WO2023206347A1 - 作为早老性痴呆诊断标志物的ddit4l剪切产物 - Google Patents

作为早老性痴呆诊断标志物的ddit4l剪切产物 Download PDF

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WO2023206347A1
WO2023206347A1 PCT/CN2022/090220 CN2022090220W WO2023206347A1 WO 2023206347 A1 WO2023206347 A1 WO 2023206347A1 CN 2022090220 W CN2022090220 W CN 2022090220W WO 2023206347 A1 WO2023206347 A1 WO 2023206347A1
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disease
qdlir
cognitive impairment
nucleic acid
alzheimer
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PCT/CN2022/090220
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English (en)
French (fr)
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李开诚
攸璞
张旭
毛颖
陈亮
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苏州瀛创生物科技有限公司
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Priority to PCT/CN2022/090220 priority Critical patent/WO2023206347A1/zh
Priority to PCT/CN2023/091199 priority patent/WO2023208119A1/zh
Publication of WO2023206347A1 publication Critical patent/WO2023206347A1/zh

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids

Definitions

  • This application relates to the field of biomedicine, specifically to the application of QDLIR, the intronic retention cleavage product of DNA damage-induced transcript 4-like transcript (DDIT4L), as a blood marker for Alzheimer's disease or mild cognitive impairment and its application Corresponding products and methods.
  • QDLIR the intronic retention cleavage product of DNA damage-induced transcript 4-like transcript
  • AD Alzheimer's disease
  • AD Alzheimer's Disease International
  • 2017 World Alzheimer's Disease Report 2017
  • the rate is 65.6%.
  • AD patients there are 5 million AD patients in the United States, and caregivers spend 18 billion hours a year caring for patients.
  • drugs that can treat, delay or prevent the progression of the disease the number of patients is expected to double every 20 years, reaching 82 million in 2030 and 150 million in 2050.
  • AD Alzheimer's disease
  • AD cognitive impairment
  • cognitive function impairment is caused by synaptic and neuronal damage, and abnormal neural circuit structure and function.
  • the main reasons for the continuous failure of clinical research on new drugs for AD are that the mechanism of synaptic and neuronal damage is still not clear enough, and there is a lack of cell/animal models that are close to the pathophysiological characteristics of human diseases (such as neurodegeneration, etc.), which leads to the effectiveness of AD.
  • Research on prevention and treatment methods and early diagnostic markers that can reflect disease prognosis is progressing slowly.
  • Metabolic abnormalities may be one of the main mechanisms of AD pathogenesis.
  • AD patients abnormal lipid metabolism represented by cholesterol and abnormal glucose metabolism represented by glucose are the most common metabolic abnormalities. They also include cerebral ischemia and hypoxia caused by insufficient blood supply to the brain, represented by degenerative changes in cerebral small blood vessels. energy metabolism disorders.
  • Mitochondria are key organelles for the metabolism of major energy substances. Abnormal mitochondrial function can lead to abnormal energy metabolism and also damage the functions of neurons and cerebral small blood vessel endothelial cells.
  • mitochondrial dysfunction is closely related to abnormalities in lipid metabolism and glucose metabolism. The relationship is not clear.
  • Mild cognitive impairment is a syndrome involving objective cognitive decline. It is a state between mild dementia and healthy aging, with memory impairment as the prominent manifestation. There is a certain degree of decline in other cognitive functions, but the ability to perform daily activities is normal and does not meet the diagnostic criteria for dementia. 10% to 15% of MCI are converted into AD every year, which is 10 times that of normal controls. As a transitional state from normal aging to dementia, MCI has attracted more and more attention. Studying MCI is an important step in the early diagnosis and early intervention of AD.
  • This application provides a new type of blood marker for Alzheimer's disease AD or mild cognitive impairment MCI.
  • the present application provides an intronic retention cleavage product QDLIR of DNA damage-inducible transcript 4-like transcript (DDIT4L), and/or a nucleic acid encoding the QDLIR, which can be used in disease diagnosis and/or disease progression. Use in the evaluation of disorders including cognitive impairment.
  • DIT4L DNA damage-inducible transcript 4-like transcript
  • the present application provides an intronic retention cleavage product QDLIR of DNA damage-inducible transcript 4-like transcript (DDIT4L), and/or a nucleic acid encoding the QDLIR, which can be used in disease diagnosis and/or disease progression. Use in the evaluation of diseases including neurodegenerative diseases.
  • DIT4L DNA damage-inducible transcript 4-like transcript
  • the QDLIR comprises the amino acid sequence set forth in SEQ ID NO: 4.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:3.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:2.
  • the cognitive impairment includes normal aging cognitive impairment, lews physical dementia (LBD), frontotemporal dementia, and/or vascular dementia.
  • LBD lews physical dementia
  • frontotemporal dementia frontotemporal dementia
  • vascular dementia vascular dementia
  • the cognitive impairment-inducing disease includes Alzheimer's disease, multi-infarct disease, Parkinson's disease, AIDS, and/or Creutzfeldt-Jakob disease (CJD).
  • Alzheimer's disease multi-infarct disease
  • Parkinson's disease AIDS
  • Creutzfeldt-Jakob disease CJD
  • the cognitive impairment includes early cognitive impairment (MCI), intermediate cognitive impairment, and late cognitive impairment.
  • MCI early cognitive impairment
  • intermediate cognitive impairment intermediate cognitive impairment
  • late cognitive impairment late cognitive impairment
  • the cognitive impairment includes amnestic MCI with multiple cognitive domain impairment (aMCI-m).
  • the neurodegenerative diseases include acute neurodegenerative diseases and chronic neurodegenerative diseases.
  • the neurodegenerative diseases include neurodegenerative diseases caused by neuronal death and glial cell homeostasis, neurodegenerative diseases caused by aging, neurodegeneration caused by affected CNS cell function. disease, neurodegenerative diseases caused by abnormal communication between cells and/or neurodegenerative diseases caused by impaired cell movement.
  • the neurodegenerative disease includes Alzheimer's disease, Parkinson's disease, multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), and/or Huntington's disease (HD). ).
  • the neurodegenerative disease includes Alzheimer's disease.
  • the neurodegenerative disease includes early Alzheimer's disease, intermediate Alzheimer's disease, and/or late Alzheimer's disease.
  • the present application provides a material for detecting the intron-retained cleavage product QDLIR of DNA damage-induced transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding the QDLIR, which is used in the preparation of diseases.
  • a material for detecting the intron-retained cleavage product QDLIR of DNA damage-induced transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding the QDLIR which is used in the preparation of diseases.
  • the present application provides a material for detecting the intron-retained cleavage product QDLIR of DNA damage-induced transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding the QDLIR, which is used in the preparation of diseases.
  • a material for detecting the intron-retained cleavage product QDLIR of DNA damage-induced transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding the QDLIR which is used in the preparation of diseases.
  • the present application provides a diagnostic kit, which contains the intron retention cleavage product QDLIR capable of detecting DNA damage-inducible transcript 4-like transcript (DDIT4L) described in the present application, and/or encoding the The nucleic acid substance of QDLIR, the diagnostic kit is used for diagnosing cognitive disorders, and/or evaluating the disease process of cognitive disorders; and/or is used for diagnosing neurodegenerative diseases, and/or evaluating neurodegenerative diseases. process.
  • QDLIR intron retention cleavage product QDLIR capable of detecting DNA damage-inducible transcript 4-like transcript (DDIT4L) described in the present application, and/or encoding the The nucleic acid substance of QDLIR
  • the diagnostic kit is used for diagnosing cognitive disorders, and/or evaluating the disease process of cognitive disorders; and/or is used for diagnosing neurodegenerative diseases, and/or evaluating neurodegenerative diseases. process.
  • the diagnostic kit includes instructions for use of the diagnostic kit to diagnose cognitive impairment, and/or to assess the disease progression of cognitive impairment; and/or to diagnose neurodegenerative diseases. , and/or methods for assessing disease progression in neurodegenerative diseases.
  • the QDLIR comprises the amino acid sequence set forth in SEQ ID NO: 4.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:3.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:2.
  • the cognitive impairment includes normal aging cognitive impairment, lews physical dementia (LBD), frontotemporal dementia, and/or vascular dementia.
  • LBD lews physical dementia
  • frontotemporal dementia frontotemporal dementia
  • vascular dementia vascular dementia
  • the cognitive impairment-inducing disease includes Alzheimer's disease, multi-infarct disease, Parkinson's disease, AIDS, and/or Creutzfeldt-Jakob disease (CJD).
  • Alzheimer's disease multi-infarct disease
  • Parkinson's disease AIDS
  • Creutzfeldt-Jakob disease CJD
  • the cognitive impairment includes early cognitive impairment (MCI), intermediate cognitive impairment, and late cognitive impairment.
  • MCI early cognitive impairment
  • intermediate cognitive impairment intermediate cognitive impairment
  • late cognitive impairment late cognitive impairment
  • the cognitive impairment includes amnestic MCI with multiple cognitive domain impairment (aMCI-m).
  • the neurodegenerative diseases include acute neurodegenerative diseases and chronic neurodegenerative diseases.
  • the neurodegenerative diseases include neurodegenerative diseases caused by neuronal death and glial cell homeostasis, neurodegenerative diseases caused by aging, neurodegeneration caused by affected CNS cell function. disease, neurodegenerative diseases caused by abnormal communication between cells and/or neurodegenerative diseases caused by impaired cell movement.
  • the neurodegenerative disease includes Alzheimer's disease, Parkinson's disease, multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), and/or Huntington's disease (HD). ).
  • the neurodegenerative disease includes Alzheimer's disease.
  • the neurodegenerative disease includes early Alzheimer's disease, intermediate Alzheimer's disease, and/or late Alzheimer's disease.
  • the substance includes reagents and/or devices capable of detecting the QDLIR, and/or nucleic acids encoding the QDLIR.
  • the substance includes a reagent that specifically detects the QDLIR, and/or a nucleic acid encoding the QDLIR.
  • the substances include antibodies capable of specifically binding QDLIR, probes capable of specifically binding QDLIR, primers capable of specifically amplifying nucleic acids encoding QDLIR, and genes capable of specifically detecting nucleic acids encoding QDLIR.
  • a chip an aptamer capable of specifically binding to the nucleic acid encoding QDLIR, and/or a guide RNA capable of specifically binding to the nucleic acid encoding QDLIR.
  • the product includes a diagnostic kit.
  • the present application provides a method for diagnosing cognitive impairment, and/or assessing the disease process of cognitive impairment, which includes the following steps: detecting DNA damage-inducing properties in a sample derived from a subject in need The content of the intronic retention cleavage product QDLIR of transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding said QDLIR.
  • the present application provides a method for diagnosing a neurodegenerative disease, and/or assessing the disease process of a neurodegenerative disease, comprising the steps of: detecting DNA damage in a sample derived from a subject in need thereof The content of the intronic retention cleavage product QDLIR of the inducible transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding said QDLIR.
  • the QDLIR comprises the amino acid sequence set forth in SEQ ID NO: 4.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:3.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:2.
  • the cognitive impairment includes normal aging cognitive impairment, lews physical dementia (LBD), frontotemporal dementia, and/or vascular dementia.
  • LBD lews physical dementia
  • frontotemporal dementia frontotemporal dementia
  • vascular dementia vascular dementia
  • the cognitive impairment-inducing disease includes Alzheimer's disease, multi-infarct disease, Parkinson's disease, AIDS, and/or Creutzfeldt-Jakob disease (CJD).
  • Alzheimer's disease multi-infarct disease
  • Parkinson's disease AIDS
  • Creutzfeldt-Jakob disease CJD
  • the cognitive impairment includes early cognitive impairment (MCI), intermediate cognitive impairment, and late cognitive impairment.
  • MCI early cognitive impairment
  • intermediate cognitive impairment intermediate cognitive impairment
  • late cognitive impairment late cognitive impairment
  • the cognitive impairment includes amnestic MCI with multiple cognitive domain impairment (aMCI-m).
  • the neurodegenerative diseases include acute neurodegenerative diseases and chronic neurodegenerative diseases.
  • the neurodegenerative diseases include neurodegenerative diseases caused by neuronal death and glial cell homeostasis, neurodegenerative diseases caused by aging, neurodegeneration caused by affected CNS cell function. disease, neurodegenerative diseases caused by abnormal communication between cells and/or neurodegenerative diseases caused by impaired cell movement.
  • the neurodegenerative disease includes Alzheimer's disease, Parkinson's disease, multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), and/or Huntington's disease (HD). ).
  • the neurodegenerative disease includes Alzheimer's disease.
  • the neurodegenerative disease includes early Alzheimer's disease, intermediate Alzheimer's disease, and/or late Alzheimer's disease.
  • the subject includes a mammal.
  • the subject includes a tumor patient.
  • the subject is aged.
  • the sample includes a blood sample and/or a tissue or cell sample.
  • the sample includes whole blood, serum, plasma, and/or cerebrospinal fluid.
  • the method includes comparing the content of the QDLIR and/or nucleic acid encoding QDLIR in a sample of the subject to a normal control value, wherein the normal control value includes a normal subject.
  • the content of the QDLIR and/or the nucleic acid encoding QDLIR in the subject includes a normal subject.
  • the normal subject does not suffer from the cognitive impairment and/or the neurodegenerative disease.
  • the subject when the QDLIR of the subject's sample, and/or the content of the nucleic acid encoding QDLIR is significantly higher than the normal control value, the subject is diagnosed as suffering from the cognitive disorder. and/or the neurodegenerative diseases described.
  • the method includes: comparing the content of the QDLIR and/or nucleic acid encoding a QDLIR in a sample of the subject to an early control value, wherein the early control value includes the same The subject's previously measured content of said QDLIR and/or nucleic acid encoding QDLIR.
  • the subject has been diagnosed with the cognitive disorder and/or the neurodegenerative disease.
  • the subject is diagnosed with the cognitive impairment and/or cognitive impairment when the QDLIR of the subject's sample, and/or the content of the nucleic acid encoding QDLIR is significantly higher than the early control value. /or exacerbation of the disease process of said neurodegenerative disease.
  • the method further includes the step of detecting the content of markers associated with cognitive impairment and/or neurodegenerative diseases in a sample derived from the subject.
  • the markers associated with cognitive impairment and/or neurodegenerative diseases include AD7C-NTP, pTau-181, pTau-217 and/or A ⁇ 1-42.
  • the method further includes the step of observing brain imaging of the subject.
  • the present application provides a system for diagnosing cognitive impairment, and/or assessing the disease process of cognitive impairment, comprising detecting intron retention splicing of DNA damage-inducible transcript 4-like transcript (DDIT4L) The product QDLIR, and/or the nucleic acid encoding said QDLIR.
  • DDIT4L DNA damage-inducible transcript 4-like transcript
  • the present application provides a system for diagnosing neurodegenerative diseases, and/or assessing the disease progression of neurodegenerative diseases, comprising detecting intron retention of DNA damage-inducible transcript 4-like transcript (DDIT4L) The cleavage product QDLIR, and/or the nucleic acid encoding said QDLIR.
  • DDIT4L DNA damage-inducible transcript 4-like transcript
  • the QDLIR comprises the amino acid sequence set forth in SEQ ID NO: 4.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:3.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:2.
  • the cognitive impairment includes normal aging cognitive impairment, lews physical dementia (LBD), frontotemporal dementia, and/or vascular dementia.
  • LBD lews physical dementia
  • frontotemporal dementia frontotemporal dementia
  • vascular dementia vascular dementia
  • the cognitive impairment-inducing disease includes Alzheimer's disease, multi-infarct disease, Parkinson's disease, AIDS, and/or Creutzfeldt-Jakob disease (CJD).
  • Alzheimer's disease multi-infarct disease
  • Parkinson's disease AIDS
  • Creutzfeldt-Jakob disease CJD
  • the cognitive impairment includes early cognitive impairment (MCI), intermediate cognitive impairment, and late cognitive impairment.
  • MCI early cognitive impairment
  • intermediate cognitive impairment intermediate cognitive impairment
  • late cognitive impairment late cognitive impairment
  • the cognitive impairment includes amnestic MCI with multiple cognitive domain impairment (aMCI-m).
  • the neurodegenerative diseases include acute neurodegenerative diseases and chronic neurodegenerative diseases.
  • the neurodegenerative diseases include neurodegenerative diseases caused by neuronal death and glial cell homeostasis, neurodegenerative diseases caused by aging, neurodegeneration caused by affected CNS cell function. disease, neurodegenerative diseases caused by abnormal communication between cells and/or neurodegenerative diseases caused by impaired cell movement.
  • the neurodegenerative disease includes Alzheimer's disease, Parkinson's disease, multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), and/or Huntington's disease (HD). ).
  • the neurodegenerative disease includes Alzheimer's disease.
  • the neurodegenerative disease includes early Alzheimer's disease, intermediate Alzheimer's disease, and/or late Alzheimer's disease.
  • the system is in the form of a kit.
  • the system includes a collection module capable of collecting samples from a subject in need thereof.
  • the subject includes a mammal.
  • the subject includes a tumor patient.
  • the subject is aged.
  • the sample includes a blood sample and/or a tissue or cell sample.
  • the sample includes whole blood, serum, plasma, and/or cerebrospinal fluid.
  • the system includes a detection module capable of detecting the content value of the QDLIR and/or the nucleic acid encoding the QDLIR.
  • the detection module detects the content value of the QDLIR and/or the nucleic acid encoding the QDLIR in the sample obtained by the collection module.
  • the detection module includes a substance capable of detecting the content of the QDLIR and/or the nucleic acid encoding the QDLIR.
  • the substances include antibodies capable of specifically binding QDLIR, probes capable of specifically binding QDLIR, primers capable of specifically amplifying nucleic acids encoding QDLIR, and genes capable of specifically detecting nucleic acids encoding QDLIR.
