Tag Archives: AEA

The endogenous cannabinoid, anandamide, induces COX-2-dependent cell death in apoptosis-resistant colon cancer cells.

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Abstract

“Despite recent advances in understanding colorectal tumour biology, there is still a need to improve the 5-year survival rate of patients with colorectal cancer as approximately 40% of patients presenting with advanced disease will remain resistant to therapy. One of the major contributing factors in resistance to therapy is the failure of colorectal tumour cells to undergo apoptosis. Hence there is an urgent need to develop novel therapeutic approaches that can target apoptosis-resistant cells. To this end, we investigated the potential efficacy of the endogenous cannabinoid anandamide to induce cell death in apoptosis-resistant colon cancer cells. Here, for the first time, we show that anandamide can induce cell death in the apoptosis-resistant HCT116 Bax-/- colorectal cell line. Importantly, we provide direct genetic evidence that this induction of cell death is dependent on COX-2 expression. Interestingly, increased COX-2 expression also sensitised the SW480 colorectal cancer cell line (low endogenous COX-2) to anandamide-induced death, whereas COX-2 suppression by RNAi inhibited anandamide-induced cell death in the HCA7 colorectal cancer cell line (high endogenous COX-2 expression). This COX-2-dependent death was independent of cannabinoid receptor engagement (CB1 or CB2), and not a direct consequence of reactive oxygen species (ROS) formation. This study demonstrates a novel utilisation for COX-2 expression, targeting apoptotic defective colorectal cancer cells for destruction by anandamide. As COX-2 is not expressed in the normal colorectal epithelium, but highly expressed in colorectal tumours and apoptosis resistance contributes to treatment failure, these data suggest that anandamide has the potential to be an effective therapeutic in colorectal cancer.”

http://www.ncbi.nlm.nih.gov/pubmed/20514410

Effects of anandamide on polyamine levels and cell growth in human colon cancer cells.

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Abstract

“BACKGROUND:

Anandamide (AEA) is an endogenous agonist for cannabinoid receptor CB1-R and seems to be involved in the control of cancer growth. Polyamines are compounds that play an important role in cell proliferation and differentiation. Our aim was to investigate the effect of AEA on the polyamine levels (putrescine, spermidine and spermine) and cell growth of three human colon cancer cell lines, positive for CB1-R.

MATERIALS AND METHODS:

After AEA treatment of DLD-1, HT-29 and SW620 cells, polyamine analysis was performed by high-performance liquid chromatography (HPLC) and cell growth was measured by 3-(4,5 di-methylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test. CB1 gene expression was determined using reverse transcription and polymerase chain reaction (RT-PCR).

RESULTS:

AEA significantly reduced polyamine levels and cell proliferation dose-dependently when the tested cell lines were exposed for 24 h and 48 h. This inhibitory effect was mediated by CB1-R, since SR 1411716A, a selective CB-1 receptor antagonist, was able to entirely antagonize the effect of AEA. CB1-R mRNA levels were enhanced after AEA treatment in DLD-1 cells, whereas no induction was found in HT-29 and SW620 cells.

CONCLUSION:

It appears that mechanisms by which AEA may affect growth of colon cancer cells involve a decrease in cell proliferation rate by reducing the polyamine levels.”

http://www.ncbi.nlm.nih.gov/pubmed/20682986

Interaction of endocannabinoid system and steroid hormones in the control of colon cancer cell growth.

