CARAMBA: a first-in-human clinical trial with SLAMF7 CAR-T cells prepared by virus-free Sleeping Beauty gene transfer to treat multiple myeloma

AbstractClinical development of chimeric antigen receptor (CAR)-T-cell therapy has been enabled by advances in synthetic biology, genetic engineering, clinical-grade manufacturing, and complex logistics to distribute the drug product to treatment sites. A key ambition of the CARAMBA project is to provide clinical proof-of-concept for virus-free CAR gene transfer using advanced Sleeping Beauty (SB) transposon technology. SB transposition in CAR-T engineering is attractive due to the high rate of stable CAR gene transfer enabled by optimized hyperactive SB100X transposase and transposon combinations, encoded by mRNA and minicircle DNA, respectively, as preferred vector embodiments. This approach bears the potential to facilitate and expedite vector procurement, CAR-T manufacturing and distribution, and the promise to provide a safe, effective, and economically sustainable treatment. As an exemplary and novel target for SB-based CAR-T cells, the CARAMBA consortium has selected the SLAMF7 antigen in multiple myeloma. SLAMF7 CAR-T cells confer potent and consistent anti-myeloma activity in preclinical assays in vitro and in vivo. The CARAMBA clinical trial (Phase-I/IIA; EudraCT: 2019-001264-30) investigates the feasibility, safety, and anti-myeloma efficacy of autologous SLAMF7 CAR-T cells. CARAMBA is the first clinical trial with virus-free CAR-T cells in Europe, and the first clinical trial that uses advanced SB technology worldwide.

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Genomic Analyses of SLAMF7 CAR-T Cells Manufactured by Sleeping Beauty Transposon Gene Transfer for Immunotherapy of Multiple Myeloma

10.1101/675009 ◽

2019 ◽

Author(s):

Csaba Miskey ◽

Maximilian Amberger ◽

Michael Reiser ◽

Sabrina Prommersberger ◽

Julia Beckmann ◽

...

Keyword(s):

Clinical Trial ◽

Multiple Myeloma ◽

T Cells ◽

T Cell ◽

Gene Transfer ◽

Copy Number ◽

Sleeping Beauty ◽

Vector System ◽

Car T Cells ◽

Car T

ABSTRACTWidespread treatment of human diseases with gene therapies necessitates the development of gene transfer vectors that integrate genetic information effectively, safely and economically. Accordingly, significant efforts have been devoted to engineer novel tools that i) achieve high-level stable gene transfer at low toxicity to the host cell; ii) induce low levels of genotoxicity and possess a ‘safe’ integration profile with a high proportion of integrations into safe genomic locations; and iii) are associated with acceptable cost per treatment and scalable/exportable vector production to serve large numbers of patients. The Sleeping Beauty (SB) transposon has been transformed into a vector system that is fulfilling these requirements.In the CARAMBA project, we use SB transposition to genetically modify T cells with a chimeric antigen receptor (CAR) specific for the SLAMF7 antigen, that is uniformly and highly expressed on malignant plasma cells in multiple myeloma. We have demonstrated that SLAMF7 CAR-T cells confer specific and very potent anti-myeloma reactivity in pre-clinical models, and are therefore preparing a Phase I/IIa clinical trial of adoptive immunotherapy with autologous, patient-derived SLAMF7-CAR T cells in multiple myeloma (EudraCT Nr. 2019-001264-30/CARAMBA-1).Here we report on the characterization of genomic safety attributes in SLAMF7 CAR-T cells that we prepared in three clinical-grade manufacturing campaigns under good manufacturing practice (GMP), using T cells that we obtained from three healthy donor volunteers. In the SLAMF7 CAR-T cell product, we determined the average transposon copy number, the genomic insertion profile, and presence of residual SB100X transposase. The data show that the SLAMF7 CAR transposon had been inserted into the T cell genome with the close-to-random distribution pattern that is typical for SB, and with an average transposon copy number ranging between 6 and 12 per T cell. No residual SB100X transposase could be detected by Western blotting in the infusion products. With these attributes, the SLAMF7 CAR-T products satisfy criteria set forth by competent regulatory authorities in order to justify administration of SLAMF7 CAR-T cells to humans in the context of a clinical trial. These data set the stage for the CARAMBA clinical trial, that will be the first in the European Union to use virus-free SB transposition for CAR-T engineering.DisclosuresThis project is receiving funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 754658 (CARAMBA).

