The Clinical Use and Immunologic Impact of Thrombin in Surgery
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1 The Clinical Use and Immunologic Impact of Thrombin in Surgery Jeffrey H. Lawson, M.D., Ph.D. Departments of Surgery and Pathology Duke University Medical Center Durham, North Carolina
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3 Bovine Thrombin Human Thrombin and Who Cares
4 The Many Faces of Thrombin KDa { Protein bands above the molecular weight of α-thrombin BT1 BT2 rht BT1: Thrombin-JMI 20 U, Lot 1 BT2: Thrombin-JMI 20 U, Lot 2 rht: rhthrombin 20 U α-thrombin 36.5 α-thrombin Thrombin B-chain Thrombin B-chain β-thrombin α-thrombin with truncated B-chain β-thrombin γ-thrombin 14 (presumed) Thrombin - B4 chain 6
5 What is Topical Bovine Thrombin?
6 Bovine Thrombin Preparations Are used in a variety of surgical procedures including, but not limited to: Cardiovascular Surgery Vascular Surgery Neurologic Surgery Orthopedic Surgery General Surgery Gynecological Surgery Estimated that more than 500,000 Americans are exposed each year More than 100 reports in the worlds literature of adverse events related to bovine thrombin exposure in humans
7 First Clinical Use of Thrombin in Humans Clinical Reports of Patients with Major Complications from Antibodies FDA Grandfathers Bovine Thrombin for Commercial Production YEAR
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9 Patient Profile 68 year old WM Myocardial Infarction MVR and CABG x Repeat MVR ,000 units of Bovine Topical Thrombin Repeat MVR ,000 units of Bovine Topical Thrombin Lawson et al., Blood, 1990
10 Patient Labs Test Name Patient Normal Range Value Prothrombin time 17s 11-13s APTT 34s 22-33s Thrombin time >150s 16-20s* (Bovine) Thrombin time 35s 16-20s* (Human) Platelet count 363,000/µL ,000/µL Fibrinogen 465mg/dL mg/dL D dimer 0.2µg/mL <0.2µg/mL FDP 10µg/mL <10µg/mL *Thrombin concentration is adjusted to give a clotting time of 16 to 20 seconds in control plasma, normal is considered ± 3 seconds of control plasma. Lawson et al., Blood, 1990
11 Isolation of Anti-thrombin Antibody from Patient Plasma Lawson et al., Blood, 1990
12 Lawson et al., Blood, 1990
13 Lawson et al., Blood, 1990
14 Lawson et al., Blood, 1990
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17 Important Drug Warning Patients with autoantibodies to bovine thrombin preparations should not be reexposed to these products. The use of topical bovine thrombin preparations has occasionally been associated with abnormalities in hemostasis
18 Immunologic Impact and Clinical Outcomes Following Surgical Exposure to Bovine Thrombin Thomas Ortel, Meg Mercer, Elizabeth Thames, Karen Moore, and Jeffrey H. Lawson Duke University Medical Center Durham, NC Ortel et al. Annals of Surgery, 233(1), 88-96, 2001
19 Study Design Patient Population Number of Individuals Normal Individuals (CONTROL) 50 CABG (Primary) 100 Valve Surgery Patients (Primary) 50 Valve Surgery Patients (Redo) 25 Total 225 Ortel et al. Annals of Surgery, 233(1), 88-96, 2001
20 Blood Sample Collection Preoperative hours Postoperative 4-7 days Postoperative 4-8 weeks Postoperative Patient Outcomes Hemorrhage Thrombosis Infection Wound Healing Transfusion Death Ortel et al. Annals of Surgery, 233(1), 88-96, 2001 Screening Laboratory Assays PT, aptt, and TCT Immunoassays: Anti- Bovine Thrombin Preparation IgG Anti- Bovine Thrombin IgG Anti- Bovine Prothrombin IgG Anti- Bovine factor V IgG Anti- Bovine factor Va IgG Anti- Human Thrombin IgG Anti- Human Prothrombin IgG Anti- Human factor V IgG
21 Results Patient Population Number of Individuals Total Enrolled 226 Incomplete Follow-up 16 Deaths 9 Total Completed 201 Normal Individuals 50 CABG (Primary) 90 Valve Surgery (Primary) 34 Redo Sternotomy 27 Ortel et al. Annals of Surgery, 233(1), 88-96, 2001
22 Pre and Post Operative Response to Bovine Antigens Following Bovine Thrombin Exposure Ortel et al. Annals of Surgery, 233(1), 88-96, 2001
23 Response to Bovine Antigens Percent Developing IgG Antibodies Primary Exposure Prior Exposure All Exposed TBT bfv bfva bfii bfiia Bovine Coagulation Factor? Anti-Bovine Ortel et al. Annals of Surgery, 233(1), 88-96, 2001
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28 The Phylogeny of αgal Natural Anti-αGal Antibodies New World Monkeys Cows αgal Old World Monkeys Mammals Apes Pigs Mice Humans αgal not expressed αgal expressed αgal
29 Contamination - Multiple Proteins and αgal Kd αgal Schoenecker et al. J. Clin. Immun. 20(6):434-44,2000. Thrombin TBT
30 Increase in Elicited IgG 1000 Change In Titer Unexposed Exposed Exposure to Bovine Thrombin? Schoenecker et al. J. Clin. Immun. 20(6):434-44,2000.
31 Conclusions - αgal Topical Bovine thrombin contains αgal. αgal Patients have xenoreactive natural antibodies which are specific for the bovine thrombin preparation proteins. Levels of elicited IgG anti- αgal are significantly increased 6 weeks following exposure to bovine thrombin preparations.
