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RU2012126138A - Специфические связывающие агенты против в7-н1 - Google Patents

Специфические связывающие агенты против в7-н1 Download PDF

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RU2012126138A
RU2012126138A RU2012126138/10A RU2012126138A RU2012126138A RU 2012126138 A RU2012126138 A RU 2012126138A RU 2012126138/10 A RU2012126138/10 A RU 2012126138/10A RU 2012126138 A RU2012126138 A RU 2012126138A RU 2012126138 A RU2012126138 A RU 2012126138A
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antibody
amino acid
acid sequence
fragment
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Кристоф КУЕВА
Майкл МОРРОУ
Скотт ХЭММОНД
Марат АЛИМЖАНОВ
Джон БЭБКУК
Ян Фолтц
Джаспал Сингх КАНГ
Лаура СЕКИРОВ
Мелани Бойл
Маттье ЧОДОРЖ
Росс СТЮАРТ
Кэтлин Энн МАЛГРУ
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Abstract

1. Выделенное антитело или его фрагмент, которое специфически связывается с В7-Н1, и антитело проявляет одно или несколько из следующих свойств, выбранных из группы, состоящей из:связывания человеческого В7-Н1 с Kменее 2 нМ, при определении методом BIAcore;перекрестного реагирования с В7-Н1 яванского макака;связывания В7-Н1 яванского макака с Kменее 2 нМ, при определении методом BIAcore;проявления активации CD4+ Т-клеток в анализе смешанных лимфоцитарных дендритных клеток-Т-клеток;ингибирования связывания человеческого В7-Н1 с PD-1, экспрессируемым на клетках ES-2, с ICменее 0,2 нМ;ингибирования связывания человеческого В7-Н1 с В7-1 при проведении анализа гомогенной TR-FRET с ICменее 0,1 нМ; иингибирования роста опухоли раковой клеточной линии в модели ксенотрансплантата по сравнению с изотипическим контрольным антителом.2. Антитело или его фрагмент по п.1, отличающееся тем, что ингибирует рост опухоли клеточной линии человеческого рака поджелудочной железы (НРАС) в модели ксенотрансплантата в день 30 более чем на 40% по сравнению с изотипическим контролем, при определении методом измерения объема опухолей.3. Антитело или его фрагмент по п.1, отличающееся тем, что ингибирует рост опухоли клеточной линии A375 (меланома) в модели ксенотрансплантата в день 30 более чем на 50% по сравнению с изотипическим контролем, при определении методом измерения объема опухолей.4. Антитело или его фрагмент по п.1, связывающееся с тем же эпитопом на внеклеточном домене человеческого В7-Н1, что и любое из антител 2.7А4, 2.14Н9 или 2.9D10, 2.7А4ОРТ или 2.14Н9ОРТ.5. Антитело или его фрагмент по п.4, отличающееся тем, что связывает эпитоп на внеклеточном домене человеческого В7-Н1, включаю

Claims (41)

1. Выделенное антитело или его фрагмент, которое специфически связывается с В7-Н1, и антитело проявляет одно или несколько из следующих свойств, выбранных из группы, состоящей из:
связывания человеческого В7-Н1 с KD менее 2 нМ, при определении методом BIAcore;
перекрестного реагирования с В7-Н1 яванского макака;
связывания В7-Н1 яванского макака с KD менее 2 нМ, при определении методом BIAcore;
проявления активации CD4+ Т-клеток в анализе смешанных лимфоцитарных дендритных клеток-Т-клеток;
ингибирования связывания человеческого В7-Н1 с PD-1, экспрессируемым на клетках ES-2, с IC50 менее 0,2 нМ;
ингибирования связывания человеческого В7-Н1 с В7-1 при проведении анализа гомогенной TR-FRET с IC50 менее 0,1 нМ; и
ингибирования роста опухоли раковой клеточной линии в модели ксенотрансплантата по сравнению с изотипическим контрольным антителом.
2. Антитело или его фрагмент по п.1, отличающееся тем, что ингибирует рост опухоли клеточной линии человеческого рака поджелудочной железы (НРАС) в модели ксенотрансплантата в день 30 более чем на 40% по сравнению с изотипическим контролем, при определении методом измерения объема опухолей.
