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Purpose A specific biomarker of post-ARDS fibroproliferation could be useful in the identification of patients who could benefit from therapies aiming to modulate fibroproliferation such as corticosteroids.The aim of this prospective study was to determine the best threshold of the N-terminal-peptidetype III procollagen (NT-PCP-III) in non-resolving ARDS to validate this threshold according to the outcome. Methods Concerning the best threshold of NT-PCP-III, all consecutive patients with a non-resolving ARDS were included if all the following criteria were fulfilled: moderate to severe ARDS lasting for at least 5 days, lung biopsy performed, serum and alveolar NT-PCP-III obtained within 1 week prior to biopsy, and no documented infection contra-indicating the corticosteroids. In the validation cohort part of the study, patients were included at day 7 if they presented a persistent moderate to severe ARDS. Results Nineteen of 32 patients had fibroproliferatio nonbiopsy. Serum and alveolar NT-PCP-III were higher in patients with fibroproliferation. Using a threshold of 9 [micro]g/L, alveolar NT-PCP-III had the highest accuracy for diagnosing fibroproliferation (sensitivity = 89.5 % and specificity = 92.3 %). Regarding the 51 patients included in the validation cohort, the mortality rate at day 60 was increased in patients presenting an alveolar NT-PCP-III level higher than 9 [micro]g/L (69 vs. 17 %, p < 0.001). The mean alveolar level of NT-PCP-III on day 7 was 8.1-fold higher in nonsurvivors (p = 0.03). Conclusions The determination of NT-PCP-III on BAL done at day 7 in persistent ARDS is able to identify patients with fibroproliferation who could be included in a trial of corticosteroids or any other treatment that might help resolve lung fibroproliferation., Author(s): Jean-Marie Forel [sup.1] [sup.2], Christophe Guervilly [sup.2], Sami Hraiech [sup.1] [sup.2], François Voillet [sup.2], Guillemette Thomas [sup.1] [sup.2], Claude Somma [sup.3], Véronique Secq [sup.1] [sup.4], Catherine Farnarier [sup.5], Marie-Josée [...]

Background: In lung adenocarcinoma, molecular profiling of actionable genes has become essential to set up targeted therapies. However, the feasibility and the relevance of molecular profiling from the cerebrospinal fluid (CSF) in the context of meningeal metastasis have been poorly assessed. Methods: We selected patients with stage IV lung adenocarcinoma harbouring metastatic cells in the CSF after cytological analysis. Seven samples from six patients were eligible for molecular testing of epidermal growth factor receptor (EGFR), V-Ki-ras2 Kirsten rat sarcoma viral oncogene homologue (KRAS), v-Raf murine sarcoma viral oncogene homologue B1 (BRAF) and human epidermal growth factor receptor 2 (HER2) mutations using quantitative polymerase chain reaction (PCR) high-resolution melting curve analysis and Sanger sequencing after DNA extraction from the cell pellets of the CSF. Results: Five patients showed mutations in one or two actionable genes, two harboured an EGFR mutation (exons 19 and 21), one only a KRAS mutation, one both EGFR and KRAS mutations and one a BRAF mutation. In all cases, the results of mutation testing provided new major information for patient management, leading to therapeutic adaptation. CSF molecular analysis identified mutations not detected in other neoplastic sites for two patients. In one case, the EGFR p.Thr790Met was identified. CSF was also the only sample available for genetic testing for almost all patients at the time of disease progression. Conclusions: When cancer cells are present in the CSF, the molecular profiling from the cell pellets is relevant, as it can detect supplemental or different mutations compared to a previous analysis of the primitive tumour or plasma cell-free DNA and allows the adaptation of the treatment strategy. [ABSTRACT FROM AUTHOR]

