Myocardial infarction augments sleep to limit cardiac inflammation and damage.

Sleep is integral to cardiovascular health1,2. Yet, the circuits that connect cardiovascular pathology and sleep are incompletely understood. It remains unclear whether cardiac injury influences sleep and whether sleep-mediated neural outputs contribute to heart healing and inflammation. Here we report that in humans and mice, monocytes are actively recruited to the brain after myocardial infarction (MI) to augment sleep, which suppresses sympathetic outflow to the heart, limiting inflammation and promoting healing. After MI, microglia rapidly recruit circulating monocytes to the brain's thalamic lateral posterior nucleus (LPN) via the choroid plexus, where they are reprogrammed to generate tumour necrosis factor (TNF). In the thalamic LPN, monocytic TNF engages Tnfrsf1a-expressing glutamatergic neurons to increase slow wave sleep pressure and abundance. Disrupting sleep after MI worsens cardiac function, decreases heart rate variability and causes spontaneous ventricular tachycardia. After MI, disrupting or curtailing sleep by manipulating glutamatergic TNF signalling in the thalamic LPN increases cardiac sympathetic input which signals through the ß2-adrenergic receptor of macrophages to promote a chemotactic signature that increases monocyte influx. Poor sleep in the weeks following acute coronary syndrome increases susceptibility to secondary cardiovascular events and reduces the heart's functional recovery. In parallel, insufficient sleep in humans reprogrammes ß2-adrenergic receptor-expressing monocytes towards a chemotactic phenotype, enhancing their migratory capacity. Collectively, our data uncover cardiogenic regulation of sleep after heart injury, which restricts cardiac sympathetic input, limiting inflammation and damage.

Hematopoietic aging promotes cancer by fueling IL-1⍺-driven emergency myelopoiesis.

Age is a major risk factor for cancer, but how aging impacts tumor control remains unclear. In this study, we establish that aging of the immune system, regardless of the age of the stroma and tumor, drives lung cancer progression. Hematopoietic aging enhances emergency myelopoiesis, resulting in the local accumulation of myeloid progenitor-like cells in lung tumors. These cells are a major source of interleukin (IL)-1⍺, which drives the enhanced myeloid response. The age-associated decline of DNA methyltransferase 3A enhances IL-1⍺ production, and disrupting IL-1 receptor 1 signaling early during tumor development normalized myelopoiesis and slowed the growth of lung, colonic, and pancreatic tumors. In human tumors, we identified an enrichment for IL-1⍺-expressing monocyte-derived macrophages linked to age, poorer survival, and recurrence, unraveling how aging promotes cancer and offering actionable therapeutic strategies.

Myeloid progenitor dysregulation fuels immunosuppressive macrophages in tumors.

Monocyte-derived macrophages (mo-macs) drive immunosuppression in the tumor microenvironment (TME) and tumor-enhanced myelopoiesis in the bone marrow (BM) fuels these populations. Here, we performed paired transcriptome and chromatin analysis over the continuum of BM myeloid progenitors, circulating monocytes, and tumor-infiltrating mo-macs in mice and in patients with lung cancer to identify myeloid progenitor programs that fuel pro-tumorigenic mo-macs. Analyzing chromatin accessibility and histone mark changes, we show that lung tumors prime accessibility for Nfe2l2 (NRF2) in BM myeloid progenitors as a cytoprotective response to oxidative stress. NRF2 activity is sustained and increased during monocyte differentiation into mo-macs in the lung TME to regulate oxidative stress, in turn promoting metabolic adaptation, resistance to cell death, and contributing to immunosuppressive phenotype. NRF2 genetic deletion and pharmacological inhibition significantly reduced mo-macs' survival and immunosuppression in the TME, enabling NK and T cell therapeutic antitumor immunity and synergizing with checkpoint blockade strategies. Altogether, our study identifies a targetable epigenetic node of myeloid progenitor dysregulation that sustains immunoregulatory mo-macs in the TME.

Single-Cell View of Tumor Microenvironment Gradients in Pleural Mesothelioma.

