US2022127326A1PendingUtilityA1
Targeted Therapeutic Lysosomal Enzyme Fusion Proteins and Uses Thereof
Est. expiryNov 27, 2032(~6.4 yrs left)· nominal 20-yr term from priority
Inventors:Mika Aoyagi-ScharberTeresa Margaret ChristiansonMelita Dvorak-EwellDaniel J. WendtShinong LongJonathan LebowitzDaniel Solomon Gold
C12Y 302/0105C12N 9/2402C07K 14/65A61K 38/47A61P 3/00A61K 38/00C07K 2319/06A61P 43/00
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Claims
Abstract
The present invention relates in general to therapeutic fusion proteins useful to treat lysosomal storage diseases and methods for treating such diseases. Exemplary therapeutic fusion proteins comprise a lysosomal enzyme, a lysosomal targeting moiety, e.g., an IGF-II peptide, and a spacer peptide. Also provided are compositions and methods for treating Mucopolysaccharidosis Type IIIB (Sanfilippo B Syndrome), comprising a targeted therapeutic fusion protein comprising alpha-N-acetylglucosaminidase (Naglu), a lysosomal targeting moiety, e.g., an IGF-II peptide, and a spacer peptide.
Claims
exact text as granted — not AI-modifiedWhat is claimed:
1 . A targeted therapeutic fusion protein comprising (a) a lysosomal enzyme, (b) a peptide tag having an amino acid sequence at least 70% identical to amino acids 8-67 of mature human IGF-II and (c) a spacer peptide between the lysosomal enzyme and the IGF-II peptide tag, wherein the spacer peptide comprises one or more GGGPS (SEQ ID NO: 14) or GGGSP (SEQ ID NO: 15) amino acid sequences, and optionally further comprises one or more of (i) GAP (SEQ ID NO: 9), (ii) GGGGS (SEQ ID NO: 12), (iii) GGGS (SEQ ID NO: 16), (iv) AAAAS (SEQ ID NO: 17), (v) AAAS (SEQ ID NO: 18), (vi) PAPA (SEQ ID NO: 19), (vii) TPAPA (SEQ ID NO: 20), (viii) AAAKE (SEQ ID NO: 21) or (ix) GGGGA (SEQ ID NO: 60).
2 . The targeted therapeutic fusion protein of claim 1 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of: (GGGGS) n (SEQ ID NOs: 12, 56, 58, 91-94), (GGGGS) n -GGGPS (SEQ ID NOs: 36, 95-100), GAP-(GGGGS) n -GGGPS (SEQ ID NOs: 101-107), GAP-(GGGGS) n -GGGPS-GAP (SEQ ID NOs: 37, 108-113), GAP-(GGGGS) n -GGGPS-(GGGGS) n -GAP (SEQ ID NOs: 114-162), GAP-GGGPS-(GGGGS) n -GAP (SEQ ID NOs: 163-169), GAP-(GGGGS) n -AAAAS-GGGPS-(GGGGS) n -AAAA-GAP (SEQ ID NOs: 170-218), GAP-(GGGGS) n -PAPAP-(Xaa) n -GAP (SEQ ID NOs: 219-267), GAP-(GGGGS) n -PAPAPT-(Xaa) n -GAP (SEQ ID NOs: 268-316), GAP-(GGGGS) n -(Xaa) n -PAPAP-(Xaa) n -(AAAKE) n -(Xaa) n -(GGGGS) n -GAP (SEQ ID NOs: 544-551), (GGGGA) n (SEQ ID NOs: 60, 79, 81, 317-320), (GGGGA) n -GGGPS (SEQ ID NOs: 321-326), GAP-(GGGGA) n -GGGPS (SEQ ID NOs: 327-333), GAP-(GGGGA) n -GGGPS-GAP (SEQ ID NOs: 334-340), GAP-(GGGGA) n -GGGPS-(GGGGA) n -GAP (SEQ ID NOs: 341-389), GAP-GGGPS-(GGGGA) n -GAP (SEQ ID NOs: 390-396), GAP-(GGGGA) n -AAAAS-GGGPS-(GGGGA) n -AAAA-GAP (SEQ ID NOs: 397-445), GAP-(GGGGA) n -PAPAP-(Xaa) n -GAP (SEQ ID NOs: 446-494), GAP-(GGGGA) n -PAPAPT-(Xaa) n -GAP (SEQ ID NOs: 495-543), GAP-(GGGGA) n -(Xaa) n -PAPAP-(Xaa) n -(AAAKE) n -(Xaa) n -(GGGGA) n -GAP (SEQ ID NOs: 552-559); wherein n is 1 to 7.
3 . The targeted therapeutic fusion protein of claim 2 , wherein n is 1 to 4.
