Dana-Farber Cancer Institute, Boston, Massachusetts, USA.
Clin Cancer Res. 2013 Jun 1;19(11):2929-40. doi: 10.1158/1078-0432.CCR-12-2307. Epub 2013 Feb 12.
Effective therapies for KRAS-mutant colorectal cancer (CRC) are a critical unmet clinical need. Previously, we described genetically engineered mouse models (GEMM) for sporadic Kras-mutant and non-mutant CRC suitable for preclinical evaluation of experimental therapeutics. To accelerate drug discovery and validation, we sought to derive low-passage cell lines from GEMM Kras-mutant and wild-type tumors for in vitro screening and transplantation into the native colonic environment of immunocompetent mice for in vivo validation.
Cell lines were derived from Kras-mutant and non-mutant GEMM tumors under defined media conditions. Growth kinetics, phosphoproteomes, transcriptomes, drug sensitivity, and metabolism were examined. Cell lines were implanted in mice and monitored for in vivo tumor analysis.
Kras-mutant cell lines displayed increased proliferation, mitogen-activated protein kinase signaling, and phosphoinositide-3 kinase signaling. Microarray analysis identified significant overlap with human CRC-related gene signatures, including KRAS-mutant and metastatic CRC. Further analyses revealed enrichment for numerous disease-relevant biologic pathways, including glucose metabolism. Functional assessment in vitro and in vivo validated this finding and highlighted the dependence of Kras-mutant CRC on oncogenic signaling and on aerobic glycolysis.
We have successfully characterized a novel GEMM-derived orthotopic transplant model of human KRAS-mutant CRC. This approach combines in vitro screening capability using low-passage cell lines that recapitulate human CRC and potential for rapid in vivo validation using cell line-derived tumors that develop in the colonic microenvironment of immunocompetent animals. Taken together, this platform is a clear advancement in preclinical CRC models for comprehensive drug discovery and validation efforts.
KRAS 突变型结直肠癌(CRC)的有效疗法是一个关键的未满足的临床需求。此前,我们描述了适合于实验治疗药物的临床前评估的散发性 Kras 突变和非突变 CRC 的基因工程小鼠模型(GEMM)。为了加速药物发现和验证,我们试图从 GEMM Kras 突变和野生型肿瘤中获得低传代细胞系,用于体外筛选,并移植到免疫活性小鼠的天然结肠环境中进行体内验证。
在定义的培养基条件下,从 Kras 突变和非突变 GEMM 肿瘤中获得细胞系。检查了生长动力学、磷酸化蛋白质组、转录组、药物敏感性和代谢。将细胞系植入小鼠中,并监测体内肿瘤分析。
Kras 突变细胞系显示出增加的增殖、有丝分裂原激活的蛋白激酶信号和磷酸肌醇-3 激酶信号。微阵列分析确定与人类 CRC 相关基因标记有显著重叠,包括 KRAS 突变和转移性 CRC。进一步的分析揭示了许多与疾病相关的生物学途径的富集,包括葡萄糖代谢。体外和体内的功能评估验证了这一发现,并强调了 Kras 突变型 CRC 对致癌信号和有氧糖酵解的依赖性。
我们已经成功地描述了一种新型的 GEMM 衍生的人 KRAS 突变型 CRC 原位移植模型。这种方法结合了使用低传代细胞系进行体外筛选的能力,这些细胞系重现了人类 CRC 的特征,并且具有使用在免疫活性动物的结肠微环境中发展的细胞系衍生肿瘤进行快速体内验证的潜力。总的来说,这个平台是 CRC 临床前模型的一个明显进步,用于全面的药物发现和验证工作。