Dimer VEGF antagonist formulations

• US 8,404,638 B2
• Filed: 01/04/2012
• Issued: 03/26/2013
• Est. Priority Date: 03/25/2005
• Status: Active Grant

• Current Assignee: Regeneron Pharmaceuticals Incorporated
• Original Assignee: Regeneron Pharmaceuticals Incorporated
• Inventors: Dix, Daniel, Frye, Kelly, Kautz, Susan
• Primary Examiner(s): STOICA, ELLY GERALD
• Application Number:
  US13/343,214
• Publication Number:
  US 20120101035A1
• Time in Patent Office: 447 Days
• Field of Search: None
• US Class Current: 514/8.1
• CPC Class Codes: A61K 31/4172 Imidazole-alkanecarboxylic acids, e.g. histidine
• CPC Class Codes: A61K 31/7012 Compounds having a free or esterified carboxyl group attached, directly or through a carbon chain, to a carbon atom of the saccharide radical, e.g. glucuronic acid, neuraminic acid gluconic acid A61K31/191; ascorbic acid A61K31/375
• CPC Class Codes: A61K 38/16 Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof enzyme inhibitors A61K38/005
• CPC Class Codes: A61K 38/179 for growth factors; for growth regulators
• CPC Class Codes: A61K 39/39591 Stabilisation, fragmentation
• CPC Class Codes: A61K 47/02 Inorganic compounds
• CPC Class Codes: A61K 47/10 Alcohols; Phenols; Salts thereof, e.g. glycerol; Polyethylene glycols [PEG]; Poloxamers; PEG/POE alkyl ethers
• CPC Class Codes: A61K 47/12 Carboxylic acids; Salts or anhydrides thereof
• CPC Class Codes: A61K 47/183 Amino acids, e.g. glycine, EDTA or aspartame
• CPC Class Codes: A61K 47/22 Heterocyclic compounds, e.g. ascorbic acid, tocopherol or pyrrolidones
• CPC Class Codes: A61K 47/26 Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
• CPC Class Codes: A61K 9/0019 Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner non-active ingredients are additionally classified in A61K47/00
• CPC Class Codes: A61K 9/08 Solutions composition of solutions A61K47/00
• CPC Class Codes: A61K 9/19 lyophilised , i.e. freeze-dried, solutions or dispersions lyophilised products with subsequent particle size reduction A61K9/14; granules or pellets made by lyphilisation A61K9/1682; solid oral dosage forms made by lyophilisation A61K9/2095; lyophilisation additives A61K47/00
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 7/00 Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07K 14/71 for growth factors; for growth regulators
• CPC Class Codes: C07K 16/22 against growth factors ; against growth regulators
• CPC Class Codes: C07K 2318/20 : Antigen-binding scaffold molecules wherein the scaffold is not an immunoglobulin variable region or antibody mimetics
• CPC Class Codes: C07K 2319/30 : Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
• CPC Class Codes: Y02A 50/30 : Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

Formulations of a vascular endothelial growth factor (VEGF)-specific fusion protein antagonist are provided including a pre-lyophilized formulation, a reconstituted lyophilized formulation, and a stable liquid formulation. Preferably, the fusion protein has the sequence of SEQ ID NO:4.

Claims

1. A non-liquid formulation of a vascular endothelial growth factor (VEGF) antagonist made according to the steps of:

(a) combining in a liquid solution the following excipients;

(i) about 50 mg/mL of a VEGF antagonist, which is a dimer consisting of two identical polypeptides consisting of amino acids 27-457 of SEQ ID NO;

4;

(ii) about 10 mM histidine, pH about 6.3-6.5;

(iii) about 1.5% polyethylene glycol (PEG) 3350;

(iv) about 5% sucrose; and

(v) about 1.5% glycine; and

(b) lyophilizing the combination of step (a), wherein, following storage of the lyophilized combination obtained after step (b) at 5°

C. for about 6 months, at least 98% of the VEGF antagonist is present in native conformation as measured by size exclusion chromatography upon reconstitution.

2. The non-liquid formulation of claim 1, wherein, following storage of the lyophilized combination obtained after step (b) at 5°

C. for about 3 months, the VEGF antagonist comprises at least 65% of maximum biological activity.

3. The non-liquid formulation of claim 2, wherein the biological activity is the ability of the VEGF antagonist to inhibit the biological effects of human VEGF on a mouse Baf/2 VEGFR1/EpoR cell line reported as IC₅₀.

4. The non-liquid formulation of claim 1, wherein, following storage of the lyophilized combination obtained after step (b) at 5°

C. for about 3 months, the VEGF antagonist binds VEGF at a level of at least 94% of maximum binding.

5. The non-liquid formulation of claim 4, wherein the binding of the VEGF antagonist to VEGF is expressed as IC₅₀ as determined by ELISA.

6. A method of producing a liquid formulation of a VEGF-specific fusion protein antagonist, comprising reconstituting the lyophilized formulation of claim 1 with liquid, wherein a liquid formulation is generated.

7. The method of claim 6, wherein the liquid is sterile water or bacteriostatic water.

8. A liquid formulation made according to the method of claim 7.