In a recent study published in the journal PLOS ONE, Researchers from the United States of America investigated the antiviral effects of valproic acid (VPA), alone and in combination with docosahexaenoic acid (DHA), against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. They found that VPA, especially when combined with DHA, reduced the rates and severity of SARS-CoV-2 infection and potentially activated intracellular antiviral mechanisms by modifying gene expression.
Record
Originally used as an inert excipient, VPA was discovered in the 1960s to have anticonvulsant properties. It is now used to treat seizure disorders, bipolar disorder and migraines. It has a known toxicity profile, including a threefold increased risk of congenital defects, possibly due to histone deacetylase (HDAC) inhibition, which makes it contraindicated in pregnant women. The HDAC inhibitory activity of VPA is also being investigated for the treatment of cancer and the treatment of human immunodeficiency virus (HIV). In addition, VPA has antiviral activity against various viruses, including DNA (short for deoxyribonucleic acid), RNA (short for ribonucleic acid), and enveloped viruses. Efforts to repurpose Food and Drug Administration (FDA)-approved drugs for coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, have identified VPA as a potential candidate due to its interaction with the viral nsp5 protease. However, precedent in vitro Viral replication assays showed no antiviral activity for VPA at the doses tested.
In the present study, the researchers conducted in vitro trial and epidemiological analysis to investigate the antiviral activity of VPA alone and in combination with DHA. They identified pitfalls in screening methods and suggested a potentially cost-effective strategy against some coronaviruses.
About the study
In the present study, the cytotoxicity of VPA was assessed using the XTT (short for methoxynitrosulfophenyl-tetrazolium carboxanilide) assay. The half-maximal inhibitory concentration of VPA (IC50) was determined using antibody-based, luciferase, and dot-blotting assays in Vero and MRC-5 cells infected with SARS-CoV-2 or human coronavirus 229E (HCoV-229E). HDAC activity was measured using a fluorometric assay kit. The impact of polyunsaturated fatty acids (PUFAs) such as DHA, eicosapentaenoic acid (EPA), linoleic acid (LA) and alpha-linolenic acid (αLA) on HCoV-229E replication was evaluated. RNA was extracted, sequenced and analyzed for differential gene expression changes. Protein expression was confirmed by Western blotting. The effects of VPA and DHA on HCoV-229E replication were studied using RNA sequencing (RNAseq). MRC-5 cells were pretreated with DHA, VPA, or both, then infected with SARS-CoV-2 to assess replication inhibition, with results analyzed by real-time reverse transcription polymerase chain reaction (RT -PCR) and RNAseq.
Further, epidemiological data from the Optum dataset were analyzed to study the outcomes of COVID-19 associated with VPA use, and serum VPA levels were obtained from LabCorp for analysis.
Results and discussion
In vitro, the IC50 of VPA for HDAC2 inhibition was found to be 2.5 mM, higher than therapeutic levels. Although initial virus inhibition assays did not show effective inhibition of viral replication at doses tested, preincubation with VPA was observed to reduce the IC50 by more than threefold, approaching therapeutic levels. The effect of VPA on gene expression suggests potential antiviral activity, although direct inhibition of SARS-CoV-2 was not demonstrated at therapeutic doses.
In addition, LA and DHA were found to significantly inhibit virus replication when tested on HCoV-229E. DHA was found to enhance the antiviral activity of VPA, lowering the IC50 for inhibition of viral replication. Combinations of VPA and DHA showed significant antiviral synergy. Testing on SARS-CoV-2 revealed that VPA, DHA and their combination inhibited virus replication, although preincubation did not improve efficacy.
In the epidemiological arm of the study, Optum data analysis revealed a 14.9% positivity rate for COVID-19 among three million patients screened between 2020 and 2021. Higher rates of black and Hispanic patients were found in the COVID-19 positive group. Less than 25% of patients had serum VPA levels above the cutoff for therapeutic levels. While univariate analysis showed no protective effect of VPA against COVID-19, multivariate models, adjusting for comorbidities, showed that VPA use was associated with 12–15% fewer emergency room (ER) visits, 17–45% fewer admissions in the hospital, 33–39% fewer cases requiring mechanical ventilation and 14–16% fewer intensive care unit (ICU) admissions related to COVID-19.
conclusion
In conclusion, the epidemiologic findings of the study suggest that VPA use is associated with lower COVID-19 test positivity, ER visits, hospitalizations, ICU admissions, and mechanical ventilation needs, with effects persisting across alpha and delta variants. Although the study does not confirm causality, the combination of VPA with DHA shows significant antiviral activity against coronaviruses, possibly due to the activation of antiviral genes. In the future, this combination may remain effective against new variants and reduce the severity and spread of the disease. Further clinical research is needed to confirm these findings and investigate PUFA and HDAC inhibitor combination therapies for COVID-19.
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