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Background Andexanet alfa (AnXa; Andexxa) was developed as a specific antidote for direct and indirect factor Xa (FXa) inhibitors. It has been approved in the US for the reversal of apixaban and rivaroxaban anticoagulation when necessary due to life-threatening or uncontrolled bleeding. Generation 2 manufacturing process of AnXa is promising for providing adequate global supply of AnXa and awaits comprehensive regulatory evaluation for comparability to the current commercial process. Purpose: In this ongoing porcine polytrauma study, we investigated and compared efficacy and safety of the first and second generation AnXa product for the reversal of apixaban anticoagulation. Method: In 35 anesthetized male pigs, a polytrauma consisting of bilateral femur fractures and a standardized blunt liver injury was inflicted after 3 days of apixaban intake (20 mg/day). 12 min after the polytrauma, pigs were randomly allocated to receive (n=7 per group) either 1000 mg bolus of 2nd generation AnXa followed by 2-hour (h) infusion of normal saline (2nd G AnXa B) or 1000 mg bolus immediately followed by infusion of 1200 mg 2nd generation AnXa in 2 h (2nd G AnXa B+). These two groups that received 2nd generation AnXa were compared with the cohorts from our previous study following precisely the same protocol with bolus and 2 h infusion of normal saline (Control group), 1000 mg bolus of 1st generation AnXa followed by 2 h infusion of normal saline (1st G AnXa B), and 1000 mg bolus immediately followed by 2 h 1200 mg infusion of 1st generation AnXa (1st G AnXa B+). Animals were then observed for 5 h or until death. The blood loss (BL) was the primary outcome. Hemodynamics and coagulation profile comprised apixaban (anti FXa activity and rotational thromboelastometry (ROTEM). Further D-dimer levels and thrombin generation (TG) were measured. Right after the premature death or at the end of the observation period, internal organs were obtained and examined for the occurrence of thromboembolic incidents. Two-way ANOVA (mean ± SD) and log-rank tests were performed to analyze the data, and the statistical significance was set at p Results: Three days of oral apixaban administration resulted in comparable anti-FXa activity levels before the trauma (195.4 ± 90.9 ng/mL, Figure 1). The increased anti-FXa activity also prolonged the clotting time (CT) on extrinsically activated ROTEM channel EXTEM (43.6 ± 5.5 to 110.1 ± 28.2 sec). BL at 12 min after the trauma was similar among the study groups (856 ± 118 mL). Anticoagulation with apixaban in control group resulted in a total BL of 3403 ± 766 mL (p Conclusion: This animal polytrauma model is the first to show that 2nd generation AnXa is as effective and well tolerated as the 1st generation in reducing blood loss and improving survival in apixaban anticoagulated pigs with massive hemorrhage. Disclosures Grottke: Boehringer Ingelheim: Consultancy, Research Funding; CSL Behring: Consultancy, Research Funding; Portola: Consultancy, Research Funding; Octapharma: Consultancy, Research Funding; Sanofi: Honoraria; Pfizer: Honoraria. Conley:Portola: Employment. Rossaint:CSL Behring: Honoraria; Boehringer Ingelheim: Honoraria.

