Chiara Laezza

Antitumor Activity of Plant Cannabinoids with Emphasis on the Effect of Cannabidiol on Human Breast Carcinoma

Δ9-Tetrahydrocannabinol (THC) exhibits antitumor effects on various cancer cell types, but its use in chemotherapy is limited by its psychotropic activity. We investigated the antitumor activities of other plant cannabinoids, i.e., cannabidiol, cannabigerol, cannabichromene, cannabidiol acid and THC acid, and assessed whether there is any advantage in using Cannabis extracts (enriched in either cannabidiol or THC) over pure cannabinoids. Results obtained in a panel of tumor cell lines clearly indicate that, of the five natural compounds tested, cannabidiol is the most potent inhibitor of cancer cell growth (IC50 between 6.0 and 10.6 μM), with significantly lower potency in noncancer cells. The cannabidiol-rich extract was equipotent to cannabidiol, whereas cannabigerol and cannabichromene followed in the rank of potency. Both cannabidiol and the cannabidiol-rich extract inhibited the growth of xenograft tumors obtained by s.c. injection into athymic mice of human MDA-MB-231 breast carcinoma or rat v-K-ras-transformed thyroid epithelial cells and reduced lung metastases deriving from intrapaw injection of MDA-MB-231 cells. Judging from several experiments on its possible cellular and molecular mechanisms of action, we propose that cannabidiol lacks a unique mode of action in the cell lines investigated. At least for MDA-MB-231 cells, however, our experiments indicate that cannabidiol effect is due to its capability of inducing apoptosis via: direct or indirect activation of cannabinoid CB2 and vanilloid transient receptor potential vanilloid type-1 receptors and cannabinoid/vanilloid receptor-independent elevation of intracellular Ca2+ and reactive oxygen species. Our data support the further testing of cannabidiol and cannabidiol-rich extracts for the potential treatment of cancer.

Suppression of Nerve Growth Factor Trk Receptors and Prolactin Receptors by Endocannabinoids Leads to Inhibition of Human Breast and Prostate Cancer Cell Proliferation

Anandamide and 2-arachidonoylglycerol (2-AG), two endogenous ligands of the CB1 and CB2 cannabinoid receptor subtypes, inhibit the proliferation of PRL-responsive human breast cancer cells (HBCCs) through down-regulation of the long form of the PRL receptor (PRLr). Here we report that 1) anandamide and 2-AG inhibit the nerve growth factor (NGF)-induced proliferation of HBCCs through suppression of the levels of NGF Trk receptors; 2) inhibition of PRLr levels results in inhibition of the proliferation of other PRL-responsive cells, the prostate cancer DU-145 cell line; and 3) CB1-like cannabinoid receptors are expressed in HBCCs and DU-145 cells and mediate the inhibition of cell proliferation and Trk/PRLr expression. β-NGF-induced HBCC proliferation was potently inhibited (IC50 = 50–600 nm) by the synthetic cannabinoid HU-210, 2-AG, anandamide, and its metabolically stable analogs, but not by the anandamide congener, palmitoylethanolamide, or the selective agonist of CB2 cannabinoid receptors, BML-190. Th...

Pharmacological Actions of Statins: A Critical Appraisal in the Management of Cancer

Statins, among the most commonly prescribed drugs worldwide, are cholesterol-lowering agents used to manage and prevent cardiovascular and coronary heart diseases. Recently, a multifaceted action in different physiological and pathological conditions has been also proposed for statins, beyond anti-inflammation and neuroprotection. Statins have been shown to act through cholesterol-dependent and -independent mechanisms and are able to affect several tissue functions and modulate specific signal transduction pathways that could account for statin pleiotropic effects. Typically, statins are prescribed in middle-aged or elderly patients in a therapeutic regimen covering a long life span during which metabolic processes, aging, and concomitant novel diseases, including cancer, could occur. In this context, safety, toxicity, interaction with other drugs, and the state of health have to be taken into account in subjects treated with statins. Some evidence has shown a dichotomous effect of statins with either cancer-inhibiting or -promoting effects. To date, clinical trials failed to demonstrate a reduced cancer occurrence in statin users and no sufficient data are available to define the long-term effects of statin use over a period of 10 years. Moreover, results from clinical trials performed to evaluate the therapeutic efficacy of statins in cancer did not suggest statin use as chemotherapeutic or adjuvant agents. Here, we reviewed the pharmacology of the statins, providing a comprehensive update of the current knowledge of their effects on tissues, biological processes, and pathological conditions, and we dissected the disappointing evidence on the possible future use of statin-based drugs in cancer therapy.

