Month: March 2017

Methods for Assessment of Liver Injury and Liver Function

March 23, 2017 Uncategorized No comments , , , , , , ,

Saving Private Ryan

Blood tests remain a mainstay for the diagnosis of a patient with suspected liver disease or to stage patients with known liver disease. In general, the goal of such tests is to detect the release of products that are characteristic of the liver into the bloodstream, thereby indicating liver cell injury. Similarly, accumulation of molecules normally excreted via the biliary system is indicative of a failure in liver function, although further tests are necessary to assess the site at which the defect is occurring.

Two enzymes referred to as transaminases are easily measured in the serum, and are sensitive markers of liver cell injury. Alanine aminotransferease (ALT) is produced by hepatocytes, and when these cells are injured, its circulating levels are increased. Aspartate aminotransferase (AST) is similarly increased in the setting of hepatocellular injury, although it is less specific for liver disease because it is also produced by other tissues. For example, AST levels are increased following cardiac injury, such as a myocardial infarction. Nevertheless, measurement of AST remains useful in the setting of clinical symptoms consistent with liver disease, particularly because it appears to be disproportionately elevated in patients whose liver injury is related to alcohol abuse. Measurements of ALT and AST over time are also used to assess the progress of established liver disease. It is important to remember, however, that elevations in transaminases can only occur if these is ongoing hepatocellular death. In cirrhosis, where large portions of the liver may be replaced by fibrous tissue, little new cell injury may be occurring and thus ALT and AST levels may not be elevated.

Two other enzymes are useful markers of injury to the biliary system. Alkaline phosphatase, while not specific to the liver (being produced also by bone, the intestine and placenta), is expressed as a membrane protein in the canaliculus. In the setting of localized obstruction to the biliary tree, alkaline phosphatase levels in the serum are increased. Similarly, gamma glutamyltranspeptidase (GGT) is localized predominantly to the apical membrane of cholangiocytes, although some is also expressed in the bile canaliculi. Serum elevations in GGT are therefore largely reflective of cholangiocyte injury.

The foregoing tests, while often referred to as “liver function” tests, are not strictly measures of actual hepatic function. To assess whether liver function is impaired, other tests are needed. One such test that is very important clinically is the measurement of bilirubin. Accumulation of bilirubin in the circulation indicates cholestasis, which can result from injury to either hepatocytes or cholangiocytes, or obstruction within the biliary system. Based on the discussion of the synthetic functions of the liver, it should be easy to understand that liver function can also be assessed by measuring levels of its various products in the circulation. The most useful tests are to measure serum albumin, and a blood clotting parameter, the prothrombin time. Tests of coagulation are not specific for liver disease and thus must be interpreted in the context of other findings. Patients with suspected liver disease are also often evaluated for serum glucose and ammonia levels, since hypoglycemia and hyperammonemia are major problems in the setting of liver failure.

Biopsies and imaging tests also play a major role in the evaluation of liver disease. Liver biopsies, which are usually obtained by inserting a needle into the liver percutaneously, can be used to evaluate the extent of liver fibrosis, or to search for evidence of rejection in a previously transplanted liver. A widely applied imaging technique is referred to as ERCP, which stands for endoscopic retrograde cholangiopancreatography. In this procedure, a special endoscope is introduced into the duodenum via the mouth, and used to insert a small tube through the sphincter of Oddi, through which contrast medium is injected. Subsequent X-rays can then visualize the drainage routes from the biliary system and pancreas, which permits the diagnosis of obstructions or strictures. Other imaging modalities, such as magnetic resonance imaging, are also assuming an increasing role in assessing liver disease and the architecture of the biliary system.

The prognosis of end-stage liver disease, and by extension the urgency of transplantation, has commonly been assessed by calculating the so-called Child-Pugh score, which takes into account 5 measures of hepatic function. The Child-Pugh score was originally designed as a predictor of surgical mortality in liver disease patients. More recently, the Child-Pugh score has been supplemented by the Model for End-Stage Liver Disease (MELD) score, which weights serum bilirubin, creatinine, and prothrombin time to predict survival. Patients awaiting liver transplantation in the United States are now prioritized on the basis of their MELD score, which results in the allocation of organs first to those who are sickest. This has reduced waiting-list mortality without impairing post-transplant outcomes.

Drug Interactions to Warfarin

March 22, 2017 Drug Informatics, Drug Interactions, Pharmacodynamics, Pharmacokinetics No comments , , , , , , ,

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Drugs may interact with warfarin sodium through pharmacodynamic or pharmacokinetic mechanisms. Pharmacodynamic mechanisms for drug interactions with warfarin sodium are synergism (impaired hemostasis, reduced clotting factor synthesis), competitive antagonism (vitamin K), and alteration of the physiologic control loop for vitamin K metabolism (hereditary resistance). Pharmacokinetic mechanisms for drug interactions with warfarin sodium are mainly enzyme induction, enzyme inhibition, and reduced plasma protein binding. It is important to note that some drugs may interact by more than one mechanism.

Pharmacodynamic:

  • Synergism
  • Competitive antagonism
  • Alteration of vitamin K cycle and metabolism

Pharmacokinetic:

  • Enzyme induction
  • Enzyme inhibition
  • Reduced plasma protein binding

CYP450 Interactions

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CYP450 isozymes involved in the metabolism of warfarin include CYP2C9, 2C19, 2C8, 2C18, 1A2, and 3A4. The more potent warfarin S-enantiomer is metabolized by CYP2C9 while the R-enantiomer is metabolized by CYP1A2 and 3A4.

  • Inhibitors of CYP2C9, 1A2, and/or 3A4 have the potential to increase the effect (increase INR) of warfarin by increasing the exposure of warfarin.
  • Inducers of CYP2C9, 1A2, and/or 3A4 have the potential to decrease the effect (decrease INR) of warfarin by decreasing the exposure of warfarin.

Examples of inhibitors and inducers of CYP2C9, 1A2, and 3A4 are below in Table 2; however, this list should not be considered all-inclusive.

Drugs that Increase Bleeding Risk

Examples of drugs known to increase the risk of bleeding are presented in Table 3. Because bleeding risk is increased when these drugs are used concomitantly with warfarin, closely monitor patients receiving any such drug with warfarin.

Antibiotics and Antifungals

There have been reports of changes in INR in patients taking warfarin and antibiotics or antifungals, but clinical pharmacokinetic studies have not shown consistent effects of these agents on plasma concentrations of warfarin.

Botanical (Herbal) Products and Foods

More frequent INR monitoring should be performed when starting or stopping botanicals.

