USOO9034822B2 (12) United States Patent (10) Patent No.: US 9,034,822 B2 Van Ryn et al. (45) Date of Patent: *May 19, 2015 (54) METHODS OF USING ANTIBODIES DURING (56) References Cited ANTCOAGULANT THERAPY OF DABGATRAN AND/OR RELATED U.S. PATENT DOCUMENTS COMPOUNDS 6,440,417 B1 8/2002 Thibaudeau et al. 6,469,039 B1 10/2002 Hauel et al. (71) Applicants: Joanne Van Ryn, Warthausen (DE); 8,486,398 B2* 7/2013 Van Ryn et al. ........... 424,133.1 2004/OO97547 A1 5, 2004 Taveras et al. John Edward Park, Warthausen (DE): 2011/0206656 A1 8/2011 Van Ryn et al. Norbert Hauel, Schemmerhofen (DE): 2012fOO27780 A1 2/2012 Van Rynet al. Ulrich Kunz, Biberach an der Riss (DE); Tobias Litzenburger, Mittelbiberach FOREIGN PATENT DOCUMENTS (DE); Keith Canada, Southbury, CT CA 2 277 949 8, 1998 (US); Sanjaya Singh, Sandy Hook, CT CA 2277949 A1 * 8, 1998 (US); Alisa Waterman, Weston, CT WO WO-9837O75 8, 1998 (US) WO WO-2011 O23.653 3, 2011 WO WO-2011 089183 T 2011 (72) Inventors: Joanne Van Ryn, Warthausen (DE); John Edward Park, Warthausen (DE): OTHER PUBLICATIONS Norbert Hauel, Schemmerhofen (DE): Schlaeppi et al., Eur J Biochem. Mar. 10, 1990:188(2):463-70.* Ulrich Kunz, Biberach an der Riss (DE); Herion et al., Blood. May 1985:65(5):1201-7.* Tobias Litzenburger, Mittelbiberach Colburn, W. A., “Specific antibodies and fab fragments to alter the (DE); Keith Canada, Southbury, CT pharmacokinetics and reverse the pharmacologic/toxicologic effects (US); Sanjaya Singh, Sandy Hook, CT of drugs.” Drug Metabolism Reviews, 1980, vol. 11, No. 2, pp. (US); Alisa Waterman, Weston, CT 223-262. Prescrire Int. Jun. 2009; 18(101):97-9.* (US) Eisert, W. G. et al., Arteriosclerosis, Thrombosis, and Vascular Biol (73) Assignee: Boehringer Igelheim International ogy, 2010, vol. 30, pp. 1885-1889. International Search Report for PCT/EP2011/073025 dated Mar. 14, GmbH, Ingelheim am Rhein (DE) 2012. International Search Report for PCT/EP2012/055397 dated May 30, (*) Notice: Subject to any disclaimer, the term of this 2012. patent is extended or adjusted under 35 Van Ryn, J. et al., “Dabigatran anticoagulantactivity is neutralized by U.S.C. 154(b) by 0 days. an antibody selective to dabigatran in in vitro and in vivo models.” This patent is Subject to a terminal dis JACC, Apr. 5, 2011, vol. 57, No. 14. claimer. Written Opinion of the International Search Authority for PCT/ EP2011 073025 dated Mar. 14, 2012. (21) Appl. No.: 13/916,994 Written Opinion of the International Searching Authority for PCT/ EP2012/055397 dated May 30, 2012. (22) Filed: Jun. 13, 2013 Van Ryn, J. et al., “Dabigatran etexilate—novel, reversible, oral direct thrombin inhibitor: interpretation of coagulation assays and (65) Prior Publication Data reversal of anticoagulant activity.” Thrombosis and Haemostasis, 2011, vol. 105, No. 3, pp. 570. US 2013/0289248A1 Oct. 31, 2013 Hardin, J. S. et al., “Pharmacodynamics of a monoclonal antiphencyclidine fab