Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Chemoproteomic selectivity profiling of PIKK and PI3K kinase inhibitors

ABSTRACT: "Here, we report the development of a novel version of Kinobeads that extends kinome coverage to PIKK and PI3K kinases. This is achieved by inclusion of two affinity probes derived from the clinical PI3K/MTOR inhibitors Omipalisib and BGT226. The new affinity matrix was used to profile 13 clinical and pre-clinical PIKK/PI3K inhibitors. The large discrepancies between the PI3K affinity values obtained and reported results from recombinant assays led us to perform a phosphoproteomic experiment showing that the chemoproteomic assay is the better approximation of PI3K inhibitor action in-vivo."

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Maria Reinecke

LAB HEAD: Bernhard Kuster

PROVIDER: PXD011719 | Pride | 2019-03-29

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	10_BGT226_final.pdf	Pdf
	10_BGT226_raw.zip	Other
	10_BGT226_txt.zip	Other
	11_CH5132799_final.pdf	Pdf
	11_CH5132799_raw.zip	Other

Items per page:

1 - 5 of 56

Publications

Chemoproteomic Selectivity Profiling of PIKK and PI3K Kinase Inhibitors.

Reinecke Maria M Ruprecht Benjamin B Poser Sandra S Wiechmann Svenja S Wilhelm Mathias M Heinzlmeir Stephanie S Kuster Bernhard B Médard Guillaume G

ACS chemical biology 20190403 4

Chemical proteomic approaches utilizing immobilized, broad-selective kinase inhibitors (Kinobeads) have proven valuable for the elucidation of a compound's target profile under close-to-physiological conditions and often revealed potentially synergistic or toxic off-targets. Current Kinobeads enrich more than 300 native protein kinases from cell line or tissue lysates but do not systematically cover phosphatidylinositol 3-kinases (PI3Ks) and phosphatidylinositol 3-kinase-related kinases (PIKKs). ...[more]

PMID: 30901187

Similar Datasets

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

Discovery of dedicated EPHA2 inhibitors - Medicinal chemistry meets chemical proteomics

Project description:The receptor tyrosine kinase EPHA2 has gained interest as therapeutical drug target in cancer and infectious diseases in the past years. However, EPHA2 research and EPHA2 based therapies have been hampered by the lack of selective small molecule EPHA2 inhibitors. We report on the synthesis and evaluation of dedicated EPHA2 inhibitors based on the clinical BCR-ABL/SRC inhibitor Dasatinib as lead structure. We designed hybrid structures of Dasatinib, CHEBI-513815, PD-173955 and a known EPHB4 inhibitor aiming at the exploitation of the ATP pocket entrance and the ribose pocket. Medicinal chemistry and inhibitor design was guided by a chemical proteomic approach allowing for early selectivity profiling of the newly synthesized inhibitor candidates. Additionally, protein crystallography delivered detailed insight into the molecular interactions that consitute structure-affinity-relationships. Finally, the anti-proliferative effect of the inhibitor candidates was confirmed in the glioblastoma cell line SF-268. In this work, we discovered a novel EPHA2 inhibitor candidate 4a comprising an improved selectivity profile while maintaining potency against EPHA2 and anti-cancer activity in SF-268 cells.

2017-05-30 | PXD006193 | Pride

Target Landscape of Clinical Kinase Inhibitors

Project description:Here, we provide a comprehensive overview of the selectivity profiles of 242 kinase inhibitors currently in clinical trials. We screened available inhibitors against human protein kinases by using the Kinobead technology resulting in a drug matrix identifying the druggable kinome and related proteins.

2017-12-01 | PXD005336 | 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

Novel Eph receptor tyrosine kinase inhibitors block colorectal cancer cell growth

Project description:Despite advances, treatment of unresectable colorectal cancer remains a clinical challenge and the incidence of this malignancy is increasing in individuals below the age of fifty. Integrated genomic, transcriptomic, proteomic, and mechanistic studies identified the tyrosine kinase receptors EphB2 and EphB4 as colorectal cancer drivers, and EphA2 activity as a cancer resistance mechanism to epidermal growth factor receptor and BRAF inhibitors. Reduction of EphB2 and EphB4 activity suppresses colorectal cancer cell growth and survival and is a potential target for colorectal cancer therapy. Here, we report the design and synthesis of two novel inhibitors, 1 and 3, that selectively bind with high affinity to A and B-type Eph receptor tyrosine kinases, inhibit A and B-type Eph tyrosine kinase activity, induce cell cycle arrest and confer a protracted state of growth reduction to colorectal cancer cells. 1 and 3 bind to EphA2 arresting the activation loop containing the conserved Asp-Phe-Gly (“DFG”)-motif in an inactive conformation and show a conserved hydrogen bond interaction with the so-called gatekeeper residue, the activation loop, alphaC helix, and hinge region of the kinase, typical of kinase inhibitors. These results demonstrate that pharmacological inhibition of Eph tyrosine kinase receptors is a valid therapeutic approach for the treatment of colorectal cancer.

2024-05-23 | PXD031880 | Pride

Single methyl group positioning opens new chemical space for EPH receptor kinase inhibition

Project description:EPHrin receptors (EPH) are transmembrane receptor tyrosine kinases. Their extracellular domains bind specifically to ephrin A/B ligands, and this binding modulates the intracellular kinase activity. EPHs are key players in bidirectional intercellular signalling, controlling cell morphology, adhesion and migration. They are increasingly recognized as cancer drug targets. We analysed the binding of the patented Novartis inhibitor NVP-BHG712 (NVP) to EPHA2 and EPHB4. Unexpectedly, all tested commercially available NVP samples were regioisomers NVPiso with a single methyl group binding to an adjacent nitrogen atom compared to the patented compound. The two compounds of identical mass reveal different binding modes. Further, both, in vitro and in vivo experiments show that the isomers differ in their kinase affinity and selectivity.

