Detection of Measurable Residual Disease Biomarkers in Extracellular Vesicles from Liquid Biopsies of Multiple Myeloma Patients-A Proof of Concept.

Monitoring measurable residual disease (MRD) is crucial to assess treatment response in Multiple Myeloma (MM). Detection of MRD in peripheral blood (PB) by exploring Extracellular Vesicles (EVs), and their cargo, would allow frequent and minimally invasive monitoring of MM. This work aims to detect biomarkers of MRD in EVs isolated from MM patient samples at diagnosis and remission and compare the MRD-associated content between BM and PB EVs. EVs were isolated by size-exclusion chromatography, concentrated by ultrafiltration, and characterized according to their size and concentration, morphology, protein concentration, and the presence of EV-associated protein markers. EVs from healthy blood donors were used as controls. It was possible to isolate EVs from PB and BM carrying MM markers. Diagnostic samples had different levels of MM markers between PB and BM paired samples, but no differences between PB and BM were found at remission. EVs concentration was lower in the PB of healthy controls than of patients, and MM markers were mostly not detected in EVs from controls. This study pinpoints the potential of PB EVs from MM remission patients as a source of MM biomarkers and as a non-invasive approach for monitoring MRD.

Perspectives on emerging technologies, personalised medicine, and clinical research for cancer control in Latin America and the Caribbean.

Challenges of health systems in Latin America and the Caribbean include accessibility, inequity, segmentation, and poverty. These challenges are similar in different countries of the region and transcend national borders. The increasing digital transformation of health care holds promise of more precise interventions, improved health outcomes, increased efficiency, and ultimately reduced health-care costs. In Latin America and the Caribbean, the adoption of digital health tools is in early stages and the quality of cancer registries, electronic health records, and structured databases are problematic. Cancer research and innovation in the region are limited due to inadequate academic resources and translational research is almost fully dependent on public funding. Regulatory complexity and extended timelines jeopardise the potential improvement in participation in international studies. Emerging technologies, artificial intelligence, big data, and cancer research represent an opportunity to address the health-care challenges in Latin America and the Caribbean collectively, by optimising national capacities, sharing and comparing best practices, and transferring scientific and technical capabilities.

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.

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.

Comprehensive Genomic Studies on the Cell Blocks of Pancreatic Cancer.

Pancreatic cancer is one of the deadliest malignancies, characterized by late-stage diagnosis and limited treatment options. Comprehensive genomic profiling plays an important role in understanding the molecular mechanisms underlying the disease and identifying potential therapeutic targets. Cell blocks (CBs), derived from EUS-FNA, have become valuable resources for diagnosis and genomic analysis. We examine the molecular profile of pancreatic ductal adenocarcinoma (PDAC) using specimens obtained from CB EUS-FNA, across a large gene panel, within the framework of next-generation sequencing (NGS). Our findings revealed that over half (55%) of PDAC CB cases provided adequate nucleic acid for next-generation sequencing, with tumor cell percentages averaging above 30%. Despite challenges such as low DNA quantification and degraded DNA, sequencing reads showed satisfactory quality control statistics, demonstrating the detection of genomic alterations. Most cases (84.6%) harbored at least one gene variant, including clinically significant gene mutation variants such as KRAS, TP53, and CDKN2A. Even at minimal concentrations, as long as the extracted DNA is of high quality, performing comprehensive molecular profiling on PDAC samples from cell blocks has remained feasible. This strategy has yielded valuable information about the diagnosis, genetic landscape, and potential therapeutic targets, aligning closely with a precision cytopathology approach.

Epithelial E- and P-cadherins: role and clinical significance in cancer.

E-cadherin and P-cadherin are major contributors to cell-cell adhesion in epithelial tissues, playing pivotal roles in important morphogenetic and differentiation processes during development, and in maintaining integrity and homeostasis in adult tissues. It is now generally accepted that alterations in these two molecules are observed during tumour progression of most carcinomas. Genetic or epigenetic alterations in E- and P-cadherin-encoding genes (CDH1 and CDH3, respectively), or alterations in their proteins expression, often result in tissue disorder, cellular de-differentiation, increased invasiveness of tumour cells and ultimately in metastasis. In this review, we will discuss the major properties of E- and P-cadherin molecules, its regulation in normal tissue, and their alterations and role in cancer, with a specific focus on gastric and breast cancer models.

