Plos One：季红斌等揭示K-ras突变体对肺癌发生及存活的作用

5月11日，Plos One在线发表了中科院上海生命科学研究院生化与细胞所季红斌研究组、刘新垣研究组与美国哈佛大学Dr. Kwok-Kin Wong合作的最新研究成果Temporal Dissection of K-rasG12D Mutant In Vitro and In Vivo Using a Regulatable K-rasG12D Mouse Allele，揭示了K-ras突变体在肺癌发生、发展及存活中的重要作用。

肺癌是严重危害我国人民生命健康的重大疾病之一，揭示其中关键的致病基因在肺癌发生发展及存活中的作用将为临床上肺癌的“个体化”分子靶向治疗提供潜在的药靶和新的策略；而可时空调控基因表达的小鼠模型能够更好地促进肺癌发病分子机理的研究。目前国际上应用最广泛的肺癌动物模型就是LSL-K-rasG12D小鼠模型，可以通过鼻腔滴入携带Cre表达基因的腺病毒或与肺上皮细胞特异性的Cre转基因小鼠杂交来实现K-ras突变体的激活，从而导致肺癌的发生；而该模型唯一的缺点就是K-ras突变体一旦激活，就无法调控其表达和活性。

最近，季红斌研究组、刘新垣研究组等在LSL-K-rasG12D小鼠模型基础上，构建了一个可时空调控K-ras突变体表达的小鼠模型LSL-ER-K-rasG12D；该小鼠的ER-KrasG12D的表达可被CRE诱导，而ER-K-rasG12D蛋白的激活可通过Tamoxifen来调控。在小鼠胚胎成纤维细胞中的研究表明，Tamoxifen处理可以诱导ER-K-rasG12D的激活，促进细胞异常增殖、恶性转化以及侵袭和浸润；而Tamoxifen去除后，小鼠胚胎成纤维细胞增殖、恶性转化及侵袭能力可基本恢复到正常水平；动物体内研究发现，通过腹腔注射小鼠Tamoxifen可持续激活K-rasG12D并促进小鼠肺部肿瘤的早期发生，而Tamoxifen撤掉后大部分的肺部肿瘤会发生细胞凋亡。

该工作在很大程度上提高了人们对K-ras突变体在肺癌发生、发展及存活中作用的认识，并为将来深入研究肺癌发病分子机理提供一个较为理想的研究体系，对新型肺癌小鼠模型的建立和发展具有重要意义。

该研究课题获得科技部、国家自然科学基金委和上海市科委的经费支持。（生物谷Bioon.com）

doi:10.1371/journal.pone.0037308
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Temporal Dissection of K-rasG12D Mutant In Vitro and In Vivo Using a Regulatable K-rasG12D Mouse Allele

Zuoyun Wang1#*, Yan Feng1#*, Nabeel Bardessy2, Kwok-Kin Wong3*, Xin-Yuan Liu1*, Hongbin Ji

Animal models which allow the temporal regulation of gene activities are valuable for dissecting gene function in tumorigenesis. Here we have constructed a conditional inducible estrogen receptor-K-rasG12D (ER-K-rasG12D) knock-in mice allele that allows us to temporally switch on or off the activity of K-ras oncogenic mutant through tamoxifen administration. In vitro studies using mice embryonic fibroblast (MEF) showed that a dose of tamoxifen at 0.05 μM works optimally for activation of ER-K-rasG12D independent of the gender status. Furthermore, tamoxifen-inducible activation of K-rasG12D promotes cell proliferation, anchor-independent growth, transformation as well as invasion, potentially via activation of downstream MAPK pathway and cell cycle progression. Continuous activation of K-rasG12D in vivo by tamoxifen treatment is sufficient to drive the neoplastic transformation of normal lung epithelial cells in mice. Tamoxifen withdrawal after the tumor formation results in apoptosis and tumor regression in mouse lungs. Taken together, these data have convincingly demonstrated that K-ras mutant is essential for neoplastic transformation and this animal model may provide an ideal platform for further detailed characterization of the role of K-ras oncogenic mutant during different stages of lung tumorigenesis.