TWI327149B - Peptides and related compounds having thrombopoietic activity - Google Patents

Description

1327149 IX. Description of the Invention: TECHNICAL FIELD OF THE INVENTION The present invention is generally related to peptides and related compounds having platelet-forming activity. The compounds of the invention may be used to increase the production of platelets or platelet precursors (e.g., megakaryocytes) in a mammal. [Previously as technology] The present invention relates to a compound having a ability to stimulate the production of platelets and precursor cells thereof, such as megakaryocytes, in vitro and in vivo, particularly peptides. Two proteins known to have platelet-forming activity: thrombopoietin (TPO) and megakaryocyte growth factor (MGDF) are set forth below. Colonization of endogenous blood-small plateogen (TPO) (Lok et al., Nature 369: 568-571 (1994); Bartley et al., Cell 77: 1117-1124 (1994); Kuter et al. , Proc. Natl. Acad. Sci. USA 91:11104-11 108 (1994); de Sauvage et al., Nature 369:533-538 (1994); Kato et al., Journal of Biochemistry 119:229-236 ( 1995); Chang et al., Journal of Biological Chemistry 270:511-514 (1995) ) has rapidly increased our understanding of megakaryocyte production (megakaryocyte production) and platelet production (platelet production). Endogenous humans TPO, a 60 to 70 kDa glycosylated protein produced in the liver and kidney, is composed of 332 amino acids (Bartley et al., Cell 77: 1117-1124 (1994); Chang et al., Journal of Biological Chemistry 270: 511-514 (1995)). The protein is highly conserved among different species and is at the amino terminus (amino acid 1 to 172) (Bartley et al., 114552.doc 1327149)

Cell 77: 1117-1124 (1994)) has a homology of 23 〇/〇 with human erythropoietin (Gurney et al., Blood 85: 981-988 (1995)). Endogenous TPO displays all of the features of a key biomodulator with platelet production. Its activity in vitro includes from purified murine hematopoietic stem cells (Zeigler et al., Blood 84: 4045-4052 (1994)) and human CD34+ cells (1^〇1<;6 (ugly 1., he 11 ^ 369:568-571 (1994); Rasko et al., Stem Cells 15:33-42 (1997)) Specific induction of both megakaryocyte populations, with increased ploidy of megakaryocytes Generation (Broudy et al., Blood 85: 402-413 (1995) ) ' and terminal megakaryocyte maturation and induction of platelet production (Zeigler et al_, Blood 84: 4045-4052 (1994); Choi et al, Blood 85 :402-413 (1995). Conversely, synthetic antisense oligodeoxynucleotides against the TPO receptor (c-mpi) significantly inhibited the ability of megakaryocyte protoplasts to colonize ( Methia et al., Blood 82: 1395-1401 (1993). In addition, c-mpl knockout mice have severe thrombocytopenia and lack megakaryocytes (Alexander et al., Blood 87: 2162-2170 (1996) )). Recombinant human MGDF (rHuMGDF, Amgen Inc., Thousand

Oaks, CA) is another TPO-associated platelet-producing peptide. It is produced by a plastid-transformed Escherichia coli (E. coU) containing a cDNA encoding a truncated protein covering the amino-terminal receptor-binding functional site of human TPO (Ulich et al., Blood 86:971) -976 (1995)). After the polypeptide is extracted, refolded and purified, the poly[ethylene glycol] (PEG) moiety is covalently bonded to the amine end. The resulting molecule is referred to herein as PEG-114552.doc 1327149 rHuMGDF or simply MGDF. Various studies using animal models (111丨(^,1\11.61&1.,81〇〇£186: 971-976(1995); Hokom, Μ.M. et al.} Blood 86:4486-4492 ( 1995)) clearly confirms the efficacy of TPO and MGDF in bone marrow transplantation and in the treatment of thrombocytopenia, a condition usually caused by chemotherapy or radiotherapy. Preliminary data in humans confirm that MGDF is used to increase various coagulants. Use of platelet counts in (Basser et al., Lancet 348-1279-81 (1996); Kato et al., Journal of Biochemistry 1 19:229-236 (1995); Ulich et al., Blood 86:971 -976 (1995)) MGDF can be used to enhance the platelet administration procedure because administration of MGDF enhances platelet counts in the circulatory system of healthy platelet donors by approximately three times the original value. TPO and MGDF are transmitted through c-mpl They bind to each other and the c-mpl receptor is mainly expressed on the surface of certain hematopoietic cells, such as megakaryocytes, platelets, CD34+ cells, and primitive protoplasts (Debili, N. et al., Blood 8 5.391 -401 (1995); de Sauvage, FJ et al., Nature 369: 533-538 (1994); Bartley, TD, et al., Cell 77:1117- 1124 (1994); Lok, S. et al" Nature 369:565-8 (1994). Like the majority of interleukin and protein hormone receptors, the c mpl receptor belongs to the Class I Interleukin Receptor Superfamily (Vig0n, I. et al) PfQe Natl_Acad. Sci. USA 89:5 640-5644 (1992). Activation of such receptors involves ligand-induced receptors Dimerization (h〇ni〇dimerization), which in turn triggers a cascade of transduction events. In general, the interaction between a protein ligand and its receptor usually occurs at the larger interface of 114552.doc 1327149. As evidenced by the fact that human growth hormone is linked to its receptors, only some of the key residues on the interface actually contribute most of the binding energy (Clackson, T. et al., Science).

267:3 83-3 86 (1995)). This fact, which is used only with the body of the remaining protein ligands to display the correct epitope in the correct topology (epjt〇pes), can be used to find active ligands with far smaller sizes. Accordingly, only molecules with a length of "peptide" (e.g., 2 to 80 amino acids) can bind to a receptor protein of a given large protein ligand. Such a peptide can mimic a large protein ligand. Activity, or through competitive binding, inhibits the biological activity of large protein ligands, and is often referred to as "peptide mimetic" or "simulated peptide". Phage display peptide libraries have emerged as identifying such peptides A powerful technique for simulating objects. See, for example, Scott, jK et al, Scienee 249 386 (1990); Devlin, JJ et al., Science 249: 404 (1990);

US Patent No. 5,223,4-9, issued on June 29; US Patent No. 5,733,731 issued March 3, 1989; US Patent No. 5,498,530 issued on March 12, 1996; issued on 1995 U.S. Patent No. 5,432, No. 18; U.S. Patent No. mg, (6) issued on August 16, 1994; U.S. Patent No. 19, issued on July 13, 1999, WO 96/side 7; And W_98/15833 published on April 16th (all of the above documents are incorporated herein by reference in their entirety). In such a library, the 'random peptide sequence is passed through the shell egg with the filamentous bacillus The fusion of the mass is shown. Typically, the peptide displayed is directed against the antibody-immobilized extracellular work of the receptor, and the retained phage is permeable to affinity purification and reproduction. The cycle is enriched. Optimal 114552.doc 1327149 The binding peptide can be sequenced to identify key residues within one or more structurally related peptide families. See, for example, Cwiria, et al., (1997) , Science 276; 1696-9. The peptide sequence can also be inferred which residues can be scanned via alanine. Mutations at the DNA level or at the DNA level are safely replaced. Mutations can be generated and screened to further optimize the sequence of the optimal binder. Lowman (1997),

Ann. Rev. Biophys. Biomol. Struct. 26:401-24 0 Structural analysis of protein-protein interactions can also be used to infer peptides that mimic large protein ligand binding activities. In such an analysis, the crystal structure can be inferred that the bulk and relative orientation of the key residues of the large protein ligand can thus be set to § eleven peptides. See, for example, Takasaki, et al" (1997), Nature Biotech, 15: 1266-70. These analytical methods can also be used to study the interaction between streptozocin and phage display peptides selected, which can be further speculated Modification to increase binding affinity. There are other methods in the peptide study that compete with the phage display method. The peptide library can be fused to the carboxy terminus of the lac repressor (repress〇r) and expressed in the colon of the large intestine. Other E. coli-based methods can be displayed on the outer membrane of cells by fusion with a peptide glycan-associated lipoprotein (PAL). In the following, these and related methods are collectively referred to as E. coli display methods. In other methods, translation of the random RNA is halted prior to ribosome release, resulting in a multi-peptide library to which the associated RNA is still attached. Hereinafter, this and related methods are collectively referred to as "ribosome display method" and other methods employ a peptide attached thereto; such as PRO fusion technology, Phyl〇s, Inc. See, for example, (4) Coffee & ^Szostak (1997) , Free. Natl. Acad. Sci. USA, 94:12297-3〇” In the following, this and related methods are collectively referred to as "rna_peptide screening 114552.doc •10· 1327149.” Chemical derivation has been developed. The peptide library, Huangzhong ^ Zhongsheng peptide is immobilized on stable, non-biological materials, such as polyethylene rods or solvent permeable resins. Another chemical derivative peptide library is photolithographically fixed by scanning. The peptides on the glass slides. Hereinafter, this and related methods are collectively referred to as "chemical-peptide screening". An advantage of chemistry-peptidic screening is that it can use hydrazine-amino acids and other non-natural analogs, as well as non-peptide components. Both biological and chemical methods are available at Wells & Lowman (1992), Curr. 〇pin. Biotechnol, 3:355-62. Conceptually, a peptide mimetic of any protein can be found using phage display, rna·peptide screening, and other methods described above. Through the use of phage display peptide library technology, a small peptide molecule that can be used as an agonist of the cucurbit receptor... (Cwirla, SE et al, Science 276: 1696-1699 (1997)) ^ In this type of study, random small The peptide sequence is shown as a fusion with the filamentous 嗟I] in vitro layer protein, and the antibody against the c_mpi receptor is lysed by the affinity of the extracellular functional site, and the retention of the retained bacterium is used for the first Round-Affinity Purification β This binding selection and re-proliferation method was repeated multiple times to enrich the tighter pool of binders. As a result, two families of c-mpl-binding peptides were first identified, and the two were independent of each other in their sequences. A library of mutations is then generated to further optimize the optimal binder, which ultimately leads to the isolation of the very active peptide with IC5() = 2 nM and its EC50-4OO nM (Cwirla, SE et al., Science 276: 1696-1699) (1997)). The 14-residue TPO mimetic peptide does not have a significant sequence homology with TPO or MGDF. The structure of this particular TPO mimetic peptide (TMP) compound is as follows: 114552.doc 1327149 lie Glu Gly Pro Thr Leu Arg Gin Trp Leu Ala Ala Arg Ala (SEQ ID NO: 1) or a single letter amino acid abbreviation denoted IEGPTLRQWLAARA .

In a previous similar study of EPO mimetic peptides, an EPO mimetic peptide (EMP) was discovered using the same technique (Wrighton, NC et al., Science, 273: 458-463 (1996)) and the peptide was found. It binds to the EPO receptor (EPOR) and acts as a dimer. According to the χ-ray crystallographic data, the (ligand) 2/(acceptor) 2 complex thus formed has (^

Weighing (Livnah, 0· et al., Science 273: 464-471 (1996)). Based on this structural information, a covalently linked EMP dimer was designed in which the C-termini of two EMP monomers were crosslinked with elastic spacers and found to have greatly enhanced binding and in vitro/in vivo biology. Activity (WHghton, NC, et al., Nature Biotechnology 15: 1261-1265 (1997)). A similar C-terminal dimerization method was applied to the TP〇 analog peptide (TMp). (Cwirla, S_E. et al., Science 276: 1696-1699 (1997)). Find

The C-terminal ligation dimer (c_c linkage) specific for a particular TPO mimetic peptide has improved 0.5 nM binding affinity in cell proliferation assays, and increased in vitro activity (EC5Q = 0.1 nM) (Cwirla, SE et al, 276: 1696-1699 (1997)). The utilization of recombinant proteins for therapeutic use leads to advances in protein upgrading to enhance or improve the properties of such proteins f as agents. Such modifications provide enhanced protein protection and reduced degradation by reducing or eliminating proteolysis. Other advantages include 'in some circumstances, increasing the stability of therapeutic proteins, circulating dark shackles 4u ~. 114552.doc 1327149 The quality of the 〇平 0 two texts as early 卩 1 * 311. 13,? Hey. 113〇11〇1'〇评1;11?3(^〇3:4-l〇(May 1992) (published by Mediscript,London,UK). Useful modifications of protein therapeutics include bonding to polymers Such modifications as polyethylene glycol (PEG) and dextran are discussed in detail in the following patent applications: "Modified Peptides as Therapeutic Agents", U.S. Patent Application Serial No. 09/428,082, PCT Application WO 00 /24782, which is incorporated herein by reference in its entirety. Another such modification is the use of immunoglobulin molecules. The antibody comprises two functionally independent moieties; a variable functional moiety, called, Fab ", which binds to the antigen, a fixed functional site is called "Fc", which provides a long plasma half-life of the Fc portion of the immunoglobulin that binds to effector functions such as complement or sputum cells. And Fab is shorter. (Cap〇n, et al

Nature 337, 525-53 1 (1989)). Fc functional sites have been used to constitute therapeutic protein products to provide longer half-life or added functions, such as F c receptor binding, protein a binding, complement fixation, and placental transplantation, all of which are present in immunoglobulins. in. (Capon, et al., Nature 337:525-53 1 (1989)). Such as,

The Fc region of the IgGl antibody is fused to CD30-L, which binds to CD3 purinoceptors expressed on Hodgkin disease tumor cells, undifferentiated lymphoma cells, τ-cell leukemia cells, and other malignant cell types. See U.S. Patent No. 5,480,981. IL-10, an anti-inflammatory and anti-rejection agent, is fused to the fusion mouse F c 2 a to increase the short-cycle half-life of interleukins (Zheng, X. et al., The J〇urnai 〇f iminun〇i〇 Gy, 154:5 590-5600 (1995)). There have also been multiple studies evaluating the use of tumor necrosis 114552.doc -13 - 1327149 factor receptors linked to human IgG 1 Fc protein for the treatment of patients with septic shock (corpse 151^1*, (:.61 pressure 1) .1^.£1^1.厂1^£1.,334:1697-1702 (1996); Van Zee, K. et al., The Journal of

Immunology, 156: 2221-2230 (1996)). There are also therapeutic proteins that fuse Fc to the CD4 receptor to produce AIDS. See Capon et al., Nature, 337: 525-531 (1989) » In addition, there is also the fusion of interleukin 2 to the Fc portion of IgG 1 or IgG3. In order to overcome the short half-life of interleukin 2 and its system. See Harvill et al., Immunotechnology, 1: 95-105 (1995). A specific platelet-causing compound, generally a peptide, having a cascade (ie N_ to c_end) orientation and a cascade of peptide dimers linked at its N-terminus is disclosed in published PCT application WO 00/24770. For carrier molecules, such as linear polymers 'oligosaccharides or Fc groups. There is a need to propose additional compounds which have excellent biological activity to stimulate platelet production (thrombotic activity) and/or production of platelet precursor cells, particularly megakaryocytes (megakaryocyte production activity). There is also a need to develop a compound which exhibits platelet-forming activity and which has excellent therapeutic properties such as long-lived period. Such compounds exhibit advantageous properties related to the production, separation, 'deuterated organism' tongueiness, stability and cycle time, and the like. The present invention proposes novel compounds and related aspects having such activities. SUMMARY OF THE INVENTION The present invention relates to a therapeutic compound that binds to a receptor (hereinafter referred to as !npl steroid). More particularly, the present invention proposes a group of compounds that exhibit enhanced binding to the c_mpl receptor and/or Transmits, ie, live H4552.doc 1327149, the ability to trigger a transmembrane signal by the 'c-mpl receptor, which is identical to the receptor for the activity of endogenous thrombopoietin (TPO). Thus, the compounds of the invention have Excellent platelet production activity, i.e., stimulation of platelet production and/or megakaryocyte production activity in vivo and in vitro, i.e., stimulation of platelet precursor production in vivo and in vitro. In addition, certain compounds of the present invention also perform well. Therapeutic properties, such as improved half-life in plasma, biological activity, and circulating time in vivo. In one aspect, the invention provides a compound that binds to an mpl receptor, the compound comprising the following sequence: X1-X2- X3-X4-GPTL-X9-X10.WL-X13-X14.X15-X16-X17-X18 wherein X1-X4, X9-X10 and Χ13_Χ18 are each independently an amino acid as defined herein, and wherein the compound The binding affinity and/or biological activity of the substance to the mpl receptor is greater than that of the following sequence: IEGPTLRQWLAARA. In another aspect, the invention provides a compound that binds to the c_mpl receptor, which has the following sequence: X1-X2 - REGPTLRQWL-X13-WRR-X17-X18 wherein XI, X2, X13, X17 and χ18 are each independently an amino acid. In another aspect, the invention provides a compound which binds to an mpl receptor, comprising And a sequence selected from the group consisting of SEQ ID NO 2 to SEQ ID NO 30. In another Φ, the invention is a dimer or polymeric Π 4552.doc 1327149 body of a compound comprising one selected from A sequence comprising the group of SEQ ID NO 2 to SEQ ID NO 30. In another aspect, the invention provides a composition of matter that binds to an mpl receptor, which has the following general formula (LNI) 1-- (TMPl)a--(LN2)m--(TMP2)b--(LN3)n--(TMP3)c-- (LN4)〇--(TMP4)d where TMP1, TMP2, TMP3 and TMP4 are each Independently selected from the group consisting of the TMPs disclosed herein; LN1, LN2, LN3, and LN4 are each independently a linker; a, b^, and d Each of which is independently an integer of 11; 1, m, n and 〇 are each independently an integer of 〇 to 2〇. In yet another aspect, the present invention provides a composition of matter that binds to an mpl receptor, Has the general formula: (Vl)v--(LNl)1-(TMPl)a..(LN2)m--(TMP2)b--(LN3)n--(TMP3)c--(LN4)0- -(XjvIP4)d--(V2)w VI and V2 are each independently a mediator, and 乂 and w are each independently an integer from 0 to 1. The compounds of the invention can be prepared by standard synthetic methods, recombinant DNA techniques or other methods of preparing peptides and fusion proteins. The compound of the present invention encompassing the non-peptide portion can be synthesized from a standard organic chemical reaction in addition to a standard dermatological reaction. The compound of the present invention can be used for treatment or prophylaxis by formulating it with a suitable pharmaceutical carrier material and administering it to a human, such as a human (or other mammal) in need thereof. The activity of the vehicle/linker peptide may be comparable to the natural ligand mimicked by the peptide, here compared to the sputum plate 114552.doc •16· 1327149 pheromone, or even larger. In another aspect, the invention provides a method of treating thrombocytopenia. In other aspects, the invention provides methods of administering megakaryocytes or platelets, and methods of producing the compounds described herein. In another aspect, the invention also provides related pharmaceutical compositions. In another aspect, the invention features a polynucleotide encoding a composition of matter disclosed herein, including a representation vector of the polynucleotide, a host cell of the present vector. This table • [Embodiment] I. Definition of terms The definitions of terms used in this specification are as follows, unless there are other restrictions in the case of ambiguity. "A" peptide refers to a molecule having about 2 to 80 amino acids, preferably 3 to 40 amino acids, and the exemplary peptide can be randomly produced by any of the methods described herein, such as Peptide library methods (such as phage display libraries) are produced by chemical man-made methods derived from protein digestion or similar. ® Terminology "Randomization" When used with a peptide sequence, it refers to a completely random sequence (eg selected from The residue of one or more of the naturally occurring molecules in the phage display or RNA-peptide screening method is replaced by an amino acid residue that is not present at that position within the naturally occurring molecule. Exemplary methods for generating and identifying randomized peptide sequences include phage display, E. coli display, ribosome display, RNA-peptide screening, chemical screening, and the like. The term "dimer" is used In the peptide, it is meant that the two peptide peptides have two peptide chains, the I2552.doc 1327149 sub-two peptide chains are covalently or non-covalently bound, with or without a linking agent. The two peptides in the peptide dimer are C-terminal to N-terminal linkages and may also be referred to as "cascade repeats" or "cascade dimers". The two peptides in the peptide dimer are c_ to C-terminal' or N- to N-terminal linkages' may also be referred to as "parallel repeats" or••parallel dimers". The term "multimer" as used in a peptide, refers to a molecule having three or more peptide chains, which are covalent, non-covalent, or both covalent and non-covalent. Covalent bond interactions, with or without linkage agents. The terms "derived" and "derivative" or "derived" relate to the following methods and resulting compounds, of which (1) compounds a cyclic moiety; such as cross-linking between cysteamine residues in a compound; (2) a compound cross-linked or having a cross-linking moiety; such as a compound having a cysteamine-based residue, thereby Or forming a cross-linked dimer in vivo; (3) one or more peptidyl linkages are replaced by non-peptidyl linkages; (4) N_-end is via _nrr1, nrc(o)ri, -NRC(0)0Rl, _NRS(0)2Rl, _NHc(〇)NHR, butyl quinone imine, or substituted or unsubstituted methoxycarbonyl-NH-, wherein R and R1 are rings a substituent 'defined as defined below; (1). The terminal is substituted with -C(0)R2 or _Nr3R4, wherein R2, r^r4 are as defined below; and (6) individual amino acid moiety in the compound It is modified by treatment with a drug that reacts with the selected side chain or terminal residues. The derivative is further described below. "The term"platelet-forming analog peptide, "τρ〇 analogue peptide""ΤΜΡ" refers to a peptide that binds to the mpl receptor and/or has platelet-forming activity, and can also stimulate & platelets or platelets before or in vitro 114552.doc -18 · 1327149 The ability to produce, including but not limited to, megakaryocytes. The term "mpl_binding functional site" refers to any amino acid sequence that binds to the mpl receptor and includes naturally occurring sequences or randomized sequences. The exemplary mpl-binding functional site can be identified or derivatized by phage display or other methods described herein. The term "mpl receptor antagonist" refers to a molecule that binds to an mpi receptor and that increases or decreases one or more assay parameters' such as endogenous thrombopoietin (eTPO), a native mpl receptor ligand. The term "includes" indicates that a compound may include an additional amino acid at one or both of the N-terminus or C-terminus of a given sequence. Of course, these additional amino acids should not significantly interfere with the activity of the compound. Further, a physiologically acceptable salt of the compound of the present invention is also encompassed therein. The term 'physiologically acceptable salt" refers to any salt known or later found to be pharmaceutically acceptable. Some specific examples are: acetates, trifluoroacetates, hydrogen hydrides such as hydrochlorides and hydrobromides, sulfates, citrates, tartrates, glycolates and oxalates. The term "vehicle" refers to a molecule that prevents degradation of therapeutic proteins and/or prolongs half-life, reduces toxicity, reduces immunogenicity, and/or increases biological activity. Exemplary vehicles include Fc functional sites (which are preferred) and linear polymers (such as polyethylene glycol (peg), polylysine, dextran, etc.), branched polymers (see, for example, 1981) Issued on September 15

U.S. Patent No. 4,289,872 to Denkenwalter et al.; U.S. Patent No. 5,229,490 issued to Tam on July 2, 1993; WO 93/21259 to Frechet et al., issued Oct. 28, 1993, and lipid 'cholesterol (eg, 114552.doc •19· 1327149 steroids), enzymes or oligosaccharides (eg glucomannans), any natural or synthetic protein, polypeptide or peptide that binds to salvage receptors; albumin, including human serum albumin (HSA), leucine zipper domain and other such proteins and protein fragments. The term "native Fc" refers to a molecule or sequence comprising a sequence of non-antigen-binding fragments derived from the digestion of the entire antibody, whether in monomeric or multimeric form. The original immunoglobulin source of native Fc is preferably a human source. And may be any immunoglobulin, although Ig〇1 and IgG2 are preferred. The natural fcs is formed by the monomeric peptide, the monomeric peptide is transmitted through a covalent bond (ie, a disulfide bond) and non-co- The valency bond binds to a dimeric or multimeric form. The number of intermolecular disulfide bonds between the haplotype subunits of the molecule is between 丨 and 4, depending on the class (eg IgG ' IgA ' IgE Or subclass (eg IgG1, IgG2, IgG3, IgAl, IgGA2). An example of a native Fc is a disulfide-bonded dimer derived from papain digesters of IgG (see EUis〇n et al (1982)) , Nucleic Acids Res. ι〇: 4〇71_9). The term natural F c ′′ as used herein is a generic term for monomers, monomers and multimers. The term "Fc variant" refers to a molecule or sequence that is modified from a native Fc but still includes a binding site for a salvage receptor (FcRn). The exemplary Fc variants, and interactions with salvage receptors, are described in the International Application, WO 97/3463, filed on Sep. 25, 1997, and WO 96/32, the entire disclosure of which is incorporated herein by reference. Thus, the term nFc variant "includes a molecule or sequence derived from a non-human native Fc. In addition, native Fc includes removable sites because the structural properties or biological activities provided by them are not the fusion molecules of the present invention. Thus, the term "Fc variant" includes a molecule or sequence lacking one or more of the natural H4552.doc-20. 1327149 residues or residues that affect or involve (1) disulfide bonds. Formation, (2) incompatibility with selected host cells, (3) N-terminal heterogeneity after expression in selected host cells '(4) glycosylation, (5) interaction with complement The role '(6) binds to the Fc receptor rather than the salvage receptor, or antibody-dependent cellular cytotoxicity (ADCC). The term "Fc functional site" includes natural and Fc variant molecules and sequences as defined above. With respect to Fc variants and native Fes, the term "Fc functional site" includes both monomeric or multimeric forms of the molecule 'whether it is digested from the entire antibody or formed by other methods. The term "dimerization A body, such as a molecule for an Fc functional site or a Fe-containing functional site, refers to a molecule having a multi-peptide chain with two covalent bonds or non-covalent bonds. The term "multimer" as used in the Fc functional site or a molecule containing a functional site means having two or more covalent bonds, non-covalent bonds or covalent bonds and non-covalent bonds. Molecules of multi-peptide chains. IgG molecules typically form dimers, IgM, pentamers; IgD, dimers; and IgA, monomeric, dimeric, trimer or tetramer. Body formation can be utilized

The sequence of the native Ig source of Fc and the resulting activity are obtained by derivatization (as defined herein) such native Fc. The term "peptide antibody" or "peptide antibody group" refers to a molecule comprising a portion of the antibody FC# binding to at least one peptide. Such a peptide antibody can be a poly- or dimer or a fragment thereof, which can be derived. II. Structure of the Compound In general, the present invention proposes a compound which binds to and/or modulates the biological activity of the mpl receptor. More particularly, the present invention proposes a group of compounds which can bind to the mpl receptor and/or permeate by the mpl receptor and/or permeate, ie, the active Kmpl receptor, which triggers transmembrane signaling, which mediates endogenous thrombopoietin (TPO). The active receptors are the same. Thus, the compounds of the invention have platelet-producing activity, i.e., the ability to stimulate platelet production in vivo and in vitro and/or have megakaryocyte-producing activity', i.e., the ability to stimulate platelet precursors, including megakaryocytes, to be produced in vivo and in vitro. Briefly, the compounds of the invention include one or more peptides of the formula: 1 : X1-X2-X3-X4-GPTL-X9-X10-WL-X13-X14-X15-X16-X17- X18; wherein X1-X4, X9-X10 and X13-X18 are each independently an amino acid. In another composition of matter prepared according to the present invention, the compounds may comprise one or more peptides having the sequence of Formula I, which are linked to each other or otherwise linked, such as to become dimers or Polymer. In another composition of matter prepared according to the invention, the compounds may comprise one or more peptides of the formula I which are linked at the N-terminus or c-terminus of the peptide or otherwise linked Above a medium. Any of these peptides may be cascaded (i.e., sequentially, N to C), or parallel (i.e., N-terminal to N-terminally or C-terminal to C-terminal), with or without a linking agent. Peptide. The compounds of the invention include TPO mimetic peptides, either alone or in combination with other TMPs, e.g., in the form of dimers or multimers. The TMPs of the present invention comprise the following sequences: 1 : X1-X2-X3-X4-G-P-T-L-X9-X10-W-L-X13-X14-X15-X16-X17-X18; 114552.doc -22- 1327149

Wherein Χ1·Χ4, X9-X10 and X13-X18 are each independently an amino acid. Preferred amino acid residues of the above sequences are further defined in Table 1 below. Table 1 - Preferred amino acid residue positions Amino acid residues XI A'V, W'M'G'Y, C, Q'E'R'h X2 a, v, l, i, g, s, c X3 L, I, P, W'G, S, D, K, R X4 L, G, Q, D, E, H X9 K, R X10 Q, E X13 A, V, L, S, Q, E, R X14 A, W, T, Y, C, Q X15 V, L, G, Y, R X16 A, L, F, G, R X17 A, V, L, M, G, C, Q, N X18 a, v, p, m, f, g, c, q, k Even better, the purine sequence of the present invention is a peptide having the following sequence: I : X1-X2-X3-X4-GPTL-X9 -X10-WL-X13-X14-X15-Χ16-Χ17-Χ18; wherein Χ1-Χ4, Χ9-Χ10 and Χ13-Χ18 are each independently an amino acid, and wherein the peptide has binding to the mpl receptor Affinity, and/or having the same or greater biological activity as: IEGPTLRQWLAARA [SEQ ID NO 1]. 114552.doc • 23– 1327149 Binding affinity can be measured by any assay available or known to the skilled artisan, including but not limited to BIAcore measurements, ELISA assays, competitive assays, and the like.

Biological activity can be measured in vivo or in vitro by any assay available or known to those skilled in the art. Exemplary assays include, but are not limited to, cell-based assays, ie, megakaryocyte proliferation assays, 32D cell assays (IL-3 dependent clones of murine 32D cells transfected with human mpl receptors, as detailed in W0 95/26746) Medium), CD34+ assay, CD61 cell assay, etc. Biological activity can also be measured by various in vivo animal assays. The preferred TMP sequences of the invention are identified in Table 2 below.

