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MBio Oct 2022The fungus Rhizopus microsporus harbors a bacterial endosymbiont () for the production of the antimitotic toxin rhizoxin. Although rhizoxin is the causative agent of...
The fungus Rhizopus microsporus harbors a bacterial endosymbiont () for the production of the antimitotic toxin rhizoxin. Although rhizoxin is the causative agent of rice seedling blight, the toxinogenic bacterial-fungal alliance is, not restricted to the plant disease. It has been detected in numerous environmental isolates from geographically distinct sites covering all five continents, thus raising questions regarding the ecological role of rhizoxin beyond rice seedling blight. Here, we show that rhizoxin serves the fungal host in fending off protozoan and metazoan predators. Fluorescence microscopy and coculture experiments with the fungivorous amoeba revealed that ingestion of R. microsporus spores is toxic to . This amoebicidal effect is caused by the dominant bacterial rhizoxin congener rhizoxin S2, which is also lethal toward the model nematode Caenorhabditis elegans. By combining stereomicroscopy, automated image analysis, and quantification of nematode movement, we show that the fungivorous nematode Aphelenchus avenae actively feeds on R. microsporus that is lacking endosymbionts, whereas worms coincubated with symbiotic R. microsporus are significantly less lively. This study uncovers an unexpected ecological role of rhizoxin as shield against micropredators. This finding suggests that predators may function as an evolutionary driving force to maintain toxin-producing endosymbionts in nonpathogenic fungi. The soil community is a complex system characterized by predator-prey interactions. Fungi have developed effective strategies to defend themselves against predators. Understanding these strategies is of critical importance for ecology, medicine, and biotechnology. In this study, we shed light on the defense mechanisms of the phytopathogenic - symbiosis that has spread worldwide. We report an unexpected role of rhizoxin, a secondary metabolite produced by the bacterium residing within the hyphae of R. microsporus. We show that this bacterial secondary metabolite is utilized by the fungal host to successfully fend off fungivorous protozoan and metazoan predators and thus identified a fundamentally new function of this infamous cytotoxic compound. This endosymbiont-dependent predator defense illustrates an unusual strategy employed by fungi that has broader implications, since it may serve as a model for understanding how animal predation acts as an evolutionary driving force to maintain endosymbionts in nonpathogenic fungi.
Topics: Animals; Burkholderia; Antimitotic Agents; Macrolides; Symbiosis; Oryza; Toxins, Biological; Seedlings; Soil
PubMed: 36005392
DOI: 10.1128/mbio.01440-22 -
Archives of Biochemistry and Biophysics Sep 2022A tritiated derivative of the sponge-derived natural product spongistatin 1 was prepared, and its interactions with tubulin were examined. [H]Spongistatin 1 was found to...
A tritiated derivative of the sponge-derived natural product spongistatin 1 was prepared, and its interactions with tubulin were examined. [H]Spongistatin 1 was found to bind rapidly to tubulin at a single site (the low specific activity of the [H]spongistatin 1, 0.75 Ci/mmol, prevented our defining an association rate), and the inability of spongistatin 1 to cause an aberrant assembly reaction was confirmed. Spongistatin 1 bound to tubulin very tightly, and we could detect no significant dissociation reaction from tubulin. The tubulin-[H]spongistatin 1 complex did dissociate in 8 M urea, so there was no evidence for covalent bond formation. Apparent K values were obtained by Scatchard analysis of binding data and by Hummel-Dreyer chromatography (3.5 and 1.1 μM, respectively). The effects of a large cohort of vinca domain drugs on the binding of [H]spongistatin 1 to tubulin were evaluated. Compounds that did not cause aberrant assembly reactions (halichondrin B, eribulin, maytansine, and rhizoxin) caused little inhibition of [H]spongistatin 1 binding. Little inhibition also occurred with the peptides dolastatin 15, its active pentapeptide derivative, vitilevuamide, or diazonamide A, nor with the vinca alkaloid vinblastine. Strong inhibition was observed with dolastatin 10, hemiasterlin, and cryptophycin 1, all of which cause aberrant assembly reactions that might actually mask the spongistatin 1 binding site. Spongistatin 5 was found to be a competitive inhibitor of [H]spongistatin 1 binding, with an apparent K of 2.2 μM. We propose that the strong picomolar cytotoxicity of spongistatin 1 probably derives from its extremely tight binding to tubulin.
