rs540635787

chr22-28694073-G-Achr22-28694073-G-Cchr22-28694073-G-T

Variant summary

Our verdict is Likely pathogenic. Variant got 7 ACMG points: 7P and 0B. PM5 PP3_Strong PP5

The NM_007194.4(CHEK2):c.1420C>T(p.Arg474Cys) variant causes a missense change involving the alteration of a non-conserved nucleotide. The variant allele was found at a frequency of 0.0000201 in 1,595,950 control chromosomes in the GnomAD database, with no homozygous occurrence. 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. R474H) has been classified as Likely pathogenic.

• Frequency:
  • Genomes: 𝑓 0.000013 (0 hom., cov: 32)
  • Exomes 𝑓: 0.000021 (0 hom.)
• Consequence: CHEK2 NM_007194.4 missense
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications P:6 U:11
• Conservation: PhyloP100: 3.16

Genes affected

CHEK2 (HGNC:16627): (checkpoint kinase 2) In response to DNA damage and replication blocks, cell cycle progression is halted through the control of critical cell cycle regulators. The protein encoded by this gene is a cell cycle checkpoint regulator and putative tumor suppressor. It contains a forkhead-associated protein interaction domain essential for activation in response to DNA damage and is rapidly phosphorylated in response to replication blocks and DNA damage. When activated, the encoded protein is known to inhibit CDC25C phosphatase, preventing entry into mitosis, and has been shown to stabilize the tumor suppressor protein p53, leading to cell cycle arrest in G1. In addition, this protein interacts with and phosphorylates BRCA1, allowing BRCA1 to restore survival after DNA damage. Mutations in this gene have been linked with Li-Fraumeni syndrome, a highly penetrant familial cancer phenotype usually associated with inherited mutations in TP53. Also, mutations in this gene are thought to confer a predisposition to sarcomas, breast cancer, and brain tumors. This nuclear protein is a member of the CDS1 subfamily of serine/threonine protein kinases. Several transcript variants encoding different isoforms have been found for this gene. [provided by RefSeq, Apr 2012]

ACMG classification

Verdict is Likely_pathogenic. Variant got 7 ACMG points.

• PM5: Other missense variant is known to change same aminoacid residue: Variant chr22-28694072-C-T is described in ClinVar as [Conflicting_classifications_of_pathogenicity]. Clinvar id is 126910.We mark this variant Likely_pathogenic, oryginal submissions are: {Uncertain_significance=7, Likely_pathogenic=7, Likely_benign=1, not_provided=1}.
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.983
• PP5: Variant 22-28694073-G-A is Pathogenic according to our data. Variant chr22-28694073-G-A is described in ClinVar as [Conflicting_classifications_of_pathogenicity]. Clinvar id is 128059.We mark this variant Likely_pathogenic, oryginal submissions are: {Likely_pathogenic=4, Uncertain_significance=9}.

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
CHEK2	NM_007194.4	c.1420C>T	p.Arg474Cys	missense_variant	13/15	ENST00000404276.6

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
CHEK2	ENST00000404276.6	c.1420C>T	p.Arg474Cys	missense_variant	13/15	1	NM_007194.4	P2

Frequencies

GnomAD3 genomes AF: 0.0000131 AC: 2 AN: 152114 Hom.: 0 Cov.: 32

Gnomad AFR AF: 0.00

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: 0.00

Gnomad MID AF: 0.00

Gnomad NFE AF: 0.0000147

Gnomad OTH AF: 0.00

GnomAD3 exomes AF: 0.0000171 AC: 4 AN: 233784 Hom.: 0 AF XY: 0.0000234 AC XY: 3 AN XY: 128346

Gnomad AFR exome AF: 0.00

Gnomad AMR exome AF: 0.0000291

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: 0.0000276

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.0000208 AC: 30 AN: 1443718 Hom.: 0 Cov.: 30 AF XY: 0.0000306 AC XY: 22 AN XY: 718616

Gnomad4 AFR exome AF: 0.00

Gnomad4 AMR exome AF: 0.0000447

Gnomad4 ASJ exome AF: 0.00

Gnomad4 EAS exome AF: 0.0000252

Gnomad4 SAS exome AF: 0.0000116

Gnomad4 FIN exome AF: 0.00

Gnomad4 NFE exome AF: 0.0000234

Gnomad4 OTH exome AF: 0.00

GnomAD4 genome AF: 0.0000131 AC: 2 AN: 152232 Hom.: 0 Cov.: 32 AF XY: 0.00 AC XY: 0 AN XY: 74422

