Haemostasis is the normal control mechanism of the body to prevent blood loss.  When a blood vessel is breached, due to some traumatic injury, blood may leak out.  The processes involved in haemostasis prevent this blood loss.  However, as with other systems of the body, these processes may also be detrimental to the overall health of the organism.
            When a vessel is compromised so that blood loss occurs, the first response is intense vasoconstriction, in an attempt to reduce blood flow to the injured area until repair can be accomplished.  Additionally, when an injury occurs, the connective tissue (primarily collagen) of the vessel is exposed.  This is the initiating sequence of clotting.  Recall that collagen may be adhesive.  Also, recall that platelets (which circulate freely in the blood) are also adhesive.  Therefore, platelets will "stick" to the exposed collagen,  an action that that thought to be dependent upon an ADP-mediated glycoprotein identified as the IIb/IIIa receptor.  This is referred to as platelet adherence and subsequent clotting activity is dependent upon this action.  As the first platelets adhere to the collagen, they in turn will become more adhesive, attracting additional platelets to the area.  Platelet aggregation refers to this migration of platelets to the injured area.  As additional platelets adhere to each other and the injured area, they will begin to lose their membrane and form a gelatinous platelet plug, which helps to block the breach in the vessel, thus reducing blood loss.  However, this plug is temporary and may be easily breached itself.  Therefore, during this time, the platelets also release thromboxane A₂ (TXA₂) which promotes the formation of fibrin from thrombin (there is also released a compound which prevents too much thrombin from forming, prostacylin I₂, which is one of the means to prevent over clotting).  The fibrin provides structure to the clot and makes it more stable and less easily breached, thus ensuring decreased blood loss.
          As the clot forms, the initial structure is primarily the platelet plug and fibrin, with some white cells to dispose of cellular debris.  This is referred to as a white thrombus or clot.  Clots in arteries are often limited to white thrombus formation (note that if the clot is large enough it may occlude blood flow distal to the injured area).  However, in vessels of low pressure (i.e. veins) the fibrin may extend in the direction of blood flow and due to the relatively slow velocity of blood flow, red cells may become incorporated into this "tail".  This is referred to as a red thrombus or clot.  This tail may continue to grow and will not adhere to intact vessel wall (where no injury and therefore no exposed collagen occurred).  The tail may break free from the thrombus and flow with the blood down the vessel (it is then called an embolus) potentially occluding the vessel distal to the point of injury.
            The formation of fibrin is central to clot formation and to the action of anti-coagulant drugs.  A diagrammatic representation of this biochemical pathway follows.  Briefly, the process involves the conversion of several biologically inactive factors into activated factors.  Each activated factor, up to the formation of fibrin is a protease that converts the next required factor in the cascade.  Note that some of these factors require more than one activated factor to cause their conversion.

NOTE:  Factor VII in the above schematic is a zymogen, you may hear this term used.  Also, Factor VIII circulates bound to Von Willebrand's factor, another component of the haemostatic pathway.

If this clotting mechanism and ultimate fibrin formation is allowed to continue unchecked, massive coagulation can occur body wide.  If this does occur, it gives rise to a disorder called disseminated intravascular coagulation and may occur following massive tissue injury (when activation of the clotting factor is occurring body wide), profound cellular lysis in certain forms of cancer, bacterial sepsis, and abruptio placentae (spontaneous abortion with premature detachment of the placenta).  Additionally, if clot formation continues, the risk of thrombus occlusion or embolism increases.  Therefore, in most bleeding situations, there are physiologic "brakes" that prevent these from happening.  Endogenous compounds such as alpha-1 antiprotease, alpha-2 macroglobulin, alpha-2 antiplasmin, and anti-thrombin III (AT-III) all contribute to the control of the clotting mechanism.

Additionally, there are mechanisms that exist to break up formed clots (those that are no longer necessary, when the injured area has healed) and to prevent the over-expression of a clot.  Plasminogen, which circulates in the plasma and may also be found in tissues, is activated by both plasma-derived and tissue-derived activating proteases to plasmin.  Plasmin may act on both fibrinogen (breaking it down to fibrinogen degradation products) so that fibrin cannot be formed and on fibrin (forming fibrin split products) thus breaking up existing clots.