Immunotherapies have shown great promise in pleural mesothelioma (PM), yet most patients still do not achieve significant clinical response, highlighting the importance of improving the understanding of the tumor microenvironment (TME). Here, we utilized high-throughput, single-cell RNA sequencing (scRNA-seq) to de novo identify 54 expression programs and construct a comprehensive cellular catalog of the PM TME. We found four cancer-intrinsic programs associated with poor disease outcome and a novel fetal-like, endothelial cell population that likely responds to VEGF signaling and promotes angiogenesis. Across cellular compartments, we observe substantial difference in the TME associated with a cancer-intrinsic sarcomatoid signature, including enrichment in fetal-like endothelial cells, CXCL9+ macrophages, and cytotoxic, exhausted, and regulatory T cells, which we validated using imaging and bulk deconvolution analyses on independent cohorts. Finally, we show, both computationally and experimentally, that NKG2A:HLA-E interaction between NK and tumor cells represents an important new therapeutic axis in PM, especially for epithelioid cases. Significance: This manuscript presents the first single-cell RNA sequencing atlas of PM tumor microenvironment. Findings of translational relevance, validated experimentally and using independent bulk cohorts, include identification of gene programs predictive of survival, a fetal-like endothelial cell population, and NKG2A blockade as a promising new immunotherapeutic intervention in PM.

Immunophenotypic correlates of sustained MRD negativity in patients with multiple myeloma.

The role of the immune microenvironment in maintaining disease remission in patients with multiple myeloma (MM) is not well understood. In this study, we comprehensively profile the immune system in patients with newly diagnosed MM receiving continuous lenalidomide maintenance therapy with the aim of discovering correlates of long-term treatment response. Leveraging single-cell RNA sequencing and T cell receptor ß sequencing of the peripheral blood and CyTOF mass cytometry of the bone marrow, we longitudinally characterize the immune landscape in 23 patients before and one year after lenalidomide exposure. We compare patients achieving sustained minimal residual disease (MRD) negativity to patients who never achieved or were unable to maintain MRD negativity. We observe that the composition of the immune microenvironment in both the blood and the marrow varied substantially according to both MRD negative status and history of autologous stem cell transplant, supporting the hypothesis that the immune microenvironment influences the depth and duration of treatment response.

Intratumoral dendritic cell-CD4+ T helper cell niches enable CD8+ T cell differentiation following PD-1 blockade in hepatocellular carcinoma.

Despite no apparent defects in T cell priming and recruitment to tumors, a large subset of T cell rich tumors fail to respond to immune checkpoint blockade (ICB). We leveraged a neoadjuvant anti-PD-1 trial in patients with hepatocellular carcinoma (HCC), as well as additional samples collected from patients treated off-label, to explore correlates of response to ICB within T cell-rich tumors. We show that ICB response correlated with the clonal expansion of intratumoral CXCL13+CH25H+IL-21+PD-1+CD4+ T helper cells ("CXCL13+ TH") and Granzyme K+ PD-1+ effector-like CD8+ T cells, whereas terminally exhausted CD39hiTOXhiPD-1hiCD8+ T cells dominated in nonresponders. CD4+ and CD8+ T cell clones that expanded post-treatment were found in pretreatment biopsies. Notably, PD-1+TCF-1+ (Progenitor-exhausted) CD8+ T cells shared clones mainly with effector-like cells in responders or terminally exhausted cells in nonresponders, suggesting that local CD8+ T cell differentiation occurs upon ICB. We found that these Progenitor CD8+ T cells interact with CXCL13+ TH within cellular triads around dendritic cells enriched in maturation and regulatory molecules, or "mregDC". These results suggest that discrete intratumoral niches that include mregDC and CXCL13+ TH control the differentiation of tumor-specific Progenitor exhasuted CD8+ T cells following ICB.

TREM2 macrophages drive NK cell paucity and dysfunction in lung cancer.

Natural killer (NK) cells are commonly reduced in human tumors, enabling many to evade surveillance. Here, we sought to identify cues that alter NK cell activity in tumors. We found that, in human lung cancer, the presence of NK cells inversely correlated with that of monocyte-derived macrophages (mo-macs). In a murine model of lung adenocarcinoma, we show that engulfment of tumor debris by mo-macs triggers a pro-tumorigenic program governed by triggering receptor expressed on myeloid cells 2 (TREM2). Genetic deletion of Trem2 rescued NK cell accumulation and enabled an NK cell-mediated regression of lung tumors. TREM2+ mo-macs reduced NK cell activity by modulating interleukin (IL)-18/IL-18BP decoy interactions and IL-15 production. Notably, TREM2 blockade synergized with an NK cell-activating agent to further inhibit tumor growth. Altogether, our findings identify a new axis, in which TREM2+ mo-macs suppress NK cell accumulation and cytolytic activity. Dual targeting of macrophages and NK cells represents a new strategy to boost antitumor immunity.

Gut-associated lymphoid tissue attrition associates with response to anti-α4ß7 therapy in ulcerative colitis.

