Role of JNK Pathway in Lung Tumorigenesis

描述（由申请人提供）：K-Ras和突变/过表达的EGFR家族成员是非小细胞肺癌（NSCLC）的显性癌基因。然而，介导这些癌基因转化的信号通路仍然不明确。此外，癌基因同时刺激促肿瘤和抗肿瘤信号。在这方面，jnk1、jnk2和jnk3编码的JNK MAPKs被生长因子和癌基因激活，文献记载了JNK在细胞转化中的积极和消极作用。MKK4是JNKs的双特异性激酶激活剂，已在多种人类癌症中作为肿瘤转移抑制因子出现。与这一发现一致，我们对JNK1和jnk2缺陷小鼠的实验显示，与野生型小鼠相比，致癌物诱导的肺肿瘤发生增加。此外，与未转化的肺上皮细胞相比，多种人类非小细胞肺癌细胞系显示JNK活性降低，转染的功能获得性JNK1抑制非锚定依赖性非小细胞肺癌生长。因此，我们提出JNK1和JNK2在肺癌中作为肿瘤抑制通路的组成部分起作用。相比之下，对JNK3缺陷小鼠的初步研究显示，致癌物诱导的肺肿瘤发生减少。此外，JNKS mRNA和蛋白在NSCLC细胞系和原发肿瘤中相对于未转化的肺上皮细胞或未累及的肺组织表达。因此，我们假设JNK3在肺癌进展过程中被诱导，并代表一种促肿瘤的JNK亚型。为了验证这些假设，我们将完成以下具体目标。目的1：确定特异性JNKs在缺乏jnk1、jnk2或jnk3小鼠肺肿瘤发生中的体内作用。特异性JNKs作为宿主肺微环境调节肺肿瘤发生的信号成分的作用也将被测试。目的2：在体外测试特异性JNKs在肿瘤抑制或细胞转化中的作用。表达JNKs分子抑制剂的未转化肺上皮细胞系将被致癌的K-Ras转导，并测量转化、分化和凋亡的标准。此外，用功能获得性JNKs或显性阴性JNKs转导的NSCLC细胞将被监测细胞转化的标准。目的3：明确NSCLC细胞系中JNK1和JNK2活性降低的机制。MKPs作为癌基因诱导的JNK1和JNK2活性负调控因子的作用将被强调。目的4：通过免疫组化和定量RT-PCR，在存档的原发性人肺肿瘤中测量JNKs的激活状态和Aims 1-3中定义的特定信号分子的表达状态，并与临床行为相关。完成这些特定目标将导致对MAP激酶家族在肺肿瘤发生中的复杂作用的全面理解，并为探索这些激酶在其他人类癌症中的作用提供一种格式。此外，对JNK MAP激酶在肺癌中的多方面作用的详细了解，对于这一途径对抗肺癌的理论基础和精确的治疗靶向是绝对必要的。

DESCRIPTION (provided by applicant): K-Ras and mutated/over-expressed EGFR family members function as dominant oncogenes in non-small cell lung cancer (NSCLC). Yet, the signal pathways that mediate transformation by these oncogenes remain ill-defined. Moreover, oncogenes stimulate both pro- and anti-tumorigenic signaling. In this regard, the JNK MAPKs, encoded by jnk1, jnk2 and jnk3, are activated by growth factors and oncogenes and the literature documents both positive and negative roles for JNKs in cellular transformation. MKK4, a dual-specificity kinase activator of the JNKs, has emerged as a tumor metastasis suppressor in diverse human cancers. Consistent with this finding, our experiments with JNK1 and JNK2-deficient mice reveal increased carcinogen-induced lung tumorigenesis relative to wild-type littermates. Also, multiple human NSCLC cell lines show decreased JNK activity relative to non-transformed lung epithelial cells and transfected gain-of-function JNK1 inhibits anchorage-independent NSCLC growth. Thus, we propose that JNK1 and JNK2 function as components of a tumor suppressor pathway in lung cancer. By contrast, preliminary studies with JNK3- deficient mice reveal decreased carcinogen-induced lung tumorigenesis. In addition, JNKS mRNA and protein is expressed in NSCLC cell lines and primary tumors relative to non-transformed lung epithelial cells or uninvolved lung tissue. Thus, we hypothesize that JNK3 is induced during lung cancer progression and represents a pro-tumorigenic JNK isoform. To test these hypotheses, we will complete the following specific aims. Aim 1: Determine the in vivo role for specific JNKs in murine lung tumorigenesis with mice lacking jnk1, jnk2 or jnk3. The role of specific JNKs as signal components of the host lung microenvironment regulating lung tumorigenesis will also be tested. Aim 2: Test the in vitro role of specific JNKs in tumor suppression or cellular transformation. Non-transformed lung epithelial cell lines expressing molecular inhibitors of the JNKs will be transduced with oncogenic K-Ras and criteria of transformation, differentiation and apoptosis will be measured. Also, NSCLC cells transduced with gain-of-function JNKs or dominant-negative JNKS will be monitored for criteria of cellular transformation. Aim 3: Define the mechanism(s) accounting for decreased JNK1 and JNK2 activity in NSCLC cell lines. The role of MKPs as oncogene-induced negative regulators of JNK1 and JNK2 activity will be highlighted. Aim 4: The activation state of JNKs and expression status of specific signaling molecules defined in Aims 1-3 will be measured in archived primary human lung tumors with IHC and quantitative RT-PCR and correlated with clinical behaviour. Completion of these specific aims will lead to a comprehensive understanding of the complex role of this family of MAP kinases in lung tumorigenesis and provide a format for exploration of the role of these kinases in other human cancers. Furthermore, a detailed understanding of the multi-faceted role of the JNK MAP kinases in lung cancer is absolutely required for rationale and precise therapeutic targeting of this pathway to combat lung cancer.