# US 11,897,914 B2 · Synthesis of 2′ protected nucleosides

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- Number: US 11,897,914 B2
- Document ID: US11897914B2
- Status: Granted
- Kind: B2
- Application number: 18/059,357
- Filed: 2022-11-28
- Published: 2023-12-28
- Granted: 2024-02-13
- Earliest claimed: 2021-11-29
- Assignees: Hongene Biotech Corporation
- Applicants: Hongene Biotech Corporation
- Inventors: Wing C. Poon; Ruiming Zou; Aldrich N. K. Lau; David Yu; Gengyu Du; Yun-Chiao Yao; Gang Zhao
- Web page: https://patentgrep.com/patent/US11897914B2

## CPC

- C07H 1/00: Processes for the preparation of sugar derivatives
- C07H 21/02: with ribosyl as saccharide radical
- C07H 19/067: with ribosyl as the saccharide radical
- C07H 19/10: with the saccharide radical esterified by phosphoric or polyphosphoric acids
- C07H 19/167: with ribosyl as the saccharide radical
- C07H 19/20: with the saccharide radical esterified by phosphoric or polyphosphoric acids
- C07H 23/00: Compounds containing boron, silicon or a metal, e.g. chelates or vitamin B12 (esters with inorganic acids C07H11/00)
- 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

Embodiments of the present application relate to the preparation of 2′-O-protected nucleoside phosphoramidites and conjugation of the 2′-O-protected nucleoside to a solid support. More specifically, the present application relates to nucleosides having a hydroxy protecting group at the 2′ position that reduces migration to the 3′ position during RNA (e.g., mRNA) synthesis and can be readily removed under mild conditions without the use of toxic metal-containing reagents. Methods of using the nucleosides described herein in RNA synthesis are also disclosed.

## Claims

### Claim 1

A compound of Formula (I):
or a pharmaceutically acceptable salt thereof,
wherein
R1 is hydrogen or a hydroxy protecting group;
R2 is hydrogen, a hydroxy protecting group, —C(═O)CH2CH2C(═O)R3, or —P(OR4)NR5R6;
alternatively, R1 and R2 are joined together to form a 6-10 membered heterocyclic ring;
RHet is optionally substituted 5-10 membered heteroaryl or optionally substituted 5-10 membered heterocyclyl;
R3 is hydroxy, —OR7 or —NR8R9;
each of R4, R5 and R6 is independently H, unsubstituted C1-C6 alkyl, or substituted C1-C6 alkyl;
R7 is unsubstituted C1-C6 alkyl, substituted C1-C6 alkyl, or a hydroxy protecting group;
each of R8 and R9 is independently H, unsubstituted or substituted C1-6 alkyl, or an amino protecting group; and
B is a natural nucleobase, a modified natural nucleobase, or an unnatural nucleobase.

### Claim 2 (depends on 1)

The compound of claim 1, wherein R1 is a trityl type of hydroxy protecting group selected from the group consisting of (4-methoxyphenyl)diphenylmethyl, bis(4-methoxyphenyl)phenylmethyl, tris(4-methoxyphenyl)methyl, 9-phenylxanthen-9-yl, and 9-(4-methoxyphenyl)xanthen-9-yl.

### Claim 3 (depends on 1)

The compound of claim 1, wherein R1 is bis(4-methoxyphenyl)phenylmethyl.

### Claim 4 (depends on 1)

The compound of claim 1, wherein R1 is hydrogen.

### Claim 5 (depends on 1)

The compound of claim 1, wherein R2 is H.

### Claim 6 (depends on 1)

The compound of claim 1, where R2 is —P(OR4)NR5R6, wherein:
R4 is substituted C1-C6 alkyl; and
R5 and R6 are each independently unsubstituted C1-C6 alkyl.

### Claim 7 (depends on 6)

The compound of claim 6, wherein R2 is

### Claim 8 (depends on 1)

The compound of claim 1, wherein R1 and R2 are joined together to form a 7 membered heterocyclic ring containing oxygen and silicon atoms.

### Claim 9 (depends on 8)

The compound of claim 8, wherein the compound of Formula (I) is also represented by Formula (Ia):
or a pharmaceutically acceptable salt thereof, wherein each of RA, RB, RC, and RD is independently H, unsubstituted C1-C6 alkyl, or substituted C1-C6 alkyl.

### Claim 10 (depends on 9)

The compound of claim 9, wherein each of RA, RB, RC, and RD is isopropyl.

### Claim 11 (depends on 1)

The compound of claim 1, wherein RHet is optionally substituted 5-10 membered heteroaryl containing one to four heteroatoms selecting from the group consisting of O, N and S.

