The use of cannabinoid-containing plant extracts as herbal medicine can be traced back to the year 500 BC. Trace back to BC. In recent years, the medical and health-related uses of one of the non-psychotic cannabinoids, cannabidiol, or CBD, have received tremendous attention. In this review we will discuss the latest findings that greatly support the continued development of CBD as a promising anti-cancer agent.
A brief explanation of CBD
Recently, cannabinoids such as cannabidiol (CBD) and Δ9-tetrahydrocannabinol (THC) have been intensively researched and scrutinized. Cannabinoids encompass a wide range of organic molecules, including those physiologically produced in humans, synthesized in laboratories, and primarily derived from the cannabis sativa plant. These organic molecules are similar both in their chemical structure and in their protein binding profile. However, there are clear differences in their mechanisms of action and clinical applications, which are briefly compared and contrasted in this overview. The mechanism of action of CBD and its possible applications in cancer therapy are the focus of this review article.
The use of cannabis sativa plant extract as herbal medicine can be traced back to the year 500 BC in Asia. Chr. Lead back. The human endocannabinoid was discovered after the discovery of cannabinoid receptors. Originally it was assumed that cannabinoids develop their physiological effects via unspecific interactions with the cell membrane; However, research on rat models in the late 1980s led to the discovery and characterization of the specific cannabinoid receptors CB1 and CB2. The CB1 receptor is expressed throughout the central nervous system (CNS), while the CB2 receptor is mainly found in the immune system and in hematopoietic cells. Soon after the discovery of CB1 and CB2, their endogenous ligands, the endocannabinoids, were also identified, including 2-arachidonolyglycerol (2-AG) and N-arachidonoylethanolamine (AEA, also called anandamide). CB1 and CB2 belong to a large family of transmembrane proteins called G-protein-coupled receptors (GPCRs) and are now believed to be responsible for most of the physiological effects of endocannabinoids. Both receptors are coupled with Gαi / o, which can inhibit adenylyl cyclase (AC). CB1 can also be coupled to Gαq / 11 and Gα12 / 13. CB2 has also been shown to work via Gαs. For a deeper understanding of the downstream effects of endocannabinoids and their receptors under physiological conditions, we refer to other excellent reviews on this topic.
The two primary endocannabinoids, 2-AG and AEA, can activate either CB1 or CB2 and are synthesized from phospholipid precursors when needed in response to an increase in intracellular calcium. In addition to CB1 and CB2, 2-AG and AEA can also bind other transmembrane proteins, including the orphan G protein-coupled receptor 55 (GPR55), peroxisome proliferator activated receptors (PPAR) and the transient receptor potential vanilloid (TRPV) -Channel type 1 (TRPV1).
The TRPV channels are of particular interest with regard to the anti-tumor functions of cannabidiol (CBD), which will be discussed in more detail later. Six different TRPV channels have been identified in humans, which can be divided into two groups: TRPV1, TRPV2, TRPV3 and TRPV4 belong to Group I, while TRPV5 and TRPV6 belong to Group II. Although the exact functions of the TRPV channels are still under intense study, they are likely involved in the regulation of cellular calcium homeostasis. For example, TRPV1 and TRPV2 are found both in the cytoplasmic membrane and in the membrane of the endoplasmic reticulum (ER). Both play an important role in regulating the cytoplasmic calcium concentration from extracellular sources as well as the calcium stored in the ER. Disruption of cellular calcium homeostasis can lead to increased reactive oxygen species (ROS) production, ER stress, and cell death.
In the cannabis sativa plant (also called hemp or marijuana or CBD buds known) there are a variety of cannabinoids. There are more than 100 different cannabinoids, of which Δ9-tetrahydrocannabinol (Δ9-THC) and CBD are the most popular. The so-called cannabis sativa drug type contains higher levels of Δ9-THC and is more commonly used for medicinal and recreational purposes, while the fiber type cannabis contains less than 0,2% Δ9-THC and is more commonly used in textiles and food. Δ9-THC is considered the psychotic cannabinoid, and many of its psychoactive effects are due to its interaction with the CB1 receptor, while its immunomodulatory properties are likely due to its interaction with the CB2 receptor. In contrast, CBD is not psychoactive and has relatively low affinity for both CB1 and CB2.
The benefits of cannabinoids in treating cancer have long been of great interest. Recently, it was found that both CB1 and CB2 are expressed in many cancers. Interestingly, both receptors were often undetectable before the neoplastic transformation at the origin of the cancer. Further evidence of the role of the endocannabinoid system in neoplasms was provided by Wang and colleagues when they showed that CB1 has a tumor suppressive function in a genetically engineered mouse model of colon cancer. On the other hand, CB1 is upregulated in hepatocellular carcinoma and Hodgkin's lymphoma, and the degree of overexpression of CB1 correlated with the severity of the disease in epithelial ovarian carcinoma. Similarly, CB2 overexpression has also been found in HER2 + breast cancer and gliomas. Finally, it has been shown that overexpression of both CB1 and CB2 correlates with a poor prognosis for stage IV colorectal cancer. In 1976, Carchman and colleagues found that administration of cannabinoids such as Δ8-THC, Δ9-THC, and CBD inhibited DNA synthesis and lung adenocarcinoma growth in cultured cells and in mouse tumor models. Similar effects have been seen in in vitro and in vivo models of several other cancers, including gliomas, breast, pancreatic, prostate, colorectal cancers, and lymphomas. There are several proposed mechanisms of action behind these results including, but not limited to: cell cycle arrest, induction of apoptosis, and inhibition of neovascularization, migration, adhesion, invasion and metastasis. Despite the numerous positive results with Δ9-THC-related cannabinoids in cancer research, the clinical use of these compounds is limited due to their psychoactive side effects.
Unlike the Δ9-THC-related cannabinoids, CBD has no known psychoactive effects and has therefore recently been extensively researched in many therapeutic areas, including cancer. Currently, the Food and Drug Administration (FDA) has only approved Epidiolex, purified CBD, for use in patients with seizures related to Lennox-Gastaut syndrome or Dravet syndrome. Worldwide, more than 40 countries have approved medical marijuana / cannabis programs, while the US has approved 34 states, the District of Columbia, Guam, Puerto Rico and the US Virgin Islands. While marijuana is a Schedule I controlled substance in the US, the Drug Enforcement Administration has ruled that CBD is a Schedule V controlled substance. If CBD is approved by the FDA, it must contain less than 0,1% Δ9-THC.
It has been found that CBD has a relatively low affinity for both CB1 and CB2. However, it has been found that CBD can act as a CB1 antagonist in vitro in the vas deferens and in mouse brain tissue. There is also evidence that CBD can act as an inverse agonist of CB2 in humans. Other cellular receptors that CBD can interact with are TRPVs, 5-HT1A, GPR55, and PPARγ. It has been hypothesized that CBD has robust anti-proliferative and pro-apoptotic effects. In addition, it can also inhibit the migration, invasion, and metastasis of cancer cells. The benefits of CBD in anti-tumor therapy and the potential mechanisms behind it are discussed in more detail below. Since much of CBD's anti-tumor activity appears to depend on its regulation of ROS, ER stress, and immune modulation, we will first summarize the interactions between ROS, ER stress, and inflammation and their known effects on various aspects of tumorigenesis. Then we will discuss the anti-tumor effects of CBD on a variety of cancers and the molecular mechanisms behind them.