Few adequate, well-controlled studies evaluating the potential for metabolic and/or pharmacologic interactions between botanicals and warfarin sodium exist. Due to a lack of manufacturing standardization with botanical medicinal preparations, the amount of active ingredients may vary. This could further confound the ability to assess potential interactions and effects on anticoagulation.

Some botanicals may cause bleeding events when taken alone and may have anticoagulant, antiplatelet, and/or fibrinolytic properties. These effects would be expected to be additive to the anticoagulant effects of warfarin sodium. Conversely, some botanicals may decrease the effects of warfarin sodium. Some botanicals and foods can interact with warfarin sodium through CYP450 interactions (e.g., echinacea, grapefruit juice, ginkgo, goldenseal, St. John’s wort).

The amount of vitamin K in food may affect therapy with warfarin sodium. Advise patients taking warfarin sodium to eat a normal, balanced diet maintaining a consistent amount of vitamin K. Patients taking warfarin sodium should avoid drastic changes in dietary habits, such as eating large amounts of green leafy vegetables.

Overview of the Hemostasis System

March 15, 2017 Hematology, Physiology and Pathophysiology No comments , , , , , , , , , , , , , , , , , , , , ,

Procoagulant Pathways

Two procoagulant pathway have been identified which converge at the “intrinsic” (accessory) fXase (fIXa*fVIIIa) complex. The contact or “intrinsic” pathway is activated by the interaction of blood with a foreign surface. This pathway is activated by the factor XIIa-high-molecular-weight kininogen (HMWK)-prekallikrein complex in association with foreign surfaces including glass, dextran sulfate, or kaolin. The complex catalyst activates factor XI leading to the factor XIa-HMWK complex which activates fIX to fIXa. The “intrinsic” fXase rapidly cleaves fX to fXa.

Factor Xa is directly but less effectively produced by the “extrinsic” fXase, which is composed of plasma-derived fVIIa and tissue factor and expressed when the latter is exposed to blood. The “extrinsic” fXase also activates fIX to fIXa as the reaction progresses, with suppression of the quaternary complex occurring by TFPI. The “intrinsic” fXase complex is kinetically superior and ultimately produces the majority of fXa.

Since bleeding pathology is not ordinarily associated with defects of the initiation protein complexes of the contact pathway, most investigations conclude that the “intrinsic” pathway is not the primary provider for hemostasis. The “intrinsic” pathway, however, has been implicated in some forms of thrombosis. The primary (“extrinsic”) and accessory (“intrinsic”) pathways, initiated by independent routes, both lead to the activation of factor IX and converge at the “intrinsic” fXase complex. Regardless of the path, the outcome is the formation of the prothrombinase complex and thrombin generation. Each reaction of the primary pathway of coagulation involves the vitamin K-dependent zymogens and serine proteases, cofactor proteins, and Ca2+ ions assembled on membranes. The complexes display reaction rates 10^5 to 10^9 times greater than the respective serine proteases alone.

Clinical laboratory tests differentiate between the pathways. The activated partial thromboplastin time (aPTT) initiates coagulation through the accessory pathway, whereas the prothrombin time (PT) assay initiates coagulation through the primary pathway. The designations of primary and accessory pathways are based on clinical evidence of bleeding diseases. Deficiencies of proteins associated with the “intrinsic” or accessory pathway (factor XII, prekallikrein, and HMWK) exist but are not associated with abnormal bleeding events, even after surgical challenge. However, deficiencies of the protein components of the “extrinsic” or primary pathway (prothrombin and factors V, VII, VIII, IX, and X) can lead to severe bleeding diatheses. Factor XI deficiency may also result in bleeding episodes subsequent to trauma or surgery. The role of the accessory pathway is therefore not clearly understood.

Factor XII, prekallikrein, and HMWK are required for activity of the contact or accessory pathway and deficiencies are reported by the aPTT. Factor XII and prekallirein are zymogens that are activated to sarin proteases, while HMWK is a cofactor. The accessory pathway factors are hypothesized to play a role in disseminated intravascular coagulation (DIC) associated with the systemic inflammatory response syndrome, and may also be involved in the promotion of thrombus stability. The accessory pathway may also be important in cardiopulmonary bypass because of contact between blood components and synthetic surfaces.

The importance of the membrane component in coagulation was initially identified by kinetic studies of the prothrombinase complex. In the absence of the membrane surface, the cofactor (factor Va)-enzyme (factor Xa) interaction is relatively weak, with dissociation constant (Kd) of 800 nmol/L. The factor Va-lipid interaction (Kd = 3 nmol/L) and factor Xa-lipid interaction (Kd = 110 nmol/L) show higher affinity. However, all of the components must be present to generate the high-affinity factor Va-factor Xa-Ca2+ membrane complex, with a Kd of 1 nmol/L. The fully assembled complex is stabilized through factor Va-factor Xa, factor Va-lipid, and factor Xa-lipid interactions. Similar properties have been observed for the fIXa/fVIIIa, TF/fVIIa, and Tm/fIIa complexes.

The primary pathway of coagulation is initiated or triggered by the interaction of circulating factor VIIa with its cofactor tissue factor (TF). In general, the serine proteases associated with hemostasis circulate in their zymogen or inactive forms; however, low levels of circulate in their zymogen or inactive forms; however, low levels of circulating factor VIIa are present in blood. This factor VIIa binds to tissue factor expressed by pathology and initiates the procoagulant response. Free factor VIIa is a poor enzyme with virtually no proteolytic activity, but as a consequence is protected from interacting with the circulating inhibitors in the absence of tissue factor. Tissue factor, an integral membrane protein not normally expressed on vascular cell surfaces, is constitutively expressed on extravascular cellular surfaces and thus becomes exposed upon damage to the endothelial cell layer. Tissue factor is also expressed on peripheral blood cells and endothelial cells stimulated by inflammatory cytokines.

Upon interaction of plasma factor VIIa and the injury/pathology present, tissue factor, the “extrinsic” fXase complex (factor VIIa-tissue factor) is formed and initiates coagulation by activating factor IX and X. Factor IXa forms a complex with its cofactor, factor VIIIa, to generate the “intrinsic” fXase complex, and factor Xa combines with factor Va to form the prothrombinase complex. The factor VIIIa-factor IXa complex more efficiently activates factor X to factor Xa, providing a robust source of the enzyme component (factor Xa) of the prothrombinase complex.

