Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Peptide containing doai
Reexamination Certificate
2000-10-27
2004-11-02
Low, Christopher S. F. (Department: 1653)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Peptide containing doai
C424S094100, C424S085100, C435S007100, C435S007200, C435S007800, C530S350000
Reexamination Certificate
active
06812203
ABSTRACT:
TECHNICAL FIELD
The invention relates to CD40 binding proteins, which can be used as modulators of the CD40 signaling pathway and/or the CD40-induced nuclear factor kappa B (NF-kB) activating pathway and thus useful in the treatment of CD40 related diseases (e.g., inflammatory diseases) and/or NF-kB related diseases and/or in the improvement of anti-tumor treatments. The current invention also relates to nucleic acid sequences coding for the CD40 interacting proteins (also called “TTRAP” (“TRAF and TNF receptor associated protein”) for CD40 receptor associated protein). The invention further relates to the use of the polypeptides derived from these CD40 interacting proteins in the treatment of CD40 and/or NF-kB related diseases and or cancer. Furthermore, the invention concerns pharmaceutical preparations comprising the CD40 interacting proteins or polypeptides derived from these proteins.
BACKGROUND
CD40 is a receptor of tumor necrosis factor (“TNF”) receptor superfamily (Banchereau et al., 1994), which is expressed at the surface of B-cells, antigen presenting cells (APC), and several non-hematopoietic cells such as endothelial cells (Hollenbaugh et al., 1995), epithelial cells (Galy & Spits, 1992), fibroblasts (Fries et al., 1995) and keratinocytes (Gaspari et al., 1996). The ligand for CD40 (CD40L) occurs mainly on activated T-cells. Up to now the role of CD40 was mainly studied in the context of the T-cell APC/B-cell interaction (for a review, see Noelle, 1996). Amongst others, the CD40-CD40L interaction seems to be important for the T-cell mediated immunity and for primary and secondary humoral immune response. These findings were confirmed by experiments in mouse models showing that treatment with anti-CD40L antibodies resulted in blocking of the development of mouse equivalents of human autoimmune diseases such as arthritis (Durie et al., 1993), oophoritis (Griggs et al., 1996) and multiple sclerosis (Gerritse et al., 1996). Activation and transduction through the CD40 pathway within this biological system is mainly responsible for B cell activation and the humoral immune response accordingly.
Apart from NF-kB, factors that can be activated by stimulation of CD40 are NF-AT (Francis et al, 1995) c-Jun, ATF-2 and IRF-1 (Karmann et al., 1996). All these factors play an important role in inflammation.
The CD40L induced signal transduction is, like TNF, mediated by the binding of TNF-Receptor Associated Factors (TRAF's) to the cytoplasmic domain of the receptor. Chaudhuri et al. (1997) demonstrated that, at least in human B cell lines, CD40 and TRAF2 are constitutively associated with each other, and that this association is inhibited by CD40 mediated signals. Apart from the binding with TRAF2, the cytoplasmic domain of CD40, which consists of 62 amino acids at positions 196-257 (mature human CD40—numbering according to Kashiwada et al., 1998), is known to associate with TRAF3, TRAFS, TRAF6 and Janus kinase 3. TRAF6 binds to the amino-terminal cytoplasmic tail of CD40 at positions 210-225, although it can not be excluded that full association of TRAF6 with CD40 may also require the carboxy-terminal part at positions 226-249 (Ishida et al., 1996). TRAF2, TRAF3 and TRAF5 bind to the carboxy-terminal CD40 cytoplasmic domain at positions 226-249 (Ishida et al., 1996).
Stimulation of CD40 results in activation of protein kinases, the nitrogen-activated protein kinase and Janus kinase 3/signal transducer and activator of Transcription 3. Moreover, stimulation of CD40 mediates critical biological effects in B cell growth survival and differentiation.
It is known that TRAF2 and TRAF5 play a role in NF-kB activation in signaling through CD40, as well as “TNF receptor type I (TNF-RI), TNF receptor type II (TNF-RII)” CD30 and lymphotoxin b receptor. TRAF6 participates in NF-kB activation signaled by CD40 and IL-1 receptor. In addition to these data, International Patent Applications WO 96/16665 and WO 96/28568 disclose a TRAF like protein that binds to the cytoplasmic domain of CD40.