4 . The targeted therapeutic fusion protein of claim 1 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of EFGGGGSTR (SEQ ID NO: 22), GAP (SEQ ID NO: 9), GGGGS (SEQ ID NO: 12), GPSGSPG (SEQ ID NO: 23), GPSGSPGT (SEQ ID NO: 24), GPSGSPGH (SEQ ID NO: 25), GGGGSGGGGSGGGGSGGGGSGGGPST (SEQ ID NO: 26), GGGGSGGGGSGGGGSGGGGSGGGPSH (SEQ ID NO: 27), GGGGSGGGGSGGGGSGGGGSGGGPSGGGGSGGGPS (SEQ ID NO: 28), GAPGGGGSGGGGSGGGGSGGGGSGGGPSGGGGSGGGPSGAP (SEQ ID NO: 29), GGGGS GGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGGS GGGGS GGGPS (SEQ ID NO: 30), GAPGGGGS GGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGGS GGGGS GGGPS GAP (SEQ ID NO: 31), GGGGSGGGGSGGGGSGGGPSGGGGSGGGGSGGGPS (SEQ ID NO: 32), GAPGGGGSGGGGSGGGGSGGGPSGGGGSGGGGSGGGPSGAP (SEQ ID NO: 33), GGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS (SEQ ID NO: 34), GAPGGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS GA P (SEQ ID NO: 35), GGGGSGGGGSGGGGSGGGGSGGGPS (SEQ ID NO: 36), GAPGGGGSGGGGSGGGGSGGGGSGGGPSGAP (SEQ ID NO: 37), GGGGSGGGGSAAAASGGGGSGGGPS (SEQ ID NO: 38), GAPGGGGSGGGGSAAAASGGGGSGGGPSGAP (SEQ ID NO: 39), GGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGPS (SEQ ID NO: 40), GAPGGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGPSG AP (SEQ ID NO: 41), GGGGSGGGGSAAAASGGGPSGGGGSAAAASGGGPSGGGGSAAAASGGGPS (SEQ ID NO: 42), GAP GGGGSGGGGSAAAASGGGPSGGGGSAAAASGGGPSGGGGSAAAASGGGPSGA P (SEQ ID NO: 43), GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPS (SEQ ID NO: 44), GAPGGSPAGSPTSTEEGTSESATPES GPGTSTEPSEGSAPGSPAGSPGPS GAP (SEQ ID NO: 45), GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPS (SEQ ID NO: 46), GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPSGAP (SEQ ID NO: 47), GGGSPAPTPTPAPTPAPTPAGGGPS (SEQ ID NO: 48), GAPGGGSPAPTPTPAPTPAPTPAGGGPSGAP (SEQ ID NO: 49), GGGSPAPAPTPAPAPTPAPAGGGPS (SEQ ID NO: 50), GAPGGGSPAPAPTPAPAPTPAPAGGGPS GAP (SEQ ID NO: 51), GGGSAEAAAKEAAAKEAAAKAGGPS (SEQ ID NO: 52), GAPGGGSAEAAAKEAAAKEAAAKAGGPSGAP (SEQ ID NO: 53), GGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGG (SEQ ID NO: 54), GAPGGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGGGAP (SEQ ID NO: 55), GGGGSGGGGSGGGGS (SEQ ID NO: 56), GAPGGGGSGGGGSGGGGS GAP (SEQ ID NO: 57), GGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 58), GAPGGGGSGGGGSGGGGSGGGGSGAP (SEQ ID NO: 59), GGGGA (SEQ ID NO: 60), GGGGAGGGGAGGGGAGGGGAGGGPST (SEQ ID NO: 61), GGGGAGGGGAGGGGAGGGGAGGGPSH (SEQ ID NO: 62), GGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGPS (SEQ ID NO: 63), GAPGGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGPSGAP (SEQ ID NO: 64), GGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGGAGGGGAGGGPS (SEQ ID NO: 65), GAP GGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGGAGGGGAGGGPS GAP (SEQ ID NO:66), GGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPS (SEQ ID NO: 67), GAPGGGGAGGGGAGGGGAGGGPS GGGGAGGGGAGGGPS GAP (SEQ ID NO: 68), GGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPSGGGGAGGGGAGGGPS (SEQ ID NO: 69), GAP GGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPSGGGGAGGGGAGGGPS GAP (SEQ ID NO: 70), GGGGAGGGGAGGGGAGGGGAGGGPS (SEQ ID NO: 71), GAPGGGGAGGGGAGGGGAGGGGAGGGPSGAP (SEQ ID NO: 72), GGGGAGGGGAAAAASGGGGAGGGPS (SEQ ID NO: 73), GAPGGGGAGGGGAAAAASGGGGAGGGPSGAP (SEQ ID NO: 74), GGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGPS (SEQ ID NO: 75), GAP GGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGPS GAP (SEQ ID NO: 76), GGGGAGGGGAAAAASGGGPSGGGGAAAAASGGGPSGGGGAAAAASGGGPS (SEQ ID NO: 77), GAPGGGGAGGGGAAAAASGGGPSGGGGAAAAASGGGPSGGGGAAAAASGGGPSG AP (SEQ ID NO: 78), GGGGAGGGGAGGGGA (SEQ ID NO: 79), GAPGGGGAGGGGAGGGGAGAP (SEQ ID NO: 80), GGGGAGGGGAGGGGAGGGGA (SEQ ID NO: 81), GAPGGGGAGGGGAGGGGAGGGGAGAP (SEQ ID NO: 82), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPS [or (GGGGA)8GGGPS] (SEQ ID NO: 83), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPSH [or (GGGGA)8GGGPSH] (SEQ ID NO: 84), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPS [or (GGGGA)9GGGPS] (SEQ ID NO: 85), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPSH [or (GGGGA)9GGGPSH] (SEQ ID NO: 86), GGGGPAPGPGPAPGPAPGPAGGGPS (SEQ ID NO: 87), GAPGGGGPAPGPGPAPGPAPGPAGGGPGGAP (SEQ ID NO: 88), GGGGPAPAPGPAPAPGPAPAGGGPS (SEQ ID NO: 89), and GAPGGGGPAPAPGPAPAPGPAPAGGGPGGAP (SEQ ID NO: 90).
