ArticlePDF AvailableLiterature Review

Cannabinoids as anticancer drugs: current status of preclinical research


Abstract and Figures

Drugs that target the endocannabinoid system are of interest as pharmacological options to combat cancer and to improve the life quality of cancer patients. From this perspective, cannabinoid compounds have been successfully tested as a systemic therapeutic option in a number of preclinical models over the past decades. As a result of these efforts, a large body of data suggests that the anticancer effects of cannabinoids are exerted at multiple levels of tumour progression via different signal transduction mechanisms. Accordingly, there is considerable evidence for cannabinoid-mediated inhibition of tumour cell proliferation, tumour invasion and metastasis, angiogenesis and chemoresistance, as well as induction of apoptosis and autophagy. Further studies showed that cannabinoids could be potential combination partners for established chemotherapeutic agents or other therapeutic interventions in cancer treatment. Research in recent years has yielded several compounds that exert promising effects on tumour cells and tissues in addition to the psychoactive Δ ⁹ -tetrahydrocannabinol, such as the non-psychoactive phytocannabinoid cannabidiol and inhibitors of endocannabinoid degradation. This review provides an up-to-date overview of the potential of cannabinoids as inhibitors of tumour growth and spread as demonstrated in preclinical studies.
Content may be subject to copyright.
Cellular and Molecular Biology
Cannabinoids as anticancer drugs: current status of preclinical
Burkhard Hinz
and Robert Ramer
© The Author(s) 2022
Drugs that target the endocannabinoid system are of interest as pharmacological options to combat cancer and to improve the life
quality of cancer patients. From this perspective, cannabinoid compounds have been successfully tested as a systemic therapeutic
option in a number of preclinical models over the past decades. As a result of these efforts, a large body of data suggests that the
anticancer effects of cannabinoids are exerted at multiple levels of tumour progression via different signal transduction
mechanisms. Accordingly, there is considerable evidence for cannabinoid-mediated inhibition of tumour cell proliferation, tumour
invasion and metastasis, angiogenesis and chemoresistance, as well as induction of apoptosis and autophagy. Further studies
showed that cannabinoids could be potential combination partners for established chemotherapeutic agents or other therapeutic
interventions in cancer treatment. Research in recent years has yielded several compounds that exert promising effects on tumour
cells and tissues in addition to the psychoactive Δ
-tetrahydrocannabinol, such as the non-psychoactive phytocannabinoid
cannabidiol and inhibitors of endocannabinoid degradation. This review provides an up-to-date overview of the potential of
cannabinoids as inhibitors of tumour growth and spread as demonstrated in preclinical studies.
British Journal of Cancer;
The use of the cannabis plant for medicinal and ritual purposes
dates back several thousand years. Accordingly, the psychoactive
effect of cannabis is already mentioned in the Pen-tsao ching, the
oldest pharmacopoeia in the world [1,2]. Further evidence for a
millennia-old use is based on cannabis-containing grave goods
found in archaeological investigations of an Ukok princess
from the Pazyryk culture [3] or the remains of cannabis fruits
identied in archaeobotanical investigations at the Laoguanshan
cemetery from the Han dynasty in Chengdu, South China [4]. The
introduction of cannabis into European medicine can be
attributed to the Irish physician William B. OShaughnessy, who
published a groundbreaking study on hemp in 1839 with his work
On the preparations of the Indian hemp or gunjah[5]. Although
the isolation of a substance called cannabinolfrom the exuded
resin of Indian hemp dates back to a communication published in
1899 [6], it was not until the 1960s that Raphael Mechoulam and
his collaborators published a series of studies that elucidated the
chemical structure and activity of Δ
-tetrahydrocannabinol (THC),
cannabidiol (CBD), and other cannabinoids [710].
Endocannabinoid system
The entry of the endocannabinoid (EC) system into modern
research as a potential target of pharmacotherapeutic interven-
tion began with the discovery and cloning of specicG
coupled cannabinoid receptors, termed CB
and CB
While CB
receptors are primarily localised in the central nervous
system, CB
receptors are mostly expressed on cells of the
immune system. Other components of the EC system discovered
in the 1990s are N-arachidonoylethanolamine (anandamide, AEA)
and 2-arachidonoylglycerol (2-AG), two endogenously synthe-
sised agonists at cannabinoid receptors [13,14]. Scientic
attention has also been paid in recent years to substances with
structural similarities to the aforementioned ECs, such as the
cannabinoid receptor ligands 2-arachidonoyl glycerol ether
(noladin ether), O-arachidonoylethanolamine (virodhamine),
N-arachidonoyldopamine and oleic acid amide (oleamide)
(reviewed in ref. [15]). However, the available data on their
biological role are very limited. Furthermore, a number of
N-acylethanolamines structurally similar to ECs, so-called EC-like
substances, such as N-palmitoylethanolamine (PEA) and
N-oleylethanolamine (OEA), have been described, which use the
biosynthesis and degradation enzymes of ECs, but do not trigger
cannabinoid receptor activation (reviewed in ref. [15]).
Around the turn of the millennium, the non-selective cation
channel transient receptor potential vanilloid 1 (TRPV1) was
described as an additional receptor target for several cannabi-
noids such as AEA [16] and the non-psychoactive phytocannabi-
noid CBD [17]. Among the phytocannabinoids, THC exhibits the
properties of an agonist at the CB
receptor and a partial
agonist at the CB
receptor [18], as well as an agonist at the G
Received: 19 August 2021 Revised: 9 December 2021 Accepted: 28 January 2022
Institute of Pharmacology and Toxicology, Rostock University Medical Centre, Schillingallee 70, 18057 Rostock, Germany. email: burkhard.hinz@med.uni-
British Journal of Cancer
Published on Behalf of CRUK
protein-coupled receptor (GPR) 55 [19]. In contrast to the high-
afnity CB
and CB
receptor binding of THC with K
values in the
low nanomolar range, CBD has been demonstrated to have
weaker afnities with K
values in the micromolar range and non-
competitive antagonistic effects at both CB
and CB
receptor [18].
In addition, CBD shows a binding preference to the CB
[20] and an antagonistic effect at GPR55 [19]. Among a variety of
receptor interactions of CBD, the compound has been shown to
increase the transcriptional activity of peroxisome proliferator-
activated receptor γ(PPARγ)[21].
Other important elements affecting the tone of the EC system are
EC-synthesising and -degrading enzymes. In this context, N-acyl-
hydrolase domain-containing 4 (ABHD4), glycerophosphodiesterase-1
(GDE1), and tyrosine protein phosphatase non-receptor type 22
(PTPN22) have been described to contribute to AEA biosynthesis.
Diacylglycerol lipase αand -β(DAGLαand -β) have been identied as
2-AG-producing enzymes (reviewed in ref. [22]). The degradation of
AEA and 2-AG is endogenously mediated by the enzyme fatty acid
amide hydrolase (FAAH) [23], whereas the hydrolysis of 2-AG
proceeds mainly via monoacylglycerol lipase (MAGL) [24]with
contribution of several other hydrolytic enzymes (ABHD6, ABHD12,
FAAH). In addition, ECs can be degraded by enzymes of the
arachidonic acid cascade, i.e. cyclooxygenase-2 (COX-2) and lipox-
ygenases (reviewed in ref. [22]).
It is worth noting that the denition of the EC system has been
subject to evolution over time. Thus, the classical EC system only
includes the two ECs AEA and 2-AG, their anabolic and catabolic
enzymes, and the two cannabinoid receptors CB
and CB
However, as more became known about its complex networks
over time, the term and concept of the EC system was expanded
accordingly, leading to the denition of an endocannabinoi-
dome[25]. This now includes also other EC-like lipid mediators,
metabolic enzymes and novel cannabinoid targets such as GPRs
(GPR18, GPR55, GPR119) and members of the transient receptor
potential cation channel subfamily (TRPV1, TRPV2, TRPV4)
(reviewed in ref. [15,25]).
Anticancer effect of cannabinoidspioneering work and
research strategies
The discovery of the anticarcinogenic effect of cannabinoid
compounds can be dated back to Munson et al. [26], who were
able to show in the mid-1970s that THC, Δ
-THC and cannabinol
inhibit Lewis lung adenocarcinoma growth in mice. However, the
discovery of cannabinoid receptors years later was only
the starting point for an extensive and detailed investigation of
the anticarcinogenic mechanisms of cannabinoid action. For
example, a study at the turn of the millennium showed that
intratumoural administration of THC and the synthetic cannabi-
noid agonist WIN 55,212-2 induced a considerable regression of
malignant gliomas in Wistar rats and in mice decient in
recombination activating gene 2 [27]. A growth inhibitory effect
of cannabinoids against glioma cells was conrmed shortly
thereafter for the selective CB
agonist JWH-133 [28]. With the
discovery of a possible role of the EC system as a factor in cancer
progression, numerous studies have also addressed a possible link
between the regulation of cannabinoid receptors, ECs, and EC-
synthesising and -degrading enzymes on the one side and disease
severity and survival on the other (reviewed in ref. [15]). However,
no clear correlation between the regulation of these parameters in
tumour tissue and disease severity could be found, so that the
aforementioned factors therefore do not show suitability as
reliable biomarkers.
In addition to the activation of cannabinoid receptors by
exogenously applied agonists, newer approaches to pharma-
cotherapeutic intervention by blocking EC turnover have been
pursued and further developed in recent years, such as the
blockade of MAGL by the selective MAGL inhibitor JZL184 [29]. In
the context of potential cancer-preventive effects, knockdown or
pharmacological inhibition of MAGL have been associated with
decreased tumour cell invasion [3032], metastasis [32] and
tumour growth [30,31]. This inhibitory effect on cancer
progression by MAGL inhibition appears to be due to a dual-
mode effect in prostate cancer cells involving both activation of
receptor by increasing 2-AG levels and reduction of
protumorigenic free fatty acids resulting from MAGL activity
[30], whereas for the anti-invasive effect on lung cancer cells, only
receptor involvement has been demonstrated [32]. On the
other hand, a recent publication concluded that inhibition of
MAGL promotes rather than retards cancer progression in mice
and that knockout of MAGL is associated with increased incidence
of lung adenocarcinoma [33]. With regard to the tumour
suppressor function of MAGL, it was shown in this context that
MAGL can inhibit the transactivation of epidermal growth factor
receptor (EGFR)-associated signalling pathways as well as proin-
ammatory proteins such as COX-2 and tumour necrosis factor
(TNF)-α. Accordingly, it is reasonable to assume that MAGL acts as
a tumour suppressor or oncoprotein depending on the tissue
type. Finally, inhibition of FAAH has also been shown to mediate
cancer-limiting effects with growth inhibitory [34,35] as well as
anti-invasive and antimetastatic properties [36]. The preclinical
work on the inhibition of FAAH and MAGL in cancer has been
summarised elsewhere (reviewed in ref. [15]).
Recently, further substances from Cannabis sativa L., which are
also found in other plants, have been investigated for their anti-
cancer effects. This concerns, for example, the sesquiterpene β-
caryophyllene, a potent CB
agonist [37]. In the hitherto
performed studies, β-caryophyllene showed antiproliferative and
proapoptotic properties on various cancer cell lines [38,39] and
enhanced the cytostatic effects of classical chemotherapeutic
agents such as doxorubicin [40] and sorafenib [41].
The rst pilot clinical trial of cannabinoids as cancer treatment in 9
glioblastoma patients was published in 2006 and found that
intracranially administered THC was safe [42]. Recently, the results
of a randomised, placebo-controlled phase 1b study with nabiximols
oromucosal spray (standardised extract of Cannabis sativa L. with an
approximate 1:1 ratio of THC and CBD) in combination with dose-
intense temozolomide in patients with recurrent glioblastoma
multiforme were published [43]. Of the 21 patients in this study,
survival at one year was 83% in the 12 patients treated with
nabiximols, while it was signicantly lower at 44% in the 9 patients
treated with placebo, leading the authors to recommend further
exploration in an adequately powered randomised controlled trial. So
far, however, randomised, placebo-controlled studies with a larger
number of cancer patients are lacking at all.
The following chapters provide an overview of the preclinical
evidence on the effect of cannabinoids on cancer growth and
metastasis, with the most relevant cannabinoid targets sum-
marised in Fig. 1(tumour cell proliferation, apoptosis and
autophagy) and Fig. 2(tumour cell invasion, angiogenesis).
Publications on growth inhibitory effects of cannabinoids have
accumulated over the last two decades. The dominant cellular
model used for this purpose in the 2000s was glioma cells.
However, over time, a wide range of different tumour cell lines of
various entities have been tested. Whereas early work focused on
mechanisms leading to cannabinoid-induced apoptosis and cell
cycle arrest of cancer cells, these investigations were comple-
mented later by studies dealing with autophagy effects. Due to
the large amount of data, only a selection could be made here.
Tumour cell proliferation
As mentioned earlier, the description of the tumour regressive
effects of THC and WIN 55,212-2 on rat and mouse glioma
B. Hinz and R. Ramer
British Journal of Cancer
xenografts represents the rst comprehensive study in this
century investigating cannabinoids as potential anticancer drugs
[27]. Since then, several mechanisms by which cannabinoids
inhibit tumour growth have been elucidated. Among these,
inhibition of protein kinase B (Akt) appears to be an important
mechanism. Accordingly, the antiproliferative action of cannabi-
noids on melanoma cells has been linked to cell cycle arrest at the
/S transition via inhibition of the prosurvival protein Akt and
hypophosphorylation of the retinoblastoma protein (pRb) tumour
suppressor protein [44]. Furthermore, CB
inhibition of Akt has been reported in ErbB2-positive breast
cancer progression in mouse mammary tumour virus (MMTV)/neu
transgenic mice as a syngeneic tumour model [45]. Meanwhile,
publications have conrmed that inactivation of the Akt pathway
is also involved in the antitumour activity of cannabinoids on
human gastric cancer [46], non-small cell lung cancer [47] and
hepatocellular carcinoma (HCC) cells [48]. In breast cancer cells,
features of cell cycle arrest induced by cannabinoid treatment
include blockade of the G
/S transition through downregulation of
cyclin-dependent kinase 1 (CDK1), induction of p21 [49], or
induction of p27kip1, decrease in cyclin (CCN) A and E,
degradation of cell division cycle 25 A (CDC25A) and inactivation
of CDK2 [50]. Furthermore, in prostate cancer cells, WIN 55,212-2
was shown to reduce proliferation and to arrest cells in the G
phase via CB
receptor-dependent signalling [51]. Studies of the
underlying mechanism showed an upregulation of p27 and a
reduction in CDK4 expression and phosphorylated pRb (P-pRb)
when cells were treated with the cannabinoid.
CBD appears to inhibit cancer cell proliferation primarily via
apoptosis signalling. Thus, in a recent study [52], CBD suppressed
the proliferation and growth of head and neck squamous cell
carcinomas by inducing the apoptotic and autophagy activity of
DUSP1 (Dual Specicity Phosphatase 1), which is known to
interfere as a negative regulator with EGFR-initiated mitogen-
activated protein kinase (MAPK) signalling and associated
proliferation. Furthermore, increased expression levels of p21,
Kruppel-like factor 6 (KFL6) and growth arrest and DNA damage-
inducible protein α(GADD45A) were observed, for which
antiproliferative effects were also described. Another recent study
investigating the in vitro effects of CBD on human gastric cancer
cells revealed an antiproliferative effect accompanied by sig-
nicant upregulation of ataxia telangiectasia mutated (ATM) gene
and p21 protein expression and downregulation of p53 protein
expression, which subsequently resulted in decreased levels of
CDK2 and CCNE and cell cycle arrest in the G
phase [53].
