Neprilysin Inhibits Coagulation through Proteolytic Inactivation of Fibrinogen

Area of Science:

• Biochemistry
• Hematology
• Neuroscience

Background:

• Neprilysin (NEP) degrades amyloid beta (Aβ), a key component in Alzheimer's disease (AD).
• Engineered NEP variants were developed as potential AD therapeutics.
• Pre-clinical safety testing revealed that engineered NEP variants increased prothrombin time (PT) and activated partial thromboplastin time (APTT).

Purpose of the Study:

• To investigate the effect of wild-type NEP and an engineered variant on blood coagulation.
• To define the mechanism by which NEP affects coagulation.

Main Methods:

• PT and APTT measurements in cynomolgus monkeys and rats treated with a human serum albumin fusion with an engineered NEP variant (HSA-NEPv).
• In vitro assays including prothrombinase, calibrated automated thrombogram (CAT), and fibrin formation assays.
• N-terminal sequencing of fibrinogen treated with wild-type or variant NEP.

Main Results:

• Both wild-type NEP and HSA-NEPv unexpectedly impaired coagulation, increasing PT and APTT in plasma samples.
• NEP abolished fibrin formation from fibrinogen.
• The mechanism involved cleavage of the N-termini of fibrinogen's Aα- and Bβ-chains, impairing thrombin-initiated fibrin formation.

Conclusions:

• Fibrinogen is identified as a novel substrate for wild-type NEP, suggesting a role in regulating fibrin formation.
• NEP's effect on coagulation may be relevant to neurovascular degeneration observed in AD and cerebral amyloid angiopathy, where NEP levels are reduced.