Targeted therapeutic proteins

Patent metadata

Field	Value
Publication number	US-9814762-B2
Application number	US-201615016930-A
Country	US
Kind code	B2
Filing date	Feb 5, 2016
Priority date	Apr 30, 2001
Publication date	Nov 14, 2017
Grant date	Nov 14, 2017

Abstract

Targeted therapeutics that localize to a specific subcellular compartment such as the lysosome are provided. The targeted therapeutics include a therapeutic agent and a targeting moiety that binds a receptor on an exterior surface of the cell, permitting proper subcellular localization of the targeted therapeutic upon internalization of the receptor. Nucleic acids, cells, and methods relating to the practice of the invention are also provided.

First claim

We claim:

1. A nucleic acid encoding a targeted therapeutic fusion protein comprising: a lysosomal enzyme; and a lysosomal targeting domain that binds human cation-independent mannose-6-phosphate receptor in a mannose-6-phosphate-independent manner; wherein the lysosomal targeting domain comprises a mutein of mature human IGF-II having an amino acid sequence at least 70% identical to mature human IGF-II (SEQ ID NO:8).

2. The nucleic acid of claim 1 , wherein the IGF-II moiety comprises a deletion or replacement of amino acids 1-7 of mature human IGF-II.

3. The nucleic acid of claim 1 , wherein the IGF-II moiety comprises:
(a) amino acids 8-67 of mature human IGF-II;
(b) amino acids 1-61 of mature human IGF-II; or
(c) amino acids 8-61 of mature human IGF-II.

4. The nucleic acid of claim 1 , wherein the IGF-II moiety comprises amino acids 8-67 of mature human IGF-II.

5. The nucleic acid of claim 1 , wherein the variant of human IGF-II comprises a deletion or replacement of amino acids 62-67 of mature human IGF-II.

6. The nucleic acid of claim 1 , wherein the variant of human IGF-II comprises a deletion or a replacement of amino acids 29-40 of mature human IGF-II.

7. The nucleic acid of claim 1 , wherein the variant of human IGF-II comprises a deletion or a replacement of amino acids 1-7 and a substitution of Tyr27Leu of mature human IGF-II.

8. The nucleic acid of claim 1 , wherein the variant of human IGF-II comprises one or more substitutions of amino acid residues Tyr27 with Leu, Leu43 with Val, and/or Ser36 with Phe of mature human IGF-II.

9. The nucleic acid of claim 1 , wherein the variant of human IGF-II comprises amino acids 41-61 of human IGF-II and a mutein of amino acids 8-28 of human IGF-II, the mutein differing from human IGF-II at a position selected from the group consisting of amino acid 9, amino acid 19, amino acid 26, and amino acid 27 of mature human IGF-II.

10. The nucleic acid of claim 1 , wherein the mutein of mature human IGF-II comprises amino acids 48-55 of mature human IGF-II.

11. The nucleic acid of claim 1 , wherein the mutein of mature human IGF-II comprises, at corresponding positions, at least three amino acids selected from the group consisting of amino acids 8, 48, 49, 50, 54, and 55 of mature human IGF-II.

12. The nucleic acid of claim 1 , wherein the amino acid sequence comprises, at positions corresponding to positions 54 and 55 of mature human IGF-II, amino acids each of which are uncharged or negatively charged at pH 7.4.

13. The nucleic acid of claim 1 , wherein the mutein of mature human IGF-II comprises the amino acid sequence phenylalanine-arginine-serine.

14. The nucleic acid of claim 1 , wherein the mutein of mature human IGF-II comprises amino acids 8-28 and 41-61 of mature human IGF-II.

15. The nucleic acid of claim 1 , wherein the fusion protein further comprises a bridge between the lysosomal enzyme and the lysosomal targeting moiety.

16. The nucleic acid of claim 15 , wherein the bridge comprises the amino acid sequence of GAP.

17. The nucleic acid of claim 1 , wherein the lysosomal enzyme is selected from the group consisting of acid-α1,4-glucosidase; β-galactosidase; β-hexosaminidase A; β-hexosaminidase B; α-galactosidase A; glucocerebrosidase; arylsulfatase B; galactosylceramidase; acid sphingomyelinase; acid ceramidase; acid lipase; α-L-iduronidase; iduronate sulfatase; heparan N-sulfatase; α-N-acetylglucosaminidase; acetyl-CoA-glucosaminide acetyltransferase; N-acetylglucosamine-6-sulfatase; galactosamine-6-sulfatase; arylsulfatase B; β-glucuronidase; α-mannosidase; β-mannosidase; α-L-fucosidase; N-aspartyl-β-glucosaminidase; α-neuraminidase; lysosomal protective protein; α-N-acetyl- galactosaminidase; N-acetylglucosamine-1-phosphotransferase; and palmitoyl-protein thioesterase.

18. A gene therapy vector comprising a nucleic acid encoding a targeted therapeutic protein of claim 1 .

19. The gene therapy vector of claim 18 that is a viral vector.

20. A cell comprising the nucleic acid of claim 1 .

21. A method of treating a patient with a lysosomal storage disease, the method comprising administering to the patient a nucleic acid encoding a therapeutic fusion protein as set out in claim 1 .

22. The method of claim 21 , wherein the lysosomal storage disease is selected from the group consisting of Pompe Disease; Tay Sachs disease; Sandhoff disease; Fabry disease; Gaucher disease; Mucopolysaccharidosis (MPS) I; MPS II, MPS IVA, MPSIVB, MPS VII; α-Mannosidosis; Wolman disease and Sanfilippo B.

23. A method of treating a patient with a lysosomal storage disease, comprising administering to the patient a gene therapy vector as set out in claim 18 .

24. The method of claim 23 , wherein the lysosomal storage disease is selected from the group consisting of Pompe Disease; Tay Sachs disease; Sandhoff disease; Fabry disease; Gaucher disease; Mucopolysaccharidosis (MPS) I; MPS II, MPS IVA, MPSIVB, MPS VII; α-Mannosidosis; Wolman disease and Sanfilippo B.

Assignees

• Biomarin Pharm Inc

Inventors

• Lebowitz Jonathan
• Beverley Stephen M

Classifications

• C07K14/65

  Insulin-like growth factors, i.e. somatomedins, e.g. IGF-1, IGF-2 · CPC title

• C07K2319/21

  containing a His-tag · CPC title

• C07K2319/02

  containing a signal sequence · CPC title

• A61K47/642

  the peptide or protein in the drug conjugate being a cytokine, e.g. IL2, chemokine, growth factors or interferons being the inactive part of the conjugate · CPC title

• A61K38/30 Primary

  Insulin-like growth factors, i.e. somatomedins, e.g. IGF-1, IGF-2 {(insulin-like growth factor binding protein A61K38/1754)} · CPC title