Dagmar Kubitza-Biomarkers for new … for new anticoagulants – vice and virtue Dagmar Kubitza, MD...

Biomarkers fornew anticoagulants – vice and virtue

Dagmar Kubitza, MD

AGAH, München 2014

Definition of Biomarkers?

….Surrogate markers are primary measures the effectiveness of investigational drugs….

Biomarkers for new oral anticoagulants… …which biomarkers?

Charles Esmon, Ann Rev Cell Biol 1993

New oral anticoagulants – mode of action?

Turpie, Current Opinion in Drug Disc & Dev 2009

Biomarkers for new anticoagulants -the „old fashioned way“?

Mani et al, Thrombosis Journal 2013

Dabigatran has a low sensitivity for PT

Douxfils et al, Thromb Heamost 2012

Samama et al, Thrombosis Haemostasis 2010

Direct inhibitors of Factor Xa prolong PT


Samama et al, Thrombosis Research 2011

Samama et al, Thrombosis Haemostasis 2010



Samama et al, Thrombosis Haemostasis 2010Douxfils et al, Thrombosis Haemostasis 2013




Fonaparinux has no effect on PT

Some direct inhibitors of Factor Xa prolong PT



Fonaparinux has no effect on PT

Calculation of INR does not decrease the variablility of the results of the different thromboplastin reagents


Background information about PT PT assays are designed to reliably measure the effect of Vitamin K

antagonists

Different reagents use different thromboplastins to start the clot formation ex vivo, e.g. rabbit brain, placenta, recombinant

Concentrations of phospholipids and ions vary between tests and evenbetween lots.

Components are added to ensure the optimal performance of tests, someassays even contain heparins.

ISI (international sensitivity index) should ensure comparability between tests, but they are optimized for Vitamin K antagonists.

What influences the PT sensitivity of direct Factor Xa inhibitors?

PT sensitivity of Rivaroxaban varies depending on the composition of thethromboplastin reagent PT sensitivity increases with Decreasing concentrations of tissue factor Increasing concentratíons of phospholipids and NaCL Presence of phosphadidylethanolamine

PT sensitivity decreases with Increasing percentage of phosphadidylserine

Smith & Morrissey, Blood 2007

Why? Hpothesis The faster that factor Xa is generated and assembled into the prothrombinase

complex, the less sensitive will the clotting time be on Rivaroxabanconcentration.

Perhaps this is because it allows factor Xa to generate thrombin before Rivaroxaban can inhibit it effectively during the very short duration of most clotting tests.

It remains unclear why Apixaban behaves different than Rivaroxaban and Edoxaban.

Smith & Morrissey, Blood 2007

Is PT a useful biomarker?PT can be used to provide insights into the pharmacodynamic effects of anticoagulants if … comparisons are made for one compound comparisons are made from preclinical to clinical study results, provided that

the same assay is used comparisons are made between studies of one compound and a central lab

has been used

Comparisons of pharmacodynamic effects between different compounds cannotbe reliably done with PT

Will a broader view on the coagulationsystem provide better insights?

Will a broader view on the coagulationsystem provide better insights?

Endogenous thrombin potential (ETP)• Is the total amount of thrombin which is generated in plasma• It reflects the sum of the activity and concentration of pro-coagulant and

anti-coagulant substances

ETP-AUCtotal amount of thrombin that can be generated

PeakMaximal hight of the TG curve

Lag timeTime interval until thrombin generation starts

Time to peakTime interval until the peak of thrombin generationoccurrs

Influence of Dabigatran on Thrombin Generation

Dabigatran mainly delayed the initiation phase with strong dose-dependent increase of lag time and time to peak and a slight dose-dependent decrease in peak hight and ETP

Douxfils et al, Thromb Haemost 2012

Influence of Rivaroxabanon Thrombin Generation

a contolb 5nM ~ 2,2 µg/Lc 10nM ~ 4,4 µg/Ld 20nM ~ 8,7µg/Le 50nM ~21,7µg/Lf 80nM ~34,9µg/Lg100nM ~43,5µg/L

Gerotziafas et al, J Thromb Haemost 2007

Influence of Rivaroxaban and Edoxabanon Thrombin Generation

a contolb 5nM ~ 2,2 µg/Lc 10nM ~ 4,4 µg/Ld 20nM ~ 8,7µg/Le 50nM ~21,7µg/Lf 80nM ~34,9µg/Lg100nM ~43,5µg/L

Gerotziafas et al, J Thromb Haemost 2007 Samama et al, Thrombosis Research 2012

Rivaroxaban and Edoxaban affect the initiation and amplificationphase of thrombin generation by affecting all parameters of TG

Influence of Apixabanon Thrombin Generation

Douxfils et al, Thrombosis Haemost 2013

Influence of Apixaban and Fondaparinuxon Thrombin Generation

Apixaban and Fondaprinux mainly act on the amplificationphase of TG by affecting the peak and the velocity rate index

Douxfils et al, Thrombosis Haemost 2013 Samama et al, Thrombosis Research 2012

Will a broader view on the coagulation systemprovide better insights? Thrombin Generation is a sensitive marker for anticoagulant effects

independent of the mode of action. Compounds with a different mode of action have different effects on TG. Compounds with the same mode of action have different effects on TG.

☻ Comparisons of pharmacodynamic effects between different compoundscannot be reliably done.

☻It remains unclear what the clinical relevance of the differences in influence of individual parameters of TG is.

Can the DOAKs be reliably measured in clinicalpractice?

Dabigatran can be measured with HTIHemoclot Thrombin Inhibitor

Hemoclot Thrombin Inhibitor shows a concentation dependent prolongation of clottingtime with linear relation and a good reproducibility

Douxfils et al, Thromb Haemost 2012

Anti-Xa assays with specific calibrators and controls forRivaroxabanshow linear concentration dependent decrease in OD

Hyphen Biomed, France

Samama et al, J Thromb Thrombolysis, 2012

Samama et al, Thromb Heamost, 2010

Anti-Xa assays with specific calibrators and controlsfor Rivaroxaban and Edoxaban show linear concentration dependent decrease in OD


Hyphen Biomed, France

Samama et al, J Thromb Thrombolysis, 2012

Samama et al, Thromb Heamost, 2010

Anti-Xa assays with specific calibrators and controlsfor Apixaban

Douxfils et al, Thrombosis Haemost 2013

Most sensitive reagents show an exponential correlation but the dynamicrange of quantification is lower.

Can the DOAKs be reliably measuredin clinical practice?

Chromogenic assays with drug specific calibrators and controls can be usedto reliably measure direct oral anticoagulants

Biomarkers for new anticoagulants –vice and virtue


Yes, the pharmacodynamic effects of the DOACs can be determined by biomarkersBUT …

Biomarkers for new anticoagulants –vice and virtue


Yes, the pharmacodynamic effects of the DOACs can be determined by biomarkersBUT

Many influencing factors have to be considered in interpreting the results.

Thank you!

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