Flora Poizat

Inherent and Tumor-Driven Immune Tolerance in the Prostate Microenvironment Impairs Natural Killer Cell Antitumor Activity

The field of immunotherapy for solid tumors, such as prostate cancer, has been recently focusing on therapies that can counter tumor-mediated immunosuppression. Precise quantification and characterization of the immune infiltrates in tumors is crucial to improve treatment efficacy. Natural killer (NK) cells, major components of the antitumor immune system, have never been isolated from prostate tumors, despite their suspected role in disease progression. Here, we examined the frequency, phenotype, and functions of NK cells infiltrating control and tumor prostate tissues. NK cell infiltrates in prostate tissues were mainly CD56 (NCAM1)-positive and displayed an unexpected immature, but activated, phenotype with low or no cytotoxic potential. Furthermore, we show that TGFβ1 (TGFB1) is highly secreted into the prostate environment and partly mediates the immunosuppressive effects on NK cells. In addition to this basal level of immunotolerance to NK cells, the prostate environment became further resistant to NK cell-mediated immunity upon cancer cell infiltration. Coculture experiments revealed that prostate cancer cells induced the expression of inhibitory receptor (ILT2/LILRB1) and downregulated the expression of activating receptors NKp46 (NCR1), NKG2D (KLRK1), and CD16 (FCGR3) by NK cells, thus preventing their recognition of tumor cells. Notably, blood levels of NKp46 were also decreased in prostate cancer patients and were inversely correlated with levels of prostate-specific antigen, the main prognostic factor in prostate cancer. Our study shows that a strong immunosuppressive environment impairs NK cell function at multiple levels in prostate cancer and provides a rationale for the design of therapies that restore NK cell efficiency in the prostate tumor microenvironment. Cancer Res; 76(8); 2153-65. ©2016 AACR.

Prognostic value of a three-grade classification in primary epithelial parotid carcinoma: Result of a histological review from a 20-year experience of total parotidectomy with neck dissection in a single institution

BACKGROUND The tumour grading of primary parotid cancers (PPCs) remains controversial. METHODS A 20-year standardised single centre treatment has been assessed retrospectively. The histological review of 155 consecutively treated parotid malignancies identified 96 suitable cases for univariate and multivariate survival analyses. RESULTS Treatment involved total parotidectomy, neck dissection and post-operative radiotherapy in, respectively, 91.7%, 83.3% and 70.4% of cases. The 5-year overall survival, disease-specific and recurrence-free survival rates were 79.4%, 83.5% and 70.8%, respectively. Univariate analysis confirmed the classical prognostic factors, i.e. age>60 years, male gender, facial palsy, hardness of the tumour, clinical stage, tumour grade, facial nerve invasion and lymph node metastases. Multivariate analysis identified a three-grade classification just after the clinical stage as the most important prognostic factor. CONCLUSION This study identifies the prognostic significance of intermediate grade tumours.

Transcriptomic Analysis Predicts Survival and Sensitivity to Anticancer Drugs of Patients with a Pancreatic Adenocarcinoma

A major impediment to the effective treatment of patients with pancreatic ductal adenocarcinoma (PDAC) is the molecular heterogeneity of this disease, which is reflected in an equally diverse pattern of clinical outcome and in responses to therapies. We developed an efficient strategy in which PDAC samples from 17 consecutive patients were collected by endoscopic ultrasound-guided fine-needle aspiration or surgery and were preserved as breathing tumors by xenografting and as a primary culture of epithelial cells. Transcriptomic analysis was performed from breathing tumors by an Affymetrix approach. We observed significant heterogeneity in the RNA expression profile of tumors. However, the bioinformatic analysis of these data was able to discriminate between patients with long- and short-term survival corresponding to patients with moderately or poorly differentiated PDAC tumors, respectively. Primary culture of cells allowed us to analyze their relative sensitivity to anticancer drugs in vitro using a chemogram, similar to the antibiogram for microorganisms, establishing an individual profile of drug sensitivity. As expected, the response was patient dependent. We also found that transcriptomic analysis predicts the sensitivity of cells to the five anticancer drugs most frequently used to treat patients with PDAC. In conclusion, using this approach, we found that transcriptomic analysis could predict the sensitivity to anticancer drugs and the clinical outcome of patients with PDAC.

Réseau RENAPE : vers une nouvelle organisation des soins pour le traitement des tumeurs rares du péritoine. Description du réseau et rôle des pathologistes

As part of the national 2009-2013 Cancer Plan, and with the support of the National cancer Institute and the French ministry of health, the National network for the treatment of rare peritoneal malignancies (RENAPE) has been organized. Its main objective is to optimize the framework for the healthcare management and treatment of rare peritoneal malignancies. This specific organization covers the whole national territory including clinical expert and specialized structures and should lead to an appropriate treatment based on expertise and proximity. Within the RENAPE network, the RENA-PATH group gathers the pathologists actively involved in the management of rare peritoneal malignancies. The actions of RENA-PATH are focused primarily on the harmonization of pathological diagnostic criteria, reporting of new cases in the RENAPE registry and histology reviewing.

