The Effect of Positive Pressure Ventilation on Acute Kidney Injury in COVID-19 Patients with Acute Respiratory Distress Syndrome: An Observational Study.

INTRODUCTION: Acute kidney injury (AKI) is frequent in critically ill COVID-19 patients and is associated with a higher mortality risk. By increasing intrathoracic pressure, positive pressure ventilation (PPV) may reduce renal perfusion pressure by reducing venous return to the heart or by increasing renal venous congestion. This study's aim was to evaluate the association between AKI and haemodynamic and ventilatory parameters in COVID-19 patients with ARDS. METHODS: This is a single-centre retrospective observational study. Consecutive patients diagnosed with COVID-19 who met ARDS criteria and required invasive mechanical ventilation were enrolled. The relationship between respiratory and haemodynamic parameters influenced by PPV and AKI development was evaluated. AKI was defined according to KDIGO criteria. AKI recovery was evaluated a month after ICU admission and patients were classified as "recovered," if serum creatinine (sCr) value returned to baseline, or as having "acute kidney disease" (AKD), if criteria for AKI stage 1 or greater persisted. The 6-month all-cause mortality was collected. RESULTS: A total of 144 patients were included in the analysis. AKI occurred in 69 (48%) patients and 26 (18%) required renal replacement therapy. In a multivariate logistic regression analysis, sex, hypertension, cumulative dose of furosemide, fluid balance, and plateau pressure were independently associated with AKI. Mortality at 6 months was 50% in the AKI group and 32% in the non-AKI group (p = 0.03). Among 36 patients who developed AKI and were discharged alive from the hospital, 56% had a full renal recovery after a month, while 14%, 6%, and 14% were classified as having an AKD of stage 0, 2, and 3, respectively. CONCLUSIONS: In our cohort, AKI was independently associated with multiple variables, including high plateau pressure, suggesting a possible role of PPV on AKI development. Further studies are needed to clarify the role of mechanical ventilation on renal function.

Prognostic significance of tumor-associated macrophages: past, present and future.

Tumor tissues are populated by a multitude of macrophages, highly different in functional activity, localization and morphology. A clear contribution to disease progression has been shown in multiple cancer types, holding promise for the development of innovative macrophage-based prognostic tools. Current studies aimed at assessing the prognostic role of macrophages have documented the relevance of the macrophage population as a whole. However, dissecting the diversity of mononuclear phagocytes in tumor tissues has provided important information about the coexistence of distinct populations of macrophages with different prognostic significance. Here we summarize evidence of macrophage prognostic function in human cancer and focus on classical and modern strategies aimed at measuring macrophage features and deciphering their diversity. The wealth of new data generated will reshape our knowledge of macrophage complexity and hopefully foster the forthcoming development of these new metrics into prognostic tools as well as new therapeutic strategies.

Multiplexed Imaging Mass Cytometry Analysis in Preclinical Models of Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers. PDAC is characterized by a complex tumor microenvironment (TME), that plays a pivotal role in disease progression and resistance to therapy. Investigating the spatial distribution and interaction of TME cells with the tumor is the basis for understanding the mechanisms underlying disease progression and represents a current challenge in PDAC research. Imaging mass cytometry (IMC) is the major multiplex imaging technology for the spatial analysis of tumor heterogeneity. However, there is a dearth of reports of multiplexed IMC panels for different preclinical mouse models, including pancreatic cancer. We addressed this gap by utilizing two preclinical models of PDAC: the genetically engineered, bearing KRAS-TP53 mutations in pancreatic cells, and the orthotopic, and developed a 28-marker panel for single-cell IMC analysis to assess the abundance, distribution and phenotypes of cells involved in PDAC progression and their reciprocal functional interactions. Herein, we provide an unprecedented definition of the distribution of TME cells in PDAC and compare the diversity between transplanted and genetic disease models. The results obtained represent an important and customizable tool for unraveling the complexities of PDAC and deciphering the mechanisms behind therapy resistance.

