High throughput analysis of B cell dynamics and neutralizing antibody development during immunization with a novel clade C HIV-1 envelope

A protective HIV-1 vaccine has been hampered by a limited understanding of how B cells acquire neutralizing activity. Our previous vaccines expressing two different HIV-1 envelopes elicited robust antigen specific serum IgG titers in 20 rhesus macaques; yet serum from only two animals neutralized the autologous virus. Here, we used high throughput immunoglobulin receptor and single cell RNA sequencing to characterize the overall expansion, recall, and maturation of antigen specific B cells longitudinally over 90 weeks. Diversification and expansion of many B cell clonotypes occurred broadly in the absence of serum neutralization. However, in one animal that developed neutralization, two neutralizing B cell clonotypes arose from the same immunoglobulin germline and were tracked longitudinally. Early antibody variants with high identity to germline neutralized the autologous virus while later variants acquired somatic hypermutation and increased neutralization potency. The early engagement of precursors capable of neutralization with little to no SHM followed by prolonged affinity maturation allowed the two neutralizing lineages to successfully persist despite many other antigen specific B cells. The findings provide new insight into B cells responding to HIV-1 envelope during heterologous prime and boost immunization in rhesus macaques and the development of selected autologous neutralizing antibody lineages 20 Indian origin rhesus macaques (Macaca mulatta), including 17 male and 3 female, were immunized intramuscularly as follows: week 0 and week 8 with pGA1-SHIV-1-R66M/Z1800M DNA; week 16 and week 24 with recombinant Modified Vaccinia Ankara(MVA)-SHIV-1-R66M/Z1800M; weeks 53 and 61 with either gp120 or trimer gp140 protein HIV-1 envelope protein. The animals were split into 2 groups of 10 animals each and one group received reagents derived from the HIV-1 T/F Env Z1800M E4.02 sequence while the other received reagents derived from the HIV-1 T/F Env R66M O20.03 sequence. For the protein immunizations, the groups of 10 were further subdivided with 5 animals receiving gp120 protein and the other 5 animals receiving the stabilized gp140 trimer.