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An efficient and novel synthetic strategy for the generation of different carbocyclic moieties by ring closing carbonyl-olefin metathesis is reported. Herein, we describe a sustainably attractive protocol for one of the most powerful carbon-carbon bond-forming reactions, based on solvent-reduction, use of InCl3 catalyst, and microwave irradiation, affording target compounds with yields up to 96%.
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Introduction: The identification of chemical compounds that interfere with SARS-CoV-2 replication continues to be a priority in several academic and pharmaceutical laboratories. Computational tools and approaches have the power to integrate, process and analyze multiple data in a short time. However, these initiatives may yield unrealistic results if the applied models are not inferred from reliable data and the resulting predictions are not confirmed by experimental evidence. Methods: We undertook a drug discovery campaign against the essential major protease (MPro) from SARS-CoV-2, which relied on an in silico search strategy -performed in a large and diverse chemolibrary- complemented by experimental validation. The computational method comprises a recently reported ligand-based approach developed upon refinement/learning cycles, and structure-based approximations. Search models were applied to both retrospective (in silico) and prospective (experimentally confirmed) screening. Results: The first generation of ligand-based models were fed by data, which to a great extent, had not been published in peer-reviewed articles. The first screening campaign performed with 188 compounds (46 in silico hits and 100 analogues, and 40 unrelated compounds: flavonols and pyrazoles) yielded three hits against MPro (IC50 ≤ 25 µM): two analogues of in silico hits (one glycoside and one benzo-thiazol) and one flavonol. A second generation of ligand-based models was developed based on this negative information and newly published peer-reviewed data for MPro inhibitors. This led to 43 new hit candidates belonging to different chemical families. From 45 compounds (28 in silico hits and 17 related analogues) tested in the second screening campaign, eight inhibited MPro with IC50 = 0.12-20 µM and five of them also impaired the proliferation of SARS-CoV-2 in Vero cells (EC50 7-45 µM). Discussion: Our study provides an example of a virtuous loop between computational and experimental approaches applied to target-focused drug discovery against a major and global pathogen, reaffirming the well-known "garbage in, garbage out" machine learning principle.
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Previously , we demonstrated that the non-antibiotic penicillin derivative TAP7f inhibited melanoma metastasis in vitro and in vivo through the downregulation of ß-catenin and integrin αVß3. As angiogenesis is required for tumor growth and metastasis, we decided to explore the possible antiangiogenic effect of TAP7f. We found that TAP7f inhibited proliferation, migration, tube formation, and actin cytoskeleton organization of human endothelial cells. In a gel plug assay, an in vivo model for angiogenesis, TAP7f also blocked vascular formation induced by fibroblast growth factor 2. Furthermore, when murine B16-F10 melanoma cells pre-treated with TAP7f were injected intradermally in mice, we observed a decrease in the number and thickness of the capillaries surrounding the tumor. Additionally, TAP7f downregulated vascular endothelial growth factor (VEGF) and platelet-derived growth factor-B (PDGF-B) expression in B16-F10 cells and VEGF receptor expression in HMEC-1 endothelial cells. When the antitumor effect of TAP7f was studied in C57BL/6 J mice challenged with B16-F10 melanoma cells, a significant reduction of tumor growth was observed. Furthermore, a decreased expression of VEGF, PDGF-B, and the endothelial cell marker CD34 was observed in tumors from TAP7f-treated mice. Together, our results suggest that the antiangiogenic activity of TAP7f contributes to its antitumor and antimetastatic action and positions this penicillin derivative as an alternative or complementary agent for the treatment of melanoma. KEY MESSAGES: ⢠TAP7f inhibits proliferation, migration, tube formation, and actin cytoskeleton organization of endothelial cells. ⢠TAP7f downregulates VEGF receptor expression in endothelial cells. ⢠TAP7f downregulates VEGF and PDGF expression in melanoma cells. ⢠TAP7f inhibits angiogenesis in vivo.
