# Patent

## Title
Therapeutic fibrin-derived peptides and uses thereof

## Abstract

The invention relates to peptides having the general formula (I), or a salt or amide thereof, wherein R1 and R2 are either the same or different, wherein R1 and R2 are each selected from the group consisting of hydrogen and a saturated or unsaturated hydrocarbon residue, said residue having from 1 to 10 carbon atoms, wherein Z1 is selected from the group consisting of histidine and proline, wherein Z2 is selected from the group consisting of an arginine and a peptide comprising an initial arginine and having from 2 to 30 amino acids. The invention also relates to methods using the peptides of the present invention in the treatment of inflammation.

## Background
CROSS REFERENCE TO RELATED APPLICATIONS
The present application is a continuation of U.S. patent application Ser. No. 10/459,030, filed Jun. 11, 2003, now U.S. Pat. No. 7,271,144, which is a continuation of International Patent Application No. PCT/AT01/00387, filed Dec. 7, 2001, published in German on Jun. 20, 2002 as International Patent Publication No. WO02/248180, which claims priority to Austrian Application No. AT A 2063/2000, filed Dec. 12, 2000, all of which are incorporated in their entireties herein.


BACKGROUND
The invention concerns peptides and/or proteins, their use for preparing a therapeutic and/or preventive pharmaceutical composition as well as a pharmaceutical composition.
Substances for the inhibition or prevention of inflammatory reactions, so-called immunosuppressants, which so far have been used for prophylaxis and therapy, generally comprise two distinct groups. Firstly, derivatives of a hormone, i.e. cortisone, naturally occurring in the body, and secondly, exogenous immunosuppressants such as cyclosporin and its derivatives, azathioprine, cyclophosphamide etc. All those substances possess anti-inflammatory effects but they show substantial side reactions in long-term therapy. Those side reactions have a limiting effect on long-term therapy, which is why those substances are used alternately or in combination in order to keep side effects on a tolerable level or in order to be able to actually proceed with the therapy. As examples of side reactions, the pathological fractures associated with cortisone are to be mentioned, which fractures are caused by the osteoporotic effect of the cortisone, or the renal failure which may be caused by cyclosporin. Those side reactions are inevitable with both groups of compounds, and hence it is merely a question of the duration of the therapy and of the total dose at what point the therapy must be stopped.
SUMMARY OF THE INVENTION
The present invention has as its object to provide new pharmaceutical products which are suitable for preventing or inhibiting inflammatory effects and which only show minor side effects. A further object consists in providing long-term therapy.
In the following, the amino acids of the peptides according to the invention are referred to by the usual abbreviations, which denote the α-amino acids.
By “analogues,” a peptide is understood which, by derivatisation, substitution, preferably homologous substitution, deletion and/or insertion, is derived from the sequence of the fibrin and in particular from the preferred sequences.
The peptides or protein according to the invention exhibit the general formula I




wherein R1 and R2, being equal or different, denote hydrogen, a saturated or unsaturated hydrocarbon residue comprising from 1 to 3, in particular up to 10, carbon atoms,

    
    
        Z1 denotes a histidine or proline residue,
        Z2 denotes an arginine residue, a peptide residue or a protein residue comprising an initial arginine residue, in particular comprising from 2 to 30 amino acids, as well as the salts thereof, and, f.i., also amides, or mixtures with each other and /or with at least one further substance for therapeutic and/or preventive use in human and/or veterinary medicine, whereby in particular only L-amino acids are provided. Sequences of formula I are listed in Table 1.
    
    


It was completely surprising that the specified amino acid sequence prevents the adhesion of cells from the bloodstream to endothelial cells of the vascular wall and/or their subsequent transmigration from the blood into the tissue.
The peptides or protein according to the invention exhibit the general formula II




wherein R1 and R2, being equal or different, denote hydrogen, a saturated or unsaturated hydrocarbon residue comprising from 1 to 3, in particular up to 10, carbon atoms,

    
    
        Z1 denotes a histidine or proline residue,
        Arg denotes an arginine,
        Z3 denotes a proline or valine residue,
        Z4 denotes a leucine or valine residue,
        Z5 denotes a protein residue or a peptide residue, in particular comprising from 2 to 30 amino acids, or an alcohol comprising from 1 to 3, in particular up to 10, carbon atoms, or an organic or inorganic base residue, as well as the salts thereof, and, f.i., also amides, or mixtures with each other and/or with at least one further substance for therapeutic and/or preventive use in human and/or veterinary medicine, whereby in particular only L-amino acids are provided. Sequences of formula II are listed in Table 2.
    
