Review
Gene to ScreenPTP1B as a drug target: recent developments in PTP1B inhibitor discovery
Gene to Screen
Introduction
Reversible protein tyrosine phosphorylation catalyzed by the coordinated actions of protein tyrosine kinases (PTKs) and phosphatases (PTPs) is of paramount importance to the regulation of the signalling events that underlie such fundamental processes as growth and proliferation, differentiation and survival or apoptosis, as well as adhesion and motility [1]. Consequently, cellular pathways regulated by tyrosine phosphorylation offer a rich source of drug targets for developing novel therapeutics 2, 3, 4. The potential of such targeted therapeutics has been well demonstrated by the successful treatment of human chronic myelogenous leukaemia and gastrointestinal stromal tumours with the PTK inhibitor STI-571 (Gleevec) 5, 6, which targets Bcr/Abl or c-kit, aberrantly activated in the malignancies.
PTPs are enzymes that catalyze protein tyrosine dephosphorylation. In humans, more than a hundred PTPs exist that can function either as negative or positive modulators in various signal transduction pathways [7]. As expected, several PTPs are found to antagonize PTK-mediated signalling. For example, mutations in the SH2-containing SHP1 tyrosine phosphatase lead to severe immune dysfunction, giving rise to the moth-eaten phenotype in mice [8]. Thus, SHP1 is an important negative regulator of cytokine signalling and its loss results in sustained tyrosine phosphorylation with consequent enhanced proliferation. Loss of SHP1 is frequently observed in myelodysplastic syndrome [9] and lymphomas [10]. A more recent analysis implicates several PTPs as tumour suppressors that are frequently mutated in colon cancer [11].
Interestingly, there is also mounting evidence that PTPs can also potentiate, rather than antagonize, the actions of PTKs. This mode of synergy enhances mitogenic signalling, leading to cell transformation. Thus CD45, through its capacity to dephosphorylate and activate src family PTKs, is essential for initiating downstream signalling processes in stimulated T and B cells [12]. SHP2 and its Drosophila homolog corkscrew are positive mediators of growth factor signalling 13, 14. Several activating (gain of function) mutations in human SHP2 have been identified as the cause of the inherited disorder Noonan syndrome [15] and some forms of leukaemia and solid tumours 16, 17. Most recently, the phosphatase of regenerating liver (PRL) phosphatases have been implicated as potential oncogenes that promote cell growth and tumour invasion [18].
As discussed above, deregulation of PTP activity contributes to the pathogenesis of several human diseases, including cancer, diabetes and immune disorders 19, 20, 21. The importance of the PTPs in diverse pathophysiology has made them the focus of intense interest as a new class of drug targets. Thus, inhibitors of the PTPs are also expected to have therapeutic value with novel modes of action 22, 23. Among various members of the PTP superfamily, PTP1B has emerged as the best-validated drug target [24]. However, it has become apparent that the conserved structural and mechanistic features of the PTP active site present substantial challenges to drug development. Nevertheless, great progress has been made to address the inherent potency, selectivity and bioavailability problems associated with targeting PTPs for therapeutic development. In the following review, we summarize the major findings that establish PTP1B as an outstanding target for the treatment of diabetes and obesity, and highlight recent developments in PTP1B inhibitor discovery.
Section snippets
Validation of PTP1B as a drug target for diabetes and obesity
PTP1B is localized to the cytoplasmic face of the endoplasmic reticulum and is expressed ubiquitously, including in the classical insulin-targeted tissues such as liver, muscle and fat [25]. Mounting evidence from biochemical, genetic and pharmacological studies support a role for PTP1B as a negative regulator in both insulin and leptin signalling (Figure 1). PTP1B can associate with and dephosphorylate activated insulin receptor (IR) or insulin receptor substrates (IRS) 26, 27, 28, 29, 30.
Challenges in developing PTP1B-based small-molecule therapeutics
Selectivity is one of the major issues in the development of PTP1B inhibitors as drugs. Because all PTPs share a high degree of structural conservation in the active site, the pTyr (phosphotyrosine)-binding pocket, designing inhibitors with both high affinity and selectivity for PTP1B poses a challenge. Fortunately, PTP substrate specificity studies have shown that pTyr alone is not sufficient for high-affinity binding, and residues flanking the pTyr are important for PTP substrate recognition
The library approach
A focused library approach was used to identify highly potent and selective PTP1B inhibitors that are capable of bridging and simultaneously associating with both the active site and an adjacent peripheral site [47]. The library contains (i) a biasing pTyr to ensure association with the active site and (ii) a structurally diverse set of 23 linkers that tether the pTyr moiety to (iii) a structurally diverse set of eight aryl acids, which were designed to associate with the peripheral subsite,
Improving bioavailability of PTP1B Inhibitors
As mentioned earlier, bioavailability represents another major challenge in transforming PTP1B inhibitors into therapeutics because most of the current active site-directed inhibitors contain substantial negative charges. Various approaches have been used to improve cell permeability of PTP1B inhibitors, and some of the commonly used strategies will now be discussed.
Breakaway tethering
There is continued interest in developing novel pTyr mimetics with more acceptable pharmacological properties. A ‘breakaway-tethering’ method was applied to search for novel pTyr mimetics [53]. In this approach, a library of small molecules containing a disulfide-bond tail was screened against PTP1B with a free thiol group engineered at position 47, near the active site. Under partially reducing conditions, if a compound in the library has an affinity for a site near the free thiol group, a
Conclusion
Mounting evidence from biochemical, genetic and small molecule studies have established PTP1B as an outstanding drug target for the treatment of diabetes and obesity. Recent studies have revealed that it is highly feasible to achieve potency and selectivity in PTP1B inhibitor development. In addition, several strategies are being explored to improve the bioavailability of PTP1B inhibitors. It is probable that potent and selective PTP1B inhibitors with optimal pharmacological properties will
Acknowledgements
This work was supported in part by NIH grants DK68447 and CA69202.
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