Leukocytes are part of the tumor microenvironment (TME) and are critical determinants of tumor progression. Because of the immunoregulatory properties of cannabinoids, the endocannabinoid system (ECS) may have an important role in shaping the TME. Members of the ECS, an entity that consists of cannabinoid receptors, endocannabinoids and their synthesizing/degrading enzymes, have been associated with both tumor growth and rejection. Immune cells express cannabinoid receptors and produce endocannabinoids, thereby forming an “immune endocannabinoid system”. Although in vitro effects of exogenous cannabinoids on immune cells are well described, the role of the ECS in the TME, and hence in tumor development and immunotherapy, is still elusive. This review/opinion discusses the possibility that the “immune endocannabinoid system” can fundamentally influence tumor progression. The widespread influence of cannabinoids on immune cell functions makes the members of the ECS an interesting target that could support immunotherapy.

Cannabinoid (CB) receptors (CB₁and CB₂) are expressed on cancer cells and their expression influences carcinogenesis in various tumor entities. Cells of the tumor microenvironment (TME) also express CB receptors, however, their role in tumor development is still unclear. We, therefore, investigated the role of TME-derived CB₁and CB₂receptors in a model of non-small cell lung cancer (NSCLC). Leukocytes in the TME of mouse and human NSCLC express CB receptors, with CB₂showing higher expression than CB₁. In the tumor model, using CB₁- (CB₁^-/-) and CB₂-knockout (CB₂^-/-) mice, only deficiency of CB₂, but not of CB₁, resulted in reduction of tumor burden vs. wild type (WT) littermates. This was accompanied by increased accumulation and tumoricidal activity of CD8⁺T and natural killer cells, as well as increased expression of programmed death-1 (PD-1) and its ligand on lymphoid and myeloid cells, respectively. CB₂^-/-mice responded significantly better to anti-PD-1 therapy than WT mice. The treatment further increased infiltration of cytotoxic lymphocytes into the TME of CB₂^-/-mice. Our findings demonstrate that TME-derived CB₂dictates the immune cell recruitment into tumors and the responsiveness to anti-PD-1 therapy in a model of NSCLC. CB₂could serve as an adjuvant target for immunotherapy.

Lung cancer is the leading cause of cancer-related death worldwide. Discoidin domain receptor 1 (DDR1), a tyrosine kinase receptor, has been associated with poor prognosis in patients with non-small cell lung cancer (NSCLC). However, its role in tumorigenesis remains poorly understood. This work aimed to explore the impact of DDR1 expression on immune cell infiltration in lung adenocarcinoma. Pharmacological inhibition and knockout of DDR1 were used in an immunocompetent mouse model of KRAS/p53-driven lung adenocarcinoma (LUAD). Tumor cells were engrafted subcutaneously, after which tumors were harvested for investigation of immune cell composition via flow cytometry. The Cancer Genome Atlas (TCGA) cohort was used to perform gene expression analysis of 509 patients with LUAD. Pharmacological inhibition and knockout of DDR1 increased the tumor burden, with DDR1 knockout tumors showing a decrease in CD8⁺ cytotoxic T cells and an increase in CD4⁺ helper T cells and regulatory T cells. TCGA analysis revealed that low-DDR1-expressing tumors showed higher FoxP3 (regulatory T-cell marker) expression than high-DDR1-expressing tumors. Our study showed that under certain conditions, the inhibition of DDR1, a potential therapeutic target in cancer treatment, might have negative effects, such as inducing a pro-tumorigenic tumor microenvironment. As such, further investigations are necessary.

The tumor microenvironment (TME) is pivotal in cancer progression and the response to immunotherapy. A “hot” tumor typically contains immune cells that promote anti-tumor immunity, predicting positive prognosis. “Cold” tumors lack immune cells, suggesting a poor outlook across various cancers. Recent research has focused on converting “cold” tumors into “hot” tumors to enhance the success of immunotherapy. A prerequisite for the studies of the TME is an accurate knowledge of the cell populations of the TME. This study aimed to describe the immune TME of lung and colorectal cancer and melanoma, focusing on lymphoid and myeloid cell populations. We induced heterotopic immunocompetent tumors in C57BL/6 mice, using KP and LLC (Lewis lung carcinoma) cells for lung cancer, MC38 cells for colorectal cancer, and B16-F10 cells for melanoma. Immune cell infiltration was analyzed using multicolor flow cytometry in single-cell suspensions after tumor excision. KP cell tumors showed an abundance of neutrophils and eosinophils; however, they contained much less adaptive immune cells, while LLC cell tumors predominated in monocytes, neutrophils, and monocyte-derived dendritic cells. Monocytes and neutrophils, along with a significant T cell infiltration, were prevalent in MC38 tumors. Lastly, B16-F10 tumors were enriched in macrophages, while showing only moderate T cell presence. In conclusion, our data provide a detailed overview of the immune TME of various heterotopic tumors, highlighting the variabilities in the immune cell profiles of different tumor entities. Our data may be a helpful basis when investigating new immunotherapies, and thus, this report serves as a helpful tool for preclinical immunotherapy research design.

Background: Cannabinoids are mainly used for recreational purposes, but also made their way into oncology, since these substances can be taken to increase appetite in tumour cachexia. Since there are some hints in the literature that cannabinoids might have some anti-cancerous effects, the aim of this study was to study if and how cannabinoids mediate pro-apoptotic effects in metastatic melanoma in vivo and in vitro and its value besides conventional targeted therapy in vivo. Methods: Several melanoma cell lines were treated with different concentrations of cannabinoids, and anti-cancerous efficacy was assessed by proliferation and apoptosis assays. Subsequent pathway analysis was performed using apoptosis, proliferation, flow cytometry and confocal microscopy data. The efficacy of cannabinoids in combination with trametinib was studied in NSG mice in vivo. Results: Cannabinoids reduced cell viability in multiple melanoma cell lines in a dose-dependent way. The effect was mediated by CB1, TRPV1 and PPARα receptors, whereby pharmacological blockade of all three receptors protected from cannabinoid-induced apoptosis. Cannabinoids initiated apoptosis by mitochondrial cytochrome c release with consecutive activation of different caspases. Essentially, cannabinoids significantly decreased tumour growth in vivo and were as potent as the MEK inhibitor trametinib. Conclusions: We could demonstrate that cannabinoids reduce cell viability in several melanoma cell lines, initiate apoptosis via the intrinsic apoptotic pathway by cytochrome c release and caspase activation and do not interfere with commonly used targeted therapy.