# US 11,078,211 B2 · Photoresponsive nucleotide analog capable of photocrosslinking in visible light region

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- Number: US 11,078,211 B2
- Document ID: US11078211B2
- Status: Granted
- Kind: B2
- Application number: 16/629,148
- Filed: 2018-07-19
- Published: 2020-10-08
- Granted: 2021-08-03
- Earliest claimed: 2017-07-26
- Assignees: Japan Advanced Institute of Science and Technology; Nicca Chemical Co., Ltd.
- Applicants: Japan Advanced Institute of Science and Technology; Nicca Chemical Co., Ltd.
- Inventors: Takahiro Inatsugi; Ayako Koto; Isao Ishimaru; Masahiko Takamura; Kenzo Fujimoto
- Web page: https://patentgrep.com/patent/US11078211B2

## CPC

- C07D 491/052: the oxygen-containing ring being six-membered
- C07F 9/6561: containing systems of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring or ring system, with or without other non-condensed hetero rings
- C07H 1/00: Processes for the preparation of sugar derivatives
- C07H 21/04: with deoxyribosyl as saccharide radical
- C07D: HETEROCYCLIC COMPOUNDS (macromolecular compounds C08)
- C07F: ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM (metal-containing porphyrins C07D487/22)
- C07H: SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS (derivatives of aldonic or saccharic acids C07C, C07D; aldonic acids, saccharic acids C07C59/105, C07C59/285; cyanohydrins C07C255/16; glycals C07D; compounds of unknown constitution C07G; polysaccharides, derivatives thereof C08B; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; sugar industry C13)

## Abstract

Provided are a compound represented by the following formula I; a novel photoreactive compound that can be used in nucleic acid photoreaction techniques by a photoreactive crosslinking agent comprising the compound; and a photoreactive crosslinking agent in which the photoreactive compound is used.

## Claims

### Claim 1

A compound represented by the following formula I:
(I)
in which formula I:
R1 is a group selected from the group consisting of a hydrogen atom, a halogen atom, a methyl group, a fluorine-substituted methyl grow, an ethyl group, a fluorine-substituted ethyl group, and a C1-C3 alkylsulfanyl group;
X is a group represented by the following formula Is or IIs:
in which formula Is:
R11 and R12 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group;
na is 1 or 2;
R21 and R22 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group;
nb is 0 or 1;
R31 and R32 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group;
nc is 0, 1, 2 or 3, and nc+nd is an integer of 0 to 3;
R41 and R42 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group;
nd is 0, 1, 2 or 3, and nc+nd is an integer of 0 to 3;
Q1 is a group selected from the group consisting of:
a hydrogen atom:
a phosphate group formed together with O bonded to Q1;
a nucleotide, nucleic acid or peptide nucleic acid linked via a phosphodiester bond formed by a phosphate group formed together with O bonded to Q1; and
a protecting group selected from:
a trityl group, a monomethoxytrityl group, a dimethoxytrityl group, a trimethoxytrityl group, a trimethylsilyl group, a triethylsilyl group, a t-butyldimethylsilyl group, an acetyl group, and a benzoyl group;
Q2 is a group selected from the group consisting of:
a hydrogen atom;
a phosphate group formed together with O bonded to Q2;
a nucleotide, nucleic acid or peptide nucleic acid linked via a phosphodiester bond formed by a phosphate group formed together with O bonded to Q2; and
a protecting group selected from:
a 2-cyanoethyl-N,N-dialkyl(C1-C4)phosphoramidite group, a methylphosphonamidite group, an ethylphosphonamidite group, an oxazaphospholidine group, a thiophosphite group, a TEA salt of —PH(═O)OH, a DBU salt of —PH(═O)OH, a TEA salt of —PH(═S)OH, and a DBU salt of —PH(═S)OH; or
in which formula Is:
R51 and R52 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group;
ne is 1, 2 or 3;
R61 and R62 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group;
nf is 0, 1, 2 or 3;
Q1 is a groups selected from the group consisting of:
a hydrogen atom;
a phosphate group formed together with O bonded to Q1;
a nucleotide, nucleic acid or peptide nucleic acid linked via a phosphodiester bond formed by a phosphate group formed together with O bonded to Q1; and
a protecting group selected from:
a trityl group, a monomethoxytrityl group, a dimethoxytrityl group, a trimethoxytrityl group, a trimethylsilyl group, a triethylsilyl group, a t-butyldimethylsilyl group, an acetyl group, and a benzoyl group;
Q2 is group selected from the group consisting of:
a hydrogen atom;
a phosphate group formed together with O bonded to Q2;
a nucleotide, nucleic acid or peptide nucleic acid linked via a phosphodiester bond formed by a phosphate group formed together with O bonded to Q2; and
a protecting group selected from:
a 2-cyanoethyl-N,N-dialkyl(C1-C4)phosphoramidite group, a methylphosphonamidite group, an ethylphosphonamidite group, an oxazaphospholidine group, a thiophosphite group, a TEA salt of —PH(═O)OH, a DBU salt of —PH(═O)OH, a TEA salt of —PH(═S)OH, and a DBU salt of —PH(═S)OH.

