JP2017534595A - ヒトp53の直線状エピトープに結合する抗体及びその診断的応用 - Google Patents
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- G01N33/6896—Neurological disorders, e.g. Alzheimer's disease
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- C07K2317/00—Immunoglobulins specific features
- C07K2317/30—Immunoglobulins specific features characterized by aspects of specificity or valency
- C07K2317/34—Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues
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- G01N2333/4748—Details p53
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Abstract
Description
健常で異常のない6/8週齢のマウスを免疫処置に用いた。抗体産生用抗原として用いたペプチドは、以下の特徴を有した。
配列:「N−末端」CRTEEENLRKKGEPHH「C末端」(配列番号2)
長さ:16アミノ酸
分子量:1960.94
純度:96.4%
形状:凍結乾燥粉末
結合:グルタルアルデヒド法によりBSAと結合
動物の脾臓細胞をマウス骨髄腫細胞(SP2/0細胞株)と融合した。融合生成物を、抗原に対してスクリーニングし、抗体産生クローンを選択した。これらのクローンの増殖を継続した。この最初のスクリーニングは、ELISA法により行った。陽性クローンを親クローンとし、3回継代培養後凍結した。ELISAプレート上に抗原コーティングを行い、続いて融合生成物の上清を添加した。免疫動物の血清をELISAにおける陽性対照として用いた(実施例2)。
散発性アルツハイマー(SAD)及び家族性アルツハイマー(FAD)と診断された患者の不死化Bリンパ球において、タンパク質の野生型アイソフォーム(PAb1620)及び立体構造変化アイソフォーム(2D3A8)を認識する2つの立体構造特異的抗体を用い、p53の立体構造状態を免疫沈降法により評価した。その後、ポリクローナル抗p53抗体(CM1)を用いたウェスタンブロットにより、免疫沈降物を可視化した。実験データを、同じ試料の2D3A8抗体陽性バンド及びPAb1620陽性バンドの強度比として表した。
Claims (10)
- 配列がRRTEEENLRKKGEPHH(配列番号1)であるヒトp53の直線状エピトープに結合することを特徴とする抗ヒトp53抗体。
- モノクローナル抗体である、請求項1に記載の抗体。
- 請求項1又は2に記載の抗体を調製する方法であって、
(i)キャリアと任意に結合し、配列がCRTEEENLRKKGEPHH(配列番号2)である前記ヒトp53タンパク質のペプチドを含む免疫原を用い、非ヒト動物に免疫処置する工程であって、前記キャリアは好ましくはウシ血清アルブミンである工程;と
(ii)免疫処置で得られた前記抗体を単離する工程と、
により特徴付けられる方法。 - 少なくとも請求項1又は2に記載の抗体、及び前記抗体のヒトp53タンパク質への結合を検出する手段を含む、イムノアッセイキット。
- 前記野生型ヒトp53タンパク質に対する翻訳後修飾により、立体構造的に変化した試料中の前記ヒトp53タンパク質のアイソフォームを検出するin vitroの方法であって、前記試料が請求項1又は2に記載の少なくとも1つの抗体と接触すること、前記ヒトp53タンパク質及び前記抗体の免疫複合体の形成が、前記試料において検出されることを特徴とする方法。
- 被験者におけるアルツハイマー病を診断するin vitroの方法であって、
(i)前記被験者の生体試料を、請求項1又は2に記載の少なくとも1つの抗ヒトp53抗体と、前記ヒトp53タンパク質及び前記少なくとも1つの抗体の免疫複合体の形成に適当な条件下で接触させる工程;と
(ii)前記ヒトp53タンパク質及び前記少なくとも1つの抗体の免疫複合体の形成を検出する工程と、により特徴付けられ、
前記免疫複合体の形成はアルツハイマー病を示す、方法。 - 軽度認知障害である被験者のアルツハイマー病を発症する疾病素質を決定するin vitroの方法であって、
(i)前記被験者の生体試料を、請求項1又は2に記載の少なくとも1つの抗ヒトp53抗体と、前記ヒトp53タンパク質及び前記少なくとも1つの抗体の免疫複合体の形成に適当な条件下で接触させる工程;と
(ii)前記ヒトp53タンパク質及び前記少なくとも1つの抗体の免疫複合体の形成を検出する工程と、により特徴付けられ、
前記免疫複合体の形成はアルツハイマー病を発症する疾病素質を示す、方法。 - 加齢に伴い認知機能不良となる被験者の疾病素質を決定するin vitroの方法であって、
(i)前記被験者の生体試料を、請求項1又は2に記載の少なくとも1つの抗ヒトp53抗体と、前記ヒトp53タンパク質及び前記少なくとも1つの抗体の免疫複合体の形成に適当な条件下で接触させる工程;と
(ii)前記ヒトp53タンパク質及び前記少なくとも1つの抗体の免疫複合体の形成を検出する工程と、により特徴付けられ、
前記免疫複合体の形成は加齢に伴い認知機能不良となる疾病素質を示す、方法。 - 前記生体試料が、血液、血漿、血清、唾液、尿、神経細胞、血液細胞又は他の細胞型である、請求項5〜8のいずれかに記載のin vitroの方法。
- 前記検出が、免疫沈降反応技術、イムノアッセイ、好ましくはELISAもしくはRIAもしくは免疫蛍光アッセイ、ウェスタンブロット、FACS分析、又は免疫細胞化学/免疫組織化学技術により実施される、請求項5〜9のいずれかに記載のin vitroの方法。
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
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ITTO20140776 | 2014-09-30 | ||
ITTO2014A000776 | 2014-09-30 | ||
PCT/EP2015/072094 WO2016050630A1 (en) | 2014-09-30 | 2015-09-25 | Antibody binding a linear epitope of human p53 and diagnostic applications thereof |
