rs201442000

chr5-173235019-T-Cchr5-173235019-T-G

Variant summary

Our verdict is Uncertain significance. Variant got 4 ACMG points: 4P and 0B. PM2 PM5

The NM_004387.4(NKX2-5):c.65A>G(p.Gln22Arg) variant causes a missense change. The variant allele was found at a frequency of 0.000271 in 1,611,708 control chromosomes in the GnomAD database, with no homozygous occurrence. Variant has been reported in ClinVar as Uncertain significance (★★). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. Q22E) has been classified as Uncertain significance.

• Frequency:
  • Genomes: 𝑓 0.00012 (0 hom., cov: 33)
  • Exomes 𝑓: 0.00029 (0 hom.)
• Consequence: NKX2-5 NM_004387.4 missense
• Clinical Significance: Uncertain significance criteria provided, multiple submitters, no conflicts P:1 U:6
• Conservation: PhyloP100: 4.74

Genes affected

NKX2-5 (HGNC:2488): (NK2 homeobox 5) This gene encodes a homeobox-containing transcription factor. This transcription factor functions in heart formation and development. Mutations in this gene cause atrial septal defect with atrioventricular conduction defect, and also tetralogy of Fallot, which are both heart malformation diseases. Mutations in this gene can also cause congenital hypothyroidism non-goitrous type 5, a non-autoimmune condition. Alternative splicing results in multiple transcript variants. [provided by RefSeq, Oct 2009]

ACMG classification

Verdict is Uncertain_significance. Variant got 4 ACMG points.

• PM2: Very rare variant in population databases, with high coverage
• PM5: Other missense variant is known to change same aminoacid residue: Variant chr5-173235020-G-T is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
NKX2-5	NM_004387.4	c.65A>G	p.Gln22Arg	missense_variant	1/2	ENST00000329198.5
NKX2-5	NM_001166176.2	c.65A>G	p.Gln22Arg	missense_variant	1/2
NKX2-5	NM_001166175.2	c.65A>G	p.Gln22Arg	missense_variant	1/2
NKX2-5	XM_017009071.3	c.65A>G	p.Gln22Arg	missense_variant	1/2

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
NKX2-5	ENST00000329198.5	c.65A>G	p.Gln22Arg	missense_variant	1/2	1	NM_004387.4	P1
NKX2-5	ENST00000424406.2	c.65A>G	p.Gln22Arg	missense_variant	1/2	1
NKX2-5	ENST00000521848.1	c.65A>G	p.Gln22Arg	missense_variant	1/2	2
NKX2-5	ENST00000517440.1	c.65A>G	p.Gln22Arg	missense_variant	1/2	4

Frequencies

GnomAD3 genomes AF: 0.000125 AC: 19 AN: 152210 Hom.: 0 Cov.: 33

Gnomad AFR AF: 0.0000723

Gnomad AMI AF: 0.00

Gnomad AMR AF: 0.0000654

Gnomad ASJ AF: 0.00

Gnomad EAS AF: 0.00

Gnomad SAS AF: 0.00

Gnomad FIN AF: 0.00

Gnomad MID AF: 0.00

Gnomad NFE AF: 0.000206

Gnomad OTH AF: 0.000479

GnomAD3 exomes AF: 0.000128 AC: 31 AN: 243024 Hom.: 0 AF XY: 0.000113 AC XY: 15 AN XY: 133092

Gnomad AFR exome AF: 0.000135

Gnomad AMR exome AF: 0.0000290

Gnomad ASJ exome AF: 0.00

Gnomad EAS exome AF: 0.00

Gnomad SAS exome AF: 0.00

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.000260

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.000286 AC: 417 AN: 1459498 Hom.: 0 Cov.: 31 AF XY: 0.000278 AC XY: 202 AN XY: 726092

