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WO2000073435A1 - Gene 441 associe a la pollinose - Google Patents

Gene 441 associe a la pollinose Download PDF

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Publication number
WO2000073435A1
WO2000073435A1 PCT/JP2000/003190 JP0003190W WO0073435A1 WO 2000073435 A1 WO2000073435 A1 WO 2000073435A1 JP 0003190 W JP0003190 W JP 0003190W WO 0073435 A1 WO0073435 A1 WO 0073435A1
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Prior art keywords
cells
preparing
antigen
compound
suppresses
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English (en)
Japanese (ja)
Inventor
Takeshi Nagasu
Yuji Sugita
Tomoko Fujishima
Tadahiro Oshida
Masaya Obayashi
Shigemichi Gunji
Izumi Obayashi
Yukiho Imai
Nei Yoshida
Kaoru Ogawa
Keiko Matsui
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Genox Research Inc
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Genox Research Inc
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    • 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
    • C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals

Definitions

  • the present invention relates to a gene associated with an antigen stimulatory response, a method for testing an allergic disease using the expression of the gene as an index, and a method for screening a candidate therapeutic compound for suppressing a T cell antigen stimulatory response.
  • Allergic diseases including hay fever, are considered multifactorial diseases. These diseases are caused by the interaction of the expression of many different genes, and the expression of these individual genes is affected by multiple environmental factors. Therefore, it is very difficult to elucidate the specific genes that cause specific diseases.
  • the differential display (DD) method is useful as such a method.
  • the differential display method was first developed in 1992 by Liang and Pardee (Science, 1992, 257: 967-971). By using this method, dozens of species at a time More than one sample can be screened, and genes with altered expression in those samples can be detected. Using such a method to examine genes with mutations or genes whose expression changes with time or environment is expected to provide important information for elucidating pathogenic genes. These genes include those whose expression is affected by environmental factors.
  • Allergic diseases such as hay fever are one of the diseases seen by many people in recent years.
  • the pathogenesis of hay fever may be related to several genes whose expression is affected by pollen, one of the environmental factors. Under such circumstances, it has been desired to isolate genes associated with allergic diseases such as hay fever. Disclosure of the invention
  • An object of the present invention is to provide a gene associated with an allergic disease. Furthermore, another object of the present invention is to provide a method for testing an allergic disease and a method for screening a candidate compound for a therapeutic agent that suppresses the antigen-stimulatory response of T cells, using the expression of the gene as an index.
  • the present inventors have proposed a method for treating a plurality of humans based on the already established “Fluorescent DD (Fluorescent DD) method” (T. I. to et al., 1994, FEBS Lett. 351: 231-236).
  • Fluorescent DD Fluorescent DD
  • the present inventors performed comparative analysis on the expression level of the isolated 441J gene in lymphocytes isolated from subjects before and after pollen scattering, and found that the gene showed a significantly low value after cedar pollen scattering. Therefore, the present inventors pointed out the expression level of the gene. As a standard, we found that it is possible to test for allergic diseases and to screen for therapeutic drug candidate compounds that suppress the T cell antigen-stimulated response.
  • the present invention relates to a gene showing a low value after pollen scattering, a method for testing allergic disease using the expression of the gene as an index, and a method for screening a candidate therapeutic compound for suppressing T cell antigen stimulation response. More specifically,
  • nucleic acid molecule comprising the nucleotide sequence of SEQ ID NO: 1;
  • nucleic acid molecule comprising the coding region of the nucleotide sequence of SEQ ID NO: 1;
  • step (f) selecting a compound that suppresses a decrease in the amount of RNA measured in step (e) as compared to a control (in the case where a test compound is not administered).
  • step (f) Amplification in step (e) compared to control (without test compound administration) Selecting a compound that suppresses the decrease in the amount of DM to be obtained.
  • step (h) selecting a compound that suppresses the decrease in the amount of RNA measured in step (g) as compared to a control (in the case where no test compound is administered).
  • step (h) selecting a compound that suppresses a decrease in the amount of DNA amplified in step (g) as compared to a control (in the case where no test compound is administered).
  • step (h) selecting a compound that suppresses a decrease in the amount of DNA amplified in step (g) as compared to a control (in the case where no test compound is administered).
  • step (g) selecting a compound that suppresses the decrease in the amount of RNA measured in step (f) as compared to a control (in the case where no test compound is administered).
  • step (g) selecting a compound that suppresses a decrease in the amount of DNA amplified in step (II) as compared to a control (in the case where no test compound is administered).
