rs6476

Positions:

• chr6-32039816-T-A
• chr6-32039816-T-C

Variant summary

Our verdict is Likely benign. Variant got -6 ACMG points: 2P and 8B. PM1 BP4_Strong BS2

The NM_000500.9(CYP21A2):​c.719T>A​(p.Met240Lys) variant causes a missense change involving the alteration of a non-conserved nucleotide. The variant allele was found at a frequency of 0.000878 in 1,613,904 control chromosomes in the GnomAD database, including 14 homozygotes. In-silico tool predicts a benign outcome for this variant. 13/17 in silico tools predict a benign outcome for this variant. Variant has been reported in ClinVar as Conflicting classifications of pathogenicity (no stars).

• Frequency:
  • Genomes: 𝑓 0.0047 (8 hom., cov: 31)
  • Exomes 𝑓: 0.00048 (6 hom.)
• Consequence: CYP21A2 NM_000500.9 missense
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications P:1 U:1 B:2
• Conservation: PhyloP100: 0.343

Genes affected

CYP21A2 (HGNC:2600): (cytochrome P450 family 21 subfamily A member 2) This gene encodes a member of the cytochrome P450 superfamily of enzymes. The cytochrome P450 proteins are monooxygenases which catalyze many reactions involved in drug metabolism and synthesis of cholesterol, steroids and other lipids. This protein localizes to the endoplasmic reticulum and hydroxylates steroids at the 21 position. Its activity is required for the synthesis of steroid hormones including cortisol and aldosterone. Mutations in this gene cause congenital adrenal hyperplasia. A related pseudogene is located near this gene; gene conversion events involving the functional gene and the pseudogene are thought to account for many cases of steroid 21-hydroxylase deficiency. Two transcript variants encoding different isoforms have been found for this gene. [provided by RefSeq, Jul 2008]

ACMG classification

Verdict is Likely_benign. Variant got -6 ACMG points.

• PM1: In a helix (size 22) in uniprot entity CP21A_HUMAN there are 11 pathogenic changes around while only 1 benign (92%) in NM_000500.9
• BP4: Computational evidence support a benign effect (MetaRNN=0.054983348).
• BS2: High Homozygotes in GnomAd4 at 8 AR gene

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	UniProt
CYP21A2	NM_000500.9	c.719T>A	p.Met240Lys	missense_variant	6/10	ENST00000644719.2
CYP21A2	NM_001128590.4	c.629T>A	p.Met210Lys	missense_variant	5/9
CYP21A2	NM_001368143.2	c.314T>A	p.Met105Lys	missense_variant	6/10
CYP21A2	NM_001368144.2	c.314T>A	p.Met105Lys	missense_variant	5/9

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Appris	UniProt
CYP21A2	ENST00000644719.2	c.719T>A	p.Met240Lys	missense_variant	6/10		NM_000500.9	P1

Frequencies

GnomAD3 genomes AF: 0.00465 AC: 707 AN: 152162 Hom.: 8 Cov.: 31

Gnomad AFR AF: 0.0158

Gnomad AMI AF: 0.00

Gnomad AMR AF: 0.00190

Gnomad ASJ AF: 0.00

Gnomad EAS AF: 0.00

Gnomad SAS AF: 0.000207

Gnomad FIN AF: 0.00

Gnomad MID AF: 0.00

Gnomad NFE AF: 0.000132

Gnomad OTH AF: 0.00574

GnomAD3 exomes AF: 0.000995 AC: 249 AN: 250242 Hom.: 1 AF XY: 0.000680 AC XY: 92 AN XY: 135366

Gnomad AFR exome AF: 0.0128

Gnomad AMR exome AF: 0.000840

Gnomad ASJ exome AF: 0.0000995

Gnomad EAS exome AF: 0.00

Gnomad SAS exome AF: 0.0000654

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.0000796

Gnomad OTH exome AF: 0.000327

GnomAD4 exome AF: 0.000481 AC: 703 AN: 1461624 Hom.: 6 Cov.: 54 AF XY: 0.000413 AC XY: 300 AN XY: 727064

Gnomad4 AFR exome AF: 0.0141

Gnomad4 AMR exome AF: 0.00132

Gnomad4 ASJ exome AF: 0.0000383

Gnomad4 EAS exome AF: 0.000101

Gnomad4 SAS exome AF: 0.000104

Gnomad4 FIN exome AF: 0.0000187

Gnomad4 NFE exome AF: 0.0000818

Gnomad4 OTH exome AF: 0.00108

GnomAD4 genome AF: 0.00469 AC: 714 AN: 152280 Hom.: 8 Cov.: 31 AF XY: 0.00447 AC XY: 333 AN XY: 74472

Gnomad4 AFR AF: 0.0160

Gnomad4 AMR AF: 0.00190

Gnomad4 ASJ AF: 0.00

Gnomad4 EAS AF: 0.00

Gnomad4 SAS AF: 0.000207

Gnomad4 FIN AF: 0.00

Gnomad4 NFE AF: 0.000132

Gnomad4 OTH AF: 0.00568

Alfa AF: 0.000306 Hom.: 0

ESP6500AA AF: 0.0182 AC: 80

ESP6500EA AF: 0.00 AC: 0

ExAC AF: 0.00137 AC: 166

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Pathogenic:1 Uncertain:1 Benign:2