    


It was completely surprising that parts of the sequence, peptides or fragments of the fibrinogen exhibit anti-inflammatory effects. Without being bound by such theoretical considerations, said effects might be based on the fact that the fibrin binds to endothelial cells via its neo-N-terminus of the Bbeta-chain and to cells in the bloodstream via the sequence of the Aalpha-chain, thereby leading to the adhesion and transmigration of cells into the tissue. Those bindings exhibit a side reaction in that the formation of fibrin is inhibited. However, said inhibition does not constitute a potential disadvantage to the patient since the blood coagulation is sufficient also in the absence of fibrin if slight injuries occur. Only in case of surgical treatment, it might optionally be suitable to stop such kind of therapy. Other side reactions may substantially be ruled out, since those substances only interact with natural ligands. Furthermore, the natural defense is not affected adversely by the leukocytes in the blood. Thus, the composition of the same, such as granulocytes, lymphocytes and monocytes, remains unaffected so that the natural defense process is maintained and the defense against infections in the blood remains unchanged.


DETAILED DESCRIPTION
Fibrinogen is produced in the liver and, in this form, is biologically inactive and normally is provided in the blood at concentrations of around 3 g/l. By proteolytic cleavage of the proenzyme prothrombin, thrombin is formed which cleaves off the fibrinopeptides A and B from the fibrinogen. In doing so, fibrinogen is transformed into its biologically active form. Fibrin and fibrin cleavage products are generated.
Thrombin is formed during each activation of the blood coagulation, i.e. with each damage to the tissue, be it of inflammatory, traumatic or degenerative genesis. The formation of fibrin as mediated by thrombin is basically a protective process with the purpose of quickly sealing any defects caused to the vascular system. However, the formation of fibrin is also a pathogenic process. The appearance of a fibrin thrombus as the triggering cause of cardiac infarction is one of the most prominent problems in human medicine.
The role which fibrin plays during the extravastation of inflammatory cells from the bloodstream into the tissue, which, on the one hand, is a desired process of the defense against pathogenic microorganisms or tumour cells occurring in the tissue, but, on the other hand, is a process which, by itself, induces or prolongues damage done to the tissue, has so far not been examined at all or not to a sufficient extent. Fibrin binds to endothelial cells via its neo-N-terminus of Bbeta by means of the sequence to Bbeta and to cells in the bloodstream by means of the sequence Aalpha, thereby leading to the adhesion and transmigration of cells into the tissue.
The peptides or proteins according to the invention may prevent the adhesion of cells from the bloodstream to endothelial cells of the vascular wall and/or their subsequent transmigration from the blood into the tissue.
A peptide or protein according to the invention of the general formula II, wherein Z5 denotes a peptide residue comprising the following amino acid sequence (SEQ ID NO 291):








Asp Lys Lys Arg Glu Glu Ala Pro Ser Leu Arg Pro



 


Ala Pro Pro Pro Ile Ser Gly Gly Gly Tyr Arg






and

    
    
        Z1 denotes a histidine residue,
        Arg denotes an arginine residue,
        Z3 denotes a proline residue,
        Z4 denotes a leucine residue,

prevents fibrin fragments from depositing on or adhering to the vascular wall. Thus, it is rendered impossible that inflammatory cells are retained at the endothelial cells of the vascular walls of arteries and veins, and such cells are prevented from remaining at the vascular walls, thus being prevented from infiltrating the tissue any further.

    
    


A peptide or protein of the general formula II, wherein Z5 denotes a peptide residue comprising the following amino acid sequence (SEQ ID NO 292):








Glu Arg His Gln Ser Ala Cys Lys Asp Ser Asp Trp



 


Pro Phe Cys Ser Asp Glu Asp Trp Asn Tyr Lys






and

    
    
        Z1 denotes a proline residue,
        Arg denotes an arginine residue,
        Z3 denotes a valine residue,
        Z4 denotes a valine residue,

has the effect of preventing the cells of the peripheral blood from adhering to fibrin or fibrin fragments, hence prohibiting their migration in the tissue.

    
    