The interactions between reactive oxygen species (ROS), ER stress, inflammation and cancer
ROS and cancers
ROS denote various oxygen-containing species that are energetically unstable and highly reactive with a variety of biomolecules, including amino acids, lipids, and nucleic acids. Common ROS include superoxide (O2-), peroxide (O2-2), hydrogen peroxide (H2O2) and hydroxyl free radicals (OH-). The most common sources of ROS are the electron transport chain in the mitochondria and the NADPH oxidase (NOX) family of transmembrane enzymes. Certain enzymes and organelles, such as peroxisomes and ER, can also produce ROS. ROS can oxidize nucleic acids, proteins and lipids directly and thus change or disrupt their functions.
In order to prevent permanent damage to the biomolecules, the ROS are compensated by various antioxidants in the cells. Major antioxidant enzymes include superoxide dismutase (SOD), catalase, peroxiredoxin (PRX), thioredoxin (TRX), and glutathione peroxidase (GPX).
In cancer, the redox balance is so disturbed that increased ROS production favors the progression and spread of the tumor and at the same time escapes cell death. The tumor-promoting effects of increased ROS formation include genomic instability and increased proliferation. ROS damage DNA by oxidizing guanine and forming 8-hydroxyguanine and 8-nitroguanine. This can lead to deletions / insertions, mutations in the base pairing and strand breaks with subsequent mutagenic repair. Genome instability plays a key role in tumor progression through the accumulation of mutations that promote uncontrolled growth and circumvent cell death. Proliferation is further promoted by the oxidation and activation of growth-promoting intracellular signaling pathways, including mitogen-activated protein kinase (MAPK) signaling pathways and the phosphatidylinositol-3-kinase (PI3K) / protein kinase B (AKT) signaling pathway. The nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), a transcription factor important for growth and migration, is also activated by ROS by inhibiting the phosphorylation of the inhibitor of NF-κB α (IκBα) or the Promotes S-glutathionylation of the inhibitor of the NF-κB kinase subunit β (IKKβ). Eventually, cancer cells can rewire their signal transduction pathways to deal with increased intracellular ROS. This can mainly be achieved through increased mitochondrial SOD activity or the inactivation of the capture enzymes.
Nevertheless, toxic ROS concentrations can trigger cell death or autophagy in cancer cells. ROS modulate the activity of calcium channels, pumps and exchangers by oxidizing their Cys residues. The increase in intracellular mitochondrial calcium or oxidation of lipids damages the mitochondrial membrane, resulting in the release of cytochrome c, a powerful activator of apoptosomes. ROS can also directly influence caspase activity and cleavage of Bcl-2 and / or increase the expression of cell death receptors such as TRAIL and Fas. Autophagy can be triggered by activating the mTOR signaling pathway.
Endoplasmic reticulum (ER) stress and cancer
The ER is an important organelle that plays a critical role in post-translational modification and folding of proteins, calcium homeostasis, and other biological processes. The accumulation of unfolded and / or misfolded proteins triggers the Unfolded Protein Response (UPR), which helps rebalance ER homeostasis. The UPR temporarily stops protein synthesis and tries to correct and refold the proteins. If the unfolded and / or misfolded proteins cannot be corrected in good time, they are degraded in a targeted manner.
The UPR is a well-studied cellular process. It is mainly regulated by the 78 kDa glucose-regulated protein, also known as immunoglobulin binding protein (BiP). Under non-stress conditions, GRP78 binds and inhibits three transmembrane proteins: Inositol-requiring enzymes 1α (IRE1α), pancreatic endoplasmic reticulum kinase (PERK) and the activating transcription factor 6 (ATF6). Under ER stress conditions, GRP78 binds the unfolded proteins, dissociates from PERK, IRE1α and ATF6 and leads to the activation of three different but interconnected signaling pathways. CHOP activity is increased downstream of the PERK and ATF6 cascades.
CHOP induces apoptosis in several ways: it increases the transcription of GADD34; It increases the transcription of ER oxidoreductase 1 alpha (ERO1α), which then reoxidizes PDI and creates ROS; It increases transcription of the inositol 1,4,5-triphosphate receptor (IP3R), which then increases calcium levels in the cytoplasm; It activates the extrinsic cell death pathway via death receptor 5 (DR5) and the caspase-8 mediated activation of truncated Bid (tBid), which then migrates into the mitochondria and promotes the release of cytochrome c; It activates the intrinsic cell death pathway by directly reducing the expression of the survival factors Bcl-2 and Bcl-xL and increasing the expression of pro-apoptotic factors such as Bax, Bak, Bim, Puma and Noxa; It activates caspase-8 via TRAIL-DR5 on the cytoplasmic membrane, which cleaves the B-cell receptor-associated protein 31 (BAP31) and forms p20. p20 then releases calcium from the ER into the cytoplasm, which is taken up by the mitochondria and leads to the further release of cytochrome c.
During development, tumors rely heavily on the UPR signaling pathway for survival, possibly due to the hypoxic environment and metabolic stress associated with the rapidly growing tumor mass. For example, PERK and ATF4 activate vascular endothelial growth factor (VEGF) and hypoxia-inducible factor 1/2 (HIF1 / 2) for angiogenesis. Silencing the XBP1 gene prevented tumor growth and metastasis of triple negative breast cancer (TNBC) in vivo. Analyzes with TNBC cell lines showed that the upregulation of XBP1 increases HIF1α expression. However, when the URP system is overwhelmed, the pro-apoptotic factors dominate, resulting in cell death.
The effects of inflammation and microenvironment on survival, migration, and immune bypassing of tumors
The microenvironment of the tissue often plays an important role in the development, spread and metastasis of tumors. The tumor microenvironment consists primarily of infiltrated leukocytes, including tumor-associated macrophages (TAMs), dendritic cells, and myeloid-derived suppressor cells (MDSC). The interaction between the infiltrated cells and the tumor cells could suppress the immune response and create a survival-friendly environment for the tumor cells.
Avoiding the attack by the immune system is critical in the development of cancer. This is achieved through dynamic interactions between different cytokines and their receptors in the microenvironment of the tumor. Tumors actively secrete various cytokines that attract a variety of infiltrating cells such as TAMs, dendritic cells, MSDCs, and immunosuppressive regulatory T cells, which in turn help the tumors evade attack by the immune system. Cytokines released by myeloid cells can also induce genomic instability in tumor cells by directly damaging DNA or epigenetically altering the expression of genes.