Deficiencies of the “intrinsic” fXase components factor VIIIa (hemophilia A) and factor IXa (hemophilia B) illustrate the significance of the factor IX activation by the fVIIa/TF complex and the enhanced rate of factor X activation by the intrinsic fXase and the inhibition of the extrinsic fXase by TFPI. Hemophilia A and B are detected using the aPTT. It would appear activation of factor X by the extrinsic fXase should compensate for the lack of factor activation by the intrinsic fXase in hemophilias A and B. However, this compensatory mechanism only occurs during clinical administration of supraphysiologic concentrations of recombinant factor VIIa during replacement therapy for hemophilia with inhibitors. The natural physiologic levels of factor VIIa are not able to provide sufficient levels of factor Xa to support normal coagulation. Factor Xa generation is suppressed to approximately one-half the level observed when factor X is the only substrate presented.

Factor IX, not factor X, appears to be the preferred substrate of the extrinsic fXase. In addition, factor IXalpha, the intermediate species in factor IX activation, is generated more rapidly in the presence of factor X. Factor IXalpha activation to the final product factor IXa occurs at a higher rate than factor IX activation, thereby providing a burst of factor IXa to form the intrinsic fXase complex. The low level of factor Xa generated by the tissue factor-factor VIIa complex most likely functions in the activation of factor IX. A model of extrinsic fXase behavior suggests that factor IX is converted to factor IXalpha by the extrinsic fXase or factor Xa-phospholipid complex. Factor IXalpha is then rapidly converted to factor IXa by the extrinsic fXase. The factor VIIIa-factor IXa complex subsequently activates the major fraction of factor X to factor Xa and provides the enzyme component for the prothrombinase complex. Measurements of second-order rate constants for factor Xa generation by the intrinsic and extrinsic fXase complexes also support this model. The rate of factor Xa generation by the tissue factor-factor VIIa complex is 1/50th the rate of factor Xa generation by the factor VIIIa-factor IXa complex. Both complexes thus have distinct roles in the procoagulant response.

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Accessory Pathway

The procoagulant proteins that make up the intrinsic or accessory pathway consist of factor XII, plasma prekallikrein, HMWK, and factor XI. These proteins are responsible for the contact activation of blood coagulation. The physiologic role of the intrinsic pathway is not clearly understood, but does not appear to be essential for hemostasis because individuals deficient in factor XII, plasma prekallirein, or HMWK do not manifest abnormal bleeding. Factor XI appears to play a more prominent role, unrelated to its activities in the intrinsic pathway, after activation by thrombin.

Update on March 15 2017

Hemostatic System

The major components of the hemostatic system are the vascular endothelium, platelets, and the coagulation and fibrinolytic systems.

Vascular Endothelium

A monolayer of endothelial cells lines the intimal surface of the circulatory tree and separates the blood from the prothrombotic sub endothelial components of the vessel wall. As such, the vascular endothelium encompasses about 10^13 cells and covers a vast surface area. Rather than serving as a static barrier, the healthy vascular endothelium is a dynamic organ that actively regulates hemostasis by inhibiting platelets, suppressing coagulation, promoting fibrinolysis, and modulating vascular tone and permeability.

Platelet Inhibition

Endothelial cells synthesize prostacyclin and nitric oxide and release them into the blood.These agents not only serve as potent vasodilators but also inhibit platelet activation and subsequent aggregation by stimulating adenylate cyclase and increasing intracellular levels of cyclic adenosine monophosphate (cAMP). In addition, endothelial cells express (CD39) on their surface, a membrane-associated ecto-adenosine diphosphatase (ADPase). By degrading ADP, which is a platelet agonist, CD39 attenuates platelet activation.

  • Prostacyclin
  • Nitric oxide
  • CD39

Anticoagulant Activity

Intact endothelial cells play an essential part in the regulation of thrombin generation through a variety of mechanisms. Endothelial cells produce heparan sulfate proteoglycans, which bind circulating antithrombin and accelerate the rate at which it inhibits thrombin and other coagulation enzymes. Tissue factor pathway inhibitor (TFPI), a naturally occurring inhibitor of coagulation, binds heparan sulfate on the endothelia cell surface. Administration of heparin or low-molecular-weight heparin (LMWH) displaces glycosaminoglycan-bound TFPI from the vascular endothelium, and released TFPI may contribute to the antithrombotic activity of these drugs.

Endothelial cells regulate thrombin generation by expressing thrombomodulin and endothelial cell protein C receptor (EPCR) on their surfaces. Thrombomodulin binds thrombin and alters this enzyme’s substrate specificity such that it no longer acts as a procoagulant but becomes a potent activator of protein C. Activated protein C serves as an anticoagulant by degrading and inactivating activated factor V (FVa) and factor VIII (FVIIIa), key cofactors involved in thrombin generation. Protein S acts as a cofactor in this reaction, and EPCR enhances this pathway by binding protein C and presenting it to the thrombin-thrombomodulin complex for activation. In addition to its role as an anticoagulant, activated protein C also regulates inflammation and preserves the barrier function of the endothelium.

  • Heparan
  • TFPI
  • Thrombomodulin
  • EPCR

Fibrinolytic Activity

The vascular endothelium promotes fibrinolysis by synthesizing and releasing tissue-type and urokinase-type plasminogen activator (t-PA and u-PA, respectively), which initiate fibrinolysis by converting plasminogen to plasmin. Endothelial cells in most vascular beds synthesize t-PA constitutively. In contrast, perturbed endothelial cells produce u-PA in the settings of inflammation and would repair.

Endothelial cell also produce type 1 plasminogen activator inhibitor 1 (PAI-1), the major regulator of both t-PA and u-PA. Therefore, net fibrinolytic activity depends on the dynamic balance between the release of plasminogen activators and PAI-1.  Fibrinolysis localizes to the endothelial cell surface because these cells express annexin II, a coreceptor for plasminogen and t-PA that promotes their interaction. Therefore, healthy vessels actively resist thrombosis and help maintain platelets in quiescent state.

  • t-PA
  • u-PA
  • PAI-1
  • Annexin II

Vascular Tone and Permeability

In addition to synthesizing potent vasodilators, such as prostacyclin and nitric oxide, endothelial cells also produce a group of counter-regulatory peptides known as endothelins that induce vasoconstriction. Endothelial cell permeability is influenced by the connections that join endothelial cells to their neighbors. Macromolecules traverse the endothelium via patent intercellular junctions, by endocytosis, or through transendothelial pores. Vasodilatation, servers thrombocytopenia, and high doses of heparin can increase endothelial permeability, which may contribute to bleeding. Activated protein C may also contribute to the barrier function of the endothelium.

