Peroxiredoxin 6 suppresses ferroptosis in lung endothelial cells.

Peroxiredoxin 6 (Prdx6) repairs peroxidized membranes by reducing oxidized phospholipids, and by replacing oxidized sn-2 fatty acyl groups through hydrolysis/reacylation by its phospholipase A2 (aiPLA2) and lysophosphatidylcholine acyltransferase activities. Prdx6 is highly expressed in the lung, and intact lungs and cells null for Prdx6 or with single-point mutations that inactivate either Prdx6-peroxidase or aiPLA2 activity alone exhibit decreased viability, increased lipid peroxidation, and incomplete repair when exposed to paraquat, hyperoxia, or organic peroxides. Ferroptosis is form of cell death driven by the accumulation of phospholipid hydroperoxides. We studied the role of Prdx6 as a ferroptosis suppressor in the lung. We first compared the expression Prdx6 and glutathione peroxidase 4 (GPx4) and visualized Prdx6 and GPx4 within the lung. Lung Prdx6 mRNA levels were five times higher than GPx4 levels. Both Prdx6 and GPx4 localized to epithelial and endothelial cells. Prdx6 knockout or knockdown sensitized lung endothelial cells to erastin-induced ferroptosis. Cells with genetic inactivation of either aiPLA2 or Prdx6-peroxidase were more sensitive to ferroptosis than WT cells, but less sensitive than KO cells. We then conducted RNA-seq analyses in Prdx6-depleted cells to further explore how the loss of Prdx6 sensitizes lung endothelial cells to ferroptosis. Prdx6 KD upregulated transcriptional signatures associated with selenoamino acid metabolism and mitochondrial function. Accordingly, Prdx6 deficiency blunted mitochondrial function and increased GPx4 abundance whereas GPx4 KD had the opposite effect on Prdx6. Moreover, we detected Prdx6 and GPx4 interactions in intact cells, suggesting that both enzymes cooperate to suppress lipid peroxidation. Notably, Prdx6-depleted cells remained sensitive to erastin-induced ferroptosis despite the compensatory increase in GPx4. These results show that Prdx6 suppresses ferroptosis in lung endothelial cells and that both aiPLA2 and Prdx6-peroxidase contribute to this effect. These results also show that Prdx6 supports mitochondrial function and modulates several coordinated cytoprotective pathways in the pulmonary endothelium.

Exhausted intratumoral Vδ2- γδ T cells in human kidney cancer retain effector function.

Gamma delta (γδ) T cells reside within human tissues including tumors, but their function in mediating antitumor responses to immune checkpoint inhibition is unknown. Here we show that kidney cancers are infiltrated by Vδ2- γδ T cells, with equivalent representation of Vδ1+ and Vδ1- cells, that are distinct from γδ T cells found in normal human tissues. These tumor-resident Vδ2- T cells can express the transcriptional program of exhausted αß CD8+ T cells as well as canonical markers of terminal T-cell exhaustion including PD-1, TIGIT and TIM-3. Although Vδ2- γδ T cells have reduced IL-2 production, they retain expression of cytolytic effector molecules and co-stimulatory receptors such as 4-1BB. Exhausted Vδ2- γδ T cells are composed of three distinct populations that lack TCF7, are clonally expanded and express cytotoxic molecules and multiple Vδ2- T-cell receptors. Human tumor-derived Vδ2- γδ T cells maintain cytotoxic function and pro-inflammatory cytokine secretion in vitro. The transcriptional program of Vδ2- T cells in pretreatment tumor biopsies was used to predict subsequent clinical responses to PD-1 blockade in patients with cancer. Thus, Vδ2- γδ T cells within the tumor microenvironment can contribute to antitumor efficacy.

Circulating monocytes associated with anti-PD-1 resistance in human biliary cancer induce T cell paralysis.

Suppressive myeloid cells can contribute to immunotherapy resistance, but their role in response to checkpoint inhibition (CPI) in anti-PD-1 refractory cancers, such as biliary tract cancer (BTC), remains elusive. We use multiplexed single-cell transcriptomic and epitope sequencing to profile greater than 200,000 peripheral blood mononuclear cells from advanced BTC patients (n = 9) and matched healthy donors (n = 8). Following anti-PD-1 treatment, CD14+ monocytes expressing high levels of immunosuppressive cytokines and chemotactic molecules (CD14CTX) increase in the circulation of patients with BTC tumors that are CPI resistant. CD14CTX can directly suppress CD4+ T cells and induce SOCS3 expression in CD4+ T cells, rendering them functionally unresponsive. The CD14CTX gene signature associates with worse survival in patients with BTC as well as in other anti-PD-1 refractory cancers. These results demonstrate that monocytes arising after anti-PD-1 treatment can induce T cell paralysis as a distinct mode of tumor-mediated immunosuppression leading to CPI resistance.

Tumor morphology and location associate with immune cell composition in pleomorphic sarcoma.

BACKGROUND: Soft-tissue sarcomas (STS) are a rare group of mesenchymal malignancies that account for approximately 1% of adult human cancer. Undifferentiated pleomorphic sarcoma (UPS) is one of the most common subtypes of adult STS. Clinical stratification of UPS patients has not evolved for decades and continues to rely on tumor-centric metrics including tumor size and depth. Our understanding of how the tumor microenvironment correlates to these clinicopathologic parameters remains limited. METHODS: Here, we performed single-cell flow cytometric immune-based profiling of 15 freshly resected UPS tumors and integrated this analysis with clinical, histopathologic, and outcomes data using both a prospective and retrospective cohort of UPS patients. RESULTS: We uncovered a correlation between physiologic and anatomic properties of UPS tumors and the composition of immune cells in the tumor microenvironment. Specifically, we identified an inverse correlation between tumor-infiltrating CD8 + T cells and UPS tumor size; and a positive correlation between tumor-infiltrating CD8 + T cells and overall survival. Moreover, we demonstrate an association between anatomical location (deep or superficial) and frequency of CD4 + PD1hi infiltrating T cells in UPS tumors. CONCLUSIONS: Our study provides an immune-based analysis of the tumor microenvironment in UPS patients and describes the different composition of tumor infiltrating lymphocytes based on size and tumor depth.