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33 Interesting Xeno-Immunology But what does that have to do with surgery and my patients? The Blue-Devil is in the Details
34 Response to Human Antigens Percent Developing IgG Antibodies Primary Exposure Prior Exposure All Exposed hfv hfii hfiia Human Coagulation Factor? Anti-Human
35 Abnormal Coagulation Studies Percent Abnormal Primary Exposure Prior Exposure 0 PT aptt TCT Total Coagulation Test Antibodies? ABNORMAL CLOTTING TESTS
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37 Pre and Post Operative Response to Bovine Antigens Following Bovine Thrombin Exposure Ortel et al. Annals of Surgery, 233(1), 88-96, 2001
38 Preoperative Antibody Response to Bovine Antigens With (+) and Without (-) Documented Prior Exposure 10% With (+) Without (-)
39 Adverse Events Related to Pre-operative Antibody Levels
40 Multivariant Analysis of Adverse Events
41 Outcomes Antibodies?? OUTCOME Re-exposure Percent Poor Outcomes After Surgery Antibody Negative Antibody Positive P= /136 (34%) Patient Group 11/15 (73%)
42 Bovine Thrombin Black Box Warning
43 What is the Cost of Exposure to Antibody Positive Patients?
44 Conclusions 1. Bovine topical thrombin is highly immunogenic when used in the setting of cardiovascular surgery. 2. Thirty percent of patients exposed to bovine thrombin develop cross-reactive antibodies with human blood coagulation factors. 3. Thirty percent of patients with anti-clotting factor antibodies develop abnormal blood coagulation tests. PEPCI
45 Conclusions 4. Anticoagulation monitoring in patients with anticlotting factor antibodies is problematic. 5. Patients with multiple elevated antibodies prior to surgery are more likely to sustain adverse clinical outcomes. Adjusted Odds Ratio: % Confidence Interval: 1.54, Patients with multiple elevated antibodies prior to surgery appear to have increased cost associated with the procedure which is independent of preoperative comorbid conditions. PEPCI
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47 Vascular Access Patients with circulating antibodies to Bovine Thrombin Preparations 50% of patients with PTFE grafts have these antibodies
48 Greater than 80% of patients with a history of graft thrombosis have elevated antibody levels to bovine thrombin reagents
49 Anticardiolipin Antibody Formation 6 Weeks after CABG Bovine Thrombin Exposure CABG CABG no Thrombin Anticardiolipin antibody IgG titer (GPL) 100 A Anticardiolipin antibody IgM titer (MPL) 100 B Anticardiolipin antibody IgG titer (GPL) 100 C Pre-op Post-op Pre-op Post-op Pre-Op Post-Op Su et al. Journal of Clinical and Laboratory Medicine, 139(6):349-56, 2002.
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51 Exposure of Mice to Bovine Thrombin
52 Antibodies against αgal and Bovine Thrombin (TGEN) Schoenecker et al. American Journal of Pathology, 159(5), , 2001
53 Development of Anti-Clotting Factor Autoantibodies and Abnormal Coagulation Tests Change in Absorbtion Change in Absorbance Unexposed Exposed Unexposed SALINE Exposed TGEN 5 aptt/pt Ratio P= Control Saline TGEN Schoenecker et al. American Journal of Pathology, 159(5), , 2001
54 Antibodies against double stranded DNA and Cardiolipin Schoenecker et al. American Journal of Pathology, 159(5), , 2001
55 Thrombin Control Schoenecker et al. American Journal of Pathology, 159(5), , 2001
56 So What about Floseal?? Lawson et al. Ann Thorac Surg, 79: , 2005
57 Lawson et al. Ann Thorac Surg, 79: , 2005
58 The Immunology of Human Fibrin Sealant and Human Thrombin
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60 Immunogenicity of rhthrombin Anti-rhThrombin Antibody Incidence % (95% CI) Placebo Only N=22 0/21 0 (0 16) Any rhthrombin N=38 1/ (0.1 14)
61 Mouse Exposure to Thrombins from Various Species
62 Case Study (II) 76 year old male admitted for Abdominal Aortic Aneurysm Repair Past Surgical History: 1971 Laryngectomy 1990 Coronary Artery Bypass Grafting x 4 Vessels 1996 Rotator Cuff Repair 1998 Left Radical Nephrectomy -Thrombin spray documented in operative report 1999 Right Radical Nephrectomy -Patient readmitted for right flank hematoma History of transient ischemic cerebrovascular attacks-taking Coumadin
63 Case Study (II) Hospital Course: 12/20/2000 Abdominal Aortic Aneurysm and Bilateral Iliac Artery Aneurysm Repair Intraoperatively received 20,000u Thrombin Spray and Flo-Seal (10,000u Thrombin) as documented in the perioperative nursing record Returned to the ICU a few days postoperatively for increased SOB, episodes of epistaxis, and difficulty breathing requiring re-intubation and mechanical ventilation. Patient developed profound coagulapathy and noted as having a factor V inhibitor
64 Antibodies Response Bovine Antigens - Post-OP Day 5
65 Antibodies Response Human Antigens - Post OP Day 5
66 Recent Case Study Con t) Treatment: Received copious amounts of blood and factor replacement. Continued to have increasing abdominal distension and hemodynamically unstable. CT scan suggested of intraperitoneal bleed 1/7/2001 Received Immune Globulin 10% (Gamimune-N) -IVIG 1/8/2001 Take back to the OR for exploratory laparotomy. Removed 3 liters of mostly clotted blood. Prolonged ICU course with Ventilator Dependence DC d (2/12/2001) to a nursing facility that takes care of ventilator dependent patients
67 Recent Case Study (Con t) Discharge Diagnosis Acquired coagulopathy secondary to factor V inhibitor, presumed secondary topical bovine thrombin.
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