1. New Anticoagulants:
Beyond Warfarin and Heparin
Rachel LaCount
Resident in Pathology
August 4, 2005

2. Overview
• Older anticoagulants: Warfarin & Heparin,
LMWH
• Why we need new drugs
• Newer drugs
– Direct Thrombin Inhibitors
– Xa inhibitors
– Heparinoids
– The future

4. Brief Review: Warfarin MOA
• Blocks vitamin K-
dependent glutamate
carboxylation of precursor
factors II, VII, IX, X
• Vit K = cofactor
• Warfarin blocks the
reduction of Vit K
• Oral administration

5. Review: Heparin MOA
• Indirect thrombin inhibitor: IV only
• Called UFH (unfractionated heparin)
• Complexes with AT (heparin co-factor I)
• AT by itself inactivates SLOWLY!
– Thrombin
– Factor Xa
– XIIa, XIa, IXa (lesser extent)
• AT + Heparin: conformational
change in AT = 1000-4000 fold acceleration in inactivation
• At high concentrations: Also binds to platelets and heparin
co-factor II—which inhibits thrombin

6. • Heparin made of polysaccharide chains of varying lengths
• Unique pentasaccharide sequence binds to AT
– Sequence is randomly distributed along heparin chains
• Inactivation of Xa—Heparin doesn’t have to bind to it
• Inactivation of Thrombin--Requires heparin to bind to both
AT and itself
– Heparin must be >18 monosaccharides long to do this
– Virtually all heparin molecules are > 18

7. Limitations of Warfarin and
Heparin:
• Both have narrow therapeutic windows
• Highly variable dose responses: requires
laboratory monitoring (PT, APTT)
– Heparin can bind to other plasma proteins making
bioavailability variable
– Warfarin has numerous food, drug interactions
• Limited ability to stop a clot from propagating:
– Heparin does not inactivate thrombin bound to fibrin
or Xa bound to platelets very well

8. LMWHs
• Molecular wt:
Heparin: 15,000 vs LMWH: 4000-5000
• LMWHs inactivate Xa but have less effect on
thrombin (some molecules not long enough)
– ratio of anti-Xa to anti-thrombin activity of 3:1
– Do not prolong PTT unless dose high
• Advantages over heparin:
– Easier to administer: sq, BID dosing
– Dosage and anticoagulant effect easier to predict; dose
based on body weight
– Lab monitoring not necessary in all patients
– Less chance of inducing immune-mediated
thrombocytopenia

10. Names of LMWHs
• Enoxaparin (Lovenox)
• Dalteparin (Fragmin)
• Tinzaparin (Innohep)
– Differ chemically and pharmacokenetically but
unsure if these differences are clinically
significant
• Other products not yet approved here:
– Fraxiparin, reviparin, nadroparin, bemiparin,
certoparin

11. LMWH Rx monitoring
• Uncomplicated patients do not require
monitoring
• Who may need to be?
– Newborns, children, pregnant women
– Conditions: obesity, renal insufficiency,
malignancy, myeloproliferative disorders
– People with hemorrhagic complications or
with initial therapy to confirm appropriate
levels

12. LMWH Rx monitoring
• Levels measured by chromogenic-based
anti-factor Xa assays
• Clot based APTT only sensitive to very
high levels of LMWHs
• Calibration is done with the same brand of
LMWH that the patient is using—and not
with Heparin!

13. Anti Xa Assay
• Pt plasma + known amount of excess
Factor Xa and antithrombin
• UFH/LMWH binds antithrombin & inhibits Factor
Xa
• Residual Factor Xa is measured
– Factor Xa cleaves a chromogenic substrate similar to
its natural substrate, releasing color detected by a
spectrophotometer
• Residual Xa is inversely proportional to the
amount of LMWH (or UFH)

14. Why new drugs?
• UFH and LMWHs are inconvenient for the
outpatient setting (IV or sq only)
• UFH and LMWHs can cause HIT:
– Risk 0.2% with LMWH vs. 2.6 % with UFH
– Pts with HIT still need to be anticoagulated

