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KR-20260068081-A - Gene therapy vectors for the treatment of Parkinson's disease and their uses

KR20260068081AKR 20260068081 AKR20260068081 AKR 20260068081AKR-20260068081-A

Abstract

Gene therapy vectors for the treatment of Parkinson's disease and their uses are provided. Specifically, an adeno-associated virus (AAV) vector for the treatment of Parkinson's disease is provided, which can promote dopamine synthesis by simultaneously expressing functional tyrosine hydroxylase (TH), GTP-cyclohydrolase 1 (GCH1), and aromatic amino acid decarboxylase (AADC). Also provided are AAV virus particles containing an AAV vector, compositions containing an AAV vector or AAV virus particles, and uses of the AAV vector, AAV virus particles, and compositions in the manufacture of drugs for the prevention or treatment of Parkinson's disease.

Inventors

  • 우, 하오콴
  • 딩, 얀푸
  • 수, 링링

Assignees

  • 강린 바이오테크 (항저우) 씨오., 엘티디.

Dates

Publication Date
20260513
Application Date
20240827
Priority Date
20230828

Claims (20)

  1. An adeno-associated virus (AAV) vector comprising a nucleic acid sequence encoding tyrosine hydroxylase (TH), GTP-cyclohydrolase 1 (GCH1), and aromatic amino acid decarboxylase (AADC), wherein the nucleic acid sequence encoding TH, GCH1, and/or AADC is in-frame linked by a nucleic acid sequence encoding a linker, wherein at least one of TH, GCH1, and AADC is a truncated form of the wild type or a functional variant thereof and has catalytic activity required for the dopamine synthesis pathway.
  2. An AAV vector according to claim 1, wherein TH is a truncated form of wild-type TH or a functional variant thereof and has activity for catalyzing the synthesis of levodopa from tyrosine; preferably, TH comprises 330 to 390 amino acid residues.
  3. An AAV vector according to claim 1 or 2, wherein TH comprises an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity with respect to sequence identification number: 11.
  4. An AAV vector according to any one of claims 1 to 3, wherein GCH1 comprises an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity with respect to sequence identification number: 9.
  5. An AAV vector according to any one of claims 1 to 4, wherein the AADC comprises an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity with respect to sequence identification number: 7.
  6. An AAV vector according to any one of claims 1 to 5, wherein the linker comprises a fusion linker, a 2A peptide linker, and/or an internal ribosome entry site (IRES).
  7. In claim 6, the fusion linker is a peptide linker; preferably, the peptide linker is selected from (GGS)n, (GGGS)n or (GGGGS)n, wherein n is an integer from 1 to 5, an AAV vector.
  8. In paragraph 7, an AAV vector in which the fusion linker is (G4S)3.
  9. An AAV vector according to claim 6, wherein the 2A peptide linker comprises a 2A peptide selected from foot-and-mouth disease virus 2A peptide (F2A), porcine Tescovirus 2A peptide (P2A), Tosea atignavirus 2A peptide (T2A), and/or equine rhinitis virus 2A peptide (E2A).
  10. In claim 9, an AAV vector wherein the N-terminus and/or C-terminus of the 2A peptide linker, preferably the N-terminus, further comprises a GSG amino acid sequence.
  11. An AAV vector according to claim 10, wherein the 2A peptide linker comprises an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity with respect to sequence identification number: 13 or sequence identification number: 15.
  12. An AAV vector according to any one of claims 1 to 11, wherein the nucleic acid sequences encoding TH, GCH1, and AADC are arranged from upstream to downstream in an order selected from any one of the following: (1) TH, GCH1, AADC; (2) AADC, GCH1, TH; (3) AADC, TH, GCH1; (4) GCH1, TH, AADC; (5) GCH1, AADC, TH; or (6) TH, AADC, GCH1.
