Tumor-infiltrating regulatory T lymphocytes (Treg) can suppress effector T cells specific for tumor antigens. Deeper molecular definitions of tumor-infiltrating-lymphocytes could thus offer therapeutic opportunities. Transcriptomes of T helper 1 (Th1), Th17, and Treg cells infiltrating colorectal or non-small-cell lung cancers were compared to transcriptomes of the same subsets from normal tissues and validated at the single-cell level. We found that tumor-infiltrating Treg cells were highly suppressive, upregulated several immune-checkpoints, and expressed on the cell surfaces specific signature molecules such as interleukin-1 receptor 2 (IL1R2), programmed death (PD)-1 Ligand1, PD-1 Ligand2, and CCR8 chemokine, which were not previously described on Treg cells. Remarkably, high expression in whole-tumor samples of Treg cell signature genes, such as LAYN, MAGEH1, or CCR8, correlated with poor prognosis. Our findings provide insights into the molecular identity and functions of human tumor-infiltrating Treg cells and define potential targets for tumor immunotherapy.

Lung adenocarcinoma (LUAD) is the most common histologic subtype of lung cancer and has a high risk of distant metastasis at every disease stage. We aimed to characterize the genomic landscape of LUAD and identify mutation signatures associated with tumor progression. We performed an integrative genomic analysis, incorporating whole exome sequencing (WES), determination of DNA copy number and DNA methylation, and transcriptome sequencing for 101 LUAD samples from the Environment And Genetics in Lung cancer Etiology (EAGLE) study. We detected driver genes by testing whether the nonsynonymous mutation rate was significantly higher than the background mutation rate and replicated our findings in public datasets with 724 samples. We performed subclonality analysis for mutations based on mutant allele data and copy number alteration data. We also tested the association between mutation signatures and clinical outcomes, including distant metastasis, survival, and tumor grade. We identified and replicated two novel candidate driver genes, POU class 4 homeobox 2 (POU4F2) (mutated in 9 [8.9%] samples) and ZKSCAN1 (mutated in 6 [5.9%] samples), and characterized their major deleterious mutations. ZKSCAN1 was part of a mutually exclusive gene set that included the RTK/RAS/RAF pathway genes BRAF, EGFR, KRAS, MET, and NF1, indicating an important driver role for this gene. Moreover, we observed strong associations between methylation in specific genomic regions and somatic mutation patterns. In the tumor evolution analysis, four driver genes had a significantly lower fraction of subclonal mutations (FSM), including TP53 (p = 0.007), KEAP1 (p = 0.012), STK11 (p = 0.0076), and EGFR (p = 0.0078), suggesting a tumor initiation role for these genes. Subclonal mutations were significantly enriched in APOBEC-related signatures (p < 2.5×10⁻⁵⁰). The total number of somatic mutations (p = 0.0039) and the fraction of transitions (p = 5.5×10⁻⁴) were associated with increased risk of distant metastasis. Our study’s limitations include a small number of LUAD patients for subgroup analyses and a single-sample design for investigation of subclonality. These data provide a genomic characterization of LUAD pathogenesis and progression. The distinct clonal and subclonal mutation signatures suggest possible diverse carcinogenesis pathways for endogenous and exogenous exposures, and may serve as a foundation for more effective treatments for this lethal disease. LUAD’s high heterogeneity emphasizes the need to further study this tumor type and to associate genomic findings with clinical outcomes.

