Rodger L. Bick

The clinical spectrum of antiphospholipid syndrome.

Antiphospholipid syndrome (APS) is a disorder characterized by a wide variety of clinical manifestations. Virtually any organ system or tissue may be affected by the consequences of large- or small-vessel thrombosis. There is a broad spectrum of disease among individuals with antiphospholipid antibodies (aPL). Patients may exhibit clinical features suggesting APS but not fulfill the International Criteria for a definite diagnosis. Seronegative APS patients demonstrate typical idiopathic thromboses but aPL are not initially detected. Patients defined with definite APS demonstrate nearly identical sites of venous and arterial thrombosis, regardless of the presence or absence of systemic lupus erythematosus. Microangiopathic APS may present with isolated tissue and organ injury or as the overwhelming thrombotic storm observed in catastrophic APS.

Management of Thrombotic and Cardiovascular Disorders in the New Millenium

Anticoagulants and antithrombotic drugs have played a key role in the prophylaxis, treatment and surgical/interventional management of thrombotic and cardiovascular disorders. There are several newer drugs which are currently developed for the anticoagulant management of cardiovascular diseases in both the medical and surgical indications. These include the low molecular weight heparins (LMWHs), antithrombin agents such as the Hirudin, Hirulog and Argatroban and indirect and direct anti-Xa drugs, represented by Pentasaccharide (Arixtra®) and DX 9065a, respectively. Several other agents such as the natural and recombinant anti-Xa drugs and anti-tissue factor agents are also developed. The antiplatelet agents include Clopidogrel, Cilostazol, Anplag and GP IIb/IIIa inhibitors. For the subcutaneous indications, unfractionated heparin is gradually replaced by the low molecular weight heparins (LMWHs). LMWHs such as the Enoxaparin and Dalteparin are commonly used for the management of acute coronary syndrome. These drugs have been approved for the treatment of unstable angina and are currently undergoing rigorous trials for interventional indications. Arixtras® is also developed for various subcutaneous indications. However, it exhibits lower anticoagulant effects and may not be optimal for intravenous and interventional purposes. At a higher dosage when administered intravenously the LMWHs produce varying degrees of anticoagulation at relatively lower activated clotting times (150-200). Several studies in vascular and cardiovascular interventions have shown that even at a relatively lower anticoagulant level the LMWHs are as effective as unfractionated heparin at the recommended dosages which produce a relatively higher level of anticoagulation (ACT > 200 secs.). Thus, these agents are currently developed for interventional and surgical indications. It should be emphasized that different LMWHs produce different degrees of anticoagulation and should therefore be individually optimized for a given interventional or surgical purposes. At a relatively high dosage the levels of LMWHs can be measured by using the ACT and APTT. When administered with such GP IIb/IIIa inhibitors as the Abciximab, Aggrastat or Eptifibratide, these drugs may require dosage adjustment. However, since the introduction of the front loading of Clopidogrel, the unqualified use of GP IIb/IIIa is debated. LMWHs will find expanded indications in both the medical and surgical management of patients with cardiovascular disorders including atrial fibrillation and congestive heart failure. The only approved anti-Xa drug is represented by a synthetic heparinomimetic, namely, Arixtra®. This drug is given for the prophylaxis of post orthopedic indications. This agent is undergoing additional clinical trials in the management of coronary artery diseases. Because of the dependence on antithrombin III (AT) and the sole anti-Xa effects, it has a narrow therapeutic index and its efficacy in this indication may be limited. Additional clinical trials are needed at this time to validate the clinical potential of this drug. The antithrombin agents (Hirudin, Hirulog and Argatroban) were initially developed for arterial indications. However, these agents are approved as a substitute anticoagulant in patients with heparin induced thrombocytopenia (HIT) and PCI. Currently all of these agents are being developed for surgical and interventional use. However, since there is no available antidote at this time, the development is somewhat limited. The antithrombin agents may be useful in patients with HIT which require further clinical validation. Many other anti-Xa agents are also developed. Most of these can be given parenterally. However, the clinical data is somewhat limited. Similarly, several of the new antiplatelet drugs can be administered parenterally and may be useful in CAD. Since most of these newer anticoagulant and antithrombotic drugs are mono-therapeutic their therapeutic index is rather limited. Only in combination these agents can mimic heparins. At this time it is safe to state that heparin and its LMW derivatives will remain the anticoagulant of choice for cardiovascular indications until these newer agents have been validated in extended dinical trials in polytherapeutc settings.

Antiphospholipid Syndrome in Pregnancy

During the past 5 years the author and his colleagues have assessed carefully 351 women referred for evaluation of thrombosis and hemostasis after they had suffered recurrent miscarriages. This article describes the flow protocol the author and associates follow to maximize success and keep the costs of evaluation of recurrent miscarriage syndrome/infertility at a minimum while providing the best chances for defining a cause and thus providing optimal therapy for successful term pregnancy outcome. It presents the outcomes of the authors protocol and those of others in treating women who have antiphospholipid syndrome and who have suffered recurrent miscarriages.