with broad selectivity for phencyclidine-like drugs.” The Journal of Pharmacology and Experimental Therapeu Related U.S. Application Data tics, 1998, vol. 285, No. 3, pp. 1113-1122. Hursting, M. J. et al., “Drug-specific fab therapy in drug overdose.” (63) Continuation of application No. 13/010,403, filed on Arch Pathol Lab Med., 1987, vol. 111, pp. 693-697. Jan. 20, 2011, now Pat. No. 8,486,398. Lapostolle, F. et al., “Assessment of digoxin antibody use in patients with elevated serum digoxin following chronic or acute exposure.” (60) Provisional application No. 61/383,914, filed on Sep. Intensive Care Med, 2008, vol. 34, pp. 1448-1453. 17, 2010. Schulman, S. et al., “Anticoagulants and their reversal.” Transfusion Medicine Reviews, Jan. 2007, vol. 21, No. 1, pp. 37-48. (30) Foreign Application Priority Data (Continued) Jan. 20, 2010 (EP) ..................................... 1O151239 Primary Examiner — Michael Szperka (51) Int. Cl. (74) Attorney, Agent, or Firm — Michael P. Morris; Wendy A 6LX39/395 (2006.01) A. Petka C07K I6/44 (2006.01) (52) U.S. Cl. (57) ABSTRACT CPC ............. C07K 16/44 (2013.01); C07K 2317/76 The present invention relates to antibody molecules against (2013.01) anticoagulants, in particular dabigatran, and their use as anti (58) Field of Classification Search dotes of Such anticoagulants. None See application file for complete search history. 9 Claims, 14 Drawing Sheets US 9,034,822 B2 Page 2 (56) References Cited reversal of anticoagulantactivity.” Thromb Haemost, 2010, vol. 103, pp. 1116-1127. Zikria, J. C. et al., “Oral anticoagulation with factor Xa and thrombin OTHER PUBLICATIONS inhibitors: on the threshold of change.” Current Opinion in Hematol ogy, 2009, vol. 16, No. 5, pp. 347-356. Van Ryn, J. et al., “Dabigatran etexilate—a novel, reversible, oral direct thrombin inhibitor: Interpretation of coagulation assays and * cited by examiner U.S. Patent May 19, 2015 Sheet 1 of 14 US 9,034,822 B2 11 O5 OO 5 O 400 500 Dabigatran (nM) FIG. 1 U.S. Patent May 19, 2015 Sheet 2 of 14 US 9,034,822 B2 g --------------------------Dabigatran 5 50 2OOnM : 40 CD E 30 2 20 5 O 10-- - - - - - - - - - - - - - - - - - - - - - - ---Control O 100 200 300 400 500 600 Antibody Concentration (pig) FIG. 2 U.S. Patent May 19, 2015 Sheet 3 of 14 US 9,034,822 B2 g ------------------------Dabigatran is 60 2OOnM c) 0. 40 O 20 O CS ------------- - - - - - O- - - - - Control O 1 OO 200 300 400 500 600 Antibody "D" concentration (pg) onspecific polyclonal Ab (1671g) + 200 nM dabigatran FIG. 3 U.S. Patent May 19, 2015 Sheet 4 of 14 US 9,034,822 B2 a "l-o------------------------- Dabigatran 2 5 OOnM 8 90 O) 0. O)E 60 an 2 30 6 ------------------------- an Control O O plasma H O 500 1OOO Antibody "D" concentration (pig) FIG. 4 U.S. Patent May 19, 2015 Sheet 6 of 14 US 9,034,822 B2 5-5ETTE?G U.S. Patent May 19, 2015 Sheet 7 of 14 US 9,034,822 B2 12O.OO 1OOOO S. a 80.00 e t 60.00 - W