Mosaic RAS/MAPK variants cause sporadic vascular malformations which respond to targeted therapy

Al-Olabi, Lara; Polubothu, Satyamaanasa; Dowsett, Katherine; Andrews, Katrina; Stadnik, Paulina; Joseph, Agnel Praveen; Knox, Rachel; Pittman, Alan; Clark, Graeme M.; Baird, William M.; Bulstrode, Neil; Glover, Mary; Gordon, Kristiana; Hargrave, Darren; Huson, Susan M.; Jacques, Thomas S.; James, Greg; Kondolf, Hannah; Kangesu, Loshan; Keppler‐Noreuil, Kim M.; Khan, Amjad Ali; Lindhurst, Marjorie J.; Lipson, Mark; Mansour, Sahar; O’Hara, Justine; Mahon, Caroline; Mosica, Anda; Moss, Celia; Murthy, Aditi; Ong, Juling; Parker, Victoria; Rivière, Jean‐Baptiste; Sapp, Julie C.; Sebire, Neil J.; Shah, Rahul; Sivakumar, Branavan; Thomas, Anna; Virasami, Alex; Waelchli, Regula; Zeng, Zhiqiang; Biesecker, Leslie G.; Barnacle, Alex; Topf, Maya; Semple, Robert K.; Patton, E. Elizabeth; Kinsler, Veronica A.

doi:10.1172/jci124649

Cited by 81 publications

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“…These, and one inactivating mutation in RASA1, suggest a more significant role (about 8% of all the included cases) of the RAS-RAF-MEK-ERK pathway in vascular malformations than may be expected from the literature. 15,16 We did not detect GNA14 mutations. This is in accordance with the literature reporting that such mutations are restricted to tufted hemangioma and kaposiform hemangioendothelioma, which were not included in our cohorts.…”

Section: Technical Challenges For Molecular Diagnosticsmentioning

confidence: 60%

“…Furthermore, it can provide clues for an underlying hereditary condition due to a germline mutation. In addition, increased knowledge of pathogenic mutations present in these lesions can stratify patients for targeted therapeutic options . A comprehensive and sensitive analysis is required because of the spectrum of genes involved and the mosaicism with low levels of mutated cells and also combined mutations.…”

Section: Resultsmentioning

confidence: 99%

“…(activating mutations in GNAQ) 13 and Maffucci syndrome (transforming mutations in IDH1 and IDH2). 14 Other somatically mutated genes comprise TEK/TIE2 (activating mutations, mainly described in venous malformations), 8 KRAS, NRAS, and HRAS (activating mutations, mainly described in arteriovenous malformations), 15,16 GLMN (disrupting mutations in glomuvenous malformations) 17 and GNA11 and GNA14 (activating mutations in vascular lesions described as vascular tumors including congenital tufted angiomas and kaposiform hemangioendotheliomas). 18 Germline RASA1-mutations have been described in patients with Parkes Weber Syndrome and Capillary Malformation-ArterioVenous Malformations (CM-AVM).…”

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confidence: 99%

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Comprehensive molecular and clinicopathological analysis of vascular malformations: A study of 319 cases

Broek

Eijkelenboom

Vleuten

et al. 2019

Genes Chromosomes & Cancer

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Vascular malformations are part of overgrowth syndromes characterized by somatic mosaic mutations or rarely by germline mutations. Due to their similarities and diversity, clinicopathological classification can be challenging. A comprehensive targeted Next Generation Sequencing screen using Unique Molecular Identifiers with a technical sensitivity of 1% mutant alleles was performed for frequently mutated positions in ≥21 genes on 319 formalin‐fixed paraffin‐embedded samples. In 132 out of 319 cases pathogenic mosaic mutations were detected affecting genes previously linked to vascular malformations e.g. PIK3CA (n=80), TEK ( TIE2 ) (n=11), AKT1 (n=1), GNAQ (n=7), GNA11 (n=4), IDH1 (n=3), KRAS (n=9), and NRAS (n=1). Six cases harbored a combination of mutations in PIK3CA and in GNA11 (n=2), GNAQ (n=2), or IDH1 (n=2). Aberrations in PTEN and RASA1 with a variant allele frequency approaching 50% suggestive of germline origin were identified in six out of 102 cases tested; four contained a potential second hit at a lower allele frequency. Ninety‐one of the total 142 pathogenic mutations were present at a variant allele frequency <10% illustrating the importance of sensitive molecular analysis. Clinicopathological characteristics showed a broad spectrum and overlap when correlated with molecular data. Sensitive screening of recurrently mutated genes in vascular malformations may help to confirm the diagnosis and reveals potential therapeutic options with a significant contribution of PIK3CA/mTOR and RAS‐MAPK pathway mutations. The co‐existence of two activating pathogenic mutations in parallel pathways illustrates potential treatment challenges and underlines the importance of multigene testing. Detected germline mutations have major clinical impact.

