Large-scale manufacturing and characterization of CMV-CD19CAR T cells.
Large-scale manufacturing and characterization of CMV-CD19CAR T cells.
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DOI：
10.1136/jitc-2021-003461
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发表时间：
2022-01
影响因子：
10.9
通讯作者：
Nakamura R
中科院分区：
文献类型：
--
作者：
Wang X;Urak R;Walter M;Guan M;Han T;Vyas V;Chien SH;Gittins B;Clark MC;Mokhtari S;Cardoso A;Diamond DJ;Zaia J;Forman SJ;Nakamura R
cd19特异性嵌合抗原受体（CD19CAR） T细胞过继转移可诱导B细胞恶性肿瘤患者的疾病急剧消退。CD19CAR - T细胞治疗可能由于植入不足和持续存在而受到限制，从而导致肿瘤复发。我们之前证明了一个原理证明，在CD19CAR转导之前，巨细胞病毒（CMV）特异性T细胞可以被分离和富集以产生CMV-CD19CAR T细胞，并且这些CMV-CD19CAR T细胞可以通过CMV疫苗在体内扩增，从而在小鼠模型中更好地控制肿瘤。在这里，我们开发了一个生成CMV-CD19CAR - T细胞的临床平台。在CliniMACS Prodigy细胞因子捕获系统（Miltenyi Biotec）中，用良好生产规范级PepTivator重叠CMVpp65肽库刺激从cmv血清阳性健康供者收集的外周血单个核细胞（PBMCs），并使用IFNγ捕集基质富集cmv应答干扰素γ (IFNγ)+T细胞。用慢病毒载体编码第二代CD19R:CD28:ζ/EGFRt CAR转导cmv特异性T细胞，并用白细胞介素2 （IL-2）和IL-15扩增15天。在9次全面鉴定中，cmv特异性T细胞从输入PBMC的0.8%±0.5富集到76.3%±11.6（输入1×109 PBMC的IFNγ+T细胞的绝对产量为4.2±3.3×106）。CMV特异性T细胞的平均CD19CAR转导效率在最终产物中为27.0%±14.2，经过快速扩增，导致具有CMV特异性（即功能双特异性）的CD19CAR转导T细胞的总细胞剂量为6.2±0.9 × 106。CMV-CD19CAR - T细胞是一种多克隆细胞，表达记忆标记物，但衰竭标记物的表达较低，对CD19和CMVpp65刺激均有快速增殖反应，对表达CD19的肿瘤和CMVpp65抗原均表现出抗原特异性效应功能。最终产品通过了临床使用的放行标准。我们证明了我们的大规模平台产生CMV-CD19CAR T细胞用于临床应用的可行性。我们计划在希望之城启动CMV- cd19car -T细胞的临床试验，在自体造血细胞移植后立即接种基于改性安卡拉牛痘（Triplex）的新型巨细胞病毒疫苗28天和56天后注射T细胞，用于治疗中高级别B细胞非霍奇金淋巴瘤患者。
Adoptive transfer of CD19-specific chimeric antigen receptor (CD19CAR) T cells can induce dramatic disease regression in patients with B cell malignancies. CD19CAR T cell therapy may be limited by insufficient engraftment and persistence, resulting in tumor relapse. We previously demonstrated a proof of principle that cytomegalovirus (CMV)-specific T cells can be isolated and enriched prior to CD19CAR transduction to produce CMV-CD19CAR T cells, and that these CMV-CD19CAR T cells can be expanded in vivo through CMV vaccination, resulting in better tumor control in a murine model. Here we developed a clinical platform for generating CMV-CD19CAR T cells. Peripheral blood mononuclear cells (PBMCs) collected from CMV-seropositive healthy donors were stimulated with a good manufacturing practices-grade PepTivator overlapping CMVpp65 peptide pool and enriched for CMV-responsive interferon γ (IFNγ)+T cells using IFNγ Catchmatrix, within the CliniMACS Prodigy Cytokine Capture System (Miltenyi Biotec). Resulting CMV-specific T cells were transduced with a lentiviral vector encoding a second generation CD19R:CD28:ζ/EGFRt CAR and expanded with interleukin 2 (IL-2) and IL-15 for 15 days before characterization. CMV-specific T cells were enriched from 0.8%±0.5 of input PBMC to 76.3%±11.6 in nine full-scale qualification runs (absolute yield of 4.2±3.3×106 IFNγ+T cells from an input of 1×109 PBMCs). Average CD19CAR transduction efficiency of CMV-specific T cells was 27.0%±14.2 in the final products, which underwent rapid expansion, resulting in a total cell dose of 6.2±0.9 × 106 CD19CAR-tranduced T cells with CMV specificity (ie, functionally bispecific). CMV-CD19CAR T cells were polyclonal, expressed memory markers but had low expression of exhaustion markers, responded to both CD19 and CMVpp65 stimulation with rapid proliferation and exhibited antigen-specific effector functions against both CD19-expressing tumors and CMVpp65 antigen. The final products passed release criteria for clinical use. We demonstrated the feasibility of our large-scale platform for generating CMV-CD19CAR T cells for clinical application. We plan to initiate a clinical trial at City of Hope using CMV-CD19CAR T cells for patients with intermediate/high-grade B cell non-Hodgkin’s lymphoma immediately after autologous hematopoietic cell transplantation followed by vaccination with a novel CMV vaccine based on Modified Vaccinia Ankara (Triplex) 28 days and 56 days post-T cell infusion.
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影响因子：
20.3
作者：
Rossig, C;Bollard, CM;Brenner, MK
通讯作者：
Brenner, MK
病毒特异性的T细胞设计为共表达肿瘤特异性受体：神经母细胞瘤个体中的持久性和抗肿瘤活性。
影响因子：
82.9
作者：
通讯作者：
--
DOI：
10.1056/nejmoa1707447
发表时间：
2017-12-28
期刊：
The New England journal of medicine
影响因子：
--
作者：
Neelapu SS;Locke FL;Bartlett NL;Lekakis LJ;Miklos DB;Jacobson CA;Braunschweig I;Oluwole OO;Siddiqi T;Lin Y;Timmerman JM;Stiff PJ;Friedberg JW;Flinn IW;Goy A;Hill BT;Smith MR;Deol A;Farooq U;McSweeney P;Munoz J;Avivi I;Castro JE;Westin JR;Chavez JC;Ghobadi A;Komanduri KV;Levy R;Jacobsen ED;Witzig TE;Reagan P;Bot A;Rossi J;Navale L;Jiang Y;Aycock J;Elias M;Chang D;Wiezorek J;Go WY
通讯作者：
Go WY
DOI：
10.1038/nrc3322
发表时间：
2012-10
期刊：
Nature reviews. Cancer
影响因子：
--
作者：
Gattinoni L;Klebanoff CA;Restifo NP
通讯作者：
Restifo NP
影响因子：
39.2
作者：
Aldoss, Ibrahim;La Rosa, Corinna;Baden, Lindsey R.;Longmate, Jeffrey;Ariza-Heredia, Ella J.;Rida, Wasima N.;Lingaraju, Chetan Raj;Zhou, Qiao;Martinez, Joy;Kaltcheva, Teodora;Dagis, Andy;Hardwick, Nicola;Issa, Nicolas C.;Farol, Len;Nademanee, Auayporn;Al Malki, Monzr M.;Forman, Stephen;Nakamura, Ryotaro;Diamond, Don J.
通讯作者：
Diamond, Don J.