Platelets

Platelets are anucleate particles released into the circulation after fragmentation of bone marrow megakaryocytes. Because they are anucleate, platelets have limited capacity to synthesize proteins. Thrombopoietin, a glycoprotein synthesized in the liver and kidneys, regulates megakaryocytic proliferation and maturation as well as platelet production. After they enter the circulation, platelet have a life span of 7 to 10 days.

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Damage to the intimal lining of the vessel exposes the underlying subendothelial matrix. Platelets home to sites of vascular disruption and adhere to the exposed matrix proteins. Adherent platelets undergo activation and not only release substances that recruit additional platelets to the site of injury but also promote thrombin generation and subsequent fibrin formation. A potent platelet agonist, thrombin amplifies platelet recruitment and activation. Activated platelets then aggregate to form a plug that seals the leak in the vasculature. An understanding of the steps in these highly integrated processes helps pinpoint the sites of action of the antiplatelet drugs and rationalizes the utility of anticoagulants for the treatment of arterial thrombosis and venous thrombosis.

Adhesion

Platelets adhere to exposed collagen and von Willebrand factor (vWF) and form a monolayer that supports and promotes thrombin generation and subsequent fibrin formation. These events depend on constitutively expressed receptors on the platelet surface, alpha2beta1 and glycoprotein (GP) VI, which bind collagen, and GPIbalpha and GPIIb/IIIa (alphaIIbbeta3), which bind vWF. The platelet surface is crowded with receptors, but those involved in adhesion are the most abundant: every platelet has about 40,000 to 80,000 copies of GPIIb/IIIa and 25,000 copies of GPIbalpha. Receptors cluster in cholesterol-enriched subdomains, which render them more mobile, thereby increasing the efficiency of platelet adhesion and subsequent activation.

  • Platelet receptors: alpha2beta1, GP VI, GPIbalpha, GPIIb/IIIa
  • Platelet receptors’ ligands: collagen (alpha2beta1, GP VI), vWF (GPIbalpha, GPIIb/IIIa)

Under low shear conditions, collagen can capture and activate platelets on its own. Captured platelets undergo cytoskeletal reorganization that causes them to flatten and to adhere more closely to the damaged vessel wall. Under high shear conditions, however, collagen and vWF must act in concert to support optimal platelet adhesion and activation. vWF synthesized by endothelial cells and megakaryocytes assembles into multimers that range from 550 to more than 10,000 kDa. When released from storage in the Weibel-Palade bodies of endothelial cells or the alpha-granules of platelets, most of the vWF enters the circulation, but the vWF released from the abluminal surface of endothelial cells accumulates in the subendothelial matrix, where it binds collagen via its A3 domain. This surface-immobilized vWF can simultaneously bind platelets via its A1 domain. In contrast, circulating vWF does not react with unstimulated platelets. This difference in reactivity likely reflects vWF conformation; whereas circulating vWF is in a coiled conformation that prevents access of its platelet-binding domain to vWF receptors on the platelet surface, immobilized vWF assumes an elongated shape that exposes its A1 domain. In this extended conformation, large vWF multimers serve as the molecular glue that tethers platelets to the damaged vessel wall with sufficient strength to withstand higher shear forces. Large vWF multimers provide additional binding sites for collagen and heighten platelet adhesion because platelets have more vWF results in platelet activation, the next step in platelet plug formation.

  • Low shear conditions
  • High shear conditions
  • Free vWF
  • Surface-immobilized vWF
  • Interaction: vWF and collagen

Activation and Secretion

Adhesion to collagen and vWF initiates signaling pathways that result in platelet activation. These pathway induce cyclooxyrgenase-1 (COX-1)-dependent synthesis and release of thromboxane A2, and trigger the release of adenosine diphosphate (ADP) from storage granules. Thromboxane A2 is a potent vasoconstrictor, and similar to ADP, locally activates ambient platelets and recruits them to the site of injury. This process results in expansion of the platelet plug. To activate platelets, thromboxane A2 and ADP must bind to their respective receptors on the platelet membrane. The thromboxane receptor (TP) is a G-protein coupled receptor that is found on platelets and on the endothelium, which explains why thromboxane A2 induces vasoconstriction as well as platelet activation. ADP interacts with a family of G protein-coupled receptors on the platelet membrane. Most important of these is P2Y12, which is the target of the theienopyridines, but P2Y1 also contributes to ADP-induced platelet activation, and maximal ADP-induced platelet activation requires activation of both receptors. A third ADP receptor, P2X1, is an adenosine triphosphate (ATP)-gated calcium channel. Platelet storage granules contain ATP as well as ADP; ATP released during the platelet activation process may contribute to the platelet recruitment process in a P2X1-dependent fashion.

  • TXA2
  • Thromboxane receptor
  • ADP
  • ADP receptors (P2Y12, P2Y1, P2X1)
  • Pathway: collagen and/or vWF > thromboxane A2 release, and ADP release

Although TP and the various ADP receptors signal through different pathways, they all trigger an increase in the intracellular calcium concentration in platelets. This in turn induces shape change via cytoskeletal rearrangement, granule mobilization and release, and subsequent platelet aggregation. Activated platelets promote coagulation by expressing phosphatidylserine on their surfaces, and anionic phospholipid that supports assembly of coagulation factor complexes. After being assembled, these clotting factor complexes trigger a burst of thrombin generation and subsequent fibrin formation. In addition to converting fibrinogen to fibrin, thrombin amplifies platelet recruitment and activation and promotes expansion of the platelet plug. Thrombin binds to protease-activated receptors types 1 and 4 (PAR1 and PAR4, respectively) on the platelet surface and cleaves their extended amino-termini, thereby generating new amino-termini that serve as tethered ligands that bind and activate the receptors. Whereas low concentrations of thrombin cleave PAR1, PAR4 cleavage requires higher thrombin concentrations. Cleavage of either receptor triggers platelet activation.

  • Pathway: thrombin (ligand), and PAR1 and PAR4 (receptors)

In addition to providing a surface on which clotting factors assemble, activated platelets also promote fibrin formation and subsequent stabilization by releasing factor V, factor XI, fibrinogen, and factor XIII. Thus, there is coordinated activation of platelets and coagulation, and the fibrin network that results from thrombin action helps anchor the platelet aggregates at the site of injury. Activated platelets also release adhesive proteins, such as vWF, thrombospondin, and fibronectin, which may augment platelet adhesion at sites of injury, as well as growth factors, such as platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGFbeta), which promote wound healing. Platelet aggregation is the final step in the formation of the platelet plug.

