Evaluation of P2X7 as a therapeutic target in autoimmune encephalomyelitis and in tumor immunity

P2 X7（一种配体门控离子通道）和ARTC 2（一种毒素相关胞外酶）是免疫细胞表面蛋白，它们在无菌炎症期间感知细胞释放的ATP和NAD+作为内源性危险信号。在骨髓细胞上，P2 X7是炎症的关键介质。在T细胞上，ARTC 2和P2 X7调节Treg功能和存活。我们在先前的合作研究中发现，通过激活ARTC 2/P2 X7轴来消耗调节性T细胞可用于促进抗肿瘤免疫应答。最近，我们开发了P2 X7阻断和P2 X7增强纳米抗体（Nbs）和基于Nb的生物制剂。我们发现，在体内用这些生物制剂阻断P2 X7功能代表了一种有前途的改善炎症的新治疗策略。这些生物制剂可以通过重复全身注射或在用编码这些Nb的腺相关病毒（AAV）载体体内转导细胞后通过持续内源性合成来施用。到目前为止，人们对这些生物制剂的体内药效学以及它们到达和阻断淋巴器官、炎症组织或肿瘤中免疫细胞靶点的能力知之甚少。血脑屏障（BBB）尤其是蛋白质生物制剂到达中枢神经系统（CNS）靶点的天然障碍。此外，P2 X7在自身免疫性炎性疾病和肿瘤微环境中的作用仍需要精确描述。本研究的中心目标是评估P2 X7作为实验性自身免疫性脑脊髓炎（EAE）模型和抗肿瘤反应中的治疗靶点。本研究的第二个目标是使用基于Nb的生物制剂优化P2 X7在CNS和肿瘤微环境中的体内靶向。在这两种模型中，我们将利用全身注射以及AAV介导的基于Nb的生物制剂的长期内源性产生，以更好地理解P2 X7的作用，并评估与其短期和长期功能调节相关的治疗益处。此外，我们将探索Fc工程和双特异性靶向以增强基于Nb的生物制剂的效应器功能的潜力。我们期望该项目的结果为P2 X7在炎症和免疫中的病理生理作用提供有价值的新见解，并验证其作为治疗靶点的相关性。

P2X7, a ligand-gated ion channel, and ARTC2, a toxin-related ecto-enzyme, are immune cell surface proteins that sense ATP and NAD+ released from cells as endogenous danger signals during sterile inflammation. On myeloid cells, P2X7 is a key mediator of inflammation. On T cells, ARTC2 and P2X7 regulate Treg function and survival. We have discovered in a previous collaborative study, that depletion of regulatory T cells by activation the ARTC2/P2X7 axis can be used to promote anti-tumor immune responses. Recently, we have developed P2X7-blocking and P2X7-potentiating nanobodies (Nbs) and Nb based biologics. We found that blocking P2X7 function with these biologics in vivo represents a promising novel therapeutic strategy to ameliorate inflammation. These biologics can be administered either by repeated systemic injections or via continued endogenous synthesis after transduction of cells in vivo with adeno-associated viral (AAV) vectors coding for these Nb. So far, little is known about the in vivo pharmacodynamics of these biologics, and about their capacity to reach and block their targets on immune cells in lymphatic organs, inflamed tissues or tumors. The blood-brain barrier (BBB) in particular poses a natural obstacle for protein-based biologics to reach their targets in the central nervous system (CNS). Moreover, the role of P2X7 in autoimmune inflammatory diseases and in the tumor microenvironment still need to be precisely delineated. The central goal of this study is to evaluate P2X7 as a therapeutic target in models of experimental autoimmune encephalomyelitis (EAE) and in anti-tumor response. A second goal of this study is to optimize the in vivo targeting of P2X7 in the CNS and in the tumor microenvironment using Nb-based biologics. In both models, we will exploit systemic injections as well as AAV-mediated long-term endogenous production of the Nb-based biologics to better understand the role P2X7 and to evaluate the therapeutic benefits associated with its short and long-term functional modulation. Moreover, we will explore the potential of Fc-engineering and of bispecific-targeting to enhance the effector functions of Nb based biologics. We expect the results of this project to provide valuable new insights into the pathophysiological roles of P2X7 in inflammation and immunity and to validate its relevance as a therapeutic target.