Molecular Basis of Congenital Factor XIII Deficiency in Iran

Transcription

1 Review Molecular Basis of Congenital Factor XIII Deficiency in Iran Clinical and Applied Thrombosis/Hemostasis 18, Vol. (2) ª The Author(s) 16 Reprints and permission: sagepub.com/journalspermissions.nav DOI: / journals.sagepub.com/home/cat Akbar Dorgalaleh, PhD 1, Vahideh Assadollahi, PhD 2, Shadi Tabibian, PhD 1, and Morteza Shamsizadeh, MSc 3 Abstract Factor XIII deficiency (FXIIID) is an extremely rare autosomal recessive disorder that has the highest incidence in Iran. The FXIIID is primarily due to mutations in the FXIII-A gene, most of which are unique. In the current study, we report all identified mutations among Iranian patients. Among 483 patients, 366 (75.8%) were molecularly analyzed; 11 different mutations were observed. Of 11, 8 (72.7%) are missense, whereas the remaining 3 (27.3%) are deletion/insertion. Among these patients, 347 (94.9%) had the unique mutation of c.562t>c and 5 (1.4%) had the c.233g>a mutation. c.1226g>a, c.2111g>a, and c.1142t>a are also common, whereas other mutations, including 3 missense and 3 deletion/insertion, were observed only in single patient. Although, in most cases, FXIII mutations are unique and restricted to a specific family, this differs in Iran where a considerable number of identified mutations, recurrently observed, appear to be due to the high rate of consanguinity. Keywords factor XIII deficiency, mutation, molecular diagnosis Introduction Coagulation factor XIII (FXIII) is a tetrameric transglutaminase, consisting of 2 A subunits (FXIII-A), with catalytic function, and 2 B subunits (FXIII-B) as a carrier (A2B2), held together by noncovalent bonds. The FXIII-A is synthesized by cells of bone marrow origin, whereas FXIII-B is produced by hepatocytes. Mature FXIII-A is a 731-amino acid (aa) protein consisting of a 37 aa activation peptide (AP-FXIII), 147 aa b-sandwich, 331 aa catalytic core, the longest domain, as well as b-barrel-1 and b-barrel-2 with 113 and 103 aa, respectively 1-4 (Figure 1). Arg260 aa in the catalytic core domain of 1 FXIII-A subunit forms a bond with Asp404 aa in the other FXIII-A monomer to form FXIII-A2 dimer. Among several cysteines in the FXIII-A subunit protein, it is Cys314, located within the Tyr-Gly-Gln- Cys-Trp sequence of the catalytic core domain, that forms the active site of FXIII protein 1,4,5 (Figure 2). During the activation of FXIII, AP-FXIII is cleaved by thrombin and released into the plasma. After activation, this coagulation factor, in the final stage of blood coagulation, stabilizes fibrin clots and protects them from the fibrinolytic system s proteolytic degradation by forming covalent cross-links between the g-glutamyl and E-lysyl residues of fibrin monomers after activation by thrombin. 1,2,6 FXIII deficiency (FXIIID) is an extremely rare bleeding disorder (RBD) with approximately 1 in 2 million prevalence in the general population Most affected patients have a deficiency of the A subunit. Clinical manifestations of FXIIID include umbilical cord bleeding (UCB), delayed wound healing, subcutaneous and soft tissue bleeding, and recurrent abortion and pregnancy loss, with intracranial bleeding the main cause of death Most bleeding episodes are seen in patients with severe deficiency. The clot solubility test in urea, acetic acid, or monochloroacetic acid was used to detect FXIIID, but this test detects only the severe form. Heterozygotes of FXIIID are diagnosed by quantitative assays such as amine incorporation and ammonia release. 9,15-17 In regions with a high rate of consanguineous marriage, the number of patients with RBDs such as FXIIID is significantly higher. The prevalence of FXIIID in Iran is very much higher than its overall frequency in the rest of the world Iran had 473 patients in 14, with an additional 10 new cases in 16, a total of ,18,19 1 Department of Hematology and Blood Transfusion, School of Allied Medicine, Iran University of Medical Sciences, Tehran, Iran 2 Student Research Committee, Kurdistan University of Medical Sciences, Sanandaj, Iran 3 School of Nursing and Midwifery, Shahroud University of Medical Sciences, Shahroud, Iran Corresponding Author: Akbar Dorgalaleh, Department of Hematology and Blood Transfusion, School of Allied Medicine, Iran University of Medical Sciences, Tehran, Iran. dorgalaleha@gmail.com

