Abstract 1319: Complement activation in pancreatic ductal adenocarcinoma

Abstract

Abstract The complement system is an innate immune cascade which recognizes and clears microbes and damaged host cells. Complement activation has been linked to the promotion of tumor growth and metastasis across various anatomical sites with context-dependent effects. We therefore aim to understand the role for complement activation in pancreatic cancer. We have generated a tamoxifen-inducible autochthonous mouse model of PDAC which recapitulates genetic mutations found in humans using LSL-KrasG12D/+; Tgfbr2fl/fl; Ptf1a-CreER (KTC) mice. Using these mice, we assessed for complement activation in pancreatic tumors by western blot and confocal immunofluorescence. Western blotting for neoepitopes generated upon cleavage of Complement Component 3 (C3) demonstrated significantly increased complement activation in KTC tumors when compared to normal pancreas from at least 4-7 months post-induction. Using confocal immunofluorescence, we observed deposition of complement fragments in tumor stroma and on ductal carcinoma cells identified by co-staining with cytokeratin 19. We therefore hypothesized complement blockade would reduce PDAC tumor growth and sought to test this using C3 knockout (C3 KO) mice which lack all downstream complement function due to the central role of C3. We assessed flank tumor growth using two Kras; Tp53 (KPC) mutant-derived tumor cell lines in wild type (WT) C57BL/6J and C3 KO mice yet did not find a biologically significant reduction in flank tumor growth between WT and C3 KO. To validate the knockout, we assessed C3 activation by western blot but surprisingly found increased complement activation in C3 KO tumors. RT-qPCR demonstrated local transcription of C3 mRNA in both WT and C3 KO tumors as well as transcription of C3 mRNA in KPC cells in culture. This demonstrates that KPC cells are capable of autocrine production of C3. Using immunocytochemistry and RT-qPCR we identified expression of the C3a receptor in KPC cells which we hypothesized could serve as a sensor for feedback to control local C3 production. We applied 10 nM recombinant C3a to KPCY cells in vitro and found a 20% reduction in C3 mRNA transcription 1-hour post-treatment. Pre-treatment with 500 nM C3aR Antagonist (SB290157) ablated this effect. In summary, complement activation is significantly increased in PDAC and appears to be activated in part by tumor cells themselves. Autocrine production of C3 by tumor cells is sufficient to produce complement activation in the absence of systemic C3. Additional experiments are warranted to determine the role of complement in PDAC using C3 mice crossed with KTC mice to assess tumorigenesis in the total absence of C3. Citation Format: Brett I. Bell, Sanjay Pandey, Patrik Asp, Chandan Guha. Complement activation in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1319.