MXPA97006873A - Hialurone receptor expressed in endothelial cells of vena umbilical huma - Google Patents

Abstract

The present invention provides nucleotide and amino acid sequences that identify and encode the hyaluronan (RH) receptor of human umbilical vein endothelial cells. The present invention also provides antisense molecules to the nucleotide sequences encoding rh, expression vectors for the production of purified RH, antibodies capable of specifically binding to RH, hybridization probes or oligonucleotides to detect up-regulation of nucleotide sequences that encode RH, host cells genetically treated for the expression of RH, diagnostic tests for cells and / or inflamed or metastatic tissues, activated, angiogenic, based on nucleic acid molecules that encode RH and antibodies capable of binding specifically to the recept

Description

HIALURONE RECEPTOR EXPRESSED IN ENDOTHELIAL CELLS OF HUMAN UMBILICAL VEIN TECHNICAL FIELD The present invention relates to polynucleotide and polypeptide sequences of a novel hyaluronan receptor and to the use of these sequences in the diagnosis, study and treatment of disease. TECHNICAL BACKGROUND All blood vessels are composed of three layers or tunics. The tunica intima consists of endothelial cells that line the vessel and rest on the basal lamina or middle layer. The subendothelial layer consists of loose connective tissue and may contain smooth muscle cells. The endothelial cells are generally polygonal and elongated in the direction of blood flow. The nucleus of the endothelial cells bulges in the capillary lumen and the Golgi complex is located in the nuclear poles. A few mitochondria, free ribosomes and rough endoplasmic reticulum are present. The endothelial cells are held together by zone occluders and occasional desmosome; there are separate junctions which offer variable permeability to macromolecules. The body of Weibel Palade, a cytoplasmic inclusion in the form of a bar is characteristic of these cells. Vascular endothelial cells play a central role in physiological homostasis, blood vessel patency, response to physiological and pathological stimuli. The endothelium is a prime target for cardiovascular risk factors such as high blood pressure, shear stress and arteriosclerosis. It is sensitive to endothelin, growth factors, interleukin 1, epinephrine, angiotensin, vasopressin arginine, heparin, bradychitinin, acetylcholine, prostacyclin, etc. Hyaluron (HA) is a highly charged, negatively charged connective tissue polysaccharide found in the extracellular matrix of most animal tissues. It is synthesized in the plasma membrane of fibroblasts and other cells and is catabolized locally as well as in the lymph nodes or sinusoids of the liver. HA is commonly isolated from the vitreous body of the eye, synovial fluid, umbilical cord and skin. It has several physiological functions including roles in water and plasma protein homeostasis; mitosis, cell migration and differentiation including angiogenesis (Rooney P and Kumar S (1994) EXS (Switzerland) 70: 179-90); and remodel tissue, either as a normal event or associated with tumors. The effects induced by the matrix in the cells are directed by a wide variety of proteins that bind to HA such as the hyaluronan (RH) receptor. The widespread presentation of RH indicates its importance in tissue organization and cell behavior control. The family is known as the hiladherines and includes those that bind to HA which act as part of the structural matrix and those that interact with HA in the plasma membrane of cell surface matrix receptors. With the recognition of the cell surface receptor of Hyaluron (RH), cell biologists, pathologists and immunologists have begun to investigate the importance of HA and RH for their potential diagnosis and therapeutic value. Hialadherins of Matrix The RH found within the cartilage matrix have been well characterized. Aggrecan is the chondroitin sulfate proteoglycan with large aggregation of cartilage that has a high affinity for AH. The binding protein is a 45-48 kDa glycoprotein that also demonstrates strong specific binding affinity. HA can bind to more than 100 aggrecans and binding protein molecules in a supramolecular complex which confers the viscoelastic properties of cartilage. Other matrix proteins such as PG-M and type VI collagen that participate in assembly and integrity may also be involved. HA binding proteins are also found in non-cartilage tissues. Versican of fibroblasts, hyururonectin of nervous and soft connective tissues, glial hyaluron binding protein in the central nervous system and neurocaine, a chondroitin sulfate proteoglycan of the brain also form strong structural complexes with HA. All hialoadherins in the matrix contain tandem repeated B loops, a structural motif that is thought to contain the HA binding domain. The hialoadherins of RH have been detected in several cell types from a wide variety of tissues.

Some reports suggest that the RH refer to the CD44 family of lymphocytes that host receptors which include the isoforms, Pgp-1, Hermes antigen, h-CAM and ECMRIII. The distal extracellular domain of CD44 has sequence homology for one of the B loop motifs of binding protein. The numerous other RH isCDs without CD44 include cell surface antigens called Ivd4 that block the binding of HA, liver endothelial cell receptors (HSC) that are involved in removing HA from the circulation and RH produced by fibroblasts that can be located on the surface. cell where the locomotion of cells induced by HA mediates. Its soluble form of 58 kDa contains an HA binding component unrelated to the loop motif of B and is known as a receptor for HA-mediated mobility (RMMHA). The important distinctions between the cell surface and matrix haloadherins are 1) the hexasaccharides of HA represent the minimum size molecule that interacts with these cell surface receptors, 2) the binding affinity increases with the increasing length of polymers, and ) the union increases with the increase in ionic resistance of the buffer solution. Cell Migration The presence of increased HA matrix has been correlated with cell migration in embolgenesis, limb regeneration, wound healing and tumor invasion. Since it has been shown that CD44 RH is associated with cytoskeletal actin, the proteins of the RH complex can affect the reorganization of the actin cytoskeleton and other activities such as cell disorder, substrate separation and locomotion necessary for cell migration, EMMHA , as one of the proteins of the RH complex, binds to HA with high affinity and is expressed only in the main lamina and perinuclear regions of the migration fibroblasts. Since RMMHA does not include a transmembrane hydrophobic region, it is assumed to be a peripheral protein associated with membrane-bound, intracellular tyrosine kinase. In studies of HA-controlled administration of a tyrosine kinase inhibitor, HA stimulated locomotion via a rapid tyrosine kinase signal transduction pathway. Invasion of the tumor and metastasis Invasive or metastatic cancer cells have the ability to exit the vascular system through the use of groups of molecules, at least one of which always has a receptor function. A number of such groups may include successive interactions between integrin of VLA-4 and endothelial E-selectin, subendothelial IV collagen and β-4 integrin, and HA of soft connective tissue and CD44 or HR interactions (Zetter BR (1993) SeinCancer Biol 4 : 215-218). Some tumor cells also have the ability to resemble enriched pericellular HA matrices which reduce cell adhesion to the outside of the growth tumor and protect the tumor from immune survival. In addition, the presence of high HA attracts endothelial cells that are active in angiogenesis. The combination of these HA functions allows the rapid establishment and growth of invasive tumor cells. The transforming oncogenic H-ras can promote cellular locomotion. Hardwick et al. (1992 J Cell Biol 117: 1343-1350) reported that H-ras actually regulates the expression of RMMHA, showed binding between HA and RMMHA and produced an antibody to the protein that is capable of inhibiting HA locomotion. RH. DESCRIPTION OF THE INVENTION The present invention provides a nucleotide sequence that encodes only a novel human hyaluron receptor. The cDNA, known as rh, was contained completely within Clone Incyte No. 3920 and encodes a polypeptide designated RH. The invention also comprises diagnostic tests for physiological or pathological compromise including the steps of testing a sample or an extract with rh nucleic acids, fragments or oligomers thereof. Aspects of the invention include rh antisense DNA; cloning or expression vectors containing rh; host cells or organisms transformed with expression vectors containing rh; a method for the production and recovery of purified RH from host cells; and purified protein, RH, which can be used to generate antibodies and other molecules for diagnosis of inflamed or metastatic, angiogenic, activated cells and / or tissues. BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 shows the amino acid alignment (aa) of RH, Sec ID NO: 2, with mouse hyaluron receptor, SEQ ID NO: 3. The amino acid without partner in the middle of the sequence can reflect the position of a mouse intron. The cDNA lacks an intron since it was constructed from mRNA. The alignments show where they were produced using the DNASTAR software multisequence alignment program (DNASTAR Inc. Madison Wl). Figure 2 shows a hydrophobicity analysis of RH based on the sequence and composition of predicted amino acids. MODES FOR CARRYING OUT THE INVENTION Definitions As used herein, the term "RH" in the above case, refers to polypeptides that are encoded by transcribed mRNAs of the cDNA described in SEQ ID NO. 1. The RH of the present invention can be naturally occurring or chemically synthesized. As used herein, the term "active" refers to choosing forms of RH that retain the biological and / or immunological activities of any naturally occurring RH. As used herein, the term "naturally occurring RH. "refers to the RH produced by human cells that have not been genetically treated and specifically contemplates several RHs that arise from post-translational modifications of the polypeptide including but not limited to acetylation, carboxylation, glycosylation, phosphorylation, lipidation and acylation.

