WO2001077393A2

WO2001077393A2 - Methods for detecting neurological disorders

Info

Publication number: WO2001077393A2
Application number: PCT/US2001/002177
Authority: WO
Inventors: Theodore E. Whitmore; Paul O. Sheppard
Original assignee: Zymogenetics, Inc.
Priority date: 2000-04-10
Filing date: 2001-01-23
Publication date: 2001-10-18
Also published as: AU2001234523A1; WO2001077393A3

Abstract

The present invention relates to polynucleotide and polypeptide molecules for ZTMPO-1, protein with a degree of homology to emerin and the thymopoietins. Human ZTMPO-1 maps to 12q24.3. The polypeptides, and polynucleotides encoding them are useful for diagnostic applications for neurological diseases such as Alzheimer's Disease.

Description

DESCRIPTION METHODS FOR DETECTING NEUROLOGICAL DISORDERS

BACKGROUND OF THE INVENTION

Alzheimer's Disease is the most common cause of degenerative dementia and the number of people suffering continues to increase the population ages. Alzheimer's Disease is a progressive degenerative disease that begins with widespread cognitive effects, including memory loss, shortened attention span, and disorientation, that develops over a number of years ultimately resulting in dementia. Pathologically, Alzheimer's is characterized by neurofibrillary tangles in the cerebral cortex and hippocampus, together with deposition of amyloid within the senile plaques and cerebral blood vessels.

Several loci have been identified as contributing to Alzheimer's Disease, including amyloid precursor protein gene (Goate et al., Nature 349:704-6, 1991), presenilin 1 gene (Herrington et al., Nature 375:754-60. 1995), presenilin 2 gene (Rogaev et al., Nature 376:755-8, 1995), which are associated with early-onset autosomal dominate Alzheimer's Disease, and the apolipoprotein E gene (Pericak-

Nance et al., Am. J. Hum. Genet. 48:1034-50, 1991) which is associated with early- onset sporadic Alzheimer's Disease. Additionally, loci have been reported in the vicinity of the centromere on chromosome 12 (Pericak-Nance et al., JAMA 278: 1237- 41, 1997) and about 120 cM proximal near D12S 1045 at 12q24.3 (Zubenko et al., Genomics 50: 121-8, 1998).

ZTMPO-1 (Sheppard et al, WO 99/54468) a protein having some homology to a family of nuclear membrane proteins that includes emerin and the thymopoietins maps -1.35 mb distal of D12S1045 at human chromosome 12q24.3 and would be useful as a marker for Alzheimer disease. The present invention provides for these and other uses that should be apparent to those skilled in the art from the teachings herein.

SUMMARY OF THE INVENTION

Within one aspect the invention provides a method of detecting a chromosome 12 abnormality in a subject comprising: (a) amplifying a polynucleotide from RNA isolated from a biological sample using primers derived from a polynucleotide of SEQ ID NO: 1 or its complement; and (b) detecting an aberration in the amplified polynucleotide by comparing the nucleic acid sequence of said amplified polynucleotide with the nucleic acid sequence of the polynucleotide of SEQ ED NO: 1 ; wherein the presence of a aberration indicates a chromosome 12 abnormality. Within one embodiment amplification is performed by polymerase chain reaction or reverse transcriptase-polymerase chain reaction.

The invention also provides a method of detecting a chromosome 12 abnormality in a subject comprising: (a) amplifying a polynucleotide from RNA isolated from a biological sample using primers derived from a polynucleotide of SEQ ID NO:l or its complement; (b) transcribing the amplified polynucleotide to express mRNA; (c) translating said mRNA to produce a polypeptide; and (d) detecting an aberration in the amino acid sequence of said transcribed polypeptide by comparison to the amino acid sequence of SEQ ID NO:2; wherein the presence of an aberration indicates a chromosome 12 abnormality.

Within another aspect the invention provides a method for diagnosing a neurological disease or susceptibility to a neurological disease in an individual, wherein said disease is related to the expression or activity of a polypeptide of SEQ ID NO:2 or fragment thereof, in said individual, comprising the step of: determining the presence of an alteration in the nucleic acid sequence of a polynucleotide encoding said polypeptide in the genome of said individual by comparison of the nucleic acid sequence of said polynucleotide with the nucleic acid sequence of SEQ ID NO: l; wherein the presence of an aberration in said nucleic acid sequence indicates a neurological disease or susceptibility to a neurological disease. Within one embodiment the neurological disease is Alzheimer's Disease.

The invention also provides a method for diagnosing a neurological disease or susceptibility to a neurological disease in an individual, comprising: (a) amplifying a polynucleotide from RNA isolated from a biological sample using primers derived from a polynucleotide of SEQ ID NO: 1 or its complement; and (b) detecting an aberration in the amplified polynucleotide by comparing the nucleic acid sequence of said amplified polynucleotide with the nucleic acid sequence of the polynucleotide of SEQ ID NO: 1 ; wherein the presence of an aberration indicates a neurological disease or susceptibility to a neurological disease. Within one embodiment the neurological disease is Alzheimer's Disease.

Within another aspect the invention provides a method for diagnosing a neurological disease or susceptibility to a neurological disease in an individual, comprising: (a) amplifying a polynucleotide from RNA isolated from a biological sample using primers derived from a polynucleotide of SEQ ID NO: l or its complement; (b) transcribing the amplified polynucleotide to express mRNA; (c) translating said mRNA to produce a polypeptide; and (d) detecting an aberration in the amino acid sequence of said transcribed polypeptide by comparison to the amino acid sequence of SEQ ID NO:2; wherein the presence of a mutation indicates a neurological disease or susceptibility to a neurological disease. Within one embodiment the neurological disease is Alzheimer's Disease.

Within another aspect the invention provides a method for diagnosing a neurological disease or susceptibility to a neurological disease in an individual, comprising: (a) contacting a polynucleotide probe with a biological sample under hybridizing conditions; wherein said polynucleotide probe is derived from a polynucleotide of SEQ ID NO: l or its complement; and (b) detecting the formation of a hybrid between said polynucleotide probe and said biological sample; wherein the presence of said hybrid indicates the presence of a ZTMPO-1 polynucleotide in said biological sample.

