Activated Clotting Time Applications, Therapeutics and Technologies Marcia L. Zucker, Ph.D. Director of Clinical Support Response Biomedical Corporation mzucker@responsebio.com ITC Educational Services, Edison, NJ

There was….. The Lee-White clotting time Add blood to glass tube, shake No activator Manual Place in heat block Examine for clot every 30 seconds Very slow Subjective clot detection

Time goes on…

1966 - Hattersley Activated Clotting Time Add blood to glass tube with dirt, shake Diatomaceous earth activator Still manual Place in heat block Visual clot detection Subjective clot detection

Particulate Contact Activation Initiation of intrinsic coagulation cascade Factor XII (Hageman factor) Prekallikrein (Fletcher factor) Shortens contact activation period Proposed as both screening assay for coagulation defects and for heparin monitoring Critical fibrin clot endpoint

Semi - Automation - 1969 HEMOCHRONOMETER Later - HEMOCHRON Add blood to tube, shake Manual sample treatment Place in test well Automated heating Objective fibrin clot detection two different activators CA510 (later FTCA510) diatomaceous earth P214 glass bead

Mechanical Detection Magnet in tube, detector in instrument Upon fibrin clot formation, magnet is deflected Clotting time displayed

Two assays for separate uses

1980’s HemoTec ACT (later Medtronics ACTII) Add blood to dual cartridge Liquid kaolin activator Place in instrument Automated mixing Objective clot detection Flag moves up and down As fibrin forms, motion slows Mechanical clot detection Instrument displays clotting time

Lower values than CA510 – differences ignored by clinicians

1980’s - ACT Differences Reported in literature >20 years Clinical evaluations of Hemochron - mid 1970’s By 1981 – poor correlation between ACT and heparin level By 1988 Hemochron and HemoTec clinically different Early ’80’s to Present Improved clinical outcome with ACT use Reviewed: Draft NACB Laboratory medicine practice guideline for point of care coagulation testing http://www.nacb.org/lmpg/poct_LMPG_draft_PDF.stm

1990’s Microsample ACTs - Hemochron Jr Add blood to sample well, press start Silica, kaolin and phospholipid (ACT+) Diatomaceous earth (ACT-LR) Automated sample measurement Automated mixing Objective clot detection Sample pumped across restriction Flow slows with clot formation Optics measure motion Clotting time displayed

Clotting Times Different

2000 Abbott - i-STAT Add blood to cartride, press start Diatomaceous earth or kaolin Insert into instrument Automated mixing No clot detection Synthetic thrombin substrate Electro-active compound formed and detected amperometrically “Clotting time” reported

Clotting Times Likely Different i-STAT ACT Package Insert Used to monitor moderate- and high-level heparin Prewarmed vs. non-prewarmed calibration modes Good correlation with Hemochron (Celite below) Intercept varied by site (+4 to -73 sec) Slope range 0.85 – 1.19 Few publications All show wide scatter vs Hemochron or Medtronics

Today Hemochron Response Hemochron Signature Elite Medtronics ACT Plus Medtronics HMS+ Helena Actalyke XL / Mini Gem PCL Abbott i-STAT

Activated Clotting Time Extrinsic Pathway Intrinsic Pathway ACT Common Pathway CLOT

Why do we use an ACT? Maintain Balance Point of Care Bleeding Thrombosis Point of Care Immediate turn around Rapidly adjust anticoagulant dosing as needed Heparin – half life varies by patient Dose required varies by patient Potency varies by lot Direct thrombin inhibitors – very short half life Require immediate intervention No antidote available

NACB Guidelines National Academy of Clinical Biochemistry Evidence Based Guidelines for POC Testing http://www.nacb.org/lmpg/poct_lmpg_draft.stm Chapter 4 – Coagulation Testing Caveats: Assume all systems equally safe / effective / precise Correlation studies excluded Few good outcome trials

NACB Guidelines Is there evidence of improved clinical outcome using ACT testing? In cardiovascular surgery? Is there evidence for optimal target times to be used with ACT monitoring? Strongly recommend ACT monitoring of heparin anticoagulation and neutralization in cardiac surgery. (Class A, Level I at least one randomized controlled trial, Level II–1 - small randomized controlled trials, non-randomized controlled trials). Insufficient evidence to recommend specific target times for use during cardiovascular surgery. (Class I – conflicting evidence across clinical trials).

