WO2023101316A1 - Composition for preventing or treating steatosis, comprising axl - Google Patents
Composition for preventing or treating steatosis, comprising axl Download PDFInfo
- Publication number
- WO2023101316A1 WO2023101316A1 PCT/KR2022/018680 KR2022018680W WO2023101316A1 WO 2023101316 A1 WO2023101316 A1 WO 2023101316A1 KR 2022018680 W KR2022018680 W KR 2022018680W WO 2023101316 A1 WO2023101316 A1 WO 2023101316A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- axl
- protein
- composition
- gene
- metabolic diseases
- Prior art date
Links
- 239000000203 mixture Substances 0.000 title claims abstract description 20
- 231100000240 steatosis hepatitis Toxicity 0.000 title abstract description 18
- 230000007863 steatosis Effects 0.000 title abstract description 4
- 102100037236 Tyrosine-protein kinase receptor UFO Human genes 0.000 claims abstract description 68
- 238000000034 method Methods 0.000 claims abstract description 38
- 230000014509 gene expression Effects 0.000 claims abstract description 32
- 238000003745 diagnosis Methods 0.000 claims abstract description 8
- 108090000623 proteins and genes Proteins 0.000 claims description 45
- 208000030159 metabolic disease Diseases 0.000 claims description 40
- 235000009200 high fat diet Nutrition 0.000 claims description 37
- 102000004169 proteins and genes Human genes 0.000 claims description 26
- 206010019708 Hepatic steatosis Diseases 0.000 claims description 19
- 101150018445 Axl gene Proteins 0.000 claims description 18
- 210000004185 liver Anatomy 0.000 claims description 18
- 208000016097 disease of metabolism Diseases 0.000 claims description 17
- 239000008194 pharmaceutical composition Substances 0.000 claims description 17
- 108091033319 polynucleotide Proteins 0.000 claims description 13
- 102000040430 polynucleotide Human genes 0.000 claims description 13
- 239000002157 polynucleotide Substances 0.000 claims description 13
- 230000015572 biosynthetic process Effects 0.000 claims description 12
- 238000003556 assay Methods 0.000 claims description 11
- 208000008338 non-alcoholic fatty liver disease Diseases 0.000 claims description 11
- 238000006243 chemical reaction Methods 0.000 claims description 9
- 238000010839 reverse transcription Methods 0.000 claims description 9
- 239000000523 sample Substances 0.000 claims description 9
- 239000013598 vector Substances 0.000 claims description 9
- 238000003127 radioimmunoassay Methods 0.000 claims description 8
- 239000000126 substance Substances 0.000 claims description 8
- 201000010099 disease Diseases 0.000 claims description 7
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 7
- 238000002965 ELISA Methods 0.000 claims description 6
- 239000012472 biological sample Substances 0.000 claims description 6
- 230000002860 competitive effect Effects 0.000 claims description 6
- 210000004027 cell Anatomy 0.000 claims description 5
- 235000005911 diet Nutrition 0.000 claims description 5
- 230000037213 diet Effects 0.000 claims description 5
- 208000001072 type 2 diabetes mellitus Diseases 0.000 claims description 5
- 238000000636 Northern blotting Methods 0.000 claims description 4
- 208000008589 Obesity Diseases 0.000 claims description 4
- 102000006382 Ribonucleases Human genes 0.000 claims description 4
- 108010083644 Ribonucleases Proteins 0.000 claims description 4
- 230000000295 complement effect Effects 0.000 claims description 4
- 230000002401 inhibitory effect Effects 0.000 claims description 4
- 239000000463 material Substances 0.000 claims description 4
- 238000005259 measurement Methods 0.000 claims description 4
- 235000020824 obesity Nutrition 0.000 claims description 4
- 210000002966 serum Anatomy 0.000 claims description 4
- 210000001519 tissue Anatomy 0.000 claims description 4
- 238000001262 western blot Methods 0.000 claims description 4
- 238000000018 DNA microarray Methods 0.000 claims description 3
- 206010022489 Insulin Resistance Diseases 0.000 claims description 3
- 230000000951 immunodiffusion Effects 0.000 claims description 3
- 238000000760 immunoelectrophoresis Methods 0.000 claims description 3
- 238000001114 immunoprecipitation Methods 0.000 claims description 3
- 238000003753 real-time PCR Methods 0.000 claims description 3
- 241000701161 unidentified adenovirus Species 0.000 claims description 3
- 239000013603 viral vector Substances 0.000 claims description 3
- 241000702421 Dependoparvovirus Species 0.000 claims description 2
- 208000032928 Dyslipidaemia Diseases 0.000 claims description 2
- 241000713666 Lentivirus Species 0.000 claims description 2
- 208000017170 Lipid metabolism disease Diseases 0.000 claims description 2
- 241000700584 Simplexvirus Species 0.000 claims description 2
- 241000700605 Viruses Species 0.000 claims description 2
- 239000013602 bacteriophage vector Substances 0.000 claims description 2
- 210000000601 blood cell Anatomy 0.000 claims description 2
- 239000013068 control sample Substances 0.000 claims description 2
- 239000013601 cosmid vector Substances 0.000 claims description 2
- 238000000684 flow cytometry Methods 0.000 claims description 2
- 238000001943 fluorescence-activated cell sorting Methods 0.000 claims description 2
- 238000012744 immunostaining Methods 0.000 claims description 2
- 210000002381 plasma Anatomy 0.000 claims description 2
- 239000013600 plasmid vector Substances 0.000 claims description 2
- 230000002265 prevention Effects 0.000 claims description 2
- 210000003296 saliva Anatomy 0.000 claims description 2
- 241001430294 unidentified retrovirus Species 0.000 claims description 2
- 210000002700 urine Anatomy 0.000 claims description 2
- 238000003757 reverse transcription PCR Methods 0.000 claims 2
- 230000000694 effects Effects 0.000 abstract description 22
- 239000003814 drug Substances 0.000 abstract description 15
- 229940124597 therapeutic agent Drugs 0.000 abstract description 9
- 238000011161 development Methods 0.000 abstract description 3
- 239000003550 marker Substances 0.000 abstract description 3
- 230000001105 regulatory effect Effects 0.000 abstract description 2
- 241000699670 Mus sp. Species 0.000 description 22
- 208000010706 fatty liver disease Diseases 0.000 description 18
- 208000004930 Fatty Liver Diseases 0.000 description 14
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 12
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 11
- 239000008103 glucose Substances 0.000 description 10
- 102000004877 Insulin Human genes 0.000 description 6
- 108090001061 Insulin Proteins 0.000 description 6
- 230000037396 body weight Effects 0.000 description 6
- 229940079593 drug Drugs 0.000 description 6
- 229940125396 insulin Drugs 0.000 description 6
- 235000021590 normal diet Nutrition 0.000 description 6
- 150000007523 nucleic acids Chemical group 0.000 description 6
- 238000011740 C57BL/6 mouse Methods 0.000 description 5
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 5
- 239000004480 active ingredient Substances 0.000 description 5
- 210000000593 adipose tissue white Anatomy 0.000 description 5
- 150000001413 amino acids Chemical group 0.000 description 5
- 210000004369 blood Anatomy 0.000 description 5
- 239000008280 blood Substances 0.000 description 5
- 238000010586 diagram Methods 0.000 description 5
- 238000011813 knockout mouse model Methods 0.000 description 5
- 239000002773 nucleotide Substances 0.000 description 5
- 125000003729 nucleotide group Chemical group 0.000 description 5
- 238000007920 subcutaneous administration Methods 0.000 description 5
- 230000006870 function Effects 0.000 description 4
- 210000005228 liver tissue Anatomy 0.000 description 4
- 108020004999 messenger RNA Proteins 0.000 description 4
- 208000007082 Alcoholic Fatty Liver Diseases 0.000 description 3
- 108020004414 DNA Proteins 0.000 description 3
- 206010016262 Fatty liver alcoholic Diseases 0.000 description 3
- 208000026594 alcoholic fatty liver disease Diseases 0.000 description 3
- 230000029142 excretion Effects 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 230000037406 food intake Effects 0.000 description 3
- 235000012631 food intake Nutrition 0.000 description 3
- 235000021588 free fatty acids Nutrition 0.000 description 3
- 210000005229 liver cell Anatomy 0.000 description 3
- 230000007246 mechanism Effects 0.000 description 3
- 238000012216 screening Methods 0.000 description 3
- 238000003786 synthesis reaction Methods 0.000 description 3
- 239000013077 target material Substances 0.000 description 3
- 239000013076 target substance Substances 0.000 description 3
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 2
- 108020000948 Antisense Oligonucleotides Proteins 0.000 description 2
- 102000053602 DNA Human genes 0.000 description 2
- 239000003298 DNA probe Substances 0.000 description 2
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Chemical compound C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 description 2
- 241000124008 Mammalia Species 0.000 description 2
- 241000699666 Mus <mouse, genus> Species 0.000 description 2
- 108091028043 Nucleic acid sequence Proteins 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 230000009471 action Effects 0.000 description 2
- 238000007792 addition Methods 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 238000010171 animal model Methods 0.000 description 2
- 239000000074 antisense oligonucleotide Substances 0.000 description 2
- 238000012230 antisense oligonucleotides Methods 0.000 description 2
- 230000031018 biological processes and functions Effects 0.000 description 2
- 210000001772 blood platelet Anatomy 0.000 description 2
- 210000001638 cerebellum Anatomy 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 230000003436 cytoskeletal effect Effects 0.000 description 2
- 230000007423 decrease Effects 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 239000003937 drug carrier Substances 0.000 description 2
- 238000007446 glucose tolerance test Methods 0.000 description 2
- 210000003494 hepatocyte Anatomy 0.000 description 2
- 210000001320 hippocampus Anatomy 0.000 description 2
- 230000001771 impaired effect Effects 0.000 description 2
- 230000002779 inactivation Effects 0.000 description 2
- 230000031146 intracellular signal transduction Effects 0.000 description 2
- HQKMJHAJHXVSDF-UHFFFAOYSA-L magnesium stearate Chemical compound [Mg+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O HQKMJHAJHXVSDF-UHFFFAOYSA-L 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 210000001616 monocyte Anatomy 0.000 description 2
- -1 nucleoside triphosphates Chemical class 0.000 description 2
- 230000001766 physiological effect Effects 0.000 description 2
- 238000006116 polymerization reaction Methods 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 102000027426 receptor tyrosine kinases Human genes 0.000 description 2
- 108091008598 receptor tyrosine kinases Proteins 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 238000006467 substitution reaction Methods 0.000 description 2
- 208000024891 symptom Diseases 0.000 description 2
- LNAZSHAWQACDHT-XIYTZBAFSA-N (2r,3r,4s,5r,6s)-4,5-dimethoxy-2-(methoxymethyl)-3-[(2s,3r,4s,5r,6r)-3,4,5-trimethoxy-6-(methoxymethyl)oxan-2-yl]oxy-6-[(2r,3r,4s,5r,6r)-4,5,6-trimethoxy-2-(methoxymethyl)oxan-3-yl]oxyoxane Chemical compound CO[C@@H]1[C@@H](OC)[C@H](OC)[C@@H](COC)O[C@H]1O[C@H]1[C@H](OC)[C@@H](OC)[C@H](O[C@H]2[C@@H]([C@@H](OC)[C@H](OC)O[C@@H]2COC)OC)O[C@@H]1COC LNAZSHAWQACDHT-XIYTZBAFSA-N 0.000 description 1
- FHVDTGUDJYJELY-UHFFFAOYSA-N 6-{[2-carboxy-4,5-dihydroxy-6-(phosphanyloxy)oxan-3-yl]oxy}-4,5-dihydroxy-3-phosphanyloxane-2-carboxylic acid Chemical compound O1C(C(O)=O)C(P)C(O)C(O)C1OC1C(C(O)=O)OC(OP)C(O)C1O FHVDTGUDJYJELY-UHFFFAOYSA-N 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 108020005544 Antisense RNA Proteins 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- QCMYYKRYFNMIEC-UHFFFAOYSA-N COP(O)=O Chemical class COP(O)=O QCMYYKRYFNMIEC-UHFFFAOYSA-N 0.000 description 1
- 206010009208 Cirrhosis alcoholic Diseases 0.000 description 1
- 206010010071 Coma Diseases 0.000 description 1
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- FBPFZTCFMRRESA-JGWLITMVSA-N D-glucitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-JGWLITMVSA-N 0.000 description 1
- 230000006820 DNA synthesis Effects 0.000 description 1
- 102000016928 DNA-directed DNA polymerase Human genes 0.000 description 1
- 108010014303 DNA-directed DNA polymerase Proteins 0.000 description 1
- 241000283073 Equus caballus Species 0.000 description 1
- 241000282326 Felis catus Species 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 102100031181 Glyceraldehyde-3-phosphate dehydrogenase Human genes 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- 206010019728 Hepatitis alcoholic Diseases 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 101001043754 Homo sapiens Inhibitor of nuclear factor kappa-B kinase subunit beta Proteins 0.000 description 1
- 102100021854 Inhibitor of nuclear factor kappa-B kinase subunit beta Human genes 0.000 description 1
- 102100034343 Integrase Human genes 0.000 description 1
- 241000087799 Koma Species 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- 229920000168 Microcrystalline cellulose Polymers 0.000 description 1
- 108091005461 Nucleic proteins Proteins 0.000 description 1
- 108020005187 Oligonucleotide Probes Proteins 0.000 description 1
- 229930040373 Paraformaldehyde Natural products 0.000 description 1
- 241000009328 Perro Species 0.000 description 1
- 108020004518 RNA Probes Proteins 0.000 description 1
- 239000003391 RNA probe Substances 0.000 description 1
- 108010092799 RNA-directed DNA polymerase Proteins 0.000 description 1
- 108020004682 Single-Stranded DNA Proteins 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 238000000692 Student's t-test Methods 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- RYYWUUFWQRZTIU-UHFFFAOYSA-N Thiophosphoric acid Chemical class OP(O)(S)=O RYYWUUFWQRZTIU-UHFFFAOYSA-N 0.000 description 1
- 238000010162 Tukey test Methods 0.000 description 1
- 208000036142 Viral infection Diseases 0.000 description 1
- 239000000205 acacia gum Substances 0.000 description 1
- 235000010489 acacia gum Nutrition 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 210000001789 adipocyte Anatomy 0.000 description 1
- 208000002353 alcoholic hepatitis Diseases 0.000 description 1
- 208000010002 alcoholic liver cirrhosis Diseases 0.000 description 1
