Table_2_Comparative kinase and cancer cell panel profiling of kinase inhibitors approved for clinical use from 2018 to 2020.xlsx

During the last two decades, kinase inhibitors have become the major drug class for targeted cancer therapy. Although the number of approved kinase inhibitors increases rapidly, comprehensive in vitro profiling and comparison of inhibitor activities is often lacking in the public domain. Here we report the extensive profiling and comparison of 21 kinase inhibitors approved by the FDA for oncology indications since June 2018 and 13 previously approved comparators on panels of 255 biochemical kinase assays and 134 cancer cell line viability assays. Comparison of the cellular inhibition profiles of the EGFR inhibitors gefitinib, dacomitinib, and osimertinib identified the uncommon EGFR p.G719S mutation as a common response marker for EGFR inhibitors. Additionally, the FGFR inhibitors erdafitinib, infigratinib, and pemigatinib potently inhibited the viability of cell lines which harbored oncogenic alterations in FGFR1-3, irrespective of the specific clinical indications of the FGFR inhibitors. These results underscore the utility of in vitro kinase inhibitor profiling in cells for identifying new potential stratification markers for patient selection. Furthermore, comparison of the in vitro inhibition profiles of the RET inhibitors pralsetinib and selpercatinib revealed they had very similar biochemical and cellular selectivity. As an exception, an NTRK3 fusion-positive cell line was potently inhibited by pralsetinib but not by selpercatinib, which could be explained by the targeting of TRK kinases in biochemical assays by pralsetinib but not selpercatinib. This illustrates that unexpected differences in cellular activities between inhibitors that act through the same primary target can be explained by subtle differences in biochemical targeting. Lastly, FLT3-mutant cell lines were responsive to both FLT3 inhibitors gilteritinib and midostaurin, and the PI3K inhibitor duvelisib. Biochemical profiling revealed that the FLT3 and PI3K inhibitors targeted distinct kinases, indicating that unique dependencies can be identified by combined biochemical and cellular profiling of kinase inhibitors. This study provides the first large scale kinase assay or cell panel profiling study for newly approved kinase inhibitors, and shows that comprehensive in vitro profiling of kinase inhibitors can provide rationales for therapy selection and indication expansion of approved kinase inhibitors.

近二十年来，激酶抑制剂（kinase inhibitors）已成为靶向癌症治疗的主流药物类别。尽管获批激酶抑制剂的数量持续快速增长，但公共领域中往往缺乏针对其活性的全面体外谱分析与对比研究。本研究针对2018年6月以来美国食品药品监督管理局（FDA）获批的21种肿瘤适应症激酶抑制剂，以及13种此前获批的对照药物，分别在255项生化激酶检测（biochemical kinase assays）与134项癌细胞系活力检测（cancer cell line viability assays）体系中开展了大规模谱分析与对比研究。对表皮生长因子受体（EGFR）抑制剂吉非替尼（gefitinib）、达可替尼（dacomitinib）与奥希替尼（osimertinib）的细胞抑制谱进行对比后，我们发现罕见的EGFR p.G719S突变可作为EGFR抑制剂的通用响应标志物。此外，成纤维细胞生长因子受体（FGFR）抑制剂厄达替尼（erdafitinib）、英菲格拉替尼（infigratinib）与培米替尼（pemigatinib）可强效抑制携带FGFR1-3致癌变异的细胞系活力，且不受该类FGFR抑制剂的具体临床适应症限制。上述结果凸显了细胞水平的激酶抑制剂体外谱分析在挖掘新的患者选择分层标志物方面的应用价值。进一步对比转染重排（RET）抑制剂普拉替尼（pralsetinib）与塞普替尼（selpercatinib）的体外抑制谱发现，二者的生化与细胞选择性高度相似。但存在一个例外：神经营养因子受体酪氨酸激酶3（NTRK3）融合阳性细胞系可被普拉替尼强效抑制，却不受塞普替尼影响——这一现象可通过普拉替尼（而非塞普替尼）在生化检测中靶向神经营养性酪氨酸受体激酶（TRK）激酶得到解释。这表明，即便作用于同一主要靶点的抑制剂，其细胞活性也可能存在意外差异，而这种差异可通过生化靶向的细微区别得到阐释。最后，FMS样酪氨酸激酶3（FLT3）突变细胞系对FLT3抑制剂吉瑞替尼（gilteritinib）、米哚妥林（midostaurin），以及磷脂酰肌醇3-激酶（PI3K）抑制剂杜韦利西布（duvelisib）均有响应。生化谱分析显示，FLT3抑制剂与PI3K抑制剂的靶向激酶各不相同，这表明通过联合开展激酶抑制剂的生化与细胞谱分析，可识别独特的肿瘤依赖特征。本研究是首个针对新近获批激酶抑制剂开展的大规模激酶检测或细胞系谱分析研究，并证实对激酶抑制剂进行全面体外谱分析，可为获批激酶抑制剂的治疗选择与适应症拓展提供科学依据。