CD160 Expression in Retinal Vessels Is Associated With Retinal Neovascular Diseases.

Purpose: Anti-angiogenic agents stand first in the treatment of neovascular diseases of the retina. CD160 appeared in several experimental studies as a marker of activated endothelial cells, suggesting it could represent a promising target for novel anti-angiogenic therapies. The aim of the present study was to assess the distribution of CD160 in the human eye, and to search for a possible correlation with retinal neovascular diseases. Methods: The physiological distribution of CD160 in the normal eye was assessed with immunolabeling in 10 human donor eyes. Then, in a retrospective cohort of 75 surgical retinal specimens, the density of CD160+ microvessels was evaluated, along with immunolabeling on serial sections against ERG (pan-endothelial cell marker), CD105 (activated endothelial cell marker), and α-SMA (pericyte cell marker). The cohort was divided into two groups: 29 patients with neovascular disease (NV+) and 46 control patients (NV-). Results: CD160 was physiologically expressed by several cell types: endothelial cells of retinal blood vessels, ganglion cells, macrophages, epithelial cells of the conjunctiva, ciliary body, and retinal pigment epithelium. In the patient cohort, the percentage of CD160+ vessels in the retina was significantly and independently higher in patients suffering from neovascular diseases (P = 0.04). On the contrary, the expression of CD105 was correlated neither with retinal neovascular diseases, nor with CD160 expression. Conclusions: CD160 was expressed in some retinal vessels in both normal and pathologic eyes. CD160 expression by endothelial cells of retinal vessels was correlated with ocular neovascular diseases. CD160 could therefore represent an interesting target for novel anti-angiogenic therapies.

Triple-negative and HER2-overexpressing breast cancer cell sialylation impacts tumor microenvironment T-lymphocyte subset recruitment: a possible mechanism of tumor escape.

INTRODUCTION: Breast cancers develop different patterns of sialylation to modulate their tumor-infiltrating lymphocyte (TIL) environment. We studied the relationship between α-2,6 sialyltransferases and the TIL in different breast cancer molecular subgroups. MATERIALS AND METHODS: Immunohistochemical preparations were made from 39 luminal (LUM), 13 human epidermal growth factor receptor 2-overexpressing (HER2) and 47 triple-negative (TN) breast carcinomas. Targeted proteins included ST6Gal-I, ST6Gal-II, ST6GalNac-I, CD8, CD4 and granzyme-B in both cytotoxic T lymphocytes and NK lymphocytes (CTL/NK). RESULTS: CTL/NK populations were significantly more frequent in TN than LUM (P <0.001). TN showed a lower level of ST6Gal-I expression than LUM or HER2 (both P > 0.001). ST6GalNac-I expression was lower in LUM than in TN or HER2 (P = 0.002 and P = 0.02, respectively). In HER2, a significant association was found between a low level of ST6Gal-I expression and a high TIL level. In TN, a significant association was observed between a high level of ST6Gal-II expression and a high TIL level. CONCLUSION: An increase in infiltrating lymphocytes could be influenced by low expression of ST6Gal-I in HER2 and by high expression of ST6Gal-II in TN breast cancers. Thus, targeting these sialylation pathways could modulate the levels of TIL.

Chemotherapy treatment induces an increase of autophagy in the luminal breast cancer cell MCF7, but not in the triple-negative MDA-MB231.

Autophagy is one of the chemotherapy resistance mechanisms in breast cancer. The aim of this study was to determine the level of recruitment of the autophagy pathway in the triple-negative breast cancer (TNBC) cell line MDA-MB231 compared with that in the control luminal breast cancer cell line MCF7 before and after treatment with chemotherapy drugs. Furthermore, we investigated the relationship between autophagy and EGFR, MUC1 and IL17-receptors as activators of autophagy. Immunohistochemistry was performed in cell culture blocks using LC3b, MUC1-C, EGFR, IL17A, IL17-RA and IL17-RB antibodies. We found that the basal autophagy level in MDA-MB231 was high, whereas it was low in MCF7. However, in contrast to MDA-MB231, the autophagy level was increased in MCF7 upon treatment with chemotherapy agents. Interestingly, we observed that the expression levels of MUC1-C, EGFR, IL17-RA, and IL17-RB were not modified by the same treatments. Furthermore, the chemotherapy treatments did not increase autophagy in TNBC cells without affecting the expression levels of MUC1-C, EGFR, IL17-RA or IL17-RB.

