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- JPWO2020033916A5 JPWO2020033916A5 JP2021506661A JP2021506661A JPWO2020033916A5 JP WO2020033916 A5 JPWO2020033916 A5 JP WO2020033916A5 JP 2021506661 A JP2021506661 A JP 2021506661A JP 2021506661 A JP2021506661 A JP 2021506661A JP WO2020033916 A5 JPWO2020033916 A5 JP WO2020033916A5
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Description
提供される任意のいくつかの態様において、組み込まれていない残りの組換え配列は、ベクタープラスミド、直鎖状相補DNA（cDNA）、自己組み込み体（autointegrant）、または末端反復配列（LTR）サークルのうちの1つまたは複数を含む。
[本発明1001]
以下の工程を含む、導入遺伝子配列のゲノム組み込みを評価するための方法：
（a）10キロベース（kb）よりも大きいかまたは約10キロベース（kb）よりも大きいデオキシリボ核酸（DNA）の高分子量画分を、1つまたは複数の細胞より単離されたDNAから分離する工程であって、該1つまたは複数の細胞が、組換えタンパク質をコードする導入遺伝子配列を含む少なくとも1つの操作された細胞を含むか、またはそれを含む疑いがある、工程；
（b）該高分子量画分から、該1つまたは複数の細胞のゲノム中に組み込まれた導入遺伝子配列の存在、不在、または量を決定する工程。
[本発明1002]
（a）における分離する工程の前に、デオキシリボ核酸（DNA）を、前記1つまたは複数の細胞から単離する、本発明1001の方法。
[本発明1003]
（b）における導入遺伝子配列の存在、不在、または量を決定する工程が、前記1つまたは複数の細胞における二倍体ゲノム当たりのまたは細胞当たりの、該導入遺伝子配列の質量、重量、またはコピー数を決定することを含む、本発明1001または本発明1002の方法。
[本発明1004]
前記1つまたは複数の細胞が、細胞の集団を含み、該集団のうちの複数の細胞が、前記組換えタンパク質をコードする前記導入遺伝子配列を含む、本発明1001～1003のいずれかの方法。
[本発明1005]
前記コピー数が、前記細胞の集団中での二倍体ゲノム当たりのまたは細胞当たりの平均（average）または平均（mean）コピー数である、本発明1003または本発明1004の方法。
[本発明1006]
（a）における分離する工程の前に、組換えタンパク質をコードする導入遺伝子配列を含むポリヌクレオチドが、前記1つまたは複数の細胞のうちの少なくとも1つの操作された細胞中に導入されている、本発明1001～1005のいずれかの方法。
[本発明1007]
前記少なくとも1つの操作された細胞が、前記導入遺伝子配列を含む前記ポリヌクレオチドの導入後に、96時間超にわたり、25℃よりも高い温度で、任意で37℃±2℃でまたは約37℃±2℃でインキュベートされていない、本発明1006の方法。
[本発明1008]
前記少なくとも1つの操作された細胞が、前記導入遺伝子配列を含む前記ポリヌクレオチドの導入後に、72時間超にわたり、25℃よりも高い温度で、任意で37℃±2℃でまたは約37℃±2℃でインキュベートされていない、本発明1006の方法。
[本発明1009]
前記少なくとも1つの操作された細胞が、前記導入遺伝子配列を含む前記ポリヌクレオチドの導入後に、48時間超にわたり、25℃よりも高い温度で、任意で37℃±2℃でまたは約37℃±2℃でインキュベートされていない、本発明1006の方法。
[本発明1010]
前記1つまたは複数の細胞が、（a）におけるDNAの高分子量画分を分離する工程の前に、凍結保存されている、本発明1001～1009のいずれかの方法。
[本発明1011]
前記1つまたは複数の細胞が、細胞株である、本発明1001～1010のいずれかの方法。
[本発明1012]
前記1つまたは複数の細胞が、対象由来のサンプルから得られた初代細胞である、本発明1001～1010のいずれかの方法。
[本発明1013]
前記1つまたは複数の細胞が、免疫細胞である、本発明1001～1012のいずれかの方法。
[本発明1014]
前記免疫細胞が、T細胞またはNK細胞である、本発明1013の方法。
[本発明1015]
前記T細胞が、CD3+、CD4+、および/またはCD8+ T細胞である、本発明1014の方法。
[本発明1016]
以下の工程を含む、対象由来の生物学的サンプルにおいて導入遺伝子配列を評価するための方法：
（a）10キロベース（kb）よりも大きいかまたは約10キロベース（kb）よりも大きいデオキシリボ核酸（DNA）の高分子量画分を、対象由来の生物学的サンプル中に存在する1つまたは複数の細胞より単離されたDNAから分離する工程であって、該生物学的サンプルが、組換えタンパク質をコードする導入遺伝子配列を含む少なくとも1つの操作された細胞を含むか、またはそれを含む疑いがある、工程； および
（b）該高分子量画分から、該生物学的サンプルの全てまたは一部分における導入遺伝子配列の存在、不在、または量を決定する工程。
[本発明1017]
（b）における導入遺伝子配列の存在、不在、または量を決定する工程が、前記生物学的サンプルの全てまたは一部分における該導入遺伝子配列の質量、重量、またはコピー数を決定することを含む、本発明1016の方法。
[本発明1018]
分離する工程の前に、DNAを、前記生物学的サンプル中に存在する1つまたは複数の細胞から単離する、本発明1016または本発明1017の方法。
[本発明1019]
前記生物学的サンプルが、前記導入遺伝子配列を含む前記少なくとも1つの操作された細胞を含む組成物が投与された対象から得られる、本発明1016～1018のいずれかの方法。
[本発明1020]
前記生物学的サンプルが、組織サンプルまたは体液サンプルである、本発明1016～1019のいずれかの方法。
[本発明1021]
前記生物学的サンプルが組織サンプルであり、該組織が腫瘍である、本発明1020の方法。
[本発明1022]
前記組織サンプルが腫瘍生検材料である、本発明1020または本発明1021の方法。
[本発明1023]
前記生物学的サンプルが体液サンプルであり、該体液サンプルが、血液または血清サンプルである、本発明1020の方法。
[本発明1024]
（a）における分離する工程の前に、前記組換えタンパク質をコードする前記導入遺伝子配列を含むポリヌクレオチドが、前記1つまたは複数の細胞のうちの前記少なくとも1つの操作された細胞中に導入されている、本発明1016～1023のいずれかの方法。
[本発明1025]
前記生物学的サンプルにおける前記1つまたは複数の細胞が、免疫細胞を含む、本発明1016～1024のいずれかの方法。
[本発明1026]
前記免疫細胞が、T細胞またはNK細胞である、本発明1025の方法。
[本発明1027]
前記T細胞が、CD3+、CD4+、および/またはCD8+ T細胞である、本発明1026の方法。
[本発明1028]
前記分離する工程が、パルスフィールドゲル電気泳動またはサイズ排除クロマトグラフィーによって行われる、本発明1001～1027のいずれかの方法。
[本発明1029]
前記分離する工程が、パルスフィールドゲル電気泳動によって行われる、本発明1001～1028のいずれかの方法。
[本発明1030]
以下の工程を含む、導入遺伝子配列のゲノム組み込みを評価するための方法：
（a）パルスフィールドゲル電気泳動によって、10キロベース（kb）よりも大きいかまたは約10キロベース（kb）よりも大きいデオキシリボ核酸（DNA）の高分子量画分を、細胞の集団より単離されたDNAから分離する工程であって、該細胞の集団が、組換えタンパク質をコードする導入遺伝子配列を各々が含むかまたはそれを含む疑いがある複数の操作された細胞を含む、工程；および
（b）該高分子量画分から、該細胞の集団のうちの該複数の操作された細胞のゲノム中に組み込まれた該導入遺伝子配列の二倍体ゲノム当たりのまたは細胞当たりの、平均（average）または平均（mean）コピー数を決定する工程。
[本発明1031]
（a）における分離する工程の前に、前記組換えタンパク質をコードする前記導入遺伝子配列を含むポリヌクレオチドが、前記細胞の集団のうちの前記複数の操作された細胞の少なくとも1つ中に導入されている、本発明1030の方法。
[本発明1032]
前記細胞の集団が、前記導入遺伝子配列を含む前記ポリヌクレオチドの、前記少なくとも1つの操作された細胞中への導入後に、96時間超にわたり、25℃よりも高い温度で、任意で37℃±2℃でまたは約37℃±2℃でインキュベートされていない、本発明1031の方法。
[本発明1033]
前記細胞の集団が、前記導入遺伝子配列を含む前記ポリヌクレオチドの、前記少なくとも1つの操作された細胞中への導入後に、72時間超にわたり、25℃よりも高い温度で、任意で37℃±2℃でまたは約37℃±2℃でインキュベートされていない、本発明1031の方法。
[本発明1034]
前記細胞の集団が、前記導入遺伝子配列を含む前記ポリヌクレオチドの、前記少なくとも1つの操作された細胞中への導入後に、48時間超にわたり、25℃よりも高い温度で、任意で37℃±2℃でまたは約37℃±2℃でインキュベートされていない、本発明1031の方法。
[本発明1035]
前記細胞の集団が、（a）におけるDNAの高分子量画分を分離する工程の前に、凍結保存されている、本発明1030～1034のいずれかの方法。
[本発明1036]
前記高分子量画分が、15キロベース（kb）よりも大きいかまたは約15キロベース（kb）よりも大きい、本発明1001～1035のいずれかの方法。
[本発明1037]
前記高分子量画分が、17.5キロベース（kb）よりも大きいかまたは約17.5キロベース（kb）よりも大きい、本発明1001～1035のいずれかの方法。
[本発明1038]
前記高分子量画分が、20キロベース（kb）よりも大きいかまたは約20キロベース（kb）よりも大きい、本発明1001～1035のいずれかの方法。
[本発明1039]
前記導入遺伝子配列が、前記組換えタンパク質をコードする核酸配列に機能的に連結された調節エレメントを含む、本発明1001～1038のいずれかの方法。
[本発明1040]
前記導入遺伝子配列の存在、不在、または量を決定する工程が、ポリメラーゼ連鎖反応（PCR）によって行われる、本発明1001～1039のいずれかの方法。
[本発明1041]
前記PCRが、定量的ポリメラーゼ連鎖反応（qPCR）、デジタルPCR、またはドロップレットデジタルPCRである、本発明1040の方法。
[本発明1042]
前記PCRがドロップレットデジタルPCRである、本発明1040または本発明1041の方法。
[本発明1043]
前記導入遺伝子配列の少なくとも一部分に相補的であるか、またはそれを特異的に増幅することができる1つまたは複数のプライマーを用いて、前記PCRが行われる、本発明1040～1042のいずれかの方法。
[本発明1044]
前記1つまたは複数のプライマーが、調節エレメントの配列に相補的であるか、またはそれを特異的に増幅することができる、本発明1043の方法。
[本発明1045]
前記導入遺伝子配列の量を決定する工程が、前記1つまたは複数の細胞から単離されたDNAの質量または重量当たりの、任意で、前記1つまたは複数の細胞から単離されたDNA 1マイクログラム当たりの、該導入遺伝子配列の質量、重量、またはコピー数を評価することを含む、本発明1001～1044のいずれかの方法。
[本発明1046]
前記導入遺伝子配列の量を決定する工程が、1つまたは複数の細胞から単離されたDNA 1マイクログラム当たりの、導入遺伝子配列のマイクログラムでの質量または重量を評価することを含む、本発明1045の方法。
[本発明1047]
前記導入遺伝子配列の量を決定する工程が、前記1つもしくは複数の細胞当たりの、任意で、CD3+、CD4+、および/もしくはCD8+細胞当たりの、ならびに/または前記組換えタンパク質を発現する細胞当たりの、該導入遺伝子配列の質量、重量、またはコピー数を評価することを含む、本発明1001～1044のいずれかの方法。
[本発明1048]