  • a chip an aptamer capable of specifically binding to the nucleic acid encoding QDLIR, and/or a guide RNA capable of specifically binding to the nucleic acid encoding QDLIR.
  • the detection module includes an instrument capable of detecting the intronic retention cleavage product QDLIR of DNA damage-inducible transcript 4-like transcript (DDIT4L), and/or the content of a nucleic acid encoding the QDLIR .
  • DIT4L DNA damage-inducible transcript 4-like transcript
  • the detection module includes substances and/or instruments capable of detecting levels of markers associated with cognitive impairment and/or neurodegenerative diseases.
  • the markers associated with cognitive impairment and/or neurodegenerative diseases include AD7C-NTP, pTau-181, pTau-217 and/or A ⁇ 1-42.
  • the detection module outputs a content value of the QDLIR and/or the nucleic acid encoding the QDLIR.
  • the system includes a judgment module, which can judge whether the subject is affected based on the content of the QDLIR and/or the nucleic acid encoding the QDLIR and the normal control value and/or early control value.
  • the diagnostic results and/or disease progression results of the subject wherein, the normal control value includes the content of the QDLIR and/or the nucleic acid encoding QDLIR in normal subjects; wherein the early control value includes the same The subject's previously measured content of said QDLIR and/or nucleic acid encoding QDLIR.
  • the normal subject does not suffer from the cognitive impairment and/or the neurodegenerative disease.
  • the judgment module diagnoses the subject as suffering from the disease. cognitive impairment and/or neurodegenerative diseases.
  • the subject has been diagnosed with the cognitive disorder and/or the neurodegenerative disease.
  • the judgment module diagnoses the subject as the Cognitive impairment and/or exacerbation of the disease process in said neurodegenerative diseases.
  • the determination module outputs a diagnosis result of the subject.
  • the system includes a display module, wherein the display module can display the diagnosis result of the judgment module.
  • the system includes a storage module, wherein the storage module can store the content value output by the detection module, and/or the diagnostic result output by the judgment module.
  • the present application provides a drug screening method, which includes the following steps: detecting changes in the content of QDLIR and/or nucleic acids encoding QDLIR in a subject after administration of a candidate drug, wherein the drug is used to prevent, Treat and/or alleviate cognitive impairment and/or neurodegenerative diseases.
  • the QDLIR comprises the amino acid sequence set forth in SEQ ID NO: 4.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:3.
  • the nucleic acid encoding QDLIR comprises the nucleotide sequence set forth in SEQ ID NO:2.
  • the cognitive impairment includes normal aging cognitive impairment, lews physical dementia (LBD), frontotemporal dementia, and/or vascular dementia.
  • LBD lews physical dementia
  • frontotemporal dementia frontotemporal dementia
  • vascular dementia vascular dementia
  • the cognitive impairment-inducing disease includes Alzheimer's disease, multi-infarct disease, Parkinson's disease, AIDS, and/or Creutzfeldt-Jakob disease (CJD).
  • Alzheimer's disease multi-infarct disease
  • Parkinson's disease AIDS
  • Creutzfeldt-Jakob disease CJD
  • the cognitive impairment includes early cognitive impairment (MCI), intermediate cognitive impairment, and late cognitive impairment.
  • MCI early cognitive impairment
  • intermediate cognitive impairment intermediate cognitive impairment
  • late cognitive impairment late cognitive impairment
  • the cognitive impairment includes amnestic MCI with multiple cognitive domain impairment (aMCI-m).
  • the neurodegenerative diseases include acute neurodegenerative diseases and chronic neurodegenerative diseases.
  • the neurodegenerative diseases include neurodegenerative diseases caused by neuronal death and glial cell homeostasis, neurodegenerative diseases caused by aging, neurodegeneration caused by affected CNS cell function. disease, neurodegenerative diseases caused by abnormal communication between cells and/or neurodegenerative diseases caused by impaired cell movement.
  • the neurodegenerative disease includes Alzheimer's disease, Parkinson's disease, multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), and/or Huntington's disease (HD). ).
  • the neurodegenerative disease includes Alzheimer's disease.
  • the neurodegenerative disease includes early Alzheimer's disease, intermediate Alzheimer's disease, and/or late Alzheimer's disease.
  • a drug candidate has a therapeutic effect if the amount of QDLIR and/or nucleic acid encoding QDLIR is reduced in a subject when the drug candidate is administered.
  • the administration includes injection.
  • the drugs include small molecule drugs and/or biomacromolecule drugs.
  • a substance for detecting the intronic retained cleavage product QDLIR encoding DNA damage-inducible transcript 4-like transcript in a subject sample prepared for use in progeria in the subject.
  • kits for diagnosis of dementia, diagnosis of mild cognitive impairment and/or assessment of progression (e.g. grading or staging) of Alzheimer's disease.
  • the subject is selected from the group consisting of humans, non-human primates, rodents (eg, rats, mice, guinea pigs), dogs, cats, horses, cattle, and sheep.
  • rodents eg, rats, mice, guinea pigs
  • dogs, cats, horses, cattle, and sheep e.g., horses, cattle, and sheep.
  • the subject does not suffer from cancer.
  • the subject is free of malignancy.
  • the QDLIR has a sequence selected from the group consisting of: a nucleotide sequence (mRNA) having SEQ ID NO: 2; a nucleotide sequence corresponding to the QDLIR peptide having SEQ ID NO: 3 Encoding nucleotide molecules; or, QDLIR peptides with the amino acid sequence shown in SEQ ID NO:4; variants obtained by one or more nucleotide and/or amino acid substitutions, deletions and/or additions to the above sequence; and the above Sequences having a high degree of homology (eg, greater than 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 99.5% sequence homology).
  • mRNA nucleotide sequence having SEQ ID NO: 2
  • a nucleotide sequence corresponding to the QDLIR peptide having SEQ ID NO: 3 Encoding nucleotide molecules or, QDLIR peptides with the amino acid sequence shown in SEQ ID NO:4; variants obtained
  • the Alzheimer's disease includes early stage (amnesia stage), middle stage (neurological disorder stage) and late stage (severe dementia stage).
  • the substance is selected from reagents, devices, or combinations thereof for detecting QDLIR.
  • the substance is a substance for detecting the QDLIR at the gene level and/or protein level.
  • the substance is a substance for use in one or more detection techniques or methods selected from the group consisting of immunohistochemistry (e.g., immunofluorescence assay, chemiluminescence assay, reverse enzyme-linked immunosorbent assay, immunocolloidal gold method), Western blotting, Northern blotting, PCR, biochip method, protein chip method.
  • immunohistochemistry e.g., immunofluorescence assay, chemiluminescence assay, reverse enzyme-linked immunosorbent assay, immunocolloidal gold method
  • Western blotting e.g., Western blotting, Northern blotting, PCR, biochip method, protein chip method.
  • the substance is selected from: substances specific to the QDLIR, such as antibodies (such as monoclonal antibodies) to the QDLIR; shear product-specific probes, gene chips, PCR primers, gRNA, etc. .
  • substances specific to the QDLIR such as antibodies (such as monoclonal antibodies) to the QDLIR; shear product-specific probes, gene chips, PCR primers, gRNA, etc. .
  • the sample is selected from: blood samples, such as whole blood, serum, plasma; tissue or cell samples; cerebrospinal fluid.
  • the blood sample is fresh blood or frozen blood.
  • an increased level of the QDLIR in the sample compared to normal control values indicates that the subject suffers from Alzheimer's disease or mild cognitive impairment.
  • the level of QDLIR in the sample is increased compared to a mild cognitive impairment control or compared to pre-control levels in a subject diagnosed with Alzheimer's disease, indicating that the subject has developed Alzheimer's disease. (e.g. higher grade or stage).
  • an increase in the level of the QDLIR in the sample as compared to a control value at a lower grade or stage is indicative of progression of Alzheimer's disease in the subject (eg, higher grade or stage).
  • the present application provides a diagnostic kit, which contains the intron retention cleavage product QDLIR capable of detecting DNA damage-inducible transcript 4-like transcript (DDIT4L) described in the present application, and/or encoding the Said QDLIR is a nucleic acid substance.
  • the present application provides a product (such as a kit) for the diagnosis of Alzheimer's disease, the diagnosis of mild cognitive impairment, and/or the assessment of the development process (such as grading or staging) of Alzheimer's disease, which includes:
  • the substance in (i) is one or more selected from the group consisting of:
  • Substances for detecting said QDLIR at the gene level and/or protein level such as substances used in one or more detection techniques or methods selected from the following group: immunohistochemistry (e.g. immunofluorescence analysis, chemical Luminescence method, reverse enzyme-linked immunosorbent assay, immunocolloidal gold method), Western blotting method, Northern blotting method, PCR, biochip method; or
  • Substances specific to the QDLIR such as its antibodies (preferably monoclonal antibodies); probes, gene chips, PCR primers, and gRNA specific to the sheared product.
  • other substances in (ii) are selected from: substances for detecting one or more other Alzheimer's disease markers, such as: AD7C-NTP, pTau-181, pTau-217, A ⁇ 1-42 ; and/or, reagents for brain amyloid imaging; and/or, reagents for subject brain imaging diagnosis.
  • substances for detecting one or more other Alzheimer's disease markers such as: AD7C-NTP, pTau-181, pTau-217, A ⁇ 1-42 ; and/or, reagents for brain amyloid imaging; and/or, reagents for subject brain imaging diagnosis.
  • a method of screening Alzheimer's disease therapeutic drugs comprising: detecting the effect of a candidate drug on QDLIR levels in a subject, wherein, after using the candidate drug, the level A decrease indicates that the drug candidate has a therapeutic effect on Alzheimer's disease.
  • the detection is performed using a product as described herein.
  • a method for diagnosing Alzheimer's disease, diagnosing mild cognitive impairment, and/or assessing the progression of cognitive impairment comprising: detecting QDLIR in a sample from the subject level, and based on this, the subject is diagnosed with Alzheimer's disease and/or the progression of Alzheimer's disease (such as grading or staging) is assessed.
  • the detection is performed using a product as described herein.
  • an increase in the level of the QDLIR in the sample compared to a control value indicates that the subject suffers from Alzheimer's disease; or, compared to a low-grade or low-stage control, the level of the QDLIR in the sample is elevated.
  • An increased level of QDLIR indicates progression of Alzheimer's disease in the subject (eg, higher grade or stage).
  • a system corresponding to the method of this application is also provided, which includes the following devices:
  • a device for analyzing data to diagnose Alzheimer's disease, diagnose mild cognitive impairment, and/or assess the progression (e.g., grade or stage) of Alzheimer's disease in the subject wherein, compared with a normal control Compared with the value, the level of the QDLIR in the sample is increased, indicating that the subject suffers from Alzheimer's disease or mild cognitive impairment; or, compared with a mild cognitive impairment control or a subject diagnosed with Alzheimer's disease An increase in the level of QDLIR in the sample compared to the previous control level indicates that the subject has developed Alzheimer's disease (eg, a higher grade or stage).
  • the system further includes (a') a device for collecting and/or receiving data on other clinical indicators or basic characteristics of the diagnosis of Alzheimer's disease and/or the progression of Alzheimer's disease in the sample.
  • the device described in (b) is used to combine QDLIR level data with other clinical indicators or basic characteristic data of Alzheimer's disease diagnosis and/or Alzheimer's disease development process to analyze the subject's Prognostic assessment of Alzheimer's disease diagnosis and/or progression of Alzheimer's disease.
  • the system further includes equipment for analyzing other clinical indicators or basic characteristic data of Alzheimer's disease diagnosis and/or Alzheimer's disease development process; and/or comparing QDLIR level data with Alzheimer's disease.
  • a device that combines and analyzes other clinical indicators or basic characteristic data of the diagnosis of dementia and/or the development process of Alzheimer's disease to evaluate the diagnosis of Alzheimer's disease and/or the development process of Alzheimer's disease of the subject.
  • the system further includes one or more devices selected from the group consisting of: a device for inputting and/or outputting and/or storing data; a device for outputting and/or storing analysis results, For example, a device for uploading and/or storing analysis results to a cloud database or its corresponding cloud database device; a device for intelligently analyzing and recommending treatment plans based on prognosis assessment results.
  • a device for inputting and/or outputting and/or storing data For example, a device for uploading and/or storing analysis results to a cloud database or its corresponding cloud database device; a device for intelligently analyzing and recommending treatment plans based on prognosis assessment results.
  • Figure 1a-1b shows the splicing mode of DDIT4L pre-mRNA and the secretion of QDLIR protein from transfected cells into the culture medium.
  • Figure a The splicing method of DDIT4L, in which the length of the mRNA obtained by removing introns through normal splicing is 582nt (SEQ ID NO: 5), and the mRNA obtained through abnormal intron retention (Intron retention) splicing is involved in coding
  • the RNA length of QDLIR is 255nt (SEQ ID NO:3);
  • FIG. b QDLIR is secreted from cells into the culture medium.
  • the pCMV-flag-QDLIR plasmid was transfected into HEK293 cells. After 48 hours of culture, it was found that the secreted QDLIR protein could be detected in the culture medium.
  • the QDLIR protein band can be seen in the QDLIR plasmid lane of the lysate, and its size is 12kDa.
  • FIGS 2a-2b show the changes in QDLIR protein levels in the blood of AD patients and patients with various other diseases.
  • Figure a Western detection of QDLIR content changes in the blood of AD patients and patients with various other diseases
  • the plasma protein in the figure is the light chain of the immunoglobulin used as an internal reference.
  • Figure b ELISA was used to detect changes in QDLIR levels in the blood of AD patients, MCI patients and non-AD/MCI patients. ***in the figure means P ⁇ 0.001, * means P ⁇ 0.05.
  • Figure 3 shows the standard curve for the ELISA detection of the standard QDLIR protein.
  • Figure 4 shows the R°C curve of ELISA detection of clinical samples.
  • Figure 5 shows the content changes of QDLIR in the blood of different types of MCI patients.
  • Figure 6 shows the content changes of QDLIR and other indicators in the blood of different categories of MCI and AD patients.
  • Figure 7 shows the correlation of QDLIR expression levels in plasma and cerebrospinal fluid.
  • DDIT4L usually refers to DNA damage-inducible transcript 4-like. It may also be called REDD2/RTP801L. Studies have found that DDIT4L can be associated with cardiac dysfunction. It can also be used to treat gliomas.
  • accession number of human DDIT4L gene in GenBank is 115265; the accession number of human DDIT4L protein in GenBank is NP_660287.1.
  • QDLIR generally refers to the intronic retention cleavage product of DDIT4L.
  • amino acid sequence of the QDLIR can be as shown in SEQ ID NO: 4.
  • expression level generally refers to the protein, RNA or mRNA level of a specific gene of interest. Any method known in the art can be used to determine the expression level of a specific relevant gene (eg, human ATP6V1B2 gene). In this application, “expression” generally refers to the process of converting the information encoded by a gene into a structure that exists in and operates in a cell.
  • Examples may include reverse transcription and amplification analysis (such as 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. Analysis can be performed directly on biological samples or on proteins/nucleic acids isolated from the samples.
  • reverse transcription and amplification analysis such as PCR, ligation RT-PCR or quantitative RT-PCT
  • hybridization analysis such as PCR, ligation RT-PCR or quantitative RT-PCT
  • Northern blotting such as PCR, ligation RT-PCR or quantitative RT-PCT
  • dot blotting dot blotting
  • in situ hybridization such as gel electrophoresis, capillary electrophoresis, Column chromatography, Western blotting, immunohistochemistry, immunostaining, or mass spectrome
  • neurodegenerative disease generally refers to cognitive disorders such as dementia resulting from the progressive loss of neuronal structure and function, including neuronal death and glial cell homeostasis.
  • age e.g., Alzheimer's disease (AD), Parkinson's disease (PD)
  • genetic mutations that affect CNS cell function e.g., Huntington's disease, early-onset AD or PD, amyotrophic Lateral sclerosis (ALS) can cause this neurodegenerative disease.
  • AD Alzheimer's disease
  • PD Parkinson's disease
  • ALS amyotrophic Lateral sclerosis
  • the neurodegenerative diseases may have changes and/or disorders selected from the group consisting of: protein misfolding and aggregation; neuroinflammation (e.g., occurring in response to toxic stimuli (e.g., protein aggregation), infection, traumatic injury, or autoimmune signaling) CNS inflammation); changes in cell signaling; acquired senescence/cell death (e.g., disrupted apoptotic signaling, mitochondrial dysfunction, impaired autophagy, and activation of necrosomes by stress/inflammation); exercise Cell damage and epigenetic changes.
  • neuroinflammation e.g., occurring in response to toxic stimuli (e.g., protein aggregation), infection, traumatic injury, or autoimmune signaling) CNS inflammation
  • changes in cell signaling e.g., acquired senescence/cell death (e.g., disrupted apoptotic signaling, mitochondrial dysfunction, impaired autophagy, and activation of necrosomes by stress/inflammation); exercise Cell damage and epigenetic
  • Alzheimer's disease usually refers to Alzheimer's disease, a neurodegenerative disease that develops slowly and worsens over time.
  • the most common early symptom is loss of short-term memory (difficulty remembering recent events).
  • the true cause of Alzheimer's disease remains unknown, but its progression may be related to the deposition of fibrillar amyloid protein plaques and tau protein in the brain.
  • There are currently no treatments that can stop or reverse the course of the disease but there are a few treatments that may temporarily relieve or improve symptoms.
  • Alzheimer's disease may include early Alzheimer's disease, middle Alzheimer's disease and/or late Alzheimer's disease.
  • the learning and memory impairments of patients with early Alzheimer's disease will become more and more obvious. In some cases, they will have language impairment, executive impairment, cognitive impairment (agnosia) and/or skill execution impairment (apraxia).
  • patients in the early stages of Alzheimer's disease will lose the ability to live independently and may be unable to perform most daily activities (in some cases, may suffer from anomia, disarranged speech, and/or hypoaesthesia).
  • patients with advanced Alzheimer's disease may develop late dependence on caregivers. For example, there may be a complete loss of language skills. For example, you may not be able to feed yourself.
  • cognitive impairment generally refers to diseases and conditions that are believed to or do involve or are related to the progressive loss of neuronal structure and/or function (including neuronal death).
