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Cellular Physiology and Biochemistry :... 2017Although proinflammatory cytokine-induced disruption of intestinal epithelial barrier integrity is associated with intestinal inflammatory disease, effective treatment...
BACKGROUND/AIMS
Although proinflammatory cytokine-induced disruption of intestinal epithelial barrier integrity is associated with intestinal inflammatory disease, effective treatment for barrier dysfunction is lacking. Previously, we demonstrated that rebeccamycin alleviates epithelial barrier dysfunction induced by inflammatory cytokines in Caco-2 cell monolayers; however, the underlying mechanism remained unclear. Here, we investigated the mechanism by which rebeccamycin protects the epithelial barrier function of Caco-2 cells exposed to TNF-α.
METHODS
To confirm the epithelial barrier function of Caco-2 cell monolayers, transepithelial electrical resistance (TER) and paracellular permeability were measured. Production levels and localization of tight junction (TJ) proteins were analyzed by immunoblot and immunofluorescence, respectively. Phosphorylated myosin light chain (pMLC) and MLC kinase (MLCK) mRNA expression levels were determined by immunoblot and quantitative RT-PCR, respectively.
RESULTS
Rebeccamycin attenuated the TNF-α-induced reduction in TER and increase in paracellular permeability. Rebeccamycin increased claudin-5 expression, but not claudin-1, -2, -4, occludin or ZO-1 expression, and prevented the TNF-α-induced changes in ZO-1 and occludin localization. Rebeccamycin suppressed the TNF-α-induced increase in MLCK mRNA expression, thus suppressing MLC phosphorylation. The rebeccamycin-mediated reduction in MLCK production and protection of epithelial barrier function were alleviated by Chk1 inhibition.
CONCLUSION
Rebeccamycin attenuates TNF-α-induced disruption of intestinal epithelial barrier integrity by inducing claudin-5 expression and suppressing MLCK production via Chk1 activation.
Topics: Caco-2 Cells; Carbazoles; Checkpoint Kinase 1; Claudin-5; Enzyme Activation; Gene Expression Regulation, Enzymologic; Humans; Intestinal Mucosa; Myosin Light Chains; Myosin-Light-Chain Kinase; Phosphorylation; Tight Junctions; Tumor Necrosis Factor-alpha
PubMed: 28391269
DOI: 10.1159/000472367 -
Marine Drugs Dec 2009Granulatimide and isogranulatimide are alkaloids obtained from marine sources which have been shown to inhibit cell-cycle G2-checkpoint, targeting more particularly... (Review)
Review
Granulatimide and isogranulatimide are alkaloids obtained from marine sources which have been shown to inhibit cell-cycle G2-checkpoint, targeting more particularly checkpoint 1 kinase (Chk1). At a structural level, they possess a characteristic pyrrolocarbazole framework also shared by the well-known rebeccamycin and staurosporine microbial metabolites which have been described to inhibit topoisomerase I and diverse kinases, respectively. This review reports precisely on the synthesis and kinase inhibitory activities of pyrrolocarbazole-based analogues of granulatimide.
Topics: Alkaloids; Carbazoles; Checkpoint Kinase 1; Humans; Imidazoles; Indoles; Marine Biology; Protein Kinase Inhibitors; Protein Kinases; Pyrroles
PubMed: 20098609
DOI: 10.3390/md7040754 -
PloS One 2016Several stressors are known to influence epithelial tight junction (TJ) integrity, but the association between DNA damage and TJ integrity remains unclear. Here we...
Several stressors are known to influence epithelial tight junction (TJ) integrity, but the association between DNA damage and TJ integrity remains unclear. Here we examined the effects of daunorubicin and rebeccamycin, two anti-tumor chemicals that induce DNA damage, on TJ integrity in human intestinal epithelial cells. Daunorubicin and rebeccamycin dose-dependently enhanced transepithelial electrical resistance (TER) and decreased flux of the 4 kDa FITC-dextran in Caco-2 cell monolayer. Daunorubicin- or rebeccamycin-induced enhancement of the TJ barrier function partly rescued attenuation of the barrier function by the inflammatory cytokines TNF-α and IFN-γ. Daunorubicin and rebeccamycin increased claudin-5 expression and the product was distributed in the actin cytoskeleton fraction, which was enriched with TJ proteins. Caffeine, which is an inhibitor of ataxia telangiectasia mutated protein (ATM) and ataxia telangiectasia mutated and Rad3-related protein (ATR), and the Chk1 inhibitor inhibited the TER increases induced by daunorubicin and rebeccamycin, whereas a Chk2 inhibitor did not. Treatment with Chk1 siRNA also significantly inhibited the TER increases. Induction of claudin-5 expression was inhibited by Chk1 inhibitor and by siRNA treatment. Our results suggest that Chk1 activation by daunorubicin and rebeccamycin induced claudin-5 expression and enhanced TJ barrier function in Caco-2 cell monolayer, which suggests a link between DNA damage and TJ integrity in the human intestine.
