Identification of Small Molecule Modulators to Enhance the Therapeutic Properties of Chimeric Antigen Receptor T Cells

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4712-4712
Author(s):  
Jonathan Rosen ◽  
Betsy Rezner ◽  
David Robbins ◽  
Ian Hardy ◽  
Eigen Peralta ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Small Molecules ◽  
Small Molecule ◽  
Employment Equity ◽  
Car T Cells ◽  
Car T ◽  
Equity Ownership

Abstract Adoptive cellular therapies using engineered chimeric antigen receptor T cells (CAR-T cells) are rapidly emerging as a highly effective treatment option for a variety of life-threatening hematological malignancies. Small molecule-mediated modulation of T cell differentiation during the in vitro CAR-T manufacturing process has great potential as a method to optimize the therapeutic potential of cellular immunotherapies. In animal models, T cells with a central or stem memory (TCM/SCM) phenotype display enhanced in vivoefficacy and persistence relative to other T cell subpopulations. We sought to identify small molecules that promote skewing towards a TCM/SCM phenotype during the CAR-T manufacturing process, with associated enhanced viability, expansion and metabolic profiles of the engineered cells. To this end, we developed a high-throughput functional screening platform with primary human T cells using a combination of high-content immunophenotyping and gene expression-based readouts to analyze cells following a high-throughput T cell culture platform that represents a scaled-down model of clinical CAR-T cell production. Multicolor flow cytometry was used to measure expansion, cell viability and the expression levels of cell surface proteins that define TCM cells (e.g., CCR7, CD62L and CD27) and markers of T cell exhaustion (e.g., PD1, LAG3, and TIM3). In parallel, a portion of each sample was evaluated using high content RNA-Seq based gene expression analysis of ~100 genes representing key biological pathways of interest. A variety of known positive and negative control compounds were incorporated into the high-throughput screens to validate the functional assays and to assess the robustness of the 384-well-based screening. The ability to simultaneously correlate small molecule-induced changes in protein and gene expression levels with impacts on cell proliferation and viability of various T cell subsets, enabled us to identify multiple classes of small molecules that favorably enhance the therapeutic properties of CAR-T cells. Consistent with results previously presented by Perkins et al. (ASH, 2015), we identified multiple PI3K inhibitors that could modify expansion of T cells while retaining a TCM/SCM phenotype. In addition, we identified small molecules, and small molecule combinations, that have not been described previously in the literature that could improve CAR-T biology. Several of the top hits from the screens have been evaluated across multiple in vitro (e.g., expansion, viability, CAR expression, serial restimulation/killing, metabolic profiling, and evaluation of exhaustion markers) and in vivo (e.g., mouse tumor models for persistence and killing) assays. Results from the initial screening hits have enabled us to further refine the optimal target profile of a pharmacologically-enhanced CAR-T cell. In addition, we are extending this screening approach to identify small molecules that enhance the trafficking and persistence of CAR-T cells for treating solid tumors. In conclusion, the approach described here identifies unique small molecule modulators that can modify CAR-T cells during in vitro expansion, such that improved profiles can be tracked and selected from screening through in vitro and in vivo functional assays. Disclosures Rosen: Fate Therapeutics: Employment, Equity Ownership. Rezner:Fate Therapeutics, Inc: Employment, Equity Ownership. Robbins:Fate Therapeutics: Employment, Equity Ownership. Hardy:Fate Therapeutics: Employment, Equity Ownership. Peralta:Fate Therapeutics: Employment, Equity Ownership. Maine:Fate Therapeutics: Employment, Equity Ownership. Sabouri:Fate Therapeutics: Employment, Equity Ownership. Reynal:Fate Therapeutics: Employment. Truong:Fate Therapeutics: Employment, Equity Ownership. Moreno:Fate Therapeutics, Inc.: Employment, Equity Ownership. Foster:Fate Therapeutics: Employment, Equity Ownership. Borchelt:Fate Therapeutics: Employment, Equity Ownership. Meza:Fate Therapeutics: Employment, Equity Ownership. Thompson:Juno Therapeutics: Employment, Equity Ownership. Fontenot:Juno Therapeutics: Employment, Equity Ownership. Larson:Juno Therapeutics: Employment, Equity Ownership. Mujacic:Juno Therapeutics: Employment, Equity Ownership. Shoemaker:Fate Therapeutics: Employment, Equity Ownership.

scholarly journals Prophylactic Itacitinib (INCB039110) for the Prevention of Cytokine Release Syndrome Induced By Chimeric Antigen Receptor T-Cells (CAR-T-cells) Therapy

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1934-1934 ◽  
Author(s):  
Eduardo Huarte ◽  
Roddy S O'Connor ◽  
Melissa Parker ◽  
Taisheng Huang ◽  
Michael C. Milone ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cytokine Release ◽  
Employment Equity ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T ◽  
Equity Ownership

