Arachidonic Acid Metabolism by Human Cardiovascular CYP2J2 Is Modulated by Doxorubicin

Abstract

Doxorubicin (DOX) is a chemotherapeutic that is used in the treatment of a wide variety of cancers. However, it causes cardiotoxicity partly due to the formation of reactive oxygen species (ROS). CYP2J2 is a human cytochrome P450 that is highly expressed in cardiomyocytes. It converts arachidonic acid (AA) into four different regioisomers of epoxyeicosatrienoic acids (EETs). Using kinetic analyses we show that AA metabolism by CYP2J2 is modulated by DOX. We show that cytochrome P450 reductase (CPR), the redox partner of CYP2J2, metabolizes DOX to 7-deoxydoxorubicin aglycone (7-de-aDOX). This metabolite then binds to CYP2J2, and inhibits and alters the preferred site of metabolism of AA leading to change in the ratio of the EET regioisomers. Furthermore, molecular dynamics (MD) simulations indicate that 7-de-aDOX and AA can concurrently bind to the CYP2J2 active site to produce these changes in the site of AA metabolism. To see if these observations are unique to DOX/7-de-aDOX, we use non-cardiotoxic DOX analogues, zorubicin (ZRN) and 5-iminodaunorubicin (IDN). ZRN and 5-IDN inhibit CYP2J2-mediated AA metabolism, but does not change the ratio of EET regioisomers. Taken together, we demonstrate that DOX and 7-de-aDOX inhibit CYP2J2-mediated AA metabolism and 7-de-aDOX binds close to the active site to alter the ratio of cardioprotective EETs. These mechanistic studies of CYP2J2 can aid in the design of new alternative DOX derivatives.

Anti-inflammatory dopamine- and serotonin-based endocannabinoid epoxides reciprocally regulate cannabinoid receptors and the TRPV1 channel

“…The MD simulations support that up to three overlapping binding sites are possible, which can help to explain the observed complex AEA and eVD interactions, such as the regioselectivity change in the AEA metabolites ( Figure S29) and the potentiation of eVD metabolism (Figures 3 and 4). We have previously observed complex inhibition and regioselectivity changes for endogenous substrates of CYP2J2 59,96,97 ; however, this is the first report of a potentiation of CYP2J2 metabolism. Due to the poor solubility of the lipid substrates used in these studies, we could not accurately determine the binding constants to understand the cooperative nature of the potentiation.…”

“…In order to characterize the molecular basis of the multi-site kinetics observed with the eVDs, we performed MD simulations starting from membrane-bound structures of CYP2J2 in complex with substrates (AEA, NADA or NA5HT). Initial molecular docking was performed with AEA and either NADA or NA5HT in a stepwise manner 97 . These models allowed us to probe the binding mode of a second molecule to CYP2J2 in the presence of another molecule bound in the active site in an unbiased manner (i.e., without any assumptions about location of peripheral binding pockets).…”

Cytochrome P450 2J2: Potential Role in Drug Metabolism and Cardiotoxicity

“…It is known that the major metabolite of DOX, doxorubicinol (DOXol), contributes to toxicity in the myocardium; however, other evidence suggests other metabolites of DOX, including DOX deoxyaglycone and DOXol hydroxyaglycone, may also contribute to the cardiotoxicity (Licata et al, 2000). In addition, P450 reductase can metabolize DOX to 7-deoxydoxyrubicin aglycone, which has been shown to inhibit AA metabolism, reducing the production of EETs and altering the regiosomers of EETs produced (Arnold et al, 2017). Therefore, increased metabolism through CYP2J2 may have a conflicting effect on the heart, elevating levels of toxic metabolites for certain drugs but increasing protection through EETs.…”

An Emerging Pathway of Doxorubicin Cardiotoxicity Mediated through CYP2J2

“…4 In order to show this, we used an LC-MS/MS method to quantify the four regioisomers of EETs produced by CYP2J2 and its redox partner cytochrome P450 reductase (CPR). We found that DOX potently inhibits AA metabolism by CYP2J2 (Figure 1).…”

“…2, 5 This is the first study demonstrating that a ligand binding directly at the active site of CYP2J2 alters the site of AA metabolism. 4 …”