Inhibitory effects of cannabinoid CB1 receptor stimulation on tumor growth and metastatic spreading: actions on signals involved in angiogenesis and metastasis

Stimulation of cannabinoid CB1 receptors by 2‐methyl‐arachidonyl‐2′‐fluoro‐ethylamide (Met‐F‐AEA) inhibits the growth of a rat thyroid cancer cell‐derived tumor in athymic mice by inhibiting the activity of the oncogene product p21ras. Here we report that Met‐F‐AEA also blocks the growth of tumors previously induced in nude mice by the s.c. injection of the same rat thyroid carcinoma cells. Met‐F‐AEA significantly inhibited, in tumors as well as transformed cells, the expression of the vascular endothelial growth factor, an angiogenetic factor known to be up‐regulated by p21ras, as well as of one of its receptors, flt‐1/VEGFR‐1. The levels of the cyclin‐dependent kinase inhibitor p27(kip1), which is down‐regulated by p21ras, were instead increased by Met‐F‐AEA. All these effects were antagonized by the selective CB1 receptor antagonist SR141716A. Met‐F‐AEA inhibited in vitro the growth of a metastasis‐derived thyroid cancer cell line more potently than a primary cancer cell line. Therefore, the hypothesis that CB1 receptor stimulation interferes not only with angiogenesis but also with metastatic processes was tested in a widely used model of metastatic infiltration in vivo, the Lewis lung carcinoma (3LL) in C57Bl/6 mice. Three weeks from the paw injection of 3LL cells, Met‐F‐AEA reduced significantly the number of metastatic nodes, in a way antagonized by SR141716A. Our findings indicate that CB1 receptor agonists might be used therapeutically to retard tumor growth in vivo by inhibiting at once tumor growth, angiogenesis, and metastasis.

A new strategy to block tumor growth by inhibiting endocannabinoid inactivation

Endocannabinoid signaling has been shown to be enhanced in several cancer tissues and malignant cells, and studies in cell lines have shown that this up‐regulation might serve the purpose of providing transformed cells with a further means to inhibit their proliferation. Here we investigated the effect of inhibitors of endocannabinoid degradation on the growth of rat thyroid tumor xenografts induced in athymic mice. VDM‐11, a selective inhibitor of endocannabinoid cellular re‐uptake, and arachidonoyl‐serotonin (AA‐5‐HT), a selective blocker of endocannabinoid enzymatic hydrolysis, both inhibited the growth in vivo of tumor xenografts induced by the subcutaneous injection of rat thyroid transformed (KiMol) cells. This effect was accompanied by significantly enhanced endocannabinoid concentrations in the tumors excised at the end of the in vivo experiments. Endocannabinoids, as well as VDM‐11 and AA‐5‐HT, inhibited the growth in vitro of the transformed rat thyroid cells used to induce the tumors in vivo, and their effect was reversed at least in part by the cannabinoid CB1 receptor antagonist SR141716A. This compound, however, when administered alone, did not enhance, but instead slightly inhibited, the growth of rat thyroid transformed cells both in vitro and in tumor xenografts induced in vivo. These findings indicate that endocannabinoids tonically control tumor growth in vivo by both CB1‐mediated and non‐CB1‐mediated mechanisms and that, irrespective of the molecular mechanism of their anti‐proliferative action, inhibitors of their inactivation might be used for the development of novel anti‐cancer drugs.

Control by the endogenous cannabinoid system of ras oncogene-dependent tumor growth

We investigated the effect of 2‐methyl‐arachidonyl‐2′‐fluoro‐ethylamide (Met‐F‐AEA), a stable analog of the endocannabinoid anandamide, on a rat thyroid epithelial cell line (FRTL‐5) transformed by the K‐ras oncogene, and on epithelial tumors derived from these cells. Met‐F‐AEA effect in vivo was evaluated in a nude mouse xenograft model, where K‐ras‐transformed (KiMol) cells were implanted subcutaneously. Met‐F‐AEA (0.5 mg/kg/dose) induced a drastic reduction in tumor volume. This effect was inhibited by the CBi receptor antagonist SR141716A (0.7 mg/kg/dose) and was accompanied by a strong reduction of K‐ras activity. Accordingly, KiMol cells and tumors express CB1 receptors. Met‐F‐AEA inhibited (IC50 ~5 μM) the proliferation in vitro and the transition to the S phase of KiMol cells and it reduced K‐ras activity; these effects were antagonized by SR141716A. Met‐F‐AEA cytostatic action was significantly smaller in nontransformed FRTL‐5 cells than in KiMol cells. Met‐F‐AEA treatment exerted opposite effects on the expression of CB1 receptors in KiMol and FRTL‐5 cells, with a strong up‐regulation in the former case and a suppression in nontransformed cells. The data suggest that: 1) Met‐F‐AEA inhibits ras oncogene‐dependent tumor growth in vivo through CB1 cannabinoid receptors; and 2) responsiveness of FRTL‐5 cells to endocannabinoids depends on whether or not they are transformed by K‐ras.

