Mutational spectrum and deep intronic variants in the factor VIII gene of haemophilia A patients

Summary

Haemophilia A (HA) is caused by a broad spectrum of different mutation types in the factor VIII gene (F8). In our patient cohort of more than 2600 HA patients as well as in other published studies, the most frequent cause are missense mutations in different F8 exons or the recurrent intron 22 inversion. Some exons and several specific nucleotide positions represent hot spots for point mutations in the examined cohort. About 4 % of cases remain without mutation after routine HA diagnostic methods including inversion PCRs, Sanger sequencing and multiplex ligation-dependent probe amplification (MLPA). Deep intronic mutations cannot be detected by current standard HA diagnostics but have been reported for several genetic disorders. However, next generation sequencing (NGS) of the whole genomic sequence of the F8 gene allows to identify deep intronic variants. Conclusion: In general, NGS provides an effective approach to screen for different HA causing mutation types in the F8 gene.

Zusammenfassung

Hämophilie A (HA) wird durch ein breites Spektrum verschiedener Mutationstypen im Faktor-VIII-Gen (F8) verursacht. In unserem Patientenkollektiv von über 2600 HA-Patienten und in anderen veröffentlichten Studien sind Missense-Mutationen in verschiedenen F8-Exons sowie die rekurrente Intron-22-Inversion die häufigsten Ursachen der HA. Einige Exons und bestimmte Nukleotid-Positionen stellen Hot-Spots für Punktmutationen in der untersuchten Kohorte dar. Etwa 4% der Fälle bleiben jedoch ungelöst nach Durchfüh-rung der Routine-Diagnostik, die die Inversions-PCRs, Sanger-Sequenzierung und MLPA (Multiplex Ligation-Dependent Probe Amplification) beinhaltet. Tiefe intronische Mutationen können im Rahmen der gegenwärtigen Routine-Diagnostik nicht entdeckt werden, wurden jedoch im Zusammenhang mit verschiedenen genetisch bedingten Erkrankungen berichtet. Im Rahmen der Analyse der kompletten genomischen Sequenz des F8-Gens mittels NGS (Next Generation Sequencing) können solche tiefen intronischen Varianten identifiziert werden. Schlussfolgerung: Generell stellt NGS einen effektiven Ansatz dar, das F8-Gen auf verschiedene HA verursachende Mutationstypen hin zu untersuchen.

Keywords

Factor VIII - haemophilia A - mutation spectrum - next generation sequencing

Schlüsselwörter

Faktor VIII - Hämophilie A - Mutations spektrum - Next-Generation-Sequenzierung

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