Reporter and uses thereof

Patent metadata

Field	Value
Publication number	US-12215381-B2
Application number	US-202318461211-A
Country	US
Kind code	B2
Filing date	Sep 5, 2023
Priority date	Mar 10, 2021
Publication date	Feb 4, 2025
Grant date	Feb 4, 2025

Abstract

The present invention relates to a fluorescent reporter capable of labeling nucleic acids including DNA and exhibiting luminescent properties at an excited energy level, and various uses thereof.

First claim

What is claimed is:

1. A reporter for labeling a nucleic acid represented by Formula 5 below:

wherein,
R 21 and R 22 are each independently selected from hydrogen, deuterium, substituted or unsubstituted C 1 -C 10 alkyl, substituted or unsubstituted C 1 -C 10 heteroalkyl containing at least one hetero atom, substituted or unsubstituted C 2 -C 10 alkenyl, substituted or unsubstituted C 2 -C 10 alkynyl, substituted or unsubstituted C 3 -C 20 cycloalkyl, substituted or unsubstituted C 3 -C 20 cycloalkenyl, substituted or unsubstituted C 2 -C 20 heterocycloalkyl, substituted or unsubstituted C 5 -C 20 aryl, and substituted or unsubstituted C 2 -C 20 heteroaryl,
R 23 to R 30 are each independently a functional group selected from hydrogen, deuterium, substituted or unsubstituted C 1 -C 10 alkyl, substituted or unsubstituted C 1 -C 10 heteroalkyl containing at least one hetero atom, substituted or unsubstituted C 2 -C 10 alkenyl, substituted or unsubstituted C 2 -C 10 alkynyl, substituted or unsubstituted C 3 -C 20 cycloalkyl, substituted or unsubstituted C 3 -C 20 cycloalkenyl, substituted or unsubstituted C 2 -C 20 heterocycloalkyl, hydroxy, oxido (—O − ), substituted or unsubstituted C 1 -C 10 alkoxy, substituted or unsubstituted C 3 -C 20 cycloalkyloxy, substituted or unsubstituted C 5 -C 20 aryloxy, substituted or unsubstituted C 2 -C 20 heteroaryloxy, substituted or unsubstituted C 5 -C 20 aryl, substituted or unsubstituted C 2 -C 20 heteroaryl, substituted or unsubstituted C 5 -C 20 aralkyl, substituted or unsubstituted C 1 -C 10 alkylthio, substituted or unsubstituted C 5 -C 20 arylthio, substituted or unsubstituted C 3 -C 20 cycloalkylthio, substituted or unsubstituted C 2 -C 20 heteroarylthio, substituted or unsubstituted acylamino, acyloxy, substituted or unsubstituted phosphino, trifluoromethylsulfonyl (—SO 2 CF 3 ), substituted or unsubstituted ammonium, nitro, substituted sulfonyl, substituted sulfonic acid ester, substituted or unsubstituted sulfonamide, substituted thioketone, trihalomethyl (—CF 3 , —CCl 3 , —CBr 3 , or —CI 3 ), haloformyl (—COCl, —COBr, or —COI), formyl (—CHO), acyl, substituted ester, substituted or unsubstituted aminocarbonyl, nitro, nitroso (—N═O), fluoro (—F), chloro (—Cl), bromo (—Br), iodo (—I), substituted or unsubstituted germanium, substituted or unsubstituted boron, substituted or unsubstituted aluminum, substituted or unsubstituted silyl, substituted or unsubstituted amide, carbamate, substituted or unsubstituted phosphine, nitrile, hydrazine, acetal, ketal, polyalkyleneoxide, and -L 1 -R 12 , or two adjacent functional groups are bonded to each other to form a ring,
R 31 to R 34 are each independently selected from hydrogen, deuterium, substituted or unsubstituted C 1 -C 10 alkyl, substituted or unsubstituted C 1 -C 10 heteroalkyl containing at least one hetero atom, and -L 1 -R 12 , and at least one of R 31 to R 34 is -L 1 -R 12 ,
L 1 includes a single bond or at least one selected from substituted or unsubstituted C 1 -C 10 alkyl, substituted or unsubstituted C 1 -C 10 heteroalkyl containing at least one hetero atom, substituted or unsubstituted C 2 -C 10 alkenyl, substituted or unsubstituted C 2 -C 10 alkynyl, substituted or unsubstituted C 3 -C 20 cycloalkyl, substituted or unsubstituted C 3 -C 20 cycloalkenyl, and substituted or unsubstituted C 2 -C 20 heterocycloalkyl, and
R 12 is a deoxyribonucleoside represented by Formula 3 below,

