Manufacture of an Allogeneic CAR-T Stem Cell Memory Product Candidate for Multiple Myeloma, P-Bcma-ALLO1, Is Robust, Reproducible and Highly Scalable

Chimeric Antigen Receptor (CAR) T cell therapy has generated unprecedented efficacy in the treatment of multiple hematologic malignancies. For relapsed/refractory Multiple Myeloma (MM), autologous CAR-T products directed against the B cell maturation antigen (BCMA), such as Poseida's P-BCMA-101, have demonstrated significant efficacy. P-BCMA-101 is comprised of a high-percentage of stem cell memory T cells (TSCM), resulting in a product that is much safer and potentially more durable than other anti-BCMA autologous product candidates. However, as individualized products, all autologous CAR-T products are expensive to manufacture and dependent upon patient T-cells of variable quality. We are developing P-BCMA-ALLO1, an off-the-shelf allogeneic (allo) BCMA-specific CAR-T product candidate derived from healthy donor material, which provides numerous advantages over autologous products, increasing patient access by being immediately available and greatly reducing manufacturing cost and variability. P-BCMA-ALLO1 is produced using two key platform technologies: the nonviral piggyBac® (PB) DNA Modification System and the high-fidelity Cas-CLOVER™ (CC) Site-Specific Gene Editing System. The mRNA coding for hyperactive, or "Super PB" transposase (SPB), and CC enzymes are codelivered with the P-BCMA-ALLO1 PB-based DNA transgene via electroporation to healthy donor T cells to stably integrate the transgene, as well as to knockout (KO) several mediators of allo graft-versus-host and host-versus-graft responses to maximize patient safety and durability of response. The P-BCMA-ALLO1 transgene encodes three genes, a BCMA-specific single-domain variable heavy chain (VH)-CAR (VCAR) gene, a drug selection gene to generate a ~100% CAR+ product, as well as a caspase-based safety switch gene to reduce or eliminate the product in vivo, if desired. The CC System is used to KO the endogenous T Cell Receptor (TCR) and beta-2 microglobulin, thereby decreasing Major Histocompatibility Complex (MHC) class I expression. KO of these key targets is aimed to prevent graft-versus-host disease, as well as reduce host-versus-graft rejection of the product. The CC System can efficiently edit resting T cells, thereby maintaining a high-percentage of TSCM cells, and does not create unwanted off-target mutations, another important consideration when creating an allo product candidate. To maximize the number of doses produced from a single manufacturing run, we have developed a proprietary "booster molecule" that allows for significant expansion of TCR-KO CAR-TSCM cells to potentially produce hundreds of doses. To date, large-scale manufacturing of significant doses of potent allo CAR-T products has been challenging for the field. P-BCMA-ALLO1 manufacturing uses a potentially unlimited number of individual serial donors. We have currently produced P-BCMA-ALLO1 at both research and near-commercial scale from >35 donors with >97% manufacturing success. While a range of TCR-KO efficiencies was observed (~50-90%), the final product was always >99% homozygous TCR-KO after a purification step. Overall expansion of TCR-KO cells ranged from ~2-20 fold, and after removal of unedited TCR+ cells ~0.42-7.04x10e9 TCR-KO cells were recovered from 0.75x10e9 starting cells. However, working at clinical production scale (starting with ~3x10e9 cells), up to 250 doses of P-BCMA-ALLO1 could be manufactured per run, at a dose of 150x10e6 cells/patient. Importantly, with this level of donor and manufacturing robustness, no significant prior screening of donor material, other than to meet standard FDA requirements, would be needed. P-BCMA-ALLO1 made from multiple donors were comprised of an exceptionally high-percentage of the desirable TSCM cells (CD45RA+CD62L+CD45RO-) and had minimal to no expression of exhaustion markers, such as PD-1 or Lag3. Furthermore, P-BCMA-ALLO1 demonstrated potent efficacy in the RPMI-8226 xenograft model in NSG mice across multiple products generated from separate individual healthy donors. Altogether, these data demonstrate a robust, reproducible and highly scalable manufacturing process. Moreover, this manufacturing process can easily be expanded for use with additional CAR targets for treatment of other hematologic or solid tumor malignancies. Disclosures Cranert: Poseida Therapeutics: Employment, Equity Ownership. Richter:Poseida Therapeutics: Employment, Equity Ownership. Tong:Poseida Therapeutics: Employment, Equity Ownership. Weiss:Poseida Therapeutics, Inc.: Employment, Equity Ownership. Tan:Poseida Therapeutics: Employment, Equity Ownership. Ostertag:Poseida Therapeutics: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Coronella:Poseida Therapeutics, Inc: Employment, Equity Ownership. Shedlock:Poseida Therapeutics, Inc.: Employment, Equity Ownership.

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

Blood ◽

10.1182/blood-2018-99-116443 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 1921-1921 ◽

Cited By ~ 2

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.

