Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Reexamination Certificate
2000-12-21
2003-11-11
Chin, Christopher L. (Department: 1641)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
C435S007100, C435S007230, C435S007400, C435S007940, C435S007950, C435S023000, C435S040520, C435S226000, C435S332000, C435S338000, C435S960000, C436S503000, C436S518000, C436S548000, C436S063000, C436S164000, C436S811000, C530S388200, C530S388260, C530S391300
Reexamination Certificate
active
06645734
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to monoclonal antibodies binding specifically to a certain serine protease, i.e., neurosin. It also relates to a production process thereof and a method for diagnosing various diseases using the monoclonal antibodies.
2. Disclosure of the Prior Art
In general, proteases are biosynthesized as inactive precursors. They undergo limited hydrolysis in molecules to convert into activated type proteases. In so far as enzymes are proteases, they have an activity for hydrolyzing a peptide bond. However, their action modes are varied according to kinds of proteases. According to particular kinds of catalytic sites, proteases are divided into serine proteases, cysteine proteases, aspartate proteases, metal proteases and the like. Proteases of each kind have a variety of properties such as, from that having general digestive properties to that having various regulatory domains and strict substrate specificity, thereby specifically hydrolyzing only characteristic proteins.
The optimal pH range of serine proteases is neutral to weak alkaline and, in general, many of them have a molecular weight of about 30,000 or lower. All proteases of blood coagulation, fibrinolysis and complement systems which have a large molecular weight belong to trypsin-like serine proteases. They have many regulator domains and form a protease cascade which is very important to reactions in a living body.
According to their primary structure, serine proteases can be divided into subtilisin family and chymotrypsin family. Those of subtilisin family are produced by only
Bacillus subtilis,
while those of chymotrypsin family are widespread in microorganisms, animals and plants. His-57, Asp-102 and Ser-195 (chymotrypsin Nos.) are concerned in their catalytic activity and, in general, they are inactivated by diisopropyl fluorophosphate (DFP). For exhibiting their activity, a catalytic triad is required, in which His influenced by Asp deprives Ser-195 of its proton to activate Ser. Further, it binds to a substrate to cause polarization of the carbonyl group, and the oxygen atom forms oxy anion. Trypsin has Asn-189 at this site and interacts with the positive charge of a basic amino acid such as Lys, Arg or the like. On the other hand, the corresponding site of chymotrypsin is Ser-189, and an aromatic amino acid such as Tyr, Phe, Trp or the like, or Leu or Met can also bind thereto. In analogy with chymotrypsin, esterase has Ser-189 and interacts with a non-aromatic amino acid such as Ala or the like. As other enzymes belonging to chymotrypsin family, there are Achromobacter protease, plasmin, medullasin, acrosin, V8protease, cathepsin G, chymase, proline specific endopeptidase, submaxillary gland protease A, XIIa, XIa, plasma kallikrein, IXa, Xa, &agr;-thrombin, VIIa, protein C, tissue plasminogen activator, urokinase, C1r, C1s, C2, B, D, I, &ggr;-seminoprotein, tissue kallikrein, C and B factors of
Limulus polyphemus
blood cells, blood coagulation enzymes and the like.
Recently, cDNA and amino acid sequences of many novel proteases have been determined by PCR using oligonucleotide primers for consensus sequences of serine proteases. According to this method, novel proteases have been found by various researchers such as Yamamura et al. (Yamanura, Y et al., Biochem. Biophys. Res. Commun., 239, 386, 1997), Gschwend, et al. (Gschwend, T. P. et al., Mol. Cell. Neurosci., 9. 207, 1997), Chen et al. (Chen, Z-L, et al., J. Neurosci., 15, 5088, 1995) and others.