Topics: Caco-2 Cells; Carbazoles; Checkpoint Kinase 1; Claudin-5; Daunorubicin; Enzyme Activation; Gene Expression Regulation, Neoplastic; Humans; Intestinal Mucosa; Protein Kinases
PubMed: 26727128
DOI: 10.1371/journal.pone.0145631 -
Molecules (Basel, Switzerland) Oct 2022The present study is to explore the anticancer effect of loonamycin (LM) in vitro and in vivo, and investigate the underlying mechanism with combined multi-omics. LM...
The present study is to explore the anticancer effect of loonamycin (LM) in vitro and in vivo, and investigate the underlying mechanism with combined multi-omics. LM exhibited anticancer activity in human triple negative breast cancer cells by promoting cell apoptosis. LM administration inhibited the growth of MDA-MB-468 tumors in a murine xenograft model of breast cancer. Mechanistic studies suggested that LM could inhibit the topoisomerase I in a dose-dependent manner in vitro experiments. Combined with the transcriptomics and proteomic analysis, LM has a significant effect on O-glycan, p53-related signal pathway and EGFR/PI3K/AKT/mTOR signal pathway in enrichment of the KEGG pathway. The GSEA data also suggests that the TNBC cells treated with LM may be regulated by p53, O-glycan and EGFR/PI3K/AKT/mTOR signaling pathway. Taken together, our findings predicted that LM may target p53 and EGFR/PI3K/AKT/mTOR signaling pathway, inhibiting topoisomerase to exhibit its anticancer effect.
Topics: Humans; Mice; Animals; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; DNA Topoisomerases, Type I; Tumor Suppressor Protein p53; Transcriptome; Proteomics; Cell Line, Tumor; TOR Serine-Threonine Kinases; Triple Negative Breast Neoplasms; Apoptosis; ErbB Receptors; Cell Proliferation
PubMed: 36296549
DOI: 10.3390/molecules27206958 -
Biomolecules Apr 2020Chagas disease and leishmaniasis are neglected tropical diseases caused by kinetoplastid parasites of and genera that affect poor and remote populations in developing...
Chagas disease and leishmaniasis are neglected tropical diseases caused by kinetoplastid parasites of and genera that affect poor and remote populations in developing countries. These parasites share similar complex life cycles and modes of infection. It has been demonstrated that the particular group of phosphorylating enzymes, protein kinases (PKs), are essential for the infective mechanisms and for parasite survival. The natural indolocarbazole staurosporine (STS, ) has been extensively used as a PKC inhibitor and its antiparasitic effects described. In this research, we analyze the antikinetoplastid activities of three indolocarbazole (ICZs) alkaloids of the family of staurosporine STS, -, and the commercial ICZs rebeccamycin (), K252a (), K252b (), K252c (), and arcyriaflavin A () in order to establish a plausive approach to the mode of action and to provide a preliminary qualitative structure-activity analysis. The most active compound was 7-oxostaurosporine (7OSTS, ) that showed IC values of 3.58 ± 1.10; 0.56 ± 0.06 and 1.58 ± 0.52 µM against and and a Selectivity Index (CC/IC) of 52 against amastigotes of compared to the J774A.1 cell line of mouse macrophages.
Topics: Animals; Antiprotozoal Agents; Biological Assay; Carbazoles; Cell Death; Cell Line; Inhibitory Concentration 50; Kinetoplastida; Membrane Potential, Mitochondrial; Metabolome; Mice; Mitochondria; Models, Molecular; Parasites; Staurosporine; Streptomyces; Toxicity Tests
PubMed: 32344693
DOI: 10.3390/biom10040657 -
Bioscience, Biotechnology, and... Oct 2009The biosynthesis of staurosporine, rebeccamycin, and goadsporin, which are produced by actinomycetes and contain characteristic heterocyclic rings, was characterized by... (Review)
Review
The biosynthesis of staurosporine, rebeccamycin, and goadsporin, which are produced by actinomycetes and contain characteristic heterocyclic rings, was characterized by genetic methods. Staurosporine and rebeccamycin contain an indolocarbazole ring synthesized from two molecules of tryptophan, with indolepyruvic acid imine and chromopyrrolic acid as biosynthetic intermediates. A tetrameric hemoprotein synthesizes chromopyrrolic acid, and cytochrome P450 peroxidase catalyzes the intramolecular C-C coupling and decarboxylation of chromopyrrolic acid to form the indolocarbazole core. Goadsporin is a thiopeptide containing thiazole and oxazole heterocyclic rings. The structural gene godA is ribosomally translated to a goadsporin precursor peptide, and oxazole, methyloxazole, and thiazole rings are derived from serine, threonine, and cystein through post-translational modifications. On the basis of these knowledges, a wide variety of indolocarbazole and goadsporin analogs through the rational gene recombination and disruption of these biosynthetic genes were successfully produced.
Topics: Actinobacteria; Animals; Anti-Bacterial Agents; Carbazoles; Humans; Intercellular Signaling Peptides and Proteins; Peptides; Staurosporine
PubMed: 19809190
DOI: 10.1271/bbb.90263 -
Frontiers in Bioengineering and... 2023, as an actinomycete, is a natural producer of the antibiotic and antitumoral drug rebeccamycin. Due to the filamentous cellular morphology handling in cultivations is...
Morphology engineering for novel antibiotics: Effect of glass microparticles and soy lecithin on rebeccamycin production and cellular morphology of filamentous actinomycete .
, as an actinomycete, is a natural producer of the antibiotic and antitumoral drug rebeccamycin. Due to the filamentous cellular morphology handling in cultivations is challenging; therefore, morphology engineering techniques are mandatory to enhance productivity. One promising approach described in the literature is the addition of mineral particles in the micrometer range to precisely adjust cellular morphology and the corresponding product synthesis (microparticle-enhanced cultivation, MPEC). Glass microparticles are introduced in this study as a novel supplementation type for bioprocess intensification in filamentous organisms. Several investigations were conducted to screen for an optimal particle setup, including particle size and concentration regarding their impact and effects on enhanced productivity, microparticle incorporation behavior into the biopellets, the viability of pellets, and morphological changes. Glass microparticles (10 g·L) with a median diameter of 7.9 µm, for instance, induced an up to fourfold increase in product synthesis accompanied by overall enhanced viability of biomass. Furthermore, structural elucidations showed that biopellets isolated from MPEC tend to have lower hyphal density than unsupplemented control pellets. In this context, oxygen microprofiling was conducted to better understand how internal structural changes interwind with oxygen supply into the pellets. Here, the resulting oxygen profiles are of a contradictive trend of steeper oxygen consumption with increasing glass microparticle supplementation. Eventually, MPEC was combined with another promising cultivation strategy, the supplementation of soy lecithin (7.5 g·L), to further increase the cultivation performance. A combination of both techniques in an optimized setup resulted in a rebeccamycin concentration of 213 mg·L after 10 days of cultivation, the highest value published so far for microparticle-supplemented shake flask cultivations of .
PubMed: 37091334
DOI: 10.3389/fbioe.2023.1171055 -
The Journal of Antibiotics May 1987An actinomycete, strain C-38,383, was selected in a screening program for the isolation of novel antitumor agents. A yellow crystalline product, named rebeccamycin, was...
An actinomycete, strain C-38,383, was selected in a screening program for the isolation of novel antitumor agents. A yellow crystalline product, named rebeccamycin, was isolated from the mycelium and was found to have activity against P388 leukemia, L1210 leukemia and B16 melanoma implanted in mice. Rebeccamycin inhibits the growth of human lung adenocarcinoma cells (A549) and produces single-strand breaks in the DNA of these cells. No DNA-protein cross-links were detected. A related antibiotic, staurosporine, is produced by Streptomyces staurosporeus and Streptomyces actuosus. Strain C-38,383 was found to resemble closely strains of Nocardia aerocolonigenes recently renamed Saccharothrix aerocolonigenes. A strain selection isolate without aerial mycelium, C-38,383-RK-1, failed to produce rebeccamycin while a strain with aerial mycelium, C-38,383-RK-2, was found to be a suitable strain for production. A description of the producing strain is presented and its taxonomic position is reviewed. A fermentor containing 37 liters of production medium gave a rebeccamycin yield of 663 mg/liter after 204 hours of incubation with strain C-38,383-RK-2.
