TO THE EDITOR:

Factor XI (FXI) is a key element for the amplification of thrombin generation, which participates in the contact pathway. This explains why FXI deficiency has minor physiological relevance but protects against thrombotic events, supporting FXI as an excellent target for antithrombotic treatments.1,2 

FXI deficiency (MIM #612416) has been considered a rare disorder that might reach relatively high prevalence in certain populations. The best example is within the Ashkenazi Jews, demonstrating an incidence of 8% of heterozygous individuals for 2 mutations: p.Glu117Ter and p.Phe283Leu, which account for 98% of alleles.3  So far, 220 different causative mutations have been identified throughout the FXI gene (F11) (http://www.factorix.org/). Recently, the exome-based data obtained from >60 000 individuals (Exome Aggregation Consortium) have revealed new information regarding genetic variation in F11, showing (a) profound differences in heterozygous frequency among populations, and (b) evidence of recurrent and ethnic-specific mutations: (1) p.Phe223Leu in Africans (23.5% of all mutated alleles); (2) p.Gln263Ter and p.Leu424CysfsTer in East Asians (28.2% and 20.5%, respectively); and (3) p.Ala412Thr in Latinos (25%).4 

The screenings of FXI deficiency for 20 years in Yecla, a small town in the southeast of Spain, have revealed a recurrent point mutation, c.166T>C p.Cys38Arg, identified in 24 out of 46 unrelated cases (52%), and reached a frequency of 2% in the general population.5  This mutation was initially described in the French Basque country as the most prevalent genetic defect causing FXI deficiency (8 out of 12 cases: 66.7%) and also with high frequency in the general population (1%).6  The same mutation was identified in French Brittany in 3 out of 12 cases (25%).7  One Portuguese patient also had this mutation.8  Here, we detected this mutation in 14 out of 55 (25.5%) unrelated cases with FXI deficiency from Barcelona (in the northeast of Spain but 487 km away from Yecla). However, it has never been described in patients with FXI deficiency from non-European populations9-11  and was also absent from large UK or Italy cohorts of patients (supplemental Figure 1, available on the Blood Web site).12,13 

Because p.Cys38Arg mutation was thought to be related to a founder effect in the French Basque country,6  the present study was performed to confirm a potential common founder effect and to estimate its occurrence time.

This study was conducted in 64 subjects with the p.Cys38Arg mutation (30 from Yecla; 27 from Barcelona; 3 from the French Brittany; and 4 from the French Basque Country) well characterized by molecular, functional, and biochemical methods previously described.5  In addition, 20 unrelated noncarrier family trios from Yecla were enrolled as controls. All subjects gave their informed consent to enter the study, which was performed according to the Declaration of Helsinki (Edinburgh 2000) (see supplemental Materials for additional information).

The whole F11 (23 Kpb) was sequenced by next-generation sequencing (supplemental Materials).5  All carriers shared a unique intragenic haplotype. The selection of single-nucleotide polymorphisms with minor allele frequency (MAF) < 0.6 revealed the common F11 haplotype containing 13 polymorphisms, rs566328623 (MAF 0.007) being the most informative variant linked to the p.Cys38Arg mutation (Table 1).

Table 1.

F11 haplotype of carriers of the p.Cys38Arg gene variation

dbSNP IDrs4253398rs3822057rs4241823rs4253405rs4241824rs566328623rs2036914p.Cys38Arg (rs121965069)rs4253409rs2055916rs4253416rs4253417rs3756011rs2289252
chr4* 187188061 187188152 187189294 187190810 187191787 187191859 187192481 187192873 187194685 187196510 187197994 187199005 187206249 187207381 
HGVSc c.-1-229T>C c.-1-138A>C c.55+949T>A c.56-1953A>G c.56-976G>A c.56-904T>A c.56-282T>C c.166T>C c.325+354C>T c.486-431G>A c.755+450C>T c.755+1461T>C c.1305-543C>A c.1481-188C>T 
Type Intron 1 Intron 1 Intron 2 Intron 2 Intron 2 Intron 2 Intron 2 Exon 3 Intron 4 Intron 5 Intron 7 Intron 7 Intron 11 Intron 12 
Frequency 0.211 0.462 0.461 0.294 0.413 0.007 0.393 0.00002 0.263 0.515 0.594 0.297 0.321 0.319 
Allele 
dbSNP IDrs4253398rs3822057rs4241823rs4253405rs4241824rs566328623rs2036914p.Cys38Arg (rs121965069)rs4253409rs2055916rs4253416rs4253417rs3756011rs2289252
chr4* 187188061 187188152 187189294 187190810 187191787 187191859 187192481 187192873 187194685 187196510 187197994 187199005 187206249 187207381 
HGVSc c.-1-229T>C c.-1-138A>C c.55+949T>A c.56-1953A>G c.56-976G>A c.56-904T>A c.56-282T>C c.166T>C c.325+354C>T c.486-431G>A c.755+450C>T c.755+1461T>C c.1305-543C>A c.1481-188C>T 
Type Intron 1 Intron 1 Intron 2 Intron 2 Intron 2 Intron 2 Intron 2 Exon 3 Intron 4 Intron 5 Intron 7 Intron 7 Intron 11 Intron 12 
Frequency 0.211 0.462 0.461 0.294 0.413 0.007 0.393 0.00002 0.263 0.515 0.594 0.297 0.321 0.319 
Allele 

Localization of intragenic polymorphic markers associated to this mutation. The MAF (data from 1000 Genomes Project) and associated alleles are shown.

*

NC_000004.11.

NM_000128.3.

The allele frequency from all populations of 1000 genomes data, April 2012 version 3.

Interestingly, rs4253413 c.486-361C>T (MAF 0.217) was exclusively detected in all carriers from Yecla, but was absent from all other subjects.

Four microsatellite markers covering 1.7 Mbp upstream and 1.8 Mbp downstream of F11 (supplemental Figure 2; supplemental Materials) were also evaluated.

Extragenic haplotypes were established from homozygous cases and by allele segregation among family. In cases without information, we chose the allele associated with the lowest number of recombination events.

Microsatellite analysis revealed at least 23 potential different extragenic haplotypes among carriers (Table 2), none present in any of the 60 healthy subjects from the family trios coming from Yecla (Fisher’s exact test, P < .0001).

Table 2.

Haplotypes defined by F11 extragenic microsatellites markers identified among carriers of the F11 p.Cys38Arg mutation

D4S3047D4S2924F11p.Cys38ArgD4S3051D4S2921Allele numberOrigin
7 (240 pb) 4 (220 pb) 1 (244 pb) 3 (161 pb) Barcelona 
−1* 7(171 pb) 
4 (234 pb) 2 (246 pb) 
5 (167 pb) French Brittany 
6 (224 pb) 6 (169 pb) Barcelona 
3 (232 pb) 
−1 7 (226 pb) 1+1 (Brittany) 
−1 −1 French Basque 
8 (228 pb) Yecla 
6 (238 pb) 
18+1 (Barcelona) 
8 (242 pb) 
Barcelona 
10 (177 pb) 
9 (244 pb) 
     47 Total 
D4S3047D4S2924F11p.Cys38ArgD4S3051D4S2921Allele numberOrigin
7 (240 pb) 4 (220 pb) 1 (244 pb) 3 (161 pb) Barcelona 
−1* 7(171 pb) 
4 (234 pb) 2 (246 pb) 
5 (167 pb) French Brittany 
6 (224 pb) 6 (169 pb) Barcelona 
3 (232 pb) 
−1 7 (226 pb) 1+1 (Brittany) 
−1 −1 French Basque 
8 (228 pb) Yecla 
6 (238 pb) 
18+1 (Barcelona) 
8 (242 pb) 
Barcelona 
10 (177 pb) 
9 (244 pb) 
     47 Total 
*

It was not possible to be established; the mutated allele could be 4 (234 bp) or 7 (240 bp).

The extragenic diversity was lower among carriers from small or isolated populations as Yecla and Brittany than in carriers from the big city of Barcelona (Table 2), as also the linkage of disequilibrium analysis showed (supplemental Table 1; supplemental Materials).

Although the extragenic haplotype was only partially determined in carriers from the Basque Country, microsatellite data showed a remarkable genetic diversity, particularly in the markers closer to F11 (supplemental Table 2).

Estimation of the p.Cys38Arg mutation age was carried out using the Bayesian method proposed by Rannala and Reeve14  and implemented by Disease Mapping Linkage disEquilibrium software (DMLE+2.3).15  The applied map distance, obtained from deCODE map,16  was 3.1 cM/Mb. The growth population rate (r) was obtained from the census results in Yecla during the last 551 years (in 1450, the population was 1400; in 2001, the population was 30 824), considering 25 years for a generation and according to the following formula: r = (LnP1 − LnP0)/g (P = population, g = generations). Thus, Yecla’s population growth rate was estimated to be 0.14. For Europe, we used the growth population rate of the last 2000 to 3000 years (r = 0.034) described elsewhere.17 

The proportion of disease-bearing chromosomes, f = 0.1% in Europe and f = 4% in Yecla, was calculated by (1) estimating the frequency of the p.Cys38Arg mutation (in Europe 1/35 000 [http://exac.broadinstitute.org/variant/4-187192873-T-C], and in Yecla 2/1005 ); (2) considering population sizes (Europe: 700 × 106 and Yecla: 35 000); and (3) the number of chromosomes analyzed (46 for the European cohort and 27 for Yecla).

The density peaked at 216 generations (95% confidence interval: 149-317) when estimating a proportion of 0.2% for the mutated allele according to data from the Exome Aggregation Consortium. The estimated age of this mutation was ∼5400 years. The estimated time of mutation occurrence in Yecla was ∼30 generations (95% confidence interval: 19-50), which represents 625 years (supplemental Figure 3).

FXI deficiency denomination as hemophilia C has certainly contributed to broaden the conclusions on the rarity of this disorder. Textbooks on general medicine, clinical hematology, and hemostasis still report a low prevalence for FXI deficiency (1/10 000-50 000) in the general population. However, increasing evidence from epidemiological studies,5,11,13  the limitation of current diagnostic methods to identify FXI deficiency,18  the clinical mildness of bleeding in even cases with severe FXI deficiency,19  and our current study strongly suggest that the incidence of this disorder might be underestimated.

Our study shows new hot spots of FXI deficiency in Europe raised by a new recurrent and significantly older mutation than that found in the Ashkenazi Jews. The recurrent Ashkenazi’s p.Glu117Ter and the p.Phe283Leu mutations, deriving from relatively few founders that have maintained rather high endogamous rates over time,3,20  were dated back from 120 to 185 and 31 to 100 generations ago, respectively.20,21  We estimated that the p.Cys38Arg variant occurred ∼149 to 317 generations ago. The remarkable extragenic genetic diversity observed in the French Basque Country suggests that this variant might have appeared in this population. Then, it was spread out to the north (reaching the French Brittany) and to the south (arriving to Spain and Portugal). Our study reveals that this variant arrived to Yecla in the thirteenth century, probably during the Christian Reconquest of Southern Spain, with a new intragenic marker. It is also important to consider possible positive selection mechanisms to explain the high prevalence of this mutation in certain populations (Yecla or the French Basque Country), as it has been described for other hemostasis genetic disorders.22  Resistance to infection, hemostasis, or iron conservation has been interpreted as an adaptive profile that might positively select certain gene variations.23  Within this framework, we think that this potential positive selection might not be explained by the antithrombotic protective effect of FXI deficiency.2  A real positive selection could be based on the evidence supporting that FXI deficiency may confer a survival advantage in sepsis by altering the cytokine response to infection and blunting activation of the contact system.24  Alternatively, drift hypothesis cannot be ruled out, because these regions were geographically isolated, with low inward migration and a high degree of consanguinity, as it has been described for other founder mutations involved in different diseases.25 

The online version of this article contains a data supplement.

The authors acknowledge the assistance of Ana Isabel Anton from the genomic platform of Instituto Murciano de Investigación Biosanitaria (IMIB-Arrixaca), and Victoriano Beltran from Hospital Virgen del Castillo, Yecla, Spain.

This work was supported by SAF2017‐91257‐EXP; PI18/00598 (ISCIII and FEDER); 19873/GERM/15 (Fundación Séneca); and the Antonio Lopez Borrasca Award from the Spanish Society of Thrombosis and Haemostasis (SETH). M.E.d.l.M.-B. holds a postdoctoral contract from University of Murcia. S.S.-A. was a fellow of the Spanish Society of Hematology and Hemotherapy during the conduction of this study. B.d.l.M.-B. is a predoctoral fellow of the Fundación Séneca. L.M.-F. is a fellow of the Fundación Española de Trombosis y Hemostasia, Madrid, Spain.

Contribution: M.E.d.l.M.-B. and S.S.-A. performed research, analyzed data, wrote, and reviewed the paper; P.C., B.d.l.M.-B., L.M.-F., F.V., and J.C. performed genetic research and analyzed the data; J.P. and A.M. carried out biochemical and genetic analysis. J.E., P.G., F.B., C.A., and R.P. recruited patient samples, designed the research, and reviewed the paper; P.C., J.C., F.B., and V.V. designed the research, analyzed the data, wrote, and reviewed the paper.

Conflict-of-interest disclosure: The authors declare no competing financial interests.

Correspondence: Pablo Carbonell, Centro de Bioquímica y Genética Clínica, Hospital Universitario Virgen de la Arrixaca, Ctra. Madrid-Cartagena, s/n, 30120 El Palmar, Murcia, Spain; e-mail: pablo.carbonell@carm.es; Javier Corral, University of Murcia, Centro Regional de Hemodonación. C/ Ronda de Garay S/N, 30003 Murcia, Spain; e-mail: javier.corral@carm.es; and Frederic Bauduer, UMR 5199 PACEA, Université de Bordeaux, Allée Geoffroy Saint-Hilaire, CS 50023, 33615 Pessac, France; e-mail: bauduer.frederic@neuf.fr.

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Author notes

*

M.E.d.l.M.-B. and S.S.-A. contributed equally to this work.

F.B., P.C., and J.C. contributed equally to this work and are considered senior authors.