WO2019000148A1 - Arnsi du gène abcb6 humain et utilisation correspondante - Google Patents
Arnsi du gène abcb6 humain et utilisation correspondante Download PDFInfo
- Publication number
- WO2019000148A1 WO2019000148A1 PCT/CN2017/089928 CN2017089928W WO2019000148A1 WO 2019000148 A1 WO2019000148 A1 WO 2019000148A1 CN 2017089928 W CN2017089928 W CN 2017089928W WO 2019000148 A1 WO2019000148 A1 WO 2019000148A1
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- WO
- WIPO (PCT)
- Prior art keywords
- sirna
- abcb6 gene
- gene
- abcb6
- sequence
- Prior art date
Links
- 108020004459 Small interfering RNA Proteins 0.000 title description 11
- 101100000375 Homo sapiens ABCB6 gene Proteins 0.000 title description 6
- 101150073228 ABCB6 gene Proteins 0.000 claims abstract description 13
- 230000014509 gene expression Effects 0.000 claims abstract description 13
- 238000012228 RNA interference-mediated gene silencing Methods 0.000 claims abstract description 7
- 230000009368 gene silencing by RNA Effects 0.000 claims abstract description 7
- 239000003814 drug Substances 0.000 claims abstract description 6
- 230000002159 abnormal effect Effects 0.000 claims abstract description 4
- 201000010099 disease Diseases 0.000 claims abstract description 4
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims abstract description 4
- 239000012634 fragment Substances 0.000 claims abstract 4
- 230000000692 anti-sense effect Effects 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 3
- 108020004999 messenger RNA Proteins 0.000 abstract description 5
- 229940079593 drug Drugs 0.000 abstract description 4
- 230000002401 inhibitory effect Effects 0.000 abstract description 2
- 239000004055 small Interfering RNA Substances 0.000 description 10
- 238000003753 real-time PCR Methods 0.000 description 6
- 238000001514 detection method Methods 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- 238000001890 transfection Methods 0.000 description 4
- 102000043966 ABC-type transporter activity proteins Human genes 0.000 description 3
- 108010006533 ATP-Binding Cassette Transporters Proteins 0.000 description 3
- 102100021503 ATP-binding cassette sub-family B member 6 Human genes 0.000 description 3
- 101000677883 Homo sapiens ATP-binding cassette sub-family B member 6 Proteins 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 2
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 2
- 108010078791 Carrier Proteins Proteins 0.000 description 2
- 238000002123 RNA extraction Methods 0.000 description 2
- 239000002299 complementary DNA Substances 0.000 description 2
- 238000010839 reverse transcription Methods 0.000 description 2
- 102000041104 ABCB family Human genes 0.000 description 1
- 108091060863 ABCB family Proteins 0.000 description 1
- ZKHQWZAMYRWXGA-KQYNXXCUSA-J ATP(4-) Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](COP([O-])(=O)OP([O-])(=O)OP([O-])([O-])=O)[C@@H](O)[C@H]1O ZKHQWZAMYRWXGA-KQYNXXCUSA-J 0.000 description 1
- ZKHQWZAMYRWXGA-UHFFFAOYSA-N Adenosine triphosphate Natural products C1=NC=2C(N)=NC=NC=2N1C1OC(COP(O)(=O)OP(O)(=O)OP(O)(O)=O)C(O)C1O ZKHQWZAMYRWXGA-UHFFFAOYSA-N 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
- 102100031181 Glyceraldehyde-3-phosphate dehydrogenase Human genes 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 108091034117 Oligonucleotide Proteins 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 206010036790 Productive cough Diseases 0.000 description 1
- 238000011530 RNeasy Mini Kit Methods 0.000 description 1
- 108091081021 Sense strand Proteins 0.000 description 1
- 108091081024 Start codon Proteins 0.000 description 1
- 241000251539 Vertebrata <Metazoa> Species 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000000903 blocking effect Effects 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 239000012091 fetal bovine serum Substances 0.000 description 1
- 108020004445 glyceraldehyde-3-phosphate dehydrogenase Proteins 0.000 description 1
- 230000003301 hydrolyzing effect Effects 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 229910021645 metal ion Inorganic materials 0.000 description 1
- 230000036457 multidrug resistance Effects 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 230000008506 pathogenesis Effects 0.000 description 1
- 239000013600 plasmid vector Substances 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 210000003802 sputum Anatomy 0.000 description 1
- 208000024794 sputum Diseases 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 235000000346 sugar Nutrition 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- 230000009261 transgenic effect Effects 0.000 description 1
- 102000035160 transmembrane proteins Human genes 0.000 description 1
- 108091005703 transmembrane proteins Proteins 0.000 description 1
- 230000032895 transmembrane transport Effects 0.000 description 1
- 230000032258 transport Effects 0.000 description 1
Classifications
-
- 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
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- 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
Definitions
- the present invention belongs to the field of molecular genetics, and particularly relates to an siRNA capable of inhibiting the expression of a human ABCB6 gene and an application thereof.
- Adenosine triphosphate binding cassette transporter is a large class of transmembrane proteins that utilize the energy of hydrolyzing A TP to transport transmembrane transport of various endogenous and exogenous biomolecules in solute. Substrates include: sugars, amino acids, metal ions, peptides, proteins, cellular metabolites, and drugs. ABCB transporters are widely present in eukaryotic and prokaryotic organisms. To date, 49 ABC transgenic superfamily members have been identified in the human genome, which are divided into seven subfamilies of A-G.
