Publications by authors named "Thomas Duflot"

25 Publications

  • Page 1 of 1

Soluble Epoxide Hydrolase Inhibition Prevents Experimental Type 4 Cardiorenal Syndrome.

Front Mol Biosci 2020 11;7:604042. Epub 2021 Mar 11.

Normandie University, UNIROUEN, INSERM U1096, FHU REMOD-VHF, Rouen, France.

Cardiovascular diseases (CVD) remain the leading cause of morbimortality in patients with chronic kidney disease (CKD). The aim of this study was to assess the cardiovascular impact of the pharmacological inhibition of soluble epoxide hydrolase (sEH), which metabolizes the endothelium-derived vasodilatory and anti-inflammatory epoxyeicosatrienoic acids (EETs) to dihydroxyeicosatrienoic acid (DHETs), in the 5/6 nephrectomy (Nx) mouse model. Compared to sham-operated mice, there was decrease in EET-to-DHET ratio 3 months after surgery in vehicle-treated Nx mice but not in mice treated with the sEH inhibitor -AUCB. Nx induced an increase in plasma creatinine and in urine albumin-to-creatinine ratio as well as the development of kidney histological lesions, all of which were not modified by -AUCB. In addition, -AUCB did not oppose Nx-induced blood pressure increase. However, AUCB prevented the development of cardiac hypertrophy and fibrosis induced by Nx, as well as normalized the echocardiographic indices of diastolic and systolic function. Moreover, the reduction in endothelium-dependent flow-mediated dilatation of isolated mesenteric arteries induced by Nx was blunted by -AUCB without change in endothelium-independent dilatation to sodium nitroprusside. Inhibition of sEH reduces the cardiac remodelling, and the diastolic and systolic dysfunctions associated with CKD. These beneficial effects may be mediated by the prevention of endothelial dysfunction, independent from kidney preservation and antihypertensor effect. Thus, inhibition of sEH holds a therapeutic potential in preventing type 4 cardiorenal syndrome.
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http://dx.doi.org/10.3389/fmolb.2020.604042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7991096PMC
March 2021

Preservation of epoxyeicosatrienoic acid bioavailability prevents renal allograft dysfunction and cardiovascular alterations in kidney transplant recipients.

Sci Rep 2021 Feb 12;11(1):3739. Epub 2021 Feb 12.

Department of Pharmacology, Rouen University Hospital, 76000, Rouen, France.

This study addressed the hypothesis that epoxyeicosatrienoic acids (EETs) synthesized by CYP450 and catabolized by soluble epoxide hydrolase (sEH) are involved in the maintenance of renal allograft function, either directly or through modulation of cardiovascular function. The impact of single nucleotide polymorphisms (SNPs) in the sEH gene EPHX2 and CYP450 on renal and vascular function, plasma levels of EETs and peripheral blood monuclear cell sEH activity was assessed in 79 kidney transplant recipients explored at least one year after transplantation. Additional experiments in a mouse model mimicking the ischemia-reperfusion (I/R) injury suffered by the transplanted kidney evaluated the cardiovascular and renal effects of the sEH inhibitor t-AUCB administered in drinking water (10 mg/l) during 28 days after surgery. There was a long-term protective effect of the sEH SNP rs6558004, which increased EET plasma levels, on renal allograft function and a deleterious effect of K55R, which increased sEH activity. Surprisingly, the loss-of-function CYP2C9*3 was associated with a better renal function without affecting EET levels. R287Q SNP, which decreased sEH activity, was protective against vascular dysfunction while CYP2C8*3 and 2C9*2 loss-of-function SNP, altered endothelial function by reducing flow-induced EET release. In I/R mice, sEH inhibition reduced kidney lesions, prevented cardiac fibrosis and dysfunction as well as preserved endothelial function. The preservation of EET bioavailability may prevent allograft dysfunction and improve cardiovascular disease in kidney transplant recipients. Inhibition of sEH appears thus as a novel therapeutic option but its impact on other epoxyfatty acids should be carefully evaluated.
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http://dx.doi.org/10.1038/s41598-021-83274-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7881112PMC
February 2021

Fostering global data sharing: highlighting the recommendations of the Research Data Alliance COVID-19 working group.

Wellcome Open Res 2020 9;5:267. Epub 2020 Nov 9.

Laurentian University, Ontario, P3E 2C6, Canada.

The systemic challenges of the COVID-19 pandemic require cross-disciplinary collaboration in a global and timely fashion. Such collaboration needs open research practices and the sharing of research outputs, such as data and code, thereby facilitating research and research reproducibility and timely collaboration beyond borders. The Research Data Alliance COVID-19 Working Group recently published a set of recommendations and guidelines on data sharing and related best practices for COVID-19 research. These guidelines include recommendations for researchers, policymakers, funders, publishers and infrastructure providers from the perspective of different domains (Clinical Medicine, Omics, Epidemiology, Social Sciences, Community Participation, Indigenous Peoples, Research Software, Legal and Ethical Considerations). Several overarching themes have emerged from this document such as the need to balance the creation of data adherent to FAIR principles (findable, accessible, interoperable and reusable), with the need for quick data release; the use of trustworthy research data repositories; the use of well-annotated data with meaningful metadata; and practices of documenting methods and software. The resulting document marks an unprecedented cross-disciplinary, cross-sectoral, and cross-jurisdictional effort authored by over 160 experts from around the globe. This letter summarises key points of the Recommendations and Guidelines, highlights the relevant findings, shines a spotlight on the process, and suggests how these developments can be leveraged by the wider scientific community.
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http://dx.doi.org/10.12688/wellcomeopenres.16378.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7808050PMC
November 2020

Pharmacogenetics of Direct Oral Anticoagulants: A Systematic Review.