SEQ ID NO: 3 of JP 9-149790 A discloses neurosin as a novel serine protease. Neurosin has also been reported in Biochimica et Byophysica Acta, 1350, 11-14, 1997. Neurosin having not more than 30% identity to known serine proteases has been obtained as a result of recognition of serine protease activity in a culture supernatant of human colon cancer COLO201 cells and isolation of a cDNA encoding neurosin. By this, there is provided a method for mass production of neurosin using the serine protease gene and a method for screening specific inhibitors using the enzyme. In addition, the screening method has been shown to be useful for screening medicines for treating various diseases.
At present, functions of neurosin are still unknown. However, since neurosin is abundantly expressed in brain, it is presumed to play an important role in maintenance of brain functions. Further, there is a possibility that further detailed functions can be elucidated by using recombinant protein.
JP 6-62855 A discloses a novel serine protease, Zyme, and this is also reported by J. Biol. Chem., 272(40), 25135-2514, 1997. The cDNA and amino acid sequences of Zyme have been determined by PCR amplification of consensus sequences having chymotrypsin-like activity to construct a cDNA library by using the brain mRNA of a patient with Alzheimer's disease. The mRNA encoding Zyme is recognized in several mammals. Further, although Zyme is expressed abundantly in brain, kidney and salivary glands, as to brain, it is not expressed in fetal brain, but is expressed only in adult brain. Further, Zyme has a gene in chromosome 19q13.3, and this region has been revealed to be a part linking to late onset familial Alzheimer's disease. Then, it is considered that Zyme would be useful for elucidating characteristics of neural diseases such as Alzheimer's disease and Down's syndromes.
WO 98/11238 discloses a novel protease, Protease M, and this is also reported in Molecular Medicine, 2(5), 624-636, 1996. Protease M cDNA is obtained from normal human mammary epithelial 76 N cell line and has a sequence very similar to kallikrein, Prostate-Specific Antigen (PSA) and trypsin. And, Protease M gene is present in chromosome 19q13.3. Protease M is considered to be a marker useful for primary breast adenocarcinoma and primary ovary cancer because, while it is downregulated in metastatic breast cancer cell line, its mRNA is strongly expressed in primary breast cancer cell line, ovary cancer tissue and cancer cell line.
Although origins of these neurosin, Zyme and Protease M are different, their cDNA sequences and amino acid sequences, and further the positions in chromosome conformation of the genes encoding them are completely identical to one another. Since there is a high possibility that these substances would be the same substance, the name “neurosin” is used herein to refer to them altogether. As described above, this serine protease has been found by different groups of researchers almost at the same time, and its pharmacological activities have been studied. Then, it is expected that its importance will be elucidated on various occasions in the future. For example, Games et al. (Games, D. et al., Nature, 373, 523, 1995) and Hsiao et al. (Hsiao, K, et al., Science, 274, 99, 1996) succeeded in expression of a large amount of &bgr;-amyloid precursor protein (&bgr;APP), and production of transgenic mice in which deposition of amyloid &bgr; protein (A&bgr;) was observed in 1995 and 1996, respectively. Then, the role of the above serine protease in Alzheimer's disease and the like will be further elucidated in the future.
As disclosed in JP 6-62855 A, Zyme (i.e., neurosin) plays an important role in Alzheimer's disease and Down's syndromes. While it has been proposed that Alzheimer's disease should be divided into that presenile Alzheimer's disease and Alzheimer type senile dementia manifesting in senescence from the pathological viewpoint, the term “Alzheimer's disease” is used herein to refer to them altogether.
Clinically, Alzheimer's disease is characterized by progressive decline of various recognition functions and the main neuropathological observation is to find abnormal structures such as senile plaque and neurofibril change in addition to nerve cell degeneration and deficiency (Trojanowski, J. Q. et al., In Current Neruology, 16, 93, 1996). Among them, while senile plaque also appears in case of normal aging, it appears much more frequently in case of Alzheimer's dise
Kominami Katsuya
Mitsui Shinichi
Okui Akira
Yamaguchi Nozomi
Chin Christopher L.
Fuso Pharmaceutical Industries Ltd.
Grun James L.
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