+

WO1994025589A1 - Recepteur des cannabinoïdes exprime dans les cellules du systeme immunitaire - Google Patents

Recepteur des cannabinoïdes exprime dans les cellules du systeme immunitaire Download PDF

Info

Publication number
WO1994025589A1
WO1994025589A1 PCT/GB1994/000863 GB9400863W WO9425589A1 WO 1994025589 A1 WO1994025589 A1 WO 1994025589A1 GB 9400863 W GB9400863 W GB 9400863W WO 9425589 A1 WO9425589 A1 WO 9425589A1
Authority
WO
WIPO (PCT)
Prior art keywords
leu
sequence
ser
ala
val
Prior art date
Application number
PCT/GB1994/000863
Other languages
English (en)
Inventor
Bruce Sean Munro
Original Assignee
Medical Research Council
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from GB939308679A external-priority patent/GB9308679D0/en
Priority claimed from GB939321697A external-priority patent/GB9321697D0/en
Application filed by Medical Research Council filed Critical Medical Research Council
Priority to AU65737/94A priority Critical patent/AU6573794A/en
Publication of WO1994025589A1 publication Critical patent/WO1994025589A1/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • C07K14/70571—Receptors; Cell surface antigens; Cell surface determinants for neuromediators, e.g. serotonin receptor, dopamine receptor