The Impact of Molecular Subtyping on Pathological Staging of Pancreatic Cancer.

BACKGROUND: The long-term outcomes following surgical resection for pancreatic ductal adenocarcinoma (PDAC) remains poor, with only 20% of patients surviving 5 years after pancreatectomy. Patient selection for surgery remains suboptimal largely due to the absence of consideration of aggressive tumor biology. OBJECTIVE: The aim of this study was to evaluate traditional staging criteria for PDAC in the setting of molecular subtypes. METHODS: Clinicopathological data were obtained for 5 independent cohorts of consecutive unselected patients, totaling n = 1298, including n = 442 that underwent molecular subtyping. The main outcome measure was disease-specific survival following surgical resection for PDAC stratified according to the American Joint Commission for Cancer (TNM) staging criteria, margin status, and molecular subtype. RESULTS: TNM staging criteria and margin status confers prognostic value only in tumors with classical pancreatic subtype. Patients with tumors that are of squamous subtype, have a poor outcome irrespective of favorable traditional pathological staging [hazard ratio (HR) 1.54, 95% confidence interval (CI) 1.04-2.28, P = 0.032]. Margin status has no impact on survival in the squamous subtype (16.0 vs 12.1 months, P = 0.374). There were no differences in molecular subtype or gene expression of tumors with positive resection margin status. CONCLUSIONS: Aggressive tumor biology as measured by molecular subtype predicts poor outcome following pancreatectomy for PDAC and should be utilized to inform patient selection for surgery.

IL-10 and macrophages orchestrate gut homeostasis.

Macrophages in the gut originate from blood precursors and have distinct functional properties. In this issue of Immunity,Zigmond et al. (2014) and Shouval et al. (2014) report that interleukin-10 drives macrophages to express homeostatic tolerogenic functions, preventing colitis.

Heterogeneity of Colorectal Cancer Progression: Molecular Gas and Brakes.

The review begins with molecular genetics, which hit the field unveiling the involvement of oncogenes and tumor suppressor genes in the pathogenesis of colorectal cancer (CRC) and uncovering genetic predispositions. Then the notion of molecular phenotypes with different clinical behaviors was introduced and translated in the clinical arena, paving the way to next-generation sequencing that captured previously unrecognized heterogeneity. Among other molecular regulators of CRC progression, the extent of host immune response within the tumor micro-environment has a critical position. Translational sciences deeply investigated the field, accelerating the pace toward clinical transition, due to its strong association with outcomes. While the perturbation of gut homeostasis occurring in inflammatory bowel diseases can fuel carcinogenesis, micronutrients like vitamin D and calcium can act as brakes, and we discuss underlying molecular mechanisms. Among the components of gut microbiota, Fusobacterium nucleatum is over-represented in CRC, and may worsen patient outcome. However, any translational knowledge tracing the multifaceted evolution of CRC should be interpreted according to the prognostic and predictive frame of the TNM-staging system in a perspective of clinical actionability. Eventually, we examine challenges and promises of pharmacological interventions aimed to restrain disease progression at different disease stages.

The Fractalkine-Receptor Axis Improves Human Colorectal Cancer Prognosis by Limiting Tumor Metastatic Dissemination.

