Cannabis -vs- Myeloma

​​​​​​cannabis hematopoietic and immune system show high levels of the cannabinoid receptor CB2, the potential effect of cannabinoids on hematologic malignancies has been poorly determined. Here we have investigated their anti-tumor effect in multiple myeloma (MM). We demonstrate that cannabinoids induce a selective apoptosis in MM cell lines and in primary plasma cells of MM patients, while sparing normal cells from healthy donors, including hematopoietic stem cells. This effect was mediated by caspase activation, mainly caspase-2, and was partially prevented by a pan-caspase inhibitor. Their pro-apoptotic effect was correlated with an increased expression of Bax and Bak, a decrease of Bcl-xL and Mcl-1, a biphasic response of Akt/PKB and an increase in the levels of ceramide in MM cells. Inhibition of ceramide synthesis partially prevented apoptosis, indicating that these sphingolipids play a key role in the pro-apoptotic effect of cannabinoids in MM cells. Remarkably, blockage of the CB2 receptor also inhibited cannabinoid-induced apoptosis. Cannabinoid derivative WIN-55 enhanced the anti-myeloma activity of dexamethasone and melphalan overcoming resistance to melphalan in vitro. Finally, administration of cannabinoid WIN-55 to plasmacytoma-bearing mice significantly suppressed tumor growth in vivo. Together, our data suggest that cannabinoids may be considered as potential therapeutic agents in the treatment of MM. myeloma (MM) is a plasma cell (PC) malignancy characterised by the accumulation of a monoclonal PC population in the bone marrow (BM). Cannabidiol (CBD) is a non-psychoactive cannabinoid with antitumoural activities, and the transient receptor potential vanilloid type-2 (TRPV2) channel has been reported as a potential CBD receptor. TRPV2 activation by CBD decreases proliferation and increases susceptibility to drug-induced cell death in human cancer cells. However, no functional role has been ascribed to CBD and TRPV2 in MM. In this study, we identified the presence of heterogeneous CD138+TRPV2+ and CD138+TRPV2- PC populations in MM patients, whereas only the CD138+ TRPV2- population was present in RPMI8226 and U266 MM cell lines. Because bortezomib (BORT) is commonly used in MM treatment, we investigated the effects of CBD and BORT in CD138+TRPV2- MM cells and in MM cell lines transfected with TRPV2 (CD138+TRPV2+). These results showed that CBD by itself or in synergy with BORT strongly inhibited growth, arrested cell cycle progression and induced MM cells death by regulating the ERK, AKT and NF-κB pathways with major effects in TRPV2+ cells. These data provide a rationale for using CBD to increase the activity of proteasome inhibitors in MM. studies support the benefit of cannabinoids in cancer therapy, especially in terms of their effects in the induction of cell death, inhibition of proliferation and anti-metastatic activity noted in different human cancer in vitro and in vivo models [1, 2]. Cannabinoids are a family of compounds that exert their biological actions via a dependent-receptors mechanism, by binding mainly to Cannabinoid receptor type-1 and -2 (CB1, CB2) and Transient Potential Vanilloid type 1 and 2 (TRPV1, TRPV2) [3]. Moreover, receptors independent cannabinoids effects have also been described in cancer [1]. The most relevant effect of cannabinoids in cancers was investigated with Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD). THC and/or CBD were able to reduce cell proliferation and induce cell death in glioblastoma (GBM), lung and breast cancers, hepatocellular carcinoma and melanoma [4–10]. In addition, CBD has been shown to reduce viability, induce necrosis as well as synergize with bortezomib (BTZ) in reducing cell proliferation and cell survival pathways in multiple myeloma (MM) cell lines [11]. THC and CBD also show anti-inflammatory activities, by decreasing the release of pro-inflammatory cytokines (IFN-γ, IFN-β, IL-1 β, IL-6) and related transcription factors (such as NF-kB and STAT-3), in normal [12] and cancer cell lines, including MM [11]. Another important feature is that treatment with cannabinoids has been shown to reduce invasiveness of cancer cells as well as CXCR4-mediated migration of immune cells [13].

