5 - Coagulation Factors Flashcards
(117 cards)
1
Q
Define Thrombosis
A
may be defined as the formation and propagation of a blood clot within the vasculature
2
Q
What is hemostasis?
A
refers to the stoppage of blood flow
3
Q
What is coagulation?
A
it’s a complex process where the blood forms solid clots in response to blood vessel damage
4
Q
What is thrombosis an important part of?
A
the normal hemostatic response that limits hemorrhage caused by vascular injury
5
Q
Under normal conditions, a thrombus is confined to the immediate areas of injury and dose not obstruct flow to critical areas, unless blood vessel lumen is already diminished, as it is in __________
A
atherosclerosis
6
Q
What can a clot cause ?
A
- Acute myocardial infarction (AMI)
- Deep vein thrombosis (DVT)
- Pulmonary embolism (PE)
- Acute ischemic stroke (AIS)
- Acute peripheral arterial occlusion
- Occlusion of indwelling catheters
7
Q
Thrombosis is a complex process and involved in which 3 factors?
A
- Blood flow and the blood vessel
- Platelet-vessel interactions related to the disruption of the endothelium
- Coagulation system
- cellular elements (platelets)
- protein elements (coagulation factors and mediators to promote thrombus formation)
8
Q
When a vascular injures, an immediate local cellular response takes place which attracts ______ to migrate to the area of injury.
A
platelets
9
Q
What do platelets do?
A
Platelets secrete several cellular factors and mediators to promote thrombus formation
10
Q
During thrombus formation, circulating prothrombin is activated to the acting clotting factor _______.
A
thrombin
11
Q
_______ is activated to fibrin by the newly activated thrombin.
A
Fibrinogen
12
Q
What does fibrin do?
A
It is then formed into the fibrin matrix.
13
Q
What are the 3 main components involved in blood clotting?
A
- platelets
- thrombin
- fibrin
14
Q
Each of the 3 components (platelets, thrombin, and fibrin) can be therapeutic targets:
List 3 drugs that can inhibit platelet activation and aggregation.
A
- aspirin
- glycoprotein (GP) 2b/3a inhibitors
- clopidogrel
15
Q
______ gathers in the fibrin matrix.
A
Plasminogen
16
Q
What does fibrin-bound plasminogen do?
A
it will be converted by thrombolytic drugs to plasmin to initiate thrombolysis
17
Q
Thrombolysis is the ______ process of thrombosis
A
opposite
18
Q
What does Thrombolysis involve?
A
It involves fibrin-specific activators to a activate plasminogen at the fibrin surface.
19
Q
What are the thrombolytic agents available today?
A
They are serine proteases that work by converting plasminogen to the natural fibrinolytic agent plasmin.
20
Q
How does plasmin lyse clots?
A
by breaking down the fibrinogen and fibrin in a clot
21
Q
List 1 naturally occurring fibrinolytic agent.
A
Tissue plasminogen activator (tPA)
22
Q
What is tPA involved in?
A
(it is a naturally occurring fibrinolytic agent) found in vascular endothelial cells and is involved in the balance between thrombosis and thrombolysis
23
Q
What does thrombosis exhibit?
A
Significant fibrin specificity and affinity
24
Q
How does the binding of tPA dissolve a clot?
A
At the site of the thrombus, the binding of tPA and plasminogen to the fibrin surface induces a conformational change that facilitates the conversion of plasminogen to plasmin and dissolves the clot
25
Fibrin specific agents produce limited plasminogen conversion in the ______ of fibrin
presence
26
List some examples of fibrin-specific agents
- Alteplase (tPA)
- Reteplase
- Tenecteplase
27
What do non-fibrin-specific agents do?
They catalyze systemic fibrinolysis
28
List an example of a non-fibrin-specific agent
Streptokinase
29
What is hemophilia?
a rare bleeding disorder in which the blood doesn't clot normally.
30
What is the problem in Hemophilia A?
genetic mutation in factor 8 gene
31
What is the problem in Hemophilia B?
lack of factor 9
32
What is the clinical application of recombinant coagulation factors?