  • step (e) selecting a compound that suppresses the decrease in the amount of RNA measured in step (d) as compared to a control (in the case where no test compound is administered).
  • step (e) selecting a compound that suppresses a decrease in the amount of DNA amplified in step (d) as compared to a control (in the case where no test compound is administered).
  • lymphocytes are prepared from peripheral blood
  • allergic disease is a general term for diseases associated with allergic reactions. More specifically, allergens have been identified, demonstrated a deep link between exposure to allergens and the development of lesions, and the immunological mechanisms involved in the lesions. It can be defined as being proved.
  • the immunological mechanism means that T cells show an immune response by allergen stimulation.
  • Representative allergic diseases can include bronchial asthma, allergic rhinitis, atopic dermatitis, hay fever, or insect allergy.
  • Allergic predisposition (all ergi cdi athes is) is a genetic factor transmitted from parents to children with allergic diseases. Allergic diseases that occur familially are also called atopic diseases, and the genetic factors that cause them are atopic predisposition.
  • nucleic acid molecule in the present invention includes DNA and RNA.
  • the present invention relates to a novel gene “441” correlated with a response of lymphocytes to a cedar pollen antigen.
  • the nucleotide sequence of "441" cDNA found by the present inventors is shown in SEQ ID NO: 1.
  • the nucleotide sequence of the "441" cDNA isolated by the present inventors is a partial distribution sequence of the "441" cDNA, and those skilled in the art will be able to obtain the sequence information of the "441" cDNA described in SEQ ID NO: 1.
  • isolation of the full-length cDNA of "441" can be usually performed. That is, a method for screening a T cell cDNA library or the like by hybridization using a sequence derived from “441” as a probe, or a method for screening a T cell cDNA library using a sequence derived from “441” as a primer.
  • nucleic acid molecule comprising the base sequence of SEQ ID NO: 1 in the present invention includes the full length of “441” which can be isolated based on the sequence information of “441” cDNA described in SEQ ID NO: 1. cDNA is included. Five
  • “441” was statistically significantly lower in subjects' lymphocytes after exposure to pollen antigen than before exposure. Therefore, using the expression of “441” gene (including transcription into mRNA and translation into protein) as an index, we will conduct tests for allergic diseases and screen candidate therapeutic compounds that suppress antigen-stimulated response of T cells. It is considered possible. Decreased expression of “441” indicates the response of T cells to antigen stimulation such as pollen, so that in patients with a history of allergic disease, the response of that patient to specific antigen exposure using the expression of “441” as an index Monitoring the progress is useful for assessing disease status and examining response to treatment.
  • cedar pollinosis is particularly preferred.
  • the detection of the expression of the "441" gene in the test for allergic disease according to the present invention can be carried out by a hybridization technique using a nucleic acid hybridizing to the "441" gene as a probe, or a DNA hybridizing to the gene of the present invention as a primer. It is possible to use the gene amplification technology.
  • a nucleic acid molecule that specifically hybridizes to the “441” gene and has a chain length of at least 15 nucleotides is used.
  • the term “specifically hybridizes” as used herein refers to a DNA that is cross-hybridized with DNA and Z or RNA encoding another gene under ordinary hybridization conditions, preferably under stringent hybridization conditions. It indicates that one shot does not occur significantly.
  • the probe and the transfer membrane are hybridized at 68 in Express Hydidi zation on Solution (manufactured by CL0NTECH), and finally mixed with 50% IX SS 0.05% SDS solution at 50%. By washing with, stringent conditions can be achieved.
  • nucleic acid molecules used in the test of the present invention may be synthetic or natural.
  • the probe DNA used for hybridization is usually labeled.
  • Labels include, for example, nicks using DNA polymerase I. Labeling with lance label, end labeling with polynucleotide kinase, fill-in labeling with cleno fragment (Berger SL, Kimmel AR. (1987) Guide to Molecular Cloning Techniques, Method in Enzymology, Academic Press; Hames BD, Higgins SJ (1985) Genes Probes: A Practical Approach. IR L Press; Sambrook J, Fritsch EF, Maniatis T. (1989) Molecular Cloning: a Laboratory Manual, 2nd Edn.
  • an RT-PCR method can be used as a method using the gene amplification technique.
  • the expression of the “441” gene can be more accurately quantified by using a PCR amplification monitor method as shown in Example 8 in the process of gene amplification.
  • probes that are labeled with different fluorescent dyes at both ends to cancel each other's fluorescence are used to hybridize to the detection target (DNA or RNA reverse transcript).
  • the detection target DNA or RNA reverse transcript.
  • the two fluorescent dyes are separated and the fluorescence is detected. This fluorescence is detected in real time.
  • the number of copies of the target in the target sample is determined by the number of linear cycles of PCR amplification by simultaneously measuring the standard sample whose copy number is clear for the target (Holland, PM et al., 1991, Proc. Natl. Acad. Sci.