P-cadherin is coexpressed with CD44 and CD49f and mediates stem cell properties in basal-like breast cancer.

Although the luminal progenitor cell of the normal mammary gland hierarchy has been proposed as the cell-of-origin for basal-like breast cancers, finding the cancer stem cell (CSC) phenotype for this malignancy has proven a difficult task, mostly due to the lack of specific markers. Recently, basal-like sporadic and familial cases of breast cancer have been linked to BRCA1 gene inactivation, which enables the upregulation of the target-repressed CDH3/P-cadherin gene, an important biomarker of basal-like breast carcinomas. Previously, we demonstrated that P-cadherin overexpression can mediate aggressive behavior in these tumors. Thus, our aim was to test whether P-cadherin mediates stem cell properties in basal-like breast carcinomas. Using a series of breast cancer cell lines and primary tumors, we showed that P-cadherin was directly associated with the expression of the breast stem markers CD44, CD49f, and aldehyde dehydrogenase 1 in the basal subtype. Moreover, cell population enriched for P-cadherin expression comprised increased in vitro mammosphere-forming efficiency and capacity to grow colonies in three-dimensional cultures as well as greater tumorigenicity. Importantly, an association was found with stem-/progenitor-like phenotypes of the breast, including the luminal progenitor population, CD49f(+) CD24(+). Additionally, P-cadherin expression conferred resistance to x-ray-induced cell death, sustaining a role for this molecule in another stem cell property. In summary, we demonstrated, for the first time, that P-cadherin mediates stem cell properties, which could be explored in order to better define the CSC phenotype of basal-like breast tumors and the cell-of-origin of this malignancy.

p40: a p63 isoform useful for lung cancer diagnosis - a review of the physiological and pathological role of p63.

At present, p63, TTF-1, and Napsin-A are the main immunochemical markers used to distinguish squamous cell carcinoma (SCC) from lung adenocarcinoma (ADC). However, studies using antibodies against p63 have demonstrated false-positive results with positivity in some ADC. In contrast, the expression of one of the p63 isoforms (ΔNp63), detected by the antibody p40, is highly specific for SCC. Since most cases of lung cancer are diagnosed in small specimens (cytology/biopsies) and saving material for molecular analysis is mandatory, we recommended the use of p40 (in adjunct with TTF-1 and/or Napsin-A) as the best approach to discriminate SCC and lung ADC. In this paper, we review the physiological and pathological role of p63 isoforms as well as their use as diagnostic markers in lung SCC.

ICI 182,780 induces P-cadherin overexpression in breast cancer cells through chromatin remodelling at the promoter level: a role for C/EBPbeta in CDH3 gene activation.

CDH3/P-cadherin is a classical cadherin. Overexpression of which has been associated with proliferative lesions of high histological grade, decreased cell polarity and poor survival of patients with breast cancer. In vitro studies showed that it can be up-regulated by ICI 182,780, suggesting that the lack of ERalpha signalling is responsible for the aberrant P-cadherin overexpression and for its role in inducing breast cancer cell invasion and migration. However, the mechanism by which ER-signalling inhibition leads to P-cadherin expression is still unknown. The aim of this study was to explore the molecular mechanism linking the ERalpha-signalling and P-cadherin-regulated expression in breast cancer cell lines. This study showed that ICI 182,780 is able to increase CDH3 promoter activity, inducing high levels of the active chromatin mark H3 lysine 4 dimethylation. We also observed, for the first time, that the transcription factor C/EBPbeta is able to up-regulate CDH3 promoter activity in breast cancer cells. Moreover, we showed that the expression of P-cadherin and C/EBPbeta are highly associated in human breast carcinomas and linked with a worse prognosis of breast cancer patients. This study demonstrates the existence of an epigenetic regulation by which ICI 182,780 up-regulates P-cadherin expression in MCF-7/AZ breast cancer cells through chromatin remodelling at CDH3 promoter, bringing forward the growing evidence that ERalpha signalling-abrogation by anti-oestrogens is able to induce the expression of ERalpha-repressed genes which, in the appropriate cell biology context, may contribute to a breast cancer cell invasion phenotype.CDH3 GenBank accession no. NT_010498.

