Nucleic acid encoding monocyte chemotactic protein 4

Details Nucleic acid encoding monocyte chemotactic protein 4 Nucleic acid encoding monocyte chemotactic protein 4

2000-04-07

2004-01-06

Mertz, Prema (Department: 1646)

Organic compounds -- part of the class 532-570 series

Organic compounds

Carbohydrates or derivatives

C435S071100, C435S071200, C435S320100, C435S325000, C435S252300, C435S254110, C435S471000, C530S324000

Reexamination Certificate

active

06673915

ABSTRACT:

BACKGROUND OF THE INVENTION
The invention relates to novel chemoattractants.
Six different types of white blood cells (leukocytes) are typically found in the blood. These are neutrophils, eosinophils, basophils, monocytes, lymphocytes, and plasma cells. The neutrophils and monocytes are primarily responsible for attacking and destroying invading bacteria, viruses, and other harmful agents. Neutrophils circulate within the bloodstream as mature, functional cells. Monocytes, however, circulate as immature cells that have a limited ability to fight infectious agents. It is only when monocytes are stimulated by chemotactic agents to move through the capillary wall into surrounding tissue that they become fully active. Once monocytes enter the tissues they begin to swell and many lysosomes and mitochondria appear in their cytoplasm. At this point, monocytes become called macrophages, which are extremely effective phagocytes. Each macrophage can engulf as many as 100 bacterial cells, as well as large particles, including whole red blood cells, malarial parasites, and necrotic tissue.
Chemokines are chemotactic cytokines that contribute to various immune and inflammatory responses by regulating the movement of selected blood-borne leukocytes into the tissues (Baggiolini et al., Adv. Immunol. 55:97-179, 1994; Oppenheim et al., Ann. Rev. Immunol. 9:617-648, 1991). All known chemokines have been assigned to one of three families on the basis of the chromosomal location of the genes that encode them and on the motif formed by conserved cysteine residues in the mature proteins. Two of these families, designated &agr; and &bgr;, have many members, all of which have four conserved cysteine residues. The first two cysteines of &agr; chemokines are separated by a single amino acid (CXC motif), and these proteins are encoded by genes clustered on human chromosome 4. In contrast, the first two cysteines of all &bgr; family chemokines are adjacent to one another (CC motif), and these proteins are encoded by genes clustered on human chromosome 17.
Chemokines having the CXC motif (&agr; chemokines) primarily affect neutrophils and lymphocytes, and they can modulate angiogenesis. Chemokines having the CC motif (&bgr; chemokines) affect monocytes, lymphocytes, eosinophils, and basophils with variable selectivity. In addition, certain chemokines in the &agr; and &bgr; family can inhibit HIV replication in T cells and monocytes, respectively.
The third chemokine family currently has only one member: the T cell-specific chemoattractant, lymphotactin (Kelner et al., Science 266:1395-1399, 1994). The gene encoding lymphotactin is located on human chromosome 1, and the protein sequence contains only the second and fourth cysteines found in the &agr; and &bgr; chemokine families (Kennedy et al., J. Immunol. 155:203-209, 1995).
The monocyte chemoattractant proteins (MCPs), which constitute a subfamily of the &bgr; chemokines described above, include the three human MCP proteins (MCP-1, MCP-2, and MCP-3). These proteins are ~65% identical, and all have Gln as their N-terminal amino acid (Van Damme et al., J. Exp. Med. 176:59-65, 1992; Yoshimura et al., J. Exp. Med. 169:1449-1459, 1989).
The chemokine eotaxin is also a &bgr; chemokine. It is related in sequence to the MCP proteins, but does not contain an amino-terminal Gln residue. Eotaxin appears to be unique among the chemokines in that it causes the selective infiltration of eosinophils when injected subcutaneously and when administered directly to the lungs of naive guinea pigs.
Five genes encoding human &bgr; chemokine receptors, referred to as CKR1 through CKR5, have been cloned. (Charo et al., Proc. Natl. Acad. Sci. USA 91:2752-2756, 1994; Combadiere et al., DNA and Cell Biol. 14:673-380, 1995; Combadiere et al., J. Leukocyte Biol. in press, 1996; Gao et al., J. Exp. Med. 177:1421-1427, 1993; Neote et al., Cell 72:415-425, 1993; Power et al., J. Biol. Chem. 270:19495-19500, 1995; Samson et al., Biochem. 35:3362-3367, 1996). CKR2a and CKR2b are splice variants of the same gene. These receptors are not ubiquitously expressed: CKR1, CKR4, and CKR5 appear to be widely expressed on leukocytes, while expression of CKR2 and CKR3 is mainly restricted to monocytes and eosinophils, respectively.
