Submissions for variant NM_007194.4(CHEK2):c.320-5T>A

gnomAD frequency: 0.00070  dbSNP: rs121908700

Total submissions: 22

Submitter	RCV	SCV	Clinical significance	Condition	Last evaluated	Review status	Method	Comment
GeneDx	RCV000119289	SCV000149920	likely benign	not provided	2021-05-19	criteria provided, single submitter	clinical testing	Patient RNA studies demonstrate aberrant splicing resulting in an in-frame transcript lacking exons 3 and 4 present at approximately 20%, as well as full-length transcript (Kraus 2017, Vargas-Parra 2020); In-silico analysis is inconclusive as to whether the variant alters gene splicing; Also known as IVS1-5T>A; This variant is associated with the following publications: (PMID: 18058223, 20643596, 21744992, 25980754, 24549055, 18996005, 27067391, 27616075, 27621404, 26506619, 29596542, 25186627, 30306255, 31422574, 28779002, 30374176, 29337092, 32906215)
Ambry Genetics	RCV000116011	SCV000172844	likely benign	Hereditary cancer-predisposing syndrome	2018-11-08	criteria provided, single submitter	clinical testing	This alteration is classified as likely benign based on a combination of the following: seen in unaffected individuals, population frequency, intact protein function, lack of segregation with disease, co-occurrence, RNA analysis, in silico models, amino acid conservation, lack of disease association in case-control studies, and/or the mechanism of disease or impacted region is inconsistent with a known cause of pathogenicity.
Labcorp Genetics (formerly Invitae), Labcorp	RCV000195943	SCV000253484	benign	Familial cancer of breast	2024-02-01	criteria provided, single submitter	clinical testing
Quest Diagnostics Nichols Institute San Juan Capistrano	RCV000212412	SCV000601165	benign	not specified	2021-12-16	criteria provided, single submitter	clinical testing
ARUP Laboratories, Molecular Genetics and Genomics, ARUP Laboratories	RCV000119289	SCV000603086	uncertain significance	not provided	2020-07-09	criteria provided, single submitter	clinical testing	The CHEK2 c.320-5T>A variant has been described in the literature in individuals affected with different cancers, including breast and/or ovarian cancer, Hodgkin lymphoma, and colorectal cancer (Castera 2014, Hampel 2018, Havranek 2011, Kleibl 2008, Kleibl 2009, Kraus 2017, Tung 2015). However, several affected individuals with this variant were found to carry an additional pathogenic variant (Hampel 2018, Tung 2015). The c.320-5T>A variant is found in the general population with an overall allele frequency of 0.05% (152/282198 alleles, including one homozygote) in the Genome Aggregation Database, including an increased frequency of 0.37% (38/10362 alleles) in the Ashkenazi Jewish population. This is an intronic variant in a moderately conserved nucleotide, and computational analyses (Alamut v.2.11) predict that this variant may impact splicing by weakening the nearby canonical acceptor splice site. Functional analyses of patient RNAs show exon skipping in a small minority of transcripts (Casadei 2019, Kraus 2017); however, it is unclear that these splicing alterations are clinically significant. While existing evidence suggests this variant is unlikely to cause disease in all carriers, given the possibility of a low penetrance effect, the clinical significance of the c.320-5T>A variant is uncertain at this time. References: Casadei S et al. Characterization of splice-altering mutations in inherited predisposition to cancer. Proc Natl Acad Sci U S A. 2019;116(52):26798-26807. Castera L et al. Next-generation sequencing for the diagnosis of hereditary breast and ovarian cancer using genomic capture targeting multiple candidate genes. Eur J Hum Genet. 2014 Nov;22(11):1305-13. Hampel H et al. Assessment of Tumor Sequencing as a Replacement for Lynch Syndrome Screening and Current Molecular Tests for Patients With Colorectal Cancer. JAMA Oncol. 2018;4(6):806-813. Havranek O et al. Alterations of CHEK2 forkhead-associated domain increase the risk of Hodgkin lymphoma. Neoplasma. 2011;58(5):392-5. Kleibl Z et al. Analysis of CHEK2 FHA domain in Czech patients with sporadic breast cancer revealed distinct rare genetic alterations. Breast Cancer Res Treat. 2008 Nov;112(1):159-64. Kleibl Z et al. The CHEK2 gene I157T mutation and other alterations in its proximity increase the risk of sporadic colorectal cancer in the Czech population. Eur J Cancer. 2009 Mar;45(4):618-24. Kraus C et al. Gene panel sequencing in familial breast/ovarian cancer patients identifies multiple novel mutations also in genes others than BRCA1/2. Int J Cancer. 2017;140(1):95-102. Mucaki EJ et al. A unified analytic framework for prioritization of non-coding variants of uncertain significance in heritable breast and ovarian cancer. BMC Med Genomics. 2016 Apr 11;9:19. Tung N et al. Frequency of mutations in individuals with breast cancer referred for BRCA1 and BRCA2 testing using next-generation sequencing with a 25-gene panel. Cancer. 2015;121(1):25-33.