NACB Guidelines Is there evidence of improved clinical outcome using ACT testing? In interventional cardiology? Is there evidence for optimal target times to be used with ACT monitoring? Strongly recommend ACT monitoring of heparin anticoagulation in interventional cardiology. (Class A, Level II–1 - small randomized controlled trials, non-randomized controlled trials, Level II-2 – Case controlled analytic studies from more than one center or research group). Recommend target times specific to ACT system Targets differ if specific platelet inhibitors are used concurrently with heparin.

NACB Guidelines Target Times: Without intravenous platelet inhibitors: >250 seconds using the Medtronics ACTII >300 seconds using the HEMOCHRON FTCA510 tube (Class B – Level II–1 - small randomized controlled trials, non-randomized controlled trials, Level II-2 – Case controlled analytic studies from more than one center or research group). With the intravenous platelet inhibitors: abciximab or eptifibatide: 200-300 seconds tirofiban: 250 – 300 (Class B – Level I- at least one randomized controlled trial).

NACB Guidelines Is there evidence of improved clinical outcome using ACT testing? In ECMO? Is there evidence for optimal target times to be used with ACT monitoring? ExtraCorporeal Membrane Oxygenation Strongly recommend ACT monitoring to control heparin anticoagulation during ECMO. (Class A – Level III – opinions of respected authorities based on clinical experience, descriptive studies or reports of expert committees). Target times for ECMO based on the ACT system. (Class B – Level III – opinions of respected authorities based on clinical experience, descriptive studies or reports of expert committees).

NACB Guidelines Is there evidence of improved clinical outcome using ACT testing? In other applications (e.g. vascular surgery, intravenous heparin therapy, dialysis, neuroradiology, etc.)? Is there evidence for optimal target times to be used with ACT monitoring? There is insufficient evidence to recommend for or against ACT monitoring in applications other than cardiovascular surgery, interventional cardiology or extracorporeal oxygenation. (Class I).

ACT - Clinical Applications Essential - outcome studies show evidence Cardiac surgery Percutaneous coronary intervention (PCI) ECMO / (Dialysis) Common – insufficient evidence for “essential” Critical care Interventional radiology Electrophysiology Vascular surgery

Different ACTs for each clinical setting

Clinical Applications Essential Cardiac surgery Percutaneous coronary intervention (PCI) ECMO / (Dialysis) Common Critical care Interventional radiology Electrophysiology Vascular surgery

Monitoring - ACT Benefits Disadvantages Industry Standard Since 1970s Recommended as 1o method in AmSECT guidelines ACT improves outcome in CPB, PCI NACB draft LMPG for POC coagulation Easy to run Disadvantages Each system yields different numbers Most sensitive to hypothermia and hemodilution Little or no correlation to heparin level especially true for pediatric patients

When to choose which ACT? Different pharmaceutical interventions Antifibrinolytics Novel anticoagulants Different patient populations Especially pediatric patients Different monitoring preferences Heparin Level Need for specialty assays Dosing Fibrinogen

Pharmaceutical Intervention Amicar or Tranexamic Acid No effect on standard celite ACT Continued debate on efficacy Multiple reports of reduction in post-operative blood loss and reduced transfusion requirements Aprotinin Significant elevation of Celite ACT Two dosing regimens – Patient size independent Full Hammersmith 2 x 106 KIU loading dose; 2 x 106 KIU pump prime; 0.5 x 106 KIU/hr infusion Half Hammersmith 1 x 106 KIU loading dose; 1 x 106 KIU pump prime; 0.25 x 106 KIU/hr infusion

ACT Monitoring-Aprotinin Treatment Celite® ACT HEMOCHRON FTCA510; Actalyke C-ACT; i-STAT ACTCelite Not recommended Still used with target times of >750 seconds Kaolin ACT HEMOCHRON FTKACT; Actalyke K-ACT; i-STAT ACTKaolin; Medtronics HR-ACT Unaffected by moderate doses of aprotinin Used with target times of > 480 seconds Combination ACT reagents HEMOCHRON Jr ACT+; Actalyke MAX-ACT; GEM PCL ACT Unaffected by ALL doses of aprotinin Used with target times of > 400 seconds

ACT Monitoring-Aprotinin Treatment Jones, et al. JECT. 2004;36:51–57

Novel Pharmaceuticals None FDA approved for cardiac surgery Direct thrombin inhibitors (DTIs) Used if patient at risk for HIT Heparin induced thrombocytopenia “Heparin allergy” Argatroban Refludan Angiomax