  • Secretion: platelet derived factor V, factor XI, fibrinogen, and factor XIII, vWF

Aggregation

Platelet aggregation links platelets to each other to form clumps. GPIIb/IIIa mediates these platelet-to-platelet linkages. On inactivated platelets, GPIIb/IIIa exhibits minimal affinity for its ligands. Upon platelet activation, GPIIb/IIIa undergoes a conformational transformation, which reflects transmission of inside-out signals from its cytoplasmic domain to its extracellular domain. This transformation enhances the affinity of GPIIb/IIIa for its ligands; fibrinogen; and, under high shear conditions, vWF. Arginine-glycine-aspartic acid (RGD) sequences located on fibrinogen and vWF, as well as platelet-binding lycine-glycine-aspartic acid (KGD) sequence on fibrinogen, mediate their interaction with GPIIb/IIIa. When subjected to high shear, circulating vWF elongates and exposes its platelet-binding domain, which enables its interaction with the conformationally activated GPIIb/IIIa. Divalent fibrinogen and multivalent vWF molecules serve as bridges and bind adjacent platelets together. After being bound to GPIIb/IIIa, fibrinogen and vWF induce outside-inside signals that augment platelet activation and result in the activation of additional GPIIb/IIIa receptors, creating a positive feedback loop. Because GPIIb/IIIa acts as the final effector in platelet aggregation, it is a logical target for potent antiplatelet drugs. Fibrin, the ultimate product of the coagulation system, tethers the platelet aggregates together and anchors them to the site of injury.

  • Platelet (activated) receptors: GPIIb/IIIb (fibrinogen, vWF)

Coagulation

Coagulation results in the generation of thrombin, which converts soluble fibrinogen to fibrin. Coagulation occurs through the action of discrete enzyme complexes, which are composed of a vitamin K-dependent enzyme and a non-enzyme cofactor, and assemble on anionic phospholipid membranes in a calcium-dependent fashion. Each enzyme complex activates a vitamin K-dependent substrate that becomes the enzyme component of the subsequent complex. Together, these complexes generate a small amount of thrombin, which amplifies its own generation by activating the non-enzyme cofactors and platelets, which then provide an anionic surface on which the complexes assemble. The three enzyme complexes involved in thrombin generation are extrinsic tenase, intrinsic tenase, and prothrombinase. Although extrinsic tenase initiates the system under most circumstances, the contact system also plays a role in some situations.

Extrinsic Tenase (FVIIa-TF complex)

This complex forms upon exposure of tissue factor-expressing cells to the blood. Tissue factor exposure occurs after atherosclerotic plaque rupture because the core of the plaque is rich in cells that express tissue factor. Denuding injury to the vessel wall also exposes tissue factor constitutively expressed by subendothelial fibroblasts and smooth muscle cells. In addition to cells in the vessel wall, circulating monocytes and monocyte-derived microparticles (small membrane fragments) also provide a source of tissue factor. When tissue factor-bearing monocytes or microparticles bind to platelets or other leukocytes and their plasma membranes fuse, tissue factor transfer occurs. By binding to adhesion molecules expressed on activated endothelial cells or to P-selectin on activated platelets, these tissue factor-bearing cells or microparticles can initiate or augment coagulation. This phenomenon likely explains how venous thrombi develop in the absence of obvious vessel wall injury.

Tissue factor is an integral membrane protein that serves as a receptor for factor VIIa. The blood contains trace amounts of factor VIIa, which has negligible activity in the absence of tissue factor. With tissue factor exposure on anionic cell surfaces, factor VIIa binds in a calcium-dependent fashion to form the extrinsic tenase complex, which is a potent activator of factor IX and X. After being activated, factor IXa and factor Xa serve as the enzyme components of intrinsic tenase and prothrombinase, respectively.

Intrinsic Tenase (FVIIIa-FIXa complex)

Factor IXa binds to factor VIIIa on anionic cell surfaces to form the intrinsic tenase complex. Factor VIII circulates in blood in complex with vWF. Thrombin cleaves factor VIII and releases it from vWF, converting it to its activated form. Activated platelets express binding sites for factor VIIIa. After being bound, factor VIIIa binds factor IXa in a calcium-dependent fashion to form the intrinsic tenase complex, which then activates factor X. The change in catalytic efficiency of factor IXa-mediated activation of factor X that occurs with deletion of individual components of the intrinsic tenase complex highlights their importance. Absence of the membrane or factor VIIIa almost completely abolishes enzymatic activity, and the catalytic efficiency of the complete complex is 10^9-fold greater than that of factor IX alone. Because intrinsic tenase activates factor X at a rate 50- to 100-fold faster than extrinsic tenase, it plays a critical role in the amplification of factor Xa and subsequent thrombin generation.

Prothrombinase (FXa-FVa complex)

Factor Xa binds to factor Va, its activated cofactor, on anionic phospholipid membrane surfaces to form the prothrombinase complex. Activated platelets release factor V from their alpha-granules, and this platelet-derived factor V may play a more important role in hemostasis than its plasma counterpart. Whereas plasma factor V requires thrombin activation to exert its cofactor activity, the partially activated factor V released from platelets already exhibits substantial cofactor activity. Activated platelets express specific factor Va binding sites on their surface, and bound factor Va serves as a receptor for factor Xa. The catalytic efficiency of factor Xa activation of prothrombin increase by 10^9-fold when factor Xa incorporates into the prothrombinase complex. Prothrombin binds to the prothrombinase complex, where it undergoes conversion to thrombin in a reaction that release prothrombin fragment 1.2. Plasma levels of F1.2, therefore, provide a marker of prothrombin activation.

Fibrin Formation

The final effector in coagulation is thrombin. Thrombin converts soluble fibrinogen into insoluble fibrin. Fibrinogen is a dimeric molecule, each half of which is composed of three polypeptide chains – the Aalpha, Bbeta, and gamma chains. Numerous disulfide bonds covalently link the chains together and join the two halves of the fibrinogen molecule. Electron micrographic studies of fibrinogen reveal a trinodular structure with a central E domain flanked by two D domains. Crystal structures show symmetry of design with the central E domain, which contains the amino termini of the fibrinogen chains, joined to the lateral D domains by coiled-coil regions.

Fibrinogen circulates in an inactive form. Thrombin binds to the amino termini of the Aalpha and Bbeta chains of fibrinogen, where it cleaves specific peptide bonds to release fibrinopeptide A and fibrinopeptide B and generates fibrin monomer. Because they are products of thrombin action on fibrinogen, plasma levels of these fibrinopeptides provide an index of thrombin activity. Fibrinopeptide release creates new amino termini that extend as knobs from the E domain of one fibrin monomer and insert into preformed holes in the D domains of other fibrin monomers. This creates long strands known as protofibrils, consisting of fibrin monomers noncovalently linked together in a half-staggered overlapping fashion.

