FIELD OF THE INVENTION

The present invention relates to the development, maintenance and repair of connective tissue. More specifically, the invention relates to the identification of novel peptides that are capable of influencing the formation and repair of skeletal tissues which include bone, cartilage and related connective tissues. The invention further relates to pharmaceutical compositions of these peptides, as well as therapeutic and prophylactic uses of such peptides.

BACKGROUND OF THE INVENTION

Throughout this application, various references are cited in parentheses to describe more fully the state of the art to which this invention pertains. The disclosure of these references are hereby incorporated by reference into the present disclosure.

Various agents have been identified and characterized for use in the stimulation of bone and/or cartilage formation and tissue repair. Examples of such agents may include for example, calcium preparations (e.g. calcium carbonate), calcitonin preparations, sex hormones (e.g. estrogen, estradiol), prostaglandin A1, bisphosphonic acids, ipriflavones, fluorine compounds (e.g. sodium fluoride), vitamin K, bone morphogenetic proteins (BMPs), fibroblast growth factor (FGF), platelet-derived growth factor (PDGF), transforming growth factor (TGF-β), insulin-like growth factors 1 and 2 (IGF-1, IGF-2), parathyroid hormone (PTH), epidermal growth factor (EGF), leukemia inhibitory factor (LIF), osteogenin, and bone resorption repressors such as estrogens, calcitonin and biphosphonates.

U.S. Pat. Nos. 5,169,837 and 5,403,825 disclose an osteogenic factor and method of extraction from crude extracts of mammalian bone. The osteogenic factor is capable of inducing bone growth as determined using a rat bone growth assay. U.S. Pat. No. 5,599,792 discloses parathyroid hormone (PTH) variants that exhibit bone stimulating activity and reduced vasoactivity.

Various patents disclose bone morphogenetic proteins (BMPs) and methods of isolating such proteins. BMPs are useful for the treatment of bone and cartilage defects. U.S. Pat. No. 4,761,471 discloses a method for purifying a mixed BMP composition from demineralized bone tissue; U.S. Pat. No. 5,013,649 discloses bone morphogenetic protein-3 (BMP-3); U.S. Pat. No. 5,106,748 discloses bone morphogenetic protein-5 (BMP-5); U.S. Pat. No. 6,187,076 discloses bone morphogenetic protein-6 (BMP-6); U.S. Pat. No. 5,141,905 discloses bone morphogenetic protein-7 (BMP-7); PCT Publication No. WO91/18098 discloses bone morphogenetic protein-8 (BMP-8); U.S. Pat. No. 6,287,816 discloses bone morphogenetic protein-9 (BMP-9); and PCT Publication No. WO94/26893 discloses bone morphogenetic protein-10 (BMP-10).

U.S. Pat. No. 6,352,973 discloses bone stimulating factor polypeptides for use in the treatment of bone disease. U.S. Pat. No. 6,344,439 discloses agents for promoting bone formation, the agents comprise the RGD binding sequence. U.S. Pat. No. 5,958,428 discloses synthetic compounds that promote cell binding with collagen.

While the aforementioned agents do have general use in aspects of bone repair, they also have various shortcomings. For example, some of the agents are relatively non-specific and thus not very effective at promoting bone tissue repair. Certain agents such as extracted factors are associated with the risk of immunogenicity, infection and/or disease transmission. Extraction from crude extracts of tissue or preparation via recombinant methods are also costly thus compromising commercial viability. Furthermore, some agents have secondary undesirable systemic or local effects. Most notable is the fact that several of the known agents may cause ectopic bone formation. Such abnormal bone formation is highly undesirable and may lead to serious medical complications.

There is therefore a need to provide novel peptides that have specific in vitro, in vivo, or ex vivo use for effectively influencing development, maintenance and repair of connective tissues.

SUMMARY OF THE INVENTION

The present invention provides a number of novel peptides and analogues thereof which are influential in skeletal biology in that the peptides serve to stimulate the normal formation of bone, cartilage and related connective tissues in vitro, in vivo, ex vivo and for tissue engineering applications. The functional analogues of these novel peptides contain one or more amino acid additions, substitutions or deletions to the disclosed peptide sequences.