2 Dorgalaleh et al 211 English studies using appropriate keywords including rare bleeding disorder in Iran, rare inherited disorder in Iran, recessive bleeding disorder in Iran, factor 13 deficiency in Iran, Factor XIII deficiency in Iran, FXIII deficiency in Iran, FXIII deficiency in Iran, and F13 deficiency in Iran in the title or abstract. These words were used separately and in combination with each other. To avoid missing data, no restrictions were set on the search. If the papers were not found in the original search, they were added manually. Results The data search identified 11 mutations among Iranian patients with FXIIID. Figure 1. X-ray structure of factor XIII-A2 dimer visualized by Swiss- PDB viewer with some editing. The PDB of factor XIII A crystal structure is downloaded from Research Collaboratory for Structural Bioinformatics (RCSB) protein data bank ( Relative position of each domain showed in factor XIII-A2 dimer. Figure 2. X-ray structure of factor XIII-A subunit dimer visualized by Swiss-PDB viewer with some editing. The PDB of factor XIII A crystal structure is downloaded from Research Collaboratory for Structural Bioinformatics (RCSB) protein data bank ( Cys314 location within Tyr-Gly-Gln-Cys-Trp was shown. Methods The aim of this study was to report all identified FXIII gene mutations among Iranian patients. For this purpose, data were obtained from all previous studies on such patients. For this review, we searched Pubmed and Web of Science databases for Molecular Basis of FXIIID Of 483 patients with FXIIID in Iran, 366 patients were molecularly analyzed. Among them, 11 different mutations were observed. Of the 11 mutations, 8 (72.7%) were missense and the remaining 3 (27.3%) were deletion/insertion. Two of these deletions lead to frameshift and, subsequently, premature termination of protein synthesis with the formation of truncated protein. 11, All of these mutations are in the FXIII-A subunit gene. Of the 366 patients, 346 are residents of Sistan and Baluchestan Province, with a unique c.562t>c (Trp187Arg) mutation in the FXIII-A subunit. Another common mutation, found in 5 patients, is c.233g>a (Arg77His) in exon 3. Other common mutations in Iranian patients are c.2111g>a (Arg703Gln) and c.1226g>a (Arg408Gln), each observed in 3 patients. Met380Lys is another missense mutation in exon 9, observed in 2 patients 11, (Table 1). Trp187Arg (c.562t>c) Trp187Arg, observed only in Iran, is the leading cause of FXIIID in the country, affecting 347 (94.9%) patients. Of these, all but one, who lived in Tehran, were resident in Sistan and Baluchestan Province. A control group of 485 normal individuals was checked for this mutation, but none was positive in heterozygote or homozygote state. The most frequent clinical presentation leading to diagnosis of FXIIID was UCB, observed in about 80%. Intracranial hemorrhage (ICH; 17%), deep soft-tissue hematoma (53%), and prolonged wound bleeding (31%) were other common presentations. The FXIII activity was tested in 63 patients, with undetectable levels in all. Most of these patients were closely related (about 70%), but about 10% descended from unrelated parents. This mutation occurs at exon 4 of the FXIII-A gene and affects the catalytic core in the FXIII-A subunit protein. -22 Trp187 is located toward the protein surface between the catalytic core and the b-sandwich domain. In native FXIII-A, Trp187 forms a stable cavity between them. Substitution by arginine in this position results in steric clashes of arginine with the side chains and formation of the hydrophobic cavity