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Melanoma Experimental , Factor A de Crecimiento Endotelial Vascular , Ratones , Humanos , Animales , Factor A de Crecimiento Endotelial Vascular/metabolismo , Células Endoteliales/metabolismo , Penicilinas/farmacología , Penicilinas/uso terapéutico , Neovascularización Patológica/metabolismo , Ratones Endogámicos C57BL , Melanoma Experimental/tratamiento farmacológico , Receptores de Factores de Crecimiento Endotelial Vascular/uso terapéutico , Inhibidores de la Angiogénesis/farmacología , Inhibidores de la Angiogénesis/uso terapéutico , Línea Celular TumoralRESUMEN
PURPOSE: To investigate the effect of TAP7f, a penicillin derivative previously characterized as a potent antitumor agent that promotes ER stress and apoptosis, in combination with thapsigargin, an ER stress inducer, on melanoma cells. METHODS: The synergistic antiproliferative effect of TAP7f in combination with thapsigargin was studied in vitro in murine B16-F0 melanoma cells, and in human A375 and SB2 melanoma cells. In vivo assays were performed with C57BL/6J mice challenged with B16-F0 cells. Immunofluorescence and Western blot assays were carried out to characterize the induction of ER stress and apoptosis. Necrotic tumor areas and the potential toxicity of the combined therapy were examined by histological analysis of tissue sections after hematoxylin-eosin staining. RESULTS: In vitro, the combination of TAP7f with thapsigargin synergistically inhibited the proliferation of murine B16-F0, and human A375 and SB2 melanoma cells. When non-inhibitory doses of each drug were simultaneously administered to C57BL/6J mice challenged with B16-F0 cells, a 50% reduction in tumor volumes was obtained in the combined group. An apoptotic response characterized by higher expression levels of Baxenhanced PARP-1 cleavage and the presence of active caspase 3 was observed in tumors from the combined treatment. In addition, higher expression levels of GADD153/CHOP and ATF4 were found in tumors of mice treated with both drugs with respect to each drug used alone, indicating the induction of an ER stress response. No signs of tissue toxicity were observed in histological sections of different organs extracted from mice receiving the combination. CONCLUSION: The synergistic and effective antitumor action of TAP7f in combination with thapsigargin could be considered as a potential therapeutic strategy for melanoma treatment.
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Antineoplásicos , Melanoma , Animales , Humanos , Ratones , Antineoplásicos/farmacología , Apoptosis , Caspasa 3/metabolismo , Línea Celular Tumoral , Melanoma/patología , Ratones Endogámicos C57BL , Penicilinas/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Tapsigargina/farmacologíaRESUMEN
We have previously examined the in vitro and in vivo antitumor action of TAP7f, a synthetic triazolylpeptidyl penicillin, on murine melanoma cells. In this work, we explored the signal transduction pathways modulated by TAP7f in murine B16-F0 and human A375 melanoma cells, and the contribution of some intracellular signals to the apoptotic cell death. TAP7f decreased ERK1/2 phosphorylation and increased phospho-p38, phospho-JNK and phospho-Akt levels. ERK1/2 blockage suppressed cell growth, while inhibition of p38, JNK and PI3K-I pathways reduced the antitumor effect of TAP7f. Pharmacological inhibition of p38 and JNK, or blockage of PI3K-I/Akt cascade with a dominant negative PI3K-I mutant diminished Bax expression levels and PARP-1 cleavage, indicating the involvement of these pathways in apoptosis. PI3K-I/Akt inhibition also favored an autophagic response, as evidenced by the higher expression levels of Beclin-1 and LC3-II detected in transfected cells exposed to TAP7f. However, although PI3K-I/Akt blockage promoted an autophagic survival response, this mechanism appears not to be critical for TAP7f antitumor action. It was also shown that TAP7f induced ER stress by enhancing the expression of ER stress-related genes and proteins. Downregulation of CHOP protein with specific siRNA increased cell growth and decreased cleavage of PARP-1, supporting its role in apoptosis. Furthermore, it was found that activation of p38, JNK and Akt occurred downstream ER perturbation. In summary, our results showed that TAP7f triggers an apoptotic cell death in melanoma cells through induction of ER stress and activation of p38, JNK and PI3K-I/Akt pathways.
