PB0202 - Molecular Characterization Of The F9 Missense Variants

Sunday, July 10, 2022

6:30 PM – 7:30 PM

Location: Exhibition

Presentation Format: In-Person

Presenting Author(s)

Bulent Bulent Zulfikar, MD

Prof. Dr. O. Bulent Zulfikar, Pediatric Hematology and Oncology
Istanbul University Oncology Institute
Istanbul, Istanbul, Turkey

Co-Author(s)

SG

Seda Genç

PhD Student
Yıldız Teknik University
İstanbul, Istanbul, Turkey
MO

Mehmet Ozbil

Assist. Prof. Dr. Mehmet Ozbil
Gebze Tecnical University
İstanbul, Istanbul, Turkey
Basak Koc, MD

Assoc. Prof.
Istanbul University, Oncology Institute
İstanbul, Istanbul, Turkey
VO

Vahap Okan

Prof. Dr. Vahap Okan
Gaziantep University, Medical Faculty, Department of Hematology
Gaziantep, Gaziantep, Turkey
KK

Kaan Kavakli

Medical Doctor
Department of Hematology, Ege University Children’s Hospital, Izmir, Turkey
Ege, Izmir, Turkey
BA

Bulent Antmen

Prof. A. Bulent Antmen, MD
Department of Pediatric Hematology/Oncology and Bone Marrow Transplantation Unit, Faculty of Medicine, Acibadem University, Adana Hospital
Adana, Adana, Turkey
FS

Fahri Sahin

Prof. Dr. Fahri Sahin
Ege University, Department of Hematology, Ege Adult Hemophilia and Thrombosis Center
İzmir, Izmir, Turkey
AM

Adalet Meral Gunes

Prof. Dr. Adalet Meral Gunes
Bursa Uludag University Pediatric Hematology
Bursa, Bursa, Turkey
YA

Yilmaz Ay

Professor
Pamukkale University Faculty of Medicine, Denizli, Turkey
Denizli, Denizli, Turkey
Canan Albayrak, MD

Professor
Ondokuz Mayis University, Faculty of Medicine, Samsun, Turkey
Samsun, Samsun, Turkey
EU

Ekrem Unal

Prof. Dr. Ekrem Ünal
Erciyes University Faculty of Medicine
Kayseri, Kayseri, Turkey
MD

Mehmet Dagli

Prof. Dr. Mehmet Dagli
Selcuk University, Faculty of Medicine
Konya, Konya, Turkey
EB

Ergul Berber

Prof. Dr. Ergul Berber, Department of Molecular Biology and Genetics
Istanbul Arel University
İstanbul, Istanbul, Turkey

Background: Genotyping in Hemophilia B (HB) is important for genetic counseling, patient management, carrier determination since carrier women also have bleeding risk. Functional characterization of gene variants is important to establish genotype-phenotype correlation. However, minority of F9 mutations were explored through in vitro expression to determine the pathogenicity mode.

Aims: The aim is to genotype F9 gene variations in HB patients in Turkey and to functionally characterize the selected F9 missense variants by in vitro expression and molecular dynamics simulation studies.

Methods: 140 HB patients in Turkey (66 Severe, 39 Moderate, 18 Mild, 19 Unknown), have been being screened for a F9 gene variation by HRM analysis and the variants were identified by direct DNA sequencing. Molecular dynamics simulations for the F9 gene variants were performed by using the GROMACS 5.1.4 software to analyze the conformational effects of the changes. Ethical approval for this study was obtained from the ethical committee of Istanbul Arel University.

Results: HRM analysis and direct DNA sequencing revealed that 60 of the patients had 27 different F9 gene variants. The most common type was missense variation with 59,25% frequency, including one novel variation (p.166Asn>Lys). The most common missense variant was p.194Thr>Ala. Among the patients having p.194Thr>Ala, 9 were severe, 11 were moderate, 4 were mild HB patients. Molecular dynamics simulations for p.194Thr>Ala, p.226 Arg>Gln and p.318 Lys>Arg revealed that p.194Thr>Ala, p.226 Arg>Gln changes caused significant alterations, compared to WT structure while p.318 Lys>Arg did not.

Conclusion(s): The preliminary data demonstrated that the most common F9 variation in the HB population in Turkey is p.194Thr>Ala (50%). Polyphen analysis revealed that it is a benign alteration. However, ESE finder analysis revealed that it creates additional SR protein binding. Molecular dynamics simulations demonstrated that it severely affects F9 protein structure. In vitro expression studies will demonstrate its effect in a heterologous expression system.