Increased activity of coagulation factor XII (Hageman factor) causes hereditary angioedema type III

Abstract

Hereditary angioedema (HAE) is characterized clinically by recurrent acute skin swelling, abdominal pain, and potentially life-threatening laryngeal edema. Three forms of HAE have been described. The classic forms, HAE types I and II, occur as a consequence of mutations in the C1-inhibitor gene. In contrast to HAE types I and II, HAE type III has been observed exclusively in women, where it appears to be correlated with conditions of high estrogen levels--for example, pregnancy or the use of oral contraceptives. A recent report proposed two missense mutations (c.1032C-->A and c.1032C-->G) in F12, the gene encoding human coagulation factor XII (FXII, or Hageman factor) as a possible cause of HAE type III. Here, we report the occurrence of the c.1032C-->A (p.Thr328Lys) mutation in an HAE type III-affected family of French origin. Investigation of the F12 gene in a large German family did not reveal a coding mutation. Haplotype analysis with use of microsatellite markers is compatible with locus heterogeneity in HAE type III. To shed more light on the pathogenic relevance of the HAE type III-associated p.Thr328Lys mutation, we compared FXII activity and plasma levels in patients carrying the mutation with that of healthy control individuals. Our data strongly suggest that p.Thr328Lys is a gain-of-function mutation that markedly increases FXII amidolytic activity but that does not alter FXII plasma levels. We conclude that enhanced FXII enzymatic plasma activity in female mutation carriers leads to enhanced kinin production, which results in angioedema. Transcription of F12 is positively regulated by estrogens, which may explain why only women are affected with HAE type III. The results of our study represent an important step toward an understanding of the molecular processes involved in HAE type III and provide diagnostic and possibly new therapeutic opportunities.

Figure 1.

Haplotype analysis with use of genetic markers at chromosomal region 5q35.2-q35.5 in a French family with HAE type III. The presence of a C or an A in the second position of codon 328 of F12 (located between D5S2111 and D5S2030) is indicated by the resulting amino acid (Thr or Lys). To avoid disclosure of carrier status of unaffected individuals, only affected individuals, obligate carriers, and founder individuals are depicted. The putative disease-associated haplotype(s) is boxed. A missing genotype is indicated by a hyphen (-). The presence of 328Lys is associated with a history of angioedema attacks.

Figure 2.

Thr328Lys mutation in factor XII in patients with HAE type III. a, Sequencing profiles showing part of exon 9 (nucleotide positions 1027–1037, according to cDNA sequence gi:9961354) in an affected female (III:2) from the French family (left panel) and in a control individual (right panel). b, Schematic picture of the primary structure of the factor XII protein, with known functionally important domains indicated as boxes. Amino acid ranges covering functional domains are given below the primary structure. The location of the Thr328Lys mutation in the proline-rich domain is indicated by an arrow.

Figure 3.

Haplotype analysis in representative parents-offspring triads from three German families (F01, F02, and F05) and one French family, showing the p.Thr328Lys mutation, with use of 10 SNPs covering the complete genomic region of the F12 gene and the disease-causing mutation (c.1032C→A). SNPs without an “rs” number were identified as informative markers by sequencing analysis of F12 in the families (flanking sequences and assay conditions for the SNPs are available on request). Our results show that the disease-causing mutation is located on the same haplotype (red) in all four families segregating the Thr328Lys mutation, which is compatible with the hypothesis that the mutation goes back to a common founder.

Figure 4.

Analysis of FXII activity in plasma from patients with HAE type III. a, Relative FXII amidolytic activity in plasma from HAE type III and healthy individuals (control), determined using the FXIIa-specific chromogenic substrate S-2303. Substrate turnover was measured photometrically via absorbance at a 405-nm wavelength. Data are presented in box-and-whisker plots showing the median (dark line in the box), 25th–75th percentiles (box), and 5th–95th percentiles (whiskers). b, Time course of S-2303 turnover in plasma of a representative patient with HAE type III (“French fam, IV:01”) from the French family, in the absence or presence of the FXIIa inhibitor PCK (2 mM) and in a patient with HAE type III (“F10, IV:11”) from German family F10. For comparison, FXII substrate cleavage in a plasma sample from a healthy control is plotted. Whereas FXII amidolytic activity is markedly increased because of the p.Thr328Lys mutation in the French patient, FXII activity is normal in the German patient.