ClinVar Genomic variation as it relates to human health

The NM_000133.3:c.835G>A variant that results in the Ala279Thr missense change is absent from gnomAD v2.1.1 and v3, meeting PM2_Supporting. It is located within the peptidase S2 domain, which is deemed critical to protein function, meeting PM1. More than 59 male patients with mild hemophilia B are reported hemizygous for this variant in the literature, meeting PS4_Very strong and PP4_Moderate criteria (PMID: 29296726, 29656491). It is noted as a founder variant but may also have arisen de novo as a recurrent variant and is found in multiple ethnicities. The variant has a REVEL score of 0.765 (>0.6) and CADD score of 23 (>21) which meet the thresholds recommended for PP3. In summary, the variant meets criteria to be classified as pathogenic. ACMG/AMP criteria applied, as specified by the Coagulation Factor Deficiency Variant Curation Expert Panel for F9: PS4_Very Strong, PM1, PP3, PP4_Moderate, PM2_Supporting.

The F9 c.835G>A; p.Ala279Thr variant (rs137852247), also known as p.Ala233Thr, is reported in the literature in numerous individuals affected with hemophilia B and is primarily associated with mild disease (Chavali 2009, Chen 1991, Hamasaki-Katagiri 2012, Kihlberg 2017, Factor IX database and references therein). This is a recurrent missense variant observed in many affected individuals due both to a founder effect and to its location in a CpG dinucleotide, which are prone to G-to-A or C-to-T transitions (Chen 1991, Lassalle 2018). Clotting activity assays indicate the p.Ala279Thr variant exhibits approximately 10-15% of wildtype activity, consistent with mild hemophilia (Chavali 2009, Chen 1991, Kihlberg 2017, Factor IX database). This variant is reported in ClinVar (Variation ID: 216926) and is absent from general population databases (Exome Variant Server, Genome Aggregation Database), indicating it is not a common polymorphism. The alanine at codon 279 is highly conserved and occurs in the catalytic serine protease domain, and computational analyses predict that this variant is deleterious (REVEL: 0.765). Additionally, another variant at this codon (c.835G>T, p.Ala279Ser) has reported in individuals with hemophilia B (Rydz 2013), suggesting this amino acid is functionally important. Based on available information, the p.Ala279Thr variant is considered to be pathogenic. References: Factor IX database: http://www.factorix.org Chavali S et al. Hemophilia B is a quasi-quantitative condition with certain mutations showing phenotypic plasticity. Genomics. 2009 Dec;94(6):433-7. Chen SH et al. CG dinucleotide transitions in the factor IX gene account for about half of the point mutations in hemophilia B patients: a Seattle series. Hum Genet. 1991 Jun;87(2):177-82. Hamasaki-Katagiri N et al. Analysis of F9 point mutations and their correlation to severity of haemophilia B disease. Haemophilia. 2012 Nov;18(6):933-40. Kihlberg K et al. Discrepancies between the one-stage clotting assay and the chromogenic assay in haemophilia B. Haemophilia. 2017 Jul;23(4):620-627. Lassalle F et al. Recurrent F8 and F9 gene variants result from a founder effect in two large French haemophilia cohorts. Haemophilia. 2018 Apr 14. Rydz N et al. The Canadian "National Program for hemophilia mutation testing" database: a ten-year review. Am J Hematol. 2013 Dec;88(12):1030-4.

This sequence change replaces alanine, which is neutral and non-polar, with threonine, which is neutral and polar, at codon 279 of the F9 protein (p.Ala279Thr). This variant is not present in population databases (gnomAD no frequency). This missense change has been observed in individuals with hemophilia B (PMID: 2066105, 23093250, 27529981, 27865967, 29656491, 34355501, 34708896). This variant is also known as Ala233Thr. ClinVar contains an entry for this variant (Variation ID: 216926). 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 F9 protein function with a negative predictive value of 80%. This variant disrupts the p.Ala279 amino acid residue in F9. Other variant(s) that disrupt this residue have been determined to be pathogenic (Invitae). 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.

PS4, PP3, PP2, PP5, PP4.

Variant summary: F9 c.835G>A (p.Ala279Thr), also reported as p.Ala233Thr, results in a non-conservative amino acid change located in the Serine proteases, trypsin domain (IPR001254) of the encoded protein sequence. Four of five in-silico tools predict a damaging effect of the variant on protein function. The variant was absent in 183314 control chromosomes. c.835G>A has been reported in the literature in multiple individuals affected with Factor IX Deficiency (Hemophilia B) ranging from mild to severe (Green_1990, Hallden_2013, Saad_1994, https://f9-db.eahad.org/). These data indicate that the variant is very likely to be associated with disease. In at least 1 study, FIX:C coagulation activities in samples from affected individuals ranged between 5-22% of controls (Green_1990). The following publications have been ascertained in the context of this evaluation (PMID: 1972560, 24219067, 8091381). ClinVar contains an entry for this variant (Variation ID: 216926). Based on the evidence outlined above, the variant was classified as pathogenic.