scholarly journals Recent Advances in Allogeneic CAR-T Cells

Biomolecules ◽  
2020 ◽  
Vol 10 (2) ◽  
pp. 263 ◽  
Author(s):  
Dong Wook Kim ◽  
Je-Yoel Cho
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
T Cell Proliferation ◽  
Downstream Signaling ◽  
Car T Cells ◽  
Cell Based Therapy ◽  
Car T ◽  
Genes Encoding ◽  
Signaling Domains

In recent decades, great advances have been made in the field of tumor treatment. Especially, cell-based therapy targeting tumor associated antigen (TAA) has developed tremendously. T cells were engineered to have the ability to attack tumor cells by generating CAR constructs consisting of genes encoding scFv, a co-stimulatory domain (CD28 or TNFRSF9), and CD247 signaling domains for T cell proliferation and activation. Principally, CAR-T cells are activated by recognizing TAA by scFv on the T cell surface, and then signaling domains inside cells connected by scFv are subsequently activated to induce downstream signaling pathways involving T cell proliferation, activation, and production of cytokines. Many efforts have been made to increase the efficacy and persistence and also to decrease T cell exhaustion. Overall, allogeneic and universal CAR-T generation has attracted much attention because of their wide and prompt usage for patients. In this review, we summarized the current techniques for generation of allogeneic and universal CAR-T cells along with their disadvantages and limitations that still need to be overcome.

scholarly journals Preinfusion polyfunctional anti-CD19 chimeric antigen receptor T cells are associated with clinical outcomes in NHL

Blood ◽  
2018 ◽  
Vol 132 (8) ◽  
pp. 804-814 ◽  
Author(s):  
John Rossi ◽  
Patrick Paczkowski ◽  
Yueh-Wei Shen ◽  
Kevin Morse ◽  
Brianna Flynn ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Clinical Response ◽  
Clinical Outcomes ◽  
Chimeric Antigen Receptor ◽  
T Cell Proliferation ◽  
Car T Cells ◽  
Key Points ◽  
Car T

Key Points The PSI of manufactured CAR T cells was associated with clinical response and toxicities. Monitoring CAR T-cell polyfunctionality as a key product attribute may complement other characteristics including T-cell proliferation.

Overcoming the breast tumor microenvironment by targeting MDSCs through CAR-T cell therapy.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. 1032-1032
Author(s):  
Saisha Abhay Nalawade ◽  
Paul Shafer ◽  
Pradip Bajgain ◽  
Katie McKenna ◽  
Arushana Ali ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Tumor Microenvironment ◽  
Nuclear Translocation ◽  
T Cell Proliferation ◽  
Tumor Site ◽  
Car T Cells ◽  
Car T

