Benralizumab有效抗嗜酸性粒细胞活性的新作用机制
2021/07/28
摘要
Benralizumab是一种人源化的抗IL-5Rα单克隆抗体，具有抗嗜酸性粒细胞活性。缺乏聚焦(afocusylation)增加了其对CD16a的亲和力，并显著增强了NK细胞介导的抗体依赖性细胞介导的细胞毒性(ADCC)。尽管benralizumab在严重哮喘和嗜酸性粒细胞增多综合征患者的临床试验中被证明是有效的，但其抗嗜酸性粒细胞增多机制的深入特征仍不清楚。在这里，我们进一步研究了benralizumab抗嗜酸性粒细胞活性的机制。在NK细胞存在的情况下，benralizumab通过上游诱导caspase 3/7和上调细胞色素c来诱导强效嗜酸性粒细胞凋亡。这个过程叫做抗体依赖性细胞吞噬(ADCP)。通过活细胞成像，我们揭示了导致嗜酸性粒细胞凋亡和被激活的巨噬细胞摄取的逐步过程。通过仔细观察细胞共培养试验，我们发现巨噬细胞来源的TNF通过激活嗜酸性粒细胞上的TNF受体1进一步增强benralizumab介导的嗜酸性粒细胞凋亡的新作用。TNF诱导的嗜酸性粒细胞凋亡与细胞色素C上调、线粒体膜去极化和caspase 3/7活性增加有关。此外，被激活的NK细胞通过分泌IFN-γ放大这条轴，随后驱动巨噬细胞表达TNF。我们的数据为benralizumab诱导的嗜酸性细胞凋亡的及时出现提供了深入的了解，并提示额外的机制可能有助于benralizumab在体内强效的抗嗜酸性细胞活性。重要的是，benralizumab的聚焦作用在所有研究机制中都强有力地增强了其效力。
Novel mechanisms of action contributing to Benralizumab's potent anti-eosinophilic activity
Rania Dagher, Varsha Kumar, Alan M Copenhaver, Sandra Gallagher, Mahboobe Ghaedi, Jonathan Boyd , Paul Newbold, Alison A Humbles, Roland Kolbeck
Abstract
Benralizumab is a humanised, anti-IL-5Rα monoclonal antibody with anti-eosinophilic activity. Lack of fucose (afucosylation) increases its affinity to CD16a and significantly enhances antibody-dependent cell-mediated cytotoxicity (ADCC) by NK cells. Although benralizumab proved clinically efficacious in clinical trials for patients with severe asthma and hypereosinophilic syndrome, in-depth characterisation of its anti-eosinophilic mechanisms of action remain elusive. Here, we further investigated the mechanisms involved in benralizumab's anti-eosinophilic activities. In the presence of NK cells benralizumab induced potent eosinophil apoptosis as demonstrated by the upstream induction of caspase 3/7 and upregulation of cytochrome C. In addition, we uncovered a previously unrecognised mechanism whereby benralizumab can induce eosinophil phagocytosis/efferocytosis by macrophages, a process called antibody-dependent cell phagocytosis (ADCP). Using live cell imaging we unravel the stepwise processes leading to eosinophil apoptosis and uptake by activated macrophages. Through careful observations of cellular co-culture assays we identified a novel role for macrophage derived TNF to further enhance benralizumab-mediated eosinophil apoptosis through activation of TNF-receptor 1 on eosinophils. TNF-induced eosinophil apoptosis was associated with Cytochrome C upregulation, mitochondrial membrane depolarisation, and increased caspase 3/7 activity. Moreover, activated NK cells were found to amplify this axis through the secretion of IFNγ, subsequently driving TNF expression by macrophages. Our data provide insights into the timely appearance of events leading to benralizumab-induced eosinophil apoptosis and suggest that additional mechanisms may contribute to the potent anti-eosinophilic activity of benralizumab in vivo Importantly, afucosylation of benralizumab strongly enhanced its potency for all mechanisms investigated.
