Fosbretabulin Disodium (CA 4P), a water-soluble prodrug of Combretastatin A4 (CA4), is a microtubule-targeting agent that binds β-tubulin (Kd: 0.4 μM). Fosbretabulin Disodium(Combretastatin A4 disodium phosphate) inhibits the polymerization of tubulin (IC50: 2.4 μM), and also disrupts tumor vasculature.

Tubulin:2.4 μM

Fosbretabulin disodium（Combretastatin A-4 phosphate disodium, CA4P disodium）是可溶于水的前药，源自非洲树木Combretum caffrum的combretastatin A4（CA4）。CA4作为一种结合微管蛋白的剂量，在β-微管蛋白的秋水仙素结合位点或其附近结合（Kd = 0.40 μM），以IC50为2.4 μM抑制微管蛋白组装。[1] CA4对增殖中而非静止的内皮细胞具有细胞毒性，对肿瘤血管具有强大且选择性的毒性。[2] CA4P（1 mM, 30分钟）破坏了内皮细胞的微管骨架并影响其形态，通过Rho/Rho激酶路径引发细胞形态改变，促进肌动蛋白应力纤维的形成及膜泡化，增加细胞层的渗透性。[3] CA4P通过打断VE-cadherin/β-catenin/Akt信号路径主要抑制内皮细胞的迁移和毛细血管管形成，从而增加内皮细胞的渗透性，并导致快速的血管崩溃和肿瘤坏死。[4]

CA4P在实验性肿瘤模型中，通过单次给药，剂量为最大耐受剂量(MTD)的10%，可迅速、广泛且不可逆地导致血管关闭。CA4P在化合物给药后6小时内，可使血管体积减少93%。[2] CA4P（100 mg/kg，给药后6小时）使肿瘤血流量大约减少了100倍，而脾脏的血流量大约减少了7倍。[5]

Tubulin assembly-disassembly: The assembly of microtubules from isolated tubulin is carried out spectrophotometrically at 350 nm and utilises the increase in turbidity which is associated with microtubule formation. Assembly is initiated by temperature increase from 10 to 35 °C. The effect of drugs on the increase in light absorption is carried. Drugs are dissolved in DMSO (<4%), which does not affect control assembly

For the proliferation assay, the minimal concentration of FBS (1%) diluted in X-VIVO medium is used to allow sufficient viability of endothelial cells. After detachment, the cells are seeded at a concentration of 2×104 HUVECs in each well of 24-well plates, allowed to adhere overnight, and then incubated with or without cytokines (5 ng/ml FGF-2 or 5 ng/ml VEGF-A). CA4P is added at 0 – 50 nM. After incubation for 12, 24, 36, and 48 hours, cells are detached by trypsin/EDTA and manually counted using trypan blue exclusion. (Only for Reference)