1032 Background: Successful targeting of solid tumors such as breast cancer (BC) using CAR T cells (CARTs) has proven challenging, largely due to the immune suppressive tumor microenvironment (TME). Myeloid derived suppressor cells (MDSCs) inhibit CART’s function and persistence within the breast TME. We generated CAR T cells targeting tumor-expressed mucin 1 (MUC1) (Bajgain P et al, 2018) for BC. To potentiate expansion and persistence of MUC1 CARTs and modulate the suppressive TME, we developed a novel chimeric co-stimulatory receptor, TR2.4-1BB, encoding a ScFv derived from a TNF-related apoptosis-inducing ligand receptor 2 (TR2) mAb followed by a 4-1BB endodomain. We hypothesize that engagement with TR2 expressed on TME-resident MDSCs, will lead to both MDSC apoptosis and CART co-stimulation, promoting T cell persistence and expansion at tumor site. Methods: Function of the novel TR2.4-1BB receptor, was assessed by exposing non-transduced (NT) and TR2.4-1BB transduced T cells to recombinant TR2 and nuclear translocation of NFκB was measured by ELISA. Functionality of in vitro generated MDSCs was determined by the suppression assay. In vitro CART/costimulatory receptor T cell function was measured by cytotoxicity assays using MUC1+ tumor targets in presence or absence of MDSCs. In vivo anti-tumor activity was assessed using MDSC enriched tumor-bearing mice using calipers to assess tumor volume and bioluminescence imaging to track T cells. Results: Nuclear translocation of NFκB was detected only in TR2.4-1BB T cells. MDSCs significantly attenuated T cell proliferation by 50±5% and IFNγ production by half compared with T cells cultured alone. Additionally, presence of MDSCs, diminished cytotoxic potential of MUC1 CARTs against MUC1+ BC cell lines by 25%. However, TR2.4-1BB expression on CAR.MUC1 T cells induced MDSC apoptosis thereby restoring the cytotoxic activity of CAR.MUC1 against MUC1+ BC lines in presence of TR2.4-1BB (67±8.5%). There was an approximate two-fold increase in tumor growth due enhanced angiogenesis and fibroblast accumulation in mice receiving tumors + MDSCs compared to tumors alone. Treatment of these MDSC-enriched tumors with MUC1.TR2.4-1BB CARTs led to superior tumor cell killing and significant reduction in tumor growth (24.54±8.55 mm3) compared to CAR.MUC1 (469.79.9±81.46mm3) or TR2.4-1BB (434.86±64.25 mm3) T cells alone (Day 28 after T cell injection). The treatment also improved T cell proliferation and persistence at the tumor site. Thereby, leading to negligible metastasis demonstrating ability of CARTs to eliminate tumor and prevent dissemination. We observed similar results using HER2.TR2.4-1BB CARTs in a HER2+ BC model. Conclusions: Our findings demonstrate that CARTs co-expressing our novel TR2.4-1BB receptor have higher anti-tumor potential against BC tumors and infiltrating MDSCs, resulting in TME remodeling and improved T cell proliferation at the tumor site.

scholarly journals Tocilizumab, but not dexamethasone, prevents CRS without affecting antitumor activity of bispecific antibodies

2020 ◽  
Vol 8 (1) ◽  
pp. e000621 ◽  
Author(s):  
Joseph Kauer ◽  
Sebastian Hörner ◽  
Lukas Osburg ◽  
Stefanie Müller ◽  
Melanie Märklin ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Antitumor Activity ◽  
Bispecific Antibodies ◽  
T Cell Proliferation ◽  
Car T Cells ◽  
Car T

Bispecific antibodies (bsAb) and chimeric antigen receptor (CAR) T cells allow for antibody guided recruitment of T cells against tumors. Both are successfully used for treatment of CD19 expressing leukemias, but may cause cytokine release syndrome (CRS) as a major dose-limiting side effect. For CRS prevention, steroids are recommended prior to bsAb treatment, despite their well-known lymphotoxic activity. The IL-6 receptor antibody tocilizumab is established for treatment of CRS induced by CAR T cells, but was not considered for CRS prevention in bsAb therapy. We here compared the influence of dexamethasone and tocilizumab on bsAb-mediated T cell proliferation and tumor lysis in vitro and in vivo and found that dexamethasone profoundly inhibited T cell proliferation and antitumor activity as induced by two different bsAb, particularly at low effector:target ratios, whereas tocilizumab did not affect efficacy. When we applied tocilizumab early during treatment of three patients with a newly developed PSMAxCD3 bsAb, significant CRS attenuation despite high IL-6 serum levels was observed. Thus, early IL-6 blockade may reduce the undesired sequelae of CRS upon bsAb therapy without affecting therapeutic activity, allowing in turn for safe application of effective doses.

CD19-Redirected Chimeric Antigen Receptor T (CART19) Cells Induce a Cytokine Release Syndrome (CRS) and Induction of Treatable Macrophage Activation Syndrome (MAS) That Can Be Managed by the IL-6 Antagonist Tocilizumab (toc).

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2604-2604 ◽  
Author(s):  
Stephan A. Grupp ◽  
David L Porter ◽  
David T Teachey ◽  
David M. Barrett ◽  
Anne Chew ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Chimeric Antigen Receptor ◽  
Tumor Lysis Syndrome ◽  
Antigen Receptor ◽  
Car T Cells ◽  
Car T ◽  
Anti Tumor Activity

