WO2002006297A1 - Nouveaux derives d'oligonucleotides antisens contre le gene de la tumeur de wilms - Google Patents

Nouveaux derives d'oligonucleotides antisens contre le gene de la tumeur de wilms Download PDF

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Publication number
WO2002006297A1
WO2002006297A1 PCT/JP2001/006049 JP0106049W WO0206297A1 WO 2002006297 A1 WO2002006297 A1 WO 2002006297A1 JP 0106049 W JP0106049 W JP 0106049W WO 0206297 A1 WO0206297 A1 WO 0206297A1
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Prior art keywords
oligonucleotide derivative
antisense
oligonucleotide
antisense oligonucleotide
present
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PCT/JP2001/006049
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English (en)
Japanese (ja)
Inventor
Takeshi Imanishi
Haruo Sugiyama
Yoshiyuki Ohsugi
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Chugai Seiyaku Kabushiki Kaisha
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Priority to AU2001271042A priority Critical patent/AU2001271042A1/en
Publication of WO2002006297A1 publication Critical patent/WO2002006297A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/32—Chemical structure of the sugar
    • C12N2310/323—Chemical structure of the sugar modified ring structure
    • C12N2310/3231—Chemical structure of the sugar modified ring structure having an additional ring, e.g. LNA, ENA

