CBD & CBG

CANNABIDIOL (CBD)

Cannabidiol Inhibits SARS-CoV-2 Replication and Promotes the Host Innate Immune Response. The rapid spread of COVID-19 underscores the need for new treatments. Here we report that cannabidiol (CBD), a compound produced by the cannabis plant, inhibits SARS-CoV-2 infection. CBD and its metabolite, 7-OH-CBD, but not congeneric cannabinoids, potently block SARS-CoV-2 replication in lung epithelial cells. CBD acts after cellular infection, inhibiting viral gene expression and reversing many effects of SARS-CoV-2 on host gene transcription. CBD induces interferon expression and up-regulates its antiviral signaling pathway. A cohort of human patients previously taking CBD had significantly lower SARSCoV-2 infection incidence of up to an order of magnitude relative to matched pairs or the general population. This study highlights CBD, and its active metabolite, 7-OH-CBD, as potential preventative agents and therapeutic treatments for SARS-CoV-2 at early stages of infection

To test the effect of CBD on SARS-CoV-2 replication, we pretreated A549 human lung carcinoma cells expressing exogenous human ACE-2 receptor (A549-ACE2) for 2 hours with 0–10 μM CBD prior to infection with SARS-CoV-2. After 48 hours, we monitored cells for expression of the viral spike protein (S). For comparison, we also treated cells over a similar dose range with an MLK inhibitor (URMC-099) previously implicated as an antiviral for HIV and KPT-9274, a PAK4/NAMPT inhibitor that our analysis suggested might reverse many changes in gene expression caused by SARS-CoV-2. All three inhibitors potently inhibited viral replication under non-toxic conditions with EC50s ranging from 0.2–2.1 μM. CBD inhibited SARS-CoV-2 replication in Vero E6 monkey kidney epithelial cells as well. No toxicity was observed at the effective doses. We also determined that CBD suppressed replication of a related beta-coronavirus, mouse hepatitis virus (MHV), under non-toxic conditions with an EC50 of ~5 μM using A549 cells that express the MHV receptor (A549-MHVR), indicating the potential for more broader viral efficacy.

Our results suggest that CBD can block SARS-CoV-2 infection at early stages of infection, and CBD administration is associated with a lower risk of SARS-CoV-2 infection in humans. Furthermore, the active compound in patients is likely to be 7-OH-CBD, the same metabolite implicated in CBD treatment of epilepsy. The substantial reduction in SARS-CoV-2 infection risk of approximately an order of magnitude in patients who took FDA-approved CBD highlights the potential efficacy of this drug in combating SARS-CoV-2 infection. Finally, the ability of CBD to inhibit replication of MHV raises the possibility that CBD may have efficacy against new pathogenic viruses arising in the future. (Nguyen, 2021).

Cannabidiol inhibits SARS-Cov-2 spike (S) protein-induced cytotoxicity and inflammation through a PPARγ-dependent TLR4/NLRP3/Caspase-1 signaling suppression in Caco-2 cell line. Given the abundancy of angiotensin converting enzyme 2 (ACE-2) receptors density, beyond the lung, the intestine is considered as an alternative site of infection and replication for severe acute respiratory syndrome by coronavirus type 2 (SARS-CoV-2). Cannabidiol (CBD) has recently been proposed in the management of coronavirus disease 2019 (COVID-19) respiratory symptoms because of its anti-inflammatory and immunomodulatory activity exerted in the lung. In this study, we demonstrated the in vitro PPAR-γ-dependent efficacy of CBD (10-9 -10-7 M) in preventing epithelial damage and hyperinflammatory response triggered by SARS-CoV-2 spike protein (SP) in a Caco-2 cells. Immunoblot analysis revealed that CBD was able to reduce all the analyzed proinflammatory markers triggered by Spike Protein incubation, such as tool-like receptor 4 (TLR-4), ACE-2, family members of Ras homologues A-GTPase (RhoA-GTPase), inflammasome complex (NLRP3), and Caspase-1. CBD caused a parallel inhibition of interleukin 1 beta (IL-1β), IL-6, tumor necrosis factor alpha (TNF-α), and IL-18 by enzyme-linked immunosorbent assay (ELISA) assay. By immunofluorescence analysis, we observed increased expression of tight-junction proteins and restoration of transepithelial electrical resistance (TEER) following CBD treatment, as well as the rescue of fluorescein isothiocyanate (FITC)-dextran permeability induced by SP. Our data indicate, in conclusion, that CBD is a powerful inhibitor of SP protein enterotoxicity in vitro.

