Normal C1 Inhibitor Activity Research Articles

Difficulties in diagnosing hereditary angioedema (HAE) with normal c1 inhibitor levels in a patient with a pathogenic mutation in the MEFV gene associated with familial Mediterranean fever

Hereditary angioedema (HAE) is an orphan disease that causes potentially life-threatening edema in various locations and remains serious problem in both pediatrics and adult clinical practice often disguised as various diseases. The HAE diagnosis with normal C1 inhibitor activity is particularly difficult. The purpose of this publication is to demonstrate unique clinical case of HAE with normal C1 inhibitor activity in young patient with a pathogenic mutation in the MEFV gene, which is associated with familial Mediterranean fever. Severe abdominal attacks in this patient have been regarded for a long time as a part of therapeutic pathology (gastroenteritis, gastritis, Familial Mediterranean fever), acute surgical and gynecological diseases, which led to late diagnosis verification. The HAE requires timely diagnosis, elimination of factors that provoke exacerbations and an immediate therapy administration, that significantly improves life quality of patients and helps prevent death.

Forms of Hereditary Angioedema That Are Independent of Kallikrein and Factor XIIa System

Hereditary angioedema (HAE) is an inherited disorder characterized by recurrent episodes of severe soft tissue swelling involving skin and mucous membranes. In most cases, edema is caused by excessive formation of the vasoactive nanopeptide bradykinin (BK) due to dysregulation of the plasma kallikrein-kinin system (KKS). The KKS consists of the zymogens prekallikrein (PK) and factor XII (FXII) and the cofactor/substrate high-molecular-weight kininogen (HK). PK and FXII reciprocally convert each other to the proteases plasma kallikrein (PKa) and FXIIa. PKa cleaves HK after Lys362 and Arg371 to release bradykinin. In most cases of HAE, increased KKS activation is due to low plasma activity of C1-Inhibitor (C1-INH), the main regulator of PKa and FXIIa. However, ~10% of HAE patients have normal C1-INH activity (HAEnC1). A relationship between fibrinolysis and kinin formation was first described in 1971. The fibrinolytic protease plasmin activates FXII and cleaves HK, although it is unclear that this results in significant bradykinin generation. Recently mutations in the PLG gene encoding plasminogen (Lys311Glu) and the KNG1 gene encoding HK and the related protein low-molecular-weight kininogen (LK, Met361Lys) were identified in patients with HAEnC1. Here we describe how the fibrinolytic system contributes to angioedema in patients with these mutations. Human Glu-plasminogen (Plg), HK and LK were expressed in HEK293 cells. Lys311 in plasminogen was changed to glutamic acid, and Met361 in HK and LK were changed to lysine, by site directed mutagenesis. Wild type (Plg-Lys311) and variant (Plg-Glu311) plasminogen were converted to plasmin (Plm-Lys311 and Plm-Glu311) with urokinase. The capacity of plasmin to cleave HK and LK were studied with SDS-PAGE, western blots and kinin ELISA. Cleavage of wild type HK-Met361 and LK-Met361 and variant HK-Lys361 and LK-Lys361 by PKa and plasmin were assessed in a similar manner. The types of kinin released during reactions were determined by mass spectroscopy. Normal plasma was supplemented with Plg-Lys311 or Plg-Glu311, and tPA was added to generate plasmin. Bradykinin release was substantially greater with Plm-Glu311 than Plm-Lys311. This held when reactions were run in plasma lacking FXII or PK, indicating activation of the KKS was not responsible for kinin production. Using plasma-derived proteins, PKa released bradykinin 100-fold faster from HK than LK, and released bradykinin from HK ~50-fold faster than did plasmin. PKa and plasmin released bradykinin at comparable relatively slow rates from LK. Plm-Glu311 released bradykinin from HK 10-fold faster, and from LK 3-fold faster, than in reactions with Plm-Lys311. In experiments in which kininogen deficient plasma was supplemented with either HK or LK to a physiologic concentration, the majority of bradykinin released by Plm-Glu311 was from LK. PKa-catalyzed release of bradykinin from wild type HK-Met361 and variant HK-Lys361 comparably. The same held for LK-Met361 and LK-Lys361. However, plasmin released kinin from HK-Lys361 and LK-Lys361 3.3-fold and 13-fold faster than from HK-Met361 and LK-Met361. While the kinin released from HK-Met361 and LK-Met361 was bradykinin, plasmin released the decapeptide Lys-bradykinin (kallidin) from HK-Lys361 and LK-Lys36. Our studies indicate that angioedema in some patients with HAEnC1 may be due to fibrinolytic activity, and not to KKS activity. Both HK and LK may be substrates in such patients. As the plasma LK concentration is ~4-fold higher than HK, it may be the main pathologic kinin source. The plasmin Lys311Glu substitution converts the protease into a more efficient kininogenase. Fibrinolytic activity is normally brisk in the mouth, perhaps explaining why patients with the Lys311Glu mutation primarily suffer from oral-lingual angioedema. The findings raise the possibility that oral-lingual edema after tPA infusion or ACE inhibitor administration may also be due to plasmin-mediated cleavage of LK and HK. The Met361Lys substitution in HK and LK introduces a novel protease cleavage site immediately upstream from a normal PKa cleavage site. The novel site is preferred over the normal site by plasmin, but not PKa. Several treatments for HAE directed at neutralizing PKa or FXIIa activity have been developed. Our results suggest that such drugs may be less effective in patients with HAEnC1 in which disease mechanisms operate independently of the KKS.

