chr3-12604188-G-A

Position:

• chr3-12604188-G-A

Variant summary

Our verdict is Pathogenic. Variant got 17 ACMG points: 17P and 0B. PM1 PM2 PM5 PP2 PP3_Moderate PP5_Very_Strong

The NM_002880.4(RAF1):​c.782C>T​(p.Pro261Leu) variant causes a missense change involving the alteration of a conserved nucleotide. The variant was absent in control chromosomes in GnomAD project. In-silico tool predicts a pathogenic outcome for this variant. Variant has been reported in ClinVar as Likely pathogenic (★★). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. P261A) has been classified as Pathogenic.

• Frequency: Genomes: not found (cov: 33)
• Consequence: RAF1 NM_002880.4 missense
• Clinical Significance: Pathogenic/Likely pathogenic criteria provided, multiple submitters, no conflicts P:6 O:1
• Conservation: PhyloP100: 10.0

Genes affected

RAF1 (HGNC:9829): (Raf-1 proto-oncogene, serine/threonine kinase) This gene is the cellular homolog of viral raf gene (v-raf). The encoded protein is a MAP kinase kinase kinase (MAP3K), which functions downstream of the Ras family of membrane associated GTPases to which it binds directly. Once activated, the cellular RAF1 protein can phosphorylate to activate the dual specificity protein kinases MEK1 and MEK2, which in turn phosphorylate to activate the serine/threonine specific protein kinases, ERK1 and ERK2. Activated ERKs are pleiotropic effectors of cell physiology and play an important role in the control of gene expression involved in the cell division cycle, apoptosis, cell differentiation and cell migration. Mutations in this gene are associated with Noonan syndrome 5 and LEOPARD syndrome 2. [provided by RefSeq, Jul 2008]

ACMG classification

Verdict is Pathogenic. Variant got 17 ACMG points.

• PM1: In a compositionally_biased_region Polar residues (size 47) in uniprot entity RAF1_HUMAN there are 14 pathogenic changes around while only 0 benign (100%) in NM_002880.4
• PM2: Very rare variant in population databases, with high coverage
• PM5: Other missense variant is known to change same aminoacid residue: Variant chr3-12604189-G-C is described in Lovd as [Pathogenic].
• PP2: Missense variant in gene, where missense usually causes diseases (based on misZ statistic), RAF1. . Gene score misZ 2.4628 (greater than the threshold 3.09). Trascript score misZ 3.4185 (greater than threshold 3.09). GenCC has associacion of gene with Noonan syndrome 5, dilated cardiomyopathy 1NN, Noonan syndrome with multiple lentigines, Noonan syndrome, cardiofaciocutaneous syndrome, familial isolated dilated cardiomyopathy, LEOPARD syndrome 2, Costello syndrome.
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.877
• PP5: Variant 3-12604188-G-A is Pathogenic according to our data. Variant chr3-12604188-G-A is described in ClinVar as [Likely_pathogenic]. Clinvar id is 120246.Status of the report is criteria_provided_multiple_submitters_no_conflicts, 2 stars. Variant chr3-12604188-G-A is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
RAF1	NM_002880.4	c.782C>T	p.Pro261Leu	missense_variant	7/17	ENST00000251849.9	NP_002871.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
RAF1	ENST00000251849.9	c.782C>T	p.Pro261Leu	missense_variant	7/17	1	NM_002880.4	ENSP00000251849	P3

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 33

ClinVar

Significance: Pathogenic/Likely pathogenic

Submissions summary: Pathogenic:6 Other:1

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Noonan syndrome Pathogenic:2

Likely pathogenic, no assertion criteria provided	literature only	Database of Curated Mutations (DoCM)	May 13, 2016	- -
Pathogenic, criteria provided, single submitter	clinical testing	Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine	Nov 19, 2013	The Pro261Leu variant in RAF1 has been reported as a de novo variant in one indi vidual with clinical features of Noonan syndrome (Pandit 2007). Recurrent missen se variants at this amino acid position (Pro261Leu, Pro261Ser, Pro261Thr, Pro261 Ala, and Pro261Arg) have been identified by our laboratory in individuals with c linical features associated with Noonan syndrome. Of note, individuals with path ogenic variants in exon 7 or 17 in RAF1 have a high incidence of hypertrophic ca rdiomyopathy (80-95%, Razzaque 2007, Pandit 2007). In summary, this variant meet s our criteria to be classified as pathogenic based on its de novo occurrence (h ttp://pcpgm.partners.org/LMM). -

Dilated cardiomyopathy 1NN Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Genomics England Pilot Project, Genomics England

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not provided Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

