Method for purification of complement factor h

Abstract

A method for purification of complement Factor H from a complement Factor H containing source such as blood or blood plasma, in particular a caprylate precipitate of a Factor H containing source, which is e.g. obtained by addition of caprylate ions to fractions of blood or plasma, comprising the steps of: a) providing a Factor H containing source, in particular reconstitution of caprylate precipitate to provide a complement Factor H containing solution; b) performing a cation exchange chromatography in particular as first chromatographic step; c) performing an anion exchange chromatography; d) performing a hydroxyl apatite chromatography; e) followed by ultra/diafiltration to obtain a complement Factor H concentrate.

1 . (canceled) 
     
     
         2 . The complement factor H of  claim 5 , wherein the purification method further comprises performing a heparin affinity chromatography step. 
     
     
         3 . The complement factor H of  claim 5  wherein the purification method comprises at least one of the following methods for pathogen removal and/or inactivation:
 a. solvent/detergent treatment; 
 b. pasteurization, 
 c. vapor heat treatment; 
 d. dry heat treatment; or 
 e. nanofiltration. 
 
     
     
         4 . The complement factor H of  claim 5  wherein the complement Factor H concentrate is lyophilized. 
     
     
         5 . A complement Factor H obtained from a complement Factor H containing source, wherein the source is a caprylate precipitate obtained by the addition of caprylate ions to fractions of blood or blood plasma, by a purification method comprising the steps of:
 a. reconstituting the caprylate precipitate to provide a complement Factor H containing solution,   b. performing cation exchange chromatography;   c. performing anion exchange chromatography;   d. performing hydroxyl apatite chromatography;   e. followed by ultra/diafiltration to obtain a complement Factor H concentrate.   
     
     
         6 . The complement Factor H of  claim 5  obtained by a method comprising
 a. reconstitution of caprylate precipitate to provide a complement Factor H containing solution; 
 b. performing virus inactivation by solvent/detergent treatment (S/D treatment). 
 c. performing a cation exchange chromatography chromatographic step under the following conditions: 
 d. binding of complement Factor H to a strong cation exchange resin of the sulphopropyl type, washing with a buffer comprising 20 mM tri-sodium citrate adjusted to pH 6.0 and elution of complement Factor H with an elution buffer comprising 20 mM tri-sodium citrate and 0.2 M NaCl adjusted to pH 6.0; 
 e. performing an anion exchange chromatography step under the following conditions:
 i. applying of the complement Factor H containing solution (conductivity 0.1-0.5 mS/cm) to a strong anion exchange resin of the quaternary ammonium type, washing with a buffer comprising 20 mM Tris adjusted to pH 8.6 and elution of complement Factor H with an elution buffer comprising 20 mM Tris and 0.2M NaCl adjusted to pH 8.6; 
 
 f. performing a ceramic hydroxyl apatite chromatography by loading the fraction of step d) on the ceramic hydroxyl apatite, optionally after an exchange of the buffer of step d) has been performed, and elution with a linear gradient of sodium chloride up to 1 M NaCl and collecting fractions eluting at a conductivity of the buffer in the range of 70-100 mS/cm; 
 g. optionally performing a heparin affinity chromatography step under the following conditions: applying of the complement Factor H containing solution to a resin with heparin immobilized on the surface, washing with a buffer comprising 20 mM tri-sodium citrate adjusted to pH 6.0 and elution of complement Factor H with an elution buffer comprising 20 mM tri-sodium citrate and 0.2M NaCl adjusted to pH 6.0; 
 h. followed by ultra/diafiltration to obtain a complement Factor H concentrate. 
 
     
     
         7 . The complement Factor H according to  claim 5  wherein the purification method comprises a virus reducing step by nanofiltration. 
     
     
         8 . The complement Factor H according to  claim 5  characterized by being a liquid or lyophilized preparation. 
     
     
         9 . A method for treating a disease related to complement Factor H deficiency or abnormal activity, comprising administering the complement Factor H of  claim 5  to a subject in need thereof. 
     
     
         10 . The method according to  claim 9  wherein the disease is selected from Membranoproliferative Glomerulonephritis, Dense Deposit Disease, Hemolytic Uremic Syndrome, Atypical Hemolytic Uremic Syndrome or Age-related Macular Degeneration. 
     
     
         11 . A method for treating Ischemia Reperfusion Injury, Chronic Nephropathy or Autoimmune Encephalomyelitis, comprising administering the complement Factor H of  claim 5  to a subject in need thereof. 
     
     
         12 . The complement Factor H of  claim 5 , wherein the first chromatographic step in the purification method is cation exchange chromatography.