Influence of the skeletal remodeling status on tumor cell dissemination and metastatic outgrowth

骨基质通过两种基本上不同的细胞类型（骨形成成骨细胞和骨再吸收破骨细胞）的平衡活动而不断重塑。我们的研究所在转基因小鼠模型的骨特异性细胞和分子表征方面拥有长期的专业知识，在过去的几年里，我们已经确定了控制骨骼重塑的几个关键机制。对于目前的提议，我们将继续与Klaus Pantel教授的小组进行合作，以解决这个问题，即受损的骨重建状态是否以及如何影响肿瘤细胞在体内的传播和转移性生长。为此，我们将人和小鼠乳腺癌细胞注射到小鼠体内，通过荧光素酶报告基因测定，µCT，不脱钙组织学和骨特异性组织形态计量学。最重要的是，与大多数先前发表的研究相反，我们的目标不是改变种子（即肿瘤细胞），而是改变土壤（即受体小鼠）。为了实现我们的目标，我们将不同的小鼠模型回交到遗传背景（BALB/c和NSG）中，允许癌细胞的异种移植或同基因移植。其中三个模型（Calca-/-、Calcr-/-和Notch 2 +/HCS）显示出同步骨形成和骨反应所需的偶联机制的特定干扰，而其他四个模型（Lrp 5 +/HBM、Col 1a 1-Sost、Col 1a 1-Krm 2和Col 1a 1-tTA; pTet-Wnt 1）显示出成骨细胞活性的选择性变化。由于所有这些转基因小鼠模型先前都已进行了分析，不仅确定了它们的骨重建状态，而且确定了观察到的表型的分子原因，因此我们预计我们的结果将清楚地确定，这种骨重建细胞类型主要参与骨和肿瘤细胞之间的有害串扰。除了这种体内方法，我们将进行离体共-用乳腺癌和骨重建细胞进行培养实验，以确定特定的分子相互作用。虽然我们的项目主要遵循一个公正的方法，有一些假设，将得到解决。例如，由于我们先前发现降钙素（CT）通过抑制破骨细胞释放鞘氨醇1-磷酸（S1 P）来控制骨形成，我们将分析CT、S1 P和/或特定S1 P受体的拮抗剂是否会影响单独培养或与骨重塑细胞类型一起培养的癌细胞的增殖和/或迁移。

The bone matrix is continuously remodeled through the balanced activities of two substantially different cell types, bone-forming osteoblasts and bone-resorbing osteoclasts. Our Institute has a long expertise in bone-specific cellular and molecular characterization of genetically modified mouse models, and we have identified several key mechanisms controlling skeletal remodeling in the last years. For the present proposal we will continue our ongoing collaboration with the group of Prof. Klaus Pantel to address the question, if and how an impaired bone remodeling status affects tumor cell dissemination and metastatic outgrowth in vivo.For that purpose we will inject human and murine breast cancer cells into mice to study skeletal metastasis formation as well as changes in the bone microenvironment by Luciferase reporter gene assays, µCT, undecalcified histology and bone-specific histomorphometry. Most importantly, and in contrast to the majority of previously published studies, we do not aim at modifying the seed (i.e. the tumor cells), but the soil (i.e. the recipient mouse). To achieve our goals we have backcrossed different mouse models into genetic backgrounds (BALB/c and NSG) allowing xenotransplantation or syngeneic transplantation of cancer cells. While three of the models (Calca-/-, Calcr-/- and Notch2+/HCS) display specific disturbances of the coupling mechanisms required to synchronize bone formation and bone resoprtion, the other four models (Lrp5+/HBM, Col1a1-Sost, Col1a1-Krm2 and Col1a1-tTA;pTet-Wnt1) display selective changes in osteoblast activity. Since all these genetically modified mouse models have been previously analyzed to define not only their bone remodeling status, but also the molecular causes of the observed phenotypes, we expect that our results will clearly identify, which bone remodeling cell type is primarily involved in the suggested detrimental crosstalk between bone and tumor cells.In addition to this in vivo approach we will perform ex vivo co-culture experiments with breast cancer and bone remodeling cells to define specific molecular interactions. Although our project principally follows an unbiased approach, there are some hypotheses that will be addressed. For instance, since we have previously found that calcitonin (CT) controls bone formation by inhibiting the release of sphingosine 1-phosphate (S1P) from osteoclasts, we will analyze if CT, S1P and/or antagonists of specific S1P receptors will influence proliferation and/or migration of cancer cells, cultured alone or together with bone remodeling cell types.