Abstract Abstract 2604 We previously reported on CART19 cells expressing a chimeric antigen receptor (CAR) with intracellular activation and costimulatory domains. Infusion of these cells results in 100 to 100,000× in vivo proliferation, tumor lysis syndrome followed by durable antitumor activity, and prolonged persistence in pts with B cell tumors. Here we report that in vivo proliferation of CART19 cells and potent anti-tumor activity is associated with CRS, leading to hemophagocytic lymphohistiocytosis (HLH), also termed MAS. We propose that MAS/HLH is a unique biomarker that is associated with and may be required for potent anti-tumor activity. Autologous T cells were lentivirally transduced with a CAR composed of anti-CD19 scFv/4-1BB/CD3-zeta, activated/expanded ex-vivo with anti-CD3/anti-CD28 beads, and then infused into ALL or CLL pts with persistent disease after 2–8 prior treatments. CART19 anti ALL activity was also modeled in a xenograft mouse model with high level of human ALL/human T cell engraftment and simultaneous detection of CAR T cells and ALL using 2-color bioluminescent imaging. We describe updated results of 10 pts who received CART19 cells elsewhere at ASH (Porter, et al), including 9 pts with CLL and 1 pediatric pt with relapsed refractory ALL. 6/9 evaluable pts had a CR or PR, including 4 sustained CRs. While there was no acute infusional toxicity, all responding pts also developed CRS. All had high fevers, as well as grade 3 or 4 hypotension/hypoxia. CRS preceded peak blood expression of CART19 cells, and then increased in intensity until the CART19 cell peak (D10–31 after infusion). The ALL pt experienced the most significant toxicity, with grade 4 hypotension and respiratory failure. Steroid therapy on D6 resulted in no improvement. On D9, noting high levels of TNFa and IL-6 (peak increases above baseline: IFNg at 6040x; IL-6 at 988x; IL-2R at 56x, IL-2 at 163× and TNFa at 17x), we administered TNFa and IL-6 antagonists entanercept and toc. This resulted in resolution of fever and hypotension within 12hr and a rapid wean from ventilator support to room air. These interventions had no apparent impact on CART19 cell expansion or efficacy: peak of CAR T cells (2539 CAR+ cells/uL; 77% of CD3 cells by flow) occurred on D11, and D23 bone marrow showed CR with negative MRD, compared to her initial on-study marrow which showed 65% blasts. Although she had no history of CNS ALL, spinal fluid showed detectable CART19 cells (21 lymphs/mcL; 78% CAR+). At 4mo post infusion, this pt remains in CR, with 17 CART19 cells/uL in the blood and 31% CAR+ CD3 cells in the marrow. Clinical assessment of subsequent responding patients shows all had evidence of MAS/HLH including dramatic elevations of ferritin and histologic evidence of HLH. Peak ferritin levels range from 44,000 to 605,000, preceding and continuing with peak T cell proliferation. Other consistent findings include rapid onset hepatosplenomegaly unrelated to disease and moderate DIC. Subsequently, 3 CLL patients have also been treated with toc, also with prompt and striking resolution of high fevers, hypotension and hypoxia. 1 received toc on D10 and achieved a CR accompanied by CART19 expansion. 1 had rapid resolution of CRS following toc administration on day 9 and follow up for response is too short. A 3rd CLL pt received toc on D3 for early fevers and had no CART-19 proliferation and no response. To model the timing of cytokine blockade, xenografts using bioluminescent primary pediatric ALL were established and then treated with extra cells from the clinical manufacture. The CART19 cells proliferated and resulted in prolonged survival. Cytokine blockade prior to T cell infusion with toc and/or etanercept abrogated disease control with less in vivo proliferation of infused CART19 cells, confirming the result seen in the one pt given early toc (D3). The optimal time and threshold to trigger cytokine blockade is currently being tested in these models. CART19 T cells can produce massive in-vivo expansion, long-term persistence, and anti-tumor efficacy, but can also induce significant CRS with features suggestive of MAS/HLH that responds rapidly to cytokine blockade. Given prior to initiation of significant CART19 proliferation, blockade of TNFa and/or IL-6 may interfere with proliferation and effector function, but if given at a point where cell proliferation is underway, toc may ameliorate the symptoms that we have observed correlate with robust clinical responses. Disclosures: Off Label Use: tocilizumab for cell therapy toxicity. Levine:University of Pennsylvania: financial interest due to intellectual property and patents in the field of cell and gene therapy. Conflict of interest is managed in accordance with University of Pennsylvania policy and oversight Patents & Royalties; TxCell: Consultancy, Membership on an entity's Board of Directors or advisory committees. Kalos:University of Pennsylvania: Patents & Royalties. June:Novartis: Research Funding, institution owned patents have been licensed by Novartis, institution owned patents have been licensed by Novartis Patents & Royalties.

