rs377729736

Positions:

• chr7-117531117-A-G

Variant summary

Our verdict is Uncertain significance. Variant got 4 ACMG points: 4P and 0B. PM2 PP3 PP5

The NM_000492.4(CFTR):​c.489+3A>G variant causes a splice donor region, intron change. The variant allele was found at a frequency of 0.000178 in 1,605,062 control chromosomes in the GnomAD database, with no homozygous occurrence. In-silico tool predicts a benign outcome for this variant. Variant has been reported in ClinVar as Conflicting classifications of pathogenicity (no stars).

• Frequency:
  • Genomes: 𝑓 0.00015 (0 hom., cov: 32)
  • Exomes 𝑓: 0.00018 (0 hom.)
• Consequence: CFTR NM_000492.4 splice_donor_region, intron
• Scores: Splicing: ADA: 0.9994
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications P:12 U:9
• Conservation: PhyloP100: 4.74

Genes affected

CFTR (HGNC:1884): (CF transmembrane conductance regulator) This gene encodes a member of the ATP-binding cassette (ABC) transporter superfamily. The encoded protein functions as a chloride channel, making it unique among members of this protein family, and controls ion and water secretion and absorption in epithelial tissues. Channel activation is mediated by cycles of regulatory domain phosphorylation, ATP-binding by the nucleotide-binding domains, and ATP hydrolysis. Mutations in this gene cause cystic fibrosis, the most common lethal genetic disorder in populations of Northern European descent. The most frequently occurring mutation in cystic fibrosis, DeltaF508, results in impaired folding and trafficking of the encoded protein. Multiple pseudogenes have been identified in the human genome. [provided by RefSeq, Aug 2017]

ACMG classification

Verdict is Uncertain_significance. Variant got 4 ACMG points.

• PM2: Very rare variant in population databases, with high coverage
• PP3: Splicing scoreres supports a deletorius effect: Scorers claiming Pathogenic: dbscSNV1_ADA, dbscSNV1_RF, max_spliceai. No scorers claiming Uncertain. No scorers claiming Benign.
• PP5: Variant 7-117531117-A-G is Pathogenic according to our data. Variant chr7-117531117-A-G is described in ClinVar as [Conflicting_classifications_of_pathogenicity]. Clinvar id is 53971.We mark this variant Likely_pathogenic, oryginal submissions are: {Likely_pathogenic=8, Uncertain_significance=7, Pathogenic=3}. Variant chr7-117531117-A-G is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
CFTR	NM_000492.4	c.489+3A>G		splice_donor_region_variant, intron_variant		ENST00000003084.11	NP_000483.3

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
CFTR	ENST00000003084.11	c.489+3A>G		splice_donor_region_variant, intron_variant		1	NM_000492.4	ENSP00000003084	P2

Frequencies

GnomAD3 genomes AF: 0.000151 AC: 23 AN: 152196 Hom.: 0 Cov.: 32

Gnomad AFR AF: 0.0000724

Gnomad AMI AF: 0.00

Gnomad AMR AF: 0.0000655

Gnomad ASJ AF: 0.00

Gnomad EAS AF: 0.00

Gnomad SAS AF: 0.00

Gnomad FIN AF: 0.0000942

Gnomad MID AF: 0.00

Gnomad NFE AF: 0.000250

Gnomad OTH AF: 0.000478

GnomAD3 exomes AF: 0.000249 AC: 61 AN: 245226 Hom.: 0 AF XY: 0.000302 AC XY: 40 AN XY: 132416

Gnomad AFR exome AF: 0.0000628

Gnomad AMR exome AF: 0.000294

Gnomad ASJ exome AF: 0.00

Gnomad EAS exome AF: 0.00

Gnomad SAS exome AF: 0.00

Gnomad FIN exome AF: 0.0000469

Gnomad NFE exome AF: 0.000427

Gnomad OTH exome AF: 0.000334

GnomAD4 exome AF: 0.000180 AC: 262 AN: 1452866 Hom.: 0 Cov.: 29 AF XY: 0.000181 AC XY: 131 AN XY: 722966

