Cetuximab treatment alters the content of extracellular vesicles released from tumor cells
Abstract
Aim: Extracellular vesicles (EVs) are attractive candidates for biomarker research, because their content reflects the parental cell status. This study aimed to examine whether tumor cell derived EVs mirrored the cellular changes caused by treatment with cetuximab, a therapeutic antibody that blocks activation of EGF receptor (EGFR). Materials & methods: A-431 cells were treated with cetuximab for 48 h. EVs were isolated using differential centrifugation and protein content was analyzed using western blotting. Results: EV levels of EGFR and phospho-EGFR were reduced after cetuximab treatment, reflecting similar changes in the parental cells. In addition, cetuximab was found associated with EVs. Conclusion: EVs could serve as biomarkers to monitor cetuximab treatment. Association of cetuximab with EVs might influence its behavior.
References
- 1 Overexpression of epidermal growth factor receptor as a prognostic factor in colorectal cancer on the basis of the Allred scoring system. Onco Targets. Ther. 6, 967–976 (2013).
- 2 Epidermal growth factor receptor (EGFR) expression is associated with a worse prognosis in gastric cancer patients undergoing curative surgery. World J. Surg. 31(7), 1458–1468 (2007).
- 3 . Structural basis for inhibition of the epidermal growth factor receptor by cetuximab. Cancer Cell. 7(4), 301–311 (2005).
- 4 . Antibody-induced epidermal growth factor receptor dimerization mediates inhibition of autocrine proliferation of A431 squamous carcinoma cells. J. Biol. Chem. 269(44), 27595–27602 (1994).
- 5 . Anti-Epidermal Growth Factor Receptor Monoclonal Antibody 225 Up-Regulates p27 KIP1 and Induces G 1 Arrest in Prostatic Cancer Cell Line DU145 Advances in Brief Anti-Epidermal Growth Factor Receptor Monoclonal Antibody 225 Up-Regulates p27KIP1 and Induces (1996). http://cancerres.aacrjournals.org/content/56/16/3666.full.pdf.
- 6 Different antiproliferative effects of matuzumab and cetuximab in A431 cells are associated with persistent activity of the MAPK pathway. Eur. J. Cancer 45(7), 1265–1273 (2009).
- 7 . Antibody-dependent cellular cytotoxicity of cetuximab against tumor cells with wild-type or mutant epidermal growth factor receptor. Cancer Sci. 98(8), 1275–1280 (2007).
- 8 . Cetuximab induce antibody-dependent cellular cytotoxicity against EGFR-expressing esophageal squamous cell carcinoma. Int. J. Cancer 120(4), 781–787 (2007).
- 9 . Molecular inhibition of angiogenesis and metastatic potential in human squamous cell carcinomas after epidermal growth factor receptor blockade. Mol. Cancer Ther. 1, 507–514 (2002).
- 10 KRAS mutation status is predictive of response to cetuximab therapy in colorectal cancer. Cancer Res. 66(8), 3992–3995 (2006).
- 11 . In vitro and in vivo antitumor activity of cetuximab in human gastric cancer cell lines in relation to epidermal growth factor receptor (EGFR) expression and mutational phenotype. Gastric Cancer 15(3), 252–264 (2012).
- 12 Gene copy number for epidermal growth factor receptor (EGFR) and clinical response to antiEGFR treatment in colorectal cancer: a cohort study. Lancet Oncol. 6(5), 279–286 (2005).
- 13 . Extracellular vesicles: emerging targets for cancer therapy. Trends Mol. Med. 20(7), 385–393 (2014).
- 14 . Exosome Mediated Communication within the Tumor Microenvironment. Elsevier B.V. http://linkinghub.elsevier.com/retrieve/pii/S0168365915300018.
- 15 Taxol®-induced phosphatidylserine exposure and microvesicle formation in red blood cells is mediated by its vehicle Cremophor® EL. Nanomedicine 8(7), 1127–1135 (2013).
- 16 Exosomal evasion of humoral immunotherapy in aggressive B-cell lymphoma modulated by ATP-binding cassette transporter A3. Proc. Natl Acad. Sci. USA 108(37), 15336–15341 (2011).
