PB1099 - Mutation Ter462GlnextTer17 introduces a tail to C-terminus of protein C and causes venous thrombosis

Tuesday, June 25, 2024

13:45 – 14:45 ICT

Room: Exhibition Hall

Presenting Author(s)

zhe lai, MA

postgraduate
shanghai ruijin hospital in China
上海市市辖区, Shanghai, China (People's Republic)

Co-Author(s)

YL

Yeling Lu

Thesis Advisor
shanghai ruijin hospital in China
shanghai, Shanghai, China (People's Republic)
XC

Xiaohong Cai

Doctoral Supervisor
ruijin hospital
shanghai, Shanghai, China (People's Republic)
JL

Jiaming Li

M.D
Department of Hematology, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Shanghai, Shanghai, China (People's Republic)
Shijie Zhou, MA

postgraduate
ruijin hospital
Shanghai, Shanghai, China (People's Republic)
Xi Wu, MD

Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
Shanghai, Shanghai, China (People's Republic)
Junwei Yuan, MA

postgraduate
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
Shanghai, Shanghai, China (People's Republic)
WW

Wenman Wu

Professor
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)
QD

Qiulan Ding

Professor
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
Shanghai, Shanghai, China (People's Republic)
XW

Xuefeng Wang

Professor
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)
JD

Jing Dai

Professor
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)

Background: Protein C (PC) is a vitamin K–dependent serine protease in plasma which is activated by thrombin-thrombomodulin(TM) complex to activated protein C (APC). APC can downregulate thrombin generation by inactivating active coagulation factors V and VIII. PC deficiency is associated with an increased risk of venous thromboembolism. We have identified two unrelated VTE patients with the same heterozygous mutation (c.1384T>C, p.Ter462GlnextTer17) in PROC.

Aims: The objective of this study was to determine the molecular basis of the anticoagulant defect in patients carrying the Ter462GlnextTer17 mutation in PROC.

Methods: Plasma was collected from probands and members of their families. PC activity was measured by chromogenic substrate and coagulation assays and PC antigen level was detected by ELISA. A hereditary thrombophilia panel was employed for genetic variant screening.Both wild-type and mutant PC's plasmids were constructed and transfected into HEK-293T cells and then ELISA and Western blot were used to detect the antigen in the supernatant. In vitro enzyme kinetic assay was used to identify the enzymatic properties of PC. APTT and thrombin generation assay were used to detect the anticoagulant function of PC and APC.

Results: Functional studies indicated the activation of the mutant by thrombin or thrombin-TM complex was significantly impaired. APC-Ter462GlnextTer17 exhibited a decreased hydrolytic activity towards the chromogenic substrate S2366. APTT and FVa degradation assays showed that the anticoagulant activity of the mutant protein was markedly reduced both in the absence and presence of protein S. These results were also confirmed by the thrombin generation assay in purified and plasma-based assay systems.

Conclusion(s): The Ter462GlnextTer17 mutation introduces a new tail in C-terminus of PC, which results in impaired activity of the PC zymogen activation and the APC’s anticoagulant activity, accounting for the thrombosis in probands carrying this compound heterozygous mutation.