16-89738943-C-T

Position:

• chr16-89738943-C-T

Variant summary

Our verdict is Likely pathogenic. Variant got 8 ACMG points: 8P and 0B. PS1_Moderate PM2 PM5 PP3 PP5

The NM_000135.4(FANCA):​c.4199G>A​(p.Arg1400His) variant causes a missense change involving the alteration of a non-conserved nucleotide. The variant allele was found at a frequency of 0.0000688 in 1,614,272 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 nucleotide change resulting in same amino acid change has been previously reported as Likely pathogenicin UniProt. Another variant affecting the same amino acid position, but resulting in a different missense (i.e. R1400C) has been classified as Pathogenic.

• Frequency:
  • Genomes: 𝑓 0.000033 (0 hom., cov: 33)
  • Exomes 𝑓: 0.000073 (0 hom.)
• Consequence: FANCA NM_000135.4 missense
• Clinical Significance: Conflicting classifications of pathogenicity criteria provided, conflicting classifications P:6 U:3
• Conservation: PhyloP100: 1.10

Genes affected

FANCA (HGNC:3582): (FA complementation group A) The Fanconi anemia complementation group (FANC) currently includes FANCA, FANCB, FANCC, FANCD1 (also called BRCA2), FANCD2, FANCE, FANCF, FANCG, FANCI, FANCJ (also called BRIP1), FANCL, FANCM and FANCN (also called PALB2). The previously defined group FANCH is the same as FANCA. Fanconi anemia is a genetically heterogeneous recessive disorder characterized by cytogenetic instability, hypersensitivity to DNA crosslinking agents, increased chromosomal breakage, and defective DNA repair. The members of the Fanconi anemia complementation group do not share sequence similarity; they are related by their assembly into a common nuclear protein complex. This gene encodes the protein for complementation group A. Alternative splicing results in multiple transcript variants encoding different isoforms. Mutations in this gene are the most common cause of Fanconi anemia. [provided by RefSeq, Jul 2008]

ZNF276 (HGNC:23330): (zinc finger protein 276) Enables sequence-specific double-stranded DNA binding activity. Predicted to be involved in regulation of transcription by RNA polymerase II. Located in kinetochore. [provided by Alliance of Genome Resources, Apr 2022]

FANCA (HGNC:):

ACMG classification

Verdict is Likely_pathogenic. Variant got 8 ACMG points.

• PS1: Transcript NM_000135.4 (FANCA) is affected with MISSENSE_VARIANT having same AA change as one Pathogenic present in UniProt
• PM2: Very rare variant in population databases, with high coverage
• PM5: Other missense variant is known to change same aminoacid residue: Variant chr16-89738944-G-A is described in Lovd as [Pathogenic].
• PP3: MetaRNN computational evidence supports a deleterious effect, 0.831
• PP5: Variant 16-89738943-C-T is Pathogenic according to our data. Variant chr16-89738943-C-T is described in ClinVar as [Conflicting_classifications_of_pathogenicity]. Clinvar id is 408175.We mark this variant Likely_pathogenic, oryginal submissions are: {Uncertain_significance=2, Pathogenic=1, Likely_pathogenic=3}. Variant chr16-89738943-C-T is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
FANCA	NM_000135.4	c.4199G>A	p.Arg1400His	missense_variant	42/43	ENST00000389301.8	NP_000126.2
ZNF276	NM_001113525.2	c.*697C>T		3_prime_UTR_variant	11/11	ENST00000443381.7	NP_001106997.1

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
FANCA	ENST00000389301.8	c.4199G>A	p.Arg1400His	missense_variant	42/43	1	NM_000135.4	ENSP00000373952.3
ZNF276	ENST00000443381.7	c.*697C>T		3_prime_UTR_variant	11/11	1	NM_001113525.2	ENSP00000415836.2

Frequencies

GnomAD3 genomes AF: 0.0000328 AC: 5 AN: 152270 Hom.: 0 Cov.: 33

Gnomad AFR AF: 0.0000482

Gnomad AMI AF: 0.00

Gnomad AMR AF: 0.0000654

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: 0.0000294

Gnomad OTH AF: 0.00

GnomAD3 exomes AF: 0.0000358 AC: 9 AN: 251486 Hom.: 0 AF XY: 0.0000368 AC XY: 5 AN XY: 135920

Gnomad AFR exome AF: 0.00

Gnomad AMR exome AF: 0.0000867

Gnomad ASJ exome AF: 0.00

Gnomad EAS exome AF: 0.0000544

Gnomad SAS exome AF: 0.0000327

Gnomad FIN exome AF: 0.00

Gnomad NFE exome AF: 0.0000352

Gnomad OTH exome AF: 0.00

GnomAD4 exome AF: 0.0000725 AC: 106 AN: 1461884 Hom.: 0 Cov.: 35 AF XY: 0.0000811 AC XY: 59 AN XY: 727246

