Effects of Phytocannabinoids on Immune Response and Autophagy During Chronic Immune-mediated Inflammatory Diseases

Cannabis, in addition to its psychotropic properties, could have anti-inflammatory and immunomodulatory effects. Phytocannabinoids (pCBs) are a group of molecules naturally secreted by the cannabis plant. The major pCBs are cannabidiol (CBD) and Δ9-tetrahydrocannabinol Δ9 (THC). Only THC has psychotropic effects, which CBD does not have. Alongside these two main components, there is a wide variety of other molecules, such as other pCBs and terpenes which could increase the effects on immune system through synergistic interactions between these different compounds ("entourage effect").In vivo, pCBs essentially interfere with the endocannabinoid system, acting on many ubiquitous receptors, present on a significant number of different cell types. Numerous published studies show that pCBs have immunomodulatory and anti-inflammatory properties by acting on several of these receptors, whether through modulation of the immune response of different cell types, effects on cytokine networks, reduction of innate and adaptive responses and/or impact on cell survival or death (autophagy, proliferation/ apoptosis). The Immune-Mediated Inflammatory Diseases (IMIDs) affect 5 to 7% of the general population in Western countries, involve different organs (joints, skin, digestive tract) but share the same inflammatory mechanisms resulting from a dysregulation of the immune response. Our research focuses on the identification of the most effective phytochemical profile of pCBs, allowing an optimal effect on chronic inflammatory pathologies of interest among immune-mediated chronic inflammatory diseases (IMIDs). The pCB-IMIDs project is therefore part of an innovative translational project, around new therapeutic applications of medical cannabis (CannAppIMIDs). In our study, we will include 100 patients with one of IMIDs among Rheumatoid Arthritis, spondylarthritis, psoriatic arthritis, Sjogren disease and systemic lupus, at different stage and with different treatments. After patient's consent we will collect for research purposes an additional 40 ml of blood during a routine care blood test. Mononuclear and polynucleated blood cells will be exposed in vitro to different full-spectrum pCB extracts (full spectrum extract) including a CBD dominant and low THC extract (\<0.2%), 1 dominant THC extract, 1 balanced THC/CBD extract and 1 dominant CBG extract. In this cross-sectional study, our objective will be to assess the biological effects of different pCB compositions on inflammatory profiles (concentrations of pro and anti-inflammatory cytokines and chemokines) and modulations of expression profiles (autophagy, apoptosis, and cannabinoid receptor expression profile).

Background:

The analgesic and anxiolytic properties of cannabis are at the origin of the French experiment on medical cannabis set up in 2020 and led by the ANSM (National Agency for the Safety of Medicines and Health Products) for 5 well-identified pathologies (neuropathic pain, certain forms of pediatric epilepsy, symptoms linked to cancer or anti-cancer treatment, palliative situations and painful spasticity linked to multiple sclerosis). Phytocannabinoids (pCBs) are a group of molecules naturally secreted by the cannabis plant. The main pCBs are cannabidiol (CBD) and Δ9-tetrahydrocannabinol Δ9-THC \[1, 2\]. Only THC has psychotropic effects, which CBD does not have. Alongside these two main components, there is a wide variety of molecules \[cannabigerol (CBG), cannabinol (CBN), cannabichromene (CBC) and other terpenes\] which are said to have an "entourage effect" by increasing the effects by synergistic interactions between these different compounds \[3-7\]. CBD of natural origin can be purified without other terpenes corresponding to CBD alone ("CBD isolate") or is combined with terpenes and phytocannabinoids but without THC or any other pCB ("broad spectrum"), or in unpurified form, CBD with terpenes and THC ("full spectrum").

In vivo, pCBs essentially interfere with the endocannabinoid system, acting on numerous receptors present on a large number of different cell types \[8\]. This means that pCBs can be used to treat a wide range of organs. Numerous published works show that pCBs have immunomodulatory and anti-inflammatory properties by acting on several of these receptors, whether via modulation of the immune response of different cell types, effects on cytokine networks, reduction of innate and adaptive responses \[9, 10\] and/or impact on cell survival or death (autophagy, proliferation/apoptosis) \[8, 9\]. Thus, in addition to its neurological effects \[10, 11\], cannabis possesses anti-inflammatory, pro-autophagic and anti-proliferative properties, which are still imperfectly characterized.

Understanding the mechanisms of action of pCBs, individually or in combination with other molecules present in cannabis, on different therapeutic targets therefore presents an immense challenge in the management of Immune-Mediated Inflammatory Diseases (IMIDs). These IMIDs affect 5 to 7% of the general population in Western countries, and involve different organs (joints, skin, gut) but share the same inflammatory mechanisms, resulting from a deregulation of the immune response.

The pCB-IMIDs project is part of an innovative translational project focusing on new therapeutic applications for medical cannabis (CannAppIMIDs).

Objectives:

The objective of this exploratory project is to evaluate the biological and immunological effects of exposure of blood cells from patients suffering from IMID to different varieties of pCB extracts on:

1. The inflammatory profile (main objective)
2. The expression profile of cannabinoid receptors
3. The autophagic and apoptotic profile

Methods: Single-center cross-sectional study without prospective follow-up of participants, classified non-interventional study. The study consists, after obtaining consent and verifying the eligibility criteria (with saliva test specific to the study), in taking an additional 40 ml of blood during a planned blood test for research purposes and to collect medical data on patients and their inflammatory pathology.

Mononuclear and polynuclear blood cells from subjects suffering from IMID will be exposed in vitro to different inflammatory inducers used individually and/ or all together (LPS- bacterial Lipopolysaccahride, TLR-4 ligand and/or poly-I:C , TLR-3 ligand and/or PAM3CSK4, TLR1/TLR2 ligand, and/or PHA-P - phytohemagglutin P, polysaccharide and glycoprotein ligand) with and without the concomitant addition of different complete spectrum of pCB extracts (full spectrum) including a dominant CBD extract and low in THC (\<0.2%), 1 dominant THC extract, 1 balanced THC/CBD extract and 1 dominant CBG extract. The same analyzes will be carried out in the presence of a recognized anti-inflammatory (for example hydrocortisone or dexamethasone) which will thus serve as a positive control \[12\].

All analyses will be carried out at CBM - UPR4301 CNRS. Results will allow us to formulate hypotheses, guide and argue for future clinical research studies evaluating the benefit(s) and tolerance of extracts rich in pCB in taking in therapeutic management of IMIDs.

Eligibility Criteria:
Inclusion Criteria:

1. Male or female ≥ 18 years old
2. Diagnosis confirmed by a rheumatologist of RA or spondyloarthropathy (with or without IBD) or psoriatic arthritis (with or without active psoriasis) or systemic lupus erythematosus or Sjögren's disease
3. Patient who has expressed consent to participate in the study
4. Patients affiliated to social security
5. Treatments authorized as part of routine care: non-steroidal anti-inflammatory drugs (NSAIDs), level 1 to 3 analgesics, local corticosteroids, oral corticosteroid therapy (daily dose ≤15 mg/d), 5-aminosalicylic acid, salazopyrine, methotrexate, leflunomide, hydroxychloroquine, biotherapies and targeted therapies.

Exclusion Criteria:

1. Patient who received intravenous corticosteroid therapy less than 4 weeks ago
2. Patient receiving oral corticosteroid therapy with a daily dose \>15 mg/day
3. Consumption of CBD and/or recreational cannabis and/or positive saliva test for cannabis consumption and/or CBD
4. Pregnant and lactating women
5. Persons under guardianship or curatorship