Antisense compounds and uses thereof
The present invention provides compounds comprising oligonucleotides complementary to a pyruvate kinase M transcript. Certain such compounds are useful for hybridizing to a pyruvate kinase M transcript, including but not limited to a pyruvate kinase M transcript in a cell. In certain embodiments, such hybridization results in modulation of splicing of the pyruvate kinase M transcript. In certain embodiments, such compounds are used to treat one or more symptoms associated with cancer.
1. A method of modulating splicing of a PK-M pre-mRNA transcript in a cell comprising contacting the cell with an antisense compound comprising a single-stranded modified oligonucleotide, wherein the single-stranded modified oligonucleotide consists of 15 to 25 linked nucleosides and has a nucleobase sequence comprising a complementary region, wherein the complementary region comprises at least 15 contiguous nucleobases and is 100% complementary to an equal-length portion of a target region of a PK-M transcript, wherein the target region of the PK-M transcript is within nucleobase 29153 and nucleobase 29281 of SEQ ID NO: 1, wherein each nucleoside of the single-stranded modified oligonucleotide comprises a modified sugar moiety, wherein the single-stranded modified oligonucleotide comprises 2′-deoxyribose unmodified sugar moieties, wherein the single-stranded modified oligonucleotide does not comprise more than 4 contiguous unmodified 2′-deoxynucleosides, and wherein inclusion of exon 9 of the PK-M transcript is increased and exclusion of exon 10 of the PK-M transcript is increased following contacting the cell with the antisense compound.
2. The method of claim 1 , wherein the nucleobase sequence of the single-stranded modified oligonucleotide is 100% complementary to an equal-length portion of the target region of the PK-M transcript.
3. The method of claim 1 , wherein the single-stranded modified oligonucleotide consists of 15 to 18 linked nucleosides.
4. The method of claim 1 , wherein at least one modified sugar moiety is a 2′-O-methoxyethyl (2′-MOE) modified sugar moiety.
5. The method of claim 1 , wherein at least one modified sugar moiety is a bicyclic modified sugar moiety.
6. The method of claim 5 , wherein the at least one bicyclic modified sugar moiety is a LNA or cEt sugar moiety.
7. The method of claim 1 , wherein each modified sugar moiety comprises the same modification.
8. The method of claim 1 , wherein each modified sugar moiety is independently selected from a 2′-MOE modified sugar moiety and a bicyclic modified sugar moiety.
9. The method of claim 8 , wherein the single-stranded modified oligonucleotide comprises a sugar motif of keekeekeekeek or kkeekeekeekeek, wherein each “k” represents a cEt bicyclic modified sugar moiety and each “e” represents a 2′-MOE modified sugar moiety.
10. The method of claim 1 , wherein the single-stranded modified oligonucleotide comprises at least one modified internucleoside linkage.
11. The method of claim 10 , wherein the at least one modified internucleoside linkage is a phosphorothioate internucleoside linkage.
12. The method of claim 10 , wherein each internucleoside linkage of the single-stranded modified oligonucleotide is a phosphorothioate internucleoside linkage.
13. The method of claim 1 , wherein the nucleobase sequence of the single-stranded modified oligonucleotide comprises a sequence as set forth in any one of SEQ ID NOs: 4 to 36.
14. The method of claim 1 , wherein the amount of PK-M2 mRNA is decreased following contacting the cell with the antisense compound.
15. The method of claim 1 , wherein the cell is in an animal.
16. The method of claim 15 , wherein the method comprises administering the antisense compound into the central nervous system of the animal.
17. The method of claim 15 , wherein the animal has one or more symptoms associated with cancer.
18. The method of claim 17 , wherein the cancer is glioblastoma.
19. The method of claim 15 , wherein the animal is a human.
20. A method of modulating splicing of a PK-M pre-mRNA transcript in a cell comprising contacting the cell with an antisense compound comprising a single-stranded modified oligonucleotide, wherein the single-stranded modified oligonucleotide consists of 15 to 25 linked nucleosides and has a nucleobase sequence comprising a complementary region, wherein the complementary region comprises at least 15 contiguous nucleobases and is 100% complementary to an equal-length portion of a target region of a PK-M transcript, wherein the target region of the PK-M transcript is within nucleobase 29153 and nucleobase 29281 of SEQ ID NO: 1, wherein the single-stranded modified oligonucleotide comprises a sugar motif of kddkddkddkddk or kkddkddkddkddk, each “k” represents a cEt bicyclic modified sugar moiety and each “d” represents a 2′-deoxyribose unmodified sugar moiety, wherein the single-stranded modified oligonucleotide does not comprise more than 4 contiguous unmodified 2′-deoxynucleosides, and wherein inclusion of exon 9 of the PK-M transcript is increased and exclusion of exon 10 of the PK-M transcript is increased following contacting the cell with the antisense compound.