Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Patent
1996-08-21
1998-08-25
Schwadron, Ronald B.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
435 71, 435960, 435331, 435335, 435336, 435337, 435338, 435332, 5303882, 53038823, 53038826, 5303892, 5303911, 935104, 935108, G01N 33573, G01N 33536, G01N 33577, C07K 1624
Patent
active
057982139
DESCRIPTION:
BRIEF SUMMARY
This application is a 371 of PCT/JP95/02661 filed Dec. 25, 1995.
TECHNICAL FIELD
The present invention relates to monoclonal antibodies which are useful for the detection of human thymidine phosphorylase and the platelet-derived endothelial cell growth factor (PD-ECGF), i.e., for the monitoring via diagnosis and treatment of a variety of tumors, diseases involving abnormal angiogenesis such as metastasis of tumors, rheumatic arthritis, diabetic retinitis, immature cataract, senile macular degeneration, etc., as well as for use in drug delivery systems for medicine, etc.
TECHNICAL BACKGROUND
Thymidine phosphorylase is an enzyme essential to the metabolism of thymidine and has been reported to manifest its activities in different tissues (liver, lungs, small intestine, large intestine, placenta, etc.) Thymidine phosphorylase has also been reported to exhibit higher levels of Bull., 34, 4225-4232 (1986)!. Because of this feature, thymidine phosphorylase is a target enzyme for an anticancer agent.
It is also reported that activity of PD-ECGF becomes elevated when abnormal (1989)!, which makes PD-ECGF a marker of diseases accompanying abnormal angiogenesis.
Recently, human thymidine phosphorylase has been reported to be genetically (1992), and J. biochem., 114, 9-14 (1993)!. Measurement of thymidine phosphorylase activity and PD-ECGF activity is useful in the diagnosis of not only malignant tumors but also diseases involving abnormal angiogenesis, such as rheumatic arthritis, diabetic retinitis, immature cataract, senile macular degeneration, etc.
Conventionally, activities of human thymidine phosphorylase and PD-ECGF have been determined by measuring them in tissue. Therefore, excision of necessary amounts of tissue and fractionation of a crude enzymatic fluid must be performed, calling for cumbersome handling techniques. In addition, detailed comparison on the intercellular level cannot be made by conventional methods, raising a great problem in both the practical and accuracy aspects.
DISCLOSURE OF THE INVENTION
Accordingly, an object of the present invention is to provide simple, widely-applicable, and practical means which enables specific measurement of thymidine phosphorylase/PD-ECGF.
The present inventors conducted careful studies, created monoclonal antibodies using as immunogens certain peptides in human thymidine phosphorylase and PD-ECGF protein, and found that they specifically recognize human thymidine phosphorylase and PD-ECGF and thus are useful for the diagnosis of various diseases accompanying elevated human thymidine phosphorylase activity and PD-ECGF activity. The present invention was accomplished based on this finding.
Accordingly, the present invention provides a monoclonal antibody against a peptide having an amino acid sequence described in Sequence No. 1, a peptide having an amino acid sequence described in Sequence No. 2, or against a peptide recognized by the presence of either one of these two peptides which serves as an antigen site.
The present invention also provides an immunoassay for human thymidine phosphorylase and/or PD-ECGF using the above monoclonal antibody.
BRIEF DESCRIPTION OF THE DRAWINGS
FIGS. 1A-1B are a chart showing profiles of reaction specificity of monoclonal antibodies of the present invention.
FIG. 2 is a chart showing results of comparison in expression between human breast cancerous tissue which reacts with a monoclonal antibody of the present invention and its adjacent normal tissue (3 cases).
FIG. 3 is a photograph showing the result of immunohistochemical staining of a slice from healthy human large intestine tissue with which a monoclonal antibody of the present invention reacts with specificity.
FIG. 4 is a photograph showing the result of immunohistochemical staining of a slice from human large intestinal cancerous tissue with which a monoclonal antibody of the present invention reacts with specificity.
BEST MODE FOR CARRYING OUT THE INVENTION
The peptide having an amino acid sequence described in Sequence No. 1 constitutes a portion of
REFERENCES:
patent: 5227302 (1993-07-01), Heldin et al.
Sumizawa, T. et al. "Thymidine Phosphorylase Acitvity Associated with Platelet-Derived Endothelial Cell Growth Factor", J. Biochem. (1993) vol. 114, No. 1, pp. 9-14.
Osamu Kanemitsu "Introduction of Antibody", Jan. 25, 1994 (25.01.94), Chijinshokan pp. 75-144.
Furukawa et al., Nature, 356:668, 1992.
Ishikawa, F. et al. "Identifiecation of Angiogenic Activity and the Cloning and Expression of Platelet-Derived Endothelial Cell Growth Factor", Nature (1989) vol. 338, pp. 557-562.
Takeuchi et al., Arth. and Rheumat., 37:662-672, 1994.
Akiyama Shin-ichi
Miyadera Kazutaka
Takebayashi Yuji
Yamada Yuji
Schwadron Ronald B.
Taiho Pharmaceutical Co. Ltd.
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