NS5A - From Obscurity to New Target for HCV Therapy

Schmitz, Uli; Tan, Seng‐Lai

doi:10.2174/157489108784746597

Cited by 84 publications

(60 citation statements)

References 44 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Indeed, genetic mutation analysis confirmed that the NS5A AH is necessary for HCV replication [63]. Together, the necessity of NS5A AH for membrane association and viral genome replication suggests that targeting membrane association of NS5A would represent a prime therapeutic target to inhibit replication [65]. Compared to other HCV targets that are prone to viral resistance, the NS5A AH possesses significant sequence homology across all HCV isolates identified to date [64].…”

Section: Challenges Of Hepatitis C Drug Developmentmentioning

confidence: 93%

Model Membrane Platforms for Biomedicine: Case Study on Antiviral Drug Development

Jackman

Cho

2012

Biointerphases

View full text Add to dashboard Cite

As one of the most important interfaces in cellular systems, biological membranes have essential functions in many activities such as cellular protection and signaling. Beyond their direct functions, they also serve as scaffolds to support the association of proteins involved in structural support, adhesion, and transport. Unfortunately, biological processes sometimes malfunction and require therapeutic intervention. For those processes which occur within or upon membranes, it is oftentimes difficult to study the mechanism in a biologically relevant, membranous environment. Therefore, the identification of direct therapeutic targets is challenging. In order to overcome this barrier, engineering strategies offer a new approach to interrogate biological activities at membrane interfaces by analyzing them through the principles of the interfacial sciences. Since membranes are complex biological interfaces, the development of simplified model systems which mimic important properties of membranes can enable fundamental characterization of interaction parameters for such processes. We have selected the hepatitis C virus (HCV) as a model viral pathogen to demonstrate how model membrane platforms can aid antiviral drug discovery and development. Responsible for generating the genomic diversity that makes treating HCV infection so difficult, viral replication represents an ideal step in the virus life cycle for therapeutic intervention. To target HCV genome replication, the interaction of viral proteins with model membrane platforms has served as a useful strategy for target identification and characterization. In this review article, we demonstrate how engineering approaches have led to the discovery of a new functional activity encoded within the HCV nonstructural 5A protein. Specifically, its N-terminal amphipathic, α-helix (AH) can rupture lipid vesicles in a size-dependent manner. While this activity has a number of exciting biotechnology and biomedical applications, arguably the most promising one is in antiviral medicine. Based on the similarities between lipid vesicles and the lipid envelopes of virus particles, experimental findings from model membrane platforms led to the prediction that a range of medically important viruses might be susceptible to rupturing treatment with synthetic AH peptide. This hypothesis was tested and validated by molecular virology studies. Broad-spectrum antiviral activity of the AH peptide has been identified against HCV, HIV, herpes simplex virus, and dengue virus, and many more deadly pathogens. As a result, the AH peptide is the first in class of broad-spectrum, lipid envelope-rupturing antiviral agents, and has entered the drug pipeline. In summary, engineering strategies break down complex biological systems into simplified biomimetic models that recapitulate the most important parameters. This approach is particularly advantageous for membrane-associated biological processes because model membrane platforms provide more direct characterization of target interactions than is possible with other methods. Consequently, model membrane platforms hold great promise for solving important biomedical problems and speeding up the translation of biological knowledge into clinical applications.

show abstract

Section: Challenges Of Hepatitis C Drug Developmentmentioning

confidence: 93%

Model Membrane Platforms for Biomedicine: Case Study on Antiviral Drug Development

Jackman

Cho

2012

Biointerphases

View full text Add to dashboard Cite

show abstract

“…Recent studies have demonstrated that domain III (residues 356 to 447) plays a critical role in infectious virion assembly but not in RNA replication (1,34) and that the former role is modulated by phosphorylation within the domain (33). High-throughput screening of small-molecule inhibitors using HCV replicon cell systems has identified NS5A as a promising therapeutic target (31).…”

mentioning

confidence: 99%

Crystal Structure of a Novel Dimeric Form of NS5A Domain I Protein from Hepatitis C Virus