Controversies and Unresolved Issues in Antiphospholipid Syndrome Pathogenesis and Management

While much is understood concerning the clinical features of patients with antiphospholipid syndrome (APS), many issues remain. The proper designation of patients with definite APS and the correct categorization of patients by both laboratory and clinical features are matters of ongoing debate. Recent proposals have identified new subsets of patients who have many typical features of APS but either do not fit the criteria for a definite diagnosis or have initially negative laboratory tests for antiphospholipid antibodies. Meanwhile, decisions about laboratory tests are based on expert opinion, rather than the results of controlled trials. As for treatment, many guidelines are offered, but few are backed by data from strong clinical trials. This article summarizes the clinical questions remaining to be answered and debates concerning pathogenesis, diagnosis, and management.

Laboratory Evaluation of the Antiphospholipid Syndrome

Antiphospholipid syndrome (APLS) is among the most common acquired blood protein defects that have been identified as leading to thrombosis. This article describes the laboratory diagnosis of APLS, including the detection of lupus anticoagulants, anticardiolipin antibodies, and subtypes of antiphospholipid antibodies.

Treatment options for patients who have antiphospholipid syndromes.

The antiphospholipid thrombosis syndrome, associated with anticardiolipin (aCL) or subgroup antibodies, can be divided into one of six subgroups (I-VI). There is little overlap (about 10% or less) between these subtypes, and patients usually conveniently fit into only one of these clinical types. Although there appears to be no correlation with the type, or titer, of aCL antibody and type of syndrome, the subclassification of thrombosis and aCL antibody patients into these groups is important from the therapy standpoint. This article also reviews the clinical presentations associated with each of these six subgroups.

Contaminant in the Recalled Unfractionated Heparin Preparations: Where is the Problem?

In a recently held press conference, the USFDA (United States Food and Drug Administration) briefed the press that the potential contaminant in the recalled Baxter product is a heparin-like substance. The USFDA commented that heparin-like molecules have been identified using high-tech methods. No specifics regarding these contaminants were given. The manufacturing process of heparin is such that other heparin-like contaminants, such as the dermatan sulfate, heparan sulfate, and chondroitin sulfate are removed effectively. These represent the main contaminants in heparin. However, these contaminants are unlikely to produce any allergic effects, nor the reported adverse reactions, because some of the antithrombotic drugs contain significant amounts of these heparin-like substances. Because of the carbohydrate nature of the contaminants identified by a special method of nuclear magnetic resonance (NMR), it is quite disturbing to note that such contaminants may have inadvertently been added to the recalled heparin. Heparin-like materials have been isolated from the shellfish, marine plants, bones, or skin of mammalian origin for several therapeutic purposes. Although structurally similar, these heparin-like substances exhibit different biochemical and pharmacologic effects. There are also various protein contaminants that can be expected along with the carbohydrates. Thus, it is likely that besides the carbohydrate contaminants, some unknown protein contaminants may also be present in these products. The statement that a heparin-like material was identified using high-tech methods is ambiguous and misleading. Such statements should be specific and clear regarding the methods used and the nature In late January, Baxter Healthcare Corporation voluntarily recalled the following lots of heparin: 107054, 117085, 047056, 097081, 107024, 107064, 107066, 107074, and 107111. These products were labeled as heparin 1000 U/mL in 10 mL vials and heparin 1000 U/mL in 30 mL vials. The reason for this recall was the reported adverse reactions, which were associated with the use of these heparin batches. These included abdominal pain, hypotension, burning, chest pain, diarrhea, dizziness, dyspepsia, dyspnea, arrhythmia, flushing, headache, hyperthyroidism, hypoesthesia, increased lacrimation, loss of consciousness, malaise, nausea, pallor, palpitation, paresthesia, pharyngeal edema, restlessness, vomiting, stomach discomfort, tachycardia, thirst, trismus, unresponsiveness to stimuli, and drug ineffectiveness. These adverse reactions were reported in hundreds of patients treated with these heparins in the USA. Initially 19 deaths were linked with the use of the recalled heparins. However, this figure has now reached 81, and it is likely that it will go even higher. These multiple adverse reactions were later attributed to heparin manufactured by Baxter’s [Deerfield, IL] suppliers using Chinese raw material from SPL Changzhou, China. Subsequently, Baxter recalled all of the remaining lots and doses of its heparin sodium injection multidose, single dose vials, and Hep-lock heparin flush product.