Background: T-cells engineered to express a chimeric antigen receptor (CAR-T-cells) are a promising cancer immunotherapy. Such targeted therapies have shown long-term relapse survival in patients with B cell leukemia and lymphoma. However, cytokine release syndrome (CRS) represents a serious, potentially life-threatening, side effect often associated with CAR-T cells therapy. The Janus kinase (JAK) tyrosine kinase family is pivotal for the downstream signaling of inflammatory cytokines, including interleukins (ILs), interferons (IFNs), and multiple growth factors. CRS manifests as a rapid (hyper)immune reaction driven by excessive inflammatory cytokine release, including IFN-g and IL-6. Itacitinib is a potent, selective JAK1 inhibitor which is being clinically evaluated in several inflammatory diseases. Aims: To evaluate in vitro and in vivo the potential of itacitinib to modulate CRS without impairing CAR-T cell anti-tumor activity. Materials and Methods: In vitro proliferation and cytotoxic activity of T cells and CAR-T cells was measured in the presence of increasing concentrations of itacitinib or tocilizumab (anti-IL-6R). To evaluate itacitinib effects in vivo, we conducted experiments involving adoptive transfer of human CD19-CAR-T-cells in immunodeficient animals (NSG) bearing CD19 expressing NAMALWA human lymphoma cells. The effect of itacitinib on cytokine production was studied on CD19-CAR-T-cells expanded in the presence of itacitinib or tocilizumab. Finally, to study whether itacitinib was able to reduce CRS symptoms in an in vivo setting, naïve mice were stimulated with Concanavalin-A (ConA), a potent T-cell mitogen capable of inducing broad inflammatory cytokine releases and proliferation. Results: In vitro, itacitinib at IC50 relevant concentrations did not significantly inhibit proliferation or anti-tumor killing capacity of human CAR-T-cells. Itacitinib and tocilizumab (anti-IL-6R) demonstrated a similar effect on CAR T-cell cytotoxic activity profile. In vivo, CD19-CAR-T-cells adoptively transferred into CD19+ tumor bearing immunodeficient animals were unaffected by oral itacitinib treatment. In an in vitro model, itacitinib was more effective than tocilizumab in reducing CRS-related cytokines produced by CD19-CAR-T-cells. Furthermore, in the in vivo immune hyperactivity (ConA) model, itacitinib reduced serum levels of CRS-related cytokines in a dose-dependent manner. Conclusion: Itacitinib at IC50 and clinically relevant concentrations did not adversely impair the in vitro or in vivo anti-tumor activity of CAR-T cells. Using CAR-T and T cell in vitro and in vivo systems, we demonstrate that itacitinib significantly reduces CRS-associated cytokines in a dose dependent manner. Together, the data suggest that itacitinib may have potential as a prophylactic agent for the prevention of CAR-T cell induced CRS. Disclosures Huarte: Incyte corporation: Employment, Equity Ownership. Parker:Incyte corporation: Employment, Equity Ownership. Huang:Incyte corporation: Employment, Equity Ownership. Milone:Novartis: Patents & Royalties: patents related to tisagenlecleucel (CTL019) and CART-BCMA; Novartis: Research Funding. Smith:Incyte corporation: Employment, Equity Ownership.

scholarly journals ALLO-715, an Allogeneic BCMA CAR T Therapy Possessing an Off-Switch for the Treatment of Multiple Myeloma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 591-591 ◽  
Author(s):  
Cesar Sommer ◽  
Bijan Boldajipour ◽  
Julien Valton ◽  
Roman Galetto ◽  
Trevor Bentley ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Expansion ◽  
Employment Equity ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T ◽  
Equity Ownership

Abstract Autologous chimeric antigen receptor (CAR) T cells targeting B-Cell Maturation Antigen (BCMA) have demonstrated promising clinical activity, inducing durable responses in patients with relapsed/refractory multiple myeloma (MM). Development of autologous CAR T therapies is however limited by logistical challenges and the time required for manufacturing, which has to be done for each patient. In addition, manufacturing may not be feasible in some patients. An allogeneic approach that utilizes engineered cells from a healthy donor could potentially expand patient access to these therapies by providing a readily available off-the-shelf product. We have previously described the screening of a library of single chain variable fragments (scFvs) with high affinity to human BCMA and the identification of candidate BCMA CARs with potent antitumor activity. Here we sought to further characterize ALLO-715, our lead allogeneic BCMA CAR T cell product, for its specificity to human BCMA, antitumor efficacy in vitro using a long-term killing assay and in xenograft mouse models with physiologic levels of human IL-7 and IL-15, and suitability for scale-up manufacturing. Allogeneic ALLO-715 CAR T cells were generated by lentiviral transduction with a second generation CAR construct incorporating a novel scFv derived from a fully-human antibody with high affinity to BCMA (KD value ~ 5 nM, determined at 37°C) and featuring a rituximab-driven off-switch. Transduced T cells were then transfected with mRNAs encoding Transcription Activator-Like Effector Nucleases (TALEN®) designed to specifically disrupt the T cell receptor alpha chain and CD52 loci. These modifications result in a cell product with a lower risk of TCR-mediated graft-versus-host disease and resistance to the CD52 antibody alemtuzumab, a lymphodepleting agent. BCMA CAR T cells exhibited robust cell expansion, with low levels of tonic signaling that resulted in minimal differentiation (> 50% Tscm/Tcm phenotype). In in vitro assays, ALLO-715 CAR T cells displayed potent cytotoxic activity when co-cultured with the target cell lines MM.1S, Molp-8, and BCMA-REH but negligible cytotoxicity against BCMA-negative REH cells. The high proliferative potential indicated by the high frequency of memory T cells was validated in long-term killing assays, where ALLO-715 CAR T cells showed substantial expansion in the presence of MM.1S cells with no evidence of exhaustion or diminished cytolytic activity after seven days of continuous exposure to target. The potency of ALLO-715 CAR T cells was unaffected by high concentrations of soluble BCMA (>10 ug/mL), which has been shown previously to interfere with the activity of some BCMA-specific CARs. In MM xenograft mouse models, ALLO-715 CAR T cells were highly efficacious at single dose. High serum IL-15 levels have been associated with CAR T cell expansion in clinical trials. To evaluate the impact of homeostatic cytokines on CAR T cell survival and antitumor activity in our xenograft models, mice were administered adeno-associated viruses (AAV) for the expression of human IL-7 and IL-15. In the presence of physiological concentrations of these cytokines, enhanced BCMA CAR T cell expansion and anti-tumor activity were observed. To assess potential off-target interactions of ALLO-715 CAR, tissue cross-reactivity studies were carried out on standard human tissue panels using a scFv-human IgG fusion protein. Consistent with the limited expression pattern of BCMA, reactivity was seen on scattered cells in lymphoid tissues such as tonsil and abundantly on BCMA-expressing cell lines, but no appreciable staining was detected in other tissues. We examined BCMA CAR T cells manufactured following a proprietary GMP-like clinical scale process and found that cell expansion and viability, T cell phenotype and in vivo antitumor efficacy were preserved. These results demonstrate the potential of ALLO-715 as a novel allogeneic BCMA CAR T therapy for the treatment of relapsed/refractory MM and other BCMA-positive malignancies. Disclosures Sommer: Allogene Therapeutics: Employment, Equity Ownership, Patents & Royalties. Boldajipour:Pfizer Inc.: Employment, Patents & Royalties. Valton:Cellectis.Inc: Employment, Equity Ownership, Patents & Royalties. Galetto:Cellectis SA: Employment, Equity Ownership, Patents & Royalties. Bentley:Allogene Therapeutics: Employment, Equity Ownership. Sutton:Allogene Therapeutics: Employment, Equity Ownership. Ni:Allogene Therapeutics: Employment, Equity Ownership. Leonard:Allogene Therapeutics: Employment, Equity Ownership. Van Blarcom:Allogene Therapeutics: Employment, Equity Ownership. Smith:Cellectis. Inc: Employment, Patents & Royalties. Chaparro-Riggers:Pfizer Inc.: Employment, Patents & Royalties. Sasu:Allogene Therapeutics: Employment, Equity Ownership, Patents & Royalties.

