NM_000500.9:c.1439

CYP21A2 · ·

GRCh37: None · GRCh38: None

Gene: CYP21A2 Transcript:

Final call

Likely Benign

BS1 supporting BS3 supporting BP6 supporting

Variant details

Gene

CYP21A2

Transcript

Protein

gnomAD AF

ClinVar

Benign

OncoKB

Classification rationale

Interpretation summary

Generated evidence synthesis

Functional studies in transiently transfected mammalian cells demonstrate the p.Arg480Leu (R479L) mutant retains 75.5% activity toward 17-hydroxyprogesterone and 79.6% activity toward progesterone compared to wild-type, with normal substrate binding kinetics.

The variant has been observed in a healthy population cohort at a frequency exceeding the benign threshold for a recessive disorder, with 1 heterozygote identified among 92 healthy Hungarian blood donors (1.09%).

Multiple independent clinical diagnostic laboratories have classified this variant as Benign (three laboratories) or Likely benign (three laboratories) in ClinVar (variation ID 445854).

The variant is also present in affected populations at a comparable low frequency (2/250 alleles, 0.8%, in a Sicilian CAH cohort), consistent with a benign polymorphism rather than a disease-causing allele.

No pathogenic or likely pathogenic classifications for this variant have been reported by any clinical laboratory or expert panel.

Final determination: Generic ACMG/AMP 2015 fallback rules support a Likely Benign classification because either one strong benign criterion plus one supporting benign criterion, or at least two supporting benign criteria, are present.

Criteria assessment

ACMG/AMP criteria review

Criteria shown when status is available

All criteria require review: For research and educational purposes only.

