Search for New Anti-angiogenic Agents Targeting on Endothelial Cell Growth or Motility.

以选择性抑制人脐静脉内皮细胞（HUVECs）生长为指导，从海洋生物提取物中寻找抗血管生成物质。溴化酪氨酸^1的环四聚体Bastadins从海绵Ianthella basta中分离得到。其中主要成分basadin 6对HUVECs表现出选择性生长抑制作用。发现Bastadin 6抑制VEGF或bfgf依赖性HUVECs的增殖（IC_<50>=0.052μM），选择性是正常成纤维细胞（3Y1）或几种肿瘤细胞（KB3-1， K562和Neuro2A）的20至100倍。Bastadin 6还能抑制VEGF或bfgf诱导的huvec小管形成（0.1μM，处理6h）和VEGF诱导的huvec迁移（1μM，处理4h）。此外，bastadin 6在小鼠角膜实验中几乎完全阻断vegf或bfgf诱导的体内新生血管形成，并抑制裸小鼠皮下接种A431实体瘤的生长（100mg/kg, i.p）。Bastadin 6诱导huvec细胞凋亡，而对VEGF诱导的VEGF受体Flt1和KDR/Flk-1的自磷酸化无影响。这些证据表明，bastadin 6的抗血管生成作用与选择性诱导内皮细胞凋亡活性密切相关。我们研究了巴斯丁素6的全合成方法，以期为进一步的生物学分析建立一种实用的合成方法。根据报告的方法。以溴酚的酶促氧化偶联为关键反应，通过改变原料和反应条件，改进了合成路线。我们还开发了一种新的Ce(NH_4)_2(NO_3)_6 （CAN）介导的2,6-二溴酚氧化偶联反应，得到了二芳基醚衍生物。两个片段的缩合和随后的分子内大环化在9步中得到巴斯丁6，总收率为26%。为了分析basadin抗血管生成活性的结构要求，对basadin 6的构效关系进行了研究。研究发现，肟部分的保护导致HUVEC的生长抑制活性完全丧失，非肟类似物的活性也明显减弱。此外，脱溴类似物的活性较弱。少

On the guidance of selective growth inhibition against human umbilical vein endothelial cells (HUVECs), we searched for anti-angiogenic substances from extracts of marine organisms.Bastadins, cyclic tetramers of brominated-tyrosine^1, were isolated from the marine sponge Ianthella basta. Among of them, bastadin 6, a major constituent, showed a selective growth inhibition against HUVECs. Bastadin 6 was found to inhibit VEGF- or bFGF-dependent proliferation (IC_<50>=0.052μM) of HUVECs, 20 to 100-fold selectively in comparison with normal fibroblast (3Y1) or several tumor cells (KB3-1, K562, and Neuro2A). Bastadin 6 also inhibited VEGF- or bFGF-induced tubular formation (0.1μM, 6h treatment) and VEGF-induced migration (1μM, 4h treatment) of HUVECs. Moreover, bastadin 6 almost completely blocked VEGF-or bFGF-induced in vivo neovascularization in mice corneal assay and suppressed growth of subcutaneously inoculated A431 solid tumor in nude mice (100mg/kg, i.p.). Bastadin 6 induced cell deat … More h of HUVECs with apoptotic phenotype, whereas it showed no effect on the VEGF-induced auto-phosphorylation of VEGF receptors, Flt1 and KDR/Flk-1. These evidences suggested that the anti-angiogenic effect of bastadin 6 is closely related to selective induction activity of apoptosis against endothelial cells.We studied total synthesis of bastadin 6 in order to establish a practical synthetic method for further biological analysis. According to the reported method., utilizing the enzymatic oxidative coupling of bromophenols as the key reaction, we developed an improved synthetic route by changing starting materials and reaction conditions. We also developed a novel Ce(NH_4)_2(NO_3)_6 (CAN)-mediated oxidative coupling reaction of 2,6-dibromophenols to give the diaryl ether derivatives. Condensation of two segments and subsequent intramolecular macrocyclization gave bastadin 6 in 9 steps, 26% overall yield.In order to analyze the structure-requirement for the anti-angiogenic activity of bastadin, structure-activity relationship of bastadin 6 was studied. Protection of oxime moiety was found to cause a complete loss of the growth inhibitory activity of HUVEC, and non-oxime analog also showed a significantly weak activity. Furthermore, debrominated analog showed a weak activity. Less