Combination of pan-RAF and MEK inhibitors in NRAS mutant melanoma.

Mohammad Atefi,Deborah Jl Wong,Charles Ng,Thomas G Graeber,David Foulad,Amanda Lassen,Helena Escuin-Ordinas,Antoni Ribas,Begonya Comin-Anduix,Earl Avramis,Bjoern Titz,Michael Cerniglia

doi:10.1186/s12943-015-0293-5

Mohammad Atefi, Deborah Jl Wong + Show 10 more

Open Access

https://doi.org/10.1186/s12943-015-0293-5

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Abstract

BackgroundApproximately 20% of melanomas contain a mutation in NRAS. However no direct inhibitor of NRAS is available. One of the main signaling pathways downstream of NRAS is the MAPK pathway. In this study we investigated the possibility of blocking oncogenic signaling of NRAS by inhibiting two signaling points in the MAPK pathway.MethodsFourteen NRAS mutated human melanoma cell lines were treated with a pan-RAF inhibitor (PRi, Amgen Compd A), a MEK inhibitor (MEKi, trametinib) or their combination and the effects on proliferation, cell cycle progression, apoptosis, transcription profile and signaling of the cells were investigated.ResultsThe majority of the cell lines showed a significant growth inhibition, with high levels of synergism of the PRi and MEKi combination. Sensitive cell lines showed induction of apoptosis by the combination treatment and there was a correlation between p-MEK levels and synergistic effect of the combination treatment. Proliferation of sensitive cell lines was blocked by the inhibition of the MAPK pathway, which also blocked expression of cyclin D1. However, in resistant cell lines, proliferation was blocked by combined inhibition of the MAPK pathway and cyclin D3, which is not regulated by the MAPK pathway. Resistant cell lines also showed higher levels of p-GSK3β and less perturbation of the apoptotic profile upon the treatment in comparison with the sensitive cell lines.ConclusionsThe combination of PRi + MEKi can be an effective regimen for blocking proliferation of NRAS mutant melanomas when there is higher activity of the MAPK pathway and dependence of proliferation and survival on this pathway.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0293-5) contains supplementary material, which is available to authorized users.

Highlights

Over activity of the RAS/RAF/MEK/ERK mitogenactivated protein kinase (MAPK) pathway is the hallmark of the majority of melanomas, which is frequently due to mutations in BRAF or NRAS in approximately 50% and 20% of cases, respectively [1]
As we investigated the effect of the treatments on the published signatures of MAPK-activation and MEKactivation [8,22], we observed a similar pattern of down regulation of these signatures in both cell lines (Figure 3A)
As we investigated the effects of pan-RAF inhibitor (PRi) and MEKi on induction of apoptosis in the sensitive cell lines versus the resistant ones, the results indicated that the combination treatment, for 48 hours, caused higher levels of cleaved poly [ADP-ribose] polymerase (PARP) (20% to 46%) in the sensitive cell line (Figure 5D)

Summary

Introduction

Over activity of the RAS/RAF/MEK/ERK mitogenactivated protein kinase (MAPK) pathway is the hallmark of the majority of melanomas, which is frequently due to mutations in BRAF or NRAS in approximately 50% and 20% of cases, respectively [1]. In melanomas with BRAFV600 mutation, the MAPK pathway, and the growth of melanoma cells, can be efficiently blocked by BRAF inhibitors such as vemurafenib or dabrafenib [2,3]. RAS is the critical switch that connects the signal of activated receptor tyrosine kinases (RTKs) to the downstream signaling network the MAPK pathway. In the MAPK pathway, RAF isoforms (CRAF, BRAF and ARAF) are the direct downstream proteins of RAS [4]. In this study we investigated the possibility of blocking oncogenic signaling of NRAS by inhibiting two signaling points in the MAPK pathway

Methods

Results

Conclusion