J Pers Med 2021 Jan 11;11(1). Epub 2021 Jan 11.

Laboratory of Pharmacology, Toxicology and Pharmacogenetic, Pharmacology Department, Rouen University Hospital, 76031 Rouen, France.

Dabigatran, rivaroxaban, apixaban, edoxaban, and betrixaban are direct oral anticoagulants (DOACs). Their inter-individual variability in pharmacodynamics and pharmacokinetics (transport and metabolism) is high, and could result from genetic polymorphisms. As recommended by the French Network of Pharmacogenetics (RNPGx), the management of some treatments in cardiovascular diseases (as antiplatelet agents, oral vitamin K antagonists, and statins) can rely on genetic testing in order to improve healthcare by reducing therapeutic resistance or toxicity. This paper is a review of association studies between single nucleotide polymorphisms (SNPs) and systemic exposure variation of DOACs. Most of the results presented here have a lot to do with some SNPs of CES1 (rs2244613, rs8192935, and rs71647871) and ABCB1 (rs1128503, rs2032582, rs1045642, and rs4148738) genes, and dabigatran, rivaroxaban, and apixaban. Regarding edoxaban and betrixaban, as well as SNPs in the and genes, literature is scarce, and further studies are needed.
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http://dx.doi.org/10.3390/jpm11010037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7826504PMC
January 2021

Ten-year Evaluation of a Large Retrospective Cohort Treated by Sacral Nerve Modulation for Fecal Incontinence: Results of a French Multicenter Study.

Ann Surg 2020 Jul 24. Epub 2020 Jul 24.

Normandie Univ, UNIROUEN, CHU Rouen, Department of Digestive Physiology and CIC-CRB 1404, Rouen, France.

Objective: The aim of this study was to assess the effectiveness of sacral nerve modulation (SNM) in a large cohort of patients implanted for at least 10 years, quantify adverse event rates, and identify predictive factors of long-term success.

Summary Background Data: Few studies have evaluated the long-term success of SNM.

Methods: Data collected prospectively from patients implanted for fecal incontinence (FI) in 7 French centers between January 1998 and December 2008 were retrospectively analyzed. Patient FI severity scores were assessed before and 10 years after implantation. The main evaluation criterion was the success of SNM defined by the continuation of the treatment without additional therapies. The secondary evaluation criteria were the rate of device revisions and explantations. Preoperative predictors of success at 10 years were sought.

Results: Of the 360 patients (27 males, mean age: 59 ± 12 years) implanted for FI, 162 (45%) had a favorable outcome 10 years post-implantation, 115 (31.9%) failed, and 83 (23.1%) were lost to follow-up. The favorable outcome derived from the time-to-event Kaplan-Meier curve at 10 years was 0.64 (95% CI 0.58-0.69). FI severity scores were significantly better 10 years post-implantation compared to preimplantation (7.4 ± 4.3 vs 14.0 ± 3.2; P < 0.0001). During the 10-year follow-up, 233 patients (64.7%) had a surgical revision and 94 (26.1%) were explanted. A history of surgery for FI and sex (male) were associated with an increased risk of an unfavorable outcome.

Conclusions: Long-term efficacy was maintained in approximately half of the FI patients treated by SNM at least 10 years post-implantation.
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http://dx.doi.org/10.1097/SLA.0000000000004251DOI Listing
July 2020

The Metabolism of Epoxyeicosatrienoic Acids by Soluble Epoxide Hydrolase Is Protective against the Development of Vascular Calcification.

Int J Mol Sci 2020 Jun 17;21(12). Epub 2020 Jun 17.

Department of Pharmacology, Rouen University Hospital, CEDEX 1, 76031 Rouen, France.

This study addressed the hypothesis that soluble epoxide hydrolase (sEH), which metabolizes endothelium-derived epoxyeicosatrienoic acids, plays a role in vascular calcification. The sEH inhibitor -4-(4-(3-adamantan-1-yl-ureido)-cyclohexyloxy)-benzoic acid (-AUCB) potentiated the increase in calcium deposition of rat aortic rings cultured in high-phosphate conditions. This was associated with increased tissue-nonspecific alkaline phosphatase activity and mRNA expression level of the osteochondrogenic marker Runx2. The procalcifying effect of -AUCB was prevented by mechanical aortic deendothelialization or inhibition of the production and action of epoxyeicosatrienoic acids using the cytochrome P450 inhibitor fluconazole and the antagonist 14,15-epoxyeicosa-5(Z)-enoic acid (14,15-EEZE), respectively. Similarly, exogenous epoxyeicosatrienoic acids potentiated the calcification of rat aortic rings through a protein kinase A (PKA)-dependent mechanism and of human aortic vascular smooth muscle cells when sEH was inhibited by -AUCB. Finally, a global gene expression profiling analysis revealed that the mRNA expression level of sEH was decreased in human carotid calcified plaques compared to adjacent lesion-free sites and was inversely correlated with Runx2 expression. These results show that sEH hydrolase plays a protective role against vascular calcification by reducing the bioavailability of epoxyeicosatrienoic acids.
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http://dx.doi.org/10.3390/ijms21124313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7352784PMC
June 2020

Development of Improved Double-Nanobody Sandwich ELISAs for Human Soluble Epoxide Hydrolase Detection in Peripheral Blood Mononuclear Cells of Diabetic Patients and the Prefrontal Cortex of Multiple Sclerosis Patients.

