Electrospray Mass Spectrometry of Iron Bleomycin: Demonstration That Activated Bleomycin Is a Ferric Peroxide Complex

Sam, Joseph; Tang, Xuejun; Peisach, Jack

doi:10.1021/ja00091a032

Cited by 222 publications

(173 citation statements)

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“…ABLM is proposed to be a low spin (ls) Fe III ─OOH species (9)(10)(11), that can be formed via the reaction of Fe III BLM with O 2 and an electron or through a shunt reaction between Fe III BLM and H 2 O 2 (8,12).…”

mentioning

confidence: 99%

Definition of the intermediates and mechanism of the anticancer drug bleomycin using nuclear resonance vibrational spectroscopy and related methods

Liu

Bell

Wong

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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Bleomycin (BLM) is a glycopeptide anticancer drug capable of effecting single-and double-strand DNA cleavage. The last detectable intermediate prior to DNA cleavage is a low spin Fe III peroxy level species, termed activated bleomycin (ABLM). DNA strand scission is initiated through the abstraction of the C-4′ hydrogen atom of the deoxyribose sugar unit. Nuclear resonance vibrational spectroscopy (NRVS) aided by extended X-ray absorption fine structure spectroscopy and density functional theory (DFT) calculations are applied to define the natures of Fe III BLM and ABLM as ðBLMÞFe III ─ OH and ðBLMÞFe III ðη 1 ─OOHÞ species, respectively. The NRVS spectra of Fe III BLM and ABLM are strikingly different because in ABLM the δFe─O─O bending mode mixes with, and energetically splits, the doubly degenerate, intense O─Fe─N ax transaxial bends. DFT calculations of the reaction of ABLM with DNA, based on the species defined by the NRVS data, show that the direct H-atom abstraction by ABLM is thermodynamically favored over other proposed reaction pathways.nonheme iron | structure/reactivity T he glycopeptide antibiotic bleomycin (BLM) exhibits high intrinsic anticancer cytotoxicity, and is used in chemotherapy against head, neck, and testicular cancers as well as Hodgkin lymphoma (1, 2). Its cytotoxicity is due to its ability to effect singleand double-strand cleavage of DNA with a number of reduced metal ions and dioxygen (3-5); Fe II displays the highest in vivo activity (6). BLM provides five nitrogen-based ligands ( Fig. 1 A and B) for coordination to the metal ion based on a crystal structure of peroxy Co III BLM (7). The four equatorial ligands are an imidazole nitrogen, a deprotonated amide, a pyrimidine nitrogen, and the secondary amine of the β-aminoalanine. The axial ligand is the primary amine of the β-aminoalanine.Activated BLM (ABLM) is the final intermediate detected before DNA double-strand scission (8). ABLM is proposed to be a low spin (ls) Fe III ─OOH species (9-11), that can be formed via the reaction of Fe III BLM with O 2 and an electron or through a shunt reaction between Fe III BLM and H 2 O 2 (8, 12).A prominent issue has been whether ABLM, a nonheme iron complex, performs heme type chemistry similar to P450 and peroxidase in which heterolytic cleavage of the O─O bond results in an Fe IV species with a radical on the BLM ligand. Experiments and calculations have argued against this heterolytic mechanism and suggested a mechanism where ABLM reacts directly in H-atom abstraction from the DNA backbone sugar (10,13,14). However, a recent theoretical study has postulated that ABLM formation involves reaction of the ðBLMÞFe III ─OH 2 complex with H 2 O 2 to form a ðBLMÞFe III ─O 2 H 2 complex (15). This study found the heterolytic cleavage pathway for the ðBLMÞFe III ─ H 2 O 2 complex to be thermoneutral and therefore feasible. Because ABLM is inaccessible to vibrational characterization by resonance Raman (rR) spectroscopy due to photodecay (16), we employed nuclear resonance vibrational spectroscopy (...

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mentioning

confidence: 99%

Definition of the intermediates and mechanism of the anticancer drug bleomycin using nuclear resonance vibrational spectroscopy and related methods

Liu

Bell

Wong

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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“…Fe II OBLM reacts with O 2 and an exogenous electron to generate activated BLM (ABLM), which is kinetically competent to cleave DNA by H-atom abstraction (25). ABLM has been determined to be a low-spin Fe III OOOH complex (26)(27)(28)(29). We have used a combination of spectroscopic techniques and density functional calculations to elucidate the geometric and electronic structure of ABLM and obtain insight into its reactivity (30).…”

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confidence: 99%

Non-heme iron enzymes: Contrasts to heme catalysis

Solomon

Decker

Lehnert

2003

Proc. Natl. Acad. Sci. U.S.A.

222

187

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Non-heme iron enzymes catalyze a wide range of O2 reactions, paralleling those of heme systems. Non-heme iron active sites are, however, much more difficult to study because they do not exhibit the intense spectral features characteristic of the porphyrin ligand. A spectroscopic methodology was developed that provides significant mechanistic insight into the reactivity of non-heme ferrous active sites. These studies reveal a general mechanistic strategy used by these enzymes and differences in substrate and cofactor interactions dependent on their requirement for activation by iron. Contributions to O2 activation have been elucidated for non-heme relative to heme ligand sets, and major differences in reactivity are defined with respect to the heterolytic and homolytic cleavage of OOO bonds.

