Junji Yatsuda

Identification of promiscuous KIF20A long peptides bearing both CD4+ and CD8+ T-cell epitopes: KIF20A-specific CD4+ T-cell immunity in patients with malignant tumor

Purpose: To identify long peptides (LP) derived from a novel tumor-associated antigen (TAA), kinesin family member 20A (KIF20A), which induce tumor-specific T-helper type 1 (TH1) cells and CTLs. Experimental Design: We combined information from a recently developed computer algorithm predicting HLA class II–binding peptides with KIF20A-derived CTL-epitope sequences presented by HLA-A2 (A*02:01) or HLA-A24 (A*24:02) to select candidate promiscuous TH1-cell epitopes containing CTL epitopes. Peripheral blood mononuclear cells (PBMC) derived from healthy donors or patients with head-and-neck malignant tumor (HNMT) were used to study the immunogenicity of KIF20A-LPs, and the in vitro cross-priming potential of KIF20A-LPs bearing CTL epitopes. We used HLA-A24 transgenic mice to address whether vaccination with KIF20A-LP induces efficient cross-priming of CTLs in vivo. The TH1-cell response to KIF20A-LPs in HNMT patients receiving immunotherapy with TAA-derived CTL-epitope peptides was analyzed using IFN-γ enzyme-linked immunospot assays. Results: We identified promiscuous KIF20A-LPs bearing naturally processed epitopes recognized by CD4+ T cells and CTLs. KIF20A-specific CTLs were induced by vaccination with a KIF20A-LP in vivo. KIF20A expression was detected in 55% of HNMT by immunohistochemistry, and significant frequencies of KIF20A-specific TH1 cell responses were detected after short-term in vitro stimulation of PBMCs with KIF20A-LPs in 50% of HNMT patients, but not in healthy donors. Furthermore, these responses were associated with KIF20A expression in HNMT tissues. Conclusions: These are the first results showing the presence of KIF20A-specific TH1 cell responses in HNMT patients and underline the possible utility of KIF20A-LPs for propagation of TH1 cells and CTLs. Clin Cancer Res; 19(16); 4508–20. ©2013 AACR.

C5aR is frequently expressed in metastatic renal cell carcinoma and plays a crucial role in cell invasion via the ERK and PI3 kinase pathways

The anaphylatoxin C5a is a chemoattractant for leukocyte migration via the C5a receptor (C5aR). We recently reported that C5aR was aberrantly expressed in a wide variety of human related cancers, while it also promotes cancer cell invasion by C5a stimulation. However, the biological significance of C5aR expression in renal cell carcinoma (RCC) has not yet been clarified. In the present study, we aimed to elucidate the biological role of C5aR in RCC progression. Clinical RCC specimens were analyzed for C5aR expression and its relationship with baseline demographic data and clinicopathological parameters was analyzed. Moreover, renal carcinoma Renca cells stably expressing C5aR were generated and used to assess the effects of C5a-C5aR axis activation on various cellular phenomena in culture. Immunohistochemistry revealed that 96.7% of the metastatic RCCs (mRCCs) showed C5aR expression, whereas only 50.5% of the non-metastatic RCCs expressed C5aR (P<0.001). Although C5a stimulation did not significantly alter anoikis of C5aR‑expressing Renca cells, C5a elicited cell morphological change and scattering of those cells accompanied with dynamic actin rearrangement, which was not observed in the Renca cells harboring the empty vector only. Moreover, C5a triggered ERK and PI3K‑dependent invasion of the C5aR-expressing renal carcinoma cells. These results are consistent with the idea that the C5a-C5aR axis plays a crucial role in renal carcinoma cell invasion, which may be one of the key steps for RCC metastasis. The present study provides proof‑of‑concept that the C5a-C5aR axis may be a useful therapeutic target for preventing RCC progression.

Identification of immunogenic LY6K long peptide encompassing both CD4+ and CD8+ T-cell epitopes and eliciting CD4+ T-cell immunity in patients with malignant disease

Identification of peptides that activate both tumor-specific helper T (Th) cells and cytotoxic T lymphocytes (CTLs) are important for the induction of effective antitumor immune responses. We focused on a long peptide (LP) derived from lymphocyte antigen 6 complex locus K (LY6K) encompassing both candidate Th epitopes and a known CTL epitope. Using IFNγ ELISPOT assays as a marker of activated T cells, we studied the immunogenicity and cross-priming potential of LY6K-LP, assaying human immune cell responses in vitro and immunologic activities in HLA-A24 transgenic mice in vivo. We identified LY6K172–191-LP as an effective immunogen spanning naturally processed epitopes recognized by T helper type 1 (Th1) cells and CTLs. LY6K-specific CTLs were induced through cross-presentation of LY6K172–191-LP in vitro and in vivo. In addition, LY6K172–191-LP enhanced induction of LY6K-specific CTLs among the peripheral blood mononuclear cells (PBMCs) of head-and-neck malignant tumor (HNMT) patients. LY6K172–191-LP-specific Th1 immunologic response following 1 week in vitro stimulation of PBMCs with LY6K172–191-LP were detected in 16 of 21 HNMT patients (76%) vaccinated with CTL-epitope peptides and 1 of 11 HNMT patients (9%) prior to vaccination, but not in 9 healthy donors. Our results are the first to demonstrate the presence of LY6K-specific Th1 cell responses in HNMT patients and underscore the possible utility of LY6K172–191-LP for the induction and propagation of both LY6K-specific Th1 cells and CTLs.

