1: J Natl Cancer Inst. 2008 Jan 2;100(1):59-69. Epub 2007 Dec 25.

Inhibition of cancer cell invasion by cannabinoids via increased expression of tissue inhibitor of matrix metalloproteinases-1.
Ramer R, Hinz B.
Institute of Toxicology and Pharmacology, University of Rostock, Schillingallee 70, Rostock D18057, Germany. BACKGROUND: Cannabinoids, in addition to having palliative benefits in cancer therapy, have been associated with anticarcinogenic effects. Although the antiproliferative activities of cannabinoids have been intensively investigated, little is known about their effects on tumor invasion. METHODS: Matrigel-coated and uncoated Boyden chambers were used to quantify invasiveness and migration, respectively, of human cervical cancer (HeLa) cells that had been treated with cannabinoids (the stable anandamide analog R(+)-methanandamide [MA] and the phytocannabinoid delta9-tetrahydrocannabinol [THC]) in the presence or absence of antagonists of the CB1 or CB2 cannabinoid receptors or of transient receptor potential vanilloid 1 (TRPV1) or inhibitors of p38 or p42/44 mitogen-activated protein kinase (MAPK) pathways. Reverse transcriptase-polymerase chain reaction (RT-PCR) and immunoblotting were used to assess the influence of cannabinoids on the expression of matrix metalloproteinases (MMPs) and endogenous tissue inhibitors of MMPs (TIMPs). The role of TIMP-1 in the anti-invasive action of cannabinoids was analyzed by transfecting HeLa, human cervical carcinoma (C33A), or human lung carcinoma cells (A549) cells with siRNA targeting TIMP-1. All statistical tests were two-sided. RESULTS: Without modifying migration, MA and THC caused a time- and concentration-dependent suppression of HeLa cell invasion through Matrigel that was accompanied by increased expression of TIMP-1. At the lowest concentrations tested, MA (0.1 microM) and THC (0.01 microM) led to a decrease in invasion (normalized to that observed with vehicle-treated cells) of 61.5% (95% CI = 38.7% to 84.3%, P < .001) and 68.1% (95% CI = 31.5% to 104.8%, P = .0039), respectively. The stimulation of TIMP-1 expression and suppression of cell invasion were reversed by pretreatment of cells with antagonists to CB1 or CB2 receptors, with inhibitors of MAPKs, or, in the case of MA, with an antagonist to TRPV1. Knockdown of cannabinoid-induced TIMP-1 expression by siRNA led to a reversal of the cannabinoid-elicited decrease in tumor cell invasiveness in HeLa, A549, and C33A cells. CONCLUSION: Increased expression of TIMP-1 mediates an anti-invasive effect of cannabinoids. Cannabinoids may therefore offer a therapeutic option in the treatment of highly invasive cancers.

2: Mol Neurobiol. 2007 Aug;36(1):60-7. Epub 2007 Jun 28.

Cannabinoids and gliomas.

Velasco G, Carracedo A, Blzquez C, Lorente M, Aguado T, Haro A, Snchez C, Galve-Roperh I, Guzmn M.
Department of Biochemistry and Molecular Biology I, School of Biology, Complutense University, 28040 Madrid, Spain. Cannabinoids, the active components of Cannabis sativa L., act in the body by mimicking endogenous substances--the endocannabinoids--that activate specific cell surface receptors. Cannabinoids exert various palliative effects in cancer patients. In addition, cannabinoids inhibit the growth of different types of tumor cells, including glioma cells, in laboratory animals. They do so by modulating key cell signaling pathways, mostly the endoplasmic reticulum stress response, thereby inducing antitumoral actions such as the apoptotic death of tumor cells and the inhibition of tumor angiogenesis. Of interest, cannabinoids seem to be selective antitumoral compounds, as they kill glioma cells, but not their nontransformed astroglial counterparts. On the basis of these preclinical findings, a pilot clinical study of

Delta(9)-tetrahydrocannabinol (THC) in patients with recurrent glioblastoma multiforme has been recently run. The good safety profile of THC, together with its possible growth-inhibiting action on tumor cells, justifies the setting up of future trials aimed at evaluating the potential antitumoral activity of cannabinoids.

