Tumor-derived endoplasmic reticulum stress reprograms immune cells toward a regulatory phenotype in renal cell carcinoma

Abstract Background Renal cell carcinoma (RCC) is characterized by a paradoxically immunosuppressive tumor-immune microenvironment despite dense immune infiltration, and resistance to immune checkpoint inhibitor (ICI) therapy remains a major clinical challenge. Endoplasmic reticulum stress (ERS) and the unfolded protein response (UPR) are persistently activated in RCC and have been implicated in tumor survival and immune dysfunction, yet whether tumor-derived ERS actively reprograms infiltrating immune cells remains undefined. Methods Human RCC cell lines (Caki-2 and TK-10) were cultured alone or co-cultured with peripheral blood mononuclear cells (PBMCs) and exposed to the ERS inducer tunicamycin (Tun) and the chemical chaperone tauroursodeoxycholic acid (TUDCA). Cell viability, migration, UPR, and immune marker expression were assessed by MTT assay, scratch wound healing, RT-qPCR, western blot, and 2D/3D immunofluorescence. Ultrastructural analysis was performed by scanning and transmission electron microscopy. To test cell-non-autonomous ERS transmission, naïve PBMCs were exposed to conditioned media (CM) collected from drug-pretreated RCC cells following complete media replacement to eliminate residual compounds; CM from unstressed RCC cells served as control. Results Tun induced robust UPR activation — confirmed across qPCR, western blot, and immunofluorescence by HSPA5 /BiP and DDIT3 /CHOP induction — in both RCC lines and in directly treated PBMCs. At the sublethal dose used, PBMC viability was preserved, yet Tun drove coordinated downregulation of CD3, CD4, CD8, PD-1, PD-L1, and FoxP3 in co-culture, indicating non-lethal immune reprogramming rather than cytotoxicity. In 3D spheroid co-cultures, Tun excluded CD3⁺ T cells from the tumor mass while TUDCA permitted immune infiltration, spatially confirming ERS-dependent immune exclusion. CM from Tun-treated — but not unstressed — RCC cells activated UPR markers ( HSPA5 /BiP and EIF2AK /PERK) and induced FoxP3 and CTLA-4 in naïve PBMCs across both donor lines, establishing that ER-stressed tumor cells transmit an immunosuppressive signal via their secretome independently of direct drug exposure. Prior ERS relief in donor cells with TUDCA attenuated this transmissible effect. Conclusions Tumor-derived ERS non-lethally remodels the RCC-immune interface and transmits a UPR-activating, FoxP3-associated immunosuppressive program to bystander immune cells via the tumor secretome. These findings reframe ERS not as a passive metabolic correlate but as an active, transmissible regulator of immune dysfunction in RCC, with direct implications for understanding ICI resistance and for rational combination of ERS-targeting strategies with immunotherapy.