前記導入遺伝子配列の存在、不在、または量を決定する工程が、前記生物学的サンプルにおける二倍体ゲノム当たりのまたは細胞当たりの、該導入遺伝子配列の質量、重量、またはコピー数を評価することを含む、本発明1016～1044のいずれかの方法。
[本発明1049]
前記コピー数が、前記生物学的サンプルにおける前記1つまたは複数の細胞中での二倍体ゲノム当たりのまたは細胞当たりの平均（average）または平均（mean）コピー数である、本発明1048の方法。
[本発明1050]
前記導入遺伝子配列の量を決定する工程が、前記生物学的サンプルの体積当たりの、任意で、前記生物学的サンプル1マイクロリットル当たりのまたは1ミリリットル当たりの、該導入遺伝子配列の質量、重量、またはコピー数を評価することを含む、本発明1016～1044のいずれかの方法。
[本発明1051]
前記導入遺伝子配列の量を決定する工程が、対象の体重または体表面積当たりの、該導入遺伝子配列の質量、重量、またはコピー数を評価することを含む、本発明1016～1044のいずれかの方法。
[本発明1052]
前記導入遺伝子配列の量を決定する工程が、前記高分子量画分における該導入遺伝子配列の質量、重量、またはコピー数を評価すること、および、該質量、重量、またはコピー数を、前記高分子量画分における参照遺伝子の質量、重量、もしくはコピー数に対して、または標準曲線に対して正規化することを含む、本発明1001～1048のいずれかの方法。
[本発明1053]
前記参照遺伝子がハウスキーピング遺伝子である、本発明1052の方法。
[本発明1054]
前記参照遺伝子が、アルブミンをコードする遺伝子（ALB）である、本発明1052または本発明1053の方法。
[本発明1055]
前記参照遺伝子が、リボヌクレアーゼPタンパク質サブユニットp30をコードする遺伝子（RPP30）である、本発明1052または本発明1053の方法。
[本発明1056]
前記単離されたDNAにおける参照遺伝子のコピー数が、参照遺伝子の少なくとも一部分に相補的であるか、またはそれを特異的に増幅することができる1つまたは複数のプライマーを用いたPCRによって行われる、本発明1052～1055のいずれかの方法。
[本発明1057]
前記導入遺伝子配列が、完全なウイルスgagタンパク質をコードしない、本発明1001～1006のいずれかの方法。
[本発明1058]
前記導入遺伝子配列が、完全なHIVゲノム、複製可能なウイルスゲノム、ならびに/または、任意でNef、Vpu、Vif、Vpr、および/もしくはVpxであるアクセサリー遺伝子を含まない、本発明1001～1057のいずれかの方法。
[本発明1059]
前記ポリヌクレオチドの導入が、該ポリヌクレオチドを含むウイルスベクターの形質導入によって行われる、本発明1006～1015、1024～1029、および1031～1057のいずれかの方法。
[本発明1060]
前記ウイルスベクターが、レトロウイルスベクターまたはガンマレトロウイルスベクターである、本発明1059の方法。
[本発明1061]
前記ウイルスベクターがレンチウイルスベクターである、本発明1059または本発明1060の方法。
[本発明1062]
前記ウイルスベクターが、任意でAAV1、AAV2、AAV3、AAV4、AAV5、AAV6、AAV7、またはAAV8ベクターの中から選択される、AAVベクターである、本発明1059の方法。
[本発明1063]
前記ポリヌクレオチドの導入が、物理的送達法によって、任意でエレクトロポレーションによって行われる、本発明1006～1015、1024～1029、および1031～1057のいずれかの方法。
[本発明1064]
前記組換えタンパク質が組換え受容体である、本発明1001～1063のいずれかの方法。
[本発明1065]
前記組換え受容体が、疾患もしくは状態に関連する抗原、または疾患もしくは状態に関連する病変の環境の細胞において発現している抗原に特異的に結合する、本発明1064の方法。
[本発明1066]
前記疾患もしくは状態ががんである、本発明1065の方法。
[本発明1067]
前記抗原が、αvβ6インテグリン（avb6インテグリン）、B細胞成熟抗原（BCMA）、B7-H3、B7-H6、炭酸脱水酵素9（CA9、別名CAIXまたはG250）、がん精巣抗原、がん/精巣抗原1B（CTAG、別名NY-ESO-1およびLAGE-2）、がん胎児性抗原（CEA）、サイクリン、サイクリンA2、C-Cモチーフケモカインリガンド1（CCL-1）、CD19、CD20、CD22、CD23、CD24、CD30、CD33、CD38、CD44、CD44v6、CD44v7/8、CD123、CD133、CD138、CD171、コンドロイチン硫酸プロテオグリカン4（CSPG4）、上皮成長因子タンパク質（EGFR）、タイプIII上皮成長因子受容体変異（EGFR vIII）、上皮糖タンパク質2（EPG-2）、上皮糖タンパク質40（EPG-40）、エフリンB2、エフリン受容体A2（EPHa2）、エストロゲン受容体、Fc受容体様5（FCRL5；別名Fc受容体ホモログ5またはFCRH5）、胎児アセチルコリン受容体（胎児AchR）、葉酸結合タンパク質（FBP）、葉酸受容体α、ガングリオシドGD2、O-アセチル化GD2（OGD2）、ガングリオシドGD3、糖タンパク質100（gp100）、グリピカン-3（GPC3）、Gタンパク質共役受容体クラスCグループ5メンバーD（GPRC5D）、Her2/neu（受容体チロシンキナーゼerb-B2）、Her3（erb-B3）、Her4（erb-B4）、erbB二量体、ヒト高分子量黒色腫関連抗原（HMW-MAA）、B型肝炎表面抗原、ヒト白血球抗原A1（HLA-A1）、ヒト白血球抗原A2（HLA-A2）、IL-22受容体アルファ（IL-22Rα）、IL-13受容体アルファ2（IL-13Rα2）、キナーゼ挿入ドメイン受容体（kdr）、カッパ軽鎖、L1細胞接着分子（L1-CAM）、L1-CAMのCE7エピトープ、ロイシンリッチリピート含有8ファミリーメンバーA（Leucine Rich Repeat Containing 8 Family Member A：LRRC8A）、Lewis Y、黒色腫関連抗原（MAGE）-A1、MAGE-A3、MAGE-A6、MAGE-A10、メソテリン（MSLN）、c-Met、マウスサイトメガロウイルス（CMV）、ムチン1（MUC1）、MUC16、ナチュラルキラーグループ2メンバーD（NKG2D）リガンド、メラン（melan）A（MART-1）、神経細胞接着分子（NCAM）、腫瘍胎児抗原、メラノーマ優先発現抗原（PRAME）、プロゲステロン受容体、前立腺特異抗原、前立腺幹細胞抗原（PSCA）、前立腺特異的膜抗原（PSMA）、受容体チロシンキナーゼ様オーファン受容体1（ROR1）、サバイビン、トロホブラスト（Trophoblast）糖タンパク質（TPBG、別名5T4）、腫瘍関連糖タンパク質72（TAG72）、チロシナーゼ関連タンパク質1（TRP1、別名TYRP1またはgp75）、チロシナーゼ関連タンパク質2（TRP2、別名ドパクロムトートメラーゼ、ドパクロムデルタイソメラーゼ、またはDCT）、血管内皮細胞増殖因子受容体（VEGFR）、血管内皮細胞増殖因子受容体2（VEGFR2）、Wilms Tumor 1（WT-1）、病原体特異的もしくは病原体発現抗原、またはユニバーサルタグに関連する抗原、および/またはビオチン化分子、および/またはHIV、HCV、HBV、もしくは他の病原体によって発現される分子から選択される、本発明1065または本発明1066の方法。
[本発明1068]
前記組換え受容体が、組換えT細胞受容体（TCR）または組換え機能的非T細胞受容体である、本発明1064～1067のいずれかの方法。
[本発明1069]
前記組換え受容体がキメラ抗原受容体（CAR）である、本発明1064～1068のいずれかの方法。
[本発明1070]
前記CARが、前記抗原に特異的に結合する細胞外抗原認識ドメインと、ITAMを含む細胞内シグナル伝達ドメインとを含む、本発明1069の方法。
[本発明1071]
前記ITAMを含む細胞内シグナル伝達ドメインが、CD3-ゼータ（CD3ζ）鎖、任意でヒトCD3-ゼータ鎖の細胞内ドメインを含む、本発明1070の方法。
[本発明1072]
前記細胞内シグナル伝達ドメインが、共刺激シグナル伝達領域をさらに含む、本発明1070または本発明1071の方法。
[本発明1073]
前記共刺激シグナル伝達領域が、CD28または4-1BB、任意でヒトCD28またはヒト4-1BBのシグナル伝達ドメインを含む、本発明1072の方法。
[本発明1074]
以下の工程を含む、組み込まれていない残りの導入遺伝子配列を評価するための方法：
（1）DNAの高分子量画分における導入遺伝子配列の存在、不在、または量を決定するために、本発明1001～1015および1028～1073のいずれかの方法を行い、それによって、該導入遺伝子配列のゲノム組み込みを評価する工程；
（2）該高分子量画分の分離を伴わずに、単離されたDNAにおける該導入遺伝子配列の存在、不在、または量を決定する工程；
（3）（1）において決定された量と（2）において決定された量とを比較し、それによって、組み込まれていない残りの組換え配列の量を決定する工程。
[本発明1075]
前記導入遺伝子配列の存在、不在、または量を決定する工程が、前記1つまたは複数の細胞における二倍体ゲノム当たりのまたは細胞当たりの、該導入遺伝子配列の質量、重量、またはコピー数を決定することを含む、本発明1074の方法。
[本発明1076]
前記1つまたは複数の細胞が細胞の集団を含み、該集団のうちの複数の細胞が、前記組換えタンパク質をコードする前記導入遺伝子配列を含むか、またはそれを含む疑いがある、本発明1074または本発明1075の方法。
[本発明1077]
前記コピー数が、細胞の集団中での二倍体ゲノム当たりのまたは細胞当たりの平均（average）または平均（mean）コピー数である、本発明1075または本発明1076の方法。
[本発明1078]
前記量を比較する工程が、（2）において決定されたコピー数から、（1）において決定されたコピー数を減算することを含む、本発明1075～1077のいずれかの方法。
[本発明1079]
前記量を比較する工程が、（1）において決定されたコピー数対（2）において決定されたコピー数の比を決定することを含む、本発明1075～1077のいずれかの方法。
[本発明1080]
（2）における存在、不在、または量を決定する工程が、ポリメラーゼ連鎖反応（PCR）によって行われる、本発明1074～1079のいずれかの方法。
[本発明1081]
前記PCRが、定量的ポリメラーゼ連鎖反応（qPCR）、デジタルPCR、またはドロップレットデジタルPCRである、本発明1080の方法。
[本発明1082]
前記PCRがドロップレットデジタルPCRである、本発明1080または本発明1081の方法。
[本発明1083]
前記導入遺伝子配列の少なくとも一部分に相補的であるか、またはそれを特異的に増幅することができる1つまたは複数のプライマーを用いて、前記PCRが行われる、本発明1080～1082のいずれかの方法。
[本発明1084]
（2）における存在、不在、または量を決定する工程が、前記高分子量画分の分離を伴わずに、前記単離されたDNAにおける前記導入遺伝子配列の質量、重量、またはコピー数を評価すること、および、前記高分子量画分の分離を伴わずに、該質量、重量、またはコピー数を、前記単離されたDNAにおける参照遺伝子の質量、重量、もしくはコピー数に対して、または標準曲線に対して正規化することを含む、本発明1074～1083のいずれかの方法。
[本発明1085]
前記参照遺伝子がハウスキーピング遺伝子である、本発明1084の方法。
[本発明1086]
前記参照遺伝子が、アルブミンをコードする遺伝子（ALB）である、本発明1084または本発明1085の方法。
[本発明1087]
前記参照遺伝子が、リボヌクレアーゼPタンパク質サブユニットp30をコードする遺伝子（RPP30）である、本発明1084または本発明1085の方法。
[本発明1088]
前記単離されたDNAにおける参照遺伝子の質量、重量、またはコピー数を決定する工程が、該参照遺伝子の少なくとも一部分に相補的であるか、またはそれを特異的に増幅することができる1つまたは複数のプライマーを用いたPCRによって行われる、本発明1084～1087のいずれかの方法。
[本発明1089]
（1）における存在、不在、または量を決定する工程と、（2）における存在、不在、または量を決定する工程とが、同じプライマーまたは同じプライマーのセットを用いたポリメラーゼ連鎖反応（PCR）によって行われる、本発明1074～1088のいずれかの方法。
[本発明1090]