Definitions

  • This invention relates to novel nucleotide and amino acid sequences, vectors comprising the novel nucleotide sequence, and a method of screening for therapeutically useful compounds.
  • Delta -Tetrahydrocannabinol which is the major active ingredient of marijuana, has been used as a psychoactive agent for thousands of years.
  • delta -THC Delta -Tetrahydrocannabinol
  • Q marijuana and delta -THC have a wide range of other effects which have attracted attention because of their therapeutic potential. These include analgesia, anti- inflammation, immunosuppression, anti-convulsion, alleviation of intraocular pressure in glaucoma, and attenuation of vomiting, and this has led to interest in the biochemical bases of their action. Progress stemmed initially from the synthesis of potent derivatives of q delta -THC, and more recently from the cloning of a gene encoding a G-protein coupled receptor for cannabinoids
  • the present inventor has isolated the DNA sequence encoding a novel receptor for cannabinoids that is not expressed in the brain but is expressed in cells of the immune system.
  • the invention provides a nucleotide sequence encoding a cannabinoid receptor not normally expressed in mammalian brain tissue, or an effective part of such a receptor.
  • the receptor of the invention is a human receptor and preferably is expressed in cells of the immune system.
  • the invention provides a nucleotide sequence encoding a polypeptide having the amino acid sequence shown in Figure 1 or functional equivalents thereof.
  • the nucleotide sequence comprises substantially nucleotides 127-1206 of the sequence shown in Figure 1 or functional equivalents thereof.
  • Such functional equivalents include, for example: those sequences which encode the same polypeptide (but which, by virtue of the degeneracy of the genetic code, possess a different nucleotide sequence); sequences which encode substantially the same polypeptide but wherein there may be one or more conserved amino acid substitutions (i.e. the substitution of an amino acid for one with similar properties); and sequences which hybridize under standard conditions to the complement of nucleotides 127-1206.
  • Such functional equivalents will display at least 70%, more preferably at least 75%, and most preferably at least 80% nucleotide sequence homology with nucleotides 127-1206 of Figure 1.
  • a particular functional equivalent is the antisense equivalent of the sequence shown in Figure 1. Whilst such antisense sequences are not generally understood to be functional equivalents, the term functional equivalent as used in this application is intended to encompass such sequences.
  • sequence also comprises a suitable 5 1 untranslated region, including a promoter, to enable expression in appropriate host cells.
  • sequence also comprises a suitable 3' untranslated region.
  • sequence when expressed in a eukaryotic host cell the sequence should comprise a polyadenylation signal. Conveniently this can comprise nucleotides 1207-1790 of the sequence shown in Figure 1.
  • the invention provides a vector comprising the nucleotide sequence of the invention.
  • the vector allows for expression, either in a eukaryotic or prokaryotic host cell, such that the polypeptide encoded thereby may be produced, preferably substantially free of other substances found in the human body.
  • the invention provides a polypeptide being a cannabinoid receptor not normally expressed in mammalian brain tissue, or an effective part thereof.
  • the polypeptide is a human cannabinoid receptor, and preferably is expressed in cells of the immune system.
  • the polypeptide has substantially the amino acid sequence shown in Figure 1 , or is a functional equivalent thereof. It is proposed that this receptor be termed CB2, but is referred to herein for present purposes as CX5. It will be appreciated by those skilled in the art that one or more amino acids may be substituted by others having similar properties without significantly altering the function of the polypeptide.
  • a functionally equivalent polypeptide will possess at last 50% amino acid homology with the amino acid sequence shown in Figure 1 , more preferably at last 55%, and most preferably at least 60% amino acid homology.
  • nucleotide sequence of the present invention compared with that of the known brain cannabinoid receptor enables the design of oligonucleotide probes to conserved regions of the receptors which could be used to isolate other cannabinoid receptors in humans and possibly in other species.
  • novel receptor of the present invention presumably mediates some of the non-psychoactive effects of cannabinoid drugs. It could thus be used to screen for compounds which bind to the novel receptor but not to the brain receptor. Such compounds would be expected to possess at least some of the known therapeutic activities of cannabinoids without the psychoactive characteristics. Examples of natural and synthetic cannabinoids are disclosed by Snyder (1990, Nature 346, p508).
  • the invention provides a method of screening for compounds having a cannabinoid activity but with reduced psychoactive effects, comprising: determining the relative binding affinity of a compound for the human brain cannabinoid receptor and for a receptor encoded by the sequence of the invention; and selecting those compounds which show lower relative binding affinity for the human brain receptor.
  • Figure 1 shows a nucleotide sequence and the amino acid sequence encoded by it, both comprising sequences in accordance with the present invention, together with part of the nucleotide sequence of a functional equivalent;
  • Figure 2 shows a comparison between a prior art amino acid sequence and the amino acid sequence of a polypeptide in accordance, with the invention
  • Figure 3a is a graph of amount of compound bound against amount of compound initially added
