Cytochrome P450 2J2 Research Articles

Cytochrome P450 2J2*7 Single-nucleotide Polymorphism and Nocturnal Hypertension in an Elderly Turkish Population

Cytochrome P450 2J2*7 Single-nucleotide Polymorphism and Nocturnal Hypertension in an Elderly Turkish Population

Effects of tyrosine kinase inhibitors used for the treatment of non-small cell lung carcinoma on cytochrome P450 2J2 activities

Cytochrome P450 (CYP) 2J2 is responsible for the epoxidation of arachidonic acid, producing epoxyeicosatrienoic acids (EETs) that are known to enhance tumorigenesis. CYP2J2 is prominently expressed in the heart and also found in the lungs. Furthermore, the expression level of CYP2J2 in tumour tissues is higher than that in adjacent normal tissues. Non-small cell lung carcinoma is a common cancer, and tyrosine kinase inhibitors (TKIs) are powerful tools for its treatment. This study aimed to elucidate the inhibitory effects of 17 TKIs on CYP2J2 activity using LC-MS/MS. Seventeen TKIs exhibited different inhibitory effects on CYP2J2-catalysed astemizole O-demethylation in recombinant CYP2J2. Pralsetinib and selpercatinib showed strong competitive inhibition, with inhibition constant values of 0.48 and 1.1 µM, respectively. They also inhibited other CYP2J2 activities, including arachidonic acid epoxidation, hydroxyebastine carboxylation, and rivaroxaban hydroxylation. In conclusion, we showed that pralsetinib and selpercatinib strongly inhibit CYP2J2 activity. Inhibition of 14,15-EET production by these TKIs may be a novel mechanism for suppressing tumour growth and proliferation. Additionally, when these TKIs are co-administered with a CYP2J2 substrate, we may consider the possibility of drug–drug interactions via CYP2J2 inhibition.

Cytochrome P450 2J2 is required for the natural compound austocystin D to elicit cancer cell toxicity.

Austocystin D is a natural compound that induces cytochrome P450 (CYP) monooxygenase-dependent DNA damage and growth inhibition in certain cancer cell lines. Cancer cells exhibiting higher sensitivity to austocystin D often display elevated CYP2J2 expression. However, the essentiality and the role of CYP2J2 for the cytotoxicity of this compound remain unclear. In this study, we demonstrate that CYP2J2 depletion alleviates austocystin D sensitivity and DNA damage induction, while CYP2J2 overexpression enhances them. Moreover, the investigation into genes involved in austocystin D cytotoxicity identified POR and PGRMC1, positive regulators for CYP activity, and KAT7, a histone acetyltransferase. Through genetic manipulation and analysis of multiomics data, we elucidated a role for KAT7 in CYP2J2 transcriptional regulation. These findings strongly suggest that CYP2J2 is crucial for austocystin D metabolism and its subsequent cytotoxic effects. The potential use of austocystin D as a therapeutic prodrug is underscored, particularly in cancers where elevated CYP2J2 expression serves as a biomarker.

A bioluminescent probe for imaging human cytochrome P450 2J2 activity in vitro and in vivo

Human Cytochrome P450 2J2 (CYP 2J2) is closely associated with various physiological diseases. However, bioluminescence (BL) tools for monitoring CYP 2J2 activity have not yet been reported. Herein, we rationally designed a BL probe, PJ-Luc, capable of real-time imaging of CYP 2J2. In vitro studies demonstrated that PJ-Luc can effectively respond to exogenous CYP 2J2 with high selectivity and a low detection limit (LOD = 0.11 nM). Further, living cell experiments indicated that PJ-Luc can accurately quantify the ratio of endogenous CYP 2J2 amounts in HepG-2-Luc and A549-Luc cells. Moreover, an ultrahigh BL signal-to-noise ratio (S/N = 2.3×105) was achieved in tumor-bearing mouse models. These results suggest that PJ-Luc has great potential in the diagnosis of tumors and other CYP-2J2-related diseases.

