The Role of FAM19A4 and Hsa-mir-124 Methylation in Predicting Prognosis of Untreated Cervical Intraepithelial Neoplasia 2 (CIN 2)

High-risk precancerous cervical lesions are divided into stage 2 and 3 cervical intraepithelial neoplasia (CIN 2 and 3). CIN 3 represents a direct pre-stage of invasive cancer, has a high rate of progression and a high degree of agreement with the final histological diagnosis. In CIN 2 lesions, the rate of agreement with the final histological diagnosis is lower and the rate of spontaneous regression is higher. Due to the higher rate of regression and possible complications after excisional treatment, conservative active monitoring can be considered in selected young CIN 2 patients. A recent meta-analysis reported a high rate of spontaneous clinical regression of CIN 2, particularly in women under 30 years old. There are currently no prospectively validated prognostic biomarkers to determine which CIN 2 will progress to higher grade and which will regress to lower grade of change. Recent research has studied HPV methylation and microbiome analysis as biomarkers. A number of studies have shown that host cell DNA methylation levels in cervical scrapes increase with underlying cervical disease severity and are highest in cervical cancer. DNA methylation involves the covalent binding of a methyl group to the 5´ position of a cytosine molecule in CpG dinucleotides. Besides global hypomethylation, the overall loss of methylation during carcinogenesis, resulting in chromosomal instability, and the silencing of tumour suppressor genes by local hypermethylation of CpG-rich promoter regions contribute to cancer development. Gene promoter methylation can be easily accessed by sensitive, quantitative methylation-specific PCR providing an objective test outcome. The aim of this study was to determine the effect of the methylation rate of two suppressor genes- FAM19A4 and hsa-mir-124 on the rate of CIN 2 regression, persistence or progression in women younger than 36 years (≤35 years old).

Study Overview

• Status: Recruiting
• Conditions: Cervical Intraepithelial Neoplasia Grade 2; DNA Methylation
• Intervention / Treatment: Diagnostic test: Testing DNA methylation test for predicting prognosis of untreated CIN 2

• Study Type: Interventional
• Enrollment (Anticipated): 100
• Phase: Not Applicable

Contacts and Locations

• Study Contact: Name: Milena Miklus, MD; Phone Number: +386 2 321 2178; Email: milena.rmus4@gmail.com

Study Locations

• Slovenia
  • Maribor, Slovenia, 2000
    • Recruiting
    • University Medical Centre Maribor
    • Contact: Milena Miklus, MD

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 20 years to 36 years (Adult)
• Accepts Healthy Volunteers: No
• Genders Eligible for Study: Female

Description

Inclusion Criteria:

• Histologically confirmed CIN 2 (with biopsy of colposcopically suspicious changes in the cervix)
• Age under 30 years
• Satisfactory colposcopy (transformation zone fully visible)
• Size of change below 75% of transformation zone
• The change in the ectocervix is fully visible
• Age 30-35 years, if the patient is non-smoker and the change in the cervix does not exceed 50% of the area of the transformation zone
• Signing an informed consent to participate in the survey
• Willingness to perform inspections every 6 months

Exclusion Criteria:

• Age 36 years or older
• Unsatisfactory colposcopy (transformation zone not fully visible)
• Size of change exceeds 75% of the transformation zone
• The change in the ectocervix is not completely visible
• Age 30-35 years for smokers or if the change exceeds 50% of the area of the transformation zone
• Suspicted glandular precancerous changes
• Histologically verified CIN 2 with cytological changes of glandular cells
• Colposcopically suspected invasive disease
• Histologically verified CIN 2 and histologically verified AIS
• Histologically verified CIN 2 and histologically verified invasive cancer elsewhere in the cervix
• Refusal to sign participation in the survey
• Unwillingness to perform control examinations
• Weakness of an immune system
• Cervical conization performed in the past
• Treatment with local immunomodulators

Study Plan

How is the study designed?

Design Details

• Primary Purpose: Diagnostic
• Allocation: N/A
• Interventional Model: Single Group Assignment
• Masking: None (Open Label)

Number of Arms

1

Arms and Interventions

Participant Group / Arm

Intervention / Treatment

Experimental: Women with CIN 2 and under 36 years old; We will analyze patients under 36 years old (≤ 35) with histological confirmed CIN 2 lesions and without any additional risk factors.

Diagnostic test: Testing DNA methylation test for predicting prognosis of untreated CIN 2; After being diagnosed with CIN 2, patients will first be contacted by telephone and invited to participate in the study. If patients agree to participate in the research, they will sign a consent to participate in the research. After that, we will perform a colposcopy and take a cervical swab for analysis with the QIAsure Methylation Test Kit (Qiagen, Gaithersburg, USA), which will determine the methylation of tumor suppressor genes FAM19A4 and has-mir-124. Patients will complete a questionnaire. The total duration of tracking in both groups will be two years. The QIAsure Methylation Test will be performed to analyze methylation. It is a methylation-specific PCR test that detects hypermethylation of the tumor promoter suppressor genes FAM19A4 and has-mir-124. The samples on which we will use this test are bisulfite-converted DNA obtained by triage test for high-risk HPV - Hybrid Capture 2 HPV DNA Test (hc2, Qiagen, Gaithersburg, USA).

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Impact of the degree of methylation on progression of CIN 2	The primary clinical outcome will be the effect of the degree of methylation on the progression of CIN 2 to CIN 3+ (CIN 3 and cervical cancer).	two years

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Rate of progression of CIN 2	The secondary clinical outcome will be the rate of progression of CIN 2 to CIN 3+, the rate of clinical decline to normal histological picture (˂ CIN 1) and the rate of persistence of precancerous change after two years (CIN 2 or persistent CIN 1).	two years

Collaborators and Investigators

• Sponsor: University Medical Centre Maribor

Study record dates

Study Major Dates

• Study Start (Actual): September 1, 2021
• Primary Completion (Anticipated): September 1, 2023
• Study Completion (Anticipated): May 1, 2024

Study Registration Dates

• First Submitted: November 14, 2022
• First Submitted That Met QC Criteria: November 14, 2022
• First Posted (Actual): November 22, 2022

Study Record Updates

• Last Update Posted (Actual): November 22, 2022
• Last Update Submitted That Met QC Criteria: November 14, 2022
• Last Verified: May 1, 2022

More Information

Terms related to this study

• Keywords: CIN 2 prognosis; DNA hypermethylation; FAM19A4; hsa-mir-124; tumor suppressor genes; diagnostic implications
• Additional Relevant MeSH Terms: Neoplasms by Histologic Type; Carcinoma; Neoplasms, Glandular and Epithelial; Neoplasms; Carcinoma in Situ; Cervical Intraepithelial Neoplasia
• Other Study ID Numbers: IRP-2021/02-04

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: Yes
• IPD Sharing Supporting Information Type: Study Protocol; Statistical Analysis Plan (SAP); Informed Consent Form (ICF); Clinical Study Report (CSR); Analytic Code

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No