JP2017538415A - Axlに対するヒトモノクローナル抗体 - Google Patents
Axlに対するヒトモノクローナル抗体 Download PDFInfo
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- JP2017538415A JP2017538415A JP2017530761A JP2017530761A JP2017538415A JP 2017538415 A JP2017538415 A JP 2017538415A JP 2017530761 A JP2017530761 A JP 2017530761A JP 2017530761 A JP2017530761 A JP 2017530761A JP 2017538415 A JP2017538415 A JP 2017538415A
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Abstract
Description
本発明は、AXLに対するヒトモノクローナル抗体及び抗体−薬剤コンジュゲートを提供する。特に、本発明の抗体は、完全にヒト抗体であり、ヒトAXLに対して高い親和性で結合し、マウス型とヒト型とのAXL間で交叉反応することができ、レセプターの内部移行及び分解を誘引し、このレセプターのホモ二量体化を支持することができ、AXLのリン酸化を阻害し、in vitroにおける細胞増殖を阻害し、抵抗性AXL過剰発現CML細胞及びイマチニブ抵抗性GIST細胞のニロチニブに対する感受性を回復させ、in nivoにおける腫瘍成長を低下させるような、1つ以上の機能性により特徴付けられる。特に、本発明は、実施例に記載されたD4抗体から得られた抗体を提供する。
D4のL−CDR2は、配列番号:2における、52位のアミノ酸残基から58位のアミノ酸残基の範囲の配列により定義される。
シクロアルケニル会合残基F、H、W、及びY
疎水性残基A、C、F、G、H、I、L、M、R、T、V、W、及びY
負に荷電した残基D及びE
極性残基C、D、E、H、K、N、Q、R、S、及びT
正に荷電した残基H、K、及びR
小型残基A、C、D、G、N、P、S、T、及びV
非常に小型の残基A、G、及びS
ターン形成に関わる残基A、C、D、E、G、H、K、N、Q、R、S、P、及びT
フレキシブルな残基Q、T、K、S、G、P、D、E、及びR
−F(ab’)2フラグメント(同フラグメントは、ヒンジ領域においてジスルフィド架橋により連結された、2つのFabフラグメントを含む二価のフラグメントである。これらは、例えば、全長抗体をペプシンで処理することにより生成することができる)、
−Fab’又はFabフラグメント(同フラグメントは、VL、VH、CL、及びCH1ドメインからなる一価のフラグメントである。Fabフラグメントは、例えば、IgG抗体をパパインで処理することにより得ることができる。Fab’フラグメントは、例えば、F(ab’)2フラグメントのジスルフィド架橋を、還元剤、例えば、ジチオスレイトールを使用して還元することにより得ることができる)、
−Fdフラグメント(同フラグメントは、VH及びCH1ドメインから本質的になる)、
−Fvフラグメント(同フラグメントは、抗体の1つのアームのVL及びVHドメインとその一本鎖抗体とから本質的になる。一本鎖抗体(一本鎖Fv(scFv)抗体としても公知)は、FvフラグメントのVL及びVHドメインが、リコンビナント法を使用して、これらのドメインを1つのタンパク質鎖として発現させることができる合成リンカーにより連結されている構築物である。同構築物中において、VL及びVH領域ペアは、一価の分子を形成する(例えば、Bird et al., Science 242, 423-426 (1988)及びHuston et al., PNAS USA 85, 5879-5883 (1988)を参照のこと))、
−配列番号:1又は配列番号:2のVL又はVH鎖ならびに配列番号:1又は配列番号:2に対して少なくとも70%の同一性を有するアミノ酸配列を含むか、又は、これらからなるフラグメント
本発明者らは、AXLレセプターの細胞外ドメインに特異的な完全なヒトmAbを、研究室において生成し、細胞膜表面上でAXLレセプターの内部移行及び分解を誘引可能であり(図1)、マウス型とヒト型とのAXL間で交叉反応可能(図6)なもの(すなわち、D4ヒトモノクローナル抗体)をスクリーニングしてきた。この抗体は、このレセプターのホモ二量体化を誘引可能である(図2)。その結果として、このmAbで処理した腫瘍細胞は、その細胞表面におけるAXLレセプターの存在を劇的に減少させるのを示した。処理された細胞は、GAS6(AXLの天然リガンド)に対して応答できない。これにより、AXLリン酸化の阻害及びin vitroにおける細胞増殖の強力な阻害ももたらされた。本発明者らは、本発明者らの抗AXL mAbによる処理により、抵抗性AXL過剰発現CML細胞のニロチニブに対する感受性が回復されることを証明した(図3)。さらに、同じ効果は、処理されたイマチニブ抵抗性GIST細胞において観察された(図4)。
全てのin vivoにおける実験を、French guidelines for experimental animal studies(協定番号C34-172-27)に遵守して行った。6週齢のメスの無胸腺ヌードマウスを、Harlanから購入した。膵臓ガン細胞(3.5×106個、BXPC3)を、無胸腺ヌードマウスの右腹部に移植した。担ガンマウスを、腫瘍が体積約100mm3に達した時点で、種々の処置群にランダム化した。マウスを、媒体(0.9% NaCl);マウス抗AXLモノクローナル抗体20G7−D9(D9)、又は、注入あたりに300μg ヒトモノクローナル抗体D4単独(4週間連続で1週間に2回)の腹腔内注入により処置した。腫瘍体積を、カリパーにより、1週間に1回測定した。抗AXL抗体、及び特に、ヒト抗体D4は、膵臓異種移植片の腫瘍成長を低下させる(図5)。
