Organic compounds -- part of the class 532-570 series – Organic compounds – Carbohydrates or derivatives
Patent
1997-07-30
1999-11-23
Jones, W. Gary
Organic compounds -- part of the class 532-570 series
Organic compounds
Carbohydrates or derivatives
536 243, 536 232, 530350, 435 6, C07H 2102, C07H 2104, C07K 1400, C12Q 168
Patent
active
059902930
DESCRIPTION:
BRIEF SUMMARY
This invention relates to a novel human metalloproteinase, to homologues and fragments thereof, to means for producing the metalloproteinase, and to means for regulating its production and activity in vivo.
A number of physiologically important processing events are mediated by metalloproteinases, which under certain circumstances may contribute to pathologies as diverse as inflammation and cancer, and it has been suggested that such enzymes would provide targets for therapeutic intervention. Thus, by varying the production of the enzyme, or inhibiting or enhancing its activity in vivo it should be possible to achieve a therapeutic effect.
In one example, tumour necrosis factor-alpha (TNF-.alpha.) is a potent pro-inflammatory and immunomodulatory mammalian cytokine produced primarily by activated monocytes and macrophages. It is initially expressed as a 233-amino-acid membrane-anchored precursor (pro-TNF-.alpha.) which is proteolytically processed to yield the mature, 157-amino-acid cytokine. Evidence has been obtained which indicates that at least one metalloproteinase-like enzyme mediates pro-TNF-.alpha. cleavage, but to date the enzyme(s) responsible for this in vivo are unknown [see for example Mohler, K M et al, Nature 370, 218-220 (1994); Gearing, A J et al 370, 555-557 (1994); McGeehan, G M et al, ibid 370, 558-561 (1994)]. A number of known matrix metalloproteinase inhibitors have been shown to block TNF-.alpha. secretion [see the above papers and International Patent Specification Publication No. WO 95/06031]. These compounds were originally designed to selectively inhibit matrix metalloproteinases such as collagenase with primary functions unrelated to pro-TNF-.alpha. cleavage. Where new inhibitors have been described these have apparently been selected on the basis of their effect on TNF-.alpha. secretion seen in cell-based assays.
In another example, L-selectin shedding is thought to be a pro-inflammatory event that is mediated by an as yet unidentified metalloproteinase [Lasky, Science, 258, 964-969 (1992)]. Some inhibitors of L-selectin proteolysis have been identified, but these have been obtained using cell based assays [Walchech et al., Nature, 380, 720-723 (1996); Feehan et al., J. Biol. Chem., 271, 7019-7024 (1996)].
In general, in order to obtain compounds capable of selectively regulating the action of a metalloproteinase implicated in human disease, for example as in the above TNF-.alpha. and L-selectin instances it would be clearly advantageous to have the enzyme unequivocally identified and obtainable in an isolated, purified and unambiguous characterised form.
Through the use of a cloning and screening approach, we have been able to identify human DNA which is responsible for coding one such metalloproteinase. This DNA has the sequence described in SEQ I.D. No: 1 below and may be of use (1) in the production of the metalloproteinase, (2) in the provision of means to regulate the activity of the metalloproteinase in vivo, and (3) in the provision of means to detect and measure a metalloproteinase in a biological system, e.g. in serum, synovial fluid or a tissue extract.
Thus according to one aspect of the invention we provide isolated human DNA comprising the nucleotide sequence of SEQ I.D. No: 1:
SEQ I.D. NO: 1 GAGAAGAGCAGACACCGTGCTCCTGGAATCACCCAGCATGTTGCAA
GGTCTCCTGCCAGTCAGTCTCCTCCTCTCTGTTGCAGTAAGTGCTAT
AAAAGAACTCCCTGGGGTGAAGAAGTATGAAGTGGTTTATCCTATAA
GACTTCATCCACTGCATAAAAGAGAGGCCAAAGAGCCAGAGCAACAG
GAACAATTTGAAACTGAATTAAAGTATAAAATGACAATTAATGGAAAAA
TTGCAGTGCTTTATTTGAAAAAAAACAAGAACCTCCTTGCACCAGGCT
ACACGGAAACATATTATAATTCCACTGGAAAGGAGATCACCACAAGC
CCACAAATTATGGATGATTGTTATTATCAAGGACATATTCTTAATGAAA
AGGTTTCTGACGCTAGCATCAGCACATGTAGGGGTCTAAGGGGCTAC
TTCAGTCAGGGGGATCAAAGATACTTTATTGAACCTTTAAGCCCCATA
CATCGGGATGGACAGGAGCATGCACTCTTCAAGTATAACCCTGATGA
AAAGAATTATGACAGCACCTGTGGGATGGATGGTGTGTTGTGGGCCC
ACGATTTGCAGCAGAACATTGCCCTACCTGCCACCAAACTAGTAAAAT
TGAAAGACAGGAAGGTTCAGGAACATGAGAAATACATAGAATATTATT
TGGTCCTGGATAATGGTGAGTTTAAAAGGTACAATGAGAATCAAGAT
GAGATCAGAAAGAGGGTATTTGAGAT
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Docherty Andrew James Penrose
Slocombe Patrick Marcel
Celltech Therapeutics Limited
Conlin David G.
Hazzard Lisa Swiszcz
Jones W. Gary
Whisenant Ethan
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