Therapeutic base editing to generate a gain-of-function F9 variant for hemophilia B

The repair of pathological gene variants is an ultimate goal in treating genetic diseases; however, developing distinct therapeutic reagents for each of the numerous variants within a gene may not be scalable. Here, we investigated whether base editing to introduce a gain-of-function variant in blood coagulation factor IX (FIX) can increase FIX activity as a targeted therapeutic approach for hemophilia B. We engineered a G:C to A:T substitution at c.1151 of F9 by cytosine base editing to generate R338Q (the Shanghai F9 variant), which markedly increases coagulation factor activity. An adeno-associated virus vector harboring the base editor converted >60% of the target G:C to A:T and increased FIX activity in HEK293 cells harboring patient-derived F9 variants as well as in knock-in mice carrying a human F9 complementary DNA. Furthermore, administration of lipid nanoparticles containing the base-editor mRNA and guide RNA increased FIX activity in mice. These data indicate that cytosine base editing to generate R338Q in FIX is a broadly applicable genome-editing strategy for hemophilia B with residual FIX activity.