Role of the type 2 cannabinoid receptor in GvHD

T-cells, as part of the adaptive immune system, have been labelled the main drivers of graft-versus-host disease (GvHD) development. Although current therapeutic strategies for GvHD are largely focused on the modulation of the adaptive immune system, it has become clear that components of the innate immune system are also essential for the progression of GvHD.

The type 2 cannabinoid receptor (CB2R) is a G protein-coupled receptor expressed on almost all immune cells, and controls cytokine release, macrophage-mediated responses, and B- and T-cell differentiation. In humans, genetic perturbances in the CB2R gene have been associated with inflammatory disorders; therefore, targeting CB2R therapeutically is an area of interest in immune-related disease states such as GvHD.

Cheng Yin Yuan and colleagues implemented a pre-clinical study to define the role of CB2R in acute and chronic GvHD (a/cGvHD) development and to determine whether pharmacological modulation of CB2R could alleviate GvHD in murine models. Their results have recently been published in Blood.¹

Study design

• CB2R^−/− mice were used in transplantation studies (as a donor or recipient) to determine the role of CB2R signaling in GvHD pathobiology.
• GvHD target organs were examined to determine the impact of CB2R expression on organ-specific immune-cell populations.
• A selective CB2R antagonist, SR144528, was used to determine the impact of CB2R signaling on disease severity.
• CB2R^EGFP reporter mice were used to identify CB2R-expressing cells in GvHD target organs.
• Two murine sclerodermatous cGvHD models were used to explore the role of CB2R in cGvHD.

Results

CB2R expression correlates with aGvHD severity in colon and liver

• No differences were observed between wild type (WT) and CB2R^−/− mice in different T- and B-cell compartments.
• Mice transplanted with bone marrow and spleen cells from CB2R^−/− donors demonstrated
  • accelerated GvHD related mortality;
  • a significant increase in CD8⁺ T cells and CD8⁺ IFN-ϒ⁺ T cells in the colon; and
  • no change in CD4⁺ T cell numbers in the colon and liver and a decrease in the lungs.
• By contrast, survival was comparable between CB2R^−/− and WT recipient mice receiving WT grafts.
• Increased GvHD mortality was also seen in transplant models where SR144528, a selective CB2R antagonist, was administered.
• Transplanted mice reconstituted with CB2R^−/− CD4⁺ or CD8⁺ T cells vs WT T cells showed
  • shorter survival;
  • elevated levels of pro-inflammatory CD8⁺ T cells in the colon, liver, and lungs, and worsened GvHD in the colon and liver when transplanted with CB2R^−/− CD8⁺ T-cells; and
  • elevated levels of CD8⁺ T cells in the colon and lungs and worsened GvHD in the liver when transplanted with CB2R^−/− CD4⁺ T cells.
• Transplantation of CB2R^EGFP reporter mice revealed a significant loss of CB2R expression in CD4⁺ and CD8⁺ T cells isolated from GvHD-affected organs.

Treatment with phytocannabinoid tetrahydrocannabinol (THC) improves survival in aGvHD models

• aGvHD did not result in reduced production of endogenous CB2R agonists.
• Treatment with a synthetic CB2R selective cannabinoid, JWH-133, had no effect on survival and did not protect mice from lethal GvHD.
• Treatment with the natural low affinity CB1R and CB2R agonist, THC, significantly improved the survival rate of GvHD mice transplanted with WT cells, while there was no survival advantage with THC treatment in mice transplanted with CB2R^−/− marrow grafts.
• Treatment with THC resulted in significantly reduced numbers of donor INF-γ-producing CD4⁺ and CD8⁺ T cells in the colon and liver.

Impact of CB2R expression in two models of sclerodermatous skin cGvHD

• Pharmacological antagonism of CB2R with SR144528 resulted in significantly worse cGvHD pathology with increased hyperkeratosis and inflammatory cell infiltrates (mostly CD3⁺ cells and macrophages) in the skin.
• Macrophages from SR144528-treated GvHD mice demonstrated significantly elevated levels of the expression of genes that are related to fibrosis, such as TGF-β, galectin 3, and alpha smooth muscle actin (α-SMA).
• GvHD-mediated damage was heightened in mice transplanted with CB2R^−/− marrow grafts as evidenced by heightened dermal fibrosis, increased infiltration of inflammatory cells, and loss of hair follicles.

Treatment with selective CB2R agonist JWH-133 reduces tissue damage in cGvHD models

• There was no difference in CB2R expression on macrophages isolated from the spleen, lung, or liver of GvHD animals vs control mice.
• Pharmacological activation of CB2R using JWH-133 significantly decreased pathological damage, and TGF-β and α-SMA levels, while THC failed to protect against cGvHD.

Key messages

• Although CB2R-knockout mice have no obvious changes in T cell compartment composition, CB2R expression on donor cells is important for the regulation of CD8⁺ T cell alloreactivity.
• Inhibition or loss of CB2R expression on donor cells aggravates aGvHD and cGvHD severity.
• In aGvHD target organs, CB2R expression on CD8⁺ T cells regulates CD8⁺ T cell expansion and inflammatory cytokine production.

• THC, but not JWH-133, was efficacious in protecting against lethal aGvHD.
• JWH-133, but not THC, was efficacious in reducing cGvHD.

Conclusion

The findings from this study highlight the role of CB2R in immune modulation. A lack of CB2R on donor T cells and pharmacological antagonism of CB2R worsened GvHD and led to the expansion of CD8⁺ T cells. This indicates that CB2R expression on T cells regulates CD8⁺ T cell alloreactivity.

The study also uncovered a decline in CB2R expression on T cells, but not macrophages, following aGvHD initiation. This provides information regarding the receptor’s expression patterns in inflammatory conditions.

The CB2R agonists, THC and JWH-133, alleviated T cell-mediated aGvHD and macrophage-driven cGvHD, respectively. The differential disease biology between the two conditions highlight the distinct mechanisms of action of the agonists. The study suggests that effective therapeutic intervention is dependent on the agonist properties in combination with the composition of pathogenic immune cells present in a/cGvHD.