As used herein, the term "derivative" refers to RHs chemically modified by such techniques as ubiquitination, labeling (e.g., with radionuclides, various enzymes, etc.), pegylation (derivatization with polyethylene glycol), and insertion or substitution by chemical synthesis of amino acids, such as ornithine, which does not normally occur in human proteins. As used herein, the term "recombinant variant" refers to any RH polypeptide that differs from an RH that occurs in nature by insertions, deletions, and amino acid substitutions created using recombinant AD techniques. The guide can be replaced to determine which amino acid residues can be replaced, added or deleted, without abolishing the activities of interest, such as cell adhesion and chemotaxis, can be found by comparing the sequence of the particular RH with that of the homologous receptors and reducing the minimum the number of amino acid sequence changes made in high homology regions. Preferably, amino acid "substitutions" are the result of replacing an amino acid with another amino acid having similar structural and / or chemical properties, such as replacing a leucine with a solucin or valine, an aspartate with a glutamate, or a threonine with a serine, that is, conservatives as replacements. "The intersections" or "deletions" are usually on the scale of approximately 1 to 5 amino acids. The allowed variation can be determined experimentally by making insertions, deletions or substitutions systematically, of amino acids in an RH molecule using recombinant DNA techniques and analyzing the resulting recombinant variants for activity. Where desired, a "leader signal or sequence" can direct the polypeptide through the membrane of a cell. Said sequence may be present in nature on the polypeptides of the present invention or provided for sources of heterologous proteins by recombinant DNA techniques. A "fragment," "portion," or "segment" of polypeptide is an extension of amino acid residues of at least about 5 amino acids, often of at least about 7 amino acids, typically at least about 9 to 13 amino acids, and, in various embodiments, at least about 17 or more amino acids. To be active, any RH polypeptide must be of sufficient length to exhibit biological and / or immunological activity. A "fragment", "portion", or "segment" of "oligonucleotide" or polynucleotide, is an extension of nucleotide residues that is long enough to be used in polymerase chain reaction (PCR) or various hybridization methods to amplify or simply reveal the related parts of mRNA or DNA molecules. The present invention includes purified RH polypeptide from natural or recombinant sources, cells transformed with nucleic acid molecules encoding RH. Various methods for the isolation of RH polypeptide can be achieved by methods well known in the art. For example, said polypeptide can be purified by immunoaffinity chromatography using the antibodies provided by the present invention. Other different protein purification methods well known in the art include those described in Deutscher M (1990) Methods in Enzymology, Vol. 182, Academic Press, San Diego: and Scopes R (1982) Protein Purification: Principies and Practice, Springer- Verlag, NYC, both incorporated here by reference. "Recombinant" can also refer to a polynucleotide that encodes RH and is prepared using recombinant DNA techniques. The DNA encoding RH may also include allelic or recombinant variants and mutants thereof. "Oligonucleotides" or "nucleic acid probes" are prepared based on the cDNA sequence encoding RH provided by the present invention Oligonucleotides comprise portions of the DNA sequence having at least about 15 nucleotides, usually at least about 20 nucleotides. Nucleic acid probes comprise portions of the sequence having fewer nucleotides than about 6 kb, usually less than about 1 kb. After the appropriate test to eliminate false positives, these probes can be used to determine whether the mRNAs encoding RH are present in a cell or had or isolate chromosomal DNA nucleic acid sequences as described by Walsh PS et al. (1992). PCR Methods Appl. 1: 241-250). The probes can be derived from single-stranded or double-stranded nucleic acids that are naturally occurring or recombinant or can be chemically synthesized. They can be labeled by nick translation, Klenow fill reaction, PCR or other methods well known in the art. The probes of the present invention, their preparation and / or labeling are elaborated in Sambrook J et al. (1980) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, NY; or Ausubel FM others (1989) Current Protocols in Molecular Biology John Wiley & Sons, NYC, both of which are incorporated here by reference. "Activated" cells, as used in this application, are those that are coupled in migration, proliferation, vascularization or differentiation as part of a normal or disease process. Recombinant variants that encode the same or similar polypeptides can be synthesized or selected using "redundancy" in the genetic code. Several codon substitutions, such as silent changes that produce several restriction sites, can be introduced to optimize cloning in a plasmid or viral vector or expression in a particular prokaryotic or eukaryotic system. Mutations can also be introduced to modify the properties of the polypeptide, to change the binding affinities of the ligand, interchain affinities or degradation of polypeptides or rate of rotation. INDUSTRIAL APPLICABILITY The present invention provides a nucleotide sequence identifying only a novel human hyaluron receptor, RH, which was highly expressed in the endothelial bank of human umbilical vein. Since RH is specifically expressed in embryonic tissue, nucleic acid (rh), polypeptide (RH) and antibodies to RH are useful in diagnostic tests for invasive cancers. Excessive expression of RH can direct, cell migration, including lymphocytes and / or other cells that respond to hyaluron. Therefore, a diagnostic test for the excess expression of RH can accelerate the diagnosis and appropriate treatment of an abnormal condition caused by viral or other infection: angiogenesis of cancerous tissues; invasive leukemias and lymphomas; or other physiological / pathological problems that deviate from normal development and result in metastatic cell migration, proliferation, vascularization, and differentiation. The nucleotide sequence encoding "RH (or its complement) has numerous applications in techniques known to those skilled in the art of molecular biology.These techniques include the use as hybridization probes, the use as oligomers for PCR, use for chromosome and gene mapping, use in the recombinant production of RH and use in the generation of antisense or RNA DNA, its similar chemical analogs The uses of the nucleotide sequences encoding RH described herein are illustrative of known techniques and are not it is intended that they limit their use in any technique known to one of ordinary skill in the art.In addition, the nucleotide sequences described herein can be used in molecular biology techniques that have not yet been developed, as long as the new techniques they refer to properties of nucleotide sequences that are currently known, such as the genetic code of triplet it is and interactions of specific base pairs. It will be appreciated by those skilled in the art that as a result of the degeneracy of the genetic code, a multitude of nucleotide sequences encoding RH can be produced, some carrying minimal homology to the nucleotide sequence of any gene known and present in nature. The invention has specifically contemplated each and every possible variation of the nucleotide sequence that could be made by selecting combinations based on possible codon choices. These combinations are made according to the normal triplet genetic code as applied to the nucleotide sequence of RH that occur in nature and all such variations should be considered as being specifically described. Although the nucleotide sequences encoding RH and its variants are preferably capable of hybridizing to the nucleotide sequence of the RH gene that is present in nature under restricted conditions, it may be advantageous to produce nucleotide sequences encoding RH or its derivatives having a substantially different codon usage. The codons can be selected to increase the rate at which expression of the peptide occurs in a particular prokaryotic or eukaryotic expression host according to the frequency with which the particular codons are used by the host. Other reasons for substantially altering the sequence of nucleotides encoding RH and its derivatives without altering the encoded amino acid sequence include the production of RNA transcripts that have more desirable properties, such as a longer half-life, than transcripts produced from the sequence that It is present in nature. The nucleotide sequence encoding RH can be linked to a variety of other nucleotide sequences by means of well-established recombinant DNA techniques (cf Sambrook J et al. (1989) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, NY) . Useful nucleotide sequences for binding to rh include a selection of cloning vectors, e.g., plasmids, cosmids, lambda phage derivatives, phagemids and the like, which are well known in the art. Vectors of interest include expression vectors, replication vectors, probe generation vectors, sequencing vectors and the like. In general, the vectors of interest may contain a functional origin of replication in at least one organism, convenient restriction endonuclease sensitive sites and selectable markers for the host cells. Another aspect of the present invention is to provide rh-specific nucleic acid hybridization probes capable of hybridizing to naturally occurring nucleotide sequences encoding RH. Such probes can also be used for the detection of sequences encoding similar hyaluron receptors and should preferably contain at least 50% of the nucleotides of this sequence encoding rh. The hybridization probes of the present invention can be derived from the nucleotide sequence of SEQ ID NO: 1 or promoter including genomic sequence, rh introns enhancing elements that are present in nature. Hybridization probes can be labeled by a variety of reporter groups, including radionuclides such as 32P or 3SS, or enzymatic labels such as alkaline phosphatase coupled with the probe via avidin / boutin coupling system and the like. PCR as described in U.S. Patent Nos. 4,683,1195; 4,800,195; and 4,965,188 provides additional uses for oligonucleotides based on the nucleotide sequences encoding RH. Said probes used in PCR can be recombinant origin, chemically synthesized, or a mixture of both and comprise a nucleotide sequence described for the diagnostic use or a degenerate well of possible sequences for identification of closely related genomic sequences.

