JP2017018125A5 - - Google Patents

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JP2017018125A5
JP2017018125A5 JP2016163762A JP2016163762A JP2017018125A5 JP 2017018125 A5 JP2017018125 A5 JP 2017018125A5 JP 2016163762 A JP2016163762 A JP 2016163762A JP 2016163762 A JP2016163762 A JP 2016163762A JP 2017018125 A5 JP2017018125 A5 JP 2017018125A5
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compact talen
talen monomer
catalytic domain
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  1. (a)興味対象の1つのDNA標的配列を二本鎖DNAの一方の鎖上で選択し;
    (b)下記:
    (i)前記興味対象のDNA標的配列に対するDNA結合特異性を有する反復可変ジペプチド領域(RVD)を含んでなる1つのコアTALE足場;
    (ii)前記二本鎖DNAに対する切断活性を有し、(i)のコアTALE足場のC末端及び/又はN末端に融合されたとき前記興味対象のDNA標的配列から数塩基対離れたDNAをプロセシングし得る少なくとも1つの触媒ドメインであって、メガヌクレアーゼバリアントを含む触媒ドメイン;
    (iii)任意に、必要な場合に(ii)の触媒ドメインを(i)のコアTALE足場に融合させるための1つのペプチドリンカー
    を含んでなり、二量体化を要することなく、前記二本鎖DNAに結合してプロセシングするように組み立てられている独特なコンパクトTALENモノマーを準備し;
    (c)前記二本鎖DNAを前記独特なモノマーと、該二本鎖が前記一本鎖標的配列から3'及び/又は5'方向に数塩基対離れた場所でプロセシングされるようにインビトロで接触させる
    ことを含んでなる、インビトロで二本鎖DNAを標的してプロセシングする方法。
  2. 前記触媒ドメインが単鎖メガヌクレアーゼであるメガヌクレアーゼバリアントを含む請求項1に記載の方法。
  3. 前記触媒ドメインが少なくとも2つのLAGLIDADGモチーフを含むメガヌクレアーゼバリアントを含む請求項1又は2に記載の方法。
  4. 前記LAGLIDADGモチーフの少なくとも1つがI-CreIに由来する請求項3に記載の方法。
  5. 前記触媒ドメインが前記コアTALE足場のC末端ドメインに融合されている請求項1〜4のいずれか1項に記載の方法。
  6. 前記触媒ドメインが前記コアTALE足場のN末端ドメインに融合されている請求項1〜5のいずれか1項に記載の方法。
  7. 前記触媒ドメインが配列番号1と少なくとも70%、好ましくは80%、より好ましくは90%、尚より好ましくは95%の配列同一性を有するポリペプチド配列を含んでなる請求項1〜6のいずれか1項に記載の方法。
  8. 前記コアTALE足場が配列番号134及び配列番号135からなる群より選択されるタンパク質配列と、少なくとも80%、より好ましくは90%、またより好ましくは95%のアミノ酸配列同一性を有するタンパク質配列を含んでなる請求項1〜7のいずれか1項に記載の方法。
  9. 前記コアTALE足場が配列番号136〜配列番号139からなる群より選択されるタンパク質配列と、少なくとも80%、より好ましくは90%、またより好ましくは95%のアミノ酸配列同一性を有するタンパク質配列を含んでなる請求項1〜7のいずれか1項に記載の方法。
  10. 前記コンパクトTALENモノマーが細胞で安定に又は一過性に発現する請求項1〜9のいずれか1項に記載の方法。
  11. 前記細胞が哺乳動物細胞である請求項10に記載の方法。
  12. 前記細胞が初代細胞である請求項11に記載の方法。
  13. 前記細胞が遺伝子操作T細胞を作製するためのT細胞である請求項10又は11に記載の方法。
  14. 前記コンパクトTALENモノマーがT細胞レセプター遺伝子中の配列を標的するように設計されている請求項13に記載の方法。
  15. 前記遺伝子操作T細胞が悪性腫瘍又は感染性疾患の治療用である請求項13又は14に記載の方法。
  16. 前記触媒ドメインが配列番号439〜441及び配列番号444〜446の群より選択されるタンパク質配列と、少なくとも80%、好ましくは90%、より好ましくは95%のアミノ酸配列同一性を有するタンパク質配列を含んでなる請求項15に記載の方法。
  17. 下記:
    (i)興味対象の特異的二本鎖DNA標的配列に対するDNA結合特異性を有する反復可変ジペプチド領域(RVD)を含んでなる1つのコアTALE足場;