3 Table 1. Spectrum of Factor XIII-A Subunit Gene Mutations in Iranian Patients With Factor XIII Deficiency. Factor XIII Activity a Number Origin Nucleotide* Exchange Amino Acid** Change Exon Domain Mutation Type Number of Participants Clinical Presentations Photometric Assay (%) Putrescine Incorp (%) References 1 Southeast Iran b c.562 T>C Trp187Arg 4 Core Missense 347 UCB, ICH, Hem 5> c 2 ND c.233g>a Arg77His 3 b-sandwich Missense 5 UCB, RPL <1-11 <1-8 3 Tehran c.1226g>a Arg408Gln 10 Core Missense 3 UCB <1 4 Tehran c.2111g>a Arg703Gln 15 Barrel 2 Missense 3 UCB <1 5 Tehran c.1142t>a Met 380 Lys 9 Core Missense 2 UCB <1 6 ND c.781c>t Arg260Cys 6 Core Missense 1 UCB, CNSB ND c.782g>a Arg260His 6 Core Missense 1 UCB <3 <1 8 ND c.1149g>t Arg382Ser 9 Core Missense 1 CNSB, Hem ND c689dela 5 Core Frame shift 1 RPL d, Meno, PSB ND C1981delC 14 Barrel 2 Frame shift 1 UCB, Hem <3 <1 11 Tehran c859_861insggg 7 Core Insertion 1 UCB <1 11, 23 Abbreviations: CNSB, central nervous system bleeding; Hem, hematoma; ICH, intracranial hemorrhage; Meno, menorrhagia; ND, not determined; PSB, postsurgical bleeding; RPL, recurrent pregnancy loss; UCB, umbilical cord bleeding. a Higher activity of factor XIII in photometric assay can be due to the lack of sarum blank. b Of 347 patients, 346 had origin of southeast Iran. c Factor XIII activity was performed in 63 patients. d Thirteen times pregnancy loss. *The nucleotide numbering is based on A nucleotide of ATG initiator as þ1. **Amino acids are numbered based on Serine amino acid as þ1 not initiator Methionine that removed post trnastaionally. 212

4 Dorgalaleh et al 213 between them. These rearrangements may contribute to the destabilization of this region. 7,23 Arg77His (c.3g>a) Another recurrent mutation is c.3g>a in exon 3 of the FXIII-A gene and the b-sandwich of FXIII protein. Of 5 patients with this mutation, 4 had UCB; the fifth experienced recurrent miscarriage (5 times). The FXIII activity was between <1% and 11% by photometric assay. The native Arg77 side chain extends in an H-bonded network. Probably, Arg77 has strong hydrogen bonds with His64 and Phe184. There is also an H bond between Arg77 and His65. Moreover, Arg77 has electrostatic interactions with Tyr69 and Leu328. When Arg77His occurs, the number of H-bonds is significantly decreased and a relatively weak hydrogen bond is formed between His77 and Asn185. This decrease in the H-bonded network probably decreases the stability of FXIII protein, with subsequent early degradation and severe FXIIID. Arg77His has been observed in patients of European origin. Two European patients, from Spain and Belgium, had this mutation in double heterozygote state. Arg77Cys is another common mutation of the FXIII-A gene in this position, observed recurrently in different parts of the world. 7, Arg382Ser (c.1149g>t) The perfect orientation of native Arg382 causes its polar hydrogens to participate in hydrogen binding. Two hydrogen bonds are formed between the Od atoms of Asp384 and the NE and NZ atoms of Arg382, which have a role in stabilizing the core domain of the FXIII-A subunit protein. In addition to these bonds, Arg382 can form 2 strong and 1 relatively weak hydrogen bonds with other amino acid side chains. 7,9 When the Arg382Ser mutation occurs, only 1 weak hydrogen bond can be formed, and the network thus created leads to the destabilization of the FXIII protein and severe FXIIID. 7 This missense mutation in exon 9 and core domain of FXIII- A subunit, leading to severe deficiency, was observed only once, in homozygote state, in an Iranian patient whose clinical presentations include central nervous system (CNS) bleeding and gastrointestinal bleeding. 9 Arg260Cys (c.781c>t) The native Arg260 has a central role in forming several hydrogen bonds. Arg260, in the catalytic core domain of 1 FXIII-A monomer, forms a salt bridge with Asp404 amino acid in another catalytic core domain to form a homodimer. When Arg260Cys occurs, hydrogen bonds are omitted and lead to abnormal folding, instability, and early degradation of FXIII protein. This mutation, which causes abnormal FXIII-A2 dimer formation, is not restricted to Iran, having been observed in patients originating in Poland, Italy, Switzerland, and Japan. 