Characterization of the premetastatic niche at different sites and in vivo imaging of granulocytic myeloid derived suppressor cells

癌症的一个重要特征是出现转移，通常代表癌症疾病的最后阶段。值得注意的是，一些肿瘤类型偏爱特定的器官进行转移扩散。与将转移形成描述为随机事件的陈旧理论相反，目前的转移扩散被认为是一个多步骤的过程，在循环中的肿瘤细胞能够种植之前，在靶器官中形成转移前生态位(PMN)。作为原发肿瘤信号传递的结果，不同类型的骨髓来源干细胞的渗透，特征黏附分子的表达增加，以及细胞外基质的重塑发生。原发肿瘤内的低氧诱导信号，导致PMN的出现增加，并伴有粒细胞髓系衍生抑制细胞(G-MDSCs)的渗透。到目前为止，最近的文献主要描述了PMN在肺内的发育情况。在这个项目中，我们将在小鼠乳腺癌转移模型(PYMT)中研究PMN在骨髓和脑中的出现情况。将使用专门生成的具有骨髓和脑转移特异性的Cherry-red/Luciferase-transgenic-PymT-breastcancer细胞。在这些实验中，用来自骨髓或脑特异性乳腺癌细胞的低氧条件培养液治疗实验动物，会导致PMN在各自部位的出现增加。本项目主要研究骨髓和脑内中性粒细胞的特性，以及G-MDSCs向PMN迁移的体内成像，可分为以下子项目：在资助期：A)骨髓和脑中的PMN内G-MDSCs的特性。B)肺、骨髓和脑转移的体内光学成像。C)建立分离的G-MDSCs的荧光染料Cy5标记。过继转移后，这些G-MDSCs在肺、骨髓和脑内的PMN的归巢将通过光学成像进行可视化。资助期后：d)建立PET(放射性CD11b单抗，单纯疱疹病毒胸苷激酶报告基因)或MRI兼容的G-MDSCs标记策略(氧化铁颗粒，19F)，用于通过同步的PET/MRI在体内检测其归巢到PMN。E)通过过继转移的G-MDSCs在不同的小鼠转移模型中同时对PMN进行PET/MRI-PMN特异性示踪剂的开发。综上所述，本项目将研究中性粒细胞在骨髓和脑内的发育，并随后建立G-MDSCs向中性粒细胞的迁移动力学，以预测未来转移的位置。拟议的试验将在2015年后的资助期后持续下去。

A significant hallmark of cancer is the appearance of metastasis, frequently representing the final stage of a cancer disease. It is conspicuous, that some tumor types prefere specific organs for metastatic spread. Contrary to obsolete theories, which describe metastasis formation as random event, the metastatic spread is considered nowadays as multi-step process, initiated by the formation of the premetastatic niche (PMN) in the target organ before circulating tumor cells are able to seed. As a consequence of signaling of the primary tumor, an infiltration of different bone marrow derived stem cell types, an increased expression of characteristic adhesion molecules and a remodeling of the extracellular matrix occurs. Hypoxia within the primary tumor induces signaling which results in an increased appearance of PMN, accompanied by an infiltration of granulocytic myeloid derived suppressor cells (G-MDSCs). So far, mainly the development of the PMN in the lung was described in recent publications. In this project the appearance of PMN in the bone marrow and brain will be investigated in a murine breast cancer-metastasismodel (PymT). Specially generated Cherry-red/Luciferase-transgenic-PymT-breastcancer cells with specificity for metastasis in the bone marrow and brain will be used. In these experiments, the treatment of experimental animals with hypoxic conditioned medium derived from bone marrow- or brain-specific breastcancer-cells induces the increased appearance of PMN at the respective sites. This project focuses on the characterization of PMN in the bone marrow and brain, as well as the in vivo imaging of the migration of G-MDSCs to the PMN and can be divided into the following subprojects:During funding period:A) Characterization of G-MDSCs within the PMN of the bone marrow and brain.B) In vivo Optical Imaging of metastatic spread in the lung, bone marrow and brain.C) Establishment of the labeling with the fluorescence dye Cy5 for isolated G-MDSCs. After adoptive transfer, the homing of these G-MDSCs to the PMN in the lung, bone marrow and brain will be visualized by Optical Imaging. After funding period:D) Establishment of PET- (radioactive CD11b-monoclonal antibodies, herpes-simplex-thymidine-kinase-reportergene) or MRI-compatible labeling strategies (iron oxide particles, 19F) for G-MDSCs for the in vivo detection of their homing to the PMN by simultaneous PET/MRI. E) Simultaneous PET/MRI of the PMN by adoptively transferred G-MDSCs in alternative murine metastasis-models and development of PMN-specific tracers. In summary, in this project the development of the PMN in the bone marrow and brain will be investigated and subsequently the migration dynamics of G-MDSCs to PMN will be established to predict the location of the future metastasis. The proposed experiments will sustain after the funding period beyond 2015.