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JP2020136359A Division JP2020203895A (ja) | 2014-09-30 | 2020-08-12 | ヒトp53の直線状エピトープに結合する抗体及びその診断的応用 |
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JP2017534595A true JP2017534595A (ja) | 2017-11-24 |
JP6938372B2 JP6938372B2 (ja) | 2021-09-22 |
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JP2020136359A Pending JP2020203895A (ja) | 2014-09-30 | 2020-08-12 | ヒトp53の直線状エピトープに結合する抗体及びその診断的応用 |
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CN (1) | CN107001453B (ja) |
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Cited By (1)
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JP2022531058A (ja) * | 2019-03-06 | 2022-07-06 | ディアデム エス.アール.エル. | アルツハイマー病の診断と予後判定におけるマーカーとしてのp53ペプチド |
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HUE042815T2 (hu) | 2014-09-30 | 2019-07-29 | Diadem S R L | Humán P53 lineáris epitópot kötõ antitest és diagnosztikai alkalmazásai |
KR101885007B1 (ko) * | 2016-12-22 | 2018-08-31 | 가천대학교 산학협력단 | 인지기능장애의 진단 방법 및 키트 |
IT202000018544A1 (it) | 2020-07-30 | 2022-01-30 | Diadem S R L | Modificazioni post-traslazionali su p-53 quali marcatori nella diagnosi e prognosi di malattia neurodegenerativa |
WO2024030982A2 (en) | 2022-08-02 | 2024-02-08 | Diadem Spa | Device for detection and prognostic assessment of neurodegenerative disorders |
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HUE042815T2 (hu) | 2014-09-30 | 2019-07-29 | Diadem S R L | Humán P53 lineáris epitópot kötõ antitest és diagnosztikai alkalmazásai |
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JP2022531058A (ja) * | 2019-03-06 | 2022-07-06 | ディアデム エス.アール.エル. | アルツハイマー病の診断と予後判定におけるマーカーとしてのp53ペプチド |
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LT3201234T (lt) | 2019-03-12 |
RU2017114626A3 (ja) | 2019-04-26 |
SI3201234T1 (sl) | 2019-03-29 |
US11208473B2 (en) | 2021-12-28 |
US10183990B2 (en) | 2019-01-22 |
TR201900550T4 (tr) | 2019-02-21 |
RS58244B1 (sr) | 2019-03-29 |
CN107001453A (zh) | 2017-08-01 |
DK3201234T3 (en) | 2019-02-25 |
JP2020203895A (ja) | 2020-12-24 |
JP6938372B2 (ja) | 2021-09-22 |
EP3201234B1 (en) | 2018-11-07 |
US10875908B2 (en) | 2020-12-29 |
WO2016050630A1 (en) | 2016-04-07 |
CN107001453B (zh) | 2021-07-20 |
US20210070848A1 (en) | 2021-03-11 |
US11203635B2 (en) | 2021-12-21 |
US20210061895A1 (en) | 2021-03-04 |
CY1121169T1 (el) | 2020-05-29 |
PL3201234T3 (pl) | 2019-04-30 |
RU2741795C2 (ru) | 2021-01-28 |
HUE042815T2 (hu) | 2019-07-29 |
PT3201234T (pt) | 2019-02-05 |
US11802148B2 (en) | 2023-10-31 |
RU2017114626A (ru) | 2018-11-02 |
US20240124565A1 (en) | 2024-04-18 |
HRP20190017T1 (hr) | 2019-03-22 |
US20190194305A1 (en) | 2019-06-27 |
EP3201234A1 (en) | 2017-08-09 |
ES2708556T3 (es) | 2019-04-10 |
US20220064273A1 (en) | 2022-03-03 |
US20180057572A1 (en) | 2018-03-01 |
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