Gnomad4 AFR AF: 0.00

Gnomad4 AMR AF: 0.0000655

Gnomad4 ASJ AF: 0.00

Gnomad4 EAS AF: 0.00

Gnomad4 SAS AF: 0.00

Gnomad4 FIN AF: 0.00

Gnomad4 NFE AF: 0.0000147

Gnomad4 OTH AF: 0.00

Bravo AF: 0.00000756

ExAC AF: 0.00000866 AC: 1

EpiCase AF: 0.0000545

EpiControl AF: 0.00

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Pathogenic:6 Uncertain:11

Revision: criteria provided, conflicting classifications

Submissions by phenotype

not provided Pathogenic:3 Uncertain:4

Likely pathogenic, criteria provided, single submitter	clinical testing	Center for Genomic Medicine, Rigshospitalet, Copenhagen University Hospital	Feb 06, 2024	- -
Likely pathogenic, criteria provided, single submitter	clinical testing	CeGaT Center for Human Genetics Tuebingen	Apr 01, 2024	CHEK2: PS3, PM1, PM5:Supporting -
Likely pathogenic, criteria provided, single submitter	clinical testing	GeneDx	Aug 22, 2023	Published functional studies demonstrate a damaging effect: decreased CHEK2 protein expression and impaired kinase activity (Kukita et al., 2016; Delimitsou et al., 2019; Stolarova et al., 2023); In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; Not observed at significant frequency in large population cohorts (gnomAD); This variant is associated with the following publications: (PMID: 25980754, 26506619, 22138346, 26094658, 19782031, 27900359, 29522266, 29520813, 28580595, 30426508, 29731985, 30851065, 30287823, 30858171, 31398194, 31159747, 33309985, 36061833, 34991090, 35661486, 32980694, 22419737, 33471991, 36243179, 37449874) -
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	Mayo Clinic Laboratories, Mayo Clinic	Apr 30, 2020	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Nov 04, 2022	The frequency of this variant in the general population, 0.000028 (3/108758 chromosomes, http://gnomad.broadinstitute.org), is uninformative in assessment of its pathogenicity. In the published literature, the variant has been reported in individuals with breast cancer (PMIDs: 30426508 (2018), 30287823 (2018), 28580595 (2018), and 33471991 (2021)), prostate cancer (PMID: 29520813 (2018)), colorectal cancer (PMID: 33309985 (2020)), as well as in healthy individuals (PMIDs: 32980694 (2020), 33309985 (2020), and 33471991 (2021)). Additionally, the variant was reported in an individual with suspected Lynch Syndrome (PMID: 25980754 (2015)). Yeast-based and mammalian studies have reported that this variant may have a deleterious effect on CHEK2 protein function. Further studies are needed to validate these findings (PMIDs: 27900359 (2016) and 30851065 (2019)). Analysis of this variant using bioinformatics tools for the prediction of the effect of amino acid changes on protein structure and function yielded predictions that this variant is damaging. Based on the available information, we are unable to determine the clinical significance of this variant. -
Uncertain significance, no assertion criteria provided	clinical testing	Clinical Genetics Laboratory, Department of Pathology, Netherlands Cancer Institute	-	- -

Familial cancer of breast Pathogenic:1 Uncertain:2

Uncertain significance, criteria provided, single submitter	clinical testing	Invitae	Jan 19, 2024	This sequence change replaces arginine, which is basic and polar, with cysteine, which is neutral and slightly polar, at codon 474 of the CHEK2 protein (p.Arg474Cys). This variant is present in population databases (rs540635787, gnomAD 0.003%). This missense change has been observed in individual(s) with breast cancer, lung cancer, Lynch syndrome-associated cancer(s), and/or prostate cancer (PMID: 25980754, 27900359, 28580595, 29520813, 30287823, 30426508, 33309985). This variant is also known as c.1549C>T (p.Arg517Cys). ClinVar contains an entry for this variant (Variation ID: 128059). An algorithm developed to predict the effect of missense changes on protein structure and function (PolyPhen-2) suggests that this variant is likely to be disruptive. Experimental studies have shown that this missense change affects CHEK2 function (PMID: 27900359, 30851065). 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 pathogenic, criteria provided, single submitter	clinical testing	Myriad Genetics, Inc.	Jun 28, 2023	This variant is considered likely pathogenic. Functional studies indicate this variant impacts protein function [PMID: 27900359]. This variant is strongly associated with more severe personal and family histories of cancer, typical for individuals with pathogenic variants in this gene [PMID: 25085752]. -
Uncertain significance, criteria provided, single submitter	clinical testing	Baylor Genetics	Jul 02, 2023	- -

