Browsing by Author "Matias, Mariana Ruivo"
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- Development of new antiepileptic drug candidates: a set of lamotrigine-related compoundsPublication . Matias, Mariana Ruivo; Alves, Gilberto Lourenço; Silvestre, Samuel Martins; Ferreira, Amílcar Celta Falcão RamosEpilepsy is one of the most common, chronic and serious neurological disorder, affecting million people worldwide. This brain disorder is characterised by recurrent spontaneous seizures, which have a considerable impact in the patients’ quality of life. The pharmacological therapy has been, and is likely to remain, the mainstay of treatment for this disorder. Although a large number of new antiepileptic drugs (AEDs) has been introduced into the market in the last years, about 30-40% of epileptic patients are still inadequately controlled by standard drug therapy. For this reason, it continues to be important to develop new and improved chemical entities through which epilepsy could be effectively controlled. In this context, the main objective of the present work was to discover new lead compounds with anticonvulsant properties for further development as AEDs. To achieve this goal, fifty dihydropyrimidin(thi)ones [DHPM(t)s] were synthesized through the Biginelli reaction, which consists in a one-pot cyclocondensation reaction among an aldehyde, a β-ketoester/acetylacetone and urea or thiourea. The products were purified and characterised by infrared and 1H- and 13C-nuclear magnetic resonance spectroscopy. High resolution mass spectrum was also obtained for the novel compounds. Afterwards, the anticonvulsant activity of the compounds was evaluated against electrically [maximal electroshock seizure (MES) test] and chemically [subcutaneous pentylenetetrazole (scPTZ) test] induced seizures in rodent models. The initial anticonvulsant screening was performed in CD-1 mice (n = 4/group) at 30 min and 4 h after the intraperitoneal administration of 30, 100 and 300 mg/kg of each compound. The investigated compounds were also evaluated in mice for neuromotor impairment (as a surrogate of minimal neurological deficit) on the rotarod performance test. Then, selected compounds previously identified as anticonvulsants in mice at the minimum dose tested were further assessed in Wistar rats (n = 4/group) at 30 min, 2 h and 4 h after the oral administration of 30 mg/kg. Additionally, the fifty DHPM(t)s were evaluated for their in vitro cytotoxicity in rat mesencephalic dopaminergic (N27), human hepatocellular carcinoma (HepaRG), human colorectal adenocarcinoma (Caco-2) and normal human dermal fibroblasts (NHDF) cell lines, through the well-established 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay at the concentration of 30 μM. Moreover, as the efficacy of a molecule is strongly dependent on its pharmacokinetics, several kinetic properties were also investigated in in vitro and in silico models. Thus, all compounds were subjected to a set of in vitro screening assays performed on a cell line overexpressing the drug efflux transporter P-glycoprotein (MDCK-MDR1 cells) and on two models of parallel artificial membrane permeability assay (PAMPA) preditive of the apparent permeability (Papp) through intestinal membrane (intestinal PAMPA model) and blood-brain barrier (PAMPA-BBB model). Lastly, several physicochemical properties of the compounds were also calculated in silico and a set of pharmacokinetic and toxicity properties were estimated employing the new computational tool, pkCSM. The target molecules that were synthesized were mainly selected based on the structure of clinically relevant AEDs, in particular the structure of lamotrigine, aiming to discover new candidates for the development of improved AEDs. The majority of the chemical reactions occurred fastly and the products were obtained in good yields. The synthetic procedure used was also extended using additional specific reagents, being the respective products, which are new to the best of our knowledge, successfully synthesized. Due to practical considerations, only forty-two compounds (twenty-eight urea derivatives and fourteen thiourea derivatives) proceeded to in vivo experiments. The results of the initial pharmacological screening in mice revealed anticonvulsant protection in the MES model for twenty-four compounds showed anticonvulsant protection in the MES model, being nine of them active at the lowest dose tested (30 mg/kg). Structurally, the most promising compounds present smaller chains at the C5 of the dihydropyrimidine ring and an unsubstituted phenyl or a para-tolyl ring at the C4. In addition, the thiourea analogues also presented slightly increased anticonvulsant activity comparing with the corresponding urea analogues. The results of the minimal neuromotor impairment obtained through the rotarod assay showed that approximately 52% of the compounds are less toxic than lamotrigine, carbamazepine and phenytoin. Compounds MM 17, MM 19 and MM 83 also protected against MES-induced seizures in 50-75% of rats after the oral administration of 30 mg/kg. Furthermore, the most active compounds did not show notable cytotoxicity in in vitro experiments conducted in the several cell lines (relative cell proliferation higher than 50% at 30 μM), which can be relevant due to the fact that