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Arin K. Greene - One of the best experts on this subject based on the ideXlab platform.
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endothelial MAP2K1 mutations in arteriovenous malformation activate the ras mapk pathway
Biochemical and Biophysical Research Communications, 2020Co-Authors: Patrick Smits, Dennis J. Konczyk, Christopher L. Sudduth, Jeremy A. Goss, Arin K. GreeneAbstract:Arteriovenous malformation (AVM) is a locally destructive congenital vascular anomaly caused by somatic mutations in MAP2K1. The mutation is isolated to endothelial cells (ECs). The purpose of this study was to determine the effects of mutant MAP2K1 on EC signaling and vascular network formation. Pathway effects were studied using both mutant MAP2K1 (K57N) human AVM tissue and human umbilical vein endothelial cells (HUVECs) engineered to overexpress the MAP2K1 (K57N) mutation. Western blot was used to determine cell signaling along the RAS/MAPK pathway. Geltrex tube formation assays were performed to assess EC vascular network formation. Cells were treated with a MAP2K1 inhibitor (Trametinib) to determine its effect on signaling and vascular tube formation. Human mutant MAP2K1-AVM ECs had similar baseline MEK1 and ERK1/2 expression with controls; however, mutant MAP2K1-AVM ECs produced significantly more phosphorylated ERK1/2 than wild-type ECs. Mutant MAP2K1 HUVECs demonstrated significantly more phosphorylated ERK1/2 than control HUVECs. Trametinib reduced the phosphorylation of ERK1/2 in mutant cells and prevented the ability of ECs to form vascular networks. AVM MAP2K1 mutations activate RAS/MAPK signaling in ECs. ERK activation and vascular network formation are reduced with Trametinib. Pharmacotherapy using MAP2K1 inhibitors may prevent the formation or progression of AVMs.
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Endothelial MAP2K1 mutations in arteriovenous malformation activate the RAS/MAPK pathway.
Biochemical and biophysical research communications, 2020Co-Authors: Patrick Smits, Dennis J. Konczyk, Christopher L. Sudduth, Jeremy A. Goss, Arin K. GreeneAbstract:Arteriovenous malformation (AVM) is a locally destructive congenital vascular anomaly caused by somatic mutations in MAP2K1. The mutation is isolated to endothelial cells (ECs). The purpose of this study was to determine the effects of mutant MAP2K1 on EC signaling and vascular network formation. Pathway effects were studied using both mutant MAP2K1 (K57N) human AVM tissue and human umbilical vein endothelial cells (HUVECs) engineered to overexpress the MAP2K1 (K57N) mutation. Western blot was used to determine cell signaling along the RAS/MAPK pathway. Geltrex tube formation assays were performed to assess EC vascular network formation. Cells were treated with a MAP2K1 inhibitor (Trametinib) to determine its effect on signaling and vascular tube formation. Human mutant MAP2K1-AVM ECs had similar baseline MEK1 and ERK1/2 expression with controls; however, mutant MAP2K1-AVM ECs produced significantly more phosphorylated ERK1/2 than wild-type ECs. Mutant MAP2K1 HUVECs demonstrated significantly more phosphorylated ERK1/2 than control HUVECs. Trametinib reduced the phosphorylation of ERK1/2 in mutant cells and prevented the ability of ECs to form vascular networks. AVM MAP2K1 mutations activate RAS/MAPK signaling in ECs. ERK activation and vascular network formation are reduced with Trametinib. Pharmacotherapy using MAP2K1 inhibitors may prevent the formation or progression of AVMs.
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Arteriovenous Malformation MAP2K1 Mutation Causes Local Cartilage Overgrowth by a Cell-Non Autonomous Mechanism.
