Publications by authors named "Mercedes Quintana"

18 Publications

  • Page 1 of 1

Efficacy of Targeted Radionuclide Therapy Using [I]ICF01012 in 3D Pigmented BRAF- and NRAS-Mutant Melanoma Models and In Vivo NRAS-Mutant Melanoma.

Cancers (Basel) 2021 Mar 20;13(6). Epub 2021 Mar 20.

INSERM U1240, University of Clermont Auvergne, 58 rue Montalembert, 63000 Clermont-Ferrand, France.

Purpose: To assess the efficiency of targeted radionuclide therapy (TRT), alone or in combination with MEK inhibitors (MEKi), in melanomas harboring constitutive MAPK/ERK activation responsible for tumor radioresistance.

Methods: For TRT, we used a melanin radiotracer ([I]ICF01012) currently in phase 1 clinical trial (NCT03784625). TRT alone or combined with MEKi was evaluated in three-dimensional melanoma spheroid models of human BRAF SK-MEL-3, murine NRAS 1007, and WT B16F10 melanomas. TRT in vivo biodistribution, dosimetry, efficiency, and molecular mechanisms were studied using the C57BL/6J-NRAS 1007 syngeneic model.

Results: TRT cooperated with MEKi to increase apoptosis in both BRAF- and NRAS-mutant spheroids. NRAS spheroids were highly radiosensitive towards [I]ICF01012-TRT. In mice bearing NRAS 1007 melanoma, [I]ICF01012 induced a significant extended survival (92 vs. 44 days, < 0.0001), associated with a 93-Gy tumor deposit, and reduced lymph-node metastases. Comparative transcriptomic analyses confirmed a decrease in mitosis, proliferation, and metastasis signatures in TRT-treated vs. control tumors and suggest that TRT acts through an increase in oxidation and inflammation and P53 activation.

Conclusion: Our data suggest that [I]ICF01012-TRT and MEKi combination could be of benefit for advanced pigmented BRAF-mutant melanoma care and that [I]ICF01012 alone could constitute a new potential NRAS-mutant melanoma treatment.
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http://dx.doi.org/10.3390/cancers13061421DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8003594PMC
March 2021

Benzamide derivative radiotracers targeting melanin for melanoma imaging and therapy: Preclinical/clinical development and combination with other treatments.

Pharmacol Ther 2021 Mar 1;224:107829. Epub 2021 Mar 1.

Université Clermont Auvergne, INSERM, Imagerie Moléculaire et Stratégies Théranostiques, UMR1240, 58 Rue Montalembert, 63005 Clermont-Ferrand, Cedex, France. Electronic address:

Cutaneous melanoma arises from proliferating melanocytes, cells specialized in the production of melanin. This property means melanin can be considered as a target for monitoring melanoma patients using nuclear imaging or targeted radionuclide therapy (TRT). Since the 1970s, many researchers have shown that specific molecules can interfere with melanin. This paper reviews some such molecules: benzamide structures improved to increase their pharmacokinetics for imaging or TRT. We first describe the characteristics and biosynthesis of melanin, and the main features of melanin tracers. The second part summarizes the preclinical and corresponding clinical studies on imaging. The last section presents TRT results from ongoing protocols and discusses combinations with other therapies as an opportunity for melanoma non-responders or patients resistant to treatments.
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http://dx.doi.org/10.1016/j.pharmthera.2021.107829DOI Listing
March 2021

Optimization of IEDDA bioorthogonal system: Efficient process to improve trans-cyclooctene/tetrazine interaction.

Eur J Med Chem 2020 Oct 15;203:112574. Epub 2020 Jul 15.