The described cleavage products are also known in the literature as peptide Bbeta and peptide Aalpha. Said above mentioned proadhesive and promigratory path is a completely new one for the system of controlling the migration of cells from the blood into the tissue. This function of the fibrin may be blocked by peptide Bbeta and also by peptide Aalpha.
Therefore, said peptides according to the invention are suitable as therapeutic agents for humans and animals in order to block the migration of cells from the blood into the tissue. Since fibrin or other fibrinogen products produced by proteolytic cleavage, such as, f.i., fibrinogen cleaved by an urokinase-plasminogen-activator, are generated only to a specific and regionally limited extent, i.e. at sites of inflammation, disturbed coagulation, arterial sclerosis, thrombosis and/or tumour growth, the effect of said therapeutic agent is regionally limited, which means that pathological side effects occurring in other places are not to be expected or only to a limited extent.
Preferable and completely unexpected fields of application for the peptides and/or proteins according to the invention consist in the preparation of pharmaceutical compositions for the therapy or prevention of local and/or generalized inflammations in the body in case of infectious genesis, based upon an auto-immune reaction, based upon a rheumatic disease, based upon a disorder in the immune system, based upon a genetic disease, for the prevention and/or therapy of the rejection occurring after organ transplants, of arterial sclerosis, of a reperfusion trauma, based upon arteriosclerotic and/or thrombotic diseases and an increased fibrin deposition. Such a peptide, in particular Bbeta, is also excellently suitable for the preparation of a pharmaceutical composition which accomplishes the transportation of a further drug substance to human or animal endothelial cells. In doing so, the drug substance to be transported is coupled to the peptide at one end and then, via VE-cadherin, deposits on a free spot of the vascular wall, i.e. on an endothelial cell.
In the following, the invention is explained in further detail by way of examples.
EXAMPLES
Example 1
Preparation of the Fibrinogen Cleavage Products
Non-polymerizing degradation products of fibrinogen were obtained by means of a decomposition involving cyanogen bromide according to Blombäck et al. (Nature 1968, 218; 130-134). The fibrinogen thus degraded largely consists of a 63 kD fragment, i.e. the N-terminal disulfide knot, NDSK, and comprises Aalpha-chain 1-51, Bbeta-chain 1-118 and gamma-chain 1-78. In order to obtain NDSK-II (NDSK minus fibrinopeptides A and B), the N-terminal amino acids of the Aalpha- and Bbeta-chains were cleaved off with thrombin (20 units/1 μg NDSK) in three hours at room temperature and subsequently were treated with diisopropylfluorophosphate in order to block thrombin activity. The NDSK-II thus obtained consisted of Aalpha-chain 17-51, Bbeta-chain 15-118 and gamma-chain 1-78.
In order to obtain NDSK-uPA, 500 μg of NDSK was treated with 200 units of urokinase-plasminogen-activator (uPA) of Messrs. Technoclone, Vienna, Austria, for one hour at 37° C. The reaction was stopped with 5 mM phenylmethylsulfonyl fluoride. The NDSK-uPA thus obtained is a NDSK and has no fibrinopeptide B.
As a negative control, a second fraction was obtained from the fibrinogen cleavage products referred to as FCB-2 according to Nieuwenhuizen et al. (Biochem Biophys Acta 1983, 755; 531-533), which cleavage products were produced by being treated with cyanogen bromide. FCB-2 is a protein having a size of 43 kD and consists of Aalpha-chain 148-208, Bbeta-chain 191-305 and gamma-chain 95-265. For control purposes, thrombin and diisopropylfluorophosphate were added to said protein. That, however, did not result in any change to the protein (in the following, referred to as FCB-2-thr).
For the purpose of further negative controls, culture medium (RPMI of Messrs. Life techn. Inc., Paisky, UK) was treated with thrombin as above and, subsequently, was inactivated (RPMI-thr) or was treated with uPA as above and was inactivated (RPMI-uPA).
Example 2
Peptide Aalpha (SEQ ID NO 293) corresponds to amino acids 1 to 28 of the alpha-chain of the fibrin and is identical to amino acids 17 to 45 of the Aalpha-chain of the fibrinogen:








Gly Pro Arg Val Val Glu Arg His Gln Ser Ala Cys



 


Lys Asp Ser Asp Trp Pro Phe Cys Ser Asp Glu Asp


 


Trp Asn Tyr Lys






Peptide Bbeta (SEQ IN NO 294) corresponds to amino acids 1 to 28 of the beta-chain of the fibrin, which is identical to amino acids 15 to 43 of the Bbeta-chain of the fibrinogen, which exhibits the following sequence:








Gly His Arg Pro Leu Asp Lys Lys Arg Glu Glu Ala



 


Pro Ser Leu Arg Pro Ala Pro Pro Pro Ile Ser Gly


 


Gly Gly Tyr Arg






By applying a fluorenylmethyloxycarbonyl (FMOC)-protective group strategy according to Carpino L. A. and Han. G Y, J. Amer. Chem. Soc. 1981; 37; 3404-3409, both peptides were synthesized by means of a solid-phase peptide synthesis according to Merrifield R. B., J. Amer. Chem. Soc. 1963; 85, 2149-2154, using a multiple peptide synthesizer. The crude peptides were purified by preparative reversed-phase HPLC via a Nucleosil 100-10, C18-column according to Engelhart H. and Müller H. Chromatography 1984 19:77 as well as Henschen A., Hupe K. P. and Lottspeich F. High Performance Liquid Chromatography VCH 1985. As control peptides, peptides of the same length but comprising a randomized amino acid sequence were used.
Example 3
HU-SCID Mouse-Model
Human skin was transplanted onto the backs of SCID mice, and two weeks later human lymphocytes were injected into the peritoneum. The proceedings were according to Petzelbauer et al. (J. Invest. Dermatol. 1996, 107; 576-581). Then, fifteen mice thus prepared were injected in their tail veins with the following:

    a) 100 μg of human NDSK-II
    b) 100 μg of human FCB-2
    c) 100 μg of peptide Aalpha
    d) 100 μg of peptide Bbeta
    e) 100 μg of randomized Aalpha
    f) 100 μg of randomized Bbeta

Twenty-four hours later, the human skin was removed and the number of inflammatory sites, expressed in cells per 0.3 mm2, was evaluated and the mean value was determined with a standard deviation.

    For a: 22+/−2.8
    for b: 9+/−2.1
    for c: 4+/−1.1
    for d: 6+/−1.1
    for e: 5+/−1.2
    for f: 7+/−1.3

That allows the conclusion that NDSK-II causes inflammations, and hence said protein was used as a pathogenic substance. The other compounds per se do not exhibit any significant increase in the amount of inflammatory cells.



