Deciphering the antiviral potential of tomatidine towards mosquito-borne viral infections

Mosquito-borne viruses have rapidly spread around the world, causing millions of clinically apparent infections with various symptoms from rashes and high fever to joint pain that can persist for years. The most common mosquito-borne virus is dengue virus, which causes an estimated 390 million infections each year. To date, there is no specific antiviral compound available to treat these viral diseases. Hence, there is a strong need for the identification and development of new treatment options towards these viruses. In this thesis, we investigated the antiviral potential of tomatidine, a natural compound found in the unripe green tomatoes, towards the mosquito-borne viruses dengue virus, zika virus, west nile virus and chikungunya virus as well as the underlying mode of action. Our work shows that tomatidine is able to reduce the infection of dengue and chikungunya virus in different human cell lines. Moreover, we found a significant but less potent antiviral activity of tomatidine towards zika virus but not towards west nile virus. Regarding the mode of action by which tomatidine exerts its antiviral activity, we found that tomatidine inhibits viral infection by interfering with the production of viral RNA and/or viral proteins which are required for the formation of new virus particles. The exact mechanism by which tomatidine exerts its antiviral activity still remains to be elucidated. Extensive pre-clinical and pharmacokinetic studies are still needed to further evaluate the potential of tomatidine as an antiviral compound in the future

Recent advances in antiviral drug development towards dengue virus

Despite the high disease burden of dengue virus, there is no approved antiviral treatment or broadly applicable vaccine to treat or prevent dengue virus infection. In the last decade, many antiviral compounds have been identified but only few have been further evaluated in pre-clinical or clinical trials. This review will give an overview of the direct-acting and host-directed antivirals identified to date. Furthermore, important parameters for further development that is, drug properties including efficacy, specificity and stability, pre-clinical animal testing, and combinational drug therapy will be discussed

Tomatidine, a natural steroidal alkaloid shows antiviral activity towards chikungunya virus in vitro

In recent decades, chikungunya virus (CHIKV) has re-emerged, leading to outbreaks of chikungunya fever in Africa, Asia and Central and South America. The disease is characterized by a rapid onset febrile illness with (poly)arthralgia, myalgia, rashes, headaches and nausea. In 30 to 40% of the cases, CHIKV infection causes persistent (poly)arthralgia, lasting for months or even years after initial infection. Despite the drastic re-emergence and clinical impact there is no vaccine nor antiviral compound available to prevent or control CHIKV infection. Here, we evaluated the antiviral potential of tomatidine towards CHIKV infection. We demonstrate that tomatidine potently inhibits virus particle production of multiple CHIKV strains. Time-of -addition experiments in Huh7 cells revealed that tomatidine acts at a post-entry step of the virus replication cycle. Furthermore, a marked decrease in the number of CHIKV-infected cells was seen, suggesting that tomatidine predominantly acts early in infection yet after virus attachment and cell entry. Antiviral activity was still detected at 24 hours post-infection, indicating that tomatidine controls multiple rounds of CHIKV replication. Solasodine and sarsasapogenin, two structural derivatives of tomatidine, also showed strong albeit less potent antiviral activity towards CHIKV. In conclusion, this study identifies tomatidine as a novel compound to combat CHIKV infection in vitro

Tomatidine reduces chikungunya virus progeny release by controlling viral protein expression

Author summaryChikungunya fever is a debilitating disease caused by the mosquito-borne Chikungunya virus. Over the past two decades the geographical spread of the virus and its mosquito vector has drastically increased thereby causing millions of infections. To date there is no antiviral drug and no vaccine available to treat/prevent Chikungunya virus infection. We recently showed that the natural steroidal alkaloid tomatidine has potent antiviral activity towards Chikungunya virus at submicromolar concentrations. In this study we dissected how tomatidine reduces the production of Chikungunya virus particles. We show that tomatidine lowers viral protein expression and we hypothesize that the effect of tomatidine on viral protein translation hampers the production of progeny viral RNA copies / number of infected cells thereby leading to a reduced production of secreted virus particles. Also, we show that Chikungunya virus does not readily become resistant to tomatidine. Collectively, we deciphered the mechanism by which tomatidine exerts antiviral activity to Chikungunya virus and our results strengthen the potential of tomatidine as an antiviral treatment strategy towards Chikungunya virus.Tomatidine, a natural steroidal alkaloid from unripe green tomatoes has been shown to exhibit many health benefits. We recently provided in vitro evidence that tomatidine reduces the infectivity of Dengue virus (DENV) and Chikungunya virus (CHIKV), two medically important arthropod-borne human infections for which no treatment options are available. We observed a potent antiviral effect with EC50 values of 0.82 mu M for DENV-2 and 1.3 mu M for CHIKV-LR. In this study, we investigated how tomatidine controls CHIKV infectivity. Using mass spectrometry, we identified that tomatidine induces the expression of p62, CD98, metallothionein and thioredoxin-related transmembrane protein 2 in Huh7 cells. The hits p62 and CD98 were validated, yet subsequent analysis revealed that they are not responsible for the observed antiviral effect. In parallel, we sought to identify at which step of the virus replication cycle tomatidine controls virus infectivity. A strong antiviral effect was seen when in vitro transcribed CHIKV RNA was transfected into Huh7 cells treated with tomatidine, thereby excluding a role for tomatidine during CHIKV cell entry. Subsequent determination of the number of intracellular viral RNA copies and viral protein expression levels during natural infection revealed that tomatidine reduces the RNA copy number and viral protein expression levels in infected cells. Once cells are infected, tomatidine is not able to interfere with active RNA replication yet it can reduce viral protein expression. Collectively, the results delineate that tomatidine controls viral protein expression to exert its antiviral activity. Lastly, sequential passaging of CHIKV in presence of tomatidine did not lead to viral resistance. Collectively, these results further emphasize the potential of tomatidine as an antiviral treatment towards CHIKV infection.Molecular basis of virus replication, viral pathogenesis and antiviral strategie