Objective: Despite recent advances in imaging and core or endoscopic biopsies, a percentage of patients have a major lung resection without diagnosis. We aimed to assess the feasibility of a rapid tissue preparation/analysis to discriminate cancerous from non-cancerous lung tissue. Methods: Fresh sample preparations were analyzed with the Microflex LTTM MALDI-TOF analyzer. Each main reference spectra (MSP) was consecutively included in a database. After definitive pathological diagnosis, each MSP was labeled as either cancerous or non-cancerous (normal, inflammatory, infectious nodules). A strategy was constructed based on the number of concordant responses of a mass spectrometry scoring algorithm. A 3-step evaluation included an internal and blind validation of a preliminary database (n = 182 reference spectra from the 100 first patients), followed by validation on a whole cohort database (n = 300 reference spectra from 159 patients). Diagnostic performance indicators were calculated. Results: 127 cancerous and 173 non-cancerous samples (144 peripheral biopsies and 29 inflammatory or infectious lesions) were processed within 30 minutes after biopsy sampling. At the most discriminatory level, the samples were correctly classified with a sensitivity, specificity and global accuracy of 92.1%, 97.1% and 95%, respectively. Conclusions: The feasibility of rapid MALDI-TOF analysis, coupled with a very simple lung preparation procedure, appears promising and should be tested in several surgical settings where rapid on-site evaluation of abnormal tissue is required. In the operating room, it appears promising in case of tumors with an uncertain preoperative diagnosis and should be tested as a complementary approach to frozen-biopsy analysis. [ABSTRACT FROM AUTHOR]

We hypothesized that inhibiting molecules that mediate the adaptation response to cellular stress can antagonize the resistance of pancreatic cancer cells to chemotherapeutic drugs. Toward this end, here, we investigated how VMP1, a stress-induced autophagy-associated protein, modulate stress responses triggered by chemotherapeutic agents in PDAC. We find that VMP1 is particularly over-expressed in poorly differentiated human pancreatic cancer. Pharmacological studies show that drugs that work, in part, via the endoplasmic reticulum stress response, induce VMP1 expression. Similarly, VMP1 is induced by known endoplasmic reticulum stress activators. Genetic inactivation of VMP1 using RNAi-based antagonize the pancreatic cancer stress response to antitumoral agents. Functionally, we find that VMP1 regulates both autophagy and chemotherapeutic resistance even in the presence of chloroquin, ATG5 or Beclin 1 siRNAs, or a Beclin 1-binding VMP1 mutant. In addition, VMP1 modulates endoplasmic reticulum stress independently of its coupling to the molecular and cellular autophagy machinery. Preclinical studies demonstrate that xenografts expressing an inducible and tractable form of VMP1 show increased resistance to the gemcitabine treatment. These results underscore a novel role for VMP1 as a potential therapeutic target for combinatorial therapies aimed at sensitizing pancreatic cancer cells to chemotherapeutic agents as well as provide novel molecular mechanisms to better understand this phenomenon. J. Cell. Physiol. 228: 1834-1843, 2013. © 2013 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Vγ9Vδ2 T cells are anti-tumor immune effectors of growing interest in cancer including Pancreatic Ductal Adenocarcinoma (PDAC), an especially aggressive cancer characterized by a hypoxic and nutrient-starved immunosuppressive microenvironment. Since Butyrophilin 3 A (BTN3A) isoforms are critical activating molecules of Vγ9Vδ2 T cells, we set out to study BTN3A expression under both basal and stress conditions in PDAC primary tumors, and in novel patient-derived xenograft and PDAC-derived cell lines. BTN3A2 was shown to be the most abundant isoform in PDAC and was stress-regulated. Vγ9Vδ2 T cells cytolytic functions against PDAC required BTN3A and this activity was strongly enhanced by the agonist anti-BTN3A 20.1 mAb even under conditions of hypoxia. In PDAC primary tumors, we established that BTN3A expression and high plasma levels of soluble BTN3A were strongly associated with a decreased survival. These findings may have important implications in the design of new immunotherapeutic strategies that target BTN3A for treating PDAC. [ABSTRACT FROM PUBLISHER]