3: Acta Oncol. 2007 Oct 12;:1-9 [Epub ahead of print]

Delta9-Tetrahydrocannabinol inhibits cell cycle progression downregulation of E2F1 in human glioblastoma multiforme cells.

by

Galanti G, Fisher T, Kventsel I, Shoham J, Gallily R, Mechoulam R, Lavie G, Amariglio N, Rechavi G, Toren A. The Mina and Everard Goodman Faculty of Life Science, Bar-Ilan University, Ramat-Gan, Israel. Background. The active components of Cannabis sativa L., Cannabinoids, traditionally used in the field of cancer for alleviation of pain, nausea, wasting and improvement of well-being have received renewed interest in recent years due to their diverse pharmacologic activities such as cell growth inhibition, anti-inflammatory activity and induction of tumor regression. Here we used several experimental approaches, which identified delta-9-tetrahydrocannabinol (Delta(9)-THC) as an essential mediator of cannabinoid antitumoral action. Methods and results. Administration of Delta(9)-THC to glioblastoma multiforme (GBM) cell lines results in a significant decrease in cell viability. Cell cycle analysis showed G(0/1) arrest and did not reveal occurrence of apoptosis in the absence of any sub-G(1) populations. Western blot analyses revealed a THC altered cellular content of proteins that regulate cell progression through the cell cycle. The cell content of E2F1 and Cyclin A, two proteins that promote cell cycle progression, were suppressed in both U251-MG and U87-MG human glioblastoma cell lines, whereas the level of p16(INK4A), a cell cycle inhibitor was upregulated. Transcription of thymidylate synthase (TS) mRNA, which is promoted by E2F1, also declined as evident by QRT-PCR. The decrease in E2F1 levels resulted from proteasome mediated degradation and was prevented by proteasome inhibitors. Conclusions. Delta(9)-THC is shown to significantly affect viability of GBM cells via a mechanism that appears to elicit G(1) arrest due to downregulation of E2F1 and Cyclin A. Hence, it is suggested that Delta(9)-THC and other cannabinoids be implemented in future clinical evaluation as a therapeutic modality for brain tumors.

4: Chem Biodivers. 2007 Aug;4(8):1729-43.

Cannabis, pain, and sleep: lessons from therapeutic clinical trials of Sativex, a cannabis-based medicine.
Russo EB, Guy GW, Robson PJ. GW Pharmaceuticals, erusso@gwpharm.com Porton Down Science Park, Salisbury, Wiltshire SP4OJQ, UK.

Cannabis sativa L. has been utilized for treatment of pain and sleep disorders since ancient times. This review examines modern studies on effects of Delta9-tetrahydrocannabinol (THC) and cannabidiol (CBD) on sleep. It goes on to report new information on the effects on sleep in the context of medical treatment of neuropathic pain and symptoms of multiple sclerosis, employing standardized oromucosal cannabis-based medicines containing primarily THC, CBD, or a 1 : 1 combination of the two (Sativex). Sleep-laboratory results indicate a mild activating effect of CBD, and slight residual sedation with THC-predominant extracts. Experience to date with Sativex in numerous Phase I-III studies in 2000 subjects with 1000 patient years of exposure demonstrate marked improvement in subjective sleep parameters in patients with a wide variety of pain conditions including multiple sclerosis, peripheral neuropathic pain, intractable cancer pain, and rheumatoid arthritis, with an acceptable adverse event

profile. No tolerance to the benefit of Sativex on pain or sleep, nor need for dosage increases have been noted in safety extension studies of up to four years, wherein 40-50% of subjects attained good or very good sleep quality, a key source of disability in chronic pain syndromes that may contribute to patients' quality of life.

5: Chem Biodivers. 2007 Aug;4(8):1664-77.

On the pharmacological properties of Delta9-tetrahydrocannabinol (THC).