scholarly journals Preclinical Evaluation of Human Placental-Derived Allogeneic CD19 CAR-T Cells Against B Cell Malignancies

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3222-3222
Author(s):  
Kathy Karasiewicz ◽  
Shuyang He ◽  
Mary Ng ◽  
Kristina Tess ◽  
Weifang Ling ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Receptor ◽  
Employment Equity ◽  
Car T Cells ◽  
Car T ◽  
Equity Ownership ◽  
Robust Process

Celularity, Inc. is developing a CD19 CAR-T Cell therapy using an allogeneic platform derived from postpartum human placental cells. T cells isolated from placenta/ umbilical cord blood and genetically modified to express CD19 chimeric antigen receptor (CAR), termed Placental-derived (P-) CD19 CAR T cells, are in development for the treatment of B cell malignancies. Unlike adult peripheral blood mononuclear cell (PBMC)-derived T cells, P-T cells are mostly naïve (CD45RA+) and can be readily expanded while maintaining an earlier differentiation phenotype such as greater expression of naïve/ memory markers, lower expression of effector/ exhaustion markers, allowing for greater proliferative potential of these cells ex vivo. These cells are also known to have greater immune tolerance to HLA mismatch and display impaired allogeneic activation, contributing to lower incidences of severe graft-verse-host disease (GvHD) (Barker, et. al. Blood, 2001; Chen, et al. Biology of Blood and Marrow Transplantation, 2006), making them an attractive cell population for use as an allogeneic, adoptive cell therapy. A robust process for the isolation, transduction, and expansion of placental-derived T cells to generate "off-the-shelf" allogeneic P-CD19 CAR T cells was developed. Twenty-One day expanded, non-modified P-T cells (N=3) were compared to adult PBMCs for their allo-reactivity in a Xenogeneic GvHD model in NCG mice. P-T cells did not induce xeno-GvHD whereas PBMCs did, as evidenced by significant weight loss and death of all mice (N=5) by Day 28 post infusion. Despite expanded P-T cells demonstrating lack of in vivo GvHD, current manufacture of P-CD19 CAR T cells does include a CRISPR-mediated T-cell receptor a constant (TRAC) knockout (KO) step as an additional risk-mitigation strategy to circumvent any potential GvHD stemming from expression of endogenous T cell receptor. CD19 CAR transduction using a retrovirus provided by Sorrento Therapeutics, Inc., followed by TRAC knockout with CRISPR results in both high efficiency of CD19 CAR expression (~30% CD19 Fc+) and TCR KO (>96% CD3-/ TCR a/b-). In vitro, the functional activity of P-CD19 CAR-TRAC KO T cells against CD19+ Burkitt's Lymphoma (Daudi) and Acute lymphoblastic Leukemia (NALM6) cell lines was assessed in cytotoxicity and cytokine release assays. P-CD19 CAR T cells specifically lyse CD19+ Daudi/ Nalm6 targets in both 4-hour endpoint FACS and ACEA kinetic cytotoxicity assays, and in most cases at levels equivalent to or greater than PBMC-derived CD19 CAR T cells. When P-CD19 CAR T cells were co-cultured with CD19+ Daudi/ Nalm6 target cells for 24-hours, they secreted pro-inflammatory cytokines and effector proteins in an antigen-specific manner. In vivo, the anti-tumor activity of P-CD19 CAR T cells was assessed using a disseminated lymphoma xenograft model in NSG mice. Luciferase expressing Daudi cells (3×106) were intravenously (IV) injected on Day 0, followed by IV injection of P-CD19 CAR T cells (14×106) on Day 7. Bioluminescence Imaging (BLI) and survival were used as primary study endpoints. P- CD19 CAR T cells were well tolerated and safe. P-CD19 CAR T cells significantly reduced tumor burden, and improved survival. Four weeks after treatment, the vehicle group had a 100% mortality rate, while all animals from P-CD19 CAR T-treated group (N=5) remained alive without clinical symptoms including weight loss or changes in their fur. In summary, Celularity has defined a robust process for the generation and expansion of CD19 CAR T cells from human placenta. These cells exhibit potent anti-tumor activity both in vitro and in vivo with little evidence of acute GvHD induction, highlighting their potential as an allogeneic, adoptive cell therapeutic agent. Future in vivo GvHD studies will include assessment of both CD19 CAR and TRAC KO genetically modified P-T cells. Disclosures Karasiewicz: Celgene: Equity Ownership; Celularity, Inc.: Employment, Equity Ownership, Patents & Royalties: Patent Inventor. He:Celularity Inc: Employment. Ng:Celularity, Inc.: Employment. Tess:Celularity, Inc.: Employment. Ling:Celularity Inc: Employment. Kaufmann:Sorrento Therapeutics, Inc.: Employment, Equity Ownership, Patents & Royalties. Zeldis:Sorrento Therapeutics Inc: Employment, Equity Ownership. Ji:Celularity, Inc.: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Sorrento Therapeutics Inc: Employment, Equity Ownership, Patents & Royalties. Hariri:Celularity Inc: Employment. Zhang:Celularity Inc: Employment.

scholarly journals Superior Efficacy of CAR-T Cells Using Marrow-Infiltrating Lymphocytes (MILsTM) As Compared to Peripheral Blood Lymphocytes (PBLs)

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 4437-4437 ◽  
Author(s):  
Eric R. Lutz ◽  
Srikanta Jana ◽  
Lakshmi Rudraraju ◽  
Elizabeth DeOliveira ◽  
Jing Zhou ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Employment Equity ◽  
Car T Cells ◽  
Car T ◽  
Equity Ownership