9,25,26 Arg260His (c.782g>a) This mutation was observed in patients from Syria, Italy, and India. Only 1 Iranian patient has this mutation in homozygote state. Arg260 is highly conserved within the transglutaminase family, and normal gene variation in this position is not reported. Similar to Arg260Cys, this mutation causes a decrease in hydrogen bonds, leading to instability and early degradation of FXIII protein. 7,27 c.689dela This single-nucleotide deletion in exon 5 of the FXIII-A subunit gene was reported in only 1 Iranian patient with severe FXIIID. This mutation led to premature stop and the formation of a truncated protein. A missense mutation (c.689a>g) in the current position was observed in an Indian patient with UCB. 7, c.1981delc Another small deletion that led to premature stop and truncated FXIII-A protein was observed solely in exon 14 of the FXIII-A subunit gene. This single-nucleotide deletion was seen only once, in 1 Iranian patient, in homozygote state. 7, Met380Lys (c.1142t>a) Met380, conserved among all homologues of transglutaminase, is not among known polymorphisms of the FXIII-A gene. When Met380Lys occurs, native Met380 is substituted by positive charge Lys that can significantly alter steric interactions with surrounding residues, resulting in destabilized FXIII-A protein and early degradation. This mutation can also affect the electrostatic balance and therefore decrease FXIII-A affinity to its substrate. This nucleotide substitution was observed for the first time in 2 Iranian patients with severe FXIIID. Arg703Gln (c.2111g>a) Substitution of wild-type Arg, as a large charged amino acid, with Gln, as a small negative-charge amino acid, in barrel-2 can cause instability of FXIII-A and early degradation. This mutation was observed for the first time in 3 unrelated Iranian patients. It causes severe FXIIID. 7,18 Arg408Gln (c.1226g>a) Similar to the previous mutation, a small negative-charge amino acid is replaced by a large positive-charge amino acid that can decrease stability of the FXIII-A protein. This mutation was seen in 3 unrelated Iranian patients with severe FXIIID. 7,18,28

5 214 Clinical and Applied Thrombosis/Hemostasis (2) c.859_861insggg (p.286insgly) A triple nucleotide insertion was not observed prior to this patient. Exon 7 has a crucial role in coding the catalytic triad and, thereby, enzymatic activity of FXIII. Any change in this exon can severely affect its activity. 7,28 Discussion With these 3 additions, 156 mutations are observed in the FXIII-A gene, most commonly observed mutations are missense (n ¼ 80, 51.3%). Deletions/insertions are the second most common mutations, representing 23.7% (n ¼ 37). Nonsense mutations are observed with a frequency of 11.5% (n ¼ 18), and splice site mutations comprise 13.5% (n ¼ 21). 1-3 Among Iranian patients, missense mutations are the most frequent of 11 reported mutations, 8 (72.7%) are missense, and the 3 (27.3%) remaining mutations are deletion/insertion. These mutations were scattered throughout the FXIII-A gene 7,28,29 (Figure 3). Of the 366 Iranian patients who were molecularly analyzed, 347 (94.9%) had the c.562t>c mutation; all except 1 are resident in southeast Iran. 7,29 Occurrence of this mutation in a considerable number of patients is suggestive of a founder effect in this population. Since c.562t>c is the sole FXIII-A gene mutation in the area, it has been used successfully for prenatal diagnosis, carrier detection, and premarital counseling. 