All-trans retinoic acid induces durable tumor immunity in IDH-mutant gliomas by rescuing transcriptional repression of the CRBP1-retinoic acid axis.

Diffuse gliomas are epigenetically dysregulated, immunologically cold, and fatal tumors characterized by mutations in isocitrate dehydrogenase (IDH). Although IDH mutations yield a uniquely immunosuppressive tumor microenvironment, the regulatory mechanisms that drive the immune landscape of IDH mutant (IDHm) gliomas remain unknown. Here, we reveal that transcriptional repression of retinoic acid (RA) pathway signaling impairs both innate and adaptive immune surveillance in IDHm glioma through epigenetic silencing of retinol binding protein 1 (RBP1) and induces a profound anti-inflammatory landscape marked by loss of inflammatory cell states and infiltration of suppressive myeloid phenotypes. Restorative retinoic acid therapy in murine glioma models promotes clonal CD4 + T cell expansion and induces tumor regression in IDHm, but not IDH wildtype (IDHwt), gliomas. Our findings provide a mechanistic rationale for RA immunotherapy in IDHm glioma and is the basis for an ongoing investigator-initiated, single-center clinical trial investigating all-trans retinoic acid (ATRA) in recurrent IDHm human subjects.

Integrated BATF transcriptional network regulates suppressive intratumoral regulatory T cells.

Human regulatory T cells (Tregs) are crucial regulators of tissue repair, autoimmune diseases, and cancer. However, it is challenging to inhibit the suppressive function of Tregs for cancer therapy without affecting immune homeostasis. Identifying pathways that may distinguish tumor-restricted Tregs is important, yet the transcriptional programs that control intratumoral Treg gene expression, and that are distinct from Tregs in healthy tissues, remain largely unknown. We profiled single-cell transcriptomes of CD4+ T cells in tumors and peripheral blood from patients with head and neck squamous cell carcinomas (HNSCC) and those in nontumor tonsil tissues and peripheral blood from healthy donors. We identified a subpopulation of activated Tregs expressing multiple tumor necrosis factor receptor (TNFR) genes (TNFR+ Tregs) that is highly enriched in the tumor microenvironment (TME) compared with nontumor tissue and the periphery. TNFR+ Tregs are associated with worse prognosis in HNSCC and across multiple solid tumor types. Mechanistically, the transcription factor BATF is a central component of a gene regulatory network that governs key aspects of TNFR+ Tregs. CRISPR-Cas9-mediated BATF knockout in human activated Tregs in conjunction with bulk RNA sequencing, immunophenotyping, and in vitro functional assays corroborated the central role of BATF in limiting excessive activation and promoting the survival of human activated Tregs. Last, we identified a suite of surface molecules reflective of the BATF-driven transcriptional network on intratumoral Tregs in patients with HNSCC. These findings uncover a primary transcriptional regulator of highly suppressive intratumoral Tregs, highlighting potential opportunities for therapeutic intervention in cancer without affecting immune homeostasis.

Immune landscape in invasive ductal and lobular breast cancer reveals a divergent macrophage-driven microenvironment.

T cell-centric immunotherapies have shown modest clinical benefit thus far for estrogen receptor-positive (ER+) breast cancer. Despite accounting for 70% of all breast cancers, relatively little is known about the immunobiology of ER+ breast cancer in women with invasive ductal carcinoma (IDC) and invasive lobular carcinoma (ILC). To investigate this, we performed phenotypic, transcriptional and functional analyses for a cohort of treatment-naive IDC (n = 94) and ILC (n = 87) tumors. We show that macrophages, and not T cells, are the predominant immune cells infiltrating the tumor bed and the most transcriptionally diverse cell subset between IDC and ILC. Analysis of cellular neighborhoods revealed an interplay between macrophages and T cells associated with longer disease-free survival in IDC but not ILC. Our datasets provide a rich resource for further interrogation into immune cell dynamics in ER+ IDC and ILC and highlight macrophages as a potential target for ER+ breast cancer.

Autoreactive CD8+ T cells are restrained by an exhaustion-like program that is maintained by LAG3.

Impaired chronic viral and tumor clearance has been attributed to CD8+ T cell exhaustion, a differentiation state in which T cells have reduced and altered effector function that can be partially reversed upon blockade of inhibitory receptors. The role of the exhaustion program and transcriptional networks that control CD8+ T cell function and fate in autoimmunity is not clear. Here we show that intra-islet CD8+ T cells phenotypically, transcriptionally, epigenetically and metabolically possess features of canonically exhausted T cells, yet maintain important differences. This 'restrained' phenotype can be perturbed and disease accelerated by CD8+ T cell-restricted deletion of the inhibitory receptor lymphocyte activating gene 3 (LAG3). Mechanistically, LAG3-deficient CD8+ T cells have enhanced effector-like functions, trafficking to the islets, and have a diminished exhausted phenotype, highlighting a physiological role for an exhaustion program in limiting autoimmunity and implicating LAG3 as a target for autoimmune therapy.

People critically ill with COVID-19 exhibit peripheral immune profiles predictive of mortality and reflective of SARS-CoV-2 lung viral burden.

