WO2002090568A2

WO2002090568A2 - Therapeutic polypeptides, nucleic acids encoding same, and methods of use

Info

Publication number: WO2002090568A2
Application number: PCT/US2002/014341
Authority: WO
Inventors: John P. Ii Alsobrook; David W. Anderson; Ferenc L. Boldog; Catherine E. Burgess; Stacie J. Casman; Schlomit R. Edinger; Karen Ellerman; Esha A. Gangolli; Valerie L. Gerlach; Linda Gorman; Erik Gunther; John L. Herrmann; Weizhen Ji; Denise M. Lepley; David A. Lewin; Li Li; John R. Macdougall; Uriel M. Malyankar; Peter D. Mezes; Muralidhara Padigaru
Original assignee: Curagen Corporation
Priority date: 2001-05-03
Filing date: 2002-05-02
Publication date: 2002-11-14
Also published as: EP1539806A2; CA2446427A1; WO2002090568A3

Abstract

Disclosed herein are nucleic acid sequences that encode G-coupled protein-receptor related polypeptides. Also disclosed are polypeptides encoded by these nucleic acid sequences, and antibodies, which immunospecifically-bind to the-polypeptide, as well as derivatives, variants, mutants, or fragments of the aforementioned polypeptide, polynucleotide, or antibody. The invention further discloses therapeutic, diagnostic and research methods for diagnosis, treatment, and prevention of disorders involving any one of these novel human nucleic acids and proteins.

Description

THERAPEUTIC POLYPEPTIDES, NUCLEIC ACIDS ENCODING SAME, AND METHODS OF USE

FIELD OF THE INVENTION

The present invention relates to novel polypeptides, and the nucleic acids encoding them, having properties related to stimulation of biochemical or physiological responses in a cell, a tissue, an organ or an organism. More particularly, the novel polypeptides are gene products of novel genes, or are specified biologically active fragments or derivatives thereof. Methods of use encompass diagnostic and prognostic assay procedures as well as methods of treating diverse pathological conditions.

BACKGROUND OF THE INVENTION

Eukaryotic cells are characterized by. biochemical and physiological processes, which under normal conditions are exquisitely balanced to achieve the preservation and propagation of the cells. When such cells are components of multicellular organisms such as vertebrates or, more particularly, organisms such as mammals, the regulation of the biochemical and physiological processes involves intricate signaling pathways. Frequently, such signaling pathways include constituted of extracellular signaling proteins, cellular receptors that bind the signaling proteins and signal transducing components located within the cells.

Signaling proteins may be classified as endocrine effectors, paracrine effectors or autocrine effectors. Endocrine effectors are signaling molecules secreted by a given organ into the circulatory system, which are then transported to a distant target organ or tissue. The target cells include the receptors for the endocrine effector, and when the endocrine effector binds, a signaling cascade is induced. Paracrine effectors involve secreting cells and receptor cells in close proximity to each other, such as two different classes of cells in the same tissue or organ. One class of cells secretes the paracrine effector, which then reaches the second class of cells, for example by diffusion through the extracellular fluid. The second class of cells contains the receptors for the paracrine effector; binding of the effector results in induction of the signaling cascade that elicits the conesponding biochemical or physiological effect. Autocrine effectors are highly analogous to paracrine effectors, except that the same cell type that secretes the autocrine effector also contains the receptor. Thus the autocrine effector binds to receptors on the same cell, or on identical neighboring cells. The binding process then elicits the characteristic biochemical or physiological effect.

Signaling processes may elicit a variety of effects on cells and tissues including, by way of nonlimiting example, induction of cell or tissue proliferation, suppression of growth or proliferation, induction of differentiation or maturation of a cell or tissue, and suppression of differentiation or maturation of a cell or tissue. Many pathological conditions involve dysregulation of expression of important effector proteins. In certain classes of pathologies the dysregulation is manifested as diminished or suppressed level of synthesis and secretion of protein effectors. In other classes of pathologies the dysregulation is manifested as increased or up-regulated level of synthesis and secretion of protein effectors. In a clinical setting a subject may be suspected of suffering from a condition brought on by altered or mis-regulated levels of a protein effector of interest. Therefore there is a need to assay for the level of the protein effector of interest in a biological sample from such a subject, and to compare the level with that characteristic of a nonpathological condition. There also is a need to provide the protein effector as a product of manufacture. Administration of the effector to a subject in need thereof is useful in treatment of the pathological condition. Accordingly, there is a need for a method of treatment of a pathological condition brought on by a diminished or suppressed levels of the protein effector of interest. In addition, there is a need for a method of treatment of a pathological condition brought on by a increased or up-regulated levels of the protein effector of interest.

SUMMARY OF THE INVENTION

The invention is based in part upon the discovery of isolated polypeptides including amino acid sequences selected from mature forms of the amino acid sequences selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62. The invention also is based in part upon variants of a mature form of the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, wherein any amino acid in the mature form is changed to a different amino acid, provided that no more than 15% of the amino acid residues in the sequence of the mature form are so changed. In another embodiment, the invention includes the amino acid sequences selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62. In another embodiment, the invention also comprises variants of the amino acid sequence selected from the group consisting of SEQ ID NO:2n, ^"wherein n is an integer between 1 and 62 wherem any amino acid specified in the chosen sequence is changed to a different amino acid, provided that no more than 15% of the amino acid residues in the sequence are so changed. The invention also involves fragments of any of the mature forms of the amino acid sequences selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, or any other amino acid sequence selected from this group. The invention also comprises fragments from these groups in which up to 15% of the residues are changed.

In another embodiment, the invention encompasses polypeptides that are naturally occurring allelic variants of the sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62. These allelic variants include amino acid sequences that are the translations of nucleic acid sequences differing by a single nucleotide from nucleic acid sequences selected from the group consisting of SEQ ID NOS: 2n-l, wherein n is an integer between 1 and 62. The variant polypeptide where any amino acid changed in the chosen sequence is changed to provide a conservative substitution.

In another embodiment, the invention comprises a pharmaceutical composition involving a polypeptide with an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 and a pharmaceutically acceptable carrier. In another embodiment, the invention involves a kit, including, in one or more containers, this pharmaceutical composition.

In another embodiment, the invention includes the use of a therapeutic in the manufacture of a medicament for treating a syndrome associated with a human disease, the disease being selected from a pathology associated with a polypeptide with an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 wherein said therapeutic is the polypeptide selected from this group.

In another embodiment, the invention comprises a method for determining the presence or amount of a polypeptide with an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 in a sample, the method involving providing the sample; introducing the sample to an antibody that binds immunospecifically to the polypeptide; and determining the presence or amount of antibody bound to the polypeptide, thereby determining the presence or amount of polypeptide in the sample. In another embodiment, the invention includes a method for determining the presence of or predisposition to a disease associated with altered levels of a polypeptide with an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 in a first mammalian subject, the method involving measuring the level of expression of the polypeptide in a sample from the first mammalian subject; and comparing the amount of the polypeptide in this sample to the amount of the polypeptide present in a control sample from a second mammalian subject known not to have, or not to be predisposed to, the disease, wherein an alteration in the expression level of the polypeptide in the first subject as compared to the control sample indicates the presence of or predisposition to the disease.

In another embodiment, the invention involves a method of identifying an agent that binds to a polypeptide with an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, the method including introducing the polypeptide to the agent; and determining whether the agent binds to the polypeptide. The agent could be a cellular receptor or a downstream effector. In another embodiment, the invention involves a method for identifying a potential therapeutic agent for use in treatment of a pathology, wherein the pathology is related to abenant expression or abenant physiological interactions of a polypeptide with an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, the method including providing a cell expressing the polypeptide of the invention and having a property or function ascribable to the polypeptide; contacting the cell with a composition comprising a candidate substance; and determining whether the substance alters the property or function ascribable to the polypeptide; whereby, if an alteration observed in the presence of the substance is not observed when the cell is contacted with a composition devoid of the substance, the substance is identified as a potential therapeutic agent.

In another embodiment, the invention involves a method for screening for a modulator of activity or of latency or predisposition to a pathology associated with a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, the method including administering a test compound to a test animal at increased risk for a pathology associated with the polypeptide of the invention, wherein the test animal recombinantly expresses the polypeptide of the invention; measuring the activity of the polypeptide in the test animal after administering the test compound; and comparing the activity of the protein in the test animal with the activity of the polypeptide in a control animal not administered the polypeptide, wherein a change in the activity of the polypeptide in the test animal relative to the control animal indicates the test compound is a modulator of latency of, or predisposition to, a pathology associated with the polypeptide of the invention. The recombinant test animal could express a test protein transgene or express the transgene under the control of a promoter at an increased level relative to a wild-type test animal The promoter may or may not b the native gene promoter of the transgene. In another embodiment, the invention involves a method for modulating the activity of a polypeptide with an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, the method including introducing a cell sample expressing the polypeptide with a compound that binds to the polypeptide in an amount sufficient to modulate the activity of the polypeptide.

In another embodiment, the invention involves a method of treating or preventing a pathology associated with a polypeptide with an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, the method including administering the polypeptide to a subject in which such treatment or prevention is desired in an amount sufficient to treat or prevent the pathology in the subject. The subject could be human.

In another embodiment, the invention involves a method of treating a pathological state in a mammal, the method including administering to the mammal a polypeptide in an amount that is sufficient to alleviate the pathological state, wherein the polypeptide is a polypeptide having an amino acid sequence at least 95% identical to a polypeptide having the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 or a biologically active fragment thereof.

In another embodiment, the invention involves an isolated nucleic acid molecule comprising a nucleic acid sequence encoding a polypeptide having an amino acid sequence selected from the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62; a variant of a mature form of the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 wherein any amino acid in the mature form of the chosen sequence is changed to a different amino acid, provided that no more than 15% of the amino acid residues in the sequence of the mature form are so changed; the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62; a variant of the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, in which any amino acid specified in the chosen sequence is changed to a different amino acid, provided that no more than 15% of the amino acid residues in the sequence are so changed; a nucleic acid fragment encoding at least a portion of a polypeptide comprising the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 or any variant of the polypeptide wherein any amino acid of the chosen sequence is changed to a different amino acid, provided that no more than 10% of the amino acid residues in the sequence are so changed; and the complement of any of the nucleic acid molecules.

In another embodiment, the invention comprises an isolated nucleic acid molecule having a nucleic acid sequence encoding a polypeptide comprising an amino acid sequence selected from the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62, wherein the nucleic acid molecule comprises the nucleotide sequence of a naturally occurring allelic nucleic acid variant. In another embodiment, the invention involves an isolated nucleic acid molecule including a nucleic acid sequence encoding a polypeptide having an amino acid sequence selected from the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62 that encodes a variant polypeptide, wherein the variant polypeptide has the polypeptide sequence of a naturally occurring polypeptide variant.

In another embodiment, the invention comprises an isolated nucleic acid molecule having a nucleic acid sequence encoding a polypeptide comprising an amino acid sequence selected from the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62, wherein the nucleic acid molecule differs by a single nucleotide from a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 2n-l, wherein n is an integer between 1 and 62.

In another embodiment, the invention includes an isolated nucleic acid molecule having a nucleic acid sequence encoding a polypeptide including an amino acid sequence selected from the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62, wherein the nucleic acid molecule comprises a nucleotide sequence selected from the group consisting of the nucleotide sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62; a nucleotide sequence wherein one or more nucleotides in the nucleotide sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62 is changed from that selected from the group consisting of the chosen sequence to a different nucleotide provided that no more than 15% of the nucleotides are so changed; a nucleic acid fragment of the sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62; and a nucleic acid fragment wherein one or more nucleotides in the nucleotide sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62 is changed from that selected from the group consisting of the chosen sequence to a different nucleotide provided that no more than 15% of the nucleotides are so changed. In another embodiment, the invention includes an isolated nucleic acid molecule having a nucleic acid sequence encoding a polypeptide including an amino acid sequence selected from the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62, wherein the nucleic acid molecule hybridizes under stringent conditions to the nucleotide sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62, or a complement of the nucleotide sequence.

In another embodiment, the invention includes an isolated nucleic acid molecule having a nucleic acid sequence encoding a polypeptide including an amino acid sequence selected from the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62, wherein the nucleic acid molecule has a nucleotide sequence in which any nucleotide specified in the coding sequence of the chosen nucleotide sequence is changed from that selected from the group consisting of the chosen sequence to a different nucleotide provided that no more than 15% of the nucleotides in the chosen coding sequence are so changed, an isolated second polynucleotide that is a complement of the first polynucleotide, or a fragment of any of them.

In another embodiment, the invention includes a vector involving the nucleic acid molecule having a nucleic acid sequence encoding a polypeptide including an amino acid sequence selected from the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62. This vector can have a promoter operably linked to the nucleic acid molecule. This vector can be located within a cell.

In another embodiment, the invention involves a method for determining the , presence or amount of a nucleic acid molecule having a nucleic acid sequence encoding a polypeptide including an amino acid sequence selected firom the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62 in a sample, the method including providing the sample; introducing the sample to a probe that binds to the nucleic acid molecule; and determining the presence or amount of the probe bound to the nucleic acid molecule, thereby determining the presence or amount of the nucleic acid molecule in the sample. The presence or amount of the nucleic acid molecule is used as a marker for cell or tissue type. The cell type can be cancerous. In another embodiment, the invention involves a method for determining the presence of or predisposition for a disease associated with altered levels of a nucleic acid molecule having a nucleic acid sequence encoding a polypeptide including an amino acid sequence selected from the group consisting of a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62 in a first mammalian subject, the method including measuring the amount of the nucleic acid in a sample from the first mammalian subject; and comparing the amount of the nucleic acid in the sample of step (a) to the amount of the nucleic acid present in a control sample from a second mammalian subject known not to have or not be predisposed to, the disease; wherein an alteration in the level of the nucleic acid in the first subject as compared to the control sample indicates the presence of or predisposition to the disease.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incoφorated by reference in their entirety. In the case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting. Other features and advantages of the invention will be apparent from the following detailed description and claims.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides novel nucleotides and polypeptides encoded thereby. Included in the invention are the novel nucleic acid sequences, their encoded polypeptides, antibodies, and other related compounds. The sequences are collectively refened to herein as "NONX nucleic acids" or "ΝOVX polynucleotides" and the conesponding encoded polypeptides are refened to as "ΝONX polypeptides" or "ΝONX proteins." Unless indicated otherwise, "NOVX" is meant to refer to any of the novel sequences disclosed herein. Table 1 provides a summary of the NONX nucleic acids and their encoded polypeptides.

TABLE 1. Sequences and Corresponding SEQ ID Numbers

Table 1 indicates homology of NOVX nucleic acids to known protein families. Thus, the nucleic acids and polypeptides, antibodies and related compounds according to the invention conesponding to a NONX as identified in column 1 of Table 1 will be useful in therapeutic and diagnostic applications implicated in, for example, pathologies and disorders associated with the known protein families identified in column 5 of Table 1.

ΝONX nucleic acids and their encoded polypeptides are useful in a variety of applications and contexts. The various ΝONX nucleic acids and polypeptides according to the invention are useful as novel members of the protein families according to the presence of domains and sequence relatedness to previously described proteins. Additionally, ΝONX nucleic acids and polypeptides can also be used to identify proteins that are members of the family to which the ΝONX polypeptides belong.

Consistent with other known members of the family of proteins, identified in column 5 of Table 1, the ΝONX polypeptides of the present invention show homology to, and contain domains that are characteristic of, other members of such protein families. Details of the sequence relatedness and domain analysis for each NONX are presented in Example A.

The ΝONX nucleic acids and polypeptides can also be used to screen for molecules, which inhibit or enhance ΝOVX activity or function. Specifically, the nucleic acids and polypeptides according to the invention may be used as targets for the identification of small molecules that modulate or inhibit diseases associated with the protein families listed in Table 1.

The ΝOVX nucleic acids and polypeptides are also useful for detecting specific cell types. Details of the expression analysis for each ΝOVX are presented in Example C. Accordingly, the ΝOVX nucleic acids, polypeptides, antibodies and related compounds according to the invention will have diagnostic and therapeutic applications in the detection of a variety of diseases with differential expression in normal vs. diseased tissues, e.g. a variety of cancers.

Additional utilities for ΝOVX nucleic acids and polypeptides according to the invention are disclosed herein.

ΝOVX clones

ΝOVX nucleic acids and their encoded polypeptides are useful in a variety of applications and contexts. The various ΝOVX nucleic acids and polypeptides according to the invention are useful as novel members of the protein families according to the presence of domains and sequence relatedness to previously described proteins.

Additionally, ΝOVX nucleic acids and polypeptides can also be used to identify proteins that are members of the family to which the ΝOVX polypeptides belong.

The ΝOVX genes and their conesponding encoded proteins are useful for preventing, treating or ameliorating medical conditions, e.g., by protein or gene therapy. Pathological conditions can be diagnosed by determining the amount of the new protein in a sample or by determining the presence of mutations in the new genes. Specific uses are described for each of the ΝOVX genes, based on the tissues in which they are most highly expressed. Uses include developing products for the diagnosis or treatment of a variety of diseases and disorders. The ΝOVX nucleic acids and proteins of the invention are useful in potential diagnostic and therapeutic applications and as a research tool. These include serving as a specific or selective nucleic acid or protein diagnostic and/or prognostic marker, wherein the presence or amount of the nucleic acid or the protein are to be assessed, as well as potential therapeutic applications such as the following: (i) a protein therapeutic, (ii) a small molecule drug target, (iii) an antibody target (therapeutic, diagnostic, drug targeting/cytotoxic antibody), (iv) a nucleic acid useful in gene therapy (gene delivery/gene ablation), and (v) a composition promoting tissue regeneration in vitro and in vivo (vi) biological defense weapon.

In one specific embodiment, the invention includes an isolated polypeptide comprising an amino acid sequence selected from the group consisting of: (a) a mature form of the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62; (b) a variant of a mature form of the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, wherein any amino acid in the mature form is changed to a different amino acid, provided that no more than 15% of the amino acid residues in the sequence of the mature form are so changed; (c) an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62; (d) a variant of the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 wherein any amino acid specified in the chosen sequence is changed to a different amino acid, provided that no more than 15% of the amino acid residues in the sequence are so changed; and (e) a fragment of any of (a) through (d). In another specific embodiment, the invention includes an isolated nucleic acid molecule comprising a nucleic acid sequence encoding a polypeptide comprising an amino acid sequence selected from the group consisting of: (a) a mature form of the amino acid sequence given SEQ ID NO:2n, wherein n is an integer between 1 and 62; (b) a variant of a mature form of the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 wherein any amino acid in the mature form of the chosen sequence is changed to a different amino acid, provided that no more than 15% of the amino acid residues in the sequence of the mature form are so changed; (c) the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62; (d) a variant of the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62, in which any amino acid specified in the chosen sequence is changed to a different amino acid, provided that no more than 15% of the amino acid residues in the sequence are so changed; (e) a nucleic acid fragment encoding at least a portion of a polypeptide comprising the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 or any variant of said polypeptide wherein any amino acid of the chosen sequence is changed to a different amino acid, provided that no more than 10% of the amino acid residues in the sequence are so changed; and (f) the complement of any of said nucleic acid molecules.

In yet another specific embodiment, the invention includes an isolated nucleic acid molecule, wherein said nucleic acid molecule comprises a nucleotide sequence selected from the group consisting of: (a) the nucleotide sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62; (b) a nucleotide sequence wherein one or more nucleotides in the nucleotide sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62 is changed from that selected from the group consisting of the chosen sequence to a different nucleotide provided that no more than 15% of the nucleotides are so changed; (c) a nucleic acid fragment of the sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62; and (d) a nucleic acid fragment wherein one or more nucleotides in the nucleotide sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62 is changed from that selected from the group consisting of the chosen sequence to a different nucleotide provided that no more than 15% of the nucleotides are so changed.

NOVX Nucleic Acids and Polypeptides

One aspect of the invention pertains to isolated nucleic acid molecules that encode NOVX polypeptides or biologically active portions thereof. Also included in the invention are nucleic acid fragments sufficient for use as hybridization probes to identify NOVX-encoding nucleic acids (e.g. , NOVX mRNAs) and fragments for use as PCR primers for the amplification and/or mutation of NOVX nucleic acid molecules. As used herein, the term "nucleic acid molecule" is intended to include DNA molecules (e.g., cDNA or genomic DNA), RNA molecules (e.g., mRNA), analogs of the DNA or RNA generated using nucleotide analogs, and derivatives, fragments and homologs thereof. The nucleic acid molecule may be single-stranded or double-stranded, but preferably is comprised double-stranded DNA.

A NOVX nucleic acid can encode a mature NOVX polypeptide. As used herein, a "mature" form of a polypeptide or protein disclosed in the present invention is the product of a naturally occurring polypeptide, precursor form, or proprotein. The naturally occurring polypeptide, precursor or proprotein includes, by way of nonlimiting example, the full-length gene product encoded by the conesponding gene. Alternatively, it may be defined as the polypeptide, precursor or proprotein encoded by an ORF described herein. The product "mature" form arises, by way of nonlimiting example, as a result of one or more naturally occurring processing steps that may take place within the cell (host cell) in which the gene product arises. Examples of such processing steps leading to a "mature" form of a polypeptide or protein include the cleavage of the N-terminal methionine residue encoded by the initiation codon of an ORF or the proteolytic cleavage of a signal peptide or leader sequence. Thus a mature form arising from a precursor polypeptide or protein that has residues 1 to N, where residue 1 is the N-terminal methionine, would have residues 2 through N remaining after removal of the N-terminal methionine. Alternatively, a mature form arising from a precursor polypeptide or protein having residues 1 to N, in which an N-terminal signal sequence from residue 1 to residue M is cleaved, would have the residues from residue M+l to residue N remaining. Further as used herein, a "mature" form of a polypeptide or protein may arise from a post-translational modification other than a proteolytic cleavage event. Such additional processes include, by way of non-limiting example, glycosylation, myristoylation or phosphorylation. In general, a mature polypeptide or protein may result from the operation of only one of these processes, or a combination of any of them.

The term "probe", as utilized herein, refers to nucleic acid sequences of variable length, preferably between at least about 10 nucleotides (nt), and 100 nt, or as many as approximately, e.g., 6,000 nt, depending upon the specific use. Probes are used in the detection of identical, similar, or complementary nucleic acid sequences. Longer length probes are generally obtained from a natural or recombinant source, are highly specific, and much slower to hybridize than shorter-length oligomer probes. Probes may be single- or double-stranded and designed to have specificity in PCR, membrane-based hybridization technologies, or ELISA-like technologies.

The term "isolated" nucleic acid molecule, as used herein, is a nucleic acid which is separated from other nucleic acid molecules which are present in the natural source of the nucleic acid. Preferably, an "isolated" nucleic acid is free of sequences which naturally flank the nucleic acid (i.e., sequences located at the 5'- and 3'-termini of the nucleic acid) in the genomic DNA of the organism from which the nucleic acid is derived. For example, in various embodiments, the isolated NOVX nucleic acid molecules can contain less than about 5 kb, 4 kb, 3 kb, 2 kb, 1 kb, 0.5 kb, 0.1 kb, or less of nucleotide sequences which naturally flank the nucleic acid molecule in genomic DNA of the cell/tissue from which the nucleic acid is derived (e.g., brain, heart, liver, spleen, etc.). Moreover, an "isolated" nucleic acid molecule, such as a cDNA molecule, can be substantially free of other cellular material, culture medium, or of chemical precursors or other chemicals.

A nucleic acid molecule of the invention, e.g., a nucleic acid molecule having the nucleotide sequence SEQ ID NOS: 2n-l, wherein n is an integer between 1 and 62, or a complement of this nucleotide sequence, can be isolated using standard molecular biology techniques and the sequence information provided herein. Using all or a portion of the nucleic acid sequence of SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, as a hybridization probe, NOVX molecules can be isolated using standard hybridization and cloning techniques (e.g., as described in Sambrook, et al., (eds.), MOLECULAR CLONING: A LABORATORY MANUAL 2^nd Ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989; and Ausubel, et al, (eds.), CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley & Sons, New York, NY, 1993).

A nucleic acid of the invention can be amplified using cDNA, mRNA or, alternatively, genomic DNA as a template with appropriate oligonucleotide primers according to standard PCR amplification techniques. The nucleic acid so amplified can be cloned into an appropriate vector and characterized by DNA sequence analysis. Furthermore, oligonucleotides conesponding to NOVX nucleotide sequences can be prepared by standard synthetic techniques, e.g., using an automated DNA synthesizer. As used herein, the term "oligonucleotide" refers to a series of linked nucleotide residues. A short oligonucleotide sequence may be based on, or designed from, a genomic or cDNA sequence and is used to amplify, confirm, or reveal the presence of an identical, similar or complementary DNA or RNA in a particular cell or tissue. Oligonucleotides comprise a nucleic acid sequence having about 10 nt, 50 nt, or 100 nt in length, preferably about 15 nt to 30 nt in length. In one embodiment of the invention, an oligonucleotide comprising a nucleic acid molecule less than 100 nt in length would further comprise at least 6 contiguous nucleotides of SEQ ID NOS :2n-l, wherein n is an integer between 1 and 62, or a complement thereof. Oligonucleotides may be chemically synthesized and may also be used as probes. In another embodiment, an isolated nucleic acid molecule of the invention comprises a nucleic acid molecule that is a complement of the nucleotide sequence shown in SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, or a portion of this nucleotide sequence (e.g., a fragment that can be used as a probe or primer or a fragment encoding a biologically-active portion of A NOVX polypeptide). A nucleic acid molecule that is complementary to the nucleotide sequence shown SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, is one that is sufficiently complementary to the nucleotide sequence shown SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62,that it can hydrogen bond with few or no mismatches to the nucleotide sequence shown SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, thereby forming a stable duplex. As used herein, the term "complementary" refers to Watson-Crick or Hoogsteen base pairing between nucleotides units of a nucleic acid molecule, and the term "binding" means the physical or chemical interaction between two polypeptides or compounds or associated polypeptides or compounds or combinations thereof. Binding includes ionic, non-ionic, van der Waals, hydrophobic interactions, and the like. A physical interaction can be either direct or indirect. Indirect interactions may be through or due to the effects of another polypeptide or compound. Direct binding refers to interactions that do not take place through, or due to, the effect of another polypeptide or compound, but instead are without other substantial chemical intermediates. "Fragments" provided herein are defined as sequences of at least 6 (contiguous) nucleic acids or at least 4 (contiguous) amino acids, a length sufficient to allow for specific hybridization in the case of nucleic acids or for specific recognition of an epitope in the case of amino acids, and are at most some portion less than a full length sequence. Fragments may be derived from any contiguous portion of a nucleic acid or amino acid sequence of choice.

A full-length NOVX clone is identified as containing an ATG translation start codon and an in-frame stop codon. Any disclosed NOVX nucleotide sequence lacking an ATG start codon therefore encodes a truncated C-terminal fragment of the respective NOVX polypeptide, and requires that the corresponding full-length cDNA extend in the 5' direction of the disclosed sequence. Any disclosed NOVX nucleotide sequence lacking an in-frame stop codon similarly encodes a truncated N-terminal fragment of the respective NOVX polypeptide, and requires that the conesponding full-length cDNA extend in the 3' direction of the disclosed sequence. "Derivatives" are nucleic acid sequences or amino acid sequences formed from the native compounds either directly, by modification, or by partial substitution. "Analogs" are nucleic acid sequences or amino acid sequences that have a structure similar to, but not identical to, the native compound, e.g. they differ from it in respect to certain components or side chains. Analogs may be synthetic or derived from a different evolutionary origin and may have a similar or opposite metabolic activity compared to wild type. Homologs , are nucleic acid sequences or amino acid sequences of a particular gene that are derived from different species.

Derivatives and analogs may be full length or other than full length. Derivatives or analogs of the nucleic acids or proteins of the invention include, but are not limited to, molecules comprising regions that are substantially homologous to the nucleic acids or proteins of the invention, in various embodiments, by at least about 70%, 80%, or 95% identity (with a preferred identity of 80-95%) over a nucleic acid or amino acid sequence of identical size or when compared to an aligned sequence in which the alignment is done by a computer homology program known in the art, or whose encoding nucleic acid is capable of hybridizing to the complement of a sequence encoding the proteins of the invention under stringent, moderately stringent, or low stringent conditions. See e.g. Ausubel, et al., CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley & Sons, New York, NY, 1993, and below. A "homologous nucleic acid sequence" or "homologous amino acid sequence," or variations thereof, refer to sequences characterized by a homology at the nucleotide level or amino acid level as discussed above. Homologous nucleotide sequences include those sequences coding for isoforms of NOVX polypeptides. Isoforms can be expressed in different tissues of the same organism as a result of, for example, alternative splicing of RNA. Alternatively, isoforms can be encoded by different genes. In the invention, homologous nucleotide sequences include nucleotide sequences encoding for A NOVX polypeptide of species other than humans, including, but not limited to vertebrates, and thus can include, e.g., frog, mouse, rat, rabbit, dog, cat, cow, horse, and other organisms. Homologous nucleotide sequences also include, but are not limited to, naturally occurring allelic variations and mutations of the nucleotide sequences set forth herein. A homologous nucleotide sequence does not, however, include the exact nucleotide sequence encoding a human NOVX protein. Homologous nucleic acid sequences include those nucleic acid sequences that encode conservative amino acid substitutions (see below) in SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, as well as a polypeptide possessing NOVX biological activity. Various biological activities of the NOVX proteins are described below.

A NOVX polypeptide is encoded by the open reading frame ("ORF") of a NOVX nucleic acid. An ORF conesponds to a nucleotide sequence that could potentially be translated into a polypeptide. A stretch of nucleic acids comprising an ORF is uninterrupted by a stop codon. An ORF that represents the coding sequence for a full protein begins with an ATG "start" codon and terminates with one of the three "stop" codons, namely, TAA, TAG, or TGA. For the purposes of this invention, an ORF may be any part of a coding sequence, with or without a start codon, a stop codon, or both. For an ORF to be considered as a good candidate for coding for a bonafide cellular protein, a minimum size requirement is often set, e.g., a stretch of DNA that would encode a protein of 50 amino acids or more.

The nucleotide sequences determined from the cloning of the human NOVX genes allows for the generation of probes and primers designed for use in identifying and/or cloning NOVX homologues in other cell types, e.g. from other tissues, as well as NOVX homologues from other vertebrates. The probe/primer typically comprises a substantially purified oligonucleotide. The oligonucleotide typically comprises a region of nucleotide sequence that hybridizes under stringent conditions to at least about 12, 25, 50, 100, 150, 200, 250, 300, 350 or 400 consecutive sense strand nucleotide sequence of SEQ ID

NOS:2n-l, wherein n is an integer between 1 and 62; or an anti-sense strand nucleotide sequence of SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62; or of a naturally occurring mutant of SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62.

Probes based on the human NOVX nucleotide sequences can be used to detect transcripts or genomic sequences encoding the same or homologous proteins. In various embodiments, the probe has a detectable label attached, e.g. the label can be a radioisotope, a fluorescent compound, an enzyme, or an enzyme co-factor. Such probes can be used as a part of a diagnostic test kit for identifying cells or tissues which mis-express A NOVX protein, such as by measuring a level of A NOVX-encoding nucleic acid in a sample of cells from a subject e.g., detecting NOVX mRNA levels or determining whether a genomic NOVX gene has been mutated or deleted.

"A polypeptide having a biologically-active portion of A NOVX polypeptide" refers to polypeptides exhibiting activity similar, but not necessarily identical, an activity of a polypeptide of the invention, including mature forms, as measured in a particular biological assay, with or without dose dependency. A nucleic acid fragment encoding a "biologically-active portion of NOVX" can be prepared by isolating a portion SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, that encodes a polypeptide having A NOVX biological activity (the biological activities of the NOVX proteins are described below), expressing the encoded portion of NOVX protein (e.g., by recombinant expression in vitro) and assessing the activity of the encoded portion of NOVX.

NOVX Nucleic Acid and Polypeptide Variants The invention further encompasses nucleic acid molecules that differ from the nucleotide sequences shown in SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, due to degeneracy of the genetic code and thus encode the same NOVX proteins as that encoded by the nucleotide sequences shown in SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62. In another embodiment, an isolated nucleic acid molecule of the invention has a nucleotide sequence encoding a protein having an amino acid sequence shown in SEQ ID NOS:2n, wherein n is an integer between 1 and 62.

In addition to the human NOVX nucleotide sequences shown in SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, it will be appreciated by those skilled in the art that DNA sequence polymorphisms that lead to changes in the amino acid sequences of the NOVX polypeptides may exist within a population (e.g., the human population). Such genetic polymorphism in the NOVX genes may exist among individuals within a population due to natural allelic variation. As used herein, the terms "gene" and "recombinant gene" refer to nucleic acid molecules comprising an open reading frame (ORF) encoding A NOVX protein, preferably a vertebrate NOVX protein. Such natural allelic variations can typically result in 1-5% variance in the nucleotide sequence of the NOVX genes. Any and all such nucleotide variations and resulting amino acid polymorphisms in the NOVX polypeptides, which are the result of natural allelic variation and that do not alter the functional activity of the NOVX polypeptides, are intended to be within the scope of the invention. Moreover, nucleic acid molecules encoding NOVX proteins from other species, and thus that have a nucleotide sequence that differs from the human SEQ ID NOS:2n-l , wherein n is an integer between 1 and 62, are intended to be within the scope of the invention. Nucleic acid molecules conesponding to natural allelic variants and homologues of the NOVX cDNAs of the invention can be isolated based on their homology to the human NOVX nucleic acids disclosed herein using the human cDNAs, or a portion thereof, as a hybridization probe according to standard hybridization techniques under stringent hybridization conditions. Accordingly, in another embodiment, an isolated nucleic acid molecule of the invention is at least 6 nucleotides in length and hybridizes under stringent conditions to the nucleic acid molecule comprising the nucleotide sequence of SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62. In another embodiment, the nucleic acid is at least 10, 25, 50, 100, 250, 500, 750, 1000, 1500, 2000 or more nucleotides in length. In yet another embodiment, an isolated nucleic acid molecule of the invention hybridizes to the coding region. As used herein, the term "hybridizes under stringent conditions" is intended to describe conditions for hybridization and washing under which nucleotide sequences at least about 65% homologous to each other typically remain hybridized to each other.

Homologs (i.e., nucleic acids encoding NOVX proteins derived from species other than human) or other related sequences (e.g., paralogs) can be obtained by low, moderate or high stringency hybridization with all or a portion of the particular human sequence as a probe using methods well known in the art for nucleic acid hybridization and cloning.

As used herein, the phrase "stringent hybridization conditions" refers to conditions under which a probe, primer or oligonucleotide will hybridize to its target sequence, but to no other sequences. Stringent conditions are sequence-dependent and will be different in different circumstances. Longer sequences hybridize specifically at higher temperatures than shorter sequences. Generally, stringent conditions are selected to be about 5 °C lower than the thermal melting point (Tm) for the specific sequence at a defined ionic strength and pH. The Tm is the temperature (under defined ionic strength, pH and nucleic acid concentration) at which 50% of the probes complementary to the target sequence hybridize to the target sequence at equilibrium. Since the target sequences are generally present at excess at Tm, 50% of the probes are occupied at equilibrium. Typically, stringent conditions will be those in which the salt concentration is less than about 1.0 M sodium ion, typically about 0.01 to 1.0 M sodium ion (or other salts) at pH 7.0 to 8.3 and the temperature is at least about 30 °C for short probes, primers or oligonucleotides (e.g., 10 nt to 50 nt) and at least about 60 °C for longer probes, primers and oligonucleotides. Stringent conditions may also be achieved with the addition of destabilizing agents, such as formamide. Stringent conditions are known to those skilled in the art and can be found in Ausubel, et al., (eds.), CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley & Sons, NY. (1989), 6.3.1-6.3.6. Preferably, the conditions are such that sequences at least about 65%, 70%, 75%, 85%, 90%, 95%, 98%, or 99% homologous to each other typically remain hybridized to each other. A non-limiting example of stringent hybridization conditions are hybridization in a high salt buffer comprising 6X SSC, 50 mM Tris-HCl (pH 7.5), 1 mM EDTA, 0.02% PVP, 0.02% Ficoll, 0.02% BSA, and 500 mg/ml denatured salmon sperm DNA at 65 °C, followed by one or more washes in 0.2X SSC, 0.01% BSA at 50 °C. An isolated nucleic acid molecule of the invention that hybridizes under stringent conditions to the sequences SEQ ID NOS:2n-l , wherein n is an integer between 1 and 62, conesponds to a naturally-occurring nucleic acid molecule. As used herein, a "naturally-occurring" nucleic acid molecule refers to an RNA or DNA molecule having a nucleotide sequence that occurs in nature (e.g., encodes a natural protein).

In a second embodiment, a nucleic acid sequence that is hybridizable to the nucleic acid molecule comprising the nucleotide sequence of SEQ ID NOS:2n-l , wherein n is an integer between 1 and 62, or fragments, analogs or derivatives thereof, under conditions of moderate stringency is provided. A non-limiting example of moderate stringency hybridization conditions are hybridization in 6X SSC, 5X Denhardt's solution, 0.5% SDS and 100 mg/ml denatured salmon sperm DNA at 55 °C, followed by one or more washes in IX SSC, 0.1% SDS at 37 °C. Other conditions of moderate stringency that may be used are well-known within the art. See, e.g., Ausubel, et al. (eds.), 1993, CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley & Sons, NY, and Kriegler, 1990; GENE TRANSFER AND EXPRESSION, A LABORATORY MANUAL, Stockton Press, NY.

In a third embodiment, a nucleic acid that is hybridizable to the nucleic acid molecule comprising the nucleotide sequences SEQ ID NOS:2n-l , wherein n is an integer between 1 and 62, or fragments, analogs or derivatives thereof, under conditions of low stringency, is provided. A non-limiting example of low stringency hybridization conditions are hybridization in 35% formamide, 5X SSC, 50 mM Tris-HCl (pH 7.5), 5 mM EDTA, 0.02% PVP, 0.02% Ficoll, 0.2% BSA, 100 mg/ml denatured salmon sperm DNA, 10% (wt/vol) dextran sulfate at 40 °C, followed by one or more washes in 2X SSC, 25 mM Tris-HCl (pH 7.4), 5 mM EDTA, and 0.1% SDS at 50 °C. Other conditions of low stringency that may be used are well known in the art (e.g., as employed for cross-species hybridizations). See, e.g., Ausubel, et al. (eds.), 1993, CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley & Sons, NY, and Kriegler, 1990, GENE TRANSFER AND EXPRESSION, A LABORATORY MANUAL, Stockton Press, NY; Shilo and Weinberg, 1981. Proc Natl Acad Sci USA 78: 6789-6792.

Conservative Mutations

In addition to naturally-occurring allelic variants of NOVX sequences that may exist in the population, the skilled artisan will further appreciate that changes can be introduced by mutation into the nucleotide sequences SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, thereby leading to changes in the amino acid sequences of the encoded NOVX proteins, without altering the functional ability of the NOVX proteins. For example, nucleotide substitutions leading to amino acid substitutions at "non-essential" amino acid residues can be made in the sequence SEQ ID NOS:2n, wherein n is an integer between 1 and 62. A "non-essential" amino acid residue is a residue that can be altered from the wild-type sequences of the NOVX proteins without altering their biological activity, whereas an "essential" amino acid residue is required for such biological activity. For example, amino acid residues that are conserved among the NOVX proteins of the invention are predicted to be particularly non-amenable to alteration. Amino acids for which conservative substitutions can be made are well known within the art. Another aspect of the invention pertains to nucleic acid molecules encoding

NOVX proteins that contain changes in amino acid residues that are not essential for activity. Such NOVX proteins differ in amino acid sequence from SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, yet retain biological activity. In one embodiment, the isolated nucleic acid molecule comprises a nucleotide sequence encoding a protein, wherein the protein comprises an amino acid sequence at least about 50% homologous to the amino acid sequences SEQ ID NOS:2n, wherein n is an integer between 1 and 62. Preferably, the protein encoded by the nucleic acid molecule is at least about 60% homologous to SEQ ID NOS:2n, wherein n is an integer between 1 and 62; more preferably at least about 70% homologous SEQ ID NOS:2n, wherein n is an integer between 1 and 62; still more preferably at least about 80% homologous to SEQ ID

NOS:2n, wherein n is an integer between 1 and 62; even more preferably at least about 90% homologous to SEQ ID NOS:2n, wherein n is an integer between 1 and 62; and most preferably at least about 95% homologous to SEQ ID NOS:2n, wherein n is an integer between 1 and 62.

An isolated nucleic acid molecule encoding A NOVX protein homologous to the protein of SEQ ID NOS:2n, wherein n is an integer between 1 and 62, can be created by introducing one or more nucleotide substitutions, additions or deletions into the nucleotide sequence of SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, such that one or more amino acid substitutions, additions or deletions are introduced into the encoded protein.

Mutations can be introduced into SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, by standard techniques, such as site-directed mutagenesis and

PCR-mediated mutagenesis. Preferably, conservative amino acid substitutions are made at one or more predicted, non-essential amino acid residues. A "conservative amino acid substitution" is one in which the amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues having similar side chains have been defined within the art. These families include amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Thus, a predicted non-essential amino acid residue in the NOVX protein is replaced with another amino acid residue from the same side chain family. Alternatively, in another embodiment, mutations can be introduced randomly along all or part of A NOVX coding sequence, such as by saturation mutagenesis, and the resultant mutants can be screened for NOVX biological activity to identify mutants that retain activity. Following mutagenesis SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, the encoded protein can be expressed by any recombinant technology known in the art and the activity of the protein can be determined.

The relatedness of amino acid families may also be determined based on side chain interactions. Substituted amino acids may be fully conserved "strong" residues or fully conserved "weak" residues. The "strong" group of conserved amino acid residues may be any one of the following groups: STA, NEQK, NHQK, NDEQ, QHRK, MILV, MILF, HY, FYW, wherein the single letter amino acid codes are grouped by those amino acids that may be substituted for each other. Likewise, the "weak" group of conserved residues may be any one of the following: CSA, ATV, SAG, STNK, STPA, SGND, SNDEQK, NDEQHK, NEQHRK, HFY, wherein the letters within each group represent the single letter amino acid code. In one embodiment, a mutant NOVX protein can be assayed for (i) the ability to form protei protein interactions with other NOVX proteins, other cell-surface proteins, or biologically-active portions thereof, (ii) complex formation between a mutant NOVX protein and A NOVX ligand; or (iii) the ability of a mutant NOVX protein to bind to an intracellular target protein or biologically-active portion thereof; (e.g. avidin proteins). In yet another embodiment, a mutant NOVX protein can be assayed for the ability to regulate a specific biological function (e.g., regulation of insulin release).

Antisense Nucleic Acids

Another aspect of the invention pertains to isolated antisense nucleic acid molecules that are hybridizable to or complementary to the nucleic acid molecule comprising the nucleotide sequence of SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, or fragments, analogs or derivatives thereof. An "antisense" nucleic acid comprises a nucleotide sequence that is complementary to a "sense" nucleic acid encoding a protein (e.g., complementary to the coding strand of a double-stranded cDNA molecule or complementary to an mRNA sequence). In specific aspects, antisense nucleic acid molecules are provided that comprise a sequence complementary to at least about 10, 25, 50, 100, 250 or 500 nucleotides or an entire NOVX coding strand, or to only a portion thereof. Nucleic acid molecules encoding fragments, homologs, derivatives and analogs of A NOVX protein of SEQ ID NOS:2n, wherein n is an integer between 1 and 62, or antisense nucleic acids complementary to A NOVX nucleic acid sequence of SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, are additionally provided.

In one embodiment, an antisense nucleic acid molecule is antisense to a "coding region" of the coding strand of a nucleotide sequence encoding A NOVX protein. The term "coding region" refers to the region of the nucleotide sequence comprising codons, which are translated into amino acid residues. In another embodiment, the antisense nucleic acid molecule is antisense to a "noncoding region" of the coding strand of a nucleotide sequence encoding the NOVX protein. The term "noncoding region" refers to 5' and 3' sequences, which flank the coding region that are not translated into amino acids (i.e., also referred to as 5' and 3' untranslated regions).

Given the coding strand sequences encoding the NOVX protein disclosed herein, antisense nucleic acids of the invention can be designed according to the rules of Watson and Crick or Hoogsteen base pairing. The antisense nucleic acid molecule can be complementary to the entire coding region of NOVX mRNA, but more preferably is an oligonucleotide that is antisense to only a portion of the coding or noncoding region of NOVX mRNA. For example, the antisense oligonucleotide can be complementary to the region sunounding the translation start site of NOVX mRNA. An antisense oligonucleotide can be, for example, about 5, 10, 15, 20, 25, 30, 35, 40, 45 or 50 nucleotides in length. An antisense nucleic acid of the invention can be constructed using chemical synthesis or enzymatic ligation reactions using procedures known in the art. For example, an antisense nucleic acid (e.g., an antisense oligonucleotide) can be chemically synthesized using naturally occurring nucleotides or variously modified nucleotides designed to increase the biological stability of the molecules or to increase the physical stability of the duplex formed between the antisense and sense nucleic acids (e.g., phosphorothioate derivatives and acridine substituted nucleotides can be used).

Examples of modified nucleotides that can be used to generate the antisense nucleic acid include: 5-fluorouracil, 5-bromouracil, 5-chlorouracil, 5-iodouracil, hypoxanthine, xanthine, 4-acetylcytosine, 5-(carboxyhydroxylmethyl) uracil, beta-D-mannosylqueosine, 5-carboxymethyIaminomethyI-2-thiouridine, 5-carboxymethylaminomethyluracil, dihydrouracil, beta-D-galactosylqueosine, inosine, N6-isopentenyladenine, 1-methylguanine, 1-methylinosine, 2,2-dimethylguanine, 2-methyladenine, 2-methylguanine, 3-methylcytosihe, 5-methyl cytosine, N6-adenine, 7-methylguanine, 5-methylaminomethyluracil, 5-memoxyaminome yl-2-thiouracil,

5'-methoxycarboxymethyluracil, 5-methoxyuracil, 2-methylthio-N6-isopentenyladenine, uracil-5-oxyacetic acid (v), wybutoxosine, pseudouracil, queosine, 2-thiocytosine, 5-methyl-2-thiouracil, 2-thiouracil, 4-thiouracil, 5-methyluracil, uracil-5-oxyacetic acid methylester, uracil-5-oxyacetic acid (v), 5-methyl-2-thiouracil, 3-(3-amino-3-N-2-carboxypropyl) uracil, (acp3)w, and 2,6-diaminopurine. Alternatively, the antisense nucleic acid can be produced biologically using an expression vector into which a nucleic acid has been subcloned in an antisense orientation (i.e., RNA transcribed from the inserted nucleic acid will be of an antisense orientation to a target nucleic acid of interest, described further in the following subsection).

The antisense nucleic acid molecules of the invention are typically administered to a subject or generated in situ such that they hybridize with or bind to cellular mRNA and/or genomic DNA encoding A NOVX protein to thereby inhibit expression of the protein (e.g., by inhibiting transcription and/or translation). The hybridization can be by conventional nucleotide complementarity to form a stable duplex, or, for example, in the case of an antisense nucleic acid molecule that binds to DNA duplexes, through specific interactions in the major groove of the double helix. An example of a route of administration of antisense nucleic acid molecules of the invention includes direct injection at a tissue site. Alternatively, antisense nucleic acid molecules can be modified to target selected cells and then administered systemically. For example, for systemic administration, antisense molecules can be modified such that they specifically bind to receptors or antigens expressed on a selected cell surface (e.g., by linking the antisense nucleic acid molecules to peptides or antibodies that bind to cell surface receptors or antigens). The antisense nucleic acid molecules can also be delivered to cells using the vectors described herein. To achieve sufficient nucleic acid molecules, vector constructs in which the antisense nucleic acid molecule is placed under the control of a strong pol II or pol III promoter are prefened. In yet another embodiment, the antisense nucleic acid molecule of the invention is an cc-anomeric nucleic acid molecule. A α-anomeric nucleic acid molecule forms specific double-stranded hybrids with complementary RNA in which, contrary to the usual β-units, the strands run parallel to each other. See, e.g., Gaultier, ei al, 1987. Nucl. Acids Res. 15: 6625-6641. The antisense nucleic acid molecule can also comprise a 2'-o-methylribonucleotide (See, e.g., Inoue, et al. 1987. Nucl. Acids Res. 15: 6131-6148) or a chimeric RNA-DNA analogue (See, e.g., Inoue, et al, 1987. FEBSLett. 215: 327-330. Ribozymes and PNA Moieties

Nucleic acid modifications include, by way of non-limiting example, modified bases, and nucleic acids whose sugar phosphate backbones are modified or derivatized. These modifications are carried out at least in part to enhance the chemical stability of the modified nucleic acid, such that they may be used, for example, as antisense binding nucleic acids in therapeutic applications in a subject.

In one embodiment, an antisense nucleic acid of the invention is a ribozyme. Ribozymes are catalytic RNA molecules with ribonuclease activity that are capable of cleaving a single-stranded nucleic acid, such as an mRNA, to which they have a complementary region. Thus, ribozymes (e.g., hammerhead ribozymes as described in Haselhoff and Gerlach 1988. Nature 334: 585-591) can be used to catalytically cleave NOVX mRNA transcripts to thereby inhibit translation of NOVX mRNA. A ribozyme having specificity for a NOVX-encoding nucleic acid can be designed based upon the nucleotide sequence of A NOVX cDNA disclosed herein (i.e., SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62). For example, a derivative of a Tetrahymena L-19 IVS RNA can be constructed in which the nucleotide sequence of the active site is complementary to the nucleotide sequence to be cleaved in a NOVX-encoding mRNA. See, e.g., U.S. Patent 4,987,071 to Cech, et al. and U.S. Patent 5,116,742 to Cech, et al. NOVX mRNA can also be used to select a catalytic RNA having a specific ribonuclease activity from a pool of RNA molecules. See, e.g., Bartel et al., (1993) Science 261:1411-1418.

Alternatively, NOVX gene expression can be inhibited by targeting nucleotide sequences complementary to the regulatory region of the NOVX nucleic acid (e.g., the NOVX promoter and or enhancers) to form triple helical structures that prevent transcription of the NOVX gene in target cells. See, e.g., Helene, 1991. Anticancer Drug Des. 6: 569-84; Helene, etal. 1992. Ann. NY. Acad. Sci. 660: 27-36; Maher, 1992. Bioassays 14: 807-15.

In various embodiments, the NOVX nucleic acids can be modified at the base moiety, sugar moiety or phosphate backbone to improve, e.g., the stability, hybridization, or solubility of the molecule. For example, the deoxyribose phosphate backbone of the nucleic acids can be modified to generate peptide nucleic acids. See, e.g., Hyrup, et al., 1996. BioorgMed Chem 4: 5-23. As used herein, the terms "peptide nucleic acids" or "PNAs" refer to nucleic acid mimics (e.g., DNA mimics) in which the deoxyribose phosphate backbone is replaced by a pseudopeptide backbone and only the four natural nucleobases are retained. The neutral backbone of PNAs has been shown to allow for specific hybridization to DNA and RNA under conditions of low ionic strength. The synthesis of PNA oligomers can be performed using standard solid phase peptide synthesis protocols as described in Hyrup, et al., 1996. supra; Perry-O'Keefe, et al., 1996. Proc. Natl. Acad. Sci. USA 93: 14670-14675.

PNAs of NOVX can be used in therapeutic and diagnostic applications. For example, PNAs can be used as antisense or antigene agents for sequence-specific modulation of gene expression by, e.g., inducing transcription or translation anest or inhibiting replication. PNAs of NOVX can also be used, for example, in the analysis of single base pair mutations in a gene (e.g., PNA directed PCR clamping; as artificial restriction enzymes when used in combination with other enzymes, e.g., Si nucleases (See, Hyrup, et al, \996.supra); or as probes or primers for DNA sequence and hybridization (See, Hyrup, et al, 1996, supra; Perry-O'Keefe, et al, 1996. supra). In another embodiment, PNAs of NOVX can be modified, e.g. , to enhance their stability or cellular uptake, by attaching lipophilic or other helper groups to PNA, by the formation of PNA-DNA chimeras, or by the use of liposomes or other techniques of drug delivery known in the art. For example, PNA-DNA chimeras of NOVX can be generated that may combine the advantageous properties of PNA and DNA. Such chimeras allow DNA recognition enzymes (e.g. , RNase H and DNA polymerases) to interact with the DNA portion while the PNA portion would provide high binding affinity and specificity. PNA-DNA chimeras can be linked using linkers of appropriate lengths selected in terms of base stacking, number of bonds between the nucleobases, and orientation (see, Hyrup, et al., 1996. supra). The synthesis of PNA-DNA chimeras can be performed as described in Hyrup, et al, 1996. supra and Finn, et al, 1996. Nucl Acids Res 24: 3357-3363. For example, a DNA chain can be synthesized on a solid support using standard phosphoramidite coupling chemistry, and modified nucleoside analogs, e.g., 5'-(4-methoxytrityl)amino-5'-deoxy-thymidine phosphoramidite, can be used between the PNA and the 5* end of DNA. See, e.g., Mag, et al, 1989. Nucl Acid Res 17: 5973-5988. PNA monomers are then coupled in a step wise manner to produce a chimeric molecule with a 5' PNA segment and a 3' DNA segment. See, e.g., Finn, et al., 1996. supra. Alternatively, chimeric molecules can be synthesized with a 5' DNA segment and a 3' PNA segment. See, e.g., Petersen, et al, 1975. Bioorg. Med. Chem. Lett. 5: 1119-11124. In other embodiments, the oligonucleotide may include other appended groups such as peptides (e.g., for targeting host cell receptors in vivo), or agents facilitating transport across the cell membrane (see, e.g., Letsinger, et al, 1989. Proc. Natl. Acad. Sci. U.S.A. 86: 6553-6556; Lemaitre, et al, 1987. Proc. Natl. Acad. Sci. 84: 648-652; PCT Publication No. WO88/09810) or the blood-brain barrier (see, e.g. , PCT Publication No. WO 89/10134). In addition, oligonucleotides can be modified with hybridization triggered cleavage agents (see, e.g., Krol, et al, 1988. BioTechniques 6:958-976) or intercalating agents (see, e.g., Zon, 1988. Pharm. Res. 5: 539-549). To this end, the oligonucleotide may be conjugated to another molecule, e.g., a peptide, a hybridization triggered cross-linking agent, a transport agent, a hybridization-triggered cleavage agent, and the like.

NOVX Polypeptides

A polypeptide according to the invention includes a polypeptide including the amino acid sequence of NOVX polypeptides whose sequences are provided in SEQ ID NOS:2n, wherein n is an integer between 1 and 62. The invention also includes a mutant or variant protein any of whose residues may be changed from the conesponding residues shown in SEQ ID NOS:2n, wherein n is an integer between 1 and 62, while still encoding a protein that maintains its NOVX activities and physiological functions, or a functional fragment thereof.

In general, A NOVX variant that preserves NOVX-like function includes any variant in which residues at a particular position in the sequence have been substituted by other amino acids, and further include the possibility of inserting an additional residue or residues between two residues of the parent protein as well as the possibility of deleting one or more residues from the parent sequence. Any amino acid substitution, insertion, or deletion is encompassed by the invention. In favorable circumstances, the substitution is a conservative substitution as defined above.

One aspect of the invention pertains to isolated NOVX proteins, and biologically-active portions thereof, or derivatives, fragments, analogs or homologs thereof. Also provided are polypeptide fragments suitable for use as immunogens to raise anti-NOVX antibodies. In one embodiment, native NOVX proteins can be isolated from cells or tissue sources by an appropriate purification scheme using standard protein purification techniques. In another embodiment, NOVX proteins are produced by recombinant DNA techniques. Alternative to recombinant expression, A NOVX protein or polypeptide can be synthesized chemically using standard peptide synthesis techniques.

An "isolated" or "purified" polypeptide or protein or biologically-active portion thereof is substantially free of cellular material or other contaminating proteins from the cell or tissue source from which the NOVX protein is derived, or substantially free from chemical precursors or other chemicals when chemically synthesized. The language "substantially free of cellular material" includes preparations of NOVX proteins in which the protein is separated from cellular components of the cells from which it is isolated or recombinantly-produced. In one embodiment, the language "substantially free of cellular material" includes preparations of NOVX proteins having less than about 30% (by dry weight) of non-NOVX proteins (also refened to herein as a "contaminating protein"), more preferably less than about 20% of non-NOVX proteins, still more preferably less than about 10% of non-NOVX proteins, and most preferably less than about 5% of non-NOVX proteins. When the NOVX protein or biologically-active portion thereof is recombinantly-produced, it is also preferably substantially free of culture medium, i.e., culture medium represents less than about 20%, more preferably less than about 10%, and most preferably less than about 5% of the volume of the NOVX protein preparation. The language "substantially free of chemical precursors or other chemicals" includes preparations of NOVX proteins in which the protein is separated from chemical precursors or other chemicals that are involved in the synthesis of the protein. In one embodiment, the language "substantially free of chemical precursors or other chemicals" includes preparations of NOVX proteins having less than about 30% (by dry weight) of chemical precursors or non-NOVX chemicals, more preferably less than about 20% chemical precursors or non-NOVX chemicals, still more preferably less than about 10% chemical precursors or non-NOVX chemicals, and most preferably less than about 5% chemical precursors or non-NOVX chemicals.

Biologically-active portions of NOVX proteins include peptides comprising amino acid sequences sufficiently homologous to or derived from the amino acid sequences of the NOVX proteins (e.g., the amino acid sequence shown in SEQ ID NOS:2n, wherein n is an integer between 1 and 62) that include fewer amino acids than the full-length NOVX proteins, and exhibit at least one activity of A NOVX protein. Typically, biologically-active portions comprise a domain or motif with at least one activity of the NOVX protein. A biologically-active portion of A NOVX protein can be a polypeptide which is, for example, 10, 25, 50, 100 or more amino acid residues in length.

Moreover, other biologically-active portions, in which other regions of the protein are deleted, can be prepared by recombinant techniques and evaluated for one or more of the functional activities of a native NOVX protein.

In an embodiment, the NOVX protein has an amino acid sequence shown SEQ ID NOS:2n, wherein n is an integer between 1 and 62. In other embodiments, the NOVX protein is substantially homologous to SEQ ID NOS:2n, wherein n is an integer between 1 and 62, and retains the functional activity of the protein of SEQ ID NOS:2n, wherein n is an integer between 1 and 62, yet differs in amino acid sequence due to natural allelic variation or mutagenesis, as described in detail, below. Accordingly, in another embodiment, the NOVX protein is a protein that comprises an amino acid sequence at least about 45% homologous to the amino acid sequence SEQ ID NOS:2n, wherein n is an integer between 1 and 62, and retains the functional activity of the NOVX proteins of SEQ ID NOS:2n, wherein n is an integer between 1 and 62.

DETERMINING HOMOLOGY BETWEEN Two OR MORE SEQUENCES

To determine the percent homology of two amino acid sequences or of two nucleic acids, the sequences are aligned for optimal comparison purposes (e.g., gaps can be introduced in the sequence of a first amino acid or nucleic acid sequence for optimal alignment with a second amino or nucleic acid sequence). The amino acid residues or nucleotides at conesponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the conesponding position in the second sequence, then the molecules are homologous at that position (i.e., as used herein amino acid or nucleic acid "homology" is equivalent to amino acid or nucleic acid "identity").

The nucleic acid sequence homology may be determined as the degree of identity between two sequences. The homology may be determined using computer programs known in the art, such as GAP software provided in the GCG program package. See, Needleman and Wunsch, 1970. JMol Biol 48: 443-453. Using GCG GAP software with the following settings for nucleic acid sequence comparison: GAP creation penalty of 5.0 and GAP extension penalty of 0.3, the coding region of the analogous nucleic acid sequences refened to above exhibits a degree of identity preferably of at least 70%, 75%, 80%, 85%, 90%, 95%, 98%, or 99%, with the CDS (encoding) part of the DNA sequence shown in SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62.

The term "sequence identity" refers to the degree to which two polynucleotide or polypeptide sequences are identical on a residue-by-residue basis over a particular region of comparison. The term "percentage of sequence identity" is calculated by comparing two optimally aligned sequences over that region of comparison, determining the number of positions at which the identical nucleic acid base (e.g., A, T, C, G, U, or I, in the case of nucleic acids) occurs in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the region of comparison (i. e., the window size), and multiplying the result by 100 to yield the percentage of sequence identity. The term "substantial identity" as used herein denotes a characteristic of a polynucleotide sequence, wherein the polynucleotide comprises a sequence that has at least 80 percent sequence identity, preferably at least 85 percent identity and often 90 to 95 percent sequence identity, more usually at least 99 percent sequence identity as compared to a reference sequence over a comparison region.

CHIMERIC AND FUSION PROTEINS

The invention also provides NOVX chimeric or fusion proteins. As used herein, A NOVX "chimeric protein" or "fusion protein" comprises A NOVX polypeptide operatively-linked to a non-NOVX polypeptide. An "NOVX polypeptide" refers to a polypeptide having an amino acid sequence conesponding to A NOVX protein SEQ ID NOS:2n, wherein n is an integer between 1 and 62, whereas a "non-NOVX polypeptide" refers to a polypeptide having an amino acid sequence conesponding to a protein that is not substantially homologous to the NOVX protein, e.g., a protein that is different from the NOVX protein and that is derived from the same or a different organism. Within A NOVX fusion protein the NOVX polypeptide can conespond to all or a portion of A NOVX protein. In one embodiment, A NOVX fusion protein comprises at least one biologically active portion of A NOVX protein. In another embodiment, A NOVX fusion protein comprises at least two biologically active portions of A NOVX protein. In yet another embodiment, A NOVX fusion protein comprises at least three biologically active portions of A NOVX protein. Within the fusion protein, the term "operatively-linked" is intended to indicate that the NOVX polypeptide and the non-NOVX polypeptide are fused in-frame with one another. The non-NOVX polypeptide can be fused to the N-terminus or C-terminus of the NOVX polypeptide.

In one embodiment, the fusion protein is a GST-NOVX fusion protein in which the NOVX sequences are fused to the C-terminus of the GST (glutathione S-transferase) sequences. Such fusion proteins can facilitate the purification of recombinant NOVX polypeptides.

In another embodiment, the fusion protein is A NOVX protein containing a heterologous signal sequence at its N-terminus. In certain host cells (e.g., mammalian host cells), expression and/or secretion of NOVX can be increased through use of a heterologous signal sequence.

In yet another embodiment, the fusion protein is a NOVX-immunoglobulin fusion protein in which the NOVX sequences are fused to sequences derived from a member of the immunoglobulin protein family. The NOVX-immunoglobulin fusion proteins of the invention can be incorporated into pharmaceutical compositions and administered to a subject to inhibit an interaction between A NOVX ligand and A NOVX protein on the surface of a cell, to thereby suppress NOVX-mediated signal transduction in vivo. The NOVX-immunoglobulin fusion proteins can be used to affect the bioavailability of A NOVX cognate ligand. Inhibition of the NOVX ligand/NOVX interaction may be useful therapeutically for both the treatment of proliferative and differentiative disorders, as well as modulating (e.g. promoting or inhibiting) cell survival. Moreover, the

NOVX-immunoglobulin fusion proteins of the invention can be used as immunogens to produce anti-NOVX antibodies in a subject, to purify NOVX ligands, and in screening assays to identify molecules that inhibit the interaction of NOVX with A NOVX ligand. A NOVX chimeric or fusion protein of the invention can be produced by standard recombinant DNA techniques. For example, DNA fragments coding for the different polypeptide sequences are ligated together in-frame in accordance with conventional techniques, e.g., by employing blunt-ended or stagger-ended termini for ligation, restriction enzyme digestion to provide for appropriate termini, filling-in of cohesive ends as appropriate, alkaline phosphatase treatment to avoid undesirable joining, and enzymatic ligation. In another embodiment, the fusion gene can be synthesized by conventional techniques including automated DNA synthesizers. Alternatively, PCR amplification of gene fragments can be carried out using anchor primers that give rise to complementary overhangs between two consecutive gene fragments that can subsequently be annealed and reamplified to generate a chimeric gene sequence (see, e.g., Ausubel, et al. (eds.) CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley & Sons, 1992). Moreover, many expression vectors are commercially available that already encode a fusion moiety (e.g., a GST polypeptide). A NOVX-encoding nucleic acid can be cloned into such an expression vector such that the fusion moiety is linked in-frame to the NOVX protein.

NOVX AGONISTS AND ANTAGONISTS

The invention also pertains to variants of the NOVX proteins that function as either NOVX agonists (i.e., mimetics) or as NOVX antagonists. Variants of the NOVX protein can be generated by mutagenesis (e.g. , discrete point mutation or truncation of the NOVX protein). An agonist of the NOVX protein can retain substantially the same, or a subset of, the biological activities of the naturally occurring form of the NOVX protein. An antagonist of the NOVX protein can inhibit one or more of the activities of the naturally occuning form of the NOVX protein by, for example, competitively binding to a downstream or upstream member of a cellular signaling cascade, which includes the NOVX protein. Thus, specific biological effects can be elicited by treatment with a variant of limited function. In one embodiment, treatment of a subject with a variant having a subset of the biological activities of the naturally occurring form of the protein has fewer side effects in a subject relative to treatment with the naturally occurring form of the NOVX proteins.

