Browsing by Author "Abreu, Rui M.V."
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- 1-aryl-3-(4-(7-methylthieno[3,2-d]pyrimidin-4-yloxy)phenyl)ureas: synthesis and molecular modelling studies using VEGFR-2Publication . Soares, Pedro; Froufe, Hugo J.C.; Abreu, Rui M.V.; Ferreira, Isabel C.F.R.; Borges, Fernanda; Queiroz, Maria João R.P.The development of anticancer drugs inhibiting angiogenesis has been an area of extensive research in the past decade. Angiogenesis is a requirement for tumor growth and metastasis and occurs through several signalling pathways. One key pathway that initiates proliferation and migration of endothelial cells is signalling through the vascular endothelial growth factor receptor-2 (VEGFR-2).1 Therefore, small molecules that block this signalling pathway through inhibition of VEGFR-2 tyrosine kinase activity could potentially inhibit angiogenesis and tumor growth. Recently works describing thienopyrimidine2 and thienopyrimidine 1,3-diarylureas3 as VEGFR-2 inhibitors have emerged in the literature. Here we present the synthesis of new 1-aryl-3-(4-(7-methylthieno[3,2-d]pyrimidin-4-yloxy)phenyl)ureas 2 in high yields by reaction of 4-[(7-methylthieno[3,2-d]pyridin-4-yl)oxy]aniline 1 with arylisocyanates. The former was prepared by regioselective nucleophilic substitution of 4-chloro-7-methylthieno[3,2-d]pyrimidine with 4-aminophenol
- 1-Aryl-3-[ 4-( thieno[3,2-d]pyrimidin-4-yloxy )phenyl]ureas as VEG FR2 inhibitors: synthesis, docking enzymatic and cellular assaysPublication . Peixoto, Daniela; Calhelha, Ricardo C.; Soares, Pedro; Abreu, Rui M.V.; Froufe, Hugo J.C.; Ferreira, Isabel C.F.R.; Costa, Raquel; Soares, Raquel; Queiroz, Maria João R.P.A number of thienopyrimidines derivatives have shown potent VEGFR2 (Vascular Endothelium Growth Factor Receptor2) tyrosine kinase inhibition activity.[ll VEGF is a sun·ogate marker of angiogenesis that activates VEGFR2 in endothelial cells. Here we present the synthesis of new 1-aryl-3-[ 4-(thieno[3,2-d]pyrimidin-4- yloxy)phenyl]ureas from the aminodi(hetero)arylether 1, also prepared by us, which was reacted with arylisocyanates to give the corresponding 1,3-diarylureas 2a-c.
- 1-Aryl-3-[4-(thieno[3,2-d]pyrimidin-4-yloxy)phenyl]ureas as potential inhibitors of VEGFR-2: synthesis and molecular modelling studiesPublication . Soares, Pedro; Froufe, Hugo J.C.; Abreu, Rui M.V.; Ferreira, Isabel C.F.R.; Borges, Fernanda; Queiroz, Maria João R.P.Angiogenesis is a requirement for tumor growth and metastasis and occurs through several signalling pathways. One key pathway that initiates proliferation and migration of endothelial cells is signalling through the vascular endothelial growth factor receptor-2 (VEGFR-2).1 Therefore, small molecules that block this signalling pathway through inhibition of the VEGFR tyrosine kinase activity could potentially inhibit angiogenesis and tumour growth. Recently works describing thienopyrimidines2 and thienopyridine ureas3 as inhibitors of VEGFR-2 have appeared in the literature. Here we present the synthesis of new 1,3-diarylureas 2 starting by regioselective nucleophilic substitution of the 4-chlorothieno[3,2-d]pyrimidine with 4-aminophenol to obtain 4-(thieno[3,2-d]pyridin-4-yloxy)aniline 1 which reacts with different arylisocyanates
