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Ogawara H. Comparison of Antibiotic Resistance Mechanisms in Antibiotic-Producing and Pathogenic Bacteria. Molecules 2019; 24:E3430. [PMID: 31546630 PMCID: PMC6804068 DOI: 10.3390/molecules24193430] [Citation(s) in RCA: 50] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2019] [Revised: 09/18/2019] [Accepted: 09/20/2019] [Indexed: 12/13/2022] Open
Abstract
Antibiotic resistance poses a tremendous threat to human health. To overcome this problem, it is essential to know the mechanism of antibiotic resistance in antibiotic-producing and pathogenic bacteria. This paper deals with this problem from four points of view. First, the antibiotic resistance genes in producers are discussed related to their biosynthesis. Most resistance genes are present within the biosynthetic gene clusters, but some genes such as paromomycin acetyltransferases are located far outside the gene cluster. Second, when the antibiotic resistance genes in pathogens are compared with those in the producers, resistance mechanisms have dependency on antibiotic classes, and, in addition, new types of resistance mechanisms such as Eis aminoglycoside acetyltransferase and self-sacrifice proteins in enediyne antibiotics emerge in pathogens. Third, the relationships of the resistance genes between producers and pathogens are reevaluated at their amino acid sequence as well as nucleotide sequence levels. Pathogenic bacteria possess other resistance mechanisms than those in antibiotic producers. In addition, resistance mechanisms are little different between early stage of antibiotic use and the present time, e.g., β-lactam resistance in Staphylococcus aureus. Lastly, guanine + cytosine (GC) barrier in gene transfer to pathogenic bacteria is considered. Now, the resistance genes constitute resistome composed of complicated mixture from divergent environments.
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Affiliation(s)
- Hiroshi Ogawara
- HO Bio Institute, 33-9, Yushima-2, Bunkyo-ku, Tokyo 113-0034, Japan.
- Department of Biochemistry, Meiji Pharmaceutical University, 522-1, Noshio-2, Kiyose, Tokyo 204-8588, Japan.
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Tenconi E, Rigali S. Self-resistance mechanisms to DNA-damaging antitumor antibiotics in actinobacteria. Curr Opin Microbiol 2018; 45:100-108. [PMID: 29642052 DOI: 10.1016/j.mib.2018.03.003] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2018] [Revised: 03/08/2018] [Accepted: 03/23/2018] [Indexed: 10/17/2022]
Abstract
Streptomyces and few other Actinobacteria naturally produce compounds currently used in chemotherapy for being cytotoxic against various types of tumor cells by damaging the DNA structure and/or inhibiting DNA functions. DNA-damaging antitumor antibiotics belong to different classes of natural compounds that are structurally unrelated such as anthracyclines, bleomycins, enediynes, mitomycins, and prodiginines. By targeting a ubiquitous molecule and housekeeping functions, these compounds are also cytotoxic to their producer. How DNA-damaging antitumor antibiotics producing actinobacteria avoid suicide is the theme of the current review which illustrates the different strategies developed for self-resistance such as toxin sequestration, efflux, modification, destruction, target repair/protection, or stochastic activity. Finally, the observed spatio-temporal correlation between cell death, morphogenesis, and prodiginine production in S. coelicolor suggests a new physiological role for these molecules, that, together with their self-resistance mechanisms, would function as new types of toxin-antitoxin systems recruited in programmed cell death processes of the producer.
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Affiliation(s)
- Elodie Tenconi
- InBioS - Center for Protein Engineering, Université de liège, Institut de Chimie B64, B-4000 Liège, Belgium
| | - Sébastien Rigali
- InBioS - Center for Protein Engineering, Université de liège, Institut de Chimie B64, B-4000 Liège, Belgium.
