Long-lasting germinal center responses to a priming immunization with continuous proliferation and somatic mutation

Summary Germinal centers (GCs) are the engines of antibody evolution. Using HIV Env protein immunogen priming in rhesus monkeys (RM) followed by a long period without further immunization, we demonstrate GC B cells (B GC ) lasted at least 6 months (29 weeks), all the while maintaining rapid proliferation. A 186-fold B GC cell increase was present by week 10 compared to a conventional immunization. Single cell transcriptional profiling revealed that both light zone and dark zone GC states were sustained throughout the 6 months. Antibody somatic hypermutation (SHM) of B GC cells continued to accumulate throughout the 29 week priming period, with evidence of selective pressure. Additionally, Env-binding B GC cells were still 49-fold above baseline 29 weeks after immunization, suggesting that they could be active for significantly longer periods of time. High titers of HIV neutralizing antibodies were generated after a single booster immunization. Fully glycosylated HIV trimer protein is a complex antigen, posing significant immunodominance challenges for B cells, among other difficulties. Memory B cells (B Mem ) generated under these long priming conditions had higher levels of SHM, and both B Mem cells and antibodies were more likely to recognize non-immunodominant epitopes. Numerous B GC cell lineage phylogenies spanning the >6-month GC period were identified, demonstrating continuous GC activity and selection for at least 191 days, with no additional antigen exposure. A long prime, adjuvanted, slow delivery (12-day) immunization approach holds promise for difficult vaccine targets, and suggests that patience can have great value for tuning GCs to maximize antibody responses.