Anales RANF

P.77 INTERACTION OF P2X7 WITH THE ER STRESS/UNFOLDED PROTEIN RESPONSE PATHWAYS S. Javed, F. Koch-Nolte, F. Haag* University Medical Center Hamburg-Eppendorf, Hamburg, Germany. Some chemotherapeutic agents such as doxorubicin (DOX) can activate the immune system by eliciting immunogenic cell death (ICD) of cancer cells. ICD is induced by triggering of the ER stress/unfolded protein response (UPR) pathways, and is characterised by the translocation of calreticulin (CRT) from the ER to the cell surface accompanied by the release of other danger signals such as ATP. In this context, P2X7 plays an important role as a receptor on antigen-presenting cells (APCs) that mediates their pro-inflammatory maturation. Complementary to its role on APCs we investigated the role of P2X7 on tumour cells with regard to the induction of ICD. In murine Yac-1 lymphoma cells low doses of extracellular ATP (eATP) synergistically enhanced the cytotoxic effects of DOX. This effect was blocked by an antagonistic and further enhanced by an agonistic nanobody to P2X7. Gating of P2X7 augmented the initial uptake of DOX into cells, but enhanced cell death was also observed when DOX was washed away before exposure to eATP, suggesting an interaction of the downstream signalling pathways. Sub-threshold doses of eATP also synergistically enhanced the cytotoxicity of the proteasome inhibitor Bortezomib, as well as DOX- mediated CRT translocation, suggesting that P2X7 might interact with the UPR. Incubation of Yac-1 cells with eATP alone for 10 min induced CRT translocation, phosphatidylserine surface exposure, and DAPI uptake. Interestingly, all these effects were blocked in the presence of GSK2606414, an inhibitor of the PKR-like ER kinase (PERK). Our results suggest that P2X7 signalling is intimately linked with the ER and provides an early input into the ER stress/UPR pathway that may modify the outcome of this signaling pathway when it is induced by other agents. * This work is part of SFB1328 “Adenine Nucleotides in Immunity and Inflammation” funded by the DFG