title: Cancer-associated fibroblasts from lung tumors maintain their immunosuppressive abilities after high-dose irradiation.
abstract: Accumulating evidence supports the notion that high-dose (>5 Gy) radiotherapy (RT) regimens are triggering stronger pro-immunogenic effects than standard low-dose (2 Gy) regimens. However, the effects of RT on certain immunoregulatory elements in tumors remain unexplored. In this study, we have investigated the effects of high-dose radiotherapy (HD-RT) on the immunomodulating functions of cancer-associated fibroblasts (CAFs). Primary CAF cultures were established from lung cancer specimens derived from patients diagnosed for non-small cell lung cancer. Irradiated and non-irradiated CAFs were examined for immunomodulation in experiments with peripheral blood mononuclear cells from random, healthy donors. Regulation of lymphocytes behavior was checked by lymphocyte proliferation assays, lymphocyte migration assays, and T-cell cytokine production. Additionally, CAF-secreted immunoregulatory factors were studied by multiplex protein arrays, ELISAs, and by LC-MS/MS proteomics. In all functional assays, we observed a powerful immunosuppressive effect exerted by CAF-conditioned medium on activated T-cells (p > 0.001), and this effect was sustained after a single radiation dose of 18 Gy. Relevant immunosuppressive molecules such as prostaglandin E2, interleukin-6, and -10, or transforming growth factor-β were found in CAF-conditioned medium, but their secretion was unchanged after irradiation. Finally, immunogenic cell death responses in CAFs were studied by exploring the release of high motility group box-1 and ATP. Both alarmins remained undetectable before and after irradiation. In conclusion, CAFs play a powerful immunosuppressive effect over activated T-cells, and this effect remains unchanged after HD-RT. Importantly, CAFs do not switch on immunogenic cell death responses after exposure to HD-RT.
authors.affiliation: Department of Clinical Medicine, University of Tromsø , Tromsø , Norway.
authors.forename: Laia
authors.initials: L
authors.lastname: Gorchs
authors.affiliation: Department of Oncology and Radiotherapy, University Hospital of Northen Norway , Tromsø , Norway.
authors.forename: Turid
authors.initials: T
authors.lastname: Hellevik
authors.affiliation: Department of Medical Biology, University of Tromsø , Tromsø , Norway.
authors.forename: Jack-Ansgar
authors.initials: JA
authors.lastname: Bruun
authors.affiliation: Department of Medical Biology, University of Tromsø , Tromsø , Norway.
authors.forename: Ketil-Andre
authors.initials: KA
authors.lastname: Camilio
authors.affiliation: Department of Medical Biology, University of Tromsø , Tromsø , Norway ; Department of Pathology, University Hospital of Northern Norway , Tromsø , Norway.
authors.forename: Samer
authors.initials: S
authors.lastname: Al-Saad
authors.affiliation: Department of Medical Biology, University of Tromsø , Tromsø , Norway.
authors.forename: Tor-Brynjar
authors.initials: TB
authors.lastname: Stuge
authors.affiliation: Department of Clinical Medicine, University of Tromsø , Tromsø , Norway.
authors.forename: Inigo
authors.initials: I
authors.lastname: Martinez-Zubiaurre
keywords: cancer-associated fibroblasts; high-dose radiation; immunogenic cell death; lung cancer; tumor immunology