Mechanisms involved in hereditary angioedema with normal C1-inhibitor activity.

Patients with the inherited disorder hereditary angioedema (HAE) suffer from episodes of soft tissue swelling due to excessive bradykinin production. In most cases, dysregulation of the plasma kallikrein-kinin system due to deficiency of plasma C1 inhibitor is the underlying cause. However, at least 10% of HAE patients have normal plasma C1 inhibitor activity levels, indicating their syndrome is the result of other causes. Two mutations in plasma protease zymogens that appear causative for HAE with normal C1 inhibitor activity have been identified in multiple families. Both appear to alter protease activity in a gain-of-function manner. Lysine or arginine substitutions for threonine 309 in factor XII introduces a new protease cleavage site that results in formation of a truncated factor XII protein (Δ-factor XII) that accelerates kallikrein-kinin system activity. A glutamic acid substitution for lysine 311 in the fibrinolytic protein plasminogen creates a consensus binding site for lysine/arginine side chains. The plasmin form of the variant plasminogen cleaves plasma kininogens to release bradykinin directly, bypassing the kallikrein-kinin system. Here we review work on the mechanisms of action of the FXII-Lys/Arg309 and Plasminogen-Glu311 variants, and discuss the clinical implications of these mechanisms.

Unnecessary Abdominal Surgeries in Attacks of Hereditary Angioedema with Normal C1 Inhibitor.

Hereditary angioedema (HAE) is an autosomal dominant disease mostly due to the deficiency of C1 inhibitor (C1-INH). HAE with normal C1-INH was first described in 2000 and associated with mutations in the coagulation factor XII in 2006. Both diseases are associated with high bradykinin production, resulting in increased vascular permeability. Gastrointestinal edema due to HAE can be misdiagnosed as acute abdomen and unnecessary surgical procedures may be performed. The present study evaluates the prevalence of surgical procedures and/or acute abdomen in HAE patients with the coagulation factor XII mutation. It is a retrospective study where patients were diagnosed with recurrent angioedema without urticaria, normal C1-INH levels, and positive family history of angioedema. All patients were evaluated for the known mutations located at exon 9 of the F12 gene. Medical records were evaluated and questionnaires were applied to 52 patients with normal C1-INH levels (age range 13-76years; 47/52, 90.38% women; 5/52, 9.61% men). F12 mutation was present in 32/52 patients (61.5%). Acute abdominal pain was diagnosed in 16/52 (30.76%) patients, appendicitis in 9/16 (56.2%), and undetermined diagnosis in 7/16 (43.7%). Among patients diagnosed with acute abdominal pain, 13/16 (81.2%) underwent surgery and 3/16 (18.7%) improved without surgical intervention. We conclude that many HAE patients with coagulation factor XII mutation were misdiagnosed with acute abdomen and subjected to unnecessary invasive procedures. It is critical to disseminate information about this rare mutation in patients with otherwise normal C1-INH activity, in order to speed up diagnosis and avoid misconduct.