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An Anti-Bcma CAR T-Cell Therapy (C-CAR088) Shows Promising Safety and Efficacy Profile in Relapsed or Refractory Multiple Myeloma

Blood ◽

10.1182/blood-2020-138734 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 29-30

Author(s):

Gang An ◽

Weiwei Sui ◽

Tingyu Wang ◽

Xiaoyan Qu ◽

Xian Zhang ◽

...

Keyword(s):

Clinical Trial ◽

Multiple Myeloma ◽

T Cells ◽

Cell Therapy ◽

Publicly Traded ◽

T Cell Therapy ◽

Car T Cells ◽

Car T Cell ◽

Car T Cell Therapy ◽

Car T

Background: C-CAR088, an anti-BCMA CAR T-cell therapy is a novel 2nd generation 4-1BB chimeric antigen receptor T (CAR-T) cell therapy targeting BCMA which is specifically and highly expressed on multiple myeloma (MM) cells. C-CAR088 is manufactured in a serum-free, automated and digital, closed system. Initial, early clinical trial results in patients with R/R MM supported preclinical findings and showed promising efficacy and manageable safety profile (Yao, Blood (2019) 134 (Supplement_1): 50.) Methods: The dose escalation and expansion studies have been conducted at four medical centers in China to evaluate the safety and efficacy of C-CAR088 in patients with R/R MM who were previously treated with at least 2 lines of therapy including proteasome inhibitors (PIs) and IMiDs. C-CAR088 is administered to patients as a single intravenous dose after a standard 3-day cyclophosphamide/fludarabine conditioning regimen. Results: As of July 15, 2020, 24 patients were infused and 21 patients had evaluable data for safety and clinical response at dose levels of 1.0 x 106 CAR-T cells/kg (n=3), 3 x106 CAR-T cells/kg (n=11) and 4.5~6x106 CAR-T cells/kg (n=7). The median vein to vein time was 16 days. The manufacturing success rate was 100%. The median age of patients dosed was 60 years (range: 45-74 years).The median number of prior lines of therapy was 4 (range: 2-12 prior therapies). There were 17 (81%) patients with at least one and 12 (57.1%) patients with at least two high risk cytogenetic tumor changes. Five patients (23.8%) had bridging therapy. C-CAR088 treatment was well tolerated. 20 of 21 (95%) patients had Grade 1-2 CRS and one patient experienced Grade 3 CRS. Median time to CRS was 6.5 days (range: 1-11 days) and median duration of CRS was 5 days (range: 2-10 days). Four patients (19%) received tocilizumab for CRS treatment. Only one patient experienced a Grade 1 neurotoxicity event. No dose-limiting toxicities were observed and all adverse events were reversible. The best overall response (BOR) included 6 complete responses (CRs), 10 very good partial responses (VGPRs) and 4 partial responses (PRs). Median follow-up was 182 days (range: 30-375 days). The median duration of response has not been reached. In the 3 x106 CAR-T cells/kg dose group, 5/11(45%) patients achieved a CR. The C-CAR088 PK profile in peripheral blood showed a trend of a dose dependent profile. AUC0~28day and Cmax increased and Tmax decreased with dose (P<0.05). Conclusion: The clinical trial results in patients with R/R MM treated with C-CAR088 show a favorable safety profile and promising signs of efficacy. We will continue to evaluate these patients to understand the long-term effect of C-CAR088 in multiple myeloma patients. Clinical trial information: NCT04322292、NCT03815383、NCT03751293、NCT04295018 Research Sponsor: Cellular Biomedicine Group, Inc. Disclosures Zhu: Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Zheng:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Yan:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Lv:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Lan:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Yang:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Huo:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Han:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Zhao:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Qin:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Wu:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Yao:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Zhu:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Ren:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Zhang:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Huang:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Humphries:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company. Yao:Cellular Biomedicine Group Inc: Current Employment, Current equity holder in publicly-traded company.

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Safety and Efficacy of Anti-Bcma CAR-T Cells for Relapsed/Refractory Multiple Myeloma: A Systematic Review of Literature

Blood ◽

10.1182/blood-2020-134529 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 5-6

Author(s):

Israr Khan ◽

Abdul Rafae ◽

Anum Javaid ◽

Zahoor Ahmed ◽

Haifza Abeera Qadeer ◽

...

