scholarly journals First immunotherapeutic CAR-T cells against the immune checkpoint protein HLA-G

2021 ◽  
Vol 9 (3) ◽  
pp. e001998
Author(s):  
François Anna ◽  
Elodie Bole-Richard ◽  
Joel LeMaoult ◽  
Marie Escande ◽  
Martin Lecomte ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Immune Escape ◽  
Lymphoblastic Leukemia ◽  
Specific Antigen ◽  
Immune Checkpoints ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T

BackgroundCAR-T cells immunotherapy is a breakthrough in the treatment of hematological malignancies such as acute lymphoblastic leukemia (ALL) and B-cell malignancies. However, CAR-T therapies face major hurdles such as the lack of tumor-specific antigen (TSA), and immunosuppressive tumor microenvironment sometimes caused by the tumorous expression of immune checkpoints (ICPs) such as HLA-G. Indeed, HLA-G is remarkable because it is both a potent ICP and a TSA. HLA-G tumor expression causes immune escape by impairing innate and adaptive immune responses and by inducing a suppressive microenvironment. Yet, to date, no immunotherapy targets it.MethodsWe have developed two anti-HLA-G third-generation CARs based on new anti-HLA-G monoclonal antibodies.ResultsAnti-HLA-G CAR-T cells were specific for immunosuppressive HLA-G isoforms. HLA-G-activated CAR-T cells polarized toward T helper 1, and became cytotoxic against HLA-G+ tumor cells. In vivo, anti-HLA-G CAR-T cells were able to control and eliminate HLA-G+ tumor cells. The interaction of tumor-HLA-G with interleukin (IL)T2-expressing T cells is known to result in effector T cell functional inhibition, but anti-HLA-G CAR-T cells were insensitive to this inhibition and still exerted their function even when expressing ILT2. Lastly, we show that anti-HLA-G CAR-T cells differentiated into long-term memory effector cells, and seemed not to lose function even after repeated stimulation by HLA-G-expressing tumor cells.ConclusionWe report for the first time that HLA-G, which is both a TSA and an ICP, constitutes a valid target for CAR-T cell therapy to specifically target and eliminate both tumor cells and HLA-G+ suppressive cells.

scholarly journals Development of Novel Anti-CD10 Target Modules for Redirection of Universal CAR T Cells Against CD10-Positive Malignancies

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 5612-5612 ◽  
Author(s):  
Anja Feldmann ◽  
Stefanie Koristka ◽  
Claudia Arndt ◽  
Liliana Raquel Loureiro ◽  
Ralf Bergmann ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Hematological Malignancies ◽  
Lymphoblastic Leukemia ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T Cell Therapy ◽  
Car T

The common acute lymphoblastic leukemia antigen CD10 is a marker for several hematological malignancies, including acute lymphoblastic leukemia as well as T and B cell lymphomas, Burkitt lymphomas, and some solid tumors like renal cell carcinomas, pancreatic tumors and melanomas. Because of its tumor related expression pattern, CD10 is an attractive target for adoptively transferred T cells that are genetically modified to express chimeric antigen receptors (CARs). Recently, conventional CAR T cell therapy targeting CD19-positive hematological malignancies was clinically approved because of its impressive effectiveness in patients. However, CAR T cells can also cause severe side effects like on-target, off-tumor reactions, tumor lysis syndrome and cytokine release syndrome. Most critically, activity of conventional CAR T cells cannot be controlled, once they are applied in patients. As CD10 is also widely expressed on normal tissues, CAR T cell reactivity has to be controllable in order to stop CAR T cell therapy in case of on-target, off-tumor toxicities occur. Especially for this purpose, we have recently established a switchable, modular and universal CAR platform technology, named UniCAR system, which can be repeatedly turned on and off. In contrast to conventional CARs, that directly recognize a tumor-associated antigen (TAA) on the tumor cell surface via their extracellular single-chain variable fragment (scFv), the UniCAR system is structured in a modular manner of two components. The first component are T cells genetically engineered to express UniCARs and the second component are target modules (TMs). Most importantly, UniCARs cannot directly bind to a TAA because their extracellular scFv is directed against the peptide epitope E5B9 which is not present on the surface of living cells. Consequently, UniCAR armed T cells are per se inert. They can be redirected towards tumor cells only via a TM. TMs consist of a scFv targeting a TAA and the epitope E5B9 recognized by UniCARs allowing a cross-linkage of UniCAR T cells with tumor cells which results in T cell activation. As TMs have a very short half-life, UniCAR T cell activity can be controlled by dosing of the TM. Once the TM is administered, UniCAR T cells can be switched on, but once the TM injection is stopped and the TM is eliminated, UniCAR T cells are switched off immediately. Here, we show proof of concept for functionality of the UniCAR system targeting CD10-positive malignancies. Therefor, a novel anti-CD10 TM was constructed which is able to redirect UniCAR T cells to eliminate CD10-expressing tumor cells. In summary, we have established a universal, switchable, modular UniCAR platform technology that can be used to target CD10-positive malignancies. Disclosures Koristka: Intellia Therapeutics: Employment. Bachmann:GEMoaB Monoclonals: Equity Ownership, Patents & Royalties.

