In Vitro Expansion of Anti-viral T Cells from Cord Blood by Accelerated Co-cultured Dendritic Cells

Abstract

Hematopoietic stem cell transplantation (HSCT) using unrelated cord blood (CB) donors is a suitable approach when an HLA-matched donor is not available. However, one important drawback is the risk of life-threatening viral infections prior to immune reconstitution, particularly from adenoviruses (AdVs). Although adoptive therapy with ex vivo expanded virus-reactive donor T cells has proven effective to treat these infections in HSCT recipients, the manufacturing process is complex and requires large numbers of cells, which is incompatible with CB donor units. Here, we have adapted our previous accelerated co-cultured dendritic cell (acDC) method, which allows to efficiently and rapidly expand peripheral blood T cells reactive to a given antigen, for use on limited CB material. Selected cytokine cocktails induced DC differentiation and maturation from unfractionated CB mononuclear cell cultures and simultaneously stimulated and expanded, within 10 days, functional CD8+ T cells specific for the model antigen MelanA or AdV immunodominant peptides. In addition, the use of G-Rex cultures yielded numbers of AdV-reactive CD8+ T cells compatible with adoptive cell therapy applications. Our acDC strategy, which uses reagents compatible with good manufacturing practices, may be promptly translated into the clinic for treating intercurrent infections in CB HSCT recipients.

Optimization of therapeutic T cell expansion in G-Rex device and applicability to large-scale production for clinical use

“…This, together with the efficient gas exchange and ready availability of abundant nutrients in culture medium, may explain the much more rapid and efficient expansion in G-Rex compared with T-flasks. G-Rex has been used by other groups for the optimal expansion of several types of blood cells, including tumor-infiltrating lymphocytes (TILs) [19,20], antigen-specific T cells [8,12,17,[33][34][35], gd T cells [23], Tregs [21,22], NKs [15], CIKs [25], CAR-Ts [16,24], megakaryocytes [36] and red blood cells [37] (reviewed in [9]). In most of these published cases, medium addition or exchange, as well as culture splitting, was performed with cell counting at various phases of the culture to define optimal dilution and collection times.…”

“…The G-Rex devices have been optimized for expansion of different types of T cells for research or clinical use [10,13,15,16]. Thus G-Rex devices have been used to produce antigen-specific T cells [8,12,17,18], tumor-infiltrating lymphocytes (TILs) [19,20], regulatory T cells (Tregs) [21,22], g/d T cells [23] and chimeric antigen receptor (CAR) T cells [16,24] as well as CIKs [25].…”

Lymphocyte expansion in bioreactors: upgrading adoptive cell therapy

“…That includes peer-reviewed guides for CTL from PBMC [ 182 , 183 ] and UCB [ 184 ], NK cells [ 185 ], TILs [ 186 ] and CAR-T cells [ 157 ]. When comparing its yield, the expansion of T cells in the G-rex generates up to 100 [ 173 ] to 1000 [ 158 ] times more cells than classical static culturing techniques, but there are lower yield exceptions [ 175 , 177 ]. In the same way, NK cells and TILs have also shown better or similar expansion performance, but the difference is moderate [ 106 , 159 , 180 ].…”

“…In TILs, the Phenotype has been found to be similar to static systems [ 180 ], with no evidence for cloning selection [ 159 ]. Functionality-wise, T cells grown in the G-rex preserve their cytolytic activity [ 158 , 160 , 176 ] and some research have found better [ 177 ] to similar [ 162 ] cytokine expression profile compared to static controls. The cytokine production of TIL grown in G-rex flasks is similar to TIL produced with 24-well plates, T-175 flasks, and bags [ 180 ].…”

“…E6 and E7 TCR transduced T-cells produced IFN-γ and TNF-α. Kuranda [ 177 ] - 2019 Used GM-CSF, IL-4, IL-1b, Flt3L, TLR8L, TNF-a, PGE2, IL-7, IL-2, IL-15, IL-7. AIMV 10% HS Ag or AdV5-reactive CD8+ from PBMC or CB 10 7 NP G-rex10 11 fold vs. std.…”