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The influence of monoacylglycerol lipase inhibition upon the expression of epidermal growth factor receptor in human PC-3 prostate cancer cells.

Cipriano M, Gouveia-Figueira S, Persson E, Nording M, Fowler CJ - BMC Res Notes (2014)

Bottom Line: CB1 receptor expression levels were found to differ greatly between two experimental series conducted using PC-3 cells.In the first series of experiments, JZL184 produced a small mitogenic effect for cells that had not been treated with EGF, whereas an anti-proliferative effect was seen for EGF-treated cells.An anti-proliferative effect for the EGF-treated cells was also seen with the CB receptor agonist CP55,940.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pharmacology and Clinical Neuroscience, Umeå University, Umeå, Sweden. cf@pharm.umu.se.

ABSTRACT

Background: It has been reported that direct activation of the cannabinoid CB1 receptor in epidermal growth factor (EGR)-stimulated PC-3 prostate cancer cells results in an anti-proliferative effect accompanied by a down-regulation of EGF receptors (EGFR). In the present study, we investigated whether similar effects are seen following inhibition of the endocannabinoid hydrolytic enzyme monoacylglycerol lipase (MGL).

Results: CB1 receptor expression levels were found to differ greatly between two experimental series conducted using PC-3 cells. The monoacylglycerol lipase inhibitor JZL184 increased levels of 2-arachidonoylglycerol in the PC-3 cells without producing changes in the levels of anandamide and related N-acylethanolamines. In the first series of experiments, JZL184 produced a small mitogenic effect for cells that had not been treated with EGF, whereas an anti-proliferative effect was seen for EGF-treated cells. An anti-proliferative effect for the EGF-treated cells was also seen with the CB receptor agonist CP55,940. In the second batch of cells, there was an interaction between JZL184 and CB1 receptor expression densities in linear regression analyses with EGFR expression as the dependent variable.

Conclusions: Inhibition of MGL by JZL184 can affect EGFR expression. However, the use in our hands of PC-3 cells as a model to investigate the therapeutic potential of MGL inhibitors and related compounds is compromised by their variability of CB1 receptor expression.

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Related in: MedlinePlus

Cell signalling by PC3 cells. The figure shows a FACS run for cells treated for a total of three weeks with 10 ng/ml EGF without medium change.
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Figure 3: Cell signalling by PC3 cells. The figure shows a FACS run for cells treated for a total of three weeks with 10 ng/ml EGF without medium change.

Mentions: The CB agonist CP55,940 (100 nM) was also investigated in the first series PC-3 cells, although in this case the cells were both counted by FACS and sorted on the basis of their pAkt and pErk immunoreactivities using the FlowCellectTM PI3K/MAPK Dual Pathway Activation and Cancer Marker Detection kit (Merck Millipore, Billerica, MA, USA). An example of the readout for EGF-treated cells is shown in Figure 3. As with JZL184, a significant interaction EGF: CP55,940 was found with the number of cells as endpoint (Figure 2C; two-way ANOVA matching for CP55,940: F1,8 (EGF × CP55,940) = 23.65, p < 0.005). The EGF treatment also increased the number of cells in the lower right FACS quadrant following cell sorting (i.e. high pAkt, low pErk expression) and this was also reduced by the CP55,940 treatment (Figure 4). Initial experiments indicated that the inhibition of cell proliferation in the EGF-treated cells by 100 nM CP55,940 was not blocked by the CB1 receptor inverse agonist AM251 (100 nM data not shown), suggesting that the effect of CP55,940 at this concentration may be mediated by a non-CB1 receptor pathway. Higher concentrations of AM251 were not tested, since they produce rapid anti-proliferative effects in PC-3 cells with low levels of CB1 receptor expression [9]. AM251 (admittedly at higher concentrations) also upregulates the mRNA for both EGFR and its ligands in PANC-1 pancreatic cancer cells, which lack CB1 receptors [10], so effects (or the lack of them) at higher concentrations of the compound would be very difficult to interpret.


