed that RNA synthesis was inhibited rapidly followed by an inhibition of protein synthesis. All four antibiotics markedly inhibited the replication of vaccinia virus and reovirus in tissue culture cells, but the production of poliovirus was resistant to the antibiotics. These actinomycins bind to DNA, resulting in an elevation of its T(m) and a decrease in the peak extinction of the actinomycins. The mode of action, as well as the structure-activity relationships among the actinomycins, are discussed relative to a previously proposed model of binding.
Topics: Bacillus subtilis; Bacterial Proteins; Carbon Isotopes; DNA; DNA, Bacterial; Dactinomycin; Escherichia coli; L Cells; Pipecolic Acids; Poliovirus; Proline; RNA; RNA Nucleotidyltransferases; RNA, Bacterial; Reoviridae; Sarcina; Spectrophotometry; Staphylococcus; Streptomyces; Thymidine; Uridine; Vaccinia virus; Valine; Virus Replication
PubMed: 4174667
DOI: 10.1128/jb.95.6.2139-2150.1968 -
Pharmacology, Biochemistry, and Behavior May 1976The influence of Actinomycin D (AMD) applied intrahippocampally at doses of 1-6 mug/animal, on the acquisition and retention of a shock-motivated brightness...
The influence of Actinomycin D (AMD) applied intrahippocampally at doses of 1-6 mug/animal, on the acquisition and retention of a shock-motivated brightness discrimination was studied on rats in a semiautomatic Y-maze. The injection of AMD 4 hr prior to training did not influence the acquisition, but causes, dose-dependent, a retention loss in relearning 24 hr after training. Twenty-eight hr after AMD application, naive rats exhibited a deterioration of acquisition performance increasing equally with the dose. At the same time, both circumscribed necroses in the hippocampus and signs of a general intoxication were observed. Considering the described pro- and retroactive effects, it is concluded that the use of the inhibitor AMD in learning experiments is not suitable to provide reliable evidence of the specific importance of the cerebral RNA synthesis for memory consolidation.
Topics: Animals; Body Temperature; Body Weight; Dactinomycin; Hippocampus; Injections; Injections, Intraventricular; Learning; Male; Memory; Rats; Time Factors
PubMed: 951429
DOI: 10.1016/0091-3057(76)90190-8 -
Biochemical Pharmacology Nov 1985Localization of protein B23 in HeLa cells after treatment with actinomycin D and its analogs was studied using indirect immunofluorescence. Bright nucleolar fluorescence...
Localization of protein B23 in HeLa cells after treatment with actinomycin D and its analogs was studied using indirect immunofluorescence. Bright nucleolar fluorescence was observed in control HeLa cells. After treatment with actinomycin D (250 ng/ml) for 2 hr, a uniform nucleoplasmic fluorescence was observed. Similar results were obtained with the actinomycin analogs, actinomycin Z5 and actinomycin K2T. Only after a much longer incubation (24 hr) with actinomycin 4-4'-gly was nucleoplasmic fluorescence observed. Actinomycin D, actinomycin Z5, and actinomycin K2T inhibited [3H]uridine incorporation into the trichloroacetic acid insoluble fraction of HeLa cells with IC50 values of 9.5 +/- 3.2, 59.1 +/- 19.6 and 1423.3 +/- 212.2 ng/ml respectively. No inhibition of [3H]uridine incorporation was observed using actinomycin 4-4'-gly (2000 ng/ml, 2-hr incubation). The order of potency for the loss of nucleolar fluorescence and the concurrent increase in nucleoplasmic fluorescence was actinomycin D greater than actinomycin Z5 greater than actinomycin K2T greater than actinomycin 4-4'-gly, which correlated with the order of their IC50 values for inhibition of [3H]uridine incorporation. Studies of the effects of actinomycin D and its analogs on RNA synthesis and localization of protein B23 indicated that there is a direct relationship between the B23 "translocation" from nucleolus to nucleoplasm and the inhibition of RNA synthesis. At 45-55% inhibition of RNA synthesis, both nuclear and nucleolar B23 immunofluorescence were observed. At 75-85% inhibition, only a uniform nucleoplasmic fluorescence was observed.
Topics: Cell Nucleolus; Dactinomycin; HeLa Cells; Humans; Nuclear Proteins; Nucleophosmin; Nucleoproteins; Phosphoproteins; RNA; Ribonucleoproteins; Time Factors
PubMed: 2415133
DOI: 10.1016/0006-2952(85)90387-9 -
Journal of Medicinal Chemistry Jul 19791,4-Oxazinone derivatives of the phenoxazinone chromophore in actinomycin D (AMD) have been synthesized by condensation of AMD with alpha-keto acids. By varying the...
