- Synthesis of Mitomycin C and Decarbamoylmitomycin C N2 deoxyguanosine-adducts
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Mitomycin C (MC) and Decarbamoylmitomycin C (DMC) - a derivative of MC lacking the carbamate on C10 - are DNA alkylating agents. Their cytotoxicity is attributed to their ability to generate DNA monoadducts as well as intrastrand and interstrand cross-lin
- Champeil, Elise,Cheng, Shu-Yuan,Huang, Bik Tzu,Conchero-Guisan, Marta,Martinez, Thibaut,Paz, Manuel M.,Sapse, Anne-Marie
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- Reaction of reductively activated mitomycin C with aqueous bicarbonate: Isolation and characterization of an oxazolidinone derivative of cis-1-hydroxy-2,7-diaminomitosene
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The reductive activation of mitomycin C in aqueous bicarbonate buffer resulted in the formation of a previously unknown compound, characterized as an oxazolidinone derivative of cis-1-hydroxy-2,7-diaminomitosene. This compound is the result of a cyclization reaction of bicarbonate with the aziridine ring of aziridinomitosene, and was observed at bicarbonate concentrations close to those present in physiological plasma.
- Paz, Manuel M.
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experimental part
p. 31 - 34
(2010/04/02)
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- A mitomycin-N6-deoxyadenosine adduct isolated from DNA.
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A minor N6-deoxyadenosine adduct of mitomycin C (MC) was isolated from synthetic oligonucleotides and calf thymus DNA, representing the first adduct of MC and a DNA base other than guanine. The structure of the adduct (8) was elucidated using submilligram quantities of total available material. UV difference spectroscopy, circular dichroism, and electrospray mass spectroscopy as well as chemical transformations were utilized in deriving the structure of 8. A series of synthetic oligonucleotides was designed to probe the specificities of the alkylation of adenine by MC. The nature and frequency of the oligonucleotide-MC adducts formed under conditions of reductive activation of MC were determined by their enzymatic digestion to the nucleoside level followed by quantitative analysis of the products by HPLC. The analyses indicated the following: (i) (A)n sequence is favored(AT)n for adduct formation; (ii) the alkylation favors the duplex structure; (iii) at adenine sites only monofunctional alkylation occurs; (iv) the adenine-to-alkylation frequency in the model oligonucleotides was 0.3-0.6 relative to guanine alkylation at the 5'-ApG sequence but only 0.02-0.1 relative to guanine alkylation at 5'-CpG. The 5'-phosphodiester linkage of the MC-adenine adduct is resistant to snake venom diesterase. The overall ratio of adenine to guanine alkylation in calf thymus DNA was 0.03, indicating that 8 is a minor MC-DNA adduct relative to MC-DNA adducts at guanine residues in the present experimental residues in the present experimental system. However, the HPLC elution time of 8 coincides with that of a major, unknown MC adduct detected previously in mouse mammary tumor cells treated with radiolabeled MC [Bizanek, R., Chowdary, D., Arai, H., Kasai, M., Hughes, C. S., Sartorelli, A. C., Rockwell, S., and Tomasz, M. (1993) Cancer Res. 53, 5127-5134]. Thus, 8 may be identical or closely related to this major adduct formed in vivo. This possibility can now be tested by further comparison.
- Palom, Jolanda,Lipman, Roselyn,Musser, Steven M.,Tomasz, Maria
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p. 203 - 210
(2007/10/03)
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- Studies on the reactivity of reductively activated mitomycin C
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Mitomycin C (1a), a clinically significant antineoplastic antiobiotic, is considered to be the prototype of bioreductive alkylating agents. It has been reported that, in the absence of DNA, reductive activation of 1a furnished both solvolytic C(1) electro
- Schiltz, Pascal,Kohn, Harold
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p. 10510 - 10518
(2007/10/02)
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- Studies on the use of Na2S2O4 for the reductive activation of mitomycin C
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Mitomycin C (1a) is considered to be the prototypical bioreductive alkylating agent. Among the numerous reductive procedures employed for the in vitro activation of mitomycin C, incremental addition of Na2S2O4 has emerged as the method of choice for generating high yields of mitomycin C-DNA adducts. The major products and distinguishing features of the incremental addition Na2S2O4-mitomycin C reductive processes in water (pH 7.4) in the absence of DNA are reported. Key observations included (1) rapid and efficient consumption of mitomycin C, (2) production of high amounts of 7-aminomitosane-9a-sulfonate (1b) in the early stages of the reaction, and (3) generation of significant amounts of C(1) and C(10) sulfonato adducts. The complexity of this transformation has been attributed in part to HSO3-, a byproduct of the Na2S2O4 reduction process. Use of buffered methanol solutions ("pH" 7.4) in place of water simplified the product profile. The poor solubility of Na2S2O4 and NaHSO3 in methanol produced only trace amounts of mitosene sulfonato adducts. There were significant differences between product profiles for the incremental addition Na2S2O4 procedure versus a protocol in which the equivalent amount of Na2S2O4 was added in a single shot. First, higher amounts of C(1) electrophilic versus C(1) nucleophilic products were observed using the single shot technique. Second, C(1) sulfonato adducts composed a larger amount of the C(1) nucleophilic product pool when the Na2S2O4 was added using the single shot protocol than with the incremental addition method. Third, higher amounts of 1a were converted to C(1), C(10) fully functionalized mitosene adducts using the incremental procedure. Select auxiliary experiments provided additional information concerning the Na2S2O4-mediated mitomycin C reductive process. Examination of the reactivity of key C(1), C(9a), and C(10) mitomycin sulfonato products demonstrated that 7-aminomitosane-9a-sulfonate (1b) was efficiently converted to C(1)- and C(10)-functionalized mitosenes under reductive conditions, whereas mitosene C(1) and C(10) sulfonates did not undergo displacement reactions and hence did not function as viable alkylating agents. On the basis of these cumulative studies, we suggest the likely mechanism for the Na2S2O4-mediated mitomycin C reductive process and the beneficial properties accrued by the use of the incremental addition technique. These notions are discussed in light of the pathway that may be operative in in vitro mitomycin C-DNA bonding transformations.
