Abstract Number: PB0617
Meeting: ISTH 2021 Congress
Background: Mild-moderate (non-severe, NS) Hemophilia A (HA) associates with minor reduction in factor VIII (FVIII:C) (1-40IU/dL) and involve about 60% of HA cases worldwide. Despite the importance of NS-HA, patients are rarely genotyped, particularly in developing countries. Clinical manifestations and laboratory phenotype are frequently mistaken with VWD2N.
Aims: Characterize the F8-genotype and phenotype, exclude VWD2N, in a series of Argentine patients with FVIII:C 1-40IU/dL.
Methods: Thirty-eight unrelated-families including 50 individuals (probands and relatives) were studied. The bleeding phenotype was obtained from clinical records.
Laboratory: FVIII:C (one-stage method), VWF:Ag (ELISA), VWF:RCo (aggregometry). Mixing studies of the patient/control plasmas to evaluate the presence of inhibitors were undertaken by APTT and VWF:RCo.
Genotyping: peripheral blood leukocyte-extracted genomic-DNA was mutational screened by PCR-amplification of all coding and regulatory regions of the F8 followed by conformation sensitive gel electrophoresis (CSGE) and selected-amplimers were characterized by Sanger sequencing. Exons 17-27 of the VWF were PCR-amplified and directly Sanger-sequenced.
N † (%)
|VWF:Ag||VWF:RCo||Major bleeding (% of patients)|
|17 (45)||A2||c.1214T>C p.(Ile405Thr)
c.1475A>G p.(Tyr492Cys) (2x)
c.1569G>T p.(Leu523Leu) (3x)
c.1834C>T p.(Arg593Cys) (3x)
c.2128G>A p.(Gly710Arg) (2x)
|1 (2.5)||a3||c.5096A>T; p.(Tyr1699Phe)||20||76||90||0|
|1 (2.5)||B||c.3637delA p.(Ile1213Phefs*5) (A run)||3||70||65||100|
|7 (18)||A3||c.5428T>C p.(Ser1810Pro) (2x)
c.5954G>A p.(Arg1985Gln) (2x)
|3 (8)||C1||c.6301C>G p.(His2101Asp)
|6 (16)||C2||c.6683G>A p.(Arg2228Gln)
c.6977G>A p.(Arg2326Gln) (2x)
|†N: number of families per specific F8 genotype; ††Human Genome Variant Society (HGSV) recommended nomenclature; RefSeq F8: NG_011403.1; F8 mRNA: NM_000132.3, FVIII Protein: NP_000123.1.|
A HA-causative F8-genotype was identified in all 38 families including 28 pathogenic variants (PV) (Table 1). Eighteen (47%) of them showed 8 recurrent PV (range 2-3 families/PV), whereas the remnant 20 (53%) showed non-recurrent (private) F8-PV (Figure 1). No variants were found in VWF. Patients with major hemorrhages: 48.6%. Laboratory phenotype is shown in Table 1.
Conclusions: Our practical approach is adequate to characterize the PV in NS-HA patients and relatives. The confirmed diagnosis of NS-HA allows an appropriate treatment and genetic counseling for X-linked inheritance highlighting the role of genetic testing and phenotypic assays. Combined HA+VWD2N was excluded in all patients.
The predominance of missense PV (30/38 cases, 79%) mostly targeting the FVIII-A2-domain and the high frequency of recurrent PVs found in NS-HA may reflect a reduced mutational loss due to the relatively conserved reproductive fitness of probands hence preserving the gene pool in the population.
To cite this abstract in AMA style:Rossetti L, Woods AI, Blanco A, Paiva J, Marchione V, Radic CP, Abelleyro MM, Ziegler B, Waisman K, Romero ML, Casinelli MM, Sánchez-Luceros A, De Brasi C. Comprehensive Genotype Characterization in Non-severe Hemophilia A. Mutational Profile and Assessment of the Bleeding Phenotype in Argentine Patients [abstract]. Res Pract Thromb Haemost. 2021; 5 (Suppl 2). https://abstracts.isth.org/abstract/comprehensive-genotype-characterization-in-non-severe-hemophilia-a-mutational-profile-and-assessment-of-the-bleeding-phenotype-in-argentine-patients/. Accessed December 6, 2023.
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