HCC Screening Using DNA Methylation Changes in ctDNA

Clinical Trials on Detection of Hepatocellular Carcinoma With Non-invasive Method Based on DNA Methylation of Circulated Tumor DNA, PBMC and T Cells

Hepatocellular Carcinoma (HCC) is the fifth most common cancer world-wide. It is particularly prevalent in Asia, and its occurrence is highest in areas where hepatitis B is prevalent, indicating a possible causal relationship. Follow up of high-risk populations such as chronic hepatitis patients and early diagnosis of transitions from chronic hepatitis to HCC would improve cure rates. In most cases HCC is detected late resulting in increased mortality and morbidity.

The purpose of this study is to develop and test non-invasive biomarkers based on methylation changes in PBMC and circulated tumor DNA in hepatocellular carcinomas patients.

Study Overview

• Status: Completed
• Conditions: Hepatocellular Carcinoma
• Intervention / Treatment: Diagnostic test: ctDNA methylation in and it's Correlation wth Development and prediction of HCC

• Study Type: Observational
• Enrollment (Actual): 403

Contacts and Locations

Study Locations

• Bangladesh
  • Dhaka, Bangladesh
    • Infectious Diseases Division

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 18 years and older (Adult, Older Adult)
• Accepts Healthy Volunteers: No

• Sampling Method: Non-Probability Sample

Study Population

HCC staging will be diagnosed according to EASL-EORTC Clinical Practice Guidelines: Management of hepatocellular carcinoma. The patients will be divided into four groups, including Stage 0 (1) (n=100), stage A (2) (n=100), stage B (3) (n=100) and stage C+D (4) (n=100). As controls we will recruit chronic hepatitis B (n=100), which diagnose will be confirmed using AASLD practice guideline for chronic Hepatitis B, and healthy sex and age matched controls (n=100).

Description

Inclusion Criteria:

• Confirmed diagnosis of HCC by EASL-EORTC guidelines
• Confirmed hepatitis B diagnosis for HepB patients using AASLD practice guidelines.

Exclusion Criteria for HCC:

• Cirrhosis, any other known inflammatory disease (bacterial or viral infection with the exception of hepatitis B or C, diabetes, asthma, autoimmune disease, active thyroid disease) which could alter T cells and monocytes characteristics
• Other cancers.

Exclusion Criteria for HepB:

• Diagnosis of HCC or any other cancer.

Exclusion Criteria for Healthy Controls:

• Any known inflammatory or infectious disease including HepB and HepC
• Chronic diseases,
• Cancer
• Medications or drug use

Study Plan

How is the study designed?

Number of groups / cohorts

3

Cohorts and Interventions

Group / Cohort	Intervention / Treatment
chronic hepatitis B; This group will include 50 with hepatitis B subjects and the diagnoses will be based on AASLD practice guideline.	Diagnostic test: ctDNA methylation in and it's Correlation wth Development and prediction of HCC; Blood sample from patients with HCC, healthy individuals and individuals with hepatitis B will be collected, and DNA will extracted from PBMC and circulated tumor DNA will be subjected to bisulfite conversion. DNA from PBMC DNA will be analyzed with primers developed for the AHNAK-STAP1 genes. Plasma samples will be subjected to EZ direct DNA extraction and bisulfite conversion kit and will be amplified with primers developed to amplify the target regions. DNA will be used for the subsequent PCR amplification with specific primer to generate PCR amplicon for sequencing using a double PCR procedure. The product of PCR reaction will be subjected to indexed MiSeq Next-Generation sequencing that will allow us quantify DNA methylation level.
HCC Cases; This group will include 350 in stage 0, stage A, stage B, Stage C+D of hepatocellular carcinoma. HCC staging will be diagnosed according to EASL-EORTC Clinical Practice Guidelines: Management of hepatocellular carcinoma	Diagnostic test: ctDNA methylation in and it's Correlation wth Development and prediction of HCC; Blood sample from patients with HCC, healthy individuals and individuals with hepatitis B will be collected, and DNA will extracted from PBMC and circulated tumor DNA will be subjected to bisulfite conversion. DNA from PBMC DNA will be analyzed with primers developed for the AHNAK-STAP1 genes. Plasma samples will be subjected to EZ direct DNA extraction and bisulfite conversion kit and will be amplified with primers developed to amplify the target regions. DNA will be used for the subsequent PCR amplification with specific primer to generate PCR amplicon for sequencing using a double PCR procedure. The product of PCR reaction will be subjected to indexed MiSeq Next-Generation sequencing that will allow us quantify DNA methylation level.
Healthy; This group will include 50 healthy sex and age matched controls.	Diagnostic test: ctDNA methylation in and it's Correlation wth Development and prediction of HCC; Blood sample from patients with HCC, healthy individuals and individuals with hepatitis B will be collected, and DNA will extracted from PBMC and circulated tumor DNA will be subjected to bisulfite conversion. DNA from PBMC DNA will be analyzed with primers developed for the AHNAK-STAP1 genes. Plasma samples will be subjected to EZ direct DNA extraction and bisulfite conversion kit and will be amplified with primers developed to amplify the target regions. DNA will be used for the subsequent PCR amplification with specific primer to generate PCR amplicon for sequencing using a double PCR procedure. The product of PCR reaction will be subjected to indexed MiSeq Next-Generation sequencing that will allow us quantify DNA methylation level.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Calculation of M Scores and HCC Probability Scores	We normalized median methylation values (range 0-100) for 'HCC-detect' (Hepatocellular Carcinoma Detection) and 'HCC-spec' (Hepatocellular Carcinoma Specificity). 'HCC-detect' is derived from CHFR, VASH2, CCNJ, and GRID2IP regions, aiming to broadly identify HCC. Theoretical range is -6.6438 to 8.6438, with observed -3.541 to 7.65; higher scores indicate increased HCC likelihood. 'HCC-spec', focusing on the F12 region, differentiates HCC from 31 other cancers and normal cells, with theoretical range -3.3219 to 6.64385 (observed -3.3219 to 6.6297). Higher scores signify greater HCC specificity. Both scores, modeled via logistic regression in Prism, predict HCC probability, linking higher scores with higher HCC likelihood or specificity	6 months to 1 year

Collaborators and Investigators

• Sponsor: HKGepitherapeutics
• Collaborators: International Centre for Diarrhoeal Disease Research, Bangladesh
• Investigators: Principal Investigator: Wasif Ali Khan, PhD, nternational Centre for Diarrhoeal Disease Research

Study record dates

Study Major Dates

• Study Start (Actual): August 20, 2018
• Primary Completion (Actual): June 1, 2020
• Study Completion (Actual): November 1, 2020

Study Registration Dates

• First Submitted: March 20, 2018
• First Submitted That Met QC Criteria: March 28, 2018
• First Posted (Actual): March 30, 2018

Study Record Updates

• Last Update Posted (Actual): July 5, 2024
• Last Update Submitted That Met QC Criteria: July 2, 2024
• Last Verified: July 1, 2024

More Information

Terms related to this study

• Additional Relevant MeSH Terms: Digestive System Diseases; Neoplasms by Histologic Type; Neoplasms; Neoplasms by Site; Adenocarcinoma; Neoplasms, Glandular and Epithelial; Digestive System Neoplasms; Liver Diseases; Liver Neoplasms; Carcinoma; Carcinoma, Hepatocellular
• Other Study ID Numbers: HKG-Bng-HCC-100

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: UNDECIDED

Drug and device information, study documents

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