Heparin

Mechanism of Action
Heparin (and heparan sulphate) are endogenous compounds that contribute to controlling the coagulation process.  Its activity is dependent upon AT-III.  AT-III binds to and inactivates factors XIIa, XIa, Xa, IXa, and IIa.  This reaction between AT-III and the factors noted occurs relatively slowly.  However, heparin may bind to AT-III, changing its conformation and exposing additional binding sites.  This in turn increases the reaction time for AT-III inactivation of these factors.  Once the inactivation has occurred, the heparin may dissociate from the AT-III (the AT-III stays bound to the factor, irreversibly inhibiting it) and be used again to inhibit another molecule of factor via an additional molecule of AT-III.  NOTE that this action would NOT be effective in clots that have already formed.  It inhibits the clotting mechanism by inhibiting fibrin formation and has no effect in reducing an existing clot.  Also note that heparin will NOT inhibit activated factor IIa (thrombin) that is already bound to fibrinogen (the AT-III:heparin complex must inactivate thrombin prior to its binding to fibrinogen).

Specific Agents --

Heparin is a mucopolysaccharide that is high in sulphate functional groups.  It is these sulphate groups that permit binding of heparin to AT-III.  Since the exact number of sugar molecules in each molecule of heparin may vary, the actual drug is a heterogenous mixture of various molecular weights.  Heparin is therefore often classified as High Molecular Weight (HMW) or Low Molecular Weight (LMW).  Heparin, labelled as such, is typically HMW (5000-30,000 Daltons) and is typically derived from either porcine intestinal mucosa or bovine lung.  HMW heparin will inhibit all factors equally.

LMW heparin is marketed primarily as three preparations:

ENOXAPARIN (2000-6000 Daltons), DALTAPARIN (2000-9000 Daltons), and DANAPROID (2000-9000 Daltons).  These agents are more effective in inhibiting thrombin than the other factors and are used primarily in the prevention of deep vein thrombosis (clot formation in the large veins).  ENOXAPARIN has recently been approved for the TREATMENT of deep vein thrombosis.  Heparin may be administered intravenously or by deep subcutaneous injection.  It should NEVER be administered intramuscularly, due to the high vascularisation of muscle and the very real possibility of deep muscle haemotoma, which is not only painful but will prevent the absorption of heparin as needed.


Adverse Effects -- The most prominent adverse reaction to heparin is in overdose situations in which bleeding occurs.  Therefore, heparin should be used cautiously in patients who have recently taken aspirin, NSAIDs, or oral anti-coagulants; pregnant patients; and is contraindicated in patients diagnosed with haemophilia.  Treatment of heparin overdose will be discussed below.  Other adverse reactions include allergies in those patients susceptible (existing allergies to beef or pork).  Additionally, a small number of patients may exhibit thrombocytopaenia. This has been associated with platelet aggregation such that some patients may actually experience heparin-induced thrombus formation.  This reaction occurs more with bovine heparin and less with porcine-derived heparin.

Monitoring of Heparin Therapy
The extent of anti-coagulant effect is poorly correlated to plasma concentrations of heparin.  Therefore, the end result (bleeding) is a more useful parameter by which to monitor heparin efficacy or overdose.  The laboratory test of choice to monitor heparin therapy is the (activated) partial thromboplastin time (PTT or aPTT).  This test measures specifically the intrinsic pathway of coagulation and is dependent upon the activity of factors I, II, V, VIII, IX, X, XI, and XII.  The normal PTT ranges from 50-80 sec.  Effective heparin therapy should produce a PTT that is 1.5-2.5 times this normal value (PTT below this would indicate ineffective dosing while values greater than this range would indicate heparin overdose).

Treatment of Heparin Overdose
The antidote for heparin overdose/toxicity is another animal-derived compound.  Protamine is purified from piscine (fish) sperm.  It will bind to and inactivate heparin, thus preventing its binding to AT-III.  However, protamine alone possesses mild anti-coagulant activity.  Therefore, only the minimum dose of protamine should be administered to inactivate heparin.  This is generally 1 mg of protamine for every 100 units of heparin in the patient (as determined by plasma analysis).