CD169-positive sinus macrophages in the lymph nodes determine bladder cancer prognosis

CD169+ macrophages are suggested to play a pivotal role in establishing anti‐tumor immunity. They capture dead tumor cell‐associated antigens and transfer their information to lymphocsytes, including CD8+ T cells, which is important for successful tumor suppression. This study aimed to determine the prognostic significance of CD169+ macrophages residing in the tumor‐draining lymph nodes from cases of bladder cancer. In this retrospective study, 44 bladder cancer patients who received radical cystectomy were examined. The abundance of CD169+ macrophages in the regional lymph nodes and the number of CD8+ T cells in the primary tumor were investigated by immunohistochemistry. A CD169 score was calculated based on the intensity of CD169 staining and the proportion of CD169+ macrophages, and the scores were compared to the patients’ clinicopathological parameters. A high CD169 score was significantly associated with low T stage and with a high number of CD8+ T cells infiltrating into the tumor. The group with high CD169 expression had significantly longer cancer‐specific survival than the group with low CD169 expression (5‐year cancer‐specific survival rate: 83.3% vs 31.3%). In a multivariate analysis, the CD169 score was identified as a strong and independent favorable prognostic factor for cancer‐specific survival. Our findings suggest that CD169+ macrophages in the lymph nodes enhance anti‐tumor immunity by expanding CD8+ T cells in bladder cancer. The CD169 score may serve as a novel marker for the evaluation of bladder cancer prognosis.

Bladder cancer-associated cancer-testis antigen-derived long peptides encompassing both CTL and promiscuous HLA class II-restricted Th cell epitopes induced CD4+ T cells expressing converged T-cell receptor genes in vitro

ABSTRACT DEP domain containing 1 (DEPDC1) and M-phase phosphoprotein 1 (MPHOSPH1) are human cancer testis antigens that are frequently overexpressed in urinary bladder cancer. In a phase I/II clinical trial, a DEPDC1- and MPHOSPH1-derived short peptide vaccine demonstrated promising efficacy in preventing bladder cancer recurrence. Here, we aimed to identify long peptides (LPs) derived from DEPDC1 and MPHOSPH1 that induced both T-helper (Th) cells and tumor-reactive cytotoxic T lymphocytes (CTLs). Stimulation of peripheral blood mononuclear cells (PBMCs) from healthy donors with the synthetic DEPDC1- and MPHOSPH1-LPs predicted to bind to promiscuous human leukocyte antigen (HLA) class II molecules by a computer algorithm induced specific CD4+ T cells as revealed by interferon-γ enzyme-linked immunospot assays. Three of six LPs encompassed HLA-A2- or -A24-restricted CTL epitopes or both, and all six LPs stimulated DEPDC1- or MPHOSPH1-specific Th cells restricted by promiscuous and frequently observed HLA class II molecules in the Japanese population. Some LPs are naturally processed from the proteins in DCs, and the capacity of these LPs to cross-prime CTLs was confirmed in vivo using HLA-A2 or -A24 transgenic mice. The LP-specific and HLA class II-restricted T-cell responses were also observed in PBMCs from patients with bladder cancer. Repeated stimulation of PBMCs with DEPDC1-LPs and MPHOSPH1-LPs yielded clonal Th cells expressing specific T-cell receptor (TCR)-α and β genes. These DEPDC1- or MPHOSPH1-derived LPs may have applications in immunotherapy in patients with bladder cancer, and the TCR genes identified may be useful for monitoring of Th cells specific to LPs in vivo.

Advanced prostate cancer discovered with cancerous peritonitis: Case report

Cancerous peritonitis occurs rarely in patients with prostate cancer since prostate cancer is not likely to cause peritoneal dissemination because of the localization of prostate itself and the low frequency of metastasis to the intraperitoneal organs from prostate cancer. This rarity of cancerous peritonitis may delay the diagnosis and treatment of prostate cancer. Herein we report a case of a patient with abdominal distension due to cancerous peritonitis wherein the primary tumor in the intraperitoneal organs could not be detected, but prostate cancer was diagnosed by the presence of adenocarcinoma cells using ascites puncture cytology.