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  Bifunctional moleculesUSPTO Application #: 20060024766Title: Bifunctional molecules Abstract: A chimeric antibody conjugate comprising an antigen binding region of a non-human antibody and an immunoglobulin constant region which comprises at least one CH domain or epitope thereof, with the proviso that the constant region is not a naturally occurring FC fragment. A bifunctional molecule for use in labelling an antibody derived from a first species, the bifunctional molecule comprising a binding-region which binds to the antibody of the first species or to one or more groups provided thereon, and a constant region derived from an antibody of a second species, the constant region comprising at least one CH domain or an epitope thereof. The present invention relates to bifunctional molecules and complexes which are useful as positive control reagents in antibody based diagnostic tests. The present invention also relates to polynucleotides encoding these bifunctional molecules, and to diagnostic assays involving the use of these molecules. (end of abstract)
Agent: Frommer Lawrence & Haug - New York, NY, US Inventors: John Leslie Atwell, Peter Leonard Devine, Gregory Coia, Alexander Andrew Kortt, Gillian Wendy Perry, Peter Gregory Bundesen USPTO Applicaton #: 20060024766 - Class: 435007320 (USPTO) Related Patent Categories: Chemistry: Molecular Biology And Microbiology, Measuring Or Testing Process Involving Enzymes Or Micro-organisms; Composition Or Test Strip Therefore; Processes Of Forming Such Composition Or Test Strip, Involving Antigen-antibody Binding, Specific Binding Protein Assay Or Specific Ligand-receptor Binding Assay, Involving A Micro-organism Or Cell Membrane Bound Antigen Or Cell Membrane Bound Receptor Or Cell Membrane Bound Antibody Or Microbial Lysate, Bacteria Or Actinomycetales The Patent Description & Claims data below is from USPTO Patent Application 20060024766. Brief Patent Description - Full Patent Description - Patent Application Claims
FIELD OF THE INVENTION [0001] The present invention relates to bifunctional molecules and complexes which are useful as a positive control reagents in antibody based diagnostic tests. The present invention also relates to polynucleotides encoding these bifunctional molecules, and to diagnostic assays involving the use of these molecules. BACKGROUND OF THE INVENTION [0002] Infection of humans by many micro-organisms leads to the initiation of a humoral immune response that can be used in the diagnosis of the disease. In the early acute phase of the infection, specific IgM class antibodies are the first to appear in serum 1-4 weeks after the onset of symptoms and last for up to three months. IgG class antibodies appear later and remain elevated throughout the patient's life. Detection of an IgM response is indicative of a recent or current infection, while the presence of an elevated IgG response is a marker for past exposure to the causative agent. Specific IgM or IgG responses to a particular infectious agent can be measured by antibody based diagnostic tests such as ELISA, immiunochromatography, particle agglutination ELISA, biosensor or other similar assays. [0003] These assays require the use of reactive human sera as a positive control. The positive control reagent is usually serum taken from a patient or animal which is known to have a positive reaction to the particular antigen under test. If the test is designed to distinguish between early and late infection (via the differentiation between immunoreactive IgM, for early infection and IgG, for late or previous infection), the positive control serum or reagent should contain immunoreactive antibody of the correct immunoglobulin class. [0004] It is becoming increasingly difficult to source sufficient quantities of immune human sera or plasma, particularly as diagnostic tests for rarer diseases become available. Collection of blood for IgM controls from patients in early stages of infection when clinical symptoms are generally most severe poses significant ethical problems, particularly if the disease primarily affects juveniles. Other drawbacks include the requirement for consistent collections from remote locations, the need to standardise each batch and to check for contamination with infectious agents such as HIV, hepatitis B and hepatitis C. There are also problems in obtaining control sera for specific endemic diseases in communities where the donation of blood or blood products is socially unacceptable. [0005] There is therefore a need for a source of positive control reagents which does not rely on being obtained from human donors. [0006] Hybridoma technology provides a plentiful supply of monoclonal antibodies, but as these are generally of murine origin, they do react with binding reagents used to quantify human antibodies. Intact, functional mouse/human chimeric antibodies have been described in the literature for some time (Boulianne et al. 1984, Nlorrison et al., 1984; Winter et al. 1991). In these constructs the antigen binding function residing in a mouse Fab or Fv fragment has been grafted on to a human Ig backbone and expressed in hebridoma cells. In some cases these reshaped molecules have been designed for human therapy, utilising the effector functions of the human Fc for targeting (Reichmann et al., 1988). Others have been designed as positive control reagent substitutes (Hamilton, 1990, 1991), where V.sub.H and V.sub.L regions from a mouse monoclonal antibody of desired specificity have been grafted onto either a human IgG or IgM backbone. [0007] Synthetic positive control reagents are available from a limited number of sources. U.S. Pat. No. 4,929,543 relates to chimeric antibody fragments where Fab or F(ab')2 