rs104894833

chrX-101403984-C-G

Variant summary

Our verdict is Likely benign. Variant got -2 ACMG points: 6P and 8B. PM1 PM5 PP3_Moderate BS1 BS2

The NM_000169.3(GLA):c.196G>C(p.Glu66Gln) variant causes a missense, splice region change involving the alteration of a conserved nucleotide. The variant allele was found at a frequency of 0.000132 in 1,203,291 control chromosomes in the GnomAD database, including 2 homozygotes. There are 49 hemizygotes in GnomAD. In-silico tool predicts a pathogenic outcome for this variant. Variant has been reported in ClinVar as Conflicting classifications of pathogenicity (no stars). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. E66G) has been classified as Likely pathogenic.

• Frequency:
  • Genomes: 𝑓 0.000036 (0 hom., 0 hem., cov: 23)
  • Exomes 𝑓: 0.00014 (2 hom. 49 hem.)
• Consequence: GLA NM_000169.3 missense, splice_region
• Scores: Splicing: ADA: 0.9018
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications U:7 B:5
• Conservation: PhyloP100: 7.86

Genes affected

GLA (HGNC:4296): (galactosidase alpha) This gene encodes a homodimeric glycoprotein that hydrolyses the terminal alpha-galactosyl moieties from glycolipids and glycoproteins. This enzyme predominantly hydrolyzes ceramide trihexoside, and it can catalyze the hydrolysis of melibiose into galactose and glucose. A variety of mutations in this gene affect the synthesis, processing, and stability of this enzyme, which causes Fabry disease, a rare lysosomal storage disorder that results from a failure to catabolize alpha-D-galactosyl glycolipid moieties. [provided by RefSeq, Jul 2008]

RPL36A-HNRNPH2 (HGNC:):

ACMG classification

Verdict is Likely_benign. Variant got -2 ACMG points.

• PM1: In a hotspot region, there are 7 aminoacids with missense pathogenic changes in the window of +-8 aminoacids around while only 0 benign, 9 uncertain in NM_000169.3
• PM5: Other missense variant is known to change same aminoacid residue: Variant chrX-101403983-T-C is described in ClinVar as [Conflicting_classifications_of_pathogenicity]. Clinvar id is 2689132.We mark this variant Likely_pathogenic, oryginal submissions are: {Likely_pathogenic=1, Uncertain_significance=1}.
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.908
• BS1: Variant frequency is greater than expected in population eas. gnomad4_exome allele frequency = 0.000142 (155/1090911) while in subpopulation EAS AF= 0.00507 (153/30189). AF 95% confidence interval is 0.00441. There are 2 homozygotes in gnomad4_exome. There are 49 alleles in male gnomad4_exome subpopulation. Median coverage is 29. This position pass quality control queck.
• BS2: High Hemizygotes in GnomAdExome at 5 XL gene

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
GLA	NM_000169.3	c.196G>C	p.Glu66Gln	missense_variant, splice_region_variant	2/7	ENST00000218516.4
RPL36A-HNRNPH2	NM_001199973.2	c.301-7952C>G		intron_variant

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
GLA	ENST00000218516.4	c.196G>C	p.Glu66Gln	missense_variant, splice_region_variant	2/7	1	NM_000169.3	P1

Frequencies

GnomAD3 genomes AF: 0.0000356 AC: 4 AN: 112326 Hom.: 0 Cov.: 23 AF XY: 0.00 AC XY: 0 AN XY: 34504

Gnomad AFR AF: 0.00

Gnomad AMI AF: 0.00

Gnomad AMR AF: 0.00

Gnomad ASJ AF: 0.00

Gnomad EAS AF: 0.00111

Gnomad SAS AF: 0.00

Gnomad FIN AF: 0.00

Gnomad MID AF: 0.00

Gnomad NFE AF: 0.00

Gnomad OTH AF: 0.00

GnomAD3 exomes AF: 0.000115 AC: 21 AN: 182566 Hom.: 0 AF XY: 0.0000745 AC XY: 5 AN XY: 67110

