The diagnostic and prognostic role of liquid-based cytology: are we ready to monitor therapy and resistance?

Here, we evaluate the diagnostic and prognostic role of liquid-based cytology (LBC) in different body lesions, including thyroid, lung, effusions and malignant breast lesions. LBC has gained consensus after being applied to both non-gynecologic and fine-needle aspiration cytology. Although some remain sceptical regarding the diagnostic efficacy of LBC, mainly when used alone, in recent years, good results have been obtained as long as it showed a high diagnostic accuracy. Here, we discuss the additional possibility of storing material for the application of ancillary techniques (immunocytochemistry-molecular analysis) with several diagnostic and prognostic advantages, which may pave the way for the challenging evaluation of both monitoring responses to treatment and resistance to targeted therapies in thyroid, lung, breast carcinoma or malignant effusions. Furthermore, it provides the use of several molecular spots as specific targets for personalized therapy.

Evaluation of indeterminate thyroid cytology by second-opinion diagnosis or repeat fine-needle aspiration: which is the best approach?

OBJECTIVE: This study investigated a published series evaluating the role of second-opinion diagnosis (SOD) or repeat fine-needle aspiration cytology (RFNA) for indeterminate thyroid aspirates. STUDY DESIGN: Twenty-three studies were selected and the following parameters were analyzed: disagreement between SOD or RFNA and the original diagnosis (OD), reclassification of OD according to the Bethesda system for reporting thyroid cytopathology, the rate of definitive diagnosis and the diagnostic performance of SOD and RFNA. RESULTS: 7,154 thyroid FNAs were retrieved from 9 studies that investigated the role of SOD, including 1,048 (14.6%) cases originally reported as indeterminate. The 14 studies that analyzed the role of thyroid RFNA comprised 67,581 FNAs and included 7,246 (10.7%) indeterminate cases. A definitive diagnosis was achieved by SOD in 450 cases (42.9%) and RFNA in 1,645 cases (57.2%, p=0.0001). Based on cases with histological follow-up, SOD demonstrated significantly higher rates of positive predictive value and accuracy than RFNA (55.8 vs. 37.7%, p=0.0001; 67.4 vs. 56.0%, p=0.0034, respectively). CONCLUSIONS: Both SOD and RFNA demonstrated an improvement in the diagnosis of initially indeterminate thyroid FNAs. RFNA achieved a definitive diagnosis for the majority of indeterminate cases. Regarding histological follow-up, SOD was shown to be more accurate than RFNA.

Is thyroid gland only a "land" for primary malignancies? role of morphology and immunocytochemistry.

BACKGROUND: Thyroid Metastases (TM) represent a rare entity with an estimated variability ranging between 0 and 24%. Fine needle aspiration cytology (FNAC) might be useful in discriminating between primary and TM nodules especially when ancillary techniques (i.e., immunocytochemistry-ICC) are carried out. METHODS: We herein appraised a series of 20 TM on FNAC analyzed between 2000 and 2013. We included eight male and 12 female patients. The cytological cases were processed with both liquid based (LBC) and conventional cytology. RESULTS: We reported 2.2% TM out of 910 malignancies. Our TM cases resulted as: six lung (LG), five gastro-intestinal (GI), five breast (B), three larynx (LX), and one clear cell renal carcinoma (CCRC) metastases. All the patients had a previous known cancer history. Although the cytological features were likely to suspect a TM, the application of ICC panels was contributive in 100% cases. None of TMs resulted as a unique localization whereby two cases underwent total thyroidectomy (including one B and one CCRC) and 18 TM were treated with radio-chemotherapy approaches. CONCLUSIONS: FNAC empowered the diagnostic workup of patients with TM avoiding useless surgical approach. The low sensitivity of cytology might be reinforced by the application of ancillary techniques. In contrast with the reported predominant rate of kidney metastatic carcinomas, our findings underlined that intestinal cancer as well as lung and breast are the most common TM. TM are frequently multifocal and in a contest of a systemic disease so that a tailored therapy seems to be the best treatment.

The basal epithelial marker P-cadherin associates with breast cancer cell populations harboring a glycolytic and acid-resistant phenotype.

