RESUMO
A new dimeric naphthylisoquinoline alkaloid, jozibrevine D (4e), was isolated from the Central-African liana Ancistrocladus ileboensis. It is a Dioncophyllaceae-type metabolite, being R-configured at C-3 and lacking an oxygen function at C-6 in both isoquinoline moieties. The two identical monomers of jozibrevine D are symmetrically linked via the sterically constrained 3',3''-positions of the naphthalene units so that the central biaryl linkage is rotationally hindered and the alkaloid is, thus, C2-symmetric. With the two outer biaryl bonds being chiral, too, 4e possesses three consecutive stereogenic axes. The absolute stereostructure of the new compound was assigned by 1D and 2D NMR, ruthenium-mediated oxidative degradation, and electronic circular dichroism (ECD) spectroscopy. Jozibrevine D (4e) is the fifth discovered isomer in a series of six possible natural atropo-diastereomeric dimers. It shows potent, and selective, antiprotozoal activity against P. falciparum (IC50 = 0.14 µM), and it also exhibits good cytotoxic activities against drug-sensitive acute lymphoblastic CCRF-CEM leukemia cells (IC50 = 11.47 µM) and their multidrug-resistant CEM/ADR5000 subline (IC50 = 16.61 µM).
Assuntos
Alcaloides , Antimaláricos , Antineoplásicos , Antiprotozoários , Caryophyllales , Antiparasitários/farmacologia , Antimaláricos/química , Estrutura Molecular , Alcaloides/química , Antiprotozoários/farmacologia , Antiprotozoários/química , Caryophyllales/químicaRESUMO
The improvement of cancer chemotherapy remains a major challenge, and thus new drugs are urgently required to develop new treatment regimes. Curcumin, a polyphenolic antioxidant derived from the rhizome of turmeric (Curcuma longa L.), has undergone extensive preclinical investigations and, thereby, displayed remarkable efficacy in vitro and in vivo against cancer and other disorders. However, pharmacological limitations of curcumin stimulated the synthesis of numerous novel curcumin analogs, which need to be evaluated for their therapeutic potential. In the present study, we calculated the binding affinities of 50 curcumin derivatives to known cancer-related target proteins of curcumin, i.e., epidermal growth factor receptor (EGFR) and nuclear factor κB (NF-κB) by using a molecular docking approach. The binding energies for EGFR were in a range of −12.12 (±0.21) to −7.34 (±0.07) kcal/mol and those for NF-κB ranged from −12.97 (±0.47) to −6.24 (±0.06) kcal/mol, indicating similar binding affinities of the curcumin compounds for both target proteins. The predicted receptor-ligand binding constants for EGFR and curcumin derivatives were in a range of 0.00013 (±0.00006) to 3.45 (±0.10) µM and for NF-κB in a range of 0.0004 (±0.0003) to 10.05 (±4.03) µM, indicating that the receptor-ligand binding was more stable for EGFR than for NF-κB. Twenty out of 50 curcumin compounds showed binding energies to NF-κB smaller than −10 kcal/mol, while curcumin as a lead compound revealed free binding energies of >−10 kcal/mol. Comparable data were obtained for EGFR: 15 out of 50 curcumin compounds were bound to EGFR with free binding energies of <−10 kcal/mol, while the binding affinity of curcumin itself was >−10 kcal/mol. This indicates that the derivatization of curcumin may indeed be a promising strategy to improve targe specificity and to obtain more effective anticancer drug candidates. The in silico results have been exemplarily validated using microscale thermophoresis. The bioactivity has been further investigated by using resazurin cell viability assay, lactate dehydrogenase assay, flow cytometric measurement of reactive oxygen species, and annexin V/propidium iodide assay. In conclusion, molecular docking represents a valuable approach to facilitate and speed up the identification of novel targeted curcumin-based drugs to treat cancer.
