Enrico Vasquez

Specific Alterations of MicroRNA Transcriptome and Global Network Structure in Colorectal Carcinoma after Cetuximab Treatment

The relationship between therapeutic response and modifications of microRNA (miRNA) transcriptome in colorectal cancer (CRC) remains unknown. We investigated this issue by profiling the expression of 667 miRNAs in 2 human CRC cell lines, one sensitive and the other resistant to cetuximab (Caco-2 and HCT-116, respectively), through TaqMan real-time PCR. Caco-2 and HCT-116 expressed different sets of miRNAs after treatment. Specifically, 21 and 22 miRNAs were differentially expressed in Caco-2 or HCT-116, respectively (t test, P < 0.01). By testing the expression of differentially expressed miRNAs in CRC patients, we found that miR-146b-3p and miR-486-5p are more abundant in K-ras–mutated samples with respect to wild-type ones (Wilcoxon test, P < 0.05). Sixty-seven percent of differentially expressed miRNAs were involved in cancer, including CRC, whereas 19 miRNA targets had been previously reported to be involved in the cetuximab pathway and CRC. We identified 25 transcription factors putatively controlling these miRNAs, 11 of which have been already reported to be involved in CRC. On the basis of these data, we suggest that the downregulation of let-7b and let-7e (targeting K-ras) and the upregulation of miR-17* (a CRC marker) could be considered as candidate molecular markers of cetuximab resistance. Global network functional analysis (based on miRNA targets) showed a significant overrepresentation of cancer-related biological processes and networks centered on critical nodes involved in epidermal growth factor receptor internalization and ubiquitin-mediated degradation. The identification of miRNAs, whose expression is linked to the efficacy of therapy, should allow the ability to predict the response of patients to treatment and possibly lead to a better understanding of the molecular mechanisms of drug response. Mol Cancer Ther; 9(12); 3396–409.

Solid pseudopapillary tumour of the pancreas.

Solid pseudopapillary tumour (Frantz’s tumour) is a rare benign or low-grade neoplasm of the pancreas with distinct clinicopathological features. The diagnosis may be difficult, but should be considered as a possibility in young women who present with a large abdominal mass involving the pancreas. In this report we describe the clinical and pathological characteristics of these lesions and discuss therapeutic options. A 27-year-old woman was admitted to our hospital in October 1999 with a 3-month history of mild asthenia, weight loss, and a feeling of heaviness localised to the left lumbar region. Routine laboratory analyses were all within the normal range; however, abdominal ultrasound revealed the presence of a mass in the superior abdomen, which was composed of high and low echoic areas. Computed tomography showed the mass to be solid, encapsulated, and about 9 cm in diameter (figure 1). It was composed of high and low density areas and was located close to the splenic hilum—above the pancreatic tail, behind the stomach, and between the spleen and the aorta (Albarran-Chatelin’s anatomosurgical quadrilateral). Nuclear magnetic resonance imaging revealed a posterior anastomosis between the mass drainage vessels and the diaphragmatic veins. The patient underwent a complete surgical resection of the tumour (enucleation) and a resection of the pancreatic tail. The spleen was removed because of vascular damage consisting of large patches of venous stasis and ischaemia which did not resolve over time or after application of warm, moist dressings. The pathological diagnosis was epithelial solid papillary cystic neoplasm with signs of incipient pseudocapsular invasion. Immunohistochemical studies established that the tumour cells were positive for cytokeratins and vimentin, and negative for S100 protein, CD34, chromogranin, neurone-specific enolase (NSE), and glial fibrillary acid protein. The postoperative course was uneventful and the patient was discharged from hospital on day 7 and referred to the Centro di Riferimento Oncologico di Aviano for counselling. No residual tumour or metastases were found with clinical imaging, so no adjuvant therapy was recommended. In November 2001, routine work-up identified no signs of tumour relapse. This case is a typical example of solid pseudopapillary tumour of the pancreas or Frantz’s tumour. The diagnostic imaging techniques used were effective in defining the location of the tumour, although a nonfunctioning neoplasm of the left adrenal gland was initially suspected. In 1959, Frantz 1

Specific alterations of the microRNA transcriptome and global network structure in colorectal cancer after treatment with MAPK/ERK inhibitors

The mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) pathway has a master control role in various cancer-related biological processes as cell growth, proliferation, differentiation, migration, and apoptosis. It also regulates many transcription factors that control microRNAs (miRNAs) and their biosynthetic machinery. To investigate on the still poorly characterised global involvement of miRNAs within the pathway, we profiled the expression of 745 miRNAs in three colorectal cancer (CRC) cell lines after blocking the pathway with three different inhibitors. This allowed the identification of two classes of post-treatment differentially expressed (DE) miRNAs: (1) common DE miRNAs in all CRC lines after treatment with a specific inhibitor (class A); (2) DE miRNAs in a single CRC line after treatment with all three inhibitors (class B). By determining the molecular targets, biological roles, network position of chosen miRNAs from class A (miR-372, miR-663b, miR-1226*) and class B (miR-92a-1*, miR-135b*, miR-720), we experimentally demonstrated that they are involved in cell proliferation, migration, apoptosis, and globally affect the regulation circuits centred on MAPK/ERK signaling. Interestingly, the levels of miR-92a-1*, miR-135b*, miR-372, miR-720 are significantly higher in biopsies from CRC patients than in normal controls; they also are significantly higher in CRC patients with mutated KRAS than in those with wild-type genotypes (Wilcoxon test, p < 0.05): the latter could be a downstream effect of ERK pathway overactivation, triggered by KRAS mutations. Finally, our functional data strongly suggest the following miRNA/target pairs: miR-92a-1*/PTEN-SOCS5; miR-135b*/LATS2; miR-372/TXNIP; miR-663b/CCND2. Altogether, these results contribute to deepen current knowledge on still uncharacterized features of MAPK/ERK pathway, pinpointing new oncomiRs in CRC and allowing their translation into clinical practice and CRC therapy.

Primitive neuroectodermal tumour of the uterus with focal cartilaginous differentiation

The uterus was enlarged (130 × 80 × 40 mm) with a smooth external surface. The endometrial cavity was filled by a white-grey, polypoidal, fleshy mass arising from the posterior wall, and measuring 60 mm in maximum dimension (Figure 1). Areas of necrosis and haemorrhage were present and, focally, the tumour invaded the myometrium but did not reach the peritoneal surface. The ovaries and Fallopian tubes were unremarkable: a paratubal cyst was present on the right. Histology revealed a neoplasm composed of solid sheets of undifferentiated round to oval cells forming numerous rosettes. The rosettes were mostly of the Flexner–Wintersteiner type (Figure 2a), but some Homer–Wright type and perivascular pseudorosettes were occasionally detected. In some areas, neuroectodermal columnar cells were arranged in medulloepithelial tubules dispersed through a loose fibrillary matrix (Figure 2b). The neoplastic cells had scanty cytoplasm, oval, hyperchromatic nuclei and inconspicuous nucleoli. Mitotic figures were absent or rare. Necrotic and haemorrhagic areas were scattered throughout the tumour. No lymphatic or vascular invasion was seen. Most of the endometrium was replaced by tumour and uninvolved endometrium was atrophic with occasional cystically dilated glands (Figure 2a). Despite extensive sampling, we found only a small island of mature cartilage which was confirmed immunohistochemically by S-100 protein positivity (Figure 2c). No other mesodermal or epithelial elements were identified within the neoplasm. At margin of the mass, tumour infiltrated the superficial myometrium. The Fallopian tubes and ovaries were atrophic, in keeping with the patient’s age. There was no evidence of metastatic tumour in the pelvic lymph nodes. The tumour was classified as stage IB (FIGO). Immunohistochemistry was carried out using standard avidin-biotin-peroxidase complex (ABC). The following antibodies were used: neuron-specific-enolase Histopathology 1997, 30, 483–485

Dysregulated miR-671-5p / CDR1-AS / CDR1 / VSNL1 axis is involved in glioblastoma multiforme

MiR-671-5p is encoded by a gene localized at 7q36.1, a region amplified in human glioblastoma multiforme (GBM), the most malignant brain cancer. To investigate whether expression of miR-671-5p were altered in GBM, we analyzed biopsies from a cohort of forty-five GBM patients and from five GBM cell lines. Our data show significant overexpression of miR-671-5p in both biopsies and cell lines. By exploiting specific miRNA mimics and inhibitors, we demonstrated that miR-671-5p overexpression significantly increases migration and to a less extent proliferation rates of GBM cells. Through a combined in silico and in vitro approach, we identified CDR1-AS, CDR1, VSNL1 as downstream miR-671-5p targets in GBM. Expression of these genes significantly decreased both in GBM biopsies and cell lines and negatively correlated with that of miR-671-5p. Based on our data, we propose that the axis miR-671-5p / CDR1-AS / CDR1 / VSNL1 is functionally altered in GBM cells and is involved in the modification of their biopathological profile.

Nuclear BMI-1 expression in laryngeal carcinoma correlates with lymph node pathological status.