  • the test for an allergic disease of the present invention may be performed by detecting the protein encoded by “441”.
  • a Western blotting method using an antibody that binds to the protein encoded by “441”, an immunoprecipitation method, an EUSA method, and the like can be used.
  • the antibody of the protein encoded by "441" of the present invention can be obtained as a polyclonal antibody or a monoclonal antibody using techniques well known to those skilled in the art (Milisten C, et al., 1983, Nature 305 (5934): 537-40).
  • a protein or a partial peptide thereof used as an antigen is prepared by incorporating the "441" gene or a part thereof into an expression vector, introducing the gene into an appropriate host cell, and preparing a transformant. Is cultured to express a recombinant protein, and the expressed recombinant protein is purified from a culture or a culture supernatant.
  • an allergen such as cedar pollen antigen.
  • the measurement of the expression level of the gene of the present invention, together with the pollen-specific antibody titer, symptoms, etc., can be used for testing for allergic diseases.
  • the expression of the "441" gene expressed in T cells is reduced after pollen antigen exposure.
  • the "441" gene is a gene whose expression level decreases as a response of the living body to the stimulation of the cedar pollen antigen.
  • a therapeutic agent for hay fever can be screened.
  • the expression level of the “441” gene is decreased by pollen antigen exposure in both healthy and hay fever patients. Presence or absence of hay fever symptoms is presumed to be due to differences since the response of the "441" gene to antigen stimulation. However, even in such cases, decreased expression of the “441” gene corresponds to enhanced T cell responsiveness. 41 "By monitoring gene expression, it is possible to screen for drugs for treating allergic diseases.
  • the method for screening a candidate therapeutic compound that suppresses the antigen-stimulated response of T cells according to the present invention can be performed in vivo or in vitro.
  • In vivo screening for example, after administering a candidate drug and stimulating with a pollen antigen to a model animal such as a mouse, T cells are separated from peripheral blood, and the transcript of “441” is measured. .
  • a model animal such as a mouse
  • lymphocytes are separated from peripheral blood, and the lymphocytes are stimulated in vitro with cedar pollen antigen or the like. T cells are separated from the lymphocytes after the stimulation, and the transcript of the “441” gene is measured.
  • a compound that suppresses a decrease in the transcription level of the “441” gene compared to a control (when no candidate drug is administered) is selected.
  • the stimulation with the pollen antigen is performed for the purpose of eliciting an antigen-specific allergic reaction in T cells and determining the therapeutic effect of the candidate compound on it.
  • the term “suppressing the decrease in the transcription level of the“ 441 ”gene” means that a higher level of transcription level is maintained by contact with a candidate compound when compared with antigen-stimulated T cells.
  • a candidate compound induces a level that exceeds the transcription level of the “441” gene before being subjected to antigen stimulation (ie, if transcription is increased), that compound is a compound to be selected in the screening method of the present invention .
  • peripheral blood lymphocytes are collected from a human mouse or the like, and the peripheral blood lymphocytes are stimulated in vitro with cedar pollen antigen or the like.
  • Candidate compounds are added during in vitro stimulation.
  • T cells are separated from the stimulated peripheral blood lymphocytes, and the “441” transcript is measured.
  • a compound that suppresses a decrease in the transcription level of the “441” gene compared to a control (when no candidate drug is contacted) is selected.
  • the screening of candidate therapeutic compounds that suppress the antigen-stimulated response of T cells of the present invention can also be performed using established T cells.
  • Molt4 cells Jurka Cell lines such as t cells T cells are stimulated in vitro with a lymphocyte stimulator.
  • lymphocyte stimulants include calcium ionophore (A23187), PMA, and phytohemagglutinin (PHA).
  • Candidate drugs are added during in vitro stimulation. Thereafter, the transcription amount of the “441” gene in the established T cells is measured. As a result of this measurement, a compound that suppresses a decrease in the transcription level of the “441” gene compared to a control (when no candidate drug is contacted) is selected.
  • Detection of the expression of the "441" gene in the screening of a candidate therapeutic drug that suppresses the T cell antigen-stimulatory response according to the present invention can be performed by detecting the nucleic acid hybridizing to the "441" gene in the same manner as in the test for an allergic disease of the present invention.
  • the hybridization can be carried out using a hybridization technique using DNA as a probe, or a gene amplification technique using DNA that hybridizes to the gene of the present invention as a primer.
  • a method utilizing the hybridization technology for example, a Northern hybridization method, a dot blot method, a method using a DNA microarray, or the like can be used.
  • a method utilizing the gene amplification technique an RT-PCR method can be used. In the RT-PCR method, more accurate quantification of the expression of the “441” gene can be performed by using a PCR amplification monitor method as shown in Example 8 in the gene amplification process.