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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 ◽

10.1182/blood.v130.suppl_1.740.740 ◽

2017 ◽

Vol 130 (Suppl_1) ◽

pp. 740-740 ◽

Cited By ~ 4

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.

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Low Dose of Human scFv-Derived BCMA-Targeted CAR-T Cells Achieved Fast Response and High Complete Remission in Patients with Relapsed/Refractory Multiple Myeloma

Blood ◽

10.1182/blood-2018-99-113220 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 960-960 ◽

Cited By ~ 23

Author(s):

Songfu Jiang ◽

Jie Jin ◽

Siguo Hao ◽

Min Yang ◽

Linjun Chen ◽

...

Keyword(s):

Multiple Myeloma ◽

T Cells ◽

Single Dose ◽

Employment Equity ◽

Car T Cells ◽

Car T ◽

Dose Infusion ◽

Human Scfv ◽

Equity Ownership ◽

Grade 3

Abstract Introduction: B Cell Mature Antigen (BCMA)-targeted chimeric antigen receptor T (CAR-T) cell therapy emerges as promising treatment for patients with relapse/refractory multiple myeloma (RRMM). Previous studies indicate patients who receive high-dose CAR-T cells may achieve better remission but have worse adverse events, like cytokine release syndrome (CRS). To solve this dilemma, we have developed novel autologous CAR-T therapeutics CT053 that are genetically modified T cells comprising an extracellular anti-BCMA human scFv and an intracellular 4-1BB costimulatory motif connected to a CD3-zeta T cell activation domain. Methods: A multi-center investigator-initiated clinical study is designed to evaluate CT053 in patients with RRMM who have failed in the prior treatment with ≥2 regimens, including a proteasome inhibitor, an immunomodulatory agent, and anti-CD38 monoclonal antibody. All patients have ≥50% BCMA expression on malignant cells. Patients are subjected to the lymphodepletion with 20-25 mg/m2 fludarabine and 300-500 mg/m2 cyclophosphamide daily for 2-4 days prior to receiving single-dose infusion of CT053 CAR-T cells. In case of progressive disease, patient may be dosed again on basis of investigators' evaluation of the disease status, BCMA expression and CAR-T persistence. Most enrolled patients received a single dose of 1.5 x 108 cells, except for 1 patient who received 0.5 x 108 cells and 1 patient who was infused with 1.8 x 108 cells. The primary outcome measure is incidence of adverse events (AEs), including dose-limiting toxicities (DLTs) and CAR T related AEs. Additional outcome measures include clinical response assessed according to the IMWG Uniform Response Criteria for Multiple Myeloma, overall and progression-free survival, pharmacokinetics and pharmacodynamic of CT053. Results: The study was performed in compliance with the declaration of Helsinki. As of the data cut-off date (July 10th, 2018), 16 patients (median 55 [39 to 67] years old) with a median of 3.9 (0.4 to 16.7) years since MM diagnosis, were infused with CT053. Patients had a median of 4 prior different regimens (range 2 to 10), and 56% (9/16) patients received prior autologous or allogeneic stem cell transplant. Among 16 patients, no neurotoxicity and no dose-limiting toxicities (DLT) were observed. The most common grade≥3 CAR-T related AEs were 3 thrombocytopenia (19%), 3 leukopenia (19%), 2 anemia (13%), 2 neutropenia (13%), 2 fever (13%) (Figure 1A). CRS was reported in 3 patients, including 1 Grade 3, 1 Grade 2 and 1 Grade 1, who had rapid recovery after Tocilizumab administration. 13/16 patients were eligible for initial evaluation of early clinical response with a median observation period of 8 (4 to 36) weeks. Overall response rate (ORR) in 13/13 patients was 100% post treatment. 12/13 patients (92%) quickly achieved partial response (4 PR), very good PR (6 VGPR), and complete response (2 CR) within 4 weeks post single-dose infusion (Figure 1B). 5/12 patients (42%) who were dosed at ≥1.5 x 108 CT053 CAR-T cells obtained CR at a median of 8 weeks post treatment. Durable responses from 4 weeks towards the data cut-off date were found in 12/13 patients (92%). One relapse from VGPR by the Week 12 was reported in a patient who had aggressive RRMM at enrollment and received the reduced dose of lymphodepletion regimen at 19 mg/m2 fludarabine and 192 mg/m2 cyclophosphamide for 2 days prior to CT053 infusion. Because positive BCMA expression on malignant cells was verified at relapse, the patient was re-dosed with CT053 at the Week 16 and subjected to the further evaluation. All patients had detectable CAR-T expansion from Day 3 post CT053 infusion. Expansion peaks were found on Day 7 (5/13), Day 14 (6/13) and Day 21 (2/13). 11/13 patients had notable persistence of CT053 CAR-T cells up to 4-6 months. The only relapsed patient had the lowest CAR-T expansion peak among 13 patients, indicating the potential correlation between CAR-T expansion and response outcome. Conclusions: Data from this early-stage clinical study showed the unparalleled safety and efficacy of CT053 CAR-T cells. Major AEs were transient, manageable, and reversible. 100% ORR in 13/13 evaluable patients were reported post single-dose infusion of 0.5~1.8 x 108 cells. 5/12 patients who were dosed at ≥1.5 x 108 CAR-T cells rapidly achieved durable CR at median of 8 weeks, suggesting CT053 could be developed as competitive therapeutics to treat patients with RRMM. Disclosures Ruan: CARsgen Therapeutics: Employment. Xiao:CARsgen Therapeutics: Employment, Equity Ownership. Wang:CARsgen Therapeutics: Employment, Equity Ownership. Li:CARsgen Therapeutics: Employment, Equity Ownership.