5 . The targeted therapeutic fusion protein of claim 4 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of
(SEQ ID NO: 36)
GGGGSGGGGSGGGGSGGGGSGGGPS,
(SEQ ID NO: 44)
GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPS,
(SEQ ID NO: 45)
GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPSG
AP,
(SEQ ID NO: 46)
GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPS,
(SEQ ID NO: 47)
GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTG
PSGAP,
(SEQ ID NO: 48)
GGGSPAPTPTPAPTPAPTPAGGGPS,
(SEQ ID NO: 49)
GAPGGGSPAPTPTPAPTPAPTPAGGGPSGAP,
(SEQ ID NO: 50)
GGGSPAPAPTPAPAPTPAPAGGGPS,
(SEQ ID NO: 51)
GAPGGGSPAPAPTPAPAPTPAPAGGGPSGAP,
(SEQ ID NO: 52)
GGGSAEAAAKEAAAKEAAAKAGGPS,
(SEQ ID NO: 53)
GAPGGGSAEAAAKEAAAKEAAAKAGGPSGAP,
(SEQ ID NO: 54)
GGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGG,
(SEQ ID NO: 55)
GAPGGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGGGAP,
and
(SEQ ID NO: 71)
GGGGAGGGGAGGGGAGGGGAGGGPS.
6 . The targeted therapeutic fusion protein of claim 5 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of
(SEQ ID NO: 36)
GGGGSGGGGSGGGGSGGGGSGGGPS,
(SEQ ID NO: 47)
GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTG
PSGAP,
(SEQ ID NO: 51)
GAPGGGSPAPAPTPAPAPTPAPAGGGPSGAP,
(SEQ ID NO: 55)
GAPGGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGGGAP,
and
(SEQ ID NO: 71)
GGGGAGGGGAGGGGAGGGGAGGGPS.
7 . A targeted therapeutic fusion protein comprising an amino acid sequence at least 85% identical to a human α-N-acetylglucosaminidase (Naglu) protein, a peptide tag having an amino acid sequence at least 70% identical to amino acids 8-67 of mature human IGF-II and a spacer peptide located between the Naglu amino acid sequence and the IGF-II peptide tag, wherein the spacer comprises the amino acid sequence GAP (SEQ ID NO: 9), GPS (SEQ ID NO: 10), or GGS (SEQ ID NO: 11).
8 . The targeted therapeutic fusion protein of claim 7 , wherein the spacer sequence comprises amino acids Gly-Pro-Ser (GPS) (SEQ ID NO: 10) between the amino acids of mature human IGF-II and the amino acids of human Naglu.
9 . The targeted therapeutic fusion protein of any of claims 7 to 8 , wherein the spacer peptide comprises one or more GGGGS (SEQ ID NO: 12), GGGGA (SEQ ID NO: 60) or GGGS (SEQ ID NO: 16) amino acid sequences.
10 . The targeted therapeutic fusion protein of any of claims 7 to 9 , wherein the spacer peptide comprises one or more GGGPS (SEQ ID NO: 14) or GGGSP (SEQ ID NO: 15) amino acid sequences.
11 . The targeted therapeutic fusion protein of any of claims 7 to 10 , wherein the spacer peptide comprises one or more AAAAS (SEQ ID NO: 17) or AAAS (SEQ ID NO: 18) amino acid sequences.
12 . The targeted therapeutic fusion protein of any of claims 7 to 11 , wherein the spacer peptide comprises one or more PAPA (SEQ ID NO: 19) or TPAPA (SEQ ID NO: 20) amino acid sequences.
13 . The targeted therapeutic fusion protein of any of claims 7 to 12 , wherein the spacer peptide comprises one or more AAAKE (SEQ ID NO: 21) amino acid sequences.
14 . The targeted therapeutic fusion protein of any of claims 7 to 8 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of: (GGGGS) n (SEQ ID NOs: 12, 56, 58, 91-94), (GGGGS) n -GGGPS (SEQ ID NOs: 36, 95-100), GAP-(GGGGS) n -GGGPS (SEQ ID NOs: 101-107), GAP-(GGGGS) n -GGGPS-GAP (SEQ ID NOs: 37, 108-113), GAP-(GGGGS) n -GGGPS-(GGGGS) n -GAP (SEQ ID NOs: 114-162), GAP-GGGPS-(GGGGS) n -GAP (SEQ ID NOs: 163-169), GAP-(GGGGS) n -AAAAS-GGGPS-(GGGGS) n -AAAA-GAP (SEQ ID NOs: 170-218), GAP-(GGGGS) n -PAPAP-(Xaa) n -GAP (SEQ ID NOs: 219-267), GAP-(GGGGS) n -PAPAPT-(Xaa) n -GAP (SEQ ID NOs: 268-316), GAP-(GGGGS) n -(Xaa) n -PAPAP-(Xaa) n -(AAAKE) n -(Xaa) n -(GGGGS) n -GAP (SEQ ID NOs: 544-551), (GGGGA) n (SEQ ID NOs: 60, 79, 81, 317-320), (GGGGA) n -GGGPS (SEQ ID NOs: 321-326), GAP-(GGGGA) n -GGGPS (SEQ ID NOs: 327-333), GAP-(GGGGA) n -GGGPS-GAP (SEQ ID NOs: 334-340), GAP-(GGGGA) n -GGGPS-(GGGGA) n -GAP (SEQ ID NOs: 341-389), GAP-GGGPS-(GGGGA) n -GAP (SEQ ID NOs: 390-396), GAP-(GGGGA) n -AAAAS-GGGPS-(GGGGA) n -AAAA-GAP (SEQ ID NOs: 397-445), GAP-(GGGGA) n -PAPAP-(Xaa) n -GAP (SEQ ID NOs: 446-494), GAP-(GGGGA) n -PAPAPT-(Xaa) n -GAP (SEQ ID NOs: 495-543), GAP-(GGGGA) n -(Xaa) n -PAPAP-(Xaa) n -(AAAKE) n -(Xaa) n -(GGGGA) n -GAP (SEQ ID NOs: 552-559); wherein n is 1 to 7.