scholarly journals Selectively targeting myeloid-derived suppressor cells through TRAIL receptor 2 to enhance the efficacy of CAR T cell therapy for treatment of breast cancer

2021 ◽  
Vol 9 (11) ◽  
pp. e003237
Author(s):  
Saisha A Nalawade ◽  
Paul Shafer ◽  
Pradip Bajgain ◽  
Mary K McKenna ◽  
Arushana Ali ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Nuclear Translocation ◽  
Suppressor Cells ◽  
T Cell Proliferation ◽  
Myeloid Derived Suppressor Cells ◽  
Tumor Site ◽  
Costimulatory Receptor ◽  
Car T Cells ◽  
Car T

BackgroundSuccessful targeting of solid tumors such as breast cancer (BC) using chimeric antigen receptor (CAR) T cells has proven challenging, largely attributed to the immunosuppressive tumor microenvironment (TME). Myeloid-derived suppressor cells (MDSCs) inhibit CAR T cell function and persistence within the breast TME. To overcome this challenge, we have developed CAR T cells targeting tumor-associated mucin 1 (MUC1) with a novel chimeric costimulatory receptor that targets tumor necrosis factor–related apoptosis-inducing ligand receptor 2 (TR2) expressed on MDSCs.MethodsThe function of the TR2.41BB costimulatory receptor was assessed by exposing non-transduced (NT) and TR2.41BB transduced T cells to recombinant TR2, after which nuclear translocation of NFκB was measured by ELISA and western blot. The cytolytic activity of CAR.MUC1/TR2.41BB T cells was measured in a 5-hour cytotoxicity assay using MUC1+ tumor cells as targets in the presence or absence of MDSCs. In vivo antitumor activity was assessed using MDSC-enriched tumor-bearing mice treated with CAR T cells with or without TR2.41BB.ResultsNuclear translocation of NFκB in response to recombinant TR2 was detected only in TR2.41BB T cells. The presence of MDSCs diminished the cytotoxic potential of CAR.MUC1 T cells against MUC1+ BC cell lines by 25%. However, TR2.41BB expression on CAR.MUC1 T cells induced MDSC apoptosis, thereby restoring the cytotoxic activity of CAR.MUC1 T cells against MUC1+ BC lines. The presence of MDSCs resulted in an approximately twofold increase in tumor growth due to enhanced angiogenesis and fibroblast accumulation compared with mice with tumor alone. Treatment of these MDSC-enriched tumors with CAR.MUC1.TR2.41BB T cells led to superior tumor cell killing and significant reduction in tumor growth (24.54±8.55 mm3) compared with CAR.MUC1 (469.79±81.46 mm3) or TR2.41BB (434.86±64.25 mm3) T cells alone. CAR.MUC1.TR2.41BB T cells also demonstrated improved T cell proliferation and persistence at the tumor site, thereby preventing metastases. We observed similar results using CAR.HER2.TR2.41BB T cells in a HER2+ BC model.ConclusionsOur findings demonstrate that CAR T cells that coexpress the TR2.4-1BB receptor exhibit superior antitumor potential against breast tumors containing immunosuppressive and tumor promoting MDSCs, resulting in TME remodeling and improved T cell proliferation at the tumor site.

Immunodulatory Capacity of Mesenquimal STROMAL CELLS IS CONTROLLED by NON-Canonical NF-Kb PATHWAY

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4865-4865
Author(s):  
Rodrigo Haddad ◽  
Felipe Saldanha-araujo ◽  
Amélia Araujo ◽  
Dimas T Covas ◽  
Marco A. Zago ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Stromal Cells ◽  
Transfection Efficiency ◽  
T Cell Proliferation ◽  
Human Mscs ◽  
Activated T Cells ◽  
Cell Based Therapy

