477一种针对人单核细胞内生性PD-L1的新型抗体促进了单核细胞抗肿瘤免疫的免疫刺激功能

Michelle Hsu, Xin Liu, Ying Li, Jacob Hirdler, Fabrice Lucien-Matteoni, Haidong Dong
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引用次数: 0

摘要

目前已经开发出PD-L1靶向抗体来阻断PD-L1与PD-1的相互作用,从而阻止T细胞毒性的抑制。然而,尽管PD-L1高表达,但在治疗癌症方面的临床成功有限。最近的报道表明,肿瘤固有的PD-L1可以在细胞内发出信号,促进细胞存活,而不依赖于PD-1的连接,这可能解释了为什么一些癌症患者对免疫检查点治疗没有反应。除肿瘤细胞外,宿主髓细胞也是PD-L1的来源,具有高度的免疫抑制作用。不幸的是,PD-L1在髓细胞中的内在功能尚未得到很好的研究。我们的目标是剖析单核细胞(骨髓细胞的一个子集)中PD-L1的内在信号传导,并研究这可能如何损害抗肿瘤免疫。方法本实验室鉴定了一种新的PD-L1抗体(克隆H1A),该抗体能在细胞表面破坏PD-L1的稳定性并诱导其降解。在我们的实验中,我们使用了从健康供体血液中分离的人类PBMCs和使用负磁选择从PBMCs中分离的单核细胞。为了研究h1a诱导的PD-L1降解对人单核细胞的影响,我们分别通过流式细胞术、免疫测定和单细胞RNA测序来评估单核细胞的表型、功能和转录谱。为了研究H1A对T细胞功能状态的间接影响,我们通过流式细胞术和T细胞聚焦板的质量细胞术评估了pbmc。为了评估T细胞的功能,我们使用了细胞毒性杀伤试验。结果经h1a处理的单核细胞在多个供体中PD-L1的总表达降低,CCL2分泌短暂增加。根据单细胞分泌的分析物的数量,H1A处理的单核细胞具有更大的多功能性。H1A处理的单核细胞有显著的转录谱变化,这与CCL2的转录激活有关。用H1A处理的pbmc导致更多的效应CD8 T细胞和更少的调节性T细胞群。我们的数据表明,单核细胞内PD-L1信号传导抑制了人单核细胞的转录激活和随后的CCL2分泌,从而限制了效应T细胞的数量。H1A抗体消除这种抑制机制,恢复效应T细胞反应。我们的研究意义有助于理解单核细胞中PD-L1的新作用机制,该机制可能导致癌症患者对抗pd -1/PD-L1治疗无反应。H1A抗体提供了一种新的工具,可以克服这些限制,增强t细胞介导的抗肿瘤免疫,延长致命癌症患者的生存时间。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
477 A novel antibody targeting human monocyte-intrinsic PD-L1 promotes immune stimulatory functions of monocytes for antitumor immunity

Background

Current PD-L1 targeting antibodies have been developed to block PD-L1’s interaction with PD-1, thereby preventing inhibition of T cell cytotoxicity. However, there has been limited clinical success in the treatment of cancers, despite high expression of PD-L1. Recent reports have demonstrated that tumor-intrinsic PD-L1 can signal intracellularly to promote cell survival independent of PD-1 ligation, potentially explaining why some cancer patients do not respond to immune checkpoint therapies. Besides tumor cells, host myeloid cells are sources of PD-L1 and can be highly immunosuppressive. Unfortunately, the intrinsic functions of PD-L1 in myeloid cells has not been well studied. We aim to dissect the intrinsic signaling of PD-L1 in monocytes, a subset of myeloid cells, and to investigate how this may be impairing antitumor immunity.

Methods

Our lab has identified a new PD-L1 antibody (clone H1A), which destabilizes PD-L1 at the cell surface and induces its degradation. In our experiments, we used human PBMCs isolated from healthy donor blood and isolated monocytes from PBMCs using negative magnetic selection. To study the effects H1A-induced PD-L1 degradation on human monocytes, we assessed monocyte phenotype, function, and transcriptional profile by flow cytometry, immunoassays, and single-cell RNA sequencing, respectively. To study the indirect effects of H1A on T cell functional states, we evaluated PBMCs by flow cytometry and mass cytometry using T cell focused panels. To evaluate T cell function, we used cytotoxic killing assays.

Results

H1A-treated monocytes resulted in decreased total expression of PD-L1 and a transient increase of CCL2 secretion across multiple donors. H1A treated monocytes had greater polyfunctionality based on the number of analytes secreted by single cells. H1A treated monocytes had significant transcriptional profile changes, related to transcriptional activation of CCL2. PBMCs treated with H1A resulted in more effector CD8 T cell and less regulatory T cell populations. Finally, H1A treatment of PBMCs resulted in greater T cell-mediated killing of tumor cells

Conclusions

Our data suggests monocyte-intrinsic PD-L1 signaling inhibits transcriptional activation and subsequent secretion of CCL2 in human monocytes, thereby restricting effector T cells populations. H1A antibody abolishes this inhibitory mechanism and restores effector T cell responses. The significance of our studies contributes to understanding a new mechanism of action of PD-L1 in monocytes that may cause cancer patients to not respond to anti-PD-1/PD-L1 therapy. The H1A antibody provides a new tool that can overcome these limitations to enhance T-cell mediated antitumor immunity and prolong survival of patients with lethal cancers.
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