CAR T cells targeting CD99 as an approach to eradicate T-cell Acute Lymphoblastic Leukemia without normal blood cells toxicity

2021 ◽  
Author(s):  
Jiangzhou Shi ◽  
Zijian Zhang ◽  
Hong Cen ◽  
Han Wu ◽  
Shangkun Zhang ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Bone Marrow Cells ◽  
Clinical Success ◽  
Lymphoblastic Leukemia ◽  
Leukemia Cells ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T ◽  
Pilot Clinical Study

Abstract CAR T cell therapy has shown dramatic clinical success in relapsed or refractory (r/r) B-ALL and other haematological malignancies. However, the loss of specific antigens, cell fratricide, T cell aplasia, and normal T cell separation are challenges in treating T cell leukemia/lymphoma with CAR T therapy. CD99 is a promising antigen to target T-ALL and AML as it is expressed on the majority of T-ALL and AML. Here, we isolated a low-affinity CD99 (12E7) antibody, which specifically recognizes leukemia cells over normal bone marrow cells. T cells transduced with an anti-CD99-specific CAR that contained the 12E7 scFv expanded with minor fratricide, maintained their cytotoxic function and mediated powerful antitumour effects. Subsequently, we conducted a pilot clinical study to evaluate the safety and feasibility of therapy with anti-CD99 CAR T cells in 4 patients with r/r T-LBL (n=1), AML (n=2) or myeloid sarcoma (MS) (n=1). The clinical overall response rate (ORR) was 50% (2/4 patients), and 1 patients (25%) achieved complete remission (CR) for 2 month. Mild cytokine release syndrome (CRS) occurred in 2 patients and the CRS no more than grade 2. Together, our results demonstrate that anti-CD99 CAR T cells specifically recognize and efficiently eliminate CD99+ leukemia cells.

CAR T-Cells for Glioblastoma: Current Concepts, Challenges and Future Perspectives

Neurology ◽  
2021 ◽  
pp. 10.1212/WNL.0000000000012193
Author(s):  
P. Karschnia ◽  
N. Teske ◽  
N. Thon ◽  
M. Subklewe ◽  
JC. Tonn ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Therapy ◽  
Immune Escape ◽  
Basic Structure ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T Cell Therapy ◽  
Car T

Glioblastoma is the most common malignant primary brain tumor and associated with a poor prognosis even after multimodal therapy. Chimeric antigen receptor (CAR) T-cells have emerged as a promising therapeutic avenue in glioblastoma. CARs incorporate antigen-recognition moieties that endow autologous T-cells with specificity against antigens expressed on glioblastoma (e.g. IL-13Rα2, EGFRvIII, and HER2). Compelling anti-tumor effects of such therapy have been shown in murine glioblastoma models. In humans, five phase I/II studies on IL-13Rα2-, EGFRvIII-, and HER2-directed CAR T-cells for the treatment of glioblastoma patients have been published suggesting an acceptable safety profile. However, anti-tumor effects fell short of expectations in these initial clinical studies. Tumor heterogeneity, antigen loss, and the immunosuppressive tumor microenvironment are among the most important factors to limit the efficacy of CAR T-cell therapy in glioblastoma. Novel target antigens, modification of CAR T-cell design, the combination of CAR T-cell therapy with other therapeutic approaches, but also the use of CAR NK-cells or CAR macrophages may optimize anti-tumor effects. Numerous clinical trials studying such approaches are ongoing, as well as several preclinical studies. With an increasing understanding of immune-escape mechanisms of glioblastoma and novel manufacturing techniques for CARs, CAR T-cells may provide clinically relevant activity in glioblastoma. This review focuses on the use of CAR T-cells in glioblastoma, but also introduces the basic structure, mechanisms of action, and relevant side effects of CAR T-cells.