Hereditary cancer-predisposing syndrome Uncertain:3

Uncertain significance, criteria provided, single submitter	clinical testing	Ambry Genetics	Nov 29, 2023	The p.R474C variant (also known as c.1420C>T), located in coding exon 12 of the CHEK2 gene, results from a C to T substitution at nucleotide position 1420. The arginine at codon 474 is replaced by cysteine, an amino acid with highly dissimilar properties. This alteration has been detected in the homozygous state in siblings with multiple primary lung cancers; the sister was also affected with breast cancer and uterine myoma and the brother was also affected with prostate and colon cancers (Kukita Y et al Cold Spring Harb. Mol. Case Stud. 2016 Nov;2(6):a001032). In addition, this alteration has been reported in the heterozygous state in multiple cohorts of affected patients with various tumor types (Wu Y et al. Prostate. 2018 Jun;78(8):607-615; Momozawa Y et al. Nat Commun, 2018 Oct;9:4083; Schubert S et al. Int. J. Cancer 2019 06;144(11):2683-2694; Xie Y et al. Clin. Genet. 2018 Jan;93(1):41-51; Tsaousis GN et al. BMC Cancer 2019 Jun;19(1):535; Dorling et al. N Engl J Med. 2021 02;384:428-439; Aksoy F et al. Hum Hered, 2022 Jan;:). This alteration behaved as non-functional in an in vivo, yeast-based growth rate assay (Delimitsou A et al. Hum. Mutat. 2019 05;40(5):631-648). In addition, this alteration was reported as functionally impaired in a study assessing CHEK2-complementation through quantification of KAP1 phosphorylation and CHK2 autophosphorylation in human RPE1-CHEK2-knockout cells (Stolarova L et al. Clin Cancer Res. 2023 Aug;29(16):3037-3050). The p.R474C variant has been shown to eliminate a salt bridge between the core kinase domain and the highly conserved activation loop, and is critical for stability and function of Ser/Thr kinases (Ambry internal analysis; Cai Z et al. Mol. Cell. 2009 Sep;35:818-29; Yang J et al. J. Mol. Biol. 2012 Jan;415:666-79). 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. -
Uncertain significance, criteria provided, single submitter	clinical testing	Color Diagnostics, LLC DBA Color Health	Jan 09, 2023	This missense variant replaces arginine with cysteine at codon 474 of the CHEK2 protein. This variant impacts a highly conserved arginine in the kinase domain of the protein (PMID: 15060014, 19782031). Computational prediction suggests that this variant may have deleterious impact on protein structure and function (internally defined REVEL score threshold >= 0.7, PMID: 27666373). Functional studies have reported that this variant may impact CHEK2 protein stability and DNA damage response (PMID: 27900359, 30851065). This variant has been reported in at least six individuals affected with breast cancer (PMID: 28580595, 29522266, 30287823, 30426508, 34991090) and in a breast cancer case-control meta-analysis in 6/60466 cases and 2/53461 unaffected individuals (PMID: 33471991; Leiden Open Variation Database DB-ID CHEK2_000093). This variant also has been reported in individuals affected with colorectal and prostate cancer (PMID: 25980754, 29520813, 33309985) and in an unaffected individual in a pancreatic cancer case-control study (PMID: 32980694). This variant has been identified in 4/233784 chromosomes in the general population by the Genome Aggregation Database (gnomAD). The available evidence is insufficient to determine the role of this variant in disease conclusively. Therefore, this variant is classified as a Variant of Uncertain Significance. -
Uncertain significance, criteria provided, single submitter	clinical testing	GeneKor MSA	Aug 01, 2018	- -

Familial cancer of breast;C0376358:Malignant tumor of prostate;C0684249:Lung carcinoma;C0699790:Carcinoma of colon Pathogenic:1

Likely pathogenic, no assertion criteria provided

research

Department of Molecular and Medical Genetics, Osaka Medical Center for Cancer and Cardiovascular Diseases

-

Two siblings, an elder sister and a younger brother, were found to have multiple primary lung cancers at the age of 60. The former subsequently developed breast cancer, and had a history of myoma uteri. The latter had initially developed prostate cancer at the age of 59, and had a history of colon cancer. This variant was homozygous in both patients. -