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Estrés del Retículo Endoplásmico , Melanoma , Animales , Apoptosis , Humanos , Melanoma/tratamiento farmacológico , Melanoma/genética , Ratones , Penicilinas/farmacología , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismoRESUMEN
Aim: Encouraged by the antitumor activity exhibited by triazolylpeptidyl penicillins, we decided to synthesize and evaluate a library of peptoid analogs. Results: The replacement of the dipeptide unit of the reference compound, TAP7f, was investigated. In addition, the effect of the triazole linking group on the biological activity of these new derivatives was evaluated, exchanging it with a glycine spacer. The cytotoxic effect of the library compounds was determined in the B16-F0 cell line and compared with the effects on normal murine mammary gland cells. Conclusion: Among the tested compounds, peptoid 4e exhibited the highest antiproliferative activity.
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Antineoplásicos/farmacología , Diseño de Fármacos , Penicilinas/farmacología , Peptoides/farmacología , Triazoles/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/química , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Ratones , Conformación Molecular , Penicilinas/síntesis química , Penicilinas/química , Peptoides/síntesis química , Peptoides/química , Triazoles/síntesis química , Triazoles/química , Células Tumorales CultivadasRESUMEN
The development of hybrid compounds led to the discovery of new pharmacologically active agents for some of the most critical diseases, including cancer. Herein, we describe a new series of oxadiazole-containing structures designed by a molecular hybridization approach. Penicillin derivatives and amino acids were linked to amino acid and aromatic moieties through the formation of a 1,2,4-oxadiazole ring. Alternatively, condensation between amino acid-derived hydrazides and an activated penicillanic acid led to a series of 1,3,4-oxadiazole penicillin-containing hybrids and non-cyclized diacylhydrazides. From the cytotoxicity assays it is highlighted that two 1,2,4-oxadiazoles and one 1,3,4-oxadiazole connecting a penicillin and aliphatic amino acids displayed a high degree of cytotoxic selectivity, ranging between being three and four times more potent against tumor cells than normal cells. The results give a very interesting perspective suggesting that these hybrid compounds can offer a novel antitumor scaffold with promising cytotoxicity profiles.
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The synthetic triazolylpeptidyl penicillin derivative, named TAP7f, has been previously characterized as an effective antitumor agent in vitro and in vivo against B16-F0 melanoma cells. In this study, we investigated the anti-metastatic potential of this compound on highly metastatic murine B16-F10 and human A375 melanoma cells. We found that TAP7f inhibited cell adhesion, migration and invasion in a dose-dependent manner. Additionally, we demonstrated that TAP7f downregulated integrin αvß3 expression and Wnt/ß-catenin pathway, a signaling cascade commonly related to tumor invasion and metastasis. Thus, TAP7f reduced both the enzymatic activity and the expression levels of matrix-metalloproteinases-2 and -9 in a time dependent manner. Moreover, TAP7f inhibited the expression of the transcription factor Snail and the mesenchymal markers vimentin, and N-cadherin, and up-regulated the expression of the epithelial marker E-cadherin, suggesting that the penicillin derivative affects epithelial-mesenchymal transition. Results obtained in vitro were supported by those obtained in a B16-F10-bearing mice metastatic model, that showed a significant TAP7f inhibition of lung metastasis. These findings suggest the potential of TAP7f as a chemotherapeutic agent for the treatment of metastatic melanoma.
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Propargylamines have gained importance in the area of anticancer research. We synthesized 1-substituted propargylic tertiary amines using the A3-coupling as the key step. Both, solution and solid-phase protocols, were used to provide a library of 1-substituted propargylic tertiary amines with interesting structural diversity. The triple negative breast cancer subtype is the most aggressive and it lacks effective therapeutic options, while pancreatic cancer is one of the neoplasms with worse prognosis and limited therapeutic possibilities. The development of tumor-selective drugs has always been a major challenge in cancer treatment. From our library, two propargylamines displayed a high degree of cytotoxic selectivity. These levels of selectivity give a very interesting perspective for further development of 1-substituted propargylic tertiary amines as new potential chemotherapeutic antitumor agents.