Table 2 - Preferred TMP sequence TMPNo PEPTIDE SEQUENCE SEQ ID NO:. TMP2 GAREGPTLRQWLEWVRVG 2 TMP3 RDLDGPTLRQWLPLPSVQ 3 TMP4 ALRDGPTLKQWLEYRRQA 4 TMP5 ARQEGPTLKEWLFWVRMG 5 TMP6 EALLGPTLREWLAWRRAQ 6 TMP7 MARDGPTLREWLRTYRMM 7 TMP8 WMPEGPTLKQWLFHGRGQ 8 TMP9 HIREGPTLRQWLVALRMV 9 TMP10 QLGHGPTLRQWLSWYRGM 10 TMP11 ELRQGPTLHEWLQHLASK 11 TMP12 VGIEGPTLRQWLAQRLNP 12 TMP13 WSRDGPTLREWLAWRAVG 13 114552.doc -24· 1327149

TMP14 AVPQGPTLKQWLLWRRCA 14 TMP15 RIREGPTLKEWLAQRRGF 15 TMP16 RFAEGPTLREWLEQRKLV 16 TMP17 DRFQGPTLREWLAAIRSV 17 TMP18 AGREGPTLREWLNMRVWQ 18 TMP19 ALQEGPTLRQWLGWGQWG 19 TMP20 YCDEGPTLKQWLVCLGLQ 20 TMP21 WCKEGPTLREWLRWGFLC 21 TMP22 CSSGGPTLREWLQCRRMQ 22 TMP23 CSWGGPTLKQWLQCVRAK 23 TMP24 CQLGGPTLREWLACRLGA 24 TMP25 CWEGGPTLKEWLQCLVER 25 TMP26 CRGGGPTLHQWLSCFRWQ 26 TMP27 CRDGGPTLRQWLACLQQK 27 TMP28 ELRSGPTLKEWLVWRLAQ 28 TMP29 GCRSGPTLREWLACREVQ 29 TMP30 TCEQGPTLRQWLLCRQGR The binding affinity and biological activity data of the 30 peptide TMP2-TMP30 are further illustrated in the examples. In order to better mimic the phage environment used to select the peptide, and to mask the charged amine and carboxy termini of the preferred 18 amino acid peptide, add two amines to each end of each peptide. Base acid "cover". In particular, valerine (Q) and cysteine (C) are added to the amine end of each TMP2-TMP30. Similarly, at the carboxy terminus of each peptide Two amino acids "cover" "...histamine (H) and serine (S). It is understood by those skilled in the art that such covers 114552.doc -25- 1327149 are only used to mask the tau terminals and are not intended to increase or decrease the binding affinity and/or biological activity of the preferred peptides. Since the peptide affinity is known to increase with the length of the peptide, the reference bioactive peptide (SEQ ID NO 1) is increased from 14 amino acids to 22 amino acids to have the same length as the test peptide TMP2-TMP30. See Example 6_11. As will be understood by those skilled in the art, the 14 amino acid sequences which are the core of the bioactive region of the peptide are identified as in SEq ID N0 i and are also referred to as TMP1. Any peptide containing a cysteamine thiol residue can be cross-linked to another cys-containing peptide, either or both of which can be linked to a vehicle. Any peptide containing more than one Cys residue can also form a disulfide bond within the peptide. Any of these peptides can be derived as described below. Other useful peptide sequences are derived from the conservative and/or non-conservative modification of the amino acid sequences of the TMPs disclosed herein. Conservative modifications result in peptides having functional and chemical characteristics similar to those of the peptides that perform such modifications. In contrast, substantial modifications to the functional and/or chemical characteristics of the peptide can be accomplished by selective substitution in the amino acid sequence such that they differ significantly in their ability to maintain the following characteristics in the substitution zone. The structure of the main molecular skeleton, such as a thin or spiral structure, the charge or hydrophobicity of the (匕) molecule at the target site, or (c) the size of the molecule. For example, "conservative amino acid substitution" can include the substitution of a natural amino acid residue with a non-native residue such that there is little or no effect on the polarity or charge of the amino acid residue at that position. In addition, any natural residue in the multi-peptide may also be substituted by alanine, as previously described in "Alanine Scanning Mutation Formation", 114552.doc '26- 1327149 (see eg MacLennan et al., 1998, Acta) Physiol. Scand. Suppl. 643: 55-67; Sasaki et al., 1998, Adv.

Biophys. 35:1-24, which discusses the formation of alanine scanning mutations). The desired amino acid substitution (whether conservative or non-conservative) can be determined by the skilled artisan when such substitution is desired. For example, an amino acid substitution can be used to identify important residues of the peptide sequence, or to increase or decrease the affinity of the peptide or vehicle-peptide molecule described herein (see the previous formula). Exemplary amino acid substitutions are listed in Table 3. #表3-Amino acid substitution

The original residue example replaces the preferred substitution Ala (A) Val, Leu, lie Val Arg (R) Lys, Gin, Asn Lys Asn (N) Gin Gin Asp (D) Glu Glu Cys (C) Ser, Ala Ser Gin ( Q) Asn Asn Glu (E) Asp Asp Gly (G) Pro ' Ala Ala His (H) Asn, Gin, Lys, Arg Arg lie (I) Leu 'Val, Met, Ala, Phe, Leucine Leu Leu (L) leucine, lie, Val, Met, Ala, Phe lie Lys (K) Arg, 1,4-diaminobutyric acid, Gin, Asn Arg Met (M) Leu, Phe, lie Leu 114552. Doc -27- 1327149

Phe(F) Leu, Val, lie, Ala, Tyr Leu Pro (P) Ala Gly Ser(S) Thr ' Ala, Cys Thr Thr(T) Ser Ser Tip (W) Tyr ' Phe Tyr Tyr (7) Trp, Phe, Thr , Ser Phe Val (V) lie, Met, Leu, Phe, Ala, and leucine Leu. In some embodiments, conservative amino acid substitutions may also encompass non-naturally occurring amino acid residues' typical The ground is propagated through chemical peptide synthesis rather than through synthesis within the biological system. Naturally occurring residues can be classified into several classes based on the general side chain properties that can be used for sequence modification. For example, a non-conservative substitution may include the exchange of one member of these classes with one of the other classes. Such a substituted residue can be introduced into a peptide region homologous to a non-human orthol(R) gS, or to a non-same region of the molecule. In addition, P or G can also be used for modification to achieve the purpose of influencing chain orientation. The hydropathic index of the amino acid can be considered when performing such modification. Each amino acid specifies a hydrophobic index based on its hydrophobicity and charge characteristics, which are: isoleucine (+4 5); valine (+4.2), leucine (+3.8); phenylalanine ( +2.8); cysteine/cystine (+2.5) 'T-thiamine S (+1.9); alanine (+1 8); glycine (_〇4); threonine (-0.7) ); serine (_0.8); tryptophan (·〇9); tyrosine (_13); proline (-1.6); histidine (_3 2); glutamic acid (_3.5) ); glutamine (-3.5), aspartic acid (-3.5); aspartame (_3 5); lysine 114552.doc • 28-1327149 (·3·9); Acid (_4.5). The importance of hydrophobic amino acid indices in the importance of interacting biological functions of proteins is well known. Kyte et al,; M〇1 Bi〇i, 157: 105-131 (1982). It is known that certain amino acids can replace other amino acids having similar hydrophobic indices or values while still maintaining similar biological activities. In the change based on the hydrophobic index, the amino acid substitution with a hydrophobic index between ±2 is preferred, preferably between ±1, between ±〇5 or even smaller. For the best. It is also understood in the art that substitutions similar to amino acids can be effectively accomplished based on hydrophilicity. The maximum local average hydrophilicity of a protein, as governed by the hydrophilicity of its adjacent amino acids, is related to its immunogenicity and antigenicity', i.e., to the biological properties of the protein. The following hydrophilicity values are assigned to amino acid residues; spermine groups (+3.〇); lysine (+3.0); aspartic acid (+3.〇±1); proline (+) 3〇±1); serine (+0.3); aspartame (+0'2); glutamic acid (+〇2); glycine; threonine (-0.4); proline (·〇.5±1); alanine (_〇5); histidine (-0.5); cysteine (-1.0); methionine (heart 3); proline (_15); Leucine (-1.8); Isoalicin (I).8); tyrosine (23); phenylalanine (-2.5); tryptoic acid (-3.4). Among the changes based on similar hydrophilicity values, it is preferred that the amino acid is substituted with a hydrophilicity value of between ±2, preferably between ±1, between ±0.5 and even more. Small is the best. The epitope can also be identified from the original amino acid sequence based on hydrophilicity. These regions are also referred to as "antigenic determinant core regions". 谙 Those skilled in the art can use known techniques to determine appropriate variants. For U4552.doc -29·4, it is possible to monitor changes in the molecule without Suitable regions for disrupting activity, and those skilled in the art may target regions that are believed to be unimportant to activity, for example, when similar polypeptides of the same or different species having similar activities are known, Comparable amino acid sequences of peptides and similar peptides. Under this comparison, residues and moieties in the molecule that are conserved relative to similar polypeptides can be identified. It is known that peptides are relative to this class. Changes in regions where the similar peptides are not conserved are less likely to negatively affect the biological activity and/or structure of the peptide. It will also be appreciated by those skilled in the art that chemically similar amine groups can be used even in fairly conserved regions. An acid replaces a naturally occurring residue while retaining activity (conservative amino acid residue substitution). Thus, conserved amino acid substitutions can be made even for biologically active or structurally important regions without disrupting biological activity or negative The surface affects the structure of the peptide. The amino acid has L or D stereochemistry (other than Gly, it is neither L nor D) 'The TMPs of the present invention may include a stereochemical combination. However, L stereochemistry is all for the TMP chain. The amino acid is preferred. The present invention also provides a reverse TMP molecule in which the amino-terminal to carboxy-terminal sequence of the amino acid is reversed. For example, a molecule having a normal XrXrXs sequence will be reversed. Is X3-X2-X1. The present invention also proposes a reverse-reverse TMP molecule in which, like the reverse TMP, the amino-terminal to carboxy-terminal sequence of the amino acid is reversed, and the TMP is usually "L" The residue of the enantiomer is changed to the "D" stereoisomer form. Derivatives of TMP are also contemplated to replace the above TMP. Such TMP derivatives include those in which one or more of the following modifications can be made: One or more of the peptidyl [-C(0)NR-] linkages are substituted by a non-peptidyl linkage, such as a -CH2-amino phthalate linkage [·〇η2-〇(:( ο)νιι-]; 114552.doc •30- 1327149 phosphate linkage; -CIV sulfonamide linkage; urea [-NHC(0)NH-] linkage; ·〇η2_secondary amine linkage; or alkyl Chemical victory Peptidyl linkage [-C(0)NR6- wherein R6 is a lower alkyl group]; • peptide, wherein the N_ terminal is derivatized to a _NRRi group; _nrc(〇)r

Base; -nrc(o)or base; _nrs(0)2R base; -NHC(9)NHR group, wherein R and R are murine atoms or low carbon number, but the restriction is that R and R1 are not all hydrogen atoms; a quinone imino group; a benzoquinone group-NH-(CBZ-NH-) group; or a benzoquinoneoxycarbonyl fluorenyl group having 1 to 3 groups selected from the group consisting of the following Substituents: lower alkyl, lower alkoxy, gas and bromine; and • peptides where the free C-terminus is _c(〇)R2, where R2 is selected from the following groups: low carbon Alkoxy and -NR3R4, wherein R3 and R4 are each independently selected from the group consisting of a hydrogen atom and a lower alkyl group. "Low carbon number" means that the group has 1 to 6 carbon atoms. Further, the modification of each amino acid can be carried out into the TMP molecule by reacting the target amino acid residue of the peptide with an organic derivatizing agent, which can be reacted with a selected side chain or terminal residue. The following are examples: The amidoxime and amine end residues can be reacted with amber or other carboxylic anhydrides. The derivatization of these agents has a reversal effect on the charge carried by the amine sulfhydryl residue. Other suitable reactants for derivatization of the α-amino group-containing residue include quinone imine g, such as methyl pyridinium imide; Pyridoxal phosphate; pyridoxal; gas borohydride; trinitrobenzenesulfonic acid; hydrazine-methylisourea; 2,4-pentanedione; and transaminase catalytic reaction of acetylate. The spermine sulfhydryl residue can be reacted with one or more conventional reactants to modify 114552.doc 1327149 '' which includes phenylethylene di-chain, 2,3-butanedione, i,2·cyclohexane di- and Ninhydrin. The derivatization of arginine residues requires a reaction under hydrazine conditions due to the high pKa of the oxime functional group. In addition, these reactants can also react with the amine group and the arginine group. A number of studies have been carried out on the specific modification of the tyramine group, with particular attention to the permeation with aromatic diazonium compounds or It was confirmed that the methyl group was introduced into the spectrum to the tyramine residue. Most commonly, N-ethenyl imidazole and tetranitromethane can be used to form the indole-acetyl tyramine species and the 3-nitro derivative, respectively. The pendant carboxyl group (aspartame or amidoxime) can be selectively modified by reaction with carbodiimide (R'-N=C=N-R'), such as 丨_cyclohexyl _ 3_(2_morpholinyl-4-ethyl)carbodiimide or ethyl_3_(4-azo- 4,4-didecylpentyl)carbodiimide. In addition, aspartame and glutamine residues can be converted to aspartame and glutamine residues by reaction with ammonium ions. The glutamine and amphoteric amine amide groups typically remove the guanamine group to form the corresponding ketone and aspartame residues. Alternatively, any form in which these residues can be removed from the guanamine group under weakly acidic conditions is within the scope of the invention. Derivatization using a bifunctional reactant can be used to link the peptide or its functional derivative to a water insoluble collateral matrix or other macromolecular carrier. Commonly used crosslinking agents include, for example, 1,1-bis(disazoethyl)-2-phenylethane, pentane dimer, N-hydroxybutylimine, such as 4-azide water An ester of a maleic acid, a homobifunctional guanidinium ester, including a disuccinimide ester such as 3,3,_disulfide 114552.doc •32· 1327149 generation double (sodium sulphate) Ester) and bifunctional maleimide, such as bis-N-maleimine-1,8-octane. Derivatizing agents such as methyl·3_[(p-azidophenyl)dithio]propanoid imidate can form photoactive intermediates capable of forming crosslink bonds in the presence of light, in addition, reactivity Water-insoluble matrices such as desertified cyanide-activated listeners and reactive matrices described in the following documents can be used for protein immobilization: U.S. Patent Nos. 3,969,287; 3,691,16; 4,195,128; 4,247,642; 4,229,537 and 4,330,440. Other possible modifications include carboxylation of lysine and lysine, phosphorylation of hydroxy groups in serine sulfhydryl or sulphalidyl residues, oxidation of sulfur atoms in cysteine, lysine, spermine Methylation of α-Amine Groups in Acid and Amine Acid Side Chains (Creighton, TE, Proteins: Structure and Molecule

Properties, W.H. Freeman & Co., San Francisco, pp. 79-86 (1983)) The acetylation of the N-terminal amine and, in some instances, the amidation of the c-terminal carboxyl group. The fraction derived from this pass preferably improves the one or the characteristics of the compound of the present invention, including platelet production activity, solubility absorbability, biological half life and the like. In addition, the derived portion may result in a compound having the same or substantially the same properties and/or properties as the underivatized compound. This moiety may additionally eliminate or impair the adverse effects of the compound and the like. The compounds of the invention may also be altered at the DNA level. The DN A sequence of any portion of the compound may be made to be more compatible with the host cell of choice. The optimized codon for E. coli from a preferred host is known. The codon can be substituted to eliminate restriction sites or include 缄 114552.doc -33· 1327149 silent restriction site, which facilitates DNA processing within the selected host cell. The DNA sequences of the vehicle, the linker and the peptide can be modified to include any of the sequence changes. Accordingly, all modifications, substitutions, derivations, and the like discussed herein are applicable to all aspects of the invention, including but not limited to peptides, peptide dimers and multimers, linkages, and vehicles. In addition, the structure-function study can be reviewed to identify residues that are important for activity and structure in similar peptides. In view of such information, the skilled artisan can predict the importance of amino acid residues in the peptide corresponding to amino acids which are important for activity or structure in similar peptides. Alternatively, a chemically similar amino acid may be substituted for such an important amino acid residue as predicted in the peptide. The skilled artisan can also analyze the two-dimensional structure and the amino acid sequence in relation to the structure of a similar multi-peptide. In view of this information, the skilled artisan may choose not to fundamentally biochemically alter the amino acid residues expected to appear on the surface of the protein, as such residues may involve important interactions with other molecules. In addition, testers can be made to produce test variants containing a single amino acid substitution in each of the desired amino acid residues, and then screening the variants using activity assays known to those skilled in the art. Such data can be used to gather information about appropriate variants. For example, if changes in one-to-one specific amino acid residues are found to result in disruption of activity, undesired reduction or inappropriate activity, variants with such changes should be avoided. In other words, based on information gathered from such routine experiments, the skilled artisan can readily determine which amino acids should be avoided, either alone or in combination with other mutations. 114552.doc -34· A large number of scientific articles are devoted to the prediction of secondary structure. See Moult J., Curr. Op. in Biotech., 7(4): 422-427 (1996), Chou et al. 5 Biochemistry, 13(2): 222-245 (1974); Chou et al., Biochemistry , 113 (2): 211-222 (1974); Chou et al., Adv.

Enzymol· Relat. Areas Mol. Biol., 47:45-148 (1978); Chou et al., Ann· Rev. Biochem., 47: 251-276 and Chou et al., Biophys. J., 26:367 -384 (1979). In addition, existing computer programs can help predict secondary structure. One method of predicting secondary structure is based on homology modeling. For example, two multi-peptides or proteins, which have a sequence identity of more than 30%, or a similarity of more than 40%, often have similar structural topologies. Recent growth in protein structure databases (PDBs) The predictability of secondary structure enhancement has been proposed, including the number of possible folds in multi-peptide or protein structures. See Holm et al., Nucl. Acid. Res., 27(1): 244-247 (1999). Brenner et al., Curr. Op. Struct. Biol., 7(3): 369-3 76 (1997) speculates that there is a finite number of folds in a given multi-peptide or protein, and once the structure is resolved Key figures to achieve very accurate structural predictions. Other methods for predicting secondary structure include "threading" (threading) (Jones, D., Curr. Opin. Struct. Biol., 7(3): 377 -87 (1997); Sippl et al., Structure, 4(1): 15-9 (1996)), "Profile Analysis "(卩1:〇^16&1^17518)(8〇\^ 6 &1.,3(^611.6,253:164-170(1991); Gribskov et al., Meth. Enzym., 183:146-159 (1990); Gribskov et al., Pr〇c Nat. Acad. Sci·, 84(13): 4355-8 (1 987)) 'and "Evolutional Linkages" (see Home, supra, and Brenner, 114552.doc • 35·1327149 supra). The general formula of the preferred peptide and peptide-linker molecules of the invention is shown in Figure 1. In addition, physiologically acceptable salts of TMPs are also contemplated. Peptide Compounds In addition to novel peptides, the present invention provides novel peptide compounds. One or more of the peptides of the present invention are followed by a linking agent ( LN) and/or vehicle (V) are 'on or otherwise bonded to each other. TMP can be cascaded (ie, sequentially 'N- to C-end) or parallelly linked (ie, N-to N-terminal or C- to C-terminus. TMP can be followed by other tmp or the same TMP with or without a linking agent. TMP can also be used with other TMPs or the same TMP, with or without a linking agent and with or without a vehicle. The peptide-bonding agent-vehicle compound of the present invention can be described by the following formula;

II (Vl)v--(LNl)1--(TMPl)a--(LN2)m--(TMP2)b--(LN3)n-- (TMP3)c-(LN4)0--(TMP4 Wherein d-(v2)w wherein V1 and V2 are vehicles; LN1, LN2, LN3 and LN4 are each independently a linking agent; TMP1, TMP2, TMP3 and TMP4 are each independently a peptide sequence of formula I; a' b, c and d and 1, m, η and 〇 are each independently an integer from 〇 to 20' and v*w are each independently an integer from 〇 to 1. The compounds of the present invention are, for example, represented by the following formula: TMP1-V1 TMP1-LN1-V1 TMP1-TMP2-V1 TMP 1-LN1-TMP2-LN2-V1 and other multimers thereof, wherein V1 is a vehicle ( Preferably, the Fc functional portion 114552.doc • 36* 1327149 bit) 'and the linkage is at the C-terminus of the TMP, with or without a bonding agent; V1-TMP1 V1-LN1-TMP1 V1-TMP1-TMP2 V1 -LN1 -TMP 1-LN2-TMP2 and other multimers thereof, wherein VI is a vehicle (preferably an Fc functional site)' and is linked at the N-terminus of TMP with or without linkage The general formula of the preferred peptide-agent and peptide-bonding agent-agent molecule of the present invention is shown in Figure 2.

Many of the preferred compounds of the invention are dimers or polymers in which they have two TMP moieties, or multimers, which have multiple TMP moieties. Each of TMP1 to TMP4 and the like may have the same or different structure. Preferably, the compound of the present invention has 2 to 5 TMP moieties, particularly preferably 2 to 3, and most preferably 2.

These compounds are preferably dimers which are bound directly or via a linkage (see below). The monomeric TMP moiety system is shown in a conventional orientation and the N-to-C-end solution is read from left to right. Accordingly, it can be seen that the compounds of the present invention can be oriented such that the C-terminus of TMP 1 binds directly or through a linkage to the N-terminus of the TMP2 (cascade dimer). In addition, the compounds of the invention may be oriented such that the C-terminus of TMP1 binds directly or through a linkage to the C-terminus of TMP2, or the N-terminus of TMP1 binds directly or through a linkage to the N of TMP2. - terminal (parallel dimer). These compounds are referred to as dimers' even though TMP 1 and TMP2 are structurally distinct. That is, it can be constructed as a homodimer and a heterodimer. Bonding Agent In another embodiment, the present invention provides one or more TMp permeable 114552.doc • 37· 1327149, a linking agent, a group (LN1, LN2, etc.) covalently bonded or otherwise The method is linked or combined with another TMP. Any linkage group is optional. When it is present, its chemical structure is not important because it acts primarily as a spacer. The linkage should be chosen such that it does not affect the biological activity of the final compound' and the immunogenicity of the final compound will not be significantly improved. The linking agent is preferably composed of an amino acid bonded together by a peptide bond. Thus, in the preferred embodiment the ' linkage agent consists of from 1 to 30 amino acids bonded by a peptide bond wherein the amino acid is selected from the group consisting of 20 naturally occurring amino acids. - Some of these amino acids are glycosylated, as is well understood by those skilled in the art. In a more preferred embodiment, from 1 to 20 amino acids are selected from the group consisting of glycine, alanine, valine, aspartame, glutamine and lysine. More preferably, the linking agent consists of a main body of amino acid which is not sterically hindered, such as glycine and alanine. Therefore, the preferred linking agent is polyglycolic acid (particularly (Gly) 4 ' (Gly) 5) 'poly(Gly-Ala) and polyalanine. Specific examples of other linking agents are: (Gly)3Lys(Gly)4 (SEQ ID NO: 96); (Gly)3AsnGlySer(Gly)2 (SEQ ID NO: 97); (Gly)3Cys(Gly)4( SEQ ID NO: 98); and GlyProAsnGlyGly (SEQ ID NO: 99) To explain the above terms, such as (Gly)3Lys(Gly)4 represents Gly-Gly-Gly-Lys-Gly-Gly-Gly-Gly-. Gly and Ala are also preferred to use. The linking agents shown herein are exemplary; the linking agents within the scope of the present invention may be longer and may include other residues. Non-peptide-linking agents are also possible. For example, an alkyl linkage agent such as -NH-114552.doc •38· 1327149 (CH2)S_C(0)·, wherein s=2-20 is available. These alkyl linking agents may be further substituted by any non-sterically hindered group such as a lower alkyl group (e.g., a lower alkyl group, a halogen (e.g., gas, bromine), CN, NH2, phenyl, etc. a non-peptide _ bond An example of a crosslinking agent is a polyethylene glycol linkage agent.

Wherein n is such that the bond has a molecular weight of from 100 to 5,000 kD, preferably from 100 to 500 kD. The peptide-bonding agent can be altered to form a derivative in the manner described above. It is known that a bonding agent having a length of about 0 to 14 units (e.g., an amino acid) is preferred for the gold platelet-forming compound of the present invention. The skin-bonding agent can be modified in the same manner as described in the above TMP to form a derivative. Further, the compound of this specific example may further be linear or cyclic. "ring type" means that at least two separate, ie non-contiguous, parts of the molecule are linked to each other. For example, the amine end and the carboxy terminus of the molecular end may be covalently bonded to form a cyclic molecule. Additionally, the molecule may comprise two or more Cys residues (as in a linking agent) which are cyclizable by the formation of a disulfide bond. It is further contemplated that more than one cascade of peptide dimers can be linked to form a dimer dimer. Thus, for example, a conjugated dimer containing a Cys residue can form an intermolecular disulfide bond with the Cys of another such dimer. EXAMPLES The peptide-bonding compound of the present invention is as shown below;

CSSGGPTLREWLQCRRMQ-GGGGG-CSSGGPTLREWLQCRRMQ I14552.doc -39· 1327149 (SEQ ID NO 100); QLGHGPTLRQWLSWYRGM-(Gly)3Lys(Gly)4-ALRDGPTLKQWLEYRRQA (SEQ ID NO 101); RFAEGPTLREWLEQRKLV-GGG(PEG)GGG-RFAEGPTLREWLEQRKLV (SEQ ID NO 102). Accordingly, in a preferred embodiment, the linking agent comprises (LN1)n, wherein LN1 is a naturally occurring amino acid or a stereoisomer thereof, and "n" is any one of 1 to 20. The formula of the preferred peptide-linker molecule is shown in Figure 1. More peptide-linker molecules include: i) TMP1-LN1-TMP2-LN2 ii) LN1-TMP1-LN2-TMP2 iii) LN1-TMP1-LN2-TMP1 iv) TMP1-LN1-TMP1-LN1-TMP1 -LN1 v) LN1-TMP1-LN2-TMP2-LN3-TMP3-LN4-TMP4 wherein LN1-LN4 are each independently a linking agent. Vehicle In another embodiment, the peptide or peptide compound of the invention may be linked or subsequently attached to a vehicle (V). A vehicle generally refers to a molecule that prevents the degradation of therapeutic proteins and/or prolongs their half-life, reduces their toxicity, reduces immunogenicity, or increases biological activity. The vehicle (V) can be passed over the peptide via the N-terminus, the C-terminus, the peptide backbone or the side chain. The vehicle (V) can be a carrier molecule, such as a linear polymer (such as polyethylene glycol, polylysine, dextran, etc.), a branched polymer (see, for example, issued on September 15, 1981) Denkenwalter et al., U.S. Patent No. 114, 552, doc, filed on Apr. 20, 1993, to U.S. Patent No. 5,229,490 issued to Tam, and on December 28, 1993, by Frechet et al. 21259), a lipid; a cholesteryl group (such as a steroid or a saccharide or an oligosaccharide. Other possible vehicles include one or more water-soluble polymer additions such as polyoxyethylene ethylene glycol, or polypropylene glycol) , as described in U.S. Patent Nos. 4,640, 835, 4, 496, 689, 4, 301, 144, 4, 670, 417, 4, 791, 192 and 4, 179, 337. Other useful polymers known in the art include monodecyloxy-polyethylene glycol, dextran, cellulose. Or other sugar-based polymers, poly(Ν-vinylpyrrolidone)-polyethylene glycol, propylene glycol homopolymer, polyoxypropylene/ethylene oxide copolymer, polyoxyethylated polyol (such as glycerin And polyvinyl alcohol, and these polymerizations Exemplary vehicles also include: • an Fc functional site; • other proteins, polypeptides or peptides that bind to salvage receptors; • human serum globulin (HSA); • leucine zipper (LZ) Domain; polyethylenol (PEG), including 5 kD, 20 kD and 30 kD PEG, and other polymers; • glucomannans; and molecules known in the art to provide extended half-life and/or Or avoid proteolytic degradation or clearance. An exemplary vehicle is polyethylene glycol (PEG) ^ PE (} family can have any convenient knife 1, and can be linear or branched. The average molecular weight of pEG is better. The range is from about 2 kDa to about i00 kDa, more preferably from about 5 kDa to about 5 〇I14552.doc 41 1327149 kDa, and the best is between about 5 kDa and about l〇kDa. The PEG family usually passes through the following reactions. Subsequent to the compounds of the invention: deuteration, reductive burning, Michael addition, thiol gasification or other chemoselective conjugate/linkage, through a reactive group on the peg moiety (eg Base, amine group, ester group, thiol, -haloethyl, maleimine or The thiol group reacts with a reactive group (such as an aldehyde group, an amine group, an ester group, a thiol group, a thiol group, a maleimide group or a thiol group) on the target compound. It can be conveniently followed by a glycosylation site known in proteins. In general, the oligosaccharide linked by 〇 _ is attached to a residue of serine or threonine (Thr), and The linked oligosaccharides are then attached to the aspartic amine (Asn) residues, which are part of the Asn-X-Ser/Thr sequence, wherein X can be any amino acid other than proline. . X is preferably one of the 19 naturally occurring amino acids' but does not include proline. The structure of the oligosaccharides bonded through the N-bond and the oxime _ and the saccharide residues found in each type are different. One type of sugar that is commonly found in both is N-ethyl thio-neuraminic acid (referred to as sialic acid). The sialic acid is often the terminal residue of the N_ linkage and the 〇_linked oligosaccharide', and the glycosylated compound can be made acidic by the negative charge it carries. Such sites may be added to the linkage agents of the present invention, and are preferably glycosylated by the cells during recombinant formation of the compound (e.g., in mammalian cells such as CHO, BHK, cos). However, such sites can be further glycosylated by synthetic or semi-synthetic procedures known in the art. In a more preferred embodiment, the vehicle (V) may comprise one or more antibody Fc functional sites. Thus, the above peptide compounds can be further fused to one or more U4552.doc • 42·1327149 Fc functional sites, either directly or via a bonding agent. The Fc vehicle can be selected from the human immunoglobulin IgG-Ι heavy chain 'see EUison, JW et al., Nucleic Acids Res. 10: 4071-4079 (1982), or any other Fc sequence known in the art (eg other The igG family includes, but is not limited to, igG-2, IgG_3, and IgG-4, or other immunoglobulins. The F c functional site of the 'antibody' antibody is composed of a monomeric multi-peptide segment that is conjugated to a dimeric or multimeric form via a disulfide or non-covalent bond. The number of intermolecular disulfide bonds between the haplotype subunits of the native F c molecule ranges from 1 to 4 ' depending on the class of antibody (eg igG, IgA, IgE), or subclass (eg IgGl, IgG2, IgG3) , IgAl, IgGA2) » The term ''Fc' as used herein is generally referred to as the monomeric, dimeric and multimeric forms of the Fc molecule. It should be noted that the Fc monomer can spontaneously dimerize when it contains a suitable Cys residue. 'Unless there are special conditions to avoid dimerization through the formation of disulfide bonds. Even Cys residues that normally form disulfide bonds in dimers are removed or replaced by other residues' monomer chains are usually It also dimerizes through non-covalent interactions. The term "Fc" is used herein to mean any such form: natural monomers, natural dimers (disulfide bonds), modified dimerization a body (disulfide and/or non-covalent bond) and a modified monomer (ie, a derivative). ^ A variant, analog or derivative of a moiety can be passed through, for example, to carry out various residues or sequences. Composed of substitutions. Variants (or analogs) Polypeptides include intervening variants, of which One or more amino acid residues complement the Fc amino acid sequence. The insertion may be located at either or both ends of the protein' or may be placed in the internal region of the Fc amino acid sequence. Addition at either or both ends Insertion variants of residues include, for example, fusion egg 114552.doc -43 - 1327149 white matter and amino acid-containing tag or labeled protein. For example, an Fc molecule may optionally comprise an N-terminal Met, particularly when molecular recombination When expressed in a bacterial cell, such as E. coli. In the Fc deletion variant, one or more amino acid residues in the Fc polypeptide are deleted. Deletion can be carried out at one or both ends of the Fc polypeptide. Or deleting one or more residues within the Fc amino acid sequence. Thus deleting the variant includes all fragments of the Fc polypeptide sequence. Among the Fc substitution variants, one or more amine groups of the Fc polypeptide The acid residue is removed and replaced with an alternative residue. In one aspect, the substitution is conservative in nature, but the invention also encompasses non-conservative substitutions. For example, a cysteine residue can be deleted or otherwise Amino acid substitution to avoid some formation of Fc sequences All disulfide linkages. Each of these cysteine residues may be removed or one or more such cysteine residues may be substituted with other amino acids, such as Ala or Ser. As another example Alternatively, it may be modified to introduce an amino acid substitution to (1) to remove the receptor binding site; (2) to remove the complement (Clq) binding site; and/or (3) to remove antibody-dependent cellular mediators. (ADCC) sites. Such sites are known in the art, and any known substitutions are within the scope of the Fc used herein. For example, see Molecular Immunology, v〇1. 29, N〇5, 633 639 ( 1992) Regarding the ADCC site in IgG1. Similarly, one or more tyrosine residues may also be substituted with amphetamine residues. In addition, insertions and deletions of other variant amino acids (e.g., from 丨25 amino acids) and/or substitutions are also contemplated and are within the scope of the invention. A conservative amino acid substitution is generally preferred. In addition, the modifications may be in the form of an amino acid of the modified 114552.doc-44-1327149, such as a peptide mimetic or a D-amino acid. The Fc sequence of the invention may also be derived, ie, with insertion and removal. Or a modification other than the substitution of an amino acid residue. Preferably, the modifications are intrinsically covalently bonded and include, for example, chemical linkages to polymers, lipids, other organic and inorganic moieties. The derivatives of the invention may be prepared to prolong the half-life of the cycle' or may be designed to improve the ability of the multi-peptide to direct to a desired cell, tissue or organ. The salvage receptor binding domain in the entire Fc molecule can also be used as the Fc portion of the compounds of the invention, as described in WO 96/32478, entitled "Ahered Polypeptides with Increased Half-Life". The other members of the molecular class are those described in W〇97/3463 1, entitled "Immunoglobulin-Like Domains with Increased Half-Lives". The two published pct applications cited in this paragraph are by reference. Into this article.