Variants of the NOVX proteins that function as either NOVX agonists (i.e., mimetics) or as NOVX antagonists can be identified by screening combinatorial libraries of mutants (e.g., truncation mutants) of the NOVX proteins for NOVX protein agonist or antagonist activity. In one embodiment, a variegated library of NOVX variants is generated by combinatorial mutagenesis at the nucleic acid level and is encoded by a variegated gene library. A variegated library of NOVX variants can be produced by, for example, enzymatically ligating a mixture of synthetic oligonucleotides into gene sequences such that a degenerate set of potential NOVX sequences is expressible as individual polypeptides, or alternatively, as a set of larger fusion proteins (e.g., for phage display) containing the set of NOVX sequences therein. There are a variety of methods, which can be used to produce libraries of potential NOVX variants from a degenerate oligonucleotide sequence. Chemical synthesis of a degenerate gene sequence can be performed in an automatic DNA synthesizer, and the synthetic gene then ligated into an appropriate expression vector. Use of a degenerate set of genes allows for the provision, in one mixture, of all of the sequences encoding the desired set of potential NOVX sequences. Methods for synthesizing degenerate oligonucleotides are well known within the art. See, e.g., Narang, 1983. Tetrahedron 39: 3; Itakura, et al., 1984. Anna. Rev. Biochem. 53: 323; Itakura, et al, 1984. Science 198: 1056; Ike, et al, 1983. Nucl. Acids Res. 11: 477.

POLYPEPTIDE LIBRARIES

In addition, libraries of fragments of the NOVX protein coding sequences can be used to generate a variegated population of NOVX fragments for screening and subsequent selection of variants of A NOVX protein. In one embodiment, a library of coding sequence fragments can be generated by treating a double stranded PCR fragment of A NOVX coding sequence with a nuclease under conditions wherein nicking occurs only about once per molecule, denaturing the double stranded DNA, renaturing the DNA to form double-stranded DNA that can include sense/antisense pairs from different nicked products, removing single stranded portions from reformed duplexes by treatment with Si nuclease, and ligating the resulting fragment library into an expression vector. By this method, expression libraries can be derived which encodes N-terminal and internal fragments of various sizes of the NOVX proteins. Various techniques are known in the art for screening gene products of combinatorial libraries made by point mutations or truncation, and for screening cDNA libraries for gene products having a selected property. Such techniques are adaptable for rapid screening of the gene libraries generated by the combinatorial mutagenesis of NOVX proteins. The most widely used techniques, which are amenable to high throughput analysis, for screening large gene libraries typically include cloning the gene library into replicable expression vectors, transforming appropriate cells with the resulting library of vectors, and expressing the combinatorial genes under conditions in which detection of a desired activity facilitates isolation of the vector encoding the gene whose product was detected. Recursive ensemble mutagenesis (REM), a new technique that enhances the frequency of functional mutants in the libraries, can be used in combination with the screening assays to identify NOVX variants. See, e.g., Arkin and Yourvan, 1992. Proc. Natl. Acad. Sci. USA 89: 7811-7815; Delgrave, et al, 1993. Protein Engineering 6:327-331. NOVX Antibodies

The term "antibody" as used herein refers to immunoglobulin molecules and immunologically active portions of immunoglobulin (Ig) molecules, i.e., molecules that contain an antigen-binding site that specifically binds (immunoreacts with) an antigen. Such antibodies include, but are not limited to, polyclonal, monoclonal, chimeric, single chain, F_ab, F_ab' and F_(ab_')₂ fragments, and an F_ab expression library. In general, antibody molecules obtained from humans relates to any of the classes IgG, IgM, IgA, IgE and IgD, which differ from one another by the nature of the heavy chain present in the molecule. Certain classes have subclasses as well, such as IgG_1? IgG₂, and others. Furthermore, in humans, the light chain may be a kappa chain or a lambda chain. Reference herein to antibodies includes a reference to all such classes, subclasses and types of human antibody species.

An isolated protein of the invention intended to serve as an antigen, or a portion or fragment thereof, can be used as an immunogen to generate antibodies that immunospecifically bind the antigen, using standard techniques for polyclonal and monoclonal antibody preparation. The full-length protein can be used or, alternatively, the invention provides antigenic peptide fragments of the antigen for use as immunogens. An antigenic peptide fragment comprises at least 6 amino acid residues of the amino acid sequence of the full length protein, such as an amino acid sequence shown in SEQ ID NOs: 2n, wherein n is an integer between 1 and 62, and encompasses an epitope thereof such that an antibody raised against the peptide forms a specific immune complex with the full length protein or with any fragment that contains the epitope. Preferably, the antigenic peptide comprises at least 10 amino acid residues, or at least 15 amino acid residues, or at least 20 amino acid residues, or at least 30 amino acid residues. Prefened epitopes encompassed by the antigenic peptide are regions of the protein that are located on its surface; commonly these are hydrophilic regions.

In certain embodiments of the invention, at least one epitope encompassed by the antigenic peptide is a region of NOVX that is located on the surface of the protein, e.g., a hydrophilic region. A hydrophobicity analysis of the human NOVX protein sequence will indicate which regions of a NOVX polypeptide are particularly hydrophilic and, therefore, are likely to encode surface residues useful for targeting antibody production. As a means for targeting antibody production, hydropathy plots showing regions of hydrophilicity and hydrophobicity may be generated by any method well known in the art, including, for example, the Kyte Doolittle or the Hopp Woods methods, either with or without Fourier transformation. See, e.g., Hopp and Woods, 1981, Proc. Nat. Acad. Sci. USA 78: 3824-3828; Kyte and Doolittle 1982, J. Mol Biol. 157: 105-142, each incorporated herein by reference in their entirety. Antibodies that are specific for one or more domains within an antigenic protein, or derivatives, fragments, analogs or homologs thereof, are also provided herein.

A protein of the invention, or a derivative, fragment, analog, homolog or ortholog thereof, may be utilized as an immunogen in the generation of antibodies that immunospecifically bind these protein components.

Various procedures known within the art may be used for the production of polyclonal or monoclonal antibodies directed against a protein of the invention, or against derivatives, fragments, analogs homologs or orthologs thereof (see, for example, Antibodies: A Laboratory Manual, Harlow E, and Lane D, 1988, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, incorporated herein by reference). Some of these antibodies are discussed below.

Polyclonal Antibodies

For the production of polyclonal antibodies, various suitable host animals (e.g., rabbit, goat, mouse or other mammal) may be immunized by one or more injections with the native protein, a synthetic variant thereof, or a derivative of the foregoing. An appropriate immunogenic preparation can contain, for example, the naturally occurring iirimunogenic protein, a chemically synthesized polypeptide representing the immunogenic protein, or a recombinantly expressed immunogenic protein. Furthermore, the protein may be conjugated to a second protein known to be immunogenic in the mammal being immunized. Examples of such immunogenic proteins include but are not limited to keyhole limpet hemocyanin, serum albumin, bovine thyroglobulin, and soybean trypsin inhibitor. The preparation can further include an adjuvant. Various adjuvants used to increase the immunological response include, but are not limited to, Freund's (complete and incomplete), mineral gels (e.g., aluminum hydroxide), surface active substances (e.g., lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, dinitrophenol, etc.), adjuvants usable in humans such as Bacille Calmette-Guerin and Corynebacterium parvum, or similar immunostimulatory agents. Additional examples of adjuvants which can be employed include MPL-TDM adjuvant (monophosphoryl Lipid A, synthetic trehalose dicorynomycolate).

The polyclonal antibody molecules directed against the immunogenic protein can be isolated from the mammal (e.g., from the blood) and further purified by well known techniques, such as affinity chromatography using protein A or protein G, which provide primarily the IgG fraction of immune serum. Subsequently, or alternatively, the specific antigen which is the target of the immunoglobulin sought, or an epitope thereof, may be immobilized on a column to purify the immune specific antibody by immunoaffinity chromatography. Purification of immunoglobulins is discussed, for example, by D. Wilkinson (The Scientist, published by The Scientist, Inc., Philadelphia PA, Vol. 14, No. 8 (April 17, 2000), pp. 25-28).

Monoclonal Antibodies

The term "monoclonal antibody" (MAb) or "monoclonal antibody composition", as used herein, refers to a population of antibody molecules that contain only one molecular species of antibody molecule consisting of a unique light chain gene product and a unique heavy chain gene product. In particular, the complementarity determining regions (CDRs) of the monoclonal antibody are identical in all the molecules of the population. MAbs thus contain an antigen binding site capable of immunoreacting with a particular epitope of the antigen characterized by a unique binding affinity for it.

Monoclonal antibodies can be prepared using hybridoma methods, such as those described by Kohler and Milstein, Nature, 256:495 (1975). In a hybridoma method, a mouse, hamster, or other appropriate host animal, is typically immunized with an immunizing agent to elicit lymphocytes that produce or are capable of producing antibodies that will specifically bind to the immunizing agent. Alternatively, the lymphocytes can be immunized in vitro.

The immunizing agent will typically include the protein antigen, a fragment thereof or a fusion protein thereof. Generally, either peripheral blood lymphocytes are used if cells of human origin are desired, or spleen cells or lymph node cells are used if non-human mammalian sources are desired. The lymphocytes are then fused with an immortalized cell line using a suitable fusing agent, such as polyethylene glycol, to form a hybridoma cell [Goding, Monoclonal Antibodies: Principles and Practice, Academic Press, (1986) pp. 59-103]. Immortalized cell lines are usually transformed mammalian cells, particularly myeloma cells of rodent, bovine and human origin. Usually, rat or mouse myeloma cell lines are employed. The hybridoma cells can be cultured in a suitable culture medium that preferably contains one or more substances that inhibit the growth or survival of the unfused, immortalized cells. For example, if the parental cells lack the enzyme hypoxanthine guanine phosphoribosyl transferase (HGPRT or HPRT), the culture medium for the hybridomas typically will include hypoxanthine, aminopterin, and thymidine ("HAT medium"), which substances prevent the growth of HGPRT-deficient cells.

Preferred immortalized cell lines are those that fuse efficiently, support stable high level expression of antibody by the selected antibody-producing cells, and are sensitive to a medium such as HAT medium. More prefened immortalized cell lines are murine myeloma lines, which can be obtained, for instance, from the Salk Institute Cell Distribution Center, San Diego, California and the American Type Culture Collection, Manassas, Virginia. Human myeloma and mouse-human heteromyeloma cell lines also have been described for the production of human monoclonal antibodies [Kozbor,

Immunol.. 133:3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, Marcel Dekker, Inc., New York, (1987) pp. 51-63].

The culture medium in which the hybridoma cells are cultured can then be assayed for the presence of monoclonal antibodies directed against the antigen. Preferably, the binding specificity of monoclonal antibodies produced by the hybridoma cells is determined by immunoprecipitation or by an in vitro binding assay, such as radioimmunoassay (RIA) or enzyme-linked immunoabsorbent assay (ELISA). Such techniques and assays are known in the art. The binding affinity of the monoclonal antibody can, for example, be determined by the Scatchard analysis of Munson and Pollard, Anal. Biochem., 107:220 (1980). It is an objective, especially important in therapeutic applications of monoclonal antibodies, to identify antibodies having a high degree of specificity and a high binding affinity for the target antigen.

After the desired hybridoma cells are identified, the clones can be subcloned by limiting dilution procedures and grown by standard methods (Goding, 1986). Suitable culture media for this purpose include, for example, Dulbecco's Modified Eagle's Medium and RPMI-1640 medium. Alternatively, the hybridoma cells can be grown in vivo as ascites in a mammal. The monoclonal antibodies secreted by the subclones can be isolated or purified from the culture medium or ascites fluid by conventional immunoglobulin purification procedures such as, for example, protein A-Sepharose, hydroxylapatite chromatography, gel electrophoresis, dialysis, or affinity chromatography. The monoclonal antibodies can also be made by recombinant DNA methods, such as those described in U.S. Patent No. 4,816,567. DNA encoding the monoclonal antibodies of the invention can be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that are capable of binding specifically to genes encoding the heavy and light chains of murine antibodies). The hybridoma cells of the invention serve as a prefened source of such DNA. Once isolated, the DNA can be placed into expression vectors, which are then transfected into host cells such as simian COS cells, Chinese hamster ovary (CHO) cells, or myeloma cells that do not otherwise produce immunoglobulin protein, to obtain the synthesis of monoclonal antibodies in the recombinant host cells. The DNA also can be modified, for example, by substituting the coding sequence for human heavy and light chain constant domains in place of the homologous murine sequences (U.S. Patent No. 4,816,567; Morrison, Nature 368, 812-13 (1994)) or by covalently joining to the immunoglobulin coding sequence all or part of the coding sequence for a non-immunoglobulin polypeptide. Such a non-immunoglobulin polypeptide can be substituted for the constant domains of an antibody of the invention, or can be substituted for the variable domains of one antigen-combining site of an antibody of the invention to create a chimeric bivalent antibody.

Humanized Antibodies

The antibodies directed against the protein antigens of the invention can further comprise humanized antibodies or human antibodies. These antibodies are suitable for administration to humans without engendering an immune response by the human against the administered immunoglobulin. Humanized forms of antibodies are chimeric immunoglobulins, immunoglobulin chains or fragments thereof (such as Fv, Fab, Fab', F(ab')₂ or other antigen-binding subsequences of antibodies) that are principally comprised of the sequence of a human immunoglobulin, and contain minimal sequence derived from a non-human immunoglobulin. Humanization can be performed following the method of Winter and co-workers (Jones et al., Nature, 321 :522-525 (1986); Riechmann et al., Nature, 332:323-327 (1988); Verhoeyen et al., Science, 239:1534-1536 (1988)), by substituting rodent CDRs or CDR sequences for the conesponding sequences of a human antibody. (See also U.S. Patent No. 5,225,539.) In some instances, Fv framework residues of the human immunoglobulin are replaced by conesponding non-human residues. Humanized antibodies can also comprise residues which are found neither in the recipient antibody nor in the imported CDR or framework sequences. In general, the humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in which all or substantially all of the CDR regions conespond to those of a non-human immunoglobulin and all or substantially all of the framework regions are those of a human immunoglobulin consensus sequence. The humanized antibody optimally also will comprise at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin (Jones et al., 1986; Riechmann et al., 1988; and Presta, Curr. On. Struct. Biol., 2:593-596 (1992)).

Human Antibodies Fully human antibodies essentially relate to antibody molecules in which the entire sequence of both the light chain and the heavy chain, including the CDRs, arise from human genes. Such antibodies are termed "human antibodies", or "fully human antibodies" herein. Human monoclonal antibodies can be prepared by the trioma technique; the human B-cell hybridoma technique (see Kozbor, et al., 1983 Immunol Today 4: 72) and the EBV hybridoma technique to produce human monoclonal antibodies (see Cole, et al., 1985 In: MONOCLONAL ANTIBODIES AND CANCER THERAPY, Alan R. Liss, Inc., pp. 77-96). Human monoclonal antibodies may be utilized in the practice of the present invention and may be produced by using human hybridomas (see Cote, et al., 1983. Proc Natl Acad Sci USA 80: 2026-2030) or by transforming human B-cells with Epstein Ban Virus in vitro (see Cole, et al., 1985 In: MONOCLONAL ANTIBODIES AND CANCER THERAPY, Alan R. Liss, Inc., pp. 77-96).

In addition, human antibodies can also be produced using additional techniques, including phage display libraries (Hoogenboom and Winter, J. Mol. Biol.. 227:381 (1991); Marks et al., J. Mol. Biol., 222:581 (1991)). Similarly, human antibodies can be made by introducing human immunoglobulin loci into transgenic animals, e.g., mice in which the endogenous immunoglobulin genes have been partially or completely inactivated. Upon challenge, human antibody production is observed, which closely resembles that seen in humans in all respects, including gene reanangement, assembly, and antibody repertoire. This approach is described, for example, in U.S. Patent Nos. 5,545,807; 5,545,806; 5,569,825; 5,625,126; 5,633,425; 5,661,016, and in Marks et al. (Bio/Technology 10, 779-783 (1992)); Lonberg et al. (Nature 368 856-859 (1994)); Morrison ( Nature 368, 812-13 (1994)); Fishwild et al.( Nature Biotechnology 14, 845-51 (1996)); Neuberger (Nature Biotechnology 14; 826 (1996)); and Lonberg and Huszar (Intern. Rev. Immunol. 13 65-93 (1995)).

Human antibodies may additionally be produced using transgenic nonhuman animals which are modified so as to produce fully human antibodies rather than the animal's endogenous antibodies in response to challenge by an antigen. (See PCT publication WO94/02602). The endogenous genes encoding the heavy and light immunoglobulin chains in the nonhuman host have been incapacitated, and active loci encoding human heavy and light chain immunoglobulins are inserted into the host's genome. The human genes are incorporated, for example, using yeast artificial chromosomes containing the requisite human DNA segments. An animal which provides all the desired modifications is then obtained as progeny by crossbreeding intermediate transgenic animals containing fewer than the full complement of the modifications. The prefened embodiment of such a nonhuman animal is a mouse, and is termed the Xenomouse™ as disclosed in PCT publications WO 96/33735 and WO 96/34096. This animal produces B cells which secrete fully human immunoglobulins. The antibodies can be obtained directly from the animal after immunization with an immunogen of interest, as, for example, a preparation of a polyclonal antibody, or alternatively from immortalized B cells derived from the animal, such as hybridomas producing monoclonal antibodies. Additionally, the genes encoding the immunoglobulins with human variable regions can be recovered and expressed to obtain the antibodies directly, or can be further modified to obtain analogs of antibodies such as, for example, single chain Fv molecules.

An example of a method of producing a nonhuman host, exemplified as a mouse, lacking expression of an endogenous immunoglobulin heavy chain is disclosed in U.S. Patent No. 5,939,598. It can be obtained by a method including deleting the J segment genes from at least one endogenous heavy chain locus in an embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector containing a gene encoding a selectable marker; and producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain the gene encoding the selectable marker.

A method for producing ah antibody of interest, such as a human antibody, is disclosed in U.S. Patent No. 5,916,771. It includes introducing an expression vector that contains a nucleotide sequence encoding a heavy chain into one mammalian host cell in culture, introducing an expression vector containing a nucleotide sequence encoding a light chain into another mammalian host cell, and fusing the two cells to form a hybrid cell. The hybrid cell expresses an antibody containing the heavy chain and the light chain.

In a further improvement on this procedure, a method for identifying a clinically relevant epitope on an immunogen, and a conelative method for selecting an antibody that binds immunospecifically to the relevant epitope with high affinity, are disclosed in PCT publication WO 99/53049.

Fab Fragments and Single Chain Antibodies According to the invention, techniques can be adapted for the production of single-chain antibodies specific to an antigenic protein of the invention (see e.g., U.S. Patent No. 4,946,778). In addition, methods can be adapted for the construction of F_ab expression libraries (see e.g., Huse, et al., 1989 Science 246: 1275-1281) to allow rapid and effective identification of monoclonal F_ab fragments with the desired specificity for a protein or derivatives, fragments, analogs or homologs thereof. Antibody fragments that contain the idiotypes to a protein antigen may be produced by techniques known in the art including, but not limited to: (i) an F(_ab-)₂ fragment produced by pepsin digestion of an antibody molecule; (ii) an F_ab fragment generated by reducing the disulfide bridges of an F(_ab_')₂ fragment; (iii) an F_ab fragment generated by the treatment of the antibody molecule with papain and a reducing agent and (iv) F_v fragments.

Bispecific Antibodies

Bispecifϊc antibodies are monoclonal, preferably human or humanized, antibodies that have binding specificities for at least two different antigens. In the present case, one of the binding specificities is for an antigenic protein of the invention. The second binding target is any other antigen, and advantageously is a cell-surface protein or receptor or receptor subunit. Methods for making bispecific antibodies are known in the art. Traditionally, the recombinant production of bispecific antibodies is based on the co-expression of two immunoglobulin heavy-chain/light-chain pairs, where the two heavy chains have different specificities (Milstein and Cuello, Nature, 305:537-539 (1983)). Because of the random assortment of immunoglobulin heavy and light chains, these hybridomas (quadromas) produce a potential mixture often different antibody molecules, of which only one has the conect bispecific structure. The purification of the conect molecule is usually accomplished by affinity chromatography steps. Similar procedures are disclosed in WO 93/08829, published 13 May 1993, and in Traunecker et al., EMBO J., 10:3655-3659 (1991).

Antibody variable domains with the desired binding specificities (antibody-antigen combining sites) can be fused to immunoglobulin constant domain sequences. The fusion preferably is with an immunoglobulin heavy-chain constant domain, comprising at least part of the hinge, CH2, and CH3 regions. It is prefened to have the first heavy-chain constant region (CHI) containing the site necessary for light-chain binding present in at least one of the fusions. DNAs encoding the immunoglobulin heavy-chain fusions and, if desired, the immunoglobulin light chain, are inserted into separate expression vectors, and are co-transfected into a suitable host organism. For further details of generating bispecific antibodies see, for example, Suresh et al.. Methods in Enzymology. 121 :210 (1986).

According to another approach described in WO 96/27011, the interface between a pair of antibody molecules can be engineered to maximize the percentage of heterodimers which are recovered from recombinant cell culture. The prefened interface comprises at least a part of the CH3 region of an antibody constant domain. In this method, one or more small amino acid side chains from the interface of the first antibody molecule are replaced with larger side chains (e.g. tyrosine or tryptophan). Compensatory "cavities" of identical or similar size to the large side chain(s) are created on the interface of the second antibody molecule by replacing large amino acid side chains with smaller ones (e.g. alanine or threonine). This provides a mechanism for increasing the yield of the heterodimer over other unwanted end-products such as homodimers.

Bispecific antibodies can be prepared as full length antibodies or antibody fragments (e.g. F(ab')₂ bispecific antibodies). Techniques for generating bispecific antibodies from antibody fragments have been described in the literature. For example, bispecific antibodies can be prepared using chemical linkage. Brennan et al., Science 229:81 (1985) describe a procedure wherein intact antibodies are proteolytically cleaved to generate F(ab') fragments. These fragments are reduced in the presence of the dithiol complexing agent sodium arsenite to stabilize vicinal dithiols and prevent intermolecular disulfide formation. The Fab' fragments generated are then converted to thionitrobenzoate (TNB) derivatives. One of the Fab'-TNB derivatives is then reconverted to the Fab'-thiol by reduction with mercaptoethylamine and is mixed with an equimolar amount of the other Fab'-TNB derivative to form the bispecific antibody. The bispecific antibodies produced can be used as agents for the selective immobilization of enzymes. Additionally, Fab' fragments can be directly recovered from E. coli and chemically coupled to form bispecific antibodies. Shalaby et al., J. EXP. Med. 175:217-225 (1992) describe the production of a fully humanized bispecific antibody F(ab') molecule. Each Fab' fragment was separately secreted from E. coli and subjected to directed chemical coupling in vitro to form the bispecific antibody. The bispecific antibody thus formed was able to bind to cells overexpressing the ErbB2 receptor and normal human T cells, as well as trigger the lytic activity of human cytotoxic lymphocytes against human breast tumor targets.

Various techniques for making and isolating bispecific antibody fragments directly from recombinant cell culture have also been described. For example, bispecific antibodies have been produced using leucine zippers. Kostelny et al., J. Immunol.

148(5): 1547- 1553 (1992). The leucine zipper peptides from the Fos and Jun proteins were linked to the Fab' portions of two different antibodies by gene fusion. The antibody homodimers were reduced at the hinge region to form monomers and then re-oxidized to form the antibody heterodimers. This method can also be utilized for the production of antibody homodimers. The "diabody" technology described by Hollinger et al., Proc. Natl. Acad. Sci. USA 90:6444-6448 (1993) has provided an alternative mechanism for making bispecific antibody fragments. The fragments comprise a heavy-chain variable domain (VH) connected to a light-chain variable domain (VL) by a linker which is too short to allow pairing between the two domains on the same chain. Accordingly, the VH and VL domains of one fragment are forced to pair with the complementary V and VH domains of another fragment, thereby forming two antigen-binding sites. Another strategy for making bispecific antibody fragments by the use of single-chain Fv (sFv) dimers has also been reported. See, Gruber et al., J. Immunol. 152:5368 (1994). Antibodies with more than two valencies are contemplated. For example, trispecific antibodies can be prepared. Tutt et al., J. Immunol. 147:60 (1991).

Exemplary bispecific antibodies can bind to two different epitopes, at least one of which originates in the protein antigen of the invention. Alternatively, an anti-antigenic arm of an immunoglobulin molecule can be combined with an arm which binds to a triggering molecule on a leukocyte such as a T-cell receptor molecule (e.g. CD2, CD3, CD28, or B7), or Fc receptors for IgG (FcγR), such as FcγRI (CD64), FcγRII (CD32) and FcγRIII (CD 16) so as to focus cellular defense mechanisms to the cell expressing the particular antigen. Bispecific antibodies can also be used to direct cytotoxic agents to cells which express a particular antigen. These antibodies possess an antigen-binding arm and an arm which binds a cytotoxic agent or a radionuclide chelator, such as EOTUBE, DPTA, DOT A, or TETA. Another bispecific antibody of interest binds the protein antigen described herein and further binds tissue factor (TF).

Heteroconjugate Antibodies

Heteroconjugate antibodies are also within the scope of the present invention. Heteroconjugate antibodies are composed of two covalently joined antibodies. Such antibodies have, for example, been proposed to target immune system cells to unwanted cells (U.S. Patent No. 4,676,980), and for treatment of HIV infection (WO 91/00360; WO 92/200373; EP 03089). It is contemplated that the antibodies can be prepared in vitro using known methods in synthetic protein chemistry, including those involving crosslinking agents. For example, immunotoxins can be constructed using a disulfide exchange reaction or by forming a thioether bond. Examples of suitable reagents for this purpose include iminothiolate and methyl-4-mercaptobutyrimidate and those disclosed, for example, in U.S. Patent No. 4,676,980.

Effector Function Engineering

It can be desirable to modify the antibody of the invention with respect to effector function, so as to enhance, e.g., the effectiveness of the antibody in treating cancer. For example, cysteine residuefs) can be introduced into the Fc region, thereby allowing interchain disulfide bond formation in this region. The homodimeric antibody thus generated can have improved internalization capability and/or increased complement-mediated cell killing and antibody-dependent cellular cytotoxicity (ADCC). See Caron et al., J. Exp Med., 176: 1191-1195 (1992) and Shopes, J. Immunol., 148: 2918-2922 (1992). Homodimeric antibodies with enhanced anti-tumor activity can also be prepared using heterobifunctional cross-linkers as described in Wolff et al. Cancer Research, 53: 2560-2565 (1993). Alternatively, an antibody can be engineered that has dual Fc regions and can thereby have enhanced complement lysis and ADCC capabilities. See Stevenson et al., Anti-Cancer Drug Design. 3: 219-230 (1989).

Immunoconjugates

The invention also pertains to immunoconjugates comprising an antibody conjugated to a cytotoxic agent such as a chemotherapeutic agent, toxin (e.g., an enzymatically active toxin of bacterial, fungal, plant, or animal origin, or fragments thereof), or a radioactive isotope (i.e., a radioconjugate).

Chemotherapeutic agents useful in the generation of such immunoconjugates have been described above. Enzymatically active toxins and fragments thereof that can be used include diphtheria A chain, nonbinding active fragments of diphtheria toxin, exotoxin A chain (from Pseudomonas aeruginosa), ricin A chain, abrin A chain, modeccin A chain, alpha-sarcin, Aleurites fordii proteins, dianthin proteins, Phytolaca americana proteins (PAPI, PAPII, and PAP-S), momordica charantia inhibitor, curcin, crotin, sapaonaria officinalis inhibitor, gelonin, mitogellin, restrictocin, phenomycin, enomycin, and the tricothecenes. A variety of radionuclides are available for the production of radiocoηjugated antibodies. Examples include ²¹²Bi, ¹³¹1, ^13IIn, ⁹⁰Y, and ¹⁸⁶Re.

Conjugates of the antibody and cytotoxic agent are made using a variety of bifunctional protein-coupling agents such as N-succinimidyl-3-(2-pyridyldithiol) propionate (SPDP), iminothiolane (IT), bifunctional derivatives of imidoesters (such as dimethyl adipimidate HCL), active esters (such as disuccinimidyl suberate), aldehydes (such as glutareldehyde), bis-azido compounds (such as bis (p-azidobenzoyl) hexanediamine), bis-diazonium derivatives (such as bis-(p-diazoniumbenzoyl)-ethylenediamine), diisocyanates (such as tolyene 2,6-diisocyanate), and bis-active fluorine compounds (such as l,5-difluoro-2,4-dinitrobenzene). For example, a ricin immunotoxin can be prepared as described in Vitetta et al., Science, 238: 1098 (1987). Carbon- 14-labeled l-isothiocyanatobenzyl-3-methyldiethylene triaminepentaacetic acid (MX-DTPA) is an exemplary chelating agent for conjugation of radionucleotide to the antibody. See WO94/11026.

In another embodiment, the antibody can be conjugated to a "receptor" (such streptavidin) for utilization in tumor pretargeting wherein the antibody-receptor conjugate is administered to the patient, followed by removal of unbound conjugate from the circulation using a clearing agent and then administration of a "ligand" (e.g., avidin) that is in turn conjugated to a cytotoxic agent.

Immunoliposomes The antibodies disclosed herein can also be formulated as immunoliposomes.

Liposomes containing the antibody are prepared by methods known in the art, such as described in Epstein et al., Proc. Natl. Acad. Sci. USA, 82: 3688 (1985); Hwang et al., Proc. Natl Acad. Sci. USA, IT.4030 (1980); and U.S. Pat. Nos. 4,485,045 and 4,544,545. Liposomes with enhanced circulation time are disclosed in U.S. Patent No. 5,013,556. Particularly useful liposomes can be generated by the reverse-phase evaporation method with a lipid composition comprising phosphatidylcholine, cholesterol, and PEG-derivatized phosphatidylethanolamine (PEG-PE). Liposomes are extruded through filters of defined pore size to yield liposomes with the desired diameter. Fab' fragments of the antibody of the present invention can be conjugated to the liposomes as described in Martin et al ., J. Biol. Chem., 257: 286-288 (1982) via a disulfide-interchange reaction. A chemotherapeutic agent (such as Doxorubicin) is optionally contained within the liposome. See Gabizon et al., J. National Cancer Inst., 81(19): 1484 (1989).

Diagnostic Applications of Antibodies Directed Against the Proteins of the Invention Antibodies directed against a protein of the invention may be used in methods known within the art relating to the localization and/or quantitation of the protein (e.g., for use in measuring levels of the protein within appropriate physiological samples, for use in diagnostic methods, for use in imaging the protein, and the like). In a given embodiment, antibodies against the proteins, or derivatives, fragments, analogs or homologs thereof, that contain the antigen binding domain, are utilized as pharmacologically-active compounds (see below).

An antibody specific for a protein of the invention can be used to isolate the protein by standard techniques, such as irnmunoaffϊnity chromatography or immunoprecipitation. Such an antibody can facilitate the purification of the natural protein antigen from cells and of recombinantly produced antigen expressed in host cells. Moreover, such an antibody can be used to detect the antigenic protein (e.g., in a cellular lysate or cell supernatant) in order to evaluate the abundance and pattern of expression of the antigenic protein. Antibodies directed against the protein can be used diagnostically to monitor protein levels in tissue as part of a clinical testing procedure, e.g., to, for example, determine the efficacy of a given treatment regimen. Detection can be facilitated by coupling (i.e., physically linking) the antibody to a detectable substance. Examples of detectable substances include various enzymes, prosthetic groups, fluorescent materials, luminescent materials, bioluminescent materials, and radioactive materials. Examples of suitable enzymes include horseradish peroxidase, alkaline phosphatase, β-galactosidase, or acetylcholinesterase; examples of suitable prosthetic group complexes include streptavidin/biotin and avidin/biotin; examples of suitable fluorescent materials include umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin; an example of a luminescent material includes luminol; examples of bioluminescent materials include luciferase, luciferin, and aequorin, and examples of suitable radioactive material include I, I, S or H.

Antibody Therapeutics Antibodies of the invention, including polyclonal, monoclonal, humanized and fully human antibodies, may used as therapeutic agents. Such agents will generally be employed to treat or prevent a disease or pathology in a subject. An antibody preparation, preferably one having high specificity and high affinity for its target antigen, is administered to the subject and will generally have an effect due to its binding with the target. Such an effect may be one of two kinds, depending on the specific nature of the interaction between the given antibody molecule and the target antigen in question. In the first instance, administration of the antibody may abrogate or inhibit the binding of the target with an endogenous ligand to which it naturally binds. In this case, the antibody binds to the target and masks a binding site of the naturally occurring ligand, wherein the ligand serves as an effector molecule. Thus the receptor mediates a signal transduction pathway for which ligand is responsible.

Alternatively, the effect may be one in which the antibody elicits a physiological result by virtue of binding to an effector binding site on the target molecule. In this case the target, a receptor having an endogenous ligand which may be absent or defective in the disease or pathology, binds the antibody as a surrogate effector ligand, initiating a receptor-based signal transduction event by the receptor.

A therapeutically effective amount of an antibody of the invention relates generally to the amount needed to achieve a therapeutic objective. As noted above, this may be a binding interaction between the antibody and its target antigen that, in certain cases, interferes with the functioning of the target, and in other cases, promotes a physiological response. The amount required to be administered will furthermore depend on the binding affinity of the antibody for its specific antigen, and will also depend on the rate at which an administered antibody is depleted from the free volume other subject to which it is administered. Common ranges for therapeutically effective dosing of an antibody or antibody fragment of the invention may be, by way of nonlimiting example, from about 0.1 mg/kg body weight to about 50 mg/kg body weight. Common dosing frequencies may range, for example, from twice daily to once a week.

Pharmaceutical Compositions of Antibodies

Antibodies specifically binding a protein of the invention, as well as other molecules identified by the screening assays disclosed herein, can be administered for the treatment of various disorders in the form of pharmaceutical compositions. Principles and considerations involved in preparing such compositions, as well as guidance in the choice of components are provided, for example, in Remington : The Science And Practice Of Pharmacy 19th ed. (Alfonso R. Gennaro, et al., editors) Mack Pub. Co., Easton, Pa. : 1995; Drug Absorption Enhancement : Concepts, Possibilities, Limitations, And Trends, Harwood Academic Publishers, Langhorne, Pa., 1994; and Peptide And Protein Drug Delivery (Advances In Parenteral Sciences, Vol. 4), 1991, M. Dekker, New York.

If the antigenic protein is intracellular and whole antibodies are used as inhibitors, internalizing antibodies are prefened. However, liposomes can also be used to deliver the antibody, or an antibody fragment, into cells. Where antibody fragments are used, the smallest inhibitory fragment that specifically binds to the binding domain of the target protein is prefened. For example, based upon the variable-region sequences of an antibody, peptide molecules can be designed that retain the ability to bind the target protein sequence. Such peptides can be synthesized chemically and/or produced by recombinant DNA technology. See, e.g., Marasco et al., Proc. Natl. Acad. Sci. USA, 90: 7889-7893 (1993). The formulation herein can also contain more than one active compound as necessary for the particular indication being treated, preferably those with complementary activities that do not adversely affect each other. Alternatively, or in addition, the composition can comprise an agent that enhances its function, such as, for example, a cytotoxic agent, cytokine, chemotherapeutic agent, or growth-inhibitory agent. Such molecules are suitably present in combination in amounts that are effective for the purpose intended.

The active ingredients can also be entrapped in microcapsules prepared, for example, by coacervation techniques or by interfacial polymerization, for example, hydroxymethylcellulose or gelatin-microcapsules and poly-(methylmethacrylate) microcapsules, respectively, in colloidal drug delivery systems (for example, liposomes, albumin microspheres, microemulsions, nano-particles, and nanocapsules) or in macroemulsions.

The formulations to be used for in vivo administration must be sterile. This is readily accomplished by filtration through sterile filtration membranes.

Sustained-release preparations can be prepared. Suitable examples of sustained-release preparations include semipermeable matrices of solid hydrophobic polymers containing the antibody, which matrices are in the form of shaped articles, e.g., films, or microcapsules. Examples of sustained-release matrices include polyesters, hydrogels (for example, poly(2-hydroxyethyl-methacrylate), or poly(vinylalcohol)), polylactides (U.S. Pat. No. 3,773,919), copolymers of L-glutamic acid and γ ethyl-L-glutamate, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers such as the LUPRON DEPOT ™ (injectable microspheres composed of lactic acid-glycolic acid copolymer and leuprolide acetate), and poly-D-(-)-3-hydroxybutyric acid. While polymers such as ethylene-vinyl acetate and lactic acid-glycolic acid enable release of molecules for over 100 days, certain hydrogels release proteins for shorter time periods.

ELISA Assay An agent for detecting an analyte protein is an antibody capable of binding to an analyte protein, preferably an antibody with a detectable label. Antibodies can be polyclonal, or more preferably, monoclonal. An intact antibody, or a fragment thereof (e.g., Fab or F(ab)2 can be used. The term "labeled", with regard to the probe or antibody, is intended to encompass direct labeling of the probe or antibody by coupling (i.e., physically linking) a detectable substance to the probe or antibody, as well as indirect labeling of the probe or antibody by reactivity with another reagent that is directly labeled. Examples of indirect labeling include detection of a primary antibody using a fluorescently-labeled secondary antibody and end-labeling of a DNA probe with biotin such that it can be detected with fluorescently-labeled streptavidin. The term "biological sample" is intended to include tissues, cells and biological fluids isolated from a subject, as well as tissues, cells and fluids present within a subject. Included within the usage of the term "biological sample", therefore, is blood and a fraction or component of blood including blood serum, blood plasma, or lymph. That is, the detection method of the invention can be used to detect an analyte mRNA, protein, or genomic DNA in a biological sample in vitro as well as in vivo. For example, in vitro techniques for detection of an analyte mRNA include Northern hybridizations and in situ hybridizations. In vitro techniques for detection of an analyte protein include enzyme linked immunosorbent assays (ELISAs), Western blots, immunoprecipitations, and immunofluorescence. In vitro techniques for detection of an analyte genomic DNA include Southern hybridizations. Procedures for conducting immunoassays are described, for example in "ELISA: Theory and Practice: Methods in Molecular Biology", Vol. 42, J. R. Crowther (Ed.) Human Press, Totowa, NJ, 1995; "Immunoassay", E. Diamandis and T. Christopoulus, Academic Press, Inc., San Diego, CA, 1996; and "Practice and Thory of Enzyme Immunoassays", P.

Tijssen, Elsevier Science Publishers, Amsterdam, 1985. Furthermore, in vivo techniques for detection of an analyte protein include introducing into a subject a labeled anti-an analyte protein antibody. For example, the antibody can be labeled with a radioactive marker whose presence and location in a subject can be detected by standard imaging techniques.

NOVX Recombinant Expression Vectors and Host Cells

Another aspect of the invention pertains to vectors, preferably expression vectors, containing a nucleic acid encoding A NOVX protein, or derivatives, fragments, analogs or homologs thereof. As used herein, the term "vector" refers to a nucleic acid molecule capable of transporting another nucleic acid to which it has been linked. One type of vector is a "plasmid", which refers to a circular double stranded DNA loop into which additional DNA segments can be ligated. Another type of vector is a viral vector, wherein additional DNA segments can be ligated into the viral genome. Certain vectors are capable of autonomous replication in a host cell into which they are introduced (e.g., bacterial vectors having a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) are integrated into the genome of a host cell upon introduction into the host cell, and thereby are replicated along with the host genome. Moreover, certain vectors are capable of directing the expression of genes to which they are operatively-linked. Such vectors are refened to herein as "expression vectors". In general, expression vectors of utility in recombinant DNA techniques are often in the form of plasmids. In the present specification, "plasmid" and "vector" can be used interchangeably as the plasmid is the most commonly used form of vector. However, the invention is intended to include such other forms of expression vectors, such as viral vectors (e.g. , replication defective retroviruses, adenoviruses and adeno-associated viruses), which serve equivalent functions.

The recombinant expression vectors of the invention comprise a nucleic acid of the invention in a form suitable for expression of the nucleic acid in a host cell, which means that the recombinant expression vectors include one or more regulatory sequences, selected on the basis of the host cells to be used for expression, that is operatively-linked to the nucleic acid sequence to be expressed. Within a recombinant expression vector, "operably-linked" is intended to mean that the nucleotide sequence of interest is linked to the regulatory sequence(s) in a manner that allows for expression of the nucleotide sequence (e.g., in an in vitro transcription/translation system or in a host cell when the vector is introduced into the host cell).

- The term "regulatory sequence" is intended to includes promoters, enhancers and other expression control elements (e.g., polyadenylation signals). Such regulatory sequences are described, for example, in Goeddel, GENE EXPRESSION TECHNOLOGY: METHODS IN ENZYMOLOGY 185, Academic Press, San Diego, Calif. (1990). Regulatory sequences include those that direct constitutive expression of a nucleotide sequence in many types of host cell and those that direct expression of the nucleotide sequence only in certain host cells (e.g., tissue-specific regulatory sequences). It will be appreciated by those skilled in the art that the design of the expression vector can depend on such factors as the choice of the host cell to be transformed, the level of expression of protein desired, etc. The expression vectors of the invention can be introduced into host cells to thereby produce proteins or peptides, including fusion proteins or peptides, encoded by nucleic acids as described herein (e.g., NOVX proteins, mutant forms of NOVX proteins, fusion proteins, etc.).

The recombinant expression vectors of the invention can be designed for expression of NOVX proteins in prokaryotic or eukaryotic cells. For example, NOVX proteins can be expressed in bacterial cells such as Escherichia coli, insect cells (using baculovirus expression vectors) yeast cells or mammalian cells. Suitable host cells are discussed further in Goeddel, GENE EXPRESSION TECHNOLOGY: METHODS IN ENZYMOLOGY 185, Academic Press, San Diego, Calif. (1990). Alternatively, the recombinant expression vector can be transcribed and translated in vitro, for example using T7 promoter regulatory sequences and T7 polymerase.

Expression of proteins in prokaryotes is most often carried out in Escherichia coli with vectors containing constitutive or inducible promoters directing the expression of either fusion or non-fusion proteins. Fusion vectors add a number of amino acids to a protein encoded therein, usually to the amino terminus of the recombinant protein. Such fusion vectors typically serve three purposes: (i) to increase expression of recombinant protein; (ii) to increase the solubility of the recombinant protein; and (iii) to aid in the purification of the recombinant protein by acting as a ligand in affinity purification. Often, in fusion expression vectors, a proteolytic cleavage site is introduced at the junction of the fusion moiety and the recombinant protein to enable separation of the recombinant protein from the fusion moiety subsequent to purification of the fusion protein. Such enzymes, and their cognate recognition sequences, include Factor Xa, thrombin and enterokinase. Typical fusion expression vectors include pGEX (Pharmacia Biotech Inc; Smith and Johnson, 1988. Gene 61: 31-40), pMAL (New England Biolabs, Beverly, Mass.) and pRIT5 (Pharmacia, Piscataway, NJ.) that fuse glutathione S-transferase (GST), maltose E binding protein, or protein A, respectively, to the target recombinant protein.

Examples of suitable inducible non-fusion E. coli expression vectors include pTrc (Amrann et al, (1988) Gene 69:301-315) and pΕT 1 Id (Siudier et al, GΕNΕ EXPRESSION TECHNOLOGY: METHODS IN ENZYMOLOGY 185, Academic Press, San Diego, Calif. (1990) 60-89).

One strategy to maximize recombinant protein expression in E. coli is to express the protein in a host bacteria with an impaired capacity to proteolytically cleave the recombinant protein. See, e.g., Gottesman, GENE EXPRESSION TECHNOLOGY: METHODS IN ENZYMOLOGY 185, Academic Press, San Diego, Calif. (1990) 119-128. Another strategy is to alter the nucleic acid sequence of the nucleic acid to be inserted into an expression vector so that the individual codons for each amino acid are those preferentially utilized in E. coli (see, e.g., Wada, et al., 1992. Nucl. Acids Res. 20: 2111-2118). Such alteration of nucleic acid sequences of the invention can be carried out by standard DNA synthesis techniques.

In another embodiment, the NOVX expression vector is a yeast expression vector. Examples of vectors for expression in yeast Saccharomyces cerivisae include pYepSecl (Baldari, et al, 1987. EMBO J. 6: 229-234), pMFa (Kurjan and Herskowitz, 1982. Cell 30: 933-943), pJRY88 (Schultz et al, 1987. Gene 54: 113-123), pYES2 (Invitrogen Corporation, San Diego, Calif.), and picZ (InVitrogen Corp, San Diego, Calif).

Alternatively, NOVX can be expressed in insect cells using baculovirus expression vectors. Baculovirus vectors available for expression of proteins in cultured insect cells (e.g., SF9 cells) include the pAc series (Smith, et al, 1983. Mol Cell. Biol. 3: 2156-2165) and the pVL series (Lucklow and Summers, 1989. Virology 170: 31-39).

In yet another embodiment, a nucleic acid of the invention is expressed in mammalian cells using a mammalian expression vector. Examples of mammalian expression vectors include pCDM8 (Seed, 1987. Nαtwre 329: 840) and pMT2PC (Kaufman, et al, 1987. EMBOJ. 6: 187-195). When used in mammalian cells, the expression vector's control functions are often provided by viral regulatory elements. For example, commonly used promoters are derived from polyoma, adenovirus 2, cytomegalovirus, and simian virus 40. For other suitable expression systems for both prokaryotic and eukaryotic cells see, e.g., Chapters 16 and 17 of Sambrook, et al, MOLECULAR CLONING: A LABORATORY MANUAL. 2nd ed., Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1989.

In another embodiment, the recombinant mammalian expression vector is capable of directing expression of the nucleic acid preferentially in a particular cell type (e.g., tissue-specific regulatory elements are used to express the nucleic acid). Tissue-specific regulatory elements are known in the art. Non-limiting examples of suitable tissue-specific promoters include the albumin promoter (liver-specific; Pinkert, et al., 1987. Genes Dev. 1: 268-277), lymphoid-specific promoters (Calame and Eaton, 1988. Adv. Immunol. 43: 235-275), in particular promoters of T cell receptors (Winoto and Baltimore, 1989. EMBOJ. 8: 729-733) and immunoglobulins (Banerji, et al, 1983. Cell 33: 729-740; Queen and Baltimore, 1983. Cell 33: 741-748), neuron-specific promoters (e.g., the neurofilament promoter; Byrne and Ruddle, 1989. Proc. Natl. Acad. Sci. USA 86: 5473-5477), pancreas-specific promoters (Edhind, et al., 1985. Science 230: 912-916), and mammary gland-specific promoters (e.g., milk whey promoter; U.S. Pat. No. 4,873,316 and European Application Publication No.264, 166). Developmentally-regulated promoters are also encompassed, e.g., the murine hox promoters (Kessel and Grass, 1990. Science 249: 374-379) and the α-fetoprotein promoter (Campes and Tilghman, 1989. Genes Dev. 3: 537-546).

The invention further provides a recombinant expression vector comprising a DNA molecule of the invention cloned into the expression vector in an antisense orientation. That is, the DNA molecule is operatively-linked to a regulatory sequence in a manner that allows for expression (by transcription of the DNA molecule) of an RNA molecule that is antisense to NOVX mRNA. Regulatory sequences operatively linked to a nucleic acid cloned in the antisense orientation can be chosen that direct the continuous expression of the antisense RNA molecule in a variety of cell types, for instance viral promoters and or enhancers, or regulatory sequences can be chosen that direct constitutive, tissue specific or cell type specific expression of antisense RNA. The antisense expression vector can be in the form of a recombinant plasmid, phagemid or attenuated virus in which antisense nucleic acids are produced under the control of a high efficiency regulatory region, the activity of which can be determined by the cell type into which the vector is introduced. For a discussion of the regulation of gene expression using antisense genes see, e.g., Weintraub, et al, "Antisense RNA as a molecular tool for genetic analysis," Reviews-Trends in Genetics, Vol. 1(1) 1986.

Another aspect of the invention pertains to host cells into which a recombinant expression vector of the invention has been introduced. The terms "host cell" and

"recombinant host cell" are used interchangeably herein. It is understood that such terms refer not only to the particular subject cell but also to the progeny or potential progeny of such a cell. Because certain modifications may occur in succeeding generations due to either mutation or environmental influences, such progeny may not, in fact, be identical to the parent cell, but are still included within the scope of the term as used herein.

A host cell can be any prokaryotic or eukaryotic cell. For example, NOVX protein can be expressed in bacterial cells such as E. coli, insect cells, yeast or mammalian cells (such as Chinese hamster ovary cells (CHO) or COS cells). Other suitable host cells are known to those skilled in the art.

Vector DNA can be introduced into prokaryotic or eukaryotic cells via conventional transformation or transfection techniques. As used herein, the terms "transformation" and "transfection" are intended to refer to a variety of art-recognized techniques for introducing foreign nucleic acid (e.g., DNA) into a host cell, including calcium phosphate or calcium chloride co-precipitation, DEAE-dextran-mediated transfection, lipofection, or electroporation. Suitable methods for transforming or transfecting host cells can be found in Sambrook, et al. (MOLECULAR CLONING: A LABORATORY MANUAL. 2nd ed., Cold Spring Harbor Laboratory, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1989), and other laboratory manuals.

For stable transfection of mammalian cells, it is known that, depending upon the expression vector and transfection technique used, only a small fraction of cells may integrate the foreign DNA into their genome. In order to identify and select these integrants, a gene that encodes a selectable marker (e.g., resistance to antibiotics) is generally introduced into the host cells along with the gene of interest. Various selectable markers include those that confer resistance to drugs, such as G418, hygromycin and methotrexate. Nucleic acid encoding a selectable marker can be introduced into a host cell on the same vector as that encoding NOVX or can be introduced on a separate vector. Cells stably transfected with the introduced nucleic acid can be identified by drug selection (e.g., cells that have incorporated the selectable marker gene will survive, while the other cells die).

A host cell of the invention, such as a prokaryotic or eukaryotic host cell in culture, can be used to produce (i.e., express) NOVX protein. Accordingly, the invention further provides methods for producing NOVX protein using the host cells of the invention. In one embodiment, the method comprises culturing the host cell of invention (into which a recombinant expression vector encoding NOVX protein has been introduced) in a suitable medium such that NOVX protein is produced. In another embodiment, the method further comprises isolating NOVX protein from the medium or the host cell.

Transgenic NOVX Animals

The host cells of the invention can also be used to produce non-human transgenic animals. For example, in one embodiment, a host cell of the invention is a fertilized oocyte or an embryonic stem cell into which NOVX protein-coding sequences have been introduced. Such host cells can then be used to create non-human transgenic animals in which exogenous NOVX sequences have been introduced into their genome or homologous recombinant animals in which endogenous NOVX sequences have been altered. Such animals are useful for studying the function and/or activity of NOVX protein and for identifying and/or evaluating modulators of NOVX protein activity. As used herein, a "transgenic animal" is a non-human animal, preferably a mammal, more preferably a rodent such as a rat or mouse, in which one or more of the cells of the animal includes a transgene. Other examples of transgenic animals include non-human primates, sheep, dogs, cows, goats, chickens, amphibians, etc. A transgene is exogenous DNA that is integrated into the genome of a cell from which a transgenic animal develops and that remains in the genome of the mature animal, thereby directing the expression of an encoded gene product in one or more cell types or tissues of the transgenic animal. As used herein, a "homologous recombinant animal" is a non-human animal, preferably a mammal, more preferably a mouse, in which an endogenous NOVX gene has been altered by homologous recombination between the endogenous gene and an exogenous DNA molecule introduced into a cell of the animal, e.g., an embryonic cell of the animal, prior to development of the animal.

A transgenic animal of the invention can be created by introducing NOVX-encoding nucleic acid into the male pronuclei of a fertilized oocyte (e.g. , by microinjection, retroviral infection) and allowing the oocyte to develop in a pseudopregnant female foster animal. The human NOVX cDNA sequences SEQ ID NOS:2n-l, wherem n is an integer between 1 and 62, can be introduced as a transgene into the genome of a non-human animal. Alternatively, a non-human homologue of the human NOVX gene, such as a mouse NOVX gene, can be isolated based on hybridization to the human NOVX cDNA (described further supra) and used as a transgene. Intronic sequences and polyadenylation signals can also be included in the transgene to increase the efficiency of expression of the transgene. A tissue-specific regulatory sequence(s) can be operably-linked to the NOVX transgene to direct expression of NOVX protein to particular cells. Methods for generating transgenic animals via embryo manipulation and microinjection, particularly animals such as mice, have become conventional in the art and are described, for example, in U.S. Patent Nos. 4,736,866; 4,870,009; and 4,873,191; and Hogan, 1986. In: MANIPULATING THE MOUSE EMBRYO, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. Similar methods are used for production of other transgenic animals. A transgenic founder animal can be identified based upon the presence of the NOVX transgene in its genome and/or expression of NOVX mRNA in tissues or cells of the animals. A transgenic founder animal can then be used to breed additional animals carrying the transgene. Moreover, transgenic animals carrying a transgene-encoding NOVX protein can further be bred to other transgenic animals carrying other transgenes.

To create a homologous recombinant animal, a vector is prepared which contains at least a portion of A NOVX gene into which a deletion, addition or substitution has been introduced to thereby alter, e.g., functionally disrupt, the NOVX gene. The NOVX gene can be a human gene (e.g. , the cDNA of SEQ ID NOS:2n-l , wherein n is an integer between 1 and 62), but more preferably, is a non-human homologue of a human NOVX gene. For example, a mouse homologue of human NOVX gene of SEQ ID NOS:2n-l , wherein n is an integer between 1 and 62, can be used to construct a homologous recombination vector suitable for altering an endogenous NOVX gene in the mouse genome. In one embodiment, the vector is designed such that, upon homologous recombination, the endogenous NOVX gene is functionally disrupted (i.e., no longer encodes a functional protein; also refened to as a "knock out" vector).

Alternatively, the vector can be designed such that, upon homologous recombination, the endogenous NOVX gene is mutated or otherwise altered but still encodes functional protein (e.g., the upstream regulatory region can be altered to thereby alter the expression of the endogenous NOVX protein). In the homologous recombination vector, the altered portion of the NOVX gene is flanked at its 5'- and 3 '-termini by additional nucleic acid of the NOVX gene to allow for homologous recombination to occur between the exogenous NOVX gene carried by the vector and an endogenous

NOVX gene in an embryonic stem cell. The additional flanking NOVX nucleic acid is of sufficient length for successful homologous recombination with the endogenous gene. Typically, several kilobases of flanking DNA (both at the 5'- and 3 '-termini) are included in the vector. See, e.g., Thomas, et al., 1987. Cell 51: 503 for a description of homologous recombination vectors. The vector is ten introduced into an embryonic stem cell line (e.g., by electroporation) and cells in which the introduced NOVX gene has homologously-recombined with the endogenous NOVX gene are selected. See, e.g., Li, et al, 1992. Cell 69: 915. The selected cells are then injected into a blastocyst of an animal (e.g., a mouse) to form aggregation chimeras. See, e.g., Bradley, 1987. In: TERATOCARCINOMAS AND EMBRYONIC STEM CELLS: A PRACTICAL APPROACH, Robertson, ed. IRL, Oxford, pp. 113-152. A chimeric embryo can then be implanted into a suitable pseudopregnant female foster animal and the embryo brought to term. Progeny harboring the homologously-recombined DNA in their germ cells can be used to breed animals in which all cells of the animal contain the homologously-recombined DNA by germline transmission of the transgene. Methods for constructing homologous recombination vectors and homologous recombinant animals are described further in Bradley, 1991. Curr. Opin. Biotechnol. 2: 823-829; PCT International Publication Nos. : WO 90/11354; WO 91/01140; WO 92/0968; and WO 93/04169.

In another embodiment, transgenic non-humans animals can be produced that contain selected systems that allow for regulated expression of the transgene. One example of such a system is the cre/loxP recombinase system of bacteriophage PL For a description of the cre/loxP recombinase system, See, e.g., Lakso, et al, 1992. Proc. Natl. Acad. Sci. USA 89: 6232-6236. Another example of a recombinase system is the FLP recombinase system of Saccharomyces cerevisiae. See, O'Gorman, et al, 1991. Science 251:1351-1355. If a cre/loxP recombinase system is used to regulate expression of the transgene, animals containing transgenes encoding both the Cre recombinase and a selected protein are required. Such animals can be provided through the construction of "double" transgenic animals, e.g., by mating two transgenic animals, one containing a transgene encoding a selected protein and the other containing a transgene encoding a recombinase.

Clones of the non-human transgenic animals described herein can also be produced according to the methods described in Wilmut, et al, 1997. Nature 385: 810-813. In brief, a cell (e.g., a somatic cell) from the transgenic animal can be isolated and induced to exit the growth cycle and enter Go phase. The quiescent cell can then be fused, e.g., through the use of electrical pulses, to an enucleated oocyte from an animal of the same species from which the quiescent cell is isolated. The reconstructed oocyte is then cultured such that it develops to morula or blastocyte and then transfened to pseudopregnant female foster animal. The offspring bome of this female foster animal will be a clone of the animal from which the cell (e.g., the somatic cell) is isolated. Pharmaceutical Compositions

The NOVX nucleic acid molecules, NOVX proteins, and anti-NOVX antibodies (also refened to herein as "active compounds") of the invention, and derivatives, fragments, analogs and homologs thereof, can be incorporated into pharmaceutical compositions suitable for administration. Such compositions typically comprise the nucleic acid molecule, protein, or antibody and a pharmaceutically acceptable carrier. As used herein, "pharmaceutically acceptable carrier" is intended to include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration. Suitable carriers are described in the most recent edition of Remington's Pharmaceutical Sciences, a standard reference text in the field, which is incorporated herein by reference. Prefened examples of such carriers or diluents include, but are not limited to, water, saline, finger's solutions, dextrose solution, and 5% human serum albumin. Liposomes and non-aqueous vehicles such as fixed oils may also be used. The use of such media and agents for pharmaceutically active substances is well known in the art. Except insofar as any conventional media or agent is incompatible with the active compound, use thereof in the compositions is contemplated. Supplementary active compounds can also be incorporated into the compositions. A pharmaceutical composition of the invention is formulated to be compatible with its intended route of administration. Examples of routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, oral (e.g., inhalation), transdermal (t^'.e., topical), transmucosal, and rectal administration. Solutions or suspensions used for parenteral, intradermal, or subcutaneous application can include the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid (EDTA); buffers such as acetates, citrates or phosphates, and agents for the adjustment of tonicity such as sodium chloride or dextrose. The pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide. The parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic. Pharmaceutical compositions suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor EL (BASF, Parsippany, N.J.) or phosphate buffered saline (PBS). In all cases, the composition must be sterile and should be fluid to the extent that easy syringeability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. The proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants. Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars, polyalcohols such as manitol, sorbitol, sodium chloride in the composition. Prolonged absoφtion of the injectable compositions can be brought about by including in the composition an agent which delays absoφtion, for example, aluminum monostearate and gelatin. Sterile injectable solutions can be prepared by incoφorating the active compound

(e.g. , A NOVX protein or anti-NOVX antibody) in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incoφorating the active compound into a sterile vehicle that contains a basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, methods of preparation are vacuum drying and freeze-drying that yields a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof.

Oral compositions generally include an inert diluent or an edible carrier. They can be enclosed in gelatin capsules or compressed into tablets. For the purpose of oral therapeutic administration, the active compound can be incoφorated with excipients and used in the form of tablets, troches, or capsules. Oral compositions can also be prepared using a fluid carrier for use as a mouthwash, wherein the compound in the fluid carrier is applied orally and swished and expectorated or swallowed. Pharmaceutically compatible binding agents, and/or adjuvant materials can be included as part of the composition. The tablets, pills, capsules, troches and the like can contain any of the following ingredients, or compounds of a similar nature: a binder such as microcrystalline cellulose, gum tragacanth or gelatin; an excipient such as starch or lactose, a disintegrating agent such as alginic acid, Primogel, or corn starch; a lubricant such as magnesium stearate or Sterotes; a glidant such as colloidal silicon dioxide; a sweetening agent such as sucrose or saccharin; or a flavoring agent such as peppermint, methyl salicylate, or orange flavoring.

For administration by inhalation, the compounds are delivered in the form of an aerosol spray from pressured container or dispenser which contains a suitable propellant, e.g., a gas such as carbon dioxide, or a nebulizer.

Systemic administration can also be by transmucosal or transdermal means. For transmucosal or transdermal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, detergents, bile salts, and fusidic acid derivatives. Transmucosal administration can be accomplished through the use of nasal sprays or suppositories. For transdermal administration, the active compounds are formulated into ointments, salves, gels, or creams as generally known in the art. The compounds can also be prepared in the form of suppositories (e.g. , with conventional suppository bases such as cocoa butter and other glycerides) or retention enemas for rectal delivery.

In one embodiment, the active compounds are prepared with carriers that will protect the compound against rapid elimination from the body, such as a controlled release formulation, including implants and microencapsulated delivery systems. Biodegradable, biocompatible polymers can.be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid. Methods for preparation of such formulations will be apparent to those skilled in the art. The materials can also be obtained commercially from Alza Coφoration and Nova Pharmaceuticals, Inc. Liposomal suspensions (including liposomes targeted to infected cells with monoclonal antibodies to viral antigens) can also be used as pharmaceutically acceptable carriers. These can be prepared according to methods known to those skilled in the art, for example, as described in U.S. Patent No. 4,522,811. It is especially advantageous to formulate oral or parenteral compositions in dosage unit form for ease of administration and uniformity of dosage. Dosage unit form as used herein refers to physically discrete units suited as unitary dosages for the subject to be treated; each unit containing a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier. The specification for the dosage unit forms of the invention are dictated by and directly dependent on the unique characteristics of the active compound and the particular therapeutic effect to be achieved, and the limitations inherent in the art of compounding such an active compound for the treatment of individuals. The nucleic acid molecules of the invention can be inserted into vectors and used as gene therapy vectors. Gene therapy vectors can be delivered to a subject by, for example, intravenous injection, local administration (see, e.g., U.S. Patent No. 5,328,470) or by stereotactic injection (see, e.g., Chen, et al., 1994. Proc. Natl. Acad. Sci. USA 91: 3054-3057). The pharmaceutical preparation of the gene therapy vector can include the gene therapy vector in an acceptable diluent, or can comprise a slow release matrix in which the gene delivery vehicle is imbedded. Alternatively, where the complete gene delivery vector can be produced intact from recombinant cells, e.g., retroviral vectors, the pharmaceutical preparation can include one or more cells that produce the gene delivery system. The pharmaceutical compositions can be included in a container, pack, or dispenser together with instructions for administration.

Screening and Detection Methods

The isolated nucleic acid molecules of the invention can be used to express NONX protein (e.g. , via a recombinant expression vector in a host cell in gene therapy applications), to detect ΝONX mRΝA (e.g., in a biological sample) or a genetic lesion in A ΝOVX gene, and to modulate ΝOVX activity, as described further, below. In addition, the ΝOVX proteins can be used to screen drugs or compounds that modulate the ΝOVX protein activity or expression as well as to treat disorders characterized by insufficient or excessive production of ΝOVX protein or production of ΝONX protein forms that have decreased or abenant activity compared to ΝONX wild-type protein (e.g.; diabetes (regulates insulin release); obesity (binds and transport lipids); metabolic disturbances associated with obesity, the metabolic syndrome X as well as anorexia and wasting disorders associated with chronic diseases and various cancers, and infectious disease(possesses anti-microbial activity) and the various dyslipidemias. In addition, the anti-NOVX antibodies of the invention can be used to detect and isolate NOVX proteins and modulate NOVX activity. In yet a further aspect, the invention can be used in methods to influence appetite, absoφtion of nutrients and the disposition of metabolic substrates in both a positive and negative fashion.

The invention further pertains to novel agents identified by the screening assays described herein and uses thereof for treatments as described, supra.

Screening Assays

The invention provides a method (also refened to herein as a "screening assay") for identifying modulators, i.e., candidate or test compounds or agents (e.g., peptides, peptidomimetics, small molecules or other drugs) that bind to NOVX proteins or have a stimulatory or inhibitory effect on, e.g., NOVX protein expression or NOVX protein activity. The invention also includes compounds identified in the screening assays described herein.

In one embodiment, the invention provides assays for screening candidate or test compounds which bind to or modulate the activity of the membrane-bound form of A NOVX protein or polypeptide or biologically-active portion thereof. The test compounds of the invention can be obtained using any of the numerous approaches in combinatorial library methods known in the art, including: biological libraries; spatially addressable parallel solid phase or solution phase libraries; synthetic library methods requiring deconvolution; the "one-bead one-compound" library method; and synthetic library methods using affinity chromatography selection. The biological library approach is limited to peptide libraries, while the other four approaches are applicable to peptide, non-peptide oligomer or small molecule libraries of compounds. See, e.g., Lam, 1997. Anticancer Drug Design 12: 145.