Wherein,
* indicates a location where the deoxyribonucleoside is bonded to L 1 ,
B is a nucleobase,
R 13 is selected from hydrogen, deuterium, P(OR 15 )(N(R 16 R 17 ), and -L 2 -R 18 ,
R 14 is an alcohol protecting group, hydrogen or P(OR 15 )(N(R 16 R 17 ), or a support or nucleic acid to which the reporter is bound, at least one of R 13 and R 14 is P(OR 15 )(N(R 16 R 17 ),
R 15 to R 17 are each independently selected from hydrogen, deuterium, substituted or unsubstituted C 1 -C 10 alkyl, and substituted or unsubstituted C 1 -C 10 heteroalkyl containing at least one hetero atom,
L 2 is a single bond, or selected from an internucleotide phosphodiester bond, substituted or unsubstituted C 1 -C 10 alkyl and substituted or unsubstituted C 1 -C 10 heteroalkyl containing at least one hetero atom,
R 18 is hydroxy or P(OR 15 )(N(R 16 R 17 ), or a support or nucleic acid to which the reporter is bound,
Y is CR 35 , each R 35 is selected from hydrogen, deuterium, substituted or unsubstituted C 1 -C 10 alkyl, substituted or unsubstituted C 1 -C 10 heteroalkyl containing at least one hetero atom, fluoro (—F), chloro (—Cl), bromo (—Br), and iodo (—I), and
n is an integer of 1 to 4,
when any functional group of R 21 to R 35 is a substituted functional group, any substituent other than hydrogen bonded to at least one carbon of the functional group,
wherein the substituent is selected from deuterium, C 1 -C 10 alkyl, C 1 -C 10 heteroalkyl containing at least one hetero atom, C 2 -C 10 alkenyl, C 2 -C 10 alkynyl, C 3 -C 20 cycloalkyl, C 3 -C 20 cycloalkenyl, C 2 -C 20 heterocycloalkyl, hydroxy, oxido (—O − ), C 1 -C 10 alkoxy, C 3 -C 20 cycloalkyloxy, C 5 -C 20 aryloxy, C 2 -C 20 heteroaryloxy, C 5 -C 20 aryl, C 2 -C 20 heteroaryl, C 5 -C 20 aralkyl, C 1 -C 10 alkylthio, C 5 -C 20 arylthio, C 3 -C 20 cycloalkylthio, C 2 -C 20 heteroarylthio, acylamino, acyloxy, phosphino, trifluoromethylsulfonyl (—SO 2 CF 3 ), ammonium, nitro, substituted sulfonyl, substituted sulfonic acid ester, sulfonamide, substituted thioketone, trihalomethyl (—CF 3 , —CCl 3 , —CBr 3 , or —CI 3 ), haloformyl (—COCl, —COBr, or —COI), formyl (—CHO), acyl, substituted ester, aminocarbonyl, nitro, nitroso (—N═O), fluoro (—F), chloro (—Cl), bromo (—Br), iodo (—I), germanium, boron, aluminum, silyl, amide, carbamate, phosphine, nitrile, hydrazine, acetal, ketal, polyalkyleneoxide, and -L 1 -R 12 .

2. The reporter of claim 1 , wherein L 1 is a linker including at least one carbon, and any carbon of the linker is a carbonyl carbon.

3. The reporter of claim 1 , wherein R 12 is a deoxyribonucleoside represented by Formula 4 below:

4. An oligonucleotide, comprising:
the reporter according to claim 1 ; and
a quencher.

5. The oligonucleotide of claim 4 , further comprising:
a minor groove binder (MGB) interposed between the reporter and the quencher.

6. A composition for detecting a nucleic acid, comprising the oligonucleotide of claim 4 .

7. A support for detecting a nucleic acid, comprising:
the reporter of claim 1 ;
a support; and
a linker connecting the reporter and the support, through any position of the compound of Formula 3.

8. The support of claim 7 , wherein the support is glass, cellulose, nylon, acrylamide gel, dextran, polystyrene, or resin.

9. A method of detecting a nucleic acid, comprising:
(a) preparing a reaction mixture including a target nucleic acid, a reagent required for amplifying the target nucleic acid, and the oligonucleotide of claim 4 ;
(b) amplifying the target nucleic acid in the reaction mixture through polymerase chain reaction; and
(c) measuring a fluorescence intensity of the reaction mixture.

10. The method of claim 9 , wherein (b) includes
(b-1) elongating the oligonucleotide hybridized to the target nucleic

Assignees

• Sfc Co Ltd

Inventors

• Lee Do Min
• Si Ho Young
• Jo Jae Hyung
• Masanta Goutam
• Song Ju Man
• Eum Min Su
• Je Jong Tae

Classifications

• C09B23/00

  Methine or polymethine dyes, e.g. cyanine dyes · CPC title

• G01N21/6486

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

• C12Q1/686

  Polymerase chain reaction [PCR] · CPC title

• C12Q1/6834 Primary

  Enzymatic or biochemical coupling of nucleic acids to a solid phase · CPC title

• C07H19/06

  Pyrimidine radicals · CPC title