Comparative Example 4
Fifteen mice according to Example 3 were injected in their tail veins with
100 μg of human NDSK-II and
100 μg of randomized peptide Aalpha.
Further proceedings were according to Example 3. Per 0.3 mm2, 23+/−3.5 inflammatory sites could be determined.
Comparative Example 5
Fifteen mice according to Example 3 were injected in their tail veins with
100 μg of human NDSK-II according to Example 1 and
100 μg of randomized peptide Bbeta.
Further proceedings were according to Example 3. Per 0.3 mm2, 24+/−2 inflammatory sites could be determined.
Example 6
Fifteen mice according to Example 3 were injected with
100 μg of human NDSK-II and
100 μg of synthesized peptide Aalpha.
Further proceedings were according to Example 3. Per 0.3 mm2, 21+/−2.2 inflammatory sites could be determined.
Example 7
Fifteen mice according to Example 3 were injected in their tail veins with
100 μg of human NDSK-II and
100 μg of synthesized peptide Bbeta.
Further proceedings were according to Example 3. Per 0.3 mm2, 14+/−2 inflammatory sites could be determined.
Examples 4 to 7 show that peptide Bbeta blocks lymphocytic inflammation.
Comparative Example 8
Endothelial cells from human umbilical veins (HUVEC) were marked with a red fluorescent dye (Cell Tracker Orange, 1 μl/ml, Molecular Probes, Eugene, Oreg.) and were dispersed on a collagen matrix (Collaborative Biomedical Products, Bedford, Mass.). Upon confluence of the endothelial cells, peripheral mononuclear blood cells (PBMC) (105 cells per 25 mm2) marked with a green fluorescent dye (Cell Tracker Green, 1 μ/ml, Molecular Probes of Messrs. Eugene, Oreg.) were superimposed. Thereafter, the cells were incubated at 37°C. for twelve hours.
Adhering cells that had transmigrated into the gel were photographed with a laser-scan microscope, were converted into pixels and were evaluated by means of an ‘NIH image” according to Gröger et al. (J. Immunol. Method 1999; 222: 101-109).
It was feasible to determine the number of adherent cells per 0.1 mm2 such as mentioned under “adhesion.” It was feasible to determine the number of migrated cells per 0.04 mm3 such as mentioned under “migration.” The mean value of three times three trials was evaluated together with the standard deviation.














adhesion
migration






















a) RPMI-uPA
0.1
μg/ml
40 +/− 4 
4 +/− 3




1.0
μg/ml
38 +/− 2 
5 +/− 2




10.0
μg/ml
32 +/− 4 
5 +/− 1



b) NDSK
0.1
μg/ml
31 +/− 18
6 +/− 3




1.0
μg/ml
35 +/− 18
5 +/− 2




10.0
μg/ml
36 +/− 24
6 +/− 3



c) NDSK-II
0.1
μg/ml
55 +/− 21
12 +/− 5 




1.0
μg/ml
67 +/− 31
19 +/− 12




10.0
μg/ml
65 +/− 31
19 +/− 10



d) NDSK-uPA
0.1
μg/ml
58 +/− 3 
10 +/− 2 




1.0
μg/ml
 60 +/− 3.5
14 +/− 3 




10.0
μg/ml
65 +/− 3 
 18 +/− 1.5



e) FCB2
0.1
μg/ml
30 +/− 26
6 +/− 4




1.0
μg/ml
10 +/− 10
3 +/− 2




10.0
μg/ml
21 +/− 7 
5 +/− 4



f) FCB-2-thr
0.1
μg/ml
20 +/− 12
6 +/− 5




1.0
μg/ml
23 +/− 13
7 +/− 5




10.0
μg/ml
26 +/− 11
4 +/− 2



g) RPMI-thr
0.1
μg/ml
29 +/− 15
4 +/− 5




1.0
μg/ml
26 +/− 14
5 +/− 5




10.0
μg/ml
41 +/− 20
5 +/− 4










That allows the conclusion that NDSK-II results in significant migrations of peripheral blood-monocellular cells (PBMC) to a greater extent than NDSK-uPA and hence exhibits pathogenic activity. None of the controls a), b), e), f) and g) resulted in any significant migration.

Example 9
100 μg of NDSK-II and Bbeta or Bbeta randomized were added to the collagen matrix according to Example 8 comprising the suspension of PBMC, and further proceedings were in accordance with Example 8.














adhesion
migration


















a) no addition of NDSK-II
38 +/− 15
 6 +/− 4


b) only 100 μg of NDSK-II
73 +/− 29
16 +/− 7


c) 10 μg of Bbeta + NDSK-II
63 +/− 33
 7 +/− 4


d) 100 μg of Bbeta + NDSK-II
47 +/− 34
 5 +/− 4


e) 1000 μg of Bbeta + NDSK-II
52 +/− 27
10 +/− 6


f) 10 μg of Bbeta randomized + NDSK-II
77 +/− 33
16 +/− 6


g) 100 μg of Bbeta randomized + NDSK-II
86 +/− 35
15 +/− 6


h) 1000 μg of Bbeta randomized + NDSK-II
78 +/− 31
13 +/− 8









As can be gathered from those test results, peptide Bbeta blocks inflammations.

