Assay for determining the type and/or status of a cell based on the epigenetic pattern and the chromatin structure

Abstract

The present invention relates to a method for identifying a specific type and/or state of a mammalian cell in a sample obtained from a mammal, comprising a) analyzing the relative amount of accessible chromatin in regions that are specific for a cell-type and/or cellular state in the genome of said cell, b) comparing said relative amount of accessible chromatin said in regions with the relative amount of accessible chromatin in regions in the genome of said cell that are unspecific for a cell-type and/or cellular state, and c) deducing the specific type and/or state of said mammalian cell in said sample based on said comparison. Preferably, said identifying further comprises a relative quantification of said specific cell type and/or state based on said comparison. The method can further comprise a diagnosis of a predisposition to a disease or a disease based on said identification. Kits and certain markers in regions of accessible chromatin in the genome are described, too.

1 . A method for identifying a specific type and/or state of a mammalian cell in a sample obtained from a mammal, comprising
 a) analyzing the relative amount of accessible chromatin in regions that are specific for a cell-type and/or cellular state in the genome of said cell,   b) comparing said relative amount of accessible chromatin in said regions with the relative amount of accessible chromatin in regions in the genome of said cell that are unspecific for a cell-type and/or cellular state,   c) optionally, normalizing the relative amount of said regions that are specific for a cell-type and/or cellular state and said regions in the genome of said cell that are unspecific for a cell-type and/or cellular state using a control plasmid, and   d) deducing the specific type and/or state of said mammalian cell in said sample based on said comparison.   
     
     
         2 . The method according to  claim 1 , wherein said identifying further comprises a relative quantification of said specific cell type and/or state based on said comparison. 
     
     
         3 . The method according to  claim 1 , further comprising a step of determining a specific cell-type and/or cellular state comprising measuring the relative amount of accessible chromatin in the genome of a cell having a known specific cell-type and/or cellular state prior to step a). 
     
     
         4 . The method according to  claim 3 , further comprising generating a knowledge base comprising information on the relative amount of accessible chromatin in the genome of cells having a known specific cell-type and/or cellular state. 
     
     
         5 . The method according to  claim 1 , wherein analysis comprises measuring the relative amount of accessible chromatin with an assay comprising DNAse I digestion, ChIP Chip®, chromatin immunoprecipitation microarray, quantitative PCR analysis based on bisulfite converted or not, selective precipitation and/or conversion of cytosines with bisulfite. 
     
     
         6 . The method according to  claim 1 , wherein said regions that are specific for a cell-type and/or cellular state in the genome of said cell are selected from regions comprising a gene selected from FOXP3, GNLY, CD3, platelet glycoprotein IX (GP9); low affinity immunoglobulin epsilon Fc receptor (FCER2); protein S100-P (S100 calcium-binding protein P); homeodomain-interacting protein kinase 3 (HIPK3); transmembrane 4 L6 family member 19 (TM4SF19); CD160 antigen precursor (Natural killer cell receptor BY55) (CD160); and LIM domain-binding protein 2 (LDB2). 
     
     
         7 . The method according to  claim 1 , wherein said regions that are unspecific for a cell-type and/or cellular state are selected from regions comprising a housekeeping gene. 
     
     
         8 . The method according to  claim 1 , wherein said cell type is selected from an immune cell; kidney cell; bone cell; neuronal cell; blood cell; lung cell; colon cell; and a precursor of any of these, excluding human embryonic stem cells. 
     
     
         9 . The method according to  claim 1 , further comprising a diagnosis of a predisposition to a disease or a disease based on said identification. 
     
     
         10 . The method of  claim 9 , wherein the disease is selected from the group consisting of immune diseases or conditions, cancer, birth defects, mental retardation, obesity, neurological disease, diabetes, and gestational diabetes. 
     
     
         11 . A method for monitoring the effect of a drug on the relative amount of a specific type and/or the state of a mammalian cell in a sample obtained from a mammal, comprising performing the method according to  claim 1  on a sample obtained from a mammal treated with said drug, and comparing the relative amount of said specific type and/or the state of said mammalian cell with an untreated sample. 
     
     
         12 . The method according to  claim 1 , wherein the sample is selected from the group consisting of blood or fractions thereof, saliva, buccal, tears, semen, urine, sweat, faecal material, skin and hair. 
     
     
         13 . The method according to  claim 11 , wherein said cell type is selected from an immune cell; kidney cell; bone cell; neuronal cell; blood cell; and a precursor of any of these, excluding human embryonic stem cells. 
     
     
         14 . The method according to  claim 11 , wherein said treatment is for a disease or condition selected from the group consisting of immune diseases or conditions, cancer, birth defects, mental retardation, obesity, neurological disease, diabetes, and gestational diabetes. 
     
     
         15 . A diagnostic kit comprising materials for performing the method according to  claim 1 . 
     
     
         16 . The method, according to  claim 7 , wherein said housekeeping gene is GAPDM. 
     
     
         17 . The method, according to  claim 8 , wherein said immune cell is selected from the group consisting of CD19+ B cells, CD3+CD8+ cytotoxic T cells, CD15+ granulocytes, CD14+ monocytes, CD56+ natural killer cells, and CD4+ helper T cells. 
     
     
         18 . The method, according to  claim 10 , wherein said cancer is selected from the group consisting of colorectal cancer, oesophageal cancer, stomach cancer, leukaemia/lymphoma, lung cancer, prostate cancer, uterine cancer, breast cancer, skin cancer, endocrine cancer, urinary cancer, pancreatic cancer, other gastrointestinal cancer, ovarian cancer, cervical cancer, head cancer, neck cancer, and adenomas. 
     
     
         19 . The method, according to  claim 13 , wherein said immune cell is selected from the group consisting of CD19+ B cells, CD3+CD8+ cytotoxic T cells, CD15+ granulocytes, CD14+ monocytes, CD56+ natural killer cells, and CD4+ helper T cells.