EGFR Expression and KRAS and BRAF Mutational Status in Intestinal-Type Sinonasal Adenocarcinoma.

Accumulation of molecular alterations, including EGFR overexpression and mutations in KRAS and BRAF, contribute to colorectal carcinogenesis. Since intestinal-type adenocarcinoma (ITAC) of the nasal cavity and paranasal sinus has morphologic and phenotypic features that are usually indistinguishable from colorectal cancer (CRC), it is likely that both tumor types share equivalent genetic alterations. Data from a series of 43 patients treated surgically for ITAC in Montpellier, France between November 1998 and December 2012 were collected. Tumors were characterized for mutations in KRAS and BRAF as well as EGFR overexpression. Kaplan-Meier survival curves were constructed using overall survival as the primary end points. Patient survival was analyzed using the hazards ratio. Twenty seven tumors (63%) showed EGFR positivity and 30% exhibited a high expression level (+2/+3). KRAS mutations were detected in 43% of cases. BRAF mutations were identified in 3.6% of specimens. Patients with age superior to 60 years, metastatic status, and KRAS mutations had significant overall survival values (p = 0.026, p = 0.001 and p = 0.03, respectively). Our results indicate that KRAS mutations and EGFR expression are frequent in ITAC and that KRAS mutations predict good patient prognosis in ITAC. Finally, EGFR directed molecular treatments could be investigated in a subset of patients affected by ITAC.

Comment améliorer la rentabilité diagnostique des biopsies de la sphère oto-rhino-laryngée dans la maladie de Wegener : analyse anatomo-clinique de 49 biopsies chez 21 patients

Resume Les biopsies de la sphere oto-rhino-laryngee (ORL) et conjonctivales sont classiquement rapportees comme peu contributives dans le diagnostic de la maladie de Wegener (MW). A partir de 49 biopsies ORL ou conjonctivales (fosses nasales 29, sinus 7, cavite buccale 4, larynx 4, conjonctive 3, oreille externe 2) realisees lors du diagnostic chez 21 patients atteints de MW, nous avons decrit les lesions histologiques evocatrices et etudie la rentabilite diagnostique de ces prelevements en fonction : de la presence de lesions macroscopiques, du site anatomique biopsie, des modalites d’obtention (anesthesie generale ou locale), de leur taille et du nombre de recoupes realisees. L’association de lesions granulomateuses (cellules geantes isolees (28,5 % des biopsies), granulomes epithelioides (28,5 %)), de lesions necrosantes (micro-abces a polynucleaires neutrophiles (16,3 %), foyers de necrose du tissu conjonctif (18,3 %)) et d’une vascularite (necrosante (12 %), leucocytoclasique (10 %), granulomateuse (6 %)) permettant d’affirmer le diagnostic n’etait presente que dans 18,3 % des biopsies (28,5 % des patients). Il nous a paru licite de proposer le diagnostic « compatible avec une MW » lorsqu’une ou deux de ces lesions histologiques etaient presentes (24,5 % des biopsies, 26 % des patients). Nos resultats montrent qu’il etait toujours preferable de biopsier les lesions macroscopiques lorsqu’elles existaient. En leur absence, il valait mieux s’orienter vers un prelevement de muqueuse sinusienne sous anesthesie generale. En effet, les nombreuses biopsies systematiques des fosses nasales realisees sous AL etaient non contributives dans 90 % des cas. Enfin, nous avons montre que la realisation de deux recoupes permettait d’augmenter la sensibilite de l’examen histologique de 7 %.

Mise au pointRéseau RENAPE : vers une nouvelle organisation des soins pour le traitement des tumeurs rares du péritoine. Description du réseau et rôle des pathologistesThe RENAPE network: Towards a new healthcare organization for the treatment of rare tumors of the peritoneum. Description of the network and role of the pathologists

As part of the national 2009-2013 Cancer Plan, and with the support of the National cancer Institute and the French ministry of health, the National network for the treatment of rare peritoneal malignancies (RENAPE) has been organized. Its main objective is to optimize the framework for the healthcare management and treatment of rare peritoneal malignancies. This specific organization covers the whole national territory including clinical expert and specialized structures and should lead to an appropriate treatment based on expertise and proximity. Within the RENAPE network, the RENA-PATH group gathers the pathologists actively involved in the management of rare peritoneal malignancies. The actions of RENA-PATH are focused primarily on the harmonization of pathological diagnostic criteria, reporting of new cases in the RENAPE registry and histology reviewing.

Endomicroscopy in bile duct: Inflammation interferes with pCLE applied in the bile duct: A prospective study of 54 patients

Background The preoperative diagnosis of biliary stenosis is associated with low accuracy. As a consequence, probe-based confocal laser endomicroscopy (pCLE), an in-vivo histological imaging technique, was applied in the bile duct. The aim of this study was to establish whether previous inflammation of the bile duct affects confocal interpretation. The findings from pCLE were compared in two groups of patients: those in whom there had been no cholangitis nor stenting and those in whom stents had been used and subsequently retrieved or who had suffered cholangitis. Patients and methods pCLE was performed on 54 patients (mean age 66 years; 31 men, 23 women) from September 2008 to July 2011. Patients were divided in two groups: group 1: 39 patients who had not undergone a biliary procedure in the month preceding the pCLE procedure; and group 2: 15 patients who had undergone stent placement or presented with cholangitis in the month preceding the pCLE procedure. Endoscopic and pCLE data were collected prospectively. pCLE results were compared to benchmark histology (surgery, endoultrasonography, percutaneous biopsy). Patients with a benign stricture who did not undergo operation were followed for 1 year. pCLE images of the bile duct were obtained during endoscopic retrograde cholangiopancreatography procedures. pCLE images were interpreted prospectively using the Miami classification in vivo and in real time. Results In group 1, sensitivity, specificity, and accuracy were 88, 83, and 87%, respectively. In group 2, sensitivity, specificity, and accuracy were 75, 71, and 73%, respectively. Diagnostic accuracy of pCLE was lower when applied to group 2 (p < 0,001). The investigation is less reliable in bile ducts affected by inflammation from cholangitis or previous stenting. Conclusions Inflammatory lesions of the bile duct interfere with interpretation of pCLE. A refined pCLE description of inflammatory lesions should improve accuracy of pCLE in bile duct stenosis.

Endoscopic ultrasound-guided needle-based confocal laser endomicroscopy in solid pancreatic masses.

BACKGROUND AND STUDY AIMS The differential diagnosis of solid pancreatic masses by endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) is currently suboptimal in centers that are not equipped with rapid on-site evaluation. Needle-based confocal laser endomicroscopy (nCLE) enables real-time in vivo microscopic imaging during endoscopy. This study aimed to describe nCLE interpretation criteria for the characterization of pancreatic masses, with histopathological correlation, and to perform the first validation of these criteria. PATIENTS AND METHODS A total of 40 patients were evaluated by EUS-FNA combined with nCLE for the diagnosis of pancreatic masses. Final diagnosis was based on EUS-FNA histology and follow-up at 1 year. Five unblinded examiners defined nCLE criteria for adenocarcinoma, chronic pancreatitis, and neuroendocrine tumor (NET) using a set of video sequences from 14 patients with confirmed pathology (Step 1). These criteria were retrospectively validated by four independent, blinded examiners using sequences from 32 patients (Step 2). RESULTS nCLE criteria were described for adenocarcinoma (dark cell aggregates, irregular vessels with leakages of fluorescein), chronic pancreatitis (residual regular glandular pancreatic structures), and NET (black cell aggregates surrounded by vessels and fibrotic areas). These criteria correlated with the histological features of the corresponding lesions. In the validation review, a conclusive nCLE result was obtained in 75 % of cases (96 % correct). Statistical evaluation provided promising results, with high specificity, and negative and positive predictive values for all types of pancreatic masses. CONCLUSION Considering the low negative predictive value of EUS-FNA, nCLE could help to rule out malignancy after a previous inconclusive EUS-FNA. Larger studies are required to confirm these findings and to establish the role of nCLE in the diagnosis of pancreatic masses. TRIAL REGISTRATION ClinicalTrials.gov (NCT01563133).

Overexpression of the Promigratory and Prometastatic PTK7 Receptor Is Associated with an Adverse Clinical Outcome in Colorectal Cancer

Biomarkers and novel therapeutic targets are urgently needed in colorectal cancer (CRC). The pseudo tyrosine kinase receptor 7 (PTK7) is involved in planar cell polarity and it is deregulated in various malignancies, including CRC. Yet, little is known about its protein expression in human CRC, or about a possible correlation of its expression with clinical endpoints. Using a clinically annotated Tissue MicroArray (TMA) produced from from 192 consecutive CRC patients treated by initial surgery, we examined PTK7 expression by immunohistochemistry in tumoral tissue and matched normal mucosae, and correlated its expression with clinico-pathological features and patient outcome. PTK7 depletion by specific shRNA in HCT116 and HCT15 CRC cell lines was found to affect cell proliferation, resistance to drugs and cell migration. Tumor growth and metastatic phenotype were investigated in vivo using a xenograft mouse model of CRC cells with modulated expression of PTK7 levels. PTK7 was significantly up-regulated in CRC tissue as compared to matched healthy mucosae, and significant overexpression was found in 34% of patients. PTK7 overexpression was significantly associated with a reduced metastasis-free survival in non-metastatic patients. In HCT116 and HCT15 cells, shRNA PTK7 reduced migration but did not affect cell proliferation and resistance to drugs. In a xenograft mouse of HCT15 cells, downregulation of PTK7 led to reduced tumor growth, whereas its overexpression in PTK7-negative cancer cells led to increased metastatic events. PTK7 expression thus represents a potential prognostic biomarker and a novel therapeutic target in CRC.