### Claim 2 (depends on 1)

The compound according to claim 1, wherein X is a group represented by the following formula It:
in which formula It:
R11 and R12 are each independently the group as defined in the formula Is;
R31 and R32 are each independently the group as defined in the formula Is;
R41 and R42 are each independently the group as defined in the formula Is; and
Q1 and Q2 are the groups as defined in the formula Is.

### Claim 3 (depends on 1)

The compound according to claim 1, wherein X is a group represented by the following formula Iu:
in which formula Iu:
R42 is the group as defined in the formula Is; and
Q1 and Q2 are the groups as defined in the formula Is.

### Claim 4 (depends on 3)

The compound according to claim 3, wherein R42 is a hydrogen atom or a methyl group.

### Claim 5 (depends on 1)

A photoreactive crosslinking agent, comprising the compound according to claim 1.

### Claim 6 (depends on 1)

A reagent for producing photoreactive modified nucleic acids, comprising the compound according to claim 1.

### Claim 7 (depends on 1)

A method for forming a photocrosslink between a photoreactive crosslinking agent and a nucleic acid base having a pyrimidine ring, comprising:
hybridizing the photoreactive crosslinking agent comprising the compound according to claim 1 and the nucleic acid base having the pyrimidine ring, and
irradiating the photoreactive crosslinking agent and the nucleic acid base having the pyrimidine ring with light.

### Claim 8

A method for producing a compound represented by the following formula V, comprising subjecting a compound represented by the following formula III:
in which formula III:
R1 is a group selected from the group consisting of a hydrogen atom, a halogen atom, a methyl group, a fluorine-substituted methyl group, an ethyl group, a fluorine-substituted ethyl group, and a C1-C3 alkylsulfanyl group;
R11 and R12 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group; and
na is 1 or 2,
to dehydration condensation with a compound represented by the formula IV:
in which formula IV:
R21 and R22 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group;
nb is 0 or 1;
R31 and R32 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group,
nc is 0, 1, 2 or 3, and nc+nd is an integer of 0 to 3;
R41 and R42 are each independently a group selected from the group consisting of a hydrogen atom, a C1-C3 alkyl group, and a C1-C3 alkoxy group;
nd is 0, 1, 2 or 3, and nc+nd is an integer of 0 to 3;
Q1 is a group selected from the group consisting of:
a hydrogen atom;
a phosphate group formed together with O bonded to Q1;
a nucleotide, nucleic acid or peptide nucleic acid linked via a phosphodiester bond formed by a phosphate group formed together with O bonded to Q1; and
a protecting group selected from:
a trityl group, a monomethoxytrityl group, a dimethoxytrityl group, a trimethoxytrityl group, a trimethylsilyl group, a triethylsilyl group, a t-butyldimethylsilyl group, an acetyl group, and benzoyl group;
Q2 is a group selected from the group consisting of:
a hydrogen atom;
a phosphate group formed together with O bonded to Q2;
a nucleotide, nucleic acid or peptide nucleic acid linked via a phosphodiester bond formed by a phosphate group formed together with O bonded to Q2; and
a protecting group selected from:
a 2-cyanoethyl-N,N-dialkyl(C1-C4)phosphoramidite group, a methylphosphonamidite group, an ethylphosphonamidite group, an oxazaphospholidine group, a thiophosphite group, a TEA salt of —PH(═O)OH, a DBU salt of —PH(═O)OH, a TEA salt of —PH(═S)OH, and a DBU salt of —PH(═S)OH,
to provide a compound represented by the formula V:
in which formula V:
R1, R11 and R12 are each independently the group as defined in the formula III;
na is the integer as defined in the formula III;
R21, R22, R31, R32, R41, R42, Q1 and Q2 are each independently the group as defined in the formula IV; and
nb, nc, and nd are each independently the integer as defined in the formula IV.

### Claim 9 (depends on 1)

The compound according to claim 1,
wherein R1 is a group selected from the group consisting of a hydrogen atom, a halogen atom, a methyl group, an ethyl group, and a C1-C3 alkylsulfanyl group.

### Claim 10 (depends on 8)

The method according to claim 8,
wherein R1 is a group selected from the group consisting of a hydrogen atom, a halogen atom, a methyl group, an ethyl group, and a C1-C3 alkylsulfanyl group.