BACKGROUND: Cancer stem cells are hypoxia-resistant and present a preponderant glycolytic metabolism. These characteristics are also found in basal-like breast carcinomas (BLBC), which show increased expression of cancer stem cell markers.Recently, we demonstrated that P-cadherin, a biomarker of BLBC and a poor prognostic factor in this disease, mediates stem-like properties and resistance to radiation therapy. Thus, the aim of the present study was to evaluate if P-cadherin expression was associated to breast cancer cell populations with an adapted phenotype to hypoxia. METHODS: Immunohistochemistry was performed to address the expression of P-cadherin, hypoxic, glycolytic and acid-resistance biomarkers in primary human breast carcinomas. In vitro studies were performed using basal-like breast cancer cell lines. qRT-PCR, FACS analysis, western blotting and confocal microscopy were used to assess the expression of P-cadherin after HIF-1α stabilization, achieved by CoCl2 treatment. siRNA-mediated knockdown was used to silence the expression of several targets and qRT-PCR was employed to evaluate the effects of P-cadherin on HIF-1α signaling. P-cadherin high and low breast cancer cell populations were sorted by FACS and levels of GLUT1 and CAIX were assessed by FACS and western blotting. Mammosphere forming efficiency was used to determine the stem cell activity after specific siRNA-mediated knockdown, further confirmed by western blotting. RESULTS: We demonstrated that P-cadherin overexpression was significantly associated with the expression of HIF-1α, GLUT1, CAIX, MCT1 and CD147 in human breast carcinomas. In vitro, we showed that HIF-1α stabilization was accompanied by increased membrane expression of P-cadherin and that P-cadherin silencing led to a decrease of the mRNA levels of GLUT1 and CAIX. We also found that the cell fractions harboring high levels of P-cadherin were the same exhibiting more GLUT1 and CAIX expression. Finally, we showed that P-cadherin silencing significantly decreases the mammosphere forming efficiency in the same range as the silencing of HIF-1α, CAIX or GLUT1, validating that all these markers are being expressed by the same breast cancer stem cell population. CONCLUSIONS: Our results establish a link between aberrant P-cadherin expression and hypoxic, glycolytic and acid-resistant breast cancer cells, suggesting a possible role for this marker in cancer cell metabolism.

Liquid-based cytology in fine-needle aspiration of breast lesions: a review.

OBJECTIVE: Fine-needle aspiration (FNA) is a safe and cost-effective technique for the diagnosis of breast lesions, especially when correlated with clinical and imaging studies. However, the success of breast FNA is highly dependent on the adequate preparation of cytological conventional smears (CS). The liquid-based cytology (LBC) technique consists of an automated method for preparing thin-layer cytological samples from cell suspensions collected in alcohol-based preservative. LBC is designed to improve CS by avoiding limiting factors such as obscuring material, air-drying and smearing artifacts. STUDY DESIGN: We performed a review of the published literature about LBC applied to breast FNA. RESULTS: LBC preparations of breast aspirates demonstrated better cellular preservation, less cell overlapping and elimination of blood and excessive inflammation compared to CS. Conversely, alterations in architecture and cell morphology as well as loss of myoepithelial cells and stromal elements have been described in LBC specimens, requiring training before applying this technique for diagnosis. Studies have shown a similar accuracy between LBC and CS for the diagnosis of breast lesions. LBC also permits the use of residual material for ancillary tests, which is an important advantage compared to CS. CONCLUSIONS: LBC can be safely applied to breast FNA, showing a similar diagnostic accuracy to CS.

Topoisomerase II-alfa gene as a predictive marker of response to anthracyclines in breast cancer.

Amplification or deletion of the topoisomerase IIα (TOP2A) gene in breast cancer has been related with responsiveness to anthracyclines-based chemotherapy. The purpose of this study was to evaluate the predictive value of TOP2A gene for the efficacy of neo-adjuvant anthracycline in a population with locally advanced breast cancer. Sixty-two patients were included, and the status of TOP2A gene was determined by in situ hybridization method. Treatment efficacy was determined by clinical and pathological response and overall survival. TOP2A gene alterations were found in 22.6% (21.0% of cases with amplification and 1.6% with deletion), and these tumors were biologically more aggressive, with higher nuclear grade, more frequently with HER2 amplification and inflammatory type. Also in these tumors response to chemotherapy appeared to be increased. There was a higher clinical and pathological response rate (complete pathological response of 21.4% vs. 8.3%), a trend toward longer progression-free survival (82.51 vs. 63.12 months) and a trend to increased overall survival (92.08 months; 95% CI 82.81-101.35 vs. 73.40 months; 95% CI 63.44-83.36; p=0.113). These results corroborate that the TOP2A gene alterations may play an important role in determining anthracycline sensitivity in breast cancer.