Despite extensive analyses, there remains an urgent need to delineate immune cell states that contribute to mortality in people critically ill with COVID-19. Here, we present high-dimensional profiling of blood and respiratory samples from people with severe COVID-19 to examine the association between cell-linked molecular features and mortality outcomes. Peripheral transcriptional profiles by single-cell RNA sequencing (RNA-seq)-based deconvolution of immune states are associated with COVID-19 mortality. Further, persistently high levels of an interferon signaling module in monocytes over time lead to subsequent concerted upregulation of inflammatory cytokines. SARS-CoV-2-infected myeloid cells in the lower respiratory tract upregulate CXCL10, leading to a higher risk of death. Our analysis suggests a pivotal role for viral-infected myeloid cells and protracted interferon signaling in severe COVID-19.

Bifurcated monocyte states are predictive of mortality in severe COVID-19.

Coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2 infection presents with varied clinical manifestations1, ranging from mild symptoms to acute respiratory distress syndrome (ARDS) with high mortality2,3. Despite extensive analyses, there remains an urgent need to delineate immune cell states that contribute to mortality in severe COVID-19. We performed high-dimensional cellular and molecular profiling of blood and respiratory samples from critically ill COVID-19 patients to define immune cell genomic states that are predictive of outcome in severe COVID-19 disease. Critically ill patients admitted to the intensive care unit (ICU) manifested increased frequencies of inflammatory monocytes and plasmablasts that were also associated with ARDS not due to COVID-19. Single-cell RNAseq (scRNAseq)-based deconvolution of genomic states of peripheral immune cells revealed distinct gene modules that were associated with COVID-19 outcome. Notably, monocytes exhibited bifurcated genomic states, with expression of a cytokine gene module exemplified by CCL4 (MIP-1ß) associated with survival and an interferon signaling module associated with death. These gene modules were correlated with higher levels of MIP-1ß and CXCL10 levels in plasma, respectively. Monocytes expressing genes reflective of these divergent modules were also detectable in endotracheal aspirates. Machine learning algorithms identified the distinctive monocyte modules as part of a multivariate peripheral immune system state that was predictive of COVID-19 mortality. Follow-up analysis of the monocyte modules on ICU day 5 was consistent with bifurcated states that correlated with distinct inflammatory cytokines. Our data suggests a pivotal role for monocytes and their specific inflammatory genomic states in contributing to mortality in life-threatening COVID-19 disease and may facilitate discovery of new diagnostics and therapeutics.

COVID-19 versus Non-COVID-19 Acute Respiratory Distress Syndrome: Comparison of Demographics, Physiologic Parameters, Inflammatory Biomarkers, and Clinical Outcomes.

Rationale: There is an urgent need for improved understanding of the mechanisms and clinical characteristics of acute respiratory distress syndrome (ARDS) due to coronavirus disease (COVID-19).Objectives: To compare key demographic and physiologic parameters, biomarkers, and clinical outcomes of COVID-19 ARDS and ARDS secondary to direct lung injury from other etiologies of pneumonia.Methods: We enrolled 27 patients with COVID-19 ARDS in a prospective, observational cohort study and compared them with a historical, pre-COVID-19 cohort of patients with viral ARDS (n = 14), bacterial ARDS (n = 21), and ARDS due to culture-negative pneumonia (n = 30). We recorded clinical demographics; measured respiratory mechanical parameters; collected serial peripheral blood specimens for measurement of plasma interleukin (IL)-6, IL-8, and IL-10; and followed patients prospectively for patient-centered outcomes. We conducted between-group comparisons with nonparametric tests and analyzed time-to-event outcomes with Kaplan-Meier and Cox proportional hazards models.Results: Patients with COVID-19 ARDS had higher body mass index and were more likely to be Black, or residents of skilled nursing facilities, compared with those with non-COVID-19 ARDS (P < 0.05). Patients with COVID-19 had lower delivered minute ventilation compared with bacterial and culture-negative ARDS (post hoc P < 0.01) but not compared with viral ARDS. We found no differences in static compliance, hypoxemic indices, or carbon dioxide clearance between groups. Patients with COVID-19 had lower IL-6 levels compared with bacterial and culture-negative ARDS at early time points after intubation but no differences in IL-6 levels compared with viral ARDS. Patients with COVID-19 had longer duration of mechanical ventilation but similar 60-day mortality in both unadjusted and adjusted analyses.Conclusions: COVID-19 ARDS bears several similarities to viral ARDS but demonstrates lower minute ventilation and lower systemic levels of IL-6 compared with bacterial and culture-negative ARDS. COVID-19 ARDS was associated with longer dependence on mechanical ventilation compared with non-COVID-19 ARDS. Such detectable differences of COVID-19 do not merit deviation from evidence-based management of ARDS but suggest priorities for clinical research to better characterize and treat this new clinical entity.

Intractable Coronavirus Disease 2019 (COVID-19) and Prolonged Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Replication in a Chimeric Antigen Receptor-Modified T-Cell Therapy Recipient: A Case Study.

A chimeric antigen receptor-modified T-cell therapy recipient developed severe coronavirus disease 2019, intractable RNAemia, and viral replication lasting >2 months. Premortem endotracheal aspirate contained >2 × 1010 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA copies/mL and infectious virus. Deep sequencing revealed multiple sequence variants consistent with intrahost virus evolution. SARS-CoV-2 humoral and cell-mediated immunity were minimal. Prolonged transmission from immunosuppressed patients is possible.