Therapeutic antitumor efficacy of anti-epidermal growth factor receptor antibody, cetuximab, against malignant pleural mesothelioma

Abstract

Epidermal growth factor receptor (EGFR) is commonly overexpressed in malignant pleural mesothelioma (MPM). Cetuximab is a chimeric mouse-human antibody targeted against EGFR and induces potent antibody-dependent cellular cytotoxicity (ADCC). The action of cetuximab against MPM cells has not been well studied. Therefore, in this study, we investigated the antitumor activity of cetuximab against MPM cell lines, particularly with respect to ADCC activity in vitro and in vivo. EGFR expression of MPM cells was measured by a quantitative flow cytometric analysis and immunohistochemistry. The effect of cetuximab on growth inhibition was assessed using a modified MTT assay. The ADCC activity was measured by a 4-h 51Cr release assay using fresh or IL-2-activated peripheral blood mononuclear cells. In vivo antitumor activity of cetuximab was evaluated using an orthotopic implantation mouse model. Cetuximab-mediated ADCC activity against MPM cells was observed at low concentration (0.25 mg/ml) and was enhanced by IL-2, whereas no direct effect on growth inhibition was detected. A logarithmic correlation was observed between the number of EGFRs on MPM cells and ADCC activity. Low EGFR expression on the MPM cells, which was weakly detectable by immunohistochemistry, was sufficient for maximum ADCC activity. In the mouse model, cetuximab treatment with or without IL-2 significantly inhibited intrathoracic tumor growth and prolonged their survival. Our study shows that cetuximab has potent anti-MPM activity both in vitro and in vivo, mainly through the immunologic mechanism of ADCC. Cetuximab has the potential to be used as a novel therapy for MPM patients.

Mutation analysis of the EGFR gene and downstream signalling pathway in histologic samples of malignant pleural mesothelioma

“…Epidermal growth factor receptor TKI inhibitors, such as gefitinib and erlotinib, inhibit MPM cell migration and proliferation, enhance the response to radiation of human MPM cell lines, and reduce motility and invasion in MPM cell lines (Kurai et al , 2012). However, the promising results obtained in in vitro studies were not reproduced in two phase II trials involving patients with pleural and peritoneal mesotheliomas, although it should be noted that neither study evaluated the mutation status of the EGFR gene and its downstream signalling transduction pathway (Govindan et al , 2005; Garland et al , 2007).…”

“…No published studies have assessed the in vivo effects of anti-EGFR mAbs on MPMs, although one recent study has found that cetuximab is highly efficacious in cultured MPM cell lines (Kurai et al , 2012). It has been demonstrated that mutations in EGFR downstream pathways can affect the efficacy of EGFR mABs in other tumours such as colorectal adenocarcinoma (Jonker et al , 2007), and we found nine patients (11.7%) with missense mutations involving the KRAS ( n =5), BRAF ( n =3), and PIK3CA genes ( n =1).…”

“…One recent study has shown that cetuximab effectively blocks the growth of MPM cells in cell cultures and mouse models (Kurai et al , 2012) and, as in the case of colorectal cancer and lung adenocarcinomas, the potential efficacy of these TKIs in MPM may depend on the mutation status of EGFR gene and its downstream effectors (Lièvre et al , 2006; Pirker et al , 2011). …”

Nicotine promotes lymph node metastasis and cetuximab resistance in head and neck squamous cell carcinoma

“…HUVECs were cultured in endothelial cell growth medium-2 (Takara Bio, Inc.) at 37˚C. Nicotine (Sigma-Aldrich; Merck KGaA; 0.5 µM) (19,20,22), mecamylamine hydrochloride (MCA; Sigma-Aldrich; Merck KGaA; 50 nM) (22), α-bungarotoxin (α-BTX; Abcam, Cambridge, UK; 1 µM) (17), cetuximab (Merck KGaA; 0.5 µg/ml) (30), SCH772984 (Santa Cruz Biotechnology, Inc., Dallas, TX, USA; 1 µM) (25), Akt inhibitor II (Merck KGaA; 10 µM) (26) and temsirolimus (Pfizer, Inc., New York, NY, USA; 10 nM) (27) were purchased. The concentration of nicotine (0.5 µM) used was decided based on previously published basic studies (19,20,22) and clinical investigations in which the concentrations of nicotine in the bloodstream of smokers exposed to nicotine at pharmacological concentrations were reported to be 0.09-1 µM (28,29).…”

“…After becoming subconfluent at 37˚C, the cells were cultured for 24 h in DMEM/F-12 without FBS. They were then cultured in the presence or absence of nicotine (0.5 µM) (19,20,22), MCA (50 nM) (22), α-BTX (1 µM) (17) or cetuximab (0.5 µg/ml) (30) in DMEM/F-12 supplemented with 0.5% FBS. The number of cells was counted with TC10TM automated cell counter (Bio-Rad Laboratories, Inc., Hercules, CA, USA) daily for 5 days.…”