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3
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Yang JL, Qin Y, Li L, Cao CY, Wang Q, Li Q, Lv YF, Wang Y. Apoptotic Melanoma B16-F1 Cells Induced by Lidamycin Could Initiate the Antitumor Immune Response in BABL/c Mice. Oncol Res 2016; 23:79-86. [PMID: 26802654 PMCID: PMC7842507 DOI: 10.3727/096504015x14478843952942] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022] Open
Abstract
In the process of tumor cell apoptosis induced by specific regents, calreticulin (CRT) was transferred from endoplasmic reticulum (ER) onto the cell membrane. These tumor cells, when used as the cellular vaccine to immunize experimental animals, could initiate effective antitumor immunoresponse against homologous tumor cells. This is referred to as immunogenic cell death. Lidamycin (LDM) is an enediyne antibiotic, which has extremely potent cytotoxicity to cancer cells. In this study, the mouse melanoma B16-F1 cancer cells were used to investigate the ability of LDM in promoting immunogenic cell death. Our data showed that LDM could induce apoptosis of B16-F1 cancer cells, accompanied by CRT translocation onto the cell membrane. These LDM-treated B16-F1 cells could be recognized and phagocytosed more efficiently by macrophage and dendritic cells. When the LDM-treated apoptotic B16-F1 cells were used as a whole-cell tumor vaccine to immune mice, the mice obtained resistance against rechallenged B16-F1 living cells. At the same time, the specific antitumor immune response was observed in these vaccinated mice. The splenocytes from the mice vaccinated with LDM-treated B16-F1 cells showed significantly enhanced NK lymphocyte activities and also faster growth rate and increased secretion of IFN-γ when encountering the cellular antigens from B16-F1 cells. All these results suggested that LDM could promote immunogenic cell death in B16-F1 cells, and these LDM-treated B16-F1 cells could be used as a sort of cell vaccine to initiate effective antitumor immunoresponse in mice.
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Affiliation(s)
- Jian-lin Yang
- China Three Gorges University Medical College, Hubei Key Laboratory of Tumor Microenvironment and Immunotherapy, Yichang, Hubei, China
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Mohamed RK, Peterson PW, Alabugin IV. Concerted Reactions That Produce Diradicals and Zwitterions: Electronic, Steric, Conformational, and Kinetic Control of Cycloaromatization Processes. Chem Rev 2013; 113:7089-129. [DOI: 10.1021/cr4000682] [Citation(s) in RCA: 166] [Impact Index Per Article: 13.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022]
Affiliation(s)
- Rana K. Mohamed
- Department of Chemistry
and Biochemistry, Florida State University, Tallahassee, Florida 32306-4390,
United States
| | - Paul W. Peterson
- Department of Chemistry
and Biochemistry, Florida State University, Tallahassee, Florida 32306-4390,
United States
| | - Igor V. Alabugin
- Department of Chemistry
and Biochemistry, Florida State University, Tallahassee, Florida 32306-4390,
United States
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Chi HW, Huang CC, Chin DH. Thiols Screened by the Neocarzinostatin Protein for Preserving or Detoxifying its Bound Enediyne Antibiotic. Chemistry 2012; 18:6238-49. [DOI: 10.1002/chem.201102825] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/09/2011] [Revised: 01/12/2012] [Indexed: 12/28/2022]
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Wen Y, Chen S, Meng Z, Gan H, Zhu X, Wu Z, Gu R, Dou G. Simultaneous determination of lidamycin enediyne chromophore (LDC) and its aromatized derivative (LDCA) using puerarin as internal standard in rat plasma by LC-MS/MS. Biomed Chromatogr 2011; 26:400-6. [PMID: 21830226 DOI: 10.1002/bmc.1676] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2011] [Accepted: 06/22/2011] [Indexed: 11/11/2022]
Abstract
Lidamycin (LDM), a promising enediyne antitumor antibiotic, was quantified by detecting lidamycin enediyne chromophore (LDC) using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the first time. A simple, rapid and reliable method was developed and validated to determine LDC and its aromatized derivative (LDCA) simultaneously in plasma. Puerarin was used as an internal standard (IS), and plasma samples were pretreated with one-step precipitation by acetonitrile. Separation was achieved on a reverse-phase C(18) column with a mobile phase composed of methanol and water containing 5 mm ammonium acetate at pH 3.5 in gradient elution mode. Detection was performed on a triple quadrupole tandem mass spectrometer using electrospray ionization (ESI) by multiple reaction monitoring (MRM) in the negative ion mode. Good linearity was obtained over the concentration range of 0.2-100 µg/mL for LDM. Precision and accuracy were validated by RSD% values in the range of 2.6-13.0% and RE% values between -4.6 and 3.8%, respectively. In addition, no specificity and matrix effects were observed. The recovery was found to be 99.2-111.0% and stability in various conditions was found to be acceptable. This method was applied in preclinical pharmacokinetic studies for routine monitoring of LDM in rat plasma.