  • the cognitive impairment may be characterized by impairment of cognition (eg, memory, attention, perception, and/or thinking). These disorders can include pathogen-induced cognitive dysfunction, such as HIV-related cognitive dysfunction and Lyme disease-related cognitive dysfunction. Examples of cognitive disorders may include Alzheimer's disease, Huntington's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS), autism, early cognitive impairment (MCI), stroke, traumatic brain injury (TBI) and/or age-related memory impairment (AAMI).
  • ALS amyotrophic lateral sclerosis
  • MCI early cognitive impairment
  • TBI traumatic brain injury
  • AAMI age-related memory impairment
  • MCI middle 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 MCI may include a subtype selected from the group consisting of: aMCI-s: amnestic MCI with impairment of a single cognitive domain; aMCI-m: amnestic MCI with impairment of multiple cognitive domains; naMCI-s : Non-amnestic MCI with single cognitive domain impairment; and naMCI-m: Non-amnestic MCI with multiple cognitive domain impairments.
  • 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.
  • lews body dementia usually refers to Lewy Body Detmentia.
  • Dementia with Lewy bodies is characterized by abnormal accumulation of proteins into lumps called Lewy bodies. Dementia with Lewy bodies causes a gradual decline in mental abilities. People with Lewy body dementia may experience visual hallucinations and changes in alertness and concentration. Other effects include muscle stiffness, slowness of movement, difficulty walking and tremors. Patients with Lewy bodies in the brain can also have plaques and tangles associated with Alzheimer's disease.
  • frontotemporal dementia usually refers to Pick's disease, a rare, progressive disease in which the tau protein affects only the frontal and temporal lobes of the brain.
  • People with frontotemporal dementia have difficulty with higher-level reasoning, expressive language, speech perception, and memory formation.
  • the frontal and temporal lobes of the brain can shrink over time.
  • vascular dementia usually refers to problems with reasoning, judgment, and memory caused by impaired blood flow to the brain.
  • the vascular dementia may include dementia due to risk factors for heart disease and stroke, such as high blood pressure and high cholesterol.
  • multiple infarct type generally refers to small noncortical infarcts caused by occlusion of a single perforating branch of a large cerebral artery.
  • the multiple infarction type may be a special type of cerebral infarction, also known as ischemic stroke.
  • the multiple infarction type may manifest as hemi-sensory impairment, aphasia, hypoarthria, slow movement, and clumsiness (especially difficulty with fine movements such as writing).
  • Parkinson's disease generally refers to a progressive neurodegenerative disease.
  • Clinical features of Parkinson's disease can include motor symptoms (eg, tremor, bradykinesia, myotonia, and postural instability), as well as neuropsychiatric and other non-motor manifestations.
  • the non-motor manifestations may include cognitive dysfunction and dementia, mood disorders (eg, depression, anxiety, apathy), and sleep disorders.
  • CJD Crohn's disease
  • MS multiple sclerosis
  • the term “multiple sclerosis (MS)” generally refers to a demyelinating neuropathy.
  • the insulating material (myelin sheath) on the surface of nerve cells in the brain or spinal cord of MS patients is damaged, and the signal transduction of the nervous system is damaged, which can lead to a series of possible symptoms, affecting the patient's activity, mind, and even spirit. state. These symptoms can include double vision, impaired vision on one side, muscle weakness, dulled sensation, or coordination problems.
  • ALS amyotrophic lateral sclerosis
  • ALS amyotrophic lateral sclerosis
  • a small number of ALS patients may develop frontotemporal lobe dementia. Some ALS patients will experience deterioration in their senses of sight, touch, smell, and taste, and a very small number of amyotrophic lateral sclerosis patients will develop dementia at the same time.
  • HD Huntington's disease
  • the term "aging stage” generally refers to a subject's stage of aging.
  • the old age stage can be over 60 years old, over 70 years old, or over 75 years old; for mice, the old age stage can be over 10 months old, for example, over 13 months old, or 18 months and above.
  • the elderly subjects may have one or more symptoms of learning deficits, memory impairment, memory deficits, and/or brain dysfunction.
  • variant generally refers to a polypeptide comprising an amino acid sequence that differs by at least one amino acid residue from the amino acid sequence of a parent or reference polypeptide (eg, a wild-type polypeptide).
  • the variant may have a high (eg, at least 80%) homology to the parent or reference polypeptide.
  • the homology may include sequence similarity or identity.
  • said homology can be determined using standard techniques known in the art (see, e.g., Smith and Waterman, Adv. Appl. Math.
  • 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]).
  • Software for performing BLAST analyzes is publicly available through the National Center for Biotechnology Information (NCBI).
  • the prerequisite for early detection and early treatment of Alzheimer's disease is to find its specific diagnostic markers.
  • the inventor found that QDLIR protein was present in the blood of Alzheimer's patients, but was not detected in the blood samples of normal people. This result shows that QDLIR is likely to become a blood marker for the diagnosis of Alzheimer's disease (possibly early diagnosis); at the same time, the inventor also tested the blood samples of other cognitive impairment patients and found that in MCI (mild cognitive impairment, mild cognitive impairment) The level of QDLIR protein in blood samples of patients with cognitive impairment such as Alzheimer's disease is significantly elevated. Therefore, elevated blood QDLIR will be a new marker in the diagnosis and treatment of Alzheimer's disease.
  • QDLIR is an abnormal protein formed by abnormal splicing in the human body and only exists in human tissues and cells.
  • QDLIR is a key molecule that can adaptively regulate the functions of cells and tissues through high expression and secretion in cellular hypoxic environments.
  • QDLIR was found in the detection of blood samples from other patients with cognitive impairment.
  • the protein level is significantly increased, so the QDLIR level in the blood can be used as a new marker in the diagnosis and treatment of AD (Alzheimer's disease).
  • 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 neurodegenerative disease may include Alzheimer's disease.
  • the neurodegenerative disease may include early Alzheimer's disease, middle Alzheimer's disease, and/or late Alzheimer's disease.
  • 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 due to normal aging, lews physical dementia (LBD), frontotemporal dementia, and/or vascular dementia.
  • LBD lews physical dementia
  • the cognitive impairment-inducing disease may include Alzheimer's disease, multi-infarct type, Parkinson's disease, AIDS, and/or Creutzfeldt-Jakob disease (CJD).
  • the cognitive impairment may include early cognitive impairment (MCI) (e.g., 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 needing reminders from others for basic self-care), medium-term cognitive impairment (for example, confusion between long-term memory and memory of real-life situations, inability to express oneself, changes in behavior or personality, or emotional instability and/or need for assistance in self-care) or Advanced cognitive impairment (e.g., 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).
  • MCI early cognitive impairment
  • MCI early cognitive impairment
  • medium-term cognitive impairment for example, confusion between long-term memory and memory of real-life situations, inability to express oneself, changes in behavior or personality, or emotional instability and/or need for assistance in self-care
  • Advanced cognitive impairment e.g., memory impairment, decline in physical activity and mental status, inability to
  • the cognitive impairment may include suffering from a disease that causes the induction of the cognitive impairment.
  • the subject may have Alzheimer's disease, multi-infarct disease, Parkinson's disease, AIDS, and/or Creutzfeldt-Jakob disease (CJD).
  • CJD Creutzfeldt-Jakob disease
  • the present application provides an intronic retention cleavage product QDLIR of DNA damage-inducible transcript 4-like transcript (DDIT4L), and/or a nucleic acid encoding the QDLIR, which can be used in disease diagnosis and/or disease progression. Use in assessment where the disease may include cognitive impairment.
  • DIT4L DNA damage-inducible transcript 4-like transcript
  • the present application provides an intronic retention cleavage product QDLIR of DNA damage-inducible transcript 4-like transcript (DDIT4L), and/or a nucleic acid encoding the QDLIR, which can be used in disease diagnosis and/or disease progression.
  • the disease may include neurodegenerative diseases.
  • DDIT4L DNA Damage Inducible Transcript 4 Like, SEQ ID NO:1
  • SEQ ID NO:1 is a protein-coding gene related to the mTOR signal transduction pathway, and its levels in macrophages can be up-regulated by oxidized LDL and hypoxia.
  • the nucleotide sequence (SEQ ID NO:5) encoding the 193aa long DDIT4L protein (SEQ ID NO:6) can be obtained.
  • this gene also has another special splicing mode, namely intron retention splicing.
  • the protein obtained by intron retention cleavage may be called "QDLIR" (as shown in Figure 1a).
  • QDLIR may include mature mRNA, spliced peptide encoding molecules or spliced product peptides (or QDLIR proteins) obtained by intron-retentive splicing of the DDIT4L gene. ).
  • the cleavage product can be mRNA
  • it can be selected from: (a) a nucleotide molecule with a sequence as shown in SEQ ID NO: 2; or (b) a nucleotide molecule with a high sequence as SEQ ID NO: 2 Nucleotide molecules that are identical (eg, sequence identity may be at least 80%, 85%, 90%, 92%, 95%, 98%, 99%, 99.5%, 99.8%).
  • the cleavage product can be a QDLIR encoding molecule
  • it can be selected from: (a) a nucleotide molecule with a sequence as shown in SEQ ID NO: 3; or (b) a nucleotide molecule having the same sequence as SEQ ID NO: 3 Nucleotide molecules with high sequence identity (eg, sequence identity may be at least 80%, 85%, 90%, 92%, 95%, 98%, 99%, 99.5%, 99.8%).
  • the cleavage product can be a QDLIR protein
  • it can be selected from: (a') a polypeptide molecule with a sequence shown in SEQ ID NO: 4; (b') having high sequence identity with the polypeptide molecule of SEQ ID NO: 4 (For example, the sequence identity can be at least 80%, 85%, 90%, 92%, 95%, 98%, 99%, 99.5%, 99.8%) protein molecules; (c') is composed of the above (a) or ( b) a protein molecule obtained by translation of any nucleotide molecule; or (d') a conservative variant polypeptide of the protein molecule described in any one of (a') to (c') above, or its homologous protein; or polypeptide, or active fragment thereof.
  • the present application provides a material for detecting the intron-retained cleavage product QDLIR of DNA damage-induced transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding the QDLIR, which is used in the preparation of diseases.
  • a material for detecting the intron-retained cleavage product QDLIR of DNA damage-induced transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding the QDLIR which is used in the preparation of diseases.
  • the present application provides a material for detecting the intron-retained cleavage product QDLIR of DNA damage-induced transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding the QDLIR, which is used in the preparation of diseases.
  • a material for detecting the intron-retained cleavage product QDLIR of DNA damage-induced transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding the QDLIR which is used in the preparation of diseases.
  • Use in products for diagnosis and/or assessment of disease progression which may include neurodegenerative diseases.
  • detection substance As used in this application, the terms “detection substance”, “detection reagent” or “reagent for detecting QDLIR” or “reagent for detecting QDLIR expression” are used interchangeably. They all refer to specific QDLIR molecules and can be used to directly or substances that indirectly detect the presence and/or content of QDLIR molecules. These detection substances can detect QDLIR at the gene level or protein level.
  • the substances may include antibodies capable of specifically binding QDLIR, probes capable of specifically binding QDLIR, primers capable of specifically amplifying nucleic acids encoding QDLIR, and gene chips capable of specifically detecting nucleic acids encoding QDLIR. , an aptamer capable of specifically binding to the nucleic acid encoding QDLIR, and/or a guide RNA capable of specifically binding to the nucleic acid encoding QDLIR.
  • detection reagents available in this application may include, but are not limited to: antibodies with detection specificity for QDLIR molecules (for example, monoclonal antibodies), probes, gene chips, PCR primers, gRNA, etc.
  • the detection reagents of the present application can also be equipped with detectable labels.
  • the detectable labels can include but are not limited to: radioactive isotopes, fluorophores, chemiluminescent moieties, enzymes, enzyme substrates, and enzyme cofactors. , enzyme inhibitors, dyes, metal ions, ligands (such as biotin or hapten), etc.
  • the detection reagent of the present application can exist in a solution, fixed on a carrier (such as a substrate, an adsorbent), or in other conventional ways in this field, as long as the way of existence is suitable for QDLIR detection in biological samples.
  • a carrier such as a substrate, an adsorbent
  • the detection reagent of the present application can be a nucleotide probe, it can exist in the form of a biochip (or "microarray").
  • the product may be in the form of a kit.
  • the present application also provides a product for diagnosis of Alzheimer's disease and/or assessment of the development process (such as grading or staging) of Alzheimer's disease, which may include: a substance for detecting QDLIR levels, and optionally, Other substances used for the diagnosis of Alzheimer's disease and/or the assessment of the progression of Alzheimer's disease (such as grading or staging), such as detection substances for existing AD markers.
  • the diagnostic kit may include instructions describing the use of the diagnostic kit to diagnose cognitive impairment, and/or to assess the disease process of cognitive impairment; and/or to diagnose neurodegenerative diseases, and/or methods for assessing disease progression in neurodegenerative diseases.
  • appropriate QDLIR detection substances can be selected and made into products suitable for the detection method used, such as kits.
  • Persons of ordinary skill in the art can adjust and change the detection method and the reagents contained in the product according to actual conditions and needs.
  • the present application also provides a product (such as a detection kit), which may include: (i) a detecting effective amount of one or more reagents for detecting QDLIR; (ii) optionally, One or more substances selected from the group consisting of containers, instructions for use, positive controls, negative controls, buffers, auxiliaries or solvents, such as solutions for suspending or fixing cells, detectable labels or markers , a solution that facilitates hybridization of nucleic acids, a solution used to lyse cells, or a solution used for nucleic acid purification.
  • a detection kit may include: (i) a detecting effective amount of one or more reagents for detecting QDLIR; (ii) optionally, One or more substances selected from the group consisting of containers, instructions for use, positive controls, negative controls, buffers, auxiliaries or solvents, such as solutions for suspending or fixing cells, detectable labels or markers , a solution that facilitates hybridization of nucleic acids, a solution used to lyse cells,
  • the present application provides a detection kit suitable for detecting the expression of QDLIR protein in a biological sample by immunohistochemistry.
  • the detection kit may include: blocking solution, such as 10% goat serum; primary antibody, such as rabbit anti-human or mouse D4LIR monoclonal antibody; secondary antibody, such as labeled (such as HRP labeled) or unlabeled goat anti-rabbit Secondary antibody; substrate buffer, such as DAB substrate buffer; chromogenic solution; and optional containers containing the above reagents and instructions for use.
  • the detection kit of the present application can also be accompanied by an instruction manual for the kit, which records how to use the kit to perform detection, and how to use the detection results to diagnose cognitive impairment, and/or evaluate the disease process of cognitive impairment, and/ or to diagnose neurodegenerative diseases, and/or to assess the disease progression of neurodegenerative diseases.
  • the kits described herein can be used for AD diagnosis and/or assessment of AD progression (eg, grading or staging).
  • Products of the present application may also contain other reagents clinically used for AD diagnosis and/or assessment of AD progression (eg, grading or staging) in subjects to assist or validate the results obtained by detecting QDLIR.
  • reagents clinically used for AD diagnosis and/or assessment of AD progression eg, grading or staging
  • grading or staging e.g., grading or staging
  • the present application provides a method for diagnosing cognitive impairment, and/or assessing the disease process of cognitive impairment, which may include the following steps: detecting DNA damage-induced DNA damage in a sample derived from a subject in need thereof The intronic retention cleavage product QDLIR of the sex transcript 4-like transcript (DDIT4L), and/or the content of the nucleic acid encoding said QDLIR.
  • the present application provides a method for diagnosing a neurodegenerative disease, and/or assessing the disease process of a neurodegenerative disease, which may include the following steps: detecting DNA in a sample derived from a subject in need thereof The content of the intronic retention cleavage product QDLIR of the damage-inducible transcript 4-like transcript (DDIT4L), and/or the nucleic acid encoding said QDLIR.
  • the subject may include a mammal.
  • the subjects may include tumor patients.
  • the subject is in the elderly stage.
  • the sample may include a blood sample and/or a tissue or cell sample.
  • the sample may include whole blood, serum, plasma and/or cerebrospinal fluid.
  • the method may include: comparing the content of the QDLIR and/or the nucleic acid encoding QDLIR in a sample of the subject with a normal control value, wherein the normal control value may include a normal subject.
  • the content of the QDLIR and/or the nucleic acid encoding QDLIR in the subject may include: comparing the content of the QDLIR and/or the nucleic acid encoding QDLIR in the subject.
  • the normal subject may not suffer from the cognitive impairment.
  • the normal subject may not suffer from the neurodegenerative disease.
  • the QDLIR of the subject's sample, and/or the content of the nucleic acid encoding QDLIR is significantly higher than (for example, can be increased by at least about 20%, at least about 50%, 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 subject may be diagnosed as suffering from the cognitive impairment and/or the neurodegenerative disease.
  • the method may include: comparing the content of the QDLIR and/or the nucleic acid encoding QDLIR in a sample of the subject with an early control value, wherein the early control value may include the same The subject's previously measured content of said QDLIR and/or nucleic acid encoding QDLIR.
  • the subject has been diagnosed with the cognitive impairment and/or the neurodegenerative disease.
  • the subject has been diagnosed with the cognitive disorder; or the subject has been diagnosed with the neurodegenerative disease.
  • the QDLIR of the subject's sample, and/or the content of the nucleic acid encoding QDLIR is significantly higher than (for example, can be increased by at least about 20%, at least about 50%, 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 early control value, the subject is diagnosed with may be an exacerbation of the disease process of the cognitive impairment and/or the neurodegenerative disease.
