Peg-Interferon Lambda Treatment Induces Robust Innate and Adaptive Immunity in Chronic Hepatitis B Patients

Abstract

IFN-lambda (IFNλ) is a member of the type III IFN family and is reported to possess anti-pathogen, anti-cancer, and immunomodulatory properties; however, there are limited data regarding its impact on host immune responses in vivo. We performed longitudinal and comprehensive immunosurveillance to assess the ability of pegylated (peg)-IFNλ to augment antiviral host immunity as part of a clinical trial assessing the efficacy of peg-IFNλ in chronic hepatitis B (CHB) patients. These patients were pretreated with directly acting antiviral therapy (entecavir) for 12 weeks with subsequent addition of peg-IFNλ for up to 32 weeks. In a subgroup of patients, the addition of peg-IFNλ provoked high serum levels of antiviral cytokine IL-18. We also observed the enhancement of natural killer cell polyfunctionality and the recovery of a pan-genotypic HBV-specific CD4⁺ T cells producing IFN-γ with maintenance of HBV-specific CD8⁺ T cell antiviral and cytotoxic activities. It was only in these patients that we observed strong virological control with reductions in both viral replication and HBV antigen levels. Here, we show for the first time that in vivo peg-IFNλ displays significant immunostimulatory properties with improvements in the main effectors mediating anti-HBV immunity. Interestingly, the maintenance in HBV-specific CD8⁺ T cells in the presence of peg-IFNλ is in contrast to previous studies showing that peg-IFNα treatment for CHB results in a detrimental effect on the functionality of this important antiviral T cell compartment.

Clinical trial registration: ClinicalTrials.gov NCT01204762.

Keywords: direct antiviral; hepatitis B; immunity; in vivo; peg-interferon lambda.

Figure 1

Study design. All 13 patients were treated with entecavir (ETV) for 12 weeks and subsequently received ETV + pegIFN-λ. The weeks of treatment reached by the patients are shown. Peripheral blood mononuclear cells (PBMC) collection are also indicated.

Figure 2

Effect of treatment on natural killer (NK) cells response in Group 1 and Group 2 patients (n = 13). Percentage of (A) tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-positive CD56^bright NK cells (B) CD107a-producing cytotoxic CD56^dim NK cells, (C) IFN-γ producing CD56^bright, (D) NKG2D-positive CD56^bright, and (E) NKG2D-positive CD56^dim were measured by flow cytometry. A total of 100,000 events were collected during FACS acquisition and the subsequent analysis was performed using FACS DIVA software. Data are shown as mean ± SEM. Two-way ANOVA followed by multiple comparison tests were performed for statistical analysis.

Figure 3

Effect of treatment on HBV-specific T cells and T regs response in Group 1 and Group 2 patients (n = 13). The frequency of IFN-γ-producing HBV-specific T cells was evaluated by ELISPOT following peripheral blood mononuclear cells (PBMC) stimulation with HBV antigens and HBV-specific overlapping peptides. PBMC stimulation with recall antigen purified protein derivate and mitogen phytohemagglutinin elicited a measurable strong response which did not change significantly during the course of the treatment. (A) ELISPOT quantitation of frequency of IFN-γ-producing HBV-specific T cells. (B) Heat map representation of the percentage of patients reacting to each individual HBV antigen and peptide pool in ELISPOT. The assessment of the functionality of T cells was performed by FACS following two rounds of stimulation with HBV antigens and HBV-specific overlapping peptides covering HBV core and HBV surface regions. (C) IFN-γ-producing HBV-specific CD4⁺ T cells, (D) IFN-γ-producing HBV-specific CD8⁺ T cells, (E) CD107a-producing HBV-specific CD8⁺ T cells, and (F) T regulatory cells were quantitated by FACS. A total of 100,000 events were collected during FACS acquisition and the subsequent analysis was performed using FACS DIVA software. Data are shown as mean ± SEM. Two-way ANOVA followed multiple comparison tests were performed for statistical analysis.

Figure 4

Effect of treatment on serum cytokines production during treatment in patients group 1 and group 2 (n = 13). Cytokines (A) IL-18, (B) IL-8, (C) IL15, (D) IL-17, (E) IP-10, (F) IFN-α, and (G) IFN-β were measured in the sera of patients by cytometric bead array or ELISA. Data are shown as mean ± SEM. Two-way ANOVA followed by multiple comparison tests were performed for statistical analysis.