Other means for producing hybridization probes specific for rh DNAs include the cloning of nucleic acid sequences encoding RH or RH derivatives into vectors for the production of mRNA probes. Such vectors are known in the art, and commercially available and can be used to synthesize RNA probes in vitro by means of the addition of the appropriate RNA polymerase such as T7 or SP6 RNA polymerase and the radioactively labeled nucleotides. It is now possible to produce a DNA sequence, or portions thereof, that encodes RH and its derivatives completely by synthetic chemistry, after which the gene can be inserted into any of the available DNA vectors using reagents and cells that are known in the matter at the time of filing this application. In addition, synthetic chemistry can be used to introduce mutations in the rh sequences or any portion thereof. The nucleotide sequence can be used to construct an assay to detect inflammation or disease associated with abnormal levels of RH expression. The nucleotide sequence can be labeled by methods known in the art and added to a sample of fluid or tissue from a patient under hybridization conditions. After an incubation period, the sample is washed with a compatible fluid which optionally contains a dye (or other tag that requires a developer) if the nucleotide has been labeled with an enzyme. After the compatible fluid is rinsed, the dye is quantified and compared to a normal one. If the amount of dye rises significantly, the nucleotide sequence has been hybridized with the sample and the analysis indicates the presence of inflammation and / or disease. The nucleotide sequence for rh can be used to construct hybridization probes to map that gene. The nucleotide sequence provided herein can be mapped to a chromosome and specific regions of a chromosome using well known genetic and / or chromosomal mapping techniques. These techniques include in situ hybridization, binding analysis against known chromosomal markers, hybridization screening with banks of chromosomal preparations selected in flux specific for known chromosomes and the like. The technique of fluorescent in situ hybridization of chromosome diffusions, among other places, has been described in Verma et al. (1988) Human Chromosomes: A Manual of Basic Techniques. Pergamon Press, New York NY. Fluorescent in situ hybridization of chromosomal preparations and other physical chromosome mapping techniques can be correlated with genetic map data. Examples of genetic map data can be found in the 1994 Genome Edition of Science (265: 1981f). The correlation between the location of rh on a map of physical chromosomes and a specific disease (or predisposition to a specific disease) can help to narrow down the DNA reaction associated with that genetic disease. The nucleotide sequence of the present invention can be used to detect differences in gene sequences between normal and carrier or affected individuals. The nucleotide sequence encoding RH can be used to produce purified RH using well known recombinant DNA technology methods. Among the many publications that teach methods for gene expression after they have been isolated is Goeddel (1990) Gene Expression Technology, Methods and Enzymology, Vol. 185, Academic Press, San Diego RH can be expressed in a variety of host cells, either prokaryotic or eukaryotic. The host cells can be of the same species in which the nucleotide sequences of rh are endogenous or of a different species. The advantages for producing RH by recombinant DNA technology include obtaining adequate amounts of the protein for purification and the availability of simplified purification procedures. Cells transformed with DNA encoding RH can be cultured under conditions suitable for the expression of hyaluronan receptors and recovery of the protein from the cell culture. The RH produced by a recombinant cell could be secreted or be contained intracellularly, depending on the particular genetic construction used. In general, it is more convenient to prepare recombinant proteins in secreted form.

The purification steps vary with the production process and the particular protein produced. In addition to recombinant production, fragments of RH can be produced by direct peptide synthesis using solid phase techniques (Stewart et al. (1969) Solid-Phase Peptide Synthesis, WH Freean Co. San Francisco: Merrifield J (1963) J Am Chem Soc 85: 2149-2154). The synthesis of in vitro proteins can be performed using manual techniques or by automation. Automated synthesis can be achieved, for example, using Applied Biosystems 431 A Peptide Synthesizer (Foster City, California) in accordance with the instructions provided by the manufacturer. Several fragments of RH can be chemically synthesized separately and combined using chemical methods to produce the full-length molecule RH for the induction of antibodies does not require biological activity; however, the protein must be immunogenic. The peptides used to induce specific antibodies can have an amino acid sequence consisting of at least five amino acids, preferably at least 10 amino acids. They must mimic a portion of the amino acid sequence of the protein and can contain the entire amino acid sequence of a small molecule present in nature similar to RH. The short extensions of RH can be fused with those other proteins such as orbital limpet hemocyanin and antibody produced against the chimeric molecule.

Antibodies specific for RH can be produced by inoculating an appropriate animal with the polypeptide or an antigenic fragment. An antibody is specific for RH if it is to be produced against an epitope of the polypeptide and binds to at least part of the natural or recombinant protein. The production of antibodies includes not only the stimulation of an immune response by injection in animals, but also the analogous steps in the production of synthetic antibodies or other specific binding molecules such as screening of recombinant immunoglobulin banks (Orlandi R. et al. 9089) PNAS 86: 3833-3837, or Huse WD et al. (1989) Science 256: 1275-1281) or in vitro stimulation of lymphocyte populations. Current technology (Winter G and Milstein C (1991) Nature 349: 293-299) provides a number of highly specific binding reagents based on the principles of antibody formation. These techniques can be adapted to produce molecules that specifically bind to RH. A further embodiment of the present invention is the use of antibodies specific for RH, inhibitors, or their analogs as bioactive agents to treat viral or other infections; angiogenesis of cancerous tissues; invasive leukemias and lymphomas; or other physiological / pathological problems that deviate from normal development and result in the migration, proliferation, vascularization and differentiation of metastatic cells. Bioactive compositions comprising RH agonists, antagonists or inhibitors can be administered in a suitable therapeutic dose determined by any of several methodologies including clinical studies in mammalian species to determine the maximum tolerable dose and in normal human subjects to determine the safe dose. Additionally, the bioactive agent can complex with a variety of well-established compounds or compositions that increase stability or pharmacological properties such as half-life. It is contemplated that the bioactive therapeutic composition may be delivered by intravenous infusion into the blood stream or any other effective means that could be used to treat invasive cancers. The following examples are provided to illustrate the present invention. These examples are provided by way of illustration and are not included for the purpose of limiting the invention. EXAMPLES I mRNA Isolation and Construction of the cDNA Bank The hyaluron cDNA sequence was identified among the sequences comprising the HUVEC library. The HUVEC cell line is an early, well-characterized, homogeneous, normal endothelial cell culture of the human umbilical vein (Cel Systems Corporation, 12815 NE 124th St. Kirkiand, WA 98034).