Definitions

  • the present invention relates to a novel antisense to the Wilms oncogene (WT 1). More specifically, the present invention relates to a WT 1 comprising an oligonucleotide derivative containing at least one nucleotide analog unit having a sugar moiety modified. A new antisense to the Wilms oncogene (WT 1).
  • Antisense oligonucleotides are one of the most promising fields in recent years, as they specifically regulate the expression of unwanted genes.
  • the antisense method is based on the concept of controlling the flow of so-called central dharma using antisense oligonucleotides, namely DNA-RNA-protein.
  • nucleic acid derivatives and analogs have been synthesized and studied. For example, phosphorothioate in which an oxygen atom on a phosphorus atom is substituted with an iodo atom, methylphosphonate in which a methyl group is substituted, or more recently one in which a phosphorus atom is also substituted with a carbon atom, and even a sugar moiety Structures that have converted the structure of nucleosides or modified nucleic acid bases have also been synthesized.
  • Japanese Patent Application Laid-Open No. 10-304889 discloses a nucleotide analog unit in which the sugar moiety of an oligonucleotide constituting antisense is modified.
  • This nucleotide analog unit has a structure represented by the following formula 1 in which a sugar moiety conformation is immobilized to an N-type.
  • an oligonucleotide derivative incorporating one or more nucleotide analog units having a sugar moiety having the above structure is synthesized, and the basic physical properties of the oligonucleotide derivative are measured extracellularly. That is, the duplex forming ability is studied by measuring the melting temperature of the oligonucleotide derivative and the annealing treatment of the sense strand composed of natural DNA or RNA. The resistance of the oligonucleotide derivative to the nuclease enzyme was measured in vitro.
  • this publication describes whether the oligonucleotide derivative exhibits nuclease resistance and acts stably as an antisense even in cells, as in the case of the extracellular experimental results. However, it does not disclose whether or not it can form a double-stranded or triple-stranded form by binding to a naturally-occurring gene, particularly in a cell, to actually inhibit the expression of a specific gene.
  • Wilms tumor is a pediatric kidney tumor caused by inactivation of both Wilms tumor gene (WT1) alleles located on chromosome 11p13 (Cal KM et al., Cell 60: 509, 1990).
  • WT1 Wilms tumor gene
  • Non-coding upstream sequence of WT1 CE Camphell et al., Oncogene 9: 583-595, 1994
  • DA Haber et al., Proc. Natl. Acad. Sc. USA, 88: 9618-9622 (1991) have already been reported and are expected to be involved in the growth and differentiation of tumors and the like (DA Haber et al., Supra).
  • WT1 is strongly expressed in some tumor cells such as human leukocyte-derived K562 cells and human gastric cancer-derived Az521 cells, and when its expression is suppressed, cells become apoptotic. It has been reported to cause cis (Kentaro Senba, Cell Engineering, 14, 5 4 6- 5 5 3 (1995).
  • Japanese Patent Application Laid-Open No. 9-104629 discloses an antisense oligonucleotide derivative against WT1.
  • This oligonucleotide derivative includes not only a natural oligonucleotide that has not been modified but also a modified oligonucleotide derivative.
  • the modified oligonucleotide derivative is obtained by modifying the phosphodiester portion of a nucleotide, for example, a modified lower alkylphosphonate such as a methylphosphonate type or an ethylphosphonate type, a modified phospholipid thioate or a modified phosphothioate. Modified oral amidates are mentioned.
  • an oligonucleotide derivative containing a nucleotide analog unit having a modified sugar moiety is not disclosed.
  • the antisense oligonucleotide derivative disclosed in this publication has various properties required for antisense, that is, whether or not it has nuclease resistance in cells as well as in the results of extracellular experiments. In addition, it does not mention whether it can bind to a naturally-occurring gene, particularly in vivo, to form a double-stranded or triple-stranded chain, and actually inhibit the expression of a specific gene.
  • the inventors of the present invention designed a nucleic acid analog having a sugar moiety conformation immobilized on a nucleic acid, which is considered to be useful in the antisense method, and synthesized a nucleotide analog unit having a unit structure thereof. It was confirmed that the oligonucleotide derivative prepared using this was extremely useful as a WT1 antisense molecule. The details of the present invention will be described below.
  • FIG. 1 is a diagram showing a nucleotide sequence of an oligonucleotide used as a sample in Experimental Example 1 and a band pattern obtained by subjecting a sample treated with nuclease (snake venom phosphodiesterase) to polyacrylamide gel electrophoresis.
  • Figure 2 shows the nucleotide sequence of the oligonucleotide used as a sample in Experimental Example 1 and the band pattern obtained by subjecting a sample treated with nuclease (fetal bovine serum: FBS) to polyacrylamide gel electrophoresis. It is.
  • nuclease fetal bovine serum
  • FIG. 3 is a graph showing the antisense effect of the natural oligonucleotide.
  • FIG. 4 is a graph showing the concentration dependence of the antisense effect of the oligonucleotide derivative of the present invention.
  • FIG. 5 is a graph showing the antisense effect of comparing the oligonucleotide derivative of the present invention with a phosphorothioate-type oligonucleotide derivative.
  • FIG. 6 is a graph showing the concentration-dependent antisense effect of the oligonucleotide derivative of the present invention and the scrambled sequence oligonucleotide.
  • FIG. 7 is a graph showing the antisense effect of the oligonucleotide derivative of the present invention with two different scrambled sequence oligonucleotides.
  • the oligonucleotide or polynucleotide derivative of the present invention has the following general formula
  • B is a pyrimidine nucleobase or a purine nucleobase or an analog thereof
  • B has one or more nucleotide analog units having a structure represented by the following formula:
  • the oligonucleotide or polynucleotide derivative of the present invention has the general formula
  • BB 2 is the same or different, is a pyrimidine nucleobase or purine nucleobase or an analog thereof, R is hydrogen, a hydroxyl group, a halogen, or an alkoxy group, and WW 2 is the same or different, hydrogen, Alkyl group, alkenyl group, alkynyl group, cycloalkyl group, aralkyl group, aryl group, acyl group, silyl group or natural nucleotide via phosphate residue or phosphate diester bond, synthetic nucleotide or these nucleotides O Rigo nucleotide if Ku is a polynucleotide, n 1 or n 2 are the same or different, 0-5 is a integer of 0 (where, n 1 or n 2 do not become zero simultaneously.
