Chemistry: molecular biology and microbiology – Animal cell – per se ; composition thereof; process of... – Method of regulating cell metabolism or physiology
Reexamination Certificate
2001-05-07
2002-09-10
LeGuyader, John L. (Department: 1635)
Chemistry: molecular biology and microbiology
Animal cell, per se ; composition thereof; process of...
Method of regulating cell metabolism or physiology
C435S006120, C435S091100, C435S325000, C435S366000, C536S023100, C536S024310, C536S024330, C536S024500
Reexamination Certificate
active
06448081
ABSTRACT:
FIELD OF THE INVENTION
The present invention provides compositions and methods for modulating the expression of Interleukin 12 p40 subunit. In particular, this invention relates to compounds, particularly oligonucleotides, specifically hybridizable with nucleic acids encoding Interleukin 12 p40 subunit. Such compounds have been shown to modulate the expression of Interleukin 12 p40 subunit.
BACKGROUND OF THE INVENTION
Cytokines are soluble factors produced by lymphocytes that regulate the survival, proliferation, differentiation, and homeostasis of cells involved in mediating the immune response. These factors do not only activate other lymphocytes, they also relay signals to non-lymphoid cells including macrophages, epithelial and stromal cells, creating a broad spectrum of cytokine activity that is critical to the maintenance of health. Consequently, much effort has been devoted to the study of these proteins.
Interleukin 12 (IL-12; formerly NKSF, for natural killer cell stimulatory factor, or CLMF, for cytotoxic lymphocyte maturation factor) is a cytokine produced by monocytes, macrophages, neutrophils, dendritic cells and antibody-producing B cells (Hall,
Science
, 1995, 268, 1432-1434) as well as keratinocytes and epidermoid carcinoma cell lines (Aragane et al.,
J. Immunol
., 1994, 153, 5366-5372).
Interleukin 12 is responsible for activation of natural killer (NK) cells, T cells and induction of increased levels of interferon-gamma, a cytokine that helps to shape the immune response (Hall,
Science
, 1995, 268, 1432-1434). The combination of interferon-gamma and interleukin 12 sends a powerful signal to native precursor cells of the T helper lineage, shifting the immune system to a T
H
1-type immune response (Hall,
Science
, 1995, 268, 1432-1434). Generally, resistance to pathogens increases when interleukin 12 is present to drive a T
H
1 response (Hall,
Science
, 1995, 268, 1432-1434).
Interleukin 12 is unique among the cytokines because it is a disulfide-linked heterodimer composed of unrelated 40-kD (p40) and 35-kD (p35) subunits that are encoded by genes on separate chromosomes. The p35 and p40 subunits are localized to chromosomes 3p12-3q13.2 and 5q31-q33 respectively (Sieburth et al.,
Genomics
, 1992, 14, 59-62). The p35 subunit shares structural similarities with the cytokines interleukin 6 and granulocyte colony-stimulating factor (Merberg et al.,
Immunol. Today
, 1992, 13, 77-78). Alternatively, the p40 subunit is structurally related to the interleukin 6 receptor (Gearing and Cosman,
Cell
, 1991, 66, 9-10).
Disclosed and claimed in U.S. Pat. No. 5,457,038 and PCT publication WO 92/05206 is a DNA sequence coding for interleukin 12 or a subunit thereof (Trinchieri et al., 1995; Trinchieri et al., 1992).
cDNAs for both subunits of interleukin 12 were cloned in 1991 from a lymphoblastoid B-cell line (Gubler et al.,
Proc. Natl. Acad. Sci. U S A
, 1991, 88, 4143-4147). Both subunits are required to obtain the biologically active heterodimer (Gubler et al.,
Proc. Natl. Acad. Sci. U S A
, 1991, 88, 4143-4147) and p35 is only secreted as part of the heterodimer (D'Andrea et al.,
J. Exp. Med
., 1992, 176, 1387-1398) whereas p40 can be induced and secreted independently (Snijders et al.,
J. Immunol
., 1996, 156, 1207-1212) and has no biological activity (Ling et al.,
J. Immunol
., 1995, 154, 116-127). Bioactive murine and human single chain interleukin 12 fusion proteins expressed from retroviral constructs have been demonstrated to retain antitumor activity in vivo (Lieschke et al.,
Nat. Biotechnol
., 1997, 15, 35-40).