前記組み込まれていない残りの組換え配列が、ベクタープラスミド、直鎖状相補DNA（cDNA）、自己組み込み体（autointegrant）、または末端反復配列（LTR）サークルのうちの1つまたは複数を含む、本発明1074～1089のいずれかの方法。
In any of the several embodiments provided, the remaining unintegrated recombination sequence is the vector plasmid, linear complementary DNA (cDNA), autointegrant, or long terminal repeat (LTR) circle. contains one or more of
[Invention 1001]
A method for assessing genomic integration of a transgene sequence comprising the steps of:
(a) separating a high molecular weight fraction of deoxyribonucleic acid (DNA) greater than or greater than about 10 kilobases (kb) from DNA isolated from one or more cells; wherein said one or more cells comprise or are suspected of comprising at least one engineered cell comprising a transgene sequence encoding a recombinant protein;
(b) determining from said high molecular weight fraction the presence, absence or amount of transgene sequences integrated into the genome of said one or more cells.
[Invention 1002]
1002. The method of Invention 1001, wherein deoxyribonucleic acid (DNA) is isolated from said one or more cells prior to the separating step in (a).
[Invention 1003]
The step of determining the presence, absence, or amount of the transgene sequence in (b) comprises mass, weight, or copies of the transgene sequence per diploid genome in said one or more cells or per cell The method of invention 1001 or invention 1002 comprising determining the number.
[Invention 1004]
1003. The method of any of inventions 1001-1003, wherein said one or more cells comprises a population of cells, a plurality of cells of said population comprising said transgene sequence encoding said recombinant protein.
[Invention 1005]
The method of invention 1003 or invention 1004, wherein said copy number is the average or mean copy number per diploid genome or per cell in said population of cells.
[Invention 1006]
prior to the separating step in (a), a polynucleotide comprising a transgene sequence encoding a recombinant protein has been introduced into at least one engineered cell of said one or more cells; The method of any one of the inventions 1001-1005.
[Invention 1007]
said at least one engineered cell is at a temperature greater than 25°C, optionally at or about 37°C ± 2°C, for more than 96 hours after introduction of said polynucleotide comprising said transgene sequence; 1006. The method of the invention 1006, which is not incubated at °C.
[Invention 1008]
Said at least one engineered cell is at a temperature greater than 25°C, optionally at or about 37°C ± 2°C, for more than 72 hours after introduction of said polynucleotide comprising said transgene sequence. 1006. The method of the invention 1006, which is not incubated at °C.
[Invention 1009]
said at least one engineered cell is at a temperature greater than 25°C, optionally at or about 37°C ± 2°C, for more than 48 hours after introduction of said polynucleotide comprising said transgene sequence; 1006. The method of the invention 1006, which is not incubated at °C.
[Invention 1010]
1009. The method of any of inventions 1001-1009, wherein the one or more cells are cryopreserved prior to the step of isolating the high molecular weight fraction of DNA in (a).
[Invention 1011]
The method of any of inventions 1001-1010, wherein said one or more cells is a cell line.
[Invention 1012]
1011. The method of any of inventions 1001-1010, wherein said one or more cells are primary cells obtained from a sample from the subject.
[Invention 1013]
1013. The method of any of inventions 1001-1012, wherein said one or more cells are immune cells.
[Invention 1014]
1013. The method of the invention 1013, wherein said immune cells are T cells or NK cells.
[Invention 1015]
1015. The method of invention 1014, wherein said T cells are CD3+, CD4+, and/or CD8+ T cells.
[Invention 1016]
A method for evaluating a transgene sequence in a biological sample from a subject comprising the steps of:
(a) a high molecular weight fraction of deoxyribonucleic acid (DNA) greater than or greater than about 10 kilobases (kb) present in a biological sample from a subject; separating from DNA isolated from a plurality of cells, wherein the biological sample contains or contains at least one engineered cell containing a transgene sequence encoding a recombinant protein suspected, the process; and
(b) determining the presence, absence or amount of transgene sequences in all or a portion of said biological sample from said high molecular weight fraction;
[Invention 1017]
wherein the step of determining the presence, absence, or amount of transgene sequences in (b) comprises determining the mass, weight, or copy number of said transgene sequences in all or a portion of said biological sample. The method of invention 1016.
[Invention 1018]
The method of invention 1016 or invention 1017, wherein prior to the separating step, DNA is isolated from one or more cells present in said biological sample.
[Invention 1019]
The method of any of inventions 1016-1018, wherein said biological sample is obtained from a subject to whom a composition comprising said at least one engineered cell containing said transgene sequence was administered.
[Invention 1020]
The method of any of inventions 1016-1019, wherein said biological sample is a tissue or body fluid sample.
[Invention 1021]
1021. The method of invention 1020, wherein said biological sample is a tissue sample and said tissue is a tumor.
[Invention 1022]
The method of invention 1020 or invention 1021, wherein said tissue sample is a tumor biopsy.