fragments of non human origin, with specificity for the desired antigen, are chemically coupled to human Fc fragments in order to confer upon the reactive non-human Fab fragments epitopes recognised by class specific anti human immunoglobulin antisera. This reference does not teach or suggest coupling non-human Fab or F(ab')2 fragments to individual CH domains in order to provide epitopes for recognition by class specific anti human immunoglobulin antisera. Furthermore, production of the chimeric fragments is entirely by synthetic routes based upon digestion of antibodies, purification of fragments and chemical linking to create the chimera. [0008] Labor Diagnostika GmbH of Heiden, Germany have produced synthetic positive control reagents which are formed by chemical attachments of non human Fab fragments and human Fc fragments onto a latex bead. These attachments confer upon the bead the twin properties required of a positive control reagent--specific antigen binding and human immunoglobulin class specific epitopes. [0009] A process for producing positive control reagents which circumvents the requirement to manipulate full length Fc fragments, or to manipulate VH and V.sub.L sequences for each new control reagent specificity, is desirable. SUMMARY OF THE INVENTION [0010] The present inventors have now developed bifunctional molecules which may be used as positive control reagents in antibody based diagnostic tests. [0011] In one aspect of the present invention, the bifunctional molecule is a chimeric antibody conjugate comprising a first region which binds a specific antigen and a second region comprising at least one constant domain sequence derived from a class specific immunoglobulin. This conjugate, which may be used directly as a positive control reagent, avoids the inconvenience of manipulating full length or naturally occurring Fc fragments. Furthermore, the conjugate may be readily produced by recombinant DNA technology. [0012] Accordingly, in a first aspect the present invention provides a chimeric antibody conjugate comprising an antigen binding region derived from a non-human antibody and a constant region which comprises at least one C.sub.H domain or epitope thereof, with the proviso that the constant region is not a naturally occurring Fc fragment. [0013] When used herein, "naturally occurring Fc fragment" means a full length naturally occurring Fc fragment which may be derived by proteolytic digestion of an intact antibody molecule. For example, a naturally occurring Fc fragment of IgM will comprise domains C.sub.H2, C.sub.H3 and C.sub.H4, whereas a naturally occurring IgG Fc fragment will comprise C.sub.H2 and C.sub.H3 domains. [0014] By "chimeric" we mean that the constant region is derived from a different species than the antigen binding region. [0015] In a preferred embodiment the non-human antigen binding region comprises or consists of a non-human Fab fragment or part thereof. The non-human antigen binding region may comprise or consist of an scFv fragment. [0016] In a further preferred embodiment the non-human antigen binding region is derived from a mouse. [0017] In a preferred embodiment the constant region is derived from a human antibody. It will be appreciated, however, that the constant region may be a non-human (such as bovine, canine, ovine, equine, feline or caprine) constant region in cases where the chimeric construct is to be used as a positive antibody control in assays involving sera derived from non-human species. [0018] The constant region may consist of a non-naturally occurring combination of Cal domains or epitopes thereof. The constant region may consist of two C.sub.H domains of the same type, for example, two C.sub.H3 domains. Alternatively, the constant region may consist of two different domains. The two different domains, or epitopes thereof may be derived from antibodies of different classes. In a preferred embodiment, the constant region consists of a single C.sub.H domain. [0019] In a particularly preferred embodiment of the present invention the chimeric antibody conjugate is suitable as a positive IgM control and the constant region comprises one or more C.sub.H3.mu. domains. [0020] In a further preferred embodiment the non-human antigen binding region binds to an epitope derived from an infectious agent selected from but not limited to dengue virus, rubella virus, herpes virus, palivovirus, human glycophorin, Rickettsia sibirica, Burlkioleria pseudomallei, Salmonella typhi or paratyphi, Leptospira interrogans, Plasmodium falciparum/vivax, Japanese encephalitis virus, Yellow fever virus, Bordetella pertlussis/parapertussis, Candida albicans/kruzei, Varicella zoster virus, HIV, Hepatitis viruses, Human papilloina virus, Epstein-Barr virus, Ross River virus, Brucella aboltis, Human herpesvirus-6, Parvovirus B19, Coxiella burnettii, Herpes simplex viruses 1&2, Rickettsia rickettsii, Conori australis, Rickettsia tsutsugamuushi. [0021] In a second aspect the present invention provides a recombinant polynucleotide molecule comprising a sequence encoding a non-human V.sub.H region, a sequence encoding a non-human V.sub.L region, a sequence encoding a flexible linker positioned between the V.sub.H region sequence and the V.sub.L region sequence, and a heterologous sequence encoding a C.sub.H domain or epitope thereof. Continue reading... Full patent description for Bifunctional molecules Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Bifunctional molecules patent application. ###  How KEYWORD MONITOR works... a FREE service from FreshPatents1. Sign up (takes 30 seconds). 2. Fill in the keywords to be monitored. 3. Each week you receive an email with patent applications related to your keywords. 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