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Proceedings of the National Academy of... Aug 1999The mitomycin C-resistance gene, mcrA, of Streptomyces lavendulae produces MCRA, a protein that protects this microorganism from its own antibiotic, the antitumor drug...
The mitomycin C-resistance gene, mcrA, of Streptomyces lavendulae produces MCRA, a protein that protects this microorganism from its own antibiotic, the antitumor drug mitomycin C. Expression of the bacterial mcrA gene in mammalian Chinese hamster ovary cells causes profound resistance to mitomycin C and to its structurally related analog porfiromycin under aerobic conditions but produces little change in drug sensitivity under hypoxia. The mitomycins are prodrugs that are enzymatically reduced and activated intracellularly, producing cytotoxic semiquinone anion radical and hydroquinone reduction intermediates. In vitro, MCRA protects DNA from cross-linking by the hydroquinone reduction intermediate of these mitomycins by oxidizing the hydroquinone back to the parent molecule; thus, MCRA acts as a hydroquinone oxidase. These findings suggest potential therapeutic applications for MCRA in the treatment of cancer with the mitomycins and imply that intrinsic or selected mitomycin C resistance in mammalian cells may not be due solely to decreased bioactivation, as has been hypothesized previously, but instead could involve an MCRA-like mechanism.
Topics: Aerobiosis; Animals; Bacterial Proteins; Biotransformation; CHO Cells; Cell Hypoxia; Cell Survival; Cloning, Molecular; Cricetinae; Drug Resistance; Mitomycin; Oxidoreductases; Porfiromycin; Prodrugs; Recombinant Proteins; Streptomyces; Transfection
PubMed: 10468636
DOI: 10.1073/pnas.96.18.10489 -
Microbiology Spectrum Feb 2023Streptococcus pyogenes prophage Φ1207.3 (formerly Tn) carries the (A)-(D) resistance genes, responsible for type M macrolide resistance. To investigate if Φ1207.3 is a...
Streptococcus pyogenes prophage Φ1207.3 (formerly Tn) carries the (A)-(D) resistance genes, responsible for type M macrolide resistance. To investigate if Φ1207.3 is a functional bacteriophage, we transferred the element from the original S. pyogenes host in a prophage-free and competence-deficient S. pneumoniae strain. Pneumococcal cultures of the Φ1207.3-carrying lysogen were treated with mitomycin C to assess if Φ1207.3 enters the lytic cycle. Mitomycin C induced a limited phage burst and a growth impairment, resulting in early entrance into the stationary phase. To determine if Φ1207.3 is able to produce mature phage particles, we prepared concentrated supernatants recovered from a mitomycin C-induced pneumococcal culture by sequential centrifugation and ultracentrifugation steps. Negative-staining transmission electron microscopy (TEM) of supernatants revealed the presence of phage particles with an icosahedral, electron-dense capsid and a long, noncontractile tail, typical of a siphovirus. Quantification of Φ1207.3 was performed by quantitative PCR (qPCR) and semiquantitatively by TEM. PCR quantified 3.34 × 10 and 6.06 × 10 excised forms of phage genome per milliliter of supernatant obtained from the untreated and mitomycin C-treated cultures, respectively. By TEM, we estimated 3.02 × 10 and 7.68 × 10 phage particles per milliliter of supernatant. The phage preparations of Φ1207.3 infected and lysogenized pneumococcal recipient strains at a frequency of 7.5 × 10 lysogens/recipient but did not show sufficient lytic activity to form plaques. Phage lysogenization efficiently occurred after 30 min of contact of the phages with the recipient cells and required a minimum of 10 phage particles. Bacteriophages play an important role in bacterial physiology and genome evolution. The widespread use of genome sequencing revealed that bacterial genomes can contain several different integrated temperate bacteriophages, which can constitute up to 20% of the genome. Most of these bacteriophages are only predicted and are never shown to be functional. In fact, it is often difficult to induce the lytic cycle of temperate bacteriophages. In this work, we show that Φ1207.3, a peculiar bacteriophage originally from Streptococcus pyogenes, which can lysogenize different streptococci and carries the macrolide resistance (A)-(D) gene pair, is capable of producing mature virions, but only at a low level, while not being able to produce plaques. This temperate phage is probably a partially functional phage, which seems to have lost lytic characteristics to specialize in lysogenization. While we are not used to conceiving phages separately from lysis, this behavior could actually be more frequent than expected.