Example 10
100 μg of NDSK-II and Aalpha or Aalpha randomized were added to the collagen matrix according to Example 8 comprising the suspension of PBMC, and further proceedings were in accordance with Example 8.














adhesion
migration


















a) no addition of NDSK-II
42 +/− 6 
10 +/− 1


b) only NDSK-II
96 +/− 11
24 +/− 3


c) 10 μg of Aalpha + NDSK-II
69 +/− 12
21 +/− 4


d) 100 μg of Aalpha + NDSK-II
73 +/− 13
15 +/− 6


e) 1000 μg of Aalpha + NDSK-II
70 +/− 6 
13 +/− 5


f) 10 μg of Aalpha randomized + NDSK-II
70 +/− 6 
25 +/− 2


g) 100 μg of Aalpha randomized + NDSK-II
65 +/− 16
24 +/− 3


h) 1000 μg of Aalpha randomized + NDSK-II
70 +/− 12
26 +/− 3









As can be gathered from the test results, peptide Aalpha blocks the migration of PBMC only partially.

Example 11
Since PBMC substantially consists of a mixture of lymphocytes and monocytes, pure lymphocytes instead of PBMC (as in Examples 8-10) were used in Example 11.
100 μg of NDSK-uPA or 100 μg of NDSK-II, respectively, and Aalpha or Bbeta, respectively, were added to the collagen matrix according to Example 8 comprising endothelial cells and lymphocytes.














adhesion
migration


















a) no addition
68 +/− 8
16 +/− 3


b) NDSK-uPA
143 +/− 11
53 +/− 5


c) NDSK-II
119 +/− 11
43 +/− 4


d) only 100 μg of Bbeta
 58 +/− 18
14 +/− 1


e) NDSK-uPA + 100 μg of Bbeta
74 +/− 8
19 +/− 2


f) NDSK-II + 100 μg of Bbeta
74 +/− 8
17 +/− 3


g) only 100 μg of Aalpha
77 +/− 4
18 +/− 1


h) NDSK-uPA + 100 μg of Aalpha
131 +/− 4 
40 +/− 3


i) NDSK-II + 100 μg of Aalpha
131 +/− 4 
44 +/− 4


j) only 100 μg of Bbeta randomized
75 +/− 5
19 +/− 1


k) NDSK-uPA + 100 μg of Bbeta randomized
134 +/− 13
46 +/− 4


l) NDSK-II + 100 μg of Bbeta randomized
120 +/− 12
42 +/− 4









Those test results show

    
    
        1) that both NDSK-II and NDSK-uPA promote lymphocytic inflammation,
        2) that peptide Bbeta completely blocks the lymphocytic adhesion and migration induced by NDSK-II and NDSK-uPA, whereas peptide Aalpha exhibits no blocking activity, which suggests that the free alpha-chain is not required for inducing the adhesion and migration of the lymphocytes.
    
    


Example 12
The proceedings were in accordance with Example 11, except for pure monocytes being used instead of lymphocytes. 100 μg of NDSK-uPA or 100 μg of NDSK-II, respectively, was added to peptide Aalpha, randomized Aalpha, Bbeta or randomized Bbeta.














adhesion
migration


















a) no addition
43 +/− 8
7 +/− 1


b) NDSK-uPA
 48 +/− 10
10 +/− 2 


c) NDSK-II
 90 +/− 11
19 +/− 6 


d) 100 μg of Bbeta
59 +/− 7
5 +/− 1


e) NDSK-uPA + 100 μg of Bbeta
 61 +/− 11
8 +/− 3


f) NDSK-II + 100 μg of Bbeta
70 +/− 7
7 +/− 5


g) 100 μg of Bbeta randomized
40 +/− 7
6 +/− 1


h) NDSK-uPA + 100 μg of Bbeta randomized
45 +/− 5
8 +/− 3


g) NDSK-II + 100 μg of Bbeta randomized
 92 +/− 10
20 +/− 7 


j) 100 μg of Aalpha
59 +/− 6
5 +/− 1


k) NDSK-uPA + 100 μg of Aalpha
62 +/− 4
8 +/− 5


l) NDSK-II + 100 μg of Aalpha
 68 +/− 10
9 +/− 6


m) 100 μg of Aalpha randomized
58 +/− 7
6 +/− 1


n) NDSK-uPA + 100 μg of Aalpha randomized
 50 +/− 10
10 +/− 4 


o) NDSK-II + 100 μg of Aalpha randomized
108 +/− 8 
21 +/− 5 









Those test results show that only NDSK-II and not NDSK-uPA promotes the migration of monocytes, which means that both the alpha-chain and the beta-chain have to exhibit a free N-terminal end and block the migration of the monocytes.