scholarly journals Sequential Infusion of Anti-CD22 and Anti-CD19 Chimeric Antigen Receptor T Cells for Adult Patients with Refractory/Relapsed B-Cell Acute Lymphoblastic Leukemia

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 846-846
Author(s):  
Liang Huang ◽  
Na Wang ◽  
Chunrui Li ◽  
Yang Cao ◽  
Yi Xiao ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Therapy ◽  
Lymphoblastic Leukemia ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T Cell Therapy ◽  
Car T ◽  
Antigen Loss

Abstract Clinical trials of second generation chimeric antigen receptor engineered T cells (CAR-T cells) have yielded unprecedented efficacy in refractory/relapsed B-cell acute lymphoblastic leukemia (B-ALL), especially in children and young adult. However, antigen loss relapse has been observed in approximately 14% of patients in anti-CD19 CAR-T cell therapy across institutions, which emerges as a challenge for the long-term disease control of this promising immunotherapy. Recently, CD19/CD20 and CD19/CD22 dual antigen targeting have been proposed to overcome antigen loss relapse after the administration of anti-CD19 CAR-T cells. This strategy may result in enhanced anti-tumor activity, while safety concern regarding the risk of cytokine release syndrome (CRS) due to significant CAR-T cell activation and cytokine release needs to be addressed. Here, we conducted an open-label, single-center and single-arm pilot study of sequential infusion of anti-CD22 and anti-CD19 CAR-T cells. We aimed to evaluate its safety and efficacy in adult patients with refractory or relapsed B-ALL. This trial is registered with ChiCTR, number ChiCTR-OPN-16008526. Between March 2016 and March 2017, 27 patients with refractory or relapsed B-ALL were enrolled in this clinical trial, with a median age of 30±12 years (range, 18-62 years). Thirteen patients (48.1%) had a history of at least two prior relapsed or primary refractory disease. Twenty-six patients received fludarabine and cyclophosphamide before the infusion of CAR-T cells. The median cell dosages of anti-CD22 and anti-CD19 CAR-T cells were 2.44 ± 1.02 × 106 /kg and 1.98 ± 1.05 × 106 /kg, respectively. 24/29 (88.9%) patients achieved CR or Cri, including 7 patients who received prior hematopoietic stem cell transplantation, and 13/27 (48.1%) patients achieved minimal residual disease negative (MRD-) CR accessed by flow cytometry. Sustained remission was achieved with a 6-month overall survival rate of 79% (95% CI, 66-97) and an event-free survival rate of 72% (95% CI, 55-95). 24/29 (88.9%) patients experienced CRS and 6/27 (22.2%) patients had reversible sever CRS (grade 3-4). And 3/27 (11.1%) patients developed neurotoxicity. Multi-color flow cytometry was used to screen and quantitate MRD in blood, bone marrow and cerebrospinal fluid. Antigen escape of CD19 and CD22 was not detected in any relapsed patient post-CAR-T cell therapy. Our results indicated that sequential infusion of third generation Anti-CD22 and Anti-CD19 CAR-T cell therapy is feasible and safe for patients with refractory/relapsed B-ALL. Dual antigen targeting should be a promising approach for overcoming antigen escape relapse, while needs to be further determined in our clinical trial. Disclosures No relevant conflicts of interest to declare.

scholarly journals New Generation Chimeric Antigen Receptor T-Cell Therapy (CoupledCAR) Induces High Rate Remissions in Solid Tumor

