研究者データベース

前川 直也(マエカワ ナオヤ)
獣医学研究院
特任助教

基本情報

所属

  • 獣医学研究院

職名

  • 特任助教

学位

  • 博士(獣医学)(北海道大学)

J-Global ID

研究キーワード

  • 獣医免疫学   

研究分野

  • ライフサイエンス / 獣医学
  • ライフサイエンス / 獣医学
  • ライフサイエンス / 獣医学

職歴

  • 2018年07月 - 現在 北海道大学 大学院獣医学研究院 特任助教
  • 2018年01月 - 2018年06月 北海道大学 大学院獣医学研究院 博士研究員
  • 2015年04月 - 2017年12月 日本学術振興会 特別研究員(DC1)

学歴

  • 2014年04月 - 2017年12月   北海道大学   大学院獣医学研究科
  • 2008年04月 - 2014年03月   北海道大学   獣医学部

所属学協会

  • 日本獣医学会   

研究活動情報

論文

  • Kato Y, Ohishi T, Kawada M, Maekawa N, Konnai S, Itai S, Yamada S, Kaneko MK
    Biochemistry and biophysics reports 17 23 - 26 2019年03月 [査読有り][通常論文]
  • Tomohiro Okagawa, Satoru Konnai, Asami Nishimori, Naoya Maekawa, Shinya Goto, Ryoyo Ikebuchi, Junko Kohara, Yasuhiko Suzuki, Shinji Yamada, Yukinari Kato, Shiro Murata, Kazuhiko Ohashi
    Veterinary research 49 1 50 - 50 2018年06月19日 [査読有り][通常論文]
     
    Bovine leukemia virus (BLV) is a retrovirus that infects B cells in cattle and causes bovine leukosis after a long latent period. Progressive exhaustion of T cell functions is considered to facilitate disease progression of BLV infection. Programmed death-1 (PD-1) and lymphocyte activation gene-3 (LAG-3) are immunoinhibitory receptors that contribute to T-cell exhaustion caused by BLV infection in cattle. However, it is unclear whether the cooperation of PD-1 and LAG-3 accelerates disease progression of BLV infection. In this study, multi-color flow cytometric analyses of PD-1- and LAG-3-expressing T cells were performed in BLV-infected cattle at different stages of the disease. The frequencies of PD-1+LAG-3+ heavily exhausted T cells among CD4+ and CD8+ T cells was higher in the blood of cattle with B-cell lymphoma over that of BLV-uninfected and BLV-infected cattle without lymphoma. In addition, blockade assays of peripheral blood mononuclear cells were performed to examine whether inhibition of the interactions between PD-1 and LAG-3 and their ligands by blocking antibodies could restore T-cell function during BLV infection. Single or dual blockade of the PD-1 and LAG-3 pathways reactivated the production of Th1 cytokines, interferon-γ and tumor necrosis factor-α, from BLV-specific T cells of the infected cattle. Taken together, these results indicate that PD-1 and LAG-3 cooperatively mediate the functional exhaustion of CD4+ and CD8+ T cells and are associated with the development of B-cell lymphoma in BLV-infected cattle.
  • Yamato Sajiki, Satoru Konnai, Tomohiro Okagawa, Asami Nishimori, Naoya Maekawa, Shinya Goto, Ryoyo Ikebuchi, Reiko Nagata, Satoko Kawaji, Yumiko Kagawa, Shinji Yamada, Yukinari Kato, Chie Nakajima, Yasuhiko Suzuki, Shiro Murata, Yasuyuki Mori, Kazuhiko Ohashi
    Infection and immunity 86 5 2018年05月 [査読有り][通常論文]
     
    Johne's disease, caused by Mycobacterium avium subsp. paratuberculosis, is a bovine chronic infection that is endemic in Japan and many other countries. The expression of immunoinhibitory molecules is upregulated in cattle with Johne's disease, but the mechanism of immunosuppression is poorly understood. Prostaglandin E2 (PGE2) is immunosuppressive in humans, but few veterinary data are available. In this study, functional and kinetic analyses of PGE2 were performed to investigate the immunosuppressive effect of PGE2 during Johne's disease. In vitro PGE2 treatment decreased T-cell proliferation and Th1 cytokine production and upregulated the expression of immunoinhibitory molecules such as interleukin-10 and programmed death ligand 1 (PD-L1) in peripheral blood mononuclear cells (PBMCs) from healthy cattle. PGE2 was upregulated in sera and intestinal lesions of cattle with Johne's disease. In vitro stimulation with Johnin purified protein derivative (J-PPD) induced cyclooxygenase-2 (COX-2) transcription, PGE2 production, and upregulation of PD-L1 and immunoinhibitory receptors in PBMCs from cattle infected with M. avium subsp. paratuberculosis Therefore, Johnin-specific Th1 responses could be limited by the PGE2 pathway in cattle. In contrast, downregulation of PGE2 with a COX-2 inhibitor promoted J-PPD-stimulated CD8+ T-cell proliferation and Th1 cytokine production in PBMCs from the experimentally infected cattle. PD-L1 blockade induced J-PPD-stimulated CD8+ T-cell proliferation and interferon gamma production in vitro Combined treatment with a COX-2 inhibitor and anti-PD-L1 antibodies enhanced J-PPD-stimulated CD8+ T-cell proliferation in vitro, suggesting that the blockade of both pathways is a potential therapeutic strategy to control Johne's disease. The effects of COX-2 inhibition warrant further study as a novel treatment of Johne's disease.
  • Maekawa N, Konnai S, Balbin MM, Mingala CN, Gicana KRB, Bernando FAEM, Murata S, Ohashi K
    Ticks and tick-borne diseases 9 2 266 - 269 2018年02月 [査読有り][通常論文]
  • Michihito Tagawa, Chihiro Kurashima, Satoshi Takagi, Naoya Maekawa, Satoru Konnai, Genya Shimbo, Kotaro Matsumoto, Hisashi Inokuma, Keiko Kawamoto, Kazuro Miyahara
    PloS one 13 7 e0201222  2018年 [査読有り][通常論文]
     
    B cell high grade lymphoma is the most common hematopoietic malignancy in dogs. Although the immune checkpoint molecules, programmed death-1 (PD-1) and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4), and immune checkpoint inhibitors have been evaluated for the treatment of various human lymphoid malignancies, the expression of those molecules and their relationship with prognosis remain unknown in canine lymphoma. The objective of this study was to evaluate the expression of costimulatory molecules on peripheral blood lymphocytes and tumor infiltrating lymphocytes, in addition to associated ligand expression in the lymph nodes of patients with B cell multicentric high grade lymphoma. Eighteen patients diagnosed with B cell high grade lymphoma and nine healthy control dogs were enrolled. Flow cytometric analysis revealed that the expression of PD-1 on CD4+ peripheral and tumor infiltrating lymphocytes and CTLA-4 on CD4+ peripheral lymphocytes was significantly higher in the lymphoma group than in the control group. The expression level of CD80 mRNA was significantly lower in the lymphoma group than in the control group. In contrast, there were no significant differences in PD-L1, PD-L2, and CD86 expression between the groups. Dogs with CTLA-4 levels below the cutoff values, which were determined based on receiver operating characteristic curves, on peripheral CD4+, CD8+, and tumor infiltrating CD4+ lymphocytes had significantly longer survival than dogs with values above the cutoff. Although it is uncertain whether the expression of immune checkpoint molecules affect the biological behavior of canine lymphoma, one possible explanation is that PD-1 and CTLA-4 might be associated with the suppression of antitumor immunity in dogs with B cell high grade lymphoma, particularly through CD4+ T cells.
  • Yamato Sajiki, Satoru Konnai, Asami Nishimori, Tomohiro Okagawa, Naoya Maekawa, Shinya Goto, Masashi Nagano, Junko Kohara, Nana Kitano, Toshihiko Takahashi, Motoshi Tajima, Hirohisa Mekata, Yoichiro Horii, Shiro Murata, Kazuhiko Ohashi
    The Journal of veterinary medical science 79 12 2036 - 2039 2017年12月22日 [査読有り][通常論文]
     