Gnomad AFR exome AF: 0.00

Gnomad AMR exome AF: 0.00

Gnomad ASJ exome AF: 0.00

Gnomad EAS exome AF: 0.00152

Gnomad SAS exome AF: 0.00

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.00

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.000142 AC: 155 AN: 1090911 Hom.: 2 Cov.: 29 AF XY: 0.000137 AC XY: 49 AN XY: 356565

Gnomad4 AFR exome AF: 0.00

Gnomad4 AMR exome AF: 0.0000284

Gnomad4 ASJ exome AF: 0.00

Gnomad4 EAS exome AF: 0.00507

Gnomad4 SAS exome AF: 0.00

Gnomad4 FIN exome AF: 0.00

Gnomad4 NFE exome AF: 0.00

Gnomad4 OTH exome AF: 0.0000218

GnomAD4 genome AF: 0.0000356 AC: 4 AN: 112380 Hom.: 0 Cov.: 23 AF XY: 0.00 AC XY: 0 AN XY: 34568

Gnomad4 AFR AF: 0.00

Gnomad4 AMR AF: 0.00

Gnomad4 ASJ AF: 0.00

Gnomad4 EAS AF: 0.00111

Gnomad4 SAS AF: 0.00

Gnomad4 FIN AF: 0.00

Gnomad4 NFE AF: 0.00

Gnomad4 OTH AF: 0.00

Bravo AF: 0.0000302

ExAC AF: 0.000107 AC: 13

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Uncertain:7 Benign:5

Revision: criteria provided, conflicting classifications

Submissions by phenotype

Fabry disease Uncertain:2 Benign:4

Likely benign, criteria provided, single submitter	clinical testing	Mendelics	May 28, 2019	- -
Likely benign, criteria provided, single submitter	clinical testing	Genome-Nilou Lab	Jul 15, 2021	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Illumina Laboratory Services, Illumina	Apr 27, 2017	This variant was observed as part of a predisposition screen in an ostensibly healthy population. A literature search was performed for the gene, cDNA change, and amino acid change (where applicable). Publications were found based on this search. However, the evidence from the literature, in combination with allele frequency data from public databases where available, was not sufficient to rule this variant in or out of causing disease. Therefore, this variant is classified as a variant of unknown significance. -
Likely benign, criteria provided, single submitter	clinical testing	3billion	May 22, 2022	This variant is observed at a frequency of 0.148% within East Asian subpopulation with 5 hemizygous male in gnomAD v2.1.1 dataset (BS1_S, BS2_S, total allele frequency: 0.01%). Missense variant of GLA gene is a common mechanism associated with Fabry disease (PP2_P). In silico prediction tools and conservation analysis predicted that this variant was probably damaging to the protein structure/function (PP3_P, 3CNET: 0.990, REVEL: 0.926). Therefore, this variant is classified as likely benign according to the recommendation of ACMG/AMP guideline. -
Uncertain significance, criteria provided, single submitter	clinical testing	Invitae	Nov 03, 2022	This sequence change replaces glutamic acid, which is acidic and polar, with glutamine, which is neutral and polar, at codon 66 of the GLA protein (p.Glu66Gln). This variant is present in population databases (rs104894833, gnomAD 0.1%), and has an allele count higher than expected for a pathogenic variant. This missense change has been observed in individual(s) with classic, non-classic, and late onset Fabry disease and/or hypertrophic cardiomyopathy and renal failure without accumulation of Gb-3 or lysosomal deposits reported on renal or cardiac biopsy (PMID: 1315715, 7575533, 11137837, 20505683, 22874111, 23146289, 26179544, 26456105, 27160240). It has also been observed to segregate with disease in related individuals. ClinVar contains an entry for this variant (Variation ID: 163548). Algorithms developed to predict the effect of missense changes on protein structure and function (SIFT, PolyPhen-2, Align-GVGD) all suggest that this variant is likely to be disruptive. Experimental studies have shown that this missense change does not substantially affect GLA function (PMID: 22305854, 26179544). Algorithms developed to predict the effect of sequence changes on RNA splicing suggest that this variant may create or strengthen a splice site. 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. -
Likely benign, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Jan 15, 2019	- -