Finally, a contribution of the eicosanoid system was also
supported by the observation that CBD reduced the activity and
content of 5-lipoxygenase (5-LOX) and its end product leukotriene
in the tumour tissue of nude mice xenografted with human
glioma cells [54]. In addition, a synergism of the viability-reducing
effect of CBD and the 5-LOX inhibitor MK-886 could be shown
in vitro, which also argues for a modulatory effect of 5-LOX on
glioma cell fate.
Conversely, a recent study has demonstrated that the CB
agonist arachidonyl-2-chloroethylamide (ACEA), the selective CB
agonist HU308 and THC promoted rather than inhibited the
progression of human papillomavirus-positive squamous cell
carcinomas of the head and neck, with THC-treated tumour
xenografts growing faster than controls [55]. This work is in line
with previous in vitro studies by other authors [5658], who also
found proliferative effects on cancer cells triggered by submicro-
molar concentrations of various cannabinoids, with corresponding
THC effects associated with transactivation of EGFR in one paper
[56]. Some other studies, in contrast, failed to observe mitogenic
Fig. 1 Mechanisms of antiproliferative, proapoptotic and proautophagic effects of cannabinoids on cancer cells. The black arrows
emanating from the cannabinoid show the respective modulated structures or levels. Coloured arrows indicate inhibitory (red) and
stimulatory (blue) effects of cannabinoids on the indicated targets. Blue dashed arrows indicate reduced stimulation of the respective effect
by cannabinoid treatment. The grey arrows indicate a shift in a parameter. The black dashed arrow indicates a functional relationship between
autophagy and apoptosis. All abbreviations are explained in the text.
B. Hinz and R. Ramer
British Journal of Cancer
effects of THC when nanomolar concentrations were tested
[42,59,60]. Overall, however, these results seem worth reconsider-
ing, as peak plasma concentrations of THC after inhalation or oral
administration do not indeed exceed 1 μM[55]. In fact, many
experimental studies investigating proapoptotic effects of canna-
binoids have been conducted with concentrations above 1 µM,
which, on the other hand, could well be achieved with intratumoral
administration [27]. Interestingly, hardly any mitogenic effects can
be observed in the case of CBD (reviewed in ref. [61]).
Tumour cell apoptosis
An important role in cannabinoid-induced apoptosis is played by
the proapoptotic sphingolipid ceramide. Initial studies showed
that THC and other cannabinoids cause the death of glioma
cells via cannabinoid receptor-dependent de novo synthesis of
ceramide [27,28,62]. The importance of ceramide for
cannabinoid-induced cytotoxic autophagy was discovered later
in 2009 [63]. Further apoptosis mechanisms were described with
receptor-dependent induced apoptosis of glioma cells
associated with an increase in the stress-associated transcriptional
coactivator p8 as an upstream regulator of the endoplasmic
reticulum (ER) stress-related proteins activating transcription
factor (ATF)-4 and tribbles pseudokinase 3 (TRB3) [64]. A similar
apoptotic effect was found for CB
receptor-induced growth
inhibition in response to Akt inhibition in translocation-positive
rhabdomyosarcoma cells [65]. Here, cannabinoid-induced loss of
viability was suppressed by transfection of p8 siRNA.
Depending on the substance and cell type, the eicosanoid
system also revealed to play an important role in the proapoptotic
effect of cannabinoids. Thus, in temporal relation to the rst
mechanistic ndings on cannabinoid-induced glioma cell death,
ceramide synthesised de novo by R(+)-methanandamide (Met-
AEA) in neuroglioma cells was shown to lead to an induction of
COX-2 expression [66], which contributes to Met-AEA-induced cell
death via proapoptotic prostaglandin (PG) E
[67]. Thereby, Met-
AEA-triggered apoptosis was found to be independent of
cannabinoid receptor and TRPV1 activation. The inuence of
COX-2 on cancer cell fate was also addressed in follow-up work on
the effect of Met-AEA [68] and the established chemotherapeutic
agents paclitaxel, cisplatin and 5-uorouracil [69] on human
cervical carcinoma cells. Here, the PPARγ-activating eicosanoids
and 15-deoxy-Δ
could be identied as apoptosis
mediators, whereby post-transcriptional knockdown of COX-2,
downstream lipocalin-type PGD synthase (L-PGDS) and transcrip-
tion factor PPARγled to inhibition of apoptosis triggered by Met-
AEA and chemotherapeutic agents. A functional role of the
aforementioned PPARγ-activating PGs could also be proven for
the CBD-triggered cannabinoid receptor and TRPV1-independent
apoptotic death of lung cancer cells [70]. CBD also led to an
upregulation of COX-2 and PPARγin tumour tissue in A549-
xenografted nude mice and to tumour regression, which was
reversed by a PPARγantagonist [70].
As already mentioned above, cannabinoid receptors CB
do not necessarily have to be involved in the apoptosis
Fig. 2 Mechanisms of anti-invasive, antimetastatic, anti-epithelial-to-mesenchymal-transition and anti-angiogenic effects of cannabi-
noids on cancer cells. The black arrows emanating from the cannabinoid show the respective modulated structures or levels. Coloured arrows
indicate the inhibitory (red) and stimulatory (blue) effects of cannabinoids on targets involved in cancer cell invasion/metastasis, angiogenesis
and epithelial-to-mesenchymal transition. Blue dashed arrows indicate reduced stimulation of each effect by cannabinoid treatment. Black lines
with circles at both ends indicate binding and dimer formation between the respective parameters. Black dashed arrow indicates functional
relationship between epithelial-to-mesenchymal transition and invasion/metastasis. All abbreviations are explained in the text.
B. Hinz and R. Ramer
British Journal of Cancer
induction of a cannabinoid. In a previous study, AEA was
demonstrated to induce apoptotic death of human neuroblas-
toma and lymphoma cells via TRPV1, which was paralleled by an
increase in intracellular calcium, mitochondrial uncoupling and
cytochrome c (Cyt c) release [71]. In other studies, AEA has been
shown to induce receptor-independent apoptosis of non-
melanoma cancer cells via ER stress [72] and to elicit reduced
viability of cholangiocarcinoma cells via activation of GPR55, with
the latter eliciting increased recruitment of the death receptor Fas
in membrane lipid rafts [73]. Furthermore, activation of TRPV2 has
been revealed to be involved in the proapoptotic effect of CBD on
human bladder cancer cells [74]. Mostly, however, the proapop-
totic effects of CBD have been associated with receptor-
independent mechanisms, although corresponding inhibitor
experiments with receptor antagonists have not always been
performed. This applies, for example, to some studies in which
CBD elicited apoptosis of mammary carcinoma [75] and human
gastric cancer cells [53] via the formation of reactive oxygen
species (ROS). Interestingly, however, CBD induced a CB
dependent mitochondrial apoptosis in human leukaemia cells,
which was accompanied by ROS production, increased expression
of the NAD(P)H oxidases Nox4 and p22phox and release of Cyt c
[76]. In a recently published study [77], CBD was also shown to
promote apoptosis of gastric cancer cells by suppressing X-linked
inhibitor apoptosis (XIAP), a member of the IAP protein family.
Thereby, CBD reduced XIAP protein levels while increasing
ubiquitination of the protein. In addition, CBD increased the
interaction between XIAP and Smac by inducing the release of
Smac from mitochondria into the cytosol and promoted
mitochondrial dysfunction. Finally, recent evidence suggests that
CBD at a relatively high concentration of 30 µM switches the
mitochondrial voltage-dependent anion channel (VDAC) from fully
open to major subconductance state, thereby arresting the
-permeable state of this channel and inducing a severe
oxidative stress, mitochondrial Ca
overload, Cyt c release into
the cytosol as well as induction of LC3-phosphatidylethanolamine
conjugate (LC3-II) and caspase activation in leukaemia cells [78].
Tumour cell autophagy
Autophagy can lead to survival or death of cells. Research in
recent years has shown that autophagy signalling pathways may
also play an important role in the toxicity of cannabinoid
compounds on cancer cells. Initial experiments conducted by
Salazar et al. [63] demonstrated that THC induces death of human
glioma cells via stimulation of autophagy. In this process, THC led
to ceramide accumulation and phosphorylation of eukaryotic
translation initiation factor 2α(eIF2α), a subsequent ER stress
response and, via this, nally to the induction of autophagy via the
TRB3-dependent inhibition of the Akt/Mammalian Target of
Rapamycin Complex 1 (mTORC1) axis. In the processes described,
autophagy preceded apoptosis. In another work, proautophagic
effects of THC and the CB
agonist JWH-015 in human HCC cells
were reported [79]. Here, autophagy was due to upregulation of
TRB3 and subsequent inhibition of the Akt/mTORC1 axis and
adenosine monophosphate-activated kinase (AMPK) stimulation,
with Ca
/calmodulin-activated kinase kinase β(CaCMKKβ) being
responsible for cannabinoid-induced AMPK activation and au-
tophagy. Importantly, the tumour regressive effects of THC and
JWH-015 on subcutaneous HCC xenografts were abolished in vivo
when autophagy was genetically or pharmacologically inhibited
[79]. A functional link between autophagic signalling pathways
and cannabinoid-induced apoptosis was conrmed by several
other studies. Accordingly, in melanoma cells, co-treatment of THC
with the autophagic ux inhibitor chloroquine or knockdown of
autophagy-related 7 (Atg7), an essential molecule for induction of
autophagy, resulted in suppression of THC-induced autophagy as
well as cell death [80]. Consistent with this, inhibition of CBD-
induced autophagy by chloroquine also led to a signicant
increase in the viability of CBD-treated human squamous cell
carcinomas of the head and neck [52].
On the other side and in contrast to previous studies on glioma
cells, a recent study reported that knockdown of autophagy genes
led to enhancement of WIN 55,212-2-induced apoptotic cell death
of human glioblastoma cells [81]. The authors concluded that
autophagy induced by cannabinoid treatment is a protective
mechanism and autophagy inhibitors may be potential agents to
enhance cannabinoid action. Evidence of cannabinoid-protective
autophagy was also provided by a glioblastoma cell culture study
in which the addition of chloroquine led to an increase in CBD-
induced cell death [82].
In glioma stem cells, a cell subpopulation of glioblastoma
multiforme implicated in chemoresistance, CBD was further
shown to induce a TRPV2-dependent autophagic process that
stimulates glioma stem cell differentiation via induction of a splice
variant of the acute myeloid leukaemia transcription factor
(Aml-1), thereby abrogating their chemoresistance to carmustine
[83]. In breast cancer cells, CBD-induced intrinsic apoptosis was
associated with autophagy and decreased levels of mTOR, its
downstream effector eukaryotic initiation factor 4E binding
protein 1 (4EBP1) and CCND1 [75]. Finally, in a recent study, a
signicant increase in LC3-II levels as an autophagy marker was
observed in two out of three medulloblastoma cell lines after CBD
and THC exposure, which was accompanied by an increase in poly
(ADP-ribose) polymerase (PARP) cleavage [84]. However, these
ndings did not translate to in vivo models in mice, as the survival
rate of the animals did not change.
As another cannabinoid-triggered upstream ion channel a
recent investigation found TRPV4 and a signalling pathway
including ATF4, DNA Damage Inducible Transcript 3 (DDIT3),
TRB3, Akt and mTOR as mediators of CBD-induced mitophagy in
glioma cells [85]. Mitophagy involves the selective degradation of
mitochondria by autophagy and is manifested in mitophagy-
related proteins such as BCL2 interacting protein 3 (BNIP3), Cyt c,
Parkin and engulfed mitochondria, with mitochondrial dysfunc-
tion and overactivation of heme oxygenase-1 (HO-1) acting
synergistically in lethal mitophagy [86].
Tumour cell invasion
A large number of publications also point to an inhibitory effect of
cannabinoid compounds on tumour cell migration, invasion and
in vivo metastasis (Fig. 2). The rst report on this referred to the
receptor-dependent inhibition of prostate cancer cell invasion
by the EC 2-AG [87]. Anti-invasive actions were also later proven
for AEA in glioma [88] and lung cancer cells [36]. A few years
earlier, the AEA derivative Met-AEA had already been shown to
exhibit anti-invasive properties on human cervical and lung cancer
cells via cannabinoid receptors and TRPV1 [89].
In one of the rst mechanistic studies in this eld, induction of
tissue inhibitor of metalloproteinase-1 (TIMP-1) expression was
shown to be a cause of the anti-invasive effect of THC, Met-AEA
and CBD in cervical and lung cancer cells, with cannabinoid
receptors (THC, Met-AEA, CBD) and TRPV1 (Met-AEA, CBD)
mediating this response [89,90]. Crucially, TIMP-1 inhibits
collagen-degrading enzymes such as matrix metalloproteinase
(MMP)-2 and MMP-9, which play an important role in promoting
cancer metastasis (reviewed in ref. [91]). Accordingly, overexpres-
sion of TIMP-1 has been associated with a reduction in tumour
growth, cancer cell invasiveness and metastasis (reviewed in ref.
[92]). An anti-invasive effect based on TIMP-1 induction in lung
cancer cells was also conrmed for the FAAH inhibitors
N-arachidonoyl-serotonin (AA-5HT) and URB597, AEA and OEA
[36] as well as for the MAGL inhibitor JZL184 and the MAGL
substrate 2-AG [32]. For the anti-invasive effect of THC, CBD and
Met-AEA on lung tumour cells, a cannabinoid-mediated increased
B. Hinz and R. Ramer
British Journal of Cancer
expression of the intercellular adhesion molecule-1 (ICAM-1) as an
upstream inducer of TIMP-1 expression could be demonstrated
In support of the importance of modulation of extracellular
matrix proteolysis in anti-invasive cannabinoid action, other
groups have demonstrated downregulation of MMP-2 by THC in
glioma cells [94] and downregulation of MMP-2 and -9 in HCC cells
treated with CB
(ACEA) or CB
agonists (CB65) [95]. Notably, the
antimigratory effect of the synthetic cannabinoid WIN 55,212-2 on
osteosarcoma cells was associated with downregulation of MMP-2
and -9 and a 700-fold upregulation of miR-29b1, a key miRNA that
downregulates MMP-2 and -9 [96]. A recent study also focused on
the role of cancer-associated broblasts as a crucial element of the
stromal compartments in the tumour microenvironment. Here,
conditioned media from WIN 55,212-2-treated cancer-associated
broblasts were shown to impair the invasive properties of
prostate cancer cells due to a cannabinoid-mediated reduction in
MMP-2 release [97].
In breast cancer, glioblastoma and salivary gland cancer cells, a
CBD-mediated downregulation of Id-1, an inhibitor of basic helix-
loop-helix transcription factors, has been reported as the cause of
the anti-invasive effect of this compound [98100]. CBD also led to
a downregulation of the sex-determining region Y (SRY)-Box 2
(Sox-2), a critical determinant of glioma tumour initiating cell
growth and downstream target of Id-1, in glioblastoma cells [99].
Finally, the antimetastatic effect on breast cancer cells demon-
strated for CBD in a mouse model [101] was directly linked to a
downregulation of Id-1 in a later work [102]. In another
investigation, siRNA, inhibitor and add-back experiments showed
that the cannabinoid receptor- and TRPV1-dependent down-
regulation of plasminogen activator inhibitor (PAI)-1 in CBD-
exposed lung cancer cells [103] is part of the anti-invasive effect of
this cannabinoid, in addition to the upregulation of TIMP-1
mentioned above. Still other work revealed that the anti-invasive
effect of THC on cholangiocarcinoma cells is associated with
reduced activation of Akt and p42/44 MAPK [104]. Furthermore, a
very recent work found that treatment with CBD in combination
with THC or CBD alone inhibited bladder urothelial carcinoma cell
migration independently of cannabinoid receptors [105].
