Cannabis (Cannabis sativa)
Cannabidiol, a major phytocannabinoid, as a potent atypical inhibitor for CYP2D6.
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Δ(9)-Tetrahydrocannabinol, cannabidiol (CBD), and cannabinol are the three major cannabinoids contained in marijuana, which are devoid of nitrogen atoms in their structures. In this study, we investigated the inhibitory effects of the major phytocannabinoids on the catalytic activity of human CYP2D6. These major cannabinoids inhibited the 3-[2-(N,N-diethyl-N-methylammonium)ethyl]-7-methoxy-4-methylcoumarin (AMMC) and dextromethorphan O-demethylase activities of recombinant CYP2D6 and pooled human liver microsomes in a concentration-dependent manner (IC(50) = 4.01-24.9 μM), indicating the strongest inhibitory potency of CBD. However, these cannabinoids showed no or weak metabolism-dependent inhibition. CBD competitively inhibited the CYP2D6 activities with the apparent K(i) values of 1.16 to 2.69 μM. To clarify the structural requirement for CBD-mediated CYP2D6 inhibition, effects of CBD-related compounds on the AMMC O-demethylase activity of recombinant CYP2D6 were examined. Olivetol (IC(50) = 7.21 μM) inhibited CYP2D6 activity as potently as CBD did (IC(50) = 6.52 μM), whereas d-limonene did not show any inhibitory effect. Pentylbenzene failed to inhibit CYP2D6 activity. Furthermore, neither monomethyl nor dimethyl ethers of CBD inhibited the activity. Cannabidivarin having a propyl side chain inhibited CYP2D6 activity; its inhibitory effect (IC(50) = 10.2 μM) was less potent than that of CBD. On the other hand, orcinol and resorcinol showed lack of inhibition. The inhibitory effect of CBD on CYP2D6 activity was more potent than those of 16 compounds without nitrogen atoms tested, such as progesterone. These results indicated that CBD caused potent direct CYP2D6 inhibition, in which two phenolic hydroxyl groups and the pentyl side chain of CBD may play important roles.
1 . Recombinant: CBD-AMMC (id=NPDI-8jlF2g)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 No
Results
Table 1
Figure 3
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
2pmol
Commercially available
200 μL
30 min
MgCl2
NADPH regenerating system
0.24-15 μM
5 min
NADPH with precipitant
2 . Recombinant: CBDV-AMMC (id=NPDI-iDzluw)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 No
Results
Figure 5
Figure 4E
- Ki at log(-4.4) (concentration estimated)
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
2pmol
Commercially available
200 μL
30 min
MgCl2
NADPH regenerating system
0.24-15 μM
5 min
NADPH with precipitant
3 . Recombinant: CBN-AMMC (id=NPDI-b-kubA)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Cell fraction Pooled human liver microsomes -7999662
Results
Table 1
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
2pmol
Commercially available
200 μL
30 min
MgCl2
NADPH regenerating system
0.24-15 μM
5 min
NADPH with precipitant
4 . Recombinant: THC-AMMC (id=NPDI-XBXjow)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 No
Results
Table 1 (AMMC)
Table 3 (20 min)
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
2pmol
Commercially available
200 μL
30 min
MgCl2
NADPH regenerating system
0.24-15 μM
5 min
NADPH with precipitant
5 . Recombinant: CBD-dextromethorphan (id=NPDI-THQe_g)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 No
Results
Table 1
Figure 3
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
0.25 pmol
Commercially available
200 μL
10 min
MgCl2
NADPH regenerating system
0.15-24 μM
5 min
NADPH with precipitant
6 . pHLM: CBD-dextromethorphan (id=NPDI-ckGWRw)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Cell fraction Pooled human liver microsomes -7999662
Results
Table 1
Figure 3
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
0.25 pmol
Commercially available
200 μL
10 min
MgCl2
NADPH regenerating system
0.15-24 μM
5 min
NADPH with precipitant
7 . Recombinant: CBN-dextromethorphan (id=NPDI-0g58zw)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 No
Results
Table 1
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
0.25 pmol
Commercially available
200 μL
10 min
MgCl2
NADPH regenerating system
0.15-24 μM
5 min
NADPH with precipitant
8 . pHLM: CBN-dextromethorphan (id=NPDI-p2jqFw)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Cell fraction Pooled human liver microsomes -7999662
Results
Table 1
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
0.25 pmol
Commercially available
200 μL
10 min
MgCl2
NADPH regenerating system
0.15-24 μM
5 min
NADPH with precipitant
9 . Recombinant: THC-dextramethorphan (id=NPDI-Hsp-vg)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 No
Results
Table 1
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
0.25 pmol
Commercially available
200 μL
10 min
MgCl2
NADPH regenerating system
0.15-24 μM
5 min
NADPH with precipitant
10 . HLMs: THC-dextromethorphan (id=NPDI-KQoVKg)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Cell fraction Pooled human liver microsomes -7999662
Results
Table 1
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
0.25 pmol
Commercially available
200 μL
10 min
MgCl2
NADPH regenerating system
0.15-24 μM
5 min
NADPH with precipitant
11 . Recombinant: CBDM-AMMC (id=NPDI-y29wsw)
In Vitro Enzyme Inhibition Experiment
Negligible Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 No
Results
Figure 4C
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
2pmol
Commercially available
200 μL
30 min
MgCl2
NADPH regenerating system
0.24-15 μM
5 min
NADPH with precipitant
12 . Recombinant: CBDD-AMMC (id=NPDI-F3DjaQ)
In Vitro Enzyme Inhibition Experiment
Negligible Inhibition was detected. Cutoff used — Not specified
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 No
Results
Figure 4E
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
2pmol
Commercially available
200 μL
30 min
MgCl2
NADPH regenerating system
0.24-15 μM
5 min
NADPH with precipitant