BCR-ABL-specific CD4+ T-helper cells promote the priming of antigen-specific cytotoxic T cells via dendritic cells.

The advent of tyrosine kinase inhibitor (TKI) therapy markedly improved the outcome of patients with chronic-phase chronic myeloid leukemia (CML). However, the poor prognosis of patients with advanced-phase CML and the lifelong dependency on TKIs are remaining challenges; therefore, an effective therapeutic has been sought. The BCR-ABL p210 fusion protein's junction region represents a leukemia-specific neoantigen and is thus an attractive target for antigen-specific T-cell immunotherapy. BCR-ABL p210 fusion-region-specific CD4+ T-helper (Th) cells possess antileukemic potential, but their function remains unclear. In this study, we established a BCR-ABL p210 b3a2 fusion-region-specific CD4+ Th-cell clone (b3a2-specific Th clone) and examined its dendritic cell (DC)-mediated antileukemic potential. The b3a2-specific Th clone recognized the b3a2 peptide in the context of HLA-DRB1*09:01 and exhibited a Th1 profile. Activation of this clone through T-cell antigen receptor stimulation triggered DC maturation, as indicated by upregulated production of CD86 and IL-12p70 by DCs, which depended on CD40 ligation by CD40L expressed on b3a2-specific Th cells. Moreover, in the presence of HLA-A*24:02-restricted Wilms tumor 1 (WT1)235-243 peptide, DCs conditioned by b3a2-specific Th cells efficiently stimulated the primary expansion of WTI-specific cytotoxic T lymphocytes (CTLs). The expanded CTLs were cytotoxic toward WT1235-243-peptide-loaded HLA-A*24:02-positive cell lines and exerted a potent antileukemic effect in vivo. However, the b3a2-specific Th-clone-mediated antileukemic CTL responses were strongly inhibited by both TKIs and interferon-α. Our findings indicate a crucial role of b3a2-specific Th cells in leukemia antigen-specific CTL-mediated immunity and provide an experimental basis for establishing novel CML immunotherapies.

Hepatocellular carcinoma cell sensitivity to Vγ9Vδ2 T lymphocyte-mediated killing is increased by zoledronate.

The limited efficacy of vaccines in hepatocellular carcinoma (HCC), due to the low frequency of tumor-infiltrating cytotoxic T lymphocytes (CTLs), indicates the importance of innate immune surveillance, which assists acquired immunity by directly recognizing and eliminating HCC. Innate Vγ9Vδ2 T cells have major histocompatibility complex-unrestricted antitumor activity and are activated by phosphoantigens, which are upregulated in cancer cells by the nitrogen-containing bisphosphonate, zoledronate (Zol). A better understanding of HCC susceptibility to Zol and downstream γδ T cell-mediated killing is essential to optimize γδ T cell-mediated immunotherapy. This study systematically examined the interactions between γδ T cells and Zol-treated HCC cell lines (HepG2, HLE, HLF, HuH-1, JHH5, JHH7, and Li-7) in vitro. All HCC cell lines expressed the DNAX accessory molecule-1 ligands, poliovirus receptor, and Nectin-2, and γδ T cell-mediated killing of these cells was significantly enhanced by Zol. Small interfering RNA-mediated knockdown of these ligands did not affect the susceptibility to γδ T cell lysis. This killing activity was partly inhibited by mevastatin, an inhibitor of the mevalonate pathway, and markedly reduced by a monoclonal antibody to γ- and δ-chain T cell receptor, indicating that this is crucial for Zol-induced HCC killing. In addition, Zol-treated HCC cell lines triggered γδ T cell proliferation and induced production of Th1 and Th2, but not Th17, cytokines. The Zol concentration that enhanced HCC cell susceptibility to γδ T cell killing was lower than that required to directly inhibit HCC proliferation. Thus, γδ T cells may be important effector cells in the presence of Zol, especially where there are insufficient number of cancer antigen-specific CTLs to eliminate HCC. Our in vitro data support the proposal that Zol-treatment, combined with adaptive γδ T cell immunotherapy, may provide a feasible and effective approach for treatment of HCC.

Vaccination with liposome-coupled glypican-3-derived epitope peptide stimulates cytotoxic T lymphocytes and inhibits GPC3-expressing tumor growth in mice.

Because therapeutic manipulation of immunity can induce tumor regression, anti-cancer immunotherapy is considered a promising treatment modality. We previously reported that glypican-3 (GPC3), an oncofetal antigen overexpressed in hepatocellular carcinoma (HCC), is a useful target for cytotoxic T lymphocyte (CTL)-mediated cancer immunotherapy, and we have performed clinical trials using the GPC3-derived peptide vaccine. Although vaccine-induced GPC3-peptide-specific CTLs were often tumor reactive in vitro and were correlated with overall survival, no complete response was observed. In the current study, we synthesized liposome-coupled GPC3-derived CTL epitope peptide (pGPC3-lipsome) and investigated its antitumor potential. Vaccination with pGPC3-liposome induced peptide-specific CTLs at a lower dose than conventional vaccine emulsified in incomplete Freund's adjuvant. Coupling of pGPC3 to liposomes was essential for effective priming of GPC3-specific CTLs. In addition, immunization with pGPC3-liposome inhibited GPC3-expressing tumor growth. Thus, vaccination with tumor-associated antigen-derived epitope peptides coupled to the surfaces of liposomes may be a novel therapeutic strategy for cancer.

Enhancement of antitumor effect by peptide vaccine therapy in combination with anti-CD4 antibody: Study in a murine model.

PURPOSE: The clinical efficacy of cancer peptide vaccine therapy is insufficient. To enhance the anti-tumor effect of peptide vaccine therapy, we combined this therapy with an anti-CD4 mAb (GK1.5), which is known to deplete CD4+ cells, including regulatory T cells (Tregs). METHODS: To determine the treatment schedule, the number of lymphocyte subsets in the peripheral blood of mice was traced by flow cytometry after administration of anti-CD4 mAb. The ovalbumin (OVA)257-264 peptide vaccine was injected intradermally and anti-CD4 mAb was administered intraperitoneally into C57BL/6 mice at different schedules. We evaluated the enhancement of OVA peptide-specific cytotoxic T lymphocyte (CTL) induction in the combination therapy using the ELISPOT assay, CD107a assay, and cytokine assay. We then examined the in vivo metastasis inhibitory effect by OVA peptide vaccine therapy in combination with anti-CD4 mAb against OVA-expressing thymoma (EG7) in a murine liver metastatic model. RESULTS: We showed that peptide-specific CTL induction was enhanced by the peptide vaccine in combination with anti-CD4 mAb and that the optimized treatment schedule had the strongest induction effect of peptide-specific CTLs using an IFN-γ ELISPOT assay. We also confirmed that the CD107a+ cells secreted perforin and granzyme B and the amount of IL-2 and TNF produced by these CTLs increased when the peptide vaccine was combined with anti-CD4 mAb. Furthermore, metastasis was inhibited by peptide vaccines in combination with anti-CD4 mAb compared to peptide vaccine alone in a murine liver metastatic model. CONCLUSION: The use of anti-CD4 mAb in combination with the OVA peptide vaccine therapy increased the number of peptide-specific CTLs and showed a higher therapeutic effect against OVA-expressing tumors. The combination with anti-CD4 mAb may provide a new cancer vaccine strategy.