Dual immunocytochemical staining of annexin A10 and p53 in low-grade and papillary urothelial carcinoma contributes to improvement of diagnostic accuracy in urine cytology.

BACKGROUND: Urothelial carcinoma (UC) is a common type of human cancer and, although urine cytology is a useful method for identifying high-grade UC (HGUC), its ability to diagnose low-grade UC (LGUC) is limited. The authors previously reported that annexin A10 (ANXA10) expression was strongly linked to both papillary and early stage LGUC and was inversely correlated with p53 expression in upper tract UC (UTUC) and bladder UC. However, it remains largely unknown whether ANXA10 is useful as a diagnostic marker for urine cytology. METHODS: In this study, the authors used 104 biopsy and 314 urine cytology samples to investigate the efficacy of ANXA10 and p53 expression by immunohistochemistry and immunocytochemistry. RESULTS: In immunohistochemistry analysis, expression levels of ANXA10 and p53 were either weak or absent in noncancerous tissues, whereas ANXA10 overexpression was observed patients with LGUC, and strong expression of p53 was identified in patients with HGUC. In immunocytochemistry analysis, sensitivity was not good for the detection of UC, especially UTUC, by cytology alone, but it was improved by combining cytology with ANXA10 and p53 to detect both bladder UC and UTUC. Receiver operating characteristic curve analysis also confirmed the diagnostic superiority of cytology combining ANXA10 and p53 for the detection of all UCs, including both HGUC and LGUC (area under the curve, 0.84). CONCLUSIONS: To the authors' knowledge, this is the first report that the combination of ANXA10 and p53 has potential application as a diagnostic immunomarker for improving the diagnostic accuracy of urine cytology.

Analysis of MicroRNA in Bile Cytologic Samples Is Useful for Detection and Diagnosis of Extrahepatic Cholangiocarcinoma.

OBJECTIVES: This study aimed to develop reliable biomarkers that improve the ability of bile cytology to diagnose cholangiocarcinoma vs benign biliary lesions. METHODS: Many studies indicate that microRNAs (miRNAs) are potential candidates for the early diagnosis of cancer. We analyzed the expression of five tumor-associated miRNAs (miR-31-5p, miR-122-5p, miR-378d, miR-182-5p, and miR-92a-3p) in cytology samples using quantitative reverse transcription polymerase chain reaction. We collected 52 surgically resected tissue samples, 84 cytologic specimens from smears (53 cases of cancer and 31 cases of noncancer), and 40 residual sediments after smearing for routine cytology at Hiroshima University Hospital. RESULTS: The expression of miR-31-5p, miR-378d, and miR-122-5p was significantly higher in cancer tissues than those in normal tissues, while miR-182-5p expression was lower. The expression of miR-31-5p, miR-378d, miR-182-5p, and miR-92a-3p was significantly higher in detached cell samples from smears of cholangiocarcinoma cases than in those from noncancer cases. CONCLUSIONS: These results suggest that the analysis of miRNAs in bile cytologic specimens is a promising auxiliary tool for distinguishing cholangiocarcinoma from benign biliary lesions.

Optimal indication of endoscopic retrograde pancreatography-based cytology in the preoperative pathological diagnosis of pancreatic ductal adenocarcinoma.

BACKGROUND: Endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) is extremely useful for pathological diagnosis of pancreatic ductal adenocarcinoma (PDAC); however, puncturing is difficult in some cases, and there is a risk of needle tract seeding. This study evaluated the indications for endoscopic retrograde pancreatography-based (ERP)-based cytology for the preoperative diagnosis of PDAC. METHODS: This study included 267 patients with PDAC who underwent preoperative ERP. The diagnostic performance of ERP-based cytology for PDAC was evaluated based on the sample collection method (pancreatic juice cytology [PJC] during ERP, brush cytology, PJC via endoscopic nasopancreatic drainage [ENPD] catheter), lesion site (pancreatic head, body/tail), and lesion size (≤10 mm, 10-20 mm, >20 mm), and compared with the diagnostic performance of EUS-FNA. RESULTS: The overall sensitivity of ERP-based cytology was 54.9%; sensitivity by the sampling method was 34.7% for PJC during ERP, 65.8% for brush cytology, and 30.8% for PJC via an ENPD catheter. The sensitivity of EUS-FNA was 85.3%. Brush cytology and PJC via an ENPD catheter were performed more often in pancreatic body/tail lesions than in head lesions (P = 0.016 and P < 0.001, respectively), and the overall sensitivity of ERP-based cytology was better for body/tail lesions (63.2% vs. 49.0%, P = 0.025). The sensitivities of ERP-based cytology and EUS-FNA in diagnosing PDAC ≤10 mm were 92.3% and 33.3%, respectively. Post-ERP pancreatitis was observed in 22 patients (8.2%) and significantly less common with ENPD catheters (P = 0.002). CONCLUSIONS: ERP-based cytology may be considered the first choice for pathological diagnosis of PDAC ≤10 mm and in the pancreatic body/tail.

Utility of cytopathological specimens and an automated image analysis for the evaluation of HER2 status and intratumor heterogeneity in breast carcinoma.

OBJECTIVES: Although updated HER2 testing guidelines have been improved by a collaboration between the American Society of Clinical Oncology (ASCO) and the College of American Pathologists (CAP) in 2013, HER2 evaluation is still problematic because of issues involving CEP17 polysomy, heterogeneity, and HER2 score 2+ cases. The aim of this retrospective study was to evaluate the relationship between HER2 gene heterogeneity, or so called CEP17 polysomy, using breast carcinoma cells sampled by scraping and the IHC score graded by automated image analysis using whole slide image. MATERIAL AND METHODS: We randomly selected 23 breast carcinoma cases with a HER2 score 0, 24 cases with a HER2 score 1+, 24 cases with HER2 score 2+, and 23 cases with HER2 score 3+ from the records of patients with breast cancer at Hiroshima University Hospital. We compared the results of fluorescent in situ hybridization (FISH) using formalin-fixed, paraffin-embedded (FFPE) tissues and cytological samples and compared the HER2 score calculated using an automated image analysis using wholly scanned slide images and visual counting. RESULTS: We successfully performed the FISH assay in 78 of 94 cases (83%) using FFPE tissues and in all 94 (100%) cases using cytological samples. Frequency of both HER2 amplification and CEP17 polysomy was higher when cytological samples were used than when FFPE tissue was used. Frequency of HER2 heterogeneity using cytological samples was higher that than using FFPE tissue, except for the IHC score 3+ cases. CONCLUSIONS: When assessment of HER2 status based on FISH using FFPE tissue cannot be accomplished, FISH using cytological samples should be considered. When intensity of HER2 is heterogeneous in the tumor tissue, particularly in cases regarded as score 2+, they should be evaluated by automated image analysis using the whole slide image.

An evaluation of the diagnostic and prognostic significance of p16(INK4a) /p21(WAF1/Cip1) immunostaining in squamous intraepithelial lesions of the uterine cervix using liquid-based cytology specimens.

Human papillomavirus (HPV) infection frequently causes squamous intraepithelial lesions (SIL) of the uterine cervix and consequently gives rise to squamous cell carcinoma. It is therefore important to identify cases that potentially develop higher grades of SIL at an early stage of the disease. In this study, we thus investigated whether immunocytochemistry for p21(WAF1/Cip1) and p16(INK4a) could be applicable in the diagnosis and the prognostic prediction of SIL in combination with genomic analyses of HPV. The genomic analysis of high-risk HPV (hrHPV), which was done by reversed dot blotting and by in situ hybridization, and immunocytochemistry were performed on liquid-based cytological specimens. A cross-sectional study comprising 145 cases of NILM, ASC-US, LSIL, and HSIL indicated that the incidence of the positive cases for p16(INK4a) and p21(WAF1/Cip1) and hrHPV increased with the grade of SIL. A double positive status for p16(INK4a) and p21(WAF1/Cip1) was a significant discriminator between HSIL and LSIL/NILM, even when applied in conjunction with the genomic test for hrHPV (P = 0.006 by logistic regression analysis). However, a prospective study employing 61 NILM/ASC-US cases, revealed that the p16(INK4a) /p21(WAF1/Cip1) immunostaining was not a significant predictor for the progression of SIL, whereas the cytological diagnosis (NILM vs. ASC-US) and the infection status of hrHPV conferred significant effects on the prognosis. Immunostaining of p16(INK4a) and p21(WAF1/Cip1) provides additional information on the cytological diagnosis of SIL. A further analysis using a larger population is warranted to obtain a conclusive result regarding the prognostic significance of p16(INK4a) /p21(WAF1/Cip1) immunocytochemistry in the diagnosis of SIL.

Tyrosine1248-phosphorylated HER2 expression and HER2 gene amplification in female invasive ductal carcinomas.

BACKGROUND: Phosphorylated HER2 (pHER2) may more accurately reflect the signaling and functional activity of the HER2 protein than detection of HER2 itself. The detection of HER2 gene amplification using fluorescence in situ hybridization (FISH) provides superior prognostic information for the diagnosis of breast cancer. However, the relationship between pHER2 expression in tissue samples and HER2 gene amplification remains unclear. METHODS: A total of 210 cases were recruited. The expression of HER2 and tyrosine (Tyr)1248-pHER2 was investigated by immunohistochemistry, and HER2 gene amplification was analyzed by FISH. Spearman's rank correlation test was employed to confirm correlation between HER2 and Tyr1248-pHER2. Chi-square and Student's t test were used to determine a significant difference between the baseline characteristics of tumors and the FISH, HER2 and Tyr1248-pHER2 results. The phosphorylation rate of HER2 was calculated using a digital-analysis system. RESULTS: HER2 expression was significantly (P < 0.001) associated with Tyr1248-pHER2 expression. HER2 gene amplification could be detected in 55 (26.2%) of the 210 tumors; 40 were HER2 positive and 32 were Tyr1248-pHER2 positive. The sensitivity and specificity of HER2 and Tyr1248-pHER2 for HER2 gene amplification were 72.7 and 58.2%, and 91.6 and 95.5%, respectively. In cases with an HER2 score of 2, and a phosphorylation score of 2 or 3, gene amplification was observed in 4 (80.0%) out of 5 tumors. CONCLUSIONS: Tyr1248-pHER2 expression is highly specific for HER2 gene amplification. The phosphorylation status might provide an adjunct to the assessment of gene amplification in patients with an HER2 score of 2.