Abstract Abstract 4865 Introduction: Mesenquimal Stromal Cells (MSCs) possess immunosuppressive properties, becoming these cells a promising subject of study for future approaches in cell-based therapy. During co-culture with activated lymphocytes and probably through T-cell derived cytokines, MSCs are activated in order to become suppressive. Once MSCs are triggered, they acquire an inhibitory profile increasing the release of immunoregulatory factors, such as IDO, IFNγ, PGE2, NO and TGF-β, responsible for T-cell inhibition. In addition, expression of adhesion molecules, such as ICAM-1, and generation of regulatory T cells subsets, such as CD69+ T cells, are important in immunosuppressive property of human MSCs. However, the exact mechanisms underlying the immunomodulatory functions of MSCs remain largely unknown. NF-kB comprises a family of inducible transcription factors that serve as important regulators of the host immune and inflammatory responses. The NF-kB signals are activated via canonical (mediated mainly by RelA-p50) and/or non-canonical (mediated by RelB-p52) pathways in response to diverse stimuli. Given the immunomodulatory properties of MSCs and the possible involvement of NF-Bk pathway in this effect, here, it was explored the role of non-canonical NF-KB signaling in the immunomodulatory capacity of MSCs cocultured with activated T cells. Methods: siRNAs targeting RelB were transfected into MSC with lipofectamine. siRNAs C- were used as negative control and siRNA-FITC as transfection control. After 24 hours, immunomagnetically purified CD3+ T cells were stained with CFSE, activated by anti-CD2/CD3/CD28 beads and cultured in the presence of MSCs. After 3 days, flow cytometry was performed to observe the transfection efficiency, T cell proliferation and percentage of CD69+ T regulatory cells. Additionally, RNA from MSCs was extracted and RelB and ICAM-1 mRNA expression were quantified by Real-time PCR. Results: The transfection efficiency was around 75% and RelB mRNA level was reduced by 80% in MSCs transfected with siRNA RelB compared to siRNAs C- transfected cells. Compared to MSCs previously transfected with siRNA C- and co-cultured with activated T cells, MSCs transfected with siRNA RelB resulted increasing of 22% in T cell proliferation and decreasing of 9,2% and 30% in the CD69+ regulatory T cells generation and ICAM-1 expression respectively. Conclusion: Non-canonical NF-kB pathway, mediated by RelB, may be partially involved in acquisition of the inhibitory profile by decreasing T cells proliferation, and increasing the expression of ICAM-1 and the generation of CD69+ regulatory T cells. Disclosures: No relevant conflicts of interest to declare.

A Novel Strategy of Switching on/Off CD19CAR Expression Under Tetracycline-Based System

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4424-4424
Author(s):  
Reona Sakemura ◽  
Seitaro Terakura ◽  
Keisuke Watanabe ◽  
Kotaro Miyao ◽  
Daisuke Koyama ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Research Funding ◽  
T Cell Proliferation ◽  
Target Cells ◽  
Car T Cells ◽  
Car T ◽  
Release Assay