Gnomad4 AFR exome AF: 0.0000300

Gnomad4 AMR exome AF: 0.0000224

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: 0.000369

Gnomad4 OTH exome AF: 0.0000829

GnomAD4 genome AF: 0.000125 AC: 19 AN: 152210 Hom.: 0 Cov.: 33 AF XY: 0.000108 AC XY: 8 AN XY: 74368

Gnomad4 AFR AF: 0.0000723

Gnomad4 AMR AF: 0.0000654

Gnomad4 ASJ AF: 0.00

Gnomad4 EAS AF: 0.00

Gnomad4 SAS AF: 0.00

Gnomad4 FIN AF: 0.00

Gnomad4 NFE AF: 0.000206

Gnomad4 OTH AF: 0.000479

Alfa AF: 0.000245 Hom.: 0

Bravo AF: 0.000147

ExAC AF: 0.000174 AC: 21

EpiCase AF: 0.0000545

EpiControl AF: 0.000178

ClinVar

Significance: Uncertain significance

Submissions summary: Pathogenic:1 Uncertain:6

Revision: criteria provided, multiple submitters, no conflicts

Submissions by phenotype

Atrial septal defect 7 Uncertain:2

Uncertain significance, criteria provided, single submitter	clinical testing	Invitae	Dec 19, 2023	This sequence change replaces glutamine, which is neutral and polar, with arginine, which is basic and polar, at codon 22 of the NKX2-5 protein (p.Gln22Arg). This variant is present in population databases (rs201442000, gnomAD 0.02%), and has an allele count higher than expected for a pathogenic variant. This missense change has been observed in individual(s) with an atrial septal defect (PMID: 19181906). ClinVar contains an entry for this variant (Variation ID: 190840). 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 not expected to disrupt NKX2-5 protein function with a negative predictive value of 80%. In summary, the available evidence is currently insufficient to determine the role of this variant in disease. Therefore, it has been classified as a Variant of Uncertain Significance. -
Uncertain significance, criteria provided, single submitter	clinical testing	Victorian Clinical Genetics Services, Murdoch Childrens Research Institute	May 06, 2021	Based on the classification scheme VCGS_Germline_v1.3.4, this variant is classified as VUS-3B. Following criteria are met: 0102 - Loss of function is a known mechanism of disease in this gene and is associated with NKX2-5-related diseases, including atrial septal defect 7, with or without AV conduction defects (MIM#108900) and tetralogy of fallot (MIM#187500). (I) 0107 - This gene is associated with autosomal dominant disease. (I) 0200 - Variant is predicted to result in a missense amino acid change from glutamine to arginine. (I) 0251 - This variant is heterozygous. (I) 0302 - Variant is present in gnomAD (v2) <0.001 for a dominant condition (33 heterozygotes, 0 homozygotes). (SP) 0309 - Alternative amino acid changes at the same position have been observed in gnomAD (v3) (10 heterozygotes, 0 homozygotes). (I) 0502 - Missense variant with conflicting in silico predictions and uninformative conservation. (I) 0604 - Variant is not located in an established domain, motif, hotspot or informative constraint region. (I) 0708 - Other missense variants comparable to the one identified in this case have conflicting previous evidence for pathogenicity. These variants (p.Gln22Lys, p.Gln22Glu) have been reported as VUS and in a family with atrial septal defect (ASD) and congenital heart disease (CHD). An additional missense variant (p.Gln22Pro) with a stronger Grantham change has also been reported as both a VUS and as pathogenic, and has been observed in a patient with CHD and tetralogy of fallot (TOF) (LOVD, ClinVar, PMID: 22179962, PMID: 31824610). (I) 0808 - Previous reports of pathogenicity for this variant are conflicting. This variant has been reported as likely pathogenic and more commonly as a VUS, and has been observed in several patients with ASD and TOF (LOVD, ClinVar, PMID: 31824610, PMID: 19181906). (I) 1208 - Inheritance information for this variant is not currently available in this individual. (I) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign -

not provided Uncertain:2

Uncertain significance, criteria provided, single submitter	clinical testing	Eurofins Ntd Llc (ga)	May 26, 2017	- -
Uncertain significance, criteria provided, single submitter	clinical testing	GeneDx	Apr 30, 2021	Identified in a patient with an atrial septal defect in published literature (Draus et al., 2009); In silico analysis, which includes protein predictors and evolutionary conservation, supports that this variant does not alter protein structure/function; Reported in ClinVar (ClinVar Variant ID# 190840; Landrum et al., 2016); This variant is associated with the following publications: (PMID: 20725931, 19181906, 33835496) -