Example 13
The proceedings were in accordance with Example 11, with pure lymphocytes being used. 100 μg of NDSK-uPA or 100 μg of NDSK-II, respectively, was added to the short peptide salts derived from Aalpha Gly Pro Arg (Pro)—NH2 acetate (Aalpha derivative) or derived from Bbeta Gly His Arg Pro-OH acetate (Bbeta derivative).














adhesion
migration



















a)
no addition
60 +/− 8
14 +/− 1


b)
NDSK-uPA
149 +/− 12
57 +/− 5


c)
NDSK-II
121 +/− 11
48 +/− 7


d)
only 100 μg of Bbeta derivative
 58 +/− 10
12 +/− 9


e)
NDSK-uPA + 100 μg of Bbeta derivative
70 +/− 8
16 +/− 3


f)
NDSK-II + 100 μg of Bbeta derivative
69 +/− 7
14 +/− 5


g)
only 100 μg of Aalpha derivative
77 +/− 4
18 +/− 1


h)
NDSK-uPA + 100 μg of Aalpha derivative
134 +/− 4 
48 +/− 5


i)
NDSK-II + 100 μg of Aalpha derivative
131 +/− 7 
49 +/− 6


j)
only 100 μg of Bbeta derivative
70 +/− 5
14 +/− 7



randomized


k)
NDSK-uPA + 100 μg of Bbeta derivative
130 +/− 12
49 +/− 6



randomized


l)
NDSK-II + 100 μg of Bbeta derivative
120 +/− 10
55 +/− 8



randomized









Said experiment allows the conclusion that, if lymphocytic migration is inhibited, those short peptides, added continuously in an appropriate manner, exhibit the same activity as do the long peptides.
Example 14
The proceedings were in accordance with Example 12, with pure monocytes being used. 100 mg of NDSK-uPA or 100 μg of NDSK-II, respectively, was added to the short peptide salts Aalpha Gly Pro Arg (Pro)—NH2 acetate (Aalpha derivative) or Bbeta Gly His Arg Pro-OH acetate (Bbeta derivative).














