WO2023246806A1 - Modèle de maladie et son utilisation - Google Patents
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Classifications
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- 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
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/027—New or modified breeds of vertebrates
-
- 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
Definitions
- the DIR and/or functionally active fragment thereof comprises any one of SEQ ID NOs: 1-2 and 4-5 The amino acid sequence shown.
- the gene encoding the DIR and/or functionally active fragment thereof is homozygous or heterozygous.
- the gene encoding the DIR and/or functionally active fragment thereof comprises the nucleic acid sequence shown in any one of SEQ ID NOs: 12-15.
- the non-human animal is a knock-in non-human animal, or is derived from a knock-in non-human animal.
- the expression of the DIR and/or functionally active fragments thereof is regulated by corresponding endogenous regulatory elements in the genome of the non-human animal.
- the non-human animal is selected from the group consisting of: mouse, rat, rabbit, ape, monkey, pig, cow, sheep, horse, chicken, dog, alpaca, and cat.
- the portions include organs, tissues and/or cells of the non-human animal.
- the portion includes body fluids.
- the plasma includes exosomes.
- the portion includes the brain or a portion thereof.
- the portion includes the olfactory mucosa.
- the portion includes a portion of the central nervous system, a portion of the peripheral nervous system, skin tissue, muscle tissue, and/or internal organs.
- the central nervous system includes the spinal cord.
- the portion includes cells, and the cells include cells selected from the group consisting of primary neurons, microglia, astrocytes, oligodendrocytes, macrophages , perivascular epithelioid cells, B cells, T cells, adult stem cells, NK cells, totipotent stem cells, unipotent stem cells, embryonic stem cells, induced pluripotent stem cells and gametes.
- the gametes include sperm and/or oocytes.
- the portion is unable to develop into a complete non-human animal.
- the application provides an isolated cell.
- the cells comprise and/or express an intron-retentive cleavage product DIR of DNA damage-inducible transcript 4-like transcript (DDIT4L) and/or a functionally active fragment thereof.
- DIR DNA damage-inducible transcript 4-like transcript
- the DIR is of human origin.
- the DIR and/or functionally active fragment thereof comprises the amino acid sequence set forth in any one of SEQ ID NOs: 1-2 and 4-5.
- the gene encoding the DIR and/or functionally active fragment thereof is homozygous or heterozygous.
- the cells are, or are derived from, gene-edited cells.
- the cell is a knock-in cell, or is derived from a knock-in cell.
- the cells are non-human animal cells.
- the non-human animal is selected from the group consisting of: mouse, rat, rabbit, ape, monkey, pig, cow, sheep, horse, chicken, dog, alpaca, and cat.
- the cells include cells selected from the group consisting of neuroblastoma cells, glioma cells, primary neurons, microglia, astrocytes, oligodendrocytes cells, macrophages, perivascular epithelioid cells, B cells, T cells, adult stem cells, NK cells, totipotent stem cells, unipotent stem cells, embryonic stem cells, induced pluripotent stem cells and gametes.
- the DIR is of human origin.
- the DIR and/or functionally active fragments thereof comprise the amino acid sequence set forth in any one of SEQ ID NOs: 1-2 and 4-5.
- the gene encoding the DIR and/or functionally active fragment thereof is homozygous or heterozygous.
- the gene encoding the DIR and/or functionally active fragment thereof comprises the nucleic acid sequence shown in any one of SEQ ID NOs: 12-15.
- the tissue is, or is derived from, a gene-edited tissue.
- the tissue is a knock-in tissue, or is derived from a knock-in tissue.
- expression of the DIR and/or functionally active fragments thereof is regulated by corresponding endogenous regulatory elements in the genome of the tissue.
- the tissue is that of a non-human animal.
- the non-human animal is a non-human mammal.
- the non-human animal is selected from the group consisting of: mouse, rat, rabbit, ape, monkey, pig, cow, sheep, horse, chicken, dog, alpaca, and cat.
- the non-human animal is a rodent or primate.
- the tissue includes body fluids.
- the body fluid is selected from the group consisting of blood, plasma, serum, urine, sweat, tears, saliva, semen, and cerebrospinal fluid.
- the plasma includes exosomes.
- the tissue includes the brain or a portion thereof.
- the portion of the brain includes a brain site selected from the group consisting of the olfactory bulb, amygdala, basal ganglia, hippocampus, thalamus, hypothalamus, cerebral cortex, medulla oblongata, and cerebellum.
- the tissue includes the olfactory mucosa.
- the tissue includes a portion of the central nervous system, a portion of the peripheral nervous system, skin tissue, muscle tissue, and/or internal organs.
- the central nervous system includes the spinal cord.
- the tissue is unable to develop into a complete non-human animal.
- the application provides a cell line or cell culture derived from a non-human animal described herein or a part thereof, derived from a progeny described herein, derived from a cell described herein, and/or derived from the organizations described in this application.
- the cell culture is a primary cell culture.
- the cell culture includes organoids.
- the application provides a composition.
- the composition includes the non-human animals described in the application or parts thereof, the progeny described in the application, the cells described in the application, the tissues described in the application, and/or the cell lines or original cells described in the application. Generation of cell cultures.
- the present application provides a kit.
- the kit includes the non-human animals described in the application or parts thereof, the progeny described in the application, the cells described in the application, the tissues described in the application, and/or the cell lines or original cells described in the application. generation cell culture; and one or more additional components selected from the group consisting of: assay buffers, controls, substrates, standards, test materials, laboratory supplies, equipment, instruments, cells, organs, tissues and user manual or instructions.
- the present application provides a method of screening and/or identifying substances, devices and/or compositions for treating, preventing and/or prognosing neurological diseases or disorders, the method comprising combining candidate substances, Devices and/or compositions are administered to non-human animals described herein or parts thereof, progeny described herein, cells described herein, tissues described herein, and/or cells described herein lines or primary cell cultures and determine the effect of the candidate substances, devices and/or compositions on one or more indications of the neurological disease or disorder.
- the present application provides a method of designing substances, devices and/or compositions for the treatment, prevention and/or prognosis of neurological diseases or disorders, the method comprising: (i) combining a candidate substance, Devices and/or compositions are administered to non-human animals described herein or parts thereof, progeny described herein, cells described herein, tissues described herein, and/or cells described herein lines or primary cell cultures; (ii) determine the effect of the candidate substances, devices and/or compositions on one or more indications of the neurological disease or disorder; (iii) select those capable of preventing, Candidate substances, devices and/or compositions that inhibit and/or alleviate one or more of the indications of the neurological disease or disorder; (iv) modify the candidate substances, devices and/or compositions of step (iii) Or the structure and/or composition of the composition to obtain modified substances, devices and/or compositions that are useful in treating, preventing and/or prognosing the neurological disease or There is improvement activity in terms of symptoms.
- the neurological disease or disorder includes cognitive impairment.
- the neurological disease or disorder includes a neurodegenerative disease or disorder.
- the neurological disease or disorder includes mild cognitive impairment (MCI) and/or Alzheimer's disease.
- MCI mild cognitive impairment
- Alzheimer's disease the neurological disease or disorder includes mild cognitive impairment (MCI) and/or Alzheimer's disease.
- the APP fragment includes: sAPP, CTF- ⁇ , CTF-a and/or AICD.
- the innate immune cells include T cells, B cells, NK cells, macrophages and/or dendritic cells.
- the vascular dysfunction includes amyloid angiopathy and/or blood-brain barrier (BBB) damage.
- BBB blood-brain barrier
- the device includes a medical device.
- the present application provides a method for screening biological targets for diagnosing and/or monitoring neurological diseases or conditions. and/or a method of biomarkers, the method comprising: measuring the non-human animals or parts thereof described herein before and after the occurrence of one or more relevant indicators of the neurological disease or disorder, respectively. , the progeny described in the application, the cells described in the application, the tissue described in the application, and/or the presence of each substance in the sample obtained from the cell line or primary cell culture described in the application and/or level, to identify a substance whose presence and/or level changes before and after the occurrence of the indication, and to identify the substance as the biological target and/or the biomarker.
- the one or more indicators of the neurological disease or disorder include: expression and/or accumulation of A ⁇ ; expression and/or accumulation of APP fragments; phosphorylation of Tau protein ation; oligomerization and/or aggregation of Tau protein; changes in brain morphology and/or brain weight; formation of brain neurofibrillary tangles; learning function, memory function, cognitive function, sensory function, motor function, emotional function and /or changes in synaptic function; brain damage; neuronal loss, necrotic body formation, neuronal death, changes in neuronal activity, neuronal apoptosis, neuronal necrosis, neuronal programmed cell necrosis, Neu-N positive nerves neurodegeneration and/or neurodegeneration; gliosis, microglial activation, astrocyte activation and/or inflammatory response; oxidative stress and/or mitochondrial dysfunction; vascular dysfunction; misfolded proteins Failure of degradation and/or autophagy; and/or changes in brain glucose
- the neurological disease or disorder includes cognitive impairment.
- the neurological disease or disorder includes a neurodegenerative disease or disorder.
- non-human animals described in the application or parts thereof, the progeny described in the application, the cells described in the application, the tissues described in the application, and/or the cell lines or primary cells described in the application Use of cell cultures in the preparation of identification and/or screening systems for the identification and/or screening of substances, devices, compositions for the treatment, diagnosis, prevention, monitoring and/or prediction of neurological diseases or conditions , biological targets and/or biomarkers.
- the non-human animal or part thereof, the progeny, the cells, the tissue, and/or the cell line or primary cell culture can be used for screening for treatment, diagnosis, prevention , substances, devices, compositions, biological targets and/or biomarkers for monitoring and/or predicting neurological diseases or conditions.
- the present application provides a method for preparing the non-human animal or part thereof, the cell described in the present application or the tissue described in the present application, the method comprising making the non-human animal or part thereof , cells or tissues containing and/or expressing the DIR and/or functionally active fragments thereof.
- the present application provides a method for preparing a disease model, the method comprising causing a non-human animal or part, cell or tissue thereof to contain and/or express DIR and/or a functionally active fragment thereof.
- the neurological disease or disorder includes cognitive impairment.
- the neurological disease or disorder includes a neurodegenerative disease or disorder.
- the neurological disease or disorder includes mild cognitive impairment (MCI) and/or Alzheimer's disease.
- MCI mild cognitive impairment
- Alzheimer's disease the neurological disease or disorder includes mild cognitive impairment (MCI) and/or Alzheimer's disease.
- the DIR and/or functionally active fragments thereof comprise the amino acid sequence shown in any one of SEQ ID NOs: 1-2 and 4-5.
- the nucleic acid sequence encoding the DIR and/or its functionally active fragment includes the nucleic acid sequence shown in any one of SEQ ID NO: 12-15.
- the knock-in further includes culturing the cell or tissue under conditions in which the one or more RNA sequences hybridize to the target sequence, and the Cas protein is present in the target sequence. The sequence is cleaved after said hybridization with said RNA sequence.
- the Cas protein is Cas9.
- Figure 4 shows exemplary behavioral phenotypic characteristics of disease models of the present application.
- Figure 6 shows that the growth, metabolism and motor functions of the disease model of the present application are not changed.
- Figure 7 shows the DIR expression of the disease model of the present application under hypoxic conditions.
- Figure 8E the expression of A ⁇ and p-Tau was significantly reduced in the hippocampal dentate gyrus of homozygous DIR-KI mice treated with DIR antibodies. *,p ⁇ 0.05.**,p ⁇ 0.01.
- homologous recombination generally refers to genetic recombination in which nucleotide sequences are exchanged between two similar or identical DNA molecules (called homologous sequences or homology arms).
- CRISPR/Cas9 system generally refers to a tool for site-specific genome targeting in an organism.
- it could be a type II CRISPR/Cas system, which is a system that uses non-coding RNA to guide the Cas9 nuclease to induce site-specific DNA breaks.
- This DNA damage can be repaired through cellular DNA repair mechanisms, or through the non-homologous end joining DNA repair pathway (NHEJ, non-homologous end joining DNA repair pathway) or homology directed repair (HDR, homology directed repair) pathway.
- NHEJ non-homologous end joining DNA repair pathway
- HDR homology directed repair
- knocking-in or “knock-in” generally refers to a genetic modification process that involves the replacement of DNA sequence information at a specific locus, or the replacement of foreign sequences with Insert into the endogenous locus.
- the knock-in may involve inserting a gene sequence into a specific site and may therefore be a "targeted” insertion.
- embryonic stem cells or "ES cells” generally refers to pluripotent stem cells derived from the inner cell mass (ICM) of the blastocyst, the early pre-implantation embryo of mammals. It can be cultured in vitro, and after it is embedded/injected into the cavity of a normal blastocyst, it can be induced to resume the normal embryonic development program to differentiate into various cell types of the adult animal body, including germ cells.
- ICM inner cell mass
- zygote generally refers to a eukaryotic cell formed by a fertilization event between gametes (eg, mammalian eggs and sperm).
- no significant difference generally means that the difference between two values or two objects is not material. For example, when comparing two values, if the difference is less than about 10%, such as less than about 9%, less than about 8%, less than about 7%, less than about 6%, less than about 5.5%, less than about 5%, less than about 4.5% , less than about 3.5%, less than about 3.5%, less than about 2.5%, less than about 2%, less than about 1.5%, less than about 1% or even lower, it can be regarded as no significant difference.
- a ⁇ oligomers generally refers to soluble amyloid ⁇ (A ⁇ ) peptide aggregates, which often form clumps or plaques.
- a ⁇ oligomers may be dimers, trimers or other multimers of A ⁇ peptide.
- a ⁇ plaque or "amyloid plaque” generally refers to fibrillar aggregates of A ⁇ peptides (e.g., A ⁇ 42 and/or A ⁇ 40) in which many copies of the A ⁇ peptide stick together to form fibrils or Fibrous deposits (eg, plaque).
- the term "large accumulation of A ⁇ peptide” generally refers to the fact that the formation of A ⁇ oligomers or amyloid plaques can be detected using common detection methods or tools, such as by staining with specific A ⁇ antibodies.
- a ⁇ or "amyloid ⁇ ” generally refers to the intramembrane protein water of amyloid precursor protein (APP).
- Amyloid beta peptide produced by hydrolysis. Sequential proteolytic reactions by ⁇ - and ⁇ -secretase produce A ⁇ peptides of different lengths, including A ⁇ 40 and A ⁇ 42. Its two superhydrophobic residues give A ⁇ 42 a higher tendency to aggregate into soluble oligomers and insoluble deposits.
- a ⁇ exists in a variety of aggregated forms, ranging from dimers to ⁇ -pleated sheet-like fibers in dense neuritic plaques. Excessive A ⁇ can induce a variety of pathological processes. A ⁇ can impair neuronal and glial cell function, synaptic physiology, neurotransmission, and cognitive function.
- hypophosphorylated refers to a state of abnormal phosphorylation at one or more sites.
- phosphorylation of Tau protein was found to cause negative regulation of its activity in promoting microtubule assembly, and abnormally hyperphosphorylated Tau is considered to be the main component of PHFs in Alzheimer's disease (AD) patients.
- AD Alzheimer's disease
- Normal brain contains 2-3 moles of phosphate per mole of Tau.
- Studies of human brain biopsies have shown that some serine and threonine residues of Tau are typically phosphorylated at low substoichiometric levels.
- the phosphorylation level of Tau isolated from brain autopsies of Alzheimer's disease (AD) patients is 3 to 4 times higher than that of normal human brains.
- Tau phosphorylation at different sites has different effects on its biological functions and pathogenic effects.
- Studies on the combination of highly phosphorylated Tau and normal Tau have shown that Ser199/Ser202/Thr205, Thr212, Thr231/Ser235, Ser262/Ser356 and Ser422 are key phosphorylation sites. The phosphorylation of these sites converts Tau into a Inhibitory molecules that dissociate microtubule-binding proteins from normal microtubules.
- neuronal loss generally refers to a reduction or weakening of the number or function of neuronal cells in an organism. Loss of neurons may manifest as neuronal cell death.