The invention also provides a method of detecting the presence of a polypeptide of SEQ ID NO:2, or a fragment thereof, in a biological sample, comprising the steps of: (a) adding an antibody, or an antibody fragment, that specifically binds with a polypeptide of SEQ ID NO:2 to said biological sample under conditions that allow the binding of said antibody or antibody fragment to said biological sample; (b) detecting any of said bound antibody or bound antibody fragment; and (c) correlating the presence of a polypeptide of SEQ ID NO:2, or a fragment thereof, with a neurological disease or susceptibility to a neurological disease. Within one embodiment the antibody or said antibody fragment further comprises a detectable label selected from the group consisting of radioisotope, fluorescent label, chemiluminescent label, enzyme label, bioluminescent label, and colloidal gold. The invention also provides a kit for the detection of a gene encoding a polypeptide of SEQ ID NO:2, comprising: a first container that comprises a polynucleotide of SEQ ID NO:l or its complement; and a second container that comprises one or more reagents capable of indicating the presence of said polynucleotide. The invention further provides a kit for the detection of a gene encoding a polypeptide of SEQ ID NO:2, comprising: a first container comprising an antibody that specifically binds to a polypeptide of SEQ ID NO:2; and a second container that comprises one or more reagents capable of indicating the presence of said antibody.

DETAILED DESCRIPTION OF THE INVENTION

As used herein, polynucleotide denotes a single- or double-stranded polymer of deoxyribonucleotide or ribonucleotide bases read from the 5' to the 3' end. Polynucleotides include RNA and DNA, and may be isolated from natural sources, synthesized in vitro, or prepared from a combination of natural and synthetic molecules. DNA includes cDNA and genomic DNA. Sizes of polynucleotides are expressed as base pairs (abbreviated "bp"), nucleotides ("nt"), or kilobases ("kb"). Where the context allows, the latter two terms may describe polynucleotides that are single- stranded or double-stranded. When the term is applied to double-stranded molecules it is used to denote overall length and will be understood to be equivalent to the term "base pairs". It will be recognized by those skilled in the art that the two strands of a double-stranded polynucleotide may differ slightly in length and that the ends thereof may be staggered as a result of enzymatic cleavage; thus all nucleotides within a double-stranded polynucleotide molecule may not be paired. Such unpaired ends will in general not exceed 20 nt in length.

Polypeptide is a polymer of amino acid residues joined by peptide bonds, whether produced naturally or synthetically. Polypeptides of less than about 10 amino acid residues are commonly referred to as "peptides".

Protein is a macromolecule comprising one or more polypeptide chains. A protein may also comprise non-peptidic components, such as carbohydrate groups. Carbohydrates and other non-peptidic substituents may be added to a protein by the cell in which the protein is produced, and will vary with the type of cell. Proteins are defined herein in terms of their amino acid backbone structures; substituents such as carbohydrate groups are generally not specified, but may be present nonetheless.

ZTMPO-1, a 2,754 bp polynucleotide which has an open reading frame encoding an 876 amino acid residue protein having regions of homology to a family of nuclear membrane proteins that includes emerin, the thymopoietins, and the lamina associated proteins (Zevin-Sonkin et al., Immuno. Letts. 31:301-10, 1992; Harris et al., Proc. Natl. Acad. Sci. USA 91:6283-87, 1994; Harris et al., Genomics 28: 198-205, 1995; Berger et al., Genome Res. 6:361-70, 1996 and Ishijima et al., Biochem. Biophys. Res. Comm. 226:431-8, 1996), (Senior and Gerace, J. Cell Biol. 107:2029-36, 1988; Worman et al., J. Cell Biol. I l l: 1535-42, 1990; Wozniak and Blobel J. Cell Biol. 119:1441-9, 1992; Foisner and Gerace, CeU 73:1267-79, 1993; Ye and Worman, J.

Biol. Chem. 269:11306-11, 1994 and Furukawa et al., EMBO J. 14:1626-36, 1995), (Bione et al., Nat. Genet. 8:323-7, 1994; Manual et al., Hum. Mol. Gen. 5:801-8, 1996 and Small et al., Mamm. Genom. 8:337-41, 1997).

The human "Z7 PO-7" nucleotide sequence is represented in SEQ ID NO: l and the deduced "ZTMPO-1" amino acid sequence in SEQ ID NO:2. Sequence analysis of the deduced amino acid sequence as represented in SEQ ID NO:2 does not indicate the presence of a secretion signal sequence or transmembrane domain. There is a putative ankyrin-like region, amino acid residues 333-385 of SEQ ID NO:2, having an ankyrin repeat (residues 347-379 of SEQ ID NO:2) which may indicate that ZTMPO-1 is retained in the plasma membrane. At the C-terminal end of ZTMPO-1 is a calcium binding protein-like region having two potential calcium binding sites (residues 678-692 and residues 719-731 of SEQ ID NO:2) similar to that seen in the sea urchin calcium binding protein LPSl-beta (Xiang et al., J. Biol. Chem. 16: 10524-33, 1991).

The ZTMPO-1 polynucleotide of SEQ ED NO: 1 encodes an 876 amino acid residue protein which is much larger than other members of the thymopoietin/emerin family. Human thymopoietin α is a 693 amino acid residue protein, human emerin is a 254 amino acid residue protein and rat LAP2 is a 452 amino acid residue protein.

Like emerin, the amino acid sequence of ZTMPO-1 does not contain the

42 amino acid thymopoietin peptide originally identified by Goldstein (Nature 247: 11 - 14, 1974) but shares discrete regions of homology with the human thymopoietins α, β and γ (Harris et al., ibid.) and the mouse thymopoietins α, β, γ, ε, δ, and ζ (Berger et al., ibid.).

As would be expected, ZTMPO-1 also shares discrete regions of homology with rat lamina associated protein 2, LAP2, (Furukawa et al., ibid.). These regions correspond to many of the same regions with which ZTMPO- 1 shares identity with the thymopoietins. ZTMPO-1 and rat LAP2 share 70% amino acid identity over the region corresponding to amino acid residues 13 to 44 of SEQ ID NO:2.

ZTMPO-1 also shares a limited degree of homology to regions of the yeast transcription factor HF alpha subunit over the region corresponding to amino acid residues 86 to 160 and amino acid residues 205 to 260 of SEQ ID NO:2.

Additionally, ZTMPO-2 shares 27% amino acid identity with Trypanosoma brucei ribonuclease HI (Hesslein and Campbell, Mol. Biochem. Parasitol. 86:121-6, 1997) over the region corresponding to amino acid residues 156 to 203 of SEQ ID NO:2. This homology, along with that shared with LAP2, as well as the possible ankyrin repeat, suggests the possibility that ZTMPO-1 possesses chromatin or

DNA binding properties.

