Modeling the KIF5B-RET Fusion Gene of Lung Cancer in Vivo

描述(申请人提供)：肺癌是世界上癌症死亡的主要原因。传统的以细胞毒为基础的化疗方案正处于其最大有效性的极限。新的治疗范例结合了针对特定驱动基因突变的抑制剂，如EGFR和ALK，已显示出更好的临床益处。这些新的治疗方法依赖于临床相关基因损伤的识别和评估，这些基因损伤驱动和维持着 恶毒。肺癌是一种异质性疾病。约50%的非小细胞肺癌(NSCLC)的临床相关突变仍有待鉴定和评估。可以预见，基因组技术的最新进展将加速发现人类癌症中新的基因变化。将新的基因组发现转化为改进的治疗方法的一个重要方面是拥有体内临床前模型，该模型忠实地复制人类疾病中新发现的基因，以评估药物反应和耐药性。KIF5B家族成员(KIF5B)与RET酪氨酸激酶(KIF5B-RET)之间的基因融合是在人类肺腺癌中发现的最新的基因突变。KIF5B-RET的临床前模型仅限于转基因的NIH3T3成纤维细胞和BA/F3前B淋巴细胞的异种移植。本研究的目的是通过Cre重组酶介导的盒交换(Cre-RMCE)技术建立多西环素诱导的、肺特异的KIF5B-RET转基因小鼠，以模拟KIF5B-RET在双转基因小鼠肺泡II型细胞中的致瘤活性。在具体目标1中，携带带有L2/L3loxP异质性位点和Cre的Teto-KIF5B-RET盒的质粒将被共同注射到来自现有Cre-RMCE转基因小鼠不同品系的受精卵(受精卵)中，以用Teto-KIF5B-RET取代原始的成群的Teto-SHP2转基因基因。在特定目的II中，将评估CCSP-RTTA/Teto-KIF5B-RET双转基因小鼠可诱导的KIF5B-RET的表达以及KIF5B-RET在肺癌发生和维持中的作用。由于我们的具有CRE-RMCE功能的SHP2转基因小鼠已经在肺中具有诱导表达和转基因功能的特征，因此从这些品系通过盒交换获得的新的转基因小鼠有望表现出可靠的可诱导转基因表达。这将加速产生和检验新的有用的转基因小鼠模型。因此，这项研究将产生针对KIF5B-RET分子亚型肺腺癌的新疗法开发急需的基因工程动物。此外，本研究将确定最有效的Cre-RMCE转基因株系(S)，该株系携带KIF5B-RET转基因基因的单一副本，用于后续研究以获得新的转基因小鼠。这将为研究界提供重要的资源，以加快新发现的分子损伤的小鼠模型的生成。

DESCRIPTION (provided by applicant): Lung cancer is a leading cause of cancer death in the world. Conventional cytotoxic-based chemotherapy regimens are at their limits of maximal effectiveness. New treatment paradigms incorporating inhibitors to specific driver mutations, such as EGFR and ALK, have shown improved clinical benefits. These new treatments depend on identification and evaluation of clinically relevant genetic lesions that drive and maintain the malignancy. Lung cancer is a heterogeneous disease. Clinically relevant mutations in ~50% of non-small cell lung cancer (NSCLC) remain to be identified and evaluated. It is envisioned that recent advances in genomic technologies will accelerate the discovery of new genetic alternations in human cancer. An important aspect in translating the new genomic findings into improved treatments is to have in vivo preclinical models that faithfully phenocopy the newly identified genes in the human diseases for evaluating drug response and resistance. Recurrent gene fusion between kinesin family member 5B (KIF5B) and the RET tyrosine kinase (KIF5B-RET) is the most recent genetic alternation identified in human lung adenocarcinoma. Preclinical models of KIF5B-RET have been limited to xenografts of transfected NIH3T3 fibroblasts and Ba/F3 pre-B- lymphocytes. The goal of this study is to generate doxycycline-inducible, lung-specific KIF5B-RET transgenic mice by Cre recombinase-mediated cassette exchange (Cre-RMCE) to model the oncogenic activity of KIF5B- RET in lung type II alveolar cells in bitransgenic mice. In Specific Aim 1, plasmids carrying a tetO-KIF5B-RET cassette flanked by heterospecific L2/L3 loxP sites and Cre will be co-injected into fertilized eggs (zygotes) derived from different lines of existing Cre-RMCE-capable transgenic mice to replace the original floxed tetO- SHP2 transgenes with tetO-KIF5B-RET. In Specific Aim II, inducible KIF5B-RET expression and effects of KIF5B-RET on lung tumor development and maintenance will be evaluated in CCSP-rtTA/tetO-KIF5B-RET bitransgenic mice. Since our Cre-RMCE-capable SHP2 transgenic mice have already been characterized for inducible expression and function of the transgene in the lung, new transgenic mice derived from these lines by cassette exchange are expected to display reliable inducible transgene expression. This will accelerate the generation and examination of novel useful transgenic mouse models. Thus, the study will generate genetically engineered animals urgently needed for development of new therapy targeting the KIF5B-RET molecular subtype of lung adenocarcinoma. Furthermore, the study will identify the most efficient Cre-RMCE- capable transgenic line(s) harboring a single copy of the KIF5B-RET transgene for subsequent studies to derive new transgenic mice. This will provide an important resource for the research community to accelerate the generation of mouse models of newly identified molecular lesions.