Keyword(s):

Clinical Trial ◽

Multiple Myeloma ◽

Clinical Trials ◽

T Cells ◽

Partial Response ◽

Residual Disease ◽

Safety And Efficacy ◽

Refractory Multiple Myeloma ◽

Car T Cells ◽

Car T

Background: Multiple myeloma (MM) is a plasma cell disorder and demonstrates overexpression of B cell maturation antigen (BCMA). Our objective is to evaluate the safety and efficacy of chimeric antigen receptor T cells (CAR-T) against BCMA in patients with relapsed/refractory multiple myeloma (RRMM). Methods: We conducted a systematic literature search using PubMed, Cochrane, Clinicaltrials.gov, and Embase databases. We also searched for data from society meetings. A total of 935 articles were identified, and 610 were screened for relevance. Results: Data from thirty-one original studies with a total of 871 patients (pts) were included based on defined eligibility criteria, see Table 1. Hu et al. reported an overall response rate (ORR) of 100% in 33 pts treated with BCMA CAR-T cells including 21 complete response (CR), 7 very good partial response (VGPR), 4 partial response (PR). Moreover, 32 pts achieved minimal residual disease (MRD) negative status. Chen et al. reported ORR of 88%, 14% CR, 6% VGPR, and 82% MRD negative status with BCMA CAR-T therapy in 17 RRMM pts. In another clinical trial by Han et al. BCMA CAR-T therapy demonstrated an ORR of 100% among 7 evaluable pts with 43% pts having ≥ CR and 14% VGPR. An ORR of 100% with 64% stringent CR (sCR) and 36% VGPR was reported with novel anti-BCMA CART cells (CT103A). Similarly, Li et al. reported ORR of 87.5%, sCR of 50%, VGPR 12.5%, and PR 25% in 16 pts. BCMA targeting agent, JNJ-4528, showed ORR of 91%, including 4sCR, 2CR, 10MRD, and 7VGPR. CAR-T- bb2121 demonstrated ORR of 85%, sCR 36%, CR 9%, VGPR 57%, and MRD negativity of 100% (among 16 responsive pts). GSK2857916, a BCMA targeting CAR-T cells yielded ORR of 60% in both clinical trials. Three studies utilizing bispecific CART cells targeting both BCMA & CD38 (LCARB38M) reported by Zhao et al., Wang et al., and Fan et al. showed ORR of 88%, 88%, & 100% respectively. Topp et al. reported ORR of 31% along with 5 ≥CR and 5 MRD negative status in 42 pts treated with Bi T-cells Engager BiTE® Ab BCMA targeting antigen (AMG420). One clinical trial presented AUTO2 CART cells therapy against BCMA with an ORR of 43%, VGPR of 14%, and PR of 28%. CT053CAR-BCMA showed 14sCR and 5CR with a collective ORR of 87.5% and MRD negative status of 85% in 24 and 20 evaluable pts, respectively. Likewise, Mikkilineni et al. reported an ORR of 83%, sCR of 16.7%, and VGPR & PR of 25% and 41% in 12 pts treated with FHVH-BCMA T cells. Similar results are also reported in other clinical trials of BCMA targeting CART therapy (Table 1). The most common adverse effects exhibited were grade 1-3 hematologic (cytopenia) and cytokine release syndrome (CRS) (mostly reversible with tocilizumab). Conclusion: Initial data from ongoing clinical trials using BCMA targeting CAR-T therapy have yielded promising results both in terms of improved outcome and tolerable toxicity profiles. Although two phase 3 trails are ongoing, additional data is warranted to further ensure the safety and efficacy of anti-BCMA CAR-T cells therapy in pts with RRMM for future use. Disclosures Anwer: Incyte, Seattle Genetics, Acetylon Pharmaceuticals, AbbVie Pharma, Astellas Pharma, Celegene, Millennium Pharmaceuticals.: Honoraria, Research Funding, Speakers Bureau.

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GPRC5D is a target for the immunotherapy of multiple myeloma with rationally designed CAR T cells

Science Translational Medicine ◽

10.1126/scitranslmed.aau7746 ◽

2019 ◽

Vol 11 (485) ◽

pp. eaau7746 ◽

Cited By ~ 49

Author(s):

Eric L. Smith ◽

Kim Harrington ◽

Mette Staehr ◽

Reed Masakayan ◽

Jon Jones ◽

...