Hemophilia (A or B)
33
What are some clinical applications of recombinant thrombolytic agents?
- Acute ischemic stroke
- Acute myocardial infarction (STEMI)
- Acute pulmonary embolism
- Restoration of central venous access devices (ex. dialysis)
34
Describe primary coagulation
Platelets from plasma bind to the exposed collagen via their surface Glycoprotein 2b/3a receptor with von Willebrand factor (vWF), platelets release their granules (ADP and TXA2) to activate more platelets. Fibrinogen crosslinks adjacent platelets forming a platelet plug at the site of injury.
35
Describe secondary coagulation
proteins in plasma (coagulation factors) respond in a complex cascade which eventually leads to fibrin formation (strengthens the platelet plug).
36
The coagulation cascade consists of which two pathways?
- The intrinsic pathway (contact activation pathway)
| - The extrinsic pathway (tissue factor pathway)
37
Describe Hemophilia
- Rare condition
- Inherited
- Usually occurs in males
- Involves long bleeding times. Some are internal, causing damage to organs/tissues (especially ankles, knees, and elbows)
- Little clotting factor which is needed for normal blood clotting
38
Hemophilia A is ______ hemophilia
classical
39
Describe Hemophilia A
- little to no clotting factor 8
| - 90% of people with the disorder have this type
40
Hemophilia B is called _______ disease
Christmas
41
Describe Hemophilia B
- missing or have low levels of clotting factor 9
- 10% of people with the disorder have this type
- named after Steven Christmas, a Canadian who in 1952 was the first person to be diagnosed with this distinct form of hemophilia
42
What is done if hemophilia is suspected?
- Personal and family medical history
- Physical exam
- Blood tests
- Prothrombin time (PT) - extrinsic pathway
- Activated partial thromboplastin time (APTT) - intrinsic and common pathway
43
What causes hemophilia in general?
Defect in one of the genes that determines how the body makes blood clotting factors 8 or 9 --> located on X chromosomes
44
What causes hemophilia in males?
abnormal gene on X chromosome results in hemophilia
45
What causes hemophilia in females?
Must have abnormal gene on both X chromosomes (very rare)
-Females can be a carrier if they have an abnormal gene on one of their X chromosomes - can pass the gene onto her children
46
Describe the severity index of hemophilia
- can range from mild to severe
| - depends on how much clotting factor is in the blood
47
How much clotting factor is in MILD hemophilia ?
5-30% of normal factor
48
How much clotting factor is in MODERATE hemophilia ?
1-5% of normal factor
49
How much clotting factor is in SEVERE hemophilia ?
less than 1% of normal factor
50
What are the symptoms of Hemophilia ?
- Easy bruising
| - Excessive bleeding (external or internal)
51
What are external bleeding symptoms?
Obvious:
- bleeding in the mouth from a cut or bite
- unexplained nosebleeds
- heavy bleeding from a minor cut
52
What are internal bleeding symptoms?
Hidden within the body:
- blood in urine or stool
- bleeding in joints or brain
53
What is factor replacement therapy?
- It is the basic treatment to stop or prevent bleeding in patients with hemophilia A and B.
- It involves the infusion of Factor 8 and 9 concentrates to prevent or control bleeding.
54
Concentrates of factors 8 and 9 come from which 2 sources?
- Human plasma (plasma-derived)
| - Genetically engineered cell line made by DNA technology (recombinant)
55
Describe Factor 8
- plasma protein
- synthesized as a single chain polypeptide for 2332 amino acids
- circulates as an heterodimer of the 80 kDa light chain and a variable (90-210 kDa) heavy chain in a metal ion-dependent complex
56
What is the risk of Recombinate (human albumin as stabilizer) ?
risk of human disease
57
What is the risk of Kogenate or Helixate or Refacto ?
no risk of passing human disease but not very stable
| uses sucrose as stabilizer
58
What is the advantage of Advate ?