Human colorectal cancer (CRC) is a frequent neoplasia in Western countries, and its metastatic progression is a major cause of cancer-related death. In search of specific molecules upregulated in CRC, with possible clinical relevance, we performed a differential gene-profiling analysis in surgery-derived CRC samples and adjacent uninvolved intestinal mucosa. The chemokine CX3CL1 and its specific receptor CX3CR1 were significantly upregulated in tumors. Higher expression of CX3CL1 and CX3CR1 was confirmed by immunohistochemistry in 100 CRC tumor samples (stages I-III). Unexpectedly, high immune scores of CX3CL1 did not correlate with the density of tumor-infiltrating CD3(+) T cells or CD68(+) macrophages. Coexpression of ligand and receptor by tumor cells (axis-positive tumors) significantly associated with longer disease-free (p = 0.01) and disease-specific survival (p = 0.001). Conversely, axis-negative tumors (with low expression of both ligand and receptor) had increased risk of tumor relapse (p = 0.02), and increased likelihood of metachronous metastasis (p = 0.001), including after stage adjustment (p = 0.006). Transduction of CX3CL1 and CX3CR1 in CRC tumor cell lines induced cell aggregation that strongly inhibited in vitro migration in chemotaxis assays. In a mouse model of spleen-liver metastases, cancer dissemination to liver was dramatically reduced in CX3CL1-CX3CR1-expressing tumors, and ligand-receptor interaction was confirmed in cancer cells in vivo by fluorescence resonance energy transfer analysis. In conclusion, tumoral expression of the CX3CL1-CX3CR1 chemokine axis functions as a retention factor, increasing homotypic cell adhesion and limiting tumor spreading to metastatic sites. Lack or low levels of expression of CX3CL1-CX3CR1 by tumor cells identifies a group of CRC patients at increased risk of metastatic progression.

Dual prognostic significance of tumour-associated macrophages in human pancreatic adenocarcinoma treated or untreated with chemotherapy.

OBJECTIVE: Tumour-associated macrophages (TAMs) play key roles in tumour progression. Recent evidence suggests that TAMs critically modulate the efficacy of anticancer therapies, raising the prospect of their targeting in human cancer. DESIGN: In a large retrospective cohort study involving 110 patients with pancreatic ductal adenocarcinoma (PDAC), we assessed the density of CD68-TAM immune reactive area (%IRA) at the tumour-stroma interface and addressed their prognostic relevance in relation to postsurgical adjuvant chemotherapy (CTX). In vitro, we dissected the synergism of CTX and TAMs. RESULTS: In human PDAC, TAMs predominantly exhibited an immunoregulatory profile, characterised by expression of scavenger receptors (CD206, CD163) and production of interleukin 10 (IL-10). Surprisingly, while the density of TAMs associated to worse prognosis and distant metastasis, CTX restrained their protumour prognostic significance. High density of TAMs at the tumour-stroma interface positively dictated prognostic responsiveness to CTX independently of T-cell density. Accordingly, in vitro, gemcitabine-treated macrophages became tumoricidal, activating a cytotoxic gene expression programme, inhibiting their protumoural effect and switching to an antitumour phenotype. In patients with human PDAC, neoadjuvant CTX was associated to a decreased density of CD206(+) and IL-10(+) TAMs at the tumour-stroma interface. CONCLUSIONS: Overall, our data highlight TAMs as critical determinants of prognostic responsiveness to CTX and provide clinical and in vitro evidence that CTX overall directly re-educates TAMs to restrain tumour progression. These results suggest that the quantification of TAMs could be exploited to select patients more likely to respond to CTX and provide the basis for novel strategies aimed at re-educating macrophages in the context of CTX.

Correlation of metabolic information on FDG-PET with tissue expression of immune markers in patients with non-small cell lung cancer (NSCLC) who are candidates for upfront surgery.