- 229940072056 alginate Drugs 0.000 description 1
- 235000010443 alginic acid Nutrition 0.000 description 1
- 229920000615 alginic acid Polymers 0.000 description 1
- 238000000540 analysis of variance Methods 0.000 description 1
- 230000000890 antigenic effect Effects 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 108010023337 axl receptor tyrosine kinase Proteins 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 229910000389 calcium phosphate Inorganic materials 0.000 description 1
- 229960001714 calcium phosphate Drugs 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 239000000378 calcium silicate Substances 0.000 description 1
- 229910052918 calcium silicate Inorganic materials 0.000 description 1
- 229960003340 calcium silicate Drugs 0.000 description 1
- 235000012241 calcium silicate Nutrition 0.000 description 1
- OYACROKNLOSFPA-UHFFFAOYSA-N calcium;dioxido(oxo)silane Chemical compound [Ca+2].[O-][Si]([O-])=O OYACROKNLOSFPA-UHFFFAOYSA-N 0.000 description 1
- 150000004657 carbamic acid derivatives Chemical class 0.000 description 1
- 230000005754 cellular signaling Effects 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 235000010980 cellulose Nutrition 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 230000007882 cirrhosis Effects 0.000 description 1
- 239000003184 complementary RNA Substances 0.000 description 1
- 239000000306 component Substances 0.000 description 1
- 238000012790 confirmation Methods 0.000 description 1
- 230000034994 death Effects 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000001934 delay Effects 0.000 description 1
- 238000012217 deletion Methods 0.000 description 1
- 230000037430 deletion Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 239000008121 dextrose Substances 0.000 description 1
- 238000004821 distillation Methods 0.000 description 1
- NAGJZTKCGNOGPW-UHFFFAOYSA-N dithiophosphoric acid Chemical class OP(O)(S)=S NAGJZTKCGNOGPW-UHFFFAOYSA-N 0.000 description 1
- 239000003995 emulsifying agent Substances 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- YQGOJNYOYNNSMM-UHFFFAOYSA-N eosin Chemical compound [Na+].OC(=O)C1=CC=CC=C1C1=C2C=C(Br)C(=O)C(Br)=C2OC2=C(Br)C(O)=C(Br)C=C21 YQGOJNYOYNNSMM-UHFFFAOYSA-N 0.000 description 1
- 230000001747 exhibiting effect Effects 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000000796 flavoring agent Substances 0.000 description 1
- 235000013355 food flavoring agent Nutrition 0.000 description 1
- 235000003599 food sweetener Nutrition 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 229940014259 gelatin Drugs 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- 108020004445 glyceraldehyde-3-phosphate dehydrogenase Proteins 0.000 description 1
- 238000007490 hematoxylin and eosin (H&E) staining Methods 0.000 description 1
- 238000009396 hybridization Methods 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 230000002452 interceptive effect Effects 0.000 description 1
- 238000007912 intraperitoneal administration Methods 0.000 description 1
- 238000000021 kinase assay Methods 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 230000003908 liver function Effects 0.000 description 1
- 239000000314 lubricant Substances 0.000 description 1
- 210000002751 lymph Anatomy 0.000 description 1
- 235000019359 magnesium stearate Nutrition 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 229920000609 methyl cellulose Polymers 0.000 description 1
- 230000011987 methylation Effects 0.000 description 1
- 238000007069 methylation reaction Methods 0.000 description 1
- 239000001923 methylcellulose Substances 0.000 description 1
- 235000010981 methylcellulose Nutrition 0.000 description 1
- LXCFILQKKLGQFO-UHFFFAOYSA-N methylparaben Chemical compound COC(=O)C1=CC=C(O)C=C1 LXCFILQKKLGQFO-UHFFFAOYSA-N 0.000 description 1
- 235000019813 microcrystalline cellulose Nutrition 0.000 description 1
- 239000008108 microcrystalline cellulose Substances 0.000 description 1
- 229940016286 microcrystalline cellulose Drugs 0.000 description 1
- 235000010446 mineral oil Nutrition 0.000 description 1
- 239000002480 mineral oil Substances 0.000 description 1
- 229930014626 natural product Natural products 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 239000002777 nucleoside Substances 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 235000003715 nutritional status Nutrition 0.000 description 1
- 239000002751 oligonucleotide probe Substances 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000012188 paraffin wax Substances 0.000 description 1
- 229920002866 paraformaldehyde Polymers 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- PTMHPRAIXMAOOB-UHFFFAOYSA-N phosphoramidic acid Chemical class NP(O)(O)=O PTMHPRAIXMAOOB-UHFFFAOYSA-N 0.000 description 1
- 150000008300 phosphoramidites Chemical class 0.000 description 1
- 239000001267 polyvinylpyrrolidone Substances 0.000 description 1
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 description 1
- 229920000036 polyvinylpyrrolidone Polymers 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 230000003449 preventive effect Effects 0.000 description 1
- 239000013615 primer Substances 0.000 description 1
- 239000002987 primer (paints) Substances 0.000 description 1
- QELSKZZBTMNZEB-UHFFFAOYSA-N propylparaben Chemical compound CCCOC(=O)C1=CC=C(O)C=C1 QELSKZZBTMNZEB-UHFFFAOYSA-N 0.000 description 1
- 229960003415 propylparaben Drugs 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 238000012552 review Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000000600 sorbitol Substances 0.000 description 1
- 235000010356 sorbitol Nutrition 0.000 description 1
- 230000009870 specific binding Effects 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 239000000375 suspending agent Substances 0.000 description 1
- 239000003765 sweetening agent Substances 0.000 description 1
- 239000006188 syrup Substances 0.000 description 1
- 235000020357 syrup Nutrition 0.000 description 1
- 239000000454 talc Substances 0.000 description 1
- 229910052623 talc Inorganic materials 0.000 description 1
- 235000012222 talc Nutrition 0.000 description 1
- 238000002626 targeted therapy Methods 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 1
- 239000001226 triphosphate Substances 0.000 description 1
- 235000011178 triphosphate Nutrition 0.000 description 1
- 230000009385 viral infection Effects 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 239000000080 wetting agent Substances 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/43—Enzymes; Proenzymes; Derivatives thereof
- A61K38/45—Transferases (2)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
- A61P1/16—Drugs for disorders of the alimentary tract or the digestive system for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/158—Expression markers
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/04—Endocrine or metabolic disorders
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/08—Hepato-biliairy disorders other than hepatitis
- G01N2800/085—Liver diseases, e.g. portal hypertension, fibrosis, cirrhosis, bilirubin
Definitions
- the present invention relates to a composition for preventing or treating fatty liver comprising an AXL protein or gene as an active ingredient.
- Fatty liver refers to a state in which fat is accumulated in hepatocytes, and the proportion of fat in a normal liver is about 5%.
- fatty liver gets worse and the fat mass inside the liver cells grows, the important components of the cells, including the nucleus, are pushed to one side and the function of the liver cells deteriorates. As a result, the circulation of blood and lymph in the liver is impaired. When this happens, liver cells cannot receive oxygen and nutrients properly, and liver function deteriorates.
- Fatty liver can be divided into alcoholic fatty liver and non-alcoholic fatty liver.
- Alcoholic fatty liver is caused by alcohol. The more alcohol you drink, the more likely it occurs, and if you continue to drink alcohol, the ability of the liver to metabolize alcohol decreases, making fatty liver more severe. It also occurs in cases of poor nutritional status. Some alcoholic fatty livers can progress to alcoholic hepatitis and cirrhosis, leading to death.
- Non-alcoholic fatty liver is a case of increased fat accumulation in the liver (exceeding 5% of hepatocytes), regardless of alcohol or viral infection, and is caused by increased fat synthesis and excretion in the liver. About 59% of non-alcoholic fatty liver patients caused by an imbalance between input and output of free fatty acids are caused by increased absorption of free fatty acids from the outside, and about 26% of patients are intrahepatic. It is known to be caused by an increase in the input of free fatty acids due to an increase in fat biosynthesis.
- AXL is a receptor tyrosine kinase (RTX), whose name is derived from the Greek word meaning 'uncontrolled'.
- RTX receptor tyrosine kinase
- AXL protein regulates biological processes such as intracellular signal transduction, cytoskeletal function and gene expression, and is known to be expressed in platelets, monocytes, liver, hippocampus and cerebellum. or its mechanism is unknown.
- an object of the present invention is to provide a pharmaceutical composition for preventing or treating metabolic diseases comprising an AXL protein or a polynucleotide encoding the AXL protein.
- Another object of the present invention is to provide a method for inhibiting fat biosynthesis in the liver by administering an AXL protein or a vector containing a polynucleotide encoding the AXL protein to a non-human animal.
- Another object of the present invention is to provide a method for treating metabolic diseases comprising administering the pharmaceutical composition described above to a subject.
- Another object of the present invention is to provide a composition for diagnosing metabolic diseases comprising a substance for measuring the level of AXL gene or AXL protein.
- Another object of the present invention is to provide an information providing method for diagnosing a metabolic disease comprising measuring the level of AXL gene or AXL protein.
- Another object of the present invention relates to a method for screening a therapeutic agent for metabolic diseases, comprising the step of selecting a substance that increases the expression or activity level of AXL in a subject.
- the present inventors confirmed that the expression of a gene related to fat synthesis in liver tissue increases when a high-fat diet is performed in an animal model in which AXL expression is suppressed.
- the present invention relates to a pharmaceutical composition for preventing or treating a metabolic disease comprising an AXL protein or a polynucleotide encoding the AXL protein, and a composition for diagnosing a metabolic disease comprising an agent for measuring the level of AXL.
- One aspect of the present invention relates to a pharmaceutical composition for preventing or treating metabolic diseases, comprising an AXL protein or a vector containing a polynucleotide encoding the AXL protein as an active ingredient.
- prevention refers to any action that suppresses or delays the onset of metabolic diseases by administration of the pharmaceutical composition according to the present invention.
- treatment refers to all activities in which symptoms of metabolic diseases are improved or beneficially changed by administration of the pharmaceutical composition according to the present invention.
- Metabolic diseases which are diseases to be prevented or treated by the composition of the present invention, are obesity, type 2 diabetes, dyslipidemia, insulin resistance, hepatic steatosis and/or fatty liver, specifically non-alcoholic fatty liver disease. Hepatic steatosis or non-alcoholic fatty liver, specifically, non-alcoholic fatty liver induced by a high fat diet is more preferable, but is not limited thereto.
- composition of the present invention provides a therapeutic agent for metabolic diseases targeting AXL as part of gene-targeted therapy.
- the AXL gene is a receptor tyrosine kinase (RTX), and its name is derived from the Greek word meaning 'uncontrolled'.
- the AXL protein is a protein encoded by the AXL gene and is also called AXL receptor tyrosine kinase.
- the AXL protein described above regulates biological processes such as intracellular signal transduction, cytoskeletal function, and gene expression, and is known to be expressed in platelets, monocytes, liver, hippocampus, and cerebellum, but treatment of fatty liver through promotion of AXL expression or activity. None has been reported about it yet.
- the nucleotide sequence of the AXL gene is shown in SEQ ID NO: 1 or 2.
- the nucleotide sequence of the gene is registered in the gene bank (Gene ID: BC058230.1), those skilled in the art will be able to easily obtain it.
- amino acid sequence of the AXL protein may be represented by SEQ ID NO: 3 or 4, but is not limited thereto.
- AXL protein according to the present invention includes a protein having the amino acid sequence of SEQ ID NO: 3 or 4 and functional equivalents of the protein.
- “Functional equivalent” means at least 70% or more, preferably 80% or more, more preferably 90% or more, even more preferably, the amino acid sequence of SEQ ID NO: 3 or 4 as a result of addition, substitution or deletion of amino acids. has a sequence homology of 95% or more, and refers to a protein exhibiting substantially the same physiological activity as the protein having the amino acid sequence of SEQ ID NO: 3 or 4.
- “Substantially homogeneous physiological activity” means preventive or therapeutic activity of metabolic diseases.
- the vector containing the polynucleotide encoding the AXL protein may be a recombinant viral vector, a plasmid vector, a cosmid vector or a bacteriophage vector, preferably a recombinant viral vector, but is not limited thereto.
- the recombinant virus may be adenovirus, adeno-associated virus, retrovirus, herpes simplex virus, and/or lentivirus, preferably adenovirus, but is not limited thereto.
- the AXL protein or the polynucleotide encoding the AXL protein has an effect of inhibiting fat biosynthesis in the liver.
- the pharmaceutical composition according to the present invention may include a pharmaceutically acceptable carrier in addition to the active ingredient.