  • Figure 3b is a graph of compound remaining bound against the concentration of competitor added
  • Figures 4a and 4b show photographs of a Northern blot
  • FIGS 5a-5c show photographs of in situ hybridisation experiments
  • Figure 6 shows the nucleotide sequence of a portion of a functional equivalent of the sequence of the invention and the amino acid sequence encoded thereby;
  • Figure 7 shows a comparison of the amino acid sequence of the invention with that of a portion of a functional equivalent.
  • PCR using degenerate primers was performed on cDNA prepared from the human promyelocytic leukemic line HL60. Treating HL60 cells with dimethyl formamide (DMF) induces granulocyte differentiation whilst 12-0- tetradecanoyphorbol 13-acetate (TPA) induces myeloid differentiation.
  • DMF dimethyl formamide
  • TPA 12-0- tetradecanoyphorbol 13-acetate
  • Oligo-dT primed cDNA was synthesised from poly(A)+RNA prepared from HL60 cells induced with 0.5% DMF for 3 days. 5ng of cDNA in 20ul was amplified with Taq polymerase using degenerate primers encoding regions conserved between many G-protein coupled receptors. These included the primers
  • GAGGGCCCATYNSNNTNGAYMGNTA (Seq. ID No. 1) and TGAAGCTTSHRTANANSANNGGRTT (Seq. ID No. 2), where each letter has the meaning ascribed in the standard IUPAC-IUB code which correspond to the encoded region shown in bold in Figure 2.
  • PCR was performed as follows: 40 cycles of 94 C C 1 min; 50 ⁇ C 2min; and 72 ⁇ C 2min, in 100mM Tris HCI pH8.3, 3mM Mg Cl 2 , 100mM tetramethylammonium chloride, 0.5% Tween 20, 0,5% NP40, 250uM dNTPs, using 20uM each primer.
  • Gel purified amplification products were digested with Apal and Hindlll and cloned into Bluescript.
  • the cloned amplification products from DMF-treated cells were sequenced and grouped into 6 classes which showed homology to the G-protein coupled receptor family. Two of these classes corresponded to previously identified receptors; the interleukin 8 receptor-B and the adenosine A3 receptor. Of the remaining four sequences, only one showed particular homology to any published receptors.
  • This clone, CX5 was related to a cannabinoid receptor cloned originally from rat brain. To investigate the functional significance of this homology, the CX5 insert was used to screen 200,000 colonies of a cDNA library from TPA-treated HL60 cells.
  • hCX5.1 (Seq. ID No. 5) and hCX5.36 (Seq. ID No. 3), the latter extending the furthest 5*.
  • the complete nucleotide sequence of hCX5.36 and the partial sequence of hCX5.1 is shown in Figure 1.
  • the polypeptide three letter code, Seq. ID No. 4
  • the polypeptide encoded by hCX5.36 exhibits 44% identity with the human brain cannabinoid receptor.
  • the transmembrane domains of G-protein coupled receptors are the most highly conserved parts, and the homology rises to 57% if these regions alone are considered (see Figure 2).
  • FIG. 1 Also shown in Figure 1 is the nucleotide sequence of part of hCX5.1 where it diverges from that of hCX5.36 at the poly (A) addition site.
  • the sequence of hCX5.1 is shown from the point equivalent to base 1714 of hCX5.36 onwards. Thus, the 3' untranslated region of hCX5.1 extends beyond that of hCX5.36.
  • Figure 2 shows the alignment between the proteins encoded by hCX5.36 and the previously reported human cannabinoid receptor (hCB-R). Identities are boxed and the seven putative transmembrane segments are underlined and numbered (Roman numerals). The most 5' ATG in CX5.36 has been assumed to be the initiator codon.
  • the cDNA clone was inserted into an expression vector and transfected into tissue culture cells, from which membrane preparations were obtained and analysed in binding studies together with control membrane preparations from cells expressing an irrelevant receptor.
  • Plasmid SCFRM2 is a cDNA for human formyl- peptide receptor (with a peptide-tag) in the same vector.
  • Figure 3a shows a graph of amount of radiolabelled CP 55,940 (a synthetic cannabinoid), in femtomoles, bound to CHOP cell membrane preparations against amount of CP 55,940 added.
  • the graph labelled 'specific' shows the specific binding taking place (i.e. binding to hCX5 minus the amount binding to the irrelevant control receptor).
  • control cells do not express receptors for cannabinoids, but expression of hCX5.36 causes the appearance of a saturable number of high affinity binding sites for CP55,940.
  • the archetypal cannabinoid (delta -THC) can compete for these binding sites as revealed by the displacement experiments (data shown in Figure 3b, which is a graph of amount of radiolabelled CP 55,940, in femtomoles, bound to COS cells against concentration of unlabelled delta 9 -THC).
  • the Kd for CP55,940 is 1nM (+/- 0.5nM)
  • the Ki for delta 9 -THC is 1OOnM (+/-50nM).
  • CX5 was used to probe Northern blots of RNA from HL60 cells and various rat tissues. The method was as follows: HL60 cells were induced with either 20nglml TPA or with 0.7% DMF.
  • the blot was exposed to Kodak XAR with an intensifying screen either for 8 days [ICAM] or 10 days [hCX5]). The blots were then stripped according to the manufacturer's instructions and reprobed with human gamma-actin, exposure 6 hours.
  • the results are shown in Figure 4a.
  • the left hand side shows a blot of RNA from cells induced with TPA (at 0-48 hours post-induction) and the right hand side shows the results with DMF-induced cells (at 0-96 hours post- induction).
  • the blot at the top was probed with CX5, the middle blot probed with the sequence encoding ICAM-1 and the bottom blot was probed with the sequence encoding gamma-actin.
  • the CX5 probe can be seen to hybridise to two transcripts of about 2.5 and 5.0 kb.
  • the longer transcript probably corresponds to use of a more 3' poly(A) addition site than that used in hCX5.36.