Topics: Bacteriophages; Anti-Bacterial Agents; Streptococcus pyogenes; Macrolides; Mitomycin; Drug Resistance, Bacterial; Prophages
PubMed: 36625667
DOI: 10.1128/spectrum.04211-22 -
Experimental Eye Research Feb 2023In this paper, we use RNAseq to identify senescence and phagocytosis as key factors to understanding how mitomyin C (MMC) stimulates regenerative wound repair. We use...
Molecular mechanisms regulating wound repair: Evidence for paracrine signaling from corneal epithelial cells to fibroblasts and immune cells following transient epithelial cell treatment with Mitomycin C.
In this paper, we use RNAseq to identify senescence and phagocytosis as key factors to understanding how mitomyin C (MMC) stimulates regenerative wound repair. We use conditioned media (CM) from untreated (CMC) and MMC treated (CMM) human and mouse corneal epithelial cells to show that corneal epithelial cells indirectly exposed to MMC secrete elevated levels of immunomodulatory proteins including IL-1α and TGFβ1 compared to cells exposed to CMC. These factors increase epithelial and macrophage phagocytosis and promote ECM turnover. IL-1α supplementation can increase phagocytosis in control epithelial cells and attenuate TGFβ1 induced αSMA expression by corneal fibroblasts. Yet, we show that epithelial cell CM contains factors besides IL-1α that regulate phagocytosis and αSMA expression by fibroblasts. Exposure to CMM also impacts the activation of bone marrow derived dendritic cells and their ability to present antigen. These in vitro studies show how a brief exposure to MMC induces corneal epithelial cells to release proteins and other factors that function in a paracrine way to enhance debris removal and enlist resident epithelial and immune cells as well as stromal fibroblasts to support regenerative and not fibrotic wound healing.
Topics: Humans; Animals; Mice; Mitomycin; Paracrine Communication; Cells, Cultured; Fibroblasts; Wound Healing; Epithelial Cells
PubMed: 36539051
DOI: 10.1016/j.exer.2022.109353 -
Molekuliarnaia Biologiia 2022DNA-methyltransferases catalyze DNA methylation in the CpG sites, which play an important role in the maintenance of genome stability. The association between DNA...
DNA-methyltransferases catalyze DNA methylation in the CpG sites, which play an important role in the maintenance of genome stability. The association between DNA methylation and genotoxic stress resulting in the action of various clastogens has been shown. Genotoxic stress is one of the triggers of endothelial dysfunction. In this study, the transcription of DNMT1, DNMT3A and DNMT3B genes in coronary (HCAEC) and internal thoracic (HITAEC) artery endothelial cells exposed to alkylating mutagen mitomycin C was studied using quantitative polymerase chain reaction. In HCAEC exposed to mitomycin C, DNMT1 transcription is 1.7-fold higher compared to the unexposed control. After elimination of the mutagen from the cultures followed by 24-hours of cultivation, a 2-fold increase of transcription of DNMT3B in HCAEC exposed to mitomycin C compared to the control was observed. At the same time, no changes in transcription of the studied DNA-methyltransferases were found in HITAEC exposed to the mutagen. Thus, increased transcription of DNA-methyltransferase may be a possible molecular mechanism underlying endothelial dysfunction in response to mutagenic load in an in vitro experiment.
Topics: DNA; DNA Methylation; DNA Methyltransferase 3A; Endothelial Cells; Mitomycin; Mutagens
PubMed: 35621104
DOI: 10.31857/S0026898422030156 -
Scientific Reports Dec 2021Optical coherence tomography angiography (OCTA) is a new technique for non-invasive imaging of blood vessels, allowing combined evaluation of both deep and surface...
Optical coherence tomography angiography (OCTA) is a new technique for non-invasive imaging of blood vessels, allowing combined evaluation of both deep and surface vessels. The purpose of this study was to evaluate the post-trabeculectomy longitudinal changes in complete avascular area (CAA) of filtering blebs using anterior segment (AS-) OCTA and their association with surgical outcomes. This study included 57 eyes of 53 patients who had undergone trabeculectomy with mitomycin C. AS-OCTA images of filtering bleb were acquired at 3 and 6 months after trabeculectomy, and at 1 month in possible cases. CAAs, regions where complete blood flow was not depicted in AS-OCTA images, were evaluated for their presence, extent, and change over time. CAAs were detected in 37 eyes (65%) and 33 eyes (58%) at 3 and 6 months postoperatively, respectively. The extent of CAAs reduced over time after surgery in most cases. No parameters related to CAAs were significantly associated with surgical success (i.e., intraocular pressure (IOP) ≤ 12 mmHg and IOP reduction > 20% without medication). In conclusion, although it is difficult to predict surgical success by CAA itself, AS-OCTA may be useful for the objective evaluation of the vascularity of filtering blebs.
