Evaluation of Rhenium and Technetium-99m Complexes Bearing Quinazoline Derivatives as Potential EGFR Agents.

Τhe Epidermal Growth Factor Receptor tyrosine kinase inhibitor (EGFR-TKI) 6-amino-4-[(3-bromophenyl) amino]quinazoline was derivatized with 6-bromohexanoyl-chloride and coupled with the tridentate chelating agents N-(2-pyridylmethyl) aminoethyl acetic acid (PAMA) and L(+)-cysteine bearing the donor atom set NNO and SNO, respectively. The rhenium precursors ReBr(CO)5 and fac-[NEt4]2[ReBr3(CO)3] were used for the preparation of the Re complexes fac-[Re(NNO)(CO)3] (5a) and fac-[Re(SNO)(CO)3] (7a) which were characterized by NMR and IR spectroscopies. Subsequently, the new potential EGFR inhibitors were labeled with the fac-[99mTc(CO)3]+ core in high yield and radiochemical purity (>90%) by ligand exchange reaction using the fac-[99mTc][Tc(OH2)3(CO)3]+ precursor. The radiolabeled complexes were characterized by comparative HPLC analysis with the analogous rhenium (Re) complexes as references. In vitro studies in the A431 cell lines showed that both ligands and Re complexes inhibit A431 cell growth. Complex 5a demonstrated the highest potency (IC50 = 8.85 ± 2.62 µM) and was further assessed for its capacity to inhibit EGFR autophosphorylation, presenting an IC50 value of 26.11 nM. Biodistribution studies of the 99mTc complexes in healthy mice showed high in vivo stability for both complexes and fast blood and soft tissue clearance with excretion occurring via the hepatobiliary system.

Synthesis and preclinical evaluation of rhenium and technetium-99m "4 + 1" mixed-ligand complexes bearing quinazoline derivatives as potential EGFR imaging agents.

Epidermal growth factor receptors (EGFR) of tyrosine kinase (TK) have shown high expression levels in most cancers and are considered a promising target for cancer diagnosis and therapy. Expanding the investigation for novel targeted radiopharmaceuticals, an EGFR inhibitor such as 4-aminoquinazoline derivatives along with a radionuclide such as technetium-99m (99mTc) could be ideal. Thus, we report herein the synthesis, characterization, and biological evaluation of new "4 + 1" mixed-ligand ReIII- and 99mTcIII-complexes of the general formula [99mTc][Tc(NS3)(CN-R)] bearing tris(2-mercaptoethyl)-amine (NS3) as the tetradentate tripodal ligand and a series of isocyanide derivatives (CN-R) of tyrosine kinase inhibitor (3-bromophenyl)quinazoline-4,6-diamine as the monodentate ligand. The quinazoline isocyanide derivatives 4a-d were prepared in two steps and reacted with the [Re(NS3)PMe2Ph] precursor leading to the final complexes 5a-d in high yield. All compounds were characterized by elemental analysis, IR, and NMR spectroscopies. In vitro studies, for their potency to inhibit the cell growth, using intact A431 cells indicate that the quinazoline derivatives 4a-d and the Re complexes 5a-d significantly inhibit the A431 cell growth. In addition, the EGFR autophosphorylation study of complex 5b shows an IC50 value in the nanomolar range. The corresponding "4 + 1" 99mTc-complexes 6a-d were prepared by employing the [99mTc]TcEDTA intermediate and the appropriate monodentate 4a-d in a two-step synthetic procedure with a radiochemical yield (RCY) from 63 to 77 % and a radiochemical purity (RCP) > 99 % after HPLC purification. Their structures have been established by HPLC comparative studies using the well-characterized Re-complexes 5a-d as reference. All 99mTc-complexes remain stable for at least 6 h, and their logD7.4 values confirmed their anticipated lipophilic character. Biodistribution studies in healthy Swiss albino mice of 99mTc-complexes showed hepatobiliary excretion and initial fast blood clearance. Complex 6b was also tested in Albino SCID mice bearing A431 tumors and showed rapid tumor uptake at 5 min (2.80 % ID/g) with a moderate tumor/muscle ratio (2.06) at 4 h p.i. The results encourage further investigation for this type of 99mTc-complexes as single-photon emission computed tomography (SPECT) radio agents for imaging tumors overexpressing EGFR.

