B cells expressing authentic naive human VRC01-class BCRs can be recruited to germinal centers and affinity mature in multiple independent mouse models

Abstract

Animal models of human antigen-specific B cell receptors (BCRs) generally depend on “inferred germline” sequences, and thus their relationship to authentic naive human B cell BCR sequences and affinities is unclear. Here, BCR sequences from authentic naive human VRC01-class B cells from healthy human donors were selected for the generation of three BCR knockin mice. The BCRs span the physiological range of affinities found in humans, and use three different light chains (VK3-20, VK1-5, and VK1-33) found among subclasses of naive human VRC01-class B cells and HIV broadly neutralizing antibodies (bnAbs). The germline-targeting HIV immunogen eOD-GT8 60mer is currently in clinical trial as a candidate bnAb vaccine priming immunogen. To attempt to model human immune responses to the eOD-GT8 60mer, we tested each authentic naive human VRC01-class BCR mouse model under rare human physiological B cell precursor frequency conditions. B cells with high (HuGL18HL) or medium (HuGL17HL) affinity BCRs were primed, recruited to germinal centers, and they affinity matured, and formed memory B cells. Precursor frequency and affinity interdependently influenced responses. Taken together, these experiments utilizing authentic naive human VRC01-class BCRs validate a central tenet of germline-targeting vaccine design and extend the overall concept of the reverse vaccinology approach to vaccine development.

Vaccine elicitation of HIV broadly neutralizing antibodies from engineered B cells

“…Allelic inclusion. To assess the surface expression of endogenous LCs in engineered VRC01+ cells, B cells from knock-in mice expressing LCs with only human κ constant chains33 were purified, activated, and engineered using the H-targeting strategy. Fc Blocker (homemade mAb 2.4g2) was added to single-cell suspensions at 0.5 mg per 10 6 cells before antibody staining.…”

Development of a VRC01-class germline targeting immunogen derived from anti-idiotypic antibodies

“…It is possible that these off-target B cell responses will be recalled and predominate the B cell responses upon subsequent Env immunizations. In line with this, adoptive transfer experiments where B cells expressing putative VRC01 precursors or intermediates with defined B cell receptor (BCR) specificity are introduced into a wild-type (WT) mouse at a controlled frequency demonstrated that immunogens with high affinity and/or avidity for the target BCR were required for successful inter-clonal competition of rare target B cells following a priming immunization (Dosenovic et al, 2018;Abbott et al, 2018;Huang et al, 2020;Kato et al, 2020).…”

Antibody Focusing to Conserved Sites of Vulnerability: The Immunological Pathways for ‘Universal’ Influenza Vaccines

“…Defined as the tendency of the immune system to respond to complex antigens in a hierarchical manner, the immunodominance patterns enforced following influenza infection and vaccination prioritize the expansion of non-neutralizing antibodies against ‘off-target’ hypervariable features at the expense of ‘on-target’ responses engaging functionally conserved epitopes [ 1 , 78 , 79 , 80 , 81 , 82 , 83 ] ( Figure 2 ). Structure-based influenza immunogens applied within transgenic mouse systems bearing user-defined B cell receptor (BCR) repertoires, along with similar approaches using HIV antigens, have enabled experimental manipulation of these parameters [ 84 , 85 , 86 , 87 , 88 , 89 , 90 , 91 , 92 ]. This work indicates that the immunogenicity of a given epitope is a function of the frequency and affinity of the on-target versus off-target BCRs present in the antigen naïve germline repertoire.…”

“…This work indicates that the immunogenicity of a given epitope is a function of the frequency and affinity of the on-target versus off-target BCRs present in the antigen naïve germline repertoire. Subdominant antibody responses thus arise when low frequency and/or low affinity on-target BCRs are unable to compete for expansion within B cell germinal centers (GCs), allowing off-target (‘immuno-distracted’) B cells to then dominate the GC reaction [ 84 , 85 , 86 , 87 , 88 , 89 , 90 , 91 ]. Such off-target anti-influenza BCRs proceed to overshadow antibody affinity maturation, the downstream serum antibody response, and the development of B cell memory [ 85 , 86 ].…”