Background The type of T cell used in generating chimeric antigen receptor (CAR) T cells is an important choice. Evidence suggests that T cells that are early in the effector/memory differentiation pathway with more stemness and greater potential to persist are better than more differentiated T cells with less stemness that are more readily exhausted and have less potential to persist. Marrow-infiltrating Lymphocytes (MILsTM) is a novel form of adoptive T cell therapy composed of patient-autologous, polyclonal CD4 and CD8 T cells that are activated and expanded from the bone marrow. Genetically unmodified MILsTM have demonstrated antitumor activity in patients with multiple myeloma and are being developed for several other tumor types, including non-small cell lung cancer and other solid tumors. Distinguishing features of bone marrow T cells used to produce MILsTM include their memory phenotype, inherent tumor antigen-specificity, higher CD8:CD4 ratio and ability to persist long-term when compared to peripheral blood lymphocytes (PBLs) which is the T cell source used to produce currently approved CAR-T therapies. Based on these differences, we hypothesize that MILsTM provide a more robust and better fit platform for CAR-T therapy compared to PBLs. Using a CD38-specific, 4-1BB/CD3z-signaling CAR as an initial model, we have demonstrated the feasibility of producing CAR-modified MILsTM (CAR-MILsTM) and showed that CAR-MILsTM demonstrate superior killing in vitro compared to CAR-T cells generated from patient-matched PBLs (CAR-PBLs). Herein, we build on our previous data and add a second BCMA-specific CAR model. We use the two multiple myeloma model systems to compare cytolytic potential, functionality, and expression of phenotypic markers of memory, stemness and exhaustion between patient-matched CAR-MILsTM and CAR-PBLs. Methods Matched pairs of CAR-MILsTM and CAR-PBLs were produced from the bone marrow and blood of multiple myeloma patients. Two different in vitro cytotoxicity assays, the RTCA xCelligence real-time impedance and FACS assays, were used to evaluate antigen-specific killing of target tumor cells. Functionality of CD4 and CD8 CAR-T cells, at the single-cell level, was evaluated by measuring the secretion of 32 cytokines and chemokines following in vitro antigen-specific stimulation using IsoPlexis IsoCode chips and analyzed using IsoPeak. Expression of markers of T cell memory (CD45RO & CCR7/CD62L), stemness (CD27) and exhaustion (PD1 & TIM3) on CAR-MILsTM and CAR-PBLs prior to and following antigen-specific stimulation was evaluated by flow-cytometry (FACS). Results CAR-MILsTM demonstrated superior killing of tumor target cells in vitro, regardless of the antigen specificity of the CAR, when compared to matched CAR-PBLs and this superiority persisted even upon repeated antigen encounter - a factor that may be critical in guaranteeing better anti-tumor efficacy and persistence. CAR-MILsTM demonstrated increased polyfunctionality (secretion of 2+ cytokines per cell) and an increased polyfunctional strength index (PSI) following antigen-stimulation compared to CAR-PBL in both CD4 and CD8 T cells. The enhanced PSI in CAR-MILsTM was predominately mediated by effector, stimulatory and chemoattractive proteins associated with antitumor activity including Granzyme B, IFNg, IL-8, MIP1a and MIP1b. Coincidentally, increased PSI and enhanced secretion of these same proteins was reported to be associated with improved clinical responses in patients with Non-Hodgkin lymphoma treated with CD19-specific CAR-T therapy. Expression of memory markers on CD4 and CD8 T cells were similar in CAR-MILsTM and CAR-PBLs both prior to and following antigen-stimulation. Although expression of CD27, PD1 and TIM3 were similar at baseline, CAR-MILs maintained higher levels of CD27 and lower levels of PD1 and TIM3 compared to CAR-PBLs following antigen-stimulation in both CD4 and CD8 T cells. Conclusions Collectively, our data suggest that CAR-MILsTM have several advantages over CAR-PBLs, including increased cytolytic potential, enhanced polyfunctionality, increased stemness and less exhaustion. Based on these differences and the inherent antitumor properties of MILsTM, we speculate that CAR-MILsTM would be more potent and effective than currently approved CAR-T products derived from PBLs. Disclosures Lutz: WindMIL Therapeutics: Employment, Equity Ownership. Jana:WindMIL Therapeutics: Employment, Equity Ownership. Rudraraju:WindMIL Therapeutics: Employment, Equity Ownership. DeOliveira:WindMIL Therapeutics: Employment, Equity Ownership. Zhou:Isoplexis: Employment, Equity Ownership. Mackay:Isoplexis: Employment, Equity Ownership. Borrello:Aduro: Patents & Royalties: intellectual property on allogeneic MM GVAX; BMS: Consultancy; WindMIL Therapeutics: Equity Ownership, Patents & Royalties, Research Funding; Celgene: Honoraria, Research Funding, Speakers Bureau. Noonan:WindMIL Therapeutics: Employment, Equity Ownership, Patents & Royalties; Aduro: Patents & Royalties: intellectual property on allogeneic MM GVAX.

scholarly journals Preclinical Evaluation of ALLO-819, an Allogeneic CAR T Cell Therapy Targeting FLT3 for the Treatment of Acute Myeloid Leukemia

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3921-3921 ◽  
Author(s):  
Cesar Sommer ◽  
Hsin-Yuan Cheng ◽  
Yik Andy Yeung ◽  
Duy Nguyen ◽  
Janette Sutton ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Target Cells ◽  
Employment Equity ◽  
T Cell Therapy ◽  
Cell Therapies ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T ◽  
Equity Ownership