    Enzootic bovine leukemia is caused by the bovine leukemia virus (BLV). BLV is transmitted vertically or horizontally through the transfer of infected cells via direct contact, through milk, insect bites and contaminated iatrogenic procedures. However, we lacked direct evidence of intrauterine infection. The purpose of this study was to confirm intrauterine BLV infection in two pregnant dams with high viral load by cesarean delivery. BLV was detected in cord and placental blood, and the BLV in the newborns showed 100% nucleotide identity with the BLV-env sequence from the dams. Notably, a newborn was seropositive for BLV but had no colostral antibodies. In this study, we presented a direct evidence of intrauterine BLV transmission in pregnant dam with a high proviral load. These results could aid the development of BLV control measures targeting viral load.
  • Goto S, Konnai S, Okagawa T, Nishimori A, Maekawa N, Gondaira S, Higuchi H, Koiwa M, Tajima M, Kohara J, Ogasawara S, Kato Y, Suzuki Y, Murata S, Ohashi K
    Immunity, inflammation and disease 5 3 355 - 363 2017年09月 [査読有り][通常論文]
     
    INTRODUCTION: Bovine mycoplasma, chiefly Mycoplasma bovis, is a pathogen that causes pneumonia, mastitis, arthritis, and otitis media in cattle. This pathogen exerts immunosuppressive effects, such as the inhibition of interferon production. However, the mechanisms involved in bovine mycoplasmosis have not been fully elucidated. In this study, we investigated the role of the programmed death-1 (PD-1)/programmed death-ligand 1 (PD-L1) pathway in immunosuppression in bovine mycoplasmosis. METHODS: In the initial experiments, we used enzyme-linked immunosorbent assay to measure interferon-γ (IFN-γ) from peripheral blood mononuclear cells (PBMCs) isolated from cattle with mycoplasmosis. RESULTS: Expectedly, IFN-γ production significantly decreased in cattle with mycoplasmosis compared with that in clinically healthy cattle. Concomitantly, flow cytometric analysis revealed that the proportions of PD-1+ CD4+ and PD-L1+ CD14+ cells significantly increased in peripheral blood of the infected cattle. Interestingly, the number of PD-1+ CD4+ and PD-1+ CD8+ T cells were negatively correlated with IFN-γ production from PBMCs in bovine mycoplasmosis. Additionally, blockade of the PD-1/PD-L1 pathway in vitro by anti-bovine PD-1- and anti-bovine PD-L1 antibodies significantly upregulated the production of IFN-γ from anti-mycoplasma-specific cells. CONCLUSIONS: These results suggest that the PD-1/PD-L1 pathway could be involved in immune exhaustion of bovine mycoplasma-specific T cells. In conclusion, our study opens up a new perspective in the therapeutic strategy for bovine mycoplasmosis by targeting the immunoinhibitory receptor pathways.
  • Asami Nishimori, Satoru Konnai, Tomohiro Okagawa, Naoya Maekawa, Shinya Goto, Ryoyo Ikebuchi, Ayako Nakahara, Yuzumi Chiba, Masaho Ikeda, Shiro Murata, Kazuhiko Ohashi
    CLINICAL AND VACCINE IMMUNOLOGY 24 9 2017年09月 [査読有り][通常論文]
     
    Bovine leukemia is classified into two types: enzootic bovine leukosis (EBL) and sporadic bovine leukosis (SBL). EBL is caused by infection with bovine leukemia virus (BLV), which induces persistent lymphocytosis and B-cell lymphoma in cattle after a long latent period. Although it has been demonstrated that BLVassociated lymphoma occurs predominantly in adult cattle of > 3 to 5 years, suspicious cases of EBL onset in juvenile cattle were recently reported in Japan. To investigate the current status of bovine leukemia in Japan, we performed immunophenotypic analysis of samples from 50 cattle that were clinically diagnosed as having bovine leukemia. We classified the samples into five groups on the basis of the analysis and found two different types of EBL: classic EBL (cEBL), which has the familiar phenotype commonly known as EBL, and polyclonal EBL (pEBL), which exhibited neoplastic proliferation of polyclonal B cells. Moreover, there were several atypical EBL cases even in cEBL, including an early onset of EBL in juvenile cattle. A comparison of the cell marker expressions among cEBL, pEBL, and B-cell-type SBL (B-SBL) revealed characteristic patterns in B-cell leukemia, and these patterns could be clearly differentiated from those of healthy phenotypes, whereas it was difficult to discriminate between cEBL, pEBL, and B-SBL only by the expression patterns of cell markers. This study identified novel characteristics of bovine leukemia that should contribute to a better understanding of the mechanism underlying tumor development in BLV infection.
  • Naoya Maekawa, Satoru Konnai, Satoshi Takagi, Yumiko Kagawa, Tomohiro Okagawa, Asami Nishimori, Ryoyo Ikebuchi, Yusuke Izumi, Tatsuya Deguchi, Chie Nakajima, Yukinari Kato, Keiichi Yamamoto, Hidetoshi Uemura, Yasuhiko Suzuki, Shiro Murata, Kazuhiko Ohashi
    SCIENTIFIC REPORTS 7 1 8951  2017年08月 [査読有り][通常論文]
     
