Insulin resistance is a pathophysiological hallmark of type 2 diabetes and nonalcoholic fatty liver disease. Under the condition of fat accumulation in the liver, suppression of hepatic glucose production by insulin is diminished. In order to gain deeper understanding of dysregulation of glucose production in metabolic diseases, in the present study, we performed an unbiased phenotypic screening in primary human hepatocytes to discover novel mechanisms that regulate gluconeogenesis in the presence of insulin. To optimize phenotypic screening process, we used a chemical genetic screening approach by building a small-molecule library with prior knowledge of activity-based protein profiling. The “positive hits” result from the screen will be small molecules with known protein targets. This makes downstream deconvolution process (i.e., determining the relevant biological targets) less time-consuming. To unbiasedly decipher the molecular targets, we developed a novel statistical method and discovered a set of genes, including DDR3 and CACNA1E that suppressed gluconeogenesis in human hepatocytes. Further investigation, including transcriptional profiling and gene network analysis, was performed to understand the molecular functions of DRD3 and CACNA1E in human hepatocytes.

胰岛素抵抗是2型糖尿病和非酒精性脂肪肝疾病的病理生理指标。在肝脏中脂肪蓄积的情况下，胰岛素对肝葡萄糖生成的抑制作用减弱。为了更深入地了解代谢疾病中葡萄糖生成的失调，在本研究中，我们在原代人肝细胞中进行了无偏见的表型筛选，以发现在胰岛素存在下调节糖异生的新机制。为了优化表型筛选过程，我们使用化学遗传筛选方法通过建立具有基于活动的蛋白质谱分析的先验知识的小分子文库。筛选产生的“阳性结果”将是具有已知蛋白质靶标的小分子。这使得下游反卷积过程（即 确定相关的生物学指标）耗时少。为了公正地破译分子靶标，我们开发了一种新颖的统计方法并发现了一组抑制人肝细胞糖异生的基因，包括DDR3和CACNA1E。进行了进一步的研究，包括转录谱分析和基因网络分析，以了解DRD3和CACNA1E在人肝细胞中的分子功能。