WO2021001377A1 - cPLA2e INDUCING AGENTS AND USES THEREOF - Google Patents
cPLA2e INDUCING AGENTS AND USES THEREOF Download PDFInfo
- Publication number
- WO2021001377A1 WO2021001377A1 PCT/EP2020/068414 EP2020068414W WO2021001377A1 WO 2021001377 A1 WO2021001377 A1 WO 2021001377A1 EP 2020068414 W EP2020068414 W EP 2020068414W WO 2021001377 A1 WO2021001377 A1 WO 2021001377A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- cpla2e
- nucleic acid
- acid construct
- vector
- promoter
- Prior art date
Links
- 230000001939 inductive effect Effects 0.000 title claims abstract description 42
- 208000010877 cognitive disease Diseases 0.000 claims abstract description 88
- 208000024827 Alzheimer disease Diseases 0.000 claims abstract description 44
- 238000011282 treatment Methods 0.000 claims abstract description 42
- 239000003795 chemical substances by application Substances 0.000 claims abstract description 37
- 206010012289 Dementia Diseases 0.000 claims abstract description 26
- 239000003814 drug Substances 0.000 claims abstract description 15
- 102100040625 Cytosolic phospholipase A2 epsilon Human genes 0.000 claims description 190
- 150000007523 nucleic acids Chemical class 0.000 claims description 153
- 102000039446 nucleic acids Human genes 0.000 claims description 149
- 108020004707 nucleic acids Proteins 0.000 claims description 149
- 108010082107 Group IV Phospholipases A2 Proteins 0.000 claims description 142
- 239000002245 particle Substances 0.000 claims description 87
- 108090000623 proteins and genes Proteins 0.000 claims description 85
- 230000003612 virological effect Effects 0.000 claims description 83
- 239000013598 vector Substances 0.000 claims description 72
- 239000002773 nucleotide Substances 0.000 claims description 64
- 125000003729 nucleotide group Chemical group 0.000 claims description 64
- 241000702423 Adeno-associated virus - 2 Species 0.000 claims description 48
- 102000004169 proteins and genes Human genes 0.000 claims description 35
- 241000282414 Homo sapiens Species 0.000 claims description 33
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 33
- 238000000034 method Methods 0.000 claims description 32
- 239000008194 pharmaceutical composition Substances 0.000 claims description 32
- 230000000694 effects Effects 0.000 claims description 29
- 230000008488 polyadenylation Effects 0.000 claims description 27
- 201000010099 disease Diseases 0.000 claims description 25
- 238000004806 packaging method and process Methods 0.000 claims description 23
- 239000013603 viral vector Substances 0.000 claims description 20
- 108090000565 Capsid Proteins Proteins 0.000 claims description 17
- 102100023321 Ceruloplasmin Human genes 0.000 claims description 17
- 230000001537 neural effect Effects 0.000 claims description 16
- 108010076504 Protein Sorting Signals Proteins 0.000 claims description 13
- 150000001875 compounds Chemical class 0.000 claims description 11
- 239000013607 AAV vector Substances 0.000 claims description 10
- 241000702421 Dependoparvovirus Species 0.000 claims description 10
- 239000001963 growth medium Substances 0.000 claims description 9
- 241001634120 Adeno-associated virus - 5 Species 0.000 claims description 8
- 239000000546 pharmaceutical excipient Substances 0.000 claims description 6
- 230000008569 process Effects 0.000 claims description 6
- 239000003937 drug carrier Substances 0.000 claims description 5
- 108010006025 bovine growth hormone Proteins 0.000 claims description 4
- 230000006698 induction Effects 0.000 claims description 4
- 238000004113 cell culture Methods 0.000 claims description 3
- 239000012228 culture supernatant Substances 0.000 claims description 3
- 238000012258 culturing Methods 0.000 claims description 3
- 238000003306 harvesting Methods 0.000 claims description 3
- 238000003556 assay Methods 0.000 claims description 2
- 102100021244 Integral membrane protein GPR180 Human genes 0.000 claims 6
- 101000821100 Homo sapiens Synapsin-1 Proteins 0.000 claims 2
- 102100021905 Synapsin-1 Human genes 0.000 claims 2
- 210000004027 cell Anatomy 0.000 description 116
- 241000699670 Mus sp. Species 0.000 description 70
- DZHSAHHDTRWUTF-SIQRNXPUSA-N amyloid-beta polypeptide 42 Chemical compound C([C@@H](C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(=O)NCC(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCCN)C(=O)NCC(=O)N[C@@H](C)C(=O)N[C@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](C(C)C)C(=O)NCC(=O)NCC(=O)N[C@@H](C(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(O)=O)[C@@H](C)CC)C(C)C)NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@@H](NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC=1N=CNC=1)NC(=O)[C@H](CC=1N=CNC=1)NC(=O)[C@@H](NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)CNC(=O)[C@H](CO)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC=1N=CNC=1)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC(O)=O)C(C)C)C(C)C)C1=CC=CC=C1 DZHSAHHDTRWUTF-SIQRNXPUSA-N 0.000 description 43
- 101710137189 Amyloid-beta A4 protein Proteins 0.000 description 42
- 101710151993 Amyloid-beta precursor protein Proteins 0.000 description 42
- 102100022704 Amyloid-beta precursor protein Human genes 0.000 description 42
- 101000614110 Homo sapiens Cytosolic phospholipase A2 epsilon Proteins 0.000 description 41
- 230000014509 gene expression Effects 0.000 description 40
- 238000012360 testing method Methods 0.000 description 36
- 239000013604 expression vector Substances 0.000 description 33
- 208000028698 Cognitive impairment Diseases 0.000 description 26
- 108090000765 processed proteins & peptides Proteins 0.000 description 24
- 241000700605 Viruses Species 0.000 description 22
- 230000015654 memory Effects 0.000 description 22
- 239000012634 fragment Substances 0.000 description 21
- 108020004999 messenger RNA Proteins 0.000 description 20
- 210000004556 brain Anatomy 0.000 description 18
- 210000001320 hippocampus Anatomy 0.000 description 18
- 210000002569 neuron Anatomy 0.000 description 18
- 239000013612 plasmid Substances 0.000 description 18
- 229920001184 polypeptide Polymers 0.000 description 18
- 102000004196 processed proteins & peptides Human genes 0.000 description 18
- 108091023045 Untranslated Region Proteins 0.000 description 16
- 241001465754 Metazoa Species 0.000 description 15
- 102000017299 Synapsin-1 Human genes 0.000 description 13
- 108050005241 Synapsin-1 Proteins 0.000 description 13
- 230000000971 hippocampal effect Effects 0.000 description 13
- 238000013518 transcription Methods 0.000 description 13
- 230000035897 transcription Effects 0.000 description 13
- 108020005345 3' Untranslated Regions Proteins 0.000 description 12
- 108700019146 Transgenes Proteins 0.000 description 12
- 230000013016 learning Effects 0.000 description 12
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 11
- 102000040430 polynucleotide Human genes 0.000 description 11
- 108091033319 polynucleotide Proteins 0.000 description 11
- 239000002157 polynucleotide Substances 0.000 description 11
- 230000001105 regulatory effect Effects 0.000 description 11
- 239000000523 sample Substances 0.000 description 11
- 108020003589 5' Untranslated Regions Proteins 0.000 description 10
- 238000011740 C57BL/6 mouse Methods 0.000 description 10
- 241000699666 Mus <mouse, genus> Species 0.000 description 10
- 102000001708 Protein Isoforms Human genes 0.000 description 10
- 108010029485 Protein Isoforms Proteins 0.000 description 10
- 238000004519 manufacturing process Methods 0.000 description 10
- 230000006386 memory function Effects 0.000 description 10
- 230000010076 replication Effects 0.000 description 10
- 230000001225 therapeutic effect Effects 0.000 description 10
- 210000001519 tissue Anatomy 0.000 description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 10
- IYGYMKDQCDOMRE-QRWMCTBCSA-N Bicculine Chemical compound O([C@H]1C2C3=CC=4OCOC=4C=C3CCN2C)C(=O)C2=C1C=CC1=C2OCO1 IYGYMKDQCDOMRE-QRWMCTBCSA-N 0.000 description 9
- 108091026890 Coding region Proteins 0.000 description 9
- AACMFFIUYXGCOC-UHFFFAOYSA-N bicuculline Natural products CN1CCc2cc3OCOc3cc2C1C4OCc5c6OCOc6ccc45 AACMFFIUYXGCOC-UHFFFAOYSA-N 0.000 description 9
- 238000004422 calculation algorithm Methods 0.000 description 9
- IYGYMKDQCDOMRE-UHFFFAOYSA-N d-Bicucullin Natural products CN1CCC2=CC=3OCOC=3C=C2C1C1OC(=O)C2=C1C=CC1=C2OCO1 IYGYMKDQCDOMRE-UHFFFAOYSA-N 0.000 description 9
- 230000014759 maintenance of location Effects 0.000 description 9
- 239000000203 mixture Substances 0.000 description 9
- 239000000047 product Substances 0.000 description 9
- 230000006886 spatial memory Effects 0.000 description 9
- 238000002672 stereotactic surgery Methods 0.000 description 9
- 101710138820 Cytosolic phospholipase A2 epsilon Proteins 0.000 description 8
- 230000001419 dependent effect Effects 0.000 description 8
- 230000006870 function Effects 0.000 description 8
- 238000001415 gene therapy Methods 0.000 description 8
- 206010027175 memory impairment Diseases 0.000 description 8
- 238000010149 post-hoc-test Methods 0.000 description 8
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 7
- 108020004414 DNA Proteins 0.000 description 7
- 239000000872 buffer Substances 0.000 description 7
- 208000035475 disorder Diseases 0.000 description 7
- 238000002347 injection Methods 0.000 description 7
- 239000007924 injection Substances 0.000 description 7
- 230000002018 overexpression Effects 0.000 description 7
- 239000000243 solution Substances 0.000 description 7
- 230000000946 synaptic effect Effects 0.000 description 7
- 241000271566 Aves Species 0.000 description 6
- 241001529936 Murinae Species 0.000 description 6
- 102000001435 Synapsin Human genes 0.000 description 6
- 108050009621 Synapsin Proteins 0.000 description 6
- 230000015572 biosynthetic process Effects 0.000 description 6
- 230000006735 deficit Effects 0.000 description 6
- -1 glycol nucleic acids Chemical class 0.000 description 6
- 238000003119 immunoblot Methods 0.000 description 6
- 230000001965 increasing effect Effects 0.000 description 6
- 230000005764 inhibitory process Effects 0.000 description 6
- 230000001404 mediated effect Effects 0.000 description 6
- 238000001543 one-way ANOVA Methods 0.000 description 6
- 239000000126 substance Substances 0.000 description 6
- 238000012549 training Methods 0.000 description 6
- 238000010175 APPswe/PSEN1dE9 Methods 0.000 description 5
- 102000004190 Enzymes Human genes 0.000 description 5
- 108090000790 Enzymes Proteins 0.000 description 5
- 241000282412 Homo Species 0.000 description 5
- 108091028043 Nucleic acid sequence Proteins 0.000 description 5
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 5
- 230000004913 activation Effects 0.000 description 5
- 150000001413 amino acids Chemical class 0.000 description 5
- 230000006399 behavior Effects 0.000 description 5
- 210000000234 capsid Anatomy 0.000 description 5
- 230000010261 cell growth Effects 0.000 description 5
- 230000003750 conditioning effect Effects 0.000 description 5
- 210000001787 dendrite Anatomy 0.000 description 5
- 238000000338 in vitro Methods 0.000 description 5
- 208000015181 infectious disease Diseases 0.000 description 5
- 210000004962 mammalian cell Anatomy 0.000 description 5
- 208000027061 mild cognitive impairment Diseases 0.000 description 5
- 210000000056 organ Anatomy 0.000 description 5
- 239000004055 small Interfering RNA Substances 0.000 description 5
- 208000024891 symptom Diseases 0.000 description 5
- 241000701447 unidentified baculovirus Species 0.000 description 5
- 208000014644 Brain disease Diseases 0.000 description 4
- 102000005636 Cyclic AMP Response Element-Binding Protein Human genes 0.000 description 4
- 108010045171 Cyclic AMP Response Element-Binding Protein Proteins 0.000 description 4
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 4
- 241000829100 Macaca mulatta polyomavirus 1 Species 0.000 description 4
- 208000026139 Memory disease Diseases 0.000 description 4
- 241000283973 Oryctolagus cuniculus Species 0.000 description 4
- 208000018737 Parkinson disease Diseases 0.000 description 4
- 108091034057 RNA (poly(A)) Proteins 0.000 description 4
- 108700005077 Viral Genes Proteins 0.000 description 4
- 206010002026 amyotrophic lateral sclerosis Diseases 0.000 description 4
- 238000004458 analytical method Methods 0.000 description 4
- 239000011575 calcium Substances 0.000 description 4
- 230000019771 cognition Effects 0.000 description 4
- 210000003520 dendritic spine Anatomy 0.000 description 4
- 239000003623 enhancer Substances 0.000 description 4
- 239000000284 extract Substances 0.000 description 4
- 230000008014 freezing Effects 0.000 description 4
- 238000007710 freezing Methods 0.000 description 4
- 239000004615 ingredient Substances 0.000 description 4
- 230000027928 long-term synaptic potentiation Effects 0.000 description 4
- 239000002609 medium Substances 0.000 description 4
- 208000015122 neurodegenerative disease Diseases 0.000 description 4
- 230000000750 progressive effect Effects 0.000 description 4
- 210000001176 projection neuron Anatomy 0.000 description 4
- 238000011160 research Methods 0.000 description 4
- 230000004044 response Effects 0.000 description 4
- 241000894007 species Species 0.000 description 4
- 230000014616 translation Effects 0.000 description 4
- 238000011144 upstream manufacturing Methods 0.000 description 4
- 208000000044 Amnesia Diseases 0.000 description 3
- 241000272525 Anas platyrhynchos Species 0.000 description 3
- 241000894006 Bacteria Species 0.000 description 3
- 241000283690 Bos taurus Species 0.000 description 3
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 3
- 241000282472 Canis lupus familiaris Species 0.000 description 3
- 108700028146 Genetic Enhancer Elements Proteins 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 241000238631 Hexapoda Species 0.000 description 3
- 102100037611 Lysophospholipase Human genes 0.000 description 3
- 108020002496 Lysophospholipase Proteins 0.000 description 3
- 108700011259 MicroRNAs Proteins 0.000 description 3
- 238000012347 Morris Water Maze Methods 0.000 description 3
- 241000699660 Mus musculus Species 0.000 description 3
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 description 3
- 208000028017 Psychotic disease Diseases 0.000 description 3
- 108020004459 Small interfering RNA Proteins 0.000 description 3
- 230000002159 abnormal effect Effects 0.000 description 3
- 229910052791 calcium Inorganic materials 0.000 description 3
- 210000003169 central nervous system Anatomy 0.000 description 3
- 238000003776 cleavage reaction Methods 0.000 description 3
- 238000012217 deletion Methods 0.000 description 3
- 230000037430 deletion Effects 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 230000018109 developmental process Effects 0.000 description 3
- 238000001378 electrochemiluminescence detection Methods 0.000 description 3
- 230000006390 fear memory Effects 0.000 description 3
- 239000000499 gel Substances 0.000 description 3
- 230000001976 improved effect Effects 0.000 description 3
- 230000002401 inhibitory effect Effects 0.000 description 3
- 208000014674 injury Diseases 0.000 description 3
- 238000003780 insertion Methods 0.000 description 3
- 230000037431 insertion Effects 0.000 description 3
- 238000007913 intrathecal administration Methods 0.000 description 3
- 150000002632 lipids Chemical class 0.000 description 3
- 239000012528 membrane Substances 0.000 description 3
- 239000002679 microRNA Substances 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 238000012986 modification Methods 0.000 description 3
- 239000013642 negative control Substances 0.000 description 3
- 210000002682 neurofibrillary tangle Anatomy 0.000 description 3
- 238000011002 quantification Methods 0.000 description 3
- 238000003753 real-time PCR Methods 0.000 description 3
- 230000007017 scission Effects 0.000 description 3
- 210000003625 skull Anatomy 0.000 description 3
- 235000000346 sugar Nutrition 0.000 description 3
- 230000008685 targeting Effects 0.000 description 3
- 230000002103 transcriptional effect Effects 0.000 description 3
- 230000032258 transport Effects 0.000 description 3
- SYSZENVIJHPFNL-UHFFFAOYSA-N (alpha-D-mannosyl)7-beta-D-mannosyl-diacetylchitobiosyl-L-asparagine, isoform B (protein) Chemical compound COC1=CC=C(I)C=C1 SYSZENVIJHPFNL-UHFFFAOYSA-N 0.000 description 2
- CPKVUHPKYQGHMW-UHFFFAOYSA-N 1-ethenylpyrrolidin-2-one;molecular iodine Chemical compound II.C=CN1CCCC1=O CPKVUHPKYQGHMW-UHFFFAOYSA-N 0.000 description 2
- ZIIUUSVHCHPIQD-UHFFFAOYSA-N 2,4,6-trimethyl-N-[3-(trifluoromethyl)phenyl]benzenesulfonamide Chemical compound CC1=CC(C)=CC(C)=C1S(=O)(=O)NC1=CC=CC(C(F)(F)F)=C1 ZIIUUSVHCHPIQD-UHFFFAOYSA-N 0.000 description 2
- 241001655883 Adeno-associated virus - 1 Species 0.000 description 2
- 241000580270 Adeno-associated virus - 4 Species 0.000 description 2
- 241000972680 Adeno-associated virus - 6 Species 0.000 description 2
- 241001164823 Adeno-associated virus - 7 Species 0.000 description 2
- 241001164825 Adeno-associated virus - 8 Species 0.000 description 2
- 241000649045 Adeno-associated virus 10 Species 0.000 description 2
- 241000649046 Adeno-associated virus 11 Species 0.000 description 2
- 241000649047 Adeno-associated virus 12 Species 0.000 description 2
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 2
- 208000037259 Amyloid Plaque Diseases 0.000 description 2
- 241000713704 Bovine immunodeficiency virus Species 0.000 description 2
- 241000282465 Canis Species 0.000 description 2
- 102000005853 Clathrin Human genes 0.000 description 2
- 108010019874 Clathrin Proteins 0.000 description 2
- 208000010859 Creutzfeldt-Jakob disease Diseases 0.000 description 2
- 241000699800 Cricetinae Species 0.000 description 2
- 230000004543 DNA replication Effects 0.000 description 2
- 206010052804 Drug tolerance Diseases 0.000 description 2
- 241000713730 Equine infectious anemia virus Species 0.000 description 2
- 241000283073 Equus caballus Species 0.000 description 2
- 241000713800 Feline immunodeficiency virus Species 0.000 description 2
- 201000011240 Frontotemporal dementia Diseases 0.000 description 2
- 241000287828 Gallus gallus Species 0.000 description 2
- JZNWSCPGTDBMEW-UHFFFAOYSA-N Glycerophosphorylethanolamin Natural products NCCOP(O)(=O)OCC(O)CO JZNWSCPGTDBMEW-UHFFFAOYSA-N 0.000 description 2
- 108091065273 Group IV family Proteins 0.000 description 2
- NYHBQMYGNKIUIF-UUOKFMHZSA-N Guanosine Chemical group C1=NC=2C(=O)NC(N)=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O NYHBQMYGNKIUIF-UUOKFMHZSA-N 0.000 description 2
- 241001135569 Human adenovirus 5 Species 0.000 description 2
- 241000725303 Human immunodeficiency virus Species 0.000 description 2
- 208000023105 Huntington disease Diseases 0.000 description 2
- 108091092195 Intron Proteins 0.000 description 2
- 208000009829 Lewy Body Disease Diseases 0.000 description 2
- 201000002832 Lewy body dementia Diseases 0.000 description 2
- 108010072388 Methyl-CpG-Binding Protein 2 Proteins 0.000 description 2
- 102100039124 Methyl-CpG-binding protein 2 Human genes 0.000 description 2
- 241000713869 Moloney murine leukemia virus Species 0.000 description 2
