Patent EA-003900-B1 (EA)
Title: Truncated tumor necrosis factor receptors, pharmaceutical composition on their base and methods os use of said factors and compositions
Inventor: FISHER ERIC F
IPC: A61K47/48
Filed: 1997-07-09
Abstract: 1. A truncated sTNFR having the following formula: R1- [Cys<19>-Cys<103>] -R2 wherein [Cys<19>-Cys<103>] represents residues 19 through 103 of sTNFR-I, the amino acid residue numbering scheme of which is provided in Figure 1 (SEQ ID NO:2) to facilitate the comparison; wherein R1 represents a methionylated or nonmethionylated amine group of Cys<19> or of aminoterminus amino acid residue(s) selected from the group: and wherein R2 represents a carboxy group of Cys<103> or of carboxy-terminal amino acid residues, selected from the group: and variants and derivatives thereof, provided however, when R1 represents a methionylated or nonmethionylated amine group of amino acid sequence VCPQGKYIHPQNNSIC or an N-terminal truncation thereof of from 1 to 15 residues, then R1-[Cys<19>-Cys<103>]-R2 is not an addition variant having the formula R1-[Cys<19>-Cys<103>]-FCCSLCL-R3, wherein R3 represents a carboxyl group of amino acid residues Asn<111>-Asn<161> of Figure 1 or a carboxy-terminal truncation of Asn<111>-Asn<161> of Figure 1. 2. The tumor necrosis binding protein according to Claim 1, selected from the group consisting of 3. A truncated sTNFR having the following formula: R4-[Cys<32>-Cys<115>]-R5 wherein [Cys<32>-Cys<115>] represents amino acid residues Cys<32> through Cys<115> of sequence SEQ ID NO:35 provided in Figure 8; wherein R4 represents a methionylated or nonmethionylated amine group of Cys<32> or of aminoterminus amino acid residue(s) selected from the group, comprising and wherein R5 represents a carboxy group of Cys<115> or of carboxy-terminal amino acid residues selected from the group, comprising and variants thereof, provided however, when R4 represents a methionylated or non-methionylated amine group of amino acid sequence TCRLREYYDQTAQMC or an N-terminal truncation thereof of from 1 to 15 residues, then R4-[Cys<32>-Cys<115>]-R5 is not an addition variant having the formula R4-[Cys<32>-Cys<115>]-APLRKCR-R6, wherein R6 represents a carboxyl group of amino acid residues Pro<123>-Thr<179> of Figure 8 or a carboxy-terminal truncation of Pro<123>-Thr<179> of Figure 8. 4. The truncated sTNFR according to any one of Claims 1 - 3, wherein it has non-glycosylated amino-acid sequence. 5. The truncated sTNFR according to any one of Claims 1 - 3, wherein it has glycosylated amino acid sequence. 6 The truncated sTNFR according to any one of Claims 1 - 3, wherein it is conjugated to a water-soluble polymer. 7. The truncated sTNFR according to Claim 6, wherein the water-soluble polymer is polyethylene glycol. 8. A polyvalent sTNFR comprising at least one of receptors according to any one of Claims 1 though 7. 9. A polyvalent sTNFR according to claim 8, wherein it contains a dimmer of truncated receptor of claim 3. 10. A polyvalent tumor necrosis binding protein having the formula R1-X-R2, wherein: X comprises a linker, wherein said linker is a water soluble polymer; and R1 and R2 are biologically-active molecules covalently bonded to said water soluble polymer, wherein at least one of R1 and R2 is a tumor necrosis binding protein according to any one of Claims 1 - 5. 11. The polyvalent receptor of Claim 10, wherein the water-soluble polymer is polyethylene glycol. 12. The polyvalent receptor of Claim 11, wherein the truncated receptor is selected from the group consisting of sTNFR-I 2.6D/C105db and sTNFR-I 2.6D/C106db. 13. A polynucleotide encoding the sTNFR according to any one of Claims 1-3, selected from group comprising: polynucleotide with sequence as shown in Fig. 2 (SEQ ID NO: 3); polynucleotide with sequence as shown in Fig. 3 (SEQ ID NO: 5); polynucleotide with sequence as shown in Fig. 4 (SEQ ID NO: 7); polynucleotide with sequence as shown in Fig. 5 (SEQ ID NO: 11); polynucleotide with sequence as shown in Fig. 6 (SEQ ID NO: 9); polynucleotide with sequence as shown in Fig. 7 (SEQ ID NO: 13); polynucleotide with sequence which is degenerated in the coding regions or portions thereof of said sequences. 14. A vector comprising a polynucleotide of any one of Claims 13 through 15 operatively linked to an expression control sequence. 15. A prokaryotic host cell containing a polynucleotide of Claim 13 or a vector of Claim 14. 16. A eukaryotic host cell containing a polynucleotide of Claim 13 or a vector of Claim 14. 17. A method of preparation of truncated sTNFR providing the growing host cells of Claims 15 or 16 in conditions, providing the expression of said truncated sTNFR by host cells, and isolation of it. 18. The method according to Claim 17, wherein said host cells are E. coli. 19. The method according to Claim 17, wherein said host cells are Chinese hamster ovary cells. 20. Truncated sTNFR according to any of claims 1-7, characterized in that it is prepared by recombinant method. 