Scientific reports, 2020Co-Authors: Dennis J. Konczyk, Patrick Smits, Christopher L. Sudduth, Jeremy A. Goss, Alyaa Al-ibraheemi, Arin K. GreeneAbstract:Extracranial arteriovenous malformation (AVM) is most commonly caused by MAP2K1 mutations in the endothelial cell. The purpose of this study was to determine if local tissue overgrowth associated with AVM is caused by direct or indirect effects of the MAP2K1 mutation (i.e., cell-autonomous or cell-non autonomous). Because cartilage does not have blood vessels, we studied ear AVMs to determine if overgrown cartilage contained AVM-causing mutations. Cartilage was separated from its surrounding tissue and isolated by laser capture microdissection. Droplet digital PCR (ddPCR) was used to identify MAP2K1 mutations. MAP2K1 (p.K57N) variants were present in the tissue adjacent to the cartilage [mutant allele frequency (MAF) 6-8%], and were enriched in endothelial cells (MAF 51%) compared to non-endothelial cells (MAF 0%). MAP2K1 mutations were not identified in the overgrown cartilage, and thus local cartilage overgrowth likely results from the effects of adjacent mutant blood vessels (i.e., cell-non autonomous).
Ambroise Wonkam - One of the best experts on this subject based on the ideXlab platform.
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Noonan Syndrome in South Africa: Clinical and Molecular Profiles.
Frontiers in genetics, 2019Co-Authors: Cedrik Tekendo-ngongang, Gloudi Agenbag, Christian Domilongo Bope, Alina Izabela Esterhuizen, Ambroise WonkamAbstract:Noonan Syndrome (NS) is a common autosomal dominant multisystem disorder, caused by mutations in more than 10 genes in the Ras/MAPK signaling pathway. Differential mutation frequencies are observed across populations. Clinical expressions of NS are highly variable and include short stature, distinctive craniofacial dysmorphism, cardiovascular abnormalities, and developmental delay. Little is known about phenotypic specificities and molecular characteristics of NS in Africa. The present study has investigated patients with NS in Cape Town (South Africa). Clinical features were carefully documented in a total of 26 patients. Targeted Next-Generation Sequencing (NGS) was performed on 16 unrelated probands, using a multigene panel comprising 14 genes: PTPN11, SOS1, RIT1, A2ML1, BRAF, CBL, HRAS, KRAS, MAP2K1, MAP2K2, NRAS, RAF1, SHOC2, and SPRED1. The median age at diagnosis was 4.5 years (range: 1 month-51 years). Individuals of mixed-race ancestry were most represented (53.8%), followed by black Africans (30.8%). Our cohort revealed a lower frequency of pulmonary valve stenosis (34.6%) and a less severe developmental milestones phenotype. Molecular analysis found variants predicted to be pathogenic in 5 / 16 cases (31.2%). Among these mutations, two were previously reported: MAP2K1-c.389A>G (p.Tyr130Cys) and PTPN11 - c.1510A>G (p.Met504Val); three are novel: CBL-c.2520T>G (p.Cys840Trp), PTPN11- c.1496C>T (p.Ser499Phe), and MAP2K1- c.200A>C (p.Asp67Ala). Molecular dynamic simulations indicated that novel variants identified impact the stability and flexibility of their corresponding proteins. Genotype-phenotype correlations showed that clinical features of NS were more typical in patients with variants in MAP2K1. This first application of targeted NGS for the molecular diagnosis of NS in South Africans suggests that, while there is no major phenotypic difference compared to other populations, the distribution of genetic variants in NS in South Africans may be different.
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Table_1_Noonan Syndrome in South Africa: Clinical and Molecular Profiles.docx
2019Co-Authors: Cedrik Tekendo-ngongang, Gloudi Agenbag, Christian Domilongo Bope, Alina Izabela Esterhuizen, Ambroise WonkamAbstract:Noonan Syndrome (NS) is a common autosomal dominant multisystem disorder, caused by mutations in more than 10 genes in the Ras/MAPK signaling pathway. Differential mutation frequencies are observed across populations. Clinical expressions of NS are highly variable and include short stature, distinctive craniofacial dysmorphism, cardiovascular abnormalities, and developmental delay. Little is known about phenotypic specificities and molecular characteristics of NS in Africa. The present study has investigated patients with NS in Cape Town (South Africa). Clinical features were carefully documented in a total of 26 patients. Targeted Next-Generation Sequencing (NGS) was performed on 16 unrelated probands, using a multigene panel comprising 14 genes: PTPN11, SOS1, RIT1, A2ML1, BRAF, CBL, HRAS, KRAS, MAP2K1, MAP2K2, NRAS, RAF1, SHOC2, and SPRED1. The median age at diagnosis was 4.5 years (range: 1 month−51 years). Individuals of mixed-race ancestry were most represented (53.8%), followed by black Africans (30.8%). Our cohort revealed a lower frequency of pulmonary valve stenosis (34.6%) and a less severe developmental milestones phenotype. Molecular analysis found variants predicted to be pathogenic in 5 / 16 cases (31.2%). Among these mutations, two were previously reported: MAP2K1-c.389A>G (p.Tyr130Cys) and PTPN11 - c.1510A>G (p.Met504Val); three are novel: CBL-c.2520T>G (p.Cys840Trp), PTPN11- c.1496C>T (p.Ser499Phe), and MAP2K1- c.200A>C (p.Asp67Ala). Molecular dynamic simulations indicated that novel variants identified impact the stability and flexibility of their corresponding proteins. Genotype-phenotype correlations showed that clinical features of NS were more typical in patients with variants in MAP2K1. This first application of targeted NGS for the molecular diagnosis of NS in South Africans suggests that, while there is no major phenotypic difference compared to other populations, the distribution of genetic variants in NS in South Africans may be different.
