Antisense composition and method for treating muscle atrophy

• US 8,785,410 B2
• Filed: 01/03/2011
• Issued: 07/22/2014
• Est. Priority Date: 02/09/2005
• Status: Active Grant

• Current Assignee: Sarepta Therapeutics, Inc.
• Original Assignee: Sarepta Therapeutics, Inc.
• Inventors: Iversen, Patrick L., Weller, Dwight D., Timmins, Alan P.
• Primary Examiner(s): ZARA, JANE J
• Application Number:
  US12/983,798
• Publication Number:
  US 20110166082A1
• Time in Patent Office: 1,296 Days
• Field of Search: 435/6, 435/1, 435/6.16, 435/91.1, 435/455, 514/1.1, 514/44, 536/23.1, 536/23.5, 536/24.5
• US Class Current: 514/44
• CPC Class Codes: A61K 31/675 having nitrogen as a ring hetero atom, e.g. pyridoxal phosphate
• CPC Class Codes: A61K 47/645 Polycationic or polyanionic oligopeptides, polypeptides or polyamino acids, e.g. polylysine, polyarginine, polyglutamic acid or peptide TAT
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C07K 7/08 having 12 to 20 amino acids gastrins C07K14/595; somatostatins C07K14/655; melanotropins C07K14/68
• 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 2310/31 of the backbone
• CPC Class Codes: C12N 2310/3145 with the nitrogen in 3' or 5'-position
• CPC Class Codes: C12N 2310/3233 Morpholino-type ring
• CPC Class Codes: C12N 2310/3513 Protein; Peptide
• CPC Class Codes: C12N 2320/30 Special therapeutic applications
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C12Q 2600/158 Expression markers
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]

Abstract

A method and compound for treating skeletal muscle mass deficiency in a human subject are disclosed. The composition is an oligomer of morpholino subunits and phosphorus-containing intersubunit linkages joining a morpholino nitrogen of one subunit to a 5′ exocyclic carbon of an adjacent subunit, contains between 10-40 nucleotide bases, has a base sequence effective to hybridize to an expression-sensitive region of processed or preprocessed human myostatin RNA transcript, identified, in its processed form, by SEQ ID NO:6, and is capable of uptake by target muscle cells in the subject. In practicing the method, the compound is administered in an amount and at a dosage schedule to produce an overall reduction in the level of serum myostatin measured in the patient, and preferably to bring the myostatin level within the a range determined for normal, healthy individuals.

Claims

1. A method of treating skeletal muscle mass deficiency in a human subject comprising

(a) measuring blood or tissue levels of myostatin in the subject,

(b) administering to the subject, a myostatin-expression-inhibiting amount of a substantially uncharged antisense 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 target muscle cells in the subject, (iii) containing between 10-40 nucleotide bases, and (iv) having a base sequence that is complementary to at least 12 contiguous bases in a target region of preprocessed human myostatin RNA transcript, identified by SEQ ID NO;

13, wherein the antisense compound includes a base sequence as set forth in SEQ ID NO;

3 and is capable of effectively blocking production of a full-length processed myostatin transcript from a preprocessed myostatin transcript,

(c) by said administering, forming within target muscle cells in the subject, a base-paired heteroduplex structure composed of human myostatin RNA transcript and the antisense compound and having a Tm of dissociation of at least 45°

C., thereby inhibiting expression of myostatin in said cells,

(d) at a selected time following said administering, measuring a blood or tissue level of myostatin in the subject, and

(e) repeating said administering, using the myostatin levels measured in (d) to adjust the dose or dosing schedule of the amount of antisense compound administered, if necessary, so as to reduce measured levels of myostatin over those initially measured and to maintain such levels of myostatin measured in step (d) within a range determined for normal, healthy individuals.

2. The method of claim 1, wherein the myostatin value measured in step (a) is above a selected threshold for normal healthy people.

3. The method of claim 1, wherein the morpholino subunits in the compound administered are joined by phosphorodiamidate linkages, in accordance with the structure:

4. The method of claim 1, wherein the compound is administered as a conjugate of the compound and an arginine-rich peptide effective to promote uptake of the compound into muscle target cells.

5. The method of claim 2, in which at least 2 and no more than half of the total number of intersubunit linkages are positively charged at physiological pH.

6. The method of claim 5, wherein said morpholino subunits are joined by phosphorodiamidate linkages, in accordance with the structure:

7. The method of claim 4, wherein the arginine-rich peptide has one of the sequences identified as SEQ ID NOS:

7-9, and the arginine-rich peptide is covalently coupled at its C terminus to the 5′

end of the antisense compound.

8. The method of claim 1, wherein said administering and measuring is carried out for a selected period of at least 2 weeks.

9. The method of claim 1, wherein said administering is by oral administration.

10. An antisense composition for use in increasing skeletal muscle mass in a human subject, comprising a substantially uncharged antisense 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 target muscle cells in the subject,

(iii) containing between 10-40 nucleotide bases, and

(iv) having a base sequence that is complementary to at least 12 contiguous bases in a target region of preprocessed human myostatin transcript identified by SEQ ID NO;

13,

wherein the antisense compound includes a base sequence as set forth in SEQ ID NO;

3 and is capable of effectively blocking production of a full-length processed myostatin transcript from a preprocessed myostatin transcript.

11. The composition of claim 10, wherein the morpholino subunits in the compound are joined by phosphorodiamidate linkages, in accordance with the structure:

12. The composition of claim 10, which includes said compound conjugated to an arginine-rich peptide effective to promote uptake of the compound into target muscle cells.

13. The composition of claim 10, in which at least 2 and no more than half of the total number of intersubunit linkages are positively charged at physiological pH.

14. The composition of claim 13, wherein said morpholino subunits are joined by phosphorodiamidate linkages, in accordance with the structure:

15. The composition of claim 12, wherein the arginine-rich peptide has one of the sequences identified as SEQ ID NOS:

7-9.

16. The composition of claim 12, arginine-rich peptide is covalently coupled at its C terminus to the 5′

end of the antisense compound.