Maria Cristina Oliveira

Characterization of weld (Reseda luteola L.) and spurge flax (Daphne gnidium L.) by high-performance liquid chromatography–diode array detection–mass spectrometry in Arraiolos historical textiles

The natural dyes, and dye sources, in two seventeenth century Arraiolos carpets from the National Museum of Machado de Castro were analysed by high-performance liquid chromatography with UV-vis diode array detection (HPLC-DAD) and HPLC-mass spectrometry (LC-MS). Weld (Reseda luteola L.), indigo and spurge flax (Daphne gnidium L.) were found to be the dye sources, in agreement with original dyeing recipes collected during the nineteenth century. In order to fully characterize the plant sources, LC-MS conditions were optimized with plant extracts and the chromatographic separation and mass detection were enhanced. Extraction of the dyes, in the Arraiolos carpet samples, was performed using mild conditions that avoid glycoside decomposition. For the blues a dimethylformamide solution proved to be efficient for indigotin recovery. For all the other colours, an improved mild extraction method (with oxalic acid, methanol, acetone and water) was used, enabling to obtain the full dye source fingerprint, namely the flavonoid glycosides in the yellow dyes.

Radioiodinated sunitinib as a potential radiotracer for imaging angiogenesis-radiosynthesis and first radiopharmacological evaluation of 5-[125I]Iodo-sunitinib.

Sunitinib® (SU11248) is a highly potent tyrosine kinase inhibitor targeting vascular endothelial growth factor receptor (VEGFR). Radiolabeled inhibitors of RTKs might be useful tools for monitoring RTKs levels in tumour tissue giving valuable information for anti-angiogenic therapy. We report here the synthesis of a (125)I-labeled derivative of sunitinib® and its first radiopharmaceutical characterization. The non-radioactive reference compound 5-iodo-sunitinib 4 was prepared by Knoevenagel condensation of 5-iodo-oxindole with the corresponding substituted 5-formyl-1H-pyrrole. In a competition binding assay against VEGFR-2 a binding constant (K(d)) of 16 nM for 4 was found. The ability of 4 to inhibit tyrosine kinase activity was demonstrated on RTK expressing cells suggesting this radiotracer as a useful tool for monitoring VEGFR expression. 5-[(125)I]lodo-sunitinib, [(125)I]-4 was obtained via destannylation of the corresponding tributylstannyl precursor with [(125)I]NaI in the presence of H(2)O(2) in high radiochemical yield (>95%) and radiochemical purity (<98%) after HPLC purification. Determination of human plasma protein binding at time intervals of 0; 1; 2; 4 and 24h suggested a low non-specific binding of 5-10%. Preliminary biodistribution studies of [(125)I]-4 in healthy CD-1 mice showed a relatively high uptake in VEGFR-2 rich tissues like kidney and lung followed by rapid washout (9.6 and 9.7; 4.5 and 3.8% ID/g of kidney and lung at 1 and 4h, respectively).

Melanocortin-1 receptor-targeting with radiolabeled cyclic α-melanocyte-stimulating hormone analogs for melanoma imaging.

Melanoma is a type of skin cancer known for its high aggressiveness, early dissemination of metastases, and poor prognosis once metastasized. Thus, early diagnosis of melanoma is a key issue for increasing patient survival. The overexpression of melanocortin-1 receptors (MC1R) in isolated melanoma cells and melanoma tissues led to the radiolabeling of several linear and cyclic MC analogs for melanoma imaging or therapy. Cyclization of α-melanocyte stimulating hormone (α-MSH) peptides has been successfully used to improve binding affinity and in vivo stability of peptides. Herein, we describe the different peptide cyclization strategies recently reported for radiolabeled α-MSH analogs and discuss how such strategies affect MC1R binding affinity, pharmacokinetic profile, and MC1R-melanoma imaging. This review also highlights how the nature of the radiometal and labeling approach influence those properties. Among the cyclized α-MSH peptides reported, (99m)Tc/(111)In-labeled metal-cyclized and lactam bridge-cyclized peptides displayed the highest melanoma and lowest renal uptake values in B16/F1 melanoma-bearing mice and became the most promising tools to be further explored as potential melanoma imaging probes.