Anal Chem 2020 05 27;92(10):7334-7342. Epub 2020 Apr 27.

Department of Entomology and Nematology and UCD Comprehensive Cancer Center, University of California, Davis, California 95616, United States.

Nanobodies have been progressively replacing traditional antibodies in various immunological methods. However, the use of nanobodies as capture antibodies is greatly hampered by their poor performance after passive adsorption to polystyrene microplates, and this restricts the full use of double nanobodies in sandwich enzyme-linked immunosorbent assays (ELISAs). Herein, using the human soluble epoxide hydrolase (sEH) as a model analyte, we found that both the immobilization format and the blocking agent have a significant influence on the performance of capture nanobodies immobilized on polystyrene and the subsequent development of double-nanobody sandwich ELISAs. We first conducted epitope mapping for pairing nanobodies and then prepared a horseradish-peroxidase-labeled nanobody using a mild conjugation procedure as a detection antibody throughout the work. The resulting sandwich ELISA using a capture nanobody (A9, 1.25 μg/mL) after passive adsorption and bovine serum albumin (BSA) as a blocking agent generated a moderate sensitivity of 0.0164 OD·mL/ng and a limit of detection (LOD) of 0.74 ng/mL. However, the introduction of streptavidin as a linker to the capture nanobody at the same working concentration demonstrated a dramatic 16-fold increase in sensitivity (0.262 OD·mL/ng) and a 25-fold decrease in the LOD for sEH (0.03 ng/mL). The streptavidin-bridged double-nanobody ELISA was then successfully applied to tests for recovery, cross-reactivity, and real samples. Meanwhile, we accidentally found that blocking with skim milk could severely damage the performance of the capture nanobody by an order of magnitude compared with BSA. This work provides guidelines to retain the high effectiveness of the capture nanobody and thus to further develop the double-nanobody ELISA for various analytes.
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http://dx.doi.org/10.1021/acs.analchem.0c01115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7744119PMC
May 2020

Exploring the microvascular impact of red blood cell transfusion in intensive care unit patients.

Crit Care 2019 Aug 30;23(1):292. Epub 2019 Aug 30.

Assistance Publique-Hôpitaux de Paris (AP-HP), Hôpital Saint-Antoine, Service de Réanimation Médicale, 184 rue du Faubourg Saint-Antoine, 75571, Paris Cedex 12, France.

Background: Red blood cell (RBC) transfusion is a common treatment for hospitalized patients. However, the effects of RBC transfusion on microvascular function remain controversial.

Methods: In a medical ICU in a tertiary teaching hospital, we prospectively included anemic patients requiring RBC transfusion. Skin microvascular reactivity was measured before and 30 min after RBC transfusion. Plasma was collected to analyze intravascular hemolysis and draw the lipidomic and cytokine profiles.

Results: In a cohort of 59 patients, the median age was 66 [55-81] years and SAPS II was 38 [24-48]. After RBC transfusion, endothelium-dependent microvascular reactivity improved in 35 (59%) patients, but worsened in 24 others (41%). Comparing clinical and biological markers revealed that baseline blood leucokyte counts distinguished improving from worsening patients (10.3 [5.7; 19.7] vs. 4.6 [2.1; 7.3] × 10/L; p = 0.001) and correlated with variations of microvascular reactivity (r = 0.36, p = 0.005). Blood platelet count was also higher in improving patients (200 [97; 280] vs 160 [40; 199] × 10/mL, p = 0.03) but did not correlate with variations of microvascular reactivity. We observed no intravascular hemolysis (HbCO, heme, bilirubin, LDH), but recorded a significant increase in RBC microparticle levels specific to improving patients after transfusion (292 [108; 531] vs. 53 [34; 99] MP/μL; p = 0.03). The improvement in microvascular dilation was positively correlated with RBC microparticle levels (R = 0.83, p < 0.001) and conversion of arachidonic acid into vasodilating eicosanoids.

Conclusions: Patients displaying an improved microvascular reactivity after RBC transfusion had high blood leukocyte counts, increased RBC microparticle formation, and enhanced metabolism of arachidonic acid into vasodilating lipids. Our data suggested a contribution of recipient leukocytes to the vascular impact of RBC transfusion.
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http://dx.doi.org/10.1186/s13054-019-2572-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6717366PMC
August 2019

Discovery of the First in Vivo Active Inhibitors of the Soluble Epoxide Hydrolase Phosphatase Domain.

J Med Chem 2019 09 17;62(18):8443-8460. Epub 2019 Sep 17.

Institute of Pharmaceutical Chemistry , Goethe-University Frankfurt , Max-von-Laue-Strasse 9 , 60438 Frankfurt am Main , Germany.

The emerging pharmacological target soluble epoxide hydrolase (sEH) is a bifunctional enzyme exhibiting two different catalytic activities that are located in two distinct domains. Although the physiological role of the C-terminal hydrolase domain is well-investigated, little is known about its phosphatase activity, located in the N-terminal phosphatase domain of sEH (sEH-P). Herein we report the discovery and optimization of the first inhibitor of human and rat sEH-P that is applicable in vivo. X-ray structure analysis of the sEH phosphatase domain complexed with an inhibitor provides insights in the molecular basis of small-molecule sEH-P inhibition and helps to rationalize the structure-activity relationships. 4-(4-(3,4-Dichlorophenyl)-5-phenyloxazol-2-yl)butanoic acid (, SWE101) has an excellent pharmacokinetic and pharmacodynamic profile in rats and enables the investigation of the physiological and pathophysiological role of sEH-P in vivo.
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http://dx.doi.org/10.1021/acs.jmedchem.9b00445DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7262789PMC
September 2019

Impact of the acute local inhibition of soluble epoxide hydrolase on diabetic skin microcirculatory dysfunction.

