Figure 7. Clonal redemption in L54/L54 individuals.

Residues implicated in L54-IGHV1-69 encoded poly- and autoreactivity, as shown for 310-39G10 Fab (A) and 310-36E6 Fab (B). HA trimers are shown in gray and antibodies are colored as red and blue. (C) HA stalk affinity for human and HC2 mouse stalk specific L54 germline Fabs with or without the L54F/R49G/I56T substitutions (See also Figures 2E, F, S5, and Tables S3, S4, S5, and S7). Human mAbs are denoted in blue while HC2 mAbs are denoted in purple (P>0.05, T-test). (D) Pathways of clonal redemption for affinity matured human (310-39G10, 310-36E6) and HC2 L54 stalk-targeting mAbs (L69_m76, and L69_m34). (E-F) Autoreactivity of mature and V gene-reverted (gHgL) antibodies as evaluated by binding to Hep-2 cells (E) and cardiolipin (F, **P<0.008, Paired T-test). Human mAbs are denoted in blue and HC2 mAbs are denoted in purple. Scale bar in (E) denotes 25μm. (G) Immunization scheme for CTLA-4 blockade in L54/L54 HC2 mice vaccinated with SS-np. (H) Endpoint titers for isotype control treated (dark blue) and anti-CTLA-4 treated (light blue) at day 14, 35 and 56 (mean and SEM, n=5 mice per group, *P<0.03, **P<0.002, ANOVA with Tukey’s test). (I) Day 56 endpoint dilution for stalk specific IgG (n=5 mice per group, mean and SEM, *P<0.05, T-test).