Although B16LS9 cells express functional TLR5 and respond to entolimod treatment with NF-κB activation (indicated by p65 translocation to the nucleus 30 min after in vitro treatment with entolimod (Supplementary Figure S1)), there was no significant difference in the size of primary melanomas in the eyes of entolimod-treated (all three treatment schedules) versus vehicle-treated mice as measured on Day 7 after tumor cell administration (Figure 1A, Supplementary Figure S2A). Here, NFKB1 is linked to melanoma.