  • N 3 is an integer from 1 to 50, provided that n 1 and Z or n 2 are B 1 and B in the case of 2 or more may not be the same, R is also an oligonucleotide or polynucleotide derived material represented by may not be the same.
  • the number of bases in the antisense oligonucleotide derivative of the present invention is not particularly limited, but is usually 5 to 50, preferably 9 to 30.
  • the pyrimidine nucleobase or purine nucleobase or an analog thereof is thymine, peracyl, cytosine, adenine, guanine and an analog thereof.
  • These analogs include purine nucleic acid salt analogs and pyrimidine nucleobase analogs.
  • the purine nucleobase analog can be preferably selected from the following compounds.
  • Guanosine diphosphate 8-oxo-adenosine, 8-oxo-guanosine, 8-fluoro-adenosine, 8-fluoro-guanosine, 8-methoxy- Adenosine, 8-methoxy-guanosine, 8-azaadenosine, 8-azaguanosine, azacytin, fludarabine phosphate, 6-MP, 6-TG, azasammlungne, aloprinol, acyclovir, ganciclovia, deoxyphormycin, arabinosilagenin ( ara-A), guanosine diphosphate tofucoose, guanosine diphosphate- 12-fluorofurose, guanosine diphosphate toe j3 L-2-aminofucose, guanosine diphosphate-D-arabinose and 2-aminoadesine And so on.
  • the pyridine nucleobase analog can be preferably selected from the following compounds. That is,
  • 5-fluoroperacil 5-chlorouracil, 5-bromouracil, dihydrouracil, 5-methylcytosine, 5-propierthymine, 5-propynylperacyl, 5-propiercytosine, 5-fluorocytosine, floxyperidine, peridine, thymine , 3'-azidodeoxythymidine, 2-fluorodeoxycytidine, 3_fluoro--3, -deoxythymidine, 3, -dideoxycytidine- 2, -ene, 3, -doxy-3, dedeo Xitymidine-2, -ene, and cytosine arabinose.
  • nucleotide analog unit which can be used as an antisense in the present invention
  • oligonucleotide derivative containing the nucleotide analog unit are described in detail in the above-mentioned Japanese Patent Application Laid-Open No. 10-304889. I have. This description shall be included in this document.
  • the antisense oligonucleotide derivative to WT1 of the present invention may be any oligo or polynucleotide that can bind to WT1 and inhibit or suppress the expression of its gene, It suffices that one or more of the nucleotides is replaced with the nucleotide analog unit of the present invention.
  • antisense oligonucleotide derivatives include those for the transcriptional capping site of WT1, those for the translation initiation region, those for exons, those for introns, and the like. More specifically, the following bases Contains an array.
  • Antisense sequence of transcription capping site AGGGTCGAAT GCGGTGGG (SEQ ID NO: 1)
  • Antisense sequence of transcription capping region (AS 2) TCAAATAAGA GGGGCCGG (SEQ ID NO: 2)
  • Antisense sequence of translation initiation region (AS 3) GTCGGAGCCC ATTTGCTG
  • the vicinity of the translation initiation region of WT1 is a region where an antisense effect is observed, and is expected as a target for cancer treatment by antisense.
  • the present inventors have synthesized the oligonucleotide derivative of the present invention having an antisense sequence in this region, and administered it to Az521 cells that strongly express WT1, using the decrease in cell number due to apoptosis as an index. The antisense effect was examined and an excellent effect was confirmed.
  • the antisense of the present invention can be used as an antitumor drug by introducing into tumor cells expressing WT1, thereby inhibiting or suppressing the expression to induce the cells to undergo apoptosis.
  • oligonucleotide derivative of the present invention are represented by the following formula:
  • 5′-GUCGGAGCCCAUUUGCUG-3 ′ (SEQ ID NO: 5) A sequence in which one or two or more bases of the antisense sequence to the translation initiation region of the WT1 translation initiation region are replaced with the nucleotide analog unit of the present invention and a sequence thereof.
  • Antisense oligonucleotide derivatives containing the same, and oligonucleotide derivatives that hybridize under stringent conditions to DNA or RNA, which is a sequence complementary to the above antisense sequence, can be mentioned. Hybridization is a well-known technique (Sambrook, l et al., Molecular Cloning 2nd ed., Cold spring Harbor Lab. Press, 1989, etc.).
  • Stringent end conditions in the present invention may be appropriately selected by those skilled in the art. Is possible, for example, under low stringent conditions.
  • the low stringent conditions are, for example, in washing after hybridization, at 42 ° C, 0.1 XSSC, 0.1% SDS, preferably 50 ° (: 0.1 XS SC, 0.1% SDS More preferred stringent conditions include high stringency conditions High stringency end conditions are 65 ° C, 6 XSSC, 0.1% SDS.
  • the oligonucleotide derivative of the present invention exhibits superior nuclease resistance as compared to a natural oligo nucleotide having the same base sequence. Therefore, it is stable even when administered intracellularly, and has the advantage of maintaining its excellent antisense effect.
  • the antisense oligonucleotide derivative of the present invention requires the use of a nucleotide analog unit in which only the sugar moiety is modified, but further modifies the other moiety, for example, the phosphodiester moiety as in the case of phosphorothioate. Nucleotide analog units may be used. Further, an oligonucleotide derivative antisense can also be prepared by combining the nucleotide analog unit having a modified sugar moiety of the present invention with a nucleotide analog unit which is another known antisense structural unit. The antisense oligonucleotide derivative of the present invention can be used as an external preparation such as a liniment or a poultice by mixing with a suitable base material which is inactive against the derivative.
  • excipients can be prepared according to a conventional method.
  • the antisense oligonucleotide derivative of the present invention is applied directly to the affected area of the patient, or is adapted to the patient so that it can reach the affected area as a result of intravenous administration or the like.
  • an antisense-encapsulated material that enhances durability and membrane permeability can be used.
  • ribosome, poly-L-lysine, lipid, cholesterol, lipofectin, or an analog thereof can be mentioned.
  • the dose of the antisense oligonucleotide derivative of the present invention depends on the patient's condition, A suitable amount can be used after appropriately adjusting according to age, sex, body weight, and the like.
  • various administration methods such as oral administration, intramuscular administration, intraperitoneal administration, intradermal administration, subcutaneous administration, intravenous administration, intravenous administration, rectal administration, etc.
