Abstract
BackgroundThe RAS/RAF/MEK/ERK pathway is one of the most downregulated pathway in cancer. Inhibitors of RAF and MEK have established clinical use while ERK inhibitors recently faced the clinic. We aimed to generate resistant cell lines which could be helpful for defining new combinations able to overcome resistance.Methodsthe human NSCLC cell line NCI-H727, sensitive to both MEK and ERK inhibitors, was treated with increasing concentrations of MEK162 (as MEK inhibitor) or SCH772984 as ERK inhibitor.Resultswe successfully obtained a MEK resistant subline (H727/MEK, after 40 passages) as well as an ERK resistant subline (H727/SCH, after 18 passages). The two resistant sublines H727/MEK and H727/SCH were cross-resistant to ERK and MEK inhibitors, respectively, but not to RAF inhibitors. The sublines maintained the responsiveness to inhibitors of the parallel PI3K/akt/mTOR pathway as well as to agents with different mechanism of action. Mechanistically, treatment of sensitive and resistant cells with MEK or ERK inhibitors was able to induce a similar inhibition of ERK phosphorylation, while only in parental cells the drugs were able to induce a downregulation of S6 and RSK phosphorylation.Conclusionsthese resistant cells represent an important tool for further studies on the mechanisms of resistance and ways to overcome it.
Highlights
The RAS/RAF/MEK/ERK pathway is one of the most downregulated pathway in cancer
Concentrations inhibiting the growth by 50% (IC50) were calculated from the curves using Graphpad Prism Version 7
The cells, after a total of 40 passages in the presence of drug showed a resistance index of approximately 10 (Table 1). This resistance was maintained after several passages in vitro in drug free medium and we considered these cells (H727/ MEK) stably resistant to the drug
Summary
The RAS/RAF/MEK/ERK pathway is one of the most downregulated pathway in cancer. Inhibitors of RAF and MEK have established clinical use while ERK inhibitors recently faced the clinic. Vemurafenib (B-RAF inhibitor) has been approved for the treatment of late stage and BRAF mutated melanoma as well as for the treatment of Erdheim-Chester Disease (a rare histiocytosis marked by recurrent BRAFV600E mutations in more than half of patients). Another B-RAF inhibitor, dabrafenib received approval for the treatment of BRAF mutated melanoma while trametinib, a MEK inhibitor has been approved for. Several mechanisms associated to resistance development have been reported [10,11,12,13,14,15,16,17] They go to changes in target splicing, upregulation, changes in phosphorylation of downstream proteins, different metabolism modulation
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