Contradictory results have been published on the role of the CB
receptor in the invasion process. While studies with selective CB
agonists [95,102] as well as inhibitor studies [36,89,90,93,103]
suggest a role for CB
in the anti-invasive effects of various
cannabinoids, a recent investigation reported that silencing of the
receptor reduced proliferation, migration and invasion of lung
cancer cells, which was associated with reduced levels of phospho-
Akt, phospho-mTOR and decreased expression of 70-kDa ribosomal
protein S6 kinase (p70S6K), a mitogen-activated Ser/Thr protein
kinase that promotes cell survival and growth [106]. With respect to
the CB
receptor, it has also been shown that the latter can form an
induced heterodimer with the G protein-coupled chemokine
receptor CXCR4 in human breast and prostate cancer cells [107].
Here, simultaneous agonist-dependent activation of CXCR4 and CB
resulted in reduced CXCR4-mediated formation of phosphorylated
p42/44 MAPK (P-p42/44) and ultimately diminished cancer cell
chemotaxis. Therefore, cannabinoids could also negatively modulate
tumour progression by interfering with CXCR4 receptor function.
A number of other publications addressed the role of epithelial-
mesenchymal transition (EMT)-lowering properties of cannabinoid
compounds as a contributing mechanism of action to reduce
cancer aggressiveness (Fig. 2). An early study on this topic showed
that the AEA derivative 2-methyl-2-F-anandamide (Met-F-AEA)
reduced cytoplasmic and nuclear protein levels of β-catenin, one
of the key factors involved in the EMT transition [108]. In this
publication, Met-F-AEA further caused signicant inhibition of
mesenchymal markers such as vimentin, N-cadherin, bronectin
and EMT markers (Snail1, Slug and Twist) and upregulation of
epithelial markers such as E-cadherin and cytokeratin 18. Recently,
CBD was also shown to reverse interleukin (IL)-1β-induced EMT of
human breast cancer cells by reprogramming invasive cells into
cells with a non-invasive phenotype [109]. Here, CBD induced the
relocalisation of E-cadherin and β-catenin at adherens junctions,
thereby preventing nuclear translocation of β-catenin and
inhibited the expression of the EMT marker ΔNp63, an isoform
of tumour protein 63 (TP63), baculoviral IAP repeat-containing
protein 3 (BIRC3) and Id-1. In the same work, studying the
malignant phenotype of breast cancer cells, CB
mediated inhibition of viability by CBD was registered, as well as
an antimigratory effect of the cannabinoid associated with
inhibition of Akt phosphorylation [109]. Others observed a reversal
of transforming growth factor (TGF)-β-induced spindle-shaped
morphology of lung cancer cells corresponding to the reorganisa-
tion of the stress bre F-actin when the cells were treated with a
combination of THC and CBD [110]. In this study, TGF-β-induced
inhibition of E-cadherin expression and upregulation of
N-cadherin and vimentin were signicantly reversed as a
characteristic EMT regulatory pattern in the presence of CBD,
THC or the combination of both. Finally, these cannabinoid-
mediated regulations were functionally linked to a reduced
migration potential of lung cancer cells.
The effect of cannabinoids in experimental metastasis models has
been described in detail elsewhere (reviewed in ref. [111]). In
short, phytocannabinoids (THC, CBD), ECs and EC analogue (AEA,
2-AG, Met-F-AEA), EC-like substances (OEA, PEA) and EC degrada-
tion inhibitors (AA-5HT, URB597, JZL184) showed an inhibitory
effect on metastatic inltration of the lung with previously
injected lung carcinoma cells [32,36,90,93,112,113]. In addition,
inhibitory effects of cannabinoids on breast cancer [101,102,114],
salivary gland cancer [100] and melanoma cell metastasis [44]
have been described. Finally, further work showed that JZL184
impairs bone metastasis of osteotropic prostate and breast cancer
cells in mice and inhibits metastasis of osteosarcoma cells [115]
and that knockdown of MAGL is associated with reduced lymph
node metastasis in MAGL-overexpressing nasopharygeal carci-
noma cells [116].
Regarding the underlying mechanisms, one investigation
demonstrated increased ICAM-1 expression as a cause of the
antimetastatic effect of CBD on lung cancer cells, with the effect
being reversed by an ICAM-1 neutralising antibody [93]. Another
work showed that CBD inhibited lung metastasis of breast cancer
cells expressing a control vector but not an Id-1-containing vector
[102]. Further studies have identied the CB
receptor as an initial
platform of the antimetastatic effect of Met-F-AEA [112] and
JZL184 [32], as the inhibition of metastasis induced by these
compounds was counteracted by CB
receptor antagonists.
Several studies have shown that cannabinoid compounds inhibit
tumour neovascularisation in mouse models with xenografts
(reviewed in ref. [117]). However, data on the exact mechanisms
underlying these effects, especially with regard to tumour-stroma
interactions, are still scarce. Early reports on the effect of cannabinoid
compounds on tumour formation found a regressive effect based on
anti-angiogenic effects through the downregulation of a number of
proangiogenic parameters such as vascular endothelial growth factor
(VEGF), placental growth factor (PlGF), angiopoietin-2 (Ang-2)
[118,119] and MMP-2 [119].Thesamegrouplaterrevealedthat
the CB
receptor agonist JWH-133 modulates several hypoxia-related
angiogenesis markers, most of which are associated with the VEGF
pathway [120]. Thus, downregulation of VEGF-A and -B, hypoxia-
inducible factor-1α(HIF-1α), connective tissue growth factor (CTGF),
midkine, Id-3, Ang-2 and its receptor tyrosine kinase with
immunoglobulin-like and EGF-like domains 1 (Tie-1), and HO-1 was
B. Hinz and R. Ramer
British Journal of Cancer
detected, while upregulation was found for the type I procollagen α1
chain (COL1A1) [120]. Remarkably, the authors of the latter study
found VEGF receptor (VEGFR)-2 downregulation in response to JWH-
133 in experimental glioma xenografts in mice. In another paper,
inhibition of MMP-2 expression in human umbilical vein endothelial
cells (HUVEC) was conrmed as part of the anti-angiogenic effects of
CBD [121]. Consistent with the aforementioned direct anti-angiogenic
effects, a further investigation identied the hexahydrocannabinol
analogues LYR-7 [(9S)-3,6,6,9-tetramethyl-6a,7,8,9,10,10a-hexahydro-
6H-benzo[c]chromen-1-ol] and LYR-8 [(1-((9S)-1-hydroxy-6,6,9-tri-
none)] as inhibitors of VEGF-induced proliferation, migration and
capillary-like tube formation of human endothelial cells as well as
VEGF-induced blood vessel formation in the chorioallantoic mem-
brane assay and VEGF release from cancer cells [122]. Thereby, the
effects of the test substances on cell proliferation and tube formation
were not abolished by cannabinoid receptor antagonists, suggesting
a cannabinoid receptor-independent mechanism.
A number of studies have also demonstrated indirect effects of
cannabinoids on endothelial cells through cannabinoid-mediated
modulation of the secretome of tumour or other non-endothelial
cells. Thus, conditioned media from AEA-treated breast cancer cells
showed an inhibitory effect on endothelial cell proliferation associated
with an approximate halving of the VEGF concentration, but also in a
signicant reduction of other proangiogenic factors such as leptin and
thrombopoietin (THPO) [123]. A further indirect effect, here mediated
by cannabinoid-induced increased formation of anti-angiogenic TIMP-
1, was reported for the effect of conditioned media from CBD-, THC-,
Met-AEA- and JWH-133-treated lung cancer cells on tube and sprout
formation and endothelial cell migration [124]. A recent report also
dealt with the effect of cannabinoids on human neutrophils with
special regard to the regulation of angiogenic factors. Thereby,
submicromolar concentrations of ACEA and JWH-133 inhibited
lipopolysaccharide (LPS)-induced VEGF-A release, which was accom-
panied by a decrease in LPS-induced angiogenic effects on bovine
aortic endothelial cell tube formation [125].
Divergent ndings have been published regarding the direct
effects of cannabinoid compounds on endothelial cells, which
depend on the cellular system, the substance and the concentration
used. Thus, in a previous study, inactivation of the CB
receptor with
the antagonist SR141716 as well as CB
knockdown using siRNA
caused inhibition of basic broblast growth factor (bFGF)-induced
proliferation, migration and capillary-like tube formation of HUVEC
[126] through pathways involving decreased activation of focal
adhesion kinase, c-Jun N-terminal kinase, RhoA and MMP-2.
Moreover, CB
antagonism was conrmed in vivo to inhibit bFGF-
induced neovascularisation in the rabbit cornea [126]. As for the
direct effects of ECs on endothelial cells, it has been shown that
bFGF-stimulated proliferation of HUVEC is upregulated by nanomo-
lar concentrations of AEA [126], while in another work of the same
group, micromolar proapoptotic concentrations of the AEA analo-
gue Met-F-AEA inhibited bFGF-stimulated proliferation of porcine
endothelial cells in a CB
receptor-dependent manner [127]. In a
further study, no anti-angiogenic effects of ECs on endothelial
proliferation were observed and proangiogenic effects of THC and
CBD were found at nanomolar concentrations [128]. Others reported
that CBD, THC, Met-AEA and JWH-133 (3 µM each) exhibited pro-
rather than anti-angiogenic effects in HUVEC directly exposed to
these substances [124]. Finally, AEA was demonstrated to induce
endothelial cell tube formation and proliferation, with both effects
being reversed by inactivation of TRPV1 [129]. The authors assumed
a TRPV1-dependent uptake of AEA into endothelial cells, which
subsequently causes a proangiogenic effect on endothelial cells via
the activation of intracellular cannabinoid receptors.
In terms of the direct inuence of CBD on the endothelium, a
recent study should also be mentioned which showed that CBD
promotes ROS-dependent HO-1 expression in HUVEC, followed by
HO-1-dependent cytoprotective autophagy [130]. This protection
was maintained up to a certain CBD concentration (in the cited
study up to 6 µM), but was then no longer sufcient to protect the
cells from apoptotic cell death, which was also HO-1-dependent. The
proapoptotic concentration of 10 µM CBD registered in this work is
within the concentration range of the previously shown anti-
angiogenic effects of CBD at concentrations M [121]. In the
latter study, however, cytostasis, but not the induction of apoptosis,
was associated with the decrease in metabolic activity. In this
context, the use of different supplements leading to phenotypic
heterogeneity [131] could explain the study-dependent varying
sensitivity of HUVEC to high CBD concentrations.
As stromal cells in the tumour microenvironment are targets for
therapeutic intervention, the role of immune cells as an important
part of the tumour stroma has been brought into focus in recent
years in relation to cannabinoid effects on cancer progression
(Fig. 3). In this context, single-cell transcriptome proling revealed
that THC signicantly affects transcriptomic subclusters in immune
cell types [132]. A recent study showed inhibition of Kras-activated
signalling pathways by p21 activated kinase 1 (PAK1) after treatment
of cells with CBD and THC. Cannabinoids were found to reduce
expression of programmed death ligand 1 (PD-L1) via decreasing
PAK1 activity, thereby enhancing immune checkpoint blockade of
pancreatic cancer cells [133]. For other peripheral cells, such as
neutrophil granulocytes, their ability to produce proangiogenic
factors such as VEGF after exposure to LPS was shown to be blocked
by treatment with CB
(ACEA) and CB
agonists (JWH-133), which in
turn led to decreased tumour growth via inhibition of angiogenesis
Regarding the effect of ECs, one study investigated the
inuence of 2-AG on different subpopulations of immune cells
involved in the progression of pancreatic ductal adenocarcinoma
using an orthotopic mouse model. Here, 2-AG increased the
proportion of CD83
, CD86
cells in CD11C
populations in the spleen of mice [134]. Dendritic cells promoted
to maturation by 2-AG exhibited higher expression of proin-
ammatory cytokines (IL-6, IL-12, interferon-α) mediated by
activation of the CB
receptor and subsequent upregulation of
the phosphorylated form of signal transducer and activator of
transcription 6 (STAT6). A concomitant activation of T cells in the
spleen was not observed. On the other hand, in spleen and
tumour tissue of mice, 2-AG also induced the proliferation of
myeloid-derived suppressor cells, which are known to suppress
the T cell response thereby promoting an immunosuppressive
microenvironment. Nevertheless, the antiproliferative effect of
2-AG on cancer cells ultimately led to an overall tumour-regressive
effect in vivo [134].
In vivo experiments with murine melanoma models indicated that
WIN 55,212-2 inhibited cancer growth more efciently in immuno-
competent compared to immunodecient mice [44]. A further in vivo
study reported reduced inltration of macrophages and neutrophils
into experimental skin tumours after treatment of mice with THC
[135]. As THC did not inhibit cancer cell proliferation in this work, the
authors concluded that THC causes inhibition of cancer growth
in vivo due to its cannabinoid receptor-activating properties in the
tumour microenvironment and not in the tumour itself. In a further
study the in vitro interaction of lung cancer cells with lymphokine-
activated killer cells was addressed [136]. Here, prior treatment of
cancer cells with CBD, THC and Met-AEA led to increased killer cell-
mediated lysis of lung cancer cells. As a cause, an ICAM-1
upregulation on tumour cells induced by the tested cannabinoids
could be identied with the consequence of an increased cross-
linking with the lymphocyte function-associated antigen-1 (LFA-1) on
the surface of natural killer cells [136]. Yet another work showed that
inhibition of endogenous AEA degradation enhanced interaction
with the tumour immune system. Accordingly, the FAAH inhibitor
B. Hinz and R. Ramer
British Journal of Cancer
URB597 suppressed the shedding of the proteins major histocompat-
ibility complex class I polypeptide-related sequence A (MICA) and B
(MICB) on the surface of human HCC cells [137]. Since MICA/B
proteins are recognised by cytotoxic lymphocytes expressing the
natural killer group 2D (NKG2D) receptor and tumour cells are
subsequently eliminated, preventing the shedding of MICA/B proteins
can improve antitumour immunity. In the paper presented here, the
effect was indirect, and was shown to be due to increased expression
of TIMP-3. The authors further found AEA, 2-AG and the CB
agonist AM1241 to enhance TIMP-3 expression accompanied be a
reduction of MICA/B shedding. Noteworthy, the TIMP-3 inducing and
inhibitory action on MICA/B shedding upon treatment with URB597
was partially prevented by the CB
receptor antagonist rimonabant.
In contrast to these positive effects, however, another early study
on this topic reported that THC increases breast cancer growth and
promotes tumour cell spread by inhibiting the antitumour immune
response via enhancement of Th2-associated cytokines [138].
