| Aggrecanase molecules -> Monitor Keywords |
|
|
 
Aggrecanase moleculesRelated Patent Categories: Chemistry: Molecular Biology And Microbiology, Measuring Or Testing Process Involving Enzymes Or Micro-organisms; Composition Or Test Strip Therefore; Processes Of Forming Such Composition Or Test Strip, Involving Nucleic AcidAggrecanase molecules description/claimsThe Patent Description & Claims data below is from USPTO Patent Application 20070128616, Aggrecanase molecules. Brief Patent Description - Full Patent Description - Patent Application Claims
RELATED APPLICATION [0001] This application relies on the benefit of priority of U.S. provisional patent application No. 60/353,680, filed on Jan. 31, 2002, the entire disclosure of which is incorporated by reference. FIELD OF THE INVENTION [0002] The present invention relates to the discovery of nucleotide sequences encoding novel aggrecanase molecules, aggrecanase proteins and fragments thereof, and processes for producing them. The invention further relates to identification and development of inhibitors of and antibodies to the aggrecanase enzymes. These inhibitors and antibodies may be useful for the treatment of various aggrecanase-associated conditions including osteoarthritis. BACKGROUND OF THE INVENTION [0003] Aggrecan is a major extracellular component of articular cartilage. It is a proteoglycan responsible for providing cartilage with its mechanical properties of compressibility and elasticity. The loss of aggrecan has been implicated in the degradation of articular cartilage in arthritic diseases. Osteoarthritis is a debilitating disease which affects at least 30 million Americans (MacLean et al., J Rheumatol 25:2213-8 (1998)). Osteoarthritis can severely reduce quality of life due to degradation of articular cartilage and the resulting chronic pain. An early and important characteristic of the osteoarthritic process is loss of aggrecan from the extracellular matrix (Brandt and Mankin, Pathogenesis of Osteoarthritis, in Textbook of Rheumatology, WB Saunders Company, Philadelphia, Pa., at 1355-1373 (1993)). The large, sugar-containing portion of aggrecan is thereby lost from the extra-cellular matrix, resulting in deficiencies in the biomechanical characteristics of the cartilage. [0004] A proteolytic activity termed "aggrecanase" is believed to be responsible for the cleavage of aggrecan thereby having a role in cartilage degradation associated with osteoarthritis and inflammatory joint disease. Research has been conducted to identify the enzymes responsible for the degradation of aggrecan in human osteoarthritic cartilage. At least two enzymatic cleavage sites have been identified within the interglobular domain of aggrecan. One enzymatic cleavage site within the interglobular domain of aggrecan (Asn.sup.341-Phe.sup.342) has been observed to be cleaved by several known metalloproteases. Flannery et al., J Biol Chem 267:1008-14 (1992); Fosang et al., Biochemical J. 304:347-351 (1994). Cleavage at a second aggrecan cleavage site within aggrecan (Glu.sup.373-Ala.sup.374) due to IL-1 induced cartilage aggrecan cleavage results in the generation of an aggrecan fragment found in human synovial fluid (Sandy et al., J Clin Invest 69:1512-1516 (1992); Lohmander et aL., Arthritis Rheum 36:1214-1222 (1993); Sandy et al., J Biol Chem 266: 8683-8685 (1991)). Aggrecan cleavage at (Glu.sup.373-Ala.sup.374) has been attributed to aggrecanase activity (Sandy et al., J Clin Invest 69:1512-1516 (1992). This Glu.sup.373-Ala.sup.374 cleavage site will be referred to as the aggrecanase cleavage site. [0005] Recently, identification of two enzymes, aggrecanase-1 (ADAMTS4) and aggrecanase-2 (ADAMTS-11) within the "Disintegrin-like and Metalloprotease with Thrombospondin type 1 motif" (ADAMTS) family have been identified which are synthesized by IL-1 stimulated cartilage and cleave aggrecan at the Glu.sup.373-Ala.sup.374 site (Tortorella et al., Science 284:1664-6 (1999); Abbaszade et al., J Biol Chem 274: 23443-23450 (1999)). It is possible that these enzymes could be synthesized by osteoarthritic human articular cartilage. It is also contemplated that there are other, related enzymes in the ADAMTS family which are capable of cleaving aggrecan at the Glu.sup.373-Ala.sup.374 bond and could contribute to aggrecan cleavage in osteoarthritis. Therefore, there is a need to identify various aggrecanase enzymes and determine ways to block their enzymatic activity. SUMMARY OF THE INVENTION [0006] The present invention is directed to the identification of novel aggrecanase protein molecules capable of cleaving aggrecan, nucleotide sequences which encode the aggrecanase enzymes, and processes for the production of aggrecanases. These enzymes are contemplated to be characterized as having proteolytic aggrecanase activity. The invention further includes compositions comprising these enzymes. [0007] The invention also includes antibodies to these enzymes, in one embodiment, for example, antibodies that block aggrecanase activity. In addition, the invention includes methods for identifying and developing inhibitors of aggrecanase which block the enzyme's proteolytic activity. These inhibitors and antibodies may be used in various assays and therapies for treatment of conditions characterized by the degradation of articular cartilage. This invention provides nucleotide molecules that encode novel aggrecanase proteins. Accordingly, in one embodiment, the invention features an isolated DNA molecule comprising a DNA sequence chosen from: nucleotide #1 to nucleotide #3663 of SEQ ID NO: 1 (FIGS. 1A and 1B); fragments of SEQ ID NO: 1 which encode polypeptides or proteins that exhibit aggrecanase activity; variants of SEQ ID NO: 1 that encode proteins or polypeptides that exhibit aggrecanase activity, and fragments thereof; sequences which hybridize under stringent conditions with SEQ ID NO: 1; naturally occurring human allelic sequences; and equivalent degenerative codon sequences [0008] In another aspect, the invention comprises an isolated aggrecanase protein comprising an amino acid sequence chosen from: amino acid #1 (methionine) to amino acid #1221 (isoleucine) of SEQ ID NO: 2 (FIG. 2); fragments of SEQ ID NO: 2 which exhibit aggrecanase activity, and variants and fragments of aggrecanase proteins that exhibit proteolytic activity, including deletion and substitution mutants. In yet another aspect, the invention provides methods for producing an isolated aggrecanase protein. One such method includes (1) transforming a host cell with a DNA sequence, such as the DNA sequence depicted in SEQ ID NO: 1; (2) culturing the host cell; and (3) purifying the aggrecanase enzyme set forth in SEQ ID NO: 2 that is encoded by the DNA sequence, from the cell culture medium. [0009] The invention also provides antibodies that bind to isolated aggrecanase proteins of the invention. In one embodiment, such an antibody reduces, inhibits or antagonizes aggrecanase activity. The invention further provides methods for developing and identifying inhibitors of aggrecanase activity comprising the use of aggrecanase protein chosen from SEQ ID NO: 2 or a fragment or a variant thereof. In one embodiment, inhibitors of aggrecanase activity prevent cleavage of aggrecan. [0010] Additionally, the invention provides pharmaceutical compositions for inhibiting the proteolytic activity of aggrecanase, wherein the compositions comprise at least one antibody according to the invention and at least one pharmaceutical carrier. The invention also provides methods for inhibiting aggrecanase activity in a mammal comprising administering to the mammal an effective amount of a pharmaceutical composition according to the invention to inhibit aggrecanase activity. [0011] Additional aspects of the disclosure will be set forth in part in the description, and in part be obvious from the description, or may be learned from practicing the invention. The invention is set forth and particularly pointed out in the claims, and the disclosure should not be construed as limiting the scope of the claims. The following detailed description includes exemplary representations of various embodiments of the invention, which are not restrictive of the invention as claimed. The accompanying figures constitute a part of this specification and, together with the description, serve to illustrate embodiments and not limit the invention. BRIEF DESCRIPTION OF THE FIGURES AND SEQUENCES [0012] FIGS. 1A and 1B showthe full-length nucleotide sequence for ADAMTS-18 (EST18). (SEQ ID NO: 1) [0013] FIG. 2 shows the full-length amino acid sequence for ADAMTS-18, based on the nucleotide sequence of SEQ ID NO: 1. (SEQ ID NO: 2) [0014] FIGS. 3A and 3B show a nucleotide sequence of ADAMTS-18 (EST18). (SEQ ID NO: 3) [0015] FIG. 4 shows the predicted amino acid sequence of ADAMTS-18 based on the nucleotide sequence of SEQ ID NO: 3. (SEQ ID NO: 4) [0016] FIGS. 5A and 5B show a virtual nucleotide sequence for ADAMTS-18, which was identified by Celera database-mining techniques. (SEQ ID NO: 5) FIG. 6A shows a schematic representation of the PCR primers used for amplification of fragments of a EST18 nucleotide sequence. FIG. 6B shows a schematic representation of the overlapping nucleotide sequence fragments of EST18 including sites for restriction enzymes. [0017] FIG. 7 shows a nucleotide sequence encoding for atruncated form of ADAMTS-18 linked to a Streptavidin-tag. (SEQ ID NO: 7) [0018] FIG. 8 shows an amino acid sequence for a truncated form of ADAMTS-18 including a Streptavidin-tag, based on SEQ ID NO: 7. (SEQ ID NO: 8) [0019] FIG. 9 shows a schematic representation of the hydrophobic plot generated for the protein of SEQ ID NO: 2 using the GCG plotstructure program. Continue reading about Aggrecanase molecules... Full patent description for Aggrecanase molecules Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Aggrecanase molecules patent application. ###  How KEYWORD MONITOR works... a FREE service from FreshPatents1. Sign up (takes 30 seconds). 2. Fill in the keywords to be monitored. 3. Each week you receive an email with patent applications related to your keywords. Start now! - Receive info on patent apps like Aggrecanase molecules or other areas of interest. ### Previous Patent Application: Therapeutic, prophylactic and diagnostic agents Next Patent Application: Alpha macroglobulin family member Industry Class: Chemistry: molecular biology and microbiology ### FreshPatents.com Support Thank you for viewing the Aggrecanase molecules patent info. IP-related news and info Results in 0.17397 seconds Other interesting Feshpatents.com categories: Software: Finance , AI , Databases , Development , Document , Navigation , Error 174 |
 * Protect your Inventions * US Patent Office filing 
PATENT INFO |
|