Topics: Adult; Aged; Computed Tomography Angiography; Female; Glaucoma; Humans; Intraocular Pressure; Longitudinal Studies; Male; Middle Aged; Mitomycin; Prospective Studies; Tomography, Optical Coherence; Trabeculectomy; Treatment Outcome
PubMed: 34862440
DOI: 10.1038/s41598-021-02871-2 -
The Cochrane Database of Systematic... Jan 2020People with urothelial carcinoma of the bladder are at risk for recurrence and progression following transurethral resection of a bladder tumour (TURBT). Mitomycin C... (Meta-Analysis)
Meta-Analysis
BACKGROUND
People with urothelial carcinoma of the bladder are at risk for recurrence and progression following transurethral resection of a bladder tumour (TURBT). Mitomycin C (MMC) and Bacillus Calmette-Guérin (BCG) are commonly used, competing forms of intravesical therapy for intermediate- or high-risk non-muscle invasive (Ta and T1) urothelial bladder cancer but their relative merits are somewhat uncertain.
OBJECTIVES
To assess the effects of BCG intravesical therapy compared to MMC intravesical therapy for treating intermediate- and high-risk Ta and T1 bladder cancer in adults.
SEARCH METHODS
We performed a systematic literature search in multiple databases (CENTRAL, MEDLINE, Embase, Web of Science, Scopus, LILACS), as well as in two clinical trial registries. We searched reference lists of relevant publications and abstract proceedings. We applied no language restrictions. The latest search was conducted in September 2019.
SELECTION CRITERIA
We included randomised controlled trials (RCTs) that compared intravesical BCG with intravesical MMC therapy for non-muscle invasive urothelial bladder cancer.
DATA COLLECTION AND ANALYSIS
Two review authors independently screened the literature, extracted data, assessed risk of bias and rated the quality of evidence according to GRADE per outcome. In the meta-analyses, we used the random-effects model.
MAIN RESULTS
We identified 12 RCTs comparing BCG versus MMC in participants with intermediate- and high-risk non-muscle invasive bladder tumours (published from 1995 to 2013). In total, 2932 participants were randomised. Time to death from any cause: BCG may make little or no difference on time to death from any cause compared to MMC (hazard ratio (HR) 0.97, 95% confidence interval (CI) 0.79 to 1.20; participants = 1132, studies = 5; 567 participants in the BCG arm and 565 in the MMC arm; low-certainty evidence). This corresponds to 6 fewer deaths (40 fewer to 36 more) per 1000 participants treated with BCG at five years. We downgraded the certainty of the evidence two levels due to study limitations and imprecision. Serious adverse effects: 12/577 participants treated with BCG experienced serious non-fatal adverse effects compared to 4/447 participants in the MMC group. The pooled risk ratio (RR) is 2.31 (95% CI 0.82 to 6.52; participants = 1024, studies = 5; low-certainty evidence). Therefore, BCG may increase the risk for serious adverse effects compared to MMC. This corresponds to nine more serious adverse effects (one fewer to 37 more) with BCG. We downgraded the certainty of the evidence two levels due to study limitations and imprecision. Time to recurrence: BCG may reduce the time to recurrence compared to MMC (HR 0.88, 95% CI 0.71 to 1.09; participants = 2616, studies = 11, 1273 participants in the BCG arm and 1343 in the MMC arm; low-certainty evidence). This corresponds to 41 fewer recurrences (104 fewer to 29 more) with BCG at five years. We downgraded the certainty of the evidence two levels due to study limitations, imprecision and inconsistency. Time to progression: BCG may make little or no difference on time to progression compared to MMC (HR 0.96, 95% CI 0.73 to 1.26; participants = 1622, studies = 6; 804 participants in the BCG arm and 818 in the MMC arm; low-certainty evidence). This corresponds to four fewer progressions (29 fewer to 27 more) with BCG at five years. We downgraded the certainty of the evidence two levels due to study limitations and imprecision. Quality of life: we found very limited data for this outcomes and were unable to estimate an effect size.