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Affiliation(s)
- Yanqing Wen
- Laboratory of Drug Metabolism and Pharmacokinetics, Beijing Institute of Blood Transfusion Medical Science, 27 Taiping Road, Beijing, 100850, China
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Zhu Y, Wang L, Du Y, Wang S, Yu T, Hong B. Heterologous expression of human interleukin-6 in Streptomyces lividans TK24 using novel secretory expression vectors. Biotechnol Lett 2010; 33:253-61. [DOI: 10.1007/s10529-010-0428-0] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/08/2010] [Accepted: 09/23/2010] [Indexed: 11/24/2022]
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Baroudi A, Mauldin J, Alabugin IV. Conformationally Gated Fragmentations and Rearrangements Promoted by Interception of the Bergman Cyclization through Intramolecular H-Abstraction: A Possible Mechanism of Auto-Resistance to Natural Enediyne Antibiotics? J Am Chem Soc 2009; 132:967-79. [DOI: 10.1021/ja905100u] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/24/2023]
Affiliation(s)
- Abdulkader Baroudi
- Department of Chemistry and Biochemistry, Florida State University, Tallahassee, Florida 32306
| | - Justin Mauldin
- Department of Chemistry and Biochemistry, Florida State University, Tallahassee, Florida 32306
| | - Igor V. Alabugin
- Department of Chemistry and Biochemistry, Florida State University, Tallahassee, Florida 32306
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Cui Z, Wang L, Wang S, Li G, Hong B. Disruption of cagA, the apoprotein gene of chromoprotein antibiotic C-1027, eliminates holo-antibiotic production, but not the cytotoxic chromophore. FEMS Microbiol Lett 2009; 301:57-68. [PMID: 19845765 DOI: 10.1111/j.1574-6968.2009.01800.x] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022] Open
Abstract
C-1027 is a chromoprotein of the nine-membered enediyne antitumour antibiotic family, comprising apoprotein to stabilize and transport the enediyne chromophore. The disruption of apoprotein gene cagA within the C-1027 biosynthetic gene cluster abolished C-1027 holo-antibiotic production detected by an antibacterial assay, as well as the expression of the apoprotein and C-1027 chromophore extracted following protein precipitation of the culture supernatant. Complementation of the cagA-disrupted mutant AKO with the intact cagA gene restored C-1027 production, suggesting that cagA is indispensable for holo-antibiotic production. Overexpression of cagA in the wild-type strain resulted in a significant increase in C-1027 production as expected. Surprisingly, electrospray ionization (ESI)-MS and ESI-MS/MS analyses suggested that the AKO mutant still produced the C-1027 enediyne chromophore [m/z=844 (M+H)(+)] and its aromatized product [m/z=846 (M+H)(+)]. Consistent with this, the results from gene expression analysis using real-time reverse transcriptase-PCR showed that transcripts of the positive regulator sgcR3 and the structural genes sgcA1, sgcC4, sgcD6 and sgcE were readily detected in the AKO mutant as well as in the wild-type and the complementation strain. These results provided, for the first time, evidence suggesting that the apoprotein of C-1027 is not essential in the self-resistance mechanism for the enediyne chromophore.
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Affiliation(s)
- Zhihui Cui
- Key Laboratory of Biotechnology of Antibiotics of Ministry of Health, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
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Cai L, Chen H, Miao Q, Wu S, Shang Y, Zhen Y. Binding capability of the enediyne-associated apoprotein to human tumors and constitution of a ligand oligopeptide-integrated protein. J Biotechnol 2009; 144:142-50. [PMID: 19737585 DOI: 10.1016/j.jbiotec.2009.09.001] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2009] [Revised: 08/16/2009] [Accepted: 09/01/2009] [Indexed: 10/20/2022]
Abstract
The molecule of lidamycin that belongs to the chromoprotein family of antitumor antibiotics is composed of an apoprotein (LDP) and an enediyne chromophore. The enediyne moiety of the molecule is responsible for the potent cytotoxicity; however, the biological function of the apoprotein moiety, particularly its interaction with cancer cells, remains unclear. In present study, the binding capability of LDP to human tumors was detected for the first time by tissue microarray. LDP bound to various human tumors with significant difference from the corresponding normal tissues. Positive correlation between binding activity and the overexpression of VEGF and EGFR was confirmed by lung carcinoma tissue microarray. A fusion protein LG-LDP that consists of LDP and a ligand oligopeptide to EGFR was constructed by DNA recombination. LG-LDP showed augmented binding to EGFR-overexpressing cancer cells. Furthermore, an energized fusion protein LG-LDP-AE was prepared by integrating the active enediyne (AE) into LG-LDP molecule. By MTT assay, LG-LDP-AE displayed extremely potent cytotoxicity to cancer cells with IC50 approximate to 0.01nM. The results indicate that LDP binds to various human tumors and it might serve as a delivery carrier by integration of ligand oligopeptide to manufacture motif-based, targeted fusion proteins for cancer.