3. Антитело или его фрагмент по п.1, отличающееся тем, что ингибирует рост опухоли клеточной линии A375 (меланома) в модели ксенотрансплантата в день 30 более чем на 50% по сравнению с изотипическим контролем, при определении методом измерения объема опухолей.
4. Антитело или его фрагмент по п.1, связывающееся с тем же эпитопом на внеклеточном домене человеческого В7-Н1, что и любое из антител 2.7А4, 2.14Н9 или 2.9D10, 2.7А4ОРТ или 2.14Н9ОРТ.
5. Антитело или его фрагмент по п.4, отличающееся тем, что связывает эпитоп на внеклеточном домене человеческого В7-Н1, включающий по меньшей мере два из следующих трех аминокислотных остатков - Asp в положении 122, Arg в положении 125 или Arg в положении 113.
6. Антитело или его фрагмент по п.5, отличающееся тем, что не связывается с эпитопом, содержащим Не в положении 54, Ser в положении 117 и Ala в положении 121, на человеческом В7-Н1, при определении способом конкурентного анализа.
7. Антитело или его фрагмент по п.5, отличающееся тем, что теряет свою способность связываться с человеческим В7-Н1, если Arg в положении 113 на указанном человеческом В7-Н1 мутирует на Ala, или на Tyr, или на Leu, при определении способом конкурентного анализа по сравнению со связыванием с В7-Н1 дикого типа.
8. Антитело или его фрагмент по п.5, отличающееся тем, что теряет свою способность связываться с человеческим В7-Н1, если Arg в положении 125 на указанном человеческом В7-Н1 мутирует на Ala, или на Gln, или на Ser, при определении способом конкурентного анализа по сравнению со связыванием с В7-Н1 дикого типа.
9. Антитело или его фрагмент по п.5, отличающееся тем, что охраняет свою способность связываться с человеческим В7-Н1, если Arg в положении 123 на указанном человеческом В7-Н1 мутирует на Ala, или на Phe, или на Thr, при определении методом конкурентного анализа по сравнению со связыванием с В7-Н1 дикого типа.
10. Антитело или его фрагмент по п.4, отличающееся тем, что связывает по меньшей мере два из следующих трех аминокислотных остатков - Phe в положении 19, Thr в положении 20 или Asp в положении 122, на человеческом В7-Н1.
11. Антитело или его фрагмент по п.10, отличающееся тем, что не связывается с эпитопом, содержащим Не в положении 54, Met в положении 115, Ser в положении 117 и Ala в положении 121, на человеческом В7-Н1, при определении методом конкурентного анализа по сравнению со связыванием с В7-Н1 дикого типа.
12. Антитело или его фрагмент по п.10, отличающееся тем, что теряет свою способность связываться с человеческим В7-Н1, если Phe в положении 19 на указанном человеческом В7-Н1 мутирует на Ala, или на Gly, или на Ser, при определении способом конкурентного анализа по сравнению со связыванием с В7-Н1 дикого типа.
13. Антитело или его фрагмент по п.10, отличающееся тем, что теряет свою способность связываться с человеческим В7-Н1, если Thr в положении 20 на указанном человеческом В7-Н1 мутирует на Ala, или на Val, или на Asp, при определении способом конкурентного анализа по сравнению со связыванием с В7-Н1 дикого типа.
14. Антитело или его фрагмент по п.10, отличающееся тем, что теряет свою способность связываться с человеческим В7-Н1, если Asp в положении 122 на указанном человеческом В7-Н1 мутирует на Asn, или на Glu, при определении способом конкурентного анализа по сравнению со связыванием с В7-Н1 дикого типа.
15. Антитело или его фрагмент по п.10, отличающееся тем, что сохраняет свою способность связываться с человеческим В7-Н1, если Arg в положении 123 на указанном человеческом В7-Н1 мутирует на Ala, или на Phe, или на Thr, при определении способом конкурентного анализа по сравнению со связыванием с В7-Н1 дикого типа.
