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RU2021118125A - Способы получения экспрессирующих химерный антигенный рецептор клеток - Google Patents

Способы получения экспрессирующих химерный антигенный рецептор клеток Download PDF

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RU2021118125A
RU2021118125A RU2021118125A RU2021118125A RU2021118125A RU 2021118125 A RU2021118125 A RU 2021118125A RU 2021118125 A RU2021118125 A RU 2021118125A RU 2021118125 A RU2021118125 A RU 2021118125A RU 2021118125 A RU2021118125 A RU 2021118125A
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cells
population
immune effector
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Фелипе БЕДОЙА
Саба ГХАССЕМИ
Карл Х. ДЖУН
Брюс Л. ЛЕВИН
Ян Дж. МИЛЕНХОРСТ
Майкл С. МАЙЛОН
Дэниэл Дж. Мл. ПАУЭЛЛ
Зоуи ЧЖЭН
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Новартис Аг
Дзе Трастиз Оф Дзе Юниверсити Оф Пенсильвания
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Claims (37)

1. Способ получения популяции иммунных эффекторных клеток, экспрессирующих химерный антигенный рецептор (CAR), причем данный способ включает
(a) обеспечение популяции иммунных эффекторных клеток, где множество иммунных эффекторных клеток в популяции содержит нуклеиновую кислоту кодирующую CAR, и
(b) контактирование популяции иммунных эффекторных клеток с интерлейкином-15 (IL-15), интерлейкином-7 (IL-7) или комбинацией IL=15 и IL-7.
2. Способ по п. 1, где (b) включает контактирование популяции иммунных эффекторных клеток с IL-15, но не с IL-7.
3. Способ по п. 1 или 2, где популяцию иммунных эффекторных клеток размножают в течение периода, составляющего менее чем 8 дней, например, менее чем 7 дней, менее чем 6 дней, менее чем 5 дней, менее чем 4 дня или менее чем 3 дня.
4. Способ по п. 1 или 2, где популяцию иммунных эффекторных клеток размножают в культуре в течение 5 дней, и получаемые клетки являются более активными, чем те же клетки, размноженные в культуре в течение 9 дней в тех же условиях культивирования.
5. Способ по п. 1 или 2, где
(i) популяция иммунных эффекторных клеток, размноженных в течение 5 дней, показывает по меньшей мере двухкратное увеличение клеточных удвоений при антигенной стимуляции по сравнению с теми же клетками, размноженными в культуре в течение 9 дней в тех же условиях культивирования;
(iii) популяция иммунных эффекторных клеток размножают в культуре в течение 5 дней, и получаемые клетки проявляют такое же или более высокое производство провоспалительных цитокинов по сравнению с теми же клетками, размноженными в культуре в течение 9 дней в тех же условиях культивирования.
6. Способ по любому из пп. 1-3, где популяцию иммунных эффекторных клеток размножают в течение периода, составляющего менее чем 3 дня.
7. Способ по любому из пп. 1-6, где популяцию иммунных клеток размножают культивированием клеток в присутствии средства, которое стимулирует ассоциированный с комплексом CD3/TCR сигнал, и лиганда, который стимулирует костимулирующую молекулу на поверхности клеток.
8. Способ по п. 7, где средство содержит гранулу, конъюгированную с анти-CD3 антителом или его фрагментом и/или анти-CD28 антителом или его фрагментом.
9. Способ по любому из предшествующих пунктов, где популяция иммунных эффекторных клеток на стадии (b) контактирует с IL-15 или комбинацией IL-15 и рецептора-альфа (Ra) IL-15.
10. Способ по п. 9, где комбинация IL-15 и IL-15Ra представляет собой hetIL-15.
11. Способ по п. 1, где способ дополнительно включает контактирование популяции клеток с анти-CD3/CD28 средством перед контактированием популяции иммунных эффекторных клеток с интерлейкином-15 (IL-15), интерлейкином-7 (IL-7) или комбинацией IL-15 и IL-7, и где популяция иммунных эффекторных клеток представляет собой клетки человека, имеющего рак.
12. Способ по п. 3, где получившаяся популяция иммунных эффекторных клеток показывает более эффективную способность клеточного киллинга, чем те же клетки, размноженные в культуре в течение 9 дней или более в тех же условиях культивирования.
13. Способ по п. 1, где обеспеченная популяция иммунных эффекторных клеток представляет собой популяцию истощенных по T-регуляторам клеток, где необязательно популяцию истощенных по Т-регуляторам клеток получают путем удаления CD25+ Т-клеток из исходной популяции имуннх эффекторных клеток, полученных у субъекта.