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 4631-4631
Author(s):  
Lei Xiao
Keyword(s):  
Clinical Trials ◽  
T Cells ◽  
Thyroid Cancer ◽  
T Cell ◽  
Cell Therapy ◽  
Solid Tumors ◽  
Tumor Cells ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T

New Generation Chimeric Antigen Receptor T-Cell Therapy ( CoupledCAR ) Induces High Rate Remissions in Solid Tumor Yu Liu1,Song Li2,Youli Luo3,Haixia Song4,Chengfei Pu5, Zhiyuan Cao 5, Cheng Lu5,Yang Hang5,Xi Huang5,Xiaogang Shen5 ,Xiaojun Hu3 , Renbin Liu1,Xiuwen Wang2,Junjie Mao3,Shihong Wei4 ,Zhao Wu5and Lei Xiao5* 1.The Third Affiliated Hospital, SUN YAT-SEN University 2.Qilu Hospital of Shandong University 3.The Fifth Affiliated Hospital, SUN YAT-SEN University 4.Gansu Procincial Cancer Hospital 5.Innovative Cellular Therapeutics *Corresponding to: Lei Xiao, [email protected] Chimeric antigen receptor (CAR) T cell therapy made significant progress for treating blood cancer such as leukemia, lymphoma, and myeloma. However, the therapy faces many challenges, such as physical barrier, tumor microenvironment immunosuppression, tumor heterogeneity, target specificity, and cell expansion in vivo for treatment of solid tumors Conventional CAR T cell therapy showed weak CAR T expansion in patients and thus achieved no or little response for treating solid tumors. Here, we generated "CoupledCAR" T cells including an anti-TSHR CAR molecule. Compared with conventional CART cells,these "CoupledCAR" T cells successfully improved the expansion of CART cells more than 100 times and enhanced CAR T cells' migration ability, allowing the CAR T cells to resist and infiltrate the tumor microenvironment and killed tumor cells. To verify the effect of "CoupledCAR" T cells on solid tumors, we have completed several clinical trials for different solid tumors, including two patients with thyroid cancer. Immunohistochemistry (IHC) results showed that thyroid stimulating hormone receptors (TSHR) were highly expressed in thyroid cancer cells. In vitro co-culture experiments showed that TSHR CAR T cells specifically recognized and killed TSHR-positive tumor cells. Animal experiments showed that TSHR CAR T cells inhibited the proliferation of TSHR-positive tumor cells. Therefore, we designed "CoupledCAR" T cells expressing a binding domain against TSHR. Further,we did clinical trials of two group patients that were successfully treated using conventional TSHR CAR T cells and the "CoupledCAR" T cells, respectively. In the first group using conventional TSHR CAR T cells, patients showed weak cell expansion and less migration ability. In the group using TSHR "CoupledCAR" T cells, patients showed rapid expansion of CAR T cells and killing of tumor cells. One month after infusion (M1), the patient was evaluated as PR(Partial Response): the lymph node metastasis disappeared, and thoracic paratracheal tumors decreased significantly. Three months after infusion (M3), the patient was evaluated as a durable response, and the tumor tissue was substantially smaller than M1. Further, two patients with colonrectal cancer were enrolled in this trial and infused "CoupledCAR" T cells. One patient achieved PR and the other one achieved SD (Stable Disease). Therefore, "CoupledCAR" T cells can effectively promote expansion, migration and killing ability of CAR T cells in patients with thyroid cancer. "CoupledCAR" T cell technology is a technological platform, which may be used to treat other cancer types. Next, we are recruiting more patients with solid tumors in clinical trials using "CoupledCAR" T cells. Disclosures No relevant conflicts of interest to declare.