7 Late diagnosis of FXIIID leads to high rates of morbidity and mortality. The FXIII activity test, however, is not routinely performed due to its high cost. The clot solubility test remains the sole screening test at this time. Identification of a unique mutation in the southeast Iran is very useful for screening and diagnosis 7. All 11 mutations are accompanied by severe, lifethreatening bleeding, most commonly UCB, that is the first presentation among all except the Arg382Ser and c689dela mutations. Although these patients did not present UCB, they had other severe bleeding such as CNS bleeding, recurrent pregnancy loss, and hematoma. Although most of these mutations are new and unique and restricted to a specific family, a few recurrent mutations were observed. In addition to c.562t>c, most occurred among related patients. 7 Recurrent mutations usually were observed among populations with high rates of consanguineous marriage. Pakistan, India, and Tunisia are such populations. Ser295Arg and c.45g>a are the 2 common mutations among Pakistani patients and c.869insc was recurrently observed among Tunisian patients, where the founder effect was seen IVS5 1G>A is one of the most recurrent mutations observed among different populations in Poland, England, Macedonia, the Czech Republic, the Netherlands, Kosovo, Germany, Turkey, and Greece. Several recurrent mutations were noted among Indian patients, and in a recent study, Trp375X was recurrently observed in northern India. 26,33,34 This mutation, similar to Trp187Arg, was Figure 3. Iranian factor XIII-A mutations were scattered throughout factor XIII gene. restricted to a geographic state of the country, although it was seen, as well, in a patient from Tehran Province. This common mutation leads to profound FXIIID with severe life-threatening bleeding such as UCB, ICH and recurrent miscarriage, hematoma, and hemarthrosis. The spectrum of FXIII-A gene mutations in Iranian patients, in contrast to other populations, includes a considerable number of recurrent mutations, and this diathesis seems to be due to the high rate of consanguinity. 7,19 Diagnosis of FXIIID is a challenge worldwide, but determination of the molecular basis of the disorder yields early and precise diagnosis, ensuring that FXIIID can be properly managed.

6 Dorgalaleh et al 215 Acknowledgments The authors appreciate Daisy Morant s work in editing the English version of this article. Declaration of Conflicting Interests The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. Funding The author(s) received no financial support for the research, authorship, and/or publication of this article. References 1. Dorgalaleh A, Rashidpanah J. Blood coagulation factor XIII and factor XIII deficiency. Blood Rev. 16;30(6): Dickneite G, Herwald H, Korte W, Allanore Y, Denton CP, Cerinic MM. Coagulation factor XIII: a multifunctional transglutaminase with clinical potential in a range of conditions. Thromb Haemost. 15;113(4): Schroeder V, Kohler H. New developments in the area of factor XIII. J Thromb Haemost. 13;11(2): Andersson C, Kvist PH, McElhinney K, et al. Factor XIII transglutaminase supports the resolution of mucosal damage in experimental colitis. PLoS One. 15;10(6):e Schroeder V, Handrková H, Dodt J, Kohler HP. Free factor XIII activation peptide affects factor XIII function. Br J Haematol. 15;168(5): Hethershaw E, Cilia La Corte A, Duval C, et al. The effect of blood coagulation factor XIII on fibrin clot structure and fibrinolysis. J Thromb Haemost. 14;12(2): Dorgalaleh A, Naderi M, Hosseini MS, et al. Factor XIII deficiency in Iran: a comprehensive review of the literature. Semin Thromb Hemost. 15;41(3): Naderi M, Dorgalaleh A, Alizadeh S, et al. Polymorphism of thrombin-activatable fibrinolysis inhibitor and risk of intracranial haemorrhage in factor XIII deficiency. Haemophilia. 14;(1): e89-e Dorgalaleh A, Tabibian S, Hosseini S, Shamsizadeh M. Guidelines for laboratory diagnosis of factor XIII deficiency. Blood Coagul Fibrinolysis. 16;27(4): Anwar R, Miloszewski KJ. Factor XIII deficiency. Br J Haematol. 1999;107(3): Naderi M, Dorgalaleh A, Alizadeh S, et al. Clinical manifestations and management of life-threatening bleeding in the largest group of patients with severe factor XIII deficiency. Int J Hematol. 14;100(5): Anwar R, Minford A, Gallivan L, Trinh CH, Markham AF. Delayed umbilical bleeding a presenting feature for factor XIII deficiency: clinical features, genetics, and management. Pediatrics. 