Autologous chimeric antigen receptor (CAR) T cells have achieved unprecedented clinical responses in patients with B-cell leukemias, lymphomas and multiple myeloma, raising interest in using CAR T cell therapies in AML. These therapies are produced using a patient's own T cells, an approach that has inherent challenges, including requiring significant time for production, complex supply chain logistics, separate GMP manufacturing for each patient, and variability in performance of patient-derived cells. Given the rapid pace of disease progression combined with limitations associated with the autologous approach and treatment-induced lymphopenia, many patients with AML may not receive treatment. Allogeneic CAR T (AlloCAR T) cell therapies, which utilize cells from healthy donors, may provide greater convenience with readily available off-the-shelf CAR T cells on-demand, reliable product consistency, and accessibility at greater scale for more patients. To create an allogeneic product, the TRAC and CD52 genes are inactivated in CAR T cells using Transcription Activator-Like Effector Nuclease (TALEN®) technology. These genetic modifications are intended to minimize the risk of graft-versus-host disease and to confer resistance to ALLO-647, an anti-CD52 antibody that can be used as part of the conditioning regimen to deplete host alloreactive immune cells potentially leading to increased persistence and efficacy of the infused allogeneic cells. We have previously described the functional screening of a library of anti-FLT3 single-chain variable fragments (scFvs) and the identification of a lead FLT3 CAR with optimal activity against AML cells and featuring an off-switch activated by rituximab. Here we characterize ALLO-819, an allogeneic FLT3 CAR T cell product, for its antitumor efficacy and expansion in orthotopic models of human AML, cytotoxicity in the presence of soluble FLT3 (sFLT3), performance compared with previously described anti-FLT3 CARs and potential for off-target binding of the scFv to normal human tissues. To produce ALLO-819, T cells derived from healthy donors were activated and transduced with a lentiviral construct for expression of the lead anti-FLT3 CAR followed by efficient knockout of TRAC and CD52. ALLO-819 manufactured from multiple donors was insensitive to ALLO-647 (100 µg/mL) in in vitro assays, suggesting that it would avoid elimination by the lymphodepletion regimen. In orthotopic models of AML (MV4-11 and EOL-1), ALLO-819 exhibited dose-dependent expansion and cytotoxic activity, with peak CAR T cell levels corresponding to maximal antitumor efficacy. Intriguingly, ALLO-819 showed earlier and more robust peak expansion in mice engrafted with MV4-11 target cells, which express lower levels of the antigen relative to EOL-1 cells (n=2 donors). To further assess the potency of ALLO-819, multiple anti-FLT3 scFvs that had been described in previous reports were cloned into lentiviral constructs that were used to generate CAR T cells following the standard protocol. In these comparative studies, the ALLO-819 CAR displayed high transduction efficiency and superior performance across different donors. Furthermore, the effector function of ALLO-819 was equivalent to that observed in FLT3 CAR T cells with normal expression of TCR and CD52, indicating no effects of TALEN® treatment on CAR T cell activity. Plasma levels of sFLT3 are frequently increased in patients with AML and correlate with tumor burden, raising the possibility that sFLT3 may act as a decoy for FLT3 CAR T cells. To rule out an inhibitory effect of sFLT3 on ALLO-819, effector and target cells were cultured overnight in the presence of increasing concentrations of recombinant sFLT3. We found that ALLO-819 retained its killing properties even in the presence of supraphysiological concentrations of sFLT3 (1 µg/mL). To investigate the potential for off-target binding of the ALLO-819 CAR to human tissues, tissue cross-reactivity studies were conducted using a recombinant protein consisting of the extracellular domain of the CAR fused to human IgG Fc. Consistent with the limited expression pattern of FLT3 and indicative of the high specificity of the lead scFv, no appreciable membrane staining was detected in any of the 36 normal tissues tested (n=3 donors). Taken together, our results support clinical development of ALLO-819 as a novel and effective CAR T cell therapy for the treatment of AML. Disclosures Sommer: Allogene Therapeutics, Inc.: Employment, Equity Ownership. Cheng:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Yeung:Pfizer Inc.: Employment, Equity Ownership. Nguyen:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Sutton:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Melton:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Valton:Cellectis, Inc.: Employment, Equity Ownership. Poulsen:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Djuretic:Pfizer, Inc.: Employment, Equity Ownership. Van Blarcom:Allogene Therapeutics, Inc.: Employment, Equity Ownership. Chaparro-Riggers:Pfizer, Inc.: Employment, Equity Ownership. Sasu:Allogene Therapeutics, Inc.: Employment, Equity Ownership.

A Novel and Highly Potent CAR T Cell Drug Product for Treatment of BCMA-Expressing Hematological Malignances

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3094-3094 ◽  
Author(s):  
Alena A. Chekmasova ◽  
Holly M. Horton ◽  
Tracy E. Garrett ◽  
John W. Evans ◽  
Johanna Griecci ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Cell Lines ◽  
Drug Product ◽  
Single Chain ◽  
Employment Equity ◽  
Car T Cells ◽  
Car T ◽  
Equity Ownership

Abstract Recently, B cell maturation antigen (BCMA) expression has been proposed as a marker for identification of malignant plasma cells in patients with multiple myeloma (MM). Nearly all MM and some lymphoma tumor cells express BCMA, while normal tissue expression is restricted to plasma cells and a subset of mature B cells. Targeting BCMA maybe a therapeutic option for treatment of patients with MM and some lymphomas. We are developing a chimeric antigen receptor (CAR)-based therapy for the treatment of BCMA-expressing MM. Our anti-BCMA CAR consists of an extracellular single chain variable fragment (scFv) antigen recognition domain derived from an antibody specific to BCMA, fused to CD137 (4-1BB) co-stimulatory and CD3zeta chain signaling domains. Selection of our development candidate was based on the screening of four distinct anti-BCMA CARs (BCMA01-04) each comprised of unique single chain variable fragments. One candidate, BCMA02 (drug product name bb2121) was selected for further studies based on the robust frequency of CAR-positive cells, increased surface expression of the CAR molecule, and superior in vitro cytokine release and cytolytic activity against the MM cell lines. In addition to displaying specific activity against MM (U226-B1, RPMI-8226 and H929) and plasmacytoma (H929) cell lines, bb2121 was demonstrated to react to lymphoma cell lines, including Burkitt's (Raji, Daudi, Ramos), chronic lymphocytic leukemia (Mec-1), diffuse large B cell (Toledo), and a Mantle cell lymphoma (JeKo-1). Based on receptor density quantification, bb2121 can recognize tumor cells expressing less than 1000 BCMA molecules per cell. The in vivo pharmacology of bb2121 was studied in NSG mouse models of human MM and Burkitt's lymphoma. NSG mice were injected subcutaneously (SC) with 107 RPMI-8226 MM cells. After 18 days, mice received a single intravenous (IV) administration of vehicle or anti-CD19Δ (negative control, anti-CD19 CAR lacking signaling domain) or anti-BCMA CAR T cells, or repeated IV administration of bortezomib (Velcade®; 1 mg/kg twice weekly for 4 weeks). Bortezomib, which is a standard of care for MM, induced only transient reductions in tumor size and was associated with toxicity, as indicated by substantial weight loss during dosing. The vehicle and anti-CD19Δ CAR T cells failed to inhibit tumor growth. In contrast, treatment with bb2121 resulted in rapid and sustained elimination of the tumors, increased body weights, and 100% survival. Flow cytometry and immunohistochemical analysis of bb2121 T cells demonstrated trafficking of CAR+ T cells to the tumors (by Day 5) followed by significant expansion of anti-BCMA CAR+ T cells within the tumor and peripheral blood (Days 8-10), accompanied by tumor clearance and subsequent reductions in circulating CAR+ T cell numbers (Days 22-29). To further test the potency of bb2121, we used the CD19+ Daudi cell line, which has a low level of BCMA expression detectable by flow cytometry and receptor quantification analysis, but is negative by immunohistochemistry. NSG mice were injected IV with Daudi cells and allowed to accumulate a large systemic tumor burden before being treated with CAR+ T cells. Treatment with vehicle or anti-CD19Δ CAR T cells failed to prevent tumor growth. In contrast, anti-CD19 CAR T cells and anti-BCMA bb2121 demonstrated tumor clearance. Adoptive T cell immunotherapy approaches designed to modify a patient's own lymphocytes to target the BCMA antigen have clear indications as a possible therapy for MM and could be an alternative method for treatment of other chemotherapy-refractory B-cell malignancies. Based on these results, we will be initiating a phase I clinical trial of bb2121 for the treatment of patients with MM. Disclosures Chekmasova: bluebird bio, Inc: Employment, Equity Ownership. Horton:bluebird bio: Employment, Equity Ownership. Garrett:bluebird bio: Employment, Equity Ownership. Evans:bluebird bio, Inc: Employment, Equity Ownership. Griecci:bluebird bio, Inc: Employment, Equity Ownership. Hamel:bluebird bio: Employment, Equity Ownership. Latimer:bluebird bio: Employment, Equity Ownership. Seidel:bluebird bio, Inc: Employment, Equity Ownership. Ryu:bluebird bio, Inc: Employment, Equity Ownership. Kuczewski:bluebird bio: Employment, Equity Ownership. Horvath:bluebird bio: Employment, Equity Ownership. Friedman:bluebird bio: Employment, Equity Ownership. Morgan:bluebird bio: Employment, Equity Ownership.