[Invention 1023]
The method of invention 1020, wherein said biological sample is a bodily fluid sample, and said bodily fluid sample is a blood or serum sample.
[Invention 1024]
prior to the separating step in (a), a polynucleotide comprising said transgene sequence encoding said recombinant protein is introduced into said at least one engineered cell of said one or more cells; The method of any of Inventions 1016-1023.
[Invention 1025]
The method of any of inventions 1016-1024, wherein said one or more cells in said biological sample comprise immune cells.
[Invention 1026]
1025. The method of the invention 1025, wherein said immune cells are T cells or NK cells.
[Invention 1027]
1026. The method of invention 1026, wherein said T cells are CD3+, CD4+, and/or CD8+ T cells.
[Invention 1028]
The method of any of inventions 1001-1027, wherein said separating step is performed by pulsed field gel electrophoresis or size exclusion chromatography.
[Invention 1029]
The method of any of inventions 1001-1028, wherein said separating step is performed by pulsed-field gel electrophoresis.
[Invention 1030]
A method for assessing genomic integration of a transgene sequence comprising the steps of:
(a) A high molecular weight fraction of deoxyribonucleic acid (DNA) greater than or greater than about 10 kilobases (kb) was isolated from a population of cells by pulsed-field gel electrophoresis. wherein the population of cells comprises a plurality of engineered cells each containing or suspected of containing a transgene sequence encoding a recombinant protein; and
(b) the average per diploid genome or per cell of said transgene sequence integrated into the genome of said plurality of engineered cells of said population of cells from said high molecular weight fraction; Or determining the mean copy number.
[Invention 1031]
prior to the separating step in (a), a polynucleotide comprising said transgene sequence encoding said recombinant protein is introduced into at least one of said plurality of engineered cells of said population of cells; The method of the invention 1030.
[Invention 1032]
Said population of cells is at a temperature greater than 25°C, optionally at 37°C ± 2, for more than 96 hours after introduction of said polynucleotide comprising said transgene sequence into said at least one engineered cell. The method of invention 1031, which has not been incubated at or at about 37°C ± 2°C.
[Invention 1033]
Said population of cells is at a temperature greater than 25°C, optionally at 37°C ± 2, for more than 72 hours after introduction of said polynucleotide comprising said transgene sequence into said at least one engineered cell. The method of invention 1031, which has not been incubated at or at about 37°C ± 2°C.
[Invention 1034]
Said population of cells is at a temperature greater than 25°C, optionally at 37°C ± 2, for more than 48 hours after introduction of said polynucleotide comprising said transgene sequence into said at least one engineered cell. The method of invention 1031, which has not been incubated at or at about 37°C ± 2°C.
[Invention 1035]
The method of any of inventions 1030-1034, wherein said population of cells has been cryopreserved prior to the step of isolating the high molecular weight fraction of DNA in (a).
[Invention 1036]
1035. The method of any of inventions 1001-1035, wherein said high molecular weight fraction is greater than 15 kilobases (kb) or greater than about 15 kilobases (kb).
[Invention 1037]
1035. The method of any of inventions 1001-1035, wherein said high molecular weight fraction is greater than 17.5 kilobases (kb) or greater than about 17.5 kilobases (kb).
[Invention 1038]
1035. The method of any of inventions 1001-1035, wherein said high molecular weight fraction is greater than 20 kilobases (kb) or greater than about 20 kilobases (kb).
[Invention 1039]
1038. The method of any of inventions 1001-1038, wherein said transgene sequence comprises a regulatory element operably linked to said recombinant protein-encoding nucleic acid sequence.
[Invention 1040]
1039. The method of any of inventions 1001-1039, wherein determining the presence, absence or amount of said transgene sequence is performed by polymerase chain reaction (PCR).
[Invention 1041]
The method of invention 1040, wherein said PCR is quantitative polymerase chain reaction (qPCR), digital PCR, or droplet digital PCR.
[Invention 1042]
The method of invention 1040 or invention 1041, wherein said PCR is droplet digital PCR.
[Invention 1043]
1043. The method of any of invention 1040-1042, wherein said PCR is performed using one or more primers that are complementary to, or capable of specifically amplifying, at least a portion of said transgene sequence. Method.
[Invention 1044]
The method of invention 1043, wherein said one or more primers are complementary to or capable of specifically amplifying the sequence of the regulatory element.
[Invention 1045]
Determining the amount of said transgene sequence optionally comprises 1 micron of DNA isolated from said one or more cells per mass or weight of DNA isolated from said one or more cells. 1044. The method of any of inventions 1001-1044, comprising assessing the mass, weight, or copy number of said transgene sequence per gram.
[Invention 1046]
The method of the invention, wherein the step of determining the amount of transgene sequence comprises assessing the mass or weight in micrograms of transgene sequence per microgram of DNA isolated from one or more cells. 1045 ways.
[Invention 1047]
determining the amount of said transgene sequence per said one or more cells, optionally per CD3+, CD4+, and/or CD8+ cell and/or per cell expressing said recombinant protein. , the method of any of the inventions 1001-1044, comprising assessing the mass, weight, or copy number of said transgene sequence.
[Invention 1048]
Determining the presence, absence, or amount of the transgene sequence comprises assessing the mass, weight, or copy number of the transgene sequence per diploid genome or per cell in the biological sample. The method of any of the inventions 1016-1044, comprising:
[Invention 1049]
1048. The method of the invention 1048, wherein said copy number is the average or mean copy number per diploid genome or per cell in said one or more cells in said biological sample. .
[Invention 1050]
The step of determining the amount of the transgene sequence comprises: the mass, weight, or the method of any of the inventions 1016-1044, comprising assessing copy number.
[Invention 1051]
1044. The method of any of inventions 1016-1044, wherein determining the amount of the transgene sequence comprises assessing the mass, weight, or copy number of the transgene sequence per body weight or body surface area of the subject. .
[Invention 1052]
Determining the amount of the transgene sequence comprises assessing the mass, weight, or copy number of the transgene sequence in the high molecular weight fraction; The method of any of the inventions 1001-1048, comprising normalizing to the mass, weight or copy number of the reference gene in the fractions or to a standard curve.
[Invention 1053]
The method of invention 1052, wherein said reference gene is a housekeeping gene.
[Invention 1054]
The method of invention 1052 or invention 1053, wherein said reference gene is the gene encoding albumin (ALB).
[Invention 1055]
The method of invention 1052 or invention 1053, wherein said reference gene is the gene encoding ribonuclease P protein subunit p30 (RPP30).
[Invention 1056]
The copy number of the reference gene in said isolated DNA is performed by PCR using one or more primers that are complementary to or capable of specifically amplifying at least a portion of the reference gene. , the method of any of the inventions 1052-1055.
[Invention 1057]
1006. The method of any of inventions 1001-1006, wherein said transgene sequence does not encode the entire viral gag protein.
[Invention 1058]
1057. Any of the inventions 1001-1057, wherein said transgene sequence does not comprise a complete HIV genome, a replication competent viral genome, and/or accessory genes that are optionally Nef, Vpu, Vif, Vpr and/or Vpx. method.
[Invention 1059]
The method of any of the inventions 1006-1015, 1024-1029, and 1031-1057, wherein introducing said polynucleotide is by transducing a viral vector containing said polynucleotide.
[Invention 1060]
1059. The method of the invention 1059, wherein said viral vector is a retroviral vector or a gammaretroviral vector.
[Invention 1061]
The method of invention 1059 or invention 1060, wherein said viral vector is a lentiviral vector.
[Invention 1062]
1059. The method of the invention 1059, wherein said viral vector is an AAV vector, optionally selected among AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, or AAV8 vectors.
[Invention 1063]
The method of any of the invention 1006-1015, 1024-1029, and 1031-1057, wherein introduction of said polynucleotide is by physical delivery methods, optionally by electroporation.
[Invention 1064]
1063. The method of any of inventions 1001-1063, wherein said recombinant protein is a recombinant receptor.
[Invention 1065]
1064. The method of invention 1064, wherein said recombinant receptor specifically binds to an antigen associated with a disease or condition or expressed in cells in the environment of a lesion associated with a disease or condition.
[Invention 1066]
1065. The method of the invention 1065, wherein said disease or condition is cancer.
[Invention 1067]
said antigen is αvβ6 integrin (avb6 integrin), B cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (CA9, aka CAIX or G250), cancer testis antigen, cancer/testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24 , CD30, CD33, CD38, CD44, CD44v6, CD44v7/8, CD123, CD133, CD138, CD171, chondroitin sulfate proteoglycan 4 (CSPG4), epidermal growth factor protein (EGFR), type III epidermal growth factor receptor mutation (EGFR vIII ), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5; also known as Fc receptor homolog 5 or FCRH5), fetal acetylcholine receptor (fetal AchR), folate binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OGD2), ganglioside GD3, glycoprotein 100 (gp100), glypican- 3 (GPC3), G protein-coupled receptor class C group 5 member D (GPRC5D), Her2/neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer body, human high molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22 receptor alpha) 22Rα), IL-13 receptor alpha 2 (IL-13Rα2), kinase insertion domain receptor (kdr), kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, containing leucine-rich repeats 8 Family Member A (Leucine Rich Repeat Containing 8 Family Member A: LRRC8A), Lewis Y, melanoma-associated antigen (MAGE)-A1, MAGE-A3, MAGE-A6, MAGE-A10, mesothelin (MSLN), c-Met , mouse cytomegalovirus (CMV), mucin 1 (MUC1) , MUC16, natural killer group 2 member D (NKG2D) ligand, melan A (MART-1), neural cell adhesion molecule (NCAM), oncofetal antigen, melanoma preferentially expressed antigen (PRAME), progesterone receptor, prostate specific antigen, prostate stem cell antigen (PSCA), prostate specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), tyrosinase-related protein 1 (TRP1, also known as TYRP1 or gp75), tyrosinase-related protein 2 (TRP2, also known as dopachrome tautomerase, dopachrome delta isomerase, or DCT), vascular endothelial cell growth factor receptor vascular endothelial cell growth factor receptor 2 (VEGFR2), Wilms Tumor 1 (WT-1), pathogen-specific or pathogen-expressed antigens, or antigens associated with universal tags, and/or biotinylated molecules, and /or a method of invention 1065 or invention 1066 selected from molecules expressed by HIV, HCV, HBV, or other pathogens.