The novel peptides herein described are generically referred to as BCSP (“bone andcartilagestimulatingpeptides”).

The invention also relates to nucleic acids encoding the isolated connective tissue promoting peptides and analogues thereof, expression vectors containing the nucleic acid molecules, host cells containing the nucleic acid molecules, compositions of such peptides and nucleic acids encoding therefor and antibodies to those peptides.

The novel peptides and nucleic acids encoding therefor can be used in compositions administered using a variety of carriers and methods to promote the formation and function of various connective tissues such as bone and cartilage. As such, the peptides and analogues thereof can be used to treat a variety of skeletal abnormalities which include but not limited to bone and cartilage defects due to trauma and/or disease.

The peptides and functional analogues thereof can be used in a variety of in vitro, in vivo or ex vivo applications and methods. Furthermore, the peptides and functional analogues thereof also have use in a variety of tissue engineering applications.

In one embodiment, the present invention provides therapeutic compositions and methods for the treatment of disorders involving abnormal bone and/or cartilage formation and associated abnormal skeletal development resulting from disease, arthritis and degenerative joint diseases involving cartilage degeneration. The therapeutic compositions and methods can also be used for the treatment of trauma induced skeletal or cartilage damage.

According to an aspect of the invention is a peptide comprising an amino acid motif selected from the group consisting of “PG”, “GP”, “PI” and “IG” and having up to 10 amino acids upstream and/or downstream of said amino acid motif, wherein “P” in the motif is proline or hydroxyproline, the peptide stimulating the development, maintenance and repair of bone, cartilage and associated connective tissue.

According to another aspect of the invention is a peptide comprising one or more “GP” and/or “PG” motifs and comprising up to a total of about 20 amino acids, wherein “P” in the motif is proline or hydroxyproline and the peptide induces the activity of bone and cartilage cells such that bone and cartilage tissue is developed, maintained and/or repaired.

The peptides of the invention stimulate the activity of bone cells, bone cell precursors and cells sharing lineage therewith. Similarly, the peptides of the invention stimulate the activity of cartilage cells, cartilage cell precursors and cells sharing lineage therewith. As such, these peptides have use in the stimulation of the formation of bone and cartilage as used in vitro, ex vivo and/or in vivo.

The peptide of the invention can be as few as three amino acids in length and more preferably, at least four amino acids in length. The peptide can further comprise one or more linkers, calcium binding motifs and agents and used for local or systemic applications.

According to another aspect of the invention, there is provided a peptide having a formula selected from the group consisting of:

(I) X—(P1)mGly(P2)n, (II) (P1)mGly(P2)n—Z, and (III) X—(P1)mGly(P2)n—Z;wherein P1and P2are selected from the group consisting of proline and hydroxyproline, m=0 or 1, n=0 or 1, where m and n are independently selected;wherein X is selected from the group consisting of leucine, glycine-leucine, proline-isoleucine, glycine-proline-isoleucine, asparagine-glycine-leucine and proline-glycine-proline-isoleucine-glycine-proline (SEQ. ID. NO.9); andwherein Z is selected from the group consisting of isoleucine, arginine, isoleucine-glycine, isoleucine-glycine-proline, isoleucine-glycine-hydroxyproline, isoleucine-glycine-proline-proline (SEQ. ID. NO.21), isoleucine-glycine-proline-proline-glycine (SEQ. ID. NO.22), isoleucine-glycine-proline-proline-glycine-proline (SEQ. ID. NO.23), isoleucine-glycine-proline-proline-glycine-proline-arginine (SEQ. ID. NO.24), isoleucine-glycine-proline-proline-glycine-proline-arginine-glycine-arginine-threonine-glycine-aspartate-alanine (SEQ. ID. NO.25) and arginine-glycine-arginine-threonine-glycine-aspartate-alanine (SEQ. ID. NO.26).

