The tyrosine phosphatase Shp2 (PTPN11) directs Neuregulin-1/ErbB signaling throughout Schwann cell development.
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ABSTRACT:
SUBMITTER: Grossmann KS
PROVIDER: S-EPMC2742405 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Activating mutations in PTPN11 (encoding SHP2), a protein tyrosine phosphatase (PTP) that plays an overall positive role in growth factor and cytokine signaling, are directly associated with the pathogenesis of Noonan syndrome and childhood leukemias. Identification of SHP2-selective inhibitors could lead to the development of new drugs that ultimately serve as treatments for PTPN11-associated diseases. As the catalytic core of SHP2 shares extremely high homology to those of SHP1 and other PTPs that play negative roles in cell signaling, to identify selective inhibitors of SHP2 using computer-aided drug design, we targeted a protein surface pocket that is adjacent to the catalytic site, is predicted to be important for binding to phosphopeptide substrates, and has structural features unique to SHP2. From computationally selected candidate compounds, #220-324 effectively inhibited SHP2 activity with an IC50 of 14 ?mol/L. Fluorescence titration experiments confirmed its direct binding to SHP2. This active compound was further verified for its ability to inhibit SHP2-mediated cell signaling and cellular function with minimal off-target effects. Furthermore, mouse myeloid progenitors with the activating mutation (E76K) in PTPN11 and patient leukemic cells with the same mutation were more sensitive to this inhibitor than wild-type cells. This study provides evidence that SHP2 is a "druggable" target for the treatment of PTPN11-associated diseases. As the small-molecule SHP2 inhibitor identified has a simple chemical structure, it represents an ideal lead compound for the development of novel anti-SHP2 drugs. Mol Cancer Ther; 12(9); 1738-48. ©2013 AACR.
| S-EPMC3769526 | biostudies-literature
Project description:Activating mutations of PTPN11 (encoding the SHP2 phosphatase) are associated with Noonan syndrome, childhood leukemias, and sporadic solid tumors. Virtual screening combined with experimental assays was performed to identify inhibitors of SHP2 from a database of natural products. This effort led to the identification of cryptotanshinone as an inhibitor of SHP2. Cryptotanshinone inhibited SHP2 with an IC50 of 22.50 ?M. Fluorescence titration experiments confirmed that it directly bound to SHP2. Enzymatic kinetic analyses showed that cryptotanshinone was a mixed-type and irreversible inhibitor. This drug was further verified for its ability to block SHP2-mediated cell signaling and cellular functions. Furthermore, mouse myeloid progenitors and patient leukemic cells with the activating mutation E76K in PTPN11 were found to be sensitive to this inhibitor. Since cryptotanshinone is used to treat cardiovascular diseases in Asian countries, this drug has a potential to be used directly or to be further developed to treat PTPN11-associated malignancies.
| S-EPMC4159095 | biostudies-literature
Project description:Both activating and inactivating mutations in protein tyrosine phosphatase Ptpn11 (encoding Shp2) are associated with tumorigenesis. However, the underlying mechanisms remain unclear. Here, we show that Shp2 plays an important role in mitosis, dysregulation of which results in chromosome instability and cancer predisposition. Depletion of Shp2 compromised the mitotic checkpoint. Shp2-depleted cells exhibited a delay in mitotic entry and an earlier mitotic exit. Moreover, Shp2 deficiency caused defective kinetochore-microtubule attachment, chromosome misalignment, chromosomal congression defects, lagging chromosomes, and chromosome missegregation. Reintroduction of wild-type Shp2, but not a catalytically deficient mutant, restored the checkpoint function and chromosome alignment at metaphase in Shp2-deficient cells, establishing a requirement for the catalytic activity of Shp2 during mitosis. Further analyses revealed that Shp2 was required for the optimal activation of the mitotic kinases PLK1 and Aurora B and thereby the proper kinetochore localization and phosphorylation of BubR1, a core mitotic checkpoint protein that is also critical for chromosome alignment. Together, our findings show a previously unrecognized role for Shp2 in the maintenance of chromosome stability and suggest a new mechanism by which dysregulation of Shp2 signaling contributes to malignancy development.
