WO2005032482A2 - Treatment of demyelinating autoimmune disease with modified ordered peptides - Google Patents
Treatment of demyelinating autoimmune disease with modified ordered peptides Download PDFInfo
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- WO2005032482A2 WO2005032482A2 PCT/US2004/032598 US2004032598W WO2005032482A2 WO 2005032482 A2 WO2005032482 A2 WO 2005032482A2 US 2004032598 W US2004032598 W US 2004032598W WO 2005032482 A2 WO2005032482 A2 WO 2005032482A2
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P37/00—Drugs for immunological or allergic disorders
- A61P37/02—Immunomodulators
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
Definitions
- MS Multiple sclerosis
- CNS central nervous system
- the concordance rate among monozygotic twins is 30%, a 10-fold increase over dizygotic twins or first-degree relatives.
- the higher incidence rate among monozygotic twins emphasizes the importance of genetic factors, but the discordance rate of 70% among identical twins illuminates the role of nongenetic factors on disease penetrance.
- HLA class II genes exert an influence, with HLA DR2 carrying a 4-fold relative risk for northern European caucasoids.
- a typical presentation of MS involves an initial course, running for several years to more than a decade, manifest by episodes of relapse followed by remission. Relapses often follow an episode of a viral infection of the upper respiratory system or gastrointestinal tract. In about one half of MS cases the disease progresses to a more chronic phase.
- Clinical problems may include disturbances in visual acuity, sometimes culminating in blindness; double vision; motor disturbances affecting walking and use of the hands; incoordination; bowel and bladder incontinence; spasticity; and sensory disturbances including loss of touch, pain, and temperature and proprioception.
- the pathology of the disease lies entirely in the central nervous system and is characterized by a classic picture of inflammation surrounding venules and extending into the myelin sheath.
- Immune responses to various components of the myelin sheath have been detected in MS patients, including myelin basic protein (MBP), proteolipid protein (PLP), transaldolase, and 2',3' cyclic nucleotide 3 * phosphodiesterases (CNP), as well as two members of the immunoglobulin supergene family found in the myelin sheath, myelin oligodendroglial glycoprotein (MOG) and myelin-associated glycoprotein (MAG) (Steinman et al. (1995) Mol. Med. Todav 1:79-83).
- MBP myelin basic protein
- PGP proteolipid protein
- CNP 2',3' cyclic nucleotide 3 * phosphodiesterases
- MOG myelin oligodendroglial glycoprotein
- MAG myelin-associated glycoprotein
- some inducible heat shock proteins can be detected in glial cells in MS lesions and can stimulate an immune response in MS patients.
- myelin proteins and epitopes were identified as targets of the autoimmune T and B cell response.
- Antibody eluted from MS brain plaques recognized myelin basic protein (MBP) peptide 83-97 (Wucherpfennig et al., J Clin Invest 100:1114-1122, 1997).
- MBP myelin basic protein
- T and B cell (brain lesion-eluted Ab) response focused on MBP 87-99 (Oksenberg et al., Nature 362, 68-70, 1993).
- MBP 87- 99 the amino acid motif HFFK is a dominant target of both the T and B cell response (Wucherpfennig et al., J Clin Invest 100, 1114-22, 1997).
- Another study observed lymphocyte reactivity against myelin-associated oligodendrocytic basic protein (MOBP), including residues MOBP 21-39 and MOBP 37-60 (Holz et al., J Immunol 164, 1103-9, 2000).
- MOBP myelin-associated oligodendrocytic basic protein
- the B cell response to MBP in MS has also been studied extensively. IgG purified from brain lesions reacted with the same region of MBP, p85-96, that is the immunodominant T cell epitope in MS patients who are HLA DR2b (DRB1*1501) and overlaps with the T cell epitope in MS patients who are DR2a (DRB5*0101).
- Literature Copolymer-1 is a mixture of polypeptides composed of alanine, glutamic acid, lysine, and tyrosine in a molar ratio of approximately 6:2:5:1 , respectively. It is synthesized by chemically polymerizing the four amino acids forming products with average molecular weights of 23,000 daltons (U.S. Pat. No. 3,849,550).
- Cop 1 binds promiscuously, with high affinity and in a peptide-specific manner to purified MS-associated HLA-DR2 (DRB1*1501) and rheumatoid arthritis- associated HLA-DR1 (DRB1*0101) or HLA-DR4 (DRB1*0401) molecules (Fridkis-Hareli et al. (1999) J Immunol 162(8):4697-704). Protruding N-terminal ends of Cop 1 bound to HLA-DR1 , -DR2, or -DR4 molecules were then treated with aminopeptidase I, followed by elution, HPLC, and pool sequencing.
- Copaxone treatment was found to lead to a significant reduction in the mean annual relapse rate and stabilization of disability.
