Marte Holmberg [1, 2], Dag H Reikvam [2, 3], Olav Dalgard [2, 4], Hans C Aass [3], Ellen Samuelsen [4], Niklas Björkström [5] and Asgeir Johannessen [1, 2].

Affiliates

Vestfold Hospital, Tønsberg, Norway [1], Institute of Clinical Medicine, University of Oslo, Oslo, Norway [2], Oslo University Hospital, Oslo, Norway [3], Akershus University Hospital, Lørenskog, Norway [4], Karolinska University Hospital, Stockholm, Sweden [5].

Abstract

Background:
Patients with HBeAg negative chronic hepatitis B may experience an immune response after stopping nucleos(t)ide analogue (NA) therapy, which may potentially trigger HBsAg loss (functional cure) or off-therapy sustained viral control. The immunological mechanisms that determine clinical outcome after treatment stop remain poorly understood, and predictors of clinical outcomes are lacking. We hypothesized that markers of systemic inflammation could identify key inflammatory pathways that are associated with and can predict clinical outcome after NA therapy cessation.

Methods:
Plasma cyto- and chemokines from 57 patients from the ongoing Nuc-Stop study were analyzed by Bio-Plex Pro Human Cytokine 27-plex Assay at baseline and three months after stopping NA therapy. All patients were HBeAg negative, non-cirrhotic, and virally suppressed for at least 24 months prior to stopping NA therapy. Outcome at three months was classified as immune control (HBV DNA ≤ 2000 IU/ml and ALT ≤ 80 U/L), viral relapse (HBV DNA > 2000 IU/ml and ALT ≤ 80 U/L), evolving clinical relapse (HBV DNA > 2000 IU/ml, ALT > 80 U/L and rising from week 8 to 12) and resolving clinical relapse (HBV DNA >2000 IU/ml, ALT >80 U/L and falling from week 8 to 12).

Clinical relapse was divided into evolving clinical relapse and resolving clinical relapse to capture the immunological dynamics during a flare. Statistical significance (p < 0.05) was determined using one-way ANOVA followed by Bonferroni post-hoc tests.

Results:
Three months after treatment cessation, there was a significant increase in interferon gamma-induced protein 10 (IP-10) in patients with evolving clinical relapse compared to the immune control and viral relapse groups (p = 0.001) (Figure 1A). A significant increase in tumor necrosis factor (TNF) compared to all other groups was also observed in patients with evolving clinical relapse (p = 0.007) (Figure 1B). There were no clinically significant changes in any of the other 25 cytokines associated with any of the four clinical outcomes

Conclusions:
Evolving clinical relapse after NA cessation was associated with specific increase in IP-10 and TNF. Further research is needed to evaluate whether these cytokines can distinguish beneficial flare from detrimental flare and predict longtime outcome in chronic hepatitis B.