    (ii)前記二本鎖DNAに対する切断活性を有し、(i)のコアTALE足場のC末端又はN末端に融合されたとき前記興味対象の二本鎖DNA標的配列から数塩基対離れたDNAをプロセシングし得る、メガヌクレアーゼバリアントを含む少なくとも1つの触媒ドメイン;
    (iii)任意に、必要な場合に(ii)の触媒ドメインを(i)の遺伝子操作コアTALE足場に融合させるための1つのペプチドリンカー
    を含んでなり、二量体化を要することなく、前記標的DNA配列に結合して二本鎖DNAをプロセシングするように組み立てられているコンパクトTALENモノマー。
  18. 前記触媒ドメインが単鎖メガヌクレアーゼであるメガヌクレアーゼバリアントを含む請求項17に記載のコンパクトTALENモノマー。
  19. 前記触媒ドメインが少なくとも2つのLAGLIDADGモチーフを含むメガヌクレアーゼバリアントを含む請求項17又は18に記載のコンパクトTALENモノマー。
  20. 前記LAGLIDADGモチーフの少なくとも1つがI-CreIに由来する請求項19に記載のコンパクトTALENモノマー。
  21. 前記触媒ドメインが前記コアTALE足場のC末端ドメインに融合されている請求項17〜20のいずれか1項に記載のコンパクトTALENモノマー。
  22. 前記触媒ドメインが前記コアTALE足場のN末端ドメインに融合されている請求項17〜20のいずれか1項に記載のコンパクトTALENモノマー。
  23. 前記触媒ドメインが配列番号1と少なくとも70%、好ましくは80%、より好ましくは90%、尚より好ましくは95%の配列同一性を有するポリペプチド配列を含んでなる請求項17〜22のいずれか1項に記載のコンパクトTALENモノマー。
  24. T細胞レセプター遺伝子中の配列を標的するように設計されている請求項17〜23に記載のコンパクトTALENモノマー。
  25. 配列番号444〜446の群より選択されるタンパク質配列と、少なくとも80%、より好ましくは90%、またより好ましくは95%のアミノ酸配列同一性を有するタンパク質配列を含んでなる請求項24に記載のコンパクトTALENモノマー。
  26. 治療剤としての請求項17〜25のいずれか1項に記載のコンパクトTALENモノマー。
  27. 悪性腫瘍又は感染性疾患の治療用治療剤として用いるための請求項17〜26のいずれか1項に記載のコンパクトTALENモノマー。
  28. 前記コアTALE足場が配列番号134及び配列番号135からなる群より選択されるタンパク質配列と、少なくとも80%、より好ましくは90%、またより好ましくは95%のアミノ酸配列同一性を有するタンパク質配列を含んでなる請求項17〜27のいずれか1項に記載のコンパクトTALENモノマー。
  29. 前記コアTALE足場が配列番号136〜配列番号139からなる群より選択されるタンパク質配列と、少なくとも80%、より好ましくは90%、またより好ましくは95%のアミノ酸配列同一性を有するタンパク質配列を含んでなる請求項17〜28のいずれか1項に記載のコンパクトTALENモノマー。
  30. 前記ペプチドリンカー配列が配列番号67〜104及び配列番号372〜配列番号415からなる群より選択され得る請求項17〜29のいずれか1項に記載のコンパクトTALENモノマー。
  31. 請求項17〜30のいずれか1項に記載のコンパクトTALENモノマーをコードする組換えポリヌクレオチド。
  32. 請求項17〜30のいずれか1項に記載のコンパクトTALENモノマー又は請求項31に記載の組換えポリヌクレオチドを含んでなる医薬組成物。
  33. 請求項31に記載の組換えポリヌクレオチドを含んでなる宿主細胞。
  34. インビトロで、細胞、組織又は非ヒト動物の遺伝子座の特異的な単一の二本鎖DNA標的配列中に導入遺伝子を挿入する方法であって、少なくとも1つの請求項17〜30のいずれか1項に記載のコンパクトTALENモノマーが前記細胞、組織又は非ヒト動物に導入される方法。
JP2016163762A 2011-04-05 2016-08-24 コンパクトtale−ヌクレアーゼを作製する方法及びその使用 Active JP6695239B2 (ja)

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US201161472065P 2011-04-05 2011-04-05
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US201161496454P 2011-06-13 2011-06-13
US61/496,454 2011-06-13
US201161499047P 2011-06-20 2011-06-20
US201161499043P 2011-06-20 2011-06-20
US61/499,043 2011-06-20
US61/499,047 2011-06-20
US201161533098P 2011-09-09 2011-09-09
US201161533123P 2011-09-09 2011-09-09
US61/533,123 2011-09-09
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