- the pharmaceutically acceptable carrier is one commonly used in the formulation, lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose , polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, and mineral oil, but are not limited thereto.
- lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents, preservatives, and the like may be further included.
- the pharmaceutical composition of the present invention may be administered orally or parenterally (for example, intravenously, subcutaneously, intraperitoneally or topically applied) depending on the desired method, and the dosage is the patient's condition and weight, the degree of disease , Depending on the drug form, administration route and time, it can be appropriately selected by those skilled in the art.
- composition of the present invention is administered in a pharmaceutically effective amount.
- pharmaceutically effective amount means an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment, and an effective amount is a patient's disease type, severity, drug activity, drug sensitivity, time of administration, route of administration and excretion rate, duration of treatment, factors including drugs used concurrently, and other factors well known in the medical field.
- the pharmaceutical composition according to the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered single or multiple times. Considering all of the above factors, it is important to administer an amount that can obtain the maximum effect with the minimum amount without side effects, which can be easily determined by those skilled in the art.
- the effective amount of the pharmaceutical composition of the present invention may vary depending on the patient's age, sex, condition, body weight, absorption rate, inactivation rate and excretion rate of the active ingredient in the body, type of disease, and concomitant drugs, and in general, body weight 0.001 to 150 mg per 1 kg, preferably 0.01 to 100 mg may be administered daily or every other day, or divided into 1 to 3 times a day.
- body weight 0.001 to 150 mg per 1 kg, preferably 0.01 to 100 mg may be administered daily or every other day, or divided into 1 to 3 times a day.
- the dosage is not limited to the scope of the present invention in any way.
- Another aspect of the present invention relates to a method of inhibiting fat biosynthesis in the liver by administering an AXL protein or a vector containing a polynucleotide encoding the AXL protein to a non-human animal.
- the method of the present invention uses the above-described AXL protein or a vector containing the polynucleotide encoding the AXL protein, descriptions of overlapping contents with those described above are omitted to avoid excessive complexity in the present specification.
- Another aspect of the present invention relates to a method for treating a metabolic disease comprising administering the aforementioned pharmaceutical composition to a subject.
- the "individual” means a subject in need of treatment of a disease, and more specifically, means a mammal such as a human or non-human primate, mouse, dog, cat, horse, and cow.
- Another aspect of the present invention relates to a composition for diagnosing metabolic diseases, including a substance for measuring the level of AXL gene or AXL protein.
- diagnosis means the action of confirming the presence or characteristics of a pathological state, that is, a metabolic disease, by administration of the composition according to the present invention.
- the material for measuring the level of the AXL gene may be a primer or probe capable of amplifying the AXL gene, and the material for measuring the level of the AXL protein may be an antibody that specifically recognizes the AXL protein, Any agent capable of measuring the expression or activity level of a gene may be included without limitation.
- a "primer” is a nucleic acid sequence having a short free 3-terminal hydroxyl group, which can form base pairs with a complementary template and refers to a short nucleic acid sequence that functions as a starting point for template strand copying.
- a primer can initiate DNA synthesis in the presence of a reagent for polymerization (i.e., DNA polymerase or reverse transcriptase) and four different nucleoside triphosphates in an appropriate buffer and temperature.
- probe refers to a nucleic acid fragment such as RNA or DNA corresponding to a few bases to several hundred bases in length that can form a specific binding with mRNA, and is labeled to detect the presence or absence of a specific mRNA. You can check.
- the probe may be manufactured in the form of an oligonucleotide probe, a single stranded DNA probe, a double stranded DNA probe, an RNA probe, or the like. Selection of suitable probes and hybridization conditions can be modified based on those known in the art.
- antibody means a specific protein molecule directed against an antigenic site.
- an antibody means an antibody that specifically binds to a marker protein, and includes both polyclonal antibodies, monoclonal antibodies and recombinant antibodies.
- nucleic acid sequences of the genes of the present invention are registered in the gene bank, those skilled in the art can design antisense oligonucleotides, primer pairs, or probes that specifically amplify specific regions of these genes based on the sequences.
- Antisense oligonucleotides, primers or probes of the present invention can be chemically synthesized using the phosphoramidite solid support method, or other well-known methods. Such nucleic acid sequences can also be modified using a number of means known in the art. Non-limiting examples of such modifications include methylation, capping, substitution of one or more natural nucleotides with homologues, and modifications between nucleotides, such as uncharged linkages such as methyl phosphonates, phosphotriesters, phosphoro amidates, carbamates, etc.) or to charged linkages (eg phosphorothioates, phosphorodithioates, etc.).
- Another aspect of the present invention relates to an information providing method for diagnosing a metabolic disease comprising the following steps.
- the expression level or the amount of protein of the AXL gene is decreased compared to the expression level or protein level of the normal control group, it can be determined that a metabolic disease, specifically, fatty liver has progressed.
- the method includes mammals, particularly humans.
- the human includes a person suspected or not suspected of having a metabolic disease and a person in need of a diagnosis of a metabolic disease.
- the “biological sample” may consist of tissues, cells, blood, serum, plasma, saliva and urine, but is not limited thereto.
- the sample may be pretreated before being used for detection, and may include, for example, filtration, distillation, extraction, concentration, inactivation of interfering components, addition of reagents, and the like.
- nucleic acids and proteins may be separated from the sample and used for detection.
- RT-PCR reverse transcription polymerase reaction
- Competitive RTPCR competitive reverse transcription polymerase reaction
- Realtime RT-PCR real-time reverse transcription polymerase reaction
- RPA RNase protection assay
- measurement of protein expression level is a process of confirming the presence and expression level of the protein expressed from the gene of the present invention in liver tissue cells in order to confirm the progress of metabolic disease, and is performed by measuring the amount of protein.
- Analysis methods for this include Western blot, enzyme linked immunosorbent assay (ELISA), radioimmunoassay (RIA), radioimmunodiffusion, Ouchterlony immunodiffusion method, rocket immunoelectrophoresis , tissue immunostaining, immunoprecipitation assay, complement fixation assay, flow cytometry (Fluorescence Activated Cell Sorter, FACS), protein chip, etc., but are not limited thereto.
- Another aspect of the present invention relates to a method for screening a therapeutic agent for metabolic diseases, comprising the step of selecting a substance that increases the expression or activity level of an AXL protein or a gene encoding the AXL protein in a subject.
- the expression or activity level of the AXL protein or the gene encoding the same when the expression or activity level of the AXL protein or the gene encoding the same is measured in the subject and the candidate for treatment of metabolic disease is administered, the expression or activity level of the AXL protein or the gene encoding the same is increased compared to the previous candidate.
- the substance can be used as a therapeutic agent for metabolic diseases.
- the method may measure the expression or activity level of the AXL protein or the gene encoding the AXL protein in the individual before treating the subject with the target material and measure and compare it after the treatment, or treat some of the subjects with the target material to compare the target material to the control group. The degree of expression or activity can also be compared.
- the step of measuring the level of expression or activity of the AXL protein or the gene encoding it is polymerase reaction (PCR), reverse transcription polymerase reaction (RT-PCR), competitive reverse transcription polymerase reaction (Competitive RT-PCR), real-time reverse transcription polymerization Enzyme reaction (Realtime RT-PCR), RNase protection assay (RPA; Northern blotting), DNA chip, Western blotting, ELISA (enzyme linked immunosorbent assay), radioimmunoassay (Radioimmunoassay, RIA), radioimmunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, immunoprecipitation assay, complement fixation assay, protein chip , and can be measured by one or more methods selected from the group consisting of in vitro kinase assay (IKKB), but is not limited thereto.
- IKKB in vitro kinase assay
- a method of treating the target substance to the subject is not particularly limited, and for example, a biological sample obtained from the subject may be treated.
- the biological sample may be any of the samples described above.
- target substance means an unknown substance used in screening to examine whether or not it affects the expression or activity level of AXL of the present invention.
- the target substance includes, but is not limited to, chemicals, nucleotides, antisense-RNA, siRNA (small interference RNA) and natural product extracts.
- the present invention relates to a composition for preventing or treating fatty liver, a composition for diagnosis, and a method for providing information for diagnosis.
- a composition for preventing or treating fatty liver a composition for diagnosis
- a method for providing information for diagnosis By regulating the expression and activity of AXL, the marker of the present invention, it is expected to be useful for the development of a therapeutic agent for fatty liver.
- FIG. 1a shows the size (FIG. 1a), body weight (FIG. 1b) and food intake of AXL knockout mice when fed with a high-fat diet (HFD) or normal diet (NCD) according to an embodiment of the present invention.
- Figure 2 is a diagram confirming the effect of AXL knockout on the size and mass of the liver of mice when fed a high-fat diet (HFD) or normal diet (NCD) according to an embodiment of the present invention.
- HFD high-fat diet
- NCD normal diet
- Figure 3 is a diagram confirming the effect of AXL knockout on the size and mass of subcutaneous white adipose tissue of mice when fed with a high-fat diet (HFD) or normal diet (NCD) according to an embodiment of the present invention.
- HFD high-fat diet
- NCD normal diet
- FIG. 4a to 4c are glucose tolerance (FIG. 4a), insulin tolerance (FIG. 4b) and fasting blood glucose/serum insulin levels of AXL knockout mice when fed a high-fat diet (HFD) according to an embodiment of the present invention.
- FIGs 5a and 5b show, according to an embodiment of the present invention, AXL knockout during high-fat diet (HFD) feeding the liver (Fig. 5a) and subcutaneous white adipose tissue (Fig. 5b) of fat biosynthesis-related proteins ( It is a diagram confirming the effect on ACC, FAS, PPARr and SREBP1c) levels.
- the present invention relates to a pharmaceutical composition for preventing or treating metabolic diseases, including an AXL protein or a vector containing a polynucleotide encoding the AXL protein.
- Example 1 High fat diet Axl Determination of the possibility of developing obesity and/or hepatic steatosis in knockout mice
- Axl wild type mice and Axl knockout (KO) mice (Axl -/- ) mice (C57BL/6) were provided and used by Professor Jeon Tae-hoon's lab at Korea University. Mice were housed in a controlled environment (12/12h light/dark cycle; humidity 50-60%; ambient temperature 22 °C). All mouse experiments were performed in a facility in accordance with institutional guidelines, and the experimental protocols were conducted by Korea Research Institute of Bioscience and Biotechnology and Chungnam. It was approved by the University Institutional Review Board.
- mice fed the high-fat diet (HFD) or normal diet (NCD) were sacrificed at the 17th week of feeding, and the liver (FIG. 2) and subcutaneous white adipose tissue (SWAT) ( The size and mass of Fig. 3) were confirmed.
- HFD high-fat diet
- NCD normal diet
- SWAT subcutaneous white adipose tissue
- Axl KO mice fed a normal diet showed similar body weight compared to WT mice (littermates) fed the same diet.
- WT mice littermates
- HFD high-fat diet
- HFD high-fat diet
- HFD high-fat diet
- mice Seven-week-old WT and Axl KO mice (C57BL/6) were fed a high-fat diet (HFD) with 60% of calories from fat (D12492) and sacrificed at 26 weeks of feeding.
- HFD high-fat diet
- ACC, FAS, PPARr and SREBP1c in liver and subcutaneous white adipose tissue (sWAT) were confirmed using Western blot.
- Anti-pACC(S79), anti-ACC, anti-FAS, anti-PPAR ⁇ , anti-SREBP1c antibody or anti-GAPDH antibody were used as primary antibodies, and HRP conjugated anti-mouse IgG antibodies were used as secondary antibodies.
- GAPDH was selected as a control group for comparing protein expression.
- anti-pACC S79
- anti-ACC anti-FAS and anti-GAPDH antibodies
- anti-PPAR ⁇ and anti-SREBP1c antibodies were obtained from Santa Cruz Biotechnology (USA)
- HRP Conjugated anti-mouse IgG antibodies were obtained from Koma biotech (Korea).
- liver and subcutaneous white adipose tissue were collected and fixed in 4% paraformaldehyde overnight at 4°C. Fixed samples were embedded in paraffin, cut into 5 m sections and mounted on slides to create a series of 6 slides with similar adjacent sections. One slide of each series was stained with H&E (hematoxylin & eosin).
- the present invention relates to a composition for preventing or treating fatty liver comprising an AXL protein or gene as an active ingredient.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Medicinal Chemistry (AREA)
- Immunology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
- Pharmacology & Pharmacy (AREA)
- Molecular Biology (AREA)
- Analytical Chemistry (AREA)
- Veterinary Medicine (AREA)
- Hematology (AREA)
- Biotechnology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Urology & Nephrology (AREA)
- Zoology (AREA)
- Gastroenterology & Hepatology (AREA)
- Wood Science & Technology (AREA)
- Microbiology (AREA)
- Biomedical Technology (AREA)
- Biochemistry (AREA)
- Physics & Mathematics (AREA)
- Pathology (AREA)
- General Chemical & Material Sciences (AREA)
- Genetics & Genomics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Obesity (AREA)
- Diabetes (AREA)
- Biophysics (AREA)
- Epidemiology (AREA)
- Cell Biology (AREA)
- General Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- General Physics & Mathematics (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
The present invention relates to a composition for preventing or treating steatosis, a diagnostic composition, and an information provision method for diagnosis. The expression and activity of AXL, which is a marker of the present invention, are regulated, and thus it is expected that AXL is effectively used in the development of a therapeutic agent for steatosis.
Description
본 발명은 AXL 단백질 또는 유전자를 유효성분으로 포함하는 지방간 예방 또는 치료용 조성물에 관한 것이다.The present invention relates to a composition for preventing or treating fatty liver comprising an AXL protein or gene as an active ingredient.