MM is a malignant disorder characterized by uncontrolled monoclonal plasma cell proliferation followed by the accumulation of malignant plasma cells in the bone marrow (BM), with possible escalation to anemia, osteolytic bone lesions, renal insufficiency, hypercalcemia and ultimately to extramedullary disease [14]. The prognosis of patients with MM has improved in the past decade, in respect of both progression-free survival (PFS) and overall survival (OS) [15], due to the introduction of a novel class of agents, such as immunomodulatory drugs (lenalidomide and pomalidomide) and proteasome inhibitors (BTZ and carfilzomib, CFZ) [16]. cannabinoids are postulated to have a potential anticancer effect working through the cb1 receptor, it would follow that the brain—where the cb1 receptor is the most densely populated seven-transmembrane domain G protein–coupled receptor—would be a good place to start the investigation. And, in fact, numerous studies in vitro and in animal models have suggested that cannabinoids can inhibit gliomas42. Other tumour cell lines are also inhibited by cannabinoids in vitro, and cannabinoid administration to nude mice curbs the growth of various tumour xenografts representing multiple solid and hematologic malignancies, including adenocarcinomas of the lung, breast, colon, and pancreas, and also myeloma, lymphoma, and melanoma. receptor-2 (CB2) is expressed dominantly in the immune system, especially on plasma cells. Cannabinergic ligands with CB2 selectivity emerge as a class of promising agents to treat CB2-expressing malignancies without psychotropic concerns. In this study, we found that CB2 but not CB1 was highly expressed in human multiple myeloma (MM) and primary CD138+ cells. A novel inverse agonist of CB2, phenylacetylamide but not CB1 inverse agonist SR141716, inhibited the proliferation of human MM cells (IC50 : 0.62 ∼ 2.5 μM) mediated by apoptosis induction, but exhibited minor cytotoxic effects on human normal mononuclear cells. CB2 gene silencing or pharmacological antagonism markedly attenuated phenylacetylamide's anti-MM effects. Phenylacetylamide triggered the expression of C/EBP homologous protein at the early treatment stage, followed by death receptor-5 upregulation, caspase activation, and β-actin/tubulin degradation. Cell cycle related protein cdc25C and mitotic regulator Aurora A kinase were inactivated by phenylacetylamide treatment, leading to an increase in the ratio inactive/active cdc2 kinase. As a result, phosphorylation of CDK substrates was decreased, and the MM cell mitotic division was largely blocked by treatment. Importantly, phenylacetylamide could overcome the chemoresistance of MM cells against dexamethasone or melphalan. Thus, targeting CB2 may represent an attractive approach to treat cancers of immune origin. against delta 1-THCA was produced by fusing hybridoma with splenocytes immunized with delta 1-THCA-BSA conjugate and hypoxanthine, aminopterine, thymidine-sensitive mouse myeloma cell line, P3-X63-Ag8-653. The cross-reaction of anti-delta 1-THCA antibody against other cannabinoids was very wide, thus many cannabinoids and a spiro-compound were reactive suggesting that 2'-hydroxyl, 6'-hydroxyl or 6'-O-alkyl, 4'-alkylbenzene ring moiety is necessary for its reactivity. It became evident that this ELISA was able to be applied to the biotransformation experiments of cannabinoids in plant tissue culture system. The metabolites of delta 6-THC such as two major metabolites, 7-oxo-delta 6-THC and 7-hydroxyl-delta 6-THC were also detectable by this ELISA. the course of a program in developing new ELISA-methods for the quantification of bioactive natural products in plants, phytomedicines and animals in a μg and ng scale, monoclonal antibodies against various natural products of medicinal and analytical importance have been developed. The ratio of hapten to bovine serum albumin (BSA) in an antigen conjugate was determined by matrix-assisted laser desorption/ionization (MALDI) of mass spectrometry. A hybridoma secreting monoclonal antibodies (MAb) was produced by fusing splenocytes immunized with an antigen-BSA conjugate with HAT-sensitive mouse myeloma cells. The cross-reaction of anti-forskolin antibodies with 7-deacetyl-forskolin was 5.6%. A very small cross-reaction appeared with other derivatives. The full measuring range of the assay extends from 5 ng to 5 μg/ml of forskolin. Immunoaffinity column chromatography using anti-forskolin MAbs appears to be far superior to previously published separation methods. The capacity of the immunoaffinity column as determined by ELISA is 9 μg/ml. Forskolin has been isolated directly from the crude extracts of tuberous roots and the callus culture of Coleus forskohlii. A MAb against Δ(1)-tetrahydrocannabinolic acid (THCA) was produced. The cross-reaction of anti-Δ(1)-THCA antibody against other cannabinoids was very wide. Many cannabinoids and a spiro-compound were reactive, but did not react with other phenolics. It became evident that this ELISA method was able to be applied to the biotransformation experiments of cannabinoids in plant tissue culture system. Anti-solamargine MAbs were produced. A method of determination for solasodine glycosides by using TLC-immunostaining was established. Solasodine glycosides separated by silica gel TLC were transferred to a polyvinylidene difluoride (PVDF) membrane. The membrane was treated with NaIO(4) solution followed by BSA, resulting in a solasodine glycoside-BSA conjugate. Immunostaining of solasodine glycosides was more sensitive compared to other staining. Immunostaining of ginsenosides in fresh ginseng root was successful using anti-ginsenoside Rb1 (G-Rb1) MAb after blotting to PVDF membrane.