Efficient gene transfer in skeletal muscle with AAV-derived bicistronic vector using the FGF-1 IRES

Area of Science:

• Molecular Biology
• Gene Therapy
• RNA Biology

Background:

• Internal ribosome entry sites (IRESs) enhance translation under global blockade and enable polycistronic mRNA expression.
• Adeno-associated virus (AAV) vectors are limited by transgene cassette size, making short IRES elements desirable.
• Encephalomyocarditis virus (EMCV) IRES is commonly used but is long and inflexible.

Purpose of the Study:

• To evaluate the efficacy of short, flexible cellular IRESs, specifically fibroblast growth factor 1 (FGF-1) IRES, in AAV-derived bicistronic vectors.
• To compare the performance of FGF-1 IRES against the commonly used EMCV IRES in vivo for gene expression.

Main Methods:

• Construction and testing of AAV-derived bicistronic vectors expressing two luciferase genes.
• Utilized either EMCV IRES or FGF-1 IRES to separate the two reporter genes.
• Administered vectors into mouse muscle to assess in vivo transgene expression.

Main Results:

• AAV vectors utilizing the FGF-1 IRES achieved efficient and stable co-expression of both transgenes for at least 120 days post-administration.
• FGF-1 IRES mediated transgene expression approximately 10-fold higher than EMCV IRES in vivo.
• Stable stoichiometry of both transgene products was observed with the FGF-1 IRES vector.

Conclusions:

• Short, flexible cellular IRESs like FGF-1 are advantageous for AAV vector design due to size constraints.
• FGF-1 IRES demonstrates superior in vivo performance compared to EMCV IRES for bicistronic gene expression in skeletal muscle.
• FGF-1 IRES-containing AAV vectors hold significant promise for gene therapy applications requiring co-expression of therapeutic genes in skeletal muscle.