Computationally designed liver-specific transcriptional modules and hyperactive factor IX improve hepatic gene...

Area of Science:

• Gene Therapy
• Hematology
• Molecular Biology

Background:

• Hemophilia B treatment requires effective gene therapy vectors.
• Current vectors necessitate high doses, posing safety concerns.
• Enhancing therapeutic efficacy at lower doses is crucial for next-generation treatments.

Purpose of the Study:

• To develop safer and more effective gene therapy vectors for hemophilia B.
• To identify regulatory elements that increase factor IX (FIX) expression in hepatocytes.
• To combine these elements with a hyperfunctional FIX transgene for enhanced efficacy.

Main Methods:

• Computational identification of hepatocyte-specific cis-regulatory modules (CRMs).
• Engineering a synthetic, codon-optimized, hyperfunctional FIX-R338L Padua transgene.
• Utilizing self-complementary adenoassociated virus serotype 9 (scAAV9) vectors for delivery in hemophilia B mice.

Main Results:

• Hepatocyte-specific CRMs increased FIX levels 11- to 15-fold.
• The FIX-R338L Padua mutation boosted FIX activity up to sevenfold without increasing thrombotic risk.
• Sustained supraphysiologic FIX activity (400%) and bleeding correction were achieved at low vector doses (5 × 10(10) vg/kg).
• Induced immune tolerance prevented inhibitory antibody formation against FIX.

Conclusions:

• Combining hepatocyte-specific CRMs with a hyperfunctional FIX transgene offers a potent gene therapy strategy for hemophilia B.
• This approach enables sustained therapeutic FIX levels at safe, low vector doses.
• The developed vectors hold promise for safe and effective hemophilia B treatment, with potential for immune tolerance induction.