- the treatment was accompanied by an elevation of serum IL-10 levels, suppression of the pro-inflammatory cytokine TNF alpha mRNA, and an elevation of the anti-inflammatory cytokines TGF-beta and IL-4 mRNAs in PBLs (Miller et al. (1998. J Neuroimmunol 92(1-2):113-21).
- Treatment of murine experimental autoimmune encephalomyelitis with a myelin basic protein peptide analog is described by Reiseter et al. (1998) J Neuroimmunol 91 (1-2): 156-70.
- J.E.M. 180:2227- 2237 demonstrate the reversal of experimental autoimmune encephalomyelitis by a soluble variant of a myelin basic protein epitope. It has been reported that administration of myelin basic protein can lead to immune tolerance (see, for example, Steinman et al. (1977) Nature 265:173; Tonegawa (1997) J Exp Med 186(4):507-15; Hafler et al. (1997) Ann N Y Acad Sci 835:120-31 ; Kennedy et al. (1997) J Immunol 159(2): 1036-44). Various forms of Ag-specific tolerance have been demonstrated, included the administration of peptide coupled splenocytes, i.p.
- MBP therapeutic ordered peptide of this invention comprises the ordered amino acid motif ⁇ SEQ ID NO:1 ⁇ [ 1 ⁇ 2 ⁇ 3 ⁇ 4 ⁇ ]n, where n is from 2 to 6, modified at the amino or carboxy terminal end.
- the ordered motif may start at residue 1 , as shown, or may start at a different position, e.g. ⁇ SEQ ID NO:2 ⁇ YYKEYYKE; ⁇ SEQ ID NO:3 ⁇ YKEYYKEY; etc.
- a MOG therapeutic ordered peptide of this invention comprises the ordered amino acid motif ( 1 Y 2 R 3 E 4 Y 5 E 6 Y 7 E) n where n is from 2 to 10.
- the MOG therapeutic ordered peptide of this invention may be modified at the amino or carboxy terminal end.
- a PLP therapeutic ordered peptide of this invention comprises the ordered amino acid motif ( 1 Y 2 G 3 K 4 E 5 L 6 G 7 E 8 Y) disciplin where n is from 2 to 10.
- the PLP therapeutic ordered peptide of this invention may be modified at the amino or carboxy terminal end.
- Other therapeutic ordered peptides of this invention include such peptides from cyclic nucleotide phosphodiesteerase (CNPase), myelin associated glycoprotein (MAG), myelin-associated oligodendrocytic basic protein (MBOP), and alpha-B-crystalin (a heat shock protein).
- Therapeutic ordered peptides of other proteins and epitopes, identified to be targets of the autoimmune T and B cell responses, can be designed and administered using the teaching of this invention.
- the compositions of the present invention may be synthesized by conventional methods known in the art, e.g.
- the therapeutic ordered peptides are formulated in a biologically acceptable carrier, and administered by a route to enhance the autoimmune suppressive effects of the treatment.
- the therapeutic ordered peptides are administered on a regular basis to patients suffering from multiple sclerosis.
- the composition is lyophilized and formed into an aqueous solution suitable for subcutaneous injection and administered on a regular basis in accordance with the method of this invention.
- Figure 1 is a graph showing the prevention of EAE in rats treated with therapeutic ordered peptides.
- MBP therapeutic ordered peptide ⁇ SEQ .D NO:4 ⁇ EYYKEYYKEYYK prevents the development of EAE in Lewis rats. Animals were injected with an emulsion of 0.1 mg of MBPp85-99 in complete Freund's adjuvant for EAE induction. Ten days later, when the clinical manifestations of disease became apparent, a single intra-peritoneal dose of MBP therapeutic ordered peptide ⁇ SEQ ID NO:4 ⁇ EYYKEYYKEYK (squares), ⁇ SEQ ID NO:5 ⁇ KYYKYYKYYKYYYYYY (triangles), or PBS (circles)was administered. Results are expressed as mean disease score of groups of six animals.
- Figure 2 is a graph depicting the reduction in relapse rates in mice with EAE treated with ordered peptides.
- Animals were induced for EAE with an emulsion of 0.1 mg of PLPp139-151 in complete Freund's adjuvant (day 0).
- Mice were randomized into equal groups at the peak of disease and were treated at days 17, 29, and 36 after EAE induction with intravenous, intra-peritoneal or subcutaneous administration of EYYKEYYKEYYK (MBP therapeutic ordered peptide) or Copaxone at two different dosages, 0.5 mg per mouse or 0.05 mg per mouse. Both the MBP therapeutic ordered peptide and Copaxone were dissolved in mannitol. Results are expressed as relapse rates per mouse, and show that the MBP therapeutic ordered peptide and Copaxone reduce relapse rates.
- Figure 3 is a table shows the ordered peptide blocking MHC binding by the native peptide.