Noncovalently linked fibrin protofibrils are unstable. The stability of the fibrin network is enhanced by platelets and procoagulant cells. Platelets not only bind fibrin via GPIIb/IIIa and promote the formation of a dense fibrin network, but the also release factor XIII. By covalently cross-linking the alpha and gamma chains of adjacent fibrin monomers, factor XIIIa stabilizes the fibrin in a calcium-dependent fashion and renders it relatively resistant to degradation. Factor XIII circulates in blood as a heterodimer consisting of pairs of A and B subunits. The active and calcium binding sites on factor XIII are localized to the A subunit. Platelets contain large amounts of factor XIII in their cytoplasms, but platelet-derived factor XIII consists only of the A subunits. Both plasma and platelet factor XIII are activated by thrombin.

Contact Pathway

Current thinking is that tissue factor exposure represents the sole pathway for activation of coagulation and that the contact system – which includes factor XII, prekallikrein, and high-molecular-weight kininogen – is unimportant for hemostasis because patients deficient in these factors do not have bleeding problems. The physiologic role of factor XI is more difficult to assess because the plasma level of factor XI does not predict the propensity for bleeding. Although the capacity of thrombin to feed back and activate platelet-bound factor XI may explain this phenomenon, platelet-derived factor XI may be more important for hemostasis than circulating factor XI.

We cannot ignore the contact pathway, however, because coronary catheters and other blood-contacting medical devices, such as stents or mechanical valves, likely trigger clotting through this mechanism. Factor XII bound to the surface of catheters or devices undergoes a conformational change that results in it activation. Factor XIIa converts prekallikrein to kallikrein in a reaction accelerated by high-molecular-weight kininogen, and factor XIIa and kallikrein then feedback to activate additional factor XII. Factor XIIa propagates coagulation by activating factor XI.

In addition to its role in device-related thrombosis, the contact pathway may also contribute to the stability of arterial and venous thrombi. DNA and RNA released from damaged cells in atherosclerotic plaques activates factor XII, and mice DNA- or RNA-degrading enzymes exhibit attenuated thrombosis at site of arterial injury. Polyphosphates released from activated platelets also activate factor XII and may provide another stimulus for contact pathway activation. Mice deficient in factor XII or factor XI form small unstable thrombi at sites of arterial or venous damage, suggesting that factor XII and factor XI contribute to thrombogenesis. It is unknown whether the same is true in humans. Patients with unstable angina have increased plasma levels of factor XIa, but it is unclear whether this reflects activation by factor XIIa or by thrombin. Although the contribution of the contact pathway to thrombin generation remains uncertain, the final product of coagulation is fibrin. Hemostasis depends on the dynamic balance between the formation of fibrin and its degradation.

Interactions Between Platelet and Coagulation System (Summary)

Platelet and coagulation system act in synergism to maintain hemostasis.

Platelet for coagulation system

  • Platelet provides membrane where clotting factors reactions could take place quickly
  • Platelet secrets FV (partially activated), FXI, fibrinogen, FXIII, and vWF
  • Polyphosphates released from activated platelets activate FXII

Coagulation system for platelet

  • Thrombin activates platelet via PAR-1 and PAR-4
  • Fibrinogen helps platelet binds adjacent platelet together

Update on Mar 25 2017

Fibrinolytic System

Fibrinolysis initiates when plasminogen activators convert plasminogen to plasmin, which then degrades fibrin into soluble fragments. Blood contains two immunologically and functionally distinct plasminogen activators, t-PA and u-PA. Whereas t-PA mediates intravascular fibrin degradation, u-PA binds to a specific u-PA receptor (u-PAR) on the surface of cells, where it activates cell-bound plasminogen. Consequently, pericellular proteolysis during cell migration and tissue remodeling and repair are the major functions of u-PA.

Regulation of fibrinolysis occurs at two levels. PAI-1, and to a lesser extent, PAI-2, inhibit the plasminogen activators, and alpha2-antiplasmin inhibits plasmin. Endothelial cells synthesize PAI-1, which inhibits both t-PA and u-PA, whereas monocytes and the placenta synthesize PAI-2, which specifically inhibits u-PA. Thrombin-activated fibrinolysis inhibitor (TAFI) also modulates fibrinolysis and provides a link between fibrinolysis and coagulation. Whereas thrombosis can occur if there is impaired activation of the fibrinolytic system, excessive activation leads to bleeding.

  • Plasminogen
  • t-PA
  • u-PA
  • PAI-1
  • PAI-2
  • TAFI

Mechanism of Action of Tissue Plasminogen Activator

Tissue plasminogen activator, a serine protease, contains five discrete domains: a fibronectin-like finger domain, and epidermal growth factor (EGF) domain, two kringle domains, and a protease domain. Synthesized as a single-chain polypeptide, plasmin readily converts t-PA into a two-chain form. Single- and two-chain forms of t-PA convert plasminogen to plasmin. Native Glu-plasminogen is a single-chain polypeptide with a Glue residue at its amino-terminus. Plasmin cleavage at the amino-terminus generates lysine-plasminogen, a truncated form with a Lys residue at its new amino terminus. t-PA cleaves a single peptide bond to convert single-chian glutamine- or lysine-plasminogen into two-chain plasmin composed of a heavy chain containing five kringle domains and a light chain containing the catalytic domain. Because its open conformation exposes the t-PA cleavage site, lysine-plasminogen is a better substrate than glutamic acid-plasminogen, which assumes a circular conformation that renders this bond less accessible.

Tissue plasminogen activator has little enzymatic activity in the absence of fibrin, but its activity increases by at least three orders of magnitude when fibrin is present. This increase in activity reflects the capacity of fibrin to serve as a template that binds t-PA and plasminogen and promotes their interaction. Whereas t-PA binds fibrin via its finger and second kringle domains, plasminogen binds fibrin via its kringle domains. Kringle domains are loop-like structures that bind Lys residues on fibrin. As fibrin undergoes degradation, more Lys residues are exposed, which provide additional binding sites for t-PA and plasminogen. Consequently, degraded fibrin stimulates t-PA activation of plasminogen more than intact fibrin.

alpha2-antiplasmin rapidly inhibits circulating plasmin by docking to its first kringle domain and then inhibiting the active site. Because plasmin binds to fibrin via its kringle domains, plasmin generated on the fibrin surface resists inhibition by alpha2-antiplasmin. This phenomenon endows fibrin-bound plasmin with the capacity to degrade fibrin. Factor XIIIa cross-links small amounts of alpha2-antiplasmin onto fibrin, which prevents premature fibrinolysis.