Diab Vasc Dis Res 2019 11 3;16(6):523-529. Epub 2019 Jul 3.

Department of Pharmacology, Rouen University Hospital, Rouen, France.

The impact of the local inhibition of soluble epoxide hydrolase, which metabolizes vasodilator and anti-inflammatory epoxyeicosanoids, on diabetic skin microvascular dysfunction was assessed. In diabetic db/db mice, basal skin blood flow assessed using laser Doppler imaging was similar to that of control mice, but thermal hyperemia was markedly reduced. At 2 h after the topical administration of an aqueous gel containing the soluble epoxide hydrolase inhibitor trans-4-[4-(3-adamantan-1-yl-ureido)-cyclohexyloxy]-benzoic acid (t-AUCB: 400 mg/L), the peak concentration of t-AUCB was detected in the skin of diabetic mice, which quickly decreased thereafter. In parallel, 2 h after application of t-AUCB treatment, thermal hyperemia was increased compared to the control gel. Quantification of t-AUCB in plasma of treated animals showed no or low systemic diffusion. Furthermore, haematoxylin and eosin histological staining of skin biopsies showed that skin integrity was preserved in t-AUCB-treated mice. Finally, for pig ear skin, a surrogate for human skin, using Franz diffusion cells, we observed a continuous diffusion of t-AUCB from 2 h after application to beyond 24 h. A single topical administration of a soluble epoxide hydrolase inhibitor improves microcirculatory function in the skin of db/db mice and might represent a new therapeutic approach for preventing the development of skin complications in diabetic patients.
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http://dx.doi.org/10.1177/1479164119860215DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7307659PMC
November 2019

Interest of adjusting urine cannabinoids to creatinine level to monitor cannabis cessation therapy in heavy smokers with psychiatric disorders.

Drug Test Anal 2019 Sep 25;11(9):1453-1459. Epub 2019 Jul 25.

Laboratory of Pharmacokinetics, Toxicology and Pharmacogenomics, Rouen University Hospital, 76000, Rouen, France.

Up to 25% of hospitalized patients in a psychiatric department exhibit troubles linked to cannabis use. Weaning patients with psychiatric disorders off drugs of abuse requires specific care to improve their clinical outcome. The present study aims to develop a predictive model of urinary excretion of creatinine-normalized cannabinoids (U ) and to determine U thresholds corresponding to the widely used cut-offs of 20 ng/mL and 50 ng/mL for cannabinoids. One hundred thirty-two patients with 452 urine samples were included between 2013 and 2017. Urinary cannabinoids and U elimination curves were computed for each patient. Using a mono-exponential mixed effect model with 88 samples from 26 subjects exhibiting at least 3 decreasing U in a row, the average calculated elimination rate constant was 0.0108 ± 0.0026 h , corresponding to a mean elimination half-life of 64 ± 12 hours. The use of U is of particular interest because of a high inter- and intra-individual variability of urinary creatinine concentration (from 0.06 to 3.81 mg/mL). Moreover, U allows for the detection of diluted or concentrated urine specimens that may lead to false positive (FP) or false negative (FN) results. Receiver operator characteristic (ROC) curves were used to assess U thresholds of 32.4 and 124.7 ng/mg that provide a strong discrimination between positive and negative samples for cannabinoids cut-offs of 20 and 50 ng/mL respectively. The developed model and the defined U thresholds allowed for the accurate prediction of the time needed to reach a negative U result that could be used by clinicians to optimize clinical care.
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http://dx.doi.org/10.1002/dta.2672DOI Listing
September 2019

Altered bioavailability of epoxyeicosatrienoic acids is associated with conduit artery endothelial dysfunction in type 2 diabetic patients.

Cardiovasc Diabetol 2019 03 18;18(1):35. Epub 2019 Mar 18.

Department of Pharmacology, Rouen University Hospital, 76000, Rouen Cedex, France.

Background: This pathophysiological study addressed the hypothesis that soluble epoxide hydrolase (sEH), which metabolizes the vasodilator and anti-inflammatory epoxyeicosatrienoic acids (EETs) to dihydroxyeicosatrienoic acids (DHETs), contributes to conduit artery endothelial dysfunction in type 2 diabetes.

Methods And Results: Radial artery endothelium-dependent flow-mediated dilatation in response to hand skin heating was reduced in essential hypertensive patients (n = 9) and type 2 diabetic subjects with (n = 19) or without hypertension (n = 10) compared to healthy subjects (n = 36), taking into consideration cardiovascular risk factors, flow stimulus and endothelium-independent dilatation to glyceryl trinitrate. Diabetic patients but not non-diabetic hypertensive subjects displayed elevated whole blood reactive oxygen species levels and loss of NO release during heating, assessed by measuring local plasma nitrite variation. Moreover, plasma levels of EET regioisomers increased during heating in healthy subjects, did not change in hypertensive patients and decreased in diabetic patients. Correlation analysis showed in the overall population that the less NO and EETs bioavailability increases during heating, the more flow-mediated dilatation is reduced. The expression and activity of sEH, measured in isolated peripheral blood mononuclear cells, was elevated in diabetic but not hypertensive patients, leading to increased EETs conversion to DHETs. Finally, hyperglycemic and hyperinsulinemic euglycemic clamps induced a decrease in flow-mediated dilatation in healthy subjects and this was associated with an altered EETs release during heating.

Conclusions: These results demonstrate that an increased EETs degradation by sEH and altered NO bioavailability are associated with conduit artery endothelial dysfunction in type 2 diabetic patients independently from their hypertensive status. The hyperinsulinemic and hyperglycemic state in these patients may contribute to these alterations. Trial registration NCT02311075. Registered December 8, 2014.
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http://dx.doi.org/10.1186/s12933-019-0843-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6423843PMC
March 2019

Thrombin generation profile in non-thrombotic factor V Leiden carriers.

J Thromb Thrombolysis 2019 Apr;47(3):473-477

Normandie Univ, UNIROUEN, INSERM U1096, Rouen University Hospital, Vascular Hemostasis Unit, F 76000, Rouen, France.

Factor V Leiden (FVL) mutation is the most common genetic risk factor for venous thromboembolism. In families with a history of thrombosis, FVL can be present in 18%. Thrombin generation test is commonly used as an evaluation tool of thrombotic risk. The objective of this study was to evaluate the thrombogenic potential of FVL in asymptomatic carriers and in patients with personal or familial history of thrombosis. This was a retrospective single center study including 160 patients. Among them, 43 had personal history of thrombosis and 117 had familial history of thrombosis. Thrombin generation (TG) was realized in frozen platelet poor plasma with 1 pM of tissue factor and 4 µM of phospholipid. FVL mutation was associated with a global increase of TG. No difference was observed between patients with provoked thrombosis and patients with first-degree familial history of thrombosis (endogenous thrombin potential (ETP): 1501.0 ± 316.4 nM min and thrombin peak: 253.4 ± 71.5 nM vs. 1520.4 ± 283.8 nM min and 268.6 ± 68.0 nM). An increase of TG was observed in patients with unprovoked thrombosis (n = 23) and in patients with provoked thrombosis (n = 20) (ETP: 1819.5 ± 319.8 nM min and peak: 332.3 ± 55.8 nM). In the unprovoked thrombosis group, patients with a pulmonary embolism had a higher ETP than patients with deep vein thrombosis (DVT) (2036 ± 343 nM min vs. 1707 ± 261 nM min). With a predictive score formula (s = 0.1315 × Age + 0.0105 × ETP) with a threshold of 22.1 as risk to develop an unprovoked thrombosis among patients with second-degree familial history. The results of our analysis suggest that measurement of thrombin generation in patients with FVL mutation may identify subjects with an increased risk of unprovoked thrombosis. Further studies are needed to examine the usefulness of predicting thrombotic presentation in asymptomatic carriers.
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http://dx.doi.org/10.1007/s11239-019-01821-0DOI Listing
April 2019

The right blood collection tube for therapeutic drug monitoring and toxicology screening procedures: Standard tubes, gel or mechanical separator?

Clin Chim Acta 2019 Jan 12;488:196-201. Epub 2018 Nov 12.

Rouen University Hospital, Department of Pharmacology - Toxicology and Pharmacogenetics, Rouen, France; Normandie University, UNIROUEN, Inserm U1096, Rouen, France. Electronic address:

Stability data of toxics or drugs in gel-based or mechanical separation blood collection tubes are lacking, especially for therapeutic drug monitoring and clinical toxicology procedures. According to ISO 15189 accreditation standard, laboratories need to master the entire preanalytical process including the stability of analytes in a specific tube. Here we explored the impact of BD PST™ II and Barricor™ separator tubes on the stability of 167 therapeutic compounds and common drugs of abuse in plasma samples using LC-MS/MS. Forty drugs were significantly affected by the use of PST™ II tubes, including antidepressants (11/26), neuroleptics (9/13), cardiovascular drugs (5/26), anxiolytics and hypnotics (4/25) and some drugs of abuse (5/26). Six compounds exhibited significant reduction by the mechanical Barricor™ tubes. Ten drugs exhibited low (<85%) but non-significant recoveries due to inter-assay variability. Besides, a logP > 3.3 was determined as a cut-off value to predict a potential lack of stability in PST™ II gel tubes with an 86.4% sensitivity and a 61.4% specificity. As a consequence, determination of drugs with a logP > 3.3 should be carried out with caution in plasma samples withdrawn on PST™ II. The study showed the Barricor™ and non-gel tubes cause less drug interference and are recommended for the drugs studied.
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http://dx.doi.org/10.1016/j.cca.2018.10.043DOI Listing
January 2019

Alteration in the availability of epoxyeicosatrienoic acids contributes with NO to the development of endothelial dysfunction in conduit arteries during aging.

Atherosclerosis 2018 08 19;275:239-245. Epub 2018 Jun 19.

Department of Pharmacology, CHU de Rouen, 76000, Rouen, France; INSERM U1096, Normandie University, UNIROUEN, 76000, Rouen, France; Centre d'Investigation Clinique (CIC)-INSERM 1404, CHU de Rouen, 76000, Rouen, France. Electronic address:

Background And Aims: The mechanisms involved in endothelial dysfunction in humans during aging are largely unknown at the level of conduit arteries. We aimed to asses the role of NO and CYP450 epoxygenases-derived epoxyeicosatrienoic acids (EETs) in the regulation of endothelium-dependent flow-mediated dilatation of conduit arteries during aging.

