pH-responsive activation of Tet-On inducible CAR-T cells enables spatially selective treatment of targeted solid tumors at reduced safety risk

Chimeric antigen receptor T (CAR-T)-cell therapy is a promising resolution for solid tumors, but its corresponding clinical translation has been hindered by unsatisfactory therapeutic potency and severe cytokine release syndrome. Herein, tetracycline (Tet)-On inducible human epidermal growth factor receptor 1 (HER1)-targeted CAR-T (Tet-HER1-CAR-T) cells were engineered to enable spatially selective activation at tumor sites by doxycycline (Doxy), which is delivered by pH-responsive stealth liposomal calcium carbonate nanoparticles (Doxy@CaCO₃-PEG). Compared with the intravenous administration of conventional HER1-CAR-T cells and Tet-HER1-CAR-T cells activated by free Doxy, concurrent intravenous administration of Tet-HER1-CAR-T cells and Doxy@CaCO₃-PEG leads to the localized tumor activation of Tet-HER1-CAR-T cells and reduced systemic secretion of inflammatory cytokines. Together with its ability to protect Tet-HER1-CAR-T cells from tumor-acidity-induced dysfunction by neutralizing tumor acidity, Doxy@CaCO₃-PEG injection synergized with Tet-HER1-CAR-T cells to effectively suppress the growth of HER1-overexpressing subcutaneous triple-negative breast cancer (TNBC) tumors, lung tumors and orthotopic lung tumors in mice. Furthermore, Doxy@CaCO₃-PEG-activated Tet-HER1-CAR-T-cell therapy synergistically suppressed HER1 inhibitor-resistant TNBC tumors and immunosuppressive Fusobacterium nucleatum (F.n.) colonized HER1-overexpressing TNBC patient-derived tumor xenografts. This study highlights that the Doxy@CaCO₃-PEG-induced pH-responsive activation of Tet-HER1-CAR-T cells is a highly spatially selective strategy for effectively eradicating targeted solid tumors with improved safety.