Assuntos
Curcumina , Neoplasias , Curcumina/química , Receptores ErbB , Humanos , Proteínas I-kappa B , Ligantes , Simulação de Acoplamento Molecular , NF-kappa B/metabolismo , Neoplasias/tratamento farmacológicoRESUMO
Background Triptolide is an active natural product, which inhibits cell proliferation, induces cell apoptosis, suppresses tumor metastasis and improves the effect of other therapeutic treatments in several cancer cell lines by affecting multiple molecules and signaling pathways, such as caspases, heat-shock proteins, DNA damage and NF-ĸB. Purpose We investigated the effect of triptolide towards NF-ĸB and GATA1. Methods We used cell viability assay, compare and cluster analyses of microarray-based mRNA transcriptome-wide expression data, gene promoter binding motif analysis, molecular docking, Ingenuity pathway analysis, NF-ĸB reporter cell assay, and electrophoretic mobility shift assay (EMSA) of GATA1. Results Triptolide inhibited the growth of drug-sensitive (CCRF-CEM, U87.MG) and drug-resistant cell lines (CEM/ADR5000, U87.MGΔEGFR). Hierarchical cluster analysis showed six major clusters in dendrogram. The sensitive and resistant cell lines were statistically significant (p = 0.65 × 10-2) distributed. The binding motifs of NF-κB (Rel) and of GATA1 proteins were significantly enriched in regions of 25 kb upstream promoter of all genes. IPA showed the networks, biological functions, and canonical pathways influencing the activity of triptolide towards tumor cells. Interestingly, upstream analysis for the 40 genes identified by compare analysis revealed ZFPM1 (friend of GATA protein 1) as top transcription regulator. However, we did not observe any effect of triptolide to the binding of GATA1 in vitro. We confirmed that triptolide inhibited NF-κB activity, and it strongly bound to the pharmacophores of IκB kinase ß and NF-κB in silico. Conclusion Triptolide showed promising inhibitory effect toward NF-κB, making it a potential candidate for targeting NF-κB.
Assuntos
Diterpenos/farmacologia , Fator de Transcrição GATA1/efeitos dos fármacos , NF-kappa B/efeitos dos fármacos , Farmacologia em Rede/métodos , Fenantrenos/farmacologia , Ligação Proteica/efeitos dos fármacos , Fatores de Transcrição/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ensaio de Desvio de Mobilidade Eletroforética , Compostos de Epóxi/farmacologia , Humanos , Simulação de Acoplamento Molecular , RNA MensageiroRESUMO
Introduction Differentiation therapy is a promising strategy for cancer treatment. The translationally controlled tumor protein (TCTP) is an encouraging target in this context. By now, this field of research is still at its infancy, which motivated us to perform a large-scale screening for the identification of novel ligands of TCTP. We studied the binding mode and the effect of TCTP blockade on the cell cycle in different cancer cell lines. Methods Based on the ZINC-database, we performed virtual screening of 2,556,750 compounds to analyze the binding of small molecules to TCTP. The in silico results were confirmed by microscale thermophoresis. The effect of the new ligand molecules was investigated on cancer cell survival, flow cytometric cell cycle analysis and protein expression by Western blotting and co-immunoprecipitation in MOLT-4, MDA-MB-231, SK-OV-3 and MCF-7 cells. Results Large-scale virtual screening by PyRx combined with molecular docking by AutoDock4 revealed five candidate compounds. By microscale thermophoresis, ZINC10157406 (6-(4-fluorophenyl)-2-[(8-methoxy-4-methyl-2-quinazolinyl)amino]-4(3H)-pyrimidinone) was identified as TCTP ligand with a KD of 0.87 ± 0.38. ZINC10157406 revealed growth inhibitory effects and caused G0/G1 cell cycle arrest in MOLT-4, SK-OV-3 and MCF-7 cells. ZINC10157406 (2 × IC50) downregulated TCTP expression by 86.70 ± 0.44% and upregulated p53 expression by 177.60 ± 12.46%. We validated ZINC10157406 binding to the p53 interaction site of TCTP and replacing p53 by co-immunoprecipitation. Discussion ZINC10157406 was identified as potent ligand of TCTP by in silico and in vitro methods. The compound bound to TCTP with a considerably higher affinity compared to artesunate as known TCTP inhibitor. We were able to demonstrate the effect of TCTP blockade at the p53 binding site, i.e. expression of TCTP decreased, whereas p53 expression increased. This effect was accompanied by a dose-dependent decrease of CDK2, CDK4, CDK, cyclin D1 and cyclin D3 causing a G0/G1 cell cycle arrest in MOLT-4, SK-OV-3 and MCF-7 cells. Our findings are supposed to stimulate further research on TCTP-specific small molecules for differentiation therapy in oncology.
