TraCe-seq was able to identify pathways that can mediate differential response to EGFR inhibitors.

  • Major Finding: TraCe-seq was able to identify pathways that can mediate differential response to EGFR inhibitors.

  • Concept: Single-cell RNA sequencing tracked the response to EGFR inhibitors with different mechanisms of action.

  • Impact: The TraCe-seq approach can be used to identify how transcriptional programs affect treatment response.

A promising class of cancer treatment is targeted therapies against oncogenic drivers. Differential responses to these therapies as well as resistance, however, continue to present challenges for patients. To this end, Chang and colleagues developed the TraCe-seq platform, a DNA barcode-based single-cell sequencing method that allows for the early capturing of the origins and adaptive processes underlying therapeutic resistance and facilitates the comparison between different therapeutic modalities at the single-cell level, providing insight into response and resistance to drugs. TraCe-Seq was used to compare a conventional EGFR kinase inhibitor and a dual EGFR inhibitor-degrader and to establish if drug-resistant clones and known resistant associated genes could be captured and whether differential resistance/response mechanisms would be observed with agents with different mechanisms of action. TraCe-seq could capture the sensitive and resistant phenotypes in an EGFR inhibitor–sensitive lung adenocarcinoma cell line with a drug-resistant subpopulation. Compared with EGFR kinase inhibitors, the EGFR inhibitor-degrader was less effective at reducing the number and diversity of TraCE-Seq barcodes despite similar levels of MAPK pathway suppression, and accordingly a greater number of cells remained after treatment. Further analysis revealed that upregulated genes were different in cells that were resistant to degraders as compared with cells that were resistant to EGFR kinase inhibitors, with degrader-resistant cells exhibiting diminished activity of genes involved in protein processing within the endoplasmic reticulum (ER). Paradoxically, the EGFR protein itself was needed for full efficacy of EGFR inhibition, as its persistent expression increased ER stress and subsequent pro-death signaling to enhance inhibitor cytotoxicity. Based on these observations, the EGFR degrader was combined with inducers of ER stress and led to near-complete elimination of residual cells in these models. This study illustrates the potential utility of the TraCe-seq platform for evaluating transcriptional responses to drug treatment and, through its use, reveals key differences between EGFR kinase inhibitors and degraders and determines that induction of ER stress is critical for full efficacy of EGFR inhibitors.

Chang MT, Shanahan F, Nguyen TTT, Staben ST, Gazzard L, Yamazoe S, et al. Identifying transcriptional programs underlying cancer drug response with TraCe-seq. Nat Biotechnol 2021 Sep 16 [Epub ahead of print].

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