21. A method of preparing of pharmaceutical composition wherein a therapeutically effective amount of the tumor necrosis factor receptor according to any one of Claims 1 -7 or Claim 20 or a polyvalent tumor necrosis factor receptor according to Claims 8-12 is mixed with one or more pharmaceutically acceptable vehicles. 22. A pharmaceutical composition comprising the tumor necrosis factor receptor according to any of Claims 1-7 or 20 in association with a pharmaceutically acceptable vehicle. 23. A pharmaceutical composition comprising the polyvalent tumor necrosis factor receptor according to any of Claims 8-12 in association with a pharmaceutically acceptable vehicle. 24. A pharmaceutical composition according to Claims 22 or 23, wherein it is continuous release composition. 25. A pharmaceutical composition according to Claims 22 or 23, wherein it is lyophilized. 26. A method of treating a TNF-mediated disease comprising administering to a patient the truncated sTNFR according to any of Claims 1-7 or 20 or polyvalent receptor according to any of Claims 8-12. 27. The method of claim 26, wherein the TNF-mediated disease is diabetes, hyperalgesia, inflammatory disease of bowels, ischemic and reperfusion injuries and rheumatic disease. 28. The method of claim 27, wherein a rheumatic disease is selected from group comprising arthritis, osteoarthritis, juvenile arthritis, ankylosing spondylitis, dermatomyositis, psoriatic arthritis, sclerodermatitis, Sjogren's sicca syndrome and vasculitis. 29. The method according to any of Claims 26-28, wherein the truncated or polyvalent sTNFR are administered intravenously, intramuscularly, intracutaneously, subcultaneously, intraarticular, or by injection. 30. The method according to any of Claims 26-29, wherein the truncated or polyvalent sTNFR are administered before, in combination and after administering of anti-inflammatory treatment. 31. The method according to Claim 30, wherein an anti-inflammatory treatment is selected from the group, comprising non-steroid anti-inflammatory treatments corticosteroids, slow antirheumatic preparations, and preparations modifying the disease. 32. The method according to Claim 30, wherein an anti-inflammatory treatment is interleikine-1 inhibitor (IL-1), selected from antagonist receptor of interleikine-1 (IL-ra) or soluble IL-1 receptor. 33. Use of truncated sTNFR according to Claims 1-7 or 20 or polyvalent receptor of Claims 8-12 for preparing of preparation for treating TNF-mediated disease. 34. Use of truncated sTNFR according to Claims 1-7 or 20 or polyvalent receptor of Claims 8-12 for preparing of preparation for treating diabetes, hyperalgesia, inflammatory disease of bowels, ischemic and reperfusion injuries and rheumatic disease. 35. Use of Claim 34, characterized in that rheumatic disease is selected from group comprising arthritis, osteoarthritis, juvenile arthritis, ankylosing spondylitis, dermatomyositis, psoriatic arthritis, sclerodermatitis, Sjogren's sicca syndrome and vasculitis. 36. The method according to any of Claims 33-35, wherein the said preparation is administered intravenously, intramuscularly, intracutaneously, subcultaneously, intraarticular, or by injection. 37. The method according to any of Claims 33-36, wherein the said preparation sTNFR is administered before, in combination and after administering of anti-inflammatory treatment. 38. The use according to Claim 37, wherein an anti-inflammatory treatment is selected from the group, comprising non-steroid anti-inflammatory treatments corticosteroids, slow antirheumatic preparations, and preparations modifying the disease. 39. The use according to Claim 37, wherein an anti-inflammatory treatment is interleikine-1 inhibitor (IL-1), selected from antagonist receptor of interleikine-1 (IL-ra) or soluble IL-1 receptor. 40. A kit for preparing an aqueous protein formulation comprising a first container with tumor necrosis factor receptor according to any one of Claims 1-7 and a second container having a physiologically acceptable solvent. 41. The method according to Claim 31, wherein an anti-inflammatory treatment is methotrexate. 42. Use according to Claim 38, wherein an anti-inflammatory treatment is methotrexate. 43. Pharmaceutical composition, according to any of Claims 22-25, wherein it further comprises an anti-inflammatory treatment. 44. Pharmaceutical composition according to Claim 44, wherein an anti-inflammatory treatment is selected from the group, comprising non-steroid anti-inflammatory treatments corticosteroids, slow antirheumatic preparations, and preparations modifying the disease. 45. Pharmaceutical composition according to Claim 44, wherein an anti-inflammatory treatment is methotrexate. 46. Pharmaceutical composition according to Claim 43, wherein an anti-inflammatory treatment is interleikine-1 inhibitor (IL-1), selected from antagonist receptor of interleikine-1 (IL-ra) or soluble IL-1 receptor. 47. Pharmaceutical composition according to Claim 46, wherein the antagonist receptor of interleikine-1 (IL-ra) comprises a sequence of human protein IL-1ra.