Details for Patent: 9,534,220

Title:	Antisense antibacterial method and compound
Abstract:	A method and antisense compound for inhibiting the growth of pathogenic bacterial cells are disclosed. The compound contains no more than 12 nucleotide bases and has a targeting nucleic acid sequence of no fewer than 10 bases in length that is complementary to a target sequence containing or within 10 bases, in a downstream direction, of the translational start codon of a bacterial mRNA that encodes a bacterial protein essential for bacterial replication. The compound binds to a target mRNA with a T.sub.m of between 50.degree. to 60.degree. C. The relatively short antisense compounds are substantially more active than conventional antisense compounds having a targeting base sequence of 15 or more bases.
Inventor(s):	Geller; Bruce L. (Corvallis, OR), Deere; Jesse D. (Davis, CA), Iversen; Patrick L. (Corvallis, OR)
Assignee:	Sarepta Therapeutics, Inc. (Corvallis, OR)
Filing Date:	Nov 19, 2012
Application Number:	13/680,790
Claims:	1. A method of inhibiting the growth of pathogenic bacterial cells, comprising exposing the bacterial cells to a growth-inhibiting amount of a substantially uncharged antisense oligonucleotide compound of 10 to 12 bases in length having a targeting sequence of at least 10 bases that are complementary to a target sequence containing or within 10 bases downstream of the translational start codon of a bacterial mRNA that encodes a bacterial protein selected from the group consisting of acyl carrier protein (acpP), gyrase A subunit (gyrA), and the cell division protein ftsZ, wherein the antisense oligonucleotide compound is a morpholino antisense oligonucleotide. 2. The method of claim 1, wherein the morpholino subunits of the antisense oligonucleotide compound are joined by phosphorus-containing linkages, in accordance with the structure: ##STR00003## where Y.sub.1.dbd.O, Z.dbd.O, Pj is a purine or pyrimidine base-pairing moiety effective to bind, by base-specific hydrogen bonding, to a base in a polynucleotide, and X is alkyl, alkoxy, thioalkoxy, amino or alkyl amino, including dialkylamino. 3. The method of claim 1, wherein the targeting sequence is complementary to a target sequence containing the translational start codon of the bacterial mRNA. 4. The method of claim 1, wherein the targeting sequence is complementary to a target sequence that is within 10 bases downstream of the translational start codon of the bacterial mRNA. 5. The method of claim 1, wherein the antisense oligonucleotide compound contains 11 bases, and has a nucleic acid sequence that is completely complementary to the mRNA target sequence. 6. The method of claim 1, for use in inhibiting a bacterial infection in a mammalian subject, wherein said exposing includes administering the antisense oligonucleotide compound in a therapeutically effective amount. 7. The method of claim 6, further comprising the step of treating the subject by administration of a non-antisense compound having antibacterial activity. 8. A substantially uncharged antisense oligonucleotide compound of 10 to 12 bases in length having a targeting sequence of at least 10 bases that are complementary to a target sequence containing or within 10 bases downstream of the translational start codon of a bacterial mRNA that encodes a bacterial protein selected from the group consisting of acyl carrier protein (acpP), gyrase A subunit (gyrA), and the cell division protein ftsZ, wherein the antisense oligonucleotide compound is a morpholino antisense oligonucleotide. 9. The antisense oligonucleotide compound of claim 8, wherein the bacterial mRNA encodes the ftsZ protein, and the targeting sequence is complementary to at least ten contiguous bases in a sequence selected from the group consisting of SEQ ID NOS: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 33, 38, 41, 44, 47, 50, 53, and 56. 10. The antisense oligonucleotide compound of claim 8, wherein the bacterial mRNA encodes the acpP protein, and the targeting sequence is complementary to at least ten contiguous bases in a sequence selected from the group consisting of SEQ ID NOS: 2, 5, 8, 11, 14, 17, 23, 28, 31, 34, 36, 39, 42, 45, 48, 51, 54, and 57. 