Remdesivir-loaded bis-MPA hyperbranched dendritic nanocarriers for pulmonary delivery.

Remdesivir is the only clinically available antiviral drug for the treatment of COVID-19. However, its very limited aqueous solubility confines its therapeutic activity and the development of novel inhaled nano-based drug delivery systems of remdesivir for enhanced lung tissue targeting and efficacy is internationally pursued. In this work 2,2-bis(hydroxymethyl)propionic acid (bis-MPA) hyperbranched dendritic nano-scaffolds were employed as nanocarriers of remdesivir. The produced nano-formulations, empty and loaded, consisted of monodisperse nanoparticles with spherical morphology and neutral surface charge and sizes ranging between 80 and 230 nm. The entrapment efficiency and loading capacity of the loaded samples were 82.0% and 14.1%, respectively, whereas the release of the encapsulated drug was complete after 48 h. The toxicity assays in healthy MRC-5 lung diploid fibroblasts and NR8383 alveolar macrophages indicated their suitability as potential remdesivir carriers in the respiratory system. The novel nano-formulations are non-toxic in both tested cell lines, with IC50 values higher than 400 µΜ after 72 h treatment. Moreover, both free and encapsulated remdesivir exhibited very similar IC50 values, at the range of 80-90 µM, while its aqueous solubility was increased, overall presenting a suitable profile for application in inhaled delivery of therapeutics.

Effective Labeling of Amine Pharmacophores through the Employment of 2,3-Pyrazinedicarboxylic Anhydride and the Generation of fac-[M(CO)3(PyA)P] and cis-trans-[M(CO)2(PyA)P2] Complexes (PyA = Pyrazine-2-carboxylate, P = Phosphine, M = Re, 99mTc).

The fac-[M(CO)3(PyA)(P)] and cis-trans-[M(CO)2(PyA)(P)2] neutral complexes (M is Re or 99mTc), based on the mixed ligand strategy with pyrazine-2-carboxylic acid (PyAH) as the bidentate N,O and triphenylphosphine as the monodentate P ligand, are presented. Through the employment of the anhydride of pyrazine-2,3-dicarboxylic acid (PyDA), the PyAH scaffold was conveniently derivatized with the model bioactive amine 1-(2-methoxyphenyl)piperazine, the active part of the 5-HT1A antagonist WAY100635. Reaction of either PyAH or the pharmacophore-bearing PyAH ligand (L1H) with fac-[M(CO)3]+ core in water yielded the intermediate fac-[M(CO)3(PyA)(H2O)] complexes. The labile aqua ligand was easily replaced by PPh3 to yield the fac-[Re(CO)3(PyA)(PPh3)] complexes, while in toluene under reflux, the cis-trans-[Re(CO)2(PyA)(PPh3)2] complexes were obtained. The latter complexes were alternatively obtained from mer-[Re(CO)3(PPh3)2Cl] by refluxing with the PyA ligand in toluene. The analogous 99mTc complexes were synthesized quantitatively, showing excellent stability in competition studies. The methodology described herein represents a practical procedure for the effective integration of the fac-[M(CO)3]+ core with amine-bearing biologically active compounds for diagnosis/therapy.

Synthesis and evaluation of new mixed "2 + 1" Re, 99mTc and 186Re tricarbonyl dithiocarbamate complexes with different monodentate ligands.

A series of "2 + 1" mixed ligand tricarbonyl complexes of the general formula fac-[Re/99mTc/186Re(CO)3(DDTC)(L)] containing diethyldithiocarbamate (DDTC) as a monoanionic bidentate ligand and a series of monodentate ligands L was synthesized, characterized and evaluated. The impact of ligand L on the radiochemical yield (RCY) and biodistribution of the final compounds was also investigated. DDTC and the appropriate L ligand [cyclohexyl isocyanide (cisc), tert-butyl isocyanide (tbi), triphenylphosphine (PPh3), methyldiphenylphosphine (PPh2Me), triphenylarsine (AsPh3), imidazole (im), and 4-aminopyridine (4AP)] readily reacted in equimolar amounts with the [Et4N]2[Re(CO)3Br3] precursor to afford fac-[Re(CO)3(DDTC)(cisc)], Re1, fac-[Re(CO)3(DDTC)(tbi)], Re2, fac-[Re(CO)3(DDTC)(PPh3)], Re3, fac-[Re(CO)3(DDTC)(PPh2Me)], Re4, fac-[Re(CO)3(DDTC)(AsPh3)], Re5, fac-[Re(CO)3(DDTC)(im)], Re6 and fac-[Re(CO)3(DDTC)(4AP)], Re7, complexes in high yields (>80%). All Re complexes were fully characterized by IR, NMR, and in addition Re4, Re5, and Re7 with X-ray crystallography. Analogous reactions as performed with Re were subsequently explored on the 99mTc and 186Re-tracer levels using the corresponding fac-[99mTc/186Re(CO)3(H2O)3]+ precursor. Complexes 99mTc1 - 99mTc5, 186Re1 and 186Re3 were obtained in high radiochemical yield (>91%), while the complexes 99mTc6, 99mTc7 and 186Re7 formed with radiochemical yields of 55%, 28%, and 75%, respectively. The 99mTc and 186Re-complexes were characterized by comparative HPLC analysis using the analogous Re complexes. During histidine and cysteine challenge experiments at 37 °C through 6 h, complexes 99mTc1 - 99mTc5 remained > 92% stable, while complexes 99mTc6 and 99mTc7 remained only 8% stable through 3 h. Similar studies for 186Re-complexes showed that 186Re1 and 186Re3 remained > 95% stable for up to 48 h, while 186Re7 had decreased to 7% after 3 h. LogD7.4 data of 99mTc1 - 99mTc5, 186Re1, and 186Re3 complexes, which ranged from 2.59 to 3.39, suggested high lipophilicity. Biodistribution studies in healthy Swiss albino mice showed hepatobiliary excretion for 99mTc1, 99mTc2, and 99mTc4, fast blood clearance for 99mTc4, while high liver uptake and retention for 99mTc3 and 99mTc5 were measured. Moreover, 99mTc2 showed high accumulation in the lungs with sustained retention (52.80% ID/g at 4 h p.i.) and significant brain uptake at 2 min p.i. (1.89% ID/g). The study showed the great influence of monodentate ligand in the synthesis and biodistribution of the mixed ligand complexes.

Synthesis and antimicrobial evaluation of a pyrazoline-pyridine silver(I) complex: DNA-interaction and anti-biofilm activity.

The emergence of resistant bacterial strains mainly due to misuse of antibiotics has seriously affected our ability to treat bacterial illness, and the development of new classes of potent antimicrobial agents is desperately needed. In this study, we report the efficient synthesis of a new pyrazoline-pyridine containing ligand L1 which acts as an NN-donor for the formation of a novel silver (I) complex 2. The free ligand did not show antibacterial activity. High potency was exhibited by the complex against three Gram-negative bacteria, namely Escherichia coli, Pseudomonas aeruginosa and Acinetobacter baumanii with the minimum inhibitory concentration (MIC) ranging between 4 and 16 µg/mL (4.2-16.7 µM), and excellent activity against the fungi Candida albicans and Cryptococcus neoformans (MIC ≤ 0.25 µg/mL = 0.26 µM). Moreover, no hemolytic activity within the tested concentration range was observed. In addition to the planktonic growth inhibition, the biofilm formation of both Methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa was significantly reduced by the complex at MIC concentrations in a dose-dependent manner for Pseudomonas aeruginosa, whereas a biphasic response was obtained for MRSA showing that the sub-MIC doses enhanced biofilm formation before its reduction at higher concentration. Finally, complex 2 exhibited strong DNA binding with a large drop in DNA viscosity indicating the absence of classical intercalation and suggesting the participation of the silver ion in DNA binding which may be related to its antibacterial activity. Taken together, the current results reveal that the pyrazoline-pyridine silver complexes are of high interest as novel antibacterial agents, justifying further in vitro and in vivo investigation.

Comparative Study of a Series of 99mTc(CO)3 Mannosylated Dextran Derivatives for Sentinel Lymph Node Detection.