Autophagy is decreased in triple-negative breast carcinoma involving likely the MUC1-EGFR-NEU1 signalling pathway.

Triple-negative breast carcinoma (TN) is a heterogeneous cancer type expressing EGFR in 75% of cases. MUC1 is a large type I sialylated glycoprotein comprising two subunits (α and ß chains, also called respectively MUC1-VNTR and MUC1-CT), which was found to regulate EGFR activity through endocytic internalisation. Endocytosis and autophagy use the lysosome pathway involving NEU1. Recently, a molecular EGFR-MUC1-NEU1 complex was suggested to play a role in EGFR pathway. In the aim to understand the relationship between EGFR-MUC1-NEU1 complex and autophagy in breast carcinoma, we compared triple negative (TN) showing a high-EGFR expression with luminal (LUM) presenting low-EGFR level. We studied the expression of MUC1-VNTR, MUC1-CT and NEU1 in comparison with those of two molecular actors of autophagy, PI3K (p110ß) and Beclin1. A total of 87 breast cancers were split in two groups following the immunohistochemical classification of breast carcinoma: 48 TN and 39 LUM. Our results showed that TN presented a high expression of EGFR and a low expression of MUC1-VNTR, MUC1-CT, NEU1, Beclin-1 and PI3Kp110ß. Moreover, in TN, a positive statistical correlation was observed between Beclin-1 or PI3Kp110ß and MUC1-VNTR or NEU1, but not with EGFR. In conclusion, our data suggest that autophagy is reduced in TN leading likely to the deregulation of EGFR-MUC1-NEU1 complex and its associated cellular pathways.

The human epidermal growth factor receptor 2 screening tests for breast cancer suggested by the new updated recommendation of the american society of clinical oncology/college of american pathologists will involve a rise of the in-situ hybridization tests for the European laboratories of pathology.

Aims. The differences between the 2007 and the 2013 ASCO/CAP HER2 guidelines have been compared. We also discussed the potential consequences in our pathological practice. Material and Methodology. 189 HER2 fluorescence in situ hybridisation (FISH) tests were performed from 1016 preliminary HER2 immunohistochemical tests (IHC). All cases were reviewed and reclassed following the 2007 and 2013 ASCO/CAP recommendations. Results. The 2013 version decreased false-negative IHC (3/118 versus 1/54, P = ns) and created more 2+ IHC (40/186 versus 89/186, P = 0.001) or more 3+ IHC (9/186 versus 39/186, P = 0.001). One false-positive IHC was described for the 2013 version (0/9 versus 1/39, P = ns). Equivocal FISH was reduced (8/186 versus 2/186, P = ns). An estimation based on our data for 1000 patients showed a rise of our FISH tests for the control of 2+ IHC (180 tests for the 2007 version versus 274 tests for the 2013 version or FISH work overflow is +52%) and for the control of 2+/3+ IHC (300 for the 2007 version versus 475 for the 2013 version or FISH work overflow is +58%). Conclusions. The new 2013 ASCO/CAP guidelines have detected more HER2 positive cases but have increased the number of FISH tests.

IL-17A is produced by breast cancer TILs and promotes chemoresistance and proliferation through ERK1/2.

