-
Acta Pharmaceutica Sinica. B Mar 2021The protein tyrosine phosphatase Src homology phosphotyrosyl phosphatase 2 (SHP2) is implicated in various cancers, and targeting SHP2 has become a promising therapeutic...
The protein tyrosine phosphatase Src homology phosphotyrosyl phosphatase 2 (SHP2) is implicated in various cancers, and targeting SHP2 has become a promising therapeutic approach. We herein described a robust cross-validation high-throughput screening protocol that combined the fluorescence-based enzyme assay and the conformation-dependent thermal shift assay for the discovery of SHP2 inhibitors. The established method can effectively exclude the false positive SHP2 inhibitors with fluorescence interference and was also successfully employed to identify new protein tyrosine phosphatase domain of SHP2 (SHP2-PTP) and allosteric inhibitors. Of note, this protocol showed potential for identifying SHP2 inhibitors against cancer-associated SHP2 mutation SHP2-E76A. After initial screening of our in-house compound library (∼2300 compounds), we identified 4 new SHP2-PTP inhibitors (0.17% hit rate) and 28 novel allosteric SHP2 inhibitors (1.22% hit rate), of which SYK-85 and WS-635 effectively inhibited SHP2-PTP (SYK-85: IC = 0.32 μmol/L; WS-635: IC = 4.13 μmol/L) and thus represent novel scaffolds for designing new SHP2-PTP inhibitors. TK-147, an allosteric inhibitor, inhibited SHP2 potently (IC = 0.25 μmol/L). In structure, TK-147 could be regarded as a bioisostere of the well characterized SHP2 inhibitor SHP-099, highlighting the essential structural elements for allosteric inhibition of SHP2. The principle underlying the cross-validation protocol is potentially feasible to identify allosteric inhibitors or those inactivating mutants of other proteins.
PubMed: 33777680
DOI: 10.1016/j.apsb.2020.10.021 -
BMC Medical Genetics Oct 2015Noonan syndrome (NS), a heterogeneous developmental disorder associated with variable clinical expression including short stature, congenital heart defect, unusual... (Review)
Review
BACKGROUND
Noonan syndrome (NS), a heterogeneous developmental disorder associated with variable clinical expression including short stature, congenital heart defect, unusual pectus deformity and typical facial features, is caused by activating mutations in genes involved in the RAS-MAPK signaling pathway.
CASE PRESENTATION
Here, we present a clinical and molecular characterization of a small family with Noonan syndrome. Comprehensive mutation analysis of NF1, PTPN11, SOS1, CBL, BRAF, RAF1, SHOC2, MAP2K2, MAP2K1, SPRED1, NRAS, HRAS and KRAS was performed using targeted next-generation sequencing. The result revealed a recurrent mutation in NRAS, c.179G > A (p.G60E), in the index patient. This mutation was inherited from the index patient's father, who also showed signs of NS.
CONCLUSIONS
We describe clinical features in this family and review the literature for genotype-phenotype correlations for NS patients with mutations in NRAS. Neither of affected individuals in this family presented with juvenile myelomonocytic leukemia (JMML), which together with previously published results suggest that the risk for NS individuals with a germline NRAS mutation developing JMML is not different from the proportion seen in other NS cases. Interestingly, 50% of NS individuals with an NRAS mutation (including our family) present with lentigines and/or Café-au-lait spots. This demonstrates a predisposition to hyperpigmented lesions in NRAS-positive NS individuals. In addition, the affected father in our family presented with a hearing deficit since birth, which together with lentigines are two characteristics of NS with multiple lentigines (previously LEOPARD syndrome), supporting the difficulties in diagnosing individuals with RASopathies correctly. The clinical and genetic heterogeneity observed in RASopathies is a challenge for genetic testing. However, next-generation sequencing technology, which allows screening of a large number of genes simultaneously, will facilitate an early and accurate diagnosis of patients with RASopathies.
Topics: Adult; Cafe-au-Lait Spots; DNA Mutational Analysis; Female; GTP Phosphohydrolases; Genes, ras; High-Throughput Nucleotide Sequencing; Humans; Lentigo; Leukemia, Myelomonocytic, Juvenile; Male; Membrane Proteins; Middle Aged; Mutation; Noonan Syndrome; Pedigree
PubMed: 26467218
DOI: 10.1186/s12881-015-0239-1 -
Viruses Nov 2022To investigate the seroprevalence of severe fever with thrombocytopenia syndrome (SFTS) among wild and companion animals on Tsushima Island, Japan, SFTS virus...