DESCRIPTION OF THE INVENTION
We show herein that at least two other proteins exist which unexpectedly interact with the cytoplasmic domain of CD40. Even more surprisingly, neither of these proteins shows significant homology, with any known CD40, interacting proteins. Further, no significant homology exists between the two proteins themselves. These proteins should therefore be considered as two new classes of CD40 interacting proteins.
The present invention thus concerns an isolated functional protein capable of interacting with the cytoplasmic domain of CD40 and/or other receptors of the TNF receptor superfamily such as CD30 or TNF receptor II, wherein the protein has no homology to TRAP-proteins.
The invention also includes an isolated functional protein either comprising an amino acid sequence with 70-100% homology to the amino acid sequence depicted in SEQ ID NO: 2 or either comprising an amino acid sequence with 70-100% homology to the amino acid sequence depicted in SEQ ID NO: 4 or in the alternative comprising an amino acid sequence with 70-100% homology to the amino acid sequence depicted in SEQ ID NO: 6.
More specifically, the functional protein comprises an amino acid sequence with 70-100% homology to the amino acids 54-362 of SEQ ID NO: 2, even more specifically the functional protein comprises an amino acid sequences with 70-100% homology to the amino acids 274-362 of SEQ ID NO: 2 or in the alternative and/or comprising an amino acid sequence with 70-100% homology to the amino acids 2-245 of the SEQ ID NO: 6.
Furthermore, the invention also includes those proteins or peptides having 70-100% homology to at least, any of the three peptides as depicted in SEQ ID NO: 2 located between the residues numbering 115-121, 145-153 and 347-352 respectively. The amino acid sequence of residue numbering 115-121 is SLITWNI; the amino acid sequence of residue numbering 145-153 is PDVIFLQEV and the amino acid sequence of residue numbering 347-352 is FPSDHW.
“Homology”, in this context, means identical or similar to the referenced sequence while obvious replacements/modifications of any of the amino acids provided, are included as well. A homology search in this respect can be performed with the BLAST-P (Basic Local Alignment Search Tool), a program well known to those of skill in the art. For the corresponding nucleic acid sequence, homology is referred to the BLASTX and BLASTN programs known in the art.
The invention thus relates to a DNA sequence encoding CD40 receptor associated protein or encoding an immunologically active and/or functional fragment of such a protein, selected from the group consisting of:
(a) DNA sequences comprising a nucleotide sequence encoding a protein comprising the amino acid sequence as given in SEQ ID NO: 2;
(b) DNA sequences comprising a nucleotide sequence as given in SEQ ID NO: 1;
(c) DNA sequences hybridizing with the complementary strand of a DNA sequence as defined in (a) or (b) and encoding an amino acid sequence which is at least 70% identical to the amino acid sequence encoded by the DNA sequence of (a) or (b);
(d) DNA sequences, the nucleotide sequence of which is degenerated as a result of the genetic code to a nucleotide sequence of a DNA sequence as defined in any one of (a) to (c); and
(e) DNA sequences encoding a fragment of a protein encoded by a DNA sequence of any one of (a) to (d).
One embodiment of the invention is a protein with SEQ ID NO: 2. Another embodiment of the invention is a protein with SEQ ID NO: 4. A further embodiment of the invention concerns a protein with SEQ ID NO: 6.
A further aspect of the invention is the use of the aforementioned proteins, or biologically active fragments thereof, to modulate and/or inhibit members of the TNF receptor superfamily such as CD40, CD30 or TNF-receptor II in their signaling activity and/or CD40-induced NF-kB activation and/or JUN-kinase activity.
The isolated functional protein according to the invention and/or a functional fragment thereof can be used to treat TRAF-related, CD40-related, NF-kB related a
Huylebroeck Danny F. E.
Pype Stefan M. C.
Remacle Jacques E. F.
Liu Samuel Wei
Low Christopher S. F.
TraskBritt
Vlaams Interuniversitair Instituut voor Biotechnologie VZW
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