The HUVEC cDNA library was designed particularly by Stratagene (11099 M. Torrey Pines Rd., La Jolla, CA 92037). The cDNA synthesis was initiated with oligo dT hexamers and the synthetic oligonucleotide adapter was ligated into the cDNA ends to allow its insertion into the A? -ZAP 'vector system (Stratagene). This allowed high unidirectional efficiency (sense orientation) construction of lambda bank and the convenience of a plasmid system with blue / white color selection to detect clones with cDNA inserts. The quality of the cDNA library was screened using probes from DNA, and then, the phagemid was removed from pBluescript® (Stratagene). This phagemid allows the use of a plasmid system for easy insertion characterization, sequencing, site-directed mutagenesis, the creation of unidirectional deletions and expression of fusion polypeptides. Subsequently, phage particles from specially constructed banks were infected in the host strain of E. Coli XLI-Blue® (Stratagene). The high transformation efficiency of this bacterial strain increases the likelihood that the cDNA library will contain rare, underrepresented clones. Alternative unidirectional vectors should include, but are not limited to, pcDNAl (Invitrogen) and pSH1ox-1 (Novagen) II Isolation of cDNA Clones Phagemid forms of individual cDNA clones were obtained by the in vitro excision process, in the which coinfected XL1-CLUE with a phage f1 of help. Proteins derived from both lambda phage and phage f1 initiated new DNA synthesis of defined sequences in the lambda target DNA and created a smaller single-row circular phagemic DNA molecule that included all the DNA sequences of the pBluescript plasmid and the cDNA insert. The phagemid DNA was released from the cells and purified, then used to reinfect fresh bacterial host cells (SOLR, Stratagene), m where the double-stranded phagemid DNA was produced. Since the phagemid carries the gene for β-lactamase, the newly transformed bacteria were selected in the medium containing ampicillin. The phagemid DNA was purified using the QIAWELL-8 plasmid purification system of the QUIAGEN® DNA Purification System (QUIAGEN Inc. 9259 Eton Ave. Chatsworth, CA 91311). This technique provides a fast and reliable high-pass method for lysing bacterial cells and isolating highly purified phagemid DNA. The DNA eluted from the purification resin was suitable for sequencing DNA and other analytical manipulations. lll Sequencing of cDNA Clones The cDNA inserts of randomized isolates from the HUVEC bank were partially sequenced. Methods for sequencing DNA are well known in the art. Conventional enzymatic methods employed Klenow fragment of DNA polymer, SEQUENASE® (US Biochemical Corp. Cleveland, OH) or Taq polymerase to extend DNA strands of an annealed oligonucleotide primer to the DNA standard of interest. The methods have been developed for the use of single row patterns as double row. The chain termination reaction products were electrophored in urea-acrylamide gels and detected by autoradiography (for precursors labeled with radionuclides) or by fluorescence (for precursors labeled with fluorescers). Recent improvements in mechanical reaction preparation, sequencing and analysis using the fluorescent detection method have allowed the expansion in the number of sequences that can be determined per day (using machines such as Catalyst 800 and the DNA sequencer of Applied Biosystems 373 ). IV Homology Searching for cDNA Clones and Deduced Proteins Each sequence thus obtained was compared with the sequences in Gen Bank using a search algorithm developed by Appiíed Biosystems Inc. and incorporated in the INHERIT ™ 670 Sequence Analysis System. The algorithm, the Model Specification Language (developed by RTW Inc.) was used to determine regions of homology. The three parameters that determine the way in which the sequence comparisons operate are window size, window deviation and error tolerance. Using a combination of three parameters, the DNA database was investigated for sequences containing regions of sequence homology with interrogation and the appropriate sequences were classified with an initial value. Subsequently, these homologous regions were examined using dot matrix homology graphs to distinguish regions of pair homology of opportunities. The Smith-Waterman alignments were used to display the results of the homology search. Homologies of peptide and protein sequences were verified using the INHERIT ™ 670 Sequence Analysis System in a manner similar to that used in DNA sequence homologies. . The Pattern Specification Language and parameter windows were used to search for protein databases for sequences containing regions of homology which they classified with an initial value. The dot matrix homology plots were examined to distinguish regions of significant homology from pairs of opportunities. V Identification, Full Length Sequencing and Translation of the Gene The INHERIT ™ analysis of the randomly collected and sequenced portions of the HUVEC bank clones identified the partial sequence of Incyte 39200 as homologues for mouse hyaluron receptor (Hardwick and gold (1992) J Cell Biol 117: 1343 1350 ). The cDNA insert comprising Incyte 19200 was completely sequenced using the same methods described above. The coding region of the insert (ATG-> TGA) was identified and shown as SEQ ID NO: 1. This sequence for human rh was translated using DNASTAR software, frame translation was identified and shown in SEQ ID NO: 2. When the three possible predicted translations of the sequence were investigated against protein databases such as SwissProt and PIR, no exact pairs were found for possible translations of rh. Figure 1 shows the degree of amino acid homology between mouse RH and RMMHA. The unpaired amino acid in the middle of the sequence can reflect the position of a mouse intron. The cDNA lacks the intron since it was constructed from mRNA. Figure 2 shows the hydrophobicity graph for HR. VI Antisense Analysis Knowledge of the correct, complete cDNA sequence encoding RH will allow its use in antisense technology in gene function research. Either the oligonucleotides, the genomic or cDNA fragments comprising the antisense row of rh can be used either in vitro or in vivo to inhibit mRNA expression. Said technology is now well known in the art, the probes can be assigned to several locations along the nucleotide sequences. By treating the whole test cells or animals with said antisense sequences, the gene of interest can be turned off effectively. Frequently, the function of the gene can be verified by observing the behavior at the cellular, tissue or organism level (eg, lethality, loss of differentiated function, changes in morphology, etc.). In addition to using sequences constructed to interrupt transcription of the open reading frame, modifications of gene expression can be obtained by designing antisense sequences for regions of introns, promoter / enhancer elements, or even downstream regulatory genes. Similarly, inhibition can be achieved using Hogeboom's base pair methodology, also known as "triple helix" base pair formation. VII Expression of RH Expression of rh can be achieved by subcloning the cDNA into appropriate expression vectors and transfecting the vectors into appropriate expression hosts. In this particular case, the cloning vector previously used for tissue bank generation also provides direct expression of rh sequences in E. coli. Upstream of this cloning site, this vector contains a β-galactosidase promoter, followed by the sequence containing amino-terminal Met and 7 subsequent residues of β-galactosidase. Immediately after these eight residues there is a treated bacteriophage promoter useful for artificial priming and transcription and a number of unique restriction sites, including Eco RI, for cloning The induction of the transfected bacterial strain, isolated with IPTG using normal methods will produce a protein of fusion corresponding to the first seven ß-galactosidase residues, approximately 15"linker" residues and the peptides encoded within the cDNA. Since inserts of cDNA clones are generated by an essentially random process, there is an opportunity in three that the included cDNA will be in the correct frame for proper translation. If the cDNA is not in the proper reading frame, it can be obtained by deletion or insertion of the appropriate number of bases by well-known methods including in vitro mutagenesis, digestion with exonuclease III or nuclease of bean or inclusion of oligonucleotide linker. The rh cDNA can be fired in other vectors that are known to be useful for protein expression in specific hosts. Oligonucleotide extenders containing cloning sites as well as a DNA segment sufficient to hybridize the extensions at both ends of white cDNA (25 bases) can be chemically synthesized by normal methods. These primers can then be used to amplify the desired gene segments by PCR. The resulting new gene segments can be digested with appropriate restriction enzymes under normal conditions and isolated by gel electrophoresis. Alternatively, segments of similar genes can be produced by digestion of the cDNA with appropriate restriction enzymes and filling the segments of missing genes with chemically synthesized oligonucleotides. Segments of the coding sequence of more than one gene can be ligated together and cloned into appropriate vectors to optimize expression of recombinant sequence. Suitable expression hosts for said chimeric molecules include but are not limited to mammalian cells such as Chinese Hamster Ovary (OHC) and 293 human cells, insect cells such as Sf9 cells, yeast cells such as Saccharomyces cerevisiae. and bacteria such as E. Coli. For each of these cellular systems, a useful expression vector may also include an origin of replication to allow propagation in bacteria and a selectable marker such as the β-lactamase antibiotic resistance gene to allow selection in bacteria. In addition, the vectors may include a second labeled marker such as the neomycin phosphotransferase gene in order to allow selection in transfected eukaryotic host cells. Vectors for use in eukaryotic expression hosts may require RNA processing elements such as a 3 'polyadenylation sequence if such are not part of the cDNA of interest. Additionally, the vector may contain promoters or enhancers with increasing expression of genes. Such promoters are host specific and include MMTV, SV40, or metallothionin promoters for CHO cells; promoters of trp, lac, tac or T7 for bacterial hosts, or alpha factor, alcohol oxidase or promoters of PGH for yeast. Transcription enhancers, such as the sarcoma rous virus (RSV) enhancer, can be used in mammalian host cells. Once the homogeneous cultures of recombinant cells are obtained through normal culture methods, large quantities of recombinantly produced RH can be recovered from the conditioned medium and analyzed using chromatographic methods known in the art. VIII RH Recombinant Isolation RH