IGHV sequencing of temporal follicular lymphoma samples

N-glycosylation sites in the variable region of the immunoglobulin heavy chain is a characteristic feature of follicular lymphoma (FL) B cells and is present in >90% of all FL patients. Unique attached oligomannoses activate B cell receptor signalling pathways following engagement with calcium-dependent lectins expressed by tissue macrophages. This novel interaction appears critical for tumour growth and survival. To elucidate the significance of N-gly site presence and loss during ongoing SHM, we tracked site behavior during tumour evolution and progression in a diverse group of patients through next-generation sequencing. A hierarchy of subclones was visualised through lineage trees based on SHM semblance between subclones and their discordance from the germline sequence. We observed conservation of N-gly sites in >96% of subclone populations within and across diagnostic, progression and transformation events. Rare N-gly-negative subclones were lost or negligible from successive events in contrast to N-gly-positive subclones which could additionally migrate between anatomical sites. Ongoing SHM of the N-gly sites resulted in subclones with different amino acid compositions across disease events, yet the vast majority of resulting DNA sequences still encoded for an N-gly site. The selection and expansion of only N-gly-positive subclones is evidence of the tumour cells dependence on sites despite the changing genomic complexity as the disease progresses. N-gly sites were gained in the earliest identified lymphoma cells, indicating they are an early and stable event of pathogenesis. Targeting the inferred mannose-lectin interaction holds therapeutic promise.