Rapid and sensitive detection of UGT1A1 polymorphisms associated with irinotecan toxicity by a novel DNA microarray

近年のヒトゲノム解析における発展は,精密で個別化された医療の可能性を大いに高めてきた.我々は精密医療実現のために,体外診断用医薬品の遺伝子解析技術として,DNAマイクロアレイを新規に開発した.本技術は,PCR法と核酸ハイブリダイゼーション法を組み合わせた方法であり,目的塩基配列のPCR法による増幅･蛍光標識と3mm角のDNAチップ上に固定化されたDNAプローブとの特異的ハイブリダイゼーション反応とを組み合わせた方法を採用している.また,チッフﾟ表面をdiamond-like carbon層にて処理することでシグナル/ノイズ比を向上させた.性能評価として,抗がん剤イリノテカンの重篤な毒性と有意に関連し,検査対象となっているUGT1A1*28及びUGT1A1*6遺伝子多型をヒト臨床検体を用いて測定した.比較として既存法の直接シーケンシングとインベーダーアッセイを併せて実施した.本技術と既存法との測定一致率は100%であった.一塩基置換であるUGT1A1*6,TAリピートであるUGT1A1*28に加えて,一塩基挿入/欠失を含む他のUGT1A多型も同時に精確な判定が可能であった.処理時間(1.5倍以上の短縮)や必要検体量(20分の1以下)はインベーダー法と比べて優れていた.本技術によって,遺伝子多型や変異を複数同時に迅速かつ正確に測定でき,臨床現場での省力･効率や信頼性の向上が期待される.

Recent developments in the field of human genomics have greatly enhanced the potential for precision and personalized medicine. We have developed a novel DNA microarray, using a 3-mm square chip coated with diamond-like carbon to enhance the signal-to-background ratio, for use as an in vitro diagnostic tool in precision medicine. To verify the genotyping effectiveness of this newly developed DNA microarray we examined UDP-glucuronosyltransferase 1A1(UGT1A1)polymorphisms in DNA extracted from patients with metastatic colorectal cancer. It is established that the polymorphisms of UGT1A1*28 and UGT1A1*6 are significantly associated with severe toxicity induced by the anti-cancer drug irinotecan. For each sample, the results obtained with the novel microarray platform were compared with those obtained using other, more established, methods, including direct sequencing and the Invader assay. The polymorphisms tested included a single nucleotide substitution(UGT1A1*6)and a TA-repeat polymorphism(UGT1A1*28),both of which were detected simultaneously and accurately using our method. Moreover, our method required 1.5-fold less time to assay and 20-fold less sample than those required by the Invader assay. In summary, our newly developed DNA microarray is more practical than established methods, and is at least as accurate

DNAマイクロアレイ

体外診断用医薬品

イリノテカン

多型

精密医療