ACT Monitoring - DTIs Argatroban Refludan Synthetic analog of L-arginine Reversible binding to thrombin PCI monitoring: ACT 300 – 450 Papers state standard ACT targets for CPB Refludan Recombinant hirudin Irreversible inhibition of thrombin Routine monitoring with aPTT No recent reports in CPB Original studies used ECT - No longer available

ACT Monitoring - DTIs Angiomax Synthetic analog hirudin Reversible binding to thrombin PCI monitoring: HEMOCHRON ACT-LR >300 Papers state standard ACT not appropriate for CPB Adapted from Zucker, et.al., JECT 2005, in press ACTT not yet available

Patient populations Pediatric Patients are NOT little adults Reports of both shorter and longer heparin half life Pediatric Patients are NOT all the same In neonates, coagulation factor levels confound dilutional effects Once patient exceeds 2 years of age, only dilutional effects need to be considered These can be very important Test volume is critical

Pediatric Monitoring Data from clinical evaluation, on file, ITC

Monitoring - Heparin Level Benefits Measures concentration, not activity Correlates with laboratory standards Disadvantages Each system yields different numbers apples and oranges do not compare Correlation to anticoagulation status is still disputed Target for neonate, pediatric and adult patients may differ

Monitoring - Heparin Level Young: <4.5 years Shayevitz, JR and O’Kelly, SW Progress in Anesthesiology, vol. IX, chapter 16 1995

Monitoring - Heparin Level αXa / αIIa Activity - laboratory only, impractical Not available with microsample systems Medtronic Hepcon HMS Indirect measure of protamine reversible heparin activity Thromboplastin based HEMOCHRON PRT ACT based protamine titration HEMOCHRON HiTT unaffected by hemodilution, hypothermia insensitive to aprotinin Thrombin Time based Correlates with αXa and αIIa activity

Heparin Monitoring Dilemma Level ACT Which Value Determines Response?

More reasons for selecting a system Heparin and Protamine Dose Optimization In vitro dosing systems Not available on microsample systems Hemochron RxDx Heparin Response Time (HRT) Protamine Response Time (PRT) PDAO HMS HDR HPT

Heparin Dose Optimization Traditional dosing regimens recommend fixed drug doses by body weight. Regimen does not account for patients whose response to heparin is different than the average “normal” patient. Patients differ in their response to heparin; may be resistant or sensitive to heparin Can represent 20 - 40% of patient population Response can vary up to 12 fold between patients

Protamine Dose Optimization Minimize protamine dose Reduce time for infusion Avoid protamine related adverse events hypotension shock bleeding platelet dysfunction

Clinical Benefits Individualize heparin dose Reduce blood products needed for post-operative transfusions i.e. red blood cells, platelets, fresh frozen plasma and cryoprecipitate Jobes DR, et al. 1995. J Thorac Cardiovasc Surg 110: 36-45 Reduced potential for protamine reaction Protamine reduced by average of 30% Zucker ML., et al. 1997. J Extra-Corp Tech. 29: 176-180. Reduced incidence of return to OR for bleeding

Economic Benefits (US$) Jobes DR., et al. 1996. Cost Effective Management Of Heparin/ Protamine In CP Bypass: Analysis By Type Of Surgery. Anesthes 85:3A.

Heparin neutralization verification Test Options ACT - global indicator of coagulation PDAO /ACT – protamine titration HEMOCHRON Response Heparinase ACT – enzymatic confirmation Medtronics ACTPlus Thrombin time – highly sensitive to heparin TT/HNTT – protamine titration aPTT - intrinsic pathway HEMOCHRON tube systems HEMOCHRON microsample systems

Clinical Applications Essential Cardiac surgery Percutaneous coronary intervention (PCI) ECMO / (Dialysis) Common Critical care Interventional radiology Electrophysiology Vascular surgery

Procedures Diagnostic Interventional Catheterization Electrophysiology locate and map vessel blockage(s) determine need for interventional procedures Electrophysiology Interventional Balloon angioplasty Atherectomy (roto-rooter) Stent placement

Diagnostic – Low dose heparin Catheterization and Electrophysiology 2500 - 5000 unit bolus dose frequently not monitored if monitored – ACT HEMOCHRON FTCA510; HEMOCHRON Jr ACT-LR; Medtronics LR-ACT: Actalyke C-ACT; GEM PCL ACT-LR Targets ~ 200 seconds aPTT