The most important inflammatory mediators for tumor proliferation and survival include NF-κB and Signal Transducer and Activator of Transcription 3 (STAT3). IL-6, which is secreted by the myeloid cells, activates STAT3, which then upregulates the cyclins D1, D2 and B as well as MYC in order to promote the proliferation of the tumor cells. STAT3, which is expressed by the tumor cells, increases the secretion of IL-6 by the myeloid cells via an increased expression of NF-κB in these inflammatory cells, creating a positive feedback loop. IL-22, which is produced by the CD11c + lymphoid cells, is also able to activate STAT3 in epithelial cells. At the same time, the secretion of TNF-α and IL-1 by leukocytes can upregulate the expression of NF-κB in tumor cells. NF-κB in turn upregulates the expression of IL-1α, IL-1R and MYD88, which can further increase the activity of NF-κB, creating a positive autocrine loop. The expression of NF-κB can be activated in immune cells directly by the inflammatory cytokines TNF-α and IL-1 as well as by TLR-MYD88 in the event of tissue damage. It has been shown that NF-κB also triggers the epithelial-mesenchymal transition (EMT) as a result of IL-6 signaling, which then promotes the migration of tumor cells. In a prostate cancer model, the interaction between the NF-κB receptor activator (RANK) on the surface of the cancer cells and the RANK ligand on the infiltrating leukocytes promoted metastasis by activating the NF-κB signaling pathway. This positive feedback loop NF-κB / IL-6 / STAT3 is present in all phases of tumor development.
In addition, the expression of STAT3 in tumor-associated leukocytes also plays a key role in immune modulation. STAT3 expression in inflammatory cells enables immune bypassing tumors, while the deletion of STAT3 in macrophages and neutrophils enhances the Th1-mediated response with increased production of IFNγ, TNF-α and IL-1. STAT3 expression in myeloid cells can inhibit dendritic cell maturation by downregulating their IL-12 expression, and suppresses the immune response by upregulating the expression of IL-23 in TAMs.
Overall, the activation of the NF-κB and STAT3 signal transmission pathways in cancer cells as well as in the inflammatory cells in the tumor microenvironment offers a great advantage for the reproduction, survival, migration and bypassing of the immune system.
The anti-cancer effects of CBD
Glioma is the most common primary malignant disease of the brain. Grade IV glioma, also known as glioblastoma multiforme (GBM) or glioblastoma, is one of the most aggressive types of cancer. The prognosis for GBM is very poor, with only 4-5% surviving within five years. Current treatment modalities include surgery followed by radiation therapy and chemotherapy with temozolomide (TMZ) or carmustine (BCNU). Unfortunately, most GBM tumors are resistant to these treatments.
Because of the urgent medical need, cannabinoids have been extensively studied in glioma. Table S1 summarizes the published studies looking at the effects of CBD on gliomas, either alone or in combination with BCNU, TMZ, tamoxifen, cisplatin, γ-irradiation, ATM inhibitors and Δ9-THC. Many GBM cell lines were used in these studies, most of them U87MG. The antiproliferative effect of CBD on GBMs is quite clear, but the mean IC50 values of CBD differ between the different cell lines: C6 (8,5 µM), U87MG (12,75 ± 9,7 µM), U373 (21,6, 3,5 ± 33,2 µM), MZC (261 µM), and GL10,67 (0,58 ± XNUMX µM). The differences between the various publications may be due to procedural differences, including the assays used to measure viability and / or duration of CBD exposure.
It has been shown that CBD alone or in combination with other active ingredients successfully induces cell death, inhibits cell migration and invasion in vitro, reduces the size, vascularization, growth and weight of the tumor and increases the rate of survival and regression of the Initiates tumor in vivo. As for the antiproliferative effects of CBD on GBM, the data show that apoptosis occurs independently of CB1, CB2, and TRPV1, but is dependent on TRPV2. Ivanov et al. found in particular that CBD, γ-irradiation and the ATM inhibitor KU60019 upregulate TNF / TNFR1 and TRAIL / TRAIL-R2 signaling together with DR5 within the extrinsic apoptotic pathway. CBD also activates the JNK-AP1 and NF-κB signaling pathways to trigger cell death. Less emphasis was placed on the role of autophagy, or cell cycle arrest, in the CBD-mediated effects on glial cells.
Many downstream effects of CBD have been studied. Several studies reported increased levels of oxidative stress in GBM cell lines treated with CBD but not with Δ9-THC. Massi et al. found that the ROS concentration increases with time and is already significant after five hours when U87MG cells were treated with 25 µM CBD. At the same time, the level of glutathione, an antioxidant, was significantly reduced after six hours of CBD treatment. In contrast, normal glial cells treated with CBD did not show a pronounced increase in ROS. Treating CBD and antioxidants at the same time, including N-acetylcysteine (NAC) and α-tocopherol (i.e., vitamin E), weakened the killing effects of CBD. Overall, the studies on GBM cell lines suggest that CBD most likely triggers cell death by upregulating ROS. Scott et al. found that CBD also increased heat shock protein (HSP) expression, which has been linked to increased production of ROS as NAC hindered the role of HSP. Interestingly, it was shown that the use of HSP inhibitors together with CBD increases the cytotoxic effect and significantly reduces the IC50 value of CBD in T98G cells from 11 ± 2,7 µM to 4,8 ± 1,9 µM. This suggests that HSP inhibitors can be used as add-on therapy to CBD. Recently, Aparicio-Blanco et al. GBM in vitro CBD in lipid nanocapsules (LNCs) to find a formula with extended CBD release. The LNCs loaded with CBD were more effective at lowering IC50 values when they were smaller and exposed for a longer period of time.
In GBMs, CBD inhibits the PI3K / AKT pathway of survival by downregulating the phosphorylation of AKT1 / 2 (pAKT) and p42 / 44 MAPKs without affecting the total levels of AKT and p42 / 44 MAPK proteins. This signaling pathway could also be responsible for the CBD-mediated autophagy in stem-like glioma cells, since in these cells PTEN is upregulated while AKT is downregulated. The PI3K pathway plays an important role in the expression of TRPV2, which is a potential target for CBD. In U251, Δ9-THC and CBD together, but not individually, downregulated p42 / 44-MAPKs. Scott et al. however, showed that CBD alone in T98G and U87MG cells, albeit in a higher concentration (20 µM), lowered the pAKT and p42 / 44-MAPK levels, even more when combined with γ-irradiation became. CBD can also activate the pro-apoptotic MAP kinase pathway. Ivanov et al. found that CBD treatment together with γ-irradiation resulted in an upregulation of the active JNK1 / 2 and p38 MAPK, especially in U87MG cells. Marcu et al. however, showed using U251 cells that Δ9-THC and CBD did not increase the activity of JNK1 / 2 or p38 MAPK. The discrepancy could be due to the genetic differences between the different GBM cell lines.
Massi et al. investigated how CBD modulates 5-lipoxygenase (5-LOX), COX-2 and the endocannabinoid system in GBMs. They found that 5-LOX, but not COX-2, was decreased by CBD in vivo. CBD treatment also resulted in increased expression of fatty acid amide hydrolase (FAAH), which lowers AEA levels, suggesting that CBD may inhibit inflammatory mediator production by indirectly depressing the endocannabinoid system in GBMs.
In addition to γ-irradiation, CBD was also tested with alkylating agents, in particular TMZ, and it turned out that both together have a synergistic antiproliferative effect on glioma cells. Kosgodage et al. found that cells treated with CBD, both alone and with TMZ, increasingly form extracellular vesicles (EV) containing the anti-oncogenic miR-126. The levels of pro-oncogenic miR-21 and prohibitin, which are responsible for chemoresistant functions and mitochondrial protective properties, were also reduced.