Abstract Introduction: Genetic modification of T cells with chimeric antigen receptor (CAR) has emerged with astonishing treatment outcomes for B cell malignancies. Clinical trials of CAR-T therapy demonstrated toxicities such as hypogammaglobulinemia due to B cell aplasia or hemophagocytic syndrome after overactivation of CAR-T cells. These toxicities are considered as major drawbacks for broader application of CAR-T therapy. To overcome these serious adverse events, further modification of CAR-T technology to control CAR expression arbitrary is needed. Therefore we aimed to develop inducible CAR expressing T cells based on tetracycline-regulation system. Methods: We developed a novel inducible CD19CAR system by infusing anti-CD19-CD3z-CD28-tEGFR into pRetroX-TetOne vector (Tet-19CAR). By using Tet-19CAR transduced SUPT1 (T cell line), expression and disappearance kinetics of CAR were determined. We also retrovirally transduced Tet-19CAR into human CD8+ T cells, and achieved more than 90% purity of CAR positive T cells after a selection with anti-EGFR mAb. These CAR-T cells were again expanded with anti-CD3/28 beads and used in 51 Cr release assay, coculture assay, cytokine release assay and T cell proliferation assay. Regarding coculture assay, CD19 transduced K562-CD19 (K562-CD19) was labeled with 0.1 nM CFSE and plated with CAR-T cells at a ratio of 1:1 without IL-2 supplementation and incubated for 96 hours. Finally we examined this system in NOG mice. We injected 0.5 x 106 Raji-ffluc (fire-fly luciferase) followed by 5.0 x 106 CAR-T cells from the tail vein, then we evaluated the tumor flux by in vivo imaging system on days 7, 14, 21, and 30. Results: With more than 100 ng/mL of Doxycycline (Dox), CD19CAR was successfully expressed on both of SUPT1 and CD8+ T cells. For maximum and minimum expression, 24 and 72 hours were needed after addition and discontinuation of Dox, respectively. To determine the cytotoxicity of Tet-19CAR-T cells according to presence or absence of Dox, we performed 51 Cr release assay and coculture assay against K562-CD19. In the presence of Dox, Tet-19CAR showed an equivalent lytic activity to conventional CD19CAR-T cells (c19CAR). In contrast, Tet-19CAR without Dox exhibited significantly lower cytotoxicity against CD19+ target cells. (Dox (-) Tet-19CAR, Dox (+) Tet-19CAR and c19CAR: 14.0±4.0%, 38.0±4.0% and 37.0±2.0% at an E:T ratio = 10:1, respectively). In the coculture assay, Tet-19CAR with Dox eradiated K562-CD19, while they failed to suppress the target cells without Dox. In the intracellular IFN-g assay against K562-CD19, a similar proportion of responder was IFN-g + in Tet-19CAR with Dox and c19CAR. On the other hand, a significantly low proportion of IFN-g + cells were observed in Tet-19CAR without Dox. (Dox (-) Tet-19CAR, 1.0%±0%, Dox (+) Tet-19CAR, 19.1%±6.0% and c19CAR 21.5%±4.0%, respectively) Similar to intracellular IFN-g assay, ELISA revealed that Tet-19CAR with Dox and c19CAR produced IL-2 and IFN-g equally well. However, Tet-19CAR without Dox hardly did. [IL-2 (ng/ml): Dox (-) Tet-19CAR, 1.00±0.060, Dox (+) Tet-19CAR, 9.25±0.30 and c19CAR 8.75±0.68; IFN-g (ng/ml): 2.32±1.24, 57.96±6.95 and 62.42±5.95] (Fig). We next analyzed CAR-T cell proliferation upon stimulation with K562-CD19 over 96 hours. Tet-19CAR with Dox showed 6-7 fold expansion, whereas Tet-19CAR without Dox failed to proliferate. Regarding in vivo model, the mice treated with c19CAR or Tet-19CAR with Dox showed significantly low tumor flux but the mice treated with Tet-19CAR without Dox showed higher tumor burden at day 21 of CAR-T cell infusion [Photons/sec: Dox (-) Tet-19CAR, 2.5 x 1010, Dox (+) Tet-19CAR, 6.4 x 108 and c19CAR, 8.4 x 108 ]. Conclusions: We generated tetracycline-inducible CAR-T cells and successfully controlled the CAR expression with Dox administration. Tet-19CAR without Dox still demonstrated some CD19CAR expression and subsequent cytotoxicity against CD19 positive cells. Nonetheless the CAR expression level of Tet-19CAR without Dox was lower than the threshold for exhibiting positive responses in the function assays such as cytokine production and proliferation. This phenomenon was also confirmed in the xenograft model. To regulate CAR expression more precisely and pursue clinical translations in combinations with other CARs, further efforts are needed to reduce any leaky CAR expression by modification of the system. Figure 1. Figure 1. Disclosures Kiyoi: Pfizer Inc.: Research Funding; Eisai Co., Ltd.: Research Funding; Yakult Honsha Co.,Ltd.: Research Funding; Alexion Pharmaceuticals: Research Funding; MSD K.K.: Research Funding; Takeda Pharmaceutical Co., Ltd.: Research Funding; Taisho Toyama Pharmaceutical Co., Ltd.: Research Funding; Teijin Ltd.: Research Funding; Astellas Pharma Inc.: Consultancy, Research Funding; Japan Blood Products Organization: Research Funding; Nippon Shinyaku Co., Ltd.: Research Funding; FUJIFILM RI Pharma Co.,Ltd.: Research Funding; Nippon Boehringer Ingelheim Co., Ltd.: Research Funding; FUJIFILM Corporation: Patents & Royalties, Research Funding; Zenyaku Kogyo Co., Ltd.: Research Funding; Sumitomo Dainippon Pharma Co., Ltd.: Research Funding; Kyowa Hakko Kirin Co., Ltd.: Consultancy, Research Funding; Bristol-Myers Squibb: Research Funding; Chugai Pharmaceutical Co., Ltd.: Research Funding; Novartis Pharma K.K.: Research Funding; Mochida Pharmaceutical Co., Ltd.: Research Funding.