Keyword(s):

Multiple Myeloma ◽

T Cells ◽

T Cell ◽

Cell Therapy ◽

B Cell ◽

T Cell Therapy ◽

Car T Cells ◽

Car T Cell Therapy ◽

Car T

Early clinical results of chimeric antigen receptor (CAR) T cell therapy targeting B cell maturation antigen (BCMA) for multiple myeloma (MM) appear promising, but relapses associated with residual low-to-negative BCMA-expressing MM cells have been reported, necessitating identification of additional targets. The orphan G protein–coupled receptor, class C group 5 member D (GPRC5D), normally expressed only in the hair follicle, was previously identified as expressed by mRNA in marrow aspirates from patients with MM, but confirmation of protein expression remained elusive. Using quantitative immunofluorescence, we determined that GPRC5D protein is expressed on CD138+ MM cells from primary marrow samples with a distribution that was similar to, but independent of, BCMA. Panning a human B cell–derived phage display library identified seven GPRC5D-specific single-chain variable fragments (scFvs). Incorporation of these into multiple CAR formats yielded 42 different constructs, which were screened for antigen-specific and antigen-independent (tonic) signaling using a Nur77-based reporter system. Nur77 reporter screen results were confirmed in vivo using a marrow-tropic MM xenograft in mice. CAR T cells incorporating GPRC5D-targeted scFv clone 109 eradicated MM and enabled long-term survival, including in a BCMA antigen escape model. GPRC5D(109) is specific for GPRC5D and resulted in MM cell line and primary MM cytotoxicity, cytokine release, and in vivo activity comparable to anti-BCMA CAR T cells. Murine and cynomolgus cross-reactive CAR T cells did not cause alopecia or other signs of GPRC5D-mediated toxicity in these species. Thus, GPRC5D(109) CAR T cell therapy shows potential for the treatment of advanced MM irrespective of previous BCMA-targeted therapy.

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First-in-Human CD4 CAR Clinical Trial on Peripheral T-Cell Lymphoma

Blood ◽

10.1182/blood-2019-122789 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 2881-2881

Author(s):

Hongyu Zhang ◽

Jia Feng ◽

Wenli Zhang ◽

Qi Chen ◽

Yuanzhen Cao ◽

...

Keyword(s):