- the ability of MBP therapeutic ordered peptide EYYKEYYKEYYK, substituted MBP therapeutic ordered peptide D-Ala- EYYKEYYKEYYK-amide, or Copaxone to block the binding of the native peptide to either a mouse or rat MHC was measured by FACS analysis. As shown in the table as indicated by the lower mean florescence intensity (MFI), the substituted D-ala form of the therapeutic ordered peptide and Copaxone was more effective in blocking either MHC than the non-D-ala form.
- MFI mean florescence intensity
- Figure 4 is a graph showing the blocking by the therapeutic ordered peptide of T cell proliferation.
- MBP therapeutic ordered peptide EYYKEYYKEYYK
- substituted MBP therapeutic ordered peptide D-Ala- EYYKEYYKEYYK-amide
- Copaxone Copaxone
- Figure 5 is a graph showing the blocking of induction of EAE by the substituted D-Ala form of the therapeutic ordered peptide.
- Lewis rats were injected with an emulsion of 0.1 mg of MBPp85-99 in complete Freund's adjuvant for EAE induction.
- the substituted D-Ala therapeutic ordered peptide was more effective in blocking EAE induction than the un-substituted therapeutic ordered peptide.
- FIG. 6 shows cytokine production from T cells derived from mice immunized with D-ala modified ordered peptide, non-modified ordered peptide, and control peptides including Cop1 and ovalbumin (OVA).
- OVA is known to cause an induction of Th1 type of T cells after immunization.
- IL4 and IL10 in the D-ala-ordered peptide (D-ala-EYYK) immunized T cell lines, but not in the non-modified ordered peptide (EYYK) immunized T cell lines.
- Cop1 caused an increase in these two Th2 cytokines as expected, and OVA did not cause an increase in these cytokines also as expected.
- D-ala modified form of the ordered peptide can cause Th2 induction but that the unmodified peptide cannot.
- DETAILED DESCRIPTION OF THE EMBODIMENTS Demyelinating autoimmune diseases, including experimental autoimmune encephalomyelitis and multiple sclerosis, are treated by administering a therapeutic ordered peptide.
- the therapeutic ordered peptides are formulated in a pharmaceutically acceptable carrier for a convenient route of administration, which may be sub-cutaneous, oral, by inhalation, etc. as known in the art.
- the subject methods are used for prophylactic or therapeutic purposes.
- the term "treating" is used to refer to both prevention of disease, and treatment of pre-existing conditions.
- the prevention of autoimmune disease is accomplished by administration of the peptide prior to development of overt disease.
- the treatment of ongoing disease, in order to stabilize or improve the clinical symptoms of the patient, is of particular interest.
- Such treatment is desirably performed prior to loss of function in the affected tissues.
- Evidence of therapeutic effect may be any diminution in the severity of disease, particularly measuring the frequency of relapses in patients being treated with the ordered peptides, which may be the length of time the patient is relapse free, or the mean relapse frequency.
- Therapeutic ordered peptides of the present invention comprise eight to eighty amino acids representing a consensus sequence of a protein identified as a target of the autoimmune T and 8, cell response.
- the myelin proteins MBP, MOG, PLP, MAG and cyclic nucleotide phosphodiesterase are examples of proteins that are the target of the autoimmune response for which a therapeutic ordered peptide would be developed according to the teaching of this invention.
- the MBP therapeutic ordered amino acid motif is ⁇ SEQ ID NO:1 ⁇ [ E 2 Y 3 Y 4 K] ⁇ , where n is from 2 to 6.
- the MBP therapeutic ordered motif may start at residue 1, as shown, or may start at a different position, e.g.
- the total length of the MBP therapeutic ordered peptide sequence will usually be at least about 8 amino acids in length and not more than about 24 amino acids in length, usually at least about 10 and not more than about 20.
- Specific MBP therapeutic ordered peptides of interest include the sequence ⁇ SEQ ID NO:4 ⁇ EYYKEYYKEYYK.
- the amino acid motif is [ 1 Y 2 R 3 E 4 Y 5 E 6 Y 7 E] n where n is from 2 to 10.
- the MOG therapeutic ordered peptide motif may start at residue 1 or may start at a different position, REYEYEYREYEYEYREYEYE, or EYEYEYREYEYEYREYEYE.
- the total length of the MOG therapeutic ordered peptide will usually be at least about 14 amino acids in length and not more than 70 amino acids in length, usually about 18 amino acids and not more than about 42.
- the amino acid motif is [ 1 Y 2 G 3 K 4 E 5 L 6 G 7 E 8 Y] n where n is from 2 to 10.
- the PLP therapeutic ordered peptide motif may start at residue 1 or may start at a different position, e.g.