この出願全体を通して、種々の参考文献が、本発明が属する技術水準を説明している。これらの参考文献の開示は、参照により、本開示に組み入れられる。
Claims (20)
- i)D4のH−CDR1に対して少なくとも90%の同一性を有するH−CDR1、ii)D4のH−CDR2に対して少なくとも90%の同一性を有するH−CDR2、及びiii)D4のH−CDR3に対して少なくとも90%の同一性を有するH−CDR3を含む重鎖、又は、i)D4のL−CDR1に対して少なくとも90%の同一性を有するL−CDR1、ii)D4のL−CDR2に対して少なくとも90%の同一性を有するL−CDR2、及びiii)D4のL−CDR3に対して少なくとも90%の同一性を有するL−CDR3を含む軽鎖を含み、そしてここで、
−D4のH−CDR1は、配列番号:1における、31位のアミノ酸残基から35位のアミノ酸残基の範囲の配列により定義され、
−D4のH−CDR2は、配列番号:1における、50位のアミノ酸残基から66位のアミノ酸残基の範囲の配列により定義され、
−D4のH−CDR3は、配列番号:1における、99位のアミノ酸残基から108位のアミノ酸残基の範囲の配列により定義され、
−D4のL−CDR1は、配列番号:2における、23位のアミノ酸残基から36位のアミノ酸残基の範囲の配列により定義され、
−D4のL−CDR2は、配列番号:2における、52位のアミノ酸残基から58位のアミノ酸残基の範囲の配列により定義され、
−D4のL−CDR3は、配列番号:2における、91位のアミノ酸残基から100位のアミノ酸残基の範囲の配列により定義される、
AXLに対するヒトモノクローナル抗体。 - i)D4のH−CDR1に対して少なくとも90%の同一性を有するH−CDR1、ii)D4のH−CDR2に対して少なくとも90%の同一性を有するH−CDR2、及びiii)D4のH−CDR3に対して少なくとも90%の同一性を有するH−CDR3を含む重鎖と、i)D4のL−CDR1に対して少なくとも90%の同一性を有するL−CDR1、ii)D4のL−CDR2に対して少なくとも90%の同一性を有するL−CDR2、及びiii)D4のL−CDR3に対して少なくとも90%の同一性を有するL−CDR3を含む軽鎖とを含む、請求項1記載のヒトモノクローナル抗体。
- i)D4のH−CDR1、ii)D4のH−CDR2、及びiii)D4のH−CDR3を有する重鎖を含む、請求項1記載のヒトモノクローナル抗体。
- i)D4のL−CDR1、ii)D4のL−CDR2、及びiii)D4のL−CDR3を有する軽鎖を含む、請求項1記載のヒトモノクローナル抗体。
- i)D4のH−CDR1、ii)D4のH−CDR2、及びiii)D4のH−CDR3を有する重鎖と、i)D4のL−CDR1、ii)D4のL−CDR2、及びiii)D4のL−CDR3を有する軽鎖とを含む、請求項1記載のヒトモノクローナル抗体。
- 配列番号:1に対して少なくとも70%の同一性を有する重鎖を含む抗体である、請求項1記載のヒトモノクローナル抗体。
- 配列番号:2に対して少なくとも70%の同一性を有する軽鎖を含む抗体である、請求項1記載のヒトモノクローナル抗体。
- 配列番号:1に対して少なくとも70%の同一性を有する重鎖と、配列番号:2に対して少なくとも70%の同一性を有する軽鎖とを含む抗体である、請求項1記載のヒトモノクローナル抗体。
- 配列番号:1に同一の重鎖を含む抗体である、請求項1記載のヒトモノクローナル抗体。
- 配列番号:2に同一の軽鎖を含む抗体である、請求項1記載のヒトモノクローナル抗体。
- 配列番号:1に同一の重鎖と、配列番号:2に同一の軽鎖とを含む抗体である、請求項1記載のヒトモノクローナル抗体。
- Fv、Fab、F(ab’)2、Fab’、dsFv、scFv、sc(Fv)2、及びディアボディからなる群より選択される、請求項1記載のヒトモノクローナル抗体のフラグメント。
- 請求項1記載のヒト抗体の重鎖及び/又は軽鎖をコードする、
核酸分子。 - 請求項13記載の核酸を含む、
ベクター。 - 請求項13記載の核酸又は請求項14記載のベクターを含む、
ホスト細胞。 - サイトトキシン、化学療法剤、サイトカイン、免疫抑制剤、免疫刺激剤、溶解型ペプチド、及び放射性同位体からなる群より選択される治療部分にコンジュゲートしている、請求項1記載のヒトモノクローナル抗体。
- オーリスタチンにコンジュゲートしている、請求項16記載のヒトモノクローナル抗体。
- 医薬として使用するための、請求項1又は16記載のヒトモノクローナル抗体。
- ガンの処置を必要する対象においてガンを処置する方法であって、
治療的に有効な量の請求項1又は16記載のヒトモノクローナル抗体を、対象に投与することを含む、
方法。 - 請求項1又は16記載のヒトモノクローナル抗体を含む、
医薬組成物。
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US10398774B2 (en) | 2019-09-03 |
ES2764299T3 (es) | 2020-06-02 |
EP3229836B1 (en) | 2019-11-13 |
EP3229836A1 (en) | 2017-10-18 |
JP6864953B2 (ja) | 2021-04-28 |
WO2016091891A1 (en) | 2016-06-16 |
US20170340734A1 (en) | 2017-11-30 |
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