    Immunotherapy targeting immune checkpoint molecules, programmed cell death 1 (PD-1) and PD-ligand 1 (PD-L1), using therapeutic antibodies has been widely used for some human malignancies in the last 5 years. A costimulatory receptor, PD-1, is expressed on T cells and suppresses effector functions when it binds to its ligand, PD-L1. Aberrant PD-L1 expression is reported in various human cancers and is considered an immune escape mechanism. Antibodies blocking the PD-1/PD-L1 axis induce antitumour responses in patients with malignant melanoma and other cancers. In dogs, no such clinical studies have been performed to date because of the lack of therapeutic antibodies that can be used in dogs. In this study, the immunomodulatory effects of c4G12, a canine-chimerised anti-PD-L1 monoclonal antibody, were evaluated in vitro, demonstrating significantly enhanced cytokine production and proliferation of dog peripheral blood mononuclear cells. A pilot clinical study was performed on seven dogs with oral malignant melanoma (OMM) and two with undifferentiated sarcoma. Objective antitumour responses were observed in one dog with OMM (14.3%, 1/7) and one with undifferentiated sarcoma (50.0%, 1/2) when c4G12 was given at 2 or 5 mg/kg, every 2 weeks. c4G12 could be a safe and effective treatment option for canine cancers.
  • Tomohiro Okagawa, Satoru Konnai, Asami Nishimori, Naoya Maekawa, Ryoyo Ikebuchi, Shinya Goto, Chie Nakajima, Junko Kohara, Satoshi Ogasawara, Yukinari Kato, Yasuhiko Suzuki, Shiro Murata, Kazuhiko Ohashi
    FRONTIERS IN IMMUNOLOGY 8 650  2017年06月 [査読有り][通常論文]
     
    Blockade of immunoinhibitory molecules, such as programmed death-1 (PD-1)/PD-ligand 1 (PD-L1), is a promising strategy for reinvigorating exhausted T cells and preventing disease progression in a variety of chronic infections. Application of this therapeutic strategy to cattle requires bovinized chimeric antibody targeting immunoinhibitory molecules. In this study, anti-bovine PD-1 rat-bovine chimeric monoclonal antibody 5D2 (Boch5D2) was constructed with mammalian expression systems, and its biochemical function and antiviral effect were characterized in vitro and in vivo using cattle infected with bovine leukemia virus (BLV). Purified Boch5D2 was capable of detecting bovine PD-1 molecules expressed on cell membranes in flow cytometric analysis. In particular, Biacore analysis determined that the binding affinity of Boch5D2 to bovine PD-1 protein was similar to that of the original anti-bovine PD-1 rat monoclonal antibody 5D2. Boch5D2 was also capable of blocking PD-1/PD-L1 binding at the same level as 5D2. The immunomodulatory and therapeutic effects of Boch5D2 were evaluated by in vivo administration of the antibody to a BLV-infected calf. Inoculated Boch5D2 was sustained in the serum for a longer period. Boch5D2 inoculation resulted in activation of the proliferation of BLV-specific CD4(+) T cells and decrease in the proviral load of BLV in the peripheral blood. This study demonstrates that Boch5D2 retains an equivalent biochemical function to that of the original antibody 5D2 and is a candidate therapeutic agent for regulating antiviral immune response in vivo. Clinical efficacy of PD-1/PD-L1 blockade awaits further experimentation with a large number of animals.
  • Asami Nishimori, Satoru Konnai, Tomohiro Okagawa, Naoya Maekawa, Ryoyo Ikebuchi, Shinya Goto, Yamato Sajiki, Yasuhiko Suzuki, Junko Kohara, Satoshi Ogasawara, Yukinari Kato, Shiro Murata, Kazuhiko Ohashi
    PLOS ONE 12 4 e0174916  2017年04月 [査読有り][通常論文]
     