- Schiltz, Pascal,Kohn, Harold
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p. 10497 - 10509
(2007/10/02)
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- Sodium dithionite-mediated mitomycin C reductive activation processes
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Sodium dithionite is the reagent of choice for the reductiue activation of mitomycin C in the presence of DNA. No comprehensive study of this transformation in the absence of DNA has appeared. The major products of this reaction have been determined and the key parameters governing this transformation identified.
- Schiltz, Pascal,Kohn, Harold
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p. 4709 - 4712
(2007/10/02)
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- 7-Aminoaziridinomitosenes: Synthesis, Structure, and Chemistry
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7-Aminoleucoaziridinomitosene (2a) has been proposed as a key intermediate in the reductive activation process for the antineoplastic agent, mitomycin C (1a).Little is known about 2a and its oxidised equivalent, 7-aminoaziridinomitosene (3a).An expedient electrochemical procedure for 3a and the corresponding N-methyl analogue 3b has been developed.NMR spectral studies of 3a in DMF-d7 and DMSO-d6 provided important information concerning the solution-state structure for this adduct.Factors controlling the aziridine ring-opening process under reductive and nonreductive conditions have been determined, as well as evidence for the intermediacy of 2a in the reductive activation cascade of 1a.
- Han, Insook,Kohn, Harold
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p. 4648 - 4653
(2007/10/02)
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- Studies on the Use of Cr(ClO4)2 for the Reductive Activation of Mitomycin C 1
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Cr(ClO4)2 has been shown to be a highly efficient reductant of the anticancer agent, mitomycin C (1). Two different Cr(ClO4)2-mediated reductive techniques were developed and utilized in buffered water and methanolic solutions. In the first procedure, Cr(ClO4)2 (1-2 equiv) was directly added to 1 at various "pH" values. Key observations included the following: (1) Consumption of mitomycin C was rapid and generated as the major products trans- and cis-10-decarbamoyl-1-hydroxy-2,7-diaminomitosenes (11 and 12), and trans- and cis-10-decarbamoyl-1,10-dimethoxymitosenes (16 and 17) in acidic-to-neutral aqueous and methanolic solutions, respectively. (2) Between "pH" 6.0 and 7.0, the difunctionalized mitosene adducts accounted for nearly half of the product profile even though noticeable amounts of unreacted 1 remained. (3) Significant amounts of C-1 electrophilic products were not observed under acidic conditions. The product profiles observed with the second Cr(ClO4)2-mediated reductive procedure were markedly different. Activation of 1 was accomplished by the prior addition of Cr(ClO4)2 to excess cis-10-decarbamoyl-1,10-dimethoxy mitosene (17) to generate the putative mitosene monochromate 20 and mitosene dichromate 21 species in situ, followed by the addition of 1 (1 equiv per Cr(ClO4)2). The products obtained by using this protocol were similar to those observed with conventional reductants in which C-1 electrophilic adducts predominated in acid, C-1 nucleophilic products were the major products under neutral and basic conditions, and little modification of the C-10 site was detected throughout the "pH" range examined. The product profiles coupled with select auxiliary experiments have provided information concerning the mechanism of both reductive procedures. The major products furnished by using the direct Cr(ClO4)2-mediated procedure under acidic and neutral conditions have been attributed to the two one-electron reductions of 1 to give the bis-CrIII-bound species 22. Complexation of the C-5 and C-8 phenolic-type oxygens in reduced 1 is believed to facilitate the loss of methanol at C-9 and C-9a in 1 and the nucleophilic substitution processes at C-1 and C-10 as well as inhibit the electrophilic transformations at both DNA bonding sites. Explanations and supporting data have also been provided to account for the other products detected in these reactions. Correspondingly, the second procedure is conjectured to occur by an outer-sphere electron-transfer process from 20 and/or 21 to 1 to give the uncomplexed hydroquinone (or semiquinone) mitomycin C species 2. Subsequent loss of methanol at C-9 and C-9a yields the activated mitosene capable of furnishing the C-1 functionalized adducts 7 and 9 + 10. The distinctive product profiles observed with the direct addition of Cr(ClO4)2 to 1 and the remarkable high yields of C-1, C-10 dinucleophilic substitution adducts suggest that similar pathways may be operative in the in vivo process to provide the DNA-mitomycin C cross-link adducts. These notions are discussed in light of the DNA sequence selectivity recently observed for the drug monoalkylation bonding process.
- Hong,Kohn, Harold
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p. 4634 - 4644
(2007/10/02)
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- DEVELOPMENT OF A HYDRAZINE-MEDIATED SYSTEM FOR THE REDUCTIVE ACTIVATION OF MITOMYCIN C
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The reactivity of mitomycin C (1) with hydrazines (2) has been examined.Monoarylhydrazines have been shown to efficiently reduce 1 leading to the activation of both proposed DNA binding sites (C-1 and C-10) within the drug.
- Russell, David J.,Kohn, Harold
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p. 223 - 227
(2007/10/02)
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