Influence of the bifunctional chelator on the pharmacokinetic properties of 99mTc(CO)3-labeled cyclic α-melanocyte stimulating hormone analog.

Aiming at the design of specific melanocortin-1 receptor (MC1R) targeted imaging probes, we report on the effect of different azolyl-ring substitution patterns (carboxylate at the 4-position and/or methyl groups at the 3,5 positions) of pyrazolyl-diamine bifunctional chelators (Pz(2)-Pz(4)) on the pharmacokinetic profile of the (99m)Tc(CO)3-labeled lactam bridge-cyclized α-melanocyte stimulating hormone derivative, βAlaNleCycMSH(hex). Three pyrazolyl-diamine-containing chelators were conjugated to βAlaNleCycMSHhex, with the resulting peptide conjugates displaying subnanomolar MC1R binding affinity. Biodistribution studies in B16F1 melanoma-bearing mice show that all radiopeptides present a good melanoma uptake. The introduction of a carboxylate group in the azolyl-ring leads to a remarkable reduction of the kidney (>89%) and liver (>91%) accumulation for (99m)Tc(CO)3-Pz(3)-βAlaNleCycMSH(hex) and (99m)Tc(CO)3-Pz(4)-βAlaNleCycMSH(hex) when compared to the radiopeptide (99m)Tc(CO)3-Pz(1)-βAlaNleCycMSH(hex), where that group is absent. The good tumor uptake and favorable tumor-to-nontarget-organs ratios of (99m)Tc(CO)3-Pz(3)-βAlaNleCycMSH(hex) and (99m)Tc(CO)3-Pz(4)-βAlaNleCycMSH(hex) highlights the potential of both compounds as melanoma imaging agents.

Fatty Acid Profile of Cheese from Dairy Goats Fed a Diet Enriched with Castor, Sesame and Faveleira Vegetable Oils

The addition of vegetable oils to the diets of dairy goats is an alternative to supplemental feeding during the dry period and improves the lipid profile of milk and by-products. Cheeses were produced using milk from cross bred goats (Saanen × Alpina) fed diets enriched with 4% vegetable oil (faveleira, sesame or castor), the fatty acid profile of cheeses was studied. Supplementation with vegetable oils did not increase the total fat percentage of the cheese (p ≥ 0.05) but did increase the percentage of CLA isomers, long-chain fatty acids (LCFA) and polyunsaturated fatty acids (PUFA); in addition, the index of desirable fatty acids (DFA - expressed as the sum of unsaturated fatty acids plus stearic acid) was increased for cheese made from milk from goats fed sesame or faveleira oil. Cheeses may have had increased percentages of cis-9,trans-11-CLA due to the supplementation of animal diets with vegetable oils rich in C18:2, such as faveleira and sesame oils. The fatty acid profile of goat cheese did not change significantly in response to the use of castor oil. Thus, the addition of sesame and faveleira oils to goat diets positively altered the fatty acid profile, which improved the nutritional characteristics of the fat present in goat cheese.

Radiohalogenated 4-anilinoquinazoline-based EGFR-TK inhibitors as potential cancer imaging agents.

INTRODUCTION The overexpression of epidermal growth factor receptor (EGFR) in tumors underlines the recent interest in EGFR as attractive target for the development of new cancer imaging agents. EGFR-tyrosine kinase inhibitors (EGFR-TKIs) based on the anilinoquinazoline scaffold have been explored as potential probes for EGFR imaging. However, up to now, no optimal radiotracer is available. Herein, we report the synthesis and biological evaluation of three novel halogenated 6-substituted 4-anilinoquinazoline based EGFR-TKIs. Radiosynthesis ((125)I and (18)F) of the corresponding analogues was also performed. METHODS 6a, 6b and 8 were obtained by reaction of 6-amino-4-anilinoquinazoline (5) with 3-/4-iodobenzoyl and 4-fluorobenzoyl chlorides. Inhibition of EGFR autophosphorylation and A431 cellular proliferation were assessed by Western blot and MTT assays. (125)I-anilinoquinazolines [(125)I]6a/b were prepared via destannylation of the corresponding tributylstannyl precursors with [(125)I]NaI. Cellular uptake studies were conducted in A431 cells. Optimization of the radiosynthesis of the (18)F-anilinoquinazoline [(18)F]8 was attempted by nucleophilic substitution of the trimethylammonium- and nitro-6-substituted 4-anilinoquinazoline precursors. RESULTS 6a, 6b and 8 were synthesized in high chemical yield. All of them are inhibitors of EGFR autophosphorylation (0.1<IC(50)<1 μM) and A431 cell proliferation (IC(50)<3.5 μM). [(125)I]6a/b, obtained in high radiochemical purity and specific activity, were highly taken up by A431 cells. Biodistribution profile in mice indicated fast blood clearance and hepatobiliary excretion. Despite all attempts, [(18)F]8 was only formed in 4% yield, hampering further biological evaluation. CONCLUSIONS This study suggests that these quinazoline derivatives can act as EGFR-TKI, warranting further modifications in the chemical structure in order to be explored as potential molecular imaging agents for single photon emission computerized tomography and positron emission tomography.

