7-140754187-T-C

Position:

• chr7-140754187-T-C

Variant summary

Our verdict is Pathogenic. Variant got 14 ACMG points: 14P and 0B. PM2 PP2 PP3 PM6_Strong PM1 PS3

This summary comes from the ClinGen Evidence Repository: The c.1741A>G (p.Asn581Asp) variant in BRAF has been reported in the literature as a non-maternity/paternity confirmed de novo occurrence in at least 2 patients with clinical features of a RASopathy (PM6_Strong; PMID 25463315). In vitro functional studies provide some evidence that the p.Asn581Asp variant may impact protein function (PS3; 19376813, 16474404). This variant was absent from large population studies (PM2; ExAC, http://exac.broadinstitute.org). The variant is located in the BRAF gene, which has been defined by the ClinGen RASopathy Expert Panel as a gene with a low rate of benign missense variants and pathogenic missense variants are common (PP2; PMID 29493581). Furthermore, this variant is located in the catalytic loop of BRAF (PM1; 16474404, 29493581). Computational prediction tools and conservation analysis suggest that the p.Asn581Asp variant may impact the protein (PP3). In summary, this variant meets criteria to be classified as pathogenic for RASopathies in an autosomal dominant manner. Rasopathy-specific ACMG/AMP criteria applied (PMID:29493581): PP2, PP3, PM1, PM2, PM6_Strong, PS3. LINK:https://erepo.genome.network/evrepo/ui/classification/CA279976/MONDO:0015280/004

• Frequency:
  • Genomes: not found (cov: 32)
  • Exomes 𝑓: 0.0 (0 hom.)
  • Failed GnomAD Quality Control
• Consequence: BRAF NM_001374258.1 missense, splice_region
• Scores: Splicing: ADA: 0.4446
• Clinical Significance: Pathogenic reviewed by expert panel P:16 O:1
• Conservation: PhyloP100: 7.85

Genes affected

BRAF (HGNC:1097): (B-Raf proto-oncogene, serine/threonine kinase) This gene encodes a protein belonging to the RAF family of serine/threonine protein kinases. This protein plays a role in regulating the MAP kinase/ERK signaling pathway, which affects cell division, differentiation, and secretion. Mutations in this gene, most commonly the V600E mutation, are the most frequently identified cancer-causing mutations in melanoma, and have been identified in various other cancers as well, including non-Hodgkin lymphoma, colorectal cancer, thyroid carcinoma, non-small cell lung carcinoma, hairy cell leukemia and adenocarcinoma of lung. Mutations in this gene are also associated with cardiofaciocutaneous, Noonan, and Costello syndromes, which exhibit overlapping phenotypes. A pseudogene of this gene has been identified on the X chromosome. [provided by RefSeq, Aug 2017]

ACMG classification

Verdict is Pathogenic. Variant got 14 ACMG points.

• PS3: For more information check the summary or visit ClinGen Evidence Repository.
• PM1: For more information check the summary or visit ClinGen Evidence Repository.
• PM2: For more information check the summary or visit ClinGen Evidence Repository.
• PM6: For more information check the summary or visit ClinGen Evidence Repository.
• PP2: For more information check the summary or visit ClinGen Evidence Repository.
• PP3: For more information check the summary or visit ClinGen Evidence Repository.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
BRAF	NM_001374258.1	c.1861A>G	p.Asn621Asp	missense_variant, splice_region_variant	15/20	ENST00000644969.2	NP_001361187.1
BRAF	NM_004333.6	c.1741A>G	p.Asn581Asp	missense_variant, splice_region_variant	14/18	ENST00000646891.2	NP_004324.2

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
BRAF	ENST00000644969.2	c.1861A>G	p.Asn621Asp	missense_variant, splice_region_variant	15/20		NM_001374258.1	ENSP00000496776
BRAF	ENST00000646891.2	c.1741A>G	p.Asn581Asp	missense_variant, splice_region_variant	14/18		NM_004333.6	ENSP00000493543	P4

