AnnexinA5 renders dead tumor cells immunogenic—implications for multimodal cancer therapies

Abstract

Multimodal tumor therapies should aim not only to kill the tumor cells, but also to stimulate a specific immune response to keep residual tumor (stem) cells and metastases under control. Apoptotic cells are mostly noninflammatory or even anti-inflammatory while necrotic cells stimulate the immune system. Whether the immunogenicity of apoptotic tumor cells can be increased by interfering with their swift and phosphatidylserine (PS)-dependent clearance by macrophages was examined. AnnexinA5 (AnxA5) is a naturally occurring highly specific ligand for PS. Proteins of the annexin family are characterized by a selective affinity for phospholipids in the presence of Ca²⁺ ions. The phagocytosis by macrophages of irradiated, apoptotic tumor cells (ITC) was partially inhibited when the ITC were preincubated with AnxA5. Activated macrophages secreted higher amounts of TNFα and IL-1β after contact with ITC plus AnxA5 in comparison with ITC alone, while the amount of TGF-β was decreased. Macrophages of AnxA5-deficient mice showed an increased phagocytosis of dead cells. Wild-type mice, where endogenous AnxA5 is present, displayed a significantly faster decline in size of allogeneic tumors in comparison with AnxA5-deficient animals. The addition of AnxA5 to ITC vaccines increased the percentage of tumor-free mice in syngeneic tumor protection and tumor cure assays. AnxA5 alone led to a retard of syngeneic tumor growth that was, however, less pronounced in comparison to treatment of the tumor with ionizing irradiation. In conclusion, AnxA5 disturbs the PS-dependent clearance by macrophages of dying tumor cells, leading to the accumulation of the latter, to the occurrence of secondary necrotic cells, and to an increased uptake of the dead cells by dendritic cells. Tumor cure appendages with dead tumor cells should be performed with AnxA5 as an immune stimulator and could be combined with irradiation, chemotherapy, and hyperthermia to induce immunogenic cell death forms in vivo or ex vivo.

Keywords::

• AnnexinA5
• apoptosis
• macrophages
• inflammation
• immunomodulation
• ionizing irradiation
• tumor cell immunogenicity

Acknowledgments

This work was supported by “Deutsche Forschungsge-meinschaft” SFB 643, by the ELAN program of the Friedrich-Alexander University of Erlangen-Nürnberg, by AIF “Otto von Guerike” e.V. PROINNO II, KF0036801ULA, by the responsif GmbH Erlangen, by the European Commissions [NOTE: (TPA4 FP6) and APOCLEAR (QLK3-CT-2002-02017)], by the Programme Alban, the European Union Program of High Level Scholarships for Latin America, scholarship no. “E04D047956VE” to L. E. M., and by the DFG research training grant GK592.

Declaration of interest: The authors report conflicts of interest. The authors alone are responsible for the content and writing of the paper.