Background: Prompt reversal of factor Xa (FXa) anticoagulation is essential in clinical situations with major bleeding or urgent surgery. Andexanet alfa (Andexxa, AnXa) is approved in the US for reversal of rivaroxaban or apixaban anticoagulation when necessary due to life-threatening or uncontrolled bleeding. In ANNEXA-4 interim report, ~85% of patients had good or excellent hemostasis after AnXa treatment (bolus + 2-h infusion). However, the minimum extent and duration of anticoagulation reversal required for a clinically relevant reduction in bleeding in humans have not been fully characterized. Purpose: The impact of escalating doses of AnXa on reversing bleeding in a porcine polytrauma model with apixaban anticoagulation was investigated. Method: After ethical approval, 40 anesthetized male pigs underwent a standardized polytrauma by blunt liver injury and bilateral femur fractures following 3-day apixaban administration (20 mg/day). 12 min post-trauma, animals were randomized (n=8 per group) to 5 groups: 250 mg bolus (AnXa 250 B), 500 mg bolus (AnXa 500 B), 250 bolus + 300 mg 2-h infusion (AnXa 250 B+), 500 mg bolus + 600 mg 2-h infusion (AnXa 500 B+), or vehicle (control). Blood loss (BL), hemodynamics and coagulation profiles were monitored over 5 h or until death; the primary outcome was total BL. Coagulation profiles consisted of apixabananti-FXa activity, global coagulation assays (PT, aPTT), rotational thromboelastometry (ROTEM) and thromboelastography (TEG). For differentiated risk assessment, thrombin generation, thrombin-antithrombin (TAT) and D-Dimer levels were measured. Data were analyzed by two-way ANOVA and log-rank tests (mean ± SD). Results: Before the polytrauma, anti-FXa levels were 178.2 ± 52.7 ng/mL (similar among 5 groups). Trauma led to comparable BL at 12 min after injury (814 ± 79 mL). Compared with control (3464 ± 719 mL) and AnXa 250 B (3155 ± 364 mL; p=0.99 vs control), a significant reduction in total BL was observed in AnXa 250 B+ (2113 ± 297 mL; p Conclusion: Dose-dependent reversal of apixaban by AnXa in this animal model was associated with decreased BL and mortality. The results indicate that AnXa regimens that decreased anti-FXa activity below a certain level (approx. 50 ng/mL), whether temporarily or sustained, was sufficient to prevent exsanguination in this animal model. This remaining low level of anticoagulation after apixaban reversal does not prevent activation of coagulation, allowing for sufficient hemostasis. Our results warrant further investigation of the extent and duration of reversal of FXa inhibition required to achieve adequate hemostasis in humans. Figure 1: Anti-FXa activity in control, AnXa 250 B and AnXa 250 B+ groups (A). Anti-FXa activity in control, AnXa 500 B and AnXa 500 B+ groups (B). Initially n=8 per group. *P Disclosures Grottke: Pfizer: Honoraria; Sanofi: Honoraria; Octapharma: Consultancy, Research Funding; Portola: Consultancy, Research Funding; CSL Behring: Consultancy, Research Funding; Boehringer Ingelheim: Consultancy, Research Funding. Conley:Portola: Employment. Rossaint:CSL Behring: Honoraria; Boehringer Ingelheim: Honoraria.

Introduction: Idarucizumab is a monoclonal humanized antibody fragment (Fab), designed to reverse dabigatran anticoagulation. It is approved for urgent reversal of dabigatran anticoagulant effects in massive bleeding or prior to urgent interventions. It is administered intravenously (i.v.), either as a bolus or a short infusion (2x2.5 g). In hemorrhagic shock gaining i.v. might be challenging, and alternative methods such as intraosseous (i.o.) administration are an alternative life-saving procedure. Due to the molecular size of idarucizumab (50 kDa), the efficacy and safety, including the passage from the i.o. compartment to circulation, are unknown. In this study we investigated the efficacy and the prothrombotic risk of idarucizumab, given i.v. and i.o. in a large animal polytrauma model. Methods: After ethical approval, 21 male pigs were administered oral dabigatran etexilate for 3 days (30 mg/kg twice daily). Following anesthesia, to achieve constant plasma levels, further dabigatran was infused on day 4 (0.77 mg/kg/h for 30 min plus 0.2 mg/kg/h for 60 min). Animals were randomized 1:1:1 to receive 1) i.o. placebo (control group), 2) i.o. idarucizumab (60 mg/kg), or 3) i.v. idarucizumab (60 mg/kg). The standardized polytrauma consisted of a blunt liver injury and a femur fracture. After trauma and hemorrhagic shock, animals were resuscitated with Ringer's solution. Idarucizumab (i.o. or i.v.) or placebo (i.o.) were administered 15 min after trauma, either via an i.o. access (left tibial plateau, EZ-IO¨), or an i.v. access. Animals were observed up to 240 min after trauma, and total blood loss was assessed after the observation period or animal's death. Coagulation variables included dabigatran plasma concentration, global coagulation assays (PT, aPTT) and thromboelastometry. For a differentiated risk assessment, thrombin generation and thrombin-antithrombin (TAT) levels, indicating the extent of coagulation activation, and D-Dimers, as markers of thromboembolic risk, were obtained. Hemodynamic monitoring comprised cardiac output, mean arterial and mean pulmonary arterial pressure. For statistical analysis, ANOVA (mean ± SD) and the log-rank test for survival were used. Results: Prior to trauma, all animals showed comparable plasma concentration of dabigatran (583 ± 139 ng/mL). Following idarucizumab administration, dabigatran plasma concentration was not detectable in both groups. Binding of dabigatran was associated with a normalization of whole blood and plasma coagulation variables. Total blood loss post-injury was comparable between both idarucizumab groups (i.o.: 1085 ± 102 mL, i.v.: 1142 ± 125 mL; P>0.05) with 100% survival. In contrast, control animals (no anticoagulation and no idarucizumab) exhibited the highest total blood loss, which was significantly higher than in idarucizumab treated pigs (4065 ± 557 mL, P0.05). Clinically no signs of thromboembolic adverse were observed. Conclusion: The route of application (i.o. or i.v.) of idarucizumab has no impact on the efficacy of the reversal of dabigatran. Despite the size of idarucizumab, therapy can be safely applied via an i.o. access. In patients with severe bleeding and hemorrhagic shock under dabigatran application, i.o. administration of idarucizumab is a safe alternative. Disclosures Honickel: Böhringer Ingelheim Pharma GmbH & Co KG: Other: travel support. ten Cate:Böhringer Ingelheim: Consultancy, Honoraria, Research Funding; CSL Behring: Research Funding; Stago: Consultancy; Bayer: Consultancy, Honoraria, Research Funding; Philips: Research Funding; Pfizer: Consultancy, Honoraria, Research Funding; Leo Pharma: Consultancy, Honoraria. Rossaint:CSL Behring: Consultancy, Honoraria; Böhringer Ingelheim: Consultancy, Honoraria; Novo Nordisk: Consultancy, Honoraria. Grottke:Böringer Ingelheim: Consultancy, Honoraria, Research Funding; Novo Nordisk: Research Funding; Biotest: Research Funding; CSL Behring: Consultancy, Honoraria, Research Funding; Nycomed: Research Funding; Bayer: Consultancy, Honoraria; Octapharma: Consultancy, Honoraria; Sanofi: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Portola: Consultancy, Honoraria.

Despite increasing use of prothrombin complex concentrate (PCC) to treat hemorrhage-associated coagulopathy, few studies have investigated PCC in trauma, and there is a particular lack of safety data. This study was performed to evaluate PCC therapy in a porcine model of coagulopathy with blunt liver injury. Coagulopathy was induced in 27 anesthetized pigs by replacing approximately 70% blood volume with hydroxyethyl starch 130/0.4 and Ringer's lactate solution; erythrocytes were collected and retransfused. Ten minutes after trauma, animals randomly received PCC (35 or 50 IU/kg) or saline. Coagulation parameters including thromboelastometry, thrombin generation, and blood loss were monitored for 2 hours. Internal organs were examined macroscopically and histologically to determine the presence of emboli and assess liver injury. Total blood loss was significantly lower and survival was higher in both PCC groups versus the control group (P < .05). These outcomes appeared to be dose-independent. Thromboembolism was found in all animals treated with 50 IU/kg PCC; 44% also showed signs of disseminated intravascular coagulation. Liver injury was similar in all animals. In conclusion, 35 IU/kg PCC safely improved coagulation and attenuated blood loss. However, the higher dose of PCC (50 IU/kg) appeared to increase the risk of thromboembolism and disseminated intravascular coagulation.

