COT drives resistance to RAF inhibition through MAP kinase pathway reactivation

Johannessen, Cory M.; Boehm, Jesse S.; Kim, So Young; Thomas, Sapana R.; Wardwell, Leslie; Johnson, Laura A.; Emery, Caroline M.; Stransky, Nicolas; Cogdill, Alexandria P.; Barretina, Jordi; Caponigro, Giordano; Hieronymus, Haley; Murray, Ryan R.; Salehi‐Ashtiani, Kourosh; Hill, David E.; Vidal, Marc; Zhao, Jean J.; Yang, Xiaoping; Alkan, Ozan; Kim, Sung-Joon; Harris, Jennifer L.; Wilson, Christopher J.; Myer, Vic E.; Finan, Peter M.; Root, David E.; Roberts, Thomas M.; Golub, Todd R.; Flaherty, Keith T.; Dummer, Reinhard; Weber, Barbara; Sellers, William R.; Schlegel, Robert; Wargo, Jennifer A.; Hahn, William C.; Garraway, Levi A.

doi:10.1038/nature09627

Cited by 1,330 publications

(1,145 citation statements)

References 29 publications

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“…5,24), with recent clinical studies reporting that highly specific BRAF inhibitors are effective in the treatment of metastatic melanoma (25)(26)(27). However, initial promise has been hampered by the development of resistance (28)(29)(30), which is characterized by the reactivation of ERK1/2 (31)(32)(33) and has been attributed to various mechanisms including activating NRAS mutations (29), CRAF overexpression (34), compensatory upregulation of MAP2K kinase COT (28), activating MEK1 mutations (35), and amplification of mutant BRAF (36).…”

Section: Introductionmentioning

confidence: 99%

Isolation of a Novel Thioflavin S–Derived Compound That Inhibits BAG-1–Mediated Protein Interactions and Targets BRAF Inhibitor–Resistant Cell Lines

Enthammer

Papadakis

Gachet

et al. 2013

Molecular Cancer Therapeutics

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Protein-protein interactions mediated through the C-terminal Bcl-2-associated athanogene (BAG) domain of BAG-1 are critical for cell survival and proliferation. Thioflavin S (NSC71948)-a mixture of compounds resulting from the methylation and sulfonation of primulin base-has been shown to dosedependently inhibit the interaction between BAG-1 and Hsc70 in vitro. In human breast cancer cell lines, with high BAG-1 expression levels, Thioflavin S reduces the binding of BAG-1 to Hsc70, Hsp70, or CRAF and decreases proliferation and viability. Here, we report the development of a protocol for the purification and isolation of biologically active constituents of Thioflavin S and the characterization of the novel compound Thio-2. Thio-2 blocked the growth of several transformed cell lines, but had much weaker effects on untransformed cells. Thio-2 also inhibited the proliferation of melanoma cell lines that had become resistant to treatment with PLX4032, an inhibitor of mutant BRAF. In transformed cells, Thio-2 interfered with intracellular signaling at the level of RAF, but had no effect on the activation of AKT. Thio-2 decreased binding of BAG-1 to Hsc70 and to a lesser extent BRAF in vitro and in vivo, suggesting a possible mechanism of action. Given that tumors frequently develop resistance to kinase inhibitors during treatment, Thio-2 and related compounds may offer promising alternative strategies to currently available therapies.

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Section: Introductionmentioning

confidence: 99%

Isolation of a Novel Thioflavin S–Derived Compound That Inhibits BAG-1–Mediated Protein Interactions and Targets BRAF Inhibitor–Resistant Cell Lines

Enthammer

Papadakis

Gachet

et al. 2013

Molecular Cancer Therapeutics

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“…Levi Garraway, an oncologist at the Dana-Farber Cancer Institute in Boston, Massachusetts, and his colleagues have now found that excessive production of a cancer-promoting protein called COT can shield cultured cells from PLX4032. High levels of COT were also found in two of three PLX4032-resistant tumours taken from patients who had received the drug 3 .…”