Frequencies

GnomAD3 genomes Cov.: 32

GnomAD4 exome Data not reliable, filtered out with message: AC0 AF: 0.00 AC: 0 AN: 1461014 Hom.: 0 Cov.: 30 AF XY: 0.00 AC XY: 0 AN XY: 726854

Gnomad4 AFR exome AF: 0.00

Gnomad4 AMR exome AF: 0.00

Gnomad4 ASJ exome AF: 0.00

Gnomad4 EAS exome AF: 0.00

Gnomad4 SAS exome AF: 0.00

Gnomad4 FIN exome AF: 0.00

Gnomad4 NFE exome AF: 0.00

Gnomad4 OTH exome AF: 0.00

GnomAD4 genome Cov.: 32

ClinVar

Significance: Pathogenic

Submissions summary: Pathogenic:16 Other:1

Revision: reviewed by expert panel

Submissions by phenotype

Cardiofaciocutaneous syndrome 1 Pathogenic:5

Pathogenic, criteria provided, single submitter	clinical testing	3billion	Oct 02, 2021	The variant has been previously reported as assumed (i.e. paternity and maternity not confirmed) de novoo in at least two similarly affected unrelated individuals (PMID:25463315, PM6_S). Functional studies provide strong evidence of the variant having a damaging effect on the gene or gene product (PMID: 19376813, 16474404). It is not observed in the gnomAD v2.1.1 dataset (PM2). A different missense change at the same codon (p.Asn621Lys) has been reported as pathogenic (ClinVar ID: VCV000044811.2, PM5). In silico tool predictions suggest damaging effect of the variant on gene or gene product (REVEL: 0.932, 3Cnet: 0.999, PP3). Therefore, this variant is classified as pathogenic according to the recommendation of ACMG/AMP guideline. -
Pathogenic, no assertion criteria provided	literature only	OMIM	Mar 01, 2006	- -
Pathogenic, no assertion criteria provided	research	Division of Human Genetics, National Health Laboratory Service/University of the Witwatersrand	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	Center for Human Genetics, Inc, Center for Human Genetics, Inc	Nov 01, 2016	- -
Pathogenic, criteria provided, single submitter	clinical testing	Victorian Clinical Genetics Services, Murdoch Childrens Research Institute	Jul 17, 2023	Based on the classification scheme VCGS_Germline_v1.3.4, this variant is classified as Pathogenic. Following criteria are met: 0101 - Gain of function is a known mechanism of disease in this gene and is associated with LEOPARD syndrome 3 (MIM#613707), cardiofaciocutaneous syndrome (MIM#115150) and Noonan syndrome 7 (MIM#613706) (PMID: 28783719, PMID: 29540830). (I) 0107 - This gene is associated with autosomal dominant disease. (I) 0200 - Variant is predicted to result in a missense amino acid change from asparagine to aspartic acid. (I) 0251 - This variant is heterozygous. (I) 0301 - Variant is absent from gnomAD (both v2 and v3). (SP) 0502 - Missense variant with conflicting in silico predictions and uninformative conservation. (I) 0602 - Variant is located in a hotspot region or cluster of pathogenic variants (DECIPHER). (SP) 0801 - This variant has strong previous evidence of pathogenicity in unrelated individuals. This variant has been reported many times as pathogenic, and observed in individuals with cardiofaciocutaneous syndrome. One of these reports is by an expert panel (ClinVar). (SP) 1102 - Strong phenotype match for this individual. (SP) 1203 - This variant has been shown to be de novo in the proband (parental status confirmed, by trio analysis). (SP) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign -

not provided Pathogenic:5

Pathogenic, no assertion criteria provided	clinical testing	Genome Diagnostics Laboratory, Amsterdam University Medical Center	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Mar 09, 2023	Not observed at significant frequency in large population cohorts (gnomAD); Published functional studies demonstrate N581D shows impaired ability to induce phosphorylation of ERK and MEK (Niihori et al., 2006); In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; Missense variants in this gene are often considered pathogenic (HGMD); This variant is associated with the following publications: (PMID: 23093928, 16439621, 34573299, 30141192, 33040082, 22876591, 19376813, 24803665, 30050098, 29907801, 33967092, 34643321, 16474404, 25463315) -
Pathogenic, criteria provided, single submitter	clinical testing	Institute of Medical Genetics and Applied Genomics, University Hospital Tübingen	Oct 23, 2020	- -
Pathogenic, no assertion criteria provided	clinical testing	Joint Genome Diagnostic Labs from Nijmegen and Maastricht, Radboudumc and MUMC+	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	Revvity Omics, Revvity	Sep 15, 2022	- -

Cardio-facio-cutaneous syndrome Pathogenic:3 Other:1

Pathogenic, reviewed by expert panel	curation	ClinGen RASopathy Variant Curation Expert Panel	Apr 03, 2017	The c.1741A>G (p.Asn581Asp) variant in BRAF has been reported in the literature as a non-maternity/paternity confirmed de novo occurrence in at least 2 patients with clinical features of a RASopathy (PM6_Strong; PMID 25463315). In vitro functional studies provide some evidence that the p.Asn581Asp variant may impact protein function (PS3; 19376813, 16474404). This variant was absent from large population studies (PM2; ExAC, http://exac.broadinstitute.org). The variant is located in the BRAF gene, which has been defined by the ClinGen RASopathy Expert Panel as a gene with a low rate of benign missense variants and pathogenic missense variants are common (PP2; PMID 29493581). Furthermore, this variant is located in the catalytic loop of BRAF (PM1; 16474404, 29493581). Computational prediction tools and conservation analysis suggest that the p.Asn581Asp variant may impact the protein (PP3). In summary, this variant meets criteria to be classified as pathogenic for RASopathies in an autosomal dominant manner. Rasopathy-specific ACMG/AMP criteria applied (PMID:29493581): PP2, PP3, PM1, PM2, PM6_Strong, PS3. -
not provided, no classification provided	phenotyping only	GenomeConnect - CFC International	-	- -
Pathogenic, criteria provided, single submitter	clinical testing	Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine	Jul 12, 2013	The Asn581Asp variant has been reported in the literature in eight individuals w ith clinical features of Cardio-facio-cutaneous syndrome and was absent from at least 180 control chromosomes (Rodriguez-Viciana 2006, Niihori 2006, Schulz 2008 , Yoon 2007, Hazan 2012). This variant was observed to have occurred de novo in at least three of these individuals (Rodriguez-Viciana 2006, Hazan 2012). The As n581Asp variant has also been identified by our laboratory in one individual wit h Cardio-facio-cutaneous syndrome (LMM unpublished data), and was not identified in this individual's parents. Additionally, the Asparagine (Asn) residue at pos ition 581 is highly conserved across evolutionarily distant species, and the var iant was not identified in large population studies. In summary, this variant me ets our criteria to be classified as pathogenic (http://pcpgm.partners.org/LMM) based upon segregation studies, de novo occurrence, and absence from controls. -
Pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Nov 04, 2016	Variant summary: The BRAF c.1741A>G (p.Asn581Asp) variant involves the alteration of a conserved nucleotide and is located in the catalytic loop of the protein kinase domain (InterPro, Niihori_2006). 3/4 in silico tools predict a damaging outcome for this variant. This variant is absent in approximately 121424 control chromosomes. In literature, this variant is reported as a pathogenic variant found in several patients with cardio-facio-cutaneous syndrome, including reported de novo occurrences (Niihori_2006, Rodriguez-Viciana_2006, Narumi_2007, Nava_2007, Schulz_2008, Pierpont _2010, Hazan_2012, Quaio_2013). In addition, multiple clinical diagnostic laboratories/reputable databases have classified this variant as pathogenic. Functional studies in zebra fish have shown that the N581D variant results in developmental defects that can be rescued by FGF-MAPK pathway inhibitors (Anastasaki_2009). Taken together, this variant is classified as Pathogenic. -