[Invention 1068]
1067. The method of any of inventions 1064-1067, wherein said recombinant receptor is a recombinant T cell receptor (TCR) or a recombinant functional non-T cell receptor.
[Invention 1069]
The method of any of inventions 1064-1068, wherein said recombinant receptor is a chimeric antigen receptor (CAR).
[Invention 1070]
1069. The method of invention 1069, wherein said CAR comprises an extracellular antigen recognition domain that specifically binds said antigen and an intracellular signaling domain comprising an ITAM.
[Invention 1071]
1070. The method of invention 1070, wherein said ITAM-comprising intracellular signaling domain comprises the intracellular domain of the CD3-zeta (CD3ζ) chain, optionally the human CD3-zeta chain.
[Invention 1072]
The method of invention 1070 or invention 1071, wherein said intracellular signaling domain further comprises a co-stimulatory signaling region.
[Invention 1073]
1073. The method of invention 1072, wherein said co-stimulatory signaling region comprises the signaling domain of CD28 or 4-1BB, optionally human CD28 or human 4-1BB.
[Invention 1074]
A method for evaluating residual unintegrated transgene sequences comprising the steps of:
(1) performing the method of any of the inventions 1001-1015 and 1028-1073 to determine the presence, absence, or amount of a transgene sequence in a high molecular weight fraction of DNA, thereby evaluating the genomic integration of
(2) determining the presence, absence, or amount of said transgene sequence in isolated DNA without separation of said high molecular weight fraction;
(3) comparing the amount determined in (1) with the amount determined in (2), thereby determining the amount of remaining unincorporated recombinant sequences.
[Invention 1075]
Determining the presence, absence or amount of said transgene sequence determines the mass, weight or copy number of said transgene sequence per diploid genome in said one or more cells or per cell. A method of the invention 1074 comprising:
[Invention 1076]
The present invention 1074 wherein said one or more cells comprises a population of cells, wherein a plurality of cells of said population contain or are suspected of containing said transgene sequence encoding said recombinant protein Or the method of the invention 1075.
[Invention 1077]
1075. The method of invention 1075 or invention 1076, wherein said copy number is the average or mean copy number per diploid genome or per cell in a population of cells.
[Invention 1078]
The method of any of inventions 1075-1077, wherein the step of comparing the amounts comprises subtracting the copy number determined in (1) from the copy number determined in (2).
[Invention 1079]
The method of any of inventions 1075-1077, wherein the step of comparing the amounts comprises determining the ratio of the copy number determined in (1) to the copy number determined in (2).
[Invention 1080]
The method of any of Inventions 1074-1079, wherein the step of determining the presence, absence or amount in (2) is performed by polymerase chain reaction (PCR).
[Invention 1081]
The method of the invention 1080, wherein said PCR is quantitative polymerase chain reaction (qPCR), digital PCR, or droplet digital PCR.
[Invention 1082]
The method of invention 1080 or invention 1081, wherein said PCR is droplet digital PCR.
[Invention 1083]
1082. The method of any of invention 1080-1082, wherein said PCR is performed using one or more primers complementary to or capable of specifically amplifying at least a portion of said transgene sequence. Method.
[Invention 1084]
The step of determining the presence, absence, or amount in (2) assesses the mass, weight, or copy number of the transgene sequence in the isolated DNA without separation of the high molecular weight fraction. and, without separation of said high molecular weight fraction, comparing said mass, weight or copy number against the mass, weight or copy number of a reference gene in said isolated DNA or a standard curve The method of any of the inventions 1074-1083, comprising normalizing against .
[Invention 1085]
1084. The method of invention 1084, wherein said reference gene is a housekeeping gene.
[Invention 1086]
The method of invention 1084 or invention 1085, wherein said reference gene is the gene encoding albumin (ALB).
[Invention 1087]
The method of invention 1084 or invention 1085, wherein said reference gene is the gene encoding ribonuclease P protein subunit p30 (RPP30).
[Invention 1088]
determining the mass, weight, or copy number of the reference gene in said isolated DNA is complementary to or capable of specifically amplifying at least a portion of said reference gene; or The method of any of inventions 1084-1087 performed by PCR using multiple primers.
[Invention 1089]
Determining the presence, absence, or amount in (1) and determining the presence, absence, or amount in (2) are performed by polymerase chain reaction (PCR) using the same primers or the same set of primers. The method of any of Inventions 1074-1088 performed.
[Invention 1090]
wherein said unintegrated residual recombination sequences comprise one or more of a vector plasmid, linear complementary DNA (cDNA), autointegrant, or long terminal repeat (LTR) circle. The method of any of Inventions 1074-1089.
Claims (48)
（a）10キロベース（kb）よりも大きいかまたは約10キロベース（kb）よりも大きいデオキシリボ核酸（DNA）の高分子量画分を、1つまたは複数の細胞より単離されたDNAから分離する工程であって、該1つまたは複数の細胞が、組換えタンパク質をコードする導入遺伝子配列を含む少なくとも1つの操作された細胞を含むか、またはそれを含む疑いがある、工程；
（b）該高分子量画分から、該1つまたは複数の細胞のゲノム中に組み込まれた導入遺伝子配列の存在、不在、または量を決定する工程。 A method for assessing genomic integration of a transgene sequence comprising the steps of:
(a) separating a high molecular weight fraction of deoxyribonucleic acid (DNA) greater than or greater than about 10 kilobases (kb) from DNA isolated from one or more cells; wherein said one or more cells comprise or are suspected of comprising at least one engineered cell comprising a transgene sequence encoding a recombinant protein;
(b) determining from said high molecular weight fraction the presence, absence or amount of transgene sequences integrated into the genome of said one or more cells.
前記1つまたは複数の細胞が、対象由来のサンプルから得られた初代細胞である、および／または
前記1つまたは複数の細胞が、免疫細胞である、
請求項1～7のいずれか一項に記載の方法。 said one or more cells is a cell line;
said one or more cells are primary cells obtained from a sample from a subject, and/or
said one or more cells are immune cells;
A method according to any one of claims 1-7 .
（a）10キロベース（kb）よりも大きいかまたは約10キロベース（kb）よりも大きいデオキシリボ核酸（DNA）の高分子量画分を、対象由来の生物学的サンプル中に存在する1つまたは複数の細胞より単離されたDNAから分離する工程であって、該生物学的サンプルが、組換えタンパク質をコードする導入遺伝子配列を含む少なくとも1つの操作された細胞を含むか、またはそれを含む疑いがある、工程； および
（b）該高分子量画分から、該生物学的サンプルの全てまたは一部分における導入遺伝子配列の存在、不在、または量を決定する工程。 A method for evaluating a transgene sequence in a biological sample from a subject comprising the steps of:
(a) a high molecular weight fraction of deoxyribonucleic acid (DNA) greater than or greater than about 10 kilobases (kb) present in a biological sample from a subject; separating from DNA isolated from a plurality of cells, wherein the biological sample contains or contains at least one engineered cell containing a transgene sequence encoding a recombinant protein and (b) determining from said high molecular weight fraction the presence, absence or amount of transgene sequences in all or a portion of said biological sample.
前記生物学的サンプルが組織サンプルであり、該組織が腫瘍である、
前記組織サンプルが腫瘍生検材料である、および／または
前記生物学的サンプルが体液サンプルであり、該体液サンプルが、血液または血清サンプルである、
請求項9～11のいずれか一項に記載の方法。 said biological sample is a tissue sample or a bodily fluid sample;
said biological sample is a tissue sample, said tissue is a tumor;
said tissue sample is a tumor biopsy, and/or
said biological sample is a bodily fluid sample, said bodily fluid sample is a blood or serum sample;
A method according to any one of claims 9-11 .
前記方法が、DNAを、前記生物学的サンプル中に存在する1つまたは複数の細胞から単離する工程を含むか、および／または
前記組換えタンパク質をコードする前記導入遺伝子配列を含むポリヌクレオチドが、前記1つまたは複数の細胞のうちの前記少なくとも1つの操作された細胞中に導入されている、
請求項9～12のいずれか一項に記載の方法。 Before the step of separating in (a),
said method comprises isolating DNA from one or more cells present in said biological sample; and/or
a polynucleotide comprising said transgene sequence encoding said recombinant protein has been introduced into said at least one engineered cell of said one or more cells;
A method according to any one of claims 9-12 .
（a）パルスフィールドゲル電気泳動によって、10キロベース（kb）よりも大きいかまたは約10キロベース（kb）よりも大きいデオキシリボ核酸（DNA）の高分子量画分を、細胞の集団より単離されたDNAから分離する工程であって、該細胞の集団が、組換えタンパク質をコードする導入遺伝子配列を各々が含むかまたはそれを含む疑いがある複数の操作された細胞を含む、工程；および
（b）該高分子量画分から、該細胞の集団のうちの該複数の操作された細胞のゲノム中に組み込まれた該導入遺伝子配列の二倍体ゲノム当たりのまたは細胞当たりの、平均（average）または平均（mean）コピー数を決定する工程。 A method for assessing genomic integration of a transgene sequence comprising the steps of:
(a) A high molecular weight fraction of deoxyribonucleic acid (DNA) greater than or greater than about 10 kilobases (kb) was isolated from a population of cells by pulsed-field gel electrophoresis. and ( b) the average per diploid genome or per cell of said transgene sequences integrated into the genomes of said plurality of engineered cells of said population of cells from said high molecular weight fraction or Determining the mean copy number.