The Fc fusion can be at the N- or C-terminus of TMP1 or TMP2, or at the N- and C-termini of TMP1 or TMP2. Similarly, Fc fusion can be at the N- or C-terminus of the fc functional site. Preferred compounds of the invention include IgGl Fc fusion dimers that are linked or otherwise followed by dimers or multimers of TMPs disclosed herein. In such instances, each F c functional site is linked to a TMP dimer or multimer, with or without a linking agent. A schematic example of such a compound is shown in Figure 2. Multiple vehicles can also be used; such as having several Fc at each end or one Fc at one end and one PEG group at the other or side chain. 114552.doc • 45· 1327149 The exemplary peptide-vehicle compounds are set forth in Table 4 below. Table 4 - Example peptide-vehicle compound amino acid sequence t SEQ ID NO: HIREGPTLRQWLVALRMV-GGG(PSG)GGG-HIREGPTLRQWLVA.LRMV 103 Fc-TCEQGPTLRQWLLCRQGR-GGGKGGG-TCEQGPTLRQWLLCRqGR-Fc 104 Fc-QLGHGPTLRQWLSWVRGM-GPNG-ELRSGPTLKEWLVWRLAq 105 CSWGGPTLKQWLQCVRAK -Fc I 1 SWGGPTLKQWLQCVRAK 106 Fc-GGGKGGG-AVPQGPTLKQWLLWRRCA 107 PEG-CSSGGPTLREWLQCRRMQ I 1 CSSGGPTLREWLQCRRMQ 108 Fc-GGGGG-YCDEGPTLKQWLVCLGLQ-GGGGG-YCDEGPTLKQWLVCLGLQ 109 cswggptlkqwlqcvrak-gggaggg-cswggptlkqwlqcvrak-gggaggg- CSWGGPTLKQWLQCVRAK-GGGAGGG-Fc 110 VGIEGPTLRQWLAQRLNP-GGGCGGG-VGIEGPTLRQWLAQRLNP-PEG 111 Fc-ELRSGPTLKEWLVWRLAq-GGGG-ELRSGPTLKEWLVWRLAQ 112 Fc-ALRDGPTLKQWLEYRRQA-GGGKGGG-ALRDGPTLKQWLEYRRQA-Fc 113

Further, Table 5 lists preferred embodiments of the present invention. Table 5. Specific examples of preferred specific amino acid sequence of SEQ ID NO: ALRDGPTLKQWLEYRRQA-ALRDGPTLKQWLEYRRQA 114 EALLGPTLREWLAWRRAQ-EALLGPTLREWLAWRRAQ 115 AVPQGPTLKQWLLWRRCA-AVPQGPTLKQWLLWRRCA 116 YCDEGPTLKQWLVCLGLQ-YCOEGPTLKQWLVCLGLQ 117 CSSGGPTLREWLQCRRMQ-CSSGGPTLREWLQCRRMQ 118 CSWGGPTLKQWLQCVRAK-CSWGGPTLKQWLQCVRAK 119 ALRDGPTLKQWLEYRRQA-GGGGG-ALRDGPTLKQWLEYRRQA 120 eallgptlrewlawrraq-ggggg -eallgptlrewlawrraq 121 avpqgptlkqwllwrrca-ggggg-avpqgptlkqwllwrrca 122 YCDEGPTLKQWLVCLGLQ-GGGGG-YCDEGPTLKQWLVCLGLQ 123 CSSGGPTLREWLQCRRMQ-GGGGG-CSSGGPTLREWLQCRRMQ 124 CSWGGPTLKQWLQCVRAK-GGGGG-CSWGGPTLKQWLQCVRAK 125 Fc-GGGGG-ALRDGPTLKQWLEYRRQA 126 FC-GGGGG-EALLGPTLREWLAWRRAQ 127 Fc-GGGGG-AVPQGPTLKQWLLWRRCA 128 Fc-GGGGG-YCDEGPTLKQWLVCLGLQ 129 Fc-GGGGG-CSSGGPTLREWLQCRRMQ 130 Fc-GGGGG-CSWGGPTLKQWLQCVRAK 131

46 - 114552.doc 1327149

Manufacturing Methods The compounds of the present invention can be produced in a variety of ways. Since many such compounds are peptides, or include peptides, the method of synthesizing peptides has a particular relevance herein. Solid phase synthesis techniques can be used. Suitable techniques are well known in the art, including those described in the following documents: Merrifield, in Chem. Polypeptides, pp. 335-61 (Katsoyannis and Panayotis eds. 1973); Merrifield, J. Am. Chem. Soc. 85: 2149 (1963); Davis et al., Biochem. Inti. 10: 394-414 (1985); Stewart and

Fc-GGGGG-ALRDGPTLKQWLEYRRQA-GGGGG-ALRDGPTLKQWLEYRRQA 132 Fc-GGGGG-EALLGPTLREWLAWRRAQ-GGGGG-EALLGPTLREWLAWRRAQ 133 Fc-GGGGG-AVPQGPTLKQWLLWRRCA-GGGGG-AVPQGPTLKQWLLWRRCA 134 Fc-GGGGG-YCDEGPTLKQWLVCLGLQ-GGGGG-YCDEGPTLKQWLVCLGLQ 135 Fc-GGGGG-CSSGGPTLREWLQCRRMQ-GGGGG-CSSGGPTLREWLQCRRMQ 136 Fc-GGGGG-CSWGGPTLKQWLQCVRAK-GGGGG-CSWGGPTLKQWLQCVRAK 137 ALRDGPTLKQWLEYRRQA-GGGGG-ALRDGPTLKQWLEYRRQA'GGGGG-FC 138 eallgptlrewlawrraq-ggggg-eallgptlrewlawrraq-ggggg-fc 139 AVPQGPTLKQWLLWRRCA-GGGGG-AVPQGPTLKQWLLWRRCA-GGGGG-Fc 140 YCDEGPTLKQWLVCLGLQ-GGGGG-YCDEGPTLKQWLVCLGLQ-GGGGG-Fc 141 CSSGGPTLREWLQCRRMQ-GGGGG-CSSGGPTLREWLQCRRMQ-GGGGG-Fc 142 CSWGGPTLKQWLQCVRAK-GGGGG-CSWGGPTLKQWLQCVRAK-GGGGG-Fc 143 ALRDGPTLKQWLEYRRQA-GGGGG-FC 144 EALLGPTLREWLAWRRAQ-GGGGG-FC 145 AVPQGPTLKQWLLWRRCA-GGGGG-FC 146 YCDEGPTLKQWLVCLGLQ-GGGGG-Fc 147 CSSGGPTLREWLQCRRMQ-GGGGG-Fc 148 CSWGGPTLKQWLQCVRAK-GGGGG-Fc 149

Young, Solid Phase Peptide Synthesis (1969); U.S. Patent No. 3,941,763; Finn et al., The Proteins, 3rd ed., vol. 2, pp. 105-253 (1976); and Erickson et al., The Proteins, 3rd ed., vol. 2, pp. 257-527 (1976). Solid phase synthesis is a preferred technique for making individual peptides because it is the most cost effective method for making small peptides. The peptide can also be produced in a transgenic host cell using recombinant DNA techniques. To carry out this, a recombinant DNA molecule encoding the peptide is prepared. Methods of preparing such DN A and/or RNA molecules are well known in the art. For example, ' • 47· 114552.doc 1327149 The sequence of the peptide can be excised from the dna using appropriate restriction enzymes. The formation of related sequences can be made using polymerase chain reaction (PCR) and adding useful restriction sites for subsequent selection. Alternatively, DNA/RNA molecules can be synthesized using chemical synthesis techniques such as the phosphoramidite method. Combinations of these or other techniques can also be used. The invention also encompasses a vector encoding a peptide in a suitable host. The vector comprises a DNA molecule encoding the peptide which is operably linked to an appropriate expression control sequence. The method of performing this operative linkage is performed before or after β is inserted into the vector by the DNA knife encoding the peptide. Expression control sequences include promoters, activators, enhancers, ribosome binding sites, initiation signals, stop signals, cap signals, polyadenylation k and other signals associated with transcriptional or translational control. A vector known to include a DNA molecule encoding a peptide is used to transform a suitable host. This transformation can be carried out using methods well known in the art. The selection of a large number of existing and well-known host cells can be used to practice the selection of the present invention depending on various factors identified by the present technology. Some of the factors include 'the compatibility of the sample with the selected expression vector, the toxicity of the peptide encoded by the DNA molecule to the host cell, the rate of transformation, the difficulty and performance characteristics of peptide recovery, biosafety and cost. To understand that not all hosts have the same effect on the performance of a particular ship sequence, the balance of these factors must be achieved. Under these general guidelines, available microbial hosts include bacteria = large intestine (4), yeast sputum (such as yeast g (~Spirit) and Piehia pastGHs) and other cultured fungi, insects. I14552.doc -48- 1327149 Plant, mammalian (including human) cells, or other host-transformed host lines known in the art, are cultured under conventional fermentation conditions to exhibit desirable peptides. Such fermentation conditions are well known in the art. The peptide is then purified from the hairy growth nucleus or from the host cell expressing the peptide. Purification methods are also well known in the art. The compound containing the derived peptide, or the compound containing the non-peptidyl group, can be synthesized by well-known organic chemical techniques. For example, solid phase synthesis techniques can be used. Suitable techniques are well known in the art, including those described in Merdfield (1973), Chem. Polypeptides, 335 61 (Katsoyannis and Panayotis eds.); Merrifield (1963), J. Am.

Chem. Soc. 85:2149; Davis et al. (1985), Biochem. Inti. 10:394-414; Stewart and Young (1969), Solid Phase Peptide

Synthesis; U.S. Patent No. 3,941,763; Finil et al. (1976), The Proteins (3rd ed.) 2:105-253; and Erickson et al. (1976), The Proteins (3rd ed.) 2: 257 -5 27. Solid phase synthesis is a preferred technique for making individual peptides' because it is the most cost effective method for making small peptides. IV. Uses of Compounds The compounds of the invention have the ability to bind to and/or activate mpl receptors, and/or can stimulate the production of platelets (in vivo and in vitro) ("platelet production activity") and production of platelet precursors ("megakaryocyte-forming activity"). To measure the activity of these compounds, standard assays can be used, as described in WO 95/26746 entitled "Compositions and Methods for Stimulating Megakaryocyte Growth and Differentiation". In vivo assays Further description is made in the Examples section of this article. U4552.doc • 49- The method of the present invention and the treatment of human sputum α, sputum are generally related to the existing megakaryocytes / blood stalks + +, lack or In the future, megakaryocytes/blood may be or are expected to be small (such as surgery or platelet donation due to 4 strokes). Such conditions may be in vivo active-ligand (temporary or permanent) deficiency, j The lack of plates is commonly referred to as gray platelet reduction, and therefore, the methods and compounds of the present invention are generally useful for the prevention or treatment of patients in need thereof. Thrombocytopenia. The World Health Organization has classified the extent of thrombocytopenia according to the number of internal circulations and small plates (Miller, et al., Cancer 47:21〇_211 (198 1)). For example, _ shed soil— Individuals who did not show thrombocytopenia (Grade 0) usually had at least 1 GG, GGG platelets/cubic millimeters. Mild thrombocytopenia (grade U) showed that platelet circulation levels ranged from 79, _ to 99, 〇〇〇/cubic millimeters. Moderate thrombocytopenia (Grade 2) shows that platelet circulation levels range from 000 to 74, GGG/cubic millimeter, and severe thrombocytopenia is characterized by between 25, _ and 49,000 per cubic millimeter. Debilitated thrombocytopenia is characterized by a circulating concentration of platelets of less than 25 嶋/m 3 . ' Thrombocytopenia (platelet deficiency) also exists for a variety of reasons, including chemotherapy and other treatments using various drugs, radiation, surgery Surgery, blood loss and other specific disease conditions. Example specific diseases involving thrombocytopenia that can be treated according to the present invention: aplastic disorders Primary anemia or immune thrombocytopenia (ITP), including primary thrombocytopenic purpura associated with breast cancer; ITp and mv-related thrombocytopenia purpura associated with mv; leading to thrombocytopenia Tumor metastasis line H4552.doc -50· 1327149 systemic lupus erythematosus; including neonatal lupus syndrome splenomegaly; Fanconi's syndrome; vitamin B12 deficiency; folate deficiency;] ^^_11% to 111 abnormalities ' Wiskott-Aldrich syndrome; chronic liver disease; myelodysplastic syndrome associated with thrombocytopenia; paroxysmal nocturnal hematuria; acute severe thrombocytopenia after C7E3 Fab (Abciximab) treatment; alloimmune thrombocytopenia, including maternal alloimmunization Thrombocytopenia; thrombocytopenia associated with antiphospholipid antibodies and A sputum; autoimmune gray platelet reduction; drug-induced thrombocytopenia, including carbamazepine-induced thrombocytopenia, heparin-induced Golden platelet reduction; fetal thrombocytopenia; thrombocytopenia during pregnancy; Hughes syndrome; lupus-like platelet reduction Symptomatic; occasional and / or massive blood loss; bone marrow hyperplasia; thrombocytopenia in patients with cachexia; thrombotic thrombocytopenic purpura, including cancer patients with thrombocytopenic purpura / hemolysis Urinary toxic syndrome, thrombotic microangiopathy, autoimmune hemolytic anemia; recessive jejunal perforation; simple red blood cell development, autoimmune thrombocytopenia; nephropathy epidemic; and rifampicin Related acute renal failure; paHs_Tr〇usseau thrombocytopenia, neonatal alloimmune thrombocytopenia; paroxysmal nocturnal urinary; blood changes in gastric cancer; early childhood hemolytic uremic syndrome; blood symptoms associated with viral infection, Including hepatitis A virus and CMV-related platelet reduction/in addition, some treatments for AIDS can also cause thrombocytopenia (such as AZT). Certain wound healing disorders can also benefit from an increase in the number of platelets. For the expected platelet deficiency, such as H4552.doc 51 1327149 due to future surgery, the compounds of the invention can be administered from a few days to several hours before platelets are required. For acute conditions, such as accidental and massive blood loss, the compounds of the invention may be administered with blood or purified platelets. The compounds of the invention may also be used to stimulate certain cell types other than megakaryocytes as long as such cells are found to exhibit mpl receptors. It is also within the scope of the present invention to be associated with such cells expressing mpl receptors, which are responsive to stimulation by mpi ligands. The compounds of the present invention are useful in any disease in which the production of platelets or platelet precursor cells is required or in which stimulation of the mpl receptor is required. Thus, for example, the compounds of the invention may be used to treat any condition in which a mammal requires platelets, megakaryocytes, and the like. Such conditions are described in detail in the following example sources: WO 95/26746; WO 95/21919; WO 95/1885 8; W0 95/21920, each of which is incorporated herein by reference. The compounds of the invention may also be used to maintain the viability or lifespan of platelets and/or megakaryocytes and associated cells. Accordingly, it is useful to include an effective amount of one or more such compounds in a composition comprising such cells. "Mammal" means any mammal, including humans, domestic animals, including dogs and deceased 'exotic and/or zoo animals including monkeys; laboratory animals including mice, rats and guinea pigs; farm animals including horses and cattle , sheep, goats and pigs; and analogues. Preferred mammals are human. V. Pharmaceutical Compositions The present invention also provides pharmaceutical compositions and methods of using the pharmaceutical compositions of the compounds of the present invention. Such pharmaceutical compositions may be administered by injection, or by oral, nasal spray, subcutaneous injection or other forms of administration, including, for example, by intravenous injection of 114552.doc-52·1J2/149, intradermal injection, intramuscular mist. Agent) or subcutaneous injection (including the main injection in the massroom 'intrapulmonary injection (such as medicine for storage for long-term release); lower, anal fistula, vagina, or read u) in the tongue or cornea. Treatment can be = surgical transplantation, such as transplantation under the spleen sac, large composition. In general, a pharmaceutical composition encompassed by the present invention or a plurality of pharmaceutical agents for a period of time includes an effective amount of a compound of the invention and hydrazine with 7^ another double mile.

Emulsifier 'adjuvant and / or; tincture, preservative 'solubilizer, liquid content (four) two ^ thinner; additives such as, heart:, salt) 'pH and ionic strength such as q detergent and solubilizer ( Such as Tween80 polysorbate 80), antioxidants (such as /, 1 ascorbic acid 'sodium metabisulfite), preservative imersol, the present methanol) and volume expansion alcohol); in the polymerized human 舲 』 σ 的 粒 粒Preparations such as polylactic acid, within the polyethylene glycol material or into the admixture f in the liposome. It is also possible to use a hyaluronic material: two may have the effect of promoting residence time in the cycle. The pharmaceutical compositions include other pharmaceutically acceptable liquid, semi-solid or solid diluents, which are used as pharmaceutical vehicles, excipients or vehicles, including but not limited to dioxetane ethyl sorbitan monolaurate , magnesium stearate, a few basic methyl and propyl formate, starch, sucrose, dextrose, arabinic acid 1 bow, mineral oil, cocoa butter and cocoa butter. Such compositions can affect the physical state, stability and rate of in vivo release of the proteins and derivatives of the invention, as well as the rate of body clearance. See, for example, Remingt〇n, s

Pharmaceutical Sciences, 18th Ed. (1990, Mack Publishing C〇·, -, PA 18〇42) pp. 1435-1712, which is incorporated by reference for preparation into a liquid form or dry powder form, such as frozen 114552.doc • 53· 1327149 Dry Open > Types of portability sustained release formulations are also contemplated, such as transdermal formulations. An oral solid dosage form contemplated for use in the present invention is generally described in the following documents: Remingtonis Pharmaceutical Sciences, 18 Ed. 1990 (Mack Publishing Co., Easton, PA 18042), Chapter 89, by way of citation Incorporated herein. Solid dosage forms include lozenges, capsules, pills, tablets or granules, cachets or granules. The vesicles or protein-like shells can also be used to formulate the compositions of the invention (e.g., protein-like microspheres, which can be used in U.S. Patent No. 4,925,673, and which can be used with various polymers. Derivatized (eg, US Patent No. 5,0 13,556). A description of possible solid dosage forms for treatment is Marshall, K., Modern Pharmaceutics, Edited by G. S.

Banker and C. T_ Rhodes Chapter 10, 1979, which is incorporated herein by reference. In general, the formulations comprise a compound of the invention, together with an inert ingredient which provides protection in the environment of the stomach and which releases the biologically active substance in the intestine. Oral dosage forms of the above compounds of the invention may also be specifically contemplated. If necessary, the compound can be chemically modified to render oral delivery effective. In general, a chemical modification that can be considered is the incorporation of at least a portion of the body to the compound molecule itself, wherein the portion of the body contributes to (4) inhibition of protein hydrolysis; (b) is taken up into the bloodstream from the stomach or intestine. It is also desirable to increase the overall stability of the compound and to extend its circulation time in the body. Examples of such partial bodies include: polyethylene glycol, copolymers of ethylene glycol and propylene glycol, carboxymethyl cellulose, dextran, polyethylene glycol, polyethylene retort, and polycholinic acid I14552.doc - 54- 1327149 (Abuchowski and Davis, Soluble Polymer-Enzyme Adducts, Enzymes as Drugs, Hocenberg and Roberts, eds., Wiley-Interscience, New York, NY, (1981), pp 367-383; Newmark, et al_, J_ Appl Biochem. 4:185-189 (1982)). Other useful polymers are poly-1,3-dioxolane and poly-i,3,6-trioxolane. For better medical use, as described above, it is a polyethylene glycol moiety.

For oral delivery dosages, it is also possible to use modified aliphatic amine salts such as N-(8-[2-hydroxybenzylidenyl]amino)octanoate (SNAC) as an enhancement of the invention. A carrier for the absorption rate of a therapeutic compound. The clinical effectiveness of the heparin formulation using SNAC has been demonstrated in the second phase of the trial by Emisphere Technologies. See U.S. Patent 5,792,45 1 » "Oral drug delivery composition and methods" 〇 The therapeutic agent can be included in the formulation as fine multiparticulates in the form of granules or pellets having a particle size of about 丨 mm. The adjustment of the substance used for the capsule

The formulation may also be a powder, a light compression suppository or even a lozenge. The therapeutic agent can be prepared by compression. Both colorants and flavor enhancers can be included. For example, the protein (or derivative) can be formulated (e.g., through a vesicle or microsphere capsule) and then further included in a ready-to-eat product, such as cold; East, a coloring and flavoring-containing beverage. TM·Reward for buying rare ..... - only -4 males can include n especially mannitol, lactose 'anhydrous lactose, sucrose, sucrose, modified glucomannan and starch. Some inorganic salts are also I14552.doc • 55· 1327149 u true fillings, including sodium trisodium sulphate, magnesium carbonate and sodium chloride. Some commercially available diluents are Fast_F1〇, Emdex, STA-RX 1500, Emcompress and Avicell. A disintegrant can be included in the formulation of the therapeutic agent to formulate a solid dosage form. Substances useful as disintegrants include, but are not limited to, house powders, including starch-based commercial disintegrants, Expl〇tab. Sodium starch glycolate, Amberlite, sodium carboxymethylcellulose, hyperamylopectin, sodium alginate, gelatin, orange peel, acid carboxymethylcellulose, natural sponge and bentonite can be used. Another form of disintegrant is an insoluble cation exchange resin. Powdered gums can be used as disintegrants and binders, and include powdered gums such as agar, paulownia gum or tragacanth. Alginic acid and its sodium salt can also act as disintegrants. Adhesives can be used to bind the therapeutic agents together to form a hard lozenge, and include materials derived from natural products such as gum arabic, tragacanth, minerals, and gelatin. Others include methyl cellulose (Mc), ethyl cellulose (EC), and carboxymethyl cellulose (CMC). Both polyvinylpyrrolidone (pvp) and hydroxypropylmethylcellulose (HPMC) can be used in an ethanol solution to granulate a therapeutic agent. An anti-friction agent may be included in the formulation of the therapeutic agent to avoid sticking during the formulation process. Lubricants can be used as a barrier between the therapeutic agent and the mold wall, which can include, but are not limited to, stearic acid, including magnesium stearate and calcium stearate, polytetrafluoroethylene (PTFE), liquid sorrow , vegetable oil and soil. Soluble lubricants such as sodium lauryl sulfate, magnesium lauryl sulfate, polyethylene glycols of various molecular weights, Carbowax 4000 and 6000 can also be used. Glidants that improve the mobility of the drug during formulation and aid in recombination during compression can be added. Glidants include powder killing 'talc, high temperature U4552, doc-56· 1327149 bauxite and hydrated strontium aluminate. In order to help the therapeutic agent dissolve in the aqueous environment, a surfactant can be added as a wetting agent. The surfactants include a variety of workers. Chess, (4) anthraquinone anion detergents, such as lauryl sulphide, succinyl acid, dioctyl succinate, and sodium dioctyl sulfonate. A cationic detergent can be used, and can include gasification or gasification. A list of potential nonionic detergents that can be included as a surfactant in the formulation is lauryl poly(vinyl alcohol 400), p〇ly〇xyl 4 decyl stearate, polyoxyethylidene hydrogenated castor oil 10, 50 and 60, glyceryl monostearate, polysorbate 40, 60, 65 and 80, sucrose fatty acid ester, f-based cellulose and carboxymethyl cellulose β these surfactants can be used alone or in a mixture of different ratios Present in a formulation of a protein or derivative. Additives which may increase the uptake of the compound are, for example, fatty acids, oleic acid, linoleic acid and linolenic acid. Controlling the release formulation is desirable. The drug can be incorporated into an inert matrix which can be released by diffusion or transfer mechanisms such as gums. The buffered metamorphic matrix can also be incorporated into the formulation, such as alginate, polysaccharides. Another controlled release form of this therapeutic agent is through a method based on the 〇r〇s treatment system (AlzaCorp.), which also encapsulates the drug into a semi-permeable membrane to allow water to enter and penetrate through a single small penetration. The opening drains the drug. Certain enteric coatings also have an extended release effect. Other coatings can be used in the formulation. These include various sugars that can be applied within the coated pan. The therapeutic agent can also be given in the form of a film-coated tablet, and the materials used in this example are classified into two classes. The first type is non-enteric substances, only JQ. Includes sulfhydryl cellulose, ethyl cellulose, hydroxyethyl cellulose, sulfhydryl Ethylene 114552.doc -57- 1327149-based cellulose, (tetra)-based cellulose, Propyl methylcellulose, slow methylcellulose, pn^vidone and polyethylene glycol. The second type of enteric material is phthalate for $. A mixture of materials can be used to provide an optimal film coating. Film coating can be carried out in a disc coater or in a fluidized bed or by compression coating. The invention also encompasses pulmonary delivery of the proteins of the invention (or derivatives thereof). The protein (or derivative) is delivered to the lungs of the mammal upon inhalation and through the pulmonary epithelial layer into the bloodstream. (Other reports on this include d al., Pharmaceutical Research 7: 5 65-5 69 (1990); Adjei et al International Journal of Pharmaceutics 63: 135-144 (199〇) (leuprolide acetate); Braquet et al., Journal of Cardiovascular

Pharmacology 13 (suppl. 5); s. 143-146 (1989) (endothelin-1); Hubbard et al., Annals of Internal Medicine 3.206-212 (1989) (1-anti-protease (i_antitrypSin) )); smith et al., J. Clin. Invest· 84:1145-1146 (1989) (1-proteinase); Oswein et al., " Aerosolization of Proteins ", Proceedings of Symposium on Respiratory Drug Delivery II, Keystone, Colorado, March, 1990 (Recombinant Human Growth Hormone); Debs et al., The Journal of Immunology 140: 3482-3488 (1988) (Interferon and Tumor Necrosis Factor) and! >1 is 2^&1., U.S. Patent No. 5,284,656 (granulocyte community stimulating factor). A wide range of mechanical devices designed to deliver therapeutic products to the lungs, including but not limited to nebulizers, graduated drug inhalers, and powder inhalers, all of which are 谙 are contemplated in the practice of the present invention. This is familiar to the skilled person at 114552.doc -58- 1327149. Some examples of commercially available equipment for use in the practice of the present invention are ultravent sprayers manufactured by Mallinckrodt, lnc., St. Louis, Missouri; Acorn® sprayers, by Marquest Medical Products, Englewo〇d, Made in Colorado; ventolin with graduated drug inhaler, manufactured by Glaxo Inc., Research Triangle Park, North Carolina; and Spinhaler powder inhaler, by Fis〇ns c〇rp, Bedf〇rd,

Made in Massachusetts.