Classification of thymic epithelial tumours (TETs) is known to be challenging; however, the level of discordances at a nationwide level between initial and expert diagnosis and their clinical consequences are currently unknown. RYTHMIC is a national network dedicated to the management of TET based on initial histological diagnosis, followed by an additional expert review of all cases. Our aim was to evaluate the discordances between initial and expert diagnoses and whether they would have led to different clinical management. We conducted a retrospective analysis of the cohort of patients discussed at RYTHMIC tumour board from January 2012 to December 2016. Assessment of disagreement was made for histological typing and for staging. The discordances were classified as major or minor based on whether they would have changed or not the proposed therapeutic strategy, respectively. Follow-up of the patients with major discordances was conducted until December 2018. Four hundred sixty-seven patients were reviewed, and 183 (39%) discordances were identified either related to histological subtype (132) and/or stage (72). Major discordances were identified in 27 patients (6%). They included 16 patients with TET for whom treatment recommendation based on the central review would have been post-operative radiotherapy, whereas it had not been the case. However, follow-up did not show any progression among the 15 patients with high-grade histology and/or stage resected thymomas. On the other hand, among the remaining 11 patients including 7 with a diagnosis other than TET, the overall management or follow-up would have been completely different with the expert diagnosis. Our real-life cohort reveals a high level of discordances considering TET diagnosis and supports expert review for optimal clinical management. • We analysed the impact of expert pathologic review of thymic epithelial tumours (TETs) in a nationwide network. • A cohort of 467 patients with TET was studied in a real-life setting. • We identified an overall discordance rate of 39% considering histotype and/or stage. • Change for post-operative radiotherapy or disease management involved 6% of patients. • Expert pathologic review of TET contributes to a better treatment decision-making. [ABSTRACT FROM AUTHOR]

Diffuse alveolar hemorrhage (DAH) is a life-threatening complication of systemic lupus erythematosus (SLE) and systemic vasculitis. Although initially described to have antibacterial properties, increasing evidence suggests that neutrophil extracellular traps (NETs) have a detrimental role in both autoimmune diseases and acute lung injury. We investigated whether NETs could be detected in a murine model of pristane-induced lupus DAH and contribute to lung injury. Such NETs might constitute a therapeutic target. NETs were characterized by immunofluorescence staining of DNA, neutrophil elastase and citrullinated histones. Evaluation of lung injury was performed by haematoxylin-eosin staining and a quantification program. Clinical status of the mice was assessed by measurement of arterial oxygen saturation and survival curves after recombinant human deoxyribonuclease-1 (Rh-DNase-1) inhalations or polymorphonuclear neutrophil (PMN) depletion. Pristane was found to promote NETs formation in vitro and in vivo. Treatment of mice with Rh-DNase-1 inhalations cleared NETs and reduced lung injury. Clinical status improved significantly, with increased arterial oxygenation and survival. Following PMN depletion, NETs were absent with a subsequent reduction of lung injury and improved arterial oxygenation. These results support a pathogenic role of PMNs and NETs in lung injury during pristane-induced DAH. Targeting NETs with Rh-DNase-1 inhalations could constitute an interesting adjuvant therapy in human DAH. • Pristane-induced diffuse alveolar hemorrhage (DAH) lupus model can be considered as a pulmonary model of NETosis. • Targeting NETs with recombinant human deoxyribonuclease reduced severity of DAH lesions and improves animal clinical status. • Recombinant human deoxyribonuclease could be an interesting adjuvant therapy in human DAH. [ABSTRACT FROM AUTHOR]

The intratumoral microenvironment, or stroma, is of major importance in the pathobiology of pancreatic ductal adenocarcinoma (PDA), and specific conditions in the stroma may promote increased cancer aggressiveness. We hypothesized that this heterogeneous and evolving compartment drastically influences tumor cell abilities, which in turn influences PDA aggressiveness through crosstalk that is mediated by extracellular vesicles (EVs). Here, we have analyzed the PDA proteomic stromal signature and identified a contribution of the annexin A6/LDL receptor-related protein 1/thrombospondin 1 (ANXA6/LRP1/TSP1) complex in tumor cell crosstalk. Formation of the ANXA6/LRP1/TSP1 complex was restricted to cancer-associated fibroblasts (CAFs) and required physiopathologic culture conditions that improved tumor cell survival and migration. Increased PDA aggressiveness was dependent on tumor cell-mediated uptake of CAF-derived ANXA6+ EVs carrying the ANXA6/LRP1/TSP1 complex. Depletion of ANXA6 in CAFs impaired complex formation and subsequently impaired PDA and metastasis occurrence, while injection of CAF-derived ANXA6+ EVs enhanced tumorigenesis. We found that the presence of ANXA6+ EVs in serum was restricted to PDA patients and represents a potential biomarker for PDA grade. These findings suggest that CAF-tumor cell crosstalk supported by ANXA6+ EVs is predictive of PDA aggressiveness, highlighting a therapeutic target and potential biomarker for PDA. [ABSTRACT FROM AUTHOR]

