Methods and compositions relating to genotoxin colibactin

Abstract

The technology described herein is directed to compounds which are substrates for ClbP, acting as fluorescent probes for ClbP activity. Further provided herein are methods for measuring ClbP activity, screening for ClbP inhibitors, detecting colibactin, and/or diagnosing cancer, which utilize the substrate compounds.

What is claimed herein is: 
     
         1 . A composition comprising a compound of Structure I: 
       
         
           
           
               
               
           
         
       
       wherein R is alkyl, alkenyl, alkynyl, acyl, aryl, heteroaryl, cyclyl, heterocyclyl, amino, alkylamino, dialkylamino, aminoalkyl, acylamino, hydroxyl, or alkoxy, each of which can be optionally substituted with one, two, three, four or more independently selected substituents 
     
     
         2 . The composition of  claim 1 , wherein R is a hydrophobic acyl. 
     
     
         3 . The composition of  claim 1 , wherein R comprises a carbon chain of at least 4 carbons in length. 
     
     
         4 . The composition of  claim 1 , wherein R comprises at least 4 carbon atoms. 
     
     
         5 . The composition of  claim 1 , wherein R comprises at least 9 carbon atoms. 
     
     
         6 . The composition of  claim 1 , wherein R is selected from:
 C 13 H 27 ; (4-Ph)-C 3 H 6 , C 3 H 7 , CH 3 , and (8-NH 2 )C 7 H 15 .   
     
     
         7 . The composition of  claim 1 , wherein R comprises: 
       
         
           
           
               
               
           
         
       
       wherein R 1  is selected from alkyl, alkenyl, alkynyl, aryl, heteroaryl, cyclyl, heterocyclyl or aminoalkyl, each of which can be optionally substituted with one, two, three or more independently selected substituents. 
     
     
         8 . The composition of  claim 7 , wherein R 1  is a C 1 -C 18 alkyl, optionally substituted with one substituent. 
     
     
         9 . The composition of  claim 7 , wherein R 1  is selected from:
 C 13 H 27 ; (4-Ph)-C 3 H 6 , C 3 H 7 , CH 3 , and (8-NH 2 )C 7 H 15 .   
     
     
         10 . The composition of  claim 7 , wherein R is selected from: 
       
         
           
           
               
               
           
         
       
     
     
         11 . A method of measuring ClbP activity in a sample, the method comprising:
 contacting the sample with a composition of  claim 1 ; and   measuring fluorescence;   wherein increased fluorescence relative to a reference sample indicates an increased level of ClbP activity relative to the reference sample.   
     
     
         12 . The method of  claim 11 , wherein an increased level of ClbP activity indicates an increased level or metabolic activity of pks +    E. coli.    
     
     
         13 . A method of diagnosing cancer, or detecting the risk of cancer in a subject, the method comprising:
 contacting a sample obtained from the subject with a composition of  claim 1 ; and measuring fluorescence;   wherein increased fluorescence relative to a reference indicates increased ClbP activity, the presence of cancer, and/or an increased risk of cancer.   
     
     
         14 . The method of  claim 13 , wherein the cancer is colorectal cancer. 
     
     
         15 . The method of  claim 12 , wherein the reference is the level of fluorescence in the sample prior to contacting it with the composition. 
     
     
         16 . The method of any  claim 13 , wherein the sample comprises bacterial cells or bacterial cell lysates. 
     
     
         17 . The method of  claim 16 , wherein the presence of fluorescence indicates the bacterial cells comprise colibactin biosynthetic machinery/pathways. 
     
     
         18 . The method of  claim 16 , wherein the presence of fluorescence indicates the bacterial cells comprise pks +    E. coli.