O18 The role of B cells and tertiary lymphoid structures for immune surveillance in cutaneous squamous cell carcinoma

Abstract Introduction and aims Cutaneous squamous cell carcinoma (cSCC), the second most common form of skin cancer, has recently been associated with B- and T-cell clusters which resembled tertiary lymphoid structures (TLSs). These structures are of particular interest due to the lack of B cells present in healthy human skin. Our group have previously identified a role for B cells, particularly IgE, in limiting cSCC growth in mice, and demonstrated a positive correlation between high-affinity IgE receptor expression and disease outcome in humans. Therefore, we hypothesize that B cells play a positive role in anti-cSCC immunity, with selection and expansion of B-cell subsets occurring locally, within TLSs. Methods We used high-parameter spectral-unmixing flow cytometry on human tissue samples, complemented by both 5-colour confocal and multiplex CellDIVE microscopy. B-cell repertoire sequencing was then used to compare the B-cell clones between matched blood, perilesional and cSCC tissue. Results We have demonstrated that patients with cSCC have elevated levels of IgE+ memory B cells in the blood, compared with healthy donors. In the tissue, there was a significantly increased population of B cells in tumours, which constituted equal ratios of classical memory cells and class-switched double-negative B cells. We also identified germinal centre-like B cells in cSCC tissue. Microscopy unveiled the proximity of these B cells both to the tumour itself, and a multitude of other immune cells, including T-cell subsets, dendritic cells and macrophages. Analysis of the repertoire sequencing is still ongoing; however, initial analysis suggests the presence of a highly diverse B-cell repertoire in the tumours, with a mix of subclasses. Conclusions This work begins to unveil the complex function of B cells and TLSs in cSCC immunity.