The Oncogenic and Tumor Suppressor Functions of the Kras isoform 4A in vivo

描述(由申请人提供)：KRAS的激活突变经常发生在一些最常见和最致命的人类癌症中，但针对KRAS用于治疗目的的努力尚未成功。KRAS通过选择性剪接产生两种高度同源的异构体(KRAS4A和KRAS4B)。虽然在小鼠中完全敲除Kras基因会导致胚胎死亡，但敲入Kras4A(Kras4AKI)或Kras4B(Kras4BKI)会导致替代亚型的损失，从而导致存活的小鼠。Kras4AKI小鼠只表达Kras4A，Kras4BKI小鼠只表达Kras4B，这使得第一次能够比较这些Kras亚型在体内转化中的单独作用。我们发现Kras4BKI小鼠对Kras突变的肺和皮肤肿瘤具有高度的抵抗力，这表明Kras4A对于两个独立组织的转化是必不可少的。我们的研究进一步表明，Kras4A也与先前报道的野生型Kras的肿瘤抑制活性有关。这项建议的总体目标是利用这些新的小鼠模型来研究Kras4A和Kras4B在肺癌发展中的功能，既作为癌基因(突变形式)，也作为肿瘤抑制基因(野生型)。这一建议的一个主要优点是可获得唯一表达每一种亚型的活菌株，代表了用于研究这一主要人类癌基因的亚型特定功能的强大和互补的体内系统。该方案的目的1将利用致癌物诱导的肺癌模型，在体内研究单个Kras亚型的致癌功能。目的研究野生型Kras4A和Kras4B在条件激活突变型Kras(即KrasLA2和KrasLSL-G12D)模型中的体内抑瘤活性。AIM 3将利用来自不同小鼠模型的肺上皮细胞系在体外详细分析Kras4A和Kras4B信号通路，以获得对其致癌活性的机制见解。目的4将评估这些单独的KRAS亚型在人KRAS突变肺癌细胞生长/维持中的作用。对KRAS异构体特定功能的洞察不仅对理解癌症的发展很重要，而且可能对治疗设计产生重大影响。

DESCRIPTION (provided by applicant): Activating mutations in KRAS occur frequently in some of the most common and deadliest of human cancers but efforts to target KRAS for therapeutic purposes have not been successful. KRAS generates two highly homologous isoforms (KRAS4A and KRAS4B) as a result of alternative splicing. While the complete Kras gene knockout in mice causes embryonic lethality, knock-in of either Kras4A (Kras4AKI) or Kras4B (Kras4BKI) at the expense of the alternate isoform results in viable mice. Kras4AKI mice express only Kras4A and Kras4BKI mice express only Kras4B, enabling for the first time a comparison of the individual roles of these Kras isoforms in transformation in vivo. We have found that Kras4BKI mice are highly resistant to the development Kras mutant lung and skin tumors, suggesting that Kras4A is essential for transformation in 2 independent tissues. Our studies further showed that Kras4A is also responsible for the previously reported tumor suppressor activity of wild type Kras. The overall goal of this proposal is to exploit these novel mouse models to study the functions of both Kras4A and Kras4B in lung cancer development, both as oncogenes (mutant form) and as tumor suppressors (wild type form). A major strength of this proposal is the availability of viable strains uniquely expressing each isoform, representing powerful and complementary in vivo systems for studies into the isoform specific functions of this major human oncogene. Aim 1 of this proposal will investigate the oncogenic function of individual Kras isoforms in vivo using models of carcinogen induced lung carcinogenesis. Aim 2 will investigate the tumor suppressor activity of wild type Kras4A and Kras4B in vivo in models of conditionally activatable mutant Kras (i.e. KrasLA2 and KrasLSL-G12D). Aim 3 will take advantage of lung epithelial cell lines derived from the various mouse models for detailed in vitro analyses of Kras4A and Kras4B signaling pathways in order to gain mechanistic insights into their oncogenic activities. Aim 4 will evaluate the roles of these individual KRAS isoforms in the growth/maintenance of human KRAS mutant lung cancer cells. Insights into the isoform specific functions of KRAS will not only be important to the understanding of cancer development but could potentially have major implications for therapeutic design.