02;109(2):E Levy JH, Greenberg C. Biology of factor XIII and clinical manifestations of factor XIII deficiency. Transfusion. 13;53(5): Dorgalaleh A, Naderi M, Shamsizadeh M. Morbidity and mortality in a large number of Iranian patients with severe congenital factor XIII deficiency. Ann Hematol. 16;95(3): Dorgalaleh A, Kazemi A, Zaker F, Shamsizadeh M, Rashidpanah J, Mollaei M. Laboratory diagnosis of factor XIII deficiency, routine coagulation tests with quantitative and qualitative methods. Clin Lab. 16;62(4): Kohler H, Ichinose A, Seitz R, Ariens R, Muszbek L. Diagnosis and classification of factor XIII deficiencies. J Thromb Haemost. 11;9(7): Naderi M, Tabibian S, Hosseini MS, et al. Congenital combined deficiency of coagulation factors: a study of seven patients. Blood Coagul Fibrinolysis. 15;26(1): Naderi M, Alizadeh S, Kazemi A, et al. Central nervous system bleeding in pediatric patients with factor XIII deficiency: a study on 23 new cases. Hematology. 15;(2): Naderi M, Alizadeh S, Tabibian S, et al. Effect of social factors on the highest global incidence of congenital factor XIII deficiency in southeast of Iran. Arch Iran Med. 15;18(5):331.. Dorgalaleh A, Tabibian SH, Boustani M, et al. Molecular genetic analysis of ten unrelated Iranian patients with congenital factor XIII deficiency. Int J Lab Hemato Hosseini S, Dorgalaleh A, Bamedi T, et al. First cases of severe congenital factor XIII deficiency in southwestern Afghanistan in the vicinity of southeast of Iran. Blood Coagul Fibrinolysis. 15; 26(8): Naderi M, Tabibian S, Menegatti M, et al. Disseminated intravascular coagulation with positive D-dimer: a controversial clinical feature in severe congenital factor XIII deficiency in southeast Iran. Blood Coagul Fibrinolysis (8): Trinh CH, Sh ElSayed W, Eshghi P, et al. Molecular analysis of sixteen unrelated factor XIIIA deficient families from south-east of Iran. Br J Haematol. 08;140(5): Peyvandi F, Tagliabue L, Menegatti M, et al. Phenotype-genotype characterization of 10 families with severe a subunit factor XIII deficiency. Hum Mutat. 04;23(1): Schroeder V, Meili E, Cung T, Schmutz P, Kohler HP. Characterisation of six novel A-subunit mutations leading to congenital factor XIII deficiency and molecular analysis of the first diagnosed patient with this rare bleeding disorder. Thromb Haemost. 06;95(1): Ivaskevicius V, Seitz R, Kohler HP, et al. International registry on factor XIII deficiency: a basis formed mostly on European data. Thromb Haemost. 07;97(6): Shanbhag S, Shetty S, Ghosh K. Mutations in coagulation factor XIII subunit A in severe factor XIII deficiency patients: five novel mutations detected. Haemophilia. 11;17(5):e843-e Naderi M, Dorgalaleh A, Alizadeh S, et al. Clinical manifestations and management of life-threatening bleeding in the largest group of patients with severe factor XIII deficiency. Int J Hematol. 14 Nov;100(5): Anwar R, Stewart AD, Miloszewski KJ, Losowsky MS, Markham AF. Molecular basis of inherited factor XIII deficiency: identification of multiple mutations provides insights into protein function. Br J Haematol. 1995;91(3): Louhichi N, Medhaffar M, HadjSalem I, et al. Congenital factor XIII deficiency caused by two mutations in eight Tunisian families: molecular confirmation of a founder effect. Ann Hematol. 10;89(5):

7 216 Clinical and Applied Thrombosis/Hemostasis (2) 31. Anwar R, Gallivan L, Miloszewski KJ, Markham AF. Factor XIII deficiency causing mutation, Ser295Arg, in exon 7 of the factor XIIIA gene. Thromb Haemost. 00;84(4): Kulkarni BP, Nair SB, Vijapurkar M, et al. Molecular pathology of rare bleeding disorders (RBDs) in India: a systematic review. PLoS One. 14;9(10):e Shanbhag S, Ghosh K, Shetty S. Genetic basis of severe factor XIII deficiency in a large cohort of Indian patients: identification of fourteen novel mutations. Blood Cells Mol Dis : Dorgalaleh A, Tabibian S, Hosseini MS, et al. Diagnosis of factor XIII deficiency. Hematology. 16;21(7):