scholarly journals CD38 Chimeric Antigen Receptor Engineered T Cells As Therapeutic Tools for Multiple Myeloma

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4759-4759 ◽  
Author(s):  
Esther Drent ◽  
Richard Groen ◽  
Willy A. Noort ◽  
Jeroen Lammerts van Bueren ◽  
Paul W.H.I. Parren ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Cells ◽  
T Cell ◽  
Research Funding ◽  
Suicide Gene ◽  
Antigen Recognition ◽  
Car T Cells ◽  
Cd38 Expression ◽  
Car T ◽  
Signaling Domains

Abstract Chimeric Antigen Receptors (CARs) are engineered constructs consisting of an antibody-derived antigen recognition domain linked to intracellular T cell signaling domains. Cytotoxic T cells transduced to express tumor-reactive CARs are highly promising tools for immunotherapy of cancer. The CD38 molecule, with its high and homogenous expression on Multiple Myeloma (MM) tumor cells, is considered a suitable target for antibody therapy of MM. Prompted by this, we evaluated the feasibility and efficacy of targeting MM cells with CD38-CAR-transduced T cells (CD38-CAR T cells). To this end, we generated three different retroviral CAR constructs based on human CD38 antibodies as antigen recognition domain, CD3zeta and 41BB (CD137) as signaling domains and transduced them into PBMCs of a healthy donor. After in vitro selection and expansion, all CD38-CAR T cells, either unsorted or CD4/CD8 sorted, effectively lysed MM cell lines in a dose-, and CD38 expression-dependent manner, with a better efficacy for the CD8+ fraction. CD38-CAR T cells also effectively eradicated primary MM cells in the bone marrow mononuclear cells derived from MM patients, indicating their clinical relevancy. Although CD38-CAR T cells also displayed cytotoxic activity against the CD38+ fraction of mature monocytes and NK cells and to a lesser extent CD38+ B and T cells, they did not affect the outgrowth of CD34+ cells into various myeloid lineages. In addition,CD38-CAR T cell activity was effectively controllable by transducing them with a caspase 9-based inducible suicide gene. More interestingly, we discovered that the CD38-CAR T cells were themselves devoid of CD38 surface expression, indicating that CD38 was not essential for T cell expansion and function. Finally, in a novel in vivo xenotransplant model (UM9 cell line), in which myeloma cells were grown in a humanized bone marrow microenvironment, i.v. as well as intra tumor administration of CD38-CAR T cells established significant anti-tumor effects, proving that CD38-CAR endowed cytotoxic T lymphocytes, even with no CD38 expression, can efficiently migrate, infiltrate and eliminate human MM tumors growing in their natural niche. These results demonstrate the feasibility and potency of CAR mediated targeting of CD38+ MM cells. Optimization of CD38-CAR and suicide-gene control of CD38 CAR T cellsmay provide next steps towards safe clinical implementation of CD38-CAR T cell immune therapy. Disclosures Drent: Genmab BV: Guest employee (unpaid) Other. Lammerts van Bueren:Genmab : Employment. Parren:Genmab: Employment, Equity Ownership. van de Donk:Genmab BV: Research Funding; J&J: Research Funding; Celgene: Research Funding. Martens:Genmab BV: Research Funding; J&J: Research Funding; Celgene: Research Funding. Lokhorst:Celgene: Research Funding; J&J: Research Funding; Genmab: Research Funding. Mutis:Celgene: Research Funding; J&J: Research Funding; Genmab BV: Research Funding.