can be expressed as a chimeric protein with one or more additional polypeptide domains added to facilitate the purification of proteins. Said purification facilitating the domain includes, but is not limited to, metal chelating peptides such as histidine-tryptophan modules that allow purification in immobilized metals, a protein domains that allow purification in immobilized immunoglobulin and the domain used in the extension / affinity purification system of "FLAGS" (Immunex Corp. Seattle WA). The inclusion of a cleavable binding sequence such as Factor ZA or enterokinase (Invitrogen, San Diego CA) between the purification domain and the rh sequence may be useful to facilitate the expression of RH. IX Production of Specific Antibodies for RH Two approaches are used to raise antibodies to RH and each approach is useful for generating any polyclonal or monoclonal antibody. In one approach, the denatured protein of the reverse phase HPLC separation is obtained in amounts up to 75 mg. This denatured protein can be used to immunize mice or rabbits using normal protocols; about 100 micrograms are suitable for immunization of a mouse, while up to 1 mg can be used to immunize a rabbit. To identify mouse hybridomas, the denatured protein can be radioiodinated and used to screen hybridoma cells of potential murine B cells for those that produce antibodies. This procedure requires only small amounts of protein, so that 20 mg may be enough to label and screen several hundred clones. In the second approach, the amino acid sequence of RH, as deduced from the translation of cDNA, is analyzed to determine regions of high immunogenicity. Oligopeptides comprising appropriate hydrophilic regions, as illustrated in Figure 2, are synthesized and used in suitable immunization protocols to raise the antibodies. The analysis for selecting appropriate epitopes is described by Ausubel FM et al. (1989, Current Protocols in Molecular Biology, John Wiley &Sons, NYC) Optimal amino acid sequences for immunization are usually in Termination C, the N terminus, and those regions hydrophilic polypeptide interventions that tend to be exposed to the external environment when the protein is in its natural conformation. Normally, the selected peptides, about 15 residues in length, are synthesized using an Applied Biosystems Model 431A Peptide Synthesizer using fmoc chemistry and coupled to orifice limpet hemocyanin (HLO, Sigma) by reaction with M-ester. maleimidobenzoyl-N-hydroxysuccinimide (MBS; cf Ausubel FM et al., supra). If necessary, a cysteine can be introduced at the N-terminus of the peptide to allow coupling to KLH. The rabbits are immunized with the peptide-KLH complex in Freund's complete adjuvant. The resulting antisera are tested for antipeptide activity by peptide-to-plastic binding, blocking with 1% BSA, reacting with antiserum, washing and reacting with specific goat anti-rabbit IgG, affinity purified, labeled (radioactive fluorescent). Hybridomas can also be prepared and sifted using standard techniques. Hybridomas of interest are detected by screening with labeled RH to identify those fusions that produce the monoclonal antibody with the desired specificity. In a normal protocol, the lacquer wells (FAST; Becton-Dickinson, Palo Alto, CA) are coated with specific anti-rabbit anti-mouse antibodies (or anti-Ig species), purified by affinity at 10 mg / ml. The coated wells are blocked with 1% BSA, washed and exposed to the supernatants of hybridomas. After incubation, the wells are exposed to labeled HR at 1 mg / ml. The antibodies that produce clones will bind to a quantity of labeled RH which is the previous detectable background. These clones are expanded and subjected to 2 cloning cycles in limiting dilution (1 cell / 3 wells). The cloned hybridomas are injected into primate mice to produce ascites, the monoclonal antibody is purified from ascitic fluid of mice by affinity chromatography on Protein A. Monoclonal antibodies with affinities of at least 10 9 to 10 10 or stronger will normally be made by normal procedures as described in Harlow and Lane (1988) Antibodies: A Laboratory Manual. Cold Spring Harbor Laboratory NY; and in Goding (1986) Monoclonal Antibodies. Principies and Practice, Academic Press, New York NY, both incorporated herein by reference. X Diagnostic Test Using Specific Antibodies for RH The particular RH antibodies are useful for the diagnosis of prepatological conditions and chronic or acute diseases that are characterized by differences in the amount or distribution of RH, respectively. To date, RH has only been found in the HUVEC bank and therefore is specific for abnormalities or pathologies that affect embryonic cells, angiogenic are invasive. Diagnostic tests for RH include methods that use the antibody and a label to detect RH in fluids of the human body, tissues or extracts of said tissues. The polypeptides and antibodies of the present invention can be used with or without modification. Frequently, polypeptides and antibodies will be labeled by joining them, either covalently or non-covalently, with a substance that provides a detectable signal. A wide variety of brands and conjugation techniques are known and have been reported extensively in both scientific and patent literature. Suitable labels include radionuclides, enzymes, substrates, cofactors, inhibitors, fluorescent agents, chemiluminescent agents, magnetic particles and the like. The patents that teach the use of said labels include the Patents of E.U.A. Nos. 3,817,837; 3,850,752; 3,939,350; 3,996,345; 4,277,437; 4,277,437; 4,275,149 and 4,366,241. Also, recombinant immunoglobulins can be produced as shown in the U.S. Patent. No. 4,816,567, incorporated herein by reference. A variety of protocols for measuring soluble or membrane-bound RH using polyclonal or monoclonal antibodies specific for the protein are known in the art. Examples include enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA) and fluorescent activated cell storage (FACS). A two-site monoclonal-based immunoassay using monoclonal antibodies reactive for two epitopes without interference to RH is preferred, but competitive binding analysis can be employed. These analyzes are described, among other places, in Maddox, DE and others (1983, J Exp Med 158: 1211). XI Native RH Purification Using Specific Antibodies The native or recombinant RH can be purified by immunoaffinity chromatography using RH-specific antibodies. In general, an immunoaffinity column is constructed by covalent coupling to the anti-RH antibody to an activated chromatographic resin. Polyclonal immunoglobulins are prepared from immune serum either by precipitation with ammonium sulfate or by purification on immobilized Protein A (Pharmacia LKB Biotechnology, Piscataway, NJ). Likewise, monoclonal antibodies are prepared from mouse ascites fluids by precipitation of ammonium sulfate or chromatography on immobilized Protein A. The partially purified immunoglobulin is covalently bound to a chromatographic resin such as Sepharose activated with CnBr (Pharmacia LKB Biotechnology). The antibody is couplant to the resin, the resin is blocked and the derived resin is washed according to the manufacturer's instructions. Said immunoaffinity columns are used in the purification of HR preparing a fraction of cells containing RH in a soluble form. This preparation is derived by solubilization of the whole cell or of a subcellular fraction obtained via differential centrifugation by the addition of detergent or by other methods well known in the art. Alternatively, the soluble RH containing a signal sequence can be secreted in a useful amount into the medium in which the cells grow. A preparation containing soluble RH was passed over the immunoaffinity column and the column was washed under conditions that allow preferential absorbance of RH (e.g., high ionic resistance of pH regulators in the presence of detergent). Then, the column was eluted under conditions that interrupt the binding of antibody / RH (e.g., a buffer solution of pH 2-3 or a high concentration of a chaotrope such as urea ion or thiocyanate) and RH was collected. XII Hyaluronan-Induced Chemotaxis for Cell Activation and Wound Healing Chemotactic interactions between HA and RH were measured in 48-well microchemotaxis chambers (see Falk WR et al. (1980) J Immunol Methods 33: 239). In each well, two compartments were prepared by a filter that allows the passage of cells in response to a chemical gradient. Cells expressing RH in a culture medium such as RPMI 1640 (Sigma, St. Louis MO) are placed on one side of a filter, usually polycarbonate and cells that produce HA or a solution enriched with HA are placed on the opposite side of the filter. Sufficient incubation time is allowed for the cells to pass through the filter in response to the concentration gradient through the filter. The filters are recovered from each well and the cells adhere to the side of the filter that looks at the HA are characterized and quantified. The cells that produce RH and migrate towards the upper end of the HA gradient are classified for chemotactic specificity. This analysis not only makes substantial the ability of cells that produce HR to respond to NH, but it also provides researchers with model systems from which they obtain and describe specific transcription factors and enhancers for use in regulating RH activity in populations of native cells. The ability to artificially supply said dissolved factors in dimethyl sulfoxide (DMSO) or some other liquid tester, to regulate the production of RH in a localized manner and to increase the migration capacity provides the use of HA as a stimulant in wound healing. . HA can be incorporated into collagen or other natural or artificial bandage materials used to treat refractory wounds. The presence of HA could attract activated endothelial cells and / or fibroblasts that could participate in the repair and healing processes. XIII Drug Screening This invention is particularly useful for screening compounds using RH or binding fragments thereof in any of a variety of drug screening techniques. The polypeptide or fragment employed in said test can be free in solution, fixed to a solid support, carried on a cell surface or located intracellularly. A drug screening method utilizes eukaryotic or prokaryotic host cells that are stably transformed with recombinant nucleic acids expressing the polypeptide or fragment. The drugs are screened against said transformed cells in competitive binding assays. Said cells, either in a viable or fixed form, can be used for normal binding analyzes. One can measure, for example, the formation of complexes between RH and the agent being tested. Alternatively, one can examine the decrease in complex formation between RH and hyaluron caused by the agent being tested. Therefore, the present invention provides methods for screening drugs or any other agents that can affect cell migration, angiogenesis or infiltration