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消化器障害バイオマーカーとしての尿素サイクル関連代謝物の有用性に関する研究

齊藤, 航 筑波大学 DOI:10.15068/00160472

2020.07.22

概要

未充足の医療ニーズを満たす医薬品を継続的に創出するためには、科学の進歩と研究開発のプロセス改善が欠かせない。医薬品の研究開発では、探索研究から製造販売承認取得までに平均10年以上の歳月を要し、その成功確率は4.9%と報告されている(1)。近年、疾患と関連する標的分子の枯渇に加え、規制当局による承認や保険者による償還の厳格化を背景に、医薬品の成功確率は低下し、研究開発費は上昇している。そのため、製薬会社には成功確率の向上と研究開発費の抑制が求められている(2)。

研究開発を効率化する取組みとして、製薬会社はトランスレーショナルリサーチ、すなわち非臨床と臨床の橋渡し研究に注力している。従来の医薬品開発では、動物を用いた非臨床試験で有効性や安全性を検証してきたが、必ずしも動物とヒトで一貫した臨床試験成績が得られなかった。そこでヒトの有効性や安全性の予測性を向上させるため、人工多能性幹細胞(iPS)細胞、血液や組織等の臨床サンプル、日常の診療記録から構成されるリアルワールドデータ、ゲノム情報等のヒト由来の試料や情報を用いた研究が進められている(Fig.1)。バイオマーカー研究は、このようなトランスレーショナルリサーチの1つである。動物とヒト共通の指標を用いて、医薬品の有効性や安全性を早期に評価し、医薬品の研究開発を効率化することが期待されている(3)。

バイオマーカーとは、『通常の生物学的過程、病理学的過程、又は治療的介入に対する薬理学的応答の指標として、客観的に測定され評価される特性』と定義されている(4)。その測定には遺伝子(DNA、RNA)、タンパク、ペプチド、内因性代謝物、イメージング等が用いられ、臨床での疾患の診断、患者層別化、薬理作用や安全性評価などの様々な目的で利用されている(Table1)。その一例として、腎毒性の安全性バイオマーカー研究が挙げられる。これまで腎毒性評価では、血中クレアチニンや尿素窒素が用いられてきたが、感度が低く、早期の腎毒性を検出できないことが知られていた。近年、Kim-1、クラスタリン、アルブミンなどの複数の尿中タンパクが、感度及び特異度の高い新規の安全性バイオマーカーであると報告されている(5,6)。現在、国内外の製薬会社は安全性バイオマーカーのコンソーシアムを形成し、米国食品医薬品局(FDA)、欧州医薬品庁(EMEA)、独立行政法人医薬品医療機器総合機構(PMDA)などの規制当局やアカデミアと連携しながら、腎臓、肝臓、骨格筋などの臓器毒性に対する新規バイオマーカーの利活用に向けた検討を進めている(7)。

医薬品の非臨床安全性試験においても、バイオマーカーの活用が期待されている(3)。病理組織検査は臓器毒性の確定診断として有用であるが、同一動物の経時的な評価はできない。また、標本作成から鏡検まで時間を要するため、効率が求められるスクリーニングには不向きである。一方、バイオマーカーによる安全性評価では、血液や尿などの低侵襲性サンプルを経時的に採取して、迅速な測定を行い、毒性を早期に検出できる。医薬品開発において、臨床試験開始直前の非臨床試験や臨床の第一相試験の成功率は低く、その開発中止理由の約4~5割は安全性に起因すると報告されている(8,9)。そのため、感度及び特異性の高いバイオマーカーを用いて、安全性の高い医薬品候補を早期に取得できれば、医薬品開発の成功確率向上及び費用削減に貢献できると考えられる。

肝臓及び消化管を含む消化器は、食物を体内に取り入れて貯蔵、消化、吸収及び不消化物の排泄を行うだけでなく、医薬品の代謝、毒性、生体防御の点で関連する。経口摂取された医薬品の多くは肝臓で代謝されるが、消化管でも肝臓の1/10程度の薬物代謝酵素を発現し、薬物の血中濃度に影響を与える。また、腸内細菌も薬物の活性化や不活性化に関与する。例えば、肝臓で抱合され胆管から排泄された薬剤が、腸内細菌叢により脱抱合され、消化管毒性を誘発することが知られている(10)。さらに、腸内細菌、エンドトキシン、又は細菌成分が腸管上皮を通過して体内に侵入し、肝硬変等の肝疾患を増悪させることが報告され(11)、肝臓と消化管は腸管軸として生体防御で重要な役割を果たしていると考えられている。

消化器毒性は医薬品の開発でよく見られる副作用であるが、これらの毒性に対して万能な安全性バイオマーカーは存在しない。肝毒性は、臨床試験中の副作用により開発中止に至る主な原因の1つである(9)。従来の低分子医薬品に加え、近年、研究開発が活発化している遺伝子発現を制御する核酸医薬品も、非特異的な肝障害を生じさせることが知られている(12,13)。アラニンアミノトランスフェラーゼ(ALT)は、主に肝臓の細胞質中に存在する代謝酵素であり、肝毒性バイオマーカーとして非臨床や臨床で広く用いられている。しかし、ALTは骨格筋障害でも軽度に上昇し、その特異性に課題がある(14)。消化管毒性は、抗がん剤や非ステロイド性消炎鎮痛薬でみられ、用量制限が必要となる毒性である。また、経口投与後の薬物の吸収にも影響を与え、疾患の管理に影響を与えることがある。消化管毒性は主に病理組織学的検査で評価されており、現時点で感度及び特異性の優れたバイオマーカーは確立されていない。このような背景から、肝臓や消化管の病理組織変化を感度よく反映し、既存のバイオマーカーより特異性の高い新規のバイオマーカーが求められている。

血中に存在するアミノ酸のうち、尿素サイクル関連代謝物であるアルギニン及びシトルリンが、消化器障害を反映する代謝性バイオマーカーとなる可能性が報告されている(Fig.2)(15,16)。一方で、代謝性バイオマーカーを非臨床安全性試験で利用するために、病理組織変化や既存のバイオマーカーと比較し、その感度及び特異性を詳細に検討した研究はない。そこで本研究では、消化器障害時の血中の尿素サイクル関連代謝物の変動とその変動機序を検証した。第二章では、肝障害と血中アルギニンの関連を調べるため、肝障害物質又は骨格筋障害物質をラットに単回経口投与し、血中アルギニン及びALTの測定と病理組織検査を実施した。また、肝障害時のアルギニンの変動機序を調べるため、血中アルギナーゼ濃度の測定とinvitroでの血中アルギナーゼ活性を調べた。第三章では、軽度の消化管障害時の血中シトルリンの変動を調べるため、消化管障害物質をラットに反復静脈内投与し、血中シトルリンの測定及び病理組織検査を実施した。さらに、消化管障害時の血中シトルリン低下の機序を確認するため、消化管中のシトルリン合成に関与する酵素の遺伝子発現と内因性代謝物の測定を行った。第四章では、これらより得られた知見と最近の文献情報を基に、アルギニンやシトルリンの安全性評価での有用性を総合的に考察した。

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