Chromosomal localization results show that ZTMPO-1 maps in the 12q24.3 chromosomal region on the integrated LDB chromosome 12 map. An Alzheimer's Disease loci also exists at 12q24.3 in the vicinity of D12S1045 (Zubenko et al., Genomics 50:121-8, 1998). ZTMPO-1 maps -1.35 mb distal from that loci.

ZTMPO-1 polynucleotides and polypeptides may be used within diagnostic systems to detect ZTMPO-1 polynucleotides, including DNA or RNA, or ZTMPO-1 proteins and polypeptides in a biological sample and would serve as a diagnostic tool for diseases where altered levels of ZTMPO-1 polynucleotides, polypeptides, or proteins are significant. The information derived from such detection methods would provide insight into the significance of ZTMPO-1 in neurological diseases, such as Alzheimer's Disease.

For example, the ZTMPO-1 gene, a probe comprising ZTMPO-1 DNA or RNA, or a subsequence thereof can be used to determine if the ZTMPO-1 gene is present on chromosome 12 or if there is a chromosomal structural abnormality associated with that region. Chromosomal structural abnormalities include, but are not limited to, insertions, inversions, deletions, duplications, rearrangements, and translocations. ZTMPO-1 gene probes can also be used to detect aberrations and alterations including, but not limited to, aneuploidy, gene copy number changes, restriction site changes, and rearrangements.

Various additional diagnostic approaches are well-known to those of skill in the art (see, for example, Mathew (ed.), Protocols in Human Molecular Genetics Humana Press, Inc., 1991; Coleman and Tsongalis, Molecular Diagnostics, Humana Press, Inc., 1996 and Elles, Molecular Diagnosis of Genetic Diseases, Humana Press, Inc., 1996).

In a basic assay, a single-stranded probe molecule is incubated with RNA, isolated from a biological sample, under conditions of temperature and ionic strength that promote base pairing between the probe and target ZTMPO-1 RNA species. After separating unbound probe from hybridized molecules, the amount of hybrid molecules are determined.

Well-established hybridization methods of RNA detection include northern analysis and dot/slot blot hybridization (see, for example, Ausubel ibid, and Wu et al. (eds.), "Analysis of Gene Expression at the RNA Level," in Methods in Gene Biotechnology, pages 225-239 (CRC Press, Inc. 1997)). Nucleic acid probes can be detectably labeled with radioisotopes such as ³²P or ³⁵S. Alternatively, ZTMPO-1 RNA can be detected with a nonradioactive hybridization method (see, for example, Isaac (ed.), Protocols for Nucleic Acid Analysis by Nonradioactive Probes, Humana Press, Inc., 1993). Typically, nonradioactive detection is achieved by enzymatic conversion of chromogenic or chemiluminescent substrates. Illustrative nonradioactive moieties include biotin, fluorescein, and digoxigenin.

ZTMPO-1 oligonucleotide probes are also useful for in vivo diagnosis. As an illustration, ¹⁸F-labeled oligonucleotides can be administered to a subject and visualized by positron emission tomography (Tavitian et al, Nature Medicine 4:467, 1998). Numerous diagnostic procedures take advantage of the polymerase chain reaction (PCR) to increase sensitivity of detection methods. Standard techniques for performing PCR are well-known (see, generally, Mathew (ed.), Protocols in Human Molecular Genetics (Humana Press, Inc. 1991), White (ed.), PCR Protocols: Current Methods and Applications (Humana Press, Inc. 1993), Cotter (ed.), Molecular Diagnosis of Cancer (Humana Press, Inc. 1996), Hanausek and Walaszek (eds.), Tumor Marker Protocols (Humana Press, Inc. 1998), Lo (ed.), Clinical Applications of PCR (Humana Press, Inc. 1998), and Meltzer (ed.), PCR in Bioanalysis (Humana Press, Inc. 1998)). PCR primers can be designed to amplify a sequence encoding a particular ZTMPO-1 domain or region of homology as described herein.

One variation of PCR for diagnostic assays is reverse transcriptase-PCR (RT-PCR). In the RT-PCR technique, RNA is isolated from a biological sample, reverse transcribed to cDNA, and the cDNA is incubated with ZTMPO-1 primers (see, for example, Wu et al. (eds.), "Rapid Isolation of Specific cDNAs or Genes by PCR," in Methods in Gene Biotechnology, CRC Press, Inc., pages 15-28, 1997). PCR is then performed and the products are analyzed using standard techniques.

As an illustration, RNA is isolated from biological sample using, for example, the guanidinium-thiocyanate cell lysis procedure described above. Alternatively, a solid-phase technique can be used to isolate mRNA from a cell lysate. A reverse transcription reaction can be primed with the isolated RNA using random oligonucleotides, short homopolymers of dT, or ZTMPO-1 anti-sense oligomers. Oligo-dT primers offer the advantage that various mRNA nucleotide sequences are amplified that can provide control target sequences. ZTMPO-1 sequences are amplified by the polymerase chain reaction using two flanking oligonucleotide primers that are typically at least 5 bases in length.

PCR amplification products can be detected using a variety of approaches. For example, PCR products can be fractionated by gel electrophoresis, and visualized by ethidium bromide staining. Alternatively, fractionated PCR products can be transferred to a membrane, hybridized with a detectably-labeled ZTMPO-1 probe, and examined by autoradiography. Additional alternative approaches include the use of digoxigenin-labeled deoxyribonucleic acid triphosphates to provide chemiluminescence detection, and the C-TRAK colorimetric assay.

Another approach is real time quantitative PCR (Perkin-Elmer Cetus,

Norwalk, Ct.). A fluorogenic probe, consisting of an oligonucleotide with both a reporter and a quencher dye attached, anneals specifically between the forward and reverse primers. Using the 5' endonuclease activity of Taq DNA polymerase, the reporter dye is separated from the quencher dye and a sequence-specific signal is generated and increases as amplification increases. The fluorescence intensity can be continuously monitored and quantified during the PCR reaction.