A "small molecule" as used herein, is meant to refer to a composition that has a molecular weight of less than about 5 kD and most preferably less than about 4 kD. Small molecules can be, e.g., nucleic acids, peptides, polypeptides, peptidomimetics, carbohydrates, lipids or other organic or inorganic molecules. Libraries of chemical and/or biological mixtures, such as fungal, bacterial, or algal extracts, are known in the art and can be screened with any of the assays of the invention. Examples of methods for the synthesis of molecular libraries can be found in the art, for example in: DeWitt, et al, 1993. Proc. Natl Acad. Sci. U.S.A. 90: 6909; Erb, et al, 1994. Proc. Natl. Acad. Sci. U.S.A. 91: 11422; Zuckermarm, et al, 1994. J. Med. Chem. 37: 2678; Cho, et al, 1993. Science 261: 1303; Canell, et al, 1994. Angew. Chem. Int. Ed. Engl 33: 2059; Carell, et al., 1994. Angew. Chem. Int. Ed. Engl. 33: 2061; and Gallop, et al., 1994. J. Med. Chem. 37: 1233.

Libraries of compounds may be presented in solution (e.g., Houghten, 1992. Biotechniques 13: 412-421), or on beads (Lam, 1991. Nature 354: 82-84), on chips (Fodor, 1993. Nature 364: 555-556), bacteria (Ladner, U.S. Patent No. 5,223,409), spores (Ladner, U.S. Patent 5,233,409), plasmids (Cull, et al, 1992. Proc. Natl. Acad. Sci. USA 89: 1865-1869) or on phage (Scott and Smith, 1990. Science 249: 386-390; Devlin, 1990. Science 249: 404-406; Cwirla, et al, 1990. Proc. Natl. Acad. Sci. U.S.A. 87: 6378-6382; Felici, 1991. J. Mol. Biol. 222: 301-310; Ladner, U.S. Patent No. 5,233,409.).

In one embodiment, an assay is a cell-based assay in which a cell which expresses a membrane-bound form of NOVX protein, or a biologically-active portion thereof, on the cell surface is contacted with a test compound and the ability of the test compound to bind to A NOVX protein determined. The cell, for example, can of mammalian origin or a yeast cell. Determining the ability of the test compound to bind to the NOVX protein can be accomplished, for example, by coupling the test compound with a radioisotope or enzymatic label such that binding of the test compound to the NOVX protein or biologically-active portion thereof can be determined by detecting the labeled compound in a complex. For example, test compounds can be labeled with ^{, 5}1, ³⁵S, ¹⁴C, or ³H, either directly or indirectly, and the radioisotope detected by direct counting of radioemission or by scintillation counting. Alternatively, test compounds can be enzymatically-labeled with, for example, horseradish peroxidase, alkaline phosphatase, or luciferase, and the enzymatic label detected by determination of conversion of an appropriate substrate to product. In one embodiment, the assay comprises contacting a cell which expresses a membrane-bound form of NOVX protein, or a biologically-active portion thereof, on the cell surface with a known compound which binds NOVX to form an assay mixture, contacting the assay mixture with a test compound, and determining the ability of the test compound to interact with A NOVX protein, wherein determining the ability of the test compound to interact with A NOVX protein comprises determining the ability of the test compound to preferentially bind to NOVX protein or a biologically-active portion thereof as compared to the known compound.

In another embodiment, an assay is a cell-based assay comprising contacting a cell expressing a membrane-bound form of NOVX protein, or a biologically-active portion thereof, on the cell surface with a test compound and determining the ability of the test compound to modulate (e.g., stimulate or inhibit) the activity of the NOVX protein or biologically-active portion thereof. Determining the ability of the test compound to modulate the activity of NOVX or a biologically-active portion thereof can be accomplished, for example, by determining the ability of the NOVX protein to bind to or interact with A NOVX target molecule. As used herein, a "target molecule" is a molecule with which A NOVX protein binds or interacts in nature, for example, a molecule on the surface of a cell which expresses A NOVX interacting protein, a molecule on the surface of a second cell, a molecule in the extracellular milieu, a molecule associated with the internal surface of a cell membrane or a cytoplasmic molecule. A NOVX target molecule can be a non-NOVX molecule or A NOVX protein or polypeptide of the invention. In one embodiment, A NOVX target molecule is a component of a signal transduction pathway that facilitates transduction of an extracellular signal (e.g. a signal generated by binding of a compound to a membrane-bound NOVX molecule) through the cell membrane and into the cell. The target, for example, can be a second intercellular protein that has catalytic activity or a protein that facilitates the association of downstream signaling molecules with NOVX.

Determining the ability of the NOVX protein to bind to or interact with A NOVX target molecule can be accomplished by one of the methods described above for determining direct binding. In one embodiment, determining the ability of the NOVX protein to bind to or interact with A NOVX target molecule can be accomplished by determining the activity of the target molecule. For example, the activity of the target molecule can be determined by detecting induction of a cellular second messenger of the

9+ target (i.e. intracellular Ca , diacylglycerol, IP₃, etc.), detecting catalytic/enzymatic activity of the target an appropriate substrate, detecting the induction of a reporter gene (comprising A NOVX-responsive regulatory element operatively linked to a nucleic acid encoding a detectable marker, e.g., luciferase), or detecting a cellular response, for example, cell survival, cellular differentiation, or cell proliferation. In yet another embodiment, an assay of the invention is a cell-free assay comprising contacting A NOVX protein or biologically-active portion thereof with a test compound and determining the ability of the test compound to bind to the NOVX protein or biologically-active portion thereof. Binding of the test compound to the NOVX protein can be determined either directly or indirectly as described above. In one such embodiment, the assay comprises contacting the NOVX protein or biologically-active portion thereof with a known compound which binds NOVX to form an assay mixture, contacting the assay mixture with a test compound, and determining the ability of the test compound to interact with A NOVX protein, wherein determining the ability of the test compound to interact with A NOVX protein comprises determining the ability of the test compound to preferentially bind to NOVX or biologically-active portion thereof as compared to the known compound.

In still another embodiment, an assay is a cell-free assay comprising contacting NOVX protein or biologically-active portion thereof with a test compound and determining the ability of the test compound to modulate (e.g. stimulate or inhibit) the activity of the NOVX protein or biologically-active portion thereof. Determining the ability of the test compound to modulate the activity of NOVX can be accomplished, for example, by determining the ability of the NOVX protein to bind to A NOVX target molecule by one of the methods described above for determining direct binding. In an alternative embodiment, determining the ability of the test compound to modulate the activity of NOVX protein can be accomplished by determining the ability of the NOVX protein further modulate A NOVX target molecule. For example, the catalytic/enzymatic activity of the target molecule on an appropriate substrate can be determined as described, supra. In yet another embodiment, the cell-free assay comprises contacting the NOVX protein or biologically-active portion thereof with a known compound which binds NOVX protein to form an assay mixture, contacting the assay mixture with a test compound, and determining the ability of the test compound to interact with A NOVX protein, wherein determining the ability of the test compound to interact with A NOVX protein comprises determining the ability of the NOVX protein to preferentially bind to or modulate the activity of A NOVX target molecule.

The cell-free assays of the invention are amenable to use of both the soluble form or the membrane-bound form of NOVX protein. In the case of cell-free assays comprising the membrane-bound form of NOVX protein, it may be desirable to utilize a solubilizing agent such that the membrane-bound form of NOVX protein is maintained in solution. Examples of such solubilizing agents include non-ionic detergents such as n-octylglucoside, n-dodecylglucoside, n-dodecylmaltoside, octanoyl-N-methylglucamide, decanoyl-N-methylglucamide, Triton^® X-l 00, Triton^® X-l 14, Thesit^®,

Isotridecypoly(ethylene glycol ether)_π, N-dodecyl— N,N-dimethyl-3-ammomo-l -propane sulfonate, 3-(3-cholamidopropyl) dimethylamminiol-1 -propane sulfonate (CHAPS), or 3-(3-cholamidopropyl)dimethylamminiol-2-hydroxy-l -propane sulfonate (CHAPSO).

In more than one embodiment of the above assay methods of the invention, it may be desirable to immobilize either NOVX protein or its target molecule to facilitate separation of complexed from uncomplexed forms of one or both of the proteins, as well as to accommodate automation of the assay. Binding of a test compound to NOVX protein, or interaction of NOVX protein with a target molecule in the presence and absence of a candidate compound, can be accomplished in any vessel suitable for containing the reactants. Examples of such vessels include microtiter plates, test tubes, and micro-centrifuge tubes. In one embodiment, a fusion protein can be provided that adds a domain that allows one or both of the proteins to be bound to a matrix. For example, GST-NOVX fusion proteins or GST-target fusion proteins can be adsorbed onto glutathione sepharose beads (Sigma Chemical, St. Louis, MO) or glutathione derivatized microtiter plates, that are then combined with the test compound or the test compound and either the non-adsorbed target protein or NOVX protein, and the mixture is incubated under conditions conducive to complex formation (e.g., at physiological conditions for salt and pH). Following incubation, the beads or microtiter plate wells are washed to remove any unbound components, the matrix immobilized in the case of beads, complex determined either directly or indirectly, for example, as described, supra. Alternatively, the complexes can be dissociated from the matrix, and the level of NOVX protein binding or activity determined using standard techniques.

Other techniques for immobilizing proteins on matrices can also be used in the screening assays of the invention. For example, either the NOVX protein or its target molecule can be immobilized utilizing conjugation of biotin and streptavidin. Biotinylated NOVX protein or target molecules can be prepared from biotin-NHS (N-hydroxy-succinimide) using techniques well-known within the art (e.g., biotinylation kit, Pierce Chemicals, Rockford, 111.), and immobilized in the wells of streptavidin-coated 96 well plates (Pierce Chemical). Alternatively, antibodies reactive with NOVX protein or target molecules, but which do not interfere with binding of the NOVX protein to its target molecule, can be derivatized to the wells of the plate, and unbound target or NOVX protein trapped in the wells by antibody conjugation. Methods for detecting such complexes, in addition to those described above for the GST-immobilized complexes, include immunodetection of complexes using antibodies reactive with the NOVX protein or target molecule, as well as enzyme-linked assays that rely on detecting an enzymatic activity associated with the NOVX protein or target molecule.

In another embodiment, modulators of NOVX protein expression are identified in a method wherein a cell is contacted with a candidate compound and the expression of NOVX mRNA or protein in the cell is determined. The level of expression of NOVX mRNA or protein in the presence of the candidate compound is compared to the level of expression of NOVX mRNA or protein in the absence of the candidate compound. The candidate compound can then be identified as a modulator of NOVX mRNA or protein expression based upon this comparison. For example, when expression of NOVX mRNA or protein is greater (i.e., statistically significantly greater) in the presence of the candidate compound than in its absence, the candidate compound is identified as a stimulator of NOVX mRNA or protein expression. Alternatively, when expression of NOVX mRNA or protein is less (statistically significantly less) in the presence of the candidate compound than in its absence, the candidate compound is identified as an inhibitor of NOVX mRNA or protein expression. The level of NOVX mRNA or protein expression in the cells can be determined by methods described herein for detecting NOVX mRNA or protein.

In yet another aspect of the invention, the NOVX proteins can be used as "bait proteins" in a two-hybrid assay or three hybrid assay (see, e.g., U.S. Patent No. 5,283,317; Zervos, et al., 1993. Cell 72: 223-232; Madura, et al, 1993. J. Biol. Chem. 268:

12046-12054; Bartel, et al., 1993. Biotechniques 14: 920-924; Iwabuchi, et al., 1993. Oncogene 8: 1693-1696; and Brent WO 94/10300), to identify other proteins that bind to or interact with NOVX ("NOVX-binding proteins" or "NOVX-bp") and modulate NOVX activity. Such NOVX-binding proteins are also likely to be involved in the propagation of signals by the NOVX proteins as, for example, upstream or downstream elements of the NOVX pathway.

The two-hybrid system is based on the modular nature of most transcription factors, which consist of separable DNA-binding and activation domains. Briefly, the assay utilizes two different DNA constructs. In one construct, the gene that codes for NOVX is fused to a gene encoding the DNA binding domain of a known transcription factor (e.g., GAL-4). In the other construct, a DNA sequence, from a library of DNA sequences, that encodes an unidentified protein ("prey" or "sample") is fused to a gene that codes for the activation domain of the known transcription factor. If the "bait" and the "prey" proteins are able to interact, in vivo, forming A NOVX-dependent complex, the DNA-binding and activation domains of the transcription factor are brought into close proximity. This proximity allows transcription of a reporter gene (e.g., LacZ) that is operably linked to a transcriptional regulatory site responsive to the transcription factor. Expression of the reporter gene can be detected and cell colonies containing the functional transcription factor can be isolated and used to obtain the cloned gene that encodes the protein which interacts with NOVX.

The invention further pertains to novel agents identified by the aforementioned screening assays and uses thereof for treatments as described herein.

Detection Assays

Portions or fragments of the cDNA sequences identified herein (and the conesponding complete gene sequences) can be used in numerous ways as polynucleotide reagents. By way of example, and not of limitation, these sequences can be used to: (/^') map their respective genes on a chromosome; and, thus, locate gene regions associated with genetic disease; (ii) identify an individual from a minute biological sample (tissue typing); and (iii) aid in forensic identification of a biological sample. Some of these applications are described in the subsections, below.

Chromosome Mapping

Once the sequence (or a portion of the sequence) of a gene has been isolated, this sequence can be used to map the location of the gene on a chromosome. This process is called chromosome mapping. Accordingly, portions or fragments of the NOVX sequences, SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, or fragments or derivatives thereof, can be used to map the location of the NOVX genes, respectively, on a chromosome. The mapping of the NOVX sequences to chromosomes is an important first step in conelating these sequences with genes associated with disease. Briefly, NOVX genes can be mapped to chromosomes by preparing PCR primers (preferably 15-25 bp in length) from the NOVX sequences. Computer analysis of the NOVX, sequences can be used to rapidly select primers that do not span more than one exon in the genomic DNA, thus complicating the amplification process. These primers can then be used for PCR screening of somatic cell hybrids containing individual human chromosomes. Only those hybrids containing the human gene conesponding to the NOVX sequences will yield an amplified fragment.

Somatic cell hybrids are prepared by fusing somatic cells from different mammals (e.g., human and mouse cells). As hybrids of human and mouse cells grow and divide, they gradually lose human chromosomes in random order, but retain the mouse chromosomes. By using media in which mouse cells cannot grow, because they lack a particular enzyme, but in which human cells can, the one human chromosome that contains the gene encoding the needed enzyme will be retained. By using various media, panels of hybrid cell lines can be established. Each cell line in a panel contains either a single human chromosome or a small number of human chromosomes, and a full set of mouse chromosomes, allowing easy mapping of individual genes to specific human chromosomes. See, e.g., D'Eustachio, et al., 1983. Science 220: 919-924. Somatic cell hybrids containing only fragments of human chromosomes can also be produced by using human chromosomes with translocations and deletions. PCR mapping of somatic cell hybrids is a rapid procedure for assigning a particular sequence to a particular chromosome. Three or more sequences can be assigned per day using a single thermal cycler. Using the NOVX sequences to design oligonucleotide primers, sub-localization can be achieved with panels of fragments from specific chromosomes. Fluorescence in situ hybridization (FISH) of a DNA sequence to a metaphase chromosomal spread can further be used to provide a precise chromosomal location in one step. Chromosome spreads can be made using cells whose division has been blocked in metaphase by a chemical like colcemid that disrupts the mitotic spindle. The chromosomes can be treated briefly with trypsin, and then stained with Gie sa. A pattern of light and dark bands develops on each chromosome, so that the chromosomes can be identified individually. The FISH technique can be used with a DNA sequence as short as 500 or 600 bases. However, clones larger than 1,000 bases have a higher likelihood of binding to a unique chromosomal location with sufficient signal intensity for simple detection. Preferably 1,000 bases, and more preferably 2,000 bases, will suffice to get good results at a reasonable amount of time. For a review of this technique, see, Verma, et al, HUMAN CHROMOSOMES: A MANUAL OF BASIC TECHNIQUES (Pergamon Press, New York 1988). Reagents for chromosome mapping can be used individually to mark a single chromosome or a single site on that chromosome, or panels of reagents can be used for marking multiple sites and/or multiple chromosomes. Reagents conesponding to noncoding regions of the genes actually are prefened for mapping purposes. Coding sequences are more likely to be conserved within gene families, thus increasing the chance of cross hybridizations during chromosomal mapping.

Once a sequence has been mapped to a precise chromosomal location, the physical position of the sequence on the chromosome can be conelated with genetic map data. Such data are found, e.g., in McKusick, MENDELIAN INHERITANCE IN MAN, available on-line through Johns Hopkins University Welch Medical Library). The relationship between genes and disease, mapped to the same chromosomal region, can then be identified through linkage analysis (co-inheritance of physically adjacent genes), described in, e.g., Egeland, et al, 1987. Nature, 325: 783-787.

Moreover, differences in the DNA sequences between individuals affected and unaffected with a disease associated with the NOVX gene, can be determined. If a mutation is observed in some or all of the affected individuals but not in any unaffected individuals, then the mutation is likely to be the causative agent of the particular disease. Comparison of affected and unaffected individuals generally involves first looking for structural alterations in the chromosomes, such as deletions or translocations that are visible from chromosome spreads or detectable using PCR based on that DNA sequence. Ultimately, complete sequencing of genes from several individuals can be performed to confirm the presence of a mutation and to distinguish mutations from polymoφhisms.

Tissue Typing

The NOVX sequences of the invention can also be used to identify individuals from minute biological samples. In this technique, an individual's genomic DNA is digested with one or more restriction enzymes, and probed on a Southern blot to yield unique bands for identification. The sequences of the invention are useful as additional DNA markers for RFLP ("restriction fragment length polymoφhisms," described in U.S. Patent No. 5,272,057).

Furthermore, the sequences of the invention can be used to provide an alternative technique that determines the actual base-by-base DNA sequence of selected portions of an individual's genome. Thus, the NOVX sequences described herein can be used to prepare two PCR primers from the 5'- and 3 '-termini of the sequences. These primers can then be used to amplify an individual's DNA and subsequently sequence it.

Panels of conesponding DNA sequences from individuals, prepared in this manner, can provide unique individual identifications, as each individual will have a unique set of such DNA sequences due to allelic differences. The sequences of the invention can be used to obtain such identification sequences from individuals and from tissue. The NOVX sequences of the invention uniquely represent portions of the human genome. Allelic variation occurs to some degree in the coding regions of these sequences, and to a greater degree in the noncoding regions. It is estimated that allelic variation between individual humans occurs with a frequency of about once per each 500 bases. Much of the allelic variation is due to single nucleotide polymoφhisms (SNPs), which include restriction fragment length polymoφhisms (RFLPs).

Each of the sequences described herein can, to some degree, be used as a standard against which DNA from an individual can be compared for identification pmposes. Because greater numbers of polymoφhisms occur in the noncoding regions, fewer sequences are necessary to differentiate individuals. The noncoding sequences can comfortably provide positive individual identification with a panel of perhaps 10 to 1,000 primers that each yield a noncoding amplified sequence of 100 bases. If predicted coding sequences, such as those in SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, are used, a more appropriate number of primers for positive individual identification would be 500-2,000.

Predictive Medicine

The invention also pertains to the field of predictive medicine in which diagnostic assays, prognostic assays, pharmacogenomics, and monitoring clinical trials are used for prognostic (predictive) puφoses to thereby treat an individual prophylactically. Accordingly, one aspect of the invention relates to diagnostic assays for determining NOVX protein and/or nucleic acid expression as well as NOVX activity, in the context of a biological sample (e.g., blood, serum, cells, tissue) to thereby determine whether an individual is afflicted with a disease or disorder, or is at risk of developing a disorder, associated with abenant NOVX expression or activity. The disorders include metabolic disorders, diabetes, obesity, infectious disease, anorexia, cancer-associated cachexia, cancer, neurodegenerative disorders, Alzheimer's Disease, Parkinson's Disorder, immune disorders, and hematopoietic disorders, and the various dyslipidemias, metabolic disturbances associated with obesity, the metabolic syndrome X and wasting disorders associated with chronic diseases and various cancers. The invention also provides for prognostic (or predictive) assays for determining whether an individual is at risk of developing a disorder associated with NOVX protein, nucleic acid expression or activity. For example, mutations in A NOVX gene can be assayed in a biological sample. Such assays can be used for prognostic or predictive pwpose to thereby prophylactically treat an individual prior to the onset of a disorder characterized by or associated with NOVX protein, nucleic acid expression, or biological activity. Another aspect of the invention provides methods for determining NOVX protein, nucleic acid expression or activity in an individual to thereby select appropriate therapeutic or prophylactic agents for that individual (refened to herein as "pharmacogenomics"). Pharmacogenomics allows for the selection of agents (e.g., drugs) for therapeutic or prophylactic treatment of an individual based on the genotype of the individual (e.g., the genotype of the individual examined to determine the ability of the individual to respond to a particular agent.)

Yet another aspect of the invention pertains to monitoring the influence of agents (e.g., drugs, compounds) on the expression or activity υf NOVX in clinical trials.

These and other agents are described in further detail in the following sections.

DIAGNOSTIC ASSAYS

An exemplary method for detecting the presence or absence of NOVX in a biological sample involves obtaining a biological sample from a test subject and contacting the biological sample with a compound or an agent capable of detecting NOVX protein or nucleic acid (e.g. , mRNA, genomic DNA) that encodes NOVX protein such that the presence of NOVX is detected in the biological sample. An agent for detecting NOVX mRNA or genomic DNA is a labeled nucleic acid probe capable of hybridizing to NOVX mRNA or genomic DNA. The nucleic acid probe can be, for example, a full-length NOVX nucleic acid, such as the nucleic acid of SEQ ID NOS:2n-l, wherein n is an integer between 1 and 62, or a portion thereof, such as an oligonucleotide of at least 15, 30, 50, 100, 250 or 500 nucleotides in length and sufficient to specifically hybridize under stringent conditions to NOVX mRNA or genomic DNA. Other suitable probes for use in the diagnostic assays of the invention are described herein.

An agent for detecting NOVX protein is an antibody capable of binding to NOVX protein, preferably an antibody with a detectable label. Antibodies can be polyclonal, or more preferably, monoclonal. An intact antibody, or a fragment thereof (e.g., Fab or F(ab')₂) can be used. The term "labeled", with regard to the probe or antibody, is intended to encompass direct labeling of the probe or antibody by coupling (/. e. , physically linking) a detectable substance to the probe or antibody, as well as indirect labeling of the probe or antibody by reactivity with another reagent that is directly labeled. Examples of indirect labeling include detection of a primary antibody using a fluorescently-labeled secondary antibody and end-labeling of a DNA probe with biotin such that it can be detected with fluorescently-labeled streptavidin. The term "biological sample" is intended to include tissues, cells and biological fluids isolated from a subject, as well as tissues, cells and fluids present within a subject. That is, the detection method of the invention can be used to detect NOVX mRNA, protein, or genomic DNA in a biological sample in vitro as well as in vivo. For example, in vitro techniques for detection of NOVX mRNA include Northern hybridizations and in situ hybridizations. In vitro techniques for detection of NOVX protein include enzyme linked immunosorbent assays (ELISAs), Western blots, immunoprecipitations, and immunofluorescence. In vitro techniques for detection of NOVX genomic DNA include Southern hybridizations. Furthermore, in vivo techniques for detection of NOVX protein include introducing into a subject a labeled anti-NOVX antibody. For example, the antibody can be labeled with a radioactive marker whose presence and location in a subject can be detected by standard imaging techniques.

In one embodiment, the biological sample contains protein molecules from the test subject. Alternatively, the biological sample can contain mRNA molecules from the test subject or genomic DNA molecules from the test subject. A prefened biological sample is a peripheral blood leukocyte sample isolated by conventional means from a subject.

In another embodiment, the methods further involve obtaining a control biological sample from a control subject, contacting the control sample with a compound or agent capable of detecting NOVX protein, mRNA, or genomic DNA, such that the presence of NOVX protein, mRNA or genomic DNA is detected in the biological sample, and comparing the presence of NOVX protein, mRNA or genomic DNA in the control sample with the presence of NOVX protein, mRNA or genomic DNA in the test sample.

The invention also encompasses kits for detecting the presence of NOVX in a biological sample. For example, the kit can comprise: a labeled compound or agent capable of detecting NOVX protein or mRNA in a biological sample; means for determining the amount of NOVX in the sample; and means for comparing the amount of NOVX in the sample with a standard. The compound or agent can be packaged in a suitable container. The kit can further comprise instructions for using the kit to detect NOVX protein or nucleic acid.

PROGNOSTIC ASSAYS

The diagnostic methods described herein can furthermore be utilized to identify subjects having or at risk of developing a disease or disorder associated with abenant NOVX expression or activity. For example, the assays described herein, such as the preceding diagnostic assays or the following assays, can be utilized to identify a subject having or at risk of developing a disorder associated with NOVX protein, nucleic acid expression or activity. Alternatively, the prognostic assays can be utilized to identify a subject having or at risk for developing a disease or disorder. Thus, the invention provides a method for identifying a disease or disorder associated with abenant NOVX expression or activity in which a test sample is obtained from a subject and NOVX protein or nucleic acid (e.g., mRNA, genomic DNA) is detected, wherein the presence of NOVX protein or nucleic acid is diagnostic for a subject having or at risk of developing a disease or disorder associated with abenant NOVX expression or activity. As used herein, a "test sample" refers to a biological sample obtained from a subject of interest. For example, a test sample can be a biological fluid (e.g., serum), cell sample, or tissue.

Furthermore, the prognostic assays described herein can be used to determine whether a subject can be admimstered an agent (e.g., an agonist, antagonist, peptidomimetic, protein, peptide, nucleic acid, small molecule, or other drug candidate) to treat a disease or disorder associated with abenant NOVX expression or activity. For example, such methods can be used to determine whether a subject can be effectively treated with an agent for a disorder. Thus, the invention provides methods for determining whether a subject can be effectively treated with an agent for a disorder associated with abenant NOVX expression or activity in which a test sample is obtained and NOVX protein or nucleic acid is detected (e.g., wherein the presence of NOVX protein or nucleic acid is diagnostic for a subject that can be administered the agent to treat a disorder associated with abenant NOVX expression or activity). The methods of the invention can also be used to detect genetic lesions in A

NOVX gene, thereby determining if a subject with the lesioned gene is at risk for a disorder characterized by abenant cell proliferation and/or differentiation. In various embodiments, the methods include detecting, in a sample of cells from the subject, the presence or absence of a genetic lesion characterized by at least one of an alteration affecting the integrity of a gene encoding A NO VX-protein, or the misexpression of the NOVX gene. For example, such genetic lesions can be detected by ascertaining the existence of at least one of: (i) a deletion of one or more nucleotides from A NOVX gene; (ii) an addition of one or more nucleotides to A NOVX gene; (iii) a substitution of one or more nucleotides of A NOVX gene, (iv) a chromosomal rearrangement of A NOVX gene; (v) an alteration in the level of a messenger RNA transcript of A NOVX gene, (vi) abenant modification of A NOVX gene, such as of the methylation pattern of the genomic DNA, (vii) the presence of a non- wild-type splicing pattern of a messenger RNA transcript of A NOVX gene, (viii) a non- wild-type level of A NOVX protein, (ix) allelic loss of A NOVX gene, and (x) inappropriate post-translational modification of A NOVX protein. As described herein, there are a large number of assay techniques known in the art which can be used for detecting lesions in A NOVX gene. A prefened biological sample is a peripheral blood leukocyte sample isolated by conventional means from a subject. However, any biological sample containing nucleated cells may be used, including, for example, buccal mucosal cells. In certain embodiments, detection of the lesion involves the use of a probe/primer in a polymerase chain reaction (PCR) (see, e.g., U.S. Patent Nos. 4,683,195 and 4,683,202), such as anchor PCR or RACE PCR, or, alternatively, in a ligation chain reaction (LCR) (see, e.g., Landegran, et al, 1988. Science 241: 1077-1080; and Nakazawa, et al, 1994. Proc. Natl. Acad. Sci. USA 91: 360-364), the latter of which can be particularly useful for detecting point mutations in the NOVX-gene (see, Abravaya, et al, 1995. Nucl. Acids Res. 23: 675-682). This method can include the steps of collecting a sample of cells from a patient, isolating nucleic acid (e.g., genomic, mRNA or both) from the cells of the sample, contacting the nucleic acid sample with one or more primers that specifically hybridize to A NOVX gene under conditions such that hybridization and amplification of the NOVX gene (if present) occurs, and detecting the presence or absence of an amplification product, or detecting the size of the amplification product and comparing the length to a control sample. It is anticipated that PCR and/or LCR may be desirable to use as a preliminary amplification step in conjunction with any of the techniques used for detecting mutations described herein.

Alternative amplification methods include: self sustained sequence replication (see, Guatelli, et al, 1990. Proc. Natl. Acad. Sci. USA 87: 1874-1878), transcriptional amplification system (see, Kwoh, et al, 1989. Proc. Natl. Acad. Sci. USA 86: 1173-1177); Qβ Replicase (see, Lizardi, et al, 1988. BioTechnology 6: 1197), or any other nucleic acid amplification method, followed by the detection of the amplified molecules using techniques well known to those of skill in the art. These detection schemes are especially useful for the detection of nucleic acid molecules if such molecules are present in very low numbers. In an alternative embodiment, mutations in A NOVX gene from a sample cell can be identified by alterations in restriction enzyme cleavage patterns. For example, sample and control DNA is isolated, amplified (optionally), digested with one or more restriction endonucleases, and fragment length sizes are determined by gel electrophoresis and compared. Differences in fragment length sizes between sample and control DNA indicates mutations in the sample DNA. Moreover, the use of sequence specific ribozymes (see, e.g., U.S. Patent No. 5,493,531) can be used to score for the presence of specific mutations by development or loss of a ribozyme cleavage site.

In other embodiments, genetic mutations in NOVX can be identified by hybridizing a sample and control nucleic acids, e.g., DNA or RNA, to high-density anays containing hundreds or thousands of oligonucleotides probes. See, e.g., Cronin, et al, 1996. Human Mutation 1: 244-255; Kozal, et al, 1996. Nat. Med. 2: 753-759. For example, genetic mutations in NOVX can be identified in two dimensional anays containing light-generated DNA probes as described in Cronin, et al., supra. Briefly, a first hybridization anay of probes can be used to scan through long stretches of DNA in a sample and control to identify base changes between the sequences by making linear anays of sequential overlapping probes. This step allows the identification of point mutations. This is followed by a second hybridization anay that allows the characterization of specific mutations by using smaller, specialized probe anays complementary to all variants or mutations detected. Each mutation anay is composed of parallel probe sets, one complementary to the wild-type gene and the other complementary to the mutant gene.

In yet another embodiment, any of a variety of sequencing reactions known in the art can be used to directly sequence the NOVX gene and detect mutations by comparing the sequence of the sample NOVX with the conesponding wild-type (control) sequence. Examples of sequencing reactions include those based on techniques developed by Maxim and Gilbert, 1977. Proc. Natl Acad. Sci. USA 74: 560 or Sanger, 1977. Proc. Natl Acad. Sci. USA 74: 5463. It is also contemplated that any of a variety of automated sequencing procedures can be utilized when performing the diagnostic assays (see, e.g., Naeve, et al, 1995. Biotechniques 19: 448), including sequencing by mass spectrometry (see, e.g., PCT International Publication No. WO 94/16101; Cohen, et al, 1996. Adv. Chromatography 36: 127-162; and Griffin, et al, 1993. Appl. Biochem. Biotechnol. 38: 147-159).

Other methods for detecting mutations in the NOVX gene include methods in which protection from cleavage agents is used to detect mismatched bases in RNA/RNA or RNA/DNA heteroduplexes. See, e.g., Myers, et al, 1985. Science 230: 1242. In general, the art technique of "mismatch cleavage" starts by providing heteroduplexes of formed by hybridizing (labeled) RNA or DNA containing the wild-type NOVX sequence with potentially mutant RNA or DNA obtained from a tissue sample. The double-stranded duplexes are treated with an agent that cleaves single-stranded regions of the duplex such as which will exist due to basepair mismatches between the control and sample strands. For instance, RNA/DNA duplexes can be treated with RNase and DNA DNA hybrids treated with S_\ nuclease to enzymatically digesting the mismatched regions. In other embodiments, either DNA/DNA or RNA/DNA duplexes can be treated with hydroxylamine or osmium tetroxide and with piperidine in order to digest mismatched regions. After digestion of the mismatched regions, the resulting material is then separated by size on denaturing polyacrylamide gels to determine the site of mutation. See, e.g., Cotton, et al, 1988. Proc. Natl. Acad. Sci. USA 85: 4397; Saleeba, et al, 1992. Methods Enzymol. 217: 286-295. In an embodiment, the control DNA or RNA can be labeled for detection.

In still another embodiment, the mismatch cleavage reaction employs one or more proteins that recognize mismatched base pairs in double-stranded DNA (so called "DNA mismatch repair" enzymes) in defined systems for detecting and mapping point mutations in NOVX cDNAs obtained from samples of cells. For example, the mutY enzyme of E. coli cleaves A at G/A mismatches and the thymidine DNA glycosylase from HeLa cells cleaves T at G/T mismatches. See, e.g., Hsu, et al., 1994. Carcinogenesis 15: 1657-1662. According to an exemplary embodiment, a probe based on A NOVX sequence, e.g., a wild-type NOVX sequence, is hybridized to a cDNA or other DNA product from a test cell(s). The duplex is treated with a DNA mismatch repair enzyme, and the cleavage products, if any, can be detected from electrophoresis protocols or the like. See, e.g., U.S. Patent No. 5,459,039.

In other embodiments, alterations in electrophoretic mobility will be used to identify mutations in NOVX genes. For example, single strand conformation polymoφhism (SSCP) may be used to detect differences in electrophoretic mobility between mutant and wild type nucleic acids. See, e.g., Orita, et al, 1989. Proc. Natl. Acad. Sci. USA: 86: 2766; Cotton, 1993. Mutat. Res. 285: 125-144; Hayashi, 1992. Genet. Anal Tech. Appl. 9: 73-79. Single-stranded DNA fragments of sample and control NOVX nucleic acids will be denatured and allowed to renature. The secondary structure of single-stranded nucleic acids varies according to sequence, the resulting alteration in electrophoretic mobility enables the detection of even a single base change. The DNA fragments may be labeled or detected with labeled probes. The sensitivity of the assay may be enhanced by using RNA (rather than DNA), in which the secondary structure is more sensitive to a change in sequence. In one embodiment, the subject method utilizes heteroduplex analysis to separate double stranded heteroduplex molecules on the basis of changes in electrophoretic mobility. See, e.g., Keen, et al, 1991. Trends Genet. 1: 5. In yet another embodiment, the movement of mutant or wild-type fragments in polyacrylamide gels containing a gradient of denaturant is assayed using denaturing gradient gel electrophoresis (DGGΕ). See, e.g., Myers, et l, 1985. Nature 313: 495. When DGGΕ is used as the method of analysis, DNA will be modified to insure that it does not completely denature, for example by adding a GC clamp of approximately 40 bp of high-melting GC-rich DNA by PCR. In a further embodiment, a temperature gradient is used in place of a denaturing gradient to identify differences in the mobility of control and sample DNA. See, e.g., Rosenbaum and Reissner, 1987. Biophys. Chem. 265: 12753. Examples of other techniques for detecting point mutations include, but are not limited to, selective oligonucleotide hybridization, selective amplification, or selective primer extension. For example, oligonucleotide primers may be prepared in which the known mutation is placed centrally and then hybridized to target DNA under conditions that permit hybridization only if a perfect match is found. See, e.g., Saiki, et al, 1986. Nature 324: 163; Saiki, et al., 1989. Proc. Natl. Acad. Sci. USA 86: 6230. Such allele specific oligonucleotides are hybridized to PCR amplified target DNA or a number of different mutations when the oligonucleotides are attached to the hybridizing membrane and hybridized with labeled target DNA.

Alternatively, allele specific amplification technology that depends on selective PCR amplification may be used in conjunction with the instant invention. Oligonucleotides used as primers for specific amplification may cany the mutation of interest in the center of the molecule (so that amplification depends on differential hybridization; see, e.g., Gibbs, et al, 1989. Nucl. Acids Res. 17: 2437-2448) or at the extreme 3 '-terminus of one primer where, under appropriate conditions, mismatch can prevent, or reduce polymerase extension (see, e.g., Prossner, 1993. Tibtech. 11 : 238). In addition it may be desirable to introduce a novel restriction site in the region of the mutation to create cleavage-based detection. See, e.g., Gasparini, et al, 1992. Mol. Cell Probes 6: 1. It is anticipated that in certain embodiments amplification may also be performed using Taq ligase for amplification. See, e.g., Barany, 1991. Proc. Natl. Acad. Sci. USA 88: 189. In such cases, ligation will occur only if there is a perfect match at the 3'-terminus of the 5' sequence, making it possible to detect the presence of a known mutation at a specific site by looking for the presence or absence of amplification. The methods described herein may be performed, for example, by μtilizing pre-packaged diagnostic kits comprising at least one probe nucleic acid or antibody reagent described herein, which may be conveniently used, e.g. , in clinical settings to diagnose patients exhibiting symptoms or family history of a disease or illness involving A NOVX gene.

Furthermore, any cell type or tissue, preferably peripheral blood leukocytes, in which NOVX is expressed may be utilized in the prognostic assays described herein. However, any biological sample containing nucleated cells may be used, including, for example, buccal mucosal cells.

PHARMACOGENOMICS

Agents, or modulators that have a stimulatory or inhibitory effect on NOVX activity (e.g., NOVX gene expression), as identified by a screening assay described herein can be administered to individuals to treat (prophylactically or therapeutically) disorders (The disorders include metabolic disorders, diabetes, obesity, infectious disease, anorexia, cancer-associated cachexia, cancer, neurodegenerative disorders, Alzheimer's Disease, Parkinson's Disorder, immune disorders, and hematopoietic disorders, and the various dyslipidemias, metabolic disturbances associated with obesity, the metabolic syndrome X and wasting disorders associated with chronic diseases and various cancers.) In conjunction with such treatment, the pharmacogenomics (i.e., the study of the relationship between an individual's genotype and that individual's response to a foreign compound or drug) of the individual may be considered. Differences in metabolism of therapeutics can lead to severe toxicity or therapeutic failure by altering the relation between dose and blood concentration of the pharmacologically active drug. Thus, the pharmacogenomics of the individual permits the selection of effective agents (e.g., drugs) for prophylactic or therapeutic treatments based on a consideration of the individual's genotype. Such pharmacogenomics can further be used to determine appropriate dosages and therapeutic regimens. Accordingly, the activity of NOVX protein, expression of NOVX nucleic acid, or mutation content of NOVX genes in an individual can be determined to thereby select appropriate agent(s) for therapeutic or prophylactic treatment of the individual.

Pharmacogenomics deals with clinically significant hereditary variations in the response to drugs due to altered drug disposition and abnormal action in affected persons. See e.g. , Eichelbaum, 1996. Clin. Exp. Pharmacol. Physiol , 23 : 983-985; Linder, 1997. Clin. Chem., 43: 254-266. In general, two types of pharmacogenetic conditions can be differentiated. Genetic conditions transmitted as a single factor altering the way drugs act on the body (altered drug action) or genetic conditions transmitted as single factors altering the way the body acts on drugs (altered drug metabolism). These pharmacogenetic conditions can occur either as rare defects or as polymoφhisms. For example, glucose-6-phosphate dehydrogenase (G6PD) deficiency is a common inherited enzymopathy in which the main clinical complication is hemolysis after ingestion of oxidant drugs (anti-malarials, sulfonamides, analgesics, nitrofurans) and consumption of fava beans. As an illustrative embodiment, the activity of drug metabolizing enzymes is a major determinant of both the intensity and duration of drug action. The discovery of genetic polymoφhisms of drug metabolizing enzymes (e.g., N-acetyltransferase 2 (NAT 2) and cytochrome Pregnancy Zone Protein Precursor enzymes CYP2D6 and CYP2C19) has provided an explanation as to why some patients do not obtain the expected drug effects or show exaggerated drug response and serious toxicity after taking the standard and safe dose of a drug. These polymoφhisms are expressed in two phenotypes in the population, the extensive metabolizer (EM) and poor metabolizer (PM). The prevalence of PM is different among different populations. For example, the gene coding for

CYP2D6 is highly polymoφhic and several mutations have been identified in PM, which all lead to the absence of functional CYP2D6. Poor metabolizers of CYP2D6 and CYP2C19 quite frequently experience exaggerated drug response and side effects when they receive standard doses. If a metabolite is the active therapeutic moiety, PM show no therapeutic response, as demonstrated for the analgesic effect of codeine mediated by its C YP2D6-formed metabolite moφhine. At the other extreme are the so called ultra-rapid metabolizers who do not respond to standard doses. Recently, the molecular basis of ultra-rapid metabolism has been identified to be due to CYP2D6 gene amplification. Thus, the activity of NOVX protein, expression of NOVX nucleic acid, or mutation content of NOVX genes in an individual can be determined to thereby select appropriate agent(s) for therapeutic or prophylactic treatment of the individual. In addition, pharmacogenetic studies can be used to apply genotyping of polymoφhic alleles encoding drug-metabolizing enzymes to the identification of an individual's drug responsiveness phenotype. This knowledge, when applied to dosing or drug selection, can avoid adverse reactions or therapeutic failure and thus enhance therapeutic or prophylactic efficiency when treating a subject with A NOVX modulator, such as a modulator identified by one of the exemplary screening assays described herein.

MONITORING OF EFFECTS DURING CLINICAL TRIALS Monitoring the influence of agents (e.g. , drugs, compounds) on the expression or activity of NOVX (e.g., the ability to modulate abenant cell proliferation and/or differentiation) can be applied not only in basic drug screening, but also in clinical trials. For example, the effectiveness of an agent determined by a screening assay as described herein to increase NOVX gene expression, protein levels, or upregulate NOVX activity, can be monitored in clinical trails of subjects exhibiting decreased NOVX gene expression, protein levels, or downregulated NOVX activity. Alternatively, the effectiveness of an agent determined by a screening assay to decrease NOVX gene expression, protein levels, or downregulate NOVX activity, can be monitored in clinical trails of subjects exhibiting increased NOVX gene expression, protein levels, or upregulated NOVX activity. In such clinical trials, the expression or activity of NOVX and, preferably, other genes that have been implicated in, for example, a cellular proliferation or immune disorder can be used as a "read out" or markers of the immune responsiveness of a particular cell.

By way of example, and not of limitation, genes, including NOVX, that are modulated in cells by treatment with an agent (e.g., compound, drug or small molecule) that modulates NOVX activity (e.g., identified in a screening assay as described herein) can be identified. Thus, to study the effect of agents on cellular proliferation disorders, for example, in a clinical trial, cells can be isolated and RNA prepared and analyzed for the levels of expression of NOVX and other genes implicated in the disorder. The levels of gene expression (i.e., a gene expression pattern) can be quantified by Northern blot analysis or RT-PCR, as described herein, or alternatively by measuring the amount of protein produced, by one of the methods as described herein, or by measuring the levels of activity of NOVX or other genes. In this manner, the gene expression pattern can serve as a marker, indicative of the physiological response of the cells to the agent. Accordingly, this response state may be determined before, and at various points during, treatment of the individual with the agent.

In one embodiment, the invention provides a method for monitoring the effectiveness of treatment of a subject with an agent (e.g., an agonist, antagonist, protein, peptide, peptidomimetic, nucleic acid, small molecule, or other drug candidate identified by the screening assays described herein) comprising the steps of (i) obtaining a pre-administration sample from a subject prior to administration of the agent, (ii) detecting the level of expression of A NOVX protein, mRNA, or genomic DNA in the preadministration sample; (iii) obtaining one or more post-administration samples from the subject; (iv) detecting the level of expression or activity of the NOVX protein, mRNA, or genomic DNA in the post-administration samples; (v) comparing the level of expression or activity of the NOVX protein, mRNA, or genomic DNA in the pre-administration sample with the NOVX protein, mRNA, or genomic DNA in the post administration sample or samples; and (vi) altering the administration of the agent to the subject accordingly. For example, increased administration of the agent may be desirable to increase the expression or activity of NOVX to higher levels than detected, i.e., to increase the effectiveness of the agent. Alternatively, decreased administration of the agent may be desirable to decrease expression or activity of NOVX to lower levels than detected, i.e., to decrease the effectiveness of the agent.

Methods of Treatment The invention provides for both prophylactic and therapeutic methods of treating a subject at risk of (or susceptible to) a disorder or having a disorder associated with abenant NOVX expression or activity. The disorders include cardiomyopathy, atherosclerosis, hypertension, congenital heart defects, aortic stenosis, atrial septal defect (ASD), atrioventricular (A-V) canal defect, ductus arteriosus, pulmonary stenosis, subaortic stenosis, ventricular septal defect (VSD), valve diseases, tuberous sclerosis, scleroderma, obesity, transplantation, adrenoleukodystrophy, congenital adrenal hypeφlasia, prostate cancer, neoplasm; adenocarcinoma, lymphoma, uterus cancer, fertility, hemophilia, hypercoagulation, idiopathic thrombocytopenic puφura, immunodeficiencies, graft versus host disease, AIDS, bronchial asthma, Crohn's disease; multiple sclerosis, treatment of Albright Hereditary Ostoeodystrophy, and other diseases, disorders and conditions of the like.

These methods of treatment will be discussed more fully, below.

DISEASES AND DISORDERS Diseases and disorders that are characterized by increased (relative to a subject not suffering from the disease or disorder) levels or biological activity may be treated with Therapeutics that antagonize (i.e., reduce or inhibit) activity. Therapeutics that antagonize activity may be administered in a therapeutic or prophylactic manner. Therapeutics that may be utilized include, but are not limited to: (i) an aforementioned peptide, or analogs, derivatives, fragments or homologs thereof; (ii) antibodies to an aforementioned peptide; (iii) nucleic acids encoding an aforementioned peptide; (iv) administration of antisense nucleic acid and nucleic acids that are "dysfunctional" (i.e., due to a heterologous insertion within the coding sequences of coding sequences to an aforementioned peptide) that are utilized to "knockout" endogenous function of an aforementioned peptide by homologous recombination (see, e.g., Capecchi, 1989. Science 244: 1288-1292); or (v) modulators ( i.e., inhibitors, agonists and antagonists, including additional peptide mimetic of the invention or antibodies specific to a peptide of the invention) that alter the interaction between an aforementioned peptide and its binding partner. Diseases and disorders that are characterized by decreased (relative to a subject not suffering from the disease or disorder) levels or biological activity may be treated with Therapeutics that increase (i.e., are agonists to) activity. Therapeutics that upregulate activity may be administered in a therapeutic or prophylactic manner. Therapeutics that may be utilized include, but are not limited to, an aforementioned peptide, or analogs, derivatives, fragments or homologs thereof; or an agonist that increases bioavailability.

Increased or decreased levels can be readily detected by quantifying peptide and or RNA, by obtaining a patient tissue sample (e.g., from biopsy tissue) and assaying it in vitro for RNA or peptide levels, structure and/or activity of the expressed peptides (or mRNAs of an aforementioned peptide). Methods that are well-known within the art include, but are not limited to, immunoassays (e.g., by Western blot analysis, immunoprecipitation followed by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis, immunocytochemistry, etc.) and/or hybridization assays to detect expression of mRNAs (e.g., Northern assays, dot blots, in situ hybridization, and the like).

PROPHYLACTIC METHODS

In one aspect, the invention provides a method for preventing, in a subject, a disease or condition associated with an abenant NOVX expression or activity, by administering to the subject an agent that modulates NOVX expression or at least one NOVX activity. Subjects at risk for a disease that is caused or contributed to by abenant NOVX expression or activity can be identified by, for example, any or a combination of diagnostic or prognostic assays as described herein. Administration of a prophylactic agent can occur prior to the manifestation of symptoms characteristic of the NOVX abenancy, such that a disease or disorder is prevented or, alternatively, delayed in its progression. Depending upon the type of NOVX abenancy, for example, A NOVX agonist or NOVX antagonist agent can be used for treating the subject. The appropriate agent can be determined based on screening assays described herein. The prophylactic methods of the invention are further discussed in the following subsections.

Therapeutic Methods

Another aspect of the invention pertains to methods of modulating NOVX expression or activity for therapeutic puφoses. The modulatory method of the invention involves contacting a cell with an agent that modulates one or more of the activities of NOVX protein activity associated with the cell. An agent that modulates NOVX protein activity can be an agent as described herein, such as a nucleic acid or a protein, a naturally-occurring cognate ligand of A NOVX protein, a peptide, A NOVX peptidomimetic, or other small molecule. In one embodiment, the agent stimulates one or more NOVX protein activity. Examples of such stimulatory agents include active NOVX protein and a nucleic acid molecule encoding NOVX that has been introduced into the cell. In another embodiment, the agent inhibits one or more NOVX protein activity. Examples of such inhibitory agents include antisense NOVX nucleic acid molecules and anti-NOVX antibodies. These modulatory methods can be performed in vitro (e.g., by culturing the cell with the agent) or, alternatively, in vivo (e.g., by administering the agent to a subject). As such, the invention provides methods of treating an individual afflicted with a disease or disorder characterized by abenant expression or activity of A NOVX protein or nucleic acid molecule. In one embodiment, the method involves administering an agent (e.g., an agent identified by a screening assay described herein), or combination of agents that modulates (e.g. , up-regulates or down-regulates) NOVX expression or activity. In another embodiment, the method involves administering A NOVX protein or nucleic acid molecule as therapy to compensate for reduced or abenant NOVX expression or activity.

Stimulation of NOVX activity is desirable in s/twations in which NOVX is abnormally downregulated and/or in which increased NOVX activity is likely to have a beneficial effect. One example of such a situation is where a subject has a disorder characterized by abenant cell proliferation and/or differentiation (e.g. , cancer or immune associated disorders). Another example of such a situation is where the subject has a gestational disease (e.g., preclampsia).

Determination of the Biological Effect of the Therapeutic In various embodiments of the invention, suitable in vitro or in vivo assays are performed to determine the effect of a specific Therapeutic and whether its administration is indicated for treatment of the affected tissue. In various specific embodiments, in vitro assays may be performed with representative cells of the type(s) involved in the patient's disorder, to determine if a given Therapeutic exerts the desired effect upon the cell type(s). Compounds for use in therapy may be tested in suitable animal model systems including, but not limited to rats, mice, chicken, cows, monkeys, rabbits, and the like, prior to testing in human subjects. Similarly, for in vivo testing, any of the animal model system known in the art may be used prior to administration to human subjects.

Prophylactic and Therapeutic Uses of the Compositions of the Invention

The NOVX nucleic acids and proteins of the invention are useful in potential prophylactic and therapeutic applications implicated in a variety of disorders including, but not limited to: metabolic disorders, diabetes, obesity, infectious disease, anorexia, cancer-associated cancer, neurodegenerative disorders, Alzheimer's Disease, Parkinson's Disorder, immune disorders, hematopoietic disorders, and the various dyslipidemias, metabolic disturbances associated with obesity, the metabolic syndrome X and wasting disorders associated with chronic diseases and various cancers.

As an example, a cDNA encoding the NOVX protein of the invention may be useful in gene therapy, and the protein may be useful when administered to a subject in need thereof. By way of non-limiting example, the compositions of the invention will have efficacy for treatment of patients suffering from: metabolic disorders, diabetes, obesity, infectious disease, anorexia, cancer-associated cachexia, cancer, neurodegenerative disorders, Alzheimer's Disease, Parkinson's Disorder, immune disorders, hematopoietic disorders, and the various dyslipidemias. Both the novel nucleic acid encoding the NOVX protein, and the NOVX protein of the invention, or fragments thereof, may also be useful in diagnostic applications, wherein the presence or amount of the nucleic acid or the protein are to be assessed. A further use could be as an anti-bacterial molecule (i.e., some peptides have been found to possess anti-bacterial properties). These materials are further useful in the generation of antibodies, which immunospecifically-bind to the novel substances of the invention for use in therapeutic or diagnostic methods.

The invention will be further described in the following examples, which do not limit the scope of the invention described in the claims. EXAMPLES

Example A: Polynucleotide And Polypeptide Sequences, And Homology Data EXAMPLE 1.

The NOVl clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 1 A.

Table 1A. NOVl Sequence Analysis

SEQ ID NO: 1 1794 bp

NOVla, AGAAGCTCCCGAGTGTCCGGCCTAGAGGCCATGAGAAGGCAGTGGGGGTCTGCCATGA CGI00041-01 GGGCGGCCGAGCAGGCGGGCTGCATGGTGAGCGCCTCCCGGGCCGGACAGCCCGAGGC DNA Sequence GGGCCCGTGGAGCTGCAGCGGGGTAATCCTGAGCCGTAGCCCGGGCCTGGTGCTTTGC

CACGGGGGCATCTTCGTCCCCTTCCTGCGAGCTGGCAGCGAAGTCCTGACCCGGCCCG

GCGCCGTCTTCCTGCCTGGCGACAGTTGCAGGGACGACCTGCGCCTGCACGTGCAGTG'

GGCCCCAACGGCAGCTGCCCCGTCTGGGAAGTGGGGAGTGCCTCTGCCCGGCCGCCCC

GTCTGGGAAGTGAGGAGTGCCTCTGCCCGGCCGCCCATCATCTGGGATGTGAGGAGCG

CCCCTCCTTCTCCTCCTCCTCCTCCCCTCCTCCTCCTGCTTGTTTCCCTTTGCTCCTT

CTTTTTAGCCCACTTTAGTCTAAAATATGGATCATGTTTTAAAAATACATTTTATTTT

GTTAGAGCAGCCCAGGCAGACACTAAGACGCTCACAGAACAGGAGGGAAGTTTAAAGA

CGCTGGGCTGGTTTGCGCTGCTGGGCGTGCGGCTAGGCCAGGAAGAGTGGAGGAGACG

CGGGCCAACGATGGCGGTGTCGCCTCTCGGGGCCGTGCCCAAGGGTGCGCCATTGCTG

GTCTGCGGCTCCCCTTTCGGCGCCTTCTGCCCCGACATCTTTCTCAACACGCTGAGCT

GCGGGGTGCTCAGCAACGTGGCCGGCCCACTGCTGCTTACCGACGCACGCTGCCTGCC

CGGCACCGAGGGCGGCGGCGTGTTCACCGCGCGGCCCGCGGGGGCGCTGGTGGCGCTG

GTGGTGGCGCCGCTCTGTTGGAAGGCCGGCGAATGGGTGGGCTTCACGCTGCTCTGCG

CCGCCGCCCCCCTTTTCCGCGCCGCCCGCGACGCGCTTCACCGCCTGCCGCACAGCAC

CGCTGCCCTGGCCGCCCTTCTGCCGCCAGAGGTGGGCGTCCCGTGGGGTCTGCCCCTC

CGAGACTCCGGGCCCCTGTGGGCAGCCGCGGCAGTGTTGGTGGAGTGCGGCACCGTAT

GGGGCTCCGGAGTGGCTGTGGCACCCCGCCTTGTAGTGACCTGTCGGCACGTGTCCCC

TCGGGAAGCAGCCAGGGTCCTGGTGCGCTCCACCACCCCCAAGAGTGTGGCCATCTGG

GGCCGTGTGGTATTTGCCACTCAGGAGACATGTCCCTATGACATAGCAGTGGTGAGCC

TGGAGGAGGACCTGGATGATGTCCCCATCCCTGTGCCCGCTGAGCACTTCCATGAAGG

CGAGGCTGTGAGTGTGGTGGGCTTTGGCGTCTTTGGCCAGTCTTGCGGGCCCTCGGTG

ACCTCAGGCATCCTTTCGGCTGTGGTGCAGGTGAATGGCACGCCCGTAATGCTGCAGA

CCACGTGTGCTGTGCACAGCGGCTCCAGTGGGGGACCCCTCTTCTCCAACCACTCAGG

AAACCTCCTTGGTATAATCACCAGCAACACCCGGGACAATAATACGGGGGCCACCTAC

CCCCACCTGAACTTCAGCATTCCCATCACGGTGCTCCAGCCGGCCCTGCAGCAGTACA

GCCAGACCCAAGACCTAGGTGGCCTCCGTGAGCTGGACCGCGCTGCTGAGCCAGTCAG

GGTGGTGTGGCGGTTGCAGCGGCCCCTGGCAGAGGCCCCGCGGAGCAAGCTCTGAGGC

TGTGTTACCACCTTTGGAAAGAAGAGTGACCTTTTTCTGCTGTAGGAAGTGATG

ORF Start: ATG at 31 ORF Stop: TGA at 1735

SEQ ID NO: 2 568 aa MW at 60004.6kD

NOVla, RRQ GSAMRAAEQAGCMVSASRAGQPEAGPWSCSGVILSRSPGLVLCHGGIFVPFLR CG100041-01 AGSEVLTRPGAVFLPGDSCRDDLRLHVQWAPTAAAPSGKWGVPLPGRPVWEVRSASAR Protein PPIIVROVRSAPPSPPPPPLLLLLVSLCSFFI-AHFSLKYGSCFKNTFYFVRAAQADTKT Sequence TEQEGSLKTLGWFALLGVRLGQEEMRRRGPTMAVSPLGAVPKGAPLLVCGSPFGAFC PDIFIJSΓTLSCGVLSNVAGPLLLTDARCLPGTEGGGVFTARPAGALVALVVAPLC KAG E VGFTLLCAAAPLFRAARDALHRLPHSTAALAALLPPEVGVP GLPLRDSGPL AAA AVLVECGTWGSGVAVAPRLWTCRHVSPREAARVLVRSTTPKSVAIWGRWFATQET CPYDIAWSLEEDLDDVPIPVPAEHFHEGEAVSWGFGVFGQSCGPSVTSGILSAWQ VNGTPVMLQTTCAVHSGSSGGPLFSIRASGNLLGIITSNTRDNNTGATYPHLNFSIPIT VLQPALQQYSQTQDLGGLREI-DRAAEPVRVVWRLQRPLAEAPRSKL Further analysis of the NOVla protein yielded the following properties shown in Table IB.

Table IB. Protein Sequence Properties NOVla

PSort analysis: 0.8741 probability located in microbody (peroxisome); 0.8266 probability located in mitochondrial inner membrane; 0.6500 probability located in plasma membrane; 0.3000 probability located in Golgi body

SignalP analysis: No Known Signal Sequence Predicted

A search of the NOVla protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table IC.

In a BLAST search of public sequence databases, the NOVla protein was found to have homology to the proteins shown in the BLASTP data in Table ID.

PFam analysis predicts that the NOVl a protein contains the domains shown in the Table IE.

EXAMPLE 2.

The NOV2 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 2A.

Table 2A. NOV2 Sequence Analysis

SEQ ID NO: 3 2274 bp

NOV2a, ATGGTCCCCGACACCGCCTGCGTTCTTCTGCTCACCCTGGCTGCCCTCGGCGCGTCCG CG105716- GACAGGGCCAGAGCCCGTTGGGCTCAGACCTGGGCCCGCAGATGCTTCGGGAACTGCA 01 DNA GGAAACCAACGCGGCGCTGCAGGACGTGCGGGACTGGCTGCGGCAGCAGGTCAGGGAG Sequence ATCACGTTCCTGAAAAACACGGTGATGGAGTGTGACGCGTGCGGGATGCAGCAGTCAG TACGCACCGGCCTACCCAGCGTGCGGCCCCTGCTCCACTGCGCGCCCGGCTTCTGCTT CCCCGGCGTGGCCTGCATCCAGACGGAGAGCGGCGGCCGCTGCGGCCCCTGCCCCGCG GGCTTCACGGGCAACGGCTCGCACTGCACCGACGTCAACGAGTGCAACGCCCACCCCT GCTTCCCCCGAGTCCGCTGTATCAACACCAGCCCGGGGTTCCGCTGCGAGGCTTGCCC GCCGGGGTACAGCGGCCCCACCCACCAGGGCGTGGGGCTGGCTTTCGCCAAGGCCAAC AAGCAGGTTTGCACGGACATCAACGAGTGTGAGACCGGGCAACATAACTGCGTCCCCA ACTCCGTGTGCATCAACACCCGGGGCTCCTTCCAGTGCGGCCCGTGCCAGCCCGGCTT CGTGGGCGACCAGGCGTCCGGCTGCCAGCGGCGCGCACAGCGCTTCTGCCCCGACGGC TCGCCCAGCGAGTGCCACGAGCATGCAGACTGCGTCCTAGAGCGCGATGGCTCGCGGT CGTGCGTGTGTGCCGTTGGCTGGGCCGGCAACGGGATCCTCTGTGGTCGCGACACTGA CCTAGACGGCTTCCCGGACGAGAAGCTGCGCTGCCCGGAGCGCCAGTGCCGTAAGGAC AACTGCGTGACTGTGCCCAACTCAGGGCAGGAGGATGTGGACCGCGATGGCATCGGAG ACGCCTGCGATCCGGATGCCGACGGGGACGGGGTCCCCAATGAAAAGGACAACTGCCC GCTGGTGCGGAACCCAGACCAGCGCAACACGGACGAGGACAAGTGGGGCGATGCGTGC GACAACTGCCGGTCCCAGAAGAACGACGACCAAAAGGACACAGACCAGGACGGCCGGG GCGATGCGTGCGACGACGACATCGACGGCGACCGGATCCGCAACCAGGCCGACAACTG CCCTAGGGTACCCAACTCAGACCAGAAGGACAGTGATGGCGATGGTATAGGGGATGCC TGTGACAACTGTCCCCAGAAGAGCAACCCGGATCAGGCGGATGTGGACCACGACTTTG TGGGAGATGCTTGTGACAGCGATCAAGACCAGGATGGAGACGGACATCAGGACTCTCG GGACAACTGTCCCACGGTGCCTAACAGTGCCCAGGAGGACTCAGACCACGATGGCCAG GGTGATGCCTGCGACGACGACGACGACAATGACGGAGTCCCTGACAGTCGGGACAACT GCCGCCTGGTGCCTAACCCCGGCCAGGAGGACGCGGACAGGGACGGCGTGGGCGACGT GTGCCAGGACGACTTTGATGCAGACAAGGTGGTAGACAAGATCGACGTGTGTCCGGAG AACGCTGAAGTCACGCTCACCGACTTCAGGGCCTTCCAGACAGTCGTGCTGGATCCTG AAGGGGATGCCCAGATCGATCCCAACTGGGTGGTCCTGAACCAGGGCATGGAGATTGT ACAGACCATGAACAGTGATCCTGGCCTGGCAGTGGGGTACACAGCTTTTAATGGAGTT GACTTCGAAGGGACCTTCCATGTGAATACCCAGACAGATGATGACTATGCAGGCTTTA TCTTTGGCTACCAAGATAGCTCCAGCTTCTACGTGGTCATGTGGAAGCAGACGGAGCA GACATATTGGCAAGCCACCCCATTCCGAGCAGTTGCAGAACCTGGCATTCAGCTCAAG GCTGTGAAGTCTAAGACAGGTCCAGGGGAGCATCTCCGGAACGCTCTGTGGCATACAG GAGACACAGAGTCCCAGGTGCGGCTGCTGTGGAAGGACCCGCGAAACGTGGGTTGGAA GGACAAGAAGTCCTATCGTTGGTTCCTGCAGCACCGGCCCCAAGTGGGCTACATCAGG GTGCGATTCTATGAGGGCCCTGAGCTGGTGGCCGACAGCAACGTGGTCTTGGACACAA CCATGCGGGGTGGCCGCCTGGGGGTCTTCTGCTTCTCCCAGGAGAACATCATCTGGGC CAACCTGCGTTACCGCTGCAATGACACCATCCCAGAGGACTATGAGACCCATCAGCTG CGGCAAGCCTAG

ORF Start: ATG at 1 ORF Stop: TAG at 2272

SEQ ID NO: 4 757 aa MW at 82915.7kD

NOV2a, MVPDTACVLLLTLAALGASGQGQSPLGSDLGPQMLRELQETNAALQDVRDWLRQQVRE CG105716- ITFLKNTVMECDACGMQQSVRTGLPSVRPLLHCAPGFCFPGVACIQTESGGRCGPCPA 01 Protein GFTGNGSHCTDVNECNAHPCFPRVRCINTSPGFRCEACPPGYSGPTHQGVGLAFAKAN Sequence KQVCTDI ECETGQHNCVPNSVCINTRGSFQCGPCQPGFVGDQASGCQRRAQRFCPDG SPSECHEHADCVLERDGSRSCVCAVG AGNGILCGRDTDLDGFPDEKLRCPERQCRKD NCVTVPNSGQEDVDRDGIGDACDPDADGDGVPNEKDNCPLVRNPDQRNTDEDK GDAC DNCRSQKNDDQKDTDQDGRGDACDDDIDGDRIR QADNCPRVPNSDQKDSDGDGIGDA CDNCPQKSNPDQADVDHDFVGDACDSDQDQDGDGHQDSRDNCPTVPNSAQEDSDHDGQ GDACDDDDDNDGVPDSRDNCRLVPNPGQEDADRDGVGDVCQDDFDADKWDKIDVCPE NAEVTLTDFRAFQTVVIiDPEGDAQIDPlsrWVVI^QGMEIVQT NSDPGI-AVGYTAFNGV DFEGTFHVNTQTDDDYAGFIFGYQDSSSFYVVM KQTEQTY QATPFRAVAEPGIQLK AVKSKTGPGEHLIWALWHTGDTESQVRLLWKDPRNVG KDKKSYR FLQHRPQVGYIR VRFYEGPELVADSlsnTv/LDTTMRGGRLGVFCFSQENII ANLRYRCNDTIPEDYETHQL RQA

Further analysis of the NOV2a protein yielded the following properties shown in Table 2B.

Table 2B. Protein Sequence Properties NOV2a

PSort analysis: 0.5278 probability located in outside; 0.1900 probability located in lysosome (lumen); 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen)

SignalP analysis: Cleavage site between residues 21 and 22

A search of the NOV2a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 2C.

In a BLAST search of public sequence databases, the NOV2a protein was found to have homology to the proteins shown in the BLASTP data in Table 2D.

PFam analysis predicts that the NOV2a protein contains the domains shown in the Table 2E.

EXAMPLE 3.

The NOV3 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 3 A.

Table 3A. NOV3 Sequence Analysis

SEQ ID NO: 5 799 bp

NOV3a, CGGCCGCGTCGACCGGGCCCCGAGGCACAGCCAGGGCACCAGGTGGAGCACCAGCTAC CGI 13569-01 GCGTGGCGCAGCGCAGCGTCCCTAGCACCGAGCCTCCCGCAGCCGCCGAGATGCTGCG DNA Sequence AACAGAGAGCTGCCGCCCCAGGTCGCCCGCCGGACAGGTGGCCGCGGCGTCCCCGCTC CTGCTGCTGCTGCTGCTGCTCGCCTGGTGCGCGGGCGCCTGCCGAGGTGCTCCAATAT TACCTCAAGGATTACAGCCTGAACAACAGCTACAGTTGTGGAATGAGGCATCCAACGC ACTGGAGGAGCTTTGCTTTATGATTATGGGAATGCTACCAAΆGCCTCAGGAACAAGAT GAAAAAGATAATACTAAAAGGTTCTTATTTCATTATTCGAAGACACAGAAGTTGGGCA AGTCAAATGTTGTGTCGTCAGTTGTGCATCCGTTGCTGCAGCTCGTTCCTCACCTGCA TGAGAGAAGAATGAAGAGATTCAGAGTGGACGAAGAATTCCAAAGTCCCTTTGCAAGT CAAAGTCGAGGATATTTTTTATTCAGGCCACGGAATGGAAGAAGGTCAGCAGGGTTCA TTTAAAATGGATGCCAGCTAATTTTCCACAGAGCAATGCTATGGAATACAAAATGTAC TGACATTTTGTTTTCTTCTGAAAAAAAATCCTTGCTAAATGTACTCTGTTGAAAATCC

CTGTGTTGTCAATGTTCTCAGTTGTAACAATGTTGTAAATGTTCAATTTGTTGAAAAT AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAACCAAAAAAAT

ORF Start: ATG at 109 ORF Stop: TAA at 583

SEQ ID NO: 6 158 aa MW at 18002.8kD

NOV3a, MLRTESCRPRSPAGQVAAASPLLLLLLLLA CAGACRGAPILPQGLQPEQQLQLW EA CGI 13569-01 SNALEELCFMIMGMLPKPQEQDE DNTKRFLFHYSKTQKLGKSNVVSSVVHPLLQLVP Protein HLHERRMKRFRVDEEFQSPFASQSRGYFLFRPRNGRRSAGFI Sequence

SEQ ID NO: 7 746 bp

NOV3b, AGTCCTGCGTCCGGGCCCCGAGGCACAGCCAGGGCACCAGGTGGAGCACCAGCTACGC CGI 13569-03 GTGGCGCAGCGCAGCGTCCCTAGCACCGAGCCTCCCGCAGCCGCCGAGATGCTGCGAA DNA Sequence CAGAGAGCTGCCGCCCCAGGTCGCCCGCCGGACAGGTGGCCGCGGCGTCCCCGCTCCT GCTGCTGCTGCTGCTGCTCGCCTGGTGCGCGGGCGCCTGCCGAGGTGCTCCAATATTA CCTCAAGGATTACAGCCTGAACAACAGCTACAGTTGTGGAATGAGGCATCCAACGCAC TGGAGGAGCTTTGCTTTATGATTATGGGAATGCTACCAAAGCCTCAGGAACAAGATGA AAAAGATAATACTAAAAGGTTCTTATTTCATTATTCGAAGACACAGAAGTTGGGCAAG TCAAATGTTGTGTCGTCAGTTGTGCATCCGTTGCTGCAGCTCGTTCCTCACCTGCATG AGAGAAGAATGAAGAGATTCAGAGTGGACGAAGAATTCCAAAGTCCCTTTGCAAGTCA AAGTCGAGGATATTTTTTATTCAGGCCACGGAATGGAAGAAGGTCAGCAGGGTTCATT TAAAATGGATGCCAGCTAATTTTCCACAGAGCAATGCTATGGAATACAAAATGTACTG ACATTTTGTTTTCTTCTGAAAAAAATCCTTGCTAAATGTACTCTGTTGAAAATCCCTG

TGTTGTCAATGTTCTCAGTTGTAACAATGTTGTAAATGTTCAATTTGTTG

ORF Start: ATG at 107 ORF Stop: TAA at 581

SEQ ID NO: 8 158 aa MW at 18002.8kD

NOV3b, MLRTESCRPRSPAGQVAAASPLLLLLLLLAWCAGACRGAPILPQGLQPEQQLQLWNEA CGI 13569-03 SN EELCFMI GML K QEQDEKD r -^FLFH SK QI< GKSNVVSSVVHP QL Protein HLHERRMKRFRVDEEFQSPFASQSRGYFLFRPRNGRRSAGFI Sequence

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 3B. Table 3B. Comparison of NOV3a against NOV3b.

NOV3a Residues/ Identities/

Protein Sequence Match Residues Similarities for the Matched Region

NOV3b 1..158 118/158 (74%) 1..158 118/158 (74%)

Further analysis of the NOV3a protein yielded the following properties shown in Table 3C.

Table 3C. Protein Sequence Properties NOV3a

PSort analysis: 0.8200 probability located in outside; 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen); 0.1000 probability located in lysosome (lumen)

SignalP analysis: Cleavage site between residues 39 and 40

A search of the NOV3a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 3D.

In a BLAST search of public sequence databases, the NOV3a protein was found to have homology to the proteins shown in the BLASTP data in Table 3E.

PFam analysis predicts that the NOV3a protein contains the domains shown in the Table 3F.

EXAMPLE 4.

The NOV4 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 4A.

Table 4A. NOV4 Sequence Analysis

SEQ ID NO: 9 1397 bp

NOV4a, TCGCTGCCGCCGCTCCCAGCAACAAATCACAGCTGCATTTTCTCGTTGATCCAGTCGA CG56602-02 TGTAGGCGGACACCCGGGTGTAGACTACCGGCTTCTTGCGGGTGTTGCAGCCCCGCCG DNA Sequence GGAGCCAGTCCTGAGCACCTAACCATGTTGGGCATCACTGTCCTCGCTGCGCTCTCCG

CCAGTGCCTCCAGCTGTGGGGTGCCCAGCTTCCCGCCCAACCTATCCGCCCGAGTGGT GGGAGGAGAGGATGCCCGGCCCCACAGCTGGCCCTGGCAGGTAAGCCTGCTCCAGTAC CTCAAGAACGACACGTGGAGGCATACGTGTGGTGGGACTTTGATTGCTAGCAACTTCG TCCTCACTGCCGCCCACTGCATCAGCAACACCCGGACCTACCGTGTGGCCGTGGGAAA GAACAACCTGGAGGTGGAAGACGAAGAAGGATCCCTGTTTGTGGGTGTGGACACCATC CACGTCCACAAGAGATGGAATGCCTTGGATTCCAGCAATGATATTGCCCTCATCAAGC TTGCAGAGCATGTGGAGCTGAGTGACACCATCCAGGTGGCCTGCCTGCCAGAGAAGGA CTCCCTGCTCCCCAAGGACTACCCCTGCTATGTCACCGGCTGGGGCCGCCTCAACGGC CCCATTGCTGATAAGCTGCAGCAGGGCCTGCAGCCCGTGGTGGATCACGCCACGTGCT CCAGGATTGACTGGTGGGGCTTCAGGGTGAAGAAAACCATGGTGTGCGCTGGGGGCGA TGGCGTCATCTCAGCCTGCAATGGGGACTCCGGTGGCCCACTGAACTGCCAGTTGGAG AACGGTTCCTGGGAGGTGTTTGGCATCGTCAGCTTTGGCTCCCGGCGGGGCTGCAACA CCCGCAAGAAGCCGGTAGTCTACACCCGGGTGTCCGCCTACATCGACTGGATCAACGA GGTGGGTGCTGCCTCCACAGCTGTCCCTGCACCTGTCAGCCCCTCCCCCTCACTCACC CATCCCCTCACTCATTCACTCATTCATGCGTTTATTCATTCATTCATTTATTCACTCA TTCATGCATTTATTCACTCATTCATGCATTCATTCATTTATTCACTTATTCAGTCACT CATTCATGTATTTATTCATTTATTCATTCACTCATGCATTTATTCATTCATTCATTTA TTCACTTATTCATTCACTCATTCATGTATTCATTCATTCATTCATGCATTTATTTACT CATTCATCCATTTATTCACTCATTCATTTGCTCATTCAGTGATTCATTCATGCACTTC TTCACACATTCACTCTCTCATTCAAGTAATATTGGTTGAGTGCTTCCAGTAGCAGGCC

TTGAGTTGGGTGCCAATAAGGAAACAGTCATTGACTCCTCCATCCATCCATTCACTGC

CTTCA

ORF Start: ATG at 141 ORF Stop: TAG at 1326

SEQ ID NO: 10 395 aa MW at 43653.5kD

NOV4a, MLGITVIJAALSASASSCGVPSFPPNLSARVVGGEDARPHS P QVSLLQYLKNDTWRH CG56602-02 CGGTLIASNI Π^TAAHCISNTRTYRVAVGKNI^EVEDEEGSLFVGVDTIHVHKRWNA Protein LDSSOT)IALIKTΙAEHVELSDTIQVACLPE ΩSLLPKDYPCYVTGWGRI_J GPIADKLQQ Sequence GLQPVVDI^TCSRID GFRV KT VCAGGDGVISACNGDSGGPLNCQLENGSWEVFG IVSFGSRRGCNTRKKPWYTRVSAYID INEVGAASTAVPAPVSPSPSLTHPLTHS I HAFIHSFIYSLIHAFIHSF HSFIYSLIQSLIHVFIHLFIHSCIYSFIHLFTYSFTHS CIHSFIHAFIYSFIHLFTHSFAHSVIHSCTSSHIHSLIQVILVECFQ

Further analysis of the NOV4a protein yielded the following properties shown in Table 4B.

Table 4B. Protein Sequence Properties NOV4a

PSort analysis: 0.4600 probability located in plasma membrane; 0.2409 probability located in microbody (peroxisome); 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen)

SignalP analysis: Cleavage site between residues 17 and 18

A search of the NOV4a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 4C.

In a BLAST search of public sequence databases, the NOV4a protein was found to have homology to the proteins shown in the BLASTP data in Table 4D.

PFam analysis predicts that the NOV4a protein contains the domains shown in the Table 4E.

EXAMPLE 5.

The NOV5 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 5 A.

Table 5A. NOV5 Sequence Analysis

SEQ ID NO: 11 3210 bp

NOV5a, CAATTCTATTCGCTTGTTATTGGACTTGAAACTCCCTTTGACCTCGGAAACTGAAGAT CG57415-01 GAGGTTGCCATGGGAACTGCTGGTACTGCAATCATTCATTTTGTGCCTTGCAGATGAT DNA Sequence TCCACACTGCATGGCCCGATTTTTATTCAAGAACCAAGTCCTGTAATGTTCCCTTTGG

ATTCTGAGGAGAAAAAAGTGAAGCTCAATTGTGAAGTTAAAGGAAATCCAAAACCTCA

TATCAGGTGGAAGTTAAATGGAACAGATGTTGACACTGGTATGGATTTCCGCTACAGT

GTTGTTGAAGGGAGCTTGTTGATCAATAACCCCAATAAAACCCAAGATGCTGGAACGT

ACCAGTGCACAGCGACAAACTCGTTTGGAACAATTGTTAGCAGAGAAGCAAAGCTTCA

GTTTGCTTATCTTGACAACTTTAAAACAAGAACAAGAAGCACTGTGTCTGTCCGTCGA

GGTCAAGGAATGGTGCTACTGTGTGGCCCGCCACCCCATTCTGGAGAGCTGAGTTATG

CCTGGATCTTCAATGAATACCCTTCCTATCAGGATAATCGCCGCTTTGTTTCTCAAGA

GACTGGGAATCTGTATATTGCCAAAGTAGAAAAATCAGATGTTGGGAATTATACCTGT

GTGGTTACCAATACCGTGACAAACCACAAGGTCCTGGGGCCACCTACACCACTAATAT

TGAGAAATGATGGAGTGATGGGTGAATATGAGCCCAAAATAGAAGTGCAGTTCCCAGA

AACAGTTCCGACTGCAAAAGGAGCAACGGTGAAGCTGGAATGCTTTGCTTTAGGAAAT

CCAGTACCAACTATTATCTGGCGAAGAGCTGATGGAAAGCCAATAGCAAGGAAAGCCA

GAAGACACAAGTCAAATGGAATTCTTGAGATCCCTAATTTTCAGCAGGAGGATGCTGG

TTTATATGAATGTGTAGCTGAAAATTCCAGAGGGAAAAATGTAGCAAGGGGACAGCTA

ACTTTCTATGCTCAACCTAATTGGATTCAAAAAATAAATGATATTCACGTGGCCATGG

AAGAAAATGTCTTTTGGGAATGTAAAGCAAATGGAAGGCCTAAGCCTACATACAAGTG

GCTAAAAAATGGCGAACCTCTGCTAACTCGGGATAGAATTCAAATTGAGCAAGGAACA

CTCAACATAACAATAGTGAACCTCTCAGATGCTGGCATGTATCAGTGTTTGGCAGAGA

ATAAACATGGAGTTATCTTTTCCAACGCAGAGCTTAGTGTTATAGCTGTAGGTCCAGA

TTTTTCAAGAACACTCTTGAAAAGAGTAACTCTTGTCAAAGTGGGAGGTGAAGTTGTC

ATTGAGTGTAAGCCAAAAGCGTCTCCAAAACCTGTTTACACCTGGAAGAAAGGAAGGG

ATATATTAAAAGAAAATGAAAGAATTACCATTTCTGAAGATGGAAACCTCAGAATCAT

CAACGTTACTAAATCAGACGCTGGGAGTTATACCTGTATAGCCACTAACCATTTTGGA

ACTGCTAGCAGTACTGGAAACTTGGTAGTGAAAGATCCAACAAGGGTAATGGTACCCC

CTTCCAGTATGGATGTCACTGTTGGAGAGAGTATTGTTTTACCGTGCCAGGTAACGCA

TGATCACTCGCTAGACATCGTGTTTACTTGGTCATTTAATGGACACCTGATAGACTTT

GACAGAGATGGGGACCACTTTGAAAGAGTTGGAGGGCAGGATTCAGCTGGTGATTTGA

TGATCCGAAACATCCAACTGAAGCATGCTGGGAAATATGTCTGCATGGTCCAAACAAG

TGTGGACAGGCTATCTGCTGCTGCAGACCTGATTGTAAGAGGTCCTCCAGGTCCCCCA

GAGGCTGTGACAATAGACGAAATCACAGATACCACTGCTCAGCTCTCCTGGAGACCCG

GGCCTGACAACCACAGCCCCATCACCATGTATGTCATTCAAGCCAGGACTCCATTCTC

CGTGGGCTGGCAAGCAGTCAGTACAGTCCCAGAACTCATTGATGGGAAGACATTCACA

GCGACCGTGGTGGGTTTGAACCCTTGGGTTGAATATGAATTCCGCACAGTTGCAGCCA

ACGTGATTGGGATTGGGGAGCCCAGCCGCCCCTCAGAGAAACGGAGAACAGAAGAAGC

TCTCCCCGAAGTCACACCAGCGAATGTCAGTGGTGGCGGAGGCAGCAAATCTGAACTG

GTTATAACCTGGGAGACGGTCCCTGAGGAATTACAGAATGGTCGAGGCTTTGGTTATG

TGGTGGCCTTCCGGCCCTACGGTAAAATGATCTGGATGCTGACAGTGCTGGCCTCAGC

TGATGCCTCTAGATACGTGTTCAGGAATGAGAGCGTGCACCCCTTCTCTCCCTTTGAG

GTTAAAGTAGGTGTCTTCAACAACAAAGGAGAAGGCCCTTTCAGTCCCACCACGGTGG

TGTATTCTGCAGAAGAAGAACCCACCAAACCACCAGCCAGTATCTTTGCCAGAAGTCT

TTCTGCCACAGATATTGAAGTTTTCTGGGCCTCCCCACTGGAGAAGAATAGAGGACGA

ATACAAGGTTATGAGGTTAAATATTGGAGACATGAAGACAAAGAAGAAAATGCTAGAA

AAATACGAACAGTTGGAAATCAGACATCAACAAAAATCACGAACTTAAAAGGCAGTGT

GCTGTATCACTTAGCTGTCAAGGCATATAATTCTGCTGGGACAGGCCCCTCTAGTGCA

ACAGTCAATGTGACAACCCGAAAGCCACCACCAAGTCAACCCCCCGGGAACATCATAT

GGAATTCATCAGACTCCAAAATTATTCTGAATTGGGATCAAGTGAAGGCCCTGGATAA

TGAGTCGGAAGTAAAAGGATACAAAGTAGTCTTGTACAGATGGAACAGACAAAGCAGC

—

Further analysis of the NOV5a protein yielded the following properties shown in

Table 5B.

A search of the NOV5a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 5C.

AAW29667 Homo sapiens DL185_ 1 clone 1..1000 617/1003 (61%) 0.0 secreted protein - Homo sapiens, 1..1001 775/1003 (76%) 1028 aa. [WO9830695-A2, 16-JUL- 1998]

AAU 18339 Human endocrine polypeptide SEQ 582..1027 445/446 (99%) 0.0 ID No 294 - Homo sapiens, 447 aa. 3..447 445/446 (99%) [WO200155364-A2, 02-AUG-2001]

AAM43534 Human polypeptide SEQ ID NO 212 578..1027 442/450 (98%) 0.0 - Homo sapiens, 456 aa. 8..456 444/450 (98%) [WO200155308-A2, 02-AUG-2001]

AAR87028 Human contactin - Homo sapiens, 25..986 439/965 (45%) 0.0 1018 aa. [WO9535373-A2, 28-DEC- 40..989 603/965 (61%) 1995]

In a BLAST search of public sequence databases, the NOV5a protein was found to have homology to the proteins shown in the BLASTP data in Table 5D.

PFam analysis predicts that the NOV5a protein contains the domains shown in the Table 5E.

EXAMPLE 6.

The NOV6 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 6A.

Table 6A. NOV6 Sequence Analysis

SEQ ID NO: 13 5115 bp

NOV6a, GAATTCCGGGAGCGGGCGGGCTGCGAGGCCGCGGGGCATGCGGGAGGCGGAGGGGTGG CG58504-01 GACCGGGTGGCTGCGCCCATTCCACACCCGCCGAAAGCGGACACTGTCAGCTGAATCA DNA Sequence CTCCCCTTTTAGGAGGAGGGAGGGGGAAAAGGTGTCTAGCTAATTTCTGCTTAAAAAA

GCACAGGAGATCGCGGGTCAGCTTTGCAGTCGCTGCCTTCTCGCGCCTGACCATGCAC

CCCTGCATCTTCCTGCTGGGCACAGGCGAGCGCTTTATTTCTGGAGCTGAGGGCTAAA

ACTTTTTTCACTTTTCTTCTCCTCAACATCTGAATCATGCCATGTGCCCAGAGGAGCT

GGCTTGCAAACCTTTCCGTGGTGGCTCAGCTCCTTAACTTTGGGGCGCTTTGCTATGG GAGACAGCCTCAGCCAGGCCCGGTTCGCTTCCCGGACAGGAGGCAAGAGCATTTTATC AAGGGCCTGCCAGAATACCACGTGGTGGGTCCAGTCCGAGTAGATGCCAGTGGGCATT TTTTGTCATATGGCTTGCACTATCCCATCACGAGCAGCAGGAGGAAGAGAGATTTGGA TGGCTCAGAGGACTGGGTGTACTACAGAATTTCTCACGAGGAGAAGGACCTGTTTTTT AACTTGACGGTCAATCAAGGATTTCTTTCCAATAGCTACATCATGGAGAAGAGATATG GGAACCTCTCCCATGTTAAGATGATGGCTTCCTCTGCCCCCCTCTGCCATCTCAGTGG CACGGTTCTACAGCAGGGCACCAGAGTTGGGACGGCAGCCCTCAGTGCCTGCCATGGA CTGACTGGATTTTTCCAACTACCACATGGAGACTTTTTCATTGAACCCGTGAAGAAGC ATCCACTGGTTGAGGGAGGGTACCACCCGCACATCGTTTACAGGAGGCAGAAAGTTCC AGAAACCAAGGAGCCAACCTGTGGATTAAAGGACAGTGTTAACATCTCCCAGAAGCAA GAGCTATGGCGGGAGAAGTGGGAGAGGCACAACTTGCCAAGCAGAAGCCTCTCTCGGC GTTCCATCAGCAAGGAGAGATGGGTGGAGACACTGGTGGTGGCCGACACAAAGATGAT TGAATACCATGGGAGTGAGAATGTGGAGTCCTACATCCTCACCATCATGAACATGGTC ACTGGGTTGTTCCATAACCCAAGCATTGGCAATGCAATTCACATTGTTGTGGTTCGGC TCATTCTACTCGAAGAAGAAGAGCAAGGACTGAAAATAGTTCACCATGCAGAAAAGAC ACTGTCTAGCTTCTGCAAGTGGCAGAAGAGTATCAATCCCAAGAGTGACCTCAATCCT GTTCATCACGACGTGGCTGTCCTTCTCACCAGAAAGGACATCTGTGCTGGTTTCAATC GCCCCTGCGAGACCCTGGGCCTGTCTCACCTTTCAGGAATGTGTCAGCCTCACCGCAG TTGTAACATCAATGAAGATTCGGGACTCCCTCTGGCTTTCACAATTGCCCATGAGCTA GGACACAGCTTCGGCATCCAGCATGATGGGAAAGAAAATGACTGTGAGCCTGTGGGCA GACATCCGTACATCATGTCCCGCCAGCTCCAGTACGATCCCACTCCGCTGACATGGTC CAAGTGCAGCGAGGAGTACATCACCCGCTTCTTGGACCGAGGCTGGGGGTTCTGTCTT GATGACATACCTAAAAAGAAAGGCTTGAAGTCCAAGGTCATTGCCCCCGGAGTGATCT ATGATGTTCACCACCAGTGCCAGCTACAATATGGACCCAATGCTACCTTCTGCCAGGA AGTAGAAAACGTCTGCCAGACACTGTGGTGCTCCGTGAAGGGCTTTTGTCGCTCTAAG CTGGACGCTGCTGCAGATGGAACTCAATGTGGTGAGAAGAAGTGGTGTATGGCAGGCA AGTGCATCACAGTGGGGAAGAAACCAGAGAGCATTCCTGGAGGCTGGGGCCGCTGGTC ACCCTGGTCCCACTGTTCCAGGACCTGTGGGGCTGGAGTCCAGAGCGCAGAGAGGCTC TGCAACAACCCCGAGCCAAAGTTTGGAGGGAAATATTGCACTGGAGAAAGAAAACGCT ATCGCTTGTGCAACGTCCACCCCTGTCGCTCAGAGGCACCAACATTTCGGCAGATGCA GTGCAGTGAATTTGACACTGTTCCCTACAAGAATGAACTCTACCACTGGTTTCCCATT TTTAACCCAGCACATCCTTGTGAGCTCTACTGCCGACCCATAGATGGCCAGTTTTCTG AGAAAATGCTGGATGCTGTCATTGATGGTACCCCTTGCTTTGAAGGCGGCAACAGCAG AAATGTCTGTATTAATGGCATATGTAAGATGGTTGGCTGTGACTATGAGATCGATTCC AATGCCACCGAGGATCGCTGCGGTGTGTGCCTGGGAGATGGCTCTTCCTGCCAGACTG TGAGAAAGATGTTTAAGCAGAAGGAAGGATCTGGTTATGTTGACATTGGGCTCATTCC AAAAGGAGCAAGGGACATAAGAGTGATGGAAATTGAGGGAGCTGGAAACTTCCTGGCC ATCAGGAGTGAAGATCCTGAAAAATATTACCTGAATGGAGGGTTTATTATCCAGTGGA ACGGGAACTATAAGCTGGCAGGGACTGTCTTTCAGTATGACAGGAAAGGAGACCTGGA AAAGCTGATGGCCACAGGTCCCACCAATGAGTCTGTGTGGATCCAGCTTCTATTCCAG GTGACTAACCCTGGCATCAAGTATGAGTACACAATCCAGAAAGATGGCCTTGACAATG ATGTTGAGCAGATGTACTTCTGGCAGTACGGCCACTGGACAGAGTGCAGTGTGACCTG CGGGACAGGTATCCGCCGCCAAACTGCCCATTGCATAAAGAAGGGCCGCGGGATGGTG ϊTo AAAGCTACATTCTGTGACCCAGAAACACAGCCCAATGGGAGACAGAAGAAGTGCCATG AAAAGGCTTGTCCACCCAGGTGGTGGGCAGGGGAGTGGGAAGCATGCTCGGCGACATG CGGGCCCCACGGGGAGAAGAAGCGAACCGTGCTGTGCATCCAGACCATGGTCTCTGAC GAGCAGGCTCTCCCGCCCACAGACTGCCAGCACCTGCTGAAGCCCAAGACCCTCCTTT CCTGCAACAGAGACATCCTGTGCCCCTCGGACTGGACAGTGGGCAACTGGAGTGAGTG TTCTGTTTCCTGTGGTGGTGGAGTGCGGATTCGCAGTGTCACATGTGCCAAGAACCAT GATGAACCTTGCGATGTGACAAGGAAACCCAACAGCCGAGCTCTGTGTGGCCTCCAGC AATGCCCTTCTAGCCGGAGAGTTCTGAAACCAAACAAAGGCACTATTTCCAATGGAAA AAACCCACCAACACTAAAGCCCGTCCCTCCACCTACATCCAGGCCCAGAATGCTGACC ACACCCACAGGGCCTGAGTCTATGAGCACAAGCACTCCAGCAATCAGCAGCCCTAGTC CTACCACAGCCTCCAAAGAAGGAGACCTGGGTGGGAAACAGTGGCAAGATAGCTCAAC CCAACCTGAGCTGAGCTCTCGCTATCTCATTTCCACTGGAAGCACTTCCCAGCCCATC CTCACTTCCCAATCCTTGAGCATTCAGCCAAGTGAGGAAAATGTTTCCAGTTCAGATA CTGGTCCTACCTCGGAGGGAGGCCTTGTAGCTACAACAACAAGTGGTTCTGGCTTGTC ATCTTCCCGCAACCCTATCACTTGGCCTGTGACTCCATTTTACAATACCTTGACCAAA GGTCCAGAAATGGAGATTCACAGTGGCTCAGGGGAAGAAAGAGAACAGCCTGAGGACA AAGATGAAAGCAATCCTGTAATATGGACCAAGATCAGAGTACCTGGAAATGACGCTCC AGTGGAAAGTACAGAAATGCCACTTGCACCTCCACTAACACCAGATCTCAGCAGGGAG TCCTGGTGGCCACCCTTCAGCACAGTAATGGAAGGACTGCTCCCCAGCCAAAGGCCCA CTACTTCCGAAACTGGGACACCCAGAGTTGAGGGGATGGTTACTGAAAAGCCAGCCAA CACTCTGCTCCCTCTGGGAGGAGACCACCAGCCAGAACCCTCAGGAAAGACGGCAAAC CGTAACCACCTGAAACTTCCAAACAACATGAACCAAACAAAAAGTTCTGAACCAGTCC TGACTGAGGAGGATGCAACAAGTCTGATTACTGAGGGCTTTTTGCTAAATGCCTCCAA TTACAAGCAGCTCACAAACGGCCACGGCTCTGCACACTGGATCGTCGGAAACTGGAGC GAGTGCTCCACCACATGTGGCCTGGGGGCCTACTGGAAAAGGGTGGAGTGCACCACCC AGATGGATTCTGACTGTGCGGCCATCCAGAGACCTGACCCTGCAAAAAGATGCCACCT CCGTCCCTGTGCTGGCTGGAAAGTGGGAAACTGGAGCAAGTGCTCCAGAAACTGCAGT GGGGGCTTCAAGATACGCGAGATTCAGTGCGTGGACAGCCGGGACCACCGGAACCTGA GGCCATTTCACTGCCAGTTCCTGGCCGGCATTCCTCCCCCATTGAGCATGAGCTGTAA CCCGGAGCCCTGTGAGGCGTGGCAGGTGGAGCCTTGGAGCCAGTGCTCCAGGTCCTGT GGAGGTGGAGTTCAGGAGAGAGGAGTGTTCTGTCCAGGAGGCCTCTGTGATTGGACAA AAAGACCCACATCCACCATGTCTTGCAATGAGCACCTGTGCTGTCACTGGGCCACTGG GAACTGGGACCTGTGTTCCACTTCCTGTGGAGGTGGCTTTCAGAAGAGGATTGTCCAA TGTGTGCCCTCAGAGGGCAATAAAACTGAAGACCAAGACCAATGTCTATGTGATCACA AACCCAGACCTCCAGAATTCAAAAAATGCAACCAGCAGGCCTGCAAGAAAAGTGCCGA TTTACTTTGCACTAAGGACAAACTGTCAGCCAGTTTCTGCCAGACACTGAAAGCCATG AAGAAATGTTCTGTGCCCACCGTGAGGGCTGAGTGCTGCTTCTCGTGTCCCCAGACAC ACATCACACACACCCAAAGGCAAAGAAGGCAACGGTTGCTCCAAAAGTCAAAAGAACT CTAAGCCCAAA

ORF Start: ATG at 327 ORF Stop: TAA at 5106

SEQ ID NO: 14 1593 aa MW at 177543.9kD

NOV6a, MPCAQRS I-ANLSVVAQLIJNFGALCYGRQPQPGPVRFPDRRQEHFIKGLPEYHVVGPV CG58504-01 RVDASGHFLSYGLHYPITSSRRKRDLDGSEDWV YRISHEEKDLFFlsrLTVNQGFIjSNS Protein YIMEKRYG LS-T^ ^MASSAPLCHLSGTVLQQGTRVGTAALSACHGLTGFFQLPHGDF Sequence FIEPVIOOIPLVEGGY IPHIVYRRQIWPETKEPTCGLKDSVNISQKQELV EK ERH L PSRSLSRRSISKERWv/ETLVVADTKMIEY^GSENVESYILTIWttJMVTGLFHNPSIGNA IHIVVVRLILLEEEEQGLKIVHHAEKTLSSFCK QKSINPKSDLNPVHHTJVAVLL.TR DICAGFNRPCETLGLSHLSGMCQPHRSCNINEDSGLPLAFTIAHELGHSFGIQHDGKE NDCEPVGRHPYI SRQLQYDPTPLTWSKCSEEYITRFLDRGWGFCLDDIPKKKGLKSK VIAPGVIYDVHHQCQLQYGPNATFCQEVENVCQTL CSVKGFCRSKLDAAADGTQCGE KKWCMAGKCITVGKKPESIPGGWGRWSPWSHCSRTCGAGVQSAERLCN PEPKFGGKY CTGER-^YRLCNVHPCRSEAPTFRQMQCSEFDTVPYKNELYHWFPIFNPAHPCELYCR PIDGQFSEKrøΛAVIDGTPCFEGGNSRNVCINGICKMVGCDYΞIDSNATEDRCGVCLG DGSSCQTVRKMFKQKEGSGYVDIGLIPKGARDIRVMEIEGAGNFLAIRSEDPΞKYYLN GGFIIQWNGNYIOiAGTVFQYDRKGDLEKLMATGPTNESVWIQLLFQVTNPGIKYEYTI QKDGLDlsrDVEQ YFWQYGHWTECSVTCGTGIRRQTAHCII GRG VKATFCDPETQPN GRQKKCHEKACPPR WAGEWEACS ATCGPHGEKKRTVLC I QTMVSDEQALPPTDCQHL LKPKTLLSCMvDILCPSDWT 7GNWSECSVSCGGGVRIRSVTCAKNHDEPCDVTRKPNS

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 6B.

Table 6B. Comparison of NOV6a against NOV6b and NOV6c.

NOV6a Residues/ Identities/

Protein Sequence Match Residues Similarities for the Matched Region

NOV6b 476..555 80/80 (100%) 3..82 80/80 (100%.)

NOV6c 476..555 79/80 (98%) 3..82 79/80 (98%)

Further analysis of the NOV6a protein yielded the following properties shown in Table 6C.

Table 6C. Protein Sequence Properties NOV6a

PSort analysis: 0.5087 probability located in outside; 0.1900 probability located in lysosome (lumen); 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen)

SignalP analysis: Cleavage site between residues 26 and 27

A search of the NOV6a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 6D.

In a BLAST search of public sequence databases, the NOV6a protein was found to have homology to the proteins shown in the BLASTP data in Table 6E.

PFam analysis predicts that the NOV6a protein contains the domains shown in the Table 6F.

EXAMPLE 7.

The NOV7 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 7A.

Table 7A. NOV7 Sequence Analysis

SEQ ID NO: 19 4324 bp

NOV7a, GCCCAATAAATGTGAACATGGGATCTGTCACGGGAGCTGTCCTCAAGACGCTACTTCT CG58586-01 GTTATCTACTCAAAATTGGAACAGAGTCGAAGCTGGGAATTCCTATGACTGTGATGAT DNA Sequence CCTCTTGTGTCTGCCTTGCCTCAGGCATCCTTCAGCAGTTCTTCCGAGCTCTCCAGCA GTCATGGTCCTGGATTTGCAAGGCTGAATAGAAGAGATGGAGCTGGTGGCTGGTCTCC ACTTGTGTCTAACAAATACCAGTGGTTGCAGATTGACCTTGGAGAGAGAATGGAGGTC ACCGCTGTGGCCACTCAAGGGGGATATGGTAGCTCCAACTGGGTGACCAGCTACCTCC TGATGTTCAGTGATAGTGGCTGGAACTGGAAACAATATCGCCAAGAGGACAGCATCTG GGGTTTTTCAGGAAATGCAAATGCAGACAGTGTTGTGTACTATAGACTCCAGCCTTCT ATCAAAGCCAGATTTCTGCGCTTCATCCCTTTGGAATGGAACCCCAAGGGCAGAATTG GAATGCGAATCGAAGTGTTCGGATGTGCATACAGATCAGAAGTGGTTGATCTTGATGG AAAAAGTTCCCTTCTCTACAGATTTGATCAAAAATCCCTGAGCCCAATAAAAGACATT ATTTCTTTGAAATTCAAAACCATGCAGAGTGATGGGATTCTACTCCACAGGGAAGGGC CAAATGGAGATCACATCACACTGCAATTAAGAAGAGCAAGACTCTTTTTACTTATTAA TTCAGGTGAAGCTAAACTGCCTTCCACTTCCACCCTGGTCAATCTCACCCTGGGCAGC CTGCTAGATGATCAGCATTGGCATTCAGTGCTCATCCAGCGTTTGGGCAAACAAGTCA ACTTCACAGTGGACGAACACAGGCATCATTTCCATGCACGGGGAGAATTCAATCTCAT GAATCTTGΆTTATGAGATCAGCTTTGGAGGGATTCCAGCACCTGGAAAATCAGTGTCA TTCCCACATAGAAATTTTCATGGATGTTTAGAAAATCTCTATTATAATGGAGTGGATA TCATTGATTTGGCCAAGCAGCAAAAACCACAGATCATTGCTATGGGAAATGTGTCATT TTCTTGTTCACAACCACAATCTATGCCCGTGACTTTTCTGAGCTCCAGGAGTTATTTA GCACTGCCAGACTTCTCTGGAGAGGAGGAGGTTTCTGCCACTTTTCAATTTCGAACTT GGAATAAGGCAGGGCTTCTGCTGTTCAGTGAACTTCAGCTGATTTCAGGGGGTATCCT CCTCTTTCTGAGTGATGGAAAACTTAAGTCGAATCTCTACCAGCCAGGAAAATTACCC AGTGACATCACAGCAGGTGTCGAATTAAATGATGGGCAGTGGCATTCTGTCTCTTTAT CTGCTAAAAAGAATCACTTGAGTGTGGCGGTGGACGGCCAGATGGCTTCTGCTGCTCC TCTGCTGGGGCCTGAGCAGATTTATTCGGGTGGCACCTATTATTTTGGAGGTTGTCCT GACAAAAGCTTTGGATCCAAΆTGTAAAAGTCCACTTGGTGGATTTCAGGGATGTATGA GGCTCATTTCTATCAGCGGCAAAGTGGTAGATCTGATTTCAGTTCAGCAGGGGTCCCT TGGGAACTTCAGTGACCTTCAGATAGACTCATGTGGCATCTCAGACAGGTGTTTGCCC AACTATTGTGAACACGGTGGGGAGTGTTCCCAGTCCTGGAGCACCTTTCATTGTAACT GTACCAACACTGGTTACAGAGGAGCTACTTGCCATAACTCTATCTATGAGCAGTCATG TGAAGCCTATAAGCACAGAGGAAATACTTCAGGGTTTTACTATATAGATTCAGATGGA AGTGGTCCCCTGGAACCATTTCTTCTATATTGCAATATGACCGAAACTGCATGGACCA TCATACAGCACAACGGCTCTGACTTAACAAGAGTCAGAAATACTAATCCAGAGAACCC ATATGCTGGGTTTTTCGAGTATGTGGCCAGCATGGAGCAACTTCAGGCCACTATTAAC CGTGCAGAGCACTGTGAACAGGAGTTTACTTATTACTGCAAGAAGTCACGGCTGGTCA ATAAGCAAGATGGAACCCCTCTGAGTTGGTGGGTAGGAAGAACCAATGAAACGCAAAC CTACTGGGGAGGTTCTTCGCCTGATCTTCAAAAATGTACTTGTGGATTAGAGGGAAAC GCATTGATTCTCAGTATTACTGCAATTGTGATGCTGACCGGAATGAATGGACCAATG ACACTGGATTGCTTGCTTATAAAGAACATCTTCCAGTAACTAAGATCGTGATTACAGA CACAGGCCGACTGCATTCAGAAGCAGCTTATAAACTGGGGCCTCTGCTCTGCCGGGGA GACAGATCATTTTGGAATTCAGCTTCCTTTGATACCGAGGCTTCATATCTTCATTTTC CTACCTTCCACGGAGAACTTAGCGCGGATGTATCTTTCTTTTTTAAGACAACAGCTTC ATCTGGGGTATTTTTAGAGAACTTGGGGATTGCTGATTTTATACGGATAGAGCTTCGC TCTCCGΆCAGTAGTGACTTTTTCATTTGATGTGGGGAATGGGCCTTTTGAAATCTCAG TGCAGTCACCCACCCACTTCAACGACAACCAGTGGCACCATGTGAGGGTTGAAAGGAA CATGAAGGAGGCCTCCCTTCAAGTGGATCAGCTGACACCAAAGACACAGCCCGCCCCC GCTGATGGGCATGTCCTGTTACAGCTCAACAGTCAGCTCTTCGTGGGTGGAACGGCCA CCAGACAGAGAGGCTTTCTGGGCTGCATTCGGTCTCTGCAGTTGAATGGGATGACCCT GGATTTGGAAGAAAGAGCCCAGGTGACTCCAGAAGTGCAGCCAGGTTGTAGGGGACAT GCAGCAGCTATGGGAAGTTATGCCGCAATGGAGGGAAATGCAGAGAAAGACCCATTG GGTTCTTTTGTGACTGCACTTTCTCTGCATACACAGGGCCATTCTGCTCAAATGAGAT TTCTGCATATTTTGGATCTGGCTCATCCGTGATATACAATTTTCAAGAAAATTATCTT TTAAGTAAAAACTCCAGCTCCCACGCTGCTTCATTTCATGGTGATATGAAGCTGAGCA GAGAΆATGATCAAATTTAGTTTCCGAACAACACGAACACCAAGCTTGCTGCTTTTTGT GAGCTCCTTTTACAAAGAATACCTTTCTGTGATCATTGCCAAAAATGGAAGTTTGCAG ATCAGGTACAAGTTAAATAAATATCAAGAGCCTGATGTTGTTAACTTTGATTTTAAAA ACATGGCTGATGGACAACTTCACCACATAATGATTAACAGAGAAGAAGGAGTGGTCTT TATAGAGATTGACGATAATAGAAGGAGACAAGTTCACCTGTCATCAGGCACAGAATTC AGTGCAGTCAAATCTCTGGTATTGGGCAGGATTTTAGAACACAGTGATGTGGACCAGG AGACTGCACTGGCAGGTGCGCAGGGCTTCACAGGCTGCCTCTCTGCAGTGCAGCTCAG CCACGTGGCCCCTCTGAAGGCAGCTCTGCACCCCAGCCACCCAGACCCTGTCACTGTT ACAGGACACGTGACCGAGTCCAGCTGTATGGCCCAGCCTGGCACTGATGCCACATCAA GGGAAAGGACACACTCGTTTGCAGATCATTCTGGAACAATAGATGACAGAGAGCCCCT TGCTAATGCAATCAAAAGTGACTCTGCAGTAATTGGAGGTCTGATAGCTGTTGTGATT TTTATCTTGCTTTGCATCACTGCCATAGCTGTTCGCATTTATCAGCAGAAAAGGTTAT TAAAAGAAGTGAGGCAAAAAGGTCAGAGAATGTAGACAGTGCTGAGGCTGTTCTGAA AAGTGAGCTTAATATACAAAATGCAGTCAATGAAAATCAGAAAGAGTACTTCTTCTGA TTGGCAGCTATGATTTAACATAAAATTATGATAGTTTGTTTTAATAGCCAGGGGTTCT

CAATGGAAAAACGAATGCTCTTACACTGAATGTACAGGCAGTGGGCTTGCAGCACTGC

CATCTTGCCATGTACAGGCTTGGGGTGGCTCCAGGAAGCCTCGTCCAGTGATATATTT

CTCATAGCATTCATTCTATGGAACAAGAAATTAGATATTGCTGTTAATTTTCAACTGT

TCTGGTATGATCTAAAACAAGTTTAACCTGCTTAATGGCTACAGTTTTTACATGTGAA

AACTGTAGCCTTGGTCTCTTAACCATGTAATACATAAGTTTTGTTAGAGGTAAAAATT

AAATTTGGACTATAATGTCCTTGCTTTATTTG

ORF Start: ATG at 18 ORF Stop: TGA at 3942

SEQ ID NO: 20 1308 aa MW at 145314.9kD

NOV7a, MGSVTGAVL TLLLLSTQNWNRVEAGNSYDCDDPLVSALPQASFSSSSELSSSHGPGF CG58586-01 ARLNRRDGAGG S PLVSNKYQWLQIDLGERMΞVTAVATQGGYGS SN VTS YLLMFSDS Protein G KQYRQEDSI GFSGNANADSWYYRLQPSIKARFL.RFIPLEWNPKGRIGMRIEV

FGCAYRSEWDLDGKSSLLYRFDQKSLSPIKDIISLKFKT QSDGILLHREGPNGDHI Sequence

TLQLRRARLFLLINSGEAKLPSTSTLVNLTLGSLLDDQHWHSVLIQRLGKQVNFTVDE

HRHHFHARGEFNLMNLD E I S FGGI PAPGKS VS FPHRNFHGCLENLYY GVD I IDLAK

QQKPQIIAMGNVSFSCSQPQSMPVTFLSSRSYLALPDFSGEEEVSATFQFRT NKAGL

LLFSELQLISGGILLFLSDGKLKSNLYQPGKLPSDITAGVEI-lTOGQVrøSVSLSAKKNH

LS VAVDGQMAS APLLGPEQI YS GGTYYFGGCPDKS FGSKCKS PLGGFQGCMRLI SIS

GKWDLISVQQGSLGNFSDLQIDSCGISDRCLPNYCEHGGECSQS STFHCNCTNTGY

RGATCHNS I YEQSCEAY HRGNTSGFYYIDSDGSGPLEPFLLYCNMTETA TI IQHNG

SDLT -TI^ ENPYAGFFE ^ASMEQLQATIN AEHCEQEFT CKKSRLV KQDGT

PLS VWGRTNETQTY^GGSSPDLQKCTCGLEGNCIDSQY^XNCDAϋl^^ TlTOTGLI-A

YKEHLPVTKIVITDTGRLHSEAAYKLGPLI.CRGDRSF SASFDTEASYLHFPTFHGE

LSADVSFFFKTTASSGVFLENLGIADFIRIELRSPTVVTFSFDVGNGPFEISVQSPTH

FlTONQVraHVRVERNMKEASLQVDQLTPKTQPAPADGHVLLQLNSQLFVGGTATRQRGF

LGCIRSLQIiNG TLDLEERAQVTPEVQPGCRGHCSSYGKLCRNGGKCRERPIGFFCDC

TFSAYTGPFCSNEISAYFGSGSSVIYl^FQElsrYLLS NSSSHAASFHGDMKLSREMIKF

SFRTTRTPSLLLFVSSFY EYLSVIIAI<^GSLQIRYKLlmYQEPDVVNFDFKNiy^

L-ffll IlsmEEGVVFIEIDDlN P QVHLSSGTEFSAVKSLVLGRILEHSDVDQETALAG

AQGFTGCLSAVQLSIWAPLI^ALHPSHPDPVTVTGHVTESSCMAQPGTDATSRERTHS

FADHSGTIDDREPLANAIKSDSAVIGGLIAWIFILLCITAIAVRIYQQKRLYKRSEA lOiSElvrVDSAEAVLKSEL IQNAVNENQKEYFF

SEQ ID NO: 21 4331 bp

NOV7b, GCCCAATAAATGTGAACATGGGATCTGTCACGGGAGCTGTCCTCAAGACGCTACTTCT CG58586-02 GTTATCTACTCAAAATTGGAACAGAGTCGAAGCTGGGAATTCCTATGACTGTGATGAT DNA Sequence CCTCTTGTGTCTGCCTTGCCTCAGGCATCCTTCAGCAGTTCTTCCGAGCTCTCCAGCA GTCATGGTCCTGGATTTGCAAGGCTGAATAGAAGAGATGGAGCTGGTGGCTGGTCTCC ACTTGTGTCTAACAAATACCAGTGGTTGCAGATTGACCTTGGAGAGAGAATGGAGGTC ACCGCTGTGGCCACTCAAGGGGGATATGGTAGCTCCAACTGGGTGACCAGCTACCTCC TGATGTTCAGTGATAGTGGCTGGAACTGGAAACAATATCGCCAAGAGGACAGCATCTG GGGTTTTTCAGGAAATGCAAATGCAGACAGTGTTGTGTACTATAGACTCCAGCCTTCT ATCAAAGCCAGATTTCTGCGCTTCATCCCTTTGGAATGGAACCCCAAGGGCAGAATTG GAATGCGAATCGAAGTGTTCGGATGTGCATACAGATCAGAAGTGGTTGATCTTGATGG AAAAAGTTCCCTTCTCTACAGATTTGATCAAAAATCCCTGAGCCCAATAAAAGACATT ATTTCTTTGAAATTCAAAACCATGCAGAGTGATGGGATTCTACTCCACAGGGAAGGGC CAAATGGAGATCACATCACACTGCAATTAAGAAGAGCAAGACTCTTTTTACTTATTAA TTCAGGTGAAGCTAAACTGCCTTCCACTTCCACCCTGGTCAATCTCACCCTGGGCAGC CTGCTAGATGATCAGCATTGGCATTCAGTGCTCATCCAGCGTTTGGGCAAACAAGTCA ACTTCACAGTGGACGAACACAGGCATCATTTCCATGCACGGGGAGAATTCAATCTCAT GAATCTTGATTATGAGATCAGCTTTGGAGGGATTCCAGCACCTGGAAAATCAGTGTCA TTCCCACATAGAAATTTTCATGGATGTTTAGAAAATCTCTATTATAATGGAGTGGATA TCATTGATTTGGCCAAGCAGCAAAAACCACAGATCATTGCTATGGGAAATGTGTCATT TTCTTGTTCACAACCACAATCTATGCCCGTGACTTTTCTGAGCTCCAGGAGTTATTTA GCACTGCCAGACTTCTCTGGAGAGGAGGAGGTTTCTGCCACTTTTCAATTTCGAACTT GGAATAAGGCAGGGCTTCTGCTGTTCAGTGAACTTCAGCTGATTTCAGGGGGTATCCT CCTCTTTCTGAGTGATGGAAAACTTAAGTCGAATCTCTACCAGCCAGGAAAATTACCC AGTGACATCACAGCAGGTGTCGAATTAAATGATGGGCAGTGGCATTCTGTCTCTTTAT CTGCTAAAAAGAATCACTTGAGTGTGGCGGTGGACGGCCAGATGGCTTCTGCTGCTCC TCTGCTGGGGCCTGAGCAGATTTATTCGGGTGGCACCTATTATTTTGGAGGTTGTCCT GACAAAAGCTTTGGATCCAAATGTAAAAGTCCACTTGGTGGATTTCAGGGATGTATGA GGCTCATTTCTATCAGCGGCAAAGTGGTAGATCTGATTTCAGTTCAGCAGGGGTCCCT TGGGAACTTCAGTGACCTTCAGATAGACTCATGTGGCATCTCAGACAGGTGTTTGCCC AACTATTGTGAACACGGTGGGGAGTGTTCCCAGTCCTGGAGCACCTTTCATTGTAACT GTACCAACACTGGTTACAGAGGAGCTACTTGCCATAACGCTATCTATGAGCAGTCATG TGAAGCCTATAAGCACAGAGGAAATACTTCAGGGTTTTACTATATAGATTCAGATGGA AGTGGTCCCCTGGAACCATTTCTTCTATATTGCAATATGACCCAAGAAACTGCATGGA CCATCATACAGCACAACGGCTCTGACTTAACAAGAGTCAGAAATACTAATCCAGAGAA CCCATATGCTGGGTTTTTCGAGTATGTGGCCAGCATGGAGCAACTTCAGGCCACTATT AACCGTGCAGAGCACTGTGAACAGGAGTTTACTTATTACTGCAAGAAGTCACGGCTGG TCAATAAGCAAGATGGAACCCCTCTGAGTTGGTGGGTAGGAAGAACCAATGAAACGCA AACCTACTGGGGAGGTTCTTCGCCTGATCTTCAAAAATGTACTTGTGGATTAGAGGGA AACTGCATTGATTCTCAGTATTACTGCAATTGTGATGCTGACCGGAATGAATGGACCA ATGACACTGGATTGCTTGCTTATAAAGAACATCTTCCAGTAACTAAGATCGTGATTAC AGACACAGGCCGACTGCATTCAGAAGCAGCTTATAAACTGGGGCCTCTGCTCTGCCGG GGAGACAGTAAGTGGTCATTTTGGAATTCAGCTTCCTTTGATACCGAGGCTTCATATC TTCATTTTCCTACCTTCCACGGAGAACTTAGCGCGGATGTATCTTTCTTTTTTAAGAC AACAGCTTCATCTGGGGTATTTTTAGAGAACTTGGGGATTGCTGATTTTATACGGATA GAGCTTCGCACAGTAGTGACTTTTTCATTTGATGTGGGGAATGGGCCTTTTGAAATCT CAGTGCAGTCACCCACCCACTTCAACGACAACCAGTGGCACCATGTGAGGGTTGAAAG GAACATGAAGGAGGCCTCCCTTCAAGTGGATCAGCTGACACCAAAG CAΓAGCCCGCC CCCGCTGATGGGCATGTCCTGTTACAGCTCAACAGTCAGCTCTTCGTGGGTGGAACGG CCACCAGACAGAGAGGCTTTCTGGGCTGCATTCGGTCTCTGCAGTTGAATGGGATGAC CCTGGATTTGGAAGAAAGAGCCCAGGTGACTCCAGAAGTGCAGCCAGGTTGTAGGGGA CATTGCAGCAGCTATGGGAAGTTATGCCGCAATGGAGGGAAATGCAGAGAAAGACCCA TTGGGTTCTTTTGTGACTGCACTTTCTCTGCATACACAGGGCCATTCTGCTCAAATGA GATTTCTGCATATTTTGGATCTGGCTCATCCGTGATATACAATTTTCAAGAAAATTAT CTTTTAAGTAAAAACTCCAGCTCCCACGCTGCTTCATTTCATGGTGATATGAAGCTGA GCAGAGAAATGATCAAATTTAGTTTCCGAACAACACGAACACCAAGCTTGCTGCTTTT TGTGAGCTCCTTTTACAAAGAATACCTTTCTGTGATCATTGCCAAAAATGGAAGTTTG CAGATCAGGTACAAGTTAAATAAATATCAAGAGCCTGATGTTGTTAACTTTGATTTTA AAAACATGGCTGATGGACAACTTCACCACATAATGATTAACAGAGAAGAAGGAGTGGT CTTTATAGAGATTGACGATAATAGAAGGAGACAAGTTCACCTGTCATCAGGCACAGAA TTCAGTGCAGTCAAATCTCTGGTATTGGGCAGGATTTTAGAACACAGTGATGTGGACC AGGAGACTGCACTGGCAGGTGCGCAGGGCTTCACAGGCTGCCTCTCTGCAGTGCAGCT CAGCCACGTGGCCCCTCTGAAGGCAGCTCTGCACCCCAGCCACCCAGACCCTGTCACT GTTACAGGACACGTGACCGAGTCCAGCTGTATGGCCCAGCCTGGCACTGATGCCACAT CAAGGGAAAGGACACACTCGTTTGCAGATCATTCTGGAACAATAGATGACAGAGAGCC CCTTGCTAATGCAATCAAAAGTGACTCTGCAGTAATTGGAGGTCTGATAGCTGTTGTG ATTTTTATCTTGCTTTGCATCACTGCCATAGCTGTTCGCATTTATCAGCAGAAAAGGT TATATAAAAGAAGTGAGGCAAAAAGGTCAGAGAATGTAGACAGTGCTGAGGCTGTTCT GAAAAGTGAGCTTAATATACAAAATGCAGTCAATGAAAATCAGAAAGAGTACTTCTTC TGATTGGCAGCTATGATTTAACATAAAATTATGATAGTTTGTTTTAATAGCCAGGGGT TCTCAATGGAAAAACGAATGCTCTTACACTGAATGTACAGGCAGTGGGCTTGCAGCAC

TGCCATCTTGCCATGTACAGGCTTGGGGTGGCTCCAGGAAGCCTCGTCCAGTGATATA

TTTCTCATAGCATTCATTCTATGGAACAAGAAATTAGATATTGCTGTTAATTTTCAAC

TGTTCTGGTATGATCTAAAACAAGTTTAACCTGCTTAATGGCTACAGTTTTTACATGT

GAAAACTGTAGCCTTGGTCTCTTAACCATGTAATACATAAGTTTTGTTAGAGGTAAAA

ATTAAATTTGGACTATAATGTCCTTGCTTTATTTGNNNN

ORF Start: ATG at 18 ORF Stop: TGA at 3945

SEQ ID NO: 22 1309 aa MW at 145488.1kD

NOV7b, MG S TGAVLKTLLLLSTQNWNRVEAGNS YDCDDPLVS ALPQASFS S SSELS S SHGPGF CG58586-02 ARI-lv RDGAGGWSPLVSNKYQWLQIDLGERMEVTAVATQGGYGSSNVr/TSYLIjMFSDS Protein GWNWKQYRQEDS I WGFSGNANADS WYYRLQPS IKARFLRFI PLE NPKGRIGMRIEV Sequence FGCAYRSEWDLDGKSSLLYRFDQKSLSPIKDIISLKFKTMQSDGILLHREGPNGDHI

TLQLRRARLFLLINSGEAKLPSTSTLVl^TLGSLLDDQH HSVLIQRLGKQVNFTVDE

HRHHFHARGEF-^i jDYEISFGGIPAPGKSVSFPHrøFHGCLENLY ^^

QQIv^QIIAMGNVSFSCSQPQSMPVTFLSSRSYXjALPDFSGEEEVSATFQFRTWN AGL

LLFSELQLISGGILLFLSDGKLKSlvTLYQPGIXPSDITAGVELNDGQWHSVSLSAKKNH

LSVAVDGQMASAAPLLGPEQIYSGGTYYFGGCPDKSFGSKCKSPLGGFQGCMRLISIS

GKVVDLISVQQGSLGNFSDLQIDSCGISDRCLPNYCEHGGECSQSWSTFHCNCTNTGY

RGATCHNAIYEQSCEAYKHRGNTSGFYYIDSDGSGPLEPFLLYC i TQETA TIIQHN

GSDLTRVRNTNPENPYAGFFEYΛ^AS EQLQATINRAEHCΞQEFTYYCKKSRLVNKQDG

TPLS VGRTNETQT^WGGSSPDLQKCTCGLEGNCIDSQYYCNCDADRNEW'TNDTGL.I.

AYKEHLPVTKIVITDTGRLHSEAAYKLGPLLCRGDSK SF NSASFDTEASYLHFPTF

HGELSADVSFFFKTTASSGVFLENLGIADFIRIELRTWTFSFDVGNGPFEISVQSPT

HFNDNQ HHVRVERNMKEASLQVDQLTPKTQPAPADGHVLLQLNSQLFVGGTATRQRG

FLGCIRSLQLNGMTLDLEERAQVTPEVQPGCRGHCSSYGKLCRNGGKCRERPIGFFCD

CTFSAYTGPFCSNEISAYFGSGSSVIYNFQENYL.LSKNSSSHAASFHGDM LSREMIK

FSFRTTRTPSLLLFVSSFY EYLSVIIAKNGSLQIRYKLNKYQEPDVV FDFKNMADG

QLHHIMINREEGWFIEIDDNRRRQVHLSSGTEFSAVKSLVLGRILEHSDVDQETALA

GAQGFTGCLSAVQLSHVAPLIVT JHPSHPDPVTVTGHVTESSCMAQPGTDATSRERTH

SFADHSGTIDDREPLIANAIKSDSAVIGGLIAVVIFILLCITAIAVRIYQQKRLYKRSE i AKRSENVDSAEAVL , KSELNIQ ZNAVNENQ 2KEYFF

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 7B.

Further analysis of the NOV7a protein yielded the following properties shown in Table 7C.

Table 7C. Protein Sequence Properties NOV7a

PSort analysis: 0.8343 probability located in mitochondrial inner membrane; 0.6400 probability located in plasma membrane; 0.4000 probability located in Golgi body; 0.3000 probability located in endoplasmic reticulum (membrane)

SignalP analysis: Cleavage site between residues 26 and 27

A search of the NOV7a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 7D.

In a BLAST search of public sequence databases, the NOV7a protein was found to have homology to the proteins shown in the BLASTP data in Table 7E.

PFam analysis predicts that the NOV7a protein contains the domains shown in the Table 7F.

EXAMPLE 8.

The NOV8 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 8A.