Background: Reversal of dabigatran anticoagulation in traumatized patients with massive bleeding using idarucizumab (IDA) or prothrombin complex concentrate (PCC) together with other resuscitation measures will be first line treatment. In cases of continuous bleeding and/or elevated dabigatran levels, further hemostatic therapy with coagulation factors (tranexamic acid: TX, fibrinogen concentrate: FGN) may be required. This study tested the safety and efficacy of IDA or PCC, given as first or second line therapy in a two hit polytrauma model under dabigatran anticoagulation. A multimodal approach using TX plus FGN was also tested. In addition to the primary endpoint, reduction in blood loss (BL), a panel of coagulation parameters and the safety of these hemostatic measurements was investigated. Methods: Dabigatran etexilate (30 mg/kg bid) was given to 28 male pigs for 3 days after ethical approval. On day 4, pigs were anesthetized and given a dabigatran infusion before blunt liver injury and bilateral femur fractures. Animals were randomized to receive either 60 mg/kg IDA or 50 U/kg PCC after the first injury. One hour later these animals received the opposite treatment post second liver injury. In a second step, TX (20 mg/kg) plus FGN (100 mg/kg) were added to hemostatic therapy (IDA or PCC) after the first injury, and received the opposite hemostatic therapy (IDA 60 mg/kg or PCC 50 U/kg) after the second liver injury. BL, hemodynamic and coagulation parameters were monitored over 5 h or until death. Results: IDA as first line treatment resulted in a significant reduction in BL (IDA: 1040±202 mL) as compared to PCC (1389 ±194 mL) 60 min post injury. Despite increasing blood loss following the second trauma, the difference between groups remained significant (IDA-PCC: 1556 ± 205 mL, PCC-IDA: 1981±361 mL, P Conclusion: Under conditions of ongoing blood loss after polytrauma and dabigatran anticoagulation, both IDA and PCC prevented exsanguination, although therapy with IDA was more effective. This can be explained by differences in mechanisms, IDA binds dabigatran and inhibits its anticoagulant effect, whereas PCC has no impact on dabigatran anticoagulation but nonspecifically enhances thrombin generation. Their different effects on coagulation parameters also reflect this. Moreover our data imply that clinically used multimodal hemostatic therapy with TX plus FGN and PCC or IDA appears safe under these conditions. Disclosures Grottke: NovoNordisk: Research Funding; Portola Pharmaceuticals: Consultancy; CSL Behring: Honoraria, Research Funding; Boehringer Ingelheim: Consultancy, Honoraria, Research Funding; Biotest: Research Funding. Rossaint:CSL Behring: Research Funding; Bayer Healthcare: Honoraria; Boehringer Ingelheim: Honoraria, Research Funding.

Background: A proportion of trauma patients presenting with coagulopathy also receive anticoagulant therapy, which can exacerbate coagulopathy and increase bleeding. Emergency reversal of anticoagulation therapy may require therapy with specific reversal agents, as with idarucizumab for dabigatran or non-specific reversal agents, such as prothrombin complex concentrates (PCC). Methods: Using a lethal polytrauma model under dabigatran anticoagulation, we investigated the mechanism and procoagulant properties of a four-factor PCC or idarucizumab in dabigatran-anticoagulated pigs following multiple injuries. Blood markers of coagulation associated with procoagulant effects, platelet activation and hyperfibrinolysis were determined over time and related to blood loss. After ethical approval, dabigatran etexilate was given orally for 3 days and iv on day 4 to 18 pigs after anesthesia. Animals were randomized to receive idarucizumab (60 mg/kg), PCC (50 IU/kg) or saline (control), administered 15 minutes after bilateral femur fracture and blunt liver injury. Plasma dabigatran was determined using diluted thrombin time. Thrombin-antithrombin (TAT) complexes and fibrinopeptide A (FPA) were quantified using ELISA and D-Dimer with a clotting assay. Thrombin generation (ETP) in plasma was measured using Calibrated Automated Thrombogram. Platelet function was evaluated as aggregation using platelet-rich plasma in response to tissue factor, collagen, or adenosine diphosphate (ADP) agonism using light transmission aggregometry. Results: Mean dabigatran levels were 543±186 ng/ml prior to injury. Blood loss was 4056±537 mL after saline treatment (control), 1822±222 mL after PCC (P Conclusion: Although four-factor PCC (50 IU/kg) is effective in reducing blood loss, its efficacy is less than therapy with idarucizumab (60 mg/kg). Most importantly the activation of coagulation is explained by different mechanisms. Supernormal activation of coagulation (e.g. TAT, FPA, ETP) was evident after PCC administration, whereas idarucizumab normalized ETP to baseline values. Since thrombin is a potent activator of platelets via PAR, restoration of active thrombin after PCC also partially restored aggregation. In contrast, idarucizumab binds dabigatran and forms an irreversible complex, inactivating dabigatran in plasma, thereby restoring hemostatic function. This difference may influence the choice of therapy when idarucizumab is clinically available. Disclosures van Ryn: Boehringer Ingelheim: Employment. Grottke:NovoNordisk: Research Funding; Portola Pharmaceuticals: Consultancy; CSL Behring: Honoraria, Research Funding; Boehringer Ingelheim: Consultancy, Honoraria, Research Funding; Biotest: Research Funding.

Background: The most frequent side effect associated with anticoagulant therapy is bleeding. Testing reversal strategies in patients is difficult due to the unpredictability, rarity and heterogeneity of these events. Currently methods to reverse this bleeding are being tested in a variety of clinical settings in volunteers by using reversal of anticoagulation as a marker of reversal of bleeding in patients. However, the limitation of volunteer studies is the lack of understanding between the assay being used to measure anticoagulation and if its reversal has any relevance for bleeding reversal in patients. A marker of bleeding that was safe to use in volunteers and also relevant for predicting reversal of bleeding in a patient would be advantageous. Objective: The feasibility of two markers of bleeding - fibrin formation (fibrinopeptide A, FPA) in blood as it exits a wound and blood loss measured via a washed blood method, both at a superficial wound site - were tested in pigs under high dabigatran anticoagulation. Reversal of dabigatran anticoagulation and bleeding following trauma injury were evaluated with idarucizumab and outcomes compared to the markers of bleeding from superficial wound. Methods: After ethical approval dabigatran etexilate (30 mg/kg p.o. twice daily, n=24) or placebo (n=6, no dabigatran anticoagulation) was administered to male pigs for 3 days. On Day 4, the pigs were anesthetized and given a 90 min infusion of active dabigatran. A standardized blunt liver injury (0 min) was performed and animals underwent hemorrhagic shock, 15 min later were randomized to receive idarucizumab (30, 60 or 120 mg/kg) or vehicle. Blood loss was measured at defined time points over 4 hrs post injury. At the same time points a standardized cut was made on the inner side of the ear using an adult Surgicutt® device, one cut for each method. Shed blood was collected from the wound site as it emerged over 4 min, placed in protease inhibitor solution and processed for FPA measurement by ELISA. For washed blood method, blood as it exited the wound in 10 sec intervals was diluted in saline into 96 wells and measured photometrically. AUC over 15 min was recorded as blood loss. Blood samples were collected over time to measure active dabigatran by diluted thrombin time (dTT). Data shown as mean ± SE. Results: Cumulative blood loss was 630 ± 56 mL in the placebo group and 2977 ± 129 mL in the dabigatran-treated group. There was a dose-dependent reduction in blood loss by 47% (1586 ± 253 mL), 64% (1064 ± 40 mL) and 62% (1140 ± 44 mL) following treatment with 30, 60 and 120 mg/kg idarucizumab (p FPA in blood from the wound in dabigatran-treated groups was decreased ~74% vs non-anticoagulated group (211.2 ± 86 vs 53.7 ± 13 ng/mL). FPA increased by 2.6-fold (136.3 ± 96 ng/mL), 1.4-fold (103.9 ± 59 ng/mL) and 6.2-fold (251.9 ± 139 ng/mL) over baseline following 30, 60 and 120 mg/kg idarucizumab, respectively. At 240 min, FPA levels were 12%, 25% and 52% of non-anticoagulated animals with increasing