adhesion
migration



















a)
no addition
40 +/− 8
5 +/− 1


b)
NDSK-uPA
54 +/− 9
7 +/− 2


c)
NDSK-II
 85 +/− 11
22 +/− 6 


d)
100 μg of Bbeta derivative
52 +/− 7
6 +/− 1


e)
NDSK-uPA + 100 μg of Bbeta derivative
 61 +/− 11
8 +/− 3


f)
NDSK-II + 100 μg of Bbeta derivative
68 +/− 7
8 +/− 4


g)
100 μg of Bbeta derivative randomized
40 +/− 7
6 +/− 1


h)
NDSK-uPA + 100 μg of Bbeta derivative
44 +/− 6
8 +/− 2



randomized


i)
NDSK-II + 100 μg of Bbeta derivative
 92 +/− 10
23 +/− 7 



randomized


j)
100 μg of Aalpha derivative
50 +/− 5
4 +/− 4


k)
NDSK-uPA + 100 μg of Aalpha derivative
60 +/− 5
7 +/− 6


l)
NDSK-II + 100 μg of Aalpha derivative
 64 +/− 11
8 +/− 2


m)
100 μg of Aalpha derivative randomized
 54 +/− 10
6 +/− 3


n)
NDSK-uPA + 100 μg of Aalpha derivative
 50 +/− 10
10 +/− 4 



randomized


o)
NDSK-II + 100 μg of Aalpha derivative
99 +/− 8
21 +/− 7 



randomized









Said experiment allows the conclusion that, if monocytic migration is inhibited, those short peptides, added continuously in an appropriate manner, exhibit the same activity as do the long peptides.
Example 15
The tests were carried out on male wistar rats weighing between 220 g and 280 g. The rats were given standard food and water. For carrying out the test, the rats were anaesthetized and artifically respirated with a frequency of 70 pulses per minute, whereby from 8 ml to 10 ml per kilogram of a gas containing 30 % by volume of oxygen and having an overpressure of from 1 mm to 2 mm mercury was emitted. The cardiac artery on the right hand side was equipped with a measuring cannula, and the blood pressure in the artery as well as the heartbeats were determined. The pressure rate was determined as a product of the blood pressure in the artery and of the heartbeat rate with the dimension mm mercury/minute/103. The vein on the right hand side was equipped with a measuring cannula for doping the test substances. After carrying out the surgical treatment, 2 ml of rat blood was supplied to the heart. Thirty minutes later, the cardiac artery on the left hand side was occluded. Another twenty-five minutes later, the occlusion was released in order to resupply the ischaemic area with blood. At that point of time, 800 μg/kg of peptide Bbeta or peptide Bbeta randomized, respectively, was intravenously administered to half of the animals, and then two hours were allowed to pass.
In order to distinguish between damaged and undamaged cardiac tissue, the cardiac artery on the left hand side was then supplied with evans blue dye at a concentration of 2% by weight. Thereupon, the removed heart was dissected by five horizontal cuts, the right hand wall of the vein was removed and the sections were treated with triphenyltetratolchloride (1% by weight) for twenty minutes at 37° C. so as to be able to distinguish between normal tissue and infarct tissue. The sections were evaluated by computer-sustained planimetry.
Because of the vascular occlusion, 62.5% of the cardiac muscle in the hearts of the reference rats was threatened, as opposed to 60% in the hearts of the test rats. In the hearts of the reference rats, 46% of the endangered tissue was dead, as opposed to 29% in the hearts of the test rats. That corresponds to a 37% reduction of dead tissue (p<0.05).
The substances according to the invention as well as the use of the substances according to the invention for preparing a pharmaceutical composition are of special significance:
For a pharmaceutical composition used in the therapy of diseases caused by the tissue-damaging effect of autoreactive lymphocytes.
Among those are diseases fitting into the sphere of autoimmunity, such as collagenoses, rheumatic diseases, psoriasis and post-/parainfectious diseases and diseases caused by a graft versus host reaction. A healing effect occurs, since said pharmaceutical composition blocks the migration of lymphocytes into the tissue. Thus, the lymphocytes remain in the bloodstream and are incapable of producing an autoreactive tissue-damaging effect.
A healing effect occurs with a drug for the therapy and/or prevention of the rejection occurring after organ transplants, since said drug prevents the migration of lymphocytes from the bloodstream into the foreign organ and hence the foreign organ cannot be destroyed by autoreactive lymphocytes.
A healing effect occurs with a drug for the therapy and/or prevention of arterial sclerosis after organ transplants, since said drug prohibits the migration of lymphocytes and monocytes into the vascular wall and hence prevents the activation of the cells of the vascular wall. In doing so, the occurrence of arterial sclerosis following organ transplants is minimized or prevented.
A healing effect occurs with a drug for the therapy and/or prevention of a reperfusion trauma following a surgically or pharmaceutically induced restoration of the blood flow such as, f.i. after cardiac infarction, apoplectic stroke, after vascular surgery, bypass surgery and organ transplants, since said drug inhibits the migration of lymphocytes and monocytes into the vascular wall. The reperfusion trauma is caused by oxygen deficiency/acidosis occurring in the cells of the vessel during the restoration of the blood flow and leads to their activation. Thereby, lymphocytes and monocytes adhere to the vascular wall and migrate into the same. The fact that lymphocytes and monocytes are prevented from adhering to and migrating into the vascular wall brings about a decrease in the hypoxia/acidosis-induced damage, without any permanent vascular damage being caused by the subsequent inflammatory reaction.
A healing effect occurs with a drug for the therapy and/or prevention of arterial sclerosis following metabolic diseases or ageing processes, since said drug inhibits the migration of lymphocytes and monocytes into the vascular wall and hence inhibits the progredience of the arteriosclerotic plaque resulting therefrom.
The pharmaceutical composition according to the invention may also be used for transporting a further drug substance. The pharmaceutical composition according to the invention specifically binds a surface molecule to endothelial cells. Thus, drug substances coupled thereto may be contacted with endothelial cells at high concentrations, without them being able to trigger side reactions in other places. The use of substances inhibiting cell division may be mentioned as an example, which substances may exhibit an antiangiogenetic effect after having been adducted specifically to endothelial cells. In that case, tumour patients experience a healing effect, since the growth of the tumour is blocked by preventing the proliferation of endothelial cells and hence by avoiding neoangiogenesis.







TABLE 1







Peptides of Formula I: Gly - His/Pro - Arg - Xaa2 - Xaa29








SEQUENCE
SEQ ID NO












Gly His Arg
1



 


Gly Pro Arg
2


 


Gly His Arg Xaa
3


 


Gly Pro Arg Xaa
4


 


Gly His Arg Xaa Xaa
5


 


Gly Pro Arg Xaa Xaa
6


 


Gly His Arg Xaa Xaa Xaa
7


 


Gly Pro Arg Xaa Xaa Xaa
8


 


Gly His Arg Xaa Xaa Xaa Xaa
9


 


Gly Pro Arg Xaa Xaa Xaa Xaa
10


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa
11


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa
12


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa
13


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa
14


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa
15


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa
16


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
17


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
18


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
19


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
20


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
21


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
22


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
23


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
24


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
25


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
26


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
27


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
28


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
29


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
30


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
31


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
32


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
33


Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
34


Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
295


Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
296


Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
35


Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
36


Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
37


Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
38


Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
39


Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
40


Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
41


Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
42


Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
43


Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
44


Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
45


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
46


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
47


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
48


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
49


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
50


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
51


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
52


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
53


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
54


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
55


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
56


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
57


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
58


Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa

















TABLE 2







Peptides of Formula II: Gly - His/Pro - Arg - Pro/Val -



Leu/Val - Xaa2 - Xaa30








SEQUENCE
SEQ ID NO












Gly His Arg Pro Leu Xaa Xaa
59



 


Gly Pro Arg Pro Leu Xaa Xaa
60


 


Gly His Arg Val Leu Xaa Xaa
61


 


Gly Pro Arg Val Leu Xaa Xaa
62


 


Gly His Arg Pro Val Xaa Xaa
63


 


Gly Pro Arg Pro Val Xaa Xaa
64


 


