Adeno-associated virus vector delivery of muscle specific micro-dystrophin to treat muscular dystrophy

A small technology for muscular dystrophy, applied in the direction of muscle protein, muscle system disease, virus/phage, etc., can solve problems such as limiting muscle regeneration, muscle fiber degeneration, necrosis, etc.

Pending Publication Date: 2021-05-18
RES INST AT NATIONWIDE CHILDRENS HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the main pathological features of DMD are muscle fiber degeneration and necrosis, fibrosis as a result of the pathology has the same consequences
Excessive production of fibrotic tissue limits muscle regeneration and contributes to progressive muscle weakness in DMD patients

Method used

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  • Adeno-associated virus vector delivery of muscle specific micro-dystrophin to treat muscular dystrophy
  • Adeno-associated virus vector delivery of muscle specific micro-dystrophin to treat muscular dystrophy
  • Adeno-associated virus vector delivery of muscle specific micro-dystrophin to treat muscular dystrophy

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0179] A) Generation of AAVrh74.MHCK7.mini-dystrophin constructs

[0180] The AAVrh74.MHCK7.microdystrophin particle contains a human microdystrophin cDNA expression cassette flanked by AAV2 inverted terminal repeats (ITRs) (see figure 1 ). The mini-dystrophin construct is characterized by an in-frame rod domain deletion (R4-R23), while hinges 1, 2 and 4 and the cysteine-rich domain still give rise to a 138 kDa protein. Expression of microdystrophin (3579bp) is directed by the MHCK7 promoter (792bp). The plasmid was constructed by removing the MCK promoter from the rAAV.MCK.microdystrophin plasmid and inserting the MHCK7 promoter. After the core promoter, a 53bp endogenous mouse MCK exon 1 (untranslated) is provided for efficient transcription initiation, followed by the SV40 late 16S / 19S splicing signal (150bp) and a small 5'UTR (61bp ). Introns and 5'UTR were derived from plasmid pCMVβ (Clontech). The mini-dystrophin cassette has a consensus sequence Kozak preceding the...

Embodiment 2

[0187] Systemic gene delivery clinical trial for Duchenne muscular dystrophy

[0188] This is a single dose controlled trial using rAAVrh74.MHCK7.microdystrophin of SEQ ID NO:3, nucleotides 55-5021, in DMD subjects. Cohort A will include six subjects aged 3 months to 3 years and Cohort B will include six subjects aged 4 to 7 years. All subjects will receive intravenous microdystrophin carrier (2 x 10 14 vg / kg, 10mL / kg) injection. rAAVrh74.MHCK7.microdystrophin in the presence of 20mM Tris (pH8.0), 1mM magnesium chloride (MgCl 2 ), 200mM sodium chloride (NaCl) and 0.001% poloxamer 188 buffer.

[0189] In this study, rAAVrh74.MHCK7.microdystrophin was infused via a peripheral arm vein, allowing access to all muscles in the body. Six DMD subjects aged 3 months to 3 years were enrolled in Cohort A, and six DMD subjects aged 4 to 7 years were enrolled in Cohort B. All subjects received intravenous microdystrophin carrier (2 × 10 14 vg / kg, 10mL / kg) injection. The capsid-encap...

Embodiment 3

[0256] A randomized double-blind placebo-controlled phase I / IIa clinical trial of systemic gene delivery

[0257] This is a randomized double-blind single-dose trial with rAAVrh74.MHCK7.microdystrophin in DMD subjects. The study included twenty-four subjects aged 4 to 7 years. Subjects were randomly assigned to treatment or placebo at enrollment. Twelve subjects received intravenous rAAVrh74.MHCK7.mini-dystrophin vector (2×10 14 vg / kg, approximately 10 mL / kg), while another twelve subjects will receive 10 mL / kg placebo (Lactated Ringer's solution). Placebo subjects will continue treatment in the same manner as the 12 previously treated subjects one year after the last treated subject is dosed. Subjects received an infusion of rAAV with microdystrophin or lactated Ringer's solution over approximately 1 hour. Needle muscle biopsies were performed on the gastrocnemius muscle before and after treatment (90 days).

[0258] The primary objective of the study was to evaluate the...

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Abstract

The invention provides gene therapy vectors, such as adeno-associated virus (AAV) vectors, expressing a miniaturized human micro-dystrophin gene and method of using these vectors to express micro-dystrophin in skeletal muscle s including diaphragm and cardiac muscle and to protect muscle fibers from injury, increase muscle strength and reduce and / or prevent fibrosis in subjects suffering from muscular dystrophy.

Description

[0001] This application claims U.S. Provisional Patent Application No. 62 / 686,668, filed June 18, 2018; U.S. Provisional Patent Application No. 62 / 740,402, filed October 2, 2018; U.S. Provisional Patent Application No. 62 / 740,402, filed October 30, 2018 62 / 752,841; U.S. Provisional Patent Application No. 62 / 823,649, filed March 25, 2019, and U.S. Provisional Patent Application No. 62 / 860,220, filed June 11, 2018, each of which are incorporated herein by reference in their entirety. [0002] Incorporation by reference of electronically submitted material [0003] As a separate part of this disclosure, this application contains a Sequence Listing in computer readable form, which is incorporated herein by reference in its entirety and identified as follows: File Name: 53169_Seqlisting.txt; Size: 60,056 bytes, Date Created: 2019 June 17th. technical field [0004] The present invention provides gene therapy vectors expressing a miniaturized human microdystrophin gene, such as ad...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/86A61K48/00
CPCC12N15/86C12N2750/14141A61K48/0058C07K14/4716A61K48/005A61K48/0075A61K48/0083A61P21/00
Inventor L·罗迪诺-克拉帕奇J·R·门德尔
Owner RES INST AT NATIONWIDE CHILDRENS HOSPITAL
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