15 . The targeted therapeutic fusion protein of claim 14 , wherein n is 1 to 4.
16 . The targeted therapeutic fusion protein of claim 7 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of EFGGGGSTR (SEQ ID NO: 22), GAP (SEQ ID NO: 9), GGGGS (SEQ ID NO: 12), GPSGSPG (SEQ ID NO: 23), GPSGSPGT (SEQ ID NO: 24), GPSGSPGH (SEQ ID NO: 25), GGGGSGGGGSGGGGSGGGGSGGGPST (SEQ ID NO: 26), GGGGSGGGGSGGGGSGGGGSGGGPSH (SEQ ID NO: 27), GGGGSGGGGSGGGGSGGGGSGGGPSGGGGSGGGPS (SEQ ID NO: 28), GAPGGGGSGGGGSGGGGSGGGGSGGGPSGGGGSGGGPSGAP (SEQ ID NO: 29), GGGGS GGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGGS GGGGS GGGPS (SEQ ID NO: 30), GAPGGGGS GGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGGS GGGGS GGGPS GAP (SEQ ID NO: 31), GGGGSGGGGSGGGGSGGGPSGGGGSGGGGSGGGPS (SEQ ID NO: 32), GAPGGGGSGGGGSGGGGSGGGPSGGGGSGGGGSGGGPSGAP (SEQ ID NO: 33), GGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS (SEQ ID NO: 34), GAPGGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS GA P (SEQ ID NO: 35), GGGGSGGGGSGGGGSGGGGSGGGPS (SEQ ID NO: 36), GAPGGGGSGGGGSGGGGSGGGGSGGGPSGAP (SEQ ID NO: 37), GGGGSGGGGSAAAASGGGGSGGGPS (SEQ ID NO: 38), GAPGGGGSGGGGSAAAASGGGGSGGGPSGAP (SEQ ID NO: 39), GGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGPS (SEQ ID NO: 40), GAPGGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGPSG AP (SEQ ID NO: 41), GGGGSGGGGSAAAASGGGPSGGGGSAAAASGGGPSGGGGSAAAASGGGPS (SEQ ID NO: 42), GAP GGGGSGGGGSAAAASGGGPSGGGGSAAAASGGGPSGGGGSAAAASGGGPSGA P (SEQ ID NO: 43), GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPS (SEQ ID NO: 44), GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPS GAP (SEQ ID NO: 45), GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPS (SEQ ID NO: 46), GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPSGAP (SEQ ID NO: 47), GGGSPAPTPTPAPTPAPTPAGGGPS (SEQ ID NO: 48), GAPGGGSPAPTPTPAPTPAPTPAGGGPSGAP (SEQ ID NO: 49), GGGSPAPAPTPAPAPTPAPAGGGPS (SEQ ID NO: 50), GAPGGGSPAPAPTPAPAPTPAPAGGGPS GAP (SEQ ID NO: 51), GGGSAEAAAKEAAAKEAAAKAGGPS (SEQ ID NO: 52), GAPGGGSAEAAAKEAAAKEAAAKAGGPSGAP (SEQ ID NO: 53), GGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGG (SEQ ID NO: 54), GAPGGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGGGAP (SEQ ID NO: 55), GGGGSGGGGSGGGGS (SEQ ID NO: 56), GAPGGGGSGGGGSGGGGSGAP (SEQ ID NO: 57), GGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 58), GAPGGGGSGGGGSGGGGSGGGGSGAP (SEQ ID NO: 59), GGGGA (SEQ ID NO: 60), GGGGAGGGGAGGGGAGGGGAGGGPST (SEQ ID NO: 61), GGGGAGGGGAGGGGAGGGGAGGGPSH (SEQ ID NO: 62), GGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGPS (SEQ ID NO: 63), GAPGGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGPSGAP (SEQ ID NO: 64), GGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGGAGGGGAGGGPS (SEQ ID NO: 65), GAP GGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGGAGGGGAGGGPS GAP (SEQ ID NO:66), GGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPS (SEQ ID NO: 67), GAPGGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPSGAP (SEQ ID NO: 68), GGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPSGGGGAGGGGAGGGPS (SEQ ID NO: 69), GAP GGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPSGGGGAGGGGAGGGPS GAP (SEQ ID NO: 70), GGGGAGGGGAGGGGAGGGGAGGGPS (SEQ ID NO: 71), GAPGGGGAGGGGAGGGGAGGGGAGGGPSGAP (SEQ ID NO: 72), GGGGAGGGGAAAAASGGGGAGGGPS (SEQ ID NO: 73), GAPGGGGAGGGGAAAAASGGGGAGGGPSGAP (SEQ ID NO: 74), GGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGPS (SEQ ID NO: 75), GAP GGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGPS GAP (SEQ ID NO: 76), GGGGAGGGGAAAAASGGGPSGGGGAAAAASGGGPSGGGGAAAAASGGGPS (SEQ ID NO: 77), GAPGGGGAGGGGAAAAASGGGPSGGGGAAAAASGGGPSGGGGAAAAASGGGPSG AP (SEQ ID NO: 78), GGGGAGGGGAGGGGA (SEQ ID NO: 79), GAPGGGGAGGGGAGGGGAGAP (SEQ ID NO: 80), GGGGAGGGGAGGGGAGGGGA (SEQ ID NO: 81), GAPGGGGAGGGGAGGGGAGGGGAGAP (SEQ ID NO: 82), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPS [or (GGGGA) 8 GGGPS] (SEQ ID NO: 83), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPSH [or (GGGGA) 8 GGGPSH] (SEQ ID NO: 84), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPS [or (GGGGA) 9 GGGPS] (SEQ ID NO: 85), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPSH [or (GGGGA) 9 GGGPSH] (SEQ ID NO: 86), GGGGPAPGPGPAPGPAPGPAGGGPS (SEQ ID NO: 87), GAPGGGGPAPGPGPAPGPAPGPAGGGPGGAP (SEQ ID NO: 88), GGGGPAPAPGPAPAPGPAPAGGGPS (SEQ ID NO: 89), and GAPGGGGPAPAPGPAPAPGPAPAGGGPGGAP (SEQ ID NO: 90).
