Date Published: April 26, 2017
Publisher: Public Library of Science
Author(s): 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, Aftab A. Ansari.
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.
Programmed death-1 (PD-1) is an immunoinhibitory receptor that is expressed on activated T cells and B cells and induces immune suppression by binding to PD-ligand 1 (PD-L1) . The PD-1/PD-L1 interaction normally works as a negative feedback system inhibiting excessive immune response, with PD-1 knockout mice developing autoimmune diseases such as lupus-like arthritis and glomerulonephritis . However, this mechanism is often exploited by various chronic diseases to avoid immune elimination. For instance, the expression of PD-L1 has been reported in many human cancers, including melanoma, squamous cell carcinoma, urothelial carcinoma, and solid tumors in the lung, ovary, kidney, pancreas, stomach, and colon [3–9]. In addition, PD-1 upregulation has been demonstrated in tumor-infiltrating T lymphocytes . Importantly, it has been shown that PD-L1 expression is strongly correlated with a progressive stage and worse prognosis, indicating a relationship with disease outcome [5–9].
In this study, we established Boch4G12, a chimeric antibody composed of the variable region from anti-bovine PD-L1 rat mAb (4G12) and the constant region from bovine IgG1. This antibody exhibited similar activities to the original antibody in terms of binding to bovine PD-L1 and interrupting the PD-1/PD-L1 interaction. The inhibition of T-cell function that is induced by PDL1/EGFP cells was restored using Boch4G12, indicating that this antibody has the potential to reactivate exhausted T cells through PD-L1 blockade. In addition, IFN-γ production was upregulated in PBMCs from BLV-infected cattle when cultured with Boch4G12. We previously demonstrated that PD-L1 on IgM+ B cells increases in line with the progression of BLV infection . Thus, this observed increase in IFN-γ in the culture supernatant was probably caused by Boch4G12 preventing an interaction between PD-1 on T cells and PD-L1 on infected B cells, reactivating the ability of T cells to respond to the BLV antigen. Finally, a BLV-infected calf that was inoculated with Boch4G12 showed a proliferation of CD4+ T cells in response to the BLV antigen and a reduction in BLV provirus loads, suggesting that Boch4G12 has therapeutic potential for controlling BLV infection.