  • the hCX5.36 polyadenylation signal (GAUAAA) is a variant of the AAUAAA consensus that is found in a small fraction of messages and which can be used, albeit inefficiently, in vitro. It is apparent that there is some slight expression of CX5 in uninduced HL60 cells, but transcript levels are markedly increased upon myeloid or granulocyte differentiation. However, the gene does not appear to be expressed in mature neutrophils isolated from blood (data not shown).
  • GGGCTCGAGGTNRAYTTYCAYGTNTT (Seq. ID No. 6) and GAGGGATCCATNSWRCARAANGCRAA (Seq. ID No. 7, letters according to IUPAC-IUB convention) that encode sequences in hCX5 which are also found in the cannabinoid receptor but not in other G-protein coupled receptors (amino acid residues VNFHVF (91-96) and FAFCSM (279-284)).
  • Cloning and sequencing of PCR products of 600-650bp produced primarily the rat cannabinoid receptor, but also a second sequence with 87% homology to hCX5, which was termed rCX5.
  • the nucleotide (Seq. ID No.
  • Figure 4b shows the blots of RNA from the various tissues probed with rCX5 (top panel), re-probed with a sequence encoding the rat cannabinoid receptor (rCB-R, middle panel) and subsequently with the sequence encoding gamma actin (bottom panel).
  • the rat probe detects an mRNA of c2.5kb. in spleen, but not in a variety of other tissues.
  • the rCX5 transcript is not detected in brain even though the 6kb mRNA encoding the rat CB-R can be readily detected in the same sample.
  • the distribution of binding sites for cannabinoids in the brain corresponds well to the expression pattern of the mRNA encoding the brain receptor.
  • CX5 is expressed in a subset of these sites and so its expression level is too low to be detected in total brain mRNA.
  • horizontal sections of rat brain were probed by in situ hybridisation with labelled oligonucleotides corresponding to rat CB-R and to rCX5 as described in Example 4 below.
  • oligonucleotides were chosen from the anti-sense of the second cytoplasmic loop of CX5 (residues 132-147) or the equivalent region of the rat cannabinoid receptor, as this region is highly divergent between the two genes and between other G- protein coupled receptors, the sequence of the probes being:
  • GGTGACGATCCTCTTATAGGCCAGAGGCCTTGTAATGGATATGTA (Seq. ID No. 11).
  • the oligonucleotides were tailed with 35S-dATP and used at 0.5ng/ml to probe 50um sections.
  • Hybridisations were performed on parallel sections in the absence or presence ("+ cold comp") of an excess of unlabelled oligonucleotide. Sections were exposed to Kodak XAR film for 7 days and then to emulsion for 1 month.
  • Figure 5a shows four autoradiographs of in situ hybridisations of labelled oligonucleotides for rat cannabinoid receptor (r-CB-R, top two panels) and rat CX5 (bottom two panels) to horizontal sections of rat brain, in the absence (left hand side) or presence (right hand side, "+ cold comp") of excess unlabelled probe.
  • Figure 5b shows four autoradiographs of in situ hybridizations of the r-CX5 probe to pairs of transverse sections of the spleen without unlabelled competitor probe (A) or with competitor probe (B).
  • Figure 5c shows a photomicrograph of an in situ hybridization of the r-CX5 probe to the periphery of the white pulp in the spleen (an arrow headmarks the central arteriole) under bright field illumination with haematoxylin/eosin staining (L) and a parallel section under dark field illumination (D).
  • Figure 5a shows that as previously reported, the brain receptor has a widespread distribution with high levels of expression in the cortex, hippocampus, striatum and cerebellum. However, when adjacent sections were probed for rCX5, no expression could be detected in these, or any other, regions.
  • the rCX5 oligonucleotide does however hybridise to localised regions of the spleen ( Figures 5b- d). The expression appears concentrated in the marginal zone around the outside of the periarteriolar lymphoid sheaths.
  • splenic macrophages/monocytes were purified using FACS and the expression of CX5 examined using PCR. Expression was detected in the macrophage/monocyte population but not in the CD5-positive population used as a control, as described in Example 5 below.
  • Example 5 PCR analysis of rCX5 expression, rat splenocytes were separated by FACS using an antibody against CD11b (MRC 0X- 42), to label myeloid cells, and an antibody against CD5 (MRC 0X-19) to label T cells (antibodies obtained from Serotec).
  • Cytoplasmic RNA prepared from 3x10 cells by detergent lysis, (20 ug of glycogen added as carrier), was treated with ribonuclease-free DNase (0.5 unit/10 minutes; RQ1, Promega), phenol extracted, ethanol precipitated, resuspended in reverse transcriptase buffer with random primers, divided in two and M-MLV reverse transcriptase was added to one set (GIBCO). After 60 min.
  • PCR amplification was performed with the primer pairs TTTCACGGTGTGGACTCC (Seq. ID No. 12) and TAGGTAGGAGATCAAGCG (Seq. ID No. 13, from rCX5, giving a 214bp product) or, as a positive control, GAAATGCACCATGAAGCT (Seq. ID No. 14) and TTACGATGCATTGTTATC (Seq. ID No. 15) (specific for Elongation factor 1-alpha [EF-1a]) using 94 ⁇ C, 1 min 54 ⁇ C, 1 min, 72 ⁇ C 1 min with the manufacturer's buffer (Promega) .
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO (ill)
  • ANTI-SENSE NO
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • SEQUENCE DESCRIPTION SEQ ID NO: 2: TGAAGC ⁇ SH RTANANSANN GGRTT 25
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • SEQUENCE DESCRIPTION SEQ ID NO: 5: TATGCTATGA TGAGGA ⁇ AA GGTG ⁇ GACT TGCCTCTTTC AGAGATAAAT GACAAGCC ⁇ 60 CAG ⁇ TGGGG CATCCTG ⁇ G TfTG 84
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • SEQUENCE DESCRIPTION SEQ ID NO: 7: GAGGGATCCA TNS RCARAA NGCRAA 26
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO
  • ANTI-SENSE NO
  • MOLECULE TYPE cDNA
  • HYPOTHETICAL NO
  • ANTI-SENSE NO