- Tau protein generally refers to the microtubule-associated protein tau (MAPT) that stabilizes microtubules.
- AGT microtubule-associated protein tau
- Tau protein is abundant in neurons of the CNS and less common elsewhere, but is also expressed at very low levels in CNS astrocytes and oligodendrocytes.
- Tau protein can control microtubule stability in two ways: through different isoforms and through different phosphorylation levels.
- the NCBI registration ID for human Tau isoform 1 is NP_058519.3.
- AD Alzheimer's disease may be related to the Tau protein, which is defective and cannot stabilize microtubules properly.
- necrosis generally refers to programmed necrosis or inflammatory cell death.
- necrosis may be related to unprogrammed cell death caused by cell damage rather than orderly programmed cell death through apoptosis.
- morphological changes generally refers to changes in the morphological structure of an organism and its specific structural characteristics.
- morphological changes may include changes in external morphology (eg, shape, structure, color, pattern, size) and/or internal morphology (eg, shape and structure of internal components).
- hippocampus generally refers to the major component of the brain located in the medial temporal lobe of the brain. In Alzheimer's disease, the hippocampus may be one of the first areas of the brain to be damaged; short-term memory loss and disorientation are among the early symptoms.
- AD Alzheimer's disease
- DSM-IV-TR Diagnostic and Statistical Manual of Mental Disorders
- cogntive impairment generally refers to cognitive impairment in patients suffering from Alzheimer's disease (AD).
- cognitive impairment may include impairment of learning functions, memory functions, cognitive functions, sensory functions, motor functions, and/or emotional functions.
- apoptosis generally refers to a genetically directed process of cellular self-destruction marked by fragmentation of nuclear DNA, which can be activated by the presence of a stimulus or the removal of an inhibitor or stimulus. Apoptosis is also known as cell suicide and programmed cell death.
- Bcl-2 family proteins are one of the main intracellular regulators of apoptosis. Intracellular proteins of the Bcl-2 family help regulate proprotease activation. Some members of the Bcl-2 family promote zymogen activation and cell death.
- the apoptosis-promoting agent Bad works by binding to and inactivating the death-suppressing members of the family, while others, such as Bax and Bak, stimulate the release of cytochrome c from mitochondria. Bax and Bak themselves are also activated by other pro-apoptotic members of the Bcl-2 family such as Bid.
- Caspase-3 is a Caspase protein that interacts with Caspase-8 and Caspase-9. It is encoded by the CASP3 gene. Sequential activation of caspases plays an important role in the execution phase of apoptosis. Caspases exist as inactive zymogens.
- Caspase-3 is the main caspase involved in the cleavage of amyloid ⁇ 4A precursor protein (also known as APP) and is associated with neuronal death in Alzheimer's disease. Elevated levels of Caspase3 precursor (“Pro-caspase3”) and its cleaved form (“Cl-caspase3”) are often associated with increased apoptosis.
- necrosis generally refers to a form of cellular damage that leads to premature cell death in living tissue through autolysis. Signaling pathways are generally thought to be responsible for executing necrosis or programmed necrosis.
- the production of TNF ⁇ during viral infection leads to stimulation of its receptor TNFR1.
- the TNFR-related death protein TRADD signals to RIPK1 (receptor-interactive protein kinases 1), which recruits RIPK3 to form necrosomes.
- RIPK1 receptor-interactive protein kinases 1
- MLKL memory containing RIPK1 and RIPK3 .
- AD Alzheimer's disease
- RIPK1-regulated genes have significant overlap with multiple independent Alzheimer's disease (AD) transcriptome signatures, indicating that RIPK1 activity is a factor that can explain Alzheimer's disease (AD) transcriptome changes. important part.
- the term "synaptic degeneration” generally refers to the loss or dysfunction of synapses, which may be reflected by the status/expression of synaptic markers and/or the postsynaptic densa.
- the postsynaptic density (PSD) is a dense protein attached to the postsynaptic membrane.
- the postsynaptic density was originally identified by electron microscopy as an electron-dense region on the membrane of the postsynaptic neuron.
- the postsynaptic high-density region is closely associated with the presynaptic active zone and ensures receptor proximity to the site of presynaptic neurotransmitter release. For example, cavities or swelling in areas of high postsynaptic density may indicate synaptic degeneration.
- Synaptic status can be detected by detecting the expression or levels of synaptic vesicle protein (Synaptophysin).
- Synaptophysin is a membrane glycoprotein present in a variety of active neurons and is also found after neurons are stimulated.
- donor nucleic acid molecule generally refers to a nucleic acid molecule that provides a heterologous nucleic acid sequence to a recipient (eg, receiving nucleic acid molecule).
- upstream when used with a DNA, RNA or gene sequence generally refers to a relative position in the DNA, RNA or gene sequence towards the 5' end.
- upstream refers to the 5' end of the coding strand of the gene.
- downstream when used with a DNA, RNA or gene sequence generally refers to a relative position in the DNA, RNA or gene sequence towards the 3' end.
- downstream refers to the 3' end of the coding strand of the gene.
- induced pluripotent stem cells or "iPS cells” generally refers to cells that are extracted from any tissue of a child or adult (usually skin or blood) and that have been genetically modified to resemble embryonic phenotypes stem cell. As the name suggests, these cells are pluripotent, meaning they have the ability to form most, if not all, adult cell types.
- tracrRNA generally refers to transactivating crRNA (tracrRNA), which is a small trans-coding RNA. TracrRNA complements and base pairs with pre-crRNA to form an RNA double strand. It is specifically nucleated by RNA Glyconuclease III cleaves, forming a crRNA/tracrRNA hybrid. This hybrid acts as a guide for the endonuclease Cas9, which cuts invading nucleic acids.
- the term "early cognitive impairment (MCI)” generally refers to a clinical state intermediate between normal cognition and cognitive impairment.
- the MCI may include cognitive impairment that meets criteria for dementia but is greater than normal aging.
- MCI is diverse in clinical presentation, etiology, prognosis, and prevalence.
- MCI can be a pathological stage of Alzheimer's disease.
- Some forms of cognitive impairment may be considered early manifestations of neurodegenerative diseases that will eventually lead to dementia.
- the term "cognitive impairment due to normal aging” generally refers to cognitive impairment due to normal aging.
- the cognitive impairments caused by normal aging can manifest as: memory loss, confusion about the location of familiar places, taking longer than usual to complete daily tasks, or changes in mood and personality.
- neuron generally refers to nerve cells, which are the main functional units of the nervous system.
- a neuron can be composed of a cell body and its neurites, an axon, and one or more dendrites. Neurons can transmit messages to other neurons or cells by releasing neurotransmitters at synapses.
- the present application provides a progeny of the non-human animal of the present application, or a part thereof, obtained by crossing the non-human animal of the present application with another non-human animal, wherein the other non-human animal
- the human animal may or may not express the DIR and/or functionally active fragments thereof.
- the other non-human animal may express and/or contain other genes associated with neurological diseases or disorders, or variants thereof (eg, APP, Tau, etc.).
- the present application provides a kit.
- the kit may include: the non-human animals described in the application or parts thereof, the progeny described in the application, the cells described in the application, the tissues described in the application and/or the cell lines or original cells described in the application. generation cell culture; and one or more additional components selected from the group consisting of: assay buffers, controls, substrates, standards, test materials, laboratory supplies, equipment, instruments, cells, organs, tissues and user manual or instructions.
- the present application provides a method of screening and/or identifying substances, devices and/or compositions useful in the treatment, prevention and/or prognosis of neurological diseases or conditions.
- the method may include administering a candidate substance, device, and/or composition to a non-human animal described herein or a part thereof, a progeny described herein, a cell described herein, a cell described herein, or a non-human animal described herein. Tissues, and/or cell lines or primary cell cultures described herein; and assaying said candidate substances, devices and/or combinations The effect of a substance on one or more indications of the neurological disease or disorder.