Accordingly, in an aspect of the invention there are provided peptides having the following amino acid sequences and designated BCSP1 which has the amino acid sequence NGLPGPIGP* (SEQ. ID. NO. 1); BCSP2 which has the amino acid sequence NGLPGPIG (SEQ. ID. NO.2); BCSP3 which has the amino acid sequence NGLP*GPIG (SEQ. ID. NO.3); BCSP4 which has the amino acid sequence NGLP*GPIGP* (SEQ. ID. NO.4); BCSP5 which has the amino acid sequence NGLPGP (SEQ ID. NO.5); BCSP6 which has the amino acid sequence LPGP (SEQ. ID. NO.6); BCSP7 which has the amino acid sequence PGPIG (SEQ ID. NO.7); BCSP8 which has the amino acid sequence NGLPGPIGP (SEQ. ID. NO.8); BCSP9 which has the amino acid sequence PGPIGP (SEQ ID. NO.9); BCSP10 which has the amino acid sequence PGPIGPPGPR (SEQ. ID. NO.10); BCSP11 which has the amino acid sequence GPIG (SEQ. ID. NO.11); BCSP12 which has the amino acid sequence PGPIGPPGPRGRTGDA (SEQ. ID. NO.12); BCSP13 which has the amino acid sequence GPRGRTGDA (SEQ. ID. NO.13); BCSP14 which has the amino acid sequence PGPIGPP (SEQ. ID. NO.14); BCSP15 which has the amino acid sequence PGPIGP* (SEQ. ID. NO.15); BCSP16 which has the amino acid sequence PGPI (SEQ. ID. NO.16); BCSP17 which has the amino acid sequence GLPGPIG (SEQ. ID. NO.17); BCSP18 which has the amino acid sequence GPIGP* (SEQ. ID. NO.18); BCSP19 which has the amino acid sequence GPIGP (SEQ. ID. NO.19); and BCSP20 which has the amino acid sequence PIGP (SEQ. ID. NO.20). As used herein, P* is used to identify hydroxyproline. Each of these peptides can be used to elicit a connective tissue promoting response in vitro, in vivo and ex vivo.

In another aspect, the invention provides functional analogues of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 for eliciting a connective tissue promoting response in vitro, in vivo and ex vivo.

In another aspect, the invention provides mimetic analogues of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 for eliciting a connective tissue promoting response in vitro, in vivo and ex vivo.

In a further aspect of the invention, there are provided compositions for stimulating a connective tissue response in a mammal, for example, a human, said compositions comprising an effective amount of one or more of the peptides selected from the group consisting of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 as well as functional analogues thereof and/or functional mimetics thereof together with a pharmaceutically acceptable diluent or carrier. Such compositions may be formulated to contain additional adjuvant(s), co-stimulatory molecules and/or stabilizers.

In accordance with another embodiment of the present invention is a composition for the stimulation of the development, maintenance and repair of bone, cartilage and related connective tissues, the composition comprising a peptide having a formula selected from the group consisting of:

(I) X—(P1)mGly(P2)n, (II) (P1)mGly(P2)n—Z and (III) X—(P1)mGly(P2)n—Z;

wherein P1and P2are selected from the group consisting of proline and hydroxyproline, m=0 or 1, n=0 or 1, where m and n are independently selected;

wherein X is selected from the group consisting of leucine, glycine-leucine, proline-isoleucine, glycine-proline-isoleucine, asparagine-glycine-leucine and proline-glycine-proline-isoleucine-glycine-proline (SEQ. ID. NO.9);

wherein Z is selected from the group consisting of isoleucine, arginine, isoleucine-glycine, isoleucine-glycine-proline, isoleucine-glycine-hydroxyproline, isoleucine-glycine-proline-proline (SEQ. ID. NO.21), isoleucine-glycine-proline-proline-glycine (SEQ. ID. NO.22), isoleucine-glycine-proline-proline-glycine-proline (SEQ. ID. NO.23), isoleucine-glycine-proline-proline-glycine-proline-arginine (SEQ. ID. NO.24), isoleucine-glycine-proline-proline-glycine-proline-arginine-glycine-arginine-threonine-glycine-aspartate-alanine (SEQ. ID. NO.25), arginine-glycine-arginine-threonine-glycine-aspartate-alanine (SEQ. ID. NO.26); and

a pharmaceutically acceptable carrier.

