THU-204
NUCLEIC ACID-BASED POLYMERS EFFECTIVE AGAINST HEPATITIS B
VIRUS INFECTION IN PATIENTS DO NOT HARBOUR IMMUNE
STIMULATORY PROPERTIES IN PRIMARY ISOLATED BLOOD OR
LIVER CELLS
C. Real1, M. Werner1, A. Paul2, G. Gerken1, J. Schlaak1,3, A. Vaillant4,
R. Broering1. 1Deptartment of Gastroenterology and Hepatology;
2Deptartment of General-, Visceral- and Transplantation Surgery,
University Duisburg-Essen, Medical Faculty, Essen; 3Evangelisches
Klinikum Niederrhein GmbH, Duisburg, Germany; 4Replicor Inc.,
Montreal, Quebec, Canada
E-mail: [email protected]
Background and Aims: Nucleic acid polymers (NAPs) block the
release of HBsAg from infected hepatocytes in vivo. Although this
mechanism is likely responsible for the activity of NAPs against
hepatitis B virus (HBV) in patients, the role of potential
immunostimulatory effects have not been explored. In this study,
the immune stimulatory properties of NAPs were examined in
primary isolated human blood and parenchymal and nonparenchymal
liver cells.
Methods: Human peripheral blood mononuclear cells (PBMCs) and
primary isolated hepatocytes (PHH) and Kupffer cells (KC) were
treated with the following NAPs: REP 2006, the prototypic
degenerate NAP (dN)40, contains residual CpG (TLR-9 stimulatory)
content, REP 2055 is clinically active having a sequence (dAdC)20
devoid of CpG content and REP 2139 (also clinically active) and REP
2165 are REP 2055 analogues further rendered immunologically
inactive by replacing cytidine with 5-methylcytidine replaces and 2′-
O methylation of riboses. Immune responsiveness was confirmed
with known ligands for TLR-7 (poly I:C), TLR-3 (ssRNA) or TLR-9 (CpG
ODN2216). Total RNA was isolated and quantitative RT-PCR was
performed to analyse gene expression indicating antiviral (IFNA4,
IFNB1, IFNG, IFNL2) and inflammatory (TNF, IL6, and IL10) effects. The
intracellular uptake of CY3-labelled NAPs was confirmed using
fluorescence microscopy.
Results: REP 2006, REP 2139 and REP 2165 induced proinflammatory
responses in PBMCs but displayed no significant
antiviral activity. In PHH, no significant inflammatory or antiviral
responses were detected for any NAP. In KC, pro-inflammatory
activity (restricted to TNF) was observed with REP 2006 and REP
2055, whereas a weak but significant induction of interferon genes
(INFA and IFNL2) was only observed with REP 2006 at the highest
concentration. These signals were comparable to those induced by
ODN2216 stimulation.
Conclusions: The data presented here suggest that NAPs optimized
to treat hepatitis B virus infection in patients do not induce antiviral
responses in primary isolated blood or parenchymal and nonparenchymal
liver cells. We therefore hypothesize that the antiviral
activity of NAPs against HBV infection cannot be explained by direct
induction of innate antiviral responses. 作者: StephenW 时间: 2016-4-3 02:50