- the present application provides a method of designing substances, devices and/or compositions for the treatment, prevention and/or prognosis of neurological diseases or disorders, the method comprising: (i) combining a candidate substance, Devices and/or compositions are administered to non-human animals described herein or parts thereof, progeny described herein, cells described herein, tissues described herein, and/or cells described herein lines or primary cell cultures; (ii) determine the effect of the candidate substances, devices and/or compositions on one or more indications of the neurological disease or disorder; (iii) select those capable of preventing, Candidate substances, devices and/or compositions that inhibit and/or alleviate one or more of the indications of the neurological disease or disorder; (iv) modify the candidate substances, devices and/or compositions of step (iii) Or the structure and/or composition of the composition to obtain modified substances, devices and/or compositions that are useful in treating, preventing and/or prognosing the neurological disease or There is improvement activity in terms of symptoms.
- the present application provides a method of screening biological targets and/or biomarkers for diagnosing and/or monitoring neurological diseases or disorders, the method comprising: respectively treating the neurological diseases or disorders.
- assays are performed on non-human animals described herein or parts thereof, progeny described herein, cells described herein, tissues described herein, and/or the presence and/or levels of each substance in samples obtained from the cell lines or primary cell cultures described herein to identify changes in the presence and/or levels before and after the occurrence of the indication. substance, and identify the substance as the biological target and/or the biomarker.
- the application provides the non-human animals described in the application or parts thereof, the progeny described in the application, the cells described in the application, the tissues described in the application, and/or the cells described in the application. lines or primary cell cultures in the preparation of identification and/or screening systems.
- the system may be used to identify and/or screen substances, devices, compositions, biological targets and/or biomarkers for the treatment, diagnosis, prevention, monitoring and/or prediction of neurological diseases or conditions.
- non-human animals described herein or parts thereof, their progeny described herein, the cells described herein, the tissues described herein, and/or the cell lines or primary cell cultures described herein can be used for Screening for substances, devices, compositions, biological targets and/or biomarkers useful in the treatment, diagnosis, prevention, monitoring and/or prediction of neurological diseases or conditions.
- the present application provides a method for preparing the non-human animal or part thereof, the cell described in the present application or the tissue described in the present application, the method comprising making the non-human animal or part thereof , cells or tissues containing and/or expressing the DIR and/or functionally active fragments thereof.
- Non-human animals their descendants or their parts
- the non-human animal may be a gene-edited non-human animal, or derived from a gene-edited non-human animal.
- the non-human animal may be a knock-in non-human animal, or derived from a knock-in non-human animal.
- the non-human animal may be a non-human mammal.
- the non-human animal is a rodent or primate.
- the non-human animal is selected from the group consisting of: mouse, rat, rabbit, ape, monkey, pig, cow, sheep, horse, chicken, dog, alpaca, and cat.
- the mouse may be of the C57/B6 strain.
- the parts of the non-human animal or its progeny may include its organs, tissues and/or cells.
- the portion includes body fluids.
- the body fluid may be selected from the group consisting of blood, plasma, serum, urine, sweat, tears, saliva, semen and cerebrospinal fluid.
- the plasma includes exosomes.
- the portion includes brain (eg, brain tissue) or a portion thereof.
- the portion of the brain may include a portion selected from the group consisting of the olfactory bulb, amygdala, basal ganglia, hippocampus, thalamus, hypothalamus, cerebral cortex, medulla oblongata, and cerebellum.
- the portion includes the olfactory mucosa.
- the portion includes a portion of the central nervous system, a portion of the peripheral nervous system, skin tissue, muscle tissue, and/or internal organs.
- the central nervous system may include the spinal cord.
- the moiety includes cells.
- the cells may include cells selected from the group consisting of primary neurons, microglia, astrocytes, oligodendrocytes, macrophages, perivascular epithelioid cells, B cells, T cells, Adult stem cells, NK cells, totipotent stem cells, unipotent stem cells, embryonic stem cells, induced pluripotent stem cells and gametes.
- the gametes may include sperm and/or oocytes.
- the part eg, the organ, tissue, or cell
- the part is unable to develop into a complete non-human animal.
- the non-human animal, its progeny, or parts thereof comprise the DIR and/or functionally active fragment thereof.
- the DIR and/or functionally active fragment thereof may be expressed by the animal, the progeny or the cell, or may be derived from other sources and then introduced (e.g., injected) into the animal, progeny or cell, thereby causing It contains the DIR and/or functionally active fragments thereof.
- the non-human animal, its progeny, or parts thereof express the DIR and/or functionally active fragment thereof.
- the expressed DIR and/or functionally active fragments thereof may be secreted or otherwise excreted outside the cell, tissue or animal body.
- the expression of DIR may include expression of a DIR gene, expression of a transcript of a DIR gene, and/or expression of a DIR protein.
- the expression may include expression of a specific gene (eg, human DIR gene) polynucleotide, mRNA, or amino acid product or protein.
- the expression may include expression of a fragment of a transcribed polynucleotide, a translated protein, or a post-translationally modified protein of a particular gene (eg, a human DIR gene).
- the expression of the functionally active fragment of DIR may include expression of a gene encoding a functionally active fragment of DIR, a transcript of a gene encoding a functionally active fragment of DIR, and/or Or the expression of functionally active fragment proteins of DIR.
- the expression may include expression of a polynucleotide, mRNA or amino acid product or protein of a particular gene, such as a gene encoding a functionally active fragment of human DIR (e.g., DIR-IR, DIR-I, and/or DIR-II).
- the expression may include a transcribed polynucleotide, a translated protein, or a post-translationally modified protein of a specific gene, such as a gene encoding a functionally active fragment of human DIR (e.g., DIR-IR, DIR-I, and/or DIR-II). expression of fragments.
- DIR-IR e.g., DIR-IR, DIR-I, and/or DIR-II
- the amino acid sequence of DIR-IR can be shown as SEQ ID NO:2.
- the amino acid sequence of DIR-I can be shown as SEQ ID NO: 4.
- the amino acid sequence of DIR-II can be as shown in SEQ ID NO: 5.
- the expression or content of the DIR can be identified by using a substance selected from the following group: primers that specifically amplify the DIR gene, nucleic acid molecules that specifically bind to the DIR gene, and DIR proteins that specifically bind. Nucleic acid molecules, small molecules that specifically bind to DIR proteins, probes that specifically bind to DIR proteins, and polypeptides that specifically bind to DIR proteins.
- the content or expression of functionally active fragments of said DIR can be identified by utilizing a substance selected from the group consisting of: specifically amplifying DIR Primers of functionally active fragment genes, nucleic acid molecules that specifically bind to functionally active fragment genes of DIR, nucleic acid molecules that specifically bind to functionally active fragments of DIR (such as DIR-IR, DIR-I and/or DIR-II), Small molecules that specifically bind to functionally active fragments of DIR (such as DIR-IR, DIR-I and/or DIR-II), functionally active fragments of DIR (such as DIR-IR, DIR-I and/or DIR-II) ) probes that specifically bind and polypeptides that specifically bind to functionally active fragments of DIR (e.g., DIR-IR, DIR-I and/or DIR-II).
- the expression and/or content of said DIR and/or functionally active fragments thereof can be determined by performing an assay selected from the group consisting of reverse transcription and amplification analysis (e.g. PCR, ligation RT-PCR or quantitative RT -PCT), hybridization analysis, Northern blotting, dot blotting, in situ hybridization, gel electrophoresis, capillary electrophoresis, column chromatography, Western blotting, immunohistochemistry, immunostaining or mass spectrometry.
- qPCR, qRT-PCR, northern Hybridization, western hybridization, and/or ELISA assays measure the expression or content of DIRs described herein.
- the expression and/or content of the DIR can also be measured by performing analysis directly on a biological sample or on proteins/nucleic acids isolated from the sample.
- non-human animals can be made to express and/or contain the DIR and/or functionally active fragments thereof by subjecting them to specific conditions or environments (eg, stress or hypoxic environment, etc.).
- specific conditions or environments eg, stress or hypoxic environment, etc.
- the DIR may be derived from a mammal, such as a primate, eg, a human.
- the DIR or functionally active fragment thereof may comprise the amino acid sequence encoded by the retained intron in DDIT4L.