of lymphomas or leukemias. These methods comprise contacting an RH polymeric agent or a fragment thereof and analyzing (i) for the presence of a complex between the agent and the RH fragment or polypeptide, or (n) for the presence of a complex between the RH polypeptide or binding assay, the RH polypeptide or fragment is normally labeled After the appropriate incubation, the free RH polypeptide or fragment is separated from the present in bound form and the amount of free or non-complexed label is a measurement of the ability of the particular agent to bind to RH or to interfere with the RH complex and agent. Another technique for drug screening provides both step screening for compounds having sufficient binding affinity to the polypeptides and is described in detail in European Patent Application 84/03564, published September 13, 1984, incorporated herein by reference. . Briefly stated, large numbers of different small peptide test compounds are synthesized on a solid substrate, such as plastic pins or some other surface. The peptide test compounds are reacted with RH polypeptides and washed. The bound RH polypeptide is detected by methods well known in the art. The purified RH can also be coated directly on plates for use in the drug screening techniques mentioned above. further, the non-neutralizing antibodies can be used to capture the peptide and immobilize it on the solid support. This invention also contemplates the use of competitive drug screen analysis in which neutralizing antibodies capable of specifically binding to RH compete with a test compound to bind to RH polypeptides or fragments thereof. In this form, these antibodies can be used to detect the presence of any peptide that shares one or more antigenic determinants with RH. XIV Rational Drug Design The goal of rational drug design is to produce structural analogs of biologically active polypeptides of interest or small molecules with which they interact, eg, agonists, antagonists, or inhibitors. Any of these examples can be used to form drugs that are more active or stable forms of the polypeptide or that by increasing they interfere with the function of a polypeptide m vivo (Hodgson J (1991) Bio / Technology 9: 19/21, incorporated herein by reference ). In one approach, the three-dimensional structure of a protein of interest, or of a protein inhibitor complex, is determined by x-ray crystallography, by computer model or, more usually, by a combination of the two approaches. Both the form and the charges of the polypeptide must be verified to elucidate the structure and to determine the active site (s) of the molecule. Less frequently, useful information regarding the structure of a polypeptide can be obtained by modeling based on the structure of homologous proteins. In both cases, the relevant structural information is used to design efficient inhibitors. Useful examples of rational drug design may include molecules having enhanced activity or stability as shown by Braxton S and Wells JA (1992 Biochemistry 31: 7796-7801) or acting as inhibitors, agonists or antagonists of native peptides as shown by Athauda SB et al. (1993 J Biochem 113: 742-746), incorporated herein by reference. It is also possible to isolate a target-specific antibody, selected by functional analysis, as described before and after to resolve its crystal structure. This approach, in principle, produces a drug matrix upon which the design of the subsequent drug can be based. It is possible to derive protein crystallography together by generating anti-idiotypic antibodies (anti-ids) to a faramcologically active, functional antibody. "As a mirror image of a mirror image, the anti-ids binding site could be expected to be an analog of the original receptor." The anti-id could then be used to identify and isolate peptides from chemically or biologically produced peptide libraries. The isolated peptides could then act as the drug matrix In accordance with the present invention, a sufficient amount of polypeptides can be made available to perform such analytical studies as X-ray crystallography. In addition, knowledge of the amino acid sequence of RH provided herein will provide guidance to those who employ computer modeling techniques in place of or in addition to X-ray crystallography. XV Identification of Other Members of the RH Complex Purified RH is useful for characterization and purification of cell surface receptors associated binding molecules: cells that respond to HA by chemotaxis and other Specific responses probably express a receptor for RH and interact with transmembrane signaling molecules such as tyrosine kinase. Radioactive labels can be incorporated into RH by various methods known in the art. A preferred embodiment is the labeling of primary amino groups in RH with Bolton-Hunter Reagent 135l (Bolton, AE and Hunterm, WM (1973) Biochem J 133: 529), which has been used to label other labeling molecules without concomitant parasite deletion. biological activity (Hebert CA et al. (1991) J BioChem 266: 18989; McColl S et al. (1993) J Immunol 150: 4550-4555). The cells carrying receptors are incubated with the labeled signaling molecules. The cells are then washed to remove unbound molecules, and the labeled molecule bound to the receptor is quantified. The data obtained using different concentrations of RH is used to calculate values for the number, affinity and association of other members of the receptor complex. RH labeled is also useful as a reagent for the purification of the molecule (s) of this complex with which it interacts. In an affinity purification mode, RH is covalently coupled to a chromatography column. Cells and their membranes are extracted HA is removed and several HA-free subcomponents are passed over the column. The molecules associated with RH are bound to the column by virtue of their biological affinity. The RH complex is recovered from the column, dissociated and the recovered molecule is subjected to N-terminal protein sequencing. This amino acid sequence is then used to identify the molecule or to design degenerate oligonucleotide probes to clone the gene from an appropriate cDNA library. In an alternative method, the mRNA is obtained from the RH complex carrier cells and formed in a cDNA library. The library is transfected into a population of cells and the cells expressing the associated molecule (s) are selected using fluorescently labeled RH. The molecule is identified by recovering and sequencing the recombinant DNA from highly labeled cells. In another alternative method, the antibodies arise against RH, specifically monoclonal antibodies. The monoclonal antibodies are screened to identify those that inhibit the binding of the labeled RH. These monoclonal antibodies are then used in affinity purification or expression cloning of the associated labeling molecule. Other soluble binding molecules are identified in a similar manner. The labeled RH is incubated with extracts or other appropriate materials derived from HUVEC cells. After incubation, the RH complexes (which are longer than the size of purified RH molecule) are identified by a size technique such as size exclusion chromatography or density gradient centrifugation and are purified by known methods in the art The soluble binding protein (s) are subjected to N-terminal sequencing to obtain sufficient information for database identification, whether the soluble protein is known, or for cloning, if the soluble protein is unknown. XVI Use and Administration of RH Antibodies, inhibitors or antagonists of RH (or other treatments for excessive production of RH, abbreviated below TERH), can provide different effects when administered therapeutically. TERS will be formulated in a non-toxic, inert, pharmaceutically acceptable aqueous vehicle medium preferably at a pH of about 5 to 8, more preferably 6 to 8. Although the pH may vary according to the characteristics of the antibody, inhibitor or antagonist that is being formulated and the condition that will be treated. The characteristics of TERH include molecule solubility, half-life and antigenicides / immunogenicity; These and other characteristics can help define an effective vehicle. Native human proteins are preferred as TERH but organic or synthetic molecules resulting from drug screens can be equally effective in particular situations. RERH can be delivered by known routes of administration including by not limited to topical creams and gels; sprinklers and transmucosal spray, transdermal patches and bandages; injectable, intravenous and washing formulations; and orally administered fluids and pills are formulated in a particular manner to resist stomach acid and enzymes. The particular formulation, exact dose and route of administration will be determined by the attending physician and will vary according to each specific situation. These determinations are made considering multiple variables such as the condition that will be treated, the RHTE that will be administered and the pharmacokinetic profile of the particular RHTE. Additional factors that may be taken into account include disease status (eg, severity) of the patient, age, weight, gender, diet, time of administration, combination of drugs, reaction sensitivities and tolerance / response to therapy, long-acting RHTE formulations can be administered every 3 to 4 days, each emanates or once every two weeks depending on the half-life and space regime of the particular TERH. The amounts of normal doses may vary from .1 to 100, oOO micrograms, up to a total dose of approximately 1 g, depending on the route of administration. Instruction is given as to the dosage of particular methods of supply in the literature; see Patents of E.U.A. Nos. 4,657,760; 5,206,344 or 5,225,212. It is anticipated that different formulations will be effective for different TEHRs and that administration targeting metastatic cancers may need to be delivered in a manner different from that delivered to vascular endothelial cells. It is contemplated that conditions or diseases that activate leukocytes can precipitate damage that is treated with TERH.

These conditions or diseases can be diagnosed specifically by the tests treated before and such tests can be performed in suspected cases of viral or other infections; angiogenesis of cancerous tissues; invasive leukemias and lymphomas; or other physiological / pathological problems that deviate from normal development and result in migration, proliferation and metastatic cell differentiation. All publications and patents mentioned in the above specification are incorporated herein by reference. The above written specification is considered sufficient to allow a person skilled in the art to practice the invention. In addition, it is intended that various modifications of the modes described above for carrying out the invention that are obvious to those skilled in the field of molecular biology or related fields are within the scope of the following claims.

