Anti-Human CCR4 Therapeutic Antibody (Mogamulizumab)

Figure 1 Mogamulizumab eliminates CCR4 + cells in both CD4 + and CD8 + T cells.

A, Representative dot plots of fluorescence-activated cell-sorter analysis of CCR4 and CD4 expression in CD3 + T cells among peripheral blood mononuclear cells from patients with human T-lymphotropic virus type 1–asso- ciated myelopathy/tropical spastic paraparesis after 5-day culture in the presence or absence of 1 µg/mL of mogamulizumab. B, Percentages of CCR4 + cells in CD3 + CD4 + T cells were compared between the untreated and mogamulizumab groups (n = 11). Statistical analysis was performed using the Wilcoxon signed-rank test. C, The population enclosed in the box in panel A (the CD3 + CD4 − CCR4 + subset) was gated and analyzed for the expression of CD8. The percentage of CD8 + cells is shown. D and E, CCR4 and CD8 expression in CD3 + T cells was analyzed as described above.

Yamauchi, J., Coler-Reilly, A., Sato, T., Araya, N., Yagishita, N., Ando, H., ... & Nishioka, K. (2014). Mogamulizumab, an Anti-CCR4 Antibody, Targets Human T-Lymphotropic Virus Type 1–infected CD8+ and CD4+ T Cells to Treat Associated Myelopathy. The Journal of infectious diseases, 211(2), 238-248.

Figure 2 Mogamulizumab and KM2760 inhibit cytokine production in peripheral blood mononuclear cells (PBMCs) from patients with human T-lymphotropic virus type 1–associated myelopathy/tropical spastic paraparesis (HAM/TSP).

PBMCs from 11 patients with HAM/TSP were cultured for 7 days without stimuli and without treatment or in the presence of mogamulizumab, KM2760, or prednisolone (PSL). The concentrations of cytokines (interferon γ [IFN-γ], interleukin 6 [IL-6], interleukin 2 [IL-2], tumor necrosis factor α [TNF-α], and interleukin 10 [IL-10]) in the supernatants were then measured. A, Direct comparison of the concentrations of these cytokines in the supernatants of untreated PBMC cultures. Horizontal bars represent the median values. B–F, The effects of the treatments on the concentrations of these cytokines. Data are presented as the mean ± SD. Statistical analyses were performed using the Friedman test followed by the Dunn test for comparison with PBMCs alone. *P < .05, **P < .01, and ***P < .001. Abbreviation: SD, standard deviation.

Yamauchi, J., Coler-Reilly, A., Sato, T., Araya, N., Yagishita, N., Ando, H., ... & Nishioka, K. (2014). Mogamulizumab, an Anti-CCR4 Antibody, Targets Human T-Lymphotropic Virus Type 1–infected CD8+ and CD4+ T Cells to Treat Associated Myelopathy. The Journal of infectious diseases, 211(2), 238-248.

Figure 3 Mogamulizumab-induced ADCC. A, four CCR4-positive and three CCR4-negative cell lines were examined.

Cytotoxicity was measured using the LDH assay in the presence of effector cells obtained from healthy volunteers and mogamulizumab (10 μg/mL) or the same volume of solvent (control). B, dose-dependent effects of mogamulizumab against SNK6. Various doses of mogamulizumab and PBMCs from three healthy individuals were prepared. The ratio of target:effector was fixed at 1:50. C, identification of the fraction responsible for ADCC against SNK6. Separated CD56-, CD3-, and CD19-positive cells were used as effector cells, respectively. The ratio of target:effector was fixed at 1:10. The mogamulizumab dose was fixed at 10 μg/mL. Error bars, SEM.

Kanazawa, T., Hiramatsu, Y., Iwata, S., Siddiquey, M., Sato, Y., Suzuki, M., ... & Kimura, H. (2014). Anti-CCR4 monoclonal antibody mogamulizumab for the treatment of EBV-associated T-and NK-cell lymphoproliferative diseases. Clinical Cancer Research.

Figure 4 Mogamulizumab induced potent antitumor activity against EBV-positive NK-cell lymphoma in the murine xenograft model.

A, NOG mice were implanted subcutaneously with SNK6 cells on day 0. From day 4, human PBMCs with mogamulizumab or saline (control) were injected intraperitoneally (6 mice per group). *, P < 0.05 (Mann–Whitney U test). Error bars, SEM. B, tumor-bearing mice at day 28. Tumors are outlined by the red dotted line. C, H&E-stained section showing tumor infiltration into the subcutaneous lesion wall. D, EBER in situ hybridization for subcutaneous tumor. Scale bars, 300 μm.

Kanazawa, T., Hiramatsu, Y., Iwata, S., Siddiquey, M., Sato, Y., Suzuki, M., ... & Kimura, H. (2014). Anti-CCR4 monoclonal antibody mogamulizumab for the treatment of EBV-associated T-and NK-cell lymphoproliferative diseases. Clinical Cancer Research.

Figure 5 Immunohistochemistry of CCR4 in patients with PTCL.

CCR4 expression level in patients with PTCL varies in each case. CCR4 expression level is immunohistochemically evaluated in three grades. 1+: 10–24% of tumor cells are positive for CCR4 (a, ×400), 2+: 25–49% of tumor cells are positive for CCR4 (b, ×400), and 3+: 50% or more tumor cells are positive (c, ×400).Abbreviations: CCR4 CC chemokine receptor 4; PTCL peripheral T-cell lymphoma.

Makita, S., & Tobinai, K. (2017). Mogamulizumab for the treatment of T-cell lymphoma. Expert opinion on biological therapy, 17(9), 1145-1153.

Figure 6 The pan-histone deacetylase inhibitor, vorinostat reduces CCR4 expression in various T-cell and natural killer- cell lymphoma cell lines.

Bars indicate the mean ± standard error of the mean (SEM) of three independent experiments. Asterisks (*) indicate statistical significance: *0.01 ≤ P < 0.05, **0.001 ≤ P < 0.01, ***P < 0.001, n.s: not siginificant. Quantitative RT-PCR analysis of CCR4 in vorinostat-treated T- and NK-cell lymphoma cell lines (5 μM for 24 h). The Student ttest was used to examine statistical significance. x-axis: cell lines. y-axis: 2-ΔCt values for microRNA expression.

Kitadate, A., Ikeda, S., Abe, F., Takahashi, N., Shimizu, N., Matsue, K., & Tagawa, H. (2018). Histone deacetylase inhibitors downregulate CCR4 expression and decrease mogamulizumab efficacy in CCR4-positive mature T-cell lymphomas. haematologica, 103(1), 126-135.

Figure 7 The pan-histone deacetylase inhibitor, vorinostat reduces CCR4 expression in various T-cell and natural killer- cell lymphoma cell lines.

Migration assay of lymphoma cells treated with 5 μM vorinostat for 24 h. A schematic illustration of the migration assay is also shown. RFU: relative fluorescence units.

Kitadate, A., Ikeda, S., Abe, F., Takahashi, N., Shimizu, N., Matsue, K., & Tagawa, H. (2018). Histone deacetylase inhibitors downregulate CCR4 expression and decrease mogamulizumab efficacy in CCR4-positive mature T-cell lymphomas. haematologica, 103(1), 126-135.