CYP2J deficiency leads to cardiac injury and presents dual regulatory effects on cardiac function in rats

Cytochrome P450 2J2 (CYP2J2) enzyme is widely expressed in aortic endothelial cells and cardiac myocytes and affects cardiac function, but the underlying mechanism is still unclear. Based on CYP2J knockout (KO) rats, we have directly studied the metabolic regulation of CYP2J on cardiac function during aging. The results showed that CYP2J deficiency significantly reduced the content of epoxyeicosatrienoic acids (EETs) in plasma, aggravated myocarditis, myocardial hypertrophy, as well as fibrosis, and inhibited the mitochondrial energy metabolism signal network Pgc-1α/Ampk/Sirt1. With the increase of age, the levels of 11,12-EET and 14,15-EET in plasma of KO rats decreased significantly, and the heart injury was more serious. Interestingly, we found that after CYP2J deletion, the heart initiated a self-protection mechanism by upregulating cardiac mechanism factors Myh7, Dsp, Tnni3, Tnni2, and Scn5a, as well as mitochondrial fusion factors Mfn2 and Opa1. However, this protective effect disappeared with aging. In conclusion, CYP2J deficiency not only reduces the amount of EETs, but also plays a dual regulatory role in cardiac function.

Investigating the relevance of CYP2J2 inhibition for drugs known to cause intermediate to high risk torsades de pointes

Cardiac cytochrome P450 2J2 (CYP2J2) metabolizes endogenous polyunsaturated fatty acid, arachidonic acid (AA), to bioactive regioisomeric epoxyeicosatrienoic acid (EET) metabolites. This endogenous metabolic pathway has been postulated to play a homeostatic role in cardiac electrophysiology. However, it is unknown if drugs that cause intermediate to high risk torsades de pointes (TdP) exhibit inhibitory effects against CYP2J2 metabolism of AA to EETs. In this study, we demonstrated that 11 out of 16 drugs screened with intermediate to high risk of TdP as defined by the Comprehensive in vitro Proarrhythmia Assay (CiPA) initiative are concurrently reversible inhibitors of CYP2J2 metabolism of AA, with unbound inhibitory constant (Ki,AA,u) values ranging widely from 0.132 to 19.9 µM. To understand the physiological relevancy of Ki,AA,u, the in vivo unbound drug concentration within human heart tissue (Cu,heart) was calculated via experimental determination of in vitro unbound partition coefficient (Kpuu) for 10 CYP2J2 inhibitors using AC16 human ventricular cardiomyocytes as well as literature-derived values of fraction unbound in plasma (fu,p) and plasma drug concentrations in clinical scenarios leading to TdP. Notably, all CYP2J2 inhibitors screened belonging to the high TdP risk category, namely vandetanib and bepridil, exhibited highest Kpuu values of 18.2 ± 1.39 and 7.48 ± 1.16 respectively although no clear relationship between Cu,heart and risk of TdP could eventually be determined. R values based on basic models of reversible inhibition as per FDA guidelines were calculated using unbound plasma drug concentrations (Cu,plasma) and adapted using Cu,heart which suggested that 4 out of 10 CYP2J2 inhibitors with intermediate to high risk of TdP demonstrate greatest potential for clinically relevant in vivo cardiac drug-AA interactions. Our results shed novel insights on the relevance of CYP2J2 inhibition in drugs with risk of TdP. Further studies ascertaining the role of CYP2J2 metabolism of AA in cardiac electrophysiology, characterizing inherent cardiac ion channel activities of drugs with risk of TdP as well as in vivo evidence of drug-AA interactions will be required prior to determining if CYP2J2 inhibition could be an alternative mechanism contributing to drug-induced TdP.

