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Sharif, Anfal Anwar (2020) TGF-β1-bearing cancer-derived microvesicles as mediators of intercellular communication in cancer. Doctoral thesis, London Metropolitan University.
Cell-to-cell communication is a prerequisite for differentiation and development in multicellular organisms. In the last decade, substantial progress has been made in the discovery of microvesicles (MVs) as an additional vector of cell-to-cell communication. MVs are intact, submicron and phospholipid-rich particles that can serve as a vehicle for transfer of signalling molecules between cells. Although it has emerged that MVs may coordinate a plethora of physiological processes, compelling evidence has recently highlighted MVs as a major contributor to oncogenesis by mediating several oncogenic mechanisms that remains poorly defined. In the present study, we attempted to investigate the oncogenic activity of cancer-derived MVs on cell proliferation and apoptotic signalling in untransformed cell lines, and to elucidate the underlying mechanism, by which cancer-derived MVs may contribute to cell survival and chemoresistance. The purity of several cancer-derived MVs isolated from Jurkat, PC-3 and A549 cell lines were confirmed through detection of PS-positive MVs, or MVs-expressing 13-actin and CD63 markers. In our findings, we found that normal cell lines, including MRC-5, PNT2 and hTERT-RPE-1 cell lines exposed to cancer-derived MVs displayed enhanced cell proliferation as a result of abolished apoptotic signalling. Accordingly, the caspase activity and the pro-apoptotic protein, Bax were attenuated; meanwhile, the anti-apoptotic protein, Bcl-2 was identified to be upregulated. In light of these findings, we reported that cancer-derived MVs could diminish the phosphorylation of SMAD3 and inhibit its nuclear translocation. In contrast, oncoproteins, including AKT and ERK1/2 were observed to be highly phosphorylated. Targeting AKT phosphorylation using MK-2206, an allosteric inhibitor of pan-AKT isoforms, resulted in the hyperphosphorylation of SMAD3. To further extent, we also revealed that cancer-derived MVs requires the acidic microenvironment to aid in these tumourigenic events, which appeared to be attributed to TGF-B1 activation. Indeed, cancer-derived MVs exhibited significant levels of active TGF-f31 under acidic condition, and that targeting Tf3RI using SB-431542, a potent and selective TBRI kinase inhibitor, counteracted the effect of MVs-mediated oncogenic transformation of normal cell lines. Furthermore, we found that MRC-5 cell lines exposed to cancer-derived MVs displayed resistance to cisplatin or doxorubicin, but restored their drug sensitivity upon SB-431542 or MK-2206 treatment. Herein, we have for the first time demonstrated that cancer-derived MVs mediates cell survival and chemoresistance through TGF-f31-AKT-SMAD3 signalling axis, and that exploiting this signalling cascade may pave the way for an attractive therapeutic avenue to combat MVs-mediated oncogenesis.
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