Gnomad4 AFR exome AF: 0.0000300

Gnomad4 AMR exome AF: 0.000293

Gnomad4 ASJ exome AF: 0.00

Gnomad4 EAS exome AF: 0.00

Gnomad4 SAS exome AF: 0.0000234

Gnomad4 FIN exome AF: 0.0000377

Gnomad4 NFE exome AF: 0.000205

Gnomad4 OTH exome AF: 0.000183

GnomAD4 genome AF: 0.000151 AC: 23 AN: 152196 Hom.: 0 Cov.: 32 AF XY: 0.000148 AC XY: 11 AN XY: 74348

Gnomad4 AFR AF: 0.0000724

Gnomad4 AMR AF: 0.0000655

Gnomad4 ASJ AF: 0.00

Gnomad4 EAS AF: 0.00

Gnomad4 SAS AF: 0.00

Gnomad4 FIN AF: 0.0000942

Gnomad4 NFE AF: 0.000250

Gnomad4 OTH AF: 0.000478

Alfa AF: 0.000352 Hom.: 0

Bravo AF: 0.000181

Asia WGS AF: 0.000866 AC: 3 AN: 3478

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Pathogenic:12 Uncertain:9

Revision: criteria provided, conflicting classifications

Submissions by phenotype

Cystic fibrosis Pathogenic:7 Uncertain:3

Likely pathogenic, criteria provided, single submitter	clinical testing	Genome-Nilou Lab	Jul 22, 2021	- -
Likely pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Jun 26, 2024	Variant summary: CFTR c.489+3A>G, legacy name c.621+3A>G, alters a conserved nucleotide located close to a canonical splice site and therefore could affect mRNA splicing, leading to a significantly altered protein sequence. 4/4 in silico tools via Alamut predict that the variant abolishes or weakens the canonical 5' splicing donor site. This is further supported by experimental evidence reporting an impact on mRNA splicing as corroborated by two independent studies that showed a reproducible distribution of transcripts resulting from activation of an alternate splice site (approx 28.3%) and transcripts resulting from complete skipping of exon 4 (approx 6.8%) in addition to levels of wild-type transcripts (approx remaining 65%) (Tzetis_2001, Forzan_2010). Furthermore, the pattern of abnormal splicing demonstrated by this variant is very similar to that of c.489+1G>T (legacy name c.621+1G>T) considered to be a well-recognized pathogenic CFTR variant (Tzetis_2001). The variant allele was found at a frequency of 0.00025 in 247812 control chromosomes. This frequency is not significantly higher than estimated for a pathogenic variant in CFTR causing Non-Classic Cystic Fibrosis (0.00025 vs 0.013), allowing no conclusion about variant significance. c.489+3A>G has been reported in the literature in individuals affected with Cystic Fibrosis or with a provisional CF diagnosis with intermediate sweat chloride levels (e.g. Tzetis_2001, Kanavakis_2003, Soltysova_2018, Claustres_2017, Angyal_2024) and in individuals with other CFTR-related diseases such as CBAVD (e.g. Amato_2012), azoospermia (e.g. Gallati_2009), sarcoidosis (Bombieri_2000), and chronic pancreatitis (Sofia_2016). These data, when ascertained conservatively, limited to patients with a confirmed diagnosis of CF, support the notion that the variant is likely to be associated with disease. At least one report of an asymptomatic 8 year old girl harboring this variant in trans with another pathogenic CFTR variant (p.Q552*) has been ascertained (Forzan_2010). This patient had inconsistent sweat chloride levels ranging from 49-75 mEq/L in three independent measurements and the possibility of subclinical disease cannot be entirely ruled out. In addition, the variant was also reported to have been observed in cis with c.4332delTG (legacy name) in a CF patient harboring c.2183AA/G (legacy name) on the other allele (Loumi_2008). However, to our knowledge, there are no additional reports of this co-occurrence to substantiate this finding. The following publications have been ascertained in the context of this evaluation (PMID: 26990548, 22020151, 22439019, 10980579, 28603918, 25304080, 25308578, 19893581, 20021716, 17850636, 20657600, 20932301, 12752573, 17572159, 27264265, 28544683, 11810271). The CFTR2 database states this variant to have varying consequences. ClinVar contains an entry for this variant (Variation ID: 53971). Based on the evidence outlined above, the variant was classified as likely pathogenic. -
Likely pathogenic, criteria provided, single submitter	clinical testing	Johns Hopkins Genomics, Johns Hopkins University	Jan 29, 2020	CFTR variant associated with variable clinical consequences. See www.CFTR2.org for phenotype information. -
