+

WO1998042841A1 - Proteine hkafe42 apparentee a l'interferon - Google Patents

Proteine hkafe42 apparentee a l'interferon Download PDF

Info

Publication number
WO1998042841A1
WO1998042841A1 PCT/GB1998/000915 GB9800915W WO9842841A1 WO 1998042841 A1 WO1998042841 A1 WO 1998042841A1 GB 9800915 W GB9800915 W GB 9800915W WO 9842841 A1 WO9842841 A1 WO 9842841A1
Authority
WO
WIPO (PCT)
Prior art keywords
polypeptide
hkafe42
seq
nucleotide sequence
amino acid
Prior art date
Application number
PCT/GB1998/000915
Other languages
English (en)
Inventor
Lee James Beeley
Stephen Lee Mathias
Original Assignee
Smithkline Beecham P.L.C.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from GBGB9706178.2A external-priority patent/GB9706178D0/en
Application filed by Smithkline Beecham P.L.C. filed Critical Smithkline Beecham P.L.C.
Priority to JP10545249A priority Critical patent/JP2000513947A/ja
Priority to EP98913911A priority patent/EP0934414A1/fr
Publication of WO1998042841A1 publication Critical patent/WO1998042841A1/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/52—Cytokines; Lymphokines; Interferons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy

Definitions

  • This invention relates to newly identified polynucleotides, polypeptides encoded by them and to the use of such polynucleotides and polypeptides, and to their production. More particularly, the polynucleotides and polypeptides of the present invention relate to interferon- related protein family, hereinafter referred to as HKAFE42. The invention also relates to inhibiting or activating the action of such polynucleotides and polypeptides.
  • Cytokine is a generic term and encompasses various families of regulators including growth factors (GF), colony- stimulating factors (CSF), interferons (IFN), interleukins (IL), lymphokines and monokines (For reviews, see Callard and Gearing, The Cytokines Facts Book, Academic Press, London, 1994 and Nicola, ed., Guidebook to Cytokines and Their Receptors, Sambrook and Tooze, Oxford, 1994). Cytokines are generallly glycoproteins with a molecular weight of 20,000-40,000 kD and work at very low concentrations (pM order).
  • the first, interferon -gamma is secreted from T- cells and natural killer cells and plays a key role in inflammatory processes. Clinically, interferon - gamma has been shown to alleviate symptoms of chronic granulomatous disease and has also shown promise for the treatment of infectious disease and neoplasias.
  • the second, Nerve Growth Factor (NGF) supports the survival of embryonic neurones and continues to be involved in the maintenance of neuronal phenotype in the mature nervous system. Intra-ventricular injection of NGF to aged, learning-deficient rats has been shown to correct the learning deficiencies (Fischer, et al., Nature 329: 65-8, 1987). Clinically, decreased levels of the mRNA of the closely related cytokine BDNF have been observed in the hippocampus of patients with Alzheimer's disease (Phillis et al., Neuron 7: 695-702, 1991).
  • the mouse PC4 gene (Varnum et al., Oncogene 4(10): 1263-65, 1989) is an immediate early gene related to IFN-gamma. PC4 is induced during the course of neronal differentiation and also in the cell line PC 12 by NGF. The relationship of PC4 to interferons and lymphokines suggests that it could play a role in regulating genetic pathways induced by NGF (Tirone and Shooter, PNAS 86(6): 2088-92, 1989). A role for PC4 has also been postulated in muscle differentiation (Guardavaccaro et al., Cell Growth Differ. 6(2): 159-69, 1995).
  • the invention relates to HKAFE42 polypeptides and recombinant materials and methods for their production.
  • Another aspect of the invention relates to methods for using such HKAFE42 polypeptides and polynucleotides.
  • Such uses include the treatment of disorders of the central nervous system (including anxiety, depression, schizophrenia and Parkinson's disease), muscle diseases (including muscular dystrophies and multiple sclerosis), Alzheimer's disease, hematopoetic disorders, allergic diseases, autoimmune diseases, immunodeficiency and cancer, among others.
  • the invention relates to methods to identify agonists and antagonists using the materials provided by the invention, and treating conditions associated with HKAFE42 imbalance with the identified compounds.
  • Yet another aspect of the invention relates to diagnostic assays for detecting diseases associated with inappropriate HKAFE42 activity or levels.
  • HKAFE42 refers, among others, generally to a polypeptide having the amino acid sequence set forth in SEQ ID NO:2 or an allelic variant thereof.
  • HKAFE42 activity or HKAFE42 polypeptide activity refers to the metabolic or physiologic function of said HKAFE42 including similar activities or improved activities or these activities with decreased undesirable side-effects. Also included are antigenic and immunogenic activities of said HKAFE42.
  • HKAFE42 gene refers to a polynucleotide having the nucleotide sequence set forth in SEQ ID NO: 1 or allelic variants thereof and/or their complements.
  • Antibodies as used herein includes polyclonal and monoclonal antibodies, chimeric, single chain, and humanized antibodies, as well as Fab fragments, including the products of an Fab or other immunoglobulin expression library.
  • Isolated means altered “by the hand of man” from the natural state. If an "isolated” composition or substance occurs in nature, it has been changed or removed from its original environment, or both.
  • a polynucleotide or a polypeptide naturally present in a living animal is not “isolated/' but the same polynucleotide or polypeptide separated from the coexisting materials of its natural state is "isolated", as the term is employed herein.
  • Polynucleotide generally refers to any polyribonucleotide or polydeoxribonucleotide, which may be unmodified RNA or DNA or modified RNA or DNA.
  • Polynucleotides include, without limitation single- and double-stranded DNA, DNA that is a mixture of single- and double-stranded regions, single- and double-stranded RNA, and RNA that is mixture of single- and double-stranded regions, hybrid molecules comprising DNA and RNA that may be single- stranded or, more typically, double-stranded or a mixture of single- and double-stranded regions.
  • polynucleotide refers to triple-stranded regions comprising RNA or DNA or both RNA and DNA.
  • the term polynucleotide also includes DNAs or RNAs containing one or more modified bases and DNAs or RNAs with backbones modified for stability or for other reasons.
  • Modified bases include, for example, tritylated bases and unusual bases such as inosine.
  • polynucleotide embraces chemically, enzymatically or metabolically modified forms of polynucleotides as typically found in nature, as well as the chemical forms of DNA and RNA characteristic of viruses and cells.
  • Polynucleotide also embraces relatively short polynucleotides, often referred to as oligonucleotides.
  • Polypeptide refers to any peptide or protein comprising two or more amino acids joined to each other by peptide bonds or modified peptide bonds, i.e., peptide isosteres.
  • Polypeptide refers to both short chains, commonly referred to as peptides, oligopeptides or oligomers, and to longer chains, generally referred to as proteins. Polypeptides may contain amino acids other than the 20 gene-encoded amino acids.
  • Polypeptides include amino acid sequences modified either by natural processes, such as posttranslational processing, or by chemical modification techniques which are well known in the art. Such modifications are well described in basic texts and in more detailed monographs, as well as in a voluminous research literature.
