Chimeric NKG2D receptor–expressing T cells as an immunotherapy for multiple myeloma

Abstract

Objective Most myeloma tumor cells from patients express NKG2D ligands. We have reported the development of a chimeric NKG2D receptor (chNKG2D), which consists of the NKG2D receptor fused to the CD3ζ chain. T cells expressing this receptor kill and produce cytokines in response to NKG2D-ligand+ tumor cells. Therefore we investigated whether human chNKG2D T cells respond against human myeloma cells. Methods ChNKG2D T cells were generated from healthy donors and myeloma patients. The effector phase of chNKG2D T cells was analyzed by cell-surface marker expression and human myeloma cell lines were tested for expression of NKG2D ligands. Lysis of myeloma cell lines and cytokine secretion by chNKG2D T cells was determined. ChNKG2D T cells grown in serum-free media, or cyropreserved, were assessed for effector cell functions. Results Myeloma cell lines expressed NKG2D ligands. ChNKG2D T cells from healthy donors and myeloma patients lysed myeloma cells, and secreted proinflammatory cytokines when cultured with myeloma cells or patient bone marrow but not with PBMCs or normal bone marrow. Lysis of myeloma cells was dependent on chNKG2D T cell expression of NKG2D and perforin. Additionally, chNKG2D T cells upregulated CD45RO, did not express CD57, and maintained expression of CD27, CD62L, and CCR7, indicating that the T cells were at an early effector stage. Finally, we showed that chNKG2D T cells generated with serum-free media, or when cryopreserved, maintained effector functions. Conclusion ChNKG2D T cells respond to human myeloma cells and can be generated using clinically applicable cell culture techniques.

Ex vivo culture of chimeric antigen receptor T cells generates functional CD8+ T cells with effector and central memory-like phenotype

“…The T-cell expansion protocols used by Barber et al 29 and our study were similar and generated both memory and 'early Eff' CD8 + T-cell phenotype. In contrast, Till et al…”

“…By the end of the transduction culture, the CD8 + LeY-T cells 30 produced CD8 + T cells that were more differentiated and largely Eff (CD62LÀ/CCR7À/ CD45RAÀ/CD127À) cells. Although the T-cell activation step (OKT3 plus IL-2 for 3 days) was the same for all three studies, in the Barber et al 29 and our study the T cells were subsequently retrovirally transduced and expanded in IL-2 for 5 or 7 days, respectively. In contrast, Till et al…”

“…Furthermore, the chimeric NKG2D T cells expressed early differentiation markers, including CD27, CCR7 and CD62L. 29 In addition, Till et al 30 described the phenotype of chimeric receptor-expressing CD8 + T cells directed to CD20. These cells were produced after a lengthy re-stimulation protocol and were largely Eff cells…”

“…However, until recently, equivalent data for CAR T cells have been absent. Barber et al 29 recently described CD8 + T cells expressing a chimeric NKG2D receptor after a short-term transduction and expansion culture. At the end of transduction culture, the chimeric NKG2D T cells had cytolytic function against human myeloma cell lines, and secreted IFN-g, granulocytemacrophage colony-stimulating factor, tumor necrosis factor (TNF)-a and CCL5 after co-culture with myeloma cell lines.…”

Costimulation of Effector CD8+ T Cells: Which Receptor is Optimal for Immunotherapy?

“…Instead it seems that the differences in T cell function induced by these receptors work in a tumor-specific manner, with one combination of signals not being ideal for every tumor type [32][33][34][35]. Thus, the best combination of anti-tumor functions will likely have to be optimized for each tumor.…”