  • test compounds used in these screenings include compound preparations synthesized by existing chemical methods such as steroid derivatives, compound preparations synthesized by combinatorial chemistry, and extracts of animal and plant tissues or Examples thereof include a mixture containing a plurality of compounds such as a microorganism culture, and a sample purified therefrom.
  • the compound isolated by the method of the present invention for screening a candidate therapeutic compound for suppressing a T cell antigen-stimulated response is a candidate for a drug that suppresses an immune response.
  • the compound isolated by the screening method of the present invention when used as a pharmaceutical, it can be used as a pharmaceutical preparation by a known pharmaceutical production method.
  • pharmacologically acceptable carriers or vehicles saline, vegetable oils, suspensions, (Activators, stabilizers, etc.).
  • Administration will be transdermal, intranasal, transbronchial, intramuscular, intravenous, or oral, depending on the nature of the compound.
  • the dose varies depending on the patient's age, body weight, symptoms, administration method and the like, but those skilled in the art can appropriately select an appropriate dose.
  • FIG. 1 is a diagram showing the antibody titers of cedar pollen-specific IgE antibodies in a total of 18 blood samples from 10 subjects who collected blood.
  • Subject A ⁇ The values of cedar pollen-specific IgE antibodies of each blood sample (sample numbers 1 to 18) were expressed in AU / ml. The pair before pollen scattering is shown on the left (white column), and the one after scattering is shown on the right (black column). Subjects A and B collected only blood after pollen scattering.
  • FIG. 2 is a diagram showing changes in the expression of “441” in the pre-scattering group and the post-scattering group when grouping was performed before and after the cedar pollen scattering time. Error bars represent standard deviation. BEST MODE FOR CARRYING OUT THE INVENTION
  • Fig. 1 shows the measured cedar pollen-specific IgE values before and after pollen scattering in each subject. As shown, most of the 10 subjects had increased serum levels of cedar pollen-specific IgE after pollen exposure. The presence of atopic predisposition was determined by whether the value of the CAP RAST test for cedar pollen-specific IgE was greater than 2. That is, subjects A to G and I were eight subjects as atopic predisposition group (hereinafter also referred to as “patient”), and subjects H and ⁇ were regarded as healthy subjects (hereinafter also referred to as “normal group”). Of the eight subjects with an atopic predisposition, seven exhibited symptoms of allergic rhinitis after pollen dispersal. '
  • the procedure was as follows. First, the wall of the syringe was uniformly treated with 1 ml of Heparin from Nopo, etc., and blood was collected in a 10 ml syringe containing a final concentration of 50 unit / ml heparin. At this time, two 22G needles were prepared for one blood sample. The injection needle was removed and transferred to a 50 ml centrifuge tube (made of polypropylene).
  • the mixture was centrifuged at 1500 rpm for 5 minutes at room temperature, 1.1 ml was collected from the surface as close as possible, and centrifuged at 15000 rpm for 5 minutes at 4 to collect 1 ml of the supernatant as plasma.
  • An equal volume (9 ml) of 0.9% NaCl containing 3% dextran (manufactured by Nacalai) was added to the rest of the collected plasma, and the mixture was gently inverted several times to mix. Then, it was left still at room temperature for 30 minutes.
  • PRP Pla teletrich plasma, platelet-rich plasma
  • PRP Pla teletrich plasma, platelet-rich plasma
  • the precipitated cells were suspended in 5 ml of Ca- and Mg-free HBSS obtained from Gibco or the like.
  • the mixture was centrifuged at 1500 rpm (equivalent to 400 Xg in a Tomy centrifuge) for 30 minutes at room temperature.
  • granulocytes and erythrocytes precipitated, and lymphocytes (lymphocytes), monocytes, and platelets were contained in the middle layer with the ficoll layer in between.
  • Collect the intermediate layer with a Pasteur pipette, add 2 to 3 volumes of BSA / PBS (0.5% BSA, 2 mM EDTA in PBS, pH 7.2; degas immediately before use), and add 1200 rpm, 4 rpm. Centrifuged at for 5 minutes.
  • the precipitate was collected and washed twice with BSA / PBS. After the second washing, the cells were suspended in 5 ml, and a part thereof was diluted 2-fold with trypan blue, and the number of cells was counted. Total cell number was about IX 10 7. This was used as the lymphocyte fraction.
  • the lymphocyte fraction obtained in Example 2 was centrifuged at 1200 ⁇ ⁇ ⁇ at 4 for 5 minutes, and suspended in BSA / PBS at 10 8 per 100 ⁇ 1. The capacity became about 201. This was transferred to an Eppendorf tube (1.5 ml), and the CD3 microbead solution was added. Then, it was left at 4-10 for 30 minutes (at this time, it was not placed on ice). This sample was treated with a magnetic cell saw Yuichi (MACS) (manufactured by Miltenyi Biotech Inc.) as follows.
  • MCS magnetic cell saw Yuichi
  • the MS + / RS + column was attached to a Mini MACS or Vario MACS separation unit (without needles). 500 1 of BSA / PBS was gently applied to the column and the buffer was drained. Next, cells labeled with CD3 microbeads were applied to the column. The column was washed three times with 500 1 (B cell fraction). The column was removed from the separation unit and placed on a tube for collecting the eluate. 1 ml of BSA / PBS was applied to the column, and positive cells were rapidly flushed out using a plunger attached to the column. This was used as the T cell fraction.
