Cell-Based Proteome Profiling of Potential Dasatinib Targets by Use of Affinity-Based Probes

Shi, Haibin; Zhang, Chong‐Jing; Chen, Grace Y. J.; Yao, Shao Q.

doi:10.1021/ja208518u

Cited by 205 publications

(216 citation statements)

References 77 publications

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“…To overcome this limitation, several groups have used activity-based protein profiling (ABPP) combined with bio-orthogonal click chemistry to identify drug targets both in vitro and in vivo (supplemental Fig. S1) (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15). ABPP probes exert their functions via covalent reactions with the target proteins or photoaffinity-based labeling via the incorporation of photoreactive groups.…”

mentioning

confidence: 99%

A Quantitative Chemical Proteomics Approach to Profile the Specific Cellular Targets of Andrographolide, a Promising Anticancer Agent That Suppresses Tumor Metastasis

Wang

Tan

Nguyễn

et al. 2014

Molecular & Cellular Proteomics

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Drug target identification is a critical step toward understanding the mechanism of action of a drug, which can help one improve the drug's current therapeutic regime and expand the drug's therapeutic potential. However, current in vitro affinity-chromatography-based and in vivo activity-based protein profiling approaches generally face difficulties in discriminating specific drug targets from nonspecific ones. Here we describe a novel approach combining isobaric tags for relative and absolute quantitation with clickable activity-based protein profiling to specifically and comprehensively identify the protein targets of andrographolide (Andro), a natural product with known anti-inflammation and anti-cancer effects, in live cancer cells. We identified a spectrum of specific targets of Andro, which furthered our understanding of the mechanism of action of the drug. Our findings, validated through cell migration and invasion assays, showed that Andro has a potential novel application as a tumor metastasis inhibitor. Moreover, we have unveiled the target binding mechanism of Andro with a combination of drug analog synthesis, protein engineering, and mass-spectrometry-based approaches and determined the drugbinding sites of two protein targets, NF-B and actin. Molecular & Cellular Proteomics 13: 10.1074/mcp. M113.029793, 876-886, 2014.As most drugs exert pharmacological effects by interacting with their target proteins, the identification of these target proteins is a critical step in unraveling the mechanisms of drug action. It is also imperative for our understanding of the pharmacodynamics of a known drug, suggesting potentially unrevealed actions and thus refining future clinical applications of the substance. Traditional approaches used to identify protein targets of a drug typically utilize immobilized drug affinity chromatography coupled with mass spectrometry (MS) 1 (1, 2). These methods can be applied to cell lysates, but not in an in vivo setting, because of the requirement of a solid support. In vitro target profiling might not accurately reflect the drug's actions in the in vivo physiological environment. To overcome this limitation, several groups have used activity-based protein profiling (ABPP) combined with bio-orthogonal click chemistry to identify drug targets both in vitro and in vivo (supplemental Fig. S1) (3-15). ABPP probes exert their functions via covalent reactions with the target proteins or photoaffinity-based labeling via the incorporation of photoreactive groups. With the increasing sensitivity of modern MS platforms, low-abundance protein targets can be successfully identified. Although both conventional affinity chromatography and recent ABPP-based methods allow us to detect a set of candidate protein targets for a drug, it remains difficult to 1 The abbreviations used are: MS, mass spectrometry; ABPP, activity-based protein profiling; ICABPP, clickable activity-based protein profiling; iTRAQ, isobaric tags for relative and absolute quantitation; DMSO, dimethyl sulfoxide; Andro, androgr...