Mitochondria must buffer the risk of proteotoxic stress to preserve bioenergetics, but the role of these mechanisms in disease is poorly understood. Using a proteomics screen, we now show that the mitochondrial unfoldase-peptidase complex ClpXP associates with the oncoprotein survivin and the respiratory chain Complex II subunit succinate dehydrogenase B (SDHB) in mitochondria of tumor cells. Knockdown of ClpXP subunits ClpP or ClpX induces the accumulation of misfolded SDHB, impairing oxidative phosphorylation and ATP production while activating “stress” signals of 5′ adenosine monophosphate-activated protein kinase (AMPK) phosphorylation and autophagy. Deregulated mitochondrial respiration induced by ClpXP targeting causes oxidative stress, which in turn reduces tumor cell proliferation, suppresses cell motility, and abolishes metastatic dissemination in vivo. ClpP is universally overexpressed in primary and metastatic human cancer, correlating with shortened patient survival. Therefore, tumors exploit ClpXP-directed proteostasis to maintain mitochondrial bioenergetics, buffer oxidative stress, and enable metastatic competence. This pathway may provide a “drugable” therapeutic target in cancer. As the powerhouse of the cell and a pivotal hub for oxidative stress, mitochondria must tightly control the state of the proteins that they contain, quickly eliminating misfolded, aggregated, or otherwise damaged proteins. Here, we show that tumor mitochondria manage their set of proteins by assembling an integrated network of protein homeostasis, or proteostasis, that controls both the folding and degradation of proteins. This protein complex is formed by the unfoldase-peptidase ClpXP, survivin, and the Hsp90-like chaperone TRAP-1, and regulates the function of the oxidative phosphorylation Complex II subunit succinate dehydrogenase B (SDHB). We find that interference with this process impairs energy production, promotes oxidative stress, and shuts down critical downstream signals important for tumor cell proliferation, invasion, and metastatic dissemination in vivo. Our results suggest that the mitochondrial proteostasis network may offer therapeutic opportunities in advanced disease.

A large number of transplantation centres consider extracorporeal membrane oxygenation as an inappropriate option for bridging critical patients to lung transplantation. Technical improvements such as the introduction of a polymethylpentene membrane, new centrifugal pumps and heparin-coated circuits have led to a safer application of extracorporeal membrane oxygenation, and an increasing number of centres are reporting their positive experiences. The aim of this study was to review our practice in bridging critical candidates to lung transplantation with extracorporeal membrane oxygenation, by comparing patients with invasive mechanical ventilation with patients with spontaneous breathing. The records of candidates for lung transplantation treated with extracorporeal membrane oxygenation have been revised. From February 2008 to 2012, 11 patients who experienced an abrupt worsening of their respiratory conditions were treated with extracorporeal membrane oxygenation; mean age: 33.9 ± 13.2 years, male/female ratio: 5/6, 6 patients were affected by cystic fibrosis, 2 had chronic rejection after transplantation, 2 had pulmonary fibrosis and 1 had systemic sclerosis. Seven patients were awake, while 4 patients received invasive mechanical ventilation. The sequential organ failure assessment score significantly increased during bridging time and this increase was significantly higher in the intubated patients. All the patients had bilateral lung transplantation. Spontaneously breathing patients showed a tendency to require a shorter duration of invasive mechanical ventilation, intensive care unit stay and hospital stay after transplantation. One-year survival rate was 85.7% in patients with spontaneous breathing vs 50% in patients with invasive mechanical ventilation. Extracorporeal membrane oxygenation in spontaneously breathing patients is a feasible, effective and safe bridge to lung transplantation.

MiR-494-3p is a novel tumor driver of lung carcinogenesis

This paper describes the initial clinical experience of ex vivo lung perfusion (EVLP) at the Fondazione Ca' Granda in Milan between January 2011 and May 2013. EVLP was considered if donor PaO2 /FiO2 was below 300 mmHg or if lung function was doubtful. Donors with massive lung contusion, aspiration, purulent secretions, pneumonia, or sepsis were excluded. EVLP was run with a low-flow, open atrium and low hematocrit technique. Thirty-five lung transplants from brain death donors were performed, seven of which after EVLP. EVLP donors were older (54 ± 9 years vs. 40 ± 15 years, EVLP versus Standard, P < 0.05), had lower PaO2 /FiO2 (264 ± 78 mmHg vs. 453 ± 119 mmHg, P < 0.05), and more chest X-ray abnormalities (P < 0.05). EVLP recipients were more often admitted to intensive care unit as urgent cases (57% vs. 18%, P = 0.05); lung allocation score at transplantation was higher (79 [40-84] vs. 39 [36-46], P < 0.05). After transplantation, primary graft dysfunction (PGD72 grade 3, 32% vs. 28%, EVLP versus Standard, P = 1), mortality at 30 days (0% vs. 0%, P = 1), and overall survival (71% vs. 86%, EVLP versus Standard P = 0.27) were not different between groups. EVLP enabled a 20% increase in available donor organs and resulted in successful transplants with lungs that would have otherwise been rejected (ClinicalTrials.gov number: NCT01967953).