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Cell Biology (AREA)
  • General Health & Medical Sciences (AREA)
  • Immunology (AREA)
  • Toxicology (AREA)
  • Zoology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Biomedical Technology (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Neurology (AREA)
  • Genetics & Genomics (AREA)
  • Medicinal Chemistry (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Engineering & Computer Science (AREA)
  • Peptides Or Proteins (AREA)
  • Saccharide Compounds (AREA)

Abstract

On décrit une séquence de nucléotides codant un récepteur des cannabinoïdes qui n'est pas normalement exprimé dans les tissus cérébraux chez les mammifères, ou une partie efficace d'un tel récepteur, ainsi qu'un vecteur et une cellule hôte contenant la séquence de l'invention.
PCT/GB1994/000863 1993-04-27 1994-04-22 Recepteur des cannabinoïdes exprime dans les cellules du systeme immunitaire WO1994025589A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU65737/94A AU6573794A (en) 1993-04-27 1994-04-22 Cannabinoid receptor expressed in cells of the immune system

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
GB939308679A GB9308679D0 (en) 1993-04-27 1993-04-27 Improvements in or relating to drug receptors
GB9308679.1 1993-04-27
GB9321697.6 1993-10-21
GB939321697A GB9321697D0 (en) 1993-10-21 1993-10-21 Improvements in or relating to drug receptors (2)

Publications (1)

Publication Number Publication Date
WO1994025589A1 true WO1994025589A1 (fr) 1994-11-10

Family

ID=26302818

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/GB1994/000863 WO1994025589A1 (fr) 1993-04-27 1994-04-22 Recepteur des cannabinoïdes exprime dans les cellules du systeme immunitaire

Country Status (2)

Country Link
AU (1) AU6573794A (fr)
WO (1) WO1994025589A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2002000712A1 (fr) * 2000-05-16 2002-01-03 Shanghai Biowindow Gene Development Inc. Nouveau polypeptide, recepteur cannabinoide central humain 22, et polynucleotide codant ce polypeptide

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1992002640A1 (fr) * 1990-08-08 1992-02-20 The United States Of America, Represented By The Secretary, United States Department Of Commerce Recepteur de cannabinoides

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1992002640A1 (fr) * 1990-08-08 1992-02-20 The United States Of America, Represented By The Secretary, United States Department Of Commerce Recepteur de cannabinoides

