B cell profiles, antibody repertoire and reactivity reveal dysregulated responses with autoimmune features in melanoma.

B cells are known to contribute to the anti-tumor immune response, especially in immunogenic tumors such as melanoma, yet humoral immunity has not been characterized in these cancers to detail. Here we show comprehensive phenotyping in samples of circulating and tumor-resident B cells as well as serum antibodies in melanoma patients. Memory B cells are enriched in tumors compared to blood in paired samples and feature distinct antibody repertoires, linked to specific isotypes. Tumor-associated B cells undergo clonal expansion, class switch recombination, somatic hypermutation and receptor revision. Compared with blood, tumor-associated B cells produce antibodies with proportionally higher levels of unproductive sequences and distinct complementarity determining region 3 properties. The observed features are signs of affinity maturation and polyreactivity and suggest an active and aberrant autoimmune-like reaction in the tumor microenvironment. Consistent with this, tumor-derived antibodies are polyreactive and characterized by autoantigen recognition. Serum antibodies show reactivity to antigens attributed to autoimmune diseases and cancer, and their levels are higher in patients with active disease compared to post-resection state. Our findings thus reveal B cell lineage dysregulation with distinct antibody repertoire and specificity, alongside clonally-expanded tumor-infiltrating B cells with autoimmune-like features, shaping the humoral immune response in melanoma.

Multiscale deep learning framework captures systemic immune features in lymph nodes predictive of triple negative breast cancer outcome in large-scale studies.

The suggestion that the systemic immune response in lymph nodes (LNs) conveys prognostic value for triple-negative breast cancer (TNBC) patients has not previously been investigated in large cohorts. We used a deep learning (DL) framework to quantify morphological features in haematoxylin and eosin-stained LNs on digitised whole slide images. From 345 breast cancer patients, 5,228 axillary LNs, cancer-free and involved, were assessed. Generalisable multiscale DL frameworks were developed to capture and quantify germinal centres (GCs) and sinuses. Cox regression proportional hazard models tested the association between smuLymphNet-captured GC and sinus quantifications and distant metastasis-free survival (DMFS). smuLymphNet achieved a Dice coefficient of 0.86 and 0.74 for capturing GCs and sinuses, respectively, and was comparable to an interpathologist Dice coefficient of 0.66 (GC) and 0.60 (sinus). smuLymphNet-captured sinuses were increased in LNs harbouring GCs (p < 0.001). smuLymphNet-captured GCs retained clinical relevance in LN-positive TNBC patients whose cancer-free LNs had on average ≥2 GCs, had longer DMFS (hazard ratio [HR] = 0.28, p = 0.02) and extended GCs' prognostic value to LN-negative TNBC patients (HR = 0.14, p = 0.002). Enlarged smuLymphNet-captured sinuses in involved LNs were associated with superior DMFS in LN-positive TNBC patients in a cohort from Guy's Hospital (multivariate HR = 0.39, p = 0.039) and with distant recurrence-free survival in 95 LN-positive TNBC patients of the Dutch-N4plus trial (HR = 0.44, p = 0.024). Heuristic scoring of subcapsular sinuses in LNs of LN-positive Tianjin TNBC patients (n = 85) cross-validated the association of enlarged sinuses with shorter DMFS (involved LNs: HR = 0.33, p = 0.029 and cancer-free LNs: HR = 0.21 p = 0.01). Morphological LN features reflective of cancer-associated responses are robustly quantifiable by smuLymphNet. Our findings further strengthen the value of assessment of LN properties beyond the detection of metastatic deposits for prognostication of TNBC patients. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

B Cells in Breast Cancer Pathology.