| S-EPMC3473170 | biostudies-literature
Project description:The ubiquitous non-receptor protein tyrosine phosphatase SHP2 (encoded by PTPN11) plays a key role in RAS/ERK signaling downstream of most, if not all growth factors, cytokines and integrins, although its major substrates remain controversial. Mutations in PTPN11 lead to several distinct human diseases. Germ-line PTPN11 mutations cause about 50% of Noonan Syndrome (NS), which is among the most common autosomal dominant disorders. LEOPARD Syndrome (LS) is an acronym for its major syndromic manifestations: multiple Lentigines, Electrocardiographic abnormalities, Ocular hypertelorism, Pulmonary stenosis, Abnormalities of genitalia, Retardation of growth, and sensorineural Deafness. Frequently, LS patients have hypertrophic cardiomyopathy, and they might also have an increased risk of neuroblastoma (NS) and acute myeloid leukemia (AML). Consistent with the distinct pathogenesis of NS and LS, different types of PTPN11 mutations cause these disorders.Although multiple studies have reported the biochemical and biological consequences of NS- and LS-associated PTPN11 mutations, their structural consequences have not been analyzed fully. Here we report the crystal structures of WT SHP2 and five NS/LS-associated SHP2 mutants. These findings enable direct structural comparisons of the local conformational changes caused by each mutation.Our structural analysis agrees with, and provides additional mechanistic insight into, the previously reported catalytic properties of these mutants. The results of our research provide new information regarding the structure-function relationship of this medically important target, and should serve as a solid foundation for structure-based drug discovery programs.
| S-EPMC4007598 | biostudies-literature
Project description:Analysis of Schwann cell (SC) development has been hampered by the lack of growing axons in many commonly used in vitro assays. As a consequence, the molecular signals and cellular dynamics of SC development along peripheral axons are still only poorly understood. Here we use a superior cervical ganglion (SCG) explant assay, in which axons elongate after treatment with nerve growth factor (NGF). Migration as well as proliferation and apoptosis of endogenous SCG-derived SCs along sympathetic axons were studied in these cultures using pharmacological interference and time-lapse imaging. Inhibition of ErbB receptor tyrosine kinases leads to reduced SC proliferation, increased apoptosis and thereby severely interfered with SC migration to distal axonal sections and colonization of axons. Furthermore we demonstrate that SC colonization of axons is also strongly impaired in a specific null mutant of an ErbB receptor ligand, Neuregulin 1 (NRG1) type III. Taken together, using a novel SC development assay, we demonstrate that NRG1 type III serves as a critical axonal signal for glial ErbB receptors that drives SC development along sympathetic axons.
| S-EPMC3241675 | biostudies-literature
Project description:The Src-homology 2 (SH2) domain-containing phosphatase 2 (SHP2), encoded by the PTPN11 proto-oncogene, is a key mediator of receptor tyrosine kinase (RTK)-driven cell signaling, promoting cell survival and proliferation. In addition, SHP2 is recruited by immune check point receptors to inhibit B and T cell activation. Aberrant SHP2 function has been implicated in the development, progression, and metastasis of many cancers. Indeed, small molecule SHP2 inhibitors have recently entered clinical trials for the treatment of solid tumors with Ras/Raf/ERK pathway activation, including tumors with some oncogenic Ras mutations. However, the current class of SHP2 inhibitors is not effective against the SHP2 oncogenic variants that occur frequently in leukemias, and the development of specific small molecules that target oncogenic SHP2 is the subject of current research. A common problem with most drug discovery campaigns involving cytosolic proteins like SHP2 is that the primary assays that drive chemical discovery are often in vitro assays that do not report the cellular target engagement of candidate compounds. To provide a platform for measuring cellular target engagement, we developed both wild-type and mutant SHP2 cellular thermal shift assays. These assays reliably detect target engagement of SHP2 inhibitors in cells. Here, we provide a comprehensive protocol of this assay, which provides a valuable tool for the assessment and characterization of SHP2 inhibitors.
