Antisense antiviral compounds and methods for treating a filovirus infection
The present invention provides antisense antiviral compounds, compositions, and methods of their use and production, mainly for inhibiting the replication of viruses of the Filoviridae family, including Ebola and Marburg viruses. The compounds, compositions, and methods also relate to the treatment of viral infections in mammals including primates by Ebola and Marburg viruses. The antisense antiviral compounds include phosphorodiamidate morpholino oligonucleotides (PMOplus) having a nuclease resistant backbone, about 15-40 nucleotide bases, at least two but typically no more than half piperazine-containing intersubunit linkages, and a targeting sequence that is targeted against the AUG start site region of Ebola virus VP35, Ebola virus VP24, Marburg virus VP24, or Marburg virus NP, including combinations and mixtures thereof.
1. A method of treating a Marburg virus infection in a mammalian subject, comprising administering to the mammalian subject a therapeutically effective amount of a morpholino antisense oligonucleotide of 23 bases comprising the base sequence of SEQ ID NO:79, wherein the morpholino antisense oligonucleotide is linked to a polyethylene glycol moiety.
2. The method of claim 1 , wherein the morpholino antisense oligonucleotide is a phosphorodiamidate oligonucleotide.
3. The method of claim 1 , wherein at least two to no more than half of the total number of phosphorus-containing intersubunit linkages are positively charged.
4. The method of claim 3 , wherein the morpholino antisense oligonucleotide comprises positively charged phosphorus-containing intersubunit linkages between bases 10 and 11, bases 12 and 13, bases 14 and 15, bases 18 and 19, and bases 19 and 20 of SEQ ID NO:79.
5. The method of claim 1 , wherein the morpholino antisense oligonucleotide comprises phosphorus-containing intersubunit linkages in accordance with the structure:
wherein P j and P i are purine or pyrimidine base-pairing moieties effective to bind, by base-specific hydrogen bonding, to a base in a polynucleotide;
X is fluoro, alkyl, alkoxy, thioalkoxy, alkyl amino, a cyclic amine, 4-morpholine, 1-piperidine, 1-piperazine, or —NR 2 , wherein each R is independently H or lower alkyl;
Y 1 is O; and
Z is O.
6. The method of claim 5 , wherein X is NH 2 , NHR or NR 2 , wherein each R is a lower alkyl.
7. The method of claim 6 , wherein X is N(CH 3 ) 2 .
8. The method of claim 5 , wherein X is 1-piperazine for two to no more than half of the total number of phosphorus-containing intersubunit linkages.
9. The method of claim 5 , wherein X is 1-piperazine at phosphorus-containing intersubunit linkages between bases 10 and 11, bases 12 and 13, bases 14 and 15, bases 18 and 19, and bases 19 and 20 of SEQ ID NO:79, and X is N(CH 3 ) 2 for the remaining phosphorus-containing intersubunit linkages.
10. The method of claim 1 , wherein the morpholino antisense oligonucleotide is formulated as a composition comprising a pharmaceutically acceptable carrier.
11. The method of claim 1 , wherein the morpholino antisense oligonucleotide comprises a formula of:
12. The method of claim 11 , wherein the morpholino antisense oligonucleotide is formulated as a composition comprising a pharmaceutically acceptable carrier.
13. A method of vaccinating a mammalian subject against a Marburg virus, comprising administering to the mammalian subject an effective amount of a morpholino antisense oligonucleotide of 23 bases comprising the base sequence of SEQ ID NO:79, wherein the morpholino antisense oligonucleotide is linked to a polyethylene glycol moiety, and exposing the mammalian subject to an attenuated Marburg virus.
14. The method of claim 13 , wherein the morpholino antisense oligonucleotide is a phosphorodiamidate oligonucleotide.
15. The method of claim 13 , wherein at least two to no more than half of the total number of phosphorus-containing intersubunit linkages are positively charged.
16. The method of claim 13 , wherein the morpholino antisense oligonucleotide comprises positively charged phosphorus-containing intersubunit linkages between bases 10 and 11, bases 12 and 13, bases 14 and 15, bases 18 and 19, and bases 19 and 20 of SEQ ID NO:79.
17. The method of claim 13 , wherein the morpholino antisense oligonucleotide comprises phosphorus-containing intersubunit linkages in accordance with the structure:
wherein P j and P i are purine or pyrimidine base-pairing moieties effective to bind, by base-specific hydrogen bonding, to a base in a polynucleotide; and
X is fluoro, alkyl, alkoxy, thioalkoxy, alkyl amino, a cyclic amine, 4-morpholine, 1-piperidine, 1-piperazine, or —NR 2 , wherein each R is independently H or lower alkyl;
Y 1 is O; and
Z is O.
18. The method of claim 17 , wherein X is NH 2 , NHR or NR 2 , wherein each R is a lower alkyl.
19. The method of claim 18 , wherein X is N(CH 3 ) 2 .
20. The method of claim 17 , wherein X is 1-piperazine for two to no more than half of the total number of phosphorus-containing intersubunit linkages.
21. The method of claim 17 , wherein X is 1-piperazine at phosphorus-containing intersubunit linkages between bases 10 and 11, bases 12 and 13, bases 14 and 15, bases 18 and 19, and bases 19 and 20 of SEQ ID NO:79, and X is N(CH 3 ) 2 for the remaining phosphorus-containing intersubunit linkages.
22. The method of claim 13 , wherein the morpholino antisense oligonucleotide is formulated as a composition comprising a pharmaceutically acceptable carrier.
23. The method of claim 13 , wherein the morpholino antisense oligonucleotide comprises a formula of: