Acquisition of a Specific and Potent PTP1B Inhibitor from a Novel Combinatorial Library and Screening Procedure

Shen, Kui; Keng, Yen Fang; Li, Wu; Guo, Xiaoyi; Lawrence, David S.; Zhang, Zhong Yin

doi:10.1074/jbc.m106568200

Cited by 194 publications

(160 citation statements)

References 52 publications

(50 reference statements)

Supporting

Mentioning

156

Contrasting

Unclassified

Order By: Relevance

“…We have recently identified the most potent and selective PTP1B inhibitor (Fig. 1, compound I) reported to date, which displays a K i value of 2.4 nM for PTP1B and exhibits several orders of magnitude of selectivity in favor of PTP1B against a panel of PTPs (28). The negatively charged difluorophosphonate functional groups participate in key active site and near active site interactions that are responsible for the high potency of compound I and selectivity toward PTP1B (29,30).…”

Section: Resultsmentioning

confidence: 99%

The Role of Protein-tyrosine Phosphatase 1B in Integrin Signaling

Liang¹,

Lee²,

Liang³

et al. 2005

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Protein-tyrosine phosphatase 1B (PTP1B) is a key negative regulator of insulin and leptin signaling and a novel therapeutic target for the treatment of type 2 diabetes, obesity, and other associated metabolic syndromes. Because PTP1B regulates multiple signal pathways and it can both enhance and antagonize a cellular event, it is important to establish the physiological relevance of PTP1B in these processes. In this study, we utilize potent and selective PTP1B inhibitors to delineate the role of PTP1B in integrin signaling. We show that down-regulation of PTP1B activity with small molecule inhibitors suppresses cell spreading and migration to fibronectin, increases Tyr 527 phosphorylation in Src, and decreases phosphorylation of FAK, p130Cas , and ERK1/2. In addition, PTP1B "substrate-trapping" mutants bind Tyr 527 -phosphorylated Src and protect it from dephosphorylation by endogenous PTP1B. These results establish that PTP1B promotes integrin-mediated responses in fibroblasts by dephosphorylating the inhibitory pTyr 527 and thereby activating the Src kinase. We also show that PTP1B forms a complex with Src and p130Cas , and that the proline-rich motif PPRPPK (residues 309 -314) in PTP1B is essential for the complex formation. We suggest that the specificity of PTP1B for Src pTyr 527 is mediated by protein-protein interactions involving the docking protein p130Cas with both Src and PTP1B in addition to the interactions between the PTP1B active site and the pTyr 527 motif.

show abstract

Section: Resultsmentioning

confidence: 99%

The Role of Protein-tyrosine Phosphatase 1B in Integrin Signaling

Liang¹,

Lee²,

Liang³

et al. 2005

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…[29][30][31]. The most potent and selective of these was isolated by screening a focused library containing a biasing phosphotyrosine (pTyr) to ensure association with the active site (32). However, inhibitors have also been identified by HTS of nonfocused compound libraries (as used in this study).…”

