Stable galenic freeze-dried pharmaceutical preparation of recombinant carbohydrate-binding polypeptides

Abstract

The invention relates to a method for the production of a medicament containing a polypeptide comprising at least one recombinant carbohydrate-binding polypeptide, or a functional fragment or derivative of said carbohydrate-binding polypeptide in a form stable for storage. The polypeptide mentioned comprises polypeptides or functional derivatives thereof, which are fused with cytotoxically effective peptides to give fusion proteins, or which are linked to another polypeptide having a cytotoxic activity. Moreover, the invention describes further formulating of the disclosed medicaments to medicaments with different pharmaceutical forms.

1 . A method for the production of a medicament containing a polypeptide comprising at least one recombinant carbohydrate-binding polypeptide that is the B-chain of a ribosome-inactivating protein or a functional fragment or derivative of said carbohydrate-binding polypeptide, wherein
 (a) the polypeptide or a functional fragment or derivative of this polypeptide which is fused to a cytotoxically effective peptide to form a fusion protein;   (b) the polypeptide or a functional fragment or derivative of this polypeptide which is linked to another polypeptide which has an enzymatic rRNA-N-glycosidase activity;   (c) the polypeptide or a functional fragment or derivative of this polypeptide which is linked to another polypeptide in which an enzymatic rRNA-N-glycosidase activity has been replaced by another cytotoxic activity; or   (d) the polypeptide or a functional fragment or derivative of this polypeptide, which is linked to a fusion protein, comprising a polypeptide with an enzymatic rRNA-N-glycosidase activity and/or another cytotoxic activity;   in a form stable for storage for at least one year, moreover optionally containing a pharmaceutically acceptable carrier,   comprising the step of cooling, freezing, spray drying or lyophilising while retaining the pharmacological properties of the polypeptide in the solution, wherein the solution is characterised in that the pH value of the solution is higher than pH 6.0 and a buffer system contained in the solvent guarantees that this pH-value is maintained.   
   
   
       2 . (canceled) 
   
   
       3 . The method of  claim 1 , wherein the further polypeptide which is linked to the recombinant carbohydrate-binding polypeptide is the A-chain of a ribosome-inactivating protein. 
   
   
       4 . The method of  claim 1 , wherein the ribosome-inactivating protein is a ribosome-inactivating protein of the type II. 
   
   
       5 . The method of  claim 4 , wherein the ribosome inactivating protein is a type II rViscumin. 
   
   
       6 . The method of  claim 1 , wherein the pH-value of the solution is between 6.0 and 9.0. 
   
   
       7 . The method of  claim 1 , wherein the pH-value of the solution is between 7.5 and 8.5. 
   
   
       8 . The method of  claim 1 , wherein the salt(s) of the buffer system is/are used in a end concentration ranging from 5 mM to 200 mM. 
   
   
       9 . The method of  claim 1 , wherein the salt(s) of the buffer system is/are used in an end concentration ranging from 100 mM to 200 mM. 
   
   
       10 . The method of  claim 1 , wherein the salt(s) of the buffer system comprise one or more salts selected from the group consisting of TRIS/HCl, TRICIN/HCl, HEPES/HCl, ammonium carbonate buffer, TRIS/glutamic acid and TRIS/aspartic acid. 
   
   
       11 . The method of  claim 1 , wherein the solution contains one or more surfactants in order to stabilise the pharmacological properties of the polypeptide. 
   
   
       12 . The method of  claim 11 , wherein the surfactants are non-ionic tensides and are used in a final concentration ranging from 0.01 to 5.0%. 
   
   
       13 . The method of  claim 12 , wherein the non-ionic tensides are selected from the group comprising: fatty alcohols, partial glycerides, polysorbates, polyoxyethylene fatty acid ethers and polyoxyethylene fatty acid esters, poloxameres (polyoxypropylene-polyoxyethylene-block polymers), saccharide fatty acid esters, polyoxyglycerol fatty acid esters and phosphatides. 
   
   
       14 . The method of  claim 13 , wherein the polysorbates are selected from the group consisting of Polysorbate 80, Polysorbate 20 and polyoxyethylene sorbitol ether. 
   
   
       15 . The method of  claim 13 , wherein the polyoxyethylene fatty acid ethers and polyoxyethylene fatty acid esters are macrogol ethers or macrogol esters. 
   
   
       16 . The method of  claim 13 , wherein the poloxamere is Pluronic F68, poloxamer 166 or poloxamer 188. 
   
   
       17 . The method of  claim 13 , wherein the phosphatides are lecithins. 
   
   
       18 . The method of  claim 11 , wherein the surfactants are amphoteric tensides and are used in a final concentration ranging from 0.01 to 5.0%. 
   
   
       19 . The method of  claim 1 , wherein one or more lyoprotectors in a final concentration ranging from 0.1 to 20% and/or cryoprotectors in a final concentration ranging from 0.01 to 1.0% are added to the solution for lyophilisation. 
   
   
       20 . The method of  claim 19 , wherein the lyoprotectors are added in a final concentration ranging from 4.0 to 10% and/or the cryoprotectors are added in a final concentration ranging from 0.05 to 0.1%. 
   
   
       21 . The method of  claim 19 , wherein the lyoprotectors are selected from the group consisting of a) low molecular saccharides; b) hexites; c) oligomeric and polymeric saccharides; d) anorganic gelling agents; and e) synthetic polymers. 
   
   
       22 . The method of  claim 19 , wherein the lyoprotector or the lyoprotectors is dextrane or are dextranes. 
   
   
       23 . The method of  claim 19 , wherein ionic substances are used as cryoprotectors. 
   
   
       24 . The method of  claim 23 , wherein the ionic substances are selected from the group consisting of sodium chloride, sodium sulphate, potassium chloride and potassium sulphate. 
   
   
       25 . The method of  claim 19 , wherein the lyoprotectors and cryoprotectors form amorphous structures during lyophilisation. 
   
   
       26 . The method of  claim 19 , wherein the solution contains one or more amino acids which are used in a final concentration of from 0.01 to 50 mg/ml. 
   
   
       27 . The method of  claim 26 , wherein the amino acids are selected from the group comprising acidic amino acids such as glutamic acid and aspartic acid, the basic amino acid arginine and the neutral amino acid valine. 
   
   
       28 . The method of  claim 1 , wherein the polypeptide comprising at least one recombinant carbohydrate-binding polypeptide or functional fragment or derivative of this polypeptide is used in a final concentration of from 10 ng/ml to 10 mg/ml. 
   
   
       29 . The method of  claim 28 , wherein the polypeptide is used in a final concentration of from 100 ng/ml to 1 mg/ml. 
   
   
       30 . The method of  claim 1 , moreover comprising the further formulation or reconstitution of the medicament as aqueous or non-aqueous solution. 
   
   
       31 . The method of  claim 30 , wherein the medicament is further formulated as injection solution, instillation solution or infusion solution. 
   
   
       32 . The method of  claim 1 , moreover comprising the further formulation or reconstitution of the medicament for gastrointestinal, oral, nasal, pulmonary, dermal, transdermal or local application. 
   
   
       33 . The method of  claim 1 , moreover comprising the further formulation of the medicament into juice, capsules, tablets, suppositories or gels. 
   
   
       34 . The method of  claim 1 , moreover comprising the further formulation of the medicament into a powder for inhalation which is administered by use of an inhalator. 
   
   
       35 . A medicament, produced according to the method of  claim 1 .