Objectives Glucose-transporter-1 (Glut-1) may be a useful marker for differentiating B3 thymomas and thymic carcinomas. Since the literature is limited, we undertook a study to evaluate its diagnostic value in a series of thymic epithelial tumors. Materials and methods Glut-1 expression was studied by the group of pathologists linked to the French national oncological network RYTHMIC. Immunostaining was performed on a whole section of one paraffin block in a series of 92 successive surgical specimens. Patterns (focal, zonal, diffuse) and intensity of Glut-1 expression were assessed and compared with WHO histological subtypes. Results Expression was mainly restricted to epithelial cells. Immature T-lymphocytes were negative. A diffuse, moderate or strong staining was observed in most thymic carcinomas (15/16). In B3 thymomas (10/11) and in B3 thymomas borderline to thymic carcinomas (5/6), a moderate to strong zonal staining was observed at distance from vessels and fibrous septa. This pattern sometimes created the aspect of an anastomosing network in large cellular lobules. In B1 thymomas, immunostaining highlighted foci of medullary differentiation (7/8). B2 thymomas ( n = 25) were heterogeneous, with a spectrum of patterns ranging between those of B1 and B3 thymomas. Type A thymomas ( n = 5) mostly presented a weak positivity but one aggressive case showed zonal moderate/strong positivity. Most AB thymomas (15/17) showed weak to moderate immunostaining in spindle cell areas. In micronodular thymomas ( n = 3), epithelial cells and B-lymphocytes were weakly positive while follicular dendritic cells were strongly highlighted. One metaplastic thymoma displayed diffuse and moderate positivity. Conclusion Glut-1 expression globally depended on histological subtypes and the staining patterns (diffuse or zonal) were different between thymic carcinomas and type B3 thymomas. A comparative study of Glut-1 expression in atypical versus conventional type A thymomas appears warranted. Otherwise, restriction to epithelial cells makes likely a correlation with clinical assessment of glucose uptake in lymphocyte-poor tumors. [ABSTRACT FROM AUTHOR]

Abstract: Background: CK2α is a signalling molecule that participates in major events in solid tumour progression. The aim of this study was to evaluate the prognostic significance of the immunohistochemical expression of CK2α in breast carcinomas. Methods: Quantitative measurements of immunohistochemical expression of 33 biomarkers using high-throughput densitometry, assessed on digitised microscopic tissue micro-array images were correlated with clinical outcome in 1000 breast carcinomas using univariate and multivariate analyses. Results: In univariate analysis, CK2α was a significant prognostic indicator (p <0.001). Moreover, a multivariable model allowed the selection of the best combination of the 33 biomarkers to predict patients’ outcome through logistic regression. A nine-marker signature highly predictive of metastatic risk, associating SHARP-2, STAT1, eIF4E, pmapKAPk-2, pAKT, caveolin, VEGF, FGF-1 and CK2α permitted to classify well 82.32% of patients (specificity 81.59%, sensitivity 92.55%, area under ROC curve 0.939). Importantly, in a node negative subset of patients an even more (86%) clinically relevant association of eleven markers was found predictive of poor outcome. Conclusion: A strong quantitative CK2α immunohistochemical expression in breast carcinomas is individually a significant indicator of poor prognosis. Moreover, an immunohistochemical signature of 11 markers including CK2α accurately (86%) well classifies node negative patients in good and poor outcome subsets. Our results suggest that CK2α evaluation together with key downstream CK2 targets might be a useful tool to identify patients at high risk of distant metastases and that CK2 can be considered as a relevant target for potential specific therapy. [ABSTRACT FROM AUTHOR]