Gnomad4 AFR exome AF: 0.00

Gnomad4 AMR exome AF: 0.0000894

Gnomad4 ASJ exome AF: 0.00

Gnomad4 EAS exome AF: 0.0000504

Gnomad4 SAS exome AF: 0.0000348

Gnomad4 FIN exome AF: 0.00

Gnomad4 NFE exome AF: 0.0000836

Gnomad4 OTH exome AF: 0.0000662

GnomAD4 genome AF: 0.0000328 AC: 5 AN: 152388 Hom.: 0 Cov.: 33 AF XY: 0.0000403 AC XY: 3 AN XY: 74512

Gnomad4 AFR AF: 0.0000481

Gnomad4 AMR AF: 0.0000653

Gnomad4 ASJ AF: 0.00

Gnomad4 EAS AF: 0.00

Gnomad4 SAS AF: 0.00

Gnomad4 FIN AF: 0.00

Gnomad4 NFE AF: 0.0000294

Gnomad4 OTH AF: 0.00

Alfa AF: 0.0000563 Hom.: 0

Bravo AF: 0.0000453

TwinsUK AF: 0.000270 AC: 1

ALSPAC AF: 0.000259 AC: 1

ESP6500AA AF: 0.00 AC: 0

ESP6500EA AF: 0.000233 AC: 2

ExAC AF: 0.0000412 AC: 5

EpiCase AF: 0.000109

EpiControl AF: 0.000119

ClinVar

Significance: Conflicting classifications of pathogenicity

Submissions summary: Pathogenic:6 Uncertain:3

Revision: criteria provided, conflicting classifications

Submissions by phenotype

Fanconi anemia complementation group A Pathogenic:3 Uncertain:3

Likely pathogenic, criteria provided, single submitter	clinical testing	Fulgent Genetics, Fulgent Genetics	Jun 17, 2024	- -
Uncertain significance, no assertion criteria provided	clinical testing	Natera, Inc.	Sep 16, 2020	- -
Uncertain significance, criteria provided, single submitter	clinical testing	Illumina Laboratory Services, Illumina	Feb 06, 2018	The FANCA c.4199G>A (p.Arg1400His) variant has been reported in two studies and is found in two individuals with Fanconi anemia, including one in a compound heterozygous state with a missense variant and one in a heterozygous state in whom a second variant was not identified (Ameziane et al. 2008; De Rocco et al. 2014). Control data are unavailable for this variant, which is reported at a frequency of 0.000045 in the European (non-Finnish) population of the Exome Aggregation Consortium. Expression of the p.Arg1400His variant in lymphoblastoid cell lines lacking functional FANCA protein demonstrated an activity similar to wild type protein, however, compensation due to overexpression of the p.Arg1400His protein could not be ruled out (Ameziane et al. 2008). The evidence for this variant is limited. The variant is therefore classified as a variant of unknown significance but suspicious for pathogenicity for Fanconi anemia. This variant was observed by ICSL as part of a predisposition screen in an ostensibly healthy population. -
Uncertain significance, criteria provided, single submitter	clinical testing	Myriad Genetics, Inc.	Nov 11, 2021	NM_000135.2(FANCA):c.4199G>A(R1400H) is a missense variant classified as a variant of uncertain significance in the context of Fanconi anemia complementation group A. R1400H has been observed in cases with relevant disease (PMID: 17924555, 29098742, 33172906). Functional assessments of this variant are available in the literature (PMID: 17924555, 33172906). R1400H has been observed in population frequency databases (gnomAD: AMR 0.01%). In summary, there is insufficient evidence to classify NM_000135.2(FANCA):c.4199G>A(R1400H) as pathogenic or benign. Please note: this variant was assessed in the context of healthy population screening. -
Likely pathogenic, no assertion criteria provided	in vitro	International Fanconi Anemia Registry, The Rockefeller University	Aug 05, 2020	The c.4199G>A/p.R1400H variant was previously reported in an individual with Fanconi anemia, but determining pathogenicity was difficult because it was reported that overexpression of the mutant allele resulted in complementation comparable to that obtained with the wild type cDNA (Ameziane et al. 2008). This suggests that overexpression of the mutant protein is able to compensate for protein instability and/or defects in FANCA protein function. We have identified c.4199G>A/p.R1400H variant in another FA family (two affected brothers) in trans with a c.3788_3790delTCT, a known pathogenic variant. Our functional data support the variant as a most likely pathogenic allele. We observe increased mislocalization of mutant protein to the cytoplasm, decreased FANCD2 ubiquitination and foci formation, and cellular sensitivity to crosslinking agents. We have examined the variant by CRISPR/Cas9 mediated genome editing in otherwise wild type fibroblast cell lines and we showed that the R1400H variant in homozygous form had an incomplete loss of function (behaved as a hypomorph), and the cellular phenotype characteristic of Fanconi anemia was further exacerbated when it was present in trans to a loss-of-function allele, indicating a dose effect of a mutant protein. -
Likely pathogenic, criteria provided, single submitter	clinical testing	Baylor Genetics	Mar 24, 2024	- -

