Chemistry: natural resins or derivatives; peptides or proteins; – Peptides of 3 to 100 amino acid residues – 8 to 10 amino acid residues in defined sequence
Reexamination Certificate
2001-01-08
2004-11-30
Weber, Jon P. (Department: 1653)
Chemistry: natural resins or derivatives; peptides or proteins;
Peptides of 3 to 100 amino acid residues
8 to 10 amino acid residues in defined sequence
C514S017400, C514S018700, C514S015800, C530S327000, C530S329000, C530S330000, C530S403000, C424S185100
Reexamination Certificate
active
06825319
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to synthetic peptides and to pharmaceutical compositions comprising them for the diagnosis and treatment of anti-phospholipid syndrome.
ABBREVIATIONS: AFC: antibody-forming cells; APS: anti-phospholipid syndrome; HUVEC: human umbilical vein endothelial cells; mAb: monoclonal antibody; MAP: multiple antigenic peptide; PBL: peripheral blood lymphocytes; SLE: systemic lupus erythematosus; St: streptavidin; &bgr;2GPI: beta-2-glycoprotein 1.
BACKGROUND OF THE INVENTION
Autoimmune diseases are disorders in which the immune system produces autoantibodies directed against an endogenous antigen, with consequent injury to tissues. These self antigens, called also autoantigens, despite being normal tissue constituents, are the target of a humoral or cell-mediated immune response that characterizes the autoimmune disease.
Several connective tissue disorders including vascular diseases, such as vasculitis, systemic lupus erythematosus (SLE), and polymyositis, neurologic diseases such as multiple sclerosis and myasthenia gravis, and hematologic diseases such as idiopathic thrombocytopenia purpura (ITP) and anti-phospholipid syndrome (APS) seem to be caused by an autoimmune reaction. For some of these disorders, the self antigen has been identified and/or pathogenic autoantibodies have been identified and isolated.
No specific drugs exist nowadays for the treatment of autoimmune diseases and patients are treated with anti-inflammatory drugs such as corticosteroids and/or immunosuppressive drugs. All research being carried out in this field is directed to the development of drugs specific for each disease.
Anti-phospholipid antibodies have been associated with a variety of clinical phenomena, including arterial and venous thrombosis, thrombocytopenia, and obstetric complications. The term “anti-phospholipid syndrome” is used to link a variety of thrombotic events to antibodies against specific proteins involved in blood coagulation. Thrombotic events are reported in approximately 30% of patients with anti-phospholipid antibodies, with an overall incidence of 2.5% patients/year. Deep vein thrombosis of the legs and/or thrombotic events, and cerebral arterial thrombosis are the most common arterial complications. Obstetric complications include recurrent spontaneous miscarriages, fetal deaths, or fetal growth retardations. Women with anti-phospholipid antibodies are particularly prone to second or third trimester fetal death.
The anti-phospholipid syndrome (APS) is characterized by the presence of high titers of anti-cardiolipin and/or anti-&bgr;2GPI (beta-2-glycoprotein 1) antibodies which might have lupus anti-coagulant activity leading to thromboembolic phenomena, thrombocytopenia, recurrent fetal loss, as well as other multisystemic involvements. APS can emerge as a primary syndrome or as secondary syndrome to SLE (Hughes et al., 1986; McNeil et al., 1991).
Anti-&bgr;2GPI antibodies bind anionic phospholipids through the &bgr;2GPI molecule (McNeil et al., 1990; Igarashi et al., 1996). &bgr;2GPI is the target antigen for the autoimmune anti-&bgr;2GPI antibodies previously entitled ‘anti-cardiolipin/anti-phospholipid &bgr;2GPI dependent antibodies’. &bgr;2GPI (50KD), initially described by Schultze et al. (1961), is composed of five respective consensus (‘sushi’ like) repeats (Kandiah and Krilis, 1994). &bgr;2GPI binds negatively charged phospholipids through a lysine-rich locus (Cys281-Cys288) located in the fifth domain (Hunt and Krilis, 1994) and possesses several in vitro properties which define it as an anticoagulant, i.e., it causes inhibition of prothrombinase activity, ADP-induced platelet aggregation, platelet factor IX production (Sheng et al., 1996). Employing site-directed mutagenesis of recombinant human &bgr;2GPI, a cluster of lysine residues that are critical for phospholipid binding and anti-cardiolipin antibody activity was identified (Sheng et al., 1996).
