The influence of B cells on the therapeutic success of immunotherapy in patients with head and neck cancer

肿瘤浸润性B细胞在实体瘤中的作用一直被低估。仅在最近几年才变得清楚的是，B细胞的存在通常与各种实体癌如头颈癌（HNC）的良好预后相关。另一方面，B细胞也被赋予肿瘤促进作用，这得到了几种鼠肿瘤模型的支持。为了进一步研究，有必要将B细胞群进一步细分为免疫活性效应B细胞和免疫抑制调节B细胞。使用检查点抑制剂（CPI）（如PD 1抗体）进行免疫治疗的可能性也导致了KHT患者生存率的增加。由于不表达PD-L1配体的肿瘤患者也受益于这种疗法，因此PD-1抗体的确切作用机制尚未阐明。在本申请的上下文中，我们正在研究不同的B细胞群如何影响CPI治疗。第一部分研究CPI如何在体外增强免疫系统，特别是B细胞功能。这是使用来自HNC患者或来自健康对照供体的血液和肿瘤样品进行的。第二部分主要研究CPI对小鼠原位肿瘤模型免疫系统的影响。在此模型中，CPI与其他抗体（抗CD 20、抗CD 73）结合或通过敲除方法删除B细胞特异性蛋白。第三部分研究了如何在肿瘤源性exosomes的帮助下修饰B细胞的功能。这些研究在体内和体外进行。使用微型尺寸排阻色谱法分离外泌体，并通过纳米颗粒tracking. Fourth部分将我们用CPI治疗的肿瘤患者的临床数据与肿瘤中B细胞群和三级淋巴结构的发生相关联。现有的肿瘤微阵列允许研究大量的肿瘤样品。使用多重血清学检查患者血清样品的抗原模式。该应用的目的是通过操纵HNC患者肿瘤微环境中的B细胞来改善肿瘤治疗。这些结果可以在进一步的过程中转移到其他类型的肿瘤。两个申请人的活动领域，一方面在诊所，另一方面在研究实验室，是互补的，从而促进了建议的实验的快速和成功的实施。

The role of tumor-infiltrating B cells in solid tumors has long been underestimated. Only in the last few years has it become clear that the presence of B cells is often associated with a good prognosis in a variety of solid cancers such as head and neck cancer (HNC). On the other hand, B cells are also assigned a tumor-promoting role, which is supported by several murine tumor models. For further investigations it is therefore necessary to further subdivide the B-cell population into immunocompetent effector B-cells and immunosuppressive regulatory B-cells.The possibilities of immunotherapy with checkpoint inhibitors (CPI), such as PD1 antibodies, have also led to an increase in survival in patients with KHT. Since patients with tumors who do not express the PD-L1 ligand, also benefit from this therapy, the exact mechanism of action of the PD-1 antibodies has not yet been clarified. Likewise, there are no reliable prognostic markers for this therapy.In the context of the present application, we are investigating how different B-cell populations can influence therapy with CPI. The first section examines how CPI can strengthen the immune system and in particular B-cell function in vitro. This is done using blood and tumor samples from patients with HNC or from healthy control donors. Mass spectrometry, UMAP flow cytometry and experiments with tumor organoids are used here.The second section focuses on the influence of CPI on the immune system in an established, orthotopic murine tumor model. In this model, CPI are combined with other antibodies (anti-CD20, anti-CD73) or B-cell-specific proteins are deleted by knockout methods.The third section examines how the function of B cells can be modified with the help of tumor-derived exosomes. These studies are carried out both in vivo and in vitro. The exosomes are isolated using mini-size exclusion chromatography and visualized by nanoparticle tracking.The fourth section correlates the clinical data of our tumor patients treated with CPI with the occurrence of B-cell populations and tertiary lymphoid structures in the tumor. Existing tumor microarrays allow the investigation of a large number of tumor samples. Serum samples from patients are examined for antigen patterns using multiplex serology.The aim of the application is to improve oncological therapy by manipulating B cells in the tumor microenvironment of patients with HNC. These results can be transferred to other types of tumors in the further course. The areas of activity of the two applicants, on the one hand in the clinic and on the other hand in the research laboratory, are complementary and thus promote the rapid and successful implementation of the suggested experiments.