Interleukin 12 has been found to be upregulated in vivo during murine lipopolysaccharide-induced endotoxemia and to stimulate the synthesis of interferon-gamma (Heinzel et al.,
Infect. Immun
., 1994, 62, 4244-4249). However, pretreatment of the mice with anti-interleukin 12 antibodies caused a reduction in interferon-gamma levels after lipopolysaccharide injection (Heinzel et al.,
Infect. Immun
., 1994, 62, 4244-4249).
Astrocyte-targeted expression of both interleukin 12 p35 and p40 genes in mice promoted the spontaneous development of a severe neuroimmunological disorder with many features resembling those of experimental allergic encephalomyelitis (EAE) (Pagenstecher et al.,
J. Immunol
., 2000, 164, 4481-4492).
Studies of interleukin 12 production and mRNA expression in mice exposed to chemical allergens indicated that the divergent T-cell cytokine responses resulting from contact and respiratory allergens are associated with differential production of the p40 protein in the absence of substantial changes in the expression of mRNA for either subunit (Warbrick et al.,
Toxicology
, 1999, 132, 57-66).
Interleukin 12 p35 and p40 antisense probes were used in in situ hybridization studies that identified enhanced interleukin 12 transcription in the gastric mucosa of pediatric patients with Crohn's disease (Berrebi et al.,
Am. J. Pathol
., 1998, 152, 667-672).
Upregulation of the interleukin 12 p40 subunit is associated with lung fibrosis induced by silica particles in mouse models (Huaux et al.,
Am. J. Respir. Cell Mol. Biol
., 1999, 20, 561-572).
The homodimer of the interleukin 12 p40 subunit (also known as (p40)
2
) has been found to be a very potent inhibitor of interleukin 12 activity (Gillessen et al.,
Eur. J. Immunol
., 1995, 25, 200-206; Ling et al.,
J. Immunol
., 1995, 154, 116-127) and functions by binding to the interleukin 12 receptor (Ling et al.,
J. Immunol
., 1995, 154, 116-127). The p40 homodimer was used as an antagonist of interleukin 12 in investigative treatments of mice with cyclophosphamide-induced diabetes and found to dampen islet formation (Rothe et al.,
Diabetologia
, 1997, 40, 641-646). Additionally, the p40 homodimer was found to inhibit the antitumor activity of the interleukin 12 heterodimer and the induction of interferon-gamma in murine bladder carcinoma in vivo. (Chen et al.,
J. Immunol
., 1997, 159, 351-359).
Carter et al. have produced antibodies to recombinant human interleukin 12 which have been demonstrated to neutralize the biological activity of interleukin 12. However, essentially all antibodies were generated to the p40 subunit, possibly due to conformational limitations of the intact interleukin 12 heterodimer (Carter et al.,
Hybridoma
, 1997, 16, 363-369). Larsson et al. have reported an immunoassay that only recognized the bioactive human interleukin 12 heterodimer and not the individual p35 and p40 subunits (Larsson and Linden,
Cytokine
, 1998, 10, 786-789). Disclosed and claimed in PCT publication WO 99/37682 are anti-human interleukin 12 antibodies that are characterized by specificity to the interleukin 12 heterodimer but do not bind to the interleukin 12 p40 subunit (Gately and Presky, 1999).
In investigations of the mechanisms of anti-inflammatory effects of corticosteriods, budesonide was found to inhibit production of bioactive interleukin 12 in human monocytes (Larsson and Linden,
Cytokine
, 1998, 10, 786-789).
The involvement of interleukin 12 p40 subunit in immune system regulation as well as viral and bacterial infections make it a potentially useful therapeutic target for intervention in autoimmune diseases. Currently, inhibitors of the interleukin 12 p40 subunit and/or the interleukin 12 heterodimer include the p40 homodimer, antibodies and corticosteroids. Consequently, there remains a long felt need for additional agents capable of effectively and selectively inhibiting the function of the interleukin 12 p40 subunit.
Antisense technology is emerging as an effective means for reducing the expression of specific gene products and may therefore prove to be uniquely useful in a number of therapeutic, diagnostic, and research applications for the modulation of expression of the interleukin 12 p40 subunit.
The present invention provides compositions and methods for modulating expression of the interleukin 12 p40 subunit.
SUMMARY OF THE INVENTION
The present invention is directed to compounds, particularly antisense oligonucleotides, which are targeted to a nucleic aci
Baker Brenda F.
Freier Susan M.
ISIS Pharmaceuticals Inc.
LeGuyader John L.
Licata & Tyrrell P.C.
Schmidt M
LandOfFree
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