Leveraging Genetics for Hereditary Angioedema: A Road Map to Precision Medicine.

Biochemical studies performed during the last decades resulted in the development of various innovative medicinal products for hereditary angioedema (HAE). These therapeutic agents target the production or the function of bradykinin-the main mediator of HAE due to C1-inhibitor (C1-INH) deficiency. However, despite these remarkable achievements, current knowledge cannot provide convincing explanations for the clinical variability of the disease. As a consequence, treatment indications apply for drugs available for C1-INH deficiency. The advent of high-throughput next-generation sequencing technologies may assist in covering the missing part of our understanding of HAE pathogenesis. During the last 3 years alone, several new entities were added to the already described genotypes. The recent discovery of four novel target genes expands our understanding of other causes which may explain recurrent angioedema in individuals and families with normal C1-INH activity. Furthermore, new genetic technologies allowed the recognition of deep intronic variants associated with the disease, and elegant functional studies characterized new variants for the C1-INH gene. Thus, evidence has been provided regarding pathogenetic aspects remaining obscure for many years, such as the defective intracellular transport of mutant C1-INH, and environmental effect on the disease expression. Therefore, it seems that the stage for Precision Medicine era in HAE management is ready. Disease endotypes are expected to be uncovered and specified targets for therapeutic intervention will be detected, promising a more effective, individualized management of the disease.

Exploration biologique des angiœdèmes à kinines

Kinin-mediated angioedema results from accumulation of kinins, vasoactive and vasopermeant peptides, on the vascular endothelium. The disease is characterized by sudden episodes of swelling in the subcutaneous and submucosal tissues; the edema may occur spontaneously or it may be precipitated by triggering factors such as physical or emotional stress, or certain medicines. The characterization of kinin formation and catabolism systems helps improve knowledge of the aetiopathogenic mechanisms involved and provides the basis for classification of kinin-mediated angioedema conditions; thus, we may distinguish between angioedema with C1 inhibitor deficiency, whether inherited or acquired, and angioedema with normal C1 inhibitor activity, associated with increased kinin-forming activity or deficiency in kinin catabolism enzymes. In support of the clinical diagnosis, the physician may request laboratory investigation for a functional and molecular definition of the disease. Laboratory diagnosis is based on the characterization of: (1) kinin production control by C1 inhibitor investigation (function, antigen levels and circulating species); (2) kinin production (kininogenase activity, kininogen cleavage species); and (3) kinin catabolism enzymes (aminopeptidase P, carboxypeptidase N, angiotensin-I converting enzyme and dipeptidyl peptidase IV). An abnormal biological phenotype is supported by examination of susceptibility genes (SERPING1, F12 and XPNPEP2) and mutation segregation in the families.

Antihistamine-resistant Angioedema in Women with Negative Family History: Estrogens and F12 Gene Mutations

BackgroundIn women with sporadic recurrent angioedema with an unknown cause who are unresponsive to antihistamines and have normal C1 inhibitor activity and a negative family history of angioedema, it is unclear whether they have idiopathic angioedema or hereditary angioedema with normal C1 inhibitor, and what impact exogenous estrogens have on their angioedema. MethodsA cohort of 147 women was analyzed for F12 exon 9 mutations and for the influence of oral contraceptives, hormonal replacement therapy, and pregnancy on their angioedema. ResultsA total of 142 women had idiopathic angioedema unresponsive to antihistamines. Five women had an F12 mutation and thereby hereditary angioedema with F12 mutations. Among the women with idiopathic angioedema, 63 had never taken estrogens. There was no estrogen impact in 42 women, a moderate impact in 15 women, and a severe impact in 22 women. The type and dose of estrogens did not differ in women with and without an estrogen impact. In 5 women, idiopathic angioedema disappeared after desogestrel use. Among the 5 women with hereditary angioedema with F12 mutations, angioedema symptoms occurred during 4 pregnancies, whereas no symptoms occurred during any of the 58 pregnancies in women with idiopathic angioedema. ConclusionsWomen with recurrent angioedema unresponsive to antihistamines may have idiopathic angioedema or, more rarely, hereditary angioedema with F12 mutations. Both conditions may be provoked or aggravated by exogenous estrogens. In idiopathic angioedema, treatment with progestins may be helpful.

