CN-121991948-A - Double-stranded oligonucleotide containing nucleotide and application thereof

CN121991948ACN 121991948 ACN121991948 ACN 121991948ACN-121991948-A

Abstract

The present disclosure provides a double-stranded oligonucleotide containing nucleotide and application thereof, which belongs to the technical field of nucleic acid medicines. The present disclosure provides for the treatment and/or prevention of pathological conditions or diseases caused by abnormal expression of specific genes in cardiomyocytes or adipocytes by introducing a nucleotide-like substance into a double-stranded oligonucleotide, which modified oligonucleotide molecule may enhance targeted delivery of the oligonucleotide drug to the myocardial tissue or adipose tissue.

Inventors

LI HAITAO
HUANG YUANYU
GAO YONGXIN

Assignees

北京炫景瑞医药科技有限公司

Dates

Publication Date: 20260508
Application Date: 20250923
Priority Date: 20241101

Claims (13)

1. A double-stranded oligonucleotide comprising a sense strand and an antisense strand, each strand having 17-23 nucleotides, and wherein the sense strand is complementary to the antisense strand or substantially complementary to form a duplex region, wherein the substantial complementarity means that the sense strand and the antisense strand do not mismatch more than 3 nucleotides in the duplex region, wherein the double-stranded oligonucleotide comprises at least two nucleotide-like sequences of formula I, or a tautomer thereof, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof: (I) Wherein " "Represents the site of attachment of the nucleotide-like sequence to an adjacent nucleotide; B is selected from a substituted or unsubstituted nucleobase A, U, G, C or T, if B contains a substituent group, each independently selected from halogen, C 1 -C 3 alkyl, C 1 -C 3 alkoxy or imido; L is selected from saturated or unsaturated C 14 -C 22 hydrocarbon groups; p is selected from 1 or 2; q is selected from 1 or 2; n is selected from 1 or 2; z is selected from hydroxyl or mercapto.
2. The double-stranded oligonucleotide of claim 1, wherein L is selected from saturated C 16 -C 20 hydrocarbyl groups, optionally L is selected from saturated C 16 、C 17 、C 18 、C 19 or C 20 hydrocarbyl groups; n is selected from 1, p is selected from 1, and q is selected from 1.
3. The double-stranded oligonucleotide of claim 2, wherein the nucleotide-like is selected from the group consisting of a structure represented by formula II, or a tautomer thereof, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof: (II) Wherein Z is selected from hydroxyl or mercapto; B is selected from unsubstituted nucleobase A, U, G, C or T; L is selected from saturated C 16 、C 18 or C 20 linear alkyl.
4. A double stranded oligonucleotide according to claim 3, wherein said nucleotide-like is each independently selected from the group consisting of the structures shown in NM1 or NM2, or a tautomer thereof, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof: (NM1)、 (NM2)。
5. The double-stranded oligonucleotide of claim 1, wherein the double-stranded oligonucleotide comprises two identical or non-identical nucleotide-like sequences and the nucleotide-like sequences are located on the sense strand, the nucleotide-like sequences being selected from any one of the following: 1) At least one nucleotide-like is located at the 5 'end or the 3' end of the sense strand; 2) One of the class of nucleotides is located at the 5 'end of the sense strand and the other class of nucleotides is located at the 3' end of the sense strand; 3) One nucleotide class is positioned at any position in the sense strand of the double-stranded region, and the other nucleotide class is positioned at the 5' -end of the sense strand; 4) One nucleotide class is positioned at any position in the sense strand of the double-stranded region, and the other nucleotide class is positioned at the 3' -end of the sense strand; 5) One of the class of nucleotides is located at any of positions 4-8 of the sense strand, starting at the 5 'end, and the other class of nucleotides is located at the 3' end of the sense strand.
6. The double-stranded oligonucleotide of claim 5, wherein the sense strand of the double-stranded oligonucleotide comprises two nucleotide classes, one at position 4 of the sense strand starting at the 5 'end and the other at the 3' end of the sense strand.
7. The double-stranded oligonucleotide according to any one of claims 1 to 6, wherein the nucleotides of the double-stranded oligonucleotide other than the nucleotide replaced by the nucleotide-like are each modified, each independently selected from the group consisting of a 2' -O-methyl modified nucleotide, a 2' -fluoro modified nucleotide, a 2' -O-methoxyethyl modified nucleotide or a 2', 2' -disubstituted modified nucleotide.
