Interleukin-1 muteins useful as vaccine adjuvants

Details Interleukin-1 muteins useful as vaccine adjuvants Interleukin-1 muteins useful as vaccine adjuvants

2000-12-04

2003-12-02

Kunz, Gary (Department: 1647)

Drug, bio-affecting and body treating compositions

Lymphokine

Interleukin

C424S085100, C530S351000, C514S002600, C514S008100, C514S021800, C514S885000, C435S069500, C435S069520

Reexamination Certificate

active

06656462

ABSTRACT:

BACKGROUND OF THE INVENTION
The immune system is regulated in part by a complex network of chemical signals. These signals include the interleukins such as IL-1&agr; and IL-1&bgr;. IL-1&bgr; is a polypeptide hormone synthesized and secreted by stimulated monocytes. The initial translation product of IL-1&bgr; is a 31 kDa precursor polypeptide having relatively low biological activity. After synthesis, the 31 kDa precursor for IL-1&bgr; is enzymatically cleaved to its highly active mature form which has a size of about 17.5 kDa. The N-terminus of mature IL-1&bgr; derived from human activated monocytes has been characterized by an N-terminal amino acid sequence beginning with Ala-Pro. The N-terminal Ala residue of human mature IL-1&bgr; is in the 117 position and an Asp residue is in the 116 position counting from the N-terminus of human precursor IL-1 &bgr; polypeptide. Mature IL-1&bgr; consists of the C-terminal 153 residues of the precursor polypeptide.
Many physiological actions and biological activities of IL-1&bgr; have been identified. IL-1&bgr; biological activity is often determined by assaying for stimulation of thymocyte proliferation. IL-1&bgr; activities also include stimulation of B-lymphocyte maturation, lymphocyte proliferation, stimulation of fibroblast growth and induction of acute-phase protein synthesis by hepatocytes.
Other biological activities have been attributed to IL-1&bgr; polypeptides. These include control of differentiation and activation of lymphocytes, stimulation of lymphokine and prostaglandin production, promotion of inflammation, induction of acute phase proteins, stimulation of bone resorption, and alteration of the level of iron and zinc in blood. Moreover, it has been found that IL-1&bgr; can stimulate the hypothalamus-pituitary-adrenal axis, suggesting that IL-1&bgr; is integrated in the complex neuroendocrine network that controls homeostasis.
Maturation and release of mature IL-1&bgr; from macrophages does not proceed by conventional means normally associated with most secretory proteins because the precursor IL-1&bgr; polypeptide lacks a hydrophobic signal sequence. Further, IL-1&bgr; is not associated with a membrane-bound compartment in monocytes. Most secretory proteins are characterized by the presence of a hydrophobic stretch of amino acids called a signal sequence. The signal sequence directs the translocation of the protein across the membrane of the endoplasmic reticulum during protein synthesis. The protein is subsequently ushered out of the cell via exocytosis. Most secreted proteins have a signal sequence at the amino terminal that is removed upon translocation. Other proteins, such as ovalbumin, have an internal signal sequence that is not removed upon translocation. The precursor form of IL-1&bgr; lacks any region (either amino terminal or internal) with sufficient hydrophobicity and length to qualify as a signal sequence.
It is very likely that the biological activity of IL-1&bgr; is tightly linked to the structural integrity of the protein molecule, for deletion of amino acids from the mature protein is accompanied by severe diminution of bioactivity. Several groups have introduced point mutations in an attempt to probe receptor ligand interactions (Jobling et al., 1988; Dechiara, T. M. et al. [1986
] Proc. Natl. Acad. Sci. USA
83:8303-8307; Mosley, B., S. K. Dower, S. Gillis, D. Cosman [1987
] Proc. Natl. Acad Sci. USA
84:4572-4576). Huang et al. (Huang, J. J. et al. [1987
] FEBS Letters
223:294-298) reported that the biological activity of IL-1 &bgr; was increased four- to seven-fold by changing the native NH
2
-terminal sequence from ala-pro-val-arg-serto
thr-met