In pre-clinical GBM mouse models, oral administration of a Sativex-like combination of Δ9-THC and CBD in a 1: 1 ratio with TMZ resulted in a decrease in tumor growth and an increase in survival time. These findings have led to two phase I / II clinical trials. Preliminary results are only available for one study and are promising. GBM patients were treated with either Sativex, CBD: Δ9-THC (1: 1), an oro-mucosal spray with high-dose TMZ, or placebo, and no Grade 3 or 4 toxicities occurred in the first part of the study. In the second part of the study, the same combination of active ingredients increased the median survival time compared to a placebo group with an increase in the one-year survival time of 83% and 56%, respectively. The most commonly reported adverse treatment effects were dizziness and nausea. Resistance to TMZ treatment can be reduced by using CBD: Δ9-THC combinations. When the full report is released, we hope the authors will discuss the safety and efficacy in more detail and help determine what adverse effects are due to Sativex compared to TMZ.
There are also some case studies that describe the use of CBD in patients with high-grade gliomas. Two patients were treated with procarbazine, lomustine, and vincristine along with CBD for about two years (one patient at 100-200 mg / day orally and the other at 300-450 mg / day orally). Neither patient had disease progression for two years after treatment. Treatment adverse effects included rash, moderate nausea, vomiting, and fatigue, but not lymphopenia, thrombocytopenia, hepatic toxicity, or neurotoxicity. In a case series in which nine patients with GBM grade IV were described, the mean survival with the combination of surgery, radio and chemotherapy, and CBD (200-400 mg / day) was increased to 22,3 months, and two patients had no evidence of disease progression for three or more years.
Overall, the published results suggest that CBD alone or in combination with Δ9-THC, TMZ, or γ-irradiation shows great promise for treating gliomas. Additionally, the unwanted effects of CBD alone or in combination with Δ9-THC appear to be relatively harmless.
Breast cancer is the leading cause of new cancer cases and the second leading cause of cancer deaths in women in the United States. The effects of CBD on breast cancer have been studied since 2006; research in this area has recently expanded. Different breast cancer cell lines have been used to show a dose-dependent response to CBD, including estrogen receptor (ER) -positive cells (MCF-7, ZR-75-1, T47D), ER-negative cells (MDA-MB-231, MDA-MB -468 and SK-BR3) and triple negative breast cancer cells (TNBC) (SUM159, 4T1up, MVT-1 and SCP2). Already 1 to 5 µM CBD caused significant cell death in MDA-MB-231 after 24 hours. The IC50 values of CBD for most cell lines are consistently low, suggesting that breast cancer cell lines are generally sensitive to the anti-proliferation effects of CBD without having a significant effect on untransformed breast epithelial cells.
CBD exerts its anti-proliferation effects on breast cancer cells through a number of mechanisms, including apoptosis, autophagy, and cell cycle arrest. Ligresti et al. reported that CBD-treated MDA-MB-231 cells induced an apoptotic effect involving caspase-3, while CBD exerted its effect on MCF-7 through cell cycle arrest at the G1 / S checkpoint. However, the cell cycle arrest at the G1 / S checkpoint was only recently detected in MDA-MB-231 and 4T1 cells after CBD treatment. Activation of the CB231 and TRPV2 receptors was found in MDA-MB-1 cells, but the effect was only partial. Recent studies have shown that the anti-proliferation effects of CBD on breast cancer cells are independent of the endocannabinoid receptors. CBD has been shown to produce ROS, which in turn inhibit proliferation and initiate cell death. CBD exerts its pro-apoptotic effects by down-regulating mTOR, AKT, 4EBP1 and cyclin D, while up-regulating PPARγ expression and its nuclear localization. Shrivastava et al. showed that the inhibition of the AKT / mTOR signaling pathway and the induction of ER stress induce not only apoptosis but also autophagy. At higher CBD concentrations or when autophagy was inhibited, apoptosis levels increased. They also showed that CBD can coordinate apoptosis and autophagy through the translocation and cleavage of Beclin-1.
CBD has also been shown to inhibit migration, invasion and metastasis in aggressive breast cancer in vivo and in vitro. McAllister et al. observed downregulation of Id-1 protein by ERK and ROS in CBD-treated MDA-MB-231 and MDA-MB-436 tumors. This down-regulation correlated with a decrease in tumor invasion and metastasis. Id-1 expression was also downregulated in metastatic lung nodules. Consistent with these observations, CBD failed to inhibit lung metastasis in Id-1 overexpressing breast cancer cells. Interestingly, the same study showed that CBD at a lower concentration (1,5 µM) that produced ROS and inhibited the expression of Id-1 in MDA-MB-231 cells did not induce autophagy or apoptosis. Recently, CBD was shown to inhibit TNBC cell proliferation, migration, and invasion by suppressing activation of the EGF / EGFR signaling pathway and its downstream targets (AKT and NF-κB). MMP, phalloidin, and actin stress fibers are important for tumor invasion and have also been suppressed by CBD. These results, which relate to the EGF / EGFR signaling pathway and the MMP, phalloidin and actin stress fibers, were also confirmed in vivo. The size of the primary tumor has been shown to decrease along with the number of lung metastases, volume, and vascularization in CBD-treated mice. Interestingly, the number of metastases and the survival time of the mice decreased when CBD was administered three times a week instead of daily, as McAllister et al. did, but the primary tumor was not reduced. The decreased angiogenesis and invasion were found to be due to a change in the microenvironment of the tumor, e.g. B. on a clear decrease in CCL3, GM-CSF and MIP-2, which led to an inhibition of the recruitment of TAMs. Finally, in another study, a synthetic cannabinoid analog, O-1663, was found to be more potent than CBD and Δ9-THC, similarly inducing cell death and autophagy. O-1663 also inhibited the aggressiveness of breast cancer in vitro and in vivo. It significantly increased the survival time of advanced breast cancer metastases, inhibited the formation of metastatic foci ≥2 mm and induced the regression of established metastatic foci, all without pronounced toxicity. Overall, the evidence suggests that there are several mechanisms by which CBD inhibits tumor migration.
Kosgodage et al. showed that breast cancer cells treated with CBD showed an increased sensitization to cisplatin. CBD significantly decreased the release of exosomes and microvesicles (EMV) (100-200 nm), which typically promote the spread of tumors and cause chemoresistance. However, an increase in the release of the larger EMV (231-201 nm) was observed in the same MDA-MB-500 cells. These cells showed a concentration-dependent increase in ROS, proton leakage, mitochondrial respiration and ATP levels. The authors attributed these effects either to a higher sensitivity or to a pseudo-apoptotic response in which the apoptotic factors such as ROS are still at a lower level, which leads to the conversion of apoptosomes into EMVs. CBD inhibited paclitaxel-induced neurotoxicity via a 5-HT1A receptor system with no conditioned reward or cognitive impairment. It also decreased the viability of 4T1 and MDA-MB-231 cells. Thus, CBD could be a useful add-on treatment for breast cancer as it can sensitize cells so that potentially lower doses of such toxic chemicals can be prescribed.
Overall, CBD has been shown to be effective in many breast cancer cells and mouse models when it comes to its anti-proliferation and pro-apoptotic effects, although the mechanisms of these effects can vary. At this point in time, there is an urgent need for clinical trials examining the anti-tumor effects of CBD in breast cancer as this appears to be the next logical step in the development of CBD as an alternative treatment for breast cancer.