LIST OF SEQUENCES (I) GENERAL INFORMATION: (i) APPLICANT: Incyte Pharmaceuticals, Inc. (¡i) TITLE OF THE INVENTION: HIALURONE RECEPTOR EXPRESSED IN ENDOTHELIAL CELLS OF HUMAN UMBILICAL VEINS (iii) NUMBER OF SEQUENCES: 3 (iv) ADDRESS OF CORRESPONDENCE: (A) RECIPIENT: INCYTE PHARMACEUTICALS, INC. (B) STREET: 3174 Porter Drive (C) CITY: Palo Alto (D) STATE: CA (E) COUNTRY: USA (F) ZP: 94304 (v) FORM READING ON THE COMPUTER: (A) TYPE OF MEANS : Soft disk (B) COMPUTER: IBM compatible PC (C) OPERATING SYSTEM: PC-DOS / MS-DOS (D) SOFTWARE: Patentln Relay # 1.0, Version # 1.30 (vi) CURRENT REQUEST DATA: (A) NUMBER OF APPLICATION FOR TCP: To be assigned (B) DATE OF SUBMISSION: March 8, 1996 (C) CLASSIFICATION: (vii) DATE OF PREVIOUS APPLICATION: (A) REQUEST SERIES NO: US 08/402 217 (B) DATE OF PRESENTATION: March 10, 1995 (viii) APPORTER / AGENT INFORMATION: (A) NAME: Luther, Barbara J. (B) REGISTRATION NUMBER: 33954 (C) REFERENCE NUMBER / CASE: PF-0028 TCP (¡x ) TELECOMMUNICATION INFORMATION: (A) TELEPHONE: 415-855-0555 (B) TELEFAX: 415-852-0195 (2) INFORMATION FOR SEC ID NO: 1: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 1056 pairs of bases (B) TYPE: nucleic acid (C) THREADS: simple (D) TOPOLOGÍ A: linear (ii) TYPE OF MOLECULES: cDNA (vii) IMMEDIATE SOURCE: (A) BANK: Human umbilical vein endothelial cell (B) CLONES: 19200 (ix) DESCRIPTION OF SEQUENCES: SEQ ID NO: 1: ATG CAA AAC TTA AAA CAG AAG TTT ATT CTT GAA CAA CAG GAA CGT GAA 48 Met Gln Asn Leu Lys Gln Lys Phe lie Leu Glu Gln Gln Glu Arg Glu 1 5 10 15 AAG CTT CAÁ CAÁ AAA GAA TTA CAA ATT GAT TCA CTT CTG CAA CAÁ GAG 96 Lys Leu Gln Gln Lys Glu Leu Gln lie Asp Ser Leu Leu Gln Gln Glu 20 25 30 AAA GAA TTA TCT AGT CTT CAT CAG AAG CTC TGT TCT TTT CAA GAG 144 Lys Glu Leu Ser Ser Leu His Gln Lys Leu Cys Ser Phe Gln Glu 35 40 45 GAA ATG GCT AAA GAG AAG AAT CTG TTT GAG GAA GAA TTA AAG CAA ACÁ 192 Glu Met Ala Lys Glu Lys Asn Leu Phe Glu Glu Glu Glu Leu Glys Thr 50 55 60 CTG GAT GAG CTT GAT AAA TTA CAG CAA AAG GAG GAA CAA GCT GAA AGG 240 Leu Asp Glu Leu Asp Lys Leu Gln Gln Lys Glu Glu Gln Wing Glu Arg 65 70 75 80 CTG GTC AAG CAA TTG GAA GAG GAA GCA AAA TCT AGA GCT GAA GAA TTA 288 Leu Val Lys Gln Leu Glu Glu Glu Ala Lys Ser Arg Ala Glu Glu Leu 85 90 95 AAA CTC CTA GAA GAA AAG CTG AAA GGG AAG GAG GCT GAA CTG GAG AAA 336 Lys Leu Leu Glu Glu Lys Leu Lys Gly Lys Glu Wing Glu Leu Glu Lys 100 105 110 AGT AGT GCT GCT CAT ACC CAG GCC ACC CTG CTT TTG GAG GAA AAG TAT 384 Ser Ser Ala Ala His Thr Glp Ala Thr Leu Leu Leu Glu Glu Lys Tyr 115 120 125 GAC AGT ATG GTG CAA AGC CTT GAA GAT GTT ACT GCT CAA TTT GAA AGC 432 Asp Ser Met Val Gln Ser Leu Glu Asp Val Thr Wing Gln Phe Glu Ser 130 135 140 TAT AAA GCG TTA ACA GCC AGT GAG ATA GAA GAT CTT AAG CTG GAG AAC 480 Tyr Lys Ala Leu Thr Ala Ser Glu lie Glu Asp Leu Lys Leu Glu Asn 145 150 155 160 TCA TCA TGA CAG GAA AAA GTG GCC AAG GCT GGG AAA AAT GCA GAG GAT 528 Ser Ser Leu Gln Glu Lys Val Ala Lys Wing Gly Lys Asn Wing Glu Asp 165 170 175 GTT CAG CAT CAG ATT TTG GCA ACT GAG AGC TCA AAT CAA GAA TAT GTA 576 Val Gln His Gln lie Leu Ala Thr Glu Ser As Asp Gln Glu Tyr Val 180 185 190 AGG ATG CTT CTA GAT CTG CAG ACC AAG TCA GCA CTA AAG GAA ACA GAA 624 Arg Met Leu Leu Asp Leu Gln Thr Lys Ser Wing Leu Lys Glu Thr Glu 195 200 205 ATT AAA GAA ATC ACÁ GTT TCT TTT CTT CAA AAA ATA ACT GAT TTG CAG 672 He Lys Glu He Thr Val Ser Phe Leu Gln Lys He Thr Asp Leu Gln 210 215 220 AAC CAA CTC AAG CAA CAG GAG GAA GAC TTT AGA AAA CAG CTG GAA GAT 720 Asn Gln Leu Lys Gln Gln Glu Glu Asp Phe Arg Lys Gln Leu Glu Asp 225 230 235 240 GAA GAA GGA AGA AAA GCT GAA AAA GAA AAT ACÁ ACÁ GCA GAA TTA ACT 768 Glu Glu Gly Arg Lys Ala Glu Lys Glu Asn Thr Thr Ala Glu Leu Thr 245 250 255 GAA GAA ATT AAC AAG TGG CGT CTC CTC TAT GAA GAA CTA TAT AAT AAA 816 Glu Glu He Asn Lys Trp Arg Leu Leu Tyr Glu Glu Leu Tyr Asn Lys 260 265 270 ACA AAA CCT TTT CAG CTA CAA CTA GAT GCT TTT GAA GTA GAA AAA CAG 864 Thr Lys Pro Phe Gln Leu Gln Leu Asp Wing Phe Glu Val Glu Lys Gln 275 280 285 GCA TTG TTG AAT GAA CAT GGT GCA GCT CAG GAA CAG CTA AAT AAA ATA 912 Wing Leu Leu Asn Glu His Gly Wing Wing Gln Glu Gln Leu Asn Lys He 290 295 300 AGA GAT TCA TAT GCT AAA TTA TTG GGT CAT CAG AAT TTG AAA CAA AAA 960 Arg Asp Ser Tyr Wing Lys Leu Leu Gly His Gln Asn Leu Lys Gln Lys 305 310 315 320 ATC AAG CAT GTT GTG AAG TTG AAA GAT GAA AAT AGC CAA CTC AAA TCG 100B He Lys His Val Val Lys Leu Lys Asp Glu Asn Ser Gln Leu Lys Ser 325 330 335 GAA GTA TCA AAA CTC CGC TGT CAG CTT GCT AAA AAA AAA ACA AAG TGA 1056 Glu Val Ser Lys Leu Arg Cys Gln Leu Wing Lys Lys Lys Thr Lys * 340 345 350 (2) INFORMATION FOR SEQ ID NO: 2: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 352 amino acids (B) TYPE: amino acids (C) TOPOLOGY: linear (i) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF SEQUENCES: SEQ ID No. 2: Met Gln Asn Leu Lys Gln Lys Phe He Leu Glu Gln Gln Glu Arg Glu 1 5 10 15 Lys Leu Gln Gln Lys Glu Leu Gln He Asp Ser Leu Leu Gln Gln Glu 20 25 30 Lys Glu Leu Ser Ser Leu His Gln Lys Leu Cys Ser Phe Gln Glu 35 40 45 Glu Met Ala Lys Glu Lys Asn Leu Phe Glu Glu Glu Leu Lys Gln Thr 50 55 60 Leu Asp Glu Leu Asp Lys Leu Gln Gln Lys Glu Glu Gln Wing Glu Arg 65 70 75 80 Leu Val Lys Gln Leu Glu Glu Glu Wing Lys Ser Arg Wing Glu Glu Leu 85 90 95 Lys Leu Leu Glu Glu Lys Leu Lys Gly Lys Glu