- 102000004868 N-Methyl-D-Aspartate Receptors Human genes 0.000 description 2
- 108090001041 N-Methyl-D-Aspartate Receptors Proteins 0.000 description 2
- 108091093037 Peptide nucleic acid Proteins 0.000 description 2
- 241000286209 Phasianidae Species 0.000 description 2
- 102000015439 Phospholipases Human genes 0.000 description 2
- 108010064785 Phospholipases Proteins 0.000 description 2
- 102000012288 Phosphopyruvate Hydratase Human genes 0.000 description 2
- 108010022181 Phosphopyruvate Hydratase Proteins 0.000 description 2
- 108091036407 Polyadenylation Proteins 0.000 description 2
- 201000010769 Prader-Willi syndrome Diseases 0.000 description 2
- 108091000106 RNA cap binding Proteins 0.000 description 2
- 102000028391 RNA cap binding Human genes 0.000 description 2
- 241000700159 Rattus Species 0.000 description 2
- 241000283984 Rodentia Species 0.000 description 2
- 206010039966 Senile dementia Diseases 0.000 description 2
- 241000713311 Simian immunodeficiency virus Species 0.000 description 2
- 108091027967 Small hairpin RNA Proteins 0.000 description 2
- 208000006011 Stroke Diseases 0.000 description 2
- RYYWUUFWQRZTIU-UHFFFAOYSA-N Thiophosphoric acid Chemical class OP(O)(S)=O RYYWUUFWQRZTIU-UHFFFAOYSA-N 0.000 description 2
- 108091036066 Three prime untranslated region Proteins 0.000 description 2
- 108091046915 Threose nucleic acid Proteins 0.000 description 2
- 108700009124 Transcription Initiation Site Proteins 0.000 description 2
- 108091023040 Transcription factor Proteins 0.000 description 2
- 102000040945 Transcription factor Human genes 0.000 description 2
- 108010067390 Viral Proteins Proteins 0.000 description 2
- 208000027418 Wounds and injury Diseases 0.000 description 2
- 230000001464 adherent effect Effects 0.000 description 2
- 230000032683 aging Effects 0.000 description 2
- 125000000539 amino acid group Chemical group 0.000 description 2
- 229960000723 ampicillin Drugs 0.000 description 2
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 2
- 230000003542 behavioural effect Effects 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 239000007975 buffered saline Substances 0.000 description 2
- 230000001413 cellular effect Effects 0.000 description 2
- 229930193282 clathrin Natural products 0.000 description 2
- 230000003920 cognitive function Effects 0.000 description 2
- 238000007596 consolidation process Methods 0.000 description 2
- 238000010276 construction Methods 0.000 description 2
- 238000007796 conventional method Methods 0.000 description 2
- 238000003745 diagnosis Methods 0.000 description 2
- 230000029087 digestion Effects 0.000 description 2
- 229940042399 direct acting antivirals protease inhibitors Drugs 0.000 description 2
- 239000012153 distilled water Substances 0.000 description 2
- 210000001671 embryonic stem cell Anatomy 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 230000002708 enhancing effect Effects 0.000 description 2
- 238000000605 extraction Methods 0.000 description 2
- 108020001507 fusion proteins Proteins 0.000 description 2
- 102000037865 fusion proteins Human genes 0.000 description 2
- 230000002068 genetic effect Effects 0.000 description 2
- 150000002327 glycerophospholipids Chemical class 0.000 description 2
- 230000026781 habituation Effects 0.000 description 2
- 238000001727 in vivo Methods 0.000 description 2
- 230000010354 integration Effects 0.000 description 2
- 238000007912 intraperitoneal administration Methods 0.000 description 2
- 238000001990 intravenous administration Methods 0.000 description 2
- PGHMRUGBZOYCAA-UHFFFAOYSA-N ionomycin Natural products O1C(CC(O)C(C)C(O)C(C)C=CCC(C)CC(C)C(O)=CC(=O)C(C)CC(C)CC(CCC(O)=O)C)CCC1(C)C1OC(C)(C(C)O)CC1 PGHMRUGBZOYCAA-UHFFFAOYSA-N 0.000 description 2
- PGHMRUGBZOYCAA-ADZNBVRBSA-N ionomycin Chemical compound O1[C@H](C[C@H](O)[C@H](C)[C@H](O)[C@H](C)/C=C/C[C@@H](C)C[C@@H](C)C(/O)=C/C(=O)[C@@H](C)C[C@@H](C)C[C@@H](CCC(O)=O)C)CC[C@@]1(C)[C@@H]1O[C@](C)([C@@H](C)O)CC1 PGHMRUGBZOYCAA-ADZNBVRBSA-N 0.000 description 2
- 230000008449 language Effects 0.000 description 2
- 238000011031 large-scale manufacturing process Methods 0.000 description 2
- 230000007786 learning performance Effects 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 230000004807 localization Effects 0.000 description 2
- 238000012423 maintenance Methods 0.000 description 2
- 230000006984 memory degeneration Effects 0.000 description 2
- 208000023060 memory loss Diseases 0.000 description 2
- 230000007334 memory performance Effects 0.000 description 2
- 230000010387 memory retrieval Effects 0.000 description 2
- YACKEPLHDIMKIO-UHFFFAOYSA-N methylphosphonic acid Chemical class CP(O)(O)=O YACKEPLHDIMKIO-UHFFFAOYSA-N 0.000 description 2
- 239000007758 minimum essential medium Substances 0.000 description 2
- 230000035772 mutation Effects 0.000 description 2
- 238000002610 neuroimaging Methods 0.000 description 2
- 230000030147 nuclear export Effects 0.000 description 2
- 230000037361 pathway Effects 0.000 description 2
- 239000000137 peptide hydrolase inhibitor Substances 0.000 description 2
- 150000008104 phosphatidylethanolamines Chemical class 0.000 description 2
- 229940067605 phosphatidylethanolamines Drugs 0.000 description 2
- 230000026731 phosphorylation Effects 0.000 description 2
- 238000006366 phosphorylation reaction Methods 0.000 description 2
- 230000004911 positive regulation of CREB transcription factor activity Effects 0.000 description 2
- KMUONIBRACKNSN-UHFFFAOYSA-N potassium dichromate Chemical compound [K+].[K+].[O-][Cr](=O)(=O)O[Cr]([O-])(=O)=O KMUONIBRACKNSN-UHFFFAOYSA-N 0.000 description 2
- 239000002243 precursor Substances 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 230000000979 retarding effect Effects 0.000 description 2
- 238000012552 review Methods 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 230000007596 spatial working memory Effects 0.000 description 2
- 239000003381 stabilizer Substances 0.000 description 2
- 238000010186 staining Methods 0.000 description 2
- 238000006467 substitution reaction Methods 0.000 description 2
- 150000008163 sugars Chemical class 0.000 description 2
- 238000001356 surgical procedure Methods 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 210000000225 synapse Anatomy 0.000 description 2
- 230000003956 synaptic plasticity Effects 0.000 description 2
- 229940124597 therapeutic agent Drugs 0.000 description 2
- 238000002560 therapeutic procedure Methods 0.000 description 2
- 238000012546 transfer Methods 0.000 description 2
- 238000011830 transgenic mouse model Methods 0.000 description 2
- 238000013519 translation Methods 0.000 description 2
- 230000008733 trauma Effects 0.000 description 2
- 210000005239 tubule Anatomy 0.000 description 2
- 238000007492 two-way ANOVA Methods 0.000 description 2
- 230000003827 upregulation Effects 0.000 description 2
- 239000003981 vehicle Substances 0.000 description 2
- 210000002845 virion Anatomy 0.000 description 2
- 230000003936 working memory Effects 0.000 description 2
- FFTVPQUHLQBXQZ-KVUCHLLUSA-N (4s,4as,5ar,12ar)-4,7-bis(dimethylamino)-1,10,11,12a-tetrahydroxy-3,12-dioxo-4a,5,5a,6-tetrahydro-4h-tetracene-2-carboxamide Chemical compound C1C2=C(N(C)C)C=CC(O)=C2C(O)=C2[C@@H]1C[C@H]1[C@H](N(C)C)C(=O)C(C(N)=O)=C(O)[C@@]1(O)C2=O FFTVPQUHLQBXQZ-KVUCHLLUSA-N 0.000 description 1
- KILNVBDSWZSGLL-KXQOOQHDSA-N 1,2-dihexadecanoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCCCCCCCCC KILNVBDSWZSGLL-KXQOOQHDSA-N 0.000 description 1
- QLOKJRIVRGCVIM-UHFFFAOYSA-N 1-[(4-methylsulfanylphenyl)methyl]piperazine Chemical compound C1=CC(SC)=CC=C1CN1CCNCC1 QLOKJRIVRGCVIM-UHFFFAOYSA-N 0.000 description 1
- UVBYMVOUBXYSFV-XUTVFYLZSA-N 1-methylpseudouridine Chemical compound O=C1NC(=O)N(C)C=C1[C@H]1[C@H](O)[C@H](O)[C@@H](CO)O1 UVBYMVOUBXYSFV-XUTVFYLZSA-N 0.000 description 1
- UVBYMVOUBXYSFV-UHFFFAOYSA-N 1-methylpseudouridine Natural products O=C1NC(=O)N(C)C=C1C1C(O)C(O)C(CO)O1 UVBYMVOUBXYSFV-UHFFFAOYSA-N 0.000 description 1
- IIZPXYDJLKNOIY-JXPKJXOSSA-N 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCC\C=C/C\C=C/C\C=C/C\C=C/CCCCC IIZPXYDJLKNOIY-JXPKJXOSSA-N 0.000 description 1
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 1
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 1
- 239000003140 4 aminobutyric acid A receptor blocking agent Substances 0.000 description 1
- KELXHQACBIUYSE-UHFFFAOYSA-N 5-methoxy-1h-pyrimidine-2,4-dione Chemical compound COC1=CNC(=O)NC1=O KELXHQACBIUYSE-UHFFFAOYSA-N 0.000 description 1
- 108091027075 5S-rRNA precursor Proteins 0.000 description 1
- 102000003678 AMPA Receptors Human genes 0.000 description 1
- 108090000078 AMPA Receptors Proteins 0.000 description 1
- 229930024421 Adenine Natural products 0.000 description 1
- GFFGJBXGBJISGV-UHFFFAOYSA-N Adenine Chemical compound NC1=NC=NC2=C1N=CN2 GFFGJBXGBJISGV-UHFFFAOYSA-N 0.000 description 1
- 102000002260 Alkaline Phosphatase Human genes 0.000 description 1
- 108020004774 Alkaline Phosphatase Proteins 0.000 description 1
- 238000010173 Alzheimer-disease mouse model Methods 0.000 description 1
- 208000009575 Angelman syndrome Diseases 0.000 description 1
- 241000272814 Anser sp. Species 0.000 description 1
- 241000153246 Anteros Species 0.000 description 1
- 208000019901 Anxiety disease Diseases 0.000 description 1
- 208000006096 Attention Deficit Disorder with Hyperactivity Diseases 0.000 description 1
- 208000036864 Attention deficit/hyperactivity disease Diseases 0.000 description 1
- 238000009020 BCA Protein Assay Kit Methods 0.000 description 1
- 101001074429 Bacillus subtilis (strain 168) Polyketide biosynthesis acyltransferase homolog PksD Proteins 0.000 description 1
- 101000936617 Bacillus velezensis (strain DSM 23117 / BGSC 10A6 / FZB42) Polyketide biosynthesis acyltransferase homolog BaeD Proteins 0.000 description 1
- 201000004569 Blindness Diseases 0.000 description 1
- 241000701822 Bovine papillomavirus Species 0.000 description 1
- 206010006187 Breast cancer Diseases 0.000 description 1
- 101100043731 Caenorhabditis elegans syx-3 gene Proteins 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 101150044789 Cap gene Proteins 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- 241000700198 Cavia Species 0.000 description 1
- 102000000844 Cell Surface Receptors Human genes 0.000 description 1
- 108010001857 Cell Surface Receptors Proteins 0.000 description 1
- 241000282693 Cercopithecidae Species 0.000 description 1
- 206010008111 Cerebral haemorrhage Diseases 0.000 description 1
- 208000000094 Chronic Pain Diseases 0.000 description 1
- 101100007328 Cocos nucifera COS-1 gene Proteins 0.000 description 1
- 108020004705 Codon Proteins 0.000 description 1
- 206010010254 Concussion Diseases 0.000 description 1
- 206010010356 Congenital anomaly Diseases 0.000 description 1
- 108091035707 Consensus sequence Proteins 0.000 description 1
- 208000011990 Corticobasal Degeneration Diseases 0.000 description 1
- 208000020406 Creutzfeldt Jacob disease Diseases 0.000 description 1
- 208000003407 Creutzfeldt-Jakob Syndrome Diseases 0.000 description 1
- MIKUYHXYGGJMLM-GIMIYPNGSA-N Crotonoside Natural products C1=NC2=C(N)NC(=O)N=C2N1[C@H]1O[C@@H](CO)[C@H](O)[C@@H]1O MIKUYHXYGGJMLM-GIMIYPNGSA-N 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
- NYHBQMYGNKIUIF-UHFFFAOYSA-N D-guanosine Natural products C1=2NC(N)=NC(=O)C=2N=CN1C1OC(CO)C(O)C1O NYHBQMYGNKIUIF-UHFFFAOYSA-N 0.000 description 1
- 102000053602 DNA Human genes 0.000 description 1
- 102000012410 DNA Ligases Human genes 0.000 description 1
- 108010061982 DNA Ligases Proteins 0.000 description 1
- 241000238557 Decapoda Species 0.000 description 1
- 201000010374 Down Syndrome Diseases 0.000 description 1
- 101100535673 Drosophila melanogaster Syn gene Proteins 0.000 description 1
- 206010013654 Drug abuse Diseases 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 241000283086 Equidae Species 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 101710091918 Eukaryotic translation initiation factor 4E Proteins 0.000 description 1
- 102100027304 Eukaryotic translation initiation factor 4E Human genes 0.000 description 1
- 101710126428 Eukaryotic translation initiation factor 4E-2 Proteins 0.000 description 1
- 101710126416 Eukaryotic translation initiation factor 4E-3 Proteins 0.000 description 1
- 101710126432 Eukaryotic translation initiation factor 4E1 Proteins 0.000 description 1
- 101710133325 Eukaryotic translation initiation factor NCBP Proteins 0.000 description 1
- 101710190212 Eukaryotic translation initiation factor isoform 4E Proteins 0.000 description 1
- 101710124729 Eukaryotic translation initiation factor isoform 4E-2 Proteins 0.000 description 1
- 241000282326 Felis catus Species 0.000 description 1
- 208000001914 Fragile X syndrome Diseases 0.000 description 1
- 208000002339 Frontotemporal Lobar Degeneration Diseases 0.000 description 1
- 229940090502 GABA A receptor antagonist Drugs 0.000 description 1
- 102000004300 GABA-A Receptors Human genes 0.000 description 1
- 108090000839 GABA-A Receptors Proteins 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Chemical compound OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 108010051696 Growth Hormone Proteins 0.000 description 1
- 241000696272 Gull adenovirus Species 0.000 description 1
- 241000713858 Harvey murine sarcoma virus Species 0.000 description 1
- 241001272567 Hominoidea Species 0.000 description 1
- 101000823051 Homo sapiens Amyloid-beta precursor protein Proteins 0.000 description 1
- 101001067140 Homo sapiens Porphobilinogen deaminase Proteins 0.000 description 1
- 101000617536 Homo sapiens Presenilin-1 Proteins 0.000 description 1
- 241000701044 Human gammaherpesvirus 4 Species 0.000 description 1
- 102000004157 Hydrolases Human genes 0.000 description 1
- 108090000604 Hydrolases Proteins 0.000 description 1
- 101150008942 J gene Proteins 0.000 description 1
- YQEZLKZALYSWHR-UHFFFAOYSA-N Ketamine Chemical compound C=1C=CC=C(Cl)C=1C1(NC)CCCCC1=O YQEZLKZALYSWHR-UHFFFAOYSA-N 0.000 description 1
- 239000012741 Laemmli sample buffer Substances 0.000 description 1
- 241000713666 Lentivirus Species 0.000 description 1
- 101710125418 Major capsid protein Proteins 0.000 description 1
- 208000002720 Malnutrition Diseases 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- 201000009906 Meningitis Diseases 0.000 description 1
- 208000019022 Mood disease Diseases 0.000 description 1
- 101001037757 Mus musculus Heat shock 70 kDa protein 1A Proteins 0.000 description 1
- 101100368134 Mus musculus Syn1 gene Proteins 0.000 description 1
- HZFDKBPTVOENNB-GAFUQQFSSA-N N-[(2S)-1-[2-[(2R)-2-chloro-2-fluoroacetyl]-2-[[(3S)-2-oxopyrrolidin-3-yl]methyl]hydrazinyl]-3-(1-methylcyclopropyl)-1-oxopropan-2-yl]-5-(difluoromethyl)-1,2-oxazole-3-carboxamide Chemical compound CC1(C[C@@H](C(NN(C[C@H](CCN2)C2=O)C([C@H](F)Cl)=O)=O)NC(C2=NOC(C(F)F)=C2)=O)CC1 HZFDKBPTVOENNB-GAFUQQFSSA-N 0.000 description 1
- 239000000020 Nitrocellulose Substances 0.000 description 1
- 240000008881 Oenanthe javanica Species 0.000 description 1
- 108091034117 Oligonucleotide Proteins 0.000 description 1
- 108700026244 Open Reading Frames Proteins 0.000 description 1
- 101150112919 PLA2G4E gene Proteins 0.000 description 1
- 208000002193 Pain Diseases 0.000 description 1
- 108091081548 Palindromic sequence Proteins 0.000 description 1
- 241000282579 Pan Species 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 208000006735 Periostitis Diseases 0.000 description 1
- 241000577979 Peromyscus spicilegus Species 0.000 description 1
- 108091000080 Phosphotransferase Proteins 0.000 description 1
- 208000000609 Pick Disease of the Brain Diseases 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 229920001213 Polysorbate 20 Polymers 0.000 description 1
- 241000282405 Pongo abelii Species 0.000 description 1
- 102100034391 Porphobilinogen deaminase Human genes 0.000 description 1
- 229920000153 Povidone-iodine Polymers 0.000 description 1
- 102100022033 Presenilin-1 Human genes 0.000 description 1
- 241000288906 Primates Species 0.000 description 1
- 102000029797 Prion Human genes 0.000 description 1
- 108091000054 Prion Proteins 0.000 description 1
- 208000024777 Prion disease Diseases 0.000 description 1
- 108010007568 Protamines Proteins 0.000 description 1
- 108091030071 RNAI Proteins 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 108020004511 Recombinant DNA Proteins 0.000 description 1
- 108010034634 Repressor Proteins Proteins 0.000 description 1
- PYMYPHUHKUWMLA-LMVFSUKVSA-N Ribose Natural products OC[C@@H](O)[C@@H](O)[C@@H](O)C=O PYMYPHUHKUWMLA-LMVFSUKVSA-N 0.000 description 1
- 241000714474 Rous sarcoma virus Species 0.000 description 1
- 108091081021 Sense strand Proteins 0.000 description 1
- 206010040047 Sepsis Diseases 0.000 description 1
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 1
- 108020004682 Single-Stranded DNA Proteins 0.000 description 1
- 208000010340 Sleep Deprivation Diseases 0.000 description 1
- UIIMBOGNXHQVGW-DEQYMQKBSA-M Sodium bicarbonate-14C Chemical compound [Na+].O[14C]([O-])=O UIIMBOGNXHQVGW-DEQYMQKBSA-M 0.000 description 1
- 208000033043 Somatoform and factitious disease Diseases 0.000 description 1
- 102100038803 Somatotropin Human genes 0.000 description 1
- 241000713880 Spleen focus-forming virus Species 0.000 description 1
- 108091081024 Start codon Proteins 0.000 description 1
- 208000037132 Subdural Chronic Hematoma Diseases 0.000 description 1
- 208000002667 Subdural Hematoma Diseases 0.000 description 1
- 231100000643 Substance intoxication Toxicity 0.000 description 1
- 241000282887 Suidae Species 0.000 description 1
- 102100038126 Tenascin Human genes 0.000 description 1
- 108010008125 Tenascin Proteins 0.000 description 1
- 239000004098 Tetracycline Substances 0.000 description 1
- 102000006601 Thymidine Kinase Human genes 0.000 description 1
- 108020004440 Thymidine kinase Proteins 0.000 description 1
- 206010070863 Toxicity to various agents Diseases 0.000 description 1
- 239000007983 Tris buffer Substances 0.000 description 1
- 102000004243 Tubulin Human genes 0.000 description 1
- 108090000704 Tubulin Proteins 0.000 description 1
- 108091034131 VA RNA Proteins 0.000 description 1
- 241000700618 Vaccinia virus Species 0.000 description 1
- 230000001594 aberrant effect Effects 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 230000036982 action potential Effects 0.000 description 1
- 239000012190 activator Substances 0.000 description 1
- 108091006088 activator proteins Proteins 0.000 description 1
- 230000001154 acute effect Effects 0.000 description 1
- 125000002252 acyl group Chemical group 0.000 description 1
- 229960000643 adenine Drugs 0.000 description 1
- 239000002671 adjuvant Substances 0.000 description 1
- 239000011543 agarose gel Substances 0.000 description 1
- HMFHBZSHGGEWLO-UHFFFAOYSA-N alpha-D-Furanose-Ribose Natural products OCC1OC(O)C(O)C1O HMFHBZSHGGEWLO-UHFFFAOYSA-N 0.000 description 1
- VREFGVBLTWBCJP-UHFFFAOYSA-N alprazolam Chemical compound C12=CC(Cl)=CC=C2N2C(C)=NN=C2CN=C1C1=CC=CC=C1 VREFGVBLTWBCJP-UHFFFAOYSA-N 0.000 description 1
- 208000020466 alteration of consciousness Diseases 0.000 description 1
- 230000001668 ameliorated effect Effects 0.000 description 1