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Abstract

“Increasing evidence suggest the role of the cannabinoid receptors (CBs) in the control of cell survival or death and signaling pathways involved in tumor progression. Cancer cell lines are characterized by a subtle modulation of CB levels which produces a modified responsiveness to specific ligands, but the molecular mechanisms underlying these events are poorly and partially understood. We previously provided evidence that the endocannabinoid (EC) anandamide (AEA) exerts anti-proliferative effect likely by modulation of the expression of genes involved in the cellular fate. In this study we focused on the role of the CB1 receptor, ECs, and steroids in the mechanisms involved in colorectal cancer (CRC) cell growth inhibition in vitro. We demonstrated that, in DLD1 and SW620 cells, 17β-estradiol induced a specific and strong up-regulation of the CB1 receptor by triggering activation of the CB1 promoting region, localized at the exon 1 of the CNR1 gene. Moreover, treatment of DLD1 and SW620 cells with Met-F-AEA, a stable AEA-analogous, or URB597, a selective inhibitor of FAAH, induced up-regulation of CB1 expression by co-localization of PPARγ and RXRα at the promoting region. Finally, increased availability of AEA, of both exogenous and endogenous sources, induced the expression of estrogen receptor-beta in both cell lines. Our results partially elucidated the role of EC system in the molecular mechanisms enrolled by steroids in the inhibition of colon cancer cell growth and strongly suggested that targeting the EC system could represent a promising tool to improve the efficacy of CRC treatments.”

http://www.ncbi.nlm.nih.gov/pubmed/21412772

Possible endocannabinoid control of colorectal cancer growth.

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Abstract

“BACKGROUND & AIMS:

The endocannabinoids anandamide and 2-arachidonoylglycerol (2-AG) inhibit cancer cell proliferation by acting at cannabinoid receptors (CBRs). We studied (1). the levels of endocannabinoids, cannabinoid CB(1) and CB(2) receptors, and fatty acid amide hydrolase (FAAH, which catalyzes endocannabinoid hydrolysis) in colorectal carcinomas (CRC), adenomatous polyps, and neighboring healthy mucosa; and (2). the effects of endocannabinoids, and of inhibitors of their inactivation, on human CRC cell proliferation.

METHODS:

Tissues were obtained from 21 patients by biopsy during colonoscopy. Endocannabinoids were measured by liquid chromatography-mass spectrometry (LC-MS). CB(1), CB(2), and FAAH expression were analyzed by RT-PCR and Western immunoblotting. CRC cell lines (CaCo-2 and DLD-1) were used to test antiproliferative effects.

RESULTS:

All tissues and cells analyzed contain anandamide, 2-AG, CBRs, and FAAH. The levels of the endocannabinoids are 3- and 2-fold higher in adenomas and CRCs than normal mucosa. Anandamide, 2-AG, and the CBR agonist HU-210 potently inhibit CaCo-2 cell proliferation. This effect is blocked by the CB(1) antagonist SR141716A, but not by the CB(2) antagonist SR144528, and is mimicked by CB(1)-selective, but not CB(2)-selective, agonists. In DLD-1 cells, both CB(1) and CB(2) receptors mediate inhibition of proliferation. Inhibitors of endocannabinoid inactivation enhance CaCo-2 cell endocannabinoid levels and block cell proliferation, this effect being antagonized by SR141716A. CaCo-2 cell differentiation into noninvasive cells results in increased FAAH expression, lower endocannabinoid levels, and no responsiveness to cannabinoids.

CONCLUSIONS:

Endocannabinoid levels are enhanced in transformed colon mucosa cells possibly to counteract proliferation via CBRs. Inhibitors of endocannabinoid inactivation may prove useful anticancer agents.”

http://www.ncbi.nlm.nih.gov/pubmed/12949714

Increased endocannabinoid levels reduce the development of precancerous lesions in the mouse colon