  13. In paragraph 12, an AAV vector in which the nucleic acid sequences encoding TH, GCH1, and AADC are linked from upstream to downstream in a manner selected from any one of the following: 1) TH-F2A-GCH1-P2A-AADC; 2) GCH1-P2A-TH-P2A-AADC; 3) GCH1-P2A-AADC-P2A-TH; 4) AADC-E2A-GCH1-(G4S)3-TH; 5) GCH1-F2A-TH-F2A-AADC; 6) AADC-(G4S)3-TH-T2A-GCH1; 7) AADC-(G4S)4-TH-T2A-GCH1; 8) AADC-P2A-GCH1-P2A-TH; 9) AADC-T2A-GCH1-(G4S)3-TH; 10) GCH1-F2A-TH-P2A-AADC; 11) GCH1-F2A-AADC-F2A-TH; 12) AADC-(G4S)3-GCH1-F2A-TH; 13) GCH1-E2A-TH-T2A-AADC; 14) GCH1-F2A-AADC-P2A-TH; 15) AADC-(G4S)3-GCH1-P2A-TH; 16) AADC-(G4S)5-GCH1-P2A-TH; 17) TH-(G4S)3-GCH1-P2A-AADC; 18) GCH1-T2A-AADC-E2A-TH; 19) AADC-(G4S)3-GCH1-T2A-TH; 20) GCH1-E2A-TH-P2A-AADC; 21) AADC-(G4S)3-GCH1-E2A-TH; 22) GCH1-E2A-AADC-P2A-TH; 23) AADC-(G4S)3-GCH1-(G4S)3-TH; 24) GCH1-T2A-TH-E2A-AADC; 25) GCH1-(G4S)3-TH-T2A-AADC; 26) GCH1-(G4S)2-TH-T2A-AADC; 27) GCH1-P2A-AADC-T2A-TH; 28) TH-P2A-GCH1-P2A-AADC; 29) TH-(G4S)3-GCH1-T2A-AADC; 30) GCH1-(G4S)3-TH-E2A-AADC; 31) GCH1-T2A-TH-(G4S)3-AADC; 32) TH-P2A-GCH1-F2A-AADC; 33) GCH1-F2A-TH-(G4S)3-AADC; 34) TH-F2A-GCH1-F2A-AADC; 35) TH-(G4S)3-GCH1-E2A-AADC; 36) TH-G4S-GCH1-E2A-AADC; 37) GCH1-P2A-TH-(G4S)3-AADC; 38) AADC-F2A-GCH1-P2A-TH; 39) TH-T2A-GCH1-(G4S)3-AADC; 40) GCH1-E2A-TH-(G4S)3-AADC; 41) THE-E2A-GCH1-(G4S)3-AADC; 42) TH-T2A-GCH1-E2A-AADC; 43) AADC-F2A-GCH1-F2A-TH; 44) TH-F2A-GCH1-(G4S)3-AADC; 45) GCH1-(G4S)3-TH-(G4S)3-AADC; 46) TH-(G4S)3-GCH1-F2A-AADC; 47) THE-E2A-GCH1-T2A-AADC; 48) AADC-P2A-TH-(G4S)3-GCH1; 49) TH-(G4S)3-GCH1-(G4S)3-AADC; 50) AADC-F2A-TH-(G4S)3-GCH1; 51) TH-F2A-AADC-F2A-GCH1; 52) TH-(G4S)3-AADC-T2A-GCH1; 53) TH-(G4S)3-AADC-E2A-GCH1; 54) AADC-P2A-GCH1-(G4S)3-TH; 55) TH-F2A-AADC-(G4S)3-GCH1; 56) AADC-F2A-GCH1-(G4S)3-TH; 57) TH-P2A-AADC-F2A-GCH1; 58) TH-P2A-AADC-(G4S)3-GCH1; 59) GCH1-(G4S)3-TH-P2A-AADC; 60) TH-T2A-AADC-(G4S)3-GCH1; 61) TH-T2A-AADC-E2A-GCH1; 62) GCH1-(G4S)3-TH-F2A-AADC; 63) THE-E2A-AADC-(G4S)3-GCH1; 64) AADC-E2A-GCH1-T2A-TH; 65) GCH1-P2A-TH-F2A-AADC; 66) TH-(G4S)3-AADC-(G4S)3-GCH1; 67) THE-E2A-AADC-T2A-GCH1; 68) GCH1-P2A-AADC-F2A-TH; 69) AADC-T2A-TH-(G4S)3-GCH1; 70) AADC-T2A-GCH1-E2A-TH; 71) TH-F2A-AADC-P2A-GCH1; 72) AADC-E2A-TH-(G4S)3-GCH1; 73) AADC-P2A-GCH1-F2A-TH; 74) TH-(G4S)3-AADC-F2A-GCH1; 75) AADC-P2A-TH-F2A-GCH1; 76) AADC-(G4S)3-TH-P2A-GCH1; 77) AADC-T2A-TH-E2A-GCH1; 78) AADC-(G4S)3-TH-E2A-GCH1; 79) AADC-E2A-TH-T2A-GCH1; or 80) AADC-(G4S)3-TH-F2A-GCH1.
  14. In paragraph 13, the AAV vector comprises a nucleic acid sequence described in any one of sequence identification numbers: 19-21.
  15. An AAV vector according to any one of claims 1 to 14, further comprising a promoter operably linked to a nucleic acid sequence encoding TH, GCH1, and AADC; preferably, wherein the promoter is selected from a CBH promoter, a chimeric synapsin I promoter, a CMV promoter, a CAG promoter, a CAGG promoter, or a CASI promoter.
  16. An AAV vector according to claim 15, wherein the promoter comprises a nucleic acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity with respect to sequence identification numbers: 1-6; preferably, the promoter comprises a nucleic acid sequence described in any one of sequence identification numbers: 1-3; and more preferably, the promoter comprises a nucleic acid sequence described in sequence identification number: 1.
  17. An AAV vector according to any one of claims 1 to 16, wherein the AAV vector further comprises one or more elements selected from a KOZAK sequence, a polyadenylation signal of the SV40 virus (SV40 poly A), and an inversion terminal repeat (ITR).
  18. An AAV virus particle comprising an AAV vector and a capsid protein according to any one of claims 1 to 17.
  19. In paragraph 18, an AAV virus particle in which the capsid protein is derived from an AAV selected from the following serotypes: AAV5, AAV9, AAVPHP.eB, AAVPHP.S, or AAVPHP.B.
  20. An AAV virus particle according to claim 19, wherein the capsid protein comprises an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity with respect to any one of sequence identification numbers: 33-37; preferably, the capsid protein comprises an amino acid sequence selected from sequence identification number: 34 or 37; and more preferably, the capsid protein comprises an amino acid sequence described in sequence identification number: 34.

