A radioisotope-nondependent high-sensitivity method for measuring the activity of glioblastoma-related O-methylguanine DNA methyltransferase |
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| Authors: | Aya Hongo Ran Gu Miho Suzuki Naoto Nemoto Koichi Nishigaki |
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| Institution: | 1. Department of Functional Materials Science, Graduate School of Science and Engineering, Saitama University, Sakura-ku, Saitama 338-8570, Japan;2. Rational Evolutionary Design of Advanced Biomolecules, Saitama (REDS), Saitama Small Enterprise Promotion Corporation, No. 552, Kawaguchi, Saitama 333-0844, Japan |
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| Abstract: | O6-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. |
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| Keywords: | MGMT RI-nondependent activity assay Sensitivity enhancement by PCR Glioblastoma TMZ |
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