Reduced Immunogenic Potential of Membrane Filtered Bovine Thrombin Preparations for Hemostatic Application

There is concern that exposure to bovine thrombin can result in the development of antibodies, usually against factor V/Va, which can lead to hemostatic abnormalities. It is thought that purer preparations of bovine thrombin might be less immunogenic. Utilizing newer methods including a membrane filtration step, bovine crude thrombin is further purified into thrombin 4A and 4B preparations which exhibit a higher specific activity and are devoid of some of the protein contaminants. Bovine crude thrombin and its purified versions were administered intravenously to individual groups of rabbits using standard immunologic protocols. Antiserum was drawn from each rabbit and the pooled antisera were purified to obtain the IgGs using protein G affinity columns. The results suggest that the reported purification process, including filtration, resulted in the removal of most of the antigens found in crude thrombin, and that none of these preparations generated any detectable antibodies against bovine factor V related antigens.

Comparison of immunogenic potentials of bovine thrombin preparations.

Using a membrane filtration step, bovine crude thrombin was purified into thrombin 4A and 4B preparations. The purpose of this study was to determine whether the improved purity of a bovine thrombin preparation can reduce its overall immunogenic potential and lower the risk of development of factor V antibodies. Bovine crude thrombin and its purified versions, thrombin 4A and 4B, were administered to individual groups of rabbits on days 0, 21, 42, 91, 123, and 151 using standard immunologic methods. Blood was drawn from each rabbit on days 30, 50, 105, 137, and 165, and the pooled antisera from individual groups were purified to obtain the Ig Gs using protein G affinity columns. Using Western blotting, the specificity of each immunoglobulin G collected at the first time point (day 30) and last time point (day 165) was determined. The results of Western blotting using the Ig Gs collected on days 30 and 165 were consistent; both demonstrating that thrombin 4B has the least immunogenic potential among the 3 thrombin preparations tested. Compared with the immunoglobulin Gs collected on day 30, the Ig Gs from day 165 did not show obvious difference regarding their ability to detect antigens in bovine thrombin samples. Neither showed cross-reactivity with human coagulation factors nor the recognition of bovine factor Va antigens. These results suggest that despite the presence of a trace amount of bovine factor Va antigen in bovine thrombin preparations, these contaminants failed to elicit the generation of antibodies against factor Va light chain in rabbit

Antigenic (Immunogenic) Profiling of Bovine Thrombin and its Purified Forms

purified and further processed by ultrafiltration. Analytical studies have shown that the current manufacturing processes are capable of removing significant amounts of extraneous proteins and result in a reduction of factor Va light chain content levels below the limit of detection of the Western blot assay (less than 92 ng/mL, when reconstituted as directed). However, the clinical significance of this reduction of factor Va–related antigen is unknown. The US FDA approval of the revised labeling regarding the purity of Thrombin JMI is a testimonial for the advanced methods that can be used to purify biologicals, such as bovine thrombin, to remove contaminants, which may lead to the generation of antibodies or may trigger other adverse reactions. Although the removal of bovine factor V, factor Va and/or their fragments from the product has been shown, it is highly likely that other potential contaminants and carryover products are also removed in this additional purification step. Therefore, a comparison of the crude thrombin and the 2 purified versions for other potential contaminants, including prothrombin, factors VII and X, may also be helpful because bovine plasma is used as the starting material for the preparation of the crude thrombin. The purified Thrombin JMI will likely show a relatively improved safety profile with much lesser immunogenic potential. Despite the fact that the reported hemostatic abnormalities with the use of this product since the 1940s are relatively rare, 2-6 the improved product may show an even lesser prevalence of these reported adverse reactions. It should remain clear that despite improved purification processes, some biological products may retain inherent immunologic properties leading to the generation of antibodies, as has been the case with many of the highly purified drugs such as recombinant hirudin, aprotinin, and recombinant Bovine thrombin is widely used as a topical hemostatic agent in various surgical procedures. To date, more than 10 million individuals have been exposed to this product. Rare case reports of adverse reactions such as antigenic reactions and apparent generation of antibodies leading to hemostatic disruption have been reported. Isolated reports on the presence of factor Va–related antigen and other contaminant proteins have also been made available. Additionally, case reports have commented on the generation of antibodies to other human coagulation factors. However, bovine thrombin has been remarkably safe than other biological products, such as aprotinin, unfractionated heparin, and erythropoietin. The article by Terrab, et al in this issue of Clinical and Applied Thrombosis/Hemostasis reports on the inclusion of a membrane-filtration step in the manufacturing process, which is performed to remove viruses from chromatographically purified bovine thrombin. It has been found that this step also results in the effective removal of extraneous protein contaminants, in particular factor V, factor Va and/or their fragments from the product. On review of the data summated for this purification process, the US Food and Drug Administration (FDA) has approved a revised labeling for Thrombin JMI (Thrombin, topical bovine, United States Pharmacopenia) to recognize the reduced factor Va light chain content (considered to be a factor for the immunogenic response). The revised labeling states that Thrombin JMI has been chromatographically