However, the study design used here was to exclude the direct
growth inhibitory effect of cannabinoids on tumour cells in order to
focus on the cannabinoid effect on the immune system. For this
reason, tumour cells whose expression of cannabinoid receptors was
low to undetectable were used in the experiments. This report was
consistent with another, even earlier study that had observed
tumour growth accelerating effects of THC based on reduced
tumour immunogenicity [139]. A recent publication further reported
that as a result of CB
receptor-induced microglial M2 polarisation,
conditioned media of cannabinoid-treated microglial cells increased
rather than inhibited the angiogenic capacities of human brain
microvascular endothelial cells [140]. The M2 polarisation induced
by JWH-133 was demonstrated here by mRNA analyses showing
increased levels of the M2 subtype markers CD206, arginase-1 (Arg1)
and the chitinase-like protein Ym1 and decreased levels of the
M1 subtype markers CD68, CD86 and inducible nitric oxide synthase
(iNOS). M2 polarisation also proved to be the mechanism by which
the CB
agonist JHW-133 attenuated its own tumour-regressive
effect in nude mice with intracranial glioma xenografts, which was
only fully expressed by knockdown of cytochrome P450 2J2
(CYP2J2), which mediates the proangiogenic effect [140]. As a
functional mediator of CB
receptor-dependent CYP2J2-induced M2
polarisation, the authors pointed to increased synthesis of 11,12-
epoxyeicosatrienoic acid (11,12-EET) by CYP2J2.
Although some newly approved anticancer drugs are also used as
monotherapy for certain indications, it seems more likely that
cannabinoid compounds will be used as a combination and add-on
option with currently employed cytostatics, assuming successful
clinical trials. Against this background, THC and CBD, which are
currently being tested in some studies as combination, have been
preclinically shown to enhance the effect of various cytostatics, such
as for vinca alkaloids, cytarabine, doxorubicin, mitoxantrone,
carmustine, temozolomide, bortezomib, carlzomib and cisplatin
(reviewed in ref. [111,141]). Thereby, combined administration of
CBD and temozolomide in patient-derived neurosphere cultures and
orthotopic mouse models was demonstrated to exert a signicant
synergistic effect in both reducing tumour size and prolonging
survival [85]. Of particular importance for the use of cannabinoids in
the treatment of glioblastoma is that the aforementioned booster
effect on temozolomide action was previously conrmed in
Fig. 3 Effects of cannabinoid compounds on tumour-immune interaction. Green arrows indicate the specic site of action of the indicated
cannabinoids. Black lines with circles at the end indicate receptor interactions. Black arrows indicate a functional or regulatory consequence of
cannabinoid treatment. The indication CB
means that the substances listed here act via both cannabinoid receptors. All abbreviations
are explained in the text.
B. Hinz and R. Ramer
British Journal of Cancer
elaborate in vivo mouse models [59]. In glioblastoma cells, CBD has
also been shown to enhance the effect of cisplatin [142]. Recently, a
synergistic effect was also conrmed for the tumour regressive
effect of CBD and cisplatin in a murine model of squamous cell
carcinoma of the head and neck as well as for the in vitro
cytotoxicity of CBD in combination with cisplatin, 5-uorouracil or
paclitaxel on human squamous cell carcinoma cells of the head and
neck [52]. However, the mechanisms of these synergies are not yet
fully understood. In this context, one study has shown that
cannabinoid-mediated enhancement of the effect of vinblastine in
resistant leukaemia cells was accompanied by THC- and CBD-
induced downregulation of P-glycoprotein [143], while the syner-
gistic cannabinoid effect over mitoxantrone in embryonic broblasts
occurred via inhibition of ATP-binding cassette transporters (ABC)G2
[144]. Another study focusing on the effect of THC on the
sensitisation of leukaemia cells to treatment with cytarabine,
doxorubicin and vincristine showed reduced p42/44 MAPK activity
as the underlying mechanism of THC-induced enhancement of
therespectivecytostaticeffect[145]. In addition, a number of
mechanistic studies have found a CBD-mediated increase in tumour
cell susceptibility to the proteasome inhibitor bortezomib [146,147],
doxorubicin [148,149] as well as temozolomide and carmustine
[148]. There has also been a report that CBD increases the uptake
into and toxicity on glioma cells of doxorubicin, temozolomide and
carmustine via an increase in TRPV2 activity and associated
increased calcium inux [148], with these results also conrmed
for doxorubicin in triple negative breast cancer cells [149]. With
regard to the synergistic effect with bortezomib, it was also shown
that the combination of CBD and THC inhibits the expression of the
immunoproteasome subunit β5i in multiple myeloma cells [147]. In
addition, the synergistic effect of the combination of CBD and THC
should also be mentioned here, which for example induces
autophagy-dependent necrosis in multiple myeloma cells and
inhibits cellular migration by downregulating the expression of the
chemokine receptor CXCR4 and the plasma membrane glycoprotein
CD147 [147]. However, in contrast to the results of these studies,
which showed an enhancement of cytostatic effects when
combined with cannabinoid compounds, a recently published
paper did not nd a survival benet in the cannabinoid treatment
group in combination with cyclophosphamide in an in vivo
medulloblastoma model [84].
In addition, several studies suggest that cannabinoid treatment
causes glioma cells to become more sensitive to ionising radiation,
as shown for the combination of THC and CBD [60] and the
combination of CBD with heat shock inhibitors [150]. Increased
radiosensitivity was conrmed in CBD-treated glioma cells in
another study [151], with the same group later reporting the
underlying mechanism to involve inhibition of ATM kinase, a
serine/threonine protein kinase that is recruited and activated by
DNA double-strand breaks [152].
Given a considerable number of in vitro and animal studies showing
that cannabinoid compounds exert tumour growth inhibitory and
antimetastatic effects, cannabinoid compounds may represent a
useful additional therapeutic option to currently used cytostatic
drugs. This view is also supported by studies indicating a synergistic
effect of cannabinoids in combination with currently used
chemotherapeutic agents and other therapeutic options. Further-
more, data increasingly suggest that cannabinoids may additionally
function as antimetastatic and anti-angiogenic tumour therapy and
support the immune system in its defence against tumours.
In addition to the partially divergent preclinical studies already
mentioned, some epidemiological studies should also be included in
the critical assessment of the potential use of cannabinoids as
systemic therapy options. Thus, a recently published prospective
observational study showed that cannabis use signicantly shortens
the time to tumour progression and overall survival of cancer
patients [153]. This study illustrates that cannabis use, in this case via
modulation of the immune system, can lead to negative and thus
life-threatening effects for cancer patients. In addition, a retro-
spective observational study showed a reduction in the response
rate to nivolumab, although the addition of cannabis here had no
effect on progression-free survival or overall survival [154]. However,
it is worth noting that the administration of cannabinoids in these
studies conducted in relatively small patient groups was through the
consumption of cannabis oil or smoked/inhaled cannabis, and in
many cases the cannabis products were also changed during the
course of the study.
On the other hand, these data are counterbalanced by an
overwhelming number of studies that clearly show that activation of
the EC system is an important factor in tumour defence and thus
could serve as a promising target for pharmacological anticancer
interventions. In this context, a collection of case reports involving
119 patients also presented impressive examples of breast cancer
and glioma patients treated with pharmaceutical-grade synthetic
CBD, demonstrating a reduction in circulating tumour cells or a
reduction in tumour size by repeat scans [155]. Furthermore, the
results of the well-conducted but very small randomised, placebo-
controlled phase 1b trial in patients with recurrent glioblastoma
multiforme mentioned at the beginning of this article and the
higher survival rate shown here in patients taking nabiximols instead
of placebo in combination with dose-intense temozolomide [43]
provide the rationale for larger and thus adequately powered
randomised placebo-controlled trials.
In summary, the property of cannabinoids, in particular, to induce
inhibition of tumour growth and spread at multiple levels of tumour
progression argues for the use of these substances as an add-on
option in tumour treatment. However, it should also be noted that
research into the efcacy, dosage and drug safety of cannabinoids in
tumour therapy still has a long way to go, especially with regard to
clinical trials to be conducted, through which alone the benets and
advantages for cancer patients but also possible risks can be
Not applicable.
1. Touwn M. The religious and medicinal uses of Cannabis in China, India and
Tibet. J Psychoact Drugs. 1981;13:2334.
2. Zuardi AW. History of cannabis as a medicine: a review. Braz J Psychiatry.
3. Liesowska, A Iconic 2,500 year old Siberian princess died from breast cancer,
reveals MRI scan
scan/ 2014. Accessed on 5 Dec 2021.
4. Bai Y, Jiang M, Xie T, Jiang C, Gu M, Zhou X, et al. Archaeobotanical evidence of
the use of medicinal cannabis in a secular context unearthed from south China.
J Ethnopharmacol. 2021;275:114114.
5. OShaughnessy, WB. On the preparations of the Indian hemp or Gunjah,
Transactions of the Medical and Physical Society of Bengal 18381840, p. 42161.
Reprint in: Mikuriya, TH (Ed.): Marijuana Medical papers 18391972, Medi-Comp
Press, Oakland, 1973.
6. Wood TB, Spivey WTN, Eastereld TH. Cannabinol. Part I. J Chem Soc, Trans.
7. Mechoulam R, Shvo Y, Hashish I. The structure of cannabidiol. Tetrahedron.
8. Gaoni Y, Mechoulam R. Isolation, structure, and partial synthesis of an active
constituent of hashish. J Am Chem Soc. 1964;86:16467.
9. Mechoulam R, Gaoni Y. The absolute conguration of delta-1-tetra-
hydrocannabinol, the major active constituent of hashish. Tetrahedron Lett.
10. Mechoulam R, Shani A, Edery H, Grunfeld Y. Chemical basis of hashish activity.
Science. 1970;169:6112.
B. Hinz and R. Ramer
British Journal of Cancer
11. Matsuda LA, Lolait SJ, Brownstein MJ, Young AC, Bonner T. Structure of a can-
nabinoid receptor and functional expression of the cloned cDNA. Nature.
12. Munro S, Thomas KL, Abu-Shaar M. Molecular characterization of a peripheral
receptor for cannabinoids. Nature. 1993;365:6165.
13. Devane WA, Hanus L, Breuer A, Pertwee RG, Stevenson LA, Grifn G, et al. Iso-
lation and structure of a brain constituent that binds to the cannabinoid
receptor. Science. 1992;258:19469.
14. Mechoulam R, Ben-Shabat S, Hanus L, Ligumsky M, Kaminski NE, Schatz AR, et al.
Identication of an endogenous 2-monoglyceride, present in canine gut, that
binds to cannabinoid receptors. Biochem Pharmacol. 1995;50:8390.
15. Schwarz R, Ramer R, Hinz B. Targeting the endocannabinoid system as a
potential anticancer approach. Drug Metab Rev. 2018;50:2653.
16. Zygmunt PM, Petersson J, Andersson DA, Chuang H, Sørgård M, Di Marzo V,
et al. Vanilloid receptors on sensory nerves mediate the vasodilator action of
anandamide. Nature. 1999;400:4527.
17. Bisogno T, Hanus L, De Petrocellis L, Tchilibon S, Ponde DE, Brandi I, et al.
Molecular targets for cannabidiol and its synthetic analogues: effect on vanilloid
VR1 receptors and on the cellular uptake and enzymatic hydrolysis of ana-
ndamide. Br J Pharmacol. 2001;134:84552.
18. Pertwee RG. The diverse CB
and CB
receptor pharmacology of three plant
cannabinoids: Δ
-tetrahydrocannabinol, cannabidiol and Δ
hydrocannabivarin. Br J Pharmacol. 2008;153:199215.
19. Ryberg E, Larsson N, Sjögren S, Hjorth S, Hermansson NO, Leonova J, et al. The
orphan receptor GPR55 is a novel cannabinoid receptor. Br J Pharmacol.
20. Rosenthaler S, Pöhn B, Kolmanz C, Huu CN, Krewenka C, Huber A, et al. Differ-
ences in receptor binding afnity of several phytocannabinoids do not explain
their effects on neural cell cultures. Neurotoxicol Teratol. 2014;46:4956. Erra-
tum in: Neurotoxicol. Teratol. 2016;54:8993.
21. OSullivan SE, Sun Y, Bennett AJ, Randall MD, Kendall DA. Time-dependent
vascular actions of cannabidiol in the rat aorta. Eur J Pharmacol.
22. Di Marzo V. The endocannabinoid system: its general strategy of action, tools for
its pharmacological manipulation and potential therapeutic exploitation. Phar-
macol Res. 2009;60:7784.
23. Deutsch DG, Chin SA. Enzymatic synthesis and degradation of anandamide, a
cannabinoid receptor agonist. Biochem Pharmacol. 1993;46:7916.
24. Blankman JL, Simon GM, Cravatt BF. A comprehensive prole of brain enzymes
that hydrolyze the endocannabinoid 2-arachidonoylglycerol. Chem Biol.
25. Di Marzo V. New approaches and challenges to targeting the endocannabinoid
system. Nat Rev Drug Discov. 2018;17:62339. Erratum in: Corrigendum: New
approaches and challenges to targeting the endocannabinoid system. Nat. Rev.
Drug Discov. 2018;17:688.
26. Munson AE, Harris LS, Friedman MA, Dewey WL, Carchman RA. Antineoplastic
activity of cannabinoids. J Natl Cancer Inst. 1975;55:597602.
27. Galve-Roperh I, Sánchez C, Cortés ML, Gómez del Pulgar T, Izquierdo M, Guzmán M.
Anti-tumoural action of cannabinoids, involvement of sustained ceramide accu-
mulation and extracellular signal-regulated kinase activation. Nat Med. 2000;6:3139.
28. Sánchez C, de Ceballos ML, Gomez del Pulgar T, Rueda D, Corbacho C, Velasco
G, et al. Inhibition of glioma growth in vivo by selective activation of the CB
cannabinoid receptor. Cancer Res. 2001;61:57849.
29. Long JZ, Li W, Booker L, Burston JJ, Kinsey SG, Schlosburg JE, et al. Selective
blockade of 2-arachidonoylglycerol hydrolysis produces cannabinoid behavioral
effects. Nat Chem Biol. 2009;5:3744.
30. Nomura DK, Long JZ, Niessen S, Hoover HS, Ng S-W, Cravatt BF. Mono-
acylglycerol lipase regulates a fatty acid network that promotes cancer patho-
genesis. Cell. 2010;140:4961.
31. Nomura DK, Lombardi DP, Chang JW, Niessen S, Ward AM, Long JZ, et al.
Monoacylglycerol lipase exerts dual control over endocannabinoid and fatty
acid pathways to support prostate cancer. Chem Biol. 2011;18:84656.
32. Prüser JL, Ramer R, Wittig F, Ivanov I, Merkord J, Hinz B. The monoacylglycerol
lipase inhibitor JZL184 inhibits lung cancer cell invasion and metastasis via the
cannabinoid receptor. Mol Cancer Ther. 2021;20:787802.
33. Liu R, Wang X, Curtiss C, Landas S, Rong R, Sheikh MS, et al. Monoglyceride
lipase gene knockout in mice leads to increased incidence of lung adeno-
carcinoma. Cell Death Dis. 2018;9:36.
34. Ligresti A, Bisogno T, Matias I, De Petrocellis L, Cascio MG, Cosenza V, et al.
Possible endocannabinoid control of colorectal cancer growth. Gastro-
enterology. 2003;125:67787.
35. Bifulco M, Laezza C, Valenti M, Ligresti A, Portella G, Di Marzo V. A new strategy
to block tumour growth by inhibiting endocannabinoid inactivation. FASEB J.
36. Winkler K, Ramer R, Dithmer S, Ivanov I, Merkord J, Hinz B. Fatty acid amide
hydrolase inhibitors confer anti-invasive and antimetastatic effects on lung
cancer cells. Oncotarget. 2016;7:1504764.
37. Gertsch J, Leonti M, Raduner S, Racz I, Chen JZ, Xie XQ, et al. Beta-caryophyllene
is a dietary cannabinoid. Proc Natl Acad Sci USA. 2008;105:9099104.
