Monitoring Inflammation Status

Abstract

Methods for monitoring inflammation status of a subject comprise determining levels of at least one neutrophil activation marker, or at least three markers, in urine samples taken from the subject at multiple time points, wherein increased levels of the at least one neutrophil activation marker, or at least one of the markers, in a urine sample are indicative of or predictive of an exacerbation of inflammation and/or wherein decreased levels of the at least one neutrophil activation marker, or at least one of the markers, in a urine sample following an increase are indicative or predictive of recovery from, or successful treatment of, an exacerbation of inflammation. Corresponding systems, test kits and computer programs are provided.

1 - 34 . (canceled) 
     
     
         35 . A method comprising determining levels of at least three markers in urine samples taken from a subject suffering from a respiratory disorder at multiple time points wherein at least one of the markers is selected from CRP, CC16, large elastin fragments (LEF), Tissue Inhibitor of metalloproteinase (TIMP), cystatin C, alpha-1 antitrypsin (A1AT), ICAM-1, IL-6, IL-1β, IL-8, N-formyl-Met-Leu-Phe (fMLP), IL-6 induced fibrinogen and cytokine induced beta-2-microglobulin (B2M). 
     
     
         36 . The method of  claim 35 , wherein at least one of the markers is selected from CRP, CC16 and large elastin fragments (LEF). 
     
     
         37 . The method of  claim 36 , further comprising determining levels of at least one further marker in the urine samples. 
     
     
         38 . The method according to  claim 35 , wherein at least one marker is selected from a protease activity, Neutrophil gelatinase-associated lipocalin (NGAL) which is either free or in complex, calprotectin, and myeloperoxidase (MPO). 
     
     
         39 . The method according to  claim 38 , wherein the protease activity is selected from matrix metalloproteinase (MMP) activity, HNE activity and cathepsin G activity, optionally wherein MMP activity comprises MMP9 and/or MMP8 activity. 
     
     
         40 . The method according to  claim 38 , wherein protease activity is determined by measuring cleavage of a peptide substrate. 
     
     
         41 . The method according to  claim 40 , wherein protease activity is determined by a method comprising:
 (a) bringing an indicator molecule into contact with the test sample, said indicator molecule comprising:
 (i) a cleavage region comprising at least one cleavage site, which can be cleaved by said protease if present; and 
 (ii) a capture site; 
   wherein cleavage of the at least one cleavage site produces a novel binding site;   (b) adding to the test sample binding molecules capable of binding to the novel binding site, wherein the binding molecules are incapable of binding to the indicator molecule unless and until cleavage has occurred;   (c) capturing the part of the indicator molecule containing the novel binding site at a capture zone through binding of capture molecules in the capture zone to the capture site; and   (d) detecting cleavage of the at least one cleavage site by determining binding of the binding molecules to the novel binding site of the indicator molecule captured in the capture zone.   
     
     
         42 . The method according to  claim 37 , wherein at least one marker is selected from chlorinated peptides, lactic acid, free fatty acids, an extracellular matrix breakdown product or elastin fragments/peptides. 
     
     
         43 . The method according to  claim 35 , wherein the respiratory disorder is chronic obstructive pulmonary disease (COPD) or cystic fibrosis (CF). 
     
     
         44 . The method according to  claim 35 , wherein at least two or three of the markers are neutrophil activation markers. 
     
     
         45 . The method according to  claim 35 , wherein the levels of at least one marker are determined by normalising against the levels of a reference marker. 
     
     
         46 . A system or test kit comprising:
 (a) one or more testing devices for determining levels of at least three markers in a urine sample from a subject wherein at least one of the markers is selected from CRP, CC16, large elastin fragments (LEF), Tissue Inhibitor of metalloproteinase (TIMP), cystatin C, alpha-1 antitrypsin (A1AT), ICAM-1, IL-6, IL-1β, IL-8, N-formyl-Met-Leu-Phe (fMLP), IL-6 induced fibrinogen and cytokine induced beta-2-microglobulin (B2M);   (b) a processor; and   (c) storage medium comprising a computer application that, when executed by the processor, is configured to:
 (i) access and/or calculate the determined levels of the at least three markers in the urine sample on the one or more testing devices; 
 (ii) calculate whether there is an increased or decreased level of the at least three markers in the urine sample relative to a reference level; and 
 (iii) output from the processor whether there is an increased or decreases level of the at least three markers in the urine sample relative to the reference level. 
   
     
     
         47 . The system or test kit of  claim 46 , further comprising a display for the output from the processor and/or wherein the one or more testing devices are disposable single use devices and/or wherein the one or more testing devices comprise lateral flow test strips, optionally comprising a lateral flow test strip for each marker that is determined. 
     
     
         48 . The system or test kit of  claim 46 , wherein at least one of the markers is selected from CRP, CC16 and large elastin fragments (LEF). 
     
     
         49 . The system or test kit of  claim 46 , wherein at least one marker is selected from a protease activity, Neutrophil gelatinase-associated lipocalin (NGAL) which is either free or in complex, calprotectin, and myeloperoxidase (MPO). 
     
     
         50 . The system or test kit of  claim 46 , wherein the protease activity is selected from matrix metalloproteinase (MMP) activity, HNE activity and cathepsin G activity, optionally wherein MMP activity comprises MMP9 and/or MMP8 activity. 
     
     
         51 . The system or test kit of  claim 50 , wherein the one or more testing devices comprises a testing device for measuring cleavage of a peptide substrate as an indicator of protease activity. 
     
     
         52 . The system or test kit of  claim 51 , wherein the testing device comprises:
 (a) an indicator molecule for adding to the urine sample, said indicator molecule comprising
 (i) a cleavage region comprising at least one cleavage site, which can be cleaved by said protease activity if present; and 
 (ii) a capture site; 
   wherein cleavage of the at least one cleavage site produces a novel binding site;   (b) a capture zone to receive the urine sample, wherein the capture zone comprises capture molecules capable of binding to the capture site of the indicator molecule in order to immobilise the indicator molecule including the novel binding site; and   (c) binding molecules capable of binding to the novel binding site, wherein the binding molecules are incapable of binding to the indicator molecule unless and until cleavage has occurred.   
     
     
         53 . The system or test kit of  claim 46 , wherein at least one marker is selected from chlorinated peptides, lactic acid, free fatty acids, an extracellular matrix breakdown product or elastin fragments/peptides. 
     
     
         54 . A computer application as defined in  claim 46 .