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Abstract

“Colorectal cancer is an increasingly important cause of death in Western countries. Endocannabinoids inhibit colorectal carcinoma cell proliferation in vitro. In this paper, we investigated the involvement of endocannabinoids on the formation of aberrant crypt foci (ACF, earliest preneoplastic lesions) in the colon mouse in vivo. ACF were induced by azoxymethane (AOM); fatty acid amide hydrolase (FAAH) and cannabinoid receptor messenger ribonucleic acid (mRNA) levels were analyzed by the quantitative reverse transcription polymerase chain reaction (RT-PCR); endocannabinoid levels were measured by liquid chromatography-mass spectrometry; caspase-3 and caspase-9 expressions were measured by Western blot analysis. Colonic ACF formation after AOM administration was associated with increased levels of 2-arachidonoylglycerol (with no changes in FAAH and cannabinoid receptor mRNA levels) and reduction in cleaved caspase-3 and caspase-9 expression. The FAAH inhibitor N-arachidonoylserotonin increased colon endocannabinoid levels, reduced ACF formation, and partially normalized cleaved caspase-3 (but not caspase-9) expression. Notably, N-arachidonoylserotonin completely prevented the formation of ACF with four or more crypts, which have been show to be best correlated with final tumor incidence. The effect of N-arachidonoylserotonin on ACF formation was mimicked by the cannabinoid receptor agonist HU-210. No differences in ACF formation were observed between CB(1) receptor-deficient and wild-type mice. It is concluded that pharmacological enhancement of endocannabinoid levels (through inhibition of endocannabinoid hydrolysis) reduces the development of precancerous lesions in the mouse colon. The protective effect appears to involve caspase-3 (but not caspase-9) activation.

Cannabinoids have been licensed for clinical use as palliative treatment of chemotherapy, but increasing evidence shows antitumor actions of cannabinoid agonists on several tumor cells in vitro and in animal models [4, 5]. The main psychotropic cannabinoid is Δ9-tetrahydrocannabinol, which exerts its biological effects mainly by activating two G protein-coupled cannabinoid receptors, named CB1 and CB2 receptors [5]. Endogenous ligands for the cannabinoid receptors have been identified; the best known are arachidonylethanolamide (anandamide) and 2-arachidonoylglycerol (2-AG) [4–6]. When released, anandamide and 2-AG are removed from extracellular compartments by a carrier-mediated reuptake process, and once within the cell, both endocannabinoids are hydrolyzed by intracellular hydrolytic enzymes of which the enzyme fatty acid amide hydrolase (FAAH) is capable of recognizing both compounds as substrates [5, 6].

The proposed mechanisms of the antitumoral effect of cannabinoids are complex and may involve induction of apoptosis in tumor cells, antiproliferative actions, and an antimetastatic effects through inhibition of angiogenesis and tumor cell migration [6]. Concerning the gastrointestinal tract, it has been shown that cannabinoid receptor agonists, mostly via CB1 activation, potently inhibit the cell proliferation of colorectal carcinoma cell lines [7]. Furthermore, compounds capable of inhibiting endocannabinoid degradation and hence of prolonging the lifespan of endocannabinoids only when and where these compounds are produced to exert physiological or pathophysiological functions also inhibit colorectal carcinoma growth in vitro [7].

In summary, the present study provides strong evidence that enhancement of colon endocannabinoid levels through pharmacological inhibition of their enzymatic hydrolysis may be protective against preneoplastic lesions in the mouse colon; a condition that, like humans adenomatous polyps and colorectal carcinoma [7], is accompanied by an elevated endocannabinoid tone. This protective effect could be due to indirect activation of one or more of the several targets proposed to date for the endocannabinoids and appears to involve caspase-3 activation and subsequent apoptosis of colon preneoplastic cells. Further studies will be required to investigate if cannabinoid CB2 receptors are involved in the protective effects of AA-5-HT and HU210 and if these effects have any therapeutic relevance for the treatment of human colon carcinoma.”

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2755791/

Cannabis for migraine treatment: the once and future prescription? An historical and scientific review.

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Abstract

“Cannabis, or Marijuana, has been used for centuries for both symptomatic and prophylactic treatment of migraine. It was highly esteemed as a headache remedy by the most prominent physicians of the age between 1874 and 1942, remaining part of the Western pharmacopoeia for this indication even into the mid-twentieth century. Current ethnobotanical and anecdotal references continue to refer to its efficacy for this malady, while biochemical studies of THC and anandamide have provided a scientific basis for such treatment. The author believes that controlled clinical trials of Cannabis in acute migraine treatment are warranted.”

http://www.ncbi.nlm.nih.gov/pubmed/9696453

 

Biochemical changes in endocannabinoid system are expressed in platelets of female but not male migraineurs.