8. The double-stranded oligonucleotide of claim 7, wherein in the duplex region of the double-stranded oligonucleotide, except for the nucleotide replaced by the nucleotide-like sequence, at least 3 positions in nucleotides 7 to 10 of the sense strand, starting from the 5' end, are selected from 2' -fluoro modified nucleotides, and the nucleotides at the remaining positions are selected from 2' -O-methyl modified nucleotides; And, at least 4 of the nucleotides at positions 2, 6, 9, 12, 14 and 16 of the antisense strand, starting at the 5' end, are selected from 2' -fluoro modified nucleotides, and the nucleotide at position 15 is selected from 2' -O-methoxyethyl or 2' -O-methyl modified nucleotides, the nucleotides at the remaining positions being selected from 2' -O-methyl modified nucleotides; The double-stranded oligonucleotide also has at least one overhang end, wherein the overhang end is not positioned at the 5' -end of the antisense strand, the overhang end consists of 1-4 nucleotides, and the nucleotides constituting the overhang end are each independently selected from a structure shown in a formula (III), or a tautomer thereof, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof: Wherein Base represents nucleobase A, U, G, C or T; z represents a hydroxyl group or a mercapto group; R 1 and R 2 are each independently selected from H, halogen, optionally substituted C 1 -C 6 alkyl or optionally substituted C 1 -C 6 alkoxy; R 1 and R 2 are each independently selected from any one of: 1) R 1 and R 2 are both selected from H; 2) R 1 is selected from methoxy, and R 2 is selected from H; 3) R 1 and R 2 are both selected from F; 4) R 1 is selected from F, and R 2 is selected from methyl; 5) R 1 is selected from methoxy and R 2 is selected from methyl.
9. The double-stranded oligonucleotide of claim 8, wherein the 5 'end of the antisense strand comprises a phosphate or a phosphate mimetic selected from the group consisting of 5' - (E) -vinyl phosphonate, and wherein the sense strand and/or the antisense strand each independently comprises one or more phosphorothioate linkages.
10. The double-stranded oligonucleotide of claim 9, wherein the internucleoside linkage between the following two adjacent nucleotides of the sense strand is a phosphorothioate linkage: The sense strand is an internucleoside linkage between nucleotide 1 and nucleotide 2, starting at the 5' end, and, An internucleoside linkage between nucleotide 2 and nucleotide 3 of the sense strand, starting at the 5' end; And The internucleoside linkage between the following two adjacent nucleotides of the antisense strand is a phosphorothioate linkage: the antisense strand is an internucleoside linkage between nucleotide 1 and nucleotide 2, starting at the 5' end, and, The antisense strand is an internucleoside linkage between nucleotide 2 and nucleotide 3 starting at the 5' end, and, The antisense strand is an internucleoside linkage between nucleotide 1 and nucleotide 2, starting at the 3' end, and, An internucleoside linkage between nucleotide 2 and nucleotide 3 of said antisense strand starting at the 3' terminus; The antisense strand contains at least 3 phosphorothioate linkages in the duplex region and the sense strand contains at least 2 phosphorothioate linkages in the duplex region; At least one phosphorothioate linkage is present in the internucleoside linkage between the overhung end and the duplex region in the double-stranded oligonucleotide, and between the nucleotides in the overhung end; The internucleoside linkage between nucleotide 10 and nucleotide 11 of the antisense strand calculated from the 5' end is a phosphorothioate linkage; And/or, the internucleoside linkage between the nucleotide in the sense strand which is base-complementarily paired with the 10 th nucleotide in the sense strand calculated from the 5 'end and the nucleotide in the antisense strand which is base-complementarily paired with the 11 th nucleotide in the sense strand calculated from the 5' end is a phosphodiester linkage.
11. A pharmaceutical composition comprising the double-stranded oligonucleotide of any one of claims 1-10, and a pharmaceutically acceptable adjuvant.
12. Use of any of the following for the manufacture of a medicament for the prevention and/or treatment of a disease associated with abnormal expression of a specific gene of a target tissue: (I) The double-stranded oligonucleotide according to claim 1 to 10, and/or (II) the pharmaceutical composition of claim 11; The target tissue is selected from adipose tissue or myocardial tissue.
13. The use according to claim 12, wherein the disease associated with abnormal expression of adipose tissue-specific genes is selected from the group consisting of lipid metabolism disorders, hypertension, cardiovascular diseases, or disorders related to overweight; The disease associated with abnormal expression of a myocardial tissue specific gene is selected from the group consisting of obstructive hypertrophic cardiomyopathy, familial hypertrophic cardiomyopathy, heart failure with retained ejection fraction, atrial fibrillation, ventricular fibrillation, angina pectoris, myocardial infarction, heart failure with reduced ejection fraction, supraventricular tachycardia, hypertrophic cardiomyopathy, dilated cardiomyopathy, arrhythmia, and congestive heart failure.

