Oncogene Mimicry as a Mechanism of Primary Resistance to BRAF Inhibitors

Sos, Martin L.; Levin, Rebecca S.; Gordan, John D.; Oses-Prieto, Juan A.; Webber, James T.; Salt, Megan B.; Hann, Byron; Burlingame, Alma L.; McCormick, Frank; Bandyopadhyay, Sourav; Shokat, Kevan M.

doi:10.1016/j.celrep.2014.07.010

Cited by 68 publications

(67 citation statements)

References 38 publications

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“…B-Raf inhibitors also paradoxically activate the mitogen-activated protein kinase (MAPK) cascade in melanoma cells expressing oncogenic mutant N-or K-Ras (4-6). Other highly promising approaches include compounds that covalently modify K-Ras proteins with a G12C mutation to abrogate effector interactions (7,8) and allosteric modulators that directly bind Ras to inhibit guanine nucleotide exchange factor (GEF)-mediated nucleotide exchange (9-11). Chronic inhibition of Ras-GEF binding reduces GTP loading of oncogenic mutant K-Ras and hence inhibits K-Ras signaling activity (11).…”

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confidence: 99%

Inhibition of Acid Sphingomyelinase Depletes Cellular Phosphatidylserine and Mislocalizes K-Ras from the Plasma Membrane

Cho

Hoeven

Zhou

et al. 2016

Molecular and Cellular Biology

143

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e K-Ras must localize to the plasma membrane for biological activity; thus, preventing plasma membrane interaction blocks KRas signal output. Here we show that inhibition of acid sphingomyelinase (ASM) mislocalizes both the K-Ras isoforms K-Ras4A and K-Ras4B from the plasma membrane to the endomembrane and inhibits their nanoclustering. We found that fendiline, a potent ASM inhibitor, reduces the phosphatidylserine (PtdSer) and cholesterol content of the inner plasma membrane. These lipid changes are causative because supplementation of fendiline-treated cells with exogenous PtdSer rapidly restores K-Ras4A and K-Ras4B plasma membrane binding, nanoclustering, and signal output. Conversely, supplementation with exogenous cholesterol restores K-Ras4A but not K-Ras4B nanoclustering. These experiments reveal different operational pools of PtdSer on the plasma membrane. Inhibition of ASM elevates cellular sphingomyelin and reduces cellular ceramide levels. Concordantly, delivery of recombinant ASM or exogenous ceramide to fendiline-treated cells rapidly relocalizes K-Ras4B and PtdSer to the plasma membrane. K-Ras4B mislocalization is also recapitulated in ASM-deficient Neimann-Pick type A and B fibroblasts. This study identifies sphingomyelin metabolism as an indirect regulator of K-Ras4A and K-Ras4B signaling through the control of PtdSer plasma membrane content. It also demonstrates the critical and selective importance of PtdSer to K-Ras4A and K-Ras4B plasma membrane binding and nanoscale spatial organization. Ras proteins are small guanine nucleotide binding proteins that oscillate between active GTP-bound and inactive GDP-bound states. Activated Ras proteins transmit signals for cell proliferation and cell survival. Importantly, ϳ15% of all human tumors express mutant Ras proteins that are locked in the GTP-bound state (1). Of the three ubiquitously expressed Ras isoforms, H-, N-, and K-Ras, oncogenic mutant K-Ras is the most prevalent, being expressed in ϳ95% of pancreatic, ϳ45% of colorectal, and ϳ35% of lung cancers (1). Despite its importance, there are currently no clinically approved drugs that directly target oncogenic K-Ras. To date, Ras drug discovery efforts have focused largely on inhibitors of Ras downstream effectors, including B-Raf, C-Raf, phosphatidylinositol 3-kinase (PI3K), MEK, and extracellular signal-regulated kinase (ERK) (2). For example, B-Raf-specific inhibitors produce excellent albeit often short-lived responses in patients with B-Raf mutant melanoma (3), in part because of a perturbation of complex negative-feedback control loops (2). B-Raf inhibitors also paradoxically activate the mitogen-activated protein kinase (MAPK) cascade in melanoma cells expressing oncogenic mutant N-or K-Ras (4-6). Other highly promising approaches include compounds that covalently modify K-Ras proteins with a G12C mutation to abrogate effector interactions (7,8) and allosteric modulators that directly bind Ras to inhibit guanine nucleotide exchange factor (GEF)-mediated nucleotide exchange (9-11). Chronic i...

