RU2013110779A - Нуклеиновая кислота, содержащая или кодирующая гистоновую структуру типа "стебель-петля" и поли(а)-последовательность или сигнал полиаденилирования, для повышения экспрессии кодируемого белка - Google Patents
Нуклеиновая кислота, содержащая или кодирующая гистоновую структуру типа "стебель-петля" и поли(а)-последовательность или сигнал полиаденилирования, для повышения экспрессии кодируемого белка Download PDFInfo
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- RU2013110779A RU2013110779A RU2013110779/10A RU2013110779A RU2013110779A RU 2013110779 A RU2013110779 A RU 2013110779A RU 2013110779/10 A RU2013110779/10 A RU 2013110779/10A RU 2013110779 A RU2013110779 A RU 2013110779A RU 2013110779 A RU2013110779 A RU 2013110779A
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- nucleic acid
- sequence
- stem
- diseases
- loop
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Abstract
1. Нуклеиновая кислота, которая содержит или кодируета) кодирующую область, которая предпочтительно кодирует пептид или белок; при условии, что кодирующая область не кодирует гистоновые белки, репортерные белки, выбранные из EGFP и люциферазы, и маркерные или применяемые для селекции белки, выбранные из альфа-глобина, галактокиназы и ксантин-гуанинфосфорибозилтрансферазы (GPT)),б) по меньшей мере одну гистоновую структуру типа «стебель-петля» ив) поли(A)-последовательность или сигнал полиаденилирования.2. Нуклеиновая кислота по п.1, где нуклеиновая кислота не содержит один или два, или по меньшей мере один или все, кроме одного, или все компоненты, выбранные из группы, включающей: последовательность, которая кодирует рибозим (предпочтительно обладающий способностью к автономному сплайсингу рибозим), вирусную нуклеотидную последовательность, сигнал процессинга гистоновой структуры типа «стебель-петля», в частности сигнал процессинга гистоновой структуры типа «стебель-петля», выведенный из гена мышиного гистона H2A614, ген Neo, инактивированную промоторную последовательность и инактивированную энхансерную последовательность.3. Нуклеиновая кислота по п.1 или 2, где нуклеиновая кислота не содержит рибозим, предпочтительно обладающий способностью к автономному сплайсингу рибозим, и один или несколько компонентов, выбранных из группы, включающей: ген Neo, инактивированную промоторную последовательность и инактивированную энхансерную последовательность, сигнал процессинга гистоновой структуры типа «стебель-петля», в частности сигнал процессинга гистоновой структуры типа «стебель-петля», выведенный из гена мышиного
Claims (17)
1. Нуклеиновая кислота, которая содержит или кодирует
а) кодирующую область, которая предпочтительно кодирует пептид или белок; при условии, что кодирующая область не кодирует гистоновые белки, репортерные белки, выбранные из EGFP и люциферазы, и маркерные или применяемые для селекции белки, выбранные из альфа-глобина, галактокиназы и ксантин-гуанинфосфорибозилтрансферазы (GPT)),
б) по меньшей мере одну гистоновую структуру типа «стебель-петля» и
в) поли(A)-последовательность или сигнал полиаденилирования.
2. Нуклеиновая кислота по п.1, где нуклеиновая кислота не содержит один или два, или по меньшей мере один или все, кроме одного, или все компоненты, выбранные из группы, включающей: последовательность, которая кодирует рибозим (предпочтительно обладающий способностью к автономному сплайсингу рибозим), вирусную нуклеотидную последовательность, сигнал процессинга гистоновой структуры типа «стебель-петля», в частности сигнал процессинга гистоновой структуры типа «стебель-петля», выведенный из гена мышиного гистона H2A614, ген Neo, инактивированную промоторную последовательность и инактивированную энхансерную последовательность.
