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

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

doi:10.1016/j.cels.2020.10.002

Cited by 87 publications

(117 citation statements)

References 72 publications

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“…Further adding an EGFR inhibitor generated high response rates and promising clinical results (Kopetz et al, 2019;Van Cutsem et al, 2019). Heterogeneity of RTK expression in genetically identical cells can also result in resistance to RAF inhibitors (Gerosa et al, 2020). Using RAF as an example, we show that negative and positive feedback loops cannot lead to a complete, steady-state revival of the pathway output activity unless the inhibitor can induce RAF dimerization (Figures 5B and 5C).…”

Section: Discussionmentioning

confidence: 91%

A systematic analysis of signaling reactivation and drug resistance

et al. 2021

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A systematic analysis of signaling reactivation and drug resistance Graphical abstract Highlights d Feedback loops cannot fully buffer drug perturbations and recover signaling output d Activating and inhibitory paths from a drug target can reactivate signaling output d Drug target dimerization combined with feedback can restore signaling output d Combining type I½ and II RAF inhibitors quells ERK reactivation in NRAS mutant cancers

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Section: Discussionmentioning

confidence: 91%

A systematic analysis of signaling reactivation and drug resistance

et al. 2021

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“…The sporadic entry of cells into S phase during drug-imposed G1 arrest has been reported in various cell culture systems ( 31, 43 ), and downregulation of high fidelity replication and repair genes has been observed in drug-arrested G1 populations ( 33, 35 ), suggesting that DNA replication could invoke a general increase in mutability. Here we examined gene expression changes in cells treated with MEKi that escape G1 and progress to S and G2, the cell cycle phases in which DNA replication and high-fidelity DNA repair factors are active.…”

Section: Discussionmentioning

confidence: 99%

“…We therefore hypothesised that cells occasionally enter the cell cycle even in the presence of selumetinib, and that these cells are at risk of genome instability. Indeed, such events were recently reported during treatment with BRAF and EGFR inhibitors (31,43).…”

Section: Individual Cells Enter the Cell Cycle Even Under Acute Mek Imentioning

confidence: 93%

“…A substantial number of genes involved in DNA replication are regulated indirectly by the ERK1/2 pathway, and suppression of ERK1/2 signalling through EGFR inhibition downregulates DNA repair genes (35,(44)(45)(46). MEK inhibition could therefore cause impaired and error prone DNA replication if not reversed on entry to S phase, but transient ERK1/2 reactivation may coincide with escape from G1 arrest as reported for BRAF inhibition (43). We therefore assayed ERK1/2 activation by high-throughput imaging for ERK1/2 phosphorylated at T202/Y204 (pERK).…”

Section: Abnormal Gene Expression During Cell Cycle Progression In Sementioning

confidence: 96%

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DNA replication during acute MEK inhibition drives acquisition of resistance through amplification of theBRAFoncogene

Channathodiyil

Segonds‐Pichon

Smith

et al. 2021

Preprint

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Mutations and gene amplifications that confer drug resistance emerge frequently during chemotherapy, but their mechanism and timing is poorly understood. Here, we investigate BRAFV600E amplification events that underlie resistance to the MEK inhibitor selumetinib (AZD6244/ARRY-142886) in COLO205 cells. We find that de novo focal BRAF amplification is the primary path to resistance irrespective of pre-existing amplifications. Although selumetinib causes long-term G1 arrest, we observe that cells stochastically re-enter the cell cycle during treatment without reactivation of ERK1/2 or induction of a normal proliferative gene expression programme. Genes encoding DNA replication and repair factors are downregulated during G1 arrest, but many are transiently induced when cells escape arrest and enter S and G2. Nonetheless, mRNAs encoding key DNA replication factors including the MCM replicative helicase complex, PCNA and TIPIN remain at very low abundance, which likely explains previous reports of replication stress and mutagenesis under long-term RAF-MEK-ERK1/2 pathway inhibition. To test the hypothesis that DNA replication in drug promotes de novo BRAF amplification, we exploited the combination of palbociclib and selumetinib to reinforce the G1 arrest. Using a palbociclib dose that suppresses cell cycle entry during selumetinib treatment but not during normal proliferation, we show that combined treatment robustly delays the emergence of drug resistant colonies. Our results demonstrate that acquisition of MEK inhibitor resistance can occur through de novo gene amplification events resulting from DNA replication in drug, and is suppressed by drug combinations that impede cell cycle entry.

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“…Examples include the signaling pulses observed from the tumor suppressor p53, the mitogen associated protein kinase (MAPK) Erk, and the immune signaling transcription factor NF-κB [1][2][3][4][5] . Pulses of Erk activity have been observed in vivo in the early mouse embryo 6,7 and in 53 tumors 8 , and self-organize into propagating waves from sites of epithelial injury in both mouse 9 54 and zebrafish 10 . The breadth of biological systems exhibiting signaling dynamics suggests that they may play important functional roles.…”

Section: Main Textmentioning

confidence: 99%

A synthetic gene circuit for imaging-free detection of dynamic cell signaling

Ravindran

McFann

Toettcher

2021

Preprint

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Cells employ intracellular signaling pathways to sense and respond to changes in their external environment. In recent years, live-cell biosensors have revealed complex pulsatile dynamics in many pathways, but studies of these signaling dynamics are limited by the necessity of live-cell imaging at high spatiotemporal resolution1. Here, we describe an approach to infer pulsatile signaling dynamics from just a single measurement in fixed cells using a pulse-detecting gene circuit. We computationally screened for circuit with pulse detecting capability, revealing an incoherent feedforward topology that robustly performs this computation. We then implemented the motif experimentally for the Erk signaling pathway using a single engineered transcription factor and fluorescent protein reporter. Our ‘recorder of Erk activity dynamics’ (READer) responds sensitively to both spontaneous and stimulus-driven Erk pulses. READer circuits thus open the door to permanently labeling transient, dynamic cell populations to elucidate the mechanistic underpinnings and biological consequences of signaling dynamics.

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

Cited by 87 publications

References 72 publications

A systematic analysis of signaling reactivation and drug resistance

A systematic analysis of signaling reactivation and drug resistance

DNA replication during acute MEK inhibition drives acquisition of resistance through amplification of theBRAFoncogene

A synthetic gene circuit for imaging-free detection of dynamic cell signaling

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