The influence of monoacylglycerol lipase inhibition upon the expression of epidermal growth factor receptor in human PC-3 prostate cancer cells.

Cipriano M, Gouveia-Figueira S, Persson E, Nording M, Fowler CJ - BMC Res Notes (2014)

Cell signalling by PC3 cells. The figure shows a FACS run for cells treated for a total of three weeks with 10 ng/ml EGF without medium change.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4109781&req=5

Figure 3: Cell signalling by PC3 cells. The figure shows a FACS run for cells treated for a total of three weeks with 10 ng/ml EGF without medium change.
Mentions: The CB agonist CP55,940 (100 nM) was also investigated in the first series PC-3 cells, although in this case the cells were both counted by FACS and sorted on the basis of their pAkt and pErk immunoreactivities using the FlowCellectTM PI3K/MAPK Dual Pathway Activation and Cancer Marker Detection kit (Merck Millipore, Billerica, MA, USA). An example of the readout for EGF-treated cells is shown in Figure 3. As with JZL184, a significant interaction EGF: CP55,940 was found with the number of cells as endpoint (Figure 2C; two-way ANOVA matching for CP55,940: F1,8 (EGF × CP55,940) = 23.65, p < 0.005). The EGF treatment also increased the number of cells in the lower right FACS quadrant following cell sorting (i.e. high pAkt, low pErk expression) and this was also reduced by the CP55,940 treatment (Figure 4). Initial experiments indicated that the inhibition of cell proliferation in the EGF-treated cells by 100 nM CP55,940 was not blocked by the CB1 receptor inverse agonist AM251 (100 nM data not shown), suggesting that the effect of CP55,940 at this concentration may be mediated by a non-CB1 receptor pathway. Higher concentrations of AM251 were not tested, since they produce rapid anti-proliferative effects in PC-3 cells with low levels of CB1 receptor expression [9]. AM251 (admittedly at higher concentrations) also upregulates the mRNA for both EGFR and its ligands in PANC-1 pancreatic cancer cells, which lack CB1 receptors [10], so effects (or the lack of them) at higher concentrations of the compound would be very difficult to interpret.

Bottom Line: CB1 receptor expression levels were found to differ greatly between two experimental series conducted using PC-3 cells.In the first series of experiments, JZL184 produced a small mitogenic effect for cells that had not been treated with EGF, whereas an anti-proliferative effect was seen for EGF-treated cells.An anti-proliferative effect for the EGF-treated cells was also seen with the CB receptor agonist CP55,940.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pharmacology and Clinical Neuroscience, Umeå University, Umeå, Sweden. cf@pharm.umu.se.

ABSTRACT

Background: It has been reported that direct activation of the cannabinoid CB1 receptor in epidermal growth factor (EGR)-stimulated PC-3 prostate cancer cells results in an anti-proliferative effect accompanied by a down-regulation of EGF receptors (EGFR). In the present study, we investigated whether similar effects are seen following inhibition of the endocannabinoid hydrolytic enzyme monoacylglycerol lipase (MGL).

Results: CB1 receptor expression levels were found to differ greatly between two experimental series conducted using PC-3 cells. The monoacylglycerol lipase inhibitor JZL184 increased levels of 2-arachidonoylglycerol in the PC-3 cells without producing changes in the levels of anandamide and related N-acylethanolamines. In the first series of experiments, JZL184 produced a small mitogenic effect for cells that had not been treated with EGF, whereas an anti-proliferative effect was seen for EGF-treated cells. An anti-proliferative effect for the EGF-treated cells was also seen with the CB receptor agonist CP55,940. In the second batch of cells, there was an interaction between JZL184 and CB1 receptor expression densities in linear regression analyses with EGFR expression as the dependent variable.

Conclusions: Inhibition of MGL by JZL184 can affect EGFR expression. However, the use in our hands of PC-3 cells as a model to investigate the therapeutic potential of MGL inhibitors and related compounds is compromised by their variability of CB1 receptor expression.

Show MeSH
Related in: MedlinePlus