Université Clermont Auvergne, Imagerie Moléculaire et Stratégies Théranostiques, BP 184, F-63005, Clermont-Ferrand, France; Inserm, U 1240, F-63000, Clermont-Ferrand, France; Centre Jean Perrin, F-63011, Clermont-Ferrand, France. Electronic address:

The antibody pretargeting approach for radioimmunotherapy (RIT) using inverse electron demand Diels-Alder cycloaddition (IEDDA) constitutes an emerging theranostic approach for solid cancers. However, IEDDA pretargeting has not reached clinical trial. The major limitation of the IEDDA strategy depends largely on trans-cyclooctene (TCO) stability. Indeed, TCO may isomerize into the more stable but unreactive cis-cyclooctene (CCO), leading to a drastic decrease of IEDDA efficiency. We have thus developed both efficient and reproducible synthetic pathways and analytical follow up for (PEGylated) TCO derivatives, providing high TCO isomeric purity for antibody modification. We have set up an original process to limit the isomerization of TCO to CCO before the mAbs' functionalization to allow high TCO/tetrazine cycloaddition.
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http://dx.doi.org/10.1016/j.ejmech.2020.112574DOI Listing
October 2020

Pretargeted radioimmunotherapy and SPECT imaging of peritoneal carcinomatosis using bioorthogonal click chemistry: probe selection and first proof-of-concept.

Theranostics 2019 19;9(22):6706-6718. Epub 2019 Sep 19.

Université Clermont Auvergne, Imagerie Moléculaire et Stratégies Théranostiques, BP 184, F-63005 Clermont-Ferrand, France. Inserm, U 1240, F-63000 Clermont-Ferrand, France. Centre Jean Perrin, F-63011 Clermont-Ferrand, France.

: Pretargeted radioimmunotherapy (PRIT) based upon bioorthogonal click chemistry has been investigated for the first time in the context of peritoneal carcinomatosis using a CEA-targeting 35A7 mAb bearing -cyclooctene (TCO) moieties and several Lu-labeled tetrazine (Tz) radioligands. Starting from three Tz probes containing PEG linkers of varying lengths between the DOTA and Tz groups (. PEG = 3, 7, or 11, respectively, for Tz-, Tz-, and Tz-), we selected [Lu]Lu-Tz as the most appropriate for pretargeted SPECT imaging and demonstrated its efficacy in tumor growth control. An orthotopic model of peritoneal carcinomatosis (PC) was obtained following the intraperitoneal (i.p.) injection of A431-CEA-Luc cells in nude mice. Tumor growth was assessed using bioluminescence imaging. Anti-CEA 35A7 mAb was grafted with 2-3 TCO immunoglobulin. Pretargeted SPECT imaging and biodistribution experiments were performed to quantify the activity concentrations of [Lu]Lu-Tz- in tumors and non-target organs to determine the optimal Tz probe for the PRIT of PC. The pharmacokinetic profiles of [Lu]Lu-Tz- alone were determined using both SPECT imaging and biodistribution experiments. These data revealed that [Lu]Lu-Tz- was cleared both the renal and hepatic systems while [Lu]Lu-Tz- and [Lu]Lu-Tz- were predominantly excreted the renal system. In addition, these results illuminated that the longer the PEG linker, the more rapidly the Tz radioligand was cleared from the peritoneal cavity. The absorbed radiation dose corresponding to pretargeting with 35A7-TCO followed 24 h later by [Lu]Lu-Tz- was higher for tumors following the administration of [Lu]Lu-Tz- (. 0.59 Gy/MBq) compared to either [Lu]Lu-Tz- (. 0.25 Gy/MBq) and [Lu]Lu-Tz- (. 0.18 Gy/MBq). In a longitudinal PRIT study, we showed that the i.p. injection of 40 MBq of [Lu]Lu-Tz- 24 hours after the systemic administration of 35A7-TCO significantly slowed tumor growth compared to control mice receiving only saline or 40 MBq of [Lu]Lu-Tz- alone. measurement of the peritoneal carcinomatosis index (PCI) confirmed that PRIT significantly reduced tumor growth (PCI = 15.5 ± 2.3 after PRIT 30.0 ± 2.3 and 30.8 ± 1.4 for the NaCl and [Lu]Lu-Tz- alone groups, respectively). : Our results clearly demonstrate the impact of the length of PEG linkers upon the biodistribution profiles of Lu-labeled Tz radioligands. Furthermore, we demonstrated for the first time the possibility of using bioorthogonal chemistry for both the pretargeted SPECT and PRIT of peritoneal carcinomatosis.
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http://dx.doi.org/10.7150/thno.35461DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6771248PMC
October 2020

Targeted Radionuclide Therapy Decreases Melanoma Lung Invasion by Modifying Epithelial-Mesenchymal Transition-Like Mechanisms.