- 229910021529 ammonia Inorganic materials 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 206010002022 amyloidosis Diseases 0.000 description 1
- 230000000202 analgesic effect Effects 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 239000012914 anti-clumping agent Substances 0.000 description 1
- 230000000692 anti-sense effect Effects 0.000 description 1
- 239000002518 antifoaming agent Substances 0.000 description 1
- 210000004507 artificial chromosome Anatomy 0.000 description 1
- 238000013528 artificial neural network Methods 0.000 description 1
- 230000035045 associative learning Effects 0.000 description 1
- 208000015802 attention deficit-hyperactivity disease Diseases 0.000 description 1
- 208000029560 autism spectrum disease Diseases 0.000 description 1
- 239000007640 basal medium Substances 0.000 description 1
- 238000009227 behaviour therapy Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 229940064804 betadine Drugs 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 239000012472 biological sample Substances 0.000 description 1
- 239000000090 biomarker Substances 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 230000003925 brain function Effects 0.000 description 1
- 239000008366 buffered solution Substances 0.000 description 1
- RMRJXGBAOAMLHD-IHFGGWKQSA-N buprenorphine Chemical compound C([C@]12[C@H]3OC=4C(O)=CC=C(C2=4)C[C@@H]2[C@]11CC[C@]3([C@H](C1)[C@](C)(O)C(C)(C)C)OC)CN2CC1CC1 RMRJXGBAOAMLHD-IHFGGWKQSA-N 0.000 description 1
- 229960001736 buprenorphine Drugs 0.000 description 1
- UAIXRPCCYXNJMQ-RZIPZOSSSA-N buprenorphine hydrochlorie Chemical compound [Cl-].C([C@]12[C@H]3OC=4C(O)=CC=C(C2=4)C[C@@H]2[C@]11CC[C@]3([C@H](C1)[C@](C)(O)C(C)(C)C)OC)C[NH+]2CC1CC1 UAIXRPCCYXNJMQ-RZIPZOSSSA-N 0.000 description 1
- 210000004899 c-terminal region Anatomy 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 235000011148 calcium chloride Nutrition 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 235000014633 carbohydrates Nutrition 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 210000005056 cell body Anatomy 0.000 description 1
- 239000013592 cell lysate Substances 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 238000002659 cell therapy Methods 0.000 description 1
- 230000005754 cellular signaling Effects 0.000 description 1
- 210000003710 cerebral cortex Anatomy 0.000 description 1
- 210000001175 cerebrospinal fluid Anatomy 0.000 description 1
- 208000013677 cerebrovascular dementia Diseases 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 210000004978 chinese hamster ovary cell Anatomy 0.000 description 1
- RCTYPNKXASFOBE-UHFFFAOYSA-M chloromercury Chemical compound [Hg]Cl RCTYPNKXASFOBE-UHFFFAOYSA-M 0.000 description 1
- 230000001684 chronic effect Effects 0.000 description 1
- 238000011260 co-administration Methods 0.000 description 1
- 238000012761 co-transfection Methods 0.000 description 1
- 230000035071 co-translational protein modification Effects 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 230000003930 cognitive ability Effects 0.000 description 1
- 230000003931 cognitive performance Effects 0.000 description 1
- 208000030251 communication disease Diseases 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 239000002299 complementary DNA Substances 0.000 description 1
- 230000009514 concussion Effects 0.000 description 1
- 230000001143 conditioned effect Effects 0.000 description 1
- 229910000366 copper(II) sulfate Inorganic materials 0.000 description 1
- 230000001054 cortical effect Effects 0.000 description 1
- 125000004122 cyclic group Chemical group 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 230000003412 degenerative effect Effects 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 230000003111 delayed effect Effects 0.000 description 1
- 238000001739 density measurement Methods 0.000 description 1
- 230000008021 deposition Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 230000000249 desinfective effect Effects 0.000 description 1
- 230000006866 deterioration Effects 0.000 description 1
- 230000001627 detrimental effect Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000009792 diffusion process Methods 0.000 description 1
- 239000003085 diluting agent Substances 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 239000002612 dispersion medium Substances 0.000 description 1
- 208000018459 dissociative disease Diseases 0.000 description 1
- NAGJZTKCGNOGPW-UHFFFAOYSA-N dithiophosphoric acid Chemical class OP(O)(S)=S NAGJZTKCGNOGPW-UHFFFAOYSA-N 0.000 description 1
- 229940052760 dopamine agonists Drugs 0.000 description 1
- 239000003136 dopamine receptor stimulating agent Substances 0.000 description 1
- 238000012377 drug delivery Methods 0.000 description 1
- 230000009977 dual effect Effects 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 230000002996 emotional effect Effects 0.000 description 1
- 206010014599 encephalitis Diseases 0.000 description 1
- 230000002121 endocytic effect Effects 0.000 description 1
- 230000002616 endonucleolytic effect Effects 0.000 description 1
- 230000008029 eradication Effects 0.000 description 1
- LYCAIKOWRPUZTN-UHFFFAOYSA-N ethylene glycol Natural products OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 1
- 210000003527 eukaryotic cell Anatomy 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 230000000763 evoking effect Effects 0.000 description 1
- 230000002964 excitative effect Effects 0.000 description 1
- 230000010326 executive functioning Effects 0.000 description 1
- 238000013401 experimental design Methods 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 230000001605 fetal effect Effects 0.000 description 1
- 238000010304 firing Methods 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 230000037406 food intake Effects 0.000 description 1
- 235000012631 food intake Nutrition 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 230000009368 gene silencing by RNA Effects 0.000 description 1
- 238000007429 general method Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 239000003102 growth factor Substances 0.000 description 1
- 239000000122 growth hormone Substances 0.000 description 1
- 229940029575 guanosine Drugs 0.000 description 1
- RQFCJASXJCIDSX-UUOKFMHZSA-N guanosine 5'-monophosphate Chemical compound C1=2NC(N)=NC(=O)C=2N=CN1[C@@H]1O[C@H](COP(O)(O)=O)[C@@H](O)[C@H]1O RQFCJASXJCIDSX-UUOKFMHZSA-N 0.000 description 1
- 210000003128 head Anatomy 0.000 description 1
- 230000008801 hippocampal function Effects 0.000 description 1
- 210000004295 hippocampal neuron Anatomy 0.000 description 1
- 230000004694 hippocampus damage Effects 0.000 description 1
- 102000046783 human APP Human genes 0.000 description 1
- 210000005260 human cell Anatomy 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- WGCNASOHLSPBMP-UHFFFAOYSA-N hydroxyacetaldehyde Natural products OCC=O WGCNASOHLSPBMP-UHFFFAOYSA-N 0.000 description 1
- 238000000126 in silico method Methods 0.000 description 1
- 238000000099 in vitro assay Methods 0.000 description 1
- 208000035231 inattentive type attention deficit hyperactivity disease Diseases 0.000 description 1
- 239000000411 inducer Substances 0.000 description 1
- 230000001524 infective effect Effects 0.000 description 1
- 230000010365 information processing Effects 0.000 description 1
- 230000000977 initiatory effect Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 208000020658 intracerebral hemorrhage Diseases 0.000 description 1
- 238000000185 intracerebroventricular administration Methods 0.000 description 1
- NBQNWMBBSKPBAY-UHFFFAOYSA-N iodixanol Chemical compound IC=1C(C(=O)NCC(O)CO)=C(I)C(C(=O)NCC(O)CO)=C(I)C=1N(C(=O)C)CC(O)CN(C(C)=O)C1=C(I)C(C(=O)NCC(O)CO)=C(I)C(C(=O)NCC(O)CO)=C1I NBQNWMBBSKPBAY-UHFFFAOYSA-N 0.000 description 1
- 229960004359 iodixanol Drugs 0.000 description 1
- 229910000359 iron(II) sulfate Inorganic materials 0.000 description 1
- VCJMYUPGQJHHFU-UHFFFAOYSA-N iron(III) nitrate Inorganic materials [Fe+3].[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O VCJMYUPGQJHHFU-UHFFFAOYSA-N 0.000 description 1
- 230000002427 irreversible effect Effects 0.000 description 1
- 239000007951 isotonicity adjuster Substances 0.000 description 1
- 229960003299 ketamine Drugs 0.000 description 1
- 206010023497 kuru Diseases 0.000 description 1
- 239000000787 lecithin Substances 0.000 description 1
- 229940067606 lecithin Drugs 0.000 description 1
- 235000010445 lecithin Nutrition 0.000 description 1
- 230000003902 lesion Effects 0.000 description 1
- 230000000670 limiting effect Effects 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 238000001294 liquid chromatography-tandem mass spectrometry Methods 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 230000003137 locomotive effect Effects 0.000 description 1
- 230000007787 long-term memory Effects 0.000 description 1
- 231100000863 loss of memory Toxicity 0.000 description 1
- 208000018769 loss of vision Diseases 0.000 description 1
- 231100000864 loss of vision Toxicity 0.000 description 1
- 210000005265 lung cell Anatomy 0.000 description 1
- 239000006166 lysate Substances 0.000 description 1
- 239000012139 lysis buffer Substances 0.000 description 1
- 230000001071 malnutrition Effects 0.000 description 1
- 235000000824 malnutrition Nutrition 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 238000004949 mass spectrometry Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 230000007595 memory recall Effects 0.000 description 1
- 230000003340 mental effect Effects 0.000 description 1
- 238000007884 metabolite profiling Methods 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 239000004530 micro-emulsion Substances 0.000 description 1
- 239000011859 microparticle Substances 0.000 description 1
- 235000013336 milk Nutrition 0.000 description 1
- 239000008267 milk Substances 0.000 description 1
- 210000004080 milk Anatomy 0.000 description 1
- 229960004023 minocycline Drugs 0.000 description 1
- 238000009126 molecular therapy Methods 0.000 description 1
- 125000004573 morpholin-4-yl group Chemical group N1(CCOCC1)* 0.000 description 1
- 108700011845 mouse Hb9 Proteins 0.000 description 1
- 238000010172 mouse model Methods 0.000 description 1
- 201000006417 multiple sclerosis Diseases 0.000 description 1
- 239000002105 nanoparticle Substances 0.000 description 1
- 230000004770 neurodegeneration Effects 0.000 description 1
- 230000000626 neurodegenerative effect Effects 0.000 description 1
- 210000004498 neuroglial cell Anatomy 0.000 description 1
- 230000000926 neurological effect Effects 0.000 description 1
- 230000008555 neuronal activation Effects 0.000 description 1
- 230000007996 neuronal plasticity Effects 0.000 description 1
- 230000003557 neuropsychological effect Effects 0.000 description 1
- 238000010855 neuropsychological testing Methods 0.000 description 1
- 229920001220 nitrocellulos Polymers 0.000 description 1
- 231100000252 nontoxic Toxicity 0.000 description 1
- 230000003000 nontoxic effect Effects 0.000 description 1
- 201000003077 normal pressure hydrocephalus Diseases 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 208000015380 nutritional deficiency disease Diseases 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 239000005022 packaging material Substances 0.000 description 1
- 239000003973 paint Substances 0.000 description 1
- 238000007911 parenteral administration Methods 0.000 description 1
- 230000007170 pathology Effects 0.000 description 1
- 210000003460 periosteum Anatomy 0.000 description 1
- 210000001428 peripheral nervous system Anatomy 0.000 description 1
- 150000008298 phosphoramidates Chemical class 0.000 description 1
- 102000020233 phosphotransferase Human genes 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920003023 plastic Polymers 0.000 description 1
- 238000007747 plating Methods 0.000 description 1
- 102000028499 poly(A) binding Human genes 0.000 description 1
- 108091023021 poly(A) binding Proteins 0.000 description 1
- 229920002401 polyacrylamide Polymers 0.000 description 1
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 229920005862 polyol Polymers 0.000 description 1
- 150000003077 polyols Chemical class 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 230000004481 post-translational protein modification Effects 0.000 description 1
- 229960001621 povidone-iodine Drugs 0.000 description 1
- 230000003518 presynaptic effect Effects 0.000 description 1
- 210000002243 primary neuron Anatomy 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 201000002212 progressive supranuclear palsy Diseases 0.000 description 1
- 230000004845 protein aggregation Effects 0.000 description 1
- 108020001580 protein domains Proteins 0.000 description 1
- 238000001243 protein synthesis Methods 0.000 description 1
- 208000020016 psychiatric disease Diseases 0.000 description 1
- 229940124811 psychiatric drug Drugs 0.000 description 1
- 210000002763 pyramidal cell Anatomy 0.000 description 1
- ZAHRKKWIAAJSAO-UHFFFAOYSA-N rapamycin Natural products COCC(O)C(=C/C(C)C(=O)CC(OC(=O)C1CCCCN1C(=O)C(=O)C2(O)OC(CC(OC)C(=CC=CC=CC(C)CC(C)C(=O)C)C)CCC2C)C(C)CC3CCC(O)C(C3)OC)C ZAHRKKWIAAJSAO-UHFFFAOYSA-N 0.000 description 1
- 239000011541 reaction mixture Substances 0.000 description 1
- 230000008707 rearrangement Effects 0.000 description 1
- 229940044551 receptor antagonist Drugs 0.000 description 1
- 239000002464 receptor antagonist Substances 0.000 description 1
- 102000005962 receptors Human genes 0.000 description 1
- 108020003175 receptors Proteins 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 230000002829 reductive effect Effects 0.000 description 1
- 230000002787 reinforcement Effects 0.000 description 1
- 101150066583 rep gene Proteins 0.000 description 1
- 210000001525 retina Anatomy 0.000 description 1
- 230000002441 reversible effect Effects 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 201000000980 schizophrenia Diseases 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000035807 sensation Effects 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 208000013223 septicemia Diseases 0.000 description 1
- 230000006403 short-term memory Effects 0.000 description 1
- 230000011664 signaling Effects 0.000 description 1
- QFJCIRLUMZQUOT-HPLJOQBZSA-N sirolimus Chemical compound C1C[C@@H](O)[C@H](OC)C[C@@H]1C[C@@H](C)[C@H]1OC(=O)[C@@H]2CCCCN2C(=O)C(=O)[C@](O)(O2)[C@H](C)CC[C@H]2C[C@H](OC)/C(C)=C/C=C/C=C/[C@@H](C)C[C@@H](C)C(=O)[C@H](OC)[C@H](O)/C(C)=C/[C@@H](C)C(=O)C1 QFJCIRLUMZQUOT-HPLJOQBZSA-N 0.000 description 1
- 229960002930 sirolimus Drugs 0.000 description 1
- 150000003384 small molecules Chemical class 0.000 description 1
- AKHNMLFCWUSKQB-UHFFFAOYSA-L sodium thiosulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=S AKHNMLFCWUSKQB-UHFFFAOYSA-L 0.000 description 1
- 235000019345 sodium thiosulphate Nutrition 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 239000000600 sorbitol Substances 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 230000035882 stress Effects 0.000 description 1
- 238000007920 subcutaneous administration Methods 0.000 description 1
- 208000011117 substance-related disease Diseases 0.000 description 1
- 150000005846 sugar alcohols Polymers 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 230000002459 sustained effect Effects 0.000 description 1
- 230000009182 swimming Effects 0.000 description 1
- 230000009782 synaptic response Effects 0.000 description 1
- 230000005062 synaptic transmission Effects 0.000 description 1
- 230000009885 systemic effect Effects 0.000 description 1
- 230000002123 temporal effect Effects 0.000 description 1
- 229960002180 tetracycline Drugs 0.000 description 1
- 229930101283 tetracycline Natural products 0.000 description 1
- 235000019364 tetracycline Nutrition 0.000 description 1
- 150000003522 tetracyclines Chemical class 0.000 description 1
- 230000004797 therapeutic response Effects 0.000 description 1
- 230000005100 tissue tropism Effects 0.000 description 1
- 238000004448 titration Methods 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 239000011573 trace mineral Substances 0.000 description 1
- 235000013619 trace mineral Nutrition 0.000 description 1
- 230000005030 transcription termination Effects 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- 230000001052 transient effect Effects 0.000 description 1
- 230000017105 transposition Effects 0.000 description 1
- 239000001226 triphosphate Substances 0.000 description 1
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 1
- 241000701161 unidentified adenovirus Species 0.000 description 1
- 241001529453 unidentified herpesvirus Species 0.000 description 1
- 229960005486 vaccine Drugs 0.000 description 1
- 210000003501 vero cell Anatomy 0.000 description 1
- 230000029812 viral genome replication Effects 0.000 description 1
- 230000000007 visual effect Effects 0.000 description 1
- 230000004393 visual impairment Effects 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 230000002747 voluntary effect Effects 0.000 description 1
- 210000002268 wool Anatomy 0.000 description 1
- BPICBUSOMSTKRF-UHFFFAOYSA-N xylazine Chemical compound CC1=CC=CC(C)=C1NC1=NCCCS1 BPICBUSOMSTKRF-UHFFFAOYSA-N 0.000 description 1
- 229960001600 xylazine Drugs 0.000 description 1
- 229910000368 zinc sulfate Inorganic materials 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
- A61K48/0075—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the delivery route, e.g. oral, subcutaneous
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells
- C12N15/86—Viral vectors
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
- C12N9/18—Carboxylic ester hydrolases (3.1.1)
- C12N9/20—Triglyceride splitting, e.g. by means of lipase
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2217/00—Genetically modified animals
- A01K2217/07—Animals genetically altered by homologous recombination
- A01K2217/072—Animals genetically altered by homologous recombination maintaining or altering function, i.e. knock in
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2227/00—Animals characterised by species
- A01K2227/10—Mammal
- A01K2227/105—Murine
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2267/00—Animals characterised by purpose
- A01K2267/03—Animal model, e.g. for test or diseases
- A01K2267/0306—Animal model for genetic diseases
- A01K2267/0312—Animal model for Alzheimer's disease
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2750/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssDNA viruses
- C12N2750/00011—Details
- C12N2750/14011—Parvoviridae
- C12N2750/14111—Dependovirus, e.g. adenoassociated viruses
- C12N2750/14141—Use of virus, viral particle or viral elements as a vector
- C12N2750/14143—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y301/00—Hydrolases acting on ester bonds (3.1)
- C12Y301/01—Carboxylic ester hydrolases (3.1.1)
- C12Y301/01004—Phospholipase A2 (3.1.1.4)
Definitions
- the present invention relates to cPLA2e inducing agents and cPLA2e inducing agents for use as a medicament, more particularly for use in the treatment of a cognitive disorder and/or disease associated with a cognitive disorder, for example dementia, and more specifically age-related dementia and/or Alzheimer’s disease.