Assuntos
Antineoplásicos/farmacologia , Drogas em Investigação/farmacologia , Neoplasias/tratamento farmacológico , Proteína Tumoral 1 Controlada por Tradução/antagonistas & inibidores , Antineoplásicos/administração & dosagem , Artesunato/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Simulação por Computador , Bases de Dados de Produtos Farmacêuticos , Relação Dose-Resposta a Droga , Drogas em Investigação/administração & dosagem , Humanos , Ligantes , Simulação de Acoplamento Molecular , Neoplasias/patologia , Proteína Tumoral 1 Controlada por Tradução/metabolismoRESUMO
Melianthus major is a medicinal plant endemic to South Africa. Its leaf extract led to the isolation of five new bufadienolides, 2ß-acetoxy-3,5-di-O-acetylhellebrigenin (1), 2ß-acetoxy-3-O-acetylhellebrigenin (2), 2ß-acetoxy-14-deoxy-15ß,16ß-epoxymelianthugenin (4), 2ß-acetoxy-14-deoxy-15ß,16ß-epoxymelianthusigenin (5), and 2ß-hydroxymelianthusigenin (6), and four known analogues. The structures of the compounds were elucidated using NMR and HRESIMS data analyses. The relative configurations were defined by single-crystal X-ray crystallography and NOESY correlations. The isolated compounds exhibited strong cytotoxicity against MCF-7 breast cancer cells and sensitive CCRF-CEM and multidrug-resistant CEM/ADR5000 leukemia cells. Compound 1 showed the most potent activity, with IC50 values of 0.1 µM toward CCRF-CEM and CEM/ADR5000 and 0.3 µM toward MCF-7.
Assuntos
Antineoplásicos Fitogênicos/isolamento & purificação , Antineoplásicos Fitogênicos/farmacologia , Bufanolídeos/isolamento & purificação , Bufanolídeos/farmacologia , Magnoliopsida/química , Folhas de Planta/química , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Análise Espectral/métodosRESUMO
From the leaves of South African medicinal plant Melianthus comosus, four previously undescribed bufadienolides, 16ß-formyloxymelianthugenin (1), 2ß-acetoxymelianthusigenin (2), 2ß-hydroxy-3ß,5ß-di-O-acetylhellebrigenin (3), and 2ß-acetoxy-5ß-O-acetylhellebrigenin (4) were isolated together with two known bufadienolides. The structural elucidation of the compounds was based on 1D and 2D NMR spectroscopy, high-resolution mass spectrometry, and other spectroscopic methods. The relative configurations were determined by single-crystal X-ray crystallography analysis and NOESY correlations. The isolated compounds displayed strong cytotoxicity against MCF-7 breast cancer cells, sensitive CCRF-CEM and multidrug-resistant CEM/ADR5000 leukemia cells. Compound 1 showed the most potent activity, with IC50 values of 0.07⯵M towards CCRF-CEM, 0.06⯵M towards CEM/ADR5000 and 0.36⯵M towards MCF-7 followed by compound 4 with IC50 values of 0.13⯵M towards CCRF-CEM, 0.08⯵M towards CEM/ADR5000 and 0.53⯵M towards MCF-7.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Bufanolídeos/farmacologia , Folhas de Planta/química , Plantas Medicinais/química , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/isolamento & purificação , Bufanolídeos/química , Bufanolídeos/isolamento & purificação , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Conformação Molecular , África do Sul , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
A group of triethylphosphine gold(I) and silver(I) complexes, structurally related to auranofin, were prepared and investigated as potential anticancer drug candidates. The antiproliferative properties of these metal compounds were assessed against two leukemia cell lines, i.e., CCRF-CEM and its multidrug-resistant counterpart, CEM/ADR5000. Interestingly, potent cytotoxic effects were disclosed for both series of compounds against leukemia cells, with IC50 values generally falling in the low-micromolar range, the gold derivatives being on the whole more effective than the silver analogues. Some initial structure-function relationships were drawn. Subsequently, the ability of the study compounds to inhibit the three main catalytic activities of the proteasome was investigated. Different patterns of enzyme inhibition emerged for the various metal complexes. Notably, gold compounds were able to inhibit effectively both the trypsin-like and chymotrypsin-like proteasome activities, being less effective toward the caspase-like catalytic activity. In most cases, a significant selectivity of the study compounds toward the proteasome proteolytic activities was detected when compared to other proteases. The implications of the obtained results are discussed.
