Antibody constant region variant

a constant region and antibody technology, applied in the field of constant region antibodies, can solve the problems of unfavorable binding to the fc receptor, unfavorable method, and posing immunogenicity risks, and achieve the effects of low heterogeneity, high heterogeneity, and reduced antibody heterogeneity

Inactive Publication Date: 2019-07-11
CHUGAI PHARMA CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0039]The present invention provides constant regions that can confer to antibodies properties desirable for pharmaceutical agents. By means of amino acid sequence alterations, the constant regions of the present invention can improve the following antibody properties to conditions favorable for pharmaceutical agents.
[0040]Decrease in Antibody Heterogeneity:
[0041]Polypeptides obtainable by expressing a DNA encoding a certain amino acid sequence should theoretically be homogeneous polypeptide molecules consisting of the same amino acid sequence. However, in practice, when a DNA encoding an antibody is expressed in suitable hosts, heterogeneous polypeptides with different structures may be formed due to various factors.
[0042]In the production of antibodies, an antibody population comprising many heterogeneous polypeptides can be referred to as having high heterogeneity. The constant regions of the present invention have the causes of heterogeneity removed by amino acid sequence alteration. Therefore, constructing antibodies using the constant regions of the present invention enables production of antibodies with low heterogeneity. Specifically, by introducing alterations provided by the present invention into heavy chain constant regions of antibodies, the homogeneity of the antibodies can be maintained at a high level. Suppressing the antibody heterogeneity to a low level means ameliorating the heterogeneity and this is an important objective in maintaining the quality of pharmaceuticals. Therefore, the constant regions of the present invention contribute to the maintenance of the quality of antibody-containing pharmaceuticals.
[0043]Improvement of Pharmacokinetics:
[0044]In a preferred embodiment, the present invention contributes to improvement of antibody pharmacokinetics. Specifically, when specific amino acid residues are altered in an antibody constant region of the present invention, blood concentration of the antibody composed of this constant region is maintained for a longer time than an antibody without amino acid sequence alterations. Maintaining blood concentration for as long a time as possible means that, when an antibody is administered as a pharmaceutical, its therapeutic effect can be maintained for a long time with a smaller amount of antibody. Alternatively, the antibody can be administered with wider intervals and smaller number of administrations.

Problems solved by technology

Thus, binding to Fcγ receptor is considered unfavorable in antibody pharmaceuticals intended for neutralizing the biological activity of an antigen from the perspective of side effects and immunogenicity.
A method for impairing the binding to Fcγ receptor is to alter the subtype of the IgG antibody from IgG1 to IgG2 or IgG4; however, this method cannot completely inhibit the binding (Non-patent Document 6).
However, these molecules contain new non-native peptide sequences of nine to twelve amino acids, which may constitute a T-cell epitope peptide and thus pose an immunogenicity risk.
Thus, it was a challenge to secure stability at high concentrations.
However, there has been no report published on the improvement of the stability of IgG at high concentrations by introducing amino acid substitutions into its constant region.
A method for prolonging the antibody half-life in plasma has been reported and it substitutes amino acids in the constant region (Non-patent Documents 14 and 15); however, introduction of non-native sequences into the constant region is unpreferable from the perspective of immunogenicity risk.
It is not easy to manufacture them as a pharmaceutical in a large scale while maintaining differences of objective substance / related substance-related heterogeneity between productions.
However, altered constant regions that meet all of the above requirements have not yet been reported.

Method used

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  • Antibody constant region variant
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Examples

Experimental program
Comparison scheme
Effect test

example 1

[Example 1] Improvement of C-Terminal Heterogeneities of IgG Molecules

[0366]Construction of an expression vector for H-chain C-terminal ΔGK antibody

[0367]Heterogeneities of the C-terminal sequence of the IgG antibody H chain that have been reported are deletion of the C-terminal amino acid lysine residue, and amidation of the C-terminal carboxyl group due to deletion of both of the two C-terminal amino acids, glycine and lysine residues (Anal Biochem. 2007 Jan. 1; 360(1): 75-83). In TOCILIZUMAB which is an anti-IL-6 receptor antibody, the main component is a sequence in which the C-terminal amino acid lysine present on the nucleotide sequence is deleted by post-translational modification, but an accessory component with remnant lysine and an accessory component with an amidated C-terminal carboxyl group produced by deletion of both glycine and lysine are also present as heterogeneities. It is not easy to manufacture such an antibody as a pharmaceutical in a large scale, while mainta...

