With recent advances in cancer vaccination therapy targeting tumor-associated antigens (TAAs), dendritic cells (DCs) are considered to play a central role as a cell-based drug delivery system in the bioactive immune environment. Ex vivo generation of monocyte-derived DCs has been conventionally applied in adherent manufacturing systems with separate loading of TAAs before clinical use. We developed DCs pre-pulsed with Wilms’ tumor (WT1) peptides in low-adhesion culture maturation (WT1-DCs). Quality tests (viability, phenotype, and functions) of WT1-DCs were performed for process validation, and findings were compared with those for conventional DCs (cDCs). In comparative analyses, WT1-DCs showed an increase in viability and recovery of the DC/monocyte ratio, displaying lower levels of IL-10 (an immune suppressive cytokine) and a similar antigen-presenting ability in an in vitro cytotoxic T lymphocytes (CTLs) assay with cytomegalovirus, despite lower levels of CD80 and PD-L2. A clinical study revealed that WT1-specific CTLs (WT1-CTLs) were detected upon using the WT1-DCs vaccine in patients with cancer. A DC vaccine containing TAAs produced under an optimized manufacturing protocol is a potentially promising cell-based drug delivery system to induce acquired immunity.

Given the recent advancements of immune checkpoint inhibitors, there is considerable interest in cancer immunotherapy provided through dendritic cell (DC)-based vaccination. Although many studies have been conducted to determine the potency of DC vaccines against cancer, the clinical outcomes are not yet optimal, and further improvement is necessary. In this study, we evaluated the potential ability of human platelet lysate (HPL) to produce interferon-α-induced DCs (IFN-DCs). In the presence of HPL, IFN-DCs (HPL-IFN-DCs) displayed high viability, yield, and purity. Furthermore, HPL-IFN-DCs displayed increased CD14, CD56, and CCR7 expressions compared with IFN-DCs produced without HPL; HPL-IFN-DCs induced an extremely higher number of antigen-specific cytotoxic T lymphocytes (CTLs) than IFN-DCs, which was evaluated with a human leukocyte antigen (HLA)-restricted melanoma antigen recognized by T cells 1 (MART-1) peptide. Additionally, the endocytic and proteolytic activities of HPL-IFN-DCs were increased. Cytokine production of interleukin (IL)-6, IL-10, and tumor necrosis factor (TNF)-α was also elevated in HPL-IFN-DCs, which may account for the enhanced CTL, endocytic, and proteolytic activities. Our findings suggest that ex-vivo-generated HPL-IFN-DCs are a novel monocyte-derived type of DC with high endocytic and proteolytic activities, thus highlighting a unique strategy for DC-based immunotherapies.

Significant recent advances in cancer immunotherapeutics include the vaccination of cancer patients with tumor antigen-associated peptide-pulsed dendritic cells (DCs). DC vaccines with homogeneous, mature, and functional activities are required to achieve effective acquired immunity; however, the yield of autologous monocyte-derived DCs varies in each patient. Priming with a low dose of recombinant human granulocyte colony-stimulating factor (rhG-CSF) 16–18 h prior to apheresis resulted in 50% more harvested monocytes, with a significant increase in the ratio of CD11c+CD80+ DCs/apheresed monocytes. The detection of antigen-specific cytotoxic T lymphocytes after Wilms’ tumor 1-pulsed DC vaccination was higher in patients treated with rhG-CSF than those who were not, based on immune monitoring using tetramer analysis. Our study is the first to report that DC vaccines for cancer immunotherapy primed with low-dose rhG-CSF are expected to achieve higher acquired immunogenicity.

Dendritic cell (DC) vaccines for cancer immunotherapy have been actively developed to improve clinical efficacy. In our previous report, monocyte−derived DCs induced by interleukin (IL)−4 with a low−adherence dish (low−adherent IL-4−DCs: la−IL-4−DCs) improved the yield and viability, as well as relatively prolonged survival in vitro, compared to IL-4−DCs developed using an adherent culture protocol. However, la−IL-4−DCs exhibit remarkable cluster formation and display heterogeneous immature phenotypes. Therefore, cluster formation in la−IL-4−DCs needs to be optimized for the clinical development of DC vaccines. In this study, we examined the effects of cluster control in the generation of mature IL-4−DCs, using cell culture vessels and measuring spheroid formation, survival, cytokine secretion, and gene expression of IL-4−DCs. Mature IL-4−DCs in cell culture vessels (cluster−controlled IL-4−DCs: cc−IL-4−DCs) displayed increased levels of CD80, CD86, and CD40 compared with that of la−IL-4−DCs. cc−IL-4−DCs induced antigen−specific cytotoxic T lymphocytes (CTLs) with a human leukocyte antigen (HLA)−restricted melanoma antigen recognized by T cells 1 (MART−1) peptide. Additionally, cc−IL-4−DCs produced higher levels of IFN−γ, possessing the CTL induction. Furthermore, DNA microarrays revealed the upregulation of BCL2A1, a pro−survival gene. According to these findings, the cc−IL-4−DCs are useful for generating homogeneous and functional IL-4−DCs that would be expected to promote long−lasting effects in DC vaccines.

Knee osteoarthritis (Knee OA) is an irreversible condition that causes bone deformity and degeneration of the articular cartilage that comprises the joints, resulting in chronic pain and movement disorders. The administration of cultured adipose-derived stem cells (ADSCs) into the knee joint cavity improves the clinical symptoms of Knee OA; however, the effect of synovial fluid (SF) filling the joint cavity on the injected ADSCs remains unclear. In this study, we investigated the effect of adding SF from Knee OA patients to cultured ADSCs prepared for therapeutic use in an environment that mimics the joint cavity. An increase in the viability of ADSCs was observed following the addition of SF. Gene expression profiling of SF-treated ADSCs using DNA microarrays revealed changes in several genes involved in cell survival. Of these genes, we focused on FOSL1, which is involved in the therapeutic effect of ADSCs and the survival and proliferation of cancer stem cells. We confirmed the upregulation of FOSL1 mRNA and protein expression using RT-PCR and western blot analysis, respectively. Next, we knocked down FOSL1 in ADSCs using siRNA and observed a decrease in cell viability, indicating the involvement of FOSL1 in the survival of ADSCs. Interestingly, in the knockdown cells, ADSC viability was also decreased by SF exposure. These results suggest that SF enhances cell viability by upregulating FOSL1 expression in ADSCs. For therapy using cultured ADSCs, the therapeutic effect of ADSCs may be further enhanced if an environment more conducive to the upregulation of FOSL1 expression in ADSCs can be established.