  • Modifications can occur anywhere in a polypeptide, including the peptide backbone, the amino acid side-chains and the amino or carboxyl termini. It will be appreciated that the same type of modification may be present in the same or varying degrees at several sites in a given polypeptide. Also, a given polypeptide may contain many types of modifications. Polypeptides may be branched as a result of ubiquitination, and they may be cyclic, with or without branching. Cyclic, branched and branched cyclic polypeptides may result from posttranslation natural processes or may be made by synthetic methods.
  • Modifications include acetylation, acylation, ADP-ribosylation, amidation, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphotidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent crosslinks, formation of cystine, formation of pyroglutamate, formylation, gamma-carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, transfer- RNA mediated addition of amino acids to proteins such as arginylation, and ubiquitination.
  • Variant is a polynucleotide or polypeptide that differs from a reference polynucleotide or polypeptide respectively, but retains essential properties.
  • a typical variant of a polynucleotide differs in nucleotide sequence from another, reference polynucleotide. Changes in the nucleotide sequence of the variant may or may not alter the amino acid sequence of a polypeptide encoded by the reference polynucleotide. Nucleotide changes may result in amino acid substitutions, additions, deletions, fusions and truncations in the polypeptide encoded by the reference sequence, as discussed below.
  • a typical variant of a polypeptide differs in amino acid sequence from another, reference polypeptide. Generally, differences are limited so that the sequences of the reference polypeptide and the variant are closely similar overall and, in many regions, identical.
  • a variant and reference polypeptide may differ in amino acid sequence by one or more substitutions, additions, deletions in any combination.
  • a substituted or inserted amino acid residue may or may not be one encoded by the genetic code.
  • a variant of a polynucleotide or polypeptide may be a naturally occurring such as an allelic variant, or it may be a variant that is not known to occur naturally. Non-naturally occurring variants of polynucleotides and polypeptides may be made by mutagenesis techniques or by direct synthesis.
  • Identity is a relationship between two or more polypeptide sequences or two or more polynucleotide sequences, as determined by comparing the sequences.
  • identity also means the degree of sequence relatedness between polypeptide or polynucleotide sequences, as the case may be, as determined by the match between strings of such sequences.
  • Preferred methods to determine identity are designed to give the largest match between the sequences tested. Methods to determine identity and similarity are codified in publicly available computer programs. Preferred computer program methods to determine identity and similarity between two sequences include, but are not limited to, the GCG program package (Devereux, J., et al., Nucleic Acids Research 12(1): 387 (1984)), BLASTP, BLASTN, and FASTA (Atschul, S.F. et al., J. Molec. Biol.
  • the BLAST X program is publicly available from NCBI and other sources (BLAST Manual, Altschul, S., et al, NCBI NLM NIH Bethesda, MD 20894; Altschul, S., et al, J. Mol. Biol. 215: 403-410 (1990).
  • the well known Smith Waterman algorithm may also be used to determine identity.
  • Preferred parameters for polypeptide sequence comparison include the following: 1) Algorithm: Needleman and Wunsch, J. Mol Biol. 48: 443-453 (1970) Comparison matrix: BLOSSUM62 from Hentikoff and Hentikoff, Proc. Natl. Acad. Sci. USA. 89: 10915-10919 (1992) Gap Penalty: 12 Gap Length Penalty: 4
  • a program useful with these parameters is publicly available as the "gap" program from Genetics Computer Group, Madison WI.
  • the aforementioned parameters are the default parameters for polynucleotide comparisons.
  • a polynucleotide sequence of the present invention may be identical to the reference sequence of SEQ ID NO: l, that is be 100% identical, or it may include up to a certain integer number of nucleotide alterations as compared to the reference sequence.
  • Such alterations are selected from the group consisting of at least one nucleotide deletion, substitution, including transition and transversion, or insertion, and wherein said alterations may occur at the 5' or 3' terminal positions of the reference nucleotide sequence or anywhere between those terminal positions, interspersed either individually among the nucleotides in the reference sequence or in one or more contiguous groups within the reference sequence.
  • the number of nucleotide alterations is determined by multiplying the total number of nucleotides in SEQ ID NO: 1 by the numerical percent of the respective percent identity(divided by 100) and subtracting that product from said total number of nucleotides in SEQ ID NO: l, or: n n ⁇ x n " (*n * y)> wherein n n is the number of nucleotide alterations, x n is the total number of nucleotides in SEQ
  • y is, for instance, 0.70 for 70%, 0.80 for 80%, 0.85 for 85%, 0.90 for 90%, 0.95 for 95%,etc, and wherein any non-integer product of x n and y is rounded down to the nearest integer prior to subtracting it from x n .
  • Alterations of a polynucleotide sequence encoding the polypeptide of SEQ ID NO:2 may create nonsense, missense or frameshift mutations in this coding sequence and thereby alter the polypeptide encoded by the polynucleotide following such alterations.
  • a polypeptide sequence of the present invention may be identical to the reference sequence of SEQ ID NO:2, that is be 100% identical, or it may include up to a certain integer number of amino acid alterations as compared to the reference sequence such that the % identity is less than 100%).
  • Such alterations are selected from the group consisting of at least one amino acid deletion, substitution, including conservative and non-conservative substitution, or insertion, and wherein said alterations may occur at the amino- or carboxy- terminal positions of the reference polypeptide sequence or anywhere between those terminal positions, interspersed either individually among the amino acids in the reference sequence or in one or more contiguous groups within the reference sequence.
  • the number of amino acid alterations for a given % identity is determined by multiplying the total number of amino acids in SEQ ID NO:2 by the numerical percent of the respective percent identity(divided by 100) and then subtracting that product from said total number of amino acids in SEQ ID NO:2, or: n a ⁇ x a - (x a • y), wherein n a is the number of amino acid alterations, x a is the total number of amino acids in SEQ ID NO:2, and y is, for instance 0.70 for 70%, 0.80 for 80%, 0.85 for 85% etc., and wherein any non-integer product of x a and y is rounded down to the nearest integer prior to subtracting it from x a .
  • the present invention relates to HKAFE42 polypeptides (or HKAFE42 proteins).
  • the HKAFE42 polypeptides includes the polypeptide of SEQ ID NO:2; as well as polypeptides comprising the amino acid sequence of SEQ ID NO: 2; and polypeptides comprising the amino acid sequence which have at least 95% identity to that of SEQ ID NO:2 over its entire length, and still more preferably at least 96% identity, and even still more preferably at least 97% identity to SEQ ID NO: 2. Furthermore, those with at least 98-99% are highly preferred.
  • HKAFE42 polypeptides having the amino acid sequence which have at least 95% identity to the polypeptide having the amino acid sequence of SEQ ID NO:2 over its entire length, and still more preferably at least 96% identity, and still more preferably at least 97% identity to SEQ ID NO:2. Furthermore, those with at least 98-99%) are highly preferred.
  • HKAFE42 polypeptide exhibit at least one biological activity of HKAFE42.
