Natalya M. Kogan

A Cannabinoid Quinone Inhibits Angiogenesis by Targeting Vascular Endothelial Cells

Recent findings on the inhibition of angiogenesis and vascular endothelial cell proliferation by anthracycline antibiotics, which contain a quinone moiety, make this type of compound a very promising lead in cancer research/therapy. We have reported that a new cannabinoid anticancer quinone, cannabidiol hydroxyquinone (HU-331), is highly effective against tumor xenografts in nude mice. For evaluation of the antiangiogenic action of cannabinoid quinones, collagen-embedded rat aortic ring assay was used. The ability of cannabinoids to cause endothelial cell apoptosis was assayed by TUNEL staining and flow cytometry analysis. To examine the genes and pathways targeted by HU-331 in vascular endothelial cells, human cDNA microarrays and polymerase chain reaction were used. Immunostaining with anti-CD31 of tumors grown in nude mice served to indicate inhibition of tumor angiogenesis. HU-331 was found to be strongly antiangiogenic, significantly inhibiting angiogenesis at concentrations as low as 300 nM. HU-331 inhibited angiogenesis by directly inducing apoptosis of vascular endothelial cells without changing the expression of pro- and antiangiogenic cytokines and their receptors. A significant decrease in the total area occupied by vessels in HU-331-treated tumors was also observed. These data lead us to consider HU-331 to have high potential as a new antiangiogenic and anticancer drug.

HU-331, a novel cannabinoid-based anticancer topoisomerase II inhibitor

Anthracyclines, a large group of quinonoid compounds, are used to treat some forms of cancer. Although highly effective in cancer therapy, the mechanism of action of these compounds is not specific; they act on cancer and other cells by numerous mechanisms. A new anticancer quinone (HU-331) was synthesized from cannabidiol. It shows significant high efficacy against human cancer cell lines in vitro and against in vivo tumor grafts in nude mice. In this study, we investigated its mode of action and present evidence on its unique mechanism. HU-331 does not cause cancer cell cycle arrest, cell apoptosis, or caspase activation. HU-331–caused cell death of human cancer cell lines is not mediated by reactive oxygen intermediates/species, as exposure to HU-331 failed to elicit the generation of reactive oxygen species. HU-331 inhibits DNA topoisomerase II even at nanomolar concentrations but has only a slight nonsignificant effect on DNA topoisomerase I action. The cannabinoid quinone HU-331 is a highly specific inhibitor of topoisomerase II, compared with most known anticancer quinones. It might represent a new potent anticancer drug. [Mol Cancer Ther 2007;6(1):173–83]

A Cannabinoid Anticancer Quinone, HU-331, Is More Potent and Less Cardiotoxic Than Doxorubicin: A Comparative in Vivo Study

Several quinones have been found to be effective in the treatment of some forms of cancer; however, their cumulative heart toxicity limits their use. The cannabinoid quinone HU-331 [3S,4R-p-benzoquinone-3-hydroxy-2-p-mentha-(1,8)-dien-3-yl-5-pentyl] is highly effective against tumor xenografts in nude mice. We report now a comparison of the anticancer activity of HU-331 and its cardiotoxicity with those of doxorubicin in vivo. General toxicity was assayed in Sabra, nude and SCID-NOD mice. The anticancer activity in vivo was assessed by measurement of the tumors with an external caliper in HT-29 and Raji tumor-bearing mice and by weighing the excised tumors. Left ventricular function was evaluated with transthoracic echocardiography. Myelotoxicity was evaluated by blood cell count. Cardiac troponin T (cTnT) plasma levels were determined by immunoassay. HU-331 was found to be much less cardiotoxic than doxorubicin. The control and the HU-331-treated groups gained weight, whereas the doxorubicin-treated group lost weight during the study. In HT-29 colon carcinoma, the tumor weight in the HU-331-treated group was 54% smaller than in the control group and 30% smaller than in the doxorubicin-treated group. In Raji lymphoma, the tumor weight in the HU-331-treated group was 65% smaller than in the control group and 33% smaller than in the doxorubicin-treated group. In contrast to doxorubicin, HU-331 did not generate reactive oxygen species in mice hearts (measured by protein carbonylation levels and malondialdehyde levels). In vivo, HU-331 was more active and less toxic than doxorubicin and thus it has a high potential for development as a new anticancer drug.

Concise Review: Energy Metabolites: Key Mediators of the Epigenetic State of Pluripotency

Recent studies suggest that the metabolic network is an important part of the molecular circuitry that underlies pluripotency. Of the metabolic pathways that were implicated in the pluripotency balance, “energy” metabolism is particularly notable. Its mechanism of action on pluripotency‐regulating genes has been partially elucidated when three metabolites, namely acetate, S‐adenosylmethionine, and O‐linked β‐N‐acetylglucosamine were recently shown to link cytosolic signals to pluripotent gene expression. The cytosolic levels of these metabolites are the result of environmental perturbations, making them sensitive messengers, which are assumed to diffuse through the nuclear pores, being small molecules. Recent work also suggests that the modulation of the levels of these metabolites in pluripotent cells controls the balance between pluripotency and early commitment via epigenetic modifications. Here, we review recent studies that link metabolism and pluripotency via epigenetic modifications that occur through these three metabolites. Stem Cells 2015;33:2374–2380