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Section: Technical Challenges For Molecular Diagnosticsmentioning

confidence: 60%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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Comprehensive molecular and clinicopathological analysis of vascular malformations: A study of 319 cases

Broek

Eijkelenboom

Vleuten

et al. 2019

Genes Chromosomes & Cancer

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“…Multiple vascular malformations were documented specifically in lztr1 del/del homozygous mutants (8 out of 8 [100%] lztr1 del/del fish analyzed), while none of the wild‐type or lztr1 del/+ controls exhibited the phenotype (Table ). These lesions observed in zebrafish resembled human vascular malformations caused by somatic activating variants in genes of the RAS/MAPK pathway: KRAS , NRAS , BRAF , and MAP2K1 (Al‐Olabi et al, ). Although patients with recessive Noonan syndrome have not yet been reported to manifest such vascular abnormalities, careful follow‐up to monitor for its emergence is of clinical significance since these vascular malformations may lead to life‐threatening bleeds, disfigurement, and/or pain.…”

Section: Resultsmentioning

confidence: 92%

Noonan syndrome‐associated biallelic LZTR1 mutations cause cardiac hypertrophy and vascular malformations in zebrafish

Nakagama

Takeda

Ogawa

et al. 2019

Molec Gen & Gen Med

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Background Variants in the LZTR1 (leucine‐zipper‐like transcription regulator 1) gene (OMIM #600574) have been reported in recessive Noonan syndrome patients. In vivo evidence from animal models to support its causative role is lacking. Methods By CRISPR‐Cas9 genome editing, we generated lztr1‐mutated zebrafish (Danio rerio). Analyses of histopathology and downstream signaling were performed to investigate the pathogenesis of cardiac and extracardiac abnormalities in Noonan syndrome. Results A frameshift deletion allele was created in the zebrafish lztr1. Crosses of heterozygotes obtained homozygous lztr1 null mutants that modeled LZTR1 loss‐of‐function. Histological analyses of the model revealed ventricular hypertrophy, the deleterious signature of Noonan syndrome‐associated cardiomyopathy. Further, assessment for extracardiac abnormalities documented multiple vascular malformations, resembling human vascular pathology caused by RAS/MAPK activation. Due to spatiotemporal regulation of LZTR1, its downstream function was not fully elucidated from western blots of adult tissue. Conclusion Our novel zebrafish model phenocopied human recessive Noonan syndrome and supported the loss‐of‐function mechanism of disease‐causing LZTR1 variants. The discovery of vascular malformations in mutants calls for the clinical follow‐up of patients to monitor for its emergence. The model will serve as a novel platform for investigating the pathophysiology linking RAS/MAPK signaling to cardiac and vascular pathology.

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“…Interestingly, recent studies showed that sporadic brain AVMs and extra-neural AVMs harbor somatic mutations of genes in Ras-mitogen-activated protein kinases (MAPK) pathways including KRAS, MAP2K1 and BRAF. These mutations were mostly present in ECs [12][13][14]. However, it is not clear how a fraction of mutant ECs leads to AVM development and if mutation in BM-derived ECs alone can cause AVM formation.…”

Section: Introductionmentioning

confidence: 99%

Alk1 mutant endothelial cells undergo clonal expansion in mouse brain arteriovenous malformations

Shaligram

Zhang

Zhu

et al. 2020

Preprint

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Total word count:Subject codes: cerebrovascular malformations, vascular disease, animal models of human disease Abstract:Rationale: Mutation in human arteriovenous malformation (AVM) causative genes in a fraction of endothelial cells (ECs) causes AVMs in mice. It is unclear how a small number of mutant ECs can lead to AVM formation. Objective:To understand how a fraction of mutant ECs causes AVM, we tested the following hypotheses: (1) activin receptor-like kinase 1 (Alk1 or Acvlr1) mutant brain ECs undergo clonal expansion upon angiogenic stimulation, (2) Alk1 mutant ECs display growth advantage, (3) the burden of Alk1 mutant ECs correlates with AVM severity, and (4) Alk1 mutant bone marrow (BM) derived ECs alone is sufficient to cause AVM. Methods and Results:We used PdgfbiCreER;Alk1 f/f ;confetti +/mice which express an EC-specific tamoxifen (TM)-inducible Cre recombinase, a Cre-regulated confetti transgene, and Alk1 floxed alleles. Brain AVMs were induced by direct brain injection of an adeno-associated viral vector expressing vascular endothelial growth factor (AAV-VEGF) followed with intra-peritoneal injection of TM two weeks later. Colorpredominance of confetti reporter in AVMs compared to control brain ECs suggested that clonal expansion was associated with AVM development. We treatedPdgfbiCreER;Alk1 f/f with different doses of TM to create a mosaic of wild-type (WT) and mutant ECs and found that equal numbers of Alk1 + and Alk1 -ECs were proliferating. Increase of TM dose increased the number of Alk1 -ECs, the abnormal vessels in brain AVMs, the number of arteriovenous shunts in the intestines, and mouse mortality. To test if mutation of Alk1 in BM-derived ECs can cause brain AVM, we transplanted WT mice with BM of PdgfbiCreER;Alk1 f/f mice. After AAV-VEGF and TM treatment, these mice developed AVMs in their brains and arteriovenous shunts in their intestines. Conclusion:Clonal expansion of Alk1 mutant ECs could partly explain why a fraction of mutant ECs causes AVM. Mutation of AVM causal genes in BM-derived ECs is sufficient to cause AVM formation.

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Mosaic RAS/MAPK variants cause sporadic vascular malformations which respond to targeted therapy

Abstract: In Table 2, the mutation for patient no. 11 in the column labeled "cDNA, aa change" is incorrect. The correct mutation is NM_002524.4:c.182A>G; NRAS p.(Q61R). The JCI regrets the error.

Cited by 81 publications

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Comprehensive molecular and clinicopathological analysis of vascular malformations: A study of 319 cases

Comprehensive molecular and clinicopathological analysis of vascular malformations: A study of 319 cases

Noonan syndrome‐associated biallelic LZTR1 mutations cause cardiac hypertrophy and vascular malformations in zebrafish

Alk1 mutant endothelial cells undergo clonal expansion in mouse brain arteriovenous malformations

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