  • the HKAFE42 polypeptides may be in the form of the "mature" protein or may be a part of a larger protein such as a fusion protein. It is often advantageous to include an additional amino acid sequence which contains secretory or leader sequences, pro-sequences, sequences which aid in purification such as multiple histidine residues, or an additional sequence for stability during recombinant production.
  • a fragment is a polypeptide having an amino acid sequence that entirely is the same as part, but not all, of the amino acid sequence of the aforementioned HKAFE42 polypeptides.
  • fragments may be "free-standing," or comprised within a larger polypeptide of which they form a part or region, most preferably as a single continuous region.
  • Representative examples of polypeptide fragments of the invention include, for example, fragments from about amino acid number 1-20, 21-40, 41-60, 61-80, 81-100, and 101 to the end of HKAFE42 polypeptide.
  • “about” includes the particularly recited ranges larger or smaller by several, 5, 4, 3, 2 or 1 amino acid at either extreme or at both extremes.
  • Preferred fragments include, for example, truncation polypeptides having the amino acid sequence of HKAFE42 polypeptides, except for deletion of a continuous series of residues that includes the amino terminus, or a continuous series of residues that includes the carboxyl terminus or deletion of two continuous series of residues, one including the amino terminus and one including the carboxyl terminus.
  • fragments characterized by structural or functional attributes such as fragments that comprise alpha-helix and alpha-helix forming regions, beta-sheet and beta-sheet-forming regions, turn and turn-forming regions, coil and coil-forming regions, hydrophilic regions, hydrophobic regions, alpha amphipathic regions, beta amphipathic regions, flexible regions, surface-forming regions, substrate binding region, and high antigenic index regions.
  • Other preferred fragments are biologically active fragments.
  • Biologically active fragments are those that mediate HKAFE42 activity, including those with a similar activity or an improved activity, or with a decreased undesirable activity. Also included are those that are antigenic or immunogenic in an animal, especially in a human.
  • all of these polypeptide fragments retain the biological activity of the HKAFE42, including antigenic activity.
  • the most preferred fragment is that having the amino acid sequence of SEQ ID NO: 4.
  • Variants of the defined sequence and fragments also form part of the present invention.
  • Preferred variants are those that vary from the referents by conservative amino acid substitutions — i.e., those that substitute a residue with another of like characteristics. Typical such substitutions are among Ala, Val, Leu and He; among Ser and Thr; among the acidic residues Asp and Glu; among Asn and Gin; and among the basic residues Lys and Arg; or aromatic residues Phe and Tyr.
  • Particularly preferred are variants in which several, 5-10, 1-5, or 1-2 amino acids are substituted, deleted, or added in any combination.
  • the HKAFE42 polypeptides of the invention can be prepared in any suitable manner.
  • Such polypeptides include isolated naturally occurring polypeptides, recombinantly produced polypeptides, synthetically produced polypeptides, or polypeptides produced by a combination of these methods. Means for preparing such polypeptides are well understood in the art.
  • HKAFE42 polynucleotides include isolated polynucleotides which encode the HKAFE42 polypeptides and fragments, and polynucleotides closely related thereto. More specifically, HKAFE42 polynucleotide of the invention include a polynucleotide comprising the nucleotide sequence contained in SEQ ID NO: 1 encoding a HKAFE42 polypeptide of SEQ ID NO: 2, and polynucleotides having the particular sequences of SEQ ID NO: 1.
  • HKAFE42 polynucleotides further include a polynucleotide comprising a nucleotide sequence that has at least 99% identity over its entire length to a nucleotide sequence encoding the HKAFE42 polypeptide of SEQ ID NO:2, and a polynucleotide comprising a nucleotide sequence that is at least 99% identical to that of SEQ ID NO: l over its entire length.
  • the invention also provides polynucleotides which are complementary to such HKAFE42 polynucleotides.
  • HKAFE42 of the invention is structurally related to other proteins of the interferon-related protein family, as shown by the results of sequencing the cDNA encoding human HKAFE42.
  • the cDNA sequence of SEQ ID NO: 1 contains an open reading frame (nucleotide number 247 to 1599) encoding a polypeptide of 451 amino acids of SEQ ID NO:2.
  • Amino acid sequence of SEQ ID NO:2 has about 95% identity (using GCG BestFit) in 450 amino acid residues with mouse PC4 (B.C. Varnum, R.W. Lim and H.R. Herschman, Oncogene 4: 1263-1265, 1989).
  • Nucleotide sequence of SEQ ID NO: 1 has about 99% identity (using blastn2) in 1767 nucleotide residues with Human mRNA for nerve growth factor-inducible PC4 homolog (GenBank Accession Y10313).
  • Preferred polypeptides and polynucleotides of the present invention are expected to have, inter alia, similar biological functions/properties to their homologous polypeptides and polynucleotides.
  • the present invention also relates to partial or other polynucleotide and polypeptide sequences which were first identified prior to the determination of the corresponding full length sequences of SEQ ID NO: 1 and SEQ ID NO:2.
  • the present invention provides for an isolated polynucleotide comprising:
  • nucleotide sequence which has at least 90% identity, preferably at least 95%> identity, more preferably at least 97-99% identity to SEQ ID NO:3 over the entire length of SEQ ID NO:3;
  • nucleotide sequence which has at least 90% identity, preferably at least 95% identity, more preferably at least 97-99% identity, to SEQ ID NO:3 over the entire length of SEQ ID NO:3;
  • the present invention further provides for a polypeptide which:
  • (a) comprises an amino acid sequence which has at least 95%> identity, preferably at least 97- 99% identity, to that of SEQ ID NO:4 over the entire length of SEQ ID NO:4;
  • (b) has an amino acid sequence which is at least at least 95% identity, preferably at least 97-99%) identity, to the amino acid sequence of SEQ ID NO:4 over the entire length of SEQ ID NO:4;
  • (c) comprises the amino acid of SEQ ID NO:4;
  • (d) is the polypeptide of SEQ ID NO:4; as well as polypeptides encoded by a polynucleotide comprising the sequence contained in SEQ ID NO:3.
  • the cDNA sequence of SEQ ID NO: 3 contains an open reading frame encoding a polypeptide of 452 amino acids.
  • Amino acid sequence of SEQ ID NO:4 has about 93%) identity (using GCG Bestfit) in 452 amino acid residues with Mouse PC4 (Varnum et al., Oncogene 4: 1263-65, 1989).
  • SEQ ID NO:4 is 19% identical and 50% similar to human interferon- gamma over 212 amino acid residues (Gray and Goeddel, Nature 295: 503-8, 1982).
  • the nucleotide sequence of SEQ ID NO:3 has about 88% > identity (using GCG Bestfit) in 181 1 nucleotide residues with Mouse PC4 (Varnum et al., Oncogene 4: 1263-65, 1989).
  • nucleotide sequence of SEQ ID NO:3 and the peptide sequence encoded thereby are derived from EST (Expressed Sequence Tag) sequences. It is recognised by those skilled in the art that there will inevitably be some nucleotide sequence reading errors in EST sequences (see Adams, M.D. et al, Nature 377 (supp) 3, 1995). Accordingly, the nucleotide sequence of SEQ ID NO:3 and the peptide sequence encoded therefrom are therefore subjec to the same inherent limitations in sequence accuracy. Furthermore, the peptide sequence encoded by SEQ ID NO:3 comprises a region of identity or close homology and/or close structural similarity (for example a conservative amino acid difference) with the closest homologous or structurally similar protein.
