Evangelos Litinas

Interpretation of Benefit-Risk of Enoxaparin as Comparator in the RECORD Program: Rivaroxaban Oral Tablets (10 milligrams) for Use in Prophylaxis in Deep Vein Thrombosis and Pulmonary Embolism in Patients Undergoing Hip or Knee Replacement Surgery

The Regulation of Coagulation in Major Orthopedic surgery reducing the Risk of DVT and PE (RECORD) clinical program of rivaroxaban consists of 4 phase III clinical trials comparing rivaroxaban with enoxaparin for the prevention of venous thromboembolism (VTE) in patients undergoing either total hip or total knee replacement surgery. Despite the comprehensive and extensive nature of this program, it had some logistic issues that included the dosing of the enoxaparin which was not only inconsistent with the recommendations but the dosages used were not optimal. The duration of treatment while consistent with rivaroxaban did vary with enoxaparin and was somewhat short. The bleeding definitions and safety evaluations were not consistent in accordance with the current recommendations. Moreover, the RECORD program has no power to show differences in major bleeding. The cardiovascular rebound phenomenon should have been adequately addressed and may require additional clinical validation to establish the safety of rivaroxaban. Although the US Food and Drug Administration (FDA) advisory committee has recommended approval of rivaroxaban, the reported analysis strongly suggests additional clinical validation on the claimed benefit/risk ratio of this monotherapeutic anticoagulant.

In Vivo Neutralization of Unfractionated Heparin and Low-Molecular-Weight Heparin by a Novel Salicylamide Derivative

Unfractionated heparin (UFH) and low-molecular-weight heparin (LMWH) are widely used anticoagulants for surgical and interventional use. Currently, the anticoagulant and bleeding effects of heparin are neutralized by protamine sulfate. There are several problems associated with the use of protamine sulfate, including allergic reactions, cardiovascular effects, heparin rebound, and incomplete neutralization of LMWHs. The objective of this investigation is to characterize the effectiveness of a novel salicylamide-derived heparin antagonist, PMX 60056, in neutralizing the antithrombotic and bleeding effects of UFH and LMWHs. Animals were first anticoagulated using an intravenous injection of UFH or LMWH followed by protamine sulfate or PMX 60056. Established animal models of hemorrhage (rat-tail transection) and thrombosis (jugular vein clamping) were then performed. Blood samples were collected for ex vivo analysis using activated partial thromboplastin time (aPTT), anti-Xa, and anti-IIa assays. We demonstrate that PMX 60056 neutralized the antithrombotic, anticoagulant, and bleeding effects of heparins as effectively as protamine sulfate and may be slightly more efficacious against LMWHs. These results suggest that PMX 60056 may provide an improved approach for the neutralization of UFH and LMWHs.

A comparison of the pharmacodynamic behavior of branded and biosimilar enoxaparin in primates.

Pharmacodynamic behavior of branded and biosimilar enoxaparin was compared in a crossover study in primates. Blood samples collected at baseline and at 1, 4, 6, and 28 hours post-subcutaneous administration of Lovenox or Fibrinox were evaluated using clot-based and amidolytic assays. Anti-Xa levels following Fibrinox and Lovenox administration were not different. Anti-IIa levels were significantly higher in Lovenox-treated animals 1 to 6 hours post-administration. Higher drug levels were measured by Heptest in Fibrinox-treated animals from 4 to 6 hours. Pharmacokinetic differences were not observed using anti-Xa or Heptest assays. The area under the curve (anti-IIa) following Lovenox treatment was significantly larger than following Fibrinox treatment. When drug levels (anti-IIa) were plotted against anti-Xa or Heptest drug levels, a hysteretic relationship which was distinct for Fibrinox- and Lovenox-treated primates was observed suggesting a lack of bioequivalence for the low-molecular-weight heparin tested. In vivo behavior is an important consideration for defining pharmacoequivalence of complex biologic drugs.