前記導入遺伝子配列の量を決定する工程が、前記1つもしくは複数の細胞当たりの、任意で、CD3+、CD4+、および/もしくはCD8+細胞当たりの、ならびに/または前記組換えタンパク質を発現する細胞当たりの、該導入遺伝子配列の質量、重量、またはコピー数を評価することを含む、
請求項1～25のいずれか一項に記載の方法。 Determining the amount of said transgene sequence optionally comprises 1 micron of DNA isolated from said one or more cells per mass or weight of DNA isolated from said one or more cells. assessing the mass, weight, or copy number of the transgene sequence per gram , or
determining the amount of said transgene sequence per said one or more cells, optionally per CD3+, CD4+, and/or CD8+ cell and/or per cell expressing said recombinant protein. , assessing the mass, weight, or copy number of the transgene sequence;
A method according to any one of claims 1-25.
前記導入遺伝子配列の量を決定する工程が、前記生物学的サンプルの体積当たりの、任意で、前記生物学的サンプル1マイクロリットル当たりのまたは1ミリリットル当たりの、該導入遺伝子配列の質量、重量、またはコピー数を評価することを含むか、または
前記導入遺伝子配列の量を決定する工程が、対象の体重または体表面積当たりの、該導入遺伝子配列の質量、重量、またはコピー数を評価することを含む、
請求項9～16および21～25のいずれか一項に記載の方法。 Determining the presence, absence, or amount of the transgene sequence comprises assessing the mass, weight, or copy number of the transgene sequence per diploid genome or per cell in the biological sample. optionally , said copy number is the average or mean copy number per diploid genome or per cell in said one or more cells in said biological sample there is
The step of determining the amount of the transgene sequence comprises: the mass, weight, or weight of the transgene sequence per volume of the biological sample, optionally per microliter or per milliliter of the biological sample; or including assessing copy number , or
wherein determining the amount of the transgene sequence comprises assessing the mass, weight, or number of copies of the transgene sequence per body weight or body surface area of the subject;
A method according to any one of claims 9-16 and 21-25.
前記導入遺伝子配列が、完全なHIVゲノム、複製可能なウイルスゲノム、ならびに/または、任意でNef、Vpu、Vif、Vpr、および/もしくはVpxであるアクセサリー遺伝子を含まない、
請求項1～29のいずれか一項に記載の方法。 said transgene sequence does not encode the complete viral gag protein ; and/or
said transgene sequence does not comprise a complete HIV genome, a replication competent viral genome and/or accessory genes that are optionally Nef, Vpu, Vif, Vpr and/or Vpx;
A method according to any one of claims 1-29 .
前記ポリヌクレオチドの導入が、物理的送達法によって、任意でエレクトロポレーションによって行われる、
請求項5～8、13～16、および18～30のいずれか一項に記載の方法。 introduction of said polynucleotide is by transduction of a viral vector containing said polynucleotide , or
introduction of said polynucleotide is by a physical delivery method, optionally by electroporation;
The method of any one of claims 5-8, 13-16 and 18-30 .
前記組換え受容体が、組換えT細胞受容体（TCR）または機能的非T細胞受容体である、および／または
前記組換え受容体がキメラ抗原受容体（CAR）である、
請求項33に記載の方法。 said recombinant receptor specifically binds to an antigen associated with a disease or condition or expressed in cells in the environment of a lesion associated with a disease or condition;
said recombinant receptor is a recombinant T-cell receptor (TCR) or a functional non-T-cell receptor, and/or
wherein said recombinant receptor is a chimeric antigen receptor (CAR);
34. The method of claim 33 .
前記抗原が、αvβ6インテグリン（avb6インテグリン）、B細胞成熟抗原（BCMA）、B7-H3、B7-H6、炭酸脱水酵素9（CA9、別名CAIXまたはG250）、がん精巣抗原、がん/精巣抗原1B（CTAG、別名NY-ESO-1およびLAGE-2）、がん胎児性抗原（CEA）、サイクリン、サイクリンA2、C-Cモチーフケモカインリガンド1（CCL-1）、CD19、CD20、CD22、CD23、CD24、CD30、CD33、CD38、CD44、CD44v6、CD44v7/8、CD123、CD133、CD138、CD171、コンドロイチン硫酸プロテオグリカン4（CSPG4）、上皮成長因子タンパク質（EGFR）、タイプIII上皮成長因子受容体変異（EGFR vIII）、上皮糖タンパク質2（EPG-2）、上皮糖タンパク質40（EPG-40）、エフリンB2、エフリン受容体A2（EPHa2）、エストロゲン受容体、Fc受容体様5（FCRL5；別名Fc受容体ホモログ5またはFCRH5）、胎児アセチルコリン受容体（胎児AchR）、葉酸結合タンパク質（FBP）、葉酸受容体α、ガングリオシドGD2、O-アセチル化GD2（OGD2）、ガングリオシドGD3、糖タンパク質100（gp100）、グリピカン-3（GPC3）、Gタンパク質共役受容体クラスCグループ5メンバーD（GPRC5D）、Her2/neu（受容体チロシンキナーゼerb-B2）、Her3（erb-B3）、Her4（erb-B4）、erbB二量体、ヒト高分子量黒色腫関連抗原（HMW-MAA）、B型肝炎表面抗原、ヒト白血球抗原A1（HLA-A1）、ヒト白血球抗原A2（HLA-A2）、IL-22受容体アルファ（IL-22Rα）、IL-13受容体アルファ2（IL-13Rα2）、キナーゼ挿入ドメイン受容体（kdr）、カッパ軽鎖、L1細胞接着分子（L1-CAM）、L1-CAMのCE7エピトープ、ロイシンリッチリピート含有8ファミリーメンバーA（Leucine Rich Repeat Containing 8 Family Member A：LRRC8A）、Lewis Y、黒色腫関連抗原（MAGE）-A1、MAGE-A3、MAGE-A6、MAGE-A10、メソテリン（MSLN）、c-Met、マウスサイトメガロウイルス（CMV）、ムチン1（MUC1）、MUC16、ナチュラルキラーグループ2メンバーD（NKG2D）リガンド、メラン（melan）A（MART-1）、神経細胞接着分子（NCAM）、腫瘍胎児抗原、メラノーマ優先発現抗原（PRAME）、プロゲステロン受容体、前立腺特異抗原、前立腺幹細胞抗原（PSCA）、前立腺特異的膜抗原（PSMA）、受容体チロシンキナーゼ様オーファン受容体1（ROR1）、サバイビン、トロホブラスト（Trophoblast）糖タンパク質（TPBG、別名5T4）、腫瘍関連糖タンパク質72（TAG72）、チロシナーゼ関連タンパク質1（TRP1、別名TYRP1またはgp75）、チロシナーゼ関連タンパク質2（TRP2、別名ドパクロムトートメラーゼ、ドパクロムデルタイソメラーゼ、またはDCT）、血管内皮細胞増殖因子受容体（VEGFR）、血管内皮細胞増殖因子受容体2（VEGFR2）、Wilms Tumor 1（WT-1）、病原体特異的もしくは病原体発現抗原、またはユニバーサルタグに関連する抗原、および/またはビオチン化分子、および/またはHIV、HCV、HBV、もしくは他の病原体によって発現される分子から選択される、
請求項34に記載の方法。 said disease or condition is cancer, and/or
said antigen is αvβ6 integrin (avb6 integrin), B cell maturation antigen (BCMA), B7-H3, B7-H6, carbonic anhydrase 9 (CA9, aka CAIX or G250), cancer testis antigen, cancer/testis antigen 1B (CTAG, also known as NY-ESO-1 and LAGE-2), carcinoembryonic antigen (CEA), cyclin, cyclin A2, CC motif chemokine ligand 1 (CCL-1), CD19, CD20, CD22, CD23, CD24 , CD30, CD33, CD38, CD44, CD44v6, CD44v7/8, CD123, CD133, CD138, CD171, chondroitin sulfate proteoglycan 4 (CSPG4), epidermal growth factor protein (EGFR), type III epidermal growth factor receptor mutation (EGFR vIII ), epithelial glycoprotein 2 (EPG-2), epithelial glycoprotein 40 (EPG-40), ephrin B2, ephrin receptor A2 (EPHa2), estrogen receptor, Fc receptor-like 5 (FCRL5; also known as Fc receptor homolog 5 or FCRH5), fetal acetylcholine receptor (fetal AchR), folate binding protein (FBP), folate receptor alpha, ganglioside GD2, O-acetylated GD2 (OGD2), ganglioside GD3, glycoprotein 100 (gp100), glypican- 3 (GPC3), G protein-coupled receptor class C group 5 member D (GPRC5D), Her2/neu (receptor tyrosine kinase erb-B2), Her3 (erb-B3), Her4 (erb-B4), erbB dimer body, human high molecular weight melanoma-associated antigen (HMW-MAA), hepatitis B surface antigen, human leukocyte antigen A1 (HLA-A1), human leukocyte antigen A2 (HLA-A2), IL-22 receptor alpha (IL-22 receptor alpha) 22Rα), IL-13 receptor alpha 2 (IL-13Rα2), kinase insertion domain receptor (kdr), kappa light chain, L1 cell adhesion molecule (L1-CAM), CE7 epitope of L1-CAM, containing leucine-rich repeats 8 Family Member A (Leucine Rich Repeat Containing 8 Family Member A: LRRC8A), Lewis Y, melanoma-associated antigen (MAGE)-A1, MAGE-A3, MAGE-A6, MAGE-A10, mesothelin (MSLN), c-Met , mouse cytomegalovirus (CMV), mucin 1 (MUC1) , MUC16, natural killer group 2 member D (NKG2D) ligand, melan A (MART-1), neural cell adhesion molecule (NCAM), oncofetal antigen, melanoma preferentially expressed antigen (PRAME), progesterone receptor, prostate specific antigen, prostate stem cell antigen (PSCA), prostate specific membrane antigen (PSMA), receptor tyrosine kinase-like orphan receptor 1 (ROR1), survivin, trophoblast glycoprotein (TPBG, also known as 5T4), tumor-associated glycoprotein 72 (TAG72), tyrosinase-related protein 1 (TRP1, also known as TYRP1 or gp75), tyrosinase-related protein 2 (TRP2, also known as dopachrome tautomerase, dopachrome delta isomerase, or DCT), vascular endothelial cell growth factor receptor vascular endothelial cell growth factor receptor 2 (VEGFR2), Wilms Tumor 1 (WT-1), pathogen-specific or pathogen-expressed antigens, or antigens associated with universal tags, and/or biotinylated molecules, and /or selected from molecules expressed by HIV, HCV, HBV, or other pathogens;