In accordance with another aspect of the present invention is a method for stimulating the development, maintenance and repair of bone, cartilage and associated connective tissue in a mammal, the method comprising administering an effective amount of a peptide comprising an amino acid motif selected from the group consisting of “PG”, “GP”, “PI” and “IG” and having up to 10 amino acids upstream and/or downstream of said amino acid motif, wherein “P” in the motif is proline or hydroxyproline.

In accordance with a further embodiment of the present invention is a therapeutic method for promoting connective tissue formation in a mammal which comprises administering to a mammal a peptide having a formula selected from the group consisting of:

(I) X—(P1)mGly(P2)n, (II) (P1)mGly(P2)n—Z and (III) X—(P1)mGly(P2)n—Z;

wherein P1and P2are selected from the group consisting of proline and hydroxyproline, m=0 or 1, n=0 or 1, where m and n are independently selected;

wherein X is selected from the group consisting of leucine, glycine-leucine, proline-isoleucine, glycine-proline-isoleucine, asparagine-glycine-leucine and proline-glycine-proline-isoleucine-glycine-proline (SEQ. ID. NO.9);

wherein Z is selected from the group consisting of isoleucine, arginine, isoleucine-glycine, isoleucine-glycine-proline, isoleucine-glycine-hydroxyproline, isoleucine-glycine-proline-proline (SEQ. ID. NO.21), isoleucine-glycine-proline-proline-glycine (SEQ. ID. NO.22), isoleucine-glycine-proline-proline-glycine-proline (SEQ. ID. NO.23), isoleucine-glycine-proline-proline-glycine-proline-arginine (SEQ. ID. NO.24), isoleucine-glycine-proline-proline-glycine-proline-arginine-glycine-arginine-threonine-glycine-aspartate-alanine (SEQ. ID. NO.25) and arginine-glycine-arginine-threonine-glycine-aspartate-alanine (SEQ. ID. NO.26);

or a functional analogue or variant of (I), (II) and/or (III).

In accordance with still a further embodiment of the invention is a therapeutic method for promoting connective tissue formation in a subject, said method comprising administering to said subject one or more of the peptides selected from the group consisting of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 as well as functional analogues thereof.

In accordance with a further embodiment, the invention provides a method for treating damaged bone, cartilage and related tissue in a subject, comprising administering to the subject an effective amount of one or more of the peptides selected from the group consisting of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 and functional analogues thereof and/or mimetic analogues thereof wherein the peptide(s) stimulate connective tissue repair and formation.

In accordance with a further embodiment, the invention provides a method for enhancing osseous integration of orthopedic or dental implants in a subject comprising administering to the subject an effective amount of one or more of the peptides selected from the group consisting of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 and functional analogues thereof.

According to a further embodiment of the invention, there is provided a method of producing cartilage at a cartilage defect site in vivo, the method comprising:

implanting into the defect site a population of chondrogenic cells which have been cultured in vitro in the presence of one or more of the peptides selected from the group consisting of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 and functional analogues thereof.

The present invention encompasses methods of promoting cartilage repair at a cartilage defect site in vivo wherein de novo cartilage prepared by tissue engineering techniques in the presence of one or more of the peptides selected from the group consisting of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 and functional analogues thereof, has been implanted into the defect site.

According to another embodiment of the invention, there is provided a method for treating a degenerative joint disease characterized by cartilage degeneration, the method comprising:

delivering a therapeutically effective amount of one or more of the peptides selected from the group consisting of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 and functional analogues thereof to a disease site.

In accordance with a further embodiment, the invention provides a method for treating bone associated disorders in a mammal, comprising administering to the mammal's cells selected from the group consisting of osteoblastic cells, pre-osteoblastic cells, chondrocytes, pre-chondrocytes, skeletal progenitor cells derived from bone, bone marrow or blood, and mixtures thereof, treated with an effective amount of one or more of the peptides selected from the group consisting of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 and functional analogues thereof.