  • the method may further comprise the step of detecting the content of markers related to cognitive impairment and/or neurodegenerative diseases in a sample derived from the subject.
  • the markers related to cognitive impairment and/or neurodegenerative diseases may include AD7C-NTP, pTau-181, pTau-217 and/or A ⁇ 1-42.
  • the detection results of the content of the marker can be used to assist in the diagnosis of cognitive disorders, and/or to evaluate the disease process of cognitive disorders, and/or to diagnose neurodegenerative diseases, and/or to evaluate the disease process of neurodegenerative diseases.
  • the method may further include the following steps: observing brain imaging of the subject.
  • the results of the brain imaging can be used to assist in the diagnosis of cognitive disorders, and/or to assess the disease process of cognitive disorders, and/or to diagnose neurodegenerative diseases, and/or to assess the disease process of neurodegenerative diseases.
  • the present application provides a system for diagnosing cognitive impairment, and/or assessing the disease process of cognitive impairment, which may include detecting intron retention splicing of DNA damage-inducible transcript 4-like transcript (DDIT4L).
  • the cleavage product is QDLIR, and/or a nucleic acid encoding said QDLIR.
  • the present application provides a system for diagnosing neurodegenerative diseases, and/or assessing the disease process of neurodegenerative diseases, which may comprise detecting the intron of DNA damage-inducible transcript 4-like transcript (DDIT4L) The retained cleavage product QDLIR, and/or the substance encoding the nucleic acid of said QDLIR.
  • DDIT4L DNA damage-inducible transcript 4-like transcript
  • the system may include a detection module capable of detecting the content value of the QDLIR and/or the nucleic acid encoding the QDLIR.
  • the detection module detects the content value of the QDLIR and/or the nucleic acid encoding the QDLIR in the sample obtained by the collection module.
  • the detection module may comprise a substance capable of detecting the content of the QDLIR and/or the nucleic acid encoding the QDLIR.
  • the substances may include antibodies capable of specifically binding QDLIR, probes capable of specifically binding QDLIR, primers capable of specifically amplifying nucleic acids encoding QDLIR, and gene chips capable of specifically detecting nucleic acids encoding QDLIR. , an aptamer capable of specifically binding to the nucleic acid encoding QDLIR, and/or a guide RNA capable of specifically binding to the nucleic acid encoding QDLIR.
  • the detection module may comprise an instrument capable of detecting the intronic retention cleavage product QDLIR of the DNA damage-inducible transcript 4-like transcript (DDIT4L), and/or the content of the nucleic acid encoding the QDLIR.
  • the instruments may include those required to perform qPCR, ELISA, western blot, and/or gene chips.
  • the detection module may comprise substances and/or instruments capable of detecting the levels of markers associated with cognitive impairment and/or neurodegenerative diseases.
  • the markers related to cognitive impairment and/or neurodegenerative diseases may include AD7C-NTP, pTau-181, pTau-217 and/or A ⁇ 1-42.
  • the substance may also include an antibody that can specifically bind to the marker, a probe that can specifically bind to the marker, a primer that can specifically amplify the nucleic acid encoding the marker, a primer that can specifically detect A gene chip encoding the nucleic acid encoding the marker, an aptamer capable of specifically binding to the nucleic acid encoding the marker, and/or a guide RNA capable of specifically binding the nucleic acid encoding the marker.
  • the instruments may include those required to perform qPCR, ELISA, western blot, and/or gene chips.
  • the detection module outputs a content value of the QDLIR and/or the nucleic acid encoding the QDLIR.
  • the system may include a judgment module, which can judge whether the subject is determined based on the content of the QDLIR and/or the nucleic acid encoding the QDLIR and the normal control value and/or early control value.
  • the before may include at least 10 days ago, at least 20 days ago, at least 1 month ago, at least 6 months ago, at least 12 months ago, at least 24 months ago, or longer.
  • the subject may have been diagnosed with the cognitive disorder and/or the neurodegenerative disease.
  • the QDLIR and/or the content of the nucleic acid encoding QDLIR in the subject can be tracked at different time stages, thereby helping to diagnose the cognitive impairment and/or the subject's cognitive impairment. or disease progression of said neurodegenerative diseases.
  • the normal subject may not suffer from the cognitive impairment and/or the neurodegenerative disease.
  • the determination module may diagnose the subject as suffering from the cognitive impairment and/or the neurodegenerative disease.
  • the judgment module may diagnose the subject as an exacerbation of the cognitive impairment and/or the neurodegenerative disease.
  • the judgment module can output the diagnosis result of the subject.
  • the system may include a display module, wherein the display module can display the diagnosis result of the judgment module.
  • the diagnostic results displayed by the display module may include qualitative results and/or quantitative results.
  • the system may include a storage module, wherein the storage module can store the content value output by the detection module, and/or the diagnosis result output by the judgment module.
  • the system may include the following devices:
  • system may further include (a') a device for collecting and/or receiving other clinical indicators or basic characteristic data of AD diagnosis and/or AD development process in the sample.
  • the device described in (b) is used to combine and analyze QDLIR level data with other clinical indicators or basic characteristic data of AD diagnosis and/or AD development process to determine the subject's AD diagnosis and/or Prognosis assessment of AD development process.
  • the system may further include equipment for analyzing other clinical indicators or basic characteristic data of AD diagnosis and/or AD development process; and/or comparing QDLIR level data with AD diagnosis and/or AD
  • a device that combines and analyzes other clinical indicators or basic characteristic data of the development process to evaluate the subject's AD diagnosis and/or AD development process.
  • the system may also include one or more devices selected from the following group: a device for inputting and/or outputting and/or storing data; a device for outputting and/or storing analysis results, For example, a device for uploading and/or storing analysis results to a cloud database or its corresponding cloud database device; a device for intelligently analyzing and recommending treatment plans based on the evaluation results.
  • a device for inputting and/or outputting and/or storing data For example, a device for uploading and/or storing analysis results to a cloud database or its corresponding cloud database device; a device for intelligently analyzing and recommending treatment plans based on the evaluation results.
  • the following methods can be used for AD diagnosis and/or AD progression progression (e.g., grading or staging) assessment: detecting the level of QDLIR molecules in a subject or a sample obtained from the subject, and comparing the level with a control level; If the comparison results show that the level of the QDLIR molecule in the subject is higher than the control level, it is suggested that the subject suffers from AD or develops AD.
  • the method of the present application optionally further includes: obtaining a test sample from the subject; contacting the test sample with a reagent or kit for detecting QDLIR levels.
  • control may include normal controls or controls for a specific developmental grade or stage of AD.
  • Control may refer to the level of a QDLIR molecule used as a reference, which includes, but is not limited to: a QDLIR molecule measured in a non-AD normal biological sample from the same subject (eg, a sample obtained from the subject who is not an AD patient or normal person) level, a statistically determined population norm level, or a standardized level.
  • control for a specific developmental grade or stage of AD refers to a QDLIR molecule level measured in the same subject at a previously determined developmental grade or stage, a statistically determined population standard level at that grade or stage, or a normalized s level.
  • the present application provides a drug screening method, which may include the following steps: detecting changes in the content of QDLIR and/or nucleic acids encoding QDLIR in a subject after administration of a candidate drug, wherein the drug is used for prevention , treatment and/or alleviation of cognitive impairment and/or neurodegenerative diseases.
  • a drug candidate has a therapeutic effect when the content of QDLIR and/or nucleic acid encoding QDLIR is reduced in a subject when the drug candidate is administered.
  • said administration may include injection.
  • the drugs may include small molecule drugs and/or biomacromolecule drugs.
  • the level of QDLIR is closely related to the presence and/or degree of cognitive impairment, and thus can be used as an evaluation index for AD diagnosis and/or AD development process (such as grading or staging).
  • the present application also provides a method of screening candidate drugs for treating AD, the method comprising testing the effect of the candidate drug on QDLIR levels in a subject or a sample obtained from the subject, wherein, after using the candidate drug , the decrease in QDLIR levels indicates that the drug candidate has the effect of treating AD.
  • Each feature involved in the candidate drug screening method of the present application may be defined or described in the present application.
  • the drug candidate may be an inhibitor of QDLIR.
  • QDLIR is specifically expressed in blood samples and/or cerebrospinal fluid of AD patients.
  • QDLIR can be detected in a small amount of serum (such as about 100 ⁇ l), which provides a promising detection method for early diagnosis of AD for clinical application.
  • serum such as about 100 ⁇ l
  • Current results suggest that the positive detection rates for diseases such as AD and MCI are above 90%, and QDLIR has great application prospects as a specific marker for early screening of AD.
  • pTau-181, pTau-217, and A ⁇ 1-42 currently used in clinical blood tests are newly developed early screening tests for AD. They have relatively low specificity and are relatively expensive.
  • the AD early screening test based on QDLIR levels in this application has a high detection rate, very good specificity, and/or low cost, and has a price advantage, such as the detection of QDLIR by ELISA.
  • the cost can be controlled at less than 10 yuan/person. Therefore, the detection indicators and methods of the present application have greater effects and cost advantages compared with existing methods.
  • QDLIR is very specific and sensitive for the diagnosis of cognitive impairment and/or neurodegenerative diseases, and can detect pTau-181, pTau-217, A ⁇ 1-42 and other target targets without significant differences. Differentiation of potential AD and/or MCI patients can lead to effective diagnosis of patients at an earlier stage.
  • a substance for detecting the intron retained cleavage product QDLIR of DNA damage-induced transcript 4-like transcript (DDIT4L) in a subject sample is prepared for use in the diagnosis of Alzheimer's disease, mild cognitive impairment diagnosis, and / Or application in products (such as kits or detection systems) for assessing the progression of Alzheimer's disease (such as grading or staging).
  • the shearing product is selected from the group consisting of: mRNA having the nucleotide sequence shown in SEQ ID NO: 2; having the nucleotide sequence shown in SEQ ID NO: 3 The nucleic acid encoding the splicing product peptide; or, the splicing product peptide having the amino acid sequence shown in SEQ ID NO:4.
  • Alzheimer's disease includes: early Alzheimer's disease (amnesia stage), middle Alzheimer's disease (nervous disorder stage) and late Alzheimer's disease (severe dementia stage).
  • the substance is selected from: substances specific to the QDLIR, such as antibodies (preferably monoclonal antibodies) to the QDLIR; QDLIR-specific probes, genes Chips, PCR primers, gRNA, etc.
  • sample is selected from: blood samples, such as whole blood, serum, plasma; cerebrospinal fluid.
  • a product (such as a kit or system) for the diagnosis of Alzheimer's disease, the diagnosis of mild cognitive impairment and/or the assessment of the development process (such as grading or staging) of Alzheimer's disease, which contains:
  • the substance in (i) is one or more selected from the following group:
  • Substances for detecting said QDLIR at the gene level and/or protein level such as substances used in one or more detection techniques or methods selected from the following group: immunohistochemistry (e.g. immunofluorescence analysis, chemical Luminescence method, reverse enzyme-linked immunosorbent assay, immunocolloidal gold method), Western blotting method, Northern blotting method, PCR, biochip method; or
  • Substances specific to the QDLIR such as antibodies (preferably monoclonal antibodies); cleavage product-specific probes, gene chips, PCR primers, gRNA; and/or
  • substances in (ii) are selected from: substances used to detect one or more other Alzheimer's disease markers, such as: AD7C-NTP, pTau-181, pTau-217, A ⁇ 1-42; and/or, with Reagents for imaging brain amyloid; and/or reagents for imaging diagnosis of a subject's brain.
  • a system for diagnosing Alzheimer's disease, diagnosing mild cognitive impairment, and/or assessing the progression (such as grading or staging) of Alzheimer's disease in a subject which includes the following device:
  • the level of the QDLIR in the sample is increased, indicating that the subject suffers from Alzheimer's disease or mild cognitive impairment; or, compared with the mild cognitive impairment control or with the diagnosed
  • An increase in the level of QDLIR in the sample compared to a previous control level for an Alzheimer's disease subject indicates that the subject's Alzheimer's disease has progressed (e.g., to a higher grade or stage).
  • a method of screening drugs for treating Alzheimer's disease comprising: detecting the effect of the candidate drug on the QDLIR level of the intronic retention cleavage product of DNA damage-inducible transcript 4-like transcript (DDIT4L) in the subject, wherein, after using the candidate drug, a decrease in the level indicates that the candidate drug has a therapeutic effect on Alzheimer's disease.
  • DIT4L DNA damage-inducible transcript 4-like transcript
  • Example 1 QDLIR protein is a secreted protein
  • Plasmid construction and cell transfection method The cDNA of human QDLIR was cloned into pCMV-flag vector to express flag-QDLIR protein.
  • HEK293 cells Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences
  • pCMV-flag plasmid and pCMV-flag-QDLIR plasmid using Lippo3000 reagent. After culturing the cells for 48 hours, harvest the cells and culture medium, add an appropriate amount of lysis solution to each, and rotate at 4°C for lysis. 30 minutes, centrifuge at 12000g for 30 minutes, take the supernatant, and take a small amount of lysate for Western Blot analysis.
  • Tissue samples AD and MCI patients, non-AD patients (hypertension, coronary heart disease or diabetes) and normal venous blood samples (58 samples in total), from Huashan Hospital affiliated to Fudan University and Xuhui District Central Hospital.
  • Methods for collecting venous blood and obtaining plasma Use vacuum blood collection technology to collect the patient's cubital fossa vein in an EDTA vacuum tube, and immediately invert the test tube gently 8 times to fully mix the blood and anticoagulant. Then transport it to the laboratory on ice, centrifuge at 1500g for 10 minutes at 4°C, aspirate the supernatant, aliquot, and store at -80°C.
  • Plasma Western Blot method aliquot the plasma and thaw it on ice, dilute it appropriately with PBS, add loading buffer and denature it in a 100°C metal bath for 5 minutes. The subsequent steps are the same as the tissue Western Blot method.
  • Plasma ELISA method The plasma sample was diluted 1:1 with 1 ⁇ diluent (OBT1998G, purchased from Bio-Rad); the pipette took 100 ⁇ l blocking solution and was blocked at 37°C for 1 hour; the pipette took 100 ⁇ l #1 ⁇ 8 Add the standard substance and diluted plasma in the centrifuge tube (double dilution) to the wells of the plate coated with capture antibody (Gill Biochemical, AB009458). Change the pipette tip every time you take a sample; incubate at 37°C for 1.5 hours; remove each well.
  • OBT1998G purchased from Bio-Rad
  • wash the liquid in each well three times with 100 ⁇ l 1 ⁇ washing solution (BUF031C, purchased from Bio-Rad); add 100 ⁇ l detection antibody (Gill Biochemical, AB008349) to each well, and incubate at 37°C for 1 hour; remove the liquid in each well.
  • Wash 3 times with 100 ⁇ l 1 ⁇ washing solution add 100 ⁇ l 1 ⁇ avidin-HRP solution to each well, and incubate at 37°C for 1 hour; remove the liquid in each well, and wash 3 times with 100 ⁇ l 1 ⁇ washing solution; add 100 ⁇ l 1 ⁇ avidin-HRP solution to each well.
  • QDLIR has high concentrations in the blood of AD and MCI patients and can be used as a blood marker to detect the presence of AD, especially for patients with mild cognitive impairment such as MCI.
  • QDLIR content is related to the severity of AD (see Figure 2b).
  • QDLIR protein (Gill Biochemical, 752084), use DPBS solution to prepare a 1 mg/ml stock solution, and dilute it with ELISA washing solution (BUF031C, purchased from Bio-Rad) before the experiment to prepare a standard QDLIR protein solution.
  • the ELISA method was used to detect the concentration of the standard QDLIR protein, and polynomial regression was used to fit it. The resulting standard curve is shown in Figure 3.
  • the R 2 value of this regression model is 0.99, indicating a high degree of fit; the shaded part shows the 95% confidence interval of the regression model, and the long and narrow confidence interval can be seen in the figure, indicating the credibility of estimating the QDLIR content in the sample based on this model. high.
  • Tissue samples venous blood samples from patients with different types of MCI and normal follow-up humans, from Huashan Hospital affiliated to Fudan University.
  • Plasma ELISA method same as Example 2.
  • MCI classification method aMCI-s: amnestic MCI with impairment of single cognitive domain; aMCI-m: amnestic MCI with impairment of multiple cognitive domains; naMCI-s: non-amnestic MCI with impairment of single cognitive domain; naMCI-m : Non-amnestic MCI is impaired in multiple cognitive domains.
  • Example 5 there were 122 samples from aMCI-m type patients; 85 samples from aMCI-s type patients; 11 samples from naMCI-m type patients; 88 samples from naMCI-s type patients; and 279 samples from normal follow-up people.
  • Tissue samples venous blood samples from different types of MCI patients, AD patients and normal follow-up humans, from Huashan Hospital affiliated to Fudan University.
  • Plasma ELISA method QDLIR is the same as in Example 2; Tau, A ⁇ 40 and A ⁇ 42 are detected by the triple kit (Product No.: 101995) of Hangzhou Jingbai Biotechnology Co., Ltd. according to its operating procedures.
  • Tissue samples venous blood samples and cerebrospinal fluid samples from the same patient (one case each of cerebral edema, meningioma and glioma), both from Huashan Hospital affiliated to Fudan University.
  • Cerebrospinal fluid collection method Collect cerebrospinal fluid from the patient during surgery.
  • Plasma and cerebrospinal fluid ELISA method same as Example 2.
  • the QDLIR content in the patient's blood is consistent with the content in the cerebrospinal fluid, indicating that the content detected in the blood can directly reflect the QDLIR content in the cerebrospinal fluid, suggesting that the QDLIR described in this application is also suitable for use as a valid sample using cerebrospinal fluid. diagnostic markers.

