Background: MicroRNAs (miRNAs) are small, noncoding RNAs that negatively regulate gene expression by binding to target mRNAs. miRNAs have not been comprehensively studied in recurrent ovarian cancer, yet an incurable disease.

Aim: To examine the potential of p16INK4A as a biomarker for dysplastic squamous and glandular cells of the cervix in tissue sections and ThinPrep™ smears. Methods: Immunocytochemical analysis of p16INK4A expression was performed on 22 normal cervical tissue samples, five cervical glandular intraepithelial neoplasia (cGIN), 38 cervical intraepithelial neoplasia 1 (CIN1), 33 CIN2, 46 CIN3, and 10 invasive cancer cases (eight squamous and two adenocarcinomas). All samples were formalin fixed and paraffin wax embedded, and immunohistochemical analysis was carried out using a mouse monoclonal anti-p16 INK4A antibody after antigen unmasking. The staining intensity was assessed using a 0 to 3 scoring system. In addition, the expression status of p16 INK4A was examined in 12 normal ThinPrep smears, one smear exhibiting cGIN, and a total of 20 smears exhibiting mild, moderate, and severe dyskaryosis. Human papillomavirus (HPV) detection was carried out using a modified SYBR green assay system. Fluorogenic polymerase chain reaction (PCR) and solution phase PCR were used for specific HPV typing. Results: p16INK4A immunoreactivity was absent in all normal cervical tissues examined. Dysplastic squamous and glandular cells were positive for p16INK4A expression in all cases included in this study, except for one CIN3 case. p16INK4A expression was mainly nuclear in CIN1 cases, and both nuclear and cytoplasmic in CIN2, CIN3, cGIN, and invasive cases. All cases positive for HPV expressed the p16 INK4A protein, although not all cases found positive for p16 INK4A were HPV positive. In general, the p16 INK4A staining intensity was lower in cases negative for HPV or those containing a low risk HPV type. Conclusion: This pattern of overexpression demonstrates the potential use of p16INK4A as a diagnostic marker for cervical squamous and also glandular neoplastic lesions. In addition, the technique can be used to identify individual dyskaryotic cells in ThinPrep smears. Thus, p16 INK4A is a useful marker of cervical dyskaryosis. C ervical cancer is one of the most common forms of cancer in women worldwide. In developing countries, cancer of the uterine cervix is ranked second, with a relative frequency of 15% of all cancers in women, whereas in developed countries cervical cancer is ranked fifth, with a relative frequency of 4.4%.1 The Papanicolaou (Pap) test, as described by G Papanicolaou, is a cytological staining technique, which allows the identification of asymptomatic women who have preneoplastic lesions or early cancer of the uterine cervix. Although the introduction of mass screening programmes in developed countries has been effective in reducing cervical cancer mortality and morbidity rates, the success of the Pap smear test is limited with respect to sensitivity and specificity. False negative rates for cervical premalignant lesions and cervical cancer lie between 15% and 50% and false positive rates of approximately 30% have been reported.2 This failure may reflect the subjectivity of cytological diagnosis. In addition...

Aim: To analyse and compare expression patterns of three potential biomarkers-p16 INK4A , CDC6, and MCM5-and evaluate their use as predictive biomarkers in squamous and glandular cervical preinvasive neoplasia. Methods: Immunocytochemical analysis of p16 INK4A, MCM5, and CDC6 expression was performed on 20 normal, 38 cervical intraepithelial neoplasia 1 (CIN1), 33 CIN2, 46 CIN3, 10 squamous cell carcinoma, 19 cervical glandular intraepithelial neoplasia (cGIN), and 10 adenocarcinoma samples. Staining intensity was assessed using a 0-3 scoring system. p16INK4A , MCM5, and CDC6 expression was also examined in ThinPrep slides exhibiting mild, moderate, and severe dyskaryosis. Human papillomavirus (HPV) was detected using a modified SYBR green assay. Fluorogenic polymerase chain reaction (PCR) and solution phase PCR were used for specific HPV typing. Results: All three markers showed a linear correlation between expression and grade of dysplasia. p16INK4A and MCM5 protein expression was upregulated in all grades of squamous and glandular dysplasia. CDC6 protein was preferentially expressed in high grade lesions and in invasive squamous cell carcinoma. Conclusion: p16INK4A expression was closely associated with high risk HPV infection-all grades of squamous and glandular cervical lesions were immunohistochemically positive. MCM5 staining intensity was independent of high risk HPV infection, highlighting its potential as a biomarker in both HPV dependent and independent cervical dysplasia. CDC6 may be a biomarker of high grade and invasive lesions of the cervix, with limited use in low grade dysplasia. p16INK4A was the most reliable marker of cervical dysplasia. Combinations of dysplastic biomarkers may be useful in difficult diagnostic cases.

MicroRNAs are a group of small non-coding RNAs approximately 22 nucleotides in length. Recent work has shown differential expression of mature microRNAs in human cancers. Production and function of microRNAs require coordinated processing by proteins of the microRNA machinery. Dicer and Drosha (RNase III endonucleases) are essential components of the microRNA machinery. Recently, the ribosome anti-association factor eIF6 has also been found to have a role in microRNA-mediated post-transcriptional silencing. We characterized the alterations in the expression of genes encoding proteins of microRNA machinery in ovarian serous carcinoma. Protein expression of eIF6 and Dicer was quantified in a tissue microarray of 66 ovarian serous carcinomas. Dicer, Drosha and eIF6 mRNA expression was analysed using quantitative reverse transcription-PCR on an independent set of 50 formalin-fixed, paraffin-embedded ovarian serous carcinoma samples. Expression profiles of eIF6 and Dicer were correlated with clinicopathological and patient survival data. We provide definitive evidence that eIF6 and Dicer are both upregulated in a significant proportion of ovarian serous carcinomas and are associated with specific clinicopathological features, most notably low eIF6 expression being associated with reduced disease-free survival. The status of eIF6 and proteins of the microRNA machinery may help predict toxicity and susceptibility to future interfering RNA-based therapy.