15. Why new drugs?
• Warfarin is underused in pts who need it
most:
– Only 47% of patients with afib are taking
warfarin
– This is often due to hemorrhagic
contraindications
– Convenience issues due to the need for
frequent monitoring
– Difficulty in maintaining optimal
anticoagulation

16. The ideal anticoagulant
• Effective
• Minimal complications/side effects
• Convenient administration (ie: oral for outpatients)
• Rapid absorption
• Fast on and offset action
• Predictable pharmacokinetics
• No interactions with food or drugs
• No HIT
• No coagulation monitoring

17. Categories of new drugs
– Direct Thrombin Inhibitors:
• hirudin, lepirudin, desirudin, bivalirudin,
argatroban, ximelagatran
– Xa inhibitors:
• fondaparinux, idraparinux
– Heparinoids:
• Danaparoid (discontinued)

18. Objectives for each drug
• Mechanism of Action
• Current uses
• Limitations
• Monitoring in the lab

19. Direct Thrombin Inhibitors

20. Direct Thrombin Inhibitors
• 3D structure of thrombin:
100s of thrombin inhibitors
in last 15 yrs
• Most are peptidomimetic
compounds
– Mimic the fibrinogen
sequence which interacts
with the thrombin active site
– Peptidomimetic: A
compound that mimics
binding and biological
activity of the natural
peptide

21. Thrombin
3 important areas:
• Active site: fibrinogen
binding
• Exosite I: major
docking site--interaction
with fibrinogen and other
receptors; fibrinogen
recognition site
3. Exosite II: interacts
with heparin

22. Hirudin
• Medicinal leeches:
– Used since ancient
times to relieve body
of “bad humors”
• Egyptians, Greeks
– Reached peak Hirudo medicinalis
popularity in mid-19th
century

23. • 1884: John Haycraft in Birmingham
demonstrated that medicinal leeches, Hirudo
medicinalis, secrete a substance that prevents
blood from clotting
• 1904: Substance named hirudin
• 1957: Markwardt isolated the active
anticoagulant substance, determined it to be a
polypeptide 65 AAs long which inhibited
thrombin

24. • Estimated to require 50,000 leeches
annually for diagnostics and treatment
• 1986: DNA isolated and cloned
• Today recombinant hirudin is made in
yeast cells
– Lepirudin, desirudin, bivalirudin

25. R-Hirudins
• All bind in active site and exosite I
• Irreversible: Lepirudin, Desirudin
• Reversible: Bivalirudin
• Minor differences in structure between them
– ie: Lepirudin has one extra oxygen molecule than
desirudin and one AA difference

26. • Lepirudin (Refludan)
– Approved for use in HIT
• No binding to platelet factor IV
• 89% of patients with rapid increase in plt count
– Monitored with APTT daily
• Measure 4 hr after dose
• Target: Pt’s APTT to be 1.5-2.5 x the lab’s median APTT
• Caution in pts with renal insufficiency
• Dose dependent relationship
– Antihirudin antibodies develop in 40-70%
• Drug is made of non-human proteins
• Can cause irritation to skin
• Not neutralizing; may enhance drug potency by delaying
clearance
• These patients need to be monitored with APTT

27. • Desirudin (Iprivask)
– Studied in DVT prophylaxis for total hip
• Lower rate of DVT than LMWH and no increase in
bleeding complications
– Also used in HIT
– Also monitored with APTT
• Monitor especially in pts with renal insufficiency
• APTT twice upper limit of normal = stop and
restart at reduced dose
• Dose dependent relationship
– Antihirudin antibodies can also develop

28. • Bivalirudin (Angiomax)
– Previously called hirulog
– Binds reversibly to thrombin
• Thrombin slowly cleaves the drug from its active site
• Short half-life 20-30 min
– Less immunogenic (is only 20 AAs long)
– FDA approved in 2002 for use in angioplasty for
patients with unstable angina

