On a path toward a broad-spectrum anti-viral: inhibition of HIV-1 and coronavirus replication by SR kinase inhibitor harmine.

IMPORTANCE: This study highlights the crucial role RNA processing plays in regulating viral gene expression and replication. By targeting SR kinases, we identified harmine as a potent inhibitor of HIV-1 as well as coronavirus (HCoV-229E and multiple SARS-CoV-2 variants) replication. Harmine inhibits HIV-1 protein expression and reduces accumulation of HIV-1 RNAs in both cell lines and primary CD4+ T cells. Harmine also suppresses coronavirus replication post-viral entry by preferentially reducing coronavirus sub-genomic RNA accumulation. By focusing on host factors rather than viral targets, our study offers a novel approach to combating viral infections that is effective against a range of unrelated viruses. Moreover, at doses required to inhibit virus replication, harmine had limited toxicity and minimal effect on the host transcriptome. These findings support the viability of targeting host cellular processes as a means of developing broad-spectrum anti-virals.

Eukaryotic translation initiation factor 4AII contributes to microRNA-122 regulation of hepatitis C virus replication.

Hepatitis C virus (HCV) is a positive sense RNA virus that persistently infects human liver, leading to cirrhosis and hepatocellular carcinoma. HCV replication requires the liver-specific microRNA-122 (miR-122). In contrast to canonical miRNA-mediated repression via 3'UTR sites, miR-122 positively regulates HCV replication by a direct interaction with the 5' untranslated region (UTR) of the viral RNA. The protein factor requirements for this unusual miRNA regulation remain poorly understood. Here, we identify eIF4AII, previously implicated in miRNA-mediated repression via 3'UTR sites, as a host factor that is important for HCV replication. We demonstrate that eIF4AII interacts with HCV RNA and that this interaction is miR-122-dependent. We show that effective miR-122 binding to, and regulation of, HCV RNA are reduced following eIF4AII depletion. We find that the previously identified HCV co-factor CNOT1, which has also been implicated in miRNA-mediated repression via 3'UTR sites, contributes to regulation of HCV by eIF4AII. Finally, we show that eIF4AI knockdown alleviates the inhibition of HCV replication mediated by depletion of either eIF4AII or CNOT1. Our results suggest a competition effect between the eIF4A proteins to influence HCV replication by modulation of miR-122 function.

Broad spectrum post-entry inhibitors of coronavirus replication: Cardiotonic steroids and monensin.

A small molecule screen identified several cardiotonic steroids (digitoxin and ouabain) and the ionophore monensin as potent inhibitors of HCoV-229E, HCoV-OC43, and SARS-CoV-2 replication with EC50s in the low nM range. Subsequent tests confirmed antiviral activity in primary cell models including human nasal epithelial cells and lung organoids. Addition of digitoxin, ouabain, or monensin strongly reduced viral gene expression as measured by both viral protein and RNA accumulation. Furthermore, the compounds acted post virus entry. While the antiviral activity of digitoxin was dependent upon activation of the MEK and JNK signaling pathways but not signaling through GPCRs, the antiviral effect of monensin was reversed upon inhibition of several signaling pathways. Together, the data demonstrates the potent anti-coronavirus properties of two classes of FDA approved drugs that function by altering the properties of the infected cell, rendering it unable to support virus replication.

Hepatitis B virus genotypes, subgenotypes, precore, and basal core promoter mutations in the two largest provinces of Pakistan.

BACKGROUND AND AIM: Hepatitis B virus (HBV) genotyping has been done in most countries, but unfortunately, in Pakistan, HBV genotypic distribution is still unclear. The aim of the present study was to determine the prevalent genotype and subgenotype in the two most populated provinces in Pakistan: Punjab and Sind. METHODS: In total, 236 HBV DNA-positive samples were selected for genotyping by polymerase chain reaction-restriction fragment length polymorphism (RFLP). The RFLP results were further confirmed with whole genome and partial genome sequencing. RESULTS: Genotype D was detected as the most prevalent (93.22%) genotype in all eight cities of both provinces; genotype C was present in 5.93% and genotype A was present in 0.85% of the samples. The D1 subtype was present in 84%, and D2 was present in 8% of 25 whole genome-sequenced samples. The C2 subtype was detected in 58.33% of S gene-sequenced samples, while D1 was detected in the remaining 41.67% of 24 samples sequenced for the S gene. Subtype D1 is the most dominant in D, while C2 is dominant in genotype C. Eight- and 15-bp deletion mutations were also detected in genotype D samples. Other precore and basal core promoter (BCP) mutations included T1915 (100%), A1679 (86.96%), T1762 (39.13%), and A1764 (30.43%), which were also detected in the genotype D samples. CONCLUSION: Genotype D subtype D1 is the most prevalent HBV strain in Pakistan with 8-bp deletion mutants the most common in HBV carriers.

Serum HBsAg changes in HBeAg positive chronic hepatitis B patients with continuous viral load reductions during treatment with adefovir or peg-interferon-alpha-2a.

Hepatitis B surface antigen (HBsAg) loss under antiviral therapy is rare in chronic hepatitis B patients and the dynamics of serum HBsAg in these patients are not available. The changes in serum HBsAg following treatment with adefovir (n=31) or peg-interferon-alpha-2a (n=23) were studied in hepatitis B e-antigen (HBeAg) positive chronic hepatitis B patients. Abbott Architect HBsAg assay was used to quantify serum HBsAg. HBsAg levels were significantly decreased during the first 12 weeks of treatment with median change of -397.0 IU/ml and -555.4 IU/ml, respectively for adefovir and peg-interferon-alpha-2a (p=0.005 and 0.001, respectively). Beyond 12 weeks, no further significant HBsAg reductions were found even in patients with sustained viral replication inhibition in either group. Three distinct patterns of HBsAg changes were observed in most patients in both treatment groups: biphasic pattern (rapid HBsAg reduction from baseline to week 12); assurgent pattern (higher HBsAg level at week 12 than at baseline); and wavy pattern (HBsAg reduction from baseline to week 12, followed by relapse at week 24 or week 28). These results might offer insights into the possible mechanism(s) underlying the unusual occurrences of HBsAg loss under antiviral therapy.