  • One polynucleotide of the present invention encoding HKAFE42 may be obtained using standard cloning and screening, from a cDNA library derived from mRNA in cells of human keratinocyte using the expressed sequence tag (EST) analysis (Adams, M.D., et al. Science (1991) 252: 1651-1656; Adams, M.D. et al, Nature, (1992) 355:632-634; Adams, M.D., et al, Nature (1995) 377 Supp:3-174).
  • Polynucleotides of the invention can also be obtained from natural sources such as genomic DNA libraries or can be synthesized using well known and commercially available techniques.
  • the nucleotide sequence encoding HKAFE42 polypeptide of SEQ ID NO:2 may be identical to the polypeptide encoding sequence contained in Table 1 (nucleotide number 247 to 1599 of SEQ ID NO: 1), or it may be a sequence, which as a result of the redundancy (degeneracy) of the genetic code, also encodes the polypeptide of SEQ ID NO:2.
  • the polynucleotide may include the coding sequence for the mature polypeptide or a fragment thereof, by itself; the coding sequence for the mature polypeptide or fragment in reading frame with other coding sequences, such as those encoding a leader or secretory sequence, a pre-, or pro- or prepro- protein sequence, or other fusion peptide portions.
  • a marker sequence which facilitates purification of the fused polypeptide can be encoded.
  • the marker sequence is a hexa- histidine peptide, as provided in the pQE vector (Qiagen, Inc.) and described in Gentz et al, Proc NatlAcadSci USA (1989) 86:821-824, or is an HA tag.
  • the polynucleotide may also contain non- coding 5' and 3' sequences, such as transcribed, non-translated sequences, splicing and polyadenylation signals, ribosome binding sites and sequences that stabilize mRNA.
  • HKAFE42 variants comprising the amino acid sequence of HKAFE42 polypeptide of Table 2 (SEQ ID NO:2) in which several, 5-10, 1-5, 1-3, 1-2 or 1 amino acid residues are substituted, deleted or added, in any combination.
  • SEQ ID NO: 3 encoding the amino acid sequence of Table 4 (SEQ ID NO: 4).
  • the present invention further relates to polynucleotides that hybridize to the herein above- described sequences.
  • the present invention especially relates to polynucleotides which hybridize under stringent conditions to the herein above-described polynucleotides.
  • stringent conditions means hybridization will occur only if there is at least 80%>, and preferably at least 90%o, and more preferably at least 95%o, yet even more preferably 97- 99% identity between the sequences.
  • Polynucleotides of the invention which are identical or sufficiently identical to a nucleotide sequence contained in SEQ ID NO: 1 or a fragment thereof (including that of SEQ ID NO:3), may be used as hybridization probes for cDNA and genomic DNA, to isolate full-length cDNAs and genomic clones encoding HKAFE42 polypeptide and to isolate cDNA and genomic clones of other genes (including genes encoding homologs and orthologs from species other than human) that have a high sequence similarity to the HKAFE42 gene.
  • Such hybridization techniques are known to those of skill in the art.
  • these nucleotide sequences are 80% identical, preferably 90%> identical, more preferably 95% identical to that of the referent.
  • the probes generally will comprise at least 15 nucleotides.
  • such probes will have at least 30 nucleotides and may have at least 50 nucleotides. Particularly preferred probes will range between 30 and 50 nucleotides.
  • to obtain a polynucleotide encoding HKAFE42 polypeptide, including homologs and orthologs from species other than human comprises the steps of screening an appropriate library under stingent hybridization conditions with a labeled probe having the SEQ ID NO: 1 or a fragment thereof (including that of SEQ ID NO: 3), and isolating full-length cDNA and genomic clones containing said polynucleotide sequence.
  • a labeled probe having the SEQ ID NO: 1 or a fragment thereof (including that of SEQ ID NO: 3
  • HKAFE42 polynucleotides of the present invention further include a nucleotide sequence comprising a nucleotide sequence that hybridize under stringent condition to a nucleotide sequence having SEQ ID NO: 1 or a fragment thereof (including that of SEQ ID NO:3). Also included with HKAFE42 polypeptides are polypeptide comprising amino acid sequence encoded by nucleotide sequence obtained by the above hybridization condition.
  • Stringent hybridization conditions are as defined above or, alternatively, conditions under overnight incubation at 42°C in a solution comprising: 50%) formamide, 5xSSC (150mMNaCl, 15mM trisodium citrate), 50 mM sodium phosphate (pH7.6), 5x Denhardt's solution, 10 % dextran sulfate, and 20 microgram/ml denatured, sheared salmon sperm DNA, followed by washing the filters in O.lx SSC at about 65°C.
  • polynucleotides and polypeptides of the present invention may be employed as research reagents and materials for discovery of treatments and diagnostics to animal and human disease.
  • the present invention also relates to vectors which comprise a polynucleotide or polynucleotides of the present invention, and host cells which are genetically engineered with vectors of the invention and to the production of polypeptides of the invention by recombinant techniques.
  • Cell-free translation systems can also be employed to produce such proteins using RNAs derived from the DNA constructs of the present invention.
  • host cells can be genetically engineered to incorporate expression systems or portions thereof for polynucleotides of the present invention.
  • Introduction of polynucleotides into host cells can be effected by methods described in many standard laboratory manuals, such as Davis et al., BASIC METHODS IN MOLECULAR BIOLOGY (1986) and Sambrook et al., MOLECULAR CLONING: A LABORATORY MANUAL, 2nd Ed, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1989) such as calcium phosphate transfection, DEAE-dextran mediated transfection, transvection, microinjection, cationic lipid-mediated transfection, electroporation, transduction, scrape loading, ballistic introduction or infection.
  • bacterial cells such as streptococci, staphylococci, E. coli, Streptomyces and Bacillus subtilis cells
  • fungal cells such as yeast cells and Aspergillus cells
  • insect cells such as Drosophila S2 and Spodoptera Sf9 cells
  • animal cells such as CHO, COS, HeLa, C127, 3T3, BHK, HEK 293 and Bowes melanoma cells
  • plant cells include bacterial cells, such as streptococci, staphylococci, E. coli, Streptomyces and Bacillus subtilis cells
  • fungal cells such as yeast cells and Aspergillus cells
  • insect cells such as Drosophila S2 and Spodoptera Sf9 cells
  • animal cells such as CHO, COS, HeLa, C127, 3T3, BHK, HEK 293 and Bowes melanoma cells
  • Such systems include, among others, chromosomal, episomal and virus-derived systems, e.g., vectors derived from bacterial plasmids, from bacteriophage, from transposons, from yeast episomes, from insertion elements, from yeast chromosomal elements, from viruses such as baculoviruses, papova viruses, such as SV40, vaccinia viruses, adenoviruses, fowl pox viruses, pseudorabies viruses and retroviruses, and vectors derived from combinations thereof, such as those derived from plasmid and bacteriophage genetic elements, such as cosmids and phagemids.
  • viruses such as baculoviruses, papova viruses, such as SV40, vaccinia viruses, adenoviruses, fowl pox viruses, pseudorabies viruses and retroviruses
  • vectors derived from combinations thereof such as those derived from plasmid and bacteriophage genetic elements, such as cosmid
  • the expression systems may contain control regions that regulate as well as engender expression.
  • any system or vector suitable to maintain, propagate or express polynucleotides to produce a polypeptide in a host may be used.
  • the appropriate nucleotide sequence may be inserted into an expression system by any of a variety of well-known and routine techniques, such as, for example, those set forth in Sambrook et al, MOLECULAR CLONING, A LABORATORY MANUAL (supra).
