Inducing a metastable tumour cell state triggers viral mimicry and enhances anti-tumour immunity

Immune checkpoint blockade (ICB) has transformed cancer therapy, yet its efficacy remains largely restricted to tumours with pre-existing T cell infiltration. Immune-cold malignancies are refractory to ICB due to low tumour immunogenicity and active immune suppression. Tumour-intrinsic calcium signalling is frequently rewired to sustain plasticity, survival and immune evasion. In particular, the calcineurin (CN)–nuclear factor of activated T cells (NFAT) axis acts as a transcriptional hub promoting tumour progression. We hypothesized that selective disruption of CN–NFAT signalling in tumour cells could enhance anti-tumour immunity and improve therapeutic outcome. To test this, we used a CN–NFAT inhibitory peptide that preserves CN catalytic activity while selectively blocking NFAT activation. Tumour cell–intrinsic CN–NFAT disruption increased the frequency of intratumoral progenitor exhausted CD8+ T cells across multiple tumour models. This effect was abrogated by CXCR3 blockade, indicating chemokine-dependent recruitment. Importantly, these CD8+ T cells remained responsive to ICB, resulting in reduced tumour growth. Lipid nanoparticle-mediated delivery of inhibitory peptide messenger RNA similarly overcame resistance to ICB. Bulk RNA sequencing revealed induction of an interferon-like transcriptional program. In vitro analyses confirmed activation of the stimulator of interferon genes pathway and upregulation of CXCL9, CXCL10 and CXCL11. Single cell RNA sequencing demonstrated that CN–NFAT disruption alters tumour cell-state transitions, leading to the accumulation of a metastable stem-like tumour cell population. These “trapped” metastable cells resulted in stress-associated activation of interferon- inducing pathways, including retinoic acid–inducible gene I (RIG-I) and melanoma differentiation–associated gene 5 (MDA5), consistent with a viral mimicry phenotype. In conclusion, cell-intrinsic activation of CN–NFAT pathway enables tumour cell-state transitions. Therefore, its inactivation drives the persistence of metastable populations that activate endogenous viral mimicry programs, converting immune-cold tumours into T cell–inflamed lesions and sensitizing them to ICB.