Single vector multiplexed shRNA provides a non-gene edited strategy to concurrently knockdown the expression of multiple genes in CAR T cells.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. 3103-3103
Author(s):  
David Edward Gilham ◽  
Simon Bornschein ◽  
Lorraine Springuel ◽  
Alexandre Michaux ◽  
Mikhail Steklov ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Retroviral Vector ◽  
Vector System ◽  
Car T Cells ◽  
Shrna Vector ◽  
Multiple Genes ◽  
Car T

3103 Background: Engineered T cells expressing chimeric antigen receptors (CAR) are now delivering clinically relevant results in patients with advanced hematological malignancies. One critical area for future development is to modulate gene expression thereby endowing the engineered T cell with specific desired features that enhance anti-tumor activity. Methods: Short-hairpin RNA (shRNA) were cloned individually or multiplexed within micro-RNA scaffolds that enabled the co-expression of the individual shRNA with a CAR and a selectable marker all driven by a PolII promoter within a single retroviral vector. Primary human T cells transduced with the CAR-shRNA vectors were selected, expanded in vitro, subjected to negative selection to eliminate any remaining TCR+ cells and examined for target gene expression and functional activity. Results: A 500bp DNA fragment incorporating a shRNA-specific for CD3ζ cloned into a retroviral vectoreffectively knocked down expression of CD3ζ in transduced BCMA-specific CAR T cells. The consequent reduction of cell surface TCR expression resulted in minimal cytokine production upon TCR stimulation in vitro providing a potential allogeneic CAR T approach. These CAR T cells showed no demonstrable evidence of GvHD induction when infused in NSG mice yet maintained BCMA-specific CAR activity in KMS-11 and RPMI-8226 established myeloma models. Initial studies further confirmed that two shRNA could be expressed from a single retroviral vector to modulate the expression of multiple genes. Further engineering of the microRNA framework reduced the size of the transgene load to 394bp while enabling the expression of up to 4 shRNA within a single vector. shRNA specific for CD3ζ, beta-2-microglobulin, CD52 and diacylglycerol kinase alpha were engineered into the framework downstream of a CD19-CAR. Transduced Jurkat cells showed concurrent knockdown of the respective gene products at the mRNA and protein levels. Conclusions: A first-in-human clinical trial evaluating the first-generation single shRNA-vector in the context of a BCMA-targeting CAR as a non-gene edited approach to allogeneic CAR T cell therapy will be initiated in 2020. The proof of principle study here shows that multiple shRNAs are active within a single viral vector thereby avoiding the need for bespoke individual clinical reagents to target multiple genes. The multiplexed shRNA vector system is now in further development to explore whether this strategy can enhance the therapeutic potential of CAR T cells.

scholarly journals Preclinical Development of CTX120, an Allogeneic CAR-T Cell Targeting Bcma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1921-1921 ◽  
Author(s):  
Henia Dar ◽  
Daniel Henderson ◽  
Zinkal Padalia ◽  
Ashley Porras ◽  
Dakai Mu ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Cells ◽  
T Cell ◽  
Cell Targeting ◽  
Employment Equity ◽  
Term Survival ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T ◽  
Equity Ownership

Abstract Autologous CAR-T cells targeting BCMA have induced robust and durable responses in patients with relapsed/refractory multiple myeloma. However, autologous cell therapies face several challenges which will likely limit the number of patients that will have access to these therapies. These limitations include manufacturing failure rates, wait time and supply constraints in addition to other factors such as reimbursement. Allogeneic CAR-T cells can potentially overcome these access challenges, and may have several other advantages over autologous therapies. Allogeneic CAR-T cells are derived from robust healthy donor T cells through a batch manufacturing process, which may result in a highly consistent product with greater potency and enable better safety management. Here we show further development and preclinical data for CTX120, an allogeneic "off the shelf" CAR-T cell targeting BCMA. CTX120 is produced using the CRISPR/Cas9 system to eliminate TCR and MHC class I, coupled with specific insertion of the CAR at the TRAC locus. CTX120 shows consistent and high percent CAR expression from this controlled insertion and exhibits target-specific cytotoxicity and cytokine secretion in response to BCMA positive cell lines. CTX120 CAR-T cells retain their cytotoxic capacity over multiple in vitro re-challenges, demonstrating durable potency and lack of exhaustion. In mouse models of multiple myeloma, CTX120 showed typical CAR-T persistence and eliminated tumors completely, resulting in long-term survival as compared to untreated animals. These data support the ongoing development of CTX120 for treatment of patients with multiple myeloma and further demonstrate the potential for our CRISPR/Cas9 engineered allogeneic CAR-T platform to generate potent CAR-T cells targeting different tumor antigens. Disclosures Dar: CRISPR Therapeutics: Employment, Equity Ownership. Henderson:CRISPR Therapeutics: Employment, Equity Ownership. Padalia:CRISPR Therapeutics: Employment, Equity Ownership. Porras:CRISPR Therapeutics: Employment, Equity Ownership. Mu:CRISPR Therapeutics: Employment, Equity Ownership. Kyungah:CRISPR Therapeutics: Employment, Equity Ownership. Police:CRISPR Therapeutics: Employment, Equity Ownership. Kalaitzidis:CRISPR Therapeutics: Employment, Equity Ownership. Terrett:CRISPR Therapeutics: Employment, Equity Ownership. Sagert:CRISPR Therapeutics: Employment, Equity Ownership.