PURPOSE: Eliciting antitumor T-cell response by targeting the PD-1/PD-L1 axis with checkpoint inhibitors has emerged as a novel therapeutic strategy in non-small cell lung cancer (NSCLC). The identification of predictors for sensitivity or resistance to these agents is, therefore, needed. Herein, we investigate the correlation of metabolic information on FDG-PET with tissue expression of immune-checkpoints and other markers of tumor-related immunity in resected NSCLC patients. MATERIALS AND METHODS: All patients referred to our institution for upfront surgical resection of NSCLC, who were investigated with FDG-PET prior to surgery, were consecutively included in the study. From January 2010 to May 2014, 55 patients (stage IA-IIIB; M:F = 42:13; mean age 68.9 years) were investigated. Sampled surgical tumor specimens were analyzed by immunohistochemistry (IHC) for CD68-TAMs (tumor-associated macrophages), CD8-TILs (tumor infiltrating lymphocytes), PD-1-TILs, and PD-L1 tumor expression. Immunoreactivity was evaluated, and scores were compared with imaging findings. FDG-PET images were analyzed to define semi-quantitative parameters: SUVmax and SUVmean. Metabolic information on FDG-PET was correlated with tissue markers expression and disease-free survival (DFS) considering a median follow-up of 16.2 months. RESULTS: Thirty-six adenocarcinomas (ADC), 18 squamous cell carcinomas (SCC), and one sarcomatoid carcinoma were analyzed. All tumors resulted positive at FDG-PET: median SUVmax 11.3 (range: 2.3-32.5) and SUVmean 6.4 (range: 1.5-13) both resulted significantly higher in SCC compared to other NSCLC histotypes (p = 0.007 and 0.048, respectively). IHC demonstrated a median immunoreactive surface covered by CD68-TAMs of 5.41 % (range: 0.84-14.01 %), CD8-TILs of 2.9 % (range: 0.11-11.92 %), PD-1 of 0.65 % (range: 0.02-5.87 %), and PD-L1 of 0.7 % (range: 0.03-10.29 %). We found a statistically significant correlation between SUVmax and SUVmean with the expression of CD8 TILs (rho = 0.31; p = 0.027) and PD-1 (rho = 0.33; p = 0.017 and rho = 0.36; p = 0.009, respectively). The other tissue markers correlated as follows: CD8 TILs and PD-1 (rho = 0.45; p = 0.001), CD8 TILs and PD-L1 (rho = 0.41; p = 0.003), CD68-TAMs and PD-L1 (rho = 0.30; p = 0.027), PD-1 and PD-L1 (rho = 0.26; p = 0.059). With respect to patients' outcome, SUVmax, SUVmean, and disease stage showed a statistically significant correlation with DFS (p = 0.002, 0.004, and <0.001, respectively). CONCLUSIONS: The present study shows a direct association between metabolic parameters on FDG-PET and the expression of tumor-related immunity markers, suggesting a potential role for FDG-PET to characterize the tumor microenvironment and select NSCLC patients candidate to checkpoint inhibitors.

Inflammation and prostate cancer: friends or foe?

INTRODUCTION: Prostate cancer is the most common non-cutaneous malignancy diagnosed in men. Moving from histological observations since a long time, it has been recognized that innate and adaptive immunity actively participates in the pathogenesis, surveillance, and progression of prostate cancer. MATERIALS AND METHODS: A PubMed and Web of Science databases search was performed for studies providing evidence on the roles of the innate and adaptive immunity during the development and progression of prostate cancer. CONCLUSIONS: There are growing evidences that chronic inflammation is involved in the regulation of cellular events in prostate carcinogenesis, including disruption of the immune response and regulation of the tumor microenvironment. This review discusses the role played by the innate and adaptive immune system in the local progression of prostate cancer, and the prognostic information that we can currently understand and exploit.

Presence of Twist1-positive neoplastic cells in the stroma of chromosome-unstable colorectal tumors.