Comparative Anticoagulant and Platelet Modulatory Effects of Enoxaparin and Sulodexide

Sulodexide represents a novel antithrombotic agent with multiple sites of action on blood coagulation and vascular processes. The purpose of this study was to compare sulodexide and enoxaparin on anticoagulant effects, tissue factor (TF)-induced activation of platelets, inhibition of microparticle generation and to investigate their effect on heparin-induced platelet aggregation (HIPA). Sulodexide was compared with enoxaparin at equigravimetric concentrations. When compared to enoxaparin, sulodexide produced a stronger anticoagulant effect in the prothrombin time (PT), activated partial thromboplastin time (APTT), Heptest, and thrombin time (TT) assays. In addition, sulodexide had a stronger inhibitory effect on TF-mediated microparticle generation (IC50 = 2.8 μg/ mL), P-selectin expression (IC50 = 4.8 μg/ml), and platelet aggregate formation (IC50 = 8.5 μg/mL) compared to higher IC50 values with enoxaparin. Sulodexide and enoxaparin exhibited a similar effect on heparin-induced thrombocytopenia (HIT) antibody-mediated platelet activation HIPA assays. These results suggest that sulodexide is a relatively stronger anticoagulant agent than enoxaparin. Sulodexide is subcutaneously absorbed. Its ability to inhibit TF-mediated platelet activation may contribute to the observed therapeutic effects of sulodexide in microvascular vasculopathy such as diabetic nephropathy. These results also suggest that inhibition of TF activation of platelets by sulodexide may be independent of its anticoagulant effects. These results warrant further investigation of sulodexide in additional preclinical and clinical studies.

Inflammatory Biomarker Profiling in Elderly Patients With Acute Hip Fracture Treated With Heparins

Hip fracture is common in the elderly patients with associated high risk of venous thromboembolic complications. Pathogenic activation results in the generation of various surrogate markers in plasma. This study is designed to identify unique biomarkers in elderly patients with hip fracture using protein chip array enzyme-linked immunosorbent assay (ELISA) methods. Plasma from a randomized hip fracture study (PK-532; n = 341) treated with either enoxaparin (40 mg once daily) or unfractionated heparin (UFH; 5000 IU twice daily) were collected prior to and at 1, 3, 5, and 7 days. A total of 52 samples were analyzed using proteomic surface-enhanced laser desorption/ ionization-time of flight (SELDI-TOF) mass spectrometry to identify unique biomarkers in the molecular weight range of 0 to 150 kd. Twenty-nine healthy volunteer’s and pooled plasma from total hip replacement/total knee replacement patients with a unique biomarker at 11.9 kd were used as quality controls. In the 29 healthy individuals, the biomarker profile did not reveal the presence of any unique peak in comparison to the reference normal human plasma (NHP). Plasma obtained prior to surgery exhibits unique biomarkers in 4 of 52 (7.6%) of the samples. On day 1 postoperatively, 41 of 51 (80.3%) showed a distinct peak at 11.9 kd. On day 3, 43 of 49 (87.8%) patients showed the presence of this biomarker most often at its strongest intensity. In all, 22 of 44 (50%) showed this biomarker on day 5 and 4 of 23 (17.9%) on day 7. C-reactive protein (CRP), tumor necrosis factor α (TNF-α), and serum amyloid A were also increased after surgery. Tissue factor pathway inhibitor (TFPI) antigen levels were increased due to the treatment modalities.

Upregulation of Inflammatory Mediators in End-Stage Renal Disease as Measured Using Biochip Array Technology

Systemic vascular changes contribute to both the pathogenesis and thrombotic comorbidities of end-stage renal disease (ESRD). This study aims to profile various biomarkers and better understand their role in the pathogenesis of ESRD. Plasma samples from 49 patients with ESRD and 56 control individuals were analyzed for markers for inflammation, specifically C-reactive protein (CRP), tumor necrosis factor receptor 1 (TNFR1), neutrophil gelatinase-associated lipocalin (NGAL); thrombomodulin (TM); neuron-specific enolase (NSE), and thrombosis-D-dimer (DD). Compared to controls, all markers studied showed a statistically significant upregulation in patients with ESRD. These results indicate a polypathologic process in patients with ESRD, leading to cardiovascular and cerebrovascular events. However, the clinical significance of previously untested markers, such as TNFR1, NGAL, and NSE, still needs to be further explored. This study further validates the role of endothelial damage and endogenous thrombotic processes in ESRD as evidenced by the increased levels of TM and DD.

