Pharmacological blockade of aquaporin-1 water channel by AqB013 restricts migration and invasiveness of colon cancer cells and prevents endothelial tube formation in vitro.
Pharmacological blockade of aquaporin-1 water channel by AqB013 restricts migration and invasiveness of colon cancer cells and prevents endothelial tube formation in vitro.
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DOI：
10.1186/s13046-016-0310-6
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发表时间：
2016-02-24
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影响因子：
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通讯作者：
Hardingham JE
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文献类型：
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作者：
Dorward HS;Du A;Bruhn MA;Wrin J;Pei JV;Evdokiou A;Price TJ;Yool AJ;Hardingham JE
水通道蛋白（AQP）是一种水通道蛋白，能够使液体穿过细胞膜，对组织环境的稳态和细胞迁移很重要。AQP 1基因敲除的人类癌症小鼠模型显示出对肿瘤诱导的血管生成的显著抑制，并且在结肠腺癌的临床前研究中，AQP 1的强制过表达显示出增加血管生成、侵袭和转移。我们已经合成了AQP 1的小分子拮抗剂。我们的假设是，抑制AQP 1将减少结肠癌细胞的迁移和侵袭能力，以及体外内皮细胞的迁移和管形成能力。采用定量PCR、免疫印迹和免疫荧光法检测细胞系中AQP 1的表达，采用免疫组化法检测人结肠肿瘤组织中AQP 1的表达。使用伤口闭合（迁移）测定、基质胶侵袭测定和增殖测定，测试不同浓度的AQP 1抑制剂AqB 013对表达高与低水平AQP 1的人结肠癌细胞系的作用。使用内皮细胞管形成测定测试AqB 013对血管生成的作用。与溶剂对照相比，160或320 μM AqB 013处理的具有高AQP 1水平的HT 29结肠癌细胞显示出显著的迁移抑制，分别为27.9% ± 2.6%（p <0.0001）和41.2% ± 2.7%（p <0.0001），而对具有低AQP 1表达的HCT-116细胞的迁移没有影响。在侵袭测定中，用160 μM AqB 013处理的HT 29细胞显示在144小时侵袭降低60.3% ± 8.5%（p < 0.0001），并且与媒介物对照相比侵袭速率显著降低（F检验，p = 0.001）。与媒介物对照相比，在80 μM AqB 013下实现了内皮管形成的几乎完全抑制（血管生成测定）（p < 0.0001）。这些数据为进一步测试抑制剂作为结肠癌治疗剂提供了良好的证据。本文的在线版本（doi：10.1186/s13046-016-0310-6）包含补充材料，可供授权用户使用。
Aquaporins (AQP) are water channel proteins that enable fluid fluxes across cell membranes, important for homeostasis of the tissue environment and for cell migration. AQP1 knockout mouse models of human cancers showed marked inhibition of tumor-induced angiogenesis, and in pre-clinical studies of colon adenocarcinomas, forced over-expression of AQP1 was shown to increase angiogenesis, invasion and metastasis. We have synthesized small molecule antagonists of AQP1. Our hypothesis is that inhibition of AQP1 will reduce migration and invasiveness of colon cancer cells, and the migration and tube-forming capacity of endothelial cells in vitro. Expression of AQP1 in cell lines was assessed by quantitative (q) PCR, western blot and immunofluorescence, while expression of AQP1 in human colon tumour tissue was assessed by immunohistochemistry. The effect of varying concentrations of the AQP1 inhibitor AqB013 was tested on human colon cancer cell lines expressing high versus low levels of AQP1, using wound closure (migration) assays, matrigel invasion assays, and proliferation assays. The effect of AqB013 on angiogenesis was tested using an endothelial cell tube-formation assay. HT29 colon cancer cells with high AQP1 levels showed significant inhibition of migration compared to vehicle control of 27.9 % ± 2.6 % (p < 0.0001) and 41.2 % ± 2.7 (p <0.0001) treated with 160 or 320 μM AqB013 respectively, whereas there was no effect on migration of HCT-116 cells with low AQP1 expression. In an invasion assay, HT29 cells treated with 160 μM of AqB013, showed a 60.3 % ± 8.5 % decrease in invasion at 144 hours (p < 0.0001) and significantly decreased rate of invasion compared with the vehicle control (F-test, p = 0.001). Almost complete inhibition of endothelial tube formation (angiogenesis assay) was achieved at 80 μM AqB013 compared to vehicle control (p < 0.0001). These data provide good evidence for further testing of the inhibitor as a therapeutic agent in colon cancer. The online version of this article (doi:10.1186/s13046-016-0310-6) contains supplementary material, which is available to authorized users.