BACKGROUND & AIMS: Cancer cells undergo an epithelial-to-mesenchymal transition (EMT) to become invasive, allowing tumors to progress. However, there is no direct evidence that human cancer cells undergo an EMT. In mouse cancer cells, up-regulation of transcription factor Twist1 was shown to promote an EMT. We searched the stroma of human colorectal tumor samples for TWIST1-positive cells with a mesenchymal phenotype and neoplastic genotype. METHODS: We measured the expression of TWIST1 in human colorectal cancer (CRC) cell lines and examined the effects of overexpression or knockdown in vitro and in mice. We used immunohistochemistry to measure levels of TWIST1 in 201 colorectal tumor samples. In 20 samples, immunostaining was combined with fluorescence in situ hybridization analyses. Levels of TWIST1 messenger RNA (mRNA) were measured in blood samples from 15 patients. RESULTS: TWIST1 was required to maintain the mesenchymal phenotype and invasiveness of the microsatellite-stable CoLo741 cells (which express endogenous TWIST1) and SW480 (expressing transgenic TWIST1). TWIST1 mRNA was not translated in CRC cells with microsatellite instability (HCT116). Syngenic TWIST1-positive colon carcinoma cells (CT26) that invaded tissues surrounding tumors acquired a mesenchymal phenotype. The presence of TWIST1-positive cells in the stroma of human colorectal tumors correlated with microsatellite stability (P = .05), stage IV cancer (P = .02), and disease-free survival time (P < .01). Trisomies of chromosome 7 and/or chromosome 20 were detected in 17 of 20 colorectal tumor samples, each of which contained TWIST1-positive cells with matching chromosomal gains in the tumor stroma (86 of 776 counted cells; 11.1%). No trisomy was observed in TWIST1-negative stromal cells (0 of 1249 cells; P < .001). Levels of TWIST1 mRNA were significantly higher in blood samples from patients with CRC than controls. CONCLUSIONS: The stroma of human colorectal tumors contains TWIST1-positive cancer cells with mesenchymal phenotypes. Patients with CRC have higher levels of TWIST1 mRNA than healthy individuals.

Macrophage diversity in cancer dissemination and metastasis.

Invasion and metastasis are hallmarks of cancer. In addition to the well-recognized hematogenous and lymphatic pathways of metastasis, cancer cell dissemination can occur via the transcoelomic and perineural routes, which are typical of ovarian and pancreatic cancer, respectively. Macrophages are a universal major component of the tumor microenvironment and, in established tumors, promote growth and dissemination to secondary sites. Here, we review the role of tumor-associated macrophages (TAMs) in cancer cell dissemination and metastasis, emphasizing the diversity of myeloid cells in different tissue contexts (lungs, liver, brain, bone, peritoneal cavity, nerves). The generally used models of lung metastasis fail to capture the diversity of pathways and tissue microenvironments. A better understanding of TAM diversity in different tissue contexts may pave the way for tailored diagnostic and therapeutic approaches.

Integrating metabolic profiling of pancreatic juice with transcriptomic analysis of pancreatic cancer tissue identifies distinct clinical subgroups.

Introduction: Metabolic reprogramming is a hallmark feature of pancreatic ductal adenocarcinoma (PDAC). A pancreatic juice (PJ) metabolic signature has been reported to be prognostic of oncological outcome for PDAC. Integration of PJ profiling with transcriptomic and spatial characterization of the tumor microenvironment would help in identifying PDACs with peculiar vulnerabilities. Methods: We performed a transcriptomic analysis of 26 PDAC samples grouped into 3 metabolic clusters (M_CL) according to their PJ metabolic profile. We analyzed molecular subtypes and transcriptional differences. Validation was performed by multidimensional imaging on tumor slides. Results: Pancreatic juice metabolic profiling was associated with PDAC transcriptomic molecular subtypes (p=0.004). Tumors identified as M_CL1 exhibited a non-squamous molecular phenotype and demonstrated longer survival. Enrichment analysis revealed the upregulation of immune genes and pathways in M_CL1 samples compared to M_CL2, the group with worse prognosis, a difference confirmed by immunofluorescence on tissue slides. Enrichment analysis of 39 immune signatures by xCell confirmed decreased immune signatures in M_CL2 compared to M_CL1 and allowed a stratification of patients associated with longer survival. Discussion: PJ metabolic fingerprints reflect PDAC molecular subtypes and the immune microenvironment, confirming PJ as a promising source of biomarkers for personalized therapy.

Integrating AI-Powered Digital Pathology and Imaging Mass Cytometry Identifies Key Classifiers of Tumor Cells, Stroma, and Immune Cells in Non-Small Cell Lung Cancer.

