Organic compounds -- part of the class 532-570 series – Organic compounds – Carbohydrates or derivatives
Patent
1996-11-12
1999-09-28
McKelvey, Terry
Organic compounds -- part of the class 532-570 series
Organic compounds
Carbohydrates or derivatives
C12N 1511
Patent
active
059590947
DESCRIPTION:
BRIEF SUMMARY
FIELD OF THE INVENTION
The present invention relates to a promoter sequence for the p75 tumor necrosis factor receptor (TNF-R), to its preparation and use.
BACKGROUND OF THE INVENTION
TNF is a multifunctional pro-inflammatory cytokine which affects a wide range of cellular functions. On the one hand, TNF is involved in the protection of the organism, but on the other hand, when over-produced, it can play a major pathogenic role in several diseases. TNF is known to be involved in inflammatory processes and to be a mediator of the damage to tissues in septic shock (1), graft-versus-host reactions (2) and in rheumatic diseases (3).
TNF exerts these effects by binding to two distinct cell surface receptors, which differ in size (about 55 and 75 kDa) and possess structurally dissimilar intracellular domains, suggesting that they signal differently (4-11). Almost all cells express TNF receptors (TNF-Rs), yet the amounts and relative proportions of the two receptors vary in different cell types. These variations are in part developmentally controlled; they are related to the phenotype of the cell and its state of differentiation, and in part can be induced transiently by cytokines and immune stimulatory components of pathogens (12-22). Studies of the function of the two TNF-Rs indicate that they have different yet interacting activities (23-28), and that their activity level may be correlated to the extent of their expression by the cell (29, 30). These findings imply that the mechanisms which affect the amounts and relative proportion of the two TNF-Rs can have significant influence both on the nature and the extent of the cellular response to TNF and thus constitute important determinants of the physiological as well as pathological manifestations of the function of this cytokine.
In order to inhibit the deleterious effects of TNF, ways were sought which would interfere with the binding of TNF to its receptors. Thus neutralizing antibodies to TNF were raised (EP 186 833). Another approach to the inhibition of the action of TNF was the provision of soluble TNF receptors which compete with the cell surface TNF-Rs for the binding of TNF (EP 308 378 and EP 398 327).
Since binding to its receptors is required for TNF in order to exert its action, if less or no cell surface receptors are expressed, it should be possible to decrease or inhibit the deleterious effects of TNF. By the same token, it may be desired in certain cases to augment the beneficial action of TNF and in such a case this could possibly be achieved by increasing the amount of cell surface receptors expressed.
SUMMARY OF THE INVENTION
The present invention provides a promoter sequence of the human p75 TNF-R gene selected from a sequence which is located upstream of the 5' end of the gene and a sequence which is located in the first intron of the gene. The 5' upstream promoter sequence (called the 5' region promoter sequence) is contained within a 2.1 kbp sequence, and the intron promoter sequence is contained within a 1.9 kbp sequence. The 5' region promoter sequence is capable of controlling the expression of the native p75 TNF-R gene product, while the intron region promoter sequence is capable of controlling the expression of a truncated p75 TNF-R gene product. Further, the intron region promoter may also function as an enhancer element for the transcription of the p75 TNF-R gene. Moreover, the intron region promoter has upstream to the intron promoter a transcription inhibitory region, which may serve as a modulator for the expression of the truncated p75 TNF-R gene product.
The invention in preferred embodiments provides the 2.18 kbp NcoI fragment, encoding the 5' region promoter sequence, and the 1.8 kbp SmaI fragment, encoding the intron promoter sequence, both fragments being derived from a genomic library containing the human p75 TNF-R. Other preferred embodiments provided by the invention are a portion of the 5' region promoter sequence containing the essential promoter activity and extending between at least nucleotides 1335 and 152
REFERENCES:
Orkin et al, Report and Recommendations of the Panel to Assess the NIH Investment in Research On Gene Therapy, NIH, published Dec. 7, 1995, various unnumbered pages.
Kadonaga, James T. et al., "Affinity purification of sequence-specific DNA binding proteins.", Proc. Natl. Acad. Sci., vol. 83, pp. 5889-5893 (Aug. 1986).
Bielinska, Anna et al., "Regulation of gene expression with double-stranded phosphorotioate oligonucleotides.", Science, vol. 250, pp. 997-1000 (Nov. 16, 1990).
Mitchell, Pamela J. et al., "Transcriptional regulation in mammalian cells by sequence-specific DNA binding proteins.", Science, vol. 245, pp. 371-378 (Jul. 28, 1989).
Singh, Harinder et al., "Molecular cloning of sequence-specific DNA binding proteins using recognition site probes.", Biotechniques, vol. 7, No. 3, pp. 252-261 (1989).
Kemper, Oliver et al., "Cloning and partial characterization of the promoter for the human p55 tumor necrosis factor (TNF) receptor.", Gene, vol. 134, pp. 209-216 (1993).
Rothe, Joachim et al., "Genomic organization and promoter function of the murine tumor necrosis factor receptor B gene.", Molecular Immunology, vol. 30, No. 2, pp. 165-175 (1993).
Smith, Craig A. et al., "A receptor for tumor necrosis factor defines an unusual family of cellular and viral proteins.", Science, vol. 248, pp. 1019-1023 (May 25, 1990).
Kohno et al., "A second tumor necrosis factor receptor gene product can shed a naturally occurring tumor necrosis factor inhibitor.", Proc. Natl. Acad. Sci., vol. 87, pp. 8331-8335 (Nov. 1990).
Kuhnert et al (1994). "Cloning, sequencing and partial functional characterization of the 5' region of the human p75 tumor necrosis factor receptor-encoding gene (TNF-R)". Gene, vol. 150, pp. 381-386.
Ehrhardt Gotz
Kemper Oliver
Kuhnert Peter
Wallach David
McKelvey Terry
Yeda Research and Development Co. Ltd.
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