Section: Discussionmentioning

confidence: 99%

High-resolution dose–response screening using droplet-based microfluidics

Miller

Harrak²,

Mangeat³

et al. 2011

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

258

217

View full text Add to dashboard Cite

A critical early step in drug discovery is the screening of a chemical library. Typically, promising compounds are identified in a primary screen and then more fully characterized in a dose-response analysis with 7-10 data points per compound. Here, we describe a robust microfluidic approach that increases the number of data points to approximately 10,000 per compound. The system exploits Taylor-Aris dispersion to create concentration gradients, which are then segmented into picoliter microreactors by droplet-based microfluidics. The large number of data points results in IC 50 values that are highly precise ( 2.40% at 95% confidence) and highly reproducible (CV 2.45%, n 16). In addition, the high resolution of the data reveals complex dose-response relationships unambiguously. We used this system to screen a chemical library of 704 compounds against protein tyrosine phosphatase 1B, a diabetes, obesity, and cancer target. We identified a number of novel inhibitors, the most potent being sodium cefsulodine, which has an IC 50 of 27 0.83 μM.high-throughput screening | HTS | small molecule library I n the early 16th century the Swiss chemist Paracelsus declared "all substances are poisons, there is none which is not a poison; only the right dose makes a substance non-poisonous." This idea that the biological effects of a chemical compound are dependent upon its concentration was quantified by A. V. Hill in 1910 (1). However, despite the fact that compounds can display complex concentration-dependent relationships, varying in potency, efficacy, and steepness of response, usually just a single measurement at a single concentration (approximately 10 μM) is obtained for each compound in the chemical library during a primary drug screen, even when using state-of-the-art robotic microplate-based screening systems. This results in high numbers of false positives and false negatives (2), as well as the inability to identify subtle, complex pharmacology, such as partial agonism or antagonism. Even when dose-response curves are generated during a quantitative primary screen (3) or, more typically, during the follow-up of a single-point screen, the time and cost limitations mean that the curves typically contain only 7-10 data points each. With ≤10 nonduplicated data points, and as many as four adjustable nonlinear parameters (e.g., in the four-parameter Hill function), the results are highly sensitive to the data quality: For example, the presence of a single outlying data point can substantially alter the fit of the data, unless the outliers are identified and removed (4).We have developed a system that uses droplet-based microfluidics to generate high-quality dose-response data during drug screening. Droplet-based microfluidics is itself a new technology for creating and manipulating picoliter-volume aqueous droplets that function as independent microreactors (for a review see ref. 5). As a result of the miniaturization inherent in this approach, our system (Fig. 1) is capable of generating doseresponse curves at materiall...

show abstract

“…17 Other recombinant PTPs were expressed and purified as described previously. [18][19][20] Non-PTP enzymes were purchased from Sigma-Aldrich and stored at −20 °C.…”

Section: Ptps and Non-ptp Proteinsmentioning

confidence: 99%

Aryl Vinyl Sulfonates and Sulfones as Active Site-Directed and Mechanism-Based Probes for Protein Tyrosine Phosphatases

et al. 2008

Self Cite

View full text Add to dashboard Cite

Protein tyrosine phosphatases (PTPs) play key roles in the regulation of normal and pathological processes ranging from cell proliferation, differentiation, metabolism, and survival to many human diseases including cancer and diabetes. Functional studies of PTP can be greatly facilitated by small molecule probes that covalently label the active site of a PTP through an activity-dependent chemical reaction. In this communication, we characterize phenyl vinyl sulfonate (PVSN) and phenyl vinyl sulfone (PVS) as a new class of mechanism-based PTP probes. PVSN and PVS inactivate a broad range of PTPs in a time-and concentration-dependent fashion. The PVSN and PVS-mediated PTP inactivation is active site-directed and irreversible, resulting from a Michael addition of the active site Cys Sγ onto the terminal carbon of the vinyl group. Structural and mechanistic analyses reveal the molecular basis for the preference of PVSN/PVS toward the PTPs, which lies in the ability of PVSN and PVS to engage the conserved structural and catalytic machinery of the PTP active site. In contrast to early α-bromobenzyl phosphonate based probes, PVSN and PVS are resistant to solvolysis and are cell permeable, and thus hold promise for in vivo applications. Collectively, these properties bode well for the development of aryl vinyl sulfonates/sulfones based PTP probes to interrogate PTP activity in complex proteomes.

show abstract

Acquisition of a Specific and Potent PTP1B Inhibitor from a Novel Combinatorial Library and Screening Procedure

Cited by 194 publications

References 52 publications

The Role of Protein-tyrosine Phosphatase 1B in Integrin Signaling

The Role of Protein-tyrosine Phosphatase 1B in Integrin Signaling

High-resolution dose–response screening using droplet-based microfluidics

Aryl Vinyl Sulfonates and Sulfones as Active Site-Directed and Mechanism-Based Probes for Protein Tyrosine Phosphatases

Contact Info

Product

Resources

About