Discovery and Characterization of a Novel HCV Inhibitor Targeting the Late Stage of HCV Life Cycle

Abstract

Background Currently approved anti-HCV drugs, the direct-acting antivirals (DAAs), are highly effective and target the viral RNA replication stage of the HCV life cycle. Due to high mutation rate of HCV, drug resistant variants can arise during DAA monotherapy. Thus, a combination of DAAs is necessary to achieve a high response rate. Novel HCV inhibitors targeting the HCV late stage such as assembly and release may further improve combination therapy with the DAAs. Here we characterize one late stage-targeting candidate compound, 6-(4-chloro-3-methylphenoxy)-pyridin-3-amine (MLS000833705). Methods: We treated HCV-infected cells with MLS000833705 and other HCV inhibitors and examined HCV RNA and infectious titres. We evaluated the colocalization of HCV core and lipid droplets by confocal microscopy. We performed HCV core-proteinase K digestion assay and several lipid assays to study the mechanism of MLS000833705. Results We showed that MLS000833705 decreased extracellular HCV RNA levels more than intracellular HCV RNA levels in HCV infectious cell culture. Similarly, MLS000833705 reduced infectious HCV titres substantially more in the culture supernatant than intracellularly. Confocal microscopy showed that MLS000833705 did not affect the colocalization of HCV core protein with cellular lipid droplets where HCV assembles. HCV core-proteinase K digestion assay showed that MLS000833705 inhibited the envelopment of HCV capsid. Conclusions Our study demonstrates that MLS000833705 is a late-stage HCV inhibitor targeting HCV morphogenesis and maturation. Therefore, MLS000833705 can be used as a molecular probe to study HCV maturation and secretion and possibly guide development of a new class of HCV antivirals.

Targeting the Virus Capsid as a Tool to Fight RNA Viruses

“…High throughput screening of about 350,000 compounds in a human hepatomaderived cell line, Huh7.5.1, a suitable infection system for hepatitis C virus, yielded 158 molecules with anti-HCV activity [160]. Later focus on their mechanism of action resulted in the identification of 6-(4-chloro-2-methylphenoxy)pyridin-3-amine (Table 3), which inhibited the viral morphogenesis [161]. Indirect inhibition of assembly was achieved by inhibition of AAK1 and GAK, serine/threonine kinases stimulating binding of clathrin adaptor protein complex 2 to cargo, and are essential for HCV assembly [162].…”

Herbal Medicines for Hepatitis C Virus Infection: The Exploratory Journey from Bench to Bedside Still Has a Long Way to Go

“…HCV life cycle includes the entry of HCV particle into hepatocytes, translation and replication of HCV RNA, and assembly, budding and secretion of HCV. 13, 14 The HCV genome is approximately 9.6 kb of uncapped RNA. 15 The HCV RNA encodes a polyprotein, which is processed by host and viral proteases into at least 10 products.…”

Evaluation of interactions between the hepatitis C virus NS3/4A and sulfonamidobenzamide based molecules using molecular docking, molecular dynamics simulations and binding free energy calculations

“…DAAs are combinations of medications that are currently being used to effectively treat hepatitis C [29]. Although effective, the therapy has its own shortcomings; (i) The cost of DAAs is quite high, adding to the economic burden on patients [30], and there is at least one report of perceived stress in HCV infected patient under DAA therapy [31], (ii) there have been reports of associated side effects [32,33], (iii) not all HCV genotypic variants are equally responsive to the current therapy [34], (iv) resistance-related variants could arise during DAA monotherapy as a result of the high HCV mutation rate [35,36], (iv) failed therapies have been reported for some di cult-to-treat patients [37], and (v) recurrences of hepatocellular carcinoma have been reported after HCV therapy with DAAs [4].…”