GeneDx

Aug 01, 2017

The P261L variant in the RAF1 gene has been reported previously in an individual with Noonan syndrome with clinical features including short stature, hypertrophic cardiomyopathy, and pectus anomaly (Pandit et al., 2007). The P261L variant is not observed in large population cohorts (Lek et al., 2016; 1000 Genomes Consortium et al., 2015; Exome Variant Server). The P261L variant is a semi-conservative amino acid substitution, which may impact secondary protein structure as these residues differ in some properties. This substitution is not within a known functional domain and occurs at a position that is conserved across species. Functional studies have shown that P261L leads to increased activity of the RAF1 protein (Molzan et al., 2010). Missense variants in the same residue (P261A, P261T, P261S, P261H, P261R) and in nearby residues (R256S, S257L, S259P, S259T, S259F, T260I, T260R, N262I, N262K, V263D, V263A) have been reported in association with RAF1-related disorders (Stenson et al., 2014), supporting the functional importance of this region of the protein. We interpret P261L as a pathogenic variant. -

Cardiovascular phenotype Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Ambry Genetics

Oct 03, 2023

The p.P261L pathogenic mutation (also known as c.782C>T), located in coding exon 6 of the RAF1 gene, results from a C to T substitution at nucleotide position 782. The proline at codon 261 is replaced by leucine, an amino acid with similar properties. This alteration has been reported as de novo in a subject with features of Noonan syndrome (Pandit B et al. Nat Genet, 2007 Aug;39:1007-12). This alteration has also been reported in subjects with features of Noonan syndrome and overlapping disorders (Kobayashi T et al. Hum Mutat, 2010 Mar;31:284-94; Justino A et al. Eur J Hum Genet, 2015 Mar;23:347-53; Lazzaro G et al. Mol Genet Genomic Med, 2020 Apr;8:e1069). Two other alterations at the same codon, p.P261A (c.781C>G) and p.P261S (c.781C>T), have been described in association with Noonan syndrome (Razzaque MA, Nat. Genet. 2007 Aug; 39(8):1013-7; Longoni M et al. Am. J. Med. Genet. A, 2010 Sep;152A:2176-84). This variant is considered to be rare based on population cohorts in the Genome Aggregation Database (gnomAD). This amino acid position is highly conserved in available vertebrate species. In addition, this alteration is predicted to be deleterious by in silico analysis. Based on the supporting evidence, this alteration is interpreted as a disease-causing mutation. -

RASopathy Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Labcorp Genetics (formerly Invitae), Labcorp

Nov 17, 2023

This sequence change replaces proline, which is neutral and non-polar, with leucine, which is neutral and non-polar, at codon 261 of the RAF1 protein (p.Pro261Leu). This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individual(s) with Noonan syndrome (PMID: 17603483). In at least one individual the variant was observed to be de novo. ClinVar contains an entry for this variant (Variation ID: 120246). Advanced modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) performed at Invitae indicates that this missense variant is expected to disrupt RAF1 protein function with a positive predictive value of 95%. Experimental studies have shown that this missense change affects RAF1 function (PMID: 20679480). This variant disrupts the p.Pro261 amino acid residue in RAF1. Other variant(s) that disrupt this residue have been determined to be pathogenic (PMID: 17603482, 20052757, 21784453, 22465605, 23885229, 29084544). This suggests that this residue is clinically significant, and that variants that disrupt this residue are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic. -

Noonan syndrome 5 Other:1

not provided, no classification provided

literature only

Institute of Molecular Pathology and Immunology of the University of Porto (IPATIMUP)

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Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	1.0
BayesDel_addAF	Pathogenic	0.43	D
BayesDel_noAF	Pathogenic	0.37
CADD	Pathogenic	32
DANN	Uncertain	1.0
DEOGEN2	Pathogenic	0.89	D;.;T
Eigen	Pathogenic	0.87
Eigen_PC	Pathogenic	0.81
FATHMM_MKL	Uncertain	0.93	D
LIST_S2	Pathogenic	1.0	D;D;D
M_CAP	Uncertain	0.20	D
MetaRNN	Pathogenic	0.88	D;D;D
MetaSVM	Uncertain	0.39	D
MutationAssessor	Pathogenic	3.0	M;M;.
MutationTaster	Benign	1.0	D;D;D;D
PrimateAI	Pathogenic	0.81	D
PROVEAN	Pathogenic	-6.3	D;D;D
REVEL	Pathogenic	0.88
Sift	Uncertain	0.0010	D;D;D
Sift4G	Uncertain	0.0020	D;D;D
Polyphen		1.0	D;.;.
Vest4		0.87
MutPred		0.66		Gain of glycosylation at S257 (P = 0.0137);Gain of glycosylation at S257 (P = 0.0137);.;
MVP		0.95
MPC		1.4
ClinPred		0.99	D
GERP RS		5.7
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.7
Varity_R		0.84
gMVP		0.73

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.0		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: rs397516828; hg19: chr3-12645687; COSMIC: COSV52583046; COSMIC: COSV52583046;