The Antiviral Alkaloid Berberine Reduces Chikungunya Virus-Induced Mitogen-Activated Protein Kinase Signaling

Abstract

Chikungunya virus (CHIKV) has infected millions of people in the tropical and subtropical regions since its reemergence in the last decade. We recently identified the nontoxic plant alkaloid berberine as an antiviral substance against CHIKV in a highthroughput screen. Here, we show that berberine is effective in multiple cell types against a variety of CHIKV strains, also at a high multiplicity of infection, consolidating the potential of berberine as an antiviral drug. We excluded any effect of this compound on virus entry or on the activity of the viral replicase. A human phosphokinase array revealed that CHIKV infection specifically activated the major mitogen-activated protein kinase (MAPK) signaling pathways extracellular signal-related kinase (ERK), p38 and c-Jun NH 2 -terminal kinase (JNK). Upon treatment with berberine, this virus-induced MAPK activation was markedly reduced. Subsequent analyses with specific inhibitors of these kinases indicated that the ERK and JNK signaling cascades are important for the generation of progeny virions. In contrast to specific MAPK inhibitors, berberine lowered virus-induced activation of all major MAPK pathways and resulted in a stronger reduction in viral titers. Further, we assessed the in vivo efficacy of berberine in a mouse model and measured a significant reduction of CHIKV-induced inflammatory disease. In summary, we demonstrate the efficacy of berberine as a drug against CHIKV and highlight the importance of the MAPK signaling pathways in the alphavirus infectious cycle. IMPORTANCEChikungunya virus (CHIKV) is a mosquito-borne virus that causes severe and persistent muscle and joint pain and has recently spread to the Americas. No licensed drug exists to counter this virus. In this study, we report that the alkaloid berberine is antiviral against different CHIKV strains and in multiple human cell lines. We demonstrate that berberine collectively reduced the virus-induced activation of cellular mitogen-activated protein kinase signaling. The relevance of these signaling cascades in the viral life cycle was emphasized by specific inhibitors of these kinase pathways, which decreased the production of progeny virions. Berberine significantly reduced CHIKV-induced inflammatory disease in a mouse model, demonstrating efficacy of the drug in vivo. Overall, this work makes a strong case for pursuing berberine as a potential anti-CHIKV therapeutic compound and for exploring the MAPK signaling pathways as antiviral targets against alphavirus infections. C hikungunya virus (CHIKV) is the causative agent of chikungunya fever, a disease spread by Aedes mosquitoes and characterized by a sudden onset of febrile illness, nausea, headache, and most importantly, severe and persistent musculoskeletal pain (1). It reemerged in tropical Asia and Africa 1 decade ago and since 2013 has infected more than 1.5 million people in the Americas (2). So far, no licensed vaccine or antiviral treatment exists to counter this disease. Over the years, many research groups have focused on...

P38 and JNK Mitogen-Activated Protein Kinases Interact With Chikungunya Virus Non-structural Protein-2 and Regulate TNF Induction During Viral Infection in Macrophages

“…The MAPKs have been shown to be activated by phosphorylation in specific positions (Ser/Tyr/Thr) by several viral infections, such as coronavirus type 2, Hepatitis C virus, Rhinovirus and Epstein-Barr virus (54)(55)(56)(57)(58). CHIKV is also known to induce MAPKs during infection in various nonimmune cells and treatment of an alkaloid berberine, reduces viral infection and joint swelling in mice (59,60).…”

Design and Validation of Novel Chikungunya Virus Protease Inhibitors

“…Targeting of several metabolic pathways has revealed anti-CHIKV effects of several licensed drugs (3). Compounds most likely targeting virus entry or host cell-specific components required for virus infection have been described (4)(5)(6)(7)(8). Several nucleosides or nucleotides, acting as pseudosubstrates for CHIKV RNA-dependent RNA polymerase and/or using another, more general mechanism of action, have shown anti-CHIKV activity (9).…”

A Biologically-Based Computational Approach to Drug Repurposing for Anthrax Infection

“…Notably, literature reports supported their activities for protecting cells from LT or ET toxicity [32,35,36,41,42,43,44,45,46,47,48,49,50,51,52,53]. On the other hand, the bottom 10 drugs listed in Table 4, except monastrol [54], ebselen [55], and genistein [56], had activities similar to those of anthrax toxins, including reduction of MAPK activities or increase of intracellular level of cAMP [32,57,58,59,60,61,62,63]. Monastrol [54] arrested cells in mitosis, unlikely providing protection if cells were under attack by anthrax toxins.…”

“…As highlighted in Table 4, berberine [58] and apigenin [62] inhibited MAPK, thereby enhancing LT toxicity. So could withaferin a, arecoline, and beclomethasone.…”