Papers by Alexandra Primikyri
ACS Chemical Biology, 2014
Xenotransplantation, Jan 27, 2016
The complement system plays a crucial role in acute xenogeneic reactions after cardiac transplant... more The complement system plays a crucial role in acute xenogeneic reactions after cardiac transplantation. We used an ex vivo perfusion model to investigate the effect of Cp40, a compstatin analog and potent inhibitor of complement at the level of C3. Fifteen wild-type pig hearts were explanted, cardiopleged, and reperfused ex vivo after 150 minutes of cold ischemia. Hearts were challenged in a biventricular working heart mode to evaluate cardiac perfusion and function. In the treatment group (n=5), the complement cascade was blocked at the level of C3 using Cp40, using diluted human blood. Untreated human and porcine blood was used for controls. Throughout the perfusion, C3 activation was inhibited when Cp40 was used (mean of all time points: 1.11 ± 0.34% vs 3.12 ± 0.48% control activation; P<.01). Compared to xenoperfused controls, the cardiac index improved significantly in the treated group (6.5 ± 4.2 vs 3.5 ± 4.8 mL/min/g; P=.03, 180 minutes perfusion), while the concentration ...
Journal of Chromatography B, 2017
Cp40 is a 14-amino acid cyclic analog of the peptidic complement inhibitor compstatin that binds ... more Cp40 is a 14-amino acid cyclic analog of the peptidic complement inhibitor compstatin that binds with sub-nanomolar affinity to complement component C3 and has already shown promise in various models of complement-related diseases. The preclinical and clinical development of this compound requires a robust, accurate, and sensitive method for quantitatively monitoring Cp40 in biological samples. In this study, we describe the development and validation of an ultra-high performance liquid chromatography electrospray mass spectrometry method for the quantitation of Cp40 in human and non-human primate (NHP) plasma. Isotope-labeled Cp40 was used as an internal standard, allowing for the accurate and absolute quantitation of Cp40. Labeled and non-labeled Cp40 were extracted from plasma using reversed phase-solid phase extraction, with recovery rates exceeding 80%, indicating minor matrix effects. The triply charged states of Cp40 and isotope-labeled Cp40 were detected at m/z 596.60 and 600.34, respectively, via a Q-TOF mass spectrometer and were used for quantitation. The method was linear in the range of 0.18-3.58μg/mL (r(2)≥0.99), with precision values below 0.71% in NHP and 0.77% in human plasma. The accuracy of the method ranged from -2.17% to 17.99% in NHP and from -0.26% to 15.75% in human plasma. The method was successfully applied to the quantitation of Cp40 in cynomolgus monkey plasma after an initial intravenous bolus of 2mg/kg followed by repetitive subcutaneous administration at 1mg/kg. The high reproducibility, accuracy, and robustness of the method developed here render it suitable for drug monitoring of Cp40, and potentially other compstatin analogs, in both human and NHP plasma samples during pharmacokinetic and pharmacodynamic studies.
Journal of Enzyme Inhibition and Medicinal Chemistry
Some new complexes of mefenamic acid with potentially interesting biological activity are describ... more Some new complexes of mefenamic acid with potentially interesting biological activity are described. The complexes of mefenamic acid [Mn(mef)(2)(H(2)O)(2)], 1, [Co(mef)(2)(H(2)O)(2)], 2, [Ni(mef)(2)(H(2)O)(2)], 3, [Cu(mef)(2)(H(2)O)](2), 4 and [Zn(mef)(2)], 5, were prepared by the reaction of mefenamic acid, a potent anti-inflammatory drug with metal salts. Optical and infrared spectral data of these new complexes are reported. Monomeric six-coordinated species were isolated in the solid state for Mn(II), Ni(II) and Co(II), dimeric five-coordinated for Cu(II) and monomeric four-coordinated for Zn(II). In DMF or CHCl(3) solution the coordination number is retained and the coordinated molecules of water are replaced by solvent molecules. The anti-oxidant properties of the complexes were evaluated using the 1,1-diphenyl-2-picrylhydrazyl, DPPH, free radical scavenging assay. The scavenging activities of the complexes were measured and compared with those of the free drug and vitamin C. ...