Assuntos
Auranofina/farmacologia , Ouro/farmacologia , Leucemia/metabolismo , Leucemia/patologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Prata/farmacologia , Ubiquitina/metabolismo , Auranofina/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Humanos , Concentração Inibidora 50RESUMO
Alzheimer's disease (AD) is the most prevalent form of dementia. The exact pathophysiology of this disease remains incompletely understood and safe and effective therapies are required. AD is highly correlated with neuroinflammation and oxidative stress in brain causing neuronal loss. Nuclear factor of activated B-cells (NF-κB) is involved in physiological inflammatory processes and thus representing a promising target for inflammation-based AD therapy. Phytochemicals are able to interfere with the NF-κB pathway. They inhibit the phosphorylation or the ubiquitination of signaling molecules, and thus, inhibit the degradation of IκB. The translocation of NF-κB to the nucleus and subsequent transcription of pro-inflammatory cytokines are inhibited by the actions of phytochemicals. Additionally, natural compounds preventing the interaction of NF-κB can block NF-κB's transcriptional activity by inhibiting its binding to target DNA. Many polyphenols including curcumin, resveratrol, pterostilbene, punicalagin, macranthoin G, salidroside, 4-O-methylhonokiol, lycopene, genistein, obovatol and gallic acid were reported as potent NF-κB inhibitors for AD treatment. Several alkaloids such as galantamine, glaucocalyxin B, tetrandrine, berberine, oridonin, anatabine have been shown anti-inflammatory effects in AD models in vitro as well as in vivo. Besides, vitamins, tanshinone IIA, artemisinin, dihydroasparagusic acid, geniposide, xanthoceraside, l-theranine, 1,8-cineole and paeoniflorin were described as promising NF-κB inhibitors. In conclusion, natural products from plants represent interesting candidates for AD treatment. They may qualify as promising compounds for the development of derivatives providing enhanced pharmacological features.
Assuntos
Alcaloides/farmacologia , Doença de Alzheimer/tratamento farmacológico , NF-kappa B/antagonistas & inibidores , Compostos Fitoquímicos/farmacologia , Polifenóis/farmacologia , Animais , Humanos , Vitaminas/farmacologiaRESUMO
Two unique pentacyclic N,C-coupled naphthylisoquinolines, the ancistrocyclinones A (5) and B (6), were discovered in the Chinese liana Ancistrocladus tectorius. Furthermore, six known, likewise N,C-coupled alkaloids, viz., ancistrocladinium A (7a) and its mono- and bisphenolic analogs 8a and 9a were isolated, along with their atropo-diastereomers 7b, 8b, and 9b. The stereostructures of 5 and 6 were determined by HRESIMS, 1D and 2D NMR, oxidative degradation, and ECD calculations. The pentacyclic ancistrocyclinones A (5) and B (6) are structurally similar to berberine alkaloids - yet arising from a most different biosynthetic pathway: they are apparently formed by N,C-coupling of their polyketide-derived molecular halves, followed by oxidative cyclo-condensation. Biomimetic conversion of the co-occurring 4'-O-demethylancistrocladinium A (8a) to ancistrocyclinone A (5) via a quinoid intermediate supported the postulated pathway. Ancistrocyclinone A (5) was found to significantly inhibit the viability of drug-sensitive human leukemia (CCRF-CEM) and multidrug-resistant tumor cells (CEM/ADR5000) with comparable efficacies.
Assuntos
Alcaloides/química , Alcaloides/farmacologia , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacologia , Caryophyllales/química , Isoquinolinas/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , HumanosRESUMO
Hybridization of natural products has high potential to further improve their activities and may produce synergistic effects between linked pharmacophores. Here we report synthesis of nine new hybrids of natural products egonol, homoegonol, thymoquinone and artemisinin and evaluation of their activities against P. falciparum 3D7 parasites, human cytomegalovirus, sensitive and multidrug-resistant human leukemia cells. Most of the new hybrids exceed their parent compounds in antimalarial, antiviral and antileukemia activities and in some cases show higher in vitro efficacy than clinically used reference drugs chloroquine, ganciclovir and doxorubicin. Combined, our findings stress the high potency of these hybrids and encourages further use of the hybridization concept in applied pharmacological research.