- 1-Aryl-3-[4-(thieno[3,2-d]pyrimidin-4-yloxy)phenyl]ureas as VEGFR-2 tyrosine kinase inhibitors: Synthesis, biological evaluation, and molecular modelling studiesPublication . Soares, Pedro; Costa, Raquel; Froufe, Hugo J.C.; Calhelha, Ricardo C.; Peixoto, Daniela; Abreu, Rui M.V.; Ferreira, Isabel C.F.R.; Soares, Raquel; Queiroz, Maria João R.P.The Vascular Endothelial Growth Factor Receptor-2 (VEGFR-2) is a tyrosine kinase receptor involved in the growth and differentiation of endothelial cells that is implicated in tumor-associated angiogenesis. In this study novel 1-aryl-3-[4-(thieno[3,2-d]pyrimidin-4-yloxy)phenyl]ureas were synthesized and evaluated for the VEGFR-2 tyrosine kinase inhibition. Three of these compounds showed good VEGFR-2 inhibition presenting low IC50 values (150-199 nM) in enzymatic assays. The latter promoted also significant inhibition of cell proliferation at low concentrations (0.5-1 µM), not affecting cell viability, of VEGF-stimulated Human Umbilical Vein Endothelial Cells (HUVECs) using the BrdU assay. The determination of the total and phosphorylated (active) VEGFR-2 was performed by western-blot, and it was possible to conclude that the compounds significantly inhibit the phosphorylation of the receptor at 1 µM pointing to their antiproliferative mechanism of action in HUVECs. The molecular rationale for inhibiting the tyrosine kinase domain of VEGFR-2 was also done and discussed using molecular docking studies.
- 1-Aryl-3-[4-(thieno[3,2-d]pyrimidin-4-yloxy)phenyl]ureas as VEGFR2 tyrosine kinase inhibitors: synthesis, docking studies, enzymatic and cellular assaysPublication . Queiroz, Maria João R.P.; Peixoto, Daniela; Soares, Pedro; Abreu, Rui M.V.; Froufe, Hugo J.C.; Calhelha, Ricardo C.; Ferreira, Isabel C.F.R.; Costa, Raquel; Soares, RaquelA number of thienopyrimidines derivatives have shown potent vascular endothelial growth factor receptor 2 (VEGFR2) inhibition activity. Here, we present the synthesis of new 1-aryl-3-[4-(thieno[3,2-d) pyrimidin-4-yloxy)phenyl]ureas by promoting t he regioselective attack of the hydroxy group of the 4-aminophenol in the chlorine nucleophilic displacement on two 4-chlorinated thieno[3,2-d]pyrimidines, obtaining compounds la and 1b which were reacted with arylisocyanates to give t he corresponding 1,3-diarylureas 2a-f (see scheme). These compounds were evaluated for inhibition of VEGFR2 tyrosine kinase activity using enzymatic assays, and 2a- c showed good inhibition ability with IC50 values in the range of hundreds of nanomolar. The rationale for the inhibition activity is also discussed using docking. To examine the activity of 2a- c in endothelial cells, human umbilical vein endothelial cells (HUVECs) were cultured in the presence or absence of each compound in different concentrations. A decrease in the proliferation of HUVECs was observed by the incorporation of BrdU quantified by ELISA assay. Given the established role of VEGFR2 in proliferation and migration of endothelial cells, these molecules are promising antiangiogenic agents that can be used for therapeutic purposes in pathological conditions where angiogenesis is exacerbated, such as cancer.
