Trapping reagents for reactive metabolites screening
View Patent ↗The present invention provides compounds of Formula (I) and (II): (I) (II) wherein R 1 , R 2 , R 4 , R 5 , R 6 , X and n are as defined herein, and wherein R 3 is hydrogen or a sulfur protecting group. Compounds of Formula (I) and (II), wherein R is hydrogen, may be useful in methods for detecting a reactive metabolite in a sample, e.g., wherein the metabolite is generated from the metabolism of a test compound, and wherein the metabolite and the compound of Formula (I) or (II) react to form a detectable adduct, e.g., detectable by mass spectrometry.
1. A method for detecting a metabolite in a sample, the method comprising:
contacting a sample comprising a metabolite and a compound of Formula (I) or (II):
wherein the metabolite and the compound of Formula (I) or (II) react to form an adduct; and detecting the adduct;
wherein:
each instance of R 1 and R 2 is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, —C(═O)R A , —C(═O)OR A , —C(═O)N(R A ) 2 , or an amino protecting group, or R 1 and R 2 are joined to form a substituted or unsubstituted heterocyclic ring;
each instance of R A is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, an oxygen protecting group when attached to an oxygen atom, or a nitrogen protecting group when attached to a nitrogen atom, or two R A groups are joined to form a substituted or unsubstituted heterocyclic ring;
R 3 is hydrogen;
R 5 and R 6 are independently selected from —OR B , —N(R B ) 2 , and a group of Formula (i):
provided at least one of R 5 and R 6 is a group of Formula (i);
each instance of R B is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, an oxygen protecting group when attached to an oxygen atom, or a nitrogen protecting group when attached to a nitrogen atom, or two R B groups are joined to form a substituted or unsubstituted heterocyclic ring;
each instance of R 4 is independently substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, or an amino protecting group, or two R 4 groups are joined to form a substituted or unsubstituted heterocyclic ring;
n is 1, 2, 3, 4, 5, or 6; and
X − is a counter anion.
2. A method for detecting a metabolite in a sample, the method comprising:
contacting a test compound, an enzyme system, and a compound of Formula (I) or (II):
wherein the test compound is metabolized by the enzyme system to provide a metabolite; and the metabolite reacts with a compound of Formula (I) or (II) to form an adduct; and detecting the adduct;
wherein:
each instance of R 1 and R 2 is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, —C(═O)R A , —C(═O)OR A , —C(═O)N(R A ) 2 , or an amino protecting group, or R 1 and R 2 are joined to form a substituted or unsubstituted heterocyclic ring;
each instance of R A is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, an oxygen protecting group when attached to an oxygen atom, or a nitrogen protecting group when attached to a nitrogen atom, or two R A groups are joined to form a substituted or unsubstituted heterocyclic ring;
R 3 is hydrogen;
R 5 and R 6 are independently selected from —OR B , —N(R B ) 2 , and a group of formula (i):
provided at least one of R 5 and R 6 is a group of formula (i);
each instance of R B is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, an oxygen protecting group when attached to an oxygen atom, or a nitrogen protecting group when attached to a nitrogen atom, or two R B groups are joined to form a substituted or unsubstituted heterocyclic ring;
each instance of R 4 is independently substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, or an amino protecting group, or two R 4 groups are joined to form a substituted or unsubstituted heterocyclic ring;
n is 1, 2, 3, 4, 5, or 6; and
X − is a counter anion.
3. The method of claim 1 , wherein the sample further comprises an enzyme system, and wherein the enzyme system is a P450 microsomal enzyme system.
4. The method of claim 1 , wherein the adduct formation is initiated by addition of an NADPH-generating system or NADPH.
5. The method of claim 1 , wherein the adduct is detected by mass spectrometry.
6. The method of claim 1 , wherein the sample comprises a metabolite and a compound of Formula (I):
wherein:
each instance of R 1 and R 2 is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, —C(═O)R A , —C(═O)OR A , —C(═O)N(R A ) 2 , or an amino protecting group, or R 1 and R 2 are joined to form a substituted or unsubstituted heterocyclic ring;
each instance of R A is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, an oxygen protecting group when attached to an oxygen atom, or a nitrogen protecting group when attached to a nitrogen atom, or two R A groups are joined to form a substituted or unsubstituted heterocyclic ring;
R 3 is hydrogen;
each instance of R 4 is independently substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, or an amino protecting group, or two R 4 groups are joined to form a substituted or unsubstituted heterocyclic ring;
n is 1, 2, 3, 4, 5, or 6; and
X − is a counteranion.
7. The method of claim 1 , wherein the sample comprises a metabolite and a compound of Formula (II):
wherein:
each instance of R 1 and R 2 is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, —C(═O)R A , —C(═O)OR A , —C(═O)N(R A ) 2 , or an amino protecting group, or R 1 and R 2 are joined to form a substituted or unsubstituted heterocyclic ring;
each instance of R A is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, an oxygen protecting group when attached to an oxygen atom, or a nitrogen protecting group when attached to a nitrogen atom, or two R A groups are joined to form a substituted or unsubstituted heterocyclic ring;
R 3 is hydrogen;
R 5 and R 6 are independently selected from —OR B , —N(R B ) 2 , and a group of formula (i):
provided at least one of R 5 and R 6 is a group of formula (i);
each instance of R B is independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, an oxygen protecting group when attached to an oxygen atom, or a nitrogen protecting group when attached to a nitrogen atom, or two R B groups are joined to form a substituted or unsubstituted heterocyclic ring;
each instance of R 4 is independently substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, or an amino protecting group, or two R 4 groups are joined to form a substituted or unsubstituted heterocyclic ring;
n is 1, 2, 3, 4, 5, or 6; and
X − is a counteranion.
8. The method of claim 6 , wherein R 1 is hydrogen, and R 2 is hydrogen, —C(═O)R A , —C(═O)OR A , —C(═O)N(R A ) 2 , or an amino protecting group.
9. The method of claim 8 , wherein R 1 is hydrogen and R 2 is —C(═O)R A , —C(═O)OR A , or —C(═O)NHR A , wherein R A is a group of formula:
wherein p is 0, 1, or 2; m is 1, 2, 3, 4, or 5; and R 7 is halogen.
10. The method of claim 9 , wherein R 7 is bromo or fluoro.
11. The method of claim 7 , wherein R B is a group of formula:
wherein p is 0, 1, or 2; m is 1, 2, 3, 4, or 5; and R 7 is halogen.
12. The method of claim 11 , wherein R 7 is bromo or fluoro.
13. The method of claim 6 , wherein R 3 is hydrogen.
14. The method of claim 6 , wherein each instance of R 4 is independently substituted or unsubstituted alkyl.
15. The method of claim 6 , wherein n is 2.
16. The method of claim 6 , wherein X − is a chloride counteranion.
17. The method of claim 6 , wherein the compound of Formula (I) is of Formula (I-c):
18. The method of claim 6 , wherein the compound of Formula (I) is selected from the group consisting of:
19. The method of claim 7 , wherein the compound of Formula (II) is of Formula (II-d) or (II-g):
20. The method of claim 7 , wherein the compound of Formula (II) is selected from the group consisting of: