Background Lifileucel (LN-144) and LN-145, adoptive cell therapies using autologous tumor-infiltrating lymphocytes (TIL), have demonstrated encouraging efficacy with acceptable safety in a variety of tumor types.1–4 The lifileucel Gen 2 clinical manufacturing process uses gas-permeable rapid expansion (G-Rex®, Wilson Wolf, Saint Paul, MN) bioreactors for TIL expansion.5 Static gas-permeable cell culture bags (EXP-Pak™, Charter Medical, Winston-Salem, NC) are alternate bioreactors that have been used for clinical manufacturing of T cells.6 In this study, TIL product characteristics were compared after expansion at small- and full-scale using EXP-Pak bags and G-Rex bioreactors.
Methods Cryopreserved pre-REP TIL were cultured with OKT-3 and irradiated peripheral blood mononuclear cells in either small-scale (G-Rex 5M flasks or EXP-50 bags) or full-scale (G-Rex 500MCS or EXP-5L bags) conditions. The same culture media formulation was used throughout the process. Final harvested TIL products were characterized for the following quality attributes: total viable cells (TVC), purity (% viability), identity (% CD45+CD3+), and activity (IFN-gamma release). Additional characterization was performed to determine the TIL differentiation, central and effector memory subsets, activation, exhaustion status, and impurities using multi-color flow cytometry. The T-cell receptor (TCR) repertoires of the final products were assessed for unique CDR3 (uCDR3) counts and shared clones using RNA-seq.
Results The median (range) TVC from 6 small-scale runs of G-Rex condition was 90.8×109 (26.9×109–98.6×109) cells, and the corresponding TVC of EXP-Pak bag condition was 119×109 (63.2×109–141×109) cells. Full-scale runs (n=3) yielded similar TVC for G-Rex and EXP-Pak bag conditions. Both conditions had comparable purity, identity, and activity (table 1). 91–99% of TCR Vbeta clones of EXP-Pak bag TIL were present in G-Rex (table 2). Cell proliferation, cell cycle, and mitochondrial function of TIL generated from EXP-Pak bags and G-Rex flask methodologies will be presented.
Conclusions The final TIL product generated in the EXP-Pak bag condition did not differ from cells produced in the G-Rex flask functionally or phenotypically and demonstrated similar growth profiles. These data support further evaluation of EXP-Pak bags for potential use in clinical and potential commercial TIL cell therapy manufacturing applications.
Acknowledgements This study and analysis were funded by Iovance Biotherapeutics, Inc. (San Carlos, CA, USA). Editorial support was provided by Amanda Kelly (Iovance).
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