AUTHORS' CONCLUSIONS
Based on our findings, BCG may reduce the risk of recurrence over time although the Confidence Intervals include the possibility of no difference. It may have no effect on either the risk of progression or risk of death from any cause over time. BCG may cause more serious adverse events although the Confidence Intervals once again include the possibility of no difference. We were unable to determine the impact on quality of life. The certainty of the evidence was consistently low, due to concerns that include possible selection bias, performance bias, given the lack of blinding in these studies, and imprecision.
Topics: Administration, Intravesical; Antibiotics, Antineoplastic; BCG Vaccine; Carcinoma, Transitional Cell; Humans; Mitomycin; Randomized Controlled Trials as Topic; Treatment Outcome; Urinary Bladder Neoplasms
PubMed: 31912907
DOI: 10.1002/14651858.CD011935.pub2 -
Molecules (Basel, Switzerland) Feb 2020Traditional medicinal plants are an important source of active compounds with potential antimutagenic activity. Bailey (Araliaceae) is a South Asian traditional herb...
Traditional medicinal plants are an important source of active compounds with potential antimutagenic activity. Bailey (Araliaceae) is a South Asian traditional herb used as an adaptogenic and cardiac drug. Extracts of contain a wide range of biologically active compounds like phenolic acids and triterpenoid saponins. In the present study. antigenotoxic potential of three naturally occurring phenolic acids and extracts of growing in vitro with the addition of elicitors was evaluated against direct (4-nitroquinoline-N-oxide (4NQO) and mitomycin C (MMC)) and indirect mutagens (2-aminoanthracene (2AA)). The evaluation was made using a bacterial -test. Moreover, the ability to prevent photogenotoxicity induced by chlorpromazine (CPZ) under UVA irradiation was measured. The phytochemical profiling of examined extracts revealed the presence of numerous compounds with the prevelance of chlorogenic, caffeic, and ferulic acid derivatives; however, saponin fractions were also determined. The antioxidant potential of extracts strictly correlated with their composition. The tested extracts exhibited high antigenotoxic activity if the assay was performed with 2AA and metabolic activation. Moreover, the extracts slightly decreased the MMC-induced genotoxicity. However, an increase of the genotoxic effect was observed in the assay performed with 4NQO. In addition, photo-antigenotoxic activity was observed. In our study, phenolic acids exhibited lower activity than the extracts.
Topics: Animals; Antimutagenic Agents; Antioxidants; Araliaceae; Chlorpromazine; DNA Damage; Male; Mitomycin; Plant Extracts; Plant Shoots; Rats; Rats, Sprague-Dawley
PubMed: 32121158
DOI: 10.3390/molecules25051090 -
Chemistry (Weinheim An Der Bergstrasse,... Sep 2018Mitomycin C (MC), an antitumor drug, and decarbamoylmitomycin C (DMC), a derivative of MC, alkylate DNA and form deoxyguanosine monoadducts and interstrand crosslinks...
Mitomycin C (MC), an antitumor drug, and decarbamoylmitomycin C (DMC), a derivative of MC, alkylate DNA and form deoxyguanosine monoadducts and interstrand crosslinks (ICLs). Interestingly, in mammalian culture cells, MC forms primarily deoxyguanosine adducts with a 1"-R stereochemistry at the guanine-mitosene bond (1"-α) whereas DMC forms mainly adducts with a 1"-S stereochemistry (1"-β). The molecular basis for the stereochemical configuration exhibited by DMC has been investigated using biomimetic synthesis. Here, we present the results of our studies on the monoalkylation of DNA by DMC. We show that the formation of 1"-β-deoxyguanosine adducts requires bifunctional reductive activation of DMC, and that monofunctional activation only produces 1"-α-adducts. The stereochemistry of the deoxyguanosine adducts formed is also dependent on the regioselectivity of DNA alkylation and on the overall DNA CG content. Additionally, we found that temperature plays a determinant role in the regioselectivity of duplex DNA alkylation by mitomycins: At 0 °C, both deoxyadenosine (dA) and deoxyguanosine (dG) alkylation occur whereas at 37 °C, mitomycins alkylate dG preferentially. The new reaction protocols developed in our laboratory to investigate DMC-DNA alkylation raise the possibility that oligonucleotides containing DMC 1"-β-deoxyguanosine adducts at a specific site may be synthesized by a biomimetic approach.