More stable !
| uses Trehalose as stabilizer
59
Dosage of recombinant Factor 9 must be individualized to the needs of the patient. It depends on which conditions:
- The severity of the deficiency
- The severity of the hemorrhage
- The presence of inhibitors
- The desired increase in Factor 3 activity
60
There are two factor 9's: What are they produced in?
- CHO (only recombinant)
| - Plasma (purified from the blood)
61
What amino acids are at 148 in CHO Factor 9?
Ala
62
What amino acids are at 148 in Plasma Factor 9?
Thr/Ala
63
Describe the carboxylation of CHO factor 9
40% is under carboxylated, lacking gamma-carboxylation at Glu40
64
Describe the carboxylation of Plasma factor 9
12 gamma-carboxyglutamic acid
65
What does the dose of recombinant Factor 9 depend on?
- the severity of factor 9 deficiency
- the location and extent of bleeding
- the clinical condition
- patient age
- the desired recovery of factor 9
66
Why was recombinant factor 7a developed ?
Because:
- 15-20% of patients with Hemophilia A
- 2-5% of patients with Hemophilia B
will develop antibodies (inhibitors) to factor 8 or 9 respectively after using recombinant factor 8 or 9.
67
Where is recombinant factor 7a produced in?
Baby hamster kidney cells
68
Recombinant factor 7a is expressed in a _____ chain form and is spontaneously activated to factor 7a during purification.
single
69
Recombinant factor 7a is very similar to plasma-derived factor 7a with regard to ?
amino acid sequence, carbohydrate composition and gamma-carboxylation
70
What is tPA?
Tissue Plasminogen Activator
| -A protein involved in the breakdown of blood clots. It is a serine protease found on endothelial cells.
71
As an enzyme, what does tPA catalyze ?
The conversion of plasminogen to plasmin
72
What does the structure of tPA include ?
- consists of 527 amino acids
- with 17 disulfide bridges
- there are 4 N-linked glycosylation sites at residues 117, 184, 218 and 448
73
tPA:
| Describe the F domain
residues 4 to 50, homologous with the finger domain mediating the fibrin affinity of fibronectin
74
tPA:
| Describe the E domain
residues 50 to 87, homologous with epidermal growth factor
75
tPA:
| Describe K1 domain
residues 87 to 176
76
tPA:
| Describe K2 domain
residues 176 to 256
77
tPA:
| What do K1 and K2 domain share?
a high degree of homology with the 5 cringes of plasminogen
78
tPA:
| Describe the P domain
residues 276 to 527, a serine protease region with the active site residues His322, Asp371 and ser478
79
tPA:
| What domains are responsible for binding to fibrin?
F domain and K2 domain
80
tPA:
| What is involved in rapid clearance in vivo with an initial half life of 6 minutes in human?
E domain and carbohydrate side chains
81
tPA:
| What domain is involved in enzymatic activity ?
P domain
82
tPA:
| What is involved in rapid inhibition by plasminogen activator inhibitor-1 ?
Lys296-His-Arg-Arg299
**if we modify this sequence and can make it resistant to plasminogen activator inhibitor 1 or to increase half life
83
tPA:
| What happens if we remove F and E domain?
more resistant to clearance in blood, increased half life
84
What are types of Alteplase?
rtPA, Activase, Cathflow
85
What is Alteplase produced in?
Chinese hamster ovary cells (CHO cells)
86
Alteplase is an enzyme that consists of ___ amino acids
527
87
Alteplase contains 4 distinct regions that contribute to it's structure-function activity. What are they?
- Fibronectin Finger
- EGF growth factor domain
- Kringle domains
- Proteinase domain
88
What does alteplase do?
binds to fibrin in a thrombus (clot) and converts plasminogen to plasmin, thereby initiating local fibrinolysis
89
In an Acute MI:
| What is alteplase used for?