Mechanism of Action
The exact mechanism of action of hirudin is not known.  It acts independently of AT-III.  It may, unlike heparin, inhibit thrombin that is already bound to fibrinogen in a forming clot, thus inhibiting fibrin and subsequent clot formation.  It appears that one molecule of lepirudin (hirudin) binds to one molecule of thrombin, thus inhibiting its thrombogenic activity.

Side Effects
The side effect profile of lepirudin is similar to that of heparin, with excess bleeding the most common adverse reaction.   Antibody formation may lead to either allergic reactions and/or loss of efficacy.  The drug is eliminated renally, therefore dose must be adjusted in patients with renal failure.

Monitoring
Hirudin therapy is monitored in the same manner as heparin, by PTT.

Mechanism of Action -- Argatroban acts as a reversible inhibitor of thrombin, preventing thrombin-induced conversion of fibrinogen to fibrin.  Similar to lepirudin, its actions are independent of  AT-III.  It will inhibit both free and fibrinogen-bound thrombin.

Side Effects -- The major side effect noted for argatroban is bleeding.  Other reported side effects include hypotension, diarrhoea, fever, and nausea and vomiting.  The risk of allergic reaction and/or antibody formation is very low.

Monitoring -- Therapy should be monitored by aPTT (desired aPTT is 1.5 to 3 times baseline, but less than 100 sec.).  Note that if warfarin therapy is initiated while the patient is undergoing argatroban therapy, the INR will initially increase.  However, when argatroban is discontinued, the INR levels will decline.

Indications -- Argatroban is indicated for the treatment or prevention of thrombosis in patients with heparin-induced thrombocytopaenia.

Oral Anti-Coagulants -- Warfarin, Dicumarol, Phenprocoumin
These agents were discovered as a result of haemorrhagic disease in cattle that had ingested spoiled sweet clover.  It was discovered that the fungi involved in the spoilage produced an anti-coagulant compound, bishydroxycoumarin, later synthesised as dicumarol.  Dicumarol is rarely used in medicine today.  Phenprocoumin has similar effects as warfarin with an extended (up to six days) half-life.  Warfarin is the most widely used of the agents.

Mechanism of Action -- These agents inhibit the formation of vitamin K-dependent clotting factors II, VII, IX, and X.  The final step in the synthesis of these factors is the addition of a carboxyl group (i.e. the conversion of descarboxyprothrombin to prothrombin, for factor II).  This addition is mediated by vitamin K which is oxidised to the dione in the reaction, as illustrated below.

Vitamin K is normally regenerated (reduced from the dione form to the dihydroxy form via an epoxide intermediary) by the enzyme vitamin K epoxide reductase.  Warfarin inhibits the action of this enzyme, thereby preventing the regeneration of active vitamin K.  This, in turn, prevents the further formation of the four dependent factors.  The onset of action for warfarin is typically 8-12 hours (in part due to the long half-life and in part due to the time taken for depletion of factors already formed and active vitamin K).  This onset may be shortened by the administration of a loading dose of warfarin.

Toxicity -- Warfarin overdose is evident mainly as excessive bleeding (which may first be noticed as bleeding gums with brushing or easy bruising).  Additionally, warfarin may produce teratogenic effects that affect primarily bone formation (the exact role of vitamin K in foetal bone development is not known).  Warfarin overdose is treated with vitamin K (dione form), which will compete with the drug for binding to the epoxide reductase.

Monitoring of Efficacy -- The most commonly used parameter to determine the efficacy of warfarin therapy is the INR or International Normalised Ratio.  This represents a standardised prothrombin time (PT) which is specific for the extrinsic pathway of coagulation (sensitive to factors I, II, V, VII, and X).  INR = (PT_pt/PT_ref)^ISI
where PT_pt = patient prothrombin time, PT_ref = specific laboratory reference PT, and ISI = international sensitivity index.  This normalisation is approximately equal to a PT of 1.6 for most American laboratories.  Effective warfarin therapy should produce an INR that is 2.5-3.5 times the normal.