Nottingham Prognostic Index in triple-negative breast cancer: a reliable prognostic tool?

BACKGROUND: A breast cancer prognostic tool should ideally be applicable to all types of invasive breast lesions. A number of studies have shown histopathological grade to be an independent prognostic factor in breast cancer, adding prognostic power to nodal stage and tumour size. The Nottingham Prognostic Index has been shown to accurately predict patient outcome in stratified groups with a follow-up period of 15 years after primary diagnosis of breast cancer. Clinically, breast tumours that lack the expression of Oestrogen Receptor, Progesterone Receptor and Human Epidermal growth factor Receptor 2 (HER2) are identified as presenting a "triple-negative" phenotype or as triple-negative breast cancers. These poor outcome tumours represent an easily recognisable prognostic group of breast cancer with aggressive behaviour that currently lack the benefit of available systemic therapy. There are conflicting results on the prevalence of lymph node metastasis at the time of diagnosis in triple-negative breast cancer patients but it is currently accepted that triple-negative breast cancer does not metastasize to axillary nodes and bones as frequently as the non-triple-negative carcinomas, favouring instead, a preferentially haematogenous spread. Hypothetically, this particular tumour dissemination pattern would impair the reliability of using Nottingham Prognostic Index as a tool for triple-negative breast cancer prognostication. METHODS: The present study tested the effectiveness of the Nottingham Prognostic Index in stratifying breast cancer patients of different subtypes with special emphasis in a triple-negative breast cancer patient subset versus non- triple-negative breast cancer. RESULTS: We demonstrated that besides the fact that TNBC disseminate to axillary lymph nodes as frequently as luminal or HER2 tumours, we also showed that TNBC are larger in size compared with other subtypes and almost all grade 3. Additionally, survival curves demonstrated that these prognostic factors are equally important to stratify different survival outcomes in non-TNBC as in TNBC. We also showed that the NPI retains the ability to stratify and predict survival of TNBC patients. CONCLUSION: The importance of this study relies on the need of prognostication improvements on TNBC, showing, at a clinical standpoint, that Nottingham Prognostic Index is as a truthful prognostic tool in TNBC.

Terminal α2,6-sialylation of epidermal growth factor receptor modulates antibody therapy response of colorectal cancer cells.

BACKGROUND: The epidermal growth factor receptor (EGFR) is a key protein involved in cancer development. Monoclonal antibodies targeting EGFR are approved for the treatment of metastatic colorectal cancer (CRC). Despite the beneficial clinical effects observed in subgroups of patients, the acquisition of resistance to treatment remains a major concern. Protein N-glycosylation of cellular receptors is known to regulate physiological processes leading to activation of downstream signaling pathways. In the present study, the role of EGFR-specific terminal ⍺2,6-sialylation was analyzed in modulation of the malignant phenotype of CRC cells and their resistance to monoclonal antibody Cetuximab-based therapy. METHODS: Glycoengineered CRC cell models with specific sialyltransferase ST6GAL1 expression levels were applied to evaluate EGFR activation, cell surface glycosylation and therapeutic response to Cetuximab. RESULTS: Glycoproteomic analysis revealed EGFR as a major target of ST6Gal1-mediated ⍺2,6-sialylation in a glycosite-specific manner. Mechanistically, CRC cells with increased ST6Gal1 expression and displaying terminal ⍺2,6-sialylation showed a marked resistance to Cetuximab-induced cytotoxicity. Moreover, we found that this resistance was accompanied by downregulation of EGFR expression and its activation. CONCLUSIONS: Our data indicate that EGFR ⍺2,6-sialylation is a key factor in modulating the susceptibility of CRC cells to antibody targeted therapy, thereby disclosing a potential novel biomarker and providing key molecular information for tailor made anti-cancer strategies.

Estrogens, MSI and Lynch syndrome-associated tumors.