Topics: Antineoplastic Agents; Binding Sites; Macrolides; Microtubules; Tubulin; Vinblastine
PubMed: 35594923
DOI: 10.1016/j.abb.2022.109296 -
Dose-response : a Publication of... 2019The role of viral infection in developing cancer was determined in the start of 20th century. Until now, 8 different virus-associated cancers have been discovered and... (Review)
Review
The role of viral infection in developing cancer was determined in the start of 20th century. Until now, 8 different virus-associated cancers have been discovered and most of them progressed in immunosuppressed individuals. The aim of the present study is to look into the benefits of natural products in treating virally infected cancers. The study focuses on bioactive compounds derived from natural sources. Numerous pharmaceutical agents have been identified from plants (vincristine, vinblastine, stilbenes, combretastatin, and silymarin), marine organisms (bryostatins, cephalostatin, ecteinascidins, didemnin, and dolastatin), insects (cantharidin, mastoparan, parectadial, and cecropins), and microorganisms (vancomycin, rhizoxin, ansamitocins, mitomycin, and rapamycin). Beside these, various compounds have been observed from fruits and vegetables which can be utilized in anticancer therapy. These include curcumin in turmeric, resveratrol in red grapes, S-allyl cysteine in allium, allicin in garlic, catechins in green tea, and β-carotene in carrots. The present study addresses various types of virally infected cancers, their mechanism of action, and the role of different cell surface molecules elicited during viral binding and entry into the target cell along with the anticancer drugs derived from natural products by targeting screening of bioactive compounds from natural sources.
PubMed: 30670935
DOI: 10.1177/1559325818813227 -
Journal of Natural Products Mar 2009This review provides an overview of the discovery, structures, and biological activities of anticancer natural products that act by inhibiting or promoting the assembly... (Review)
Review
This review provides an overview of the discovery, structures, and biological activities of anticancer natural products that act by inhibiting or promoting the assembly of tubulin to microtubules. The emphasis is on providing recent information on those compounds in clinical use or in advanced clinical trials. The vinca alkaloids, the combretastatins, NPI-2358, the halichondrin B analogue eribulin, dolastatin 10, noscapine, hemiasterlin, and rhizoxin are discussed as tubulin polymerization inhibitors, while the taxanes and the epothilones are the major classes of tubulin polymerization promoters presented, with brief treatments of discodermolide, eleutherobin, and laulimalide. The challenges and future directions of tubulin-interactive natural products-based drug discovery programs are also discussed briefly.
Topics: Antineoplastic Agents; Biological Products; Diketopiperazines; Imidazoles; Microtubules; Molecular Structure; Piperazines; Tubulin; Vinca Alkaloids
PubMed: 19125622
DOI: 10.1021/np800568j -
Medicinal Research Reviews Sep 2015Although fungi produce highly structurally diverse metabolites, many of which have served as excellent sources of pharmaceuticals, no fungi-derived agent has been... (Review)
Review
Although fungi produce highly structurally diverse metabolites, many of which have served as excellent sources of pharmaceuticals, no fungi-derived agent has been approved as a cancer drug so far. This is despite a tremendous amount of research being aimed at the identification of fungal metabolites with promising anticancer activities. This review discusses the results of clinical testing of fungal metabolites and their synthetic derivatives, with the goal to evaluate how far we are from an approved cancer drug of fungal origin. Also, because in vivo studies in animal models are predictive of the efficacy and toxicity of a given compound in a clinical situation, literature describing animal cancer testing of compounds of fungal origin is reviewed as well. Agents showing the potential to advance to clinical trials are also identified. Finally, the technological challenges involved in the exploitation of fungal biodiversity and procurement of sufficient quantities of clinical candidates are discussed, and potential solutions that could be pursued by researchers are highlighted.
Topics: Androstadienes; Animals; Antineoplastic Agents; Aphidicolin; Biological Products; Clinical Trials as Topic; Cyclohexanes; Diketopiperazines; Disease Models, Animal; Drug Design; Drug Resistance, Neoplasm; Fatty Acids, Unsaturated; Female; Fungi; Humans; Macrolides; Male; Mice; Neoplasms; Polycyclic Sesquiterpenes; Sesquiterpenes; Trichothecenes; Wortmannin
PubMed: 25850821
DOI: 10.1002/med.21348 -
Annals of Oncology : Official Journal... Mar 2000Rhizoxin (NSC 332598) is a novel macrolide antitumor antibiotic that inhibits microtubule assembly and also depolymerizes preformed microtubules. In preclinical... (Clinical Trial)
Clinical Trial
BACKGROUND
Rhizoxin (NSC 332598) is a novel macrolide antitumor antibiotic that inhibits microtubule assembly and also depolymerizes preformed microtubules. In preclinical evaluations, rhizoxin demonstrated broad antitumor activity in vitro and in vivo including both vincristine- and vindesine-resistant human lung cancers. Prolonged exposure schedules in xenograft models demonstrated optimal efficacy indicating schedule-dependent antitumor activity. The early phase I and II evaluations a five-minute bolus infusion schedule was studied, however, only modest anti-tumor activity was noted, possibly due to rapid systemic clearance. To overcome these limitations and to exploit the potential for schedule-dependent behavior of rhizoxin, the feasibility of administering rhizoxin as a 72-hour continuous intravenous (i.v.) infusion was evaluated.
