Reactive labelling compounds and uses thereof

Patent metadata

Field	Value
Publication number	US-9759726-B2
Application number	US-201514698660-A
Country	US
Kind code	B2
Filing date	Apr 28, 2015
Priority date	Mar 27, 2014
Publication date	Sep 12, 2017
Grant date	Sep 12, 2017

Abstract

Provided are azido-BODIPY compounds of formula (I), cyclooctyne-based fluorogenic probes of formula (IV), and activity-based probes of formula (VI). These compounds undergo azide-alkyne cycloadditions (AAC) with to form triazolyl products. The provided compounds are useful for detection and imaging of alkyne-, or azide-containing molecules. Methods for detection and imaging biomolecules using compounds of the present disclosure are disclosed.

First claim

We claim:

1. A compound of Formula (I):

or a pharmaceutically acceptable salt thereof,
wherein:
B is boron;
each instance of G 1 , G 2 ,G 3 , G 5 , G 6 , G 7 and G 8 is independently hydrogen, optionally substituted C 1-6 alkyl, optionally substituted C 1-6 alkenyl, optionally substituted C 1-6 alkynyl, optionally substituted aryl, optionally substituted acyl, —OR A , —CH 2 OR A , —OC(O)R A , —SR A , —N(R B ) 2 , —N(R A )C(O)R A , —C(O)N(R B ) 2 , —CN, —NO 2 , —C(O)R A , —C(O)OR A , —S(O)R A , —SO 2 R A , —SO 3 R A , —SO 2 N(R B ) 2 , and —NHSO 2 R B ;
each R A is independently selected from hydrogen, optionally substituted C 1 -C 6 alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted heterocyclyl, and optionally substituted aryl;
each R B is independently selected from hydrogen, optionally substituted C 1 -C 6 alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted heterocyclyl, and optionally substituted aryl, or two R B taken together with the intervening nitrogen form a heterocycle;
each instance of G 4a and G 4b is fluoro, alkyl, alkoxy, aryloxy, or alkynyl;
wherein alkyl and alkoxy groups are unbranched, saturated, and have 1-4 carbon atoms; aryl groups and aryl groups of aryloxy can be either carbocyclic aryl or heterocyclic aryl; carbocyclic aryl groups have a total of 6-20 carbon atoms, including carbon atoms of substituents; heterocyclic aryl groups have a total of 5-20 carbon atoms, including carbon atoms of substituents; carboalkoxy groups are alkyl esters of a carboxylic acid wherein alkyl groups are as defined above; each alkyl, aryl, alkoxy, aryloxy, benzo, and carboalkoxy, independently, may be unsubstituted or substituted with one or more substituent; alkyl substituents are halo, hydroxyl, amino, or aryl; aryl substituents are halo, hydroxyl, amino, alkyl, aryl, nitro, or carboxyl; and halo substituents are fluoro or chloro; and
n is 0, 1, 2, 3, or 4.

2. The compound of claim 1 , wherein G 1 and G 3 are methyl.

3. The compound of claim 1 , wherein G 5 and G 7 are methyl.

4. The compound of any one of claims 1 - 3 , wherein the compound is of Formula (II):

or a pharmaceutically acceptable salt, solvate, or hydrate thereof.

5. The compound of claim 4 , wherein the compound is described by a formula selected from Table 1.

6. The compound of claim 4 , wherein the compound is selected from the group consisting of:

7. A method for imaging an alkyne-containing molecule comprising
(a) contacting a compound of claim 1 with a sample containing the alkyne-containing molecule under conditions for ligation of the compound to an alkyne group of the molecule to form a triazole product; and
(b) measuring a fluorescent signal released from the triazole product.

8. The method of claim 7 , wherein the contacting step is carried out in the presence of a metal catalyst.

9. The method of claim 8 , wherein the metal catalyst is copper(I).

10. The method of claim 7 , wherein the compound is covalently linked to the alkyne group.

11. The method of claim 7 , wherein the sample contains cells and the alkyne-containing molecule is located on the cell surface or inside the cells.

12. The method of claim 7 , wherein the compound is described by a formula selected from Table 1.

13. The method of claim 7 , wherein the compound is selected from a compound according to claim 6 .

14. The method of any one of claims 7 - 11 , wherein the alkyne-containing molecule is a biomolecule.

15. The method of claim 14 , wherein the biomolecule is a DNA, RNA, protein or glycan.

16. The method of claim 14 , wherein the biomolecule is located on or near the surface of a cell.

17. The method of claim 14 , wherein the biomolecule is an intracellular biomolecule.

18. A method for detecting an alkyne-containing molecule in a sample, comprising:
(a) contacting the compound of claim 1 with a sample suspected of having an alkyne-containing molecule;
(b) measuring the level of a fluorescent signal released from the sample mixture, and
(c) determining the presence of the alkyne-containing molecule in the sample, wherein an enhanced fluorescent signal as compared to a level of the fluorescent signal in the absence of the molecule indicates presence of the alkyne-containing molecule.

19. The method of claim 18 , wherein the contacting step is carried out in the presence of a metal catalyst.

20. The method of claim 19 , wherein the metal catalyst is copper(I).

21. The method of any one of claims 18 - 20 , wherein the sample contains cells and the alkyne-containing molecule is located on or near the surface of a cell or inside a cell.

22. The method of claim 18 , wherein the compound is described by a formula selected from Table 1.

23. The method of claim 18 , wherein the compound is selected from a compound according to claim 6 .

24. The method of claim 18 , wherein the alkyne-containing molecule is a biomolecule.

25. The method of claim 24 , wherein the biomolecule is a DNA, RNA, protein or glycan.

26. The method of claim 24 , wherein the biomolecule is located on or near the surface of a cell.

27. The method of claim 24 , wherein the biomolecule is an intracellular biomolecule.

28. The method of claim 7 or 18 wherein the method comprises the use of the compound for dual fluorescence imaging in a cell.

Assignees

• Academia Sinica

Inventors

• Wong Chi-Huey
• Fang Jim-Min
• Shie Jiun-Jie

Classifications

• C07D309/14

  Nitrogen atoms not forming part of a nitro radical · CPC title

• C07D311/02

  ortho- or peri-condensed with carbocyclic rings or ring systems · CPC title

• G01N2458/30

  Electrochemically active labels · CPC title

• G01N33/582 Primary

  with fluorescent label · CPC title

• C07F5/027

  Organoboranes and organoborohydrides · CPC title