지방간(steatosis, fatty liver)은 간세포 속에 지방이 축적된 상태를 말하며, 정상 간은 지방이 차지하는 비율은 5% 정도인데, 이보다 많은 지방이 축적된 상태를 지방간이라고 한다. 지방간이 악화되어 간세포 속의 지방 덩어리가 커지면 핵을 포함한 세포의 중요한 구성성분이 한쪽으로 밀려 간세포의 기능이 저하되며, 세포 내에 축적된 지방으로 인하여 팽창된 간세포들이 간세포 사이에 있는 미세 혈관과 임파선을 압박하여 간 내의 혈액과 임파액의 순환에 장애가 생기게 된다. 이렇게 되면 간세포는 산소와 영양공급을 적절히 받을 수 없어 간기능이 저하된다.Fatty liver (steatosis, fatty liver) refers to a state in which fat is accumulated in hepatocytes, and the proportion of fat in a normal liver is about 5%. When fatty liver gets worse and the fat mass inside the liver cells grows, the important components of the cells, including the nucleus, are pushed to one side and the function of the liver cells deteriorates. As a result, the circulation of blood and lymph in the liver is impaired. When this happens, liver cells cannot receive oxygen and nutrients properly, and liver function deteriorates.
지방간은 알코올성 지방간과 비알코올성 지방간으로 나눌 수 있다. 알코올성 지방간은 술이 원인이다. 술을 많이 마실수록 잘 발생하며, 지속적으로 술을 섭취하면 간에서 알코올을 대사하는 능력이 떨어져 지방간 발생이 더 심해진다. 또 영양상태가 나쁜 경우에도 잘 발생한다. 알코올성 지방간 중 일부는 알코올성 간염과 간경변으 로 진행되어 사망할 수 있다.Fatty liver can be divided into alcoholic fatty liver and non-alcoholic fatty liver. Alcoholic fatty liver is caused by alcohol. The more alcohol you drink, the more likely it occurs, and if you continue to drink alcohol, the ability of the liver to metabolize alcohol decreases, making fatty liver more severe. It also occurs in cases of poor nutritional status. Some alcoholic fatty livers can progress to alcoholic hepatitis and cirrhosis, leading to death.
비알코올성 지방간은 술 또는 바이러스 감염과 상관없이 간 내 지방축적이 증가한 경우로(간세포의 5% 초과), 간에서 지방 합성 증가 및 배출의 문제로 발생한다. 유리지방산(fatty acid)의 투입(input)과 배출(output)의 불균형에 의해 유발되는 비알코올성 지방간 환자의 약 59%는 외부로부터 유리지방산의 흡수 증가에 의해 발생하고, 약 26% 환자는 간 내 지방 생합성의 증가에 따른 유리지방산의 투입(input) 증가를 원인으로 유발되는 것으로 알려져 있다.Non-alcoholic fatty liver is a case of increased fat accumulation in the liver (exceeding 5% of hepatocytes), regardless of alcohol or viral infection, and is caused by increased fat synthesis and excretion in the liver. About 59% of non-alcoholic fatty liver patients caused by an imbalance between input and output of free fatty acids are caused by increased absorption of free fatty acids from the outside, and about 26% of patients are intrahepatic. It is known to be caused by an increase in the input of free fatty acids due to an increase in fat biosynthesis.
지방간은 병의 원인과 심각성이 밝혀졌고, 식이 조절과 운동을 통하여 증상을 개선해야 하지만, 지방간 환자가 이를 실천하지 못하는 경우가 많다. 따라서 효과적인 지방간 치료 약물의 개발 필요성이 요구되고 있다.The cause and severity of fatty liver disease have been identified, and the symptoms should be improved through diet control and exercise, but in many cases, patients with fatty liver fail to practice it. Therefore, there is a need for the development of effective drugs for treating fatty liver.
한편, AXL은 수용체 티로신 키나아제(receptor tyrosine kinase, RTX)로, 명칭은 '통제되지 않는'을 의미하는 그리스어에서 파생되었다. AXL 단백질은 세포 내 신호 전달, 세포 골격 기능 및 유전자 발현과 같은 생물학적 과정을 조절하며, 혈소판, 단핵구, 간, 해마 및 소뇌에서 발현되는 것으로 알려져 있으나, AXL의 지방간과의 관계, 대사와의 관계, 또는 이의 기전 등에 대해서는 알려진 바 없다.Meanwhile, AXL is a receptor tyrosine kinase (RTX), whose name is derived from the Greek word meaning 'uncontrolled'. AXL protein regulates biological processes such as intracellular signal transduction, cytoskeletal function and gene expression, and is known to be expressed in platelets, monocytes, liver, hippocampus and cerebellum. or its mechanism is unknown.
[선행기술문헌][Prior art literature]
[특허문헌][Patent Literature]
KR 10-2018-0118384 A1KR 10-2018-0118384 A1
본 발명자들은 AXL 유전자 및 이의 기전에 대하여 연구하던 중, AXL의 발현이 억제된 동물 모델에서 고지방 식이 조건을 실시하는 경우 간 조직 내의 지방 합성과 관련된 유전자 발현이 증가한다는 것을 확인함으로써, 본 발명을 완성하였다.While studying the AXL gene and its mechanism, the present inventors completed the present invention by confirming that the expression of genes related to fat synthesis in liver tissue increases when a high-fat diet is performed in an animal model in which the expression of AXL is suppressed. did
따라서, 본 발명의 목적은 AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 대사성 질환의 예방 또는 치료용 약학 조성물을 제공하는 것이다.Accordingly, an object of the present invention is to provide a pharmaceutical composition for preventing or treating metabolic diseases comprising an AXL protein or a polynucleotide encoding the AXL protein.
본 발명의 다른 목적은 AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 벡터를 인간을 제외한 동물에게 투여하여 간 내 지방 생합성을 억제시키는 방법을 제공하는 것이다.Another object of the present invention is to provide a method for inhibiting fat biosynthesis in the liver by administering an AXL protein or a vector containing a polynucleotide encoding the AXL protein to a non-human animal.
본 발명의 또 다른 목적은 상술한 약학 조성물을 개체에 투여하는 단계를 포함하는 대사성 질환의 치료방법을 제공하는 것이다.Another object of the present invention is to provide a method for treating metabolic diseases comprising administering the pharmaceutical composition described above to a subject.
본 발명의 또 다른 목적은 AXL 유전자의 수준 또는 AXL 단백질의 수준을 측정하는 물질을 포함하는 대사성 질환 진단용 조성물을 제공하는 것이다.Another object of the present invention is to provide a composition for diagnosing metabolic diseases comprising a substance for measuring the level of AXL gene or AXL protein.
본 발명의 또 다른 목적은 AXL 유전자의 수준 또는 AXL 단백질의 수준을 측정하는 단계를 포함하는 대사성 질환의 진단을 위한 정보제공방법을 제공하는 것이다.Another object of the present invention is to provide an information providing method for diagnosing a metabolic disease comprising measuring the level of AXL gene or AXL protein.
본 발명의 또 다른 목적은 개체 내 AXL의 발현 또는 활성 정도를 증가시키는 물질을 선별하는 단계를 포함하는 대사성 질환 치료제의 스크리닝 방법에 관한 것이다.Another object of the present invention relates to a method for screening a therapeutic agent for metabolic diseases, comprising the step of selecting a substance that increases the expression or activity level of AXL in a subject.
본 발명자들은 AXL 유전자 및 이의 기전에 대하여 연구하던 중, AXL의 발현이 억제된 동물 모델에서 고지방 식이 조건을 실시하는 경우 간 조직 내의 지방 합성과 관련된 유전자 발현이 증가한다는 것을 확인하였다.While studying the AXL gene and its mechanism, the present inventors confirmed that the expression of a gene related to fat synthesis in liver tissue increases when a high-fat diet is performed in an animal model in which AXL expression is suppressed.
따라서, 본 발명은 AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 대사성 질환의 예방 또는 치료용 약학 조성물 및 AXL의 수준을 측정하는 제제를 포함하는 대사성 질환 진단용 조성물에 관한 것이다.Accordingly, the present invention relates to a pharmaceutical composition for preventing or treating a metabolic disease comprising an AXL protein or a polynucleotide encoding the AXL protein, and a composition for diagnosing a metabolic disease comprising an agent for measuring the level of AXL.
이하, 본 발명을 더욱 자세히 설명하고자 한다.Hereinafter, the present invention will be described in more detail.
본 발명의 일 양태는 AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 벡터를 유효성분으로 포함하는, 대사성 질환의 예방 또는 치료용 약학 조성물에 관한 것이다.One aspect of the present invention relates to a pharmaceutical composition for preventing or treating metabolic diseases, comprising an AXL protein or a vector containing a polynucleotide encoding the AXL protein as an active ingredient.
본 발명에서 "예방"이란 본 발명에 따른 약학 조성물의 투여에 의해 대사성 질환의 발병을 억제시키거나 발병을 지연시키는 모든 행위를 의미한다.In the present invention, "prevention" refers to any action that suppresses or delays the onset of metabolic diseases by administration of the pharmaceutical composition according to the present invention.
본 발명에서 "치료"란 본 발명에 따른 약학 조성물의 투여에 의해 대사성 질환에 대한 증세가 호전되거나 이롭게 변경되는 모든 행위를 의미한다.In the present invention, "treatment" refers to all activities in which symptoms of metabolic diseases are improved or beneficially changed by administration of the pharmaceutical composition according to the present invention.
본 발명의 조성물에 의한 예방, 치료 대상 질병인 대사성 질환은 비만, 제2형 당뇨, 이상지질혈증, 인슐린저항성, 간 지방증(hepatic steatosis) 및/또는 지방간(fatty liver), 구체적으로 비알코올성 지방간일 수 있고, 간 지방증 또는 비알코올성 지방간, 구체적으로 고지방(high fat) 식이에 의해 유도되는 비알코올성 지방간인 것이 보다 바람직하나, 이에 한정되지 않는다. Metabolic diseases, which are diseases to be prevented or treated by the composition of the present invention, are obesity, type 2 diabetes, dyslipidemia, insulin resistance, hepatic steatosis and/or fatty liver, specifically non-alcoholic fatty liver disease. Hepatic steatosis or non-alcoholic fatty liver, specifically, non-alcoholic fatty liver induced by a high fat diet is more preferable, but is not limited thereto.
본 발명의 조성물은 유전자 표적 치료의 일환으로서, AXL을 표적으로 하는 대사성 질환 치료제를 제공한다.The composition of the present invention provides a therapeutic agent for metabolic diseases targeting AXL as part of gene-targeted therapy.
상기 AXL 유전자는 수용체 티로신 키나아제(receptor tyrosine kinase, RTX)로, 명칭은 '통제되지 않는'을 의미하는 그리스어에서 파생되었다. AXL 단백질은 상기 AXL 유전자에 의해 암호화되는 단백질로서, AXL 리셉터 타이로신 키나아제(AXL receptor tyrosine kinase)라고도 불리운다. 상기한 AXL 단백질은 세포 내 신호 전달, 세포 골격 기능 및 유전자 발현과 같은 생물학적 과정을 조절하며, 혈소판, 단핵구, 간, 해마 및 소뇌에서 발현되는 것으로 알려져 있으나, AXL의 발현 또는 활성 촉진을 통한 지방간 치료에 대해서는 아직 보고된 바가 없다.The AXL gene is a receptor tyrosine kinase (RTX), and its name is derived from the Greek word meaning 'uncontrolled'. The AXL protein is a protein encoded by the AXL gene and is also called AXL receptor tyrosine kinase. The AXL protein described above regulates biological processes such as intracellular signal transduction, cytoskeletal function, and gene expression, and is known to be expressed in platelets, monocytes, liver, hippocampus, and cerebellum, but treatment of fatty liver through promotion of AXL expression or activity. Nothing has been reported about it yet.
상기 AXL 유전자의 뉴클레오티드 서열을 서열번호 1 또는 2에 나타내었다. 또한, 상기 유전자는 뉴클레오티드 서열이 유전자 은행에 등록(Gene ID: BC058230.1)되어 있으므로 당업자라면 쉽게 입수가 가능할 것이다.The nucleotide sequence of the AXL gene is shown in SEQ ID NO: 1 or 2. In addition, since the nucleotide sequence of the gene is registered in the gene bank (Gene ID: BC058230.1), those skilled in the art will be able to easily obtain it.
상기 AXL 단백질의 아미노산 서열을 서열번호 3 또는 4로 표시될 수 있으나, 이에 제한되는 것은 아니다. The amino acid sequence of the AXL protein may be represented by SEQ ID NO: 3 or 4, but is not limited thereto.
본 발명에 따른 AXL 단백질의 범위는 서열번호 3 또는 4의 아미노산 서열을 갖는 단백질 및 상기 단백질의 기능적 동등물을 포함한다. "기능적 동등물"이란 아미노산의 부가, 치환 또는 결실의 결과, 상기 서열번호 3 또는 4의 아미노산 서열과 적어도 70% 이상, 바람직하게는 80% 이상, 더욱 바람직하게는 90% 이상, 더 더욱 바람직하게는 95% 이상의 서열 상동성을 갖는 것으로, 서열번호 3 또는 4의 아미노산 서열을 갖는 단백질과 실질적으로 동질의 생리활성을 나타내는 단백질을 말한다. "실질적으로 동질의 생리활성"이란 대사성 질환의 예방 또는 치료 활성을 의미한다.The scope of the AXL protein according to the present invention includes a protein having the amino acid sequence of SEQ ID NO: 3 or 4 and functional equivalents of the protein. "Functional equivalent" means at least 70% or more, preferably 80% or more, more preferably 90% or more, even more preferably, the amino acid sequence of SEQ ID NO: 3 or 4 as a result of addition, substitution or deletion of amino acids. has a sequence homology of 95% or more, and refers to a protein exhibiting substantially the same physiological activity as the protein having the amino acid sequence of SEQ ID NO: 3 or 4. "Substantially homogeneous physiological activity" means preventive or therapeutic activity of metabolic diseases.
