Merocyanine-based compounds, and dyes, kits and contrast medium compositions for labelling biomolecules comprising the same

Patent metadata

Field	Value
Publication number	US-9969887-B2
Application number	US-201715490971-A
Country	US
Kind code	B2
Filing date	Apr 19, 2017
Priority date	Apr 20, 2016
Publication date	May 15, 2018
Grant date	May 15, 2018

Abstract

The present disclosure relates to a novel merocyanine-based compound capable of labeling biomolecules by intercalating biomolecules, and to a dye, kit and contrast medium composition for labelling biomolecules comprising the same.

First claim

What is claimed is:

1. A merocyanine-based compound having a structure represented by the following formula 1:

wherein
Ar is a substituted or unsubstituted aromatic ring;
Y 1 and Y 2 are each independently selected from sulfur, oxygen, selenium, NR 8 and —CR 8 =CR 9 -;
R 1 to R 9 are each independently selected from the group consisting of hydrogen, deuterium, substituted or unsubstituted C 1 -C 10 alkyl, substituted or unsubstituted C 2 -C 10 heteroalkyl containing at least one heteroatom, substituted or unsubstituted C 2 -C 10 alkenyl, substituted or unsubstituted C 2 -C 10 alkynyl, substituted or unsubstituted C 1 -C 10 alkoxy, substituted or unsubstituted aryloxy, substituted or unsubstituted C 1 -C 10 haloalkyl, halogen, cyano, hydroxyl, substituted or unsubstituted amino, substituted or unsubstituted amide, carbamate, sulthydryl, nitro, carboxyl, carboxylate, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted aralkyl, quaternary ammonium, phosphoric acid, phosphate, phosphonate, —COR 10 , aldehyde, —COOR 10 , acyl chloride, sulfonic acid, sulfonate, polyalkylene oxide, and -L-Z functional groups;
when R a , wherein a is an integer selected from 1 to 9, is a —COR 10 group or a —COOR 10 group, R 10 is selected from the group consisting of substituted or unsubstituted C 1 -C 10 alkyl, substituted or unsubstituted C 2 -C 10 heteroalkyl containing at least one heteroatom, substituted or unsubstituted C 2 -C 10 alkenyl, substituted or unsubstituted C 2 -C 10 alkynyl, substituted or unsubstituted C 1 -C 10 alkoxy, substituted or unsubstituted C 1 -C 10 haloalkyl, and substituted or unsubstituted C 1 -C 10 aminoalkyl;
when R b , wherein b is an integer selected from 1 to 10, is substituted, any carbon or terminal carbon in the functional groups may be substituted with at least one substituent selected from the group consisting of sulfonic acid, sulfonate, ketone, aldehyde, carboxylic acid, carboxylate, phosphoric acid, phosphate, phosphonate, acyl chloride, polyalkylene oxide, quaternary ammonium salt, ester, and amide;
m is an integer of 1 to 3;
L is a linker comprising 3 to 150 non-hydrogen atoms;
Z is a fluorescent moiety capable of generating a fluorescent signal, or has a structure represented by formula 1; and
wherein the structure represented by the formula 1 has one or two -L-Z functional groups.

2. The merocyanine-based compound of claim 1 , wherein Z is selected from the group consisting of phenanthridium, coumarins, cyanine, bodipy, fluoresceins, rhodamines, pyrenes, carbopyronin, oxazine, xanthenes, thioxanthene, and acridine.

3. The merocyanine-based compound of claim 1 , wherein the -L-Z functional group is represented by the following formula 2:
-L 1 -[A 1 -(CH 2 ) x1 -] y1 [A 2 -(CH 2 ) x2 -] y2 [A 3 -(CH 2 ) x3 -] y3 [A 4 -(CH 2 ) x4 -] y4 [A 5 -(CH 2 ) x5 -] y5 [A 6 -(CH 2 ) x6 -] y6 [A 7 -(CH 2 ) x7 -] y7 [A 8 -(CH 2 ) x8 -] y8 [A 9 -(CH 2 ) x9 -] y9 -A 10 -L 2 -Z  [Formula 2],

