The Role of Endothelial ACKR1 in Triple-Negative Breast Cancer Metastasis

转移是乳腺癌死亡的最常见原因。在乳腺癌的亚型中，三重- 阴性乳腺癌（TNBC）是最致命的，因为其转移的可能性增加。肿瘤细胞 外渗是转移的关键步骤， 远端组织清楚地了解肿瘤细胞外渗的主要调节因子将为以下方面提供见解： TNBC转移的进展。 TNBC细胞外渗的一种潜在调节剂是ACKR 1。在许多情况下，ACKR 1表达是 内皮细胞（EC）需要白细胞外渗。内皮细胞ACKR 1结合CXCL 2，一种促迁移蛋白 趋化因子，并将其定位于EC连接处以引导中性粒细胞通过白细胞外渗。CXCL2 在TNBC细胞中的表达对于肿瘤细胞从肺微血管外渗和肿瘤转移也是必需的。 转移我们的初步数据显示，在TNBC的至少一种基质细胞类型中需要ACKR 1 从原发肿瘤转移到肺部。这些数据表明，内皮细胞ACKR 1-CXCL 2相互作用 可能介导TNBC转移中的肿瘤细胞外渗。因此，我们假设内皮细胞ACKR 1 通过将CXCL 2保留在EC连接处促进TNBC细胞外渗，导致增加的 TNBC的转移性扩散。 为了解决这一假设，我们将研究内皮细胞ACKR 1表达的体内意义， 使用我们验证的ACKR 1内皮细胞特异性敲除小鼠模型。我们将测试以下要求： 内皮ACKR 1用于原位植入TNBC肿瘤转移至肺部远端部位以及用于 循环肿瘤细胞外渗到肺组织中。我们将确定哪些步骤的外渗需要 通过使用Ibidi流式共培养系统评估ACKR 1-低和ACKR 1-过表达的EC， 概括了肺微血管的剪应力条件。我们将研究这些步骤是否 通过将CXCL 2中和抗体引入Ibidi流动系统并观察 它们对每个外渗步骤的影响。 我们提出的研究将确定内皮ACKR 1在TNBC转移进展中的作用， 确定肿瘤细胞外渗的具体步骤，其中内皮ACKR 1和CXCL 2发挥作用。 了解这些过程可能会指导开发ACKR 1作为转移和转移的预后标志物， 可以提供对待评估的候选趋化因子和趋化因子受体抑制剂的机理洞察 用于治疗乳腺癌。

Metastasis is the most common cause of breast cancer mortality. Of the breast cancer subtypes, triple- negative breast cancer (TNBC) is the deadliest due to its increased likelihood to metastasize. Tumor cell extravasation is a critical step of metastasis and allows circulating tumor cells to exit the vasculature and seed distant tissues. Clear understanding of the major regulators of tumor cell extravasation will provide insights into the progression of TNBC metastasis. One potential regulator of TNBC cell extravasation is ACKR1. In many contexts, ACKR1 expression is required in endothelial cells (EC) for leukocyte extravasation. Endothelial ACKR1 binds CXCL2, a promigratory chemokine, and localizes it to EC junctions to guide neutrophils through leukocyte extravasation. CXCL2 expression in TNBC cells is also necessary for tumor cell extravasation from lung microvasculature and for tumor metastasis. Our preliminary data show that ACKR1 is required in at least one stromal cell type for TNBC metastasis from the primary tumor to the lung. These data suggest that endothelial ACKR1-CXCL2 interactions may mediate tumor cell extravasation in TNBC metastasis. Therefore, we hypothesize that endothelial ACKR1 promotes TNBC cell extravasation by retaining CXCL2 at EC junctions, resulting in the increased metastatic spread of TNBC. To address this hypothesis, we will examine the in vivo significance of endothelial ACKR1 expression using our validated ACKR1 endothelial cell-specific knockout mouse model. We will test the requirement for endothelial ACKR1 for metastasis of orthotopically implanted TNBC tumors to distant sites in the lung and for extravasation of circulating tumor cells into lung tissue. We will determine which steps of extravasation require ACKR1 by evaluating ACKR1-low and ACKR1-overexpressing ECs using an Ibidi flow co-culture system that recapitulates the shear stress conditions of pulmonary microvasculature. We will examine whether these steps are dependent on CXCL2 by introducing CXCL2-neutralizing antibodies to the Ibidi flow system and observing their effects on each extravasation step. Our proposed studies will establish the role of endothelial ACKR1 in TNBC metastatic progression and determine the specific steps of tumor cell extravasation in which endothelial ACKR1 and CXCL2 function. Understanding these processes may guide development of ACKR1 as a prognostic marker for metastasis and can provide mechanistic insight into candidate chemokine and chemokine receptor inhibitors under evaluation for treatment of breast cancer.