29. • Bivalirudin, cont…
– REPLACE-2 (2004)
• Randomized Evaluation in PCI Linking Angiomax
to Reduced Clinical Events
• Pts randomized to receive either bivalirudin or
heparin + GP IIb/IIIa inhibitor during PCI
• Significant reduction in in-hospital bleeding
• Endpoints about equal: MI, urgent repeat
revascularization
• Trend towards decreased mortality
– Undergoing study for use in cardiac surgery
for use both “on pump” and “off pump”
– Monitored with APTT or ACT

30. Direct Thrombin Inhibitors, cont…
• Argatroban (Novastan)
– Binds at active site reversibly
– Does not bind at exosites
– First introduced in Japan in 1990 for treatment of
peripheral vascular disorders

31. • Argatroban, cont…
– FDA approved in 2000: Anticoagulant for pts
with HIT needing prophylaxis or treatment for
thrombosis
– 2002 approved for pts with or at risk for HIT
undergoing PCI
– Small, without immunogenicity
– Monitoring with APTT or ACT recommended
• Desired APTT: 1.5 to 3 times the initial baseline
value (not to exceed 100 seconds)

32. • Ximelagatran (Exanta)
– ORAL !!!!! BID dosing
– Only binds to active site; reversible
– Prodrug for melagatran
– Numerous Phase III trials (10):
• 6 for prophylaxis of venous thromboembolism [VTE]
due to orthopedic surgery
• 1 for initial treatment of VTE
• 1 for long-term prevention of VTE recurrence
• 2 for stroke prophylaxis due to atrial fibrillation
– The good news: Some studies have shown
benefit over warfarin or LMWH

33. Ximelagatran, cont…
The bad news:
• Studies have shown increase in bilirubin, ALT
• FDA: Estimates rate of liver injury 1 in 200
• 10% could progress to liver failure, need liver
transplantation, or death
– 1 in 2000 patients tx’d long-term could have overt
liver failure
• 3 of 6948 patients did die under circumstances
that FDA felt could reasonably be related to
ximelagatran
– Consistent with the 1-in-2000 rate

34. DTIs…a bit more on lab testing
• Interference with APTT:
– Other drugs: warfarin, abx
– Pts with LA
– High factor VIII levels leading to falsely low APTT
• Why not do a Factor IIa assay to measure drug
activity?
– For example, chromogenic anti-factor IIa assay
– Limited reproducibility, linearity, and sensitivity

35. DTIs…
• Why not use PT/INR?
– Little to no sensitivity to some Rx and less
than optimal to others
– INR can be markedly different depending on
reagent used
– One recent study of lepirudin, argatroban, &
bivalirudin observed a dose dependent effect
on the INR
• PT/INR most affected by argatroban at therapeutic
concentrations
• Lepirudin had the least overall effect on PT/INR

36. DTIs & Lab tests, cont…
• TCT (thrombin clotting time)
– Seems logical, but is overly sensitive to these drugs
and doesn’t provide useful clinical info
• ACT (activated clotting time)
– Has been used successfully in the OR
• ECT (ecarin clotting time)
– Not standardized for clinical use, but shows promise
– Venom from snake Echis carinatus:
• Converts prothrombin to a meizothrombin that is sensitive to
thrombin inhibitors

37. Factor Xa inhibitors

38. Factor Xa inhibitors
• Fondaparinux (Arixtra)
– Synthetic polysaccharide:
– The drug is the unique pentasaccharide sequence that
UFH and LMWH use to bind to AT
– Reacts with strong affinity to AT (reversible) →
Induces conformational change in AT →
Increased ability to inactivate Xa

39. Fondaparinux: Too short to inactivate thrombin (much like
LMWH); need >18 saccharide units to inactivate thrombin

40. • Fondaparinux, cont…
– Does not interact with plasma proteins,
platelets, or platelet factor IV = useful in HIT
(although not yet formally approved)
– FDA approved in 2001
• Prevention of post op VTE (DVT and PE) in
orthopedic surgery
– Hip fracture, hip replacement, knee replacement
– Fondaparinux vs. enoxaparin in one study decreased
VTE in knee replacement from 12.5 to 27.8%
• 2004/5 approval:
– VTE treatment if administered with warfarin
– Anticoagulation in abdominal surgery
• Potential uses being studied: MI, PCI, UA

