Diagnostic methods for asthma

Abstract

Screening methods for asthma and analogous diseases in which activated eosinophils are found at the disease site are provided. The methods involle assaying for the presence of brominated tyrosine species in a bodily sample which has been obtained from a test subject. The brominated tyrosine species are either free in the sample or protein bound. In one embodiment, the assay involve measuring the amount of a brominated tyrosine species, particularly 3-bromotyrosine, 3,5-dibromotyrosine, or combinations thereof (referred to hereinafter collectively as the “diagnostic marker”) in a bodily sample from the test subject. In another embodiment for determining the prognosis of asthma in a test subject, the concentration or content of the diagnostic marker is determined in bodily samples taken from the test subject over successive time intervals. The concentrations are compared to determine the prognosis of the asthma. In another embodiment of the invention for monitoring the response of the test subject to treatment with an anti-asthmatic drug, the concentration or content of the diagnostic marker is measured in bodily samples obtained from the test subject before and after such treatment. The present invention also relates to a diagnostic kit and to a diagnostic reagent for diagnosing asthma and analogous diseases which are associated with activated eosinophils.

What is claimed is:  
     
         1 . A method for diagnosing diseases associated with activated eosinophils in a subject comprising: 
 assaying for the presence of a brominated tyrosine species in a test sample of a body fluid or tissue obtained from the subject.    
     
     
         2 . The method of  claim 1  in which the test sample is selected from the group consisting of bronchoalveolar lavage, serum, plasma, urine, tissue biopsy and sputum, induced sputum, blood, pericardial fluid, pleural fluid, cerebrospinal fluid, stool, or broncho-alveolar lavage; or cells containing metabolic products of activated eosinophils and phagocytes.  
     
     
         3 . The method of  claim 1  wherein the brominated tyrosine species is selected from the group consisting of 3-bromotyrosine, 3,5-dibromotyrosine, and combinations thereof.  
     
     
         4 . The method of  claim 1  wherein the disease is asthma and the assay comprises measuring the amount of the brominated species in the test subject.  
     
     
         5 . The method of  claim 1  wherein the disease is asthma and the assay comprises determining the concentration or content of the brominated species in a test subject.  
     
     
         6 . The method of  claim 5  further comprising the step of comparing the concentration or content of the brominated species in the test sample to the concentration or content of the brominated species in corresponding samples from normal subjects lacking asthma.  
     
     
         7 . The method of  claim 4  wherein the concentration or content of the brominated species in the test sample is compared to a baseline concentration or content of the brominated species in a corresponding sample from the test subject.  
     
     
         8 . The method of  claim 1  wherein the disease is asthma and wherein the assay comprises determining the concentration or content of the brominated species in a plurality of test samples obtained at successive time intervals from the subject.  
     
     
         9 . The method of  claim 1  wherein the disease is asthma and wherein the assay comprises determining the concentration or content of the brominated species in a test sample obtained from the subject at a time prior to treatment with an anti-asthmatic drug and determining the concentration or content of the brominated species in a corresponding test sample obtained from the subject at a time following treatment with an anti-asthmatic agent.  
     
     
         10 . The method of  claim 1  wherein the presence of the brominated species in the test sample is assayed by a technique selected from mass spectrometric analysis, HPLC with electrochemical detection, HPLC with fluorescence detection following pre- or post column derivatization, Capillary Electrophoresis, and high resolution NMR spectroscopy.  
     
     
         11 . The method of  claim 1  wherein the presence of the brominated species in the sample is assayed by contacting the sample with an anti-bromotyrosine monoclonal antibody and assaying for the formation of an antigen-antibody complex between said antibody and a protein in said sample.  
     
     
         12 . A diagnostic kit for diagnosing diseases in which activated eosinophils are present at the disease site comprising an antibody reactive with a peptide bound brominated tyrosine species.  
     
     
         13 . The diagnostic kit of  claim 11  wherein said antibody is a monoclonal antibody reactive with proteins containing a bromotyrosine selected from the group consisting of 3-bromotyrosine, 3,5-dibromotyrosine, and combinations thereof.  
     
     
         14 . The diagnostic kit of  claim 12  further comprising a synthetic peptide or a synthetic protein containing bromotyrosine.  
     
     
         15 . A reagent for diagnosing diseases in which activated eosinophils are present at the disease site said reagent selected from the group consisting 3-bromotyrosine, 3,5-dibromotyrosine, a peptide containing 3-bromotyrosine, a peptide containing 3,5-dibromotyrosine, and combinations thereof.  
     
     
         16 . The reagent of  claim 14  wherein said reagent is a heavy isotope labeled 3-bromotyrosine or a heavy isotope labeled 3,5-dibromotyrosine. .  
     
     
         17 . The reagent of  claim 16  wherein said reagent is labeled with  13 C 6 ,  2 H 4 , or  13 C 9  and  15 N 1 .  
     
     
         18 . The method of  claim 1  in which the presence the brominated tyrosine species is detected by an immunoassay which employs antibodies to 3-bromotyrosine or 3,5 di-bromotyrosine.  
     
     
         19 . The method of  claim 1  wherein the disease is associated with the presence of increased concentrations of 3-bromotyrosine or 3,5-dibromotryosine in samples from subjects having the disease relative to samples from normal subjects lacking the disease.  
     
     
         20 . A method for diagnosing diseases associated with protein damage by brominating oxidants in a subject comprising: 
 assaying for the presence of a brominated tyrosine species in a test sample of a body fluid or tissue obtained from the subject, wherein the brominated tyrosine species is selected from the group consisting of 3-bromotyrosine, 3,5-dibromotyrosine, and combinations thereof.