Figure 5.

Modeling Spontaneous Mutations or a Recombination Event

(A) LD₅₀ values for serially passaged and engineered viruses. A theoretical concern with a recombinant is that secondary mutations could develop during replication, thus limiting its safety and clinical utility. To assess this, we serially passaged C134 in vivo and found that after six passages in the CNS of mice, the virus retained it pre-passage LD₅₀ profile. Another theoretical concern with an oHSV vector containing a foreign gene is that a recombination event could produce an HSV with greater virulence or encephalitic potential than the WT species. To test this hypothesis, we mimicked such a recombination event and created C124 (WT+HCMV IRS1). LD₅₀ studies show that insertion of HCMV IRS1 into the U_L3/U_L4 intergenic region of WT HSV does not increase HSV-1 (LD₅₀ of 86 PFU versus 18 PFU) virulence. (B and C) Furthermore, using dose-matched groups, 330 PFU (B) and 1,000 PFU (C) HSV-1 was more virulent than C124 and led to earlier death in the IC-administered mice in survival studies. A full summary of animal numbers and timing of deaths for all dose groups is provided in Figure S4.