Artificial intelligence (AI)-powered approaches are becoming increasingly used as histopathologic tools to extract subvisual features and improve diagnostic workflows. On the other hand, hi-plex approaches are widely adopted to analyze the immune ecosystem in tumor specimens. Here, we aimed at combining AI-aided histopathology and imaging mass cytometry (IMC) to analyze the ecosystem of non-small cell lung cancer (NSCLC). An AI-based approach was used on hematoxylin and eosin (H&E) sections from 158 NSCLC specimens to accurately identify tumor cells, both adenocarcinoma and squamous carcinoma cells, and to generate a classifier of tumor cell spatial clustering. Consecutive tissue sections were stained with metal-labeled antibodies and processed through the IMC workflow, allowing quantitative detection of 24 markers related to tumor cells, tissue architecture, CD45+ myeloid and lymphoid cells, and immune activation. IMC identified 11 macrophage clusters that mainly localized in the stroma, except for S100A8+ cells, which infiltrated tumor nests. T cells were preferentially localized in peritumor areas or in tumor nests, the latter being associated with better prognosis, and they were more abundant in highly clustered tumors. Integrated tumor and immune classifiers were validated as prognostic on whole slides. In conclusion, integration of AI-powered H&E and multiparametric IMC allows investigation of spatial patterns and reveals tissue relevant features with clinical relevance. SIGNIFICANCE: Leveraging artificial intelligence-powered H&E analysis integrated with hi-plex imaging mass cytometry provides insights into the tumor ecosystem and can translate tumor features into classifiers to predict prognosis, genotype, and therapy response.

Immune cells: plastic players along colorectal cancer progression.

Inflammatory cells are involved in tumour initiation and progression. In parallel, the adaptive immune response plays a key role in fighting tumour growth and dissemination. The double-edged role of the immune system in solid tumours is well represented in colorectal cancer (CRC). The development and progression of CRC are affected by the interactions between the tumour and the host's response, occurring in a milieu named tumour microenvironment. The role of immune cells in human CRC is being unravelled and there is a strong interest in understanding their dynamics as to tumour promotion, immunosurveillance and immunoevasion. A better definition of immune infiltration would be important not only with respect to the 'natural history' of CRC, but in a clinically relevant perspective in the 21st century, with respect to its post-surgical management, including chemotherapy responsiveness. While it is becoming established that the amount of tumour-infiltrating lymphocytes influences the post-surgical progression of early-stage CRC, the relevance of this immune parameter as to chemotherapy responsiveness remains to be clarified. Despite recent experimental work supporting the notion that infiltrating immune cells may influence chemotherapy-mediated tumour cell death, tumour-infiltrating cells are not employed to identify patients who are more likely to benefit from adjuvant treatment. This review focuses on studies addressing the role of innate and adaptive immune cells along the occurrence and the progression of potentially curable CRC.

Clinical frailty scale as a predictor of outcome in elderly patients affected by moderate or severe traumatic brain injury.

Background: Older age is a well-known risk factor for unfavorable outcome in traumatic brain injury (TBI). However, many older people with TBI respond well to aggressive treatments, suggesting that chronological age and TBI severity alone may be inadequate prognostic markers. Frailty is an age-related homeostatic imbalance of loss of physiologic and cognitive reserve resulting in both limitation in autonomy of activities of daily living and vulnerability to adverse events. We hypothesized that frailty would be associated with 6-month adverse functional outcome in older people affected by moderate or severe TBI. Methods: This was a single-center prospective observational study. We enrolled consecutive patients aged ≥65 years after TBI with Glasgow Coma Scale ≤13 and admitted to our Neurosurgical Intensive Care Unit. Frailty was evaluated by Clinical Frailty Scale (CFS). Relationships between TBI severity, frailty and extended Glasgow Outcome Scale (GOSE) at 6-month were evaluated. Results: Sixty patients were studied, 65% were males, their age was 76 years (IQR 70-80) and their admission GCS was 8 (IQR 6-11) with a GCS motor score of 5 (IQR 4-5). Twenty eight were vulnerable-frail (defined as CFS ≥ 4). Vulnerable-frail patients showed greater 6-month mortality and unfavorable outcome compared to non-frail [87% vs. 30% OR and 95% CI: 15.7 (3.9-55.2), p < 0.0001 and 92% vs. 51% OR and 95% CI: 9.9 (2.1-46.3), p = 0.002]. In univariate analysis patients with unfavorable outcome were more frequently male and vulnerable-frail, had a higher prevalence of pre-existing neurodegenerative disease, abnormal pupil, lower GCS and had worst CT scan characteristics. At multivariate analysis, only CFS ≥ 4 and traumatic subarachnoid hemorrhage remained associated to 6-month outcome. Conclusion: Frailty was associated with 6 month-outcome, suggesting that the pre-injury functional status could represent an additional indicator to stratify patient's severity and to predict outcome.