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JCARH125, Anti-BCMA CAR T-cell Therapy for Relapsed/Refractory Multiple Myeloma: Initial Proof of Concept Results from a Phase 1/2 Multicenter Study (EVOLVE)

Blood ◽

10.1182/blood-2018-99-113548 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 957-957 ◽

Cited By ~ 48

Author(s):

Sham Mailankody ◽

Myo Htut ◽

Kelvin P. Lee ◽

William Bensinger ◽

Todd Devries ◽

...

Keyword(s):

Multiple Myeloma ◽

T Cells ◽

Research Funding ◽

Phase 1 ◽

Employment Equity ◽

Car T Cells ◽

Car T ◽

Equity Ownership ◽

Grade 3

Abstract Introduction: B-cell maturation antigen (BCMA) is expressed on malignant plasma cells and is an attractive therapeutic target for multiple myeloma. BCMA CAR T-cells, antibody drug conjugates and bispecific T-cell engagers have demonstrated substantial preclinical and clinical activity to date. JCARH125 is a BCMA-targeting CAR T product containing a lentiviral CAR construct with a fully human scFv, optimized spacer, 4-1BB co-stimulatory and CD3z activation domains. The construct has shown minimal tonic signaling and lack of inhibition by soluble BCMA. JCARH125 is generated using a manufacturing process developed to optimize various aspects, including increased consistency of cell health, in the drug product. Methods: EVOLVE (NCT03430011) is a multi-center, phase 1/2 trial of JCARH125 in patients with relapsed and/or refractory multiple myeloma, who have received 3 or more prior regimens, which must include autologous stem cell transplant, a proteasome inhibitor, immunomodulatory drug and an anti-CD38 monoclonal antibody, unless not a candidate (i.e. contraindicated) to receive one or more of the above treatments. Lymphodepleting chemotherapy (LDC) consisting of 3 days of fludarabine (30 mg/m2) and cyclophosphamide (300 mg/m2) is given 2 to 7 days prior to JCARH125 infusion. A single dose of JCARH125 is given on day 1. Dose escalation is determined using the modified toxicity probability interval 2 (mTPI-2). A minimum of 3 patients are evaluated at each dose level (DL). The first 2 DLs evaluated were 50 and 150x 106 CAR+ T cells. Additional DLs are planned, followed by an expansion at the recommended phase 2 dose (RP2D). The primary objectives of the phase 1 portion are safety and identifying a RP2D. Results: At the time of the July 12, 2018 data analysis, 19 patients have been enrolled (i.e. apheresed) and 13 patients dosed with JCARH125. Only one patient was unable to receive JCARH125, due to sepsis after LDC, leading to death before JCARH125 administration. Eight patients were evaluable for safety (≥ 1 mo follow-up). (n = 5 DL1; n = 3 DL2). Three patients (all from DL1) were evaluable for confirmed response (≥ 2 mo follow-up) per International Myeloma Working Group (IMWG) criteria. Data reported here are from these initial 8 patients. Median follow-up is 5 weeks (range 4 - 13 weeks). Median age is 53 years (range 36 - 66) with a median time from diagnosis of 4 years (range 2 - 12). Patients had received a median of 10 prior regimens (range 4 - 15). Of these 8 patients, 4 (50%) were refractory (no response or progression within 60 days of last therapy) to bortezomib, carfilzomib, lenalidomide, pomalidomide and an anti-CD38 monoclonal antibody. Seven of 8 (88%) had prior autologous stem cell transplant and 4 of 8 (50%) have IMWG high risk cytogenetics. As of the data cut, no DLTs have been observed at the first 2 DLs. Cytokine release syndrome (CRS), all grade 1 or 2, was observed in 6 of 8 (75%) patients. Median onset of CRS was 9 days (range 4 - 10) with a median duration of 4.5 days (range 2 - 19 days). None of the patients with grade 2 CRS required vasopressor support and only 1 patient received tocilizumab. No patients had grade ≥ 3 CRS. Three of 8 (38%) patients experienced neurologic adverse events (AE). Two patients had grade 1 events, and 1 had a grade 3 event (lethargy), which resolved within 24 hours after receiving steroids. Onset of neurologic AEs was 9,11 and 12 days with a duration of 2, 3 and 1 days respectively. Notably, the patient who experienced grade 3 neurotoxicity (NT), developed secondary plasma cell leukemia (PCL) just prior to receiving LDC. All 8 patients have evidence of objective response (≥ MR), including the patient with secondary PCL. 3 patients, all treated at DL1 (50 x 106 CAR+ T-cells), have confirmed responses (1 PR, 2 sCR) with the remainder unconfirmed (1 CR, 2 VGPR, 1 PR, 1 MR). As of the data cut, no patients have progressed. Additional clinical and translational data on at least 30 patients and additional follow up of at least 4 months will be available at time of presentation. Conclusion: At initial lower dose levels, JCARH125 showed an acceptable safety profile with no DLTs reported thus far. Incidence of grade ≥ 3 NT was low and no grade ≥ 3 CRS has occurred with clear clinical activity. Although durability of response and response rate in a greater number of patients remain to be determined, early experience with JCARH125 support a favorable risk-benefit profile and rapid clinical development. Disclosures Mailankody: Takeda: Research Funding; Janssen: Research Funding; Physician Education Resource: Honoraria; Juno: Research Funding. Bensinger:celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; amgen: Speakers Bureau; Takeda: Speakers Bureau; Janssen: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Devries:Junot Therapeutics: Employment. Piasecki:Juno Therapeutics: Employment, Equity Ownership; Cascadian Therapeutics: Patents & Royalties; Amgen: Patents & Royalties. Ziyad:Juno Therapeutics: Employment, Equity Ownership. Blake:Celgene: Employment, Equity Ownership. Byon:Juno Therapeutics: Employment, Equity Ownership. Jakubowiak:Janssen: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria; Karyopharm: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Adaptive Biotechnologies: Consultancy, Honoraria; SkylineDx: Consultancy, Honoraria.