the toxicity is a common problem of the available AEDs. The data obtained showed that 82% of the investigated compounds are expected to have good intestinal permeability (Papp > 1.1×10-6 cm/s), and 66% of which good brain penetration (Papp > 2.0×10-6 cm/s), which can suggest a high passive transcellular permeability. In both cases, thiourea derivatives presented higher permeability values than the respective urea analogues, which can be associated with their higher lipophilicity. This finding can explain, at least in part, the higher activity of the thiourea derivatives in the anticonvulsant screening after both intraperitoneal and oral administrations. In addition, 44% of the compounds did not significantly modulate (inhibit or induce) P-glycoprotein at 10 and 50 μM. This is an interesting finding since P-glycoprotein is physiologically expressed in several tissues and organs relevant from a pharmacokinetics perspetive. Finally, in silico studies indicated that all compounds respect the Lipinski’s rule-of-five, suggesting that they possess favourable properties that fulfil the druglikeness criteria. The pkCSM in silico tool also estimated that the DHPM(t)s have good human intestinal absorption (67.73-93.91%) and an apparent volume of distribution at the steady-state in the same range of values of the AEDs. The in silico predictions also suggested a low plasma protein binding percentage for the target compounds, which is considered to be therapeutically favourable, minimizing the risk of drug interactions. These results corroborate those obtained with the intestinal PAMPA assay that showed that probably none of the tested compounds have a binding to plasma proteins higher than 90% (Papp ≤ 1.0×10−5 cm/s). The thiourea derivatives were also predicted as compounds that permeate better through biological barriers (e.g., Caco-2 cell monolayers and blood-brain barrier), similarly to the observed in the experimental PAMPA assays. However, the prediction model suggested that 14% of the urea derivatives have tendency for cytochrome P450 inhibition versus 36% of the thiourea derivatives. On the other hand, concerns on the disruption of normal liver function were predicted for half of the compounds. Overall, the set of studies carried out provide new information about the anticonvulsant activity of this class of heterocycles, along with pharmacokinetic and toxicity data. More than half of the investigated molecules showed anticonvulsant protection against electrically-induced seizures (MES model), confirming the interest of the pharmacophoric model for the design of new anticonvulsant agents. The data gathered here allowed to identify important structural features of this attractive scaffold that can be responsible for the anticonvulsant activity, which should be maintained or better explored in order to produce more active analogues in further hit-to-lead optimization. However, the results presented in this thesis are just the “tip of the iceberg” in the discovery and development of the DHPM(t)s as potential AEDs.
- Reconciliação da medicação no pós-operatório da cirurgia ortopédicaPublication . Matias, Mariana Ruivo; Aperta, Jorge Manuel GonçalvesA reconciliação da medicação é um processo sistemático e contínuo que visa promover a segurança do doente e aumentar a qualidade de vida deste através da redução de erros de medicação e eventos adversos. O objetivo deste estudo consistiu na realização da reconciliação da medicação no pós-operatório da cirurgia ortopédica, determinando quais as classes de fármacos envolvidas nas discrepâncias e identificando as discrepâncias preponderantes. Este estudo foi conduzido no serviço de ortopedia do Centro Hospitalar Médio Tejo – Unidade de Abrantes, entre 3 de abril e 20 de maio de 2011. As listas de medicação domiciliares foram obtidas através da aplicação de inquéritos aos doentes e da consulta dos registos de enfermagem. A medicação prescrita foi obtida a partir dos processos clínicos dos doentes. As listas foram comparadas. Identificaram-se as discrepâncias (qualquer variação não intencional entre os medicamentos utilizados antes da admissão e os medicamentos prescritos após a intervenção cirúrgica) e discriminaram-se os fármacos. O processamento dos dados foi realizado utilizando estatística descritiva (frequências, percentagens, médias, desvios padrão) e estatística indutiva (qui-quadrado, odds ratio). Dos 36 doentes (média de idade 74 anos, 30 mulheres e 6 homens) incluídos no estudo, 23 (64,0%) apresentaram ≥1 discrepâncias. A medicação cardiovascular foi a que registou maior envolvimento nas discrepâncias (32,6%) seguida dos medicamentos que atuam no sistema nervoso central (30,2%). Dos fármacos envolvidos nas discrepâncias, 14,0% foram considerados medicação de alto risco. As discrepâncias mais frequentes foram a omissão (50,0%) e a interação com potencial para causar dano ao doente (19,4%). Verificou-se, neste estudo, a existência de uma correlação entre o número de medicamentos domiciliares e o número de discrepâncias identificadas (OR=8,5; IC95%, 1,84-39,23). O processo da reconciliação da medicação mostrou ter um grande potencial para identificar discrepâncias de medicação no pós-operatório de doentes hospitalizados, o que evidencia a importância e necessidade de criação e implementação deste processo.