16. Антитело или его фрагмент по п.1, отличающееся тем, что связывает человеческий В7-Н1 с KD менее 1,0 нМ, при определении методом BIAcore.
17. Антитело или его фрагмент по п.16, отличающееся тем, что связывает человеческий В7-Н1 с KD менее 200 пМ, при определении методом BIAcore.
18. Антитело или его фрагмент по п.1, отличающееся тем, что представляет собой моноклональное антитело.
19. Антитело или его фрагмент по п.18, отличающееся тем, что представляет собой полностью человеческое моноклональное антитело.
20. Антитело или его фрагмент по п.1, где антитело является любым из 2.9D10, 2.7А4, 2.14Н9, 2.7А4ОРТ или 2.14Н9ОРТ.
21. Антитело или его фрагмент по п.20, где фрагмент представляет собой связывающий фрагмент, выбранный из группы, состоящей из Fab, Fab', F(ab')2, Fv и фрагмента dAb.
22. Антитело или его фрагмент по п.20, отличающееся тем, что имеет аминокислотную последовательность, содержащую:
аминокислотную последовательность вариабельного домена тяжелой цепи, кодируемую полинуклеотидом в плазмиде, обозначенной 2.7A4_G, которая была депонирована в NCIMB под депозитарным номером 41598, и аминокислотную последовательность вариабельного домена легкой цепи, кодируемую полинуклеотидом в плазмиде, обозначенной 2.7A4_G, которая была депонирована в NCIMB под депозитарным номером 41598; или
аминокислотную последовательность вариабельного домена тяжелой цепи, кодируемую полинуклеотидом в плазмиде, обозначенной 2.14H9_G, которая была депонирована в NCIMB под депозитарным номером 41597, и аминокислотную последовательность вариабельного домена легкой цепи, кодируемую полинуклеотидом в плазмиде, обозначенной 2.14H9_G, которая была депонирована в NCIMB под депозитарным номером 41597; или
аминокислотную последовательность вариабельного домена тяжелой цепи, кодируемую полинуклеотидом в плазмиде, обозначенной 2.9D10_NG, которая была депонирована в NCIMB под депозитарным номером 41599, и аминокислотную последовательность вариабельного домена легкой цепи, кодируемую полинуклеотидом в плазмиде, обозначенной 2.9D10_NG, которая была депонирована в NCIMB под депозитарным номером 41599.
23. Антитело или его фрагмент по п.20, отличающееся тем, что имеет аминокислотную последовательность, содержащую:
VH CDR1, имеющий аминокислотную последовательность SEQ ID NO:3; и
VH CDR2, имеющий аминокислотную последовательность SEQ ID NO:4; и
VH CDR3, имеющий аминокислотную последовательность SEQ ID NO:5; и
VL CDR1, имеющий аминокислотную последовательность SEQ ID NO:8; и
VL CDR2, имеющий аминокислотную последовательность SEQ ID NO:9; и
VL CDR3, имеющий аминокислотную последовательность SEQ ID NO:10; или
VH CDR1, имеющий аминокислотную последовательность SEQ ID NO:23; и
VH CDR2, имеющий аминокислотную последовательность SEQ ID NO:24; и
VH CDR3, имеющий аминокислотную последовательность SEQ ID NO:25; и
VL CDR1, имеющий аминокислотную последовательность SEQ ID NO:28; и
VL CDR2, имеющий аминокислотную последовательность SEQ ID NO:29; и
VL CDR3, имеющий аминокислотную последовательность SEQ ID NO:30; или
VH CDR1, имеющий аминокислотную последовательность SEQ ID NO:13; и
VH CDR2, имеющий аминокислотную последовательность SEQ ID NO:14; и
VH CDR3, имеющий аминокислотную последовательность SEQ ID NO:15; и
VL CDR1, имеющий аминокислотную последовательность SEQ ID NO:18; и
VL CDR2, имеющий аминокислотную последовательность SEQ ID NO:19; и
VL CDR3, имеющий аминокислотную последовательность SEQ ID NO:20; или
VH CDR1, имеющий аминокислотную последовательность SEQ ID NO:63; и
VH CDR2, имеющий аминокислотную последовательность SEQ ID NO:64; и
VH CDR3, имеющий аминокислотную последовательность SEQ ID NO:65; и
VL CDR1, имеющий аминокислотную последовательность SEQ ID NO:68; и
VL CDR2, имеющий аминокислотную последовательность SEQ ID NO:69; и
VL CDR3, имеющий аминокислотную последовательность SEQ ID NO:70; или
VH CDR1, имеющий аминокислотную последовательность SEQ ID NO:73; и
VH CDR2, имеющий аминокислотную последовательность SEQ ID NO:74; и
VH CDR3, имеющий аминокислотную последовательность SEQ ID NO:75; и
VL CDR1, имеющий аминокислотную последовательность SEQ ID NO:78; и
VL CDR2, имеющий аминокислотную последовательность SEQ ID NO:79; и
VL CDR3, имеющий аминокислотную последовательность SEQ ID NO:80.