Transl Oncol 2019 Nov 14;12(11):1442-1452. Epub 2019 Aug 14.

UMR 1240 INSERM, University of Clermont Auvergne, Clermont-Ferrand, France. Electronic address:

Melanin-radiolabeled molecules for targeted radionuclide therapy (TRT) provide a promising approach for the treatment of pigmented melanoma. Among these radiolabeled molecules, the iodinated melanin-specific binding molecule ([I]ICF01012) has shown a significant antitumor effect on metastatic melanoma preclinical models. We report herein that [I]ICF01012 decreases the epithelial-mesenshymal transition-like (EMT-like) markers in both in vivo and in vitro three-dimensional (3D) melanoma spheroid models. [I]ICF01012 spheroids irradiation resulted in reduced clonogenic capacity of all pigmented spheroids accompanied by increased protein expression levels of phosphorylated H2A.X, p53 and its downstream target p21. In addition, [I]ICF01012 treatment leads to a significant increase of cell pigmentation as demonstrated in SK-MEL3 human xenograft model. We also showed that [I]ICF01012 decreases the size and the number of melanoma lung colonies in the syngeneic murine B16BL6 in vivo model assessing its potentiality to kill circulating tumor cells. Taken together, these results indicate that [I]ICF01012 reduces metastatic capacity of melanoma cells presumably through EMT-like reduction and cell differentiation induction.
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http://dx.doi.org/10.1016/j.tranon.2019.07.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6704444PMC
November 2019

Determination of eumelanin and pheomelanin in melanomas using solid-phase extraction and high performance liquid chromatography-diode array detection (HPLC-DAD) analysis.

J Chromatogr B Analyt Technol Biomed Life Sci 2019 Apr 11;1113:60-68. Epub 2019 Mar 11.

UMR 1240 INSERM UCA, 58 Rue Montalembert, 63005 Clermont-Ferrand Cedex, France. Electronic address:

Determination of eumelanin and pheomelanin in melanomas that exhibit different pigmentation was carried using a solid-phase extraction (SPE) preparation method based on weak anion exchange chemistry. This extraction significantly enhanced the chromatographic profile obtained by reverse phase high performance liquid chromatography-diode array detection (RP-HPLC-DAD). The SPE method was developed using aqueous standards of melanin markers: thiazole-2,4,5-tricarboxylic acid (TTCA), thiazole-4,5-dicarboxylic acid (TDCA), pyrrole-2,3-dicarboxylic acid (PDCA) and pyrrole-2,3,5-tricarboxylic acid (PTCA) and non-pigmented cell lines spiked with those markers. An excellent average recovery, above 90%, was obtained for the four markers with a relative standard deviation below 7%. We have also optimized the stationary phase and the mobile phase (phosphate concentration and pH) to improve sensitivity and to reduce the analysis time. Elution of the four markers is achieved in 5 min and total analysis of biological samples is completed in 15 min. The quantification limits for TDCA, TTCA, PDCA and PTCA are 60, 50, 47 and 48 ng/mL respectively. Furthermore, DAD detection improves the marker identification in complex matrices through the analysis of UV spectra. We have successfully applied this method to melanoma tumors and cells. Murine B16BL6 tumor are highly pigmented with mostly eumelanin (98.1% of eumelanin) while human SK-MEL-3 tumor contain about 30% pheomelanin. B16BL6 and B16F10 are eumelanic cells lines and NHEM melanocytes contain about 24% of pheomelanin.
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http://dx.doi.org/10.1016/j.jchromb.2019.03.010DOI Listing
April 2019

Antibody PEGylation in bioorthogonal pretargeting with trans-cyclooctene/tetrazine cycloaddition: in vitro and in vivo evaluation in colorectal cancer models.

Sci Rep 2017 11 2;7(1):14918. Epub 2017 Nov 2.

Université Clermont Auvergne, INSERM U1240, Imagerie Moléculaire et Stratégies Théranostiques, F-63000, Clermont Ferrand, France.