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Abstract

“The endogenous cannabinoid anandamide (AEA) plays important roles in modulating pain. Head pain is an almost universal human experience, yet primary headache disorders, such as migraine without aura (MoA) or episodic tension-type headache (ETTH), can represent a serious threat to well-being when frequent and disabling. We assessed the discriminating role of endocannabinoids among patients with ETTH or MoA, and control subjects. We measured the activity of AEA hydrolase and AEA transporter, and the level of cannabinoid receptors in peripheral platelets from MoA, ETTH and healthy controls. Sixty-nine headache patients and 36 controls were selected. Diagnosis of headache type was made according to the International Headache Society criteria. We observed significant sex differences concerning AEA membrane transporter and fatty acid amide hydrolase activity in all groups. An increase in the activity of AEA hydrolase and AEA transporter was found in female but not male migraineurs. Cannabinoid receptors were the same in all groups. Here we show that the endocannabinoid system in human platelets is altered in female but not male migraneurs. Our results suggest that in migraineur women an increased AEA degradation by platelets, and hence a reduced concentration of AEA in blood, might reduce the pain threshold and possibly explain the prevalence of migraine in women. The involvement of the endocannabinoid system in migraine is new and broadens our knowledge of this widespread and multifactorial disease.”

http://www.ncbi.nlm.nih.gov/pubmed/16472333

Endocannabinoids in platelets of chronic migraine patients and medication-overuse headache patients: relation with serotonin levels.

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Abstract

“BACKGROUND:

Chronic migraine (CM) and medication-overuse headaches (MOH) are well-recognized disabling conditions affecting a significant portion of the headache population attending centers specialized in treating headaches. A dysfunctioning of the serotonergic system has been demonstrated in MOH and CM patients. Here we report on our assessment of the dysfunctioning of the endocannabinoid system as a potential underlying factor in pathogenic mechanisms involved in CM and MOH.

METHOD:

To test the hypothesis of an impairment in the endocannabinoid system in patients with MOH and CM and to assess its relationship with any disruption of the serotonergic system, we determined the levels of the two main endogenous cannabinoids, anandamide (AEA) and 2-acylglycerol (2-AG), in platelets of 20 CM patients, 20 MOH patients and 20 control subjects and also measured the platelet serotonin levels in the same patients.

RESULTS:

We found that 2-AG and AEA levels were significantly lower in MOH patients and CM patients than in the control subjects, without significant differences between the two patient groups. Serotonin levels were also strongly reduced in the two patient groups and were correlated with 2-AG levels, with higher values for MOH patients.

CONCLUSION:

These data support the potential involvement of a dysfunctioning of the endocannabinoid and serotonergic systems in the pathology of CM and MOH. These systems appear to be mutually related and able to contribute to the chronification of both CM and MOH.”

http://www.ncbi.nlm.nih.gov/pubmed/18004553

Acute Reduction of Anandamide-Hydrolase (FAAH) Activity is Coupled With a Reduction of Nociceptive Pathways Facilitation in Medication-Overuse Headache Subjects After Withdrawal Treatment.