24. Антитело или его фрагмент по п.22, отличающееся тем, что дополнительно включает вариант Fc, в котором Fc-участок содержит, по меньшей мере, одну неприродную аминокислоту, выбранную из группы, состоящей из 234F, 235F и 331S, пронумерованных в соответствии с индексом EU, как описано Кабат.
25. Молекула нуклеиновой кислоты, кодирующая антитело по п.24.
26. Клетка-хозяин, трансфицированная вектором, содержащим молекулу нуклеиновой кислоты по п.25.
27. Антитело, продуцируемое по способу, включающему культивацию указанной клетки-хозяина по п.26.
28. Композиция, содержащая антитело по п.20.
29. Фармацевтическая композиция, включающая антитело по п.20 и фармацевтически приемлемый носитель.
30. Способ репрессии В7-Н1-медиируемого ингибирования Т-клеток у животного, включающий введение животному, нуждающемуся в этом, эффективного количества композиции по п.28.
31. Способ лечения злокачественной опухоли у животного, включающий: выбор животного, нуждающегося в лечении злокачественной опухоли; и введение указанному животному терапевтически эффективного количества композиции по п.29.
32. Способ по п.31, отличающийся тем, что указанную злокачественную опухоль выбирают из группы, состоящей из: меланомы, немелкоклеточного рака легкого, гепатоцеллюлярной карциномы, желудочного рака, рака мочевого пузыря, рака легкого, почечно-клеточной карциномы, рака шейки матки, рака ободочной кишки, колоректального рака и рака поджелудочной железы.
33. Способ по п.30, отличающийся тем, что указанное животное является человеком.
34. Способ по п.31, отличающийся тем, что указанное животное является человеком.
35. Антитело или его фрагмент по п.20, отличающееся тем, что указанное антитело является моноклональным антителом 2.14Н9ОРТ.
36. Очищенное антитело или фрагмент антитела, отличающееся тем, что указанное антитело или фрагмент антитела иммуноспецифически связывается с В7-Н1 и содержит вариабельный домен тяжелой цепи, имеющий по меньшей мере 90% идентичность относительно аминокислотной последовательности SEQ ID NO:72, и содержит вариабельный домен легкой цепи, имеющий по меньшей мере 90% идентичность относительно аминокислотной последовательности SEQ ID NO:77, причем указанное антитело обладает активностью связывания с В7-Н1.
37. Композиция, содержащая антитело по п.36.
38. Фармацевтическая композиция, содержащая антитело по п.36 и фармацевтически приемлемый носитель.
39. Способ репрессии В7-Н1-медиируемого ингибирования Т-клеток у животного, включающий введение животному, нуждающемуся в этом, эффективного количества композиции по п.37.
40. Способ лечения злокачественной опухоли у животного, включающий: выбор животного, нуждающегося в лечении злокачественной опухоли; и введение указанному животному терапевтически эффективного количества композиции по п.38.
41. Способ по п.40, отличающийся тем, что указанную злокачественную опухоль выбирают из группы, состоящей из меланомы, немелкоклеточного рака легкого, гепатоцеллюлярной карциномы, желудочного рака, рака мочевого пузыря, рака легкого, почечно-клеточной карциномы, рака шейки матки, рака ободочной кишки, колоректального рака и рака поджелудочной железы.
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