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Gapmer型アンチセンスの肝毒性発現に関する研究

釘宮, 啓 東京大学 DOI:10.15083/0002006086

2023.03.20

概要





















釘宮



アンチセンス核酸やsi RNA など核酸医薬品は、 従来からの低分子医薬品や抗体医薬品などが
アクセスすることが困難な遺伝子転写産物(RNA)を創薬ター ゲットにできる特徴を有してお
り、次世代の創薬モダリティとして期待されている。それらの核酸医薬品の中で もGapmer型ア
ンチセンス(以下、Gapmerと呼ぶ)は、 現在までに3品目上市されており注目を集めている。
一方、Gapmerは臨床および非臨床において肝毒性懸念があることが知られている。 この肝毒性
発現メカニズムについては、 従来から研究がおこなわれているものの未だ詳細に解明されてい
ない。 今後、Gapmerを安全な核酸医薬品として継続的に活用していくためには毒性発現の要因
を明らかにすることが不可避となっている。 そこで「Gapmer型アンチセンスの肝毒性発現に関
する研究」と題する本論文おいて釘宮は、その要因について主にメカニズム解明に焦点をあてた
研究をおこなった。 具体的には、 薬理活性および肝毒性の双方が強力なLNA修飾Gapmerをモ
デルとして用いて、 核酸化合物自体が持つ毒性やRNA 切断機構と肝毒性の関連性、 標的 RNA
に対する配列特異性解析などについて検討した。
第1章では、 肝毒性を示すGapmerの中央部、 標的mRNAの切断に重要な部分を化学修飾し
たNon-Gapmerを設計· 合成しマウスに投与することで、 その薬理活性と肝毒性発現について
Gapmerと比較検討した。その結果、Non-Gapmer投与群でのみノックダウン活性と肝毒性が同時
に消失したことから、 核酸化合物自体による毒性惹起ではないことを明らかとした。 第2章で
は、Gapmerの薬理活性に必須な肝臓中RNaseHIの発現をsiRNAにより抑制したところ、Gapmer
投与によるノックダウン活性と肝毒性 が減弱もしくは消失することを見出した。 これらのこと
から、Gapmerによる肝毒性はRNase HIの活性に依存することを示した。 第3章では、 肝壽性
RNA領域を標的とするsiRNAを設計・合成し、 マウスに投与した。
を示すGapmerと同一 のm
その結果、siRNAを用いて肝臓中の標的遣伝子をGapmerと同程度にノックダウンしても肝毒性
が惹起されなかったことから、Gapmerによる肝毒性はオンター ゲット作用ではないことを明ら
かにした。 第4章では、Gapmerはpre-m
RNAを切断することから作用部位は核内であることを
示した。 第5章では、 ゲノムデー タベ ー スやマイクロアレイデー タを用いて、Gapmerによるオ
フター ゲット作用を検討した。その結果、 肝毒性を示すGapmerはオフター ゲット作用により核
内で標的RNA以外の標的類似配列の非特異切断を引き起こし、 肝毒性を示さないGapmerは標
的類似配列の非特異切断を引き起こさないことを見出した。 以上の結果から釘宮は、 Gapmerに
よる肝毒性発現の主要因はRNase HIを介したRNAの非特異的な切断であることを明らかにし
た。また、 毒性を示さないGapmerを創製するためには、 ゲノムレベルで配列特異性の担保や標
的特異性の確保が重要となる可能性を示すことができた。
これらの研究成果は核酸医薬品の研究開発に貢献するものであり、 よって本論文は博士(薬科
学)の学位請求論文として合格と認められる。

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