Clinical Trial ◽

T Cells ◽

T Cell ◽

Sézary Syndrome ◽

Sezary Syndrome ◽

T Cell Therapy ◽

Car T Cells ◽

Car T Cell ◽

Car T Cell Therapy ◽

Car T

Background CD19 CAR T cell therapy has achieved success in treating acute lymphoblastic leukemia. However, the treatment for Sezary syndrome, an aggressive form of cutaneous peripheral T cell lymphoma (PTCL) has remained a challenge. Despite patients with Sezary syndrome typically receiving multiple treatments within their disease progression, the prognosis is poor with 5 year survival rate of only 24%. Therefore, it is crucial to establish a novel treatment for PTCLs. CD4 is uniformly expressed on most mature T cell lymphoma, which makes it a promising target for treating PTCLs. Here, we present the efficacy of CD4 CAR T cell in our preclinical study and the success in level 1 dose escalation clinical trial on patients with Sezary syndrome. Methods We engineered a CD4 CAR with scFv (single-chain variable fragment) with CD28 and 4-1BB co-activators fused to CD3zeta and a leader sequence of CD8. The efficacy of CD4 CAR was tested with CD4+ leukemic cell line, primary CD4+ PTCL patient samples and multiple mouse models. An alemtuzumab safety switch has also been established to ensure the elimination of CAR T cells following tumor eradication. Children and adults with PTCLs were enrolled in our phase 1 dose escalation trial to evaluate the safety and efficacy of CD4 CAR T cell antitumor activity. Results Coculture assays results showed that CD4 CAR T cells displayed profound tumor killing effects in leukemia cell lines, primary patient samples and multiple mouse model systems. Our preclinical findings suggest that CD4 CAR T cells is an effective approach in treating PTCLs. Patients enrolled in the phase 1 dose escalation trial have shown remarkable response to CD4 CAR T cells treatment. Noticeably, a 54-yr-old patient diagnosed with Sezary syndrome had achieved complete remission with CD4 CAR T cell therapy. Prior to admission, he had been having symptoms of erythroderma, pruritus and scaling of the skin for over 10 years and had been resistant to multiple lines of chemotherapy. Before the initiation of CAR therapy, patient's body skin has extensive leukemia infiltrate (Fig. 1A) confirmed with skin biopsy (Fig. 1B) with bone marrow and blood comprising 50% leukemic cells (Fig. 1C). Patient received a total dose of 3x10^6 /kg single dose CAR T cells, following which fluconazole and valacyclovir were administrated for infection prophylaxis. Since patient received CD4 CAR T cell infusion, the percentage of CAR T cells (Fig 1. D) in peripheral blood had continue to increase as well as NK cells. (Fig 1. E) On day 13, patient had achieved complete remission with the percentage of leukemia cells in blood decreased to zero (Fig. 1C). On day 28, the appearance of the skin had undergone drastic change from what was before the treatment. Noticeable skin regeneration on both legs of the patients was observed (Fig 1. F). Flow cytometry of bone marrow and peripheral blood confirmed the absence of tumor cells. In addition, Skin biopsy on multiple sites demonstrated absence of leukemia infiltrates post CAR treatment (Fig. 1G). Patient was subsequently discharged with no additional medication needed. Throughout the treatment, patient had developed no infections with Grade II CRS toxicity noted. No other toxicities were observed. Updated results on other patients enrolled in this clinical trial including adverse events will be presented. Conclusion Our first-in-human clinical trial demonstrates promising efficacy of CD4 CAR T cell therapy in treating patients with refractory Sezary syndrome. cCAR is able to eradicate leukemia blasts, exerting a profound tumor killing effect that is superior to traditional chemotherapies. Disclosures Pinz: iCell Gene Therapeutics LLC: Employment. Ma:iCAR Bio Therapeutics Ltd: Employment. Wada:iCell Gene Therapeutics LLC: Employment. Ma:iCell Gene Therapeutics LLC: Consultancy, Equity Ownership, Research Funding; iCAR Bio Therapeutics Ltd: Consultancy, Equity Ownership, Research Funding.

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Paving the Way toward Successful Multiple Myeloma Treatment: Chimeric Antigen Receptor T-Cell Therapy

Cells ◽

10.3390/cells9040983 ◽

2020 ◽

Vol 9 (4) ◽

pp. 983 ◽

Cited By ~ 1

Author(s):

Ewelina Grywalska ◽

Barbara Sosnowska-Pasiarska ◽

Jolanta Smok-Kalwat ◽

Marcin Pasiarski ◽

Paulina Niedźwiedzka-Rystwej ◽

...

Keyword(s):

Multiple Myeloma ◽

T Cells ◽

T Cell ◽

Side Effects ◽

Cell Therapy ◽

T Cell Therapy ◽

Car T Cells ◽

Car T Cell ◽

Car T Cell Therapy ◽

Car T

Despite the significant progress of modern anticancer therapies, multiple myeloma (MM) is still incurable for the majority of patients. Following almost three decades of development, chimeric antigen receptor (CAR) T-cell therapy now has the opportunity to revolutionize the treatment landscape and meet the unmet clinical need. However, there are still several major hurdles to overcome. Here we discuss the recent advances of CAR T-cell therapy for MM with an emphasis on future directions and possible risks. Currently, CAR T-cell therapy for MM is at the first stage of clinical studies, and most studies have focused on CAR T cells targeting B cell maturation antigen (BCMA), but other antigens such as cluster of differentiation 138 (CD138, syndecan-1) are also being evaluated. Although this therapy is associated with side effects, such as cytokine release syndrome and neurotoxicity, and relapses have been observed, the benefit–risk balance and huge potential drive the ongoing clinical progress. To fulfill the promise of recent clinical trial success and maximize the potential of CAR T, future efforts should focus on the reduction of side effects, novel targeted antigens, combinatorial uses of different types of CAR T, and development of CAR T cells targeting more than one antigen.

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Transposon-Based CAR T Cells in Acute Leukemias: Where Are We Going?