  • secretion signals may be incorporated into the desired polypeptide. These signals may be endogenous to the polypeptide or they may be heterologous signals.
  • the polypeptide be produced at the surface of the cell.
  • the cells may be harvested prior to use in the screening assay. If HKAFE42 polypeptide is secreted into the medium, the medium can be recovered in order to recover and purify the polypeptide; if produced intracellularly, the cells must first be lysed before the polypeptide is recovered.
  • HKAFE42 polypeptides can be recovered and purified from recombinant cell cultures by well- known methods including ammonium sulfate or ethanol precipitation, acid extraction, anion or cation exchange chromatography, phosphocellulose chromatography, hydrophobic interaction chromatography, affinity chromatography, hydroxylapatite chromatography and lectin chromatography. Most preferably, high performance liquid chromatography is employed for purification. Well known techniques for refolding proteins may be employed to regenerate active conformation when the polypeptide is denatured during isolation and or purification.
  • This invention also relates to the use of HKAFE42 polynucleotides for use as diagnostic reagents. Detection of a mutated form of HKAFE42 gene associated with a dysfunction will provide a diagnostic tool that can add to or define a diagnosis of a disease or susceptibility to a disease which results from under-expression, over-expression or altered expression of HKAFE42. Individuals carrying mutations in the HKAFE42 gene may be detected at the DNA level by a variety of techniques.
  • Nucleic acids for diagnosis may be obtained from a subject's cells, such as from blood, urine, saliva, tissue biopsy or autopsy material.
  • the genomic DNA may be used directly for detection or may be amplified enzymatically by using PCR or other amplification techniques prior to analysis.
  • RNA or cDNA may also be used in similar fashion. Deletions and insertions can be detected by a change in size of the amplified product in comparison to the normal genotype. Point mutations can be identified by hybridizing amplified DNA to labeled HKAFE42 nucleotide sequences. Perfectly matched sequences can be distinguished from mismatched duplexes by RNase digestion or by differences in melting temperatures.
  • DNA sequence differences may also be detected by alterations in electrophoretic mobility of DNA fragments in gels, with or without denaturing agents, or by direct DNA sequencing. See, e.g., Myers et al, Science (1985) 230: 1242. Sequence changes at specific locations may also be revealed by nuclease protection assays, such as RNase and SI protection or the chemical cleavage method. See Cotton et al, Proc Natl Acad Sci USA (1985) 85: 4397-4401.
  • an array of oligonucleotides probes comprising HKAFE42 nucleotide sequence or fragments thereof can be constructed to conduct efficient screening of e.g., genetic mutations.
  • Array technology methods are well known and have general applicability and can be used to address a variety of questions in molecular genetics including gene expression, genetic linkage, and genetic variability. (See for example: M.Chee et al, Science, Vol 274, pp 610-613 (1996)).
  • the diagnostic assays offer a process for diagnosing or determining a susceptibility to disorders of the central nervous system (including anxiety, depression, schizophrenia and Parkinson's disease), muscle diseases (including muscular dystrophies and multiple sclerosis), Alzheimer's disease, hematopoetic disorders, allergic diseases, autoimmune diseases, immunodeficiency and cancer through detection of mutation in the HKAFE42 gene by the methods described.
  • disorders of the central nervous system including anxiety, depression, schizophrenia and Parkinson's disease
  • muscle diseases including muscular dystrophies and multiple sclerosis
  • Alzheimer's disease hematopoetic disorders
  • allergic diseases autoimmune diseases
  • immunodeficiency and cancer can be diagnosed by methods comprising determining from a sample derived from a subject an abnormally decreased or increased level of HKAFE42 polypeptide or HKAFE42 mRNA. Decreased or increased expression can be measured at the RNA level using any of the methods well known in the art for the quantitation of polynucleotides, such as, for example, PCR, RT-PCR, RNase protection, Northern blotting and other hybridization methods.
  • Assay techniques that can be used to determine levels of a protein, such as an HKAFE42 polypeptide, in a sample derived from a host are well-known to those of skill in the art. Such assay methods include radioimmunoassays, competitive-binding assays, Western Blot analysis and ELISA assays.
  • the present invention relates to a diagonostic kit for a disease or suspectability to a disease, particularly disorders of the central nervous system (including anxiety, depression, schizophrenia and Parkinson's disease), muscle diseases (including muscular dystrophies and multiple sclerosis), Alzheimer's disease, hematopoetic disorders, allergic diseases, autoimmune diseases, immunodeficiency and cancer, which comprises:
  • HKAFE42 polynucleotide preferably the nucleotide sequence of SEQ ID NO: 1, or a fragment thereof ;
  • HKAFE42 polypeptide preferably the polypeptide of SEQ ID NO: 2, or a fragment thereof;
  • kits an antibody to a HKAFE42 polypeptide, preferably to the polypeptide of SEQ ID NO: 2. It will be appreciated that in any such kit, (a), (b), (c) or (d) may comprise a substantial component.
  • the nucleotide sequences of the present invention are also valuable for chromosome identification.
  • the sequence is specifically targeted to and can hybridize with a particular location on an individual human chromosome.
  • the mapping of relevant sequences to chromosomes according to the present invention is an important first step in correlating those sequences with gene associated disease. Once a sequence has been mapped to a precise chromosomal location, the physical position of the sequence on the chromosome can be correlated with genetic map data. Such data are found, for example, in V. McKusick, Mendelian Inheritance in Man (available on line through Johns Hopkins University Welch Medical Library).
  • genes and diseases that have been mapped to the same chromosomal region are then identified through linkage analysis (coinheritance of physically adjacent genes).
  • linkage analysis coinheritance of physically adjacent genes.
  • the differences in the cDNA or genomic sequence between affected and unaffected individuals can also be determined. If a mutation is observed in some or all of the affected individuals but not in any normal individuals, then the mutation is likely to be the causative agent of the disease.
  • the HKAFE42 gene has been mapped to chromosome 7q31.1-2.
  • polypeptides of the invention or their fragments or analogs thereof, or cells expressing them can also be used as immunogens to produce antibodies immunospecific for the HKAFE42 polypeptides.
  • immunospecific means that the antibodies have substantiall greater affinity for the polypeptides of the invention than their affinity for other related polypeptides in the prior art.
  • Antibodies generated against the HKAFE42 polypeptides can be obtained by administering the polypeptides or epitope-bearing fragments, analogs or cells to an animal, preferably a nonhuman, using routine protocols.
  • any technique which provides antibodies produced by continuous cell line cultures can be used. Examples include the hybridoma technique (Kohler, G. and Milstein, C, Nature (1975) 256:495-497), the trioma technique, the human B-cell hybridoma technique (Kozbor et al, Immunology Today (1983) 4:72) and the EBV-hybridoma technique (Cole et al, MONOCLONAL ANTIBODIES AND CANCER THERAPY, pp. 77-96, Alan R. Liss, Inc., 1985).
  • the above-described antibodies may be employed to isolate or to identify clones expressing the polypeptide or to purify the polypeptides by affinity chromatography.