Macrophage morphology and distribution are strong predictors of prognosis in resected colorectal liver metastases: results from an external retrospective observational study.

INTRODUCTION: Tumor-associated macrophages (TAMs) are key components of a tumoral microenvironment and have been shown to impact prognosis in different cancers. Previously reported data showed that TAM morphology correlates with prognosis in colorectal liver metastases (CLMs) after hepatectomy, with smaller TAMs (S-TAMs) conferring a more favorable prognosis than larger ones (L-TAMs). This study aims to externally validate this finding. MATERIAL AND METHODS: The external cohort consisted of 84 formalin-fixed and paraffin-embedded surgical samples of CLMs and peritumoral tissue. Two-micrometer-section slides were obtained; the area and perimeter of 21 macrophages in each slide were recorded. The endpoints were TAMs morphometrics and their prognostic significance in relation to disease-free survival (DFS). RESULTS: The average macrophage perimeter was 71.5±14.1 µm whilst the average area was 217.7±67.8 µm 2 . At univariate analysis, the TAM area demonstrated a statistically significant association with DFS ( P =0.0006). Optimal area cutoff value was obtained, showing a sensitivity and specificity of 92 and 56%, respectively. S-TAMs and L-TAMs were associated with 3-year DFS rates of 60 and 8.5%, respectively ( P <0.001). Multivariate analysis confirmed the predictive role of TAM area for DFS [hazard ratio (HR)=5.03; 95% CI=1.70-14.94; P =0.003]. Moreover, in a subset of patients ( n =12) characterized by unfavorable ( n =6, recurrence within 3 months) or favorable ( n =6, no recurrence after 48 months) prognosis, TAMs showed a different distribution: L-TAMs were more abundant and closer to the tumor invasive margin in patients that encountered early recurrence and tended to cluster in foci significantly larger ( P =0.02). CONCLUSIONS: This external validation confirms that morphometric characterization of TAMs can serve as a simple readout of their diversity and allows to reliably stratify patient outcomes and predict disease recurrence after hepatectomy for CLMs.

High-Resolution Analysis of Mononuclear Phagocytes Reveals GPNMB as a Prognostic Marker in Human Colorectal Liver Metastasis.