- Lyses of occlusive coronary artery thrombi
- Reduction in mortality
- Improve cardiac function
- Reduce incidence of congestive heart failure
90
Alteplase has to be used within ___ hr of MI
12
91
Alteplase has 2 treatment regimens: a 90 minute accelerated infusion and a 3 hour infusion
Describe the 90 minute accelerated infusion
**don't need to know doses
-recommend up to 6 hours of AMI symptoms
92
Alteplase has 2 treatment regimens: a 90 minute accelerated infusion and a 3 hour infusion
Describe the 3 hour infusion
(still 100mg total)
Effective up to 12 hours of AMI symptoms
93
What is Cathflo ?
Used to restore the function of central venous access devices that have been occluded by thrombus formation
94
What is TNKase?
Tenecteplase is a 527 amino acid glycoprotein
95
TNKase is a modified form of ?
the endogenous tissue plasminogen activation (tPA) molecule
96
What are the 4 domains of TNKase ?
- fibronectin finger
- EGF growth factor domain
- kringle domains
- proteinase domain
97
TNKase is produced in ____ cells
CHO
98
Describe the TNKase use in an AMI (acute myocardial infarction)
- 40mg as a single bolus over a 5-10 second period
| - Never more than 50 mg
99
KNOW slide 37
okay
100
For TNKase, replace the K with __ ______ so can be resistant to plasminogen activation inhibitor 1
4 alanine
101
TNKase:
| What substitution causes glycosylation occurs at 103 position?
a sub of threonine 103 with asparagine means that glycosylation occurs at 103 position
102
TNKase:
| What sub means that the high mannose sugar residue is absent at 117 position?
sub of asparagus 117 with glutamine
103
TNKase:
| What do mutations within the kringle 1 domain do?
- increase plasma half life (T50)
| - 5 to 7 times longer half life vs tPA allowing for singles 5s bolus administration
104
TNKase:
| What does a tetra-alanine substitution at amino acids 296-299 in the protease domain cause ?
- Increased fibrin specificity (14 fold greater fibrin selectivity over tPA)
- Greater resistance to plasminogen activator inhibitor-1 (PAL-1) - 80 fold more resistant to PAL-1 than tPA
105
What are the reasons to develop the second generation of recombinant thrombolytic agents?
- Rapid clearance of recombinant tPA (Alteplase) from the liver
- The first generation of recombinant tPA may cause some general fibrinolysis although it is administered locally
- It has relatively complicated administration procedure
106
What is Reteplase?
It is a 355 amino acid deletion variant of natural t-PA, which consists of 527 amino acids
107
What 2 domains does Reteplase have?
- Protease
| - Kringle 2
108
Reteplase:
| What leads to low fibrin binding affinity?
The kringle 2 domain and the lack of the finger domain lead to low fibrin binding affinity
109
Reteplase:
| What leads to fibrin specificity?
the protease domain
110
Reteplase:
| What type of protein is it?
non-glycoslylated protein
111
Reteplase:
| Is synthesized in ____ as a single chain about 40 kDa peptide
E. coli
112
Reteplase:
| How is it used in acute myocardial infarction in adults (STEMI) ?
- Two doses of 10 unites (18.1 mg) as slow intravenous infusion not to exceed two minutes.
- The second dose is given 30 minutes after the first dose.
113
Describe the general monitoring of thrombolytic agents
- aPPT - activated partial thromboplastin time
- CBC count
- Signs and symptoms of bleeding
- ECG
114
What is internal bleeding ?
Internal: intracranial, retroperitoneal sites, GI, genitourinary, or respiratory tracts
115
What is superficial bleeding?
Superficial: observed mainly at invaded or disturbed sites. Also including epistaxis (nose bleeding) and hematuria (blood cell in urine)
116
What are complications of thrombolytic therapy?
hemorrhage, allergic reactions, embolism, stroke, and repercussion arrhythmias, among others
117
What are some risk factors for hemorrhagic complications?
- Increasing age
- Lower body weight
- Elevated pulse pressure
- Uncontrolled hypertension
- Recent stroke or surgery
- Presence of a bleeding diathesis
- Severe congestive heart failure