  • a suitable method can be used as appropriate from the administration method.
  • Example 1 Nuclease resistance of oligonucleotide derivatives containing nucleotide analog units of the present invention
  • Oligonucleotide derivatives containing five nucleotide analog units of the present invention were prepared and used as samples for the nuclease resistance test. On the other hand, a natural oligonucleotide having the same sequence was prepared and used as a control.
  • the nuclease of the oligonucleotide derivative of the present invention is obtained by using a snake venom phosphodiesterase as a nuclease, treating the oligonucleotide labeled with radioisotope with 32 P with nuclease, and separating the oligonucleotide by polyacrylamide gel electrophoresis. Monozyme resistance was evaluated.
  • Figure 1 shows the sequence of the oligonucleotide prepared as a sample and the results of electrophoresis after nuclease treatment.
  • the ratio of the undegraded oligonucleotide was less than 30% for the natural oligonucleotide.
  • the undegraded oligonucleotide completely disappeared.
  • the oligonucleotide derivative of the present invention about 90% of undegraded oligonucleotide was confirmed even 8 hours after the start of the nuclease treatment. From these results, it was found that the oligonucleotide derivative of the present invention has at least 100 times or more the nuclease resistance performance as compared with the natural oligonucleotide.
  • the oligonucleotide derivatives of the present invention showed extremely strong resistance to degradation by nuclease.
  • the following oligonucleotide derivative to WT1 synthesized by the method described in the example of JP-A-10-304889 was used as an antisense strand, and was subjected to annealing treatment with a sense strand consisting of natural DNA or RNA.
  • Tm value melting temperature
  • a sample solution (5001) with final concentrations of 100 mM NaC, 10 mM sodium phosphate buffer (pH 7.2), 4 M for antisense strand and 4 M for sense strand was bathed in boiling water. And slowly cooled to room temperature over 10 hours.
  • a nitrogen stream was passed through the cell chamber of the spectrophotometer (Bec kman DU650) to prevent dew condensation, the sample solution was gradually cooled to 5 ° C, and kept at 5 ° C for another 20 minutes before measuring. Started.
  • the sample temperature was increased by 0.2 ° C per minute to 90 ° C, and ultraviolet absorption at 260 nm was measured at 0.1 ° C intervals.
  • Tm value Melting temperature (ATm / mod.) /.
  • Antisense molecules 1, 2, 3, and 4 correspond to SEQ ID NOs: 3, 6, 7, and 1, respectively.
  • the oligomer (Test 2 (referred to as 5bc_AS) and Test 3) in which five nucleotide analog units (underlined) of the present invention were inserted into the natural DNA chain was found to be complementary to the complementary DNA oligomer.
  • Hypride forming ability was increased 7-8 degrees over the native chain (Test 1) as assessed by Tm values.
  • the Tm value of the oligomer of the present invention was found to be 5 to 9 degrees higher than that of the natural chain (D-origo).
  • D-origo no analogs whose Tm value is so markedly higher than that of the natural chain have been known.
  • the oligonucleotide derivative antisense of the present invention showed an excellent double strand-forming ability to both DNA and RNA.
  • Example 3 Antisense effect of oligonucleotide derivative on WT1
  • the antisense effect of oligonucleotide derivative of the present invention on WT1 was confirmed using cultured cells.
  • oligonucleotide derivative having an antisense sequence near the translation initiation region of WT1 was prepared in the same manner as in Example 2.
  • the oligonucleotide derivative of the present invention having the antisense sequence was administered to Az521 cells, and the antisense effect was evaluated using the decrease in the number of cells as an index.
  • experiments were performed under the same conditions for natural (D-origo) and phosphorothioate-type oligonucleotides (S-oligo).
  • oligonucleotide solution in the absence of serum (1.25 / 1) was added to 21 (final concentration 25 fig / m 1), and cultured for 2 hours. Then, 10 il of FBS (final concentration 10%) was added, followed by further culturing. Oligonucleotide solution 11 (final concentration + 12.5 g / ml) was added on the second, third, and fourth days, respectively, and the cells were further cultured for 2 days. (In experiments where the oligonucleotide concentration was varied, an appropriate volume of the oligonucleotide solution was used.) On day 6, the number of living cells was counted using a microscope.
  • the antisense effect of the oligonucleotide derivative of the present invention was examined using a system in which the number of cells was not reduced by D-oligo in (1) above. At this time, the oligonucleotide derivative was also administered to PC 14 cells that did not express WT1, and changes in the number of cells were compared. Fig. 4 shows the results. In the case of Az521 cells, the number of cells was not reduced when the D-oligo sequence was administered, but the number of cells was decreased when the oligonucleotide derivative of the present invention was administered. .
  • the oligonucleotide derivative of the present invention exhibited an antisense effect, but the S-oligo and D-oligo did not show an antisense effect. From these results, it was confirmed that the oligonucleotide derivative of the present invention had a higher antisense effect than S-oligo widely known as antisense. This result shows a good correlation with the value obtained in the in vitro experiment (Tm value).
  • the oligonucleotide derivative antisense of the present invention was administered to Az521 cells in the same manner as in Example 3, and the antisense effect was evaluated using the decrease in the number of cells as an index. For comparison, an experiment was performed under the same conditions using an oligonucleotide derivative having a scrambled sequence represented by the following formula.
  • FIG. 5 shows the growth inhibitory effect of Az521 cells when the antisense oligonucleotide derivative (5bc-AS) of the present invention was administered at different concentrations. At the same time, the Sc-B sequence was tested under the same conditions.
  • the antisense oligonucleotide derivative of the present invention exhibited a remarkable cell growth inhibitory effect as compared with the two scrambled sequence oligonucleotides.
  • An antisense oligonucleotide derivative having the following base sequence in which peracyl (U) of the above antisense oligonucleotide derivatives (first and second) is substituted with thymine (T) is expected to have the same antisense effect as the above antisense.

