Compositions and Methods for in Vivo Imaging

Abstract

The present disclosure provides lipid-probe compounds, and compositions comprising the compounds. A subject lipid-probe compound is useful for various imaging applications, which are also provided.

1 . A compound of the formula:
   Q-L-X—Y
   wherein Q is a lipid;   X is an optional leaving group (e.g., O, S, or NH);   L is a cleavable linker comprising a cleavable bond that provides for release of Y or X—Y following cleavage of the linker; and   Y comprises a detectable moiety that, after release, generates a direct or indirect detectable signal.   
     
     
         2 . The compound of  claim 1 , wherein:
 Q is selected from an unsaturated fatty acid, a polyunsaturated fatty acid, a saturated fatty acid, an essential fatty acid, a trans fatty acid, a glycerolipid, a triglyceride, a diglyceride, a monoglyceride, a very long-chain fatty acid, a long-chain fatty acid, a medium-chain fatty acid, a short-chain fatty acid, a free fatty acid, a sterol, an oxisterol, a cholesterol ester, cholesterol, a bile acid, a steroid hormone, a vitamin derived fatty acid, vitamin E, vitamin K, a phospholipid, a sphingolipid, a ganglioside, a prenol lipid, a carotinoid, and a ubiquinone.   
     
     
         3 . The compound of  claim 1 , wherein:
 L comprises an enzyme substrate and is cleaved using an enzyme.   
     
     
         4 . The compound of  claim 1 , wherein:
 L is susceptible to cleavage under particular physiological conditions selected from reducing conditions, oxidizing conditions, acidic pH, and basic pH.   
     
     
         5 . The compound of  claim 1 , wherein:
 cleavage of the cleavable bond of linker L unmasks a functional group that triggers the release of Y or X—Y.   
     
     
         6 . The compound of  claim 1 , wherein:
 upon cleavage of the cleavable bond of linker L, a nucleophilic moiety is unmasked, that provides for intramolecular reaction at an electrophilic site adjacent to the leaving group X leading to the release of X—Y.   
     
     
         7 . The compound of  claim 1 , wherein:
 L is described by one of the following structures:   
       
         
           
           
               
               
           
         
         
           wherein m and n are independently 1, 2 or 3; 
           R 15 , R 26  and R 51  are independently selected from O, S and NR, where R is hydrogen or alkyl; 
           T 1  is a single bond or a linking group that is bound to Q; 
           R 31  and R 52  are independently one or more groups, each R 31  and R 52  independently selected from H, an alkyl, an aliphatic, an amino, an aryl, an acyl, an alkoxy, an aryloxy, an acyloxy, a carbonyl, a cyano, a halogen, hydroxyl, a heterocyclic group, a nitro, a thio, a sulfinyl, a sulfonyl, and a trifluoromethyl; and 
           R 11 , R 12 , R 13 , R 14 , R 21 , R 22 , R 23 , R 24 , R 25 , R 32 , R 33 , R 41 , R 42 , R 43 , R 44 , R 45 , R 46 , R 53  and R 54  are independently selected from hydrogen, an alkyl, an aryl, a heterocyclic group, and an amino acid sidechain group. 
         
       
     
     
         8 . The compound of  claim 1 , wherein:
 Y is selected from a luciferin, an aminoluciferin, coelenterazine, a modified coelenterazine, a coelenterazine analog, a membrane permeant coelenterazine analog, a dihydroluciferin, a luciferin 6′ methylether, a luciferin 6′ chloroethylether.   
     