Another approach for detection of ZTMPO-1 expression is cycling probe technology (CPT), in which a single-stranded DNA target binds with an excess of DNA-RNA-DNA chimeric probe to form a complex, the RNA portion is cleaved with RNase H, and the presence of cleaved chimeric probe is detected (see, for example, Beggs et al., J. Clin. Microbiol. 34:2985, 1996 and Bekkaoui et al., Biotechniques 20:240, 1996). Alternative methods for detection of ZTMPO-1 sequences can utilize approaches such as nucleic acid sequence-based amplification (NASBA), cooperative amplification of templates by cross-hybridization (CATCH), and the ligase chain reaction (LCR) (see, for example, Marshall et al., U.S. Patent No. 5,686,272 (1997), Dyer et al., J. Virol. Methods 60:161, 1996; Ehricht et al., Eur. J. Biochem. 243:358, 1997 and Chadwick et al., J. Virol. Methods 70:59, 1998). Other standard methods are known to those of skill in the art. ZTMPO-1 probes and primers can also be used to detect and to localize

ZTMPO-1 gene expression in tissue samples. Methods for such in situ hybridization are well-known to those of skill in the art (see, for example, Choo (ed.), In Situ Hybridization Protocols, Humana Press, Inc., 1994; Wu et al. (eds.), "Analysis of Cellular DNA or Abundance of mRNA by Radioactive In Situ Hybridization (RISH)," in Methods in Gene Biotechnology. CRC Press, Inc., pages 259-278, 1997 and Wu et al.

(eds.), "Localization of DNA or Abundance of mRNA by Fluorescence In Situ Hybridization (RISH)," in Methods in Gene Biotechnology, CRC Press, Inc., pages 279- 289, 1997).

Other suitable assay methods include restriction fragment length polymoφhism (RFLP) analysis, short tandem repeat (STR) analysis employing PCR techniques, ligation chain reaction (Barany, PCR Methods and Applications L5-16, 1991), ribonuclease protection assays, and other genetic linkage analysis techniques known in the art (Sambrook et al., ibid. ; Ausubel et. al., ibid. ; Marian, Chest 108:255- 65, 1995). Ribonuclease protection assays (see, e.g., Ausubel et al., ibid., ch. 4) comprise the hybridization of an RNA probe to a patient RNA sample, after which the reaction product (RNA-RNA hybrid) is exposed to RNase. Hybridized regions of the RNA are protected from digestion. Within PCR assays, a patient's genetic sample is incubated with a pair of polynucleotide primers, and the region between the primers is amplified and recovered. Changes in size or amount of recovered product are indicative of mutations in this gene. Another PCR-based technique that can be employed is single strand conformational polymoφhism (SSCP) analysis (Hayashi, PCR Methods and Applications 1:34-8, 1991). The invention provides diagnostic methods comprising the steps of (a) obtaining a biological sample from a patient; (b) incubating the biological sample with a ZTMPO-1 polynucleotide probe or primer, under conditions wherein the polynucleotide will hybridize to complementary polynucleotide sequence creating a hybridized biological sample; and (c) detecting the hybridized biological sample. Deletions, translocations, duplications, changes in size, or amount of the hybridized biological sample is indicative of a abnormality in the patient that can then be correlated with the incidence of a particular disease such as neurological diseases like Alzheimer's Disease. Biological samples include, but are not limited to, samples derived from or comprised of cells, cell components or cell products, including, but not limited to, cell culture supernatants, cell lysates, cleared cell lysates, cell extracts, tissue, tissue extracts, whole blood, plasma, serum, and fractions thereof; from a patient. Cell components or products include, but are not limited to, nucleotides, polypeptides, chromosomes, and proteins.

The invention provides isolated and purified ZTMPO-1 polynucleotide probes or primers for use in hybridization. Techniques for developing polynucleotide probes and hybridization techniques are known in the art, see for example, Ausubel et al., eds., Current Protocols in Molecular Biology, John Wiley and Sons, Inc., NY, 1991. Such polynucleotide probes can be RNA or DNA. DNA can be either cDNA or genomic DNA. Polynucleotide probes are single or double-stranded DNA or RNA, generally synthetic oligonucleotides, but may be generated from cloned cDNA or genomic sequences. Analytical probes will generally be at least 20 nucleotides in length, although somewhat shorter probes (14-17 nucleotides) can be used. PCR primers are at least 5 nucleotides in length, preferably 15 or more nucleotides, more preferably 20-30 nucleotides in length. Short polynucleotides can be used when a small region of the gene is targeted for analysis. For gross analysis of genes, a polynucleotide probe may comprise an entire exon or more.

Probes can be labeled to provide a detectable signal, such as with an enzyme, biotin, a radionuclide, fluorophore, chemiluminescer, paramagnetic particle and the like, which are commercially available from many sources, such as Molecular Probes, Inc., Eugene, OR, and Amersham Coφ., Arlington Heights, EL, using techniques that are well known in the art. Probes can be directly labeled by random priming, end labeling, PCR, or nick translation. Suitable direct labels include radioactive labels such as ³²P, ³H, and ³⁵S and non-radioactive labels such as fluorescent markers (e.g., fluorescein, Texas Red, AMCA blue (7-amino-4-methyl-coumanine-3- acetate), lucifer yellow, rhodamine, etc.), cyanin dyes which are detectable with visible light, enzymes, and the like. Probes labeled with an enzyme can be detected through a colorimetric reaction by providing a substrate for the enzyme. In the presence of various substrates, different colors are produced by the reaction, and these colors can be visualized to separately detect multiple probes if desired. The present invention also contemplates kits for performing a diagnostic assay for ZTMPO-1 gene expression or to detect mutations in the ZTMPO-1 gene. Such kits comprise nucleic acid probes, such as double-stranded nucleic acid molecules comprising the nucleotide sequence of SEQ ID NO:l, or a portion thereof, as well as single-stranded nucleic acid molecules having the complement of the nucleotide sequence of SEQ ED NO: l, or a portion thereof. Probe molecules may be DNA, RNA, oligonucleotides, and the like. Kits can comprise nucleic acid primers for performing PCR or oligonucleotides for performing the ligase chain reaction, in situ hybridizations and the like.

Preferably, such a kit contains all the necessary elements to perform diagnostic assays as described herein. A kit will comprise at least one container comprising a ZTMPO-1 probe or primer. The kit may also comprise a second container comprising one or more reagents capable of indicating the presence of ZTMPO-1 sequences. Examples of such indicator reagents include detectable labels such as radioactive labels, fluorochromes, chemiluminescent agents, and the like. A kit may also comprise a means for conveying to the user that the ZTMPO-1 probes and primers are used to detect ZTMPO-1 gene expression. For example, written instructions may state that the enclosed nucleic acid molecules can be used to detect either a nucleic acid molecule that encodes ZTMPO-1, or a nucleic acid molecule having a nucleotide sequence that is complementary to an ZTMPO- 1 -encoding nucleotide sequence. The written material can be applied directly to a container, or the written material can be provided in the form of a packaging insert.

Methods for preparing cDNA and genomic clones are well known and within the level of ordinary skill in the art, and include the use of the sequence disclosed herein, or parts thereof, for probing or priming a library. Additionally, the polynucleotides of the present invention can be synthesized using gene machines.

