Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Reexamination Certificate
1999-11-15
2002-07-23
Stucker, Jeffrey (Department: 1647)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
C435S007920, C435S069100, C435S069500, C530S351000, C530S388200, C530S389200
Reexamination Certificate
active
06423500
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates generally to nucleic acids and polypeptides characteristic of receptors for mammalian interleukin-10, and more particularly to their uses in preparing reagents useful for diagnosing or treating various receptor-related medical conditions.
BACKGROUND OF THE INVENTION
Activated hemopoietic cells secrete numerous proteins. Cytokines are a subset of these proteins and play a variety of important roles in regulation of immune responses by controlling proliferation, differentiation, and the effector functions of immune cells. Most cytokines have more than one biological activity and which activity is the most important likely depends on the local context in which the cytokine is produced.
These cytokines are intercellular signaling molecules whose actions are typically mediated through specific receptor molecules found on target cells. The structure and mechanism of action of these receptors on target cells is not well understood, though many are composed of at least two separate proteins. These earlier described heterodimeric receptors are often composed of polypeptides which are related both to each other and to receptors for other cytokines. Components of receptors for other cytokines have been described. See, e.g., Gearing, et al. (1989)
EMBO J
. 8:3667-3676 (low affinity &agr; chain of a human GM-CSF receptor); Itoh, et al. (1990)
Science
247:324-327 (low affinity &agr; chain of a mouse IL-3 receptor); and Hayashida, et al. (1990)
Proc. Nat'l Acad. Sci. USA
87:9655-9659 (a &bgr; chain of a human GM-CSF receptor); and Tavernier, et al. (1991)
Cell
66:1175-1184 (IL-5 receptor, &agr; and &bgr; chains). The various components of the earlier identified receptors appear to share properties useful in defining a receptor superfamily of related proteins. See, e.g., Bazan (1990)
Immunology Today
11:350-354; and Bazan (1990)
Proc. Nat'l Acad. Sci. USA
87:6934-6938. However, the structure and mechanism of action of a receptor for a mammalian interleukin-10 (IL-10) could not be predicted with reliability based merely upon speculated similarity to receptors for other cytokines.
As soluble intercellular messenger molecules, the cytokines likely bind to cellular receptors, e.g., cell surface receptors. Receptor molecules have been identified and isolated for G-CSF, GM-CSF, EPO, TNF, IFN-&ggr;, IL-2, IL-3, IL-4, IL-5, IL-6, and IL-7. Many of these receptors have two chains, both of which are members of the hemopoietic receptor superfamily. In such cases, typically one chain, designated the &agr; chain, can bind its ligand with low affinity. This interaction may or may not result in transduction to the cell of a signal. Another chain, designated the &bgr; chain, when associated with the &agr; chain, confers higher affinity binding of the heterodimeric receptor to the cytokine. The &bgr; chain by itself usually lacks significant ligand binding affinity. The dimeric form of receptor is capable of transducing a signal into the cell as a consequence of ligand, e.g., cytokine, binding. However, any similarity between the structural and functional features of those receptors generally and a receptor for IL-10 is speculative. Additional subunits may also be associated with the receptors.
A cytokine synthesis inhibitory factor (CSIF) activity led to assays which allowed the isolation of a cytokine designated interleukin-10 (IL-10). See Fiorentino, et al. (1989)
J. Exptl. Med
. 170:2081-2095; and Mosmann, et al. (1991)
Immunol. Today
12:A49-A53. Both mouse and human counterparts have been isolated. See Moore, et al. (1990)
Science
248:1230-1234; and Vieira, et al. (1991)
Proc. Nat'l Acad. Sci. USA
88:1172-1176. A human viral analog, known as either vIL-10 or BCRF1, has been described which shares many characteristic activities of the natural human form. See Hsu, et al. (1990)
Science
250:830-832. Another viral homolog has been described from an equine herpes virus. See Rode, et al. (1993)
Virus Genes
7:111-116.
Human IL-10 (hIL-10) has an N-terminal hydrophobic signal sequence of 18 amino acids, one potential N-linked glycosylation site, 4 cysteine residues, and seven methionine residues. It shares strong DNA and amino acid sequence homology with mouse IL-10 and an open reading frame in Eppstein-Barr virus, BCRF1, and an open reading frame in an equine herpes virus, type II. It inhibits cytokine synthesis by activated T cells, stimulates growth for thymocytes and mast cells, induces class II MHC expression, and sustains viability in culture of small dense resting mouse B cells. A mouse counterpart has also been described, and equivalent proteins would be found in other mammalian species. IL-10 binding to cell surface receptors is thought to be an initiating step for various specific cellular responses, as described below.
One means to modulate IL-10 effect upon binding to its receptor, and therefore potentially useful in treating inappropriate immune responses, e.g., autoimmune, inflammation, sepsis, and cancer situations, is to inhibit the receptor signal transduction. Unfortunately, finding reagents capable of serving as an antagonist or agonist has been severely hampered by the absence of large quantities of IL-10 receptor, preferably purified and in an active form. In order to characterize the structural properties of the IL-10 receptor in greater detail and to understand the mechanism of action at the molecular level, purified receptor will be very useful.
Moreover, similarities to other cytokine functions exist. In particular, the IL-10 receptor likely shares many functions and characteristics with other receptors, but also exhibits different structural and functional properties. The receptors provided herein, by comparison to other receptors or by combining structural components, will provide further understanding of signal transduction induced by ligand binding.
The isolated receptor gene should provide means to generate an economical source of the receptor, allow expression of more receptors on a cell leading to increased assay sensitivity, promote characterization of various receptor subtypes and variants, and allow correlation of activity with receptor structures. Moreover, fragments of the receptor may be useful as agonists or antagonists of ligand binding. See, e.g., Harada, et al. (1992)
J. Biol. Chem
. 267:22752-22758.
Thus, a need exists for the isolation and characterization of nucleic acids encoding components of receptors for IL-10. The present invention provides these and the means for preparing many other useful reagents.
SUMMARY OF THE INVENTION
The present invention provides nucleic acid and protein sequences of components of a receptor for IL-10. Both a human IL-10 receptor component and a mouse counterpart are exemplified, though equivalent components from other mammalian species will be found by similar methods or based upon other properties derived therefrom.
The present invention provides recombinant or isolated nucleic acids comprising a sequence exhibiting homology to a sequence encoding a mammalian receptor for IL-10, a fragment thereof, or a unique portion thereof. In preferred embodiments, the nucleic acids will comprise deoxyribonucleic acid, will be isolated, further comprise a regulatory sequence from the 5′ or 3′ sequence adjacent a gene encoding a receptor for IL-10, or are operably linked to a genetic control element. In alternative embodiments the receptors, fragments, or portions thereof have a biological activity, e.g., one characteristic of a receptor for IL-10, or are from a mammal, including a mouse or human.
In particular embodiments, the nucleic acids, are capable of hybridizing at high stringency to SEQ ID NO: 1 or 3, or are isolated using a probe which hybridizes at high stringency to a human receptor for IL-10. The invention also embraces nucleic acids capable of hybridizing to these sequences which contain mutations selected from the group consisting of nucleotide substitutions, nucleotide deletions, nucleotide insertions, and inversions of nucleotide
Chou Chuan-Chu
Ho Alice Suk-Yue
Hsu Di-Hwei
J. Bazan Fernando
Liu Ying
Ching Edwin P.
Mohan-Peterson Sheela
Schering Corporation
Seharaseyon Jegatheesan
Stucker Jeffrey
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