Frizzled (FZD) proteins, a family of Wnt receptors, are involved in carcinogenesis in different organs. One interesting FZD protein is FZD-10 highly expressed in embryogenesis but completely absent in the membrane or cytosol of healthy proliferated cells. We studied in detail the expression level and the location of Frizzled-10 protein in different cancerous tissues, such as colon, melanoma and gastric cancer and in function of different staging of the tumor and in metastases. We observed a correlation between cancer evolution and FZD-10 expression, and localization of protein during carcinogenesis. In colon, we have an increase of cytoplasmic FZD-10 expression from hyperplastic mucosa to metastatic tissues, while the amount in the nucleus decreases significantly in T3 and T4 staging tumors as well as in metastases. In melanoma and gastric cancer, we observed the opposite trend of FZD-10 protein in the cytosol but both show a decrease in the T3 and T4 stage of the tumor and in metastases. However, the decrease is less prominent in gastric cancer.Our findings indicate an important role of FZD-10 in tumor progression especially in the later stages of tumor. The nuclear expression of FZD-10 or its absence can give a new tool for tumor staging to pathologists. For target therapy, at least for colon cancer, the high presence of FZD-10 in the later stages of tumor progression and the absence in healthy tissue present a promising new approach.

BACKGROUND: Anaplastic lymphoma kinase (ALK) rearrangement represents a landmark in the targeted therapy of non–small cell lung cancer (NSCLC). Immunohistochemistry (IHC) is a sensitive and specific method to detect ALK protein expression, possibly an alternative to fluorescence in situ hybridization (FISH). In this study, the concordance of FISH and IHC to determine ALK status was evaluated, particularly focusing on discordant cases. MATERIALS AND METHODS: ALK status was tested by FISH and the IHC validated method (Ventana ALK (D5F3) CDx Assay) in 95 NSCLCs. Discordant cases were analyzed also by next-generation sequencing (NGS). The response to crizotinib of treated patients was recorded. RESULTS: Seven (7.3%) discordant cases were ALK FISH positive and IHC negative. They showed coexistent split signals pattern, with mean percentage of 15.4%, and 5′ deletions pattern, with mean percentage 31.7%. Two cases had also gene amplification pattern. In three cases (42.8 %), the polysomy was observed. The NGS assay confirmed IHC results. In these patients, the treatment with crizotinib was ineffective. CONCLUSIONS: In our discordant cases, a coexistent complex pattern (deleted, split, and amplified/polysomic) of ALK gene was observed by FISH analysis. These complex rearranged cases were not detectable by IHC, and it could be speculated that more complex biological mechanisms could modulate protein expression. These data highlight the role of IHC and underscore the complexity of the genetic pattern of ALK. It could be crucial to consider these findings in order to best select patients for anti-ALK treatment in daily clinical practice.

Diagnostic assays for human epidermal growth factor receptor 2 (HER2) expression have a high predictive value because patients with HER2-positive tumors could benefit from HER2-targeted therapy. The aim of the present study is to analyze the incidence of HER2 gene amplification in selected tumors with adverse features that scored 1+ by immunohistochemistry (IHC). For that purpose, 331 consecutive invasive breast cancers (IBCs) were tested by IHC for HER2 expression between January and December 2013, 102 of which (31%) scored 1+. Of these 102 women with IBC who underwent surgery, 75 entered the study (73.5%). A total of 48 out of 75 (64%) IBC samples (patients' median age, 60.75 years) were selected according to ≥1 unfavorable tumor characteristics, and tested by fluorescence in situ hybridization (FISH). Of these 48 IBC samples scoring 1+ by IHC, 22 (46%) exhibited high histological grade (G3), 23 (48%) had a high proliferative index (Ki-67, >30%), 27 (56%) showed vascular invasion and 32 out of 41 evaluable cases (78%) were node-positive. Regarding hormone receptor expression, 3 (6%) and 10 (21%) cases were negative for estrogen and progesterone receptors expression, respectively. FISH was performed on 48 IBC cases scoring 1+ by IHC, and 7 infiltrating ductal carcinomas (IDCs) (14.6%) demonstrated HER2 amplification with a high proliferative index. In 42 IDC samples, statistical analysis evidenced a significant association between histological grade and high proliferative index (P=0.0200). In addition, in 48 HER2 scoring 1+ IBCs, Fisher's exact test evidenced a significant association between the presence of gene amplification and high proliferative index (P=0.0033). Based on these biopathological parameters, particularly a high proliferative index, the present results indicate that it is possible to of identify tumors scoring 1+ by IHC with HER2 amplification by FISH, thus aiding the selection of patients who are suitable for HER2-targeted therapy according to an acceptable cost/benefit ratio.

Cellular resistance in advanced gastric cancer (GC) might be related to function of multidrug resistance (MDR) proteins. The adaptor protein NHERF1 (Na C /H C exchanger regulatory factor) is an important player in cancer progression for a number of solid malignancies, even if its role to develop drug resistance remains uncertain. Herein, we aimed to analyze the potential association between NHERF1 expression and P-gp, sorcin and HIF-1a MDR-related proteins in advanced GC patients treated with epirubicin/oxaliplatin/capecitabine (EOX) chemotherapy regimen, and its relation to response. Total number of 28 untreated patients were included into the study. Expression and subcellular localization of all proteins were assessed by immunohistochemistry on formalin-fixed paraffin embedded tumor samples. We did not found significant association between NHERF1 expression and the MDR-related proteins. A trend was observed between positive cytoplasmic NHERF1 (cNHERF1) expression and negative nuclear HIF-1a (nHIF-1a) expression (68.8% versus 31.3% respectively, P D 0.054). However, cytoplasmic P-gp (cP-gp) expression was positively correlated with both cHIF-1a and sorcin expression (P D 0.011; P D 0.002, respectively). Interestingly, nuclear NHERF1 (nNHERF1) staining was statistically associated with clinical response. In detail, 66.7% of patients with high nNHERF1 expression had a disease control rate, while 84.6% of subjects with negative nuclear expression of the protein showed progressive disease (P D 0.009). Multivariate analysis confirmed a significant correlation between nNHERF1 and clinical response (OR 0.06, P D 0.019). These results suggest that nuclear NHERF1 could be related to resistance to the EOX regimen in advanced GC patients, identifying this marker as a possible independent predictive factor.