Small bowel angioedema due to acquired C1 inhibitor deficiency

Acquired angioedema is a rare disorder caused by an acquired deficiency of C1 inhibitor. It is characterized by nonpitting, nonpruritic subcutaneous or submucosal edema of the skin, or of the respiratory or gastrointestinal tract. When localized in the gastrointestinal tract, it can cause severe abdominal pain, mimicking an acute surgical abdomen, or chronic recurrent pain of moderate intensity. We report a case of a 48-year-old man presenting with recurrent episodes of hypotension and abdominal pain. Computed tomography of the abdomen showed edema of the small bowel. The first determinations of C1 inhibitor level and activity, measured in a symptom-free period, were normal. Repetition of the laboratory tests in the acute phase, however, showed a low C1 inhibitor level. Further diagnostic work-up indicated an acquired C1 inhibitor deficiency caused by a monoclonal gammopathy. He was treated with tranexamic acid as prophylaxis for his frequent attacks and to date, he has remained symptom free. Acquired C1 inhibitor deficiency is a rare cause of angioedema and is, among others, related to autoantibodies and abnormal B-cell proliferation, for example monoclonal gammopathy. The diagnosis of acquired C1 inhibitor deficiency is made on the basis of the medical history and on the level and activity of plasma C4, C1q, and C1 inhibitor. In case of high suspicion and a normal C1 inhibitor activity, it is recommended to repeat this test during an angioedema attack. Early diagnosis is important for the treatment of severe, potentially life-threatening attacks and to start prophylactic treatment in patients with frequent or severe angioedema attacks.

Clinical, biochemical, and genetic characterization of type III hereditary angioedema in 13 Northwest Spanish families

BackgroundA new variant of hereditary angioedema has been reported during the last decade. Three main characteristics distinguish it from classic hereditary angioedema: normal C1 inhibitor activity, predominance in women, and different genetic alterations. ObjectiveTo assess the symptoms, laboratory findings, and treatment of a population with type III hereditary angioedema from Northwest Spain. MethodsWe studied 29 patients (26 female and 3 male) from 13 different families. ResultsThe 26 female patients showed a similar clinical pattern to the classic forms of hereditary angioedema, and 22 of these patients had the estrogen-dependent phenotype. Three patients had a negative family history, and 1 of the parents was confirmed as an asymptomatic carrier in 2 of them. All had functional C1 inhibitor activity within the normal range in periods without high estrogen levels, but during attacks (in female patients) and pregnancy, activity decreased to below 50%. One male patient had normal C1 inhibitor activity during attacks, and he was initially diagnosed as having idiopathic angioedema. The C4 and antigenic C1 inhibitor levels were always normal. All studied patients had the c.1032C>A, Thr309Lys mutation in the factor XII gene. The mutation was also found in asymptomatic relatives: 5 of 6 men studied and 1 of 8 women studied. ConclusionPositive family history is a diagnosis criterion, but it could be lacking because there may be asymptomatic relatives, primarily males.