Noonan syndrome 1 Pathogenic:1

Pathogenic, no assertion criteria provided

clinical testing

Molecular Genetics, Centre for Human Genetics

-

- -

Cardiovascular phenotype Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Ambry Genetics

May 25, 2023

The c.1741A>G (p.N581D) alteration is located in exon 14 (coding exon 14) of the BRAF gene. This alteration results from an A to G substitution at nucleotide position 1741. This variant was not reported in population-based cohorts in the Genome Aggregation Database (gnomAD). This alteration has been reported in multiple individuals with features consistent with BRAF-related RASopathy, including multiple cases with reported de novo occurrences (Niihori, 2006; Rodriguez-Viciana, 2006; Yoon, 2007; Hazan, 2012; Ciara, 2014; Battaglia, 2021; Pierpont, 2022). Another alteration at the same codon, c.1741A>C (p.N581H), has been reported de novo in an individual with features of BRAF-related RASopathy (Sparks, 2020). This amino acid position is highly conserved in available vertebrate species. The p.N581D amino acid is located in the catalytic loop of BRAF (Niihori, 2006). Experimental studies on the effect of cardiofaciocutaneous syndrome mutant alleles in an in vivo zebrafish model system showed that the p.N581D alteration can result in developmental abnormalities in zebrafish when expressed during early development. The mutant embryos were shown to respond to treatment with inhibitors of the FGF-MAPK pathway (Anastasaki, 2009). The in silico prediction for this alteration is inconclusive. Based on the available evidence, this alteration is classified as pathogenic. -

RASopathy Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Labcorp Genetics (formerly Invitae), Labcorp

Jul 25, 2022

This variant is not present in population databases (gnomAD no frequency). This sequence change replaces asparagine, which is neutral and polar, with aspartic acid, which is acidic and polar, at codon 581 of the BRAF protein (p.Asn581Asp). This missense change has been observed in individual(s) with features of cardio-facio-cutaneous (CFC) syndrome (PMID: 16439621, 16474404, 17366577, 18042262, 22876591, 24037001, 25463315). In at least one individual the variant was observed to be de novo. For these reasons, this variant has been classified as Pathogenic. Experimental studies have shown that this missense change affects BRAF function (PMID: 19376813). Algorithms developed to predict the effect of missense changes on protein structure and function are either unavailable or do not agree on the potential impact of this missense change (SIFT: "Deleterious"; PolyPhen-2: "Benign"; Align-GVGD: "Class C15"). ClinVar contains an entry for this variant (Variation ID: 13979). -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.99
BayesDel_addAF	Pathogenic	0.27	D
BayesDel_noAF	Pathogenic	0.15
CADD	Pathogenic	33
DANN	Uncertain	1.0
DEOGEN2	Pathogenic	0.96	.;.;D;.
Eigen	Pathogenic	0.97
Eigen_PC	Pathogenic	0.88
FATHMM_MKL	Pathogenic	0.99	D
LIST_S2	Benign	0.80	T;T;T;T
M_CAP	Pathogenic	0.55	D
MetaRNN	Pathogenic	0.99	D;D;D;D
MetaSVM	Pathogenic	0.94	D
MutationAssessor	Pathogenic	3.4	.;.;M;.
MutationTaster	Benign	1.0	A
PrimateAI	Pathogenic	0.88	D
REVEL	Pathogenic	0.93
Polyphen		1.0	.;.;D;.
MutPred		0.98		Loss of ubiquitination at K578 (P = 0.1113);.;Loss of ubiquitination at K578 (P = 0.1113);.;
MVP		0.92
MPC		1.3
ClinPred		0.99	D
GERP RS		5.3
Varity_R		0.93
gMVP		1.0

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Benign	0.44
SpliceAI score (max)		0.15		Details are displayed if max score is > 0.2

Find out detailed SpliceAI scores and Pangolin per-transcript scores at spliceailookup.broadinstitute.org

Publications

LitVar

Other links and lift over

dbSNP: rs180177040; hg19: chr7-140453987; COSMIC: COSV56286119;