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

Blood ◽

10.1182/blood-2019-130629 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 4437-4437 ◽

Cited By ~ 2

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.

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Developing a Novel Anti-Bcma CAR-T for Relapsed or Refractory Multiple Myeloma

Blood ◽

10.1182/blood-2019-125372 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 50-50 ◽

Cited By ~ 1

Author(s):

Xin Yao ◽

Shigui Zhu ◽

Jiaqi Huang ◽

Xiaoyan Qu ◽

Judy Zhu ◽

...

Keyword(s):

Clinical Trial ◽

Multiple Myeloma ◽

T Cells ◽

Tumor Cells ◽

Human Monoclonal Antibody ◽

Employment Equity ◽

Early Clinical Trial ◽

Car T Cells ◽

Car T ◽

Equity Ownership

CBMG has developed C-CAR088, a novel chimeric antigen receptor (CAR)-T cell therapy targeting BCMA, which is specifically and highly expressed on multiple myeloma cells. C-CAR088 is designed to improve efficacy through increasing the specificity and reducing immunogenicity by fusing a scFv from high-affinity human monoclonal antibody to a CD3ζ/4-1BB signaling domain. In preclinical study, the human T cells transduced with the lentiviral vector encoding C-CAR088 exhibited specific functions in vitro including CAR-T proliferation, cytokine production, cytotoxicity to BCMA positive tumor cells. C-CAR088 cells were not activated by soluble BCMA protein and MM patient serums. However, they can eradicate BCMA positive tumor cells in vivo including BMCA positive multiple myeloma tumor model RPMI-8226. C-CAR088 is manufactured in a serum free, automated and digital, closed system which produce CAR-T cells with stable and high percentage of Tcm phenotype. C-CAR088 showed a very good dose dependent tumor inhibition effect and survival benefit in animal studies. A Phase 1, 3+3 dose escalation trial is being conducted in patients with r/r MM (≥ 3 prior lines, having received treatment and proteasome inhibitors (PI) and IMiD or double refractory) to assess the safety and efficacy of C-CAR088 (NCT03815383). Patients are apheresed to harvest T cells. C-CAR088 is then manufactured and administered to patients as a single intravenous dose after a standard 3-day cyclophosphamide/fludarabine conditioning regimen. As of July 5, 2019 cutoff date, 3 patients have been treated with C-CAR088 at the dose of 1.0 x 106 CAR-T cells/kg. Patients were heavily pre-treated (7 prior lines of therapy), and all failed IMiDs and proteasome inhibitor therapies. After C-CAR088 treatment, all three patients showed clinical improvement as early as two weeks post treatment. Furthermore, C-CAR088 proliferation & expansion in the peripheral blood correlated with the decrease of tumor burden. Two patients reached VGPR at 4 weeks and 8 weeks respectively, and the third patient reached PR as early as 2 weeks post C-CAR088 infusion. C-CAR088 treatment was well tolerated, no dose-limiting toxicities (DLTs), reversible Grade 1~2 CRS observed. In conclusion, early clinical trial results in patients with r/r MM for C-CAR088 support preclinical findings that the drug shows promising efficacy and manageable safety profile.The very early clinical efficacy signal at low, suboptimal dose is encouraging and compares favorably to many other anti-BCMA CAR-T products at similar dose. The promising trend needs to be confirmed by the ongoing clinical trial. Disclosures Yao: Cellular Biomedicine Group Inc: Employment, Equity Ownership. Zhu:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Huang:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Zhu:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Wei:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Lan:Cellular Biomedicine Group Inc: Employment, Equity Ownership. LV:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Wu:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Wang:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Yang:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Zheng:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Zhao:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Zhang:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Chen:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Li:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Ren:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Zhang:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Humphries:Cellular Biomedicine Group Inc: Employment, Equity Ownership. Yao:Cellular Biomedicine Group Inc: Employment, Equity Ownership.