Currently the method of choice is the phosphoramidite method. If chemically synthesized double stranded DNA is required for an application such as the synthesis of a gene or a gene fragment, then each complementary strand is made separately. The production of short genes (60 to 80 bp) is technically straightforward and can be accomplished by synthesizing the complementary strands and then annealing them. For the production of longer genes (>300 bp), however, special strategies must be invoked, because the coupling efficiency of each cycle during chemical DNA synthesis is seldom 100%. To overcome this problem, synthetic genes (double-stranded) are assembled in modular form from single-stranded fragments that are from 20 to 100 nucleotides in length. See Glick, Bernard R. and Jack J. Pasternak, Molecular Biotechnology, Principles & Applications of Recombinant DNAJASM Press, Washington, D.C. 1994), Itakura, K. et al. Synthesis and use of synthetic oligonucleotides. Annu. Rev. Biochem. 53: 323-56, 1984, and Climie, S. et al., Proc. Natl. Acad. Sci. USA 87 :633-7, 1990.

The ZTMPO-1 polypeptides of the present invention, including full- length polypeptides, biologically active fragments, and fusion polypeptides, can be produced in genetically engineered host cells according to conventional techniques as described in Sheppard et al., WIPO International Patent Publication WO99/54,468, 1999. Suitable host cells are those cell types that can be transformed or transfected with exogenous DNA and grown in culture, and include bacteria, fungal cells, and cultured higher eukaryotic cells. Eukaryotic cells, particularly cultured cells of multicellular organisms, are preferred. Techniques for manipulating cloned DNA molecules and introducing exogenous DNA into a variety of host cells are disclosed by Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989, and Ausubel et al., eds., Current Protocols in Molecular Biology, John Wiley and Sons, Inc., NY, 1987.

Other higher eukaryotic cells can also be used as hosts, including plant cells, insect cells and avian cells. The use of Agrobacterium rhizogenes as a vector for expressing genes in plant cells has been reviewed by Sinkar et al., J. Biosci. (Bangalore) 1 47-58, 1987. Transformation of insect cells and production of foreign polypeptides therein is disclosed by Guarino et al., U.S. Patent No. 5,162,222, WIPO publication WO 94/06463, and Glick and Pasternak, Molecular Biotechnology: Principles and Applications of Recombinant DNA, ASM Press, Washington, D.C, 1994.

Fungal cells, including yeast cells, can also be used within the present invention. Yeast species of particular interest in this regard include Saccharomyces cerevisiae, Pichia pastoris, and Pichia methanolica. Methods for transforming S. cerevisiae cells with exogenous DNA and producing recombinant polypeptides therefrom are disclosed by, for example, Kawasaki, U.S. Patent No. 4,599,311;

Kawasaki et al., U.S. Patent No. 4,931,373; Brake, U.S. Patent No. 4,870,008; Welch et al., U.S. Patent No. 5,037,743; Murray et al., U.S. Patent No. 4,845,075; Raymond, 5,955,349; Raymond, 5,888,768; Raymond 6,001,597; and Raymond et al., 5,965,389.

Prokaryotic host cells, including strains of the bacteria Escherichia coli, Bacillus and other genera are also useful as host cells within the present invention.

Expressed recombinant ZTMPO-1 polypeptides can be purified using fractionation and/or conventional purification methods and media. Ammonium sulfate precipitation and acid or chaotrope extraction may be used for fractionation of samples. Exemplary purification steps may include hydroxyapatite, size exclusion, FPLC and reverse-phase high performance liquid chromatography. Suitable anion exchange media include derivatized dextrans, agarose, cellulose, polyacrylamide, specialty silicas, and the like. PEI, DEAE, QAE and Q derivatives are preferred, with DENE Fast-Flow Sepharose (Pharmacia, Piscataway, ΝJ) being particularly preferred. Exemplary chromatographic media include those media derivatized with phenyl, butyl, or octyl groups, such as Phenyl-Sepharose FF (Pharmacia), Toyopearl butyl 650 (Toso Haas, Montgomeryville, PA), Octyl-Sepharose (Pharmacia) and the like; or polyacrylic resins, such as Amberchrom CG 71 (Toso Haas) and the like. Suitable solid supports include glass beads, silica-based resins, cellulosic resins, agarose beads, cross-linked agarose beads, polystyrene beads, cross-linked polyacrylamide resins and the like that are insoluble under the conditions in which they are to be used. These supports may be modified with reactive groups that allow attachment of proteins by amino groups, carboxyl groups, sulfhydryl groups, hydroxyl groups and/or carbohydrate moieties. Examples of coupling chemistries include cyanogen bromide activation, Ν- hydroxysuccinimide activation, epoxide activation, sulfhydryl activation, hydrazide activation, and carboxyl and amino derivatives for carbodiimide coupling chemistries. These and other solid media are well known and widely used in the art, and are available from commercial suppliers. Methods for binding receptor polypeptides to support media are well known in the art. Selection of a particular method is a matter of routine design and is determined in part by the properties of the chosen support. See, for example, Affinity Chromatography: Principles & Methods, Pharmacia LKB Biotechnology, Uppsala, Sweden, 1988. The polypeptides of the present invention can be isolated by exploitation of their structural features. For example, immobilized metal ion adsoφtion (EVIAC) chromatography can be used to purify histidine-rich proteins or proteins having a His- tag. Briefly, a gel is first charged with divalent metal ions to form a chelate (E. Sulkowski, Trends Biochem. 3:1-7, 1985). Histidine-rich proteins will be adsorbed to this matrix with differing affinities, depending upon the metal ion used, and will be eluted by competitive elution, lowering the pH, or use of strong chelating agents. Other methods of purification include purification of glycosylated proteins by lectin affinity chromatography and ion exchange chromatography (Meth. Enzymol., Vol. 182, "Guide to Protein Purification", Deutscher, (ed.), Acad. Press, San Diego, 1990, pp.529-39). Within additional embodiments of the invention, a fusion of the polypeptide of interest and an affinity tag (e.g., polyhistidine, maltose-binding protein, Glu-Glu tag, FLAG tag, an immunoglobulin domain) may be constructed to facilitate purification. Protein refolding (and optionally reoxidation) procedures may be advantageously used. It is preferred to purify the protein to >80% purity, more preferably to >90% purity, even more preferably >95%, and particularly preferred is a pharmaceutically pure state, that is greater than 99.9% pure with respect to contaminating macromolecules, particularly other proteins and nucleic acids, and free of infectious and pyrogenic agents. Preferably, a purified protein is substantially free of other proteins, particularly other proteins of animal origin.