scholarly journals Durable Clinical Responses in Heavily Pretreated Patients with Relapsed/Refractory Multiple Myeloma: Updated Results from a Multicenter Study of bb2121 Anti-Bcma CAR T Cell Therapy

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 740-740 ◽  
Author(s):  
Jesus G. Berdeja ◽  
Yi Lin ◽  
Noopur Raje ◽  
Nikhil Munshi ◽  
David Siegel ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Cells ◽  
T Cell ◽  
Research Funding ◽  
Employment Equity ◽  
Car T Cells ◽  
Car T ◽  
Clinical Responses ◽  
Equity Ownership ◽  
Dose Levels

Abstract Introduction: Chimeric antigen receptor (CAR) T cell therapies have demonstrated robust and sustained clinical responses in several hematologic malignancies. Data suggest that achieving acceptable benefit:risk profiles depends on several factors, including the specificity of the antigen target and characteristics of the CAR itself, including on-target, off-tumor activity.To test the safety and efficacy of CAR T cells in relapsed and/or refractory multiple myeloma (RRMM), we have designed a second-generation CAR construct targeting B cell maturation antigen (BCMA) to redirect T cells to MM cells. BCMA is a member of the tumor necrosis factor superfamily that is expressed primarily by malignant myeloma cells, plasma cells, and some mature B cells. bb2121 consists of autologous T cells transduced with a lentiviral vector encoding a novel CAR incorporating an anti-BCMA scFv, a 4-1BB costimulatory motif and a CD3-zeta T cell activation domain. Methods: CRB-401 (NCT02658929) is a multi-center phase 1 dose escalation trial of bb2121 in patients with RRMM who have received ≥ 3 prior regimens, including a proteasome inhibitor and an immunomodulatory agent, or are double-refractory, and have ≥ 50% BCMA expression on malignant cells. Peripheral blood mononuclear cells are collected via leukapheresis and shipped to a central facility for transduction, expansion, and release testing prior to being returned to the site for infusion. Patients undergo lymphodepletion with fludarabine (30 mg/m2) and cyclophosphamide (300 mg/m2) daily for 3 days then receive 1 infusion of bb2121. The study follows a standard 3+3 design with planned dose levels of 50, 150, 450, 800, and 1,200 x 106 CAR+ T cells. The primary outcome measure is incidence of adverse events (AEs), including dose-limiting toxicities (DLTs). Additional outcome measures were quality and duration of clinical response assessed according to the IMWG Uniform Response Criteria for Multiple Myeloma, evaluation of minimal residual disease (MRD), overall and progression-free survival, quantification of bb2121 in blood, and quantification of circulating soluble BCMA over time. Results: Asof May 4, 2017, 21 patients (median 58 [37 to 74] years old) with a median of 5 (1 to 16) years since MM diagnosis, had been infused with bb2121, and 18 patients were evaluable for initial (1-month) clinical response. Patients had a median of 7 prior lines of therapy (range 3 to 14), all with prior autologous stem cell transplant; 67% had high-risk cytogenetics. Fifteen of 21 (71%) had prior exposure to, and 6 of 21 (29%) were refractory to 5 prior therapies (Bort/Len/Car/Pom/Dara). Median follow-up after bb2121 infusion was 15.4 weeks (range 1.4 to 54.4 weeks). As of data cut-off, no DLTs and no treatment-emergent Grade 3 or higher neurotoxicities similar to those reported in other CAR T clinical studies had been observed. Cytokine release syndrome (CRS), primarily Grade 1 or 2, was reported in 15 of 21 (71%) patients: 2 patients had Grade 3 CRS that resolved in 24 hours and 4 patients received tocilizumab, 1 with steroids, to manage CRS. CRS was more common in the higher dose groups but did not appear related to tumor burden. One death on study, due to cardiopulmonary arrest more than 4 months after bb2121 infusion in a patient with an extensive cardiac history, was observed while the patient was in sCR and was assessed as unrelated to bb2121. The overall response rate (ORR) was 89% and increased to 100% for patients treated with doses of 150 x 106 CAR+ T cells or higher. No patients treated with doses of 150 x 106 CAR+ T cells or higher had disease progression, with time since bb2121 between 8 and 54 weeks (Table 1). MRD negative results were obtained in all 4 patients evaluable for analysis. CAR+ T cell expansion has been demonstrated consistently and 3 of 5 patients evaluable for CAR+ cells at 6 months had detectable vector copies. A further 5 months of follow up on reported results and initial data from additional patients will be presented. Conclusions: bb2121 shows promising efficacy at dose levels above 50 x 106 CAR+ T cells, with manageable CRS and no DLTs to date. ORR was 100% at these dose levels with 8 ongoing clinical responses at 6 months and 1 patient demonstrating a sustained response beyond one year. These initial data support the potential of CAR T therapy with bb2121 as a new treatment paradigm in RRMM. CT.gov study NCT02658929, sponsored by bluebird bio and Celgene Disclosures Berdeja: Teva: Research Funding; Janssen: Research Funding; Novartis: Research Funding; Abbvie: Research Funding; Celgene: Research Funding; BMS: Research Funding; Takeda: Research Funding; Vivolux: Research Funding; Amgen: Research Funding; Constellation: Research Funding; Bluebird: Research Funding; Curis: Research Funding. Siegel: Celgene, Takeda, Amgen Inc, Novartis and BMS: Consultancy, Speakers Bureau; Merck: Consultancy. Jagannath: MMRF: Speakers Bureau; Bristol-Meyers Squibb: Consultancy; Merck: Consultancy; Celgene: Consultancy; Novartis: Consultancy; Medicom: Speakers Bureau. Turka: bluebird bio: Employment, Equity Ownership. Lam: bluebird bio: Employment, Equity Ownership. Hege: Celgene Corporation: Employment, Equity Ownership. Morgan: bluebird bio: Employment, Equity Ownership, Patents & Royalties. Quigley: bluebird bio: Employment, Equity Ownership, Patents & Royalties. Kochenderfer: Bluebird bio: Research Funding; N/A: Patents & Royalties: I have multiple patents in the CAR field.; Kite Pharma: Research Funding.