38. Dahham SS, Tabana YM, Iqbal MA, Ahamed MB, Ezzat MO, Majid AS, et al. The
anticancer, antioxidant and antimicrobial properties of the sesquiterpene β-
caryophyllene from the essential oil of aquilaria crassna. Molecules.
39. Irrera N, DAscola A, Pallio G, Bitto A, Mannino F, Arcoraci V, et al. β-
Caryophyllene inhibits cell proliferation through a direct modulation of CB
receptors in glioblastoma cells. Cancers (Basel). 2020;12:1038.
40. Di Giacomo S, Di Sotto A, Mazzanti G, Wink M. Chemosensitizing properties of β-
caryophyllene and β-caryophyllene oxide in combination with doxorubicin in
human cancer cells. Anticancer Res. 2017;37:11916.
41. Di Giacomo S, Briz O, Monte MJ, Sanchez-Vicente L, Abete L, Lozano E, et al.
Chemosensitization of hepatocellular carcinoma cells to sorafenib by β-
caryophyllene oxide-induced inhibition of ABC export pumps. Arch Toxicol.
42. Guzmán M, Duarte MJ, Blázquez C, Ravina J, Rosa MC, Galve-Roperh I, et al. A
pilot clinical study of Δ
-tetrahydrocannabinol in patients with recurrent glio-
blastoma multiforme. Br J Cancer. 2006;95:197203.
43. Twelves C, Sabel M, Checketts D, Miller S, Tayo B, Jove M, et al. GWCA1208 study
group. A phase 1b randomised, placebo-controlled trial of nabiximols canna-
binoid oromucosal spray with temozolomide in patients with recurrent glio-
blastoma. Br J Cancer. 2021;124:137987.
44. Blázquez C, Carracedo A, Barrado L, Real PJ, Fernández-Luna JL, Velasco G, et al.
Cannabinoid receptors as novel targets for the treatment of melanoma. FASEB J.
45. Caffarel MM, Andradas C, Mira E, Pérez-Gómez E, Cerutti C, Moreno-Bueno G,
et al. Cannabinoids reduce ErbB2-driven breast cancer progression through Akt
inhibition. Mol Cancer. 2010;9:196.
46. Xian XS, Park H, Cho YK, Lee IS, Kim SW, Choi MG, et al. Effect of a synthetic
cannabinoid agonist on the proliferation and invasion of gastric cancer cells. J
Cell Biochem. 2010;110:32132.
47. Boyacıoğlu Ö, Bilgiç E, Varan C, Bilensoy E, Nemutlu E, Sevim D, et al. ACPA
decreases non-small cell lung cancer line growth through Akt/PI3K and JNK
pathways in vitro. Cell Death Dis. 2021;12:56.
48. Rao M, Chen D, Zhan P, Jiang J. MDA19 a novel CB
agonist inhibits hepato-
cellular carcinoma partly through inactivation of AKT signaling pathway. Biol
Direct. 2019;14:9.
49. Caffarel MM, Sarrió D, Palacios J, Guzmán M, Sánchez C. Δ
hydrocannabinol inhibits cell cycle progression in human breast cancer cells
through Cdc2 regulation. Cancer Res. 2006;66:661521.
50. Laezza C, Pisanti S, Crescenzi E, Bifulco M. Anandamide inhibits Cdk2 and
activates Chk1 leading to cell cycle arrest in human breast cancer cells. FEBS
Lett. 2006;580:607682.
51. Roberto D, Klotz LH, Venkateswaran V. Cannabinoid WIN 55,212-2 induces cell
cycle arrest and apoptosis, and inhibits proliferation, migration, invasion, and
tumor growth in prostate cancer in a cannabinoid-receptor 2 dependent
manner. Prostate. 2019;79:1519.
52. Go YY, Kim SR, Kim DY, Chae SW, Song JJ. Cannabidiol enhances cytotoxicity of
anti-cancer drugs in human head and neck squamous cell carcinoma. Sci Rep.
53. Zhang X, Qin Y, Pan Z, Li M, Liu X, Chen X, et al. Cannabidiol induces cell cycle
arrest and cell apoptosis in human gastric cancer SGC-7901 cells. Biomolecules.
54. Massi P, Valenti M, Vaccani A, Gasperi V, Perletti G, Marras E, et al.
5-Lipoxygenase and anandamide hydrolase (FAAH) mediate the antitumor
activity of cannabidiol, a non-psychoactive cannabinoid. J Neurochem.
55. Liu C, Sadat SH, Ebisumoto K, Sakai A, Panuganti BA, Ren S, et al. Cannabinoids
promote progression of HPV-positive head and neck squamous cell carcinoma
via p38 MAPK activation. Clin Cancer Res. 2020;26:2693703.
56. Hart S, Fischer OM, Ullrich A. Cannabinoids induce cancer cell proliferation via
tumor necrosis factor alpha-converting enzyme (TACE/ADAM17)-mediated
transactivation of the epidermal growth factor receptor. Cancer Res.
57. Miyato H, Kitayama J, Yamashita H, Souma D, Asakage M, Yamada J, et al.
Pharmacological synergism between cannabinoids and paclitaxel in gastric
cancer cell lines. J Surg Res. 2009;155:407.
58. Martínez-Martínez E, Martín-Ruiz A, Martín P, Calvo V, Provencio M, García JM.
cannabinoid receptor activation promotes colon cancer progression via
AKT/GSK3βsignaling pathway. Oncotarget. 2016;7:6878191.
B. Hinz and R. Ramer
British Journal of Cancer
59. Torres S, Lorente M, Rodríguez-Fornés F, Hernández-Tiedra S, Salazar M, García-
Taboada E, et al. A combined preclinical therapy of cannabinoids and temo-
zolomide against glioma. Mol Cancer Ther. 2011;10:90103.
60. Scott KA, Dalgleish AG, Liu WM. The combination of cannabidiol and Δ
hydrocannabinol enhances the anticancer effects of radiation in an orthotopic
murine glioma model. Mol Cancer Ther. 2014;13:295567.
61. Fowler CJ. Delta
-tetrahydrocannabinol and cannabidiol as potential curative
agents for cancer: a critical examination of the preclinical literature. Clin Phar-
macol Ther. 2015;97:58796.
62. Gómez del Pulgar T, Velasco G, Sánchez C, Haro A, Guzmán M. De novo-
synthesized ceramide is involved in cannabinoid-induced apoptosis. Biochem J.
63. Salazar M, Carracedo A, Salanueva IJ, Hernández-Tiedra S, Lorente M, Egia A,
et al. Cannabinoid action induces autophagy-mediated cell death through sti-
mulation of ER stress in human glioma cells. J Clin Invest. 2009;119:135972.
64. Carracedo A, Gironella M, Lorente M, Garcia S, Guzmán M, Velasco G, et al.
Cannabinoids induce apoptosis of pancreatic tumour cells via endoplasmic
reticulum stress-related genes. Cancer Res. 2006;66:674855.
65. Oesch S, Walter D, Wachtel M, Pretre K, Salazar M, Guzmán M, et al. Cannabinoid
receptor 1 is a potential drug target for treatment of translocation-positive
rhabdomyosarcoma. Mol Cancer Ther. 2009;8:183845.
66. Ramer R, Weinzierl U, Schwind B, Brune K, Hinz B. Ceramide is involved in
R(+)-methanandamide-induced cyclooxygenase-2 expression in human neu-
roglioma cells. Mol Pharmacol. 2003;64:118998.
67. Hinz B, Ramer R, Eichele K, Weinzierl U, Brune K. Up-regulation of
cyclooxygenase-2 expression is involved in R(+)-methanandamide-induced
apoptotic death of human neuroglioma cells. Mol Pharmacol. 2004;66:164351.
68. Eichele K, Ramer R, Hinz B. R(+)-methanandamide-induced apoptosis of human
cervical carcinoma cells involves a cyclooxygenase-2-dependent pathway.
Pharm Res. 2009;26:34655.
69. Eichele K, Ramer R, Hinz B. Decisive role of cyclooxygenase-2 and lipocalin-type
prostaglandin D synthase in chemotherapeutics-induced apoptosis of human
cervical carcinoma cells. Oncogene. 2008;27:303244.
70. Ramer R, Heinemann K, Merkord J, Rohde H, Salamon A, Linnebacher M, et al.
COX-2 and PPAR-γconfer cannabidiol-induced apoptosis of human lung cancer
cells. Mol Cancer Ther. 2013;12:6982.
71. Maccarrone M, Lorenzon T, Bari M, Melino G, Finazzi-Agro A. Anandamide
induces apoptosis in human cells via vanilloid receptors. Evidence for a pro-
tective role of cannabinoid receptors. J Biol Chem. 2000;275:3193845.
72. Soliman E, Van Dross R. Anandamide-induced endoplasmic reticulum stress and
apoptosis are mediated by oxidative stress in non-melanoma skin cancer: Receptor-
independent endocannabinoid signaling. Mol Carcinog. 2016;55:180721.
73. Huang L, Ramirez JC, Frampton GA, Golden LE, Quinn MA, Pae HY, et al. Ana-
ndamide exerts its antiproliferative actions on cholangiocarcinoma by activation
of the GPR55 receptor. Lab Invest. 2011;91:100717.
74. Yamada T, Ueda T, Shibata Y, Ikegami Y, Saito M, Ishida Y, et al. TRPV2 activation
induces apoptotic cell death in human T24 bladder cancer cells: a potential
therapeutic target for bladder cancer. Urology. 2010;76:509.e1509.e7.
75. Shrivastava A, Kuzontkoski PM, Groopman JE, Prasad A. Cannabidiol induces
programmed cell death in breast cancer cells by coordinating the cross-talk
between apoptosis and autophagy. Mol Cancer Ther. 2011;10:116172.
76. McKallip RJ, Jia W, Schlomer J, Warren JW, Nagarkatti PS, Nagarkatti M.
Cannabidiol-induced apoptosis in human leukemia cells: A novel role of can-
nabidiol in the regulation of p22phox and Nox4 expression. Mol Pharmacol.
77. Jeong S, Jo MJ, Yun HK, Kim DY, Kim BR, Kim JL, et al. Cannabidiol promotes
apoptosis via regulation of XIAP/Smac in gastric cancer. Cell Death Dis.
78. Olivas-Aguirre M, Torres-López L, Valle-Reyes JS, Hernández-Cruz A, Pottosin I,
Dobrovinskaya O. Cannabidiol directly targets mitochondria and disturbs cal-
cium homeostasis in acute lymphoblastic leukemia. Cell Death Dis. 2019;10:779.
79. Vara D, Salazar M, Olea-Herrero N, Guzmán M, Velasco G, Díaz-Laviada I. Anti-
tumoural action of cannabinoids on hepatocellular carcinoma, role of AMPK-
dependent activation of autophagy. Cell Death Differ. 2011;18:1099111.
80. Armstrong JL, Hill DS, McKee CS, Hernandez-Tiedra S, Lorente M, Lopez-Valero I,
et al. Exploiting cannabinoid-induced cytotoxic autophagy to drive melanoma
cell death. J Invest Dermatol. 2015;135:162937.
81. Ellert-Miklaszewska A, Ciechomska IA, Kaminska B. Synthetic cannabinoids
induce autophagy and mitochondrial apoptotic pathways in human glio-
blastoma cells independently of deciency in TP53 or PTEN tumour suppressors.
Cancers (Basel). 2021;13:419.
82. Ivanov VN, Grabham PW, Wu CC, Hei TK. Inhibition of autophagic ux differently
modulates cannabidiol-induced death in 2D and 3D glioblastoma cell cultures.
Sci Rep. 2020;10:2687.
83. Nabissi M, Morelli MB, Amantini C, Liberati S, Santoni M, Ricci-Vitiani L, et al.
Cannabidiol stimulates Aml-1a-dependent glial differentiation and inhibits
glioma stem-like cells proliferation by inducing autophagy in a TRPV2-
dependent manner. Int J Cancer. 2015;137:185569.
84. Andradas C, Byrne J, Kuchibhotla M, Ancliffe M, Jones AC, Carline B, et al.
Assessment of cannabidiol and Δ
-tetrahydrocannabiol in mouse models of
medulloblastoma and ependymoma. Cancers (Basel). 2021;13:330.
85. Huang T, Xu T, Wang Y, Zhou Y, Yu D, Wang Z, et al. Cannabidiol inhibits human
glioma by induction of lethal mitophagy through activating TRPV4. Autophagy.
86. Meyer N, Zielke S, Michaelis JB, Linder B, Warnsmann V, Rakel S, et al. AT 101
induces early mitochondrial dysfunction and HMOX1 (heme oxygenase 1) to
trigger mitophagic cell death in glioma cells. Autophagy. 2018;14:1693709.
87. Nithipatikom K, Endsley MP, Isbell MA, Falck JR, Iwamoto Y, Hillard CJ, et al. 2-
arachidonoylglycerol, a novel inhibitor of androgen-independent prostate
cancer cell invasion. Cancer Res. 2004;64:882630.
88. Ma C, Wu TT, Jiang PC, Li ZQ, Chen XJ, Fu K, et al. Anti-carcinogenic activity of
anandamide on human glioma in vitro and in vivo. Mol Med Rep.
89. Ramer R, Hinz B. Inhibition of cancer cell invasion by cannabinoids via increased
expression of tissue inhibitor of matrix metalloproteinases-1. J Natl Cancer Inst.
90. Ramer R, Merkord J, Rohde H, Hinz B. Cannabidiol inhibits cancer cell invasion
via upregulation of tissue inhibitor of matrix metalloproteinases-1. Biochem
Pharmacol. 2010;79:95566.
91. Stamenkovic I. Matrix metalloproteinases in tumor invasion and metastasis.
Semin Cancer Biol. 2000;10:41533.
92. Cruz-Munoz W, Khokha R. The role of tissue inhibitors of metalloproteinases in
tumorigenesis and metastasis. Crit Rev Clin Lab Sci. 2008;45:291338.
93. Ramer R, Bublitz K, Freimuth N, Merkord J, Rohde H, Haustein M, et al. Canna-
bidiol inhibits lung cancer cell invasion and metastasis via intercellular adhesion
molecule-1. FASEB J. 2012;26:153548.
94. Blázquez C, Salazar M, Carracedo A, Lorente M, Egia A, González-Feria L, et al.
Cannabinoids inhibit glioma cell invasion by down-regulating matrix
metalloproteinase-2 expression. Cancer Res. 2008;68:194552.
95. Pourkhalili N, Ghahremani MH, Farsandaj N, Tavajohi S, Majdzadeh M, Parsa M,
et al. Evaluation of anti-invasion effect of cannabinoids on human hepato-
carcinoma cells. Toxicol Mech Methods. 2013;23:1206.
96. Notaro A, Emanuele S, Geraci F, DAnneo A, Lauricella M, Calvaruso G, et al.
WIN55212-2-induced expression of mir-29b1 favours the suppression of
osteosarcoma cell migration in a SPARC-independent manner. Int J Mol Sci.
97. Pietrovito L, Iozzo M, Bacci M, Giannoni E, Chiarugi P. Treatment with canna-
binoids as a promising approach for impairing broblast activation and prostate
cancer progression. Int J Mol Sci. 2020;21:787.
98. McAllister SD, Christian RT, Horowitz MP, Garcia A, Desprez PY. Cannabidiol as a
novel inhibitor of Id-1 gene expression in aggressive breast cancer cells. Mol
Cancer Ther. 2007;6:29217.
