NM_000465.4:c.1694G>A

BARD1 · NP_000456.2:p.(Arg565His) · NM_000465.4

GRCh37: chr2:215610562 C>T · GRCh38: chr2:214745838 C>T

Gene: BARD1 Transcript: NM_000465.4

Final call

Likely Benign

BS2 supporting benign BS3 strong benign BP4 supporting benign

Variant details

Gene

BARD1

Transcript

NM_000465.4

Protein

NP_000456.2:p.(Arg565His)

gnomAD AF

0.00021065074973080074 (v4.1)

ClinVar

Likely benign

OncoKB

Unknown Oncogenic Effect

Classification rationale

Interpretation summary

Generated evidence synthesis

BARD1 c.1694G>A (p.Arg565His) is a missense variant in exon 8 located in the inter-domain region between the ankyrin repeat and BRCT domains.

Direct functional testing in a validated homology-directed repair (HDR) assay demonstrated that BARD1 R565H is fully functional, with HDR activity comparable to wild-type. The variant was explicitly classified as 'functional in HDR' alongside known benign variants (PMID:30925164).

This variant is present in gnomAD v2.1 at 105/282,714 alleles (AF=0.037%) and in gnomAD v4.1 at 340/1,614,046 alleles (AF=0.021%), with 3 homozygous individuals observed in v4.1. Homozygosity in a general population database is inconsistent with a highly penetrant pathogenic variant in a tumor suppressor gene.

Multiple in silico tools support a benign effect: REVEL score 0.179, BayesDel score -0.24551, and SpliceAI max delta 0.01 (no splicing impact).

ClinVar reports this variant as Likely benign based on submissions from 10 clinical laboratories, with an additional 2 laboratories reporting Benign and 4 reporting Uncertain significance (Variation ID 127721).

The variant was observed in 1 of 354 sporadic breast cancer cases and 0 of 258 controls (PMID:17972171), which does not establish significant case enrichment. A single FCCTX colorectal cancer case carrying this variant has also been reported, but colorectal cancer is not within the primary BARD1-associated disease spectrum (PMID:32984025).

No evidence of de novo occurrence, co-segregation with disease, or location within a critical functional domain or mutational hotspot was identified. Residue 565 lies in a linker region where variants tested in functional studies were generally proficient in DNA repair.

