Pharmaceutical compositions containing sc(Fv)2

• US 9,241,994 B2
• Filed: 06/09/2006
• Issued: 01/26/2016
• Est. Priority Date: 06/10/2005
• Status: Active Grant

• Current Assignee: Chugai Seiyaku Kabushiki Kaisha (Roche Holding AG)
• Original Assignee: Chugai Seiyaku Kabushiki Kaisha (Roche Holding AG)
• Inventors: Igawa, Tomoyuki
• Primary Examiner(s): Sang, Hong
• Application Number:
  US11/916,979
• Publication Number:
  US 20090214535A1
• Time in Patent Office: 3,518 Days
• US Class Current: 1/1
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61K 39/39591 Stabilisation, fragmentation
• CPC Class Codes: A61K 47/02 Inorganic compounds
• CPC Class Codes: A61K 47/183 Amino acids, e.g. glycine, EDTA or aspartame
• 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 9/19 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 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 16/2866 against receptors for cytokines, lymphokines, interferons
• CPC Class Codes: C07K 16/2869 against hormone receptors for antibodies against neuromediator receptors C07K16/286
• CPC Class Codes: C07K 16/30 from tumour cells
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/31 multispecific
• CPC Class Codes: C07K 2317/622 Single chain antibody (scFv)
• CPC Class Codes: C07K 2317/626 Diabody or triabody
• CPC Class Codes: C07K 2317/75 Agonist effect on antigen
• CPC Class Codes: C07K 2317/76 Antagonist effect on antigen, e.g. neutralization or inhibition of binding

Abstract

The present inventor discovered stabilizing agents/stabilizing conditions for suppressing isomerization reactions of sc(Fv)2. It was also discovered that the above-mentioned isomerization reactions can be suppressed through use of freeze-dried formulations. As disclosed herein, by applying the discovered stabilizing agents/stabilizing conditions or the freeze-dried formulation, the isomerization reaction of an sc(Fv)2-type molecule from the bivalent scFv type to the single chain diabody type, and/or the isomerization reaction from a single chain diabody type to a bivalent scFv type can be suppressed in both directions or one direction.

Claims

1. A method for suppressing the isomerization of a specific conformational isomer of sc(Fv)2 in a pharmaceutical composition comprising sc(Fv)2, the sc(Fv)2 having four variable regions connected by three peptide linkers arranged in order V1, linker, V2, linker, V3, linker, and V4, wherein each linker comprises 15 amino acids, and wherein the method comprises:

(a) providing the pharmaceutical composition, wherein the content ratio of the specific conformational isomer is 80% or more in the composition;

(b) adding a pH adjusting agent to the composition, thereby adjusting the pH of the composition to a pH in the range of 6.0 to 9.0; and

(c) adjusting the salt concentration of the composition to a concentration of about 300 mM, thereby producing a pharmaceutical composition in which isomerization of sc(Fv)2 is suppressed.

2. The method of claim 1, wherein the salt is selected from the group consisting of sodium chloride and magnesium chloride.

3. The method of claim 1, wherein the pH adjusting agent is selected from the group consisting of a sodium citrate buffer and histidine hydrochloride.

4. The method of claim 1, further comprising freeze-drying the pharmaceutical composition produced in (c).

5. A method for producing a pharmaceutical sc(Fv)2 composition comprising a specific conformational isomer of sc(Fv)2, the specific conformational isomer of sc(Fv)2 being either a bivalent scFv-type sc(Fv)2 or a single chain diabody type sc(Fv)2, wherein the content ratio of the specific conformational isomer is 80% or more in the pharmaceutical sc(Fv)2 composition, wherein the method comprises:

(a) providing a first composition comprising both bivalent scFv-type sc(Fv)2 and single chain diabody type sc(Fv)2 conformational isomers of an sc(Fv)2, wherein both conformational isomers of the sc(Fv)2 have four variable regions connected by three peptide linkers arranged in order V1, linker, V2, linker, V3, linker, and V4, wherein each linker comprises 15 amino acids, and wherein, in the bivalent scFv-type sc(Fv)2, V1 is associated with V2, and V3 is associated with V4, and, in the single chain diabody type sc(Fv)2, V1 is associated with V4, and V2 is associated with V3, and wherein the content ratio of the specific conformational isomer of sc(Fv)2 in the first composition is 20% or less;

(b) incubating the first composition at a pH in the range of 3.0 to 6.0 and at a salt concentration of 500 mM or less, for two or more days, to produce a second composition in which the content ratio of the specific conformational isomer of sc(Fv)2 is increased compared to the content ratio of the specific conformational isomer of sc(Fv)2 in the first composition;

(c) isolating the specific conformational isomer of sc(Fv)2 from the second composition, thereby forming a third composition comprising the isolated specific conformational isomer at a content ratio of 80% or more;

(d) stabilizing the specific conformational isomer of sc(Fv)2 in the third composition; and

(e) formulating the stabilized specific conformational isomer of sc(Fv)2 as a pharmaceutical composition.

6. The method of claim 5, wherein, in step (b), the salt concentration is 150 mM or less.

7. The method of claim 5, wherein step (d) comprises adjusting the pH and salt concentration of the third composition to a pH in the range of 6.0 to 9.0 and a salt concentration in the range of 50 mM to 1000 mM.

8. The method of claim 7, wherein, in step (d), the salt concentration is adjusted to 150 mM to 300 mM.

9. The method of claim 5, wherein the specific conformational isomer is a single chain diabody-type sc(Fv)2.

10. The method of claim 5, wherein the specific conformational isomer is a bivalent scFv-type sc(Fv)2.

11. The method of claim 5, wherein the method further comprises:

(f) freeze-drying the composition of (e).