Assuntos
Antimaláricos/química , Antineoplásicos/química , Antivirais/química , Produtos Biológicos/química , Animais , Anisóis/química , Anisóis/farmacologia , Antimaláricos/síntese química , Antimaláricos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Antivirais/síntese química , Antivirais/farmacologia , Artemisininas/química , Artemisininas/farmacologia , Benzofuranos/química , Benzofuranos/farmacologia , Benzoquinonas/química , Benzoquinonas/farmacologia , Produtos Biológicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cristalografia por Raios X , Citomegalovirus/efeitos dos fármacos , Humanos , Conformação Molecular , Plasmodium falciparum/efeitos dos fármacosRESUMO
Four new dimeric naphthylisoquinoline alkaloids, michellamine A5 (2) and mbandakamines C-E (4-6), were isolated from the Congolese plant Ancistrocladus ealaensis, along with the known dimer mbandakamine A (3). They represent constitutionally unsymmetric dimers, each consisting of two 5,8'-coupled naphthylisoquinoline monomers. While the molecular halves of michellamine A5 (2) are linked via C-6' of both of the naphthalene moieties, i.e., via the least-hindered positions, so that the central biaryl axis is configurationally unstable and not an additional element of chirality, the mbandakamines 3-6 possess three consecutive stereogenic axes. Their monomeric units are linked through an unprecedented 6',1â³-coupling in the binaphthalene core, leading to a high steric load, since the central axis is located in one of the peri-positions, neighboring one of the outer axes. In addition, four new 5,8'-coupled monomeric naphthylisoquinolines, viz., ancistroealaines C-F (7-10), were identified, along with four "naphthalene-devoid" tetra- and dihydroisoquinolines, named ealaines A-D (11-14). The new mbandakamines C (4) and D (5) showed pronounced activities against the malaria parasite Plasmodium falciparum, and they were likewise found to display strong cytotoxic activities against human leukemia (CCRF-CEM) and multi-drug-resistant tumor cells (CEM/ADR5000).
Assuntos
Alcaloides/farmacologia , Antiparasitários/farmacologia , Caryophyllales/química , Isoquinolinas/farmacologia , Naftalenos/farmacologia , Quinolinas/farmacologia , África Central , Antimaláricos/farmacologia , Antineoplásicos Fitogênicos , Linhagem Celular Tumoral , Humanos , Plasmodium falciparum/efeitos dos fármacosRESUMO
Here, we describe an efficient and diversity-oriented entry to both (-)-artemisinin (1) and its natural antipode (+)-artemisinin, starting from commercially and readily available S-(+)- and R-(-)-citronellene, respectively. Subsequently, we answered the still open question regarding the specificity of artemisinins action. By using a drug-sensitive Plasmodium falciparum NF54 strain, we showed that the antimalarial activity of artemisinin is not stereospecific. Our straightforward and biomimetic approach to this natural endoperoxide enables the synthesis of artemisinin derivatives that are not accessible through applying current methods and may help to address the problem of emerging resistance of Plasmodium falciparum towards artemisinin.
Assuntos
Antimaláricos/síntese química , Artemisininas/química , Antimaláricos/química , Antimaláricos/farmacologia , Artemisininas/síntese química , Artemisininas/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ciclização , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Humanos , Monoterpenos/química , Plasmodium falciparum/efeitos dos fármacos , EstereoisomerismoRESUMO
A D-ring contracted analogue of the strongly cytotoxic marine pyrrole alkaloid lamellarin D was synthesized and investigated for its antiproliferative action towards a wild type and a multidrug resistant (MDR) cancer cell line. The compound was found to inhibit tumor cell growth at submicromolar concentrations and showed a lower relative resistance in the MDR cell line than the antitumor drug camptothecin to which lamellarin D shows cross resistance and with which lamellarin D shares the same binding site.
Assuntos
Cumarínicos/síntese química , Cumarínicos/farmacologia , Compostos Heterocíclicos de 4 ou mais Anéis/síntese química , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Isoquinolinas/síntese química , Isoquinolinas/farmacologia , Inibidores da Topoisomerase I/síntese química , Antineoplásicos/síntese química , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Sítios de Ligação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cumarínicos/metabolismo , DNA Topoisomerases Tipo I/química , DNA Topoisomerases Tipo I/genética , DNA Topoisomerases Tipo I/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/metabolismo , Humanos , Isoquinolinas/metabolismo , Pontos de Checagem da Fase M do Ciclo Celular/efeitos dos fármacos , Simulação de Acoplamento Molecular , Mutagênese , Estrutura Terciária de Proteína , Inibidores da Topoisomerase I/metabolismo , Inibidores da Topoisomerase I/farmacologiaRESUMO
A series of new artemisinin-derived hybrids which incorporate cholic acid moieties have been synthesized and evaluated for their antileukemic activity against sensitive CCRF-CEM and multidrug-resistant CEM/ADR5000 cells. The new hybrids 20-28 showed IC50 values in the range of 0.019µM-0.192µM against CCRF-CEM cells and between 0.345µM and 7.159µM against CEM/ADR5000 cells. Amide hybrid 25 proved the most active compound against both CCRF-CEM and CEM/ADR5000 cells with IC50 value of 0.019±0.001µM and 0.345±0.031µM, respectively. A relatively low cross resistance to hybrids 20-28 in the range of 5.7-fold to 46.1-fold was measured. CEM/ADR5000 cells showed higher resistance than CCRF-CEM to all the tested compounds. Interestingly, the lowest cross resistance to 23 was observed (5.7-fold), whereas hybrid 25 showed 18.2-fold cross-resistant to CEM/ADR5000 cells. Hybrid 25 which proved even more potent than clinically used doxorubicin against CEM/ADR5000 cells may serve as a promising antileukemic agent against both sensitive and multidrug-resistant cells.
