Project 4: Modulating the natural killer cell response to oHSV1 in recurrent human GBM

项目摘要--项目4 这项建议的最终目标是了解先天免疫在溶瘤的背景下的作用。 单纯疱疹病毒(OHSV)治疗胶质母细胞瘤(GBM)，一种高度致命的脑瘤。单纯疱疹病毒感染治疗基底细胞瘤 依赖于癌症特异性病毒的复制，导致肿瘤破坏，对邻近肿瘤的毒性最小 非肿瘤组织。它在患者身上的安全性已经得到证实，但是否有显著疗效的证据仍有待证实。 已经成立了。项目4将重点放在接种OHSV后宿主的自然杀伤(NK)细胞反应。 由于其强大的抗病毒特性，NK细胞是OHSV治疗的潜在障碍。或者，也可以使用 抗肿瘤NK细胞反应有可能增强OHSV治疗的肿瘤清除特性。我们的 先前发表的研究表明：1)NK细胞对OHSV快速反应并在感染前清除病毒 它可以有效地复制和传播到整个GBM；2)瞬时免疫调节延缓 NK细胞对OHSV的反应，即使是一次注射转化生长因子-β，也显著增强了OHSV的复制， 接种后的扩散和抗肿瘤效果。因此，我们假设NK细胞协调一种 最初接种OHSV后产生的强大炎症反应限制了OHSV的复制、传播和 肿瘤溶解，从而为有效的OHSV治疗基底膜创造了障碍。这一假设的推论是 通过了解NK细胞如何识别OHSV感染的GBM，我们可以最好地调节这一过程，以便 优化这种高度选择性的治疗GBM的方法。因此，我们最近开发了一种新的分析方法，通过这种方法，我们可以 将每个HSV1基因克隆到载体中，使我们能够确定其诱导或抑制NK细胞的能力 激活。我们先前发现了NK细胞识别和破坏HSV感染靶点的机制 与初级免疫反应的发展有关。我们将这种基本的、新发现的被动抗体称为被动抗体 依赖性细胞毒性或被动ADCC，由此免疫球蛋白(IgGFc)的Fc片段形成桥梁 在NK细胞针对IgGFc的Fc受体CD16和表达HSV US8的GBM细胞之间，后者编码 Fc结合蛋白糖蛋白E(Ge)。CD16-IgGFc-Ge三元复合体激活NK细胞破坏 OHSV感染的GBM不需要免疫球蛋白的抗原特异性Fab部分。在本建议书中，项目 4将：(1)进一步研究被动ADCC，以最好地了解如何调节这一过程以增强 (2)确定转化生长因子-β信号转导通路在调节血管生成中的作用。 NK细胞与OHSV编码病毒蛋白的相互作用，特别是与被动ADCC和Noch相关 项目3的信号传递；(3)评估复发的GBM患者的NK细胞免疫反应 在项目2中接受我们的OHSV，rQNestin34.5。通过阐明NK细胞如何识别和破坏OHSV- 在感染肿瘤的其他促炎和抗炎过程中，我们将最好地理解如何 优化这种对GBM的神经选择性治疗，并识别导致NK的关键机制信号 细胞介导的病毒感染的清除。

PROJECT SUMMARY – PROJECT 4 The ultimate goal of this proposal is to understand the role of innate immunity within the context of oncolytic herpes simplex viral (oHSV) therapy for glioblastoma (GBM), a highly fatal brain tumor. oHSV treatment of GBM relies on cancer-specific replication of the virus leading to tumor destruction with minimal toxicity to adjacent non-neoplastic tissue. Its safety in patients has been proven, yet evidence for significant efficacy remains to be established. Project 4 will focus on the host's natural killer (NK) cell response following oHSV administration. Due to their strong antiviral properties, NK cells represent a potential barrier to oHSV therapy. Alternatively, the anti-tumor NK cell response has the potential of augmenting the tumor clearing properties of oHSV therapy. Our previously published studies demonstrated that: 1) NK cells rapidly respond oHSV and eliminate the virus before it can effectively replicate and disseminate throughout the GBM; 2) transient immune modulation delaying the NK cell response to oHSV, even with a single injection of TGF-beta, significantly enhances oHSV replication, spread and anti-tumor efficacy following inoculation. As a result, we hypothesize that NK cells coordinate a robust inflammatory response following initial oHSV administration that limits oHSV replication, spread, and tumor lysis, thereby creating a barrier to effective oHSV therapy for GBM. A corollary to this hypothesis is that by understanding how NK cells recognize oHSV-infected GBM we can best modulate this process so as to optimize this highly selective therapy for GBM. We therefore recently developed a novel assay whereby we cloned each HSV1 gene into a vector that allowed us to determine its ability to either induce or inhibit NK cell activation. We discovered the mechanism by which NK cells recognize and destroy HSV-infected targets prior to the development of a primary immune response. We term this fundamental, novel discovery passive antibody dependent cellular cytotoxicity or passive ADCC, whereby the Fc fragment of IgG (IgGFc) forms a bridge between the NK cell Fc receptor for IgGFc, called CD16, and the GBM cell expressing HSV Us8, which encodes the Fc binding protein glycoprotein E (gE). The CD16-IgGFc-gE ternary complex activates NK cells to destroy oHSV-infected GBM without the requirement for the antigen-specific Fab portion of IgG. In this proposal, Project 4 will: (1) further investigate passive ADCC to best understand how to modulate the process so as to enhance the efficacy of oHSV therapy for GBM with Project 1; (2) determine the role of TGF-β signaling in regulating the NK cell interaction with oHSV-encoded viral proteins, specifically as relates to passive ADCC and to NOTCH signaling with Project 3; (3) To assess the NK cell immune response in patients with recurrent GBM who will be receiving our oHSV, rQNestin34.5 in Project 2. By elucidating how NK cells recognize and destroy oHSV- infected tumors in the context of other pro- and anti-inflammatory processes, we will best understand how to optimize this neuro-selective treatment for GBM as well as discern the key mechanistic signals leading to NK cell-mediated clearance of viral infection.