Immunomodulatory agents and methods of use

• US 7,989,608 B2
• Filed: 12/24/2008
• Issued: 08/02/2011
• Est. Priority Date: 12/28/2007
• Status: Active Grant

• Current Assignee: Sarepta Therapeutics, Inc.
• Original Assignee: Avi Biopharma, Inc.
• Inventors: Iversen, Patrick L., Mourich, Dan V.
• Primary Examiner(s): Navarro; Albert
• Application Number:
  US12/344,143
• Publication Number:
  US 20090246221A1
• Time in Patent Office: 951 Days
• Field of Search: None
• US Class Current: 536/23.1
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 37/04 Immunostimulants
• CPC Class Codes: C12N 15/1136 against growth factors, growth regulators, cytokines, lymphokines or hormones
• CPC Class Codes: C12N 2310/11 Antisense
• CPC Class Codes: C12N 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2310/3233 Morpholino-type ring

Abstract

An antisense oligonucleotide compound, composition, vaccine and methods for treating a variety of conditions characterized by up-regulation of IL-10 in a mammalian subject are disclosed. The compound (i) is composed of morpholino subunits and phosphorus-containing intersubunit linkages joining a morpholino nitrogen of one subunit to a 5′ exocyclic carbon of an adjacent subunit, (ii) is capable of uptake by monocytes, lymphocytes, and dendritic cells in a mammalian subject, (iii) contains between 10-40 nucleotide bases, and (iv) has a base sequence effective to hybridize to at least 12 contiguous bases of a target sequence contained in an exon-2 or exon-4 slice site region of human IL-10 pre-mRNA.

Claims

1. A method of treating a mammalian subject infected with a pathogen which acts to up-regulate IL-10 during infection in a mammalian host, as evidenced by increased serum levels of IL-10, comprising administering to the subject, a therapeutically effective amount of an antisense composition containing an antisense oligonucleotide compound (i) composed of morpholino subunits and phosphorus-containing intersubunit linkages joining a morpholino nitrogen of one subunit to a 5′

exocyclic carbon of an adjacent subunit, (ii) capable of uptake by monocytes, lymphocytes, and dendritic cells in a mammalian subject, (iii) containing between 10-40 nucleotide bases, and (iv) having a targeting sequence complementary to at least 12 contiguous bases of the 5′

-most 25 bases of exon 2 or exon 4 of the preprocessed human IL-10 transcript contained within SEQ ID NO;

6, and identified by SEQ ID NOS;

2 and 5, respectively.

2. The method of claim 1, wherein the targeting sequence is complementary to at least 12 contiguous bases of SEQ ID NO:

5 in SEQ ID NO;

6.

3. The method of claim 1, wherein the targeting sequence is complementary to at least 12 contiguous bases of SEQ ID NO:

2 in SEQ ID NO;

6.

4. The method of claim 1, wherein the target sequence is contained entirely within SEQ ID NOS:

2 or 5.

5. The method of claim 1, wherein the composition administered contains a second oligonucleotide antisense compound having a targeting sequence complementary to at least 12 contiguous bases of another splice-site target sequence of the preprocessed human IL-10 transcript contained within SEQ ID NO:

6.

6. The method of claim 1, wherein the antisense oligonucleotide compound which is administered is conjugated to an arginine-rich polypeptide effective to promote uptake of the compound into monocytes, lymphocytes, and dendritic cells.

7. The method of claim 1 for use in treating a mammalian subject infected with a viral pathogen, which further includes administering to the subject, an anti-viral compound effective to inhibit replication of the viral pathogen in the mammalian host.

8. The method of claim 1 for use in treating a mammalian subject infected with a bacterial pathogen, which further includes administering to the subject, an anti-bacterial compound effective to inhibit replication of the bacterial pathogen in the mammalian host.

9. A pharmaceutical composition for treatment of a pathogen which acts to up-regulate IL-10 during infection in a mammalian host, as evidenced by increased serum levels of IL-10, comprising (a) a pathogen antigenic component capable of eliciting an immune response against the pathogen, (b) an antisense oligonucleotide compound (i) composed of morpholino subunits and phosphorus-containing intersubunit linkages joining a morpholino nitrogen of one subunit to a 5′

exocyclic carbon of an adjacent subunit, (ii) capable of uptake by monocytes, lymphocytes, and dendritic cells in a mammalian subject, (iii) containing between 10-40 nucleotide bases, and (iv) having a targeting sequence complementary to at least 12 contiguous bases of the 5′

-most 25 bases of exon 2 or exon 4 of the preprocessed human IL-10 transcript contained within SEQ ID NO;

6, and identified by SEQ ID NOS;

2 and 5, respectively, and (c) an adjuvant in which the antigenic component and antisense compound are formulated.

10. The composition of claim 9, which further includes a second antisense oligonucleotide compound having a targeting sequence complementary to at least 12 contiguous bases of another splice-site target sequence of the preprocessed human IL-10 transcript contained within SEQ ID NO:

6.

11. The composition of claim 9, wherein the antisense oligonucleotide compound is conjugated to an arginine-rich polypeptide effective to promote uptake of the compound into monocytes, lymphocytes, and dendritic cells.

12. The composition of claim 9, wherein the arginine-rich polypeptide has the sequence defined by SEQ ID NO:

36 or SEQ ID NO;

40.

13. The composition of claim 9, wherein the intersubunit linkages linking the morpholino subunits are phosphorodiamidate linkages having the structure:

14. An antisense oligonucleotide compound (i) composed of morpholino subunits and phosphorus-containing intersubunit linkages joining a morpholino nitrogen of one subunit to a 5′

exocyclic carbon of an adjacent subunit, (ii) capable of uptake by monocytes, lymphocytes, and dendritic cells in a mammalian subject, (iii) containing between 10-40 nucleotide bases, and (iv) having a targeting sequence complementary to at least 12 contiguous bases of a target sequence composed of 5′

-end 25 bases of exon 2 or exon 4 of the preprocessed human IL-10 transcript contained within SEQ ID NO;

6, and identified by SEQ ID NOS;

2 and 5, respectively.

15. The compound of claim 14, wherein the targeting sequence is complementary to at least 12 contiguous bases of SEQ ID NO:

5 in SEQ ID NO;

6.

16. The compound of claim 14, wherein the targeting sequence is complementary to at least 12 contiguous bases of SEQ ID NO:

2 in SEQ ID NO;

6.

17. The compound of claim 14, wherein the target sequence is contained entirely within SEQ ID NOS:

2 or 5.

18. The compound of claim 14, which is conjugated to an arginine-rich polypeptide effective to promote uptake of the compound into monocytes, lymphocytes, and dendritic cells.

19. The compound of claim 18, wherein the arginine-rich polypeptide has the sequence defined by SEQ ID NO:

36 or SEQ ID NO;

40.

20. The compound of claim 14, wherein the intersubunit linkages linking the morpholino subunits are phosphorodiamidate linkages having the structure:

21. The compound of claim 20, wherein the intersubunit linkages are uncharged linkages interspersed with piperazine-containing linkages, where the total number of piperazine-containing linkages is between 2 and no more than half of the total number of linkages.

22. The compound of claim 21, wherein the piperazine-containing linkages have the structure X is 1-piperazinyl.