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Affiliation(s)
- Lin Cai
- Department of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
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Chen J, Wu SY, Ou-Yang ZG, Zhen YS. Synergy of gemcitabine and lidamycin associated with NF-kappaB downregulation in pancreatic carcinoma cells. Acta Pharmacol Sin 2008; 29:614-9. [PMID: 18430374 DOI: 10.1111/j.1745-7254.2008.00774.x] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
AIM To investigate the effects on human pancreatic cancer PANC-1 and SW1990 cells using a combination of lidamycin (LDM) and gemcitabine. METHODS A 3- (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was used to determine the growth inhibition of drugs in PANC-1 and SW1990 cells. The effects on apoptosis were measured by terminal uridine deoxynucleotidyl transferase dUTP nick end labeling assay and flow cytometry combined with fluorescein- isothiocyanate-Annexin V/propidium iodide staining. The activity of caspase-3 was measured with a special assay kit. The mitochondrial membrane potential was determined by confocal microscopy analyses. The level of mRNA encoding K-ras in the cells was determined by RT-PCR analysis. The expression of K-ras, NF-kappaB, and Bcl-2 was detected by Western blotting analysis. RESULTS There was a significant reduction in proliferation in the pancreatic cancer cell lines treated with a combination of gemcitabine and LDM. The overall growth inhibition directly correlated with apoptotic cell death. LDM potentiated the gemcitabine-induced cell killing by reducing mitochondrial membrane potential and increasing the caspase-3 activity. Notably, the K-ras mRNA level was significantly reduced with the combination of gemcitabine and LDM. The results for K-ras, NF-kappaB, and Bcl-2 proteins also showed downregulation in the combination group relative to the single-agent treatment and the untreated control. CONCLUSION LDM can potentiate the growth inhibition induced by gemcitabine in human pancreatic cancer cells, and the synergy may be associated with NF-kappaB downregulation.
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Affiliation(s)
- Jing Chen
- Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
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12
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Abstract
The enediyne polyketides are secondary metabolites isolated from a variety of Actinomycetes. All members share very potent anticancer and antibiotic activity, and prospects for the clinical application of the enediynes has been validated with the recent marketing of two enediyne derivatives as anticancer agents. The biosynthesis of these compounds is of interest because of the numerous structural features that are unique to the enediyne family. The gene cluster for five enediynes has now been cloned and sequenced, providing the foundation to understand natures' means to biosynthesize such complex, exotic molecules. Presented here is a review of the current progress in delineating the biosynthesis of the enediynes with an emphasis on the model enediyne, C-1027.
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Affiliation(s)
- Steven G Van Lanen
- Division of Pharmaceutical Sciences, University of Wisconsin, Madison, WI, 53705.
