JP6916776B2 - Ang−2に対する重鎖のみ抗体 - Google Patents
Ang−2に対する重鎖のみ抗体 Download PDFInfo
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Description
本出願は、2015年7月29日に出願された米国仮特許出願第62/198,518号及び2015年8月14日に出願された米国仮特許出願第62/205,185号に対する利益を主張するものであり、それら両方の全記載内容は、参照により本明細書に組み込まれる。
(1)疎水性:ノルロイシン、Met、Ala、Val、Leu、Ile;
(2)中性親水性:Cys、Ser、Thr;
(3)酸性:Asp、Glu;
(4)塩基性:Asn、Gin、His、Lys、Arg;
(5)鎖の配向に影響を与える残基:Gly、Pro;及び
(6)芳香族:Trp、Tyr、Phe。
動物を用いたすべての手順は、Institutional Animal Care and Use Committeeによって作成されたガイドラインに従って実施した。Harbour Antibodies(マサチューセッツ州ケンブリッジ)由来HCAbマウスを使用して抗ANG−2抗体を作製した。
トータルRNAを所望のハイブリドーマから単離し、特定のプライマーを使用してPCR増幅を行った。各ハイブリドーマから単離されたVH領域(図3)をIgG1ヒンジ領域及びCH2/CH3領域と融合させ、GS SYSTEM(商標)(Lonza、スイス、バーゼル)発現プラスミドにHCAb配列を生成させた。発現プラスミドをCHO細胞株にトランスフェクションし、完全ヒト組換えHCAbを作製した。
抗hIgG Fc捕捉(AHC)バイオセンサーチップ及びストレプトアビジンでコートしたバイオセンサー(forteBIO、カリフォルニア州メンローパーク)が装備されたforteBIO OCTET(登録商標)QKeシステムを使用して親和性測定を実施した。AHC親和性測定では、精製抗ANG−2 HCAb A33A8を、そのAHCセンサーチップとの結合能について試験した。チップに20μg/mlの抗ANG−2 A33A8 HCAbを用いて担持させた。担持を300秒進行させたところ、捕捉レベルは1.8〜2nmであった。ANG−2(R&D systems)抗原を濃度20〜500nMまで1xPBSで希釈して結合分析用に調製した。会合が開始し、200秒間監視を行い、その後、Factorタンパク質(Gibco、PBS pH7.2)を含まない1xPBS緩衝液にチップを移し、解離を監視した。その後、バイオレイヤー干渉分析を使用して、抗ANG−2 A33A8 HCAbの結合活性をリアルタイムで監視した。測定した会合定数(「Kassoc」)と解離(「Kdissoc」)定数に基づいて、被験ANG−2の抗原結合親和性を計算した。OCTET(登録商標)データ解析ソフトウェア6.4を使用してデータの処理と解析を行った(forteBIO)(図4)。A33A8はANG−2に1ピコモル未満のKdで結合する。ラットANG−2と交差反応しないが、ウサギANG−2とは交差反応する。さらに、A33A8はANG−1と交差反応する。
1ウェルにつき1ミリリットルあたり2マイクログラムのTie−2−hFc(R&D Systems)で96ウェルマイクロタイタープレートを4℃で一晩コートした。別のプレートに、抗HCAbの系列希釈を、2nMのビオチン化ヒトANG−2(R&D Systems)と共に1時間インキュベートした。100マイクロリットルの抗ANG−2 HCAb−ANG−2混合物をTie−2−hFcでコートしたマイクロタイタープレートに1時間塗布した。その後、検出抗体(抗ビオチンHRP)を1時間加え、化学発光HRP基質でELISAを作成し、ELISAマイクロプレートリーダーで読み取った。データをGraphPad Prism 6を使用して分析した(図12)。
アラニンスキャニングを使用して、修飾を受けると、抗ANG−2 HCAb A33A8の結合親和性が改変される、CDR配列内のアミノ酸位置を同定した。
Claims (9)
- ANG−2に対する抗原結合特異性を有する重鎖のみ抗体(HCAb)であって、配列番号9のアミノ酸配列を有する、前記重鎖のみ抗体。
- ANG−2に対する抗原結合特異性を有するHCAbであって、可変領域(VH)が配列番号25の配列を有する、前記HCAb。
- ANG−2に対する抗原結合特異性を有するHCAbであって、相補性決定領域(CDR)は配列番号12、14、及び16を含む、前記HCAb。
- それを必要とする対象の眼科障害を治療するための医薬組成物であって、請求項1〜3のいずれか一項に記載のVH領域を有するHCAbを含む、前記医薬組成物。
- 前記眼科障害は、萎縮型加齢黄斑変性、滲出型加齢黄斑変性、脈絡膜新生血管(CNV)、嚢胞様黄斑浮腫(CME)、近視性脈絡膜新生血管、血管線条、糖尿病黄斑浮腫(DME)、黄斑の浮腫、網膜静脈閉塞、角膜血管新生異常、結膜翼状片、網膜下浮腫、及び網膜内浮腫からなる群から選択される、請求項4に記載の医薬組成物。
- 前記角膜血管新生異常は、角膜炎、角膜移植、角膜形成術または低酸素症の結果として生じる、請求項5に記載の医薬組成物。
- それを必要とする対象の眼科障害を治療するための医薬品の製造における、請求項1〜3のいずれか一項に記載のVH領域を有するHCAbの使用。
- 前記眼科障害は、萎縮型加齢黄斑変性、滲出型加齢黄斑変性、脈絡膜新生血管(CNV)、嚢胞様黄斑浮腫(CME)、近視性脈絡膜新生血管、血管線条、糖尿病黄斑浮腫(DME)、黄斑の浮腫、網膜静脈閉塞、角膜血管新生異常、結膜翼状片、網膜下浮腫、または網膜内浮腫からなる群から選択される、請求項7に記載の使用。
- 前記角膜血管新生異常は、角膜炎、角膜移植、角膜形成術または低酸素症の結果として生じる、請求項8に記載の使用。
Applications Claiming Priority (5)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US201562198518P | 2015-07-29 | 2015-07-29 | |
US62/198,518 | 2015-07-29 | ||
US201562205185P | 2015-08-14 | 2015-08-14 | |
US62/205,185 | 2015-08-14 | ||
PCT/US2016/044838 WO2017020001A2 (en) | 2015-07-29 | 2016-07-29 | Heavy chain only antibodies to ang-2 |
Publications (3)
Publication Number | Publication Date |
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JP2018527327A JP2018527327A (ja) | 2018-09-20 |
JP2018527327A5 JP2018527327A5 (ja) | 2019-07-18 |
JP6916776B2 true JP6916776B2 (ja) | 2021-08-11 |