Bioorthogonal chemistry represents a challenging approach in pretargeted radioimmunotherapy (PRIT). We focus here on mAb modifications by grafting an increase amount of trans-cyclooctene (TCO) derivatives (0 to 30 equivalents with respect to mAb) bearing different polyethylene glycol (PEG) linkers between mAb and TCO (i.e. PEG (1), PEG (2) and PEG (3)) and assessing their functionality. We used colorectal xenograft (HT29/Ts29.2) and peritoneal carcinomatosis (A431-CEA-Luc/35A7) as tumor cells/mAbs models and fluorescent tetrazines (TZ). MALDI-TOF MS shows that grafting with 2,3 increases significantly the number of TCO per mAb compared with no PEG. In vitro immunofluorescence showed that Ts29.2 and 35A7 labeling intensity is correlated with the number of TCO when using 1,3 while signals reach a maximum at 10 equivalents when using 2. Under 10 equivalents conditions, the capacity of resulting mAbs-1-3 for antigen recognition is similar when reported per grafted TCO and comparable to mAbs without TCO. In vivo, on both models, pretargeting with mAbs-2,3 followed by TZ injection induced a fluorescent signal two times lower than with mAbs-1. These findings suggest that while PEG linkers allow a better accessibility for TCO grafting, it might decrease the number of reactive TCO. In conclusion, mAb-1 represents the best candidate for PRIT.
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http://dx.doi.org/10.1038/s41598-017-15051-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5668303PMC
November 2017

Tetraspanin 8 (TSPAN 8) as a potential target for radio-immunotherapy of colorectal cancer.

Oncotarget 2017 Mar;8(13):22034-22047

INSERM, U 1240, 63005 Clermont-Ferrand, France.

Tetraspanin 8 (TSPAN8) overexpression is correlated with poor prognosis in human colorectal cancer (CRC). A murine mAb Ts29.2 specific for human TSPAN8 provided significant efficiency for immunotherapy in CRC pre-clinical models. We therefore evaluate the feasability of targeting TSPAN8 in CRC with radiolabeled Ts29.2. Staining of tissue micro-arrays with Ts29.2 revealed that TSPAN8 espression was restricted to a few human healthy tissues. DOTA-Ts29.2 was radiolabeled with 111In or 177Lu with radiochemical purities >95%, specific activity ranging from 300 to 600 MBq/mg, and radioimmunoreactive fractions >80%. The biodistribution of [111In]DOTA-Ts29.2 in nude mice bearing HT29 or SW480 CRC xenografts showed a high specificity of tumor localization with high tumor/blood ratios (HT29: 4.3; SW480-TSPAN8: 3.9 at 72h and 120h post injection respectively). Tumor-specific absorbed dose calculations for [177Lu]DOTA-Ts29.2 was 1.89 Gy/MBq, establishing the feasibility of using radioimmunotherapy of CRC with this radiolabeled antibody. A significant inhibition of tumor growth in HT29 tumor-bearing mice treated with [177Lu]DOTA-Ts29.2 was observed compared to control groups. Ex vivo experiments revealed specific DNA double strand breaks associated with cell apoptosis in [177Lu]DOTA-Ts29.2 treated tumors compared to controls. Overall, we provide a proof-of-concept for the use of [111In/177Lu]DOTA-Ts29.2 that specifically target in vivo aggressive TSPAN8-positive cells in CRC.
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http://dx.doi.org/10.18632/oncotarget.15787DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5400644PMC
March 2017

Proteoglycans as Target for an Innovative Therapeutic Approach in Chondrosarcoma: Preclinical Proof of Concept.

Mol Cancer Ther 2016 11 29;15(11):2575-2585. Epub 2016 Aug 29.

Clermont Université, Université d'Auvergne, Imagerie Moléculaire et Thérapie Vectorisée, Clermont-Ferrand, France.