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
GERARD CM;MOLLEREAU C;VASSART G;PARMENTIER M;: "Molecular cloning of a human cannabinoid receptor which is also expressed in testis.", THE BIOCHEMICAL JOURNAL, vol. 279, no. 1, 1 October 1991 (1991-10-01), LONDON, GB, pages 129 - 134 *
MUNRO S;THOMAS KL;ABU-SHAAR M;: "Molecular characterization of a peripheral receptor for cannabinoids", NATURE., vol. 365, 2 September 1993 (1993-09-02), LONDON GB, pages 61 - 65 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2002000712A1 (fr) * 2000-05-16 2002-01-03 Shanghai Biowindow Gene Development Inc. Nouveau polypeptide, recepteur cannabinoide central humain 22, et polynucleotide codant ce polypeptide

Also Published As

Publication number Publication date
AU6573794A (en) 1994-11-21

Similar Documents

Publication Publication Date Title
Jones et al. Molecular cloning of five GTP-binding protein cDNA species from rat olfactory neuroepithelium.
Fong et al. Distinct forms of the beta subunit of GTP-binding regulatory proteins identified by molecular cloning.
Fukumoto et al. Characterization and expression of human HepG2/erythrocyte glucose-transporter gene
Klein et al. trkB, a novel tyrosine protein kinase receptor expressed during mouse neural development.
Munro et al. Molecular characterization of a peripheral receptor for cannabinoids
WO1994002605A1 (fr) Adn codant le recepteur de l'adenosine a1 humaine
Swaroop et al. Differential expression of novel Gsα signal transduction protein cDNA species
Mclean et al. Mapping of the α4 subunit gene (GABRA4) to human chromosome 4 defines an α2—α4—β1—γ1 gene cluster: Further evidence that modern GABAA receptor gene clusters are derived from an ancestral cluster
AU680852B2 (en) Stably transfected cell lines expressing GABA-A receptors
Satokata et al. Molecular basis of group A xeroderma pigmentosum: a missense mutation and two deletions located in a zinc finger consensus sequence of the XPAC gene
Iwata et al. Structure of the mouse tyrosine hydroxylase gene
AU657780B2 (en) Cloning and characterization of a cardiac adenylyl cyclase
WO1994025589A1 (fr) Recepteur des cannabinoïdes exprime dans les cellules du systeme immunitaire
Burghelle-Mayeur et al. Sequences of the swine 21-hydroxylase gene (CYP21) and a portion of the opposite-strand overlapping gene of unknown function previously described in human
Raming et al. Molecular cloning, sequence and expression of cDNA encoding a G0-protein from insect
Hibbs et al. Identification of a duplication of the mouse Lyn gene
CA2393447A1 (fr) Nouvelles proteines membranaires humaines et polynucleotides codant pour ces proteines
AU753400B2 (en) Orphan receptors
Yamashita et al. Genomic structure and chromosomal localization of the mouse LIM domain-binding protein 1 gene, Ldb1
CA2265467A1 (fr) Proteines g sensibles a l'anatoxine coquelucheuse, leur preparation et leur utilisation
US6570003B1 (en) Human 7TM proteins and polynucleotides encoding the same
EP1330524A2 (fr) Nouvelles proteines humaines 7tm et polynucleotides codant pour ces proteines
WO1995029234A1 (fr) Sous unite de recepteur gamma 3 de gaba-a humain et lignees cellulaires co-transfectees de maniere stable
WO1999012961A1 (fr) Recepteur d'oestrogene
RUSSEK Mapping of the cy4 Subunit Gene (GABRA4) to Human Chromosome 4 Defines an cu2-cw4-p,-y, Gene Cluster: Further Evidence That Modern GABA* Receptor Gene Clusters Are Derived from an Ancestral Cluster

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AT AU BB BG BR BY CA CH CN CZ DE DK ES FI GB GE HU JP KG KP KR KZ LK LU LV MD MG MN MW NL NO NZ PL PT RO RU SD SE SI SK TJ TT UA US UZ VN

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): AT BE CH DE DK ES FR GB GR IE IT LU MC NL PT SE BF BJ CF CG CI CM GA GN ML MR NE SN TD TG

DFPE Request for preliminary examination filed prior to expiration of 19th month from priority date (pct application filed before 20040101)
121 Ep: the epo has been informed by wipo that ep was designated in this application
REG Reference to national code

Ref country code: DE

Ref legal event code: 8642

122 Ep: pct application non-entry in european phase
NENP Non-entry into the national phase

Ref country code: CA

点击 这是indexloc提供的php浏览器服务，不要输入任何密码和下载