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“Objectives.- We investigated (1) a possible relationship between the functional activity of the endocannabinoid system and the facilitation of pain processing in migraineurs with medication-overuse headache, and (2) the effect of withdrawal treatment on both. Background.- The endocannabinoid system antinociception effect includes prevention of nociceptive pathways sensitization. The sensitization of the pain pathways has been demonstrated to be pivotal in the development and maintenance of chronic form of migraine, including medication-overuse headache. Methods.- We used the temporal summation threshold of the nociceptive withdrawal reflex to explore the spinal cord pain processing, and the platelet activity of the enzyme fatty acid amide hydrolase to detect the functional state of the endocannabinoid system in 27 medication-overuse headache subjects before and 10 and 60 days after a standard withdrawal treatment and compared results with those of 14 controls. Results.- A significantly reduced temporal summation threshold and increased related pain sensation was found in subjects before withdrawal treatment when compared with controls. A significant fatty acid amide hydrolase activity reduction coupled with a significant improvement (reduction) in facilitation of spinal cord pain processing (increase in temporal summation threshold and reduction in related pain sensation) was found in medication-overuse headache subjects at both 10 and 60 days after withdrawal treatment when compared with medication-overuse headache subjects before withdrawal treatment. Conclusions.- We demonstrated a marked facilitation in spinal cord pain processing in medication-overuse headache before withdrawal treatment when compared with controls. Furthermore, the acute reduction of the fatty acid amide hydrolase activity coupled with a reduction of the facilitation in pain processing immediately (10 days) after withdrawal treatment and its persistence 60 days after withdrawal treatment could represent the consequence of a mechanism devoted to acutely reduce the degradation of endocannabinoids and aimed to increase the activity of the endocannabinoid system that results in an antinociceptive effect.”

http://www.ncbi.nlm.nih.gov/pubmed/22670561

Anandamide Is Able to Inhibit Trigeminal Neurons Using an in Vivo Model of Trigeminovascular-Mediated Nociception

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“Arachidonylethanolamide (anandamide, AEA) is believed to be the endogenous ligand of the cannabinoid CB(1) and CB(2) receptors. CB(1) receptors have been found localized on fibers in the spinal trigeminal tract and spinal trigeminal nucleus caudalis. Known behavioral effects of anandamide are antinociception, catalepsy, hypothermia, and depression of motor activity, similar to Delta(9)-tetrahydocannanbinol, the psychoactive constituent of cannabis. It may be a possible therapeutic target for migraine. In this study, we looked at the possible role of the CB(1) receptor in the trigeminovascular system, using intravital microscopy to study the effects of anandamide against various vasodilator agents. Anandamide was able to inhibit dural blood vessel dilation brought about by electrical stimulation by 50%, calcitonin gene-related peptide (CGRP) by 30%, capsaicin by 45%, and nitric oxide by 40%. CGRP(8-37) was also able to attenuate nitric oxide (NO)-induced dilation by 50%. The anandamide inhibition was reversed by the CB(1) receptor antagonist AM251. Anandamide also reduced the blood pressure changes caused by CGRP injection, this effect was not reversed by AM251. It would seem that anandamide acts both presynaptically, to prevent CGRP release from trigeminal sensory fibers, and postsynaptically to inhibit the CGRP-induced NO release in the smooth muscle of dural arteries. CB(1) receptors seem to be involved in the NO/CGRP relationship that exists in causing headache and dural blood vessel dilation. It also seems that some of the blood pressure changes caused by anandamide are mediated by a noncannabinoid receptor, as AM251 was unable to reverse these effects. It can be suggested that anandamide is tonically released to play some form of modulatory role in the trigeminovascular system.

The known behavioral effects of anandamide are similar to that of Δ9-tetrahydrocannabinol, the psychoactive constituent of cannabis, being antinociception, catalepsy, hypothermia, and depression of motor activity (Dewey, 1986; Adams et al., 1998). Although there is a history of anecdotal evidence suggesting the use of cannabinoids is effective at reducing headache and providing other pain relief, its potential as an acute migraine treatment and even preventive has never been scientifically studied in animal studies or clinical trial (Russo, 1998). However, one anonymous standardized survey found that of those using cannabis medicinally, over 10% were using it to relieve headache or migraine (Schnelle et al., 1999). Although many aspects of the study are open to debate, such as the highly selected nature of patient group, it is nevertheless an interesting observation.”

http://jpet.aspetjournals.org/content/309/1/56.long