The proinflammatory cytokine Interleukin 17A (hereafter named IL-17A) or IL-17A producing cells are elevated in breast tumors environment and correlate with poor prognosis. Increased IL-17A is associated with ER(-) or triple negative tumors and reduced Disease Free Survival. However, the pathophysiological role of IL-17A in breast cancer remains unclear although several studies suggested its involvement in cancer cell dissemination. Here we demonstrated that a subset of breast tumors is infiltrated with IL-17A-producing cells. Increased IL-17A seems mainly associated to ER(-) and triple negative/basal-like tumors. Isolation of tumor infiltrating T lymphocytes (TILs) from breast cancer biopsies revealed that these cells secreted significant amounts of IL-17A. We further established that recombinant IL-17A recruits the MAPK pathway by upregulating phosphorylated ERK1/2 in human breast cancer cell lines thereby promoting proliferation and resistance to conventional chemotherapeutic agents such as docetaxel. We also confirmed here that recombinant IL-17A stimulates migration and invasion of breast cancer cells as previously reported. Importantly, TILs also induced tumor cell proliferation, chemoresistance and migration and treatment with IL-17A-neutralizing antibodies abrogated these effects. Altogether these results demonstrated the pathophysiological role of IL-17A-producing cell infiltrate in a subset of breast cancers. Therefore, IL-17A appears as potential therapeutic target for breast cancer.

Muc1/Cd227 immunohistochemistry in routine practice is a useful biomarker in breast cancers.

Over-expression of MUC1/CD227 is observed in 90% of breast tumors. Classical morphologic description and semi-quantitative digital measurement of MUC1 were performed from immunohistochemical stained slides of 123 routine histological samples. Measures of MUC1 expression showed statistical differences between non tumoral (NT) breast tissue and Ductal Carcinoma In Situ (DCIS) or infiltrating carcinoma (IC), p < 0.0001. Loss of MUC1 was correlated with high Ki67 index (p = 0.001) and loss of hormonal receptors (p = 0.03), whereas no correlations were found with HER2 expression. High-grade DCIS or IC showed increasing loss of apical polarised and cytoplasmic expression of MUC1.

Expression of MUC1 (Ma695) in noninvasive papillary urothelial neoplasm according to the 2004 World Health Organization classification of the noninvasive urothelial neoplasm. An immunologic tool for the pathologist?

OBJECTIVE: To examine the association between MUC1 (Ma695) and noninvasive papillary urothelial neoplasm according to the 2004 World Health Organization classification. STUDY DESIGN: Histologic evaluation was performed in 46 patients with nontumoral benign bladder urothelium (n = 11), papillary urothelial neoplasm of low malignant potential (PUNLMP) (n = 14), low-grade papillary urothelial carcinoma (LgPUC) (n = 11), and noninvasive high-grade papillary urothelial carcinoma (HgPUC) (n = 10). Classical morphologic description and semiquantitative digital measurement were performed from immunohistochemical-stained slides using an anti-MUC1 antibody (Ma695). RESULTS: Measures showed an obvious statistical difference between benign urothelium and LgPUC (p = 0.0004) or HgPUC (p = 0.0041). MUC1 is expressed less in PUNLMP than in LgPUC (p = 0.04). Benign urothelium and PUNLMP more often showed apical and superficial MUC1 expression. Basal cells with cytoplasmic and/or circumferential membrane positivity were more often observed in LgPUC and HgPUC (p < 0.001). CONCLUSION: MUC1 could be an interesting tool for the pathologist to differentiate between PUNLMP and LgPUC or HgPUC.

Fine needle aspiration cytology of lymph node: experience of 2 university hospitals with conventional smears and liquid-based cytology.

OBJECTIVE: To compare 2 different series of fine needle aspirations (FNA) performed by conventional smears (CS) and liquid-based cytology (TriPath PREP). STUDY DESIGN: From January 1, 2001, to December 31, 2005, we selected 139 FNA samples of lymph node, if the diagnosis was histologically proven after the initial cytologic diagnosis. Samples came from 2 university hospitals from Brussels: UZ-Brussel (n=96) using liquid-based cytology (LBC) and Hospital Erasme ULB (n=43) using CS. RESULTS: The number of inadequate samples was greater for LBC than CS, but there was no statistical difference (17.7% vs. 4.7%, p = 0.059). No differences were found between LBC and CS in sensitivity (respectively, 85.0% vs. 85.2%), specificity (84.2% vs. 85.7%) and efficiency (84.8% vs. 85.4%). CONCLUSION: Despite the cost, the efficiency of lymph node FNA cytology is identical between the CS and LBC performed by the TriPath PREP system. The quality of the smears, both LBC and CS, depends mainly on practitioner dexterity. Nevertheless, one advantage of LBC is that it provides an ancillary technique to improve FNAC diagnosis. In the future, a combination of these technologies with FNAC alone could in some cases take the place of surgical lymph node excision.