University of Washington Department of Laboratory Medicine, University of Washington	RCV000195943	SCV000611874	likely benign	Familial cancer of breast	2018-03-28	criteria provided, single submitter	research	The CHEK2 variant designated as NM_007194.3:c.320-5T>A is classified as likely benign. Cosegregation analysis of one observed family was performed using analyze.myvariant.org (Rañola et al, 2018, PMID:28965303) and shows a likelihood ratio of 1.38 to 1 (Thompson, et al., 2003, PMID:12900794). This likelihood ratio indicates weak evidence that the allele co-segregates disease in this family. However, RNA studies in two individuals unrelated from different families show no significant change in RNA splicing or protein transcripts, thus providing strong evidence that this variant does not alter the CHEK2 protein. This variant has been classified as likely benign by other laboratories (ClinVar Variation ID: 128070). Bayesian analysis integrating all of this data (Tavtigian et al, 2018, PMID:29300386) gives less than 1% probability of pathogenicity, which is consistent with a classification of likely benign. This variant is not predicted to alter CHEK2 function or modify cancer risk. This reclassification analysis was performed in conjunction with the family studies project as part of the University of Washington Find My Variant Study.
Women's Health and Genetics/Laboratory Corporation of America, LabCorp	RCV000212412	SCV000698794	benign	not specified	2019-04-11	criteria provided, single submitter	clinical testing	Variant summary: CHEK2 c.320-5T>A alters a conserved nucleotide located close to a canonical splice site and therefore could affect mRNA splicing, leading to a significantly altered protein sequence. 5/5 computational tools predict the variant slightly weakens a 3 acceptor site. An in silico analysis also predicted the variant to result in a reduced splicing efficiency that could activate pre-existing cryptic acceptor 92 nucleotide upstream of the natural site, leading to the inclusion of 92 nucleotide of the intron. However this effect was predicted to be partial, with the variant mRNA relatively less abundant compared to the wild type (Mucaki 2016). Two publications reported experimental evidence evaluating splicing impact: one identified an in frame transcript variant lacking exon 3 and 4, however, the amount of the shortened transcript compared to wild type was less than 20%, suggesting that this variant reduces but does not eliminate the usage of the consensus splice acceptor and can therefore be regarded as a hypomorphic allele (Kraus 2017); whereas the other study reported splice analysis from 2 patients showing no significant change in RNA splicing, however without providing experimental evidence (Tsai 2018). The variant allele was found at a frequency of 0.00053 in 288862 control chromosomes (gnomAD and publication data), including 1 homozygote. The observed variant frequency is approximately 1.7 fold of the estimated maximal expected allele frequency for a pathogenic variant in CHEK2 causing Breast Cancer phenotype (0.00031), strongly suggesting that the variant is benign. In addition, this variant has been reported in 10/9884 American women who are older than age 70 and cancer free (FLOSSIES database). c.320-5T>A has been reported in the literature in individuals affected with various tumor phenotypes, including breast-/ovarian-, colorectal- and pancreas cancer, non-Hodgkin lymphoma and Lynch syndrome (Kleibl 2008, Kleibl 2009, Mohelnikova-Duchonova 2010, Havranek 2015, Castera 2014, Tung 2015, Yurgelun 2015, Kraus 2017, Decker 2017, Bonache 2018, Hampel 2018). These reports however do not provide unequivocal conclusions about association of the variant with Breast Cancer. In addition, in two of these patients co-occurrences with other pathogenic variants have been reported (CHEK2 exon 2 deletion, Tung 2015; MLH1 c.1381A>T/K461X, Hampel 2018), providing supporting evidence for a benign role. Though one case-control study suggested that the variant might be associated with breast cancer, the carrier frequency in healthy controls recruited in this study was much lower than observed in population datasets, therefore this postulation might be the result of sampling error (Decker 2017). Nine other clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation. Multiple laboratories reported the variant with conflicting assessments (6 calling it a VUS, 2 classifying as likely benign, and 1 as benign). Based on the evidence outlined above, the variant was classified as benign.