     
         9 . The compound of  claim 1 , wherein:
 Y comprises:   a) an optionally substituted luciferin moiety, where luciferin moiety has one of the following structures:   
       
         
           
           
               
               
           
         
         
           wherein R 3  is hydrogen, alkyl or substituted alkyl; 
         
         b) an optionally substituted coelenterazine moiety, where coelenterazine has the structure: 
       
       
         
           
           
               
               
           
         
         
           wherein R 3  and R 4  are independently selected from hydrogen, an acyl, an acyloxy, and an acylamino; 
         
         c) the following structure: 
       
       
         
           
           
               
               
           
         
         
           wherein R 1 , R 2 , R 3 , and R 4  are independently H, alkyl, heteroalkyl, aryl, or combinations thereof, wherein the structure is attached to the linker via any one of R 1 , R 2 , R 3  and R 4 ; 
         
         d) an optionally substituted membrane-permeant coelenterazine moiety of one of the following structures: 
       
       
         
           
           
               
               
           
         
         
           wherein R 4  and R 5  are independently an alkyl, an aryl, an aralkyl, an alkoxy, or a heterocyclic group; 
         
         e) an optionally substituted membrane-permeant coelenterazine moiety of one of the following structures: 
       
       
         
           
           
               
               
           
         
         
           wherein p is an integer of about 1 to 20; 
         
         f) an optionally substituted membrane-permeant coelenterazine moiety of one of the following structures: 
       
       
         
           
           
               
               
           
         
         
           wherein R 1 , R 2 , and R 3  are independently an alkyl, an alkenyl, or an aralkyl; 
         
         g) an optionally substituted membrane-permeant coelenterazine moiety of one of the following structures: 
       
       
         
           
           
               
               
           
         
         
           wherein r is an integer of about 1 to 20; 
         
         h) an optionally substituted membrane-permeant coelenterazine moiety of one of the following structures: 
       
       
         
           
           
               
               
           
         
         
           wherein r is an integer of about 1 to 20; and 
           R 6  is an alkyl, an aryl, an aralkyl, or an alkoxyalkyl; 
         
         i) one of the following structures: 
       
       
         
           
           
               
               
           
         
         
           wherein R 2  is N or CH; 
           R 3  is hydrogen, a halo, an alkyl, an alkoxy, an amino, —CH 2 N═R, or CH 2 NRR′, wherein R is alkyl and R′ is alkyl; and 
           R 4  is hydrogen, alkyl or substituted alkyl; 
         
         j) one of the following structures: 
       
       
         
           
           
               
               
           
         
         
           wherein R 2  is N or CH; and R 3  is hydrogen, alkyl or substituted alkyl; 
         
         k) one of the following structures: 
       
       
         
           
           
               
               
           
         
         
           wherein R 2  is O or S; 
           R 3  is hydrogen, a halo, an alkyl, an alkoxy, an amino, —CH 2 N═R, or CH 2 NRR′, wherein R is alkyl and R′ is alkyl; and 
           R 4  is hydrogen, alkyl or substituted alkyl; or 
         
         l) one of the following structures: 
       
       
         
           
           
               
               
           
         
         
           wherein R 2  is N or CH; 
           R 3  is hydrogen, a halo, an alkyl, an alkoxy, an amino, —CH 2 N═R, or CH 2 NRR′, wherein R is alkyl and R′ is alkyl; 
           and R 4  is hydrogen, alkyl or substituted alkyl. 
         
       
     
     
         10 . The compound of  claim 8 , wherein:
 release of Y produces a luciferin of one of the following structures:   
       
         
           
           
               
               
           
         
         wherein R 3  is hydrogen, alkyl or substituted alkyl; and R 4  is hydrogen, alkyl, substituted alkyl or alkoxy. 
       
     
     
         11 .- 21 . (canceled) 
     
     
         22 . The compound of  claim 1 , wherein:
 Y comprises;   a) a contrast agent or a radioisotope suitable for use in imaging;   b) a detectable moiety that comprises  123 I (iodine),  18 F (fluorine),  99 Tc (technetium),  111 In (indium), and  67 Ga (gallium), gadolinium (Gd), dysprosium, or iron;   c) a chelating ligand for a gadolinium ion;   d) a chelating ligand of the structure:   
       
         
           
           
               
               
           
         
         e) a PET or SPECT radiotracer; or 
         f) a fluorophore. 
       
