RU2013103322A - КРИСТАЛЛИЧЕСКИЕ АНТИТЕЛА ПРОТИВ hTNFα - Google Patents

КРИСТАЛЛИЧЕСКИЕ АНТИТЕЛА ПРОТИВ hTNFα Download PDF

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RU2013103322A
RU2013103322A RU2013103322/10A RU2013103322A RU2013103322A RU 2013103322 A RU2013103322 A RU 2013103322A RU 2013103322/10 A RU2013103322/10 A RU 2013103322/10A RU 2013103322 A RU2013103322 A RU 2013103322A RU 2013103322 A RU2013103322 A RU 2013103322A
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crystallization
antibody
crystallization method
mixture
crystal
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Дэвид В. БОРХАНИ
Вольфганг ФРАУНХОФЕР
Ханс-Юрген КРАУЗЕ
Анетте КЕНИГСДОРФЕР
Герхард ВИНТЕР
Штефан ГОТТШАЛЬК
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Эббви Байотекнолоджи Лтд.,Вм
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    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/24—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against cytokines, lymphokines or interferons
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Abstract

1. Способ порционной кристаллизации для кристаллизования человеческого антитела против hTNFα, включающий стадии:(a) предоставления водного раствора указанного антитела в смеси с солью неорганического фосфата в качестве кристаллизационного агента; и(b) инкубации указанной водной кристаллизационной смеси до тех пор, пока не образуются кристаллы указанного антитела.2. Способ кристаллизации по п.1, где рН указанной водной кристаллизационной смеси находится в диапазоне примерно от рН 3 до 5.3. Способ кристаллизации по п.1 или 2, где указанная водная кристаллизационная смесь содержит буфер.4. Способ кристаллизации по п.3, где указанный буфер содержит ацетатный буфер.5. Способ кристаллизации по п.4, где буфер содержит ацетат натрия.6. Способ кристаллизации по п.3, где концентрация буфера в указанной водной кристаллизационной смеси составляет от 0М до 0,5М.7. Способ кристаллизации по п.1, где фосфатная соль представляет собой гидрофосфатную соль.8. Способ кристаллизации по п.7, где фосфатная соль представляет собой соль щелочного металла или смесь, по меньшей мере, двух различных солей щелочных металлов.9. Способ кристаллизации по п.1, где концентрация фосфатной соли в кристаллизационной смеси находится в диапазоне примерно от 1 до 6M.10. Способ кристаллизации по п.9, где концентрация соли в кристаллизационной смеси находится в диапазоне примерно от 1,0 до 3,0М.11. Способ кристаллизации по п.1, где соблюдено, по меньшей мере, одно из следующих дополнительных условий кристаллизации:a) инкубацию осуществляют в течение примерно от 1 часа до примерно 60 дней;b) инкубацию осуществляют при температуре примерно от 4°С до примерно 37°С;c) концентрация а�

Claims (25)