Sentinel lymph node detection (SLND) is rapidly entering common practice in the management of patients with tumors. The introduction of mannose molecules to 99mTc-labeled dextrans, so far, showed that the sentinel node could trap these agents due to their recognition by the mannose receptors of lymph node macrophages. The current study aimed to synthesize, characterize, and biologically evaluate a series of mannosylated dextran derivatives labeled with 99mTc for potential use in SLND. The compounds were designed to have a dextran with a molecular weight of 10-500 kDa as a backbone, S-derivatized cysteines, efficient SNO chelators, and mannose moieties for binding to mannose receptors. They were successfully synthesized, thoroughly characterized using NMR techniques, and labeled with the fac-[99mTc(CO)3]+ synthon. Labeling with high yields and radiochemical purities was achieved with all derivatives. In vivo biodistribution and imaging studies demonstrated high uptake in the first lymph node and low uptakes in the following node and confirmed the ability to visualize the SLN. Among the compounds studied, 99mTc-D75CM demonstrated the most attractive biological features, and in combination with the high radiochemical yield and stability of the compound, its further evaluation as a new radiopharmaceutical for sentinel lymph node detection was justified.

Liposomes Decorated with 2-(4'-Aminophenyl)benzothiazole Effectively Inhibit Aß1-42 Fibril Formation and Exhibit in Vitro Brain-Targeting Potential.

The potential of 2-benzothiazolyl-decorated liposomes as theragnostic systems for Alzheimer's disease was evaluated in vitro, using PEGylated liposomes that were decorated with two types of 2-benzothiazoles: (i) the unsubstituted 2-benzothiazole (BTH) and (ii) the 2-(4-aminophenyl)benzothiazole (AP-BTH). The lipid derivatives of both BTH-lipid and AP-BTH-lipid were synthesized, for insertion in liposome membranes. Liposomes (LIP) containing three different concentrations of benzothiazoles (5, 10, and 20%) were formulated, and their stability, integrity in the presence of serum proteins, and their ability to inhibit ß-amyloid (1-42) (Αß42) peptide aggregation (by circular dichroism (CD) and thioflavin T (ThT) assay), were evaluated. Additionally, the interaction of some LIP with an in vitro model of the blood-brain barrier (BBB) was studied. All liposome types ranged between 92 and 105 nm, with the exception of the 20% AP-BTH-LIP that were larger (180 nm). The 5 and 10% AP-BTH-LIP were stable when stored at 4 °C for 40 days and demonstrated high integrity in the presence of serum proteins for 7 days at 37 °C. Interestingly, CD experiments revealed that the AP-BTH-LIP substantially interacted with Αß42 peptides and inhibited fibril formation, as verified by ThT assay, in contrast with the BTH-LIP, which had no effect. The 5 and 10% AP-BTH-LIP were the most effective in inhibiting Αß42 fibril formation. Surprisingly, the AP-BTH-LIP, especially the 5% ones, demonstrated high interaction with brain endothelial cells and high capability to be transported across the BBB model. Taken together, the current results reveal that the 5% AP-BTH-LIP are of high interest as novel targeted theragnostic systems against AD, justifying further in vitro and in vivo exploitation.

Synthesis, characterization and evaluation of 68Ga labelled monomeric and dimeric quinazoline derivatives of the HBED-CC chelator targeting the epidermal growth factor receptor.

Tyrosine kinase (TK) receptors including epidermal growth factor receptors (EGFRs) are known to be overexpressed in a wide variety of solid tumors associated with poor prognosis. The HBED-CC chelator N,N'-bis[2-hydroxy-5-(carboxyethyl)benzyl]ethylenediamine-N,N'-diacetic acid 1 was coupled via one or both its propionic acid moieties with the quinazoline EGFR-TK inhibiting pharmacophore 4-amino-N-(4-((3-bromophenyl)amino)quinazolin-6-yl)butanamide 3 resulting in either a monomeric 4 or a dimeric 5 species. Ligands 4 and 5 reacted with Ga3+ generating the corresponding complexes Ga4 and Ga5. Both ligands and complexes were characterized with mass spectrometry and NMR spectroscopy and evaluated in vitro with MTT assays in A431 cells, where they showed IC50 values in the range 51.6 to 68.8 µM. Labeling of ligands 4 and 5 with the PET radionuclide 68Ga was quantitative and resulted in tracers [68Ga]Ga4 and [68Ga]Ga5 with radiochemical purities greater than 98%, which were also characterised by comparative RP-HPLC studies with Ga4 and Ga5 respectively. Radiotracers [68Ga]Ga4 and [68Ga]Ga5 were stable (intact tracer over 98%) in the reaction mixture (120 min) and in human serum (30 min). Both tracers were evaluated in vivo with biodistribution experiments in SCID mice bearing A431 tumors presenting tumor uptake of 1.34 for [68Ga]Ga4 and 1.01 %ID/g for [68Ga]Ga5 at 5 min, which was slightly decreased at 60 min p.i. and then remained stable until 120 min p.i. To the best of our knowledge, this is the first report of monomeric and dimeric quinazoline conjugates with the chelator HBED-CC, which can serve as a basis for further development of EGFR-TKI targeting tracers.