Uncertain significance, criteria provided, single submitter	clinical testing	Illumina Laboratory Services, Illumina	Dec 07, 2018	The CFTR c.489+3A>G variant, also referred to as c.621+3A>G, has been reported in six studies in which it is identified in a compound heterozygous state in at least six individuals and in a heterozygous state in at least five individuals with varying clinical consequences including cystic fibrosis, CBAVD, azoospermia, and sarcoidosis (Bombieri et al. 2000; Tzetis et al. 2001; Loumi et al. 2008; Gallati et al. 2009; Forzan et al. 2010; Amato et al. 2012). However, one study found that the frequency of affected individuals carrying the variant was lower than expected based on the allele frequency in the control population (Forzan et al. 2010). The c.489+3A>G variant occurs in the splice region and RT-PCR analysis revealed that the variant generates both correctly spliced and incorrectly spliced products, suggesting aberrant splicing with a possibility of some residual activity of the CFTR protein (Tzetis et al. 2001). The c.489+3A>G variant has been reported at a frequency of 0.000592 in the European (non-Finnish) population of the Exome Aggregation Consortium. Based on the conflicting evidence and the clinical variability found in patients with this variant, the c.489+3A>G variant is classified as a variant of unknown significance but suspicious for pathogenicity for CFTR-related disorders. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population. -
Pathogenic, criteria provided, single submitter	clinical testing	Institute of Human Genetics, University of Leipzig Medical Center	May 22, 2024	Criteria applied: PS3_VSTR,PM3,PP4 -
Uncertain significance, criteria provided, single submitter	clinical testing	Mendelics	Nov 05, 2018	- -
Likely pathogenic, criteria provided, single submitter	clinical testing	Baylor Genetics	-	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Ambry Genetics	Dec 18, 2023	The c.489+3A>G intronic variant results from an A to G substitution 3 nucleotides after coding exon 4 in the CFTR gene. This variant (reported as 621+3A>G) was described in four Greek individuals with severe cystic fibrosis (CF), each with a second pathogenic mutation; however, the phase of these alterations is uncertain (Tzetis M et al. Hum Genet, 2001 Dec;109:592-601). In addition, this variant has been reported with a second pathogenic variant in other individuals with intermediate sweat chloride levels (Forzan M et al. J Hum Genet, 2010 Jan;55:23-6; Sermet-Gaudelus I et al. Thorax, 2010 Jun;65:539-44; Esposito MV et al. J Clin Med, 2020 Nov;9:). However, it has also been detected in trans with a pathogenic variant in multiple individuals with normal sweat chloride levels (Ambry internal data). Functional studies using RNA from patients and minigene assay showed that this variant results in partial and full exon skipping (Tzetis M et al. Hum Genet, 2001 Dec;109:592-601; Forzan M et al. J Hum Genet, 2010 Jan;55:23-6). In another RNA study, this variant has approximately 90% of wild type quantity and function (The Clinical and Functional TRanslation of CFTR (CFTR2) database available at http://cftr2.org. Accessed 12/15/2023). This nucleotide position is highly conserved in available vertebrate species. In silico splice site analysis predicts that this alteration will weaken the native splice donor site. Based on available evidence to date, this variant is unlikely to be causative of classic cystic fibrosis; however, its contribution to the development of a CFTR-related disorder is uncertain. This alteration is thus classified as a variant of unknown significance. -
Pathogenic, criteria provided, single submitter	clinical testing	Labcorp Genetics (formerly Invitae), Labcorp	Jan 31, 2024	This sequence change falls in intron 4 of the CFTR gene. It does not directly change the encoded amino acid sequence of the CFTR protein. RNA analysis indicates that this variant induces altered splicing and likely results in a shortened protein product. This variant is present in population databases (rs377729736, gnomAD 0.04%). This variant has been observed in individual(s) with cystic fibrosis, sarcoidosis, azoospermia, or congenital absence of the vas deference (PMID: 10980579, 11810271, 19893581, 20021716, 22020151). This variant is also known as 621+3A>G. ClinVar contains an entry for this variant (Variation ID: 53971). Variants that disrupt the consensus splice site are a relatively common cause of aberrant splicing (PMID: 17576681, 9536098). Studies have shown that this variant results in skipping of 4, but is expected to preserve the integrity of the reading-frame (PMID: 11810271). For these reasons, this variant has been classified as Pathogenic. -
Likely pathogenic, criteria provided, single submitter	clinical testing	MGZ Medical Genetics Center	Jan 13, 2022	- -