Description

Double-stranded oligonucleotide containing nucleotide and application thereof Technical Field The present disclosure relates to the technical field of nucleic acid medicines, and in particular relates to a double-stranded oligonucleotide containing nucleotide and application thereof. Background Diseases associated with abnormal expression of adipose tissue-specific genes including lipid metabolism disorders, hypertension, cardiovascular diseases, obesity, etc. are increasing worldwide. At present, it is difficult to efficiently target the delivery of oligonucleotides to adipose tissue for the treatment of diseases, and thus, development of an oligonucleotide molecule and a method thereof that can target the delivery to adipose cells in vivo or have relatively high selectivity to adipose tissue cells has been desired. Disclosure of Invention The present disclosure provides a class of nucleotides (or nucleotide analogs), double-stranded oligonucleotides containing such nucleotides, and uses thereof. The present disclosure provides for the treatment and/or prevention of pathological conditions or diseases caused by abnormal expression of specific genes in cardiomyocytes or adipocytes by introducing a nucleotide-like substance into a double-stranded oligonucleotide, which modified oligonucleotide molecule may enhance targeted delivery of the oligonucleotide drug to the myocardial tissue or adipose tissue. The technical scheme is as follows: In a first aspect of the present disclosure, the present disclosure provides a double-stranded oligonucleotide comprising a sense strand and an antisense strand, each strand having 17-23 nucleotides, and the sense strand being complementary or substantially complementary to the antisense strand to form a duplex region, the substantial complementarity meaning that the sense strand and the antisense strand do not have more than 3 nucleotides mismatched at the duplex region, wherein the double-stranded oligonucleotide comprises at least two (preferably two or three) nucleotides of the type (Ns) shown in formula I, or a tautomer thereof, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof: (I) Wherein " "Represents the site of attachment of the nucleotide-like sequence to an adjacent nucleotide; B is selected from a substituted or unsubstituted nucleobase A, U, G, C or T, if B contains a substituent group, each independently selected from halogen, C 1-C3 alkyl, C 1-C3 alkoxy or imido; L is selected from saturated or unsaturated C 14-C22 hydrocarbon groups; p is selected from 1 or 2; q is selected from 1 or 2; n is selected from 1 or 2; z is selected from hydroxyl or mercapto. In some embodiments of the present disclosure, L is selected from saturated C 16-C20 hydrocarbyl groups. In some embodiments of the disclosure, L is selected from saturated C 16、C17、C18、C19 or C 20 hydrocarbyl groups. In some embodiments of the disclosure, n is selected from 1. In some embodiments of the disclosure, p is selected from 1 and q is selected from 1. In some embodiments of the present disclosure, the nucleotide-like is selected from the group consisting of structures represented by formula II, or a tautomer thereof, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof: (II) Wherein Z is selected from hydroxyl or mercapto; B is selected from unsubstituted nucleobase A, U, G, C or T; L is selected from saturated C 16、C18 or C 20 linear alkyl. In some embodiments of the disclosure, the nucleotide-like groups are each independently selected from the structures shown in NM1 or NM2, or a tautomer thereof, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof: (NM1)、(NM2)。 in some embodiments of the disclosure, the double-stranded oligonucleotide comprises two identical or non-identical nucleotide-like regions, and the nucleotide-like regions are located in the duplex region. In some embodiments of the disclosure, the nucleotide-like is located on the sense strand. In some embodiments of the disclosure, at least one nucleotide-like is located at the 5 'end or the 3' end of the sense strand. In some embodiments of the disclosure, one nucleotide class is located at the 5 'end of the sense strand and the other nucleotide class is located at the 3' end of the sense strand. In some embodiments of the disclosure, one nucleotide class is located at any position in the sense strand of the double-stranded region and the other nucleotide class is located at the 5' end of the sense strand. In some embodiments of the disclosure, one nucleotide class is located at any position in the sense strand of the double-stranded region and the other nucleotide class is located at the 3' end of the sense strand. In some embodiments of the present disclosure, the sense strand of the double-stranded oligonucleotide comprises two nucleotide classes, one at any of positions 4-8 of the sense strand, starting at the 5 'end, and the other at the 3' end of the sense strand.