- A amilóide A no leite é útil no diagnóstico de mastites subclínicas em caprinos?Publication . Quintas, Helder; Sequeira, Ana; Coelho, Ana Cláudia; Abreu, Rui M.V.; Alegria, Nuno; Mendonça, ÁlvaroAs mastites são responsáveis por perdas económicas extremamente elevadas nos ruminantes leiteiros e, deste ponto de vista, a forma subclínica é a mais preocupante. Assim, o diagnóstico realizado de forma precoce e eficaz torna-se numa das medidas mais importantes no controlo desta doença. A mensuração das proteínas da fase aguda da inflamação (PFA), para deteção de mastites, tem sido alvo de várias pesquisas em bovinos e diversos estudos comprovam um incremento na produção de haptoglobina e Amilóide A (AA) séricas em vacas com mastites clínicas e subclínicas. Apesar da reduzida literatura disponível em pequenos ruminantes, a Amilóide A sérica está indicada como a PFA mais sensível em ovinos. Recentemente foi descoberta uma isoforma específica de AA no leite de bovinos e ovinos, denominada por Amilóide A do leite. Esta é produzida pelas células epiteliais e aumenta precocemente em situações de mastite. No presente estudo foi avaliada a utilidade do doseamento de Amilóide A do leite, através de um ensaio imunoenzimático, no diagnóstico de mastites em caprinos. Semanalmente, durante seis semanas consecutivas, foram recolhidas amostras de leite de 12 cabras de raça Serrana, de manhã, antes de se proceder à ordenha. No total foram recolhidas 144 amostras de metades mamárias. Com base nos resultados microbiológicos e da contagem de células somáticas, as metades mamárias foram divididas em 4 grupos: saudáveis, com mastites subclínicas e com mastites subclínicas duvidosas – latentes e inespecíficas. Os resultados das concentrações de Amilóide A do leite mostraram diferenças significativas (P<0,05) entre as metades mamárias saudáveis e as com mastites subclínicas e entre o grupo com mastites subclínicas inespecíficas e os restantes grupos. O doseamento da Amilóide A do leite revelou-se uma técnica útil na diferenciação das metades mamárias saudáveis das com mastites subclínicas. Porém, esta distinção foi condicionada pelas mastites subclínicas inespecíficas.
- A amilóide A no leite é útil no diagnóstico de mastites subclínicas em caprinos?Publication . Quintas, Helder; Sequeira, Ana; Coelho, Ana Cláudia; Abreu, Rui M.V.; Alegria, Nuno; Mendonça, ÁlvaroA mensuração das proteínas da fase aguda da inflamação (PFA), para deteção de mastites, tem sido alvo de vários estudos em bovinos e diversos estudos comprovam um incremento na produção de haptoglobina e Amilóide A (AA) séricas em vacas com mastites clínicas e subclínicas. Apesar da reduzida literatura disponível em pequenos ruminantes, a Amilóide A sérica está indicada como a PFA mais sensível em ovinos. Recentemente foi descoberta uma isoforma específica de AA no leite de bovinos e ovinos, denominada por Amilóide A do leite. Esta é produzida pelas células epiteliais e aumenta de forma precoce em situações de mastite. Com este estudo pretendeu-se avaliar a utilidade do doseamento de Amilóide A do leite no diagnóstico de mastites em caprinos.
- Aminodi(hetero)arylamines in the thieno[3,2-b]pyridine series: synthesis, effects in human tumor cells growth, cell cycle analysis, apoptosis and evaluation of toxicity using non-tumor cellsPublication . Calhelha, Ricardo C.; Ferreira, Isabel C.F.R.; Peixoto, Daniela; Abreu, Rui M.V.; Vale-Silva, Luís A.; Pinto, Eugénia; Lima, Raquel T.; Alvelos, M. Inês; Vasconcelos, M. Helena; Queiroz, Maria João R.P.Three aminodi(hetero)arylamines were prepared via a palladium-catalyzed C-N Buchwald-Hartwig coupling of methyl 3-aminothieno[3,2-b]pyridine-2-carboxylate with different bromonitrobenzenes, followed by reduction of the nitro groups of the coupling products to the corresponding amino compounds. The aminodi(hetero)arylamines thus obtained were evaluated for their growth inhibitory effect on four human tumor cell lines MCF-7 (breast adenocarcinoma), A375-C5 (melanoma), NCI-H460 (non-small cell lung cancer) and HepG2 (hepatocellular carcinoma). The toxicity to non-tumor cells was also evaluated using a porcine liver primary cell culture (PLP1), established by us. The aminodi(hetero)arylamine with the NH2 group in the ortho position and an OMe group in the para position to the NH of the di(hetero)arylamine, is the most promising compound giving the lowest GI50 values (1.30–1.63 μM) in all the tested human tumor cell lines, presenting no toxicity to PLP1 at those concentrations. The effect of this compound on the cell cycle and induction of apoptosis was analyzed in the NCI-H460 cell line. It was observed that it altered the cell cycle profile causing a decrease in the percentage of cells in the G0/G1 phase and an increase of the apoptosis levels.