t S. 40.00 W 2OOO W o &six & OOO - O.1OO 1.OOO 1OOOO Antibody (ig/ml) -- Whole Blood 2U/ml -- Whole Blood 1.5U/ml A Plasma 2U/ml -o- Plasma 1.5U/m FIG. 7 U.S. Patent May 19, 2015 Sheet 8 of 14 US 9,034,822 B2 Š 9. E 50.00 o es an s t an U.S. Patent May 19, 2015 Sheet 9 of 14 US 9,034,822 B2 1OOOO 9 OOO 8OOO S 7000 - pre-r .9 6000 - X --Antibody + Dabigatran E 5OOO ... I --Antibody.Acylglucuronide Dablatan 30.00 S 1.O.OO OOO O.1O 1.OO 1.O.OO Antibody (g/ml) FIG. 9 U.S. Patent May 19, 2015 Sheet 10 of 14 US 9,034,822 B2 100 90 IC50 s 80 -o- Fab 18/15 O.1247 E SR 70 s 60 ICso 2.6 nM SYSS 50 CD E 40 2 30 a 20 O 10 O O.O 0.5 1.0 1.5 2.0 Antibody concentration (g/ml) FIG. 10 U.S. Patent May 19, 2015 Sheet 11 of 14 US 9,034,822 B2 1. 5 t O O - - - . O - Hy -20 5 O 5 O 25 30 Time (min) -- CO to -H 0.3 + 0.1 uM/kg.hr -- Fab 0.3 Mkg Dabigatran bolus (0.3 uMkg) + infusion (0.1 uMkg/hr) Thrombin time: 3.0 Uml thrombin used in the assay Data expressed as meant SE, n=4. FIG. 11 U.S. Patent May 19, 2015 Sheet 12 of 14 US 9,034,822 B2 35 30 E O 15 Hi----- - IIII -20 5 O 5 20 25 30 Time (min) -- Control –H 0.3 + O.1 uM/kg/hr -- Fab O.3 uM.kg Dabigatran. Bolus (0.3 uMkg) + infusion (0.1 uMkg/hr) Data expressed as mean E SE, n=4. FIG. 12 U.S. Patent May 19, 2015 Sheet 13 of 14 US 9,034,822 B2 125 1OO 7 5 50 2 5 O - III -O O. 5 O 5 O 25 30 Time (min) -- Control -H 0.3 + 0.1 uM/kg/hr -W Fab O. 15 M/kg -- Fab 0.3 uM/kg Dabigatran bolus (0.3 uM/kg) + infusion (0.1 uM/kg/hr) Thrombin time: 3.0 U.ml thrombin used in the assay Data expressed as mean E SE, n=4 FIG. 13 U.S. Patent May 19, 2015 Sheet 14 of 14 US 9,034,822 B2 Time (min) -- Control -H 0.3 + 0.1 uM/kg/hr -- Fab O.3 uw.kg - W. Fab O. 15uM/kg Dabigatran bolus (0.3 uM/kg) + infusion (0.1 uM/kg/hr) Data expressed as mean E SE, n=4 FIG. 14 US 9,034,822 B2 1. 2 METHODS OF USING ANTIBODES DURING There is a need to provide improved antidotes for antico ANTICOAGULANT THERAPY OF agulant therapy, and in particular to provide antidotes for DABGATRAN AND/OR RELATED direct thrombin inhibitors like dabigatran for which no spe COMPOUNDS cific antidotes have been disclosed so far. BACKGROUND OF THE INVENTION BRIEF SUMMARY OF THE INVENTION 1. Technical Field In one aspect, the present invention relates to an antibody The present invention pertains to the field of medicine, in molecule capable of neutralizing the activity of an anticoagul particular to the field of anticoagulant therapy. 10 lant. 2. Background Information In a further aspect, the antibody molecule has binding Anticoagulants are substances that prevent coagulation; specificity for the anticoagulant. that is, they stop blood from clotting. Anticoagulants are widely used in human therapy as a medication for thrombotic In a further aspect, the anticoagulant is a direct thrombin disorders, for example primary and secondary prevention of 15 inhibitor, a Factor Xa inhibitor, or a vitamin Kantagonist.