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Abstract

本申请涉及作为早老性痴呆诊断标志物的DDIT4L剪切产物，及其在早老性痴呆的诊断中的应用。具体而言，本申请涉及检测来自对象样品中编码DNA损伤诱导性转录物4样转录物（DDIT4L）的内含子滞留剪切产物QDLIR的物质在制备用于所述对象早老性痴呆诊断、轻度认知障碍诊断和/或认知障碍发展进程（例如分级或分期）评估的产品（例如试剂盒或检测系统）中的应用，及其相关产品以及利用QDLIR来筛选药物的方法。

Description

作为早老性痴呆诊断标志物的DDIT4L剪切产物 技术领域
本申请涉及生物医药领域，具体的涉及DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR作为早老性痴呆或轻度认知障碍的血液标志物的应用及其相应的产品和方法。
背景技术
作为危害人类大脑的疾病中发病率最高的神经退行性疾病，早老性痴呆，又称阿尔茨海默病(Alzheimer Disease,AD)给患者本身造成了极大的痛苦，同时也给家庭和社会带来了沉重的负担。
据国际阿尔兹海默病协会(ADI)的“世界阿尔茨海默病2017年报告”，全球痴呆病患者照料费用已从2010年的6,040亿美元增加到了2017年的10，000亿美元，增长率为65.6％。其中，美国AD患者500万，一年需花费照料者180亿小时照料患者。2017年全球AD患者为5000万，随着世界老龄化人口的增加，这一数字仍将不断增加。在目前尚没有能够治疗、延缓或阻止该病进展的药物的前提下，预计患者数量将以每20年递增1倍的速度增加，2030年将达到8200万，2050年将达1.5亿。
根据中国发展基金会(国务院发展研究中心发起成立的全国性社团组织)发布的《中国发展报告2020：中国人口老龄化的发展趋势和政策》，2020年中国65岁及以上的老年人约有1.8亿，约占总人口的13％；2025年“十四五”规划完成时，65岁及以上的老年人将超过2.1亿，占总人口数的约15％；2035年和2050年时，中国65岁及以上的老年人将达到3.1亿和接近3.8亿，占总人口比例则分别达到22.3％和27.9％。
AD的发病率与年龄相关，在整个人群中的发病率为2％～4％。尤其对于年龄在65岁以上的中老年者，其患病率为5％，且随着年龄增长，患病率呈倍数增长。据分析，85岁以上的老人，患病率可增加至25％，而95岁以上，则患病率高达60％。我国目前已逐步进入老龄化社会，据统计，我国AD患者已近千万，且每年增加30万以上的新发病例。
AD主要症状为认知功能损害。认知功能损害归根到底是突触和神经元损伤、神经环路结构和功能异常所致。目前AD新药临床研究连续失败的主要原因是突触和神经元损伤的机制仍然不够明确、缺乏接近人类疾病病理生理特征(如神经退行性变等)的细胞/动物模型，并 由此导致AD有效防治方法和能反映疾病转归的早期诊断标志物研究进展缓慢。
代谢异常可能是AD发病的主要机制之一。在AD病人中，胆固醇等代表的脂代谢异常和葡萄糖等代表的糖代谢异常是最为常见的代谢异常，还包括脑小血管的退行性变化为代表的对大脑供血不足造成的脑缺血缺氧的能量代谢紊乱。但是大脑脂代谢和糖代谢异常是如何影响AD的发生和发展过程，其机制并不清楚。线粒体是主要能量物质代谢的关键细胞器，线粒体功能异常会导致能量代谢异常，同样损伤神经元及脑小血管内皮细胞的功能，但是在早期AD发生发展中，线粒体功能异常与脂代谢和糖代谢异常的关系并不清楚。
轻度认知功能障碍(Mild cognitive impairment，MCI)是一种涉及客观认知能力下降的综合征，是介于轻度痴呆和健康衰老之间的一种状态，以记忆障碍为突出表现，可有其他认知功能某种程度的减退，但日常活动能力正常，不符合痴呆诊断标准。每年有10％～15％的MCI转化为AD，是正常对照的10倍。作为正常老化向痴呆的过渡状态，MCI越来越受到人们的关注，研究MCI是AD早期诊断和早期干预的重要环节。
目前的AD诊断标志物主要包括脑脊液Aβ和Tau含量检测、PET和MRI神经影像学手段等。这些方法虽然具有较高、甚至很高的诊断效价，但也存在操作困难、昂贵、损伤和疾病转归关联性不强等缺点。由此，开发出新型简便易行、性价比高、适合大规模人群筛查且能较好地用于AD早期诊断标志物的需求十分迫切。
发明内容
本申请提供了一种新型早老性痴呆AD或轻度认知障碍MCI的血液标志物。
一方面，本申请提供了一种DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸，在疾病诊断和/或疾病进程评估中的用途，其中所述疾病包括认知障碍。
一方面，本申请提供了一种DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸，在疾病诊断和/或疾病进程评估中的用途，其中所述疾病包括神经退行性疾病。
在某些实施方式中，所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
在某些实施方式中，所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
在某些实施方式中，所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
在某些实施方式中，所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知障碍。
在某些实施方式中，所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
在某些实施方式中，所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
另一方面，本申请提供一种检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质，在制备用于疾病诊断和/或疾病进程评估的产品中的用途，其中所述疾病包括认知障碍。
另一方面，本申请提供一种检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质，在制备用于疾病诊断和/或疾病进程评估的产品中的用途，其中所述疾病包括神经退行性疾病。
另一方面，本申请提供一种诊断试剂盒，其包含本申请所述的能够检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质，所述诊断盒用于诊断认知障碍，和/或评估认知障碍的疾病进程；和/或用于诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程。
在某些实施方式中，所述诊断试剂盒包含说明书，所述说明书记载了利用所述诊断试剂盒诊断认知障碍，和/或评估认知障碍的疾病进程；和/或诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程的方法。
在某些实施方式中，所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
在某些实施方式中，所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
在某些实施方式中，所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
在某些实施方式中，所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知障碍。
在某些实施方式中，所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
在某些实施方式中，所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
在某些实施方式中，所述物质包括能够检测所述QDLIR，和/或编码所述QDLIR的核酸的试剂和/或设备。
在某些实施方式中，所述物质包括特异性检测所述QDLIR，和/或编码所述QDLIR的核酸的试剂。
在某些实施方式中，所述物质包括能够特异性结合QDLIR的抗体、能够特异性结合QDLIR的探针、能够特异性扩增编码QDLIR的核酸的引物、能够特异性检测编码QDLIR的核酸的基因芯片、能够特异性结合编码QDLIR的核酸的适体、和/或能够特异性结合编码QDLIR的核酸的向导RNA。
在某些实施方式中，所述产品包括诊断试剂盒。
另一方面，本申请提供了一种诊断认知障碍，和/或评估认知障碍的疾病进程的方法，其包括以下步骤：检测源自有需要的受试者的样本中，DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的含量。
另一方面，本申请提供了一种诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程的方法，其包括以下步骤：检测源自有需要的受试者的样本中，DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的含量。
在某些实施方式中，所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
在某些实施方式中，所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
在某些实施方式中，所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
在某些实施方式中，所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知障碍。
在某些实施方式中，所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
在某些实施方式中，所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
在某些实施方式中，所述受试者包括哺乳动物。
在某些实施方式中，所述受试者包括肿瘤患者。
在某些实施方式中，所述受试者处于老龄阶段。
在某些实施方式中，所述样本包括血液样品和/或组织或细胞样品。
在某些实施方式中，所述样本包括全血、血清、血浆和/或脑脊液。
在某些实施方式中，所述方法包括：将所述受试者的样本的所述QDLIR和/或编码QDLIR的核酸的含量与正常对照值进行比较，其中所述正常对照值包括正常的受试者中所述QDLIR 和/或编码QDLIR的核酸的含量。
在某些实施方式中，所述正常的受试者不患有所述认知障碍和/或所述神经退行性疾病。
在某些实施方式中，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于所述正常对照值，所述受试者被诊断为患有所述认知障碍和/或所述神经退行性疾病。
在某些实施方式中，所述方法包括：将所述受试者的样本的所述QDLIR和/或编码QDLIR的核酸的含量与早期对照值进行比较，其中所述早期对照值包括同一所述受试者之前测得的所述QDLIR和/或编码QDLIR的核酸的含量。
在某些实施方式中，所述受试者已确诊患有所述认知障碍和/或所述神经退行性疾病。
在某些实施方式中，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于所述早期对照值，所述受试者被诊断为所述认知障碍和/或所述神经退行性疾病的疾病进程加剧。
在某些实施方式中，所述的方法还包括以下的步骤：检测源自所述受试者的样本中，与认知障碍和/或神经退行性疾病相关的标志物的含量。
在某些实施方式中，所述与认知障碍和/或神经退行性疾病相关的标志物包括AD7C-NTP、pTau-181、pTau-217和/或Aβ1-42。
在某些实施方式中，所述的方法还包括以下的步骤：观察所述受试者的脑部成像。
另一方面，本申请提供一种诊断认知障碍，和/或评估认知障碍的疾病进程的系统，其包含检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质。
另一方面，本申请提供一种诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程的系统，其包含检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质。
在某些实施方式中，所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
在某些实施方式中，所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
在某些实施方式中，所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
在某些实施方式中，所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知 障碍。
在某些实施方式中，所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
在某些实施方式中，所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
在某些实施方式中，所述的系统以试剂盒的形式存在。
在某些实施方式中，所述的系统包含收集模块，所述收集模块能够收集源自有需要的受试者的样本。
在某些实施方式中，所述受试者包括哺乳动物。
在某些实施方式中，所述受试者包括肿瘤患者。
在某些实施方式中，所述受试者处于老龄阶段。
在某些实施方式中，所述样本包括血液样品和/或组织或细胞样品。
在某些实施方式中，所述样本包括全血、血清、血浆和/或脑脊液。
在某些实施方式中，所述的系统包含检测模块，所述检测模块能够检测所述QDLIR和/或编码所述QDLIR的核酸的含量值。
在某些实施方式中，所述检测模块检测所述收集模块获得的所述样本中所述QDLIR和/或编码所述QDLIR的核酸的含量值。
在某些实施方式中，所述检测模块包含能够检测所述QDLIR和/或编码所述QDLIR的核酸的含量的物质。
在某些实施方式中，所述物质包括能够特异性结合QDLIR的抗体、能够特异性结合QDLIR的探针、能够特异性扩增编码QDLIR的核酸的引物、能够特异性检测编码QDLIR的核酸的基因芯片、能够特异性结合编码QDLIR的核酸的适体、和/或能够特异性结合编码QDLIR的核酸的向导RNA。
在某些实施方式中，所述检测模块包含能够检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的含量的仪器。
在某些实施方式中，所述检测模块包含能够检测与认知障碍和/或神经退行性疾病相关的标志物的含量的物质和/或仪器。
在某些实施方式中，所述与认知障碍和/或神经退行性疾病相关的标志物包括AD7C-NTP、pTau-181、pTau-217和/或Aβ1-42。
在某些实施方式中，所述检测模块输出所述QDLIR和/或编码所述QDLIR的核酸的含量值。
在某些实施方式中，所述的系统包含判断模块，所述判断模块能够根据所述QDLIR和/或编码所述QDLIR的核酸的含量与正常对照值和/或早期对照值，判断所述受试者的诊断结果和/或疾病进程结果；其中，所述正常对照值包括正常的受试者中所述QDLIR和/或编码QDLIR的核酸的含量；其中，所述早期对照值包括同一所述受试者之前测得的所述QDLIR和/或编码QDLIR的核酸的含量。
在某些实施方式中，所述正常的受试者不患有所述认知障碍和/或所述神经退行性疾病。
在某些实施方式中，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于所述正常对照值，所述判断模块将所述受试者诊断为患有所述认知障碍和/或所述神经退行性疾病。
在某些实施方式中，所述受试者已确诊患有所述认知障碍和/或所述神经退行性疾病。
在某些实施方式中，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于所述早期对照值，所述判断模块将所述受试者诊断为所述认知障碍和/或所述神经退行性疾病的疾病进程加剧。
在某些实施方式中，所述判断模块输出所述受试者的诊断结果。
在某些实施方式中，所述的系统包括显示模块，其中所述显示模块能够显示出所述判断模块的诊断结果。
在某些实施方式中，所述的系统包括存储模块，其中所述存储模块能够存储所述检测模块输出的所述含量值，和/或所述判断模块输出的诊断结果。
另一方面，本申请提供一种药物筛选方法，其包括以下的步骤：检测施用候选药物后，受试者中QDLIR和/或编码QDLIR的核酸的含量的变化，其中所述药物用于预防、治疗和/或缓解认知障碍和/或神经退行性疾病。
在某些实施方式中，所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
在某些实施方式中，所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
在某些实施方式中，所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
在某些实施方式中，所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
在某些实施方式中，所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知障碍。
在某些实施方式中，所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
在某些实施方式中，所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
在某些实施方式中，所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆。
在某些实施方式中，所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
在某些实施方式中，当施用所述候选药物中，受试者中QDLIR和/或编码QDLIR的核酸的含量降低，则所述候选药物具有治疗效果。
在某些实施方式中，所述施用包括注射。
在某些实施方式中，所述药物包括小分子药物和/或生物大分子药物。
在本申请的第一方面中，提供了检测来自对象样品中编码DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR的物质在制备用于所述对象早老性痴呆诊断、轻度认知障碍诊断和/或早老性痴呆发展进程(例如分级或分期)评估的产品(例如试剂盒)中的应用。
在一些实施方式中，所述对象选自：人、非人灵长类动物、啮齿类动物(如大鼠、小鼠、豚鼠)、犬、猫、马、牛、羊。
在一些实施方式中，所述对象未罹患癌症。
在一些实施方式中，所示对象未罹患恶性肿瘤。
在一些实施方式中，所述QDLIR具有选自下组的序列：具有SEQ ID NO:2的核苷酸序列(mRNA)；具有SEQ ID NO:3所示核苷酸序列的与QDLIR肽对应的编码核苷酸分子；或者，具有SEQ ID NO:4所示氨基酸序列的QDLIR肽；以上序列经一个或多个核苷酸和/或氨基酸取代、缺失和/或添加获得的变体；与以上序列具有高度同源性(例如高于80％、85％、90％、95％、96％、97％、98％、99％、99.5％序列同源性)的序列。
在一些实施方式中，所述早老性痴呆包含早期(健忘期)、中期(神经紊乱期)和晚期(严重痴呆期)。
在一些实施方式中，所述物质选自用于检测QDLIR的试剂、设备或其组合。
在一些实施方式中，所述物质是用于在基因水平和/或蛋白质水平上检测所述QDLIR的物质。例如，所述物质是用于选自下组的一种或多种检测技术或方法中的物质：免疫组织化学法(如免疫荧光分析、化学发光法、反向酶联免疫吸附、免疫胶体金法)、Western印迹法、Northern印迹法、PCR、生物芯片法、蛋白质芯片法。
在一些实施方式中，物质选自：对所述QDLIR具有特异性的物质，例如所述QDLIR的抗体(例如单克隆抗体)；剪切产物特异性的探针、基因芯片、PCR引物、gRNA等。
在一些实施方式中，所述样品选自：血液样品，如全血、血清、血浆；组织或细胞样品；脑脊液。
在一些实施方式中，所述血液样品是新鲜血液或冻存血。
在一些实施方式中，与正常对照值相比，所述样品中所述QDLIR的水平提高，表明所述对象罹患早老性痴呆或轻度认知障碍。
在一些实施方式中，与轻度认知障碍对照相比或与已诊断为早老性痴呆对象的前期对照水平相比，所述样品中QDLIR的水平提高，表明所述对象早老性痴呆有所发展(例如分级或分期更高)。
在一些实施方式中，与低分级或低分期时的对照值相比，所述样品中所述QDLIR的水平提高，表明所述对象早老性痴呆有所发展(例如分级或分期更高)。
另一方面，本申请提供一种诊断试剂盒，其包含本申请所述的能够检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质。
例如，本申请提供了一种用于早老性痴呆诊断、轻度认知障碍诊断和/或早老性痴呆发展进程(例如分级或分期)评估的产品(例如试剂盒)，其包含：
(i)用于检测对象样品中QDLIR的物质；以及
(ii)可任选的，用于早老性痴呆诊断、轻度认知障碍诊断和/或早老性痴呆发展进程(例如分级或分期)评估的其他物质，例如用于检测现有早老性痴呆标志物的检测物质。
在一些实施方式中，(i)中的物质为选自下组中的一种或多种：
用于检测所述QDLIR的试剂、设备或其组合；
用于在基因水平和/或蛋白质水平上检测所述QDLIR的物质，如用于选自下组的一种或多种检测技术或方法中的物质：免疫组织化学法(如免疫荧光分析、化学发光法、反向酶联免疫吸附、免疫胶体金法)、Western印迹法、Northern印迹法、PCR、生物芯片法；或者
对所述QDLIR具有特异性的物质，例如其抗体(优选单克隆抗体)；剪切产物特异性的探针、基因芯片、PCR引物、gRNA。
在一些实施方式中，(ii)中的其他物质选自：用于检测一种或多种其他早老性痴呆标志物的物质，例如：AD7C-NTP、pTau-181、pTau-217、Aβ1-42；和/或，用于大脑淀粉样蛋白成像的试剂；和/或，用于对象脑部成像诊断的试剂。
在本申请的一些方面中，提供了一种筛选早老性痴呆治疗药物的方法，所述方法包括：检测候选药物对对象中QDLIR水平的影响，其中，在使用所述候选药物后，所述水平降低则表明所述候选药物具有早老性痴呆治疗效果。
在一些实施方式中，所述检测采用如本申请所述的产品进行。
在本申请的一些方面中，提供了用于早老性痴呆诊断、轻度认知障碍诊断和/或认知障碍发展进程评估的方法，所述方法包括：检测来自所述对象的样品中QDLIR的水平，并据此对该对象进行早老性痴呆诊断和/或早老性痴呆发展进程(例如分级或分期)评估。
在一些实施方式中，所述检测采用如本申请所述的产品进行。
在一些实施方式中，与对照值相比，所述样品中所述QDLIR的水平提高，表明所述对象罹患早老性痴呆；或者，与低分级或低分期对照相比，所述样品中所述QDLIR的水平提高，表明所述对象早老性痴呆有所发展(例如分级或分期更高)。
在本申请的一些方面中还提供了一种与本申请方法对应的系统，其包括如下装置：
(a)采集和/或接收样品中QDLIR水平数据的装置；
(b)对数据进行分析以对所述对象的早老性痴呆进行诊断、轻度认知障碍进行诊断和/或早老性痴呆发展进程(例如分级或分期)进行评估的装置，其中，与正常对照值相比，所述样品中所述QDLIR的水平提高，表明所述对象罹患早老性痴呆或轻度认知障碍；或者，与轻度认知障碍对照相比或与已诊断为早老性痴呆对象的前期对照水平相比，所述样品中 QDLIR的水平提高，表明所述对象早老性痴呆有所发展(例如分级或分期更高)。
在一些实施方式中，所述系统还进一步包括(a')采集和/或接收样品中早老性痴呆诊断和/或早老性痴呆发展进程的其他临床指标或基本特征数据的装置。
在一些实施方式中，(b)中所述的装置用于将QDLIR水平数据与早老性痴呆诊断和/或早老性痴呆发展进程的其他临床指标或基本特征数据进行结合分析以对所述对象的早老性痴呆诊断和/或早老性痴呆发展进程进行预后评估。