Journal of inorganic biochemistry
The novel triphenyltin(IV) esters of flufenamic acid (1), Hflu, [Ph(3)Sn(flu)] (2), and of [2-(2,... more The novel triphenyltin(IV) esters of flufenamic acid (1), Hflu, [Ph(3)Sn(flu)] (2), and of [2-(2,3-dichlorophenylamino)benzoic acid] (3), Hdcpa, [Ph(3)Sn(dcpa)] (4) have been structurally characterized by means of vibrational and (1)H, (13)C NMR spectroscopic studies. The crystal and molecular structures of [SnPh(3)(dcpa)(DMSO)] 4a are described. The molecular structure of 4a reveals that the Sn atom has a distorted trigonal bipyramidal coordination geometry with equatorial phenyl groups and the carboxylate and dimethylsulfoxide oxygen atoms occupying axial positions. The crystal structure of 4a is self-assembled by C-H-π and π-π stacking interactions. The in vitro cytotoxic activity of 1-4 and of the related non-steroidal anti-inflammatory drugs, NSAIDs, [2-(2,6-dimethylphenylamino)benzoic acid], Hdmpa (5), [Ph(3)Sn(dmpa)] (6), [2-(2,3-dimethylphenylamino)benzoic acid], mefenamic acid, Hmef (7) and [Ph(3)Sn(mef)] (8) has been evaluated against the cancer cell lines MCF-7, T-24, A-5...
European Journal of Cancer Supplements, 2008
XRF). methods. The compounds were incubated with target cells for 72 h. At the end of this incuba... more XRF). methods. The compounds were incubated with target cells for 72 h. At the end of this incubation period, antiproliferative activity in vitro was determined by the MTT assay. In this work we have analysed the cell cycle phase distribution of compounds treated (2IC50) and untreated malignant cells, 24h, 48h, and 72h, after the treatment. After cell staining with propidium iodide cells were analyzed using a Becton Dickinson FAC-Scan flow cytometer.
European Journal of Cancer Supplements, 2008
this strategy, we intend to enhance the efficiency of cancer therapies, and cover up possible res... more this strategy, we intend to enhance the efficiency of cancer therapies, and cover up possible resistance to individual treatments. Studies are in progress to assess the in vitro efficacy of these combined treatments. In parallel, we perform preclinical trials to investigate the importance of these mechanisms for tumor formation and metastasis in vivo. The efficacy of these treatments is first investigated in chicken metastasis assays. Besides, we have generated transgenic mice over-expressing Met in a temporally and spatially regulated manner. Met-overexpressing cells also express the luciferase, for non-invasive monitoring of primary tumours and metastasis. These mice are precious tools to better understand Mettriggered tumorigenesis and validate the efficiency of combined treatment to un-favour cell survival in Met-triggered cancers.
European Journal of Cancer Supplements, 2010
Journal of Inorganic Biochemistry, 2009
The complexes [Me 2 (Meclo)SnOSn(Meclo)Me 2 ] 2 (2) and [Ph 3 Sn(Meclo)] (3) where HMeclo is mecl... more The complexes [Me 2 (Meclo)SnOSn(Meclo)Me 2 ] 2 (2) and [Ph 3 Sn(Meclo)] (3) where HMeclo is meclofenamic acid, N-(2,6-dichloro-m-tolylanthranilic acid)], have been prepared and structurally characterized by means of vibrational, 1 H and 13 C NMR spectroscopies. The crystal structure of complexes (2) and (3) have been determined by X-ray crystallography. Three distannoxane rings are present to the dimeric tetraorganodistannoxane of planar ladder arrangement of (2). The structure is centro symmetric and features a central rhombus Sn 2 O 2 unit two additional tin atoms linked at the oxygen atoms. Five-and six-coordinated tin centers are present in the dimer distannoxane. X-ray analysis of (3) revealed a penta-coordinated structure containing Ph 3 Sn coordinated to the chelated carboxylato group. The polar imino hydrogen atom participates in intra-molecular hydrogen bonds. Complexes and (3) are self-assembled via p ? p, C-H-p, stacking interactions and intra-molecular hydrogen bonds. Meclofenamic acid and [Ph 3 Sn(Meclo)] have been evaluated for antiproliferative activity in vitro against three human cancer cell lines: MCF-7 (human breast cancer cell line), T24 (bladder cancer cell line), A-549 (non-small cell lung carcinoma) and a mouse L-929 (a fibroblast-like cell line cloned from strain L). The [Ph 3 Sn(Meclo)] complex exhibited high cytotoxic activity against all the cancer cell lines. Meclofenamic and [Ph 3 Sn(Meclo)]
