Biophysical constraints on antibody affinity maturation to SARS-CoV-2

PROJECT SUMMARY/ABSTRACT The objective of this proposal is to develop a quantitative understanding of how the biophysical properties of antibodies impact their capacity to evolve affinity to divergent SARS-CoV-2 spike variants. Though there is substantial evidence that mutations acquired during affinity maturation impact antibody expression, affinity for distinct viral variants, and self-reactivity, we lack a quantitative understanding of (1) how mutations impact these biophysical properties and (2) how these properties, and trade-offs between them, collectively determine the fate of the corresponding B-cell lineage. Here, we propose three Aims to test our hypothesis that mutations differentially impact antibody expression, affinity, and self-reactivity, resulting in biophysical trade-offs that constrain the evolution of antibodies that bind divergent SARS-CoV-2 spike variants. In Aim 1, we quantitate the biophysical effects of mutations in anti-SARS-CoV-2 spike antibodies, using high-throughput mammalian cell- display methods we recently developed. By measuring the expression, affinity, and self-reactivity for millions of anti-spike antibodies, including broadly neutralizing antibodies (bnAbs) that bind divergent spike variants, their evolutionary predecessors, and systematically mutagenized antibody sequences, we will unveil biophysical constraints that shape affinity maturation to rapidly evolving viral antigens. In Aim 2, we evaluate the contributions of antibody biophysical properties to B-cell fitness, or proliferation, using longitudinally-sampled patient B-cells following exposure to divergent strains of SARS-CoV-2. This approach will reveal the relative importance of distinct antibody biophysical properties in driving B-cell evolutionary dynamics in human repertoires and enable development of quantitative models for predicting the outcomes of affinity maturation. In Aim 3, we define the impact of selection pressure during affinity maturation on the biophysical properties of the resulting antibodies, focusing on selection regimes known to favor the maturation of bnAbs that bind distinct spike variants. To this end, we leverage a B-cell directed evolution platform that mimics the mutagenic load of somatic hypermutation, enables fine-tuning of the antibody selection conditions, and supports longitudinal B-cell sampling to profile the evolutionary dynamics of the B-cell response and the biophysical properties of the corresponding antibody lineages. The resulting data will be used to define the impact of the selection regime on the biophysical determinants of B-cell fitness. Successful completion of these Aims will yield quantitative insight into (1) how antibody biophysical properties change during affinity maturation, (2) how they collectively determine B-cell fate in human repertoires, and (3) how their relative importance varies across distinct selection regimes. Thus, this work will advance our fundamental understanding of the biophysical mechanisms that shape antibody affinity maturation to rapidly evolving pathogens like SARS-CoV-2, supporting efforts to design and elicit antibodies that bind existing and novel viral variants.