Childhood pulmonary Langerhans cell histiocytosis (PLCH) is a rare disease. Its pulmonary histopathology, according to comprehensive clinical-radiological findings and BRAF V600E mutation status, has not yet been thoroughly documented. From the 167 childhood PLCH cases entered in the French National Histiocytosis Registry (1983-2016), we retrieved lung biopsies from a consecutive retrospective series of 17 patients, diagnosed when they were 2 weeks to 16 years old (median, 9.4 years), and report the clinical and histopathological findings herein. Histological analyses of biopsies (16 surgical and 1 postmortem) found the following features, alone or associated: Langerhans cell (LC) nodules with cavitation (9/17), cysts (14/17), fibrotic scars (2/17), peribronchiolar topographic distribution of the lesions (10/17), and accessory changes, like stretch emphysema (7/17). Those characteristics closely resemble those describing adult PLCH. However, unusual findings observed were 2 large nodules and a diffuse interstitial LC infiltrate. BRAF V600E mutation was detected in 4 of 12 samples tested, notably in the 3 with unusual features. In conclusion, childhood PLCH mostly shares the common histology features already described in adult PLCH, regardless of age. Because smoking is considered the major trigger in PLCH pathogenesis, the findings based on this series suggest other inducers of bronchiolar LC recruitment, especially in very young patients., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Primary defects in lung branching morphogenesis, resulting in neonatal lethal pulmonary hypoplasias, are incompletely understood. To elucidate the pathogenetics of human lung development, we studied a unique collection of samples obtained from deceased individuals with clinically and histopathologically diagnosed interstitial neonatal lung disorders: acinar dysplasia (n = 14), congenital alveolar dysplasia (n = 2), and other lethal lung hypoplasias (n = 10). We identified rare heterozygous copy-number variant deletions or single-nucleotide variants (SNVs) involving TBX4 (n = 8 and n = 2, respectively) or FGF10 (n = 2 and n = 2, respectively) in 16/26 (61%) individuals. In addition to TBX4, the overlapping ∼2 Mb recurrent and nonrecurrent deletions at 17q23.1q23.2 identified in seven individuals with lung hypoplasia also remove a lung-specific enhancer region. Individuals with coding variants involving either TBX4 or FGF10 also harbored at least one non-coding SNV in the predicted lung-specific enhancer region, which was absent in 13 control individuals with the overlapping deletions but without any structural lung anomalies. The occurrence of rare coding variants involving TBX4 or FGF10 with the putative hypomorphic non-coding SNVs implies a complex compound inheritance of these pulmonary hypoplasias. Moreover, they support the importance of TBX4-FGF10-FGFR2 epithelial-mesenchymal signaling in human lung organogenesis and help to explain the histopathological continuum observed in these rare lethal developmental disorders of the lung., (Copyright © 2018 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Pancreatic ductal adenocarcinoma (PDAC) is characterized by extensive stroma and pathogenic modifications to the peripheral nervous system that elevate metastatic capacity. In this study, we show that the IL6-related stem cell-promoting factor LIF supports PDAC-associated neural remodeling (PANR). LIF was overexpressed in tumor tissue compared with healthy pancreas, but its receptors LIFR and gp130 were expressed only in intratumoral nerves. Cancer cells and stromal cells in PDAC tissues both expressed LIF, but only stromal cells could secrete it. Biological investigations showed that LIF promoted the differentiation of glial nerve sheath Schwann cells and induced their migration by activating JAK/STAT3/AKT signaling. LIF also induced neuronal plasticity in dorsal root ganglia neurons by increasing the number of neurites and the soma area. Notably, injection of LIF-blocking antibody into PDAC-bearing mice reduced intratumoral nerve density, supporting a critical role for LIF function in PANR. In serum from human PDAC patients and mouse models of PDAC, we found that LIF titers positively correlated with intratumoral nerve density. Taken together, our findings suggest LIF as a candidate serum biomarker and diagnostic tool and a possible therapeutic target for limiting the impact of PANR in PDAC pathophysiology and metastatic progression. Significance: This study suggests a target to limit neural remodeling in pancreatic cancer, which contributes to poorer quality of life and heightened metastatic progression in patients. Cancer Res; 78(4); 909-21. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Vγ9Vδ2 T cells are anti-tumor immune effectors of growing interest in cancer including Pancreatic Ductal Adenocarcinoma (PDAC), an especially aggressive cancer characterized by a hypoxic and nutrient-starved immunosuppressive microenvironment. Since Butyrophilin 3 A (BTN3A) isoforms are critical activating molecules of Vγ9Vδ2 T cells, we set out to study BTN3A expression under both basal and stress conditions in PDAC primary tumors, and in novel patient-derived xenograft and PDAC-derived cell lines. BTN3A2 was shown to be the most abundant isoform in PDAC and was stress-regulated. Vγ9Vδ2 T cells cytolytic functions against PDAC required BTN3A and this activity was strongly enhanced by the agonist anti-BTN3A 20.1 mAb even under conditions of hypoxia. In PDAC primary tumors, we established that BTN3A expression and high plasma levels of soluble BTN3A were strongly associated with a decreased survival. These findings may have important implications in the design of new immunotherapeutic strategies that target BTN3A for treating PDAC.