According to epidemiological studies by the American Cancer Society, lung cancer is the second most common cancer in both men and women. Lung cancer is divided into small cell lung cancer (SCLC, 13%) and non-small cell lung cancer (NSCLC, 84%), which in turn can be divided into adenocarcinoma, squamous cell carcinoma, and large cell carcinoma.
Ramer and colleagues have published numerous studies on the effects of CBD on lung cancer. They all used the WST-1 assay to assess lung cancer viability. CBD decreased the viability of two NSCLC cell lines, A549 (a lung adenocarcinoma cell line) and H460 (a large cell lung cancer cell line), with IC50 values of 3,47 µM and 2,80 µM, respectively. The invasion of A549 was reduced by 72% and 0,001% after incubation for 0,1 hours with 29 µM and 63 µM CBD, respectively. No significant cell death was found in A549 cells after treatment with 0,001 µM or 0,1 µM CBD. Various lung cancer cell lines (e.g. A549, H358 and H460) have been shown to express CB1, CB2 and TRPV1, on which the anti-invasive function of CBD is partly based. CBD also significantly reduced tumor size and metastatic nodules in the lung (from an average of 6 nodules to just 1 nodule) in an A549 xenograft tumor model.
One mechanism for the pro-apoptotic effect of CBD is the activation of COX-2, a signaling pathway for the breakdown of endocannabinoids, and PPAR-γ. CBD treatment with 3 µM in A549, H460 and primary lung tumor cells of a patient with brain metastases led to an upregulation of COX-2 and PPAR-γ in both the mRNA and the protein. These observations were also confirmed in vivo. Products derived from COX-2 (PGE2, PGD2 and 15d-PGJ2) were also increased in lung cancer cells treated with CBD. By suppressing the COX-2 and PPAR-γ activity with antagonists or siRNA, the pro-apoptotic and cytotoxic effects of CBD were greatly attenuated. In a lung tumor mouse model, the PPAR-γ inhibition by GW9662 abolished the tumor suppressive effect of CBD.
Ramer et al. discussed the pro- and anti-tumor effects, respectively, of plasminogen activator inhibitor-1 (PAI-1), but provided evidence for the former. With 1 µM CBD there was a decrease in PAI-1 mRNA and the protein in A549, H358 and H460. This was confirmed in vivo in the A549 mouse model with 5 mg / kg CBD three times a week. In vitro, the anti-invasive property of CBD was reduced by siRNA knockdown of PAI-1 and increased by treatment with recombinant PAI-1. The CBD-mediated decrease in PAI-1 is due in part to activation of CB1, CB2, and TRPV1 as their antagonists cancel the effects. Therefore, in lung cancer, CBD acts as an agonist of CB1, CB2, and TRPV1.
The TIMPs (Tissue Inhibitor of MMPs) have been investigated and are related to the anti-invasive effects of CBD. It has been found that they are induced by CBD in a time and concentration dependent manner. The CBD-mediated upregulation of TIMP-1 was attributed to the activation of CB1, CB2 and TRPV1. CBD also activated p38 MAPK and p42 / 44 MAPK, two downstream targets of TRPV1. To link CB1, CB2 and TRPV1 to the activation of MAPK and TIMP-1, Ramer et al. the expression and function of the intercellular adhesion molecule-1 (ICAM-1), a transmembrane glycoprotein that is involved in tumor metastasis. A time- and concentration-dependent increase in ICAM-1 was observed in A549, H358 and H460 cells treated with CBD and in cells from a patient with brain metastatic NSCLC. An increase in the expression of TIMP-1 mRNA was also observed, but it occurred after the increase in ICAM-1 mRNA. The expression of ICAM-1 was dependent on the activation of p42 / 44 MAPK and p38 MAPK. In the in vivo model A549, which shows the anti-invasive properties of CBD, both ICAM-1 and TIMP-1 were upregulated. Inactivating ICAM-1 with a neutralizing antibody and siRNA led to a decrease in TIMP-1 activation as well as a decrease in the anti-invasive properties of CBD. These data suggest that the MAPKs activate ICAM-1, which then stimulates the function of TIMP-1, which in turn suppresses tumor invasion.
In a separate study, Haustein et al. CBD-induced ICAM-1 expression on lymphokine-activated kill-cell-mediated cytotoxicity (LAK). Treatment with 3 µM CBD induced ICAM-1 expression and LAK cell-mediated lysis of tumor cells in A549 and H460 as well as in metastatic cells of a patient with NSCLC. The increased susceptibility to adhesion and lysis by LAK in CBD-treated cells was abolished by a neutralizing ICAM-1 antibody. This effect of cell lysis was neutralized by using ICAM-1 siRNA together with CB1, CB2 and TRPV1 antagonists. The lymphocyte function association antigen (LFA-1) reversed the CBD-induced killing effect on LAK cells, suggesting that it acts as a counter-receptor to ICAM-1. Finally, CBD did not induce LAK cell-mediated lysis and upregulation of ICAM-1 in non-tumorous bronchial epithelial cells, suggesting that this effect is specific for cancer cells.
Taken together, these studies suggest that CBD activates p1 MAPK and p2 / 1 MAPK via CB38, CB42 and TRPV44 receptors, which induce ICAM-1 first and then TIMP-1. The upregulation of ICAM-1 and TIMP-1 then attenuates the invasion of lung cancer.
There are currently no published results from a clinical study using CBD to treat lung cancer patients. In a recent case report, however, an 81-year-old patient attempted his pulmonary adenocarcinoma CBD Oil treat yourself. Upon initial diagnosis of a 2,5 × 2,5 cm mass and several mediastinal masses, the patient was refused chemotherapy and radiation therapy due to his age and the toxicity profile of these treatments. However, a year later, computed tomography (CT) showed that the tumor and mediastinal lymph nodes were beginning to regress. During this time, the intake of 2% CBD oil was changed in particular. The undesirable effects included mild nausea and an unpleasant taste.
In the United States, colorectal cancer (CRC) is the third leading cause of cancer death in both men and women. Studies with two colon cancer cell lines, Caco-2 and DLD-1, and with healthy and cancerous tissue from nine colon cancer patients suggest that endocannabinoid production is markedly increased in precancerous adenomatous polyps and, to a lesser extent, in cancerous colon tissue. Normal human colon tissue expresses both CB1 and CB2 as well as AEA, 2-AG and endocannabinoid metabolizing enzymes such as FAAH. Transformed adenomatous polyps have increased levels of 2-AG compared to normal colorectal tissue. While DLD-1 cells express both CB1 and CB2, Caco-2 cells express only CB1. Depending on the stage of the cancer, endocannabinoids can either inhibit or promote the growth of colon cancer. Therefore, depending on the stage of cancer, both activators and inhibitors of the endocannabinoid system can be useful in fighting colon cancer.
The effects of CBD on CRC are summarized in Table S4. The dose-dependent killing of CRC cells by CBD has been demonstrated in many studies, with the IC50 values of SW480 between 5,95 µM and 16,5 µM over a period of 48 hours. This dose-dependent killing response is specific for CRC cells and not for normal human colorectal cells. The IC50 value for CaCo-2 was given as 7,5 ± 1,3 µM. Under the physiological oxygen conditions in the large intestine estimated at around 5%, Caco-2s were even more sensitive to CBD and showed a decrease in proliferation at 0,5 µM compared to 1 µM under atmospheric oxygen (~ 20%). In the same study, it was found that the anti-proliferation effects of CBD under physiological oxygen conditions are likely due to its ability to induce mitochondrial ROS. Apoptosis has been described as the primary route of cell death by CBD in CRC.
