Tumor-B-Cell Interactions Promote Isotype Switching to an Immunosuppressive IgG4 Antibody Response Through Upregulation of IL-10 in Triple Negative Breast Cancers

Abstract

Background: Triple negative breast cancer (TNBC) is an aggressive breast cancer for which there is currently no targeted therapy. Tumor-infiltrating B-cells (TIB) have been observed in tumor tissues of TNBC patients, but their functional role is unclear. IgG4 is one of four antibody subclasses of IgG expressed and secreted by B cells. Unlike other IgG isotypes, IgG4 has an immunosuppressive function and is induced by Th2-type cytokines. In cancers such as melanoma, IgG4 has been linked with advanced disease and poor patient survival. Therefore, we sought to determine the role of IgG4 in TNBC. Methods: We performed co-culture assays to examine expression of Th2 cytokines by TNBC cells with and without the presence of B cells. We also performed in vitro class switching experiments with peripheral B cells with and without co-culture with TNBC cells in the presence or absence of an IL-10 blocking antibody. We examined expression of CD20 + TIB, IgG4 and Th2 cytokines by immunohistochemistry in 152 TNBC samples. Statistical analysis was done using Log-Rank and Cox-proportional hazards tests. Results: Our findings indicate that B cells interact with TNBC to drive chronic inflammatory responses through increased expression of inflammatory cytokines including the TH2 cytokines IL-4 and IL-10. In vitro class switching studies show that interactions between TNBC cell lines and B cells drive isotype switching to the IgG4 isotype in an IL-10 dependent manner. In patient tissues, expression of IgG4 correlates with CD20 and tumor expression of IL-10. Both IgG4 and tumor IL-10 are associated to shorter recurrence free survival (RFS) and overall survival (OS) in TNBC. In a multi-variant analysis, IL-10 was associated with poor outcomes indicating that tumor IL-10 may drive immune escape. Conclusions: These findings indicate that interactions between TIB and TNBC results in activation of chronic inflammatory signals that suppress antibody driven immune responses

Both COVID-19 infection and vaccination induce high-affinity cross-clade responses to SARS-CoV-2 variants

“…Our observation was corroborated by repeating the measurements using the same IgG4-specific secondary antibody as well as by using the same clone from an alternative source (including differences in concentrations and storage buffers), which makes us confident that the results presented are valid. Moreover, we presented one of the most comprehensive cross-validations to date of all secondary antibodies -widely used in research on SARS-CoV-2 (Crowley et al, 2021;Fraley et al, 2021;Kim et al, 2021;Lee et al, 2021;Newell et al, 2021;Phelan et al, 2021;Pullen et al, 2021;Yates et al, 2021) and in other fields (Minassian et al, 2021;Sanchez Vargas et al, 2021;Tschismarov et al, 2021;Jennewein et al, 2022;Toney et al, 2022) -employed in this study and have confirmed their specificity to their target. We have noted that the sensitivity of the anti-human IgG1 antibody used can benefit from a higher concentration; however, it would come at a cost of compromising its specificity as the antibody shows cross-reactivity to other IgG subtypes at slightly increased concentrations, therefore, presenting a less favorable sensitivity-to-specificity profile than the other secondary antibodies.…”