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Image_1_Noonan Syndrome in South Africa: Clinical and Molecular Profiles.TIF
2019Co-Authors: Cedrik Tekendo-ngongang, Gloudi Agenbag, Christian Domilongo Bope, Alina Izabela Esterhuizen, Ambroise WonkamAbstract:Noonan Syndrome (NS) is a common autosomal dominant multisystem disorder, caused by mutations in more than 10 genes in the Ras/MAPK signaling pathway. Differential mutation frequencies are observed across populations. Clinical expressions of NS are highly variable and include short stature, distinctive craniofacial dysmorphism, cardiovascular abnormalities, and developmental delay. Little is known about phenotypic specificities and molecular characteristics of NS in Africa. The present study has investigated patients with NS in Cape Town (South Africa). Clinical features were carefully documented in a total of 26 patients. Targeted Next-Generation Sequencing (NGS) was performed on 16 unrelated probands, using a multigene panel comprising 14 genes: PTPN11, SOS1, RIT1, A2ML1, BRAF, CBL, HRAS, KRAS, MAP2K1, MAP2K2, NRAS, RAF1, SHOC2, and SPRED1. The median age at diagnosis was 4.5 years (range: 1 month−51 years). Individuals of mixed-race ancestry were most represented (53.8%), followed by black Africans (30.8%). Our cohort revealed a lower frequency of pulmonary valve stenosis (34.6%) and a less severe developmental milestones phenotype. Molecular analysis found variants predicted to be pathogenic in 5 / 16 cases (31.2%). Among these mutations, two were previously reported: MAP2K1-c.389A>G (p.Tyr130Cys) and PTPN11 - c.1510A>G (p.Met504Val); three are novel: CBL-c.2520T>G (p.Cys840Trp), PTPN11- c.1496C>T (p.Ser499Phe), and MAP2K1- c.200A>C (p.Asp67Ala). Molecular dynamic simulations indicated that novel variants identified impact the stability and flexibility of their corresponding proteins. Genotype-phenotype correlations showed that clinical features of NS were more typical in patients with variants in MAP2K1. This first application of targeted NGS for the molecular diagnosis of NS in South Africans suggests that, while there is no major phenotypic difference compared to other populations, the distribution of genetic variants in NS in South Africans may be different.
Yoichi Matsubara - One of the best experts on this subject based on the ideXlab platform.
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A novel heterozygous MAP2K1 mutation in a patient with Noonan syndrome with multiple lentigines.