not provided Pathogenic:1 Uncertain:4

Likely pathogenic, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Jul 05, 2023	The CFTR c.489+3A>G variant has been reported in the published literature in individuals with cystic fibrosis (CF) who carried a CFTR pathogenic variant on the opposite chromosome (PMID: 11810271 (2001)). It has also been observed in individuals with sarcoidosis (PMID: 10980579 (2000)), azoospermia (PMID: 20021716 (2009)), bronchiectasis (PMID: 33260873 (2020)), and congenital bilateral absence of the vas deferens (CBAVD) (PMID: 22020151 (2012)). However, it has also been observed in an asymptomatic individual who carried a CFTR pathogenic variant on the opposite chromosome (PMID: 1989381 (2010)). RNA studies have shown that it causes the synthesis of aberrantly spliced CFTR mRNA transcripts, but it does not prevent the synthesis of normally spliced CFTR mRNA (PMID: 11810271 (2001), 19893581 (2010)). The frequency of this variant in the general population, 0.0021 (23/11152 chromosomes (Genome Aggregation Database, http://gnomad.broadinstitute.org)), is uninformative in the assessment of its pathogenicity. Analysis of this variant using software algorithms for the prediction of the effect of nucleotide changes on splicing yielded predictions that this variant may affect proper CFTR mRNA splicing. Based on the available information, this variant is classified as likely pathogenic. -
Uncertain significance, no assertion criteria provided	clinical testing	Clinical Genetics DNA and cytogenetics Diagnostics Lab, Erasmus MC, Erasmus Medical Center	-	- -
Uncertain significance, criteria provided, single submitter	clinical testing	GeneDx	Jul 28, 2017	The c.489+3A>G variant in the CFTR gene, also referred to as c.621+3A>G, has been reported in the heterozygous state patients with sarcoidosis, CAVD, and azoospermia without CAVD (Bombieri et al., 2000; Amato et al., 2012; Gallati et al., 2009). This variant reduces the quality of the splice donor site in intron 4, and may cause abnormal gene splicing. The c.489+3A>G variant is observed in 30/50716 (0.06%) alleles in the ExAC dataset (Lek et al., 2016). We interpret c.489+3A>G as a variant of uncertain significance. -
Uncertain significance, no assertion criteria provided	clinical testing	Joint Genome Diagnostic Labs from Nijmegen and Maastricht, Radboudumc and MUMC+	-	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Eurofins Ntd Llc (ga)	Nov 22, 2017	- -

CFTR-related disorder Pathogenic:2

Pathogenic, criteria provided, single submitter	curation	CFTR-France	Mar 09, 2018	- -
Likely pathogenic, no assertion criteria provided	clinical testing	PreventionGenetics, part of Exact Sciences	Mar 05, 2024	The CFTR c.489+3A>G variant is predicted to interfere with splicing. This variant has been reported in patients with cystic fibrosis, congenital bilateral absence of vas deferens (CBAVD), azoospermia, or sarcoidosis, including in patients with a pathogenic variant on the opposite chromosome (Amato et al. 2011. PubMed ID: 22020151; Bombieri et al. 2000. PubMed ID: 10980579; Gallati et al. 2009. PubMed ID: 20021716; Giuliani et al. 2010. PubMed ID: 20657600; Kanavakis et al. 2003. PubMed ID: 12752573; Sermet-Gaudelus et al. 2010. PubMed ID: 20522854; Soltysova et al. 2017. PubMed ID: 28544683; Tzetis et al. 2001. PubMed ID: 11810271). This variant was also reported in trans with a second pathogenic variant in an asymptomatic female child; however, her sweat chloride testing suggested possible subclinical disease (49-75 mEq/L) (Forzan et al. 2009. PubMed ID: 19893581). Additionally, functional studies demonstrate that this variant leads to aberrant splicing with a defect consistent with other known splicing variants at this location (c.489+1G>T aka 621+1G>T; Tzetis et al. 2001. PubMed ID: 11810271; Forzan et al. 2009. PubMed ID: 19893581). This variant is reported in 0.039% of alleles in individuals of European (Non-Finnish) descent in gnomAD. Taken together, this variant is interpreted as likely pathogenic. -