Increased Risk of Infection with Severe Fever with Thrombocytopenia Virus among Animal Populations on Tsushima Island, Japan, Including an Endangered Species, Tsushima Leopard Cats.
To investigate the seroprevalence of severe fever with thrombocytopenia syndrome (SFTS) among wild and companion animals on Tsushima Island, Japan, SFTS virus (SFTSV)-specific ELISA and virus-neutralizing tests were conducted on 50 wild boars, 71 Sika deer, 84 dogs, 323 domestic cats, and 6 Tsushima leopard cats. In total, 1 wild boar (1.8%), 2 dogs (2.4%), 7 domestic cats (2.2%), and 1 Tsushima leopard cat (16.7%) were positive for anti-SFTSV antibodies. Among the 11 positive animals, 10 were collected after 2019, and all were found on the southern part of the island. SFTSV, thus far, seems to be circulating within a limited area of Tsushima Island. To protect humans and animals, including endangered Tsushima leopard cats, from SFTSV infection, countermeasures are needed to prevent the spread of SFTSV on Tsushima Island.
Topics: Humans; Animals; Cats; Dogs; Endangered Species; Japan; Seroepidemiologic Studies; Deer; Phlebovirus; Severe Fever with Thrombocytopenia Syndrome; Panthera; Thrombocytopenia; Bunyaviridae Infections
PubMed: 36560635
DOI: 10.3390/v14122631 -
JACC. Case Reports Aug 2022We describe the case of a 5-year-old girl with genetically confirmed LEOPARD syndrome (LS) who presented with multiple lentigines, ocular hypertelorism, retardation of...
We describe the case of a 5-year-old girl with genetically confirmed LEOPARD syndrome (LS) who presented with multiple lentigines, ocular hypertelorism, retardation of growth, myocardial hypertrophy, and diffuse coronary artery dilatation. This case highlights the importance of multimodality imaging for the assessment of LS-associated cardiovascular alterations and follow-up. ().
PubMed: 36062059
DOI: 10.1016/j.jaccas.2022.07.002 -
Circulation. Heart Failure Oct 2017Alterations in autophagy have been reported in hypertrophic cardiomyopathy (HCM) caused by Danon disease, Vici syndrome, or LEOPARD syndrome, but not in HCM caused by...
BACKGROUND
Alterations in autophagy have been reported in hypertrophic cardiomyopathy (HCM) caused by Danon disease, Vici syndrome, or LEOPARD syndrome, but not in HCM caused by mutations in genes encoding sarcomeric proteins, which account for most of HCM cases. , encoding cMyBP-C (cardiac myosin-binding protein C), is the most frequently mutated HCM gene.
METHODS AND RESULTS
We evaluated autophagy in patients with HCM carrying mutations and in a -targeted knockin HCM mouse model, as well as the effect of autophagy modulators on the development of cardiomyopathy in knockin mice. Microtubule-associated protein 1 light chain 3 (LC3)-II protein levels were higher in HCM septal myectomies than in nonfailing control hearts and in 60-week-old knockin than in wild-type mouse hearts. In contrast to wild-type, autophagic flux was blunted and associated with accumulation of residual bodies and glycogen in hearts of 60-week-old knockin mice. We found that Akt-mTORC1 (mammalian target of rapamycin complex 1) signaling was increased, and treatment with 2.24 mg/kg·d rapamycin or 40% caloric restriction for 9 weeks partially rescued cardiomyopathy or heart failure and restored autophagic flux in knockin mice.
CONCLUSIONS
Altogether, we found that (1) autophagy is altered in patients with HCM carrying mutations, (2) autophagy is impaired in -targeted knockin mice, and (3) activation of autophagy ameliorated the cardiac disease phenotype in this mouse model. We propose that activation of autophagy might be an attractive option alone or in combination with another therapy to rescue HCM caused by mutations.
Topics: Animals; Autophagy; Cardiomyopathies; Cardiomyopathy, Hypertrophic; Carrier Proteins; Disease Models, Animal; Gene Knock-In Techniques; Genotype; Heart Failure; Humans; Mice, Transgenic; Mutation; Myocardium; Phenotype
PubMed: 29021349
DOI: 10.1161/CIRCHEARTFAILURE.117.004140 -
Genes Nov 2022Tissue fragility, skin hyperextensibility and joint hypermobility are defining characteristics of Ehlers-Danlos syndrome (EDS). Human EDS is subclassified into fourteen...
