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

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.

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