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WO 2013/185113 Al 12 December 2013 (12.12.2013) P O P C T

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WO 2013/185113 Al 12 December 2013 (12.12.2013) P O P C T (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization I International Bureau (10) International Publication Number (43) International Publication Date WO 2013/185113 Al 12 December 2013 (12.12.2013) P O P C T (51) International Patent Classification: (81) Designated States (unless otherwise indicated, for every A61K 38/02 (2006.01) A61P 7/02 (2006.01) kind of national protection available): AE, AG, AL, AM, A61K 38/36 (2006.01) AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY, BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DK, DM, (21) Number: International Application DO, DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, PCT/US20 13/044841 HN, HR, HU, ID, IL, IN, IS, JP, KE, KG, KN, KP, KR, (22) International Filing Date: KZ, LA, LC, LK, LR, LS, LT, LU, LY, MA, MD, ME, 7 June 2013 (07.06.2013) MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, OM, PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SC, (25) Filing Language: English SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, (26) Publication Language: English TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW. (30) Priority Data: (84) Designated States (unless otherwise indicated, for every 61/657,688 8 June 2012 (08.06.2012) kind of regional protection available): ARIPO (BW, GH, 61/800,626 15 March 2013 (15.03.2013) GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, SZ, TZ, UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, RU, TJ, (71) Applicant: BIOGEN IDEC MA INC. [US/US]; 14 Cam TM), European (AL, AT, BE, BG, CH, CY, CZ, DE, DK, bridge Center, Cambridge, Massachusetts 02142 (US). EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU, LV, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK, SM, (72) Inventors: HONG, Vu Phong; 27 Wheeler Street #217, TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ, GW, Cambridge, Massachusetts 02138 (US). MEZO, Adam R.; KM, ML, MR, NE, SN, TD, TG). 12449 Meeting House Road, Carmel, Indiana 46032 (US). SALAS, Joe; 9 D'Angelo Road, Wayland, Massachusetts Published: 01778 (US). PETERS, Robert; 51 Newfield Street, West — with international search report (Art. 21(3)) Roxbury, Massachusetts 02132 (US). — before the expiration of the time limit for amending the (74) Agents: STEFFE, Eric K. et al; Sterne, Kessler, Gold claims and to be republished in the event of receipt of stein & Fox P.L.L.C., 1100 New York Avenue, NW, amendments (Rule 48.2(h)) Washington, District of Columbia 20005 (US). — with sequence listing part of description (Rule 5.2(a)) (54) Title: PROCOAGULANT COMPOUNDS (57) Abstract: The present disclosure provides protease-activatable procoagulant compounds comprising a procoagulant poly peptide, e.g., a procoagulant peptide and/or clotting factor, and a linker comprising a protease-cleavable substrate (e.g., a synthetic thrombin substrate) and a self-immolative spacer (e.g., p-amino benzyl carbamate). Upon cleavage of the protease-cleavable sub - strate by a protease (e.g., thrombin), the self-immolative spacer cleaves itself from the procoagulant polypeptide such that the poly- peptide is in an underivatized and active form. Also provided are pharmaceutical compositions, methods for treating bleeding dis - orders using the disclosed compounds, methods of enhancing in vivo efficacy of procoagulant polypeptides, methods of increasing the efficacy of proteolytic cleavage of compounds comprising procoagulant polypeptides, methods of activating procoagulant poly- peptides, and methods of releasing a procoagulant polypeptide from a heterologous moiety such as PEG. PROCOAGULANT COMPOUNDS BACKGROUND Field of the Disclosure [0001] The present invention relates to procoagulant compounds useful for the treatment of bleeding diseases or disorders. Background [0002] The blood coagulation pathway, in part, involves the formation of an enzymatic complex of Factor Villa (FVIIIa) and Factor IXa (FIXa) (Xase complex) on the surface of platelets. FIXa is a serine protease with relatively weak catalytic activity without its cofactor FVIIIa. The Xase complex cleaves Factor X (FX) into Factor Xa (FXa), which in turn interacts with Factor Va (FVa) to cleave prothrombin and generate thrombin. Hemophilia A is a bleeding disorder caused by mutations and/or deletions in the factor VIII (FVIII) gene resulting in a deficiency of FVIII activity (Peyvandi et al. 2006). Hemophilia B (also known as Christmas disease) is one of the most common inherited bleeding disorders in the world. It results in decreased in vivo and in vitro blood clotting activity and requires extensive medical monitoring throughout the life of the affected individual. 