  • Antibodies against HKAFE42 polypeptides may also be employed to treat disorders of the central nervous system (including anxiety, depression, schizophrenia and Parkinson's disease), muscle diseases (including muscular dystrophies and multiple sclerosis), Alzheimer's disease, hematopoetic disorders, allergic diseases, autoimmune diseases, immunodeficiency and cancer, among others.
  • Vaccines include anxiety, depression, schizophrenia and Parkinson's disease, muscle diseases (including muscular dystrophies and multiple sclerosis), Alzheimer's disease, hematopoetic disorders, allergic diseases, autoimmune diseases, immunodeficiency and cancer, among others.
  • Another aspect of the invention relates to a method for inducing an immunological response in a mammal which comprises inoculating the mammal with HKAFE42 polypeptide, or a fragment thereof, adequate to produce antibody and/or T cell immune response to protect said animal from disorders of the central nervous system (including anxiety, depression, schizophrenia and Parkinson's disease), muscle diseases (including muscular dystrophies and multiple sclerosis), Alzheimer's disease, hematopoetic disorders, allergic diseases, autoimmune diseases, immunodeficiency and cancer, among others.
  • disorders of the central nervous system including anxiety, depression, schizophrenia and Parkinson's disease
  • muscle diseases including muscular dystrophies and multiple sclerosis
  • Alzheimer's disease including hematopoetic disorders
  • allergic diseases autoimmune diseases
  • immunodeficiency and cancer among others.
  • Yet another aspect of the invention relates to a method of inducing immunological response in a mammal which comprises, delivering HKAFE42 polypeptide via a vector directing expression of HKAFE42 polynucleotide in vivo in order to induce such an immunological response to produce antibody to protect said animal from diseases.
  • composition which, when introduced into a mammalian host, induces an immunological response in that mammal to a HKAFE42 polypeptide wherein the composition comprises a HKAFE42 polypeptide or HKAFE42 gene.
  • the vaccine formulation may further comprise a suitable carrier. Since HKAFE42 polypeptide may be broken down in the stomach, it is preferably administered parenterally (including subcutaneous, intramuscular, intravenous, intradermal etc. injection).
  • Formulations suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation instonic with the blood of the recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents or thickening agents.
  • the formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampoules and vials and may be stored in a freeze-dried condition requiring only the addition of the sterile liquid carrier immediately prior to use.
  • the vaccine formulation may also include adjuvant systems for enhancing the immunogenicity of the formulation, such as oil- in water systems and other systems known in the art. The dosage will depend on the specific activity of the vaccine and can be readily determined by routine experimentation.
  • the HKAFE42 polypeptide of the present invention may be employed in a screening process for compounds which activate (agonists) or inhibit activation of (antagonists, or otherwise called inhibitors) the HKAFE42 polypeptide of the present invention.
  • polypeptides of the invention may also be used to assess identify agonist or antagonists from, for example, cells, cell- free preparations, chemical libraries, and natural product mixtures.
  • agonists or antagonists may be natural or modified substrates, ligands, enzymes, receptors, etc, as the case may be, of the polypeptide of the present invention; or may be structural or functional mimetics of the polypeptide of the present invention. See Coligan et al, Current Protocols in Immunology l(2):Chapter 5 (1991).
  • HKAFE42 polypeptides are responsible for many biological functions, including many pathologies. Accordingly, it is desirous to find compounds and drugs which stimulate HKAFE42 polypeptide on the one hand and which can inhibit the function of HKAFE42 polypeptide on the other hand.
  • agonists are employed for therapeutic and prophylactic purposes for such conditions as disorders of the central nervous system (including anxiety, depression, schizophrenia and Parkinson's disease), muscle diseases (including muscular dystrophies and multiple sclerosis), Alzheimer's disease, hematopoetic disorders, allergic diseases, autoimmune diseases, immunodeficiency and cancer.
  • Antagonists may be employed for a variety of therapeutic and prophylactic purposes for such conditions as disorders of the central nervous system (including anxiety, depression, schizophrenia and Parkinson's disease), muscle diseases (including muscular dystrophies and multiple sclerosis), Alzheimer's disease, hematopoetic disorders, allergic diseases, autoimmune diseases, immunodeficiency and cancer.
  • disorders of the central nervous system including anxiety, depression, schizophrenia and Parkinson's disease
  • muscle diseases including muscular dystrophies and multiple sclerosis
  • Alzheimer's disease including muscular dystrophies and multiple sclerosis
  • hematopoetic disorders including allergic diseases, autoimmune diseases, immunodeficiency and cancer.
  • such screening procedures may involve using appropriate cells which express the HKAFE42 polypeptide or respond to HKAFE42 polypeptide of the present invention.
  • Such cells include cells from mammals, yeast, Drosophila or E. coli.
  • Cells which express the HKAFE42 polypeptide (or cell membrane containing the expressed polypeptide) or respond to HKAFE42 polypeptide are then contacted with a test compound to observe binding, or stimulation or inhibition of a functional response.
  • the ability of the cells which were contacted with the candidate compounds is compared with the same cells which were not contacted for HKAFE42 activity.
  • the assays may simply test binding of a candidate compound wherein adherence to the cells bearing the HKAFE42 polypeptide is detected by means of a label directly or indirectly associated with the candidate compound or in an assay involving competition with a labeled competitor. Further, these assays may test whether the candidate compound results in a signal generated by activation of the HKAFE42 polypeptide, using detection systems appropriate to the cells bearing the HKAFE42 polypeptide. Inhibitors of activation are generally assayed in the presence of a known agonist and the effect on activation by the agonist by the presence of the candidate compound is observed.
  • the assays may simply comprise the steps of mixing a candidate compound with a solution containing a HKAFE42 polypeptide to form a mixture, measuring HKAFE42 activity in the mixture, and comparing the HKAFE42 activity of the mixture to a standard.
  • the HKAFE42 cDNA, protein and antibodies to the protein may also be used to configure assays for detecting the effect of added compounds on the production of HKAFE42 mRNA and protein in cells.
  • an ELISA may be constructed for measuring secreted or cell associated levels of HKAFE42 protein using monoclonal and polyclonal antibodies by standard methods known in the art, and this can be used to discover agents which may inhibit or enhance the production of HKAFE42 (also called antagonist or agonist, respectively) from suitably manipulated cells or tissues.
  • the HKAFE42 protein may be used to identify membrane bound or soluble receptors, if any, through standard receptor binding techniques known in the art. These include, but are not limited to, ligand binding and crosslinking assays in which the HKAFE42 is labeled with a radioactive isotope (eg 1251), chemically modified (eg biotinylated), or fused to a peptide sequence suitable for detection or purification, and incubated with a source of the putative receptor (cells, cell membranes, cell supernatants, tissue extracts, bodily fluids). Other methods include biophysical techniques such as surface plasmon resonance and spectroscopy.