SUMMARY OF THE INVENTION
The invention features substantially pure nucleic acid molecules that encode MCP-4 and MCP-5, two novel members of the &bgr;-chemokine family. The invention also includes polypeptides encoded by these nucleic acid molecules.
The nucleic acid molecules may consist of genomic DNA, cDNA, or mRNA, and, due to the degenerate nature of the genetic code, may vary in sequence provided that the encoded polypeptides are MCP-4 or MCP-5, as shown, for example, in
FIGS. 1A and 1B
, respectively.
Substantially pure MCP-4 and MCP-5 polypeptides, biologically active fragments of these polypeptides, including immunogenic fragments, are also considered within the scope of the invention. The biological activity of any given fragment of MCP-4 or MCP-5 may be readily determined by conducting an assay of, e.g., monocyte chemotaxis, as described herein. The preferred polypeptides of the invention are both substantially homologous to MCP-4 or MCP-5 and retain the biological activity of the relevant polypeptide, as described herein.
The nucleic acid molecules can encode a mammalian MCP-4 or MCP-5 polypeptide, such as those from a human, mouse, rat, guinea pig, cow, sheep, horse, pig, rabbit, monkey, dog, or cat. Preferably, the nucleic acid molecule encoding MCP-4 encodes human MCP-4 and the nucleic acid molecule encoding MCP-5 encodes murine MCP-5 or, more preferably, human MCP-5.
The nucleic acid molecules can be placed under the control of a promoter, which may be constitutively active or induced by one or more external agents. The promoter can provide the means to achieve tissue-specific or cell type-specific expression of the nucleic acid molecules of the invention. Alternatively, or in addition, the nucleic acid molecules can be operably linked to a DNA regulatory sequence. Skilled artisans will recognize that the nucleic acid molecules can be placed into a vector construct, such as a plasmid or viral vector, which may in turn be used to transduce living cells with the nucleic acid molecules of the invention. Cells can be transfected with plasmid vectors by standard methods including, but not limited to, liposome-, polybrene-, or DEAE dextran-mediated transfection (see, e.g., Feigner et al., Proc. Natl. Acad. Sci. USA 84:7413, 1987; Ono et al., Neurosci. Lett. 117:259, 1990; Brigham et al., Am. J. Med. Sci. 298:278, 1989), electroporation (Neumann et al., EMBO J. 7:841, 1980), calcium phosphate precipitation (Graham et al., Virology 52:456, 1973; Wigler et al., Cell 14:725, 1978; Felgner et al., supra) microinjection (Wolff et al., Science 247:1465, 1990), or velocity driven microprojectiles (“biolistics”). Viruses known to be useful for gene transfer include adenoviruses, adeno associated virus, herpes virus, mumps virus, poliovirus, retroviruses, Sindbis virus, and vaccinia virus such as canary pox virus.
The nucleic acid constructs described above are useful in a variety of ways. For example, the constructs may be used as a source of recombinantly produced MCP-4 or MCP-5 polypeptides. Alternatively, the constructs may be administered themselves in a therapeutic approach, as described below. In addition, the constructs may be used to generate transgenic animals that overexpress, or fail to express, MCP-4 or MCP-5. The animal can be a mouse, a worm, or any other animal considered useful for research or drug development. Transgenesis has become routine in the art of molecular biology.
MCP-4 or MCP-5 polypeptides expressed by transfected cells can be purified and injected into an animal, such as a rabbit, in order to generate polyclonal antibodies that specifically bind MCP-4 or MCP-5. Monoclonal antibodies may also be prepared using standard hybridoma technology (see, e.g., Kohler et al., Nature 256:495, 1975; Kohler et al., Eur. J. Immunol. 6:292 and 6:511, 197

Affiliated with

Also associated with

Rating

Nucleic acid encoding monocyte chemotactic protein 4 does not yet have a rating. At this time, there are no reviews or comments for this patent.

If you have personal experience with Nucleic acid encoding monocyte chemotactic protein 4, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Nucleic acid encoding monocyte chemotactic protein 4 will most certainly appreciate the feedback.

Rate now

Comments { 0 }

Profile ID: LFUS-PAI-O-3243966