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	This is a missense variant (c.1694G>A, p.Arg565His). PVS1 applies only to null variants (nonsense, frameshift, canonical ±1,2 splice sites). The variant does not fall into any of the default generic PVS1 null-variant buckets per ClinGen SVI PVS1 recommendations (PMC6185798).	pvs1_generic_framework
PS1	Not met	No evidence of a different nucleotide change at codon 565 resulting in the same amino acid substitution (p.Arg565His) that has been classified as pathogenic. PM5 candidate harvesting found no eligible comparators at this residue.	pm5_candidates
PS2	Not met	No de novo observation with confirmed maternity and paternity has been reported for this variant in any reviewed publication or database.
PS3	Not met	Direct functional testing of BARD1 R565H in a validated homology-directed repair (HDR) assay demonstrated that the variant is functional and proficient in DNA repair, comparable to wild-type. The variant was explicitly classified among 'functional in HDR' variants alongside known benign variants (PMID:30925164). This functional evidence contradicts a damaging effect and instead supports a benign interpretation.	PMID:30925164
PS4	Not met	Observed in 1 of 354 sporadic breast cancer cases and 0 of 258 controls (PMID:17972171), which does not establish statistically significant enrichment. The variant is present in gnomAD at 105/282,714 alleles (v2.1, AF=0.037%) and 340/1,614,046 alleles (v4.1, AF=0.021%), indicating it is established in the general population at frequencies incompatible with a highly penetrant pathogenic variant. No case-control study demonstrates significant enrichment in affected individuals.	gnomad_v2 gnomad_v4 PMID:17972171
PS5	Not met	No reputable source classifies this variant as pathogenic. ClinVar consensus classification is Likely benign (10 clinical laboratories reporting Likely benign, 2 reporting Benign). No expert panel has reviewed this variant.	clinvar
PM1	Not met	Residue Arg565 lies in the inter-domain region between the ankyrin repeat domain (residues 427-525) and the first BRCT domain (residues 616-653). It is not located within a well-characterized critical functional domain. Cancerhotspots.org does not identify this residue as a statistically significant mutational hotspot. Functional data (PMID:30925164) shows variants in this inter-domain region are proficient in DNA repair, further arguing against a critical domain assignment.	PMID:30925164 PMID:23056176
PM2	Not met	Although total allele frequency in gnomAD v2.1 is 0.037% (below 0.1% PM2 threshold), the variant is present at 105 alleles in v2.1 and 340 alleles in v4.1 with 3 homozygous individuals observed in v4.1. The presence of homozygotes in a general population database and occurrence across multiple continental populations indicates this variant is established in the population and does not meet the 'absent from controls' standard of PM2.	gnomad_v2 gnomad_v4
PM5	Not met	No known pathogenic missense variant at codon 565 with a different amino acid change has been established. R565C was tested in the HDR functional assay (PMID:30925164) and showed borderline activity just below the 0.6 cutoff, but it is not classified as pathogenic in ClinVar and does not satisfy the PM5 requirement for an established pathogenic comparator at the same residue.	pm5_candidates PMID:30925164
PM6	Not met	No de novo observation (maternity and paternity unconfirmed) has been reported for this variant in any reviewed source.
PP1	Not met	No co-segregation data are available. The variant was reported in a single sporadic breast cancer case (PMID:17972171) without family studies, and in a single FCCTX colorectal cancer case (PMID:32984025) without segregation analysis. The Gorringe et al. study specifically tested segregation of other BARD1 variants but did not analyze R565H for segregation.	PMID:17972171 PMID:32984025
PP2	Not met	BARD1 has a high rate of benign missense variation. Multiple common missense polymorphisms have been documented including P24S, M507V, C557S, and I738V (PMID:17972171). The gene does not exhibit a low rate of benign missense variation, and pathogenic missense variants are also well-documented (PMID:30925164), indicating missense changes are not inherently pathogenic in this gene.	PMID:17972171 PMID:30925164
PP3	Not met	Multiple in silico tools predict a benign effect. REVEL score is 0.179 (well below the typical ≥0.5 damaging threshold). BayesDel score is -0.24551 (negative, supporting a benign interpretation). SpliceAI delta score is 0.01 (no predicted splicing impact). Although PolyPhen predicted damaging in an earlier in silico screen (PMID:23056176), the preponderance of evidence from validated ensemble predictors (REVEL, BayesDel) supports a benign computational profile.	revel bayesdel spliceai PMID:23056176