- the total length of the PLP therapeutic ordered peptide sequence will usually be at least about 16 amino acids in length and not more than about 80 amino acids in length, usually at least about 20 and not more than about 48. Included within the scope of therapeutic ordered peptides as that term is used herein are amino acid sequence variants. Such therapeutic ordered peptides are referred to herein as "substituted” or "modified” therapeutic ordered peptide. The amino acid sequence variants of therapeutic ordered peptides fall into two classes; either substitutional or additional.
- the therapeutic ordered peptide variants may be prepared by site specific mutagenesis of nucleotides in the DNA encoding the therapeutic ordered peptide if a recombinant expression system is used or by altering the synthetic scheme in solid phase peptide synthesis.
- Amino acid sequence variants are characterized by the predetermined nature of the variation.
- the therapeutic ordered peptide variants typically exhibit the same qualitative biological activity as the therapeutic ordered peptide, for example MHC binding, effect on T cell proliferation, effect on disease severity and relapse rate in EAE etc.
- the peptide may consist only of the ordered repeats, or may be extended at either terminusby the addition of other amino acid residues.
- Modification and changes may be made in the structure of the ordered peptide and still obtain a molecule having the desired characteristic of suppressing demyelinating autoimmune disease.
- the desired properties may be determined, at least in part, in an in vitro assay, where binding to the MHC antigen HLA-DR, particularly HLA-DR2 (DRB1*1501), is indicative of the relevant biological activity.
- the modified ordered peptides of this invention are likely to bind MHC antigen HLA-DR2 as well as other MHC antigens as would be known to one of ordinary skill in the art.
- HLA-DR2 modified PLP ordered peptides may also bind HLA-DR15.
- modified MOG ordered peptides in addition to their likely binding of HLA-DR2 may also bind HLA-DRB1 , HLA-DRB5, and HLA-DR4.
- certain amino acids may be substituted for other amino acids in a protein structure without appreciable loss of function.
- changes such as to protein stability or efficiency
- the resultant protein will be considered a biologically functional equivalent for the purposes of the invention.
- Amino acid substitutions are typically of single residues: insertions usually will be on the order of about from 1 to 4 amino acid residues; and deletions will range about from 1 to 4 residues. Deletions or insertions preferably are made in adjacent pairs, i.e. a deletion of 2 residues or insertion of 2 residues. Substitutions, deletions, insertions or any combination thereof may be combined to arrive at a final substituted therapeutic ordered peptide. Substitutional variants are those in which at least one residue of a therapeutic ordered peptide has been removed and a different residue inserted in its place. Such substitutions generally are made in accordance with the following table when it is desired to finely modulate the characteristics of a therapeutic ordered peptide.
- electropositive side chain e.g., lysyl, arginyl, or histidyl
- an electronegative residue e.g., glutamyl or aspartyl
- a residue having a bulky side chain e.g., phenylalanine
- Deletions of cysteine or other labile residues also may be desirable, for example in increasing the oxidative stability of the ordered peptide.
- Deletions or substitutions of potential proteolysis sites e.g. Arg Arg, is accomplished by deleting one of the basic residues or substituting one by glutaminyl or histidyl residues.
- the stability of the ordered peptide may be improved by D-amino acid additions or substitutions.
- D-amino acid additions at the N- and/or C-terminal of the ordered peptide, as well as internal D-amino acid substitutions, are made to maintain the helical structure of the peptide, and to maintain the biological characteristics of the ordered peptide while improving the stability and half-life of the ordered peptide.
- Certain D-amino acid additions or substitutions may enhance the biological activity of the therapeutic ordered peptide as described herein.
- the therapeutic ordered peptides may be provided in a variety of ways, being joined to non-wild-type flanking regions, as fused proteins, joined by linking groups or directly covalently linked through cysteine (disulfide) or peptide linkages.
- the therapeutic ordered peptides may be joined to a single amino acid, either a D- or L- amino acid, at the N- or C-terminus or a chain of amino acids.
- the fused peptides may be extended to provide convenient linking sites, e.g. cysteine or lysine, to enhance stability, to bind to particular receptors, to provide for site-directed action, to provide for ease of purification, to alter the physical characteristics (e.g. solubility, charge, etc.), to stabilize the conformation, etc.
- the therapeutic ordered peptide may be N-terminal, C-terminal or internal in relation to these added sequences.
- the therapeutic ordered peptide may be linked through a variety of bi- functional agents, such as maleimidobenzoic acid, methyldithioacetic acid, mercaptobenzoic acid, S-pyridyl dithiopropionate, etc.
- the oligopeptides may be linked to proteins to provide site-directed action.
- the oligopeptides may be iinked, particularly by an intracellular cleavable linkage, to antibodies for site directed action.
- conjugation techniques see, for example, U.S. Pat Nos. 3,817,837; 3,853,914;. 3,850,752; 3,905,654; 4,156,081; 4,069,105; and 4,043,989, which are incorporated herein by reference.
- the oligopeptides may also be modified by incorporation into the lumen of vesicles, e.g. Iiposomes, which in turn may be bound to ligands or receptors for direction to particular cells or tissue.