Concerns on Generic Enoxaparin use in Acute Coronary Syndrome

Correspondencia: Eduardo Ramacciotti • Vascular Surgery University of Michigan Medical School A570D, MSRB II, 1150 W. Medical Center Dr, Ann Arbor, MI 48109 E-mail: eduardoramacciotti@gmail.com Articulo recibido en 30/07/10, revisado recibido en 30/07/10; aceptado en 16/08/10. Palabras clave Enoxaparina/efectos adversos, medicamentos genericos, sindrome coronario agudo. antiplaquetarios, con dosajes variables. De esa forma, ese procedimiento tambien representa un regimen de tratamiento complicado, el cual presenta un bajo margen de seguridad..In addition, the immunogenicity of different branded and generic products may be contributory to the safety outcome with these agents. Although patients are treated for up to 10 days, antibodies can last for up to 6 to 8 weeks and can alter the anticoagulant efficacy of these compounds. However, such data are not available. Some of the concerns related to the safety of generic enoxaparin usage in ACS are given below:1. A full dose of 1 mg/kg BID resulting in high circulating levels may have a higher risk of bleeding due to several complicating factors.2. Interactions with antiplatelet agents, which may vary with different drugs and drug combinations.3. Interaction with thrombolytic agents may also vary, depending upon the type of the agent used and the timing.4. Potential unknown interactions with newer anticoagulants such as dabigatran, rivaroxaban and apixaban. 5. Drug over-accumulation in renal-impaired patients.6. Conversion from clinical treatment of ACS to PCI or CABG and the consequences of potential interactions between on-board enoxaparin and UFH IV, while on full dosage7. The primary outcomes and incidence of peri-procedural complications, such as primary ischemic outcomes (MI), bleeding and catheter thrombosis have been addressed for the branded enoxaparin and UFH. These concerns, however, have not been adequately studied for the branded and generic products and may result in significant differences between these products. This requires clinical validation in adequately powered trials.8. Therefore, at this time, the use of generic enoxaparin products in this critical indication is not recommended. Moreover, simply carrying out studies of bio-equivalence and limited VTE prophylaxis in healthy volunteer with relatively lower doses of 40 mg OD SQ are not adequate to justify the use of generic agents in such critical cardiovascular indications as ACS.A previous study has compared the efficacy and safety of a branded enoxaparin, namely Lovenox, with a generic form of this agent