PATIENTS AND METHODS
Patients with advanced solid malignancies were entered into this phase I study, in which both the infusion duration and dose of rhizoxin were increased. The starting dose was 0.2 mg/m2 over 12 hours administered every 3 weeks. In each successive dose level, the dose and infusion duration were incrementally increased in a stepwise fashion. Once a 72-hour i.v. infusion duration was reached, rhizoxin dose-escalations alone continued until a maximum tolerated dose (MTD) was determined.
RESULTS
Nineteen patients were entered into the study. Rhizoxin was administered at doses ranging from 0.2 mg/m2 i.v. over 12 hours to 2.4 mg/m2 i.v. over 72 hours every 3 weeks. The principal dose-limiting toxicities (DLT) were severe neutropenia and mucositis, and the incidence of DLT was unacceptably high at rhizoxin doses above 1.2 mg/m2, which was determined to be the MTD and dose recommended for phase II studies. At these dose levels, rhizoxin could not be detected in the plasma by a previously validated and sensitive high-performance liquid chromatography assay with a lower limit of detection of 1 ng/ml. No antitumor responses were observed.
CONCLUSIONS
Rhizoxin can be safely administered using a 72-hour i.v. infusion schedule. The toxicity profile is similar to that observed previously using brief infusion schedules. Using this protracted i.v. infusion schedule the maximum tolerated dose is 1.2 mg/m2/72 hours.
Topics: Adult; Aged; Antibiotics, Antineoplastic; Humans; In Vitro Techniques; Lactones; Macrolides; Middle Aged; Neoplasms; Neutropenia
PubMed: 10811501
DOI: 10.1023/a:1008398725442 -
British Journal of Cancer Dec 1996Rhizoxin is a macrocyclic lactone compound that binds to tubulin and inhibits microtubule assembly. Rhizoxin demonstrated preclinical anti-tumour activity against a... (Clinical Trial)
Clinical Trial
Rhizoxin is a macrocyclic lactone compound that binds to tubulin and inhibits microtubule assembly. Rhizoxin demonstrated preclinical anti-tumour activity against a variety of human tumour cell lines and xenograft models. Phase I evaluation found a maximum tolerated rhizoxin dose of 2.6 mg m-2, with reversible, but dose-limiting, mucositis, leucopenia and diarrhoea. Clinical trials were then initiated by the EORTC ECSG in melanoma, breast, head and neck, and non-small-cell lung cancers with the recommended phase II rhizoxin dose of 2 mg m-2. Pharmacological studies were instituted with the phase II trials to complement the limited pharmacokinetic data available from the phase I trial. Blood samples were obtained from 69 of 103 eligible patients enrolled in phase II rhizoxin studies, and these were evaluable for pharmacokinetic analysis in 36 patients. Plasma rhizoxin concentrations were determined by high-performance liquid chromatography (HPLC), and post-distribution pharmacokinetic parameters were estimated by a one-compartment model. Rhizoxin was rapidly eliminated from plasma, with a median systemic clearance of 8.41 min-1 m-2 and an elimination half-life of 10.4 min. Rhizoxin area under the concentration-time curve (AUC) was higher in patients obtaining a partial response or stable disease than in those with progressive disease (median 314 vs 222 ng ml-1 min; P = 0.03). As predicted from previous studies, haematological and gastrointestinal toxicity was observed, but could not be shown to be related to rhizoxin AUC. This study demonstrated the rapid and variable elimination of rhizoxin from the systemic circulation. The presence of pharmacodynamic relationships and the low level of systemic toxicity suggest that future trials of rhizoxin with alternative dosage or treatment schedules are warranted.
Topics: Antibiotics, Antineoplastic; Area Under Curve; Breast Neoplasms; Carcinoma, Non-Small-Cell Lung; Female; Head and Neck Neoplasms; Humans; Injections, Intravenous; Lactones; Lung Neoplasms; Macrolides; Male; Melanoma; Treatment Outcome
PubMed: 8980394
DOI: 10.1038/bjc.1996.657 -
Molecules (Basel, Switzerland) Oct 2020Efforts are described towards the total synthesis of the bacterial macrolide rhizoxin F, which is a potent tubulin assembly and cancer cell growth inhibitor. A...
