Anco Molijn

Molecular and pathological basis of HPV‐negative cervical adenocarcinoma seen in a global study

AbstractInternational surveys find HPV‐negativity in up to 30% of cervical adenocarcinomas. We investigated the pathological diagnosis by expert consensus with immunohistochemistry and the presence of somatic mutations in recognised tumour genes in HPV‐positive and negative cervical adenocarcinomas (CADC). A sample was selected of 45 paraffin‐embedded cervical blocks diagnosed locally as usual cervical adenocarcinoma from a global study. These represented different diagnoses made at previous diagnostic review and HPV status. All were suitable for analysis for somatic tumour associated gene mutations. Three pathologists examined H/E slides and immunohistochemistry for p16, progesterone receptor and p53 and classified the cases. L1 genes from high‐risk HPVs and low‐risk HPVs were analysed by SPF10 PCR‐DEIA‐LiPA25 version 1 in whole tissue sections and microdissected tumour and retested by PCR for E6/E7 genes of hrHPVs if negative. Cases were analysed for microsatellite instability and next‐generation sequencing mutation analysis. From the 45 cases, 20 cases of usual CADC were confirmed of which 17 (85%) were HPV‐positive in tumour cells. The other 25 cases were reclassified as endometrial, serous, clear‐cell and gastric‐type adenocarcinomas and all were HPV‐negative in tumour cells. Careful retesting for HPV DNA and IHC leads to more accurate identification of HPV‐positive usual cervical adenocarcinomas. Endometrioid endometrial adenocarcinomas, other uterine adenocarcinoma with multiple somatic mutations were important in misclassification of HPV‐negative cases locally managed as cervical adenocarcinoma, as was gastric‐type adenocarcinoma with germline STK11 mutation in East Asia. Few consensuses confirmed HPV‐negative usual cervical adenocarcinomas showed somatic tumorigenic mutations also seen in some HPV‐positive usual CADC.

Performance of DNA methylation analysis of ASCL1, LHX8, ST6GALNAC5, GHSR, ZIC1 and SST for the triage of HPV‐positive women: Results from a Dutch primary HPV‐based screening cohort

AbstractMethylation of host‐cell deoxyribonucleic acid (DNA) has been proposed as a promising biomarker for triage of high‐risk (hr) human papillomavirus (HPV) positive women at screening. Our study aims to validate recently identified host‐cell DNA methylation markers for triage in an hrHPV‐positive cohort derived from primary HPV‐based cervical screening in The Netherlands. Methylation markers ASCL1, LHX8, ST6GALNAC5, GHSR, ZIC1 and SST were evaluated relative to the ACTB reference gene by multiplex quantitative methylation‐specific PCR (qMSP) in clinician‐collected cervical samples (n = 715) from hrHPV‐positive women (age 29‐60 years), who were enrolled in the Dutch IMPROVE screening trial (NTR5078). Primary clinical end‐point was cervical intraepithelial neoplasia grade 3 (CIN3) or cancer (CIN3+). The single‐marker and bi‐marker methylation classifiers developed for CIN3 detection in a previous series of hrHPV‐positive clinician‐collected cervical samples were applied. The diagnostic accuracy was visualised using receiver operating characteristic (ROC) curves and assessed through area under the ROC curve (AUC). The performance of the methylation markers to detect CIN3+ was determined using the predefined threshold calibrated at 70% clinical specificity. Individual methylation makers showed good performance for CIN3+ detection, with highest AUC for ASCL1 (0.844) and LHX8 (0.830). Combined as a bi‐marker panel with predefined threshold, ASCL1/LHX8 yielded a CIN3+ sensitivity of 76.9% (70/91; 95% CI 68.3‐85.6%) at a specificity of 74.5% (465/624; 95% CI 71.1‐77.9%). In conclusion, our study shows that the individual host‐cell DNA methylation classifiers and the bi‐marker panel ASCL1/LHX8 have clinical utility for the detection of CIN3+ in hrHPV‐positive women invited for routine screening.

Comparison Of Cytology And Molecular Screening For Detecting Cervical Reactive Cellular Changes In General Population

This study compares the efficacy of cytology (Pap smear) with the molecular screening in their ability to detect reactive cellular changes in the cervix among an open population