Table 8A. NOV8 Sequence Analysis

SEQ ID NO: 23 5878 bp

NOV8a, TCTTAAAGAAACTTATTTTGGGCGGGGGGGGTGGGTTTGCTCTGGGCATTTGCTTTGC CG93453-01 CCAGTAGTTGGAAAGTGAACTCGACTCGTGATGGTTCTCCTGTCACTTTGGTTGATAG DNA Sequence CAGCCGCTCTGGTAGAGGTTAGGACTTCAGCTGATGGACAAGCTGGTAATGAAGAAAT GGTGCAAATAGATTTACCAATAAAGAGATATAGAGAGTATGAGCTGGTGACTCCAGTC AGCACAAATCTAGAAGGACGCTATCTCTCCCATACTCTTTCTGCGAGTCACAAAAAGA GGTCAGCGAGGGACGTGTCTTCCAACCCTGAGCAGTTGTTCTTTAACATCACGGCATT TGGAAAAGATTTTCATCTGCGACTAAAGCCCAACACTCAACTAGTAGCTCCTGGGGCT GTTGTGGAGTGGCATGAGACATCTCTGGTGCCTGGGAATATAACCGATCCCATTAACA ACCATCAACCAGGAAGTGCTACGTATAGAATCCGGAGAACAGAGCCTTTGCAGACTAA CTGTGCTTATGTTGGTGACATCGTGGACATTCCAGGAACCTCTGTTGCCATCAGCAAC TGTGATGGTCTGGCTGGAATGATAAAAAGTGATAATGAAGAGTATTTCATTGAACCCT GGAAAGAGGTAAACAGATGGAGGAAGAAAAAGGAAGGATTCATGTTGTCTACAAGAG ATCAGCTGTAGAACAGGCTCCCATAGACATGTCCAAAGACTTCCACTACAGAGAGTCG GACCTGGAAGGCCTTGATGATCTAGGTACTGTTTATGGCAACATCCACCAGCAGCTGA ATGAAACAATGAGACGCCGCAGACACGCGGGAGAAAACGATTACAATATCGAGGTACT GCTGGGAGTGGATGACTCTGTGGTCCGTTTCCATGGCAAAGAGCACGTCCAAAACTAC CTCCTGACCCTAATGAACATTGTGAATGAAATTTACCATGATGAGTCCCTCGGAGTGC ATATAAATGTGGTCCTGGTGCGCATGATAATGCTGGGATATGCAAAGTCCATCAGCCT CATAGAAAGGGGAAACCCATCCAGAAGCTTGGAGAATGTGTGTCGCTGGGCGTCCCAA CAGCAAAGATCTGATCTCAACCACTCTGAACACCATGACCATGCAATTTTTTTAACCA GGCAAGACTTTGGACCTGCTGGAATGCAAGGATATGCTCCAGTCACCGGCATGTGTCA TCCAGTGAGAAGTTGTACCCTGAATCATGAGGATGGTTTTTCATCTGCTTTTGTAGTA GCCCATGAAACGGGCCATGTGTTGGGAATGGAGCATGATGGACAAGGCAACAGGTGTG GTGATGAGACTGCTATGGGAAGTGTCATGGCTCCCTTGGTACAAGCAGCATTCCATCG TTACCACTGGTCCCGATGCAGTGGTCAAGAACTGAAAAGATATATCCATTCCTATGAC TGTCTCCTTGATGACCCTTTTGATCATGATTGGCCTAAACTCCCAGAACTTCCTGGAA TCAATTATTCTATGGATGAGCAATGTCGTTTTGATTTTGGTGTTGGCTATAAAATGTG CACCGCGTTCCGAACCTTTGACCCATGTAAACAGCTGTGGTGTAGCCATCCTGATAAT CCCTACTTTTGTAAGACTAAAAAGGGACCTCCACTTGATGGGACTGAATGTGCTGCTG GAAAATGGTGCTATAAGGGTCATTGCATGTGGAAGAATGCTAATCAGCAAAAACAAGA TGGCAATTGGGGGTCATGGACTAAATTTGGCTCCTGTTCTCGGACATGTGGAACTGGT GTTCGTTTCAGAACACGCCAGTGCAATAATCCCATGCCCATCAATGGTGGTCAGGATT GTCCTGGTGTTAATTTTGAGTACCAGCTTTGTAACACAGAAGAATGCCAAAAACACTT TGAGGACTTCAGAGCACAGCAGTGTCAGCAGCGAAACTCCCACTTTGAATACCAGAAT ACCAAACACCACTGGTTGCCATATGAACATCCTGACCCCAAGAAAAGATGCCACCTTT ACTGTCAGTCCAAGGAGACTGGAGATGTTGCTTACATGAAACAACTGGTGCATGATGG AACGCACTGTTCTTACAAAGATCCATATAGCATATGTGTGCGAGGAGAGTGTGTGAAA GTGGGCTGTGATAAAGAAATTGGTTCTAATAAGGTTGAGGATAAGTGTGGTGTCTGTG GAGGAGATAATTCCCACTGCCGAACCGTGAAGGGGACATTTACCAGAACTCCCAGGAA GCTTGGGTACCTTAAGATGTTTGATATACCCCCTGGGGCTAGACATGTGTTAATCCAA GAAGACGAGGCTTCTCCTCATATTCTTGCTATTAAGAACCAGGCTACAGGCCATTATA TTTTAAATGGCAAAGGGGAGGAAGCCAAGTCGCGGACCTTCATAGATCTTGGTGTGGA GTGGGATTATAACATTGAAGATGACATTGAAAGTCTTCACACCGATGGACCTTTACAT GATCCTGTTATTGTTTTGATTATACCTCAAGAAAATGATACCCGCTCTAGCCTGACAT ATAAGTACATCATCCATGAAGACTCTGTACCTACAATCAACAGCAACAATGTCATCCA GGAAGAATTAGATACTTTTGAGTGGGCTTTGAAGAGCTGGTCTCAGTGTTCCAAACCC TGTGGTGGAGGTTTCCAGTACACTAAATATGGATGCCGTAGGAAAAGTGATAATAAAA TGGTCCATCGCAGCTTCTGTGAGGCCAACAAAAAGCCGAAACCTATTAGACGAATGTG CAATATTCAAGAGTGTACACATCCACTCTGGGTAGCAGAAGAATGGGAACACTGCACC AAAACCTGTGGAAGTTCTGGCTATCAGCTTCGCACTGTACGCTGCCTTCAGCCACTCC TTGATGGCACCAACCGCTCTGTGCACAGCAAATACTGCATGGGTGACCGTCCCGAGAG CCGCCGGCCCTGTAACAGAGTGCCCTGCCCTGCACAGTGGAAAACAGGACCCTGGAGT GAGTGTTCAGTGACCTGCGGTGAAGGAACGGAGGTGAGGCAGGTCCTCTGCAGGGCTG GGGACCACTGTGATGGTGAAAAGCCTGAGTCGGTCAGAGCCTGTCAACTGCCTCCTTG TAATGATGAACCATGTTTGGGAGACAAGTCCATATTCTGTCAAATGGAAGTGTTGGCA CGATACTGCTCCATACCAGGTTATAACAAGTTATGTTGTGAGTCCTGCAGCAAGCGCA GTAGCACCCTGCCACCACCATACCTTCTAGAAGCTGCTGAAACTCATGATGATGTCAT CTCTAACCCTAGTGACCTCCCTAGATCTCTAGTGATGCCTACATCTTTGGTTCCTTAT CATTCAGAGACCCCTGCAAAGAAGATGTCTTTGAGTAGCATCTCTTCAGTGGGAGGTC CAAATGCATATGCTGCTTTCAGGCCAAACAGTAAACCTGATGGTGCTAATTTACGCCA GAGGAGTGCTCAGCAAGCAGGAAGTAAGACTGTGAGACTGGTCACCGTACCATCCTCC CCACCCACCAAGAGGGTCCACCTCAGTTCAGCTTCACAAATGGCTGCTGCTTCCTTCT TTGCAGCCAGTGATTCAATAGGTGCTTCTTCTCAGGCAAGAACCTCAAAGAAAGATGG AAAGATCATTGACAΆCAGACGTCCGACAAGATCATCCACCTTAGAAAGATGAGAAAGT GAACCAAAAAGGCTAGAAACCAGAGGAAAACCTGGACAACCTCTCTCTTCCCATGGTG

CATATGCTTGTTTAAAGTGGAAATCTCTATAGATCGTCAGCTCATTTTATCTGTAATT

GGAAGAACAGAAAGTGCTGGCTCACTTTCTAGTTGCTTTCATCCTCCTTTTGTTCTGC

ATTGACTCATTTACCAGAATTCATTGGAAGAAATCACCAAAGATTATTACAAAAGAAA

AATATGTTGCTAAGATTGTGTTGGTCGCTCTCTGAAGCAGAAAAGGGACTGGAACCAA

TTGTGCATATCAGCTGACTTTTTGTTTGTTTTAGAAAAGTTACAGTAAAAATTAAAAA

GAGATACCAATGGTTTACACTTTAACAAGAAATTTTGGATATGGAACAAAGAATTCTT

AGACTTGTATTCCTATTTATCTATATTAGAAATATTGTATGAGCAAATTTGCAGCTGT

TGTGTAAATACTGTATATTGCAAAAATCAGTATTATTTTAAGAGATGTGTTCTCAAAT

GATTGTTTACTATATTACATTTCTGGATGTTCTAGGTGCCTGTCGTTGAGTATTGCCT

TGTTTGACATTCTATAGGTTAATTTTCAAAGCAGAGTATTACAAAAGAGAAGTTAGAA

TTACAGCTACTGACAATATAAAGGGTTTTGTTGAATCAACAATGTGATACGTAAATTA AGAAAAAGAAAAGAAACACAAAAGCTATAGATATACAGATATCAGCTTACCTATTGC

CTTCTATACTTATAATTTAAAGGATTGGTGTCTTAGTACACTTGTGGTCACAGGGATC

AACGAATAGTAAATAATGAACTCGTGCAAGACAAAACTGΆAACCCTCTTTCCAGGACC

TCAGTAGGCACCGTTGAGGTGTCCTTTGTTTTTGTGTGTGTGTGTTCTTTTTTAATTT

TCGCATTGTTGACAGATACAAACAGTTATACTCAATGTACTGTAATAATCGCAAAGGA

AAAAGTTTTGGGATAACTTATTTGTATGTTGGTAGCTGAGAAAAATATCATCAGTCTA

GAATTGATATTTGAGTATAGTAGAGCTTTGGGGCTTTGAAGGCAGGTTCAAGAAAGCA

TATGTCGATGGTTGAGATATTTATTTTCCATATGGTTCATGTTCAAATGTTCACAACC

ACAATGCATCTGACTGCAATAATGTGCTAATAATTTATGTCAGTAGTCACCTTGCTCA

CAGCAAAGCCAGAAATGCTCTCTCCAGGGAGTAGATGTAAAGTACTTGTACATAGAAT

TCAGAACTGAAGATATTTATTAAAAGTTGATTTTTTTTTCTTGATAGTATTTTTATGT

ACTAAATATTTACACTAATATCAATTACATATTTTGGTAAACTAGAGAGACATAATTA

GAGATGCATGCTTCGTTCTGTGCATAGAGACCTTTAAGCAAACTACTACAGCCAACTC

AAAAGCTAAAACTGAACAAATTTGATGTTATACAAACATCTTGCATTTTTAGTAGTTG

ATATTAAGTTGATGACTTGTTTCCCTTCAAGGAAACATTAAATTGTATGGACTCAGCT

AGCTGTTCAATGAAATTGTGAATTAGAAACATTTTTAAAAGTTTTTGAAAGAGATAAG

TGCATCATGAATTACATGTACATGAGAGGAGATAGTGATATCAGCATAATGATTTTGA

GGTCAGTACCTGAGCTGTCTAAAAATATATTATACAAACTAAAATGTAGATGAATTAA

CCTCTCAAAGCACAGAATGTGCAAGAACTTTTGCATTTTAATCGTTGTAAACTAACAG

CTTAAACTATTGACTCTATACCTCTAAAGAATTGCTGCTACTTTGTGCAAGAACTTTG

AAGGTCAAATTAGGCAAATTCCAGATAGTAAAACAATCCCTAAGCCTTAAGTCTTTTT

TTTTTCCTAAΆAATTCCCATAGAATAAAATTCTCTCTAGTTTACTTGTGTGTGCATAC TCTCATCCACAGGGGAAGATAAAGATGGTCACACAAACAGTTTCCATAAAGATGTAC

ATATTCATTATACTTCTGACCTTTGGGCTTTCTTTTCTACTAAGCTAAAAATTCCTTT

TTATCAAAGTGTACACTACTGATGCTGTTTGTTGTACTGAGAGCACGTACCAATAAAA

ATGTTAACAAAATATAAAAA

ORF Start: ATG at 89 ORF Stop: TGA at 3704

SEQ ID NO: 24 1205 aa MW at 135601.5kD

NOV8a, MVLLSLWLIAAALVEVRTSADGQAGNEEMVQIDLPIKRYREYELVTPVSTN EGRYLS

CG93453-01 HTLSASHKKRSARDVSSNPEQLFFNITAFGKDFHLRLKPNTQLVAPGAWEWHETSLV

Protein PGNITDPINLRØQPGSATYRIRRTEPLQTKCAYVGDIVTJIPGTSVAISNCDGLAGMIKS DLSREEYFIEPLERGKQMEEEKGRIH ARYKRSAVEQAPIDMSKDFHYRESDLEGLDDLGT Sequence WGNIHQQIjNETMRRRRHAGΞJ>roγ^EVLLGTO^

IY^mESLGVHINVVLVRMIMLGYA SISLIERGNPSRSLENVCR ASQQQRSDLNHSE

HHDHAIFLTRQDFGPAGMQGYAPVTGMCHPVRSCTLNHEDGFSSAFWAHETGHVLGM

EHDGQGNRCGDETAMGSVMAPLVQAAFHRYHWSRCSGQELKRYIHSYDCLLDDPFDHD PKLPELPGI YSMDEQCRFDFGVGYKMCTAFRTFDPCKQLWCSHPDNPYFCKTKKGP

PLDGTECAAG-OTCYKGHClVrø.αA QQKQDGNWGS TKFGSCSRTCGTGVRFRTRQCNN

PMPINGGQDCPGVNFEYQLv^NTEECQIvΗFEDFFJVQQCQQRNSHFEYQNTKHH LPYEH

PDPKKRCHLYCQSKETGDVAYMKQLVHDGTHCSYOPYSICVRGECV-VGCøKEIGSN

KVEDKCGVCGGDNSHCRTVKGTFTRTPRKLGYLKMFDIPPGARHVLIQEDEASPHILA

IKNQATGHYIL GKGEEAKSRTFIDLGVEWDYNIEDDIESLHTDGPLHDPVIVLIIPQ

ENDTRSSLTYKYIIHEDSVPTINSN VIQEELDTFEWALKSWSQCSKPCGGGFQYTKY

GCRRKSDNKMVHRSFCEANKKPKPIRRMCNIQECTHPLWVAEE EHCTKTCGSSGYQL

RTVRCLQPLLDGTlTOSVHSKYCMGDRPESRRPClSrRVPCPAQWKTGPWSECSVTCGEGT

EVRQVLCRAGDHCDGEKPESVRACQLPPCtTOEPCLGDKSIFCQMEVLARYCSIPGYNK

LCCESCSKRSSTLPPPYLLEAAETHDDVISNPSDLPRSLVMPTSLVPYHSETPAKKMS

LSSISSVGGPNAYAAFRPNSKPDGANLRQRSAQQAGSKTVRLVTVPSSPPTKRVHLSS

ASQMAAASFFAASDSIGASSQARTSKKDGKIIDNRRPTRSSTLER

SEQ ID NO: 25 2286 bp

NOV8b, CTCGAGACAAGGAGGCAGTTGACAGGCTCTGACCGACTCAGGCTTTTCACCATCACAG CG93453-02 TGGTCCCCAGCCCTGCAGAGGACCTGCCTCACCTCCGTTCCTTCACCGCAGGTCACTG DNA Sequence AACACTCACTCCAGGGTCCTGTTTTCCACTGTGCAGGGCAGGGCACTCTGTTACAGGG

CCGGCGGCTCTCGGGACGGTCACCCATGCAGTATTTGCTGTGCACAGAGCGGTTGGTG

CCATCAAGGAGTGGCTGAAGGCAGCGTACAGTGCGAAGCTGATAGCCAGAACTTCCAC

AGGTTTTGGTGCAGTGTTCCCATTCTTCTGCTACCCAGAGTGGATGTGTACACTCTTG

AATATTGCACATTCGTCTAATAGGTTTCGGCTTTTTGTTGGCCTCACAGAAGCTGCGA

TGGACCATTTTATTATCACTTTTCCTACGGCATCCATATTTAGTGTACTGGAAACCTC

CACCACAGGGTTTGGAACACTGAGACCAGCTCTTCAAAGCCCACTCAAAAGTATCTAA

TTCTTCCTGGATGACATTGTTGCTGTTGATTGTAGGTACAGAGTCTTCATGGATGATG

TACTTATATGTCAGGCTAGAGCGGGTATCATTTTCTTGAGGTATAATCAAAACAATAA

CAGGATCATGTAAAGGTCCATCGGTGTGAAGACTTTCAATGTCATCTTCAATGTTATA

ATCCCACTCCACACCAAGATCTATGAAGGTCCGCGACTTGGCTTCCTCCCCTTTGCCA

TTTAAAATATAATGGCCTGTAGCCTGGTTCTTAATAGCAAGAATATGAGGAGAAGCCT

CGTCTTCTTGGATTAACACATGTCTAGCCCCAGGGGGTATATCAAACATCTTAAGGTA

CCCAAGCTTCCTGGGAGTTCTGGTAAATGTCCCCTTCACGGTTCGGCAGTGGGAATTA TCTCCTCCACAGACACCACACTTATCCTCAACCTTATTAGAACCAATTTCTTTATCAC AGCCCACTTTCACACACTCTCCTCGCACACATATGCTATATGGATCTTTGTAAGAACA GTGCGTTCCATCATGCACCAGTTGTTTCATGTAAGCAACATCTCCAGTCTCCTTGGAC TGACAGTAAAGGTGGCATCTTTTCTTGGGGTCAGGATGTTCATATGGCAACCAGTGGT GTTTGGTATTCTGGTATTCAAAGTGGGAGTTTCGCTGCTGACACTGCTGTGCTCTGAA GTCCTCAAAGTGTTTTTGGCATTCTTCTGTGTTACAAAGCTGGTACTCAAAATTAACA CCAGGACAATCCTGACCACCATTGATGGGCATGGGATTATTGCACTGGCGTGTCCTGA AACGAACACCAGTTCCACATGTCCGAGAACAGGAGCCAAATTTAGTCCATGACCCCCA ATTGCCATCTTGTTTTTGCTGATTAGCATTCTTCCACATGCAATGACCCTTATAGCAC CATTTTCCAGCAGCACATTCAGTCCCATCAAGTGGAGGTCCCTTTTTAGTCTTACAAA AGTAGGGATTATCAGGATGGCTACACCACAGCTGTTTACATGGGTCAAAGGTTCGGAA CGCGGTGCACATTTTATAGCCAACACCAAAATCAAAACGACATTGCTCATCCATAGAA TAATTGATTCCAGGAAGTTCTGGGAGTTTAGGCCAATCATGATCAAAAGGGTCATCAA GGAGACAGTCATAGGAATGGATATATCTTTTCAGTTCTTGACCACTGCATCGGGACCA GTGGTAACGATGGAATGCTGCTTGTACCAAGGGAGCCATGACACTTCCCATAGCAGTC TCATCACCACACCTGTTGCCTTGTCCATCATGCTCCATTCCCAACACATGGCCCGTTT CATGGGCTACTACAAAAGCAGATGAAAAACCATCCTCATGATTCAGGGTACAACTTCT CACTGGATGACACATGCCGGTGACTGGAGCATATCCTTGCATTCCAGCAGGTCCAAAG TCTTGCCTGGTTAAAAAAATTGCATGGTCATGGTGTTCAGAGTGGTTGAGATCAGATC TTTGCTGTTGGGACGCCCAGCGACACACATTCTCCAAGCTTCTGGATGGGTTTCCCCT TTCTATGAGGCTGATGGACTTTGCATATCCCAGCATTATCATGCGCACCAGGACCACA TTTATATGCACTCCGAGGGACTCATCATGGTAAATTTCATTCACAATGTTCATTAGGG TCAGGAGGTAGTTTTGGACGTGCTCTTTGCCATGGAAACGGACCACAGAGTCATCCAC TCCCAGCAGTACCTCGATGGATCC ORF Start: at 829 ORF Stop: end of sequence

SEQ ID NO: 26 758 aa MW at 86176.4kD

NOV8b, II3 L GVDDSV FHGI<^HVQN L^-NIVNEIYlrDES GVHI VV MIMLβ K CG93453-02 SISLIERGNPSRSLΞlWCRWASQQQRSDIuNHSEHHDHAIFLTRQDFGPAGMQGYAPVT Protein G CHPVRSCTLNHEDGFSSAFVVAHETGHVLGMEHDGQG RCGDΞTAMGSVMAPNQQA Sequence AFHRYHWSRCSGQELI^YIHSYTJCLLDDPFDimWP-vXPELPGIi^ryrSMDEQCRFDFGVG

YKMCTAFRTFDPCKQLWCSHPDNPYFCKTKKGPPLDGTECAAGK CYKGHC)>WK A Q Q QDGNWGS TKFGSCSRTCGTGVRFRTRQCNNPMPINGGQDCPGV FEYQLCNTEEC QKHFEDFRAQQCQQRNSHFEYQNTKHHWLPYEHPDPKKRCHLYCQSKETGDVAYMKQL VHDGTHCSYKDPYSIC TOGECVKVGCDISΈIGSNKΛ EDKCGVCGGDNSHCRTVKGTFTR TPRIVXGYT-KMFDIPPGAL^VLIQEDEASPHII-AIKLVTQATGHYILNGKGEEAKSRTFID GVE DY IEDDIESLHTDGPLHDPVIVLIIPQE DTRSSLTYKYIIHEDSVPTINSN JRVIQEELDTFE ALKSWSQCSKPCGGGFQYTKYGCRRKSDN-MVHRSFCEANK PKPI RFJ^CNIQECTHPLWVAEE EHCTKTCGSSGYQLRTVRCLQPLLDGTNRSVHSKYCMGD RPΞSRRGCNRVGCRAQWKTAGRSECSVTCGEGTEVRQVLCRAGDHCDGEKPESVRACQ

LPGC

SEQ ID NO: 27 2286 bp

NOV8c, GGATCCATCGAGGTACTGCTGGGAGTGGATGACTCTGTGGTCCGTTTCCATGGCAAAG 210387874 AGCACGTCCAAAACTACCTCCTGACCCTAATGAACATTGTGAATGAAATTTACCATGA DNA Sequence TGAGTCCCTCGGAGTGCATATAAATGTGGTCCTGGTGCGCATGATAATGCTGGGATAT GCAAAGTCCATCAGCCTCATAGAAAGGGGAAACCCATCCAGAAGCTTGGAGAATGTGT GTCGCTGGGCGTCCCAACAGCAAAGATCTGATCTCAACCACTCTGAACACCATGACCA TGCAATTTTTTTAACCAGGCAAGACTTTGGACCTGCTGGAATGCAAGGATATGCTCCA GTCACCGGCATGTGTCATCCAGTGAGAAGTTGTACCCTGAATCATGAGGATGGTTTTT CATCTGCTTTTGTAGTAGCCCATGAAACGGGCCATGTGTTGGGAATGGAGCATGATGG ACAAGGCAACAGGTGTGGTGATGAGACTGCTATGGGAAGTGTCATGGCTCCCTTGGTA CAAGCAGCATTCCATCGTTACCACTGGTCCCGATGCAGTGGTCAAGAACTGAAAAGAT ATATCCATTCCTATGACTGTCTCCTTGATGACCCTTTTGATCATGATTGGCCTAAACT CCCAGAACTTCCTGGAATCAATTATTCTATGGATGAGCAATGTCGTTTTGATTTTGGT GTTGGCTATAAAATGTGCACCGCGTTCCGAACCTTTGACCCATGTAAACAGCTGTGGT GTAGCCATCCTGATAATCCCTACTTTTGTAAGACTAAAAAGGGACCTCCACTTGATGG GACTGAATGTGCTGCTGGAAAATGGTGCTATAAGGGTCATTGCATGTGGAAGAATGCT AATCAGCAAAAACAAGATGGCAATTGGGGGTCATGGACTAAATTTGGCTCCTGTTCTC GGACATGTGGAACTGGTGTTCGTTTCAGGACACGCCAGTGCAATAATCCCATGCCCAT CAATGGTGGTCAGGATTGTCCTGGTGTTAATTTTGAGTACCAGCTTTGTAACACAGAA GAATGCCAAAAACACTTTGAGGACTTCAGAGCACAGCAGTGTCAGCAGCGAAACTCCC ACTTTGAATACCAGAATACCAAACACCACTGGTTGCCATATGAACATCCTGACCCCAA GAAAAGATGCCACCTTTACTGTCAGTCCAAGGAGACTGGAGATGTTGCTTACATGAAA CAACTGGTGCATGATGGAACGCACTGTTCTTACAAAGATCCATATAGCATATGTGTGC GAGGAGAGTGTGTGAAAGTGGGCTGTGATAAAGAAATTGGTTCTAATAAGGTTGAGGA TAAGTGTGGTGTCTGTGGAGGAGATAATTCCCACTGCCGAACCGTGAAGGGGACATTT ACCAGAACTCCCAGGAAGCTTGGGTACCTTAAGATGTTTGATATACCCCCTGGGGCTA GACATGTGTTAATCCAAGAAGACGAGGCTTCTCCTCATATTCTTGCTATTAAGAACCA GGCTACAGGCCATTATATTTTAAATGGCAAAGGGGAGGAAGCCAAGTCGCGGACCTTC ATAGATCTTGGTGTGGAGTGGGATTATAACATTGAAGATGACATTGAAAGTCTTCACA CCGATGGACCTTTACATGATCCTGTTATTGTTTTGATTATACCTCAAGAAAATGATAC CCGCTCTAGCCTGACATATAAGTACATCATCCATGAAGACTCTGTACCTACAATCAAC AGCAACAATGTCATCCAGGAAGAATTAGATACTTTTGAGTGGGCTTTGAAGAGCTGGT CTCAGTGTTCCAAACCCTGTGGTGGAGGTTTCCAGTACACTAAATATGGATGCCGTAG GAAAAGTGATAATAAAATGGTCCATCGCAGCTTCTGTGAGGCCAACAAAAAGCCGAAA CCTATTAGACGAATGTGCAATATTCAAGAGTGTACACATCCACTCTGGGTAGCAGAAG AATGGGAACACTGCACCAAAACCTGTGGAAGTTCTGGCTATCAGCTTCGCACTGTACG CTGCCTTCAGCCACTCCTTGATGGCACCAACCGCTCTGTGCACAGCAAATACTGCATG GGTGACCGTCCCGAGAGCCGCCGGCCCTGTAACAGAGTGCCCTGCCCTGCACAGTGGA AAACAGGACCCTGGAGTGAGTGTTCAGTGACCTGCGGTGAAGGAACGGAGGTGAGGCA GGTCCTCTGCAGGGCTGGGGACCACTGTGATGGTGAAAAGCCTGAGTCGGTCAGAGCC TGTCAACTGCCTCCTTGTCTCGAG

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 8B.

Further analysis of the NOV8a protein yielded the following properties shown in Table 8C.

Table 8C. Protein Sequence Properties NOV8a

PSort analysis: 0.5708 probability located in outside; 0.1900 probability located in lysosome (lumen); 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen)

SignalP analysis: I Cleavage site between residues 21 and 22

A search of the NOVSa protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 8D.

In a BLAST search of public sequence databases, the NOV8a protein was found to have homology to the proteins shown in the BLASTP data in Table 8E.

PFam analysis predicts that the NOV8a protein contains the domains shown in the Table 8F.

EXAMPLE 9.

The NOV9 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 9A.

Further analysis of the NOV9a protein yielded the following properties shown in

Table 9B.

Table 9B. Protein Sequence Properties NOV9a

PSort analysis: 0.3798 probability located in outside; 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen); 0.1000 probability located in lysosome (lumen)

SignalP analysis: I Cleavage site between residues 22 and 23

A search of the NOV9a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 9C.

In a BLAST search of public sequence databases, the NOV9a protein was found to have homology to the proteins shown in the BLASTP data in Table 9D.

PFam analysis predicts that the NOV9a protein contains the domains shown in the Table 9E.

EXAMPLE 10.

The NOVIO clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 10A.

Table 10A. NOVIO Sequence Analysis

SEQ ID NO: 31 3121 bp

NOVlOa, TTTTGACAGCTGCCACAGTCTCTGAGCTCCAGCCTCGCGCCTGAACCCGGTCCCTGCC CG95250-01 ATGGGGCCCCCTTCCAGCTCAGGCTTCTATGTGAGCCGCGCAGTGGCCCTGCTGCTGG DNA Sequence CTGGGCTGGTAGCCGCCCTCCTGCTGGCGCTGGCCGTACTCGCCGCCTTGTACGGCCA CTGCGAGCGCGTCCCACCGTCGGAGCTGCCTGGACTCAGGGACTTGGAAGCCGAGTCT TCCCCTCCCCTCAGGCAGAAGCCGACGCCAACCCCGAAACCCAGCAGTGCACGCGAGC TAGCGGTGACGACCACCCCGAGCAACTGGCGACCCCCGGGGCCCTGGGACCAGCTACG CCTGCCGCCCTGGCTCGTGCCGCTGCACTACGATCTGGAGCTGTGGCCGCAGCTGAGG CCCGACGAGCTTCCGGCCGGGTCTTTGCCCTTCACTGGCCGCGTGAACATCACGGTGC GCTGCACGGTGGCCACCTCTCGACTGCTGCTGCATAGCCTCTTCCAGGACTGCGAGCG CGCCGAGGTGCGGGGACCCCTTTCCCCGGGCACTGGGAACGCCACAGTGGGCCGCGTG CCCGTGGACGACGTGTGGTTCGCGCTGGACACGGAATACATGGTGCTGGAGCTCAGTG AGCCCCTGAAACCTGGTAGCAGCTACGAGCTGCAGCTTAGCTTCTCGGGCCTGGTGAA GGAAGACCTCAGGGAGGGACTCTTCCTCAACGTCTACACCGACCAGGGCGAGCGCAGG GCCCTGTTAGCGTCCCAGCTGGAACCAACATTTGCCAGGTATGTTTTCCCTTGTTTTG ATGAGCCAGCTCTGAAGGCAACTTTTAATATTACAATGATTCATCATCCAAGTTATGT GGCCCTTTCCAACATGCCAAAGAATTCTCAGTCTGAAAAAGAAGATGTGAATGGAAGC AAATGGACTGTTACAACCTTTTCCACTACGCCCCACATGCCAACTTACTTAGTCGCAT TTGTTATATGTGACTATGACCACGTCAACAGAACAGAAAGGGGCAAGGAGGTGATACG CATCTGGGCCCGGAAAGATGCAATTGCAAATGGAAGTGCAGACTTTGCTTTGAACATC ACAGGTCCCATCTTCTCTTTTCTGGAGGATTTGTTTAATATCAGTTACTCTCTTCCAA AAACAGATATAATTGCCTTGCCTAGTTTTGACAACCATGCAATGGAAAACTGGGGACT AATGATATTTGATGAATCAGGATTGTTGTTGGAACCAAAAGATCAACTGACAGAAAAA AAGACTCTGATCTCCTATGTTGTCTCCCACGAGATTGGACACCAGTGGTTTGGAAACT TGGTTACCATGAATTGGTGGAACAATATCTGGCTCAACGAGGGTTTTGCATCTTATTT TGAGTTTGAAGTAATTAACTACTTTAATCCTAAACTCCCAAGAGTAAGTAATGAGATC TTTTTTTCTAACATTTTACATAATATCCTCAGAGAAGATCACGCCCTGGTGACTAGAG CTGTGGCCATGAAGGTGGAAAATTTCAAAACAAGTGAAATACAGGAACTCTTTGACAT ATTTACTTACAGCAAGGTAAAAGCAGTTAGAAATTTCCTTTGGTTTTGTACTCTGGTA GAAAGTCTATATCATCATACATTACAGTCATATTTGAAGACATTTTCCTACTCAAACG CTGAGCAAGATGATCTATGGAGGCATTTTCAACAGGCCATAGATGACCAGAGTACAGT TATTTTGCCAGCAACAATAAAAAACATAATGGACAGTTGGACACACCAGAGTGGTTTT CCAGTGATCACTTTAAATGTGTCTACTGGCGTCATGAAACAGGAGCCATTTTATCTTG AAAACATTAAAAATCGGACTCTTCTAACCAGCAAGGACACATGGATTGTCCCTATTCT TTGGATAAAAAATGGAACTACACAACCTTTAGTCTGGCTAGATCAAAGCAGCAAAGTA TTCCCAGAAATGCAAGTTTCAGATTCTGACCATGACTGGGTGATTTTGAATTTGAATA TGACTGGATATTATAGAGTTAATTATGATAAATTAGGTTGGAAGAAACTAAATCAACA ACTTGAAAAGGATCCTAAGGCTATTCCTGTTATTCACAGACTGCAGTTGATTGATGAT GCCTTTTCCTTGTCTAAGAAGTTATTGAGCTTGTCCCGAACTTTGCCTTTGGACCACT TCTTCTTTTTGGCCTTGCCCCCGGATTTGTTCACTGGGTCTTTGTCTTTCTTGGCTGA CTTTCCAGCGTCCTTCTTCTCGTCGTCCTTGGGCGCTGCTCAGGAAAAAAAAAGATTC CTTCCAAAATATTACTGCTCATTGATGATGCACTTGGTTACACAAGCACTAATGATGG AGATGTACAAGGAGATTAATGTTGTTTTCATGCCCACTAACACAATATCAATTCTTCA GCCCAGGGATCAAGGAGTAACTGCGTGTTGGTTGGGCCTTGAAGACTGCCTTCAGCTG TCAAAAGAACTTTTCGCAAAATGGGTGGATCATCCAGAAAATAGAATACCTTATCCAA TTAAAGATGTGGTTTTATGTTATGGCATTGCCTTGGGAAGTGATAAAGAGTGGGACAT CTTGTTAAATACTTACACTAATACAACAAACAAAGAAGAAAAGATTCAACTTGCTTAT GCAATGAGCTGCAGCAAAGACCCATGGATACTTAACAGGAGATATATGGAGTATGCCA TCAGCACATCTCCATTCACTTCTAATGAAACAAATATAATTGAGGTTGTGGCTTCATC TGAAGTTGGCCGGTATGTCGCAAAAGACTTCTTAGTCAACAACTGGCAAGCTGTGAGT CACACTTTGAGCAGGTATGGAACACAATCATTGATTAATCTAATATATACAATAGGGA GAACCGTAACTACAGATTTACAGATTGTGGAGCTGCAGCAGTTTTTCAGTAACATGTT GGAGGAACACCAGAGGATCAGAGTTCATGCCAACTTACAGACAATAAAGAATGAAAAT CTGAAAAACAAGAAGCTAAGTGCCAGGATAGCTGCGTGGCTAAGGAGAAACACATAGC TTGTGGCTATCTTTCAGCACTCCTCTTGCATATTATAATGTAGTTTG

ORF Start: ATG at 59 ORF Stop: TAG at 3071

SEQ ID NO: 32 1004 aa MW at 114692.2kD

NOVlOa, MGPPSSSGFY tSRAVALLI-AGLVAALLl-ALAVLAALYGHCERVPPSELPGLRDLEAES CG95250-01 SPPLRQKPTPTPKPSSARELAVTTTPSN RPPGPWDQLRLPP LVPLHYDLELWPQLR Protein PDELPAGSLPFTGRVNITVRCTVATSRLLLHSLFQDCERAEVRGPLSPGTGNATVGRV Sequence PVDDVWFALDTEYiVLELSEPLKPGSSYELQLSFSGLVIvΕDLREGLFI.NVYTDQGERR

ALIASQLEPTFARYVFPCFDEPALKATF ITMIHHPSY/ALSNMPKNSQSEKEDVNGS

KWTVTTFSTTPHMPTYLVAFVICDYDHV1TOTΞRGKEVIRIWARKDAIANGSADFALNI

TGPIFSFLEDLFNISYSLPKTDIIALPSFDNHAME GL IFDESGLLLEPKDQLTEK

KTLISYVVSHEIGHQ FGNLVT lvriWNNIWLNEGFASYFEFEVINYFNPKLPRVSNEI

FFSNILHITILREDHALVTRAVAMK NFKTSEIQELFDIFTYSKVKAVRlvTFL FCTLV

ESLYHHTLQSYLKTFSYSNAEQDDL RHFQQAIDDQSTVILPATIKNI DS THQSGF

PVITLNVSTGVMKQEPFYLENIKlvTRTLLTSKDTWIVPIL IKNGTTQPLVWLDQSSKV

FPΞMQVSDSDHD IL^LlSMTGYΛrRVNTO

AFSLSKKLLSLSRTLPLDHFFFLALPPDLFTGSLSFLADFPASFFSSSLGAAQEKKRF

LPKYYCSI-MMHLVTQAL MEMYKEIjrVVFMPTNTISILQPRDQGVTAC LGLEDCLQL

SKELFAKW/DHPEKRIPYPIi VVLCYGIALGSD.ΕWD^

AMSCSKDP ILMRRYMEYAISTSPFTSNETNIIFΛ/ASSEVGRY/AKDFLVNNWQAVS

HTLSRYGTQSLINLIYTIGRTVTTDLQIVELQQFFSNMLEEHQRIRVHAIvrLQTIKNEN

LKN KLSARIAA LRRNT

SEQ ID NO: 33 2880 bp

NOVl 0b, TTTTGACAGCTGCCACAGTCTCTGAGCTCCAGCCTCGCGCCTGAACCCGGTCCCTGCC CG95250-02 ATGGGGCCCCCTTCCAGCTCAGGCTTCTATGTGAGCCGCGCAGTGGCCCTGCTGCTGG DNA Sequence CTGGGCTGGTAGCCGCCCTCCTGCTGGCGCTGGCCGTACTCGCCGCCTTGTACGGCCA CTGCGAGCGCGTCCCACCGTCGGAGCTGCCTGGACTCAGGGACTTGGAAGCCGAGTCT TCCCCTCCCCTCAGGCAGAAGCCGACGCCAACCCCGAAACCCAGCAGTGCACGCGAGC TAGCGGTGACGACCACCCCGAGCAACTGGCGACCCCCGGGGCCCTGGGACCAGCTACG CCTGCCGCCCTGGCTCGTGCCGCTGCACTACGATCTGGAGCTGTGGCCGCAGCTGAGG CCCGACGAGCTTCCGGCCGGGTCTTTGCCCTTCACTGGCCGCGTGAACATCACGGTGC GCTGCACGGTGGCCACCTCTCGACTGCTGCTGCATAGCCTCTTCCAGGACTGCGAGCG CGCCGAGGTGCGGGGACCCCTTTCCCCGGGCACTGGGAACGCCACAGTGGGCCGCGTG CCCGTGGACGACGTGTGGTTCGCGCTGGACACGGAATACATGGTGCTGGAGCTCAGTG AGCCCCTGAAACCTGGTAGCAGCTACGAGCTGCAGCTTAGCTTCTCGGGCCTGGTGAA GGAAGACCTCAGGGAGGGACTCTTCCTCAACGTCTACACCGACCAGGGCGAGCGCAGG GCCCTGTTAGCGTCCCAGCTGGAACCAACATTTGCCAGGTATGTTTTCCCTTGTTTTG ATGAGCCAGCTCTGAAGGCAACTTTTAATATTACAATGATTCATCATCCAAGTTATGT GGCCCTTTCCAACATGCCAAAGAATTCTCAGTCTGAAAAAGAAGATGTGAATGGAAGC AAATGGACTGTTACAACCTTTTCCACTACGCCCCACATGCCAACTTACTTAGTCGCAT TTGTTATATGTGACTATGACCACGTCAACAGAACAGAAAGGGGCAAGGAGGTGATACG CATCTGGGCCCGGAAAGATGCAATTGCAAATGGAAGTGCAGACTTTGCTTTGAACATC ACAGGTCCCATCTTCTCTTTTCTGGAGGATTTGTTTAATATCAGTTACTCTCTTCCAA AAACAGATATAATTGCCTTGCCTAGTTTTGACAACCATGCAATGGAAAACTGGGGACT AATGATATTTGATGAATCAGGATTGTTGTTGGAACCAAAAGATCAACTGACAGAAAAA AAGACTCTGATCTCCTATGTTGTCTCCCACGAGATTGGACACCAGTGGTTTGGAAACT TGGTTACCATGAATTGGTGGAACAATATCTGGCTCAACGAGGGTTTTGCATCTTATTT TGAGTTTGAAGTAATTAACTACTTTAATCCTAAACTCCCAAGAGTAAGTAATGAGATC TTTTTTTCTAACATTTTACATAATATCCTCAGAGAAGATCACGCCCTGGTGACTAGAG CTGTGGCCATGAAGGTGGAAAATTTCAAAACAAGTGAAATACAGGAACTCTTTGACAT ATTTACTTACAGCAAGGGAGCGTCTATGGCCCGGATGCTTTCTTGTTTCTTGAATGAG CATTTATTTGTCAGTGCATTACAGTCATATTTGAAGACATTTTCCTACTCAAACGCTG AGCAAGATGATCTATGGAGGCATGATTTTTTAAAACAGGCCATAGATGACCAGAGTAC AGTTATTTTGCCAGCAACAATAAAAAACATAATGGACAGTTGGACACACCAGAGTGGT TTTCCAGTGATCACTTTAAATGTGTCTACTGGCGTCCTGATACAGGAGCCATTTTATC TTGAAAACATTAAAAATCGGACTCTTCTAACCAGCAATGACACATGGATTGTCCCTAT TCTTTGGATAAAAAATGGAACTACACAACCTTTAGTCTGGCTAGATCAAAGCAGCAAA GTATTCCCAGAAATGCAAGTTTCAGATTCTGACCATGACTGGGTGATTTTGAATTTGA ATATGACTGGATATTATAGAGTTAATTATGATAAATTAGGTTGGAAGAAACTAAATCA ACAACTTGAAAAGGATCCTAAGGCTATTCCTGTTATTCACAGACTGCAGTTCATTGAT GATGCCTTTTCCTTGTCTAAAAACAATTATATTGAGATTGAAACAGCACTTGAGTTAA CCAAGTACCTTGCTGAAGAAGATGAAATTATAGTATGGCATACAGTCTTGGTAAACTT GGTAACCAGGGATCTTGTTTCTGAGGTGAACATCTATGATATATACTCATTATTAAAG ACTGCGTGTTGGTTGGGCCTTGAAGACTGCCTTCAGCTGTCAAAAGAACTTTTCGCAA AATGGGTGGATCATCCAGAAAATATTAAAGATGTGGTTTTATGTTATGGCATTGCCTT GGGAAGTGATAAAGAGTGGGACATCTTGTTAAATACTTACACTAATACAACAAACAAA GAAGAAAAGATTCAACTTGCTTATGCAATGAGCTGCAGCAAAGACCCATGGATACTTA ACAGGTATATGGAGTATGCCATCAGCACATCTCCATTCACTTCTAATGAAACAAATAT AATTGAGGTTGTGGCTTCATCTGAAGTTGGCCGGTATGTCGCAAAAGACTTCTTAGTC AACAACTGGCAAGCTGTGAGTAAAAGGTAAGAAGGAAAGTGAGACCTTTCTTTCATTT

AGGCCACTGGTTTGGCACTGGAAGCTCAGCTTTAGTCTAGCTTGGAAGCTCAGCTTTA

GTCTAGCTAGGCCACAAACGTCCTTTGCATTGACTAGAAAAGTTATCATTTTTCCTTT

GTTTAGTCTCACTACAAACTGCCTGTGTTATGGAAGAG

ORF Start: ATG at 59 ORF Stop: TAA at 2696

SEQ ID NO: 34 879 aa MW at 100068.0kD

NOVl Ob, MGPPSSSGFYVSRAVALLrAGLVAALLL-ALAVLAALYGHCERVPPSELPGLRDLEAES CG95250-02 SPPLRQKPTPTPKPSSAREIAVTTTPSNWRPPGPWDQLRLPP LVPLHYDLEL PQLR Protein PDELPAGSLPFTGRVNITVRCTVATSRLLLHSLFQDCERAEVRGPLSPGTGNATVGRV Sequence PVDDVWFALDTEYMVLELSEPLKPGSSYELQLSFSGLVKEDLREGLFIJ^rVYTDQGERR

ALI-ASQLEPTFARYVFPCFDEPALIvATFNIT IHHPSYVALSNMPK SQSEKEDVNGS

K TVTTFSTTPHMPTYLVAFVICDYDHVNRTERGKEVIRI ARKDAIANGSADFALNI

TGPIFSFLEDLFNISYSLPKTDIIALPSFDNHAME GLMIFDESGLLLEPKDQLTEK

KTLISYΛTVSHEIGHQWFGlrLVTMIW^

FFSNILH ILREDHALVTRAVAMKVENFKTSEIQΞLFDIFTYSKGASMARMLSCFLNE

HLFVSALQSYLKTFSYSNAEQDDL RHDFLKQAIDDQSTVILPATIKNIMDS THQSG

FPVITiαWSTGVLIQEPFYLENIKNRTLLTSlsroT IVPILWIKIvTGTTQPLVWLDQSSK

VFPEMQVSDSDHDWVITJvjLNMTGYYRVNYDKLG I^LNQQLEKDPKAIPVII^

DAFSLSKMSTYIEIETALELTKYLAEEDEIIVVraTVLVlsrLVTRDLVSEVNIYDIYSLLK

TAC LGLEDCLQLSKELFAKWVDHPENIIvTJVVLCYGIALGSDKEVroiLLNTYTNTTNK

EEKIQLAYAMSCSKDP IIiNRYMEYAISTSPFTSNETNIIEVVASSEVGRYΛ^AKDFLV

NN QAVSKR

SEQ ID NO: 35 1695 bp

NOVlOc, ATGGGGCCCCCTTCCAGCTCAGGCTTCTATGTGAGCCGCGCAGTGGCCCTGCTGCTGG CG95250-03 CTCGGCTGGTAGCCGCCCTCCTGCTGGCGCTGGCCGTACTCGCCGCCTTGTACGGCCA DNA Sequence CTGCGAGCGCGTCCCACCGTCGGAGCTGCCTGGACTCAGGGACTTGGAAGCCGAGTCT TCCCCTCCCCTCAGGCAGAAGCCGACGCCAACCCCGAAACCCAGCAGTGCACGCGAGC TAGCGGTGACGACCACCCCGAGCAACTGGCGACCCCCGGGGCCCTGGGACCAGCTACG CCTGCCGCCCTGGCTCGTGCCGCTGCACTACGATCTGGAGCTGTGGCCGCAGCTGAGG CCCGACGAGCTTCCGGCCGGGTCTTTGCCCTTCACTGGCCGCGTGAACATCACGGTGC GCTGCACGGTGGCCACCTCTCGACTGCTGCTGCATAGCCTCTTCCAGGACTGCGAGCG CGCCGAGGTGCGGGGACCCCTTTCCCCGGGCACTGGGAACGCCACAGTGGGCCGCGTG CCCGTGGACGACGTGTGGTTCGCGCTGGACACGGAATACATGGTGCTGGAGCTCAGTG AGCCCCTGAAACCTGGTAGCAGCTACGAGCTGCAGCTTAGCTTCTCGGGCCTGGTGAA GGAAGACCTCAGGGAGGGACTCTTCCTCAACGTCTACACCGACCAGGGCGAGCGCAGG GCCCTGTTAGCGTCCCAGCTGGAACCAACATTTGCCAGGTATGTTTTCCCTTGTTTTG ATGAGCCAGCTCTGAAGGCAACTTTTAATATTACAATGATTCATCATCCAAGTTATGT GGCCCTTTCCAACATGCCAAAGCTAGGTCAGTCTGAAAAAGAAGATGTGAATGGAAGC AAATGGACTGTTACAACCTTTTCCACTACGCCCCACATGCCAACTTACTTAGTCGCAT TTGTTATATGTGACTATGACCACGTCAACAGAACAGAAAGGGGCAAGGAGATACGCAT CTGGGCCCGGAAAGATGCAATTGCAAATGGAAGTGCAGACTTTGCTTTGAACATCACA GGTCCCATCTTCTCTTTTCTGGAGGATTTGTTTAATATCAGTTACTCTCTTCCAAAAA CAGATATAATTGCCTTGCCTAGTTTTGACAACCATGCAATGGAAAACTGGGGACTAAT

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 10B.

Table 10B. Comparison of NOVlOa against NOVlOb and NOVlOc.

NOVlOa Residues/ Identities/

Protein Sequence Match Residues Similarities for the Matched Region

NOVl Ob 1..928 783/931 (84%) 1..877 808/931 (86%)

NOVlOc 1..555 509/555 (91%) 1..551 512/555 (91%)

Further analysis of the NOVlOa protein yielded the following properties shown in Table IOC.

Table IOC. Protein Sequence Properties NOVlOa

PSort analysis: 0.8000 probability located in mitochondrial inner membrane; 0.6500 probability located in plasma membrane; 0.6199 probability located in microbody (peroxisome); 03000 probability located in Golgi body

SignalP analysis: Cleavage site between residues 35 and 36

A search of the NOVlOa protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 10D.

In a BLAST search of public sequence databases, the NOVlOa protein was found to have homology to the proteins shown in the BLASTP data in Table 10E.

PFam analysis predicts that the NOVlOa protein contains the domains shown in the Table 1 OF.

Table 10F. Domain Analysis of NOVlOa

Identities/

Pfam Domain NOVlOa Match Region Similarities Expect Value for the Matched Region

Peptidase_Ml 98..509 150/451 (33%) 9.1e-144 323/451 (72%) EXAMPLE 11.

The NOVl 1 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 11 A.

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 1 IB.

Further analysis of the NOVl la protein yielded the following properties shown in Table HC.

Table HC. Protein Sequence Properties NOVlla

PSort analysis: 0.6400 probability located in microbody (peroxisome); 0.5057 probability located in mitochondrial matrix space; 0.2277 probability located in mitochondrial inner membrane; 0.2277 probability located in mitochondrial intermembrane space

SignalP analysis: Cleavage site between residues 20 and 21

A search of the NOVl la protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 1 ID.

In a BLAST search of public sequence databases, the NOVl la protein was found to have homology to the proteins shown in the BLASTP data in Table 1 IE.

PFam analysis predicts that the NOVl la protein contains the domains shown in the Table 1 IF.

EXAMPLE 12.

The NOVl 2 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 12 A.

Further analysis of the NOVl 2a protein yielded the following properties shown in Table 12B.

Table 12B. Protein Sequence Properties NOV12a

PSort analysis: 0.5729 probability located in mitochondrial matrix space; 0.2867 probability located in mitochondrial inner membrane; 0.2867 probability located in mitochondrial intermembrane space; 0.2867 probability located in mitochondrial outer membrane

SignalP analysis: Cleavage site between residues 24 and 25

A search of the NOVl 2a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 12C.

In a BLAST search of public sequence databases, the NOVl 2a protein was found to have homology to the proteins shown in the BLASTP data in Table 12D.

PFam analysis predicts that the NOVl 2a protein contains the domains shown in the Table 12E.

EXAMPLE 13.

The NOVl 3 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 13 A.

NOVl 3a, TGGACTCCTGTTACCATGAGGTTCCTGATCCTGTTCCTAGCCCTGTCCCTAGGAGGGA CG95804-01 TTGATGCTGCACCTCCTGTCCAGTCTCGAATTGTTGGAGGATTTAACTGTGAGAAGAA DNA Sequence TTCCCAGCCCTGGCAAGTGGCTGTGTACCGCTTCACCAAATATCAATGTGGGGGTATC CTGCTGAACGCCAACTGGGTTCTCACAGCTGCCCACTGCCATAATGACAAGTACCAGG TGTGGCTGGGCAAAAACAACTTTTTGGAGGATGAACCCTCTGCCCAACACCGGCTTGT CAGCAAAGCCATCCCTCACCCTGACTTCAACATGAGCCTCCTGAATGAGCACACCCCA CAACCTGAGGATGACTACAGCAATGACCTGATGCTGCTCCGCCTCAAAAAGCCTGCTG ACATCACAGATGTTGTGAAGCCCATCGACCTGCCCACTGAGGAGCCCAAGCTGGGGAG CACATGCCTAGCCTCAGGCTGGGGCAGCATTACACCCGTCAAATATGAATACCCAGAT GAGCTCCAGTGTGTGAACCTCAAGCTCCTGCCTAATGAGGACTGTGCCAAAGCCCACA TAGAGAAGGTGACAGATGACATGTTGTGTGCAGGAGATATGGATGGAGGCAAAGACAC TTGTGCGGGTGACTCAGGAGGCCCACTGATCTGTGATGGTGTTCTCCAAGGTATCACA TCATGGGGCCCTAAGCCTTGCGGTAAACCCAATGTGCCGGGTATCTACACCAGAGTTT TAAATTTCAACACCTGGATAAGAGAAACTATGGCTGAAAATGACTGAGTATCACATTG TCCCAT

ORF Start: ATG at 16 ORF Stop: TGA at 799

SEQ ID NO: 70 261 aa MW at 28815.6kD

NOV13a, MRFLILFLALSLGGIDAAPPVQSRIVGGFNCEKNSQPWQVAVYRFTKYQCGGILLNAN CG95804-01 VLTAAHCHNDKYQVWLGKNNFLEDEPSAQHRLVSKAIPHPDFNMSLLNEHTPQPEDD Protein YSNDLMLLRLKKPADITDVVKPIDLPTEEPKLGSTCLASG GSITPVKYEYPDELQCV Sequence NLKLLPNEDCAlvAHIEKVTDDMLCAGDIvlDGGKDTCAGDSGGPLICDGVLQGITS GPK PCGKPNVPGIYTRVLNFNTWIRETMAEND

Further analysis of the NOV13a protein yielded the following properties shown in

Table 13B.

Table 13B. Protein Sequence Properties NOV13a

PSort analysis: 0.6281 probability located in outside; 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen); 0.1000 probability located in lysosome (lumen)

SignalP analysis: Cleavage site between residues 18 and 19

A search of the NOVl 3a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 13C.

In a BLAST search of public sequence databases, the NOVl 3a protein was found to have homology to the proteins shown in the BLASTP data in Table 13D.

PFam analysis predicts that the NOVl 3a protein contains the domains shown in the Table 13E.

EXAMPLE 14.

The NOVl 4 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 14 A. Table 14A. NOV14 Sequence Analysis

SEQ ID NO: 71 2691 bp

NOV14a, GCTTGCCCGTCGGTCGCTAGCTCGCTCGGTGCGCGTCGTCCCGCTCCATGGCGCTCTT CG95861-01 CGTGCGGCTGCTGGCTCTCGCCCTGGCTCTGGCCCTGGGCCCCGCCGCGACCCTGGCG DNA Sequence GGTCCCGCCAAGTCGCCCTACCAGCTGGTGCTGCAGCACAGCAGGCTCCGGGGCCGCC AGCACGGCCCCAACGTGTGTGCTGTGCAGAAGGTTATTGGCACTAATAGGAAGTACTT CACCAACTGCAAGCAGTGGTACCAAAGGAAAATCTGTGGCAAATCAACAGTCATCAGC TACGAGTGCTGTCCTGGATATGAAAAGGTCCCTGGGGAGAAGGGCTGTCCAGCAGCCC TACCACTCTCAAACCTTTACGAGACCCTGGGAGTCGTTGGATCCACCACCACTCAGCT GTACACGGACCGCACGGAGAAGCTGAGGCCTGAGATGGAGGGGCCCGGCAGCTTCACC ATCTTCGCCCCTAGCAACGAGGCCTGGGCCTCCTTGCCAGCTGAAGTGCTGGACTCCC TGGTCAGCAATGTCAACATTGAGCTGCTCAATGCCCTCCGCTACCATATGGTGGGCAG GCGAGTCCTGACTGATGAGCTGAAACACGGCATGACCCTCACCTCTATGTACCAGAAT TCCAACATCCAGATCCACCACTATCCTAATGGGATTGTAACTGTGAACTGTGCCCGGC TCCTGAAAGCCGACCACCATGCAACCAACGGGGTGGTGCACCTCATCGATAAGGTCAT CTCCACCATCACCAACAACATCCAGCAGATCATTGAGATCGAGGACACCTTTGAGACC CTTCGGGCTGCTGTGGCTGCATCAGGGCTCAACACGATGCTTGAAGGTAACGGCCAGT ACACGCTTTTGGCCCCGACCAATGAGGCCTTCGAGAAGATCCCTAGTGAGACTTTGAA CCGTATCCTGGGCGACCCAGAAGCCCTGAGAGACCTGCTGAACAACCACATCTTGAAG TCAGCTATGTGTGCTGAAGCCATCGTTGCGGGGCTGTCTGTAGAGACCCTGGAGGGCA CGACACTGGAGGTGGGCTGCAGCGGGGACATGCTCACTATCAACGGGAAGGCGATCAT CTCCAATAAAGACATCCTAGCCACCAACGGGGTGATCCACTACATTGATGAGCTACTC ATCCCAGACTCAGCCAAGACACTATTTGAATTGGCTGCAGAGTCTGATGTGTCCACAG CCATTGACCTTTTCAGACAAGCCGGCCTCGGCAATCATCTCTCTGGAAGTGAGCGGTT GACCCTCCTGGCTCCCCTGAATTCTGTATTCAAAGATGGAACCCCTCCAATTGATGCC CATACAAGGAATTTGCTTCGGAACCACATAATTAAAGACCAGCTGGCCTCTAAGTATC TGTACCATGGACAGACCCTGGAAACTCTGGGCGGCAAAAAACTGAGAGTTTTTGTTTA TCGTAATAGCCTCTGCATTGAGAACAGCTGCATCGCGGCCCACGACAAGAGGGGGAGG TACGGGACCCTGTTCACGATGGACCGGGTGCTGACCCCCCCAATGGGGACTGTCATGG ATGTCCTGAAGGGAGACAATCGCTTTAGCATGCTGGTAGCTGCCATCCAGTCTGCAGG ACTGACGGAGACCCTCAACCGGGAAGGAGTCTACACAGTCTTTGCTCCCACAAATGAA GCCTTCCGAGCCCTGCCACCAAGAGAACGGAGCAGACTCTTGGGAGATGCCAAGGAAC TTGCCAACATCCTGAAATACCACATTGGTGATGAAATCCTGGTTAGCGGAGGCATCGG GGCCCTGGTGCGGCTAAAGTCTCTCCAAGGTGACAAGCTGGAAGTCAGCTTGAAAAAC AATGTGGTGAGTGTCAACAAGGAGCCTGTTGCCGAGCCTGACATCATGGCCACAAATG GCGTGGTCCATGTCATCACCAATGTTCTGCAGCCTCCAGCCAACAGACCTCAGGAAAG AGGGGATGAACTTGCAGACTCTGCGCTTGAGATCTTCAAACAAGCATCAGCGTTTTCC AGGGCTTCCCAGAGGTCTGTGCGACTAGCCCCTGTCTATCAAAAGTTATTAGAGAGGA TGAAGCATTAGCTTGAAGCACTACAGGAGGAATGCACCACGGCAGCTCTCCGCCAATT TCTCTCAGATTTCCACAGAGACTGTTTGAATGTTTTCAAAACCAAGTATCACACTTTA

ATGTACATGGGCCGCACCATAATGAGATGTGAGCCTTGTGCATGTGGGGGAGGAGGGA

GAGAGATGTACTTTTTAAATCATGTTCCCCCTAAACATGGCTGTTAACCCACTGCATG

CAGAAACTTGGATGTCACTGCCTGACATTCACTTCCAGAGAGGACCTATCCCAAATGT

GGAATTGACTGCCTATGCCAAGTCCCTGGAAAAGGAGCTTCAGTATTGTGGGGCTCAT

AAAACATGAATCAAGCAATCCAGCCTCATGGGAAGTCCTGGCACAGTTTTTGTAAAGC

CCTTGCACAGCTGGAGAAATGGCATCATTATAAGCTATGAGTTGAAATGTTCTGTCAA

ATGTGTCTCACATCTACACGTGGCTTGGAGGCTTTTATGGGGCCCTGTCCAGGTAGAA

AAGAAATGGTATGTAGAGCTTAGATTTCCCTATTGTGACAGAGCCATGGTGTGTTTGT

AATAATAAAACCAAAGAAACATA

ORF Start: ATG at 48 ORF Stop: TAG at 2097

SEQ ID NO: 72 683 aa MW at 74680.0kD

NOV14a, MALFVRLliAlALAI ALGPAATI-AGPAKSPYQLVLQHSRLRGRQHGPNVCAVQKVIGTN CG95861-01 RKYFTNCKQWYQRKICGKSTVISYECCPGYE VPGEKGCPAALPLSNLYETLGWGST Protein TTQLYTDRTEKXiRPEMEGPGSFTIFAPSNEAWASLPAEVLDSLVSNVNIELLNALRYH Sequence 1WGRRVLTDELKHGMTLTS ΪQNSNIQIHHYPNGIVTVNCARLLKADHHATNGVVHLI DKVISTITNNIQQIIEIEDTFETLRAAVAASGLNTMLEGNGQYTLLAPTNEAFEKIPS ETLNRILGDPFJVLIsDLrjvraHILKSAMCAEAIVAGLSVETLEGTTLEVGCSGDMLTING

KAIISN DILATNGVIHYIDELLIPDSAKTLFELAAESDVSTAIDLFRQAGLGNHLSG

SERLTLIAPIJvTSVFKDGTPPIDAHTRNLLRNHIIKDQriASKYLYHGQTLETLGGKKLR

VFVYRNSLCIENSCIAAHDKRGRYGTLFTJ>©RVLTPP GTVMDVLKGDNRFSMLVAAI

QSAGLTET1-NREGVYTVFAPTNEAFRALPPRERSRLLGDAKELANILKYHIGDEILVS

GGIGAL LKSLQGDKLWSLKNNVVSVNKEPVAEPDIMATNGV^

PQERGDELADSALEIFKQASAFSRASQRSVRLAPVYQKLLERMKH

Further analysis of the NOVl 4a protein yielded the following properties shown in Table 14B.

Table 14B. Protein Sequence Properties NOV14a

Psort analysis: 0.8200 probability located in endoplasmic reticulum (membrane); 0.1900 probability located in plasma membrane; 0.1000 probability located in endoplasmic reticulum (lumen); 0.1000 probability located in outside

SignalP analysis: I Cleavage site between residues 24 and 25

A search of the NOVl 4a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 14C.

In a BLAST search of public sequence databases, the NOVl 4a protein was found to have homology to the proteins shown in the BLASTP data in Table 14D.

PFam analysis predicts that the NOVl 4a protein contains the domains shown in the Table 14E.

EXAMPLE 15.

The NOVl 5 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 15A.