99. Soroceanu L, Murase R, Limbad C, Singer E, Allison J, Adrados I, et al. Id-1 is a key
transcriptional regulator of glioblastoma aggressiveness and a novel ther-
apeutic target. Cancer Res. 2013;73:155969.
100. Murase R, Sumida T, Kawamura R, Onishi-Ishikawa A, Hamakawa H, McAllister
SD, et al. Suppression of invasion and metastasis in aggressive salivary cancer
cells through targeted inhibition of ID1 gene expression. Cancer Lett.
101. McAllister SD, Murase R, Christian RT, Lau D, Zielinski AJ, Allison J, et al. Pathways
mediating the effects of cannabidiol on the reduction of breast cancer cell
proliferation invasion and metastasis. Breast Cancer Res Treat. 2011;129:3747.
Erratum in: Breast Cancer Res Treat. 2012;133:4014.
102. Murase R, Kawamura R, Singer E, Pakdel A, Sarma P, Judkins J, et al. Targeting
multiple cannabinoid anti-tumour pathways with a resorcinol derivative leads to
inhibition of advanced stages of breast cancer. Br J Pharmacol.
103. Ramer R, Rohde A, Merkord J, Rohde H, Hinz B. Decrease of plasminogen acti-
vator inhibitor-1 may contribute to the anti-invasive action of cannabidiol on
human lung cancer cells. Pharm Res. 2010;27:216274.
104. Leelawat S, Leelawat K, Narong S, Matangkasombut O. The dual effects of Δ
tetrahydrocannabinol on cholangiocarcinoma cells, anti-invasion activity at low
concentration and apoptosis induction at high concentration. Cancer Invest.
105. Anis O, Vinayaka AC, Shalev N, Namdar D, Nadarajan S, Anil SM, et al. Cannabis-
derived compounds cannabichromene and Δ
-tetrahydrocannabinol interact and
exhibit cytotoxic activity against urothelial cell carcinoma correlated with inhibition
of cell migration and cytoskeleton organization. Molecules. 2021;26:465.
B. Hinz and R. Ramer
British Journal of Cancer
106. Xu S, Ma H, Bo Y, Shao M. The oncogenic role of CB
in the progression of non-
small-cell lung cancer. Biomed Pharmacother. 2019;117:109080.
107. Coke CJ, Scarlett KA, Chetram MA, Jones KJ, Sandifer BJ, Davis AS, et al.
Simultaneous activation of induced heterodimerization between CXCR4 che-
mokine receptor and cannabinoid receptor 2 (CB
) reveals a mechanism for
regulation of tumour progression. J Biol Chem. 2016;291:999110005.
108. Laezza C, DAlessandro A, Paladino S, Maltano AM, Proto MC, Gazzerro P, et al.
Anandamide inhibits the Wnt/β-catenin signalling pathway in human breast
cancer MDA MB 231 cells. Eur J Cancer. 2012;48:311222.
109. García-Morales L, Castillo AM, Tapia Ramírez J, Zamudio-Meza H, Domínguez-Robles
MDC, Meza I. CBD reverts the mesenchymal invasive phenotype of breast cancer
cells induced by the inammatory cytokine IL-1β. Int J Mol Sci. 2020;21:2429.
110. Milian L, Mata M, Alcacer J, Oliver M, Sancho-Tello M, Martín de Llano JJ, et al.
Cannabinoid receptor expression in non-small cell lung cancer. Effectiveness of
tetrahydrocannabinol and cannabidiol inhibiting cell proliferation and
epithelial-mesenchymal transition in vitro. PLoS ONE. 2020;15:e0228909.
111. Ramer R, Hinz B. Cannabinoids as anticancer drugs. Adv Pharmacol.
112. Portella G, Laezza C, Laccetti P, De Petrocellis L, Di Marzo V, Bifulco M. Inhibitory
effects of cannabinoid CB
receptor stimulation on tumor growth and meta-
static spreading: actions on signals involved in angiogenesis and metastasis.
FASEB J. 2003;17:17713.
113. Preet A, Ganju RK, Groopman JE. Δ
-Tetrahydrocannabinol inhibits epithelial
growth factor-induced lung cancer cell migration in vitro as well as its growth
and metastasis in vivo. Oncogene. 2008;27:33946.
114. Qamri Z, Preet A, Nasser MW, Bass CE, Leone G, Barsky SH, et al. Synthetic
cannabinoid receptor agonists inhibit tumor growth and metastasis of breast
cancer. Mol Cancer Ther. 2009;8:311729.
115. Marino S, de Ridder D, Bishop RT, Renema N, Ponzetti M, Sophocleous A, et al.
Paradoxical effects of JZL184, an inhibitor of monoacylglycerol lipase, on bone
remodelling in healthy and cancer-bearing mice. EBioMedicine. 2019;44:45266.
116. Hu WR, Lian YF, Peng LX, Lei JJ, Deng CC, Xu M, et al. Monoacylglycerol lipase
promotes metastases in nasopharyngeal carcinoma. Int J Clin Exp Pathol.
117. Ramer R, Hinz B. New insights into antimetastatic and antiangiogenic effects of
cannabinoids. Int Rev Cell Mol Biol. 2015;314:43116.
118. Casanova ML, Blázquez C, Martínez-Palacio J, Villanueva C, Fernández-Aceñero
MJ, Huffman JW, et al. Inhibition of skin tumour growth and angiogenesis
in vivo by activation of cannabinoid receptors. J Clin Invest. 2003;111:4350.
119. Blázquez C, Casanova ML, Planas A, Gómez Del Pulgar T, Villanueva C,
Fernández-Aceñero MJ, et al. Inhibition of tumour angiogenesis by cannabi-
noids. FASEB J. 2003;17:52931.
120. Blázquez C, González-Feria L, Alvarez L, Haro A, Casanova ML, Guzmán M.
Cannabinoids inhibit the vascular endothelial growth factor pathway in gliomas.
Cancer Res. 2004;64:561723.
121. Solinas M, Massi P, Cantelmo AR, Cattaneo MG, Cammarota R, Bartolini D, et al.
Cannabidiol inhibits angiogenesis by multiple mechanisms. Br J Pharmacol.
122. Thapa D, Lee JS, Heo SW, Lee YR, Kang KW, Kwak MK, et al. Novel hexahy-
drocannabinol analogs as potential anti-cancer agents inhibit cell proliferation
and tumour angiogenesis. Eur J Pharmacol. 2011;650:6471.
123. Picardi P, Ciaglia E, Proto M, Pisanti S. Anandamide inhibits breast tumour-
induced angiogenesis. Transl Med UniSa. 2014;10:812.
124. Ramer R, Fischer S, Haustein M, Manda K, Hinz B. Cannabinoids inhibit angiogenic
capacities of endothelial cells via release of tissue inhibitor of matrix
metalloproteinases-1 from lung cancer cells. Biochem Pharmacol. 2014;91:20216.
125. Braile M, Cristinziano L, Marcella S, Varricchi G, Marone G, Modestino L, et al.
LPS-mediated neutrophil VEGF-A release is modulated by cannabinoid receptor
activation. J Leukoc Biol. 2021;109:62131.
126. Pisanti S, Picardi P, Prota L, Proto MC, Laezza C, McGuire PG, et al. Genetic and
pharmacologic inactivation of cannabinoid CB
receptor inhibits angiogenesis.
Blood. 2011;117:554150.
127. Pisanti S, Borselli C, Oliviero O, Laezza C, Gazzerro P, Bifulco M. Antiangiogenic
activity of the endocannabinoid anandamide, correlation to its tumour-
suppressor efcacy. J Cell Physiol. 2007;211:495503.
128. Kogan NM, Blázquez C, Alvarez L, Gallily R, Schlesinger M, Guzmán M, et al. A
cannabinoid quinone inhibits angiogenesis by targeting vascular endothelial
cells. Mol Pharmacol. 2006;70:519.
129. Hofmann NA, Barth S, Waldeck-Weiermair M, Klec C, Strunk D, Malli R, et al.
TRPV1 mediates cellular uptake of anandamide and thus promotes endothelial
cell proliferation and network-formation. Biol Open. 2014;3:116472.
130. Böckmann S, Hinz B. Cannabidiol promotes endothelial cell survival by heme
oxygenase-1-mediated autophagy. Cells. 2020;9:1703.
131. Aird WC. Endothelial cell heterogeneity. Cold Spring Harb Perspect Med 2012;2:
132. Hu Y, Ranganathan M, Shu C, Liang X, Ganesh S, Osafo-Addo A, et al.
Single-cell transcriptome mapping identies common and cell-type specic
genes affected by acute delta9-tetrahydrocannabinol in humans. Sci Rep.
133. Yang Y, Huynh N, Dumesny C, Wang K, He H, Nikfarjam M. Cannabinoids
inhibited pancreatic cancer via P-21 activated kinase 1 mediated pathway. Int J
Mol Sci. 2020;21:8035.
134. Qiu C, Yang L, Wang B, Cui L, Li C, Zhuo Y, et al. The role of
2-arachidonoylglycerol in the regulation of the tumour-immune microenviron-
ment in murine models of pancreatic cancer. Biomed Pharmacother.
135. Glodde N, Jakobs M, Bald T, Tüting T, Gaffal E. Differential role of cannabinoids
in the pathogenesis of skin cancer. Life Sci 2015;138:3540.
136. Haustein M, Ramer R, Linnebacher M, Manda K, Hinz B. Cannabinoids increase
lung cancer cell lysis by lymphokine-activated killer cells via upregulation of
ICAM-1. Biochem Pharmacol. 2014;92:31225.
137. Sekiba K, Otsuka M, Seimiya T, Tanaka E, Funato K, Miyakawa Y, et al. The fatty-
acid amide hydrolase inhibitor URB597 inhibits MICA/B shedding. Sci Rep.
138. McKallip RJ, Nagarkatti M, Nagarkatti PS. Δ-9-tetrahydrocannabinol enhances
breast cancer growth and metastasis by suppression of the antitumour immune
response. J Immunol. 2005;174:32819.
139. Zhu LX, Sharma S, Stolina M, Gardner B, Roth MD, Tashkin DP, et al. Δ-9-
tetrahydrocannabinol inhibits antitumour immunity by a CB
ated, cytokine-dependent pathway. J Immunol. 2000;165:37380.
140. Lei X, Chen X, Quan Y, Tao Y, Li J. Targeting CYP2J2 to enhance the anti-glioma
efcacy of cannabinoid receptor 2 stimulation by inhibiting the pro-
angiogenesis function of M2 microglia. Front Oncol. 2020;10:574277.
141. Hinz B, Ramer R. Anti-tumour actions of cannabinoids. Br J Pharmacol.
142. Deng L, Ng L, Ozawa T, Stella N. Quantitative analyses of synergistic responses
between cannabidiol and DNA-damaging agents on the proliferation and via-
bility of glioblastoma and neural progenitor cells in culture. J Pharmacol Exp
Ther. 2017;360:21524.
143. Holland ML, Panetta JA, Hoskins JM, Bebawy M, Roufogalis BD, Allen JD, et al.
The effects of cannabinoids on P-glycoprotein transport and expression in
multidrug resistant cells. Biochem Pharmacol. 2006;71:114654.
144. Holland ML, Lau DT, Allen JD, Arnold JC. The multidrug transporter ABCG2
(BCRP) is inhibited by plant-derived cannabinoids. Br J Pharmacol.
145. Liu WM, Scott KA, Shamash J, Joel S, Powles TB. Enhancing the in vitro cytotoxic
activity of Δ
-tetrahydrocannabinol in leukemic cells through a combinatorial
approach. Leuk Lymphoma. 2008;49:18009.
146. Morelli MB, Ofdani M, Alesiani F, Discepoli G, Liberati S, Olivieri A, et al. The
effects of cannabidiol and its synergism with bortezomib in multiple myeloma
cell lines. A role for transient receptor potential vanilloid type-2. Int J Cancer.
147. Nabissi M, Morelli MB, Ofdani M, Amantini C, Gentili S, Soriani A, et al. Can-
nabinoids synergize with carlzomib, reducing multiple myeloma cells viability
and migration. Oncotarget. 2016;7:7754357.
148. Nabissi M, Morelli MB, Santoni M, Santoni G. Triggering of the TRPV2 channel by
cannabidiol sensitizes glioblastoma cells to cytotoxic chemotherapeutic agents.
Carcinogenesis. 2013;34:4857.
149. Elbaz M, Ahirwar D, Xiaoli Z, Zhou X, Lustberg M, Nasser MW, et al. TRPV2 is a
novel biomarker and therapeutic target in triple negative breast cancer.
Oncotarget. 2016;9:3345970.
150. Scott KA, Dennis JL, Dalgleish AG, Liu WM. Inhibiting heat shock proteins can
potentiate the cytotoxic effect of cannabidiol in human glioma cells. Anticancer
Res. 2015;35:582737.
151. Ivanov VN, Wu J, Hei TK. Regulation of human glioblastoma cell death by
combined treatment of cannabidiol, γ-radiation and small molecule inhibitors of
cell signaling pathways. Oncotarget. 2017;8:7406895.
152. Ivanov VN, Wu J, Wang TJC, Hei TK. Inhibition of ATM kinase upregulates
levels of cell death induced by cannabidiol and γ-irradiation in human glio-
blastoma cells. Oncotarget. 2019;10:82546. Erratum in: Oncotarget.
153. Bar-Sela G, Cohen I, Campisi-Pinto S, Lewitus GM, Oz-Ari L, Jehassi A, et al.
Cannabis consumption used by cancer patients during immunotherapy corre-
lates with poor clinical outcome. Cancers (Basel). 2020;12:2447.
154. Taha T, Meiri D, Talhamy S, Wollner M, Peer A, Bar-Sela G. Cannabis impacts
tumour response rate to nivolumab in patients with advanced malignancies.
Oncologist. 2019;24:54954.
155. Kenyon J, Liu W, Dalgleish A. Report of objective clinical responses of cancer
patients to pharmaceutical-grade synthetic cannabidiol. Anticancer Res.
B. Hinz and R. Ramer
British Journal of Cancer
The corresponding authors preclinical experimental work on the effects of cannabinoids
and chemotherapeutic agents is currently funded by the German Research Foundation
(DFG, HI 813/9-1), the joint research project ONKOTHER-H (supported by the European
Social Fund, reference: ESF/14-BM-A55-0002/18, and the Ministry of Education, Science
and Culture of Mecklenburg-Vorpommern, Germany) and by the joint research projects
RESPONSE FV18 (reference: 03ZZ0928A) and RESPONSE TV3 (reference: 03ZZ0933A) of
the Federal Ministry of Education and Research (BMBF), Germany.
BH and RR: conceptualisation, formal analysis, methodology, writing and editing.
Open Access funding enabled and organized by Projekt DEAL.
Not applicable.
Not applicable.
The authors declare no competing interests.
Correspondence and requests for materials should be addressed to Burkhard Hinz.
Reprints and permission information is available at
Publishers note Springer Nature remains neutral with regard to jurisdictional claims
in published maps and institutional afliations.
Open Access This article is licensed under a Creative Commons
Attribution 4.0 International License, which permits use, sharing,
adaptation, distribution and reproduction in any medium or format, as long as you give
appropriate credit to the original author(s) and the source, provide a link to the Creative
Commons license, and indicate if changes were made. The images or other third party
material in this article are included in the articles Creative Commons license, unless
indicated otherwise in a credit line to the material. If material is not included in the
articles Creative Commons license and your intended use is not permitted by statutory
regulation or exceeds the permitted use, you will need to obtain permission directly
from the copyright holder. To view a copy of this license, visit http://creativecommons.