Assuntos
Antineoplásicos/farmacologia , Artemisininas/farmacologia , Leucemia/tratamento farmacológico , Antineoplásicos/síntese química , Antineoplásicos/química , Artemisininas/síntese química , Artemisininas/química , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Leucemia/patologia , Estrutura Molecular , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Dioncophylline F (1), the first 5,8'-coupled dioncophyllaceous alkaloid (i.e., lacking an oxygen function at C-6 and possessing an R-configuration at C-3), was isolated from the recently described Congolese liana Ancistrocladus ileboensis. Two further, likewise Dioncophyllaceae-type, alkaloids, the dioncophyllines C2 (2) and D2 (3), were identified, along with the Ancistrocladaceae-type compound ancistrocladisine B (4), which is oxygenated at C-6 and S-configured at C-3. The structures of the new compounds were determined by spectroscopic, chemical, and chiroptical methods. The stereostructure of 1 was further confirmed by total synthesis. As a consequence of the lack of a methyl group ortho to their biaryl axes, both dioncophylline F (1) and the 7,8'-coupled dioncophylline D2 (3) occur as pairs of configurationally semistable and, thus, slowly interconverting atropo-diastereomers, whereas dioncophylline C2 (2), with its 5,1'-linkage, is configurationally stable at the axis. Eight further known naphthylisoquinolines were isolated from A. ileboensis, among them dioncophylline A (P-10), its 4'-O-demethyl analogue P-11, and 5'-O-methyldioncophylline D (7), which were found to display strong cytotoxic activities against multiple myeloma INA-6 cells (P-10 even stronger than the standard drug melphalan) and against drug-sensitive acute lymphoblastic CCRF-CEM leukemia cells and their multidrug-resistant subline, CEM/ADR5000. Moreover, the dioncophyllines 1, 3, and 7 showed high-and specific-activities against the malaria parasite Plasmodium falciparum.
Assuntos
Alcaloides/isolamento & purificação , Alcaloides/farmacologia , Antimaláricos/isolamento & purificação , Antimaláricos/farmacologia , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Isoquinolinas/isolamento & purificação , Isoquinolinas/farmacologia , Magnoliopsida/química , Mieloma Múltiplo/tratamento farmacológico , Plasmodium falciparum/efeitos dos fármacos , Alcaloides/química , Antimaláricos/química , Antineoplásicos/química , Humanos , Isoquinolinas/química , Estrutura Molecular , Ressonância Magnética Nuclear Biomolecular , Relação Estrutura-AtividadeRESUMO
The 6-epimer of the plakortide H acid (1), along with the endoperoxides plakortide E (2), plakortin (3), and dihydroplakortin (4) have been isolated from a sample of the Caribbean sponge Plakortis halichondrioides. To perform a comparative study on the cytotoxicity towards the drug-sensitive leukemia CCRF-CEM cell line and its multi-drug resistant subline CEM/ADR5000, the acid of plakortin, namely plakortic acid (5), as well as the esters plakortide E methyl ester (6) and 6-epi-plakortide H (7) were synthesized by hydrolysis and Steglich esterification, respectively. The data obtained showed that the acids (1, 2, 5) exhibited potent cytotoxicity towards both cell lines, whereas the esters showed no activity (6, 7) or weaker activity (3, 4) compared to their corresponding acids. Plakortic acid (5) was the most promising derivative with half maximal inhibitory concentration (IC50) values of ca. 0.20 µM for both cell lines.
