The impact of GPIbα on platelet-targeted FVIII gene therapy in hemophilia A mice with pre-existing anti-FVIII immunity

Abstract

Essentials Platelet-specific FVIII gene therapy is effective in hemophilia A mice even with inhibitors. The impact of platelet adherence via VWF/GPIbα binding on platelet gene therapy was investigated. GPIbα does not significantly affect platelet gene therapy of hemophilia A with inhibitors. Platelet gene therapy induces immune tolerance in hemophilia A mice with pre-existing immunity. Summary: Background We have previously demonstrated that von Willebrand factor (VWF) is essential in platelet-specific FVIII (2bF8) gene therapy of hemophilia A (HA) with inhibitory antibodies (inhibitors). At the site of injury, platelet adherence is initiated by VWF binding to the platelet GPIb complex. Objective To investigate the impact of GPIbα on platelet gene therapy of HA with inhibitors. Methods Platelet-FVIII expression was introduced by 2bF8 lentivirus (2bF8LV) transduction of hematopoietic stem cells (HSCs) from GPIbα null (Ib null ) mice or rhF8-primed FVIII null (F8 null ) mice followed by transplantation into lethally irradiated rhF8-primed F8 null recipients. Animals were analyzed by flow cytometry, FVIII assays and the tail bleeding test. Results After transplantation, 99% of platelets were derived from donors. The macrothrombocytopenia phenotype was maintained in F8 null mice that received 2bF8LV-transduced Ib null HSCs (2bF8-Ib null /F8 null ). The platelet-FVIII expression level in 2bF8-Ib null /F8 null recipients was similar to that obtained from F8 null mice that received 2bF8LV-transduced F8 null HSCs (2bF8-F8 null /F8 null ). The tail bleeding test showed that the remaining hemoglobin level in the 2bF8-Ib null /F8 null group was significantly higher than in the F8 null control group, but there was no significant difference between the 2bF8-Ib null /F8 null and 2bF8-F8 null /F8 null groups. The half-life of inhibitor disappearance time was comparable between the 2bF8-Ib null /F8 null and 2bF8-F8 null /F8 null groups. The rhF8 re-challenge did not elicit a memory immune response once inhibitor titers dropped to undetectable levels after 2bF8 gene therapy. Conclusion GPIbα does not significantly impact platelet gene therapy of HA with inhibitors. 2bF8 gene therapy restores hemostasis and promotes immune tolerance in HA mice with pre-existing immunity.