或者，在一些实施方式中，所述系统还进一步包括对早老性痴呆诊断和/或早老性痴呆发展进程的其他临床指标或基本特征数据进行分析的设备；和/或将QDLIR水平数据与早老性痴呆诊断和/或早老性痴呆发展进程的其他临床指标或基本特征数据进行结合分析以对所述对象的早老性痴呆诊断和/或早老性痴呆发展进程进行评估的装置。
在一些实施方式中，所述系统还包括选自下组的一种或多种装置：用于输入和/或输出和/或存储数据的装置；用于输出和/或存储分析结果的装置，例如用于将分析结果上传和/或存储到云数据库的装置或其相应的云数据库装置；根据预后评估结果智能分析和推荐治疗方案的装置。
应理解，本领域技术人员可根据需要在系统中添加其他常规装置或程序。
本领域技术人员能够从下文的详细描述中容易地洞察到本申请的其它方面和优势。下文的详细描述中仅显示和描述了本申请的示例性实施方式。如本领域技术人员将认识到的，本申请的内容使得本领域技术人员能够对所公开的具体实施方式进行改动而不脱离本申请所涉及发明的精神和范围。相应地，本申请的附图和说明书中的描述仅仅是示例性的，而非为限制性的。
附图说明
本申请所涉及的发明的具体特征如所附权利要求书所显示。通过参考下文中详细描述的示例性实施方式和附图能够更好地理解本申请所涉及发明的特点和优势。对附图简要说明书如下：
图1a-1b显示的是DDIT4L mRNA前体的剪接方式及QDLIR蛋白从转染细胞分泌到培养基中的情况。其中：
a图：DDIT4L的剪接方式，其中通过正常剪切去除内含子获得mRNA的长度为582nt(SEQ ID NO:5)，通过异常的内含子滞留(Intron retention)剪接方式获得的mRNA中参与编码QDLIR的RNA长度为255nt(SEQ ID NO:3)；
b图：QDLIR从细胞中分泌到培养基中，其中将pCMV-flag-QDLIR质粒转染到HEK293细胞中，培养48小时后发现培养基中可以检测到分泌的QDLIR蛋白。裂解液QDLIR质粒泳道中可见QDLIR蛋白条带，其大小为12kDa。
图2a-2b显示的是QDLIR蛋白在AD病人以及多种其他疾病病人血液中含量的变化。
a图：通过Western检测QDLIR在AD病人以及多种其他疾病病人血液中含量的变化；
图中血浆蛋白(plasma protein)是作为内参的免疫球蛋白的轻链。
b图：通过ELISA检测QDLIR在AD病人、MCI病人以及非AD/MCI病人的血液中含量的变化。图中***表示P<0.001，*表示P<0.05。
图3显示的是标准品QDLIR蛋白的ELISA法检测标准曲线。
图4显示的是临床样品ELISA检测的R℃曲线。
图5显示的是QDLIR在不同类别MCI病人血液中的含量变化。
图6显示的是QDLIR及其他指标在不同类别MCI和AD病人血液中的含量变化。
图7显示的是QDLIR在血浆和脑脊液中的表达量的相关性。
具体实施方式
以下由特定的具体实施例说明本申请发明的实施方式，熟悉此技术的人士可由本说明书所公开的内容容易地了解本申请发明的其他优点及效果。
术语定义
术语“DDIT4L”通常是指DNA damage-inducible transcript 4-like，DNA损伤诱导转录本4样。其也可以称为REDD2/RTP801L。研究发现，DDIT4L可以与心脏功能障碍相关。也可以用于治疗胶质瘤。人DDIT4L基因在GenBank的登录号为115265；人DDIT4L蛋白在GenBank的登录号为NP_660287.1。
术语“QDLIR”通常是指DDIT4L的内含子滞留剪切产物。在本申请中，所述QDLIR的氨基酸序列可以如SEQ ID NO:4所示。
术语“表达水平”通常是指特定相关基因的蛋白质、RNA或mRNA水平。可以采用本领域中已知的任何方法来测定特定相关基因(例如人ATP6V1B2基因)的表达水平。在本申请中，所述“表达”通常是指将基因编码的信息转变成存在于细胞中并在细胞中操作的结构的过程。例如可以包括逆转录和扩增分析(例如PCR、连接RT-PCR或定量RT-PCT)、杂交分析、RNA印迹法(Northern blotting)、斑点印迹法、原位杂交、胶凝电泳、毛细管电泳、柱色谱、蛋白 质印迹法、免疫组织化学、免疫染色或质谱。可以直接对生物样品或对从样品中分离出来的蛋白质/核酸执行分析。
术语“神经退行性疾病”通常是指由神经元结构和功能逐渐丧失，包括神经元死亡和胶质细胞平衡，所导致的痴呆等认知障碍。在某些情况下，年龄(例如阿尔茨海默病(AD)、帕金森病(PD))或影响CNS细胞功能的基因突变(例如亨廷顿氏舞蹈病、早发性AD或PD、肌萎缩性脊髓侧索硬化症(ALS))可以引起所述神经退行性疾病。所述神经退行性疾病可以具有选自以下的变化和/或病症：蛋白错误折叠和聚集；神经炎症(例如在毒刺激(如蛋白聚集)、感染、创伤性损伤或自身免疫等信号刺激下出现的CNS炎症)；细胞信号转导的改变；获得性衰老/细胞死亡(例如被中断的凋亡信号转导、线粒体功能障碍、自噬受损以及坏死小体被应激/炎症激活)；运动细胞受损以及表观遗传学变化。
术语“阿尔兹海默症”通常是指早老性痴呆、老年痴呆，是一种发病进程缓慢、随着时间不断恶化的神经退化性疾病。最常见的早期症状为丧失短期记忆(难以记住最近发生的事)，当疾病逐渐进展，可能逐渐出现下述症状中的至少一种：语言障碍、定向障碍(例如，容易迷路)、情绪不稳、丧失动机、无法自理和行为问题。阿尔茨海默病的真正成因至今仍然不明，其进程可能与大脑中的纤维状类淀粉蛋白质斑块沉积和Tau蛋白有关。目前并没有可以阻止或逆转病程的治疗，只有少数方法或许可以暂时缓解或改善症状。
术语“早老性痴呆”在本申请中可以与术语“阿尔兹海默症”互换使用。所述早老性痴呆可以包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。例如，所述早老性痴呆早期患者的学习与记忆障碍会愈见明显，在某些情况下，会具有语言障碍、执行障碍、认识障碍(失认症)和/或技能执行障碍(失用症)。例如，所述早老性痴呆中期患者将失去独立生活的能力，并且可能无法进行大部分的日常活动(在某些情况下，可以患有命名不能症、言语错乱症、和/或病觉缺失症)。例如，所述早老性痴呆晚期患者可能会晚期依赖照护者。例如，可能完全失去了语言能力。例如，可能无法自行进食。
术语“认知障碍”通常是指被认为或确实牵涉于神经元结构和/或功能的进行性损失(包括神经元死亡)或与上述有关的疾病和病症。例如，所述认知障碍的特征可以包括认知(例如，记忆、注意力、感知和/或思维)的损伤。这些障碍可以包括病原体-诱导的认知功能障碍，例如HIV相关的认知功能障碍和莱姆病相关的认知功能障碍。认知障碍的实例可以包括阿尔茨海默病、亨廷顿病、帕金森病、肌萎缩侧索硬化(ALS)、孤独症、早期认知功能损害(MCI)、中风、创伤性脑损伤(TBI)和/或与年龄相关的记忆损害(AAMI)。
术语“早期认知障碍(MCI)”通常是指一种介于正常认知与认知障碍的中间临床状态。在 某些情况下，所述MCI可以包括满足痴呆标准但程度超过正常衰老的认知损害。MCI在临床表现、病因、预后和患病率方面存在多样化。在某些情况下，MCI可以为阿尔茨海默症的一个病理阶段。某些形式的认知损害可视为神经变性疾病的早期表现，最终将导致痴呆。在某些情况下，所述MCI可以包括选自下组的亚型：aMCI-s：遗忘型MCI单认知域受损；aMCI-m：遗忘型MCI多认知域受损；naMCI-s：非遗忘型MCI单认知域受损；以及naMCI-m：非遗忘型MCI多认知域受损。
术语“正常衰老引起的认知障碍”通常是指由于正常衰老引起的认知损害。例如，所述正常衰老引起的认知障碍可以表现为：记忆丧失、对熟悉地方的位置感到困惑、完成日常工作需要比平时更长时间、或者情绪和性格的变化。
术语“lews身体痴呆(LBD)”通常是指Lewy Body Detmentia，路易体痴呆。路易体痴呆的特征是蛋白质异常积聚成为称为路易体的肿块。路易体痴呆症导致心智能力逐渐下降。路易体痴呆症患者可能会出现幻视以及警觉性和注意力的变化。其他影响包括肌肉僵硬、运动缓慢、行走困难和颤抖。大脑中有路易体的患者也可以有与阿尔茨海默病相关的斑块和缠结。
术语“额颞叶痴呆”通常是指皮克病，其是一种tau蛋白只影响大脑的额叶和颞叶的渐进性的罕见病。额颞叶痴呆患者在更高层次的推理、表达语言、语言感知和记忆形成方面都有困难。额颞叶痴呆患者的大脑的额叶和颞叶可以随着时间推移而萎缩。
术语“血管性痴呆”通常是指由于大脑血液流动受损而导致的推理、判断和记忆方面的问题。例如，所述血管性痴呆可以包括由于心脏病和中风风险的因素，如高血压和高胆固醇而导致的痴呆。
术语“多发性梗死型”通常是指由大型脑动脉的单个穿通支闭塞引起的非皮质性小梗死。所述多发性梗死型可以是脑梗塞的一种特殊类型，又称缺血性中风。所述多发性梗死型可以表现为偏身感觉障碍、失语、构音不全、动作缓慢、笨拙(尤以精细动作如书写更为困难)。
术语“帕金森氏症”通常是指一种进行性神经变性疾病。所述帕金森氏症(PD)的临床特征可以包括运动症状(例如震颤、运动徐缓、肌强直和姿势不稳)，以及神经精神性及其他非运动表现。例如，所述非运动表现可以包括认知功能障碍和痴呆、心境障碍(例如抑郁、焦虑、情感淡漠)和睡眠障碍。
术语“克雅氏病(CJD)”通常是指一种发生在人类身上的传染性海绵状脑病。CJD是由朊病毒感染引起的疾病。CJD患者可以表现为偏执行为，意识模糊、食欲及体重下降、抑郁，少数患者有视觉或听觉异常；在进展期，表现为进行性的神经系统病情恶化(例如感觉异常、语言障碍和失语)。
术语“多发性硬化(MS)”通常是指一种脱髓鞘性神经病变。所述MS患者脑或脊髓中的神经细胞表面的绝缘物质(即髓鞘)受到破坏，神经系统的信号转导受损，可以导致一系列可能发生的症状，影响患者的活动、心智、甚至精神状态。这些症状可以包括复视、单侧视力受损、肌肉无力、感觉迟钝，或协调障碍。
术语“肌萎缩性脊髓侧索硬化症(ALS)”通常是指渐冻人症、运动神经元病，是一种渐进且致命的神经退行性疾病。其中少数ALS患者可以出现额颞叶痴呆。部分ALS患者的觉、视觉、触觉、嗅觉和味觉会出现退化，极少数肌萎缩侧索硬化症患者会同时出现痴呆症。
术语“亨廷顿病(HD)”，即亨廷顿舞蹈症，通常是指一种会导致脑细胞死亡的遗传性疾病。HD患者随着疾病的进展，身体运动的不协调变得更加明显，能力逐渐恶化直到运动变得困难，无法说话。心智能力则通常会衰退为痴呆症。
术语“老龄阶段”通常是指受试者的老龄化阶段。例如，对人而言，所述老龄阶段可以为60岁以上，70岁以上或75岁以上；对小鼠而言，所述老龄阶段可以为10月龄以上，例如可以为13月龄以上或者18月龄以上。在某些情况下，所述老龄阶段的受试者可以具有学习缺陷、记忆力障碍、记忆力缺陷和/或脑功能障碍中的一种或多种症状。
术语“变体”通常是指包含与亲本或参考多肽(例如野生型多肽)的氨基酸序列相差至少一个氨基酸残基的氨基酸序列的多肽。在本申请中，所述变体可以与所述亲本或参考多肽具有较高的(例如至少80％)的同源性。所述同源性可以包括序列的相似性或同一性。在本申请中，所述同源性可以用本领域已知的标准技术确定(参见例如Smith和Waterman，Adv.Appl.Math.《应用数学进展》)；多核苷酸或多肽序列共有的同一性百分数通过分子之间序列信息的直接比较测定，所述比较通过序列比对并使用本领域已知的方法确定同一性来进行。适用于确定序列相似性的算法的例子是BLAST算法(参见Altschul等人，J.Mol.Biol.《分子生物学杂志》，215:403-410[1990])。用于进行BLAST分析的软件可通过国家生物技术信息中心(NCBI)公开获得。
术语“约”通常是指在比特定值多或少20％的数值范围。例如，“约X”包括为X的±20％、±10％、±5％、±2％、±1％、±0.5％、±0.2％或±0.1％的数值范围，其中X为数值。
发明详述
早老性痴呆的早发现、早治疗的前提条件是找到其特异性的诊断标志物。发明人在研究中发现早老性痴呆病人血液中存在QDLIR蛋白，而在正常人的血样中没有检测到。这个结果表明QDLIR很有可能成为早老性痴呆诊断(有可能是早期诊断)的血液标志物；同时发明人对其他认知障碍病人的血样也进行了检测，发现在MCI(mild cognitive impairment，轻度认 知障碍)等认知障碍的病人血样中QDLIR蛋白水平明显升高，因此血中QDLIR升高将是早老性痴呆诊断和治疗过程中的一个新标志物。
具体而言，在病理状态下(如肿瘤、神经退行性疾病等)，病变组织缺少血供，导致低氧，引起一系列基因表达的变化，同时还有异常剪切体出现。如何能找到其中特异性的基因及其剪切体有可能是打开治疗疾病的关键钥匙。发明人在前期的研究中发现：QDLIR是人体内异常剪接形成的异常蛋白，且只在人体组织和细胞才存在。QDLIR是在细胞低氧环境下，可通过高表达和分泌对细胞和组织的功能进行适应性调控的关键分子。
由此，发明人提出了假设：QDLIR有可能通过病变组织中的血供系统进入血液中。随后，发明人通过如下试验进一步证明了该假设的正确性：发明人首先通过外转QDLIR的质粒到HEK293细胞中，然后在培养基中检测到大量的QDLIR蛋白的存在，证明QDLIR可以分泌到细胞外；随后在AD和MCI病人血浆中检测到QDLIR，而在正常人和非AD病人的血浆中则未检出或检测极低水平的QDLIR。这个结果表明QDLIR特异性存在于AD和MCI患者的血液中，从而可作为AD诊断(如早期诊断)的血液标志物；同时，在对其他一些存在认知障碍的病人血样的检测中，发现QDLIR蛋白水平明显升高，因此血中QDLIR水平可作为AD(早老性痴呆)诊断和治疗过程中的新标志物。
本申请中提供的所有数值范围旨在清楚地包括落在范围端点之间的所有数值及它们之间的数值范围。可对本申请提到的特征或实施例提到的特征进行组合。本说明书所揭示的所有特征可与任何组合物形式并用，说明书中所揭示的各个特征，可以任何可提供相同、均等或相似目的的替代性特征取代。因此除有特别说明，所揭示的特征仅为均等或相似特征的一般性例子。
如本申请所用，“含有”、“具有”或“包括”包括了“包含”、“主要由……构成”、“基本上由……构成”、和“由……构成”；“主要由……构成”、“基本上由……构成”和“由……构成”属于“含有”、“具有”或“包括”的下位概念。
在本申请中，所述神经退行性疾病可以包括急性神经退行性疾病和慢性神经退行性疾病。例如，所述神经退行性疾病可以包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
在本申请中，所述神经退行性疾病可以包括早老性痴呆。例如，所述神经退行性疾病可以包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
在本申请中，所述认知障碍可以包括早期认知障碍(MCI)、中期认知障碍和晚期认知障 碍。例如，所述认知障碍可以包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。例如，所述认知障碍的诱导疾病可以包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
在本申请中，所述认知障碍可以包括早期认知障碍(MCI)(例如，失去短期记忆、表达或理解抽象事情时感困难、情绪或行为变幻无常、学习新事物及跟随复杂指令感到困难、判断力减退和/或基本自理需要旁人提醒)、中期认知障碍(例如，混淆远期记忆和现实情况记忆、词不达意、行为性格转变或情绪不稳和/或需要旁人协助自理)或晚期认知障碍(例如，记忆缺损、身体活动及精神状况出现衰退、不能有效表达或沟通、不能自理和/或生物钟混乱)。
在本申请中，所述认知障碍可以包括患有能够引起诱导所述认知障碍的疾病。例如，所述受试者可以患有阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
诊断靶点QDLIR
一方面，本申请提供了一种DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸，在疾病诊断和/或疾病进程评估中的用途，其中所述疾病可以包括认知障碍。
一方面，本申请提供了一种DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸，在疾病诊断和/或疾病进程评估中的用途，其中所述疾病可以包括神经退行性疾病。
DDIT4L(DNA Damage Inducible Transcript 4 Like，SEQ ID NO:1)是一种蛋白质编码基因，其与mTOR信号转导途径相关，在巨噬细胞中其水平可通过氧化LDL和低氧上调。
DDIT4L基因通过正常剪切(normal splicing)去除内含子后可得到编码193aa长DDIT4L蛋白(SEQ ID NO:6)的核苷酸序列(SEQ ID NO:5)。然而，本申请人出乎意料地发现，该基因还存在另一种特殊的剪接方式，即内含子滞留式(Intron retention)剪接。通过该特殊的剪接方式获得2212nt的核苷酸序列，并由于内含子内可以包含终止子而导致最终获得84aa的剪切产物肽(SEQ ID NO:4)。在本申请中称所述通过内含子滞留式剪切获得的蛋白质可以为“QDLIR”(如图1a所示)。
如本申请所用，除非另有说明或可明确判定，术语“QDLIR”可可以包括DDIT4L基因通过内含子滞留式剪切获得的成熟mRNA、剪切肽编码分子或剪接产物肽(或称QDLIR蛋白)。
当剪切产物可以为mRNA时，其可选自：(a)序列如SEQ ID NO：2所示的核苷酸分子；或(b)与SEQ ID NO:2的核苷酸分子具有高序列相同性(例如序列相同性至少可以为 80％、85％、90％、92％、95％、98％、99％、99.5％、99.8％)的核苷酸分子。
当剪切产物可以为QDLIR编码分子时，其可选自：(a)序列如SEQ ID NO：3所示的核苷酸分子；或(b)与SEQ ID NO:3的核苷酸分子具有高序列相同性(例如序列相同性至少可以为80％、85％、90％、92％、95％、98％、99％、99.5％、99.8％)的核苷酸分子。
当剪切产物可以为QDLIR蛋白时，其可选自：(a')序列如SEQ ID NO：4所示的多肽分子；(b')与SEQ ID NO:4的多肽分子具有高序列相同性(例如序列相同性至少可以为80％、85％、90％、92％、95％、98％、99％、99.5％、99.8％)的蛋白质分子；(c')由如上(a)或(b)中任一核苷酸分子翻译得到的蛋白质分子；或(d')如上(a')～(c')中任一所述的蛋白质分子的保守性变异多肽、或其同源蛋白或多肽、或其活性片段。
诊断物质和诊断试剂盒
另一方面，本申请提供一种检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质，在制备用于疾病诊断和/或疾病进程评估的产品中的用途，其中所述疾病可以包括认知障碍。
另一方面，本申请提供一种检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质，在制备用于疾病诊断和/或疾病进程评估的产品中的用途，其中所述疾病可以包括神经退行性疾病。
如本申请所用，术语“检测用物质”、“检测试剂”或“检测QDLIR的试剂”或“检测QDLIR表达量的试剂”可互换使用，均是指特异性针对QDLIR分子，且可用于直接或间接检测出QDLIR分子的存在和/或含量的物质。这些检测物质可在基因水平或蛋白质水平上检测QDLIR。
在本申请中，所述物质可以包括能够特异性结合QDLIR的抗体、能够特异性结合QDLIR的探针、能够特异性扩增编码QDLIR的核酸的引物、能够特异性检测编码QDLIR的核酸的基因芯片、能够特异性结合编码QDLIR的核酸的适体、和/或能够特异性结合编码QDLIR的核酸的向导RNA。
根据QDLIR分子的序列，本领域普通技术人员可基于常规手段制备或通过市售获得特异性针对QDLIR分子的试剂。例如，本申请中可用的检测试剂可以包括但不限于：对QDLIR分子具有检测特异性的抗体(例如，可以为单克隆抗体)、探针、基因芯片、PCR引物、gRNA等。
并且，可以为了便于检测，本申请的检测试剂还可带有可检测标记，所述可检测标记可以包括但不限于：放射性同位素、荧光团、化学发光部分、酶、酶底物、酶辅因子、酶抑制剂、染料、金属离子、配体(如，生物素或半抗原)等。
本申请的检测试剂可存在于溶液中、固定于载体(如基片、吸附物)上或以其它本领域中常规的方式存在，只要该存在方式适于对生物样品中QDLIR检测即可。例如，当本申请的检测试剂可以为核苷酸探针时，其可以生物芯片(或称“微阵列”)的形式存在。
在本申请中，所述产品可以以试剂盒的形式存在。
本申请中还提供了一种用于早老性痴呆诊断和/或早老性痴呆发展进程(例如分级或分期)评估的产品，其可以包含：用于检测QDLIR水平的物质，以及可任选的，用于早老性痴呆诊断和/或早老性痴呆发展进程(例如分级或分期)评估的其他物质，例如现有AD标志物的检测物质。
在本申请中，所述诊断试剂盒可以包含说明书，所述说明书记载了利用所述诊断试剂盒诊断认知障碍，和/或评估认知障碍的疾病进程；和/或诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程的方法。
根据所用检测方法的需要，可选择适当的QDLIR检测物质，并将其制成适于所用检测方法的产品，如试剂盒。本领域普通技术人员可根据实际条件和需要对检测方式和产品中所含试剂进行调整和改变。
由此，本申请中还提供了一种产品(如检测试剂盒)，其可以包含：(i)检测有效量的用于检测QDLIR的一种或多种试剂；(ii)可任选地，选自下组的一种或多种物质：容器、使用说明书、阳性对照物、阴性对照物、缓冲剂、助剂或溶剂，例如用于混悬或固定细胞的溶液，可检测的标签或标记，使核酸易于杂交的溶液，用于裂解细胞的溶液，或用于核酸纯化的溶液。
在一个示例中，本申请提供了一种适于通过免疫组织化学方法检测生物样品中QDLIR蛋白的表达的检测试剂盒。该检测试剂盒可可以包含：封闭液，例如10％山羊血清；一抗，例如兔抗人或小鼠D4LIR单克隆抗体；二抗，例如标记(如HRP标记的)或未标记的羊抗兔二抗；底物缓冲液，例如DAB底物缓冲液；显色液；以及可选的装有上述试剂的容器及使用说明书。
本申请的检测试剂盒中还可附有试剂盒的使用说明书，其中记载了如何采用试剂盒进行检测，以及如何利用检测结果诊断认知障碍，和/或评估认知障碍的疾病进程，和/或诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程。例如，可以利用本申请所述的试剂盒进行AD诊断和/或AD进程(例如分级或分期)的评估。
本申请的产品还可可以包含临床上用于对象中AD诊断和/或AD发展进程(例如分级或分期)评估的其它试剂，以辅助或验证通过检测QDLIR所得到的结果。本领域普通技术人员 可根据具体需要进行常规选择。
诊断方法
另一方面，本申请提供了一种诊断认知障碍，和/或评估认知障碍的疾病进程的方法，其可以包括以下步骤：检测源自有需要的受试者的样本中，DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的含量。
另一方面，本申请提供了一种诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程的方法，其可以包括以下步骤：检测源自有需要的受试者的样本中，DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的含量。
在本申请中，所述受试者可以包括哺乳动物。
在本申请中，所述受试者可以包括肿瘤患者。
在本申请中，所述受试者处于老龄阶段。
在本申请中，所述样本可以包括血液样品和/或组织或细胞样品。
在本申请中，所述样本可以包括全血、血清、血浆和/或脑脊液。
在本申请中，所述方法可以包括：将所述受试者的样本的所述QDLIR和/或编码QDLIR的核酸的含量与正常对照值进行比较，其中所述正常对照值可以包括正常的受试者中所述QDLIR和/或编码QDLIR的核酸的含量。
在本申请中，所述正常的受试者可以不患有所述认知障碍。所述正常的受试者可以不患有所述神经退行性疾病。
在本申请中，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于(例如可以提高至少约20％、至少约50％、至少约60％、至少约65％、至少约70％、至少约75％、至少约80％、至少约85％、至少约90％、至少约95％、至少约100％、至少约200％、至少约500％或更多)所述正常对照值，所述受试者被诊断可以为患有所述认知障碍和/或所述神经退行性疾病。