American journal of medical genetics. Part A, 2014Co-Authors: Eriko Nishi, Seiji Mizuno, Yuka Nanjo, Tetsuya Niihori, Yoshimitsu Fukushima, Yoichi Matsubara, Yoko Aoki, Tomoki KoshoAbstract:Noonan syndrome with multiple lentigines (NSML), formerly referred to as LEOPARD syndrome, is a rare autosomal-dominant condition, characterized by multiple lentigines, electrocardiographic conduction abnormalities, ocular hypertelorism, pulmonary stenosis, abnormal genitalia, growth retardation, and sensorineural deafness. To date, PTPN11, RAF1, and BRAF have been reported to be causal for NSML. We report on a 13-year-old Japanese boy, who was diagnosed with NSML. He was found to have a novel heterozygous missense variant (c.305A > G; p.E102G) in MAP2K1, a gene mostly causal for cardio-facio-cutaneous syndrome (CFCS). He manifested fetal macrosomia, and showed hypotonia and poor sucking in the neonatal period. He had mild developmental delay, and multiple lentigines appearing at approximately age 3 years, as well as flexion deformity of knees bilaterally, subtle facial characteristics including ocular hypertelorism, sensorineural hearing loss, and precocious puberty. He lacked congenital heart defects or hypertrophic cardiomyopathy, frequently observed in patients with NSML, mostly caused by PTPN11 mutations. He also lacked congenital heart defects, characteristic facial features, or intellectual disability, frequently observed in those with CFCS caused by MAP2K1 or MAP2K2 mutations. This may be the first patient clinically diagnosed with NSML, caused by a mutation in MAP2K1.
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A novel heterozygous MAP2K1 mutation in a patient with Noonan syndrome with multiple lentigines.
American Journal of Medical Genetics Part A, 2014Co-Authors: Eriko Nishi, Seiji Mizuno, Yuka Nanjo, Tetsuya Niihori, Yoshimitsu Fukushima, Yoichi Matsubara, Yoko Aoki, Tomoki KoshoAbstract:Noonan syndrome with multiple lentigines (NSML), formerly referred to as LEOPARD syndrome, is a rare autosomal-dominant condition, characterized by multiple lentigines, electrocardiographic conduction abnormalities, ocular hypertelorism, pulmonary stenosis, abnormal genitalia, growth retardation, and sensorineural deafness. To date, PTPN11, RAF1, and BRAF have been reported to be causal for NSML. We report on a 13-year-old Japanese boy, who was diagnosed with NSML. He was found to have a novel heterozygous missense variant (c.305A > G; p.E102G) in MAP2K1, a gene mostly causal for cardio-facio-cutaneous syndrome (CFCS). He manifested fetal macrosomia, and showed hypotonia and poor sucking in the neonatal period. He had mild developmental delay, and multiple lentigines appearing at approximately age 3 years, as well as flexion deformity of knees bilaterally, subtle facial characteristics including ocular hypertelorism, sensorineural hearing loss, and precocious puberty. He lacked congenital heart defects or hypertrophic cardiomyopathy, frequently observed in patients with NSML, mostly caused by PTPN11 mutations. He also lacked congenital heart defects, characteristic facial features, or intellectual disability, frequently observed in those with CFCS caused by MAP2K1 or MAP2K2 mutations. This may be the first patient clinically diagnosed with NSML, caused by a mutation in MAP2K1. © 2014 Wiley Periodicals, Inc.
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The RAS/MAPK syndromes: novel roles of the RAS pathway in human genetic disorders.
Human mutation, 2008Co-Authors: Yoko Aoki, Tetsuya Niihori, Yoko Narumi, Shigeo Kure, Yoichi MatsubaraAbstract:The RAS proteins and their downstream pathways play pivotal roles in cell proliferation, differentiation, survival and cell death, but their physiological roles in human development had remained unknown. Noonan syndrome, Costello syndrome, and cardio-facio-cutaneous (CFC) syndrome are autosomal dominant multiple congenital anomaly syndromes characterized by a distinctive facial appearance, heart defects, musculocutaneous abnormalities, and mental retardation. A variety of mutations in protein tyrosine phosphatase, non-receptor type 11(PTPN11) has been identified in 50% of Noonan patients. Specific mutations in PTPN11 have been identified in LEOPARD (multiple lentigines, electrocardiographic conduction abnormalities, ocular hypertelorism, pulmonary stenosis, abnormal genitalia, retardation of growth, and sensorineural deafness) syndrome. In 2005, we discovered Harvey-RAS (HRAS) germline mutations in patients with Costello syndrome. This discovery provided a clue to identification of germline mutations in Kirsten-RAS (KRAS), BRAF and mitogen-activated protein kinase kinase 1 and 2 (MAP2K1/MAP2K2) in patients with CFC syndrome. These genes encode molecules in the RAS/RAF/MEK/extracellular signal-regulated kinase (ERK) pathway, leading to a new concept that clinically related disorders, i.e., Noonan, Costello, and CFC syndromes are caused by dysregulation of the RAS/mitogen activated protein kinase (MAPK) pathway. In the present review, we summarize mutations in HRAS, KRAS, BRAF, MAP2K1/2, and PTPN11, the phenotypes of patients with these mutations, the functional properties of mutants and animal models. Finally we suggest that disorders with mutations of molecules in the RAS/MAPK cascade (Noonan, LEOPARD, Costello, and CFC syndromes and neurofibromatosis type I) may be comprehensively termed "the RAS/MAPK syndromes." Details on mutations will be updated in the RAS/MAPK Syndromes Homepage (www.medgen.med.tohoku.ac.jp/RasMapk syndromes.html).