- the DIR or functionally active fragment thereof comprises the amino acid sequence shown in any one of SEQ ID NOs: 1-2 and 4-5, or a functional variant or truncation thereof.
- the functional variant may comprise 1 or more (eg, 1 or several, eg, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or more) amino acids substitution, deletion or addition.
- the genes encoding the DIR and/or functionally active fragments thereof are homozygous. In certain embodiments, the genes encoding the DIR and/or functionally active fragments thereof are hybrid. In certain embodiments, the expression of the DIR and/or functionally active fragments thereof is regulated by corresponding endogenous regulatory elements in the genome of the non-human animal or cell. For example, expression of the DIR and/or functionally active fragments thereof may be regulated by regulatory elements (eg, promoter and/or enhancer elements) of the DDIT4L-encoding gene in the genome of the non-human animal or cell.
- regulatory elements eg, promoter and/or enhancer elements
- identity shared by polynucleotide or polypeptide sequences Percentages are determined by direct comparison of sequence information between molecules by aligning the sequences and determining identity using methods known in the art.
- An example of an algorithm suitable for determining sequence similarity is the BLAST algorithm (see Altschul et al., J. Mol. Biol. 215:403-410 [1990]). Used to enter Software for performing BLAST analyzes is publicly available through the National Center for Biotechnology Information (NCBI).
- the biological activity may include the ability to reduce the frequency of excitatory postsynaptic currents (EPSCs), and/or the ability to reduce the amplitude of EPSCs.
- said reducing may comprise administering said DIR and/or functionally active fragments thereof and/or encoding said DIR and/or functionally active fragments thereof compared to the original biological activity of said DIR and/or functionally active fragments thereof in said subject.
- Nucleic acids of DIR and/or functionally active fragments thereof can reduce the frequency of excitatory postsynaptic currents (EPSCs) in a subject, and/or reduce the amplitude of EPSCs in a subject.
- it can be reduced by at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least About 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 100%, at least about 200%, at least About 500% or more.
- the cells may be non-human animal cells.
- the tissue may be a tissue of a non-human animal.
- the cells or tissues are unable to develop into a complete non-human animal.
- the cell may not be a totipotent stem cell or an embryonic stem cell.
- the cells may be knock-in cells, or derived from knock-in cells.
- the expression of the DIR and/or functionally active fragments thereof can be regulated by corresponding endogenous regulatory elements in the genome of the cell.
- the tissue may be a gene-edited tissue (e.g., at least a portion of the cells within the tissue may be gene-edited cells), or derived from a gene-edited tissue (e.g., derived from a gene-edited cell). tissue differentiation or culture).
- compositions of the present application may include the non-human animals described in the present application or parts thereof, the progeny described in the present application, the cells described in the present application, the tissues described in the present application and/or the cell lines described in the present application, or Primary cell cultures.
- the composition may also include one or more other adjuvants or excipients, such as pharmaceutically acceptable buffers, other reagents, excipients, etc.
- the user manual or instructions may record the screening, detection or identification methods described in this application.
- the buffer can be used to analyze the performance of the non-human animal, parts thereof, the tissue, cell, cell line or cell culture, and the like.
- the control substance can be used as a control substance for non-human animals, tissues, cells, cell lines, cell cultures, etc. of the present application to detect or evaluate the non-human animals, tissues, cells, cell lines, cell cultures, etc. nature or activity.
- the substrates, standards, detection materials, laboratory supplies, equipment, instruments, etc. can also be used or coordinated with the non-human animals, parts thereof, tissues, cells, cell lines or cell cultures of the present application. Used for analysis, screening or determination.
- the present application also provides methods for preparing the non-human animals or parts thereof, the cells described in the present application, or the tissues described in the present application, the methods comprising making the non-human animals or parts thereof, cells or tissues comprise and/or express said DIR and/or functionally active fragments thereof.
- the method may comprise introducing into the non-human animal or part thereof, the cell, tissue, cell line and/or cell culture the DIR, a functionally active fragment thereof, a nucleic acid molecule encoding the DIR, and /or a nucleic acid molecule encoding a functionally active fragment of DIR.
- the introduction may include, for example, injection (eg, injection of a protein or nucleic acid molecule) into the non-human animal, a part thereof, the tissue or cells.
- the introduction may also include nucleic acid molecule transfection, viral transduction and other means of introducing exogenous proteins, exogenous nucleic acid molecules, etc. into cells, tissues or animals.
- the method may comprise knocking in the DIR and/or functionally active fragment thereof in the non-human animal or part thereof, the cell, tissue, cell line and/or cell culture. nucleic acid sequence.
- nucleases can be used to assist in the modification of target gene sites. Such nucleases can promote homologous recombination between donor nucleic acid molecules and target gene sites.
- the nuclease includes a nuclease Endonuclease.
- the recognition site can be about 9 to 12 nucleotides in length, about 12 to 15 nucleotides in length, about 15 to 18 nucleotides in length, or about 18 to 18 nucleotides in length. 21 nucleotides, and any combination of these subranges (e.g., 9-18 nucleotides).
- the recognition site can be a palindrome, that is, the sequence on one strand is identical to the sequence in the opposite direction on the complementary strand.
- a specific nuclease can bind to the recognition site and cleave the binding site, or the nuclease can bind to a different sequence than the recognition site.
- the term "recognition site” may encompass both a nuclease binding site and a nick/cleavage site, whether the nick/cleavage site is within or outside the nuclease binding site.
- nuclease cleavage can occur at nucleotide positions opposite each other to create blunt-end cleavages, or in other cases, the cuts can be staggered to create single-stranded overhangs, also known as "sticky ends.” It can be a 5' overhang or a 3' overhang.
- nuclease that produces a nick or double-stranded break at the desired recognition site can be used in the methods of the present application.
- Naturally occurring or native nucleases can be used as long as the nuclease induces a nick or double-strand break at the desired recognition site.
- modified or engineered nucleases may be used, "engineered nucleases” including nucleases that are engineered (modified or derived) from their native form to specifically recognize and generate nicks or double-stranded breaks at the desired recognition site .
- engineered nucleases may be derived from natural or naturally occurring nucleases, or may be artificially produced or synthesized.
- the nuclease may be a transcription activator-like effector nuclease (Transcription Activator-Like Effector Nuclease, TALEN).
- TAL effector nucleases are a class of sequence-specific nucleases that can be used to generate double-strand breaks at specific target sequences in prokaryotic or eukaryotic genomes.
- TAL effector nucleases can be generated by fusing natural or engineered transcription activator-like (TAL) effectors, or functional portions thereof, into the catalytic domain of an endonuclease (eg, FokI).
- TAL effector nucleases can be generated by fusing natural or engineered transcription activator-like (TAL) effectors, or functional portions thereof, into the catalytic domain of an endonuclease (eg, FokI).
- TAL effector DNA-binding domains enable the design of proteins with any specific DNA recognition specificity.
- the DNA-binding domains of TAL effector nucleases can be engineered to recognize specific DNA target sites and thus be used to make double-strand breaks at desired target sequences.
- WO 2010/079430 see WO 2010/079430; Morbitzer et al., (2010) PNAS 10.1073/pnas.1013133107; Scholze & Boch (2010) Virulence 1: 428-432; Christian et al., (2010) Genetics 186: 757-761; Li et al. , (2010) Nuc. Acids Res., doi: 10.1093/nar/gkq704; Miller et al., (2011) Nature Biotechnology 29: 143-148; the above are hereby incorporated by reference.
- the nuclease can be a zinc finger nuclease (Zinc-finger nuclease, ZFN).
- ZFN zinc finger nuclease
- each monomer of ZFN can contain 3 or more zinc finger-based DNA binding domains, where Each zinc finger-based DNA-binding domain can bind to a 3 bp subsite.
- the ZFN may be a chimeric protein comprising a zinc finger-based DNA binding domain operably linked to an independent nuclease.
- the independent endonuclease can be FokI endonuclease.
- the nuclease can comprise a first ZFN and a second ZFN, wherein each of the first ZFN and the second ZFN is operably linked to a FokI nuclease, wherein the first ZFN and the second ZFN recognize Two consecutive target DNA sequences on each single strand of the target DNA sequence, the sequences being separated by a cleavage site of about 6 bp to about 40 bp or a cleavage site of about 5 bp to about 6 bp, and wherein FokI nuclease dimerizes and produce double-strand breaks.
- the nuclease may be a meganuclease.
- Giant nucleases have been classified into four families based on conserved sequence motifs. These different structural formulas (motifs) are involved in the coordination of metal ions and the hydrolysis of phosphodiester bonds.
- HEase is known for its long recognition site and tolerance to certain sequence polymorphisms in its DNA substrate.
- Giant nuclease domains, structure and function are known, see for example: Guhan and Muniyappa (2003) Rev Biochem Mol Biol 38: 199-248; Lucas et al., (2001) Nucleic Acids Res 29: 960-9; Jurica and Stoddard , (1999) Cell Mol Life Sci 55: 1304-26; Stoddard, (2006) Q Rev Biophys 38: 49-95; and Moure et al., (2002) Nat Struct Biol 9: 764.
- the sgRNA expression component and the Cas9 expression component can then be introduced into the cell. See: for example, Mali P et al., (2013) Science, 339(6121): 823-6; Jinek M et al., (2012) Science, 337(6096): 816-21; Hwang WY et al., (2013) Nat Biotechnol, 31 (3): 227-9; Jiang W et al., (2013) Nat Biotechnol, 31(3): 233-9, and Cong L et al., (2013) Science, 339(6121): 819-23. The foregoing are incorporated herein by reference.
- the knock-in may include: contacting the genome of the non-human animal, cell or tissue with the following substances in the presence of a donor nucleic acid molecule comprising the nucleic acid sequence: 1) CRISPR-associated (Cas) proteins; and 2) one or more ribonucleic acid (RNA) sequences comprising: i) a portion complementary to a target site and ii) a binding site for the Cas protein.
- a donor nucleic acid molecule comprising the nucleic acid sequence: 1) CRISPR-associated (Cas) proteins; and 2) one or more ribonucleic acid (RNA) sequences comprising: i) a portion complementary to a target site and ii) a binding site for the Cas protein.
- the binding site of the Cas protein may comprise a tracrRNA sequence.
- the RNA sequence comprises the nucleic acid sequence set forth in SEQ ID NO: 6.
- the one or more RNA sequences may be provided in the form of one or more RNA molecules or one or more DNA molecules encoding the RNA sequences.
- the Cas protein can be a Type I Cas protein; in certain embodiments, the Cas protein can be a Type II Cas protein; in certain embodiments, the Cas protein can be Cas9.
- the method may further comprise identifying a modified cell or non-human animal containing the knock-in heterologous nucleic acid sequence.
- Donor nucleic acid molecules containing heterologous nucleic acid sequences may also contain 5' homology arms and 3' homology arms.
- 5' homology arms and 3' homology arms may flank heterologous nucleic acid sequences encoding said DIR and/or functionally active fragments thereof.
- the homology arms (e.g., 5' homology arms or 3' homology arms) in the donor nucleic acid molecule can have sufficient strength to facilitate homologous recombination with the corresponding region in the endogenous target site (e.g., endogenous DDIT4L).
- the donor nucleic acid molecule includes a 5' homology arm that can be about 800-900 bp in length and a 3' homology arm that can be about 800-950 bp in length.
- the degree of sequence identity shared by the homology arms of the donor nucleic acid molecule (or fragment thereof) and the target site (or fragment thereof) can be at least about 50%, at least about 55%, at least about 60%, at least about 65% %, at least about 70%, at least about 75%, at least about 80%, at least about 81%, at least about 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%%, at least about 87 %, at least about 88%, at least about 89%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%%, at least about 97%, at least about 98%, at least about 99%, or at least about 100% for homologous recombination of the sequence.
- the DNA encoding the Cas protein may be in the form of a first expression vector comprising a first promoter operably linked to the nucleic acid encoding the Cas protein.
- the DNA encoding an RNA portion complementary to the target gene can be in the form of a second expression vector comprising a second promoter operably linked to the nucleic acid encoding the RNA portion.
- the first, second and third promoters may be active in the non-human animal or cell.
- the identification, screening or design method of the present application may be an in vitro method, an ex vivo method or an in vivo method.
- Candidate substances, devices and/or compositions are selected for further investigation (e.g., as potential therapeutic agents/devices/compositions for treating AD or inhibiting A ⁇ accumulation).
- screening methods can also be performed using non-human animal tissues and/or cells according to the present application, such as neuronal cells, brain regions, or other tissues containing/corresponding to AD lesions, or part of tissues thereof (for example, the olfactory bulb, amygdala, basal ganglia, hippocampus, thalamus, hypothalamus, cerebral cortex, medulla oblongata, central nervous system, peripheral nervous system, spinal cord, cerebellum, skin tissue, muscle tissue, and/or internal organs).
- non-human animal tissues and/or cells such as neuronal cells, brain regions, or other tissues containing/corresponding to AD lesions, or part of tissues thereof (for example, the olfactory bulb, amygdala, basal ganglia, hippocampus, thalamus, hypothalamus, cerebral cortex, medulla oblongata, central nervous system, peripheral nervous system, spinal cord, cerebellum, skin tissue, muscle tissue, and/or internal organs).
- tissues and/or cells may be cultured in vitro or ex vivo, and then, after incubation for an appropriate period of time, candidate substances, devices, and/or compositions may be administered to the cultured tissues and/or cells (e.g., several hours, days, weeks, or months) and examine the treated brain tissue/cells using the methods described above.
- non-human animals derived from the present application, parts thereof, or their progeny may be harvested upon the development of signs associated with neurological diseases or disorders (e.g., A ⁇ deposition, neurological Fibrillary tangles, morphological or functional abnormalities (atrophic) synapses, neuronal cell death, or impairment of memory and learning functions) before and after samples (e.g., cells, tissues or other samples containing DNA or RNA, proteins containing samples, and/or metabolite-containing samples), followed by a comprehensive analysis of gene transcripts (transcriptome), gene translation products (proteome), lipids (lipidome) or metabolites (metabolome) from the sample , and identify substances that changed before and after the relevant indications appeared.
- the sample may comprise a body fluid of the non-human animal, such as rat blood, plasma (e.g., Exosome-containing plasma), serum, urine, sweat, tears, saliva, semen, and/or cerebros
- the substance can be considered a biomarker for a neurological disease or condition, which can then be used for early diagnosis (especially for Preclinical diagnosis of neurological diseases or conditions).
- the identified biomarkers can be further tested using specific reagents or detection methods.
- the biomarker is a protein or peptide, it can be detected by immunoassay using specific antibodies.
- the biomarker is a nucleic acid molecule (such as a transcript), it can be detected by Northern blot analysis using specific probes or by RT-PCR using specific primers.
- Co-localization of necrosomes and microglia can be detected in non-human animals, parts (e.g., tissues, organs, or other parts), tissues, cells, or cell cultures of the present application. as revealed by colocalization of expression.
- neuronal cell loss can be detected in non-human animals, parts thereof (eg, tissues, organs, or other parts), tissues, cells, or cell cultures of the present application.
- a decrease in the number of Neu-N positive neurons may be detected in the non-human animal, part thereof, tissue, cell or cell culture.
- gliosis can be detected in the non-human animal, part thereof, cell, tissue or cell culture according to the present application.
- Gliosis can include microgliosis and/or astrogliosis.
- microgliosis and/or astrogliosis may be associated with amyloid plaques. For example, astrocytes and microglia gather around amyloid plaques, and their numbers may also increase.
- synaptic degeneration is detectable in the non-human animal, part thereof, cell, tissue or cell culture according to the present application.
- synaptic degeneration may be revealed by swelling and/or cavitation of the postsynaptic dense zone and/or the status/expression of synaptic markers. Swelling and/or cavities in this postsynaptic density and/or presynaptic density can be detected with anti-PSD95 staining. The status/expression of this synaptic marker can be detected using anti-synaptophysin staining.
- changes in brain morphology and/or weight are detectable in the non-human animal or part thereof compared to the corresponding wild-type non-human animal.