- a cognitive disorder and/or disease associated with a cognitive disorder for example dementia, and more specifically age-related dementia and/or Alzheimer’s disease.
- Mild cognitive impairment is characterized by deficits in memory, language and/or other essential cognitive functions that do not interfere with an individual’s daily life.
- the condition often evolves towards dementia, which is characterized by a global deterioration of cognitive abilities to an extent that does interfere with daily life.
- AD Alzheimer’s disease
- NFTs neurofibrillary tangles
- Cognitive impairment is a condition associated with a large number of brain disorders.
- Brain disorders can have many causes, e.g., degenerative conditions, heredity, trauma, infection, malnutrition and others.
- cognitive impairment can be associated with aging and/or neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, Huntington's disease, Amyotrophic Lateral Sclerosis (ALS), psychosis, Parkinson's disease psychosis, Alzheimer's disease psychosis, Lewy-body dementia, prionic neurodegenerative disorders such as Creutzfeld- Jacob disease and kuru disease, corticobasal degeneration, frontotemporal lobar degeneration, multiple sclerosis, normal pressure hydrocephalus, organic chronic brain syndrome, Pick's disease, progressive supranuclear palsy, or senile dementia.
- neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, Huntington's disease, Amyotrophic Lateral Sclerosis (ALS), psychosis, Parkinson's disease psychosis, Alzheimer's disease psychosis,
- Cognitive impairment may also have a congenital basis, e.g., Prader-Willi syndrome, Down Syndrome, Fragile X Syndrome, Angelman syndrome and autism spectrum disorder. Cognitive impairment may also be associated with trauma to the brain, such as that caused by chronic subdural hematoma, concussion, stroke, intracerebral hemorrhage, or with other injury to the brain, such as that caused by infection (e.g., encephalitis, meningitis, and septicemia) or drug intoxication or abuse.
- infection e.g., encephalitis, meningitis, and septicemia
- Cognitive impairment may also be associated with other conditions which impair or otherwise affect normal functioning of the central nervous system, including sleep deprivation, psychiatric disorders such as anxiety disorders, dissociative disorders, mood disorders, schizophrenia, treatment with psychiatric medications, treatment with dopamine agonists and somatoform and factitious disorders; it may also be associated with conditions of the peripheral nervous system, such as chronic pain. In some cases, the cause of a cognitive impairment may be unknown or uncertain.
- Cognitive impairment can be manifest in many ways, e.g., deficits in learning and/or memory including, but not limited to, attention, information acquisition, information processing, working memory, short-term memory, long-term memory, anterograde memory, retrograde memory, memory retrieval, discrimination learning, decision-making, language retrieval, inhibitory response control, attentional set-shifting, delayed reinforcement learning, reversal learning, the temporal integration of voluntary behavior, and expressing an interest in one's surroundings and self-care.
- Cognitive impairment may be characterized by progressive loss of memory, cognition, reasoning, executive functioning, planning, judgment and emotional stability
- hippocampal PLA2G4E also named cytosolic phospholipase A2 epsilon (cPLA2e) mediated by treatment with AAV2/9-mPLA2G4E (a viral vector encoding cPLA2e)
- cPLA2e cytosolic phospholipase A2 epsilon
- AAV2/9-mPLA2G4E a viral vector encoding cPLA2e
- the invention relates to a nucleic acid construct that comprises a nucleotide sequence encoding a cytosolic phospholipase A2 epsilon (cPLA2e).
- the cPLA2e is a human cPLA2e; typically human cPLA2e of SEQ ID NO: 1 or SEQ ID NO:3, or a variant human cPLA2e having at least 70%, sequence identity with respect to human cPLA2e SEQ ID NO: 1 or SEQ ID NO:3.
- the nucleotide sequence encoding cPLA2e is SEQ ID NO:2 or SEQ ID NO:4.
- said nucleic acid construct further comprises a promoter operably-linked to the nucleotide sequence encoding a cPLA2e.
- the promoter operably- linked to the nucleotide sequence encoding a cPLA2e is a neuronal-specific promoter; notably said promoter is a SYN1 promoter or hybrid SYN1 promoter.
- said nucleic acid construct further comprises a polyadenylation signal sequence; notably a polyadenylation signal sequence of bovine growth hormone gene.
- said nucleic acid construct comprises a 5'ITR and a 3’ITR sequences; preferably a 5'ITR and a 3 'ITR sequences of an adeno-associated virus, more preferably a 5’ITR and a 3’ITR sequences from the AAV2 serotype.
- the nucleic acid construct of the invention is an RNA, notably an mRNA.
- the invention relates to a vector that comprises a nucleic acid construct of the invention; preferably said vector is a viral vector; more preferably an AAV vector.
- the invention relates to a viral particle that includes a nucleic acid construct of the invention.
- said viral particle is selected among AAV particles, preferably including capsid proteins selected from the group consisting of AAV2, AAV5, AAV9, and AAV EE serotypes.
- the invention also relates to a host cell comprising a nucleic acid construct or an expression vector of the invention.
- the invention relates to a process for producing viral particles comprising:
- the invention in another aspect, relates to a pharmaceutical composition
- a pharmaceutical composition comprising a nucleic acid construct, a vector, or a viral particle, or a host cell of the invention; and a pharmaceutically acceptable carrier or excipient.
- the invention relates to a nucleic acid construct, a vector, a viral particle, or host cell of the invention, or to a pharmaceutical composition comprising said nucleic acid construct, vector, viral particle or host cell for use as a medicament.
- the invention relates to a cPLA2e inducing agent for use as a medicament.
- the invention relates to a cPLA2e inducing agent for use in the treatment of cognitive disorders and/or diseases associated with cognitive disorders in a subject in need thereof.
- said disease associated with cognitive disorders is dementia.
- said disease associated with cognitive disorders is an age-related dementia or an Alzheimer’s disease.
- said cPLA2e inducing agent is selected from the group consisting of: a) a nucleic acid construct of the invention; b) a vector comprising a nucleic acid construct of the invention; c) a viral particle comprising a nucleic acid construct or vector of the invention; d) a host cell comprising a nucleic acid construct or vector of the invention; e) a cPLA2e polypeptide or protein; and f) a pharmaceutical composition comprising any of said nucleic acid construct, vector comprising the nucleic acid construct, viral particle comprising the nucleic acid construct or vector, and cPLA2e polypeptide or protein.
- said cPLA2e inducing agent is a nucleic acid construct, a vector or a viral particle of the invention, or a pharmaceutical composition comprising said nucleic acid, vector or viral particle.
- said cPLA2e inducing agent is a protein with cPLA2e activity; preferably a protein of SEQ ID NO: 1 or SEQ ID NO:3.
- Figure 1A Escape latency to the hidden-platform in the MWM test for elderly WT (negative control), APP/PSl sham (sham-injected APP/PSl mice) and APP/PSl AAV2/9- mPLA2G4E (APP/PSl mice treated with AAV2/9-mPLA2G4E) two months after stereotactic surgery.
- Figure IB Percentage of time spent in the correct quadrant during the 15s and 60s probe trials on day 6th for elderly WT, APP/PSl sham and APP/PSl AAV2/9-mPLA2G4E two months after hippocampal injections.
- Figure 4A Diagram showing the experimental design of the Fear Conditioning paradigm used to elucidate the role of PLA2G4E in memory function. Graph indicating the percentage of freezing behavior of TT mice during the training and test phase respectively
- the invention relates to a cytosolic phospholipase A2 epsilon inducing agent for use as a medicament, and more specifically for use in the treatment of a cognitive disorder and/or a disease associated with a cognitive disorder in a subject in need thereof.
- the terms“cytosolic phospholipase A2 epsilon”, and“phospholipase A2 group IVE”,“PLA2G4E”, or“cPLA2e” refer interchangeably to a Calcium-dependent enzyme member of the cytosolic phospholipase A2 group IV family that selectively hydrolyzes glycerophospholipids in the sn-2 position. Members of this family are involved in regulation of membrane tubule-mediated transport. This enzyme plays a role in trafficking through the clathrin-independent endocytic pathway. The enzyme regulates the recycling process via formation of tubules that transport internalized clathrin-independent cargo proteins back to the cell surface (Capestrano M.
- PLA2G4E can catalyze the calcium-dependent formation of N-acyl phosphatidylethanolamines (NAPEs) using phosphatidylethanolamine (PE) as the acyl chain donor (Ogura Y. et al. Nat Chem Biol. 2016; 12(9): 669-671).
- NAPEs N-acyl phosphatidylethanolamines
- PE phosphatidylethanolamine
- Human cPLA2e is naturally encoded by PLA2G4E gene; human cPLA2e is recorded for example at UniprotKB (https://www.uniprot.org/) with entry Accession number Q3MJ16.
- This entry describes 2 isoforms produced by alternative splicing: a) isoform 1 (with identifier: Q3MJ16-3) that is chosen as the 'canonical' sequence (SEQ ID NO: l); and b) isoform 2 (with identifier: Q3MJ16-2), that differs from the canonical sequence in that amino acids 1-376 are missing in isoform 2 (SEQ ID NO:3).
- the term“cPLA2e” refers to the enzyme and any additional co translation or post-translational modifications thereof.
- cPLA2e inducing agent refers to an agent (molecule or composition) that when administered to a cell, directly or indirectly produces a gaining of cPLA2e activity within the cell; particularly an agent that produces a gaining in expression of the enzyme cPLA2e, such as a cPLA2e transgene (i.e. a nucleotide sequence encoding a cPLA2e), or an expression product of said transgene.
- a cPLA2e transgene i.e. a nucleotide sequence encoding a cPLA2e
- the cPLA2 inducing agent for the use of the invention is or comprises a nucleic acid construct that comprises a nucleotide sequence encoding a cytosolic phospholipase A2 epsilon (cPLA2e).
- the invention relates to a nucleic acid construct that comprises a nucleotide sequence encoding a cytosolic phospholipase A2 epsilon (cPLA2e).
- nucleic acid and “polynucleotide” or “nucleotide sequence” are interchangeably used herein to refer to any molecule composed of or comprising monomeric nucleotides.
- a nucleic acid may be an oligonucleotide or a polynucleotide.
- a nucleotide sequence may be a DNA or RNA.
- a nucleotide sequence may be chemically modified or artificial. Nucleotide sequences include peptide nucleic acids (PNA), morpholinos and locked nucleic acids (LNA), as well as glycol nucleic acids (GNA) and threose nucleic acid (TNA).
- PNA peptide nucleic acids
- LNA locked nucleic acids
- GAA glycol nucleic acids
- TAA threose nucleic acid
- Each of these sequences is distinguished from naturally-occurring DNA or RNA by changes to the backbone of the molecule.
- phosphorothioate nucleotides may be used.
- Other deoxynucleotide analogs include methylphosphonates, phosphoramidates, phosphorodithioates, N3'P5'-phosphoramidates and oligoribonucleotide phosphorothioates and their 2'-0-allyl analogs and 2'-0-methylribonucleotide methylphosphonates which may be used in a nucleotide of the invention.
- nucleic acid construct refers to a non-naturally occurring nucleic acid resulting from the use of recombinant DNA technology.
- a nucleic acid construct is a nucleic acid molecule, either single- or double- stranded, which has been modified to contain segments of nucleic acid sequences, which are combined or juxtaposed in a manner which would not otherwise exist in nature.
- the nucleic acid construct of the invention comprises a nucleotide sequence encoding a naturally-occurring cPLA2e (wild type cPLA2e); e.g., a naturally-occurring human cPLA2e (e.g. iso form 1, or 2), a primate, murine or other mammalian known cPLA2e.
- the cPLA2e is a variant, a peptide or a polypeptide containing a substitution, and insertion and/or an addition, a deletion and/or a covalent modification with respect to a naturally-occurring cPLA2e, typically with respect to human cPLA2e isoform 1, or 2.
- cPLA2e encoded by the nucleic acid construct of the invention is a fusion protein or polypeptide in which some amino acids (e.g. tags) or polypeptides (e.g. a carrier polypeptide), can be added to the encoded cPLA2e (e.g., at the N-terminal or C-terminal ends), e.g., for localization or targeting.
- cPLA2e encoded by the nucleic acid construct is a fragment cPLA2e to which amino acid residues located at the carboxy, amino terminal, or internal regions of the cPLA2e, typically human cPLA2e isoform 1, or 2, can optionally be deleted.
- the nucleic acid construct of the present invention comprises a nucleotide sequence encoding a human cPLA2e, preferably a human cPLA2e of SEQ ID NO: l or SEQ ID NO:3, that correspond respectively to isoform 1 or 2; or a variant human cPLA2e having at least 70%, 75%, 80%, 85%, 90%; 95% or 99% sequence identity with respect to the coding sequence of a naturally-occurring or recombinant cPLA2e; typically with respect to human cPLA2e SEQ ID NO: 1 or SEQ ID NO:3.
- human cPLA2e iso form 1, or 2 protein fragments, functional protein domains, variants, and homologous proteins (orthologs) are also considered to be within the scope of the cPLA2e of the nucleic acid construct of the invention. It is understood the different embodiments of the cPLA2e have substantially the same cPLA2e activity as human cPLA2e isoform 1 or 2. Substantially the same activity can be for instance an activity of up to ⁇ 5%, including ⁇ 4, ⁇ 3, ⁇ 2%, ⁇ 1% or less.
- cPLA2e is a calcium-dependent enzyme member of the cytosolic phospholipase A2 group IV family that selectively hydrolyzes glycerophospholipids in the sn-2 position. It has been described to exhibit very low phospholipase (PLA) activity. Instead it has been shown to have a calcium-dependent N-acyltransferase (Ca-NAT) activity that produces N-acyl phosphatidylethanolamines (NAPEs) and N-acyl ethanolamines (NAEs) in mammalian cells. Its transacylase properties have been associated to a serine hydrolase activity (Ogura et al, Nat Chem Biol. 2016, 12(9), 669-671).
- Ca-NAT activity can be determined by measuring in a biological sample (e.g. cell lysate) the production of NAPEs. For instance, N-C16:0 DOPE production in reactions with DPPC (40 mM) and DOPE (75 mM) for 30 min at 37 °C with or without CaCl2 (3 mM) added to the reaction mixture. Ca-independent activities are substracted in the calculations of Ca- dependent activity.
- targeted analysis of NAPEs production e.g. 13 C16:0- containing NAPEs
- mammalian cells e.g. HEK293T cells
- the nucleotide sequence encoding a cPLA2e is SEQ ID NO:2 or SEQ ID NO:4; or a variant nucleotide sequence having at least 75%, at least 80%, at least 85%, at least 90% or at least 95% identity with respect to SEQ ID NO:2 or 4.
- sequence identity refers to the number of matches (identical nucleic acid residues or amino acid residues) in positions from an alignment of two polynucleotide sequences or two polypeptide sequences.
- sequence identity is determined by comparing the sequences when aligned so as to maximize overlap and identity while minimizing sequence gaps.
- sequence identity may be determined using any of a number of mathematical global or local alignment algorithms, depending on the length of the two sequences. Sequences of similar lengths are preferably aligned using a global alignment algorithm (e.g.
- Needleman and Wunsch algorithm Needleman and Wunsch, 1970, J Mol Biol.;48(3):443-53 which aligns the sequences optimally over the entire length, while sequences of substantially different lengths are preferably aligned using a local alignment algorithm (e.g. Smith and Waterman algorithm (Smith and Waterman, 1981, J Theor Biol. ;91(2):379-80) or Altschul algorithm (Altschul SF et al., 1997, Nucleic Acids Res.;25(17):3389-402; Altschul SF et al, 2005, Bioinformatics;21(8): 1451-6).
- a local alignment algorithm e.g. Smith and Waterman algorithm (Smith and Waterman, 1981, J Theor Biol. ;91(2):379-80) or Altschul algorithm (Altschul SF et al., 1997, Nucleic Acids Res.;25(17):3389-402; Altschul SF et al, 2005, Bioinformatics;21(8): 145
- the nucleic acid construct described herein may have different uses: among others, it may be used to generate a viral vector for gene therapy; or to generate a non-viral vector also for gene therapy, such as a nucleic acid construct with mRNA structure.
- the nucleic acid construct according to the present invention includes a nucleotide sequence encoding a cPLA2e and at least suitable nucleic acid elements for its expression in a host cell.
- the nucleic acid construct comprises a nucleotide sequence encoding a cPLA2e and one or more control sequence(s) required for expression of said coding sequence in the relevant target cell types or tissues.
- the nucleic acid construct comprises a coding sequence and regulatory sequences preceding (5' non-coding sequences) and following (3' non-coding sequences) the coding sequence that are required for expression of the selected gene product.
- said nucleic acid construct comprises at least (i) a nucleotide sequence encoding a cPLA2e under the control of (ii) a promoter and (iii) a 3' untranslated region that usually contains a polyadenylation signal sequence and/or transcription terminator.
- the nucleic acid construct may also comprise additional regulatory elements such as, for example, enhancer sequences, introns, microRNA targeted sequence, a polylinker sequence facilitating the insertion of a DNA fragment within a vector and/or splicing signal sequences.
- the nucleic acid construct of the invention also comprises a promoter. Said promoter initiates transgene expression upon introduction into a host cell.
- transgene refers to nucleic acid molecule, DNA or cDNA encoding a gene product for use as the active principle in gene therapy.
- the gene product may be an RNA, peptide or protein; transgene may encode a natural gene product or a recombinant non naturally occurring gene product e.g. the cPLA2e.
- promoter refers to a regulatory element that directs the transcription of a nucleic acid (transgene) to which it is operably linked.
- a promoter can regulate both rate and efficiency of transcription of an operably linked nucleic acid.
- a promoter may also be operably linked to other regulatory elements which enhance (“enhancers”) or repress (“repressors”) promoter-dependent transcription of a nucleic acid.
- enhance enhance
- repressors repress
- These regulatory elements include, without limitation, transcription factor binding sites, repressor and activator protein binding sites, and any other sequences of nucleotides known to one of skill in the art to act directly or indirectly to regulate the amount of transcription from the promoter, including e.g. attenuators, enhancers, and silencers.
- the promoter is located near the transcription start site of the gene or coding sequence to which is operably linked, on the same strand and upstream of the DNA sequence (towards the 5' region of the sense strand).
- a promoter can be about 100-1000 base pairs long. Positions in a promoter are designated relative to the transcriptional start site for a particular gene (i.e., positions upstream are negative numbers counting back from -1, for example -100 is a position 100 base pairs upstream).
- operably linked refers to a linkage of polynucleotide (or polypeptide) elements in a functional relationship.
- a nucleic acid is“operably linked” when it is placed into a functional relationship with another nucleic acid sequence.