ZTMPO-1 polypeptides or fragments thereof may also be prepared through chemical synthesis. ZTMPO-1 polypeptides may be monomers or multimers; glycosylated or non-glycosylated; pegylated or non-pegylated; and may or may not include an initial methionine amino acid residue.

Antibodies that specifically bind to ZTMPO-1 epitopes, peptides or polypeptides for use within the methods described herein can be prepared by means known in the art and as described in Sheppard et al., WIPO International Patent Publication WO99/54,468, 1999. The ZTMPO-1 polypeptide, or a fragment, thereof serves as an antigen (immunogen) to inoculate an animal and elicit an immune response. Suitable antigens would be the ZTMPO-1 polypeptide encoded by SEQ ID NO:2 from amino acid number 1 to amino acid number 876, or contiguous 9 to 25 amino acid residue fragments thereof. Antibodies generated from this immune response can be isolated and purified as described herein. Methods for preparing and isolating polyclonal and monoclonal antibodies are well known in the art. See, for example, Current Protocols in Immunology. Cooligan, et al. (eds.), National Institutes of Health, John Wiley and Sons, Inc., 1995; Sambrook et al., Molecular Cloning: A Laboratory Manual, Second Edition, Cold Spring Harbor, NY, 1989; and Hurrell, (Ed.), Monoclonal Hvbridoma Antibodies: Techniques and Applications, CRC Press, Inc., Boca Raton, FL, 1982.

As used herein, the term "antibodies" includes polyclonal antibodies, affinity-purified polyclonal antibodies, monoclonal antibodies, and antigen-binding fragments, such as F(ab')2 and Fab proteolytic fragments. Genetically engineered intact antibodies or fragments, such as chimeric antibodies, Fv fragments, single chain antibodies and the like, as well as synthetic antigen-binding peptides and polypeptides, are also included. Non-human antibodies may be humanized by grafting non-human CDRs onto human framework and constant regions, or by incoφorating the entire non- human variable domains (optionally "cloaking" them with a human-like surface by replacement of exposed residues, wherein the result is a "veneered" antibody). In some instances, humanized antibodies may retain non-human residues within the human variable region framework domains to enhance proper binding characteristics. Through humanizing antibodies, biological half-life may be increased, and the potential for adverse immune reactions upon administration to humans is reduced.

A variety of assays known to those skilled in the art can be utilized to detect antibodies and binding proteins which specifically bind to ZTMPO-1 proteins or peptides. Exemplary assays are described in detail in Antibodies: A Laboratory Manual, Harlow and Lane (Eds.), Cold Spring Harbor Laboratory Press, 1988. Representative examples of such assays include: concurrent immunoelectrophoresis, radioimmunoassay, radioimmuno-precipitation, enzyme-linked immunosorbent assay (ELISA), dot blot or Western blot assay, inhibition or competition assay, and sandwich assay. In addition, antibodies can be screened for binding to wild-type versus mutant ZTMPO-1 protein or polypeptide. Antibodies to ZTMPO-1 may be used for detecting or quantitating soluble ZTMPO-1 as marker of underlying pathology or disease.

Antibodies or polypeptides herein may also be directly or indirectly conjugated to drugs, toxins, radionuclides and the like, and these conjugates used for in vivo diagnostic or therapeutic applications. More specifically, ZTMPO-1 polypeptides or anti-ZTMPO-1 antibodies, or bioactive fragments or portions thereof, can be coupled to detectable or cytotoxic molecules and delivered to a mammal having cells, tissues or organs that express the anti-complementary molecule.

The invention provided a method of detecting the presence of ZTMPO-1 in a biological sample, comprising the steps of: a) contacting the biological sample with an antibody, or an antibody fragment, that specifically binds with a polypeptide having the amino acid sequence of SEQ ID NO:2, or a fragment there of; wherein the contacting is performed under conditions that allow the binding of the antibody or antibody fragment to the biological sample, and b) detecting any of the bound antibody or bound antibody fragment.

In one type of in vitro assay, anti-ZTMPO-1 antibodies are used in liquid phase. For example, the presence of ZTMPO-1 in a biological sample can be tested by mixing the biological sample with a trace amount of labeled ZTMPO-1 and an anti- ZTMPO-1 antibody under conditions that promote binding between ZTMPO-1 and its antibody. Complexes of ZTMPO-1 and anti-ZTMPO-1 in the sample can be separated from the reaction mixture by contacting the complex with an immobilized protein which binds with the antibody, such as an Fc antibody or Staphylococcus protein A. The concentration of ZTMPO-1 in the biological sample will be inversely proportional to the amount of labeled ZTMPO-1 bound to the antibody and directly related to the amount of free labeled ZTMPO- 1.

Alternatively, in vitro assays can be performed in which anti-ZTMPO-1 antibody is bound to a solid-phase carrier. For example, antibody can be attached to a polymer, such as aminodextran, in order to link the antibody to an insoluble support such as a polymer-coated bead, a plate or a tube. Other suitable in vitro assays will be readily apparent to those of skill in the art.

In another approach, anti-ZTMPO-1 antibodies can be used to detect ZTMPO-1 in tissue sections prepared from a biopsy specimen. Such immunochemical detection can be used to determine the relative abundance of ZTMPO-1 and to determine the distribution of ZTMPO-1 in the examined tissue. General immunochemistry techniques are well established (see, for example, Ponder, "Cell Marking Techniques and Their Application," in Mammalian Development: A Practical Approach, Monk (ed.), pages 115-38, IRL Press 1987; Ausubel ibid.; and Manson (ed.), Methods In Molecular Biology, Vol.10: Immunochemical Protocols, The Humana Press, Inc., 1992).

Immunochemical detection can be performed by contacting a biological sample with an anti-ZTMPO-1 antibody, and then contacting the biological sample with a detectably labeled molecule which binds to the antibody. For example, the detectably labeled molecule can comprise an antibody moiety that binds to anti-ZTMPO-1 antibody. Alternatively, the anti-ZTMPO-1 antibody can be conjugated with avidin/streptavidin (or biotin) and the detectably labeled molecule can comprise biotin (or avidin/streptavidin). Numerous variations of this basic technique are well-known to those of skill in the art.