Estrogens play a major role in the biology of hormone-responsive tissues but also in the normal physiology of various non-typical hormone-responsive tissues. In disease, estrogens have been associated with tumor development, in particular with tumors such as breast, endometrium, ovary and prostate. In this paper we will review the molecular mechanisms by which estrogens are involved in cancer development, with a special focus in Lynch syndrome related neoplasia. Further, we discuss the role estrogens might have on cell proliferation and apoptosis, how estrogens metabolites can induce DNA damage and we discuss a possible connection between estrogens and changes in DNA (hypo- and hyper) methylation. In this review we will also address the protective effect that high levels of estrogens have in MMR related neoplasias.

Monocarboxylate transporter 1 is up-regulated in basal-like breast carcinoma.

AIMS: Monocarboxylate transporters (MCTs) have been considered promising targets for cancer therapy, since they facilitate lactate efflux in glycolytic tumours. However, their role in solid tumours is still poorly understood. Thus, the present work aimed to contribute to understanding the involvement of MCT1 and MCT4 in breast cancer progression as well as MCT regulation by CD147. METHODS AND RESULTS: The expression of the membrane transporters MCT1 and MCT4 was analysed in a series of breast carcinomas (249 cases) and their clinicopathological significance investigated. Additionally, we analysed the significance of CD147 co-expression, as an important regulator of MCT expression and activity. MCT1 was significantly increased in breast carcinomas when compared with normal breast tissue and, importantly, both MCT1 and CD147 were associated with poor prognostic variables such as basal-like subtype and high grade tumours. CONCLUSIONS: These results provide evidence for a prognostic value of MCT1 in breast carcinoma and support the exploitation of the complex MCT1/CD147 as a promising target for cancer therapy, especially in basal-like breast carcinoma.

Expression of FOXA1 and GATA-3 in breast cancer: the prognostic significance in hormone receptor-negative tumours.

INTRODUCTION: The expression of additional genes, other than oestrogen receptor (ER), may be important to the hormone-responsive phenotype of breast cancer. Microarray analyses have revealed that forkhead box A1 (FOXA1) and GATA binding protein 3 (GATA-3) are expressed in close association with ERalpha, both encoding for transcription factors with a potential involvement in the ERalpha-mediated action in breast cancer. The purpose of this study was to explore if the expression of FOXA1 and GATA-3 may provide an opportunity to stratify subsets of patients that could have better outcome, among the ERalpha-negative/poor prognosis breast cancer group. METHODS: We evaluate FOXA1 and GATA-3 expression in 249 breast carcinomas by immunohistochemistry, associating it with breast cancer molecular markers, clinicopathological features and patient's survival. The clinicopathological features and immunohistochemical markers of the tumours were compared using the chi-square test and ANOVA. Disease-free survival was analysed through Kaplan-Meier survival curves and Cox regression. RESULTS: FOXA1 expression was demonstrated in 42% of invasive carcinomas, while GATA-3 was detected in 48% of the cases. FOXA1 expression was inversely associated with tumour size, Nottingham Prognostic Index, histological grade, lymph vascular invasion, lymph node stage and human epidermal growth factor receptor-2 (HER-2) overexpression, while GATA-3 expression showed inverse association with histological grade and HER-2. Both FOXA1 and GATA-3 were directly associated with ERalpha and progesterone receptor. Among FOXA1-positive tumours, 83.1% are comprised in the luminal A subtype, similar to GATA-3 where 87.7% of positive tumours were classified within this molecular subtype. In the subset of ERalpha-negative patients, those who were FOXA1-negative had a 3.61-fold increased risk of breast cancer recurrence when compared with the FOXA1-positive. CONCLUSIONS: FOXA1 was a significant predictor of good outcome in breast cancer, whereas GATA-3 was an important luminal marker. The expression of FOXA1 may be used for risk stratification among ERalpha-negative patients.

A [Glyco]biomarker that Predicts Failure to Standard Therapy in Ulcerative Colitis Patients.