Wing Glu Leu Glu Lys 100 105 110 Being Wing Wing His Thr Gln Wing Thr Leu Leu Leu Glu Glu Lys Tyr 115 120 125 Asp Ser Met Val Gln Ser Leu Glu Asp Val Thr Wing Gln Phe Glu Ser 130 135 140 Tyr Lys Ala Leu Thr Wing Ser Glu He Glu Asp Leu Lys Leu Glu Asn 145 150 155 160 Being Ser Leu Gln Glu Lys Val Wing Lys Wing Gly Lys Asn Wing Glu Asp 165 170 175 Val Gln His Gln He Leu Wing Thr Glu Ser Asn Gln Glu Tyr Val 18J 185 190 Arg Met Leu Leu Asp Leu Gln Thr Lys Ser Wing Leu Lys Glu Thr Glu 195 200 205 He Lys Glu He Thr Val Ser Phe Leu Gln Lys He Thr Asp Leu Gln 210 215 220 Asn Gln Leu Lys Gln Gln Glu Glu Asp Phe Arg Lys Gln Leu Glu Asp 225 230 235 240 Glu Glu Gly Arg Lys Ala Glu Lys Glu Asn Thr Thr Ala Glu Leu Thr 245 250 255 Glu Glu He Asn Lys Trp Arg Leu Leu Tyr Glu Glu Leu Tyr Asn Lys 260 265 270 Thr Lys Pro Phe Gln Leu Gln Leu Asp Wing Phe Glu Val Glu Lys Gln 275 280 285 Wing Leu Leu Asn Glu His Gly Wing Wing Gln Glu Gln Leu Asn Lys He 290 295 300 Arg Asp Ser Tyr Ala Lys Leu Leu Gly His Gln Asn Leu Lys Gln Lys 305 310 315 320 He Lys His Val Val Lys Leu Lys Asp Glu Asn Ser Gln Leu Lys Ser 325 330 335 Glu Val Ser Lys Leu Arg Cys Gln Leu Wing Lys Lys Lys Thr Lys 340 345 350 (2) INFORMATION FOR SEQ ID NO: 3: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 477 amino acids (B) TYPE: amino acids (C) THREES: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULES: Protein (vii) IMMEDIATE SOURCE: (A) BANK: mouse (B) CLONES: Gl 53979 (ix) DESCRIPTION OF SEQUENCES: SEQ ID NO: 3 Met Gln He Leu Thr Glu Arg Leu Wing Leu Glu Arg Gln Glu Tyr Glu 1 5 10 15 Lys Leu Gln Gln Lys Glu Leu Gln Ser Gln Ser Leu Leu Gln Gln Glu 20 25 30 Lys Glu Leu Be Wing Arg Leu Gln Gln Gln Leu Cys Ser Phe Gln Glu 35 40 45 Glu Met Thr Ser Glu Lys Asn Val Phe Lys Glu Glu Leu Lys Leu Wing 50 55 60 Leu Wing Glu Leu Asp Wing Val Gln Gln Lys Glu Glu Gln Ser Glu Arg 65 70 75 80 Leu Val Lys Gln Leu Glu Glu Glu Arg Lys Ser Thr Ala Glu Gln Leu 85 90 95 Thr Arg Leu Asp Asn Leu Leu Arg Glu Lys Glu Val Glu Leu Glu Lys 100 105 110 His He Ala Ala Ala Ala Gln Ala He Leu He Ala Gln Glu Lys Tyr 115 120 125 Asn Asp Thr Ala Gln Ser Leu Arg Asp Val Thr Ala Gln Leu Glu Ser 130 135 140 Val Gln Glu Lys Tyr Asn Asp Thr Wing Gln Ser Leu Arg Asp Val Thr 145 150 155 160 Wing Gln Leu Glu Ser Glu Gln Glu Lys Tyr Asn Asp Thr Wing Gln Ser 165 170 175 Leu Arg Asp Val Thr Ala Gln Leu Glu Ser Glu Gln Glu Lys Tyr Asn 180 185 190 Asp Thr Wing Gln Ser Leu Arg Asp Val Thr Wing Gln Leu Glu Ser Val 195 200 205 Gln Glu Lys Tyr Asn Asp Thr Wing Gln Ser Leu Arg Asp Val Ser Wing 210 215 220 Gln Leu Glu Ser Tyr Lys Ser Ser Thr Leu Lys Glu He Glu Asp Leu 225 230 235 240 Lys Leu Glu Asn Leu Thr Leu Gln Glu Lys Val Wing Met Wing Glu Lys 245 250 255 Ser Val Glu Asp Val Gln Gln Gln He Leu Thr Wing Glu Ser Thr Asn 260 265 270 Gln Glu Tyr Wing Arg Met Val Gln Asp Leu Gln Asn Arg Ser Thr Leu 275 280 285 Lys Glu Glu Glu He Lys Glu He Thr Ser Ser Phe Leu Glu Lys He 290 295 300 Thr Asp Leu Lys Asn Gln Leu Arg Gln Gln Asp Glu Asp Phe Arg Lys 305 310 315 320 Gln Leu Glu Glu Lys Gly Lys Arg Thr Wing Glu Lys Glu Asn Val Met 325 330 335 Thr Glu Leu Thr Met Glu He Asn Lys Trp Arg Leu Leu Tyr Glu Glu 340 345 350 Leu Tyr Glu Lys Thr Lys Pro Phe Gln Gln Gln Leu Asp Wing Phe Glu 355 360 365 Wing Glu Lys Gln Wing Leu Leu Asn Glu His Gly Wing Thr Gln Glu Gln 370 375 380 Leu Asn Lys He Arg Asp Ser Tyr Ala Gln Leu Leu Gly His Gln Asn 385 390 395 400 Leu Lys Gln Lys He Lys His Val Val Lys Leu Lys Asp Glu Asn Ser 405 410 415 Gln Leu Lys Ser Glu Val Ser Lys Leu Arg Ser Gln Leu Val Lys Arg 420 425 430 Lys Gln Asn Glu Leu Arg Leu Gln Gly Glu Leu Asp Lys Wing Leu Gly 435 440 445 He Arg His Phe Asp Pro Ser Lys Wing Phe Cys His Wing Ser Lys Glu 450 455 460 Asn Phe Thr Pro Leu Lys Glu Gly Asn Pro Asn Cys Cys 465 470 475

Claims (12)

1. CLAIMS 1. An isolated DNA molecule comprising a sequence of nucleotides encoding human hyururon receptor (SEQ ID NO: 2).
2. 2. The DNA molecule of claim 1, wherein the nucleotide sequence comprises SEQ ID NO: 1 or its complement.
3. 3. A diagnostic test for the regulation of rh comprising the steps of: a) providing a biological sample; b) combining the biological sample with DNA of claim 1 or a fragment thereof.
4. 4. The diagnostic test of claim 3, wherein the regulation of rh is characteristic of activated, angiogenic, inflamed or metastatic cells and / or tissues.
5. 5. An expression vector comprising the DNA molecule of claim 1.
6. 6. A host cell transformed with the expression vector of claim 5.
7. 7. The antisense DNA of the DNA molecule of claim 1.
8. A method for producing hyaluronan receptor (RH) polypeptide, said method comprising the steps of: a) culturing host cells of claim 6, under conditions suitable for the expression of rh; and b) recovering RH from the cell culture.
9. 9. A purified RH polypeptide having the sequence as shown in SEQ ID NO: 2.
10. 10. An antibody specific for the polypeptide of the reiindication 9.
11. 11. A diagnostic test for the regulation of RH comprising the steps of a) providing a biological sample; and b) combining the biological sample with antibody of claim 9. The diagnostic test of claim 11, wherein the regulation of RH is characteristic of activated, angiogenic, inflamed or metastatic cells and / or tissues.