Water-soluble triazapyridinophane-based complexing agents and corresponding fluorescent lanthanide complexes

Patent metadata

Field	Value
Publication number	US-10597409-B2
Application number	US-201616060291-A
Country	US
Kind code	B2
Filing date	Dec 9, 2016
Priority date	Dec 9, 2015
Publication date	Mar 24, 2020
Grant date	Mar 24, 2020

Abstract

The invention can be used for marking biological molecules.

First claim

The invention claimed is:

1. A complexing agent of formula (I):

wherein:
the R 1 groups are identical and are either: —CO 2 H, or —PO(OH)R, wherein R is selected from the group consisting of: phenyl optionally substituted by an —SO 3 H group benzyl, methyl, ethyl, propyl, n-butyl, sec-butyl, isobutyl, and tent-butyl;
the A 1 , A 2 groups are identical or different and are selected from the group consisting of: a group of formula -L 1 -E; a group of formula (II) and a group of formula (II′);
the A 3 group is selected from the group consisting of: a group of formula —O-L 3 -G, a group of formula (II), a group of formula (II′), a group of formula (III), and a group of formula (III′);

L 1 , L 2 and L 3 are identical or different and each represent a divalent linkage group;
E is a group that renders the complexing agent water-soluble, chosen selected from the group consisting of: —SO 3 H, —PO(OH) 2 , —CO 2 H, —N + Alk 1 Alk 2 Alk 3 , a carbohydrate residue, a sulfobetaine; and a PEG group;
G is a reactive group selected from the group consisting of: an acrylamide, an amine, an ester, an aldehyde, an alkyl halide, an anhydride, an aniline, an azide, an aziridine, a carboxylic acid, a diazoalkane, a haloacetamide, a halotriazine, dichlorotriazine, a hydrazine, an imido ester, an isocyanate, an isothiocyanate, a maleimide, a sulfonyl halide, a thiol, a ketone, an acid halide, a succinimidyl ester, a hydroxysuccinimidyl ester, a hydroxysulfosuccinimidyl ester, an azidonitrophenyl, an azidophenyl, a glyoxal, a triazine, and an acetylenic group;
Alk 1 , Alk 2 , Alk 3 , which may be identical or different, each represent a (C 1 -C 6 )alkyl.

2. The complexing agent as claimed in claim 1 , wherein A 1 and A 2 are identical and are a group of formula (II) and A 3 is a group of formula (III).

3. The complexing agent as claimed in claim 1 , wherein A 1 is a group of formula (II), A 2 is an -L 1 -E group and A 3 is a group of formula (III).

4. The complexing agent as claimed in claim 1 , wherein A 1 and A 2 are identical and are groups of formula (II), and A 3 is a group of formula —O-L 3 -G.

5. The complexing agent as claimed in claim 1 , wherein A 1 and A 2 are identical and are groups of formula -L 1 -E and A 3 is a group of formula (III).

6. The complexing agent as claimed in claim 1 , wherein A 1 and A 2 are identical and are groups of formula (II′) and A 3 is a group of formula (III′).

7. The complexing agent as claimed in claim 1 , wherein A 1 is a group of formula (II′), A 2 is an -L 1 -E group and A 3 is a group of formula (III′).

8. The complexing agent as claimed in claim 1 , wherein A 1 and A 2 are identical and are groups of formula (II′), and A 3 is a group of formula —O-L 3 -G.

9. The complexing agent as claimed in claim 1 , wherein A 1 and A 2 are identical and are groups of formula -L 1 -E and A 3 is a group of formula (III′).

10. The complexing agent as claimed in claim 1 , wherein L 1 , L 2 and L 3 are selected from the group consisting of:
a linear or branched C 1 -C 20 alkylene group optionally containing one or more double or triple bonds;
a C 5 -C 8 cycloalkylene group,
a C 6 -C 14 arylene group,
said alkylene, cycloalkylene or arylene groups optionally containing one or more hetero atoms, or one or more carbamoyl or carboxamido groups, and said alkylene, cycloalkylene or arylene groups being optionally substituted by C 1 -C 8 alkyl, C 6 -C 14 aryl, sulfonate or oxo groups; and
a group having one of the following formulae:

wherein n, m, p, q are integers from 1 to 5.

11. The complexing agent as claimed in claim 1 , wherein the -L 3 -G group consists of (i) a reactive group G selected from the group consisting of: a carboxylic acid, an amine, a succinimidyl ester, a haloacetamide, a hydrazine, an isothiocyanate, and a maleimide group, and (ii) a spacer arm L 3 consisting of an alkylene group comprising from 1 to 5 carbon atoms.

12. The complexing agent as claimed in claim 1 , wherein L 1 is a divalent group of formula:

where n is an integer from 1 to 5.

13. The complexing agent as claimed in claim 1 , wherein L 2 is a divalent group of formula:

where n and m are integers from 1 to 5.

14. The complexing agent as claimed in claim 1 , wherein L 3 is a divalent group of formula:
—(CH 2 ) n —

where n is an integer from 1 to 5.

15. A fluorescent lanthanide complex comprising a complexing agent as claimed in claim 1 and a lanthanide selected from the group consisting of: Eu 3+ , Sm 3+ , and Tb 3+ .

16. A fluorescent lanthanide complex of formula (IV), (V), (VI) or (VII) below:

wherein:
Ln 3+ is chosen from: Eu 3+ , Tb 3+ , Sm 3+ ;
R 3 is chosen from the following groups: OH;

17. A fluorescent conjugate of a molecule of interest selected from the group consisting of: an amino acid, a peptide, a protein, an antibody, a sugar, a carbohydrate chain, a nucleoside, a nucleotide, an oligonucleotide, and an enzyme substrate, said molecule being covalently conjugated with a lanthanide complex as claimed in claim 15 .

18. The complexing agent as claimed in claim 1 , wherein the reactive group G is a group having one of the following formulae:

wherein w varies from 0 to 8 and v is equal to 0 or 1, and Ar is a saturated or unsaturated 5- or 6-membered heterocycle, comprising 1 to 3 heteroatoms, optionally substituted by a halogen atom.

19. The fluorescent conjugate of claim 17 , wherein the molecule is the enzyme substrate and is selected from the group consisting of: a benzylguanine, a benzylcytosine, a chloroalkane and a coenzyme A.

Assignees

• Cisbio Bioassays
• Univ Toulouse
• Centre Nat Rech Scient

Inventors

• Lamarque Laurent
• Picard Claude
• Galaup Chantal
• Leygue Nadine
• Zwier Jurriaan
• Bourrier Emmanuel

Classifications

• C07D401/14

  containing three or more hetero rings · CPC title

• C07F5/003 Primary

  without C-Metal linkages · CPC title

• G01N2458/40

  Rare earth chelates · CPC title

• G01N33/582

  with fluorescent label · CPC title