Methods: Radial artery diameter and mean wall shear stress were determined by echotracking coupled with Doppler in 83 subjects (19-71 years old) during a sustained flow increase induced by hand skin heating, with the brachial infusion of saline or NO-synthase and cytochrome P450 epoxygenase inhibitors (L-NNMA and fluconazole respectively). Local blood sampling was performed for the quantification of NO metabolite nitrite and EETs.

Results: The magnitude of flow-mediated dilatation was independently and negatively correlated with age, baseline artery diameter and systolic blood pressure, and positively correlated with the increase in shear stress induced by heating. There was an increase in nitrite level during heating until the age of 35-40 years, which declined thereafter. However, the inhibitory effect of L-NMMA on flow-mediated dilatation progressively decreased during aging, demonstrating a decrease in functional NO availability. Moreover, aging progressively reduced the increase in EET level during heating as well as the inhibitory effect of fluconazole on flow-mediated dilatation.

Conclusions: These results show that aging impairs the availability of EETs and NO and epoxyeicosatrienoic acids in peripheral conduit arteries, contributing to the development of endothelial dysfunction.
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http://dx.doi.org/10.1016/j.atherosclerosis.2018.06.865DOI Listing
August 2018

Possible role of CYP2B6 genetic polymorphisms in ifosfamide-induced encephalopathy: report of three cases.

Fundam Clin Pharmacol 2018 Jun 25;32(3):337-342. Epub 2018 Jan 25.

Department of Pharmacology, Rouen University Hospital, 1 rue de Germont, F 76000, Rouen, France.

Ifosfamide (IFA) is a potent alkylating antitumoral agent, but its use is limited by neurological side effects. IFA is a racemic mixture of two enantiomeric forms, R-IFA and S-IFA with a stereoselective metabolism by CYP3A4 and CYP2B6, leading either to bioactive or to toxic pathways. In three consecutive cases of pediatric patients who exhibited IFA-induced encephalopathy (IIE), genotyping of clinically relevant single-nucleotide polymorphisms associated with decreased CYP3A4 and CYP2B6 activities was performed. Genetic investigations revealed the presence of CYP2B6 rs4803419 (C>T) in one patient while the two others carried the CYP2B6*6 allelic variant. All patients carried CYP3A4 wild-type genotype (CYP3A4*1/*1). Because CYP2B6-deficient alleles may be responsible for an increased conversion of S-IFA into neurotoxic metabolites, screening for CYP2B6 polymorphisms may help to avoid IIE and improve clinical outcomes.
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http://dx.doi.org/10.1111/fcp.12345DOI Listing
June 2018

Impact of CYP3A4 Genotype on Voriconazole Exposure.

Clin Pharmacol Ther 2018 02 29;103(2):185-186. Epub 2017 Aug 29.

Rouen University Hospital, Department of Pharmacology - Toxicology and Pharmacogenetics, Rouen, France.

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http://dx.doi.org/10.1002/cpt.809DOI Listing
February 2018

Multiple and Opposite Effects of Angiotensin II Receptor Blockers on the Bioavailability of Epoxyeicosatrienoic Acids.

Basic Clin Pharmacol Toxicol 2017 10 22;121(4):213-214. Epub 2017 Jun 22.

Department of Pharmacology & INSERM U1096, Rouen University Hospital, Normandie University, Rouen, France.

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http://dx.doi.org/10.1111/bcpt.12810DOI Listing
October 2017

Physiological role of endothelin-1 in flow-mediated vasodilatation in humans and impact of cardiovascular risk factors.

J Hypertens 2017 06;35(6):1204-1212

aDepartment of Pharmacology, Rouen University HospitalbInstitut National de la Santé et de la Recherche Médicale (INSERM) U1096cInstitute for Research and Innovation in Biomedicine, Normandy University, University of RouendCentre d'Investigation Clinique (CIC)-INSERM 1404, Rouen University HospitaleEquipe d'Accueil (EA) 4651, Rouen, France.

Objectives: The current study addressed the hypothesis that the local decrease in endothelin-1 (ET-1) bioavailability during sustained flow increases contributes to endothelium-dependent, flow-mediated dilatation (FMD) of conduit arteries and is altered in presence of cardiovascular risk factors.

Methods And Results: In nine young healthy individuals, the decrease in local ET-1 plasma levels and radial artery FMD in response to hand skin heating (from 34 to 44 °C) was not affected by endothelin type A (ETA) receptor blockade, achieved using the brachial infusion of BQ-123 (100 nmol/min per l of forearm), as compared with physiological saline (0.9% NaCl) infusion. In contrast, endothelin type B (ETB) receptor blockade with BQ-788 (10 nmol/min per l) suppressed the decrease in plasma ET-1 during heating and reduced FMD, without altering nitric oxide release. The coinfusion of BQ-123 did not affect the inhibitory effect of ETB receptor blockade on the decrease in ET-1 plasma levels during heating but prevented the reduction in FMD. Basal radial artery parameters, systemic hemodynamics, and endothelium-independent dilatation to glyceryl trinitrate were not modified by ETA and/or ETB blockade. In a general population of 40 participants without treatment or major cardiovascular diseases, including the nine healthy individuals, the reduction in endothelin-1 level during heating was correlated with FMD (r = -0.55, P < 0.001) and decreased with increased age (r = 0.49, P = 0.001), mean arterial blood pressure (r = 0.48, P = 0.002), and total cholesterol level (r = 0.37, P = 0.024).

