Lyophilized formulations for factor xa antidote

Abstract

The present disclosure relates to solutions and methods of preparing lyophilized formulations of factor Xa (fXa) antidotes. A suitable aqueous formulation suitable for lyophilization can include a fXa antidote, a solubilizing agent, a stabilizer, and a crystalline component, wherein the formulation does not collapse during lyophilization.

1 . An aqueous formulation, comprising from 10 mM to 55 mM arginine, from 1% to 3% sucrose (w/v), from 2% to 8% mannitol (w/v) and at least 5 mg/mL of a two-chain polypeptide comprising a first chain comprising the amino acid sequence of SEQ ID NO. 4, a second chain comprising the amino acid sequence of SEQ ID NO. 5, and a disulfide bond between a first Cysteine residue at position 98 (Cys98) of SEQ ID NO. 4 and a second Cysteine residue at position 108 (Cys108) of SEQ ID NO. 5, wherein the formulation has a pH from 7.5 to 8. 
     
     
         2 . The formulation of  claim 1 , comprising from 40 mM to 50 mM arginine, from 1.5% to 2.5% sucrose (w/v), from 4.5% to 5.5% mannitol (w/v) and at least 10 mg/mL of the polypeptide. 
     
     
         3 . The formulation of  claim 2 , comprising at least 18 mg/mL of the polypeptide. 
     
     
         4 . The formulation of  claim 2 , wherein the polypeptide comprises an amino acid residue that is modified to be different from natural amino acids. 
     
     
         5 . The formulation of  claim 4 , wherein residue Asp29 of the first chain is modified to (3R)-3-hydroxyAsp at Asp29. 
     
     
         6 . The formulation of  claim 1 , wherein the polypeptide comprises at least an intra-chain disulfide bond for each of the first and second chains. 
     
     
         7 . The formulation of  claim 1 , comprising at least about 10 mg/mL of the two-chain polypeptide. 
     
     
         8 . The formulation of  claim 1 , comprising at least about 20 mg/mL of the two-chain polypeptide. 
     
     
         9 . An aqueous formulation, comprising from 10 mM to 55 mM arginine, from 1% to 3% sucrose (w/v), from 2% to 8% mannitol (w/v) and at least 5 mg/mL of a two-chain polypeptide comprising a first chain comprising the amino acid sequence of SEQ ID NO. 4 or having at least 90% sequence identity to SEQ ID NO. 4, a second chain of the amino acid sequence of SEQ ID NO. 5 or having at least 90% sequence identity to SEQ ID NO. 5, and a disulfide bond between a first Cysteine residue at position 98 (Cys98) of SEQ ID NO. 4 and a second Cysteine residue at position 108 (Cys108) of SEQ ID NO. 5, wherein the formulation has a pH from 7.5 to 8. 
     
     
         10 . The formulation of  claim 9 , wherein the two-chain polypeptide has modifications to the Gla domain and the active site as compared to the wild-type fXa protein, is able to bind to a fXa inhibitor but does not assemble into a prothrombinase complex. 
     
     
         11 . A method of preparing a lyophilized formulation, comprising lyophilizing the aqueous formulation of  claim 1 . 
     
     
         12 . A lyophilized composition obtainable by lyophilizing the aqueous formulation of  claim 1 . 
     
     
         13 . A lyophilized composition comprising at least 10% (w/w) of a two-chain polypeptide comprising a first chain of the amino acid sequence of SEQ ID NO. 4, a second chain of the amino acid sequence of SEQ ID NO. 5, and a disulfide bond between a first Cysteine residue at position 98 (Cys98) of SEQ ID NO. 4 and a second Cysteine residue at position 108 (Cys108) of SEQ ID NO. 5, and L-arginine HCl:sucrose:mannitol in a weight ratio of the range (0.5-1.4):(1-3):(2-8). 
     
     
         14 . The lyophilized composition of  claim 13 , comprising at least 18% (w/w) of the two-chain polypeptide. 
     
     
         15 . The lyophilized composition of  claim 13 , wherein the weight ratio of L-arginine HCl:sucrose:mannitol is in the range of (0.9-1):(1.5-2.5):(4.5-5.5). 
     
     
         16 . The lyophilized composition of  claim 13 , wherein the weight ratio of L-arginine HCl:sucrose:mannitol is about 0.95:2:5. 
     
     
         17 . A method of reducing bleeding in a subject undergoing anticoagulant therapy with a factor Xa inhibitor comprising administering to the subject a solution prepared by dissolving the lyophilized composition of  claim 13  in an aqueous solvent. 
     
     
         18 . The method of  claim 17 , wherein the factor Xa inhibitor is apixaban, rivaroxaban or betrixaban. 
     
     
         19 . An aqueous formulation, comprising a polypeptide comprising the amino acid sequence of SEQ ID NO. 3 or an amino acid sequence having at least 95% sequence identity to SEQ ID NO. 3, a solubilizing agent, a stabilizer, and a crystalline component, wherein the formulation does not collapse during lyophilization. 
     
     
         20 . The formulation of  claim 19 , wherein the crystalline component is mannitol present in a concentration from 2% to 8% (w/v), the solubilizing agent is arginine and the stabilizer is sucrose. 
     
     
         21 . The formulation of  claim 19 , further comprising a surfactant and a buffer.