Methylation of the hsa-miR-124, SOX1, TERT, and LMX1A genes as biomarkers for precursor lesions in cervical cancer.

OBJECTIVES: The methylation profile of genes in precursor lesions in cervical cancer was characterized to improve screening techniques for high-grade intraepithelial neoplasia. METHODS: A total of 447 cervical cytology samples obtained from women who underwent colposcopy were examined. The cases were distributed as follows: (1) cervices without cervical intraepithelial neoplasia (CIN; n = 152); (2) cervices with a CIN grade of 1 (CIN 1; n = 147); and (3) cervices with a CIN grade of 2 or 3 (CIN 2/3; n = 148). The methylation pattern for a panel of 15 genes was analysed by quantitative methylation-specific PCR (qMSP) and compared between the groups (≤CIN 1 vs. CIN 2+). RESULTS: In the validation set, seven genes presented significantly different methylation profiles according to diagnosis, namely, DAPK1 (p = 0.001), EPB41L3 (p = 0.001), HIC1 (p = 0.028), hsa-miR-124-2 (p = 0.001), LMX1A (p = 0.001), SOX1 (p = 0.001), and TERT (p = 0.001). Six genes showed a significant increase in the frequency of methylation in the presence of hr-HPV, namely, DAPK1 (p = 0.001), EPB41L3 (p = 0.001), hsa-miR-124-2 (p = 0.001), LMX1A (p = 0.001), SOX1 (p = 0.001), and TERT (p = 0.001). The methylation of the hsa-miR-124 gene showed sensitivity and specificity (86.7% and 61.3%, respectively) similar to that of the HPV test (91.3% and 50.0%, respectively). The independent factors associated with the diagnosis of CIN 2+ and the methylation of the hsa-miR-124-2 (OR = 5.1), SOX1 (OR = 2.8), TERT (OR = 2.2), and LMX1A (OR = 2.0) genes were a positive test for hr-HPV (odds ratio [OR] = 5.5). CONCLUSIONS: Hypermethylation of the hsa-miR-124-2, SOX1, TERT, and LMX1A genes may be a promising biomarker for precursor lesions in cervical cancer regardless of the hr-HPV status.

The Role of Self-Collection by Vaginal Lavage for the Detection of HPV and High-Grade Intraepithelial Neoplasia.

OBJECTIVE: To compare the results of cervical cytology and high-risk HPV tests using samples obtained using two different collection modalities in a population of Brazilian women: self-collection (vaginal lavage) and cervical Pap testing. METHODS: We enrolled 204 women who were aged 18-64 years and had previously obtained abnormal cervical cytology test results; 83.8% of them agreed to participate. The sample was divided into two aliquots: one for the cytological study and one for the molecular analysis of high-risk HPV. RESULTS: Fifty-eight percent of the participants preferred to utilize self-collection as an alternative screening method. However, we noticed that the HPV positivity rate was significantly lower in self-collected samples when compared to those obtained using the conventional collection method (p = 0.035). The cytology tests of the samples obtained via self-collection were sensitive and had a positive predictive value and an area under the curve (AUC) that were significantly lower than those of the Pap test. However, the specificity and negative predictive value of these tests were similar. When compared with the HPV test, the self-collected samples demonstrated lower accuracy in predicting high-grade cervical intraepithelial neoplasia or worse, with a significantly lower sensitivity, positive predictive value, and AUC than the cervical Pap test samples. CONCLUSION: Self-collection by vaginal lavage is simple and well accepted by women. Due to its limitations, however, self-collection by lavage should be utilized with caution.

A Low-Cost HPV Immunochromatographic Assay to Detect High-Grade Cervical Intraepithelial Neoplasia.

OBJECTIVE: To evaluate the reproducibility and accuracy of the HPV16/18-E6 test. METHODS: The study population was comprised of 448 women with a previously abnormal Pap who were referred to the Barretos Cancer Hospital (Brazil) for diagnosis and treatment. Two cervical samples were collected immediately before colposcopy, one for the hr-HPV-DNA test and cytology and the other for the HPV16/18-E6 test using high-affinity monoclonal antibodies (mAb). Women with a histologic diagnosis of cervical intraepithelial neoplasia grade 2 or 3 were considered to be positive cases. Different strategies using a combination of screening methods (HPV-DNA) and triage tests (cytology and HPV16/18-E6) were also examined and compared. RESULTS: The HPV16/18-E6 test exhibited a lower positivity rate compared with the HPV-DNA test (19.0% vs. 29.3%, p<0.001) and a moderate/high agreement (kappa = 0.68, 95%CI: 0.60-0.75). It also exhibited a significantly lower sensitivity for CIN2+ and CIN3+ detection compared to the HPV-DNA test and a significantly higher specificity. The HPV16/18-E6 test was no different from cytology in terms of sensitivity, but it exhibited a significantly higher specificity in comparison to ASCH+. A triage test after HPV-DNA detection using the HPV16/18-E6 test exhibited a significantly higher specificity compared with a triage test of ASCH+ to CIN2+ (91.8% vs. 87.4%, p = 0.04) and CIN3+ (88.6% vs. 84.0%, p = 0.05). CONCLUSION: The HPV16/18-E6 test exhibited moderate/high agreement with the HPV-DNA test but lower sensitivity and higher specificity for the detection of CIN2+ and CIN3+. In addition, its performance was quite similar to cytology, but because of the structural design addressed for the detection of HPV16/18-E6 protein, the test can miss some CIN2/3+ lesions caused by other high-risk HPV types.

Cervicovaginal cytology in patients undergoing pelvic radiotherapy using the Focalpoint system: results from the RODEO study.

BACKGROUND: Evaluate the performance of the Focalpoint system in identifying and classifying cervical cytology alterations from samples collected from patients treated with Radiotherapy (RT). METHODS: The reproducibility of manual and automated screening by cytotechnologists using the BD Focalpoint GS Imaging System was examined. Samples were collected from May 2010 to August 2011. RESULTS: A total of 378 treated with RT and 8,967 cytology samples from patients without previous RT, were evaluated. The kappa values for cytological diagnoses read manually and automated in cases without previous RT were as follows: < ASC-H vs. ≥ ASC-H = 0.71; < LSIL vs. ≥ LSIL = 0.66; and < HSIL vs. ≥ HSIL = 0.67. The kappa for cytological diagnoses in post-RT women have showed: < ASC-H vs. ≥ ASC-H = 0.71; < LSIL vs. ≥ LSIL = 0.65; < HSIL vs. ≥ HSIL = 0.57. CONCLUSIONS: There was no significant difference among the kappa values we found. Post-RT cytology showed small diagnostic agreement between manual and automated examination. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/13000_2014_231.

Performance and reproducibility of gynecologic cytology interpretation using the FocalPoint system: results of the RODEO Study Team.

OBJECTIVES: To assess whether automated screening in the cytologic examination of Papanicolaou smear slides results in smaller margins of error than manual screening. METHODS: We compared cytotechnologists' performance and reproducibility of manual and automated screening of 10,165 consecutive cervical cytology slides examined at Barretos Cancer Hospital using the FocalPoint system. RESULTS: In total, 83% of atypical squamous cells of undetermined significance and greater were classified as quintiles 1 and 2; no high-grade squamous intraepithelial lesions and greater were observed in quintile 5. No statistically significant differences were found between manual and automated screening, using cervical biopsy specimens as the gold standard. CONCLUSIONS: FocalPoint safely screened high-grade lesions, which can be valuable for high-workload routines.