The anti-&bgr;2GPI antibodies have been considered to exert a direct pathogenic effect by interfering with hemostatic reactions occurring on the surface of platelets or vascular endothelial cells (Shi et al, 1993; Simantov et al., 1995). Passive transfer of these antibodies into naive mice or mice prone to develop APS, resulted in induction of experimental APS in mice (Blank et al., 1991). It has been shown recently (Del Papa et al., 1997; George et al., 1998) that human polyclonal and monoclonal anti-&bgr;2GPI antibodies react in vitro with endothelial cells through adherent &bgr;2GPI and induce differential endothelial cell activation. It is not clear to which epitopes on the &bgr;2GPI molecule these anti-&bgr;2GPI antibodies are directed, and the correlation to their biological activity.
Attempts have been made to find peptides that could mimic the self antigen-epitope and would inhibit the autoantibody/self antigen binding and consequent injury to the tissue. Thus, recently, peptides selected from phage-epitope libraries through binding to pathogenic monoclonal autoantibodies were shown to provide a surrogate antigen or mimotope that inhibits binding to the original antigen. Such peptides reflect the sequence or conformation of the antigen-binding site, and the fine specificity of the autoantibodies to the protein and non-protein, e.g. polysaccharides or dsDNA, antigens (Scott and Smith, 1990; Scott et al., 1992; Yayon et al., 1993).
SUMMARY OF THE INVENTION
It is an object of the present invention to provide means for diagnosis and specific treatment of the autoimmune disorder anti-phospholipid syndrome (APS).
A further object of the invention is to provide means for inactivating B-cells responsible for the production of autoantibodies appearing in APS patients.
The present invention relates to synthetic peptides suitable for the diagnosis and treatment of APS, more particularly to synthetic peptides and derivatives thereof capable of inhibiting the biological activity of anti-&bgr;2GPI mabs in vitro. and of inhibiting induction of experimental APS in mice by anti-&bgr;2GPI mAbs.
In a preferred embodiment, the peptides of the invention and their derivatives are selected from the group consisting of:
(i) a peptide of at least 4 amino acid residues comprising a sequence selected from:
(a) Thr-Pro-Arg-Val
(b) Lys-Ala-Thr-Phe
(c) Leu-Arg-Val-Tyr
(ii) a cyclic derivative of a peptide of (i);
(iii) a peptide according to (i) or (ii) in which one or more amino acid residues have been replaced by the corresponding D-isomer or by a non-natural amino acid residue;
(iv) a chemical derivative of a peptide according to (i)-(iii);
(v) a multichain peptide-oligomer/polymer conjugate comprising two or more of the same or different peptides or peptide derivatives (i) to (iv) attached to a native or synthetic multifunctional oligomeric or macromolecular backbone; and
(vi) a multiple antigen peptide (MAP) in which two to eight same or different peptides or peptide derivatives (i) to (iv) are attached to a diaminoalkanoic acid core.
In one embodiment, a peptide according to (i)(a) above has a sequence selected from:
(a1) Leu-Lys-Thr-Pro-Arg-Val
(a2) Lys-Thr-Pro-Arg-Val-Thr
(A) Asn-Leu-Lys-Thr-Pro-Arg-Val-Gly-Gly
In another embodiment, a peptide according to (i)(b) above has a sequence selected from:
(b1) Lys-Asp-Lys-Ala-Thr-Phe
(B) Lys-Asp-Lys-Ala-Thr-Phe-Gly-Thr-His-Asp-Gly
In a further embodiment, a peptide according to (i)(c) above has a sequence selected from:
(c1) Thr-Leu-Arg-Val-Tyr-Lys
(c2) Thr-Lys-Leu-Arg-Val-Tyr
(c3) Thr-Leu-Leu-Arg-Val-Tyr
(C) Cys-Ala-Thr-Leu-Arg-Val-Tyr-Lys-Gly-Gly
(i) a peptide of at least 4 amino acid residues comprising a sequence selected from:
(a) Thr-Pro-Arg-Val (residues 1-4 of SEQ ID NC:1)
(b) Lys-Ala-Thr-Phe (residues 3-6 of SEQ ID NO:4)
(c) Leu-Arg-Val-Tyr (residues 4-7 of SEQ ID NO:7)
It has further been found according to the present invention that when a number of the same or different peptides or peptide derivatives (i) to (iv) which recognize and bind to autoantibodies secreted by specific B cells in APS are attached to a multifunctional oligo
Blank Miri
Cabilly Shmuel
Katchalski-Katzir Ephraim
Shoenfeld Yehuda
Browdy and Neimark PLLC
Lukton David
Weber Jon P.
Yeda Research and Development Co. Ltd.
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