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Alquinos/farmacología , Antineoplásicos/farmacología , Propilaminas/farmacología , Pirrolidinas/farmacología , Alquinos/síntesis química , Animales , Antineoplásicos/síntesis química , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Ratones , Estructura Molecular , Propilaminas/síntesis química , Pirrolidinas/síntesis química , Bibliotecas de Moléculas Pequeñas/síntesis química , Bibliotecas de Moléculas Pequeñas/farmacología , Relación Estructura-ActividadRESUMEN
AIM: According to the need for the development of new anticancer agents, we have synthetized novel bioactive compounds and aimed to determine their antitumor action. MATERIALS & METHODS: We describe in vitro studies evaluating the effect of 35 novel chemical compounds on two triple negative murine mammary adenocarcinoma tumors. RESULTS & CONCLUSION: Three compounds were selected because of their high antitumor activity and their low toxicity to normal cells. Their effect on tumor cells apoptosis, clonogenicity and migratory capacity, were determined. We found that the selected compounds showed inhibition of viability and clonogenic capacity, and promotion of apoptosis. They also decreased the migratory capacity of tumor cells. The results obtained suggest the likelihood of their future use as antitumor and/or antimetastatic agents.
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A series of novel mimetic peptides were designed, synthesised and biologically evaluated as inhibitors of Aß42 aggregation. One of the synthesised peptidic compounds, termed compound 7 modulated Aß42 aggregation as demonstrated by thioflavin T fluorescence, acting also as an inhibitor of the cytotoxicity exerted by Aß42 aggregates. The early stage interaction between compound 7 and the Aß42 monomer was investigated by replica exchange molecular dynamics (REMD) simulations and docking studies. Our theoretical results revealed that compound 7 can elongate the helical conformation state of an early stage Aß42 monomer and it helps preventing the formation of ß-sheet structures by interacting with key residues in the central hydrophobic cluster (CHC). This strategy where early "on-pathway" events are monitored by small molecules will help the development of new therapeutic strategies for Alzheimer's disease.
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Péptidos beta-Amiloides/antagonistas & inhibidores , Oligopéptidos/farmacología , Fragmentos de Péptidos/antagonistas & inhibidores , Peptidomiméticos/farmacología , Conformación Proteica en Hélice alfa/efectos de los fármacos , Péptidos beta-Amiloides/química , Péptidos beta-Amiloides/metabolismo , Línea Celular Tumoral , Humanos , Simulación del Acoplamiento Molecular , Oligopéptidos/síntesis química , Oligopéptidos/metabolismo , Oligopéptidos/toxicidad , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Peptidomiméticos/síntesis química , Peptidomiméticos/metabolismo , Peptidomiméticos/toxicidad , Unión ProteicaRESUMEN
The transition-metal-catalyzed [2 + 2 + 2]-cycloaddition of alkynes has become a powerful atom-economical strategy for aromatic ring construction. Unfortunately, the control of the stereo-, regio-, and chemoselectivity of these processes is usually challenging, and these reactions can potentially lead to complex unuseful mixtures. While solid-phase chemistry has proven to be a successful tool for decreasing the number of cycloadducts formed and for facilitating the purification step, an integral use of the outcoming products in this complex reaction is described herein. By using an immobilized monoalkyne, the transition-metal-catalyzed [2 + 2 + 2]-cycloaddition with soluble 1,6-diyne-esters led to the simultaneous preparation of soluble and solid-supported phthalides, showing a new way to benefit from solid-phase synthetic methodologies.
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The application of the reagent-based diversification strategy for generation of libraries of biologically promising ß-lactam derivatives is described. Key features are the versatility of the linker used and the cross-metathesis functionalization at the cleavage step. From an immobilized primary library, diversity was expanded by applying different cleavage conditions, leading to a series of cholesterol absorption inhibitor analogues together with interesting hybrid compounds through incorporation of a chalcone moiety.
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Alquenos/química , Bibliotecas de Moléculas Pequeñas/síntesis química , beta-Lactamas/síntesis química , Anticolesterolemiantes/síntesis química , Anticolesterolemiantes/química , Catálisis , Estructura Molecular , Bibliotecas de Moléculas Pequeñas/química , Extracción en Fase Sólida , beta-Lactamas/químicaRESUMEN
The gold-catalyzed intermolecular hydroalkylation of olefins with ß-ketoesters represents a conceptually attractive and useful synthetic tool; however, it has been scarcely applied, remaining a challenge for chemists. The aim of the current study was to investigate the addition of these 1,3-diketo-compounds to alkenes under gold catalysis conditions, in order to establish the electronic and steric effects of the alkenyl substrates in the reaction outcome. The screening of different catalyst systems and diverse olefins enabled defining the alkenyl requirements and the best reaction conditions to efficiently achieve the coupled products.