To date, surgery remains the only option for the treatment of chondrosarcoma, which is radio- and chemoresistant due in part to its large extracellular matrix (ECM) and poor vascularity. In case of unresectable locally advanced or metastatic diseases with a poor prognosis, improving the management of chondrosarcoma still remains a challenge. Our team developed an attractive approach of improvement of the therapeutic index of chemotherapy by targeting proteoglycan (PG)-rich tissues using a quaternary ammonium (QA) function conjugated to melphalan (Mel). First of all, we demonstrated the crucial role of the QA carrier for binding to aggrecan by surface plasmon resonance. In the orthotopic model of Swarm rat chondrosarcoma, an in vivo biodistribution study of Mel and its QA derivative (Mel-QA), radiolabeled with tritium, showed rapid radioactivity accumulation in healthy cartilaginous tissues and tumor after [H]-Mel-QA injection. The higher T/M ratio of the QA derivative suggests some advantage of QA-active targeting of chondrosarcoma. The antitumoral effects were characterized by tumor volume assessment, in vivo Tc-NTP 15-5 scintigraphic imaging of PGs, H-HRMAS NMR spectroscopy, and histology. The conjugation of a QA function to Mel did not hamper its in vivo efficiency and strongly improved the tolerability of Mel leading to a significant decrease of side effects (hematologic analyses and body weight monitoring). Thus, QA conjugation leads to a significant improvement of the therapeutic index, which is essential in oncology and enable repeated cycles of chemotherapy in patients with chondrosarcoma. Mol Cancer Ther; 15(11); 2575-85. ©2016 AACR.
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http://dx.doi.org/10.1158/1535-7163.MCT-16-0003DOI Listing
November 2016

Compatibility of Injectable Anticoagulant Agents in Ethanol; In Vitro Antibiofilm Activity and Impact on Polyurethane Catheters of Enoxaparin 400 U/mL in 40% v/v Ethanol.

PLoS One 2016 21;11(7):e0159475. Epub 2016 Jul 21.

UMR CNRS 6023, Laboratoire Microorganismes: Génome et Environnement, Clermont Université, Université d'Auvergne, Clermont Ferrand, 63000, France.

Background And Objectives: Interdialytic lock solutions should maintain catheter patency and prevent catheter infections. We aimed to determine in which conditions injectable anticoagulant agents (IAAs) combined with ethanol are compatible and to assess the antibiofilm activity of the selected combination and its effects on dialysis catheters (DC).

Methods: The solubility and compatibility of unfractionated heparin (UFH), low molecular weight heparins (LMWHs), heparinoids and fondaparinux (50 to 2,500 U/mL) in 30 to 70% ethanol were determined by visual observation. The stability of enoxaparin in ethanol and the ethanol content were assessed by high performance liquid chromatography (HPLC) and titrimetric control, respectively. The bactericidal effect was determined on 24h-old biofilms embedded in silicone-DC. The integrity of polyurethane-DC immersed in anticoagulant-ethanol was assessed by gas chromatography-mass spectrometry (GC-MS) and compared with previously published results.

Results: The compatibility of IAAs and ethanol varied according to IAA type and concentration, and ethanol content. UFH in 40% ethanol was not compatible, whatever the UFH concentration used. Established limits of compatibility of enoxaparin, nadroparin, dalteparin and tinzaparin in 40% ethanol were 1350, 575, 307 and 207 U/ml, respectively, and up to 300 U/ml for danaparoid and 1 mg/mL for fondaparinux. Enoxaparin 400 U/mL in 40% ethanol (Enox/Eth) eradicated biofilm after 4 hours of exposure for Staphylococcus epidermidis, Pseudomonas aeruginosa and Candida albicans and after 24 hours for Klebsiella pneumoniae and S. aureus. Aliphatic carbonate and alcohol compounds were released by polyurethane-DC after Enox/Eth exposure, as after 40% ethanol or saline exposure. There was no significant difference between the amounts released after 30 minutes of exposure to Enox/Eth and 15 days to saline.

Conclusions: A 40% ethanol solution can be combined with all IAAs but UFH. Enox/Eth was effective as an anti-biofilm agent with minor impacts on DC integrity and could be a useful interdialytic lock solution.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0159475PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4956118PMC
July 2017

Fibronectin contributes to pathological cardiac hypertrophy but not physiological growth.

Basic Res Cardiol 2013 Sep 4;108(5):375. Epub 2013 Aug 4.

Heart Institute, and Biology Department, SDSU Integrated Regenerative Research Institute, Life Sciences North, Room 426, 5500 Campanile Drive, San Diego, CA 92182, USA.

