Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Peptide containing doai
Reexamination Certificate
1997-07-17
2002-11-19
Prouty, Rebecca E. (Department: 1652)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Peptide containing doai
C530S300000, C530S324000, C536S023100, C536S023500, C435S069200, C435S255200, C435S320100
Reexamination Certificate
active
06482798
ABSTRACT:
The present invention relates to aprotinin variants having improved enzyme-inhibitory, immunological and pharmacokinetic properties and their preparation.
Aprotinin, which is also called bovine pancreatic trypsin inhibitor (BPTI), belongs to the family of serine protease inhibitors of the Kunitz type. The spectrum of the inhibitable serine proteases includes, for example, trypsin, chymotrypsin, plasmin and plasma kallikrein (W. Gebhard, H. Tschesche and H. Fritz, Proteinase Inhibitors, Barrett and Salvesen (eds.), Elsevier Science Publ. BV 375-387, 1986).
Aprotinin consists of 58 amino acids. The three-dimensional structure of the protein was elucidated with the aid of X-ray structural analysis and NMR spectroscopy (Wlodawer et al., J. Mol. Biol. 198 (3), 469-480, 1987; Wagner et al., J. Mol. Biol. 196 (1), 227-231, 1987; Berndt et al., Biochemistry 32 (17), 4564-4570, 1993).
Under the trade name Trasylol®, natural aprotinin was originally employed for the treatment of pancreatitis. Trasylol® is today used in cardiac surgery, since clinical studies have shown that treatment with aprotinin significantly decreases the need for transfusion in operations of this type and leads to the reduction of post-operative bleeding (D. Royston, J. Cardiothorac. Vasc. Anesth. 6; 76-100, 1992).
It was possible to show that the replacement of the amino acid in position 15 defining the inhibitory specificity leads to useful aprotinin variants having improved inhibitory properties (German Patent Specification 3 339 693). Depending on the amino acid introduced, in this way potent inhibitors can be produced which, for example, inhibit the elastase from pancreas or from leucocytes.
It was further possible to show that the inhibitory properties of aprotinin and of the variants produced by replacement in position 15 is also determined by further amino acid residues in the contact region between the target protease to be inhibited and the inhibitor molecule. These especially include the additional amino acid residues in positions 14, 16, 17, 18, 19, 34, 38 and 39. Aprotinin variants having improved properties due to replacement of one or more of these amino acid residues in the area of the contact region were described, inter alia, in the following patent applications by way of example: WO 89/01968, WO 89/10374, EP 0 307 592, EP 683 229.
Interestingly, it was possible to improve the pharmacokinetic properties of aprotinin and its variants by amino acid replacements which define the physicochemical properties of the substance. It was thus possible, by lowering the positive net charge of the molecule, significantly to lower the renal binding. Variants of this type were described in the Patent Application WO 92/06111.
For reasons of better industrial preparability, it is convenient in certain cases to carry out a modification to the N-terminal end of the inhibitor molecule. Such modifications can be N-terminal contractions or extensions or deletions of one or more amino acids. N-terminally modified aprotinin variants were described in EP 419 878.
Aprotinin Variants According to the Invention
The aprotinin variants described in the present patent application are distinguished by the following features:
1. Replacement of one or more amino acids in the active centre of the molecule to improve the activity properties.
2. Replacement of amino acids to decrease the positive net charge with the aim of improving the immunological and pharmacokinetic properties.
3. Modification of the N-terminal amino acid sequence for reasons of industrial preparability.
REFERENCES:
patent: 4595674 (1986-06-01), Tschesche et al.
patent: 5118668 (1992-01-01), Auerswald et al.
patent: 5164482 (1992-11-01), Ebbers et al.
patent: 5231010 (1993-07-01), Ebbers et al.
patent: 5591603 (1997-01-01), Bjørn et al.
patent: 5621074 (1997-04-01), Bjørn et al.
patent: 132732 (1985-02-01), None
patent: 339942 (1989-11-01), None
patent: 419878 (1991-04-01), None
patent: 89/02463 (1989-03-01), None
patent: 90/10075 (1990-09-01), None
patent: 92/0611 (1992-04-01), None
patent: 92/06111 (1992-04-01), None
Deng, et al., Anal. Biochem., 200, pp. 81-88, “Site-Directed Mutagenesis of Virtually Any Plasmid by Eliminating a Unique Site”,(1992).
J.G. Bieth, Biochemical Medicine, 32, pp. 387-397, “In Vivo Significance of Kinetic Constants of Protein Proteinase Inhibitors”, (1984).
U.K. Laemmli, Nature 227, pp. 680-685, “Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4”, (1970).
C.R. Merril, M.L. Dunau, D. Goldmann, Anal. Biochem. 110, pp. 201-207,“ A Rapid Sensitive Silver Stain for polypeptides in Polyacrylamide Gels”, (1981).
J. Kurjan and Herskowitz, Cell, 30, pp. 933-943, “Structure of a Yeast Pheromone Gene (Mf&agr;)-Factor Precursor Contains Four Tandem Copies of Mature &agr;-Factor”, (1982).
Yarger, et al., Mol. Cell. Biol., 6, pp. 1095-1101, “Transcription Terminator-Like Element within aSaccharomyces cerevisiaePromotor Region”, (1986).
W. Gebbhard, H. Tschesche and H. Fritz, Proteinase Imhibitors, Barrett and Salvesen (eds.), Elsevier Science Publ. BV 375-387, “Biochemistry of aprotinin and aprotinin-like inhibitors”, (1986).
Wlodawer, et al., J. Mol. Biol. 198, (1), pp. 227-231, “Structure of Form III Crystals of Bovine Pancreatic Trypsin Inhibitor”, (1987).
Berndt, et al., Biochemistry 32 (17), pp. 4564-4570, “Designed Replacement of an Internal Hydration Water Molecule in BPTI: Structural and Functional Implications of a Glycine-to-Serine Mutation”, (1993).
Wagner, et al., J. Mol. Biol. 196, (1), pp. 227-231, “Reinvestigation of the Aromatic Side-chains in the Basic Pancreatic Trypsin Inhibitor by Heteronuclear Two-dimensional Nuclear Magnetic Resonance”, (1987).
D. Royston, J. Cardithorac, Vasc. Anesth. 6; pp. 76-100, “High-Dose Aprotinin Therapy: A Review o the First Five Years' Experience”, (1992).
Björn Sören
Christensen Niels Dyhr
Diness Viggo
Norris Kjeld
Nørkskov-Lauritsen Leif
Bayer Aktiengesellschaft
Briscoe Kurt G.
Norris McLaughlin & Marcus P.A.
Prouty Rebecca E.
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