- a promoter or transcription regulatory sequence is operably linked to a coding sequence if it affects the transcription of the coding sequence.
- Operably linked means that the polynucleotide sequences being linked are typically contiguous; where it is necessary to join two protein encoding regions, they are contiguous and in reading frame.
- the nucleic acid construct of the invention further comprises a promoter operably-linked to the nucleotide sequence encoding a cPLA2e.
- said promoter operably linked to the cPLA2e coding sequence is an heterologous promoter.
- heterologous when used as an attribute of a nucleotide or a peptide sequence (e.g.
- heterologous promoter means a sequence that is not naturally operably linked to another nucleotide or peptide sequence; in this particular case,“heterologous promoter” means a promoter sequence that it is not naturally operably linked to a nucleotide sequence encoding a cPLA2e.
- such promoter may be tissue or cell type specific promoter, or an organ- specific promoter, or a promoter specific to multiple organs or a systemic or ubiquitous promoter.
- the nucleic acid construct of the invention further comprises a promoter operably-linked to the nucleotide sequence encoding a cPLA2e and wherein said promoter directs the expression of encoded cPLA2e at least in neurons of the hippocampus.
- the promoter operably linked to the nucleotide sequence encoding a cPLA2e is a neuronal-specific promoter.
- the term “specific promoter” relates to a promoter that is not necessarily restricted in activity to a single cell type but which nevertheless shows selectivity in that it is active in one group of cells or tissues and less active or silent in another group. However, it may be preferred that the promoter of the nucleic acid construct of the invention shows strict cell-specificity in that they are only active at detectable levels in neuronal cells.
- a“neuronal-specific promoter” is a promoter that controls expression of genes that are uniquely or predominantly expressed in neuronal cells or in cells derived from neuronal cells.
- a neuronal-specific promoter directs expression of a gene in neuronal cells or in cells derived from neuronal cells, but does not substantially direct expression of that same gene in other cell types, for example glial cells, thus having neuronal specific transcriptional activity.
- Such a promoter may be a strong promoter or it may be a weak promoter, and it may direct constitutive expression of a gene in a neuronal cell or a cell derived from a neuronal cell, or it may direct expression in response to certain conditions, signals or cellular events.
- a neuronal-specific promoter allows an active expression in the neurons of the gene linked to it and prevents its expression in other cells or tissues.
- the promoter operably linked to the nucleotide sequence encoding a cPLA2e is a neuronal- specific promoter selected from the group consisting of: Synapsin 1 (SYN1) gene promoter (Kiigler S et al. Gene Ther. 2003; 10(4): 337-47), neuron specific enolase (NSE) gene promoter (Forss-Petters S et al. Neuron. 1990;5(2): 187-97; Twyman RM et a. J Mol Neurosci. 1997;8(l):63-73), Hb9 gene promoter (Eur J Neurosci.
- SYN1 Synapsin 1
- NSE neuron specific enolase
- such promoter may be the complete promoter, which includes core, proximal and distal promoter elements; a fragment of the promoter, e.g. core promoter, or any other fragment sufficient to direct gene expression in target cell, tissue or organ; or a chimeric or hybrid promoter, e.g. a promoter that includes the core promoter of a gene, and heterologous enhancer sequences, from another gene or synthetic.
- the term“core promoter” refers herein to the minimal portion of the promoter required to properly initiate transcription. It consists of a transcription start site and functional sequences for binding the transcription start complex (TATA-box) inside a cell or a host organism.
- Non-limiting examples of suitable neuronal- specific hybrid promoters are hybrid promoters based on SYN1 promoter, e.g. hybrid promoters resulting from the fusion of promoter elements of SYN1 and CMV genes (e.g., Matsuzaki Y. et al. J Neurosci Methods 2014; 223 : 133-143); Hb9 promoter, e.g. mouse Hb9 enhancer fused to Hsp68 minimal promoter (Singh NR et al. Exp Neurol. 2005; 196(2):224- 234) or to CMV minimal promoter (Lukashchuk V. et al. Mol Ther Methods Clin Dev. 2016; 3: 15055), among others.
- SYN1 promoter e.g. hybrid promoters resulting from the fusion of promoter elements of SYN1 and CMV genes
- Hb9 promoter e.g. mouse Hb9 enhancer fused to Hsp68 minimal promoter
- the promoter operably linked to the nucleotide sequence encoding a cPLA2e is a SYN 1 promoter, or a hybrid S YN 1 promoter, such as a hybrid SYN1 promoter that comprises core SYN1 promoter fused to CMV gene promoter elements.
- the nucleic acid construct of the invention comprises a hybrid SYN1 promoter operably linked to a nucleotide sequence encoding a cPLA2e, typically a cPLA2e of SEQ ID NO:l or 3; wherein preferably coding nucleotide sequence is SEQ ID NO:2 or 4.
- All these promoter sequences have properties of allowing expression of cPLA2e encoded by the nucleic acid construct in at least the neurons of the hippocampus.
- the promoter for use in the nucleic acid construct of the invention may be a chemical inducible promoter.
- a chemical inducible promoter is a promoter that is regulated by the in vivo administration of a chemical inducer to said subject in need thereof.
- suitable chemical inducible promoters include without limitation Tetracycline/Minocycline inducible promoter (Chtarto 2003, Neurosci Lett. 352: 155-158) or rapamycin inducible systems (Sanftner 2006, Mol Ther.13: 167-174).
- Nucleic acid construct embodiments may also include a polyadenylation signal sequence; together or not with other optional nucleotide elements.
- polyadenylation signal or“poly(A) signal” refers to a specific recognition sequence within 3’ untranslated region (3’ UTR) of the gene, which is transcribed into precursor mRNA molecule and guides the termination of the gene transcription.
- Poly(A) signal acts as a signal for the endonucleolytic cleavage of the newly formed precursor mRNA at its 3’-end, and for the addition to this 3’-end of a RNA stretch consisting only of adenine bases (polyadenylation process; poly(A) tail).
- Poly(A) tail is important for the nuclear export, translation, and stability of mRNA.
- the polyadenylation signal is a recognition sequence that can direct polyadenylation of mammalian genes and/or viral genes, in mammalian cells.
- Poly(A) signals typically consist of a) a consensus sequence AAUAAA, which has been shown to be required for both 3 '-end cleavage and polyadenylation of premessenger RNA (pre-mRNA) as well as to promote downstream transcriptional termination, and b) additional elements upstream and downstream of AAUAAA that control the efficiency of utilization of AAUAAA as a poly(A) signal.
- pre-mRNA pre messenger RNA
- the polyadenylation signal sequence of the nucleic acid construct of the invention is a polyadenylation signal sequence of a mammalian gene or a viral gene.
- Suitable polyadenylation signals include, among others, a SV40 early polyadenylation signal, a SV40 late polyadenylation signal, a HSV thymidine kinase polyadenylation signal, a protamine gene polyadenylation signal, an adenovirus 5 Elb polyadenylation signal, a growth hormone polyadenylation signal, a PBGD polyadenylation signal, in silico designed polyadenylation signal (synthetic) and the like.
- the polyadenylation signal sequence of the nucleic acid construct is a polyadenylation signal sequence based on bovine growth hormone gene.
- the nucleic acid construct according to the present invention includes a hybrid SYN1 promoter operably linked to a nucleotide sequence encoding a cPLA2e of SEQ ID NO:l or 3, and the polyadenylation signal sequence of bovine growth hormone gene; in a preferred embodiment nucleotide sequence encoding said cPLA2e is SEQ ID NO:2 or 4.
- the nucleic acid construct of the invention is an RNA.
- the nucleic acid construct has the structure of an mRNA.
- the mRNA can be modified. Modifications of mRNA nucleic acids are described in further detail in U.S. Patent Publication Nos. US20140206752 and US20150086614 and US20160304552 and PCT Publication Nos. W02016011226, WO2016014846, and W02016011306.
- the nucleic acid construct comprising a nucleotide sequence encoding a cPLA2e can further comprise at least one of the following features: a) a 5’ cap structure; b) a 5’UTR; or c) a 3’UTR.
- the polynucleotide further comprises two of the features.
- the polynucleotide can further comprise all three of these features.
- the UTRs can be homologous or heterologous to the nucleotide sequence encoding the cPLA2e.
- Untranslated regions are nucleic acid sections of a polynucleotide before a start codon (5’UTR) and after a stop codon (3’UTR) that are not translated.
- a nucleic acid construct of the invention comprising a nucleotide sequence encoding a cPLA2e further comprises UTR (e.g., a 5’UTR or functional fragment thereof, a 3’UTR or functional fragment thereof, or a combination thereof).
- the nucleic acid construct comprises two or more 5’UTRs or functional fragments thereof, each of which has the same or different nucleotide sequence. In some embodiments, the nucleic acid construct comprises two or more 3’UTRs or functional fragments thereof, each of which has the same or different nucleotide sequence. In some embodiments, the 5’UTR or functional fragment thereof, 3’UTR or functional fragment thereof, or any combination thereof is sequence optimized. In some embodiments, the 5’UTR or functional fragment thereof, 3’UTR or functional fragment thereof, or any combination thereof comprises at least one chemically modified nucleobase, e.g., 1-methylpseudouridine or 5-methoxyuracil. In some embodiments, a functional fragment of a 5’UTR or 3’UTR comprises one or more regulatory features of a full length 5’ or 3’UTR, respectively.
- the 5’UTR and the 3’UTR can be heterologous. In some embodiments, the 5’UTR can be derived from a different species than the 3’UTR.
- Wild-type UTRs derived from any gene or mRNA can be incorporated into the nucleic acid construct of the invention.
- a UTR can be altered relative to a wild type or native UTR to produce a variant UTR, e.g., by changing the orientation or location of the UTR relative to the coding nucleotide sequence; or by inclusion of additional nucleotides, deletion of nucleotides, swapping or transposition of nucleotides.
- variants of 5’ or 3’ UTRs can be utilized, for example, mutants of wild type UTRs, or variants wherein one or more nucleotides are added to or removed from a terminus of the UTR.
- one or more synthetic UTRs can be used in combination with one or more non synthetic UTRs. See, e.g., Mandal and Rossi, Nat. Protoc. 2013 8(3):568-82, and sequences available at www.addgene.org/Derrick_Rossi/, the contents of each are incorporated herein by reference in their entirety.
- the 5’ cap structure of a natural mRNA is involved in nuclear export, increasing mRNA stability and binds the mRNA Cap Binding Protein (CBP), which is responsible for mRNA stability in the cell and translation competency through the association of CBP with poly(A) binding protein to form the mature cyclic mRNA species.
- CBP mRNA Cap Binding Protein
- the cap further assists the removal of 5’ proximal introns during mRNA splicing.
- Endogenous mRNA molecules can be 5’ -end capped generating a 5’ -ppp-5’ -triphosphate linkage between a terminal guanosine cap residue and the 5’-terminal transcribed sense nucleotide of the mRNA molecule.
- This 5’- guanylate cap can then be methylated to generate an N7-methyl-guanylate residue.
- the ribose sugars of the terminal and/or ante-terminal transcribed nucleotides of the 5’end of the mRNA can optionally also be 2'-0-methylated.
- 5 '-decapping through hydrolysis and cleavage of the guanylate cap structure can target a nucleic acid molecule, such as an mRNA molecule, for degradation.
- nucleic acid construct of the present invention incorporates as 5’ cap moiety or structure.
- the nucleic acid construct of the present invention i.e., a polynucleotide comprising a nucleotide sequence encoding a cPLAe
- the nucleic acid construct of the present invention further comprises a poly- A tail.
- the nucleic acid construct of the invention may be comprised in an expression vector; thus, in an aspect the invention relates to an expression vector that comprises a nucleic acid construct of the invention.
- expression vector refers to a nucleic acid molecule used as a vehicle to transfer genetic material, and in particular to deliver a nucleic acid into a host cell, either in vitro or in vivo.
- Expression vector also refers to a nucleic acid molecule capable of effecting expression of a gene (transgene) in host cells or host organisms compatible with such sequences.
- Expression vectors typically include at least suitable transcription regulatory sequences and optionally, 3’ transcription termination signals. Additional factors necessary or helpful in effecting expression may also be present, such as expression enhancer elements able to respond to a precise inductive signal (endogenous or chimeric transcription factors) or specific for certain cells, organs or tissues.
- Vectors include, but are not limited to, plasmids, phasmids, cosmids, transposable elements, viruses, and artificial chromosomes (e.g., YACs).
- the vector of the invention is a vector suitable for use in gene or cell therapy, and in particular is suitable to target neuronal cells.
- the expression vector is a viral vector, such as vectors derived from Moloney murine leukemia virus vectors (MoMLV), MSCV, SFFV, MPSV or SNV, lentiviral vectors (e.g. derived from human immunodeficiency virus (HIV), simian immunodeficiency virus (SIV), feline immunodeficiency virus (FIV), bovine immunodeficiency virus (BIV) or equine infectious anemia virus (EIAV)), adenoviral (Ad) vectors, adeno-associated viral (AAV) vectors, simian virus 40 (SV-40) vectors, bovine papilloma virus vectors, Epstein-Barr virus, herpes virus vectors, vaccinia virus vectors, Harvey murine sarcoma virus vectors, murine mammary tumor virus vectors, Rous sarcoma virus vectors.
- lentiviral vectors e.g. derived from human immunodeficiency virus (
- suitable sequences should be introduced in the vector of the invention for obtaining a functional viral vector, such as AAV ITRs for an AAV vector, or LTRs for lentiviral vectors.
- a functional viral vector such as AAV ITRs for an AAV vector, or LTRs for lentiviral vectors.
- said vector is an AAV vector.
- AAV has arisen considerable interest as a potential vector for human gene therapy.
- favorable properties of the virus are its lack of association with any human disease, its ability to infect both dividing and non-dividing cells, and the wide range of cell lines derived from different tissues that can be infected.
- the AAV genome is composed of a linear, single-stranded DNA molecule which contains 4681 bases (Berns and Bohenzky, 1987, Advances in Virus Research (Academic Press, Inc.) 32:243-307).
- the genome includes inverted terminal repeats (ITRs) at each end, which function in cis as origins of DNA replication and as packaging signals for the virus.
- the ITRs are approximately 145 bp in length.
- the internal non-repeated portion of the genome includes two large open reading frames, known as the AAV rep and cap genes, respectively. These genes code for the viral proteins involved in replication and packaging of the virion. In particular, at least four viral proteins are synthesized from the AAV rep gene, Rep 78, Rep 68, Rep 52 and Rep 40, named according to their apparent molecular weight.
- the AAV cap gene encodes at least three proteins, VP1, VP2 and VP3.
- the nucleic acid construct or expression vector of the invention [comprising nucleotide sequence encoding cPLA2e] further comprises a 5’ITR and a 3’ITR sequences, preferably a 5’ITR and a 3’ITR sequences of an adeno- associated virus.
- ITR inverted terminal repeat
- AAV ITRs for use in the viral vector of the invention may have a wild-type nucleotide sequence or may be altered by insertion, deletion or substitution.
- the serotype of the inverted terminal repeats (ITRs) of the AAV may be selected from any known human or nonhuman AAV serotype.
- the nucleic acid construct or viral expression vector may be carried out by using ITRs of any AAV serotype, including AAV1, AAV2, AAV3 (including types 3A and 3B), AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, avian AAV, bovine AAV, canine AAV, equine AAV, ovine AAV, and any other AAV serotype now known or later discovered.
- AAV1, AAV2, AAV3 including types 3A and 3B
- AAV4 AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, avian AAV, bovine AAV, canine AAV, equine AAV, ovine AAV, and any other AAV serotype now known or later discovered.
- the nucleic acid construct or expression vector further comprises a 5’ITR and a 3’ITR of an AAV of a serotype AAV2.
- the nucleic acid construct or expression vector of the invention can be carried out by using synthetic 5’ITR and/or 3’ITR; and also by using a 5’ITR and a 3’ITR which come from viruses of different serotype. All other viral genes required for viral vector replication can be provided in trans within the virus-producing cells (packaging cells) as described below. Therefore, their inclusion in the viral vector is optional.
- the nucleic acid construct or viral vector of the invention comprises a 5’ITR, a y packaging signal, and a 3’ITR of a virus “Y packaging signal” is a cis- acting nucleotide sequence of the virus genome, which in some viruses (e.g. adenoviruses, lentiviruses ...) is essential for the process of packaging the virus genome into the viral capsid during replication.
- viruses e.g. adenoviruses, lentiviruses
- AAV viral particles The construction of recombinant AAV viral particles is generally known in the art and has been described for instance in US 5,173,414 and US5, 139,941; WO 92/01070, WO 93/03769, Lebkowski et al. (1988) Molec. Cell. Biol. 8:3988-3996; Vincent et al. (1990) Vaccines 90 (Cold Spring Harbor Laboratory Press); Carter, B. J. (1992) Current Opinion in Biotechnology 3:533-539; Muzyczka, N. (1992) Current Topics in Microbiol and Immunol. 158:97-129; and Kotin, R. M. (1994) Human Gene Therapy 5:793-801.
- the viral particle is the viral particle
- the nucleic acid construct or the expression vector of the invention may be packaged into a virus capsid to generate a "viral particle”, also named“viral vector particle”.
- the present invention relates to a viral particle comprising a nucleic acid construct or an expression vector of the invention.
- the invention relates to a viral particle that includes a nucleic acid construct or expression vector comprising a promoter, operably-linked to a nucleotide sequence encoding a cPLA2e; and to a viral particle that includes a nucleic acid construct or expression vector comprising a) a promoter operably-linked to a nucleotide sequence encoding a cPLA2e, b) a polyadenylation signal sequence, and c) 5’ITR and a 3’ITR; optionally in combination with one or more features of the various embodiments described above or below.
- the viral particle of the invention is an AAV particle comprising capsid proteins of adeno-associated virus, i.e. the nucleic acid construct or the expression vector of the invention is packaged into an AAV-derived capsid to generate an "adeno-associated viral particle" or "AAV particle".
- AAV particle encompasses any recombinant AAV particle or mutant AAV particle, genetically engineered.
- a recombinant AAV particle may be prepared by encapsidating the nucleic acid construct or viral expression vector including ITR(s) derived from a particular AAV serotype on a viral particle formed by natural or mutant Cap proteins corresponding to an AAV of the same or different serotype.
- Proteins of the viral capsid of an adeno-associated virus include the capsid proteins VP1, VP2, and VP3. Differences among the capsid protein sequences of the various AAV serotypes result in the use of different cell surface receptors for cell entry. In combination with alternative intracellular processing pathways, this gives rise to distinct tissue tropisms for each AAV serotype.
- an AAV particle according to the invention may be prepared by encapsidating the viral vector of an AAV vector/genome derived from a particular AAV serotype on a viral particle formed by natural Cap proteins corresponding to an AAV of the same particular serotype.
- AAV viral particles according to the invention include the nucleic acid construct comprising the nucleotide sequence encoding the cPLA2e flanked by ITR(s) of a given AAV serotype packaged, for example, into: a) a viral particle constituted of capsid proteins derived from the same or different AAV serotype [e.g. AAV2 ITRs and AAV9 capsid proteins; AAV2 ITRs and AAV TT capsid proteins; etc]; b) a mosaic viral particle constituted of a mixture of capsid proteins from different AAV serotypes or mutants [e.g.
- AAV2 ITRs with a capsid formed by proteins of two or multiple AAV serotypes comprising c) a chimeric viral particle constituted of capsid proteins that have been truncated by domain swapping between different AAV serotypes or variants [e.g. AAV2 ITRs with AAV5 capsid proteins with AAV3 domains]; or d) a targeted viral particle engineered to display selective binding domains, enabling stringent interaction with target cell specific receptors.
- examples of AAV serotype of the capsid proteins of AAV particle according to the present invention include AAV2, AAV5, AAV9, and AAV TT.