- the therapeutic ordered peptides may be administered topically or parenterally, e.g. by injection at a particular site, including subcutaneously, intraperitoneally, intravasculariy, or the like or transdermally, as by electrotransport. In a preferred embodiment, subcutaneous injection is used to deliver the therapeutic ordered peptide.
- the oligopeptides may also be administered in a sustained release formulation or osmotic pump, to provide a depot of active peptide for slow release over an extended period.
- the therapeutic oligopeptides of this invention may be prepared in accordance with conventional techniques, such as synthesis, recombinant 5 techniques, or the like.
- solid-phase peptide synthesis involves the successive addition of amino acids to create a linear peptide chain (see Merrifield (1963) J. Am. Chem. Soc. 85:2149-2154).
- Production of the peptide by recombinant DNA technology may also be performed.
- This coding sequence is operably connected to suitable control elements for expression, e.g. promoters, terminators, ATG start codon, and the like as known in the art.
- suitable control elements for expression e.g. promoters, terminators, ATG start codon, and the like as known in the art.
- DNA sequences encoding certain functional polypeptide elements such as signal sequences or proteins for targeting the peptide to specific intracellular compartments may be joined to the peptide within the
- This expression construct is introduced into a suitable host cell, and the recombinant protein that is produced is isolated.
- the coding sequence is introduced into the host to be treated for long term therapy, for example by inserting an expression construct into muscle or long-lived hematopoietic cells for therapy.
- the expression vector may be a plasmid, viral
- compositions including retrovirus, adenovirus, etc., and may be introduced by transduction, DNA vaccination, etc.
- Pharmaceutically acceptable salts of the peptides also fall within the scope of the compounds as disclosed herein.
- pharmaceutically acceptable salts as used herein means an inorganic acid addition salt such as
- organic acid addition salt such as acetate, maleate, fumarate, tartrate, and citrate.
- pharmaceutically acceptable metal salts are alkali metal salts such as sodium salt and potassium salt, alkaline earth metal salts such as magnesium salt and calcium salt, aluminum salt, and zinc salt. Examples of pharmaceutically acceptable
- ammonium salts are ammonium salt and tetramethylammonium salt.
- examples of pharmaceutically acceptable organic amine addition salts are salts with morpholine and piperidine.
- examples of pharmaceutically acceptable amino acid addition salts are salts with lysine, glycine, and phenylalanine.
- the subject methods are used to treat individuals suffering from demyelinating autoimmune disease. Diagnosis of suitable patients may utilize a variety of criteria known to those of skill in the art. A quantitative increase in myelin D autoreactive T cells with the capacity to secrete IFN-gamma is associated with the pathogenesis of MS and EAE.
- Such criteria rely on the presence of two attacks at least one month apart, where an attack is a sudden appearance of or worsening of an MS symptom or symptoms which lasts at least 24 hours; and more than one area of damage to central nervous system myelin.
- the damage to myelin must have occurred at more than one point in time and not have been caused by any other disease that can cause demyelination or similar neurologic symptoms.
- MRI magnetic resonance imaging
- Other symptoms include disability in mental, emotional, and language functions, movement and coordination, vision, balance, and the functions of the five senses.
- Evoked potential tests are electrical diagnostic studies which can show if there is a slowing of messages in the various parts of the brain, and may provide evidence of scarring along nerve pathways that is not apparent on a neurologic exam.
- Cerebrospinal fluid usually taken by a spinal tap, may be tested for levels of cytokines, and for the presence of oligoclonal antibody band. The therapeutic effect may be measured in terms of clinical outcome, or may rely on immunological or biochemical tests.
- Suppression of the deleterious T-cell activity can be measured by enumeration of myelin-reactive Th1 cells in spinal fluid, by quantitating the release of cytokines at the sites of lesions, or using other assays for the presence of autoimmune T cells known in the art.
- a reduction in symptoms of a disease such as the damage to neural tissue observed in MS, or the decrease in-the number or severity of attacks of MS suffered by MS patients.
- Damage to neural tissue can be assessed for example by magnetic resonance imaging (MRI) and measurement of the number and severity of lesions visible therein.
- Reduction in MS attack number or severity can be assessed for example by clinical evaluation of patients. Methods for both MRI and clinical evaluation are well-known in the art.
- Various methods for administration may be employed.
- the formulation may be given orally, by inhalation, or may be injected, e.g. intravascular, intra- tumor, subcutaneous, intraperitoneal, intramuscular, etc.
- the dosage of the therapeutic formulation will vary widely, depending upon the nature of the disease, the frequency of administration, the manner of administration, the clearance of the agent from the host, and the like.
- the initial dose may be larger, followed by smaller maintenance doses.