Costa B. Department of Biotechnology and Bioscience, University of Milano-Bicocca, Piazza della Scienza 2, I20126 Milano. barbara.costa@unimib.it Cannabis is one of the first plants used as medicine, and the notion that it has potentially valuable therapeutic properties is a matter of current debate. The isolation of its main constituent, Delta9tetrahydrocannabinol (THC), and the discovery of the endocannabinoid system (cannabinoid receptors CB1 and CB2 and their endogenous ligands) made possible studies concerning the pharmacological activity of cannabinoids. This paper reviews some of the most-important findings in the field of THC pharmacology. Clinical trials, anecdotal reports, and experiments employing animal models strongly support the idea that THC and its derivatives exhibit a wide variety of therapeutic applications. However, the psychotropic effects observed in laboratory animals and the adverse reactions reported during human trials, as well as the risk of tolerance development and potential dependence, limit the application of THC in therapy. Nowadays, researchers focus on other therapeutic strategies by which the endocannabinoid system might be modulated to clinical advantage (inhibitor or activator of endocannabinoid biosynthesis, cellular uptake, or metabolism). However, emerging evidence highlights the beneficial effects of the whole cannabis extract over those observed with single components, indicating cannabis-based medicines as new perspective to revisit the pharmacology of this plant.

6: Oncogene. 2008 Jan 10;27(3):339-46. Epub 2007 Jul 9.

Delta9-Tetrahydrocannabinol inhibits epithelial growth factor-induced lung cancer cell migration in vitro as well as its growth and metastasis in vivo.
Preet A, Ganju RK, Groopman JE. Division of Experimental Medicine, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA. Delta(9)-Tetrahydrocannabinol (THC) is the primary cannabinoid of marijuana and has been shown to either potentiate or inhibit tumor growth, depending on the type of cancer and its pathogenesis. Little is known about the activity of cannabinoids like THC on epidermal growth factor receptor-overexpressing lung cancers, which are often highly aggressive and resistant to chemotherapy. In this study, we characterized the effects of THC on the EGF-induced growth and metastasis of human non-small cell lung cancer using the cell lines A549 and SW-1573 as in vitro models. We found that these cells express the cannabinoid receptors CB(1) and CB(2), known targets for THC action, and that THC inhibited EGF-induced growth, chemotaxis and chemoinvasion. Moreover, signaling studies indicated that THC may act by inhibiting the EGF-induced phosphorylation of ERK1/2, JNK1/2 and AKT. THC also induced the phosphorylation of focal adhesion kinase at tyrosine 397. Additionally, in in vivo studies in severe combined immunodeficient mice, there was significant inhibition of the subcutaneous tumor growth and lung metastasis of A549 cells in THC-treated animals as compared to vehicle-treated controls. Tumor samples from THC-treated animals revealed antiproliferative and antiangiogenic

effects of THC. Our study suggests that cannabinoids like THC should be explored as novel therapeutic molecules in controlling the growth and metastasis of certain lung cancers.

7: Int J Cancer. 2007 Nov 15;121(10):2172-80. Comment in:

Int J Cancer. 2008 Apr 15;122(8):1920-1.

The cannabinoid delta(9)-tetrahydrocannabinol inhibits RASMAPK and PI3K-AKT survival signalling and induces BADmediated apoptosis in colorectal cancer cells.
Greenhough A, Patsos HA, Williams AC, Paraskeva C. Department of Cellular and Molecular Medicine, Cancer Research UK, Colorectal Tumour Biology Group, School of Medical Sciences, University of Bristol, University Walk, Bristol, United Kingdom. Deregulation of cell survival pathways and resistance to apoptosis are widely accepted to be fundamental aspects of tumorigenesis. As in many tumours, the aberrant growth and survival of colorectal tumour cells is dependent upon a small number of highly activated signalling pathways, the inhibition of which elicits potent growth inhibitory or apoptotic responses in tumour cells. Accordingly, there is considerable interest in therapeutics that can modulate survival signalling pathways and target cancer cells for death. There is emerging evidence that cannabinoids, especially Delta(9)-tetrahydrocannabinol (THC), may represent novel anticancer agents, due to their ability to regulate signalling pathways critical for cell growth and survival. Here, we report that CB1 and CB2 cannabinoid receptors are expressed in human colorectal adenoma and carcinoma cells, and show for the first time that THC induces apoptosis in colorectal cancer cells. THC-induced apoptosis was rescued by pharmacological blockade of the CB1, but not CB2, cannabinoid receptor. Importantly, THC treatment resulted in CB1mediated inhibition of both RAS-MAPK/ERK and PI3K-AKT survival signalling cascades; two key cell survival pathways frequently deregulated in colorectal tumours. The inhibition of ERK and AKT activity by THC was accompanied by activation of the proapoptotic BCL-2 family member BAD. Reduction of BAD protein expression by RNA interference rescued colorectal cancer cells from THC-induced apoptosis. These data suggest an important role for CB1 receptors and BAD in the regulation of apoptosis in colorectal cancer cells. The use of THC, or selective targeting of the CB1 receptor, may represent a novel strategy for colorectal cancer therapy. (c) 2007 Wiley-Liss, Inc.