- said AAV serotype of the capsid proteins are selected from AAV9 and AAV TT serotype.
- the viral particle is an AAV particle including a nucleic acid construct or expression vector that comprises 5’ITR and 3’ITR sequences from an AAV virus; preferably AAV particle comprises capsid proteins of an AAV2, AAV5, AAV9, or AAV TT serotype, more preferably of an AAV9 serotype or of an AAV TT serotype; and/or 5’ITR and 3’ITR sequences of an AAV2 serotype.
- the viral particle includes a nucleic acid construct or expression vector comprising a nucleotide sequence encoding a human cPLA2e of amino acid SEQ ID NO: l or SEQ ID NO:3 under the control of a promoter, said promoter allowing expression of said human cPLA2e in at least neurons of the hippocampus, and said viral particle is selected among viral particles that targets at least neurons of the hippocampus, typically an AAV particle including capsid proteins selected from the group consisting of AAV2, AAV5, AAV9, or AAV TT serotypes; preferably the nucleotide sequence encoding human cPLA2e is SEQ ID NO:2 or SEQ ID NO:4, and/or the promoter is a neuronal-specific promoter, more preferably a SYN1 promoter or a hybrid SYN1 promoter.
- such recombinant AAV particle according to the present invention comprises capsid proteins of the AAV9 or AAV TT serotype and an AAV vector comprising (i) a nucleic acid construct comprising a hybrid SYN1 promoter operably linked to a nucleotide sequence SEQ ID NO:2 or 4 encoding a human cPLA2e, and (ii) AAV ITRs, such as 5’ and 3’ ITRs of AAV2, flanking said nucleic acid construct.
- the AAV viral particle according to the present invention may comprise capsid proteins from any AAV serotype including AAV1, AAV2, AAV3 (including types 3A and 3B), AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, avian AAV, bovine AAV, canine AAV, equine AAV, ovine AAV, synthetic AAV variants such as NP40, NP59, NP84 (Paulk et al. Mol ther. 2018.26(1):289- 303), LK03 (Wang L et al. Mol Ther. 2015. 23(12): 1877-87), AAV3-ST (Vercauteren et al. Mol Ther. 2016.24(6): 1042-1049), Anc80 (Zinn E et al., Cell Rep. 2015;12(6): 1056-68) and any other AAV serotype now known or later discovered.
- AAV1, AAV2, AAV3 including types 3A
- Production of viral particles carrying the expression viral vector as disclosed above can be performed by means of conventional methods and protocols, which are selected taking into account the structural features chosen for the actual embodiment of expression vector and viral particle of the vector to be produced.
- viral particles can be produced in a host cell, more particularly in specific virus-producing cell (packaging cell), which is transfected with the nucleic acid construct or expression vector to be packaged, in the presence of a helper vector or virus or other DNA construct(s).
- packaging cell specific virus-producing cell
- helper vector or virus or other DNA construct(s) in the presence of a helper vector or virus or other DNA construct(s).
- packaging cells refers to a cell or cell line which may be transfected with a nucleic acid construct or expression vector of the invention and provides in trans all the missing functions which are required for the complete replication and packaging of a viral vector.
- the packaging cells express in a constitutive or inducible manner one or more of said missing viral functions.
- Said packaging cells can be adherent or suspension cells.
- a process of producing viral particles comprises the following steps: a) culturing a packaging cell comprising a nucleic acid construct or expression vector as described above in a culture medium; and b) harvesting the viral particles from the cell culture supernatant and/or inside the cells.
- AAV viral particles which consist on transient cell co-transfection with nucleic acid construct or expression vector (e.g. a plasmid) carrying the transgene of the invention; a nucleic acid construct (e.g., an AAV helper plasmid) that encodes rep and cap genes, but does not carry ITR sequences; and with a third nucleic acid construct (e.g., a plasmid) providing the adenoviral functions necessary for AAV replication.
- Viral genes necessary for AAV replication are referred herein as viral helper genes.
- said genes necessary for AAV replication are adenoviral helper genes, such as E1A, E1B, E2a, E4, or VA RNAs.
- the adenoviral helper genes are of the Ad5 or Ad2 serotype.
- AAV particles can also be carried out for example by infection of insect cells with a combination of recombinant baculoviruses (Urabe et al. Hum. Gene Ther. 2002; 13: 1935-1943).
- SF9 cells are co-infected with two or three baculovirus vectors respectively expressing AAV rep, AAV cap and the AAV vector to be packaged.
- the recombinant baculovirus vectors will provide the viral helper gene functions required for virus replication and/or packaging.
- Smith et al 2009 (Molecular Therapy, vol.17, no.11, pp 1888-1896) further describes a dual baculovirus expression system for large-scale production of AAV particles in insect cells.
- Suitable culture media will be known to a person skilled in the art.
- the ingredients that compose such media may vary depending on the type of cell to be cultured. In addition to nutrient composition, osmolarity and pH are considered important parameters of culture media.
- the cell growth medium comprises a number of ingredients well known by the person skilled in the art, including amino acids, vitamins, organic and inorganic salts, sources of carbohydrate, lipids, trace elements (CuS04, FeS04, Fe(N03)3, ZnS04%), each ingredient being present in an amount which supports the cultivation of a cell in vitro (i.e., survival and growth of cells).
- Ingredients may also include different auxiliary substances, such as buffer substances (like sodium bicarbonate, Hepes, Tris%), oxidation stabilizers, stabilizers to counteract mechanical stress, protease inhibitors, animal growth factors, plant hydrolyzates, anti-clumping agents, anti-foaming agents. Characteristics and compositions of the cell growth media vary depending on the particular cellular requirements.
- Examples of commercially available cell growth media are: MEM (Minimum Essential Medium), BME (Basal Medium Eagle) DMEM (Dulbecco’s modified Eagle’s Medium), Iscoves DMEM (Iscove’s modification of Dulbecco’s Medium), GMEM, RPMI 1640, Leibovitz L-15, McCoy’s, Medium 199, Ham (Ham’s Media) F10 and derivatives, Ham F12, DMEM/F12, etc.
- Viral Vectors for Gene Therapy Methods and Protocols. Series: Methods in Molecular Biology, Vol. 737. Merten and Al-Rubeai (Eds.); 2011 Humana Press (Springer); Gene Therapy. M. Giacca. 2010 Springer- Verlag; Heilbronn R. and Weger S. Viral Vectors for Gene Transfer: Current Status of Gene Therapeutics. In: Drug Delivery, Handbook of Experimental Pharmacology 197; M. Schafer-Korting (Ed.). 2010 Springer- Verlag; pp. 143-170; Adeno-Associated Virus: Methods and Protocols. R.O. Snyder and P. Moulllier (Eds).
- the invention relates to a host cell comprising a nucleic acid construct or an expression vector of the invention.
- the host cell according to the invention is a specific virus- producing cell, also named packaging cell, which is transfected with the nucleic acid construct or expression vector according to the invention, in the presence of a helper vector or virus or other DNA constructs and provides in trans all the missing functions which are required for the complete replication and packaging of a viral particle.
- Said packaging cells can be adherent or suspension cells
- said packaging cells may be eukaryotic cells such as mammalian cells, including simian, human, dog and rodent cells.
- human cells are PER.C6 cells (WOOl/38362), MRC-5 (ATCC CCL-171), WI-38 (ATCC CCL-75), HEK-293 cells (ATCC CRL-1573), HeLa cells (ATCC CCL2) and fetal rhesus lung cells (ATCC CL- 160).
- non-human primate cells are Vero cells (ATCC CCL81), COS-1 cells (ATCC CRL-1650) or COS-7 cells (ATCC CRL-1651).
- dog cells are MDCK cells (ATCC CCL-34).
- rodent cells are hamster cells, such as BHK21-F, HKCC cells, or CHO cells.
- the packaging cells for producing the viral particles may be derived from avian sources such as chicken, duck, goose, quail or pheasant.
- avian cell lines include avian embryonic stem cells (WOO 1/85938 and W003/076601), immortalized duck retina cells (W02005/042728), and avian embryonic stem cell derived cells, including chicken cells (W02006/108846) or duck cells, such as EB66 cell line (W02008/129058 & WO2008/142124).
- the cells can be any cells permissive for baculovirus infection and replication packaging cells.
- said cells are insect cells, such as SF9 cells (ATCC CRL-1711), Sf21 cells (IPLB-Sf21), MG1 cells (BTI-TN-MG1) or High FiveTM cells (BTI-TN-5B1-4).
- the host cell comprises: a nucleic acid construct or expression vector comprising a nucleotide sequence encoding a cPLA2e according to the invention (e.g., the AAV vector according to the invention); a nucleic acid construct, for example a plasmid, encoding AAV rep and/or cap genes which does not carry the ITR sequences; and/or a nucleic acid construct, for example a plasmid or virus, comprising viral helper genes.
- a nucleic acid construct or expression vector comprising a nucleotide sequence encoding a cPLA2e according to the invention (e.g., the AAV vector according to the invention); a nucleic acid construct, for example a plasmid, encoding AAV rep and/or cap genes which does not carry the ITR sequences; and/or a nucleic acid construct, for example a plasmid or virus, comprising viral helper genes.
- the invention in another aspect, relates to a host cell transduced with an expression vector or viral particle of the invention and the term“host cell” as used herein refers to any cell line that is susceptible to infection by a virus of interest, and amenable to culture in vitro.
- host cell of the invention can be used for therapeutic purposes, e.g. for the therapeutic uses disclosed herein.
- Another aspect of the present invention is a pharmaceutical composition
- a pharmaceutical composition comprising a nucleic acid construct as mentioned above, a vector as mentioned above, a host cell as mentioned above, or a viral particle as mentioned above, in combination with one or more pharmaceutical acceptable excipients.
- the invention also refers to a pharmaceutical composition comprising a cPLA2e inducing agent in any of the embodiments disclosed above or below, for use or administration in the treatment of a cognitive disorder and/or a disease associated with a cognitive disorder.
- the term "pharmaceutically acceptable” means approved by a regulatory agency or recognized pharmacopeia such as European Pharmacopeia, for use in animals and/or humans.
- excipient refers to a diluent, adjuvant, carrier, or vehicle with which the therapeutic agent is administered.
- the pharmaceutical composition or medicament of the invention typically comprises the therapeutic agent (e.g. a vector or viral particle of the invention) in an effective amount, sufficient to provide a desired therapeutic effect, and a pharmaceutically acceptable carrier or excipient.
- the therapeutic agent e.g. a vector or viral particle of the invention
- the invention relates then to a pharmaceutical composition that comprises a vector or viral particle as disclosed above, and a pharmaceutically acceptable carrier.
- compositions are typically sterile and stable under the conditions of manufacture and storage.
- Pharmaceutical compositions may be formulated as solutions (e.g. saline, dextrose solution, or buffered solution, or other pharmaceutically acceptable sterile fluids), microemulsions, liposomes, or other ordered structure suitable to accommodate a high product concentration (e.g. microparticles or nanoparticles).
- the carrier may be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof.
- the proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants.
- isotonic agents for example, sugars, polyalcohols such as mannitol, sorbitol, or sodium chloride in the composition.
- said pharmaceutical composition is formulated as a solution, more preferably as an optionally buffered saline solution.
- said pharmaceutical composition is formulated as a solution, more preferably as an optionally buffered saline solution.
- Supplementary active compounds can also be incorporated into the pharmaceutical compositions of the invention. Guidance on co administration of additional therapeutics can for example be found in the Compendium of Pharmaceutical and Specialties (CPS) of the Canadian Pharmacists Association.
- the pharmaceutical composition is a composition suitable for intraparenchymal, intracerebral, intravenous, or intrathecal administration. These pharmaceutical compositions are exemplary only and do not limit the pharmaceutical compositions suitable for other parenteral and non-parenteral administration routes.
- the pharmaceutical compositions described herein can be packaged in single unit dosage or in multidosage forms.
- another aspect of the invention relates to a method for treating a cognitive disorder and/or a disease associated with a cognitive disorder, for example dementia, in particular age-related dementia or Alzheimer’s disease, in a subject in need thereof, said method comprising administering to said subject a therapeutically effective amount of a cPLA2e inducing agent.
- a cognitive disorder and/or a disease associated with a cognitive disorder for example dementia, in particular age-related dementia or Alzheimer’s disease
- the invention relates to a cPLA2e inducing agent, for use as a medicament in a subject in need thereof, and more specifically, for use in the treatment of a cognitive disorder and/or disease associated with a cognitive disorder, for example dementia, and more specifically age-related dementia or Alzheimer’s disease, in a subject in need thereof.
- the invention pertains to the use of a cPLA2e inducing agent for the manufacturing of a medicament, more specifically, for the treatment of a cognitive disorder and/or disease associated with a cognitive disorder, for example dementia, and more specifically age-related dementia or Alzheimer’s disease.
- the cPLA2e inducing agent for use or administration in the treatment of cognitive disorders and diseases associated with a cognitive disorder according to the invention can be selected, among others, from the group consisting of: a) a nucleic acid construct of the invention, as mentioned above; b) a vector comprising a nucleic acid construct of the invention, as mentioned above; c) a viral particle comprising a nucleic acid construct or vector of the invention, as mentioned above; d) a host cell comprising a nucleic acid construct or vector according to the invention; e) a cPLA2e polypeptide or protein; and f) a pharmaceutical composition comprising any of said nucleic acid construct, vector comprising the nucleic acid construct, viral particle comprising the nucleic acid construct or vector, and cPLA2e polypeptide or protein.
- any and all the embodiments and preferred embodiments mentioned above for the cPLA2e encoded by the nucleotide sequence included in the nucleic acid construct of the invention are embodiments within the scope of the cPLA2e polypeptide or protein for use or administration in the treatment according to the invention; in particular, a cPLA2e comprising or consisting of amino acid SEQ ID NO: l or 3 or a variant with at least 70% sequence identity thereto; optionally as a fusion protein with another polypeptide(s), such as a tag or carrier polypeptide.
- the cPLA2e inducing agent for the therapeutic use of the invention is a vector of the invention, more preferably a viral vector, or a viral particle (e.g. an AAV particle), or the pharmaceutical composition that comprises it.
- cognition disorder and “cognitive impairment,” interchangeably refer to any impairment of cognition such as a condition characterized by one or more of the following behaviors: inhibition of at least one form of learning (e.g., associative learning), inhibition of at least one form of memory function (e.g., executive function), inhibition of learning, inhibition of memory acquisition, inhibition of memory recall, suppression of long term potentiation (LTP) in the hippocampus, or a combination thereof.
- learning e.g., associative learning
- memory function e.g., executive function
- LTP long term potentiation
- any suitable method for testing and neuroimaging of the hippocampus or brain function can be used to determine or assess cognition and its potential impairment.
- cognition e.g., memory or learning
- its potential impairment can be measured using any suitable psychological test, including without limitation,“Kiel Locomotor Maze”, containing features of the Radial Arm Maze and the Morris Water Maze, in order to assess spatial memory and orientation, which has been optimized for school age children, or Cambridge Neuropsychological Test Automated Battery (CANTAB), a computerized battery of nonverbal visually-presented neuropsychological tests, designed to test spatial memory span, spatial working memory and spatial recognition.
- CANTAB Cambridge Neuropsychological Test Automated Battery
- the outcome of methods related to cognitive impairment disclosed herein can be shown via comparative testing in animals (e.g., rats or mice), using the same composition administered to humans.
- the cPLA2e inducing agents of the invention are particularly useful for the treatment of cognitive disorders associated to conditions which impair or otherwise affect normal functioning of the central nervous system and diseases associated with cognitive disorders, such as dementias (e.g. age-associated dementia (senile dementia), cerebrovascular dementia, and/or neurodegenerative dementing disease with aberrant protein aggregations as specially Alzheimer's disease, Parkinson disease, ALS, or prion diseases, as Creutzfeldt-Jakob disease or Gerstmann-Straussler-Scheinher disease); mild cognitive impairment, attention deficit disorder, among others.
- dementias e.g. age-associated dementia (senile dementia), cerebrovascular dementia, and/or neurodegenerative dementing disease with aberrant protein aggregations as specially Alzheimer's disease, Parkinson disease, ALS, or prion diseases, as Creutzfeldt-Jakob disease or Gerstmann-Straussler-Scheinher disease
- mild cognitive impairment attention deficit disorder
- the cPLA2e inducing agent is particularly useful for the treatment of a cognitive disorder in a patient with Alzheimer’s disease.
- Alzheimer’s disease refers to a progressive neurodegenerative disorder of the central nervous system of an unknown origin.
- the defining characteristic is cognitive impairment.
- AD is defined as a neurodegenerative disorder that constitutes the main common form of dementia in the elderly: it is characterized by accumulation of two abnormal proteins in the brain: beta-amyloid peptide and hyperphosphorylated tau in form of amyloid plaques and neurofibrillary tangles respectively.
- NINCDS National Institute of Neurological and Communicative Disorders and Stroke
- ARRDA Alzheimer’s disease and Related Disorders Association
- MCI Mild Cognitive Impairment
- subject or“patient” refers to mammals.
- Mammalian species that can benefit from the disclosed methods of treatment include, but are not limited to, humans, non-human primates such as apes, chimpanzees, monkeys, and orangutans, domesticated animals, including dogs and cats, as well as livestock such as horses, cattle, pigs, sheep, and goats, or other mammalian species including, without limitation, mice, rats, guinea pigs, rabbits, hamsters, and the like.
- “treatment”,“treating” or“treat” refer to: (i) preventing or retarding a disease, disorder or condition from occurring in a subject which may be predisposed to the disease, disorder and/or condition but has not yet been diagnosed as having it; (ii) inhibiting the disease, disorder or condition, i.e., arresting or slowing down its development or progression; and/or (iii) relieving the disease, disorder or condition, i.e., causing regression of the disease, disorder and/or condition. In certain embodiments, such term refers to the amelioration or eradication of a disease or symptoms associated with a disease.
- “treatment”,“treating” or“treat” refer to: (i) preventing or retarding cognitive impairment from occurring in a subject which may be predisposed to cognitive impairment but has not yet been diagnosed as having it; (ii) inhibiting cognitive impairment, i.e., arresting or slowing down its development or progression; (iii) relieving cognitive impairment, i.e., causing its regression; and/or (iv) enhancing cognitive performance.
- the treatment of cognitive impairment relates, in particular, to the treatment of learning and memory impairments, and to the enhancement of learning and memory performance.
- “Enhancing learning and memory performance” refers to improving or increasing the mental faculty by which to register, retain or recall past experiences, knowledge, ideas, sensations, thoughts or impressions.
- a "therapeutically effective amount” refers to an amount effective, at dosages and for periods of time necessary to achieve the desired therapeutic result, such as one or more of the following therapeutic results: a significant delay of the onset or progression of the disease; a significant reduction of the severity of one or more symptoms; a significant reduction of hallmarks of AD, amyloid and/or tau pathology; a significant increase in synaptic plasticity; a significant attenuation of mortality associated to aging and/or AD.
- a therapeutically effective amount is also typically one in which any toxic or detrimental effect of the product or pharmaceutical composition is outweighed by the therapeutically beneficial effects.
- nucleic acid construct, expression vector, viral particle, host cell, cPLA2e polypeptide or protein or pharmaceutical composition for its therapeutic use according to the invention is administered to the subject or patient by a parenteral route, e.g. by intraparenchymal, intracerebral, intracerebroventricular (icv), intrathecal, intranasal, intravenous, or subcutaneous route.
- a parenteral route e.g. by intraparenchymal, intracerebral, intracerebroventricular (icv), intrathecal, intranasal, intravenous, or subcutaneous route.
- a therapeutically effective amount of said nucleic acid construct, expression vector, viral particle, host cell, cPLA2e polypeptide or protein, or pharmaceutical composition is preferably administered by intrathecal or intraparenchymal route, the latter preferably to brain areas such as the hippocampal formation or cerebral cortex.