Sreevalsan, et al. used SW480 cells with 15 µM CBD to show that apoptosis is phosphatase and endocannabinoid dependent. After 24 hours, CBD induced the expression of various dual-specific phosphatases and protein tyrosine phosphatases, including DUSP1, DUSP10, serum ACPP, cellular ACPP, and PTPN6. Consistent with the hypothesis, the use of a phosphatase inhibitor, sodium orthovanadate (SOV), reduced apoptosis. Switching off CB1 and CB2 also inhibited apoptosis. Taken together, these studies suggest that the apoptotic effects of CBD in CRC are via the endocannabinoid system and activation of its downstream targets, including various phosphatases.
CBD has been shown to induce noxa-mediated apoptosis by generating ROS and excessive ER stress. In HCT116 and DLD-1 cells, CBD treatment induced an overproduction of ROS, particularly mitochondrial superoxide anions, which has been linked to the activation of noxa. Jeong et al. also found that noxa-activated apoptosis is dependent on excessive ER stress from ATF3 and ATF4. These proteins bind the Noxa promoter and stimulate its expression. In vivo, CBD-treated CRC tumors also led to a significant decrease in tumor size and to induction of apoptosis by Noxa.
Using HCT115 and Caco-2 cells, Aviello et al. found that 10 µM CBD has anti-proliferation effects through several mechanisms. CBD could work by indirectly activating receptors by increasing endocannabinoids, especially 2-AG, in CRC cell lines. In vivo, CBD at a dose of 1 mg / kg significantly reduced azoxymethane-induced aberrant crypt foci, polyps, tumors and the percentage of mice with polyps. The anti-tumor mechanism of CBD was determined by the down-regulation of the PI3K / AKT signaling pathway and the up-regulation of caspase-3.
In some studies, CBD was also examined as an additive to chemotherapy for CRC. CRC is often treated surgically in conjunction with the combination of 5-fluorouracil, leucovorin, and oxaliplatin (FOLFOX). In an effort to overcome potential resistance to FOLFOX, Jeong et al. oxaliplatin-resistant DLD-1 and colo205 cells with oxaliplatin and CBD (4 µM) and found that CBD played a role in oxaliplatin-mediated autophagy through decreased phosphorylation of NOS3, which is involved in the production of nitric oxide (NO) which plays oxaliplatin resistance. The combination of oxaliplatin and CBD caused mitochondrial dysfunction (decreased oxygen consumption rate, mitochondrial membrane potential, mitochondrial complex I activity and the number of mitochondria) through decreased SOD2 expression. These results were also confirmed in vivo.
An alternative targeted therapy for colorectal cancer, the TNF-related apoptosis-inducing ligand (TRAIL), has also shown resistance that can be overcome by the addition of CBD (4 µM) in HCT116, HT29 and DLD-1 cells . CBD and TRAIL increased apoptosis by activating genes related to ER stress, including PERK, CHOP, and DR5. In vivo, TRAIL with CBD showed a significant decrease in tumor growth and an increased number of apoptotic cells. Overall, these studies on FOLFOX and TRAIL therapy suggest that CBD could be considered as a therapeutic option for CRC, or perhaps as an adjunct treatment that works synergistically with conventional chemotherapy. There are currently no clinical studies on CBD in CRC, but these results are very promising in terms of the synergistic effects of CBD with chemotherapy and speak for a future clinical study.
Leukemia / lymphoma
Our knowledge of the effects of CBD on leukemia and lymphoma has expanded in recent years. EL-4 and Jurkat cell lines are the most common models for lymphoma and leukemia, respectively. CBD induced dose- and time-dependent killing effects on these leukemia and lymphoma cell lines, while peripheral blood monomolecular cells were more resistant to CBD.
McKallip et al. found that the anti-proliferation effects of CBD in both EL-4 and Jurkat cells are mediated by CB2, but are independent of CB1 and TRPV1. In another study, Olivas-Aguirre et al. however, that the effect of CBD is independent of the endocannabinoid receptors and the Ca2 + channels of the plasma membrane in Jurkat cells. These conflicting results need to be clarified through future studies. Nonetheless, the majority of research on leukemia / lymphoma has confirmed apoptosis as the mechanism by which CBD-mediated cell death occurs, either alone or in combination with other treatment modalities, including γ-irradiation, Δ9-THC, vincristine, and cytarbine. One study also showed that CBD reduced tumor burden and triggered apoptosis in vivo. Calendaroglou et al. found that CBD can induce a cell cycle arrest in Jurkat cells, with the number of cells increasing in the G1 phase. CBD treatment also resulted in changes in cell morphology, including decreased cell size, extensive vacuolization, swollen mitochondria, disassembled ER and Golgi, and a distended plasma membrane.
Similar to other cancers, CBD also induced ROS in leukemia and lymphoma. Treatment of Jurkat and MOLT-4, another leukemia cell line, with ≥2,5 µM CBD for 24 hours induced increased ROS values. Treating the cells with the ROS scavengers α-tocopherol and NAC reduced the killing effects of CBD. CBD exposure also increased NOX4 and p22phox, while inhibition of NOX4 and p22phox decreased ROS levels and inhibited CBD-induced cell toxicity. In agreement with these observations, the ROS concentrations in EL-4 cells were already significantly increased after two hours of CBD treatment, which was accompanied by a simultaneous decrease in cellular thiols.
Calendaroglou et al. examined the effects of CBD on the mTOR signaling pathway in Jurkat cells. They found that CBD reduced the phosphorylation of AKT and the ribosomal protein S6. They also tested the effects of CBD under various nutrient and oxygen conditions and found that the antiproliferative effects of CBD alone or with doxorubicin were greater with 1% serum than with 5% serum. Olivas-Aguirre, et al. found that when Jurkat cells were treated with lower CBD concentrations, proliferation still took place (at 1 µM CBD) and autophagy was increased at 10 µM CBD. At higher concentrations (30 µM), however, the intrinsic apoptotic pathway was activated, which led to the release of cytochrome c and a Ca2 + overload of the mitochondria. In the Burkitt lymphoma cell lines Jiyoye and Mutu I, AF1q stimulated cell proliferation and reduced ICAM-1 expression, making the cells resistant to chemotherapy. After 24 hours of exposure to CBD, the chemoresistant effects were drastically weakened.
Prostate cancer is the most common type of cancer and the second leading cause of cancer-related death in men. A detailed summary of the studies describing the effects of CBD on prostate cancer can be found in Table S6. The prostate cancer cell lines used in these studies can be divided into androgen receptor (AR) positive (LNCaP and 22RV1) and AR negative (DU-145 and PC-3). CBD can inhibit the expression of the androgen receptor in AR-positive cell lines. As for the endocannabinoid receptors, depending on the cancer cell type, either CB1 or CB2 or both are upregulated in prostate cancer cells compared to normal prostate cells. Specifically, 22RV1 only expresses CB1, while DU-145 expresses only CB2. Although CB1 and CB2 are found in both LNCaP and PC-3, they are much more pronounced in PC-3. TRPV1 is expressed in all four prostate cancer cell lines, with the highest expression being found in DU-145 cells.