17 . The targeted therapeutic fusion protein of claim 16 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of
(SEQ ID NO: 36)
GGGGSGGGGSGGGGSGGGGSGGGPS,
(SEQ ID NO: 44)
GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPS,
(SEQ ID NO: 45)
GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPS
GAP,
(SEQ ID NO: 46)
GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPS,
(SEQ ID NO: 47)
GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTG
PSGAP,
(SEQ ID NO: 48)
GGGSPAPTPTPAPTPAPTPAGGGPS,
(SEQ ID NO: 49)
GAPGGGSPAPTPTPAPTPAPTPAGGGPSGAP,
(SEQ ID NO: 50)
GGGSPAPAPTPAPAPTPAPAGGGPS,
(SEQ ID NO: 51)
GAPGGGSPAPAPTPAPAPTPAPAGGGPSGAP,
(SEQ ID NO: 52)
GGGSAEAAAKEAAAKEAAAKAGGPS,
(SEQ ID NO: 53)
GAPGGGSAEAAAKEAAAKEAAAKAGGPSGAP,
(SEQ ID NO: 54)
GGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGG,
(SEQ ID NO: 55)
GAPGGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGGGAP,
and
(SEQ ID NO: 71)
GGGGAGGGGAGGGGAGGGGAGGGPS.
18 . The targeted therapeutic fusion protein of claim 17 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of
(SEQ ID NO: 36)
GGGGSGGGGSGGGGSGGGGSGGGPS,
(SEQ ID NO: 47)
GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTG
PSGAP,
(SEQ ID NO: 51)
GAPGGGSPAPAPTPAPAPTPAPAGGGPSGAP,
(SEQ ID NO: 55)
GAPGGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGGGAP,
and
(SEQ ID NO: 71)
GGGGAGGGGAGGGGAGGGGAGGGPS.
19 . The targeted therapeutic fusion protein of any one of the preceding claims, wherein the peptide tag is an N-terminal tag or a C-terminal tag.
20 . The targeted therapeutic fusion protein of claim 19 , wherein the peptide tag is a C-terminal tag.
21 . The targeted therapeutic fusion protein of any one of the preceding claims, wherein the spacer comprises a Gly-Ala-Pro (GAP) (SEQ ID NO: 9) or Gly-Pro-Ser (GPS) (SEQ ID NO: 10) amino acid sequence.
22 . The targeted therapeutic fusion protein of any one of the preceding claims, wherein the lysosomal targeting domain comprises amino acids 8-67 of mature human IGF-II.
23 . The targeted therapeutic fusion protein of any one of the preceding claims, wherein the spacer comprises an alpha-helical structure or a rigid structure.
24 . The targeted therapeutic fusion protein of any one of the preceding claims, wherein the IGF-II peptide tag comprises a mutation at residue Arg37.
25 . The targeted therapeutic fusion protein of claim 23 , wherein the mutation is a substitution of alanine for arginine.
26 . The targeted therapeutic fusion protein of any one of the preceding claims, further comprising a pharmaceutically acceptable carrier, diluent or excipient.
27 . A pharmaceutical composition suitable for treating lysosomal storage disease comprising a targeted therapeutic fusion protein of any one of the preceding claims.
28 . A nucleic acid encoding the targeted therapeutic fusion protein of any one of the preceding claims.
29 . A cell containing the nucleic acid of claim 27 .
30 . A method of producing a targeted therapeutic fusion protein comprising a step of: culturing mammalian cells in a cell culture medium, wherein the mammalian cells carry the nucleic acid of claim 28 ; and the culturing is performed under conditions that permit expression of the targeted therapeutic fusion protein.
31 . A method for treating a lysosomal storage disease in a subject comprising administering to the subject a therapeutically effective amount of a pharmaceutical composition comprising a fusion protein comprising a lysosomal enzyme, a peptide tag having an amino acid sequence at least 70% identical to amino acids 8-67 of mature human IGF-II and a spacer peptide located between the lysosomal enzyme amino acid sequence and the IGF-II peptide tag, wherein the spacer peptide comprises one or more GGGPS (SEQ ID NO: 14) or GGGSP (SEQ ID NO: 15) amino acid sequences, and optionally further comprises one or more of (i) GAP (SEQ ID NO: 9), (ii) GGGGS (SEQ ID NO: 12), (iii) GGGS (SEQ ID NO: 16), (iv) AAAAS (SEQ ID NO: 17), (v) AAAS (SEQ ID NO: 18), (vi) PAPA (SEQ ID NO: 19), (vii) TPAPA (SEQ ID NO: 20), (viii) AAAKE (SEQ ID NO: 21) or (ix) GGGGA (SEQ ID NO: 60).
