Shikonin and its derivatives inhibit the epidermal growth factor receptor signaling and synergistically kill glioblastoma cells in combination with erlotinib

Overexpression and mutation of the epidermal growth factor receptor (EGFR) gene play a causal role in tumorigenesis and resistance to treatment of glioblastoma (GBM). EGFR inhibitors such as erlotinib are currently used for the treatment of GBM; however, their efficacy has been limited due to drug resistance. New treatment strategies are therefore urgently needed. Shikonin, a natural naphthoquinone, induces both apoptosis and necroptosis in human glioma cells, but the effectiveness of erlotinib-shikonin combination treatment as well as the underlying molecular mechanisms is unknown yet. In this study, we investigated erlotinib in combination with shikonin and 14 shikonin derivatives in parental U87MG and transfected U87MG.EGFR GBM cells. Most of the shikonin derivatives revealed strong cytotoxicity. Shikonin together with five other derivatives, namely deoxyshikonin, isobutyrylshikonin, acetylshikonin, ,-dimethylacrylshikonin and acetylalkannin showed synergistic cytotoxicity toward U87MG.EGFR in combination with erlotinib. Moreover, the combined cytotoxic effect of shikonin and erlotinib was further confirmed with another three EGFR-expressing cell lines, BS153, A431 and DK-MG. Shikonin not only dose-dependently inhibited EGFR phosphorylation and decreased phosphorylation of EGFR downstream molecules, including AKT, P44/42MAPK and PLC1, but also together with erlotinib synergistically inhibited EGFR phosphorylation in U87MG.EGFR cells as determined by Loewe additivity and Bliss independence drug interaction models. These results suggest that the combination of erlotinib with shikonin or its derivatives might be a potential strategy to overcome drug resistance to erlotinib. What's new? Patients with glioblastoma can fight back with EGFR inhibitors, such as erlotinib, but drug resistance hobbles their usefulness over the long term. Now, researchers may have found a wingman for these EGFR inhibitors. The natural naphthoquinone shikonin also kills cancer cells, and in this study, the authors investigated whether the addition of various different shikonins could boost the efficacy of erlotinib. Shikonin, they found, also inhibits EGFR signaling, and when used in combination, the two agents halted EGFR phosphorylation and killed tumor cells far more effectively than either alone.