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mentioning

confidence: 99%

Inhibition of Acid Sphingomyelinase Depletes Cellular Phosphatidylserine and Mislocalizes K-Ras from the Plasma Membrane

Cho

Hoeven

Zhou

et al. 2016

Molecular and Cellular Biology

143

View full text Add to dashboard Cite

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“…Once those treated patients obtained acquired resistance, new therapeutics should be applied [85]. The primary resistance is beyond the scope of this review [86]. Based on whether other signal pathways participate in acquired resistance, acquired resistance has three simplified conditions.…”

Section: Acquired Resistance Mechanism Of Brafi and Mekimentioning

confidence: 99%

The RAF-MEK-ERK Pathway: Targeting ERK to Overcome Obstacles of Effective Cancer Therapy

et al. 2015

Future Med. Chem.

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“…Online targeted detection of endogenous kinases by a highly multiplexed label-free PRM method provides high sensitivity and specificity of detection compatible with automated data processing. MIB Kinase Enrichment and Tryptic Digestion-Compounds for Multiplex inhibitor beads were synthesized or commercially purchased and conjugated to resin as previously described (8). 10 mg of total protein per condition from lysed cell pellets was diluted to 10 ml final volume in high salt buffer (50 mM HEPES pH 7.5, 0.5% Triton X-100, 1 M NaCl, 1 mM EDTA, 1 mM EGTA) containing protease and phosphatase inhibitors (Roche).…”

Section: Methodsmentioning

confidence: 99%

“…To date, several workflows focused on this task have been described. Typically, around 110 -125 kinases can be uniquely quantified by data dependent acquisition (DDA) LC-MS/MS in any single MIB enrichment with stringent exclusion of peptides shared by multiple kinases (6,8). In addition, isobaric tags for relative and absolute quantitation (iTRAQ) (9), have been used downstream of MIB enrichment.…”

mentioning

confidence: 99%

An Optimized Chromatographic Strategy for Multiplexing In Parallel Reaction Monitoring Mass Spectrometry: Insights from Quantitation of Activated Kinases

Tlsty

Levin

Gordan

et al. 2017

Molecular & Cellular Proteomics

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Kinases are involved in the regulation of most cellular processes and are implicated in numerous human disease states. Five hundred and eighteen human kinases are known, and between 200 and 400 are typically expressed in a given cell type (1-3). Quantitative measurement of kinase activity is of critical importance in numerous biological applications, including studies of cellular signaling, cancer biomarker discovery, and drug development. An ideal strategy for functional kinase analysis must combine an active state kinase enrichment component and a methodology to quantify their levels of activation, because kinase regulation is a major determinant of cellular function and disease phenotypes.We and other groups have previously reported on the development and use of Multidrug Inhibitor Beads (MIB) 1 strategy for active kinase enrichment (4 -8). In this approach, kinase affinity enrichment scaffolds based on known kinase inhibitor structures targeted to conformationally active kinases are chemically coupled to a resin, which is used to capture broad classes of kinases from cell lysates (see recent review of this and related approaches by Daub (4)). When

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Oncogene Mimicry as a Mechanism of Primary Resistance to BRAF Inhibitors

Cited by 68 publications

References 38 publications

Inhibition of Acid Sphingomyelinase Depletes Cellular Phosphatidylserine and Mislocalizes K-Ras from the Plasma Membrane

Inhibition of Acid Sphingomyelinase Depletes Cellular Phosphatidylserine and Mislocalizes K-Ras from the Plasma Membrane

The RAF-MEK-ERK Pathway: Targeting ERK to Overcome Obstacles of Effective Cancer Therapy

An Optimized Chromatographic Strategy for Multiplexing In Parallel Reaction Monitoring Mass Spectrometry: Insights from Quantitation of Activated Kinases

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