US 5,807,706 AGranted

Method for making heteromultimeric polypeptides

AssigneeGenentech, Inc.

InventorsPaul J. Carter, Leonard G. Presta, John B. Ridgway

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Abstract

The invention relates to a method of preparing heteromultimeric polypeptides such as bispecific antibodies, bispecific immunoadhesins and antibody-immunoadhesin chimeras. The invention also relates to the heteromultimers prepared using the method. Generally, the method involves introducing a protuberance at the interface of a first polypeptide and a corresponding cavity in the interface of a second polypeptide, such that the protuberance can be positioned in the cavity so as to promote heteromultimer formation and hinder homomultimer formation. "Protuberances" are constructed by replacing small amino acid side chains from the interface of the first polypeptide with larger side chains (e.g. tyrosine or tryptophan). Compensatory "cavities" of identical or similar size to the protuberances are created in the interface of the second polypeptide by replacing large amino acid side chains with smaller ones (e.g. alanine or threonine). The protuberance and cavity can be made by synthetic means such as altering the nucleic acid encoding the polypeptides or by peptide synthesis.

Claims26

Dependent claims

A method of preparing a heteromultimer which comprises a first polypeptide and a second polypeptide which meet at an interface which has been genetically engineered to promote heteromultimer formation comprising:

(a) altering a first nucleic acid encoding said interface of the first polypeptide so that an original amino acid residue in the interface of the first polypeptide is replaced with an import amino acid residue having a larger side chain volume therefrom, thereby generating a protuberance on the interface of the first polypeptide;
(b) altering a second nucleic acid encoding said interface of the second polypeptide so that an original amino acid residue in the interface of the second polypeptide is replaced with an import amino acid residue having a smaller side chain volume therefrom, thereby generating a cavity in the interface of the second polypeptide, wherein the protuberance is position able in the cavity;
(c) introducing into a single host cell or separate host cells the first and second nucleic acids and culturing the cell or cells so that expression of the first polypeptide and second polypeptide occurs;
and
(d) allowing the heteromultimerto form in the single host cell and recovering the heteromultimer formed from the single host cell culture or recovering the first polypeptide and the second polypeptide from the separate host cell cultures and preparing the heteromultimer vitro.

2 more elements

Dependencies are inferred from the claim text.

Facts

Application: 08/433,105Filed 3 May 1995 · division
Priority: 1 Mar 1995
Granted: 15 Sep 1998
Term ends: Not estimatedThe source records don’t support an estimate.

Classification14 codes

C07K 16/468
Hybrid immunoglobulins: immunoglobulins having two or more different antigen binding sites, e.g. multifunctional antibodies
A61P 31/04
Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics: antibacterial agents
A61P 31/12
Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics: antivirals
C07K 14/70514
Immunoglobulin superfamily: cd4