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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Is RK-682 a promiscuous enzyme inhibitor? Synthesis and in vitro evaluation of protein tyrosine phosphatase inhibition of racemic RK-682 and analogues

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Carneiro, Vania M. T. [1] ; Trivella, Daniela B. B. [2] ; Scorsato, Valeria [1] ; Beraldo, Viviane L. [2] ; Dias, Mariana P. [1] ; Sobreira, Tiago J. P. [2] ; Aparicio, Ricardo [1] ; Pilli, Ronaldo A. [1]
Total Authors: 8
[1] Univ Estadual Campinas, Inst Chem, UNICAMP, BR-13084971 Campinas, SP - Brazil
[2] Brazilian Biosci Natl Lab, Natl Ctr Res Energy & Mat, BR-13083100 Campinas, SP - Brazil
Total Affiliations: 2
Document type: Journal article
Web of Science Citations: 4

RK-682 (1) is a natural product known to selectively inhibit protein tyrosine phosphatases (PTPases) and is used commercially as a positive control for phosphatase inhibition in in vitro assays. Protein phosphatases are involved in several human diseases including diabetes, cancer and inflammation, and are considered important targets for pharmaceutical development. Here we report the synthesis of racemic RK-682 (rac-1) and a focused set of compounds, including racemic analogues of I, dihydropyranones and C-acylated Meldrum's acid derivatives, the later obtained in one synthetic step from commercially available starting material. We further characterized the behavior of some representative compounds in aqueous solution and evaluated their in vitro PTPase binding and inhibition. Our data reveal that rac-1 and some derivatives are able to form large aggregates in solution, in which the aggregation capacity is dependent on the acyl side chain size. However, compound aggregation per se is not able to promote PTPase inhibition. Our data disclose a novel family of PTPase inhibitors (C-acylated Meldrum's acid derivatives) and that rac-1 and derivatives with an exposed latent negatively charged substructure (e.g.: the tetronic acid core of 1) can bind to the PTPase binding site, as well promiscuously to protein surfaces. The combined capacity of compounds to bind to proteins together with their intrinsic capacity to aggregate in solution seems essential to promote enzyme aggregation and thus, its inhibition. We also observed that divalent cations, such as magnesium frequently used in enzyme buffer solutions, can deplete the inhibitory activity of rac-1, thus influencing the enzyme inhibition experiment. Overall, these data help to characterize the mechanism of PTPase inhibition by rac-1 and derivatives, revealing that enzyme inhibition is not solely dependent on compound binding to the PTPase catalytic site as generally accepted in the literature. In addition, our results point to promiscuous mechanisms that influence significantly the in vitro evaluation of enzyme inhibition by rac-1. Therefore, we recommend caution when using natural or synthetic RK-682 (1) as an internal control for evaluating PTPase inhibition and selectivity, since many events can modulate the apparent enzyme inhibition. (C) 2015 Elsevier Masson SAS. All rights reserved. (AU)

FAPESP's process: 11/00457-5 - Synthesis of Dihydropyranones, evaluation of the inhibitory activity of CDC25 and cytotoxicity in tumor cells and structural studies
Grantee:Vânia Maria Teixeira Carneiro
Support Opportunities: Scholarships in Brazil - Post-Doctoral
FAPESP's process: 10/17544-5 - Structural and molecular bases of inhibitor recognition by the CDC-25 and LMW-PTP human protein phosphatases involved in cancer
Grantee:Daniela Barretto Barbosa Trivella
Support Opportunities: Scholarships in Brazil - Post-Doctoral
FAPESP's process: 09/51602-5 - Chemical biology: new natural and synthetic molecular targets against cancer, structural studies, biological evaluation and mode of action
Grantee:Ronaldo Aloise Pilli
Support Opportunities: Research Projects - Thematic Grants
FAPESP's process: 13/07607-8 - OCRC - Obesity and Comorbidities Research Center
Grantee:Licio Augusto Velloso
Support Opportunities: Research Grants - Research, Innovation and Dissemination Centers - RIDC