BACKGROUND AND AIMS: There is a clinical need to identify biomarkers able to select patients who are most likely to develop aggressive/complicated disease, for early selection for appropriate therapy. Changes in the glycosylation profile of intestinal lymphocytic infiltrate were previously demonstrated to regulate T cell activity, being associated with disease severity in ulcerative colitis [UC] patients. We interrogated whether this heterogeneous expression of branched N-glycans in intestinal inflammatory infiltrate predicts therapy response early in disease course. METHODS: The expression levels of the branched N-glycans in colonic biopsies collected around time of diagnosis from a well-characterised cohort of 131 UC patients were correlated with response to standard therapy. Receiver operating characteristic analysis and specificity/sensitivity were determined. RESULTS: Branched N-glycans levels around time of diagnosis predict non-response to conventional therapy with 75% specificity. Moreover, high levels of branched N-glycans predict 78% of UC patients who will display a favourable disease course [exclusively under 5-aminosalicylate therapy for more than 5 years of disease]. The best predictive performance was observed in severe UC patients with Mayo endoscopic subscore 3 and in those that were naïve to therapy. Multivariable analysis revealed that low levels of branched N-glycans and high levels of C-reactive protein [CRP] around time of diagnosis act as independent predictors of non-response to standard therapy. A powerful effect of the combined use of the branched N-glycans and CRP was observed. CONCLUSIONS: Our results reveal a potential [glyco]biomarker that predicts, early in the disease course, patients who will fail to respond to standard therapy, benefiting thereby from other therapeutic strategies such as biologics.

The transcription factor FOXO3a is a crucial cellular target of gefitinib (Iressa) in breast cancer cells.

Gefitinib is a specific inhibitor of the epidermal growth factor receptor (EGFR) that causes growth delay in cancer cell lines and human tumor xenografts expressing high levels of EGFR. An understanding of the downstream cellular targets of gefitinib will allow the discovery of biomarkers for predicting outcomes and monitoring anti-EGFR therapies and provide information for key targets for therapeutic intervention. In this study, we investigated the role of FOXO3a in gefitinib action and resistance. Using two gefitinib-sensitive (i.e., BT474 and SKBR3) as well as three other resistant breast carcinoma cell lines (i.e., MCF-7, MDA-MB-231, and MDA-MB-453), we showed that gefitinib targets the transcription factor FOXO3a to mediate cell cycle arrest and cell death in sensitive breast cancer cells. In the sensitive cells, gefitinib treatment causes cell cycle arrest predominantly at the G(0)-G(1) phase and apoptosis, which is associated with FOXO3a dephosphorylation at Akt sites and nuclear translocation, whereas in the resistant cells, FOXO3a stays phosphorylated and remains in the cytoplasm. The nuclear accumulation of FOXO3a in response to gefitinib was confirmed in tumor tissue sections from breast cancer patients presurgically treated with gefitinib as monotherapy. We also showed that knockdown of FOXO3a expression using small interfering RNA (siRNA) can rescue sensitive BT474 cells from gefitinib-induced cell-proliferative arrest, whereas reintroduction of active FOXO3a in resistant MDA-MB-231 cells can at least partially restore cell-proliferative arrest and sensitivity to gefitinib. These results suggest that the FOXO3a dephosphorylation and nuclear localization have a direct role in mediating the gefitinib-induced proliferative arrest and in determining sensitivity to gefitinib.

P-cadherin expression in breast cancer: a review.

P-cadherin is frequently over-expressed in high-grade invasive breast carcinomas and has been reported to be an enhancer of migration and invasion of breast cancer cells, being correlated with tumour aggressiveness. In addition, expression of P-cadherin is well established as an indicator of poor prognosis in human breast cancer, which has stimulated our interest in studying its role in this setting. This review describes the most important findings on P-cadherin expression and function in normal mammary tissue and breast cancer cells, emphasizing that further research is required to elucidate the role played by this protein in human mammary tumours.

P-cadherin overexpression is an indicator of clinical outcome in invasive breast carcinomas and is associated with CDH3 promoter hypomethylation.