Hereditary pancreatitis Pathogenic:1 Uncertain:1

Likely pathogenic, criteria provided, single submitter	clinical testing	Eurofins-Biomnis	Oct 20, 2022	- -
Uncertain significance, criteria provided, single submitter	curation	Sema4, Sema4	Apr 21, 2021	- -

Bronchiectasis with or without elevated sweat chloride 1 Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

Baylor Genetics

Mar 28, 2024

- -

not specified Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

ARUP Laboratories, Molecular Genetics and Genomics, ARUP Laboratories

Apr 06, 2019

The CFTR c.489+3A>G variant (rs377729736), also known as 621+3A>G, is reported in the literature in individuals affected with cystic fibrosis, congenital bilateral absence of vas deferens (CBAVD), and sarcoidosis, including individuals with a severe pathogenic variant on the opposite chromosome (Amato 2012, Gallati 2009, Giuliani 2010, Sermet-Gaudelus 2010, Soltysova 2018, Tzetis 2001). However, this variant has also been reported in an asymptomatic individual who carries a pathogenic nonsense variant in trans (Forzan 2010), and in cis with a truncating variant in an individual with cystic fibrosis (Loumi 2008). This variant is reported in ClinVar (Variation ID: 53971), and is found in the general population with an overall allele frequency of 0.023% (63/276,630 alleles) in the Genome Aggregation Database. This is an intronic variant in a highly conserved nucleotide, and computational analyses (Alamut v.2.11) predict that this variant may impact splicing by weakening the nearby canonical donor splice site. Functional studies show aberrant mRNA splicing making up about 35% of transcripts, but it is unknown if this level altered transcripts is sufficient to cause disease (Forzan 2010, Tzetis 2001). Due to conflicting information, the clinical significance of the c.489+3A>G variant is uncertain at this time. References: Amato F et al. Extensive molecular analysis of patients bearing CFTR-related disorders. J Mol Diagn. 2012 Jan;14(1):81-9. Forzan M et al. Is CFTR 621+3 A>G a cystic fibrosis causing mutation? J Hum Genet. 2010 Jan;55(1):23-6. Gallati S et al. Cystic fibrosis transmembrane conductance regulator mutations in azoospermic and oligospermic men and their partners. Reprod Biomed Online. 2009 Nov;19(5):685-94. Giuliani R et al. Identification of the second CFTR mutation in patients with congenital bilateral absence of vas deferens undergoing ART protocols. Asian J Androl. 2010 Nov;12(6):819-26. Loumi O et al. CFTR mutations in the Algerian population. J Cyst Fibros. 2008 Jan;7(1):54-9. Epub 2007 Jun 14. Sermet-Gaudelus I et al. Measurement of nasal potential difference in young children with an equivocal sweat test following newborn screening for cystic fibrosis. Thorax. 2010 Jun;65(6):539-44. Soltysova A et al. Comprehensive genetic study of cystic fibrosis in Slovak patients in 25 years of genetic diagnostics. Clin Respir J. 2018 Mar;12(3):1197-1206. Tzetis M et al. Qualitative and quantitative analysis of mRNA associated with four putative splicing mutations (621+3A-->G, 2751+2T-->A, 296+1G-->C, 1717-9T-->C-D565G) and one nonsense mutation (E822X) in the CFTR gene. Hum Genet. 2001 Dec;109(6):592-601. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_noAF	Benign	-0.18
CADD	Benign	17
DANN	Benign	0.97

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.95
SpliceAI score (max)		0.72		Details are displayed if max score is > 0.2
DS_DL_spliceai		0.72		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: rs377729736; hg19: chr7-117171171;