Interventional Angioplasty, Atherectomy, Stent placement HEMOCHRON FTCA510; HEMOCHRON Jr ACT-LR; Medtronics LR-ACT or HR-ACT: Actalyke C-ACT, MAX-ACT; GEM PCL ACT-LR; i-STAT ACTCelite 10,000 unit bolus dose or 2 - 2.5 mg/kg target ACT 300 - 350 seconds Target must be reduced if ReoPro Used ReoPro is one of 3 “GPIIb/IIIa” Inhibitors GPIIb/IIIa receptor critical in platelet aggregation

Novel Pharmaceuticals Platelet Inhibitors Oral Aspirin Plavix Parenteral ReoPro Integrilin Aggrastat Low Molecular Weight Heparins (LMWH) None FDA approved for interventional cardiology Lovenox Fragmin

Platelet Structure PLAVIX ASPIRIN Parenteral platelet inhibitors Courtesy of Helena Laboratories

Intervention promotes aggregation Picture courtesy of Reopro.com

ACT Monitoring - Platelet Inhibitors Aspirin No reported effects on ACT No anecdotal reports of effects on ACT Plavix Anecdotal reports Increased bleeding at sheath pull ACTs don’t return to “normal” prior to sheath pull Plateau ~ 180 – 220 seconds

ACT Monitoring - Platelet Inhibitors ReoPro elevates ACTs Study used FTCA510 and Medtronics HR-ACT target time = 250 sec with ReoPro determined using FTCA510 tube Integrelin No reported clinically significant effects on ACT Slight decrease in ACT Study used FTCA510 and ACT-LR Aggrastat No reported effects on ACT

ACT Monitoring - LMWH Fragmin Lovenox Reports of efficacy of ACT monitoring during PCI Studies used FTCA510; ACT-LR; HR-ACT; LR-ACT Lovenox Reports that ACT not useful for monitoring in PCI Adapted from el Rouby, et.al., J Thromb Thrombolys 2005, in press HEMONOX not yet available

Targets can change with ACT Choose ACT by application and Intended Use:

Clinical Applications Essential Cardiac surgery Percutaneous coronary intervention (PCI) ECMO / (Dialysis) Common Critical care Interventional radiology Electrophysiology Vascular surgery

ECMO (Dialysis) ACT used to monitor low dose heparin P214 glass activated ACT has been the “standard” HemoTec HR and LR also used Targets generally 180 - 220 seconds Changing ACT requires changing target Target often 220-260 with ACT-LR Not all ACTs have low heparin indication Also indicated for low dose: GEM PCL ACT-LR; Actalyke G-ACT

Clinical Applications Essential Cardiac surgery Percutaneous coronary intervention (PCI) ECMO / (Dialysis) Common Critical care Interventional radiology Electrophysiology Vascular surgery

ACT or aPTT Determine when to pull the femoral sheath HEMOCHRON FTCA510; HEMOCHRON Jr ACT-LR; Medtronics LR-ACT; Actalyke C-ACT; GEM PCL ACT-LR Premature sheath pull can lead to bleeding. Delayed removal can increase time in CCU. Target set at each site. ACT targets range from 150 – 220 seconds aPTT targets range from 40 – 70 seconds Monitor heparin therapy Target times determined by each facility

Clinical Applications Essential Cardiac surgery Percutaneous coronary intervention (PCI) ECMO / (Dialysis) Common Critical care Interventional radiology Electrophysiology Vascular surgery

Low dose heparin Interventional Radiology Targets generally ~250 - 300 Sometimes target “twice baseline” Twice baseline can differ by ACT heparin sensitivity More centers targeting “increase from baseline” e.g. >130 second increase from baseline

Clinical Applications Essential Cardiac surgery Percutaneous coronary intervention (PCI) ECMO / (Dialysis) Common Critical care Interventional radiology Electrophysiology Vascular surgery

Low dose heparin Vascular Surgery and Electrophysiology Targets generally ~200 – 250 frequently no monitoring Change from baseline is critical factor Is heparin having an effect? Is patient responding “normally”? ~20% patients “resistant” <80 sec/unit/ml blood ~60% patients “normal” 80-120 sec/unit/ml blood ~20% patients “sensitive” >120 sec/unit/ml blood

Summary ACT is NOT a standardized test Each assay yields different results Activator differences Endpoint detection differences Current targets developed on Hemochron First available non-manual system Targets must be adjusted for other systems

Summary While all systems correlate, they yield different results Targets may need to be adjusted by clinical application Inter-operator differences minimized with microcoagulation technology

The future of ACT?