All of these devices require the use of a formulation suitable for dispensing the compounds of the invention. Typically, each formulation will be specific to the type of device employed and, in addition to the diluents, adjuvants and/or carriers available in the treatment, may include the use of a proper aerosol base. The most advantageous compound of the invention is in the form of a granular form having an average particle size of less than 10 microns ' optimally being from 5 to 5 microns for most efficient delivery to the lung terminals.

Carriers include sugars such as Haiping, Sun # 凓 甘 甘 甘 甘, xylitol 'sucrose, lactose and sorbitol. Other ingredients that can be used for formulation include DPPC, DOPE, DSPC and DOPC. Natural or synthetic surfactants can be used. Polyethylene glycol can be used (even in addition to its use in derivatized proteins or the like). Glucan can be used, such as cycloglycan, a related enhancer. Cellulose and cellulose may be used to derivatize the hydrazine; with an amino acid, for example, in a buffer to prepare a mist. In addition, it is also contemplated to use vesicles, microcapsules or microspheres, inclusion complexes, or other types of carriers. U4D52.doc • 59-1327149 Formulations suitable for use with spray or ultrasonic type nebulizers typically comprise a compound of the invention dissolved in water at a concentration of from about G.1 to 25 mg of bioactive protein per ml of solution. Formulations may also include buffers and two early sugars (e.g., for protein f stabilization and osmotic pressure regulation). The nebulizer formulation may also contain a surfactant to reduce or avoid induced agglomeration of the surface of the protein f which is caused by the atomization of the aerosol. Formulations for use with graduated drug inhaler devices typically include a hair extension

Monthly. Subdivided powder of the powder suspended in the agent matrix I aerosol base by means of a surfactant can be any conventional substance used for this purpose, such as chlorofluorocarbons, hydrogen fluorocarbons, hydrofluorocarbons a compound, or a surfactant comprising a combination of trifluorofluoromethane, dihalofluorodifluorocarbon, dioxane, and m2-tetrahydroethane, or a combination thereof, including sorbitan trioleate Ester and soy indigo ester. Oleic acid can also be used as an interface activity |J. Formulations dispensed from a powder inhaler device include finely divided dry powders containing a compound of the invention, and may also include an adenoid, such as lactose 'sorbitol, sorbitol, mannitol, trehalose or xylitol, in an amount of Helps powder dispersion from the device, such as from 50 to 90% by weight of the formulation. Delivery via the # of the compounds of the invention is also contemplated. The nasal delivery allows the protein to enter the bloodstream directly after nasal administration of the therapeutic product without the need to deposit the product in the lungs. Formulations that are delivered via the nasal spray include those containing dextran or cyclamate^ that cover the use of other (four) carriers; 'medical treatments, including those used in the treatment of these diseases, by 114552.doc It is true that it will consider the factors that influence the age of the music, such as the patient's age, physical condition, weight, sex & 'A', Dip #饮&, the severity of the infection, the time of the operation and other Clinical factors. The compound of the present invention can be used in the first step of the treatment of the therapeutic cycle level of β-r. In another specific example, the compound of the present invention can be used once.仏田'·°. μ The skilled person can easily optimize the effective dose and the suitability according to the excellent-dry work and the clinical situation of individual patients. The frequency of administration depends on the pharmacokinetic parameters of the agent and the means of administration. The preferred pharmaceutical formulation can be determined by the skilled artisan according to the route of administration and the desired dosage. See, for example, Remingt〇n, s ph_ceutical fences, 18- Ed. (1990, Mack Pushing Co., Easton, ΡΛ 8042) pages 1435_1712, the disclosure of which is incorporated herein by reference. Such formulations can affect the physical state, stability, rate of internal release, and rate of in vivo clearance of the agent being administered. Depending on the route of administration, the appropriate dosage can be calculated based on body weight, body surface area or organ volume. Further precision in the calculations required to determine the appropriate agent for treatment with each of the above formulations may be routinely accomplished by the skilled artisan without undue experimentation, particularly in accordance with the dosage information and assays disclosed herein, and in Pharmacokinetic data observed in the above human clinical trials. The appropriate dose can be determined by using the existing assay for measuring the level of the liquid level and using the appropriate dose-response data. The final pharmacy will be decided by the attending physician's consideration of various factors that alter the action of the drug, such as the specific activity of the drug's severity of the injury and the patient's responsiveness, the patient's age' physical condition, weight's gender and diet, any The severity of the infection, the time of application 114552.doc 61 1327149 and other clinical causes. Lu, and Yan Yanjiu's progress will provide information on the appropriate dose levels and duration of treatment for various diseases and conditions.

The methods, compositions and compounds of the invention may also be used, either alone or in combination with other interleukins, soluble mpl receptors, hematopoietic factors, interleukin growth factors or antibodies for the treatment of other symptoms and platelet deficiency. Characteristics of the disease state. The compounds of the invention are expected to be useful in combination with common hematopoietic stimulators such as IL-3 or GM-CSF for the treatment of certain forms of platelet reduction. Other megakaryocyte stimulating factors such as meg-CSF, stem cell factor (SCF), leukemia inhibitory factor (10), 〇nec statin M) (〇SM), or megakaryocyte stimulating activity: other molecules may also Used with mpl ligands. Examples of 'interferon' or hematopoietic factors used for such co-administration include H ", n call' π-2 IL-3 ' IL_4 , IL_5 , IL_6 , lLu , community stimulating factor _ i (csf-i) , m_csf, scf, GM csf, granulocyte community stimulating factor ^ (10)), interferon _α (ΙΡΝ·α), alizarin, trace factory bribe -7, m8, IL.9, mi2, L13 \ human blood f Formation of serotonin, vascular endothelial growth factor (vegf), blood: = bone type formation protein seven bone type 2 = white f-2, bone type formation protein surface 3 'bone type formation protein shell · 4 , the monthly form of protein formation. 5, bone type formation protein formation protein-7, bone type formation protein [" morphological 烕 protein _9, 骨 剞 剞 疋 疋 疋 amp amp # # # # # # # # # # # # 〇, bone type formation I14552.doc •62· Protein-11, bone form formation into protein 贞-12, bone type detachment 忐 a a quality-13, bone type formation egg protein 1C ^ 白贝_14 , bone type formation protein 15, bone type formation protein 森贝 _ 质 体, bone type formation protein receptor IB, brain derived god management And ciliary neurotrophic factor by Bian 'acceptor fiber Maoying Ji, L. cytokine induced by the primary. Neutrophil-like leukocytic chemokine 1, interleukin-induced neutrophil-induced serotonin 2, endothelial cell growth factor, endothelin 1, dermal growth factor, epithelial neutrophil leukocytosis, fibrils Cell growth factor 4, 镳 her# ^ ε ε fibroblast growth factor 5, fibroblast growth factor ό, fibroblasts + tian gong g 飑 growth factor 7, fibroblast growth factor 8, fibroblast birth county On the field. l Sub 8b, fibroblast growth factor 8c, fibroblast growth factor 9, Min Shi # Λ & fibroblast growth factor 1 纤维, fibroblast growth factor acid, fibroblast growth factor test, glial, - Cell line Hesheng neurotrophin receptor i, glial cell line derived god: strophic factor 2, growth-related peptone, growth-related protein, growth-related protein, growth-related protein f, heparin Bonded growth factor, hepatocyte growth factor, hepatocyte growth factor receptor 'like insulin growth factor!, like the growth factor receptor, the like growth factor Π 'like the growth factor binding Protein f, keratinocyte growth factor, leukemia inhibitory factor, leukemia inhibitory factor receptor, nerve growth factor, nerve growth factor receptor, neurotrophin _3, neurotrophin-4, placental growth factor, placental growth factor? Platelet-derived endothelial cell growth factor, platelet-derived growth factor, platelet-derived growth factor A chain platelet-derived growth factor AA, platelet-derived growth factor AB platelet-derived growth factor B chain, platelet-derived growth factor BB, 114552 .doc •63 · 1327149 Platelet Derived Growth Factor Receptor's Platelet Derived ~ Long Factor Factor, Portuguese

B cell growth stimulating factor, stem cell factor receptor, Ding Ν ρ, including Ding delete, TNF1, TNF2, transforming growth factor, transforming growth factor growth factor 1, transforming growth factor K2, transforming growth factor 2, transformation Growth factor 3, transforming growth factor 5, recessive transforming growth factor! , transforming growth factor binding protein! , transforming growth factor-binding protein, transforming growth factor binding protein m, tumor necrosis factor receptor type, tumor necrosis factor $body type II, urokinase ▲ plasminogen activator receptor, vascular endothelium Growth factors, and chimeric proteins and biological or immunologically active fragments thereof. It is also useful to simultaneously or sequentially administer an effective amount of soluble mammal = receptor, which has the effect of causing megakaryocyte segmentation to form gray platelets when megakaryocytes become mature. Thus, administration of a compound of the invention (to increase the number of mature megakaryocytes) followed by administration of a soluble mpl receptor (with de-ligand activity and production of platelets by mature megakaryocytes) is expected to be a particularly effective pathway for stimulating platelet production. The above dosages can be adjusted to compensate for such additional ingredients in the therapeutic composition. The progress of the treated patient can be monitored by conventional methods. In the case where the compound of the present invention is to be added to a composition of platelets and/or megakaryocytes and related cells, the amount to be included is usually determined experimentally by techniques and assays known in the art. An example of this amount ranges from 0-1 micrograms to 1 milligram of the compound of the invention per 1 〇 6 cells. It will be appreciated by those skilled in the art that the application to the particular problem or circumstance is within the capabilities of the skilled artisan. Examples of the products of the invention and representative methods for their isolation, use and manufacture are described in the following paragraph D4552.doc-64- 1327149. EXAMPLES Exemplary methods of making and identifying the compounds disclosed herein are described below. Example 1 1. Composition of a secondary peptide library A. Preparation of an electrically competent E. coli cell: overnight culture of large intestine rod (TG1 strain, Amersham Pharmacia Biotech, Piscataway, New Jersey) at 37 °C 2xYT medium (1.6% Bacto Tryptone, 1% yeast extract, 85.5 mM NaCl) in ι〇 ml was prepared. One milliliter of this overnight culture was used to inoculate 1 liter of 2xYT medium containing 0.4% glucose and 1 mM MgCh, and the one liter culture was grown to OD6 at 37 ° C in an oscillator = 0.8. The culture was cooled on ice for 15 minutes and centrifuged at 4 ° C for 20 minutes at 4 ° C (Beckman JA-10 rotor). The pellet was resuspended in 500 ml of ice-cold 10% glycerol solution, and the resulting mixture was centrifuged at 4000 rpm for 20 minutes at 4 °C. The bacterial pellet was resuspended again in 500 ml of ice-cold 10% glycerol solution, and the resulting mixture was again centrifuged at 4000 rpm for 20 minutes at 4 °C. The pellet was then resuspended in 25 ml of ice-cold 10% glycerol solution. The concentrated bacterial sample was transferred to an ice-cold 50 ml conical tube and centrifuged at 3500 rpm for 5 minutes at 4 °C in a tabletop centrifuge (Beckman CS-6R). Resuspend the cell pellet in a small volume of ice-cold glycerol solution and immediately freeze 100 or 300 microliters of bacterial stock in ethanol/dry 114552.doc •65· 1327149 ice bath and store at -80° C in the freezer. B. Modification of pCES1 vector

PCR was carried out using Extend Long Template PCR Systems (Roche Diagnostics Corp., Indianapolis, Indiana) with 1 μg of pCES1 vector (TargetQuest Inc.) as a template. The volume of the PCR mixture was 100 μl, containing 1 x PCR buffer, two primers of 200 nM each, 5, -CAAACGAATGGATCCTCATTAAA GCCAGA-3' and 5'-GGTGGTG CGGCCGCACTCGAGACTGTT GAAAGTTGTTTAGCA-3', 200 nM dNTP , 3 units of Tag DNA polymerase. The following procedure was run using the TRIO-Thermoblock (Biometra) PCR system: 5 minutes at 94 ° C; 30 cycles (30 seconds at 94 ° C, 30 seconds at 50 ° C, 45 seconds at 72 ° C); 72 ° C for 10 minutes; cool to 4 °C. The PCR product was run on a 1% agarose gel and purified using QIAGEN Spin Column according to the manufacturer's instructions (QIAGEN Inc., Valencia, California). The second PCR reaction was carried out under the same PCR conditions as described above using 5 μl of PCR product and two primers of 50 nM each of 5'-CAAACGAATGGATC CTCATTAAAGCCAGA-3' and 5丨-AACAAAAAGTGCACAGGG TGGAGGTGGTGGTGCGGCCGCACT-3'. The PCR product and the original pCES1 vector were separately contained in 100 μl of a reaction solution containing lx NEB2 buffer, 60 units of ApaLI (New England Biolabs, Beverly, Massachusetts), 60 units of BamHI (New England Biolabs). Digestion was carried out at 37 ° C for 1 hour. The two digested DNAs were ligated with QIAGEN Spin Column pure 114552.doc • 66 - 1327149 and incubated in 40 μl of reaction solution overnight at room temperature containing 1X ligation buffer and 4 units. T4 DNA ligase (New England Biolabs). The vector was transfected into E. coli and at 37. (: cultivate overnight. Select the isolated single colony and purify the plastid with qIAGeN Spin Column. The correct insertion line is determined by DNA sequence. C. Preparation of vector DNA One microgram of modified pCES1 vector DNA (1B segment) ) Transformed to 1 〇〇 microliter of electricity competent TG1 E. coli (ία segment), which uses Gene Pu丨ser Π (BIO-RAD, Hercules, California), set to 2500 volts (V), 25 °F ' With 200 ohms, the transformed bacterial sample was immediately transferred to a 900 μl SOC (2% pancreas '0.5% yeast extract, 1 mM sodium sulphate, 2.5 mM chlorpyrifos) In a test tube of 20 mM glucose, 1 mM magnesium sulphate, 1 mM magnesium sulphate, and the culture was shaken for 1 hour at 37 C. The cells were then dispersed into 2 xytaG (2xYT ' Containing 1 μg/ml ampicillin and 2% glucose) in a lipid plate and incubating overnight at 37 ° C. Using a single flora at 37 ° C, inoculate 1 liter of 2xYTAG medium to shake the whole Night. The plastid vector DNA was purified using QIAGEN Plasmid Maxi Kit according to the manufacturer's instructions. Digested DNA of body

50 μg of vector DNA (Phase 1C) was digested overnight at 37 ° C in 400 μl of a reaction mixture containing 2,200 units of ApaLI and 200 units of xh〇I in lx NEB buffer. The restriction digestion reaction solution was incubated at 37 ° C overnight and analyzed in a prefabricated 1% agarose gel (Embi Tec, San Diego, California) 114552.doc - 67 · 1327149. The linearized vector DNA was cleaved from the gel and extracted using a QIAquick Gel Extraction Kit (QIAGEN Inc.) according to the manufacturer's instructions. E. Preparation of an oligomeric nucleocapnic acid library A two-oligonucleotide acid library (immobilized and additive-added) was designed. Synthetic library oligonucleotide

5'-CACAGTGCACAGGGTNNKNNKNNKNNKGGTCCTACT CTGMRKSARTGGCTGNNKNNKNNKNNKNNKNNKCATTC TCTCGAGATCG-3', and the additive oligonucleotide 5'-CACAGTGCAC-AGGGTNNKNNKNNKNNKggKcc-KacKc tKNNKNNKtgKNNKNNKNNKNNKNNKNNKNNKCATTCTC TCGAGATCG-3, (lowercase letters represent a base of 70% and 10% a mixture of one of the other three nucleotides). These oligonucleotides were used as templates in the polymerase chain reaction, respectively.

The Expand High Fidelity PCR System kit (Roche Diagnos sties Corp.) was used in the PCR reaction. Each PCR reaction is in a volume of 100 μl and contains 10 nM of oligomeric nuclei: y: acid bank, IX PCR buffer, each primer of 300 nM 5'-CACAGTGCACAGGGT-3' and 5,-TGATCTCGAGAGAATG -3 ', 200 nM dNTPs, 2 mM calcium carbonate and 5 units of Exp and polymerase. A thermal cycler (GeneAmp PCR System 9700, Applied Biosystem) was used to perform the following procedure: 5 minutes at 94 ° C; 30 cycles (30 seconds at 94 ° C, 30 seconds at 55 ° C, 45 seconds at 72 ° C) ); 7 minutes at 72 ° C; cooled to 4 ° C. The free nuclear acid was removed using the QIAquick Nucleotide Removal Kit (QIAGEN Inc.) according to the manufacturer's instructions 114552.doc -68- 1327149. F. Digestion of the oligonucleotide library 5 micrograms of each PCR product (paragraph 1E) in 400 microliters of reaction solution containing 1 x 2,0 units of ApaLI and 200 units of Xhol in lx NEB buffer Medium 'digested overnight at 37 ° C. The digested DNA was separated on a 3% agarose gel (Embi Tec). The target DNA bands obtained from each reaction solution were excised from the gel and extracted with QlAquick Gel Extraction Kit to 0 G. The vector linearization vector (1D segment, 25 μg) and each of each was ligated with an oligonucleotide library. The digested PCR product (1F segment '5 μg) was ligated overnight at i6 ° C in 400 μl of reaction solution containing ΐχ NEB ligation buffer and 80 units of T4 DNA ligase. The ligation product was incubated at 65 °C for 20 minutes to inhibit activation of DNA ligase, and then incubated with 8 units of Notl for 2 hours at 37 °C to minimize autologous ligation of the vector. The ligation product was then purified using standard phenol/gas imitation extract (Moleeular Cloning, Maniatis et al 3rd Edition) and resuspended in 30 microliters of water. H. Electrical Penetration Transformation Ten electrical penetration reactions were performed for each pool. In each transformation, 3 microliters of ligation vector DNa (1G segment) and 300 microliters of TG1 cells (1A segment) were mixed in a 0.2 cmi tube (BI0-Rad). The resulting mixture was pulsed with a Gene pulser(R), set to 25 volts, 25 uI^2 ohms. The transformed bacteria samples from ten electroporation reactions were combined and transferred to a solution containing 27 ml. In the SOC flask, at 37. (: Incubate a small U4552.doc -69· 1327149 and then add the cells to 170 ml of 2xYTAG and grow for 3 hours at 37 ° C. The cells are at 5 ° rpm at 4 ° Centrifuge for 1 minute at C. Then resuspend the cell pellet in 1 mL of 15% glycerol/2χγτ 'stored at -8 0 ° C. This is the initial stock solution of the library. The titration shows the immobilized library and In the addition library, the independent size of the library is 丨〇X 1 〇9 and the independent transformation of 2·4 X 1〇9. 2·Expansion of the library 制造. The initial library cell stock (1 Η) was inoculated with 1300 ml (for immobilization) and 2600 ml (for dosing) 2xYTAG medium, so that the starting 〇E>6qd=〇.1. Let the culture at 37 Gently grow at °C for several hours until 〇D600=〇.5. Take a 120 ml portion of the immobilized reservoir and a 240 ml portion of the pooled solution in separate flasks at 37. Growth for 2 hours. Centrifuge these subcultures at 5000 rpm (Beckman JA-14 rotor) at 4 C for 1 min and resuspend the bacterial pellets at 1 〇 (for each library) within 15% glycerol/2xYT and stored at -80 ° C. B. Phage induction of M13K07 helper phage (Amersham Pharmacia 31 (^ (; 11) to 〇〇 6〇〇 =0.5 was added to the remaining bacterial culture (2:8) to a final concentration of 3 X 109 pfu/ml. The helper bacteria were infected with bacteria for 30 minutes at 37 °C without shaking and 3 min with slow shock infection. The infected cells were centrifuged at 5000 rpm for 10 minutes at 4 ° C. The cell pellets were resuspended in 1300 ml (immobilized pool) and 2600 ml (doped pool) of 2xYTAK (2YT and 1 μg/g/ ML ampicillin and 4 〇 micrograms/ml card lU552.doc • 70· 1327149 kanamycin » oscillate overnight at 37 ° C to produce phagemid production ° C. phage harvesting bacterial culture (2B) was centrifuged at 5000 rpm for 1 C at 4 ° C. Transfer the supernatant to a new bottle, add 0.2 volume of 20 〇 / 〇 PEG / 2.5 Μ sodium chloride, and on ice Incubate for 1 hour to precipitate the bacterium of the bacterium. The precipitated phage is at 8000 rpm at 4. (: 20 points under centrifugation) And carefully resuspended in 1 ml of cold PBs. The remaining cells were separated by centrifugation of the sputum plastid solution at 8000 rpm for 10 minutes at 4 °C, and 0.2 volume of 20% PEG/2.5 加入 was added. Gasification of sodium to precipitate phage is further purified. The sputum plastids were centrifuged at 8 ° rpm for 20 minutes at 4 ° C, and the sputum plastids were resuspended in 12 ml of cold pbs. 4 ml of 60% glycerol solution was added to the phage solution and stored at _8 〇t. The titer of the plastids was determined by standard procedures (M〇lecular Cloning, Maniatis et al. 3rd Edition). 3 · Selection of human MPL-binding phage A. Biotinylation of human MPL 1 mg of recombinant human MPL using ugly B-1^ spit 8111 &->^8-1^-

The Biotinylation Kit (PIERCE, Rockford, Illinois) was biotinylated according to the manufacturer's instructions. B. Immobilization of MPL receptors on magnetic spheres MEL after biotination (3A) is immobilized on Dynabead Μ·28〇

On Streptavidin (DYNAL, Lake Success, New York), the concentration was 1 microgram MPL/100 microliters of pellets from the manufacturer. After the beads 114552.doc 71 1327149 were directed to one side of the tube with a magnet and the liquid was aspirated, the magnetic beads were washed twice with saline buffered saline (PBS) and resuspended in phosphate buffered saline. The biotinylated MPL protein was added to the washed magnetic beads at the same concentration and incubated at room temperature for 1 hour. B S A was then added to a final concentration of 2% to block the MPL coated magnetic beads and incubated at 4 °C overnight. The resulting MPL coated magnetic beads were then washed twice with PBST (phosphate buffered saline and 〇.〇5 % Tween-20) and subjected to a selection step. C. Using MPL coated magnetic beads for selection. Use 100 ml of 2% BSA in PBS to block about 100 times of library equivalent phage (2C, Ulcncfu for immobilization, 2.4xlOncfu for doping Library) 1 hour. A negative selection step was performed on the blocked phage sample, which was added to the empty beads (the same beads as the 3B segment, but not coated with MPL), and the mixture was rotated at room temperature. Cultivate for 1 hour. The phage-containing supernatant was withdrawn by a magnet and transferred to a new test tube containing MPL coated magnetic beads (Section 3B), and the mixture was incubated at room temperature for 1 hour. After removing the supernatant, the phage-bound magnetic beads were washed 10 times with PBST and washed 10 times with PBS. The sputum plastids were then solubilized in 1 ml of 100 mM triethylamine solution (Sigma, St. L〇uis Missouri) in a rotator for 10 minutes. The pH of the phage-containing solution was neutralized by adding 0.5 ml of 1 MTdS-HCl (Ph7.5). Using the obtained sputum plastids, 5 ml of newly grown butyl (1) fine (OD6 (10) was about 0.5) for 30 minutes was inoculated without shaking at 37 t and then infected by slow shaking for 3 minutes. All infected TG1 cells were plated on a large 2xYTAG plate and incubated overnight at 30 °C. U4552.doc -72- 1327149 D. Phage induction and harvesting 10 ml of 2xYTAG medium was added to the plate (3C section) to resuspend the floating T G1 cells. All T G1 cells were collected in a test tube, and 250 μL of the cells were added to 25 ml of 2xYTAG and grown at 37 ° C until 0 〇 6. 〇 = 0.5. The M13K07 helper bacteria were added to a final concentration of 3 X 109 cfu/ml, and incubated at 37 ° C for 30 minutes without shaking, followed by slow shaking for 30 minutes. The cells were centrifuged at 5 ° rpm for 10 minutes at 4 ° C and resuspended in 25 ml of 2x YTAK. These bacteria were shaken and grown overnight at 30 °C. The induced phage plastids were harvested and purified as described in Section 2C. E. The second round selection The second round of selection is performed as described in paragraphs 3B to 3C, with only the following differences. A phage solution obtained from the 3D fraction of about 0.5 ml of the aliquot was used as the feed phage. Only 0.1 microgram of biotinylated MPL (3A segment) was used to coat Dynabead M-2 80 Streptavidin. The bacterium-bonded-magnetic beads were washed 16 times with PBST, wherein the final wash included incubation in PBST for 30 minutes at room temperature. The magnetic beads were washed 10 times with PBS. 4. Analysis of the colonies A. Preparation of the main board The single colony selected from the second round was taken out and inoculated into 96-hole plates, each containing 120 μl of 2xYTAG. The 96-well plates were incubated overnight at 30 ° C in an incubator. 40 ml of 60% glycerol was added to each well and stored at -80 °C. B. Phagocytic ELISA assay 114552.doc -73· 1327149 Inoculate approximately 3 microliters of cells (4 A segments) obtained from the autonomous plate in a new 96-well plate containing 12 μL per well 2xYTAG, the new cell plate was grown at 37 ° C until 〇d6〇q was approximately 0.5. 40 ml of 2xYTAG containing M13K07 helper phage (1.2 X 1010 cfu/ml) was added to each well 'at 37 The 96-well plate was incubated for 30 minutes without shaking at ° C, and then slowly shaken for 30 minutes. The plate was centrifuged at 2000 rpm (Beckman CS-6R tabletop centrifuge) for 10 minutes at 4 °C. Remove the supernatant from the hole and resuspend each cell pellet with 1 60 μl of 2 χΥΤΑΚ per well. The plates were incubated overnight at 3 °C for phage display. Recombinant human MPL at a concentration of 5 μg/ml in 0.1 M carbonate buffer and pH 9.6 was applied to 96-well Maxisorp plates (NUNC) overnight at 4 °C. As a control, BSA (Sigma) was coated on a separate Maxisorp plate at a concentration of 5 μg/ml. On the next day, overnight cell cultures were centrifuged at 2000 rpm for 10 minutes at 4 °C. Transfer 20 ml of the supernatant from each well to a new 96-well plate containing 180 μl of 2% BSA/PBS solution per well. The resulting mixture was incubated at room temperature for 1 hour to block the phage. At the same time, the MPL coated panels were used at 200 microliters 2°/. The BSA/PBS solution/hole was clogged for 1 hour at room temperature. The BS A solution was removed and each well was washed three times with PBST solution. After the last washing step, 50 μl of the blocked plastid solution was added to the hole of each MPL coated plate and the control plate, and incubated for 1 hour at room temperature. The liquid was removed and each well was washed three times with PBST solution. Fifty microliters of HRP-conjugated anti-M13 mAb (Amersham Pharmacia Biotech) was added to the holes of each MPL coated plate and the holes of the control plate at a 1: 15,000 dilution of 114552 doc • 74· 1327149. Incubate for 1 hour at room temperature. The liquid was removed again and each well was washed three times with PBST solution. Fifty microliters of LumiGLO chemiluminescent matrix (Kirkegaard & Perry Laboratories, Gaithersburg, Maryland) was added to the wells and each well was read with a Luminoskan Ascent DLRearly machine (Labsystems, Franklin, Massachusetts). C. Sequencing of phage populations PCR was carried out using 1 μl of bacteria (4A) of each well of the autonomous plate as a template. The volume of each PCR mixture is 20 μl, containing lx PCR buffer, two primers of 300 nM each 5'-GTTAGCTCACTCATTAGG CAC-3' and 5'-GTACCGTAACAC TGAGTTTCG-3', 200 nM dNTP, 2 mM The gasification is about 5 units of tag DNA polymerase (Roche Molecular Biochemicals). The following program was run using The GeneAmp PCR System 9700 (Applied Biosystem): 5 minutes at 94 ° C; 40 cycles of [45 seconds at 94 ° C, 45 seconds at 55 ° C, 90 seconds at 72 ° C]; 72 ° C for 1 minute; cool to 4 °C. The PCR product was purified using QlAquick 96 PCR Purification Kit (QIAGEN Inc.) according to the manufacturer's instructions. "All purified PCR products were using primer 5, -CGGATAACAATTTCACACAGG-3' with ABI 3770 Sequencer (perkin Elmer) according to the manufacturer's instructions. Perform sequencing. 5. Sequence grading will correlate the peptide sequence translated from the above nucleus: y: acid sequence with ELISA data. ° High OD readings are shown in the 1<1^ coating hole in the sho, and low in the BSA coating hole. The 0D read strain is considered a candidate for further study. The sequence of 114552.doc -75· 1327149 is also considered to be a candidate for further study. The phage population selected based on these criteria was further identified in an ELISA titration experiment. See Figure 9 (£18 18 dose-response curves for selected phage colonies). Example 2 Preparation of peptides All peptides were prepared by existing stepwise solid phase synthesis. Merrifield (1963), J. Am. Chem. Soc. 85:2149. Steward and

Young (1969), Solid Phase Peptide Synthesis. Fmoc-protected amino acids were used as constituent elements, and ABI or Symphony peptide synthesizers were used to construct the peptide chain. Typically, the peptide synthesis begins with a pre-filled Wang resin to produce a peptide having a free carboxylic acid at the c-terminus (or a Rink resin can be used to produce a peptide having a c-terminal guanamine function). Fmoc removal was performed using the standard piperidine procedure. A chemical reaction of a urea cation (e.g., HBTU) or a carbodiimide (e.g., DCC/HOBt) is used to facilitate coupling. Side chain protection *g: Glu (0-t-Bu), Asp (0-t-Bu), Ser (t-Bu), Thr (t-Bu), Arg (Pbf), Asn (Trt), Gln ( Trt), His(Trt), Lys(t-Boc), Trp(t-Boc) and Cys(Trt). The final deprotection and cleavage of the peptide-based resin is 2.5% water, 5% phenol, 2.5% triisopropyl decane and 2.5% thioanisole or sulfhydryl at room temperature. Ethyl trifluoroacetic acid (TFA) was carried out for 4 hours. After removing the trifluoroacetic acid, the cut peptide was precipitated with cold anhydrous diethyl ether. For the peptide containing the disulfide bond, the formation of the cyclic product was carried out directly on the crude material using 15% DMSO (ρΗ7·5) in water. All crude peptides were purified by reverse phase HPLC and analyzed by ESI-MS and amino acid analysis to determine the peptide of the purified peptide. 114552.doc • 76· 1327149. Example 3 Preparation of TMP-Fc Peptidomimetic Compounds Several peptides were selected to be expressed as a peptide-Fc fusion (i.e., Fc followed by the C-terminus of the peptide) (c-terminal fusion). The DNA sequence encoding the Fc region contained in human IgG1 fused to each of the τρο mimetic peptides was placed under the control of the luxPR promoter gene contained in the plastid expression vector pAMG2 1 as follows. The plastid encoding tmpkFc (Amgen strain, #3788) was altered to include an ApaLI site and a xh〇I site for selection of the short peptide obtained from paired oligonucleotides. The primers 2396-69 were used to add ApaLI and Xhol restriction enzyme sites. The PCR system was carried out on a 3788 DNA template using Expand Long Polymerase ' at 2396-69 and General 3, primer 191-24. The primer sequence is as follows:

2396-69 ACAAACAAACATATGGGTGCACAGAAAGCGG CCGCAAAAAAACTCGAGGGTGGAGGCGGTGG GGACA

191-24 GGTCATTACTGGACCGGATC The resulting PCR fragment was digested with Ndel and BsrGI, gel purified, and used as an insert. The plastids obtained from #3788 strain were also digested with Ndel and BsrGI, gel purified, and used as a carrier. The vector and insert were ligated together and the resulting ligation mixture was electroporated into GM221 cells (see below). A single colony is removed, plastid DNA is prepared, and the DNA is sequenced. One of the resulting plasmids, 200003 180, was shown to have the correct DNA sequence and was used as a vector for constructing the TMP-Fc fusion. This carrier is shown in Figure 6. 114552.doc • 77- 1327149 The plastid 200003180 was digested with human 卩&1^1 and 乂11〇1 and used as a carrier. Each pair of oligomeric nucleosides (see Figure 7) was paired to form a double strand of ApaLI and Xhol sticky ends. These molecules are ligated into a vector to produce the target fusion protein. The ApaLI and Xhol fragments of each corresponding oligomeric nucleoside pair are presented in Figure 7. TMPs 1-23, 25, 26 and 28 are all expressed as C-terminal fusions. Example 4

Preparation of Fc-TMP Peptidomimetic Compounds Some peptides behave as Fc-peptide fusions (i.e., Fc followed by the N-terminus of the peptide) (N-terminal fusion). The plastid encoding Fc-TMP 1 (Amgen strain, #3728) was modified to include an ApaLI site and an Xhol site for the selection of the short peptide obtained from the cooled oligonucleotide oligonucleotide "Introduction 2396- The 70 line was designed to incorporate ApaLI and Xhol restriction enzyme sites. The PCR system was carried out on a 3728 DNA template using Expand Long polymerase at 2396-70 and the universal 5' primer 1209-85. The primer sequence was as follows: 1209-85 CGTACAGGTTTACGCAAGAAAATGG 2396-70 TTTGTTGGATCCATTACTCGAGTTTTTTTGCGG CCGCTTTCTGTGCACCACCACCTCCACCTTTAC The resulting PCR fragment was digested with BsrGI and BamHI, gel purified, and used as an insert. The plastid obtained from #3728 strain was also digested with 85 "1 and 6&11^1, gel purified, and used as a carrier. The vector and insert were ligated together and the resulting ligation mixture was electroporated into GM22 1 cells. A single colony is taken, plastid DNA is prepared, and DNA is sequenced. A resulting plastid, 200003182 (Figure 8), shows the correct DNA sequence and is used as a construct 114552.doc •78-

The vector used for the Fc-TMP fusion. The plastid 200003182 was digested with eight ports of 3 [1 and 乂11〇1 and served as a carrier. A cooled conjugated oligomer having ApaLI and Xhol viscous ends was attached to the vector to produce the target fusion. Take! TMP20, TMP24, TMP27, TMP29 spear TMP30 is an N-terminal fusion. Transformation Each of the above-described conjugates was transformed into host species GM22 1 by electroporation as described below. The selection is screened for the ability to produce recombinant protein products and possess the ability to fuse with the correct nucleotide sequence. The pAMG21 expression plastid PAMG21 is available from ATCC under accession number 98113, and its registration period is July 24, 1996. GM221 (Amgen Host Strain #2596)

Amgen host strain #2596 is the E. coli K-12 strain which has been modified to include the temperature-sensitive lambda inhibitor cI857s7 in the early ebg region and the lacIQ inhibitor in the late ebg region (68 minutes). The presence of these two repressor genes allows the host to be used in a variety of expression systems, although the two repressors are independent of the performance of the IuxPr. Untransformed hosts are not resistant to antibiotics. The ribosome binding site of the cI8 5 7s7 gene has been modified to include an enhanced RBS. It is inserted into the ebg operon, which is between the nucleotide positions 1170 and 1411, numbered M64441Gb_Ba in the Genbank accession number, in which the intervening ebg sequence is deleted. 114552.doc -79- 1327149 The construct was delivered into the chromosome using recombinant phage, MMebg-cI857s7 enhanced RBS #4 into F'tet/393. After recombination and resolution, only the above chromosomal inserts remain in the cells. It was renamed to F'tet/GMIOl. The F'tet/GMIO1 is then modified by transporting the laciQ construct into the 6匕§ operon between the nucleotide positions 2493 and 2937, and the access number is numbered M64441G b_B in 〇61^&11 a, which removes the intervening ebg sequence. The construct was delivered to the chromosome using recombinant phage which was AGebg-LacIQ#5 into F'tet/GMIOl » after recombination and resolution. Only the above chromosomal insert remained in the cell. Rename it to F’tet/GM221. The F'tet gene epithet was incubated with the acridine orange at a concentration of 25 μg/ml in LB. The recovered strain was identified as a tetracycline-sensitive strain and stored as GM221. Performance Cultures of GM221, which can express each fusion, were grown in Luria Broth medium at 37 °C. The step of displaying the gene-induced gene product from the iuxPR promoter is carried out by adding a synthetic autoinducer N_(3-ketohexylenyl)-DL-homoserine to the final concentration of 2 〇 ng/ml. And at 3 7 C for another 3 hours to reach the winner. After 3 hours, the fine nutrients were observed through a microscope to observe the presence of inclusion bodies, and then centrifuged to collect them. Refractive inclusion bodies were observed in the induced cultures, indicating that the fusion protein is most likely to produce an insoluble fraction in E. coli. The pellets were directly lysed by resuspending in LaemmU sample buffer containing 1% cold-mercaptoethanol and analyzed by SDS-PAGE. A strong Coomassie staining band (approximately 30 kDa) of appropriate size was observed for each protein • S0·114552.doc 1327149. Example 5 Purification of peptide compounds The cells were disrupted by high pressure homogenization (2 passes at 14,000 PSI) in water (1/10), and the inclusion bodies were harvested by centrifugation (4,200 rpm in J-6B). ,1 hour). The inclusion bodies were dissolved in 6 Μ 胍, 50 mM Tris, 8 mM DTT at pH 8.7 for 1 hour. The dissolved mixture was diluted 20-fold to 2 脲 urea, 50 mM Tris, 160 mM arginine, 3 mM cysteine, pH 8.5. The mixture was stirred overnight at low temperature and then the mixture was concentrated by ultrafiltration for about 丨〇 times. It was then diluted 3 times with a 10 mM Tris, 1·5 guanidine urea, pH 9. The pH of the mixture was then adjusted to pH 5 with acetic acid. The precipitate was removed by centrifugation and the supernatant was loaded onto a Sp_Sephar〇se Fast Fl〇w column equilibrated with 20 mM NaAc, 100 mM sodium chloride, pH 5 (10 mg/ml protein) Loading capacity, at room temperature). Protein was eluted using a 2 column column volume with 100 mM sodium chloride to 5 mM sodium sulphate gradient in the same buffer. Dilute the combined fraction obtained from the column by 3 times.

It was loaded onto an SP-Sepharose HP column, 20 mM NaAc, 150 mM sodium hydride, pH 5 (10 mg/ml protein loading, room temperature). Protein was eluted using a 20 column volume with 15 mM sodium chloride to 400 sodium chloride gradient in the same buffer. The peaks were combined and filtered. Example 6 Peptide affinity binding study U4552.doc 1327149 Experiments were carried out at room temperature using BIACORE 3000 to determine the binding affinities of several TMP peptides (TMP1-TMP23). Hu-mpl was immobilized on the surface of the sensor wafer (CM5) using an amine coupling procedure (activated with NHS/EDC and blocked with ethanolamine). A 0.78 nM to 100 nM TMP peptide was injected onto the Hu-mpl surface. BIACORE Working Buffer was PBS plus 0.005% Surfactant P20. The sample was also injected onto a blank surface as a control. The experimental data was analyzed using the BIAEVALUATION 3.1 suite of software. As discussed above, in order to better mimic the phage environment in which the peptide is selected, and to eliminate the charged amine and carboxy termini of the 18 amino acid preferred peptides (TMP2-TMP30) from the receptor, Amino acid "cover" is added to each carboxy terminal and amine end of each peptide: glutamine-cysteine (QC) to the amine end, and histidine-silyl group (HS) to the carboxy terminus, changing the length of each peptide to 22 amino acids. Since the peptide affinity is known to increase with the length of the peptide, the signature biologically active 14 amino acid sequence (SEQ ID NO 1) is also increased to a total of 22 amino acids. However, the bioactive region of each peptide remains unchanged, as shown in bold in the table below. TMPNo. Peptide sequence KD (nM) with respect to the TMP1 affinity TMP1 SAQGIEGPTLRQWLAARALETV 5.40 TMP2 QGGAREGPTLRQWLEWVRVGHS 1.60 3.38 TMP3 QGRDLDGPTLRQWLPLPSVQHS 45.00 0.12 TMP4 QGALRDGPTLKQWLEYRRQAHS 0.86 6.28 TMP5 QGARQEGPTLKEWLFWVRMGHS 6.66 0.81 TMP6 QGEALLGPTLREWLAWRRAQHS 0.37 14.59 114552.doc -82 · 1327149

TMP7 QGMARDGPTLREWLRTYRMMHS 1.20 4.50 TMP8 QGWMPEGPTLKQWLFHGRGQHS 23.20 0.23 TMP9 QGHIREGPTLRQWLVALRMVHS 1.67 3.23 TMP10 QGQLGHGPTLRQWLSWYRGMHS 1.22 4.43 TMP11 QGELRQGPTLHEWLQHLASKHS 35.90 0.15 TMP12 QGVGIEGPTLRQWLAQRLNPHS 5.20 1.04 TMP13 QGWSRDGPTLREWLAWRAVGHS 4.44 1.22 TMP14 QGAVPQGPTLKQWLLWRRCAHS 0.88 6.14 TMP15 QGRIREGPTLKEWLAQRRGFHS 1.03 5.24 TMP16 QGRFAEGPTLREWLEQRKLVHS 6.58 0.82 TMP17 QGDRFQGPTLREWLAAIRSVHS 12.90 0.42 TMP18 QGAGREGPTLREWLNMRVWQHS 12.80 0.42 TMP19 QGALQEGPTLRQWLGWGQWGHS 78.50 0.07 TMP20 QGYCDEGPTLKQWLVCLGLQHS 0.56 9.64 TMP21 QGWCKEGPTLREWLRWGFLCHS 1.53 3.53 TMP22 QGCSSGGPTLREWLQCRRMQHS <0.1 >54 TMP23 QGCSWGGPTLKQWLQCVRAKHS <0.1 > 54 Example 7 Peptide Bioactivity Study The biological activity of the peptide TMP1-TMP23 was determined using a cell-based assay. Murine 32D cell proliferation assays include murine 32D cells transfected with human mpl receptor. The following results are reported relative to TMP1. The CD61 cell assay involves the use of pro-human CD34+ cells cultured for several days in the presence of the peptide TMP1-TMP23. These cells were then divided into 114552.doc -83 * 1327149 to determine the percentage of cells that exhibited megakaryocyte-specific markers (CD61) on the cell surface. Markers of red blood cell precursor (CD36+) and neutrophil precursor (CD15+) were still at baseline when active compounds were used to stimulate the appearance of these platelet precursor cells in a dose-dependent manner. The qualitative results of the CD61 cell assay are shown below, which present an average of three different concentrations.

Peptide mouse 32D cell proliferation assay (relative to TMP1) CD61 cell assay (relative to TMP1) TMP01 100% -/+ TMP02 290% + TMP03 39% ++ TMP04 42% - TMP05 .85% ++ TMP06 569% ++ TMP07 289% ++ TMP08 39% + TMP09 2% - TMP10 12% - TMP11 21% - TMP12 10% - TMP13 328% ++ TMP14 635% +++ TMP15 35% - TMP16 32% + 114552.doc -84- TMP17 21% - TMP18 337% ++ TMP19 27% + TMP20 Undetectable / + TMP21 312% -/+ TMP22 Undetectable - TMP23 Cannot detect +++ Example 8 Peptide binding studies on BIAcore Binding assays were used to test the binding activity of several TMP peptides to hu-MPL. The experiment was carried out at 25 ° C using BIAcore 2000 (BIACORE Inc.). The running buffer was 0.005% surfactant plus P20. Recombinant protein G (Pierce 21193ZZ) was immobilized on the surface of a CM5 wafer using an amine coupling procedure (activated with NHS/EDC and blocked with ethanolamine) to capture the TMP peptide to about 400 RU. Recombinant hu-MPL (Lot 273 15-5 3) was serially diluted from 1 // Μ to 0.15 nM in sample buffer (?63 plus 0.005 °/. of surfactant? 20 and 100 μg/ml BSA) was then injected on the surface of the captured peptide body at 50 μl/min for 3 minutes. The rhu-MPL sample was also injected onto the blank protein G surface to subtract any non-specific binding background. The protein G surface was sequentially injected with 100 μl of ImmunoPure IgG Lysis Buffer (Pierce 21009ZZ, pH 2) and 100 μl of 8 mM Glycine, pH 1.5, 1 at 50 μL/min between two cycles. The vaporized copper of the earth is regenerated. The binding affinity (KD) of the peptide to rhu-MPL was determined by nonlinear regression analysis of the data using BIAevaluation 3 · 1 set 114552.doc -85 · 1327149 software (BIACORE Inc.). The results are summarized as follows: Polypeptide (TMP-Fc) ka(l/Ms) kd(l/s) Κ0(Μ) TMP20-Fc 5.06 x 1〇4 7.34 χ ΙΟ'3 1.45χ10-7 Fc-TMP24 4.01 χ 1〇4 8.75χ1〇-3 2.18 χ ΙΟ'7 TMP25-Fc 2.35 χ ΙΟ4 1.40 χ 1〇*3 5.97 χ ΙΟ'8 TMP26-Fc 2.58 χ ΙΟ4 5.72 χ ϊ〇^ 2.22 χ ΙΟ'7 Fc-TMP27 1.3 χ 1〇5 8.42 χ 1〇'3 6.49 χ ΙΟ·8 TMP28-Fc 6.78 χ ΙΟ4 2.52 χ 1〇'2 3.71 χ ΙΟ·7 Example 9 Peptide activity assay Putting human CD34 + cells in several TMp_Fc Cultured in the presence of the fusion protein for several days. These cells are then sorted to determine the percentage of cells that can exhibit megakaryocyte-specific D61 on the cell surface. Signs of red blood cell precursor (CD36+) (not shown) and neutrophil precursor (CD15+) (not shown) when using live & beta agents to stimulate the appearance of these platelet precursor cells in a dose-dependent manner The object is still at baseline. See Figure 1〇, “and 12 (CD61 Cell Assay). Example 10 In vivo Activity Using normal BDF1 mice, approximately 10-12 weeks old, in vivo activity studies” The bolus treatment was carried out. The subcutaneous injection was delivered in a volume of 0.23⁄4 liter. The compound was diluted in pBs plus I14552.doc -86 - 1327149. All experiments included a control marker "cutting". 'And ten mice were treated on day 0. The distance between the two groups was 4 and the number of mice was 20 mice per group. Five rats were placed at each time point, and at least three times of blood was discharged from the mother. The mice were anesthetized with fluorosis, and the total volume of blood was i4〇_i6〇 microliters by puncturing the eye. The blood was run on the echmcon H1E blood analyzer to run the mouse blood analysis software.

number. The parameters measured were white blood cells, red blood cells, hematocrit, hemoglobin, platelets, and neutrophils. See Figures 13 and 14. The present invention has been fully described herein, and it is understood that the invention may be modified, and the invention may be practiced without departing from the spirit and scope of the invention. BRIEF DESCRIPTION OF THE DRAWINGS Many other aspects and advantages of the present invention will become apparent from the following detailed description.

Figure 1 which has been treated with only this diluent shows an exemplary structure of the peptide and peptide-bonder compounds of the present invention. Figure 2 shows an exemplary structure of the peptide-vehicle and peptide-bonder-vehicle compound of the present invention. Figure 3 shows the nucleic acid and amino acid sequence of human lgQ1 Fc (Seq id NO: 3 1 and 32, respectively) which can be used as a preferred vehicle in the present invention. Figure 4 shows exemplary F c monomers and dimeric compounds of the invention, which may be derived from IgG1 antibodies. "fc" represents any Fc variant in the figure that is consistent with the meaning of the functional site herein. "Peptide" stands for any peptide, linker peptide, peptide-peptide combination, or any combination thereof, such as As disclosed herein. 114552.doc -87- 1327149 Specific dimers are as follows: Figures 4A and 4D show a single disulfide-bonded dimer. IgGl antibodies typically have two disulfides in the more stranded regions of the antibody. The Fc functional site in Figures 4A and 4D can be formed via truncation between two disulfide bond sites or via substitution of a cysteamine sulfhydryl residue with a non-reactive residue such as an alanine group. In Figure 4A, the Fc functional site is bonded to the amino terminus of the peptide; in Figure 4D, it is at the slow terminus of the peptide. Figures 4B and 4E show the dual disulfide-bonded dimer. By truncating the parent antibody to retain two cysteamine-branched residues in the Fc functional site chain, or by expression from a construct comprising sequences encoding such Fc functional sites. In Figure 4B, Fc function The site is bonded to the amino terminus of the peptide; in Figure 4E, at the carboxy terminus of the peptide. Figure 4C And 4F shows a non-covalent bond dimer. The Fc functional site can be formed by truncation or substitution to eliminate cysteamine residues. Suitably, the cysteamine thiol residue can be eliminated to avoid caspase An impurity formed by reacting an amine sulfhydryl residue with a cysteamine thiol residue contained in other proteins in the host cell. The non-covalent bond at the functional site is sufficient to hold the dimer together. The mer can be formed using Fc functional sites derived from different types of antibodies (eg, IgG2, igM). Figures 4G and 4H show binding to the N• terminus of the peptide (Fig. 4G) and the c_ terminus of the peptide (Fig. 4G) Figure 4H) Single-stranded Fc functional site. Figure 5 shows an exemplary structure of a preferred compound of the invention, characterized by a cascading repeat of a pharmaceutically active peptide that binds to an Fc functional site. Figure 5A shows a single strand (or The Fc monomer) molecule has a cascade of peptide dimerization 114552.doc -88 - 1327149 bound to it and also shows the DNA construct of the molecule. Figure 5B shows the ^ dimer 'where the linkage The peptide-peptide moiety is only present on one strand of the Fc dimer. Figure 5C shows on both strands An Fc dimer having a peptide moiety (in this case 'is a cascade peptide dimer.) The dimerization system of Figure 5C, in certain host cells, encodes a single strand of DNA as shown in Figure 5A. After the expression of the construct, it is naturally formed. In other host cells, the cells may be placed in a condition suitable for forming a dimer, or the dimer may be formed in vitro. Figures 5D to 51 represent other example sheets. Preferred specific examples of the strand (Fc monomer) and double strand (Fc dimer). Figure 6 shows the nucleotide sequence (SEQ ID 1 0 0 3 3) and amino acid of a preferred vector (20003 180). Sequence (5 £ (^10 1^0 34) for constituting butyl]\^-?; fusion compound, as shown in Example 3 herein. Figure 7 shows an exemplary oligomeric core: y: acid pair fragment for use in forming the preferred peptide of the invention, as shown in Example 3. The nuclei were separately presented: y: acid sequence and amino acid sequence (SEQ ID NO: 35-93). Figure 8 shows the nucleotide sequence (SEQ ID NO 94) and the amino acid sequence (SEq id NO 95) of the exemplary vector (20003 182) for constituting a C-terminal Fc fusion compound (ie, binding at the N-terminus) The peptide at the C-terminus of Fc) Figure 9 shows the ELISA dose-response of selected phage colonies. Figures 10, 11 and 12 show the biological activity of selected compounds of the invention. Figures 13 and 14 show the counts of platelets in vivo after a single injection of selected compounds of the invention in mice. U4552.doc *89· 1327149 Sequence Listing <11〇> American company Anmeiji <120>Peptide-generating peptide and related compound <I30> TW 091123464 <IS0> US 60/328,666 <151> 2001-10-11 <160> 199 <170> Patentln version 3.1

<210> 1 <2Ii> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Gin Gly lie Glu Gly Pro Thr Leu Arg Gin Trp Leu Ala Ala Arg Ala 15 10 15

Leu Glu

<210> 2 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400> 2

Gly Ala Glu Gly Pro Thr Leu Arg Gin Trp Leu Glu Trp Val Arg 1 5 10 15

Val Gly <210> 3 114552.doc 1327149 <211> 18 <212> PRT <213> artificial sequence <220><223> synthetic peptide sequence <400>

Arg Asp Leu Asp Gly Pro Thr Leu Arg Gin Trp Leu Pro Leu Pro Ser 15 10 15

Val Gin

<210> 4 <21i> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Leu Arg Asp Gly Pro Thr Leu Lys Gin Trp Leu Glu Tyr Arg Arg 15 10 15

Gin Ala

<210> 5 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Arg Gin Glu Gly Pro Thr Leu Lys Glu Trp Leu Phe Trp Val Arg 15 10 15

Met Gly <210> 6 2-114552.doc 1327149 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Glu Ala Leu Leu Gly Pro Thr Leu Arg Glu Trp Leu Ala Trp Arg Arg 15 10 15 <210> 7 <211> 18 <212> PRT <213>

Ala Gin <220><223> Synthetic peptide sequence <400> 7

Met Ala Arg Asp Gly Pro Thr Leu Arg Glu Trp Leu Arg Thr Tyr Arg 15 10 15

Met Met

<210> 8 <211> 18 <2i2> PRT <2i3> Artificial sequence <220><223> Synthetic peptide sequence <400>

Trp Met Pro Glu Gly Pro Thr Leu Lys Gin Trp Leu Phe His Gly Arg 15 10 15

Gly Gin <210> 9 114552.doc 1327149 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

His Πε Arg Glu Gly Pro Thr Leu Arg Gin Trp Leu Val Ala Leu Arg 15 10 15

Met Val

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Gin Leu Gly His Gly Pro Thr Leu Arg Gin Trp Leu Ser Trp Tyr Arg 15 10 15

Gly Met

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Glu Leu Arg Gin Gly Pro Thr Leu His Glu Trp Leu Gin His Leu Ala 15 10 15

Ser Lys <210> 12 4- H4552.doc 1327149 <21i> 18 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Val Gly He Glu Gly Pro Tar Leu Arg Gin Trp Leu Ala Gin Arg Leu 15 10 15

Asa Pro

<210> 13 <21i> 18 <212> PRT <2I3> Artificial sequence <220><223> Synthetic peptide sequence <400>

Trp Ser Arg Asp Gly Pro Thr Leu Arg Glu Trp Leu Ala Trp Arg Ala 15 10 15

Val Gly

<210> 14 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Val Pro Gin Gly Pro Thr Leu Lys Gin Trp Leu Leu Trp Arg Arg 15 10 15

Cys Ala <210> 15 114552.doc 1327149 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Arg lie Arg Glu Gly Pro Thr Leu Lys Glu Trp Leu Ala Gin Arg Arg 15 10 15

Gly Phe

<210> 16 <21l> 18 <2I2> PRT <213> Artificial sequence <220>'<223> Synthetic peptide sequence <400> 16

Arg Phe Ala Glu Gly Pro Thr Leu Arg Glu Trp Leu Glu Gin Arg Lys 15 10 15

Leu Val

<210> 17 <21I> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic victory J large sequence <400>

Asp Arg Phe Gin Gly Pro Thr Leu Arg Glu Trp Leu Ala Ala He Arg 15 10 15

Ser Val <210> 18 6-114552.doc 1327149 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic victory belly sequence <400>

Ala Gly Arg Glu Gly Pro Thr Leu Arg Glu Trp Leu Asn Met Arg Val 15 10 15

Trp Gin <210> 19 <211> 18 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Leu Gla Glu Gly Pro Thr Leu Arg Gin Trp Leu Gly Trp Gly Gin 15 10 15

Trp Gly

<210> 20 <211> 13 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp Leu Val Cys Leu Gly 15 10 15

Leu Gin <2i0> 21 114552.doc 1327149 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic surname peptide sequence <400> 21

Trp Cys Lys Glu Gly Pro Thr Leu Arg Glu Trp Leu Axg Trp Gly Phe 15 10 15

Leu Cys

<210> 22 <211> 18 <212> PRT ' <213> artificial sequence <220><223>synthetic peptide sequence <400>

Cys Ser Ser Gly Gly Pro Thr Leu Arg Glu Tip Leu Gin Cys Arg Arg 15 10 15

Met Gin

<210> 23 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Cys Ser Trp Giy Gly Pro Thr Leu Lys Gin Trp Leu Gin Cys Val Arg 15 10 15

Ala Lys <210> 24 114552.doc 1327149 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Cys Gin Leu Gly Gly Pro Thr Leu Arg Glu Trp Leu Ala Cys Arg Leu 15 10 15

Gly Ala

<210> 25 <211> 18 <212> PRT <213> artificial sequence <220><223> synthetic winning sequence <400>

Cys Trp Glu Gly Gly Pro Thr Leu Lys Glu Trp Leu Gin Cys Leu Val 15 10 15

Glu Arg

<210> 26 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Cys Arg Gly Gly Gly Pro Thr Leu His Gin Trp Leu Ser Cys Phe Arg 15 10 15

Trp Gin <210> 27 9-114552.doc 1327149 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Cys Arg Asp Gly Gly Pro Thr Leu Arg Gin Trp Leu Ala Cys Leu Gin 15 10 15

Gin Lys

<210> 28 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Glu Leu Arg Ser Gly Pro Thr Leu Lys Glu Trp Leu Val Trp Arg Leu 15 10 15

Ala Gin <210> 29 φ <21l> 18 <212> PRT <213> artificial sequence <220><223> synthetic victory belly sequence <400>

Gly Cys Arg Ser Gly Pro Thr Leu Arg Glu Trp Leu Ala Cys Arg Glu 15 10 15

Val Gin <210> 30 10-114552.doc 1327149 <211> 18 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400> 30

Thr Cys Glu Gin Gly Pro Thr Leu Arg Gin Trp Leu Leu Cys Arg Gin 15 10 15

Gly Arg

<210> 31 <211> 684 <212> DNA <213> Artificial sequence <220><223> Synthetic peptide sequence <220><221> CDS <222> (1). (684) <223><400> 31 atg gac aaa act cac aca tgt cca cct tgt cca get ccg gaa etc ctg 48 Met Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu i 5 10 15

ggS £ga ccg tea gtc ttc ccc ttc ccc cca aaa ccc aag gac acc etc 96 Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 20 25 30 atg ate tee egg acc cct gag gtc aca tgc gtg gtg gtg Gac gtg age 144 Met lie Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser 35 40 45 cac gaa gac cct gag gtc aag ttc aac tgg tac gtg gac ggc gtg gag 192 His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 50 55 60 gtg cat aat gee aag aca aag ccg egg gag gag cag tac aac age aeg 240 Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asn Ser Thr 65 70 75 80 tac cgt gtg gtc Age gtc etc acc gtc ctg cac cag gac tgg ctg aat 288 Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 85 90 95 -11 - 114552.doc 1327149 ggc aag gag tac aag tgc aag gtc tcc aac aaa Gcc etc cca gee ccc 336 Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 100 105 110 ate gag aaa acc ate tec aaa gcc aaa ggg cag ccc ega gaa cca cag 384 He Glu Lys Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 115 120 125 gtg tac acc ctg ccc cca tec egg gat gag ctg acc aag aac cag gtc 432 Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gin Val 130 135 140 age ctg acc tgc ctg gtc aaa ggc ttc Tat ccc age gac ate gcc gtg 480 Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp lie Ala Val 145 150 155 160

Gag tgg gag age aat ggg cag ccg gag aac aac tac aag acc aeg cct 528 Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro 165 170 175 ccc gtg ctg gac tec gac ggc tec ttc ttc etc tac. Aag etc acc 576 Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 180 185 190 gtg gac aag age agg tgg cag cag ggg aac gtc ttc tea tgc tec gtg 624 Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 195 200 205 atg cat gag get ctg cac aac cac tac aeg cag aag age etc tec ctg 672 Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 210 215 220 tet ccg ggt aaa 684

Ser Pro Gly Lys 225

<210> 32 <211> 228 <212> PRT <213> Artificial sequence .<220><223> Synthetic peptide sequence <400>

Met Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu 15 10 15

Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 20 25 30 •12· 114552.doc 1327149

Met lie Ser Arg Tnr Pro Glu Val Thr Cys Val Val Val Asp Val Ser 35 40 45

His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 50 55 60

Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asn Ser Thr 65 70 75 80

Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 85 90 95

Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 100 105 Π0

He Glu Lys Thr He Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 115 120 125

Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gin Val 130 135 140

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp lie Ala Val 145 150 155 160

Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro 165 170 175

Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 180 185 190

Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 195 200 205

Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 210 215 220

Ser Pro Gly Lys 225 -13 - 114552.doc 1327149 <210> 33 <211> 835 <212> DNA <213>Artificial sequence <220><223> Synthetic peptide sequence <220>;22i> CDS <222> (60)..(791) <223><400> 33 tagtcgatta atcgatttga ttctagattt gmtaacta attaaaggag gaataacat 59