Safe and Effective Re-Induction Of Complete Remissions In Adults With Relapsed B-ALL Using 19-28z CAR CD19-Targeted T Cell Therapy

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 69-69 ◽  
Author(s):  
Marco L Davila ◽  
Isabelle Riviere ◽  
Xiuyan Wang ◽  
Shirley Bartido ◽  
Jolanta Stefanski ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Phase I ◽  
B Cell ◽  
Deep Sequencing ◽  
Residual Disease ◽  
The Other ◽  
Car T Cells ◽  
Cell Based Therapy ◽  
Car T

Abstract Although most adults diagnosed with B-ALL can be induced into a first complete remission (CR1 rates of approximately 80-90%), a majority will relapse and develop chemorefractory disease. Novel, non-chemotherapy based treatments are needed for this group of patients. We have developed a novel CD19-targeted T cell-based therapy for patients with B cell malignancies. We isolate T cells from patients with relapsed/refractory B-ALL and genetically modify them with a chimeric antigen receptor (CAR) construct, termed 19-28z, which comprises a CD19 binding domain fused to the signaling domains of the CD28 costimulatory receptor and the ζ chain of the CD3 complex. CD19 is a universal B cell antigen expressed on all normal and malignant B cells. Expression of the 19-28z CAR by a T cell promotes binding of the CD19 antigen and triggers cytotoxicity, cytokine release and proliferation upon engagement of CD19. We are conducting a Phase I clinical protocol in adults with relapsed/refractory B-ALL (NCT01044069). Enrolled patients are leukapheresed and then re-induced with salvage chemotherapy. A T cell infusion (3 x 106 19-28z CAR T cells/kg) is administered following conditioning chemotherapy. We now describe the results from this Phase I protocol. Thirteen adults have been treated to date: eleven were enrolled with relapsed/refractory disease, while the other 2 were enrolled and leukapheresed during CR1, but not treated until they relapsed. The age of the patients ranged from 23 to 74 with a median age of 42. Three of the 13 patients had Philadelphia-chromosome positive B-ALL, which is considered the most negative genetic risk factor for adults with B-ALL. We were able to achieve the required T cell dose, despite collecting patients with high blast counts or marked lymphopenias, in all but one patient. Seven of the 13 patients were infused with 19-28z CAR T cells while they had gross residual disease (>5 to 70% blasts in the BM). The remaining patients had MRD, detected by flow cytometry or deep sequencing, at the time of 19-28z CAR T cell infusion. Six patients developed toxicities including high-grade fevers (>40°C), hypotension, hypoxia, mental status changes, and seizures. These episodes ran for approximately one week before they were halted by treatment with steroids or tocilizumab. The other 7 patients did not experience toxicities. All patients completely recovered and were able to leave the hospital. The occurrence of toxicities correlated with tumor burden so that patients with gross residual disease (>5% blasts in BM) developed toxicities, while patients with MRD had no evidence of toxicities. Ten out of the 12 patients with detectable disease before T cell infusion developed MRD- responses such that 5 of the patients with gross residual disease (blasts > 5% in BM) became MRD- and 5 MRD+ patients became MRD-. The rapidity of the responses was quite remarkable with MRD- results obtained as early as 7 -14 days after T cell infusion. Furthermore, despite the poor predicted outcomes of relapsed Ph+ B-ALL we were able to get these patients MRD-, as detected by both deep sequencing for the IgH rearrangement and qPCR for the bcr-abl transcript. Conclusion The potent induction of MRD- responses and the reversibility of toxicities occurring in a subset of patients strongly support integrating this therapy in the modern paradigm for B-ALL therapy, by facilitating access to allogeneic-stem cell transplantation (SCT) for patients with relapsed B-ALL. Indeed, out of the 13 patients treated to date, 4 underwent an allo-SCT and 5 are being prepared for an allo-SCT. Of the remaining 4 patients, 1 is a non-responder (a patient with extramedullary disease, but no disease in the bone marrow), 1 relapsed (not eligible for allo-SCT), 1 is in CR2 (medically not eligible for allo-SCT), and 1 has not been evaluated for treatment response yet. Overall, the results from this Phase I protocol demonstrate that toxicities associated with this therapy are predictable and manageable. Furthermore, the remarkable MRD- re-induction rate and facilitation for allo-SCT we report here warrant further evaluation of this therapy in a Phase II protocol. Disclosures: No relevant conflicts of interest to declare.

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