-val-arg-ser; however, further alteration of arginine
120
to generate thr-met-val-
glu
-ser effectively abolished bioactivity. Circular dichroism data demonstrated no major structural differences among the proteins. Gronenborn et al. (Gronenborn, A. M., P. T. Wingfield, H. F. McDonald, U. Schmeissner, G.M. Clore [1988
] FEBS Letters
231:135-138) mutated IL-1-alpha histidine and tryptophan residues without effect upon receptor binding affinity, while MacDonald et al. (MacDonald, H. R. et al. [1986
] FEBS Letters
209:295-298) reported IL-1 histidine muteins with 2-100 fold less competitive binding activity than the wild-type protein.
Excessive or unregulated IL-1 has been implicated in various diseases. These include rheumatoid arthritis (see, e.g., Fontana et al. [1982
] Arthritis Rheum.
22:49-53); osteoarthritis (see, e.g., Wood et al. [1983
] Arthritis Rheum.
26:975); toxic shock syndrome (see, e.g., Ikejima and Dinarello [1985
] J. Leukocyte Biology
37:714); other acute or chronic inflammatory disease states such as the inflammatory reaction induced by endotoxin (see, e.g., Habicht and Beck [1985
] J. Leukocyte Biology
37:709); and other chronic inflammatory disease states such as tuberculosis (see, e.g., Chesque et al. [1985] J. Leukocyte Biology 37:690). Benjamin et al. ([1985] “Annual Reports in Medicinal Chemistry—20,” Chapter 18, pp. 173-183, Academic Press, Inc.) disclose that excessive IL-1 production is implicated in psoriatic arthritis, Reiter's syndrome, rheumatoid arthritis, osteoarthritis, gout, traumatic arthritis, rubella arthritis, and acute synovitis.
Dinarello ([1985
] J. Clinical Immunol.
5(5):287-297)reviews the biological activities which have been attributed to IL-1.
Another form of IL-1, the naturally-occurring receptor antagonist protein (IL-1ra) (Carter, D. B., M. R. Deibel, C. J. Dunn et aL [1990
] Nature
344:633-638; Hannum, C. H., C. J. Wilcox, W. P. Arend et al. [1990
] Nature
343:336-340; Seckinger, P. K. Williamson, J.-F. Balavoine et al. [1987
] J. Immunol.
139:1541-1545) may play an important role in modulating the effects of IL-1. Interleukin-1 biological activity is initiated by interaction with either the type I or type II cellular IL-1 receptors (Dower, S. K., S. R. Kronheim, C. J. March et al. [1984
] J. Exp. Med.
162:501; Sims, J. E., S. K. Dower [1990
] Year in Immunology
6:112-126; Sims, J. E., C. J. March, D. Cosman et al. [1988
] Science
241:585-588; Spriggs, M. K., P. J. Lioubin, J. Slack et al. [1990
] J. Biol. Chem.
265:22499-22505), each of which possesses three immunoglobulin-fold extracellular ligand-binding domains. The IL-1ra also binds to IL-1 receptors, but this protein has not been reported to elicit biological responses (Dripps, D. J., B. J. Brandhuber, R. C. Thompson, S. P. Eisenberg [1991
] J. Biol. Chem.
266:10331-10336). The three-dimensional structures of both IL-1&agr; and IL-1&bgr; have been reported (Clore, G. M., P. T. Wingfield, A. M. Gronenbom [1991
] Biochemistry
30:2315-2323; Driscoll, P. C., G. M. Clore, D. Marion et al. [1990
] Biochemistry
29:3542-3556; Finzel et al., supra; Graves et al. [1990],supra; Priestle et al. [1988], supra; Priestle, J. P., Schar, H. P., M. G. Grutter [1990
] Cytokines and Lipocortins
349:297-307) and both receptor types have been molecularly cloned and expressed (McMahan, C. J., J. L. Slack, B. Mosely et al. [1991
] EMBO J.
10:2821; Sims et al. [1988], supra; Spriggs et al., supra).
By changing the R
127
in &bgr;-strand 1 of the native IL-1&bgr; to a glycine, it has been determined that the receptor binding and biological activity domains of the protein are at least partially distinct. Others have subsequently reported similar effects resulting from mutating other amino acids located in the hydrogen-bonded antiparallel &bgr;-strand 1 and 12 of IL-1&bgr; (Ju, G., E. Labriolatompkins, C. A. Campen et al. [1991
] Proc. Natl. Acad. Sci. USA
88:2658-2662; Young, P., V. Kumar, J. Lillquist et al. [1990
] Lymphokine Res.
9:599). Ju et al., supra, also showed that the IL-1ra protein can be converted to a partial agonist by mutating a residue in a similar

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