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Global Manufacturing of CAR T Cell Therapy.

ABSTRACT: Immunotherapy using chimeric antigen receptor-modified T cells has demonstrated high response rates in patients with B cell malignancies, and chimeric antigen receptor T cell therapy is now being investigated in several hematologic and solid tumor types. Chimeric antigen receptor T cells are generated by removing T cells from a patient's blood and engineering the cells to express the chimeric antigen receptor, which reprograms the T cells to target tumor cells. As chimeric antigen receptor T cell therapy moves into later-phase clinical trials and becomes an option for more patients, compliance of the chimeric antigen receptor T cell manufacturing process with global regulatory requirements becomes a topic for extensive discussion. Additionally, the challenges of taking a chimeric antigen receptor T cell manufacturing process from a single institution to a large-scale multi-site manufacturing center must be addressed. We have anticipated such concerns in our experience with the CD19 chimeric antigen receptor T cell therapy CTL019. In this review, we discuss steps involved in the cell processing of the technology, including the use of an optimal vector for consistent cell processing, along with addressing the challenges of expanding chimeric antigen receptor T cell therapy to a global patient population.

SUBMITTER: Levine BL

PROVIDER: S-EPMC5363291 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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Global Manufacturing of CAR T Cell Therapy.

Levine Bruce L BL Miskin James J Wonnacott Keith K Keir Christopher C

Molecular therapy. Methods & clinical development 20161231

Immunotherapy using chimeric antigen receptor-modified T cells has demonstrated high response rates in patients with B cell malignancies, and chimeric antigen receptor T cell therapy is now being investigated in several hematologic and solid tumor types. Chimeric antigen receptor T cells are generated by removing T cells from a patient's blood and engineering the cells to express the chimeric antigen receptor, which reprograms the T cells to target tumor cells. As chimeric antigen receptor T cel ...[more]

PMID: 28344995

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Project description:BackgroundChimeric antigen receptor (CAR)-T cell has revolutionary efficacy against relapsed/refractory multiple myeloma (R/R MM). However, current CAR-T cell therapy has several limitations including long vein-to-vein time and limited viability.MethodsA 4-1BB-costimulated B-cell maturation antigen (BCMA) CAR-T integrating an independently-expressed OX40 (BCMA-BBZ-OX40) was designed and generated by a traditional manufacturing process (TraditionCART) or instant manufacturing platform (named InstanCART). The tumor-killing efficiency, differentiation, exhaustion, and expansion level were investigated in vitro and in tumor-bearing mice. An investigator-initiated clinical trial was performed in patients with R/R MM to evaluate the outcomes of both TraditionCART and InstanCART. The primary objective was safety within 1 month after CAR-T cell infusion. The secondary objective was the best overall response rate.ResultsPreclinical studies revealed that integrated OX40 conferred BCMA CAR-T cells with superior cytotoxicity and reduced exhaustion levels. InstanCART process further enhanced the proliferation and T-cell stemness of BCMA-BBZ-OX40 CAR-T cells. BCMA-BBZ-OX40 CAR-T cells were successfully administered in 22 patients with R/R MM, including 15 patients with TraditionCART and 7 patients with InstanCART. Up to 50% (11/22) patients had a high-risk cytogenetic profile and 36% (8/22) had extramedullary disease. CAR-T therapy caused grade 1-2 cytokine release syndrome in 19/22 (80%) patients, grade 1 neurotoxicity in 2/22 (9%) patients and led to ≥grade 3 adverse events including neutropenia (20/22, 91%), thrombocytopenia (15/22, 68%), anemia (12/22, 55%), creatinine increased (1/22, 5%), hepatic enzymes increased (5/22, 23%), and sepsis (1/22, 5%). The best overall response rate was 100%, and 64% (14/22) of the patients had a complete response or better. The median manufacturing time was shorter for InstanCART therapy (3 days) than for TraditionCART therapy (10 days). Expansion and duration were dramatically higher for InstanCART cells than for TraditionCART cells.ConclusionsBCMA-BBZ-OX40 CAR-T cells were well tolerated and exhibited potent responses in patients with R/R MM. InstanCART shortened the manufacturing period compared to TraditionCART, and improved the cellular kinetics. Our results demonstrated the potency and feasibility of OX40-modified BCMA CAR-T cells using InstanCART technology for R/R MM therapy.Trial registration numberThis trial was registered at www.Clinicaltrialsgov as #NCT04537442.

Project description:Genetic reprogramming of immune cells to recognize and target tumor cells offers a possibility of long-term cure. Cell therapies, however, lack simple and affordable manufacturing workflows, especially to genetically edit immune cells to more effectively target cancer cells and avoid immune suppression mechanisms. Microfluidics is a pathway to improve the manufacturing precision of gene modified cells. However, to date, it remains to be demonstrated that microfluidic treatment preserves the functionality of T cell products in a complete workflow. In this study, we used microfluidics to perform CRISPR/Cas9 gene editing of CD5, a negative T-cell regulator, followed by the insertion of a chimeric antigen receptor (CAR) transgene via lentiviral vector transduction to generate CAR T cells targeted against the B cell antigen CD19. As part of the workflow, we have optimized a microfluidic device that relies on convective volume exchange between cells and surrounding fluid to deliver guide RNA and Cas9 ribonucleoprotein to primary T cells. We comprehensively tested critical design features of the device to improve the gene-edited product yield. By combining high-speed video and cell mechanics measurements using the atomic force microscope, we validate a model that relates the device design features to cell properties. Our findings showed enhanced performance was obtained by focusing the cells to counteract the flow resistance caused by the ridge constrictions, providing a ridge layout that allows sufficient cycles of compression and time for volume recovery, and including a gutter to clear aggregates that could reduce cell viability. The optimized device was used in a workflow to generate CD5-knockout CD19 CAR T cells. The microfluidics approach resulted in >60% CD5 editing efficiency, ≥80% cell viability, similar memory phenotype composition as unprocessed cells, and superior cell growth. The microfluidics workflow yielded 4-fold increase of edited T cells compared to an electroporation workflow post-expansion. The transduced CAR T cells showed similar transduction efficiency and cytotoxicity against CD19-positive leukemia cells. Moreover, patient-derived T cells showed the ability to be similarly edited, though their distinct biomechanics resulted in slightly lower outcomes. Microfluidics-based manufacturing is a promising path towards more productive clinical manufacturing of gene edited CAR T cells.

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Global Manufacturing of CAR T Cell Therapy.

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Global Manufacturing of CAR T Cell Therapy.

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