Cells ◽

10.3390/cells9061337 ◽

2020 ◽

Vol 9 (6) ◽

pp. 1337 ◽

Cited By ~ 1

Author(s):

Chiara F. Magnani ◽

Sarah Tettamanti ◽

Gaia Alberti ◽

Ilaria Pisani ◽

Andrea Biondi ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Gene Transfer ◽

Cell Therapy ◽

Viral Vector ◽

T Cell Therapy ◽

Car T Cells ◽

Car T Cell ◽

Car T Cell Therapy ◽

Car T

Chimeric Antigen Receptor (CAR) T-cell therapy has become a new therapeutic reality for refractory and relapsed leukemia patients and is also emerging as a potential therapeutic option in solid tumors. Viral vector-based CAR T-cells initially drove these successful efforts; however, high costs and cumbersome manufacturing processes have limited the widespread clinical implementation of CAR T-cell therapy. Here we will discuss the state of the art of the transposon-based gene transfer and its application in CAR T immunotherapy, specifically focusing on the Sleeping Beauty (SB) transposon system, as a valid cost-effective and safe option as compared to the viral vector-based systems. A general overview of SB transposon system applications will be provided, with an update of major developments, current clinical trials achievements and future perspectives exploiting SB for CAR T-cell engineering. After the first clinical successes achieved in the context of B-cell neoplasms, we are now facing a new era and it is paramount to advance gene transfer technology to fully exploit the potential of CAR T-cells towards next-generation immunotherapy.

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Chimeric Antigen Receptor T-Cell Therapy for Multiple Myeloma

Cancers ◽

10.3390/cancers11122024 ◽

2019 ◽

Vol 11 (12) ◽

pp. 2024 ◽

Cited By ~ 4

Author(s):

Naoki Hosen

Keyword(s):

Multiple Myeloma ◽

T Cells ◽

T Cell ◽

Cell Therapy ◽

B Cell ◽

Chimeric Antigen Receptor ◽

T Cell Therapy ◽

Car T Cells ◽

Car T Cell Therapy ◽

Car T

CD19 Chimeric antigen receptor (CAR) T cell therapy has been shown to be effective for B cell leukemia and lymphoma. Many researchers are now trying to develop CAR T cells for various types of cancer. For multiple myeloma (MM), B-cell maturation antigen (BCMA) has been recently proved to be a promising target. However, cure of MM is still difficult, and several other targets, for example immunoglobulin kappa chain, SLAM Family Member 7 (SLAMF7), or G-protein coupled receptor family C group 5 member D (GPRC5D), are being tested as targets for CAR T cells. We also reported that the activated integrin β7 can serve as a specific target for CAR T cells against MM, and are preparing a clinical trial. In this review, we summarized current status of CAR T cell therapy for MM and discussed about the future perspectives.

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Donor-derived CD19 CAR-T cell therapy of relapse of CD19-positive B-ALL post allotransplant

Leukemia ◽

10.1038/s41375-020-01056-6 ◽

2020 ◽

Cited By ~ 1

Author(s):

Cheng Zhang ◽

Xiao-Qi Wang ◽

Rong-Li Zhang ◽

Fang Liu ◽

Yi Wang ◽

...

Keyword(s):

T Cells ◽

Lymphoblastic Leukemia ◽

Multiorgan Failure ◽

High Rate ◽

T Cell Therapy ◽

Immune Effector ◽

Car T Cells ◽

Car T ◽

Grade 3 ◽

Histological Remission

Abstract Safety and efficacy of allogeneic anti-CD19 chimeric antigen receptor T cells (CAR-T cells) in persons with CD19-positive B-cell acute lymphoblastic leukemia (B-ALL) relapsing after an allotransplant remain unclear. Forty-three subjects with B-ALL relapsing post allotransplant received CAR-T cells were analyzed. 34 (79%; 95% confidence interval [CI]: 66, 92%) achieved complete histological remission (CR). Cytokine release syndrome (CRS) occurred in 38 (88%; 78, 98%) and was ≥grade-3 in 7. Two subjects died from multiorgan failure and CRS. Nine subjects (21%; 8, 34%) developed ≤grade-2 immune effector cell-associated neurotoxicity syndrome (ICANS). Two subjects developed ≤grade-2 acute graft-versus-host disease (GvHD). 1-year event-free survival (EFS) and survival was 43% (25, 62%). In 32 subjects with a complete histological remission without a second transplant, 1-year cumulative incidence of relapse was 41% (25, 62%) and 1-year EFS and survival, 59% (37, 81%). Therapy of B-ALL subjects relapsing post transplant with donor-derived CAR-T cells is safe and effective but associated with a high rate of CRS. Outcomes seem comparable to those achieved with alternative therapies but data from a randomized trial are lacking.