Small Multitarget Molecules Incorporating the Enone Moiety.

Chalcones represent a class of small drug/druglike molecules with different and multitarget biological activities. Small multi-target drugs have attracted considerable interest in the last decade due their advantages in the treatment of complex and multifactorial diseases, since "one drug-one target" therapies have failed in many cases to demonstrate clinical efficacy. In this context, we designed and synthesized potential new small multi-target agents with lipoxygenase (LOX), acetyl cholinesterase (AChE) and lipid peroxidation inhibitory activities, as well as antioxidant activity based on 2-/4- hydroxy-chalcones and the bis-etherified bis-chalcone skeleton. Furthermore, the synthesized molecules were evaluated for their cytotoxicity. Simple chalcone b4 presents significant inhibitory activity against the 15-human LOX with an IC50 value 9.5 µM, interesting anti-AChE activity, and anti-lipid peroxidation behavior. Bis-etherified chalcone c12 is the most potent inhibitor of AChE within the bis-etherified bis-chalcones followed by c11. Bis-chalcones c11 and c12 were found to combine anti-LOX, anti-AchE, and anti-lipid peroxidation activities. It seems that the anti-lipid peroxidation activity supports the anti-LOX activity for the significantly active bis-chalcones. Our circular dichroism (CD) study identified two structures capable of interfering with the aggregation process of Aß. Compounds c2 and c4 display additional protective actions against Alzheimer's disease (AD) and add to the pleiotropic profile of the chalcone derivatives. Predicted results indicate that the majority of the compounds with the exception of c11 (144 Å) can cross the Blood Brain Barrier (BBB) and act in CNS. The results led us to propose new leads and to conclude that the presence of a double enone group supports better biological activities.

Dicarbonyl cis-[M(CO)2(N,O)(C)(P)] (M = Re, 99mTc) Complexes with a New [2 + 1 + 1] Donor Atom Combination.

The synthesis and characterization of the dicarbonyl mixed ligand cis-[Re(CO)2(quin)(cisc)(PPh3)] complex, 4, where quin is the deprotonated quinaldic acid, cisc is cyclohexyl isocyanide, and PPh3 is triphenylphosphine, is presented. The synthesis of 4 proceeds in three steps. In the first, the intermediate fac-[Re(CO)3(quin)(H2O)] aqua complex 2 is generated from the fac-[NEt4]2[Re(CO)3Br3] precursor, together with the brominated products fac-[Re(CO)3(quinH)(Br)] 1a and fac-[NEt4][Re(CO)3(quin)(Br)] 1b, in low yield. In the following step, replacement of the aqua ligand of complex 2 by the monodentate isocyanide ligand leads to the formation of fac-[Re(CO)3(quin)(cisc)], 3. In the third step replacement of the species trans to the isocyanide carbonyl group of 3 by a phosphine generates complex 4. The Re complexes 2-4 were prepared in high yield and fully characterized by elemental analysis, spectroscopic methods, and X-ray crystallography. At the technetium-99m (99mTc) tracer level, the analogous complexes 3' and 4' were produced in high radiochemical purity, characterized by comparative reverse phase high-performance liquid chromatography and showed high resistance to transchelation by histidine or cysteine. This new [N,O][C][P] donor atom combination with the cis-[M(CO)2]+ core (M = Re, 99mTc) is a promising scaffold for the development of novel diagnostic and therapeutic targeted radiopharmaceuticals.

New labeled derivatives of the neuroprotective peptide colivelin: synthesis, characterization, and first in vitro and in vivo applications.