Topics: Actinomycetales; Adenocarcinoma; Aminoglycosides; Animals; Anti-Bacterial Agents; Antibiotics, Antineoplastic; Carbazoles; Carbohydrate Metabolism; Cell Survival; Cells, Cultured; Cisplatin; DNA Damage; Glycosides; Humans; Indoles; Intercalating Agents; Leukemia L1210; Leukemia P388; Melanoma, Experimental; Mice; Microbial Sensitivity Tests; Mitomycin; Mitomycins; Nucleic Acids; Protein Biosynthesis
PubMed: 3112080
DOI: 10.7164/antibiotics.40.668 -
Cancer Chemotherapy and Pharmacology Dec 2009Advanced cancers of the bile duct and gallbladder carry an ominous prognosis. Rebeccamycin analogue (RA) is a novel antitumor antibiotic where phase I trials suggested...
PURPOSE
Advanced cancers of the bile duct and gallbladder carry an ominous prognosis. Rebeccamycin analogue (RA) is a novel antitumor antibiotic where phase I trials suggested clinical efficacy in patients with biliary cancers.
METHODS
The primary objective was to determine the response rate to RA in patients with advanced gallbladder and bile duct tumors. Secondary endpoints were survival and pharmacokinetic characterization. RA was given at a dose 165 mg/(m(2) day) x 5 days every 3 weeks.
RESULTS
Forty-six patients were enrolled. Nine patients were removed from study before their first planned imaging study for response. Two patients had partial responses and 16 had stable disease. On an intent-to-treat analysis the median survival was 6.3 months. A >20% drop in CA19.9 was seen in 43% of patients with initial high levels. Grade 4 neutropenia and thrombocytopenia were seen in 35 and 5% of patients, respectively. Febrile neutropenia occurred in 16% of patients. The pharmacokinetic profile of this trial closely resembles those of prior phase I trials. Measured biliary concentrations of RA were as much as 100x greater than simultaneous plasma concentration.
CONCLUSION
Although RA has a response rate of 5% in advanced biliary cancers, it is associated with significant numbers of patients experiencing prolonged stable disease. Biliary concentrations of RA are significantly greater than plasma concentrations.
Topics: Aged; Antineoplastic Agents; Bile Duct Neoplasms; CA-19-9 Antigen; Carbazoles; Female; Fever; Gallbladder Neoplasms; Glucosides; Humans; Male; Middle Aged; Neutropenia; Survival Rate; Thrombocytopenia
PubMed: 19399502
DOI: 10.1007/s00280-009-1005-x -
Proceedings of the National Academy of... Jan 2005Rebeccamycin and staurosporine are natural products with antitumor properties, which belong to the family of indolocarbazole alkaloids. An intense effort currently...
Rebeccamycin and staurosporine are natural products with antitumor properties, which belong to the family of indolocarbazole alkaloids. An intense effort currently exists for the generation of indolocarbazole derivatives for the treatment of several diseases, including cancer and neurodegenerative disorders. Here, we report a biological process based on combinatorial biosynthesis for the production of indolocarbazole compounds (or their precursors) in engineered microorganisms as a complementary approach to chemical synthesis. We have dissected and reconstituted the entire biosynthetic pathway for rebeccamycin in a convenient actinomycete host, Streptomyces albus. This task was achieved by coexpressing different combinations of genes isolated from the rebeccamycin-producing microorganism. Also, a gene (staC) was identified in staurosporine-producing microbes and was shown to have a key role to differentiate the biosynthetic pathways for the two indolocarbazoles. Last, incorporation of the pyrH and thal genes, encoding halogenases from different microorganisms, resulted in production of derivatives with chlorine atoms at novel positions. We produced >30 different compounds by using the recombinant strains generated in this work.
Topics: Antineoplastic Agents; Carbazoles; Combinatorial Chemistry Techniques; Indoles; Staurosporine; Streptomyces
PubMed: 15625109
DOI: 10.1073/pnas.0407809102