Fanconi anemia Pathogenic:2

Likely pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Dec 13, 2023	Variant summary: FANCA c.4199G>A (p.Arg1400His) results in a non-conservative amino acid change in the encoded protein sequence. Five of five in-silico tools predict a damaging effect of the variant on protein function. The variant allele was found at a frequency of 3.6e-05 in 251486 control chromosomes (gnomAD). c.4199G>A has been reported in the literature in multiple compound heterozygous individuals diagnosed with Fanconi Anemia, where the diagnosis had been established by chromosomal breakage test (Ameziane_2008, Kimble_2018, Lach_2020), however 2 of these cases (brothers) had an atypical, delayed phenotype (i.e. presenting with esophageal malignancies at the age of 51y, with the lack of hematologic failure), and were subsequently diagnosed to have FA by chromosomal breakage and molecular testing (Lach_2020). Publications have also reported experimental evidence evaluating an impact on protein function. In an early study, expression of the R1400H variant in lymphoblastoid cell lines lacking functional FANCA protein demonstrated similar activity to the wild type, however due to overexpression of the construct, a potential hypomorphic effect couldn't be ruled out (Ameziane_2008). A later, more detailed study demonstrated a reduced protein level, and increased cytoplasmic localization, together with a mild decrease in Fancd2 ubiquitination and foci formation, and increased sensitivity to DNA cross-linking agents, providing evidence for a hypomorphic effect (Lach_2020). The following publications have been ascertained in the context of this evaluation (PMID: 17924555, 29098742, 33172906, 28717661, 33686268, 26580448). Six submitters have cited clinical-significance assessments for this variant to ClinVar after 2014 with conflicting assessments. Three submitters classified the variant as pathogenic/likely pathogenic and three classified it as uncertain significance. Based on the evidence outlined above, the variant was classified as likely pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Labcorp Genetics (formerly Invitae), Labcorp	Dec 30, 2023	This sequence change replaces arginine, which is basic and polar, with histidine, which is basic and polar, at codon 1400 of the FANCA protein (p.Arg1400His). This variant is present in population databases (rs149851163, gnomAD 0.006%). This missense change has been observed in individuals with Fanconi anemia (PMID: 17924555, 29098742). ClinVar contains an entry for this variant (Variation ID: 408175). 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 expected to disrupt FANCA protein function with a positive predictive value of 80%. This variant disrupts the p.Arg1400 amino acid residue in FANCA. Other variant(s) that disrupt this residue have been determined to be pathogenic (PMID: 15643609, 21273304, 24584348, 28102861, 28717661, 29098742). This suggests that this residue is clinically significant, and that variants that disrupt this residue are likely to be disease-causing. For these reasons, this variant has been classified as Pathogenic. -

not provided Pathogenic:1

Likely pathogenic, criteria provided, single submitter

clinical testing

GeneDx

Aug 10, 2022

Observed with a second FANCA variant in unrelated patients with Fanconi anemia in published literature, but it is not known whether the variants occurred on the same (in cis) or on different (in trans) chromosomes (Ameziane 2008, Kimble 2018); Published functional studies demonstrate decreased FANCA levels and impaired ubiquitination and foci formation upon MMC-induced damage; however, functional complementation assays show that overexpresson of this variant exhibited a level of activity similar to WT, suggesting that this may be a hypomorphic variant (Ameziane 2008, Lach 2020); In silico analysis supports that this missense variant has a deleterious effect on protein structure/function; This variant is associated with the following publications: (PMID: 29098742, 33172906, 17924555, 33822308) -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
AlphaMissense	Benign	0.18
BayesDel_addAF	Pathogenic	0.26	D
BayesDel_noAF	Pathogenic	0.32
CADD	Uncertain	25
DANN	Pathogenic	1.0
DEOGEN2	Uncertain	0.67	D
Eigen	Uncertain	0.34
Eigen_PC	Benign	0.21
FATHMM_MKL	Uncertain	0.83	D
LIST_S2	Uncertain	0.88	D
M_CAP	Uncertain	0.17	D
MetaRNN	Pathogenic	0.83	D
MetaSVM	Uncertain	0.65	D
MutationAssessor	Uncertain	2.7	M
PrimateAI	Benign	0.35	T
PROVEAN	Uncertain	-3.0	D
REVEL	Pathogenic	0.66
Sift	Uncertain	0.0020	D
Sift4G	Uncertain	0.0040	D
Polyphen		1.0	D
Vest4		0.78
MVP		0.97
ClinPred		0.60	D
GERP RS		5.4
Varity_R		0.39
gMVP		0.46

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: rs149851163; hg19: chr16-89805351;