Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Reexamination Certificate
1998-08-14
2001-09-18
Ceperley, Mary E. (Department: 1641)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
C435S007500, C530S388900, C530S389800, C530S391100, C530S391300, C530S404000, C530S405000, C530S807000
Reexamination Certificate
active
06291198
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to an antibody recognizing a pyrazine derivative, which is a reaction product of a 1,2-dicarbonyl derivatives that is an intermediate of advanced glycated protein in vivo, an immunogen that induces said antibody, and a method for immunological assay of a 1,2-dicarbonyl derivative using said antibody.
BACKGROUND OF THE INVENTION
Diabetes is a metabolic disease attributed to insulin and produces various kinds of complications, including, for example, eye diseases, kidney diseases, neurological disorders, cardiovascular complications, and gangrenes, since it is liable to become chronic. Prognosis of the patients is greatly influenced by the presence or absence and the degree of complications.
Deoxyglucosone is an intermediate produced in glycation of protein, which is suspected to be the potent cause of diabetes, and serves as a cross-linker between proteins to enhance production of an advanced glycated end proteins. This substance has attracted attention as an index for assessment of symptoms of diabetic complications from the facts that its blood level increased in patients with diabetes who are likely to maintain hyperglycemic conditions, especially patients with diabetes complicated with kidney diseases (Biochem. Biophys. Res. Commun. 196, 837-843, 1993) and patients with diabetic arteriosclerosis (DIABETES, 45, SP3, S81-83, 1996). It has also been suggested that the methylglyoxal blood level increased in patients with diabetic arteriosclerosis (DIABETES, 45, SP3, S81-83, 1996). Accordingly, attention has been paid to behavior of 1,2-dicarbonyl derivatives in cases of diabetes.
However, since 1,2-dicarbonyl derivatives, represented by deoxyglucosone, are very reactive, the compounds have been so far determined by converting them to derivatives or stable forms, extracting the conversion products, and measuring the products by chromatographic analysis such as GS/MS, HPLC, or the like. In such a method, operation of extracting 1,2-dicarbonyl derivatives and procedures for chromatographic analysis are complicated and time-consuming. Therefore, it has been desired to develop a simple method for measuring 1,2-dicarbonyl derivatives in multiple specimens.
SUMMARY OF THE INVENTION
An object of the present invention is to provide a simple method for measuring 1,2-dicarbonyl derivatives in multiple specimens. Another object of the present invention is to provide antibodies to be used in the above method, and immunogens that induce the production of the antibodies.
As a result of intensive studies, the present inventors found that a 1,2-dicarbonyl derivative in a specimen could be measured by reacting the 1,2-dicarbonyl derivative with a diamino derivative to convert it to a stable pyrazine derivative and measuring the pyrazine derivative by an immunological assay using an antibody that recognizes the pyrazine derivative. The present inventors succeeded to provide the above antibody and, thus, the present invention was completed.
The present invention provides an antibody that recognizes at least a region comprising R
1
, R
2
, carbon atoms in the pyrazine ring to which R
1
and R
2
are respectively attached, and nitrogen atoms in the pyrazine ring attached to said carbon atoms, of a pyrazine derivative represented by the formula (I):
wherein R
1
and R
2
independently represent a hydrogen atom, a methyl group, a hydroxymethyl group, a hydroxyethyl group, a dihydroxyethyl group, a dihydroxypropyl group, a trihydroxypropyl group, or a trihydroxybutyl group, A represents a group that binds to the pyrazine ring to form a 6-membered aromatic hydrocarbon group, a 5- or 6-membered aromatic heterocyclic ring group, or a 5- or 6-membered alicyclic hydrocarbon group, and R
3
represents a linking residue, where said 5-membered ring formed by A may have 1 or 2 substituents and said 6-membered ring may have 1 to 3 substituents, in addition to R
3
.
The present invention also provides an immunogen comprising the pyrazine derivative represented by the formula (I) or the pyrazine derivative to which a carrier is bound via R
3
.
The present invention further provides a method for measuring 1,2-dicarbonyl derivative, which is an intermediate of glycation in vivo, by an immunological assay, comprising the steps of:
reacting a 1,2-dicarbonyl derivative in a specimen with a diamino derivative represented by the formula (II):
wherein R
3
and A are as defined in the formula (I), to produce a pyrazine derivative represented by the formula (I); and
measuring the pyrazine derivative by an immunological assay utilizing the antigen-antibody reaction between the pyrazine derivative and the above antibody according to the present invention to thereby measure the 1,2-dicarbonyl derivative in the specimen.
Furthermore, the present invention provides a kit for immunological determination of 1,2-dicarbonyl derivative, comprising the antibody as described above and the diamino derivative represented by the formula(II).
REFERENCES:
Kevin J. Knecht et al.; “Detection of 3-Deoxyfructose and 3-Deoxyglucosone in Human Urine and Plasma: Evidence for Intermediate Stages of the Maillard Reaction in Vivo1”; Archives of Biochemistry and Biophysics; vol. 294, No. 1, 1992, pp.130-137.
Toshimitsu Niwa et al.;“Presence of 3-Deoxyglucosone, A Potent Protein Crosslinking Intermediate of Maillard Reaction, In Diabetic Serum”; Biochemical and Biophysical Research Communications; vol. 196, No. 2, 1993, pp. 837-843.
Hiroyuki Yamada et al.; “Increase in 3-Deoxyglucosone Levels in Diabetic Rat Plasma”; The Journal of Biological Chemistry; vol. 269, No. 32, 1994, pp. 20275-20280.
Ei Fujji et al.; “Quantitation of the Glycation Intermediate 3-Deoxyglucosone By Oxidation with Rabbit Liver Oxoaldehyde Dehydrogenase to 2-Keto-3-Deoxygluconic Acid Followed By High-Performance Liquid Chromatography”; Journal of Chromatography B; vol. 660, 1994, pp. 265-270.
Toshimitsu Niwa et al.; “Elevated Serum Levels of 3-Deoxyglucosone, a Potent Protein-Cross-Linking Intermediate of the Maillard Reaction, in Uremic Patients”; NEPHRON; vol. 69, 1995, pp. 438-443.
Sundeep Lal et al.; “Quantitation of 3-Deoxyglucosone Levels in Human Plasma”; Archives of Biochemistry and Biophysics; vol. 342, No. 2, 1997, pp. 254-260.
Naoyuki Taniguchi et al.; “Involvement of Glycation and Oxidative Stress in Diabetic Macroangiopathy”; Diabetes; vol. 45, Sp3, 1996, s81-83.
Chemical Abstracts, vol. 118, 1993, Reference 231710.
Ito Satoru
Kurano Yoshihiro
Uchida Yoshiaki
Banner & Witcoff Ltd
Ceperley Mary E.
The Fujirebio Inc.
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