Love

Brodsky

Hickey

et al. 2009

J Virol

222

288

View full text Add to dashboard Cite

Hepatitis C virus (HCV) is a member of the Flaviviridae family of enveloped, positive-strand RNA viruses (23). It is responsible for persistent infections in humans, with associated risk of chronic liver diseases, including cirrhosis and hepatocellular carcinoma. Nearly 3% of the global population is chronically infected with HCV, and there are no clinically proven vaccines. Antiviral therapeutic agents are at an early stage of clinical evaluation, and standard treatments (interferon and ribavirin combinations) are associated with suboptimal response rates and/or high incidence of side effects. Complicating the discovery of new therapies is the highly complex and incompletely understood nature of the viral life cycle. The HCV genome consists of a single strand of RNA of about 9,600 nucleotides encoding a polypeptide precursor of about 3,000 amino acids (26). Co-and posttranslational proteolytic cleavage of this precursor by cellular and viral enzymes yields structural proteins involved in viral assembly, along with nonstructural (NS) proteins NS2, NS3, NS4A, NS4B, NS5A, and NS5B, which are required for membrane-associated RNA replication (14).Nonstructural protein NS5A is a critical component of HCV replication and is involved in several cellular processes, such as interferon resistance (3, 13) and apoptotic regulation (9). It is a phosphoprotein of 447 residues with three domains (35), and while no clear enzymatic functions have been assigned, it appears to function through interactions with other HCV proteins and host cell factors (17). Domain I (residues 1 to 213) contains a zinc-binding motif (35) and an amphipathic N-terminal helix which promotes membrane association (4, 12, 30), possibly through specific interaction of the helix with target membrane proteins (8). Domain II (residues 250 to 342) has regulatory functions, such as interactions with protein kinase PKR and PI3K (13), as well as NS5B (32); contains the interferon sensitivity-determining region (13); and appears to lack major elements of secondary structure (22). Recent studies have demonstrated that domain III (residues 356 to 447) plays a critical role in infectious virion assembly but not in RNA replication (1,34) and that the former role is modulated by phosphorylation within the domain (33). High-throughput screening of small-molecule inhibitors using HCV replicon cell systems has identified NS5A as a promising therapeutic target (31).A crystal structure of domain I lacking the amphipathic helix and spanning residues 25 to 215 showed two subdomains and a homodimeric association and was interpreted as having a potential role in RNA binding (36). Although specific binding to domain I was not described, RNA binding to full-length NS5A has been reported, using, for example, the 3Ј nontranslated region of HCV (15). Efforts in our laboratory to study the structure of NS5A have yielded an alternative arrangement of the domain I homodimer (residues 33 to 202) that differs substantially from that previously described. The observation that the NS5A do...

show abstract

“…5 Although symmetry is not essential for inhibition of HCV replication in genotype 1b replicon, as reported elsewhere, genotype 1a inhibition is most optimally achieved in symmetrically disposed compounds. 32 In summary, we describe here fundamental aspects associated with the HCV inhibitory activity of a series of C-5-phenylated iminothiazolidinones discovered in a high throughput replicon screen, compounds that target the NS5A protein.…”

mentioning

confidence: 99%

Inhibitors of HCV NS5A: From Iminothiazolidinones to Symmetrical Stilbenes

Romine

Laurent

Leet

et al. 2011

ACS Med. Chem. Lett.

View full text Add to dashboard Cite

ABSTRACT:The iminothiazolidinone BMS-858 (2) was identified as a specific inhibitor of HCV repli cation in a genotype 1b replicon assay via a high-throughput screening campaign. A more potent analogue, BMS-824 (18), was used in resistance mapping studies, which revealed that inhibitory activity was related to disrupting the function of the HCV nonstructural protein 5A. Despite the development of coherent and interpretable SAR, it was subsequently discovered that in DMSO 18 underwent an oxidation and structural rearrangement to afford the thiohydantoin 47, a compound with reduced HCV inhibitory activity. However, HPLC bioassay fractionation studies performed after incubation of 18 in assay media led to the identification of fractions containing a dimeric species 48 that exhibited potent antiviral activity. Excision of the key elements hypothesized to be responsible for antiviral activity based on SAR observations reduced 48 to a simplified, symmetrical, pharmacophore realized most effectively with the stilbene 55, a compound that demonstrated potent inhibition of HCV in a genotype 1b replicon with an EC 50 = 86 pM.

show abstract

NS5A - From Obscurity to New Target for HCV Therapy

Cited by 84 publications

References 44 publications

Model Membrane Platforms for Biomedicine: Case Study on Antiviral Drug Development

Model Membrane Platforms for Biomedicine: Case Study on Antiviral Drug Development

Crystal Structure of a Novel Dimeric Form of NS5A Domain I Protein from Hepatitis C Virus

Inhibitors of HCV NS5A: From Iminothiazolidinones to Symmetrical Stilbenes

Contact Info

Product

Resources

About