32 . The method of claim 31 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of: (GGGGS) n (SEQ ID NOs: 12, 56, 58, 91-94), (GGGGS) n -GGGPS (SEQ ID NOs: 36, 95-100), GAP-(GGGGS) n -GGGPS (SEQ ID NOs: 101-107), GAP-(GGGGS) n -GGGPS-GAP (SEQ ID NOs: 37, 108-113), GAP-(GGGGS) n -GGGPS-(GGGGS) n -GAP (SEQ ID NOs: 114-162), GAP-GGGPS-(GGGGS) n -GAP (SEQ ID NOs: 163-169), GAP-(GGGGS) n -AAAAS-GGGPS-(GGGGS) n -AAAA-GAP (SEQ ID NOs: 170-218), GAP-(GGGGS) n -PAPAP-(Xaa) n -GAP (SEQ ID NOs: 219-267), GAP-(GGGGS) n -PAPAPT-(Xaa) n -GAP (SEQ ID NOs: 268-316), GAP-(GGGGS) n -(Xaa) n -PAPAP-(Xaa) n -(AAAKE) n -(Xaa) n -(GGGGS) n -GAP (SEQ ID NOs: 544-551), (GGGGA) n (SEQ ID NOs: 60, 79, 81, 317-320), (GGGGA) n -GGGPS (SEQ ID NOs: 321-326), GAP-(GGGGA) n -GGGPS (SEQ ID NOs: 327-333), GAP-(GGGGA) n -GGGPS-GAP (SEQ ID NOs: 334-340), GAP-(GGGGA) n -GGGPS-(GGGGA) n -GAP (SEQ ID NOs: 341-389), GAP-GGGPS-(GGGGA) n -GAP (SEQ ID NOs: 390-396), GAP-(GGGGA) n -AAAAS-GGGPS-(GGGGA) n -AAAA-GAP (SEQ ID NOs: 397-445), GAP-(GGGGA) n -PAPAP-(Xaa) n -GAP (SEQ ID NOs: 446-494), GAP-(GGGGA) n -PAPAPT-(Xaa) n -GAP (SEQ ID NOs: 495-543), GAP-(GGGGA) n -(Xaa) n -PAPAP-(Xaa) n -(AAAKE) n -(Xaa) n -(GGGGA) n -GAP (SEQ ID NOs: 552-559); wherein n is 1 to 7.
33 . The method of claim 32 , wherein n is 1 to 4.
34 . A method for treating Mucopolysaccharidosis Type IIIB (Sanfilippo B Syndrome) in a subject comprising administering to the subject a therapeutically effective amount of a pharmaceutical composition comprising a fusion protein comprising an amino acid sequence at least 85% identical to a human α-N-acetylglucosaminidase (Naglu) protein, a peptide tag having an amino acid sequence at least 70% identical to amino acids 8-67 of mature human IGF-II and a spacer peptide located between the Naglu amino acid sequence and the IGF-II peptide tag, wherein the spacer comprises the amino acid sequence GAP (SEQ ID NO: 9), GPS (SEQ ID NO: 10), or GGS (SEQ ID NO: 11).
35 . The method of claim 34 , wherein the spacer sequence comprises amino acids Gly-Pro-Ser (GPS) (SEQ ID NO: 10) between the amino acids of mature human IGF-II and the amino acids of human Naglu.
36 . The method of any of claims 34 to 35 , wherein the spacer peptide comprises one or more GGGGS (SEQ ID NO: 12), GGGGA (SEQ ID NO: 60) or GGGS (SEQ ID NO: 16) amino acid sequences.
37 . The method of any of claims 34 to 36 , wherein the spacer peptide comprises one or more GGGPS (SEQ ID NO: 14) or GGGSP (SEQ ID NO: 15) amino acid sequences.
38 . The method of any of claims 34 to 37 , wherein the spacer peptide comprises one or more AAAAS (SEQ ID NO: 17) or AAAS (SEQ ID NO: 18) amino acid sequences.
39 . The method of any of claims 34 to 38 , wherein the spacer peptide comprises one or more PAPA (SEQ ID NO: 19) or TPAPA (SEQ ID NO: 20) amino acid sequences.
40 . The method of any of claims 34 to 39 , wherein the spacer peptide comprises one or more AAAKE (SEQ ID NO: 21) amino acid sequences.
41 . The method of any of claim 31 or 34 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of: (GGGGS) n (SEQ ID NOs: 12, 56, 58, 91-94), (GGGGS) n -GGGPS (SEQ ID NOs: 36, 95-100), GAP-(GGGGS) n -GGGPS (SEQ ID NOs: 101-107), GAP-(GGGGS) n -GGGPS-GAP (SEQ ID NOs: 37, 108-113), GAP-(GGGGS) n -GGGPS-(GGGGS) n -GAP (SEQ ID NOs: 114-162), GAP-GGGPS-(GGGGS) n -GAP (SEQ ID NOs: 163-169), GAP-(GGGGS) n -AAAAS-GGGPS-(GGGGS) n -AAAA-GAP (SEQ ID NOs: 170-218), GAP-(GGGGS) n -PAPAP-(Xaa) n -GAP (SEQ ID NOs: 219-267), GAP-(GGGGS) n -PAPAPT-(Xaa) n -GAP (SEQ ID NOs: 268-316), GAP-(GGGGS) n -(Xaa) n -PAPAP-(Xaa) n -(AAAKE) n -(Xaa) n -(GGGGS) n -GAP (SEQ ID NOs: 544-551), (GGGGA) n (SEQ ID NOs: 60, 79, 81, 317-320), (GGGGA) n -GGGPS (SEQ ID NOs: 321-326), GAP-(GGGGA) n -GGGPS (SEQ ID NOs: 327-333), GAP-(GGGGA) n -GGGPS-GAP (SEQ ID NOs: 334-340), GAP-(GGGGA) n -GGGPS-(GGGGA) n -GAP (SEQ ID NOs: 341-389), GAP-GGGPS-(GGGGA) n -GAP (SEQ ID NOs: 390-396), GAP-(GGGGA) n -AAAAS-GGGPS-(GGGGA) n -AAAA-GAP (SEQ ID NOs: 397-445), GAP-(GGGGA) n -PAPAP-(Xaa) n -GAP (SEQ ID NOs: 446-494), GAP-(GGGGA) n -PAPAPT-(Xaa) n -GAP (SEQ ID NOs: 495-543), GAP-(GGGGA) n -(Xaa) n -PAPAP-(Xaa) n -(AAAKE) n -(Xaa) n -(GGGGA) n -GAP (SEQ ID NOs: 552-559); wherein n is 1 to 7.