Effects of genetic polymorphisms of CYP2J2, CYP2C9, CYP2C19, CYP4F2, CYP4F3 and CYP4A11 enzymes in preeclampsia and gestational hypertension

IntroductionThe aim of this study was to investigate the effects of cytochrome P450 (CYP) 2J2, CYP2C9, CYP2C19 and CYP4F2, CYP4F3 and CYP4A11 genetic polymorphisms in preeclampsia and gestational hypertension (GHT) patients in a sample of Turkish population. Materials-methodsPatients (n = 168; 110 GHT and 58 preeclampsia) and healthy pregnant women (n = 155, controls) participated in the study. For genotyping, polymerase chain reaction (PCR) and restriction analysis (RFLP) were used. Substance levels were measured using LC-MS. ResultsPlasma DHET levels in GHT and preeclampsia patients were significantly lower than those in the control group (62.7%, 66.3% vs.100.0%, respectively, p < 0.0001). An increase in CYP2J2*7 allele frequency was observed in the preeclampsia group, as compared to GHT group (12.1% vs. 4.5%; odds ratio, O.R. = 2.88, p < 0.01). The frequencies of CYP2C19*2 and*17 alleles were higher in GHT group as compared to the control group (17.7% vs. 11.6%, O.R. = 1.99, p < 0.01; and 28.6% vs.18.4%, O.R. = 2.03, p < 0.01, respectively). An increased frequency of CYP4F3 rs3794987 G allele was found in GHT group as compared to the control group (48.0% vs. 38.0%; O.R. = 1.53, p < 0.01). DiscussionDHET plasma levels were significantly reduced in hypertensive pregnant groups as compared to the control group. The allele frequency distributions for CYP2J2*7, CYP2C19 *2, *17 and CYP4F3 rs3794987 were significantly different in hypertensive pregnant patients as compared to the healthy control subjects. Our results may suggest that investigated genetic polymorphisms may be useful in diagnosis and clinical management of GHT and preeclampsia patients.

Identification of Biomarkers Associated with Heart Failure Caused by Idiopathic Dilated Cardiomyopathy Using WGCNA and Machine Learning Algorithms.

The genetic factors and pathogenesis of idiopathic dilated cardiomyopathy-induced heart failure (IDCM-HF) have not been understood thoroughly; there is a lack of specific diagnostic markers and treatment methods for the disease. Hence, we aimed to identify the mechanisms of action at the molecular level and potential molecular markers for this disease. Gene expression profiles of IDCM-HF and non-heart failure (NF) specimens were acquired from the database of Gene Expression Omnibus (GEO). We then identified the differentially expressed genes (DEGs) and analyzed their functions and related pathways by using "Metascape". Weighted gene co-expression network analysis (WGCNA) was utilized to search for key module genes. Candidate genes were identified by intersecting the key module genes identified via WGCNA with DEGs and further screened via the support vector machine-recursive feature elimination (SVM-RFE) method and the least absolute shrinkage and selection operator (LASSO) algorithm. At last, the biomarkers were validated and evaluated the diagnostic efficacy by the area under curve (AUC) value and further confirmed the differential expression in the IDCM-HF and NF groups using an external database. We detected 490 genes exhibiting differential expression between IDCM-HF and NF specimens from the GSE57338 dataset, with most of them being concentrated in the extracellular matrix (ECM) of cells related to biological processes and pathways. After screening, 13 candidate genes were identified. Aquaporin 3 (AQP3) and cytochrome P450 2J2 (CYP2J2) showed high diagnostic efficacy in the GSE57338 and GSE6406 datasets, respectively. In comparison to the NF group, AQP3 was significantly down-regulated in the IDCM-HF group, while CYP2J2 was significantly up-regulated. As far as we know, this is the first study that combines WGCNA and machine learning algorithms to screen for potential biomarkers of IDCM-HF. Our findings suggest that AQP3 and CYP2J2 could be used as novel diagnostic markers and treatment targets of IDCM-HF.

Real-time tracking of endogenous CYP2J2 by red-emitting aggregation-induced emission probe with large Stokes shift

The abnormal expression of cytochrome P450 2J2 (CYP2J2) is closely related to the proliferation of human cancer cells. In this study, a red-emitting probe with an aggregation-induced emission (AIE) effect for the quantification of CYP2J2 was developed by modifying the typical AIE fluorescent group tetraphenylethylene (TPE) through two-step synthesis. Upon addition of CYP2J2, the probe underwent O-demethylation and spontaneously 1,6-elimination reactions of p-hydroxybenzyl that allowed it to alter its photophysical properties, thus realizing light-up detection of CYP2J2 with extremely large Stokes shift (236 nm), excellent sensitivity and selectivity, low detection limit (1.09 nM). Moreover, this new AIE-active probe with good biocompatibility was admirably employed to tracking endogenous CYP2J2 in living cells.