not specified Uncertain:2 Benign:1

Uncertain significance, no assertion criteria provided	clinical testing	Laboratory for Molecular Medicine, Mass General Brigham Personalized Medicine	Feb 11, 2013	proposed classification - variant undergoing re-assessment, contact laboratory -
Uncertain significance, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Mar 09, 2023	Variant summary: GLA c.196G>C (p.Glu66Gln) results in a conservative amino acid change in the encoded protein sequence. Four of five in-silico tools predict a damaging effect of the variant on protein function. The variant allele was found at a frequency of 0.0023 in 249895 control chromosomes, predominantly at a frequency of 0.008 within the Japanese subpopulation, and including at least 88 hemizygous males and 3 homozygous females (gnomAD v2, Lee_2010, jMorp database, Tadaka_2021). The observed variant frequency within the Japanese subpopulation is approximately 1.6 fold of the estimated maximal expected allele frequency for a pathogenic variant in GLA causing Fabry disease (0.005) in control chromomes, suggesting that the variant is benign. c.196G>C has been reported in the literature in sequencing studies of individuals affected with a variety of Fabry disease-related phenotypes. A comprehensive review of literature spanning over two decades identified its occurrence in patients with reported/suspected diagnosis of Fabry disease (example, Ishii_1992, Park_2009, Lee_2010, Shimotori_2007, Sakuraba_2018), cohorts of male hemodialysis patients (Doi_2012, Maruyama_2013), patients on maintenance dialysis (Nishino_2012), male ischemic stroke (Nakamura_2013, Nagamatsu_2017) and newborn screening (Hwu_2009). It has also been observed in at least one case control study of patients with chronic kidney disease in whom no association with disease progression was identified (Watanabe_2015). Examples of isolated case reports of patients with this variant include, cerebral hemorrhage (Nakamura_2010), hemodialysis (Kikumoto_2012), a male with interstitial Nephritis and no pathological or cellular characteristics of Fabry disease (Satomura_2015), a male with suspected Fabry disease due to end stage renal failure and cardiomegaly in whom no pathological characteristics of Fabry were identified (Kobayashi_2012) and Parkinsonism without classic symptoms of Fabry (Tomizawa_2015). Notably many studies ascertained above also reported this variant in patients with normal levels of lyso Gb3, a common biomarker for Fabry disease (example, Sakuraba_2018). The variant was reported in two hemizygous brothers with suspected cardiac Fabry disease (Yoshitama_2001) and also to co-segregate with the phenotype of chronic glomerulonephritis in one large Chinese family (Peng_2016). Additional reports of similar co-segregation in other kindreds will help corroborate these findings. In summary, these reports do not provide evidence for an unequivocal association of this variant with the phenotype of classic Fabry disease. At least one reported co-occurrence in cis with another pathogenic variant in the GLA gene in a male patient with classic Fabry disease has been reported (GLA c.334C>T, p.Arg112Cys), providing supporting evidence for a benign role (Ishii_1992). Several publications report experimental evidence evaluating an impact on protein function with variable findings on GLA enzyme activities in-vitro in transfected cells and patient leukocytes. The most pronounced variant effect results in less than 10% activity in some studies (example, Peng_2016, patient leukocytes),but 30%-50% of normal activity in many others (example, Park_2009, Shimotori_2007, Hwu_2009 in transfected cells). Furthermore, at least one in-vitro study evaluating kinetic parameters reported some instability at a neutral pH of 7.5 (reflective of the environment in the ER lumen) despite no impact on the enzyme kinetic properties at pH 4.5 (example, Ishii_2007). Therefore, given the wide variability in reported activities across a cross section of studies evaluated, an exact in-vivo impact of these findings on the associated pathophysiology of Fabry disease is not apparent. Eight ClinVar submitters (evaluation after 2014) have cited the variant, at-least one of whom reported other distinct and separate publications from those summarized above. These submitters reported the variant with con -
Likely benign, criteria provided, single submitter	clinical testing	GeneDx	Apr 05, 2017	This variant is considered likely benign or benign based on one or more of the following criteria: it is a conservative change, it occurs at a poorly conserved position in the protein, it is predicted to be benign by multiple in silico algorithms, and/or has population frequency not consistent with disease. -

not provided Uncertain:2

Uncertain significance, criteria provided, single submitter	clinical testing	Revvity Omics, Revvity	Jul 28, 2023	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Eurofins Ntd Llc (ga)	Aug 07, 2017	- -