- the dose may be administered as infrequently as weekly or biweekly or monthly or on a schedule determined by the ordinarily skilled physician when administering a vaccine-like therapeutic, or fractionated into smaller doses and administered daily, semi-weekly, etc. to maintain an effective dosage level. In many cases, oral administration will require a higher dose than if administered intravenously.
- the therapeutic ordered peptides of the invention can be incorporated into a variety of formulations for therapeutic administration. More particularly, the .
- complexes can be formulated into pharmaceutical compositions by combination with appropriate, pharmaceutically acceptable carriers or diluents, and may be formulated into preparations in solid, semi-solid, liquid or gaseous forms, such as tablets, capsules, powders, granules, ointments, solutions, suppositories, injections, inhalants, gels, microspheres, and aerosols.
- administration of the peptides can be achieved in various ways, including oral, buccal, rectal, parenteral, intraperitoneal, intradermal, transdermal, intracheal, etc., administration.
- the peptides may be systemic after administration or may be localized by the use of an implant that acts to retain the active dose at the site of implantation.
- the peptides may be administered in the form of their pharmaceutically acceptable salts, or they may also be used alone or in appropriate association, as well as in combination with other pharmaceutically active compounds.
- the following methods and excipients are merely exemplary and are in no way limiting.
- the therapeutic ordered peptides can be used alone or in combination with appropriate additives to make tablets, powders, granules or capsules, for example, with conventional additives, such as lactose, mannitol, corn starch or potato starch; with binders, such as crystalline cellulose, cellulose derivatives, acacia, corn starch or gelatins; with disintegrators, such as corn starch, potato starch or sodium carboxymethylcellulose; with lubricants, such as talc or magnesium stearate; and if desired, with diluents, buffering agents, moistening agents, preservatives and flavoring agents.
- conventional additives such as lactose, mannitol, corn starch or potato starch
- binders such as crystalline cellulose, cellulose derivatives, acacia, corn starch or gelatins
- disintegrators such as corn starch, potato starch or sodium carboxymethylcellulose
- lubricants such as talc or magnesium stearate
- the therapeutic ordered peptides can be formulated into preparations for injections by dissolving, suspending or emulsifying them in an aqueous or nonaqueous solvent, such as vegetable or other similar oils, synthetic aliphatic acid glycerides, esters of higher aliphatic acids or propylene glycol; and if desired, with conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifying agents, stabilizers and preservatives.
- the peptides can be utilized in aerosol formulation to be administered via inhalation.
- the compounds of the present invention can be formulated into pressurized acceptable propellants such as dichlorodifluoromethane, propane, nitrogen and the like.
- the therapeutic ordered peptides can be made into suppositories by mixing with a variety of bases such as emulsifying bases or water-soluble bases.
- the therapeutic ordered peptides of the present invention can be administered rectally via a suppository.
- the suppository can include vehicles such as cocoa butter, carbowaxes and polyethylene glycols, which melt at body temperature, yet are solidified at room temperature.
- Unit dosage forms for oral or rectal administration such as syrups, elixirs, and suspensions may be provided wherein each dosage unit, for example, teaspoonful, tablespoonful, tablet or suppository, contains a predetermined amount of the composition containing one or more compounds of the present invention.
- unit dosage forms for injection or intravenous administration may comprise the compound of the present invention in a composition as a solution in sterile water, normal saline or another pharmaceutically acceptable carrier.
- Implants for sustained release formulations are well-known in the art. Implants are formulated as microspheres, slabs, etc. with biodegradable or " non- biodegradable polymers. For example, polymers of lactic acid and/or glycolic acid form an erodible polymer that is well-tolerated by the host. The implant containing therapeutic ordered peptides is placed in proximity to the site of action, so that the local concentration of active agent is increased relative to the rest of the body.
- unit dosage form refers to physically discrete units suitable as unitary dosages for human and animal subjects, each unit containing a predetermined quantity of peptides of the present invention calculated in an amount sufficient to produce the desired effect in association with a pharmaceutically acceptable diluent, carrier or vehicle.
- the specifications for the novel unit dosage forms of the present invention depend on the particular complex employed and the effect to be achieved, and the pharmacodynamics associated with each complex in the host.
- the pharmaceutically acceptable excipients such as vehicles, adjuvants, carriers or diluents, are readily available to the public.
- compositions of the invention may also contain other therapeutically active agents, e.g. immunomodulators, immunosuppressants, ⁇ -interferon, steroids, statins etc.
- therapeutically active agents e.g. immunomodulators, immunosuppressants, ⁇ -interferon, steroids, statins etc.
- agents capable of additive or synergistic effect in achieving a therapeutic result e.g. where a different or complementary pathway is affected by each of the active agents.
- Immunosuppressants of interest include cyclosporins A and G, FK-506, mycophenylate mofetil, rapamycin, azathioprine, antibodies for plasma membrane proteins associated with graft rejection, such as antibodies to CD4, CD8, CD2, LFA-1, ICAM-1, CD28, and the like; and immunosuppressive oligopeptides derived from MHC molecules.
