Molecular evaluation of a multiplex methylation panel for epigenetic analysis of FNAB samples from Greek patients with suspicious breast lesions.

PURPOSE: Aberrant DNA methylation in promoter regions has been found in many cancers, including breast cancer (BC). A Methylation Specific PCR (MSP) was applied in breast Fine Needle Aspiration Biopsy (FNAB) material, which has been rarely used in the literature, to estimate the methylation frequencies of CND2, APC, HIN1 & CDH13 and to assess whether this multiplex methylation panel can be possibly used as an indicator-biomarker for BC detection in a Greek population. METHODS: A total of 104 participants were subjected to FNAB and both cytological evaluation and epigenetic analysis were carried out. DNA was extracted from FNAB samples and was subjected to bisulfite conversion. MSP was carried out with primers specific for either the methylated or unmethylated status for each gene. The final MSP products were analyzed in 2% agarose gels with electrophoresis. RESULTS: Hypermethylation was observed in 74%, 69.2%, 59.6% and 63.4% of the samples for CND2, HIN1, APC and CDH13, respectively. CND2 was the most hypermethylated in C5 cases (90%) and APC and HIN1 in C4 cases (88.2%). A significant correlation between histologic evaluation and the methylation frequencies for all 4 genes was calculated (p<0.001). Odds ratio for breast malignancy was 8.267 for CND2, 5.235 for APC, 7.852 for HIN1 and 22.920 for CDH13, underlying that their methylation is positively related to breast malignancy. Also, it seems that the combination of all genes into a multiplex methylation panel has significantly higher SP and PPV than any single gene methylation. CONCLUSIONS: Our study shows that breast FNAB combined with methylation data from the collected aspirates has a promising potential as a biomarker for the early detection of BC risk in women with suspicious lesions.

Cervical, anal and oral HPV detection and HPV type concordance among women referred for colposcopy.

BACKGROUND: Infection with human papillomaviruses (HPVs) can cause benign and malignant tumours in the anogenital tract and the oropharynx both in men and women. The aim of the presented study was to investigate cervical, anal, and oral HPV-detection rates among women referred to colposcopy for abnormal Cervical Cancer (CaCx) screening results and assess the concordance of HPV-types among these anatomical sites. METHODS: Women referred to colposcopy at a single centre due to abnormal cytology, conducted for CaCx screening, were subjected to cervical Liquid-based Cytology (LBC) smear testing, anal and oral sampling. Routine colposcopy consisted in multiple biopsies and/or Endocervical Curettage (ECC). HPV-detection was performed by PCR genotyping in all three anatomical sites. In high-risk (hr) HPV-DNA positive samples either from anal canal or oral cavity, anal LBC cytology and anoscopy were performed, or oral cavity examination respectively. Descriptive statistics was used for the analysis of HPV-detection rates and phi-coefficient for the determination of HPV-positivity concordance between the anatomical sites. RESULTS: Out of 118 referred women, hr. HPV-DNA was detected in 65 (55.1%), 64 (54.2%) and 3 (2.5%) at cervix, anal canal and oral cavity respectively while low-risk HPV-DNA was detected in 14 (11.9%) and 11 (9.3%) at cervix and anal canal respectively. The phi-coefficient for cervix/anal canal was 0.392 for HPV16, 0.658 for HPV31, 0.758 for HPV33, - 0.12 for HPV45, 0.415 for HPV52 and 0.473 for HPV58. All values were statistically significant (p < 0.001). CONCLUSIONS: The results suggest that most HPV-types, high-risk and low-risk, detected in the cervix of women with prevalent cervical dysplasia, correlate with the ones detected in their anal canal. This particularly applies for the HPV-types included in the nonavalent HPV-vaccine (HPVs 6/11/16/18/31/33/45/52/58).