Stress-induced inflammation evoked by immunogenic cell death is blunted by the IRE1α kinase inhibitor KIRA6 through HSP60 targeting

Mounting evidence suggests that immunogenic therapies leveraging the unfolded protein response (UPR) during endoplasmic reticulum (ER) stress enhance cancer cell-immune interactions by promoting the release of immunomodulatory and proinflammatory factors from stressed or dying cancer cells. While UPR-driven transcription of proinflammatory cytokines and chemokines can either support or hinder tumor growth and antitumor immunity, the mechanisms regulating the inflammatory output of cancer cells during immunogenic therapies remain poorly understood.

In this study, we analyzed the transcriptomes of cancer cells exposed to immunogenic and weakly immunogenic treatments. Bioinformatics pathway analysis revealed that immunogenic treatments triggered an NF-κB/AP-1-mediated inflammatory stress response, distinct from both cell death and UPR activation. This stress-induced inflammation was selectively abolished by the IRE1α-kinase inhibitor KIRA6. Cancer cells co-treated with immunogenic chemotherapy and KIRA6 produced supernatants lacking proinflammatory and chemoattractant factors, which impaired neutrophil mobilization and dendritic cell maturation. In vivo, KIRA6 significantly reduced the vaccination potential of dying cancer cells exposed to immunogenic chemotherapy.

Mechanistic investigations showed that KIRA6’s anti-inflammatory effects persisted in IRE1α-deficient cells, suggesting an off-target effector. Using a KIRA6-clickable photoaffinity probe, mass spectrometry, and co-immunoprecipitation, we identified cytosolic HSP60 as an off-target of KIRA6. HSP60 was shown to regulate the IKK-driven NF-κB pathway, positioning it as a key mediator of the NF-κB/AP-1 inflammatory stress response induced by immunogenic treatments.

In summary, our findings identify HSP60 as a novel upstream regulator of NF-κB/AP-1-mediated inflammation that is inhibited by KIRA6. These results highlight the complexity of interpreting the anti-inflammatory effects of IRE1α inhibitors and call for careful consideration of off-target effects in future studies.