- Anti-hepatocellular carcinoma activity using human HepG2 cells and hepatotoxicity of 6-substituted methyl 3-aminothieno[3,2-b]pyridine-2- carboxylate derivatives: In vitro evaluation, cell cycle analysis and QSAR studiesPublication . Abreu, Rui M.V.; Ferreira, Isabel C.F.R.; Calhelha, Ricardo C.; Lima, Raquel T.; Vasconcelos, M. Helena; Adega, Filomena; Chaves, Raquel; Queiroz, Maria João R.P.Hepatocellular carcinoma (HCC) is a highly complex cancer, resistant to commonly used treatments and new therapeutic agents are urgently needed. A total of thirty-two thieno[3,2-b]pyridine derivatives of two series: methyl 3-amino- -(hetero)arylthieno[3,2-b]pyridine-2-carboxylates (1ae1t) and methyl 3-amino-6-[(hetero)arylethynyl]thieno[3,2-b]pyridine-2-carboxylates (2ae2n), previously prepared by some of us, were evaluated as new potential anti-HCC agents by studying their in vitro cell growth inhibition on human HepG2 cells and hepatotoxicity using a porcine liver primary cell culture (PLP1). The presence of amino groups linked to a benzene moiety emerges as the key element for the anti-HCC activity. The methyl 3-amino-6-[(3-aminophenyl)ethynyl]thieno[3,2-b]pyridine-2-carboxylate (2f) is the most potent compound presenting GI50 values on HepG2 cells of 1.2 mM compared to 2.9 mM of the positive control ellipticine, with no observed hepatotoxicity (PLP1 GI50 > 125 mM against 3.3 mM of ellipticine). Moreover this compound changes the cell cycle profile of the HepG2 cells, causing a decrease in the % of cells in the S phase and a cell cycle arrest in the G2/M phase. QSAR studies were also performed and the correlations obtained using molecular and 1D descriptors revealed the importance of the presence of amino groups and hydrogen bond donors for anti-HCC activity, and hydrogen bond acceptors for hepatotoxicity. The best correlations were obtained with 3D descriptors belonging to different subcategories for anti-HCC activity and hepatotoxicity, respectively. These results point to different molecular mechanisms of action of the compounds in anti-HCC activity and hepatotoxicity. This work presents some promising thieno[3,2-b]pyridine derivatives for potential use in the therapy of HCC. These compounds can also be used as scaffolds for further synthesis of more potent analogs.
- Anti-inflammatory activity of mushrooms extracts, identified phenolic acids and their possible metabolitesPublication . Taofiq, Oludemi; Calhelha, Ricardo C.; Heleno, Sandrina A.; Barros, Lillian; Martins, Anabela; Abreu, Rui M.V.; Queiroz, Maria João R.P.; Ferreira, Isabel C.F.R.Mushrooms are rich sources of many bioactive compounds, such as phenolic acids, that play an important role in the organism, acting as antioxidants, antitumors, antimicrobials, immunomodulators, among others. However, their anti-inflammatory activity has not been deeply studied. In the present study, the ethanolic extracts of fourteen edible mushroom species were firstly characterized in terms of phenolic acids and related compounds by HPLC-PDA, followed by the study of the anti-inflammatory activity of those extracts and corresponding identified compounds, by using LPS (lipopolysaccharide) activated RAW 264.7 macrophages and measuring the inhibition in NO production. Furthermore, methylated and glucuronated derivatives of the identified compounds (p-hydroxybenzoic, p-coumaric and cinnamic acids) were synthesised and evaluated for the same bioactivity to understand the contribution of these compounds for the overall activity of the extracts, and to establish structure-activity relationships. Pleurotus ostreatus, Macrolepiota procera, Boletus impolitus and Agaricus bisporus revealed the strongest anti-inflammatory potential, presenting also the highest concentration in cinnamic acid, which was also the individual compound displaying the highest activity. The derivative compounds of p-coumaric acid revealed the strongest properties, especially the compound CoA-M1 (presenting an ester instead of the carboxylic group), that exhibited a very similar activity to the one showed by dexamethasone, used as anti-inflammatory standard. On the contrary, p-hydroxybenzoic acid derivatives revealed the lowest activity. Overall, the conjugation reactions change the chemical structure of phenolic acids and may increase or decrease their activity; nevertheless, the glucuronated and methylated derivatives of the studied compounds are still displaying anti-inflammatory activity.