Alternatively, an anti-ZTMPO-1 antibody can be conjugated with a detectable label to form an anti-ZTMPO-1 immunoconjugate. Suitable direct tags or labels include radionuclides, enzymes, substrates, cofactors, inhibitors, fluorescent markers, chemiluminescent markers, magnetic particles and the like; indirect tags or labels may feature use of biotin-avidin or other complement/anti-complement pairs as intermediates. Antibodies herein may also be directly or indirectly conjugated to drugs, toxins, radionuclides and the like, and these conjugates used for in vivo diagnostic or therapeutic applications. Moreover, antibodies to ZTMPO-1 or fragments thereof may be used in vitro to detect denatured ZTMPO-1 or fragments thereof in assays, for example, Western Blots or other assays known in the art.

The detectable label can be a radioisotope that is detected by autoradiography. Isotopes that are particularly useful for the puφose of the present invention are ³H, ¹²⁵I, ^{I 1}I, ³⁵S and ^l4C.

Anti-ZTMPO-1 immunoconjugates can also be labeled with a fluorescent compound. The presence of a fluorescently-labeled antibody is determined by exposing the immunoconjugate to light of the proper wavelength and detecting the resultant fluorescence. Fluorescent labeling compounds include fluorescein isothiocyanate, rhoda- mine, phycoerytherin, phycocyanin, allophycocyanin, o-phth-aldehyde and fluorescamine. Alternatively, anti-ZTMPO-1 irnmunoconjugates can be detectably labeled by coupling an antibody component to a chemiluminescent compound. The presence of the chemiluminescent-tagged immunoconjugate is determined by detecting the presence of luminescence that arises during the course of a chemical reaction. Examples of chemi- luminescent labeling compounds include luminol, isoluminol, an aromatic acridinium ester, an imidazole, an acridinium salt and an oxalate ester.

Similarly, a bioluminescent compound can be used to label anti-ZTMPO-1 irnmunoconjugates of the present invention. Bioluminescence is a type of chemiluminescence found in biological systems in which a catalytic protein increases the efficiency of the chemiluminescent reaction. The presence of a bioluminescent protein is determined by detecting the presence of luminescence. Bioluminescent compounds that are useful for labeling include luciferin, luciferase and aequorin.

Alternatively, anti-ZTMPO-1 irnmunoconjugates can be detectably labeled by linking an anti-ZTMPO-1 antibody component to an enzyme. When the anti-ZTMPO- 1 -enzyme conjugate is incubated in the presence of the appropriate substrate, the enzyme moiety reacts with the substrate to produce a chemical moiety which can be detected, for example, by spectrophotometric, fluorometric or visual means. Examples of enzymes that can be used to detectably label polyspecific irnmunoconjugates include β-galactosidase, glucose oxidase, peroxidase and alkaline phosphatase. Those of skill in the art will know of other suitable labels which can be employed in accordance with the present invention. The binding of marker moieties to anti-ZTMPO-1 antibodies can be accomplished using standard techniques known to the art. Typical methodology in this regard is described by Kennedy et al., Clin. Chim. Acta 70:1, 1976; Schurs et al., Clin. Chim. Acta 81:1, 1977; Shih et al.. Intl. J. Cancer 46: 1101. 1990; and Stein et al., Cancer Res. 50:1330,1990.

Moreover, the convenience and versatility of immunochemical detection can be enhanced by using anti-ZTMPO-1 antibodies that have been conjugated with avidin, streptavidin, and biotin (see, for example, Wilchek et al. (eds.), "Avidin-Biotin Technology," Methods In Enzvmology, Vol. 184 (Academic Press 1990), and Bayer et al., "Immunochemical Applications of Avidin-Biotin Technology," in Methods In Molecular

Biology, Vol. 10, Manson (ed.), pages 149-162 (The Humana Press, Inc. 1992).

Methods for performing immunoassays are well-established. See, for example, Cook and Self, "Monoclonal Antibodies in Diagnostic Immunoassays," in

Monoclonal Antibodies: Production, Engineering, and Clinical Application, Ritter and Ladyman (eds.), pages 180-208, (Cambridge University Press, 1995), Perry, "The Role of

Monoclonal Antibodies in the Advancement of Immunoassay Technology," in Monoclonal Antibodies: Principles and Applications, Birch and Lennox (eds.), pages 107- 120 (Wiley-Liss, Inc. 1995), and Diamandis, Lmmunoassay (Academic Press, Inc. 1996).

The present invention also contemplates the use of irnmunoconjugates for in vivo detection of ZTMPO-1 polypeptide. As an illustration, the method of diagnostic imaging with radiolabeled monoclonal antibodies is well-known. Examples of radioisotopes that can be bound to antibodies and are appropriate for diagnostic imaging include K-emitters and positron-emitters such as ⁹⁹Tc, ⁹⁴Tc, ⁶⁷Ga, ^Cu, ^{H 1}In, i23_L i24_L i2_ i3i_j 51^ 89^ i8_{p and 6}8_{Ga Qther suitable ra} ioisotopes are known to those of skill in the art. Methods for performing immunoscintigraphy are described, for example, by Srivastava (ed.), Radiolabeled Monoclonal Antibodies For Imaging And Therapy (Plenum Press 1988), Chase, "Medical Applications of Radioisotopes," in Remington's Pharmaceutical Sciences, Volume π, Gennaro et al. (eds.), pp. 843-65 (Mack Publishing Co., 1995), and Brown, "Clinical Use of Monoclonal Antibodies," in Biotechnology and Pharmacy, pages 227-49, Pezzuto et al. (eds.) (Chapman & Hall 1993).

Anti-ZTMPO-1 antibodies also can be labeled with paramagnetic ions for puφoses of in vivo diagnosis. Elements that are particularly useful for magnetic resonance imaging include Gd, Mn, Dy and Fe ions.

The invention is further illustrated by the following non-limiting example.

EXAMPLES

Example 1 Chromosomal Assignment and Placement of ZTMPO-1

ZTMPO-1 was mapped in human by polymerase chain reaction (PCR) to chromosome 12 using both the low resolution GeneBridge 4 and the medium resolution Stanford G3 radiation hybrid (RH) mapping panels (Research Genetics, Inc., Huntsville, AL). The GeneBridge 4 RH Panel contains DNA from each of 93 radiation hybrid clones, plus two control DNAs (the HFL donor and the A23 recipient). The Stanford G3 RH Panel contains DNA from each of 83 radiation hybrid clones, plus two control DNAs (the RM donor and the A3 recipient). Publicly available WWW servers (http://www-genome.wi. mit.edu/,http://shgc-www. stanford.edu/, and http://www.ncbi. nlm.nih.gov/gene map99/, respectively) allow chromosomal placement in respect to already mapped genes and markers.

