Acidic environment-detecting fluorescent probe

Patent metadata

Field	Value
Publication number	US-9784732-B2
Application number	US-201214343033-A
Country	US
Kind code	B2
Filing date	Sep 6, 2012
Priority date	Sep 7, 2011
Publication date	Oct 10, 2017
Grant date	Oct 10, 2017

Abstract

Provided is a compound represented by formula (I), which can be used as a fluorescent probe that becomes highly fluorescent only in an intracellular acidic environment and can be adapted to the fluorescent imaging of an intracellular vesicle that is a microstructure.

In the formula, R 1 and R 2 independently represent a hydrogen atom, or an alkyl group which may be substituted (wherein R 1 and/or R 2 represents an alkyl group which may be substituted); R 3 , R 4 , R 5 , R 6 , R 7 and R 8 independently represent a hydrogen atom, a halogen atom, or the like; X represents a functional group to which a labeling site or a target-accumulating site can be introduced, or the like; Y represents a halogen atom, an alkyl group which may be substituted, or the like; m represents an integer of 0 to 5; and n represents an integer of 0 to 5.

First claim

What is claimed is:

1. A compound represented by the following general formula (I) or a salt thereof:

wherein R 1 and R 2 each independently represents hydrogen or an alkyl group that may be substituted with at least one substituent selected from the group consisting of halogen atom, hydroxyl group, alkoxyl group, amino group, carboxyl group, sulfo group, and alkylsulfonyloxy group (at least one of R 1 and R 2 being an alkyl group that may be substituted with at least one substituent selected from the group consisting of halogen atom, hydroxyl group, alkoxyl group, amino group, carboxyl group, sulfo group, and alkylsulfonyloxy group); R 3 , R 4 , R 5 , R 6 , R 7 , and R 8 each independently represents hydrogen, a halogen, or an alkyl group that may be substituted with at least one substituent selected from the group consisting of halogen atom, hydroxyl group, alkoxyl group, amino group, carboxyl group, sulfo group, and alkylsulfonyloxy group; X represents a functional group into which a labeling site or a target-accumulating site can be introduced, or a substituent into which a labeling site or a target-accumulating site has been introduced, wherein the functional group into which a labeling site or a target-accumulating site can be introduced is a carboxyl group, or an alkyl group substituted with a carboxyl group, and wherein the substituent into which a labeling site or a target-accumulating site has been introduced is represented by —(X′-T-S) (X′ represents a group into which a labeling site or target-accumulating site is introduced, wherein X′ is a carbonyl group, an alkyl carbonyl group, or a carboxamido group; T, when present, represents a crosslinking group, wherein the crosslinking group is selected from the group consisting of a divalent hydrocarbon group, an ethylene glycol group, a diethylene glycol group, a triethylene glycol group, and a divalent piperidine ring group, and the crosslinking group optionally modified by a functional group selected from the group consisting of an amino group, a carbonyl group, a carbonyloxy group, and a carboxamido group at one or both terminals; and S represents a labeling site or target-accumulating site, wherein S is (2,5-dioxo-1-pyrrolidinyl)oxy, a 2-(2-((6-chlorohexyl)oxy)ethoxy)ethane amino group, a maleimido, an isothiocyanato group, a chlorosulfonyl group, a haloalkyl group, a haloacetamido group, an azido group, an alkynyl group and a polyethylene glycol group having not more than three ethoxy units and optionally substituted by a group selected from the group consisting of an amino group, a carbonyl group and a carboxyl group at one or both terminals; Y represents a halogen, an alkyl group that may be substituted with at least one substituent selected from the group consisting of halogen atom, hydroxyl group, alkoxyl group, amino group, carboxyl group, sulfo group, and alkylsulfonyloxy group, an alkoxyl group that may be substituted with at least one substituent selected from the group consisting of halogen atom, hydroxyl group, alkoxyl group, amino group, carboxyl group, sulfo group, and alkylsulfonyloxy group, or a cyano group; m represents an integer from 1 to 5; X may optionally be identical or different when m is 2 or higher; n represents an integer from 0 to 5; and Y may optionally be identical or different when n is 2 or higher and the sum of m and n being an integer equal to 5 or less.

2. The compound or salt thereof according to claim 1 , wherein each of R 1 and/or R 2 in general formula (I) is a C 1-4 alkyl group that may be substituted with at least one substituent selected from the group consisting of halogen atom, hydroxyl group, alkoxyl group, amino group, carboxyl group, sulfo group, and alkylsulfonyloxy group.

3. The compound or salt thereof according to claim 1 , wherein the compound is represented by the following formula:

4. A fluorescent probe comprising the compound or salt thereof according to claim 1 .

5. A method of measuring pH changes in intracellular acidic regions in living cells or tissues, the method comprising:
(a) introducing the compound or salt thereof according to claim 1 into a cell; and
(b) measuring fluorescence emitted within the cell by the compound or salt thereof.

6. The method according to claim 5 , wherein acidic regions in which intracellular acidic organelles are present are measured.

Assignees

• Urano Yasuteru
• Nagano Tetsuo
• Asanuma Daisuke
• Hirose Kenzo
• Namiki Shigeyuki
• TAKAOKA Yousuke
• Univ Tokyo

Inventors

• Urano Yasuteru
• Nagano Tetsuo
• Asanuma Daisuke
• Hirose Kenzo
• Namiki Shigeyuki
• TAKAOKA Yousuke

Classifications

• C09K11/06

  containing organic luminescent materials · CPC title

• C09K2211/1088

  containing oxygen as the only heteroatom · CPC title

• C07D405/04

  directly linked by a ring-member-to-ring-member bond · CPC title

• C09B69/103

  containing a diaryl- or triarylmethane dye · CPC title

• G01N33/52 Primary

  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