Saunderset al.,Science 366 , eaay7199 (2019) 6 December 2019 5of17
Fig. 2. CH848 10.17DT nanoparticle immunization of DH270 UCA knock-in
mice induces serum neutralizing antibody responses and the improbable
G57R mutation.(A) Negative-stain electron microscopy of CH848 10.17DT–
stabilized Env trimers conjugated to ferritin nanoparticles. A magnified image of
the Env nanoparticles (top) and 2D class average (bottom) of the Env nano-
particles are shown at the right. (B) Antigenic profile of CH848 10.17DT–
stabilized Env trimers conjugated to ferritin nanoparticles or unconjugated.
Binding was determined by biolayer interferometry and normalized to PGT151
binding values. The normalized binding is shown in the heat map, with a value of
1 being equal to PGT151 binding. OD glycan, outer domain glycans in the high-
mannose patch. (C) CH848 10.17DT Env trimer nanoparticle (red) induction of
calcium flux in homozygous (VH+/+,VL+/+) DH270 UCA double knock-in (double KI)
mouse B cells. (D)Heterozygous(VH+/–,VL+/–)DH270UCAdoubleKImouse
immunization regimen with CH848 10.17DT SOSIP trimer nanoparticles. (E)Quan-
tification of the frequency of splenic germinal center B cells (top) and follicular
helper T cells (bottom) in vaccinated DH270 UCA mice. Unvaccinated, wild-type
C57BL/6 mice (BL6) served as baseline controls. Means ± SEM are shown by black
bars. *P< 0.05 (Wilcoxon exact test). (F) Serum antibody neutralization of
autologous tier 2 CH848 10.17DT (top) and CH848 10.17 (bottom) virus infection of
TZM-bl cells. Group geometric mean ID 50 titers are shown (nanoparticle,n=5;
adjuvant only,n= 6). Arrows indicate immunization time points. (G) Top: Enumeration
by VHnext-generation sequencing of the frequency of unique DH270 sequences
encoding the G57R amino acid change. Frequencies of G57R were determined in
total splenocytes 1 week after the sixth immunization. Each mouse is shown by
an individual symbol; horizontal bars indicate the group mean. Bottom: Relative
(fold) increase in G57R frequency in heterozygous DH270 UCA KI mice immunized
with CH848 10.17DT Env nanoparticle and adjuvant compared to adjuvant-only
control mice. Box-and-whisker plots indicate the minimum, maximum, median,
and interquartile range fold increase for each group. Each symbol represents an
individual mouse. *P< 0.05 (Wilcoxon exact test).RESEARCH | RESEARCH ARTICLE
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