Ability of the heart to undergo pathological or physiological hypertrophy upon increased wall stress is critical for long-term compensatory function in response to increased workload demand. While substantial information has been published on the nature of the fundamental molecular signaling involved in hypertrophy, the role of extracellular matrix protein Fibronectin (Fn) in hypertrophic signaling is unclear. The objective of the study was to delineate the role of Fn during pressure overload-induced pathological cardiac hypertrophy and physiological growth prompted by exercise. Genetic conditional ablation of Fn in adulthood blunts cardiomyocyte hypertrophy upon pressure overload via attenuated activation of nuclear factor of activated T cells (NFAT). Loss of Fn delays development of heart failure and improves survival. In contrast, genetic deletion of Fn has no impact on physiological cardiac growth induced by voluntary wheel running. Down-regulation of the transcription factor c/EBPβ (Ccaat-enhanced binding protein β), which is essential for induction of the physiological growth program, is unaffected by Fn deletion. Nuclear NFAT translocation is triggered by Fn in conjunction with up-regulation of the fetal gene program and hypertrophy of cardiomyocytes in vitro. Furthermore, activation of the physiological gene program induced by insulin stimulation in vitro is attenuated by Fn, whereas insulin had no impact on Fn-induced pathological growth program. Fn contributes to pathological cardiomyocyte hypertrophy in vitro and in vivo via NFAT activation. Fn is dispensable for physiological growth in vivo, and Fn attenuates the activation of the physiological growth program in vitro.
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http://dx.doi.org/10.1007/s00395-013-0375-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3813434PMC
September 2013

Fibronectin is essential for reparative cardiac progenitor cell response after myocardial infarction.

Circ Res 2013 Jul 7;113(2):115-25. Epub 2013 May 7.

San Diego State University Integrated Regenerative Research Institute, San Diego, CA 92182, USA.

Rationale: Adoptive transfer of cardiac progenitor cells (CPCs) has entered clinical application, despite limited mechanistic understanding of the endogenous response after myocardial infarction (MI). Extracellular matrix undergoes dramatic changes after MI and therefore might be linked to CPC-mediated repair.

Objective: To demonstrate the significance of fibronectin (Fn), a component of the extracellular matrix, for induction of the endogenous CPC response to MI.

Methods And Results: This report shows that presence of CPCs correlates with the expression of Fn during cardiac development and after MI. In vivo, genetic conditional ablation of Fn blunts CPC response measured 7 days after MI through reduced proliferation and diminished survival. Attenuated vasculogenesis and cardiogenesis during recovery were evident at the end of a 12-week follow-up period. Impaired CPC-dependent reparative remodeling ultimately leads to continuous decline of cardiac function in Fn knockout animals. In vitro, Fn protects and induces proliferation of CPCs via β₁-integrin-focal adhesion kinase-signal transducer and activator of transcription 3-Pim1 independent of Akt.

Conclusions: Fn is essential for endogenous CPC expansion and repair required for stabilization of cardiac function after MI.
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http://dx.doi.org/10.1161/CIRCRESAHA.113.301152DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3815660PMC
July 2013

Mass spectrometry-based metabolomics for the discovery of biomarkers of fruit and vegetable intake: citrus fruit as a case study.

J Proteome Res 2013 Apr 5;12(4):1645-59. Epub 2013 Mar 5.

INRA , UMR 1019, UNH, CRNH Auvergne, F-63000 Clermont-Ferrand, France.

Elucidation of the relationships between genotype, diet, and health requires accurate dietary assessment. In intervention and epidemiological studies, dietary assessment usually relies on questionnaires, which are susceptible to recall bias. An alternative approach is to quantify biomarkers of intake in biofluids, but few such markers have been validated so far. Here we describe the use of metabolomics for the discovery of nutritional biomarkers, using citrus fruits as a case study. Three study designs were compared. Urinary metabolomes were profiled for volunteers that had (a) consumed an acute dose of orange or grapefruit juice, (b) consumed orange juice regularly for one month, and (c) reported high or low consumption of citrus products for a large cohort study. Some signals were found to reflect citrus consumption in all three studies. Proline betaine and flavanone glucuronides were identified as known biomarkers, but various other biomarkers were revealed. Further, many signals that increased after citrus intake in the acute study were not sensitive enough to discriminate high and low citrus consumers in the cohort study. We propose that urine profiling of cohort subjects stratified by consumption is an effective strategy for discovery of sensitive biomarkers of consumption for a wide range of foods.
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http://dx.doi.org/10.1021/pr300997cDOI Listing
April 2013

Fibrinolytic activity is associated with presence of cystic medial degeneration in aneurysms of the ascending aorta.