MiR-3646 accelerates inflammatory response of Ang II-induced hVSMCs via CYP2J2/EETs axis in hypertension model

ABSTRACT Background Inflammatory response of human vascular smooth muscle cells (hVSMCs) is a driving factor in hypertension progression. It has been reported that miR-3646 was significantly up-regulated in serum samples from patients with coronary artery disease and acute myocardial infarction mice. However, its role and underlying molecular mechanism related to inflammatory response of angiotensin II (Ang II)-induced hVSMCs remain unclear. Objective We aimed to explore the potential molecular mechanisms related to inflammatory response of angiotensin II (Ang II)-induced hVSMCs. Methods Ang II–induced hypertension model was established after hVSMCs treated with 1 μM Ang II at 24 h. The interaction between microRNA 3646 (miR-3646) and cytochrome P450 2J2 (CYP2J2) was assessed by dual-luciferase reporter gene assay. MTS assay, Lipid Peroxidation MDA Assay Kit, ELISA, Western blot, and qRT-PCR were performed to examine viability, malondialdehyde (MDA) level, inflammatory cytokine levels, and the level of genes and proteins. Results Our findings illustrated that miR-3646 was up-regulated but CYP2J2 was down-regulated in Ang II–induced hVSMCs. Mechanically, miR-3646 negatively targeted to CYP2J2 in Ang II–induced hVSMCs. These findings indicated that miR-3646 regulated inflammatory response of Ang II–induced hVSMCs via targeting CYP2J2. Moreover, functional researches showed that CYP2J2 overexpression alleviated inflammatory response of Ang II–induced hVSMCs via epoxyeicosatrienoic acids/peroxisome proliferator-activated receptor-γ (EETs/PPARγ) axis, and miR-3646 aggravated inflammatory response of Ang II–induced hVSMCs via mediating CYP2J2/EETs axis. Conclusion MiR-3646 accelerated inflammatory response of Ang II–induced hVSMCs via CYP2J2/EETs axis. Our findings illustrated the specific molecular mechanism of miR-3646 regulating hypertension.

Regioselective metabolism of cannabinoids by cytochrome P450 2J2.

Cannabichromene (CBC) is a major non-psychoactive phytocannabinoid extracted from the cannabis plant. CBC has been shown to have anti-inflammatory effects; hence it can serve as a non-narcotic alternative to tetrahydrocannabinol (THC). Nevertheless, the metabolic pathways and metabolites of CBC have not been fully characterized. Metabolism of phytocannabinoids typically occurs via a superfamily of hemoproteins called cytochrome P450s (CYP450s). Our liquid chromatography-mass spectrometry (LC/MS) experiments show that CBC is metabolized to three different products by various human CYP450s. We identified that CYP450 2J2 (CYP2J2) predominantly produces 8'-hydroxy-CBC, an allylic hydroxide, out of the three metabolites. To investigate the CBC metabolic regioselectivity in CYP2J2, we leveraged atomistic molecular dynamics (MD) and modeling of the system. Using 200 unique conformations of CYP2J2 obtained from MD, we performed ensemble molecular docking of CBC to reveal three putative binding modes. We then assessed the stability of the obtained CBC binding modes by simulating them individually for 1 μs. Our simulations captured an average distance of 4.8 Å between the CYP2J2 heme and the 8’ carbon atom on CBC for one of the predominant binding modes, at which the allylic hydroxide metabolite occurs experimentally. At the regioselective binding mode, we observe conserved hydrogen bonding networks, mainly between the CBC hydroxyl group and the backbone oxygen of I487, as well as N379, stabilized by a water molecule. We further corroborated the stability of the regioselective binding modes by calculating relative binding free energies using alchemical free energy perturbation calculations. In conjunction, our computational and experimental methodologies elucidate a likely mode for regioselective binding and metabolism of CBC by CYP2J2, furthering the way towards accessible plant-based non-psychoactive anti-inflammatory therapeutics.

