WO1987003205A1

WO1987003205A1 - Enzyme-coupled antibodies

Info

Publication number: WO1987003205A1
Application number: PCT/GB1986/000711
Authority: WO
Inventors: Selby John Starkie
Original assignee: Coral Sociedade Brasileira De Pesquisas E Desenvol
Priority date: 1985-11-22
Filing date: 1986-11-21
Publication date: 1987-06-04
Also published as: GB8528761D0; EP0250477A1

Abstract

An enzyme-coupled antibody comprises an enzyme, e.g. coupled to a monoclonal antibody, which can catalyse reactions which result in the death of cells bearing antigenic sites with which the antibody can bind. Such products have therapeutic value.

Description

ENZYME-COUPLED ANTIBODIES This invention relates to antibodies and their use in killing specific organisms or tissue cells which carry appropriate antigens. The invention is thus of potential value in therapy.

Antibodies bind specifically with appropriate antigens. However, the presence of specific antibodies in body fluids, in vivo, and their binding with antigens of organisms or tissue cells, are not necessarily sufficient to kill the organisms or tissue cell. The primary causes of killing are phagocytosis, macrophages, and lysis by complement. Antibodies bound to antigens on the organism or tissue cell often mediate opsonisation or the activation of complement. The role of antibodies in this respect is incompletely understood, but may be related to the class or subclass of the antibody, the nature and distribution of the antigen, the availability and activation of various kinds of phagocytic and accessory cells and the concentration and nature of the components and inhibitors of complement.

It is of potential value to make the killing of cells carrying specific antigens dependent only on the relative concentration of antibody (or modified antibody) to antigen, and independent of other effect or agents. WO-A-8303679 discloses "polydomas", i.e. the product of the fusion of a hybrido a with a B-lymphocyte or another hybridoma. The polydomas produce a hybrid monoclonal antibody having dual specificity, e.g. one specificity directed against a target antigen and the other against a moiety which permits a diagnosis to be made or which delivers an agent lethal to the target antigen or associated tissue.

It is well known to couple enzymes to antibodies. For example, enzymes such as alkaline phosphatase and horseradish peroxidase have been coupled to antibodies, and the properties of the enzyme used in order to detect antibody-antigen binding. For the aim of detection, it is appropriate that the substrate of the enzyme should undergo a readily-detectable (e.g. colour) change in the presence of the enzyme.

Toxins such as ricin, gelonin or diphtheria toxin may be bound to antibodies. Such toxins are enzymes and result in the death of cells by degradation of components of the protein synthetic mechanisms within the cell. To achieve this effect, the complex of antibody and enzyme requires introduction into the cytoplasm of the cell, usually by a mechanism of the cell itself characteristic of a fluid cell membrane. In general, organisms with non-fluid cell walls do not internalise complex molecules of this kind, so that toxin-conjugated antibodies would not kill some organisms.

In an enzyme-coupled antibody according to the present invention, the enzyme has the ability to catalyse degradative or synthetic reactions which result in the death of the cell.

The present invention is based on the concept that an enzyme attached to an antibody is brought into close proximity with the cell when the antibody binds with the antigen on the cell. The substrate of the enzyme is a substance close to, or part of, the cell or the antigen, or the fluid around the cell or the antigen. The antibody need not have been produced from a polydoma, but can retain a single specificity.

Examples of suitable enzymes are lipases, phospholipases, proteases and glycases, substrates for which might be lipids, phospholipids, proteins, sugars and glycoproteins in the cell walls. Glucose oxidase is an example of an enzyme which has a natural plasma substrate, glucose; one of the products of glucose oxidation is hydrogen peroxide which can kill cells by oxidation of components of the cell wall.

Many of the components of complement are enzymes. When some forms of antibody bind with some forms of antigen, the enzymatic components of complement are activated in a cascade, i.e. each activated component catalyses the activation of the next component. The final components, activated C789, constitute a phospholipase which digests parts of certain membranes, such as cell walls and basement membranes. The classical pathway of complement activation is initiated by the activation of the first components by antibody bound to an antigen, but not by a free antibody. By contrast, the concept of the present invention is that the enzyme is irreversibly bound to the antibody.

It will readily be appreciated that an enzyme- coupled antibody according to the invention can be used therapeutically. For example, in order to kill an organism or cell which has been detected in a subject, an enzyme selected to kill the cell can be coupled to an antibody, e.g. a monoclonal antibody, specific to an antigen of the organism or cell, and the coupled product administered to the subject. Coupling can be by any convenient, conventional method, e.g. the glutaraldehyde or the SPDP method. Administration may also be by any suitable conventional method, e.g. inoculation; for inoculation, a conventional composition may be formulated which comprises the product and a physiologically- acceptable excipient. Example

A monoclonal antibody against Shigella as described in Example 1 of WO-A-8600646 is coupled, not with alkaline phosphatase as described in section G therein, but with glucose oxidase. Administration of the enzyme-coupled antibody into plasma containing the antigen (as determined by conventional procedures) leads to (i) antibody-antigen reaction and thus the proximity of enzyme and antigen-bearing cell, and (ii) the local production of cell-toxic hydrogen peroxide by enzymatic oxidation of glucose in the plasma.

Claims

1. An enzyme-coupled antibody, in which the enzyme is ---^■ characterised by ability to catalyse reactions which result in the death of cells bearing antigenic sites with ^ 5 which the antibody can bind.

2. An enzyme-coupled antibody according to claim 1, in which the enzyme catalyses destruction of the cell wall.

3. An enzyme-coupled antibody according to claim 1 , in which the substrate for the enzyme is present in plasma

10 and the cells are destroyed by a product of the enzymatic reaction.

4. An enzyme-coupled antibody according to any preceding claim, in which the antibody is monoclonal.

5. An enzyme-coupled antibody according to any 15 preceding claim, for therapeutic use.

6. A method for treating a subject hosting an antigen, which comprises administering to the subject an effective amount of an enzyme-coupled antibody, in which the antibody is specific for the antigen and the enzyme has

_.20 the ability to cause destruction of cells bearing the antigen.

7. A method according to claim 6, in which the enzyme-coupled antibody is as defined in any of claims 2 to 4.

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