S. A. Surzhikov - One of the best experts on this subject based on the ideXlab platform.
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Detection of BRAF, NRAS, KIT, GNAQ, GNA11 and MAP2K1/2 mutations in Russian melanoma patients using LNA PCR clamp and biochip analysis.
Oncotarget, 2017Co-Authors: Marina Emelyanova, Lilit Ghukasyan, I. S. Abramov, Oxana Ryabaya, Evgenia Stepanova, Anna V. Kudryavtseva, Asiya F. Sadritdinova, Cholpon Dzhumakova, Tatiana S. Belysheva, S. A. SurzhikovAbstract:// Marina Emelyanova 1 , Lilit Ghukasyan 1 , Ivan Abramov 1, 2 , Oxana Ryabaya 2 , Evgenia Stepanova 2 , Anna Kudryavtseva 1, 3 , Asiya Sadritdinova 1, 3 , Cholpon Dzhumakova 2 , Tatiana Belysheva 2 , Sergey Surzhikov 1 , Lyudmila Lyubchenko 2 , Alexander Zasedatelev 1, 2 and Tatiana Nasedkina 1, 2 1 Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russian Federation 2 Blokhin Cancer Research Center, Ministry of Health of the Russian Federation, Moscow, Russian Federation 3 P. Hertsen Moscow Oncology Research Institute, Moscow, Russian Federation Correspondence to: Marina Emelyanova, email: emel_marina@mail.ru Keywords: biochip, somatic mutations, melanoma, diagnostic tool, targeted therapy Received: December 30, 2016 Accepted: March 30, 2017 Published: April 10, 2017 ABSTRACT Target inhibitors are used for melanoma treatment, and their effectiveness depends on the tumor genotype. We developed a diagnostic biochip for the detection of 39 clinically relevant somatic mutations in the BRAF , NRAS , KIT , GNAQ , GNA11 , MAP2K1 and MAP2K2 genes. We used multiplex locked nucleic acid (LNA) PCR clamp for the preferable amplification of mutated over wild type DNA. The amplified fragments were labeled via the incorporation of fluorescently labeled dUTP during PCR and were hybridized with specific oligonucleotides immobilized on a biochip. This approach could detect 0.5% of mutated DNA in the sample analyzed. The method was validated on 253 clinical samples and six melanoma cell lines. Among 253 melanomas, 129 (51.0%) BRAF , 45 (17.8%) NRAS , 6 (2.4%) KIT , 4 (1.6%) GNAQ , 2 (0.8%) GNA11 , 2 (0.8%) MAP2K1 and no MAP2K2 gene mutations were detected by the biochip assay. The results were compared with Sanger sequencing, next generation sequencing and ARMS/Scorpion real-time PCR. The specimens with discordant results were subjected to LNA PCR clamp followed by sequencing. The results of this analysis were predominantly identical to the results obtained by the biochip assay. Infrequently, we identified rare somatic mutations. In the present study we demonstrate that the biochip-based assay can effectively detect somatic mutations in approximately 70% of melanoma patients, who may require specific targeted therapy.