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P09.08 Clinical-grade manufacturing of ROR1 CAR T cells using a novel virus-free protocol

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-itoc7.108 ◽

2020 ◽

Vol 8 (Suppl 2) ◽

pp. A55.2-A56

Author(s):

K Mestermann ◽

M Eichler ◽

M Machwirth ◽

K Kebbel ◽

U Köhl ◽

...

Keyword(s):

T Cells ◽

Gene Transfer ◽

Drug Product ◽

Scale Up ◽

Sleeping Beauty ◽

Clinical Grade ◽

Protocol Optimization ◽

Car T ◽

Genomic Analyses ◽

Biosafety Level

BackgroundImmunotherapy with T cells that were modified by gene-transfer to express a ROR1-specific chimeric antigen receptor (ROR1 CAR-T) has therapeutic potential in ROR1+ malignancies in hematology and oncology. The ROR1 tumor antigen has a favorable expression profile with absence in vital normal human tissues. In this study, we sought to establish and validate clinical-grade manufacturing of ROR1 CAR-T to enable a Phase I/IIa clinical trial. In particular, we sought to integrate virus-free gene-transfer based on Sleeping Beauty transposition into this manufacturing protocol to permit scale-up and export to point-of-care manufacturing, and to reduce turn-around time, complexity and regulatory burden associated with conventional viral gene-transfer (biosafety level 2 to biosafety level 1).Materials and MethodsBuffy coats or leukaphereses were obtained from healthy donors to perform protocol optimization (n=7) and scale-up runs (n=1). CD4+ and CD8+ T cells were isolated separately by magnetic selection and stimulated with CD3/CD28 TransACT® reagent. T cells were transfected with mRNA encoding hyperactive Sleeping Beauty transposase (SB100X) and minicircle DNA (MC) encoding a pT2 transposon comprising the ROR1 CAR and an EGFRt marker gene using the MaxCyte GTx ® electroporation platform. Following transfection, T cells were expanded for 10–13 days in G-REX® bioreactors and then harvested and formulated into the drug product at a 1:1 ratio of CAR-expressing CD4:CD8 T cells. The drug product underwent comprehensive phenotypic, functional and genomic analyses as part of product qualification.ResultsThe set of protocol optimization runs resulted in a highly robust process. On average, the stable gene-transfer rate at the end of the manufacturing process was 71% in CD4+ (n=5) and 54% in CD8+ T cells (n=7). The average yield of ROR1 CAR-T relative to the number of input T cells was 12.6-fold for CD4+ and 9.4-fold for CD8+ after 12–15 days of expansion, with an average viability of 84% for CD4+ and 82% of CD8+ T cells. The scale-up run was performed with a leukapheresis product from which 52.5 × 10^6 CD4+ and 109 × 10^6 CD8+ T cells were transfected. At the end of the manufacturing process (day 12), there were 844 × 10^6 CAR-expressing CD4+ (~16-fold expansion) and 857 × 10^6 CAR-expressing CD8+ T cells (8-fold expansion). In functional testing, ROR1 CAR-T showed specific recognition and potent elimination of ROR1+ target cells, as well as antigen-dependent cytokine production and productive proliferation in in vitro analyses. Experiments to determine the anti-tumor potency of the drug product in vivo and detailed genomic analyses are ongoing. Preliminary analyses suggest a favorable genomic insertion profile of the CAR transposon, and a transposon copy number that is well within the range acceptable for clinical use of the drug product.ConclusionsWith this novel protocol, we aim to obtain the first manufacturing license for CAR-T in Europe that integrates our optimized approach with SB100X mRNA and transposon MC for CAR gene-transfer on the MaxCyte transfection platform. The quality and yield of the drug product support the design and dose escalation of the proposed clinical trial with ROR1 CAR-T, and will serve as a blueprint for other CAR-T products from our pipeline.Disclosure InformationK. Mestermann: None. M. Eichler: None. M. Machwirth: None. K. Kebbel: None. U. Köhl: None. H. Einsele: None. C. Müller: None. J. Lehmann: None. T. Raskó: None. F. Lundberg: None. Z. Izsvák: None. G. Schmiedeknecht: None. M. Hudecek: None.

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