Journal of Inorganic Biochemistry, 2010
a b s t r a c t 2-Acetyl pyridine thiosemicarbazone containing an 1-(4-fluorophenyl)-piperazinyl ... more a b s t r a c t 2-Acetyl pyridine thiosemicarbazone containing an 1-(4-fluorophenyl)-piperazinyl ring incorporated at N(4)-position, HAcPipPheF (1) and the zinc(II) complexes [Zn(AcPipPheF) 2 ] (2) and [Zn(OAc)(AcPip-PheF)] 2 (3) have been prepared and structurally characterized by means of vibrational and NMR ( 1 H and 13 C) spectroscopy. The crystal structures of the compounds 1-3 have been determined by X-ray crystallography. The metal coordination geometry of [Zn(AcPipPheF) 2 ] is described as distorted octahedral configuration in a trans-N-cis-N-cis-S configuration. In [Zn(OAc)(AcPipPheF)] 2 one of the acetato group exhibits monoatomic bridge and the other bridges in a bidentate manner. The zinc(1) metal ion is coordinated in a distorted octahedral configuration while the metal coordination of Zn(2) is described as distorted square pyramidal. Biomedical studies revealed that, compounds 1-3 displayed potent anticancer activity. The antiproliferative activity of 1-3 was found to be considerably stronger than that of cis-platin. The IC 50 values range from 26 to 90 nM, against all cell lines tested, while for cis-platin the IC 50 values range from 2 to 17 lM and for the zinc salt, ZnCl 2 , the IC 50 values range from 81 to 93 lM. The complex 3 shows the highest activity against all four cancer cell lines and the highest selectivity against K562 and MDA-MB-453 cancer cell lines. The compounds inhibited tumor cell proliferation by arresting the cell cycle progression at the S phase.
Chemistry & Biodiversity, 2009
Some new complexes of tolfenamic acid ( ¼ 2-[(2-methyl-3-chlorophenyl)amino]benzoic acid; Htolf) ... more Some new complexes of tolfenamic acid ( ¼ 2-[(2-methyl-3-chlorophenyl)amino]benzoic acid; Htolf) with potentially interesting biological activities are described. The complexes [Mn(tolf) 2 (H 2 O) 2 ], [Co(tolf) 2 (H 2 O) 2 ], [Ni(tolf 2 (H 2 O) 2 ], [Cu(tolf) 2 (H 2 O)] 2 , and [Zn(tolf) 2 (H 2 O)] were prepared by the reaction of tolfenamic acid, a potent anti-inflammatory drug, with metal salts. The radical-scavenging activities of the complexes were evaluated using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radicalscavenging assay. Their ability to inhibit soybean lipoxygenase, b-glucuronidase, and trypsin-induced proteolysis was studied. Their inhibitory effects on rat paw edema induced by carrageenin was studied and compared with those of tolfenamic acid. The complex [Zn(tolf) 2 (H 2 O)] exhibited the strongest in vivo inhibitory effect at 0.1 mm/kg Body Weight (BW; 93.0 AE 0.9%), superior than the inhibition induced by tolfenamic acid at the same molar dose (76.0 AE 0.9%). Tolfenamic acid and its metal complexes have been evaluated for antiproliferative activity in vitro against the cells of three human cancer cell lines, MCF-7 (breast cancer cell line), T24 (bladder cancer cell line), and A-549 (non-small cell lung carcinoma), and a mouse fibroblast L-929 cell line. The complexes [Mn(tolf) 2 (H 2 O) 2 ] and [Cu-(tolf) 2 (H 2 O)] 2 have shown selectivity against T24 cell line. The IC 50 values of these two complexes against T24 cancer cell lines are in a micromolar range similar or better to that of the antitumor drug cisplatin.