Benralizumab是一种人源化的抗IL-5Rα单克隆抗体，具有抗嗜酸性粒细胞活性。缺乏聚焦(afocusylation)增加了其对CD16a的亲和力，并显著增强了NK细胞介导的抗体依赖性细胞介导的细胞毒性(ADCC)。尽管benralizumab在严重哮喘和嗜酸性粒细胞增多综合征患者的临床试验中被证明是有效的，但其抗嗜酸性粒细胞增多机制的深入特征仍不清楚。在这里，我们进一步研究了benralizumab抗嗜酸性粒细胞活性的机制。在NK细胞存在的情况下，benralizumab通过上游诱导caspase 3/7和上调细胞色素c来诱导强效嗜酸性粒细胞凋亡。这个过程叫做抗体依赖性细胞吞噬(ADCP)。通过活细胞成像，我们揭示了导致嗜酸性粒细胞凋亡和被激活的巨噬细胞摄取的逐步过程。通过仔细观察细胞共培养试验，我们发现巨噬细胞来源的TNF通过激活嗜酸性粒细胞上的TNF受体1进一步增强benralizumab介导的嗜酸性粒细胞凋亡的新作用。TNF诱导的嗜酸性粒细胞凋亡与细胞色素C上调、线粒体膜去极化和caspase 3/7活性增加有关。此外，被激活的NK细胞通过分泌IFN-γ放大这条轴，随后驱动巨噬细胞表达TNF。我们的数据为benralizumab诱导的嗜酸性细胞凋亡的及时出现提供了深入的了解，并提示额外的机制可能有助于benralizumab在体内强效的抗嗜酸性细胞活性。重要的是，benralizumab的聚焦作用在所有研究机制中都强有力地增强了其效力。
(中日友好医院呼吸与危重症医学科 张清 摘译 林江涛 审校)
(Eur Respir J. 2021 Jul 21;2004306. doi: 10.1183/13993003.04306-2020)
(Eur Respir J. 2021 Jul 21;2004306. doi: 10.1183/13993003.04306-2020)
Novel mechanisms of action contributing to Benralizumab's potent anti-eosinophilic activity
Rania Dagher, Varsha Kumar, Alan M Copenhaver, Sandra Gallagher, Mahboobe Ghaedi, Jonathan Boyd , Paul Newbold, Alison A Humbles, Roland Kolbeck
Abstract
Benralizumab is a humanised, anti-IL-5Rα monoclonal antibody with anti-eosinophilic activity. Lack of fucose (afucosylation) increases its affinity to CD16a and significantly enhances antibody-dependent cell-mediated cytotoxicity (ADCC) by NK cells. Although benralizumab proved clinically efficacious in clinical trials for patients with severe asthma and hypereosinophilic syndrome, in-depth characterisation of its anti-eosinophilic mechanisms of action remain elusive. Here, we further investigated the mechanisms involved in benralizumab's anti-eosinophilic activities. In the presence of NK cells benralizumab induced potent eosinophil apoptosis as demonstrated by the upstream induction of caspase 3/7 and upregulation of cytochrome C. In addition, we uncovered a previously unrecognised mechanism whereby benralizumab can induce eosinophil phagocytosis/efferocytosis by macrophages, a process called antibody-dependent cell phagocytosis (ADCP). Using live cell imaging we unravel the stepwise processes leading to eosinophil apoptosis and uptake by activated macrophages. Through careful observations of cellular co-culture assays we identified a novel role for macrophage derived TNF to further enhance benralizumab-mediated eosinophil apoptosis through activation of TNF-receptor 1 on eosinophils. TNF-induced eosinophil apoptosis was associated with Cytochrome C upregulation, mitochondrial membrane depolarisation, and increased caspase 3/7 activity. Moreover, activated NK cells were found to amplify this axis through the secretion of IFNγ, subsequently driving TNF expression by macrophages. Our data provide insights into the timely appearance of events leading to benralizumab-induced eosinophil apoptosis and suggest that additional mechanisms may contribute to the potent anti-eosinophilic activity of benralizumab in vivo Importantly, afucosylation of benralizumab strongly enhanced its potency for all mechanisms investigated.
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