The landscape of drug resistant CDK4/6 mutants

These saturating mutagenesis screens of CDK4 and CDK6 resistance to palbociclib were part of a larger study defining the landscape of ATP-competitive inhibitor resistance residues in protein kinases. We utilized the structural conservation of protein kinase domains, the similar mechanism of action for ATP-competitive kinase inhibitors, and a systematic comparison of mutagenesis assays to prospectively identify common sites of drug resistance in kinases. To do this we perform hotspot analysis of 8 different mutagenesis screens for resistance to ATP-competitive inhibitors. Two screens, identifying CDK6 and CDK4 resistance to palbociclib, are novel saturating mutagenesis screens presented here for the first time and included in this submission; two screens are saturating mutagenesis screens previously reported by our group; and 4 screens are sub-saturating datasets obtained from the literature. Using this approach we identified multiple sites that drive drug resistance when mutated and validate them in an extended panel of kinases. Unexpectedly, we uncovered a common position where mutation leads to activation, and subsequently drug resistance in protein kinases. In the future, comparative mutagenic approaches such as those described here may be used to improve drug efficacy, support biological discovery of kinase targets, and potentially may help identify activating or drug-resistant mutants that arise in disease.

针对细胞周期蛋白依赖性激酶4（CDK4）与细胞周期蛋白依赖性激酶6（CDK6）对帕博西尼（palbociclib）的耐药性开展的饱和诱变筛选，属于一项旨在阐明蛋白激酶中ATP竞争性抑制剂耐药残基分布特征的更大规模研究的组成部分。我们依托蛋白激酶结构域的保守性、ATP竞争性激酶抑制剂相似的作用机制，以及对诱变实验的系统性对比，前瞻性地鉴定出激酶中常见的耐药位点。为此，我们针对8组针对ATP竞争性抑制剂耐药性的不同诱变筛选开展热点分析。其中，用于鉴定CDK4与CDK6对帕博西尼耐药性的2组筛选为本次首次公开的新型饱和诱变筛选，已纳入本提交数据集；另外2组为本团队此前报道的饱和诱变筛选；剩余4组为从公开文献中获取的亚饱和数据集。通过该研究策略，我们鉴定出多个突变后可介导药物耐药性的位点，并在扩展的激酶检测面板中对这些位点进行了验证。出乎意料的是，我们发现了一类共通位点：该位点发生突变后可激活蛋白激酶，进而引发药物耐药性。未来，类似本文所述的比较诱变方法可用于提升药物疗效、助力激酶靶点的生物学研究，还有望帮助识别疾病中出现的激活突变或耐药突变体。