Receptor-Driven ERK Pulses Reconfigure MAPK Signaling and Enable Persistence of Drug-Adapted BRAF-Mutant Melanoma Cells.

Luca Gerosa,Sreeram Vallabhaneni,Christopher Chidley,Jia-Yun Chen,Thomas S.C Ng,Fabian Fröhlich,Miles A Miller,Douglas A Lauffenburger,Sang Kyun Lim,Allison Claas,H Steven Wiley,Jeremy Muhlich,Lily A Chylek,Denis Schapiro,Gregory J Baker,Peter K Sorger,Tujin Shi,Ralph Weissleder,Weijun Qian ,Mariya Atanasova ,Yi Lee ,Carrie Nicora ,Gabriela Sánchez ,Rainer H Köhler

doi:10.1016/j.cels.2020.10.002

Abstract

Targeted inhibition of oncogenic pathways can be highly effective in halting the rapid growth of tumors but often leads to the emergence of slowly dividing persister cells, which constitute a reservoir for the selection of drug-resistant clones. In BRAFV600E melanomas, RAF and MEK inhibitors efficiently block oncogenic signaling, but persister cells emerge. Here, we show that persister cells escape drug-induced cell-cycle arrest via brief, sporadic ERK pulses generated by transmembrane receptors and growth factors operating in an autocrine/paracrine manner. Quantitative proteomics and computational modeling show that ERK pulsing is enabled by rewiring of mitogen-activated protein kinase (MAPK) signaling: from an oncogenic BRAFV600E monomer-driven configuration that is drug sensitive to a receptor-driven configuration that involves Ras-GTP and RAF dimers and is highly resistant to RAF and MEK inhibitors. Altogether, this work shows that pulsatile MAPK activation by factors in the microenvironment generates a persistent population of melanoma cells that rewires MAPK signaling to sustain non-genetic drug resistance.

Highlights

Mutated BRAF is found in 50% of melanomas and results in constitutive activation of the mitogen-activated protein kinase (MAPK) signaling cascade, which comprises the RAF, MEK, and ERK kinases and thereby promotes proliferation
The previously described phenomenon of MAPK rebound in drugadapted BRAF-mutant melanoma appears to be caused by pERK reactivation in rare cells found in clusters
We found that the inducibility of pERK and pMEK correlated directly with the degree of (C) Schematic of reduction in spontaneous ERK pulses achieved by co-targeting MAPK cascade components in addition to vemurafenib. (D) Receptor abundance by immunofluorescence and ELISA in A375 cells with and without vemurafenib treatment for 24 h. (E) pERK in cells exposed to different concentrations of growth factors in A375 cells exposed to vemurafenib for 24 h. (F) pERK levels in cells exposed to growth factors (100 ng/mL) for 15 min at different times after the addition of vemurafenib

Summary

Introduction

Mutated BRAF (canonically BRAFV600E) is found in 50% of melanomas and results in constitutive activation of the mitogen-activated protein kinase (MAPK) signaling cascade, which comprises the RAF, MEK, and ERK kinases and thereby promotes proliferation. Emerging evidence suggests that rapid adaptation to targeted drugs by non-genetic mechanisms promotes sustained survival of persister cells, contributes to residual disease, and facilitates emergence of resistance mutations responsible for disease recurrence in patients (Pazarentzos and Bivona, 2015; Russo et al, 2019; Cipponi et al, 2020). The role of negative feedback is well established in the case of BRAFV600E cancers: when BRAFV600E signaling is inhibited by drugs, synthesis of dual activity serinethreonine phosphatases (DUSPs) and other negative regulators of the MAPK cascade falls. This makes cells more sensitive to MAPK reactivation, for example, by growth factors in the tumor

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