Neovasculogenic effect of 11,12-epoxyeicosatrienoic acid involves the Akt/eNOS signaling pathways in human endothelial progenitor cells.

The 11,12-epoxy-eicosatrienoic acid (11,12-EET) is formed from arachidonic acid (AA) by cytochrome P450 2J2 (CYP 2J2) epoxygenase and function as an effector in blood vessels. Human endothelial progenitor cells (hEPCs), a preceding cell source for endothelial cells (ECs), involve in the vascular tissue repairing by postnatal neovasculogenesis. However, the effect of 11, 12-EET on hEPCs and neovasculogenesis is not well known. In the current study, we examined the function of 11, 12-EET in hEPCs-mediated neovasculogenesis by using tubular formation analysis, Western Blotting assay, immunofluorescence staining, flow cytometry analysis and zymogram analysis. The results suggest that 11, 12-EET significantly induces neovasculogenesis through the phosphorylation of phosphoinositide 3-kinase (PI3-K)/Akt, endothelial-nitric oxide synthase (e-NOS) and extracellular signal-regulated kinase 1/2 (ERK 1/2) signaling pathways. 11, 12-EET up-regulates the expression of cyclin D1, cyclin-dependent kinase 4 (CDK4) and nuclear factor kappa B (NF-κB) proteins. Moreover, 11, 12-EET augments the expression of VE-cadherin and CD31 proteins in hEPCs. 11, 12-EET also augmented Rac1/Rho A signaling cascades, cell migration and an up-regulation of matrix metalloproteinase (MMP) -2 and -9 proteins. These results demonstrate that 11, 12-EET exerts a significant function in the neovasculogenesis of hEPCs.

The Role of Cytochrome P450 2J2 in Cancer: Cell Protector, Therapeutic Target, or Prognostic Marker?

Cytochrome P450 2J2 (CYP2J2) is one of the recently discovered enzymes that is involved in the metabolism of several drugs. It is mainly an extrahepatic enzyme which can, to some extent, explain some the drugs’ side effects such as cyclophosphamide (Cy). Cyclophosphamide is an alkylating agent that is commonly used in conditioning prior to stem cell transplantation. It is a prodrug that is bio-activated by several CYPs including CYP2J2. Cy is known to have several side effects and its extrahepatic activation by CYP2J2 may explain the mechanism of some of these side effects. On the other hand, CYP2J2 is overexpressed in several types of cancer and its inhibition (e.g. by Cy) reduces the cancer cells survival. Taken together, CYP2J2 is considered as a potential therapeutic target and prognostic marker for cancer patients in addition to its role in drugs extrahepatic metabolism.

Effects of multi-kinase inhibitors on the activity of cytochrome P450 2J2

1. Cytochrome P450 2J2 (CYP2J2) shows high expression in extrahepatic tissues, including the heart and kidney and in tumours. Inhibition of CYP2J2 has attracted attention for cancer treatment because it metabolises arachidonic acid (AA) to epoxyeicosatrienoic acid (EET), which inhibits apoptosis and promotes tumour growth. Multi-kinase inhibitor (MKI) is a molecular-targeted drug with antitumor activities. This study aimed to clarify the inhibitory effects of MKIs on CYP2J2 activity. We also investigated whether MKIs affected CYP2J2-catalysed EET formation from AA. 2. Twenty MKIs showed different inhibitory potencies against astemizole O-demethylation in CYP2J2. In particular, apatinib, motesanib, and vatalanib strongly inhibited astemizole O-demethylation. These three MKIs exhibited competitive inhibition with inhibition constant (K i) values of 9.3, 15.4, and 65.0 nM, respectively. Apatinib, motesanib, and vatalanib also inhibited CYP2J2-catalysed 14,15-EET formation from AA. 3. In simulations of docking to CYP2J2, the U energy values of apatinib, motesanib, and vatalanib were low, and measured −84.5, −69.9, and −52.3 kcal/mol, respectively. 4. In conclusion, apatinib, motesanib, and vatalanib strongly inhibited CYP2J2 activity, suggesting that the effects of a given CYP2J2 substrate may be altered upon the administration of these MKIs.

Profiling the Cytochrome P450 2J2 Active Site

Human cytochrome P450 2J2 (CYP2J2) is a membrane bound, heme‐containing monooxygenase primary expressed in the heart. This enzyme oxidizes the double bonds in polyunsaturated fatty acids such as arachidonic acid and linoleic acid to epoxides, generating epoxyeicosatrienoic acids (EETs). Various EET products are implicated in cardioprotective effects, but also the promotion of cancer tumor growth and metastasis. Additionally, CYP2J2 can act on a range of pharmaceuticals. Thus, CYP2J2 is of clinical interest, but there is no structural information to understand how the CYP2J2 active site accommodates this wide variety of ligands.In this study, the CYP2J2 active site was defined by its ability to bind an array of &gt;75 substrates, inhibitors, and xenobiotics, including a broad panel of azole compounds. The binding modes and affinities were determined by monitoring the shifts of the CYP2J2 heme Soret absorbance, which is indicative of heme environment perturbation. From these experiments the following spectral shift responses were observed: displacement of the heme‐coordinated water (type‐I binding mode), compound coordination to the heme iron (type‐II), a red‐shifted type‐I binding mode, compound coordination to heme coordinated water (reverse type‐I binding mode), or no spectral shift. As the latter result can be consistent with no binding or binding in the active site in a way that does not perturb the heme center, such compounds were investigated via a luminescence‐based inhibition assay to verify their interaction with CYP2J2. As imidazole‐containing ligands provide a fixed, known reference point for their interaction with the CYP2J2 heme iron, cheminformatics approaches were applied to these compounds to help provide insight into the chemical makeup of the CYP2J2 active site. The physical properties of these ligands were analyzed to identify key ligand features that drive CYP2J2 ligand preference. In the absence of protein structure information, this strategy provides a route to defining the CYP2J2 ligand binding site, potentially supporting the development of selective inhibitors that can help define the role of CYP2J2 modulation in vivo and potentially facilitate therapeutic strategies.

Site-directed deuteration of dronedarone preserves cytochrome P4502J2 activity and mitigates its cardiac adverse effects in canine arrhythmic hearts.

Cytochrome P4502J2 (CYP2J2) metabolizes arachidonic acid (AA) to cardioprotective epoxyeicosatrienoic acids (EETs). Dronedarone, an antiarrhythmic drug prescribed for treatment of atrial fibrillation (AF) induces cardiac adverse effects (AEs) with poorly understood mechanisms. We previously demonstrated that dronedarone inactivates CYP2J2 potently and irreversibly, disrupts AA-EET pathway leading to cardiac mitochondrial toxicity rescuable via EET enrichment. In this study, we investigated if mitigation of CYP2J2 inhibition prevents dronedarone-induced cardiac AEs. We first synthesized a deuterated analogue of dronedarone (termed poyendarone) and demonstrated that it neither inactivates CYP2J2, disrupts AA-EETs metabolism nor causes cardiac mitochondrial toxicity in vitro. Our patch-clamp experiments demonstrated that pharmacoelectrophysiology of dronedarone is unaffected by deuteration. Next, we show that dronedarone treatment or CYP2J2 knockdown in spontaneously beating cardiomyocytes indicative of depleted CYP2J2 activity exacerbates beat-to-beat (BTB) variability reflective of proarrhythmic phenotype. In contrast, poyendarone treatment yields significantly lower BTB variability compared to dronedarone in cardiomyocytes indicative of preserved CYP2J2 activity. Importantly, poyendarone and dronedarone display similar antiarrhythmic properties in the canine model of persistent AF, while poyendarone substantially reduces beat-to-beat variability of repolarization duration suggestive of diminished proarrhythmic risk. Our findings prove that deuteration of dronedarone prevents CYP2J2 inactivation and mitigates dronedarone-induced cardiac AEs.

