Effects of a Commonly Occurring Genetic Polymorphism of Human CYP3A4 (I118V) on the Metabolism of Anandamide

Abstract

ABSTRACT:The endocannabinoid system plays an important role in numerous physiological processes including mood, appetite, and pain sensation. A critical compound in maintaining cannabinoid tone is the endocannabinoid anandamide (AEA). We have recently shown that AEA is metabolized by several different human cytochromes P450 (P450) to form a number of metabolites, one of which exhibits increased biological activity. CYP3A4, one of the major P450s involved in the metabolism of AEA, produces four major metabolites. One of these metabolites, 5,6-epoxyeicosatrienoic acid ethanolamide (5,6-EET-EA), exhibits a much higher affinity than AEA for the cannabinoid 2 receptor (CB-2), which leads to a marked decrease in intracellular cAMP levels in cells expressing CB-2. There are multiple human alleles of CYP3A4, and the CYP3A4.4 allele has been shown to exhibit a significant decrease in activity.Recombinant CYP3A4*4 was expressed in Escherichia coli and was demonstrated to produce 60% less 6-hydroxytestosterone than the wild-type (WT) 3A4 in a reconstituted system. The metabolism of AEA by the WT and the CYP3A4.4 variant was investigated. The mutant produced 60% less of the four EET-EA metabolites than the WT. The mutant also produced a new peak on liquid chromatography-mass spectrometry not seen with the WT, which corresponded to 19-hydroxyeicosatetraenoic acid-ethanolamide. In addition, the mutant produces four novel peaks at m/z 380, which correspond to the addition of two oxygen atoms, possibly to form a peroxide bond. These data indicate that individuals expressing the CYP3A4.4 allele may exhibit significant variations in the metabolism of AEA as well as any other compounds resembling AEA.

Functional Gene Variants of CYP3A4

“…CYP3A4*4 was identified in three subjects, *5 in two, and *6 in one of 102 Chinese individuals 41 . In an Escherichia coli in vitro expression system, the variant CYP3A4 enzymes showed reduced activity, 42 and in a clinical study, stronger lipid‐lowering effects of simvastatin were observed, although the number of subjects was extremely small 43 . It remains unclear whether CYP3A4*6 finally results in an active enzyme because the insertion of an A nucleotide causes a frameshift of the open reading frame.…”

Endocannabinoid metabolism by cytochrome P450 monooxygenases

“…CYP4F2 and CYP4F3B are expressed in the liver and kidneys, with CYP4F2 representing approximately 15% of all hepatic CYPs [127][128][129][130]. Additionally, CYP4F3B is expressed in leukocytes [131].…”

Metabolism of Anandamide by Human Cytochrome P450 2J2 in the Reconstituted System and Human Intestinal Microsomes

“…Although some of the metabolites had significantly different values for their kinetic parameters, similar deviations in these kinetic values have been seen for the metabolism of AEA by other P450s and for the different regioisomers formed during P450-mediated metabolism of AA (Daikh et al, 1994;Kiss et al, 2000;Snider et al, 2008;Pratt-Hyatt et al, 2010;Sridar et al, 2011). The metabolism of AEA by CYP2J2 is not as efficient, based on the k cat /K M values, as that by other P450s such as CYP2D6, CYP2B6, and CYP3A4 (Snider et al, 2008;Pratt-Hyatt et al, 2010;Sridar et al, 2011); however, in the presence of 50 mM AEA, the total turnover rate for the formation of all the products by recombinant CYP2J2 was calculated to be about 7 nmol of total product formed/nmol of CYP2J2/min. For comparison, in the presence of 100 mM AA, two separate laboratories have determined that the turnover for AA was 0.065 and 0.165 nmol of total product formed/nmol of CYP2J2/min King et al, 2002).…”

“…The metabolism of AEA by human P450s has been shown to give the following metabolites: 19-and 20-hydroxyeicosatetraenoic acid ethanolamides (HETE-EAs) and the 5,6-, 8,9-, 11,12-, and 14,15-epoxyeicosatrienoic acid ethanolamides (EET-EAs) (Alberich Jorda et al, 2004;Snider et al, 2007Snider et al, , 2008Snider et al, , 2009Stark et al, 2008;Pratt-Hyatt et al, 2010;Sridar et al, 2011). CYP2J2 is a P450 epoxygenase that is expressed mainly in the cardiovascular system , but it is also expressed in the intestines, stomach, and other tissues Zeldin et al, 1997b;Enayetallah et al, 2004;Paine et al, 2006).…”