Anti-CCR7 Monoclonal antibody, clone 4E23 [PE]

The finding that dendritic cells (DCs) orchestrate innate and adaptive immune responses has stimulated research on harnessing DCs for developing more effective vaccines for DC therapy. The expression of cytomegalovirus (CMV) antigens in glioblastoma multiforme (GBM) presents a unique opportunity to target these viral proteins for tumour immunotherapy. Here, we demonstrate that V9T cells, innate immune cells activated by zoledronate (Z) and V24 natural killer (V24NK) cells, innate/adaptive immune cells activated by -galactosylceramide (G) can link innate and adaptive immunities through cross-talk with interferon (IFN) DCs from patients with glioblastoma multiforme (GBM) and healthy donors in a manner that can amplify the activation and proliferation of CMVpp65-specific CD8(+) T cells. The IFN DCs derived from patients with GBM used in this study express lower levels of programmed cell death ligand (PD)-L1 and PD-L2 and higher levels of C-C receptor 7 (CCR7) than the most commonly used mature interleukin (IL)-4 DCs. The expression level of programmed cell death 1 (PD-1) on CD8(+) T cells, including CMVpp65-specific CD8(+) T cells, expanded by IFN DCs pulsed with the CMVpp65-peptide and Z plus G (IFN DCs/P+Z+G), was lower than that expanded by IFN DCs pulsed with the peptide alone (IFN DCs/P). Multi-functional T cells, including human leucocyte antigen (HLA)-A*0201-restricted CMVpp65-specific CD8(+) T cells, V9T cells and V24NKT cells, efficiently kill the HLA-A*0201-positive GBM cell line expressing CMVpp65 protein (T98G). These findings indicate that DC therapy using IFN DCs/P+Z+G and/or CTL therapy using CMVpp65-specific CD8(+) T cells expanded by IFN DCs/P+Z+G may lead to a good clinical outcome for patients with GBM.

Improvements in the quality and fitness of chimeric antigen receptor (CAR)-engineered T cells, through CAR design or manufacturing optimizations, could enhance the therapeutic potential of CAR-T cells. One parameter influencing the effectiveness of CAR-T cell therapy is the differentiation status of the final product: CAR-T cells that are less-differentiated and less exhausted are more therapeutically effective. In the current study, we demonstrate that CAR-T cells expanded in IL15 (CAR-T/IL15) preserve a less-differentiated stem cell memory (Tscm) phenotype, defined by expression of CD62L(+) CD45RA(+) CCR7(+), as compared with cells cultured in IL2 (CAR-T/IL2). CAR-T/IL15 cells exhibited reduced expression of exhaustion markers, higher antiapoptotic properties, and increased proliferative capacity upon antigen challenge. Furthermore, CAR-T/IL15 cells exhibited decreased mTORC1 activity, reduced expression of glycolytic enzymes and improved mitochondrial fitness. CAR-T/IL2 cells cultured in rapamycin (mTORC1 inhibitor) shared phenotypic features with CAR-T/IL15 cells, suggesting that IL15-mediated reduction of mTORC1 activity is responsible for preserving the Tscm phenotype. CAR-T/IL15 cells promoted superior antitumor responses in vivo in comparison with CAR-T/IL2 cells. Inclusion of cytokines IL7 and/or IL21 in addition to IL15 reduced the beneficial effects of IL15 on CAR-T phenotype and antitumor potency. Our findings show that IL15 preserves the CAR-T cell Tscm phenotype and improves their metabolic fitness, which results in superior in vivo antitumor activity, thus opening an avenue that may improve future adoptive T-cell therapies.