title: A radioisotope-nondependent high-sensitivity method for measuring the activity of glioblastoma-related O(6)-methylguanine DNA methyltransferase.
abstract: O(6)-Methylguanine DNA methyltransferase (MGMT) cancels the anticancer effect of temozolomide (drug for glioblastoma), which introduces methylation to DNA. Therefore, developing an MGMT inhibitor is a promising strategy for the treatment of this cancer. For this purpose, a sensitive detection method that does not depend on the conventional radioisotope (RI) method was developed. This was realized by a fluorescence-based method that measured the amount of cleavable restriction sites demethylated by the action of MGMT; this method was enhanced by introducing a polymerase chain reaction (PCR) amplification step. As an assay of enzyme activity, 20-fold higher sensitivity (subnanomolar) was attained compared with our and others' fluorescence-based approaches.
authors.affiliation: Department of Functional Materials Science, Graduate School of Science and Engineering, Saitama University, Sakura-ku, Saitama 338-8570, Japan.
authors.forename: Aya
authors.initials: A
authors.lastname: Hongo
authors.affiliation: Department of Functional Materials Science, Graduate School of Science and Engineering, Saitama University, Sakura-ku, Saitama 338-8570, Japan.
authors.forename: Ran
authors.initials: R
authors.lastname: Gu
authors.affiliation: Department of Functional Materials Science, Graduate School of Science and Engineering, Saitama University, Sakura-ku, Saitama 338-8570, Japan; Rational Evolutionary Design of Advanced Biomolecules, Saitama (REDS), Saitama Small Enterprise Promotion Corporation, No. 552, Kawaguchi, Saitama 333-0844, Japan.
authors.forename: Miho
authors.initials: M
authors.lastname: Suzuki
authors.affiliation: Department of Functional Materials Science, Graduate School of Science and Engineering, Saitama University, Sakura-ku, Saitama 338-8570, Japan; Rational Evolutionary Design of Advanced Biomolecules, Saitama (REDS), Saitama Small Enterprise Promotion Corporation, No. 552, Kawaguchi, Saitama 333-0844, Japan.
authors.forename: Naoto
authors.initials: N
authors.lastname: Nemoto
authors.affiliation: Department of Functional Materials Science, Graduate School of Science and Engineering, Saitama University, Sakura-ku, Saitama 338-8570, Japan; Rational Evolutionary Design of Advanced Biomolecules, Saitama (REDS), Saitama Small Enterprise Promotion Corporation, No. 552, Kawaguchi, Saitama 333-0844, Japan. Electronic address: koichi@fms.saitama-u.ac.jp.
authors.forename: Koichi
authors.initials: K
authors.lastname: Nishigaki
mesh_terms: D004789:Enzyme Activation; D005453:Fluorescence; D005909:Glioblastoma / Q000201:enzymology* / Q000378:metabolism; D006801:Humans; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000032:analysis* / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011868:Radioisotopes
chemical_list: D011868:Radioisotopes; D019853:O(6)-Methylguanine-DNA Methyltransferase
keywords: Glioblastoma; MGMT; RI-nondependent activity assay; Sensitivity enhancement by PCR; TMZ