3. Нуклеиновая кислота по п.1 или 2, где нуклеиновая кислота не содержит рибозим, предпочтительно обладающий способностью к автономному сплайсингу рибозим, и один или несколько компонентов, выбранных из группы, включающей: ген Neo, инактивированную промоторную последовательность и инактивированную энхансерную последовательность, сигнал процессинга гистоновой структуры типа «стебель-петля», в частности сигнал процессинга гистоновой структуры типа «стебель-петля», выведенный из гена мышиного гистона H2A614.
4. Нуклеиновая кислота по п.1 или 2, где нуклеиновая кислота представляет собой РНК, предпочтительно мРНК.
5. Нуклеиновая кислота по п.1 или 2, в которой по меньшей мере одна гистоновая структура типа «стебель-петля» выбрана из структур, имеющих следующие формулы (I) или (II):
формула (I) (последовательность структуры типа «стебель-петля» без пограничных элементов «стебля»):
формула (II) (последовательность структуры типа «стебель-петля» с пограничными элементами «стебля»):
в которых:
пограничные элементы stem1 или stem2 N1-6 обозначают непрерывную последовательность, состоящую из 1-6, предпочтительно 2-6, более предпочтительно 2-5, еще более предпочтительно 3-5, наиболее предпочтительно 4-5 или 5 N, где N каждый независимо друг от друга выбран из нуклеотида, выбранного из A, U, T, G и C или их нуклеотидного аналога;
stem1 [N0-2GN3-5] обозначает последовательность, обратно комплементарную или частично обратно комплементарную элементу stem2, и представляет собой непрерывную последовательность, состоящую из 5-7 нуклеотидов;
в которой N0-2 обозначает непрерывную последовательность, состоящую из 0-2, предпочтительно 0-1, более предпочтительно 1 N, где N каждый независимо друг от друга выбран из нуклеотида, выбранного из A, U, T, G и C или его нуклеотидного аналога;
в которой N3-5 обозначает непрерывную последовательность, состоящую из 3-5, предпочтительно 4-5, более предпочтительно 4 N, где N каждый независимо друг от друга выбран из нуклеотида, выбранного из A, U, T, G и C или его нуклеотидного аналога, и в которой G обозначает гуанозин или его аналог и необязательно может быть заменен на цитидин или его аналог при условии, что комплементарный ему нуклеотид цитидин в stem2 заменен на гуанозин;
последовательность «петли» [N0-4(U/T)N0-4] локализована между элементами stem1 и stem2 и обозначает непрерывную последовательность, состоящую из 3-5 нуклеотидов, более предпочтительно 4 нуклеотидов;
в которой N0-4 каждый независимо друг от друга обозначает непрерывную последовательность, состоящую из 0-4, предпочтительно 1-3, более предпочтительно 1-2 N, где N каждый независимо друг от друга выбран из нуклеотида, выбранного из A, U, T, G и C или его нуклеотидного аналога; и в которой U/T обозначает уридин или необязательно тимидин;
stem2 [N3-5CN0-2] обозначает последовательность, обратно комплементарную или частично обратно комплементарную элементу stem1, и представляет собой непрерывную последовательность, состоящую из 5-7 нуклеотидов;
в которой N3-5 обозначает непрерывную последовательность, состоящую 3-5, предпочтительно 4-5, более предпочтительно 4 N, где N каждый независимо друг от друга выбран из нуклеотида, выбранного из A, U, T, G и C или его нуклеотидного аналога;
в которой N0-2 обозначает непрерывную последовательность, состоящую из 0-2, предпочтительно 0-1, более предпочтительно 1 N, где N каждый независимо друг от друга выбран из нуклеотида, выбранного из A, U, T, G или C или его нуклеотидного аналога; и
в которой C обозначает цитидин или его аналог и необязательно может быть заменен на гуанозин или его аналог при условии, что комплементарный ему нуклеотид гуанозин в stem1 заменен на цитидин;
где
у stem1 и stem2 может происходить спаривание оснований друг с другом
с образованием обратно комплементарной последовательности, где спаривание оснований может иметь место между stem1 и stem2, или
с образованием частично обратно комплементарной последовательности, если может происходить неполное спаривание оснований между stem1 и stem2.