wherein
L 1 and L 2 are each independently a C 1 -C 12 polymethylene unit optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur, or an aryl optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur;
A 1 to A 10 are each independently a chain alkyl or branched alkyl optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur, or a five- or six-membered ring optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur;
x1 to x9 are each independently 0 or an integer of 1 to 20;
y1 to y9 are each independently 0 or an integer of 1 to 20; and
Z is selected from the group consisting of phenanthridium, coumarins, cyanine, bodipy, fluoresceins, rhodamines, pyrenes, carbopyronin, oxazine, xanthenes, thioxanthene, and acridine.

4. The merocyanine-based compound of claim 3 , wherein one of A 1 to A 10 is represented by the following formula 3:

wherein R 11 is an aryl optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur;
R 12 is represented by the following formula 4:
-[A 11 -(CH 2 ) x11 -] y11 [A 12 -(CH 2 ) x12 -] y12 [A 13 -(CH 2 ) x13 -]A 14 -L 3 -Z   [Formula 4],

wherein L 3 is a C 1 -C 12 polymethylene unit optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur, or an aryl optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur;
A 11 to A 14 are each independently a chain alkyl or branched alkyl optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur, or a five- or six-membered ring optionally containing at least one heteroatom selected from nitrogen, oxygen and sulfur;
x11 to x13 are each independently 0 or an integer of 1 to 20; and
y11 to y13 are each independently 0 or an integer of 1 to 20.

5. A dye for labeling biomolecules comprising the merocyanine-based compound according to claim 1 .

6. The dye for labeling biomolecules of claim 5 , wherein the merocyanine-based compound is a compound that is intercalated into a nucleic acid which is a biomolecule.

7. The dye for labeling biomolecules of claim 6 , wherein the biomolecule is at least one nucleic acid selected from a single-stranded RNA, a double-stranded RNA, a single-stranded DNA, and a double-stranded DNA.

8. A kit for labeling biomolecules comprising the merocyanine-based compound according to claim 1 .

9. The kit of claim 8 , wherein the kit is an electrophoresis kit, wherein the electrophoresis kit comprises the merocyanine-based compound; and a buffer, a gel matrix, at least one material for forming a gel matrix, or at least one material for forming a surface.

10. A method for determining the presence or absence of nucleic acids in a sample, comprising:
when the nucleic acids are present in a sample,
exposing the nucleic acids to the merocyanine-based compound according to claim 1 , such that the merocyanine-based compound is intercalated into the nucleic acids to form a complex; and
determining the presence or absence of fluorescence of the merocyanine-based compound.

11. A method for determining the presence or absence of an amplified target nucleic acid, comprising:
when performing a nucleic acid amplification reaction,
providing a reaction mixture comprising a target nucleic acid, a reagent necessary for amplifying the target nucleic acid, and the merocyanine-based compound according to claim 1 ;
subjecting the reaction mixture to polymerization under conditions suitable for the formation of the amplified target nucleic acid;
illuminating the reaction mixture with light; and
detecting a fluorescence emission from the reaction mixture.

12. The kit of claim 8 , wherein the kit is for determining the viability of cells in a sample, the kit comprises the merocyanine-based compound; and when apoptotic cells are present in a sample, fluorescence is detected by intercalating them with the merocyanine-based compound.

Assignees

• Sfc Co Ltd

Inventors

• Kim Su-Jin
• Lee Do-Min
• Je Jong-Tae

Classifications

• G01N33/52

  Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper {and including single- and multilayer analytical elements (immunological elements G01N33/54386; involving labelled immunochemicals G01N33/58; for haemoglobin or occult blood G01N33/72)} · CPC title

• G01N27/447

  using electrophoresis · CPC title

• G01N21/6486

  Measuring fluorescence of biological material, e.g. DNA, RNA, cells (G01N21/6428 takes precedence) · CPC title

• C12Q1/6844

  Nucleic acid amplification reactions · CPC title

• C12Q1/6813

  Hybridisation assays · CPC title