Linezolid Metabolism Is Catalyzed by Cytochrome P450 2J2, 4F2, and 1B1.

The oxazolidinone antibacterial linezolid has been in clinical use for over 20 years, yet knowledge of the contributions of specific cytochrome (CYP) 450 enzymes to the metabolic clearance of this drug were mostly unknown. In this investigation, it was revealed that three P450 enzymes that had not been previously explored in linezolid metabolism, CYP2J2, CYP4F2, and CYP1B1, catalyzed the 2-hydroxylation and de-ethyleneation of the morpholine moiety of linezolid. The intrinsic clearance for linezolid metabolism in pooled human liver microsomes was low at 0.51 μL/min/mg protein, consistent with its in vivo clearance in humans, and the KM was high (>200 μM). In recombinant human P450 enzymes, a rank order of intrinsic clearance values for linezolid 2-hydroxylation were CYP2J2 ≫ CYP4F2 > CYP2C8 > CYP1B1 ≈ CYP2D6 ≈ CYP3A4 > CYP1A1 > CYP3A5, with nine other P450 enzymes showing no linezolid metabolism. The effect of selective inhibitors for these eight P450 enzymes on linezolid metabolism in pooled human liver microsomes was evaluated to provide estimates of the relative fractional contributions of these enzymes to linezolid metabolism. These experiments suggest that CYP2J2 and CYP4F2 contribute about 50% each to linezolid hepatic metabolism. It is proposed that the oxidative metabolic clearance of linezolid is primarily catalyzed by these two unusual P450 enzymes and that this explains the lack of observation of meaningful effects of common perpetrators of drug interactions on linezolid pharmacokinetics. SIGNIFICANCE STATEMENT: Linezolid is an important antibacterial drug, but the enzymes involved in its oxidative metabolism were unknown. In this study, evidence is shown that supports an important role for two enzymes not frequently associated with the metabolism of drugs: cytochrome P450 2J2 and cytochrome P450 4F2. These observations offer insight to understand the results of clinical drug-drug interaction studies conducted on linezolid.

Anthracycline derivatives inhibit cardiac CYP2J2

Anthracycline chemotherapeutics are highly effective, but their clinical usefulness is hampered by adverse side effects such as cardiotoxicity. Cytochrome P450 2J2 (CYP2J2) is a cytochrome P450 epoxygenase in human cardiomyocytes that converts arachidonic acid (AA) to cardioprotective epoxyeicosatrienoic acid (EET) regioisomers. Herein, we performed biochemical studies to understand the interaction of anthracycline derivatives (daunorubicin, doxorubicin, epirubicin, idarubicin, 5-iminodaunorubicin, zorubicin, valrubicin, and aclarubicin) with CYP2J2. We utilized fluorescence polarization (FP) to assess whether anthracyclines bind to CYP2J2. We found that aclarubicin bound the strongest to CYP2J2 despite it having large bulky groups. We determined that ebastine competitively inhibits anthracycline binding, suggesting that ebastine and anthracyclines may share the same binding site. Molecular dynamics and ensemble docking revealed electrostatic interactions between the anthracyclines and CYP2J2, contributing to binding stability. In particular, the glycosamine groups in anthracyclines are stabilized by binding to glutamate and aspartate residues in CYP2J2 forming salt bridge interactions. Furthermore, we used iterative ensemble docking schemes to gauge anthracycline influence on EET regioisomer production and anthracycline inhibition on AA metabolism. This was followed by experimental validation of CYP2J2-mediated metabolism of anthracycline derivatives using liquid chromatography tandem mass spectrometry fragmentation analysis and inhibition of CYP2J2-mediated AA metabolism by these derivatives. Taken together, we use both experimental and theoretical methodologies to unveil the interactions of anthracycline derivatives with CYP2J2. These studies will help identify alternative mechanisms of how anthracycline cardiotoxicity may be mediated through the inhibition of cardiac P450, which will aid in the design of new anthracycline derivatives with lower toxicity.

Polymorphisms in EPHX2 and CYP2J2 moderate associations between white matter hyperintensities and temporal lobe atrophy.