Detection of common and less frequent EGFR mutations in cytological samples of lung cancer.

OBJECTIVE: Lung cancer represents the leading cause of cancer death. EGFR mutations, detected in 10-40% of lung adenocarcinomas, are an essential key to therapeutic management. EGFR-activated mutations comprise mainly deletions in exon 19 and point mutations in exon 21. Although histology is the traditional method of detection, we investigated the role of cytology in EGFR mutations. STUDY DESIGN: A total of 774 lung cancers were studied for EGFR mutations (676 histological and 98 cytological samples), including 424 adenocarcinomas, 326 non-small cell lung carcinomas not otherwise specified, and 24 squamous cell carcinomas. RESULTS: We had a total of 164 (21.2%) cases of mutations. Common mutations were short in-frame deletions in exon 19 (53.7%) and single-nucleotide substitutions in exon 21 (34.1%); less frequent mutations included single-nucleotide substitutions in exon 18 (3.7%) and in-frame insertions/deletions in exon 20 (8.5%). Histologically, EGFR mutations in exons 19 and 21 occurred in 19.4% and in exons 18 and 20 in 2.2%, while the rates cytologically were 13.3% for exons 19 and 21 and 5.1% for exons 18 and 20. CONCLUSIONS: The sensitivity for the detection of EGFR mutations in cytological samples overlaps histology, so the use of cytological material constitutes an adequate approach for treatment selection in patients with locally advanced or metastatic lung cancer.

High diagnostic accuracy and reproducibility of fine-needle aspiration cytology for diagnosing salivary gland tumors: cytohistologic correlation in 182 cases.

OBJECTIVE: The purpose of this study was to assess the efficacy and reproducibility of the cytologic diagnosis of salivary gland tumors (SGTs) using fine-needle aspiration cytology (FNAC). The study aimed to determine diagnostic accuracy, sensitivity, and specificity and to evaluate the extent of interobserver agreement. STUDY DESIGN: We retrospectively evaluated SGTs from the files of the Division of Pathology at the Clinics Hospital of São Paulo and Piracicaba Dental School between 2000 and 2006. RESULTS: We performed cytohistologic correlation in 182 SGTs. The sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy were 94%, 100%, 100%, 100%, and 99%, respectively. The interobserver cytologic reproducibility showed significant statistical concordance (P < .0001). CONCLUSIONS: FNAC is an effective tool for performing a reliable preoperative diagnosis in SGTs and shows high diagnostic accuracy and consistent interobserver reproducibility. Further FNAC studies analyzing large samples of malignant SGTs and reactive salivary lesions are needed to confirm their accuracy.

The value of second opinion in thyroid cytology: a review.

BACKGROUND: Second-opinion diagnosis (SOD) on pathological material is an accepted practice before definitive therapy is considered for referred patients. The thyroid gland is an anatomical site prone to diagnostic disagreement between pathologists. We performed a review of the literature that addressed the role of interinstitutional SOD on thyroid fine-needle aspirations (FNAs). METHODS: Nine studies comprising second opinions on thyroid FNAs were selected. The parameters analyzed included: discordances between the initial diagnoses (IDs) and SODs; cytohistologic correlation; changes in the clinical management of the patients with thyroid nodules after SOD. The same parameters were applied to the "indeterminate" diagnostic category comprising cases initially reported as "atypia," "atypia of undetermined significance/follicular lesion of undetermined significance," "suspicious for a follicular neoplasm," "follicular neoplasm," "suspicious," and "suspicious for malignancy." RESULTS: A total of 7154 thyroid FNAs were retrieved, showing an overall discordance rate between ID and SOD of 28.6%. In general, SOD was better supported by clinical follow-up and histological diagnosis, showing higher diagnostic accuracy in comparison with ID. Almost one-third (30.4%) of the discordant cases resulted in changes in the clinical management of patients with thyroid nodules. Numerous thyroid FNAs initially categorized as "indeterminate" were definitively classified as benign or malignant by SOD, with an overall diagnostic resolution rate of 42.5%, sensitivity of 97.9%, and diagnostic accuracy of 73.7%. CONCLUSIONS: Second-opinion review of thyroid FNA improves diagnostic accuracy and potentially changes clinical management. SOD also demonstrates a significative rate of diagnostic resolution for thyroid FNAs originally diagnosed as "indeterminate."