CB2 cannabinoid receptor agonist enantiomers HU-433 and HU-308: An inverse relationship between binding affinity and biological potency

Significance The significance of the results reported is in two areas. (i) Because the cannabinoid receptor type 2 (CB2) agonists seem to be general protective agents, HU-433, a new specific CB2 agonist, may be of major therapeutic importance. (ii) Enantiomers usually have different activity profiles. We report now that HU-433 and its enantiomer HU-308 are both specific CB2 agonists, but whereas HU-433 is much more potent than HU-308 in the rescue of ovariectomy-induced bone loss and ear inflammation, its binding to the CB2 receptor (through which the activity of both enantiomers takes place) is substantially lower compared with HU-308. This situation questions the usefulness of universal radioligands for comparative binding studies. Activation of the CB2 receptor is apparently an endogenous protective mechanism. Thus, it restrains inflammation and protects the skeleton against age-related bone loss. However, the endogenous cannabinoids, as well as Δ9-tetrahydrocannabinol, the main plant psychoactive constituent, activate both cannabinoid receptors, CB1 and CB2. HU-308 was among the first synthetic, selective CB2 agonists. HU-308 is antiosteoporotic and antiinflammatory. Here we show that the HU-308 enantiomer, designated HU-433, is 3–4 orders of magnitude more potent in osteoblast proliferation and osteoclast differentiation culture systems, as well as in mouse models, for the rescue of ovariectomy-induced bone loss and ear inflammation. HU-433 retains the HU-308 specificity for CB2, as shown by its failure to bind to the CB1 cannabinoid receptor, and has no activity in CB2-deficient cells and animals. Surprisingly, the CB2 binding affinity of HU-433 in terms of [3H]CP55,940 displacement and its effect on [35S]GTPγS accumulation is substantially lower compared with HU-308. A molecular-modeling analysis suggests that HU-433 and -308 have two different binding conformations within CB2, with one of them possibly responsible for the affinity difference, involving [35S]GTPγS and cAMP synthesis. Hence, different ligands may have different orientations relative to the same binding site. This situation questions the usefulness of universal radioligands for comparative binding studies. Moreover, orientation-targeted ligands have promising potential for the pharmacological activation of distinct processes.

HU-444, a Novel, Potent Anti-Inflammatory, Nonpsychotropic Cannabinoid

Cannabidiol (CBD) is a component of cannabis, which does not cause the typical marijuana-type effects, but has a high potential for use in several therapeutic areas. In contrast to Δ9-tetrahydrocannabinol (Δ9-THC), it binds very weakly to the CB1 and CB2 cannabinoid receptors. It has potent activity in both in vitro and in vivo anti-inflammatory assays. Thus, it lowers the formation of tumor necrosis factor (TNF)-α, a proinflammatory cytokine, and was found to be an oral antiarthritic therapeutic in murine collagen-induced arthritis in vivo. However, in acidic media, it can cyclize to the psychoactive Δ9-THC. We report the synthesis of a novel CBD derivative, HU-444, which cannot be converted by acid cyclization into a Δ9-THC–like compound. In vitro HU-444 had anti-inflammatory activity (decrease of reactive oxygen intermediates and inhibition of TNF-α production by macrophages); in vivo it led to suppression of production of TNF-α and amelioration of liver damage as well as lowering of mouse collagen-induced arthritis. HU-444 did not cause Δ9-THC–like effects in mice. We believe that HU-444 represents a potential novel drug for rheumatoid arthritis and other inflammatory diseases.

Cannabidiol, a Major Non‐Psychotropic Cannabis Constituent Enhances Fracture Healing and Stimulates Lysyl Hydroxylase Activity in Osteoblasts

Cannabinoid ligands regulate bone mass, but skeletal effects of cannabis (marijuana and hashish) have not been reported. Bone fractures are highly prevalent, involving prolonged immobilization and discomfort. Here we report that the major non‐psychoactive cannabis constituent, cannabidiol (CBD), enhances the biomechanical properties of healing rat mid‐femoral fractures. The maximal load and work‐to‐failure, but not the stiffness, of femurs from rats given a mixture of CBD and Δ9‐tetrahydrocannabinol (THC) for 8 weeks were markedly increased by CBD. This effect is not shared by THC (the psychoactive component of cannabis), but THC potentiates the CBD stimulated work‐to‐failure at 6 weeks postfracture followed by attenuation of the CBD effect at 8 weeks. Using micro–computed tomography (μCT), the fracture callus size was transiently reduced by either CBD or THC 4 weeks after fracture but reached control level after 6 and 8 weeks. The callus material density was unaffected by CBD and/or THC. By contrast, CBD stimulated mRNA expression of Plod1 in primary osteoblast cultures, encoding an enzyme that catalyzes lysine hydroxylation, which is in turn involved in collagen crosslinking and stabilization. Using Fourier transform infrared (FTIR) spectroscopy we confirmed the increase in collagen crosslink ratio by CBD, which is likely to contribute to the improved biomechanical properties of the fracture callus. Taken together, these data show that CBD leads to improvement in fracture healing and demonstrate the critical mechanical role of collagen crosslinking enzymes.