Colivelin (CL), first reported in 2005, is the most potent member of the humanin family of neuroprotective peptides with in vitro and in vivo rescuing action against insults associated with Alzheimer's disease (AD). The objective of the present work is the design, synthesis and characterization of specific CL derivatives that can be used as molecular probes in the investigation of the unknown mechanism of CL action. Within this framework, three CL derivatives bearing suitable tags, i.e., the fluorescent moiety FITC, the streptavidin-counterpart biotinyl-group, and the (99m)Tc-radiometal chelating unit dimethylGly-Ser-Cys, were developed and subsequently applied in biological evaluation experiments. Specifically, the FITC-labeled derivative of CL was used in confocal microscopy, where specific binding at the periphery of F11 cells was observed; the biotin-labeled derivative of CL was used in an in-house developed ELISA-type assay, where specific and concentration-dependent binding with the ß-amyloid peptide of AD was shown; finally, the (99m)Tc-radiolabeled derivative of CL was used in in vivo biodistribution studies in healthy Swiss Albino mice, where 0.58% of the radioactivity administered was measured in the mouse brain 2min after injection. The above first successful applications of the CL probes demonstrate their potential to contribute in the field of neuroprotective peptides.

Amphiphilic Chlorin-ß-cyclodextrin Conjugates in Photo-Triggered Drug Delivery: The Role of Aggregation.

Conjugates of chlorins with ß-cyclodextrin connected either directly or via a flexible linker were prepared. In aqueous medium these amphiphilic conjugates were photostable, produced singlet oxygen at a rate similar to clinically used temoporfin and formed irregular nanoparticles via aggregation. Successful loading with the chemotherapeutic drug tamoxifen was evidenced in solution by the UV-Vis spectral changes and dynamic light scattering profiles. Incubation of MCF-7 cells with the conjugates revealed intense spotted intracellular fluorescence suggestive of accumulation in endosome/lysosome compartments, and no dark toxicity. Incubation with the tamoxifen-loaded conjugates revealed also practically no dark toxicity. Irradiation of cells incubated with empty conjugates at 640 nm and 4.18 J/cm2 light fluence caused >50 % cell viability reduction. Irradiation following incubation with tamoxifen-loaded conjugates resulted in even higher toxicity (74 %) indicating that the produced reactive oxygen species had triggered tamoxifen release in a photochemical internalization (PCI) mechanism. The chlorin-ß-cyclodextrin conjugates displayed less-lasting effects with time, compared to the corresponding porphyrin-ß-cyclodextrin conjugates, possibly due to lower tamoxifen loading of their aggregates and/or their less effective lodging in the cell compartments' membranes. The results suggest that further to favorable photophysical properties, other parameters are important for the in vitro effectiveness of the photodynamic systems.

A Monocarbonyl Curcuminoid Derivative Inhibits the Activity of Human Glutathione Transferase A4-4 and Chemosensitizes Glioblastoma Cells to Temozolomide.

Human glutathione transferase A4-4 (hGSTA4-4) displays high catalytic efficiency towards 4-hydroxyalkenals and other cytotoxic and mutagenic products of radical reactions and lipid peroxidation. Its role as a target for the chemosensitization of cancer cells has not been investigated so far. In this study, the inhibitory potency of twelve selected natural products and ten monocarbonyl curcumin derivatives against hGSTA4-4 was studied. Among natural products, ellagic acid turned out to be the strongest inhibitor with an IC50 value of 0.44 ± 0.01 µM. Kinetic analysis using glutathione (GSH) and 1-chloro-2,4-dinitrobenzene (CDNB) as variable substrates showed that ellagic acid behaved as a competitive inhibitor towards both GSH and CDNB, with Ki values of 0.39 ± 0.02 and 0.63 ± 0.03 µM, respectively. Among the curcumin derivatives studied, three proved to be the most potent inhibitors, in the order DM151 > DM101 > DM100, with IC50 values of 2.4 ± 0.1 µM, 12.7 ± 1.1 µΜ and 16.9 ± 0.4 µΜ, respectively. Further kinetic inhibition analysis of the most active derivative, DM151, demonstrated that this compound is a mixed inhibitor towards CDNB with inhibition constants of Ki = 4.1 ± 0.5 µM and Ki' = 0.536 ± 0.034 µM, while it is a competitive inhibitor towards GSH with a Ki = 0.98 ± 0.11 µM. Molecular docking studies were performed to interpret the differences in binding of ellagic acid and curcumin derivatives to hGSTA4-4. The in silico measured docking scores were consistent with the obtained experimental data. Hydrogen bonds appear to be the main contributors to the specific binding of monocarbonyl curcumin derivatives, while π-π stacking interactions play a key role in the enzyme-ellagic acid interaction. In vitro cytotoxicity assessment of the worst (DM148) and the best (DM151) inhibitors was performed against glioblastoma cell lines U-251 MG and U-87 MG. The results revealed that DM151 displays considerably higher cytotoxicity against both glioblastoma cell lines, while the glioblastoma cytotoxicity of DM148 was very limited. Furthermore, low and non-toxic doses of DM151 sensitized U-251 MG cells to the first-line glioblastoma chemotherapeutic temozolomide (TMZ), allowing us to propose for the first time that hGSTA4-4 inhibitors may be attractive therapeutic partners for TMZ to optimize its clinical effect in glioblastoma chemotherapy.