Molecules capable of reducing the hyperinflammatory response that characterizes the most severe clinical forms of COVID‐19 are strategically essential in supporting therapies against SARS‐CoV‐2 infections. Such molecules can significantly reduce the damage deriving from the cytokine storm and might strongly reduce the need for intensive care units for the most compromised patients. CBD is suitable for this purpose, and if orally administered, as the antiepileptic CBD‐based drug Epidiolex, it demonstrated itself capable of reducing the susceptibility caused by SARS‐CoV‐2. Moreover, as an anti‐inflammatory and immunoregulatory agent, CBD has shown the therapeutic potential of certain advantages in controlling the picture of lung inflammation from COVID‐19 and proving capable of markedly improve the symptoms of SARS‐CoV‐2 infection‐mimicking model by poly (I: C)‐induced ARDS in mice. Moreover, CBD demonstrated itself capable of increasing the expression of apelin, a peptide with a significant role in the central and peripheral regulation of the immune system, CNS, metabolic, and cardiovascular system. These effects, added to its known anti‐inflammatory properties identified at the lung level, are in line with other preclinical models of ARDS. In this sense, acting at the PPAR‐γ site, CBD has been proposed as a drug of potential applicability in anti‐COVID‐19 protocols, as demonstrated by an increasing number of clinical trials that examine its protective effect in SARS‐CoV‐2 infections (NCT04731116; NCT04686539). In addition to these previous publications, our study first demonstrates, as far we know, new and potential effects of CBD in the intestinal epithelium, potentially able to counteract SARS‐CoV‐2 hyper‐inflammation in the gut. We consider this point extremely interesting since the emerging importance of the intestine in viral replication, especially for SARS‐CoV‐2 variants, the role exerted by intestinal mucosa in the immune response to viral attack mediated by SP in this site. Although our data refer to an in vitro model, and further in vivo investigations are needed, they indicate additional potential beneficial effects by CBD against COVID‐19 that integrate with previously established lung protection. Moreover, by reducing ACE‐2 expression and down‐regulating RhoA‐GTPase/NLRP3/Caspase‐1 pathway, by a selective PPAR‐γ involvement, CBD may potentially counteract viral entry and replication by the intestine. At the same time, this molecule can reduce the over‐release of proinflammatory cytokine at the basis of systemic inflammatory dysfunction in most severe COVID‐19 cases.  (Corpetti, 2021).

Reference:

(Nguyen, 2021). Nguyen LC, Yang D, Nicolaescu V, et al. Cannabidiol Inhibits SARS-CoV-2 Replication and Promotes the Host Innate Immune Response. Preprint. bioRxiv. 2021;2021.03.10.432967. Published 2021 Mar 10. doi:10.1101/2021.03.10.432967 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7987002/

(Corpetti, 2021). Corpetti C, Del Re A, Seguella L, Palenca I, Rurgo S, De Conno B, Pesce M, Sarnelli G, Esposito G. Cannabidiol inhibits SARS-Cov-2 spike (S) protein-induced cytotoxicity and inflammation through a PPARγ-dependent TLR4/NLRP3/Caspase-1 signaling suppression in Caco-2 cell line. Phytother Res. 2021 Dec;35(12):6893-6903. doi: 10.1002/ptr.7302. Epub 2021 Oct 12. PMID: 34643000; PMCID: PMC8662250. https://pubmed.ncbi.nlm.nih.gov/34643000/

 

CANNABIGEROL (CBG)

CBG (Cannabigerol) mainly interacts with CB2 Receptors. CB2 receptors are predominantly found in the immune system. In late December 2019, a novel coronavirus (SARS-CoV-2 or CoV-19) appeared in Wuhan, China, causing a global pandemic. SARS-CoV-2 causes mild to severe respiratory tract inflammation, often developing into lung fibrosis with thrombosis in pulmonary small vessels and causing even death. Coronavirus Disease (COVID-19) patients manifest exacerbated inflammatory and immune responses, cytokine storm, prevalence of pro-inflammatory M1 macrophages and increased levels of resident and circulating immune cells. Men show higher susceptibility to SARS-CoV-2 infection than women, likely due to estrogens production. The protective role of estrogens, as well as an immune-suppressive activity that limits the excessive inflammation, can be mediated by cannabinoid receptor type 2 (CB2). The role of this receptor in modulating inflammation and immune response is well documented in fact in several settings. The stimulation of CB2 receptors is known to limit the release of pro-inflammatory cytokines, shift the macrophage phenotype towards the anti-inflammatory M2 type and enhance the immune-modulating properties of mesenchymal stromal cells. For these reasons, we hypothesize that CB2 receptor can be a therapeutic target in COVID-19 pandemic emergency.

The immune system has complex mechanisms to fulfill its function and respond to a variety of signaling molecules including hormones, neurotransmitters, and specific lipids, such as endocannabinoids (eCBs). The biological effects of cannabinoids are mediated through the activation of G-protein-coupled cannabinoid (CB) receptors. The endocannabinoid system (ECS) includes the cannabinoid receptor type 1 (CB1) and 2 (CB2), the endogenous cannabinoids, and the enzymes for their metabolism. CB1 is mostly expressed in the central nervous system and is strongly associated with the psychoactive effects of cannabinoids. CB1 is also expressed at low levels in peripheral tissues. Instead, CB2 is highly expressed by immune cells (B cells, natural killer cells, monocytes, neutrophils, CD8 lymphocytes, CD4 lymphocytes) and in several organs and tissues such as liver, spleen, nasal epithelium, thymus, brain, lung and kidney. Both CB1 and CB2 receptors have been widely demonstrated to be important modulators of the immune system, potentially inducing immunosuppression. CB2 is widely known for its immunomodulatory role, which is related to four events: (i) induction of apoptosis, (ii) suppression of cell proliferation, (iii) inhibition of proinflammatory cytokines production and increase in anti-inflammatory cytokines and (iv) induction of regulatory T cells. It is therefore conceivable that, also in COVID-19, the activation of the ECS plays a role in preventing and/or influencing the development and the severity of the disease. (Rossi, 2020).

