This study identifies mechanisms mediating responses to immune checkpoint inhibitors using mouse models of triple-negative breast cancer. By creating new mammary tumor models, we find that tumor mutation burden and specific immune cells are associated with response. Further, we developed a rich resource of single-cell RNA-seq and bulk mRNA-seq data of immunotherapy-treated and non-treated tumors from sensitive and resistant murine models. Using this, we uncover that immune checkpoint therapy induces T follicular helper cell activation of B cells to facilitate the anti-tumor response in these models. We also show that B cell activation of T cells and the generation of antibody are key to immunotherapy response and propose a new biomarker for immune checkpoint therapy. In total, this work presents resources of new preclinical models of breast cancer with large mRNA-seq and single-cell RNA-seq datasets annotated for sensitivity to therapy and uncovers new components of response to immune checkpoint inhibitors.

Although the transcription factors IRF-3 and IRF-7 are considered master regulators of type I interferon (IFN) induction and IFN stimulated gene (ISG) expression, Irf3−/−×Irf7−/− double knockout (DKO) myeloid dendritic cells (mDC) produce relatively normal levels of IFN-β after viral infection. We generated Irf3−/−×Irf5−/−×Irf7−/− triple knockout (TKO) mice to test whether IRF-5 was the source of the residual induction of IFN-β and ISGs in mDCs. In pathogenesis studies with two unrelated positive-sense RNA viruses (West Nile virus (WNV) and murine norovirus), TKO mice succumbed at rates greater than DKO mice and equal to or approaching those of mice lacking the type I IFN receptor (Ifnar−/−). In ex vivo studies, after WNV infection or exposure to Toll-like receptor agonists, TKO mDCs failed to produce IFN-β or express ISGs. In contrast, this response was sustained in TKO macrophages following WNV infection. To define IRF-regulated gene signatures, we performed microarray analysis on WNV-infected mDC from wild type (WT), DKO, TKO, or Ifnar−/− mice, as well as from mice lacking the RIG-I like receptor adaptor protein MAVS. Whereas the gene induction pattern in DKO mDC was similar to WT cells, remarkably, almost no ISG induction was detected in TKO or Mavs−/− mDC. The relative equivalence of TKO and Mavs−/− responses suggested that MAVS dominantly regulates ISG induction in mDC. Moreover, we showed that MAVS-dependent induction of ISGs can occur through an IRF-5-dependent yet IRF-3 and IRF-7-independent pathway. Our results establish IRF-3, -5, and -7 as the key transcription factors responsible for mediating the type I IFN and ISG response in mDC during WNV infection and suggest a novel signaling link between MAVS and IRF-5.

Although interferon-λ [also known as type III interferon or interleukin-28 (IL-28)/IL-29] restricts infection by several viruses, its inhibitory mechanism has remained uncertain. We used recombinant interferon-λ and mice lacking the interferon-λ receptor (IFNLR1) to evaluate the effect of interferon-λ on infection with West Nile virus, an encephalitic flavivirus. Cell culture studies in mouse keratinocytes and dendritic cells showed no direct antiviral effect of exogenous interferon-λ, even though expression of interferon-stimulated genes was induced. We observed no differences in West Nile virus burden between wild-type and Ifnlr1−/− mice in the draining lymph nodes, spleen, or blood. We detected increased West Nile virus infection in the brain and spinal cord of Ifnlr1−/− mice, yet this was not associated with a direct antiviral effect in mouse neurons. Instead, we observed an increase in blood-brain barrier permeability in Ifnlr1−/− mice. Treatment of mice with pegylated interferon-λ2 resulted in decreased blood-brain barrier permeability, reduced West Nile virus infection in the brain without affecting viremia, and improved survival against lethal virus challenge. An in vitro model of the blood-brain barrier showed that interferon-λ signaling in mouse brain microvascular endothelial cells increased transendothelial electrical resistance, decreased virus movement across the barrier, and modulated tight junction protein localization in a protein synthesis– and signal transducer and activator of transcription 1 (STAT1)–independent manner. Our data establish an indirect antiviral function of interferon-λ in which noncanonical signaling through IFNLR1 tightens the blood-brain barrier and restricts viral neuroinvasion and pathogenesis.

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Memory CD4 T cells in tissues fulfill numerous functions that are critical for local immune homeostasis and protection against pathogens. Previous studies have highlighted the phenotypic and functional heterogeneity of circulating and tissue-resident memory CD4 T cells across different human tissues such as skin, lung, liver and colon. Comparatively little is known in regard to memory CD4 T cells across tissues of the female reproductive tract (FRT). We examined CD4 T cells in donor-matched vaginal, ecto- and endo-cervical tissues, which differ in mucosal structure and exposure to external environmental stimuli. We hypothesized that this could be reflected by tissue-specific differences in the memory CD4 T cell compartment. We found differences in CD4 subset distribution across these tissues. Specifically, CD69 + CD103 + CD4 T cells were significantly more abundant in vaginal than cervical tissues. In contrast, the transcriptional profiles of CD4 subsets were fairly conserved across FRT tissues. CD69 + CD103 + CD4 T cells showed a T H 17 bias independent of tissue niche. Our data suggest that FRT tissues affect T cell subset distribution but have limited effects on the transcriptome of each subset. We discuss the implications for barrier immunity in the FRT.

Unique circulating regulatory T cell phenotypes distinguish hospitalized patients with SARS-CoV-2.