White matter hyperintensities (WMH) of presumed vascular origin commonly coincide with neurodegeneration. Previous studies have reported heterogeneous relationships between WMH and atrophy. Cytochrome P450 (CYP) 2J2 (CYP2J2) is involved in the vascular ischemic response and soluble epoxide hydrolase (EPHX2) metabolizes its products, which have been implicated in vascular pathology and WMH. Here we investigate the potential moderation effect of CYP2J2 and EPHX2 single nucleotide polymorphisms (SNPs), individually and interactively, on the association between WMH and atrophy. Patients with vascular cognitive impairment or neurodegenerative diagnoses (n=447) from the Sunnybrook Dementia Study (NCT01800214), Vascular Brain Health (VBH) study, and the Brain-Eye Amyloid Memory (BEAM) study were genotyped for SNPs in EPHX2 and CYP2J2. SNPs were genotyped by the Illumina Neurochip and were selected a priori based on literature. All participants were confirmed to have European ancestry via multi-dimensional scaling using the genotype data. Imaging analysis was performed on 3D T1 and T2/PD weighted images for brain segmentation and volumetric acquisition using SABRE, and WMH volumes were acquired on T2/PD/FLAIR images using Lesion Explorer. Linear regression models controlling for age, sex, Mini Mental State Exam, and head size were used to assess moderation effects of the SNPs (WMH×SNP interactions) on associations between WMH and temporal atrophy using SPSS (v.26). Presence of minor alleles were assessed using a dominant genetic model. The intron variant EPHX2 rs7816586 G/A polymorphism had a trending moderation effect on the association between periventricular WMH and left temporal atrophy where the minor A-allele (frequency=11.52%) was protective (F=3.958, p=0.047) in extensive WMH. The promoter variant CYP2J2 rs10889162 C/T had a significant moderation effect on the association between deep WMH and right temporal atrophy where the minor T-allele (frequency=8.61%) was protective in cases of extensive WMH (F=5.771, p=0.017). No interaction between the EPHX2 and CYP2J2 SNPs was detected. We identified potential moderation effects of genetic variation in EPHX2 and CYP2J2 on the association between WMH and temporal lobe atrophy across neurodegenerative diagnoses. The lack of SNP×SNP interactions suggests that these genes may act independently, consistent with WMH×SNP interaction effects that occurred in association with different anatomical WMH locations.

Cytochrome P450 2J2 inhibits the proliferation and angiogenesis of retinal vascular endothelial cells by regulating the Notch signaling pathway in a hypoxia-induced retinopathy model

ABSTRACT Retinopathy of Prematurity (ROP), a type of retinal neovascularization in premature infants, has become a serious problem that drastically affects the quality of life of premature infants. ROP is associated with angiogenesis and neovascularization. Here, we aimed to explain the function and latent roles of Cytochrome P450 2J2 (CYP2J2) in hypoxia-induced retinopathy in retinal vascular endothelial cells (HRVECs). HRVECs were stimulated with hypoxia for 24 h to establish an in vitro retinopathy model. Cell viability and migration were evaluated using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and Transwell assays, respectively. Protein and gene expression was determined by reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) and Western blot analysis. We observed that pcDNA3.1(+)-CYP2J2 promoted CYP2J2 and Jagged1 expression, while Dll4 was down-regulated in hypoxia-stimulated HRVECs. Additionally, pcDNA3.1(+)-CYP2J2 inhibited HRVEC viability, reduced PCNA expression, and inhibited the migration of HRVECs. Further, the Notch pathway was inhibited in the Hypoxia+pcDNA3.1(+)-CYP2J2 group. Opposite results were observed upon Terfenadone treatment in hypoxia induced HRVECs. Finally, our findings further verified that DAPT promotes the effects of CYP2J2 on cell viability, migration, and Notch signaling in hypoxia-induced HRVECs, while EDTA reversed the inhibitory effects of CYP2J2 on hypoxia-induced HRVECs. In conclusions, CYP2J2 was found to inhibit the viability and angiogenesis of HRVECs by inhibiting Notch signaling in a hypoxia-induced retinopathy model.