The methods of the invention may involve providing systemic and/or local administration of the selected peptide(s) or functional analogues of the invention.

According to another aspect of the invention, there is provided a morphogenetic device for implantation at a bone site in a vertebrate, the device comprising:

an implantable biocompatible carrier; and

one or more of the peptides selected from the group consisting of BCSP1, BCSP2, BCSP3, BCSP4, BCSP5, BCSP6, BCSP7, BCSP8, BCSP9, BCSP10, BCSP11, BCSP12, BCSP13, BCSP14, BCSP15, BCSP16, BCSP17, BCSP18, BCSP19 and BCSP20 and functional analogues thereof provided or dispersed within or on said carrier.

According to another aspect of the invention is a method for stimulation of bone development in a mammal, said method comprising administering an effective amount of one or more peptides selected from the group consisting of SEQ. ID. NO.1, SEQ. ID. NO.4, SEQ. ID. NO.7, SEQ. ID. NO.8, SEQ. ID. NO.9, SEQ. ID. NO.10, SEQ. ID. NO.12, SEQ. ID. NO.15 and SEQ. ID. NO.18 as well as functional analogues and mimetic analogues thereof.

According to yet another aspect of the invention is a method for stimulation of cartilage development in a mammal, said method comprising administering an effective amount of one or more peptides selected from the group consisting of SEQ. ID. NO.1, SEQ. ID. NO.4, SEQ. ID. NO.7, SEQ. ID. NO.8, SEQ. ID. NO.9, SEQ. ID. NO.10, SEQ. ID. NO.11 as well as functional analogues and mimetic analogues thereof.

According to yet another aspect of the invention is a method for screening for a peptide that stimulates bone and/or cartilage formation and associated connective tissue, said method comprising the steps of:

(a) contacting a first biological sample capable of undergoing bone and/or cartilage formation with a test peptide;

(b) separately contacting a second biological sample capable of undergoing bone and/or cartilage formation with a biologically effective amount of a peptide selected from the group consisting of NGLPGPIGP* (SEQ. ID. NO. 1); NGLPGPIG (SEQ. ID. NO.2); NGLP*GPIG (SEQ. ID. NO.3); NGLP*GPIGP* (SEQ. ID. NO.4); NGLPGP (SEQ ID. NO.5); LPGP (SEQ. ID. NO.6); PGPIG (SEQ ID. NO.7); NGLPGPIGP (SEQ. ID. NO.8); PGPIGP (SEQ ID. NO.9); PGPIGPPGPR (SEQ. ID. NO.10); GPIG (SEQ. ID. NO.11); PGPIGPPGPRGRTGDA (SEQ. ID. NO.12); GPRGRTGDA (SEQ. ID. NO.13); PGPIGPP (SEQ. ID. NO.14); PGPIGP* (SEQ. ID. NO.15); PGPI (SEQ. ID. NO.16); GLPGPIG (SEQ. ID. NO.17); GPIGP* (SEQ. ID. NO.18); GPIGP (SEQ. ID. NO.19); and PIGP (SEQ. ID. NO.20) thereby providing a control;

(c) assessing the level of bone and/or cartilage formation resulting from step (a) and from step (b); and

(d) comparing the levels of bone and/or cartilage formation assessed in step (c), whereby a peptide capable of stimulating bone and/or cartilage formation is identified by its ability to alter the level of bone and/or cartilage formation when compared to the control of step (b).