     
     
         23 .- 27 . (canceled) 
     
     
         28 . The compound of  claim 1 , wherein:
 L is T 1 -Z-T 2  such that the compound is of the structure:
   Q-T 1 -Z-T 2 -X—Y
 
   wherein T 1  and T 2  are independently a single bond or a linking group; and   Z comprises a cleavable bond that after cleavage unmasks a functional group that provides for release of Y or X—Y.   
     
     
         29 . The compound of  claim 28 , wherein:
 T 2  comprises an electrophilic center adjacent to X; and   the functional group is a nucleophilic group that provides for intramolecular reaction at the electrophilic center to release X—Y.   
     
     
         30 . The compound of  claim 28 , wherein the cleavable bond is part of the T 1 -Z 1 -T 2  backbone. 
     
     
         31 . The compound of  claim 28 , wherein the cleavable bond is NOT part of the T 1 -Z 1 -T 2  backbone. 
     
     
         32 . The compound of  claim 28 , wherein:
 cleavage of the cleavable bond provides for spontaneous release of X—Y, via electron pair donation.   
     
     
         33 . The compound of  claim 28 , wherein the compound is of the structure:
 (a)   
       
         
           
           
               
               
           
         
         
           where m is 1, 2 or 3; 
           R 15  is selected from O, S and NH; and 
           R 11 , R 12 , R 13  and R 14  are independently selected from hydrogen, an alkyl, an aryl and a heterocyclic group; 
         
         (b) 
       
       
         
           
           
               
               
           
         
         
           wherein n is 1, 2 or 3; 
           R 26  is selected from O, S and NH; and 
           R 21 , R 22 , R 23 , R 24  and R 25  are independently selected from hydrogen, an alkyl, an aryl and a heterocycle; 
         
         c) 
       
       
         
           
           
               
               
           
         
         
           wherein R 31  is one or more groups, each R 31  independently selected from H, an alkyl, an aliphatic, an amino, an aryl, an acyl, an alkoxy, an aryloxy, an acyloxy, a carbonyl, a cyano, a halogen, hydroxyl, a heterocyclic group, a nitro, a thio, a sulfinyl, a sulfonyl, and a trifluoromethyl; and 
           R 32  and R 33  are independently selected from hydrogen, an alkyl, an aryl and a heterocycle; 
         
         d) 
       
       
         
           
           
               
               
           
         
         
           wherein R 41  and R 44  are independently selected from hydrogen, an alkyl, an aryl and a heterocycle; and 
           R 42 , R 43 , R 45  and R 46  are independently selected from hydrogen an alkyl, an aryl a heterocyclic group and an amino acid sidechain; or 
         
         e) 
       
       
         
           
           
               
               
           
         
         
           wherein R 51  is O, S or NH; 
           R 52  is one or more groups, each R 52  independently selected from H, an alkyl, an aliphatic, an amino, an aryl, an acyl, an alkoxy, an aryloxy, an acyloxy, a carbonyl, a cyano, a halogen, hydroxyl, a heterocyclic group, a nitro, a thio, a sulfinyl, a sulfonyl, and a trifluoromethyl; and 
           R 53  and R 54  are independently selected from hydrogen, an alkyl, an aryl, and a heterocyclic group. 
         
       
     
     
         34 . The compound of  claim 33 , wherein the compound is of the structure (a), and wherein:
 m is 2;   R 11 , R 12 , R 13  and R 14  are hydrogen;   R 15  is O; and   T 1  is a single bond connecting Q and S.   
     
     
         35 . The compound of  claim 33 , wherein the compound is of the structure (a), and wherein:
 Q is a saturated fatty acid comprising 20 carbons or less; and   X—Y is a luciferin.   
     