Putting an eye on cytological specimens: an audit of the clinical impact of thyroid fine-needle aspiration in different health care settings.

There is published evidence showing less cost-benefit approaches in the evaluation of thyroid nodules. We performed an institutional audit of the cytologic diagnosis of thyroid fine-needle aspiration (FNA) in an attempt to perceive the clinical impact of this technique on the management of thyroid nodules and to compare it in two different types of health care: Primary Care Medicine and Endocrinology. We performed a retrospective analysis to the electronic records of patients referred from General Practitioners (GP) and Endocrinologists (E) for thyroid FNA between 2010 and 2012. Request forms for cytological reports where retrieved for analysis of clinical and cytological data. The database search retrieved 1655 patients (female gender: 88.2%; GP references: 51.8%). Preprocedure clinical information was available from 157 out of 2005 nodules (7.8%). Significant differences in cytological diagnosis were seen in "Nondiagnostic" (GP: 11.6%; E: 7.5%, χ(2) = 0.002) and "Benign" categories (GP: 75%; E: 81.8%, χ(2) < 0.001). The main potential cause of "Nondiagnostic" samples was nodules smaller than one centimeter (total: 14 cases; GP: 7; E: 7). Reasons to request FNA for these nodules were provided in 6 out of 27 cases (GP: 0/16; E: 6/11, P < 0.001). The rate of insufficient samples was inversely correlated with nodule size (τ = -0.242, P = 0.001). When evaluating thyroid nodules, clinicians should take into account the limitations of FNA, the international recommendations for better cost-benefit approaches and the importance of a well-informed cytopathologist for better cytological diagnostic results.

Analysis of nondiagnostic results in a large series of thyroid fine-needle aspiration cytology performed over 9 years in a single center.

OBJECTIVE: Thyroid fine-needle aspiration cytology (FNAC) is the most valuable, cost-effective and accurate method for the evaluation of patients with thyroid nodules. One of its limitations is that up to 20% of results are nondiagnostic or unsatisfactory. The aim of this study was to analyze the number of thyroid FNAC specimens with nondiagnostic results obtained on an outpatient basis and how many of these had to be repeated according to their results. STUDY DESIGN: This was a retrospective analysis of diagnostic reports of nondiagnostic thyroid FNAC specimens obtained between 1 January 2004 and 31 December 2012 which were retrieved by means of a computerized search. The FNAC results and the age and sex of the patients were collected. RESULTS: From a total of 15,292 thyroid FNAC specimens, 6.8% (n = 1,033) corresponded to nondiagnostic cases. Eligible diagnostic reports for analysis included 877 cases (106 were repetitions of previous nondiagnostic FNAC). After an initial nondiagnostic finding for 771 FNAC smears, 29.5% (n = 225) were repeated with the following results: 43.6% insufficient, 49.3% benign, 6.2% follicular neoplasm, 0.4% suspicious for malignancy and 0.4% malignant. Twenty-two patients underwent a second repeated FNAC. Here the findings were: 36.4% insufficient, 59.1% benign, 4.5% follicular neoplasm, 0.0% suspicious for malignancy and 0.0% malignant. CONCLUSIONS: There was a low rate of repeated FNAC among the group of nondiagnostic cases. With repeated FNAC, the rate of nondiagnostic cases and the number of results that potentially demand surgery diminish.

P-cadherin signals through the laminin receptor α6ß4 integrin to induce stem cell and invasive properties in basal-like breast cancer cells.