14. Способ по п. 13, где исходная популяция имунных эффекторных клеток представляет собой клетки субъекта, имеющего рак, например, субъекта, имеющего CD25-экспрессирующий рак, такой как, например, хронический лимфоцитарный лейкоз (ХЛЛ).
15. Способ по п. 14, где обеспеченная популяция иммунных эффекторных клеток содержит
(i) менее чем 50% CD25+ Т-клеток; или
(ii) менее чем 50% CD25+ Т-клеток и менее чем 50% CD25-экспрессирующих раковых клеток,
по сравнению с исходной популяцией иммунных эффекторных клеток, полученных у субъекта, имеющего рак.
16. Способ по п. 15, где обеспеченная популяция иммунных эффекторных клеток содержит менее чем 15%, 10%, 5%, 4%, 3%, 2% или 1% CD25+ клеток и менее чем 15%, 10%, 5%, 4%, 3%, 2% или 1% раковых клеток, например, CD25-экспрессирующих раковых клеток, например, клеток ХЛЛ, или где обеспеченная популяция иммунных эффекторных клеток содержит менее чем 10%, 5%, 4%, 3%, 2% или 1% CD25+ клеток и менее чем 10%, 5%, 4%, 3%, 2% или 1% раковых клеток, например, CD25-экспрессирующих раковых клеток, например, клеток ХЛЛ, по сравнению с исходной популяцией иммунных эффекторных клеток, полученных у субъекта, имеющего рак.
17. Способ по п. 16, где Т-регуляторные клетки, например, CD25+ T-клетки, удаляют из исходной популяции иммунных эффекторных клеток, используя анти-CD25 антитело или его фрагмент, необязательно, где анти-CD25 антитело или его фрагмент конъюгированы с субстратом, например, гранулой.
18. Способ по любому из пп. 13-17, где исходная популяция иммунных эффекторных клеток получена от субъекта, имеющего гематологический рак, например, лейкоз, например, хронический лимфоцитарный лейкоз (ХЛЛ), острый лимфоцитарный лейкоз (ОЛЛ), или лимфому, например, мантийноклеточную лимфому (МКЛ).
19. Способ по любому из предшествующих пунктов, в котором популяцию иммунных эффекторных клеток активируют перед контактированием с популяцией иммунных эффекторных клеток с IL-15 млм IL-7.
20. Способ по п. 13, где популяция иммунных эффекторных клеток подвергается по меньшей мере 200-кратному (например, 200-кратному, 250-кратному, 300-кратному, 350-кратному) увеличению количества клеток в течение 14-дневного периода размножения.
21. Способ по любому из предшествующих пунктов, дополнительно включающий контактирование популяции иммунных эффекторных клеток с нуклеиновой кислотой, кодирующей субъединицу теломеразы, например, hTERT, и где нуклеиновая кислота представляет собой, например, ДНК или РНК.
22. Способ по любому из предшествующих пунктов, где нуклеиновая кислота кодирует CAR CD19.
23. Реакционная смесь, содержащая
(а) популяцию иммунных эффекторных клеток, где множество клеток популяции в реакционной смести содержит молекулу нуклеиновой кислоты, например, описанную здесь молекулу нуклеиновой кислоты, которая содержит последовательность, кодирующую CAR, например, последовательность, кодирующую CAR CD19, например, как описано здесь, и
(b) IL-7 и/или IL-15.
24. Реакционная смесь по п. 23, в которой молекула нуклеиновой кислоты содержит ДНК, РНК, плазмиду, лентивирусный вектор, аденовирусный вектор или ретровирусный вектор.
25. Реакционная смесь по п. 23 или 24, дополнительно содержащая средство, которое активирует и/или размножает клетки популяции, например, средство, которое стимулирует ассоциированный с комплексом CD3/TCR сигнал, и/или лиганд, который стимулирует костимулирующую молекулу на поверхности клеток, например, в которой средство представляет собой гранулу, конъюгированную с анти-CD3 антителом или его фрагментом и/или анти-CD28 антителом или его фрагментом.
26. Реакционная смесь по любому из пп. 23-25, в которой реакционная смесь содержит популяцию истощенных по T-регуляторам клеток, содержащую менее чем 50%, 40%, 30%, 25%, 20%, 15%, 10%, 5%, 4%, 3%, 2% или 1% CD25+ клеток и менее чем 50%, 40%, 30%, 25%, 20%, 15%, 10%, 5%, 4%, 3%, 2% или 1% экспрессирующих ингибитор контрольных точек клеток, например, PD1+ клеток, LAG3+ клеток или TIM3+ клеток, по сравнению с реакционной смесью, содержащей исходную популяцию иммунных эффекторных клеток, полученных у субъекта.
27. Реакционная смесь по любому из пп. 23-26, которая содержит IL-15, но не содержит IL-7.
28. Реакционная смесь по любому из п.п. 23-27, дополнительно содержащая криопротектор или стабилизатор, такой как, например, сахарид, олигосахарид, полисахарид и полиол (например, трегалозу, маннит, сорбит, лактозу, сахарозу, глюкозу и декстран), соли и краун-эфиры, необязательно, где криопротектор представляет собой декстран.
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