- the intraparenchymal route may facilitate preferred local administration to hippocampus and cortex as compared to other area of the brain.
- a“preferred local administration to hippocampus” does not mean that all the cPLA2e inducing agent is administered to said areas of the brain, but a majority, for example at least 50%, at least 60%, at least 70%, or at least 80% of the cPLA2e inducing agent is administered to said areas.
- the therapeutically effective amount of the cPLA2e inducing agent e.g. a nucleic acid construct, expression vector, viral particle, host cell, or cPLA2e polypeptide or protein
- the pharmaceutical composition that comprises it may vary according to factors such as the disease state, age, sex, and weight of the individual, and the ability of the product or pharmaceutical composition to elicit a desired response in the individual. Dosage regimens may be adjusted to provide the optimum therapeutic response.
- dosage regimens may be adjusted over time according to the individual needs and the professional judgment of the person administering or supervising the administration of the compositions. Dosage ranges set forth herein are exemplary only and do not limit the dosage ranges that may be selected by medical practitioners.
- an AAV viral particle according to the invention can be administered to the human subject or patient for the treatment of a cognitive disorder or a disease associated with a cognitive disorder, such as Alzheimer’s disease, in an amount or dose comprised within a range of 10 8 to 10 14 vg / kg (vg: viral genomes; kg: subject’s or patient’s body weight), for example 1 x 10 10 to 5 x 10 14 vg/kg.
- an amount comprised within a range of 1 x 10 12 to 1 x 10 13 vg/kg is administered.
- an amount or dose comprised within a range of 1 x 10 9 to 1 x 10 11 iu/kg is administered.
- the invention further relates to a kit comprising a nucleic acid construct, expression vector, host cell, viral particle of the invention, or a pharmaceutical composition comprising said nucleic acid construct, vector, host cell or viral particle, in one or more containers.
- the kit may include instructions or packaging materials that describe how to administer the nucleic acid construct, expression vector, viral particle, host cell or pharmaceutical compositions contained within the kit to a patient.
- Containers of the kit can be of any suitable material, e.g., glass, plastic, metal, etc., and of any suitable size, shape, or configuration.
- the kits may include one or more ampoules or syringes that contain the products of the invention in a suitable liquid or solution form.
- the inventors have also put their efforts on developing systems to screen compound candidates, such as a peptide, polypeptide (e.g., antibodies) or small molecule candidates, taking advantage of the fact that cPLA2e induction or increase results in a very clear increase of the calcium-dependent N-acyltransferase (Ca-NAT) activity.
- compound candidates such as a peptide, polypeptide (e.g., antibodies) or small molecule candidates
- the present invention also provides a method for identifying a compound as a candidate for the treatment of a cognitive disorder and/or disease associated with a cognitive disorder which comprises the steps of:
- a possible embodiment of the method of the invention is carrying out an in vitro method, wherein the assayed cells are mammalian cells, such as HEK293T cells. These cells are cultured in a medium suitable for cell growth and proliferation in the presence of the candidate compound (e.g. for 30 minutes or 1 hour), with or without ionomycin (e.g. 2 pm) and Ca-NAT activity is tested (e.g. by targeted metabolite profiling) with regard to a control that has had no contact with the candidate compound. If Ca-NAT activity is found to be increased with respect to that of control cells, the compound is identified as a possible candidate for the treatment of a cognitive disorder and/or disease associated with a cognitive disorder.
- the candidate compound e.g. for 30 minutes or 1 hour
- ionomycin e.g. 2 pm
- Ca-NAT activity is tested (e.g. by targeted metabolite profiling) with regard to a control that has had no contact with the candidate compound. If Ca-NAT activity is found to be increased
- cPLA2e-transfected cells e.g., with a nucleic acid construct of the invention
- induce or increase may refer to the ability to cause an overall increase, preferably of 20% or greater, more preferably of 50% or greater, and most preferably of 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or greater.
- PLA2G4E has a role in learning and memory functions
- the inventors overexpressed PLA2G4E in the brain of a) APP/PSl mice (model for AD) and b) (aged- wild type animals), which are both affected with cognitive impairment.
- they constructed an AAV vector carrying as transgene Mus musculus cPLA2e and administered it to the animals. Learning and memory functions were then assessed by the MWM method.
- AAV2/9-mPLA2G4E vector was constructed, which includes as transgene the nucleotide sequence SEQ ID NO: 5 encoding a murine PLA2G4E fused to a flag sequence through a linker.
- a 4163 bp backbone fragment was obtained from plasmid pAAV-ha- synucleinA53T (kindly gifted by Dr. J. Gerez), by running a digestion with Xhol (in CutSmart® Buffer) and a subsequent treatment with Klenow polymerase, dNTPs and NEB2.1 buffer. After purification, the 4163 backbone fragment was then digested with SaCL (in CutSmart® Buffer) and dephosphorylated using the Shrimp Alkaline Phosphatase rSAP (New England Biolabs, MA, USA; Weissig, H. et al. Biochem. J. 1993; 290: 503-508) to avoid vector re-ligation. The 4163 pb backbone fragment was finally separated, extracted and purified as described above for 3108 bp fragment.
- the 3108 bp fragment was cloned into the 4163 bp backbone fragment by means of a treatment with a T4 DNA ligase (Invitrogen).
- the pAAV2-mPLA2G4E with desired construct was produced, it was then subjected to several amplification steps to generate an appropriate amount of plasmid for final virus production.
- E. coli chemically-competent bacteria were transformed with the plasmid using TOP 10 electro-competent cells (Invitrogen) and the bacteria that had incorporated the plasmid were selected by plating on LB medium with ampicillin (50mg/ml). Then, the plasmid was obtained and purified from the bacteria using a QIAprep® Spin Miniprep kit (QUIAGEN).
- the AAV vector particles were produced by double transfection into HEK-293T cells of the plasmid pAAV2- mPLA2G4E and of a pDP9 helper plasmid that expresses adenoviral molecules required for production and packaging of the AAV: AAV9 cap and AAV2 rep (Durocher, Y., S. Perret, and A. Kamen., 2002, Nucleic Acids Research, 30(2):9e-9).
- the vector particles were finally purified by iodixanol gradient and titrated by quantitative PCR. Viral titration, expressed as viral particles (vp)/ml, was obtained through quantitative PCR (q-PCR) using primers for mouse PLA2G4E,
- the murine APP/PS1 model expresses human transgenes for the amyloid precursor protein (APP) bearing the Swedish mutation (K595N/M596L) and for the PSEN1 containing a L166P mutation, both driven by the Thyl promoter. These mice are on an inbred C57BL/6J genetic background.
- the AD murine model APP/PS1 is a more accelerated amyloidosis model than the Tg2576. In these mice, the expression of human APP transgene is approximately three times higher than that of endogenous murine APP and human Ab42 is preferentially generated over Ab40.
- amyloid plaque deposition starts in the hippocampus at 3-4 months (Radde et al., 2006, EMBO reports, 7(9): 940- 946)(Maia LF et al., 2013, Science translational medicine, 5(194):94-194) and cognitive impairment is presented from 7 months (Semeels L et al, 2009, Science, 324(5927):639- 642).
- APP/PS1 and their correspondent negative littermates both male and female 16-19 months old mice were used.
- Aged wild type mice Wild type mice present age-related memory deficits. Specifically, in the Morris water maze, aged wild type mice do not form a robust memory of the platform location in the hidden phase because they perform significantly worse than young mice during probe trials. These mice were on an inbred C57BL/6/SJL genetic background.
- mice were administered with AAV2/9-mPLA2G4E in the CA1 region of the hippocampus through a stereotactic surgery.
- This procedure is based on a three-dimensional system of axes and spatial coordinates that allows the localization of specific points in the mouse brain (given as three-dimensional distances in millimeters (mm)) taking bregma or lambda, two easily identifiable points in the brain, as reference.
- mice were placed in the stereotactic device with the head completely immobilized. After disinfecting the area with 96° alcohol, an antero posterior cut was made in the skin using a scalpel, releasing thus the skull from its periosteum and leaving visible the bregma and lambda reference points. Next, with the help on a drill bill, a hole was made in the skull and a 5 pi Hamilton syringe was placed on the stereotaxic arm loaded with the vector viral particles (2.6 x 10E8 genomic copies) or unloaded (for sham procedure).
- mice Once positioned on the exact coordinate, 1 ml of the solution was injected at 0.2 m ⁇ /min and then, the syringe was maintained there for another 2 min to allow correct diffusion of the virus before withdrawing slowly the syringe. For sham-injected mice, syringe remained in the brain 5 min before the withdrawing. The same procedure was repeated for the other hemisphere.
- the wound was sutured and povidone iodine (Betadine®) was administered topically. Then, animals were placed on an electric blanket to avoid heat lost until their awakening. Finally, they were stabled individually in clean cages, with easy access to softened-in water food to facilitate food intake after surgery. Throughout the intervention, physiological serum was continuously applied in mice eyes to avoid their dryness and consequent loss of vision.
- This test is carried out in a circular pool (diameter 1.2 m) filled with water at 20 °C and made opaque by the addition of non-toxic white paint.
- the pool is divided into four imaginary quadrants, in one of which is located the platform that the mouse must learn to locate in order to escape from water and be safe.
- In each of the four walls that surround the pool there is a picture of a geometric figure that would serve as a guide for the mouse and that would be cover or uncover depending on the phase of the test.
- mice behavior was monitored by a camera anchored in the ceiling, just above the pool, and recorded with an HVS system to allow the subsequent analysis of escape latencies, swimming speed, path length and percentage of time spent in each quadrant of the pool using the software SMART -LD (Panlab).
- Visible-platform phase in this phase, the platform was located in the center of one of the quadrants, elevated 1 cm above the water and identified by a piece clearly visible to the animal in order to facilitate its location.
- mice should became familiar with the pool and learn to go to the platform to escape from water, so the visual clues remain hidden.
- the visible-platform phase mice were trained 8 times per day during 3 consecutive days. In each trial, mice had 60 s to locate the platform; if they could not reach it during this period, they were placed on it. Once the animal was on the platform, he was allowed to inspect it for 15 s before being returned to his cage.
- Probe trial memory retention was evaluated in probe trials carried out on day 6 th and 8 th of the hidden-platform phase, just before starting the hidden-platform trials of those days. For this test, the platform was removed from the pool and animals were allowed to swim for 60 s. The time that mice spend in the quadrant where the platform was placed during the hidden-platform phase is considered an estimate of memory retention degree. Retention rates higher than 25% are considered indicative of learning while those lower than 25% are considered random.
- slices were incubated in 70° ethanol, washed with distilled water, reduced in 16% ammonia for one hour and fixed in 1% sodium thiosulfate for 7 min. After another wash, slices were placed in microscope slides, dehydrated in an increasing alcohol graduation and mounted with DPX Mountant (VWR, BDH Prolabo®).
- FC paradigm was used to analyze the effect of PLA2G4E expression on fear memory. This behavioral test consists of three phases: habituation, training and test. It was carried out in a StartFear system (Panlab). During habituation phase, mice were habituated to the conditioning chamber for 3 min with no stimuli presented. After twenty-four hours, during the training phase, they were placed again in the same chamber and were allowed to explore for 2 min. After that, they received two footshocks (0.3 mA) of 2 s separated by an interval of 30 s and were returned to their home cages after another 30 s. The following day, mice were returned to the conditioning chamber and allowed to explore the context for 2 min. Freezing behavior was recorded during this time and freezing scores were expressed as percentages. The T24 group was sacrificed 24 h after the training and the TT group 1 h after the test. The naive group was sacrificed without performing any step of the paradigm.
- Protein samples were mixed with 6x Laemmli sample buffer, boiled for 5 min at 95 °C, resolved onto SDS-polyacrylamide gels and transferred to nitrocellulose membranes.
- Membranes were then blocked with 5% milk in TBS and incubated overnight with the following primary antibodies: rabbit polyclonal anti-pGluAl-Ser831(1 : 1000, Millipore), rabbit monoclonal anti-pCREB (Serl33) (1 : 1000, Cell Signaling), mouse monoclonal anti- synapsin I (1 : 1000, Synaptic Systems), rabbit polyclonal anti-PLA2G4E (1 : 1000, Proteintech) and mouse monoclonal anti-b-actin (1 : 100000, Synaptic Systems) in the corresponding buffer.
- rabbit polyclonal anti-pGluAl-Ser831(1 : 1000, Millipore) rabbit monoclonal anti-pCREB (Serl33) (1 : 1000, Cell Signaling)
- mouse monoclonal anti- synapsin I (1 : 1000, Synaptic Systems
- rabbit polyclonal anti-PLA2G4E (1 : 1000, Proteintech
- RNAi small interfering RNA
- AAV9-shPLA2G4E adeno-associated virus serotype 9
- siRNA delivery Unitat de Producci ⁇ de Vectors, Barcelona.
- PLA2G4E adeno-associated virus serotype 9
- Primary neuronal cultures were obtained from the hippocampus and cortex of embryonic day 16 (E16) wild type (WT) mice (A.
- mice treated with the AAV2/9-mPLA2G4E spent more time in the right quadrant than sham-injected mice during the probe trial on day 6 th .
- the Golgi-Cox method was used to analyze whether the behavioral recovery induced by PLA2G4E overexpression was reflected by structural changes in dendritic spine density. Specifically, apical dendrites from pyramidal neurons of the CA1 region of the hippocampus were studied.
- mice treated with AAV2/9-mPLA2G4E spent more time in the right quadrant during the probe trials performed on day 5 th ( Figure 3B) and 7 th (data not shown) than sham-injected mice, indicating that viral PLA2G4E hippocampal overexpression improves memory retention rates in elderly WT mice.
- hippocampal PLA2G4E overexpression mediated by AAV2/9-mPLA2G4E treatment improved memory retention in elderly C57BL/6/SJL WT mice three months after injection.
- PLA2G4E expression was regulated in the fear conditioning (FC) test. This task requires hippocampal-dependent transcription and protein synthesis, and it has been widely used to characterize the biochemical requirements for memory formation (Huff et al, 2006 J. Neurosci., 26, pp. 1616-1623).
- pCREB was first analyzed as indicative of neural plasticity in the hippocampus of the animals. An up- regulation of pCREB in the hippocampus was observed in the group of mice re-introduced into the cage.
- PLA2G4E expression was also stronger in both of these areas in this group of mice compare to the others (Fig. 4C).
- PLA2G4E in memory function, in vitro assays were conducted to further characterize its role on synaptic activity.
- CREB activation phosphorylation of CREB at the activator site residue Ser 133 was analyzed (Ginty et al., 1993 Science 260: 238-241). As depicted in Fig. 5 and described by several authors (Hardingham et al., 2002 Nat. Neurosci., 5: 405-414), we demonstrated that bicuculline (through the activation of NMDA receptors) caused a sustained CREB phosphorylation at Seri 33 as well as an increase of AMPA receptor activation (Rao et al., 2006 Nat. Neurosci., 9: 887-895) which was analyzed measuring pGluAl levels.
- Synapsin I levels were also analyzed since this presynaptic protein increases in the hippocampus during long term potentiation (LTP) (Sato et al., 2000 Brain Res., 872: 219-222) and plays a fundamental role in the formation, maintenance and rearrangements of synaptic contacts (review in Cesca et al, 2010 Prog. Neurobiol, 91 : 313-348).
- LTP long term potentiation
- a significant increase of synapsin I was also observed in neuronal cultures activated with bicuculline.
