Despite the promise of immune checkpoint therapies, treatment-resistant cancers remain a stubborn frontier, making it difficult to predict treatment responsiveness and optimize treatment protocols. A major obstacle? Our limited ability to track how tumor-infiltrating T cells expand over time. Now, researchers reveal a gene signature that could decode this critical immune burst—and reshape how we design and evaluate cancer immunotherapies.
A team of researchers led by Satoshi Ueha, PhD, and Kouji Matsushima, MD, PhD, from the Research Institute for Biomedical Sciences at Tokyo University of Science (TUS), has developed a novel method to monitor CD8⁺ T cell activity over time within tumors. Their study, published in Nature Communications and titled “A pan-immunotherapy signature to predict intratumoral CD8⁺ T cell expansions,” uncovers new insights into how tumor-infiltrating T cells expand, how this expansion can be predicted, and how these cells might be therapeutically reactivated.
“The development of immunotherapies has been hindered by our inability to comprehensively monitor their effects on immune cells—particularly cancer-fighting T cells—over time,” explained Ueha. “Building on our previous work, we developed a method to track these cells longitudinally in the tumor, allowing us to gain deeper insights into the burst of proliferation that drives effective anti-tumor responses.”
To achieve this, the team implanted tumors at distinct anatomical locations in mice, developing “a multi-site tumor mouse model system to track hundreds of expanding and contracting CD8⁺ T cell clones over multiple timepoints in tumors of the same individual,” the authors wrote. By leveraging unique T cell receptor (TCR) sequences as natural barcodes, the researchers tracked hundreds of individual CD8⁺ T cell clones over time—capturing a dynamic, clonal-level view of the immune response that had previously been inaccessible.
The researchers profiled thousands of tumor-infiltrating CD8⁺ T cells, capturing a clonal level-view of their expansion dynamics using next-generation single-cell RNA and TCR sequencing. This approach enabled them to identify a consistent gene expression pattern—termed the “expansion signature”—that appeared in T cells prior to proliferation. The signature was enriched in expanding clones across multiple tumor sites and timepoints, and it remained predictive of T cell expansion in both untreated and immunotherapy-treated mice. Importantly, when applied to human datasets, the expansion signature correlated with improved survival in patients undergoing PD-1 blockade therapy, underscoring its translational potential as a pan-immunotherapy biomarker.
Although the expansion signature diminished as CD8⁺ T cells entered contraction, the researchers observed that a subset of cells within the tumor retained transcriptional features associated with proliferative potential. To probe whether these cells could be therapeutically reactivated, the team administered LAG-3 blockade. This intervention re-induced the expansion signature and triggered renewed clonal proliferation.
These findings position the expansion signature not only as a dynamic biomarker for monitoring immune responses, but also as a potential lever for designing therapies that restore anti-tumor T cell activity. “Our work opens the door to a dynamic understanding of how immunotherapies succeed or fail in real time,” said Ueha. “We hope that the expansion signature can serve not only as a predictor of treatment response but also as a guide for designing new therapies that can reawaken the immune system when it begins to falter.”
This study offers a high-resolution view of T cell expansion dynamics within tumors and introduces a gene signature with broad utility across immunotherapy platforms. By enabling real-time tracking and potential reactivation of tumor-fighting T cells, the expansion signature potentially lays the groundwork for next-generation immunodynamic therapies and a more personalized approach to cancer treatment.