41. • Fondaparinux, cont…
– Drug monitoring:
• APTT and PT are insensitive
• PT/INR may or may not be proportional to the
clinical safety or efficacy—more studies needed
• Anti-factor Xa assay –must be calibrated with
fondaparinux
– Long half-life (17 hours) = qd dosing
(LMWH = BID)
• Idraparinux
– Longer acting analogue (q week dosing)
currently being developed

42. Danaparoid (Orgaran)
• A LWM heparinoid
• Derived from porcine gut mucosa
– (can’t use if have a pork allergy)
• A mixture of heparan sulfate, dermatan
sulfate, chondroitin sulfate
• Does not have heparin or heparin
fragments (heparan differs from heparin
by sulfur groups on the sugar molecules)

43. Danaparoid: MOA
• Binds to AT and heparin cofactor II
• More selective at inhibiting Xa than LMWH, not as
selective as fondaparinux:
Drug Anti-Xa/anti-IIa
ratio
UFH 1:1
LMWH 2-4:1
Danaparoid 22:1
Fondaparinux ∞
• Ability to prevent extension of thrombi
• Minimal effect on platelet function and aggregability

44. Danaparoid
• FDA approved in 1996
– Proven effective in DVT prophylaxis in pts
undergoing hip surgery
• Used off label in patients with HIT
• Discontinued manufacturing in USA
4/2002 due to problems obtaining raw
material
– Available in Germany

45. Future possible drugs
• Ticks
Isolated from the saliva or
blood of various species:
– Thrombin inhibitors
– Factor Xa inhibitors,
– Tissue factor pathway
inhibitor (TFPI)

46. Future possible drugs
• Razaxaban
– Oral drug
– Inhibitor of factor Xa without requiring AT
– In phase II trials
• DX-9065a
– Xa inhibitor; also in phase II trials; IV
• Potential targets being developed
– TF/fVIIa
• Recombinant tissue factor inhibitor (FTPI)
• Other specific TF/fVIIa or fVIIa inhibitors being developed
– Recombinant APC
• In phase III trials—inactivates Va and VIIIa

47. Future possible drugs
• Aptamers:
– From root word aptus (“fit”)
– Single stranded nucleic acids that fold into
specific 3D structures which bind and inhibit a
protein target
– Anti factor VIIa, IXa, and thrombin aptamers
have been developed
– Should be nonimmunogenic—small and
similar to endogenous molecules
– Possible use in HIT

48. References
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Thrombolysis. 2003;16 (1/2): 49-54.
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Anesthesiologists. 2003 Dec.
(http://www.scahq.org/sca3/newsletters/2003dec/drug4.shtml)
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lepirudin, and argatroban, on prothrombin time and INR values. Am J Clin
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49. References, cont…
• Markwardt, F. Past, present and future of hirudin. Haemostasis.
1991;21 Suppl 1:11-26.
• Martel, N. et al. Risk of heparin induced thrombocytopenia with
unfractionated and low molecular weight heparin thromboprophylaxis: a
meta-analysis. Blood. 2005 Jun 28 (Epub ahead of print).
• Nimjee, S. et al. The potential of aptamers as anticoagulants. TCM.
2005; 15(1):41-45.
• Starke, K. The beginnings of hirudin. Trends Pharmacol Sci. 1989 Mar;
10(3):99.
• Walenga, J. et al. Monitoring the new antithrombotic drugs. Seminars in
Thrombosis and Hemostasis. 2004; 30(6): 683-695.
• Weitz, J. et al. Treatment of venous thromboembolism: New
anticoagulants for treatment of venous thromboembolism. Circulation.
2004;110:I-19 – I-26.
• White, CM. Thrombin-directed inhibitors: Pharmacology and clinical
use. American Heart Journal. 2005 Jan;149(15):S54-60.
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heparin. Uptodate