Assuntos
Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacologia , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Leucemia/tratamento farmacológico , Plakortis/química , Poríferos/química , Ácidos/química , Animais , Região do Caribe , Linhagem Celular Tumoral , Dioxanos/química , Dioxanos/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Ésteres/química , HumanosRESUMO
Drug resistance and the severe side effects of chemotherapy necessitate the development of novel anticancer drugs. Natural products are a valuable source for drug development. Scopoletin is a coumarin compound, which can be found in several Artemisia species and other plant genera. Microarray-based RNA expression profiling of the NCI cell line panel showed that cellular response of scopoletin did not correlate to the expression of ATP-binding cassette (ABC) transporters as classical drug resistance mechanisms (ABCB1, ABCB5, ABCC1, ABCG2). This was also true for the expression of the oncogene EGFR and the mutational status of the tumor suppressor gene, TP53. However, mutations in the RAS oncogenes and the slow proliferative activity in terms of cell doubling times significantly correlated with scopoletin resistance. COMPARE and hierarchical cluster analyses of transcriptome-wide mRNA expression resulted in a set of 40 genes, which all harbored binding motifs in their promoter sequences for the transcription factor, NF-κB, which is known to be associated with drug resistance. RAS mutations, slow proliferative activity, and NF-κB may hamper its effectiveness. By in silico molecular docking studies, we found that scopoletin bound to NF-κB and its regulator IκB. Scopoletin activated NF-κB in a SEAP-driven NF-κB reporter cell line, indicating that NF-κB might be a resistance factor for scopoletin. In conclusion, scopoletin might serve as lead compound for drug development because of its favorable activity against tumor cells with ABC-transporter expression, although NF-κB activation may be considered as resistance factor for this compound. Further investigations are warranted to explore the full therapeutic potential of this natural product.
Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , NF-kappa B/biossíntese , Neoplasias/tratamento farmacológico , Extratos Vegetais/administração & dosagem , Escopoletina/administração & dosagem , Transportadores de Cassetes de Ligação de ATP/biossíntese , Transportadores de Cassetes de Ligação de ATP/genética , Artemisia/química , Linhagem Celular Tumoral , Resistência a Múltiplos Medicamentos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Simulação de Acoplamento Molecular , NF-kappa B/genética , Neoplasias/genética , Farmacogenética , Extratos Vegetais/química , Análise Serial de Proteínas , Escopoletina/química , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição RelA/biossínteseRESUMO
Ergot alkaloids are psychoactive and vasoconstricting agents of the fungus Claviceps purpurea causing poisoning such as ergotism in medieval times (St. Anthony's Fire). This class of substances also inhibits tumor growth in vitro and in vivo, though the underlying mechanisms are unclear as yet. We investigated six ergot alkaloids (agroclavine, ergosterol, ergocornin E, ergotamine, dihydroergocristine, and 1-propylagroclavine tartrate) for their cytotoxicity towards tumor cell lines of the National Cancer Institute, USA. 1-Propylagroclavine tartrate (1-PAT) revealed the strongest cytotoxicity. Out of 76 clinically established anticancer drugs, cross-resistance was found between the ergot alkaloids and 6/7 anti-hormonal drugs (=85.7 %) and 5/15 DNA-alkylating drugs (=33.3 %). The IC50 values for the six alkaloids were not correlated to well-known determinants of drug resistance, such as proliferative activity (as measured by cell doubling times, PCNA expression, and cell cycle distribution), the multidrug resistance-mediating P-glycoprotein/MDR1 and expression or mutations of oncogenes and tumor suppressor genes (EGFR, RAS, TP53). While resistance of control drugs (daunorubicin, cisplatin, erlotinib) correlated with these classical resistance mechanisms, ergot alkaloids did not. Furthermore, COMPARE and hierarchical cluster analyses were performed of mRNA microarray data to identify genes correlating with sensitivity or resistance to 1-PAT. Twenty-three genes were found with different biological functions (signal transducers, RNA metabolism, ribosome constituents, cell cycle and apoptosis regulators etc.). The expression of only 3/66 neuroreceptor genes correlated with the IC50 values for 1-PAT, suggesting that the psychoactive effects of ergot alkaloids may not play a major role for the cytotoxic activity against cancer cells. In conclusion, the cytotoxicity of ergot alkaloids is not involved in classical mechanisms of drug resistance opening the possibility to bypass resistance and to treat otherwise drug-resistant and refractory tumors. The modes of action are multifactorial, which is a typical feature of many natural compounds.