Histopathology 2010 Dec;57(6):917-32

Inserm Unit 698, Paris, University of Paris 7 (Denis Diderot), Paris, France.

Aims: Thoracic ascending aortic aneurysms (TAA) are characterized by elastic fibre breakdown and cystic medial degeneration within the aortic media, associated with progressive smooth muscle cell (SMC) rarefaction. The transforming growth factor (TGF)-β/Smad2 signalling pathway is involved in this process. Because the pericellular fibrinolytic system activation is able to degrade adhesive proteins, activate matrix metalloproteinase (MMP), induce SMC disappearance and increase the bioavailability of TGF-β, the aim was to investigate the plasminergic system in TAA.

Methods And Results: Ascending aortas [21 controls and 19 TAAs (of three different aetiologies)] were analysed. Immunohistochemistry showed accumulation of t-PA, u-PA and plasmin in TAAs, associated with residual SMCs. Overexpression of t-PA and u-PA was confirmed by reverse transcription-polymerase chain reaction (RT-PCR), immunoblotting and zymography on TAA extracts and culture medium conditioned by TAA. Plasminogen was present on the SMC surface and inside cytoplasmic vesicles, but plasminogen mRNA was undetectable in the TAA medial layer. Plasmin-antiplasmin complexes were detected in TAA-conditioned medium and activation of the fibrinolytic system was associated with increased fibronectin turnover. Fibronectin-related material was detected immunohistochemically in dense clumps around SMCs and colocalized with latent TGF-β binding protein-1.

Conclusions: The fibrinolytic pathway could play a critical role in TAA progression, via direct or indirect impact on ECM and consecutive modulation of TGF-β bioavailability.
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http://dx.doi.org/10.1111/j.1365-2559.2010.03719.xDOI Listing
December 2010

Inter-individual variability of protein patterns in saliva of healthy adults.

J Proteomics 2009 Jul 23;72(5):822-30. Epub 2009 May 23.

INRA, UMR 1129 FLAVIC, F-21000 Dijon, France.

In order to document inter-individual variability in salivary protein patterns, unstimulated whole saliva was obtained from 12 subjects at 10 am and 3 pm of the same day. Saliva proteins were separated using two-dimensional gel electrophoresis, and semi-quantified using image analysis. One-way ANOVA was used to test the effects "time of sampling" and "subject". Data were further explored by multivariate analyses (PCA, hierarchical clustering). Spots of interest were identified by mass spectrometry (MALDI-TOF MS/MS and nanoLC ESI-IT MS/MS). A dataset of 509 spots matched in all gels was obtained. There was no diurnal statistical effect on salivary patterns while inter-individual variability was high with 47 spots differentially expressed between subjects (p<1%). Clustering of these spots revealed that subjects could be discriminated first based on several proteins participating to the non-specific immune response (cystatins, lipocalin 1, parotid-secretory protein and prolactin-induced protein). Independently, subjects were also differentiated by their level of proteins originating from serum and involved in the immune system (complement C3, transferrin, IgG2), as well as the relative abundance of enzymes involved in carbohydrates metabolism (amylase and glycolytic enzymes). Inter-individual variability should be accounted for when searching for salivary biomarkers or when studying in-mouth biochemical mechanisms.
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http://dx.doi.org/10.1016/j.jprot.2009.05.004DOI Listing
July 2009

Determination of normal human intrathyroidal iodine in Caracas population.

J Trace Elem Med Biol 2009 23;23(1):9-14. Epub 2009 Jan 23.

Analytical Chemistry Center, School of Chemistry, Faculty of Science, Central University of Venezuela, P.O. Box 40764, Caracas 1053, Venezuela.