- PLA2G4E expression was next analyzed on the same conditions and, interestingly, we observed that it was strongly induced by bicuculline which indicates that neuronal activation indeed upregulated PLA2G4E expression.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Biomedical Technology (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Medicinal Chemistry (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Biochemistry (AREA)
- Neurosurgery (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Pharmacology & Pharmacy (AREA)
- Neurology (AREA)
- Microbiology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Psychiatry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Hospice & Palliative Care (AREA)
- Epidemiology (AREA)
- Physics & Mathematics (AREA)
- Biophysics (AREA)
- Virology (AREA)
- Plant Pathology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Enzymes And Modification Thereof (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
Description
Claims
Priority Applications (6)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP20735571.0A EP3993871A1 (en) | 2019-07-02 | 2020-06-30 | Cpla2e inducing agents and uses thereof |
CA3145446A CA3145446A1 (en) | 2019-07-02 | 2020-06-30 | Cpla2e inducing agents and uses thereof |
JP2021577986A JP2022544740A (en) | 2019-07-02 | 2020-06-30 | cPLA2e inducers and uses thereof |
AU2020299718A AU2020299718A1 (en) | 2019-07-02 | 2020-06-30 | cPLA2e inducing agents and uses thereof |
US17/624,178 US20220233722A1 (en) | 2019-07-02 | 2020-06-30 | cPLA2e INDUCING AGENTS AND USES THEREOF |
CN202080049050.2A CN114222818A (en) | 2019-07-02 | 2020-06-30 | cPLA2e inducer and use thereof |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP19382563.5 | 2019-07-02 | ||
EP19382563 | 2019-07-02 |
Publications (1)
Publication Number | Publication Date |
---|---|
WO2021001377A1 true WO2021001377A1 (en) | 2021-01-07 |
Family
ID=67402895
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/EP2020/068414 WO2021001377A1 (en) | 2019-07-02 | 2020-06-30 | cPLA2e INDUCING AGENTS AND USES THEREOF |
Country Status (7)
Country | Link |
---|---|
US (1) | US20220233722A1 (en) |
EP (1) | EP3993871A1 (en) |
JP (1) | JP2022544740A (en) |
CN (1) | CN114222818A (en) |
AU (1) | AU2020299718A1 (en) |
CA (1) | CA3145446A1 (en) |
WO (1) | WO2021001377A1 (en) |
Citations (19)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO1992001070A1 (en) | 1990-07-09 | 1992-01-23 | The United States Of America, As Represented By The Secretary, U.S. Department Of Commerce | High efficiency packaging of mutant adeno-associated virus using amber suppressions |
US5139941A (en) | 1985-10-31 | 1992-08-18 | University Of Florida Research Foundation, Inc. | AAV transduction vectors |
US5173414A (en) | 1990-10-30 | 1992-12-22 | Applied Immune Sciences, Inc. | Production of recombinant adeno-associated virus vectors |
WO1993003769A1 (en) | 1991-08-20 | 1993-03-04 | THE UNITED STATES OF AMERICA, represented by THE SECRETARY, DEPARTEMENT OF HEALTH AND HUMAN SERVICES | Adenovirus mediated transfer of genes to the gastrointestinal tract |
WO1995017183A1 (en) * | 1993-12-23 | 1995-06-29 | Eli Lilly And Company | Use of pla2 inhibitors as treatment for alzheimer's disease |
WO2001085938A1 (en) | 2000-05-11 | 2001-11-15 | Institut National De La Recherche Agronomique | Modified es cells and es cell-specific gene |
WO2003076601A1 (en) | 2002-03-08 | 2003-09-18 | Vivalis | Avian cell lines for the production of useful substances |
WO2005042728A2 (en) | 2003-11-03 | 2005-05-12 | Probiogen Ag | Immortalized avian cell lines for virus production |
WO2006108846A1 (en) | 2005-04-11 | 2006-10-19 | Vivalis | Process of manufacturing viral vaccines in suspension avian embryonic derived stem cell lines |
WO2008129058A1 (en) | 2007-04-24 | 2008-10-30 | Vivalis | Duck embryonic derived stem cell lines for the production of viral vaccines |
WO2008142124A1 (en) | 2007-05-21 | 2008-11-27 | Vivalis | Recombinant protein production in avian ebx® cells |
US20120121571A1 (en) * | 2008-06-30 | 2012-05-17 | The Regents Of The University Of Michigan | Lysosomal Phospholipase A2 (LPLA2) Activity as a Diagnostic and Therapeutic Target for Identifying and treating Systemic Lupus Erythematosis |
US20140206752A1 (en) | 2011-05-17 | 2014-07-24 | Moderna Therapeutics, Inc. | Engineered nucleic acids and methods of use thereof for non-human vertebrates |
WO2014164253A1 (en) | 2013-03-09 | 2014-10-09 | Moderna Therapeutics, Inc. | Heterologous untranslated regions for mrna |
US20150086614A1 (en) | 2012-04-02 | 2015-03-26 | Moderna Therapeutics, Inc. | Modified polynucleotides for the production of cytoplasmic and cytoskeletal proteins |
WO2016011306A2 (en) | 2014-07-17 | 2016-01-21 | Moderna Therapeutics, Inc. | Terminal modifications of polynucleotides |
WO2016011226A1 (en) | 2014-07-16 | 2016-01-21 | Moderna Therapeutics, Inc. | Chimeric polynucleotides |
WO2016014846A1 (en) | 2014-07-23 | 2016-01-28 | Moderna Therapeutics, Inc. | Modified polynucleotides for the production of intrabodies |
US20160304552A1 (en) | 2013-12-13 | 2016-10-20 | Moderna Therapeutics, Inc. | Modified nucleic acid molecules and uses thereof |
Family Cites Families (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
IL101507A0 (en) * | 1991-04-17 | 1992-12-30 | Lilly Co Eli | Compounds,vectors and methods for expressing human,cytosolic phospholipase a2 |
WO2002031160A2 (en) * | 2000-10-10 | 2002-04-18 | Bayer Aktiengesellschaft | Regulation of human phospholipase a2-like enzyme |
CA2388659A1 (en) * | 2002-05-31 | 2003-11-30 | Mcgill University | Phospholipase a2 expression and activity and use thereof for diagnosis, prognostication, prevention and treatment of neural inflammatory and demyelinating disease |
-
2020
- 2020-06-30 US US17/624,178 patent/US20220233722A1/en active Pending
- 2020-06-30 CA CA3145446A patent/CA3145446A1/en active Pending
- 2020-06-30 WO PCT/EP2020/068414 patent/WO2021001377A1/en unknown
- 2020-06-30 AU AU2020299718A patent/AU2020299718A1/en active Pending
- 2020-06-30 EP EP20735571.0A patent/EP3993871A1/en active Pending
- 2020-06-30 CN CN202080049050.2A patent/CN114222818A/en active Pending
- 2020-06-30 JP JP2021577986A patent/JP2022544740A/en active Pending
Patent Citations (19)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5139941A (en) | 1985-10-31 | 1992-08-18 | University Of Florida Research Foundation, Inc. | AAV transduction vectors |
WO1992001070A1 (en) | 1990-07-09 | 1992-01-23 | The United States Of America, As Represented By The Secretary, U.S. Department Of Commerce | High efficiency packaging of mutant adeno-associated virus using amber suppressions |
US5173414A (en) | 1990-10-30 | 1992-12-22 | Applied Immune Sciences, Inc. | Production of recombinant adeno-associated virus vectors |
WO1993003769A1 (en) | 1991-08-20 | 1993-03-04 | THE UNITED STATES OF AMERICA, represented by THE SECRETARY, DEPARTEMENT OF HEALTH AND HUMAN SERVICES | Adenovirus mediated transfer of genes to the gastrointestinal tract |
WO1995017183A1 (en) * | 1993-12-23 | 1995-06-29 | Eli Lilly And Company | Use of pla2 inhibitors as treatment for alzheimer's disease |
WO2001085938A1 (en) | 2000-05-11 | 2001-11-15 | Institut National De La Recherche Agronomique | Modified es cells and es cell-specific gene |
WO2003076601A1 (en) | 2002-03-08 | 2003-09-18 | Vivalis | Avian cell lines for the production of useful substances |
WO2005042728A2 (en) | 2003-11-03 | 2005-05-12 | Probiogen Ag | Immortalized avian cell lines for virus production |
WO2006108846A1 (en) | 2005-04-11 | 2006-10-19 | Vivalis | Process of manufacturing viral vaccines in suspension avian embryonic derived stem cell lines |
WO2008129058A1 (en) | 2007-04-24 | 2008-10-30 | Vivalis | Duck embryonic derived stem cell lines for the production of viral vaccines |
WO2008142124A1 (en) | 2007-05-21 | 2008-11-27 | Vivalis | Recombinant protein production in avian ebx® cells |
US20120121571A1 (en) * | 2008-06-30 | 2012-05-17 | The Regents Of The University Of Michigan | Lysosomal Phospholipase A2 (LPLA2) Activity as a Diagnostic and Therapeutic Target for Identifying and treating Systemic Lupus Erythematosis |
US20140206752A1 (en) | 2011-05-17 | 2014-07-24 | Moderna Therapeutics, Inc. | Engineered nucleic acids and methods of use thereof for non-human vertebrates |
US20150086614A1 (en) | 2012-04-02 | 2015-03-26 | Moderna Therapeutics, Inc. | Modified polynucleotides for the production of cytoplasmic and cytoskeletal proteins |
WO2014164253A1 (en) | 2013-03-09 | 2014-10-09 | Moderna Therapeutics, Inc. | Heterologous untranslated regions for mrna |
US20160304552A1 (en) | 2013-12-13 | 2016-10-20 | Moderna Therapeutics, Inc. | Modified nucleic acid molecules and uses thereof |
WO2016011226A1 (en) | 2014-07-16 | 2016-01-21 | Moderna Therapeutics, Inc. | Chimeric polynucleotides |
WO2016011306A2 (en) | 2014-07-17 | 2016-01-21 | Moderna Therapeutics, Inc. | Terminal modifications of polynucleotides |
WO2016014846A1 (en) | 2014-07-23 | 2016-01-28 | Moderna Therapeutics, Inc. | Modified polynucleotides for the production of intrabodies |
Non-Patent Citations (64)
Title |
---|
"Adenovirus: Methods and Protocols", 2014, HUMANA PRESS |
"Viral Vectors for Gene Therapy, Methods and Protocols. Series: Methods in Molecular Biology", vol. 737, 2011, HUMANA PRESS |
A. RICOBARAZA, NEUROPSYCHOPHARMACOLOGY, vol. 34, 2009, pages 1721 - 1732 |
ADACHI M. ET AL., HUM MOL GENET., vol. 14, no. 23, 2005, pages 3709 - 3722 |
ALTSCHUL SF ET AL., BIOINFORMATICS, vol. 21, no. 8, 2005, pages 1451 - 6 |
ALTSCHUL SF ET AL., NUCLEIC ACIDS RES., vol. 25, no. 17, 1997, pages 3389 - 402 |
ARNOLD ET AL., J. PHYSIOL., vol. 564, 2005, pages 3 - 19 |
BERNSBOHENZKY: "Advances in Virus Research", vol. 32, 1987, ACADEMIC PRESS, INC., pages: 243 - 307 |
BIINNING H. ET AL.: "Recent developments in adeno-associated virus technology", J. GENE MED., vol. 10, 2008, pages 717 - 733 |
BUNNING H ET AL., J GENE MED, vol. 10, 2008, pages 717 - 733 |
CAPESTRANO M. ET AL., JOURNAL OF CELL SCIENCE, vol. 127, 2014, pages 977 - 993 |
CARTER, B. J., CURRENT OPINION IN BIOTECHNOLOGY, vol. 3, 1992, pages 533 - 539 |
CESCA ET AL., PROG. NEUROBIOL., vol. 91, 2010, pages 313 - 348 |
CHTARTO, NEUROSCI LETT., vol. 352, 2003, pages 155 - 158 |
COLIN P.J. GLOVER ET AL: "Adenoviral-Mediated, High-Level, Cell-Specific Transgene Expression: A SYN1-WPRE Cassette Mediates Increased Transgene Expression with No Loss of Neuron Specificity", MOLECULAR THERAPY, vol. 5, no. 5, 1 May 2002 (2002-05-01), US, pages 509 - 516, XP055729217, ISSN: 1525-0016, DOI: 10.1006/mthe.2002.0588 * |
DATABASE Geneseq [online] 11 June 2007 (2007-06-11), "Full length human cDNA useful for treating neurological disease Seq 894.", XP002800270, retrieved from EBI accession no. GSN:ADR07388 Database accession no. ADR07388 * |
DATABASE Geneseq [online] 15 June 2007 (2007-06-15), "Human protein useful for treating neurological disease Seq 2850.", XP002800271, retrieved from EBI accession no. GSP:ADR09344 Database accession no. ADR09344 * |
D'HOOGEDEYN, BRAIN RESEARCH REVIEWS, vol. 36, no. 1, 2001, pages 60 - 90 |
DITTGEN T ET AL., PROC NATL ACAD SCI USA, vol. 101, 2004, pages 18206 - 18211 |
DUROCHER, Y.S. PERRETA. KAMEN., NUCLEIC ACIDS RESEARCH, vol. 30, no. 2, 2002, pages 9e - 9 |
FORSS-PETTERS S ET AL., NEURON, vol. 5, no. 2, 1990, pages 187 - 97 |
GINTY ET AL., SCIENCE, vol. 260, 1993, pages 238 - 241 |
GLASEREDMUND M.HENDRIK VAN DER LOOS, JOURNAL OF NEUROSCIENCE METHODS, vol. 4, no. 2, 1981, pages 117 - 25 |
GLOSTER A. ET AL., J NEUROSC, vol. 14, 1994, pages 7319 |
GRAY S.J. ET AL., HUM GENE THER., vol. 22, no. 9, 2011, pages 1143 - 1153 |
HARDINGHAM ET AL., NAT. NEUROSCI., vol. 5, 2002, pages 405 - 414 |
HEILBRONN R.WEGER S.: "Drug Delivery, Handbook of Experimental Pharmacology", vol. 197, 2010, SPRINGER-VERLAG, article "Viral Vectors for Gene Transfer: Current Status of Gene Therapeutics", pages: 143 - 170 |
HUFF ET AL., J. NEUROSCI., vol. 26, 2006, pages 1616 - 1623 |
HUSSAIN ZAHIR ET AL: "Phosphatidylserine-stimulated production ofN-acyl-phosphatidylethanolamines by Ca2+-dependentN-acyltransferase", BIOCHIMICA ET BIOPHYSICA ACTA (BBA) - MOLECULAR AND CELL BIOLOGY OF LIPIDS, ELSEVIER, AMSTERDAM, NL, vol. 1863, no. 5, 12 February 2018 (2018-02-12), pages 493 - 502, XP085359046, ISSN: 1388-1981, DOI: 10.1016/J.BBALIP.2018.02.002 * |
KIIGLER S ET AL., GENE THER., vol. 10, no. 4, 2003, pages 337 - 47 |
KOTIN, R. M., HUMAN GENE THERAPY, vol. 5, 1994, pages 793 - 801 |
LEBKOWSKI ET AL., MOLEC. CELL. BIOL., vol. 8, 1988, pages 3988 - 3996 |
LUKASHCHUK V. ET AL., MOL THER METHODS CLIN DEV., vol. 3, 2016, pages 15055 |
M. CAPESTRANO ET AL: "Cytosolic phospholipase A2 drives recycling through the clathrin-independent endocytic route", JOURNAL OF CELL SCIENCE, vol. 127, no. 5, 1 March 2014 (2014-03-01), Cambridge, pages 977 - 993, XP055727435, ISSN: 0021-9533, DOI: 10.1242/jcs.136598 * |
MAIA LF ET AL., SCIENCE TRANSLATIONAL MEDICINE, vol. 5, no. 194, 2013, pages 94 - 194 |
MANDALROSSI, NAT. PROTOC., vol. 8, no. 3, 2013, pages 568 - 82 |
MATSUZAKI Y. ET AL., J NEUROSCI METHODS, vol. 223, 2014, pages 133 - 143 |
MEGIAS, M.Z. EMRITF FREUNDAI GULYAS, NEUROSCIENCE, vol. 102, no. 3, 2001, pages 527 - 40 |
MUZYCZKA, N., CURRENT TOPICS IN MICROBIOL. AND IMMUNOL., vol. 158, 1992, pages 97 - 129 |
NAZIA MAROOF ET AL: "Reductions in Endocannabinoid Levels and Enhanced Coupling of Cannabinoid Receptors in the Striatum are Accompanied by Cognitive Impairments in the A[beta]PPswe/PS1[Delta]E9 Mouse Model of Alzheimer's Disease", JOURNAL OF ALZHEIMER'S DISEASE, vol. 42, no. 1, 11 August 2014 (2014-08-11), NL, pages 227 - 245, XP055728805, ISSN: 1387-2877, DOI: 10.3233/JAD-131961 * |
NEEDLEMANWUNSCH, J MOL BIOL., vol. 48, no. 3, 1970, pages 443 - 53 |
OGURA Y ET AL., NAT. CHEM. BIOL., vol. 12, no. 9, 2016, pages 669 - 671 |
OGURA Y. ET AL., NAT CHEM BIOL., vol. 12, no. 9, 2016, pages 669 - 671 |
PAULK ET AL., MOL THER., vol. 26, no. 1, 2018, pages 289 - 303 |
PÉREZ-GONZÁLEZ MARTA ET AL: "PLA2G4E, a candidate gene for resilience in Alzheimer s disease and a new target for dementia treatment", PROGRESS IN NEUROBIOLOGY, ELSEVIER, AMSTERDAM, NL, vol. 191, 5 May 2020 (2020-05-05), XP086184967, ISSN: 0301-0082, [retrieved on 20200505], DOI: 10.1016/J.PNEUROBIO.2020.101818 * |
RADDE ET AL., EMBO REPORTS, vol. 7, no. 9, 2006, pages 940 - 946 |
RAO ET AL., NAT. NEUROSCI., vol. 9, 2006, pages 887 - 895 |
SANFTNER, MOL THER., vol. 13, 2006, pages 167 - 174 |
SATO ET AL., BRAIN RES., vol. 872, 2000, pages 219 - 222 |
SERNEELS L ET AL., SCIENCE, vol. 324, no. 5927, 2009, pages 639 - 642 |
SINGH NR ET AL., EXP NEUROL., vol. 196, no. 2, 2005, pages 224 - 234 |
SMITH ET AL., MOLECULAR THERAPY, vol. 17, no. 11, 2009, pages 1888 - 1896 |
SMITHWATERMAN, J THEOR BIOL., vol. 91, no. 2, 1981, pages 379 - 80 |
SMRITI SULTANA BINTE MUSTAFIZ ET AL: "The role of intracellular anionic phospholipids in the production of N -acyl-phosphatidylethanolamines by cytosolic phospholipase A2[epsilon]", JOURNAL OF BIOCHEMISTRY, vol. 165, no. 4, 1 April 2019 (2019-04-01), GB, pages 343 - 352, XP055727438, ISSN: 0021-924X, DOI: 10.1093/jb/mvy104 * |
TWYMAN RM, J MOL NEUROSCI, vol. 8, no. 1, 1997, pages 63 - 73 |
URABE ET AL., HUM. GENE THER., vol. 13, 2002, pages 1935 - 1943 |
VERCAUTEREN ET AL., MOL THER., vol. 24, no. 6, 2016, pages 1042 - 1049 |
VINCENT ET AL.: "Vaccines 90", 1990, COLD SPRING HARBOR LABORATORY PRESS |
WANG L ET AL., MOL THER., vol. 23, no. 12, 2015, pages 1877 - 87 |
WEBER P ET AL., EUR J NEUROSCI, vol. 14, 2001, pages 1777 |
WEISSIG, H. ET AL., BIOCHEM. J., vol. 290, 1993, pages 503 - 508 |
YUJI OGURA ET AL: "A calcium-dependent acyltransferase that produces N-acyl phosphatidylethanolamines", NATURE CHEMICAL BIOLOGY, vol. 12, no. 9, 11 July 2016 (2016-07-11), New York, pages 669 - 671, XP055727421, ISSN: 1552-4450, DOI: 10.1038/nchembio.2127 * |
ZHANG Y Y ET AL: "Effect of phosphatidylserine on memory in patients and rats with Alzheimer's disease", GENETICS AND MOLECULAR RESEARCH, FUNDACAO DE PESQUISAS CIENTIFICAS DE RIBEIRAO PRETO, BR, vol. 14, no. 3, 1 January 2015 (2015-01-01), pages 9325 - 9333, XP009506001, ISSN: 1676-5680, DOI: 10.4238/2015.AUGUST.10.13 * |
ZINN E ET AL., CELL REP., vol. 12, no. 6, 2015, pages 1056 - 68 |
Also Published As
Publication number | Publication date |
---|---|
CA3145446A1 (en) | 2021-01-07 |
AU2020299718A1 (en) | 2022-02-24 |
CN114222818A (en) | 2022-03-22 |
JP2022544740A (en) | 2022-10-21 |
US20220233722A1 (en) | 2022-07-28 |
EP3993871A1 (en) | 2022-05-11 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
KR20200039617A (en) | Adeno-associated virus capsid variants and methods of use thereof | |
Cain et al. | Gene therapy corrects brain and behavioral pathologies in CLN6-Batten disease | |
ES2871527T3 (en) | Adeno-associated virus virion for use in the treatment of epilepsy | |
WO2019068854A1 (en) | Gene therapy of neurodegenerative diseases using aav vectors | |
Kim et al. | Rescue of behavioral and electrophysiological phenotypes in a Pitt-Hopkins syndrome mouse model by genetic restoration of Tcf4 expression | |
EP2212348B1 (en) | Materials and methods for gene mediated therapy of psychiatric disorders | |
Gabery et al. | Characterization of a rat model of Huntington’s disease based on targeted expression of mutant huntingtin in the forebrain using adeno‐associated viral vectors | |
US20220233722A1 (en) | cPLA2e INDUCING AGENTS AND USES THEREOF | |
US20220098617A1 (en) | Ascl1 vector | |
US11364309B2 (en) | Neuronal enhancers | |
US11597936B2 (en) | Recombinant Dgkk gene for fragile X syndrome gene therapy | |
US20220098254A1 (en) | NEUROD1 and DLX2 VECTOR | |
US20240216536A1 (en) | Secreted ube3a for treatment of neurological disorders | |
US20220364118A1 (en) | Targeting deltafosb for treatment of dyskinesia | |
KR20240099360A (en) | Nucleic acid constructs, viral vectors, and viral particles | |
US20220106614A1 (en) | Dlx2 vector | |
US20180327778A1 (en) | Animal models for brain inflammation and white matter degeneration | |
Aimiuwu | Modeling Gene Therapy for Intractable Developmental and Epileptic Encephalopathy | |
Miller | Utilizing gene therapy methods to probe the genetic requirements to prevent spinal muscular atrophy | |
CA3195052A1 (en) | Nucleic acid constructs, viral vectors and viral particles | |
Sindarto | An investigation into the role of mesoaccumbal GABAA receptor a2 subunit in mediating cocaine-facilitated conditioned behaviours using the RNA interference system | |
CN116723868A (en) | Methods of treating neurological disorders | |
Al-Rafiah | PLASTIN3 AS A THERAPEUTIC TARGET IN SPINAL MUSCULAR ATROPHY | |
Moghaddam | Virus-Mediated Delivery of the Fmr1 Gene as a Tool for the Treatment of Fragile X Syndrome |
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: 20735571 Country of ref document: EP Kind code of ref document: A1 |
|
ENP | Entry into the national phase |
Ref document number: 2021577986 Country of ref document: JP Kind code of ref document: A |
|
ENP | Entry into the national phase |
Ref document number: 3145446 Country of ref document: CA |
|
NENP | Non-entry into the national phase |
Ref country code: DE |
|
ENP | Entry into the national phase |
Ref document number: 2020735571 Country of ref document: EP Effective date: 20220202 |
|
ENP | Entry into the national phase |
Ref document number: 2020299718 Country of ref document: AU Date of ref document: 20200630 Kind code of ref document: A |