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CN-115820740-B - Recombinant adeno-associated viral vectors for the treatment of mucopolysaccharidosis type II and uses thereof

CN115820740BCN 115820740 BCN115820740 BCN 115820740BCN-115820740-B

Abstract

The invention discloses a recombinant adeno-associated virus vector carrying an IDS gene expression frame and application thereof in treating type II mucopolysaccharide storage disease.

Inventors

  • WU XIAOBING
  • MA WENHAO
  • WU ZHIJIE

Assignees

  • 北京锦篮基因科技有限公司

Dates

Publication Date
20260512
Application Date
20210916

Claims (20)

  1. 1. A recombinant AAV9 viral vector, comprising an expression construct of iduronate-2-sulfatase IDS gene comprising the following elements functionally linked to each other in the direction of transcription: (1) A promoter sequence shown in SEQ ID NO. 1; (2) Nucleic acid encoding human iduronate-2-sulfatase, (3) A human miR-142-3p target sequence, which consists of 2 nucleotide sequences shown in SEQ ID NO. 7 in tandem.
  2. 2. The recombinant AAV9 viral vector of claim 1, comprising in the genome: a.5 'and 3' AAV inverted terminal repeat ITR sequences, and B. the expression construct of claim 1 located between the 5 'and 3' itrs.
  3. 3. The recombinant AAV9 viral vector of claim 2, wherein the expression construct comprises the following elements functionally linked to each other in the transcriptional direction: a promoter sequence as shown in SEQ ID No.1, The sequence of-Kozak, A polynucleotide encoding human iduronate-2-sulfatase hIDS, A human miR-142-3p target sequence shown in SEQ ID NO. 3, -A transcription terminator.
  4. 4. The recombinant AAV9 viral vector of claim 3, wherein the transcription terminator is a bovine growth hormone polyA sequence.
  5. 5. The recombinant AAV9 viral vector of claim 2, wherein the 5 'and 3' ITRs are wild-type AAV2 ITR sequences.
  6. 6. The recombinant AAV9 viral vector of claim 1, wherein the nucleic acid encoding human iduronate-2-sulfatase encodes a human iduronate-2-sulfatase selected from the amino acid sequence of: (a) Amino acid 1 to amino acid 550 of SEQ ID NO. 10; (b) Amino acid 26 to amino acid 550 of SEQ ID NO 10, or (C) Amino acid 34 to amino acid 550 of SEQ ID NO. 10.
  7. 7. The recombinant AAV9 viral vector of claim 1, wherein the nucleic acid encoding human iduronate-2-sulfatase is a nucleotide sequence selected from the group consisting of: (i) Nucleotide sequence shown as SEQ ID NO. 2, or (Ii) A nucleotide sequence which encodes the same iduronate-2-sulfatase as the nucleotide sequence of (i) but which differs from the nucleotide sequence of (i) by the degeneracy of the genetic code.
  8. 8. A pharmaceutical composition comprising the recombinant AAV9 viral vector of any one of claims 1-7 and a pharmaceutically acceptable carrier.
  9. 9. The pharmaceutical composition of claim 8, wherein the pharmaceutical composition is for intravenous administration or lateral ventricle injection administration.
  10. 10. The pharmaceutical composition of claim 9, wherein the pharmaceutical composition is for lateral ventricle injection administration.
  11. 11. The pharmaceutical composition of any one of claims 8-10, wherein the recombinant AAV9 viral vector has a titer of at least 1.0e13 GC/ml.
  12. 12. Use of a recombinant AAV9 viral vector according to any one of claims 1-7 or a pharmaceutical composition according to any one of claims 8-11 in the manufacture of a medicament for treating mucopolysaccharidosis type II (MPSII) in a subject.
  13. 13. The use of claim 12, wherein the subject is a neonatal MPSII patient, or a MPSII patient less than 5 years old or 12 years old, or a MPSII patient less than 16 years old, or an adult MPSII patient.
  14. 14. The use of claim 13, wherein the patient is MPSII patient with CNS involvement.
  15. 15. The use of any one of claims 12-14, wherein the pharmaceutical composition is for increasing the level of functionality hIDS in a cell or one or more tissues or organs in the body of a subject.
  16. 16. The use of any one of claims 12-14, wherein the pharmaceutical composition is for expressing hIDS in the heart, liver, spleen, lung, kidney, muscle, gut, and brain of a subject.
  17. 17. The use according to any one of claims 12-14, wherein the pharmaceutical composition is formulated for systemic or topical administration.
  18. 18. The use of claim 17, wherein the pharmaceutical composition is formulated for intravenous administration or lateral ventricular injection administration.
  19. 19. The use of claim 17, wherein the pharmaceutical composition is formulated for lateral ventricle injection administration.
  20. 20. The use according to any one of claims 12-14, wherein the pharmaceutical composition is for increasing the level of functionality hIDS in peripheral tissues and/or in the brain of a subject.