Oleuropein, an anti-oxidant polyphenol constituent of olive promotes α-secretase cleavage of the amyloid precursor protein (AßPP).

Over the past decade, intense focus has been dedicated on investigating processes involved in the proteolysis of amyloid precursor protein (AßPP) and ß-amyloid (Aß) peptide metabolism, as possible targets for Alzheimer's disease (AD) therapy. To this goal, considerable research has been targeted on potential therapeutic use of compounds promoting non-amyloidogenic processing of AßPP. One of these compounds, oleuropein, a polyphenol constituent of extra virgin olive oil exhibiting a wide range of pharmacological properties, was shown to interact non-covalently with Aß, an interaction that might be related to a potential protective role of oleuropein against Aß aggregation. In the present study, it was demonstrated that oleuropein treatment of HEK293 cells stably transfected with the isoform 695 of human AßPP (APP695) leads to markedly elevated levels of sAPPα and to significant reduction of Aß oligomers. These effects were associated with increased activity of matrix metalloproteinase 9 (MMP-9), whereas no significant alterations in the expression of secretases TACE, ADAM-10 or BACE-1 were observed. Similar results were obtained using the human neuroblastoma cell line SK-N-SH. The experimental data reveal an anti-amyloidogenic effect of oleuropein and suggest a possible protective role for oleuropein against AD, extending the spectrum of beneficial properties of this naturally occurring polyphenol.

Synthesis and characterization of fac-[M(CO)3(P)(OO)] and cis-trans-[M(CO)2(P)2(OO)] complexes (M = Re, (99m)Tc) with acetylacetone and curcumin as OO donor bidentate ligands.

The synthesis and characterization of neutral mixed ligand complexes fac-[M(CO)3(P)(OO)] and cis-trans-[M(CO)2(P)2(OO)] (M = Re, (99m)Tc), with deprotonated acetylacetone or curcumin as the OO donor bidentate ligands and a phosphine (triphenylphosphine or methyldiphenylphosphine) as the monodentate P ligand, is described. The complexes were synthesized through the corresponding fac-[M(CO)3(H2O)(OO)] (M = Re, (99m)Tc) intermediate aqua complex. In the presence of phosphine, replacement of the H2O molecule of the intermediate complex at room temperature generates the neutral tricarbonyl monophosphine fac-[Re(CO)3(P)(OO)] complex, while under reflux conditions further replacement of the trans to the phosphine carbonyl generates the new stable dicarbonyl bisphosphine complex cis-trans-[Re(CO)2(P)2(OO)]. The Re complexes were fully characterized by elemental analysis, spectroscopic methods, and X-ray crystallography showing a distorted octahedral geometry around Re. Both the monophosphine and the bisphosphine complexes of curcumin show selective binding to ß-amyloid plaques of Alzheimer's disease. At the (99m)Tc tracer level, the same type of complexes, fac-[(99m)Tc(CO)3(P)(OO)] and cis-trans-[(99m)Tc(CO)2(P)2(OO)], are formed introducing new donor combinations for (99m)Tc(I). Overall, ß-diketonate and phosphine constitute a versatile ligand combination for Re(I) and (99m)Tc(I), and the successful employment of the multipotent curcumin as ß-diketone provides a solid example of the pharmacological potential of this system.

Small anticancer drug release by light: Photochemical internalization of porphyrin-ß-cyclodextrin nanoparticles.

Aiming to engineer simple, neutral, strongly amphiphilic photoactive nanoparticles (NPs) to specifically target cancer cell lysosomes for drug transport and light-controlled release, new conjugates of ß-cyclodextrin with highly hydrophobic triphenylporphyrin bearing different alkyl chains, were synthesized. Although differently sized, all conjugates self-assemble into ~60 nm NPs in water and display similar photoactivity. The NPs target selectively the lysosomes of breast adenocarcinoma MCF-7 cells, embedding in vesicular membranes, as experiments with model liposomes indicate. Either empty or drug-loaded, the NPs lack dark toxicity for 48 h. They bind with differently structured anticancer drugs tamoxifen and gemcitabine as its N-adamantyl derivative. Red light irradiation of cells incubated with drug-loaded NPs results in major reduction of viability (>85 %) for 48 h displaying significant synergy of photo-chemotoxicity, as opposed to empty NPs, and to loaded non-irradiated NPs, in manifestation of photochemical internalization (PCI). Our approach expands the field of PCI into different small molecule chemotherapeutics.

Hydrophilic bis-MPA hyperbranched dendritic scaffolds as nanocarriers of a fully characterized flavonoid morin-Zn(II) complex for anticancer applications.

In this work the first crystallographically characterized complex of the bioactive flavonoid morin with the Zn(II) ion is presented along with its complete physico-chemical characterization. In view of the antioxidant activity of morin and its toxicity against respiratory tract cancers, the encapsulation of the complex in the hydrophilic bis(methylol)propionic acid hyperbranched dendritic scaffolds (bis-MPA HDSs) was effected. The produced nano-formulations were characterized with physico-chemical and electron microscopy techniques, and biologically evaluated for their antioxidant and anticancer activity against human A549 and H520 lung cancer cells, as well as healthy human MRC-5 lung fibroblasts. The obtained results demonstrate that encapsulation increases the solubility, and thus bioavailability, of the complex in physiological media and enhances anticancer action. They also highlight the importance of the non-toxic bis-MPA HDSs as nanocarriers of bioactive flavonoid metal complexes for anticancer therapeutic applications.

The Prophylactic and Multimodal Activity of Two Isatin Thiosemicarbazones against Alzheimer's Disease In Vitro.

Alzheimer's disease (AD) is a multifactorial disorder strongly involving the formation of amyloid-ß (Aß) oligomers, which subsequently aggregate into the disease characteristic insoluble amyloid plaques, in addition to oxidative stress, inflammation and increased acetylcholinesterase activity. Moreover, Aß oligomers interfere with the expression and activity of Glycogen synthase kinase-3 (GSK3) and Protein kinase B (PKB), also known as AKT. In the present study, the potential multimodal effect of two synthetic isatin thiosemicarbazones (ITSCs), which have been previously shown to prevent Aß aggregation was evaluated. Both compounds resulted in fully reversing the Aß-mediated toxicity in SK-NS-H cells treated with exogenous Aß peptides at various pre-incubation time points and at 1 µM. Cell survival was not recovered when compounds were applied after Aß cell treatment. The ITSCs were non-toxic against wild type and 5xFAD primary hippocampal cells. They reversed the inhibition of Akt and GSK-3ß phosphorylation in 5xFAD cells. Finally, they exhibited good antioxidant potential and moderate lipoxygenase and acetylcholinesterase inhibition activity. Overall, these results suggest that isatin thiosemicarbazone is a suitable scaffold for the development of multimodal anti-AD agents.

Unsymmetrical, monocarboxyalkyl meso-arylporphyrins in the photokilling of breast cancer cells using permethyl-ß-cyclodextrin as sequestrant and cell uptake modulator.

In the search for photosensitizers with chemical handles to facilitate their integration into complex drug delivery nanosystems, new, unsymmetrically substituted, water insoluble meso-tetraphenylporphyrin and meso-tetra(m-hydroxyphenyl)porphyrin derivatives bearing one carboxyalkyl side chain were synthesized. Permethyl-ß-cyclodextrin (pMßCD) was their ideal monomerizing host and highly efficient shuttle to transfer them into water. New assembly modes of the extremely stable (Kbinding > 1012 M-2) 2:1 complexes were identified. The complexes are photostable and do not disassemble in FBS-containing cell culture media for 24 h. Incubation of breast cancer MCF-7 cells with the complexes results in intense intracellular fluorescence, strongly enhanced in the endoplasmic reticulum (ER), high photokilling efficiency (~90%) and low dark toxicity. pMßCD stands out as a very capable molecular isolator of mono-carboxyalkyl-arylporphyrins that increases uptake and modulates their localization in the cells. The most efficient porphyrins are envisaged as suitable photosensitizers that can be linked to biocompatible drug carriers for photo- and chemo-therapy applications.