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Miao QF, Liu XY, Shang BY, Ouyang ZG, Zhen YS. An enediyne-energized single-domain antibody-containing fusion protein shows potent antitumor activity. Anticancer Drugs 2007; 18:127-37. [PMID: 17159599 DOI: 10.1097/cad.0b013e3280112779] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
Abstract
Single-domain antibodies are attractive as tumor-targeting vehicles because of their much smaller size than intact antibody molecules. Lidamycin is a macromolecular antitumor antibiotic, which consists of a labile enediyne chromophore (AE) and a noncovalently bound apoprotein (LDP). An enediyne-energized fusion protein VH-LDP-AE composed of single-domain antibody directed against type IV collagenase and lidamycin was prepared by a novel two-step method including DNA recombination and molecular reconstitution. VH-LDP-AE demonstrated extremely potent cytotoxicity to cancer cells and marked antiangiogenic activity in vitro. In the mouse hepatoma 22 model, drugs were administered intravenously as a single dose on day 1 with maximal tolerated doses. VH-LDP-AE (0.25 mg/kg) suppressed the tumor growth by 95.9%, whereas lidamycin (0.05 mg/kg) and mitomycin (1 mg/kg) by 79.6 and 51.1%, respectively. In the HT-1080 xenograft model in nude mice, drugs were given intravenously as a single dose on day 4 after tumor implantation. VH-LDP-AE at 0.25 mg/kg suppressed tumor growth by 76% (P<0.05) compared with that of lidamycin at 0.05 mg/kg (53%) on day 18. No obvious toxic effects were observed in all groups during treatments. The results showed that energized fusion protein VH-LDP-AE was more effective than lidamycin and mitomycin. These properties, together with its much smaller size than conventional antibody-based agents, suggested that VH-LDP-AE would be a promising candidate for cancer-targeting therapy. In addition, the two-step approach could serve as a new technology platform for making a series of highly potent engineered antibody-based drugs for a variety of cancers.
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Affiliation(s)
- Qing-fang Miao
- Department of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences & Peking Union Medical College, #1 Tiantan Xili, Beijing 100050, PRC
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Inoue M, Usuki T, Lee N, Hirama M, Tanaka T, Hosoi F, Ohie S, Otani T. Antitumor Enediyne Chromoprotein C-1027: Mechanistic Investigation of the Chromophore-Mediated Self-Decomposition Pathway. J Am Chem Soc 2006; 128:7896-903. [PMID: 16771503 DOI: 10.1021/ja060724w] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
C-1027 is an extremely potent antitumor agent that causes double-stranded DNA cleavages. It is a unique small molecule-protein complex composed of a highly reactive enediyne chromophore, which upon binding reacts with its target molecule DNA through radical-mediated hydrogen abstraction and an apoprotein that encapsulates the chromophore serving as its carrier to reach DNA. Although C-1027 has favorable properties as an effective drug delivery system, it slowly self-decomposes due to the reactivity of the chromophore toward the apoprotein. Understanding how the C-1027 destroys itself may enable design of its analogues that overcome this limitation. In this paper, mechanistic insights into the self-reactivity of C-1027 that facilitates its own decomposition are described. We provide evidence that the formation of the Gly96 radical, which promotes the oxidative protein scission and the subsequent chromophore release, is the major pathway for the self-decomposition of C-1027. On the basis of the newly isolated products of the self-decomposition, we propose that the apoprotein effectively protects two different structural elements of the chromophore that are essential for its biological activity: the nine-membered enediyne moiety (necessary for DNA cleavage) and the benzoxazine moiety (necessary for DNA intercalation). Using an engineered apoprotein analogue kinetically more stable toward the chromophore radical, we show that enhanced overall properties can be achieved for the natural C-1027 with respect to stability and antitumor activities. The results present the first example of a rationally designed C-1027 analogue reported to display superior in vitro antitumor activity to the natural C-1027. Our findings may have implications for design of proteins that can stably encapsulate highly reactive small molecules.
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Affiliation(s)
- Masayuki Inoue
- Department of Chemistry, Graduate School of Science, Tohoku University, Sendai 980-8578, Japan.
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Inoue M. Exploring the Chemistry and Biology of Antitumor Enediyne Chromoprotein C-1027. BULLETIN OF THE CHEMICAL SOCIETY OF JAPAN 2006. [DOI: 10.1246/bcsj.79.501] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
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Abstract
AIM: To study the in vitro and in vivo antitumor effect of lidamycin (LDM) on hepatoma and the active moiety of its molecule.
METHODS: MTT assay was used to determine the growth inhibition of human hepatoma BEL-7402 cells, SMMC-7721 cells and mouse hepatoma H22 cells. The in vivo therapeutic effects of lidamycin and mitomycin C were determined by transplantable hepatoma 22 (H22) in mice and human hepatoma BEL-7402 xenografts in athymic mice.