     
         36 . (canceled) 
     
     
         37 . The compound of  claim 33 , wherein the compound is of the structure (b), and wherein:
 n is 1;   R 21  is methyl;   R 22 , R 23 , R 24  and R 25  are hydrogen; and   R 26  is O or NCH 3 .   
     
     
         38 . The compound of  claim 33 , wherein the compound is of the structure (b), and wherein
 Q is a saturated fatty acid comprising 20 carbons or less; and   X—Y is a luciferin.   
     
     
         39 . (canceled) 
     
     
         40 . The compound of  claim 33 , wherein the compound is of the structure (c), and wherein:
 R 31 , R 32  and R 33  are hydrogen.   
     
     
         41 . The compound of  claim 33 , wherein the compound is of the structure (c), and wherein:
 Q is a saturated fatty acid comprising 20 carbons or less; and   X—Y is a luciferin.   
     
     
         42 . (canceled) 
     
     
         43 . The compound of  claim 33 , wherein the compound is of the structure (d), and wherein:
 R 41  and R 44  are hydrogen; and   R 42 , R 43 , R 45  and R 46  are independently selected from hydrogen and an amino acid sidechain, wherein at least one of R 42  and R 43  are hydrogen, and at least one of R 45  and R 46  are hydrogen.   
     
     
         44 . The compound of  claim 33 , wherein the compound is of the structure (d), and wherein:
 Q is a saturated fatty acid comprising 20 carbons or less; and   X—Y is a luciferin.   
     
     
         45 . (canceled) 
     
     
         46 . The compound of  claim 33 , wherein the compound is of the structure (e), and wherein:
 R 51  is NH or O; and   R 52 , R 53  and R 54  are hydrogen.   
     
     
         47 . The compound of  claim 33 , wherein the compound is of the structure (e), and wherein:
 Q is a saturated fatty acid comprising 20 carbons or less; and   X—Y is luciferin.   
     
     
         48 . The compound of  claim 1 , wherein the compound is of one of the following structures: 
       
         
           
           
               
               
           
         
       
     
     
         49 . A method of detecting uptake of a lipid into a living cell in vitro, or into a living cell of multicellular organism in vivo, the method comprising:
 a) contacting the in vitro cell, or the multicellular organism, with a compound of  claim 1 ; and   b) detecting a signal produced following uptake of the compound into the cell.   
     
     
         50 . The method of  claim 49 , wherein:
 the cell or the multicellular organism comprises an enzyme that cleaves the cleavable linker.   
     
     
         51 . The method of  claim 49 , wherein:
 the cell or the multicellular organism comprises physiological conditions sufficient to cleave the cleavable linker L.   
     
     
         52 . The method of  claim 49 , wherein the cell:
 a) is in vitro;   b) is in vivo;   c) has been exposed to an internal or external stimulus;   d) is diseased;   e) is a mammalian cell.   
     
     
         53 .- 56 . (canceled) 
     
     
         57 . The method of  claim 49 , wherein the multicellular organism is a mammal. 
     
     
         58 . A method of detecting fatty acid uptake into a living cell in vitro, or into a living cell in a multicellular organism in vivo, the method comprising:
 a) contacting the cell or the multicellular organism with a compound of  claim 1 ; and   b) detecting a signal produced by the compound upon uptake into the cell.   
     
     
         59 . The method of  claim 49 , wherein
 the detecting comprises imaging the signal using magnetic resonance imaging.   
     
     
         60 . A method of identifying an agent that modulates lipid uptake into a living cell in vitro, or into a living cell in a multicellular organism in vivo, the method comprising:
 contacting the living cell in vitro, or administering to the multicellular organism, a compound of  claim 1  and a test agent; and   determining whether said test agent modulates uptake of the compound of  claim 1  into the living cell in vitro, or into the living cell in the multicellular organism in vivo.   
     
     
         61 . The method of  claim 60 , where the multicellular organism is a transgenic non-human mammal comprising a transgene integrated into the genome of the mammal, wherein the transgene comprises a nucleotide sequence encoding a luciferase.