The reciprocal relationship shared between breast cancer cells and cells of the tumour microenvironment is crucial in relation to metastasis. MDA-MB-231 breast cancer cells have been shown to release the S1P2 receptor in exosomes, where it is taken up by fibroblasts and N-Terminally processed to a shorter, constitutively active form. The aim of the investigation was to examine the effect of truncated S1P2 on the activation of the ERK-1/2 pathway in various cell lines. Indeed, using truncated (1x and 2x HA tagged) S1P2 receptor constructs we have demonstrated constitutive activation of the ERK-1/2 pathway in HEK293 cells, MEFs but not MDA-MB-231 cells. Sustained ERK-1/2 activation could facilitate expansion of the fibroblast population and may prove an important step in the metastatic cascade. Indeed, our results have shown the 2x HA-tagged truncated S1P2 to promote DNA synthesis in HEK293 cells and MEFs but not MDA-MB-231 cells. This would suggest that ERK-1/2 activated by the truncated receptor is responsible for promoting DNA synthesis. The S1P2 receptor sub-type selective agonist, CYM5520, was shown to inhibit DNA synthesis in HEK293 cells (p = 0.1), MEFs (p = 0.01) and MDA-MB-231 cells (p < 0.05). Similarly, the S1P4 selective agonist CYM50308 was also shown to significantly inhibit DNA synthesis in HEK293 cells (p < 0.01), MEFs (p < 0.001) and MDA-MB-231 cells (p < 0.05). Since these agonists were shown to induce ERK-1/2 activation in HEK293 cells, there is a correlation between activation of this kinase pathway and inhibition of DNA synthesis in these cells. However, there was no activation of ERK-1/2 by these agonists in MEFs or MDA-MB-231 cells which indicates there may be activation or inhibition of alternative pathways to achieve this response. The inhibition of DNA synthesis by CYM5520 and CYM50308 could be explained by the agonists promoting the migration of these cells. Migrating cells require growth arrest and therefore DNA synthesis would be inhibited. Indeed, we have shown CYM5520 to induce migration of MDA-MB-231 cells (p < 0.05). Therefore, we speculate here that MDA-MB-231 cells may release the S1P2 receptor early in the metastatic cascade so as to increase DNA synthesis and subsequent proliferation. When taken together, our results show a clear function for this unique form of GPCR signalling in breast cancer tumorigenesis.
|Date of Award||11 Mar 2019|
- University Of Strathclyde
|Supervisor||Nigel Pyne (Supervisor)|