PP4	Not met	The variant has been observed in a colorectal cancer case (FCCTX, PMID:32984025) and a sporadic breast cancer case (PMID:17972171). The colorectal cancer phenotype is not within the primary BARD1-associated disease spectrum (hereditary breast and ovarian cancer). No specific phenotypic data or family history highly specific for BARD1-related disease is available for either proband.	PMID:17972171 PMID:32984025
PP5	Not met	ClinVar review status for this variant is 'criteria provided, single submitter' (1-star). Per adjudication rules, PP5 applies at supporting strength only for 3-star expert panel submissions. No expert panel (CSPEC/VCEP or ClinGen) has reviewed this variant. The overall ClinVar classification is Likely benign, which would support BP6 rather than PP5.	clinvar
BA1	Not met	Allele frequency in gnomAD v2.1 is 0.037% (total) with maximum subpopulation frequency 0.22% (South Asian). In gnomAD v4.1, total AF is 0.021% with max subpopulation AF 0.17% (South Asian). All frequencies are far below the 1% BA1 threshold.	gnomad_v2 gnomad_v4
BS1	Not met	Allele frequency in gnomAD v2.1 is 0.037% (total), which is below the 0.3% BS1 threshold (non-VCEP). The maximum subpopulation frequency in South Asians is 0.22%, also below the threshold. Grpmax FAF in v2.1 is 0.18%, also below 0.3%.	gnomad_v2 gnomad_v4
BS2	Met	Observed in the homozygous state in 3 individuals in gnomAD v4.1, a general population database. BARD1 is a tumor suppressor gene where biallelic loss-of-function would be expected to have significant phenotypic consequences. The observation of homozygosity in presumably healthy population controls suggests the variant is tolerated even in the homozygous state.	gnomad_v4
BS3	Met	Direct functional testing of BARD1 R565H in a validated homology-directed repair (HDR) assay demonstrated unequivocally normal function. Adamovich et al. (PMID:30925164) tested 76 BARD1 missense variants in a well-characterized HDR assay that accurately discriminates known pathogenic from known benign BARD1 variants. R565H was proficient in HDR with activity comparable to wild-type and was explicitly grouped with other functional variants that were interpreted as 'likely benign.' The HDR assay has been validated as predictive of BRCA1/BARD1 pathogenicity, with truncating variants and known pathogenic missense variants showing HDR deficiency while known benign variants remain functional.	PMID:30925164
BS4	Not met	No co-segregation data are available for this variant. While the Gorringe et al. study (PMID:17972171) tested segregation for other BARD1 variants (T598I, I692T) and found no segregation with disease, R565H was identified in a single sporadic case and was not included in segregation analysis.	PMID:17972171
BP1	N/A	BP1 applies when a missense variant occurs in a gene for which primarily truncating variants are known to cause disease. In BARD1, both truncating and missense variants are established mechanisms of disease. Multiple missense variants in BARD1 have been demonstrated to be HDR-deficient and are classified as pathogenic (PMID:30925164). Therefore, a missense change in BARD1 cannot be assumed benign solely on the basis of variant type.	PMID:30925164
BP2	Not met	No data are available regarding observation of this variant in trans with a known pathogenic variant in BARD1.
BP4	Met	Multiple lines of computational evidence support a benign effect. REVEL score is 0.179 (well below damaging thresholds). BayesDel score is -0.24551 (negative, supporting benign). SpliceAI max delta score is 0.01 (no predicted splicing impact). The preponderance of in silico evidence from validated ensemble predictors supports a benign interpretation, consistent with the functional data showing normal HDR activity.	revel bayesdel spliceai
BP5	Not met	No data are available demonstrating that a case carrying this variant has an alternate molecular basis for disease. The variant was observed in an FCCTX patient (PMID:32984025) who also carried BRCA2 variants, but these were classified as VUS and do not constitute an alternate definitive molecular diagnosis.	PMID:32984025
BP6	Not met	Although ClinVar reports this variant as Likely benign with strong clinical laboratory consensus (10 labs reporting Likely benign, 2 reporting Benign, 4 reporting VUS), the review status is 'criteria provided, single submitter' (1-star). Per adjudication rules, BP6 applies at supporting strength only for 3-star expert panel submissions. No ClinGen or VCEP expert panel has reviewed this variant.	clinvar
BP7	N/A	This is a missense variant (c.1694G>A, p.Arg565His), not a synonymous or intronic variant. BP7 applies only to synonymous variants with no predicted splice impact.
BP3	N/A	BP3 applies to in-frame insertions/deletions in repetitive regions. This is a single-nucleotide substitution.
PM3	N/A	PM3 applies to recessive disorders where the variant is observed in trans with a pathogenic variant. BARD1-associated disease is autosomal dominant; PM3 is not applicable in this context.
PM4	N/A	PM4 applies to protein length changes (in-frame deletions/insertions, stop-loss). This is a single-nucleotide missense substitution.

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