Criterion	Status	Rationale	Evidence used
PVS1	N/A	PVS1 is not applicable to missense variants. NM_000500.9:c.1439G>T is a missense change (p.Arg480Leu) and does not fall into the null-variant buckets of nonsense, frameshift, or canonical ±1,2 splice consensus variants per ClinGen SVI PVS1 recommendations (PMC6185798).	pvs1_generic_framework
PS1	Not assessed	PS1 requires a different nucleotide change at the same codon producing the same amino acid change (p.Arg480Leu). Only c.1439G>T produces this change at codon 480; no alternative nucleotide substitution yielding p.Arg480Leu has been identified.
PS2	Not met	No evidence of a de novo occurrence of NM_000500.9:c.1439G>T with confirmed maternity and paternity has been reported. CYP21A2 mutations are typically inherited in an autosomal recessive pattern, and de novo occurrences are rare.
PS3	Not met	Functional studies by Robins et al. 2007 (PMID:17119906) demonstrated that the R479L (p.Arg480Leu) mutant retains 75.5% activity toward 17-hydroxyprogesterone and 79.6% activity toward progesterone relative to wild-type, with normal substrate binding kinetics. This near-normal enzyme activity does not support a damaging effect and therefore does not meet PS3.	PMID:17119906
PS4	Not met	The variant has been observed in both affected and healthy populations. It was detected in 2 of 250 CYP21A2 alleles (0.8%) in a Sicilian CAH cohort (PMID:21169732) and in 1 of 92 healthy Hungarian blood donors (PMID:19505723). The similar prevalence in affected and general populations does not support statistically significant enrichment in cases versus controls.	PMID:21169732 PMID:19505723
PS5	Not assessed	PS5 requires PM2_Supporting as a prerequisite per generic ACMG/AMP 2015 framework. PM2 is not met (variant observed in healthy population, gnomAD data unavailable), so PS5 is not applicable.
PM1	Not met	PM1 applies to variants located in a mutational hot spot or well-established critical functional domain without benign variation. Structural modeling by Robins et al. 2006 (PMID:16788163) places residue Arg479 (Arg480 in NM numbering) in a surface-exposed loop region, not in the heme-binding pocket, steroid-binding site, or redox partner interaction interface. This residue is not part of a recognized critical functional domain without benign variation.	PMID:16788163
PM2	Not met	PM2 requires the variant to be absent from or at extremely low frequency (<0.1%) in population databases. The variant was observed in 1 of 92 healthy Hungarian individuals (1.09%) by Blasko et al. 2009 (PMID:19505723). gnomAD v2.1 and v4.1 data are unavailable for this variant. The observation of the variant in a healthy population at >0.1% precludes application of PM2.	PMID:19505723
PM5	N/A	PM5 requires a different missense change at the same amino acid residue that has been established as pathogenic. No same-residue comparator variants with established pathogenicity could be identified for position Arg480 (PM5 candidate harvesting returned no results).
PM6	Not met	No evidence of a de novo occurrence of NM_000500.9:c.1439G>T without confirmation of maternity and paternity has been identified in the reviewed literature.
PP1	Not met	No co-segregation data for NM_000500.9:c.1439G>T in multiple affected family members has been reported. The R479L carrier in Robins et al. 2007 (PMID:17119906) was heterozygous with wild-type on the other allele and lacked affected family members for segregation analysis.
PP2	Not met	PP2 requires the gene to have a low rate of benign missense variation with missense variants as a common disease mechanism. CYP21A2 has multiple well-characterized benign missense variants (e.g., p.Leu9del, p.Arg102Lys, p.Asn493Ser) occurring at appreciable population frequencies, indicating a high rate of benign missense variation. Therefore PP2 is not met.	PMID:19505723
PP3	Not met	Multiple lines of computational evidence are required for PP3. REVEL, BayesDel, and SpliceAI scores are unavailable for this variant (mutalyzer normalization did not produce the required genomic coordinates). Without in silico predictions, PP3 cannot be assessed as met.
PP4	Not met	PP4 requires the patient's phenotype or family history to be highly specific for the disease. No patient-specific clinical data for NM_000500.9:c.1439G>T carriers with highly specific phenotype was available for adjudication.
PP5	Not met	PP5 requires a reputable source to have recently reported the variant as pathogenic. In ClinVar (ID 445854), the variant is classified as Benign by 3 clinical laboratories, Likely benign by 3 clinical laboratories, and Uncertain significance by 2 clinical laboratories. No clinical laboratory or expert panel has classified this variant as pathogenic.	clinvar
BA1	Not met	BA1 requires allele frequency >1% in a general population database. gnomAD v2.1 and v4.1 data are unavailable. The variant was observed at 1.09% (1/92) in a small healthy Hungarian cohort (PMID:19505723), but this single small study does not meet the rigorous population database standard required for BA1.	PMID:19505723
BS1	Met	BS1 applies when allele frequency is >0.3% in a general population database for a recessive disorder. The variant was observed in 1 of 92 healthy Hungarian blood donors (1.09%) by Blasko et al. 2009 (PMID:19505723), a healthy population cohort. Although gnomAD data are unavailable, this observation in an unselected healthy population at >0.3% supports benign classification. Note: sample size is limited (n=92), warranting caution.	PMID:19505723
BS2	Not met	BS2 requires observation of the variant in a healthy adult individual when full penetrance is expected at an early age, either in trans with a known pathogenic variant (for recessive disorders) or in the homozygous state. No such observation has been reported for NM_000500.9:c.1439G>T.
BS3	Met	Well-established in vitro functional studies by Robins et al. 2007 (PMID:17119906) demonstrate that the R479L (p.Arg480Leu) mutant retains 75.5% activity toward 17-hydroxyprogesterone and 79.6% activity toward progesterone compared to wild-type enzyme, with normal substrate binding kinetics (Km and Vmax not significantly different). Protein expression was confirmed by Western blot to be comparable to wild-type. This near-normal enzymatic activity in a well-validated assay using both natural substrates indicates no damaging effect on protein function, supporting a benign interpretation.	PMID:17119906
BS4	Not met	BS4 requires lack of segregation of the variant with disease in affected family members. No segregation data for NM_000500.9:c.1439G>T in affected families are available from the reviewed literature.
BP1	N/A	BP1 applies to missense variants in genes where truncating variants are the primary known cause of disease. CYP21A2 has numerous well-characterized pathogenic missense variants that are a primary disease mechanism in congenital adrenal hyperplasia; therefore BP1 is not applicable.
BP2	Not met	BP2 requires observation in trans with a pathogenic variant in a healthy individual (for recessive disorders) or observation in cis with a pathogenic variant. No such observations have been reported for NM_000500.9:c.1439G>T.
BP4	Not met	BP4 requires multiple lines of computational evidence suggesting no impact on the gene or gene product. REVEL, BayesDel, and SpliceAI scores are unavailable for this variant due to failed genomic coordinate normalization. Without in silico predictions, BP4 cannot be assessed as met.
BP5	Not met	BP5 requires the variant to be found in a case with an alternate molecular basis for disease. No such case has been identified in the reviewed literature for NM_000500.9:c.1439G>T.
BP6	Met	BP6 applies when a reputable source (e.g., clinical diagnostic laboratory) has recently reported the variant as benign. In ClinVar (variation ID 445854), NM_000500.9:c.1439G>T has been classified as Benign by three clinical diagnostic laboratories (Center for Pediatric Genomic Medicine, Mendelics, GeneDx) and as Likely benign by three additional clinical laboratories (Quest Diagnostics, Athena Diagnostics, and one other submitter). Multiple independent clinical laboratories have reached a benign or likely benign conclusion, supporting BP6 at the supporting level.	clinvar
BP7	N/A	BP7 applies to synonymous (silent) variants for which splicing prediction algorithms predict no impact on the splice consensus sequence or the creation of a new splice site. NM_000500.9:c.1439G>T is a missense variant (p.Arg480Leu), not a synonymous variant. BP7 is not applicable.
BP3	N/A	BP3 applies to in-frame deletions/insertions in repetitive regions without known function. This variant is a single-nucleotide missense change, not an in-frame indel.
PM3	N/A	Skipped per adjudication instructions. PM3 was listed under trivially not_applicable criteria.
PM4	N/A	Skipped per adjudication instructions. PM4 was listed under trivially not_applicable criteria.

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