© The Author(s) 2022
B. Hinz and R. Ramer
British Journal of Cancer
... For transcriptome data analysis, we utilized publicly available data from The Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) dataset, obtained using the TCGAbiolinks package in R software [19]. Additionally, we obtained data from the GSE4024 (accessed on 12 July 2023), GSE89377 (accessed on 12 July 2023), and GSE6764 (accessed on 12 July 2023) datasets from the NCBI GEO database (, accessed on 12 July 2023). ...
... CBD has been previously found to show anti-cancer activity and possess apoptotic properties when used in a high concentration (e.g., >40 µM) against various cancer cells, including breast cancer, lung cancer, and HCC [12,20,21]. To determine the optimal concentration of CBD associated with anti-cancer activity, we first evaluated the half-maximal inhibitory concentration (IC 50 ) value of CBD in HepG2 cells, which was found to be 32.52 µM ( Figure 1A). ...
... Previous studies have highlighted the anti-cancer effects of CBD, including its apoptotic properties, on various cancer cells, including breast cancer, lung cancer, and HCC [12,20,21]. Similarly, we observed a concentration-dependent increase in CBDinduced apoptosis in HepG2 cells. ...
Full-text available
Cannabidiol (CBD), a primary constituent in hemp and cannabis, exerts broad pharmacological effects against various diseases, including cancer. Additionally, cabozantinib, a potent multi-kinase inhibitor, has been approved for treating patients with advanced hepatocellular carcinoma (HCC). Recently, there has been an increase in research on combination therapy using cabozantinib to improve efficacy and safety when treating patients. Here, we investigated the effect of a combination treatment of cabozantinib and CBD on HCC cells. CBD treatment enhanced the sensitivity of HCC cells to cabozantinib-mediated anti-cancer activity by increasing cytotoxicity and apoptosis. Phospho-kinase array analysis demonstrated that the apoptotic effect of the combination treatment was mainly related to p53 phosphorylation regulated by endoplasmic reticulum (ER) stress when compared to other kinases. The inhibition of p53 expression and ER stress suppressed the apoptotic effect of the combination treatment, revealing no changes in the expression of Bax, Bcl-2, cleaved caspase-3, cleaved caspase-8, or cleaved caspase-9. Notably, the effect of the combination treatment was not associated with cannabinoid receptor 1 (CNR1) and the CNR2 signaling pathways. Our findings suggest that the combination therapy of cabozantinib and CBD provides therapeutic efficacy against HCC.
... The plethora of effects can also be explained by the fact that cannabinoids bind to both CB1R and CB2R, and non-cannabinoid receptors, such as the adrenergic receptors, vanilloid receptor 1 (TVRP1), transient receptor ankyrin 1 potential (TRPA1), peroxisome proliferator-activated receptor-gamma or glitazone receptor (PPAR-γ), G55 proteincoupled receptor (GPR55), and nuclear receptor (NRs) (Huang et al., 2020). This broader view of ligands and enzymes involved in the endocannabinoid system led to the concept of the endocannabinoidome, which encompasses hundreds of lipid mediators and tens of enzymes and molecular targets (Huestis et al., 2019;Pryimak et al., 2021;Hinz and Ramer, 2022). As such, the potential effect of cannabinoids on the most prominent cancer-associated symptoms, such as pain, nausea, vomiting, cachexia, anorexia, depression or anxiety has been investigated in several clinical and preclinical studies. ...
... A large body of available data shows that cannabinoids can reduce cancer cell proliferation through protein kinase B (Akt) inhibition (Hinz and Ramer, 2022). Akt is a serine/threonine kinase that plays a key role in growth factor-induced cell survival. ...
... It is a critical mediator of the canonical PI3K signaling pathway, which has long been recognized for its major oncogenic role within the cell (Revathidevi and Munirajan, 2019). Other important mechanisms include retinoblastoma protein hypophosphorylation (Hinz and Ramer, 2022), promotion of reactive oxygen species (Dando et al., 2013), modulation of the mitogen-activated protein kinases (MAPK) pathway and apoptosis signaling. Several preclinical studies report that cannabinoids induce cell cycle arrest by downregulation or inactivation of cyclindependent kinases (CDK) and cyclin modulation (Thoma et al., 2021). ...
Full-text available
Cannabis enjoyed a “golden age” as a medicinal product in the late 19th, early 20th century, but the increased risk of overdose and abuse led to its criminalization. However, the 21st century have witnessed a resurgence of interest and a large body of literature regarding the benefits of cannabinoids have emerged. As legalization and decriminalization have spread around the world, cancer patients are increasingly interested in the potential utility of cannabinoids. Although eager to discuss cannabis use with their oncologist, patients often find them to be reluctant, mainly because clinicians are still not convinced by the existing evidence-based data to guide their treatment plans. Physicians should prescribe cannabis only if a careful explanation can be provided and follow up response evaluation ensured, making it mandatory for them to be up to date with the positive and also negative aspects of the cannabis in the case of cancer patients. Consequently, this article aims to bring some clarifications to clinicians regarding the sometimes-confusing various nomenclature under which this plant is mentioned, current legislation and the existing evidence (both preclinical and clinical) for the utility of cannabinoids in cancer patients, for either palliation of the associated symptoms or even the potential antitumor effects that cannabinoids may have.
... : Alkyl, π-alkyl, π-σ bond, C-H bond, van der Waals. CB 2 : Alkyl, π-alkyl, π-π-T-shaped, π-σ bond 15 9.1 [91,111] results for the three cannabinoids. However, for the CB1 receptor, R-HHC (1) and Δ9-THC (35b) displayed similar high calculated binding affinities, while Δ8-THC (36b) and S-HHC (7) bound to this receptor with lower affinity. ...
... Given the emergence of in vivo studies on the use of saturated cannabinoids in the treatment of various diseases, including cancer [15,[114][115][116][117][118], neurological disorders [64,119,120], and diabetes [121,122], but also the prevalence of the consumption of these compounds [28], there is a crucial need to better comprehend their pharmacology and toxicology. In particular, the role of intrinsic efficacy in abuse-related effects, major metabolites, and adverse effects should be the subject of future study. ...
Full-text available
Natural and non-natural hexahydrocannabinols (HHC) were first described in 1940 by Adam and in late 2021 arose on the drug market in the United States and in some European countries. A background on the discovery, synthesis, and pharmacology studies of hydrogenated and saturated cannabinoids is described. This is harmonized with a summary and comparison of the cannabinoid receptor affinities of various classical, hybrid, and non-classical saturated cannabinoids. A discussion of structure–activity relationships with the four different pharmacophores found in the cannabinoid scaffold is added to this review. According to laboratory studies in vitro, and in several animal species in vivo, HHC is reported to have broadly similar effects to Δ9-tetrahydrocannabinol (Δ9-THC), the main psychoactive substance in cannabis, as demonstrated both in vitro and in several animal species in vivo. However, the effects of HHC treatment have not been studied in humans, and thus a biological profile has not been established.
... It may also have anti-migratory, antiinvasive, anti-metastatic, and maybe anti-angiogenic effects. Based on the findings, evidence is mounting that CBD is a powerful inhibitor of cancer growth and metastasis [Hinz and Ramer et al., 2022]. According to a recent study, CBD significantly decreased the proliferation of gastric cancer SGC-7901 cells in a concentration-dependent manner, with an IC50 value of 23.4 g/mL after 24 hours of treatment . ...
Full-text available
According to World Health Organisation data, cancer is one of the most influential diseases that goes head to head with heart diseases in the ranking of causes of death, causing approximately 10 million deaths annually and accounting for 20% of all deaths. Lung cancer is a leading cause of cancer-related deaths globally for both men and women. In some pre-clinical studies in cancer cell line in vitro experiments, some results have been obtained that Aloe Vera Extract (AVE) obtained from Aloe vera plant and Cannabindiol (CBD) obtained from Cannabis sativa may have anticancer effect against cancer, but more analysis is required for the reliability of these results. In this study, the cytotoxic and apoptotic effects of AVE and CBD in human lung cancer (A549 and BEAS) cells were studied in a multifaceted manner. Diagnosis of lung cancer disease is approximately 15% in the early stage and 85% in the late stage or metastatic stage. Therefore, despite the use of targeted drugs today, the 5-year survival rate of patients can only be 5-10%. In this study, the cytotoxic and apoptotic effects of AVE and CBD on human lung cancer cells A549 and healthy normal BEAS-2B cells were studied in a versatile and comparative manner. Cytotoxic effect of AVE and CBD was evaluated by xCELLigence RTCA System and AnnexinV-FITC/PI Apoptosis Assay by Flow Cytometry. In the current investigation, we discovered that AVE and CBD might incite a relative higher ratio of cell death in lung cancer cells (A549) than in non-cancer human epithelial lung cell line BEAS-2B, which may be achieved through regulating mitochondrial metabolism. In summary, our research is under Patent submission and shows that CBD and AVE have the potential to develop into a potent new anti-lung cancer drug.
... Phyto-cannabinoids are chemical compounds with psychoactive and non-psychoactive effects obtained from Cannabis sativa [9]. CBD represents the main non-psychoactive compound that is legally approved for use in research and has been frequently used as a palliative in inflammatory diseases and as an inhibitor of cancer progression in patients with different types of cancer [10][11][12]. In our previous in vitro study using 6D cells, the role of CBD was demonstrated to be an inhibitor of the EMT induced by IL-1β, leading to downregulated expression of proteins and features identified as malignant markers of the EMT processes. ...
Full-text available
Inflammation is a critical component of cancer development. Previously, we showed in vitro that IL-1β treatment of non-invasive human breast cancer MCF-7 cells promoted their transition to a malignant phenotype (6D cells). This epithelial–mesenchymal transition was reverted by exposure to cannabidiol (CBD). We show in a murine model that subcutaneous inoculation of 6D cells induced formation and development of tumors, the cells of which keep traits of malignancy. These processes were interrupted by administration of CBD under two schemes: therapeutic and prophylactic. In the therapeutic scheme, 6D cells inoculated mice developed tumors that reached a mean volume of 540 mm3 at 45 days, while 50% of CBD-treated mice showed gradual resorption of tumors. In the prophylactic scheme, mice were pre-treated for 15 days with CBD before cells inoculation. The tumors formed remained small and were eliminated under continuous CBD treatment in 66% of the animals. Histological and molecular characterization of tumors, from both schemes, revealed that CBD-treated cells decreased the expression of malignancy markers and show traits related with apoptosis. These results confirm that in vivo CBD blocks development of breast cancer tumors formed by cells induced to malignancy by IL-1β, endorsing its therapeutic potential for cancer treatment.
... Since its discovery in 1964, tetrahydrocannabinol (THC) and related analogs such as cannabidiol (CBD), natural and non-natural saturated cannabinoids have caught the attention of research groups all over the world [12][13][14][15]. Hexahydrocannbinol (HHC) is a newer cannabinoid to hit the cannabis consumer market, but it is not exactly a new cannabinoid. ...
Full-text available
Natural and non-natural hexahydrocannabinols (HHC) were first described in 1940 by Adam and in late 2021 arose on the drug market in the United States and in some European countries. A background on the discovery, synthesis, and pharmacology studies of hydrogenated and saturated cannabinoids is described. This is harmonized with a summary and comparison of the cannabinoid receptor affinities of various classical, hybrid, and non-classical saturated cannabinoids. A discussion of structure-activity relationships with the four different pharmacophores found in the cannabinoid scaffold is added to this review. According to laboratory studies in vitro, and in several animal species in vivo, HHC is reported to have broadly similar effects to Δ9-tetrahydrocannabinol (Δ9-THC), the main psychoactive substance in cannabis, as demonstrated both in vitro and in several animal species in vivo. However, the effects of HHC treatment have not been studied in humans, and thus a biological profile has not been established.
Full-text available
Background Breast cancer is one of the world’s most frequently diagnosed malignancy and the second highest cause of all cancers in females. This cancer mortality is mostly due to lung metastasis. High dose chemotherapy is one of therapeutic strategies beside others. Doxorubicin (DOX) is a widely used chemotherapy agent. however, use of DOX is limited due to its dose- depended cardiotoxic effects. Recently, cannabidiol (CBD) shows anti-tumoral and cardioprotective effects so we aimed to CBD administration with high-dose DOX chemotherapy can improve anticancer effect and reduce cardiotoxicity side effect. Method Mice breast cancer model established by injecting 4T1 cell lines. One group did not injected by 4T1 cells as a not cancerous group and administrated normal saline (1ml). In cancerous groups one group consider as cancerous control and administrated normal saline (1ml), other one administrated only DOX (5 mg/kg, IV) on day 1,7,14, other administrated CBD (5 mg/kg, IP) as DOX schedule, nex group administrated CBD and DOX at same time with previews doses and schedule, the last one administrated CBD one day before DOX as pretreatment. On day 21 all mice sacrificed, heart and lungs obtained and histological section preformed. SOD2, iNOS, MMP2, MMP9 evaluated through western blot and TUNEL test preformed for breast tumor. Results TUNEL test demonstrated CBD + DOX and pretreatment group was significantly green even compare to DOX group (P < 0.05). In CBD + DOX SOD2 increased and iNOS, MM2 and MMP9 decreased. Conclusions This study shows CBD + DOX at same time can increase anti-tumoral effect and reduce DOX cardiotoxicity effect. However CBD alone had a cardioroxic effect.
Consumer use of hemp-derived products continues to rise, underscoring the need to establish evidence-based safety guidance. The present study sought to develop recommendations for oral upper intake limits of cannabidiol (CBD) isolate. Sufficiently robust and reliable data for this purpose were identified from published human clinical trials and guideline-compliant toxicity studies in animal models. Based on the metrics used in this assessment, a potential Acceptable Daily Intake (ADI) value of 0.43 mg/kg-bw/d (e.g., 30 mg/d for 70-kg adult) was determined for the general population based on liver effects in human studies. This value applies to the most sensitive subpopulations, including children, over a lifetime of exposure and from all sources, including food. For dietary supplements with adequate product labeling intended for use by healthy adults only, a potential Upper Intake Limit (UL) of 70 mg/d was determined based on reproductive effects in animals. For healthy adults, except those trying to conceive, or currently pregnant or lactating, a conservative dietary supplement UL of 100 mg/d was identified based on liver effects; however, as the target population excludes individuals at risk for liver injury, an alternative dietary supplement UL of 160 mg/d for this population can also be considered.
Full-text available
In T-cell acute lymphoblastic leukemia (T-ALL) more than 50% of cases display autoactivation of Notch1 signaling, leading to oncogenic transformation. We have previously identified a specific chemovar of Cannabis that induces apoptosis by preventing Notch1 maturation in leukemia cells. Here, we isolated three cannabinoids from this chemovar that synergistically mimic the effects of the whole extract. Two were previously known, Cannabidiol (CBD) and Cannabidivarin (CBDV); whereas the third cannabinoid, which we termed 331-18A, was identified and fully characterized in this study. We demonstrated that these cannabinoids act through Cannabinoid receptor type 2 and TRPV1 to activate the integrated stress response pathway by depleting intracellular Ca2+. This is followed by increased mRNA and protein expression of ATF4, CHOP and CHAC1, which is hindered by inhibiting the upstream initiation factor eIF2α. The increased abundance of CHAC1 prevents Notch1 maturation, thereby reducing the levels of the active Notch1 intracellular domain, and consequently decreasing cell viability and increasing apoptosis. Treatment with the three isolated molecules resulted in reduced tumor size and weight in-vivo and slowed leukemia progression in mice models. Altogether, this study elucidated the mechanism of action of three distinct cannabinoids in modulating the Notch1 pathway, and constitutes an important step in the establishment of a new therapy for treating NOTCH1 -mutated diseases and cancers such as T-ALL.