Drug Interactions -- Warfarin is the subject of numerous drug interactions that could interfere with proper therapy and patient health.  Drugs that increase the effect of warfarin include those which displace it from plasma protein binding sites (NSAIDs, phenytoin), drugs which inhibit its metabolism (cimetidine) (NOTE both of the preceding interactions are pharmacokinetic in nature) and drugs that also prolong bleeding time (such as ASA, NSAIDs -- a pharmacodynamic interaction).  Other drugs that may increase the effect of warfarin include amiodarone, disulfiram, and 3rd generation cephalosporins.  Co-administration of these drugs may require a reduction in the dose of warfarin.  Interactions that decrease the efficacy of warfarin (hence, increase the risk of clot formation) include those agents that induce its metabolism (barbiturates), those that inhibit its absorption (cholestyramine, rifampin) and those that hasten its excretion (diuretics).

Fibrinolytics -- All of these agents result in the conversion of plasminogen to plasmin.  Therapy with any of these agents is expensive; however, the cost increases with the relative degree of specificity.  They are used primarily in the prevention of multiple pulmonary emboli, deep vein thrombosis, and post-MI thrombo-emboli.  The primary side effect which they all share is enhanced bleeding (more often resulting from inadvertent overdose).

Streptokinase -- This agent is synthesised from the Streptococcus bacteria.  It is a protein that binds to circulating plasminogen and catalyses its conversion to plasmin, which will degrade both fibrinogen and fibrin, as discussed previously.  The action of streptokinase is relatively non-specific.  It will convert to plasmin all plasminogen to which it binds.  This decreases its efficacy somewhat (the molecule is wasted on a circulating plasminogen that is not near the targeted clot) and may cause undesirable bleeding (by enhancing clot breakdown where a clot may be needed).  Streptokinase may produce fever, allergic reactions, and decreased efficacy in patients who have anti-streptococcus antibodies.

Urokinase -- This enzyme (derived from urine) is a protease that actually converts plasminogen to plasmin.  It, too, is relatively non-specific in its actions, converting both circulating plasminogen and plasminogen associated with the clot formation.

Sauroplase, a prourokinase, is in clinical trials and appears to be somewhat more selective in its activity.

Anistreplase -- This agent, which is a complex of plasminogen and streptokinase, is relatively more selective in its actions.  Since it is a complex, it converts only those molecules of plasminogen to which the streptokinase is bound (it has no effect on endogenous plasminogen).  Since plasminogen/plasmin has greater affinity for forming clots, anistreplase shows some affinity for forming clots.  This action results in less body-wide clot prevention and relative selectivity.

Tissue Plasminogen Activator (tPA, Alteplase, Reteplase) -- these drugs are the most selective of the fibrinolytics.  They preferentially activate plasminogen that is bound to fibrin.  They have no action on circulating plasminogen.  This relative selectivity provides greater efficacy and fewer adverse effects than the relatively non-specific fibrinolytics.

Anti-Thrombotics -- These agents interfere with platelet aggregation, to slow clot formation.

ASA -- inhibits TXA₂, which increases platelet aggregation.  This drug has already been discussed (refer to previous notes).  The standard dose of ASA as an anti-thrombotic is 1-5 gr 3 times weekly up to q.d.

Dipyridamole -- This agent exerts anti-platelet effects by 1) inhibition of phosphodiesterase, 2) blocking adenosine re-uptake, and/or 3) alterations in ADP function.  Regardless of the mechanism, it has exhibited some effect in preventing platelet aggregation.  This drug has also been discussed previously.

Prostacyclin (PGI₂) -- This eicosanoid is synthesised and released along with TXA₂ as a controller or "brake" to prevent excessive platelet aggregation.  It may sometime be administered post-MI to prevent excessive clot formation.

Ticlopidine

Pharmacologic Response -- This drug prevents binding of fibrinogen to activated platelets.  Fibrinogen binds specifically to the IIb/IIIa glycoprotein on platelet membranes.

Mechanism of Action -- The exact mechanism of action of ticlopidine is unclear, but it may either change the conformation of gp IIb/IIIa, decreasing its binding affinity (perhaps by altering ADP-dependent processes) or it may directly block the binding site, preventing fibrinogen binding.