- Antibacterial, antiviral and antifungal drugs may also be co-formulated in order to minimize the effects of immunosuppression.
- the therapeutic ordered peptides will generally be administered in dosages of 0.01 mg to 500 mg V/kg body weight per day, e.g. about 20 mg/day for an average person.
- the range is broad, since in general the efficacy of a therapeutic effect for different mammals varies widely with doses typically being 20, 30 or even 40 times smaller (per unit body weight) in man than in the rat. Similarly the mode of administration can have a large effect on dosage. Thus for example oral dosages in the rat may be ten times the injection dose. A typical dosage may be one injection daily.
- dose levels can vary as a function of the specific compound, the severity of the symptoms and the susceptibility of the subject to side effects. Some of the specific peptides are more potent than others.
- Preferred dosages for a given complex are readily determinable by those of skill in the art by a variety of means. A preferred means is to measure the physiological potency of a given compound.
- MBP myelin basic protein
- the MBP p87-99 peptide was immunodominant in all cases and it inhibited autoantibody binding to MBP by more than 95%. Residues contributing to autoantibody binding were located in a 10-amino acid segment p86-95 ( ⁇ SEQ ID NO:9 ⁇ WHFFKNIVT) that also contained the MHC -T cell receptor contact residues for T cells recognizing MBP in the context of DRB1*1501 and DQB1 * 0602. In the epitope center, the same residues, ⁇ SEQ ID NO:10 ⁇ VHFFK, were important for T cell binding and MHC recognition.
- Peptides For immunization and disease reversal, peptides were synthesized on a peptide synthesizer (model 9050: MilliGen, Burlington, MA) by standard 9-fluorenylmethoxycarbonyl chemistry. Peptides were purified by HPLC. Structure was confirmed by amino acid analysis and mass spectroscopy. Peptides used for the experiments were: ⁇ SEQ ID NO:11 ⁇ ENPWHFFKNIVTPR (MBPp85-99), ⁇ SEQ ID NO:4 ⁇ EYYKEYYKEYK, ⁇ SEQ ID NO:5 ⁇ KYYKYYKYYKYYYYYYY.
- Synthetic peptide MBPp85-99 was dissolved in PBS to a concentration of 2 mg/ml and emulsified with and equal volume of Incomplete Freund's Adjuvant (I FA), supplemented with 4 mg/ml heat-killed Mycobacterium tuberculosis H37Ra (Difco Laboratories, Detroit, Ml). Rats were injected subcutaneously with 0.1 ml of the peptide emulsion. Experimental animals were scored as follows: 0, no clinical disease; 1 , tail weakness or paralysis; 2, hind limb weakness; 3, hind limb paralysis; 4, forelimb weakness or paralysis; 5, moribund or dead animal.
- ordered peptide treatment was administered in a mouse model of ongoing-EAE (Figure 2) at days 17, 29, and 36 after EAE induction with intravenous, intra-peritoneal or subcutaneous administration of EYYKEYYKEYYK (therapeutic ordered peptide) or Copaxone at two different dosages, 0.5 mg per mouse or 0.05 mg per mouse. Both the therapeutic ordered peptide and Copaxone were dissolved in mannitol. Results are expressed as relapse rates per mouse, and show that the therapeutic ordered peptide and Copaxone reduce relapse rates similarly.
- APCs antigen presenting cells
- CD52 antibodies specific for T cells
- CD45R macrophages
- NKR-P1A antibodies specific for T cells
- T cells CD52
- CD45R macrophages
- NKR-P1A NK cells
- Therapeutic ordered peptides were used as inhibitors are added to the wells at different concentrations (ranging from 0.01 to 0.24 mM) in a volume of 0.05 ml.
- MOG therapeutic ordered peptide of, YREYEYE either singly or in multimer form is prepared and tested in rat and mouse models of EAE.
- a modified therapeutic ordered peptide D-alaYREYEYE is tested in both rat and mouse models of EAE.
- the PLP therapeutic ordered peptide, YGKELGEY either singly or in multimer form is prepared and tested in rat and mouse models of EAE.
- a modified therapeutic ordered peptide D-ala YGKELGEY is tested in both rat and mouse models of EAE.
- Example 6 Administraion of MBP and MOG Therapeutic Ordered Peptides in Combination and Their Effect in EAE
- Cop1 caused an increase in these two Th2 cytokines as expected, and OVA did not cause an increase in these cytokines also as expected.