    Programmed death-1 (PD-1), an immunoinhibitory receptor on T cells, is known to be involved in immune evasion through its binding to PD-ligand 1 (PD-L1) in many chronic diseases. We previously found that PD-L1 expression was upregulated in cattle infected with bovine leukemia virus (BLV) and that an antibody that blocked the PD-1/PD-L1 interaction reactivated T-cell function in vitro. Therefore, this study assessed its antivirus activities in vivo. First, we inoculated the anti-bovine PD-L1 rat monoclonal antibody 4G12 into a BLV-infected cow. However, this did not induce T-cell proliferation or reduction of BLV provirus loads during the test period, and only bound to circulating IgM(+) B cells until one week post-inoculation. We hypothesized that this lack of in vivo effects was due to its lower stability in cattle and so established an anti-PD-L1 rat-bovine chimeric antibody (Boch4G12). Boch4G12 was able to bind specifically with bovine PD-L1, interrupt the PD-1/PD-L1 interaction, and activate the immune response in both healthy and BLV-infected cattle in vitro. Therefore, we experimentally infected a healthy calf with BLV and inoculated it intravenously with 1 mg/kg of Boch4G12 once it reached the aleukemic (AL) stage. Cultivation of peripheral blood mononuclear cells (PBMCs) isolated from the tested calf indicated that the proliferation of CD4(+) T cells was increased by Boch4G12 inoculation, while BLV provirus loads were significantly reduced, clearly demonstrating that this treatment induced antivirus activities. Therefore, further studies using a large number of animals are required to support its efficacy for clinical application.
  • Naoya Maekawa, Satoru Konnai, Tomohiro Okagawa, Asami Nishimori, Ryoyo Ikebuchi, Yusuke Izumi, Satoshi Takagi, Yumiko Kagawa, Chie Nakajima, Yasuhiko Suzuki, Yukinari Kato, Shiro Murata, Kazuhiko Ohashi
    PLOS ONE 11 6 e0157176  2016年06月 [査読有り][通常論文]
     
    Spontaneous cancers are common diseases in dogs. Among these, some malignant cancers such as oral melanoma, osteosarcoma, hemangiosarcoma, and mast cell tumor are often recognized as clinical problems because, despite their high frequencies, current treatments for these cancers may not always achieve satisfying outcomes. The absence of effective systemic therapies against these cancers leads researchers to investigate novel therapeutic modalities, including immunotherapy. Programmed death 1 (PD-1) is a costimulatory receptor with immunosuppressive function. When it binds its ligands, PD-ligand 1 (PD-L1) or PD-L2, PD-1 on T cells negatively regulates activating signals from the T cell receptor, resulting in the inhibition of the effector function of cytotoxic T lymphocytes. Aberrant PD-L1 expression has been reported in many human cancers and is considered an immune escape mechanism for cancers. In clinical trials, anti-PD-1 or anti-PD-L1 antibodies induced tumor regression for several malignancies, including advanced melanoma, non-small cell lung carcinoma, and renal cell carcinoma. In this study, to assess the potential of the PD-1/PD-L1 axis as a novel therapeutic target for canine cancer immunotherapy, immunohistochemical analysis of PD-L1 expression in various malignant cancers of dogs was performed. Here, we show that dog oral melanoma, osteosarcoma, hemangiosarcoma, mast cell tumor, mammary adenocarcinoma, and prostate adenocarcinoma expressed PD-L1, whereas some other types of cancer did not. In addition, PD-1 was highly expressed on tumor-infiltrating lymphocytes obtained from oral melanoma, showing that lymphocytes in this cancer type might have been functionally exhausted. These results strongly encourage the clinical application of PD-1/PD-L1 inhibitors as novel therapeutic agents against these cancers in dogs.
  • Ohira K, Nakahara A, Konnai S, Okagawa T, Nishimori A, Maekawa N, Ikebuchi R, Kohara J, Murata S, Ohashi K
    Immunity, inflammation and disease 4 1 52 - 63 2016年03月 [査読有り][通常論文]
  • Michihito Tagawa, Naoya Maekawa, Satoru Konnai, Satoshi Takagi
    PLOS ONE 11 2 e0150030  2016年02月 [査読有り][通常論文]
     