Atg ggt gca cag aaa gcg gcc gca aaa aaa etc gag ggt gga ggc ggt 107 Met Gly Ala Gin Lys Ala Ala Ala Lys Lys Leu Glu Gly Gly Gly 15 10 15 ggg gac aaa act cac aca tgt cca cct tgc cca gca cct gaa Etc ctg 155 Gly Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu 20 25 30 ggg gga ccg tea gtt ttc etc ttc ccc cca aaa ccc aag gac acc etc 203 Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 35 40 45 atg ate tee egg acc cct gag gtc aca tgc gtg gtg gtg gac gtg age 251 Met lie Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser 50 55 60 cac gaa gac cct gag gtc Aag ttc aac tgg tac gtg gac ggc gtg gag 299 His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 65 70 75 80

Gtg cat aat gcc aag aca aag ccg egg gag gag cag tac aac age aeg 347 Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asn Ser Thr 85 90 95 tac cgt gtg gtc age gtc etc acc gtc ctg cac cag gac tgg Ctg aat 395 Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 100 105 110 ggc aag gag tac aag tgc aag gtc tee aac aaa gcc etc cca gcc ccc 443 Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 115 120 125 ate gag aaa acc ate tee aaa gcc aaa ggg cag ccc ega gaa cca cag 491 lie Glu Lys Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 130 135 140 gtg tac acc ctg ccc cca Tee egg gat gag ctg acc aag aac cag gtc 539 Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gin Val • 14- 114552.doc 1327149 145 150 155 160 age ctg acc tgc ctg gtc aaa ggc ttc tat ccc age Gac ate gee gtg 587 Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Scr Asp lie Ala Val 165 170 175 gag tgg gag age aat ggg cag ccg gag aac aac tac aag acc aeg cct 635 Glu Trp Glu Ser Asa Gly Gin Pro Glu Asn A Sn Tyr Lys Thr Thr Pro 180 185 190 ccc gtg ctg gac tee gac ggc tee ttc ttc etc tac age aag etc acc 683 Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 195 200 205 gtg gac aag age agg Tgg cag cag ggg aac gtc ttc tea tgc tee gtg 731 Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 210 215 220

At'g cat gag get ctg cac aac cac tac aeg cag aag age etc tee ctg 779 Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 225 230 235 240 tet ccg ggt aaa taatggatcc gcggaaagaa gaagaagaag aagaaagccc gaaa 835 Ser Pro Gly Lys <210> 34 <211> 244 <212> PRT < 213 > artificial sequence <220><223> synthetic peptide sequence <400>

Met Gly Ala Gin Lys Ala Ala Ala Lys Lys Leu Glu Gly Gly Gly Giy 15 10 15

Gly Asp Lys Thr His Thr Cys Pro ?vo Cys Pro Ala Pro Glu Leu Leu 20 30

Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Tnr Leu 35 40 45

Met lie Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser 50 55 60

His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu -15- 114552.doc 1327149 65 70 75 80

Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asn Ser Thr 85 90 95

Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 100 105 110

Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 115 120 125 lie Giu Lys Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 130 135 140

Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gin Val 145 150 155 160

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp He Ala Val 165 170 175

Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro 180 185 190

Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 195 200 205

Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 210 215 220

Met His Glu A.la Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 225 230 235 240

Ser Pro Gly Lys <210> 35 <211> 66 <212> DNA <213>Artificial Sequence<220><223> Synthetic peptide sequence-16-114552.doc 1327149 <220>221> CDS <222>(4)..(66) <223><400> 35 cat acg ggt gca cag ggt ate gaa ggt ccg act ctg cgt cag tgg ctg 48 Met Gly Ala Gin Gly lie Glu Gly Pro Thr Leu Arg Gin Tip Leu 15 10 15 get get cgt get etc gag 66

Ala Ala Arg Ala Leu Glu 20

<210> 36 <2il> 21 <212> PRT <213:> Artificial sequence <220><223> Synthetic peptide sequence <400> 36

Met Gly Ak Gin Gly lie Glu Gly Pro Thr Leu Arg Gin Trp Leu Ala 15 10 15

Aia Arg Ala Leu Glu 20

<210> 37 <211> 71 <212> PRT <213> Artificial shore column <220><223> Synthetic peptide sequence <400>

Tnr Gly Cys Ala Cys Ala Ala Gly Gly Thr Gly Gly Ala Gly Cys Ala 15 10 15

Cys Gly Thr Gly Ala Ala Gly Gly Ala Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Cys Gly Thr Cys Ala Ala Thr Gly Gly Cys Thr Thr Gly Ala 35 40 45 -17- 114552.doc 1327149

Ala Thr Gly Gly Gly Thr Thr Cys Gly Thr Gly Thr Thr Gly Gly Thr 50 55 60

Cys Ala Thr Thr Cys Thr Cys 65 70 <210> 38 <211> 71 <212> PRT <213> Artificial Sequence <220>

<223> Synthetic peptide sequence <4〇〇> 38

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Ala Cys Cys Ala Ala 15 10 15

Cys Ala Cys Gly Ala Ala Cys Cys Cys Ala Thr Thr Cys Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Gly Ala Cys Gly Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Thr Cys Cys Thr Thr Cys Ala Cys Gly Thr Gly Cys Thr Cys 50 55 60

Cys Aia Cys Cys Thr Thr Gly 65 70 <210> 39 <211> 77 <212> DNA <213>Artificial Sequence <220><223> Synthetic Peptide Sequence <220><221> CDS <222> (3)..(77) <223><400> 39 18-114552.doc 1327149 gt gca caa ggt gga gca cgt gaa gga cca act ctt cgt caa tgg ctt 47 Ala Gin Gly Gly Ala Arg Glu Gly Pro Thr Leu Arg Gin Trp Leu 15 10 15 gaa tgg gtt cgt gtt ggt cat tct etc gag 77

Glu Tr? Val Arg Val Gly His Ser Leu Glu 20 25 <210> 40 <211> 25 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400> 40

Ala Gin Gly Gly Ala Aig Glu Gly Pro Thr Leu Arg Gin Trp Leu Glu 15 10 15

Trp Val Arg Val Gly His Ser Leu Glu 20 25 <210> 41 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Cys Gly Thr Gly Ala Thr 15 10 15

Cys Thr Thr Gly Ala Thr Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Tar Cys Gly Thr Cys Ala Ala Thr Gly Gly Cys Thr Thr Cys Cys 35 40 45

Ala Cys Thr Thr Cys Cys Ala Tnr Cys Thr Gly Tnr Thr Cys Ala Ala 50 55 60

Cys Ala Thr Tnr Cys Thr Cys -19- 114552.doc 1327149 65 70 <210> 42 <211> 71 <212> PRT <213>Artificial Sequence<220><223> Synthetic Skin Sequence<223>;400> 42

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Thr Thr Gly Ala Ala 15 10 15

Cys Ala Gly Ala Thr Gly Gly Ala Ala Gly Thr Gly Gly Ala Ala Gly

Cys Cys Ala Thr Thr Gly Ala Cys Gly Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Ala Thr Cys Ala Ala Gly Ala Thr Cys Ala Cys 50 55 60

Gly Thr Cys Cys Thr Thr Gly 65 70

<210> 43 <211> 77 <212> DNA <213>Artificial sequence <220><223> Synthetic peptide sequence <220><221> CDS <222> (3). (77) <223><400> 43 gt gca caa gga cgt gat ctt gat ggt cca act ctt cgt caa tgg ctt 47 Ala Gin Gly Arg Asp Leu Asp Gly Pro Thr Leu Arg Gin Trp Leu 15 10 15 cca ctt Cca tct gn caa cat tct etc gag 77

Pro Leu Pro Ser Val Gin His Ser Leu Glu 20 25 -20- 114552.doc 1327149 <210> 44 <21l> 25 <212> PRT <213> Artificial Sequence <220><223> Peptide sequence <400> 44

Ala Gin Gly Arg Asp Leu Asp Gly Pro Thr Leu Arg Gin Trp Leu Pro 15 10 15

Leu Pro Ser Val Gin His Ser Leu Glu 20 25

<210> 45 <211> 71 <212> PRT <213> artificial sequence <220>c223> synthetic peptide sequence <400> 45

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Gly Cys Thr Thr Thr Ala 15 10 15

Cys Gly Thr Gly Ala Thr Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Ala Ala Ala Cys Ala Ala Thr Gly Gly Thr Thr Ala Gly Ala 35 40 45

Ala Thr Ala Thr Cys Gly Thr Cys Gly Thr Cys Ala Ala Gly Cys Thr 50 55 60

Cys Ala Thr Thr Cys Ala Cys 65 70 <210> 46 <211> 7i <2i2> PRT <213>Artificial Sequence-21 - 114552.doc 1327149 <220><223> Synthetic peptide sequence <223>;400> 46

Thr Cys Gly Ala Gly Thr Gly Ala Ala Thr Gly Ala Gly Cys Thr Thr 15 10 15

Gly Ala Cys Gly Ala Cys Gly Ala Thr Ala Thr Thr Cys Thr Ala Ala 20 25 30

Cys Cys Ala Thr Thr Gly Thr Thr Thr Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Ala Thr Cys Ala Cys Gly Thr Ala Ala Ala Gly

Cys Thr Cys Cys Thr Thr Gly 65 70 <210> 47 <211> 77 <212> 'DNA <213> Artificial Sequence <220><223> Synthetic Peptide Sequence <220><221>; CDS <222> (3)..(77)

<223><400> 47 gt gca caa gga get tta cgt gat ggt cca act ett aaa caa tgg tta 47 Ala Gin Gly Ala Leu Arg Asp Gly Pro Thr Leu Lys Gin Trp Leu 15 10 15 gaa tat cgt cgt caa get Cat tea etc gag 77

Glu Tyr Arg Arg Gin Ala His Ser Leu Glu 20 25 <210> 48 <211> 25 <212> PRT <213>Artificial Sequence-22-114552.doc 1327149 <220><223> Peptide sequence <400> 48

Ala Gin Gly Ala Leu Arg Asp Gly Pro Tnr Leu Lys Gin Trp Leu Glu 15 10 15

Tyr Arg Arg Gin Ala His Ser Leu Glu 20 25 <210> 49 <211> 71 <212> PRT <213> Artificial sequence

<220><223> Synthetic animal belly sequence <400> 49

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Gly Cys Ala Cys Gly Thr 15 10 15

Cys Ala Ala Gly Ala Ala Gly Gly Ala Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Ala Ala Ala Gly Ala Ala Thr Gly Gly Thr Thr Ala Thr Thr 35 40 45

Thr Thr Gly Gly Gly Thr Thr Cys Gly Thr Ala Thr Gly Gly Gly Thr

Cys Ala Thr Thr Cys Ala Cys 65 70 <210> 50 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Cys Gly Ala Giy Thr Gly Ala Ala Thr Gly Ala Cys Cys Cys Ala -23 - 114552.doc 1327149 15 10 15

Thr Ala Cys Gly Ala Ala Cys Cys Cys Ala Ala Ala Ala Thr Ala Ala 20 25 30

Cys Cys Ala Tnr Thr Cys Thr Thr Thr Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Thr Cys Cys Thr Thr Cys Thr Thr Gly Ala Cys Gly Thr Gly 50 55 60

Cys Thr Cys Cys Thr Thr Gly 65 70

<210> 51 <211> 77 <212> DNA <213> Artificial sequence <220><223> Synthetic victory belly sequence <220><221> CDS <222> (3). (77) <223><400> 51 gt gca caa gga gca cgt caa gaa gga cca act cu aaa gaa tgg tta 47 Ala GUi Gly Ala Arg Gin Glu Gly Pro Thr Leu Lys Glu Trp Leu 15 10 15

Ttt tgg gtt cgt atg ggt cat tea etc gag 77

"health sequence <220><223&gt

Ala Gin Gly Ala Arg Gin Glu Gly Pro Thr Leu Lys Glu Trp Leu Phe 1 5 10 15 -24- 114552.doc 1327149

Trp Val Arg Met Gly His Ser Leu Glu 20 25 <210> 53 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Gly Ala Ala Gly Cys Thr 15 10 15

Thr Thr Ala Thr Thr Ala Gly Gly Thr Cys Cys Ala Ala Cys Thr Thr 20 25 30

Thr Ala Cys Gly Thr Gly Ala Ala Thr Gly Gly Cys Thr Thr Gly Cys 35 40 45

Thr Thr Gly Gly Cys Gly Thr Cys Gly Thr Gly Cys Ala Cys Ala Ala 50 55 60

Cys Ala Thr Thr Cys Thr Cys 65 70

<210> 54 <2ll> 71 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <4〇0> 54

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Thr Thr Gly Thr Gly 15 10 15

Cys Ala Cys Gly Ala Cys Gly Cys Cys Ala Ala Gly Cys Ala Ala Gly 20 25 30-25- 114552.doc 1327149

Cys Cys Ala Thr Thr Cys Ala Cys Gly Thr Ala Ala Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Thr Ala Ala Thr Ala Ala Ala Gly Cys Thr Thr 50 55 60

Cys Thr Cys Cys Thr Thr Gly 65 70 <210> 55 <211> 77 <212> DNA <213> Artificial sequence

<220><223> Synthetic peptide sequence <220><221> CDS <222> (3).. (77) <223><400> 55 gt gca caa gga gaa get tta tta Ggt cca act tta cgt gaa tgg ett 47 Ala Gin Gly Glu Ala Leu Leu Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15 get tgg cgt cgt gca caa cat tet etc gag 77

Ala Trp Arg Arg Ala Gin His Ser Leu Glu 20 25

<210> 56 <211> 25 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400> 56

Ala Gin Gly Glu Ala Leu Leu Gly Pro Thr Leu Arg Glu Trp Leu Ala 15 10 15

Trp Arg Arg Ala Gin His Ser Leu Glu 20 25 <210> 57 -26- 114552.doc 1327149 <2Π> 71 <212> PRT <213> Artificial Sequence <220><223> Synthetic peptide Sequence <400> 57

Thr Gly Cys Ala Cys Ala Ala Gly Gly Thr Ala Ίΐτ Gly Gly Cys Ala 15 10 15

Cys Gly Thr Gly Ala Thr Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Cys Gly Thr Gly Ala Ala Thr Gly Gly Cys Thr Thr Cys Gly

Thr AJa Cys Thr Thr Ala Thr Cys Gly Thr Ala Thr Gly Ala Thr Gly 50 55 60

Cys Ala Thr Thr Cys Thr Cys 65 70 <210> 58 <211> 71 <212> PRT <213> Artificial Sequence <220>

<223> Synthetic peptide sequence <400> 58

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Cys Ala Thr Cys Ala 15 10 15

Thr Ala Cys Gly Ala Thr Ala Ala Gly Thr Ala Cys Gly Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Cys Ala Cys Gly Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Ala Thr Cys Ala Cys Gly Thr Gly Cys Cys Ala 50 55 60 -27- 114552.doc 1327149

Thr Ala Cys Cys Thr Thr Gly 65 70 <210> 59 <21l> 77 <212> DNA <213>Artificial Sequence <220><223> Synthetic Peptide Sequence <220><221> CDS <222>(3)..(77) <223>

<400> 59 gt gca caa ggt atg gca cgt gat ggt cca act ctt cgt gaa tgg ctt 47 Ala Gla Gly Met Ala Arg Asp Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15 cgt act tat cgt atg atg cat tct etc gag 77

Arg Thr Tyr Arg Met Met His Ser Leu Glu 20 25 <210> 60 <211> 25 <212> PRT <213> Artificial Sequence <220>

<223> Synthetic peptide sequence <400> 60

Ala Gin Gly Met Ala Arg Asp Gly Pro Thr Leu Arg Glu Trp Leu Arg 15 10 15

Thr Tyr Arg Met Met His Ser Leu Glu 20 25 <210> 61 <211> 71 <212> PRT <213>Artificial Sequence<220><223> Synthetic peptide sequence-28-114552.doc 1327149 <400> 61

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Thr Gly Gly Ala Thr Gly 15 10 15

Cys Cys Ala Gly Ala Ala Gly Gly Ala Cys Cys Ala Ala Cys Ala Thr 20 25 30

Thr Ala Ala Ala Ala Cys Ala Ala Thr Gly Gly Cys Thr Thr Thr Thr 35 40 45

Thr Cys Ala Thr Gly Gly Thr Cys Gly Thr Gly Gly Thr Cys Ala Ala 50 55 60

Cys Ala Thr Thr Cys Thr Cys 65 70 <210> 62 <2U> 71 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Thr Thr Gly Ala Cys 15 10 15

Cys Ala Cys Gly Ala Cys Cys Ala Thr Gly Ala Ala Ala Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Gly Thr Thr Thr Thr Ala Ala Thr Gly Thr Thr 35 40 45

Gly Gly Thr Cys Cys Thr Thr Cys Thr Giy Gly Cys Ala Thr Cys Cys 50 55 60 • Ala Thr Cys Cys Thr Thr Gly 65 70 <210> 63 <211> 77 -29- 114552.doc 1327149 <212> DNA <213> artificial sequence <220><223> synthetic peptide sequence <220><221> CDS <222> (3).. (77) <223><400> 63 gt Gca caa gga tgg atg cca gaa gga cca aca tta aaa caa tgg ctt 47 Ala Gb Gly Trp Met Pro Glu Gly Pro Thr Leu Lys Gin Trp Leu 15 10 15 ttt cat ggt cgt ggt caa cat tct etc gag 77

"health sequence" <220> artificial sequence &lt

Ala Gin Gly Trp Met Pro Glu Gly Pro Thr Leu Lys Gin Tip Leu Phe 15 10 15

<212> 71 <212&gt

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Cys Ala Thr Ala Thr Thr 15 10 15

Cys Gly Thr Gly Ala Ala Gly Gly Thr Cys Cys Ala Ala Cys Ala Thr -30- 114552.doc 1327149 20 25 30

Thr Ala Cys Gly Thr Cys Ala Ala Thr Gly Gly Cys Thr Thr Gly Thr 35 40 45

Thr Gly Cys Thr Cys Thr Thr Cys Gly Thr Ala Thr Gly Gly Thr Thr 50 55 60

Cys Ala Thr Thr Cys Thr Cys 65 70

<2i0> 66 <211> 71 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Ala Ala Cys Cys Ala 15 10 15

Thr Ala Cys Gly Ala Ala Gly Ala Gly Cys Ala Ala Cys Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Gly Ala Cys Gly Thr Ala Ala Thr Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Thr Thr Cys Ala Cys Gly Ala Ala Thr Ala Thr 50 55 60

Gly Thr Cys Cys Thr Thr Gly 65 70 <210> 67 <211> 77 <212> DNA <213>Artificial sequence <220><223> Synthetic peptide sequence <220> •31 · 114552 .doc 1327149 20 25 30

Thr Ala Cys Gly Thr Cys Ala Ala Thr Gly Gly Cys Thr Thr Gly Thr 35 40 45

Thr Gly Cys Γητ Cys Thr Thr Cys Gly Thr Ala Thr Gly Gly Thr Thr 50 55 60

Cys Ala Thr Thr Cys Thr Cys 65 70

<210> 66 <211> 71 <212> PRT < 213 > artificial sequence <220><223> synthetic peptide sequence • <400>

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Ala Ala Cys Cys Ala 15 10 15

Thr Ala Cys Gly Ala Ala Gly Ala Gly Cys Ala Ala Cys Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Gly Ala Cys Gly Thr Ala Ala Thr Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Thr Thr Cys Ala Cys Gly Ala Ala Thr Ala Thr 50 55 60

Gly Thr Cys Cys Thr Thr Gly 65 70 <210> 67 <211> 77 <212> DNA <213>Artificial Sequence <220><223> Synthetic peptide sequence <220> -32·114552 .doc 1327149

Thr Thr Gly Gly Thr Ala Thr Cys Gly Thr Gly Gly Thr Ala Thr Gly 50 55 60

70, <210>&lt

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Cys Ala Thr Ala Cys 15 10 15

Cys Ala Cys Gly Ala Thr Ala Cys Cys Ala Ala Gly Ala Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Gly Ala Cys Gly Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Ala Thr Gly Thr Cys Cys Thr Ala Ala Thr Thr 50 55 60

Gly Ala Cys Cys Thr Thr Gly 65 70

<210> 71 <211> 77 <212> DNA <213> artificial sequence <220><223> synthetic peptide sequence <220><221> CDS <222> (3). (77) <223><400> 71 gt gca caa ggt caa tta gga cat ggt cca act ctt cgt caa tgg ctt 47 Ala Gin Gly Gin Leu Gly His Gly Pro Thr Leu Arg Gin Trp Leu • 33- 114552. Doc 11327149 15 10 15 tct tgg tat cgt ggt atg cat tct etc gag 77

Ser Trp Tyr Arg Gly Met His Ser Leu Glu 20 25 <210> 72 <211> 25 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Gin Gly Gin Leu Gly His Gly Pro Thr Leu Arg Gin Trp Leu Ser 1 5 10 15

Trp Tyr Arg Gly Met His Ser Leu Glu 20 25 <210> 73 <21l> 71 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Gly Ala Ala Thr Thr Ala 15 10 15

Cys Gly Thr Cys Ala Ala Gly Gly Ala Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Cys Ala Thr Gly Ala Ala Thr Gly Gly Cys Thr Thr Cys Ala 35 40 45

Ala Cys Ala Thr Thr Thr Ala Gly Cys Ala Ala Gly Cys Ala Ala Ala 50 55 60

Cys Ala Thr Tnr Cys Thr Cys 65 70 -34- 114552.doc 1327149 <210> 74 <211> 71 <212> PRT <213:► artificial sequence <220><223> synthetic peptide sequence <400> 74

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Thr Thr Tnr Gly Cys 15 10 15

Thr Thr Gly Cys Thr Ala Ala Ala Thr Gly Thr Thr Gly Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Cys Ala Thr Gly Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Thr Cys Cys Thr Tnr Gly Ala Cys Gly Thr Ala Ala Thr Thr 50 55 60

Cys Thr Cys Cys Thr Thr Gly 65 70 <210> 75 <211> 77 <212> DNA <213> Artificial sequence

<220><223> Synthetic peptide sequence <220><221> CDS <222> (3).. (77) <223><400> 75 gt gca caa gga gaa tta cgt caa Gga cca act ctt cat gaa tgg ctt 47 Ala Gin Gly Glu Leu Arg Gin Gly Pro Thr Leu His Glu Trp Leu 15 10 caa cat tta gca age aaa cat tet etc gag 77

Gin His Leu Ala Ser Lys His Ser Leu Glu 20 25 -15- 114552.doc 1327149 <210> 76 <211> 25 <212> PRT <213> Artificial Sequence <220><223> Synthetic Limb Belly sequence <400> 76

Ala Gin Gly Glu Leu Arg Gin Gly Pro Thr Leu His Glu Trp Leu Gin 15 10 15

His Leu Ala Ser Lys His Ser Leu Glu 20 25 φ <210> 77 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400> 77

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Gly Thr Ala Gly Gly Thr 15 10 15

Ala Thr Thr Gly Ala Ala Gly Gly Thr Cys Cys Ala Ala Cys Ala Thr 20 25 30

Thr Ala Cys Gly Thr Cys Ala Ala Thr Gly Gly Thr Thr Ala Gly Cys 35 40 45

Thr Cys Ala Ala Cys Gly Thr Cys Thr Thr Ala Ala Thr Cys Cys Ala 50 55 60

Cys Ala Thr Thr Cys Thr Cys 65 70 <210> 78 <211> 71 <212> PRT <213>Artificial Sequence<220> -36-114552.doc 1327149 <223> Synthetic peptide sequence<;400> 78

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Thr Gly Gly Ala Thr 1 5 10 15

Thr Ala Ala Gly Ala Cys Gly Thr Thr Gly Ala Gly Cys Thr Ala Ala 20 25 30

Cys Cys Ala Thr Thr Gly Ala Cys Gly Thr Ala Ala Thr Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Thr Thr Cys Ala Ala Thr Ala Cys Cys Thr Ala 50 55 60

Cys Thr Cys Cys Thr Thr Gly 65 70 <210> 79 <211> 77 <212> DNA <213>Artificial Sequence <220><223> Synthetic Peptide Sequence <220><221> CDS <222> 0)..(77)

<223><400> 79 gt gca caa gga gta ggt att gaa ggt cca aca tta cgt caa tgg tta 47 Ala Gin Gly Val Gly He Glu Gly Pro Thr Leu Arg Gin Trp Leu 15 10 15 get caa cgt ett aat cca Cat tet etc gag 77

Ala Gin Arg Leu Asn Pro His Ser Leu Glu 20 25 <210> 80 <211> 25 <212> PRT <213>Artificial Sequence<220><223> Synthetic peptide sequence 37-114552.doc 1327149 <400> 80

Ala Gin Gly Val Gly lie Glu Gly Pro Thr Leu Arg Gin Trp Leu Ala 15 10 15

Gin Arg Leu Asn Pro His Ser Leu Glu 20 25 <210> 81 <21I> 71 <212> PRT <213> Artificial Sequence <220>

<223> Synthetic peptide sequence <400> 81

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Thr Gly Gly Thr Cys Ala 15 10 15

Cys Gly Thr Gly Ala Thr Gly Gly Thr Cys Cys Ala Ala Cys Ala Cys 20 25 30

Thr Thr Cys Gly Thr Gly Ala Ala Thr Gly Gly Cys Thr Thr Gly Cys 35 40 45

Thr Thr Gly Gly Cys Gly Thr Gly Cys Thr Gly Thr Thr Gly Gly Ala 50 55 60

Cys Ala Thr Ala Gly Thr Cys 65 70 <210> 82 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic surname peptide sequence <400>

Thr Cys Gly Ala Gly Ala Cys Thr Ala Thr Gly Thr Cys Cys Ala Ala 15 10 15 114552.doc 1327149

Cys Ala Gly Cys Ala Cys Gly Cys Cys Ala Ala Gly Cys Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Cys Ala Cys Gly Ala Ala Gly Thr Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Ala Thr Cys Ala Cys Gly Thr Gly Ala Cys Cys 50 55 60

Ala Thr Cys Cys Thr Tnr Gly 65 70

<210> 83 <2U> 77 <212> DNA <213> Artificial Sequence <220><223> Synthetic Victory At Sequence<220><22i> CDS <222> (3). .07) <223><40O> 83 gt gca caa gga tgg tea cgt gat ggt cca aca ett cgt gaa tgg ett 47 Ala Gin Gly Trp Ser Arg Asp Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15

Get tgg cgt get gtt gga cat agt etc gag 77

Ala Trp Arg Ala Val Gly His Ser Leu Glu 20 25 <210> 84 <211> 25 <212>PRT_<213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Gin Gly Trp Ser Arg Asp Gly Pro Thr Leu Arg Glu Trp Leu Ala 15 10 15 -39- 114552.doc 1327149

Trp Axg Ala Val Gly His Ser Leu Glu 20 25 <210> 85 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Gly Cys Ala Gly Thr Thr 1 5 10 15

Cys Cys Ala Cys Ala Ala Gly Gly Ala Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Ala Ala Ala Cys Ala Gly Thr Gly Gly Thr Thr Ala Thr Thr 35 40 45

Ala Thr Gly Gly Cys Gly Thr Cys Gly Thr Thr Gly Thr Gly Cys Ala 50 55 60

Cys Ala Thr Thr Cys Thr Cys 65 70

<210> 86 <211> 71 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Thr Gly Cys Ala Cys 15 10 15

Ala Ala Cys Gly Ala Cys Gly Cys Cys Ala Thr Ala Ala Thr Ala Ala 20 25 30

Cys Cys Ala Cys Thr Gly Γητ Thr Thr Ala Ala Gly Ala Gly Thr Thr 35 40 45 -4υ · 114552.doc 1327149

Gly Gly Thr Cys Cys Thr Thr Gly Thr Gly Gly Ala Ala Cys Thr Gly 50 55 60

Cys Thr Cys Cys Thr Thr Gly 65 70 <210> 87 <211> 77 <212> DNA <213>Artificial Sequence <220><223> Synthetic Name Sequence <220><221> CDS <222> (3)..(77) <223><400> 87 gt gca caa gga gca gtt cca caa gga cca act ctt aaa cag tgg tta 47 Ala Gin Gly Ala Val Pro Gin Gly Pro Thr Leu Lys Gin Ttp Leu 15 10 15 tta tgg cgt cgt tgt gca cat tct etc gag 77

Leu Trp Arg Arg Cys Ala His Ser Leu Glu 20 25

<210> 88 <211> 25 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Gin Gly Ala Val Pro Gin Gly Pro Thr Leu Lys Gin T卬 Leu Leu l 5 10 15

Trp Arg Arg Cys Ala His Ser Leu Glu 20 25

<210> 89 <211> 71 <212> PRT -41 - 114552.doc 1327149 <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Thr Cys Gly Thr Ala Thr Thr 15 10 15

Cys Gly Thr Gly Ala Ala Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Ala Ala Ala Gly Ala Ala Thr Gly Gly Cys Thr Thr Gly Cys 35 40 45

Thr Cys Ala Ala Cys Gly Thr Cys Gly Thr Gly Gly Thr Thr Thr Thr 50 55 60

Cys Ala Thr Ala Gly Thr Cys 65 70 <210> 90 <211> 71 <212> PRT <213>Artificial Sequence <220><223> Synthetic Winning Sequence <400>

Thr Cys Gly Ala Gly Ala Cys Thr Ala Thr Gly Ala Ala Ala Ala Cys 15 10 15

Cys Ala Cys Gly Ala Cys Gly Thr Thr Gly Ala Gly Cys Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Cys Thr Thr Thr Ala Ala Gly Aia Giy Thr Thr 35 40 45

Gly Gly Ala Cys Cys Thr Thr Cys Ala Cys Gly Ala Ala Thr Ala Cys 50 55 60 .

