Demethylated and/or oxidized membrane DNA

Abstract

A process for the preparation of oxidized and/or demethylated antigens comprising the steps of treating a cell with a stress factor selected from the group consisting of UV-radiation, oxidizing reagents, heavy metal salts, drugs, nucleoside and nucleotide analogs, and enzyme inhibitors lyses of the cell to give a cell lysate purification of oxidized and/or demethylated antigens from the cell lysate.

1 . A process for the preparation of oxidized and/or demethylated membrane DNA (mDNA) comprising the steps of 
 treating a cell with a stress factor selected from the group consisting of UV-irradiation, oxidizing reagents, heavy metal salts, drugs, nucleoside and nucleotide analogs, enzyme inhibitors and pH-shift    lyses of the cell to give a cell lysate    purification of oxidized and/or demethylated mDNA from the cell lysate.    
     
     
         2 . The process of  claim 1 , wherein the stressfactor is selected from dichromate, hydrogen peroxide, permanganate, quinidine, D-pencillamine, hydralazine, procainimide, RNA/DNA metabolites, nucleoside and nucleotide analogs such as 5-aza-cytdine, 5-aza-2′-deoxycytidine, inhibitors of DNA methylases, inhibitors of histone deacetylases such as butyrates or trichostatin A, inhibitors of histone arginine and histone lysine methyltransferases, inhibitors of protein kinases such as staurosporine or calphostin C or K-252a or H-89, activators of protein kinases such as phorbol esters or bryostatin 1, inhibitors of protein phosphatases such as calyculin A or okadaic acid, inhibitors of poly(ADP-ribose) polymerase such as 3-aminobenzamide or m-Iodobenzylguanidine hemisulfate, inhibitors of ubiquitin conjugating enzymes such as methylated ubiquitin, inhibitors of ubiquitin C-terminal hydrolase such as ubiquitin aldehyde, inhibitors of enzymes involved in correction of replicational errors and/or spontaneous DNA damage such as cadmium ions, inhibitors of N-glycosylation such as tunicamycin, inhibitors of 20S or 26S proteasomes such as lactacystin, inhibitors of farnesylation such as alpha-hydroxy-farnesylphosphonic acid, inhibitors of geranylgeranylation, inhibitors of protein methylation such as ebelactone B, inhibitors of catalase, inhibitors of superoxide dismutase, inhibitors of glutathione peroxidase, or combinations thereof.  
     
     
         3 . The process of  claim 1  wherein the oxidized and/or demethylated mDNA is associated with nucleic acid binding proteins.  
     
     
         4 . Oxidized and/or demethylated membrane DNA (mDNA) obtainable according to the method of any one of  claims 1  to  3 .  
     
     
         5 . mDNA according to  claim 4  associated with DNA-binding proteins, especially histones.  
     
     
         6 . mDNA according to  claim 5 , wherein the DNA-binding proteins are hypo- or hyperacetylated and/or hypo- or hyperphosphorylated, and/or hypo- or hypermethylated, and/or hypo- or hyper-ubiquitinated, and/or hypo- or hyper-(poly-ADP-ribosylated), and/or hypo- or hyperglycosylated or combinations thereof.  
     
     
         7 . Cells comprising mDNA according to any one of  claims 4  to  6 .  
     
     
         8 . A process for preparing cells according to  claim 7  comprising the step of treating a cell with a stress factor selected from the group consisting of UV-irradiation, oxidizing reagents, heavy metal salts, drugs, nucleoside and nucleotide analogs, enzyme inhibitors and pH-shift.  
     
     
         9 . A diagnostic agent comprising the mDNA according to any one the  claims 4  to  6 , fragments of the mDNA according to any one of  claims 4  to  6  or the cells according to  7 .  
     
     
         10 . A process for the in-vitro detection of a disease comprising the steps of 
 bringing a biological fluid of an animal containing antibodies into contact with the diagnostic agent of  claim 9     measuring binding of said antibodies with said diagnostic agent    wherein binding indicates a disease of the animal.    
     
     
         11 . The process of  claim 10 , wherein the diagnostic agent are cells and the cells are attached to a solid support.  
     
     
         12 . A process for the in-vitro detection of a disease comprising the steps of bringing a biological sample of an animal containing cells into contact with ligands especially antibodies or DNA-binding proteins such as formamidopyrimidine DNA-glycosylase directed against the oxidized and/or demethylated mDNA of  claim 4   measuring binding of said ligands, antibodies or DNA-binding proteins with said cells    wherein binding indicates a disease of the animal.    
     
     
         13 . A process according to  claim 10  or  12 , wherein the disease is an autoimmune disease.  
     
     
         14 . Use of the diagnostic agent of  claim 9  to detect binding of antibodies to mDNA.  
     
     
         15 . Use according to  claim 14 , wherein binding of antibodies from a animal indicates an autoimmune disease of the animal.  
     
     
         16 . A method for screening drugs by measuring the binding of antibodies to the mDNA according to any one the  claims 4  to  6  or the cells according to  7  in the presence of a drug.  
     
     
         17 . A pharmaceutical formulation comprising the mDNA according to any one the  claims 4  to  6  or the cells according to  7 .  
     
     
         18 . The pharmaceutical formulation of  claim 17  in form of a vaccine.  
     
     
         19 . Use of the pharmaceutical formulation of  claim 10  for treating autoimmune diseases in an animal.