Table 15A. NOV15 Sequence Analysis

SEQ ID NO: 73 8279 bp

NOVl 5a, AGTCGGGCCCCTCGGGGCCGCGTGGCCAATCAGATCCCCCCTGAGATCCTGAAGAACC CG96412-01 CTCAGCTGCAGGCAGCAATCCGGGTCCTGCCTTCCAACTACAACTTTGAGATCCCCAA DNA GACCATCTGGAGGATCCAACAAGCCCAGGCCAAGAAGGTGGCCTTGCAAATGCCGGAA Sequence GGCCTCCTCCTCTTTGCCTGTACCATTGTGGATATCTTGGAAAGGTTCACGGAGGCCG AAGTGATGGTGATGGGTGACGTGACCTACGGGGCTTGCTGTGTGGATGACTTCACAGC GAGGGCCCTGGGAGCTGACTTCTTGGTGCACTACGGCCACAGTTGCCTGAGTATGGTA GATCTTTCCTTTGGATTTGGTTGCCTTGGCAACGGTGCTCTGTCCCAGATGCAGGTGT TTGAAAGGCTGTTGGTTGTAGAGCAGGCTGGGCCCCGGCCGGTTCCCATGGACACCTC GGCCCAAGACTTCCGGGTGCTGTACGTCTTTGTGGACATCCGGATAGACACTACACAC CTCCTGGACTCTCTCCGCCTCACCTTTCCCCCAGCCACTGCCCTTGCCCTGGTCAGCA CCATTCAGTTTGTGTCATTCCTACAGGCAGCCGCCCAGGAGCTGAAAGCCGAGTATCG TGTGAGTGTCCCACAGTGCAAGCCCCTGTCCCCTGGAGAGATCCTGGGCTGCACATCC CCCCGACTGTCCAAAGAGGTGGAGGCCGTTGTATCCGCCGCGGCATTAGATTCTTGTA GGAGCTCAAACCCTATCGTGACCTGTACATGCGAGGGATCTAGGTTGCATGATCTTTA TGAGATCCTAATGCCTGATGATCTGAGGTATCTTGGAGATGGCCGCTTCCATCTGGAG TCTGTCATGATTGCCAACCCCAATGTCCCCGCTTACCGGTATGGGCTGGGCCGGGCTG GGCTGACCAGCTGGTATGACCCATATAGCAAAGTCCTATCCAGAGAACACTATGACCA CCAGCGCATGCAGGCTGCTCGCCAAGAAGCCATAGCCACTGCCCGCTCAGCTAAGTCC TGGGGCCTTATTCTGGGCACTTTGGGCCGCCAGGGCAGTCCTAAGATCCTGGAGCACC TGGAATCTCGACTCCGAGCCTTGGGCCTTTCCTTTGTGAGGCTGCTGCTCTCTGAGAT CTTCCCCAGCAAGCTTAGCCTACTTCCTGAGGTGGATGTGTGGGTGCAGGTGGCATGT CCACGTCTCTCCATTGACTGGGGCACAGCCTTCCCCAAGCCGCTGCTGACACCCTATG AGGTAACACCAAGCTCTGGGAGAGAGTGGGCTTTGGACGTGGTTCTCAAAGGCGGCCG TGGCTCTGAGGGACATTTCCTGGCAGCAGCCCTACCCGATGGACTTCTACGCTGGCAG CTCCTTGGGGCCCTGGACGGTGAACCACGGCCAGGACCGCCGTCCCCACGCCCCGGGC CGGCCCGCGCGGGGGAAGGAGGGGTCCGCGCGTCCCCCTTCGGCCGTGGCTTGCGAGG ACTGCAGCTGCAGGGACGAGAAGGTGGCGCCGCTGGCTCCTTGACGCGCTCCCGGGCC TCAGACCGCTTCCGGTGCTTCCGTCGCTCCTTGCCGGGCATAATGGCCGCGCAGCGAC CCCTGCGGGTCCTGTGCCTGGCGGGCTTCCGGCAGAGCGAGCGGGGCTTCCGTGAGAA GACCGGGGCGCTGAGGAAGGCGCTGCGGGGTCGCGCCGAGCTCGTGTGCCTCAGCGGC CCGCACCCGGTCCCCGACCCCCCGGGCCCCGAGGGCGCCAGATCAGACTTCGGGTCCT GCCCTCCGGAGGAGCAGCCTCGAGGCTGGTGGTTTTCAGAGCAGGAGGCCGACGTTTT CTCCGCATTGGAAGAGCCCGCCGTCTGCAGGGGCCTGGAGGAATCACTGGGGATGGTG GCACAGGCACTGAACAGGCTGGGGCCTTTTGACGGCCTTCTTGGTTTCAGCCAAGGGG CTGCGCTAGCAGCCCTTGTGTGTGCCCTGGGCCAGGCAGGCGATCCCCGCTTCCCCTT GCCACGGTTTATCCTCTTGGTGTCTGGTTTCTGTCCCCGGGGCATTGGGTTCAAGGAA TCCATCCTGCAAAGGCCCTTGTCATTGCCTTCGCTCCATGTTTTTGGGGACACTGACA AAGTCATCCCCTCTCAGGAGAGTGTGCAACTGGCCAGCCAATTTCCCGGAGCCATCAC CCTCACCCACTCTGGTGGCCACTTCATTCCAGCAGCTGCACCCCAGCGTCAGGCCTAC CTCAAGTTCTTGGACCAGTTTGCAGAGTGAAAGATCAAGAAATGTCTCTGCTCCTACA TCCAGCTCCTCTAGGGGCAGCCTCCGTCATCCATGCCCTCCCAGGACCCTCCACTCAC TGCTGTGAGTGCGCCTCACCAGAACCAGTTAAGAGACAACTATCAATTCTTGAGACCC AAATTATAAGGGCCCTGCCCTGTACTGAAGAAAAGGGGAGCACAAGGCCTTAATGGAC ATTGACTTGTGAAAACGCAAACATGAATATGGTTGGAGAGCCCTGGATTAGGAGGGTG ACATGGGGAAGGCAGAGGCTGGCACCATGGTGACTGCCACATAATAAAGTGGTGATTT GGATTTTGAGCATCTTTTTCCTGGTACACACAGAAAAACATTTTATAATGGAAGTCGG TTTCTTGGCCATCTACATAGTTTTCTGGGCAGCGCCAAGCAGGGAGGTGTCTCCAGTT GTGAGTCCTCGGACAGGCTGCTGCATGGGTGCACATACTCACGTTATTGGTGGAAGTT TTAAGTCCCAAACTGAAGGGAAGGAGGCCTAGGTGGCACAATCAGGGAGGAATATACA TCTGAGAAGTTTTGGGAAGACATCACCTGGCAAGGCTGCCTGAACCACAGTAATTTAG TCTTCCTCTATCCAGATCACTGAGAGTTGTGTGCTGGTCTTCTCTCAAACCCTGGATG

TGTTACCTGCTTCTCCCTAAGTGCTCTAACCTCGCTGAACTATGAGCCTGAAGGGTGG

GTCGATGCCAGACCTTAGGTGGAGGCCAAAGACAGTACAGAATAAACAGCTTCTTCCT

AAATTGACCCCATCTTGAGGGTTTCTGAGATTGATGCCATTCTTTAGTGACTACAGCC

AAGGCCTAAGCAATCCAGCTGGTTTTCCCCTTGGGGCGTGTAGTTGTTCTTGGATACT

TTGAGGATTGCTCACTCTTTTTTTTTTTTTTTTTGAGACAGGCTCACTCTGTCACCCA

GGCTGGAGAGCAGTGGCCAAGATCCTGGCTCACTGCAACCTCAGCCTCCTGGGTTCAA

GCAGTTCTGTCTCAGCCTCCTGAGTAGGTGGCGTTACAGGCATGCGCCACCACGCCTG

GCTAATTTCTATATTTTTTTGTAGAGATAGGGTTTTGCCACGTTGCCCAGGCTGGTCT

CAAAACTCTTTAGCTCAAACAATCCACCTGCCTCAGCCTCCCAAAGTGCTAGGATTAT

AGGCCTGAGCCACTGCACCTACCTAGGATTGCCCACTCTTTTTTTGTTTTCTTTACTT

TTAAAGACAAGGTCTGACTCTATCACCCAGGCTGGAGTGCAGTGGCACAATCTTGACA

GCCTCCACCTCCCGGGCTCAAGCCATCCATCCTCCCACCTCAGCCTCCCAAGCAGCTG

GGGCTGCAGGCGCAAATCACCATGCCTGGCTAATTTTTGTATTTGTTGTAGAGAAGGG

GTCTCACCATGTTGTGCGGATGGTCTCCAACTTGCGAGCTCAAGCAATCCACCTGTCC

TGGCCTCCCAAAGTGCTGGAATTAACAGGCGTGAGCCACCTCACCTGGCCAATTGCCC

ACTCTTAATGAGTTCCCTCCCATTTCCAGTGCCACCTGACACCCCCCACCCCCTGACA

CACACACACACACTCTTGTTTTCTGACTTCCTGCCTCTGCAAGACAAGAAGCCTGTTC

TTTTTCTGGAAGATCACACCAACTCCTGTTTAGCCCTCAGACATAGTTGCCAGGGAGC

TAAGCAACTGAGCAGGAATTATGGTCAGGGACACAGCACAAATACTCCAGCCCATTTT

TCTGACAACCAGACTAATATCCCTTACATTACCGAAAGTTCTGGGTTCCTCACTACAG AGATTAGCTGTCCAAAATTAAAAAAAACAAAACAAAACAGAATTATCCGCCCAGAGAC CTAGTTACTGGGTGTGGAAAGTGTGTGTGGGGAGGGGTGGGAGGTGGTTCTCCCTTGC TTCAGTTGGCTTCTGCCTTGTTTTAAGGAATCTTCTCCTTTAGCAAAAGGAGGAGACT TTGGAATGGATTGATTACACAGACTGTGGTATCTGACCCATTGAATTTTAGAAAAAAT TCTGATTTAAAGTTTCAGAAATTGCACAGAAAATTTCAGATTTCTGGCTTCTCTGGAA ATGATAGATTTGCAGTTCAGGGCTCCCATACCCTCATGGTAATCATAGGCTGCCCCCT TTAGCACGGTCCCAAGAGGTGAAAACGACTTGTACACCCCAGCTGCTTGGGATTATTG AGGACGAAAGTAGAGAGAAGGGGAGAAATATTGGGGAGTGAATTTTGAGCCAAGGCTT AAAATTAAAAGTGGGGGAGGTAGAGCCCAGCAGTAGTAGGTGGAGGAGAAGGGCTCCT GGCCGGGGGTGAGGATTCCTCCTGAGAACCATAGTGTCCAAGCTAGAGGGAAACATGG GGTCCACTAGTTCTCGACAAGTGGAACAGGTCACTTCCCATCATGCCGTCCAGGAGCA AGAGAGACTCTTAGGGGGTTGTGCCCCTCCCCACCCCCAATACGCAGATTCTGTGTAC CATTTACATCCAGAACATTGGTTAAAACCTGAATTCTGGCCCTGCGTGGTGGCTCATG CCTGTAATCCCAGCACTTTGGGAGGCGGAGGCGGGCAGATCACCTGAGGTTGGGAATT TGAGGCCAGCCAGACCAACAGAGAGAAACTCTGTCTCTACTAAAAATACAAAATTAGC CAACCATGGTGGCGCATGCCTGTAATCCCAGCTACTTGGAAGGCTGAAGCAGGAGAAT TGCTTGAACCTGGGAGGCAAAGGTTGCGGTAAGCCGAGATCACGCTATTGCACTCCAA CCTGGGCAACGAGAGCGAAACTGTCTCAAAAAAAAAAAAAAAAAAAAGTAGCTGGGCG TGGTGGCATGCACCTGTAATCCTAGCTACTTGGTAGGCTGAGGCAGGAGAATTGCTTG AACCCAGGAGGCGGAGGTTGCAGTGAGCCGAAATTGTGCCACTGCACTCCAGCCTGGG TGACAGAGTGAGACTCCATCTCAAAAΆACCAACTTGAATTCTGGGGCCAACTCCAGAT GTACTGAGTCAGAATCACTGGGACCCAGGAATCTGCATTTTGGCAAATTGCCCCCATC ATTCCCAAGCAGGGATTTGAAAAGCCACTGCCTTAGGGGTTGAATGAAAGCCTCGGGT GAGGGTGTGTTATCTGAAAAACCCCTAGTGTGACTCTCTTGCAACCCACCAGGGAGGG GAGCAAGCTCCCCAGAACAATCACTGCTTAAACATTTTCAATTTATAGCAAΆGGAAAA TTAGGTCTGAAGGATAGAAATAGGAATAGCATTTATTTATTTACTTACTTATTTGAGA CGGAGTCTCGCTCTGTCACCCAGGCTGGAGTGCAGTGGTACAGTCTCCGCTCTCTGCG ACCTCCGCCTCCTGAGTTCGAGCGATCCGCCTGCCTCAGCCTCCAGAGTAGCTGGGAT TACAGGTGTGCATCACCACTCCCAGCTAATTTTTGCATTTTCAGTAGAGACAGGGTTT CGCCATGTTGGCCAGGCTGGTCTTGAACTCCTGATCTCAAGTGATTCGCCCACCTCAG CCTCCCAAAGTGTTGGGATTACAGGTGTGAGCCACTGCACCCGGCCAGAAGAGCATTT ATTGAGTAATAACTGTATACCAGACATTGTAATAAGCAAGAGCTTTAATGCGGCCTAA TTTAGGGCCAGGTGCGATTGCTCATGCCTATAATCCCAGCACTTTGGGAGGCTAAGGC ATGAGGATCAATTGAGCCCAGGAGTTCAGAACCAGCCTGGGAAACATAGTGAGACCCC CTCTCTACAAATAATTTAAAAGTTAGCTGGGTGTGGTGGCATGCACCTGTGGTCCCAG CTACTCGGGAGGCTCAGGTGGGAGGATCACTTGAGCCCAGGAGTTTTGAGGCTGCAGT AAGCTGTGATTGTGCCACTGTATTCCAGCCTGGGAGACAGAGCTAGACCCTGTCTCCA AAAAAAAAGAAAAAAAAACAAACTACAAAAATTGGCTGGGGGTGGTGGCACACACCTG TAGTCCTAACTACTCAGGAGGCTGAGACAGGAGAATTGCTTGAACCTGGGAGGCGGAC

GTTGCAGTGAGCCGAGATCATGCTACTGCACTTCAGCCTGGGCAACAGAGCAAGACTC

CATCTCAGAAGAAAAAAACAAATAAAATTATATATATATAACACATATATATTTTATA

TATATTAGGCAATAAATATATATAGCTTAAACCTTACAACTTCATGTTTAAACTCACA

ACTTCATGAGGTAGGTACCATTATTATTATTATTATTTGAGACGGAGTGTCACTCAGC

CACCCAGGCTGGAGTTCAGTGGCACGATCACGGCTCACTGCACCTCCGTCTCCTGGGT

TCAAGTGATTCTAGTGCCTCAGCCTCCCAAGCAGCTGGAATTACTGGCAAGCACCACC

ATGGCTGGCTAATTTTTTGTGTTATTAGTAGAGACGGAGTCTGACCATATTGGCCAGG

CTGGATTATTATTAAGACAGGGTCTACCTCTGTCACCAGGTGCAGGATCATGGCTCAC

TGCAGCCTCGACCTCTTCGGCTCAAGTGACCCTCCCGCTTCAGCCTCCTGAGTAGCTT

AGCTGAGACTACAGGTGCCACCTATCACACTGGCTTTTTTTTTTTTAAGACATGGGGT

CTCACTATGTTGCCCAGGCTGGTCTCGAATTTCTGGGCTCAAGCAGTCCTCCAGCCTC

TGCCCCGCCCCGCAAAATGGTGTGATTACAGGCATGAGCCATGTCTCCCAGCCCTAAA

TAATGATTTTTTTTTTTTGAGACAGGGTCTCACTCTGTCATCCAGGCTGGAGTACAGT

GGTGCAATCACAGCTCACTGCAGCCTCAACCTCCCAGCCTCAAGCAATCCTCCCACCT

CAACTTCCCAAGTAGCTGGGACTACAGGTGTGCACAACTAATTTTCTTTCTTTTCTTT

CCTTTCTTCTTTCCTTTCTTTCTTTTTTGTAGAGATGGGAGTCGCATTGTATTGCCCA

GGCTGGTCTCGAACTCCTGGGCTCAAGAGATCCTCCCACCTATGCCTCCCAAAGTGCT

GGGATTACAGGCGTGAGCCACTGTGCCCTGCCCTAAATGATTTCTAAGATGCTAAACA

ACCCTGGCGTTCGGTCAAACCTCCGGTTCTTTCCCCCCCTACCACATTCGTGACCTTT

TCTCAAATAATCCCAACCATTCTGTTTTTCCACCTCTGATTCCACCTTGGATTCCTAG

AGCTCTAGAGAGAGAACCCCCTTAGCAAGAACCTCTCACAGCCCCAGGCCCAGAGTGA

ACCCGGAAGGCGGAGGTTGCAGTGAGCCGAGATAGTGCCACTGCACTCCAGCCTGGGC

GACAGAGCGAGACTGTCTCAGAAGAAAAAAAATAAAAGTATATGTATGTATAACATAT

ATATAATATAGATTATATATATATTAGCCAATAAATATATATAGCCTAAAGCTCAAGG

TGTACCCCCCACATCCCGTTCATTCTGCCTAAGTGCACAGAGCTGGGGTCCCCTCCCA

ACACCACCCCTTCACAAACATGTAGTAGCCTAACAGAATACTTAAGATCAAAATAACA

TTCTGCTGTTTTTGGATTTTTACTTACATGATCCTATTTAATCCTCACAGTGCACCTG

CCGAGGAGAAGAGATTATTATCCCGTTTTACAGAAAGCTCAGAAACAGAATATGGCCC

CTTAACCACCAGTTGCTGATAATGCCAGGTTGGACCTCTGGCTTCCCAGAGCACTACA

CCCAGCAGTCCACTGCAGTCCACCCACTGACATCCCCTGGAACCCAGAAGCAAGTGCG

TTTTAATCTGCAAGACCTCGGGGCCCTGGGGAGGTGGGATGGCTAGCATGTGGGTGTT

GATTAACTGGGAAACCGGCACGAGTGTCTTAGAAGTACTTCACAAAGGAGCCGGGTGG

GGAGTGAAGGAGGGGGTGGGGCGTGAGACGTTAAGAAAAATTG

ORF Start: ATG at 166 ORF Stop: TGA at 2290

SEQ ID NO: 74 708 aa MW at 76785.2kD

NOVl 5a, PEGLLLFACTIVDILERFTEAEVMV GDVTYGACCVDDFTARALGADFLVHYGHSCL CG96412-01 SMVDLSFGFGCLGNGALSQMQVFERLLWEQAGPRPVPMDTSAQDFRVLYVFVDIRID Protein TTHLLDSLRLTFPPATALALVSTIQFVSFLQAAAQELKAEYRVSVPQCKPLSPGEILG Sequence CTSPRLSKEVΞAWSAAALDSCRSSNPIVTCTCEGSRLHDLYΞILMPDDLRYLGDGRF HLESVMIANPNVPAYRYGLGRAGLTS YDPYSKVLSREHYDHQRMQAARQEAIATARS A S GLILGTLGRQGSPKILEHLESRLRALGLSFVRLLLSEIFPSKLSLLPΞVDV VQ VACPRLSIDWGTAFPKPLLTPYEVTPSSGRE ALDWLKGGRGSEGHFLAAALPDGLL RWQLLGALDGEPRPGPPSPRPGPARAGEGGVRASPFGRGLRGLQLQGREGGAAGSLTR SRASDRFRCFRRSLPGIMAAQRPLRVLCLAGFRQSERGFREKTGALRKALRGRAELVC LSGPHPVPDPPGPEGARSDFGSCPPEEQPRGWWFSEQEADVFSALEEPAVCRGLEESL GMVAQAI-NRLGPFDGLLGFSQGAALAALVCALGQAGDPRFPLPRFILLVSGFCPRGIG FKESILQRPLSLPSLHVFGDTDKVIPSQESVQLASQFPGAITLTHSGGHFIPAAAPQR QAYLKFLDQFAE

SEQ ID NO: 75 987 bp

NOV15b, ATGCCGGAAGGCCTCCTCCTCTTTGCCTGTACCATTGTGGATATCTTGGAAAGGTGTG CG96412-03 TGGATGACTTCACAGCGAGGGCCCTGGGAGCTGACTTCTTGGTGCACTACGGCCACAG DNA TTGCCTGGTTCCCATGGACACCTCGGCCCAAGACTTCCGGGTGCTGTACGTCTTTGTG Sequence GACATCCGGATAGACACTACACACCTCCTGGACTCTCTCCGCCTCACCTTTCCCCCAG CCACTGCCCTTGCCCTGGTCAGCACCATTCAGTTTGTGTCGACCTTGCAGGCAGCCGC CCAGGAGCTGAAAGCCGAGTATCGTGTGAGTGTCCCACAGTGCAAGCCCCTGTCCCCT GGAGAGATCCTGGGCTGCACATCCCCCCGACTGTCCAAAGAGGTGGAGGCCGTTGTGG

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 15B.

Further analysis of the NOVl 5a protein yielded the following properties shown in Table 15C.

Table 15C. Protein Sequence Properties NOV15a

PSort 0.7300 probability located in plasma membrane; 0.6400 probability located in analysis: endoplasmic reticulum (membrane); 0.2279 probability located in microbody (peroxisome); 0.1000 probability located in endoplasmic reticulum (lumen)

SignalP Cleavage site between residues 23 and 24 analysis:

A search of the NOVl 5a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 15D.

In a BLAST search of public sequence databases, the NOVl 5a protein was found to have homology to the proteins shown in the BLASTP data in Table 15E.

PFam analysis predicts that the NOVl 5a protein contains the domains shown in the Table 15F.

EXAMPLE 16.

The NOVl 6 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 16 A.

Table 16A. NOV16 Sequence Analysis

SEQ ID NO: 81 1492 bp

NOVlδa, ATGACCTGGGTACTCAGGACTCCTCCAGCACTCCTCCTGCTAGCTGTCATTGTTCTGG CG96511-01 GCACATTTGGTAAGCCAACTGTGGCTTATGCTGAACTCCGCTGCATGTGTATAAAGAC DNA Sequence AACCTCTGGAATTCATCCCAAAAACATCCAAAGTTTGGAAGTGATCGGGAAAGGAACC CATTGCAACCAAGTCGAAGTGATGGCCACACTGAAGGATGGGAGGAAAATCTGCCTGG ACCCAGATGCTCCCAGAATCAAGAAAATTGTAGGCTCTGTTTCCACAACCTTTCCCCA TTCTCTTGGCAGAGAACAGTCCCCACCTTCTTCAGCACTTATTTATATGCAGATACAA GAGCCAAAGTCATTCTTTCTCAGAGAAGGGAAGACTGTCCATATGGATCTCTTCCCCT TTCCTGCCAAAGTGGTCTCAGGGGATTTTTCTGACTTGGCCAATTCCAGCTCCATCAA TTTCCTGCGTGACTTCAGCCAAGTCACTCAACCTCTCTTAAACTCTATAATCTTATCC AGAAAAATGGGCAAAAATAGCTGTTGGGAGGATTAGGTAAATATAATTCATAAAACAC ACTTGGAACAAAGTAGTTGTCATGAAATATTAGTTGTTATTATATATGATAAGGTGCA

GTGGGCTAGTGTGGAATTATGAGAGAAAAAGAAAAAAAAGAGATTTAGAAAACTCAAG ACATTGAGCTTATTCTGCCACAAATGTTTAGATAAGGAAGAGCTTTCTTGTCTGTTAA

AATACTTATTTAGAGTAAAGGGCAAGTACAATACCAACAACTATAATCTTTCTAGTGT TGACCTAGCTAGACCTTTATATTTCATGAACAGAGGTAAGATTAATCAGATGAAATGC

CAAATGAAAAGAGGAAGACAGTTGAGTAAAACAAAAAGTGTGATTTAGCAGAGGATTT

AGTGCGTCAAGTTATACCACATCCTGTTAGTCTTTTATGCCGTTATCCAGTGTTGTGT

GTCCCAGCAGCATAACAAAGGTTAAGAACAAGTTTATTCAACATCAGCAATGATTGCC

AGAGGGAAGCAACTCACTTTAGTCAACATGGCTATCAAATGACAAAGAAAGGCACAAG

TGAGAAATCTGTTTACAACCCTGGCATTTGGAGACACAACAGATGAATTATAGCATTT

TGTAGATATTGAATTATATTTTTACATTCACTCACAATACTTGGAGTGAAGAGATCAA

AAATTCTGTTAGAACAAATTGTAATGCATTTTCTCTCTTGTTTCTATTTTTCCCCACT

TCTCCAATATACTTACATACTTTTAATAGGTCGGTAAATCTCCTAAATATTAGCTCAC

ATCCAATCATACTGTTTAGAGAGCTTCTATGATTCATAAACATGCCTGAATGGATTTG

GAGCGAATTCACAATTTCAGTATTTACCAAAATGTTAAATACTATGATTTGTATCACA

GGTATCAACAGAGCTAAAGTAGCCAGATTAGTAAAACTTCTC

ORF Start: ATG at 1 ORF Stop: TAG at 556

SEQ ID NO: 82 185 aa MW at 20434.7kD

NOVlδa, MTVITV^IRTPPALLLI-AVIVLGTFGKPTVAYAELRCMCIKTTSGLHPKNIQSLEVIGKGT CG96511-01 HCNQVEVMATLKΠGRKICLDPDAPRIKKIVGSVSTTFPHSLGREQSPPSSALIY QI Protein EPKSFFLREGKTVHMDLFPFPA VVSGDFSDLALVTSSSINFLRDFSQVTQPLLNSIILS Sequence RKMGKNSCWED

Further analysis of the NOVl 6a protein yielded the following properties shown in

Table 16B.

Table 16B. Protein Sequence Properties NOV16a

PSort analysis: 0.7427 probability located in outside; 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen); 0.1000 probability located in lysosome (lumen)

SignalP analysis: Cleavage site between residues 24 and 25 A search of the NOVl 6a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 16C.

In a BLAST search of public sequence databases, the NOVlδa protein was found to have homology to the proteins shown in the BLASTP data in Table 16D.

PFam analysis predicts that the NOVl 6a protein contains the domains shown in the Table 16E.

EXAMPLE 17.

The NOVl 7 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 17 A.

Table 17A. NOV17 Sequence Analysis

SEQ ID NO: 83 3030 bp

NOVl 7a, CCTGGCTCTCCCCTATGGTTCTTCTTTCTTTAGCTTTGGCCCCGGGGGCCACGGCAGA CG96522-01 CCTGGCAGGGCGGGCGGCACAGGGGAGAGGCAGGAAGAAACTGGATCCCTGGGGACCT DNA Sequence GTGGTAGGGTCGGCAGGAAGAAACTGGATCCCTGGGGACCTGTGGCCACGGGCCCTCC CTGAGCACCGCGCGCAAAGGCCCGGCCCCAGGGCCAGGCAACTCCAGCGCCGAGGCCG TCCAGTGCGGGGCCAGGCCCCGGGGGTGCCCTGCTGCACCGACTCCGGGCGGAGATTG GCTGTCCTCGGCCCGCAATGCTGTGCGCCCCGCCCCGCGCGCCGCTTCCGGCGGAGTC AGGCTTGGCTAATCGAGCGCGCGGGCTGGCGGCTCGGCGTTCGTTTGGCCGCGCCTGC CCGTGTGGTGGTTTCCGGCGGAGGTGGTGAGAGCCGGCGGGCAGGTGGGCTTGGCCGC GCTGTGGGTGCCTGGGACCCGCAGGGAGGATGGGCGCGGTGGCGCGGCCTGGCGGGGG GCTCGTCTCCGGGGTCCCCGGGTCCTGGTGAGAGCGGGGTCCCTCGACGCCGTGGCGG TCTCGAACCTGTGGATCTGAGGAGGGGATGCACACACAGCAGCCAGCCCAGTGTGGTG CCGAGAAACAGAGCCCCGAGGCCCTGGTCCTCAGAAAGGTCCCTCCCCTGCCTTCCTG TCCCTGCAGAGGTCATGCAGAAATTCTCTGGCTGGCCCGAAGTCCAGCTCAGGGCCAT GAAGAGGCTTGTGGCCGTGGGCCCCGATGTCTTCCAGGCTCACCAGGAGGACACAGAG CGCTATGTGCTCACCAACCTCAACATCGGGGCAGAACTGCTTCGGGACCCGTCCCTGG GGGCTCAGTTTCGGGTGCACCTGGTGAAGATGGTCATTCTGACAGAGCCTGAGGGTGC CCCAAATATCACAGCCAACCTCACCTCGTCCCTGCTGAGCGTCTGTGGGTGGAGCCAG ACCATCAACCCTGAGGACGACACGGATCCTGGCCATGCTGACCTGGTCCTCTATATCA CTAGGAGGTTTGACCTGGAGTTGCCTGATGGTAACCGGCAGGTGCGGGGCGTCACCCA GCTGGGCGGTGCCTGCTCCCCAACCTGGAGCTGCCTCATTACCGAGGACACTGGCTTC GACCTGGGAGTCACCATTGCCCATGAGATTGGGCACAGGTATGTAGCCCCACCAGCTG TCCCCAGGATCTGGCAAGGAGCTGACCTGGGTACCCAGGGTGGAGGTGGTCTTAGCAA GCAGTGGGTCCTTGTAGAGTTTCTCCAGAGGAGCCTGTACCCCTCACCCCGACAGACT CAGGTGAGCTTCGGCCTGGAGCACGACGGCGCGCCCGGCAGCGGCTGCGGCCCCAGCG GACACGTGATGGCTTCGGACGGCGCCGCGCCCCGCGCCGGCCTCGCCTGGTCCCCCTG CAGCCGCCGGCAGCTGCTGAGCCTGCTCGGACGGGCGCGCTGCGTGTGGGACCCGCCG CGGCCTCAACCCGGGTCCGCGGGGCACCCGCCGGATGCGCAGCCTGGCCTCTACTACA GCGCCAACGAGCAGTGCCGCGTGGCCTTCGGCCCCAAGGCTGTCGCCTGCGATATGTG CCAGGCCCTCTCCTGCCACACAGACCCGCTGGACCAAAGCAGCTGCAGCCGCCTCCTC GTTCCTCTCCTGGATGGGACAGAATGTGGCGTGGAGAAGTGGTGCTCCAAGGGTCGCT GCCGCTCCCTGGTGGAGCTGACCCCCATAGCAGCAGTGCATGGGCGCTGGTCTAGCTG GGGTCCCCGAAGTCCTTGCTCCCGCTCCTGCGGAGGAGGTGTGGTCACCAGGAGGCGG CAGTGCAACAACCCCAGGTACCGCAGGGAGGGTGCTTTTCTGTCAGGGTGTCCTGGGG GGAAGCCGGAAGTGAGTCACAGTCAGCTCTTCCGAGCCTCCAGTGTGCACGCCTGTAA GCTGGGATCGGTCCTCAGCGATGTCCATCAGTGCAGACACATGTGCCGGGCCATTGGC GAGAGCTTCATCATGAAGCGTGGAGACAGCTTCCTCGATGGGACCCGGTGTATGCCAA GTGGCCCCCGGGAGGACGGGACCCTGAGCCTGTGTGTGTCGGGCAGCTGCAGGGTAGG CGGCTGTGATGGTAGGATGGACTCCCAGCAGGTATGGGACAGGTGCCAGGTGTGTGGT GGGGACAACAGCACGTGCCACGGCGTCGAGGGACCCCGCTCTCACCAGGACCCGGGGA CCCCGGAGACGAGCCCCCCGCCAGGCCGCGCCACCGCGCCCATCCTCCCTGCGGGTCC CAGGCAGGCTTGCGGCACTGGCCAGATGTGGGCATCGAGGGGGCAGGTGCGGAATG C ACCACCTCTCCCATACCCGCAAGGCCGATCTGCCTTCAGCTCCCAGCAAGTGTGGGGC AGCGCGGGCCACAGAGTAGGGTGCAGGGATGGGGCCCCGGGAGGAGCAGGCCCACCTC ACTGAACTCTATCCCAGACAGCCTGCCTAGCACAACACAGGGAGGCCCCCAAATGGCT CATTCCTCAGCCATCAGCAGCAGCCTGCATAGAGGACACTGGGGTTACCAGGGGATGG TTACCTGGTCCCCAAATCACCTTGTTGTGGCAAGTGCCAGAATTCCTAAGCCACGGCA GGCCTGGGTGTGGGCCGCTGTGCGTGGGCCCTGCTCGGTGAGCTGTGGGGCAGGTGAG ACCTGGGGAAGGCTCATCCACAGCACGGCTTGCGTGGAGGCCCAGGGCAGCCTCCTGA AGACATTGCCCCCAGCCCGGTGCAGAGCAGGGGCCCAGCAGCCAGCTGTGGCGCTGGA AACCTGCAACTCTGAGTCTCAATTTCCCATCTGTGAAATGGAGATAATAGCAGTAGGT

Further analysis of the NOVl 7a protein yielded the following properties shown in

Table 17B.

Table 17B. Protein Sequence Properties NOVl 7a

PSort analysis: 0.5500 probability located in lysosome (lumen); 0.3700 probability located in outside; 0.1132 probability located in microbody (peroxisome); 0.1000 probability located in endoplasmic reticulum (membrane)

SignalP analysis: Cleavage site between residues 15 and 16

A search of the NOVl 7a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 17C.

In a BLAST search of public sequence databases, the NOVl 7a protein was found to have homology to the proteins shown in the BLASTP data in Table 17D.

PFam analysis predicts that the NOVl 7a protein contains the domains shown in the Table 17E.

EXAMPLE 18.

The NOVl 8 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 18A.

Table 18A. NOV18 Sequence Analysis

SEQ ID NO: 85 1103 bp

NOV18a, ATGCATTAGGAAGATCCTGGACCTAGAGAACAAGTCCCCCGAACGCTGAGTTGGAGGC CG96535-01 GGGACTTCGGGTGCGCGTTGGCGGGAGCATGCTGGGGCTCTGGGGGCAGCGGCTCCCC DNA Sequence GCGGCGTGGGTCCTGCTTCTGTTGCCTTTCCTGCCGCTGCTGCTGCTTGCAGCCCCCG CGCCCCACCGCGCGCCCTACAAGCCGGTCATCGTGGTGCATGGGCTCTTCGACAGCTC GTACAGCTTCCGCCACCTGCTGGAATACATCAATGAGACACACCCCGGGACTGTGGTG ACAGTGCTCGATCTCTTCGATGGGAGAGAGAGCTTGCGACCCCTGTGGGAACAGGTGC AAGGGTTCCGAGAGGCTGTGGTCCCCATCATGGCAAAGGCCCCTCAAGGGGTGCATCT CATCTGCTACTCGCAGGGGGGCCTTGTGTGCCGGGCTCTGCTTTCTGTCATGGATGAT CACAACGTGGATTCTTTCATCTCCCTCTCCTCTCCACAGATGGGACAGTATGGAGACA CGGACTACTTGAAGTGGCTGTTCCCCACCTCCATGCAGTCTAACCTCTATCGGATCTG CTATAGCCCCTGGGGCCAGGAATTCTCCATCTGCAACTACTGGCATGATCCCCACCAC GATGACTTGTACCTCAATGCCAGCAGCTTCCTGGCCCTGATCAATGGGGAAAGAGACC ATCCCAATGCCGCAGTATGGCGGAAGAACTTTCTGCGTGTGGGCCACCTGGTGCTGAT TGGGGGCCCTGATGATGGTGTTATTACTCCCTGGCAGTCCAGCTTCTTTGGTTTCTAT GATGCAAATGAGACCGTCCTGGAGATGGAGGAGCAACTGCCTGCCAGGCCCACCCACC AGTCTGAGCTGCTTCTGCTGAGGCTGGTCTGCTTGAAGCCTCCCAGGAGAAAGAAGCC AGGTGGGAATGGAGAGAGAGAGGAAGCCTGTAGGGTCCAGCGTCAAAGCGAATCATGG GGCCCAGGGCTGAGCTGTGCACTCTCTTAGGCGGATTCTCCTTCCTCCTGCTACTGAT ACCAGGCGAGGGGGCCAAGGGTGGATCCCTCAGAGAGAGGTGACAACAGAGGGGGTAG

ORF Start: ATG at 87 ORF Stop: TAG at 1014

SEQ ID NO: 86 309 aa MW at 34918.7kD

NOVl 8a, MLGL GQRLPAA VLLLLPFLPLLLLAAPAPHRAP YKPVI WHGLFDS SYS FRHLLE Y CG96535-01 INETHPGTVVTVIJDLFDGRESLRPLWEQVQGFREAVVPIMZVKAPQGVHLICYSQGGLV Protein CRAL SV roDH VDSFIS SSPQ GQ GDTDYL- W FP SMQS LYRICYSP GQEFS Sequence ICNY^HDPHHDDLYI-NASSFLALINGERDHPNAAVWRKNFLRVGHLVLIGGPDDGVIT PWQSSFFGFYDANETVLEMEEQLPARPTHQSELLLLRLVCLKPPRRKKPGGNGERΞEA CRVQRQSESWGPGLSCALS

SEQ ID NO: 87 1103 bp

NOVl 8b, ATGCATTAGGAAGATCCTGGACCTAGAGAACAAGTCCCCCGAACGCTGAGTTGGAGGC CG96535-02 GGGACTTCGGGTGCGCGTTGGCGGGAGCATGCTGGGGCTCTGGGGGCAGCGGCTCCCC DNA Sequence GCGGCGTGGGTCCTGCTTCTGTTGCCTTTCCTGCCGCTGCTGCTGCTTGCAGCCCCCG CGCCCCACCGCGCGTCCTACAAGCCGGTCATCGTGGTGCATGGGCTCTTCGACAGCTC GTACAGCTTCCGCCACCTGCTGGAATACATCAATGAGACACACCCCGGGACTGTGGTG ACAGTGCTCGATCTCTTCGATGGGAGAGAGAGCTTGCGACCCCTGTGGGAACAGGTGC AAGGGTTCCGAGAGGCTGTGGTCCCCATCATGGTAAAGGCCCCTCAAGGGGTGCATCT CATCTGCTACTCGCAGGGGGGCCTTGTGTGCCGGGCTCTGCTTTCTGTCATGGATGAT CACAACGTGGATTCTTTCATCTCCCTCTCCTCTCCACAGATGGGACAGTATGGAGACA CGGACTACTTGAAGTGGCTGTTCCCCACCTCCATGCGGTCTAACCTCTATCGGATCTG CTATAGCCCCTGGGGCCAGGAATTCTCCATCTGCAACTACTGGCATGATCCCCACCAC GATGACTTGTACCTCAATGCCAGCAGCTTCCTGGCCCTGATCAATGGGGAAAGAGACC ATCCCAATGCCGCAGTATGGCGGAAGAACTTTCTGCGTGTGGGCCACCTGGTGCTGAT TGGGGGCCCTGATGATGGTGTTATTACTCCCTGGCAGTCCAGCTTCTTTGGTTTCTAT GATGCAAATGAGACCGTCCTGGAGATGGAGGAGCAACTGCCTGCCAGGCCCACCCACC AGTCTGAGCTGCTTCTGCTGAGGCTGGTCTGCTTGAAGCCTCCCAGGAGAAAGAAGCC AGGTGGGAATGGAGAGAGAGAGGAAGCCTGTAGGGTCCAGCGTCAAAGCGAATCATGG GGCCCAGGGCTGAGCTGTGCACTCTCTTAGGCGGATTCTCCTTCCTCCTGCTACTGAT

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 18B.

Table 18B. Comparison of NOV18a against NOV18b.

NOV18a Residues/ Identities/

Protein Sequence Match Residues Similarities for the Matched Region

NOVl 8b 1.309 266/309 (86%) 1.309 267/309 (86%)

Further analysis of the NOVl 8a protein yielded the following properties shown in Table 18C.

Table 18C. Protein Sequence Properties NOV18a

PSort analysis: 0.8200 probability located in outside; 0.6850 probability located in plasma membrane; 0.4882 probability located in lysosome (lumen); 0.1370 probability located in microbody (peroxisome)

SignalP analysis: Cleavage site between residues 28 and 29

A search of the NOVl 8a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 18D.

In a BLAST search of public sequence databases, the NOVl 8a protein was found to have homology to the proteins shown in the BLASTP data in Table 18E.

PFam analysis predicts that the NOVl 8a protein contains the domains shown in the Table 18F.

Table 18F. Domain Analysis of NO lSa

Identities/

Pfam Domain NOV18a Match Region Similarities Expect Value for the Matched Region

Palm thioest 177..242 26/66 (39%) 4. le-07 41/66 (62%) EXAMPLE 19.

The NOVl 9 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 19A.

Further analysis of the NOVl 9a protein yielded the following properties shown in Table 19B.

Table 19B. Protein Sequence Properties NOVl 9a

SignalP analysis: j Cleavage site between residues 32 and 33

A search of the NOVl 9a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 19C.

In a BLAST search of public sequence databases, the NOVl 9a protein was found to have homology to the proteins shown in the BLASTP data in Table 19D.

PFam analysis predicts that the NOVl 9a protein contains the domains shown in the Table 19E.

Table 19E. Domain Analysis of NOV19a

Identities/

Pfam Domain NOV19a Match Region Similarities Expect Value for the Matched Region

EXAMPLE 20.

The NOV20 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 20A.

Sequence compa son of t e above protein sequences yie s the o ow ng sequence relationships shown in Table 20B.

Table 20B. Comparison of NOV20a against NOV20b.

NOV20a Residues/ Identities/

Protein Sequence Match Residues Similarities for the Matched Region

NOV20b 1..116 72/120 (60%) 1..92 78/120 (65%) Further analysis of the NOV20a protein yielded the following properties shown in Table 20C.

Table 20C. Protein Sequence Properties NOV20a

PSort 0.7141 probability located in outside; 0.1000 probability located in endoplasmic analysis: reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen); 0.1000 probability located in lysosome (lumen)

SignalP Cleavage site between residues 22 and 23 analysis:

A search of the NOV20a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 20D.

In a BLAST search of public sequence databases, the NOV20a protein was found to have homology to the proteins shown in the BLASTP data in Table 20E.

PFam analysis predicts that the NOV20a protein contains the domains shown in the Table 20F.

EXAMPLE 21.

The NOV21 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 21 A.

GCTTCCTGGAGGTGTCGTACCGCGTTCTACGCCACCTTGCCCAGCCCTGAGCCAAGCC CTCCCCATCCCATGTATTTATCTCTATTTAATATTTATGTCTATTTAAGCCTCATATT

TAAAGACAGGGAAGAGCAGAACGGAGCCCCAGGCCTCTGTGTCCTTCCCTGCATTTCT

GAGTTTCATTCTCCTGCCTGTAGCAGTGAGAAAAAGCTCCTGTCCTCCCATCCCCTGG

ACTGGGAGGTAGATAGGTAAATACCAAGTATTTATTACTATGACTGCTCCCCAGCCCT

GGCTCTGCAATGGGCACTGGGATGAGCCGCTGTGAGCCCCTGGTCCTGAGGGTCCCCA

CCTGGGACCCTTGAGAGTATCAGGTCTCCCACGTGGGAGACAAGAAATCCCTGTTTAA

TATTTAAACAGCAGTGTTCCCCATCTGGGTCCTTGCACCCCTCACTCTGGCCTCAGCC

GACTGCACAGCGGCCCCTGCATCCCCTTGGCTGTGAGGCCCCTGGACAAGCAGAGGTG

GCCAGAGCTGGGAGGCATGGCCCTGGGGTCCCACGAATTTGCTGGGGAATCTCGTTTT

TCTTCTTAAGACTTTTGGGACATGGTTTGACTCCCGAACATCACCGACGTGTCTCCTG

TTTTTCTGGGTGGCCTCGGGACACCTGCCCTGCCCCCACGAGGGTCAGGACTGTGACT

CTTTTTAGGGCCAGGCAGGTGCCTGGACATTTGCCTTGCTGGACGGGGACTGGGGATG

TGGGAGGGAGCAGACAGGAGGAATCATGTCAGGCCTGTGTGTGAAAGGAAGCTCCACT

GTCACCCTCCACCTCTTCACCCCCCACTCACCAGTGTCCCCTCCACTGTCACATTGTA

ACTGAACTTCAGGATAATAAAGTGTTTGCCTCCAAAAAAAAAAAAAAA

ORF Start: ATG at 193 ORF Stop: TGA at 802

SEQ ID NO: 100 203 aa|MW at 21858.0kD

NOV21c, MSPEPALSPALQLLL HSALWTVQEATPLGPASSLPQSFLLKCLEQVRKIQGDGAALQ CG97274-04 EKLVSECATYKLCHPEELVLLGHSLGIP APLSSCPSQALQLAGCLSQLHSGLFLYQG Protein LLQALEGISPELGPTLDTLQLDVADFATTIWQQMEELGMAPALQPTQGAMPAFASAFQ Sequence RRAGGVLVASHLQSFLEVSYRVLRHLAQP

SEQ ID NO: 101 426 bp

NOV21d, GGATCCACCCCCCTGGGCCCTGCCAGCTCCCTGCCCCAGAGCTTCCTGCTCAAGTGCT 197208289 TAGAGCAAGTGAGGAAGATCCAGGGCGATGGCGCAGCGCTCCAGGAGAAGCTGGCAGG DNA Sequence CTGCTTGAGCCAACTCCATAGCGGCCTTTTCCTCTACCAGGGGCTCCTGCAGGCCCTG GAAGGGATCTCCCCCGAGTTGGGTCCCACCTTGGACACACTGCAGCTGGACGTCGCCG ACTTTGCCACCACCATCTGGCAGCAGATGGAAGAACTGGGAATGGCCCCTGCCCTGCA GCCCACCCAGGGTGCCATGCCGGCCTTCGCCTCTGCTTTCCAGCGCCGGGCAGGAGGG GTCCTAGTTGCCTCCCATCTGCAGAGCTTCCTGGAGGTGTCGTACCGCGTTCTACGCC ACCTTGCCCAGCCCCTCGAG

ORF Start: at 1 ORF Stop: end of sequence

SEQ ID NO: 102 142 aaiMW at l51993kD

NOV21d, GSTPLGPASSLPQSFLLKCLEQVRKIQGDGAALQEKLAGCLSQLHSGLFLYQGLLQAL 197208289 EGISPELGPTLDTLQLDVADFATTI QQMEELGMAPALQPTQGAMPAFASAFQRRAGG Protein VLVASHLQSFLEVSYRVLRHLAQPLE Sequence

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 2 IB.

Table 21B. Comparison of NOV21a against NOV21b through NOV21d.

NOV21a Residues/ Identities/

Protein Sequence Match Residues Similarities for the Matched Region

NOV21b 21..157 137/137 (100%) 1..137 137/137 (100%)

NOV21c 1..158 158/197 (80%)

7..203 158/197 (80%)

NOV21d 20..158 138/139 (99%) 2..140 139/139 (99%)

Further analysis of the NOV2 la protein yielded the following properties shown in Table 21 C.

Table 21C. Protein Sequence Properties NOV21a

PSort analysis: j 0.5567 probability located in microbody (peroxisome); 0.4273 probability j located in mitochondrial matrix space; 03175 probability located in ] lysosome (lumen); 0.1052 probability located in mitochondrial inner 1 membrane

SignalP analysis: j Cleavage site between residues 21 and 22

A search of the NOV2 la protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 2 ID.

In a BLAST search of public sequence databases, the NOV2 la protein was found to have homology to the proteins shown in the BLASTP data in Table 2 IE.

PFam analysis predicts that the NOV21a protein contains the domains shown in the Table 2 IF.

EXAMPLE 22.

The NOV22 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 22 A.

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 22B.

Table 22B. Comparison of NOV22a against NOV22b.

NOV22a Residues/ Identities/

Protein Sequence Match Residues Similarities for the Matched Region

NOV22b 1..82 62/82 (75%) 1..82 62/82 (75%)

Further analysis of the NOV22a protein yielded the following properties shown in Table 22C. Table 22C. Protein Sequence Properties NOV22a

PSort analysis: 0.6000 probability located in endoplasmic reticulum (membrane); 0.5994 probability located in mitochondrial inner membrane; 0.3647 probability located in mitochondrial intermembrane space; 0.1802 probability located in mitochondrial matrix space

SignalP analysis: Cleavage site between residues 35 and 36

A search of the NOV22a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 22D.

Table 22D. Geneseq Results for NOV22a

Identities/

NOV22a

Similarities

Geneseq Protein/Organism/Length Residues/ Expect for the Identifier [Patent #, Date] Match Value

Matched Residues

Region

AAB15804 Human chemokine PF4 SEQ ID NO: 1..132 106/137 (77%)) 4e-43 46 - Homo sapiens, 128 aa. 1-128 109/137 (79%) [WO200042071-A2, 20-JUL-2000]

AAW96716 A platelet basic protein (PBP) - Homo 1..132 106/137 (77%) 4e-43 sapiens, 128 aa. [US5871723-A, 16- 1..128 109/137 (79%) FEB-1999]

AAR13519 Leukocyte derived growth factor - 1..132 106/137 (77%) 4e-43 Homo sapiens, 128 aa. [WO911 515- 1..128 109/137 (79%) A, 08-AUG-1991]

AAR05767 Precursor of platelet basic protein 1..132 106/137 (77%) 4e-43 (PBP) - Synthetic, 128 aa. 1-128 109/137 (79%) [WO9006321-A, 14-JUN-1990]

AAR13520 Leukocyte derived growth factor 1..132 104/137 (75%) 2e-42 analogue - Homo sapiens, 128 aa. 1-128 109/137 (78%) [WO9111515-A, 08-AUG-1991]

In a BLAST search of public sequence databases, the NOV22a protein was found to have homology to the proteins shown in the BLASTP data in Table 22E.

PFam analysis predicts that the NOV22a protein contains the domains shown in the Table 22F.

EXAMPLE 23.

The NOV23 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 23A.

Further anayss o t e 23a proten yede t e o owng propertes s own Table 23B.

Table 23B. Protein Sequence Properties NOV23a

PSort analysis: 0.5804 probability located in outside; 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen); 0.1000 probability located in lysosome (lumen)

SignalP analysis: j Cleavage site between residues 27 and 28

A search of the NOV23a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 23 C.

In a BLAST search of public sequence databases, the NOV23a protein was found to have homology to the proteins shown in the BLASTP data in Table 23D.

PFam analysis predicts that the NOV23a protein contains the domains shown in the Table 23E.

EXAMPLE 24.

The NOV24 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 24A.

Further analysis of the NOV24a protein yielded the following properties shown in Table 24B.

Table 24B. Protein Sequence Properties NOV24a

PSort analysis: 0.3700 probability located in outside; 0.1080 probability located in nucleus; 0.1000 probability located in endoplasmic reticulum (membrane); 0.1000 probability located in endoplasmic reticulum (lumen)

SignalP analysis: J Cleavage site between residues 18 and 19

A search of the NOV24a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 24C.

In a BLAST search of public sequence databases, the NOV24a protein was found to have homology to the proteins shown in the BLASTP data in Table 24D.

PFam analysis predicts that the NOV24a protein contains the domains shown in the Table 24E.

EXAMPLE 25.

The NOV25 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 25A.

Furt er anayss o t e NOV25a protein yelde t e olowng propertes s own Table 25B.

Table 25B. Protein Sequence Properties NOV25a

PSort analysis: 0.3600 probability located in microbody (peroxisome); 0.3000 probability located in nucleus; 0.1000 probability located in mitochondrial matrix space; 0.1000 probability located in lysosome (lumen)

SignalP analysis: No Known Signal Sequence Predicted

A search of the NOV25a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 25 C.

In a BLAST search of public sequence databases, the NOV25a protein was found to have homology to the proteins shown in the BLASTP data in Table 25D.

PFam analysis predicts that the NOV25a protein contains the domains shown in the Table 25E.

EXAMPLE 26.

The NOV26 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 26A.

Sequence comparison of the above protein sequences yields the following sequence relationships shown in Table 26B.

Table 26B. Comparison of NOV26a against NOV26b and NOV26c.

NOV26a Residues/ Identities/

Protein Sequence Match Residues Similarities for the Matched Region

NOV26b 77..280 191/204 (93%) 10..213 191/204 (93%)

NOV26c 1..280 268/280 (95%) 1..280 268/280 (95%)

Further analysis of the NOV26a protein yielded the following properties shown in Table 26C.

Table 26C. Protein Sequence Properties NOV26a

PSort analysis: 0.8650 probability located in lysosome (lumen); 0.6854 probability located in outside; 0.1092 probability located in microbody (peroxisome); 0.1000 probability located in endoplasmic reticulum (membrane)

SignalP analysis: j Cleavage site between residues 19 and 20

A search of the NOV26a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 26D.

In a BLAST search of public sequence databases, the NOV26a protein was found to have homology to the proteins shown in the BLASTP data in Table 26E.

PFam analysis predicts that the NOV26a protein contains the domains shown in the Table 26F.

EXAMPLE 27.

The NOV27 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 27A.

Table 27 A. NOV27 Sequence Analysis

SEQ ID NO: 119 3290 bp

NOV27a, GCCGGGAGGGCCGCGTGAGGAGAGCGAAGAGGGAGCCCGAGCTCTGCGGCCCCGGGTG CG98092-01 GCGGGCCGGGGGCGCCCGTGAGCAGAGACCTCCCCGTCGACGGGGGGCGATGTTCCCC DNA Sequence TCACCCGTGGGCTCTTCGGGCAGCCAGGGCATCCCGCCGCTGGGACCTGGACCCGCTC

AGTGGGCACCCCTCAGGCTGCTCCATGGAGACCCTGTGCCCTGCGCCCCGCCTGGCAG

TGCCGGCGTCCCCGCGAGGGTCGCCCTGCTCCCCCACGCCCCGGAAGCCGTGTCGGGG

GACCCAGGAATTCTCTCCGCTGTGCCTGCGTGCCCTCGCCTTCTGCGCCCTTGCCAAG

CCCCGGGCGTCCTCTCTGGGCCCGGGGCCTGGGGAGCTGGCGGCGCGGTCCCCAGTGC

TGCGGGGCCCTCAGGCCCCCCTGCGCCCTGGCGGCTGGGCCCCGGATGGCCTGAAGCA

CCTCTGGGCACCGACCGGGCGGCCCGGCGTTCCTAACACCGCCGCCGGCGAGGATGCG

GACGTCGCAGCGTGCCCCCGCCGCGGAGAGGAGGAAGAGGGCGGAGGCGGTTTCCCGC

ACTTCGGCGTTCGCTCCTGTGCACCTCCGGGCCGCTGCCCTGCGCCCCCGCACCCTCG

GGAATCTACGACCAGCTTCGCCTCGGCCCCGCCTCGCCCGGCCCCGGGTCTCGAGCCT

CAGCGTGGCCCAGCCGCCAGCCCGCCTCAGGAACCCAGTTCCCGGCCTCCGTCGCCAC

CTGCGGGCCTCTCCACCGAGCCCGCGGGTCCCGGGACGGCGCCGCGGCCGTTCCTGCC

CGGCCAGCCTGCCGAAGTCGATGGAAACCCCCCGCCGGCCGCCCCCGAGGCTCCAGCG

GCCAGCCCCTCGACGGCCAGCCCGGCTCCGGCCGCACCCGGAGATCTCCGCCAGGAAC

ATTTCGATCGTCTGATCCGCCGGTCGAAACTTTGGTGTTACGCGAAGGGCTTCGCCTT

GGACACTCCGAGTTTGCGCCGGGGGCCAGAGCGGCCGCCTGCGAAAGGGCCGGCTCGG

GGAGCCGCCAAGAAACGCCGGCTGCCGGCGCCCCCTCCGCGCACCGCGCAGCCCCGCC

GCCCTGCACCGACGCTCCCCACCACGAGCACCTTCAGCCTCCTCAACTGCTTCCCCTG

CCCCCCGGCCCTGGTGGTGGGGGAAGACGGAGACCTAAAGCCGGCATCCTCGCTTCGC

CTCCAGGGAGACTCTAAGCCCCCGCCCGCCCACCCGCTGTGGAGGTGGCAGATGGGGG

GTCCCGCTGTCCCTGAGCCCCCTGGCCTCAAATTCTGGGGGATCAACATGGATGAAAG

CTGACCGTGGGACTTCTGCCAAAGGGGAAAAGTTGGGACCATGGCCAAACCGCGGGCT

TGAGGAGGGAGCCCCGTTTCTCACATTTGTCCCCTTCCTTTACATTTTAGGAGCTGTG

GGCAGAGGGACCTAAATAACAGTGATCTTCATTCAAGCACCTAAGTTTTCGGGGTGAC

AGTCCCTCCCCCTCATCCTTTGCAGAGGAACCCAGGGCTGGAGTCGGGAGAAGGCTGA

TGACATAGATTCCAATCCCTGCCTCCTTCCATCTCGGACCGTTGGAGGCAGGGCCTGC

ACCCCAGTGGGAGCAAAGGAGGCCACCGCTCAAAGACACCCCCCCACCCAAAAAAAAG

GGGAGAAGAGAGAGACCTCGGTGATGGACAAACCGGTTGTTACTGTGTCTGTGGGCGA

GCCTGGGGTGCGGGGCTGTGGTGGGGGTGGGGAGATGATTGGCAGCTCCCTGGGGGCA

TCCCCCACCCCCACTGTCCAGGCCTTTAACCCTTTGCTCCCCTCAGGCCTTCCCTAAC

GCTCCAAGCACCGCTGGAGCCTTTAATGGGTGAGGGAACTTGGGTAAGAGGAAGATCA

CCCCCTTCCTGTCCCCTTTCTAGGCCCCCTCAAGTGCAGGTGACCCTTAATTGGTGAG

ATCTTCAGCCTCAGCCGCCGACCTTTCCCTTTTGTCCAGTTTTGGAGTTCCCGTTTTT

TCCTTGTTTGCTTTCCGAGTGTAAGGTCTGGCCGGTGAGAAAGATTTCCCCCAACCTT

GATTAATCAGCCCCCTCCCCCAACTTACTTCCCTTAGGACGGGTAGGGCTGAGGGACC

TCCTCTCCTGGAAAGTGCTTACTTTGCCTGGGGAAGGGGCTAGACACTGTCCCAGGGA

AAGTAATAGAAGGTGGAAGAAATCAATAAAATCAGACCAAACAAGTCGCCTTTCGAGG

GCCTCCACCGATTTATGGATGAGAGGGGGTGGAGGTGGAAGGCAGGCCCAAGTCCATT

CTTTGGACACCCAAACTCAGCCCCCTTAAAGAGTGGAAACAAAACAAGCTGCACTTTG

CAGAGGTGGTAAATGAAAGGACTCTTGGCCTAACTTCAAGAGTCCCCTGGGGTTTGAA

GGGGCAAAGTTTGAGTCTGGATGGAACCTGGGCTGAGGTACCTTAAGCTTCCCCCCGC

AACACCCCAGCCTCAGGGATTGCGGGAGTTGTCAGAGATCTGATGGATCCGAAAGGGG

CAGGGCCAGGGGATTAGGTTTGGGGTCAGAGGTTCTGTTTTCCAGGGGAGGGGTGAGA

TAGGCCTGGATCATGCCCTCTGCCATGCCCTCCAGCTAGGAGGATCTTGAGTCAGAGA

GGATTGGAAGTGCTTTCTCCTCCACCCAGGTGAGGTCAGGGGAGCTTAGGTCTTAGGG

AGATGGCAAGTTGAGGTATGAAGGGAAGCTGGGGCTTTTGGAGCTGCCGAACAACTGA.

GGGACCCAGTGCGCCTTCCATCCCGCACTAGTGAATAGCGCCCCCTCTTCCCCCGAAA

ACGAGGTGCGAGAGGAACAATTCCCACGCTGGGGAAGGACTTGTCTCCTTTTCTGTGA AAATGCTTTGTAAAAAGTTGTTATTGTTTGCATAGAGCAGATTCTTGAGAAAAACTGT

TTTGGACCATAAAAGTTTTGTTTGTTTTAAAAACTGTCTCCTTTCATTTTTCTTTCCT TGGGGGTGAGGGTGGGGGTGGGGTGGGTGGGCACTTCTCTCTTTTTCCTTAACATCTG

GCCTCTGTGAACCCTGCTGACCTCCCCTCCCCCTCCAGCTGTGTTGTGGGAGGAGGAG

GAAGAAGGGGTGGGGGAGTGCCTTCCACCCTGTGCTTCGGGAGTCTCCATCTTATTTT

GCCCCCCAAGAATGGAGAACGGGAGGAAGAAGACACAAGGGGTGGGGAGAGAATCGCT

GTGAAGAGGGGGGCTGTCAGAAGTCTGGAAAGGCAGACTCCC

ORF Start: ATG at 199 ORF Stop: TGA at 1336

SEQ ID NO: 120 379 aa MW at 39321.2kD

NOV27a, METLCPAPRLAVPASPRGSPCSPTPRKPCRGTQEFSPLCLRALAFCALAKPRASSLGP CG98092-01 GPGELAARSPVLRGPQAPLRPGGAPDGLKHLWAPTGRPGVPNTAAGEDADVAACPRR Protein GEEEEGGGGFPHFGVRSCAPPGRCPAPPHPRESTTSFASAPPRPAPGLEPQRGPAASP Sequence PQEPSSRPPSPPAGLSTEPAGPGTAPRPFLPGQPAEVDGNPPPAAPEAPAASPSTASP APAAPGDLRQEHFDRLIRRSKLWCYAKGFALDTPSLRRGPERPPAKGPARGAAKKRRL PAPPPRTAQPRRPAPTLPTTSTFSLLNCFPCPPALWGEDGDLKPASSLRLQGDSKPP PAHPL RWQMGGPAVPEPPGLKF GINMDES

Further analysis of the NOV27a protein yielded the following properties shown in Table 27B.

Table 27B. Protein Sequence Properties NOV27a

PSort analysis: 0.3000 probability located in microbody (peroxisome); 0.3000 probability located in nucleus; 0.2584 probability located in lysosome (lumen); 0.1000 probability located in mitochondrial matrix space

SignalP analysis: Cleavage site between residues 50 and 51

A search of the NOV27a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 27C.

In a BLAST search of public sequence databases, the NOV27a protein was found to have homology to the proteins shown in the BLASTP data in Table 27D.

PFam analysis predicts that the NOV27a protein contains the domains shown in the Table 27E.

Table 27E. Domain Analysis of NOV27a

Identities/

Pfam Domain NOV27a Match Region Similarities Expect Value for the Matched Region

EXAMPLE 28.

The NOV28 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 28 A.

Table 28A. NOV28 Sequence Analysis

SEQ ID NO: 121 1520 bp

NOV28a, GGCAGCGGGGCAGGCGCGTGGCCGGGCCGCGGCGCGATGAGTGGGGCCCGGGCGGCGC CG98121-01 CCGGGGCCGCGGGCAACGGCGCGGTCCGGGGGCTGCGGGTGGACGGGCTGCCCCCGCT DNA Sequence GCCAAAGAGCTTGAGCGGGCTGCTGCACTCGGCGTCGGGCGGCGGCGCGTCTGGGGGC TGGCGGCACCTGGAGCGGCTGTACGCGCAGAAGTCGCGCATCCAGGACGAGCTGAGCC GCGGGGGCCCGGGCGGCGGCGGGGCCCGGGCGGGCAGCGCTGCCCGCCAAGCCTCCCA ACCTGGACGCCGCTCTGGCGCTGCTCCGCAAAGAGATGGTTGGTCTCCGCCAGCTGGA CATGTCCTTGCTCTGCCAACTGTACAGCCTCTACGAGTCGATTCAGGAGTACAAGGGG GCATGCCAGGCAGCCTCCAGCCCAGACTGCACTTACGCTCTGGAGAACGGCTTCTTCG ATGAAGAGGAGGAATATTTCCAGGAGCAGAACTCCCTGCACGACAGGAGGGACCGAGG CCCTCCTCGGGACTTGTCACTGCCTGTCTCCTCCCTCTCCAGCAGCGACTGGATTCTG

GAGTCCATCTAGAGGGTCTTGGGAGGGATGTGACTGTTGGGAAGCCCTTCCTACTGGA

CACGCTGTCATCATTTGCTGCTTCTCTTGCAAGAAAGCACCTCCGTTGTGGACGGTCC

TCGGGCACAGGGGATGAGCGCTACCAGTTTCATTTGTAGGCAGGGAGTTCTCCGCGGA

TGCATGGTGGCAGTCTGCTTTGATGGCAGCAGTTTCTGCTTAGGTGACCTAGAGGTCC

TCAGCAGTATCCTCCACACCTATTTATTGAGGTGCACCTGCTGGGGATTCATAATGAG

AATATAACAAGAGGATCTCGGTGAAAGGCCTTAGTGGGTGTTTTGTGTGAGGTGGCTT

GTAGCTAGCTACTTCCTTACAGATGGTAGAGTATTCCAATCCTCTTTGTGTTAGGGTT

CTTGCTTCCAGTTTGGGATGTATTAAAACCACCATTTCACTGCTTCCCTTCCTCAATA

TGCTCTGCAGCTTTTCTTGCTGTTTAAACCTCTCGCCTCAGCTTTATTTATTTGTAAG

CTGCATTACTAACTGCCCAGTGATTCGGTGAAAGCTTTTTACTGAAAAAGTTAACATT

TCTAGTCATCCCAATCAACTGGCTTTTTTCAACCAAAATTTTATATCATTCTTTGTCT

ATCAGATACGAGAGGAAGGAAGATAATACGAAGACATGTTGAATAGTGAAAAAAAAAA

AAAAGAACCACAAAAACTGGGGCAAGCCAATGTGATGTATCACTCACTGTAAGATGGC

AAATGTTTTCATTTTTAAGATTCCGAATGTAAACTAGTGTGCTAGAAAGCAAACCACC

CGCCACTCAAACCAGTAATTACCTTAAGCCTTAATATATTTATTAAAATACTTTATGA

GAACATTACACTTTGTAGGTTAAAAATGAGGATAAAATGCTAAACTATCAAAAAAAAA

AAAAAAGAAAAA

ORF Start: ATG at 37 ORF Stop: TGA at 466

SEQ ID NO: 122 143 aa MW at 14441. lkD

NOV28a, CG98121-01 RIQDELSRGGPGGGGARAGSAARQASQPGRRSGAAPQRDGWSPPAGHVLALPTVQPLR Protein VDSGVQGGMPGSLQPRLHLRSGERLLR Sequence

Further analysis of the NOV28a protein yielded the following properties shown in Table 28B.

Table 28B. Protein Sequence Properties NOV28a

PSort analysis: 0.8231 probability located in lysosome (lumen); 0.6500 probability located in cytoplasm; 0.1000 probability located in mitochondrial matrix space; 0.0580 probability located in microbody (peroxisome)

SignalP analysis: No Known Signal Sequence Predicted A search of the NOV28a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 28C.

In a BLAST search of public sequence databases, the NOV28a protein was found to have homology to the proteins shown in the BLASTP data in Table 28D.

PFam analysis predicts that the NOV28a protein contains the domains shown in the Table 28E.

Table 28E. Domain Analysis of NOV28a

Identities/

Pfam Domain NOV28a Match Region Similarities ExDect Value for the Matched Region

EXAMPLE 29.

The NOV29 clone was analyzed, and the nucleotide and encoded polypeptide sequences are shown in Table 29A.

Table 29A. NOV29 Sequence Analysis

SEQ ID NO: 123 970 bp

NOV29a, GCTGCTGGTTTTGAAACATGAATCTTTCGCTCGTCCTGGCTGCCTTTTGCTTGGGAAT CG99662-01 AGCCTCCGCTGTTCCAAAATTTGACCAAAATTTGGATACAAAGTGGTACCAGTGGAAG DNA Sequence GCAACACACAGAAGATTATATGGCGCGAATGAAGAAGGATGGAGGAGAGCAGTGTGGG AAAAGAATATGAAAATGATTGAACTGCACAATGGGGAATACAGCCAAGGGAAACATGG CTTCACAATGGCCATGAATGCTTTTGGTGACATGACCAATGAAGAATTCAGGCAGATG ATGGGTTGCTTTCGAAACCAGAAATTCAGGAAGGGGAAAGTGTTCCGTGAGCCTCTGT TTCTTGATCTTCCCAAATCTGTGGATTGGAGAAAGAAAGGCTACGTGACGCCAGTGAA GAATCAGAATCTGGTGGACTGTTCGCGTCCTCAAGGCAATCAGGGCTGCAATGGTGGC TTCATGGCTAGGGCCTTCCAGTATGTCAAGGAGAACGGAGGCCTGGACTCTGAGGAAT CCTATCCATATGTAGCAGTGGATGAAATCTGTAAGTACAGACCTGAGAATTCTGTTGC TAATGACACTGGCTTCACAGTGGTCGCACCTGGAAAGGAGAAGGCCCTGATGAAAGCA GTCGCAACTGTGGGGCCCATCTCCGTTGCTATGGATGCAGGCCATTCGTCCTTCCAGT TCTACAAATCAGGCATTTATTTTGAACCAGACTGCAGCAGCAAAAACCTGGATCATGG TGTTCTGGTGGTTGGCTACGGCTTTGAAGGAGCAAATTCGAATAACAGCAAGTATTGG CTCGTCAAAAACAGCTGGGGTCCAGAATGGGGCTCGAATGGCTATGTAAAAATAGCCA AAGACAAGAACAACCACTGTGGAATCGCCACAGCAGCCAGCTACCCCAATGTGTGAGC TGATGGATGGTGAGGAGGAAGGACTTAAGGACAGCATGTCTA

ORF Start: ATG at 18 ORF Stop: TGA at 924

SEQ ID NO: 124 302 aa MW at 33897.2kD

NOV29a, NLSLVIAAFCLGIASAVPKFDQNLDTKWYQWKATHRRLYGANEEGWRRAV EKN KM CG99662-01 IELMIGE SQGKHGF )YI AFGDMTNEEFRQMMGCFRNQKFRKGK FREPLFLD PK Protein SVD RKKGYVTPVKNQNLVDCSRPQGNQGCNGGFMARAFQYVKENGGLDSEESYPYVA Sequence VDEICKYRPENSVALVTDTGFTVVAPGLNΕKAIJVIKAVATVGPISVAMDAGHSSFQFYKSGI YFEPDCSSKNLDHGVLVVGYGFEGANSNNSKYWLVKNSWGPEWGSNGYVKIAKDIOSΓNH CGIATAASYPNV

Further analysis of the NOV29a protein yielded the following properties shown in

Table 29B.

Table 29B. Protein Sequence Properties NOV29a

PSort analysis: 0.8200 probability located in outside; 0.1900 probability located in lysosome (lumen); 0.1598 probability located in microbody (peroxisome); 0.1000 probability located in endoplasmic reticulum (membrane)

SignalP analysis: j Cleavage site between residues 18 and 19

A search of the NOV29a protein against the Geneseq database, a proprietary database that contains sequences published in patents and patent publication, yielded several homologous proteins shown in Table 29C.

In a BLAST search of public sequence databases, the NOV29a protein was found to have homology to the proteins shown in the BLASTP data in Table 29D.

PFam analysis predicts that the NOV29a protein contains the domains shown in the Table 29E.

Example B: Sequencing Methodology and Identification of NOVX Clones 1. GeneCalling™ Technology: This is a proprietary method of performing differential gene expression profiling between two or more samples developed at CuraGen and described by Shimkets, et al., "Gene expression analysis by transcript profiling coupled to a gene database query" Nature Biotechnology 17:198-803 (1999). cDNA was derived from various human samples representing multiple tissue types, normal and diseased states, physiological states, and developmental states from different donors. Samples were obtained as whole tissue, primary cells or tissue cultured primary cells or cell lines. Cells and cell lines may have been treated with biological or chemical agents that regulate gene expression, for example, growth factors, chemokines or steroids. The cDNA thus derived was then digested with up to as many as 120 pairs of restriction enzymes and pairs of linker-adaptors specific for each pair of restriction enzymes were ligated to the appropriate end. The restriction digestion generates a mixture of unique cDNA gene fragments. Limited PCR amplification is performed with primers homologous to the linker adapter sequence where one primer is biotinylated and the other is fluorescently labeled. The doubly labeled material is isolated and the fluorescently labeled single strand is resolved by capillary gel electrophoresis. A computer algorithm compares the electropherograms from an experimental and control group for each of the restriction digestions. This and additional sequence-derived information is used to predict the identity of each differentially expressed gene fragment using a variety of genetic databases. The identity of the gene fragment is confirmed by additional, gene-specific competitive PCR or by isolation and sequencing of the gene fragment.