Description

Gene therapy vectors for the treatment of Parkinson's disease and their uses The present disclosure relates to the field of gene therapy, particularly to gene therapy vectors and compositions for the treatment of Parkinson's disease. Parkinson's disease (PD) is a common neurodegenerative disorder. Patients with PD are characterized by the loss of dopaminergic neurons within the substantia nigra region. Dopaminergic neurons are a type of neuron primarily capable of producing dopamine, which can transmit neural impulse signals to the striatum, the region that primarily controls skeletal muscle movement. Consequently, reduced dopamine synthesis leads to motor impairments. Key clinical findings in patients include bradykinesia, dystonia, resting tremor, and abnormal posture and gait. PD affects approximately 1% of the global population aged 55 and older. With the aging of society, the patient population is expected to continue expanding. The conventional treatment for Parkinson's disease is the oral administration of levodopa, a dopamine precursor. While this drug provides good relief during the initial stages of treatment, once the "honeymoon period" of treatment ends, levodopa can cause various motor complications. In contrast, dopamine agonist drugs can alleviate symptoms to some extent, but long-term use can lead to adverse reactions, such as gastrointestinal reactions and psychiatric symptoms. In the case of surgical treatment, it shows improvement in symptoms but may not provide a cure. Post-operative medication is still necessary, and while it may lead to associated complications such as paralysis of the tongue and hands, impaired distance perception, and unconsciousness, it also has certain limitations. Gene therapy offers unique advantages for the treatment of Parkinson's disease. On the one hand, relevant genes can be introduced into the lesion site via a vector to repair, supplement, and modify cells within the target region, thereby restoring normal physiological function in the affected area. This is primarily achieved by introducing neurotrophic factors (including glial cell-derived neurotrophic factor (GDNF), neuturin, and brain-derived neurotrophic factor (BDNF)) to promote the survival and protect the affected neurons. However, a large number of dopaminergic neurons are lost at the onset of PD in most patients. Therefore, treating this disease by increasing dopamine synthesis in the affected nuclei may be key. Dopamine synthesis primarily involves the following enzymes: tyrosine hydroxylase (TH), aromatic amino acid decarboxylase (AADC), and GTP-cyclohydrolase 1 (GCH1). Tyrosine is catalyzed to levodopa by tyrosine hydroxylase (TH). Subsequently, levodopa is converted to dopamine by aromatic amino acid decarboxylase (AADC). TH requires tetrahydrobiopterin as a coenzyme, which is synthesized by GTP-cyclohydrolase 1 (GCH1). Therefore, the expression of TH, AADC, and GCH1 can enable efficient dopamine synthesis. Currently, gene therapy strategies under development for Parkinson's disease targeting dopamine synthetase involve using lentiviral vectors to deliver expression vectors containing TH, GCH1, and AADC into the lesion site. By achieving high expression of TH, GCH1, and AADC, this strategy increases the effective concentration of local dopamine in the midbrain striatum and thus treats Parkinson's disease. This strategy has been adopted in the drug under development, AXO-Lenti-PD (OXB-102), a Parkinson's disease gene therapy drug targeting dopamine synthetase jointly developed by Oxford Biomedica (UK) and Axovant Gene Therapy (US). Currently, this drug is in Phase I/II clinical trials. Evidence from animal and clinical trial data has demonstrated that the drug can effectively increase local dopamine in the midbrain striatum and improve associated Parkinson's disease symptoms. Long-term pharmacodynamic studies are still under investigation. However, 3-gene vectors packaged with lentiviruses exhibit low transduction efficiency and low dopamine synthesis efficiency in neurons in vivo, which does not lead to sufficient improvement in efficacy in preclinical animal models and clinical efficacy. Adeno-associated virus (AAV) vectors belong to the genus Dependovirus of the family Parvoviridae and are a type of very small, envelope-less, linear single-stranded DNA virus. Due to high transfection efficiency and high safety, they have become one of the most widely used viral vectors in gene therapy. Due to the packaging capacity limitations of AAV vectors, they can only package DNA fragments of 4.7 kb or less. Currently, there are no ongoing clinical trials for AAVs packaging three genes to express dopamine synthetase for the treatment of Parkinson's disease. The U.S. company Voyager Therapeutics achieves the treatment of Parkinson's disease by using AAV2-hAADC as a vector to efficiently express AADC and injecting it intracranially into the patient's putamen to improve the efficiency of converting levodo