Full-text available
Glioma is the most common primary malignant brain tumor with poor survival and limited therapeutic options. The non-psychoactive phytocannabinoid cannabidiol (CBD) has been shown to be effective against glioma; however, the molecular target and mechanism of action of CBD in glioma are poorly understood. Here we investigated the molecular mechanisms underlying the antitumor effect of CBD in preclinical models of human glioma. Our results showed that CBD induced autophagic rather than apoptotic cell death in glioma cells. We also showed that CBD induced mitochondrial dysfunction and lethal mitophagy arrest, leading to autophagic cell death. Mechanistically, calcium flux induced by CBD through TRPV4 (transient receptor potential cation channel subfamily V member 4) activation played a key role in mitophagy initiation. We further confirmed TRPV4 levels correlated with both tumor grade and poor survival in glioma patients. Transcriptome analysis and other results demonstrated that ER stress and the ATF4-DDIT3-TRIB3-AKT-MTOR axis downstream of TRPV4 were involved in CBD-induced mitophagy in glioma cells. Lastly, CBD and temozolomide combination therapy in patient-derived neurosphere cultures and mouse orthotopic models showed significant synergistic effect in both controlling tumor size and improving survival. Altogether, these findings showed for the first time that the antitumor effect of CBD in glioma is caused by lethal mitophagy and identified TRPV4 as a molecular target and potential biomarker of CBD in glioma. Given the low toxicity and high tolerability of CBD, we therefore propose CBD should be tested clinically for glioma, both alone and in combination with temozolomide. Abbreviations: 4-PBA: 4-phenylbutyrate; AKT: AKT serine/threonine kinase; ATF4: activating transcription factor 4; Baf-A1: bafilomycin A1; CANX: calnexin; CASP3: caspase 3; CAT: catalase; CBD: cannabidiol; CQ: chloroquine; DDIT3: DNA damage inducible transcript 3; ER: endoplasmic reticulum; GBM: glioblastoma multiforme; GFP: green fluorescent protein; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; PARP1: poly(ADP-ribose) polymerase; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; SLC8A1: solute carrier family 8 member A1; SQSTM1: sequestosome 1; TCGA: The cancer genome atlas; TEM: transmission electron microscopy; TMZ: temozolomide; TRIB3: tribbles pseudokinase 3; TRPC: transient receptor potential cation channel subfamily C; TRPV4: transient receptor potential cation channel subfamily V member 4
Full-text available
Background Preclinical data suggest some cannabinoids may exert antitumour effects against glioblastoma (GBM). Safety and preliminary efficacy of nabiximols oromucosal cannabinoid spray plus dose-intense temozolomide (DIT) was evaluated in patients with first recurrence of GBM. Methods Part 1 was open-label and Part 2 was randomised, double-blind, and placebo-controlled. Both required individualised dose escalation. Patients received nabiximols (Part 1, n = 6; Part 2, n = 12) or placebo (Part 2 only, n = 9); maximum of 12 sprays/day with DIT for up to 12 months. Safety, efficacy, and temozolomide (TMZ) pharmacokinetics (PK) were monitored. Results The most common treatment-emergent adverse events (TEAEs; both parts) were vomiting, dizziness, fatigue, nausea and headache. Most patients experienced TEAEs that were grade 2 or 3 (CTCAE). In Part 2, 33% of both nabiximols- and placebo-treated patients were progression-free at 6 months. Survival at 1 year was 83% for nabiximols- and 44% for placebo-treated patients ( p = 0.042), although two patients died within the first 40 days of enrolment in the placebo arm. There were no apparent effects of nabiximols on TMZ PK. Conclusions With personalised dosing, nabiximols had acceptable safety and tolerability with no drug–drug interaction identified. The observed survival differences support further exploration in an adequately powered randomised controlled trial. Clinical trial registration Part 1– NCT01812603; Part 2– NCT01812616.
Full-text available
Glioblastomas (GBMs) are aggressive brain tumors with frequent genetic alterations in TP53 and PTEN tumor suppressor genes rendering resistance to standard chemotherapeutics. Cannabinoid type 1 and 2 (CB1/CB2) receptor expression in GBMs and antitumor activity of cannabinoids in glioma cells and animal models, raised promises for a targeted treatment of these tumors. The susceptibility of human glioma cells to CB2-agonists and their mechanism of action are not fully elucidated. We determined CB1 and CB2 expression in 14 low-grade and 21 high-grade tumor biopsies, GBM-derived primary cultures and established cell lines. The non-selective CB receptor agonist WIN55,212-2 (but not its inactive enantiomer) or the CB2-selective agonist JWH133 induced apoptosis in patient-derived glioma cultures and five established glioma cell lines despite p53 and/or PTEN deficiency. Growth inhibitory efficacy of cannabinoids correlated with CB1/CB2 expression (EC50 WIN55,212-2: 7.36–15.70 µM, JWH133: 12.15–143.20 µM). Treatment with WIN55,212-2 or JWH133 led to activation of the apoptotic mitochondrial pathway and DNA fragmentation. Synthetic cannabinoid action was associated with the induction of autophagy and knockdown of autophagy genes augmented cannabinoid-induced apoptotic cell death. The high susceptibility of human glioblastoma cells to synthetic cannabinoids, despite genetic defects contributing to apoptosis resistance, makes cannabinoids promising anti-glioma therapeutics.
Full-text available
Cannabis sativa contains more than 500 constituents, yet the anticancer properties of the vast majority of cannabis compounds remains unknown. We aimed to identify cannabis compounds and their combinations presenting cytotoxicity against bladder urothelial carcinoma (UC), the most common urinary system cancer. An XTT assay was used to determine cytotoxic activity of C. sativa extracts on T24 and HBT-9 cell lines. Extract chemical content was identified by high-performance liquid chromatography (HPLC). Fluorescence-activated cell sorting (FACS) was used to determine apoptosis and cell cycle, using stained F-actin and nuclei. Scratch and transwell assays were used to determine cell migration and invasion, respectively. Gene expression was determined by quantitative Polymerase chain reaction (PCR). The most active decarboxylated extract fraction (F7) of high-cannabidiol (CBD) C. sativa was found to contain cannabichromene (CBC) and Δ9-tetrahydrocannabinol (THC). Synergistic interaction was demonstrated between CBC + THC whereas cannabinoid receptor (CB) type 1 and type 2 inverse agonists reduced cytotoxic activity. Treatments with CBC + THC or CBD led to cell cycle arrest and cell apoptosis. CBC + THC or CBD treatments inhibited cell migration and affected F-actin integrity. Identification of active plant ingredients (API) from cannabis that induce apoptosis and affect cell migration in UC cell lines forms a basis for pre-clinical trials for UC treatment.
Full-text available
Children with medulloblastoma and ependymoma are treated with a multidisciplinary approach that incorporates surgery, radiotherapy, and chemotherapy; however, overall survival rates for patients with high-risk disease remain unsatisfactory. Data indicate that plant-derived cannabinoids are effective against adult glioblastoma; however, preclinical evidence supporting their use in pediatric brain cancers is lacking. Here we investigated the potential role for Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD) in medulloblastoma and ependymoma. Dose-dependent cytotoxicity of medulloblastoma and ependymoma cells was induced by THC and CBD in vitro, and a synergistic reduction in viability was observed when both drugs were combined. Mechanistically, cannabinoids induced cell cycle arrest, in part by the production of reactive oxygen species, autophagy, and apoptosis; however, this did not translate to increased survival in orthotopic transplant models despite being well tolerated. We also tested the combination of cannabinoids with the medulloblastoma drug cyclophosphamide, and despite some in vitro synergism, no survival advantage was observed in vivo. Consequently, clinical benefit from the use of cannabinoids in the treatment of high-grade medulloblastoma and ependymoma is expected to be limited. This study emphasizes the importance of preclinical models in validating therapeutic agent efficacy prior to clinical trials, ensuring that enrolled patients are afforded the most promising therapies available.
Full-text available
Therapeutic agents used for non-small cell lung cancer (NSCLC) have limited curative efficacy and may trigger serious adverse effects. Cannabinoid ligands exert antiproliferative effect and induce apoptosis on numerous epithelial cancers. We confirmed that CB1 receptor (CB1R) is expressed in NSCLC cells in this study. Arachidonoylcyclopropylamide (ACPA) as a synthetic, CB1R-specific ligand decreased proliferation rate in NSCLC cells by WST-1 analysis and real-time proliferation assay (RTCA). The half-maximal inhibitory concentration (IC50) dose of ACPA was calculated as 1.39 × 10 ⁻¹² M. CB1 antagonist AM281 inhibited the antiproliferative effect of ACPA. Flow cytometry and ultrastructural analyzes revealed significant early and late apoptosis with diminished cell viability. Nano-immunoassay and metabolomics data on activation status of CB1R-mediated pro-apoptotic pathways found that ACPA inhibited Akt/PI3K pathway, glycolysis, TCA cycle, amino acid biosynthesis, and urea cycle and activated JNK pathway. ACPA lost its chemical stability after 24 hours tested by liquid chromatography-mass spectrometry (LC–MS/MS) assay. A novel ACPA-PCL nanoparticle system was developed by nanoprecipitation method and characterized. Sustained release of ACPA-PCL nanoparticles also reduced proliferation of NSCLC cells. Our results demonstrated that low dose ACPA and ACPA-PCL nanoparticle system harbor opportunities to be developed as a novel therapy in NSCLC patients that require further in vivo studies beforehand to validate its anticancer effect.
Full-text available
Enhancing the therapeutic efficacy of anti-tumor drugs is essential for cancer management. Although cannabinoid receptor 2 (CB2R) stimulation exerts anti-tumor action in glioma cells by regulating cellular proliferation, differentiation, or apoptosis, selective CB2R agonist alone does not achieve a satisfactory therapeutic outcome. Herein, we aimed to evaluate the possible strategy for enhancing the anti-glioma efficacy of JWH133, a selective CB2R agonist. In this study, immunofluorescence and qRT-PCR were used to investigate microglia polarization. Tumor growth was monitored via bioluminescent imaging using the IVIS Spectrum System. The angiogenesis of human brain microvascular endothelial cells (HBMECs) was detected by the tube formation assay. qRT-PCR was used to investigate cytochrome P450 2J2 (CYP2J2) and 11,12-epoxyeicosatrienoic acid (11,12-EET) expression. Our results showed that administration of JWH133 significantly promoted microglial M2 polarization both in vitro and in vivo . The medium supernatant of M2 microglia induced by JWH133 treatment facilitated angiogenesis of HBMECs. CYP2J2 expression and 11,12-EET release in the supernatant of JWH133-induced M2 microglia were significantly upregulated. Treatment with 11,12-EET prompted HBMEC angiogenesis and glioma growth. CYP2J2 knockdown restrained the release of 11,12-EET and significantly enhanced the anti-tumor effect of JWH133 on glioma. This study showed that targeting CYP2J2 might be a beneficial strategy to enhance the anti-glioma efficacy of JWH133 by inhibiting the pro-angiogenesis function of M2 microglia.
Full-text available
Cannabidiol (CBD) has anti-tumorigenic activity. However, the anti-cancer effect of CBD on head and neck squamous cell carcinoma (HNSCC) remains unclear. The cytotoxicity of CBD on HNSCC was analyzed using cell survival and colony-forming assays in vitro. RNA-seq was used for determining the mechanism underlying CBD-induced cell death. Xenograft mouse models were used to determine CBD’s effects in vivo. CBD treatment significantly reduced migration/invasion and viability of HNSCC cells in a dose- and time-dependent manner. HNSCC mouse xenograft models revealed anti-tumor effects of CBD. Furthermore, combinational treatment with CBD enhanced the efficacy of chemotherapy drugs. Apoptosis and autophagy processes were involved in CBD-induced cytotoxicity of HNSCCs. RNA-seq identified decreased expression of genes associated with DNA repair, cell division, and cell proliferation, which were involved in CBD-mediated cytotoxicity toward HNSCCs. We identified CBD as a new potential anti-cancer compound for single or combination therapy of HNSCC.
Ethnopharmacological relevance: As one of the first plants used by ancient people, cannabis has been used for medicinal purposes for thousands of years. The long history of medicinal cannabis use contrasts with the paucity of archaeobotanical records. Moreover, physical evidence of medicinal cannabis use in a secular context is much rarer than evidence of medicinal cannabis use in religious or ritual activities, which impedes our understanding of the history of medicinal cannabis use. Aim of the study: This study aims to provide archeobotanical evidence of medicinal cannabis use and analyse the specific medicinal usage of cannabis in a secular context in ancient times. Materials and methods: Plant remains were collected from the Laoguanshan Cemetery of the Han Dynasty in Chengdu, South China, with the archaeological flotation process and were identified based on morphological and anatomical characteristics. The examination of the medicinal significance of the remains relied on the investigation of the documentation on unearthed medical bamboo slips, the diseases of the tomb occupants, the cemetery's cultural background and Chinese historical records. Results: The botanical remains were accurately identified as cannabis. More than 120 thousand fruits were found, which represents the largest amount of cannabis fruit remains that have been statistically analysed from any cemetery in the world thus far. The cannabis fruits are suspected to have been used for medical purposes in a secular context and were most likely used to stop severe bleeding of the uterus and treat lumbago and/or arthralgia. Conclusions: The cannabis fruit remains reported here likely represent the first physical evidence of medicinal cannabis use for the treatment of metrorrhagia, severe lumbago, and/or arthralgia. This study emphasizes the importance of the evidence of the diseases suffered by the occupants of the tomb in determining the medicinal use of cannabis in a secular context and contributes to a comprehensive understanding of the ancient history of medicinal cannabis.
A targeted modulation of the endocannabinoid system is currently discussed as a promising strategy for cancer treatment. An important enzyme for the endocannabinoid metabolism is the monoacylglycerol lipase (MAGL), which catalyzes the degradation of 2-arachidonoylglycerol (2-AG) to glycerol and free fatty acids. In this study we investigated the influence of MAGL inhibition on lung cancer cell invasion and metastasis. Using LC-MS, significantly increased 2-AG levels were detected in A549 cells treated with the MAGL inhibitor JZL184. In athymic nude mice, JZL184 suppressed metastasis of A549 cells in a dose-dependent manner, whereby the antimetastatic effect was cancelled by the CB1 receptor antagonist AM-251. In vitro, JZL184 induced a time- and concentration-dependent reduction of A549 cell invasion through Matrigel-coated membranes, which was likewise reversed by AM-251. A MAGL inhibition-associated reduction of free fatty acids as a cause of the anti-invasive effect could be excluded by add-back experiments with palmitic acid. Both JZL184 and the MAGL substrate 2-AG led to an increased formation of the tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), whereby a TIMP-1 knockdown using siRNA significantly attenuated the anti-invasive effects of both substances. Decreased invasion and TIMP-1 upregulation was also caused by the MAGL inhibitors JW651 and MJN110 or transfection with MAGL siRNA. A CB1- and TIMP-1-dependent anti-invasive effect was further confirmed for JZL184 in H358 lung cancer cells. In conclusion, MAGL inhibition led to a CB1-dependent decrease in human lung cancer cell invasion and metastasis via inhibition of 2-AG degradation, with TIMP-1 identified as a mediator of the anti-invasive effect.