Assuntos
Antineoplásicos/farmacologia , Alcaloides de Claviceps/farmacologia , Neoplasias/tratamento farmacológico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Análise em Microsséries , Neoplasias/genética , RNA Mensageiro/genéticaRESUMO
Geldanamycin is an ansamycin-derivative of a benzoquinone isolated from Streptomyces hygroscopicus. It inhibits tyrosine kinases and heat shock protein 90 (HSP90). Geldanamycin and 11 derivatives were subjected to molecular docking to HSP90, and 17-desmethoxy-17-N,N-dimethylamino-geldanamycin (17-DMAG) was the compound with the highest binding affinity (-7.73 ± 0.12 kcal/mol) and the lowest inhibition constant (2.16 ± 0.49 µM). Therefore, 17-DMAG was selected for further experiments in comparison to geldanamycin. Multidrug resistance (MDR) represents a major problem for successful cancer therapy. We tested geldanamycin and 17-DMAG against various drug-resistant cancer cell lines. Although geldanamycin and 17-DMAG inhibited the proliferation in all cell lines tested, multidrug-resistant P-glycoprotein-overexpressing CEM/ADR5000 cells were cross-resistant, ΔEGFR-overexpressing tumor cells and p53 knockout cells were sensitive to these two compounds. COMPARE and hierarchical cluster analyses were performed, and 60 genes were identified to predict the sensitivity or resistance of 59 NCI tumor cell lines towards geldanamycin and 17-DMAG. The distribution of cell lines according to their mRNA expression profiles indicated sensitivity or resistance to both compounds with statistical significance. Moreover, bioinformatic tools were used to study possible mechanisms of action of geldanamycin and 17-DMAG. Galaxy Cistrome analyses were carried out to predict transcription factor binding motifs in the promoter regions of the candidate genes. Interestingly, the NF-ĸB DNA binding motif (Rel) was identified as the top transcription factor. Furthermore, these 60 genes were subjected to Ingenuity Pathway Analysis (IPA) to study the signaling pathway interactions of these genes. Interestingly, IPA also revealed the NF-ĸB pathway as the top network among these genes. Finally, NF-ĸB reporter assays confirmed the bioinformatic prediction, and both geldanamycin and 17-DMAG significantly inhibited NF-κB activity after exposure for 24 h. In conclusion, geldanamycin and 17-DMAG exhibited cytotoxic activity against different tumor cell lines. Their activity was not restricted to HSP90 but indicated an involvement of the NF-KB pathway.
Assuntos
NF-kappa B , Neoplasias , Lactamas Macrocíclicas/farmacologia , Simulação de Acoplamento Molecular , Benzoquinonas/farmacologia , Linhagem Celular Tumoral , Proteínas de Choque Térmico HSP90/metabolismoRESUMO
The probiotic Escherichia coli strain Nissle 1917 (EcN) is one of the few probiotics licensed as a medication in several countries. Best documented is its effectiveness in keeping patients suffering from ulcerative colitis (UC) in remission. This might be due to its ability to induce the production of human ß-defensin 2 (HBD2) in a flagellin-dependent way in intestinal epithelial cells. In contrast to ulcerative colitis, for Crohn's disease (CD) convincing evidence is lacking that EcN might be clinically effective, most likely due to the genetically based inability of sufficient defensin production in CD patients. As a first step in the development of an alternative approach for the treatment of CD patients, EcN strains were constructed which were able to produce human α-defensin 5 (HD5) or ß-defensin 2 (HBD2). For that purpose, codon-optimized defensin genes encoding either the proform with the signal sequence of human α-defensin 5 (HD5) or the gene encoding HBD2 with or without the signal sequence were cloned in an expression vector plasmid under the control of the T7 promoter. Synthesis of the encoded defensins was shown by Western blots after induction of expression and lysis of the recombinant EcN strains. Recombinant mature HBD2 with an N-terminal His-tag could be purified by Ni-column chromatography and showed antimicrobial activity against E. coli, Salmonella enterica serovar Typhimurium and Listeria monocytogenes. In a second approach, that part of the HBD2 gene which encodes mature HBD2 was fused with the yebF gene. The resulting fusion protein YebFMHBD2 was secreted from the encoding EcN mutant strain after induction of expression. Presence of YebFMHBD2 in the medium was not the result of leakage from the bacterial cells, as demonstrated in the spent culture supernatant by Western blots specific for ß-galactosidase and maltose-binding protein. The dialyzed and concentrated culture supernatant inhibited the growth of E. coli, S. enterica serovar Typhimurium and L. monocytogenes in radial diffusion assays as well as in liquid culture. This demonstrates EcN to be a suitable probiotic E. coli strain for the production of certain defensins.