Li-Fraumeni syndrome 2 Pathogenic:1

Likely pathogenic, no assertion criteria provided

clinical testing

Department of Genetics, HCU Lozano Blesa

May 01, 2023

Variant summary: CHEK2 c.1420C>T results in the replacement of Arg474 by a Cys residue (p.Arg474Cys). The variant was identified in 1 out of 396 patients analysed (freq: 0.0025, doi: 10.3389/fgene.2023.1274108). The patient was a woman that developed breast cancer at the age of 69 years (luminal-A tumor phenotype). Two second-degree relatives were also diagnosed with BC, although a co-segregation study could not be performed. Arg474 is located in the kinase domain and appears well conserved in vertebrate species. The replacement of Arg474 by a cysteine could affect the interaction with some residues at the dimerization interface, particularly with Ala392. It thus may affect the protein integrity and therefore its function. In addition, Arg474, forms a cation/π interaction with Trp411 (at the same monomer) and a salt bridge with Glu394 (at the partner monomer). Replacement of Arg474 with a cysteine residue will therefore remove the indicated stabilizing interactions of Arg474. The variant is reported in gnomAD v4 in 32 cases out of 1595950 alleles analysed (freq=2.0x10-3 %). In ClinVar a dozen of reports appear classifying the variant as of Uncertain Significance, whereas in 3 reports the submitters have suggested the classification of Likely Pathogenic. Other replacements found at this position (R474L, R474G, R474S and R474H) are classified by ClinVar as of either Uncertain Significance or Conflicting Interpretation. The metapredictor PirePred (relies on verdicts from other 15 predictor or metapredictor tools) and the AI-based predictor AlphaMissense classify Arg474Cys as Pathogenic, whereas the ACMG classification tool Franklin suggests this variant as Likely Pathogenic. Our in-silico study on the protein stability based on relaxation molecular dynamics simulations (doi: 10.3389/fgene.2023.1274108) indicates that Arg474Cys induces conformational unstability on CHEK2 protein. Moreover, Stolarova L. et al’s functional study on KAP1 phosphorylation and CHEK2 autophosphorylation protein capability reports this variant as functionally impaired (PMID: 37449874). In summary, we believe this variant have more chances of being Pathogenic. -

Familial cancer of breast;C0376358:Malignant tumor of prostate;C0585442:Bone osteosarcoma;C1836482:Li-Fraumeni syndrome 2 Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

Fulgent Genetics, Fulgent Genetics

Oct 31, 2018

- -

not specified Uncertain:1

Uncertain significance, criteria provided, single submitter

clinical testing

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

Mar 16, 2022

Variant summary: CHEK2 c.1420C>T (p.Arg474Cys) results in a non-conservative amino acid change located in the protein kinase-like domain of the encoded protein sequence. Five of five in-silico tools predict a damaging effect of the variant on protein function. This variant reportedly alters the tertiary structure of CHEK2 by disrupting the salt bridge between p.R474 and p.E394 suggesting that protein stability of both inactive and active states might be disturbed (Kukita_2016, Delimitsou_2019). The variant allele was found at a frequency of 1.7e-05 in 233784 control chromosomes (gnomAD). It has also been reported in the FLOSSIES database of women over the age of 70 with no history of cancer. c.1420C>T has been reported in the literature in individuals affected with a a variety of cancer types such as colorectal cancer, prostate cancer, breast cancer and lung cancer (e.g. Kukita_2016, Wu_2016, Schubert_2019), including two homozygous siblings who experienced multiple primary lung cancers as well as cancers in other organs (Kukita_2016). These data indicate that the variant may be associated with disease. The variant was determined to be damaging in an in vivo yeast functional assay (Delimitsou_2019). Kukita_2016 also found the variant expression protein did not increase nor activate in response to UV damage. Nine ClinVar submitters have assessed this variant since 2014: eight classified the variant as of uncertain significance and one as likely pathogenic. Some submitters cite overlapping evidence utilized in the context of this evaluation. Based on the evidence outlined above, the variant was classified as VUS-possibly pathogenic. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Pathogenic	0.97
BayesDel_addAF	Pathogenic	0.37	D
BayesDel_noAF	Pathogenic	0.37
Cadd	Pathogenic	27
Dann	Pathogenic	1.0
DEOGEN2	Uncertain	0.51	D;.;D;.;D;.;D;.;.
Eigen	Pathogenic	0.79
Eigen_PC	Uncertain	0.63
FATHMM_MKL	Uncertain	0.85	D
M_CAP	Uncertain	0.14	D
MetaRNN	Pathogenic	0.98	D;D;D;D;D;D;D;D;D
MetaSVM	Pathogenic	0.92	D
MutationAssessor	Pathogenic	4.8	H;.;H;.;H;.;H;.;.
MutationTaster	Benign	1.0	D;D;D;D;D;D;D;D;D;D;D
PrimateAI	Uncertain	0.52	T
PROVEAN	Pathogenic	-7.4	D;D;D;.;D;D;.;D;D
REVEL	Pathogenic	0.90
Sift	Pathogenic	0.0	D;D;D;.;D;D;.;D;D
Sift4G	Pathogenic	0.0	D;D;D;.;D;D;.;D;D
Polyphen		1.0	D;D;D;.;D;D;D;D;D
Vest4		0.96
MutPred		0.98		Loss of disorder (P = 0.0362);.;Loss of disorder (P = 0.0362);.;Loss of disorder (P = 0.0362);.;Loss of disorder (P = 0.0362);.;.;
MVP		0.95
MPC		0.16
ClinPred		1.0	D
GERP RS		4.4
RBP_binding_hub_radar		0.0
RBP_regulation_power_radar		1.1
Varity_R		0.97
gMVP		0.94

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.010		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: rs540635787; hg19: chr22-29090061; COSMIC: COSV60418058; COSMIC: COSV60418058;