  • the obtained T cell fraction was centrifuged at 1200 rpm at 4 for 5 minutes.
  • the precipitate was washed twice with BSA / PBS. After the second washing, the cells were suspended in 1 ml, a part thereof was diluted 2-fold with trypan blue, and the number of cells was counted. The total cell number was about 4 ⁇ 10 6 .
  • Example 4 Preparation of total RNA from T cells
  • RNeasy Mini manufactured by Qiagen
  • All operations were performed at room temperature, wearing gloves.
  • Four times the volume of ethanol was added to Posh Buffer-RPE.
  • 10 w 1 / ml 2-mercaptoethanol was added to the lysis buffer RLT.
  • the cell suspension was centrifuged at 1000-1200 rpm for 5 minutes, and the supernatant was removed by evaporation.
  • To the precipitate was added 350 1 lysis buffer RLT (containing 2-mercaptoethanol) solution.
  • the lysate of cells in the RLT buffer could be stored at _70 ° C.
  • the cell lysate had been stored frozen, incubate at 37 for 10-15 minutes, and if insolubles were visible, centrifuge for 3 minutes at maximum speed to collect only the supernatant.
  • the lysate was homogenized with a syringe equipped with a 20 G force terran needle and then treated with Q IAshredder. (That is, a normal cell lysate was applied to a Kyaschlets unit using a Pittman. This was centrifuged at 1500 i "pm for 2 minutes, and the effluent was collected.) 3501 70% ethanol was added.
  • the column was placed in a new 1.5 ml tube, DEPC-treated water was applied, the lid was capped, and the tube was allowed to stand for 10 minutes, and centrifuged at 11,500 rpm for 10 minutes to obtain total RNA. If the volume was low, re-attach the column to a new 1.5 ml tube, apply DEPC-treated water30, cap the lid for 10 minutes, and centrifuge at 11500 rpm for 10 minutes.
  • Example 5 DNase treatment of total RNA DNase treatment was performed to remove DNA from total RNA prepared from T cells. The reaction was performed with 2 units of DNase (Futtsubon Gene) and 50 units of RNase inhibitor
  • DD fluorescent differential display
  • the PCR reaction conditions were as follows: 1 minute at 95 minutes, 3 minutes at 40, 5 minutes at 725, and 7 minutes at 72 minutes after 30 cycles of ⁇ 95 seconds, 2 minutes at 40, and 1 minute at 72 ''. For 5 minutes, and then continuously at 4.
  • the primer pair used was an arbitrary primer for each of the primers GT15A (SEQ ID NO: 2), GT15C (SEQ ID NO: 3), and GT15G (SEQ ID NO: 4), AG 1 to 110 and AG 111, respectively. 199 and AG 200-287 were combined for a total of 287 reactions.
  • an oligomer composed of 10 nucleotides having a GC content of 50% was designed, synthesized, and used.
  • a 6% denaturing polyacrylamide gel was prepared, 2.5 1 samples were applied, and electrophoresed at 40 W for 210 minutes. Thereafter, the gel plate was scanned using Hitachi Fluorescence Image Analyzer -FMBI0 II, and electrophoretic images were obtained by fluorescence detection.
  • Two DD analyzes were performed using a number of arbitrary primers. Bands that differed before and after pollen dispersal or between the patient and healthy groups were selected and reproducible bands were excised from the gel in two experiments.
  • the gel containing the "441" band was cut out, stored in a TE solution, and heated at 60 for 10 minutes to elute DNA from the gel.
  • PCR was performed under the same conditions as DD-PCR, and a DNA fragment of about 250 bp was amplified.
  • GT15A was used as an anchor primer, and AG54 was used as an optional primer.
  • the amplified DNA fragment was cloned into a plasmid vector pCR2.1 (Invitrogen) to obtain a plasmid p44-13 carrying a DNA fragment of about 250 bp.
  • the nucleotide sequence of the DNA fragment was determined according to a conventional method.
  • SEQ ID NO: 1 shows the nucleotide sequence of "441"
  • the expression level of "441" was quantified by the TaqMan method using ABI-PRISM7700. This method uses a fluorescent dye to quantitatively detect the PCR-amplified DNA strand in real time. For the purpose of quantification, in the spring of 1998, a new blood sample before and after the cedar pollen was dispersed was collected from 22 volunteers, T cells were prepared, and total RNA was extracted. The expression level of the target gene was quantified using a total of 44 total RNA samples.
  • primer 44 ⁇ C TTCTCTATGGACCAATTCAACTTTGZ SEQ ID NO: 6
  • 441r AAGGGCCATTTTTTTACCATAATCAA / SEQ ID NO: 7
  • TaqMan probe P441 TCTGGATAATTAGTAGGATTTAAGCTG TGTTACAAGGCAZ SEQ ID NO: 8
  • FAM 6-carboxyfluorescein
  • TAMRA 6-carboxy-tetramethytri rhodamine
  • cDNA transcribed reversely from 44 kinds of total RNA using poly T (12 to 18) as a primer was used.
  • a serial dilution of plasmid p44-13 obtained in Example 7 was used as a type III reaction.
  • Table 3 shows the composition of the reaction mixture for monitoring PCR amplification.
  • the same quantitative analysis was performed on the ⁇ -actin (3-actin) gene, and correction was performed based on the copy number of those genes to obtain a copy of the target gene (441). The number was calculated.
  • Reaction composition of ABI-PRISM 7700 (reaction volume per 1 ⁇ ) Sterile distilled water 25.66 (ill)
  • Table 4 shows the number (copy number) of “441” in each sample corrected for the copy number of 3) -actin. For the correction, the average copy of 3-actin in all samples was determined, and the copy number of "441" in each sample was divided by the relative value of -actin in each sample when it was set to 1.
  • a novel gene that can be used as an indicator of the response of T cells to antigen stimulation such as cedar pollen was provided.