상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 벡터는 재조합 바이러스 벡터, 플라스미드 벡터, 코스미드 벡터 또는 박테리오파아지 벡터일 수 있으며, 바람직하게는 재조합 바이러스 벡터일 수 있으나, 이에 한정되지 않는다. The vector containing the polynucleotide encoding the AXL protein may be a recombinant viral vector, a plasmid vector, a cosmid vector or a bacteriophage vector, preferably a recombinant viral vector, but is not limited thereto.
상기 재조합 바이러스는 아데노 바이러스, 아데노 부속 바이러스(adeno-associated virus), 레트로 바이러스, 헤르페스 심플렉스 바이러스 및/또는 렌티바이러스일 수 있으며, 바람직하게는 아데노 바이러스일 수 있으나, 이에 한정되지 않는다.The recombinant virus may be adenovirus, adeno-associated virus, retrovirus, herpes simplex virus, and/or lentivirus, preferably adenovirus, but is not limited thereto.
상기 AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드는 간에서 지방 생합성을 억제하는 효과가 있다.The AXL protein or the polynucleotide encoding the AXL protein has an effect of inhibiting fat biosynthesis in the liver.
본 발명에 따른 약학 조성물은 상기 유효성분 이외에 약제학적으로 허용되는 담체를 포함할 수 있다. 이때, 약제학적으로 허용되는 담체는 제제 시에 통상적으로 이용되는 것으로서, 락토스, 덱스트로스, 수크로스, 솔비톨, 만니톨, 전분, 아카시아고무, 인산 칼슘, 알기네이트, 젤라틴, 규산 칼슘, 미세 결정성셀룰로스, 폴리비닐피로리돈, 셀룰로스, 물, 시럽, 메틸 셀룰로스, 메틸히드록시벤조에이트, 프로필 히드록시벤조에이트, 활석, 스테아르산 마그네슘 및 미네랄 오일 등을 포함하나, 이에 한정되는 것은 아니다. 또한, 상기성분들 이외에 윤활제, 습윤제, 감미제, 향미제, 유화제, 현탁제, 보존제 등을 추가로 포함할 수 있다.The pharmaceutical composition according to the present invention may include a pharmaceutically acceptable carrier in addition to the active ingredient. At this time, the pharmaceutically acceptable carrier is one commonly used in the formulation, lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose , polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, and mineral oil, but are not limited thereto. In addition to the above components, lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents, preservatives, and the like may be further included.
본 발명의 약학 조성물은 목적하는 방법에 따라 경구 투여하거나 비경구투여(예를 들어, 정맥 내, 피하, 복강 내 또는 국소에 적용)할 수 있으며, 투여량은 환자의 상태 및 체중, 질병의 정도, 약물형태, 투여경로 및 시간에 따라 다르지만, 당업자에 의해 적절하게 선택될 수 있다.The pharmaceutical composition of the present invention may be administered orally or parenterally (for example, intravenously, subcutaneously, intraperitoneally or topically applied) depending on the desired method, and the dosage is the patient's condition and weight, the degree of disease , Depending on the drug form, administration route and time, it can be appropriately selected by those skilled in the art.
본 발명의 약학 조성물은 약학적으로 유효한 양으로 투여한다. 본 발명에 있어서 "약학적으로 유효한 양"은 의학적 치료에 적용 가능한 합리적인 수혜/위험 비율로 질환을 치료하기에 충분한 양을 의미하며, 유효량은 환자의 질환의 종류, 중증도, 약물의 활성, 약물에 대한 민감도, 투여 시간, 투여 경로 및 배출비율, 치료기간, 동시 사용되는 약물을 포함한 요소 및 기타 의학 분야에 잘 알려진 요소에 따라 결정될 수 있다. The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment, and an effective amount is a patient's disease type, severity, drug activity, drug sensitivity, time of administration, route of administration and excretion rate, duration of treatment, factors including drugs used concurrently, and other factors well known in the medical field.
본 발명에 따른 약학 조성물은 개별 치료제로 투여하거나 다른 치료제와 병용하여 투여될 수 있고, 종래의 치료제와는 순차적 또는 동시에 투여될 수 있으며, 단일 또는 다중 투여될 수 있다. 상기 요소들을 모두 고려하여 부작용 없이 최소한의 양으로 최대 효과를 얻을 수 있는 양을 투여하는 것이 중요하며, 이는 당업자에 의해 용이하게 결정될 수 있다.The pharmaceutical composition according to the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered single or multiple times. Considering all of the above factors, it is important to administer an amount that can obtain the maximum effect with the minimum amount without side effects, which can be easily determined by those skilled in the art.
구체적으로 본 발명의 약학 조성물의 유효량은 환자의 연령, 성별, 상태, 체중, 체내에 활성 성분의 흡수도, 불활성율 및 배설속도, 질병종류, 병용되는 약물에 따라 달라질 수 있으며, 일반적으로는 체중 1kg 당 0.001 내지 150mg, 바람직하게는 0.01 내지 100mg을 매일 또는 격일 투여하거나, 1일 1 내지 3회로 나누어 투여할 수 있다. 그러나 투여 경로, 비만의 중증도, 성별, 체중, 연령 등에 따라서 증감 될 수 있으므로 상기 투여량이 어떠한 방법으로도 본 발명의 범위를 한정하는 것은 아니다.Specifically, the effective amount of the pharmaceutical composition of the present invention may vary depending on the patient's age, sex, condition, body weight, absorption rate, inactivation rate and excretion rate of the active ingredient in the body, type of disease, and concomitant drugs, and in general, body weight 0.001 to 150 mg per 1 kg, preferably 0.01 to 100 mg may be administered daily or every other day, or divided into 1 to 3 times a day. However, since it may increase or decrease depending on the route of administration, severity of obesity, gender, weight, age, etc., the dosage is not limited to the scope of the present invention in any way.
본 발명의 다른 일 양태는 AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 벡터를 인간을 제외한 동물에게 투여하여 간 내 지방 생합성을 억제시키는 방법에 관한 것이다.Another aspect of the present invention relates to a method of inhibiting fat biosynthesis in the liver by administering an AXL protein or a vector containing a polynucleotide encoding the AXL protein to a non-human animal.
본 발명의 방법은 상술한 AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 벡터를 이용하기 때문에, 상술한 내용과 중복된 내용은 본 명세서의 과도한 복잡성을 피하기 위하여 그 기재를 생략한다.Since the method of the present invention uses the above-described AXL protein or a vector containing the polynucleotide encoding the AXL protein, descriptions of overlapping contents with those described above are omitted to avoid excessive complexity in the present specification.
본 발명의 또 다른 일 양태는 상술한 약학 조성물을 개체에 투여하는 단계를 포함하는 대사성 질환의 치료방법에 관한 것이다.Another aspect of the present invention relates to a method for treating a metabolic disease comprising administering the aforementioned pharmaceutical composition to a subject.
상기 "개체"란 질병의 치료를 필요로 하는 대상을 의미하고, 보다 구체적으로는, 인간 또는 비-인간인 영장류, 마우스(mouse), 개, 고양이, 말 및 소 등의 포유류를 의미한다.The "individual" means a subject in need of treatment of a disease, and more specifically, means a mammal such as a human or non-human primate, mouse, dog, cat, horse, and cow.
본 발명의 또 다른 일 양태는 AXL 유전자의 수준 또는 AXL 단백질의 수준을 측정하는 물질을 포함하는, 대사성 질환 진단용 조성물에 관한 것이다.Another aspect of the present invention relates to a composition for diagnosing metabolic diseases, including a substance for measuring the level of AXL gene or AXL protein.
본 발명에서 "진단"이란 본 발명에 따른 조성물의 투여에 의해 병리 상태, 즉 대사성 질환의 존재 또는 특징을 확인하는 행위를 의미한다.In the present invention, "diagnosis" means the action of confirming the presence or characteristics of a pathological state, that is, a metabolic disease, by administration of the composition according to the present invention.
본 발명에서 상기 AXL 유전자의 수준을 측정하는 물질은 AXL 유전자를 증폭할 수 있는 프라이머 또는 프로브일 수 있고, 상기 AXL 단백질의 수준을 측정하는 물질은 AXL 단백질을 특이적으로 인식하는 항체일 수 있으나, 유전자의 발현 또는 활성 수준을 측정할 수 있는 제제라면 제한 없이 포함될 수 있다.In the present invention, the material for measuring the level of the AXL gene may be a primer or probe capable of amplifying the AXL gene, and the material for measuring the level of the AXL protein may be an antibody that specifically recognizes the AXL protein, Any agent capable of measuring the expression or activity level of a gene may be included without limitation.
본 발명에서 "프라이머"는 짧은 자유 3말단 수산화기를 가지는 핵산 서열로 상보적인 주형(template)과 염기쌍을 형성할 수 있고 템플레이트 가닥 복사를 위한 시작 지점으로 기능을 하는 짧은 핵산 서열을 의미한다. 프라이머는 적절한 완충용액 및 온도에서 중합반응(즉, DNA 중합효소 또는 역전사효소)을 위한 시약 및 상이한 4가지 뉴클레오사이드 트리포스페이트의 존재하에서 DNA 합성을 개시할 수 있다.In the present invention, a "primer" is a nucleic acid sequence having a short free 3-terminal hydroxyl group, which can form base pairs with a complementary template and refers to a short nucleic acid sequence that functions as a starting point for template strand copying. A primer can initiate DNA synthesis in the presence of a reagent for polymerization (i.e., DNA polymerase or reverse transcriptase) and four different nucleoside triphosphates in an appropriate buffer and temperature.
본 발명에서 "프로브"란 mRNA와 특이적 결합을 이룰 수 있는 짧게는 수 염기 내지 길게는 수백 염기에 해당하는 RNA 또는 DNA 등의 핵산 단편을 의미하며 표지(Labelling)되어 있어서 특정 mRNA의 존재 유무를 확인할 수 있다. 프로브는 올리고 뉴클레오티드 프로브, 단쇄 DNA(single stranded DNA) 프로브, 이중쇄 DNA(double stranded DNA) 프로브, RNA 프로브 등의 형태로 제작될 수 있다. 적당한 프로브의 선택 및 혼성화 조건은 당업계에 공지된 것을 기초로 변형할 수 있다.In the present invention, "probe" refers to a nucleic acid fragment such as RNA or DNA corresponding to a few bases to several hundred bases in length that can form a specific binding with mRNA, and is labeled to detect the presence or absence of a specific mRNA. You can check. The probe may be manufactured in the form of an oligonucleotide probe, a single stranded DNA probe, a double stranded DNA probe, an RNA probe, or the like. Selection of suitable probes and hybridization conditions can be modified based on those known in the art.
본 발명에서 "항체"란 항원성 부위에 대해서 지시되는 특이적인 단백질 분자를 의미한다. 본 발명의 목적상, 항체는 마커 단백질에 대해 특이적으로 결합하는 항체를 의미하며, 다클론 항체, 단클론 항체 및 재조합 항체를 모두 포함한다.In the present invention, "antibody" means a specific protein molecule directed against an antigenic site. For the purposes of the present invention, an antibody means an antibody that specifically binds to a marker protein, and includes both polyclonal antibodies, monoclonal antibodies and recombinant antibodies.
본 발명의 유전자는 핵산 서열이 유전자 은행에 등록되어 있으므로 당업자는 상기 서열을 바탕으로 이들 유전자의 특정 영역을 특이적으로 증폭하는 안티센스 올리고뉴클레오티드, 프라이머 쌍 또는 프로브를 디자인할 수 있다.Since the nucleic acid sequences of the genes of the present invention are registered in the gene bank, those skilled in the art can design antisense oligonucleotides, primer pairs, or probes that specifically amplify specific regions of these genes based on the sequences.
본 발명의 안티센스 올리고뉴클레오티드, 프라이머 또는 프로브는 포스포르아미다이트 고체 지지체 방법, 또는 기타 널리 공지된 방법을 사용하여 화학적으로 합성할 수 있다. 이러한 핵산 서열은 또한 당해 분야에 공지된 많은 수단을 이용하여 변형시킬 수 있다. 이러한 변형의 비-제한적인 예로는 메틸화, 캡화, 천연 뉴클레오티드 하나 이상의 동족체로의 치환, 및 뉴클레오티드 간의 변형, 예를 들면, 하전되지 않은 연결체(예: 메틸 포스포네이트, 포스포트리에스테르, 포스포로아미데이트, 카바메이트 등) 또는 하전된 연결체(예: 포스포로티오에이트, 포스포로디티오에이트 등)로의 변형이 있다.Antisense oligonucleotides, primers or probes of the present invention can be chemically synthesized using the phosphoramidite solid support method, or other well-known methods. Such nucleic acid sequences can also be modified using a number of means known in the art. Non-limiting examples of such modifications include methylation, capping, substitution of one or more natural nucleotides with homologues, and modifications between nucleotides, such as uncharged linkages such as methyl phosphonates, phosphotriesters, phosphoro amidates, carbamates, etc.) or to charged linkages (eg phosphorothioates, phosphorodithioates, etc.).