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Characterization of Lentiviral Vector Derived Anti-Bcma CAR T Cells Reveals Key Parameters for Robust Manufacturing of Cell-Based Gene Therapies for Multiple Myeloma

Blood ◽

10.1182/blood.v126.23.3243.3243 ◽

2015 ◽

Vol 126 (23) ◽

pp. 3243-3243

Author(s):

Graham Lilley ◽

Alden Ladd ◽

Daniel Cossette ◽

Laura Viggiano ◽

Gregory Hopkins ◽

...

Keyword(s):

Multiple Myeloma ◽

T Cells ◽

T Cell ◽

Lentiviral Vector ◽

Employment Equity ◽

Cell Product ◽

Car T Cells ◽

Car T Cell ◽

Car T ◽

Equity Ownership

Abstract T cells engineered with chimeric antigen receptors (CAR) specific to CD19 have caused rapid and durable clinical responses in ~90% of patients with acute lymphoblastic leukemia. These data support the development of additional CAR T cell products for the treatment of other hematological malignancies. 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 non-Hodgkin's lymphoma tumor cells express BCMA, while normal tissue expression is restricted to plasma cells and a subset of mature B cells. Therefore, BCMA is an attractive CAR T cell target to treat patients with MM and some B cell lymphomas. To this end, using lentiviral vector technology, we successfully generated CAR T cells specific to BCMA that exhibit potent anti-tumor activity to both multiple myeloma and Burkitt's lymphoma in animal models. Manufacture of CAR T cells for individual patient treatment requires the establishment of a robust and reproducible process - since variability in manufacturing could impact the potency of each cell product. To begin to understand the parameters of the manufacturing process that might contribute to the activity of the final product, we first tested the impact of lentiviral vector (LVV) multiplicity of infection (MOI) on CAR T cell phenotype and function. Using a broad range of MOIs (0.625 to 40) across multiple independent PBMC donors we observed no differences in population doubling or cell size throughout the ~10 day manufacturing process, irrespective of the MOI used. As expected, the number of anti-BCMA CAR expressing cells, the level of CAR expression per cell and the average vector copy number (VCN) in the cell product increased proportionally with MOI. Similarly, T cell function, as determined by an IFNg cytokine release assay in response to BCMA-expressing K562 target cells, was also correlated with the LVV MOI. Notably, increased IFNg expression was readily observable at MOIs as low as 1.25 and reached a plateau with T cells generated using an MOI of 20 or more - highlighting the sensitivity of this functional assay. Analogous data demonstrating MOI dependent in vitro killing activity were obtained using a BCMA-expressing tumor cell cytotoxicity assay. Varying the LVV MOI used during transduction simultaneously alters both the amount of anti-BCMA CAR molecules expressed per cell as well as the number of T cells in the cell product that express anti-BCMA CAR. To evaluate each variable in isolation we generated T cell products containing the same frequency of anti-BCMA CAR T cells (26 ± 4% CAR+ T cells) but different levels of anti-BCMA expression per cell by diluting T cell products made with MOIs from 5 to 40 with donor-matched untransduced cells. While these populations had markedly different levels of CAR surface expression per cell (based on anti-BCMA CAR MFI levels measured by flow cytometry) both low and high expressing anti-BCMA CAR T cell products exhibited identical levels of cytotoxicity against BCMA-expressing tumor cells. These data suggest it is the number of CAR expressing cells that is the critical driver of higher functional activity (perhaps due to the efficiency of LVV mediated anti-BCMA CAR expression per transduced cell). Finally, using this information the variability in manufacturing of anti-BCMA CAR T cells was evaluated across 11 independent normal PBMC donors. All 11 products demonstrated very similar properties with respect to cell growth (population doublings, cell volume), and VCN. Importantly, using our standard MOI we obtained a consistent and high level of anti-BCMA CAR expressing T cells that resulted in robust IFNg cytokine release when co-cultured with BCMA-expression cells. Together, our data highlight the frequency of anti-BCMA CAR T cells per cell product as a key parameter for anti-tumor activity in vitro. Moreover, these data suggest that our LVV driven T cell engineering process can reproducibly generate robust anti-BCMA CAR expressing T cell products in a donor independent manner. A phase I clinical trial to evaluate this technology as a cell-based gene therapy for MM is under development. Disclosures Lilley: bluebird bio, Inc: Employment, Equity Ownership. Ladd:bluebird bio, Inc: Employment, Equity Ownership. Cossette:bluebird bio, Inc: Employment, Equity Ownership. Viggiano:bluebird bio, Inc: Employment, Equity Ownership. Hopkins:bluebird bio, Inc: Employment, Equity Ownership. Evans:bluebird bio, Inc: Employment, Equity Ownership. Li:bluebird bio, Inc: Employment, Equity Ownership. Latimer:bluebird bio: Employment, Equity Ownership. Miller:bluebird bio: Employment, Equity Ownership. Kuczewski:bluebird bio: Employment, Equity Ownership. Bakeman:bluebird bio, Inc: Employment, Equity Ownership. MacLeod:bluebird bio, Inc: Employment, Equity Ownership. Friedman:bluebird bio: Employment, Equity Ownership. Maier:bluebird bio, Inc: Employment, Equity Ownership. Paglia:bluebird bio, Inc: Employment, Equity Ownership. Morgan:bluebird bio: Employment, Equity Ownership. Angelino:bluebird bio, Inc: Employment, Equity Ownership.