8: AIDS Care. 2007 Feb;19(2):295-301.

Marijuana as therapy for people living with HIV/AIDS: social and health aspects.
Fogarty A, Rawstorne P, Prestage G, Crawford J, Grierson J, Kippax S. National Centre in HIV Social Research, University of New South Wales, Sydney, Australia. a.fogarty@unsw.edu.au Therapeutic use of marijuana has emerged as an important issue for people living with cancer,

HIV/AIDS and multiple sclerosis. This paper examines therapeutic use of marijuana in the Positive Health cohort study, a longitudinal cohort study of men and women living with HIV/AIDS in NSW and Victoria, Australia. Factors that distinguish therapeutic use of marijuana from recreational use were assessed by comparisons on a range of social and health-related variables. The results show that among 408 participants, 59.8% reported some use of marijuana in the past six months. Of those participants (n=244), 55.7% reported recreational use only of marijuana and 44.3% report mixed use of marijuana for therapeutic and recreational purposes. Multivariate logistic regression analysis showed that participants who used marijuana for therapeutic purposes were significantly more likely than recreational-only users to have used other complementary or alternative therapies, experienced HIV/AIDS-related illness or other illnesses in the past 12 months, had higher CD4/T-cell counts, had lower incomes, be younger in age and less likely to have had a casual partner in the six months prior to interview. These results show that a substantial proportion of people living with HIV/AIDS (PLWHA) use marijuana for therapeutic purposes, despite considerable legal barriers, suggesting marijuana represents another option in their health management. Rather than solely using marijuana in response to illness, the experience of illness may influence a person's understanding of their marijuana use, so that they come to understand it as therapeutic. Further research might consider possible interactions between cannabinoids and antiretroviral treatments, potential use of oral THC and the difficulties faced by clinicians and PLWHA in discussing marijuana in the current legal context.

9: Cancer Res. 2006 Jul 1;66(13):6615-21.

Delta9-tetrahydrocannabinol inhibits cell cycle progression in human breast cancer cells through Cdc2 regulation.
Caffarel MM, Sarri D, Palacios J, Guzmn M, Snchez C. Department of Biochemistry and Molecular Biology I, School of Biology, Complutense University, 28040 Madrid, Spain. It has been proposed that cannabinoids are involved in the control of cell fate. Thus, these compounds can modulate proliferation, differentiation, and survival in different manners depending on the cell type and its physiopathologic context. However, little is known about the effect of cannabinoids on the cell cycle, the main process controlling cell fate. Here, we show that Delta(9)-tetrahydrocannabinol (THC), through activation of CB(2) cannabinoid receptors, reduces human breast cancer cell proliferation by blocking the progression of the cell cycle and by inducing apoptosis. In particular, THC arrests cells in G(2)-M via down-regulation of Cdc2, as suggested by the decreased sensitivity to THC acquired by Cdc2-overexpressing cells. Of interest, the proliferation pattern of normal human mammary epithelial cells was much less affected by THC. We also analyzed by real-time quantitative PCR the expression of CB(1) and CB(2) cannabinoid receptors in a series of human breast tumor and nontumor samples. We found a correlation between CB(2) expression and histologic grade of the tumors. There was also an association between CB(2) expression and other markers of prognostic and predictive value, such as estrogen receptor, progesterone receptor, and ERBB2/HER-2 oncogene. Importantly, no significant CB(2) expression was detected in nontumor breast tissue. Taken together, these data might set the bases for a cannabinoid therapy for the management of breast cancer.