Gly Ala Cys Cys Thr Thr Gly -42- 114552.doc 1327149 65 70 <210> 91 <211> 77 <212> DNA <213>Artificial Sequence<220><223> Synthetic belly sequence<223>;220><221> CDS <222>(3)..(77) <223><400> 91

Gt gca caa ggt cgt att cgt gaa ggt cca act ctt aaa gaa tgg ctt 47 Ala Gin Gly Arg lie Axg Glu Gly Pro Thr Leu Lys Glu Trp Leu 15 10 15 get caa cgt cgt ggt ttt cat agt etc gag 77

Ala Gin Arg Arg Gly Phe His Ser Leu Glu 20 25 <210> 92 <211> 25 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Gin Gly Arg lie Arg Glu Giy Pro Thr Leu Lys Glu Trp Leu Ala 15 10 15

Gin Arg Arg Gly Phe His Ser Leu Glu 20 25 <210> 93 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400> 93 43 - 114552.doc 1327149

Thr Giy Cys Ala Cys Ala Ala Gly Gly Thr Cys Gly T'nr Thr Thr Cys 15 10 15

Gly Cys Thr Gly Ala Ala Gly Gly Thr Cys Cys Ala Ala Cys Ala Cys 20 25 30

Thr Thr Cys Gly Thr Giy Ala Ala Thr Gly Gly Thr Thr Ala Gly Ala 35 40 45

Ala Cys Ala Ala Cys Gly Thr Ala Ala Ala Cys Thr Thr Gly Thr Thr 50 55 60

Cys Ala Thr Ala Gly Thr Cys 65 70

<210> 94 <211> 71 <212> PRT <213> artificial sequence <220><223> synthetic surname sequence <400>

Thr Cys Gly Ala Gly Ala Cys Thr Ala Thr Gly Ala Ala Cys Ala Ala 15 10 15

Gly Thr Thr Thr Ala Cys Gly Thr Thr Gly Thr Thr Cys Thr Ala Ala 20 25 30

Cys Cys Ala Thr Thr Cys Ala Cys Gly Ala Ala Gly Thr Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Thr Thr Cys Ala Gly Cys Gly Ala Ala Ala Cys 50 55 60

Gly Ala Cys Cys Thr Thr Gly 65 70 <210> 95 <211> 77 <212> DNA <213>Artificial Sequence -44 - 114552.doc 1327149 <220><223> Synthetic peptide sequence <223>;220><221> CDS <222> (3)..(77) <223><400> 95 gt gca caa ggt cgt ttc get gaa ggt cca aca ett cgt gaa tgg tta 47 Ala Gin Gly Arg Phe Ala Glu Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15 gaa caa cgt aaa ett gtt cat agt etc gag 77

Glu Gin Arg Lys Leu Val His Ser Leu Glu 20 25

<210> 96 <211> 25 <212> PRT <213> Artificial sequence <220><223> Synthetic skin sequence <400>

Ala Gin Gly Arg Phe Ala Glu Gly Pro Thr Leu Arg Glu Trp Leu Glu 15 10 15

Gla Arg Lys Leu Val His Ser Leu Glu 20 25

<210> 97 <211> 71 <212> PRT <213> artificial sequence <220><223> synthetic surname sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Thr Gly Ala Thr Cys Gly Thr 15 10 15

Thr Thr Cys Cys Ala Ala Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30 -45- 114552.doc 1327149 <220><223> Synthetic peptide sequence <220><221> CDS <222> (3)..(77) <223><4〇〇> 95 gt gca caa ggt cgt ttc get gaa ggt cca aca ett cgt gaa tgg tta 47 Ala Gin Gly Arg Phe Ala Glu Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15 gaa caa cgt aaa ett gtt cat agt etc gag 77

Glu Gin Arg Lys Leu Val His Ser Leu Glu 20 25

<210> 96 <211> 25 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Gin Gly Arg Phe Ala Glu Gly Pro Thr Leu Arg Glu Trp Leu Glu 15 10 15

Gin Arg Lys Leu Val His Ser Leu Glu 20 25

<210> 97 <2ll> 71 <212> PRT <213> Artificial shore column <220><223> Synthetic victory belly sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Thr Gly Ala Thr Cys Gly Thr 15 10 15

Thr Thr Cys Cys Ala Ala Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30 -46- 114552.doc <223>1327149 <400> 99 gt gca caa ggt gat cgt tic caa ggt cca act ctt cgt gaa tgg Ctt 47 Ala Gin Gly Asp Arg Phe Gin Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15 get gca ate cgt age gta cat agt etc gag 77

Ala Ala lie Arg Ser Val His Ser Leu Glu 20 25 <210> 100 <211> 25 <212> PRT <2I3> artificial sequence <220>

<223> Synthetic peptide sequence <4〇0> 100

Ala Gin Gly Asp Arg Phe Gin Gly Pro Thr Leu Arg Glu Trp Leu Ala 15 10 15

Ala He Arg Ser Val His Ser Leu Glu 20 25 <210> 101 <211> 71 <212> PRT <213> Artificial Sequence <220>

<223> Synthetic surname peptide sequence <4〇〇> 101

Thr Gly Cys Ala Cys Ala Ala Gly Gly Thr Gly Cys Thr Gly Gly Thr 15 10 15

Cys Gly Thr Gly Ala Ala Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr' Ala Cys Gly Thr Gly Ala Ala Thr Gly Gly Cys Thr Tnr Ala Ala 35 40 45

Thr Ala Thr Gly Cys Gly Thr Gly Thr Thr Thr Gly Gly Cys Ala Ala 50 55 60 -47- 114552.doc 1327149

Cys Ala Thr Thr Cys Thr Cys 65 70 <210> 102 <2ll> 71 <212> PRT <213> Artificial Sequence <220><223> Synthetic Name Peptide Sequence <400>

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Thr Thr Gly Cys Cys 15 10 15

Ala Ala Ala Cys Ala Cys Gly Cys Ala Thr Ala Thr Thr Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Cys Ala Cys Gly Thr Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Thr Thr Cys Ala Cys Gly Ala Cys Cys Ala Gly 50 55 60

Cys Ala Cys Cys Thr Thr Gly 65 70

<210> 103 <211> 77 <212> DNA <213>Artificial sequence <220><223> Synthetic peptide sequence <220><221> CDS <222> (3). (77) <223><400> 103 gt gca caa ggt get ggt cgt gaa ggt cca act eta cgt gaa tgg ett 47 Ala Gin Gly Ala Gly Arg Glu Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15 -48 - 114552.doc 1327149 aat atg cgt gtt tgg caa cat cct etc gag 77

Asn Met Arg Val Trp Gin His Ser Leu Glu 20 25 <210> 104 <211> 25 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Gin Gly Ala Gly Arg Glu Gly Pro Thr Leu Arg Glu Trp Leu Asn 15 10 15

Met Arg Val Trp Gin His Ser Leu Glu 20 25 <210> 105 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Gly Cys Thr Thr Thr Ala 15 10 15

Cys Ala Ala Gly Ala Ala Gly Gly Ala Cys Cys Ala Ala Cys Ala Thr 20 25 30

Thr Ala Cys Gly Thr Cys Ala Ala Thr Gly Gly Thr Thr Ala Gly Gly 35 40 45

Ala Thr Gly Gly Gly Gly Thr Cys Ala Ala Thr Gly Gly Gly Gly Ala 50 55 60

Cys Ala Cys Thr Cys Thr Cys 65 70 <210> 106 49-114552.doc 1327149 <211> 71 <212> PRT <213>Artificial Sequence<220><223><223> Synthetic peptide sequence<400> 106

Thr Cys Gly Ala Gly Ala Gly Ala Gly Thr Gly Thr Cys Cys Cys Cys 15 10 15

Ala Thr Thr Gly Ala Cys Cys Cys Cys Ala Thr Cys Cys Thr Ala Ala 20 25 30

Cys Cys Ala Thr Thr Gly Ala Cys Gly Thr Ala Ala Thr Gly Thr Thr

Gly Gly Tnr Cys Cys Thr Thr Cys Thr Thr Gly Thr Ala Ala Ala Gly 50 55 60

Cys Thr Cys Cys Thr Thr Gly 65 70 <210> 107 <211> 77 <212> DNA <213> Artificial Sequence <220>

<223> Synthetic peptide sequence <220><221> CDS <222> (3)..(77) <223><400> 107 gt gca caa gga get tta caa gaa gga cca aca tta Cgt caa egg tta 47 Ala Gin Gly Ala Leii Gin Glu Gly Pro Thr Leu Arg Gin Trp Leu 15 10 15 gga tgg gst caa tgg gga cac tet etc gag 77

Gly Trp Gly Gin Trp Gly His Ser Leu Glu 20 25 <210> 108 <211> 25 50-114552.doc 1327149 <212> PRT <213> Artificial Sequence <220><223> Synthetic peptide Sequence <400> 108

Ala Gin Gly Ala Leu Gin Glu Gly Pro Thr Leu Arg Gin Trp Leu Gly 15 10 15

Trp Gly Gin Trp Gly His Ser Leu Glu 20 25

<210> 109 <2U> 71 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <4〇〇> 109

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Thr Ala Cys Thr Gly Thr 15 10 15

Gly Ala Thr Gly Ala Ala Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Ala Ala Ala Cys Ala Ala Thr Gly Gly Thr Thr Ala Gly Thr 35 40 45

Ala Thr Gly Thr Cys Thr Thr Gly Gly Thr Thr Thr Ala Cys Ala Ala 50 55 60

Cys Ala Thr Ala Gly Tiir Cys 65 70 <210> 110 <211> 71 <212> PRT <213>Artificial Sequence<220><223> Synthetic peptide sequence -51 - 114552.doc 1327149 <;40〇> 110

Thr Cys Gly Ala Gly Ala Cys Thr Ala Thr Gly Thr Thr Gly Thr Ala 15 10 15

Ala Ala Cys Cys Ala Ala Gly Ala Cys Ala Thr Ala Cys Thr Ala Ala 20 25 30

Cys Cys Ala Thr Thr Gly Thr Thr Thr Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Thr Thr Cys Ala Thr Cys Ala Cys Ala Gly Thr 50 55 60

Ala Thr Cys Cys Thr Thr Gly 65 70 <210> 111 <211> 77 <212> DNA <213>Artificial sequence <220><223> Synthetic peptide sequence <220><221> CDS <222>(3)..(77) <223><400> 111

Gt gca caa gga tac tgt gat gaa ggt cca act ctt aaa caa tgg tla 47 Ala Gin Gly Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp Leu 15 10 15 gta tgt ctt ggt tta caa cat agt etc gag 77

Val Cys Leu Gly Leu Gin His Ser Leu Glu 20 25 <210> 112 <211> 25 <212> PRT <213>Artificial Sequence<220><223> Synthetic Peptide Sequence <400> 52-114552.doc 1327149

Ala Gin Gly Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp Leu Val 15 10 15

Cys Leu Gly Leu Gin His Ser Leu Glu 20 25 <210> 113 <2ll> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Thr Gly Thr Ala Gly Thr 15 10 15

Thr Cys Ala Gly Gly Ala Gly Gly Thr Cys Cys Ala Ala Cys Thr Thr 20 25 30

Thr Ala Cys Gly Thr Gly Ala Ala Thr Gly Gly Thr Thr Ala Cys Ala 35 40 45

Ala Thr Gly Thr Cys Gly Thr Cys Gly Thr Ala Thr Gly Cys Ala Ala 50 55 60

Cys Ala Thr Thr Cys Thr Cys

<210> 114 <211> 7i <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Thr Thr Gly Cys Ala 15 10 15

Thr Ala Cys Gly Ala Cys Gly Ala Cys Ala Thr Thr Gly Thr Ala Ala -53- ;14552.doc 1327149 20 25 30

Cys Cys Ala Thr Thr Cys Ala Cys Gly Thr Ala Ala Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Thr Cys Cys Thr Gly Ala Ala Cys Thr Ala Cys 50 55 60

Ala Thr Cys Cys Thr Tnr Gly 65 70

<210> 115 <211> 77 <212> DNA <2I3> artificial sequence <220><223> synthetic peptide sequence <220><221> CDS <222> (3). (77) <223><400> 115 gt gca caa gga tgt agt tea gga ggt cca act tta cgt gaa tgg tta 47 Ala Gin Gly Cys Ser Ser Gly Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15 caa tgt Cgt cgt atg caa cat tet etc gag 77

Gin Cys Arg Arg Met Gin His Ser Leu Glu 20 25

<210> 116 <211> 25 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Gin Gly Cys Ser Ser Gly Gly Pro Thr Leu Arg Glu Trp Leu Gin 15 10 15

Cys Arg Arg Met Gin His Ser Leu Glu 20 25 -54- 114552.doc 1327149 <210> 117 <211> 71 <212>PRT <213>Artificial sequence <220><223> Sequence <400> 117

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Thr Gly Thr Thr Cys Ala 15 10 15

Thr Gly Gly Gly Gly Thr Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Ala Ala Ala Cys Ala Ala Thr Gly Gly Thr Thr Ala Cys Ala 35 40 45

Ala Thr Gly Thr Gly Thr Thr Cys Gly Thr Gly Cys Thr Ala Ala Ala 50 55 60

Cys Ala Thr Thr Cys Thr Cys 65 70 <210> 118 <211> 71 <212> PRT <213>

<223> Synthetic peptide sequence <400> 118

Thr Cys Gly Ala Gly Ala Gly Ala Ala Thr Gly Thr Thr Thr Ala Gly 15 10 15

Cys Ala Cys Gly Ala Ala Cys Ala Cys Ala Thr Thr Gly Thr Ala Ala 20 25 30

Cys Cys Ala Thr Thr Gly Thr Thr Thr Ala Ala Gly Ala Gly Thr Thr 35 40 45 114552.doc 1327149

Gly Gly Ala Cys Cys Ala Cys Cys Cys Cys Ala Thr Gly Ala Ala Cys 50 55 60

Ala Thr Cys Cys Thr Thr Gly 65 70 <210> 119 <211> 77 <212> DNA <213>Artificial sequence <220><223> Synthetic peptide sequence <220>

<221> CDS <222> (3)..(77) <223><400> 119 gt gca caa gga tgt tea tgg ggt ggt cca act ett aaa caa tgg tta 47 Ala Gin Gly Cys Ser Trp Gly Gly Pro Thr Leu Lys Gin Trp Leu 15 10 15 caa tgt gtt cgt get aaa cat tet etc gag 77

Gin Cys Val Arg Ala Lys His Ser Leu Glu 20 25 <210> 120 <2ll> 25 <212> PRT <213> Artificial Sequence Lu <220><223> Synthetic peptide sequence <400> 120

Ala Gin Gly Cys Ser Trp Gly Gly Pro Thr Leu Lys Gin Trp Leu Gin 15 10 15

Cys Val Arg Ala Lys His Ser Leu Glu 20 25 <210> 121 <21l> 71 <212> PRT <213>Artificial Sequence -56-114552.doc 1327149 <220><223> Synthetic peptide Sequence <400> 121

Thr Gly Cys .Ma Cys Ala Ala Gly Gly Ala Thr Gly Thr Cys Ala Ala 15 10 15

Tnr Thr Ala Gly Gly Thr Gly Gly Tnr Cys Cys Gly Ala Cys Thr Cys 20 25 30

Thr Thr Cys Gly Thr Gly Ala Ala Thr Gly Gly Cys Thr Thr Gly Cys 35 40 45

Thr Thr Gly Thr Cys Gly Thr Cys Thr Thr Gly Gly Thr Gly Cys Thr 50 55 60

Cys Ala Thr Thr Cys Ala Cys 65 70 <210> 122 <211> 71 <212> PRT <213> Artificial Sequence <220><223> Synthetic Skin Sequence <400>

Thr Cys Gly Ala Gly Thr Gly Ala Ala Thr Gly Ala Gly Cys Ala Cys 15 10 15

Cys Ala Ala Gly Ala Cys Gly Ala Cys Ala Ala Gly Cys Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Cys Ala Cys Gly Ala Ala Gly Ala Gly Thr Cys 35 40 45

Gly Gly Ala Cys Cys Ala Cys Cys Thr Ala Ala Thr Tnr Gly Ala Cys 50 55 60

Ala Thr Cys Cys Thr Tnr Gly 65 70 -57- 114552.doc 1327149 <210> 123 <211> 77 <212> DNA <213>Artificial Sequence<220><223> Synthetic peptide sequence<223>;220><221> CDS <222>(3)..(77) <223><400> 123 gt gca caa gga tgt caa tta ggt ggt ccg act cn cgt gaa tgg ctt 47 Ala Gin Gly Cys Gin Leu Gly Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15

Get tgt cgt etc ggt get cat tea etc gag 77

Ala Cys Arg Leu Gly Ala His Ser Leu Glu 20 25 <210> 124 <211> 25 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Gin Gly Cys Gin Leu Gly Gly Pro Thr Leu Arg Glu Trp Leu Ala 15 10 15

Cys Arg Leu Gly Ala His Ser Leu Glu 20 25 <210> 125 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400> 125

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Thr Gly Thr Thr Gly Gly 15 10 15 • 58 - 114552.doc 1327149

Gly Ala Ala Gly Giy Tlu Gly Giy Thr Cys Cys Thr Ala Cys Ala Cys 20 25 30

Thr Tnr Ala Ala Ala Gly Ala Ala Thr Gly Gly Cys Thr Thr Cys Ala 35 40 45

Ala Thr Gly Thr Cys Thr Thr Gly Thr Ala Gly Ala Ala Cys Gly Thr 50 55 60

Cys Ala Thr Thr Cys Ala Cys 65 70 <210> 126 <211> 71 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Cys Gly Ala Gly Thr Gly Ala Ala Thr Gly Ala Cys Gly Thr Thr 15 10 15

Cys Thr Ala Cys Ala Ala Gly Ala Cys Ala Thr Thr Gly Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Cys Thr Thr Thr Ala Ala Gly Thr Gly Thr Ala

Gly Gly Ala Cys Cys Ala Cys Cys Thr Thr Cys Cys Cys Ala Ala Cys 50 55 60

Ala Thr Cys Cys Thr Thr Gly 65 70 <210> 127 <211> 77 <212> DNA <213>Artificial Sequence<220> -59*114552.doc 1327149 <223> Synthetic peptide sequence<;220><221> CDS <222> (3)..(77) <223><400> 127 gt gca caa gga tgt tgg gaa ggt ggt cct aca ctt aaa gaa tgg ctt 47 Ala Gin Gly Cys Tip Glu Gly Gly Pro Thr Leu Lys Glu Trp Leu 15 10 15 caa tgt ctt gta gaa cgc cat tea etc gag 77

Gin His Ser Leu Glu 20 25 <210&gt

Ala Gin Gly Cys Trp Glu Gly Gly Pro Thr Leu Lys Glu Trp Leu Gin 1 5 10 15

Cys Leu Val Glu Arg His Ser Leu Glu 20 25

<210> 129 <211> 71 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Thr Thr Gly Thr Cys Gly Thr 15 10 15

Gly Gly Thr Gly Gly Tnr Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Cys Ala Thr Cys Ala Ala Thr Gly Gly Cys Thr Thr Thr Cys -60- 114552.doc 1327149 35 40 45

Thr Thr Gly Thr Thr Thr Thr Cys Gly Thr Thr Gly Gly Cys Ala Ala 50 55 60

Cys Ala Thr Thr Cys Ala Cys 65 70 <210> 130 <2U> 71 <212> PRT <213> Artificial Sequence <220>

<223> Synthetic peptide sequence <400> 130

Thr Cys Gly Ala Gly Thr Gly Ala Ala Thr Gly Thr Thr Gly Cys Cys 15 10 15

Ala Ala Cys Gly Ala Ala Ala Ala Cys Ala Ala Gly Ala Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Gly Ala Thr Gly Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Ala Cys Cys Ala Cys Cys Ala Cys Gly Ala Cys 50 55 60

Ala Ala Cys Cys Thr Thr Gly 65 70 <210> 131 <211> 77 <212> DNA <213>Artificial Sequence <220><223> Synthetic Glycopeptide Sequence <220><221> CDS <222> (3)..(77) <223> -61 - 114552.doc 1327149 <400> 131 gt gca caa ggt tgt cgt ggt ggt ggt cca act ctt cat caa tgg ctt 47 Ala Gin Gly Cys Arg Gly Gly Gly Pro Thr Leu His Gin Trp Leu 15 10 15 tct tgt ctt cgt tgg caa cat tea etc gag 77

Ser Cys Phe Arg Trp Gin His Ser Leu Glu 20 25 <210> 132 <211> 25 <212> PRT <213> Artificial Sequence <220>

<223> Synthetic surname peptide sequence <400> 132

Ala Gin Gly Cys Arg Gly Gly Gly Pro Thr Leu His Gin Trp Leu Ser 15 10 15

Cys Phe Arg Trp Gin His Ser Leu Glu 20 25 <210> 133 <211> 71 <212> PRT <213>Artificial sequence <220>

<223> Synthetic peptide sequence <4〇〇> 133

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Thr Gly Thr Cys Gly Thr 15 10 15

Gly Ala Thr Gly Gly Thr Gly Gly Thr Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Thr Ala Gly Ala Cys Ala Ala Thr Gly Gly Cys Thr Thr Gly Cys 35 40 45

Thr Thr Gly Thr Cys Tnr Thr Cys Ala Ala Cys Ala Ala Ala Ala Ala 50 55 60 -62- U4552.doc 1327149

Cys Ala Thr Thr Cys Ala Cys 65 70 <210> 134 <211> 71 '<212>PRT <213>Artificial Sequence<220><223>Synthesis Peptide Sequence <4〇〇> 134

Thr Cys Gly Ala Gly Thr Gly Ala Ala Thr Gly Thr Thr Thr Thr Thr 15 10 15

Gly Thr Thr Gly Ala Ala Gly Ala Cys Ala Ala Gly Cys Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Gly Thr Cys Thr Ala Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Ala Cys Cys Ala Thr Cys Ala Cys Gly Ala Cys 50 55 60

Ala Thr Cys Cys Thr Thr Gly 65 70

<210> 135 <211> 77 <212> DNA <213>Artificial sequence <220><223> Synthetic surname peptide sequence <220><22i> CDS <222> (3). (77) <223><400> 135 gt gca caa gga tgt cgt gat ggt ggt cca act ctt aga caa tgg ctt 47 Ala Gla Gly Cys Arg Asp Gly Gly Pro Thr Leu Arg Gin Trp Leu 15 10 15 get tgt Ett caa caa aaa cat tea etc gag 77

Ala Cys Leu Gin Gin Lys His Ser Leu Gla • 63 114552.doc 1327149 20 25 <210> 136 <211> 25 <212> PRT <213>Artificial Sequence <220><223> Synthetic Name Peptide Sequence <400> 136

Ala Gin Gly Cys Arg Asp Gly Gly Pro Thr Leu Arg Gin Trp Leu Ala 15 10 15

Cys Leu Gin Gin Lys His Ser Leu GIu 20 25

<210> 137 <211> 71 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Cys Gly Ala Gly Thr Gly Ala Ala Thr Giy Thr Thr Gly Ala Gly 15 10 15

Cys Ala Ala Gly Ala Cys Gly Cys Cys Ala Ala Ala Cys Ala Ala Gly 20 25 30

Cys Cys Ala Thr Thr Cys Thr Thr Thr Thr Ala Ala Ala Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Ala Gly Ala Thr Cys Thr Thr Ala Ala Thr Thr 50 55 60

Cys Thr Cys Cys Thr Thr Gly 65 70

<210> 138 <2U> 71 <212> PRT 64-114552.doc 1327149 <213> Artificial sequence <220><223> Synthetic skin sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Gly Ala Ala Thr Thr Ala 15 10 15

Ala Gly Ala Thr Cys Thr Gly Gly Thr Cys Cys Ala Ala Cys Thr Thr 20 25 30

Thr Ala Ala Ala Ala Gly Ala Ala Thr Gly Gly Cys Thr Thr Gly Thr 35 40 45

Thr Thr Gly Gly Cys Gly Thr Cys Thr Thr Gly Cys Thr Cys Ala Ala 50 55 60

Cys Ala Thr Thr Cys Ala Cys 65 70 <210> 139 <211> 77 <212> DNA <213>Artificial Sequence <220><223> Synthetic Surname Sequence <220>

<22l> CDS <W> 〇)..W) <223><400> 139 gt gca caa gga gaa tta aga tct ggt cca act tta aaa gaa tgg ctt 47 Ala Gin Gly Glu Leu Arg Ser Gly Pro Thr Leu Lys Glu Trp Leu 15 10 15 gtt tgg cgt ctt get caa cat tea etc gag 77

Val Trp Arg Leu Ala Gin His Ser Leu Glu 20 25 <210> 140 <211> 25 <212> PRT <213>Artificial sequence 65-114552.doc 1327149 <220><223> Sequence <400> 140

Ala Gin Gly Glu Leu Arg Ser Gly Pro Thr Leu Lys GIu Trp Leu Val 15 10 15

Trp Arg Leu Ala Gin His Ser Leu GIu 20 25 <210> 141 <211> 71 <212> PRT <213> Artificial sequence Lu <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Ala Gly Gly Ala Thr Gly Thr 15 10 15

Ala Gly Ala Thr Cys Thr Gly Gly Thr Cys Cys Ala Ala Cys Ala Cys 20 25 30

Thr Thr Cys Gly Thr Gly Ala Ala Thr Gly Gly Thr Thr Ala Gly Cys 35 40 45

Thr Thr Gly Thr Ala Gly Ala Gly Ala Gly Gly Thr Thr Cys Ala Ala 50 55 60

Cys Ala Cys Thr Cys Thr Cys 65 70 <210> 142 <211> 71 <212> PRT <213>Artificial Sequence<220><223> Synthetic Victory Sequence<400> 142 66- 114552.doc 1327149

Thr Cys Gly Ala Gly Ala Gly Ala Gly Thr Gly Thr Thr Gly Ala Ala 15 10 15

Cys Cys Thr Cys Thr Cys Thr Ala Cys Ala Ala Gly Cys Thr Ala Ala 20 25 30

Cys Cys Ala Thr Thr Cys Ala Cys Gly Ala Ala Gly Thr Gly Thr Thr 35 40 45

Gly Gly Ala Cys Cys Ala Gly Ala Thr Cys Tnr Ala Cys Ala Thr Cys 50 55 60

Cys Thr Cys Cys Thr Thr Gly 65 70

<210> 143 <211> 77 <212> DNA <213>Artificial sequence <220><223> Synthetic peptide sequence <220><221> CDS <222> (3). (77) <223><400> 143 gt gca caa gga gga tgt aga tct ggt cca aca ctt cgt gaa tgg tta 47 Ala Gin Gly Gly Cys Arg Ser Gly Pro Thr Leu Arg Glu Trp Leu

Get tgt aga gag gtt caa cac tci etc gag 77

Ala Cys Arg Glu Val Gin His Ser Leu Glu 20 25 <2i0> 144 <211> 25 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Gin Gly Gly Cys Arg Ser Gly Pro Thr Leu Arg Glu Trp Leu Ala -67- 114552.doc 1327149 15 10 15

Cys Arg Glu Val Gin His Ser Leu Glu 20 25 <210> 145 <211> 71 <212> PRT <213:. Artificial sequence <220><223> Synthetic peptide sequence <400>

Thr Gly Cys Ala Cys Ala Ala Gly Gly Thr Ala Cys Ala Thr Gly Cys 15 10 15

Gly Ala Ala Cys Ala Ala Gly Gly Ala Cys Cys Ala Ala Cys Thr Cys 20 25 30

Thr Ala. Ala Gly Ala Cys Ala Ala Thr Gly Gly Cys Thr Ala Cys Thr 35 40 45

Ala Thr Gly Thr Ala Gly Ala Cys Ala Ala Gly Gly Ala Ala Gly Ala 50 55 60

Cys Ala Cys Thr Cys Ala Cys 65 70

<210> 146 <211> 71 <212> PRT <213r artificial sequence <220><223> synthetic peptide sequence <400>

Thr Cys Gly Ala Gly Thr Gly Ala Gly Thr Gly Thr Cys Thr Thr Cys 15 10 15

Cys Thr Thr Gly Thr Cys Tnr Ala Cys Ala Thr Ala Gly Thr Ala Gly 20 25 30 • 68 - 114552.doc 1327149

Cys Cys Ala Thr Thr Gly Thr Cys Thr Thr Ala Gly Ala Gly Thr Thr 35 40 45

Gly Gly Thr Cys Cys Thr Γητ Gly Thr Thr Cys Gly Cys Ala Thr Gly 50 55 60

Thr Ala Cys Cys Thr Thr Gly 65 70 <210> 147 <211> 77 <212> DNA <213> Artificial Sequence

<220><223> Synthetic peptide sequence <220><221> CDS <222> (3).. (77) <223><400> 147 gt gca caa ggt aca tgc gaa caa Gga cca act eta aga caa tgg eta 47 Ala Gin Gly Thr Cys Glu Gin Gly Pro Thr Leu Arg Gin Trp Leu 15 10 15 eta tgt aga caa gga aga cac tea etc gag 77

Leu Cys Arg Gin Gly Arg His Ser Leu Glu 20 25

<210> 148 <211> 25 <212> PRT <213> artificial sequence <220><223> synthetic belly sequence <40C>

Ala Gin Gly Thr Cys Glu Gin Gly Pro Tnr Leu Arg Gin Trp Leu Leu 15 10 15

Cys Arg Gin Gly Arg His Ser Leu Glu 20 25 -69- 114552.doc 1327149 <210> 149 <211> 77 <212> DNA <213> Artificial Sequence <220><223> Synthetic peptide Sequence <220><221> CDS <222> (3)..(77) <223><400> 149 gt gca cag ggt tgg tgt aag gag ggt cct act ctg cgt gag tgg ctg 47 Ala Gin Gly Trp Cys Lys Glu Gly Pro Thr Leu Arg Glu Trp Leu 15 10 15

Egg tgg ggt ttt ctg tgt cat tet etc gag 77

Arg Trp Gly Phe Leu Cys His Ser Leu Glu 20 25 <210> 150 <211> 25 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <4〇〇 > 150

Ala Gin Gly Trp Cys Lys Glu Gly Pro Thr Leu Arg Glu Trp Leu Arg 15 10 15

Trp Gly Phe Leu Cys His Ser Leu Glu 20 25 <210> 151 <211> 837 <212> DNA <213>Artificial Sequence <220><223> Synthetic Victory Sequence <220>< 22 CDS <222> (57)..(785) <223> 70-114552.doc 1327149 <400> 151 tcgattaatc gatttgattc tagatttgtt ttaactaatt aaaggaggaa caacat atg 59