Tissue fragility, skin hyperextensibility and joint hypermobility are defining characteristics of Ehlers-Danlos syndrome (EDS). Human EDS is subclassified into fourteen types including dermatosparactic EDS, characterized by extreme skin fragility and caused by biallelic mutations. We report two novel, variants in DNA from EDS-affected dogs. Separate whole-genome sequences from a Pit Bull Terrier and an Alapaha Blue Blood Bulldog each contained a rare, homozygous variant (11:2280117delC, CanFam3.1), predicted to produce a frameshift in the transcript from the first coding exon (c.10delC) and a severely truncated protein product, p.(Pro4ArgfsTer175). The clinical features of these dogs and 4 others with the same homozygous deletion included multifocal wounds, atrophic scars, joint hypermobility, narrowed palpebral fissures, skin hyperextensibility, and joint-associated swellings. Due to severe skin fragility, the owners of all 6 dogs elected euthanasia before the dogs reached 13 weeks of age. Cross sections of collagen fibrils in post-mortem dermal tissues from 2 of these dogs showed hieroglyphic-like figures similar to those from cases of severe dermatosparaxis in other species. The whole-genome sequence from an adult Catahoula Leopard Dog contained a homozygous missense mutation, [11:2491238G>A; p.(Arg966His)]. This dog exhibited multifocal wounds, atrophic scars, and joint hypermobility, but has survived for at least 9 years. This report expands the spectrum of clinical features of the canine dermatosparactic subtype of EDS and illustrates the potential utility of subclassifying canine EDS by the identity of gene harboring the causal variant.
Topics: Animals; Dogs; ADAMTS Proteins; Atrophy; Cicatrix; Ehlers-Danlos Syndrome; Homozygote; Joint Instability; Phenotype; Sequence Deletion
PubMed: 36421833
DOI: 10.3390/genes13112158 -
Actas Dermo-sifiliograficas Apr 2024
Topics: Humans; LEOPARD Syndrome; Noonan Syndrome; Pigmentation Disorders; Mutation; Protein Tyrosine Phosphatase, Non-Receptor Type 11
PubMed: 36273552
DOI: 10.1016/j.ad.2022.05.028 -
Scientific Reports Aug 2016The genetics of both syndromic (SHL) and non-syndromic hearing loss (NSHL) is characterized by a high degree of genetic heterogeneity. We analyzed whole exome sequencing...
The genetics of both syndromic (SHL) and non-syndromic hearing loss (NSHL) is characterized by a high degree of genetic heterogeneity. We analyzed whole exome sequencing data of 102 unrelated probands with apparently NSHL without a causative variant in known NSHL genes. We detected five causative variants in different SHL genes (SOX10, MITF, PTPN11, CHD7, and KMT2D) in five (4.9%) probands. Clinical re-evaluation of these probands shows that some of them have subtle syndromic findings, while none of them meets clinical criteria for the diagnosis of the associated syndrome (Waardenburg (SOX10 and MITF), Kallmann (CHD7 and SOX10), Noonan/LEOPARD (PTPN11), CHARGE (CHD7), or Kabuki (KMT2D). This study demonstrates that individuals who are evaluated for NSHL can have pathogenic variants in SHL genes that are not usually considered for etiologic studies.
Topics: Adolescent; Child; Child, Preschool; Cohort Studies; Connexins; DNA Helicases; DNA-Binding Proteins; Deafness; Exome; Female; Genetic Heterogeneity; Genetic Predisposition to Disease; Genetic Variation; Humans; Male; Microphthalmia-Associated Transcription Factor; Mutation; Neoplasm Proteins; Pedigree; Protein Tyrosine Phosphatase, Non-Receptor Type 11; SOXE Transcription Factors; Syndrome
PubMed: 27562378
DOI: 10.1038/srep31622 -
Biochemistry Jul 2014The Src homology 2 (SH2) domain-containing protein tyrosine phosphatase 2 (SHP2) is a critical signal transducer downstream of growth factors that promotes the...