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Abstract

L'invention concerne des dérivés d'oligonucléotides antisens contre le gène de la tumeur de Wilms (WT1), lesdits dérivés renfermant au moins une unité d'analogue de nucléotide modifiée au niveau du sucre représentée par la formule générale (2). Dans cette dernière, B représente une base d'acide nucléique de pyrimidine, une base d'acide nucléique de purine ou un analogue correspondant. Ces dérivés d'oligonucléotides antisens contre le gène de la tumeur de Wilms (WT1) peuvent se lier, à un degré d'affinité élevé, audit gène dans des cellules afin de réguler ou inhiber efficacement l'expression de ce gène et être très résistant aux nucléases.
PCT/JP2001/006049 2000-07-19 2001-07-12 Nouveaux derives d'oligonucleotides antisens contre le gene de la tumeur de wilms WO2002006297A1 (fr)

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AU2001271042A AU2001271042A1 (en) 2000-07-19 2001-07-12 Novel antisense oligonucleotide derivatives against wilms's tumor gene

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11234995B2 (en) 2016-01-07 2022-02-01 Osaka University α-synuclein expression inhibitor

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0841068A1 (fr) * 1995-06-01 1998-05-13 Kishimoto, Tadamitsu Inhibiteur de la croissance de cellules leucemiques contenant des derives oligonucleotidiques antisens agissant contre le gene de la tumeur de wilms (wt1)
EP1004319A1 (fr) * 1997-07-16 2000-05-31 Haruo Sugiyama Medicaments contre une tumeur solide, contenant des inhibiteurs de l'expression du gene (wt1) de la tumeur de wilms
EP1013661A1 (fr) * 1997-03-07 2000-06-28 Takeshi Imanishi Nouveaux analogues de bicyclonucleoside et d'oligonucleotide

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0841068A1 (fr) * 1995-06-01 1998-05-13 Kishimoto, Tadamitsu Inhibiteur de la croissance de cellules leucemiques contenant des derives oligonucleotidiques antisens agissant contre le gene de la tumeur de wilms (wt1)
EP1013661A1 (fr) * 1997-03-07 2000-06-28 Takeshi Imanishi Nouveaux analogues de bicyclonucleoside et d'oligonucleotide
EP1004319A1 (fr) * 1997-07-16 2000-05-31 Haruo Sugiyama Medicaments contre une tumeur solide, contenant des inhibiteurs de l'expression du gene (wt1) de la tumeur de wilms

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11234995B2 (en) 2016-01-07 2022-02-01 Osaka University α-synuclein expression inhibitor

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