7. Нуклеиновая кислота по п.п.1, 2 или 6, в которой поли(A)-последовательность включает последовательность, которая содержит от примерно 25 до примерно 400 аденозиновых нуклеотидов, предпочтительно последовательность, которая содержит от примерно 50 до примерно 400 аденозиновых нуклеотидов, более предпочтительно последовательность, которая содержит от примерно 50 до примерно 300 аденозиновых нуклеотидов, еще более предпочтительно последовательность, которая содержит от примерно 50 до примерно 250 аденозиновых нуклеотидов, наиболее предпочтительно последовательность, которая содержит от примерно 60 до примерно 250 аденозиновых нуклеотидов.
8. Нуклеиновая кислота по п.п.1, 2 или 6, в которой сигнал полиаденилирования содержит консенсусную последовательность NNUANA, предпочтительно AAUAAA или AUUAAA.
9. Нуклеиновая кислота по п.п.1, 2 или 6, где нуклеиновая кислота кодирует терапевтически активный белок или пептид, адъювантный белок, антиген, опухолевый антиген, патогенный антиген, антиген из животных, вирусный антиген, антиген простейших, бактериальный антиген, аллергенный антиген, аутоиммунный антиген, аллерген, антитело, иммуностимулирующий белок или пептид или антигенспецифический T-клеточный рецептор.
10. Нуклеиновая кислота по п.п.1, 2 или 6, где нуклеиновая кислота представляет собой моноцистронную, бицистронную или даже мультицитронную нуклеиновую кислоту.
11. Применение нуклеиновой кислоты по одному из п.п.1-10 для повышения уровня экспрессии кодируемого пептида или белка.
12. Нуклеиновая кислота по п.п.1, 2 или 6, предназначенная для применения в качестве лекарственного средства.
13. Фармацевтическая композиция, содержащая нуклеиновую кислоту по одному из п.п.1-10 и необязательно фармацевтически приемлемый носитель.
14. Вакцина, содержащая нуклеиновую кислоту по одному из п.п.1-10, которая кодирует антиген или содержит антиген.
15. Вакцина по п.14, в которой антиген выбран из опухолевых антигенов, аллергенных антигенов, аутоиммунных аутоантигенов, патогенных антигенов, вирусных антигенов, бактериальных антигенов, грибных антигенов, антигенов простейших, антигенов животных или аллергенных антигенов.
16. Применение нуклеиновой кислоты по одному из п.п.1-10 для повышения уровня экспрессии белка при лечении раковых или опухолевых заболеваний, инфекционных болезней, предпочтительно (вирусных, бактериальных или протозойных) инфекционных болезней, аутоиммунных заболеваний, аллергий или аллергических заболеваний, моногенных заболеваний, т.е. (наследственных) заболеваний или генетических заболеваний в целом, заболеваний, которые имеют генетически наследуемую основу и которые, как правило, вызываются дефектом одного гена и наследуются согласно законам Менделя, сердечно-сосудистых заболеваний, заболеваний нервной системы, болезней дыхательной системы, болезней пищеварительной системы, заболеваний кожи, скелетно-мышечных нарушений, нарушений соединительной ткани, неоплазм, иммунодефицита, эндокринных, трофических и метаболических заболеваний, глазных болезней или ушных болезней.
17. Способ in vitro или ex vivo повышения уровня экспрессии белка, заключающийся в том, что:
а) получают нуклеиновую кислоту по одному из п.п.1-10,
б) вносят или вводят нуклеиновую кислоту в бесклеточную экспрессионную систему, в клетку (например, предназначенную для экспрессии клетку-хозяина или соматическую клетку), ткань или организм.
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EPPCT/EP2010/004998 | 2010-08-13 | ||
PCT/EP2011/004077 WO2012019780A1 (en) | 2010-08-13 | 2011-08-12 | Nucleic acid comprising or coding for a histone stem-loop and a poly(a) sequence or a polyadenylation signal for increasing the expression of an encoded protein |
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2017
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2020
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