Division of Gene Regulation, Institute for Advanced Medical Research, School of Medicine, Keio University, Tokyo, Japan.
PLoS One. 2013;8(1):e54424. doi: 10.1371/journal.pone.0054424. Epub 2013 Jan 14.
Most targeted anticancer drugs have been identified by screening at the molecular or cellular level in vitro. However, many compounds selected by such costly and time-consuming screening do not prove effective against tumors in vivo. The development of anticancer drugs would thus be facilitated by the availability of an in vivo screening system based on a multicellular organism. We have now established a transgenic line of the freshwater fish medaka in which melanophores (melanocytes) proliferate in a manner dependent on heat shock-induced signaling by a human RAS oncoprotein. The human HRAS(G12V) oncogene was expressed under the control of a melanophore-specific gene promoter in order to allow visualization of tumor growth in live fish maintained in a water tank. The expression of HRAS(G12V) was induced as a result of Cre-mediated recombination by exposure of the fish to a temperature of 37°C for 30 min, given that the Cre gene was placed under the control of a medaka heat shock promoter. One of the stable transgenic lines developed abnormal pigment cell proliferation in the eyes and epidermis with 100% penetrance by 6 months postfertilization. Sorafenib, an inhibitor of RAS signaling, was administered to the transgenic fish and was found both to reduce the extent of melanophore proliferation and to improve survival. The transgenic medaka established here thus represents a promising in vivo system with which to screen potential anticancer drugs that target RAS signaling, and this system can readily be adapted for the screening of agents that target other oncogenes.
大多数靶向抗癌药物是通过在体外的分子或细胞水平筛选确定的。然而，通过这种昂贵且耗时的筛选选择的许多化合物在体内对肿瘤并不有效。因此，如果有一种基于多细胞生物的体内筛选系统，抗癌药物的开发将会得到促进。我们现在已经建立了一种转基因淡水鱼斑马鱼系，其中黑色素细胞（melanocytes）的增殖依赖于人类 RAS 癌蛋白诱导的热休克信号。为了能够在水箱中维持的活体鱼中观察肿瘤生长，人 HRAS(G12V) 癌基因在黑色素细胞特异性基因启动子的控制下表达。通过将 Cre 基因置于斑马鱼热休克启动子的控制下，将鱼暴露在 37°C 30 分钟以进行 Cre 介导的重组，从而诱导 HRAS(G12V) 的表达。其中一条稳定的转基因系在受精后 6 个月时，眼睛和表皮中的黑色素细胞出现异常增殖，具有 100%的外显率。RAS 信号抑制剂索拉非尼(sorafenib)被给予转基因鱼，发现它既可以减少黑色素细胞的增殖程度，又可以提高存活率。因此，这里建立的转基因斑马鱼代表了一种很有前途的体内系统，可以筛选靶向 RAS 信号的潜在抗癌药物，并且该系统可以很容易地适应筛选靶向其他癌基因的药物。
