Development of an Activity‐Based Probe for Steroid Sulfatases

Lu, Chun‐Ping; Ren, Chien‐Tai; Wu, Shih‐Hsiung; Chu, Chih-Yuan; Lo, Lee‐Chiang

doi:10.1002/cbic.200700279

Cited by 32 publications

(18 citation statements)

References 45 publications

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“…An alternative strategy based on the use of the protected sulfonation reagent 8 [25,26] was investigated [27,28]. Following the procedure of Taylor [29], treatment of phenol 7 (0.3 M concentration in the reaction medium) with an excess of 8 in the presence of triethylamine and DMAP afforded the sulfate diester 10 in 94% yield.…”

Section: Synthesis Of Sulfated Metanephrinesmentioning

confidence: 99%

Synthetic calibrators for the analysis of total metanephrines in urine: Revisiting the conditions of hydrolysis

Simonin

Gerber‐Lemaire

Centeno

et al. 2012

Clinica Chimica Acta

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Background: The quantification of total (free + sulfated) metanephrines in urine is recommended to diagnose pheochromocytoma. Urinary metanephrines include metanephrine itself, normetanephrine and methoxytyra-mine, mainly in the form of sulfate conjugates (60-80%). Their determination requires the hydrolysis of the sul-fate ester moiety to allow electrochemical oxidation of the phenolic group. Commercially available urine calibrators and controls contain essentially free, unhydrolysable metanephrines which are not representative of native urines. The lack of appropriate calibrators may lead to uncertainty regarding the completion of the hy-drolysis of sulfated metanephrines, resulting in incorrect quantification. Methods: We used chemically synthesized sulfated metanephrines to establish whether the procedure most fre-quently recommended for commercial kits (pH 1.0 for 30 min over a boiling water bath) ensures their complete hydrolysis. Results: We found that sulfated metanephrines differ in their optimum pH to obtain complete hydrolysis. Highest yields and minimal variance were established for incubation at pH 0.7-0.9 during 20 min. Conclusion: Urinary pH should be carefully controlled to ensure an efficient and reproducible hydrolysis of sul-fated metanephrines. Synthetic sulfated metanephrines represent the optimal material for calibrators and pro-ficiency testing to improve inter-laboratory accuracy.

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Section: Synthesis Of Sulfated Metanephrinesmentioning

confidence: 99%

Synthetic calibrators for the analysis of total metanephrines in urine: Revisiting the conditions of hydrolysis

Simonin

Gerber‐Lemaire

Centeno

et al. 2012

Clinica Chimica Acta

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“…This trapping device has previously been used to probe protein tyrosine phosphatase activity. [23][24][25][26][27] Upon ATXmediated cleavage of the phosphodiester bond in the probe, the released intermediate undergoes rapid 1,6-elimination of the fluoride and generates a reactive quinone methide species that, in turn, traps nearby nucleophiles in the ATX active site; this results in covalent labelling of ATX in an activity-dependent manner. Incorporation of a fluorescent dye allows visualisation of active ATX.…”

Section: Probe Designmentioning

confidence: 99%

Development of an Activity‐Based Probe for Autotaxin

et al. 2010

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Autotaxin (ATX), or ecto-nucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2), is a secreted lysophospholipase D that hydrolyses lysophosphatidylcholine into the lipid mediator lysophosphatidic acid (LPA), a mitogen and chemoattractant for many cell types. ATX has been implicated in tumour progression and inflammation, and might serve as a biomarker. Here we describe the development of a fluorescent activity-based probe that covalently binds to the active site of ATX. The probe consists of a lysophospholipid-based backbone linked to a trapping moiety that becomes reactive after phosphate ester hydrolysis, and a Cy5 fluorescent dye to allow visualisation of active ATX. The probe reacts specifically with the three known isoforms of ATX, it competes with small-molecule inhibitors for binding to ATX and allows ATX activity in plasma to be determined. Our activity-based reporter will be useful for monitoring ATX activity in biological fluids and for inhibitor screening.

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“…This has included the development of probes that target non-enzymatic proteins such as GABA receptors 60 and acetylcholine receptors. 61 In addition, there have been recent reports of probes that target other diverse enzyme classes including sulfatases, 62 deaminases, 35 dimethylaminohydrolases, 63 lipases, 64,65 esterases, 65,66 nitrilases, 67 cytochrome P450s, 68 gylcosidases, 69 gylcanases 70 and beta galactosidase. 71 This ever-expanding list of enzyme families and receptors that can be targeted by ABPs suggests that the field continues to advance.…”

Section: Other Classes Of Enzymesmentioning

confidence: 99%

Applications for Activity-based Probes in Drug Discovery

Edgington

Bogyo

2010

New Frontiers in Chemical Biology

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The path to a new therapeutic drug is long and difficult and involves many stages including validation of a target, design and selection of a lead compound and finally development of the lead into a drug. This chapter will discuss a relatively new technology that makes use of small molecules termed activity based probes (ABPs). These probes bind in the active site of a target enzyme or class of enzymes in an activity dependent fashion. Thus probe labeling serves as an indirect readout of enzyme activity, allowing the dynamic regulation of the target enzyme to be monitored using a number of biochemical and cell biological methods. In addition, labeled targets can be directly isolated by affinity methods, thereby allowing identification of potentially valuable drug targets based solely on their ability to bind a small molecule. Finally, because of the high degree of selectivity of ABPs for a given target protein class, they can be used for studies of drug binding and efficacy in complex cellular mixtures, intact cells and even in whole animals. These attributes of ABPs make them extremely valuable reagents for use at multiple points in the drug discovery process.

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Development of an Activity‐Based Probe for Steroid Sulfatases

Cited by 32 publications

References 45 publications

Synthetic calibrators for the analysis of total metanephrines in urine: Revisiting the conditions of hydrolysis

Synthetic calibrators for the analysis of total metanephrines in urine: Revisiting the conditions of hydrolysis

Development of an Activity‐Based Probe for Autotaxin

Applications for Activity-based Probes in Drug Discovery

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