This study focuses on the determination of iodine content in healthy thyroid samples on male population from Caracas, Venezuela. Contribution to establish a baseline of iodine content in thyroid glands and hence to compare the iodine thyroid concentration of the Venezuelan population with other countries is also our objective. Male post-mortem individual samples were analyzed using a spectrophotometric flow injection method, based on the Sandell-Kolthoff reaction. The median intrathyroidal iodine concentration was 1443+/-677 microg/g (wet weight), ranging from 419 to 3430 microg/g, which corresponds to a median of total iodine content of 15+/-8 mg (ranging from 4 to 37). These results were higher than those values reported in the literature. No correlation of iodine content with age or weight of the healthy gland was found.
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http://dx.doi.org/10.1016/j.jtemb.2008.11.002DOI Listing
July 2009

Analysis of size characterized manganese species from liver extracts using capillary zone electrophoresis coupled to inductively coupled plasma mass spectrometry (CZE-ICP-MS).

Anal Chim Acta 2006 Jul 4;573-574:172-80. Epub 2006 Apr 4.

Department of Chemistry, University of Carracas, Venezuela.

Mn is of toxicological concern because overexposure can lead to progressive, permanent neurodegenerative damage. Monomethyl-Mn-pentadienyl-tricarbonyl (MMT) is used as an anti-knock agent in fuel. Exhausted Mn compounds are absorbed in the lung and transported to the liver. Extended exposure causes an overflow of the liver with Mn species moving e.g. to the brain, causing irreversible central nervous system (CNS) disorders like Manganism. This paper focuses on experiments for getting more information on Mn species in liver extracts. The investigations are performed with respect to (1) a size characterization and (2) a subsequent identification of the Mn species in liver extracts using preparative size exclusion chromatography (SEC) followed by capillary zone electrophoresis coupled to inductively coupled plasma mass spectrometry (CZE-ICP-MS). First, extracts were analyzed using a mass calibrated SEC column coupled to ICP-MS detection. The chromatogram showed the 55 Mn-trace and proved main Mn elution between ca. 60-150 kDa. Second, liver extracts were fractionated on the same SEC column, however, now the effluent was directed to a fraction collector. This resulted in fractions containing pre-purified, size characterized Mn species per fraction. It turned out that the Mn concentrations per fraction reflected roughly the previous on-line Mn trace. Third, the fractions were subject to CZE-ICP-MS, where the MS was operated additionally with dynamic reaction cell (DRC) technique. From size characterization (with SEC coupled on-line to ICP-MS or connected to a fraction collector and subsequent Mn determination in fractions) it was shown that most Mn species from liver extract were of high molecular mass (HMM) nature as they eluted mostly between 50 and 80 min, corresponding to ca. 60-150 kDa. With the two-dimensional speciation approach employing first SEC and then CZE-ICP-DRC-MS together with standard addition method, a series of Mn species was identified. Mn species predominantly were Mn-enzymes e.g. arginase, isocitric dehydrogenase, galactosyltransferase, prolidase, pyruvate carboxylase and oxalate oxidase. A typical Mn-transporter--Mn-albumin-- was also seen, whilst Mn-transferrin obviously was degraded during SEC separation. This Mn-compound (independent whether as a standard or from liver extract) was not stable during SEC even at the finally chosen physiological conditions.
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http://dx.doi.org/10.1016/j.aca.2006.02.045DOI Listing
July 2006

Determination of selenium and iodine in human thyroids.

J Trace Elem Med Biol 2005 ;19(1):23-7

Analytical Chemistry Center, Chemistry School, Science Faculty, Central University of Venezuela, Zip Code 47102, Caracas 1041-A, Venezuela.

This study focuses on the determination of selenium and iodine in human thyroids. The glands were digested using nitric acid in a microwave oven. Selenium was determined by inductively coupled plasma optical emission spectrometry (ICP-OES) using a new sample introduction system consisting of a reduction system coupled to a hydride generation nebulizer (DHGN). Iodine was determined by using the Sandell-Kolthoff procedure. The detections limits were 0.2 ng/mL and 0.3 ng/mL for the determination method of selenium and iodine, respectively. The amount of iodine in the whole gland was 3.44 +/- 1.11 microg/g. The lowest iodine level was 2.34 microg/g and the highest 5.21 microg/g. The lowest selenium concentration for a single sample was 505 +/- 51 ng/g and the highest 1495 +/- 204 ng/g depending on the fraction of the gland selected.
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http://dx.doi.org/10.1016/j.jtemb.2005.07.005DOI Listing
November 2005