본 발명의 또 다른 일 양태는 다음의 단계를 포함하는 대사성 질환의 진단을 위한 정보제공방법에 관한 것이다.Another aspect of the present invention relates to an information providing method for diagnosing a metabolic disease comprising the following steps.
생물학적 시료에 존재하는 AXL 유전자의 발현량 또는 단백질의 양을 측정하는 단계; 및 Measuring the amount of expression or protein of the AXL gene present in the biological sample; and
상기 측정 결과를 정상 대조군 시료의 AXL 유전자의 발현량 또는 단백질의 양과 비교하는 단계.Comparing the measurement result with the amount of expression or protein of the AXL gene in the normal control sample.
상기 AXL 유전자의 발현량 또는 단백질의 양이 정상 대조군의 AXL 유전자의 발현량 또는 단백질의 양과 비교하여 감소하는 경우 대사성 질환, 구체적으로 지방간이 진행된 것으로 판정할 수 있다.When the expression level or the amount of protein of the AXL gene is decreased compared to the expression level or protein level of the normal control group, it can be determined that a metabolic disease, specifically, fatty liver has progressed.
상기 방법은 포유류, 특히 인간을 대상으로 포함한다.The method includes mammals, particularly humans.
구체적으로는, 상기 인간은 대사성 질환이 발병했을 것으로 의심되는 사람 또는 의심되지 않는 사람으로 대사성 질환 진단 여부가 필요한 사람을 포함한다.Specifically, the human includes a person suspected or not suspected of having a metabolic disease and a person in need of a diagnosis of a metabolic disease.
상기 "생물학적 시료"는 조직, 세포, 혈액, 혈청, 혈장, 타액 및 뇨로 이루어질 수 있으나, 이에 한정되지 않는다.The "biological sample" may consist of tissues, cells, blood, serum, plasma, saliva and urine, but is not limited thereto.
상기 시료는 검출에 사용하기 전에 전처리할 수 있으며, 예를 들어, 여과, 증류, 추출, 농축, 방해 성분의 불활성화, 시약의 첨가 등을 포함할 수 있다. 또한, 상기 시료로부터 핵산 및 단백질을 분리하여 검출에 사용할 수 있다.The sample may be pretreated before being used for detection, and may include, for example, filtration, distillation, extraction, concentration, inactivation of interfering components, addition of reagents, and the like. In addition, nucleic acids and proteins may be separated from the sample and used for detection.
본 발명에서 "유전자 발현량 측정"이란 대사성 질환의 진행 여부를 확인하기 위하여 간 조직 세포에서 상기 유전자의 mRNA 존재 여부와 발현 정도를 확인하는 과정으로, mRNA의 양을 측정하여 이루어진다. 이를 위한 분석 방법으로는 예를 들어, 역전사 중합효소반응(RT-PCR), 경쟁적 역전사 중합효소반응(Competitive RTPCR), 실시간 역전사 중합효소반응(Realtime RT-PCR), RNase 보호 분석법(RPA; RNase protection assay), 노던 블랏팅(Northern blotting), DNA 칩 등이 있으나, 이에 한정되지 않는다.In the present invention, "measurement of gene expression level" is a process of confirming the presence or absence of mRNA of the gene and the expression level of the gene in liver tissue cells in order to confirm the progress of metabolic disease, and is performed by measuring the amount of mRNA. Analytical methods for this include, for example, reverse transcription polymerase reaction (RT-PCR), competitive reverse transcription polymerase reaction (Competitive RTPCR), real-time reverse transcription polymerase reaction (Realtime RT-PCR), RNase protection assay (RPA; RNase protection assay), Northern blotting, DNA chip, etc., but are not limited thereto.
본 발명에서 "단백질 발현량 측정"이란 대사성 질환의 진행 여부를 확인하기 위하여 간 조직 세포에서 본 발명의 유전자로부터 발현된 단백질의 존재 여부와 발현 정도를 확인하는 과정으로, 단백질의 양을 측정하여 이루어진다. 이를 위한 분석 방법으로는 웨스턴 블랏, ELISA(enzyme linked immunosorbent assay), 방사선면역분석(RIA: Radioimmunoassay), 방사 면역 확산법(radioimmunodiffusion), 오우크테로니(Ouchterlony) 면역확산법, 로케트(rocket) 면역전기영동, 조직면역 염색, 면역침전 분석법(Immunoprecipitation Assay), 보체 고정 분석법(Complement Fixation Assay), 유세포분석(Fluorescence Activated Cell Sorter, FACS), 단백질 칩(protein chip) 등이 있으나, 이에 한정되지 않는다.In the present invention, "measurement of protein expression level" is a process of confirming the presence and expression level of the protein expressed from the gene of the present invention in liver tissue cells in order to confirm the progress of metabolic disease, and is performed by measuring the amount of protein. . Analysis methods for this include Western blot, enzyme linked immunosorbent assay (ELISA), radioimmunoassay (RIA), radioimmunodiffusion, Ouchterlony immunodiffusion method, rocket immunoelectrophoresis , tissue immunostaining, immunoprecipitation assay, complement fixation assay, flow cytometry (Fluorescence Activated Cell Sorter, FACS), protein chip, etc., but are not limited thereto.
본 발명의 또 다른 일 양태는 개체 내 AXL 단백질 또는 이를 암호화하는 유전자의 발현 또는 활성 정도를 증가시키는 물질을 선별하는 단계를 포함하는 대사성 질환 치료제의 스크리닝 방법에 관한 것이다.Another aspect of the present invention relates to a method for screening a therapeutic agent for metabolic diseases, comprising the step of selecting a substance that increases the expression or activity level of an AXL protein or a gene encoding the AXL protein in a subject.
상기 방법에서 개체 내 AXL 단백질 또는 이를 암호화하는 유전자의 발현 또는 활성 정도를 측정하고 대사성 질환 치료용 후보물질 투여 시, 상기 AXL 단백질 또는 이를 암호화하는 유전자의 발현 또는 활성 정도가 기존 대비 증가되는 경우 상기 후보물질은 대사성 질환 치료제로 이용될 수 있다.In the above method, when the expression or activity level of the AXL protein or the gene encoding the same is measured in the subject and the candidate for treatment of metabolic disease is administered, the expression or activity level of the AXL protein or the gene encoding the same is increased compared to the previous candidate. The substance can be used as a therapeutic agent for metabolic diseases.
상기 방법은 개체에 대상 물질 처리 전에 개체 내 AXL 단백질 또는 이를 암호화하는 유전자의 발현 또는 활성 정도를 측정하고 처리 후에 이를 측정하여 비교할 수도 있고, 여러 개체 중 일부 개체에 대상 물질을 처리하여 비교군과 상기 발현 또는 활성 정도를 비교할 수도 있다.The method may measure the expression or activity level of the AXL protein or the gene encoding the AXL protein in the individual before treating the subject with the target material and measure and compare it after the treatment, or treat some of the subjects with the target material to compare the target material to the control group. The degree of expression or activity can also be compared.
상기 AXL 단백질 또는 이를 암호화하는 유전자의 발현 또는 활성 정도를 측정하는 단계는 중합효소반응(PCR), 역전사 중합효소반응(RT-PCR), 경쟁적 역전사 중합효소반응(Competitive RT-PCR), 실시간 역전사 중합효소반응(Realtime RT-PCR), RNase 보호 분석법(RPA; RNase protection assay), 노던 블롯팅(Northern blotting), DNA 칩, 웨스턴 블롯팅(Western blotting), ELISA(enzyme linked immunosorbent assay), 방사선 면역 분석(Radioimmunoassay, RIA), 방사 면역 확산법(radioimmunodiffusion), 오우크테로니(Ouchterlony) 면역확산법, 로케트(rocket) 면역전기영동, 면역침전 분석법(Immunoprecipitation Assay), 보체 고정 분석법(Complement Fixation Assay), 단백질 칩, 및 In vitro kinase assay(IKKB)로 이루어진 군으로부터 선택된 1종 이상의 방법으로 측정할 수 있으나, 이에 제한되는 것은 아니다.The step of measuring the level of expression or activity of the AXL protein or the gene encoding it is polymerase reaction (PCR), reverse transcription polymerase reaction (RT-PCR), competitive reverse transcription polymerase reaction (Competitive RT-PCR), real-time reverse transcription polymerization Enzyme reaction (Realtime RT-PCR), RNase protection assay (RPA; Northern blotting), DNA chip, Western blotting, ELISA (enzyme linked immunosorbent assay), radioimmunoassay (Radioimmunoassay, RIA), radioimmunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, immunoprecipitation assay, complement fixation assay, protein chip , and can be measured by one or more methods selected from the group consisting of in vitro kinase assay (IKKB), but is not limited thereto.
상기 방법에서 개체에 대상 물질을 처리하는 방법은 특별히 한정되지 않으며, 예를 들면 개체로부터 얻어진 생물학적 시료에 처리할 수 있다. 생물학적 시료는 전술한 바의 시료일 수 있다.In the above method, a method of treating the target substance to the subject is not particularly limited, and for example, a biological sample obtained from the subject may be treated. The biological sample may be any of the samples described above.
상기 "대상 물질"은 본 발명의 AXL의 발현 또는 활성 정도에 영향을 미치는지 여부를 검사하기 위하여 스크리닝에서 이용되는 미지의 물질을 의미한다. 상기 대상 물질은 화학물질, 뉴클레오타이드, 안티센스-RNA, siRNA(small interference RNA) 및 천연물 추출물을 포함하나, 이에 한정되는 것은 아니다.The "target substance" means an unknown substance used in screening to examine whether or not it affects the expression or activity level of AXL of the present invention. The target substance includes, but is not limited to, chemicals, nucleotides, antisense-RNA, siRNA (small interference RNA) and natural product extracts.
본 발명은 지방간 예방 또는 치료용 조성물, 진단용 조성물, 진단을 위한 정보제공방법에 관한 것이다. 본 발명의 마커인 AXL의 발현 및 활성을 조절하여 지방간 치료제 개발에 유용하게 사용할 수 있을 것으로 기대된다.The present invention relates to a composition for preventing or treating fatty liver, a composition for diagnosis, and a method for providing information for diagnosis. By regulating the expression and activity of AXL, the marker of the present invention, it is expected to be useful for the development of a therapeutic agent for fatty liver.
도 1a 내지 1c는 본 발명의 일 실시예에 따라, 고지방 식이(HFD) 또는 일반 식이(NCD) 급이 시 AXL 녹아웃(knockout)이 마우스의 크기(도 1a), 체중(도 1b) 및 음식 섭취량(도 1c)에 미치는 영향을 확인한 도이다. WT (n=6), AXL KO (n=7)Figures 1a to 1c show the size (FIG. 1a), body weight (FIG. 1b) and food intake of AXL knockout mice when fed with a high-fat diet (HFD) or normal diet (NCD) according to an embodiment of the present invention. (FIG. 1c) is a diagram confirming the effect. WT (n=6), AXL KO (n=7)
도 2는 본 발명의 일 실시예에 따라, 고지방 식이(HFD) 또는 일반 식이(NCD) 급이 시 AXL 녹아웃(knockout)이 마우스의 간의 크기 및 질량에 미치는 영향을 확인한 도이다. WT (n=6), AXL KO (n=7)Figure 2 is a diagram confirming the effect of AXL knockout on the size and mass of the liver of mice when fed a high-fat diet (HFD) or normal diet (NCD) according to an embodiment of the present invention. WT (n=6), AXL KO (n=7)
도 3은 본 발명의 일 실시예에 따라, 고지방 식이(HFD) 또는 일반 식이(NCD) 급이 시 AXL 녹아웃(knockout)이 마우스의 피하 백색 지방조직의 크기 및 질량에 미치는 영향을 확인한 도이다. WT (n=6), AXL KO (n=7)Figure 3 is a diagram confirming the effect of AXL knockout on the size and mass of subcutaneous white adipose tissue of mice when fed with a high-fat diet (HFD) or normal diet (NCD) according to an embodiment of the present invention. WT (n=6), AXL KO (n=7)
도 4a 내지 4c는 본 발명의 일 실시예에 따라, 고지방 식이(HFD) 급이 시 AXL 녹아웃(knockout)이 마우스의 포도당 내성(도 4a), 인슐린 내성(도 4b) 및 공복 혈당/혈청 인슐린 수준(도 4c)에 미치는 영향을 확인한 도이다. WT (n=6), AXL KO (n=7)4a to 4c are glucose tolerance (FIG. 4a), insulin tolerance (FIG. 4b) and fasting blood glucose/serum insulin levels of AXL knockout mice when fed a high-fat diet (HFD) according to an embodiment of the present invention. (FIG. 4c) is a diagram confirming the effect. WT (n=6), AXL KO (n=7)
도 5a 및 5b는 본 발명의 일 실시예에 따라, 고지방 식이(HFD) 급이 시 AXL 녹아웃(knockout)이 마우스의 간(도 5a) 및 피하 백색 지방조직(도 5b)의 지방 생합성 관련 단백질(ACC, FAS, PPARr 및 SREBP1c) 수준에 미치는 영향을 확인한 도이다. WT (n=6), AXL KO (n=7)Figures 5a and 5b show, according to an embodiment of the present invention, AXL knockout during high-fat diet (HFD) feeding the liver (Fig. 5a) and subcutaneous white adipose tissue (Fig. 5b) of fat biosynthesis-related proteins ( It is a diagram confirming the effect on ACC, FAS, PPARr and SREBP1c) levels. WT (n=6), AXL KO (n=7)
본 발명은 AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 벡터를 포함하는, 대사성 질환의 예방 또는 치료용 약학 조성물에 관한 것이다. The present invention relates to a pharmaceutical composition for preventing or treating metabolic diseases, including an AXL protein or a vector containing a polynucleotide encoding the AXL protein.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하고자 한다. 이들 실시예는 오로지 본 발명을 보다 구체적으로 설명하기 위한 것으로, 본 발명의 요지에 따라 본 발명의 범위가 이들 실시예에 의해 제한되지 않는다는 것은 당업계에서 통상의 지식을 가진 자에 있어서 자명할 것이다.Hereinafter, the present invention will be described in more detail through examples. These examples are only for explaining the present invention in more detail, and it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples according to the gist of the present invention. .