Organic & Biomolecular Chemistry
Unprecedented regioselective acylation of flavonoid aglycones was achieved using Candida antarcti... more Unprecedented regioselective acylation of flavonoid aglycones was achieved using Candida antarctica lipase B (CALB). The rapid screening of product formation was performed by the use of the high resolution phenol-type OH (1)H NMR spectral region recorded after the addition of picric acid.
ABSTRACT Deregulation of apoptosis contributes largely to the pathogenesis of lymphoid neoplasms ... more ABSTRACT Deregulation of apoptosis contributes largely to the pathogenesis of lymphoid neoplasms allowing the expansion of cell clones that develop resistance to drug-induced cell death. Therefore, subverting inherent resistance to apoptosis is a very challenging therapeutic issue and developing small molecules that can mimic the BH3 domain of the pro-apoptotic Bcl-2 family proteins appears to offer such potentials. Natural products, a rich source of compounds with BH3 mimetic activity, make a sound basis for such an approach. Quercetin, a natural polyphenol, has drawn much attention because it exerts anticancer cytotoxic effects, while sparing normal cells. Pro-apoptotic effects of quercetin have been shown in human lymphoma and leukemic cells and have been associated with Bcl-2 and Mcl-1 downregulation and Bax conformational activation. We undertook a multidisciplinary approach to elucidate the mode of action of quercetin, including cytotoxicity cell assays, biochemical approaches (pull down assays), NMR spectroscopy and docking calculations. We demonstrate that quercetin binds directly to the BH3 domain of the anti-apoptotic Bcl-2 and Bcl-xL proteins, exhibiting BH3-mimetic properties. Specifically, functional cytotoxicity assays of quercetin in Jurkat T-cell leukemic lines, Jurkat Bcl-2 and Jurkat Puro, indicated that quercetin binds to Bcl-2 protein. The direct binding of quercetin to Bcl-2 and to Bcl-xL was biochemically validated using pull down assays. NMR chemical shift perturbation experiments and docking calculations finally revealed that quercetin was bound to the pro-apoptotic BH3-binding site of the anti-apoptotic Bcl-2 family proteins (Figure 1). This property classifies quercetin as a natural flavonoid with BH3 mimetic activity, capable of driving leukemic cells to apoptosis. These results explain the cytotoxic effects of quercetin on Jurkat cells. These data can serve as a basis to consider studying BH3 mimetic natural products as adjuncts in the therapeutic approaches of lymphoid neoplasms in combination with chemotherapy, and can also provide a starting structure for developing novel potent analogues for the treatment of blood cancers.
ACS Chemical Biology, 2014
Bcl-2 family proteins are important regulators of apoptosis and its antiapoptotic members, which ... more Bcl-2 family proteins are important regulators of apoptosis and its antiapoptotic members, which are overexpressed in many types of cancer, are of high prognostic significance, establishing them as attractive therapeutic targets. Quercetin, a natural flavonoid, has drawn much attention because it exerts anticancer effects, while sparing normal cells. A multidisciplinary approach has been employed herein, in an effort to reveal its mode of action including dose-response antiproliferative activity and induced apoptosis effect, biochemical and physicochemical assays, and computational calculations. It may be concluded that, quercetin binds directly to the BH3 domain of Bcl-2 and Bcl-xL proteins, thereby inhibiting their activity and promoting cancer cell apoptosis.
