Genetic and non-genetic heterogeneity within cancer cell populations represent major challenges to anticancer therapies. We currently lack robust methods to determine how preexisting and adaptive features affect cellular responses to therapies. Here, by conducting clonal fitness mapping and transcriptional characterization using expressed barcodes and single-cell RNA sequencing (scRNA-seq), we have developed tracking differential clonal response by scRNA-seq (TraCe-seq). TraCe-seq is a method that captures at clonal resolution the origin, fate and differential early adaptive transcriptional programs of cells in a complex population in response to distinct treatments. We used TraCe-seq to benchmark how next-generation dual epidermal growth factor receptor (EGFR) inhibitor–degraders compare to standard EGFR kinase inhibitors in EGFR-mutant lung cancer cells. We identified a loss of antigrowth activity associated with targeted degradation of EGFR protein and an essential role of the endoplasmic reticulum (ER) protein processing pathway in anti-EGFR therapeutic efficacy. Our results suggest that targeted degradation is not always superior to enzymatic inhibition and establish TraCe-seq as an approach to study how preexisting transcriptional programs affect treatment responses.

癌细胞群体中的遗传和非遗传异质性代表了抗癌治疗的主要挑战。我们目前缺乏可靠的方法来确定先前存在的和适应性特征如何影响细胞对治疗的反应。在这里，通过使用表达的条形码和单细胞 RNA 测序 (scRNA-seq) 进行克隆适应性作图和转录表征，我们开发了通过 scRNA-seq (TraCe-seq) 跟踪差异克隆反应。TraCe-seq 是一种以克隆分辨率捕获复杂群体中细胞响应不同治疗的起源、命运和差异早期适应性转录程序的方法。我们使用 TraCe-seq 来比较下一代双重表皮生长因子受体 (EGFR) 抑制剂-降解剂与 EGFR 突变肺癌细胞中标准 EGFR 激酶抑制剂的比较。我们确定了与 EGFR 蛋白靶向降解相关的抗生长活性丧失以及内质网 (ER) 蛋白加工途径在抗 EGFR 治疗效果中的重要作用。我们的结果表明，靶向降解并不总是优于酶促抑制，并将 TraCe-seq 建立为一种研究先前存在的转录程序如何影响治疗反应的方法。