Noninvasive early detection of breast cancer through the use of biomarkers is urgently needed since the risk of recurrence, morbidity, and mortality is closely related to disease stage at the time of primary surgery. A crucial issue in this approach is the availability of relevant markers and corresponding monoclonal antibodies suitable for the development of effective immunodiagnostic modalities. The identification of such markers from human pathological lesions and the isolation of specific antibodies using conventional approaches remain major challenges. Camelids produce functional antibodies devoid of light chains in which the single N-terminal domain of the heavy chain is fully capable of antigen binding. When produced as an independent domain, these so-called single-domain antibody fragments (sdAbs) or nanobodies have several advantages for biotechnological applications owing to their unique properties of size (13 kDa), stability, solubility, and expression yield. In this work, we have generated phage display libraries from animals immunized with breast cancer biopsies. These libraries were used to isolate sdAbs against known and relevant antigens such as HER2, or several cancer-specific sdAbs against unknown targets. We describe the identification of one these targets, cytokeratin 19, using affinity purification in combination with mass spectrometry. Some of these sdAbs were used in several straightforward diagnostic applications such as immunohistochemical analysis of tumor samples, multiplexed cytometric bead array analysis of crude samples, or an immune enrichment procedure of rare cells. Here, we demonstrate that phage display-based selection of single-domain antibodies is an efficient and high-throughput compatible approach to generate binders with excellent characteristics for the fast development of diagnostic and prognostic modalities.

We aimed in this study at identifying prognostic immunohistochemical molecular signatures indicative of disease outcome, also relevant for development of new specific therapies, in triple-negative (ER, PR, c-erbB2- negative) breast carcinoma subtypes. We evaluated 42 markers in tissue micro-arrays from a series of 924 breast carcinomas including 184 triple-negative tumors using standardized quantitative immunocytochemical assays and correlated the data with patients' outcome (mean follow-up of 79 months). When 27/42 markers including basal-like markers first found to be individually significant for prognosis in a univariate analysis (log-rank test) in 924 tumors, were secondly evaluated in the triple-negative tumor subtype (184/924), eleven including maspin, P21, P27, PTEN, caveolin, EGFR, FAK, P38, pMAPK, STAT1 and CD10 were 89.2% predictive of disease outcome in logistic regression. When markers reported in the literature as expressed in basal-like subtype were evaluated in the 924 series, only eight (EGFR, CK14, moesin, caveolin, cMet, ckit, CD44v6, C10) were prognosis predictive in univariate analysis (log-rank test) and in logistic regression were predictive of disease outcome in 66.3% independently of ER, PR and c-erbB2 expression and in 72% in triple-negative tumor subset. The results suggest that the category of 'triple-negative' breast carcinomas does not exactly overlap the basal-like subtype, and that immunoprofiling of triple-negative tumors (not similar to that of basal-like tumors) may be helpful to select patients for more aggressive treatment and provides a basis for development of tailored therapy.