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JP2006534217A JP2007507541A (en) | 2003-10-03 | 2004-10-01 | Method of treating demyelinating autoimmune diseases with modified regular peptides |
CA002540890A CA2540890A1 (en) | 2003-10-03 | 2004-10-01 | Treatment of demyelinating autoimmune disease with modified ordered peptides |
EP04809854A EP1677737A4 (en) | 2003-10-03 | 2004-10-01 | Treatment of demyelinating autoimmune disease with modified ordered peptides |
US10/589,067 US7585843B2 (en) | 2003-10-03 | 2004-10-01 | Treatment of demyelinating autoimmune disease with modified ordered peptides |
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Cited By (4)
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WO2007120834A2 (en) | 2006-04-13 | 2007-10-25 | Peptimmune, Inc. | Methods for designing and synthesizing directed sequence polymer compositions via the directed expansion of epitope permeability |
WO2010079464A1 (en) | 2009-01-12 | 2010-07-15 | Novartis Ag | Cna_b domain antigens in vaccines against gram positive bacteria |
EP2586460A1 (en) | 2007-10-16 | 2013-05-01 | Peptimmune, Inc. | Method for designing and preparing vaccines comprising directed sequence polymer composition via the directed expansion of epitopes |
US8546532B2 (en) | 2008-04-17 | 2013-10-01 | Declion Pharmaceuticals, Inc. | Synthesis of directed sequence polymer compositions and antibodies thereof for the treatment of protein conformational disorders |
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JP4750419B2 (en) * | 2002-11-21 | 2011-08-17 | ベイヒル セラピューティクス インコーポレーティッド | Methods and immunomodulatory nucleic acid compositions for preventing and treating diseases |
AU2005308396B2 (en) * | 2004-11-29 | 2011-06-09 | Yeda Research And Development Co. Ltd. | Induction of neurogenesis and stem cell therapy in combination with Copolymer 1 |
CA2667637C (en) | 2006-10-31 | 2021-04-20 | East Carolina University | Fusion proteins comprising an anti-inflammatory cytokine and an antigen for treatment of immune disorders |
EP3348275A3 (en) | 2009-03-31 | 2018-10-24 | East Carolina University | Cytokines and neuroantigens for treatment of immune disorders |
US8920373B2 (en) | 2009-07-15 | 2014-12-30 | Teva Pharmaceutical Industries, Ltd. | Reduced volume formulation of glatiramer acetate and methods of administration |
DK2275086T3 (en) * | 2009-07-15 | 2012-07-09 | Teva Pharma | Reduced volume formulation of glatiramer acetate and methods of administration thereof |
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AU5360796A (en) * | 1995-03-09 | 1996-10-02 | Neurocrine Biosciences, Inc. | Peptide analogues of human myelin basic protein useful in treating multiple sclerosis |
US6531130B1 (en) * | 1999-07-06 | 2003-03-11 | The Board Of Trustees Of The Leland Stanford University | Treatment of demyelinating autoimmune disease with ordered peptides |
WO2001093893A2 (en) * | 2000-06-07 | 2001-12-13 | Yeda Research And Development Co. Ltd. | The use of copolymer 1 and related peptides and polypeptides and t cells treated therewith for neuroprotective therapy |
WO2002059143A2 (en) * | 2001-01-24 | 2002-08-01 | President And Fellows Of Harvard College | Therapeutic peptides for demyelinating conditions |
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- 2004-10-01 WO PCT/US2004/032598 patent/WO2005032482A2/en active Application Filing
- 2004-10-01 US US10/589,067 patent/US7585843B2/en not_active Expired - Fee Related
- 2004-10-01 EP EP04809854A patent/EP1677737A4/en not_active Withdrawn
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Cited By (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2007120834A2 (en) | 2006-04-13 | 2007-10-25 | Peptimmune, Inc. | Methods for designing and synthesizing directed sequence polymer compositions via the directed expansion of epitope permeability |
US8378072B2 (en) | 2006-04-13 | 2013-02-19 | Declion Pharmaceuticals, Inc. | Methods for designing and synthesizing directed sequence polymer compositions via the directed expansion of epitope permeability |
EP2586460A1 (en) | 2007-10-16 | 2013-05-01 | Peptimmune, Inc. | Method for designing and preparing vaccines comprising directed sequence polymer composition via the directed expansion of epitopes |
US8546532B2 (en) | 2008-04-17 | 2013-10-01 | Declion Pharmaceuticals, Inc. | Synthesis of directed sequence polymer compositions and antibodies thereof for the treatment of protein conformational disorders |
WO2010079464A1 (en) | 2009-01-12 | 2010-07-15 | Novartis Ag | Cna_b domain antigens in vaccines against gram positive bacteria |
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EP1677737A2 (en) | 2006-07-12 |
WO2005032482A8 (en) | 2008-03-06 |
CA2540890A1 (en) | 2005-04-14 |
JP2007507541A (en) | 2007-03-29 |
US20070275899A1 (en) | 2007-11-29 |
US7585843B2 (en) | 2009-09-08 |
WO2005032482A3 (en) | 2009-06-25 |
EP1677737A4 (en) | 2008-04-16 |
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