Domain Structure of Clotting Factor VIII (FVIII)

Synthesis as Peptide Chain

•   FVIII is synthesized as a single chain polypeptide of 2351 amino acids
• A 19-amino acid signal peptide is cleaved by a protease shortly after synthesis

Arrangement of single chain of FVIII

• Domain structure of FVIII is as under:
• A1 – a1 – A2 – a2 – B – a3 – A3 – C1 – C2
• FVIII amino acid sequence suggests that the molecule is composed of
• A triplicated homology region
• A1 domain
• A2 domain
• A3 domain
• A duplicated homology region
• C1 domain
• C2 domain
• Heavily glycosylated B domain  
• A1 domain
• Cleavage site (Arg336/337 & 562/563) by APC
• FVIII binds with
• Phospholipids via C1 & C2 domains
• VWF via a3 domain

Significance & Physiological Role of Tissue Factor Pathway Inhibitor (TFPI)

• Tissue Factor Pathway Inhibitor (TFPI) is located

• On Endothelial Cells (ECs)
• On Monocytes
• In Platelets 
• In plasma 

• Bulk of TFPI is associated with ECs

• Measurement of low TFPI in plasma doesn’t mean TFPI deficiency 
• TFPI associated with ECs → 4X ↑in TFPI upon Heparin injection
• Most of circulating TFPI is bound to lipoproteins & the C terminals are variably degraded  

• Physiological Role 

• TFPI inhibits FVIIa in a FXa-dependent manner
• TFPI binds to & directly inhibits TF-FVIIa-FXa complex → So, it effectively halts TF initiated activation of coagulation but only after sufficient FXa has been generated to propagate coagulation
• Plasma TFPI has reduced anticoagulant activity
• So, likely not very important anticoagulant role in vivo

• Therapeutic Role 

• TFPI binds to Heparan Sulphate on EC surface
• UFH or LMWH displace this bound TFPI → This released TFPI contributes to antithrombotic activities of these drugs

• Isoforms of TFPI

• Two isoforms exist 
• TFPIα
• TFPIβ 
• TFPIα has 3 Kunitz type protease inhibitory domains → K1, K2, K3 
• K1 domain inhibits FVIIa in a FXa dependent manner 
• K2 directly binds & inhibits FXa
• Protein S (PS) acts as cofactor in the formation of TFPI-FXa complex

Physiological Inhibitors of Coagulation System

There are many physiological inhibitors impacting various stages of coagulation system. Most important inhibitors with known physiology are enumerated as under: 

• Antithrombin 
• Protein C
• Protein S 
• Heparin
• Heparin cofactor II
• TFPI (Kunitz type inhibitor)
• PZ & PZ dependent inhibitor 
• 𝛂1-Antitrypsin
• 𝛂2-Antiplasmin
• α2-Macroglobulin
• Protease Nexin 2 
• C1-esterase inhibitor

Afibrinogenemia & Important Manifestations/Complications

• Afibrinogenemia is an inherited Quantitative Autosomal Recessive disorder 

• Plasma and platelet levels of the fibrinogen are not measurable 
• Plasma levels <0.1 g/L or <10 mg/dL

• Homozygous or Compound heterozygous

• FGA gene is most commonly involved

• Clinical manifestations

• Bleeding

• Neonatal bleedings like umbilical stump bleeding

• Gastrointestinal bleeding

• Urological bleeding 

• Central nervous sytsem bleeding

• Spleen (with splenic rupture)

• Abnormal wound healing

• Obstetric complications

• 1^st trimester fetal loss

• Antepartum hemorrhage

• Postpartum hemorrhage

• Spontaneous abortion

• Placental abruption

• Bone cysts

• Mostly in long bones → possible effects of bleeding

NOTE: Arterial or venous thrombosis is possible 

Disorders of Fibrinogen (Clotting Factor I)

Quantitative Disorders 

o   Inherited

§  Autosomal Recessive Afibrinogenemia

§  Autosomal Dominant Hypofibrinogenemia 

o   Acquire

§  Hypofibrinogenemia

§  Hyperfibrinogenemia

Qualitative Disorders 

o   Inherited

§  Autosomal Dominant Dysfibrinogenemia

§  Autosomal Dominant Hypodysfibrinogenemia

o   Acquired

§  Liver disease

§  Malignancies 

§  Antifibrinogen antibodies 

Management Principles of Subtherapeutic INR

• Patient education regarding 
• Compliance

• Drug/Food interactions with warfarin  

• Effects of low INR 

• When truly genetic resistance 

• Increase the dose of warfarin 

• Titrate the dose carefully to >100 mg/day with regular monitoring of INR 

• Change the anticoagulant 

• UFH, LMWH, Fondaparinux 

• Rivaroxaban, Dabigatran 

• Other Vitamin K antagonists 

• Bishydroxycoumarin

• Phenprocoumon 

• Acenocoumarol

• Phenindione

Investigating Subtherapeutic & Supratherapeutic INR

•  Full history regarding 

• Compliance
• Drugs
• Diet
• Associated medical conditions 

•  Plasma levels of Warfarin 

• Therapeutic levels 
• At specific intervals after administration 

• FII & FX assays 

• Pharmacogenetic analysis 

• P450 CYP2C9*2 and CYP2C9*3 polymorphism →↑ INR
• Missense mutations of VKORC1 gene CYP 2D6 & CYP2A6 →↓ INR
• Polymorphism in VKORC1 gene → Variable INR