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Lisocabtagene Maraleucel (liso-cel) Manufacturing Process Control and Robustness across CD19+ Hematological Malignancies

Blood ◽

10.1182/blood-2019-127150 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 593-593 ◽

Cited By ~ 2

Author(s):

Jeffrey Teoh ◽

Timothy G. Johnstone ◽

Brian Christin ◽

Rachel Yost ◽

Neil A. Haig ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cell ◽

Manufacturing Process ◽

Starting Material ◽

Employment Equity ◽

Cell Product ◽

Car T Cell ◽

Car T ◽

Equity Ownership

Background Lisocabtagene maraleucel (liso-cel) is an investigational, CD19-directed, genetically modified, autologous cellular immunotherapy administered as a defined composition of CD8+ and CD4+ components to deliver target doses of viable chimeric antigen receptor (CAR) T cells from both components. The CAR comprises a CD19-specific scFv and 4-1BB-CD3ζ endodomain. Liso-cel is being developed for the treatment of multiple B cell malignancies, including relapsed/refractory large B cell non-Hodgkin lymphoma (NHL) and chronic lymphocytic leukemia (CLL)/small lymphocytic lymphoma (SLL). The liso-cel manufacturing process design includes controls that enable robustness across heterogeneous patient populations and disease indications, minimizing between-lot variability. This is highlighted by consistency in process duration, reduction of terminally differentiated T cells present in the T cell starting material, and consistency in T cell purity across B cell NHL and CLL/SLL indications. Methods The liso-cel manufacturing process involves selection of CD8+ and CD4+ T cells from leukapheresis, followed by independent CD8+ and CD4+ activation, transduction, expansion, formulation, and cryopreservation. Liso-cel was manufactured in support of the TRANSCEND NHL 001 (NCT02631044) and TRANSCEND CLL 004 (NCT03331198) clinical trials. Phenotypic analysis of T cell and B cell composition from leukapheresis, T cell starting material, and CAR T cell product was performed by flow cytometry. Molecular characterization of T cell receptor (TCR) clonality was estimated from the T cell starting material and CAR T cell product through transcriptional profiling. Results Liso-cel manufacturing process optimizations have been implemented in advance of commercialization. These optimizations have significantly improved process duration consistency (Figure 1; F test P=4.1×10−36). Both phenotypic and molecular TCR clonality analyses demonstrated a significant reduction in terminally differentiated CD8+ T cells across the manufacturing process. Frequencies of CD45RA+ CCR7− populations were measured by flow cytometry in CD8+ T cell starting material (median=35.1%) and CAR T cell product (median=11.7%; Wilcoxon rank sum P=3.1×10−25). Characterization of TCR clonality showed a significant decrease in clonality in the CAR T cell product compared with T cell starting material (Wilcoxon rank sum P=5.6×10−6), suggesting selective expansion of clonally diverse, less differentiated T cell populations. These findings are supported by the predominant memory T cell composition observed in liso-cel. Manufacturing process robustness enabled by in-process T cell selection is further demonstrated by the capability to produce highly pure T cell products across heterogeneous patient populations and different disease indications. T cell and B cell composition were characterized in the leukapheresis, selected T cell material, and CAR T cell product, demonstrating consistent clearance of non-T cells, including CD19+ B cells in both B- cell NHL and CLL/SLL patient cohorts. Although the CD19+ B cell composition is significantly higher in leukapheresis from patients with CLL/SLL (median=10.0% of leukocytes) compared with B cell NHL patients (median=0.0% of leukocytes, Wilcoxon rank sum P=1.6×10−9), CAR T cell products manufactured from both CLL/SLL and B cell NHL patient populations consistently demonstrated clearance of non-T cells, including CD19+ cells, to below levels of quantitation. Conclusion Despite variation between B cell NHL and CLL/SLL patient leukapheresis, T cell enrichment before activation and transduction enables consistent downstream process performance and T cell purity, and a substantially reduced risk of transducing residual tumor cells. In addition, the reduction of terminally differentiated effector T cells and capacity to retain T cell diversity further improved consistency in product quality. Taken together, process modifications have enabled consistent manufacturing duration and quality of liso-cel product, which support operational efficiency and scalability for commercial production. Disclosures Teoh: Juno Therapeutics, a Celgene Company: Employment, Equity Ownership. Johnstone:Juno Therapeutics, a Celgene Company: Employment, Patents & Royalties: Author on a number of patent applications and invention disclosures relating to cell therapy and immunosequencing. Christin:Juno Therapeutics, a Celgene Company: Employment, Equity Ownership. Yost:Juno Therapeutics, a Celgene Company: Employment, Equity Ownership. Haig:Juno Therapeutics, a Celgene Company: Employment, Equity Ownership. Mallaney:Juno Therapeutics, a Celgene Company: Employment. Radhakrishnan:Juno Therapeutics, a Celgene Company: Employment, Equity Ownership. Gillenwater:Juno Therapeutics, a Celgene Company: Employment, Equity Ownership. Albertson:Juno Therapeutics, a Celgene Company: Employment, Equity Ownership. Guptill:Juno Therapeutics, a Celgene Company: Employment. Brown:Juno Therapeutics, a Celgene Company: Employment. Ramsborg:Juno Therapeutics, a Celgene Company: Employment, Equity Ownership, Patents & Royalties: Numerous patents. Hause:Juno Therapeutics, a Celgene Company: Employment, Equity Ownership. Larson:Juno Therapeutics, a Celgene Company: Employment, Equity Ownership.