For the mapping of ZTMPO-1 with either panel, 20 μl reactions were set up in a microtiter plate compatible for PCR (Stratagene, La Jolla, CA) and used in a RoboCycler Gradient 96 thermal cycler (Stratagene). Each of the 95 PCR reactions consisted of 2 μl 10X KlenTaq PCR reaction buffer (Clontech, Palo Alto, CA), 1.6 μl dNTPs mix (2.5 mM each, Perkin-Elmer, Foster City, CA), 1 μl sense primer, ZC 15,487 (SEQ ID NO: 3), 1 μl antisense primer, ZC 15,486 (SEQ ID NO:4), 2 μl RediLoad (Research Genetics, Inc., Huntsville, AL), 0.4 μl 50X Advantage KlenTaq Polymerase Mix (Clontech Laboratories, Inc.), 25 ng of DNA from an individual hybrid clone or control and distilled water for a total volume of 20 μl. The reactions were overlaid with an equal amount of mineral oil and sealed. The PCR cycler conditions were as follows: an initial 1 cycle 5 minute denaturation at 95°C, 35 cycles of a 1 minute denaturation at 95°C, 1 minute annealing at 62°C and 1.5 minute extension at 72°C, followed by a final 1 cycle extension of 7 minutes at 72°C. The reactions were separated by electrophoresis on a 2% agarose gel (EM Science, Gibbstown, NJ) and visualized by staining with ethidium bromide.

The RH mapping results placed ZTMPO-1 22.44 cR_3,000 distal of framework marker D12S367 on the GeneBridge 4 RH map and 12 cR_10,000 distal of framework marker AFM295ye9 on the Stanford G3 RH map. These position ZTMPO-1 in the 12q24.3 chromosomal region and approximately 1.35 mb distal of the Alzheimer's Disease loci linked marker D12S1045 which mapped 10 cR_ 10,000 distal of the Stanford G3 RH map framework marker D12S97 (AFM210zd6). From the foregoing, it will be appreciated that, although specific embodiments of the invention have been described herein for puφoses of illustration, various modifications may be made without deviating from the spirit and scope of the invention. Accordingly, the invention is not limited except as by the appended claims.

Claims

What is claimed is:

1. A method of detecting a chromosome 12 abnormality in a subject comprising:

(a) amplifying a polynucleotide from RNA isolated from a biological sample using primers derived from a polynucleotide of SEQ ID NO: 1 or its complement; and

(b) detecting an aberration in the amplified polynucleotide by comparing the nucleic acid sequence of said amplified polynucleotide with the nucleic acid sequence of the polynucleotide of SEQ ID NO:l; wherein the presence of a aberration indicates a chromosome 12 abnormality.

2. The method according to claim 1, wherein amplification is performed by polymerase chain reaction or reverse transcriptase-polymerase chain reaction.

3. A method of detecting a chromosome 12 abnormality in a subject comprising:

(a) amplifying a polynucleotide from RNA isolated from a biological sample using primers derived from a polynucleotide of SEQ ID NO: 1 or its complement;

(b) transcribing the amplified polynucleotide to express mRNA;

(c) translating said mRNA to produce a polypeptide; and

(d) detecting an aberration in the amino acid sequence of said transcribed polypeptide by comparison to the amino acid sequence of SEQ ID NO:2; wherein the presence of an aberration indicates a chromosome 12 abnormality.

4. A method for diagnosing a neurological disease or susceptibility to a neurological disease in an individual, wherein said disease is related to the expression or activity of a polypeptide of SEQ ED NO:2 or fragment thereof, in said individual, comprising the step of: determining the presence of an alteration in the nucleic acid sequence of a polynucleotide encoding said polypeptide in the genome of said individual by comparison of the nucleic acid sequence of said polynucleotide with the nucleic acid sequence of SEQ ED NO:l; wherein the presence of an aberration in said nucleic acid sequence indicates a neurological disease or susceptibility to a neurological disease. A method of claim 4, wherein said neurological disease is Alzheimer's

Disease.

6. A method for diagnosing a neurological disease or susceptibility to a neurological disease in an individual, comprising:

(b) detecting an aberration in the amplified polynucleotide by comparing the nucleic acid sequence of said amplified polynucleotide with the nucleic acid sequence of the polynucleotide of SEQ ID NO: 1 ; wherein the presence of an aberration indicates a neurological disease or susceptibility to a neurological disease.

A method of claim 6, wherein said neurological disease is Alzheimer's

Disease.

8. A method for diagnosing a neurological disease or susceptibility to a neurological disease in an individual, comprising:

(b) transcribing the amplified polynucleotide to express mRNA;

(c) translating said mRNA to produce a polypeptide; and

(d) detecting an aberration in the amino acid sequence of said transcribed polypeptide by comparison to the amino acid sequence of SEQ ID NO:2; wherein the presence of a mutation indicates a neurological disease or susceptibility to a neurological disease.

9. The method for according to claim 8, wherein the neurological disease is Alzheimer's Disease.

10. A method for diagnosing a neurological disease or susceptibility to a neurological disease in an individual, comprising:

(a) contacting a polynucleotide probe with a biological sample under hybridizing conditions; wherein said polynucleotide probe is derived from a polynucleotide of SEQ ID NO: 1 or its complement; and

(b) detecting the formation of a hybrid between said polynucleotide probe and said biological sample; wherein the presence of said hybrid indicates the presence of a ZTMPO-1 polynucleotide in said biological sample.

11. A method of detecting the presence of a polypeptide of SEQ ID NO:2, or a fragment thereof, in a biological sample, comprising the steps of:

(a) adding an antibody, or an antibody fragment, that specifically binds with a polypeptide of SEQ ID NO: 2 to said biological sample under conditions that allow the binding of said antibody or antibody fragment to said biological sample;

(b) detecting any of said bound antibody or bound antibody fragment; and

(c) correlating the presence of a polypeptide of SEQ ID NO:2, or a fragment thereof, with a neurological disease or susceptibility to a neurological disease.

12. The method of claim 11, wherein said antibody or said antibody fragment further comprises a detectable label selected from the group consisting of radioisotope, fluorescent label, chemiluminescent label, enzyme label, bioluminescent label, and colloidal gold.

13. A kit for the detection of a gene encoding a polypeptide of SEQ ID NO:2, comprising: a first container that comprises a polynucleotide of SEQ ID NO: l or its complement; and a second container that comprises one or more reagents capable of indicating the presence of said polynucleotide.

14. A kit for the detection of a gene encoding a polypeptide of SEQ ED NO:2, comprising: a first container comprising an antibody that specifically binds to a polypeptide of SEQ ID NO:2; and a second container that comprises one or more reagents capable of indicating the presence of said antibody.