Gly His Arg Val Val Xaa Xaa
65


 


Gly Pro Arg Val Val Xaa Xaa
66


 


Gly His Arg Pro Leu Xaa Xaa Xaa
67


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa
68


 


Gly His Arg Val Leu Xaa Xaa Xaa
69


 


Gly Pro Arg Val Leu Xaa Xaa Xaa
70


 


Gly His Arg Pro Val Xaa Xaa Xaa
71


 


Gly Pro Arg Pro Val Xaa Xaa Xaa
72


 


Gly His Arg Val Val Xaa Xaa Xaa
73


 


Gly Pro Arg Val Val Xaa Xaa Xaa
74


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa
75


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa
76


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa
77


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa
78


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa
79


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa
80


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa
81


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa
82


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa
83


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa
84


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa
85


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa
86


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa
87


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa
88


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa
89


 


Gly Pro Arg Val Val Xaa Xa.a Xaa Xaa Xaa
90


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa
91


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa
92


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa
93


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa
94


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa
95


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa
96


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa
97


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa
98


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa
99


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa
100


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa
101


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa
102


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa
103


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa
104


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa
105


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa
106


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
107


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
108


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
109


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
110


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
111


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
112


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
113


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
114


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
115


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
116


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
117


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
118


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
119


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
120


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
121


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
122


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
123


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
124


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
125


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
126


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
127


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
128


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
129


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
130


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
131


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
132


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
133


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
134


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
135


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
136


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
137


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
138


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
139


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
140


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
141


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
142


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
143


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
144


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
145


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
146


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
147


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
148


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
149


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
150


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
151


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
152


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
153


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
154


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
155


Xaa


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
156


Xaa


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
157


Xaa


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
158


Xaa


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
159


Xaa


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
160


Xaa


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
161


Xaa


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
162


Xaa


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
163


Xaa Xaa


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
164


Xaa Xaa


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
165


Xaa Xaa


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
166


Xaa Xaa


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
167


Xaa Xaa


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
168


Xaa Xaa


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
169


Xaa Xaa


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
170


Xaa Xaa


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
171


Xaa Xaa Xaa


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
172


Xaa Xaa Xaa


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
173


Xaa Xaa Xaa


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
174


Xaa Xaa Xaa


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
175


Xaa Xaa Xaa


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
176


Xaa Xaa Xaa


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
177


Xaa Xaa Xaa


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
178


Xaa Xaa Xaa


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
179


Xaa Xaa Xaa Xaa


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
180


Xaa Xaa Xaa Xaa


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
181


Xaa Xaa Xaa Xaa


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
182


Xaa Xaa Xaa Xaa


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
183


Xaa Xaa Xaa Xaa


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
184


Xaa Xaa Xaa Xaa


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
185


Xaa Xaa Xaa Xaa


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
186


Xaa Xaa Xaa Xaa


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
187


Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
188


Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
189


Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
190


Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
191


Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
192


Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
193


Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
194


Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
195


Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
196


Xaa Xaa Xaa Xaa Xaa Xaa


 


Uty His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
197


Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
198


Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
199


Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
200


Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
201


Xaa Xaa Xaa Xaa Xa.a Xaa


 


Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
202


Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
203


Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
204


Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
205


Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
206


Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
207


Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly Pro Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
208


Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
209


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Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


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Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Pro Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Val Leu Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly His Arg Pro Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa


 


Gly His Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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Gly Pro Arg Val Val Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa
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## Claims

1. A method of treating rejection of a transplanted tissue in a subject comprising administering to the subject a peptide








(SEQ ID NO:294)









Gly His Arg Pro Leu Asp Lys Lys Arg Glu Glu Ala



 


Pro Ser Leu Arg Pro Ala Pro Pro Pro Ile Ser Gly


 


Gly Gly Tyr Arg






or a salt or amide thereof, in an amount effective to treat rejection of transplanted tissue, wherein the amino terminus is



wherein R1 and R2 are either the same or different,
wherein R1 and R2 are each selected from the group consisting of hydrogen and a saturated or unsaturated hydrocarbon residue, said residue having from 1 to 10 carbon atoms, and
wherein the peptide, or a salt or amide thereof, comprises a peptide derived from a source selected from the group consisting of the Aalpha-chain of fibrin and the Bbeta chain of fibrin.

2. A method of inhibiting inflammation of a transplanted tissue in a subject comprising administering to the subject a peptide








(SEQ ID NO:294)









Gly His Arg Pro Leu Asp Lys Lys Arg Glu Glu Ala



 


Pro Ser Leu Arg Pro Ala Pro Pro Pro Ile Ser Gly


 


Gly Gly Tyr Arg






or a salt or amide thereof, in an amount effective to treat rejection of transplanted tissue, wherein the amino terminus is



wherein R1 and R2 are either the same or different,
wherein R1 and R2 are each selected from the group consisting of hydrogen and a saturated or unsaturated hydrocarbon residue, said residue having from 1 to 10 carbon atoms.

3. The method of claim 2, wherein the peptide, or a salt or amide thereof, comprises a peptide derived from a source selected from the group consisting of the Aalpha-chain of fibrin and the Bbeta chain of fibrin.