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Production of Anti-CD19 CAR T Cells for ZUMA-3 and -4: Phase 1/2 Multicenter Studies Evaluating KTE-C19 in Patients With Relapsed/Refractory B-Precursor Acute Lymphoblastic Leukemia (R/R ALL)

Blood ◽

10.1182/blood.v128.22.1227.1227 ◽

2016 ◽

Vol 128 (22) ◽

pp. 1227-1227 ◽

Cited By ~ 4

Author(s):

Marianna Sabatino ◽

Kenny Choi ◽

Vijay Chiruvolu ◽

Marc Better

Keyword(s):

Clinical Trials ◽

T Cells ◽

T Cell ◽

Manufacturing Process ◽

Pediatric Patients ◽

Phase 1 ◽

Target Dose ◽

Employment Equity ◽

Car T ◽

Equity Ownership

Abstract Introduction: ZUMA-3 and -4 (NCT02614066 and NCT02625480) are ongoing phase 1-2 multicenter trials evaluating KTE-C19, a CD28/CD3ζ anti-CD19 chimeric antigen receptor (CAR) T cell therapy, in adult and pediatric patients respectively with R/R ALL. The manufacturing process developed to support the clinical trials aimed to accommodate variability in the incoming apheresis material, maximize manufacturing success rate, and minimize timing between leukapheresis collection and KTE-C19 administration at bedside. Methods: Once eligibility criteria were satisfied, patients underwent leukapheresis to process 12-15 L of blood (2-5 blood volumes for pediatric patients) to target collection of 10 × 109 mononuclear cells (MNCs). After collection, leukapheresis material was shipped to the central manufacturing site and processed to enrich T cells using an automated and closed system. The T cell enrichment step was implemented to remove cellular impurities such as leukemic blasts and myeloid cells that may negatively impact T cell activation and expansion. Enriched T cells were activated using an antigen presenting cell (APC)-free and bead-free method and cultured in serum-free medium containing 300 IU/mL of interleukin-2. Activated T cells were transduced with a gamma retroviral vector that encodes the CAR gene and further expanded to achieve target dose of 2 × 106 CAR-positive T cells/kg body weight (minimum of 1 × 106). Overall target duration of the manufacturing process was 8 days with an allowed range for harvest set between process days 6 and 10. Final KTE-C19 product was washed, formulated, cryopreserved and tested for identity, potency, and adventitious agents. After acceptance criteria were met, KTE-C19 was shipped to the clinical site using a validated cryoshipper. Results: As of July 2016, 5 of the 6 patients enrolled in the clinical trials received the target dose of 2 × 106 CAR-positive cells/kg body weight. The assigned dose could not be generated for one patient enrolled in the ZUMA-4 trial. In this patient, analysis performed on the leukapheresis material revealed >99% leukemic blasts. For the other 5 patients, the target dose was achieved within 6 - 7 process days notwithstanding a wide range in T cell composition of the incoming leukapheresis material (range, 4.7% to 81%), and wide variation in percent leukemic blasts (range, 11% to 92%). Median frequency of CAR expression in the KTE-C19 final product was consistent across product lots. The manufacturing process led to final products highly enriched for T cells with a median CD3+ cell content of >99%. Expansion of T cells was consistent among the 5 product lots with a mean 7-fold expansion from transduction to harvest. Median time from leukapheresis collection to final product release was 14 days. Conclusions: Reproducible manufacturing of high-quality, clinical-grade autologous CAR T cell products may be challenged by the inherent variability of starting material in patients with high content of leukemic blasts in peripheral blood. The bead- and serum-free manufacturing process developed to support the ZUMA-3 and -4 clinical trials was able to generate products within 6 -7 days with a low failure rate (including from patients with high percentages of leukemic blasts), and most importantly, a short duration from leukapheresis collection to shipment of KTE-C19 final product back to the clinical center for patient administration. Disclosures Sabatino: Kite: Employment, Equity Ownership. Choi:Kite Pharma: Employment, Equity Ownership. Chiruvolu:Kite Pharma: Employment, Equity Ownership. Better:Kite Pharma: Employment, Equity Ownership.