Hereditary Angioedema in Childhood

Hereditary angioedema (HAE) is an inherited disorder characterized by recurrent, circumscribed, non-pitting, non-pruritic, and rather painful subepithelial swelling of sudden onset, which fades during the course of 48-72 hours, but can persist for up to 1 week. Lesions can be solitary or multiple, and primarily involve the extremities, larynx, face, esophagus, and bowel wall. Patients with HAE experience angioedema because of a defective control of the plasma kinin-forming cascade that is activated through contact with negatively charged endothelial macromolecules leading to binding and auto-activation of coagulation factor XII, activation of prekallikrein to kallikrein by factor XIIa, and cleavage of high-molecular-weight kininogen by kallikrein to release the highly potent vasodilator bradykinin. Three forms of HAE have currently been described. Type I and type II HAE are rare autosomal dominant diseases due to mutations in the C1-inhibitor gene (SERPING1). C1-inhibitor mutations that cause type I HAE occur throughout the gene and result in truncated or misfolded proteins with a deficiency in the levels of antigenic and functional C1-inhibitor. Mutations that cause type II HAE generally involve exon 8 at or adjacent to the active site, resulting in an antigenically intact but dysfunctional mutant protein. In contrast, type III HAE (also called estrogen-dependent HAE) is characterized by normal C1-inhibitor activity. The diagnosis of HAE is suggested by a positive family history, the absence of accompanying pruritus or urticaria, the presence of recurrent gastrointestinal attacks of colic, and episodes of laryngeal edema. Estrogens may exacerbate attacks, and in some patients attacks are precipitated by trauma, inflammation, or psychological stress. For type I and type II HAE, diminished C4 concentrations are highly suggestive for the diagnosis. Further laboratory diagnosis depends on demonstrating a deficiency of C1-inhibitor antigen (type I) in most kindreds, but some kindreds have an antigenically intact but dysfunctional protein (type II) and require a functional assay to establish the diagnosis. There are no particular laboratory findings in type III HAE. Prophylactic administration of either 17alpha-alkylated androgens or synthetic antifibrinolytic agents has proven useful in reducing the frequency or severity of attacks. Plasma-derived C1-inhibitor concentrate, recombinant C1-inhibitor, ecallantide (DX88; a plasma kallikrein inhibitor) and icatibant (a bradykinin B(2) receptor antagonist) have demonstrated significant efficacy in the treatment of acute attacks, whereas the C1-inhibitor concentrate has also provided a significant benefit as long-term prophylaxis. However, these drugs are not licensed in all countries and are not always readily available.

Diagnosis and treatment of hereditary angioedema with normal C1 inhibitor

Until recently it was assumed that hereditary angioedema is a disease that results exclusively from a genetic deficiency of the C1 inhibitor. In 2000, families with hereditary angioedema, normal C1 inhibitor activity and protein in plasma were described. Since then numerous patients and families with that condition have been reported. Most of the patients by far were women. In many of the affected women, oral contraceptives, hormone replacement therapy containing estrogens, and pregnancies triggered the clinical symptoms. Recently, in some families mutations in the coagulation factor XII (Hageman factor) gene were detected in the affected persons.

Hereditary angioedema with normal C1 inhibition

Until recently, it was assumed that hereditary angioedema was a disease resulting exclusively from a genetic deficiency of the C1 inhibitor. In 2000, families with hereditary angioedema, normal C1 inhibitor activity, and protein in plasma were described. Since then, numerous patients and families with this condition have been reported. Most of the patients were women. In many of the affected women, oral contraceptives, hormone replacement therapy containing estrogens, and pregnancies triggered the clinical symptoms. In some families, mutations in the coagulation factor XII (Hageman factor) gene were detected in the affected persons.

Increased Activity of Coagulation Factor XII (Hageman Factor) Causes Hereditary Angioedema Type III

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.

Hereditary angioedema with normal C1-inhibitor activity in women

Hereditary angioedema (HAE) is a well defined autosomal dominant disease (Mendelian Inheritance in Man #106100) that results from an inherited deficiency of C1 (the activated first component of complement) inhibitor function. We report an unusual variant of HAE with normal biochemical C1-inhibitor function, occurring only in women. We screened 574 patients with recurrent angioedema of the skin for presence of HAE. 283 patients were selected, in whom angioedema was associated with abdominal pain attacks or recurrent life-threatening episodes of upper-airway obstruction, or both, rather than with urticaria. We measured C1-inhibitor concentration and functional activity as well as complement C4 concentration and took pedigrees to characterise patients. 94 HAE cases with C1-inhibitor deficiency, positive family history, or both were identified. Biochemical testing showed that 84 patients from 49 families had a functional C1-inhibitor deficiency. 11 of these patients had no affected family members (probably representing de-novo mutations). Ten women with HAE, from ten families, had normal C1-inhibitor protein concentrations and function, and normal C4 concentration. A more detailed study of these families identified another 26 affected members, who were also all women. Of those women, 14 could be studied and also had normal C1-inhibitor concentration and function. The disease was seen in successive generations, and in offspring of affected mothers, the sex ratio (M/F) was shifted to 1/1.5. HAE with normal C1-inhibitor concentration and function represents a unique genetic disease arising only in women. The formal genetics of this entity are suggestive of an X-linked dominant mode of inheritance. For this disorder we propose the term hereditary angioedema type 3 (HAE III).