### Claim 12 (depends on 11)

The compound of claim 11, wherein RHet is selected from the group consisting of pyridinyl, pyrimidinyl, furanyl, pyrrolyl, thienyl, thiazolyl, isothiazolyl, imidazolyl, pyrazolyl, oxazolyl, isoxazolyl, benzofuranyl, benzoimidazolyl, benzopyrazolyl, benzothiazolyl, benzooxazolyl, indolyl, and quinolinyl, each optionally substituted with one, two or three substituents Q; wherein each Q is independently selected from the group consisting of halo, cyano, hydroxy, amino, C1-C6 alkyl, C1-C6 haloalkyl, hydroxy(C1-C6 alkyl), amino(C1-C6 alkyl), (C1-C6 alkyl)amino, C1-C6 alkoxy, C1-C6 haloalkoxy, (C1-C6 alkoxy)C1-C6 alkyl and —O(C1-C6 alkoxy)C1-C6alkyl.

### Claim 13 (depends on 12)

The compound of claim 12, wherein each Q is independently selected from the group consisting of —F, —Cl, —Br, —CH3, —CH2CH3, —CF3, —CH3, —OCH3, and —OCH2CH3.

### Claim 14 (depends on 1)

The compound of claim 1, wherein RHet is optionally substituted 5-10 membered heterocyclyl containing one to four heteroatoms selecting from the group consisting of O, N and S.

### Claim 15 (depends on 14)

The compound of claim 14, wherein RHet is selected from the group consisting of tetrahydofuranyl, tetrahydropyranyl, pyrrolidinyl, piperidinyl, piperazinyl, and morpholinyl, each optionally substituted with one, two or three substituents Q; wherein each Q is independently selected from the group consisting of halo, cyano, hydroxy, amino, C1-C6 alkyl, C1-C6 haloalkyl, hydroxy(C1-C6 alkyl), amino(C1-C6 alkyl), (C1-C6 alkyl)amino, C1-C6 alkoxy, C1-C6 haloalkoxy, (C1-C6 alkoxy)C1-C6 alkyl and —O(C1-C6 alkoxy)C1-C6 alkyl.

### Claim 16 (depends on 1)

The compound of claim 1, wherein B is:
wherein Rx is hydrogen, unsubstituted or substituted C1-C6 alkyl, or an amino protecting group, or the hydrogen in —NHRx is absent and Rx is a divalent amino protecting group.

### Claim 17 (depends on 16)

The compound of claim 16, wherein Rx is —C(═O)Ph (Bz), —C(═O)CH3 (Ac) or —C(═O)CH(CH3)2 (iBu), or the hydrogen in —NHRx is absent and Rx is

### Claim 18 (depends on 1)

The compound of claim 1, selected from the group consisting of:
wherein R2 is H,
—C(═O)CH2CH2C(═O)OH or —C(═O)CH2CH2C(═O)NH2;
each
is optionally substituted with one to three substituent Q, and wherein each Q is independently selected from the group consisting of —F, —Cl, —Br, —CH3, —CH2CH3, —CF3, —CH3, —OCH3, and —OCH2CH3.

### Claim 19 (depends on 18)

The compound of claim 18, wherein B is
wherein Rx is —C(═O)Ph (Bz), —C(═O)CH3 (Ac) or —C(═O)CH(CH3)2 (iBu).

### Claim 20 (depends on 1)

A method of preparing a synthetic oligonucleotide, comprising reacting a compound of claim 1, with an oligonucleotide.

### Claim 21 (depends on 20)

An oligonucleotide or polynucleotide prepared by the method of claim 20.

### Claim 22

A method of deprotecting an oligonucleotide or polynucleotide comprising at least one 2′ protected nucleotide residue comprising the structure of Formula (II):
wherein
X is O or S;
R4 is H, unsubstituted C1-C6 alkyl or substituted C1-C6 alkyl;
B is a natural nucleobase, a modified natural nucleobase, or an unnatural nucleobase; and
RHet is optionally substituted 5-10 membered heteroaryl or optionally substituted 5-10 membered heterocyclyl;
the method comprising:
contacting the oligonucleotide or polynucleotide with a composition comprising an amine or ammonia to deprotect the 2′ protected nucleotide residue.

### Claim 23

A solid support bound oligonucleotide or polynucleotide comprising at least one 2′ protected nucleotide residue comprising the structure of Formula (II):
wherein
X is O or S;
R4 is H, unsubstituted C1-C6 alkyl or substituted C1-C6 alkyl;
B is a natural nucleobase, a modified natural nucleobase, or an unnatural nucleobase; and
RHet is optionally substituted 5-10 membered heteroaryl or optionally substituted 5-10 membered heterocyclyl.