P-cadherin is a classical cell-cell adhesion molecule that, in contrast to E-cadherin, has a positive role in breast cancer progression, being considered a poor prognostic factor in this disease. In previous reports, we have shown that this protein induces cancer stem cell and invasive properties to basal-like breast cancer cells. Here, we clarify the downstream signaling pathways that are triggered by P-cadherin to mediate these effects. We demonstrated that P-cadherin inhibition led to a significant decreased adhesion of cancer cells to the basement membrane substrate laminin, as well as to a major reduction in the expression of the laminin receptor α6ß4 integrin. Remarkably, the expression of this heterodimer was required for the invasive capacity and increased mammosphere forming efficiency induced by P-cadherin expression. Moreover, we showed that P-cadherin transcriptionally up-regulates the α6 integrin subunit expression and directly interacts with the ß4 integrin subunit. We still showed that P-cadherin downstream signaling, in response to laminin, involves the activation of focal adhesion (FAK), Src and AKT kinases. The association between the expression of P-cadherin, α6ß4 heterodimer and the active FAK and Src phosphorylated forms was validated in vivo. Our data establish that there is a crosstalk between P-cadherin and the laminin receptor α6ß4 integrin signaling pathway, which link has never been previously described. The activation of this heterodimer explains the stem cell and invasive properties induced by P-cadherin to breast cancer cells, pointing to a new molecular mechanism that may be targeted to counteract the effects induced by this adhesion molecule.

Clinicopathological significance of ERCC1 expression in breast cancer.

The excision repair cross-complementation 1 (ERCC1) enzyme plays an essential role in the nucleotide excision repair pathway and is associated with resistance to platinum-based chemotherapy in different types of cancer. The aim of the present study was to evaluate the clinicopathological significance of ERCC1 expression in breast cancer patients. We analyzed the immunohistochemical expression of ERCC1 in a tissue microarray from 135 primary breast carcinomas and correlated the immunohistochemical findings with clinicopathological factors and outcome data. ERCC1 expression analysis was available for 109 cases. In this group, 58 (53.2%) were positive for ERCC1. ERCC1-positive expression was correlated with smaller tumor size (P=0.007) and with positivity for estrogen receptor (P=0.040), but no correlation was found with other clinicopathological features. Although not statistically significant, triple negative breast cancers were more frequently negative for ERCC1 (61.5% of the cases) compared to the non-triple negative breast cancer cases (41.5%). In conclusion, ERCC1 expression correlated significantly with favorable prognostic factors, such as smaller tumor size and ER-positivity, suggesting a possible role for ERCC1 as a predictive and/or prognostic marker in breast cancer.

Thyroid fine-needle aspiration cytology: is there a place to virtual cytology?

Telecytology has been used for education, training, and consultation. Cytological studies from gynecological, nongynecological and fine-needle aspiration cytology (FNAC) specimens (including studies of thyroid FNAC) analyzed the diagnostic accuracy and reproducibility of telecytology-based predominantly on static digital images. The aim of this study was to evaluate the diagnostic reproducibility of virtual cytology by measuring intraobserver and interobserver agreements among two cytopathologists, using the Bethesda System for Reporting Thyroid Cytopathology (BSRTC) nomenclature. 502 glass slides from 222 cases of thyroid FNAC were retrieved and scanned by a high-resolution scanner generating whole slides images (virtual cytology). Conventional and virtual cytology were analyzed by a skilled cytopathologist and the intraobserver agreement rate was 77.5% with the corresponding κ value of 0.54, suggesting a moderate agreement between both methods. A second cytopathologist analyzed the same slides only by virtual cytology and the interobserver agreement rate was 80.2% with the corresponding κ value of 0.57, suggesting a moderate agreement between both cytopathologists. The virtual cytology resulted in a higher proportion of aspirates classified as nondiagnostic (20.3 and 14.9% for the first and second cytopathologist, respectively) as compared to conventional cytology (8.1%). Regarding specific diagnostic categories as defined by the BSRTC nomenclature, the follicular lesion of undetermined significance category presented the lowest concordance rates, corresponding to 5.9% intraobserver agreement and no (0.0%) interobserver agreement. We suggest that virtual cytology can be an alternative to conventional cytology in assessment of thyroid FNAC specimens, but nondiagnostic aspirates obtained by virtual cytology should be reassessed by conventional cytology.