Targeting the α4ß7-MAdCAM-1 axis with vedolizumab (VDZ) is a front-line therapeutic paradigm in ulcerative colitis (UC). However, mechanism(s) of action (MOA) of VDZ remain relatively undefined. Here, we examined three distinct cohorts of patients with UC (n=83, n=60, and n=21), to determine the effect of VDZ on the mucosal and peripheral immune system. Transcriptomic studies with protein level validation were used to study drug MOA using conventional and transgenic murine models. We found a significant decrease in colonic and ileal naïve B and T cells and circulating gut-homing plasmablasts (ß7+) in VDZ-treated patients, pointing to gut-associated lymphoid tissue (GALT) targeting by VDZ. Murine Peyer's patches (PP) demonstrated a significant loss cellularity associated with reduction in follicular B cells, including a unique population of epithelium-associated B cells, following anti-α4ß7 antibody (mAb) administration. Photoconvertible (KikGR) mice unequivocally demonstrated impaired cellular entry into PPs in anti-α4ß7 mAb treated mice. In VDZ-treated, but not anti-tumor necrosis factor-treated UC patients, lymphoid aggregate size was significantly reduced in treatment responders compared to non-responders, with an independent validation cohort further confirming these data. GALT targeting represents a novel MOA of α4ß7-targeted therapies, with major implications for this therapeutic paradigm in UC, and for the development of new therapeutic strategies.

A shift in lung macrophage composition is associated with COVID-19 severity and recovery.

Although it has been more than 2 years since the start of the coronavirus disease 2019 (COVID-19) pandemic, COVID-19 continues to be a worldwide health crisis. Despite the development of preventive vaccines, therapies to treat COVID-19 and other inflammatory diseases remain a major unmet need in medicine. Our study sought to identify drivers of disease severity and mortality to develop tailored immunotherapy strategies to halt disease progression. We assembled the Mount Sinai COVID-19 Biobank, which was composed of almost 600 hospitalized patients followed longitudinally through the peak of the pandemic in 2020. Moderate disease and survival were associated with a stronger antigen presentation and effector T cell signature. In contrast, severe disease and death were associated with an altered antigen presentation signature, increased numbers of inflammatory immature myeloid cells, and extrafollicular activated B cells that have been previously associated with autoantibody formation. In severely ill patients with COVID-19, lung tissue-resident alveolar macrophages not only were drastically depleted but also had an altered antigen presentation signature, which coincided with an influx of inflammatory monocytes and monocyte-derived macrophages. In addition, we found that the size of the alveolar macrophage pool correlated with patient outcome and that alveolar macrophage numbers and functionality were restored to homeostasis in patients who recovered from COVID-19. These data suggest that local and systemic myeloid cell dysregulation are drivers of COVID-19 severity and modulation of alveolar macrophage numbers and activity in the lung may be a viable therapeutic strategy for the treatment of critical inflammatory lung diseases.

Comprehensive Characterization of the Multiple Myeloma Immune Microenvironment Using Integrated scRNA-seq, CyTOF, and CITE-seq Analysis.

As part of the Multiple Myeloma Research Foundation (MMRF) immune atlas pilot project, we compared immune cells of multiple myeloma bone marrow samples from 18 patients assessed by single-cell RNA sequencing (scRNA-seq), mass cytometry (CyTOF), and cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) to understand the concordance of measurements among single-cell techniques. Cell type abundances are relatively consistent across the three approaches, while variations are observed in T cells, macrophages, and monocytes. Concordance and correlation analysis of cell type marker gene expression across different modalities highlighted the importance of choosing cell type marker genes best suited to particular modalities. By integrating data from these three assays, we found International Staging System stage 3 patients exhibited decreased CD4+ T/CD8+ T cells ratio. Moreover, we observed upregulation of RAC2 and PSMB9, in natural killer cells of fast progressors compared with those of nonprogressors, as revealed by both scRNA-seq and CITE-seq RNA measurement. This detailed examination of the immune microenvironment in multiple myeloma using multiple single-cell technologies revealed markers associated with multiple myeloma rapid progression which will be further characterized by the full-scale immune atlas project. Significance: scRNA-seq, CyTOF, and CITE-seq are increasingly used for evaluating cellular heterogeneity. Understanding their concordances is of great interest. To date, this study is the most comprehensive examination of the measurement of the immune microenvironment in multiple myeloma using the three techniques. Moreover, we identified markers predicted to be significantly associated with multiple myeloma rapid progression.