2018-09-05 | PXD009188 | Pride

SOX4 AND SMARCA4 COOPERATIVELY REGULATE PI3K SIGNALING THROUGH TRANSCRIPTIONAL ACTIVATION OF TGFBR2 IN TRIPLE NEGATIVE BREAST CANCER

Project description:Dysregulation of PI3K/Akt signaling is a dominant feature in basal-like or triple-negative breast cancers (TNBC). However, the mechanisms regulating this pathway are largely unknown in this subset of aggressive tumors. Here we demonstrate that the transcription factor SOX4 is a key regulator of PI3K signaling in TNBC. Genomic and proteomic analyses coupled with mechanistic studies identified TGFBR2 as a direct transcriptional target of SOX4 and demonstrated that TGFBR2 is required to mediate SOX4-dependent PI3K signaling. We further report that SOX4 and the SWI/SNF ATPase SMARCA4, which are uniformly overexpressed in basal-like tumors, form a previously unreported complex that is required to maintain an open chromatin conformation at the TGFBR2 regulatory regions in order to mediate TGFBR2 expression and PI3K signaling. Collectively, our findings delineate the mechanism by which SOX4 and SMARCA4 cooperatively regulate PI3K/Akt signaling and suggest that this complex may play an essential role in TNBC genesis and/or progression. Kinase enrichment proteomic analysis was performed using HCC1143 breast cancer cells treated with a control siRNA pool or a pool targeting SOX4 in biological triplicate to evaluate the effects on the functional kinome.

2021-03-03 | PXD022596 | Pride

Differential Chemoproteomics Reveals MARK2/3 as Cell Migration-Relevant Targets of the ALK Inhibitor Brigatinib

Project description:Metastasis poses a major challenge in cancer management, including EML4-ALK-rearranged non-small cell lung cancer (NSCLC). As cell migration is a critical step during metastasis, we assessed the anti-migratory activities of several clinical ALK inhibitors in NSCLC cells and observed differential anti-migratory capabilities despite similar ALK inhibition, with brigatinib displaying superior anti-migratory effects over other ALK inhibitors. Applying an unbiased in-situ mass spectrometry-based chemoproteomics approach, we determined the proteome-wide target profile of brigatinib in EML4-ALK+ NSCLC cells. Dose-dependent and cross-competitive chemoproteomics suggested MARK2 and MARK3 as relevant brigatinib kinase targets. Functional validation showed that combined pharmacological inhibition or genetic modulation of MARK2/3 inhibited cell migration. Consistently, brigatinib treatment induced inhibitory YAP1 phosphorylation downstream of MARK2/3. Collectively, our data suggest that brigatinib exhibits unusual cross-phenotype polypharmacology as despite similar efficacy for inhibiting EML4-ALK-dependent cell proliferation as other ALK inhibitors, it more effectively prevented migration of NSCLC cells due to co-targeting of MARK2/3.

2024-04-05 | PXD036716 | Pride

Proteomic and Genomic Analyses of the Rvb1 and Rvb2 Interaction Network upon Deletion of R2TP Complex Components

Project description:The Saccharomyces cerevisiae R2TP protein complex consists of Rvb1, Rvb2, Pih1 and Tah1. The R2TP complex has been implicated in various cellular processes such as assembly of snoRNP complex, RNA polymerase II complex, apoptosis and PIKK signaling. The involvement of R2TP in assembling various complexes seems to be in part due to Pih1 and Tah1, which serve as adapter/recruiter proteins. Here, we have performed high resolution RNA-seq. analyses to identify differential expression levels between wild type and PIH1 and TAH1 deletion strains of Saccharomyces cerevisiae that can help in unraveling other functions of Pih1 and Tah1. Both wild type and deletion strains contained TAP (tandem affinity purification) tag at the C-terminal end of either RVB1 or RVB2. 3 biological replicates were performed for each strains

2016-02-02 | E-GEOD-69220 | biostudies-arrayexpress

Effect of depleting GNB1L on PIKK mRNA levels to explore the regulatory mechanism of GNB1L on PIKK proteins

Project description:Our screens identified GNB1L, a high-confidence hit, as a novel positive regulator of DDR signaling. We therefore generated HEK293A-GNB1L-dTAG cells which the GNB1L-dTAG-HA protein decreased to markedly lower levels in the presence of dTAGv-1 than that in cells treated with dTAG-NEG , we next examined whether depletion of GNB1L affects DDR signaling. Notably, depletion of GNB1L diminished all IR-induced DDR signaling. Moreover, depletion of GNB1L also significantly reduced basal DDR signaling as well as PIKKs. To determine whether GNB1L depletion specifically affects PIKKs instead of other DDR factors, we carried out label-free quantitative proteomics analysis by comparing the proteomes in NEG-treated versus in dTAGv-1-treated GNB1L-dTAG cells. The data showed that GNB1L depletion specifically reduced PIKK kinases and ATRIP rather than other DDR factors. Obviously, PIKK kinases, ATRIP and mTOR are major downregulated substrates affected by GNB1L depletion. Therefore, we performed RNA-seq to rule out the possibility that GNB1L regulates PIKK protein levels via transcriptional regulation. The differentially expressed genes (DEGs) as well as the GO analysis confirmed that depleting GNB1L had no effect on PIKK mRNA levels . These data suggested that GNB1L specifically regulates PIKK protein levels instead of other DDR factors.

2023-06-23 | GSE235200 | GEO