Cancer stem cells markers CD44, CD24 and ALDH1 in breast cancer special histological types.

AIMS: CD44, CD24 and ALDH1 are the most consistently used biomarkers to identify and characterise the breast cancer stem cell (CSC) phenotype. However, most studies performed until now analysed samples of invasive ductal carcinomas of no special type (IDC-NST). Therefore, prevalence and clinical significance of these CSC markers in breast carcinomas of special histological types (SHT) is largely unknown. For that reason, this study aims to determine the distribution of the breast CD44, CD24 and ALDH1 CSC markers among a series of invasive breast carcinomas of SHT, in comparison with a series of IDC-NST. METHODS: 117 invasive SHT breast carcinomas were analysed for the expression of CD44, CD24 and ALDH1, by immuhohistochemistry. The distribution of these CSC markers was evaluated among the distinct histological special types, and the results were compared with a series of 466 IDC-NST. RESULTS: The expression prevalence of the breast CSC markers differed between special types and IDC-NST. Medullary, papillary and tubular carcinomas were enriched in the CSC phenotype CD44(+)/CD24(-/low) (80.0%, 100.0% and 100.0%, respectively, vs 45.3% in IDC-NST). Considering the ALDH1 cytoplasmic tumour expression, only medullary and metaplastic carcinomas displayed significant increase in CD44(+)/CD24(-/low)/ALDH1(+) CSC phenotype frequency (36.4% and 28.6%, respectively, vs 4.8% in IDC-NST). CONCLUSIONS: The expression distribution of breast CSC markers is largely dependent on histological type. Interestingly, within the distinct SHT, medullary and metaplastic carcinomas are the two types highly associated with high-grade carcinomas, basal-like and claudin-low molecular subtypes, and to the CSC phenotype CD44(+)/CD24(-/low)/ALDH1(+).

P-cadherin functional role is dependent on E-cadherin cellular context: a proof of concept using the breast cancer model.

P-cadherin overexpression is associated with worse breast cancer survival, being a poor prognostic marker as well as a putative therapeutic target for the aggressive triple-negative and basal-like carcinomas (TNBCs). Previously, we have shown that P-cadherin promotes breast cancer invasion of cells where membrane E-cadherin was maintained; however, it suppresses invasion in models without endogenous cadherins, like melanomas. Here, we investigated if P-cadherin expression would interfere with the normal adhesion complex and which were the cellular/molecular consequences, constituting, in this way, a new mechanism by which E-cadherin invasive-suppressor function was disrupted. Using breast TNBC models, we demonstrated, for the first time, that P-cadherin co-localizes with E-cadherin, promoting cell invasion due to the disruption caused in the interaction between E-cadherin and cytoplasmic catenins. P-cadherin also induces cell migration and survival, modifying the expression profile of cells expressing wild-type E-cadherin and contributing to alter their cellular behaviour. Additionally, E- and P-cadherin co-expressing cells significantly enhanced in vivo tumour growth, compared with cells expressing only E- or only P-cadherin. Finally, we still found that co-expression of both molecules was significantly correlated with high-grade breast carcinomas, biologically aggressive, and with poor patient survival, being a strong prognostic factor in this disease. Our results show a role for E- and P-cadherin co-expression in breast cancer progression and highlight the potential benefit of targeting P-cadherin in the aggressive tumours expressing high levels of this protein.

Differentiated thyroid carcinomas may elude the immune system by B7H1 upregulation.

