Identification of tumor-cell intrinsic and microenvironmental factors influencing lymph node colonization in melanoma.

大多数癌症早期通过淋巴管扩散到同侧区域淋巴结盆的淋巴结。黑色素瘤扩散到区域淋巴结盆地的前哨淋巴结（sln）和非前哨淋巴结（nsln）是黑色素瘤预后的主要预测因素，这些组织为研究控制播散性癌细胞选择和适应的肿瘤细胞内在和微环境因素提供了独特的机会。我们之前的研究表明，癌细胞播散是黑色素瘤进展的早期阶段，播散性黑色素瘤细胞在播散时缺乏典型的驱动变化，如BRAF突变，但在前哨淋巴结的集落形成过程中继续进化并获得它们。我们首次开发了一种工作流程，允许将患者淋巴结肿瘤细胞负荷的精确定量评估与免疫细胞和播散性癌细胞（DCCs）的功能和分子分析相结合，我们的项目旨在了解肿瘤细胞内在和微环境因素如何促进DCCs的生长和淋巴结定植。我们将研究与淋巴结定植和播散性癌细胞遗传成熟相关的免疫微环境的变化。由于我们之前的工作已经暗示了外泌体途径对DCCs定植的重要性，我们将更详细地研究其对免疫监视的影响。我们将利用从黑色素瘤患者的淋巴结定殖dcc中建立的细胞系，表征它们的外泌体miRNA和选定的蛋白质含量，并评估它们对免疫细胞表型和功能的影响。对于免疫细胞在抗肿瘤免疫反应中产生的细胞因子TNF和IFNg，已经报道了抑制和促进肿瘤生长的作用。因此，我们将研究它们在来自非定植和定植淋巴结的播散性癌细胞的不同遗传成熟状态下对生长和存活的作用。总之，我们对dcc和免疫细胞的综合表型和功能分析将揭示淋巴结定植的规律，并从根本上提高我们对辅助治疗情况下治疗方案的理解。

Most cancers spread early through lymphatic vessels to lymph nodes in the ipsilateral, regional lymph node basin. Melanoma spread to sentinel (SLNs) and non-sentinel nodes (NSLNs) of the regional nodal basin are major predictors for melanoma outcome and these tissues offer the unique opportunity to study tumor cell-intrinsic and microenvironmental factors governing selection and adaptation of disseminated cancer cells. Our previous work revealed that cancer cell dissemination is an early step in melanoma progression and disseminated melanoma cells lack typical driver changes, such as BRAF mutations at dissemination, but continue to evolve and acquire them during colony formation in the sentinel lymph node. Having developed a work flow allowing for the first time to combine the precise quantitative assessment of the tumor cell load of patient lymph nodes with functional and molecular analyses of immune cells and disseminated cancer cells (DCCs), our project aims at understanding how tumor-cell intrinsic and microenvironmental factors contribute to outgrowth of DCCs and lymph node colonization. We will investigate changes in the immune microenvironment associated with lymph node colonization and genetic maturation of disseminated cancer cells. As our previous work has implicated the importance of the exosomal pathway for colonizing DCCs, we will investigate its influence on immunosurveillance in more detail. We will utilize cell lines established from lymph node-colonizing DCCs of melanoma patients, characterize their exosomal miRNA and selected protein-content and evaluate their influence on the phenotype and functionality of immune cells. For the cytokines TNF and IFNg, which are produced during an anti-tumoral immune response by immune cells, both inhibitory and tumor-growth promoting effects have been reported. We will therefore investigate their role on growth and survival as a function of the different genetic maturation status of disseminated cancer cells from non-colonized and colonized lymph nodes. In summary, our combined phenotypic and functional analyses of DCCs and immune cells will unravel the rules of lymph node colonization and fundamentally improve our understanding of therapeutic options in the adjuvant therapeutic situation.