JCI Insight (06/20/2019) Vol. 4, No. 12. Samelson-Jones, Benjamin J.; Finn, Jonathan D.; George, Lindsey A.; et al.
The hyperactive factor IX Padua variant (FIX-R338L) is being evaluated in multiple adeno-associated-viral (AAV) vector liver-directed gene therapy trials for patients with hemophilia B. This high specific activity variant may allow for reduced dosing of the vector relative to FIX-WT and reduced potential for AAV-related hepatotoxicity, while still providing clinical benefit. However, because the biochemical mechanism of this variant’s hyperactivity is not well defined, there are safety questions related to unregulated coagulation and the potential for thrombotic complications. The authors compared purified recombinant protein of FIX-WT and FIX-R338L variants to evaluate differences in enzymatic and clotting activity, activation, inactivation, and cofactor dependence. Their results from studies in purified-protein and plasma-based systems suggest similar molecular regulation of FIX-R338L and FIX-WT, and demonstrate that FIX-R338L hyperactivity derives from an enhanced interaction with FVIIIa that results in accelerated FX activation. The mechanism of enhanced allosteric activation of FIXa-R338L by FVIIIa (relative to FIX-WT) for the gain-of-function associated with this variant helps to reduce safety concerns of potential thrombotic complications in the context of AAV-based clinical hemophilia B gene therapy studies.