42 . The method of claim 41 wherein n is 1 to 4.
43 . A method for reducing glycosaminoglycan levels in vivo comprising administering to a subject suffering from Mucopolysaccharidosis Type IIIB (Sanfilippo B Syndrome) an effective amount of a fusion protein comprising i) an amino acid sequence at least 85% identical to a human α-N-acetylglucosaminidase (Naglu) protein, ii) a peptide tag having an amino acid sequence at least 70% identical to amino acids 8-67 of mature human IGF-II, and iii) a spacer peptide located between the Naglu amino acid sequence and the IGF-II peptide tag.
44 . The method of any one of claim 31 , 34 or 43 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of EFGGGGSTR (SEQ ID NO: 22), GAP (SEQ ID NO: 9), GGGGS (SEQ ID NO: 12), GPSGSPG (SEQ ID NO: 23), GPSGSPGT (SEQ ID NO: 24), GPSGSPGH (SEQ ID NO: 25), GGGGSGGGGSGGGGSGGGGSGGGPST (SEQ ID NO: 26), GGGGSGGGGSGGGGSGGGGSGGGPSH (SEQ ID NO: 27), GGGGSGGGGSGGGGSGGGGSGGGPSGGGGSGGGPS (SEQ ID NO: 28), GAPGGGGSGGGGSGGGGSGGGGSGGGPSGGGGSGGGPSGAP (SEQ ID NO: 29), GGGGS GGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGGS GGGGS GGGPS (SEQ ID NO: 30), GAPGGGGS GGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGGS GGGGS GGGPS GAP (SEQ ID NO: 31), GGGGSGGGGSGGGGSGGGPSGGGGSGGGGSGGGPS (SEQ ID NO: 32), GAPGGGGSGGGGSGGGGSGGGPSGGGGSGGGGSGGGPSGAP (SEQ ID NO: 33), GGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS (SEQ ID NO: 34), GAPGGGGS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS GGGGS GGGGS GGGPS GA P (SEQ ID NO: 35), GGGGSGGGGSGGGGSGGGGSGGGPS (SEQ ID NO: 36), GAPGGGGSGGGGSGGGGSGGGGSGGGPSGAP (SEQ ID NO: 37), GGGGSGGGGSAAAASGGGGSGGGPS (SEQ ID NO: 38), GAPGGGGSGGGGSAAAASGGGGSGGGPSGAP (SEQ ID NO: 39), GGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGPS (SEQ ID NO: 40), GAPGGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGGSGGGGSAAAASGGGPSG AP (SEQ ID NO: 41), GGGGSGGGGSAAAASGGGPSGGGGSAAAASGGGPSGGGGSAAAASGGGPS (SEQ ID NO: 42), GAP GGGGSGGGGSAAAASGGGPSGGGGSAAAASGGGPSGGGGSAAAASGGGPSGA P (SEQ ID NO: 43), GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPS (SEQ ID NO: 44), GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPS GAP (SEQ ID NO: 45), GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPS (SEQ ID NO: 46), GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPSGAP (SEQ ID NO: 47), GGGSPAPTPTPAPTPAPTPAGGGPS (SEQ ID NO: 48), GAPGGGSPAPTPTPAPTPAPTPAGGGPSGAP (SEQ ID NO: 49), GGGSPAPAPTPAPAPTPAPAGGGPS (SEQ ID NO: 50), GAPGGGSPAPAPTPAPAPTPAPAGGGPS GAP (SEQ ID NO: 51), GGGSAEAAAKEAAAKEAAAKAGGPS (SEQ ID NO: 52), GAPGGGSAEAAAKEAAAKEAAAKAGGPSGAP (SEQ ID NO: 53), GGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGG (SEQ ID NO: 54), GAPGGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGGGAP (SEQ ID NO: 55), GGGGSGGGGSGGGGS (SEQ ID NO: 56), GAPGGGGSGGGGSGGGGS GAP (SEQ ID NO: 57), GGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 58), GAPGGGGSGGGGSGGGGSGGGGSGAP (SEQ ID NO: 59), GGGGA (SEQ ID NO: 60), GGGGAGGGGAGGGGAGGGGAGGGPST (SEQ ID NO: 61), GGGGAGGGGAGGGGAGGGGAGGGPSH (SEQ ID NO: 62), GGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGPS (SEQ ID NO: 63), GAPGGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGPSGAP (SEQ ID NO: 64), GGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGGAGGGGAGGGPS (SEQ ID NO: 65), GAP GGGGAGGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGGAGGGGAGGGPS GAP (SEQ ID NO:66), GGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPS (SEQ ID NO: 67), GAPGGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPSGAP (SEQ ID NO: 68), GGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPSGGGGAGGGGAGGGPS (SEQ ID NO: 69), GAP GGGGAGGGGAGGGGAGGGPSGGGGAGGGGAGGGPSGGGGAGGGGAGGGPS GAP (SEQ ID NO: 70), GGGGAGGGGAGGGGAGGGGAGGGPS (SEQ ID NO: 71), GAPGGGGAGGGGAGGGGAGGGGAGGGPSGAP (SEQ ID NO: 72), GGGGAGGGGAAAAASGGGGAGGGPS (SEQ ID NO: 73), GAPGGGGAGGGGAAAAASGGGGAGGGPSGAP (SEQ ID NO: 74), GGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGPS (SEQ ID NO: 75), GAP GGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGGAGGGGAAAAASGGGPS GAP (SEQ ID NO: 76), GGGGAGGGGAAAAASGGGPSGGGGAAAAASGGGPSGGGGAAAAASGGGPS (SEQ ID NO: 77), GAP GGGGAGGGGAAAAASGGGPSGGGGAAAAASGGGPSGGGGAAAAASGGGPSG