Recently, Canadian researchers have tested CBD extracts of 800 different C. sativa lines on 3D human models of oral, airways, and intestinal tissues and found 13 low THC/high CBD lines that modulated ACE2 and TMPRSS2 levels, which might lower the virus load (128). ACE2-reducing activity of cannabis-derived products were confirmed by a different group. They extracted a CBD, CBG, and THCV-containing fraction of a C. sativa strain and tested it in vitro in comparison to a standard phytocannabinoid agent. Both products reduced the secretion of pro-inflammatory cytokines IL-6, IL-8, CCL2, and CCL7 from the alveolar epithelial cell line A549, induced polarization of the macrophage cell line KG1 and increased the phagocytosis. CD36 and type II receptor for the Fc region of IgG (FcγRII) were upregulated. The researchers reported a certain superiority of the standard phytocannabinoids compared to the cannabis-derived fraction but cannot give recommendations for usage of cannabis in the treatment of COVID-19. Another recent study simulated viral infections using the synthetic RNA Poly I:C and could show that Poly I:C-induced ARDS could be prevented by CBD through the upregulation of apelin, a peptide regulating central and peripheral immunity that was severely downregulated in a murine model of ARDS. In the following two sections we will present the processes of regulation of the immune responses of endo- and phytocannabinoids (Paland, 2021).

 

Protective Effects of Cannabigerol on Cells of the Blood–Brain Barrier

Cannabidiol (CBD), one of the chemicals found in Cannabis sativa, has displayed a range of neuroprotective qualities, preventing neuronal loss, attenuating astrocyte reactivity, and dampening the neuroinflammatory response. Unlike delta 9 tetrahydrocannabinol (D9 -THC), CBD does not activate the central cannabinoid receptors, CB1 or CB2, but activates a plethora of other targets including PPAR-c, TRPV1, and 5-HT1A receptors. CBD has formulations (alone and with D9 -THC) licensed by GW pharmaceuticals to treat rare childhood epilepsies and spasticity associated with multiple sclerosis. The protective effects of CBD in stroke models has been well documented, specifically CBD has been shown to reduce infarct volume, reduce glutamate toxicity, attenuate mitochondrial dysfunction and glial activation. In a co-culture BBB model CBD preserved barrier integrity after oxygen-glucose deprivation (OGD), which was mediated at least in part by PPAR-c and 5-HT 1A receptors. Cannabigerol (CBG) and cannabidivarin (CBDV) are neutral cannabinoids present in cannabis and studies have found these compounds share similar pharmacological characteristics to CBD. Like CBD, they do not produce feelings of euphoria and display antioxidant and anti-inflammatory properties, as well as interacting with a range of target proteins including TRPV1, 13 PPAR-c, 20 5-HT1A , and CB2 . Recently our group conducted a systematic review focusing on the neuroprotective properties of minor phytocannabinoids (other than D9 -THC or CBD) and found that CBG and CBDV show efficacy in models of Huntington’s disease, Alzheimer’s, and epilepsy, with CBG mediating its protective effects through PPAR-c activation, the same mechanism by which we have shown that CBD protects BBB integrity. CBG has displayed prominent anti-inflammatory and antioxidant capabilities.

In astrocytes CBG and CBDV attenuated levels of interleukin-6 (IL-6) spike protein and lactate dehydrogenase (LDH). In astrocytes, CBG decreased levels of DNA damage proteins (spike proteins) (Stone, 2021).

Reference:

(Rossi, 2020). Rossi F, Tortora C, Argenziano M, Di Paola A, Punzo F. Cannabinoid Receptor Type 2: A Possible Target in SARS-CoV-2 (CoV-19) Infection? Int J Mol Sci. 2020 May 27;21(11):3809. doi: 10.3390/ijms21113809. PMID: 32471272; PMCID: PMC7312493. Cannabinoid Receptor Type 2: A Possible Target in SARS-CoV-2 (CoV-19) Infection? – PubMed (nih.gov).

(Paland, 2021). Paland N, Pechkovsky A, Aswad M, et al. The Immunopathology of COVID-19 and the Cannabis Paradigm. Front Immunol. 2021;12:631233. Published 2021 Feb 12. doi:10.3389/fimmu.2021.631233

(Stone, 2021). Nicole L. Stone, Timothy J. England, and Saoirse E. O’Sullivan.Cannabis and Cannabinoid Research.Aug 2021.315-326.http://doi.org/10.1089/can.2020.0159