GraphPad Prism을 통계 분석에 사용하였으며, 실험의 결과는 최소 3개의 개별 실험으로부터 얻은 Mean±Standard error로 표기하였다. 그룹 간의 차이는 Tukey 사후 테스트를 통해 ANOVA로 분석되었으며, 데이터 쌍의 비교는 Student's t-test로 하였다. p 값이 0.05 미만일 때 통계적으로 유의한 것으로 판단하였다. 실험 결과의 정량 분석은 Image J 소프트웨어(버전 1.47)를 사용하여 수행되었다.GraphPad Prism was used for statistical analysis, and the results of the experiments were expressed as the mean ± standard error obtained from at least three separate experiments. Differences between groups were analyzed by ANOVA with Tukey's post hoc test, and pairwise comparisons of data were performed by Student's t-test. A p value of less than 0.05 was considered statistically significant. Quantitative analysis of the experimental results was performed using Image J software (version 1.47).
실시예 1. 고지방 식이 Example 1. High fat diet
AxlAxl
녹아웃 마우스의 비만 및/또는 간 지방증 발병 가능성 확인 Determination of the possibility of developing obesity and/or hepatic steatosis in knockout mice
Axl 야생형(wild type, WT) 마우스 및 Axl 녹아웃(knockout, KO)(Axl-/-) 마우스(C57BL/6)는 고려대학교(Korea university) 전태훈 교수 랩에서 제공받아 사용하였다. 마우스는 통제된 환경(12/12h 명/암 사이클; 습도 50-60%; 주변 온도 22 ℃에 수용되었다. 모든 마우스 실험은 기관 지침에 따른 시설에서 수행되었으며, 실험 프로토콜은 한국생명공학연구원 및 충남대학교 기관심사위원회의 승인을 받았다.Axl wild type (WT) mice and Axl knockout (KO) (Axl -/- ) mice (C57BL/6) were provided and used by Professor Jeon Tae-hoon's lab at Korea University. Mice were housed in a controlled environment (12/12h light/dark cycle; humidity 50-60%; ambient temperature 22 °C). All mouse experiments were performed in a facility in accordance with institutional guidelines, and the experimental protocols were conducted by Korea Research Institute of Bioscience and Biotechnology and Chungnam. It was approved by the University Institutional Review Board.
먼저, 7주령 WT 및 Axl KO 마우스(C57BL/6)에 대하여, 60%의 칼로리가 지방으로 구성된 고지방 식이(high fat diet, HFD)(D12492; Research Diets, USA) 또는 일반 식이(normal chow diet, NCD)를 급이하고, 급이 13주(NCD) 또는 17주(HFD) 차의 크기(도 1a), 체중 변화(도 1b) 및 음식 섭취량(도 1c)을 확인하였다.First, for 7-week-old WT and Axl KO mice (C57BL/6), a high fat diet (HFD) (D12492; Research Diets, USA) or a normal chow diet (normal chow diet, NCD) was fed, and the size of the difference (Fig. 1a), weight change (Fig. 1b) and food intake (Fig. 1c) at 13 weeks (NCD) or 17 weeks (HFD) of feeding were confirmed.
다음으로, 상기의 고지방 식이(HFD) 또는 일반 식이(NCD)를 급이한 마우스를 급이 17주 차에 희생시키고, 간(도 2) 및 피하 백색 지방조직(Subcutaneous white adipose tissue, sWAT)(도 3)의 크기 및 질량을 확인하였다.Next, the mice fed the high-fat diet (HFD) or normal diet (NCD) were sacrificed at the 17th week of feeding, and the liver (FIG. 2) and subcutaneous white adipose tissue (SWAT) ( The size and mass of Fig. 3) were confirmed.
도 1a 내지 1c에서 확인할 수 있듯이, 일반 식이(NCD)를 먹인 Axl KO 마우스는 동일한 식이를 먹인 WT 마우스(littermates)와 비교하여 유사한 체중을 나타내었다. 반면, 고지방 식이(HFD)를 먹인 Axl KO 마우스의 경우 음식 섭취량에는 차이가 없었으나, 크기와 체중은 유의하게 증가하였다.As can be seen in Figures 1a to 1c, Axl KO mice fed a normal diet (NCD) showed similar body weight compared to WT mice (littermates) fed the same diet. On the other hand, in the case of Axl KO mice fed a high-fat diet (HFD), there was no difference in food intake, but size and weight significantly increased.
또한, 도 2 및 3에서 확인할 수 있듯이, 일반 식이(NCD)를 먹인 Axl KO 마우스에서는 아무 변화가 없었으나, 고지방 식이(HFD)를 먹인 Axl KO 마우스의 경우 간 및 sWAT 질량이 모두 유의하게 증가하였다.In addition, as can be seen in Figures 2 and 3, there was no change in Axl KO mice fed a normal diet (NCD), but in the case of Axl KO mice fed a high-fat diet (HFD), both liver and sWAT mass increased significantly .
실시예 2. 고지방 식이 Example 2. High fat diet
AxlAxl
녹아웃 마우스의 포도당 내성(GTT) 및 인슐린 내성(ITT) 평가 Assessment of glucose tolerance (GTT) and insulin tolerance (ITT) in knockout mice
[Glucose tolerance test (GTT)][Glucose tolerance test (GTT)]
4주령 WT 및 Axl KO 마우스(C57BL/6)에 대하여, 60%의 칼로리가 지방으로 구성된 고지방 식이(HFD)(D12492)를 16주 동안 급이한 후, 16 시간 동안 금식시키고 공복 혈당(fasting glucose level)을 측정하였다. 그 다음, 복강 내 포도당(1g/kg body weight)을 투여하고 일정 시간마다(15, 30, 45 및 60 분) 혈당을 측정하였다.4-week-old WT and Axl KO mice (C57BL/6) were fed a high-fat diet (HFD) (D12492) consisting of 60% of calories from fat for 16 weeks, followed by fasting for 16 hours and fasting glucose level) was measured. Next, glucose (1 g/kg body weight) was administered intraperitoneally, and blood glucose was measured at regular intervals (15, 30, 45, and 60 minutes).
[Insulin tolerance test (ITT)][Insulin tolerance test (ITT)]
4주령 WT 및 Axl KO 마우스(C57BL/6)에 대하여, 60%의 칼로리가 지방으로 구성된 고지방 식이(HFD)(D12492)를 16주 동안 급이한 후, 16 시간 동안 금식시키고 공복 혈당(fasting glucose level)을 측정하였다. 그 다음, 복강 내 인슐린(0.75 unit/kg body weight; 91077C; Sigma-Aldrich)을 투여하고 일정 시간마다(15, 30, 45 및 60 분) 혈당을 측정하였다.4-week-old WT and Axl KO mice (C57BL/6) were fed a high-fat diet (HFD) (D12492) consisting of 60% of calories from fat for 16 weeks, followed by fasting for 16 hours and fasting glucose level) was measured. Then, intraperitoneal insulin (0.75 unit/kg body weight; 91077C; Sigma-Aldrich) was administered, and blood glucose was measured at regular intervals (15, 30, 45, and 60 minutes).
도 4a 내지 4c에서 확인할 수 있듯이, 고지방 식이(HFD)를 먹인 Axl KO 마우스의 경우 WT 마우스에 비하여 포도당 내성 및 인슐린 내성이 모두 유의하게 손상되었으며(도 4a 및 4b), 또한 Axl KO 마우스의 공복 혈당 및 혈청 인슐린 수준은 WT 마우스(littermates)와 비교하여 유의하게 더 높은 것으로 나타났다(도 4c). As can be seen in Figures 4a to 4c, in the case of Axl KO mice fed a high-fat diet (HFD), both glucose tolerance and insulin resistance were significantly impaired compared to WT mice (Figures 4a and 4b), and fasting blood glucose of Axl KO mice and serum insulin levels were significantly higher compared to WT mice (littermates) (FIG. 4c).
실시예 3. 고지방 식이 Example 3. High fat diet
AxlAxl
녹아웃 마우스의 지방 생합성 관련 단백질의 변화 확인 Confirmation of changes in fat biosynthesis-related proteins in knockout mice
7주령 WT 및 Axl KO 마우스(C57BL/6)에 대하여, 60%의 칼로리가 지방으로 구성된 고지방 식이(HFD)(D12492)를 급이하고, 급이 26주 차에 희생시켰다.Seven-week-old WT and Axl KO mice (C57BL/6) were fed a high-fat diet (HFD) with 60% of calories from fat (D12492) and sacrificed at 26 weeks of feeding.
먼저, 간 및 피하 백색 지방조직(sWAT) 내 ACC, FAS, PPARr 및 SREBP1c의 단백질 발현량을 웨스턴 블랏을 이용하여 확인하였다. 1차 항체로서 항-pACC(S79), 항-ACC, 항-FAS, 항-PPARγ, 항-SREBP1c 항체 또는 항-GAPDH 항체를 사용하였고, 2차 항체로서 HRP 접합 항-마우스 IgG 항체를 사용하였다. 단백질의 발현을 비교하기 위한 대조군으로는 GAPDH를 대상으로 선정하였다. 이때, 항-pACC(S79), 항-ACC, 항-FAS 및 항-GAPDH 항체는 Cell Signaling Technology(USA)에서, 항-PPARγ 및 항-SREBP1c 항체는 Santa Cruz Biotechnology(USA)에서 입수하였으며, HRP 접합 항-마우스 IgG 항체는 Koma biotech(Korea)로부터 입수하였다.First, the protein expression levels of ACC, FAS, PPARr and SREBP1c in liver and subcutaneous white adipose tissue (sWAT) were confirmed using Western blot. Anti-pACC(S79), anti-ACC, anti-FAS, anti-PPARγ, anti-SREBP1c antibody or anti-GAPDH antibody were used as primary antibodies, and HRP conjugated anti-mouse IgG antibodies were used as secondary antibodies. . GAPDH was selected as a control group for comparing protein expression. At this time, anti-pACC (S79), anti-ACC, anti-FAS and anti-GAPDH antibodies were obtained from Cell Signaling Technology (USA), anti-PPARγ and anti-SREBP1c antibodies were obtained from Santa Cruz Biotechnology (USA), and HRP Conjugated anti-mouse IgG antibodies were obtained from Koma biotech (Korea).
다음으로, 간 및 피하 백색 지방조직(sWAT)을 수집하여 4 ℃에서 밤새 4 % 파라포름알데하이드에 고정하였다. 고정된 샘플을 파라핀에 삽입하고, 5 m 섹션으로 절단한 후 슬라이드에 장착하여 유사한 인접 섹션이 있는 일련의 6개 슬라이드를 제작하였다. 각 시리즈의 한 슬라이드는 H&E(hematoxylin & eosin) 염색하였다.Next, liver and subcutaneous white adipose tissue (sWAT) were collected and fixed in 4% paraformaldehyde overnight at 4°C. Fixed samples were embedded in paraffin, cut into 5 m sections and mounted on slides to create a series of 6 slides with similar adjacent sections. One slide of each series was stained with H&E (hematoxylin & eosin).
도 5a 및 5b에서 확인할 수 있듯이, 고지방 식이(HFD)를 먹인 Axl KO 마우스에서 지방 생합성 관련 단백질(ACC, FAS, PPARr 및 SREBP1c) 수준이 증가하였다. 또한, H&E 염색 결과 sWAT 지방세포의 크기 및 간 지방증이 Axl WT 마우스에 비하여 Axl KO 마우스에서 더 많이 유도되었음을 알 수 있었다. As can be seen in Figures 5a and 5b, the levels of fat biosynthesis-related proteins (ACC, FAS, PPARr and SREBP1c) increased in Axl KO mice fed a high-fat diet (HFD). In addition, as a result of H&E staining, it was found that the size of sWAT adipocytes and hepatic steatosis were more induced in Axl KO mice than in Axl WT mice.
본 발명은 AXL 단백질 또는 유전자를 유효성분으로 포함하는 지방간 예방 또는 치료용 조성물에 관한 것이다.The present invention relates to a composition for preventing or treating fatty liver comprising an AXL protein or gene as an active ingredient.
Claims (16)
- AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 벡터를 포함하는, 대사성 질환의 예방 또는 치료용 약학 조성물.A pharmaceutical composition for preventing or treating metabolic diseases comprising a vector containing an AXL protein or a polynucleotide encoding the AXL protein.
- 제 1항에 있어서, 상기 대사성 질환은 비만, 제2형 당뇨, 이상지질혈증, 인슐린 저항성, 간 지방증(hepatic steatosis) 및 비알코올성 지방간(fatty liver)으로 구성된 군으로부터 선택되는 것을 특징으로 하는, 대사성 질환의 예방 및 치료용 약학적 조성물.The metabolic disease according to claim 1, characterized in that the metabolic disease is selected from the group consisting of obesity, type 2 diabetes, dyslipidemia, insulin resistance, hepatic steatosis and non-alcoholic fatty liver. A pharmaceutical composition for preventing and treating diseases.