Dietary fiber pectin directly blocks toll-like receptor 2-1 and prevents doxorubicin-induced ileitis

Dietary carbohydrate fibers are known to prevent immunological diseases common in Western countries such as allergy and asthma but the underlying mechanisms are largely unknown. Until now beneficial effects of dietary fibers are mainly attributed to fermentation products of the fibers such as anti-inflammatory short-chain fatty acids (SCFAs). Here, we found and present a new mechanism by which dietary fibers can be anti-inflammatory: a commonly consumed fiber, pectin, blocks innate immune receptors. We show that pectin binds and inhibits, toll-like receptor 2 (TLR2) and specifically inhibits the proinflammatory TLR2-TLR1 pathway while the tolerogenic TLR2-TLR6 pathway remains unaltered. This effect is most pronounced with pectins having a low degree of methyl esterification (DM). Low-DM pectin interacts with TLR2 through electrostatic forces between non-esterified galacturonic acids on the pectin and positive charges on the TLR2 ectodomain, as confirmed by testing pectin binding on mutated TLR2. The anti-inflammatory effect of low-DM pectins was first studied in human dendritic cells and mouse macrophages in vitro and was subsequently tested in vivo in TLR2-dependent ileitis in a mouse model. In these mice, ileitis was prevented by pectin administration. Protective effects were shown to be TLR2-TLR1 dependent and independent of the SCFAs produced by the gut microbiota. These data suggest that low-DM pectins as a source of dietary fiber can reduce inflammation through direct interaction with TLR2-TLR1 receptors

Resveratrol and Pterostilbene Inhibit SARS-CoV-2 Replication in Air-Liquid Interface Cultured Human Primary Bronchial Epithelial Cells

The current COVID-19 pandemic is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and has an enormous impact on human health and economy. In search for therapeutic options, researchers have proposed resveratrol, a food supplement with known antiviral, anti-inflammatory, and antioxidant properties as an advantageous antiviral therapy for SARS-CoV-2 infection. Here, we provide evidence that both resveratrol and its metabolically more stable structural analog, pterostilbene, exhibit potent antiviral properties against SARS-CoV-2 in vitro. First, we show that resveratrol and pterostilbene antiviral activity in African green monkey kidney cells. Both compounds actively inhibit virus replication within infected cells as reduced virus progeny production was observed when the compound was added at post-inoculation conditions. Without replenishment of the compound, antiviral activity was observed up to roughly five rounds of replication, demonstrating the long-lasting effect of these compounds. Second, as the upper respiratory tract represents the initial site of SARS-CoV-2 replication, we also assessed antiviral activity in air–liquid interface (ALI) cultured human primary bronchial epithelial cells, isolated from healthy volunteers. Resveratrol and pterostilbene showed a strong antiviral effect in these cells up to 48 h post-infection. Collectively, our data indicate that resveratrol and pterostilbene are promising antiviral compounds to inhibit SARS-CoV-2 infection. Because these results represent laboratory findings in cells, we advocate evaluation of these compounds in clinical trials before statements are made whether these drugs are advantageous for COVID-19 treatment

Tomatidine and analogs thereof for use as antiviral agent

The invention relates to the fields of medicine and virology, more in particular to means and methods for treating a viral disease caused by flaviviruses or alphaviruses. Provided is tomatidine or an analog thereof for use in a method of treating a viral infection caused by a flavivirus or an alphavirus, such as dengue virus or Chikungunya virus. Also provided is a pharmaceutical composition comprising tomatidine or an analog thereof, and at least one further antiviral agent