Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Patent
1993-10-18
1996-11-12
Saunders, David
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
435 791, 435 792, 4351722, 43524027, 435972, 435975, 436518, 436548, 436819, 5303873, 5303911, G01N 33535, G01N 33577
Patent
active
055739202
DESCRIPTION:
BRIEF SUMMARY
This invention relates to antibodies and is particularly, though not exclusively, concerned with diagnostic and therapeutic methods using monoclonal or polyspecific, such as bi-or tri-specific antibodies.
Monoclonal-based antibody assays have not achieved full potential as they generally have to be performed by trained operators in a laboratory. Even relatively simple assays require washing steps and multiple manual addition of reagents. There is a need for a one-step system, which would have a wide field of applications.
Monoclonal antibodies have also found application in treatment of disease. For example, monoclonal antibody conjugates have been used to localize and treat tumours in the body, destroying the tumour with toxic agents, including ricin and radioiodine, attached to the antibody protein.
Bispecific antibodies have been developed from monoclonal antibody technology, in the example of bispecific immunoglobulin G each bispecific antibody has two antigen binding sites of differing specificities. Bispecific antibodies can be produced by fusing two different hybridomas which respectively secrete monoclonal antibodies against the antigens of interest to form a single hybrid-hybridoma or "fusoma", (sometimes called a "polydoma") (Songsivilai, S and Lachmann P.J (1990) Clin Exp Immunol 79, 315 and Suresh MR et al (1986) Proc. Natl Acad Sci USA 83, 7989 and GB2169921A). Parent hybridomas can be removed by standard HAT selection or introduction of selectable drug resistance (De Lau B.M, et al (1989) J. Immunol Methods 117 ,1). The first bispecific antibodies produced were used in a conventional immunoassay (Milstein C., and Cuello A.C., (1983) Nature 305 537). The antibodies were produced by fusing a monoclonal antibody-secreting cell with splenocytes from an immune mouse. The first binding site was specific for an antigen of interest. The second binding site of the enzyme was specific for a marker enzyme. The immunoassay demonstrated increased assay sensitivity, reduction in signal-to-noise ratio, simplification of staining procedures, and preservation of ultrastructural detail.
Bispecific antibodies have found extensive application in novel therapy regimes targeting effector toxins, which remain bound to the antibody, to rumours (Corvalan JRF et al (1987) Cancer Immunol Immunother 24 133), crosslinking cellular antigens on cytotoxic killer cells to rumour targets (Nitta T., et al (1990) Lancet 335 368) and Fanger MW and Guyre PM, Tibtech 9, 375-380 (1991). Other methods of producing bi-and trispecific antibodies, such as chemical linkage, are reviewed in the latter paper.
WO90/07714 discloses an immunoassay in which an enzyme is stabilised by binding to a bispecific antibody against heat denaturation. W091/09134 discloses a bispecific antibody capable of binding both to an enzyme that converts an inactive anticancer prodrug into its active form and to a human cancer cell. An immunocomplex comprising the antibody and the enzyme can be administered to cancer patients together with the inactive prodrug to selectively kill cancer cells with minimal side effects. The enzyme remains bound to the antibody in an active form. Also disclosed are methods of producing polydomas.
It is an object of the invention to provide an immunoassay method involving fewer, preferably only one, reaction steps than conventional immunoassay methods.
According to one aspect of the invention, there is provided a method in which binding of an antigen to one antibody antigen binding site causes release of another antigen from an adjacent second antibody antigen binding site. Whilst not wishing to be bound by theory the applicants believe that steric hindrance between the incoming antigen and the bound antigen causes release of the bound antigen from the second antibody binding site. The first and second antibody antigen binding sites may be provided by the same multispecific antibody or by different antibodies which are physically adjacent. The term "multispecific" embraces all antibodies having more than one antigen binding site su
REFERENCES:
patent: 4446233 (1984-05-01), Auditore-Hargreaves et al.
patent: 4578360 (1986-03-01), Smith
patent: 4686181 (1987-08-01), Dona435 7
Suresh et al., 1986, Advantages of bispecific hybridomas in one-step immunocytochemistry and immunoassays, Proc. Natl. Acad. Sci. USA, 83:7989-7993.
De Lau et al., 1989, Production of hybrid hybridomas based on HAT.sup.S -neomycin.sup.r double mutants, J. Immunol. Methods, 117:1-8.
Saunders David
Surface Active Limited
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