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Section: Discussionmentioning

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“…It is now recognized that a discordance may exist between the appearance of the classic morphological and biochemical features of apoptosis and loss of cellular clonogenic potential (29,30). To assess the biological consequences of p21 WAF1/CIP1 dysregulation, the viability of ara-C-treated empty vector and p21 antisense-expressing cells was compared using trypan blue exclusion and clonogenic assays (Fig.…”

Section: P21 Dysfunction Potentiates Ara-c-induced Apoptosismentioning

confidence: 99%

“…It is noteworthy that p21 antisenseexpressing cells exposed to ara-C displayed a relatively modest enhancement in ROS generation, consistent with the notion that induction of ROS lags behind and represents a consequence of earlier rnitochondrial damage (41). It is also important to note that the increased susceptibility of p21 WAF1/CIP1 antisense-expressing cells to ara-C-mediated rnitochondrial dysfunction was accompanied by a loss in leukemic cell self-renewal capacity, particularly in view of reports suggesting a discordance between apoptosis and loss of clonogenic potential (27,30, 44).Although disruption of p21 WAF1/CIP1 function has been shown to increase the sensitivity of tumor cells to various agents (8-10), this has not been a universal finding. For example, antisense-mediated disruption of p21 WAF1/CIP1 reduced antioxidant-related apoptosis and potentiation of 5-FU-mediated lethality in human colon cancer cells (HCT 15 and HCT 116) in a p53-independent manner (45).…”

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confidence: 99%

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DNA Cleavage/Repair and Signal Transduction Pathways in Irradiated Breast Tumor Cells

Gewirtz¹

2001

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Public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing this collection of information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing this burden to Washington Headquarters Services, Directorate for Information Operations and Reports, 1215 Jefferson Davis Highway, Suite 1204, Arlington, VA 22202-4302, and to the Office of Management and Budget, Paperwork Reduction Project (0704-0188), Washington, DC 20503 AGENCY USE ONLY (Leave blank) REPORT DATE September 1999 REPORT TYPE AND DATES COVEREDannual (1 Sep 98 -31 Aug 99) TITLE AND SUBTITLE DNA Cleavage/Repair and Signal Transduction Pathways in Irradiated Breast Tumor Cells AUTHOR(S)David A. Gewirtz, Ph.D. FUNDING NUMBERS DAMD17-96-1-6167 PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) Virginia Commonwealth UniversityRichmond, Virginia 23298-0568 E-MAIL: gewirtz@hsc.vcu.edu We have determined that neither p53 status nor alterations in levels of the Myc protein are critical factors in radiosensitivity or in sensitivity to adriamycin. The refractoriness of breast tumor cells to DNA damage induced apoptosis may be related, in part, to upregulation of p21 as well as to stimulation of MAP kinase activity. Our current studies using cells carrying a p21 antisense vector should clarify the role of p21 while studies relating to the MAP kinase pathway are being initiated. Pretreatment of p53 wild type breast tumor cells with Vitamin D3 compounds sensitizes the cells to ionizing radiation and to adriamycin, in part through the promotion of apoptosis -suggesting that the Vitamin D compounds can be used to enhance the effectiveness of radiotherapy and chemotherapy in the clinical treatment of breast cancer. Finally, we are attempting to determine how p53 status influences the fidelity of double-strand break repair in apoptosis-proficient 184B5 breast epithelial cells, and the possible relation between apoptosis and tolerance for misrepair. Such studies may suggest additional candidates for transgenic manipulation of the response to radiation. y Where copyrighted material is quoted, permission has been obtained to use such material. PERFORMING ORGANIZATION REPORT NUMBER SPONSORING / MONITORING AGENCY NAME(S) AND ADDRESS(ES)U SUBJECT TERMS Breast CancerWhere material from documents designated for limited distribution is quoted, permission has been obtained to use the material.Citations of commercial organizations and trade names in this report do not constitute an official Department of Army endorsement or approval of the products or services of these organizations.N/A In conducting research using animals, the investigator(s) adhered to the "Guide for the Care and Use of Laboratory Animals," prepared by the Committee on Care and use of Laboratory Animals of the Institute of Laboratory Resources, ...

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Electrospray Mass Spectrometry of Iron Bleomycin: Demonstration That Activated Bleomycin Is a Ferric Peroxide Complex

Cited by 222 publications

References 4 publications

Definition of the intermediates and mechanism of the anticancer drug bleomycin using nuclear resonance vibrational spectroscopy and related methods

Definition of the intermediates and mechanism of the anticancer drug bleomycin using nuclear resonance vibrational spectroscopy and related methods

Non-heme iron enzymes: Contrasts to heme catalysis

DNA Cleavage/Repair and Signal Transduction Pathways in Irradiated Breast Tumor Cells

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