11. The antisense oligonucleotide compound of claim 8, wherein the bacterial mRNA encodes the gyrA protein, and the targeting sequence is complementary to at least ten contiguous bases in a sequence selected from the group consisting of SEQ ID NOS: 3, 6, 9, 12, 15, 18, 20, 24, 29, 32, 35, 37, 40, 43, 46, 49, 52, 55, and 58. 12. The antisense oligonucleotide compound of claim 8, wherein the morpholino subunits of the antisense oligonucleotide compound are joined by phosphorus-containing linkages, in accordance with the structure: ##STR00004## where Y.sub.1.dbd.O, Z.dbd.O, Pj is a purine or pyrimidine base-pairing moiety effective to bind, by base-specific hydrogen bonding, to a base in a polynucleotide, and X is alkyl, alkoxy, thioalkoxy, amino or alkyl amino, including dialkylamino. 13. The antisense oligonucleotide compound of claim 8, wherein the targeting sequence is complementary to a target sequence containing the translational start codon of the bacterial mRNA. 14. The antisense oligonucleotide compound of claim 8, wherein the targeting sequence is complementary to a target sequence that is within 10 bases downstream of the translational start codon of the bacterial mRNA. 15. The antisense oligonucleotide compound of claim 8, which contains 11 bases, and has a nucleic acid sequence that is completely complementary to the mRNA target sequence. 16. The antisense oligonucleotide compound of claim 8, which contains 10 bases, and has a nucleic acid sequence that is completely complementary to the mRNA target sequence. 17. The method of claim 1, wherein the bacterial mRNA encodes the ftsZ protein, and the targeting sequence is complementary to at least ten contiguous bases in a sequence selected from the group consisting of SEQ ID NOS: 1, 4, 7, 10, 13, 16, 19, 22, 27, 30, 33, 38, 41, 44, 47, 50, 53, and 56. 18. The method of claim 1, wherein the bacterial mRNA encodes the acpP protein, and the targeting sequence is complementary to at least ten contiguous bases in a sequence selected from the group consisting of SEQ ID NOS: 2, 5, 8, 11, 14, 17, 23, 28, 31, 34, 36, 39, 42, 45, 48, 51, 54, and 57. 19. The method of claim 1, wherein the bacterial mRNA encodes the gyrA protein, and the targeting sequence is complementary to at least ten contiguous bases in a sequence selected from the group consisting of SEQ ID NOS: 3, 6, 9, 12, 15, 18, 20, 24, 29, 32, 35, 37, 40, 43, 46, 49, 52, 55, and 58. 20. The method of claim 2, wherein the phosphorus-containing linkages are phosphorodiamidate linkages. 21. The method of claim 20, wherein each X is NR.sub.2 wherein each R is CH.sub.3. 22. The method of claim 2, wherein up to one of every five phosphorus-containing linkages is positively charged, wherein X is 1-piperazine. 23. The antisense oligonucleotide compound of claim 12, wherein the phosphorus-containing linkages are phosphorodiamidate linkages. 24. The antisense oligonucleotide compound of claim 23, wherein each X is NR.sub.2 wherein each R is CH.sub.3. 25. The antisense oligonucleotide compound of claim 12, wherein up to one of every five phosphorus-containing linkages is positively charged, wherein X is 1-piperazine. 26. An antisense oligonucleotide of 11 nucleotide bases for inhibiting the growth of pathogenic bacterial cells comprising a base sequence of CTTCGATAGTG (SEQ ID NO: 109) that is completely complementary to a bacterial mRNA that encodes the acyl carrier protein (acpP), wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide. 27. The antisense oligonucleotide of claim 26, wherein the antisense oligonucleotide is conjugated to a polyethylene glycol moiety. 28. An antisense oligonucleotide of 10 nucleotide bases for inhibiting the growth of pathogenic bacterial cells comprising a base sequence of TTCGATAGTG (SEQ ID NO: 110) that is completely complementary to a bacterial mRNA that encodes the acyl carrier protein (acpP), wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide. 29. The antisense oligonucleotide of claim 28, wherein the antisense oligonucleotide is conjugated to a polyethylene glycol moiety.