03] Treatment of hemophilia is by replacement therapy targeting restoration of clotting activity. There are plasma-derived and recombinant clotting factor products available to treat bleeding episodes on-demand or to prevent bleeding episodes from occurring by treating prophylactically. Based on the half-life of these products, treatment regimens require frequent intravenous administration. Such frequent administration is painful and inconvenient. Strategies to extend the half-life of clotting factors include pegylation (Rostin J, et al, Bioconj. Chem. 2000;1 1:387-96), glycopegylation (Stennicke HR, et al, Thromb. Haemost. 2008;100:920-8), formulation with pegylated liposomes (Spira J, et al, Blood 2006; 08:3668-3673, Pan J, et al, Blood 2009; 114:2802-281 1) and conjugation with albumin (Schulte S., Thromb. Res. 2008;122 Suppl 4-.S14-9). However, modification of coagulation factors and procoagulant peptides with half-life extending moieties (e.g., PEG) and other similar strategies to extend their half-lives can lead to compromised activity. In order to rescue their activity, a cleavable linker can be inserted between the protein or peptide of interest and its modifier. The chosen cleavable linker must be cleaved efficiently and rapidly by a protease, for example, a protease involved in the coagulation cascade. Thrombin being the activator of many clotting factors is the most popular choice. However, all known substrate sequences composed of natural amino acids (e.g., LVPR, ALRPR (SEQ ID NO: 7), etc.) are not optimal substrates. Furthermore, covalent binding of the cleavable linker to a coagulation factors or procoagulant peptide can result in steric hindrances (e.g., due to the presence of amino acids such as such as proline, isoleucine or arginine C-terminal to the cleavage site) that can prevent an efficient enzymatic cleavage reaction. BRIEF SUMMARY The present disclosure provides procoagulant compounds comprising a protease- cleavable substrate (e.g., a synthetic thrombin substrate) and a self-immolative spacer (e.g., PABC) linked to a procoagulant polypeptide, e.g., a clotting factor or a procoagulant peptide. Accordingly, in some embodiments, the present disclosure provides A procoagulant compound having a formula: (Het2)-(Pep2)-(Hetl)-(L)-Zy-Bx-Pepl (Formula I) wherein, Hetl is a first heterologous molecule, which is either absent or present; Het2 is a second heterologous molecule, which is either absent or present; L is a linker, which is either absent or present; Zy is a protease-cleavable substrate; Bx is a self-immolative spacer; Pepl is a polypeptide; and, Pep2 is a polypeptide, which is either absent or present; wherein, Pepl or Pep2 comprises a clotting factor or a fragment thereof, or a synthetic procoagulant peptide. [0005] In some embodiments, the self-immolative spacer in the procoagulant compound of the invention undergoes 1,4 elimination after the enzymatic cleavage of the protease- cleavable substrate. In some embodiments, the self-immolative spacer in the procoagulant compound of the invention undergoes 1,6 elimination after the enzymatic cleavage of the protease-cleavable substrate. In some embodiments, the self-immolative spacer is a p- amino benzyl carbamate (PABC), a p-amino benzyl ether (PABE), or a p-amino benzyl carbonate. In certain embodiments, the self-immolative spacer comprises an aromatic group. In some embodiments, the aromatic group is selected from the group consisting of benzyl, cinnamyl, naphthyl, and biphenyl. In some embodiments, the aromatic group is heterocyclic. In other embodiments, the aromatic group comprises at least one substituent. In some embodiments, at least one substituent is selected from F, CI, I, Br, OH, methyl, 3+ methoxy, N0 2, NH2, N0 , NHCOCH 3, N(CH3)2, NHCOCF3, alkyl, haloalkyl, Ci-C alkylhalide, carboxylate, sulfate, sulfamate, sulfonate, or any combinations thereof In other embodiments, at least one C in the aromatic group is substituted with N, O, or C-Ri, wherein R i is independently selected from H, F, CI, I, Br, OH, methyl, methoxy, N0 2, + NH2, N0 , NHCOCH 3, N(CH3) , NHCOCF3, alkyl, haloalkyl, Ci-C alkylhalide, carboxylate, sulfate, sulfamate, and sulfonate. [0006] In some embodiments, the protease-cleavable substrate in the procoagulant compound of the invention comprises a coagulation cascade protease substrate. In some embodiments, the coagulation cascade protease is selected from thrombin, thromboplastin, Factor Va, Factor Vila, Factor Villa, Factor IXa, Factor Xa, Factor XIa, Factor Xlla, or any combinations thereof. In other embodiments, the coagulation cascade protease substrate is a thrombin substrate. In some embodiments, the thrombin substrate is a synthetic thrombin substrate. In other embodiments, the synthetic thrombin substrate comprises the sequence of D-Phe-Pip-Arg. In some embodiments, the thrombin substrate is selected from D-Phe-Pro-Arg, D-Ala-Leu-Val-Pro-Arg (SEQ ID NO: 17), Ala-Leu- Val-Pro-Arg (SEQ
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