A Factor VIII Variant With Elimination of the N-glycosylation Site at 2118 Reduced Anti-FVIII Immune Responses in Gene Therapy Treated Hemophilia A Mice

Highlights From the ASGCT 25th Annual Meeting

A Factor VIII Variant With Elimination of the N-glycosylation Site at 2118 Reduced Anti-FVIII Immune Responses in Gene Therapy Treated Hemophilia A Mice

Presented by: Carol H. Miao, MD, Seattle Children’s Research Institute, Seattle, Washington, United States

Carol H. Miao^1,4, Meng-Ni Fan¹, Shuaishuai Wang², Junping Zhang³, Xiaohe Cai¹, Ting-Yen Chao¹, Lei Li², Weidong Xiao³, Barbara A. Konkle⁴

¹Seattle Children’s Research Institute, Seattle, WA
²Georgia State University, Atlanta, GA
³Indiana University, Indianapolis, IN
⁴University of Washington, Seattle, WA

Key Data Points

Inhibitor Development After Infusion of WT-BDD-FVIII and Mutated FVIII Variants
CD4+T-cell Proliferation in Response to Overlapping Mannosylated Peptides

Inhibitor Development After Infusion of WT-BDD-FVIII and Mutated FVIII Variants

Hemophilia A mice were injected hydrodynamically with plasmids encoding mutated FVIII variants and WT BDD-FVIII, respectively. Second challenges were performed via hydrodynamic injection in all groups on Day 86. (A) From left to right, the wild-type (WT)-BDD-FVIII plasmids used for N to Q mutagenesis, the resulting FVIII variant constructs used for in vivo gene therapy, and relative FVIII antibody response. (B) FVIII-specific IgG levels were analyzed by ELISA following first and second plasmid challenges. Anti-FVIII immune responses were significantly reduced in mice treated with the plasmid carrying FVIII N2118Q variant.

CD4+T-cell Proliferation in Response to Overlapping Mannosylated Peptides

(A) Overlapping peptides surrounding the N2118 site, 15 amino acids in length, were synthesized with high mannose glycan Man6GlcNAc2 attachments. (B) CD4+ T-cell proliferation levels were measured in response to mannosylated peptides MP1 (left panel), MP2 (middle panel), and MP3 (right panel) and their non-glycosylated counterparts (NGP1, NGP2, and NGP3). The data are presented as means with standard deviation from three separate experiments (**P < 0.01, ***P < 0.001).