The Src homology 2 (SH2) domain-containing protein tyrosine phosphatase 2 (SHP2) is a critical signal transducer downstream of growth factors that promotes the activation of the RAS-ERK1/2 cascade. In its basal state, SHP2 exists in an autoinhibited closed conformation because of an intramolecular interaction between its N-SH2 and protein tyrosine phosphatase (PTP) domains. Binding to pTyr ligands present on growth factor receptors and adaptor proteins with its N-SH2 domain localizes SHP2 to its substrates and frees the active site from allosteric inhibition. Germline mutations in SHP2 are known to cause both Noonan syndrome (NS) and LEOPARD syndrome (LS), two clinically similar autosomal dominant developmental disorders. NS-associated SHP2 mutants display elevated phosphatase activity, while LS-associated SHP2 mutants exhibit reduced catalytic activity. A conundrum in how clinically similar diseases result from mutations to SHP2 that have opposite effects on this enzyme's catalytic functionality exists. Here we report a comprehensive investigation of the kinetic, structural, dynamic, and biochemical signaling properties of the wild type as well as all reported LS-associated SHP2 mutants. The results reveal that LS-causing mutations not only affect SHP2 phosphatase activity but also induce a weakening of the intramolecular interaction between the N-SH2 and PTP domains, leading to mutants that are more readily activated by competing pTyr ligands. Our data also indicate that the residual phosphatase activity associated with the LS SHP2 mutant is required for enhanced ERK1/2 activation. Consequently, catalytically impaired SHP2 mutants could display gain-of-function properties because of their ability to localize to the vicinity of substrates for longer periods of time, thereby affording the opportunity for prolonged substrate turnover and sustained RAS-ERK1/2 activation.
Topics: Enzyme Activation; HEK293 Cells; Humans; Kinetics; LEOPARD Syndrome; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Models, Molecular; Mutation; Protein Conformation; Protein Tyrosine Phosphatase, Non-Receptor Type 11; src Homology Domains
PubMed: 24935154
DOI: 10.1021/bi5002695 -
Cardiovascular Drugs and Therapy Aug 2022Noonan syndrome with multiple lentigines (NSML) is an autosomal dominant disorder presenting with hypertrophic cardiomyopathy (HCM). Up to 85% of NSML cases are caused...
PURPOSE
Noonan syndrome with multiple lentigines (NSML) is an autosomal dominant disorder presenting with hypertrophic cardiomyopathy (HCM). Up to 85% of NSML cases are caused by mutations in the PTPN11 gene that encodes for the Src homology 2 (SH2) domain-containing protein tyrosine phosphatase 2 (SHP2). We previously showed that low-dose dasatinib protects from the development of cardiac fibrosis in a mouse model of NSML harboring a Ptpn11 mutation. This study is performed to determine the pharmacokinetic (PK) and pharmacodynamic (PD) properties of a low-dose of dasatinib in NSML mice and to determine its effectiveness in ameliorating the development of HCM.
METHODS
Dasatinib was administered intraperitoneally into NSML mice with doses ranging from 0.05 to 0.5 mg/kg. PK parameters of dasatinib in NSML mice were determined. PD parameters were obtained for biochemical analyses from heart tissue. Dasatinib-treated NSML mice (0.1 mg/kg) were subjected to echocardiography and assessment of markers of HCM by qRT-PCR. Transcriptome analysis was performed from the heart tissue of low-dose dasatinib-treated mice.
RESULTS
Low-dose dasatinib exhibited PK properties that were linear across doses in NSML mice. Dasatinib treatment of between 0.05 and 0.5 mg/kg in NSML mice yielded an exposure-dependent inhibition of c-Src and PZR tyrosyl phosphorylation and inhibited AKT phosphorylation. We found that doses as low as 0.1 mg/kg of dasatinib prevented HCM in NSML mice. Transcriptome analysis identified differentially expressed HCM-associated genes in the heart of NSML mice that were reverted to wild type levels by low-dose dasatinib administration.
CONCLUSION
These data demonstrate that low-dose dasatinib exhibits desirable therapeutic PK properties that is sufficient for effective target engagement to ameliorate HCM progression in NSML mice. These data demonstrate that low-dose dasatinib treatment may be an effective therapy against HCM in NSML patients.
Topics: Animals; Cardiomyopathy, Hypertrophic; Dasatinib; Disease Models, Animal; LEOPARD Syndrome; Mice; Mutation
PubMed: 33689087
DOI: 10.1007/s10557-021-07169-z