Novel fibrinogen mutations (Aα17Gly→Cys and Aα381Ser→Phe) occurring with a 312Thr→Ala polymorphism

Abstract

The aim of the study was to determine the molecular cause of dysfibrinogenaemia in a woman with a prolonged thrombin time. Functional fibrinogen abnormalities can be benign or may lead to bleeding or thrombotic conditions. In complex cases, phenotypes may be acquired or involve interplay between several coinherited mutations. The authors developed a new whole-protein time-of-flight mass spectrometry (TOF MS) approach to direct targeted DNA sequencing of the fibrinogen genes and determine the phase of multiple substitutions in a single individual. TOF MS analysis of the individual's fibrinogen indicated normal Bβ, γ, and alternately transcribed γ' chain isoforms, but aberrant Aα chain masses. Subsequent fibrinogen Aα gene (FGA) sequencing indicated the presence of three different mutations. Two of the substitutions, Aα17Gly→Cys (at the thrombin cleavage site) and Aα381Ser→Phe (in the αC connector) were novel and the third, Aα312Thr→Ala, was a known benign polymorphism. Accurate mass measurements of isolated control Aα chains showed the predicted Aα polypeptide at 66 132 Da and additional phosphorylated species at + 80 and + 160 Da. Patient's Aα chains on the other hand had masses of 66 103 and 66 241 Da indicating that she had one 312Ala allele (-30 Da) and one 312Thr allele which carried both the 17Gly→Cys (+ 46 Da) and 381Ser→Phe (+ 60) Da mutations. Cotransmission of these new mutations was confirmed by Aα chain TOF MS of plasma fibrinogen and targeted FGA nucleotide sequencing for 10 additional family members.

Dysfibrinogenemia—Potential Impact of Genotype on Thrombosis or Bleeding

“…In fibrinogen Austin Aα Arg17Cys is combined with Aα Ser381-Phe and p.Thr331Ala (rs6050). 44 This patient suffered from menorrhagia only at two occasions. However, the rs6050 SNP was previously shown to be related to thrombotic complication in a patient with hypodysfibrinogenemia (heterozygous for the AαIVS4 þ 1G > T mutation and the Aα4841delC truncation [AαPerth]) and otherwise with a mild bleeding phenotype.…”

“…Individuals with these mutations suffer from bleeding episodes varying from severe menorrhagia, history of easy bruising, subcutaneous hematomas, bleeding after tooth extractions, and post-caesarian bleeding. 13,41,[43][44][45] At residue Pro18, only the Pro!Leu mutation is reported (►Table 4). Patients with this mutation are either asymptomatic though with impaired fibrin monomer polymerization 46,47 or suffer from massive genital bleeding after delivery without any apparent precipitating cause (Kyoto II).…”

Genetics, diagnosis and clinical features of congenital hypodysfibrinogenemia: a systematic literature review and report of a novel mutation

“…More recently, S.O. Brennan's group described the direct analysis of fibrinogen by HPLC time of flight MS . This technique provides a higher resolution, permitting the detection of very subtle molecular changes, as well as the quantification of expression levels of fibrinogen variants .…”

Extension of the Human Fibrinogen Database with Detailed Clinical Information—The αC-Connector Segment

“…There was at least one member of each family in this sub-cohort. We excluded the carrier of p.[G32E];[S333F] and a seven-member family carrying p.[G36C;S400F] as the mutation outside of the αC-connector is considered to cause dysfibrinogenemia [ 52 , 59 ], (see Section 3.5 ). Among this sub-cohort, there are 32 carriers of homozygous mutations (two compound mutations) and 39 carriers of heterozygous mutations (including eight compound mutations).…”