    Histiocytic sarcoma is a rapidly progressive and fatal neoplastic disease in dogs. It is unclear whether costimulatory molecules, including CD28, cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4), and programmed death-1 (PD-1), are expressed on peripheral blood lymphocytes (PBLs) of canine patients with histiocytic sarcoma. The objective of this study was to evaluate the expression of CD28, CTLA-4, and PD-1 molecules on PBLs of patients with histiocytic sarcoma, patients with other tumors, and healthy controls. Twenty-six dogs were included in the study, with eight, ten, and eight dogs in the histiocytic sarcoma, other tumor, and healthy control groups, respectively. PBLs and serum were prospectively obtained from patients diagnosed histopathologically with histiocytic sarcoma, other tumors and healthy controls. The surface expression of CTLA-4, CD28, and PD-1 on T lymphocytes was examined using flow cytometric analysis. Serum samples were frozen at -30 degrees C until serum interferon-gamma (IFN-gamma) was measured by enzyme-linked immunosorbent assay. The expression level of CTLA-4 on CD4+ lymphocytes was significantly higher in the histiocytic sarcoma group than in the control group. The expression of CTLA-4 on CD8+ lymphocytes was significantly higher in the histiocytic sarcoma group than in the other two groups. In addition, the expression of PD-1 on CD8+ lymphocytes was significantly higher in the histiocytic sarcoma group than in the control group. However, no significant differences in CD28 expressions and serum IFN-gamma levels were observed. The present results provided evidence showing that the expression levels of CTLA-4 on both CD4+ and CD8+ lymphocytes and PD-1 on CD8+ lymphocytes in peripheral blood obtained from dogs with histiocytic sarcoma were upregulated. The overexpressions of CTLA 4 and PD-1 suggested that antitumor immunity may be suppressed in dogs with histiocytic sarcoma.
  • Maekawa N, Konnai S, Ikebuchi R, Okagawa T, Adachi M, Takagi S, Kagawa Y, Nakajima C, Suzuki Y, Murata S, Ohashi K
    PloS one 9 6 e98415  2014年06月 [査読有り][通常論文]
     
    Programmed death 1 (PD-1), an immunoinhibitory receptor, and programmed death ligand 1 (PD-L1), its ligand, together induce the "exhausted'' status in antigen-specific lymphocytes and are thus involved in the immune evasion of tumor cells. In this study, canine PD-1 and PD-L1 were molecularly characterized, and their potential as therapeutic targets for canine tumors was discussed. The canine PD-1 and PD-L1 genes were conserved among canine breeds. Based on the sequence information obtained, the recombinant canine PD-1 and PD-L1 proteins were constructed; they were confirmed to bind each other. Antibovine PD-L1 monoclonal antibody effectively blocked the binding of recombinant PD-1 with PD-L1-expressing cells in a dose-dependent manner. Canine melanoma, mastocytoma, renal cell carcinoma, and other types of tumors examined expressed PD-L1, whereas some did not. Interestingly, anti-PD-L1 antibody treatment enhanced IFN-gamma production from tumor-infiltrating cells. These results showed that the canine PD-1/PD-L1 pathway is also associated with T-cell exhaustion in canine tumors and that its blockade with antibody could be a new therapeutic strategy for canine tumors. Further investigations are needed to confirm the ability of anti-PD-L1 antibody to reactivate canine antitumor immunity in vivo, and its therapeutic potential has to be further discussed.

受賞

  • 2018年09月 日本獣医学会 獣医学奨励賞
     
    受賞者: 前川 直也
  • 2016年08月 The 11th International Veterinary Immunology Symposium IUIS VIC/EVIG Travel Award
     
    受賞者: 前川 直也
  • 2014年03月 北海道獣医師会 北海道獣医師会長賞
     
    受賞者: 前川 直也

共同研究・競争的資金等の研究課題

  • 伴侶動物の腫瘍における免疫抑制因子を標的とした新規免疫療法の開発
    日本学術振興会:特別研究員奨励費
    研究期間 : 2015年04月 -2017年12月 
    代表者 : 前川 直也


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