PURPOSE: P-cadherin overexpression has been reported in breast carcinomas, where it was associated with proliferative high-grade histological tumors. This study aimed to analyze P-cadherin expression in invasive breast cancer and to correlate it with tumor markers, pathologic features, and patient survival. Another purpose was to evaluate the P-cadherin promoter methylation pattern as the molecular mechanism underlying this gene regulation. EXPERIMENTAL DESIGN: Using a series of invasive breast carcinomas, P-cadherin expression was evaluated and correlated with histologic grade, estrogen receptor, MIB-1, and p53 and c-erbB-2 expression. In order to assess whether P-cadherin expression was associated with changes in CDH3 promoter methylation, we studied the methylation status of a gene 5'-flanking region in these same carcinomas. This analysis was also done for normal tissue and for a breast cancer cell line treated with a demethylating agent. RESULTS: P-cadherin expression showed a strong correlation with high histologic grade, increased proliferation, c-erbB-2 and p53 expression, lack of estrogen receptor, and poor patient survival. This overexpression can be regulated by gene promoter methylation because the 5-Aza-2'-deoxycytidine treatment of MCF-7/AZ cells increased P-cadherin mRNA and protein levels. Additionally, we found that 71% of P-cadherin-negative cases showed promoter methylation, whereas 65% of positive ones were unmethylated (P = 0.005). The normal P-cadherin-negative breast epithelial cells showed consistent CDH3 promoter methylation. CONCLUSIONS: P-cadherin expression was strongly associated with tumor aggressiveness, being a good indicator of clinical outcome. Moreover, the aberrant expression of P-cadherin in breast cancer might be regulated by gene promoter hypomethylation.

Maspin expression in normal skin and usual cutaneous carcinomas.

Maspin is a serine protease inhibitor whose gene is located on 3q27. Several lines of evidence point towards its putative role as tumor suppressor gene and angiogenesis inhibitor; however, there are compelling data showing that maspin is also expressed in the nuclear compartment and might be associated with the differentiation of specific cell lineages. No systematic study of maspin expression in normal skin and usual skin carcinomas have been published so far. We semiquantitatively analyzed the distribution and immunoreactivity pattern of maspin in 14 squamous cell carcinomas (SCCs) and 16 basal cell carcinomas (BCCs) and in the adjacent normal epidermis of all cases. We also examined the correlation of maspin expression with histological type, grade, vascular invasion, perineural infiltration, and mitotic counting. Cytoplasmic expression of maspin was observed in suprabasal, prickle, and granular cell layers of normal epidermis; cells of the germinative hair matrix, Henle's and Huxley's layers, and cuticle of hair follicles; mature sebaceous cells and sweat gland's secretory cells. Nuclear expression was detected in some basal/myoepithelial cells of the sweat glands and scattered mature sebaceous cells. All SCCs but one grade IV SCC showed maspin expression, and it was correlated with the differentiation of these neoplasms. BCCs presented variable maspin expression, while metatypical carcinomas showed moderate to intense maspin expression, nodular BCCs variable contents of maspin and displayed a peculiar distribution, confined to the center of the neoplastic nodules. Two BCCs and one SCC showed maspin nuclear expression. No correlation with other clinical pathological features was observed. Our findings do not support the role of maspin as a tumor suppressor gene and suggest that this serpin is probably associated with specific lines of differentiation.

Naked nuclei revisited: p63 Immunoexpression.

The presence of naked nuclei (NN) in cytological preparations of fibroadenomas is a well-known finding. Regardless of their importance on the differential diagnosis of fibroadenoma and other benign lesions of the breast, the origin of NN remains elusive. Despite previous efforts to characterize them, the lack of a reliable nuclear marker for myoepithelial cells impaired definitive conclusions. We performed a systematic evaluation of p63 expression in cytological and histological preparations of 10 fibroadenoma specimens. We observed that in histological sections, p63 was restricted to the nuclei of myoepithelial/basal cells in lobules and ducts of normal breast. In fibroadenomas, p63 decorated the nuclei of myoepithelial cells in the periphery of epithelial duct-like formations and slit-like formations. No p63 immunoreactivity was observed in stromal or epithelial cells. In cytological samples, almost all NN and cells surrounding epithelial cell clusters were stained; no stromal cell admixed with fibrillary matrix or epithelial cell was stained with p63. Based on our findings, we strongly suggest that most, if not all, NN are myoepithelial in origin.