Efforts are described towards the total synthesis of the bacterial macrolide rhizoxin F, which is a potent tubulin assembly and cancer cell growth inhibitor. A significant amount of work was expanded on the construction of the rhizoxin core macrocycle by ring-closing olefin metathesis (RCM) between C(9) and C(10), either directly or by using relay substrates, but in no case was ring-closure achieved. Macrocycle formation was possible by ring-closing alkyne metathesis (RCAM) at the C(9)/C(10) site. The requisite diyne was obtained from advanced intermediates that had been prepared as part of the synthesis of the RCM substrates. While the direct conversion of the triple bond formed in the ring-closing step into the C(9)-C(10) double bond of the rhizoxin macrocycle proved to be elusive, the corresponding Z isomer was accessible with high selectivity by reductive decomplexation of the biscobalt hexacarbonyl complex of the triple bond with ethylpiperidinium hypophosphite. Radical-induced double bond isomerization, full elaboration of the C(15) side chain, and directed epoxidation of the C(11)-C(12) double bond completed the total synthesis of rhizoxin F.
Topics: Acids; Alkenes; Alkynes; Cyclization; Macrolides; Models, Molecular
PubMed: 33023218
DOI: 10.3390/molecules25194527 -
The ISME Journal Jun 2008The rice seedling blight fungus Rhizopus microsporus harbors endosymbiotic Burkholderia sp. for the production of the virulence factor, the antimitotic agent rhizoxin....
The rice seedling blight fungus Rhizopus microsporus harbors endosymbiotic Burkholderia sp. for the production of the virulence factor, the antimitotic agent rhizoxin. Since the toxin highly efficiently blocks mitosis in most eukaryotes, it remained elusive how self-resistance emerged in the fungal host. In this study, rhizoxin sensitivity was systematically correlated with the nature of beta-tubulin sequences in the kingdom Fungi. A total of 49 new beta-tubulin sequences were generated for representative species of Ascomycota, Basidiomycota and Zygomycota. Rhizoxin sensitivity assays revealed two further amino acids at position 100 (Ser-100 and Ala-100), in addition to the known Ile-100 and Val-100, which convey rhizoxin resistance. All sensitive strains feature Asn-100. This hot spot was verified by modeling studies, which support the finding that rhizoxin preferentially interacts with the tubulin molecule in a cavity near position 100. Ancestral character state reconstructions conducted in a Bayesian framework suggest that rhizoxin sensitivity represents the ancestral character state in fungi, and that evolution of rhizoxin resistance took place in the ancestor of extant resistant Zygomycota. These findings support a model according to which endosymbiosis became possible through a parasitism--mutualism shift in insensitive fungi.
Topics: Amino Acid Sequence; Bacterial Toxins; Burkholderia; Drug Resistance, Fungal; Evolution, Molecular; Fungal Proteins; Fungi; Macrolides; Models, Molecular; Molecular Sequence Data; Phylogeny; Protein Binding; Rhizopus; Symbiosis; Tubulin
PubMed: 18309361
DOI: 10.1038/ismej.2008.19 -
Applied and Environmental Microbiology May 2008The products synthesized from a hybrid polyketide synthase/nonribosomal peptide synthetase gene cluster in the genome of Pseudomonas fluorescens Pf-5 were identified...
The products synthesized from a hybrid polyketide synthase/nonribosomal peptide synthetase gene cluster in the genome of Pseudomonas fluorescens Pf-5 were identified using a genomics-guided strategy involving insertional mutagenesis and subsequent metabolite profiling. Five analogs of rhizoxin, a 16-member macrolide with antifungal, phytotoxic, and antitumor activities, were produced by Pf-5, but not by a mutant with an insertion in the gene cluster. The five rhizoxin analogs, one of which had not been described previously, were differentially toxic to two agriculturally important plant pathogens, Botrytis cinerea and Phytophthora ramorum. The rhizoxin analogs also caused swelling of rice roots, a symptom characteristic of rhizoxin itself, but were less toxic to pea and cucumber roots. Of the rhizoxin analogs produced by Pf-5, the predominant compound, WF-1360 F, and the newly described compound 22Z-WF-1360 F were most toxic against the two plant pathogens and three plant species. These rhizoxin analogs were tested against a panel of human cancer lines, and they exhibited potent but nonselective cytotoxicity. This study highlights the value of the genomic sequence of the soil bacterium P. fluorescens Pf-5 in providing leads for the discovery of novel metabolites with significant biological properties.
Topics: Antifungal Agents; Antineoplastic Agents; Botrytis; Cell Line, Tumor; Cucumis; Genomics; Humans; Inhibitory Concentration 50; Macrolides; Magnetic Resonance Spectroscopy; Microbial Sensitivity Tests; Molecular Structure; Multigene Family; Mutagenesis, Insertional; Oryza; Pisum sativum; Peptide Synthases; Phytophthora; Plant Roots; Polyketide Synthases; Pseudomonas fluorescens
PubMed: 18344330
DOI: 10.1128/AEM.02848-07