OC 30.4 - Efficacy and Safety of AAV Delivery of a Novel Factor VIII Variant with Improved Secretion in a Non-Human Primate Model

Monday, June 26, 2023

11:30 – 11:45 ET

Room: 517

Session Available As: Live-Stream and On-Demand

Co-Author(s)

MS

Maria C. Seleme

Research Scientist
The Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States

Presenting Author(s)

Jonathan R. Lindgren, B.S. (he/him/his)

Research Technician
The Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States

Co-Author(s)

GN

Giang N. Nguyen

Research Associate
The Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
AH

Allysen C. Henriksen

Research Technician
The Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
CA

Charles-Antoine Assenmacher

Technical Director, Comparative Pathology Core
School of Veterinary Medicine, University of Pennsylvania
Philadelphia, Pennsylvania, United States
AM

Amy Muehlmatt

Operations Manager
Comparative Medicine Services Core, The Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
GS

Geary R. Smith

Technical Director
Comparative Medicine Services Core, The Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
MM

Melanie McFadden

Technical Director
Comparative Medicine Services Core, The Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
Denise E. Sabatino, PhD (she/her/hers)

Research Associate Professor of Pediatrics
The Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States

Background: Current AAV-mediated approaches for hemophilia A that are in clinical development utilize codon-optimized wild type B-domain deleted human FVIII (hFVIII-BDD) transgenes. hFVIII-BDD is not secreted well in vitro or in vivo. Strategies to improve secretion of codon-optimized hFVIII would improve circulating FVIII protein levels at a lower vector dose.

Aims: We have characterized a novel hFVIII variant that has a deletion of the furin cleavage site (del3) and a modification of a cleavage site at 1657-58 (SP/DE). Studies of AAV delivery of this variant (hFVIII-del3-SP/DE) in hemophilia A mice and hemophilia A dogs demonstrated that hFVIII-del3-SP/DE is expressed at higher levels than hFVIII-BDD. To further evaluate this variant, we performed a dose-escalation study in non-human primates to compare AAV administration of hFVIII-BDD with hFVIII-del3-SP/DE.

Methods: African green monkeys were intravenously administered AAV8-TTRm-hFVIII-BDD or AAV8-TTRm-hFVIII-del3-SP/DE at three doses (2e12 vg/kg, 5e12 vg/kg, 1e13 vg/kg) and followed for 90 days. hFVIII antigen levels, liver function tests (ALT, AST), D-dimer analysis and liver pathology were evaluated.

Results: The median peak hFVIII antigen levels were 3 to 5-fold higher in the AAV8-TTRm-hFVIII-del3-SP/DE treated animals than in the animals treated with AAV8-TTRm-hFVIII-BDD. Although there was some variability in ALT and AST levels with some elevations observed, the elevations were not clinically concerning and remained < 3x the upper limit of normal. D-dimer levels did not increase above baseline levels. Liver histology (H&E) showed baseline levels of inflammation with no major changes in the status of the liver over the course of the study. Additional analysis of the vector genomes and FVIII protein are ongoing.

Conclusion(s): AAV delivery of the hFVIII-del3-SP/DE variant improves FVIII expression 3 to 5-fold without concerns in liver function, thrombosis or pathological findings. This variant provides a novel strategy for reducing the AAV vector dose and improving the safety profile of AAV-hFVIII.

Learning Objectives:

Explain why there is a need to develop more efficacious next generation AAV-mediated gene therapy systems for hemophilia A.
Understand how engineering minimal changes in the amino-acid sequence of wild type Factor VIII to generate the FVIII-delta3-SP/DE variant improves the secretion of the protein in vitro and after AAV-delivery in small and large hemophilia A animal models compared to wild type FVIII.
Appreciate that while AAV-delivered wild type FVIII and FVIII-delta3-SP/DE are equally safe in non-human primates, the latter also has better secretion in this large animal model and provides a promising alternative for next generation AAV-gene therapy clinical studies for hemophilia A.