Similar to fibrin, annexin II on endothelial cells binds t-PA and plasminogen and promotes their interaction. Cell-surface gangliosides and alpha-enolase also may bind plasminogen and promote its activation by altering its conformation into the more readily activated open form. Plasminogen binds to endothelial cells via its kringle domains. Lipoprotein a, which also possesses kringle domains, impairs cell-based fibrinolysis by competing with plasminogen for cell-surface binding. This phenomenon may explain the association between elevated lipoprotein a levels and atherosclerosis.

Mechanism of Action of Urokinase-Type Plasminogen Activator

Synthesized as a single-chain polypeptide, single-chain u-PA (scu-PA) has minimal enzymatic activity. Plasmin converts scu-PA into a two-chain form that is enzymatically active and capable of binding u-PAR, the u-PA receptor on cell surfaces. Further cleavage at the amino terminus of two-chain u-PA yields a truncated, lower-molecular-weight form that lacks the u-PAR binding domain.

Two-chain forms of u-PA readily convert plasminogen to plasmin in the absence or presence of fibrin. In contrast, scu-PA does not activate plasminogen in the absence of fibrin, but it can activate fibrin-bound plasminogen because plasminogen adopts a more open and readily activatable conformation when immobilized on fibrin. Similar to the higher-molecular-weight form of two-chain u-PA, scu-PA binds to cell surface u-PAR, where plasmin can activate it. Many tumor cells elaborate u-PA and express u-PAR on their surfaces. Plasmin generated on these cells endows them with the capacity for metastasis.

Mechanism of Action of TAFI

Thrombin-activated fibrinolysis inhibitor, a procarboxpeptidase B-like molecule synthesized in the liver, circulates in blood in a latent form while thrombin bound to thrombomodulin can activate it. Unless bound to thrombomodulin, thrombin activates TAFI inefficiently. TAFIa attenuates fibrinolysis by cleaving lysine residues form the carboxy termini of chains of degrading fibrin, thereby removing binding sites for plasminogen, plasmin, and t-PA. TAFI links fibrinolysis to coagulation because the thrombin-thrombomodulin complex not only activates TAFI, which attenuates fibrinolysis, but also activates protein C, which mutes thrombi in generation.

Activated TAFI (TAFIa) has a short half-life in plasma because the enzyme is unstable. Genetic polymorphisms can result in synthesis of more stable forms of TAFIa. Persistent attenuation of fibrinolysis by these variant forms of TAFIa may render patients susceptible to thrombosis.

Factors That Alter Clearance

March 11, 2017 Pharmacokinetics No comments , , , ,

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Body Surface Area (BSA)

Most literature values for clearance are expressed as volume/kg/time or as volume/70 kg/time. There is some evidence, however, that drug clearance is best adjusted on the basis of BSA rather than weight.

The patient’s BSA can be obtained from a nomogram, estimated from below:

BSA in m2 = [(Patient’s Weight in Kg / 70 kg)^0.7]*(1.73 m2)

or

BSA in m2 = (W^0.425)(H^0.725)*0.007184

The following formulas can be used to adjust the clearance values reported in the literature for specific patients. There are other equations one can use depending on units used in the literature for clearance.

  • Patient’s Cl = (Literature Cl per m2)(Patient’s BSA)
  • Patient’s Cl = (Literature Cl per 70 kg) (Patient’s BSA / 1.73 m2)
  • Patient’s Cl = (Literature Cl per 70 kg)(Patient’s Weight in Kg / 70 kg)
  • Patient’s Cl = (Literature Cl per kg)(Patient’s Weight in kg)

When patients do not differ significantly from 70 kg, the difference between using weight versus BSA becomes less significant.

The underlying assumption in using weight or BSA to adjust clearance is that the patient’s liver and kidney size (and hopefully function) vary in proportion to these physical measurements. This may not always be the case; therefore, clearance values derived from the patient populations having a similar age and size should be used whenever possible. If the patient’s weight is reasonably close to 70 kg (BSA = 1.73 m2), the patient’s calculated clearance will be similar whether weight or BSA are used to calculate clearance. If, however, the patient’s weight differs significantly from 70 kg, then the use of weight or surface area is likely to generate substantially different estimates of the patient’s clearance. When a patient’s size is substantially greater or less than the standard 70 kg, or 1.73 m2, a careful assessment t should be made to determine if the patient’s body stature is normal, obese, or emaciated. In obese and emaciated patients, neither weight nor surface area is likely to be helpful in predicting clearance, since the patient’s body size will not reflect the size or function of the liver and kidney.

Plasma Protein Binding

For highly protein-bound drugs, diminished plasma protein binding is associated with a decrease in reported steady-state plasma drug concentrations (total of unbound plus free drug) for any given dose that is administered. It would be misleading, however, to assume that because the calculated clearance is increased, the amount eliminated per unit of time has increased. Actually the amount eliminated per unit of time equals is the production of both Cl and C. In summary, when the same daily dose of a drug is given in the presence of diminished protein binding, an amount equal to that dose will be eliminated from the body each day at steady state despite a diminished steady-state plasma concentration and an increase in the calculated clearance. This is one way to explain the un-changed RE (rate of elimination). In another way to explain, when Css ave changes, the free or unbound fraction of drug in the plasma generally increases (even though Css ave decreases) with diminished plasma protein binding. As a result, the amount of free drug eliminated per unit of time remains unchanged.

And also what is important is that the pharmacologic effect achieved will be similar to that produced by the higher serum concentration observed under normal protein binding conditions. This example re-emphasizes the principle that clearance alone is not a good indicator of the amount of drug eliminated per unit of time (RE).

Extraction Ratio

The direct proportionality between calculated clearance and fraction unbound (fu) does not apply to drugs that are so efficiently metabolized or excreted that some (perhaps all) of the drug bound to plasma protein is removed as it passes through the eliminating organ. In this situation the plasma protein acts as a “transport system” for the drug, carrying it to the eliminating organs, and clearance becomes dependent on the blood or plasma flow to the eliminating organ. To determine whether the clearance for a drug with significant plasma binding will be influenced primarily by blood flow or plasma protein binding, its extraction ratio is estimated and compared to its fu value.