idarucizumab doses and correlated with levels of dabigatran at 240 min. Washed blood loss (AUC) in dabigatran-treated animals was increased ~7-fold prior to injury vs non-anticoagulated groups (2110 ± 420 vs 309.1 ± 11.2 OD*sec). There was a dose-dependent reduction in washed blood loss by 2.5% (1682 ± 688 OD*sec), 48% (889 ± 304 OD*sec), and 79% (370 ± 97 OD*sec) 30 min following 30, 60 and 120 mg/kg idarucizumab vs dabigatran-treatment (1724 ± 298 OD*sec). Washed blood loss was comparable to the non-anticoagulated group (186 ± 59 vs 281 ± 201 OD*sec) with 120 mg/kg idarucizumab after 240 min, also correlating with lack of dabigatran anticoagulant activity. Conclusions: This study shows that bleeding and its reversal with a dabigatran-specific antidote can be evaluated using alternative methods that measure FPA levels in shed blood and washed blood loss at non-trauma superficial wound sites. The reversal of blood loss and FPA generation correlated with reversal of bleeding from a trauma wound. These methods may serve as potential markers in experimental bleeding and trauma models to sensitively assess blood loss and to also monitor reversal agents. Disclosures Grottke: Boehringer Ingelheim : Consultancy, Research Funding; CSL Behring: Research Funding. Honickel:Boehringer Ingelheim : Travel support Other. Schurer:Boehringer Ingelheim Pharma GmbH & Co KG: Employment. van Ryn:Boehringer Ingelheim Pharma: Employment.

Background: Urgent surgery or life-threatening bleeding requires prompt reversal of the anticoagulant effects of the direct thrombin inhibitor dabigatran. As no specific reversal agent in situations of life-threatening bleeding with dabigatran is currently available, this study assessed the ability of a four-factor prothrombin complex concentrate (PCC) as mono-therapy as well as PCC in combination with tranexamic acid (TX) and fibrinogen in an experimental polytrauma pig model. Methods: After ethical approval dabigatran etexilate (30 mg/kg p.o. twice daily, n=24) was administered to male pigs for 3 days. On day 4, the pigs were anesthetized and given a 90 min infusion of active dabigatran to achieve consistent, supratherapeutic plasma levels. A standardized polytrauma including bilateral femur fractures and a blunt liver injury was induced. Animals underwent hemorrhagic shock and were resuscitated using Ringer’s solution (1 L). Twelve min after polytrauma, animals received either placebo (control group, n=6); TX, 20 mg/kg plus fibrinogen (80 mg/kg; TX+F group, n=6); PCC alone (PCC group, 50 U/kg, n=6); or TX (20 mg/kg) plus fibrinogen (80 mg/kg) plus PCC (50 U/kg; TX+F+PCC group, n=6) according to randomized group allocation. Coagulation was assessed by thromboelastometry, coagulation parameters and diluted TT. Blood loss (BL) was measured over the observation period of 240 min. Data were analyzed by ANOVA (± SD). Results: Dabigatran levels were 504 ± 171 ng/mL prior to injury and plasma levels remained significantly elevated in all animals throughout the observation period. The degree of injury was similar among animals with comparable BL of 803 ± 46 mL 12 min post injury. Anticoagulation with dabigatran without intervention resulted in a total BL of 3521 ± 600 mL, with 100% mortality and mean survival time of 101 ± 34 min (range: 67 - 148 min; p Conclusions: This study shows that bleeding in a lethal trauma model can be reduced with mono-therapy using a four-factor PCC. Prior correction of hypofibrinogenanemia with fibrinogen concentrate and tranexamic acid further improves PCC efficacy. Although the effects of PCCs on dabigatran reversal are not well understood, one possible explanation is that by elevating thrombin concentrations through PCC administration, (i.e. PCCs contain prothrombin) this may competitively bind to dabigatran, thereby reducing its anti-thrombin anticoagulant activity. This hypothesis is supported by the lack of effect on blood loss by mono-therapy with TX plus fibrinogen in this study. Disclosures Grottke: Boehringer Ingelheim: Consultancy, Research Funding; CSL Behring: Research Funding. Honickel:Boehringer Ingelheim: Travel support Other. Spronk:Boehringer Ingelheim: Research Funding. van Ryn:Boehringer Ingelheim Pharma: Employment. Rossaint:CSL Behring: Honoraria.