  • a radioactive isotope eg 1251
  • chemically modified eg biotinylated
  • these binding assays can be used to identify agonists and antagonists of HKAFE42 which compete with the binding of HKAFE42 to its receptors, if any. Standard methods for conducting screening assays are well understood in the art.
  • HKAFE42 polypeptide antagonists include antibodies or, in some cases, oligonucleotides or proteins which are closely related to the ligands, substrates, enzymes, receptors, etc, as the case may be, of the HKAFE42 polypeptide, e.g., a fragment of the ligands, substrates, enzymes, receptors, etc.; or small molecules which bind to the polypeptide of the present invention but do not elicit a response, so that the activity of the polypeptide is prevented.
  • the present invention relates to a screening kit for identifying agonists, antagonists, ligands, receptors, substrates, enzymes, etc. for HKAFE42 polypeptides; or compounds which decrease or enhance the production of HKAFE42 polypeptides, which comprises:
  • HKAFE42 polypeptide preferably that of SEQ ID NO:2;
  • This invention provides methods of treating abnormal conditions such as, disorders of the central nervous system (including anxiety, depression, schizophrenia and Parkinson's disease), muscle diseases (including muscular dystrophies and multiple sclerosis), Alzheimer's disease, hematopoetic disorders, allergic diseases, autoimmune diseases, immunodeficiency and cancer, related to both an excess of and insufficient amounts of HKAFE42 polypeptide activity.
  • disorders of the central nervous system including anxiety, depression, schizophrenia and Parkinson's disease
  • muscle diseases including muscular dystrophies and multiple sclerosis
  • Alzheimer's disease including hematopoetic disorders
  • allergic diseases autoimmune diseases
  • immunodeficiency and cancer related to both an excess of and insufficient amounts of HKAFE42 polypeptide activity.
  • HKAFE42 polypeptide is in excess, several approaches are available.
  • One approach comprises administering to a subject an inhibitor compound (antagonist) as hereinabove described along with a pharmaceutically acceptable carrier in an amount effective to inhibit the function of the HKAFE42 polypeptide, such as, for example, by blocking the binding of ligands, substrates, enzymes, receptors, etc, or by inhibiting a second signal, and thereby alleviating the abnormal condition.
  • soluble forms of HKAFE42 polypeptides still capable of binding the ligand, substrate, enzymes, receptors, etc. in competition with endogenous HKAFE42 polypeptide may be administered. Typical embodiments of such competitors comprise fragments of the HKAFE42 polypeptide.
  • HKAFE42 polypeptides still capable of binding the ligand in competition with endogenous HKAFE42 polypeptide may be administered.
  • Typical embodiments of such competitors comprise fragments of the HKAFE42 polypeptide.
  • expression of the gene encoding endogenous HKAFE42 polypeptide can be inhibited using expression blocking techniques.
  • Known such techniques involve the use of antisense sequences, either internally generated or separately administered. See, for example, O'Connor, J Neurochem (1991) 56:560 in Oligodeoxynucleotides as Antisense Inhibitors of Gene Expression. CRC Press, Boca Raton, FL (1988).
  • oligonucleotides which form triple helices with the gene can be supplied. See, for example, Lee et al, Nucleic Acids Res (1979) 3:173; Cooney et al, Science (1988) 241 :456; Dervan et al, Science (1991) 251 : 1360. These oligomers can be administered per se or the relevant oligomers can be expressed in vivo.
  • HKAFE42 For treating abnormal conditions related to an under-expression of HKAFE42 and its activity, several approaches are also available.
  • One approach comprises administering to a subject a therapeutically effective amount of a compound which activates HKAFE42 polypeptide, i.e., an agonist as described above, in combination with a pharmaceutically acceptable carrier, to thereby alleviate the abnormal condition.
  • gene therapy may be employed to effect the endogenous production of HKAFE42 by the relevant cells in the subject.
  • a polynucleotide of the invention may be engineered for expression in a replication defective retroviral vector, as discussed above.
  • the retroviral expression construct may then be isolated and introduced into a packaging cell transduced with a retroviral plasmid vector containing RNA encoding a polypeptide of the present invention such that the packaging cell now produces infectious viral particles containing the gene of interest.
  • These producer cells may be administered to a subject for engineering cells in vivo and expression of the polypeptide in vivo.
  • Another approach is to administer a therapeutic amount of HKAFE42 polypeptides in combination with a suitable pharmaceutical carrier.
  • Peptides such as the soluble form of HKAFE42 polypeptides, and agonists and antagonist peptides or small molecules, may be formulated in combination with a suitable pharmaceutical carrier.
  • suitable pharmaceutical carrier include but are not limited to, saline, buffered saline, dextrose, water, glycerol, ethanol, and combinations thereof. Formulation should suit the mode of administration, and is well within the skill of the art.
  • the invention further relates to pharmaceutical packs and kits comprising one or more containers filled with one or more of the ingredients of the aforementioned compositions of the invention.
  • Polypeptides and other compounds of the present invention may be employed alone or in conjunction with other compounds, such as therapeutic compounds.
  • systemic administration of the pharmaceutical compositions include injection, typically by intravenous injection.
  • Other injection routes such as subcutaneous, intramuscular, or intraperitoneal, can be used.
  • Alternative means for systemic administration include transmucosal and transdermal administration using penetrants such as bile salts or fusidic acids or other detergents.
  • penetrants such as bile salts or fusidic acids or other detergents.
  • oral administration may also be possible.
  • Administration of these compounds may also be topical and/or localized, in the form of salves, pastes, gels and the like.
  • the dosage range required depends on the choice of peptide, the route of administration, the nature of the formulation, the nature of the subject's condition, and the judgment of the attending practitioner. Suitable dosages, however, are in the range of 0.1-100 ⁇ g/kg of subject. Wide variations in the needed dosage, however, are to be expected in view of the variety of compounds available and the differing efficiencies of various routes of administration. For example, oral administration would be expected to require higher dosages than administration by intravenous injection. Variations in these dosage levels can be adjusted using standard empirical routines for optimization, as is well understood in the art.
  • Polypeptides used in treatment can also be generated endogenously in the subject, in treatment modalities often referred to as "gene therapy" as described above.
  • cells from a subject may be engineered with a polynucleotide, such as a DNA or RNA, to encode a polypeptide ex vivo, and for example, by the use of a retroviral plasmid vector. The cells are then introduced into the subject.
  • a polynucleotide such as a DNA or RNA
  • a nucleotide sequence of a human HKAFE42. A nucleotide sequence of a human HKAFE42..
  • 165 170 175 lie Cys Asp Gly Ser Ala Ser Met Gin Ala Arg Gin Thr Cys Ala Thr
  • Lys Glu Lys Asp Thr Thr Val lie Cys Ser Thr Pro Asn Thr Val Leu 225 230 235 240
  • 405 410 415 lie Ser Arg Phe Glu Arg His Leu Tyr Asn Ser Ala Ala Phe Lys Ala