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Combination Immunotherapy with NY-ESO-1-Specific CAR+ T Cells with T-Cell Vaccine Improves Anti-Myeloma Effect

Blood ◽

10.1182/blood.v128.22.3366.3366 ◽

2016 ◽

Vol 128 (22) ◽

pp. 3366-3366 ◽

Cited By ~ 7

Author(s):

Krina Patel ◽

Simon Olivares ◽

Harjeet Singh ◽

Lenka V. Hurton ◽

Mary Helen Huls ◽

...

Keyword(s):

Multiple Myeloma ◽

T Cells ◽

T Cell ◽

Sleeping Beauty ◽

Employment Equity ◽

Effector Cells ◽

T Effector Cells ◽

Car T Cells ◽

Car T ◽

Equity Ownership

Abstract Adoptive transfer of T cells expressing chimeric antigen receptor (CAR) has demonstrated clinical effectiveness in early phase clinical trials, with persistence of effector cells typically leading to improved outcomes. Most CARs directly dock with cell-surface antigens, but this limits the number of tumor-derived targets. Thus, we have adapted two technologies to target intracellular antigens and improve survival of infused T cells. This was accomplished by expressing a CAR on T effector cells that functions as a mimetic of T-cell receptor (TCR) to recognize NY-ESO-1 in the context of HLA A2 and adapting HLA-A2+ T cells to serve as antigen presenting cells (T-APC) by expressing NY-ESO-1 antigen. NY-ESO-1 is a desirable target for T-cell therapy of high risk multiple myeloma (MM) with efficacy in trials infusing T cells expressing TCR recognizing this antigen. We hypothesized combined immunotherapy with NY-ESO-1-specific CAR+ T cells and an NY-ESO-1+ T-APC vaccine will lead to enhanced anti-myeloma efficacy due to improved persistence of the CAR+ T effector cells. An NY-ESO-1-specific CAR and control TCR were expressed on primary T cells using the Sleeping Beauty (SB) transposon/transposase system. T-APC was generated by electro-transfer of DNA plasmids from SB system coding for NY-ESO-1 and membrane-bound IL-15 (mbIL15). The tethered cytokine functions as co-stimulatory molecule to improve the potency of the vaccine. In vitro studies confirmed the NY-ESO-1-specific CAR+ (and TCR+) T cells could be numerically expanded upon co-culture with T-APC. A mouse model of NY-ESO-1+HLA-A2+(CD19neg) multiple myeloma was used to compare tumor growth for CAR+ T effector cells with and without T-APC. The NY-ESO-1-specific CAR+ T effector cells displayed anti-tumor effect that was superior to control mice without T cells and mice receiving CD19-specific control CAR+ T cells. Mice receiving both NY-ESO-1-specific CAR+T effector cells and T-APC exhibited further improvement in anti-myeloma activity. This group demonstrated superior persistence of T effector cells with recovered cells exhibiting a memory phenotype. In summary, T cells can target intracellular NY-ESO-1 using a TCR mimetic CAR. Improved anti-tumor effect attributed to better persistence can be achieved by co-infusion of T-APC vaccine. These data provide the foundation to assess T cells targeting NY-ESO-1 in a clinical trial. Disclosures Patel: Ziopharm Oncology: Equity Ownership, Patents & Royalties; Intrexon: Equity Ownership, Patents & Royalties. Olivares:Ziopharm Oncology: Equity Ownership, Patents & Royalties; Intrexon: Equity Ownership, Patents & Royalties. Singh:Ziopharm Oncology: Equity Ownership, Patents & Royalties; Immatics: Equity Ownership, Patents & Royalties; Intrexon: Equity Ownership, Patents & Royalties. Hurton:Ziopharm Oncology: Equity Ownership, Patents & Royalties; Intrexon: Equity Ownership, Patents & Royalties. Huls:Ziopharm Oncology: Equity Ownership, Patents & Royalties; Intrexon: Employment, Equity Ownership, Patents & Royalties. Cooper:City of Hope: Patents & Royalties; Intrexon: Equity Ownership; Ziopharm Oncology: Employment, Equity Ownership, Patents & Royalties; Targazyme, Inc.,: Equity Ownership; Immatics: Equity Ownership; Sangamo BioSciences: Patents & Royalties; MD Anderson Cancer Center: Employment; Miltenyi Biotec: Honoraria.

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