Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Peptide containing doai
Reexamination Certificate
2000-02-15
2001-09-11
Carlson, Karen Cochrane (Department: 1653)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Peptide containing doai
C514S008100, C514S023000, C514S053000, C530S322000, C536S007100, C536S007200, C536S013800
Reexamination Certificate
active
06288028
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to new application forms of synthetic BPC (Body Protection Compound) peptides comprising 8 to 15 amino acid residues with a molecular weight of 900 to 1,600 daltons, which have organo-protective activity, to processes for their preparation and their use in diagnosis and therapy.
2. Description of the Related Art
Proteins and peptides which are useful for the treatment of various diseases in humans and animals are known. Many of these agents are produced in vivo and may be extracted from animals or humans to prepare pharmaceutical compositions. Examples for pharmaceutically useful proteins or peptides are insulin, erythropoietin, BMPs, interferons, etc. Another example is a gastric juice protein with mucosal protective activity which was recently isolated and named BPC. WO 92/04368 relates to BPC which exhibits body-protective activity and has a molecular weight of about 40,000 daltons, its preparation and its use. WO 93/24521 and WO 94/11394 disclose BPC peptides which have organo-protective activity of the same type as known from the parent protein BPC. Sikiric et al. in:
Digestive Diseases and Sciences
, 41(1996)7, 1518-1526, describe pentadecapeptide BPC 157, which exhibits when dissolved in water and saline salutary and prophylactic effects on acute pancreatitis and concomitant gastroduodenal lesions in rats.
Thus, the BPC peptides are known for a wide variety of pharmaceutical applications. However, the physicochemical stability of these peptides, for instance in normal saline, is not satisfactory. Furthermore, the application of BPC peptides, in particular by injection of an aqueous solution or in normal saline, causes pain and/or necrosis.
Salts of proteins and peptides are well known in the art. It is for instance known from Bertrand, M. et al. in:
Journal of Peptide Research
, 49 (1997)3, 269-272, that the effects of particular salts are very selective in respect to the stability and structure of peptides. For instance, the addition of monovalent cations, such as NH
4
+
to a final 0.1 M peptide (poly (Glu-Leu)) solution, induces a transition to a water soluable beta-structure. In contrast, no transition was observed using Li
+
, Na
+
, or Cs
+
ions.
SUMMARY OF THE INVENTION
The role of surface-accessable ion pairs in protein stability was investigated by determining the effects of added salts (KCl, MgCl
2
and LaCl) at neutral and acidic pH upon the stability of de novo designed two-stranded alpha-helical coiled-coils. The results show that added salt may have complex effects on protein stability, involving stabilizing and destabilizing contributions, whereby the net effect depends upon the nature of the charged residues and ionic interactions present in the protein (Kohn et al. in:
Journal of Molecular Biology
, 267 (1997)4,1039-1052)
Previous studies of model peptides have also shown that salt bridges spaced at i,i +4 along the peptide chain are more stabilizing than those spaced at i,i +3, with a preference for the order acid-base rather than base-acid from the N- to the C- terminus. However, at present it is not known whether surface salt bridges have a strong stabilizing effect on the native structure in proteins (Berger et al. in:
Journal of Biomolecular Structure and Dynamics
, 14 (1996)3, 285-291).
Thus, the properties of peptide salts with respect to their stability, structure and function depend very much upon the specific ions involved in forming the specific salt and other intrinsic or extrinsic factors. At present, it cannot be foreseen which particular properties a specific salt of a specific peptide might have.
DESCRIPTION OF PREFERRED EMBODIMENT
The technical problem underlying the present invention is to provide BPC peptides in more stable form and a diagnostic and/or pharmaceutical composition comprising BPC peptides exhibiting improved stability and at least the same pharmaceutical activities as the BPC peptides themselves. Furthermore, the technical problem underlying the present invention is to provide a pharmaceutical composition which overcomes the drawbacks mentioned above, in particular, allowing painless injection of BPC peptides. The present invention solves these problems by providing BPC peptide salts and a pharmaceutical or diagnostic composition comprising a pharmaceutically or diagnostically effective amount of the salts of BPC peptides, wherein the anion of the salt is a negatively charged peptide comprising 8 to 15 amino acids, exhibiting a molecular weight of 900 to 1,600 daltons and having the general formula (I)
[Zaa Pro Pro Pro Xaa Yaa Pro Ala]
(−) or (2−)
(I)
wherein Xaa is a neutral aliphatic amino acid residue, in particular Ala, bAla, Leu, lie, Gly, Val, Nle or Nva,
Yaa is a basic amino acid residue, in particular Lys, Arg, Orn or His and
Zaa is an acidic amino acid residue, in particular Glu, Asp, Aad or Apm
and wherein the cation of the salt is the cation of an inorganic or organic nontoxic and pharmaceutically acceptable base. In particular, the cation of the salt is an alkali metal or an alkaline earth metal, for instance Na
+
, K
+
, Li
+
, Cs
+
, Ca
+
, or another metal, such as Zn
2+
, or a primary, secondary or tertiary amine or organic compound, such as NH
4
+
, triethanolamine
+
, cyclohexylamine
+
, 2-AMP
+
(2-amino-1-propanol) or TRIS
+
(Tris-(hydroxymethyl)-aminomethan), as long as these cations are physiologically acceptable.
Surprisingly, the BPC peptide salts of the present invention show at least the same pharmaceutical activity as the BPC peptides and, additionally, a remarkably increased physicochemical stability in comparison with the free BPC peptides or acetates of BPC peptides. The cation used according to the present invention does not influence the activity of the BPC peptides, but increases their stability. The BPC peptide salts of the present invention are, for instance, more stable than BPC peptides or BPC peptide acetates in normal saline or water. Furthermore, the salts of the present invention are well suited for peroral use and do not exhibit any undesired side effects such as pain or necrosis during or after application, in particular, by injection. The BPC peptide salts of the present invention therefore allow an improved enteral and parenteral application. Furthermore, the salts of the present invention are very favorable due to the absence of any signs of toxicity up to doses of 50 mg/kg b.w., (body weight).
The salts of the invention can be obtained by dissolving the free BPC peptide in an aqueous or aqueous/alcoholic solvent or in other suitable solvents with an appropriate base and then isolating the obtained salt of the invention by evaporating the solution, by freezing and lyophilization or by addition of another solvent, e.g. diethylether, to the aqueous and/or alcoholic solution of the BPC peptide salt inducing the separation of unsoluble crude salt. For salt formation, usually one or maximal two mols of base, i.e. cation, and one mol of the free BPC peptide are used. For the preparation of alkali BPC peptide salts, alkali metal carbonates or hydrogencarbonates are preferably used. The prepared peptide salts are freely soluble in water. Thus, the present invention also relates to a process for the preparation of the BPC peptide salts.
In the context of the present invention, a base is considered as a substance capable of forming a cation in a solution, particularly in an aqueous and aqueous/alcoholic solution.
In the context of the present invention, pharmaceutical activity encompasses prophylactic and therapeutic activities. Accordingly, a pharmaceutical composition refers to compositions exhibiting prophylactic and/or therapeutic activities.
Additionally, the invention relates to a pharmaceutical or diagnostic composition comprising the BPC peptide salts of the present invention, optionally in conjunction with one or more pharmaceutically acceptable carriers, as well as proc
Duvnjak Marko
Grabarevic Zeljko
Mise Stjepan
Petek Marijan
Rotkvic Ivo
Carlson Karen Cochrane
Ostrolenk Faber Gerb & Soffen, LLP
Robinson Patricia
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