1,4-Oxazinone derivatives of the phenoxazinone chromophore in actinomycin D (AMD) have been synthesized by condensation of AMD with alpha-keto acids. By varying the starting alpha-keto acid, the substitutions on the oxazinone ring and, consequently, the lipophilicity of the molecule could be altered. These oxazinone derivatives revert to AMD in physiological media and it appears that these oxazinones are "depot" forms of AMD and possess physicochemical and DNA-binding properties which are significantly different from those of AMD. The oxazinones, which have bulky and lipophilic substituents at position 3, demonstrate more pronounced antitumor activity against P388 mouse leukemia and are less toxic than AMD.
Topics: Animals; Antineoplastic Agents; Chemical Phenomena; Chemistry; Chemistry, Physical; DNA; Dactinomycin; Dose-Response Relationship, Drug; Esterases; Humans; In Vitro Techniques; Leukemia, Experimental; Male; Mice; Rats; Structure-Activity Relationship
PubMed: 448678
DOI: 10.1021/jm00193a009 -
Biophysical Journal Oct 2000Earlier calorimetric studies had indicated that despite the absence of a GpC sequence, the self-complementary octamer d(CGTCGACG) binds strongly to actinomycin D (ACTD)... (Comparative Study)
Comparative Study
Earlier calorimetric studies had indicated that despite the absence of a GpC sequence, the self-complementary octamer d(CGTCGACG) binds strongly to actinomycin D (ACTD) with high cooperativity and a 2:1 drug/duplex ratio. A subsequent optical spectral study with related oligomers led us to suggest that ACTD may likely stack at the G. C basepairs of the duplex termini. New findings are reported herein to indicate that despite the lack of complete self-complementarity, oligomers of d(CGXCGXCG) [X = A or T] motif exhibit unusually strong ACTD affinities with binding constants of roughly 2 x 10(7) M(-1) and binding densities of 1 drug molecule per strand. The ACTD binding affinity for the corresponding heteroduplex obtained by annealing these two oligomers is, however, considerably reduced. Although spectroscopic results with related oligomers obtained by removing, replacing, or appending bases at the termini appear to be consistent with the end-stacking model, capillary electrophoretic (CE) evidence provides additional insights into the binding mode. CE experiments with the self-complementary oligomers d(CGAGCTCG) and d(CGTCGACG) revealed contrasting migration patterns in the presence of ACTD, with mobility retardation and acceleration exhibited by the GpC- and non-GpC-containing octamers, respectively, whereas the X/X-mismatched d(CGXCGXCG) experienced retardation. These results, along with those of related oligomers, suggest that ACTD may in fact stack at the duplex stem end of a monomeric hairpin or at the 3'-end of dG as a single strand. The seemingly cooperative ACTD binding and the curved Scatchard plot for the self-complementary d(CGTCGACG) may thus be attributed to the drug-induced duplex denaturation resulting from strong binding to single strands of d(CGXCGYCG) motif. Detailed structural information on the ACTD-DNA complexes, however, must await further NMR investigations.
Topics: Antibiotics, Antineoplastic; Base Sequence; Binding Sites; Biophysical Phenomena; Biophysics; Circular Dichroism; Dactinomycin; Electrophoresis, Capillary; In Vitro Techniques; Kinetics; Oligodeoxyribonucleotides; Spectrometry, Fluorescence
PubMed: 11023913
DOI: 10.1016/S0006-3495(00)76457-5 -
Journal of Cellular Physiology Dec 1992Pretreatment plus concomitant treatment with 10 micrograms/ml cycloheximide protected Chinese hamster ovary cells and Swiss 3T3 cells against the cytotoxicity of...
Pretreatment plus concomitant treatment with 10 micrograms/ml cycloheximide protected Chinese hamster ovary cells and Swiss 3T3 cells against the cytotoxicity of actinomycin D. The cycloheximide treatment reduced the intracellular concentration of actinomycin D by reducing the level of actinomycin D bound to the acid precipitable fraction of the cell. Levels of unbound actinomycin D were unaffected by cycloheximide, indicating that the plasma membrane permeability to AD was not reduced. Actinomycin D inhibited total transcription but did not reduce cytoplasmic levels of rRNA nor of most tested mRNA; however, cytoplasmic levels of c-myc mRNA were reduced below detectability. Cycloheximide treatment further inhibited total transcription and had no effect on cytoplasmic levels of rRNA nor of most tested mRNA. Cytoplasmic levels of c-myc were elevated by cycloheximide and remained so even in the presence of actinomycin D. These data suggested that a reduction in cytoplasmic levels of short lived, essential mRNA, such as c-myc mRNA, was one lethal lesion of actinomycin D. Furthermore, cycloheximide's protection may result, in part, from its ability to stabilize and/or elevate cytoplasmic levels of these mRNA, thus counteracting their depletion by actinomycin D. Protection may also result from the cycloheximide-induced reduction of actinomycin D bound to the acid precipitable fraction of the cells.
Topics: Animals; Blotting, Northern; CHO Cells; Cell Line; Cell Survival; Cricetinae; Cycloheximide; Dactinomycin; Intracellular Membranes; RNA; Temperature; Uridine
PubMed: 1280278
DOI: 10.1002/jcp.1041530310