B cells have recently become a focus in breast cancer pathology due to their influence on tumour regression, prognosis, and response to treatment, besides their contribution to antigen presentation, immunoglobulin production, and regulation of adaptive responses. As our understanding of diverse B cell subsets in eliciting both pro- and anti-inflammatory responses in breast cancer patients increases, it has become pertinent to address the molecular and clinical relevance of these immune cell populations within the tumour microenvironment (TME). At the primary tumour site, B cells are either found spatially dispersed or aggregated in so-called tertiary lymphoid structures (TLS). In axillary lymph nodes (LNs), B cell populations, amongst a plethora of activities, undergo germinal centre reactions to ensure humoral immunity. With the recent approval for the addition of immunotherapeutic drugs as a treatment option in the early and metastatic settings for triple-negative breast cancer (TNBC) patients, B cell populations or TLS may resemble valuable biomarkers for immunotherapy responses in certain breast cancer subgroups. New technologies such as spatially defined sequencing techniques, multiplex imaging, and digital technologies have further deciphered the diversity of B cells and the morphological structures in which they appear in the tumour and LNs. Thus, in this review, we comprehensively summarise the current knowledge of B cells in breast cancer. In addition, we provide a user-friendly single-cell RNA-sequencing platform, called "B singLe cEll rna-Seq browSer" (BLESS) platform, with a focus on the B cells in breast cancer patients to interrogate the latest publicly available single-cell RNA-sequencing data collected from diverse breast cancer studies. Finally, we explore their clinical relevance as biomarkers or molecular targets for future interventions.

Interleukin-22 regulates neutrophil recruitment in ulcerative colitis and is associated with resistance to ustekinumab therapy.

The function of interleukin-22 (IL-22) in intestinal barrier homeostasis remains controversial. Here, we map the transcriptional landscape regulated by IL-22 in human colonic epithelial organoids and evaluate the biological, functional and clinical significance of the IL-22 mediated pathways in ulcerative colitis (UC). We show that IL-22 regulated pro-inflammatory pathways are involved in microbial recognition, cancer and immune cell chemotaxis; most prominently those involving CXCR2+ neutrophils. IL-22-mediated transcriptional regulation of CXC-family neutrophil-active chemokine expression is highly conserved across species, is dependent on STAT3 signaling, and is functionally and pathologically important in the recruitment of CXCR2+ neutrophils into colonic tissue. In UC patients, the magnitude of enrichment of the IL-22 regulated transcripts in colonic biopsies correlates with colonic neutrophil infiltration and is enriched in non-responders to ustekinumab therapy. Our data provide further insights into the biology of IL-22 in human disease and highlight its function in the regulation of pathogenic immune pathways, including neutrophil chemotaxis. The transcriptional networks regulated by IL-22 are functionally and clinically important in UC, impacting patient trajectories and responsiveness to biological intervention.

Leveraging the Dynamic Immune Environment Triad in Patients with Breast Cancer: Tumour, Lymph Node, and Peripheral Blood.

During the anti-tumour response to breast cancer, the primary tumour, the peripheral blood, and the lymph nodes each play unique roles. Immunological features at each site reveal evidence of continuous immune cross-talk between them before, during and after treatment. As such, immune responses to breast cancer are found to be highly dynamic and truly systemic, integrating three distinct immune sites, complex cell-migration highways, as well as the temporal dimension of disease progression and treatment. In this review, we provide a connective summary of the dynamic immune environment triad of breast cancer. It is critical that future studies seek to establish dynamic immune profiles, constituting multiple sites, that capture the systemic immune response to breast cancer and define patient-selection parameters resulting in more significant overall responses and survival rates for breast cancer patients.

Tumor-Infiltrating B Lymphocyte Profiling Identifies IgG-Biased, Clonally Expanded Prognostic Phenotypes in Triple-Negative Breast Cancer.

