Studies of Alternations of c-MET in SCLC

描述（由申请人提供）：拟议项目将在Ravi Salgia博士的指导下进行，他在小细胞肺癌的酪氨酸激酶、细胞骨架和侵袭/转移方面拥有丰富的专业知识。我的长期职业目标是成为一名独立的临床科学家，专注于靶向治疗的转化癌症研究。我发现c-Met在SCLC中表达，功能性，有时突变。我有一个新的免疫组织化学发现，激活的磷酸化蛋氨酸优先定位在肿瘤浸润前沿。在最近完成的c-Met突变分析小细胞肺癌，我已经确定了新的选择性剪接的转录本和体细胞突变的c-Met，特别是在Sema域和质膜（JM）域。发现JM突变改变c-Met信号传导，增加酪氨酸磷酸化，包括关键的粘着斑蛋白桩蛋白。发现这与JM突变增强的肿瘤发生和细胞运动/迁移相关。这些结果表明JM结构域在c-Met信号传导中具有独特的作用，对细胞骨架功能具有显著影响。两种可用的特异性c-Met抑制剂（SU 11274和PHA 665752）的初步研究显示出对SCLC活力和运动性的有希望的抑制。我现在的目标是进一步确定各种c-Met突变在小细胞肺癌中的功能意义，如活力，存活/凋亡和转化。c-Met突变对下游PI 3 K/Akt信号传导途径的影响将通过PI 3 K-激酶测定、IP/IB以及PI 3 K抑制剂的组合来确定。将建立表达野生型或突变Met的细胞系并用于功能分析。将研究各种生物测定，如散射、运动/迁移和侵袭。利用磷酸特异性抗体对抗黏着斑蛋白，和延时视频显微镜，研究将集中在细胞骨架功能。c-Met及其突变体的各种抑制剂在SCLC中的作用将被进一步定义，旨在转化为新的靶向治疗剂。

DESCRIPTION (provided by applicant): The proposed project will be performed under the mentorship of Dr. Ravi Salgia, who has great expertise in the tyrosine kinases, cytoskeleton, and invasion/metastasis in SCLC. My long term career goal is to become an independent clinician-scientist in translational cancer research with focus on targeted therapeutics. I have found that c-Met is expressed, functional and sometimes mutated in SCLC. I have novel immunohistochemistry finding of the activated phospho-Met localized preferentially at the tumor invasive front. In the recently completed c-Met mutational analysis in SCLC, I have identified novel alternatively spliced transcripts and somatic mutations of c-Met, particularly in the Sema domain and the juxtamembrane (JM) domain. The JM-mutations were found to alter c-Met signaling with increased tyrosine phosphorylation including the key focal adhesion protein paxillin. This was found correlating with enhanced tumorigenesis and cell motility/migration by the JM mutations. These results suggest a unique role of the JM-domain in c-Met signaling with dramatic effects in cytoskeletal functions. Preliminary studies with two available specific c-Met inhibitors (SU11274 and PHA665752) have shown promising inhibition of viability and motility of SCLC. I now aim to further determine the functional implications of the various c-Met mutations in SCLC, such as viability, survival/ apoptosis, and transformation. Effects of the c-Met mutations on the downstream PI3K/Akt signaling pathway would be determined with combinations of PI3K-kinase assay, IP/IB, and also PI3K inhibitors. Cell lines expressing wild-type or mutated Met would be established and utilized in functional analyses. Various biological assays, such as scattering, motility/migration, and invasion would be investigated. Utilizing phosphospecific antibodies against focal adhesion proteins, and time-lapse video-microscopy, studies would be focused on cytoskeletal functions. The role of various inhibitors of c-Met and its mutants in SCLC would be further defined aiming to translate into novel targeted-therapeutics.