35. The method of claim 34 .
前記ITAMを含む細胞内シグナル伝達ドメインが、CD3-ゼータ（CD3ζ）鎖、任意でヒトCD3-ゼータ鎖の細胞内ドメインを含む、
前記細胞内シグナル伝達ドメインが、共刺激シグナル伝達領域をさらに含む、および／または
前記共刺激シグナル伝達領域が、CD28または4-1BB、任意でヒトCD28またはヒト4-1BBのシグナル伝達ドメインを含む、
請求項34に記載の方法。 said CAR comprises an extracellular antigen recognition domain that specifically binds said antigen and an intracellular signaling domain comprising an ITAM;
wherein said ITAM-comprising intracellular signaling domain comprises a CD3-zeta (CD3ζ) chain, optionally the intracellular domain of the human CD3-zeta chain;
said intracellular signaling domain further comprises a co-stimulatory signaling region , and/or
said co-stimulatory signaling region comprises the signaling domain of CD28 or 4-1BB, optionally human CD28 or human 4-1BB;
35. The method of claim 34 .
（1）DNAの高分子量画分における導入遺伝子配列の存在、不在、または量を決定するために、請求項1～8および16～36のいずれか一項に記載の方法を行い、それによって、該導入遺伝子配列のゲノム組み込みを評価する工程；
（2）該高分子量画分の分離を伴わずに、単離されたDNAにおける該導入遺伝子配列の存在、不在、または量を決定する工程；
（3）（1）において決定された量と（2）において決定された量とを比較し、それによって、組み込まれていない残りの組換え配列の量を決定する工程。 A method for evaluating residual unintegrated transgene sequences comprising the steps of:
(1) performing the method of any one of claims 1-8 and 16-36 to determine the presence, absence or amount of a transgene sequence in a high molecular weight fraction of DNA, thereby assessing genomic integration of said transgene sequence;
(2) determining the presence, absence, or amount of said transgene sequence in isolated DNA without separation of said high molecular weight fraction;
(3) comparing the amount determined in (1) with the amount determined in (2), thereby determining the amount of remaining unincorporated recombinant sequences.
（2）において決定されたコピー数から、（1）において決定されたコピー数を減算すること、または
（1）において決定されたコピー数対（2）において決定されたコピー数の比を決定すること
を含む、請求項38または39に記載の方法。 Comparing the amounts comprises:
subtracting the copy number determined in (1) from the copy number determined in (2) , or
Determining the ratio of the copy number determined in (1) to the copy number determined in (2)
40. The method of claim 38 or 39 , comprising
前記参照遺伝子が、アルブミンをコードする遺伝子（ALB）である、および／または
前記参照遺伝子が、リボヌクレアーゼPタンパク質サブユニットp30をコードする遺伝子（RPP30）である、
請求項28～36および44のいずれか一項に記載の方法。 wherein said reference gene is a housekeeping gene;
said reference gene is the gene encoding albumin (ALB), and/or
wherein the reference gene is the gene encoding the ribonuclease P protein subunit p30 (RPP30);
The method of any one of claims 28-36 and 44 .
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Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115916223A (en) | 2020-04-10 | 2023-04-04 | 朱诺治疗学股份有限公司 | Methods and uses related to cell therapy engineered with chimeric antigen receptors targeting B cell maturation antigens |
| EP4138930A4 (en) * | 2020-04-24 | 2024-05-22 | Adoc Ssf Llc | Methods of determining gene editing efficiencies in cells |
| CN115852064A (en) * | 2020-09-30 | 2023-03-28 | 浙江大学 | Primer and probe for detecting T216C mutation site of HBV gene |
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Family Cites Families (80)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4452773A (en) | 1982-04-05 | 1984-06-05 | Canadian Patents And Development Limited | Magnetic iron-dextran microspheres |
| FR2540122B1 (en) | 1983-01-27 | 1985-11-29 | Centre Nat Rech Scient | NOVEL COMPOUNDS COMPRISING A SEQUENCE OF OLIGONUCLEOTIDE LINKED TO AN INTERCALATION AGENT, THEIR SYNTHESIS PROCESS AND THEIR APPLICATION |
| US4690915A (en) | 1985-08-08 | 1987-09-01 | The United States Of America As Represented By The Department Of Health And Human Services | Adoptive immunotherapy as a treatment modality in humans |
| US4795698A (en) | 1985-10-04 | 1989-01-03 | Immunicon Corporation | Magnetic-polymer particles |
| IN165717B (en) | 1986-08-07 | 1989-12-23 | Battelle Memorial Institute | |
| US5219740A (en) | 1987-02-13 | 1993-06-15 | Fred Hutchinson Cancer Research Center | Retroviral gene transfer into diploid fibroblasts for gene therapy |
| AU4746590A (en) | 1988-12-28 | 1990-08-01 | Stefan Miltenyi | Methods and materials for high gradient magnetic separation of biological materials |
| US5200084A (en) | 1990-09-26 | 1993-04-06 | Immunicon Corporation | Apparatus and methods for magnetic separation |
| CA2096222C (en) | 1990-11-13 | 1998-12-29 | Stephen D. Lupton | Bifunctional selectable fusion genes |
| DE4228458A1 (en) | 1992-08-27 | 1994-06-01 | Beiersdorf Ag | Multicistronic expression units and their use |
| EP0804590A1 (en) | 1993-05-21 | 1997-11-05 | Targeted Genetics Corporation | Bifunctional selectable fusion genes based on the cytosine deaminase (cd) gene |
| US5827642A (en) | 1994-08-31 | 1998-10-27 | Fred Hutchinson Cancer Research Center | Rapid expansion method ("REM") for in vitro propagation of T lymphocytes |
| WO1996013593A2 (en) | 1994-10-26 | 1996-05-09 | Procept, Inc. | Soluble single chain t cell receptors |
| WO1996018105A1 (en) | 1994-12-06 | 1996-06-13 | The President And Fellows Of Harvard College | Single chain t-cell receptor |
| US20020150914A1 (en) | 1995-06-30 | 2002-10-17 | Kobenhavns Universitet | Recombinant antibodies from a phage display library, directed against a peptide-MHC complex |
| US5801115A (en) | 1995-09-05 | 1998-09-01 | Kataleuna Gmbh | Catalyst composition and methods for using and preparing same |
| DE19608753C1 (en) | 1996-03-06 | 1997-06-26 | Medigene Gmbh | Transduction system based on rep-negative adeno-associated virus vector |
| WO1997034634A1 (en) | 1996-03-20 | 1997-09-25 | Sloan-Kettering Institute For Cancer Research | Single chain fv constructs of anti-ganglioside gd2 antibodies |
| AU6467796A (en) | 1996-04-24 | 1997-05-15 | Claude Fell | Cell separation system for biological fluids like blood |
| CA2305630C (en) | 1997-10-02 | 2012-11-06 | Sunol Molecular Corporation | Soluble single-chain t-cell receptor proteins |
| TR200003391T2 (en) | 1998-05-19 | 2001-02-21 | Avidex Limited | Soluble T cell receptor |
| JP2002524081A (en) | 1998-09-04 | 2002-08-06 | スローン − ケッタリング インスティチュート フォー キャンサー リサーチ | Fusion receptor specific for prostate-specific membrane antigen and uses thereof |
| AU2472400A (en) | 1998-10-20 | 2000-05-08 | City Of Hope | CD20-specific redirected T cells and their use in cellular immunotherapy of CD20+ malignancies |
| ES2224726T3 (en) | 1998-12-24 | 2005-03-01 | Biosafe S.A. | BLOOD SEPARATION SYSTEM INDICATED IN PARTICULAR FOR THE CONCENTRATION OF HEMATOPOYETIC MOTHER CELLS. |
| US20040191260A1 (en) | 2003-03-26 | 2004-09-30 | Technion Research & Development Foundation Ltd. | Compositions capable of specifically binding particular human antigen presenting molecule/pathogen-derived antigen complexes and uses thereof |
| CA2410510A1 (en) | 2000-06-02 | 2001-12-13 | Memorial Sloan-Kettering Cancer Center | Artificial antigen presenting cells and methods of use thereof |
| AU2001297703B2 (en) | 2000-11-07 | 2006-10-19 | City Of Hope | CD19-specific redirected immune cells |
| US7070995B2 (en) | 2001-04-11 | 2006-07-04 | City Of Hope | CE7-specific redirected immune cells |
| US20090257994A1 (en) | 2001-04-30 | 2009-10-15 | City Of Hope | Chimeric immunoreceptor useful in treating human cancers |
| PL208712B1 (en) | 2001-08-31 | 2011-05-31 | Avidex Ltd | Soluble t cell receptor |
| US7939059B2 (en) | 2001-12-10 | 2011-05-10 | California Institute Of Technology | Method for the generation of antigen-specific lymphocytes |
| US6992176B2 (en) | 2002-02-13 | 2006-01-31 | Technion Research & Development Foundation Ltd. | Antibody having a T-cell receptor-like specificity, yet higher affinity, and the use of same in the detection and treatment of cancer, viral infection and autoimmune disease |
| CA2476625A1 (en) | 2002-02-20 | 2003-08-28 | Dyax Corp. | Mhc-peptide complex binding ligands |