Introduction New oral anticoagulants are effective alternatives to warfarin; however, no specific reversal agents are currently available for life-threatening bleeding or emergency surgery. This study measured the effects of prothrombin complex concentrate (PCC), activated PCC (aPCC) and recombinant factor VIIa (rFVIIa) and compared this to reversal with the specific antibody fragment for dabigatran after high-dose dabigatran etexilate (DE) treatment in a porcine model of trauma, to assess reversal of anticoagulation. Methods Studies were performed in 5 male pigs after ethical approval. DE was given orally for 3 days (30 mg/kg bid) and, on the 4th day, dabigatran was infused at 0.77mg/kg/h for 30 min followed by 0.52 mg/kg/min for 60 min. The infusion was administered immediately before trauma (blunt liver insult). Blood samples were taken before dabigatran infusion and 60 min post-injury. Two doses of PCC (30 and 60 IU/kg), aPCC (30 and 60 IU/kg) and rVIIa (90 and 180 μg/kg) and two doses anti-Dabi Fab (30 and 60 mg/kg) were added to blood samples ex vivo. Coagulation was assessed by thromboelastometry (ExTEM assay) and PT and dabigatran levels with diluted TT. Data were analyzed by ANOVA (± SEM). Results Oral DE prolonged clot formation (CT: 389 ± 128 sec; CFT: 159 ± 39 sec) and PT (27 ± 9 sec), but coagulopathy was more severe after achieving high dabigatran plasma levels (943 ± 147 ng/ml) and inducing trauma (CT: 2378 ± 480 sec; CFT: 3374 ± 625 sec; PT: 167 ± 40 sec). At this time point, 30 IU/kg PCC and 30 IU/kg aPCC significantly accelerated clot formation (PCC: CT 416 ± 70 sec, CFT 250 ± 91 sec; aPCC: CT 525 ± 126 sec, CFT 309 ± 60 sec; p Conclusion The ex vivo addition of either PCC or aPCC is effective in reducing coagulopathy in a porcine model of high-dose dabigatran therapy and trauma. However the specific aDabi-Fab is the most promising agent to reverse the anticoagulant effects of dabigatran. The potential for reducing blood loss remains to be investigated. Disclosures: van Ryn: Boehringer Ingelheim Pharma GmbH & Co. KG: Employment. Spronk:Boehringer Ingelheim Pharma GmbH & Co. KG: Research Funding. Rossaint:Boehringer Ingelheim: Research Funding. Grottke:Boehringer Ingelheim: Research Funding; CSL Behring: Consultancy.

