Follistatin in treating duchenne muscular dystrophy

1. A method of treating Duchenne muscular dystrophy (DMD) comprising administering to an individual who is suffering from or susceptible to DMD an effective amount of a recombinant follistatin protein comprising an amino acid sequence of SEQ ID NO: 1 or 2 fused to an Fc domain via a peptide linker comprising 10 or more amino acids such that at least one symptom or feature of DMD is reduced in intensity, severity, or frequency, or has delayed onset, and wherein the peptide linker comprises the amino acid sequence of SEQ ID NO: 5, 6, or 7.

2. The method of claim 1 , wherein the recombinant follistatin protein comprises a deletion of amino acids residues 212-288 of SEQ ID NO:1.

3. The method of claim 1 , wherein the Fc domain comprises an amino acid sequence of

(SEQ ID NO: 3)

EPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVV

DVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDW

LNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQ

VSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT

VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK.

4. The method of claim 1 , wherein the recombinant follistatin protein is produced from mammalian cells.

5. The method of claim 1 , wherein the recombinant follistatin protein is administered systemically.

6. The method of claim 1 , wherein the administration of the recombinant follistatin protein results in muscle regeneration, increased muscle strength, increased flexibility, increased range of motion, increased stamina, reduced fatigability, increased blood flow, improved cognition, improved pulmonary function, inflammation inhibition, reduced muscle fibrosis, and/or reduced muscle necrosis.

7. The method of claim 1 , wherein the at least one symptom or feature of DMD is selected from the group consisting of muscle wasting, muscle weakness, muscle fragility, muscle necrosis, muscle fibrosis, joint contracture, skeletal deformation, cardiomyopathy, impaired swallowing, impaired bowel and bladder function, muscle ischemia, cognitive impairment, behavioral dysfunction, socialization impairment, scoliosis, and impaired respiratory function.

8. A recombinant follistatin fusion protein comprising

a follistatin polypeptide;

an Fc domain; and

a linker that associates the follistatin polypeptide with the Fc domain,

wherein the follistatin polypeptide comprises an amino acid sequence of SEQ ID NO.: 1 or 2; wherein the linker comprises the amino acid sequence of SEQ ID NO: 5, 6, or 7; and further wherein the recombinant follistatin fusion protein is capable of binding to activin, myostatin and/or GDF-11 and has an in vivo half-life ranging from about 0.5-10 days.

9. The recombinant follistatin fusion protein of claim 8 , wherein the follistatin polypeptide contains a deletion of amino acids residues 212-288 of SEQ ID NO:1.

10. The recombinant follistatin fusion protein of claim 8 , wherein the Fc domain is an IgG1 Fc domain.

11. The recombinant follistatin fusion protein of claim 8 , wherein the Fc domain has an amino acid sequence of

(SEQ ID NO: 3)

EPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVV

DVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDW

LNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQ

VSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT

VDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK.

12. The recombinant follistatin fusion protein of claim 8 , wherein the recombinant follistatin fusion protein comprises an amino acid sequence of

SEQ ID NO: 8

GNCWLRQAKNGRCQVLYKTELSKEECCSTGRLSTSWTEEDVNDNTLFKW

MIFNGGAPNCIPCKETCENVDCGPGKKCRMNKKNKPRCVCAPDCSNITW

KGPVCGLDGKTYRNECALLKARCKEQPELEVQYQGRCKKTCRDVFCPGS

STCVVDQTNNAYCVTCNRICPEPASSEQYLCGNDGVTYSSACHLRKATC

LLGRSIGLAYEGKCIKAKSCEDIQCTGGKKCLWDFKVGRGRCSLCDELC

PDSKSDEPVCASDNATYASECAMKEAACSSGVLLEVKHSGSCNSISEDT

EEEEEDEDQDYSFPISSILEWGAPGGGGGAAAAAGGGGGGAPGGGGGAA

AAAGGGGGGAPGGGGGAAAAAGGGGGGAPKTHTCPPCPAPELLGGPSVF

LFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTK

PREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKA

KGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPE

NNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYT

QKSLSLSPGK (SEQ ID NO: 8),

or

SEQ ID NO: 9

GNCWLRQAKNGRCQVLYKTELSKEECCSTGRLSTSWTEEDVNDNTLFKW

MIFNGGAPNCIPCKETCENVDCGPGKKCRMNKKNKPRCVCAPDCSNITW

KGPVCGLDGKTYRNECALLKARCKEQPELEVQYQGRCKKTCRDVFCPGS

STCVVDQTNNAYCVTCNRICPEPASSEQYLCGNDGVTYSSACHLRKATC

LLGRSIGLAYEGKCIKAKSCEDIQCTGGKKCLWDFKVGRGRCSLCDELC

PDSKSDEPVCASDNATYASECAMKEAACSSGVLLEVKHSGSCNSISEDT

EEEEEDEDQDYSFPISSILEWGAPGGGGGAAAAAGGGGGGAPGGGGGAA

AAAGGGGGGAPGGGGGAAAAAGGGGGGAPEPKSCDKTHTCPPCPAPELL

GGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEV

HNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIE

KTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWE

SNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEA

LHNHYTQKSLSLSPGK (SEQ ID NO: 9).

13. The recombinant follistatin fusion protein of claim 8 , wherein the recombinant follistatin fusion protein comprises an amino acid sequence of

SEQ ID NO: 10

GNCWLRQAKNGRCQVLYKTELSKEECCSTGRLSTSWTEEDVNDNTLFKW

MIFNGGAPNCIPCKETCENVDCGPGKKCRMNKKNKPRCVCAPDCSNITW

KGPVCGLDGKTYRNECALLKARCKEQPELEVQYQGRCKKTCRDVFCPGS

STCVVDQTNNAYCVTCNRICPEPASSEQYLCGNDGVTYSSACHLRKATC

LLGRSIGLAYEGKCISISEDTEEEEEDEDQDYSFPISSILEWGAPGGGG

GAAAAAGGGGGGAPGGGGGAAAAAGGGGGGAPGGGGGAAAAAGGGGGGA

PKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHE

DPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKE

YKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTC

LVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSR

WQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 10),

or

SEQ ID NO: 11

GNCWLRQAKNGRCQVLYKTELSKEECCSTGRLSTSWTEEDVNDNTLFKW

MIFNGGAPNCIPCKETCENVDCGPGKKCRMNKKNKPRCVCAPDCSNITW

KGPVCGLDGKTYRNECALLKARCKEQPELEVQYQGRCKKTCRDVFCPGS

STCVVDQTNNAYCVTCNRICPEPASSEQYLCGNDGVTYSSACHLRKATC

LLGRSIGLAYEGKCISISEDTEEEEEDEDQDYSFPISSILEWGAPGGGG

GAAAAAGGGGGGAPGGGGGAAAAAGGGGGGAPGGGGGAAAAAGGGGGGA

PEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVV

VDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQD

WLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKN

QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKL

TVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 11).

14. A nucleic acid comprising a nucleotide sequence encoding the recombinant follistatin fusion protein of claim 8 .

15. A cell comprising a nucleic acid of claim 14 .

16. A pharmaceutical composition comprising a recombinant follistatin fusion protein of claim 8 and a pharmaceutically acceptable carrier.

17. A method of treating Duchenne muscular dystrophy (DMD) comprising administering to an individual who is suffering from or susceptible to DMD an effective amount of a recombinant follistatin protein comprising an amino acid sequence SEQ ID NOs: 8, 9, 10 or 11, such that at least one symptom or feature of DMD is reduced in intensity, severity, or frequency, or has delayed onset.