- changes in brain morphology and/or weight may include a reduction in brain size, the development of ventricular cavities and/or ventricular enlargement, and/or damage to the hippocampus.
- a reduction in brain size may be detected in the non-human animal or part thereof.
- cognitive impairment is detectable in the non-human animal compared to a corresponding wild-type non-human animal.
- cognitive impairment can be detected by open field testing, Morris maze testing, and/or T-maze working memory testing.
- cognitive functions may include learning functions, memory functions, cognitive functions, sensory functions, motor functions, and/or emotional functions.
- Impairment of learning function can be detected in the open field test, Morris maze test and/or T-maze working memory test.
- impairment of memory function can be detected in the open field test, Morris maze test and/or T-maze working memory test.
- Cognitive impairment can be detected in the open field test, Morris maze test, and/or T-maze working memory test.
- impairment of sensory function can be detected in the open field test, Morris maze test, and/or T-maze working memory test.
- the neurological disease or disorder may include cognitive impairment.
- the neurological diseases or conditions described herein may include neurodegenerative diseases or conditions.
- the neurological diseases or conditions described herein may include mild cognitive impairment (MCI) and/or Alzheimer's disease.
- MCI mild cognitive impairment
- the neurodegenerative diseases may include acute neurodegenerative diseases and chronic neurodegenerative diseases.
- the neurodegenerative diseases may include neurodegenerative diseases caused by neuronal death and glial cell homeostasis, neurodegenerative diseases caused by aging, neurodegenerative diseases caused by CNS cell function being affected, neurodegenerative diseases caused by Neurodegenerative diseases caused by abnormal communication between cells and/or neurodegenerative diseases caused by impaired cell movement.
- the cognitive impairment may include early cognitive impairment (MCI), intermediate cognitive impairment and late cognitive impairment.
- MCI early cognitive impairment
- the cognitive impairment may include cognitive impairment caused by normal aging and the like.
- symptoms of early cognitive impairment (MCI) may include: loss of short-term memory, difficulty expressing or understanding abstract things, erratic mood or behavior, difficulty learning new things and following complex instructions, impaired judgment and/or basic Self-care requires reminders from others.
- symptoms of mid-term cognitive impairment may include: confusion between long-term memory and memory of real-life situations, inability to communicate clearly, changes in behavior or personality, or emotional instability, and/or the need for assistance with self-care; for example, symptoms of late-stage cognitive impairment may include: Memory impairment, decline in physical activity and mental status, inability to express or communicate effectively, inability to take care of oneself, and/or disruption of the biological clock, etc.
- Cas9 mRNA and sgRNA for knock-in (SEQ ID NO: 6) through in vitro transcription; then, construct a homologous recombination vector as a donor, which contains about 3 kb of 5' homology arm (SEQ ID NO:16), a nucleic acid sequence encoding the last 64 amino acids of human DIR (SEQ ID NO:18), and a 3' homology arm of approximately 3 kb (SEQ ID NO:17).
- the mRNA encoding Cas9, sgRNA and donor vector were microinjected into the fertilized eggs of C57BL/6J mice to obtain F0 generation mice.
- the F0 generation mice identified as positive by PCR were mated with C57BL/6J mice to obtain positive F1 generation mice.
- F2 generation wild-type, heterozygous and homozygous mice can be obtained by mating F1 generation mice.
- the body weight of homozygous (HO) DIR-KI mice was not significantly different from that of wild-type (WT) mice, indicating that the growth and metabolism of the mutant mice were normal (Fig. 6A).
- WT wild-type mice
- Fig. 6B there was no difference in walking distance and speed between WT and homozygous (HO) DIR-KI mice in the open field test, indicating that the locomotor function of the mutant mice was not altered
- Genome identification Collect the tails of the DIR-KI mice in Example 1, use TIANamp Genomic DNA Kit (TIANGEN) to extract the mouse genomic DNA, and design identification primers P1 (SEQ ID NO: 7), P2 (SEQ ID NO: 8), and identify the mouse genotype through PCR reaction combined with gel electrophoresis.
- a 476bp PCR product can be obtained from wild-type (WT) mice
- two PCR products of 476bp and 671bp can be obtained from DIR heterozygous (HE) mice
- a 671bp PCR product can be obtained from DIR homozygous (HO) mice.
- Protein detection Take the brain tissue of DIR-KI mice and wild-type mice and lyse it with lysis solution (50mM Tris, 150mM NaCl, 1% Triton X-100, 1% sodium deoxycholate, 0.1% SDS, pH 7.4) Then, centrifuge the supernatant, mix it with an appropriate amount of 4X loading buffer, and denature it at 100°C for 10 minutes. Finally, an appropriate amount of samples were taken for a Western blot experiment, and the results showed that DIR protein could be detected in the brain tissue of DIR-KI mice (Figure 2, C).
- a Leica SP8 two-photon confocal microscope was used to image the labeled tissue slices, and it was found that DIR was highly enriched in neurons.
- DG hippocampal dentate gyrus
- the activity of neurons in DIR-KI mice was detected by electrophysiological recording, and immunohistological staining was used to determine whether the increase in p-Tau and the accumulation of A ⁇ occurred.
- Immunological tissue staining Adult DIR-KI mice were anesthetized, perfused with PBS, and fixed with 4% paraformaldehyde solution. Then the brain tissue of the mice was removed and cut into approximately 100 pieces under a Leica freezing microtome. 50 ⁇ m thick tissue sections. Take out an appropriate amount of sections and attach them to a slide that has been previously coated with gelatin. After blocking with donkey serum, add p-Tau (Abcam, Cat. No. ab32057) and A ⁇ (Agrisera, Cat. No. AS0932) antibodies for labeling, and add Fluorescent secondary antibodies were labeled with p-Tau and A ⁇ antibodies.
- p-Tau Abcam, Cat. No. ab32057
- a ⁇ Agrisera, Cat. No. AS0932
- mice Before conducting the behavioral experiment, the mice were allowed to adapt to the breeding environment for one week. On the day of the experiment, allow the mice to adapt to the experimental room 30 minutes in advance. After the experiment started, the mice were placed in the starting arm A of the Y-shaped maze facing the wall and allowed to freely explore the maze for 10 minutes. When located in the center of the maze, mice can choose any direction to enter. Cognitively normal mice tended to enter unexplored arms during the experiment. The entire process was recorded using camera video. After the experiment, the mice were returned to their cages. Use 75% alcohol to clean the experimental device between experiments.
- New object recognition experiment Allow the mice to adapt to the experimental room 30 minutes in advance on the day of the experiment.
- the experiment is divided into adaptation period, training period and experimental period.
- Adaptation period Take the mice out of the cage, place them in the center of the plastic box, and freely explore the experimental site for 10 minutes.
- the training period is carried out 12 to 24 hours after the adaptation period; training period: place 2 identical batteries 5 cm away from the wall. Place the mouse in the center of the arena and explore freely for 10 minutes.
- the experimental period will be conducted within 3 to 4 hours after the end of the training period; during the experimental period: a toy is randomly replaced with a battery and placed in the same position as the original object. The mice were placed in the center of the arena to explore freely for 10 minutes.
- mice were returned to their cages. The entire process was recorded using camera video. Use 75% alcohol to clean the experimental device between experiments.
- the time spent by mice exploring old and new objects in the video was manually counted and the time recognition index of new objects was calculated.
- the tip of the mouse's nose is within 2 cm of an object, and the mouse sniffs or touches the object, it is considered an exploratory behavior. Biting or climbing on the object is not considered an exploratory behavior.
- the mouse's recognition index of new objects (time to explore new objects - time to explore old objects)/(time to explore new objects + time to explore old objects).
- the total exploration time of the mouse is less than 15 seconds Data will be excluded.
- the experimental operators and data analysts were double-blind. The results showed that the ability of DIR-KI mice to recognize new objects was weaker than that of wild-type mice ( Figure 4, B).
- Water maze test Allow the mice to adapt to the experimental room 30 minutes in advance on the day of the experiment.
- the experiment is divided into training day and experimental day.
- the training day is 5 to 6 days, and the experimental day is 1 day.