The Journal of Physical Chemistry B, 2015
The Zn(II) chelation with natural flavonoids, quercetin and luteolin, was investigated by the use... more The Zn(II) chelation with natural flavonoids, quercetin and luteolin, was investigated by the use of NMR spectroscopy and various levels of ab initio calculations. Very sharp phenolic OH (1)H resonances in DMSO-d6 were observed for both free and complexed quercetin which allowed (i) the unequivocal assignment with the combined use of (1)H-(13)C HSQC and HMBC experiments and (ii) the determination of complexation sites which were found to be the CO-4 carbonyl oxygen and the deprotonated C-5 OH group of quercetin and CO-4 carbonyl oxygen and the deprotonated C-5 OH group of luteolin. DOSY experiments allowed the determination of the effective molecular weight of the Zn-quercetin complex which was shown to be mainly 1:1. DFT calculations of the 1:1 complex in the gas phase demonstrated that the C-3 O(-) and CO-4 sites are favored for quercetin at both GGA and LDA approximations and the C-5 O(-) and CO-4 groups of luteolin at the LDA approximation. Quantum chemical calculations were also performed by means of the conductor polarizable model in DMSO by employing various functionals. The energetically favored Zn chelation sites of the 1:1 complex were found to be either the C-3 O(-) and CO-4 or C-5 O(-) and CO-4 sites, depending on the functional used, for quercetin and the C-5 O(-) and CO-4 sites for luteolin.
Tetrahedron, 2012
ABSTRACT A convenient DOSY methodology was developed that can be applied directly in crude reacti... more ABSTRACT A convenient DOSY methodology was developed that can be applied directly in crude reaction products or mixtures containing polyphenol organic compounds, for the rapid identification of their various components without any prior separation or isolation. The method is based on the resolution enhancement of the resonances of the –OH protons and the fine-tuning of their diffusion coefficients to the molecular diffusion coefficient; this can be achieved in DMSO-d6 in combination with the addition of picric acid and the use of temperatures near the freezing point of the solution. This method, which does not modify the apparent molecular diffusion, allowed the recording of high resolution DOSY spectra, both in crude enzymatic reactions and mixtures of organic compounds based on the phenolic OH NMR spectral region which is much less crowded and, thus, much more informative compared to the aromatic region.
Organic & Biomolecular Chemistry, 2012
Unprecedented regioselective acylation of flavonoid aglycones was achieved using Candida antarcti... more Unprecedented regioselective acylation of flavonoid aglycones was achieved using Candida antarctica lipase B (CALB). The rapid screening of product formation was performed by the use of the high resolution phenol-type OH (1)H NMR spectral region recorded after the addition of picric acid.
Organic & Biomolecular Chemistry, 2013
Correlations between hydrogen bonds and solvent effects on phenol -OH proton shieldings, temperat... more Correlations between hydrogen bonds and solvent effects on phenol -OH proton shieldings, temperature coefficients (Δδ/ΔT) and effects on OH diffusion coefficients for numerous phenolic acids, flavonols, flavones, and oleuropein derivatives of biological interest were investigated in several organic solvents and were shown to serve as reliable indicators of hydrogen bonding and solvation state of -OH groups. The temperature coefficients span a range of -0.5 to -12.3 ppb K(-1). Shielding differences of 2.0 to 2.9 ppm at 298 K were observed for solvent exposed OH groups between DMSO-d(6) and CD(3)CN which should be compared with a shielding range of ~7 ppm. This demonstrates that the solvation state of hydroxyl protons is a key factor in determining the value of the chemical shift. For -OH protons showing temperature gradients more positive than -2.5 ppb K(-1), shielding changes between DMSO-d(6) and CD(3)CN below 0.6 ppm, and diffusion coefficients significantly different from those of traces of H(2)O, there is an intramolecular hydrogen bond predictivity value of 100%. The C-3 OH protons of flavonols show very significant negative temperature coefficients and shielding changes between DMSO-d(6) and CD(3)CN of ~2.3 ppm, which indicate the absence of persistent intramolecular hydrogen bonds, contrary to numerous X-ray structures.
Journal of Natural Products, 2011
A general method is demonstrated for obtaining ultra-high resolution in the phenolic hydroxy grou... more A general method is demonstrated for obtaining ultra-high resolution in the phenolic hydroxy group 1 H NMR spectroscopic region, in DMSO-d 6 solution, with the addition of picric acid. Line-width reduction by a factor of over 100 was observed, which resulted in line-widths ranging from 1.6 to 0.6 Hz. This unprecedented resolution, in combination with the shielding sensitivity of the hydroxy group absorptions to substituent effects at least up to 11 bonds distant and the application of 2D 1 H− 13 C HMBC techniques, allows the unequivocal structure analysis of natural products with phenolic hydroxy groups in complex plant extracts.
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Papers by Alexandra Primikyri