Met 1 gac aaa act cac aca tgt cca cct tgt cca fict ccg gaa etc ctg ggg 107 Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly 5 10 15 gga ccg tea gtc ttc etc etc ccc cca aaa ccc aag Gac acc etc atg 155 Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met 20 25 30 ate tee egg acc cct gag gtc aca tgc gtg gtg gtg gac gtg age cac 203 lie Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His 35 40 45

Gaa gac cct gag gtc aag ttc aac tgg tac gtg gac ggc gtg gag gtg 251 Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val 50 55 60 65 cat aat gee aag aca ag ccg egg gag gag cag tac aac Age aeg tac 299 His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asn Ser Thr Tyr 70 75 80 cgt gtg gtc age gtc etc acc gtc ctg cac cag gac tgg ctg aat ggc 347 Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn Gly 85 90 95 aag gag tac aag tgc aag gtc tee aac aaa gee etc cca gee ccc ate 395 Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro He 100 105 110 gag aaa acc ate tee Aaa gee aaa ggg cag ccc ega gaa cca cag gtg 443 Glu Lys Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin Val 115 120 125

Tac acc ctg ccc cca tee egg gat gag ctg acc aag aac cag gtc age 491 Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gin Val Ser 130 135 ' 140 145 ctg acc tgc ctg gtc aaa ggc ttc tat ccc age gac Ate gee gtg gag 539 Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp lie Ala Val Glu 150 155 160 tgg gag age aat ggg cag ccg gag aac aac tac aag acc aeg cct ccc 587 Trp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro 165 170 175 gtg ctg gac lee gac ggc tee ttc ttc etc tac age aag etc acc gtg 635 Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val 180 185 190 gac aag age agg Tgg cag cag ggg aac gtc ttc tea tgc tee gtg atg 683 -71 - 114552.doc 1327149

Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val Met 195 200 205 cat gag get ctg cac aac cac tac aeg cag aag age etc tee ctg tet 731 His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu Ser 210 215 220 225 ccg ggt aaa ggt gga ggt ggt ggt gca cag aaa geg gee gca aaa aaa 779 Pro Gly Lys Gly Gly Gly Gly Gly Ala Gin Lys Ala Ala Ala Lys Lys 230 235 240 ccc gag taatggatcc gcggaaagaa gaagaagaag aagaaagccc gaaaggaagc tg 83y Leu Glu

<210> 152 <211> 243 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Met Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu 15 10 15

Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 20 25 30

Met lie Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser

His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 50 55 60

Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asa Ser Thr 65 70 75 80

Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 85 90 95

Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 100 105 110 •72· I14552.doc 1327149 lie Glu Lys Thr He Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 115 120 125

Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gin Val 130 135 140

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp lie Ala Val 145 150 155 160

Glu Trp Giu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro 165 170 175

Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 180 185 190

Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 195 200 205

Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 210 215 220

Ser Pro Gly Lys Gly Gly Gly Gly Gly Ala Gin Lys Ala Ala Ala Lys 225 230 235 240

Lys Leu Glu

<210> 153 <2ll> 45 <212> PRT < 213 > artificial sequence <220><223> synthetic winning sequence <220><221> misc_feature <222> (22) ..(22) <223> At Position 22, PEG linker <400> 153

His lie Arg Glu Gly Pro Thr Leu Arg Gin Trp Leu Val Ala Leu Arg 15 10 15 -73- 114552.doc 1327149

Met Val Gly Gly Gly Pro Glu Gly Gly Gly Gly His lie Arg Glu Gly 20 25 30

Pro Thr Leu Arg Gin Trp Leu Val Ala Leu Arg Met Val 35 40 45 <210> 154 <211> 43 <212> PRT <2I3> Artificial Sequence <220>

<223> Synthetic peptide sequence <220><221> misc.feature <222> (1)..(1) <223> At Position 1, Fc at N-terminus <220><221> misc_feature <222> (43)..(43) <223> At position 43, Fc on C-terminus <400> 154

Thr Cys Glu Gin Gly Pro Thr Leu Axg Gin Trp Leu Leu Cys Arg Gin 15 10 15

Gly Arg Gly Gly Gly Lys Gly Gly Gly Thr Cys Glu Gin Gly Pro Thr 20 25 30

Leu Arg Gin Trp Leu Leu Cys Arg Gin Gly Arg 35 40 <210> 155 <211> 40 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <220><221> misc_feature • 74· 114552.doc 1327149 <222> (1)..(1) <223> At position I, Fc at N-tenriinus <400>

Gin Leu Gly His Gly Pro Thr Leu Arg Gin Trp Leu Ser Trp Tyr Arg 15 10 15

Gly Met Gly Pro Asn Gly Glu Leu Axg Ser Gly Pro Thr Leu Lys Glu 20 25 30

Trp Leu Val Trp Arg Leu Ala Gin 35 40

<210> 156 <211> 18 <212> PRT < 213 > artificial sequence <220><223> synthetic winning sequence <220><221> misc_feature <222>(19). (19) <223> At position 19, Fc at C-terminus <400> 156

Cys Ser Trp Gly Gly Pro Thr Leu Lys Gin Trp Leu Gin Cys Val Arg 15 10 15

Ala Lys <210> 157 <211> 25 <212> PRT < 213 > artificial sequence <220><223> synthetic winning sequence <220><221> misc_fcaturc <222> )..(1) <223> At position 1, Fc at N-terminus •75· 114552.doc 1327149 <400> 157

Giy Gly Gly Lys Gly Gly Gly Ala Val Pro Gin Gly Pro Thr Leu Lys 15 10 15

Gin Trp Leu Leu Trp Arg Arg Cys Ala 20 25 <210> 158 <211> 18 <212> PRT <213>Artificial Sequence<220><223> Synthetic Animal Sequence Lu <220><;221> misc^feature <222> (1)..(1) <223> At position l, PEG group attached <400> 158

Cys Ser Ser Gly Gly Pro Thr Leu Axg Glu Trp Leu Gin Cys Arg Arg 15 10 15

Met Gin

<2i0> 159 <211> 46 <212; · PRT < 213 > artificial sequence <220><223> synthetic peptide sequence <220><221> misc.feature <222> 1)..(1) <223> At position 1, Fc at N-ierminus <4〇〇> 159

Gly Gly Gly Gly Gly Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp 15 10 15 •76· 114552.doc 1327149

Leu Val Cys Leu Gly Leu Gin Gly Gly Gly Gly Gly Tyr Cys Asp Glu 20 25 30

Gly Pro Thr Leu Lys Gin Trp Leu Val Cys Leu Gly Leu Gin 35 40 45 <210> 160 <211> 75 <212> PRT <213>Artificial Sequence<220><223> Synthetic peptide sequence <220>

<221> misc_feature <222>(76)..(76) <223> At position 76, Fc at C-terrainus <400> 160

Cys Ser Trp Gly Gly Pro Thr Leu Lys Gin Trp Leu Gin Cys Val Arg 15 10 15

Ala Lys Gly Gly Gly Ala Gly Gly Gly Cys Ser Τφ Gly Gly Pro Thr 20 25 30

Leu Lys Gin Trp Leu Gin Cys Val Arg Ala Lys Gly Gly Gly Ala Gly 35 40 45

Gly Gly Cys Ser Trp Gly Gly Pro Tnr Leu Lys Gin Trp Leu Gin Cys 50 55 60

Val Arg Ala Lys Gly Gly Gly Ala Gly Gly Gly 65 70 75 <210> 161 <21l> 43 <212> PRT <213>Artificial Sequence <220><223> Synthetic peptide sequence • 77· 114552.doc 1327149 <220><221> misc_feature <222> (44)..(44) <223> At position 44, PEG group at C-terminas <4〇0> 161

Val G!y lie Glu Gly Pro Tnr Leu Arg Gin Trp Leu Ala Gin Arg Leu 15 10 15

Asn Pro Gly Gly Gly Cys Gly Gly Gly Val Gly lie Glu Gly Pro Thr 20 25 30

Leu Arg Gin Trp Leu Ala Gin Arg Leu Asn Pro 35 40

<210> 162 <211> 40 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <220><221> rnisc_fcature <222> (1). (1) <223> At position 1, Fc at N-terminus <400> 162

Glu Leu Arg Ser Gly Pro Thr Leu Lys Glu Trp Leu Val Trp Arg Leu 15 10 15

Ala Gin Gly Gly Gly Gly Glu Leu Arg Ser Gly Pro Thr Leu Lys Glu 20 25 30

Trp Leu Val Trp Arg Leu Ala Gin 35 40 <210> 163 <211> 43 <212> PRT <213>Artificial Sequence<220> 78-114552.doc 1327149 <223> Synthetic peptide sequence<;220><221> misc_feature <222> (1)..(1) <223> Ac position 1, Fc at N-terminus <220><221> misc_feature <222> (44). .(44) <223> At position 44, Fc at C-terminus <4〇0> 163

Ala Leu Arg Asp Gly Pro Thr Leu Lys Gin Trp Leu Glu Tyr Axg Arg 15 10 15

Gin Ala Gly Gly Gly Lys Gly Gly Gly Ala Leu Arg Asp Gly Pro Thr 20 25 30

Leu Lys Gin Trp Leu Glu Tyr Arg Arg Gin Ala 35 40 <210> 164 <211> 36 <212> PRT <213>Artificial Sequence <220>

<223> Synthetic peptide sequence <400> 164

Ala Leu Arg Asp Gly Pro Thr Leu Lys Gin Trp Leu Glu Tyr Arg Arg 15 10 15

Gin Ala Ala Leu Arg Asp Gly Pro Thr Leu Lys Gin Trp Leu Glu Tyr 20 25 30

Arg Arg Gin Ala 35 <210> 165 <211> 36 <212> PRT <213> Artificial sequence -79-114552.doc 1327149 <220><223> Synthetic winning sequence <4〇〇 > 165

Glu Ala Lea Leu Gly Pro Thr Leu Arg Glu Trp Leu Ala Trp Arg Arg 15 10 15

Ala Gin Glu Ala Leu Leu Gly Pro Thr Leu Arg Glu Trp Leu Ala Trp 20 25 30 .Arg Arg Ala Gin 35

<210> 166 <211> 36 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Ala Val Pro Gin Gly Pro Thr Leu Lys Gin Trp Leu Leu Trp Arg Arg 15 10 15

Cys Ala Ala Val Pro Gin Gly Pro Thr Leu Lys Gin Trp Leu Leu Trp 20 25 30

Arg Arg Cys Ala 35 <210> 167 <211> 36 <212> PRT <213> artificial sequence <220><223> synthetic trait sequence <400>

Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp Leu Val Cys Leu Gly 15 10 15 -80 - 114552.doc 1327149

Leu Gin Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp Leu Val Cys 20 25 30

Leu Gly Leu Gin 35 <210> 168 <211> 36 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <400>

Cys Ser Ser Gly Gly Pro Thr Leu Arg Glu Trp Leu Gin Cys Arg Arg 15 10 15

Met Gin Cys Ser Ser Gly Gly Pro Thr Leu Arg Glu Trp Leu Gin Cys 20 25 30

Arg Arg Met Gin 35 <210> 169 <2U> 36 <212> PRT <213> Artificial sequence

<220><223> Synthetic peptide sequence <400> 169

Cys Ser Trp Gly Gly Pro Thr Leu Lys Gin Trp Leu Gin Cys Val Arg 15 10 15

Ala Lys Cys Ser Trp Gly Gly Pro Tnr Leu Lys Gin Trp Leu Gin Cys 20 25 30

Val Arg Ala Lys 35 <210> 170 <211> 41 -81 - 114552.doc 1327149 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400 170

Ala Leu Arg Asp Gly Pro Thr Leu Lys Gin Trp Leu Glu Tyr Arg Arg 15 10 15

Gin Ala Gly Gly Gly Gly Gly Ala Leu Arg Asp Gly Pro Thr Leu Lys 20 25 30

Gin Trp Leu Glu Tyr Arg Arg Gin Ala 35 40

<210> 171 <211> 41 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Glu Ala Leu Leu Gly Pro Thr Leu Arg Glu T卬 Leu Ala Trp Arg Arg 15 10 15

Ala Gin Gly Gly Gly Gly Gly Glu Ala Leu Leu Gly Pro Thr Leu Arg 20 25 30

Lu Arg Ala Gla 35 40 <210>

Ala Val Pro Gin Gly Pro Thr Leu Lys Gin Trp Leu Leu Trp Arg Arg 15 10 15 -82 * 114552.doc 1327149

Cys Ala Gly Gly Gly Gly Gly Ala Val Pro Gin Gly Pro Thr Leu Lys 20 25 30

Gin Trp Leu Leu Trp Arg Arg Cys Ala 35 40

<210> 173 <211> 41 <212> PRT <2ϋ> Artificial sequence <220><223> Synthetic victory belly sequence Lu <400>

Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp Leu Val Cys Leu Gly 15 10 15

Leu Gin Gly Gly Gly Gly Gly Tyr Cys Asp Glu Gly Pro Thr Leu Lys 20 25 30

Gin Trp Leu Val Cys Leu Gly Leu Gin 35 40

<2i0> 174 <211> 41 <212> PRT <213> Artificial sequence <220><223> Synthetic peptide sequence <400>

Cys Ser Ser Gly Gly Pro Thr Leu Arg Glu Trp Leu Gin Cys Arg Arg 15 10 15

Met Gin Gly Gly Gly Gly Gly Cys Ser Ser Giy Gly Pro Thr Leu Arg 20 25 30

Glu Trp Leu Gin Cys Arg Arg Met Gin 35 40 -83- 114552.doc 1327149 <210> 175 <211> 41 <212> PRT <2I3> Artificial Sequence <220><223> Synthetic peptide Slice column <400> 175

Cys Ser Trp Gly Gly Pro Tnr Leu Lys Gb Trp Leu Gin Cys Val Arg 15 10 15

Ala Lys Gly Gly Gly Gly Gly Cys Ser Trp Gly Gly Pro Thr Leu Lys 20 25 30

Gin Trp Leu Gin Cys Val Arg Ala Lys 35 40 <210> 176 <211> 23 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <220>221> misc.feature <222> (1)..(1) <223> At position 1, Fc at N-terminus

<4〇〇> 176

Gly Gly Gly Gly Gly Ala Leu Arg Asp Gly Pro Thr Leu Lys Gin Trp 15 10 15

Leu Glu Tyr Arg Arg Gin Ala 20 <210> 177 <211> 23 <212> PRT · <213>Artificial sequence <220><223> Synthetic peptide sequence -84 - 114552.doc 1327149 <;220><221> misc^feature <222> (1)..(1) <223> At position 1, Fc at N-terminus <400> 177

Gly Gly Gly Gly Gly GIu Ala Leu Leu Gly Pro Thr Leu Arg Glu Trp 15 10 15

Leu Ala Trp Arg Arg Ala Gin 20

<210> 178 <211> 23 <212> PRT < 213 > artificial sequence <220><223> synthetic peptide sequence <220><221> misc.feature <222> )..(1) <223> At position 1, Fc at N-terminus <400> 178

Gly Gly Gly Gly Gly Ala Val Pro Gin Gly Pro Thr Leu Lys Gin Trp 15 10 15

Leu Leu Trp Arg Arg Cys Ala 20 • <210> 179 <211> 23 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <220><22l> Misc.feature <222> (1)..(1) <223> At position 1, Fc at N-terminus -85- 114552.doc 1327149 <400> 179

Gly Gly Gly Gly Gly Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp 15 10 15

Leu Val Cys Leu Gly Leu Gin 20 <210> 180 <211> 23 <212> PRT <213>Artificial Sequence <220>

<223> Synthetic skin sequence <220><221> misc_feacure <222> (1)..(1) <223> At position 1, Fc at N-terrninus <400>

Gly Gly Gly Gly Gly Cys Ser Ser Gly Gly Pro Thr Leu Arg Glu Trp 15 10 15

Leu Gin Cys Arg Arg Met Gin 20

<210> 181 <211> 23 <212> PRT < 213 > artificial sequence <220><223> synthetic peptide sequence <220><221> misc_feature <222> (1). (1) <223> At position 1, Fc at N-terminus <400> 181

Gly Gly Gly Gly Gly Cys Ser Trp Gly Gly Pro Thr Leu Lys Gin Trp 15 10 15 -86- 114552.doc 1327149

Leu Gin Cys Val Arg Ala Lys 20 <210> 182 <211> 46 <212> PRT <213>Artificial Sequence<220><223> Synthetic Winning Sequence<220><221> ^feature <222> (1)..(1) <223> At position 1, Fc at N-terminus

<400> 182

Gly Gly Gly Gly Gly Ala Leu Arg Asp Gly Pro Thr Leu Lys Gin Trp 15 10 15

Leu Glu Tyr Arg Arg Gb Ala Gly Gly Gly Gly Gly Ala Leu Arg Asp 20 25 30

Gly Pro Thr Leu Lys Gin Trp Leu Glu Tyr Arg Arg Gin Ala 35 40 45 <210> 183 <211> 46 <212> PRT <213>

<220><223> Synthetic peptide sequence <220><221> miscjeature <222> (1)..(1) <223> At position 1, Fc at N-terminus <400> 183

Gly Gly Gly Gly Gly Glu Ala Leu Leu Gly Pro Thr Leu Arg Glu Trp 15 10 15

Leu Ala Trp Arg Arg Ala Gin Giy Gly Gly Gly Gly Glu Ala Leu Leu 20 25 30 -87· 114552.doc 1327149

Gly Pro Thr Leu Arg Glu Trp Leu Ala Trp Arg Arg Ala Gin 35 40 45

<210> 184 <2U> 46 <212> PRT <2Π> artificial sequence <220><223> synthetic belly sequence <220><221> misc_feature <222> (1). .(1)

<223> At position 1, Fc at N-terminus <400> 184

Gly Gly Gly Gly Gly Ala Val Pro Gin Gly Pro Thr Leu Lys Gin Trp 15 10 15

Leu Leu Trp Arg Arg Cys Ala Gly Gly Gly Gly Gly Ala Val Pro Gin 20 25 30

Gly Pro Thr Leu Lys Gin Trp Leu Leu Trp Arg Arg Cys Ala 35 40 45

<210> 185 <2ll> 46 <212> PRT < 213 > artificial sequence <220><223> will be a peptide sequence <220><221> miscjeature <222> (1) ..(1) <223> At position 1, Fc at N*terminus <400> 185

Gly Gly Gly Gly Gly Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp 15 10 15 -88 - 114552.doc 1327149

Leu Val Cys Leu Gly Leu Gin Gly Gly Gly Gly Gly Tyr Cys Asp Glu 20 25 30

Gly Pro Thr Leu Lys Gin Trp Leu Val Cys Leu Gly Leu Gin 35 40 45 <210> 186 <211> 46 <212> PRT <213>Artificial Sequence<220><22;3> Peptide sequence <220>

<221 > misc_feature <222>(1)..(1) <223> At position 1, Fc at N-terminus <400> 186

Gly Gly Gly Gly Gly Cys Ser Ser Gly Gly Pro Thr Leu Arg Glu Trp 15 10 15

Leu Gin Cys Arg Arg Met Gin Gly Gly Gly Gly Gly Cys Ser Ser Gly 20 25 30

Gly Pro Thr Leu Arg Glu Trp Leu Gin Cys Arg Arg Met Gin 35 40 45

<210> 187 <211> 46 <212> PRT < 213 > artificial sequence <220><2!D> synthetic peptide sequence <220><22l> miscjeature <222> )..(1) <223> At position 1, Fc at N-tcrminus <4〇0> 187

Gly Gly Gly Gly Gly Cys Ser Trp Gly Gly Pro Thr Leu Lys Gin Trp 15 10 15 -89- 114552.doc 1327149

Leu Gin Cys Val Arg Ala Lys Gly Gly Gly Gly Gly Cys Ser Trp Gly 20 25 30

Gly Pro Thr Leu Lys Gin Trp Leu Gin Cys Val Arg Ala Lys 35 40 45 <210> 188 <21!> 46 <212> PRT <213>Artificial Sequence<220><223> Peptide sequence Lu <220><221> misejeature <222> (47)..(47) <223> At position 47, Fc at C-terminus <4〇0> 188

Ala Leu Arg Asp Gly Pro Thr Leu Lys Gin Trp Leu Glu Tyr Arg Arg 15 10 15

Gin Ala Gly Gly Gly Gly Gly Ala Leu Arg Asp Gly Pro Thr Leu Lys 20 25 30

Gin Gin Gly Gly Gly Gly Gly Gly 35 40 45 <210><220><221> misejeature <222> (47)..(47) <223> At position 47, Fc at C-temiinus <4〇0> 189 -90- 114552.doc 1327149

Glu Ala Leu Leu Gly Fro Thr Leu Arg Glu Trp Leu Ala Trp Arg Arg 1 5 10 15

Ala Gin Gly Gly Gly Gly Gly Glu Ala Leu Leu Gly Pro Thr Leu Arg 20 25 30 .Glu Trp Leu Ala Trp Arg Arg Ala Gin Gly Gly Gly Gly Gly 35 40 45 <210> 190 <21i> 46 <212&gt ; PRT <213> artificial sequence

<220><223> Synthetic peptide sequence <220><221> misc.feature <222> (47)..(47) <223> At position 47, Fc at C-terminus <400> 190

Ala Val Pro Gin Gly Pro Thr Leu Lys Gin Trp Leu Leu Trp Arg Arg 15 10 15

Cys Ala Gly Gly Gly Gly Gly Ala Val Pro Gin Gly Pro Thr Leu Lys 20 25 30

Gin Trp Leu Leu Trp Arg Arg Cys Ala Gly Gly Gly Gly Gly 35 40 45 <210> 191 <211> 46 <212> PRT <213>Artificial Sequence<220><2ij> Synthetic peptide sequence <220><221> misc.feature <222> (47)..(47) <223> At position 47, Fc at C-tenninus •91 _ 114552.doc 1327149 <400> 191

Tyr Cys Asp Glu Gly Pro Thr Leu Lys Gin Trp Leu Val Cys Leu Gly 15 10 15

Leu Gin Gly Gy Gly Gly Gly Tyr Cys Asp Glu Gly Pro Thr Leu Lys 20 25 30

Gin Trp Leu Val Cys Leu Gly Leu Gin Gly Gly Gly Gly Gly 35 40 45 <210> 192 <211> 46 <212> PRT <213>

<220><223> Synthetic peptide sequence <220><221> misc_feature <222> (47)..(47) <223> At position 47, Fc at C-terminus <400> 192

Cys Ser Ser Gly Gly Pro Thr Leu Arg Glu Trp Leu Gla Cys Arg Arg 15 10 15

Met Gin Gly Gly Gly Gly Gly Cys Ser Ser Gly Gly Pro Thr Leu Arg 20 25 30

Glu Trp Leu Gin Cys Arg Arg Met Gin Gly Gly Gly Gly Gly 35 40 45 <210> 193 <21i> 46 <212> PRT <213>Artificial Sequence<220><223> Synthetic peptide sequence <220><22i> misc.feature <222> (47)..(47) <223> At position 47, Fc at C-terminus -92· U4552.doc 1327149 <400:. 193

Cys Ser Trp Gly Gly Pro Thr Leu Lys Gin Trp Leu Gin Cys Val Arg 15 10 15

Ala Lys Gly Gly Gly Gly Gly Cys Ser Trp Gly Gly Pro Thr Leu Lys 20 25 30

Gin Trp Leu Gin Cys Val Ar〇Ala Lys Gly Gly Gly Gly Gly 35 40 45 <210> 194 <211> 23 <212> PRT <213>Artificial Sequence<220><223> Sequence <220><221> misc^feature <222>(24)..(24) <223> At position 24, Fc at C-tenninus <4CX)> 194

Ala Leu Aig Asp Gly Pro Thr Leu L.ys Gin Trp Leu Glu Tyr Arg Arg 15 10 15

Gin Ala Gly Gly Gly Gly Gly 20 <210> 195 <211> 23 <212> PRT <213>Artificial Sequence, 220><223> Synthetic Limb Peptide Sequence <220><22i> miscjeature <222>(24)..(24) <223> At position 24, Fc at C-terrainus -93- 114552.doc 1327149 <400> 195

Glu Ala Leu Leu Gly Pro Thr Leu Ai*g Glu Trp Leu Ala Trp Arg Arg I 5 10 15

Ala Gin Gly Gly Gly Gly Gly 20 <210> 196 <211> 23 <2i2> PRT <213>Artificial Sequence <220>

<223> Synthetic peptide sequence <220><221> rniscjeature <222> (24)..(24) <223> At position 24, Fc at C-teminus <400>

Glu Ala Leu Leu Gly Pro Thr Leu Arg Glu Trp Leu Ala Trp Axg Arg 15 10 15

Ala Gin Gly Gly Gly Giy Gly 20

<210> 197 <211> 23 <212> PRT < 213 > artificial sequence <220><223> synthetic peptide sequence <220><22ni$c_featurc<222> (24). .(24) <223> At position 24, Fc at C-terminus <400> 197

Tyr Cys Asp Glu Gly Pro Tnr Leu Lys Gin Trp Leu Val Cys Leu Gly ί 5 10 15 -94· I14552.doc 1327149

Leu Gla Gly Gly Gly Gly Gly 20 <210> 193 <211> 23 <212> PRT <213>Artificial sequence <220><223> Synthetic peptide sequence <220><221> .feature <222> (24)..(24) <223> At position 24, Fc at C^tenninus

<400> 198

Cys Ser Ser Gly Gly Pro Thr Leu Arg GIu Trp Leu Gin Cys Arg Arg 15 10 15

Met Gin Gly Gly Gly Gly Gly 20 <2I0> 199 <211> 23 <212> PRT <213>Artificial Sequence <220>

<223> Synthetic peptide sequence <220><221> misc_feature <222> (24)..(24) <223> At position 24, Fc at C-tenr.inus <4〇Q> 199

Cys Scr Trp Gly Gly Pro Thr Leu Lys Gin Trp Leu Gin Cys Val Arg 15 10 15

Aia Lys Gly Gly Gly Gly Gly 20 U4552.doc -95-

Claims (1)

1327149 "Λ ' * 撕.n? -1 年月曰修(更)本本 ii Patent application No. 095133491... _> Chinese patent application scope replacement (99 years · 3' months) '_ 10. Patent Application Range: 1. A compound that binds to an mpl receptor, comprising a sequence selected from the group consisting of SEQ ID NO 2 to SEQ ID NO 30, comprising SEQ ID NO 2 and SEQ ID NO 30 peptide sequence of SEQ ID NO: GAREGPTLRQWLEWVRVG 2 RDLDGPTLRQWLPLPSVQ 3 ALRDGPTLKQWLEYRRQA 4 ARQEGPTLKEWLFWVRMG 5 EALLGPTLREWLAWRRAQ 6 MARDGPTLREWLRTYRMM 7 WMPEGPTLKQWLFHGRGQ 8 HIREGPTLRQWLVALRMV 9 QLGHGPTLRQWLSWYRGM 10 ELRQGPTLHEWLQHLASK 11 VGIEGPTLRQWLAQRLNP 12 WSRDGPTLREWLAWRAVG 13 AVPQGPTLKQWLLWRRCA 14 RIREGPTLKEWLAQRRGF 15 RFAEGPTLREWLEQRKLV 16 DRFQGPTLREWLAAIRSV 17 AGREGPTLREWLNMRVWQ 18 ALQEGPTLRQWLGWGQWG 19 YCDEGPTLKQWLVCLGLQ 20 114552-990308.doc 1327149 WCKEGPTLREWLRWGFLC 21 CSSGGPTLREWLQCRRMQ 22 CSWGGPTLKQWLQCVRAK 23 CQLGGPTLREWLACRLGA 24 CWEGGPTLKEWLQCLVER 25 CRGGGPTLHQWLSCFRWQ 26 CRDGGPTLRQ'WLACLQQK 27 ELRSGPTLKEWLVWRLA Q 28 GCRSGPTLREWLACREVQ 29 TCEQGPTLRQWLLCRQGR 30 2. A composition for binding to an mpl receptor, wherein the composition is selected from the group consisting of: (Vl)v-(LNl)1--(TMPl)a -(LN2)m-(TMP2)b-(LN3)n-(TMP3)c-(LN4)〇-(TMP4)d-(V2)w where TMP1, TMP2, TMP3 and TMP4 are each independently selected from SEQ a group consisting of ID NO 1 to SEQ ID NO 23, and a pharmaceutically acceptable salt thereof; LN1, LN2, LN3 and LN4 are each independently a linker comprising a peptide; V1 and V2 are each independently a Fc-containing a vehicle of a functional site; a, b, c and d are each independently an integer from 0 to 20, and at least one of a to d is not 0; 1, m, η and 〇 are each independently 0 to 20 An integer; and, 乂 and ... are each independently an integer of 〇1, and at least one of ν and w is not zero. 3. The composition according to claim 2, wherein VI and/or V2 comprises an IgGl Fc functional site. A di- or multi-polymer comprising a composition according to claim 1 of the scope of claim 114552-990308.doc -2-, or a composition of claim 2 of the patent. A pharmaceutical composition which comprises an effective amount of a composition according to the scope of the patent application 及其 2 and a pharmaceutically acceptable carrier thereof. 6 / The use of a composition according to item 2 of the patent application for the preparation of a medicament for the treatment of thrombocytopenia in a mammal. 7. The use of a composition according to item 2 of the scope of the patent application for the preparation of a medicament for increasing megakaryocytes or platelets of a patient in need thereof. g A compound that binds to an mpl receptor, which comprises the formula: '(Vl)v-(TMPl)a-(V2)w wherein VI and V2 are each an IgGl Fc functional site, but the restriction is at v At 1 o', w is 〇, and when v is 〇, ~ is 1, and ΤΜρι is a peptide having seq ID NO 2 to 30; a is an integer from 〇 to 2〇. 114552-990308.doc 1327149 Patent Application No. 095133491 Replacement Page (March 99) W _More) 正_ I 1 Ίmi σ> ο τ· cm co Body T-CMCO Inch 'T-CMCVJCMCMia QQQQQQQQQS H t Μ Η I On S brother ao 114552-fig-990308.doc -27-