Considerações sobre o uso de enoxaparina genérica em síndrome coronariana aguda

Correspondencia: Eduardo Ramacciotti • Vascular Surgery University of Michigan Medical School A570D, MSRB II, 1150 W. Medical Center Dr, Ann Arbor, MI 48109 E-mail: eduardoramacciotti@gmail.com Articulo recibido en 30/07/10, revisado recibido en 30/07/10; aceptado en 16/08/10. Palabras clave Enoxaparina/efectos adversos, medicamentos genericos, sindrome coronario agudo. antiplaquetarios, con dosajes variables. De esa forma, ese procedimiento tambien representa un regimen de tratamiento complicado, el cual presenta un bajo margen de seguridad..In addition, the immunogenicity of different branded and generic products may be contributory to the safety outcome with these agents. Although patients are treated for up to 10 days, antibodies can last for up to 6 to 8 weeks and can alter the anticoagulant efficacy of these compounds. However, such data are not available. Some of the concerns related to the safety of generic enoxaparin usage in ACS are given below:1. A full dose of 1 mg/kg BID resulting in high circulating levels may have a higher risk of bleeding due to several complicating factors.2. Interactions with antiplatelet agents, which may vary with different drugs and drug combinations.3. Interaction with thrombolytic agents may also vary, depending upon the type of the agent used and the timing.4. Potential unknown interactions with newer anticoagulants such as dabigatran, rivaroxaban and apixaban. 5. Drug over-accumulation in renal-impaired patients.6. Conversion from clinical treatment of ACS to PCI or CABG and the consequences of potential interactions between on-board enoxaparin and UFH IV, while on full dosage7. The primary outcomes and incidence of peri-procedural complications, such as primary ischemic outcomes (MI), bleeding and catheter thrombosis have been addressed for the branded enoxaparin and UFH. These concerns, however, have not been adequately studied for the branded and generic products and may result in significant differences between these products. This requires clinical validation in adequately powered trials.8. Therefore, at this time, the use of generic enoxaparin products in this critical indication is not recommended. Moreover, simply carrying out studies of bio-equivalence and limited VTE prophylaxis in healthy volunteer with relatively lower doses of 40 mg OD SQ are not adequate to justify the use of generic agents in such critical cardiovascular indications as ACS.A previous study has compared the efficacy and safety of a branded enoxaparin, namely Lovenox, with a generic form of this agent

Relative Purity of Different Bovine Thrombin Preparations

The aim of this study was to compare the relative purity of bovine crude thrombin and its purified forms, namely, thrombin 4A and thrombin 4B (the products of King Pharma, Middleton, Wisconsin) by virtue of the detection of bovine prothrombin-related antigens in these preparations. Bovine prothrombin was administered intravenously to 3 individual rabbits on days 0, 21, 42, 91, 123, and 151 using standard immunologic method. Blood was drawn from each rabbit on days 30, 50, 105, 137, and 165, and the pooled antisera from 3 rabbits were purified to isolate immunoglobulin G (IgG) using protein G affinity columns. Using Western blotting method, serially diluted bovine crude thrombin, thrombin 4A, and 4B preparations were probed using the prothrombin IgGs obtained from each time point to explore prothrombin-related antigens in these preparations. The results revealed that compared with the prothrombin IgG collected on day 30, the IgGs collected on days 50 to 165 showed a time-dependent increase in their ability to detect the prothrombin-related antigens in 3 bovine thrombin preparations studied. The lowest amount of crude thrombin, thrombin 4A, and 4B preparations that prothrombin IgG could detect was 0.125, 10, and 20 U, respectively. The rank order of the number of immunoreactive bands detectable in 3 bovine thrombin preparations probed by the prothrombin IgGs collected from any given time point was always the same: crude thrombin > thrombin 4A > thrombin 4B. The results indicate that thrombin 4B preparation contains the least amount of antigens detectable by prothrombin IgG, suggesting that relatively thrombin 4B represents the most purified thrombin preparation among the 3 thrombin preparations studied.

Interactions of Oversulfated Chondroitin Sulfate (OSCS) From Different Sources With Unfractionated Heparin

In 2008, oversulfated chondroitin sulfate (OSCS) was identified as the main contaminant in recalled heparin. Oversulfated chondroitin sulfate can be prepared from bovine (B), porcine (P), shark (Sh), or skate (S) origin and may produce changes in the antithrombotic, bleeding, and hemodynamic profile of heparins. This study examines the interactions of various OSCSs on heparin in animal models of thrombosis and bleeding, as well as on the anticoagulant and antiprotease effects in in vitro assays. Mixtures of 70% unfractionated heparin (UFH) with 30% OSCS from different sources were tested. In the in vitro activated partial thromboplastin time (aPTT) assay, all contaminant mixtures showed a decrease in clotting times. In addition, a significant increase in bleeding time compared to the control (UFH/saline) was observed. In the thrombosis model, no significant differences were observed. The OSCSs significantly increased anti-Xa activity in ex vivo blood samples. These results indicate that various sources of OSCS affect the hemostatic properties of heparin.