CBD induced an antiproliferative effect and apoptosis-mediated cell death (via the intrinsic pathway) in prostate cancer cells, possibly caused by CB2, but not by CB1, and the receptor for the transient receptor potential cation channel of subfamily M, member 8 (TRPM8 ), in LNCaP cells. In addition, treatment with CBD has been shown to down-regulate the expression of prostate specific antigen (PSA), vascular endothelial growth factor (VEGF) and proinflammatory cytokines. In LNCaP and PC3 cells, the CBD treatment led to an arrest of the cell cycle at the G0 / G1 transition and in DU-145 cells at the G1 / S transition.
Similar to the CRCs, Sreevalsan et al. found that dual-specific phosphatases and protein tyrosine phosphatases were also induced by CBD in LNCaP cells. The inhibition of phosphatases with the phosphatase inhibitor SOV reduced PARP cleavage. In addition, CBD increased the phosphorylation of p38 MAPK. Recently, Kosgodage et al. found that treatment with CBD (1 µM and 5 µM) in PC3 reduced the release of EMV. CBD has also been shown to reduce mitochondria-associated proteins, prohibitin, and STAT3, which may be responsible for the decrease in EMV.
So far, only one study on the effectiveness of CBD in prostate cancer has been carried out in vivo. Before the clinical trial phase can begin, higher quality studies with mouse models are required.
The effects of CBD on a number of other cancers have also been reported, but to a lesser extent. Cervical cancer cell lines treated with CBD exhibited time- and concentration-dependent killing effects that have been shown to be mediated by apoptosis and independent of cell cycle arrest. Treatment with CBD resulted in an upregulation of p53 and Bax, a pro-apoptotic protein, and a downregulation of RBBP6 and Bcl-2, two anti-apoptotic proteins, in SiHa, HeLa and ME-180 cells. CBD also decreased the invasion of HeLa and C33A, which was dependent on CB1, CB2 and TRPV1. Ramer et al. also found that this anti-invasive property of CBD was related to the upregulation of p38 MAPK and p42 / 44 MAPK and their downstream target, TIMP-1, which is similar to lung cancer as described above.
CBD (1 µM and 5 µM) also decreased the cell viability of a hepatocellular carcinoma cell line, Hep G2, in a dose-dependent manner after 24 hours. Similar to the breast and prostate cell lines MDA-MB-231 and PC3, respectively, treatment of Hep G2 cells with CBD reduced the release of EMV and the expression of CD63, prohibitin and STAT3. In addition, treatment of Hep G2 cells with CBD sensitized them to cisplatin. Neumann-Raizel et al. used the mouse hepatocellular carcinoma cell line, BNL1 ME, expressing functional TRPV2 channels to demonstrate the effects of CBD in conjunction with doxorubicin. It has been shown that CBD (10 µM) activates TRPV2 and inhibits the P-glycoprotein ATPase transporter, which means that doxorubicin can increasingly penetrate the cell and accumulate there, as it is transported via TRPV2 through the cytoplasmic membrane and with the help of the P-glycoprotein -ATPase transporter is pumped out of the cell. These effects were likely responsible for CBD's ability to reduce the dose of doxorubicin required to reduce cell viability and proliferation.
In thyroid cancer, CBD in KiMol induced a proliferation-inhibiting effect by activating apoptosis and cell cycle arrest. KiMol was shown to contain increased levels of CB1, CB2, and TRPV1, but inhibitors of CB1, CB2, and TRPV1 only marginally decreased the antiproliferative effects of CBD. CBD (5 mg / kg twice a week) also had anti-tumor effects in a mouse thyroid tumor model.
Taha et al. studied patients with stage IV non-small cell lung cancer, clear cell renal cell carcinoma, and advanced melanoma who were treated with nivolumab immunotherapy (anti-PD-1 agent), and patients who had also consumed cannabis, including CBD and Δ9-THC . They showed a lower rate of response to treatment in the groups who consumed cannabis with nivolumab, while patients who did not use cannabis were 3,17 times more likely to respond to treatment with nivolumab. However, cannabis use did not result in a significant difference in overall survival and progression-free survival. This group suggested that there may be a negative interaction between cannabis and immunotherapy.
CBD decreased cell proliferation and colony formation in gastric cancer cells in a concentration-dependent manner without affecting normal gastric cells. The gastric adenocarcinoma cell line, AGS, has abundant expression of TRPV1 with no evidence of CB1 or CB2. Zhang et al. found that CBD caused cell cycle arrest in SGC-7901, another gastric cancer cell line, by inhibiting the expression of CDK2 and cyclin E. In addition, CBD increased the expression of ATM and p21 while decreasing that of p53. The antiproliferative effects of CBD in SGC-7901 have also been attributed to mitochondrial dependent apoptosis, as it increased the activity of caspase-3 and caspase-9, the release of cytochrome c and the expression of the proteins Apaf-1, Bad and Bax, and the expression of Bcl-2 decreased. The CBD-induced cell cycle arrest and apoptosis were associated with increased ROS levels. Jeong et al. showed in several gastric cancer cell lines that CBD triggers apoptosis by inducing ER stress, which then upregulates the second mitochondria-derived caspase activator (Smac). The upregulation of Smac led to a downregulation of the X-linked Inhibitor of Apoptosis (XIAP) through ubiquitination / proteasome activation. CBD has also been shown to induce mitochondrial dysfunction, as demonstrated by the CBD-related decrease in oxygen consumption rate, ATP production, mitochondrial membrane potential and the NADH dehydrogenase ubiquinone 1α subcomplex subunit 9. In vivo, mice injected with MKN45, another gastric cancer cell line, showed slower tumor growth and smaller tumor size when treated with CBD (20 mg / kg) three times a week. As in the in vitro studies, CBD promoted apoptosis and decreased the expression of XIAP in the tumors.
Melanoma cell lines express the endocannabinoid receptors CB1 and CB2. Previous studies have also shown that activation of these receptors with Δ9-THC decreased melanoma growth, proliferation, angiogenesis, and metastasis in vivo. While Δ9-THC looks promising as a treatment for melanoma, the effects of CBD on melanoma have hardly been studied. In a more recent study by Simmerman et al. CBD was tested in a melanic model for mice (B16F10). They set up three groups of mice: control (ethanol and PBS treatment), cisplatin treatment (5 mg / kg intraperitoneally once a week), and CBD treatment (5 mg / kg intraperitoneally twice a week). Survival time was significantly increased and tumor size was significantly reduced in CBD-treated mice compared to control mice, but to a lesser extent than in cisplatin-treated mice. Quality of life was subjectively described and it was found that CBD-treated mice had better quality of life, better mobility, and fewer hostile interactions / fights compared to control mice and cisplatin-treated mice. In this study, no group was treated with a combination of CBD and cisplatin. More research is needed to understand the effects of CBD on human melanoma cells.
