Division of Endocrinology and Diabetology, Department of Internal Medicine, University Hospital of Wuerzburg, Würzburg, Germany.
Institute of Human Genetics, University of Wuerzburg, Würzburg, Germany.
Front Endocrinol (Lausanne). 2020 Apr 16;11:219. doi: 10.3389/fendo.2020.00219. eCollection 2020.
Adrenocortical carcinomas (ACC) are aggressive tumors with a heterogeneous prognosis and limited therapeutic options for advanced stages. This study aims to identify novel drug targets for a personalized treatment in ACC. RNA was isolated from 40 formalin-fixed paraffin-embedded ACC samples. We evaluated gene expression of 84 known cancer drug targets by reverse transcriptase quantitative real time-PCR and calculated fold change using 5 normal adrenal glands as reference (overexpression by fold change >2.0). The most promising candidate cyclin-dependent kinase 4 (CDK4) was investigated at protein level in 104 ACC samples and tested by experiments in two ACC cell lines (NCI-H295R and MUC1). The most frequently overexpressed genes were (100% of cases, median fold change = 16.5), (95%, fold change = 52.9), (80%, fold change = 6.7) (62%, fold change = 2.6) (60%, fold change = 2.8), and (52%, fold change = 2.3). CDK4 was chosen for functional validation, as it is actionable by approved CDK4/6-inhibitors (e.g., palbociclib). Nuclear immunostaining of CDK4 significantly correlated with mRNA expression (R = 0.52, < 0.005). We exposed both NCI-H295R and MUC1 cell lines to palbociclib and found a concentration- and time-dependent reduction of cell viability, which was more pronounced in the NCI-H295R cells in line with higher CDK4 expression. Furthermore, we tested palbociclib in combination with insulin-like growth factor 1/insulin receptor inhibitor linsitinib showing an additive effect. In conclusion, we demonstrate that RNA profiling is useful to discover potential drug targets and that CDK4/6 inhibitors are promising candidates for treatment of selected patients with ACC.
肾上腺皮质癌（ACC）是一种侵袭性肿瘤，具有异质性的预后，且晚期阶段的治疗选择有限。本研究旨在为 ACC 患者的个体化治疗寻找新的药物靶点。
从 40 例福尔马林固定石蜡包埋的 ACC 样本中提取 RNA。我们通过逆转录定量实时 PCR 评估了 84 种已知癌症药物靶点的基因表达，并使用 5 个正常肾上腺作为参考计算了 fold change（fold change >2.0 表示过表达）。在 104 例 ACC 样本中对最有前途的候选基因 cyclin-dependent kinase 4（CDK4）进行了蛋白水平检测，并在 2 个 ACC 细胞系（NCI-H295R 和 MUC1）中进行了实验测试。
过表达最频繁的基因是 （100%的病例，中位数 fold change = 16.5）、 （95%，fold change = 52.9）、 （80%，fold change = 6.7）、 （62%，fold change = 2.6）和 （60%，fold change = 2.8），以及 （52%，fold change = 2.3）。由于 CDK4 可被批准的 CDK4/6 抑制剂（如 palbociclib）靶向作用，因此选择其进行功能验证。CDK4 的核免疫染色与 mRNA 表达显著相关（R = 0.52， < 0.005）。我们将 palbociclib 暴露于 NCI-H295R 和 MUC1 细胞系中，发现细胞活力呈浓度和时间依赖性下降，在 CDK4 表达较高的 NCI-H295R 细胞中更为明显。此外，我们还测试了 palbociclib 与胰岛素样生长因子 1/胰岛素受体抑制剂 linsitinib 联合使用，显示出相加作用。
总之，我们证明 RNA 谱分析可用于发现潜在的药物靶点，CDK4/6 抑制剂是治疗选定 ACC 患者的有前途的候选药物。