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mentioning

confidence: 99%

A Quantitative Chemical Proteomics Approach to Profile the Specific Cellular Targets of Andrographolide, a Promising Anticancer Agent That Suppresses Tumor Metastasis

Wang

Tan

Nguyễn

et al. 2014

Molecular & Cellular Proteomics

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“…The tag may incorporate a moiety, such as an alkyne or azide, for subsequent modification, such as by the Cu(I)-catalyzed azide-alkyne [3+2] cycloaddition (CuAAC), to introduce a reporter (15,16). ABPP probes have been developed to monitor changes of specific enzymes associated with certain biological states, including serine hydrolases (17,18), cysteine proteases (19)(20)(21)(22), protein phosphatases (23)(24)(25), oxidoreductases (26), histone deacetylases (27), kinases (28,29), metalloproteases (30)(31)(32), and glycosidases (33)(34)(35)(36).…”

mentioning

confidence: 99%

Cell-permeable probe for identification and imaging of sialidases

Tsai

Yen

Lin

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Alkyne-hinged 3-fluorosialyl fluoride (DFSA) containing an alkyne group was shown to be a mechanism-based target-specific irreversible inhibitor of sialidases. The ester-protected analog DFSA (PDFSA) is a membrane-permeable precursor of DFSA designed to be used in living cells, and it was shown to form covalent adducts with virus, bacteria, and human sialidases. The fluorosialyl–enzyme adduct can be ligated with an azide-annexed biotin via click reaction and detected by the streptavidin-specific reporting signals. Liquid chromatography-mass spectrometry/mass spectrometry analysis on the tryptic peptide fragments indicates that the 3-fluorosialyl moiety modifies tyrosine residues of the sialidases. DFSA was used to demonstrate influenza infection and the diagnosis of the viral susceptibility to the anti-influenza drug oseltamivir acid, whereas PDFSA was used for in situ imaging of the changes of sialidase activity in live cells.

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“…This in situ capturing of interactions between Dasatinib and its cellular targets was shown to be more effective than the previous approach performed in cell lysates. [39] In order to minimize interference upon binding to the target proteins, the same group developed Dasatinib and Staurosporine probes with "minimalist" linkers, in which both the photo-reactive and reporter groups are made as small as possible.…”

Section: Activity-and Affinity-based Probes For Proteomic Profilingmentioning

confidence: 99%

“…[70] Twenty specific targets of this b-lactone probe library were identified as different enzymes belonging to four major families including ligases, oxidoreductases, hydrolases, and transferases. [71] Later, the same group carried out an elegant study by extending this "labelfree" design to another group of natural products: b-lactams, including natural b-lactam probes based on the antibiotics penicillin (37), aztreonam (38), and cephalosporin (39). [72] The synthesized b-lactam probe library (40) was applied to label and identify b-lactam-binding enzymes under in vivo conditions.…”

Section: Abpp Probes Derived From and Inspired By Natural Products Fomentioning

confidence: 99%

Activity‐Based Protein Profiling: Recent Advances in Probe Development and Applications

Yang

2015

ChemBioChem

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The completion of the human genome sequencing project has provided a wealth of new information regarding the genomic blueprint of the cell. Although, to date, there are roughly 20,000 genes in the human genome, the functions of only a handful of proteins are clear. The major challenge lies in translating genomic information into an understanding of their cellular functions. The recently developed activity-based protein profiling (ABPP) is an unconventional approach that is complementary for gene expression analysis and an ideal utensil in decoding this overflow of genomic information. This approach makes use of synthetic small molecules that covalently modify a set of related proteins and subsequently facilitates identification of the target protein, enabling rapid biochemical analysis and inhibitor discovery. This tutorial review introduces recent advances in the field of ABPP and its applications.

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Cell-Based Proteome Profiling of Potential Dasatinib Targets by Use of Affinity-Based Probes

Cited by 205 publications

References 77 publications

A Quantitative Chemical Proteomics Approach to Profile the Specific Cellular Targets of Andrographolide, a Promising Anticancer Agent That Suppresses Tumor Metastasis

A Quantitative Chemical Proteomics Approach to Profile the Specific Cellular Targets of Andrographolide, a Promising Anticancer Agent That Suppresses Tumor Metastasis

Cell-permeable probe for identification and imaging of sialidases

Activity‐Based Protein Profiling: Recent Advances in Probe Development and Applications

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