In pancreatic cancer, especially ductal adenocarcinoma of the pancreas (PDAC), the chances of treatment and survival have barely improved. Ferro et al. used PDAC cell lines, including ASPC1, HPAFII, BXPC3 and PANC1, and KRASWt / G12D / TP53WT / R172H / Pdx1-Cre + / + (KPC) mice as PDAC models to show that GPR55 accumulates in PDAC tissue and that its interruption results in improved survival and decreased proliferation both in vivo and in vitro. This was mainly done by slowing down the cell cycle at the G1 / S junction by reducing the expression of cyclins without increasing apoptosis. In addition, they found that downstream MAPK / ERK signaling is inhibited in cells lacking GPR55. In vivo, treatment of KPC mice with CBD (100 mg / kg) prolonged survival in a manner similar to gemcitabine (GEM) (100 mg / kg), and when CBD and GEM were used together, survival increased compared to the control around three times. With this combination, cell proliferation was also reduced. CBD was also able to counteract increased ERK activation by GEM, a proposed mechanism of acquired GEM resistance.
Summary and conclusions
As can be seen from the extensive literature, CBD has shown robust antiproliferative and pro-apoptotic effects in a variety of cancers in both cultured cancer cell lines and mouse tumor models. In comparison, CBD generally has milder effects on normal cells from the same tissue / organ. The anti-tumor mechanisms vary depending on the type of tumor and range from an interruption of the cell cycle to autophagy to cell death or a combination thereof. In addition, CBD can also inhibit the migration, invasion and neovascularization of tumors, suggesting that CBD not only acts on tumor cells but can also affect the tumor's microenvironment, e.g. B. by modulating infiltrating mesenchymal cells and immune cells. CBD's reliance on the endocannabinoid receptors CB1 and CB2 or the TRPV family of calcium channels is also different, suggesting that CBD may have multiple cellular targets and / or different cellular targets in different tumors. Mechanistically, CBD appears to disrupt cellular redox homeostasis and trigger a dramatic increase in ROS and ER stress, which could then cause cell cycle arrest, autophagy, and cell death. For future studies, it is crucial to elucidate the interactions between different signaling pathways such as ROS, ER stress, and inflammation in order to better understand how CBD treatment disrupts cellular homeostasis in both tumor cells and infiltrating cells, leading to death of cancer cells and inhibits tumor migration, invasion, metastasis and angiogenesis. The final step in developing CBD as an oncological drug is extensive and well-designed clinical trials that are urgently needed.
The cellular points of attack of CBD
Although the affinity of CBD for CB1 and CB2 is considered to be relatively low, both CB1 and CB2 could be targets for CBD in certain cancer cells and in infiltrating cells in the tumor microenvironment. Other identified cellular targets of CBD are TRPV1, TRPV2, GPR55, and possibly other GPCRs or non-GPCRs. As summarized in Table 1, these cellular targets can vary depending on the type of cancer. For example, the effects of CBD on gliomas depend on TRPV2, but not on CB1, CB2 and TRPV1. On the other hand, the effects of CBD on lung, colon, prostate and cervical cancers are largely dependent on a combination of CB1, CB2 and TRPV1. The simple presence of these receptors on the surface of cancer cells is not necessarily a good predictor of sensitivity to CBD. For example, CB1, CB2 and TRPV1 are strongly expressed on the cell surface of the thyroid cancer cell line SkiMol; however, the inhibition of these receptors only marginally affected the antiproliferative effects of CBD in SkiMol.
CBD induces intracellular ROS and ER stress and increases the immune response
Although the cellular response to CBD treatment can be quite complex, certain issues have emerged that explain the anti-tumor effects of CBD. A common feature of cancer cells treated with CBD is the dramatic increase in ROS, which is likely caused by disruption of intracellular calcium homeostasis and / or mitochondrial function. ER stress and ROS production are closely related and are regulated by ERO1 activity. Either pathway can activate the other, but ultimately they culminate in activation of mitochondrial-mediated cell death due to increased intracellular calcium. The upstream regulation of ROS- and ER-stress-induced apoptosis is largely unknown. One of the possible mechanisms is via the TRPV channels. Wang et al. demonstrated, for example, that treatment of ovarian cancer cells with the TRPV1 antagonist DWP05195 increased ROS production by upregulating NOX; the increased ROS activity led to an upregulation of the CHOP activity and thus to ER stress-mediated apoptosis. Interestingly, the TRPV1 antagonist did not drastically change calcium levels. This suggests another possible mechanism of intracellular calcium regulation - the NOX enzymes. Calcium release from the ER has been shown to activate NOX, which leads to ROS production in endothelial cells. Whether the CBD-induced ER stress and ROS formation are mediated by the activation of CB1, CB2, TRPV1 or other channels remains to be investigated. CBD could regulate intracellular calcium via transmembrane channels or ER release, leading to apoptosis.
The endocannabinoid receptors CB1 and CB2 are highly expressed on inflammatory cells, including B cells, NK cells, monocytes, T cells, and neutrophils. In addition, CB2 is expressed in different ways when B cells and macrophages are activated. Studies of the immunomodulatory role of the endocannabinoid system have shown that CB2 activation inhibits the production of TNF-α, IL-6 and IL-8 in monocytes and macrophages. Unsurprisingly, CBD reduced TNF-α production in macrophages after LPS stimulation. In addition, CBD also decreased the secretion of IL-1β and TNF-α from activated lymphocytes and monocytes in the peripheral blood.
The secretion of cytokines is also largely mediated through the production of ROS, a major source being the NOX2-expressing immune cells. In many types of cancer, MSDCs produce ROS through an increased expression of NOX2, which is regulated by STAT3. CBD has been shown to lower STAT3 levels in colon cancer, prostate cancer, hepatocellular carcinoma, breast cancer, leukemia, and lymphoma. MDSCs lacking NOX2 were unable to prevent T cell proliferation and IFNγ production. The inhibition of STAT3 by CBD strengthens the Th1 immune response and is an important source for the ROS production, which leads to the death of the tumor cells. Whether the downregulation of STAT3 in tumor-associated immune cells is mediated by the CBD agonist or the inverse agonist at CB2 receptors remains to be investigated.
Safety of CBD in humans
Most of the research into the effects of CBD on cancer has not yet reached the clinical trial stage, so we have limited knowledge of the safety profile at the doses required to inhibit tumor growth. In the study by Twelves et al. over CBD and Δ9-THC in the treatment of GBM, dizziness and nausea have been reported as the most common undesirable effects. Outside of cancer treatment, CBD has been shown to be safe with no changes in heart rate, blood pressure, neurological tests, or blood tests. Unlike other controlled substances, patients do not seem to develop tolerance for CBD. Since CBD also affects the expression of various CYP enzymes, it can interact with other drugs; therefore caution should be exercised in patients taking drugs that are metabolized in the liver.
An urgent need for clinical trials
As mentioned earlier, there is extensive pre-clinical research suggesting that CBD, either alone or in conjunction with other cannabinoids, chemotherapy, and radiation therapies is an effective anticancer agent. Although CBD causes mild hepatotoxicity in mice and cats, preliminary toxicity studies suggest that there may still be a therapeutic window for cancer therapy in humans. Therefore, systematic clinical trials of CBD, examining its safety and effectiveness in a wide variety of cancers, are the next logical step in the development of CBD as a cancer drug. This could be done with CBD alone or in combination with established therapeutic modalities.