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detection of braf nras kit gnaq gna11 and MAP2K1 2 mutations in russian melanoma patients using lna pcr clamp and biochip analysis
Oncotarget, 2017Co-Authors: Marina Emelyanova, Lilit Ghukasyan, I. S. Abramov, Oxana Ryabaya, Evgenia Stepanova, Anna V. Kudryavtseva, Asiya F. Sadritdinova, Cholpon Dzhumakova, Tatiana S. Belysheva, S. A. SurzhikovAbstract:// Marina Emelyanova 1 , Lilit Ghukasyan 1 , Ivan Abramov 1, 2 , Oxana Ryabaya 2 , Evgenia Stepanova 2 , Anna Kudryavtseva 1, 3 , Asiya Sadritdinova 1, 3 , Cholpon Dzhumakova 2 , Tatiana Belysheva 2 , Sergey Surzhikov 1 , Lyudmila Lyubchenko 2 , Alexander Zasedatelev 1, 2 and Tatiana Nasedkina 1, 2 1 Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russian Federation 2 Blokhin Cancer Research Center, Ministry of Health of the Russian Federation, Moscow, Russian Federation 3 P. Hertsen Moscow Oncology Research Institute, Moscow, Russian Federation Correspondence to: Marina Emelyanova, email: emel_marina@mail.ru Keywords: biochip, somatic mutations, melanoma, diagnostic tool, targeted therapy Received: December 30, 2016 Accepted: March 30, 2017 Published: April 10, 2017 ABSTRACT Target inhibitors are used for melanoma treatment, and their effectiveness depends on the tumor genotype. We developed a diagnostic biochip for the detection of 39 clinically relevant somatic mutations in the BRAF , NRAS , KIT , GNAQ , GNA11 , MAP2K1 and MAP2K2 genes. We used multiplex locked nucleic acid (LNA) PCR clamp for the preferable amplification of mutated over wild type DNA. The amplified fragments were labeled via the incorporation of fluorescently labeled dUTP during PCR and were hybridized with specific oligonucleotides immobilized on a biochip. This approach could detect 0.5% of mutated DNA in the sample analyzed. The method was validated on 253 clinical samples and six melanoma cell lines. Among 253 melanomas, 129 (51.0%) BRAF , 45 (17.8%) NRAS , 6 (2.4%) KIT , 4 (1.6%) GNAQ , 2 (0.8%) GNA11 , 2 (0.8%) MAP2K1 and no MAP2K2 gene mutations were detected by the biochip assay. The results were compared with Sanger sequencing, next generation sequencing and ARMS/Scorpion real-time PCR. The specimens with discordant results were subjected to LNA PCR clamp followed by sequencing. The results of this analysis were predominantly identical to the results obtained by the biochip assay. Infrequently, we identified rare somatic mutations. In the present study we demonstrate that the biochip-based assay can effectively detect somatic mutations in approximately 70% of melanoma patients, who may require specific targeted therapy.
Dong Chen - One of the best experts on this subject based on the ideXlab platform.
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LncRNA SNHG12 contributes to multidrug resistance through activating the MAPK/Slug pathway by sponging miR-181a in non-small cell lung cancer
Oncotarget, 2017Co-Authors: Pei Wang, Dong ChenAbstract:Small nucleolar RNA host gene 12 (SNHG12), as one of the long non-coding RNAs (lncRNAs), plays an oncogenic role in various cancers, however, its role in the chemoresistance of non-small cell lung cancer (NSCLC) is unclear. In this study, we investigated the effect of SNHG12 on multidrug resistance (MDR) in NSCLC. The results showed that SNHG12 was high-expressed and miR-181a was low-expressed in NSCLC tumor tissues and cell lines. Knockdown of SNHG12 reversed the resistance to cisplatin, paclitaxel and gefitinib in A549/DDP, A549/PTX and PC9/AB2 cells through inducing cell apoptosis. Moreover, SNHG12 silencing suppressed MAPK1 and MAP2K1 expression by upregulating miR-181a, leading to inhibition of the MAPK/Slug pathway through decreasing phosphorylated MAPK1 (p-MAPK1), phosphorylated MAP2K1 (p-MAP2K1) and Slug levels. Furthermore, downregulation of SNHG12 enhanced the sensitivity of NSCLC cells to cisplatin in nude mice. Overall, our study is the first to identify a SNHG12-miR-181a-MAPK/Slug axis to elucidate in part how SNHG12 exert functions in NSCLC MDR, providing a novel therapeutic target to overcome MDR in NSCLC.