RESULTS: In terms of IC50 values, the cytotoxicity of LDM was 10 000-fold more potent than that of mitomycin C (MMC) and adriamycin (ADM) in human hepatoma BEL-7402 cells and SMMC-7721 cells. LDM molecule consists of two moieties, an aproprotein (LDP) and an enediyne chromophore (LDC). In terms of IC50 values, the potency of LDC was similar to LDM. However, LDP was 105-fold less potent than LDM and LDC to hepatoma cells. For mouse hepatoma H22 cells, the IC50 value of LDM was 0.025 nmol/L. Given by single intravenous injection at doses of 0.1, 0.05 and 0.025 mg/kg, LDM markedly suppressed the growth of hepatoma 22 in mice by 84.7%, 71.6% and 61.8%, respectively. The therapeutic indexes (TI) of LDM and MMC were 15 and 2.5, respectively. By 2 iv. injections in two experiments, the growth inhibition rates by LDM at doses of 0.1, 0.05, 0.025, 0.00625 and 0.0125 mg/kg were 88.8-89.5%, 81.1-82.5%, 71.2-74.9%, 52.3-59.575%, and 33.3-48.3%, respectively. In comparison, MMC at doses of 5, 2.5, and 1.25 mg/kg inhibited tumor growth by 69.7-73.6%, 54.0-56.5%, and 31.5-52.2%, respectively. Moreover, in human hepatoma BEL-7402 xenografts, the growth inhibition rates by LDM at doses of 0.05 mg/kg × 2 and 0.025 mg/kg × 2 were 68.7% and 27.2%, respectively. However, MMC at the dose of 1.25 mg/kg × 2 showed an inhibition rate of 34.5%. The inhibition rate of tumor growth by LDM was higher than that by MMC at the tolerated dose.
CONCLUSION: Both LDM and its chromophore LDC display extremely potent cytotoxicity to hepatoma cells. LDM shows a remarkable therapeutic efficacy against murine and human hepatomas in vivo.
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Affiliation(s)
- Yun-Hong Huang
- Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, 1 Tiantan Xili, Beijing 100050, China
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Galm U, Hager MH, Van Lanen SG, Ju J, Thorson JS, Shen B. Antitumor Antibiotics: Bleomycin, Enediynes, and Mitomycin. Chem Rev 2005; 105:739-58. [PMID: 15700963 DOI: 10.1021/cr030117g] [Citation(s) in RCA: 431] [Impact Index Per Article: 21.6] [Reference Citation Analysis] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Affiliation(s)
- Ute Galm
- Division of Pharmaceutical Sciences and Department of Chemistry, University of Wisconsin, Madison, Wisconsin 53705, USA
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Usuki T, Inoue M, Hirama M, Tanaka T. Rational design of a supra C-1027: kinetically stabilized analogue of the antitumor enediyne chromoprotein. J Am Chem Soc 2004; 126:3022-3. [PMID: 15012111 DOI: 10.1021/ja039635z] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
C-1027, an extremely potent antitumor agent, is composed of a highly reactive chromophore and an apoprotein. While the chromophore causes DNA cleavage, the apoprotein functions as its carrier. Despite these ideal properties as an anticancer agent, C-1027 slowly self-decomposes through chromophore-mediated abstraction of hydrogens from the apoprotein. In this paper, we report the design and preparation of an engineered C-1027 apoprotein that decelerates this self-decomposition pathway. Our design is based on the kinetic isotope effect, and deuterium is incorporated instead of protium into the hydrogen-abstraction site. The deuterated supra C-1027 was found to have a 4-fold longer lifetime than the natural C-1027.