AP (SEQ ID NO: 78), GGGGAGGGGAGGGGA (SEQ ID NO: 79), GAPGGGGAGGGGAGGGGAGAP (SEQ ID NO: 80), GGGGAGGGGAGGGGAGGGGA (SEQ ID NO: 81), GAPGGGGAGGGGAGGGGAGGGGAGAP (SEQ ID NO: 82), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPS [or (GGGGA) 8 GGGPS] (SEQ ID NO: 83), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPSH [or (GGGGA) 8 GGGPSH] (SEQ ID NO: 84), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPS [or (GGGGA) 9 GGGPS] (SEQ ID NO: 85), GGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGGAGGGPSH [or (GGGGA) 9 GGGPSH] (SEQ ID NO: 86), GGGGPAPGPGPAPGPAPGPAGGGPS (SEQ ID NO: 87), GAPGGGGPAPGPGPAPGPAPGPAGGGPGGAP (SEQ ID NO: 88), GGGGPAPAPGPAPAPGPAPAGGGPS (SEQ ID NO: 89), and GAPGGGGPAPAPGPAPAPGPAPAGGGPGGAP (SEQ ID NO: 90).
45 . The method of claim 44 , wherein the spacer comprises an amino acid sequence selected from the group consisting of GGGGSGGGGSGGGGSGGGGSGGGPS (SEQ ID NO: 36), GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPS (SEQ ID NO: 44), GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPGPSGAP (SEQ ID NO: 45), GGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPS (SEQ ID NO: 46), GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPSGAP (SEQ ID NO: 47), GGGSPAPTPTPAPTPAPTPAGGGPS (SEQ ID NO: 48), GAPGGGSPAPTPTPAPTPAPTPAGGGPSGAP (SEQ ID NO: 49), GGGSPAPAPTPAPAPTPAPAGGGPS (SEQ ID NO: 50), GAPGGGSPAPAPTPAPAPTPAPAGGGPSGAP (SEQ ID NO: 51), GGGSAEAAAKEAAAKEAAAKAGGPS (SEQ ID NO: 52), GAPGGGSAEAAAKEAAAKEAAAKAGGPSGAP (SEQ ID NO: 53), GGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGG (SEQ ID NO: 54), GAPGGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGGGAP (SEQ ID NO: 55), and GGGGAGGGGAGGGGAGGGGAGGGPS (SEQ ID NO: 71).
46 . The method of claim 45 , wherein the spacer peptide comprises an amino acid sequence selected from the group consisting of GGGGSGGGGSGGGGSGGGGSGGGPS (SEQ ID NO: 36), GAPGGSPAGSPTSTEEGTSESATPESGPGTSTEPSEGSAPGSPAGSPTSTGPSGAP (SEQ ID NO: 47), GAPGGGSPAPAPTPAPAPTPAPAGGGPSGAP (SEQ ID NO: 51), GAPGGGSPAEAAAKEAAAKEAAAKEAAAKEAAAKAPSGGGGAP (SEQ ID NO: 55), and GGGGAGGGGAGGGGAGGGGAGGGPS (SEQ ID NO: 71).
47 . The method of any one of claims 31 to 46 , wherein the spacer sequence comprises amino acids Gly-Ala-Pro (GAP) (SEQ ID NO: 9) sequence.
48 . The method of any one of claims 31 to 47 , wherein the peptide tag is an N-terminal tag or a C-terminal tag.
49 . The method of claim 48 , wherein the peptide tag is a C-terminal tag.
50 . The method of any one of claims 31 to 49 , wherein the spacer comprises an alpha-helical structure or a rigid structure.
51 . The method of any one of claims 31 to 50 , wherein the lysosomal targeting domain comprises amino acids 8-67 of mature human IGF-II.
52 . The method of any one of claims 31 to 51 , wherein the IGF-II peptide tag comprises a mutation at residue Arg37.
53 . The method of claim 52 , wherein the mutation is a substitution of alanine for arginine.
54 . The method of any one of claims 31 to 53 , wherein the fusion protein comprises amino acids 1-743 or amino acids 24-743 of human Naglu ( FIG. 1 ).
55 . The method of any one of claims 31 to 54 , wherein the effective amount is in the range of 2.5-20 mg per kilogram of body weight of the subject.
56 . The method of any one of claims 31 to 55 , wherein the fusion protein is administered intrathecally, optionally further comprising administering the fusion protein intravenously.
57 . The method of claim 56 , wherein the administering comprises introducing the fusion protein into a cerebral ventricle, lumbar area, or cisterna magna.
58 . The method of any one of claims 31 to 55 , wherein the fusion protein is administered intravenously.
59 . The method of any one of claims 31 to 58 , wherein the fusion protein is administered bimonthly, monthly, triweekly, biweekly, weekly, daily, or at variable intervals.
60 . The method of any one of claims 31 to 59 , wherein the treatment results in reducing glycosaminoglycan (GAG) levels in a brain tissue.
61 . The method of any one of claims 31 to 60 , wherein the treatment results in reducing lysosomal storage granules in a brain tissue.Join the waitlist — get patent alerts
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