Counsyl	RCV000195943	SCV000786412	uncertain significance	Familial cancer of breast	2018-04-26	criteria provided, single submitter	clinical testing
Mendelics	RCV003492504	SCV000839498	likely benign	Hereditary cancer	2024-01-23	criteria provided, single submitter	clinical testing
Color Diagnostics, LLC DBA Color Health	RCV000116011	SCV000902592	likely benign	Hereditary cancer-predisposing syndrome	2016-05-13	criteria provided, single submitter	clinical testing
Illumina Laboratory Services, Illumina	RCV004556727	SCV000915970	uncertain significance	CHEK2-related cancer predisposition	2017-04-28	criteria provided, single submitter	clinical testing	The CHEK2 c.320-5T>A splice region variant has been reported in a heterozygous state in at least five individuals with various cancers (Kleibl et al. 2008; Kleibl et al. 2009; Mohelnikova-Duchonova et al. 2010; Havranek et al. 2011; Kraus et al. 2016; Mucaki et al. 2016). Of these five individuals, the variant was found in one patient with sporadic breast cancer, one with colorectal cancer, one with sporadic pancreatic cancer, one with Hodgkin lymphoma, and one with hereditary breast or ovarian cancer. The c.320-5T>A variant was absent from 1366 control chromosomes (Kleibl et al. 2008), but is reported at a frequency of 0.01402 in the Iberian population in Spain from the 1000 Genomes Project. Kraus et al. (2016) demonstrated using RT-PCR analysis that the c.320-5T>A variant resulted in an in-frame transcript lacking exons 3 and 4 that was stably expressed, however at low levels (about 20%) compared to the level of wild type transcript, suggesting a leaky splicing defect and a hypomorphic allele. Based on the evidence, the c.320-5T>A variant is classified as a variant of unknown significance but suspicious for pathogenicity for CHEK2-related cancer susceptibility. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population.
CeGaT Center for Human Genetics Tuebingen	RCV000119289	SCV001245717	likely benign	not provided	2022-10-01	criteria provided, single submitter	clinical testing	CHEK2: BP4, BS1:Supporting, BS2
CHEO Genetics Diagnostic Laboratory, Children's Hospital of Eastern Ontario	RCV001798354	SCV002043401	uncertain significance	Breast and/or ovarian cancer	2023-04-27	criteria provided, single submitter	clinical testing
Genetic Services Laboratory, University of Chicago	RCV000212412	SCV002070850	likely benign	not specified	2021-01-08	criteria provided, single submitter	clinical testing
Sema4, Sema4	RCV000116011	SCV002537418	likely benign	Hereditary cancer-predisposing syndrome	2021-07-21	criteria provided, single submitter	curation
Center for Genomic Medicine, Rigshospitalet, Copenhagen University Hospital	RCV000212412	SCV002761116	likely benign	not specified	2023-08-15	criteria provided, single submitter	clinical testing
Myriad Genetics, Inc.	RCV000195943	SCV004020194	likely benign	Familial cancer of breast	2023-03-09	criteria provided, single submitter	clinical testing	This variant is considered likely benign. This variant is strongly associated with less severe personal and family histories of cancer, typical for individuals without pathogenic variants in this gene [PMID: 25085752].
Institute. of Biochemistry and Experimental Oncology, First Faculty of Medicine, Charles University in Prague	RCV000119289	SCV000154113	not provided	not provided		no assertion provided	not provided
PreventionGenetics, part of Exact Sciences	RCV004528807	SCV000806878	uncertain significance	CHEK2-related disorder	2023-12-06	no assertion criteria provided	clinical testing	The CHEK2 c.320-5T>A variant is predicted to interfere with splicing. This variant has been reported in individuals with breast and colorectal cancer and Hodgkin lymphoma (Kleibl et al. 2008. PubMed ID: 18058223; Kleibl et al. 2009. PubMed ID: 18996005; Havranek et al. 2011. PubMed ID: 21744992). RNA studies on blood from a patient carrying this variant identified a novel CHEK2 transcript with in-frame omission of exons 3 and 4. However, this transcript was detected at a level <20% of normal, prompting the authors to conclude it may be a hypomorphic allele (Kraus et al. 2017. PubMed ID: 27616075). This variant is reported in 0.37% of alleles in individuals of Ashkenazi Jewish descent in gnomAD. This variant has also been reported in ClinVar with conflicting interpretations from benign to uncertain (https://www.ncbi.nlm.nih.gov/clinvar/variation/128070/). Although we suspect that this variant may be benign, at this time, the clinical significance of this variant is uncertain due to the absence of conclusive functional and genetic evidence.
King Laboratory, University of Washington	RCV001171460	SCV001251371	pathogenic	Familial cancer of breast; Li-Fraumeni syndrome 2	2019-09-01	no assertion criteria provided	research
Department of Pathology and Laboratory Medicine, Sinai Health System	RCV000119289	SCV001551631	likely benign	not provided		no assertion criteria provided	clinical testing	The CHEK2 c.320-5T>A variant was identified in 40 of 33888 proband chromosomes (frequency: 0.001) from individuals or families with breast, ovarian, pancreatic, and colon cancer, as well as non-Hodkin lymphoma and the variant was present in 1 of 10976 control chromosomes (frequency: 0.00009) from healthy individuals (Decker 2017, Havranek 2015, Mucaki 2016, Yurgelun 2015, Kraus 2017, Kleibl 2008, Kleibl 2009, Mohelnikova-Duchonova 2010). The variant was also identified in dbSNP (ID: rs121980700) as “With uncertain significance”. The variant was also identified in ClinVar (as benign by Invitae, likely benign by University of Washington, LabCorp, and Ambry and uncertain significance by Quest, ARUP and Gene Dx). The variant was not identified in Cosmic, MutDB or Zhejiang University databases. The variant was identified in control databases in 151 of 276520 chromosomes (1 homozygous) at a frequency of 0.0005 (Genome Aggregation Database Feb 27, 2017). It was observed in the following populations: “Other” in 6 of 6458 chromosomes (freq: 0.009), Latino in 33 of 34412 chromosomes (freq: 0.001), European Non-Finnish in 71 of 126074 chromosomes (freq: 0.0006), Ashkenazi Jewish in 40 of 10146 chromosomes (freq: 0.004), and South Asian in 1 of 30776 chromosomes (freq: 0.00003). The variant was not observed in the African, East Asian, or Finnish populations. The c.320-5T>A variant is located in the 3' splice region but does not affect the invariant -1 and -2 positions. However, positions -3 and -5 to -12 are part of the splicing consensus sequence and variants involving these positions sometimes affect splicing. Reverse transcription PCR analysis on blood from an individual with breast or ovarian cancer demonstrated that this variant leads to an in frame transcript lacking exon 3 and 4; however the shortened transcript was present at levels less than 20% that of the wild-type transcript suggesting that this variant does not abolish normal splicing (Kraus, 2017). However, 3 of 5 in silico or computational prediction software programs (SpliceSiteFinder, MaxEntScan, NNSPLICE, GeneSplicer, HumanSpliceFinder) predict a greater than 10% difference in splicing. In addition the variant was identified as co-occurring with a pathogenic BRCA2 variant (c.5857G>T, p.Glu1953X) by our laboratory in an affected individual, indicating that the variant c.320-5T>A may not have clinical significance. In summary, based on the above information the clinical significance of this variant cannot be determined with certainty at this time although we would lean towards a more benign role for this variant. This variant is classified as likely benign.
Institute for Biomarker Research, Medical Diagnostic Laboratories, L.L.C.	RCV000116011	SCV001977053	benign	Hereditary cancer-predisposing syndrome	2021-09-27	no assertion criteria provided	clinical testing