- ABCB1 is the first human ABC transporter to be discovered. Because multiple members are associated with multidrug resistance (MD R), the ABCB subfamily is also known as the ABC transporter MDR family. As one of them, A BCB6 is not only related to MDR, but also related to the pathogenesis of various tumors. However, the function and role of ABCB6 in it have not been clarified so far, and further research is needed.
- RNA interference RNA interference
- RNAi small interfering RNA
- siABCB6 sequence is as follows: [0007] Justice Chain: 5'- GGCAUCUGGAUCAAGUUCA -3' (SEQ ID NO: 1)
- Antisense strand 5,- UGAACUUGAUCCAGAUGCC -3, (SEQ ID NO: 2).
- the present invention provides siABCB6 which has the advantages of high interference efficiency, high-efficiency and specific inhibition of ABCB6 gene expression, and can be used as a powerful tool for the preparation of a medicament for treating a disease associated with abnormal expression of ABCB6 gene.
- 1 is a schematic diagram showing the results of quantitative PCR detection of ABCB6 gene expression levels after transfection of A375 cells with siABCB6.
- A375 cells purchased from Biovector Plasmid Vector Culture Cell Gene Collection
- DMEM complete medium containing 10% fetal bovine serum
- 6-well plates were plated at a ratio of 150,000 cells/well at 37°C. C, 5 % C02 was cultured for 18 h.
- Cell transfection was performed using the Lipofectamine 3000 Transfection Kit (Invitrogen) according to the product instructions.
- RNA extraction Total RNA from normal and transfected siABCB6 ABCB6 cells was extracted using the QIAGEN RNeasy Mini Kit.
- Reverse Transcription Reverse transcription was performed using FastQuant RT Super Mix.
- Quantitative PCR was carried out, and the reaction system was 20 ⁇ , and 1 L cDNA was added as a template for each reaction.
- the reaction procedure is: (1) 95 °C 30 s, (2) 95 °C 5s, (3) 60 °C 30s, (2)-(3), 40 cycles. With GAPDH as the internal reference, the results are shown in Figure 1.
- the quantitative PCR primers used are shown in Table 1:
- the mRNA expression level of ABCB6 gene in A375 cells transfected with siABCB6 was significantly decreased compared with normal A375 cells, indicating that siABCB6 of the present invention can efficiently and specifically inhibit the expression of ABCB6 gene.
- the siABCB6 provided by the invention has high interference efficiency and can efficiently and specifically inhibit the ABCB6 gene expression.
- the advantages of the drug can be used as a powerful tool for the preparation of drugs for the treatment of diseases related to abnormal expression of ABCB6 gene.
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Genetics & Genomics (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Biomedical Technology (AREA)
- General Health & Medical Sciences (AREA)
- Molecular Biology (AREA)
- Organic Chemistry (AREA)
- Biotechnology (AREA)
- Public Health (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Medicinal Chemistry (AREA)
- Veterinary Medicine (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- Zoology (AREA)
- General Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Physics & Mathematics (AREA)
- Microbiology (AREA)
- Plant Pathology (AREA)
- Biophysics (AREA)
- Biochemistry (AREA)
- Epidemiology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
L'invention concerne un fragment d'ARN interférant qui inhibe efficacement et spécifiquement le niveau d'expression d'ARNm du gène ABCB6, ledit fragment pouvant être utilisé pour préparer un médicament pour traiter une maladie associée à l'expression anormale du gène ABCB6.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| PCT/CN2017/089928 WO2019000148A1 (fr) | 2017-06-26 | 2017-06-26 | Arnsi du gène abcb6 humain et utilisation correspondante |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| PCT/CN2017/089928 WO2019000148A1 (fr) | 2017-06-26 | 2017-06-26 | Arnsi du gène abcb6 humain et utilisation correspondante |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| WO2019000148A1 true WO2019000148A1 (fr) | 2019-01-03 |
Family
ID=64742784
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| PCT/CN2017/089928 WO2019000148A1 (fr) | 2017-06-26 | 2017-06-26 | Arnsi du gène abcb6 humain et utilisation correspondante |
Country Status (1)
| Country | Link |
|---|---|
| WO (1) | WO2019000148A1 (fr) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116064527A (zh) * | 2022-08-26 | 2023-05-05 | 广东省第二人民医院（广东省卫生应急医院） | 抑制ABCB6基因表达的siRNA及其应用 |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103849623A (zh) * | 2012-11-28 | 2014-06-11 | 天津华大基因科技有限公司 | Abcb6基因突变体及其应用 |
-
2017
- 2017-06-26 WO PCT/CN2017/089928 patent/WO2019000148A1/fr active Application Filing
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103849623A (zh) * | 2012-11-28 | 2014-06-11 | 天津华大基因科技有限公司 | Abcb6基因突变体及其应用 |
Non-Patent Citations (2)
| Title |
|---|
| DATABASE Nucleotide [O] 28 March 2017 (2017-03-28), ALLIKMETS, R. ET AL., XP055558442, retrieved from ncbi Database accession no. NM_005689 * |
| ZHAO, SHIGUANG ET AL.: "Increased Expression of ABCB6 Enhances Protoporphyrin IX Accumulation and Photodynamic Effect in Human Glioma", ANNALS OF SURGICAL ONCOLOGY, vol. 20, no. 13, 12 June 2012 (2012-06-12), pages 4379 - 4388, XP055558440, ISSN: 1068-9265 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116064527A (zh) * | 2022-08-26 | 2023-05-05 | 广东省第二人民医院（广东省卫生应急医院） | 抑制ABCB6基因表达的siRNA及其应用 |
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