Revision: criteria provided, conflicting classifications

Submissions by phenotype

not provided Pathogenic:1 Uncertain:1

Pathogenic, criteria provided, single submitter	clinical testing	Clinical Genetics and Genomics, Karolinska University Hospital	May 23, 2017	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Invitae	Oct 31, 2022	This sequence change replaces methionine, which is neutral and non-polar, with lysine, which is basic and polar, at codon 240 of the CYP21A2 protein (p.Met240Lys). The frequency data for this variant in the population databases (gnomAD) is considered unreliable due to the presence of homologous sequence, such as pseudogenes or paralogs, in the genome. The c.719T>A (p.Met240Lys) variant alone has not been reported in the literature in individuals with CYP21A2-related conditions. However, it has been reported to co-occur with the c.710T>A (p.Ile237Asn) and c.713T>A (p.Val238Glu) variants in cis (on the same chromosome), which is known as the c.[710T>A;713T>A;719T>A] haplotype, E6 cluster, or CL6. This haplotype has been reported in the literature as homozygous or in combination with other CYP21A2 variants in individual(s) affected with classic salt-wasting, simple virilizing, and non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency (PMID: 12915679, 1644925, 23359698, 26804566). This variant is also known as p.M239K. ClinVar contains an entry for this variant (Variation ID: 242688). 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 CYP21A2 protein function. While the c.719T>A (p.Met240Lys) variant alone does not substantially affect CYP21A2 protein function, the c.[710T>A;713T>A;719T>A] haplotype has been reported to abolish enzyme activity (PMID: 15623806). 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. -

not specified Benign:1

Likely benign, criteria provided, single submitter

clinical testing

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

Dec 07, 2023

Variant summary: CYP21A2 c.719T>A (p.Met240Lys) results in a non-conservative amino acid change in the encoded protein sequence. Three of five in-silico tools predict a benign effect of the variant on protein function. The variant allele was found at a frequency of 0.001 in 250242 control chromosomes, predominantly at a frequency of 0.013 within the African or African-American subpopulation in the gnomAD database, including 1 homozygotes (14 total homozygotes in gnomAD V4). This observation needs to be cautiously considered due to the possibility of the CYP21A1P pseudogene being captured, although the observation of 14 homozygotes decreases the likelihood of a pseudogene artifact. p.M240K is found to be part of a recurrent cluster of 3 variants (I237N, V238E and M240K), which belongs to a group of common, pseudogene-derived mutations that are found in patients with classical CAH. This cluster of 3 variants is assumed to be transferred together from the CYP21A1P pseudogene to CYP21A2 in a continuous stretch of DNA (Robins_2005). To our knowledge, p.M240K has been reported in the literature as part of this cluster of 3 variants in individuals affected with Congenital Adrenal Hyperplasia (e.g. Higashi_1991, Barbat_1995, Chang_2011, Kirac_2014, Essawi_2020, Wang_2021, Turan_2022). These reports do not provide unequivocal conclusions about association of the p.M240K variant alone with Congenital Adrenal Hyperplasia. Experimental evidence evaluating an impact on protein function demonstrated that the p.M240K variant alone had no effect on enzyme activity and consequently does not contribute to the disease (Robins_2005). The following publications have been ascertained in the context of this evaluation (PMID: 7749410, 32616876, 21117955, 32614782, 1869518, 25227725, 15623806, 2249999, 33864926, 31586465). Three submitters have cited clinical-significance assessments for this variant to ClinVar after 2014. One submitter classified the variant as pathogenic, one submitter classified the variant as VUS and one submitter classified the variant as likely benign. Based on the evidence outlined above, the variant was classified as likely benign. -

Classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency Benign:1

Likely benign, criteria provided, single submitter

clinical testing

3billion

May 22, 2022

The variant is observed as homozygous in at least two unrelated individuals/adults in the gnomAD v.2.1.1 dataset, being considered benign, however it is informative in the circumstances for the cluster of variants for pathogenicity. In silico tool predictions suggest no damaging effect of the variant on gene or gene product (REVEL: 0.18; 3Cnet: 0.04). Same nucleotide change resulting in same amino acid change has been previously reported to be associated with CYP21A2 related disorder (ClinVar ID: VCV000242688). Therefore, this variant is classified as pathogenic according to the recommendation of ACMG/AMP guideline. -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Benign	-0.45	T
BayesDel_noAF	Benign	-0.40
CADD	Benign	11
DANN	Benign	0.88
Eigen	Benign	-1.4
Eigen_PC	Benign	-1.3
FATHMM_MKL	Benign	0.0012	N
MetaRNN	Benign	0.055	T;T;T;T
MetaSVM	Benign	-1.1	T
MutationTaster	Benign	0.0000027	A;A
PrimateAI	Benign	0.30	T
PROVEAN	Benign	2.8	N;.;N;.
REVEL	Benign	0.18
Sift	Benign	1.0	T;.;T;.
Sift4G	Benign	1.0	T;.;T;.
Polyphen		0.0	B;B;.;B
Vest4		0.14
MVP		0.76
MPC		0.72
ClinPred		0.0032	T
GERP RS		1.6
gMVP		0.48

Splicing

Name	Calibrated prediction	Score	Prediction
SpliceAI score (max)		0.030		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: rs6476; hg19: chr6-32007593;