The DNA damage-dependent expression of ligands for cytotoxic NK-cell receptors: Impact of the DNA damage response on inside out signaling in CLL

细胞毒性 NK 细胞受体配体的 DNA 损伤依赖性表达：DNA 损伤反应对 CLL 中由内而外信号传导的影响

• 批准号: 234151796
• 负责人: Professorin Dr. Elke Pogge von Strandmann
• 金额: --
• 依托单位: Zentrum für Tumor- und Immunbiologie (ZTI)
• 依托单位国家: 德国
• 项目类别: Clinical Research Units
• 财政年份: 2013
• 资助国家: 德国
• 起止时间: 2012-12-31 至 2016-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/234151796?language=en
• 关键词: DNA; damage; dependent; expression; ligands

Aim of this project is to elucidate the role of genetic lesions and the cellular DNA damage response for the immune escape mechanisms in CLL. Normal cells respond to DNA damage in two ways. One response includes cell cycle arrest followed by DNA repair or, after severe damage, by apoptosis. The other response aims at signaling the comprised state of the dangerous cells to the innate immune system. This communication occurs through the induced expression of cellular ligands for activating immune receptors such as NKG2D, NKp30 and NKp46.The expression of ligands or "danger molecules" on released vesicles (exosomes) and on the surface of affected cells allows recognition and killing of the damaged cell by NK cells. Recently, we and others observed that the activity of NK cells from CLL patients is impaired and serum levels for soluble ligands engaging NKG2D and NKp30 are significantly elevated in comparison to healthy donors. Soluble ligands cause a general NK cell inhibition and high serum levels correlate with a poor prognosis indicating that NK cells are critically involved in immunosurveillance of CLL. The molecular mechanisms directing the inducible expression of ligands for NKG2D are not fully defined but known to be ATM/ATR-dependent. To uncover the impact of the DDR for ligand upregulation in CLL we will analyze the proposed ATM-dependent induction of the ligands using RNAi technology and established knockout mice. Specifically, we will cross the TCL1 transgenic mouse with ATM, TP53 and CBP/P300 knockouts. These compound mutant mice will be compared to TCL1 transgenic counterparts to test the hypothesis that components of the DDR contribute to the inducible ligand expression and to the generation of an anti-tumor immune response. Based on previous data we expect that loss of ATM, p53 and CBP/p300 in the CLL mouse model is essential to sensitize CLL cells to the NK cell attack and promotes tumor rejection by the innate immune system.Finally we plan to restore NK cell function by a rescue of ligand expression on malignant CLL cells to prove that NK cell inhibition results from the impaired expression of NKR-ligands. We will design bispecific fusion proteins (immunoligands) of ligands for NKG2D fused to an antibody fragment detecting the B-CLL-specific antigen ROR1. The construct is used to decorate ROR1-expressing CLL cells with NKG2D ligands rendering the cells more susceptible to NK cell-mediated killing. The effect of the restored ligand expression/decoration will be evaluated in the CLL mouse models and also ex vivo using NK cells derived from healthy donors or CLL patients. We aim to derive novel, genetically-informed combination therapies consisting of frontline DNA-damaging agents with innovative approaches that harness NK cells for cancer therapy.

该项目的目的是阐明CLL中免疫逃逸机制的遗传病变和细胞DNA损伤反应的作用。正常细胞以两种方式应对DNA损伤。一种反应包括细胞周期停滞，然后进行DNA修复，或在严重损害后通过细胞凋亡。另一个响应旨在向先天免疫系统发出包含的危险细胞状态。这种通信是通过诱导的细胞配体表达发生的，用于激活诸如NKG2D，NKP30和NKP46的免疫受体。在释放的囊泡（外部）和受影响的细胞表面上，配体或“危险分子”的表达允许识别和杀死。 NK细胞受损的细胞。最近，我们和其他人观察到，与健康供体相比，与NKG2D和NKP30相比，CLL患者的NK细胞的活性受损，可溶性配体的血清水平显着升高。可溶性配体引起一般的NK细胞抑制作用，高血清水平与预后不良相关，表明NK细胞与CLL的免疫监视至关重要。指导NKG2D配体诱导表达的分子机制未完全定义，但已知是ATM/ATR依赖性的。为了揭示DDR对CLL配体上调的影响，我们将使用RNAi技术和建立的基因敲除小鼠分析提出的ATM依赖性诱导。具体而言，我们将使用ATM，TP53和CBP/P300敲除TCL1转基因小鼠穿越TCL1转基因小鼠。这些复合突变小鼠将与TCL1转基因对应物进行比较，以检验DDR的成分有助于诱导的配体表达以及抗肿瘤免疫反应的产生的假设。根据先前的数据，我们预计CLL小鼠模型中ATM，p53和CBP/P300的损失对于将CLL细胞敏感到NK细胞攻击并促进先天免疫系统的肿瘤排斥。我们计划恢复NK细胞功能。通过拯救恶性CLL细胞上的配体表达，证明NK细胞抑制是由NKR配体的表达受损而导致的。我们将针对融合到抗体片段的NKG2D的配体的双特异性融合蛋白（免疫原体），以检测B-CLL特异性抗原ROR1。该构建体用于用NKG2D配体来装饰表达ROR1的CLL细胞，使细胞更容易受到NK细胞介导的杀伤的影响。将在CLL小鼠模型中评估恢复的配体表达/装饰的效果，并使用来自健康供体或CLL患者的NK细胞进行离体评估。我们旨在得出新型的，具有遗传信息的组合疗法，这些疗法由前线DNA损害剂组成，其创新方法可以利用NK细胞进行癌症治疗。