2. SeqCalling™ Technology: cDNA was derived from various human samples representing multiple tissue types, normal and diseased states, physiological states, and developmental states from different donors. Samples were obtained as whole tissue, primary cells or tissue cultured primary cells or cell lines. Cells and cell lines may have been treated with biological or chemical agents that regulate gene expression, for example, growth factors, chemokines or steroids. The cDNA thus derived was then sequenced using CuraGen's proprietary SeqCalling technology. Sequence traces were evaluated manually and edited for corrections if appropriate. cDNA sequences from all samples were assembled together, sometimes including public human sequences, using bioinformatic programs to produce a consensus sequence for each assembly. Each assembly is included in CuraGen Corporation's database. Sequences were included as components for assembly when the extent of identity with another component was at least 95% over 50 bp. Each assembly represents a gene or portion thereof and includes information on variants, such as splice forms single nucleotide polymorphisms (SNPs), insertions, deletions and other sequence variations. 3. PathCalling™ Technology:

The NOVX nucleic acid sequences are derived by laboratory screening of cDNA library by the two-hybrid approach. cDNA fragments covering either the full length of the DNA sequence, or part of the sequence, or both, are sequenced. In silico prediction was based on sequences available in CuraGen Corporation's proprietary sequence databases or in the public human sequence databases, and provided either the full length DNA sequence, or some portion thereof.

The laboratory screening was performed using the methods summarized below: cDNA libraries were derived from various human samples representing multiple tissue types, normal and diseased states, physiological states, and developmental states from different donors. Samples were obtained as whole tissue, primary cells or tissue cultured primary cells or cell lines. Cells and cell lines may have been treated with biological or chemical agents that regulate gene expression, for example, growth factors, chemokines or steroids. The cDNA thus derived was then directionally cloned into the appropriate two-hybrid vector (Gal4-activation domain (Gal4-AD) fusion). Such cDNA libraries as well as commercially available cDNA libraries from Clontech (Palo Alto, CA) were then transferred from E.coli into a CuraGen Corporation proprietary yeast strain (disclosed in U. S. Patents 6,057,101 and 6,083,693, incorporated herein by reference in their entireties).

Gal4-binding domain (Gal4-BD) fusions of a CuraGen Corportion proprietary library of human sequences was used to screen multiple Gal4-AD fusion cDNA libraries resulting in the selection of yeast hybrid diploids in each of which the Gal4-A.D fusion contains an individual cDNA. Each sample was amplified using the polymerase chain reaction (PCR) using non-specific primers at the cDNA insert boundaries. Such PCR product was sequenced; sequence traces were evaluated manually and edited for corrections if appropriate. cDNA sequences from all samples were assembled together, sometimes including public human sequences, using bioinf rmatic programs to produce a consensus sequence for each assembly. Each assembly is included in CuraGen Corporation's database. Sequences were included as components for assembly when the extent of identity with another component was at least 95% over 50 bp. Each assembly represents a gene or portion thereof and includes information on variants, such as splice forms single nucleotide polymorphisms (SNPs), insertions, deletions and other sequence variations.

Physical clone: the cDNA fragment derived by the screening procedure, covering the entire open reading frame is, as a recombinant DNA, cloned into pACT2 plasmid (Clontech) used to make the cDNA library. The recombinant plasmid is inserted into the host and selected by the yeast hybrid diploid generated during the screening procedure by the mating of both CuraGen Corporation proprietary yeast strains N106' and YULH (U. S. Patents 6,057,101 and 6,083,693).

4. RACE: Techniques based on the polymerase chain reaction such as rapid amplification of cDNA ends (RACE), were used to isolate or complete the predicted sequence of the cDNA of the invention. Usually multiple clones were sequenced from one or more human samples to derive the sequences for fragments. Various human tissue samples from different donors were used for the RACE reaction. The sequences derived from these procedures were included in the SeqCalling Assembly process described in preceding paragraphs.

5. Exon Linking: The NOVX target sequences identified in the present invention were subjected to the exon linking process to confirm the sequence. PCR primers were designed by starting at the most upstream sequence available, for the forward primer, and at the most downstream sequence available for the reverse primer. In each case, the sequence was examined, walking inward from the respective termini toward the coding sequence, until a suitable sequence that is either unique or highly selective was encountered, or, in the case of the reverse primer, until the stop codon was reached. Such primers were designed based on in silico predictions for the full length cDNA, part (one or more exons) of the DNA or protein sequence of the target sequence, or by translated homology of the predicted exons to closely related human sequences from other species. These primers were then employed in PCR amplification based on the following pool of human cDNAs: adrenal gland, bone marrow, brain - amygdala, brain - cerebellum, brain - hippocampus, brain - substantia nigra, brain - thalamus, brain -whole, fetal brain, fetal kidney, fetal liver, fetal lung, heart, kidney, lymphoma - Raji, mammary gland, pancreas, pituitary gland, placenta, prostate, salivary gland, skeletal muscle, small intestine, spinal cord, spleen, stomach, testis, thyroid, trachea, uterus. Usually the resulting amplicons were gel purified, cloned and sequenced to high redundancy. The PCR product derived from exon linking was cloned into the pCR2.1 vector from Invitrogen. The resulting bacterial clone has an insert covering the entire open reading frame cloned into the pCR2.1 vector. The resulting sequences from all clones were assembled with themselves, with other fragments in CuraGen Corporation's database and with public ESTs. Fragments and ESTs were included as components for an assembly when the extent of their identity with another component of the assembly was at least 95% over 50 bp. In addition, sequence traces were evaluated manually and edited for corrections if appropriate. These procedures provide the sequence reported herein.

6. Physical Clone: Exons were predicted by homology and the intron/exon boundaries were determined using standard genetic rules. Exons were further selected and refined by means of similarity determination using multiple BLAST (for example, tBlastN, BlastX, and BlastN) searches, and, in some instances, GeneScan and Grail. Expressed sequences from both public and proprietary databases were also added when available to further define and complete the gene sequence. The DNA sequence was then manually corrected for apparent inconsistencies thereby obtaining the sequences encoding the full-length protein.

The PCR product derived by exon linking, covering the entire open reading frame, was cloned into the pCR2.1 vector from Invitrogen to provide clones used for expression and screening purposes.

Example C: Quantitative expression analysis of clones in various cells and tissues

The quantitative expression of various clones was assessed using microtiter plates containing RNA samples from a variety of normal and pathology-derived cells, cell lines and tissues using real time quantitative PCR (RTQ PCR). RTQ PCR was performed on an Applied Biosystems ABI PRISM® 7700 or an ABI PRISM® 7900 HT Sequence Detection System. Various collections of samples are assembled on the plates, and referred to as Panel 1 (containing normal tissues and cancer cell lines), Panel 2 (containing samples derived from tissues from normal and cancer sources), Panel 3 (containing cancer cell lines), Panel 4 (containing cells and cell lines from normal tissues and cells related to inflammatory conditions), Panel 5D/5I (containing human tissues and cell lines with an emphasis on metabolic diseases), AI_comprehensive_panel (containing normal tissue and samples from autoimmune diseases), Panel CNSD.01 (containing central nervous system samples from normal and diseased brains) and CNS_neurodegeneration_panel (containing samples from normal and Alzheimer's diseased brains).

RNA integrity from all samples is controlled for quality by visual assessment of agarose gel electropherograms using 28S and 18S ribosomal RNA staining intensity ratio as a guide (2:1 to 2.5:1 28s:18s) and the absence of low molecular weight RNAs that would be indicative of degradation products. Samples are controlled against genomic DNA contamination by RTQ PCR reactions run in the absence of reverse transcriptase using probe and primer sets designed to amplify across the span of a single exon. First, the RNA samples were normalized to reference nucleic acids such as constitutively expressed genes (for example, β-actin and GAPDH). Normalized RNA (5 ul) was converted to cDNA and analyzed by RTQ-PCR using One Step RT-PCR Master Mix Reagents (Applied Biosystems; Catalog No. 4309169) and gene-specific primers according to the manufacturer's instructions. In other cases, non-normalized RNA samples were converted to single strand cDNA (sscDNA) using Superscript II (Invitrogen Corporation; Catalog No. 18064-147) and random hexamers according to the manufacturer's instructions. Reactions containing up to 10 μg of total RNA were performed in a volume of 20 μl and incubated for 60 minutes at 42°C. This reaction can be scaled up to 50 μg of total RNA in a final volume of 100 μl. sscDNA samples are then normalized to reference nucleic acids as described previously, using IX TaqMan® Universal Master mix (Applied Biosystems; catalog No. 4324020), following the manufacturer's instructions.

Probes and primers were designed for each assay according to Applied Biosystems Primer Express Software package (version I for Apple Computer's Macintosh Power PC) or a similar algorithm using the target sequence as input. Default settings were used for reaction conditions and the following parameters were set before selecting primers: primer concentration = 250 nM, primer melting temperature (Tm) range = 58°-60°C, primer optimal Tm = 59°C, maximum primer difference = 2°C, probe does not have 5'G, probe Tm must be 10°C greater than primer Tm, amplicon size 75bp to lOObp. The probes and primers selected (see below) were synthesized by Synthegen (Houston, TX, USA). Probes were double purified by HPLC to remove uncoupled dye and evaluated by mass spectroscopy to verify coupling of reporter and quencher dyes to the 5' and 3' ends of the probe, respectively. Their final concentrations were: forward and reverse primers, 900nM each, and probe, 200nM.

PCR conditions: When working with RNA samples, normalized RNA from each tissue and each cell line was spotted in each well of either a 96 well or a 384-well PCR plate (Applied Biosystems). PCR cocktails included either a single gene specific probe and primers set, or two multiplexed probe and primers sets (a set specific for the target clone and another gene-specific set multiplexed with the target probe). PCR reactions were set up using TaqMan® One-Step RT-PCR Master Mix (Applied Biosystems, Catalog No. 4313803) following manufacturer's instructions. Reverse transcription was performed at 48°C for 30 minutes followed by amplification/PCR cycles as follows: 95°C 10 min, then 40 cycles of 95°C for 15 seconds, 60°C for 1 minute. Results were recorded as CT values (cycle at which a given sample crosses a threshold level of fluorescence) using a log scale, with the difference in RNA concentration between a given sample and the sample with the lowest CT value being represented as 2 to the power of delta CT. The percent relative expression is then obtained by taking the reciprocal of this RNA difference and multiplying by 100.

When working with sscDNA samples, normalized sscDNA was used as described previously for RNA samples. PCR reactions containing one or two sets of probe and primers were set up as described previously, using IX TaqMan® Universal Master mix (Applied Biosystems; catalog No. 4324020), following the manufacturer's instructions. PCR amplification was performed as follows: 95°C 10 min, then 40 cycles of 95°C for 15 seconds, 60°C for 1 minute. Results were analyzed and processed as described previously. Panels 1, 1.1, 1.2, and 1.3D The plates for Panels 1, 1.1, 1.2 and 1.3D include 2 control wells (genomic DNA control and chemistry control) and 94 wells containing cDNA from various samples. The samples in these panels are broken into 2 classes: samples derived from cultured cell lines and samples derived from primary normal tissues. The cell lines are derived from cancers of the following types: lung cancer, breast cancer, melanoma, colon cancer, prostate cancer, CNS cancer, squamous cell carcinoma, ovarian cancer, liver cancer, renal cancer, gastric cancer and pancreatic cancer. Cell lines used in these panels are widely available through the American Type Culture Collection (ATCC), a repository for cultured cell lines, and were cultured using the conditions recommended by the ATCC. The normal tissues found on these panels are comprised of samples derived from all major organ systems from single adult individuals or fetuses. These samples are derived from the following organs: adult skeletal muscle, fetal skeletal muscle, adult heart, fetal heart, adult kidney, fetal kidney, adult liver, fetal liver, adult lung, fetal lung, various regions of the brain, the spleen, bone marrow, lymph node, pancreas, salivary gland, pituitary gland, adrenal gland, spinal cord, thymus, stomach, small intestine, colon, bladder, trachea, breast, ovary, uterus, placenta, prostate, testis and adipose.

In the results for Panels 1 , 1.1 , 1.2 and 1 D, the following abbreviations are used: ca. = carcinoma,

* = established from metastasis, met = metastasis, s cell var = small cell variant, non-s = non-sm = non-small, squam = squamous, pi. eff = pi effusion = pleural effusion, glio = glioma, astro = astrocytoma, and neuro = neuroblastoma.

General_screeningjpanel_vl.4 and General_screening panel_vl.5 The plates for Panels 1.4 and 1.5 include 2 control wells (genomic DNA control and chemistry control) and 94 wells containing cDNA from various samples. The samples in Panels 1.4 and 1.5 are broken into 2 classes: samples derived from cultured cell lines and samples derived from primary normal tissues. The cell lines are derived from cancers of the following types: lung cancer, breast cancer, melanoma, colon cancer, prostate cancer, CNS cancer, squamous cell carcinoma, ovarian cancer, liver cancer, renal cancer, gastric cancer and pancreatic cancer. Cell lines used in Panel 1.4 are widely available through the American Type Culture Collection (ATCC), a repository for cultured cell lines, and were cultured using the conditions recommended by the ATCC. The normal tissues found on Panels 1.4 and 1.5 are comprised of pools of samples derived from all major organ systems from 2 to 5 different adult individuals or fetuses. These samples are derived from the following organs: adult skeletal muscle, fetal skeletal muscle, adult heart, fetal heart, adult kidney, fetal kidney, adult liver, fetal liver, adult lung, fetal lung, various regions of the brain, the spleen, bone marrow, lymph node, pancreas, salivary gland, pituitary gland, adrenal gland, spinal cord, thymus, stomach, small intestine, colon, bladder, trachea, breast, ovary, uterus, placenta, prostate, testis and adipose. Abbreviations are as described for Panels 1, 1.1, 1.2, and 1.3D.

Panels 2D and 2.2

MISSING AT THE TIME OF PUBLICATION

cell lines or tissues related to inflammatory conditions. Total RNA from control normal tissues such as colon and lung (Stratagene, La Jolla, CA) and thymus and kidney (Clontech) was employed. Total RNA from liver tissue from cirrhosis patients and kidney from lupus patients was obtained from BioChain (Biochain Institute, Inc., Hayward, CA). Intestinal tissue for RNA preparation from patients diagnosed as having Crohn's disease and ulcerative colitis was obtained from the National Disease Research Interchange (NDRI) (Philadelphia, PA).

Astrocytes, lung fibroblasts, dermal fibroblasts, coronary artery smooth muscle cells, small airway epithelium, bronchial epithelium, microvascular dermal endothelial cells, microvascular lung endothelial cells, human pulmonary aortic endothelial cells, human umbilical vein endothelial cells were all purchased from Clonetics (Walkersville, MD) and grown in the media supplied for these cell types by Clonetics. These primary cell types were activated with various cytokines or combinations of cytokines for 6 and/or 12-14 hours, as indicated. The following cytokines were used; IL-1 beta at approximately l-5ng/ml, TNF alpha at approximately 5-1 Ong/ml, IFN gamma at approximately

20-50ng/ml, IL-4 at approximately 5-1 Ong/ml, IL-9 at approximately 5-1 Ong/ml, IL-13 at approximately 5-1 Ong/ml. Endothelial cells were sometimes starved for various times by culture in the basal media from Clonetics with 0.1 % serum.

Mononuclear cells were prepared from blood of employees at CuraGen Corporation, using Ficoll. LAK cells were prepared from these cells by culture in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco/Life Technologies, Rockville, MD), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5x10^'5M (Gibco), and lOmM Hepes (Gibco) and Interleukin 2 for 4-6 days. Cells were then either activated with 10-20ng/ml PMA and l-2μg/ml ionomycin, IL-12 at 5-1 Ong/ml, IFN gamma at 20-50ng/ml and IL-18 at 5-1 Ong/ml for 6 hours. In some cases, mononuclear cells were cultured for 4-5 days in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5x10^"5M (Gibco), and lOmM Hepes (Gibco) with PHA (phytohemagglutinin) or PWM (pokeweed mitogen) at approximately 5μg/ml. Samples were taken at 24, 48 and 72 hours for RNA preparation. MLR (mixed lymphocyte reaction) samples were obtained by taking blood from two donors, isolating the mononuclear cells using Ficoll and mixing the isolated mononuclear cells 1:1 at a final concentration of approximately 2xl0⁶cells/ml in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol (5.5x10^"5M) (Gibco), and lOmM Hepes (Gibco). The MLR was cultured and samples taken at various time points ranging from 1- 7 days for RNA preparation.

Monocytes were isolated from mononuclear cells using CD 14 Miltenyi Beads, +ve VS selection columns and a Vario Magnet according to the manufacturer's instructions. Monocytes were differentiated into dendritic cells by culture in DMEM 5% fetal calf serum (FCS) (Hyclone, Logan, UT), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5xlO^"5M (Gibco), and lOmM Hepes (Gibco), 50ng/ml GMCSF and 5ng/ml IL-4 for 5-7 days. Macrophages were prepared by culture of monocytes for 5-7 days in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5x10^"5M (Gibco), lOmM Hepes (Gibco) and 10% AB Human Serum or MCSF at approximately 50ng/ml. Monocytes, macrophages and dendritic cells were stimulated for 6 and 12-14 hours with lipopolysaccharide (LPS) at lOOng/ml. Dendritic cells were also stimulated with anti-CD40 monoclonal antibody (Pharmingen) at lOμg/ml for 6 and 12-14 hours. CD4 lymphocytes, CD8 lymphocytes and NK cells were also isolated from mononuclear cells using CD4, CD8 and CD56 Miltenyi beads, positive VS selection columns and a Vario Magnet according to the manufacturer's instructions. CD45RA and CD45RO CD4 lymphocytes were isolated by depleting mononuclear cells of CD8, CD56, CD14 and CD19 cells using CD8, CD56, CD14 and CD19 Miltenyi beads and positive selection. CD45RO beads were then used to isolate the CD45RO CD4 lymphocytes with the remaining cells being CD45RA CD4 lymphocytes. CD45RA CD4, CD45RO CD4 and CD8 lymphocytes were placed in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco)j ImM sodium pyruvate (Gibco), mercaptoethanol 5.5x10^"5M (Gibco), and lOmM Hepes (Gibco) and plated at 10⁶cells/ml onto Falcon 6 well tissue culture plates that had been coated overnight with 0.5μg/ml anti-CD28 (Pharmingen) and 3ug/ml anti-CD3 (OKT3, ATCC) in PBS. After 6 and 24 hours, the cells were harvested for RNA preparation. To prepare chronically activated CD8 lymphocytes, we activated the isolated CD8 lymphocytes for 4 days on anti-CD28 and anti-CD3 coated plates and then harvested the cells and expanded them in DMEM 5% FCS (Hyclone), 1 OOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5x10^"5M (Gibco), and lOmM Hepes (Gibco) and IL-2. The expanded CD8 cells were then activated again with plate bound anti-CD3 and anti-CD28 for 4 days and expanded as before. RNA was isolated 6 and 24 hours after the second activation and after 4 days of the second expansion culture. The isolated NK cells were cultured in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5x10^"5M (Gibco), and lOmM Hepes (Gibco) and IL-2 for 4-6 days before RNA was prepared. To obtain B cells, tonsils were procured from NDRI. The tonsil was cut up with sterile dissecting scissors and then passed through a sieve. Tonsil cells were then spun down and resupended at 10⁶cells/ml in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5x10^"5M (Gibco), and lOmM Hepes (Gibco). To activate the cells, we used PWM at 5 μg/ml or anti-CD40 (Pharmingen) at approximately lOμg/ml and IL-4 at 5-1 Ong/ml. Cells were harvested for RNA preparation at 24,48 and 72 hours.

To prepare the primary and secondary Thl/Th2 and Tri cells, six- well Falcon plates were coated overnight with lOμg/ml anti-CD28 (Pharmingen) and 2μg/ml OKT3 (ATCC), and then washed twice with PBS. Umbilical cord blood CD4 lymphocytes (Poietic Systems, German Town, MD) were cultured at 10⁵-10⁶cells/ml in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5x10^"5M (Gibco), lOmM Hepes (Gibco) and IL-2 (4ng/πil). IL-12 (5ng/mι) and anti-IL4 (1 μg/ml) were used to direct to Thl, while IL-4 (5ng/ml) and anti-IFN gamma (1 μg/ml) were used to direct to Th2 and IL-10 at 5ng/ml was used to direct to Tri . After 4-5 days, the activated Thl , Th2 and Tri lymphocytes were washed once in DMEM and expanded for 4-7 days in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5xlO^"5M (Gibco), lOmM Hepes (Gibco) and IL-2 (lng/ml). Following this, the activated Thl, Th2 and Tri lymphocytes were re-stimulated for 5 days with anti-CD28/OKT3 and cytokines as described above, but with the addition of anti-CD95L (1 μg/ml) to prevent apoptosis. After 4-5 days, the Thl, Th2 and Tri lymphocytes were washed and then expanded again with IL-2 for 4-7 days. Activated Thl and Th2 lymphocytes were maintained in this way for a maximum of three cycles. RNA was prepared from primary and secondary Thl , Th2 and Tri after 6 and 24 hours following the second and third activations with plate bound anti-CD3 and anti-CD28 mAbs and 4 days into the second and third expansion cultures in Interleukin 2.

The following leukocyte cells lines were obtained from the ATCC: Ramos, EOL-1, KU-812. EOL cells were further differentiated by culture in O.lmM dbcAMP at 5xl0⁵cells/ml for 8 days, changing the media every 3 days and adjusting the cell concentration to 5xl0⁵cells/ml. For the culture of these cells, we used DMEM or RPMI (as recommended by the ATCC), with the addition of 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5xlO^"5M (Gibco), lOmM Hepes (Gibco). RNA was either prepared from resting cells or cells activated with PMA at 1 Ong/ml and ionomycin at 1 μg/ml for 6 and 14 hours. Keratinocyte line CCDl 06 and an airway epithelial tumor line NCI-H292 were also obtained from the ATCC. Both were cultured in DMEM 5% FCS (Hyclone), lOOμM non essential amino acids (Gibco), ImM sodium pyruvate (Gibco), mercaptoethanol 5.5xlO^"5M (Gibco), and lOmM Hepes (Gibco). CCDl 106 cells were activated for 6 and 14 hours with approximately 5 ng/ml TNF alpha and lng/ml IL-1 beta, while NCI-H292 cells were activated for 6 and 14 hours with the following cytokines: 5ng/ml IL-4, 5ng/rnl IL-9, 5ng/ml IL-13 and 25ng/ml IFN gamma.

For these cell lines and blood cells, RNA was prepared by lysing approximately 10⁷cells/ml using Trizol (Gibco BRL). Briefly, 1/10 volume of bromochloropropane

(Molecular Research Corporation) was added to the RNA sample, vortexed and after 10 minutes at room temperature, the tubes were spun at 14,000 rpm in a Sorvall SS34 rotor. The aqueous phase was removed and placed in a 15ml Falcon Tube. An equal volume of isopropanol was added and left at -20°C overnight. The precipitated RNA was spun down at 9,000 rpm for 15 min in a Sorvall SS34 rotor and washed in 70% ethanol. The pellet was redissolved in 300μl of RNAse-free water and 35μl buffer (Promega) 5μl DTT, 7μl RNAsin and 8μl DNAse were added. The tube was incubated at 37°C for 30 minutes to remove contaminating genomic DNA, extracted once with phenol chloroform and re-precipitated with 1/10 volume of 3M sodium acetate and 2 volumes of 100% ethanol. The RNA was spun down and placed in RNAse free water. RNA was stored at -80°C. AI_comprehensive panel vl.O

The plates for AI_comprehensive panel_vl.O include two control wells and 89 test samples comprised of cDNA isolated from surgical and postmortem human tissues obtained from the Backus Hospital and Clinomics (Frederick, MD). Total RNA was extracted from tissue samples from the Backus Hospital in the Facility at CuraGen. Total RNA from other tissues was obtained from Clinomics.

Joint tissues including synovial fluid, synovium, bone and cartilage were obtained from patients undergoing total knee or hip replacement surgery at the Backus Hospital. Tissue samples were immediately snap frozen in liquid nitrogen to ensure that isolated RNA was of optimal quality and not degraded. Additional samples of osteoarthritis and rheumatoid arthritis joint tissues were obtained from Clinomics. Normal control tissues were supplied by Clinomics and were obtained during autopsy of trauma victims. Surgical specimens of psoriatic tissues and adjacent matched tissues were provided as total RNA by Clinomics. Two male and two female patients were selected between the ages of 25 and 47. None of the patients were taking prescription drugs at the time samples were isolated.

Surgical specimens of diseased colon from patients with ulcerative colitis and Crohns disease and adjacent matched tissues were obtained from Clinomics. Bowel tissue from three female and three male Crohn's patients between the ages of 41-69 were used. Two patients were not on prescription medication while the others were taking dexamethasone, phenobarbital, or tylenol. Ulcerative colitis tissue was from three male and four female patients. Four of the patients were taking lebvid and two were on phenobarbital.

Total RNA from post mortem lung tissue from trauma victims with no disease or with emphysema, asthma or COPD was purchased from Clinomics. Emphysema patients ranged in age from 40-70 and all were smokers, this age range was chosen to focus on patients with cigarette-linked emphysema and to avoid those patients with alpha- lanti-trypsin deficiencies. Asthma patients ranged in age from 36-75, and excluded smokers to prevent those patients that could also have COPD. COPD patients ranged in age from 35-80 and included both smokers and non-smokers. Most patients were taking corticosteroids, and bronchodilators.

In the labels employed to identify tissues in the AI_comprehensive panel vl .0 panel, the following abbreviations are used:

Al = Autoimmunity

Syn = Synovial

Normal = No apparent disease

Rep22 /Rep20 = individual patients RA = Rheumatoid arthritis

Backus = From Backus Hospital

OA = Osteoarthritis

(SS) (BA) (MF) = Individual patients

Adj = Adjacent tissue Match control = adjacent tissues

-M = Male

-F = Female COPD = Chronic obstructive pulmonary disease

Panels 5D and 51

The plates for Panel 5D and 51 include two control wells and a variety of cDNAs isolated from human tissues and cell lines with an emphasis on metabolic diseases.

Metabolic tissues were obtained from patients enrolled in the Gestational Diabetes study. Cells were obtained during different stages in the differentiation of adipocytes from human mesenchymal stem cells. Human pancreatic islets were also obtained.

In the Gestational Diabetes study subjects are young (18 - 40 years), otherwise healthy women with and without gestational diabetes undergoing routine (elective) Caesarean section. After delivery of the infant, when the surgical incisions were being repaired/closed, the obstetrician removed a small sample (<1 cc) of the exposed metabolic tissues during the closure of each surgical level. The biopsy material was rinsed in sterile saline, blotted and fast frozen within 5 minutes from the time of removal. The tissue was then flash frozen in liquid nitrogen and stored, individually, in sterile screw-top tubes and kept on dry ice for shipment to or to be picked up by CuraGen. The metabolic tissues of interest include uterine wall (smooth muscle), visceral adipose, skeletal muscle (rectus) and subcutaneous adipose. Patient descriptions are as follows:

Patient 2: Diabetic Hispanic, overweight, not on insulin Patient 7-9: Nondiabetic Caucasian and obese (BMI>30)

Patient 10: Diabetic Hispanic, overweight, on insulin Patient 11 : Nondiabetic African American and overweight Patient 12: Diabetic Hispanic on insulin Adipocyte differentiation was induced in donor progenitor cells obtained from

Osirus (a division of Clonetics/BioWhittaker) in triplicate, except for Donor 3U which had only two replicates. Scientists at Clonetics isolated, grew and differentiated human mesenchymal stem cells (HuMSCs) for CuraGen based on the published protocol found in Mark F. Pittenger, et al., Multilineage Potential of Adult Human Mesenchymal Stem Cells Science Apr 2 1999: 143-147. Clonetics provided Trizol lysates or frozen pellets suitable for mRNA isolation and ds cDNA production. A general description of each donor is as follows:

Donor 2 and 3 U: Mesenchymal Stem cells, Undifferentiated Adipose Donor 2 and 3 AM: Adipose, AdiposeMidway Differentiated Donor 2 and 3 AD: Adipose, Adipose Differentiated Human cell lines were generally obtained from ATCC (American Type Culture Collection), NCI or the German tumor cell bank and fall into the following tissue groups: kidney proximal convoluted tubule, uterine smooth muscle cells, small intestine, liver HepG2 cancer cells, heart primary stromal cells, and adrenal cortical adenoma cells. These cells are all cultured under standard recommended conditions and RNA extracted using the standard procedures. All samples were processed at CuraGen to produce single stranded cDNA.

Panel 51 contains all samples previously described with the addition of pancreatic islets from a 58 year old female patient obtained from the Diabetes Research Institute at the University of Miami School of Medicine. Islet tissue was processed to total RNA at an outside source and delivered to CuraGen for addition to panel 51.

In the labels employed to identify tissues in the 5D and 51 panels, the following abbreviations are used:

GO Adipose = Greater Omentum Adipose SK = Skeletal Muscle

UT = Uterus

PL = Placenta

AD = Adipose Differentiated

AM = Adipose Midway Differentiated U = Undifferentiated Stem Cells

Panel CNSD.01

The plates for Panel CNSD.01 include two control wells and 94 test samples comprised of cDNA isolated from postmortem human brain tissue obtained from the Harvard Brain Tissue Resource Center. Brains are removed from calvaria of donors between 4 and 24 hours after death, sectioned by neuroanatomists, and frozen at -80°C in liquid nitrogen vapor. All brains are sectioned and examined by neuropathologists to confirm diagnoses with clear associated neuropathology.

Disease diagnoses are taken from patient records. The panel contains two brains from each of the following diagnoses: Alzheimer's disease, Parkinson's disease,

Huntington's disease, Progressive Supernuclear Palsy, Depression, and "Normal controls". Within each of these brains, the following regions are represented: cingulate gyrus, temporal pole, globus palladus, substantia nigra, Brodman Area 4 (primary motor strip), Brodman Area 7 (parietal cortex), Brodman Area 9 (prefrontal cortex), and Brodman area 17 (occipital cortex). Not all brain regions are represented in all cases; e.g., Huntington's disease is characterized in part by neurodegeneration in the globus palladus, thus this region is impossible to obtain from confirmed Huntington's cases. Likewise Parkinson's disease is characterized by degeneration of the substantia nigra making this region more difficult to obtain. Normal control brains were examined for neuropathology and found to be free of any pathology consistent with neurodegeneration. In the labels employed to identify tissues in the CNS panel, the following abbreviations are used:

PSP = Progressive supranuclear palsy Sub Nigra = Substantia nigra Glob Palladus= Globus palladus Temp Pole = Temporal pole

Cing Gyr = Cingulate gyrus BA 4 = Brodman Area 4

Panel CNS_Neurodegeneration_V1.0 The plates for Panel CNS_Neurodegeneration_V 1.0 include two control wells and

47 test samples comprised of cDNA isolated from postmortem human brain tissue obtained from the Harvard Brain Tissue Resource Center (McLean Hospital) and the Human Brain and Spinal Fluid Resource Center (VA Greater Los Angeles Healthcare System). Brains are removed from calvaria of donors between 4 and 24 hours after death, sectioned by neuroanatomists, and frozen at -80°C in liquid nitrogen vapor. All brains are sectioned and examined by neuropathologists to confirm diagnoses with clear associated neuropathology.

Disease diagnoses are taken from patient records. The panel contains six brains from Alzheimer's disease (AD) patients, and eight brains from "Normal controls" who showed no evidence of dementia prior to death. The eight normal control brains are divided into two categories: Controls with no dementia and no Alzheimer's like pathology (Controls) and controls with no dementia but evidence of severe Alzheimer's like pathology, (specifically senile plaque load rated as level 3 on a scale of 0-3; 0 = no evidence of plaques, 3 = severe AD senile plaque load). Within each of these brains, the following regions are represented: hippocampus, temporal cortex (Brodman Area 21), parietal cortex (Brodman area 7), and occipital cortex (Brodman area 17). These regions were chosen to encompass all levels of neurodegeneration in AD. The hippocampus is a region of early and severe neuronal loss in AD; the temporal cortex is known to show neurodegeneration in AD after the hippocampus; the parietal cortex shows moderate neuronal death in the late stages of the disease; the occipital cortex is spared in AD and therefore acts as a "control" region within AD patients. Not all brain regions are represented in all cases.

In the labels employed to identify tissues in the CNS_Neurodegeneration_V1.0 panel, the following abbreviations are used:

AD = Alzheimer's disease brain; patient was demented and showed AD-like pathology upon autopsy

Control = Control brains; patient not demented, showing no neuropathology

Control (Path) = Control brains; pateint not demented but showing sever AD-like pathology

SupTemporal Ctx = Superior Temporal Cortex

Inf Temporal Ctx = Inferior Temporal Cortex

A. CG100041-01: Trypsin Protease

Expression of gene CGI 00041-01 was assessed using the primer-probe sets Ag4360 and Ag4361, described in Tables AA and AB. Table AA. Probe Name Ag4360

Table AB. Probe Name Ag4361

General_screening_panel_vl.4 Summary: Ag4361 Expression of the CG100041-01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown).

Panel CNS_1 Summary: Ag4360 Expression of the CGI 00041-01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown). B. CG105716-01: germline oligomeric matrix protein

Expression of gene CGI 05716-01 was assessed using the primer-probe set Ag2362, described in Table BA. Results of the RTQ-PCR runs are shown in Tables BB, BC, BD, BE, BF, BG, BH and Bl.

Table BA. Probe Name Ag2362

Table BC. General_screeningjpanel_vl.5

Table BD. HASS Panel vl.O

Table BF. Panel 2D

Table BG. Panel 3D

Table BH. Panel 4D

AI_comprehensive panel_vl.0 Summary: Ag2362 Highest expression of the CG105716-01 gene is detected in cartilage from osteoarthritis patient (CT=19). In addition, high expression of this gene is also seen in synovium and bone samples from the osteoarthritis patient. Furthermore, low but significant expression of this gene is also detected in synovium, bone and cartilage samples of rheumatoid arthritis patients. The CGI 05716-01 gene codes for cartilage oligomeric matrix protein (COMP). COMP is a noncollagenous extracellular matrix (ECM) protein which consists of five identical glycoprotein subunits, each with EGF-like and calcium-binding (thrombospondin-like) domains. COMP has been implicated in inflammatory diseases including osteochondrodysplasias and arthritis (Neidhart et al., 1997, Br J Rheumatol 36(11):1151- 60, PMID: 9402858; Baitner et al, 2000, J Pediatr Orthop 20(5):594-605, PMID: 11008738; Clark et al., 1999, Arthritis Rheum 1999 Nov;42(l l):2356-64, PMID: 10555031). Therefore, therapeutic modulation of this gene product through the use of small molecule drugs, protein therapeutics or antibodies, might be beneficial in the treatment of inflammatory diseases such as rheumatoid and osteoarthritis, and osteochondrodysplasia.

General_screeningjpanel_vl.5 Summary: Ag2362 Highest expression of the CGI 05716-01 gene is detected in melanoma sample (CT=24). Thus, expression of this gene can be used to distinguish this sample from other samples in this panel. In addition, significant expression of this gene is seen in colon cancer tissue, a colon cancer, lung cancer, liver cancer, and CNS cancer cell line (CTs=31-34). The CGI 05716-01 gene codes for cartilage oligomeric matrix protein (COMP). Cartilage oligomeric matrix protein (COMP) is a noncollagenous extracellular matrix (ECM) protein which consists of five identical glycoprotein subunits, each with EGF-like and calcium-binding (thrombospondin-like) domains. COMP contains an RGD sequence. The RGD domain in other proteins has been shown to affect cell adhesion, migration , survival and proliferation.

Mutations of COMP can cause the osteochondrodysplasias pseudochondroplasia (PSACH) and multiple epiphyseal dysplasia (MED) (Kleerekoper et al., 2002, J Biol Chem 2002 Jan 8; [epub ahead of print], PMID: 11782471). Based on this profile, COMP may play a role in tumor cell growth and survival based upon the cells ability to interact with the extracellular matrix. Thus, therapeutic targeting with a human monoclonal antibody might block the interaction of cancer cells, or supporting sfromal elements, with extracellular matrix and thus promote cell death rather than cell survival especially in these cancers. Additionally, this gene is expressed in two melanoma cell lines that mimic some of characteristics of activated tumor endothelial cells. Hence, antibody directed against this gene may affect endothelial growth and survival in the tumor and prevent tumor growth.

In addition, recently COMP has also been implicated in vascular calcification and fibrosis especially associated with with advanced complicated atherosclerosis (Canfield et al., 2002, J Pathol 196(2):228-34, PMID: 11793375). Therefore, therapeutic modulation of this gene could also be beneficial in the treatement of vascular calcification and fibrosis.

Among tissues with metabolic or endocrine function, this gene is expressed at high to moderate levels in pancreas, adipose, thyroid, skeletal muscle, heart, and the gastrointestinal tract. Therefore, therapeutic modulation of the activity of this gene may prove useful in the treatment of endocrine/metabolically related diseases, such as obesity and diabetes.

HASS Panel vl.O Summary: Ag2362 The expression of this gene appears to be highest in astrocytes (Ct=28.95). There is a slight induction in expression of this gene when LnCAP cells are serum-starved and subjected to a reduced oxygen concentration and a decreased pH. These conditions resemble those typically found in tumors and suggest that in the tumors from which LnCAp cells are derived, expression of this gene may be regulated by these conditions.

Panel 1.3D Summary: Ag2362 Two experiments with same primer and probe set are in excellent agreement, with highest expression of the CG105716-01 gene in colon cancer ODO3866 sample (CTs=29). High expression of this gene are also associated with melanoma, and a liver cancer cell line. In addition, moderate expression of this gene is also seen adipose, brain, bone marrow, skeletal muscle heart, placenta, lung, testis and prostate. Please see panel 1.4 for the utility of this gene.

Panel 2D Summary: Ag2362 The expression of this gene appears to be highest in a sample derived from a breast cancer(CT=27). In addition, there appears to be substantial expression in other samples derived from breast cancer, gastric cancer, ovarian cancer, bladder cancer, thyroid cancer, kidney cancer, lung cancer, prostate cancer, liver cancer and colon cancer. Therapeutic modulation of this gene, through the use of small molecule drugs, protein therapeutics or antibodies could be of benefit in the treatment of breast, gastric, ovarian, bladder, thyroid, kidney, lung, prostate, liver or colon cancer. Panel 3D Summary: Ag2362 Highest expression of the CGI 05716-01 gene is detected in cerebellum (CT=27). Low to moderate expression of this gene is associated with small cell lung cancer, lung carcinoid, and osteosarcoma. Please see panel 1.4 for the utility of this gene.

Panel 4D Summary: Ag2362 Highest expression of the CG105716-01 gene is detected in IL4 treated dermal fibroblast cells (CT=29.2). High expression of this gene is seen in all the dermal fibroblast samples (CTs=29-31). Thus expression of this gene can be used to distinguish the dermal fibroblast from other samples used in this panel.

Furthermore, therapeutic modulation of this gene product could be beneficial in the treatment of skin disorders, including psoriasis. In addition, low to moderate expression of this gene is also with lung and colon.

Therefore therapeutic modulation of this gene could be useful in treatment of lung and colon related diseases such as lupus and glomerulonephritis, and inflammatory bowel diseases.

Panel 5D Summary: Ag2362 Highest expression of the CG105716-01 gene is detected in a adipose sample (CT=25). In addition, high expression of this gene is seen in other adipose samples, as well as skeletal muscle. Thus, expression of this gene could be used to distinguish this sample from other samples in this panel.

C. CG57415-01: neural cell adhesion protein

Expression of gene CG574I5-01 was assessed using the primer-probe sets Agl030, Ag3231, Ag971, Ag994 and Ag275, described in Tables CA, CB, CC, CD and CE. Results of the RTQ-PCR runs are shown in Tables CF, CG, CH, CI and CJ. Table CA. Probe Name Agl030

Table CB. Probe Name Ag3231

Table CC. Probe Name Ag971

Table CD. Probe Name Ag994

Table CE. Probe Name Ag275

Table CF. CNS_neurodegeneration_vl.O

Table CG. General_screening_panel_yl.4

Table CH. Panel 1

CNSjneurodegeneration vl.O Summary: Ag3231 The CG57415-01 gene is homologus to a neural cell adhesion molecule, a membrane-bound glycoprotein that plays a role in cell-cell and cell-matrix adhesion. NCAM related proteins, such as Nr-CAM, play a critical role in neurite extension. (Sakurai T. J Cell Biol 2001 Sep 17;154(6):1259-73) In addition, NCAMs are involved in plasticity mechanisms critical for learning, memory, and regeneration and have been implicated in brain pathology, including Alzheimer's disese. (Mikkonen M. Rev Neurosci 2001 ;12(4):311-25) Furthermore, this gene appears to be slightly downregulated in the temporal cortex of Alzheimer's patients when compared to expression in control brains. Therefore, therapeutic modulation of the expression or function of this gene may foster focal neurite outgrowth and have utility in therapeutically countering neurite degeneration of neurodegenerative diseases such as Alzheimer's, ataxias, and Parkinson's disease.

General_screening_panel_vl.4 Summary: Ag3231 The CG57415-01 gene is most highly expressed in a colon cancer cell line (CT=29.4) with sigmficant expression also seen in a breast cancer cell line. Thus, expression of this gene could be used to differentiate between these samples and other samples on this panel and as a marker to detect the presence of these cancers. Furthermore, therapeutic modulation of the expression or function of this protein may be useful in the treatment of colon and breast cancers.

Among tissues with metabolic function, this gene is expressed at moderate to low levels in pituitary, adipose, pancreas, thyroid, fetal liver and adult and fetal skeletal muscle and heart. This widespread expression among these tissues suggests that this gene product may play a role in normal neuroendocrine and metabolic function and that disregulated expression of this gene may contribute to neuroendocrine disorders or metabolic diseases, such as obesity and diabetes.

This gene also shows moderate to low expression in all regions of the CNS examined. Please see CNS_neurodegeneration_vl .0 for discussion of utility of this gene in the CNS. Panel 1 Summary: Ag275 Expression of the CG57415-01 gene is highest in samples derived from cerebellum (CT = 24.5) and testis (CT = 25.1). Thus, expression of this gene may be used to distinguish cerebellum and testis from other tissues. In addition, therapeutic modulation of this gene product, either through the use of purified protein to increase levels or through antibodies or small molecule drugs to inhibit function, might be of use to treat diseases of the testis, such as infertility or testicular cancer. However, expression of this gene is also detected in other samples on this panel, although expression is largely restricted to normal tissues. In addition to the high expression seen in cerebellum, this gene is also more moderately expressed in other CNS tissues including amygdala, hippocampus, substantia nigra, thalamus, hypothalamus and spinal cord. This gene shows homology to BIG-2, an axon-associated cell adhesion molecule (AxCAM) (Yoshihara Y. J Neurobiol 28:51-69 ). AxCAMs are critical for the development and maintenance of neural networks within the brain. In the response to injury and/or neuronal death, gene expression during the process of compensatory synaptogenesis in many ways mirrors that seen during development. Thus, the therapeutic expression of this gene or its protein product may be beneficial in the treatment of CNS injury (stroke, head trauma, spinal cord injury) or neurodegenerative diseases (Alzheimer's disease, Parkinson's disease, Huntington's disease, spinocerebellar ataxia, multiple sclerosis, ALS, or any disease resulting in neuronal atrophy or death).

The 30675585_EXT3 gene is also moderately expressed in all metabolic tissues on this panel ) including pancreas (CT = 29), adrenal gland (CT = 32), thyroid (CT = 28), heart (CT = 31), skeletal muscle (CT = 33), liver (CT = 30) and fetal liver (CT = 32). Therefore, this gene product may have a role in cell-cell communication in these tissues and thus be an antibody target for the treatment of diseases involving any or all of these tissues.

Panel 2.2 Summary: Ag3231 Expression of the CG57415-01 gene is highest in a sample derived from a kidney cancer (CT = 32.2), although the overall levels of expression are low. In addition, there is significant expression detected in samples derived from two breast cancer metastases and normal stomach. Overall this pattern of expression, suggests that this gene might be useful in distinguishing kidney, metastatic breast cancer and stomach from other tissues. In addition, therapeutic modulation of the function of this gene product might be of use in the treatment of metastatic breast cancer or kidney cancer. Panel 4D Summary: Ag3231 The CG57415-01 gene is expressed at low levels in normal thymus, lung, kidney and colon (CTs = 31 -32). Interestingly, there is lower expression in IBD colitis and Crohns disease samples as well as in lupus kidney, suggesting that this gene may play a role in these diseases. Thus, this gene may be used to distinguish normal kidney from lupus kidney as well as normal colon from colon affected by IBD or Crohns disease. In addition, this gene is expressed in an untreated eosinophil (EOL) cell line; however, EOL cells freated with PMA and ionomycin express this gene at much lower levels. This gene encodes a protein that is related to BIG2, a neural adhesion molecule. Transcript expression is detected primarily in untreated tissues and is down regulated upon inflammation. Based on t the function of BIG2 as an adhesion and signaling molecule, the 30675585_EXT3 protein may be important in the devlopment of normal organ stirucmre and on the normal trafficking of eosinophils from the bone marrow into peripheral tissues. Therapies using the protein encoded by this transcript may therefore be important in reducing inflammation or in wound healing; similar therapies using other adhesion molecules which encourge neurite outgrowth have been proposed ((Nogelezang M.G. J. Neurosci.21 : 6732-6744.) .

D. CG58504-01: ADAMTS12

Expression of gene CG58504-01 was assessed using the primer-probe set Ag2475, described in Table DA. Results of the RTQ-PCR runs are shown in Tables DB, DC, DD, DE and DF.

Table DA. Probe Name Ag2475

Table DB. HASS Panel vl.O

Table DC. Panel 1.3D

HASS Panel vl.O Summary: Ag2475 This gene is expressed in glioma samples and primary asfrocytes in culture (highest expression CT=27.8) suggesting a role in cell growth. Expression of this gene in U87-MG (a mixed glial/asfrocytoma cell line) is repressed by reducing the oxygen content of the environment. Serum starvation of these cells induces expression. This effect is not observed in T24 (bladder cancer) cells and thus may reflect tissue specific regulation of this gene.

Panel 1.3D Summary: Ag2475 Highest expression of the CG58504-01 gene is seen in fetal skeletal muscle (CT=28.4). This expression is significantly higher than expression seen in the corresponding adult tissue (CT=36.9). Thus, expression of this gene could be used to differentiate between the fetal and adult sources of this tissue. In addition, the relative overexpression of this gene in fetal skeletal muscle suggests that the protein product may enhance muscular growth or development in the fetus and thus may also act in a regenerative capacity in the adult. Therefore, therapeutic modulation of the protein encoded by this gene could be useful in treatment of muscle related diseases. More specifically, freatment of weak or dystrophic muscle with the protein encoded by this gene could restore muscle mass or function.

Low levels of expression are also seen in other metabolic tissues, including adipose and fetal heart, suggesting a potential role for this gene in obesity and/or diabetes. Moderate levels of expression are also seen in cell lines derived from brain cancer, breast cancer, renal cancer, lung cancer, colon cancer and melanoma. Since cell lines and fetal tissues are, on the whole, more proliferative than normal tissues, this expression profile suggests that this gene might be involved in cell proliferation. Therefore, modulation of the expression or function of this gene may be a therapeutic avenue for the treatment of cancer or other disease that involve cell proliferation. Furthermore, therapeutic targeting of this gene product with a monoclonal antibody is anticipated to limit or block the extent of tumor cell migration and invasion and tumor metastasis, particularly in brain cancer, breast cancer, renal cancer, lung cancer, colon cancer and melanoma. This gene might also be an effective marker for the diagnosis and detection of these cancers.

Panel 2D Summary: Ag2475 Highest expression of the CG58504-01 gene is seen in a lung cancer (CT=28.3). This gene encodes a putative member of the ADAMS family. The ADAMS family of proteins has multiple domains associated with function; A fibronectin domain involved cell/extracellular matrix interaction, a thrombospondin domain involved in angiogenesis and a metalloproteinase domain involved in matrix degredation. This multi-domain structure has implications for this molecule in several tumorigenic processes, including invasion and metastasis and proliferation and cell survival. Thus, the metalloproteinase domain might play a role in cell invasion and metastasis, the fibronectin domain may play a role in cell adhesion or survival and the thrombospondin domain might play a role in angiogenesis. ADAM 12-S cleaves insulinlike growth factor binding protein-3 (IGFBP-3). IGFBP-3 enhances the p53-dependent apoptotic response of colorectal cells to DNA damage. IGF-BP3 is inversely, associated with risk for colorectal cancer. Expression of IGFBP-3 induces Growth inhibition and differentiation of the human colon carcinoma cell line, Caco-2. All these data indicate that CG58504-01 may act by cleaving and inactivating IGFBP-3 limiting its anti-tumor activity.

Thus, therapeutic targeting with a human monoclonal antibody of CG58504-01 may inhibit any or all of the listed activities therefore blocking the angiogenic, invasion/metastasis or growth/survival promoting activities of this molecule especially in those cancer types, like colon, lung, kidney, bladder ovarian and gastric tumors where the gene is overexpressed in the tumor compared to the normal adjacent tissue.

Panel 3D Summary: Ag2475 Highest expression of the CG58504-01 gene is seen in a leiomyosarcoma cell line (CT=30.4). Significant levels of expression are also seen in other cell lines including samples derived from bladder, ovarian, lung and brain cancers. Thus, expression of this gene could be used to differentiate these samples from other samples on this panel. Please see Panel 2D for detailed discussion of utility of this gene in cancer. Panel 4D Summary: Ag2475 Highest expression of the CG58504-01 gene is seen in resting coronary artery smooth muscle cells (CT=27.3). Moderate to low levels of expression are also seen in resting astrocytes and TNFalpha + IL-lbeta treated astrocytes and coronary artery smooth muscle cells, TNF alpha and IL-4 treated dermal fibroblasts, and lung. Lower levels of expression are seen in treated and untreated lung fibroblasts. This expression suggests that this gene may be a marker of smooth muscle. In addition, expression in fibroblasts and asfrocytes suggests that this gene product may be involved in inflammatory conditions that involve these cells. This gene encodes a putative ADAMTS molecule which has been implicated in extracellular proteolysis and may play a critical role in the tissue degradation seen in arthritis and other inflammatory conditions. (Kuno K. : J Biol Chem 1997 Jan 3;272(l):556-62) Therefore, therapeutic modulation of this gene product may be useful in the treatment of pathological and inflammatory lung and skin disorders that include chronic obstructive pulmonary disease, asthma, allergy, psoriasis and emphysema. Panel 5 Islet Summary: Ag2475 Results from one experiment with the CG58504-01 gene are not included. The amp plot indicates that there were experimental difficulties with this run.

E. CG58586-01 and CG58586-02: CASPR4B

Expression of gene CG58586-01 and variant CG58586-02 was assessed using the primer-probe set Ag3379, described in Table EA. Results of the RTQ-PCR runs are shown in Tables EB, EC, ED and EE.

Table EA. Probe Name Ag3379

Table EC. General_screening_panel_vl.4

Table ED. Panel 4D

Table EE. general oncology screening panel_y_2.4

CNS__neurodegeneration_vl.O Summary: Ag3379 This panel confirms the expression of the CG58586-01 gene at significant levels in the brain in an independent group of individuals. This gene is found to be slighltly upregulated in the temporal cortex of Alzheimer's disease patients. Blockade of this receptor may be of use in the treatment of this disease and decrease neuronal death.

General_screeningjpanel_yl.4 Summary: Ag3379 Highest expression of the CG58586-01 is detected in spinal cord sample (CT=26.3). In addition, high expression of this gene is exclusively seen in all the region of central nervous system examined including amygdala, hippocampus, substantia nigra, thalamus, cerebellum, cerebral cortex, and spinal cord. The CG58586-01 gene codes for contactin associated protein-like 4 precursor (Cell recognition molecule, Caspr4). Caspr (paranodin) family of proteins play a cenfral role in in the assembly of multiprotein complexes necessary for the formation and maintenance of paranodal junctions (Denisenko-Nehrbass et al., 2002, J Physiol Paris 96(l-2):99-103, PMID: 11755788). Therefore, therapeutic modulation of this gene could be useful in the treatment of central nervous system disorders such as Alzheimer's disease, Parkinson's disease, epilepsy, multiple sclerosis, schizophrenia and depression. In addition, significant expression of this gene is seen in a colon cancer and two lung cancer cell lines. Therefore, therapeutic modulation of the activity of this gene or its protein product, through the use of small molecule drugs, protein therapeutics or antibodies, might be beneficial in the treatment of lung cancer or colon cancer.

This gene also shows moderate expression in fetal liver and skeletal muscle (CTs=31). Interestingly, this gene is expressed at much higher levels in fetal when compared to adult liver and skeletal muscle (CTs=35-40). This observation suggests that expression of this gene can be used to distinguish fetal from the coπesponding adult tissue. In addition, the relative overexpression of this gene in fetal tissue suggests that the protein product may enhance growth or development of liver and skeletal muscle in the fetus and thus may also act in a regenerative capacity in the adult. Therefore, therapeutic modulation of the protein encoded by this gene could be useful in freatment of muscle and liver related diseases.

Panel 4D Summary: Ag3379 Highest expression of the CG58586-01 is detected exclusively in Ramos B cells (CTs=27-28). Thus, expression of this gene can be used to distinguish the Ramos B cells from other samples used in this panel. B cells represent a principle component of immunity and contribute to the immune response in a number of important functional roles, including antibody production. Production of antibodies against self-antigens is a major component in autoimmune disorders. In addition, low but significant expression of this gene is also seen in thymus (CT=33). Therefore, therapeutic modulation of this gene product may reduce or eliminate the symptoms of patients suffering from asthma, allergies, chronic obstructive pulmonary disease, emphysema, Crohn's disease, ulcerative colitis, rheumatoid arthritis, psoriasis, osteoarthritis, systemic lupus erythematosus and other autoimmune disorders.

In addition, low but significant expression of this gene is also seen in colon samples (CT=34), Interestingly, expression of this gene is decreased in colon samples from patients with IBD colitis and Crohn's disease (CTs=36-37)relative to normal colon. Therefore, therapeutic modulation of the activity of the protein encoded by this gene may be useful in the freatment of inflammatory bowel disease. general oncology screening panel_v_2.4 Summary: Ag3379 Highest expression of the CG58586-01 is detected exclusively in lung cancer (OD06850-03C) sample (CT=30.5). In addition, low levels of expression of this gene is also seen in two of the metastatic melanoma samples. Therefore, expression of this gene may be used as diagnostic marker for detection of lung and metastatic melanoma. Furthermore, therapeutic modulation of this gene product may be beneficial in the treatment of lung cancer and melanoma.

F. CG93453-01 and CG93453-02: ADAM-TS 3 PRECURSOR (KIAA0366)

Expression of gene CG93453-01 and full length clone CG93453-02 was assessed using the primer-probe set Ag2085, described in Table FA. Results of the RTQ-PCR runs are shown in Tables FB, FC and FD.

Table FA. Probe Name Ag2085

Table FC. Panel 2D

Panel 1.3D Summary: Ag2085 Highest expression of the CG93453-01 gene, an ADAM TS3 homolog, is seen in a renal cancer cell line (CT=30.1). Thus, expression of this gene could be used to differentiate between this sample and other samples on this panel. Low but significant levels of expression are also seen in cell lines derived from brain, lung, breast, ovarian, and melanoma cancers. Thus, therapeutic modulation of the expression or function of this gene may also be effective in the treatment of these cancers.

This gene is also expressed at low levels in the CNS, including the hippocampus, and amygdala. Therefore, therapeutic modulation of the expression or function of this gene may be useful in the treatment of neurologic disorders, such as Alzheimer's disease, Parkinson's disease, schizophrenia, multiple sclerosis, stroke and epilepsy.

Among tissues with metabolic function, this gene is expressed at low levels in adipose, thyroid, and fetal liver. This expression suggests that this gene product may play a role in normal neuroendocrine and metabolic function and that disregulated expression of this gene may contribute to neuroendocrine disorders or metabolic diseases, such as obesity and diabetes.

Panel 2D Summary: Ag2085 Highest expression of the CG93453-01 gene is seen in a kidney cancer (CT=29.5), in agreement with expression in Panel 1.3D. Significant levels of expression are also seen in kidney cancer, breast cancer and gastric cancer. Thus, expression of this gene could be used to differentiate between the renal cancer and other samples on this panel, especially normal kidney tissue. The ADAMS family of proteins has multiple domains associated with function, including a thrombospondin domain involved in angiogenesis and a metalloproteinase domain involved in matrix degredation. This multi-domain structure has implications for this molecule in several tumorigenic processes, including invasion and metastasis and proliferation and cell survival. Thus, the metalloproteinase domain might play a role in cell invasion and metastasis, and the thrombospondin domain might play a role in angiogenesis. Therefore, therapeutic modulation of the expression or function of this gene may also be effective in the treatment of kidney cancer, breast cancer and gastric cancer. Panel 4D Summary: Ag2085 Highest expression of the CG93453-01 gene is seen in the KU-812 basophil cell line treated with PMA/ionomycin (CT=26.3). This transcript appears to be induced in the PMA and ionomycin treated basophil cell line, when compared to expression in resting basophils (CT=29.4). Basophils release histamines and other biological modifiers in reponse to allergens and play an important role in the pathology of asthma and hypersensitivity reactions. In addition, this gene encodes a putative ADAMTS molecule which has been implicated in extracellular proteolysis and may play a critical role in the tissue degradation seen in arthritis and other inflammatory conditions. (Kuno K. : J Biol Chem 1997 Jan 3;272(l):556-62) Therefore, therapeutics designed against the putative protein encoded by this gene may reduce or inhibit inflammation by blocking basophil function in these diseases. In addition, these cells are a reasonable model for the inflammatory cells that take part in various inflammatory lung and bowel diseases, such as asthma, Crohn's disease, and ulcerative colitis. Therefore, therapeutics that modulate the function of this gene product may reduce or eliminate the symptoms of patients suffering from asthma, Crohn's disease, and ulcerative colitis.

G. CG95145-01: Clq-related Gliacolin

Expression of gene CG95145-01 was assessed using the primer-probe set Ag4503, described in Table GA. Results of the RTQ-PCR runs are shown in Tables GB, GC and GD.

Table GA. Probe Name Ag4503

Table GB. CNS_neurodegeneration_vl.O

Table GC. General_screening_panel_vl.4

Table GD. Panel 4.1D

CNS_neurodegeneration_vl.0 Summary: Ag4503 Two experiments with the same probe and primer set produce results that are in excellent agreement. Highest expression of the CG54503-05 gene is seen in the parietal cortex of a control patient (CTs=27-28.6). This protein is found to be down-regulated in the temporal cortex of Alzheimer's disease patients. This protein appears to be a member of the complement family, specifically containing a Clq domain and homology to Clq-related factor. Clq is a subunit of the complex that activates the complement system. The complement system has been implicated in Alzheimer's disease because complement proteins are found in senile plaques and neuroinflammation in response to plaques appears to be a major cause of neuronal death in AD. Therefore, up-regulation of this gene or its protein product may be of use in reversing the dementia/memory loss and neuronal death associated with this disease.

References:

Lue LF, Rydel R, Brigham EF, Yang LB, Hampel H, Murphy GM Jr, Brachova L, Yan SD, Walker DG, Shen Y, Rogers J. Inflammatory repertoire of Alzheimer's disease and nondemented elderly microglia in vitro. Glia 2001 Jul;35(l):72-9

H. CG95250-01 and CG95250-02: Aminopeptidase N - isoform 2

Expression of gene CG95250-01 and variant CG95250-02 was assessed using the primer-probe sets Agl355 and Ag4501, described in Tables HA and HB. Results of the RTQ-PCR runs are shown in Tables HC, HD, HE, HF, HG and HH. Please note that the probe and primer set Ag4501 corresponds to the CG95250-02 variant only.

Table HA. Probe Name Agl355

Table HB. Probe Name Ag4501

Table HP. CNS_neurodegeneration_vl.O

Table HE. General_screening_panel_vl.4

Table HF. Panel 1.2

Table HG. Panel 2.2

Table HH. Panel 4. ID

AI_comprehensive panel vl.O Summary: Agl355 Low to moderate levels of expression of the CG95250-01 gene are detected in most of the samples used in this panel, with highest expression in a psoriasis sample (CT=27). Significant expression of this gene is also detected in bone, cartilage, synovium and synovia! fluid samples, normal lung samples, COPD lung, emphysema, atopic asthma, asthma, allergy, Crohn's disease (normal matched confrol and diseased), ulcerative colitis(normal matched confrol and diseased), and psoriasis (normal matched control and diseased). Therefore, therapeutic modulation of this gene product may ameliorate symptoms/conditions associated with autoimmune and inflammatory disorders including psoriasis, allergy, asthma, inflammatory bowel disease, rheumatoid arthritis and osteoarthritis

CNS_neurodegeneration_vl.O Summary: Agl355/Ag4501 Two experiment with different probe and primer sets are in excellent agreement with highest expression of the CG95250-01 gene in a temporal cortex sample derived from an Alzheimer's disease patient (CT=31.5). This gene is found to be slighltly upregulated in the temporal cortex of Alzheimer's disease patients. Therefore, therapeutic modulation of this gene product may be of useful in the freatment of this disease and decrease neuronal death.

General_screening_panel_vl.4 Summary: Agl355/Ag4501 Two experiment with different probe and primer sets are in excellent agreement with highest expression of the CG95250-01 gene in placenta and an ovarian cancer cell line (CTs=30). Therefore, therapeutic modulation of this gene product may be useful in treatment of reproductive disorders and ovarian cancer.

In addition, significant expression of this gene is seen in a ovarian cancer, breast cancer, lung cancer, pancreatic cancer, and colon cancer cell lines. The CG95250-01 gene codes for aminopeptidase N (APN) like protein. Recently, APN has shown to play a role in cell motility and angiogenesis, and it is a useful indicator of a poor prognosis for node- positive patients with colon cancer (Hashida et al., 2002, Gasfroenterology 2002 Feb;122(2):376-86, PMID: 11832452). Therefore, therapeutic modulation of the protein encoded by this gene, through the use of small molecule drugs, protein therapeutics or antibodies, could be useful in the freatement of these cancers.

Among tissues with metabolic or endocrine function, this gene is expressed at high to moderate levels in adipose, adrenal gland, skeletal muscle, and stomach. Therefore, therapeutic modulation of the activity of this gene may prove useful in the treatment of endocrine/metabolically related diseases, such as obesity and diabetes. Results from one experiment (run 213323381) with the CG95250-01 gene are not included. The amp plot indicates that there were experimental difficulties with this run.

Panel 1.2 Summary: Agl355 Highest expression of the CG95250-01 gene is seen in placenta (CT=22). In addition, significant expression of this gene is seen in a ovarian cancer, breast cancer, lung cancer, pancreatic cancer, prostate cancer, renal cancer, CNS cancer, melanoma and colon cancer cell lines. Among tissues with metabolic or endocrine function, this gene is expressed at high to moderate levels in pancrease, liver, heart, adrenal gland, skeletal muscle, small intestine and stomach. Please see panel 1.4 for discussion on the potential utility of this gene.

In addition, this gene is expressed at high levels in all regions of the central nervous system examined, including amygdala, hippocampus, substantia nigra, thalamus, cerebellum, cerebral cortex, and spinal cord. Therefore, this gene may play a role in cenfral nervous system disorders such as Alzheimer's disease, Parkinson's disease, epilepsy, multiple sclerosis, schizophrenia and depression. Panel 2.2 Summary: Agl355 Highest expression of the CG95250-01 gene is seen in ovarian margin sample (CT=32.7). Low but significant expression of this gene is also seen in ovarian cancer, normal breast and the cancer metastasis, and in normal liver samples. Please see panel 1.4 for the discussion of the utility of the gene. Panel 4.1D Summary: Agl355 Highest expression of the CG95250-01 gene is seen in IL-4 treated dermal fibroblast sample (CT=34). In addition, significant expression of this gene is also detected in thymus, TNFalpha + ILl beta freated bronchial epithelium, LPS freated monocytes and resting dermal fibroblasts. LPS freated monocytes contribute to the innate and specific immunity by migrating to the site of tissue injury and releasing inflammatory cytokines. Cytokine activated epithelial and dermal fibroblast cells contribute to the inflammation process. The CG95250-01 gene codes for aminopeptidase N (APN) like protein. APN is shown to induce chemotactic migration of leukocytes (Tani et al., 2001, J Med Invest 48:133-41). Thus, APN-induced leukocyte chemotaxis and activation may play an important role in immunologic events of inflammatory and allergic diseases.

Ag4501 Expression of this gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown).

I. CG95430-01: AdipoQ-like

Expression of gene CG95430-01 was assessed using the primer-probe set Ag4020, described in Table IA. Results of the RTQ-PCR runs are shown in Tables IB, IC, ID and IE.

Table IA. Probe Name Ag4020

Table IB. CNS_neurodegeneration_vl.O

Table ID. Panel 5 Islet

Table IE. general oncology screening panel_v_2.4

CNS_neurodegeneration_vl.O Summary: Ag4020 This panel does not show differential expression of the CG95430' 01 gene in Alzheimer's disease. However, this expression profile confirms the presence of this gene in the brain, with highest expression in the hippocampus of an Alzheimer's patient (CT=31.4). Therefore, therapeutic modulation of the expression or function of this gene may be useful in the freatment of neurologic disorders, such as Alzheimer's disease, Parkinson's disease, schizophrenia, multiple sclerosis, stroke and epilepsy.

General_screeningjpanel_vl.4 Summary: Ag4020 Results from one experiment with the CG95430-01 gene are not included. The amp plot indicates that there were experimental difficulties with this run.

