title: Minimal tumor necrosis factor receptor binding protein: optimum biological activity of a truncated p55 soluble tumor necrosis factor receptor-IgG fusion protein.
abstract: We have previously demonstrated, using expressed deletion constructs, that the fourth membrane proximal cysteine-rich repeat of the p55 TNF receptor (TNF-R) is not required for binding of tumour necrosis factor-alpha (TNF) or lymphotoxin-alpha (LT; tumour necrosis factor-beta). We and others have also shown that the soluble p55 TNF-R, rendered dimeric by fusion to an IgG backbone is extremely effective at neutralizing the harmful effects of TNF overproduction, such as in toxic shock. Here we address the question of how the TNF binding properties of the truncated TNF-R comprising the three distal cysteine-rich repeats (delta4 TNF-R), when fused with an IgG backbone, compare with those of the full length soluble receptor. We constructed several versions of the soluble delta4 TNF-R, on a complete IgG heavy chain backbone and on an IgG lacking the CH1 (first constant region) domain. The constructs were expressed with an Ig or native TNF receptor leader sequence and altered or native N terminal sequence, to compare efficiency of expression. When compared with a full length, soluble receptor Ig fusion protein, the affinity of all for TNF was identical, as were their activities in in vitro binding and cytotoxicity assays. In vivo studies showed that the delta4 and wild type fusion proteins afforded equivalent protection against LPS-induced lethality. However, the delta4 proteins exhibited a significantly lower affinity for LT, and reduced activity in LT binding and cytotoxicity assays. We conclude that the truncated TNF receptor IgG fusion protein is as effective at neutralizing TNF activity as the full length soluble receptor fusion protein. Its lower affinity for LT may make it a more selective agent in blocking the action of TNF, while causing less interference with the action of LT. Also its smaller size may make it a more useful therapeutic agent as it may be less immunogenic than the full length receptor.
authors.affiliation: The Mathilda and Terence Kennedy Institute of Rheumatology, Sunley Division, Hammersmith, London, England.
authors.forename: A E
authors.initials: AE
authors.lastname: Corcoran
authors.forename: B J
authors.initials: BJ
authors.lastname: Scallon
authors.forename: H
authors.initials: H
authors.lastname: Trinh
authors.forename: Y
authors.initials: Y
authors.lastname: Chernajovsky
authors.forename: J
authors.initials: J
authors.lastname: Ghrayeb
authors.forename: M
authors.initials: M
authors.lastname: Feldmann
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D002470:Cell Survival / Q000187:drug effects; D019281:Dimerization; D005786:Gene Expression Regulation; D006801:Humans; D007127:Immunoglobulin Constant Regions; D007074:Immunoglobulin G / Q000737:chemistry / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D008070:Lipopolysaccharides / Q000037:antagonists & inhibitors / Q000633:toxicity; D008233:Lymphotoxin-alpha / Q000633:toxicity; D051379:Mice; D008958:Models, Molecular; D016133:Polymerase Chain Reaction; D018124:Receptors, Tumor Necrosis Factor / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D047888:Receptors, Tumor Necrosis Factor, Type I; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism / Q000627:therapeutic use; D014162:Transfection; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: D015703:Antigens, CD; D007127:Immunoglobulin Constant Regions; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D008070:Lipopolysaccharides; D008233:Lymphotoxin-alpha; D018124:Receptors, Tumor Necrosis Factor; D047888:Receptors, Tumor Necrosis Factor, Type I; D011993:Recombinant Fusion Proteins; D014409:Tumor Necrosis Factor-alpha