Osteosarcoma is the most common bone tumour and affects younger patients. The combination of chemotherapy with aggressive surgical resection results in survival rates achieving 60%-70% in patients with localized disease. Unfortunately, 30% of osteosarcoma patients present metastatic disease at diagnosis and only 20-30% will become long-term survivors. Therefore, finding new drugs to treat these patients is a research priority. Preclinical and clinical studies suggest involvement of ErbB network aberrations in the aetiology of osteosarcomas. The present study assessed the effect of Afatinib, an irreversible ErbB family blocker, in osteosarcoma cell lines.
Cell viability was assessed in osteosarcoma cell lines HOS, SJSA-1, Saos-2, U-2OS and MNNG using Afatinib at increasing concentration (0, 0.5, 1, 1.5, 2, 2.5, 3 and 5 μM). Motility was measure by scratch assay and invasion was performed using transwell chamber. Western blot was used to evaluate EGFR/HER2 pathway.
In cell viability assays Afatinib displayed micro Molar (μM) anti-proliferative activity in all cell lines tested; IC50 values (2.03 +/- 0.35) for SJSA-1, (1.62 +/- 0.38) for U-2OS, (1.8 +/- 0.24) for HOS, (2.08 +/- 0.48) for Saos-2 and (1.8 +/- 0.37) for MNNG cell lines. Motility scratch assays revealed that Afatinib is able, in a statistically significant and dose dependent manner, to reduce the width of the scratch after 24 hrs in all cell lines (p < 0.01). The scratch line width decreased by +/-45 and +/-28 microns when treated with 1 μM or 2 μM, respectively. In the Transwell cell invasion assay, Afatinib, at different concentrations was found to inhibit the invasion behavior of the non-metastatic cell line HOS and the metastatic cell line MNNG in a dose-dependent manner (p < 0.05). The number of invasive osteosarcoma cells in Afatinib treated groups was gradually reduced as the concentration of Afatinib was increased. The inhibitory effect of Afatinib on EGFR and HER2 pathway was evaluated in HOS osteosarcoma line. Afatinib showed a significant reduction in EGFR and HER2 phosphorylation and downstream signalling (p < 0.01).
Afatinib shows anti proliferative activity and decreases the migration capacity of osteosarcoma tumours cells lines.
Boehringer Ingelheim RCV GmbH & Co KG, Mexico
Boehringer Ingelheim RCV GmbH & Co KG, Mexico
M. Cruz Ramos: For this work Boehringer Ingelheim provided afatinib drug for in vitro experiments and partial financial. F. Solca: Works at Pharmacology and Translational Research, Boehringer Ingelheim RCV GmbH & Co KG. All other authors have declared no conflicts of interest.