Cannabinoids and cancer: pros and cons of an antitumour strategy

In the last two decades, research has dramatically increased the knowledge of cannabinoids biology and pharmacology. In mammals, compounds with properties similar to active components of Cannabis sativa, the so called ‘endocannabinoids’, have been shown to modulate key cell‐signalling pathways involved in cancer cell growth, invasion and metastasis. To date, cannabinoids have been licensed for clinical use as palliative treatment of chemotherapy, but increased evidences showed direct antiproliferative actions of cannabinoid agonists on several tumour cells in vitro and in animal models. In this article, we will review the principal molecular pathways modulated by cannabinoids on cancer and summarize pros and cons evidence on the possible future use of endocannabinoid‐based drugs in cancer therapy.

The Cannabinoid CB1 Receptor Antagonist Rimonabant (SR141716) Inhibits Human Breast Cancer Cell Proliferation through a Lipid Raft-Mediated Mechanism

The endocannabinoid system has been shown to modulate key cell-signaling pathways involved in cancer cell growth. In this study, we show that cannabinoid receptor type 1 (CB1) antagonist Rimonabant (SR141716) inhibited human breast cancer cell proliferation, being more effective in highly invasive metastatic MDA-MB-231 cells than in less-invasive T47D and MCF-7 cells. The SR141716 antiproliferative effect was not accompanied by apoptosis or necrosis and was characterized by a G1/S-phase cell cycle arrest, decreased expression of cyclin D and E, and increased levels of cyclin-dependent kinase inhibitor p27KIP1. We have also shown that SR141716 exerted a significant antiproliferative action, in vivo, by reducing the volume of xenograft tumors induced by MDA-MB-231 injection in mice. On the other hand, at the concentration range in which we observed the antiproliferative effect in tumor cells, we did not observe evidence of any genotoxic effect on normal cells. Our data also indicate that the SR141716 antiproliferative effect requires lipid raft/caveolae integrity to occur. Indeed, we found that CB1 receptor (CB1R) is completely displaced from lipid rafts in SR141716-treated MDA-MB-231 cells, and cholesterol depletion by methyl-β-cyclodextrin strongly prevented SR141716-mediated antiproliferative effect. Taken together, our results suggest that SR141716 inhibits human breast cancer cell growth via a CB1R lipid raft/caveolae-mediated mechanism.

The endocannabinoid signaling system in cancer

Changes in lipid metabolism are intimately related to cancer. Several classes of bioactive lipids play roles in the regulation of signaling pathways involved in neoplastic transformation and tumor growth and progression. The endocannabinoid system, comprising lipid-derived endocannabinoids, their G-protein-coupled receptors (GPCRs), and the enzymes for their metabolism, is emerging as a promising therapeutic target in cancer. This report highlights the main signaling pathways for the antitumor effects of the endocannabinoid system in cancer and its basic role in cancer pathogenesis, and discusses the alternative view of cannabinoid receptors as tumor promoters. We focus on new players in the antitumor action of the endocannabinoid system and on emerging crosstalk among cannabinoid receptors and other membrane or nuclear receptors involved in cancer. We also discuss the enzyme MAGL, a key player in endocannabinoid metabolism that was recently recognized as a marker of tumor lipogenic phenotype.

The cannabinoid CB1 receptor antagonist rimonabant stimulates 2-deoxyglucose uptake in skeletal muscle cells by regulating the expression of phosphatidylinositol-3-kinase.

The endocannabinoid system regulates food intake, energy, and glucose metabolism at both central and peripheral levels. We have investigated the mechanism by which it may control glucose uptake in skeletal muscle cells. Detectable levels of the cannabinoid receptor type 1 (CB1) were revealed in L6 cells. Exposure of differentiated L6 myotubes to the CB1 antagonist rimonabant (SR141716) selectively increased 2-deoxyglucose uptake (2-DG) in a time- and dose-dependent manner. A similar effect was induced by genetic silencing of CB1 by small interfering RNA. Protein expression profiling revealed that both the regulatory p85 and the catalytic p110 subunits of the phosphatidylinositol-3-kinase (PI3K) were increased by SR141716. No significant change in the cellular content of other known molecules regulating PI3K was observed. However, phosphoinositide-dependent kinase-1, Akt/protein kinase B, and protein kinase Cζ activities were rapidly induced after SR141716 treatment of L6 cells in a PI3K-dependent manner. The stimulatory effect of SR141716 on PI3K expression and activity was largely prevented by N-[2-(4-bromocinnamylamino)ethyl]-5-isoquinoline (H-89), an inhibitor of the cAMP-dependent protein kinase. Moreover, SR141716-stimulated 2-DG uptake was blunted by the coincubation either with H-89 or with the PI3K inhibitor 2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride (LY294002), both in L6 cells and in mouse primary myocytes. Thus, modulation of CB1 regulates glucose uptake at the level of the PI3K signaling system in skeletal muscle cells. Interfering with CB1 signaling may therefore ameliorate glucoregulatory functions in peripheral tissues.