Introduction A specific antibody fragment (Fab) is currently in development to reverse the effects of dabigatran in cases of life-threatening bleeding or emergency surgery. In trauma, different volume expanders are used for resuscitation to compensate for blood loss and hemorrhagic shock. The influence of volume expanders on binding of dabigatran to the Fab, however, is not known. Therefore, in this study we investigated whether frequently used infusion solutions affect binding of dabigatran to the Fab antidote and the resulting effects on coagulation assays. Methods All studies were performed after ethical approval. Dabigatran etexilate was given to pigs for 3 days (30 mg/kg bid) and on day 4 a 90 min infusion of dabigatran (30 min:0.77mg/kg/h; 60 min:0.26 mg/kg/min) was performed in anesthetized pigs. Then ∼50% (100 mL/min) of total blood volume was removed and animals were randomized to (n=5/group): balanced Ringer’s solution, 6% HES 130/0.4, 6% HES 200/0.5, 4% gelatin, retransfusion of washed red blood cells (RBCs) or control (no haemodilution). Resuscitation consisted of, 1:1 to blood loss for crystalloids, 25 mL/kg for colloids, and 12 mL/kg for RBC. Antidote was then given (30 mg/kg iv) and serial blood samples were taken for up to 24 hrs to measure active dabigatran as diluted TT (Hemoclot) and total dabigatran (LC-MS/MS). Coagulation was assessed by means of aPTT, diluted PT (dPT, Neoplastin R, 1:100), i-Stat ACT celite (ACT), and ROTEM, ExTem and InTem reagents. Data expressed as mean ± SE and analyzed with ANOVA. Results Hemodilution resulted in a substantial reduction of hemostatic parameters (Hemoglobin, platelets and leukocytes) with all agents, except after RBC transfusion vs control. Mean plasma dabigatran levels measured at dTT were 640 ± 60 ng/mL after infusion and 625 ± 123 ng/mL after hemodilution without significant differences between groups. Administration of 30 mg/kg iv Fab resulted in a comparable reduction of active dabigatran in all groups, with a calculated half-life of 3.2 ± 0.1 hrs. The best correlation between dTT and other coagulation assays was obtained for the dPT (r2=0.6321), the ACT-celite (r2=0.6605) and clotting time from ExTEM and InTEM ROTEM (r2=0.5169 and 0.7303, respectively). The correlation between aPTT and dTT was moderate: r2=0.2321. Overall, coagulation was prolonged upon dabigatran treatment as indicated by the dPT (from 32.3±10.6 to 162.7±18.1 sec), aPTT (fom 44.6±6.2 to 67.9±10.3 sec), ACT (from 97±6 to 297±25 sec), CT ExTem (from 36.5±3.7 to 1205.4±598.3 sec) and CT InTem (from 122.0±11.9 to 989.3±218.9 sec). Fab treatment restored coagulation by on average 73% (between 58 and 95%, depending on the applied assay) at 5 minutes following administration. The dPT shortened to 43.8±24.1 sec, the aPTT to 30.6±5.5 sec, the ACT to 126±25 sec, the CT ExTem to 124.3±83.4 sec, and the CT InTem to 300.7±168.7 sec, with no significant differences in coagulation parameters between the various volume expanders. However, resuscitation with 4% gelatin and Fab treatment resulted in a significant lesser reduction of the CT InTem compared to the other volume expanders: 631.6±360.4 sec for 4% gelatin and 203.8±22.4, 251.7±157.2, and 280.8±235.3 sec for 6% HES 200/0.5, 6% HES 130/0.4, and Ringers, respectively. All coagulation parameters were comparable between the various volumes expanders from 60 minutes onward. Twenty four hours after Fab administration all coagulation parameters were almost back to baseline values with 26.9±62.8 ng/mL dabigatran, 50.9±12.6 sec for the dPT, 48.6±13.5 sec for the aPTT, 121±6 sec for ACT, 50.0±3.4 sec for CT ExTem, and 189.7±24.5 sec for CT InTem. Conclusions Clinically used infusion solutions for volume resuscitation do not interfere with binding of dabigatran to its specific antidote. Inhibition of dabigatran with its specific Fab antidote at this particular dose, restored coagulation by almost 70% This effect was independent of the volume applied for resuscitation. Disclosures: Spronk: Boehringer Ingelheim Pharma GmbH & Co. KG: Research Funding. van Ryn:Boehringer Ingelheim Pharma GmbH & Co. KG: Employment. Grottke:Boehringer Ingelheim Pharma GmbH & Co. KG: Research Funding.