Panel 4.1D Summary: Ag4020 The CG95430-01 gene is most highly expressed in kidney (CT=32.3). Low but significant levels of expression are also seen in untreated and cytokine activated lung fibroblasts, and thymus. This expression profile suggests that this gene may be involved in the homeostasis of the lung, thymus, and kidney. Expression of this gene appears to be slightly downregulated in cytokine activated lung fibroblasts suggesting that modulation of this gene product may help to maintain or restore function to the lung during inflammation.

Panel 5 Islet Summary: Ag4020 The CG95430-01 gene is expressed in adipose and skeletal muscle (CTs=31,8-34). This gene encodes a putative adiponectin (also known as adipocyte complement-related protein (ACRP-30), AdipoQ, apMl (adipose most abundant transcript 1) or GBP28 (28 kDa gelatin binding protein)), a member of the Clq family. This protein is induced over 100-fold in adipocyte differentiation (Scherer et al., J Biol Chem 1995 Nov 10;270(45):26746-9) and is involved in adipocyte signaling (Hu et al., J Biol Chem 1996 May 3;271(18):10697-703). Like other members of the Clq family, it forms a homotrimer and the crystal structure indicates that it likely arose from tumor necrosis factor (TNF; Shapiro and Scherer,Cuπ Biol 1998 Mar 12;8(6):335-8). Ionomycin increases expression of adiponectin and dibutyryl cAMP and TNF-alpha reduce expression and secretion in 3T3-L1 adipocytes (Kappes and Loffler, Horm Metab Res 2000 Nov-Dec;32(l l-12):548-54). Levels of adiponectin are decreased in obese humans (Arita et al., Biochem Biophys Res Commun 1999 Apr 2;257(l):79-83) and mice (Hu et al., J Biol Chem 1996 May 3;271(18):10697-703). A proteolytic cleavage product of adiponectin is reported to increase fatty acid oxidation in muscle and causes weight loss in mice. (Fruebis et al., Proc Natl Acad Sci U S A 2001 Feb 13;98(4):2005-10). A missense mutation in the protein was coπelated with a markedly low plasma adiponectin level (Takahashi et al., Int J Obes Relat Metab Disord 2000 Jul;24(7):861-8). Recent papers have shown that adiponectin reverses insulin resistance in mouse models of lipoatrophy and obesity (Yamauchi et al., Nature Med 2000; 7(8): 941-6), and that it enhances insulin action on the liver (Berg et al., ibid, 947-53). In addition, circulating levels of adiponectin have been shown to be lower in obese than in lean subjects and lower in diabetic patients than in non-diabetic patients, with particularly low levels in subjects with coronary artery disease. Furthermore, in patients who were subjected to a weight loss program that resulted in a 10% reduction of their body mass index, circulating adiponectin levels increased significantly. (Berg AH. Trends Endocrinol Metab. 2002 Mar; 13 (2): 84-9) Therefore based on its homology to adiponectin and its expression profile, this protein may function as a potential therapeutic for the treatment of obesity, type II diabetes and/or their secondary complications.

Adiponectin also seems to have additional cardiovascular and immune system effects. Levels of this protein are reduced in a cohort of Japanese patients with coronary artery disease (CAD), which correlates with the modulation of endothelial adhesion molecules on treatment of human aortic endothelial cells with adiponectin (Ouchi et al., Circulation 1999 Dec 21-28;100(25):2473-6). This protein is found adhering to vascular walls after injury (Okamoto et al. Horm Metab Res 2000 Feb;32(2):47-50) and presence of adiponectin suppresses the macrophage to foam cell transformation (Ouchi et al., Circulation 2001 Feb 27;103(8):1057-63).In addition, levels of adiponectin were lower in diabetic subjects with CAD relative to non-diabetic subjects or diabetic subjects without CAD (Hotta et al., Arterioscler Thromb Vase Biol 2000 Jun;20(6): 1595-9), indicating that lower levels of adiponectin may be an indicator of macroangiopathy in diabetes. Moreover, this protein negatively regulates the growth of myelomonocytic precursors (in part by inducing apoptosis) and macrophage function (Yokota et al., Blood 2000 Sep 1;96(5): 1723-32). This effect seems to be via the complement 1Q receptor ClqRp.

The Clq family of proteins involves members such as the complement subunit Clq, gliacolin, Clq-related protein, cerebellin, CORS26 etc., all of which are secreted. They show the presence of a common domain, the Clq domain, at the C terminus and collagen triple helix repeats at the C terminus. The repeats enable the proteins to form homotrimers and possibly oligomers. Members of this family have been implicated in tissue differentiation, immune regulation, energy homeostasis, synaptic function and in diseases such as obesity and neurodegeneration. Therefore, therapeutic modulation of the expression or function of this gene through the use of monoclonal antibodies may be useful in the prevention and/or treatment of obesity and diabetes. Furthermore, development of human monoclonal antibodies which inhibit this Adipo-Q like protein may also prove useful in the therapeutic treatment of cachexia that occurs in many forms of cancer. References:

Biol Chem 1995 Nov 10;270(45):26746-26749, A novel serum protein similar to Clq, produced exclusively in adipocytes. Scherer PE, Williams S, Fogliano M, Baldini G, Lodish HF.

We describe a novel 30-kDa secretory protein, Acrp30 (adipocyte complement- related protein of 30 kDa), that is made exclusively in adipocytes and whose mRNA is induced over 100-fold during adipocyte differentiation. Acrp30 is structurally similar to complement factor Clq and to a hibernation-specific protein isolated from the plasma of Siberian chipmunks; it forms large homo-oligomers that undergo a series of post- translational modifications. Like adipsin, secretion of Acrp30 is enhanced by insulin, and Acrp30 is an abundant serum protein. Acrp30 may be a factor that participates in the delicately balanced system of energy homeostasis involving food intake and carbohydrate and lipid catabolism. Our experiments also further corroborate the existence of an insulin- regulated secretory pathway in adipocytes.

J Biol Chem 1996 May 3;271(18):10697-10703, AdipoQ is a novel adipose- specific gene dysregulated in obesity. Hu E, Liang P, Spiegelman BM.

Adipose differentiation is accompanied by changes in cellular morphology, a dramatic accumulation of intracellular lipid and activation of a specific program of gene expression. Using an mRNA differential display technique, we have isolated a novel adipose cDNA, termed adipoQ. The adipoQ cDNA encodes a polypeptide of 247 amino acids with a secretory signal sequence at the amino terminus, a coUagenous region (Gly- X-Y repeats), and a globular domain. The globular domain of adipoQ shares sigmficant homology with subunits of complement factor Clq, collagen alpha 1(X), and the brain- specific factor cerebellin. The expression of adipoQ is highly specific to adipose tissue in both mouse and rat. Expression of adipoQ is observed exclusively in mature fat cells as the sfromal-vascular fraction of fat tissue does not contain adipoQ mRNA. In cultured 3T3-F442A and 3T3-L1 preadipocytes, hormone-induced differentiation dramatically increases the level of expression for adipoQ. Furthermore, the expression of adipoQ mRNA is significantly reduced in the adipose tissues from obese mice and humans. Whereas the biological function of this polypeptide is presently unknown, the tissue- specific expression of a putative secreted protein suggests that this factor may function as a novel signaling molecule for adipose tissue.

Horm Metab Res 2000 Nov;32(l l-12):548-554, Influences of ionomycin, dibutyryl-cycloAMP and tumour necrosis factor-alpha on intracellular amount and secretion of apMl in differentiating primary human preadipocytes. Kappes A, Loffler G.

3T3-L1 -adipocytes produce the adipocyte complement related protein of 30 kD (Acrp30), which is also designated as AdipoQ. In order to study the expression and secretion of the human homologue of this protein, apMl (adipose Most abundant gene transcript 1 , also named gelatin-binding protein of 28 kD [GBP28] or adiponectin), a polyclonal antibody was produced. Both expression and secretion can be detected beginning with day 4 after induction of differentiation. The amount of expressed apMl coπelates with the specific activity of the differentiation marker glycerol-3 -phosphate dehydrogenase. Secretion of apMl is increased by the addition of ionomycin. Both the nonhydrolysable dibutyryl-cycloAMP and tumour necrosis factor alpha reduce the expression and secretion of apMl.

Biochem Biophys Res Commun 1999 Apr 2;257(l):79-83, Paradoxical decrease of an adipose-specific protein, adiponectin, in obesity. Arita Y, Kihara S, Ouchi N, Takahashi M, Maeda K, Miyagawa J, Hotta K, Shimo ura I, Nakamura T, Miyaoka K, Kuriyama H, Nishida M, Yamashita S, Okubo K, Matsubara K, Muraguchi M, Ohmoto Y, Funahashi T, Matsuzawa Y.

We isolated the human adipose-specific and most abundant gene transcript, apMl (Maeda, K., et al., Biochem. Biophys. Res. Commun. 221, 286-289, 1996). Tne apMl gene product was a kind of soluble matrix protein, which we named adiponectin. To quantitate the plasma adiponectin concentration, we have produced monoclonal and polyclonal antibodies for human adiponectin and developed an enzyme-linked immunosorbent assay (ELISA) system. Adiponectin was abundantly present in the plasma of healthy volunteers in the range from 1.9 to 17.0 mg/ml. Plasma concentrations of adiponectin in obese subjects were significantly lower than those in non-obese subjects, although adiponectin is secreted only from adipose tissue. The ELISA system developed in this study will be useful for elucidating the physiological and pathophysiological role of adiponectin in humans. Nat Med 2001 Aug;7(8):941-946, The fat-derived hormone adiponectin reverses insulin resistance associated with both lipoatrophy and obesity. Yamauchi T, Ka on J, Waki H, Terauchi Y, Kubota N, Hara K, Mori Y, Ide T, Murakami K, Tsuboyama- Kasaoka N, Ezaki O, Akanuma Y, Gavrilova O, Vinson C, Reitman ML, Kagechika H, Shudo K, Yoda M, Nakano Y, Tobe K, Nagai R, Kimura S, Tomita M, Froguel P, Kadowaki T.

Adiponectin is an adipocyte-derived hormone. Recent genome- wide scans have mapped a susceptibility locus for type 2 diabetes and metabolic syndrome to chromosome 3q27, where the gene encoding adiponectin is located. Here we show that decreased expression of adiponectin correlates with insulin resistance in mouse models of altered insulin sensitivity. Adiponectin decreases insulin resistance by decreasing triglyceride content in muscle and liver in obese mice. This effect results from increased expression of molecules involved in both fatty-acid combustion and energy dissipation in muscle. Moreover, insulin resistance in lipoatrophic mice was completely reversed by the combination of physiological doses of adiponectin and leptin, but only partially by either adiponectin or leptin alone. We conclude that decreased adiponectin is implicated in the development of insulin resistance in mouse models of both obesity and lipoatrophy. These data also indicate that the replenishment of adiponectin might provide a novel freatment modality for insulin resistance and type 2 diabetes. general oncology screening panel_v_2.4 Summary: Ag4020 The CG95430-01 gene is most highly expressed in a metastatic melanoma (CT=32.7). Significant levels of expression are also seen in a lung cancer and a kidney cancer when compared to normal adjacent tissue. Thus, expression of this gene may be useful as a diagnostic marker of the presence of these cancers. Furthermore, therapeutic modulation of the expression or function of this gene may be useful in the treatment of kidney cancer, lung cancer, and melanoma. J. CG95794-01: TRYPSIN III, CATIONIC PRECURSOR

Expression of gene CG95794-01 was assessed using the primer-probe set Ag4029, described in Table JA. Results of the RTQ-PCR runs are shown in Table JB. Table JA. Probe Name Ag4029

CNS_neurodegeneration_vl.O Summary: Ag4029 Expression of the CG95794- 01 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.)

General_screeningjpanel_vl.4 Summary: Ag4029 Expression of the CG95794- 01 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.) Panel 4.1D Summary: Ag4029 Significant expression of the CG95794-01 gene, which encodes a trypsin homolog, is limited to the kidney and thymus (CTs=32-33). Adminisfration of trypsin has been shown to decrease the presence of TGF-betal in the kidney, a significant factor in the development of diabetic nephropathy (Paczek L. Drugs Exp Clin Res 2001;27(4):141-9). Thus, based on this selective expression profile, expression of this gene could be used as to differentiate between these samples and other samples on this panel and as a marker of these tissues. Furthermore, therapeutic modulation of the expression or function of this gene product may be useful in maintaining or restoring function to these organs during inflammation or disease, specifically diabetes.

K. CG95804-01: KALLIKREIN

Expression of gene CG95804-01 was assessed using the primer-probe set Ag4030, described in Table KA. Results of the RTQ-PCR runs are shown in Table KB. Table KA. Probe Name Ag4030

Table KB. Panel 4. ID

CNS_neurodegeneration_vl.O Summary: Ag4030 Expression of the CG95804-

01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown). General_screening_panel_vl.4 Summary: Ag4030 Expression of the CG95804- 01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown).

Panel 4.1D Summary: Ag4030 Highest expression of the CG95804-01 gene is detected exclusively in IL-13 treated NCI-H292 (CT=33). Thus, expression of this gene can be used to distinguish this sample from other samples used in this panel. The NCI- H292 cell line is a human airway epithelial cell line that produces mucins. Mucus overproduction is an important feature of bronchial asthma and chronic obstructive pulmonary disease samples. The expression of this gene in this mucoepidermoid cell line that is often used as a model for airway epithelium (NCI-H292 cells) suggests that this gene may be important in the proliferation or activation of airway epithelium. Therefore, therapeutics designed with the protein encoded by the transcript may reduce or eliminate symptoms caused by inflammation in lung epithelia in chronic obstructive pulmonary disease, asthma, allergy, and emphysema.

L. CG95861-01: TRANSFORMING GROWTH FACTOR-BETA INDUCED PROTEIN

Expression of gene CG95861-01 was assessed using the primer-probe set Ag2049, described in Table LA. Results of the RTQ-PCR runs are shown in Tables LB, LC, LD, LE, LF and LG.

Table LA. Probe Name Ag2049

Table LB. General_screening panel_vl.4

Table LC. Panel 1.3D

Table LG. general oncology screening panel_v_2.4

CG95861-01 gene is seen in melanoma (CT=1 .5). Overall, expression is much higher in cancer cell lines than in normal tissues. This pattern is also seen in Panels 1.3D and 2D. Please see Panel 2D for discussion of utility of this gene in cancer. Panel 1.3D Summary: Ag2049 Two experiments with the same probe and primer set produce results that are in excellent agreement. Overall, the expression of this gene in this panel is largely limited to samples derived from cancer cell lines. In particular there is a strong association (8 of 9) with expression of this gene prodcut, a transforming growth factor-beta induced protein IG-H3 precursor (BETA IG-H3) homolog, and cell lines derived from CNS cancer tissue, with highest expression of the CG95861-01 gene in cell lines derived from brain cancer (CTs=22-23.7). In addition, there is little expression in the normal brain samples when compared to the CNS derived cell lines. This putative BIGH3 is also expressed in a number of other cell lines derived from cancer tissues including melanoma, ovarian, breast, lung and renal cancers. Relatively low expression is observed in RNA samples corresponding to normal counterparts to these cancer cell lines. Please see Panel 2D for discussion of utility of this gene in cancer.

Panel 2D Summary: Ag2049 Highest expression of this putative BIG-H3 protein is seen in kidney cancer (CT=22). In addition, expression of this gene produt is seen in a number of cancer tissues, but not adjacent uninvolved sample. These include samples from gastric, bladder, kidney, lung and colon cancers. This suggests that the expression of BIGH3 might be associated with cancer.

Beta ig-h3 (BIGH3), was identified originally as a novel gene induced in a human adenocarcinoma cell line after treatment with transforming growth factor-beta (Skonier et al. DNA Cell Biol 1992 Sep;l 1(7):511-22). Most notable, with respect to potential roles in cancer biology, is a follow-up report by Skonier et al. in which it was reported that soluble BIGH3 was found to be capable of inhibiting the attachment of A549 lung carcinoma, HeLa cervical carcinoma, and WI-38 fibroblast cells in an in vitro adhesion assay. Moreover, Skonier et al reported that ectopic overexpression of BIGH3 was found to be capable of suppressing the growth of CHO cells in nude mice (DNA Cell Biol 1994

Jun;13(6):571-84). Finally, Skonier et al mapped BIGH3 to human chromosome 5q31, a region frequently deleted in preleukemic myeiodysplasia and leukemias, providing casual support to the notion that BIGH3 is a tumor suppressor.

Our data indicates that BIGH3 is unlikely to function as a tumor growth suppressor, but rather as a pro-active effector of tumor growth and perhaps resistance to cytotoxic therapy. BIGH3 (gbh_m77349) was identified in a GeneCalling analysis (Shimkets et al. Nat Biotechnol 1999 Aug;17(8):798-803) of differential gene expression comparison between MCF-7 cells (ER positive,, p53 positive, vimentin negative, noninvasive) and MCF-7/Adr (estrogen receptor negative, p53 negative, vimentin positive, invasive and hormone resistant) cells (Fairchild et al. Cancer Res 1987 Oct 1;47(19):5141- 8). MCF-7 Cells were derived from MCF-7 cells by virtue of their relative resistance to the front-line therapeutic agent for patients with breast cancer, adriamycin (doxorubicin), a DNA intercalating agent. In the GeneCalling analysis BIGH3 was found to be approximately 100-fold upregulated in MCF-7/ Adr cells relative to MCF-7 cells. Data obtained from real-time quantitative polymerase chain reaction (RTQ_PCR) analysis supports a pro-active role for BIGH3 in tumor etiology and/or progression. By RTQ-PCR we find that BIGH3 is expressed at high levels by most tumor cell lines relative to normal tissues or origins (see Panel 1.3D) and is frequently overexpressed in tumor tissues relative to histopathologically normal tissues obtained from the surgical margin adjacent to the tumor (see Panel 2D). This is most notable for clear cell carcinomas of the kidney, colon adenocarcinomas, non-small cell carcinomas of the lung, and carcinomas of the stomach. The striking and consistently high expression by cell lines derived form gliomas, astrocytomas and mixed glioma/astrocytomas strongly supports the role in the development and/or progression of malignancies of the CNS.

Six autosomal dominant corneal dystrophies are caused by mutations in the TGFBI (BIGH3) gene on chromosome 5q31 : three types of lattice corneal dystrophy (LCD), including type I and type IIIA, granular, Avellino (ACD), and Reis-Bucklers. The embryonic expression of the mouse ortholog in Bigh3 is observed in the mesenchyme of the first and second branchial arches as early as dpc 11.5 and is particularly strong in the mesenchyme of numerous tissues throughout all the development stages (Biochem Biophys Res Commun 2000 Aug 2;274(2):267-74). Mesenchymal cells are characterized by high growth rate, motility and invasion. The strong expression of this gene by most tumor cell lines in Panel 1.3D indicate that it is part of the epithelial to mesenchymal switch that tumor cells undergo during tumorogenic cellular transformation. Therefore, we postulate that this gene product has a role in tumor invasion, cell migration and growth, metastasis. Because of its activity and overexpression in many tumor types in panel 2D, therapeutic targeting of thsi gene product with a human monoclonal antibody is anticipated to limit or block the extent of tumor cell migration, invasion, growth and metastasis, preferably in gastric and colon, kidney, lung, bladder and ovarian tumors.

Panel 3D Summary: Ag2049 Highest expression of the CG95861-01 gene is seen in a glioma cell line (CT=23.6). Expression in this panel confirms expression of this gene product in cancer cell lines and suggests its central role in cell survival and growth. Please see Panel 2D for detailed discussion of utility of this gene in cancer.

Panel 4D Summary: Ag2049 The expression of the CG95861-01 gene, which encodes a putative BIGH3 molecule, is seen in many ceϋs involved in the immune system including endothelial cells, astrocytes, fibroblasts and monocytes, macrophages and dendritic cells. Highest expression is seen in resting coronary artery smooth muscle cells (CT=22.2). Expression in a cluster of samples derived from cells in the lung and skin suggest that this gene product may be involved in pathological and inflammatory conditions of the lung and skin, including psoriasis, asthma, emphysema and allergy. general oncology screening panel_v_2.4 Summary: Ag2049 Two experiments with the same probe and primer set produce results that are in excellent agreement.

Highest expression of the CG95861-01 gene is seen in kidney cancer (CTs=21-23). This expression is in agreement with expression in previous panels. Significant levels of expression are also seen in colon, lung and bladder cancers when compared to expression in the normal adjacent tissue. This gene is also expressed at a moderate level in the melanoma samples in this panel. Please see Panel 2D for additional discussion of utility of this gene in cancer, [sedinger, 18-Mar-02]

M. CG96412-01: diphthamide synthesis protein

Expression of gene CG96412-01 was assessed using the primer-probe sets Agl985 and Ag4055, described in Tables MA and MB. Results of the RTQ-PCR runs are shown in Tables MC, MD, ME, MF, MG, MH and MI.

Table MA. Probe Name Agl 985

Table MB. Probe Name Ag4055

Table MC. AI_comprehensive panel_vl .0

Table ME. General_screeningjpanel_vl.4

Table MF. Panel 1.3D

Table MH. Panel 4.1D

AI_comprehensive panel_vl.0 Summary: Agl985 Expression of the CG96412- 01 gene, which encodes a putative dipthamide synthesis protein, is highest in lung (CT=29.3). This gene is expressed widely in this panel, confirming its presence in tissues involved in the immune response. Please see Panel 4. ID for discussion of utility of this gene in inflammation.

CNS_neurodegeneration_vl.0 Summary: Agl985 This gene appears to be slightly upregulated in the temporal cortex of patients suffering from Alzheimer's disease. Thus, modulation of the expression or function of this gene may decrease neuronal death and be of use in the treatment of this disease. Results from a second experiment with the probe and primer set Ag4055 are not included. The amp plot indicates there were experimental difficulties with this run.

General_screening_panel_vl.4 Summary: Ag4055 The CG96412-01 gene is widely expressed in this panel, with highest expression in the cerebellum (CT=28.13). The high levels of expression in the cerebellum suggest that this gene product may be a useful and specific target of drugs for the treatment of CNS disorders that have this brain region as the site of pathology, such as autism and the ataxias. Moderate levels of expression are seen across the CNS, including the amygdala, hippocampus, cerebral cortex, substantia nigra and thalamus. Therefore, therapeutic modulation of the expression or function of this gene may be useful in the freatment of neurologic disorders, such as Alzheimer's disease, Parkinson's disease, schizophrenia, multiple sclerosis, stroke and epilepsy.

Among tissues with metabolic function, this gene is expressed at moderate to low levels in pituitary, adipose, adrenal gland, pancreas, thyroid, and adult and fetal skeletal muscle, heart, and liver. This widespread expression among these tissues suggests that this gene product may play a role in normal neuroendocrine and metabolic function and that disregulated expression of this gene may contribute to neuroendocrine disorders or metabolic diseases, such as obesity and diabetes.

Panel 1.3D Summary: Agl985 Expression of the CG96412-01 gene is highest in the pituitary (CT=31), with low but significant levels also evident in the amygdala, cerebellum, cerebral cortex and spinal cord. Please see the previous panels for discussion of utility of this gene in the CNS.

Among other tissues with metabolic function, this gene is expressed at low levels in adipose, adrenal gland, and fetal skeletal muscle and heart. This expression suggests that this gene product may play a role in normal neuroendocrine and metabolic function and that disregulated expression of this gene may contribute to neuroendocrine disorders or metabolic diseases, such as obesity and diabetes. In addition, expression of this gene product could be used to differentiate between fetal skeletal muscle (CT=32.4) and adult skeletal muscle (CT=40).

Panel 2D Summary: Agl985 The CG96412-01 gene is most highly expressed in uterine cancer (CT=31.1). In addition, higher levels of expression are seen in normal bladder, ovary, lung, kidney, and prostate when compared to expression in the coπesponding adjacent tumor. Thus, expression of this gene could be used as a marker of these tissues. Furthermore, therapeutic modulation of the expression or function of this gene may be effective in the freatment of bladder, ovary, lung, kidney, uterine and prostate cancers. Panel 4.1D Summary: Agl985/Ag4055 Two experiments with two different probe and primer sets produce results that are in excellent agreement. The CG96412-01 gene is widely expressed in this panel, with highest expression in LAK cells freated with PMA and ionomycin (CTs=27-27.6). Lower levels of induction are seen in macrophages and monocytes after treatment with LPS and in EOL cells (eosinophil cell line) after freatment with PMA/ionomycin. Thus, this transcript encodes a protein that appears to be expressed in response to activation and may be involved in the immune function or proliferation of LAK cells, macrophages and monocytes. It is also detected in tumor margins (Panel 2D) that often contain activated LAK, monocytes, and macrophages. Therefore, antibody or small molecule antagonist therapies directed against the protein encoded by this transcript could reduce or inhibit inflammation in diseases such as asthma, emphysema, allergy, psoriasis, diabetes and arthritis. These treatments may also reduce or prevent tissue rejection after organ transplant. Agonistic therapies in contrast, my up regulate the function of LAK cells and aid in cancer treatments.

Panel 4D Summary: Agl985 Expression of the CG96412-01 gene is in reasonable agreement with the results in Panel 4. ID. Highest expression of the gene is seen in LAK cells treated with PMA/ionomycin (CT=28.6). Please see Panel 4. ID for discussion of utility of this gene in inflammation.

N. CG96511-01: Hs ECHE

Expression of gene CG96511-01 was assessed using the primer-probe sets Ag4063, Ag4171 and Ag4172, described in Tables NA, NB and NC. Results of the RTQ- PCR runs are shown in Tables ND and NE. Table NA. Probe Name Ag4063

Table NB. Probe Name Ag4171

Table NC. Probe Name Ag4172

Table ND. General_screening_panel_vl.4

Table NE. Panel 4.1D

CNS_neurodegeneration_vl.O Summary: Ag4063/Ag4171/Ag4172 Results from these three experiment with the CG96511-01 gene are not included. The amp plot indicates that there were experimental difficulties with this run.

General_screening_panel_vl.4 Summary: Ag4063 Highest expression of the CG96511-01 gene is detected in gastric cancer Kato III sample (CT=30.8). In addition, significant expression of this gene is also detected in gastric (liver metastasis) cancer, lung cancer, renal cancer and breast cancer cell lines. Thus, expression of this gene can be used to distinguish between these samples and other samples in this panel. In addition, therapeutic modulation of the small chemokine encoded by this gene, through the use of small molecule drugs, protein therapeutics or antibodies, might be beneficial in the freatment of these cancers.

Ag4171/Ag4172 Results from these two experiment with the CG96511-01 gene are not included. The amp plot indicates that there were experimental difficulties with this run.

Panel 4.1D Summary: Ag4063 The CG96511-01 gene is only expressed at detectable levels in the kidney (CT=33.7). Therefore, espression of this gene could be used to distinguish the kidney sample from other samples used in this panel. In addition, antibody or small molecule therapies designed with the protein encoded for by this gene could modulate kidney function and be important in the treatment of inflammatory or autoimmune diseases that affect the kidney, including lupus and glomerulonephritis.

Ag4171/Ag4172 Expression of the CG96511-01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown).

O. CG96522-01: ADAM-TS 7 PRECURSOR

Expression of gene CG96522-01 was assessed using the primer-probe sets Ag4084, Ag4322 and Ag4084, described in Tables OA, OB and OC. Table OA. Probe Name Ag4084

Table OB. Probe Name Ag4322

CNS_neurodegeneration_vl.O Summary: Ag4322 Expression of the CG96522- 01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown).

Ag4084 Results from one experiment with the CG96522-01 gene are not included. The amp plot indicates that there were experimental difficulties with this run.

Generaϊ_screeningjpanel_vl.4 Summary: Ag4084/Ag4322 Expression of the CG96522-01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown).

Panel 4.1D Summary: Ag4084/Ag4322 Expression of the CG96522-01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown).

P. CG96637-01: SMALL INDUCIBLE CYTOKINE A23 PRECURSOR Expression of gene CG96637-01 was assessed using the primer-probe sets Ag4081 and Ag4345, described in Tables PA and PB. Results of the RTQ-PCR runs are shown in Tables PC and PD.

Table PA. Probe Name Ag4081

Table PB. Probe Name Ag4345

Table PD. Panel 4. ID

CNS_neurodegeneration_vl.O Summary: Ag4345 This panel confirms the expression of the CG96637-01 gene at low levels in the brains of an independent group of individuals. There is no differential expression of this gene between Alzheimer's diseased postmortem brains and those of non-demented controls in this experiment. However, this gene may play a role in other central nervous system disorders such as Parkinson's disease, epilepsy, multiple sclerosis, schizophrenia and depression.

Ag4081 Results from one experiment with the CG96637-01 gene are not included. The amp plot indicates that there were experimental difficulties with this run.

General_screening_panel_vl.4 Summary: Ag4081 Expression of the CG96637- 01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown).

Panel 4.1D Summary: Ag4345 Two experiments with same probe and primer sets are in excellent agreement with highest expression of the CG96637-01 gene in dentritic cells and LPS freated monocytes (CTs=29.4-30.6). In addition, expression of this gene is stimulated in PHA-L freated PBMC cells, IFN gamma treated HUVEC, and LPS freated monocytes. Moderate expression of this gene is also detected in resting neutrophils, colon, lung, HPAEC, and dendritic cells. Therefore, therapeutic modulation of this gene product may be useful in the freatment of lupus erythematosus, asthma, emphysema, Crohn's disease, ulcerative colitis, rheumatoid arthritis, osteoarthritis, and psoriasis.

Ag4081 Expression of the CG96637-01 gene is low/undetectable (CTs > 35) across all of the samples on this panel (data not shown).

Q. CG97274-01: GRANULOCYTE COLONY-STIMULATING FACTOR PRECURSOR

Expression of full length clone CG97274-01 was assessed using the primer-probe sets Ag4104 and Ag4120, described in Tables QA and QB. Results of the RTQ-PCR runs are shown in Tables QC, QD, QE, QF and QG.

Table OA. Probe Name Ag4104

Table OB. Probe Name Ag4120

Table QC. AI_comprehensive panel_vl.O

Table OD. General_screening_panel_vl.4

Table OE. Panel 2.2

Table OG. Panel 4. ID

AI_comprehensive pane vl.O Summary: Ag4120 Two experiments with the same probe and primer show results that are in excellent agreement. Highest expression of the CG97274-01 is seen in samples derived from synovium of RA patients (CTs=33.4- 33.5). In contrast, transcripts of variant CG97274-04 are produced at lower levels and in fewer tissue samples from RA patients. This shows disease tissue specific expression of this gene variant and distinguishes the expression pattern of CG97274-01 and CG97274- 04. Results from a third experiment swith the same probe and primer show low/undetectable levels of expression (CTs>35) and are not included.

CNSjαeurodegeneration_vl.0 Summary: Ag4120 Expression of the CG97274- 01 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.) A third experiment with the probe and primer set Ag4104 is not included. The amp plot indicates that there were experimental difficulties with this run.

General_screeningjpanel_vl.4 Summary: Ag4120 Two experiments with the same probe and primer produce results that are in excellent agreement. Highest expression of the CG97274-01 gene is seen in a brain cancer cell line (CT=29-31.6) with significant expression also seen in a melanoma cell line. Thus, expression of this gene could be used to differentiate between these samples and other samples on this panel and as a marker to detect the presence of brain and melanoma cancers. Furthermore, therapeutic modulation of the expression or function of this gene may be effective in the treatment of these cancers.

This gene is also expressed at moderate to low levels in adipose and fetal lung. In comparison, CG97274-04 is expressed at significant levels only in the cancer cell lines.

Panel 2.2 Summary: Ag4120 Expression of the CG97274-01 gene is restricted to samples derived from normal lung tissue (CTs=34). Thus, expression of this gene could be used to differentiate between these samples and other samples on this panel and as a marker of lung tissue. Furthermore, therapeutic modulation of the expression or function of this gene may be effective in the treatment of lung cancer.

Panel 3D Summary: Ag4120 Expression of the CG97274-01 gene is restricted to a sample derived from a bladder cancer cell line (CT=33.2). Thus, expression of this gene could be used to differentiate between this sample and other samples on this panel and as a marker to detect the presence of bladder cancer. Furthermore, therapeutic modulation of the expression or function of this gene may be effective in the freatment of bladder cancer. Panel 4.1D Summary: Ag4120 Three experiments with the same probe and primer set produce results that are in very good agreement. Highest expression of the CG97274-01 gene is seen in TNF-alpha and IL-1 beta treated coronary artery SMCs and HPAECs (CTs=29.4-31.7) and IL-1 beta treated dermal fibroblasts (CT=33). The transcript is also induced in microvascular dermal endothelial cells, and lung microvascular endothelial cells and LPS treated monocytes. Antibody therapeutics designed against the protein encoded by the CG97274-01 gene may reduce or inhibit inflammation or the tissue damage that results from inflammatory processes by blocking the function of this protein, a putative member of the granulocytes colony stimulating factor family. Members of this family have been shown to promote survival, proliferation and terminal differentiation of granulocyte precursors (Metcalf, D. Science 229: 16-22, 1985. Souza LM, Science 1986 Apr 4;232(4746):61-5). Treatment with G-CSF protein also induces allergic reactions (Sullivan AK, Int J STD AIDS 1997 Feb;8(2): 135-6. Glass LF, J Am Acad Dermatol 1996 Mar;34(3):455-9). Thus, the CG97274-01 gene product may have similar functions as G-CSF in hematopoiesis and distinct functions in autoimmune disorders such as rheumatoid arthritis. Based on the expression profile of the CG97274-01 gene and on the known function of G-CSF, blocking the biological activity of this protein with antibody therapeutics may reduce granulocyte participation in diseases, particularly rheumatoid arthritis (based on expression in the A/I panel) and in asthma, emphysema, allergy and other inflammatory conditions (based on panel 4.1). In addition, protein therapeutics could also be designed with the protein encoded for by CG97274-01. The protein could be inhibitory and block G-CSF interactions with the G- CSF receptor^nd serve as an antagonistic protein therapeutic to block the specific functions of G-CSF. The protein encoded by the CG97274-01 gene could also serve as an agonistic protein therapeutic if it utilizes the GCSF receptor and functions similarly to clinically utilized forms of G-CSF. Thus, it may play an important role in restoring myelopoiesis after immunomodulatory treatments such as chemotherapy for cancer and bone maπow transplant or in stimulating myelopoiesis in neufropenic disorders (Weaver C, Bone Marrow Transplant 2001 May;27 Suppl 2:S23-9 Dale DC, Stem Cells 1995 Mar;13(2):94-100). Alternatively, based on expression in Panel 1.4 this gene product may have a unique receptor and have agonistic protein therapeutic functions distinct from CG97274-04, such as promoting lung development. Panel 5D Summary: Ag4120 Expression of the CG97274-01 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.)

Panel CNS_1 Summary: Ag4120 Expression of the CG97274-01 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.)

R. CG97274-04: Granulocyte colony-stimulating factor precursor (G-

CSF3)

Expression of gene CG97274-04 was assessed using the primer-probe set Ag4124, described in Table RA. Results of the RTQ-PCR runs are shown in Tables RB, RC and RD.

Table RA. Probe Name Ag4124

Table RD. Panel 4.1D

AI_comprehensive paneLvl.O Summary: Ag4124 Expression of the CG97274- 04 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.) Please note that this lack of detectable expression is in confrast with the expression profile of the CG97274-01 variant. CNS_neurodegeneration_vl.0 Summary: Ag4124 Expression of the CG97274- 04 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.)

General_screeningjpanel_vl.4 Summary: Ag4124 Expression of the CG97274- 04 gene is restricted to samples derived from a brain cancer and a melanoma cell line (CTs=30-32). Thus, expression of this gene could be used to differentiate between these samples and other samples on this panel and as a marker to detect the presence of brain and melanoma cancers. Furthermore, therapeutic modulation of the expression or function of this gene may be effective in the freatment of these cancers.

Panel 3D Summary: Ag4124 Expression of the CG97274-04 gene is restricted to a sample derived from a bladder cancer cell line (CT=33). Thus, expression of this gene could be used to differentiate between this sample and other samples on this panel and as a marker to detect the presence of bladder cancer. Furthermore, therapeutic modulation of the expression or function of this gene may be effective in the treatment of bladder cancer.

Panel 4.1D Summary: Ag4124 Expression of the CG97274-04 gene is highest in TNF-alpha and IL-1 beta HPAEC cells (CT=29.4). In addition, expression is seen in other TNF-alpha and/or IL-1 beta treated samples including dermal fibroblasts, lung fibroblasts, coronary artery SMCs, lung microvasculature, small airway epithelium and keratinocytes. Since expression of this transcript appears to be up regulated by the cytokine TNF-a, therapeutic modulation of the function or expression of the protein encoded by this gene may reduce or eliminate inflammation and tissue damage that result from diseases associated with hyperactivated T cells including lupus erythematosus, rheumatoid arthritis, and inflammatory bowel diseases.

Panel CNS_1 Summary: Ag4124 Expression of the CG97274-04 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.) general oncology screening panel_v_2.4 Summary: Ag4124 Expression of the

CG97274-04 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.)

S. CG97288-01 and CG97288-02: HsPLP2Long

Expression of gene CG97288-01 and variant CG97288-02 was assessed using the primer-probe set Ag4173, described in Table SA. Results of the RTQ-PCR runs are shown in Tables SB, SC and SD.

Table SA. Probe Name Ag4173

Table SC. General_screening_panel_vl.4

CNS_neurodegeneration_vl.O Summary: Ag4173 This panel confirms the expression of the CG97288-01 gene at low levels in the brains of an independent group of individuals. However, no differential expression of this gene was detected between Alzheimer's diseased postmortem brains and those of non-demented controls in this experiment. Please see Panel 1.4 for a discussion of the potential utility of this gene in treatment of cenfral nervous system disorders.

Generaϊ_screening_panel_vl.4 Summary: Ag4173 Highest expression of the CG97288-01 is detected in fetal liver (CT=26). Interestingly, this gene is expressed at much higher levels in fetal when compared to adult liver(CT=34). This observation suggests that expression of this gene can be used to distinguish fetal from adult liver. In addition, the relative overexpression of this gene in fetal liver suggests that the protein product may enhance liver growth or development in the fetus and thus may also act in a regenerative capacity in the adult. Therefore, therapeutic modulation of the protein encoded by this gene could be useful in treatment of liver related diseases.

Among tissues with metabolic or endocrine function, this gene is expressed at low to moderate levels in pancreas, adipose, adrenal gland, thyroid, pituitary gland, skeletal muscle, heart, liver and the gastrointestinal tract. Therefore, therapeutic modulation of the activity of this gene may prove useful in the treatment of endocrine/metabolically related diseases, such as obesity and diabetes.

Low to moderate expression of this gene is also detected in colon cancer tissue, a breast cancer and an ovarian cancer cell lines. Therefore, therapeutic modulation of this gene product could be useful in the freatment of these cancers.

In addition, this gene is expressed at moderate levels in cerebellum, fetal and adult whole brain. Therefore, this gene may play a role in central nervous system disorders such as Alzheimer's disease, Parkinson's disease, epilepsy, multiple sclerosis, schizophrenia and depression.

Interestingly, this gene is expressed at much higher levels in fetal (CTs=27-30) when compared to adult heart, lung and kidney(CTs=34). This observation suggests that expression of this gene can be used to distinguish these fetal from the corresponding adult tissue. In addition, the relative overexpression of this gene in fetal tissue suggests that the protein product may play a role in lung, heart and kidney development in the fetus and thus may also act in a regenerative capacity in the adult. Therefore, therapeutic modulation of the protein encoded by this gene could be useful in treatment of lung, heart and kidney related diseases.

Panel 4.1D Summary: Ag4173 Highest expression of the CG97288-01 is detected in LPS treated monocytes (CT=28.5). Significant expression of this gene is thymus, PBMC cells, neutrophils, macrophages, LAK cells, CD4 and secondary CD8 lymphocytes. In addition, expression of this gene is stimulated in LPS treated monocytes, PMA/ionomycing freated EOL-1 dbcAMP, TNF alpha + IL-1 beta freated HPAEC cells. Therefore, therapeutic modulation of this gene product could be beneficial in the treatment of autoimmune and inflammatory diseases such as asthma, allergies, inflammatory bowel disease, lupus erythematosus, psoriasis, rheumatoid arthritis, and osteoarthritis.

In addition, moderate expression is also detected in kidney (CT=30). Therefore, antibody or small molecule therapies designed with the protein encoded for by this gene could modulate kidney function and be important in the treatment of inflammatory or autoimmune diseases that affect the kidney, including lupus and glomerulonephritis.

T. CG97550-01: Serine Protease

Expression of gene CG97550-01 was assessed using the primer-probe sets Agl 156, Agl411 and Ag384, described in Tables TA, TB and TC. Results of the RTQ- PCR runs are shown in Tables TD, TE and TF.

Table TA. Probe Name Agl 156

Table TB. Probe Name Agl411

Table TC. Probe Name Ag384

Table TD. CNS_neurodegeneration_vl.O

CNS_neurodegeneration_vl.0 Summary: Ag384 his panel confirms the expression of the CG97550-01 gene at low levels in the brains of an independent group of individuals. However, no differential expression of this gene was detected between Alzheimer's diseased postmortem brains and those of non-demented controls in this experiment. Low levels of expression in the brain suggests that this gene may play a role in cenfral nervous system disorders such as Parkinson's disease, epilepsy, multiple sclerosis, schizophrenia and depression.

Panel 1.1 Summary: Ag384 Two experiments with excellent agreement with highest expression of the CG97550-01 gene in heart (CTs=22). Expression of this gene is seen exclusively in almost all the normal tissue samples used in this panel. This expression in normal tissues suggests that this gene product may play an important role in cellular function. Interestingly, high expression of this gene is also detected in a lung cancer (non- small cell) HOP-62 cell line and prostate cancer bone metastasis cell line PC3. Thus, expression of this gene can be used as a diagnostic marker for lung and prostate cancer. Also, therapeutic modulation of the activity of this gene or its protein product, through the use of small molecule drugs, protein therapeutics or antibodies, might be beneficial in the freatment of lung cancer or prostate cancer.

Among tissues with metabolic or endocrine function, this gene is expressed at high to moderate levels in pancreas, adrenal gland, thyroid, pituitary gland, skeletal muscle, heart, liver and the gastrointestinal tract. Therefore, therapeutic modulation of the activity of this gene may prove useful in the freatment of endocrine/metabolically related diseases, such as obesity and diabetes.

Panel 4.1D Summary: Ag384 Highest expression of the CG97550-01 gene is detected in lung (CTs=30.15). Moderate expression of this gene is also seen in dermal fibroblasts, HPAEC cells, coronery artery SMC cells, HUVEC cells, lung and dermal microvascular EC cells and liver ciπhosis sample. In addition, expression of this gene is stimulated in LPS freated monocytes, TNFalpha + IL-lbeta treated asfrocytes, and PMA/ionomycin freated basophils. The CG97550-01 gene codes for a serine protease. Proteins belonging to serine protease family have been implicated in numerous inflammatory and malignant diseases wherein inflammatory cells such as monocytes/macrophages are involved. Such diseases include lung cancer, chronic inflammatory bowel disease, vasculitis and connective tissue disease, bacterial sepsis, and septic shock (Heiden et al., 1996, Semin Thromb Hemost 22(6) :497-501, PMID: 9122714). Therefore, therapeutic modulation of the serine protease encoded by this gene could be beneficial in the freatement of asthma, allergies, inflammatory bowel disease, lupus erythematosus, psoriasis, rheumatoid arthritis, and osteoarthritis.

Significant expression of this gene is also seen in normal colon, lung, thymus and kidney tissue. This pattern is in agreement with the expression profile in General_screening_panel_vl .4 and also suggests a role for the gene product in cell survival and proliferation.

Therefore, therapeutic modulation of this gene product may be useful in treatment of inflammatory disease of these tissues.

U. CG97800-01 and CG97800-02 and CG97800-03: Elastase IV-like

Expression of gene CG97800-01 and full length clones CG97800-02 and CG97800-03 was assessed using the primer-probe set Ag4156, described in Table UA. Results of the RTQ-PCR runs are shown in Tables UB and UC. Please note that CG97800-03 represents a full-length physical clone of the CG97800-01 gene, validating the prediction of the gene sequence.

Table UA. Probe Name Ag4156

Table UB. General_screening panel_vl.4

Table UC. Panel 4.1D

CNS_neurodegeneration_vl.0 Summary: Ag4156 Expression of the CG97800- 01 gene is low/undetectable in all samples on this panel (CTs>35). (Data not shown.)

General_screening__panel_vl.4 Summary: Ag4156 Expression of the CG97800- 01 gene, an elastase homolog, is limited to the pancreas and bladder (CTs=24.3-25.1). Elastases are proteinases that dissolve elastin. They have been implicated in the pathology of infections and inflammation. Based on the pattern of expression of this putative elastin, expression of this gene could be used as a marker of bladder and pancreatic tissue and to distinguish these samples from other samples on this panel. In addition, therapeutic modulation of the expression or function of this gene product may be useful in the freatment of infection, inflammation and cancer of these tissues.

Panel 4.1D Summary: Ag4156 Expression of the CG97800-01 gene is limited to a few samples with highest expression in LAK cells freated with PMA/ionomycin (CT=33.5). Low, but significant expression is also seen in TNF-alpha and LPS treated neutrophils, untreated dendritic cells, LPS treated monocytes and IL-2 treated resting NK cells. LAK cells are involved in tumor immunology and cell clearance of virally and bacterial infected cells as well as tumors. Therefore, modulation of the function of the protein encoded by this gene through the application of a small molecule drug or antibody may alter the functions of these cells and lead to improvement of symptoms associated with these conditions.

V. CG98092-01: Collagen like protein

Expression of gene CG98092-01 was assessed using the primer-probe sets Agl 964 and Ag4143, described in Tables VA and VB. Results of the RTQ-PCR runs are shown in Tables VC, VD, VE, VF, VG, VH and VI.

Table VA. Probe Name Agl 964

Table VB. Probe Name Ag4143

Table VC. CNS_neurodegeneration_ l.O

Table VD. General_screening_panel_vl.4

Table VE. Panel 1.3D

Table VF. Panel 2D

Table VG. Panel 4. ID

Table VH. Panel 4D

Table VI. general oncology screening panel_y_2.4

CNS_neurodegeneration_vl.O Summary: Agl964/Ag4143 Two experiments with two different probe and primer sets produce results that are in very good agreement. Highest expression of the CG98092-01 gene is seen in the temporal cortex of an Alzheimer's patient in one experiment (CT=28.2) and in a confrol brain in the second (CT=30). Overall, this gene appears to be slightly down-regulated in the temporal cortex of Alzheimer's disease patients. Therefore, up-regulation of this gene or its protein product, or freatment with specific agonists may be of use in reversing the dementia,memory loss and neuronal death associated with this disease.

General_screening_panel_vl.4 Summary: Ag4143 Highest expression of the CG98092-01 gene is seen in a lung cancer cell line (CT=26.4). This gene is widely expressed in the samples on this panel, with prominent expression in all the cancer cell lines. Thus, expression of this gene could be used to differentiate between the lung cancer cell line and other samples on this panel and as a marker for lung cancer. Since cell lines are generally more proliferative than tissues, this gene might be involved in cell proliferation. Therefore, modulation of the expression or function of this gene may useful for the treatment of cancer or other disease that involve cell proliferation. Among tissues with metabolic function, this gene is expressed at moderate to low levels in pituitary, adipose, adrenal gland, pancreas, thyroid, fetal skeletal muscle and , heart, and adult and fetal liver. This widespread expression among these tissues suggests that this gene product may play a role in normal neuroendocrine and metabolic function and that disregulated expression of this gene may contribute to neuroendocrine disorders or metabolic diseases, such as obesity and diabetes.

In addition, this gene is expressed at much higher levels in fetal lung (CT=32) when compared to expression in the adult counterpart (CT=35). Thus, expression of this gene may be used to differentiate between the fetal and adult source of this tissue. This gene is also expressed at moderate levels in the CNS, including the hippocampus, thalamus, substantia nigra, amygdala, cerebellum and cerebral cortex.

Therefore, therapeutic modulation of the expression or function of this gene may be useful in the freatment of neurologic disorders, such as Alzheimer's disease, Parkinson's disease, schizophrenia, multiple sclerosis, stroke and epilepsy.

Panel 1.3D Summary: Agl964 Highest expression of the CG98092-01 gene is seen in the hippocampus (CT=30.2). Moderate to low levels of expression are also seen in the amygdala, thalamus, and cortex. Please see Panel 1.4 for discussion of utility in the CNS. Among metabolic tissues, low but significant levels of expression are seen in fetal skeletal muscle and the adrenal gland. Thus, this gene product may be involved in the pathogenesis and/or diagnosis of metabolic diseases that affect these tissues.

Moderate levels of expression are also seen in clusters of samples derived from lung cancer, brain cancer, and colon cancer cell lines. Please see Panel 1.4 for further discussion of utility of this gene in cancer.

Panel 2D Summary: Agl964 Highest expression of the CG98092-01 gene is seen in a lung cancer (CT=28). Significant levels of expression are also seen in lung, bladder, and colon cancers when compared to coπesponding normal adjacent tissue. Thus, expression of this gene could be used as a marker to detect the presence of these cancers. Furthermore, therapeutic modulation of the expression or function of this gene may be effective in the treatment of bladder, lung, and colon cancer.

Panel 4.1D Summary: Ag4143 Highest expression of the CG98092-01 gene is seen in LAK cells freated with PMA/ionomycin (CT=28.1). In addition, moderate levels of expression are seen many hematopoietic cell types, including activated primary and secondary T cells, CD8 and CD4 lymphocytes, PWM treated PBMC and B lymphocytes, and activated dendritic cells, macrophages and monocytes. Thus, therapeutic modulation of the expression or function of this gene may reduce or eliminate symptoms in patients with autoimmune and inflammatory diseases such as, but not limited to, including Crohn's disease, ulcerative colitis, multiple sclerosis, chronic obstructive pulmonary disease, asthma, emphysema, rheumatoid arthritis, lupus erythematosus, or psoriasis.

Panel 4D Summary: Agl964 Highest expression of the CG98092-01 gene is seen in LAK cells freated with PMA/ionomycin (CT=27.3). In addition, moderate levels of expression are seen many hematopoietic cell types, including activated primary and secondary T cells, CD8 and CD4 lymphocytes, PWM treated PBMC and B lymphocytes, and activated dendritic cells, macrophages and monocytes. Thus, therapeutic modulation of the expression or function of this gene may reduce or eliminate symptoms in patients with autoimmune and inflammatory diseases such as, but not limited to, including Crohn's disease, ulcerative colitis, multiple sclerosis, chronic obstructive pulmonary disease, asthma, emphysema, rheumatoid arthritis, lupus erythematosus, or psoriasis.

General oncology screening panel_v_2.4 Summary: Ag4143 Expression of the CG98092-01 gene appears to be limited to prostate derived tissue, with highest expression in a prostate cancer sample (CT=28). Thus, expression of this gene could be used to differentiate between prostate derived tissue and other samples on this panel and as a marker of prostate tissue.

W. CG98121-01: MMTV-R-like cytokine

Expression of gene CG98121-01 was assessed using the primer-probe set Agl 984, described in Table WA. Results of the RTQ-PCR runs are shown in Tables WB, WC, WD and WE.

Table WA. Probe Name Agl 984

Table WB. CNS_neurodegeneration_vl.O

Table WC. Panel 1.3D

Table WE. Panel 4D

CNS_neurodegeneration_vl.O Summary: Agl 984 This panel confirms the expression of the CG98121-01 gene at moderate levels in the brain in an independent group of individuals. This gene is found to be upregulated in the temporal cortex of Alzheimer's disease patients. Therefore, therapeutic inhibition of this protein may be of use in the freatment of this disease and decrease neuronal death.

Panel 1.3D Summary: Agl984 Highest expression of the CG98121-01 gene is seen in a neuroblastoma cell line(CT=29.7). Thus, expression of this gene could be used to differentiate between this sample and other samples and as a marker for this cancer.

Furthermore, therapeutic modulation of the expression or function of this gene may be effective in the treatment of brain cancer.

Among tissues with metabolic function, this gene is expressed at moderate to low levels in pituitary, adipose, adrenal gland, fetal liver, and adult and fetal skeletal muscle and heart. This expression suggests that this gene product may play a role in normal neuroendocrine and metabolic function and that disregulated expression of this gene may contribute to neuroendocrine disorders or metabolic diseases, such as obesity and diabetes. This gene is also expressed at low levels in the CNS, including the hippocampus, thalamus, substantia nigra, amygdala, and cerebral cortex. Therefore, therapeutic modulation of the expression or function of this gene may be useful in the freatment of neurologic disorders, such as Alzheimer's disease, Parkinson's disease, schizophrenia, multiple sclerosis, stroke and epilepsy.

Panel 2D Summary: Agl984 Highest expression of the CG98121-01 gene is seen in normal lung (CT=31.1). Significant expression is also seen in colon cancer when compared to normal adjacent tissue and conversely in kidney when compared to adjacent tumor. Thus, therapeutic modulation of the expression or function of this protein may be useful in the treatment of kidney and colon cancer.

Panel 4D Summary: Agl984 The CG98121-01 transcript is induced in the PMA and ionomycin freated basophil cell line KU-812 (CT=27.5). Basophils release histamines and other biological modifiers in reponse to allergens and play an important role in the pathology of asthma and hypersensitivity reactions. Therefore, therapeutics designed against the putative protein encoded by this gene may reduce or inhibit inflammation by blocking basophil function in these diseases. In addition, these cells are a reasonable model for the inflammatory cells that take part in various inflammatory lung and bowel diseases, such as asthma, Crohn's disease, and ulcers.

Example D: Identification of Single Nucleotide Polymorphisms in NOVX nucleic acid sequences Variant sequences are also included in this application. A variant sequence can include a single nucleotide polymorphism (SNP). A SNP can, in some instances, be referred to as a "cSNP" to denote that the nucleotide sequence containing the SNP originates as a cDNA. A SNP can arise in several ways. For example, a SNP may be due to a substitution of one nucleotide for another at the polymoφhic site. Such a substitution can be either a transition or a transversion. A SNP can also arise from a deletion of a nucleotide or an insertion of a nucleotide, relative to a reference allele. In this case, the polymorphic site is a site at which one allele bears a gap with respect to a particular nucleotide in another allele. SNPs occurring within genes may result in an alteration of the amino acid encoded by the gene at the position of the SNP. Infragenic SNPs may also be silent, when a codon including a SNP encodes the same amino acid as a result of the redundancy of the genetic code. SNPs occurring outside the region of a gene, or in an intron within a gene, do not result in changes in any amino acid sequence of a protein but may result in altered regulation of the expression pattern. Examples include alteration in temporal expression, physiological response regulation, cell type expression regulation, intensity of expression, and stability of transcribed message.

SeqCalling assemblies produced by the exon linking process were selected and extended using the following criteria. Genomic clones having regions with 98% identity to all or part of the initial or extended sequence were identified by BLASTN searches using the relevant sequence to query human genomic databases. The genomic clones that resulted were selected for further analysis because this identity indicates that these clones contain the genomic locus for these SeqCalling assemblies. These sequences were analyzed for putative coding regions as well as for similarity to the known DNA and protein sequences. Programs used for these analyses include Grail, Genscan, BLAST, HMMER, FAST A, Hybrid and other relevant programs.

Some additional genomic regions may have also been identified because selected SeqCalling assemblies map to those regions. Such SeqCalling sequences may have overlapped with regions defined by homology or exon prediction. They may also be included because the location of the fragment was in the vicinity of genomic regions identified by similarity or exon prediction that had been included in the original predicted sequence. The sequence so identified was manually assembled and then may have been extended using one or more additional sequences taken from CuraGen Corporation's human SeqCalling database. SeqCalling fragments suitable for inclusion were identified by the CuraTools™ program SeqExtend or by identifying SeqCalling fragments mapping to the appropriate regions of the genomic clones analyzed.

The regions defined by the procedures described above were then manually integrated and corrected for apparent inconsistencies that may have arisen, for example, from miscalled bases in the original fragments or from discrepancies between predicted exon junctions, EST locations and regions of sequence similarity, to derive the final sequence disclosed herein. When necessary, the process to identify and analyze SeqCalling assemblies and genomic clones was reiterated to derive the full length sequence (Alderborn et al., Determination of Single Nucleotide Polymorphisms by Real-time Pyrophosphate DNA Sequencing. Genome Research. 10 (8) 1249-1265, 2000).

Variants are reported individually but any combination of all or a select subset of variants are also included as contemplated NOVX embodiments of the invention.

Table 1. Variants of NOV2a (CGI 05716-01).

Table 2. Variants of NOV5a (CG57415-01).

Table 3. Variants of NOV7b (CG58586-02).

Table 7. Variants of NOV14a (CG95861-01).

Table 8. Variants of NOV15a (CG96412-01).

Table 9. Variant of NOVl 9a (CG96567-02).

Table 10. Variants of NOV21a (CG97274-01).

Table 11. Variant of NOV24a (CG97550-01).

Table 12. Variants of NOV26a (CG97800-01).

Table 13: Variants of NOV29a (CG99662-01).

OTHER EMBODIMENTS

Although particular embodiments have been disclosed herein in detail, this has been done by way of example for purposes of illustration only, and is not intended to be limiting with respect to the scope of the appended claims, which follow. In particular, it is contemplated by the inventors that various substitutions, alterations, and modifications may be made to the invention without departing from the spirit and scope of the invention as defined by the claims. The choice of nucleic acid starting material, clone of interest, or library type is believed to be a matter of routine for a person of ordinary skill in the art with knowledge of the embodiments described herein. Other aspects, advantages, and modifications considered to be within the scope of the following claims. The claims presented are representative of the inventions disclosed herein. Other, unclaimed inventions are also contemplated. Applicants reserve the right to pursue such inventions in later claims.

Claims

We claim:

What is claimed is: 1. An isolated polypeptide comprising the mature form of an amino acid sequenced selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62

2. An isolated polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62.

3. An isolated polypeptide comprising an amino acid sequence which is at least 95% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62.

4. An isolated polypeptide, wherein the polypeptide comprises an amino acid sequence comprising one or more conservative substitutions in the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62.

5. The polypeptide of claim 1 wherein said polypeptide is naturally occurring.

6. A composition comprising the polypeptide of claim 1 and a carrier.

7. A kit comprising, in one or more containers, the composition of claim 6.

8. The use of a therapeutic in the manufacture of a medicament for treating a syndrome associated with a human disease, the disease selected from a pathology associated with the polypeptide of claim 1, wherein the therapeutic comprises the polypeptide of claim 1.

9. A method for determining the presence or amount of the polypeptide of claim 1 in a sample, the method comprising:

(a) providing said sample; (b) introducing said sample to an antibody that binds immunospecifically to the polypeptide; and

(c) determining the presence or amount of antibody bound to said polypeptide, thereby determining the presence or amount of polypeptide in said sample.

10. A method for determining the presence of or predisposition to a disease associated with altered levels of expression of the polypeptide of claim 1 in a first mammalian subject, the method comprising: a) measuring the level of expression of the polypeptide in a sample from the first mammalian subject; and b) comparing the expression of said polypeptide in the sample of step (a) to the expression of the polypeptide present in a control sample from a second mammalian subject known not to have, or not to be predisposed to, said disease, wherein an alteration in the level of expression of the polypeptide in the first subject as compared to the control sample indicates the presence of or predisposition to said disease.

11. A method of identifying an agent that binds to the polypeptide of claim 1 , the method comprising: (a) introducing said polypeptide to said agent; and

(b) determining whether said agent binds to said polypeptide.

12. The method of claim 11 wherein the agent is a cellular receptor or a downstream effector.

13. A method for identifying a potential therapeutic agent for use in treatment of a pathology, wherein the pathology is related to abenant expression or aberrant physiological interactions of the polypeptide of claim 1, the method comprising:

(a) providing a cell expressing the polypeptide of claim 1 and having a property or function ascribable to the polypeptide;

(b) contacting the cell with a composition comprising a candidate substance; and (c) determining whether the substance alters the property or function ascribable to the polypeptide; whereby, if an alteration observed in the presence of the substance is not observed when the cell is contacted with a composition in the absence of the substance, the substance is identified as a potential therapeutic agent.

14. A method for screening for a modulator of activity of or of latency or predisposition to a pathology associated with the polypeptide of claim 1, said method comprising: (a) administering a test compound to a test animal at increased risk for a pathology associated with the polypeptide of claim 1, wherein said test animal recombinantly expresses the polypeptide of claim 1; (b) measuring the activity of said polypeptide in said test animal after administering the compound of step (a); and (c) comparing the activity of said polypeptide in said test animal with the activity of said polypeptide in a control animal not administered said polypeptide, wherein a change in the activity of said polypeptide in said test animal relative to said control animal indicates the test compound is a modulator activity of or latency or predisposition to, a pathology associated with the polypeptide of claim 1.

15. The method of claim 14, wherein said test animal is a recombinant test animal that expresses a test protein fransgene or expresses said transgene under the confrol of a promoter at an increased level relative to a wild-type test animal, and wherein said promoter is not the native gene promoter of said transgene.

16. A method for modulating the activity of the polypeptide of claim 1 , the method comprising contacting a cell sample expressing the polypeptide of claim 1 with a compound that binds to said polypeptide in an amount sufficient to modulate the activity of the polypeptide.

17. A method of freating or preventing a pathology associated with the polypeptide of claim 1, the method comprising administering the polypeptide of claim 1 to a subject in which such treatment or prevention is desired in an amount sufficient to treat or prevent the pathology in the subject.

18. The method of claim 17, wherein the subject is a human.

19. A method of treating a pathological state in a mammal, the method comprising administering to the mammal a polypeptide in an amount that is sufficient to alleviate the pathological state, wherein the polypeptide is a polypeptide having an amino acid sequence at least 95% identical to a polypeptide comprising the amino acid sequence selected from the group consisting of SEQ ID NO:2n, wherein n is an integer between 1 and 62 or a biologically active fragment thereof.

20. An isolated nucleic acid molecule comprising a nucleic acid sequence selected from the group consisting of SEQ ID NO:2n-l , wherein n is an integer between 1 and 62.

21. The nucleic acid molecule of claim 20, wherein the nucleic acid molecule is naturally occuπing.

22. A nucleic acid molecule, wherein the nucleic acid molecule differs by a single nucleotide from a nucleic acid sequence selected from the group consisting of SEQ ID NO: 2n-l, wherein n is an integer between 1 and 62.

23. An isolated nucleic acid molecule encoding the mature form of a polypeptide having an amino acid sequence selected from the group consisting of SEQ ID

NO:2n, wherem n is an integer between 1 and 62.

24. An isolated nucleic acid molecule comprising a nucleic acid selected from the group consisting of 2n-l, wherein n is an integer between 1 and 62.

25. The nucleic acid molecule of claim 20, wherein said nucleic acid molecule hybridizes under stringent conditions to the nucleotide sequence selected from the group consisting of SEQ ID NO: 2n-l, wherein n is an integer between 1 and 62, or a complement of said nucleotide sequence.

26. A vector comprising the nucleic acid molecule of claim 20.

27. The vector of claim 26, further comprising a promoter operably linked to said nucleic acid molecule.

28. A cell comprising the vector of claim 26.

29. An antibody that immunospecifically binds to the polypeptide of claim 1.

30. The antibody of claim 29, wherein the antibody is a monoclonal antibody.

31. The antibody of claim 29, wherein the antibody is a humanized antibody.

32. A method for determining the presence or amount of the nucleic acid molecule of claim 20 in a sample, the method comprising:

(a) providing said sample; (b) introducing said sample to a probe that binds to said nucleic acid molecule; and

(c) determining the presence or amount of said probe bound to said nucleic acid molecule, thereby determining the presence or amount of the nucleic acid molecule in said sample.

33. The method of claim 32 wherein presence or amount of the nucleic acid molecule is used as a marker for cell or tissue type.

34. The method of claim 33 wherein the cell or tissue type is cancerous.

35. A method for determining the presence of or predisposition to a disease associated with altered levels of expression of the nucleic acid molecule of claim 20 in a first mammalian subject, the method comprising: a) measuring the level of expression of the nucleic acid in a sample from the first mammalian subject; and b) comparing the level of expression of said nucleic acid in the sample of step (a) to the level of expression of the nucleic acid present in a control sample from a second mammalian subject known not to have or not be predisposed to, the disease; wherein an alteration in the level of expression of the nucleic acid in the first subject as compared to the control sample indicates the presence of or predisposition to the disease.

36. A method of producing the polypeptide of claim 1 , the method comprising culturing a cell under conditions that lead to expression of the polypeptide, wherein said cell comprises a vector comprising an isolated nucleic acid molecule comprising a nucleic acid sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62.

37. The method of claim 36 wherein the cell is a bacterial cell.

38. The method of claim 36 wherein the cell is an insect cell.

39. The method of claim 36 wherein the cell is a yeast cell.

40. The method of claim 36 wherein the cell is a mammalian cell.

41. A method of producing the polypeptide of claim 2, the method comprising culturing a cell under conditions that lead to expression of the polypeptide, wherein said cell comprises a vector comprising an isolated nucleic acid molecule comprising a nucleic acid sequence selected from the group consisting of SEQ ID NO:2n-l, wherein n is an integer between 1 and 62.

42. The method of claim 41 wherein the cell is a bacterial cell.

43. The method of claim 41 wherein the cell is an insect cell.

44. The method of claim 41 wherein the cell is a yeast cell.

45. The method of claim 41 wherein the cell is a mammalian cell.