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bioRxiv · 10.1101/2025.08.06.668639

Spatiotemporal lineage mapping of tumor immune escape with eTRACER

Yang, J.·Hou, L.·Wang, X.·Zhang, N.·Bian, Y.·Lu, Z.·Chen, Y.·Xie, D.·Fang, Y.·Wang, K.·Wan, R.·Jin, Y.·Cai, X.·Lee, L. T. O.·Hu, Z.·Ji, H.
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Abstract

The plasticity-first hypothesis posits that phenotypic plasticity emerges first in novel environments, followed by (epi)genetic changes that stabilize adaptive phenotypes. Whether tumor evolution follows this model, and how non-genetic mechanisms sustain long-term therapeutic resistance, remain unclear. Here, we develop eTRACER, an endogenously targeted, high-resolution and spatially-resolved CRISPR-Cas9 lineage-tracing system, to study tumor adaptation during longitudinal CD8+ T cell-mediated immunotherapy. Integrated clonal and single-cell transcriptomic analyses show that tumor immune evasion is governed by clonal state composition rather than clonal identity, arguing against selection from pre-existing genetic heterogeneity. Spatiotemporal lineage tracing reveals highly reversible state transitions prior to therapy, followed by consolidated epithelial-mesenchymal transition (EMT) during therapy. Single-cell multiomics identifies AP-1 as a key regulator of EMT stabilization and its inhibition enhances tumor sensitivity to immune killing. Together, these findings show that tumor plasticity precedes stable immune evasion, revealing an AP-1-centered mechanism underlying plasticity-first tumor evolution.

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Yang, J., Hou, L., Wang, X., Zhang, N., Bian, Y., Lu, Z., Chen, Y., Xie, D., Fang, Y., Wang, K., Wan, R., Jin, Y., Cai, X., Lee, L. T. O., Hu, Z., Ji, H.. 2025-08-09. Spatiotemporal lineage mapping of tumor immune escape with eTRACER. https://doi.org/10.1101/2025.08.06.668639

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