Warren Janine S A, Feustel Paul J, Lamar John M
Department of Molecular and Cellular Physiology, Albany Medical College.
Department of Neuroscience and Experimental Therapeutics, Albany Medical College.
J Vis Exp. 2019 Dec 19(154). doi: 10.3791/60687.
Metastasis is the main cause of cancer-related deaths and there are limited therapeutic options for patients with metastatic disease. The identification and testing of novel therapeutic targets that will facilitate the development of better treatments for metastatic disease requires preclinical in vivo models. Demonstrated here is a syngeneic mouse model for assaying breast cancer metastatic colonization and subsequent growth. Metastatic cancer cells are stably transduced with viral vectors encoding firefly luciferase and ZsGreen proteins. Candidate genes are then stably manipulated in luciferase/ZsGreen-expressing cancer cells and then the cells are injected into mice via the lateral tail vein to assay metastatic colonization and growth. An in vivo imaging device is then used to measure the bioluminescence or fluorescence of the tumor cells in the living animals to quantify changes in metastatic burden over time. The expression of the fluorescent protein allows the number and size of metastases in the lungs to be quantified at the end of the experiment without the need for sectioning or histological staining. This approach offers a relatively quick and easy way to test the role of candidate genes in metastatic colonization and growth, and provides a great deal more information than traditional tail vein metastasis assays. Using this approach, we show that simultaneous knockdown of Yes associated protein (YAP) and transcriptional co-activator with a PDZ-binding motif (TAZ) in breast cancer cells leads to reduced metastatic burden in the lungs and that this reduced burden is the result of significantly impaired metastatic colonization and reduced growth of metastases.
转移是癌症相关死亡的主要原因，对于转移性疾病患者，治疗选择有限。要确定和测试有助于开发更好的转移性疾病治疗方法的新治疗靶点，需要临床前体内模型。本文展示了一种用于检测乳腺癌转移定植和后续生长的同基因小鼠模型。用编码萤火虫荧光素酶和ZsGreen蛋白的病毒载体稳定转导转移性癌细胞。然后在表达荧光素酶/ZsGreen的癌细胞中稳定操纵候选基因，接着通过侧尾静脉将细胞注射到小鼠体内，以检测转移定植和生长情况。然后使用体内成像设备测量活体动物体内肿瘤细胞的生物发光或荧光，以量化转移负荷随时间的变化。荧光蛋白的表达使得在实验结束时无需切片或组织学染色就能对肺内转移灶的数量和大小进行量化。这种方法提供了一种相对快速简便的方式来测试候选基因在转移定植和生长中的作用，并且比传统的尾静脉转移试验提供了更多信息。使用这种方法，我们发现乳腺癌细胞中Yes相关蛋白（YAP）和具有PDZ结合基序的转录共激活因子（TAZ）的同时敲低会导致肺内转移负荷降低，并且这种降低的负荷是转移定植显著受损和转移灶生长减少的结果。