Conclusion: The uptake of endothelin-1 by ETB receptors contributes to conduit artery FMD, preventing its vasoconstrictor action mediated by ETA receptors. The alteration of this mechanism by cardiovascular risk factors may contribute to endothelial dysfunction.
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http://dx.doi.org/10.1097/HJH.0000000000001307DOI Listing
June 2017

Pharmacogenetics in cardiovascular diseases: State of the art and implementation-recommendations of the French National Network of Pharmacogenetics (RNPGx).

Therapie 2017 Apr 30;72(2):257-267. Epub 2017 Jan 30.

Service de pharmacologie, laboratoire de pharmacologie-toxicologie et pharmacogénétique, centre hospitalier universitaire, 76031 Rouen, France; Inserm unité 1096, pharmacologie des dysfonctionnements endothéliaux et myocardiques, faculté de médecine et de pharmacie, université de Rouen, 76000 Rouen, France.

The use of genomic markers to predict drug response and effectiveness has the potential to improve healthcare by increasing drug efficacy and minimizing adverse effects. Polymorphisms associated with inter-individual variability in drug metabolism, transport, or pharmacodynamics of major cardiovascular drugs have been identified. These include single nucleotide polymorphisms (SNP) affecting clinical outcomes in patients receiving antiplatelet agents, oral anticoagulants and statins. Based on clinical evidence supporting genetic testing in the management of cardiovascular diseases using these drug classes, this short review presents clinical guidance regarding current pharmacogenetics implementation in routine medical practice.
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http://dx.doi.org/10.1016/j.therap.2016.09.017DOI Listing
April 2017

Pharmacogénétique en cardiovasculaire : état des connaissances et des pratiques – recommandations du Réseau national de pharmacogénétique (RNPGx).

Therapie 2017 Apr 3;72(2):245-255. Epub 2017 Jan 3.

Service de pharmacologie, laboratoire de pharmacologie-toxicologie et pharmacogénétique, institut de biologie clinique (I.B.C.), 2e étage, hôpitaux de Rouen, centre hospitalier universitaire, 1, rue de Germont, 76031 Rouen, France; Inserm unité 1096, pharmacologie des dysfonctionnements endothéliaux et myocardiques, faculté de médecine et de pharmacie, université de Rouen, 76000 Rouen, France.

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http://dx.doi.org/10.1016/j.therap.2016.09.010DOI Listing
April 2017

A sensitive LC-MS/MS method for the quantification of regioisomers of epoxyeicosatrienoic and dihydroxyeicosatrienoic acids in human plasma during endothelial stimulation.

Anal Bioanal Chem 2017 Mar 15;409(7):1845-1855. Epub 2016 Dec 15.

Department of Pharmacology, Rouen University Hospital, 1 rue de Germont, 76031, Rouen, France.

Epoxyeicosatrienoic acids (EETs) are vasodilating lipid mediators metabolized into dihydroxyeicosatrienoic acids (DHETs) by soluble epoxide hydrolase. We aimed to develop a LC-MS/MS method to quantify EETs and DHETs in human plasma and monitored their levels during vascular endothelial stimulation. Plasma samples, collected from 14 healthy and five hypertensive subjects at baseline and during radial artery endothelium-dependent flow-mediated dilatation, were spiked with internal standards. Lipids were then extracted by a modified Bligh and Dyer method and saponified to release bound EETs and DHETs. Samples were purified by a second liquid-liquid extraction and analyzed by LC-MS/MS. The assay allowed identification of (±)8(9)-epoxy-5Z,11Z,14Z-eicosatrienoic acid (8,9-EET); (±)11(12)-epoxy-5Z,8Z,14Z-eicosatrienoic acid (11,12-EET); (±)14(15)-epoxy-5Z,8Z,11Z-eicosatrienoic acid (14,15-EET); (±)8,9-dihydroxy-5Z,11Z,14Z-eicosatrienoic acid (8,9-DHET); (±)11,12-dihydroxy-5Z,8Z,14Z-eicosatrienoic acid (11,12-DHET); and (±)14,15-dihydroxy-5Z,8Z,11Z-eicosatrienoic acid (14,15-DHET). (±)5(6)-epoxy-5Z,11Z,14Z-eicosatrienoic acid (5,6-EET) was virtually undetectable due to its chemical instability. The limits of quantification were 0.25 ng/mL for DHETs and 0.5 ng/mL for EETs. Intra- and inter-assay variations ranged from 1.6 to 13.2%. Heating induced a similar increase in 8,9-EET, 11,12-EET, and 14,15-EET levels and in corresponding DHET levels in healthy but not in hypertensive subjects. We validated a sensitive LC-MS/MS method for measuring simultaneously plasma EET and DHET regioisomers in human plasma and showed its interest for assessing endothelial function.
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http://dx.doi.org/10.1007/s00216-016-0129-1DOI Listing
March 2017

Impact of CYP2C19 genetic polymorphisms on voriconazole dosing and exposure in adult patients with invasive fungal infections.

Int J Antimicrob Agents 2016 Feb 21;47(2):124-31. Epub 2015 Dec 21.

Department of Pharmacology and Toxicology, University Hospital, Rouen, France; INSERM U1096, University of Rouen, School of Medicine, Rouen, France.

