Performance of HPV E4 and p16^INK4a biomarkers in predicting regression of cervical intraepithelial neoplasia grade 2 (CIN2): protocol for a historical cohort study

Abstract

IntroductionCervical intraepithelial neoplasia grade 2 (CIN2) represents a spectrum of lesions with variable progression and regression. Pathological diagnosis of CIN2 is subjective and poorly reproducible. Accurate diagnosis and identification of different patterns of CIN2 related to outcome are essential to reduce the risks of overtreatment or undertreatment. It is important to explore novel methods for risk stratification of CIN2 to enable targeted treatment of women at high risk of progression or persistent disease and follow-up of women at low risk. The combination of the novel biomarker human papillomavirus (HPV) E4 with p16INK4a targets steps in the transition from a productive oncogenic HPV infection (CIN1) to a transformed lesion (CIN3) within CIN2. Previous cross-sectional studies suggest that HPV E4 combined with p16INK4a may be valuable for risk assessment of CIN2. However, data on HPV E4/p16INK4a as a predictor for CIN2 regression is lacking.Methods and analysisWe will conduct a historical cohort study including 500 women aged 23–40 years with a first CIN2 diagnosis in Aarhus, Denmark during 2000–2010. Women will be eligible if they have undergone active surveillance and have no previous record of hysterectomy, cone biopsy, and CIN2 or worse. Women will be randomly selected through the Danish Pathology Databank. Tissue samples from women included will be sectioned for p16INK4a and HPV E4 immunohistochemical staining in addition to conventional hematoxylin and eosin (H&E) staining. A positive result will be defined as HPV E4 positive. Through the Danish Pathology Databank, we will collect results on all subsequent cervical biopsies. Regression will be used as the primary outcome.Ethics and disseminationThe study has been approved by the Ethical Committee in Central Denmark Region (1-10-72-60-20) and registered at the Faculty of Health, Aarhus University. Results will be published in a peer-reviewed journal and presented at scientific meetings.Trial registration numberNCT05049252.

Validation of an on-chip p16ink4a/Ki-67 dual immunostaining cervical cytology system using microfluidic device technology

“…Primary screening for cervical cancer using HPV testing, which is currently used in Western countries, requires triaging to determine which patients who are positive for HPV should be referred for further testing 41 , 42 . Cytology diagnostic methods, which have been used for primary screening worldwide, can be used for triaging patients, p16/Ki-67 dual-stain cytology analyses, and testing HPVE6/E7 mRNAs, which are proposed as biomarkers for cervical cancer 43 – 45 .…”

High prevalence of HPV16 and high‐grade cytology in women undergoing active surveillance for cervical intraepithelial neoplasia grade 2

“…We conducted a single‐center cross‐sectional study on women undergoing active surveillance for CIN2. The source population consisted of women with a record of a community CIN2 diagnosis ( N = 3623) in the Danish Pathology Data Bank (DPDB) at the Department of Pathology, Aarhus University Hospital from January 1, 2000, to December 31, 2010 23 . The DPDB stores data on all cyto‐ and histopathological examinations since 1978, with data being complete since 1998 24 .…”

“…Of note, this study is a secondary analysis of data collected for the purpose of a historical cohort study aiming to explore various biomarkers for risk of regression in women undergoing active surveillance for CIN2. In the current cross‐sectional study, we present baseline characteristics on this CIN2 cohort 23 …”

“…The source population consisted of women with a record of a community CIN2 diagnosis ( N = 3623) in the Danish Pathology Data Bank (DPDB) at the Department of Pathology, Aarhus University Hospital from January 1, 2000, to December 31, 2010. 23 The DPDB stores data on all cyto‐ and histopathological examinations since 1978, with data being complete since 1998. 24 We identified women in the source population using systematized nomenclature of medicine codes for moderate dysplasia (M74B09) (i.e., CIN2) and cervical biopsy (T83110).…”

“…This study was approved by the Danish Scientific Ethical Committee in Central Denmark Region on July 17, 2020 (journal number 1‐10‐72‐60‐20) and registered at Aarhus University (2016‐051‐000001) on March 19, 2020. 23 As judged by the Danish Scientific Ethical Committee, it was not required to obtain a written informed consent from study participants since biological material had already been collected in relation to community diagnosis.…”