Donor-derived CD7 CAR T cells for T-cell acute lymphoblastic leukemia.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. 10001-10001
Author(s):  
Jing Pan ◽  
Jing Pan ◽  
Biping Deng ◽  
Zhuojun Ling ◽  
Weiliang Song ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Residual Disease ◽  
Lymphoblastic Leukemia ◽  
Cellular Immunotherapy ◽  
Hematologic Toxicity ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T ◽  
Grade 3

10001 Background: Despite the success of chimeric antigen receptor T cell therapy in B cell malignances, there is currently no proved CAR T treatment for T cell neoplasms. We provide first evidence support the use of donor derived CAR T cells in T cell leukemia. Methods: In this phase 1 trial, CD7 CAR T cells were manufactured with T cells from prior SCT prior to a single infusion at doses of 5 × 105 or 1 × 106 (± 30%) cells per kilogram of body weight. donors, or from new donors who were HLA-matched or haploidentical, via leukopheresis and transduced with a lentiviral vector which carries a CD7 CAR construct. The primary endpoint was safety. Short-term efficacy was also assessed. Results: Results of 20 enrolled patients who received infusion are reported. Of 20 patients, 12 received previous HSCT-donor derived CAR T cells and 8 received fresh haplo-identical donor derived CAR T cells and plan to received transplantation as consolidation after remission.Adverse events included grade 3-4 hematologic toxicity in all (100%), grade 3-4 and grade 1-2 cytokine release syndrome in 2 (10%) and in 18 (90%), grade 1 neurotoxicity in 3 (15%), grade 1-2 graft-versus-host disease in 12 (60%), and grade 1 viral activation in 3 (15%) patients. Nineteen (95%) patients had a response, including 18 (90%) with complete remission and 1 (5%) with partial remission. Of 19 responders, 7 were bridged to SCT and remained minimal residual disease (MRD)-negative until last visit; 12 were followed up at a medium of 4.4 months, among whom 9 remained MRD-negative, 1 had a relapse, 1 discontinued for other treatment, and 1 died of pulmonary fungal infection at 5.5 months. CAR cells mostly persisted beyond 3 months. Patient CD7-positive healthy T cells were depleted, while CD7-negative T cells increased. Conclusions: We report the initial toxicity profile and anti-leukemia activity of a donor-derived CD7-targeted cellular immunotherapy for patients with relapsed or refractory T-ALL. (Funded by the National Key R&D program; ChiCTR.org number, ChiCTR2000034762). Clinical trial information: ChiCTR2000034762. [Table: see text]

scholarly journals Efficacy and safety of CAR19/22 T-cell cocktail therapy in patients with refractory/relapsed B-cell malignancies

Blood ◽  
2020 ◽  
Vol 135 (1) ◽  
pp. 17-27 ◽  
Author(s):  
Na Wang ◽  
Xuelian Hu ◽  
Wenyue Cao ◽  
Chunrui Li ◽  
Yi Xiao ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Target Antigen ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T Cell Therapy ◽  
Car T ◽  
Negative Tumor

Relapse following chemeric antigen receptor (CAR) T-cell therapy can arise from progressive loss of the CAR T cells or from loss of the target antigen by tumor cells. Wang et al report that using a mix of CAR T cells targeting CD19 and CD22 reduces relapse with antigen-negative tumor cells. However, a lack of CAR T-cell persistence leads to increased relapse with antigen-positive cells.

scholarly journals Engineered Removal of PD-1 From the Surface of CD19 CAR-T Cells Results in Increased Activation and Diminished Survival

2021 ◽  
Vol 8 ◽  
Author(s):  
R. S. Kalinin ◽  
V. M. Ukrainskaya ◽  
S. P. Chumakov ◽  
A. M. Moysenovich ◽  
V. M. Tereshchuk ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Therapy ◽  
Tumor Cells ◽  
Functional Impairment ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T Cell Therapy ◽  
Car T

CAR-T cell therapy is the most advanced way to treat therapy resistant hematologic cancers, in particular B cell lymphomas and leukemias, with high efficiency. Donor T cells equipped ex vivo with chimeric receptor recognize target tumor cells and kill them using lytic granules. CAR-T cells that recognize CD19 marker of B cells (CD19 CAR-T) are considered the gold standard of CAR-T therapy and are approved by FDA. But in some cases, CD19 CAR-T cell therapy fails due to immune suppressive microenvironment. It is shown that tumor cells upregulate expression of PD-L1 surface molecule that binds and increases level and signal provided by PD-1 receptor on the surface of therapeutic CAR-T cells. Induction of this negative signaling results in functional impairment of cytotoxic program in CAR-T cells. Multiple attempts were made to block PD-1 signaling by reducing binding or surface level of PD-1 in CAR-T cells by various means. In this study we co-expressed CD19-CAR with PD-1-specific VHH domain of anti-PD-1 nanobody to block PD-1/PD-L1 signaling in CD19 CAR-T cells. Unexpectedly, despite increased activation of CAR-T cells with low level of PD-1, these T cells had reduced survival and diminished cytotoxicity. Functional impairment caused by disrupted PD-1 signaling was accompanied by faster maturation and upregulation of exhaustion marker TIGIT in CAR-T cells. We conclude that PD-1 in addition to its direct negative effect on CAR-induced signaling is required for attenuation of strong stimulation leading to cell death and functional exhaustion. These observations suggest that PD-1 downregulation should not be considered as the way to improve the quality of therapeutic CAR-T cells.