scholarly journals Development and Evaluation of a Human Single Chain Variable Fragment (scFv) Derived Bcma Targeted CAR T Cell Vector Leads to a High Objective Response Rate in Patients with Advanced MM

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 742-742 ◽  
Author(s):  
Eric L Smith ◽  
Sham Mailankody ◽  
Arnab Ghosh ◽  
Reed Masakayan ◽  
Mette Staehr ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Board Of Directors ◽  
Research Funding ◽  
Employment Equity ◽  
Advisory Committees ◽  
Car T Cells ◽  
Car T Cell ◽  
Car T ◽  
Equity Ownership

Abstract Patients with relapsed/refractory MM (RRMM) rarely obtain durable remissions with available therapies. Clinical use of BCMA targeted CAR T cell therapy was first reported in 12/2015 for RRMM, and based on small numbers, preliminary results appear promising. Given that host immune anti-murine CAR responses have limited the efficacy of repeat dosing (Turtle C. Sci Trans Med 2016), our goal was to develop a human BCMA targeted CAR T cell vector for clinical translation. We screened a human B cell derived scFv phage display library containing 6x1010 scFvs with BCMA expressing NIH 3T3 cells, and validated results on human MM cell lines. 57 unique and diverse BCMA specific scFvs were identified containing light and heavy chain CDR's each covering 6 subfamilies, with HCDR3 length ranges from 5-18 amino acids. 17 scFvs met stringent specificity criteria, and a diverse set was cloned into CAR vectors with either a CD28 or a 4-1BB co-stimulatory domain. Donor T cells transduced with BCMA targeted CAR vectors that conveyed particularly desirable properties over multiple in vitro assays, including: cytotoxicity on human MM cell lines at low E:T ratios (>90% lysis, 1:1, 16h), robust proliferation after repeat antigen stimulation (up to 700 fold, stimulation q3-4d for 14d), and active cytokine profiling, were selected for in vivo studies using a marrow predominant human MM cell line model in NSG mice. A single IV injection of CAR T cells, either early (4d) or late (21d) after MM engraftment was evaluated. In both cases survival was increased when treated with BCMA targeted CAR T cells vs CD19 targeted CAR T cells (median OS at 60d NR vs 35d p<0.05). Tumor and CAR T cells were imaged in vivo by taking advantage of luciferase constructs with different substrates. Results show rapid tumor clearance, peak (>10,000 fold) CAR T expansion at day 6, followed by contraction of CAR T cells after MM clearance, confirming the efficacy of the anti-BCMA scFv/4-1BB containing construct. Co-culture with primary cells from a range of normal tissues did not activate CAR T cells as noted by a lack of IFN release. Co-culture of 293 cells expressing this scFv with those expressing a library of other TNFRSF or Ig receptor members demonstrated specific binding to BCMA. GLP toxicity studies in mice showed no unexpected adverse events. We generated a retroviral construct for clinical use including a truncated epithelial growth factor receptor (EGFRt) elimination gene: EGFRt/hBCMA-41BBz. Clinical investigation of this construct is underway in a dose escalation, single institution trial. Enrollment is completed on 2/4 planned dose levels (DL). On DL1 pts received cyclophosphamide conditioning (3g/m2 x1) and 72x106 mean CAR+ T cells. On DL2 pts received lower dose cyclophosphamide/fludarabine (300/30 mg/m2 x3) and 137x106 mean CAR+ T cells. All pts screened for BCMA expression by IHC were eligible. High risk cytogenetics were present in 4/6 pts. Median prior lines of therapy was 7; all pts had IMiD, PI, high dose melphalan, and CD38 directed therapies. With a data cut off of 7/20/17, 6 pts are evaluable for safety. There were no DLT's. At DL1, grade 1 CRS, not requiring intervention, occurred in 1/3 pts. At DL2, grade 1/2 CRS occurred in 2/3 pts; both received IL6R directed Tocilizumab (Toci) with near immediate resolution. In these 2 pts time to onset of fever was a mean 2d, Tmax was 39.4-41.1 C, peak CRP was 25-27mg/dl, peak IL6 level pre and post Toci were 558-632 and 3375-9071 pg/ml, respectively. Additional serum cytokines increased >10 fold from baseline in both pts include: IFNg, GM CSF, Fractalkine, IL5, IL8, and IP10. Increases in ferritin were limited, and there were no cases of hypofibrinogenemia. There were no grade 3-5 CRS and no neurotoxicities or cerebral edema. No pts received steroids or Cetuximab. Median time to count recovery after neutropenia was 10d (range 6-15d). Objective responses by IMWG criteria after a single dose of CAR T cells were observed across both DLs. At DL1, of 3 pts, responses were 1 VGPR, 1 SD, and 1 pt treated with baseline Mspike 0.46, thus not evaluable by IMWG criteria, had >50% reduction in Mspike, and normalization of K/L ratio. At DL2, 2/2 pts had objective responses with 1 PR and 1 VGPR (baseline 95% marrow involvement); 1 pt is too early to evaluate. As we are employing a human CAR, the study was designed to allow for an optional second dose in pts that do not reach CR. We have treated 2 pts with a second dose, and longer follow up data is pending. Figure 1 Figure 1. Disclosures Smith: Juno Therapeutics: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties: BCMA targeted CAR T cells, Research Funding. Almo: Cue Biopharma: Other: Founder, head of SABequity holder; Institute for Protein Innovation: Consultancy; AKIN GUMP STRAUSS HAUER & FELD LLP: Consultancy. Wang: Eureka Therapeutics Inc.: Employment, Equity Ownership. Xu: Eureka Therapeutics, Inc: Employment, Equity Ownership. Park: Amgen: Consultancy. Curran: Juno Therapeutics: Research Funding; Novartis: Consultancy. Dogan: Celgene: Consultancy; Peer Review Institute: Consultancy; Roche Pharmaceuticals: Consultancy; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Seattle Genetics: Consultancy, Membership on an entity's Board of Directors or advisory committees. Liu: Eureka Therpeutics Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Brentjens: Juno Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding.

Sign in / Sign up

Export Citation Format

Share Document