Ex vivo expansion of human T cells for adoptive immunotherapy using the novel Xeno-free CTS Immune Cell Serum Replacement

The manufacture of clinical grade cellular products for adoptive immunotherapy requires ex vivo culture and expansion of human T cells. One of the key components in manufacturing of T cell therapies is human serum (HS) or fetal bovine serum (FBS), which can potentially expose immunotherapy recipient to adventitious infectious pathogens and are thus considered as non-cGMP compliant for adoptive therapy. Here we describe a novel xeno-free serum replacement (SR) with defined components that can be reproducibly used for the production of clinical grade T-cell therapies in combination with several different cell culture media. Dynabeads CD3/CD28 Cell Therapy System (CTS)-activated or antigen-specific T cells expanded using the xeno-free SR, CTS Immune Cell SR, showed comparable growth kinetics observed with cell culture media supplemented with HS or FBS. Importantly the xeno-free SR supplemented medium supported the optimal expansion of T cells specific for subdominant tumour-associated antigens and promoted expansion of T cells with central memory T-cell phenotype, which is favourable for in vivo survival and persistence following adoptive transfer. Furthermore, T cells expanded using xeno-free SR medium were highly amenable to lentivirus-mediated gene transduction for potential application for gene-modified T cells. Taken together, the CTS Immune Cell SR provides a novel platform strategy for the manufacture of clinical grade adoptive cellular therapies

Antibody-mediated delivery of T-cell epitopes to antigen-presenting cells induce strong CD4 and CD8 T-cell responses

Targeted delivery of antigen to antigen-presenting cells (APCs) enhances antigen presentation and thus, is a potent strategy for making more efficacious vaccines. This can be achieved by use of antibodies with specificity for endocytic surface molecules expressed on the APC. We aimed to compare two different antibody-antigen fusion modes in their ability to induce T-cell responses; first, exchange of immunoglobulin (Ig) constant domain loops with a T-cell epitope (Troybody), and second, fusion of T-cell epitope or whole antigen to the antibody C-terminus. Although both strategies are well-established, they have not previously been compared using the same system. We found that both antibody-antigen fusion modes led to presentation of the T-cell epitope. The strength of the T-cell responses varied, however, with the most efficient Troybody inducing CD4 T-cell proliferation and cytokine secretion at 10–100-fold lower concentration than the antibodies carrying antigen fused to the C-terminus, both in vitro and after intravenous injection in mice. Furthermore, we exchanged this loop with an MHCI-restricted T-cell epitope, and the resulting antibody enabled efficient cross-presentation to CD8 T cells in vivo. Targeting of antigen to APCs by use of such antibody-antigen fusions is thus an attractive vaccination strategy for increased activation of both CD4 and CD8 peptide-specific T cells

Novel APC-like properties of human NK cells directly regulate T cell activation

Initiation of the adaptive immune response is dependent on the priming of naive T cells by APCs. Proteomic analysis of unactivated and activated human NK cell membrane–enriched fractions demonstrated that activated NK cells can efficiently stimulate T cells, since they upregulate MHC class II molecules and multiple ligands for TCR costimulatory molecules. Furthermore, by manipulating antigen administration, we show that NK cells possess multiple independent unique pathways for antigen uptake. These results highlight NK cell–mediated cytotoxicity and specific ligand recognition by cell surface–activating receptors on NK cells as unique mechanisms for antigen capturing and presentation. In addition, we analyzed the T cell–activating potential of human NK cells derived from different clinical conditions, such as inflamed tonsils and noninfected and CMV-infected uterine decidual samples, and from transporter-associated processing antigen 2–deficient patients. This in vivo analysis revealed that proinflammatory, but not immune-suppressive, microenvironmental requirements can selectively dictate upregulation of T cell–activating molecules on NK cells. Taken together, these observations offer new and unexpected insights into the direct interactions between NK and T cells and suggest novel APC-like activating functions for human NK cells