BackgroundThe main cause of treatment failure and death in laryngeal squamous cell carcinoma is metastasis to the regional lymph nodes. The current clinical staging criteria fail to differentiate patients with occult metastasis from patients without metastasis. Identifying molecular markers of the disease might improve our understanding of the molecular mechanisms underlying the pathogenesis and development of laryngeal carcinoma and may help improve clinical staging and treatment.MethodsSixty-four previously untreated patients who underwent surgical excision of laryngeal squamous cell carcinoma with neck dissection were included in this study. The expression of B cell-specific Moloney murine leukemia virus integration site 1 (BMI-1) was examined immunohistochemically on formalin-fixed paraffin-embedded primary tissue specimens.ResultsNuclear expression of BMI-1 (nBMI-1) was detected in 32 of the 64 tumors (50%), cytoplasmic expression of BMI-1 (cBMI-1) was detected in 22 (34.4%), and 10 tumors (15.6%) showed no BMI-1 immunoreactivity. High nBMI-1 expression levels (≥10) were detected in 28 of the 32 (87.5%) nBMI-1-positive patients. Multivariate analysis including age at diagnosis, grade, tumor location, TNM status, and nBMI-1 expression showed that a high nBMI-1 expression level was an independent prognostic factor for lymph node metastasis.ConclusionThe expression of BMI-1 in patients with laryngeal carcinoma seems to correlate with lymph node metastasis.

Expression of BMI1 and p16 in laryngeal squamous cell carcinoma

The clinical evolution of laryngeal squamous cell carcinoma (SCC) is undetectable with the current staging criteria. To more completely understand the biology of laryngeal SCC, we assessed the expression of the proteins B‐cell–specific Moloney murine leukemia virus integration site 1 (BMI1) and p16.

EPITHELIAL INCLUSION IN AXILLARY LYMPH NODE ASSOCIATED WITH A BREAST CARCINOMA : REPORT OF A CASE WITH A REVIEW OF THE LITERATURE

Since epithelial inclusions in axillary lymph nodes are rare, some diagnostic problems concerning their nature have arisen. We report a case of a cystic squamous epithelial inclusion in an axillary lymph node of a patient affected by a high grade breast carcinoma with focal squamous differentiation. Although the lesion was histologically benign, the possibility that it represents a well-differentiated squamous metastasis cannot be completely ruled out. To our knowledge, this is the first case of an epithelial lymph node inclusion associated with a breast carcinoma showing morphological evidence of squamous differentiation. Here, we discuss the diagnostic problems and give a review of the literature.

Wilms tumor 1 (WT1) protein: Diagnostic utility in pediatric tumors

Despite Wilms tumor 1 (WT1) protein was originally considered as a specific immunomarker of Wilms tumor, with the increasing use of immunohistochemistry, there is evidence that other tumors may share WT1 protein expression. This review focuses on the immunohistochemical profile of WT1 protein in the most common malignant tumors of children and adolescents. The variable expression and distribution patterns (nuclear vs cytoplasmic) in the different tumors, dependent on the antibodies used (anti-C or N-terminus WT1 protein), will be emphasized by providing explicative illustrations. Potential diagnostic pitfalls from unexpected WT1 protein expression in some tumors will be discussed in order to avoid diagnostic errors, especially when dealing with small biopsies.

Comparative Immunohistochemical Analysis of the Expression of Cytokeratins, Vimentin and α-smooth Muscle Actin in Human Foetal Mesonephros and Metanephros

The human mesonephros is currently regarded as a simplified version of the foetal metanephros, primarily due to the close morphological resemblance between these two structures. The aim of the present study was to define whether human mesonephric and foetal metanephric nephrons share immunophenotypical traits in their corresponding structures (glomeruli, proximal and distal tubules). For this purpose we first investigated immunohistochemically the overall expression and topographical distribution of cytokeratins 7, 8, 18, 19, and 20, vimentin and α-smooth muscle actin in mature mesonephric nephrons and compared the results with those obtained in maturing-stage foetal metanephric nephrons. No expression of cytokeratins 7 and 20 was found. Cytokeratins 8, 18, and 19 and vimentin showed a restricted and basically coincident expression along the different components of both mesonephric and metanephric nephrons. These findings indicate that the intermediate filament protein profile of human mature mesonephric nephrons closely recapitulates that observed in developing metanephros and thereby strengthens the concept that human mesonephros, a transient ontogenic structure, is largely similar to the foetal metanephros.The sole difference between human mesonephros and foetal metanephros was the divergent expression of α-smooth muscle actin. This protein exhibited an increasingly accentuated mesangial expression paralleling the morphological maturation of metanephric glomerulus, whereas it was absent from the mesonephric one. This would suggest that the mesangial cells in these two renal structures have a different function during the foetal life.