Voriconazole (VCZ) use is limited by its narrow therapeutic range and significant interpatient variability in exposure. This study aimed to assess (i) the impact of CYP2C19 genotype on VCZ exposure and (ii) the doses required to achieve the therapeutic range in adult patients with invasive fungal infections (IFIs). Therapeutic drug monitoring (TDM) of VCZ, based on trough concentration measurement, and CYP2C19 genotyping were used to guide VCZ dosing in Caucasian patients with IFIs. The two common polymorphisms in Caucasians (CYP2C19*2 and *17), associated with decreased or increased CYP2C19 activity, respectively, were correlated with the daily VCZ dose, pharmacokinetic parameters and concentration-to-dose ratio. In total, 111 trough concentration measurements from 35 genotyped patients were analysed using linear mixed-effect models. The mean VCZ doses required to achieve target concentrations were significantly higher in CYP2C19*17 carriers compared with CYP2C19*1/*1 individuals (P<0.001): 2.57±0.25mg/kg twice daily in CYP2C19*1/*1 patients versus 3.94±0.39mg/kg and 6.75±0.54mg/kg in *1/*17 and *17/*17 patients, respectively. In addition, exposure to VCZ correlated with the CYP2C19*17 variant. Indices of exposure for CYP2C19*2 carriers were in line with the functional effect of this polymorphism compared with CYP2C19*1/*1 individuals, however comparisons of doses required to achieve target concentrations were not statistically different. The CYP2C19*17 allele predicted both VCZ exposure and dose required to achieve effective and non-toxic concentrations. CYP2C19 genotyping appears useful to guide VCZ initial dosing when coupled with TDM and to explain subtherapeutic concentrations frequently observed in clinical practice.
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http://dx.doi.org/10.1016/j.ijantimicag.2015.12.003DOI Listing
February 2016

Impact of soluble epoxide hydrolase inhibition on early kidney damage in hyperglycemic overweight mice.

Prostaglandins Other Lipid Mediat 2015 Jul 27;120:148-54. Epub 2015 May 27.

Institut National de la Santé et de la Recherche Médicale (INSERM) U1096, Rouen, France; University of Rouen, Institute for Research and Innovation in Biomedicine (IRIB), Rouen, France; Department of Pharmacology, Rouen University Hospital, Rouen, France. Electronic address:

This study addressed the hypothesis that inhibition of the EETs degrading enzyme soluble epoxide hydrolase affords renal protection in the early stage of diabetic nephropathy. The renal effects of the sEH inhibitor t-AUCB (10mg/l in drinking water) were compared to those of the sulfonylurea glibenclamide (80mg/l), both administered for 8 weeks in FVB mice subjected to a high-fat diet (HFD, 60% fat) for 16 weeks. Mice on control chow diet (10% fat) and non-treated HFD mice served as controls. Compared with non-treated HFD mice, HFD mice treated with t-AUCB had a decreased EET degradation, as shown by their higher plasma EETs-to-DHETs ratio, and an increased EET production, as shown by the increase in EETs+DHETs levels, which was associated with induction of CYP450 epoxygenase expression. Both agents similarly reduced fasting glycemia but only t-AUCB prevented the increase in the urinary albumine-to-creatinine ratio in HFD mice. Histopathological analysis showed that t-AUCB reduced renal inflammation, which was associated with an increased mRNA expression of the NFκB inhibitor Iκ≡ and related decrease in MCP-1, COX2 and VCAM-1 expressions. Finally, there was a marginally significant increase in reactive oxygen species production in HFD mice, together with an enhanced NOX2 expression. Both agents did not modify these parameters but t-AUCB increased the expression of the antioxidant enzyme superoxide dismutase 1. These results demonstrate that, independently from its glucose-lowering effect, sEH inhibition prevents microalbuminuria and renal inflammation in overweight hyperglycemic mice, suggesting that this pharmacological strategy could be useful in the management of diabetic nephropathy.
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http://dx.doi.org/10.1016/j.prostaglandins.2015.04.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4575616PMC
July 2015

Design and discovery of soluble epoxide hydrolase inhibitors for the treatment of cardiovascular diseases.

Expert Opin Drug Discov 2014 Mar 4;9(3):229-43. Epub 2014 Feb 4.

Rouen University Hospital, Department of Pharmacology , Rouen , France.

Introduction: Cardiovascular diseases are a leading cause of death in developed countries. Increasing evidence shows that the alteration in the normal functions of the vascular endothelium plays a major role in the development of cardiovascular diseases. However, specific agents designed to prevent endothelial dysfunction and related cardiovascular complications are still lacking. One emerging strategy is to increase the bioavailability of epoxyeicosatrienoic acids (EETs), synthesized by cytochrome P450 epoxygenases from arachidonic acid. EETs are endothelium-derived hyperpolarising and relaxing factors and display attractive anti-inflammatory and metabolic properties. Genetic polymorphism studies in humans, and experiments in animal models of diseases, have identified soluble epoxide hydrolase (sEH), the major enzyme involved in EET degradation, as a potential pharmacological target.

Areas Covered: This review presents EET pathway and its functions and summarises the data supporting the development of sEH inhibitors for the treatment of cardiovascular and metabolic diseases. Furthermore, the authors present the different chemical families of sEH inhibitors developed and their effects in animal models of cardiovascular and metabolic diseases.

Expert Opinion: Several generations of sEH inhibitors have now been designed to treat endothelial dysfunction and cardiovascular complications for a variety of diseases. The safety of these drugs remains to be carefully investigated, particularly in relation to carcinogenesis. The increasing knowledge of the biological role of each of the EET isomers and of their metabolites may improve their pharmacological profile. This, in turn, could potentially lead to the identification of new pharmacological agents that achieve the cellular effects needed without the deleterious side effects.
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http://dx.doi.org/10.1517/17460441.2014.881354DOI Listing
March 2014