1. Способ порционной кристаллизации для кристаллизования человеческого антитела против hTNFα, включающий стадии:
(a) предоставления водного раствора указанного антитела в смеси с солью неорганического фосфата в качестве кристаллизационного агента; и
(b) инкубации указанной водной кристаллизационной смеси до тех пор, пока не образуются кристаллы указанного антитела.
2. Способ кристаллизации по п.1, где рН указанной водной кристаллизационной смеси находится в диапазоне примерно от рН 3 до 5.
3. Способ кристаллизации по п.1 или 2, где указанная водная кристаллизационная смесь содержит буфер.
4. Способ кристаллизации по п.3, где указанный буфер содержит ацетатный буфер.
5. Способ кристаллизации по п.4, где буфер содержит ацетат натрия.
6. Способ кристаллизации по п.3, где концентрация буфера в указанной водной кристаллизационной смеси составляет от 0М до 0,5М.
7. Способ кристаллизации по п.1, где фосфатная соль представляет собой гидрофосфатную соль.
8. Способ кристаллизации по п.7, где фосфатная соль представляет собой соль щелочного металла или смесь, по меньшей мере, двух различных солей щелочных металлов.
9. Способ кристаллизации по п.1, где концентрация фосфатной соли в кристаллизационной смеси находится в диапазоне примерно от 1 до 6M.
10. Способ кристаллизации по п.9, где концентрация соли в кристаллизационной смеси находится в диапазоне примерно от 1,0 до 3,0М.
11. Способ кристаллизации по п.1, где соблюдено, по меньшей мере, одно из следующих дополнительных условий кристаллизации:
a) инкубацию осуществляют в течение примерно от 1 часа до примерно 60 дней;
b) инкубацию осуществляют при температуре примерно от 4°С до примерно 37°С;
c) концентрация антитела находится в диапазоне примерно от 0,5 до примерно 100 мг/мл.
12. Способ кристаллизации по п.1, дополнительно включающий стадию сушки указанных кристаллов.
13. Способ кристаллизации по п.1, дополнительно включающий стадию замены кристаллизационного маточного раствора другим буфером.
14. Способ кристаллизации по п.1, где объем порции находится в диапазоне примерно от 1 мл до 20,000 л.
15. Кристалл антитела против hTNFα, который можно получить при помощи способа кристаллизации, как определено в любом из предшествующих пунктов.
16. Кристалл антитела против hTNFα, при условии, что указанное антитело не является INFLIXIMAB.
17. Кристалл по п.15 или 16, обладающий игольчатоподобной морфологией с максимальной длиной 1 примерно 2-500 мкм и соотношением 1/d примерно от 3 до 30.
18. Кристалл по п.15 или 16, где указанное антитело представляет собой поликлональное антитело или моноклональное антитело.
19. Кристалл по п.18, где указанное антитело выбрано из группы, состоящей из химерных антител, гуманизированных антител, негликозилорованных антител, человеческих антител и мышиных антител.
20. Кристалл по п.17, где указанное антитело представляет собой антитело IgG.
21. Кристалл по п.19, где указанное антитело выбирают из группы, состоящей из: антител IgG1, IgG2, IgG3 и IgG4.
22. Кристалл по п.21, где указанное антитело представляет собой антитело против TNFα человека группы IgG1.
23. Кристалл по п.22, полученный из выделенного антитела человека, которое диссоциирует от hTNFα с Kd 1∙10-8M или меньше и константой скорости Ka 1·10-3 с-1 или меньше, каждая из которых определена при помощи поверхностного плазменного резонанса, и нейтрализует цитотоксичность hTNFα в стандартном анализе in vitro L929 с IC50 1∙10-7 M или меньше.
24. Кристалл по п.22, полученный из выделенного антитела человека со следующими характеристиками: а) диссоциирует от TNFα человека с константой скорости Koff 1·10-3 с-1 или меньше, как определено при помощи поверхностного плазменного резонанса; b) обладает доменом легкой цепи CDR3, включающим аминокислотную последовательность SEQ ID NO:3 или последовательность, модифицированную из SEQ ID NO:3 путем одной замены аланина в положении 1, 4, 5, 7 или 8 или путем одной-пяти консервативных аминокислотных замен в положениях 1, 3, 4, 6, 7, 8 и/или 9; с) обладает доменом тяжелой цепи CDR3, включающим аминокислотную последовательность SEQ ID NO:4 или последовательность, модифицированную из SEQ ID NO:4 путем одной замены аланина в положении 2, 3, 4, 5, 6, 8, 9, 10 или 11 или путем одной-пяти консервативных аминокислотных замен в положениях 2, 3, 4, 5, 6, 8, 9, 10, 11 и/или 12.
25. Кристалл по п.22, полученный из выделенного антитела человека с вариабельной областью легкой цепи (LCVR), включающей аминокислотную последовательность SEQ ID NO:1, и вариабельной областью тяжелой цепи (HCVR), включающей аминокислотную последовательность SEQ ID NO:2.
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