Abstract
Chronic hepatitis B (CHB) infection is a disease affecting approximately 300 million people worldwide. Chronic hepatitis B is caused by infection of liver cells with hepatitis B virus (HBV) and kills more people than HIV, malaria and tuberculosis put together. HBV infection is predominantly contracted at birth from infected mothers. Although HBV infection is silent in childhood and causes no or minimal liver damage, it is a persistent viral infection, which leads to progressive liver disease in 40% of patients and accounts for 1 million deaths every year. Screening for HBV during pregnancy is widely available to reduce the risk of perinatal transmission, along with other precautions such as antiviral therapy during pregnancy, birth dose vaccination and application of hepatitis B immunoglobulin are also available. However, even with these preventative methods in place in most communities, still 10% of mothers with highly active virus levels pass the infection to their infants.This thesis investigates the role of serological and virological markers in predicting long term clinical outcomes in patients suffering from chronic hepatitis B virus infection who acquired it perinatally. I will particularly focus on novel markers: Hepatitis B core-related antigen (HBcrAg) and pregenomic Ribonucleic acid (pgRNA) which may be beneficial in HBV management and grant us a step towards curing HBV. Overall, I aim to evaluate the predictive nature of these markers in relation to liver damage progression and to identify their place in chronic HBV management and treatment.
In my first study I investigated whether pgRNA and HBcrAg could predict clinical outcomes in HBeAg negative chronic hepatitis B patients suppressed on nucleos(t)ide analogue therapy. Serum samples from patients who were diagnosed in childhood and followed up through adulthood were collected. Serological markers HBeAg, HBsAg, and HBcrAg and virological markers HBV DNA, and pgRNA were determined, along with liver function tests including ALT, APRI and FIB4 scores. Patients were divided into three cohorts based on their treatment status. The first cohort consisted of patients on maintenance suppressive therapy with tenofovir (TDF); the second cohort consisted of patients on long-term Nucleos(t)ide analogue (NA) suppressive therapy for at least three years, and in whom NA treatment was withdrawn before HBsAg loss. The third cohort consisted of patients with HBsAg loss on long-term suppressive NA therapy, in whom therapy was withdrawn after HBsAg loss. Results from the first cohort demonstrated that out of 66 patients treatment baseline HBcrAg could be detected in 71% (n=47) of patients and pgRNA could be detected in 83% (n=55) of patients. After 3 years of antiviral therapy 33% (n=22) of patients still had detectable HBcrAg in their serum and 30% (n=20) still had detectable pgRNA in their serum even after HBV DNA could no longer be detected. After 5 years of antiviral therapy HBcrAg could still be detected in 27% (n=18) of patients and pgRNA could be detected in 14% (n=9) of patients. In the second cohort (n=23), detectable levels of HBcrAg was observed in 17% (n=4) of patient serum and pgRNA was observed in 13% (n=3) of patient serum after nucleos(t)ide (NA) withdrawal. Additionally, pgRNA detection at the time of NA withdrawal correlated with ALT flares after stopping NA therapy. In the third cohort (n=19), at the follow up appointment HBsAg was undetectable in all patients after NA withdrawal. However, 11% (n=2) of patients had relapsed as they had detectable HBV DNA in their serum and had increased ALT activity but not detectable HBsAg levels. This group re-commenced NA therapy.
The aim of my second project was to determine whether HBcrAg and HBV pgRNA could be used to assess virological response and liver disease progression in patients who achieve HBeAg seroconversion versus those who do not while on Nucleos(t)ide analogue (NA) therapy. Other markers associated with CHB – Interferon-γ-inducible protein 10 (IP-10 or CXCL10) and Programmed cell death protein 1 (PD-1), were also determined at baseline, and at follow ups. 58 patient samples were collected at 3 different time points (at therapy baseline, at 5 years and 10 years of antiviral therapy). Results demonstrated that DNA suppression and ALT normalization improved steadily with NA therapy. HBcrAg and HBV pgRNA could still be detected in patients even after HBV DNA suppression and achieving anti-HBe status. This suggested that cccDNA was still active in hepatocytes and liver disease could still occur if antiviral therapy is discontinued. High levels of IP-10 and PD1 were associated with increased levels of liver fibrosis.
In my third study I investigated whether markers of cccDNA transcriptional activity: HBcrAg and pgRNA could help to predict future disease progression (need for therapy) in patients with perinatally acquired Chronic Hepatitis B infection followed from childhood to adulthood. Serological markers including HBeAg, HBsAg and HBcrAg and virological markers HBV DNA and pgRNA were compared between diagnosis and their last clinical visit (median follow-up duration 6 years, range 3-12 years). Liver function markers including ALT, AST, ALP, GGT, bilirubin and albumin were also compared. This cohort included 29 paediatric patients (range: 2-18 years old). It was found that pgRNA and HBcrAg were present in all patient serum at baseline (before the antiviral therapy initiation) and in particularly high concentrations in the patients that were placed on antiviral therapy. During antiviral therapy HBV DNA reduced significantly and markers of cccDNA transcriptional activity – HBcrAg and pgRNA also decreased substantially. Overall, the novel markers of cccDNA (HBcrAg and pgRNA) could be utilised with existing markers for predictive purposes to determine which patients are more likely to require antiviral therapy.
In my fourth study I investigated the clinical relevance of ultrasensitive HBsAg assay. Serum samples from 203 chronic hepatitis B (CHB) patients, all HBeAg negative were tested on the Abbott Architect and then tested using the CLEIA HBsAg-HQ assay (Fujirebio, Europe, Ghent, Belgium) on the Fujirebio Lumipulse platform. Results showed that the Lumipulse HBsAg-HQ assay could consistently determine HBsAg in all HBV genotypes tested and results were very similar to the Abbott assay (median HBsAg levels by Fujirebio :17.7, range 0.0001-150 IU/ml vs Abbott: 18.6, range 0.1-164 IU/mL). There was also a strong bi-variate correlation observed between the two assays (r=0.977, p<0.001).
| Date of Award | 1 Oct 2024 |
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| Original language | English |
| Awarding Institution |
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| Supervisor | Ivana Carey (Supervisor) & Emer Fitzpatrick (Supervisor) |