Phenotype-genotype correlations in hemophilia A carriers are consistent with the binary role of the phase between F8 and X-chromosome inactivation.

BACKGROUND: The recessive X-linked disorder hemophilia A (HA) is rarely expressed in female carriers, most of whom express about half of normal factor VIII activity ( FVIII: C). OBJECTIVE: To propose an integrative assessment model for the binary role of the phase between the mutated F8 and the active X-chromosome (Xa) in FVIII: C in HA carriers. METHODS: We studied 67 females at risk of severe HA, comprising five symptomatic females ( FVIII: C < 1.5 IU dL(-1) ) and 14 controls. A correlation study between FVIII: C (observed vs. expected) and X-chromosome inactivation (XCI) patterns (XIPs; androgen receptor gene [AR] system) in blood leukocyte DNA was performed in carriers, by comparison of a model correlating FVIII: C and XIP with arbitrary models devoid of biological significance, and with FVIII: C levels in non-carriers (mean model) as a proxy from background data dispersion not influenced by XIP. RESULTS: We provide proof-of-concept example from a family presenting with extremely skewed XIPs in which the severe HA phenotype appeared in a heterozygous carrier of a crossover between AR and F8 loci that phased the mutated F8 with the maternally inherited Xa. Furthermore, four cases of severe HA affected women who had a combination of a heterozygous F8 mutation and extremely skewed XIPs in leukocytes or oral mucosa are presented. Correlation analyses between FVIII: C levels and XIPs in carriers (n = 38) but not in non-carriers (n = 20) showed highly significant differences between the proposed correlation model and models without biological significance. The data support a binary influence of XCI, either increasing or decreasing the FVIII: C, subject to the underlying phase set between the F8 mutation and XCI. CONCLUSIONS: Our evidence suggests that the phase between XCI and mutated F8 acts as a molecular switch conditioning FVIII: C levels and HA expression in carriers.

Factor VIII genotype characterization of haemophilia A affected patients with transient and permanent inhibitors: a comprehensive Argentine study of inhibitor risks.

Inhibitor development against exogenous factor VIII is a severe impairment of replacement therapy affecting 18% of Argentine patients with severe haemophilia A (HA). To study the molecular predisposition for inhibitor development, we genotyped 260 HA patients with and without inhibitors, countrywide. The inhibitor-positive population (19 transients, 15 low responders, LR and 70 high responders, HR) of 104 severe-HA patients showed 59 Inv22 (intron 22 inversions), 18 small ins/del-frameshifts, 12 gross deletions, 12 nonsense, one splicing defect and two missense, p.Arg531Pro and p.Leu575Pro, both LR and thought to impair FVIII A2 domain secondary structure. In addition, a patient with mild HA and HR showed the missense p.Glu1704Lys associated with two neutral intronic substitutions potentially affecting the A3 domain. A case/control study (84/143) permitted estimation of F8 genotype-specific inhibitor risks [OR; prevalence (CI)] in severe-HA patients classifying a high-risk group including multi-exon deletions [3.66; 55% (19-100)], Inv22 [1.8; 24% (19-100)] and nonsense in FVIII-LCh [1.2; 21% (7-59)]; an average risk group including single-exon deletions, indel frameshifts and nonsense-HCh; and a low-risk group represented by missense defects [0.14; 3% (0.6-11)]. Analysis of inhibitor concordance/discordance in related patients indicated additional genetic factors other than F8 genotype for inhibitor formation. No significant inhibitor-predisposing factors related to FVIII product exposure were found in age- and F8 genotype-stratified populations of severe-HA patients. In conclusion, the Argentine HA patient series presents similar global and mutation-specific inhibitor risks than the HA database and other published series. This case-specific information will help in designing fitted therapies and follow-up protocols in Argentina.