在本申请中，所述方法可以包括：将所述受试者的样本的所述QDLIR和/或编码QDLIR的核酸的含量与早期对照值进行比较，其中所述早期对照值可以包括同一所述受试者之前测得的所述QDLIR和/或编码QDLIR的核酸的含量。
在本申请中，所述受试者已确诊患有所述认知障碍和/或所述神经退行性疾病。例如，所述受试者已确诊患有所述认知障碍；或者，所述受试者已确认患有所述神经退行性疾病。
在本申请中，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于 (例如可以提高至少约20％、至少约50％、至少约60％、至少约65％、至少约70％、至少约75％、至少约80％、至少约85％、至少约90％、至少约95％、至少约100％、至少约200％、至少约500％或更多)所述早期对照值，所述受试者被诊断可以为所述认知障碍和/或所述神经退行性疾病的疾病进程加剧。
在本申请中，所述的方法还可以包括以下的步骤：检测源自所述受试者的样本中，与认知障碍和/或神经退行性疾病相关的标志物的含量。
在本申请中，所述与认知障碍和/或神经退行性疾病相关的标志物可以包括AD7C-NTP、pTau-181、pTau-217和/或Aβ1-42。所述标志物的含量的检测结果可以用于辅助诊断认知障碍，和/或评估认知障碍的疾病进程，和/或诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程。
在本申请中，所述的方法还可以包括以下的步骤：观察所述受试者的脑部成像。所述脑部成像的结果可以用于辅助诊断认知障碍，和/或评估认知障碍的疾病进程，和/或诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程。
诊断系统
另一方面，本申请提供一种诊断认知障碍，和/或评估认知障碍的疾病进程的系统，其可以包含检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质。
另一方面，本申请提供一种诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程的系统，其可以包含检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质。
在本申请中，所述的系统可以包含检测模块，所述检测模块能够检测所述QDLIR和/或编码所述QDLIR的核酸的含量值。
在本申请中，所述检测模块检测所述收集模块获得的所述样本中所述QDLIR和/或编码所述QDLIR的核酸的含量值。
在本申请中，所述检测模块可以包含能够检测所述QDLIR和/或编码所述QDLIR的核酸的含量的物质。
在本申请中，所述物质可以包括能够特异性结合QDLIR的抗体、能够特异性结合QDLIR的探针、能够特异性扩增编码QDLIR的核酸的引物、能够特异性检测编码QDLIR的核酸的基因芯片、能够特异性结合编码QDLIR的核酸的适体、和/或能够特异性结合编码QDLIR的核酸的向导RNA。
在本申请中，所述检测模块可以包含能够检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的含量的仪器。例如，所述仪器可以包括进行qPCR、ELISA、western blot所需的仪器，和/或基因芯片。
在本申请中，所述检测模块可以包含能够检测与认知障碍和/或神经退行性疾病相关的标志物的含量的物质和/或仪器。
在本申请中，所述与认知障碍和/或神经退行性疾病相关的标志物可以包括AD7C-NTP、pTau-181、pTau-217和/或Aβ1-42。例如，所述物质还可以包括能够特异性结合所述标志物的抗体、能够特异性结合所述标志物的探针、能够特异性扩增编码所述标志物的核酸的引物、能够特异性检测编码所述标志物的核酸的基因芯片、能够特异性结合编码所述标志物的核酸的适体、和/或能够特异性结合编码所述标志物的核酸的向导RNA。例如，所述仪器可以包括进行qPCR、ELISA、western blot所需的仪器，和/或基因芯片。
在本申请中，所述检测模块输出可以所述QDLIR和/或编码所述QDLIR的核酸的含量值。
在本申请中，所述的系统可以包含判断模块，所述判断模块能够根据所述QDLIR和/或编码所述QDLIR的核酸的含量与正常对照值和/或早期对照值，判断所述受试者的诊断结果和/或疾病进程结果；其中，所述正常对照值可以包括正常的受试者中所述QDLIR和/或编码QDLIR的核酸的含量；其中，所述早期对照值可以包括同一所述受试者之前测得的所述QDLIR和/或编码QDLIR的核酸的含量。
在本申请中，所述之前可以包括至少10天前、至少20天前、至少1个月前、至少6个月前、至少12个月前、至少24个月前或更久之前。在本申请中，所述受试者可以已确诊患有所述认知障碍和/或所述神经退行性疾病。利用本申请所述的系统，可以追踪所述受试者在不同时间阶段的所述QDLIR和/或编码QDLIR的核酸的含量，从而可以帮助诊断所述受试者的所述认知障碍和/或所述神经退行性疾病的疾病进程。
在本申请中，所述正常的受试者可以不患有所述认知障碍和/或所述神经退行性疾病。
例如，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于(例如可以提高至少约20％、至少约50％、至少约60％、至少约65％、至少约70％、至少约75％、至少约80％、至少约85％、至少约90％、至少约95％、至少约100％、至少约200％、至少约500％或更多)所述正常对照值，所述判断模块将所述受试者诊断可以为患有所述认知障碍和/或所述神经退行性疾病。
例如，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于(例如 可以提高至少约20％、至少约50％、至少约60％、至少约65％、至少约70％、至少约75％、至少约80％、至少约85％、至少约90％、至少约95％、至少约100％、至少约200％、至少约500％或更多)所述早期对照值，所述判断模块将所述受试者诊断可以为所述认知障碍和/或所述神经退行性疾病的疾病进程加剧。
在本申请中，所述判断模块可以输出所述受试者的诊断结果。
在本申请中，所述的系统可以包括显示模块，其中所述显示模块能够显示出所述判断模块的诊断结果。例如，所述显示模块所显示的诊断结果可以包括定性结果和/或定量结果。
在本申请中，所述的系统可以包括存储模块，其中所述存储模块能够存储所述检测模块输出的所述含量值，和/或所述判断模块输出的诊断结果。
在本申请中，所述系统可以包括如下装置：
(a)采集和/或接收样品中QDLIR水平数据的装置；
(b)对数据进行分析以对所述对象的AD诊断和/或AD发展进程进行评估的装置，其中若QDLIR高于对照水平则表明所述对象罹患AD和/或AD有所发展。
在本申请中，所述系统还进一步可以包括(a')采集和/或接收样品中AD诊断和/或AD发展进程的其他临床指标或基本特征数据的装置。
在本申请中，(b)中所述的装置用于将QDLIR水平数据与AD诊断和/或AD发展进程的其他临床指标或基本特征数据进行结合分析以对所述对象的AD诊断和/或AD发展进程进行预后评估。
或者，在本申请中，所述系统还进一步可以包括对AD诊断和/或AD发展进程的其他临床指标或基本特征数据进行分析的设备；和/或将QDLIR水平数据与AD诊断和/或AD发展进程的其他临床指标或基本特征数据进行结合分析以对所述对象的AD诊断和/或AD发展进程进行评估的装置。
在本申请中，所述系统还可以包括选自下组的一种或多种装置：用于输入和/或输出和/或存储数据的装置；用于输出和/或存储分析结果的装置，例如用于将分析结果上传和/或存储到云数据库的装置或其相应的云数据库装置；根据评估结果智能分析和推荐治疗方案的装置。
应理解，本领域技术人员可根据需要在系统中添加其他常规装置或程序。
通常，可采用如下方法进行AD诊断和/或AD发展进程(例如分级或分期)评估：检测待测对象或获自该对象的样品中QDLIR分子的水平，并将该水平与对照水平相比较；若比较结果显示对象中的QDLIR分子的水平高于对照水平，则提示所述对象罹患AD或AD有所发展。在一些实施方式中，本申请的方法还可选地包括：从对象获得待测样品；使待测样品与 检测QDLIR水平的试剂或试剂盒接触。
如本申请所用，术语“对照”可包括正常对照或AD特定发展分级或分期的对照。“对照”可指用作参照的QDLIR分子的水平，其包括但不限于：由同一对象的非AD正常生物样品(例如获自该对象非AD病人或正常人的样品)中测得的QDLIR分子水平、通过统计学确定的群体标准水平、或经标准化的水平。术语“AD特定发展分级或分期的对照”是指由同一对象之前已确定的发展分级或分期时测得的QDLIR分子水平、通过统计学确定的该分级或分期时的群体标准水平、或经标准化的水平。
药物筛选方法
另一方面，本申请提供一种药物筛选方法，其可以包括以下的步骤：检测施用候选药物后，受试者中QDLIR和/或编码QDLIR的核酸的含量的变化，其中所述药物用于预防、治疗和/或缓解认知障碍和/或神经退行性疾病。
在本申请中，当施用所述候选药物中，受试者中QDLIR和/或编码QDLIR的核酸的含量降低，则所述候选药物具有治疗效果。
在本申请中，所述施用可以包括注射。
在本申请中，所述药物可以包括小分子药物和/或生物大分子药物。
根据本申请中所公开的内容，QDLIR的水平与认知障碍存在与否和/或程度密切相关，从而可作可以为AD诊断和/或AD发展进程(例如分级或分期)评估指标。
此外，本申请还提供了一种筛选治疗AD的候选药物的方法，所述方法包括测试所述候选药物对对象或获自对象的样品中QDLIR水平的影响，其中，在使用所述候选药物后，QDLIR水平降低表明所述候选药物具有治疗AD的效果。本申请候选药物筛选方法所涉及的各特征可如本申请中所限定或阐述。在一些实施方式中，候选药物可以为QDLIR的抑制剂。
本申请发现QDLIR特异性表达在AD病人血样和/或脑脊液中。重要的一点，QDLIR可以在少量的血清(如100μl左右)中被检测到，这为临床应用提供了很有前景的AD早期诊断的检测手段。目前结果提示AD和MCI等疾病的阳性检出率均在90％以上，QDLIR极具成为AD早筛特异性标志物的应用前景。
目前临床血液检测中开展的pTau-181、pTau-217、Aβ1-42是新开展的AD早筛的检测项目，特异性相对较低，且费用比较昂贵。而与这些现有AD早筛的检测相比，本申请中基于QDLIR水平的AD早筛检测检出率高、特异性非常好，和/或成本低，具有价格优势，例如通过ELISA检测QDLIR的水平其成本可控制在低于10元/人。由此，本申请的检测指标和方法与现有方法相比具有较大的效果和成本优势。
此外，本申请发现QDLIR对于认知障碍和/或神经退行性疾病的诊断非常具有特异性和灵敏性，可以在pTau-181、pTau-217、Aβ1-42等靶点差异不显著的情况下，区分出潜在的AD和/或MCI患者，总而可以在更早期对患者进行有效地诊断。
序列表信息：
SEQ ID NO: 序列名称 长度
1 DDIT4L基因全长序列 4599nt
2 QDLIR内含子保留剪切成熟mRNA 4163nt
3 QDLIR蛋白编码分子 255bp
4 QDLIR蛋白 84aa
5 DDIT4L蛋白编码分子 582nt
6 DDIT4L蛋白 193aa
本申请提供了以下的实施方式：
1.检测来自对象样品中DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR的物质在制备用于所述对象早老性痴呆诊断、轻度认知障碍诊断和/或早老性痴呆发展进程(例如分级或分期)评估的产品(例如试剂盒或检测系统)中的应用。
2.如实施方式1所述的应用，其中：所述剪切产物选自下组：具有SEQ ID NO:2所示核苷酸序列的mRNA；具有SEQ ID NO:3所示核苷酸序列的剪接产物肽编码核酸；或者，具有SEQ ID NO:4所示氨基酸序列的剪接产物肽。
3.如实施方式1所述的应用，其中，所述早老性痴呆包括：早老性痴呆早期(健忘期)、早老性痴呆中期(神经紊乱期)和早老性痴呆晚期(严重痴呆期)。
4.如实施方式1所述的应用，其中，所述物质选自用于检测QDLIR的试剂、设备或其组合。
5.如实施方式1所述的应用，其中，所述物质选自：对所述QDLIR具有特异性的物质，例如所述QDLIR的抗体(优选单克隆抗体)；QDLIR特异性的探针、基因芯片、PCR引物、gRNA等。
6.如实施方式1所述的应用，其中，所述样品选自：血液样品，如全血、血清、血浆；脑脊液。
7.如实施方式1所述的应用，其中，与正常对照值相比，所述样品中所述QDLIR的水平提高，表明所述对象罹患早老性痴呆或轻度认知障碍；或者，与轻度认知障碍对照相比或与已诊断为早老性痴呆对象的前期对照水平相比，所述样品中QDLIR的水平提高，表明所述对象早老性痴呆有所发展(例如分级或分期更高)。
8.一种用于早老性痴呆诊断、轻度认知障碍诊断和/或早老性痴呆发展进程(例如分级或分期)评估的产品(例如试剂盒或系统)，其包含：
(i)用于检测对象样品中DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR的物质；以及
(ii)可任选的，用于早老性痴呆诊断、轻度认知障碍诊断和/或早老性痴呆发展进程(例如分级或分期)评估的其他物质，例如用于检测现有早老性痴呆标志物的检测物质，
例如，在一些情况下：
(i)中的物质为选自下组中的一种或多种：
用于检测所述QDLIR的试剂、设备或其组合；
用于在基因水平和/或蛋白质水平上检测所述QDLIR的物质，如用于选自下组的一种或多种检测技术或方法中的物质：免疫组织化学法(如免疫荧光分析、化学发光法、反向酶联免疫吸附、免疫胶体金法)、Western印迹法、Northern印迹法、PCR、生物芯片法；或者
对所述QDLIR具有特异性的物质，例如其抗体(优选单克隆抗体)；剪切产物特异性的探针、基因芯片、PCR引物、gRNA；和/或
(ii)中的其他物质选自：用于检测一种或多种其他早老性痴呆标志物的物质，例如：AD7C-NTP、pTau-181、pTau-217、Aβ1-42；和/或，用于大脑淀粉样蛋白成像的试剂；和/或，用于对象脑部成像诊断的试剂。
9.一种用于对象早老性痴呆诊断、轻度认知障碍诊断和/或早老性痴呆发展进程(例如分级或分期)评估的系统，其包括如下装置：
(a)采集和/或接收样品中DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR水平数据的装置；
(b)对数据进行分析以对所述对象的早老性痴呆进行诊断、轻度认知障碍进行诊断和/或早老性痴呆发展进程(例如分级或分期)进行评估的装置，
其中，与正常对照值相比，所述样品中所述QDLIR的水平提高，表明所述对象罹患早老性痴呆或轻度认知障碍；或者，与轻度认知障碍对照相比或与已诊断为早老性痴呆对象的前期对照水平相比，所述样品中QDLIR的水平提高，表明所述对象早老性痴呆有所发展(例如分级或分期更高)。
10.一种筛选早老性痴呆治疗药物的方法，所述方法包括：检测候选药物对对象中DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR水平的影响，其中，在使用所述候选药物后，所述水平降低则表明所述候选药物具有早老性痴呆治疗效果。
不欲被任何理论所限，下文中的实施例仅仅是为了阐释本申请的检测方法、应用等，而不用于限制本申请发明的范围。
实施例
下面结合具体实施例，进一步阐述本申请。应理解，这些实施例仅用于说明本申请而不用于限制本申请的范围。本领域技术人员可对本申请做出适当的修改、变动，这些修改和变动都在本申请的范围之内。
下列实施例中未注明具体条件的实验方法，可采用本领域中的常规方法，例如参考《分子克隆实验指南》(第三版，纽约，冷泉港实验室出版社，New York：Cold Spring Harbor Laboratory Press，1989)或按照供应商所建议的条件。DNA的测序方法为本领域常规的方法，也可由商业公司提供测试。
除非另外说明，否则百分比和份数按重量计算。除非另行定义，文中所使用的所有专业与科学用语与本领域熟练人员所熟悉的意义相同。此外，任何与所记载内容相似或均等的方法及材料皆可应用于本申请方法中。文中所述的较佳实施方法与材料仅作示范之用。
实施例1 QDLIR蛋白是一种分泌蛋白
试验材料与方法
质粒构建和细胞转染方法：将人QDLIR的cDNA克隆到pCMV-flag载体中以表达flag-QDLIR蛋白。在HEK293细胞(中科院生化细胞所)上，通过Lippo3000试剂转染pCMV-flag质粒和pCMV-flag-QDLIR质粒，细胞培养48小时后，收获细胞和培养基，各加入适量裂解液，4℃旋转裂解30分钟，12000g离心30分钟后取上清，取少量裂解液以备Western Blot分析。
Western Blot实验：按照SDS-PAGE电泳、转膜、免疫反应和化学发光的步骤完成Western Blot实验(所用抗体购自吉尔生化(上海)有限公司)。
结果与讨论
在HEK293细胞培养48小时后，转染pCMV-flag-QDLIR质粒的细胞中有大量的QDLIR的表达，同时在细胞的培养基中也检测到有大量的QDLIR存在(图1b)；这表明QDLIR在细胞中转录、翻译成蛋白质后，可以通过胞内的分泌途径被分泌到细胞外的培养基中，从分泌量来看，胞内合成的QDLIR大部分被分泌到了细胞外的培养基中，这提示QDLIR在人体内也会被分泌到细胞外，从而进入血液循环系统，这为它成为某些病理条件下的一个血液标志物提供了理论上的基础。
实施例2 QDLIR蛋白在AD病人血液中的含量变化
试验材料与方法
组织标本：AD和MCI患者、非AD患者(高血压、冠心病或糖尿病患者)和正常人静脉血标本(共计58个样本)，来自复旦大学附属华山医院、徐汇区中心医院。
静脉血采集和获取血浆方法：使用真空采血技术采集患者肘窝静脉于EDTA真空管中，立即将试管轻轻颠倒8次使血液与抗凝剂充分混匀。之后冰上运输至实验室，4℃1500g离心10分钟，吸出上清分装，-80℃保存。
血浆Western Blot方法：分装血浆冰上融化后用PBS适当稀释，加入上样缓冲液100℃金属浴变性5分钟，后续步骤同组织Western Blot方法。
血浆ELISA方法：血浆标本以1×稀释液(OBT1998G，购自Bio-Rad)按1:1进行稀释；移液器取100μl封闭液37℃下封闭1小时；移液器取100μl#1～8离心管内(倍比稀释)标准品、稀释的血浆加入到捕捉抗体(吉尔生化，AB009458)包被好的板孔内，每次取样时更换枪头；37℃下孵育1.5小时；移去各孔内液体，以100μl 1×洗涤液(BUF031C，购自Bio-Rad)洗涤3次；向各孔加100μl检测抗体(吉尔生化，AB008349)，37℃下孵育1小时；移去各孔内液体，以100μl 1×洗涤液洗涤3次；向各孔加100μl 1×亲和素-HRP溶液，37℃下孵育1小时；移去各孔内液体，以100μl 1×洗涤液洗涤3次；向各孔加100μl 1×底物反应液(BUF062C，购自Bio-Rad)，室温下避光下孵育10-30分钟；利用酶标仪在655nm波长下检测，并建立标准曲线。
结果与讨论
通过临床收集AD和MCI患者、非AD患者和正常人的血样，对不同病人进行了检测，发现AD和MCI病人血样中QDLIR的含量最高，而且在约95％的患者中可以检测到；MCI病人血样中也含有较高量的QDLIR，有90％左右的患者可以检测到；而在无认知障碍患者的血样中检测不到或检测到很微量的QDLIR，正常人的血样中检测不到QDLIR(图2a)。
通过ELISA的方法，发现AD病人血样中QDLIR的含量比较高，可以达到10-100ug/ml的水平，而非AD患者的QDLIR的含量均在pg/ml的水平，正常人检测不到，故AD患者的QDLIR水平是其他患者的103-104倍。这提示QDLIR的特异性非常高(图2a和2b)。
以上结果表明，QDLIR在AD和MCI患者的血液中具有高浓度，可作为血液标志物检测AD的存在，尤其是针对MCI等轻度认知功能障碍的患者。
并且，QDLIR含量与AD的严重程度相关(见图2b)。
实施例3 QDLIR蛋白检测的方法学研究
取QDLIR蛋白(吉尔生化，752084)，用DPBS溶液配置成1mg/ml的原液，在实验前用ELISA洗涤液(BUF031C，购自Bio-Rad)稀释，制得标准品QDLIR蛋白溶液。
采用ELISA法检测标准品QDLIR蛋白的浓度，并采用多项式回归拟合。所得标准曲线如图3所示。
该回归模型的R 2值为0.99，说明拟合程度很高；阴影部分显示了该回归模型的95％置信区间，图中可见置信区间狭长，说明根据该模型估计样品中QDLIR含量的可信度高。
实施例4 QDLIR作为临床检测指标的R℃曲线
针对临床收集AD和MCI患者、非AD患者和正常人的血样(同实施例2)，进行了ELISA检测，依据检测结果利用R软件(版本号3.6.2)的“pROC”包分析，以检测到的QDLIR浓度建立Logistic回归模型，采用自助抽样法分析可得ROC曲线。
结果如图4所示。结果显示：只用QDLIR单参数建模即可获得较理想的预测结果，AUC＝0.807(95％置信区间为0.688-0.926)。
实施例5 QDLIR蛋白在不同类别MCI病人血液中的含量变化
试验材料与方法
组织标本：不同类型MCI患者、正常随访人静脉血标本，来自复旦大学附属华山医院。
静脉血采集和获取血浆方法：同实施例2。
血浆ELISA方法：同实施例2。
MCI分类方法：aMCI-s：遗忘型MCI单认知域受损；aMCI-m：遗忘型MCI多认知域受损；naMCI-s：非遗忘型MCI单认知域受损；naMCI-m：非遗忘型MCI多认知域受损。
在实施例5中，其中aMCI-m型患者122例样本；aMCI-s型患者85例样本；naMCI-m型患者11例样本；naMCI-s型患者88例样本；正常随访人279例样本。
结果与讨论
通过临床收集不同类型MCI患者、正常随访人群的血样，对其进行了检测，发现MCI患者中aMCI-m类型的病人血样中QDLIR的含量升高，与正常随访人群相比有显著性差异(图5)。
通过ELISA的方法，发现aMCI-m类型的病人血中QDLIR的含量比较高，而aMCI-m类型病人相对于其他类型MCI病人更容易进展成为AD，提示了QDLIR这个指标有区分aMCI-m类型的能力。
上述结果表明，QDLIR可作为AD的有效诊断标志物。
实施例6 QDLIR蛋白在不同类别MCI病人血液中的含量变化
试验材料与方法
组织标本：不同类型MCI患者、AD患者和正常随访人静脉血标本，来自复旦大学附属华山医院。
静脉血采集和获取血浆方法：同实施例2。
血浆ELISA方法：QDLIR同实施例2；Tau、Aβ40和Aβ42由杭州晶佰生物技术有限公司的三重试剂盒(货号：101995)按照其操作流程委托检测。
MCI分类方法：同实施例5。
结果与讨论
通过临床收集不同类型MCI患者、AD患者和正常随访人群的血样，我们对其进行了检测，发现QDLIR在aMCI-m类型的病人和AD病人的血样中其含量升高，与正常随访人群以及其余MCI类型相比有显著性差异，而已有的检测血样中Tau、Aβ40和Aβ42等指标，尽管部分指标(比如Aβ40)有在aMCI-m类型的病人和AD病人血中升高的趋势，但组间均未检测到显著差异(参见图6)。
通过ELISA的方法，发现aMCI-m类型的病人和AD病人血中QDLIR的含量比较高，可见QDLIR与已有辅助诊断指标Tau、Aβ40和Aβ42相比，能够在早期更明显地区分aMCI-m类型的病人、AD病人与其他类型MCI病人或正常人员。因此，QDLIR可以作为aMCI-m和AD的有效诊断标志物。
实施例7 QDLIR蛋白在患者血浆和脑脊液中的含量比较
试验材料与方法
组织标本：分别源自同一患者(脑水肿、脑膜瘤和胶质瘤各1例)的静脉血标本和脑脊液标本，均来自复旦大学附属华山医院。
静脉血采集和获取血浆方法：同实施例2。
脑脊液采集方法：患者手术中留取脑脊液。
血浆和脑脊液ELISA方法：同实施例2。
结果与讨论
血浆和脑脊液中QDLIR含量一致性很高，R 2值为0.99(参见图7)。
通过ELISA的方法，病人血中QDLIR的含量与脑脊液中的含量一致，说明血液检测到 的含量可以直接反映脑脊液中的QDLIR的含量，提示本申请所述的QDLIR同样适用于以脑脊液为样本作为有效的诊断标志物。
在本申请提及的所有文献都在本申请中引用作为参考，就如同每一篇文献被单独引用作为参考那样。此外应理解，在阅读了本申请的上述讲授内容之后，本领域技术人员可以对本申请作各种改动或修改，这些等价形式同样落于本申请所附权利要求书所限定的范围。