- 17 Inhibition of oncogenic epidermal growth factor receptor kinase triggers release of exosome-like extracellular vesicles and impacts their phosphoprotein and DNA content. J. Biol. Chem. 290(40), 24534–24546 (2015).
- 18 . Endothelial expression of autocrine VEGF upon the uptake of tumor-derived microvesicles containing oncogenic EGFR. Proc. Natl Acad. Sci. USA 106(10), 3794–9 (2009).
- 19 ImClone LLC. ERBITUX (Cetuximab). www.erbitux.com.
- 20 Phase I studies of anti-epidermal growth factor receptor chimeric antibody C225 alone and in combination with cisplatin. J. Clin. Oncol. 18(4), 904–914 (2000).
- 21 . The monoclonal antibody 225 activates caspase-8 and induces apoptosis through a tumor necrosis factor receptor family-independent pathway. Oncogene 20(28), 3726–3734 (2001).
- 22 . Epidermal growth factor receptor blockade with C225 modulates proliferation, apoptosis, and radiosensitivity in squamous cell carcinomas of the head and neck. Cancer Res. 59(8), 1935–1940 (1999).
- 23 Early redistribution of plasma membrane phosphatidylserine is a general feature of apoptosis regardless of the initiating stimulus: inhibition by overexpression of Bcl–2 and Abl. J. Exp. Med. 182(5), 1545–1556 (1995).
- 24 . Analytical challenges of extracellular vesicle detection: a comparison of different techniques. Cytom. Part A
doi:10.1002/cyto.a.22795 (2015) (Epub ahead of print). - 25 Possibilities and limitations of current technologies for quantification of biological extracellular vesicles and synthetic mimics. J. Control. Release 200, 87–96 (2015).
- 26 . Extracellular vesicle sizing and enumeration by nanoparticle tracking analysis. J. Extracell. Vesicles
doi:10.3402/jev.v2i0.19671 (2013). - 27 . Effect of the anti-receptor ligand-blocking 225 monoclonal antibody on EGF receptor endocytosis and sorting. Exp. Cell Res. 312(15), 2778–2790 (2006).
- 28 Potential role of HER2–overexpressing exosomes in countering trastuzumab-based therapy. J. Cell. Physiol. 227(2), 658–667 (2012).
- 29 Abnormal lysosomal trafficking and enhanced exosomal export of cisplatin in drug-resistant human ovarian carcinoma cells. Mol. Cancer Ther. 4(10), 1595–1604 (2005).
- 30 . Expulsion of small molecules in vesicles shed by cancer cells: association with gene expression and chemosensitivity profiles. Cancer Res. 63(15), 4331–4337 (2003).
- 31 . Exosomes: composition, biogenesis and function. Nat. Rev. Immunol. 2(8), 569–579 (2002).
- 32 Analysis of AKT and ERK1/2 protein kinases in extracellular vesicles isolated from blood of patients with cancer. J. Extracell. Vesicles 3, 25657 (2014).
- 33 Highly skewed distribution of miRNAs and proteins between colorectal cancer cells and their exosomes following Cetuximab treatment: biomolecular, genetic and translational implications. Oncoscience 1(2), 132–157 (2014).
- 34 The tetraspanin CD9 modulates epidermal growth factor receptor signaling in cancer cells. J. Cell. Physiol. 216(1), 135–143 (2008).
- 35 . Exosomes packaging APOBEC3G confer human immunodeficiency virus resistance to recipient cells. J. Virol. 83(2), 512–521 (2009).
- 36 Serum-free culture alters the quantity and protein composition of neuroblastoma-derived extracellular vesicles. J. Extracell. Vesicles 1, 1–12 (2015).
- 37 . Cetuximab/C225-induced intracellular trafficking of epidermal growth factor receptor. Cancer Res. 69(15), 6179–6183 (2009).
- 38 . Exosomes in cancer development, metastasis, and drug resistance: a comprehensive review. Cancer Metastasis Rev. 32(3–4), 623–642 (2013).
- 39 Exosome release and low pH belong to a framework of resistance of human melanoma cells to cisplatin. PLoS ONE 9(2), e88193 (2014).