Description

Recombinant adeno-associated viral vectors for the treatment of mucopolysaccharidosis type II and uses thereof Technical Field The invention relates to the field of biotechnology, in particular to a recombinant adeno-associated virus vector carrying an IDS gene expression frame and application thereof in treating type II mucopolysaccharidosis. Background Mucopolysaccharidoses (mucopolysaccharidosis, MPS) are a single genetic metabolic disease of severe disability and mortality caused by the accumulation of GAGs and their intermediate metabolites in the body due to the inability or incomplete degradation of acidic mucopolysaccharides (also known as glycosaminoglycans, GAGs) by the lack or reduced activity of the lysosomal associated acid hydrolases. MPS IS currently classified into 7 large forms, including MPSI type (containing IH, IS, IH/S subtypes), MPSII type (containing IIA, IIB subtypes), MPSI type (containing IIIA, IIIB, IIIC, IIID subtypes), MPSIV type (containing IVA, IVB subtypes), MPSVI type (containing VIA, VIB subtypes), MPSVII type, MPSIX type, depending on the defects of lysosomal degrading enzyme and the kind of storage mucopolysaccharide. China is more common in MPSI, II, IV, VI types. Type II mucopolysaccharidoses (Mucopolysaccharidosis type II, MPSII), also known as Hunter syndrome, are the only X chromosome-linked recessive genetic disease in mucopolysaccharidoses, a rare genetic disease with a morbidity of about 0.38/10 ten thousand to 1.09/10 ten thousand, and European countries typically have a lower morbidity than eastern countries, where MPSII accounts for about 50% of all Mucopolysaccharidoses (MPS). Patient lysosomes lack iduronate 2-sulfatase (iduronate 2-sulphatase, I2S) resulting in pathological accumulation of both glycosaminoglycans (GAGs) of Dermatan Sulfate (DS) and Heparan Sulfate (HS) and dysfunction of most organ systems, including the brain. MPSII patients will be affected to varying degrees in most organ systems and the disease will exhibit significant heterogeneity. Clinical manifestations according to MPSII are classified into light and heavy, with heavy accounting for about 2/3. Common clinical signs and symptoms include altered levels of urinary GAGs (uGAGs), facial roughness, skeletal deformity and joint stiffness, short stature caused by growth retardation, respiratory and cardiac injuries including diffuse valvular disease, inguinal and umbilical hernias, hepatosplenomegaly, and the like. The patient also has ear-nose-throat manifestations including hearing loss, adenotonsil hypertrophy, frequent ear and upper respiratory tract infections, sleep disorders, obstructive apneas, and retinal degeneration. Heavy patients have neurological diseases, mainly manifested by cognitive impairment and serious behavioral problems. Patients typically appear normal at birth, signs begin to appear usually at the age of 2 to 4 years, signs in heavy patients usually appear earlier, light patients develop slowly, there are no or fewer cognitive problems, and there are no behavioral disorders. In major clinical changes, cardiopulmonary failure is often the cause of death, with severe forms occurring before adulthood, and mild forms surviving to late adulthood. Etiology MPSII is due to mutations in the IDS gene that result in a decrease or disappearance of I2S enzyme activity in the lysosome, which in turn results in accumulation of HS, DS in the lysosome, resulting in cell and organ damage that can lead to progressive cell and multi-organ dysfunction in patients. IDS gene, located at Xq28, CDS full length 1653bp, encodes a 550 amino acid polypeptide that is processed to form iduronate 2-sulfatase (iduronate-sulphatase, I2S) protein. The mature form is monomeric, 76kDa in size, and consists of two subdomains (a heavy chain of 42kDa and a light chain of 14 kDa). Hydrolysis of the C2-sulfate linkage of the 2-O-sulfo-alpha-L-iduronic acid residue in Dermatan Sulfate (DS) and Heparan Sulfate (HS) is catalyzed in vivo. IDS is a housekeeping gene and is widely expressed in various organs throughout the body, with a high overall expression level in the central nervous system. MPSII treatments have been palliative, focusing on the treatment of signs and symptoms. Since the discovery of the biochemical and genetic basis of the disease in the 1970 s and 1990 s, respectively, many studies have been conducted, with different strategies being employed with the aim of developing specific therapies for the disease. These efforts have led to Hematopoietic Stem Cell Transplantation (HSCT) in the 1980 s and Enzyme Replacement Therapy (ERT) in 2006 coming into clinical practice. Although these therapeutic strategies (mainly ERT) have been used today as therapeutic alternatives for MPS, there are a number of problems yet to be solved regarding the effectiveness and safety of their application. MPSII ERT treatment currently uses two different recombinases Elaprase, approved by the United states food and dr