Landscapes

  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Organic Chemistry (AREA)
  • Medicinal Chemistry (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Zoology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Toxicology (AREA)
  • Peptides Or Proteins (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

L'invention concerne les polypeptides HKAFE42 et des polynucléotides ainsi que des procédés de production de ces polypeptides par génie génétique. L'invention concerne également des méthodes d'utilisation des polypeptides HKAFE42 et des polynucléotides dans la conception de protocoles de traitement de troubles du système nerveux central (notamment de l'anxiété, de la dépression, de la schizophrénie et de la maladie de Parkinson), de maladies musculaires (notamment des myopathies primitives progressives et la sclérose en plaque), de la maladie d'Alzheimer, de troubles hématopoïétiques, de maladies allergiques, de maladies auto-immunes, dans le traitement de l'immunodéficience et du cancer, entre autres, et dans des méthodes de diagnostic de ces états.
PCT/GB1998/000915 1997-03-25 1998-03-25 Proteine hkafe42 apparentee a l'interferon WO1998042841A1 (fr)

Priority Applications (2)

Application Number Priority Date Filing Date Title
JP10545249A JP2000513947A (ja) 1997-03-25 1998-03-25 インターフェロン関連蛋白ｈｋａｆｅ４２
EP98913911A EP0934414A1 (fr) 1997-03-25 1998-03-25 Proteine hkafe42 apparentee a l'interferon

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
GBGB9706178.2A GB9706178D0 (en) 1997-03-25 1997-03-25 Novel compound
GB9706178.2 1997-03-25
EP97308921 1997-11-06
EP97308921.2 1997-11-06

Publications (1)

Publication Number Publication Date
WO1998042841A1 true WO1998042841A1 (fr) 1998-10-01

Family

ID=26147688

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/GB1998/000915 WO1998042841A1 (fr) 1997-03-25 1998-03-25 Proteine hkafe42 apparentee a l'interferon

Country Status (4)

Country Link
EP (1) EP0934414A1 (fr)
JP (1) JP2000513947A (fr)
CA (1) CA2247075A1 (fr)
WO (1) WO1998042841A1 (fr)

Non-Patent Citations (8)

* Cited by examiner, † Cited by third party
Title
B.C. VARNUM ET AL: "Characterization of TIS7, a gene indiced in Swiss 3T3 cells by the tumor promoter tetradecanoyl phorbol acetate", ONCOGENE, vol. 4, no. 10, October 1989 (1989-10-01), pages 1263 - 1265, XP002073833 *
D. GUARDAVACCARO ET AL: "Inhibition of differentiation in myoblasts deprived of the Interferon-related protein PC4", CELL GROWTH AND DIFFERENTIATION, vol. 6, no. 2, February 1995 (1995-02-01), pages 159 - 169, XP002073832 *
D. GUARDAVACCARO ET AL: "Nerve Growth Factor regulates the subcellular localization of the nerve growth Factor-inducible protein PC4 in PC12 cells", JOURNAL OF NEUROSCIENCE RESEARCH, vol. 37, no. 5, 1 April 1994 (1994-04-01), pages 660 - 674, XP002073831 *
F. LATIF ET AL: "The human homolog of the rodent immediate early response genes, PC4 and TIS7, resides in the lung cancer tumor suppressor gene region on chromosome 3q21", HUMAN GENETICS, vol. 99, no. 3, 17 February 1997 (1997-02-17), pages 334 - 341, XP002073836 *
F. TIRONE ET AL: "Early gene regulation by nerve growth factor in PC12 cells: Induction of an interferon-related gene", PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF USA., vol. 86, March 1989 (1989-03-01), WASHINGTON US, pages 2088 - 2092, XP002073830 *
L. HILLIER ET AL: "The WashU-Merck EST project. zh68f09.s1 Soares fetal liver spleen 1NFLS S1 Homo sapiens cDNA clone 417257 3' similar to SW: PC4- mouse P19182 Interferon-related protein PC4", EMBL DATABASE ENTRY HSW87947, ACCESSION NUMBER W87947, 5 July 1996 (1996-07-05), XP002073834 *
L. HILLIER ET AL: "The WashU-NCI human EST project. zq0g12.r1 Stratagene neuroepithelium (#937231) Homo sapiens cDNA clone 645094 5' similar to SW: PC4-rat P20695 Interferon-related protein PC4.", EMBL DATABASE ENTRY HSAA9385, ACCESSION NUMBER AA209385, 1 February 1997 (1997-02-01), XP002073835 *
P. BUANNE ET AL: "H. sapiens mRNA for nerve Growth Factor-inducible PC4 homologue", EMBL DATABASE ENTRY HSNGIPC4, ACCESSION NUMBER Y10313, 2 June 1997 (1997-06-02), XP002073837 *

Also Published As

Publication number Publication date
EP0934414A1 (fr) 1999-08-11
CA2247075A1 (fr) 1998-10-01
JP2000513947A (ja) 2000-10-24

Similar Documents

Publication Publication Date Title
EP0855404B1 (fr) Antagoniste beta d'un recepteur de l'interleukine-1 (il-1rabeta)
WO1999021991A1 (fr) Bmzf12: gene a doigt de zinc clone a partir de la moelle osseuse
WO1999036541A1 (fr) ANTAGONISTE DU RECEPTEUR DE L'INTERLEUKINE-1 BETA (IL-1RAβ)
EP0897002A2 (fr) U62317, une protéine avec un domaine se liant à JNK
WO1999046293A1 (fr) Proteine a doigt de zinc derivee de cellules hematopoietiques
EP0879889A2 (fr) Un membre de la famille de l'interleukine-1,iL-1 delta
EP0953638A1 (fr) Canal cationique d'origine humaine homologue au récepteur vanilloide
US5879908A (en) CRFG-1a, a target and marker for chronic renal failure
EP0908515A2 (fr) Polypeptide pancréatique
EP0894855A2 (fr) Facteur de transcription I kappa B-beta humain
US6187560B1 (en) Polynucleotides and polypeptides belonging to the uncoupling proteins family
EP0894856A1 (fr) Variante d'épissure de sMAD3 humaine
WO1999046290A1 (fr) Gene humain de type recepteur d'angiotensine ii/vasopreessine (aii/avp) (cbdakd01)
EP0934414A1 (fr) Proteine hkafe42 apparentee a l'interferon
WO1999022006A1 (fr) Cblafc02: sous-unite de h(+)-atpase vacuolaire
EP0890578A2 (fr) Polynucleotides codant le géne d'horloge humaine et leur utilisation
WO1999021988A1 (fr) Gene sec22b humain de cbfbba01 de la proteine d'echange vesiculaire
US20020019520A1 (en) CBFBGA09: a human SL15 homolog
EP0879886A2 (fr) HLDAT86, Protéine de la transduction de signaux et homoloque humaine de Wnt-4
EP1224220A2 (fr) Gene 1 de reponse primaire a l'erythropoietine (epo), gene eprg1
US6255471B1 (en) CRFG-1b, a target and marker for chronic renal failure
WO1999021885A1 (fr) Gene transporteur 7 abc humain (habc7)
WO1999021982A1 (fr) Gene m6b1 humain
WO1999046291A1 (fr) Gene homologue humain du facteur de maturation des cellules gliales (gmf) type beta (cbfboe11)
WO1999022007A1 (fr) Cbfaie10: gene skd1 humain

Legal Events

Date Code Title Description
ENP Entry into the national phase

Ref document number: 2247075

Country of ref document: CA

Ref country code: CA

Ref document number: 2247075

Kind code of ref document: A

Format of ref document f/p: F

WWE Wipo information: entry into national phase

Ref document number: 1998913911

Country of ref document: EP

AK Designated states

Kind code of ref document: A1

Designated state(s): CA JP US

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): AT BE CH DE DK ES FI FR GB GR IE IT LU MC NL PT SE

121 Ep: the epo has been informed by wipo that ep was designated in this application
WWP Wipo information: published in national office

Ref document number: 1998913911

Country of ref document: EP

WWE Wipo information: entry into national phase

Ref document number: 09125239

Country of ref document: US

WWW Wipo information: withdrawn in national office

Ref document number: 1998913911

Country of ref document: EP

点击 这是indexloc提供的php浏览器服务，不要输入任何密码和下载