According to still another aspect of the invention is a method for screening for a modulator that stimulates bone and/or cartilage formation and associated connective tissue, said method comprising the steps of:

(a) contacting a first biological sample capable of undergoing bone and/or cartilage formation with a test compound and a biologically effective amount of a peptide selected from the group consisting of NGLPGPIGP* (SEQ. ID. NO. 1); NGLPGPIG (SEQ. ID. NO.2); NGLP*GPIG (SEQ. ID. NO.3); NGLP*GPIGP* (SEQ. ID. NO.4); NGLPGP (SEQ ID. NO.5); LPGP (SEQ. ID. NO.6); PGPIG (SEQ ID. NO.7); NGLPGPIGP (SEQ. ID. NO.8); PGPIGP (SEQ ID. NO.9); PGPIGPPGPR (SEQ. ID. NO.10); GPIG (SEQ. ID. NO.11); PGPIGPPGPRGRTGDA (SEQ. ID. NO.12); GPRGRTGDA (SEQ. ID. NO.13); PGPIGPP (SEQ. ID. NO.14); PGPIGP* (SEQ. ID. NO.15); PGPI (SEQ. ID. NO.16); GLPGPIG (SEQ. ID. NO.17); GPIGP* (SEQ. ID. NO.18); GPIGP (SEQ. ID. NO.19); and PIGP (SEQ. ID. NO.20);

(b) separately contacting a second biological sample with a biologically effective amount of a peptide selected from the group consisting of NGLPGPIGP* (SEQ. ID. NO. 1); NGLPGPIG (SEQ. ID. NO.2); NGLP*GPIG (SEQ. ID. NO.3); NGLP*GPIGP* (SEQ. ID. NO.4); NGLPGP (SEQ ID. NO.5); LPGP (SEQ. ID. NO.6); PGPIG (SEQ ID. NO.7); NGLPGPIGP (SEQ. ID. NO.8); PGPIGP (SEQ ID. NO.9); PGPIGPPGPR (SEQ. ID. NO.10); GPIG (SEQ. ID. NO.11); PGPIGPPGPRGRTGDA (SEQ. ID. NO.12); GPRGRTGDA (SEQ. ID. NO.13); PGPIGPP (SEQ. ID. NO.14); PGPIGP* (SEQ. ID. NO.15); PGPI (SEQ. ID. NO.16); GLPGPIG (SEQ. ID. NO.17); GPIGP* (SEQ. ID. NO.18); GPIGP (SEQ. ID. NO.19); and PIGP (SEQ. ID. NO.20) thereby providing a control;

(c) assessing the level of bone and/or cartilage formation resulting from step (a) and from step (b); and

(d) comparing the levels of bone and/or cartilage formation assessed in step (c), whereby a modulator of bone and/or cartilage formation is identified by its ability to alter the level of bone and/or cartilage formation when compared to the control of step (b).

According to yet another aspect of the invention is a method for identifying peptides that stimulate the development, maintenance and repair of bone, cartilage and associated connective tissue, said method comprising:

determining whether said peptide has comparable activity to one or more of the peptides selected from the group consisting of NGLPGPIGP* (SEQ. ID. NO. 1); NGLPGPIG (SEQ. ID. NO.2); NGLP*GPIG (SEQ. ID. NO.3); NGLP*GPIGP* (SEQ. ID. NO.4); NGLPGP (SEQ ID. NO.5); LPGP (SEQ. ID. NO.6); PGPIG (SEQ ID. NO.7); NGLPGPIGP (SEQ. ID. NO.8); PGPIGP (SEQ ID. NO.9); PGPIGPPGPR (SEQ. ID. NO.10); GPIG (SEQ. ID. NO.11); PGPIGPPGPRGRTGDA (SEQ. ID. NO.12); GPRGRTGDA (SEQ. ID. NO.13); PGPIGPP (SEQ. ID. NO.14); PGPIGP* (SEQ. ID. NO.15); PGPI (SEQ. ID. NO.16); GLPGPIG (SEQ. ID. NO.17); GPIGP* (SEQ. ID. NO.18); GPIGP (SEQ. ID. NO.19); and PIGP (SEQ. ID. NO.20);

wherein the activity is assessed by the analysis of one or more criteria selected from the group consisting of bone mineral density, bone mineral content, alkaline phosphatase activity, proliferation of osteoblasts, bone nodule formation, bone nodule mineralization, chondrocyte proliferation, collagen assay and proteoglycan assay.

Other features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples while indicating embodiments of the invention are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from said detailed description.