Topics: Alkylation; Animals; Base Sequence; Chromatography, High Pressure Liquid; DNA; DNA Adducts; DNA, Bacterial; Deoxyadenosines; Deoxyguanosine; Mice; Micrococcus luteus; Mitomycin; Mitomycins; Stereoisomerism; Temperature
PubMed: 29958326
DOI: 10.1002/chem.201802038 -
Cancer Genomics & Proteomics 2007The modern era of targeted therapeutics offers the potential to tailor therapy to individual patients whose tumours express a specific target. Previous attempts to... (Review)
Review
The modern era of targeted therapeutics offers the potential to tailor therapy to individual patients whose tumours express a specific target. Previous attempts to forecast tumour response to conventional chemotherapeutics based on similar principles have however been disappointing. Mitomycin C (MMC), for example, is a bioreductive drug that requires metabolic activation by cellular reductases for activity. The enzyme NAD(P)H:Quinone oxidoreductase-1 (NQO1) can reduce MMC to DNA damaging species but attempts to establish the relationship between tumour response to MMC and NQO1 expression have generated conflicting reports of good and poor correlations. Several other reductases are known to activate MMC. This, in conjunction with the fact that various physiological and biochemical factors influence therapeutic response, suggests that the mechanism of action of MMC is too complex to allow tumour response to be predicted on the basis of a single enzyme. Alternative approaches using more complex biological and pharmacological systems that reflect the spectrum of reductases present within the tumour have been developed and it remains to be seen whether or not the predictive value of these approaches is enhanced. With regards to targeted therapeutics, the experience with MMC suggests that prediction of tumour response based on analysis of a single target may be too simplistic. Multiple mechanisms of action and the influence of tumour microenvironment on cell biology and drug delivery are likely to influence the final outcome of therapy. The challenge for the future progression of this field is to develop assays that reflect the overall biological and pharmacological processes involved in drug activation whilst retaining the simplicity and robustness required for routine chemosensitivity testing in a clinical setting.
Topics: Antibiotics, Antineoplastic; DNA Damage; Drug Screening Assays, Antitumor; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Humans; Mitomycin; NAD(P)H Dehydrogenase (Quinone); Neoplasms; Treatment Outcome
PubMed: 17878521
DOI: No ID Found -
Molecules (Basel, Switzerland) Nov 2021Mitomycin has a unique chemical structure and contains densely assembled functionalities with extraordinary antitumor activity. The previously proposed mitomycin C...
Mitomycin has a unique chemical structure and contains densely assembled functionalities with extraordinary antitumor activity. The previously proposed mitomycin C biosynthetic pathway has caused great attention to decipher the enzymatic mechanisms for assembling the pharmaceutically unprecedented chemical scaffold. Herein, we focused on the determination of acyl carrier protein (ACP)-dependent modification steps and identification of the protein-protein interactions between MmcB (ACP) with the partners in the early-stage biosynthesis of mitomycin C. Based on the initial genetic manipulation consisting of gene disruption and complementation experiments, genes , , and were identified as the essential functional genes in the mitomycin C biosynthesis, respectively. Further integration of biochemical analysis elucidated that MitE catalyzed CoA ligation of 3-amino-5-hydroxy-bezonic acid (AHBA), MmcB-tethered AHBA triggered the biosynthesis of mitomycin C, and both MitB and MitF were MmcB-dependent tailoring enzymes involved in the assembly of mitosane. Aiming at understanding the poorly characterized protein-protein interactions, the pull-down assay was carried out by monitoring MmcB individually with MitB and MitF. The observed results displayed the clear interactions between MmcB and MitB and MitF. The surface plasmon resonance (SPR) biosensor analysis further confirmed the protein-protein interactions of MmcB with MitB and MitF, respectively. Taken together, the current genetic and biochemical analysis will facilitate the investigations of the unusual enzymatic mechanisms for the structurally unique compound assembly and inspire attempts to modify the chemical scaffold of mitomycin family antibiotics.
Topics: Acyl Carrier Protein; Amino Acid Sequence; Aminobenzoates; Anti-Bacterial Agents; China; Escherichia coli; Escherichia coli Proteins; Hydroxybenzoates; Mitomycin; Mitomycins; Protein Interaction Mapping; Protein Interaction Maps; Streptomyces
PubMed: 34833880
DOI: 10.3390/molecules26226791