Department of Medicine, Division of Pulmonary and Critical Care, University of California San Diego, La Jolla, CA 92037 USA.
Mol Cancer. 2010 Dec 20;9:316. doi: 10.1186/1476-4598-9-316.
Lymph node metastasis constitutes a key event in tumor progression. The molecular control of this process is poorly understood. Heparan sulfate is a linear polysaccharide consisting of unique sulfate-modified disaccharide repeats that allow the glycan to bind a variety of proteins, including chemokines. While some chemokines may drive lymphatic trafficking of tumor cells, the functional and genetic importance of heparan sulfate as a possible mediator of chemokine actions in lymphatic metastasis has not been reported.
We applied a loss-of-function genetic approach employing lymphatic endothelial conditional mutations in heparan sulfate biosynthesis to study the effects on tumor-lymphatic trafficking and lymph node metastasis. Lymphatic endothelial deficiency in N-deacetylase/N-sulfotransferase-1 (Ndst1), a key enzyme involved in sulfating nascent heparan sulfate chains, resulted in altered lymph node metastasis in tumor-bearing gene targeted mice. This occurred in mice harboring either a pan-endothelial Ndst1 mutation or an inducible lymphatic-endothelial specific mutation in Ndst1. In addition to a marked reduction in tumor metastases to the regional lymph nodes in mutant mice, specific immuno-localization of CCL21, a heparin-binding chemokine known to regulate leukocyte and possibly tumor-cell traffic, showed a marked reduction in its ability to associate with tumor cells in mutant lymph nodes. In vitro modified chemotaxis studies targeting heparan sulfate biosynthesis in lymphatic endothelial cells revealed that heparan sulfate secreted by lymphatic endothelium is required for CCL21-dependent directional migration of murine as well as human lung carcinoma cells toward the targeted lymphatic endothelium. Lymphatic heparan sulfate was also required for binding of CCL21 to its receptor CCR7 on tumor cells as well as the activation of migration signaling pathways in tumor cells exposed to lymphatic conditioned medium. Finally, lymphatic cell-surface heparan sulfate facilitated receptor-dependent binding and concentration of CCL21 on the lymphatic endothelium, thereby serving as a mechanism to generate lymphatic chemokine gradients.
This work demonstrates the genetic importance of host lymphatic heparan sulfate in mediating chemokine dependent tumor-cell traffic in the lymphatic microenvironment. The impact on chemokine dependent lymphatic metastasis may guide novel therapeutic strategies.
淋巴结转移是肿瘤进展中的一个关键事件。这个过程的分子控制机制还不太清楚。肝素硫酸盐是一种线性多糖，由独特的硫酸化二糖重复序列组成，使聚糖能够结合多种蛋白质，包括趋化因子。虽然一些趋化因子可能会促进肿瘤细胞的淋巴转移，但肝素硫酸盐作为趋化因子在淋巴转移中作用的可能介质的功能和遗传重要性尚未得到报道。
我们采用了一种丧失功能的遗传方法，利用淋巴内皮条件性突变来研究肝素硫酸盐生物合成对肿瘤-淋巴转移和淋巴结转移的影响。在参与硫酸化新生肝素硫酸盐链的关键酶 N-去乙酰基/N-磺基转移酶-1（Ndst1）的淋巴内皮中缺乏，导致携带基因靶向肿瘤的小鼠的淋巴结转移发生改变。这种情况发生在携带泛内皮 Ndst1 突变或诱导性淋巴内皮特异性 Ndst1 突变的小鼠中。除了突变小鼠中肿瘤转移到局部淋巴结的明显减少外，肝素结合趋化因子 CCL21 的特异性免疫定位也显示出其与肿瘤细胞结合的能力明显降低，已知 CCL21 调节白细胞，可能还有肿瘤细胞的转移。在针对淋巴内皮细胞中肝素硫酸盐生物合成的体外修饰趋化性研究中，发现淋巴内皮分泌的肝素硫酸盐是 CCL21 依赖的定向迁移所必需的，无论是对小鼠还是人肺癌细胞都是如此，向靶向的淋巴内皮。淋巴肝素硫酸盐也是 CCL21 与其在肿瘤细胞上的受体 CCR7 结合以及暴露于淋巴条件培养基的肿瘤细胞中迁移信号通路激活所必需的。最后，淋巴细胞表面肝素硫酸盐促进了 CCL21 在淋巴内皮上的受体依赖性结合和浓缩，从而作为一种机制来产生淋巴趋化因子梯度。
这项工作证明了宿主淋巴肝素硫酸盐在介导淋巴微环境中趋化因子依赖性肿瘤细胞转移中的遗传重要性。对趋化因子依赖性淋巴转移的影响可能为新的治疗策略提供指导。