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JP2018504124A Active JP6916776B2 (ja) | 2015-07-29 | 2016-07-29 | Ang−2に対する重鎖のみ抗体 |
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US (3) | US10266589B2 (ja) |
EP (2) | EP3328886B1 (ja) |
JP (1) | JP6916776B2 (ja) |
CN (1) | CN107849126B (ja) |
AU (2) | AU2016298398A1 (ja) |
BR (1) | BR112018001762A2 (ja) |
CA (1) | CA2992660A1 (ja) |
DK (1) | DK3328886T3 (ja) |
ES (1) | ES2828694T3 (ja) |
HK (1) | HK1253947A1 (ja) |
HU (1) | HUE051165T2 (ja) |
PL (1) | PL3328886T3 (ja) |
RU (1) | RU2749674C2 (ja) |
WO (1) | WO2017020001A2 (ja) |
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US20220185875A1 (en) * | 2019-03-18 | 2022-06-16 | Jiangsu Hengrui Medicine Co., Ltd. | Bispecific antibody specifically bound to vegf and ang2 |
US20230405121A1 (en) * | 2020-11-04 | 2023-12-21 | Fate Therapeutics, Inc. | Engineered ipsc and immune effector cells for heterogenous tumor control |
CN116003591B (zh) * | 2022-12-12 | 2023-09-19 | 三门峡市眼科医院 | Ang-2抗体及其应用 |
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AU2015345322A1 (en) * | 2014-11-10 | 2017-04-06 | F. Hoffmann-La Roche Ag | Anti-PDGF-B antibodies and methods of use |
EP3328886B1 (en) * | 2015-07-29 | 2020-09-16 | Allergan, Inc. | Heavy chain only antibodies to ang-2 |
JP2018523673A (ja) * | 2015-08-14 | 2018-08-23 | アラーガン、インコーポレイテッドAllergan,Incorporated | Pdgfに対する重鎖のみ抗体 |
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2016
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- 2016-07-29 HU HUE16748025A patent/HUE051165T2/hu unknown
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- 2016-07-29 ES ES16748025T patent/ES2828694T3/es active Active
- 2016-07-29 CN CN201680044324.2A patent/CN107849126B/zh active Active
- 2016-07-29 AU AU2016298398A patent/AU2016298398A1/en not_active Abandoned
- 2016-07-29 RU RU2018103960A patent/RU2749674C2/ru active
- 2016-07-29 BR BR112018001762A patent/BR112018001762A2/pt not_active IP Right Cessation
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2018
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2019
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Also Published As
Publication number | Publication date |
---|---|
HUE051165T2 (hu) | 2021-03-01 |
BR112018001762A2 (pt) | 2018-09-18 |
DK3328886T3 (da) | 2020-11-02 |
US20170107279A1 (en) | 2017-04-20 |
EP3328886B1 (en) | 2020-09-16 |
JP2018527327A (ja) | 2018-09-20 |
CA2992660A1 (en) | 2017-02-02 |
AU2016298398A1 (en) | 2018-02-08 |
EP3328886A2 (en) | 2018-06-06 |
HK1253947A1 (zh) | 2019-07-05 |
RU2018103960A3 (ja) | 2020-01-16 |
RU2018103960A (ru) | 2019-08-28 |
US20210324064A1 (en) | 2021-10-21 |
RU2749674C2 (ru) | 2021-06-16 |
EP3792279A3 (en) | 2021-07-07 |
AU2022259761A1 (en) | 2022-12-01 |
CN107849126B (zh) | 2022-04-08 |
PL3328886T3 (pl) | 2021-02-08 |
US11046756B2 (en) | 2021-06-29 |
CN107849126A (zh) | 2018-03-27 |
EP3792279A2 (en) | 2021-03-17 |
ES2828694T3 (es) | 2021-05-27 |
US10266589B2 (en) | 2019-04-23 |
US20190248882A1 (en) | 2019-08-15 |
WO2017020001A3 (en) | 2017-03-16 |
WO2017020001A2 (en) | 2017-02-02 |
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