- 제 2항에 있어서, 상기 비알코올성 지방간은 고지방(high fat) 식이에 의해 유도되는 비알코올성 지방간인 것인, 대사성 질환의 예방 또는 치료용 약학 조성물.According to claim 2, wherein the non-alcoholic fatty liver is a high-fat (high fat) non-alcoholic fatty liver induced by a diet would be, a pharmaceutical composition for the prevention or treatment of metabolic diseases.
- 제 1항에 있어서, 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 벡터는 재조합 바이러스 벡터, 플라스미드 벡터, 코스미드 벡터 및 박테리오파아지 벡터로 이루어진 군으로부터 선택되는 것인, 대사성 질환의 예방 또는 치료용 약학 조성물.The method of claim 1, wherein the vector containing the polynucleotide encoding the AXL protein is selected from the group consisting of a recombinant viral vector, a plasmid vector, a cosmid vector and a bacteriophage vector. composition.
- 제 4항에 있어서, 상기 재조합 바이러스는 아데노 바이러스, 아데노 부속 바이러스(adeno-associated virus), 레트로 바이러스, 헤르페스 심플렉스 바이러스 및 렌티바이러스로 이루어진 군으로부터 선택되는 것인, 대사성 질환의 예방 또는 치료용 약학 조성물.The method of claim 4, wherein the recombinant virus is selected from the group consisting of adenovirus, adeno-associated virus, retrovirus, herpes simplex virus, and lentivirus. composition.
- 제 1항에 있어서, 상기 AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드는 간에서 지방 생합성을 억제하는 것을 특징으로 하는, 대사성 질환의 예방 또는 치료용 약학 조성물.The pharmaceutical composition for preventing or treating metabolic diseases according to claim 1, wherein the AXL protein or the polynucleotide encoding the AXL protein inhibits fat biosynthesis in the liver.
- AXL 단백질 또는 상기 AXL 단백질을 암호화하는 폴리뉴클레오티드를 포함하는 벡터를 인간을 제외한 동물에게 투여하여 간 내 지방 생합성을 억제시키는 방법.A method of inhibiting fat biosynthesis in the liver by administering an AXL protein or a vector containing a polynucleotide encoding the AXL protein to a non-human animal.
- AXL 유전자의 수준 또는 AXL 단백질의 수준을 측정하는 물질을 포함하는, 대사성 질환 진단용 조성물.A composition for diagnosing metabolic diseases, comprising a substance for measuring the level of AXL gene or AXL protein.
- 제 8항에 있어서, 상기 대사성 질환은 고지방(high fat) 식이에 의해 유도되는 비알코올성 지방간인 것인, 대사성 질환 진단용 조성물.The composition for diagnosing metabolic diseases according to claim 8, wherein the metabolic disease is non-alcoholic fatty liver induced by a high fat diet.
- 제 8항에 있어서, 상기 AXL 유전자의 수준을 측정하는 물질은 AXL 유전자를 증폭시킬 수 있는 프라이머 또는 프로브인 것인, 대사성 질환 진단용 조성물.The composition for diagnosing metabolic diseases according to claim 8, wherein the material for measuring the level of the AXL gene is a primer or probe capable of amplifying the AXL gene.
- 제 8항에 있어서, 상기 AXL 단백질의 수준을 측정하는 물질은 AXL 단백질을 특이적으로 인식하는 항체인 것인, 대사성 질환 진단용 조성물.The composition for diagnosing metabolic diseases according to claim 8, wherein the material for measuring the level of the AXL protein is an antibody specifically recognizing the AXL protein.
- 다음의 단계를 포함하는 대사성 질환의 진단을 위한 정보제공방법:Information provision method for diagnosis of metabolic disease comprising the following steps:생물학적 시료에 존재하는 AXL 유전자의 발현량 또는 단백질의 양을 측정하는 단계; 및Measuring the amount of expression or protein of the AXL gene present in the biological sample; and상기 측정 결과를 정상 대조군 시료의 AXL 유전자의 발현량 또는 단백질의 양과 비교하는 단계.Comparing the measurement result with the amount of expression or protein of the AXL gene in the normal control sample.
- 제 12항에 있어서, 상기 대사성 질환은 고지방(high fat) 식이에 의해 유도되는 비알코올성 지방간인 것인, 대사성 질환의 진단을 위한 정보제공방법.The method of claim 12, wherein the metabolic disease is nonalcoholic fatty liver induced by a high fat diet.
- 제 12항에 있어서, 상기 생물학적 시료는 조직, 세포, 혈액, 혈청, 혈장, 타액 및 뇨로 이루어진 군으로부터 선택되는 것인, 대사성 질환의 진단을 위한 정보제공방법.The method of claim 12, wherein the biological sample is selected from the group consisting of tissue, cell, blood, serum, plasma, saliva and urine.
- 제 12항에 있어서, 상기 유전자의 발현 수준을 측정하는 방법은 역전사 중합효소반응(RT-PCR), 경쟁적 역전사 중합효소반응(Competitive RT-PCR), 실시간 역전사 중합효소반응(Realtime RT-PCR), RNase 보호 분석법(RPA; RNaseprotection assay), 노던 블롯팅(Northern blotting) 및 DNA 칩으로 이루어진 군으로부터 선택된 1개 이상의 방법인, 대사성 질환의 진단을 위한 정보제공방법.The method of claim 12, wherein the method for measuring the expression level of the gene is reverse transcription polymerase reaction (RT-PCR), competitive reverse transcription polymerase reaction (Competitive RT-PCR), real-time reverse transcription polymerase reaction (Realtime RT-PCR), RNase protection assay (RPA; RNaseprotection assay), Northern blotting (Northern blotting), and one or more methods selected from the group consisting of DNA chips, an information providing method for the diagnosis of metabolic diseases.
- 제 12항에 있어서, 상기 단백질의 발현 수준을 측정하기 위한 방법은 웨스턴 블롯, ELISA(enzyme linked immunosorbent assay), 방사선면역분석(RIA: Radioimmunoassay), 방사 면역 확산법(radioimmunodiffusion), 오우크테로니(Ouchterlony) 면역확산법, 로케트(rocket) 면역전기영동, 조직 면역 염색, 면역침전 분석법(Immunoprecipitation Assay), 보체 고정 분석법(Complement Fixation Assay), 유세포분석(Fluorescence Activated Cell Sorter, FACS) 및 단백질칩(protein chip)으로 이루어진 군으로부터 선택된 1개 이상의 방법인, 대사성 질환의 진단을 위한 정보제공방법.The method of claim 12, wherein the method for measuring the expression level of the protein is Western blot, ELISA (enzyme linked immunosorbent assay), radioimmunoassay (RIA: Radioimmunoassay), radioimmunodiffusion, Ouchterlony ) Immunodiffusion method, rocket immunoelectrophoresis, tissue immunostaining, immunoprecipitation assay, complement fixation assay, flow cytometry (Fluorescence Activated Cell Sorter, FACS) and protein chip One or more methods selected from the group consisting of, information providing method for the diagnosis of metabolic diseases.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020210167877A KR20230080560A (en) | 2021-11-30 | 2021-11-30 | Composition for preventing and treating fatty liver comprising AXL |
| KR10-2021-0167877 | 2021-11-30 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| WO2023101316A1 true WO2023101316A1 (en) | 2023-06-08 |
Family
ID=86612546
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| PCT/KR2022/018680 WO2023101316A1 (en) | 2021-11-30 | 2022-11-24 | Composition for preventing or treating steatosis, comprising axl |
Country Status (2)
| Country | Link |
|---|---|
| KR (1) | KR20230080560A (en) |
| WO (1) | WO2023101316A1 (en) |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030157573A1 (en) * | 2002-02-12 | 2003-08-21 | Orna Mor | Use of Axl receptor for diagnosis and treatment of renal disease |
| KR20170134641A (en) * | 2015-04-10 | 2017-12-06 | 샤카이후쿠시호진 온시자이단사이세이카이 | How to determine the condition of liver disease |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101925020B1 (en) | 2017-04-21 | 2018-12-04 | 연세대학교 산학협력단 | Pharmaceutical composition for metabolic disease comprising expression or activity inhibitor of MKRN1 as an active ingredient |
-
2021
- 2021-11-30 KR KR1020210167877A patent/KR20230080560A/en not_active Application Discontinuation
-
2022
- 2022-11-24 WO PCT/KR2022/018680 patent/WO2023101316A1/en unknown
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030157573A1 (en) * | 2002-02-12 | 2003-08-21 | Orna Mor | Use of Axl receptor for diagnosis and treatment of renal disease |
| KR20170134641A (en) * | 2015-04-10 | 2017-12-06 | 샤카이후쿠시호진 온시자이단사이세이카이 | How to determine the condition of liver disease |
Non-Patent Citations (4)
| Title |
|---|
| BHAVE SAYALI, HO HAN KIAT: "Exploring the Gamut of Receptor Tyrosine Kinases for Their Promise in the Management of Non-Alcoholic Fatty Liver Disease", BIOMEDICINES, vol. 9, no. 12, pages 1776, XP093070510, DOI: 10.3390/biomedicines9121776 * |
| LEE HYUN-JI: "The role and function ofAXL, a new target of obese fatty liver", CHUNGNAM NATIONAL UNIVERSITY HOSPITAL NEWSLETTER - HAPPY KEEPER, 1 January 2021 (2021-01-01), XP093070508, Retrieved from the Internet <URL:http://webzine.cnuh.co.kr/2021/07/sec3/study> * |
| TUTUSAUS ANNA, DE GREGORIO ESTEFANÍA, CUCARULL BLANCA, CRISTÓBAL HELENA, ARESTÉ CRISTINA, GRAUPERA ISABEL, COLL MAR, COLELL ANNA, : "A Functional Role of GAS6/TAM in Nonalcoholic Steatohepatitis Progression Implicates AXL as Therapeutic Target", CMGH CELLULAR AND MOLECULAR GASTROENTEROLOGY AND HEPATOLOGY, vol. 9, no. 3, 1 January 2020 (2020-01-01), pages 349 - 368, XP093070513, ISSN: 2352-345X, DOI: 10.1016/j.jcmgh.2019.10.010 * |
| ZHU CHENJING, WEI YUQUAN, WEI XIAWEI: "AXL receptor tyrosine kinase as a promising anti-cancer approach: functions, molecular mechanisms and clinical applications", MOLECULAR CANCER, BMC, vol. 18, no. 1, 1 December 2019 (2019-12-01), pages 153, XP093013311, DOI: 10.1186/s12943-019-1090-3 * |
Also Published As
| Publication number | Publication date |
|---|---|
| KR20230080560A (en) | 2023-06-07 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Li et al. | Leptin regulates exon-specific transcription of the Bdnf gene via epigenetic modifications mediated by an AKT/p300 HAT cascade | |
| US20110236897A1 (en) | Novel markers for diagnosing brain disease caused by brain injury and use thereof | |
| US6406853B1 (en) | Interventions to mimic the effects of calorie restriction | |
| US20160115540A1 (en) | Prodrugs and drugs | |
| WO2017217807A9 (en) | Biomarker comprising nckap1 as effective ingredient for diagnosing colorectal cancer or for predicting metastasis and prognosis of colorectal cancer | |
| KR20110014979A (en) | Antiviral therapy | |
| WO2017026843A1 (en) | Method for providing information on chronic myeloid leukemia | |
| US20030124540A1 (en) | Interventions to mimic the effects of calorie restriction | |
| WO2022019604A1 (en) | Biomarker specific for liver cancer, and use thereof | |
| WO2018105921A2 (en) | Use of leucine-zipper protein for diagnosis or treatment of fatty liver | |
| WO2023101316A1 (en) | Composition for preventing or treating steatosis, comprising axl | |
| WO2018004240A1 (en) | Use of nupr1 in diagnosis and treatment of brain tumor | |
| WO2017026691A1 (en) | Composition for diagnosing obesity and uses therefor | |
| WO2022025387A1 (en) | Biomarker for diagnosing nonalcoholic steatohepatitis using microrna combination | |
| WO2018048257A1 (en) | Use of akap12 as hepatic fibrosis diagnostic marker | |
| KR101611004B1 (en) | Composition for diagnosis of leukemia using TrkA | |
| WO2019103456A2 (en) | Biomarker composition for diagnosing radiation-resistant cancer or for predicting prognosis of radiation therapy containing pmvk as active ingredient | |
| WO2020138561A1 (en) | Method for providing information for diagnosis of meningitis | |
| US8895011B2 (en) | Insulin-resistance-improving drug | |
| WO2018155948A1 (en) | Marker for prediction of survival prognosis of lung cancer patient | |
| WO2021101257A1 (en) | Composition for diagnosing, preventing, or treating cognitive dysfunction comprising cotl1 as active ingredient | |
| WO2023287079A1 (en) | Biomarker composition for predicting triple negative breast cancer metastasis, containing csde1 as active ingredient | |
| WO2019045432A1 (en) | Biomarker for squamous cell carcinoma of lung and method for diagnosing squamous cell carcinoma of lung by using same | |
| WO2024147473A1 (en) | Composition for diagnosis or prognosis prediction of non-alcoholic fatty liver disease comprising lipocalin-2 gene or protein and diagnosis method using same | |
| KR102202120B1 (en) | Use of Ube2h for Diagnosis or Treatment of Alzheimer's Disease |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| 121 | Ep: the epo has been informed by wipo that ep was designated in this application |
Ref document number: 22901674 Country of ref document: EP Kind code of ref document: A1 |
|
| NENP | Non-entry into the national phase |
Ref country code: DE |