The extraction ratio is the fraction of the drug presented to the eliminating organ that is cleared after a single pass through that organ. It can be estimated by dividing the blood or plasma clearance of a drug by the blood or plasma flow to the elimination organ. At rest, the blood flow to the liver via the portal vein is at a rate of 1300 mL/min, and the other 500 mL/min is suppled by the hepatic artery. If the extraction ratio exceeds the free fraction (fu), then the plasma proteins are acting as a transport system and clearance will not change in proportion to fu. If, however, the extraction ratio is less than fu, clearance is likely to increase by the same proportion that fu changes. This approach does not take into account other factors that may affect clearance such as red blood cell binding, elimination from red blood cells, or changes in metabolic function.

Renal and Hepatic Function

Drugs can be eliminated or cleared as unchanged drug through the kidney (renal clearance) and by metabolism in the liver (metabolic clearance). These two routes of clearance are assumed to be independent of one another and additive.

Clt = Clm + Clr (total Cl = metabolic CI + renal Cl)

Because the kidneys and liver function independently, it is assumed that a change in one does not affect the other. Thus, Clt can be estimated in the presence of renal or hepatic failure or both. Because metabolic function is difficult to quantitate, Clt is most commonly adjusted when there is decreased renal function:

Screen Shot 2017 03 09 at 9 41 09 PM

A clearance that has been adjusted for renal function can be used to estimate the maintenance dose for a patient with diminished renal function. This adjusted clearance equation, however, is only valid if the drug’s metabolites are inactive and if the metabolic clearance is indeed unaffected by renal dysfunction as assumed. A decrease in the function of an organ of elimination is most significant when that organ serves as the primary route of drug elimination. However, as the major elimination pathway becomes increasingly compromised, the “minor” pathway becomes more significant because it assumes a greater proportion of the total clearance. For example, a drug that is usually 67% eliminated by the renal route and 33% by the metabolic route will be 100% metabolized in the event of complete renal failure; the total clearance, however, will only be one-third of the normal value.

As an alternative to adjusting Clt to calculate dosing rate, one can substitute fraction of the total clearance that is metabolic and renal for Clm and Clr. Using this technique the equation below can be derived.

Screen Shot 2017 03 09 at 10 21 01 PM

The Dosing Rate Adjustment Factor can be used to adjust the maintenance dose for a patient with altered renal function.

Most pharmacokinetic adjustments for drug elimination are based on renal function because hepatic function is usually more difficult to quantitate. Elevated liver enzymes do reflect liver damage but are not a good measure of function. Hepatic function is often evaluated using the prothrombin time (or INR), serum albumin concentration, and serum bilirubin concentration. Unfortunately, each of these laboratory tests is affected by variables other than altered hepatic function. For example, the serum albumin may be low due to decreased protein intake or increased renal or GI loss, as well as decreased hepatic function. Although liver function tests do not provide quantitative data, pharmacokinetic adjustments must still take into consideration liver function because this route of elimination is important for a significant number of drugs.

Cardiac Output

Cardiac output also affects drug metabolism. Hepatic or metabolic clearances for some drugs can be decreased by 25% to 50% in patients with congestive heart failure. For example, the metabolic clearances of theophylline and digoxin are reduced by approximately one-half in patients with congestive heart failure. Since the metabolic clearance for both of these drugs is much lower than the hepatic blood or plasma flow (low extraction ratio), it would not have been predicted that their clearances would have been influenced by cardiac output or hepatic blood flow to this extent. The decreased cardiac output and resultant hepatic congestion must, in some way, decrease the intrinsic metabolic capacity of the liver.

Creatinine Clearance Estimation – Non-Steady State

March 8, 2017 Clinical Skills, Laboratory Medicine, Nephrology, Pharmacokinetics, Practice 2 comments , ,

Using non-steady-state serum creatinine values to estimate creatinine clearance is difficult, and a number of approaches have been proposed. The author use Equation 1 below to estimate creatinine clearance when steady-state conditions have not been achieved.

ClCr (mL/min) = { (Production of Creatinine in mg/day) – [(SCr 2 – SCr 1)(V Cr) / t]*(10 dL/L)} * [(1000 mL/L) / (1440 min/day)] / [(SCr 2)(10 dL/L)] [Equation 1]

Screen Shot 2017 03 08 at 7 25 08 PM

The daily production of creatinine in milligram is calculated by multiplying the daily production value in mg/kg/day from Table 5 by the patient’s weight in kg. The serum creatinine values in Equation 1 are expressed in units of mg/dL; t is the number (or fraction) of days between the first serum creatinine measurement (SCr1) and the second (SCr2). The volume of distribution of creatinine (Vcr) is calculated by multiplying the patient’s weight in kg times 0.65 L/kg. Equation 1 (or 79) is essentially a modification of the mass balance equation (we will discuss it in another thread later).

Screen Shot 2017 03 08 at 9 11 53 PM

where the daily production of creatinine in milligram has replaced the infusion rate of the drug and the second serum creatinine value replaced C ave. The second serum creatinine is used primarily because Equation 1 is most commonly applied when creatinine clearance is decreasing (serum creatinine rising), and using the higher of the two serum creatinine values results in a lower, more conservative estimate of renal function. Some have suggested that the iterative search process, as represented by the combination of Equation 28 and 37 (won’t be discussed here; if needed, please contact Tom for detail), be used:

Screen Shot 2017 03 08 at 9 29 36 PMwhere C2 represents SCr2, and C represents SCr1. (S)(F)(Dose/tau) represents the daily production of creatinine, and t represents the time interval between the first and second serum creatinine concentrations. Cl represents the creatinine clearance with the corresponding elimination rate constant K being Cl/V or the creatinine clearance divided by the creatinine volume of distribution. As discussed previously, the solution would require an iterative search, and the inherent errors in the calculation process probably do not warrant this type of calculation.

Screen Shot 2017 03 02 at 10 47 36 PM

Although Equation 1 (or Equation 79) can be used to estimate a patient’s creatinine clearance when a patient’s serum creatinine is rising or falling, there are potential problems associated with this and all other approaches using non-steady-state serum creatinine values. First, a rising  serum creatinine concentration may represent a continually declining renal function. To help compensate for the latter possibility, the second creatinine (SCr2) rather than the average is used in the denominator of Equation 1/79. Furthermore, there are non-renal routes of creatinine elimination that become significant in patients with significantly diminished renal function. Because as much as 30% of a patient’s daily creatinine excretion is the result of dietary intake, the ability to predict a patient’s daily creatinine production in the clinical setting is limited. One should also consider the potential errors in estimating creatinine production for the critical ill patient, the errors in serum creatinine measurements, and the uncertainty in the volume of distribution estimate for creatinine. Estimating creatinine clearance in a patient with a rising or falling serum creatinine should be viewed as a best guess under difficult conditions, and ongoing reassessment of the patient’s renal function is warranted.