Cardiovascular phenotype Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Ambry Genetics

Nov 04, 2022

The p.E66Q variant (also known as c.196G>C), located in coding exon 2 of the GLA gene, results from a G to C substitution at nucleotide position 196. The glutamic acid at codon 66 is replaced by glutamine, an amino acid with highly similar properties. This alteration has been reported in an individual with classical Fabry disease; however, he was also identified to have an additional alteration in GLA (Ishii S et al. Hum Genet, 1992 Apr;89:29-32). Additionally, this alteration was detected in several individuals with concerns for late onset Fabry disease (Yoshitama T et al. Am J Cardiol, 2001 Jan;87:71-5; Nakao S et al. Kidney Int, 2003 Sep;64:801-7; Shimotori M et al. Hum Mutat, 2008 Feb;29:331; Nakamura K et al. J Hum Genet, 2010 Apr;55:259-61; Lee BH et al. J Hum Genet, 2010 Aug;55:512-7; Kobayashi M et al. Mol Genet Metab, 2012 Dec;107:711-5; Doi K et al. J Hum Genet, 2012 Sep;57:575-9; Tomizawa Y et al. Intern Med, 2015 Oct;54:2503-6; Satomura A et al. Intern Med, 2015 Jul;54:1819-24; Oikawa M et al. BMC Cardiovasc Disord, 2016 May;16:83; Sakuraba H et al. Mol Genet Metab Rep, 2018 Dec;17:73-79; Kinoshita N et al. J Stroke Cerebrovasc Dis, 2018 Dec;27:3563-3569; Kim WS et al. J Korean Med Sci, 2019 Feb;34:e63). In vitro studies showed this alteration may not impact protein function (Hwu WL et al. Hum Mutat, 2009 Oct;30:1397-405; Park JY et al. Exp Mol Med, 2009 Jan;41:1-7). Based on data from gnomAD, the C allele has an overall frequency of 0.0108% (22/204601) total alleles studied, with 5 hemizygote(s) observed. The highest observed frequency was 0.1480% (22/14865) of East Asian alleles. This amino acid position is highly conserved in available vertebrate species. In addition, this alteration is predicted to be deleterious by in silico analysis. Since supporting evidence is limited at this time, the clinical significance of this alteration remains unclear. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Benign	0.23
CardioboostCm	Uncertain	0.89
BayesDel_addAF	Pathogenic	0.17	D
BayesDel_noAF	Pathogenic	0.44
Cadd	Pathogenic	33
Dann	Uncertain	1.0
DEOGEN2	Pathogenic	0.96	D;.
FATHMM_MKL	Pathogenic	1.0	D
LIST_S2	Uncertain	0.95	D;T
M_CAP	Pathogenic	0.99	D
MetaRNN	Pathogenic	0.89	D;D
MetaSVM	Pathogenic	0.85	D
MutationAssessor	Uncertain	2.8	M;.
MutationTaster	Benign	1.0	A
PrimateAI	Uncertain	0.59	T
PROVEAN	Uncertain	-2.8	D;.
REVEL	Pathogenic	0.93
Sift	Uncertain	0.0020	D;.
Sift4G	Uncertain	0.0030	D;.
Polyphen		0.99	D;.
Vest4		0.80
MutPred		0.96		Loss of ubiquitination at K67 (P = 0.1);.;
MVP		0.98
MPC		1.7
ClinPred		0.29	T
GERP RS		5.5
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.1
Varity_R		0.96
gMVP		0.99

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Benign	0.90
dbscSNV1_RF	Pathogenic	0.79
SpliceAI score (max)		0.44		Details are displayed if max score is > 0.2
DS_AG_spliceai		0.44		Position offset: -3

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

Publications

LitVar

Other links and lift over

dbSNP: rs104894833; hg19: chrX-100658972;