| S-EPMC8080703 | biostudies-literature
Project description:During peripheral nerve development, each segment of a myelinated axon is matched with a single Schwann cell. Tight regulation of Schwann cell movement, proliferation and differentiation is essential to ensure that these glial cells properly associate with axons. ErbB receptors are required for Schwann cell migration, but the operative ligand and its mechanism of action have remained unknown. We demonstrate that zebrafish Neuregulin 1 (Nrg1) type III, which signals through ErbB receptors, controls Schwann cell migration in addition to its previously known roles in proliferation and myelination. Chimera analyses indicate that ErbB receptors are required in all migrating Schwann cells, and that Nrg1 type III is required in neurons for migration. Surprisingly, expression of the ligand in a few axons is sufficient to induce migration along a chimeric nerve constituted largely of nrg1 type III mutant axons. These studies also reveal a mechanism that allows Schwann cells to fasciculate axons regardless of nrg1 type III expression. Time-lapse imaging of transgenic embryos demonstrated that misexpression of human NRG1 type III results in ectopic Schwann cell migration, allowing them to aberrantly enter the central nervous system. These results demonstrate that Nrg1 type III is an essential signal that controls Schwann cell migration to ensure that these glia are present in the correct numbers and positions in developing nerves.
| S-EPMC3190382 | biostudies-literature
Project description:Invadopodia are actin-enriched membrane protrusions that are important for extracellular matrix degradation and invasive cell motility. Src homolog domain-containing phosphatase 2 (SHP2), a non-receptor protein tyrosine phosphatase, has been shown to play an important role in promoting cancer metastasis, but the underlying mechanism is unclear. In this study, we found that depletion of SHP2 by short-hairpin RNA suppressed invadopodia formation in several cancer cell lines, particularly in the SAS head and neck squamous cell line. In contrast, overexpression of SHP2 promoted invadopodia formation in the CAL27 head and neck squamous cell line, which expresses low levels of endogenous SHP2. The depletion of SHP2 in SAS cells significantly decreased their invasive motility. The suppression of invadopodia formation by SHP2 depletion was restored by the Clostridium botulinum C3 exoenzyme (a Rho GTPase inhibitor) or Y27632 (a specific inhibitor for Rho-associated kinase). Together, our results suggest that SHP2 may promote invadopodia formation through inhibition of Rho signaling in cancer cells.
| S-EPMC4695156 | biostudies-literature
Project description:The nonreceptor protein-tyrosine phosphatase (PTP) SHP2 is encoded by the proto-oncogene PTPN11 and is a ubiquitously expressed key regulator of cell signaling, acting on a number of cellular processes and components, including the Ras/Raf/Erk, PI3K/Akt, and JAK/STAT pathways and immune checkpoint receptors. Aberrant SHP2 activity has been implicated in all phases of tumor initiation, progression, and metastasis. Gain-of-function PTPN11 mutations drive oncogenesis in several leukemias and cause developmental disorders with increased risk of malignancy such as Noonan syndrome. Until recently, small molecule-based targeting of SHP2 was hampered by the failure of orthosteric active-site inhibitors to achieve selectivity and potency within a useful therapeutic window. However, new SHP2 allosteric inhibitors with excellent potency and selectivity have sparked renewed interest in the selective targeting of SHP2 and other PTP family members. Crucially, drug discovery campaigns focusing on SHP2 would greatly benefit from the ability to validate the cellular target engagement of candidate inhibitors. Here, we report a cellular thermal shift assay that reliably detects target engagement of SHP2 inhibitors. Using this assay, based on the DiscoverX InCell Pulse enzyme complementation technology, we characterized the binding of several SHP2 allosteric inhibitors in intact cells. Moreover, we demonstrate the robustness and reliability of a 384-well miniaturized version of the assay for the screening of SHP2 inhibitors targeting either WT SHP2 or its oncogenic E76K variant. Finally, we provide an example of the assay's ability to identify and characterize novel compounds with specific cellular potency for either WT or mutant SHP2.
| S-EPMC7049970 | biostudies-literature
Project description:Src homology containing protein tyrosine phosphatase 2 (SHP2) represents a noteworthy target for various diseases, serving as a well-known oncogenic phosphatase in cancers. As a result of the low cell permeability and poor bioavailability, the traditional inhibitors targeting the protein tyrosine phosphate catalytic sites are generally suffered from unsatisfactory applied efficacy. Recently, a particularly large number of allosteric inhibitors with striking inhibitory potency on SHP2 have been identified. In particular, few clinical trials conducted have made significant progress on solid tumors by using SHP2 allosteric inhibitors. This review summarizes the development and structure-activity relationship studies of the small-molecule SHP2 inhibitors for tumor therapies, with the purpose of assisting the future development of SHP2 inhibitors with improved selectivity, higher oral bioavailability and better physicochemical properties.
| S-EPMC7838030 | biostudies-literature