Expression of MUC1 and sialyl-Tn in benign prostatic glands, high-grade prostate intraepithelial neoplasia and malignant prostatic glands: a preliminary study.

OBJECTIVE: To postulate a possible role of MUC1 and sialylated mucins (sTn) in prostate malignancy. STUDY DESIGN: One sample histologic paraffin block of 24 radical prostatectomies was selected for presence of both benign and malignant glands; 17 samples also included PIN. Serial cuts were stained with MUC1 and sTn mouse monoclonal antibodies. Description and percentage of cell expression for MUC1 and sTn antibodies were obtained by light microscopy. RESULTS: MUC1 immunostaining was more often positive for neoplastic cells, progressively from benign (4.7+/-5.3%) to PIN (27.1+/-19.7%) and malignant glands (34.5+/-28.4%). The same observation was made for sTn, respectively, from 3.9+/-4.9% to 54.8+/-26.1% and 62.8+/-25.6%. Both antibodies showed a statistical difference between benign glands and PIN or malignant glands but not between PIN and malignant. CONCLUSION: MUC1 and sTn were expressed more intensely in PIN and malignant prostate glands than in benign glands. sTn seemed more specific in favor of cell malignancy.

Evaluation of combined general primer-mediated PCR sequencing and type-specific PCR strategies for determination of human papillomavirus genotypes in cervical cell specimens.

A strategy combining human papillomavirus general primer (mainly the PGMY primers)-directed PCR sequencing and type-specific PCR is presented. DNA samples were first tested in general primer-mediated PCR. The amplified fragments of positive samples after ethidium bromide-stained DNA gel analysis were further sequenced, and corresponding DNA samples were further analyzed by PCR using type-specific primers for human papillomavirus (HPV) types 16, 18, 31, and 52. The comparison of the results of 157 samples analyzed by this strategy in parallel with the Hybrid Capture 2 tests and with the HPV INNO-LiPA (Innogenetics line probe assay) shows that this method is suitable for HPV detection and genotyping in cervical cell samples. Although the PCR sequencing method is as sensitive as the HPV INNO-LiPA for HPV detection, our method allows the identification of a broader range of HPV types. In contrast, the HPV INNO-LiPA was less time-consuming and better identified coinfections.

Efficiency of an inexpensive liquid-based cytology performed by cytocentrifugations: a comparative study using the histology as reference standard.

BACKGROUND: Although liquid-based cytology (LBC) is now recommended for cervical cancer screening, it requires expensive automated devices and materials. To evaluate the efficiency of inexpensive LBC methods relying on an inexpensive fixative liquid, Easyfix, we compared the results obtained by the liquid-based cytology (LBC) diagnoses performed by cytocentrifugations (Papspin and Turbitec) with those obtained by histology. Furthermore, we evaluated the efficiency of the fixative liquid, Easyfix, to preserve HPV DNA in the collected samples. METHOD: 266 LBC were compared with 174 colposcopies and 91 Loop Electrosurgical Excision Procedure (LEEP). Among the LBC, 51 were performed using the Papspin system and 215 were performed using the Turbitec system. To control the quality of the preservation liquid, Easyfix, we correlated the results of HCII assays with those of HPV PCR. RESULTS: For Papspin and Turbitec systems, the sensitivities were respectively 82.6% (95% CI: 61.2-95.0%, p < 0.001) and 75.0% (95% CI: 64.4-89.8%, p < 0.001) and the specificities were 92.6% (95%CI: 76.5-99.1%, p < 0.001) and 96.2% (95% CI: 91.3-98.7%, p < 0.001). We find no statistical difference between the results of the both systems (p = ns). The sensitivity of the HCII was 86.4% (95% IC: 77.4-92.8%, p < 0.001) and the specificity was 39.4% (95% CI: 31.2-48.1%, p < 0.001). The comparison between HCII and HPV-PCR shows a good correlation: the kappa was 0.89. CONCLUSION: LBC performed by cytocentrifugations are inexpensive, reduce inadequate smears, show excellent efficiency and allow HPV detection by molecular biology.