Claims (115)

  1. DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸，在疾病诊断和/或疾病进程评估中的用途，其中所述疾病包括认知障碍。
  2. DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸，在疾病诊断和/或疾病进程评估中的用途，其中所述疾病包括神经退行性疾病。
  3. 根据权利要求1-2中任一项所述的用途，其中所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
  4. 根据权利要求1-3中任一项所述的用途，其中所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
  5. 根据权利要求1-4中任一项所述的用途，其中所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
  6. 根据权利要求1-5中任一项所述的用途，其中所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
  7. 根据权利要求1-6中任一项所述的用途，其中所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
  8. 根据权利要求1-7中任一项所述的用途，其中所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知障碍。
  9. 根据权利要求1-8中任一项所述的用途，其中所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
  10. 根据权利要求2-9中任一项所述的用途，其中所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
  11. 根据权利要求2-10中任一项所述的用途，其中所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
  12. 根据权利要求2-11中任一项所述的用途，其中所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
  13. 根据权利要求2-12中任一项所述的用途，其中所述神经退行性疾病包括早老性痴呆。
  14. 根据权利要求2-13中任一项所述的用途，其中所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
  15. 检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质，在制备用于疾病诊断和/或疾病进程评估的产品中的用途，其中所述疾病包括认知障碍。
  16. 检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质，在制备用于疾病诊断和/或疾病进程评估的产品中的用途，其中所述疾病包括神经退行性疾病。
  17. 根据权利要求15-16中任一项所述的用途，其中所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
  18. 根据权利要求15-17中任一项所述的用途，其中所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
  19. 根据权利要求15-18中任一项所述的用途，其中所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
  20. 根据权利要求15-19中任一项所述的用途，其中所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
  21. 根据权利要求15-20中任一项所述的用途，其中所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
  22. 根据权利要求15-21中任一项所述的用途，其中所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知障碍。
  23. 根据权利要求15-22中任一项所述的用途，其中所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
  24. 根据权利要求16-23中任一项所述的用途，其中所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
  25. 根据权利要求16-24中任一项所述的用途，其中所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
  26. 根据权利要求16-25中任一项所述的用途，其中所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
  27. 根据权利要求16-26中任一项所述的用途，其中所述神经退行性疾病包括早老性痴呆。
  28. 根据权利要求16-27中任一项所述的用途，其中所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
  29. 根据权利要求15-28中任一项所述的用途，其中所述物质包括能够检测所述QDLIR，和/或编码所述QDLIR的核酸的试剂和/或设备。
  30. 根据权利要求15-29中任一项所述的用途，其中所述物质包括特异性检测所述QDLIR，和/或编码所述QDLIR的核酸的试剂。
  31. 根据权利要求15-30中任一项所述的用途，其中所述物质包括能够特异性结合QDLIR的抗体、能够特异性结合QDLIR的探针、能够特异性扩增编码QDLIR的核酸的引物、能够特异性检测编码QDLIR的核酸的基因芯片、能够特异性结合编码QDLIR的核酸的适体、和/或能够特异性结合编码QDLIR的核酸的向导RNA。
  32. 根据权利要求15-31中任一项所述的用途，其中所述产品包括诊断试剂盒。
  33. 诊断试剂盒，其包含权利要求15-32中任一项所述的物质，所述诊断盒用于诊断认知障碍，和/或评估认知障碍的疾病进程；和/或用于诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程。
  34. 根据权利要求33所述的诊断试剂盒，其包含说明书，所述说明书记载了利用所述诊断试剂盒诊断认知障碍，和/或评估认知障碍的疾病进程；和/或诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程的方法。
  35. 诊断认知障碍，和/或评估认知障碍的疾病进程的方法，其包括以下步骤：检测源自有需要的受试者的样本中，DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的含量。
  36. 诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程的方法，其包括以下步骤：检测源自有需要的受试者的样本中，DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的含量。
  37. 根据权利要求35-36中任一项所述的方法，其中所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
  38. 根据权利要求35-37中任一项所述的方法，其中所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
  39. 根据权利要求35-38中任一项所述的方法，其中所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
  40. 根据权利要求35-39中任一项所述的方法，其中所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
  41. 根据权利要求35-40中任一项所述的方法，其中所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
  42. 根据权利要求35-41中任一项所述的方法，其中所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知障碍。
  43. 根据权利要求35-42中任一项所述的方法，其中所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
  44. 根据权利要求36-43中任一项所述的方法，其中所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
  45. 根据权利要求36-44中任一项所述的方法，其中所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
  46. 根据权利要求36-45中任一项所述的方法，其中所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
  47. 根据权利要求36-46中任一项所述的方法，其中所述神经退行性疾病包括早老性痴呆。
  48. 根据权利要求36-47中任一项所述的方法，其中所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
  49. 根据权利要求35-48中任一项所述的方法，其中所述受试者包括哺乳动物。
  50. 根据权利要求35-49中任一项所述的方法，其中所述受试者包括肿瘤患者。
  51. 根据权利要求35-50中任一项所述的方法，其中所述受试者处于老龄阶段。
  52. 根据权利要求35-51中任一项所述的方法，其中所述样本包括血液样品和/或组织或细胞样品。
  53. 根据权利要求35-52中任一项所述的方法，其中所述样本包括全血、血清、血浆和/或脑脊液。
  54. 根据权利要求35-53中任一项所述的方法，其中所述方法包括：将所述受试者的样本的所述QDLIR和/或编码QDLIR的核酸的含量与正常对照值进行比较，其中所述正常对照值包括正常的受试者中所述QDLIR和/或编码QDLIR的核酸的含量。
  55. 根据权利要求54所述的方法，其中所述正常的受试者不患有所述认知障碍和/或所述神经退行性疾病。
  56. 根据权利要求54-55中任一项所述的方法，其中，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于所述正常对照值，所述受试者被诊断为患有所述认知障碍和/或所述神经退行性疾病。
  57. 根据权利要求35-56中任一项所述的方法，其中所述方法包括：将所述受试者的样本的所 述QDLIR和/或编码QDLIR的核酸的含量与早期对照值进行比较，其中所述早期对照值包括同一所述受试者之前测得的所述QDLIR和/或编码QDLIR的核酸的含量。
  58. 根据权利要求57所述的方法，其中所述受试者已确诊患有所述认知障碍和/或所述神经退行性疾病。
  59. 根据权利要求57-58中任一项所述的方法，其中，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于所述早期对照值，所述受试者被诊断为所述认知障碍和/或所述神经退行性疾病的疾病进程加剧。
  60. 根据权利要求35-59中任一项所述的方法，其还包括以下的步骤：检测源自所述受试者的样本中，与认知障碍和/或神经退行性疾病相关的标志物的含量。
  61. 根据权利要求60所述的方法，其中所述与认知障碍和/或神经退行性疾病相关的标志物包括AD7C-NTP、pTau-181、pTau-217和/或Aβ1-42。
  62. 根据权利要求35-61中任一项所述的方法，其还包括以下的步骤：观察所述受试者的脑部成像。
  63. 诊断认知障碍，和/或评估认知障碍的疾病进程的系统，其包含检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质。
  64. 诊断神经退行性疾病，和/或评估神经退行性疾病的疾病进程的系统，其包含检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的物质。
  65. 根据权利要求64所述的系统，其中所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
  66. 根据权利要求64-65中任一项所述的系统，其中所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
  67. 根据权利要求64-66中任一项所述的系统，其中所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
  68. 根据权利要求64-67中任一项所述的系统，其中所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
  69. 根据权利要求64-68中任一项所述的系统，其中所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
  70. 根据权利要求64-69中任一项所述的系统，其中所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知障碍。
  71. 根据权利要求64-70中任一项所述的系统，其中所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
  72. 根据权利要求64-71中任一项所述的系统，其中所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
  73. 根据权利要求64-72中任一项所述的系统，其中所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
  74. 根据权利要求64-73中任一项所述的系统，其中所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
  75. 根据权利要求64-74中任一项所述的系统，其中所述神经退行性疾病包括早老性痴呆。
  76. 根据权利要求64-75中任一项所述的系统，其中所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
  77. 根据权利要求64-76中任一项所述的系统，其以试剂盒的形式存在。
  78. 根据权利要求64-77中任一项所述的系统，其包含收集模块，所述收集模块能够收集源自有需要的受试者的样本。
  79. 根据权利要求78所述的系统，其中所述受试者包括哺乳动物。
  80. 根据权利要求78-79中任一项所述的系统，其中所述受试者包括肿瘤患者。
  81. 根据权利要求78-80中任一项所述的系统，其中所述受试者处于老龄阶段。
  82. 根据权利要求78-81中任一项所述的系统，其中所述样本包括血液样品和/或组织或细胞样品。
  83. 根据权利要求78-82中任一项所述的系统，其中所述样本包括全血、血清、血浆和/或脑脊液。
  84. 根据权利要求64-83中任一项所述的系统，其包含检测模块，所述检测模块能够检测所述QDLIR和/或编码所述QDLIR的核酸的含量值。
  85. 根据权利要求84所述的系统，其中所述检测模块检测所述收集模块获得的所述样本中所述QDLIR和/或编码所述QDLIR的核酸的含量值。
  86. 根据权利要求84-85中任一项所述的系统，其中所述检测模块包含能够检测所述QDLIR和/或编码所述QDLIR的核酸的含量的物质。
  87. 根据权利要求86所述的系统，其中所述物质包括能够特异性结合QDLIR的抗体、能够特异性结合QDLIR的探针、能够特异性扩增编码QDLIR的核酸的引物、能够特异性检测编 码QDLIR的核酸的基因芯片、能够特异性结合编码QDLIR的核酸的适体、和/或能够特异性结合编码QDLIR的核酸的向导RNA。
  88. 根据权利要求84-87中任一项所述的系统，所述检测模块包含能够检测DNA损伤诱导性转录物4样转录物(DDIT4L)的内含子滞留剪切产物QDLIR，和/或编码所述QDLIR的核酸的含量的仪器。
  89. 根据权利要求84-88中任一项所述的系统，其中所述检测模块包含能够检测与认知障碍和/或神经退行性疾病相关的标志物的含量的物质和/或仪器。
  90. 根据权利要求89所述的系统，其中所述与认知障碍和/或神经退行性疾病相关的标志物包括AD7C-NTP、pTau-181、pTau-217和/或Aβ1-42。
  91. 根据权利要求84-90中任一项所述的系统，其中所述检测模块输出所述QDLIR和/或编码所述QDLIR的核酸的含量值。
  92. 根据权利要求64-91中任一项所述的系统，其包含判断模块，所述判断模块能够根据所述QDLIR和/或编码所述QDLIR的核酸的含量与正常对照值和/或早期对照值，判断所述受试者的诊断结果和/或疾病进程结果；
    其中，所述正常对照值包括正常的受试者中所述QDLIR和/或编码QDLIR的核酸的含量；
    其中，所述早期对照值包括同一所述受试者之前测得的所述QDLIR和/或编码QDLIR的核酸的含量。
  93. 根据权利要求92所述的系统，其中所述正常的受试者不患有所述认知障碍和/或所述神经退行性疾病。
  94. 根据权利要求92-93中任一项所述的系统，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于所述正常对照值，所述判断模块将所述受试者诊断为患有所述认知障碍和/或所述神经退行性疾病。
  95. 根据权利要求92-94中任一项所述的系统，其中所述受试者已确诊患有所述认知障碍和/或所述神经退行性疾病。
  96. 根据权利要求92-95中任一项所述的系统，其中，当所述受试者的样本的QDLIR，和/或编码QDLIR的核酸的含量显著高于所述早期对照值，所述判断模块将所述受试者诊断为所述认知障碍和/或所述神经退行性疾病的疾病进程加剧。
  97. 根据权利要求92-96中任一项所述的系统，其中所述判断模块输出所述受试者的诊断结果。
  98. 根据权利要求64-97中任一项所述的系统，其包括显示模块，其中所述显示模块能够显示出所述判断模块的诊断结果。
  99. 根据权利要求64-98中任一项所述的系统，其包括存储模块，其中所述存储模块能够存储 所述检测模块输出的所述含量值，和/或所述判断模块输出的诊断结果。
  100. 药物筛选方法，其包括以下的步骤：检测施用候选药物后，受试者中QDLIR和/或编码QDLIR的核酸的含量的变化，其中所述药物用于预防、治疗和/或缓解认知障碍和/或神经退行性疾病。
  101. 根据权利要求100所述的方法，其中所述QDLIR包含SEQ ID NO:4所示的氨基酸序列。
  102. 根据权利要求100-101中任一项所述的方法，其中所述编码QDLIR的核酸包含SEQ ID NO:3所示的核苷酸序列。
  103. 根据权利要求100-102中任一项所述的方法，其中所述编码QDLIR的核酸包含SEQ ID NO:2所示的核苷酸序列。
  104. 根据权利要求100-103中任一项所述的方法，其中所述认知障碍包括正常衰老引起的认知障碍、lews身体痴呆(LBD)、额颞叶痴呆和/或血管性痴呆。
  105. 根据权利要求100-104中任一项所述的方法，其中所述认知障碍的诱导疾病包括阿尔兹海默症、多发性梗死型、帕金森氏症、艾滋病和/或克雅氏病(CJD)。
  106. 根据权利要求100-105中任一项所述的方法，其中所述认知障碍包括早期认知障碍(MCI)、中期认知障碍和晚期认知障碍。
  107. 根据权利要求100-106中任一项所述的方法，其中所述认知障碍包括遗忘型MCI多认知域受损(aMCI-m)。
  108. 根据权利要求100-107中任一项所述的方法，其中所述神经退行性疾病包括急性神经退行性疾病和慢性神经退行性疾病。
  109. 根据权利要求100-108中任一项所述的方法，其中所述神经退行性疾病包括由神经元死亡和胶质细胞稳态引起的神经退行性疾病、由衰老引起的神经退行性疾病、由CNS细胞功能被影响引起的神经退行性疾病、由细胞间异常通信引起的神经退行性疾病和/或由细胞运动受损引起的神经退行性疾病。
  110. 根据权利要求100-109中任一项所述的方法，其中所述神经退行性疾病包括阿尔兹海默症、帕金森氏症、多发性硬化(MS)、肌萎缩性脊髓侧索硬化症(ALS)和/或亨廷顿病(HD)。
  111. 根据权利要求100-110中任一项所述的方法，其中所述神经退行性疾病包括早老性痴呆。
  112. 根据权利要求100-111中任一项所述的方法，其中所述神经退行性疾病包括早老性痴呆早期、早老性痴呆中期和/或早老性痴呆晚期。
  113. 根据权利要求100-112中任一项所述的方法，其中，当施用所述候选药物中，受试者 中QDLIR和/或编码QDLIR的核酸的含量降低，则所述候选药物具有治疗效果。
  114. 根据权利要求100-113中任一项所述的方法，其中所述施用包括注射。
  115. 根据权利要求100-114中任一项所述的方法，其中所述药物包括小分子药物和/或生物大分子药物。
PCT/CN2022/090220 2022-04-29 2022-04-29 作为早老性痴呆诊断标志物的ddit4l剪切产物 WO2023206347A1 (zh)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111699386A (zh) * 2017-12-08 2020-09-22 神经Gx有限责任公司 用于阿尔茨海默病的同步化细胞周期基因表达测试及相关治疗方法
CN113286895A (zh) * 2018-09-05 2021-08-20 阿莫内塔诊断股份公司 用于诊断和治疗脑疾患，尤其是认知障碍的长的非编码RNA(lncRNA)

Family Cites Families (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6800455B2 (en) * 2000-03-31 2004-10-05 Scios Inc. Secreted factors
JP5858785B2 (ja) * 2009-07-29 2016-02-10 シスメックス株式会社 ヒトｉｌ−１７産生ヘルパーｔ細胞検出用マーカーおよび試薬、並びにヒトｉｌ−１７産生ヘルパーｔ細胞の検出方法
MX2019009346A (es) * 2017-02-10 2019-10-02 Genmab Bv Variantes de polipeptidos y usos de los mismos.
CN112168969B (zh) * 2020-09-29 2023-09-29 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

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111699386A (zh) * 2017-12-08 2020-09-22 神经Gx有限责任公司 用于阿尔茨海默病的同步化细胞周期基因表达测试及相关治疗方法
CN113286895A (zh) * 2018-09-05 2021-08-20 阿莫内塔诊断股份公司 用于诊断和治疗脑疾患，尤其是认知障碍的长的非编码RNA(lncRNA)

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
DATABASE Nucleotide 24 October 2000 (2000-10-24), " Homo sapiens genomic DNA, chromosome 4q22-q24, clone:2190A3, complete sequence", XP009550072, retrieved from NCBI Database accession no. AP001961.2 *
INDIRA, T.H. ET AL.: "DNA Damage Inducible Transcript 4 Gene: the Switch of the Metabolism as Potential Target in Cancer", FRONTIERS IN ONCOLOGY, vol. 8, 12 April 2018 (2018-04-12), XP055931499, DOI: 10.3389/fonc.2018.00106 *

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