Abnormal cardiovascular system morphology Pathogenic:1

Likely pathogenic, flagged submission

clinical testing

Genetic Services Laboratory, University of Chicago

Feb 08, 2013

- -

not specified Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Women's Health and Genetics/Laboratory Corporation of America, LabCorp

Sep 24, 2022

Variant summary: NKX2-5 c.65A>G (p.Gln22Arg) results in a conservative amino acid change in the encoded protein sequence. Four of five in-silico tools predict a benign effect of the variant on protein function. The variant allele was found at a frequency of 0.00013 in 243024 control chromosomes (gnomAD), predominantly at a frequency of 0.00026 within the Non-Finnish European subpopulation in the gnomAD database. The observed variant frequency within Non-Finnish European control individuals in the gnomAD database is approximately 52 fold of the estimated maximal expected allele frequency for a pathogenic variant in NKX2-5 causing Congenital Heart Disease (5e-06), strongly suggesting that the variant is a benign polymorphism found primarily in populations of Non-Finnish European origin. c.65A>G has been reported in the literature in individuals affected with Atrial Septal Defects (Draus_2009) and Left Bundle Branch Block (Kohli_2021). These data indicate that the variant may be associated with disease. To our knowledge, no experimental evidence demonstrating an impact on protein function has been reported. Five ClinVar submitters have assessed the variant since 2014: all classified the variant as of uncertain significance. Based on the evidence outlined above, the variant was classified as VUS-possibly benign. -

Cardiovascular phenotype Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Ambry Genetics

Oct 14, 2022

The p.Q22R variant (also known as c.65A>G), located in coding exon 1 of the NKX2-5 gene, results from an A to G substitution at nucleotide position 65. The glutamine at codon 22 is replaced by arginine, an amino acid with highly similar properties. This alteration has been reported in one individual with an atrial septal defect (Draus JM, J. Med. Genet. 2009 Feb; 46(2):115-22). This amino acid position is well conserved in available vertebrate species; however, arginine is the reference amino acid in other vertebrate species. In addition, the in silico prediction for this alteration is inconclusive. Since supporting evidence is limited at this time, the clinical significance of this variant remains unclear. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Benign	0.18
BayesDel_addAF	Uncertain	0.049	T
BayesDel_noAF	Uncertain	0.13
Cadd	Uncertain	24
Dann	Uncertain	1.0
DEOGEN2	Uncertain	0.47	T;.;.;T
Eigen	Benign	-0.13
Eigen_PC	Benign	-0.0065
FATHMM_MKL	Uncertain	0.93	D
LIST_S2	Benign	0.75	T;D;D;D
M_CAP	Pathogenic	0.59	D
MetaRNN	Uncertain	0.61	D;D;D;D
MetaSVM	Uncertain	-0.20	T
MutationAssessor	Benign	1.2	L;L;L;.
MutationTaster	Benign	1.0	D;D;D
PrimateAI	Pathogenic	0.86	D
PROVEAN	Benign	-1.6	N;N;N;N
REVEL	Pathogenic	0.65
Sift	Benign	0.13	T;D;T;T
Sift4G	Benign	0.20	T;T;T;.
Polyphen		0.015	B;.;.;B
Vest4		0.76
MVP		0.95
MPC		1.2
ClinPred		0.077	T
GERP RS		2.3
Varity_R		0.17
gMVP		0.76

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: rs201442000; hg19: chr5-172662022; COSMIC: COSV61299297;