Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
2000-08-18
2002-04-30
Horlick, Kenneth R. (Department: 1656)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C435S007100, C435S069100, C435S070100, C435S455000, C436S501000
Reexamination Certificate
active
06379894
ABSTRACT:
BACKGROUND OF THE INVENTION
The present invention relates to a method for identifying compounds capable of inhibiting activation by cytokines, in particular by interferon &ggr;, of expression of the CIITA gene which itself is involved in controlling and regulating the expression of genes coding for MHC class II molecules.
Molecules of the major histocompatibility complex (hereinafter designated MHC) class II are transmembrane heterodimeric glycoproteins which are directly involved in the activation of CD4+ T helper lymphocytes during the immune response.
In man, this class II complex is represented by molecules belonging to the HLA (human leukocyte antigen) system. Genes coding for the &agr; and &bgr; chains constituting the HLA-DR, HLA-DQ and HLA-DP molecules are located in region D of chromosome 6.
Expression of these genes is perfectly regulated. In contrast to genes coding for MHC class I molecules which are expressed ubiquitously, the genes coding for MHC class II molecules are expressed either constitutively uniquely in some cell types such as B lymphocytes, activated T lymphocytes, macrophages, thymic epithelium cells, or dentritic cells such as Langerhans cells, or inductively after stimulation, for example by cytokines, and more particularly by interferon &ggr; (INF &ggr;) or interleukin 4 (IL4), in several other cell types such as cells from the macrophage or monocyte line, endothelial cells, fibroblasts, muscle cells or cancer cells such as melanoma cells.
Further, in B lymphocytes, the expression of genes coding for MHC class II molecules is transitory. Differentiation of B cells into plasmocytes producing immunoglobulins is accompanied by extinction of certain genes, including those coding for MHC class II.
Similarly, it has been shown that the amount of expression of MHC class II molecules constitutes a determining factor in the T cell activation process.
As a result, it is clear that molecular mechanisms for regulating the expression of these genes constitute a key element in the effectiveness of the immune response. Any defect in this regulation process can result in substantial immunological problems, or autoimmune diseases. Thus in some cases, abnormal expression of MHC class II genes has been observed on the surface of cells which normally should not express such genes. Similarly, an over-expression of these genes can be observed, leading to aberrant and uncontrolled activation of CD4+ lymphocytes (BOTTAZZO et al., 1986, Immunol. Rev., 94, 137-169). Such manifestations could be at least partially responsible for diseases such as insulin dependent diabetes, multiple sclerosis, rheumatoid arthritis or lupus erythematosus. In contrast, in some patients immunodeficiency has been demonstrated resulting from problems with expression of MHC class II genes. An example which can be cited is BLS syndrome (bare lymphocyte syndrome), which is a recessive autosomal disease in which expression of MHC class II genes is very limited or even non-existent, resulting in an absence of cellular and humoral immune response and accompanied by numerous infections which are often fatal.
A number of scientific teams have analysed the mechanisms of regulation of MHC class II gene expression and have identified a certain number of transactivating molecules which can directly or indirectly bind to specific promoter sequences of said genes (for a review, see MACH et al., 1996, Annu. Rev. Immunol. 14, 301-331).
The Applicant has previously identified and characterized one of these factors, the CIITA factor (class II transactivator) [STEIMLE et al., 1993, Cell 75, 135-146 and EP-A-0 648 836]. Further, International patent application WO-A-9606107 has shown that there are two domains in the CIITA factor which are more involved in activation of transcription of MHC class II genes. However, surprisingly and in contrast to that which has been observed for the other factors involved in regulating the expression of MHC class II genes [COGSWELL et al., 1991, Crit. Rev. Immunol. 11, 87-112], STEIMLE et al have shown that expression of the CIITA factor coincides closely with expression of MHC class II genes and is absolutely required both for constitutive expression and for induction of said MHC genes. Further, SILACCI et al (1994, J. Exp. Med., 180, 1329-1336) have shown that extinction of MHC class II genes during differentiation of plasmocytes is associated with extinction of the gene coding for the CIITA factor.
Further, LENNON et al (1997, Immunogenetics, 45, 266-273) have identified the promoter sequence of a CIITA gene which is responsible for differential expression of this factor in B cells. However, the existence of this single sequence does not explain why differential expression of the CIITA factor is observed in different cell types. Further, it does not account for induction by cytokines.
In previous studies, the Applicant used samples from different tissues of human origin to identify the complex organisation of sequences providing control of expression of the CIITA factor, the Applicant isolated and characterized several promoter regions and the Applicant demonstrated the existence of different forms of the CIITA factor and also different CIITA genes. These studies have formed the basis of a publication (MUHLETHALER-MOTTET et al., 1997, EMBO J., 16, 2851-2860) and form the subject matter of French patent application 97/04954 the contents of which are hereby incorporated into the present application. The inventors have thus shown that the different promoters identified can be activated selectively: two of the promoters are responsible for constitutive expression of the CIITA gene in dendritic cells (promoter I) and in B lymphocytes (promoter III) while promoter IV is involved in expressing the CIITA gene after induction by a cytokine, in particular interferon &ggr;.
More particularly, the inventors have identified a sequence capable of expressing a transcriptional promoter activity after induction by a cytokine, such as interferon &ggr; or interleukin 4. Such a sequence is represented by the sequence comprising all or part of a sequence identified as SEQ ID NO:1 (set forth on Table 1), or its complementary sequence. An analysis of this sequence has identified several regions corresponding to cis acting regulation expression sites, such as the NF-GMa site, the GAS element, the E-box or the IRF-1 factor binding site (MUHLETHALER-MOTTET et al, 1997, EMBO J., 16, 2851-2860 and FIG.
1
).
More recently, a number of studies have provided a deeper understanding of the succession of events and signals involved in activating genes expressed in response to induction by a cytokine, in particular by interferon y. An activation scheme has been proposed by DARNELL, (1997, Science 277, 1630-1635). In that model, firstly the activating cytokine, for example interferon &ggr;, binds to its surface cell receptors thus enabling activation of cellular tyrosine kinases JAK1 and JAK2. Then the tyrosine residues of the STAT1 transcription factor, located in the cell cytoplasm, are phosphorylated by activated JAK kinases. This phosphorylation then enables the activated STAT1 factor to migrate into the nucleus where it binds to the GAS box of promoters inducible by cytokines (for example interferon &ggr;) thus enabling activated expression of genes under the control of such promoters.
The implication of such a JAK/STAT1 activation system in the control of expression of CIITA genes inducible by interferon &ggr; has been the subject of studies which have established that, as with other genes which are inducible by interferon &ggr;, expression of the CIITA factor cannot be induced in cell lines which are deficient for JAK1 (CHANG et al., 1994, J. Exp. Med., 180, 1367-1374).
Similarly, MERAZ et al. (1996, Cell, 84, 431-442) have shown that CIITA gene expression is not induced by interferon Y in bone marrow macrophages from the STAT1
−
/
−
mouse, suggesting a determining role for STAT1 in inducing the expression of the CIITA gene by interferon &ggr;.
Further, LEE and BEN
Horlick Kenneth R.
Lerner David Littenberg Krumholz & Mentlik LLP
Novimaune S.A.
LandOfFree
Method for screening compounds capable of inhibiting binding... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Method for screening compounds capable of inhibiting binding..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Method for screening compounds capable of inhibiting binding... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2907209