- fill the water tank with water to a depth of 30 cm, maintain the water temperature at 19-22°C, and add talcum powder to the water to make it turbid.
- the water tank is divided into four directions: N, S, E and W.
- the platform is placed in the NE quadrant.
- the mice's daily water entry points are selected from non-repeating combinations of S, W, NW and SE.
- the room setup remained unchanged during the experiment, with specific patterns posted around the water maze as distal cues.
- the room light is indirect light.
- mice Train 4 times a day for a total of 5 to 6 days.
- Experimental day The experiment will be conducted 24 hours after the end of the training day. Remove platform. Mice entered the water from the SW quadrant, facing the tank wall. After 1 minute, the mice were taken out, dried, and returned to the cage. After the experiment, the mouse behavioral video was analyzed using Etho Vision The experimental operators and data analysts were double-blind. The results showed that the number of times DIR-KI mice crossed the quadrant of the platform was significantly reduced, and the time to enter the quadrant of the platform was significantly increased, indicating that the spatial memory ability of DIR-KI mice was weakened (Figure 4, C).
- Brain slice electrophysiological recording The mouse brain tissue was placed in pre-cooled artificial cerebrospinal fluid (ACSF) and continuously filled with 95% O 2 and 5% CO 2 , and then cut into approximately 350 ⁇ m thick using a Leica vibrating microtome. of tissue pieces. After selecting suitable brain slices, they are placed in an observation chamber filled with artificial cerebrospinal fluid. The observation chamber is fixed under the field of view of the microscope, and fresh artificial cerebrospinal fluid is continuously perfused to maintain the activity of the brain tissue. Finally, electrodes were used to record postsynaptic currents from the hippocampus.
- ASF artificial cerebrospinal fluid
- Example 6 The growth, metabolism and motor functions of the disease model are not changed
- mice By weighing the body weight of mice, we found no significant difference between wild-type mice (WT) and homozygous mice (HO), indicating that DIR knock-in did not affect the growth and metabolism of mice (Figure 6, A).
- Open field experiment Allow the mice to adapt to the experimental room 30 minutes in advance on the day of the experiment. During the experiment, the mice were taken out of the breeding cage, placed in the center of the plastic box, and allowed to freely explore the experimental site for 10 minutes. After the experiment, the mice were returned to their cages. The entire process was recorded using camera video. Use 75% alcohol to clean the experimental device between experiments. After the experiment, use Etho Vision XT 14 software to analyze the experimental video and count the time the mice stayed in the central area and peripheral area. The results showed no obvious difference between wild-type mice (WT) and homozygous mice (HO), indicating that the locomotor ability of DIR knock-in mice was not changed.
- WT wild-type mice
- HO homozygous mice
- hypoxic treatment Mice were placed in laboratory mouse cages, and hypoxic conditions (8% oxygen) were continued to be provided through the air inlet for 8 hours.
- An oxygen analyzer (Shanghai Precision Instrument Co., Ltd.) was used to detect the air outlet. Ensure hypoxic conditions in the cage.
- Figure 7 shows DIR expression in homozygous (HO) DIR-KI mice under hypoxic conditions.
- DIR mAb that recognizes the last 27 AA of DIR (PPLPLCRRCHKIHLRRLLSKFSNIFSP), such as A2D10 antibody, which has HCDR of the amino acid sequence shown in SEQ ID NO:19, HCDR2 of the amino acid sequence shown in SEQ ID NO:20, SEQ ID HCDR3 with the amino acid sequence shown in NO:21, the heavy chain variable region of the amino acid shown in SEQ ID NO:22; it has LCDR1 with the amino acid sequence shown in SEQ ID NO:24, the amino acid sequence shown in SEQ ID NO:25: YTS LCDR2, LCDR3 with the amino acid sequence shown in SEQ ID NO:26, and light chain variable region with the amino acid sequence shown in SEQ ID NO:27.
- DIR PPLPLCRRCHKIHLRRLLSKFSNIFSP
- A2D10 antibody which has HCDR of the amino acid sequence shown in SEQ ID NO:19, HCDR2 of the amino acid sequence shown in SEQ ID NO:20, SEQ ID
- mice Homozygous (HO) DIR-KI mice were injected intraperitoneally with DIR mAb (0.5 mg/day) or IgG for 5 days.
- DIR mAb 0.5 mg/day
- IgG IgG
- mAb treatment reduced escape latency, and both working memory and novel object recognition were improved ( Figures 8A, 8B).
- mice treated with mAb entered shorter latencies and spent longer in the target quadrant (Fig. 8C).
- mAb treatment reduced the expression of A ⁇ and p-Tau ( Figures 8D, 8E) but did not significantly change the expression of GFAP and IBA1 in the hippocampus of syngeneic DIR-KI mice.
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Abstract
L'invention concerne un modèle de maladie et son utilisation. L'invention concerne spécifiquement un modèle d'une maladie ou d'un trouble du système nerveux, le modèle comprenant et/ou exprimant un produit de clivage-retenue d'intron de transcrit de type 4 de transcription inductible par des dommages à l'ADN ou un fragment de celui-ci.
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Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20160237455A1 (en) * | 2013-09-27 | 2016-08-18 | Editas Medicine, Inc. | Crispr-related methods and compositions |
CN111936628A (zh) * | 2018-03-20 | 2020-11-13 | 清华大学 | 阿尔茨海默病动物模型及其应用 |
CN112168969A (zh) * | 2020-09-29 | 2021-01-05 | 上海联慧脑智工程中心 | Ddit4l及其功能小肽对胶质母细胞瘤的抑制作用 |
KR20220082545A (ko) * | 2020-12-10 | 2022-06-17 | 중앙대학교 산학협력단 | 전사체 분석을 이용한 인트론 유지 검출을 통해 퇴행성 뇌 질환을 진단하는 방법 |
WO2023143425A1 (fr) * | 2022-01-25 | 2023-08-03 | 上海魁特迪生物科技有限公司 | Méthode d'amélioration de troubles cognitifs |
-
2023
- 2023-06-20 WO PCT/CN2023/101488 patent/WO2023246806A1/fr unknown
Patent Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20160237455A1 (en) * | 2013-09-27 | 2016-08-18 | Editas Medicine, Inc. | Crispr-related methods and compositions |
CN111936628A (zh) * | 2018-03-20 | 2020-11-13 | 清华大学 | 阿尔茨海默病动物模型及其应用 |
CN112168969A (zh) * | 2020-09-29 | 2021-01-05 | 上海联慧脑智工程中心 | Ddit4l及其功能小肽对胶质母细胞瘤的抑制作用 |
KR20220082545A (ko) * | 2020-12-10 | 2022-06-17 | 중앙대학교 산학협력단 | 전사체 분석을 이용한 인트론 유지 검출을 통해 퇴행성 뇌 질환을 진단하는 방법 |
WO2023143425A1 (fr) * | 2022-01-25 | 2023-08-03 | 上海魁特迪生物科技有限公司 | Méthode d'amélioration de troubles cognitifs |
Non-Patent Citations (2)
Title |
---|
LI HONG‐DONG, FUNK CORY C., MCFARLAND KAREN, DAMMER ERIC B., ALLEN MARIET, CARRASQUILLO MINERVA M., LEVITES YONA, CHAKRABARTY PARA: "Integrative functional genomic analysis of intron retention in human and mouse brain with Alzheimer's disease", ALZHEIMER'S & DEMENTIA, ELSEVIER, NEW YORK, NY, US, vol. 17, no. 6, 1 June 2021 (2021-06-01), US , pages 984 - 1004, XP093080999, ISSN: 1552-5260, DOI: 10.1002/alz.12254 * |
ZHENG JIAN-TAO, LIN CUI-XIANG, FANG ZHAO-YU, LI HONG-DONG: "Intron Retention as a Mode for RNA-Seq Data Analysis", FRONTIERS IN GENETICS, FRONTIERS RESEARCH FOUNDATION, SWITZERLAND, vol. 11, Switzerland , XP093121113, ISSN: 1664-8021, DOI: 10.3389/fgene.2020.00586 * |
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