In research recently published in the journal of Science Advances, an interdisciplinary team of researchers from the University of Chicago assessed the potential role of cannabidiol (CBD), a natural product extracted from the cannabis plant, to prevent and inhibit severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections in human cells and mice.
Study: Cannabidiol inhibits SARS-CoV-2 replication through induction of the host ER stress and innate immune responses. Image Credit: Dmytro Tyshchenko / Shutterstock
The ongoing coronavirus disease 2019 (COVID-19) pandemic has caused widespread morbidity and mortality across the globe ever since it started in late 2019. Despite the availability of a selection of COVID-19 vaccines, SARS-CoV-2 and its newly emerging variants are spreading worldwide, stressing the need for alternative solutions such as effective treatments. Unfortunately, despite a collective global effort, only limited therapies have been identified so far that can block SARS-CoV-2 replication and viral production.
About the study
In the current study, researchers used human A549 lung carcinoma, human lung Calu3 cells, and Vero E6 monkey kidney epithelial cells for assessing the effect of CBD on SARS-CoV-2 replication. CBD and related compounds were isolated and identified by 1D- 1H nuclear magnetic resonance (NMR) analysis and compared with the 1H iterative full spin analysis (HiFSA) profile of CBD.
For assessing the influence of CBD on the endoplasmic reticulum (ER) stress response, gene set enrichment analysis (GSEA) was used for quantifying X-box binding protein 1 (XBP1) splicing using ribonucleic acid (RNA) sequencing gene expression data, followed by confirmation using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Inositol-requiring enzyme 1 α (IRE1α) knockout cells and anti-interferon blocking antibodies were used for assessing the effect of CBD on IRE1α and interferons.
The researchers observed that pre-treatment of A549-ACE-2 human lung carcinoma cell lines with 0-10 µM CBD (97% purity with congeneric cannabinoids <1%) before infection with SARS-CoV-2 or α, β or γ variants potently inhibited SARS-CoV-2 viral replication with an EC50 of 1µM. In addition, CBD also inhibited SARS-CoV-2 replication in human Calu3 lung and Vero E6 monkey kidney epithelial cells.
According to researchers, except for CBD, none of the closely related CBD congeners-such as tetrahydrocannabinol (THC), cannabidivarin (CBDV), cannabidiolic acid (CBDA), cannabichromene (CBC), or cannabigerol inhibited SARS-CoV-2 infection. Perhaps combining CBD with THC (1:1) significantly suppressed CBD efficacy consistent with competitive inhibition. A metabolite of CBD- 7-hydroxy-cannabidiol (7-OH-CBD) effectively inhibited SARS-CoV-2 replication in A549 angiotensin-converting enzyme 2 (ACE2) cells at a non-toxic level.
No effect of CBD was observed on ACE2 receptor expression on three cells lines used in the study. CBD showed no effect on viral entry but was 95-99% effective at inhibiting SARS-CoV-2 spike protein expression in host cells post entry. CBD did not affect viral protein processing by viral main protease (Mpro) or papain-like protease (PLpro), showing that CBD targets host cell processes.
The study revealed remarkable suppression of induction of SARS-CoV-2- genes for a spike, envelope, and nucleocapsid protein by 99% by CBD in contrast to 60% with CBDV. CBDV caused fewer transcriptomic changes compared to CBD and was mostly ineffective in reversing transcriptional changes induced by SARS-CoV-2. CBDV induced autophagy and lipid metabolism and suppressed processes such as protein translation and DNA replication.
The team found that CBD caused gene expression of three transmembrane proteins of unfolded protein response (UPR) pathway – IRE1α, protein kinase R-like endoplasmic reticulum kinase (PERK), and activating transcription factor 6 (ATF6). CBD strongly activated the IRE1α pathway in the presence or absence of the virus, but not by SARS-CoV-2 alone. In contrast, PERK was activated by both SARS-CoV-2 and CBD.
CBD, in the presence of SARS-CoV-2, more strongly induced genes involved in the interferon signaling pathway- ISG15, IFIT1, IFIT3, SOCS1, and OAS1 compared to CBD alone. CBD effectively reversed viral induction of cytokines leading to cytokine storm at later stages of infection. On the other hand, the inactive homolog CBDV does not significantly prevent cytokine induction. The exposure of ACE2-A549 cells to a mixture of antibodies against Type I (α,β,o) and Type II (γ) interferons before 2.5 mM CBD treatment and infection with SARS-CoV-2 showed that anti-interferon antibodies reduced CBD’s anti-viral effects and rescue SARS-CoV-2 infection partially.
CBD administration at lower and higher doses decreased viral titers in lungs (4.8 and 40 folds) and nasal turbinate (3.7 and 4.8 fold) in K18-hACE2 female mice subjected to intranasal challenge with SARS-CoV-2 (2×104 plaque-forming units [PFU]).
From an analysis of a patient group with records of consumption of CBD (100 mg/ml) from the National COVID Cohort Collaborative (N3C), researchers observed that CBD consumption was significantly associated with fewer SARS-CoV-2 positive test results.
The study results showed that among all cannabinoids congeners, only CBD and its metabolite 7-OH-CBD, potently block SARS-CoV-2 replication in lung epithelial cells. In addition, CBD exerted its effect post-viral entry, and inhibited viral genome expression. CBD also inhibited SARS-CoV-2 replication by the activation of host IRE1α, RNase ER stress response, and interferon pathways.
Pre-clinical studies have established the efficacy of CBD against SARS-CoV-2 during the early stages of infection. This study demonstrated the role of CBD as a potential agent for combating SARS-CoV-2 infection during the early stages. However, additional studies and clinical trials will be needed for determining precise dosing, safety profile, and potential side effects of CBD as a treatment agent against SARS-CoV-2 infection in humans.
The researchers cautioned that the anti-viral effects of CBD demonstrated in this study apply only for high purity, specially formulated doses taken in specific situations. The study’s findings do not suggest that the consumption of commercially available products of CBD with varying potency and quality can prevent or treat COVID-19.
- Cannabidiol inhibits SARS-CoV-2 replication through induction of the host ER stress and innate immune responses. Long Chi Nguyen, Dongbo Yang, Vlad Nicolaescu, Thomas J. Best, Haley Gula, Divyasha Saxena, Jon D. Gabbard, Shao-Nong Chen, Takashi Ohtsuki, John Brent Friesen, Nir Drayman, Adil Mohamed, Christopher Dann, Diane Silva, Lydia Robinson-Mailman, Andrea Valdespino, Letícia Stock, Eva Suárez, Krysten A. Jones, Saara-Anne Azizi, Jennifer K. Demarco, William E. Severson, Charles D. Anderson, James Michael Millis, Bryan C. Dickinson, Savaş Tay, Scott A. Oakes, Guido F. Pauli, Kenneth E. Palmer, David O. Meltzer, Glenn Randall, Marsha Rich Rosner. Science Advances. 2022, DOI: 10.1126/sciadv.abi6110, https://www.science.org/doi/10.1126/sciadv.abi6110