In breast cancer, humoral immune responses may contribute to clinical outcomes, especially in more immunogenic subtypes. Here, we investigated B lymphocyte subsets, immunoglobulin expression, and clonal features in breast tumors, focusing on aggressive triple-negative breast cancers (TNBC). In samples from patients with TNBC and healthy volunteers, circulating and tumor-infiltrating B lymphocytes (TIL-B) were evaluated. CD20+CD27+IgD- isotype-switched B lymphocytes were increased in tumors, compared with matched blood. TIL-B frequently formed stromal clusters with T lymphocytes and engaged in bidirectional functional cross-talk, consistent with gene signatures associated with lymphoid assembly, costimulation, cytokine-cytokine receptor interactions, cytotoxic T-cell activation, and T-cell-dependent B-cell activation. TIL-B-upregulated B-cell receptor (BCR) pathway molecules FOS and JUN, germinal center chemokine regulator RGS1, activation marker CD69, and TNFα signal transduction via NFκB, suggesting BCR-immune complex formation. Expression of genes associated with B lymphocyte recruitment and lymphoid assembly, including CXCL13, CXCR4, and DC-LAMP, was elevated in TNBC compared with other subtypes and normal breast. TIL-B-rich tumors showed expansion of IgG but not IgA isotypes, and IgG isotype switching positively associated with survival outcomes in TNBC. Clonal expansion was biased toward IgG, showing expansive clonal families with specific variable region gene combinations and narrow repertoires. Stronger positive selection pressure was present in the complementarity determining regions of IgG compared with their clonally related IgA in tumor samples. Overall, class-switched B lymphocyte lineage traits were conspicuous in TNBC, associated with improved clinical outcomes, and conferred IgG-biased, clonally expanded, and likely antigen-driven humoral responses. SIGNIFICANCE: Tumor-infiltrating B lymphocytes assemble in clusters, undergoing B-cell receptor-driven activation, proliferation, and isotype switching. Clonally expanded, IgG isotype-biased humoral immunity associates with favorable prognosis primarily in triple-negative breast cancers.

Immune Crosstalk Between Lymph Nodes and Breast Carcinomas, With a Focus on B Cells.

Lymph nodes (LNs) are highly organized secondary lymphoid organs, and reflective of immune responses to infection, injuries, or the presence of cancer. Extensive molecular and morphological analyses of immune and stromal features in tumors and LNs of breast cancer patients have revealed novel patterns indicative of disease progression. Within LNs, there are dynamic structures called germinal centers (GCs), that act as the immunological hubs for B cell development and generation of affinity matured memory B and antibody-producing plasma cells. Acting as a bridge between systemic and local immunity, associations are observed between the frequency of GCs within cancer-free LNs, the levels of stromal tumor infiltrating lymphocytes, and cancer progression. Scattered throughout the tumor microenvironment (TME) or aggregated in clusters forming tertiary lymphoid structures (TLS), the occurrence of tumor infiltrating B cells (TIL-Bs) has been linked mostly to superior disease trajectories in solid cancers. Recent TIL-Bs profiling studies have revealed a plethora of different TIL-B populations, their functional roles, and whether they are derived from GC reactions in the LN, and/or locally from GC-like structures within the TME remains to be investigated. However, parallels between the immunogenic nature of LNs as a pre-metastatic niche, TIL-B populations within the TME, and the presence of TLS will help to decipher local and widespread TIL-Bs responses and their influence on cancer progression to the lymphatics. Therapies that enhance TIL-Bs responses in the LN GC and/or in GC-like structures in the TME are thus emerging management strategies for breast and other cancer patients.

Circulating and intrahepatic antiviral B cells are defective in hepatitis B.

B cells are increasingly recognized as playing an important role in the ongoing control of hepatitis B virus (HBV). The development of antibodies against the viral surface antigen (HBV surface antigen [HBsAgs]) constitutes the hallmark of resolution of acute infection and is a therapeutic goal for functional cure of chronic HBV (CHB). We characterized B cells directly ex vivo from the blood and liver of patients with CHB to investigate constraints on their antiviral potential. Unexpectedly, we found that HBsAg-specific B cells persisted in the blood and liver of many patients with CHB and were enriched for T-bet, a signature of antiviral potential in B cells. However, purified, differentiated HBsAg-specific B cells from patients with CHB had defective antibody production, consistent with undetectable anti-HBs antibodies in vivo. HBsAg-specific and global B cells had an accumulation of CD21-CD27- atypical memory B cells (atMBC) with high expression of inhibitory receptors, including PD-1. These atMBC demonstrated altered signaling, homing, differentiation into antibody-producing cells, survival, and antiviral/proinflammatory cytokine production that could be partially rescued by PD-1 blockade. Analysis of B cells within healthy and HBV-infected livers implicated the combination of this tolerogenic niche and HBV infection in driving PD-1hiatMBC and impairing B cell immunity.