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Alquenos/química , Ésteres/química , Oro/química , Cetonas/química , Catálisis , Estructura Molecular , Espectroscopía de Protones por Resonancia Magnética , EstereoisomerismoRESUMEN
In this study, we explored the in-vitro and in-vivo mechanism of antitumor action of a novel synthetic nonantibiotic triazolylpeptidyl penicillin derivative, named TAP7f, on B16-F0 murine melanoma cells. In-vitro assays showed that TAP7f caused an inhibition of S phase progression and a concomitant decrease of the percentage of cells in G0/G1 phase. We also found that TAP7f treatment induced an apoptotic response characterized by an increase of the sub-G1 fraction of B16-F0 hypodiploid cells, the occurrence of cells with picnotic nuclei, and the detection of phosphatidylserine exposure on the outer side of the plasma membrane. Apoptotic cell death was further characterized by the activation of caspase-8, caspase-9, and caspase-3; the increase in the proapoptotic/antiapoptotic ratio of Bcl-2 family proteins; the higher expression levels of Fas receptor and TRAIL ligand; and the cleavage of poly(ADP-ribose) polymerase, a caspase-3 substrate. The in-vivo effect of TAP7f was studied in a syngeneic C57BL/6J mouse melanoma model. Results showed that TAP7f inhibited melanoma cell proliferation in vivo, as determined by a decreased expression of proliferating cell nuclear antigen, inducing a significant reduction of tumor growth. Apoptosis in vivo was assessed by detecting active caspase-3 in tumor slices from treated mice and the expression levels of Fas, TRAIL, and Bcl-2 proteins in tumor lysates. The administration of 80 mg/kg of TAP7f to non-tumor-bearing mice showed no histopathological effects on different organ tissues. Our results suggest that TAP7f might be considered as a potential therapeutic agent for cancer treatment.
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Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales/métodos , Penicilinas/farmacología , Triazoles/farmacología , Animales , Antineoplásicos/química , Apoptosis/fisiología , Caspasas/metabolismo , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Femenino , Humanos , Melanoma Experimental/tratamiento farmacológico , Ratones Endogámicos C57BL , Penicilinas/química , Triazoles/química , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
A versatile palladium-catalyzed tandem synthetic sequence to afford E-stilbenes libraries has been developed. Excellent regio- and stereocontrol have been achieved by means of the sequence of Hiyama and Heck cross-couplings. Undesirable homocoupling byproducts were avoided employing immobilized substrates.
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Paladio/química , Bibliotecas de Moléculas Pequeñas/síntesis química , Estilbenos/síntesis química , CatálisisRESUMEN
Conjugated trienones and isomeric 2H-pyrans were found to engage in a novel cycloisomerization cascade toward cyclopenta[b]furan derivatives. Knoevenagel chemistry and pericyclic reactions meet again to expand the polyene-carbonyl manifold.
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Solid-phase synthetic strategies toward the generation of libraries of biologically relevant molecules were developed using olefin cross-metathesis as a key step. It is remarkably the formal alkane metathesis based on a one-pot, microwave-assisted, ruthenium-catalyzed cross-metathesis and reduction to obtain Csp3-Csp3 linkages.
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Alcanos/síntesis química , Alquenos/química , Alcanos/química , Catálisis , Microondas , Estructura Molecular , Rutenio/químicaRESUMEN
The solid-phase version of the Pd-catalyzed Hiyama reaction between a variety of aryltriethoxysilanes and immobilized aryl halides was developed. Smooth cross-coupling was achieved to afford the corresponding biaryl products in moderate to excellent yields. The described protocol would be particularly useful for the construction of 4'-substituted 1,1'-biphenyl derivatives.
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Compuestos de Bifenilo/síntesis química , Paladio/química , Silanos/química , Técnicas de Síntesis en Fase Sólida/métodos , Compuestos de Bifenilo/química , Catálisis , Silanos/síntesis químicaRESUMEN
An efficient solid-phase protocol for the rapid generation of libraries of biologically promising 1,2,4-benzotriazines, including amino acid-derived components, is described.