example 2

[Example 2] Novel Constant Regions with Reduced Heterogeneity, which Retain the Stability of Natural IgG2

Heterogeneity of Natural IgG1 and Natural IgG2

[0373]For antibody pharmaceuticals against cancer such as those that kill target cells with effector functions and such, IgG1 constant region (isotype) having effector function is preferred. However, for antibody pharmaceuticals that neutralize the functions of a target antigen or antibody pharmaceuticals that bind to target cells but do not kill them, binding to Fcγ receptors is not preferred.

[0374]As methods for decreasing the binding to Fcγ receptors, the method of changing the IgG antibody isotype from IgG1 to IgG2 or IgG4 has been considered (Ann Hematol. 1998 June; 76(6): 231-48), and from the viewpoint of binding to Fcγ receptor I and pharmacokinetics of each isotype, IgG2 was considered to be more desirable than IgG4 (Nat Biotechnol. 2007 December; 25(12): 1369-72). On the other hand, when developing antibodies into pharmaceut...

example 3

[Example 3] Pharmacokinetics-Improving Effect of Novel Constant Region M58-k0 Pharmacokinetics of IgG-Type Antibodies

[0394]The prolonged retention (slow elimination) of IgG molecule in plasma is known to be due to the function of FcRn which is known as a salvage receptor of IgG molecule (Nat. Rev. Immunol. 2007 September; 7(9): 715-25). When taken up into endosomes via pinocytosis, IgG molecules bind to FcRn expressed in endosomes under the acidic conditions within the endosomes (approx. pH 6.0). While IgG molecules that do not bind to FcRn are transferred and degraded in lysosomes, those bound to FcRn are translocated to the cell surface and then released from FcRn back into plasma again under the neutral conditions in the plasma (approx. pH 7.4).

[0395]Known IgG-type antibodies include the IgG1, IgG2, IgG3, and IgG4 isotypes. The plasma half-lives of these isotypes in human are reported to be about 36 days for IgG1 and IgG2; about 29 days for IgG3; and 16 days for IgG4 (Nat. Biotec...

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Abstract

By altering amino acid sequences, the present inventors successfully produced constant regions that can confer antibodies with particularly favorable properties for pharmaceutical agents. When used to produce antibodies, the altered constant regions produced according to the present invention significantly reduce heterogeneity. Specifically, the antibody homogeneity can be achieved by using antibody heavy chain and light chain constant regions introduced with alterations provided by the present invention. More specifically, the alterations can prevent the loss of homogeneity of antibody molecules due to disulfide bond differences in the heavy chain. Furthermore, in a preferred embodiment, the present invention can improve antibody pharmacokinetics as well as prevent the loss of homogeneity due to C-terminal deletion in antibody constant region.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]This application is a divisional application of U.S. application Ser. No. 14 / 680,250, filed on Apr. 7, 2015, which is a divisional of U.S. application Ser. No. 13 / 257,145, having a 371 (c) date of Nov. 22, 2011, which is the National Stage of International Application Serial No. PCT / JP2010 / 054767, filed on Mar. 19, 2010, which claims priority to Japanese Application Serial Nos. 2009-068631 and 2009-213901, filed on Mar. 19, 2009, and Sep. 16, 2009, respectively.TECHNICAL FIELD[0002]The present invention relates to antibody constant regions with an altered amino acid sequence, and antibodies comprising these constant regions.BACKGROUND[0003]Antibodies are drawing attention as pharmaceuticals as they are highly stable in plasma (blood) and have few side effects. Of these, a number of IgG-type antibody pharmaceuticals are available on the market and many antibody pharmaceuticals are currently under development (Non-patent Documents 1 and 2)....

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): C07K16/00C07K16/28
CPCC07K16/00C07K16/2866C07K16/2875C07K2317/528C07K2317/524C07K2317/52C07K2317/515C07K2317/92C07K2317/53C07K2317/526C07K2317/522C07K2317/76C07K2317/71C07K2317/72C07K2317/94
Inventor IGAWA, TOMOYUKIKURAMOCHI, TAICHIMAEDA, ATSUHIKOSHIRAIWA, HIROTAKE
Owner CHUGAI PHARMA CO LTD
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