p16(INK4a) is superior to high-risk human papillomavirus testing in cervical cytology for the prediction of underlying high-grade dysplasia.

BACKGROUND: The primary goal of this study was to compare the clinical performance of an optimized and rigorously controlled immunocytochemical (ICC) assay for p16(INK4a) to high-risk (HR) human papillomavirus (HPV) detection by polymerase chain reaction (PCR) as diagnostic adjuncts for cytology specimens from colposcopy patients. METHODS: : The study included 403 cervical cytology specimens collected within 3 months of colposcopy. The colposcopic impression and cervical biopsy diagnosis served as the standards for correlation with cytological, p16(INK4a), and HPV data. p16(INK4a) was evaluated using an immunoperoxidase-based assay that was linear over 4 logs for the detection of HeLa-spiked positive control cytology specimens, using a threshold for positive test results that was based on receiver operating characteristic curve analysis. HR-HPV was detected by multiplex PCR using genotype-specific primers. RESULTS: : In all combined diagnostic categories (negative for intraepithelial lesion and malignancy, atypical glandular cells, atypical squamous cells of undetermined significance, atypical squamous cells cannot exclude high-grade squamous intraepithelial lesion, low-grade squamous intraepithelial lesion, and high-grade squamous intraepithelial lesion), the p16(INK4a) ICC and HR-HPV assays, respectively, had sensitivity of 81.7% and 83.3% (P = .81) and specificity of 78.1% and 50.9% (P < .001) for the detection of underlying > or =grade 2 cervical intraepithelial neoplasia (CIN) lesions on biopsy. Furthermore, the positive predictive value of p16(INK4a) ICC was greater than that of HR-HPV for patients with biopsies > or =CIN-2 (41.2% and 24.2%, respectively, P = .001). CONCLUSIONS: : This p16(INK4a) immunocytochemical assay has superior specificity but similar sensitivity to HR-HPV testing to predict underlying high-grade dysplastic lesions in patients who are referred for colposcopy. The determination of the overall performance characteristics of p16(INK4a) immunocytochemistry, as an independent test or in combination with HPV testing in low-risk screening populations, however, will require subsequent large-scale prospective clinical trials.

Molecular mechanism of chemoresistance by miR-215 in osteosarcoma and colon cancer cells.

BACKGROUND: Translational control mediated by non-coding microRNAs (miRNAs) plays a key role in the mechanism of cellular resistance to anti-cancer drug treatment. Dihydrofolate reductase (DHFR) and thymidylate synthase (TYMS, TS) are two of the most important targets for antifolate- and fluoropyrimidine-based chemotherapies in the past 50 years. In this study, we investigated the roles of miR-215 in the chemoresistance to DHFR inhibitor methotrexate (MTX) and TS inhibitor Tomudex (TDX). RESULTS: The protein levels of both DHFR and TS were suppressed by miR-215 without the alteration of the target mRNA transcript levels. Interestingly, despite the down-regulation of DHFR and TS proteins, ectopic expression of miR-215 resulted in a decreased sensitivity to MTX and TDX. Paradoxically, gene-specific small-interfering RNAs (siRNAs) against DHFR or TS had the opposite effect, increasing sensitivity to MTX and TDX. Further studies revealed that over-expression of miR-215 inhibited cell proliferation and triggered cell cycle arrest at G2 phase, and that this effect was accompanied by a p53-dependent up-regulation of p21. The inhibitory effect on cell proliferation was more pronounced in cell lines containing wild-type p53, but was not seen in cells transfected with siRNAs against DHFR or TS. Moreover, denticleless protein homolog (DTL), a cell cycle-regulated nuclear and centrosome protein, was confirmed to be one of the critical targets of miR-215, and knock-down of DTL by siRNA resulted in enhanced G2-arrest, p53 and p21 induction, and reduced cell proliferation. Additionally, cells subjected to siRNA against DTL exhibited increased chemoresistance to MTX and TDX. Endogenous miR-215 was elevated about 3-fold in CD133+HI/CD44+HI colon cancer stem cells that exhibit slow proliferating rate and chemoresistance compared to control bulk CD133+/CD44+ colon cancer cells. CONCLUSIONS: Taken together, our results indicate that miR-215, through the suppression of DTL expression, induces a decreased cell proliferation by causing G2-arrest, thereby leading to an increase in chemoresistance to MTX and TDX. The findings of this study suggest that miR-215 may play a significant role in the mechanism of tumor chemoresistance and it may have a unique potential as a novel biomarker candidate.