Complement C5a Fosters Tumor-promoting Immunity by Recruiting regulatory T cells in hepatocellular carcinoma

Anaphylatoxins are active complement components that regulate immune cells. However, their role in shaping the tumor immune microenvironment (TIME) in hepatocellular carcinoma (HCC) remains poorly understood. This study utilized CRISPR/Cas9 technology to knock out complements proteins C3 or C5 in mouse liver cancer cells. Both in vivo and in vitro experiments revealed that C5 knockout significantly inhibited tumor growth in mouse models. Moreover, C5ar1-knockout mice were generated using C57BL/6J mice, and immune cell profiles within subcutaneous tumors were analyzed using mass spectrometry and flow cytometry. The results demonstrated that C5ar1 knockout significantly reduced the infiltration of regulatory T cells (Tregs) and M2-like tumor-associated macrophages (TAMs) into the TIME. Conversely, the infiltration of effector CD8⁺ T cells, known for their tumor-killing activity, was markedly enhanced. Mechanistically, C5a was shown to promote the M2-like polarization of macrophages and the C-C-motif chemokine ligand 5 (CCL5) expression by activating the AKT signaling pathway. Further studies identified TAM-derived CCL5 as a key driver of the chemotactic activity of Tregs. Finally, blocking the C5a/C5ar1 axis synergistically enhanced the anti-tumor effects of PD-1 monoclonal antibody in both transplanted and spontaneous liver cancer mouse models. Overall, these findings reveal that cancer cell-derived anaphylatoxin C5a plays a pivotal role in promoting an inhibitory TIME in liver tumor tissues. Blocking the C5a/C5ar1 axis in combination with immune checkpoint inhibitors could represent a promising strategy for treating HCC.