Adverse Effects -- Nausea/vomiting/diarrhoea (20%); excessive bleeding (10%), leukopenia (1%) .  Additionally, a new potentially dangerous adverse reaction has recently been reported for ticlopidine.  It is reported to produce a thrombotic thrombocytopeanic purpurea (very similar to heparin-induced thrombi formation associated with thrombocytopaenia) in a small portion (1% or less) of patients.

Clinical Uses -- Ticlopidine is used primarily for the prevention of thrombosis in cerebral and/or coronary vascular disease, thus reducing the risk of stroke, angina, or myocardial infarction.  It is especially useful in those patients who do not tolerate aspirin (due to allergy, hypersensitivity, or gastric ulcer disease), however it may also work well with ASA, due to their divergent mechanisms.
 

Clopidogrel (Plavix®) -- clopidogrel is a relatively small MW compound available as an oral tablet. Clopidogrel specifically inhibits the binding of ADP to its platelet receptor, thus inhibiting ADP-induced activation of the glycoprotein IIb/IIIa complex and subsequent platelet aggregation.  This inhibition is non-competitive and irreversible.  The side effect profile of  clopidogrel is similar to and not significantly different than than of aspirin.
 
Abciximab -- abciximab is an antibody that is specific for the IIb/IIIa gp.

Eptifibatide (Integrilin® ) is a peptide that has high binding affinity for the IIb/IIIa gp.  Both drugs block the glycoprotein, thus preventing fibrinogen binding and subsequent clot formation.

Tirofiban (Aggrastat®) is an agent similar in action to eptifibatide, an anti-thrombotic that inhibits fibrin binding with platelets by preventing its binding with the IIb/IIIa receptor on the platelet membrane.  Tirofiban is administered intravenously.

Anagrelide

Mechanism of Action -- Anagrelide inhibits platelet counts by an unknown mechanism, apparently by inhibiting the synthesis of platelets.  Other cells are not effected to a clinically signficant level, although red cells may also slightly decrease in number.    Higher doses of anagrelide may also inhibit platelet cell function, specifically inhibiting aggregation.  This occurs as a result of inhibition of phosphodiesterase (similar to dipyridamole) and inhibition of ADP and collegen-induced aggregation (similar to clopidogrel).

Side Effects -- Most common side effects include headache, diarrhoea, oedema, palpitations, and abdominal pain.  These are generally mild and transient.  Rare, but life-threatening, adverse reactions involve the cardiovascular system and include failure, heart block, arrhythmias, pericarditis, pulmonary hypertention and pulmonary fibrosis.

Indications and Use -- Anagrelide is used in the treatment of myeloproliferative disorders that result in excessive platelet formation, including polycythemia vera.
 

Vitamin K

Vitamin K is available in three forms:
Vitamin K₁ -- phytonadione -- the active form of vitamin K as found in food sources.  This is the most commonly prescribed/dispensed form of vitamin K.

Vitamin K₂ -- menaquinone -- this is the form of vitamin K that is obtained by intestinal microflora.

Vitamin K₃ -- menadione

The action of all three forms are the same and have been discussed in the section of warfarin.  Vitamin K is used primarily for warfarin overdose.  It may also be given as a supplement in vitamin K deficiency (which is relatively rare but may occur in chronic hospital patients on enteral/parenteral nutrition).  It is sometimes used as an adjunct in some forms of haemophilia.

Inhibitors of Fibrinolysis -- Aminocaproic Acid, Traxenamic Acid
These drugs inactivate the conversion of plasminogen to plasmin, thereby allowing clots to form undisturbed.  They are used as adjuncts in haemophilia, in overdoses of fibrinolytic drugs, and to prevent re-bleeding in patients with intracranial haemorrhage.  They have also recently been suggested for use in patients on warfarin prior to dental visits.  The adverse effects most commonly seen with these agents include thrombosis, hypotension, myopathy, and GI disturbances.

Apoprotin -- This drug has already been discussed as a bradykinin/kallikrein inhibitor.  In addition to its ability to inhibit kininogen activation, it also inhibits free plasmin-induced thrombolysis.  In this regard it is a non-specific serine protease inhibitor (serpin) that decreases bleeding and perhaps decreases clot destruction.  It is used primarily to prevent bleeding in open heart surgery and coronary bypass grafts.

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