Abstract
Complement C5a is a potent inflammatory chemoattractant and might be a beneficial therapeutic target for the induction of an effective anti-tumour response. C5a agonist and antagonist modulation have been demonstrated to either promote or inhibit tumour development in EMT6 murine mammary cancer cells in both in vitro and in vivo studies. For the in vitro studies Alamar Blue cell viability assay was used for cell viability determination, and an immunofluorescence assay was used to determine the location of C5aR expression on the EMT6 cell line. For the in-vivo experiment, female Balb/c mice were subcutaneously injected with EMT6 tumour cells and subsequently treated with both C5aR agonist and antagonist peptides. At the end of the in vivo study period of 14 days, liver and tumour samples were obtained for an ELISA assay to quantify the levels of TNF-α, caspase, C5a and VEGF-A signals following treatment with both C5aR agonist and antagonist. The in vitro experiment revealed that an expression of C5aR was found on the cell membrane of the EMT6 cells, and treatment with EP54, which is a C5aR agonist, showed low cell viability 48 h posttreatment. For the in vivo experiment, the ELISA assay outcome showed that EP54 significantly promoted higher numbers of circulating signalling proteins except for VEGF-A, suggesting that the C5aR agonist modulation might inhibit tumour growth and also trigger the induction of apoptosis.
Highlights
Complement Component 5a (C5a) is a potent inflammatory chemoattractant and might be a beneficial therapeutic target for the induction of an effective anti-tumour response
A green color appeared in the EMT6 cells and indicated expression of C5aR on the plasma membrane only, which was differentiated from the cell nucleus by the blue fluorescent color (Fig. 1)
EMT6 cells treated with PMX205 showed a higher % of Alamar Blue (AB) reduction, compared to cells treated with EP54 or tamoxifen (Table 2)
Summary
Complement C5a is a potent inflammatory chemoattractant and might be a beneficial therapeutic target for the induction of an effective anti-tumour response. Complement is a key mediator of inflammation [6, 7] and activation of the complement system results in the formation of a pore-like membrane attack complex (MAC) [8] and the release of the potent C5a peptide [9]. C5a induces inflammation following interaction with its specific receptor, C5aR [10, 11] and has been associated with various disease processes, such as rheumatoid arthritis [12], brain disorders [13, 14], inflammatory bowel diseases [15], cancers [16], sepsis [17] and ischemia reperfusion injuries [18]. The functional role of complement in cancer immunotherapy is poorly understood, since C5a has been reported to enhance lung cancer in mice by inhibiting myeloid-derived suppressor cells [19], while inducing regression of mammary tumours in mice [20]. It has been reported that the presence of the complement regulatory protein enhanced the immunemediated clearance of tumour cells [25]
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