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Aquired resistance mechanisms to EGFR-specific kinase inhibitors using chemical proteomics

ABSTRACT: Here we used kinase affinity enrichment (Kinobeads) and intensity-based mass spectrometry to elucidate aquired resistance mechanisms to EGFR inhibition in cancer cells. First, kinase expression differences between untreated control cells and long term treated resistant cells were investigated using Kinobeads and LC-MS/MS. Overexpressed Kinases were then further investigated by cellular assays to evaluate targets and development of combination treatment strategies.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

DISEASE(S): Non-small Cell Lung Carcinoma

SUBMITTER: Heiner Koch

LAB HEAD: Prof. Bernhard Kuster

PROVIDER: PXD001825 | Pride | 2015-05-20

REPOSITORIES: Pride

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Chemical Proteomics Uncovers EPHA2 as a Mechanism of Acquired Resistance to Small Molecule EGFR Kinase Inhibition.

Koch Heiner H Busto M Estela Del Castillo ME Kramer Karl K Médard Guillaume G Kuster Bernhard B

Journal of proteome research 20150526 6

Tyrosine kinase inhibitors (TKIs) have become an important therapeutic option for treating several forms of cancer. Gefitinib, an inhibitor of the epidermal growth factor receptor (EGFR), is in clinical use for treating non-small cell lung cancer (NSCLC) harboring activating EGFR mutations. However, despite high initial response rates, many patients develop resistance to gefitinib. The molecular mechanisms of TKI resistance often remain unclear. Here, we describe a chemical proteomic approach co ...[more]

PMID: 25963923

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2017-10-13 | PXD007920 | Pride

Defining the landscape of EPHA2 Inhibition by Clinical Kinase Inhibitors

Project description:The receptor tyrosine kinase EPHA2 plays important roles in oncogenesis, metastasis and treatment resistance but therapeutic targeting, drug discovery or investigation of EPHA2 biology is hampered by the lack of appropriate inhibitors and structural information on how these interact with the protein. Here, we used chemical proteomics to survey 235 clinical kinase inhibitors and identified 24 with sub-micromolar potency for EPHA2. We generated nine high resolution co-crystal structures to identify drug-protein interactions at the atomic level. The combination of compound selectivity, structure determinantion and kinome-wide sequence alignment allowed us to develop a classification system in which amino acids in the drug binding site are categorized into key, scaffold, potency and selectivity residues. This scheme should be generally applicable in kinase drug discovery and we anticipate that the provided information will greatly facilitate the development of selective EPHA2 inhibitors in particular and the repurposing of clinical kinase inhibitors in general.

2016-12-23 | PXD004112 | Pride

Chemical Proteomics Reveals Ferrochelatase as a Common Off-target of Kinase Inhibitors

Project description:Protein kinases are key components in signal transduction pathways and are established drug targets in oncology. Consequently, small molecule kinase inhibitors are on the rise but often show a rather broad target spectrum, potentially leading to toxic side effects. As a result, a broad assessment of the target space is desirable for proper interpretation of observed biological effects. The enzyme Ferrochelatase (FECH), which catalyzes the conversion of protoporphyrin IX into heme, was recently found to be an off-target of the BRAF inhibitor Vemurafenib potentially explaining the often severe phototoxicity associated with this drug in melanoma patients. However, the extent to which kinase inhibitors bind to FECH in general is currently unclear. Here, we used a chemical proteomics approach based on the kinobead technology to profile 226 clinical kinase inhibitors for their potential to bind FECH. Surprisingly, low or sub-micromolar FECH binding was detected for 29 (13%) of all compounds tested and isothermal dose response measurements confirmed drug binding to FECH in cells. We also show that Vemurafenib, Linsitinib, Neratinib and MK-2461 reduce heme levels in K562 cells, verifying that drug binding leads to loss of FECH activity. Further experiments identified the protoporphyrin pocket in FECH as one major binding site for small molecule inhibitors. Since genetic loss of FECH function leads to photosensitivity in humans, we suggest that FECH inhibition by kinase inhibitors is the molecular mechanism triggering photosensitivity in patients and should therefore be part of the pre-clinical tox package for kinase inhibitors.

2017-03-22 | PXD003373 | Pride

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2018-05-02 | PXD008632 | Pride

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2017-05-30 | PXD006193 | Pride

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2017-07-05 | E-MTAB-4548 | biostudies-arrayexpress

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