Hepatitis Viruses Flashcards

Question

Genome gymnastics in HBV replication

Answer 1

1. Initiation at 5 ́ by P covalently linked to priming nucleotides 2. Switch of P-linked primer to DR1, annealing and ( -) strand DNA synthesis 3. (+)-RNA genome degradation by RNaseH (part of P) 4. 5 ́part of (+) RNA which is not degraded, serves as primer for (+)-DNA synthesis 5. Elongation till completion; in 5-10% of cases („in situ priming“; none productive way) 6. 90-95% of (+) DNA synthesis: Second template switch leads to elongation to 5 ́R sequence followed by 7. Third template switch by hybridization between copy of 5 ́R in (+)-DNA strand and 3 ́R of (-)-DNA strand (+)-strand synthesis continues for variable distance (-> incomplete (+) strand!) The resulting partially ds-DNA serves as genome found in mature virions Formation of stemloop in DR1 in (-) DNA „truncates“ sequence complementary to primer -> longer complementary sequence now in DR2 -> more stable base pairing with DR2 Thermodynamics determine the consecutive annealing steps!

Answer 2

Receptor candidates (binding to aa 21–47 of preS1) published so far: - human immunoglobulin A receptor - glyceraldehyde-3-phosphate dehydrogenase (GAPDH) - interleukin-6 - a 31-kDa protein HBV binding factor - asialoglycoprotein receptor - nascent polypeptide-associated complex α polypeptide - lipoprotein lipase - hepatocyte-associated heparan sulfate proteoglycans - glucose-regulated protein 75. The only bona fide receptor is NTCP (transporter for uptake of bile salts into hepatocytes)

Answer 3

PreS1 domain binds to NTCP receptor on hepatocyte Subcutaneous application of myristylated preS1 peptide in humanized uPA-mouse: - specific accumulation of peptide in liver - prevention of HBV infection of human liver cells - prevention of Hepatitis Delta infection Effective when a HBV-positive person gets a liver transplantation Peptide drug: Myrcludex B

Answer 4

- 1.7 kB RNA (neg. polarity) - needs Hepatitis B Virus capsid proteins for packaging * codes for hepatitis delta antigen (HD Ag; L and S form) * S-HD Ag important for genome replication, L for virion assembly * cell. RNA Pol II (DdRp) is involved in genome replication - genome slightly larger than in viroids - Coding one ORF but no capsid proteins - The virusoid-nucleic-acid is packaged by the capsid proteins of HBV - HD-Ag forms RNP with RNA genome; import into nucleus via NLS in HD-Ag - Rolling circle amplification leads to concatameric + strand (anti-genome) which is cut to genome size + RNAs by a ribozyme at the 3 ́end of the anti-genome; circularisation (ligation) - Rolling circle amplification of - RNA (genomic) concatamer; ribozyme cleavage....

Answer 5

The therapeutic option for patients with chronic hepatitis delta virus infection (CHD) is limited to interferon alpha with rare curative outcome. Myrcludex B is a first-in-class entry inhibitor inactivating the hepatitis B virus (HBV) and hepatitis D virus (HDV) receptor sodium taurocholate co-transporting polypeptide. We report the interim results of a pilot trial on chronically infected HDV patients treated with myrcludex B, or pegylated interferon alpha (PegIFNα-2a) or their combination. METHODS: Twenty-four patients with CHD infection were equally randomized (1:1:1) to receive myrcludex B, or PegIFNα-2a or their combination. Patients were evaluated for virological and biochemical response and tolerability of the study drugs at weeks 12 and 24. CONCLUSIONS: Myrcludex B showed a strong effect on HDV RNA serum levels and induced ALT normalization under monotherapy. Synergistic antiviral effects on HDV RNA and HBV DNA in the Myr-IFN cohort indicated a benefit of the combination of entry inhibition with PegIFNα-2a to treat CHD patients. LAY SUMMARY: Myrcludex B is a new drug to treat hepatitis B and D infection. After 24weeks of treatment with myrcludex B and/or pegylated interferon α-2a, HDV RNA, a relevant marker for hepatitis D infection, decreased in all patients with chronic hepatitis B and D. Two of eight patients which received either myrcludex B or pegylated interferon α-2a, became negative for HDV RNA, and five of seven patients who received both drugs at the same time became negative. The drug was well tolerated. NEW NAME: Hepcludex

Answer 6

Hepatitis D virus (HDV) doesn't encode envelope proteins for packaging of its ribonucleoprotein (RNP) and typically relies on the surface glycoproteins (GPs) from hepatitis B virus (HBV) for virion assembly, envelopment and cellular transmission. HDV RNA genome can efficiently replicate in different tissues and species, raising the possibility that it evolved, and/or is still able to transmit, independently of HBV. Here we show that alternative, HBV-unrelated viruses can act as helper viruses for HDV. In vitro, envelope GPs from several virus genera, including vesiculovirus, flavivirus and hepacivirus, can package HDV RNPs, allowing efficient egress of HDV particles in the extracellular milieu of co-infected cells and subsequent entry into cells expressing the relevant receptors. Furthermore, HCV can propagate HDV infection in the liver of co- infected humanized mice for several months. Further work is necessary to evaluate whether HDV is currently transmitted by HBV-unrelated viruses in humans.

Answer 7

Hepatitis B Virus (HBV) infection can be prevented by vaccination. Vaccines containing the small (S) envelope protein are currently used in universal vaccination programs and achieve protective immune response in more than 90% of recipients. However, new vaccination strategies are necessary for successful immunization of the remaining non- or low-responders... Here we describe the transient expression of the S/preS121-47 antigen in an edible plant, Lactuca sativa, for potential development of an oral HBV vaccine. Our study shows that oral administration of adjuvant-free Lactuca sativa expressing the S/preS121-47 antigen, three times, at 1 μg/dose, was sufficient to trigger a humoral immune response in mice. Importantly, the elicited antibodies were able to neutralize HBV infection in an NTCP- expressing infection system (HepG2-NTCP cell line) more efficiently than those induced by mice fed on Lactuca sativa expressing the S protein. These results support the S/preS121-47 antigen as a promising candidate for future development as an edible HBV vaccine.

Answer 8

- core protein self-assembles into particles – easy to purify, highly antigenic - since long time trials to present foreign antigens on core to immune system - problem: many large inserts in core protein interfere with particle assembly - solution: cleavage of core with large insert (by addition of protease) into N-term. and C-term. half followed by assembly into icosahedral hepatitis B virus capsids - large inserts do no longer interfer with particle formation - programmable viral nanoparticles