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Affiliation(s)
- Toyonobu Usuki
- Department of Chemistry, Graduate School of Science, Tohoku University, Sendai 980-8578, Japan
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Tanaka T, Fukuda-Ishisaka S, Hirama M, Otani T. Solution structures of C-1027 apoprotein and its complex with the aromatized chromophore. J Mol Biol 2001; 309:267-83. [PMID: 11491295 DOI: 10.1006/jmbi.2001.4621] [Citation(s) in RCA: 46] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/17/2023]
Abstract
C-1027 is one of the most potent antitumor antibiotic chromoproteins, and is a 1:1 complex of an enediyne chromophore having DNA-cleaving ability and a carrier apoprotein. The three-dimensional solution structures of the 110 residue (10.5 kDa) C-1027 apoprotein and its complex with the aromatized chromophore have been determined separately by homonuclear two-dimensional nuclear magnetic resonance methods. The apoprotein is mainly composed of three antiparallel beta-sheets: four-stranded beta-sheet (43-45, 52-54; 30-38; 92-94; 104-106), three-stranded beta-sheet (4-6; 17-22; 61-66), and two-stranded beta-sheet (70-72; 83-85). The overall structure of the apoprotein is very similar to those of other chromoprotein apoproteins, such as neocarzinostatin and kedarcidin. A hydrophobic pocket with approximate dimensions of 14 A x 12 A x 8 A is formed by the four-stranded beta-sheet and the three loops (39-42; 75-79; 97-100). The holoprotein (complex form with the aromatized chromophore) structure reveals that the aromatized chromophore is bound to the hydrophobic pocket found in the apoprotein. The benzodihydropentalene core of the chromophore is located in the center of the pocket and other substituents (beta-tyrosine, benzoxazine, and aminosugar moieties) are arranged around the core. Major binding interactions between the apoprotein and the chromophore are likely the hydrophobic contacts between the core of the chromophore and the hydrophobic side-chains of the pocket-forming residues, which is supplemented by salt bridges and/or hydrogen bonds. Based on the holoprotein structure, we propose possible mechanisms for the stabilization and the release of chromophore by the apoprotein.
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Affiliation(s)
- T Tanaka
- Institute of Applied Biochemistry, University of Tsukuba, Ibaraki, Japan.
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Liu W, Shen B. Genes for production of the enediyne antitumor antibiotic C-1027 in Streptomyces globisporus are clustered with the cagA gene that encodes the C-1027 apoprotein. Antimicrob Agents Chemother 2000; 44:382-92. [PMID: 10639366 PMCID: PMC89687 DOI: 10.1128/aac.44.2.382-392.2000] [Citation(s) in RCA: 71] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/1999] [Accepted: 11/10/1999] [Indexed: 11/20/2022] Open
Abstract
C-1027, the most potent member of the enediyne antitumor antibiotic family, is produced by Streptomyces globisporus C-1027 and consists of an apoprotein (encoded by the cagA gene) and a nonpeptidic chromophore. The C-1027 chromophore could be viewed as being derived biosynthetically from a benzoxazolinate, a deoxyamino hexose, a beta-amino acid, and an enediyne core. By adopting a strategy for cloning of the C-1027 biosynthesis gene cluster by mapping a putative dNDP-glucose 4,6-dehydratase (NGDH) gene to cagA, we have localized 75 kb of contiguous DNA from S. globisporus. DNA sequence analysis of two regions of the cloned gene cluster revealed two genes, sgcA and sgcB, that encode an NGDH enzyme and a transmembrane efflux protein, respectively, and confirmed that the cagA gene resides approximately 14 kb upstream of the sgcAB locus. The involvement of the cloned gene cluster in C-1027 biosynthesis was demonstrated by disrupting the sgcA gene to generate C-1027-nonproducing mutants and by complementing the sgcA mutants in vivo to restore C-1027 production. These results represent the first cloning of a gene cluster for enediyne antitumor antibiotic biosynthesis and provide a starting point for future genetic and biochemical investigations of C-1027 biosynthesis.
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Affiliation(s)
- W Liu
- Department of Chemistry, University of California, Davis, California 95616, USA
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Imajo S, Ishiguro M, Tanaka T, Hirama M, Teplyakov A. On the conformation of Phe78 of a chromoprotein antibiotic, neocarzinostatin. Bioorg Med Chem 1995; 3:429-36. [PMID: 8581426 DOI: 10.1016/0968-0896(95)00032-c] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
A structure of neocarzinostatin, an antitumor chromoprotein antibiotic, has been built using X-ray crystallographic data and NMR data, particularly NOE data observed between the apoprotein and the chromophore. Chemical shift changes of protons of the chromophore upon binding to the apoprotein indicated that the aromatic plane of Phe52 has the conformation almost perpendicular to the C-2-C-3 triple bond of the core of the chromophore while Phe78 takes multiple conformations in solution although one of the stable conformations has been assigned for Phe78 in a crystal structure.
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Affiliation(s)
- S Imajo
- Suntory Institute for Biomedical Research, Osaka, Japan
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