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Abstract

On a réussi à isoler un nouveau gène se caractérisant par une expression notablement réduite après la dispersion des pollens, en préparant des lymphocytes T prélevés sur des sujets avant et après la saison de dispersion des pollens et en recherchant ledit gène par la technique dite du DD (differential display). On a découvert que ce gène est utilisable dans l'examen des affections allergiques et dans le criblage de composés candidats pour des médicaments capables de réguler la réponse des lymphocytes T au stimulus d'un antigène.
PCT/JP2000/003190 1999-05-27 2000-05-18 Gene 441 associe a la pollinose Ceased WO2000073435A1 (fr)

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Cited By (1)

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JPH11332567A (ja) * 1998-05-22 1999-12-07 Dai Ichi Seiyaku Co Ltd アトピー体質の判定方法
WO2000020575A1 (fr) * 1998-10-06 2000-04-13 Genox Research, Inc. Gene associe a la pollinose

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* Cited by examiner, † Cited by third party
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JPH11332567A (ja) * 1998-05-22 1999-12-07 Dai Ichi Seiyaku Co Ltd アトピー体質の判定方法
WO2000020575A1 (fr) * 1998-10-06 2000-04-13 Genox Research, Inc. Gene associe a la pollinose

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JP2003322610A (ja) * 2002-04-30 2003-11-14 Omron Corp 光学装置、被測定体載置部品、光出射位置制御装置、分析システム、本人照合方法及びアレルギー・副作用検査方法

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