The authors acknowledge support by Cancer Research UK (C30122/A11527; C30122/A15774: SNK, JFS, DHJ, HJB, LS) (C16736/A8371: FON, KMI); the Medical Research Council (MR/L023091/1) (SNK, FON); the Dermatrust (KEL, IUE, FON); the British Skin Foundation (S633) (KEL, IUE, FON, SNK); Mary Dunhill Trust (FON); Breakthrough Breast Malignancy/Breast Cancer Now (147) (KMI, SNK, FON); the Academy of Medical Sciences (DHJ, JFS, SNK); CR UK//NIHR in England/DoH for Scotland, Wales and Northern Ireland Experimental Cancer Medicine Centre (C10355/A15587) (FON, JFS, SNK, SL); CR UK/EPSRC/MRC/NIHR KCL/UCL Comprehensive Cancer Imaging Centre (C1519/A10331) (DHJ, JFS, SNK)

The authors acknowledge support by Cancer Research UK (C30122/A11527; C30122/A15774: SNK, JFS, DHJ, HJB, LS) (C16736/A8371: FON, KMI); the Medical Research Council (MR/L023091/1) (SNK, FON); the Dermatrust (KEL, IUE, FON); the British Skin Foundation (S633) (KEL, IUE, FON, SNK); Mary Dunhill Trust (FON); Breakthrough Breast Malignancy/Breast Cancer Now (147) (KMI, SNK, FON); the Academy of Medical Sciences (DHJ, JFS, SNK); CR UK//NIHR in England/DoH for Scotland, Wales and Northern Ireland Experimental Cancer Medicine Centre (C10355/A15587) (FON, JFS, SNK, SL); CR UK/EPSRC/MRC/NIHR KCL/UCL Comprehensive Cancer Imaging Centre (C1519/A10331) (DHJ, JFS, SNK). from circulating B cells. Consistent with AID, comparable somatic hypermutation frequencies and class-switching indicated affinity-matured antibodies in normal and malignant skin. A melanoma-associated antibody subset featured shorter complementarity-determining (CDR3) regions relative to those from circulating B cells. Clonal amplification in melanoma-associated antibodies and homology modeling indicated differential potential antigen recognition profiles between normal skin and melanoma sequences, suggesting distinct antibody repertoires. Evidence for IgG-expressing B cells, class switching and antibody maturation in normal and malignant skin and clonally-expanded antibodies in melanoma, support the involvement of mature B cells in cutaneous immunity. Despite being important immune sentinels in inflammation, antigen presentation, and adaptive immunity through antibody production, the recruitment and functions of B cells and the humoral immune compartment in cancer immune surveillance and in normal tissue homeostasis are insufficiently understood. B cells that are exposed to antigens in peripheral tissues can undergo clonal growth and class switching to mature antibody classes (IgG1-4, IgA1-2, IgE). Antigen challenge triggers daughter cells to undergo somatic hypermutation (SHM) and Vinflunine Tartrate to express antibodies with increasing affinity for the specific antigen. Class switch recombination (CSR) and SHM, involving the enzyme Activation-induced cytidine Deaminase (AID) can occur both in lymph node germinal centers and also in tissues (e.g. lung, nasal mucosa) in response to antigenic challenge. This provides an enriched antibody repertoire with reactivity and affinity against encountered antigens and of different isotypes, conferring the potential to generate antibodies with a variety of Fc-mediated immune effector functions1,2,3. The presence and nature of skin-resident B cells are ill-defined due to low cutaneous B cell infiltrate numbers. Preliminary findings describe a ENPP3 subset of B cells distinct from those in lymph nodes, circulating through sheep skin4. Moreover, potential functions for B cells in cutaneous inflammation, allergy and autoimmunity and in skin malignancy are reported5,6,7, suggesting immune surveillance in the context of inflammation or antigenic challenge in skin. In cutaneous melanomas, IgG-producing B cells may infiltrate tumors and form a part of tertiary lymphoid structures8,9. Clonal growth of IgG-expressing clones against tumor-associated antigens has been reported to correspond to clinical tumor regression10. Taken together, these studies support potential functions for mature humoral responses in normal and inflamed cutaneous sites. We provide the first report of the human mature skin-resident B cell compartment and its IgG-expressing profiles in cutaneous melanoma and in normal skin. We describe evidence for the presence of cutaneous mature B cells, distinct IgG subclass distribution profiles, clonal growth, somatic hypermutation in the IgG heavy chain variable regions, and predicted antigen binding site characteristics of the mature humoral response repertoire in cutaneous malignant melanoma lesions and in normal skin. Results B cells are present in melanoma lesions and normal skin We aimed to investigate B cell surveillance in cutaneous sites. We found that a small proportion of circulating CD45+CD3-CD14-CD19+CD22+B cells in healthy volunteers (n?=?24) and patients with melanoma (n?=?49) express the skin-homing Cutaneous Leucocyte-associated Antigen (CLA) (Fig. 1a, Physique S1a). Immunohistochemical evaluations revealed CD22+ cells in normal skins and melanomas (n?=?189, Fig. 1b, Physique S1b). We detected low frequencies of CD22+ infiltrates in 31.3% of normal skin samples (n?=?16). CD22+ infiltrates were found in 37.6% of melanomas (27% cutaneous lesions, 49.1% lymph node metastases, 38% distant metastases), with ~10% of melanomas featuring denser B cell infiltrating populations (>10 cells per high powered field, Fig. 1b,c). Cutaneous B cell infiltrates from non-malignant skin and melanoma lesion samples were also confirmed by flow cytometric analyses of CD45+CD19+CD22+B Vinflunine Tartrate cells (Fig. 1d, for matched normal skin and melanoma lesion B cells and peripheral blood B cells from a single Vinflunine Tartrate donor, representative of n?=?4; Physique S2 for further examples of CD45+CD19+B.