Section: By H E I D I L E D F O R Dmentioning

confidence: 90%

The roots of resistance

Ledford¹

2010

Nature

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“…For V5-A-Raf, A-Raf from pDONR223-ARAF (Addgene Plasmid 23725) 86 was fused N-terminal with a V5-tag of pcDNA6.2/V5-DEST using Gatway technology.…”

Section: Methodsmentioning

confidence: 99%

Differential localization of A-Raf regulates MST2-mediated apoptosis during epithelial differentiation

et al. 2016

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A-Raf belongs to the family of oncogenic Raf kinases that are involved in mitogenic signaling by activating the MEK-ERK pathway. Low kinase activity of A-Raf toward MEK suggested that A-Raf might have alternative functions. We recently identified A-Raf as a potent inhibitor of the proapoptotic mammalian sterile 20-like kinase (MST2) tumor suppressor pathway in several cancer entities including head and neck, colon, and breast. Independent of kinase activity, A-Raf binds to MST2 thereby efficiently inhibiting apoptosis. Here, we show that the interaction of A-Raf with the MST2 pathway is regulated by subcellular compartmentalization. Although in proliferating normal cells and tumor cells A-Raf localizes to the mitochondria, differentiated non-carcinogenic cells of head and neck epithelia, which express A-Raf at the plasma membrane. The constitutive or induced re-localization of A-Raf to the plasma membrane compromises its ability to efficiently sequester and inactivate MST2, thus rendering cells susceptible to apoptosis. Physiologically, A-Raf re-localizes to the plasma membrane upon epithelial differentiation in vivo. This re-distribution is regulated by the scaffold protein kinase suppressor of Ras 2 (KSR2). Downregulation of KSR2 during mammary epithelial cell differentiation or siRNA-mediated knockdown re-localizes A-Raf to the plasma membrane causing the release of MST2. By using the MCF7 cell differentiation system, we could demonstrate that overexpression of A-Raf in MCF7 cells, which induces differentiation. Our findings offer a new paradigm to understand how differential localization of Raf complexes affects diverse signaling functions in normal cells and carcinomas. A-Raf is a member of the Raf family of serine-threonine protein kinases, which comprises A-Raf, B-Raf, and Raf-1. Raf kinases are at the apex of the three-tiered Raf/MEK/ extracellular signal-regulated kinase (ERK) (mitogen-activated protein kinase (MAPK)) pathway regulating fundamental cellular functions, including differentiation, transformation, apoptosis, proliferation, and metabolism. 1,2 Activation of Ras GTPases at the cell membrane initiates Raf kinase activation and sequential phosphorylation and activation of the serinethreonine kinases MEK1/2 and ERK1/2. 3,4 In comparison to Raf-1 and B-Raf, A-Raf is only weakly activated by oncogenic H-Ras and Src 5 and is a poor MEK kinase, 5-8 which is due to unique non-conserved amino acid substitutions in the N-region. 9 Only recently, the first somatic oncogenic mutations of A-Raf were identified in lung adenocarcinomas 10 and Langerhans cell histiocytosis. 11,12 We recently showed that A-Raf and Raf-1, independent of their kinase activity, bind the proapoptotic mammalian sterile 20-like kinase (MST2) thereby suppressing MST2 activation and MST2-induced apoptosis. 13-17 MST2 employs several mechanisms to induce apoptosis including the transcriptional induction of PUMA, 14 a BH3 domain protein that causes mitochondrial depolarization and subsequent cell death. 18 Although Raf-1 counteracts MS...

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COT drives resistance to RAF inhibition through MAP kinase pathway reactivation

Cited by 1,330 publications

References 29 publications

Isolation of a Novel Thioflavin S–Derived Compound That Inhibits BAG-1–Mediated Protein Interactions and Targets BRAF Inhibitor–Resistant Cell Lines

Isolation of a Novel Thioflavin S–Derived Compound That Inhibits BAG-1–Mediated Protein Interactions and Targets BRAF Inhibitor–Resistant Cell Lines

The roots of resistance

Differential localization of A-Raf regulates MST2-mediated apoptosis during epithelial differentiation

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