scholarly journals CAR T cells targeting CD99 as an approach to eradicate T-cell acute lymphoblastic leukemia without normal blood cells toxicity

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Jiangzhou Shi ◽  
Zijian Zhang ◽  
Hong Cen ◽  
Han Wu ◽  
Shangkun Zhang ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Blood Cells ◽  
Clinical Success ◽  
Lymphoblastic Leukemia ◽  
Leukemia Cells ◽  
Normal Blood ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T

AbstractCAR T cell therapy has shown dramatic clinical success in relapsed or refractory B-ALL and other hematological malignancies. However, the loss of specific antigens, cell fratricide, T cell aplasia, and normal T cell separation are challenges in treating T cell leukemia/lymphoma with CAR T therapy. CD99 is a promising antigen to target T-ALL and AML as it is strongly expressed on the majority of T-ALL and AML. Here, we isolated a low-affinity CD99 (12E7) antibody, which specifically recognizes leukemia cells over normal blood cells. Moreover, T cells transduced with an anti-CD99-specific CAR that contained the 12E7 scFv expanded with minor fratricide and without normal blood cells toxicity. We observed that our anti-CD99 CAR T cells showed robust cytotoxicity specifically against CD99+ T-ALL cell lines and primary tumor cells in vitro and significantly prolonged cell line-derived xenografts (CDXs) or patient-derived xenografts (PDXs) models survival in vivo. Together, our results demonstrate that anti-CD99 CAR T cells could specifically recognize and efficiently eliminate CD99+ leukemia cells.

scholarly journals CAR T-cells that target acute B-lineage leukemia irrespective of CD19 expression

Leukemia ◽  
2020 ◽  
Vol 35 (1) ◽  
pp. 75-89 ◽  
Author(s):  
Kristen Fousek ◽  
Junji Watanabe ◽  
Sujith K. Joseph ◽  
Ann George ◽  
Xingyue An ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Tracking ◽  
Lymphoblastic Leukemia ◽  
Target Cells ◽  
T Cell Therapy ◽  
Car T Cells ◽  
Car T ◽  
B Lineage ◽  
Cd19 Expression

AbstractChimeric antigen receptor (CAR) T-cells targeting CD19 demonstrate remarkable efficacy in treating B-lineage acute lymphoblastic leukemia (BL-ALL), yet up to 39% of treated patients relapse with CD19(−) disease. We report that CD19(−) escape is associated with downregulation, but preservation, of targetable expression of CD20 and CD22. Accordingly, we reasoned that broadening the spectrum of CD19CAR T-cells to include both CD20 and CD22 would enable them to target CD19(−) escape BL-ALL while preserving their upfront efficacy. We created a CD19/20/22-targeting CAR T-cell by coexpressing individual CAR molecules on a single T-cell using one tricistronic transgene. CD19/20/22CAR T-cells killed CD19(−) blasts from patients who relapsed after CD19CAR T-cell therapy and CRISPR/Cas9 CD19 knockout primary BL-ALL both in vitro and in an animal model, while CD19CAR T-cells were ineffective. At the subcellular level, CD19/20/22CAR T-cells formed dense immune synapses with target cells that mediated effective cytolytic complex formation, were efficient serial killers in single-cell tracking studies, and were as efficacious as CD19CAR T-cells against primary CD19(+) disease. In conclusion, independent of CD19 expression, CD19/20/22CAR T-cells could be used as salvage or front-line CAR therapy for patients with recalcitrant disease.

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