| US20030170238A1 (en) | 2002-03-07 | 2003-09-11 | Gruenberg Micheal L. | Re-activated T-cells for adoptive immunotherapy |
| US7446190B2 (en) | 2002-05-28 | 2008-11-04 | Sloan-Kettering Institute For Cancer Research | Nucleic acids encoding chimeric T cell receptors |
| AU2003271904B2 (en) | 2002-10-09 | 2009-03-05 | Adaptimmune Limited | Single chain recombinant T cell receptors |
| US20050129671A1 (en) | 2003-03-11 | 2005-06-16 | City Of Hope | Mammalian antigen-presenting T cells and bi-specific T cells |
| US20050025763A1 (en) | 2003-05-08 | 2005-02-03 | Protein Design Laboratories, Inc. | Therapeutic use of anti-CS1 antibodies |
| CA2555185C (en) | 2004-02-06 | 2020-03-24 | Morphosys Ag | Anti-cd38 human antibodies and uses therefor |
| US20090304679A1 (en) | 2004-05-27 | 2009-12-10 | Weidanz Jon A | Antibodies as T cell receptor mimics, methods of production and uses thereof |
| EP1773383B1 (en) | 2004-05-27 | 2012-09-12 | Jon A. Weidanz | Antibodies as t cell receptor mimics, methods of production and uses thereof |
| US20090226474A1 (en) | 2004-05-27 | 2009-09-10 | Weidanz Jon A | Antibodies as T cell receptor mimics, methods of production and uses thereof |
| JP5563194B2 (en) | 2004-06-29 | 2014-07-30 | イムノコア リミテッド | Cells expressing modified T cell receptors |
| KR101319135B1 (en) | 2005-03-23 | 2013-10-17 | 바이오세이프 쏘시에떼아노님 | Integrated System for Collecting, Processing and Transplanting Cell Subsets, including Adult Stem Cells, for Regenerative Medicine |
| SI2567976T1 (en) | 2005-03-23 | 2017-11-30 | Genmab A/S | Antibodies against CD38 for treatment of multiple myeloma |
| EP1914242A1 (en) | 2006-10-19 | 2008-04-23 | Sanofi-Aventis | Novel anti-CD38 antibodies for the treatment of cancer |
| WO2008121420A1 (en) | 2007-03-30 | 2008-10-09 | Memorial Sloan-Kettering Cancer Center | Constitutive expression of costimulatory ligands on adoptively transferred t lymphocytes |
| ES2640216T3 (en) | 2007-12-07 | 2017-11-02 | Miltenyi Biotec Gmbh | Systems and methods for cell processing |
| US8479118B2 (en) | 2007-12-10 | 2013-07-02 | Microsoft Corporation | Switching search providers within a browser search box |
| PL2801584T3 (en) | 2007-12-26 | 2019-12-31 | Biotest Ag | Agents targeting CD138 and uses thereof |
| JP5173594B2 (en) | 2008-05-27 | 2013-04-03 | キヤノン株式会社 | Management apparatus, image forming apparatus, and processing method thereof |
| ES2961498T3 (en) | 2008-08-26 | 2024-03-12 | Hope City | Method and compositions for enhanced performance of anti-tumor effect of T cells |
| WO2010051391A1 (en) | 2008-10-31 | 2010-05-06 | Facet Biotech Corporation | Use of anti-cs1 antibodies for treatment of rare lymphomas |
| PT2406284T (en) | 2009-03-10 | 2016-09-29 | Biogen Ma Inc | Anti-bcma antibodies |
| RU2587621C2 (en) | 2009-04-01 | 2016-06-20 | Дженентек, Инк. | ANTI-FcRH5 ANTIBODIES, IMMUNOCONJUGATES THEREOF AND METHODS FOR USE THEREOF |
| CA2777053A1 (en) | 2009-10-06 | 2011-04-14 | The Board Of Trustees Of The University Of Illinois | Human single-chain t cell receptors |
| JP5956342B2 (en) | 2009-11-03 | 2016-07-27 | シティ・オブ・ホープＣｉｔｙ ｏｆ Ｈｏｐｅ | Truncated epidermal growth factor receptor (EGFRt) for transduction T cell selection |
| BR122021026169B1 (en) | 2010-12-09 | 2023-12-12 | The Trustees Of The University Of Pennsylvania | USE OF A CELL |
| JOP20210044A1 (en) | 2010-12-30 | 2017-06-16 | Takeda Pharmaceuticals Co | Anti-cd38 antibodies |
| ES2841983T3 (en) | 2011-03-23 | 2021-07-12 | Hutchinson Fred Cancer Res | Method and compositions for cellular immunotherapy |
| KR20140033029A (en) | 2011-04-01 | 2014-03-17 | 메모리얼 슬로안-케터링 캔서 센터 | T cell receptor-like antibodies specific for a wt1 peptide presented by hla-a2 |
| US8398282B2 (en) | 2011-05-12 | 2013-03-19 | Delphi Technologies, Inc. | Vehicle front lighting assembly and systems having a variable tint electrowetting element |
| BR112014011417B1 (en) | 2011-11-11 | 2021-10-13 | Fred Hutchinson Cancer Research Center | ISOLATED POLYPEPTIDE CAPABLE OF ELIGITING AN ANTIGEN-SPECIFIC T-CELL RESPONSE TO HUMAN CYCLIN A1 IMMUNOGENIC COMPOSITION INCLUDING SUCH POLYPEPTIDE, AS WELL AS A METHOD TO PREPARE CELLS PRESENTING SUPER ANTIGEN, ANTIGEN, ANTIGEN-USED ANTIGEN-USED CELLS |
| EP2814846B1 (en) | 2012-02-13 | 2020-01-08 | Seattle Children's Hospital d/b/a Seattle Children's Research Institute | Bispecific chimeric antigen receptors and therapeutic uses thereof |
| WO2013126726A1 (en) | 2012-02-22 | 2013-08-29 | The Trustees Of The University Of Pennsylvania | Double transgenic t cells comprising a car and a tcr and their methods of use |
| KR20150009556A (en) | 2012-05-03 | 2015-01-26 | 프레드 헛친슨 켄서 리서치 센터 | Enhanced affinity t cell receptors and methods for making the same |
| WO2014031687A1 (en) | 2012-08-20 | 2014-02-27 | Jensen, Michael | Method and compositions for cellular immunotherapy |
| CN112430580A (en) | 2012-10-02 | 2021-03-02 | 纪念斯隆-凯特琳癌症中心 | Compositions and methods for immunotherapy |
| CA2887341C (en) | 2012-10-12 | 2021-03-16 | Sage Science, Inc. | Side-eluting molecular fractionator |
| TWI654206B (en) | 2013-03-16 | 2019-03-21 | 諾華公司 | Treatment of cancer with a humanized anti-CD19 chimeric antigen receptor |
| AR096687A1 (en) | 2013-06-24 | 2016-01-27 | Genentech Inc | ANTI-FCRH5 ANTIBODIES |
| KR20220136455A (en) | 2014-04-23 | 2022-10-07 | 주노 쎄러퓨티크스 인코퍼레이티드 | Methods for isolating, culturing, and genetically engineering immune cell populations for adoptive therapy |
| US10301370B2 (en) | 2014-05-02 | 2019-05-28 | The Trustees Of The University Of Pennsylvania | Compositions and methods of chimeric autoantibody receptor T cells |
| CN107109487B (en) | 2014-10-15 | 2022-03-04 | 赛琪科学股份有限公司 | Apparatus, method and system for automated processing of nucleic acids and electrophoretic sample preparation |
| AU2015343121B2 (en) | 2014-11-05 | 2020-06-18 | Juno Therapeutics, Inc. | Methods for transduction and cell processing |
| RS61781B1 (en) | 2014-12-05 | 2021-06-30 | Memorial Sloan Kettering Cancer Center | Antibodies targeting b-cell maturation antigen and methods of use |
| SG11201704549UA (en) | 2014-12-05 | 2017-07-28 | Memorial Sloan Kettering Cancer Center | Chimeric antigen receptors targeting b-cell maturation antigen and uses thereof |
| AU2015357535B2 (en) | 2014-12-05 | 2020-05-14 | Eureka Therapeutics, Inc. | Antibodies targeting G-protein coupled receptor and methods of use |
| SI3227339T1 (en) | 2014-12-05 | 2022-02-28 | Memorial Sloan-Kettering Cancer Center | Chimeric antigen receptors targeting g-protein coupled receptor and uses thereof |
| KR102584280B1 (en) | 2016-04-01 | 2023-10-04 | 카이트 파마 인코포레이티드 | Chimeric antigen and t cell receptors and methods of use |
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2019
- 2019-08-09 EP EP19770209.5A patent/EP3833759A1/en active Pending
- 2019-08-09 CA CA3108698A patent/CA3108698A1/en active Pending
- 2019-08-09 WO PCT/US2019/046048 patent/WO2020033916A1/en active Application Filing
- 2019-08-09 MX MX2021001519A patent/MX2021001519A/en unknown
- 2019-08-09 SG SG11202101130VA patent/SG11202101130VA/en unknown
- 2019-08-09 EA EA202190469A patent/EA202190469A1/en unknown
- 2019-08-09 BR BR112021002245-7A patent/BR112021002245A2/en unknown
- 2019-08-09 US US17/266,995 patent/US20210230671A1/en active Pending
- 2019-08-09 AU AU2019316647A patent/AU2019316647A1/en active Pending
- 2019-08-09 JP JP2021506661A patent/JP2021533747A/en active Pending
- 2019-08-09 CN CN201980064962.4A patent/CN112805378A/en active Pending
- 2019-08-09 KR KR1020217006943A patent/KR20210059715A/en active Search and Examination
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2021
- 2021-01-31 IL IL280528A patent/IL280528A/en unknown
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