Novel estradiol based metal complexes of Tc-99m

Aiming to contribute to the design of technetium imaging agents for estrogen receptor (ER) positive breast tumors, we have synthesized and evaluated the novel organometallic estradiol complexes (fac-[M(CO)(3)(κ(3)-10)](+) and fac-[M(CO)(3)(κ(3)-12) M=Re/(99m)Tc) using two different bifunctional tridentate ligands (4 and 8). The rhenium complexes (13 and 14) were fully characterized by IR, (1)H NMR, (13)C NMR, mass spectrometry and elemental analyses. The (99m)Tc complexes (15 and 16) were obtained with high radiochemical purity and exhibited high in vitro radiochemical stability. To get a first insight into the relevance of these complexes for targeting ER positive tumors, ER binding affinity assays and cellular internalization studies in an ER expressing cell line, MCF-7, have also been performed suggesting a non ER mediated uptake.

Steroid receptor ligands for breast cancer targeting: an insight into their potential role as PET imaging agents.

The design and development of radiolabelled steroid derivatives has been an important area of research due to their wellknown value in breast cancer targeting. The estrogen receptor (ER) and progesterone receptor (PR) are important biomarkers in the diagnosis, prognosis and follow-up of the therapeutic response of breast tumours. Thus, several radioligands based on estrogens and progestins have been proposed for targeted functional ER imaging. The aim of this review is to survey and analyze the developments in this field, which have led to the design of a number of PET steroid-based imaging agents, a few of which seem to be promising as radiopharmaceuticals for detection of ER-positive breast tumours.

Theoretical spectroscopic studies and identification of metal-citrate (Cd and Pb) complexes by ESI-MS in aqueous solution

The combined use of ESI-MS, FTIR-ATR and theoretical calculations for the determination of metal-citrate (metal=Cd and Pb) structures are reported. Mass spectrometry allowed to determine the stoichiometry 1:1 and 2:1 of the complexes, corroborating the theoretical calculations. The species found in the ratio 2:1 had their molecular structures readjusted, since the deprotonation of citric acid differed from what was simulated. The calculations of thermodynamic stability (ΔH(0)(aq.)) for the complexes obtained by B3LYP/LANL2DZ were more exoenergetic than those found by PM6. However, for both methods, the stability of the complexes follows a trend, that is, the lowest-energy isomers in PM6 are also the most stable in B3LYP/LANL2DZ. The infrared analysis suggested that carboxyl groups are complexation sites and hydrogen bonds can help in the stability of the complexes. The vibrational frequencies in B3LYP/LANL2DZ had a good correlation with the experimental infrared results.

Estrogen Receptor Ligands for Targeting Breast Tumours: A Brief Outlook on Radioiodination Strategies

The design and development of radiolabelled estradiol derivatives has been an important area of research due to their recognized value in breast cancer management. The estrogen receptor (ER) is a relevant biomarker in the diagnosis, prognosis and prediction of the therapeutic response in estrogen receptor positive breast tumours. Hence, many radioligands based on estradiol derivatives have been proposed for targeted functional ER imaging. The main focus of this review is to survey the current knowledge on estradiol-based radioiodinated receptor ligands synthesis for breast tumour functional imaging. The main preclinical and clinical achievements in the field will also be briefly presented to make the manuscript more comprehensive.