B7H1 is consistently associated with inhibition of the immune system in many solid tumors. However, there is no report about its impact on differentiated thyroid carcinoma (DTC) presentation, aggressiveness, or evolution. Aiming to investigate the role of B7H1 in DTC and correlate this protein with other tumor-infiltrating immune cells, we studied 407 thyroid nodule tissue samples including 293 from DTC patients, all managed according to a same standard protocol. In addition, we obtained 5 normal and 114 benign thyroid lesions. Eighteen out of the 253 papillary thyroid carcinomas were paired with respective metastatic lymph node tissues. B7H1 (CD274) protein expression was assessed by immunohistochemistry and the gene expression was quantified by real-time PCR. Malignant tissues displayed a more intense B7H1 staining and higher mRNA levels than benign tissues (both P<0.0001). We observed a positive linear correlation between higher age at diagnosis and B7H1 mRNA levels (P=0.02896). Elevated levels of B7H1 protein were associated with the presence of CD4+, CD8+, CD20+, and FoxP3+ lymphocytes (all P<0.05); tumor-associated macrophages (P<0.0001); and the presence of myeloid-derived suppressor cells (P=0.03256). Stage II-IV patients presented higher B7H1 mRNA levels than stage I cases (P=0.03522). On the contrary, a decreased expression of B7H1 protein was observed in lymph node metastasis (P=0.0152). In conclusion, our data demonstrate that B7H1 expression is associated with features of aggressiveness, suggesting that this is an immune evasion mechanism of DTC cells.

Claudin expression in breast cancer: high or low, what to expect?

The evaluation of claudins (CLDNs) expression pattern in tumours can be important to understand breast carcinogenesis. The study of CLDNs became more appealing since it was found that CLDN3 and CLDN4 are putative therapeutic targets for Clostridium perfrigens enterotoxin (CPE), as well as for monoclonal antibody-based therapy. Moreover, the recently characterized CLDN-low molecular subgroup of breast tumours increased the interest in these molecules. Based on these facts, our aim was to explore the pattern of expression of CLDNs among a large series of invasive breast carcinomas. We also analysed the correlation between the combinatorial expression of CLDN3/CLDN4 and classical prognostic factors and biological markers. In addition, we also compared the characteristics of tumours with low expression of CLDN3, CLDN4 and CLDN7, assessed by immunohistochemistry (IHC), and the ones from CLDN-low subgroup of tumours previously defined by genomic assays. The combinatorial analysis of the expression of CLDN3/CLDN4 showed a significant association between high CLDN3/CLDN4 levels and triple-negative tumours, as well as with worse patient outcome. This combined analysis may provide useful information for breast carcinomas, since these two CLDN members are putative therapeutic targets. Comparing tumours with low expression of CLDN3, CLDN4 and CLDN7 with tumours previously referred to as CLDN-low by genomic assays, we demonstrated that the single IHC evaluation of these three specific CLDNs is insufficient to identify the CLDN-low molecular subtype of breast tumours. The analysis of several other molecular markers, such as EMT (epithelial-to-mesenchymal transition) and CSC (cancer stem cell) markers should probably be added to improve the identification of this subgroup of tumours by IHC, which probably are enriched in carcinomas with metaplastic differentiation.

Infiltration of a mixture of immune cells may be related to good prognosis in patients with differentiated thyroid carcinoma.

OBJECTIVE: Immune responses against differentiated thyroid carcinomas (DTC) have long been recognized. We aimed to investigate the role of immune cell infiltration in the progression of DTC. DESIGN: We studied 398 patients - 253 with papillary and 13 with follicular thyroid cancers, as well as 132 with nonmalignant tissues. PATIENTS AND MEASUREMENTS: Immune cell infiltration was identified using CD3, CD4, CD8, CD20, CD68 and FoxP3 immunohistochemical markers. In addition, we assessed colocalization of CD4 and IL-17 to identify Th17 lymphocytic infiltration and colocalization of CD33 and CD11b to identify infiltration of myeloid-derived suppressor cells (MDSC). RESULTS: Immune cells infiltrated malignant tissues more often than benign lesions. The presence of chronic lymphocytic thyroiditis (CLT) concurrent to DTC, CD68+, CD4+, CD8+, CD20+, FoxP3+ and Th17 lymphocytes but not MDSCs was associated with clinical and pathological features of lower tumour aggressiveness and a more favourable patient outcome. A log-rank test confirmed an association between concurrent CLT, tumour-associated macrophage infiltration, and CD8+ lymphocytes and an increased in disease-free survival, suggesting that evidence of these immune reactions is associated with a favourable prognosis. CONCLUSION: Our data suggest that the tumour or peri-tumoural microenvironment may act to modify the observed pattern of immune response. Immune cell infiltration and the presence of concurrent CLT helped characterize specific tumour histotypes associated with favourable prognostic features.