Patients with colorectal liver metastasis (CLM) present with heterogenous clinical outcomes and improved classification is needed to ameliorate the therapeutic output. Macrophages (Mϕ) hold promise as prognostic classifiers and therapeutic targets. Here, stemming from a single-cell analysis of mononuclear phagocytes infiltrating human CLM, we identified two Mϕ markers associated with distinct populations with opposite clinical relevance. The invasive margin of CLM was enriched in pro-inflammatory monocyte-derived Mϕ (MoMϕ) expressing the monocytic marker SERPINB2, and a more differentiated population, tumor-associated Mϕ (TAM), expressing glycoprotein nonmetastatic melanoma protein B (GPNMB). SERPINB2+ MoMϕ had an early inflammatory profile, whereas GPNMB+ TAMs were enriched in pathways of matrix degradation, angiogenesis, and lipid metabolism and were found closer to the tumor margin, as confirmed by spatial transcriptomics on CLM specimens. In a cohort of patients, a high infiltration of SERPINB2+ cells independently associated with longer disease-free survival (DFS; P = 0.033), whereas a high density of GPNMB+ cells correlated with shorter DFS (P = 0.012) and overall survival (P = 0.002). Cell-cell interaction analysis defined opposing roles for MoMϕ and TAMs, suggesting that SERPINB2+ and GPNMB+ cells are discrete populations of Mϕ and may be exploited for further translation to an immune-based stratification tool. This study provides evidence of how multi-omics approaches can identify nonredundant, clinically relevant markers for further translation to immune-based patient stratification tools and therapeutic targets. GPNMB has been shown to set Mϕ in an immunosuppressive mode. Our high dimensional analyses provide further evidence that GPNMB is a negative prognostic indicator and a potential player in the protumor function of Mϕ populations.

hMENA isoforms regulate cancer intrinsic type I IFN signaling and extrinsic mechanisms of resistance to immune checkpoint blockade in NSCLC.

BACKGROUND: Understanding how cancer signaling pathways promote an immunosuppressive program which sustains acquired or primary resistance to immune checkpoint blockade (ICB) is a crucial step in improving immunotherapy efficacy. Among the pathways that can affect ICB response is the interferon (IFN) pathway that may be both detrimental and beneficial. The immune sensor retinoic acid-inducible gene I (RIG-I) induces IFN activation and secretion and is activated by actin cytoskeleton disturbance. The actin cytoskeleton regulatory protein hMENA, along with its isoforms, is a key signaling hub in different solid tumors, and recently its role as a regulator of transcription of genes encoding immunomodulatory secretory proteins has been proposed. When hMENA is expressed in tumor cells with low levels of the epithelial specific hMENA11a isoform, identifies non-small cell lung cancer (NSCLC) patients with poor prognosis. Aim was to identify cancer intrinsic and extrinsic pathways regulated by hMENA11a downregulation as determinants of ICB response in NSCLC. Here, we present a potential novel mechanism of ICB resistance driven by hMENA11a downregulation. METHODS: Effects of hMENA11a downregulation were tested by RNA-Seq, ATAC-Seq, flow cytometry and biochemical assays. ICB-treated patient tumor tissues were profiled by Nanostring IO 360 Panel enriched with hMENA custom probes. OAK and POPLAR datasets were used to validate our discovery cohort. RESULTS: Transcriptomic and biochemical analyses demonstrated that the depletion of hMENA11a induces IFN pathway activation, the production of different inflammatory mediators including IFNß via RIG-I, sustains the increase of tumor PD-L1 levels and activates a paracrine loop between tumor cells and a unique macrophage subset favoring an epithelial-mesenchymal transition (EMT). Notably, when we translated our results in a clinical setting of NSCLC ICB-treated patients, transcriptomic analysis revealed that low expression of hMENA11a, high expression of IFN target genes and high macrophage score identify patients resistant to ICB therapy. CONCLUSIONS: Collectively, these data establish a new function for the actin cytoskeleton regulator hMENA11a in modulating cancer cell intrinsic type I IFN signaling and extrinsic mechanisms that promote protumoral macrophages and favor EMT. These data highlight the role of actin cytoskeleton disturbance in activating immune suppressive pathways that may be involved in resistance to ICB in NSCLC.

Cancer-promoting tumor-associated macrophages: new vistas and open questions.

Tumor-associated macrophages (TAMs) are key components of the tumor macroenvironment. Cancer- and host cell-derived signals generally drive the functions of TAMs towards an M2-like polarized, tumor-propelling mode; however, when appropriately re-educated. TAMs also have the potential to elicit tumor destructive reactions. Here, we discuss recent advances regarding the immunobiology of TAMs and highlight open questions including the mechanisms of their accumulation (recruitment versus proliferation), their diversity and how to best therapeutically target these cells.