RESUMO
BACKGROUND: Identifying drug-binding targets and their corresponding sites is crucial for drug discovery and mechanism studies. Limited proteolysis-coupled mass spectrometry (LiP-MS) is a sophisticated method used for the detection of compound and protein interactions. However, in some cases, LiP-MS cannot identify the target proteins due to the small structure changes or the lack of enrichment of low-abundant protein. To overcome this drawback, we developed a thermostability-assisted limited proteolysis-coupled mass spectrometry (TALiP-MS) approach for efficient drug target discovery. RESULTS: We proved that the novel strategy, TALiP-MS, could efficiently identify target proteins of various ligands, including cyclosporin A (a calcineurin inhibitor), geldanamycin (an HSP90 inhibitor), and staurosporine (a kinase inhibitor), with accurately recognizing drug-binding domains. The TALiP protocol increased the number of target peptides detected in LiP-MS experiments by 2- to 8-fold. Meanwhile, the TALiP-MS approach can not only identify both ligand-binding stability and destabilization proteins but also shows high complementarity with the thermal proteome profiling (TPP) and machine learning-based limited proteolysis (LiP-Quant) methods. The developed TALiP-MS approach was applied to identify the target proteins of celastrol (CEL), a natural product known for its strong antioxidant and anti-cancer angiogenesis effect. Among them, four proteins, MTHFD1, UBA1, ACLY, and SND1 were further validated for their strong affinity to CEL by using cellular thermal shift assay. Additionally, the destabilized proteins induced by CEL such as TAGLN2 and CFL1 were also validated. SIGNIFICANCE: Collectively, these findings underscore the efficacy of the TALiP-MS method for identifying drug targets, elucidating binding sites, and even detecting drug-induced conformational changes in target proteins in complex proteomes.
Assuntos
Proteólise , Humanos , Espectrometria de Massas/métodos , Lactamas Macrocíclicas/farmacologia , Lactamas Macrocíclicas/química , Benzoquinonas/química , Benzoquinonas/farmacologia , Temperatura , Triterpenos Pentacíclicos/química , Ciclosporina/farmacologia , Ciclosporina/química , Ciclosporina/metabolismo , Estaurosporina/farmacologia , Estaurosporina/metabolismo , Ligantes , Descoberta de Drogas , Sítios de LigaçãoRESUMO
The induction of heat stress response (HSR) mediated by the generation of heat shock proteins (HSPs) on exposure to magnetic hyperthermia-mediated cancer therapy (MHCT) decreases the efficacy of localized heat treatment at the tumor site, and thus therapy remains a significant challenge. Hence, the present study examined differential HSR elicited in glioma cells post-MHCT under different tumor microenvironment conditions (2D monolayers, 3D monoculture, and coculture spheroids) to recognize target genes that, when downregulated, could enhance the therapeutic effect of MHCT. Gene expression analysis following MHCT revealed that HSP90 was upregulated as compared to HSP70. Hence, to enhance the efficacy of the treatment, a combinatorial strategy using 17-DMAG as an inhibitor of HSP90 following MHCT was investigated. The effects of combinatorial therapy in terms of cell viability, HSP levels by immunofluorescence and gene expression analysis, oxidative stress generation, and alterations in cellular integrity were evaluated, where combinatorial therapy demonstrated an enhanced therapeutic outcome with maximum glioma cell death. Further, in the murine glioma model, a rapid tumor inhibition of 65 and 53% was observed within 8 days at the primary and secondary tumor sites, respectively, in the MCHT + 17-DMAG group, with abscopal effect-mediated complete tumor inhibition at both the tumor sites within 20 days of MHCT. The extracellularly released HSP90 from dying tumor cells further suggested the induction of immune response supported by the upregulation of IFN-γ and calreticulin genes in the MHCT + 17-DMAG group. Overall, our findings indicate that MHCT activates host immune systems and efficiently cooperates with the HSP90 blockade to inhibit the growth of distant metastatic tumors.
Assuntos
Benzoquinonas , Glioma , Proteínas de Choque Térmico HSP90 , Hipertermia Induzida , Lactamas Macrocíclicas , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Proteínas de Choque Térmico HSP90/metabolismo , Glioma/terapia , Glioma/patologia , Glioma/imunologia , Glioma/tratamento farmacológico , Animais , Camundongos , Lactamas Macrocíclicas/farmacologia , Lactamas Macrocíclicas/química , Humanos , Benzoquinonas/farmacologia , Benzoquinonas/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Microambiente Tumoral/efeitos dos fármacosRESUMO
The efficacy of photodynamic therapy (PDT)-related cancer therapies is significantly restricted by two irreconcilable obstacles, i.e., low reactive oxygen species (ROS) generation capability and hypoxia which constrains the immune response. Herein, this work develops a self-assembled clinical photosensitizer indocyanine green (ICG) and the HSP90 inhibitor 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) nanoparticles (ISDN) without any excipient. This work discovers that the hydrophobic interaction forces between ICG and 17-DMAG promote the photostability of ICG and its intersystem crossing (ISC) process, thereby improving the ROS quantum yield from 0.112 to 0.46. Augmented ROS generation enhances PDT efficacy and further enhances immunogenic cell death (ICD) effects. 17-DMAG inhibits the HSP90/hypoxia-inducible factor 1α (HIF-1α) axis to dramatically reverse the immunosuppressive tumor microenvironment caused by PDT-aggravated hypoxia. In a mouse model of pancreatic cancer, ISDN markedly improve cytotoxic T lymphocyte infiltration and MHC I and MHC II activation, demonstrating the superior ICD effects in situ tumor and the powerful systematic antitumor immunity generation, eventually achieving vigorous antitumor and recurrence resistance. This study proposes an unsophisticated and versatile strategy to significantly improve PDT efficacy for enhancing systemic antitumor immunity and potentially extending it to multiple cancers.
Assuntos
Imunoterapia , Verde de Indocianina , Nanopartículas , Fotoquimioterapia , Fármacos Fotossensibilizantes , Espécies Reativas de Oxigênio , Fotoquimioterapia/métodos , Animais , Espécies Reativas de Oxigênio/metabolismo , Camundongos , Verde de Indocianina/química , Verde de Indocianina/farmacologia , Linhagem Celular Tumoral , Nanopartículas/química , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêutico , Humanos , Nanomedicina Teranóstica , Proteínas de Choque Térmico HSP90/metabolismo , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Microambiente Tumoral/efeitos dos fármacos , Benzoquinonas/química , Benzoquinonas/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/terapia , Neoplasias Pancreáticas/metabolismo , Lactamas Macrocíclicas/química , Lactamas Macrocíclicas/farmacologia , Morte Celular Imunogênica/efeitos dos fármacos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/terapia , Neoplasias/metabolismoRESUMO
Obtaining a microorganism strain with a broad-spectrum resistance property and highly efficient antifungal activity is important to the biocontrol strategy. Herein, a marine Streptomyces sp. HNBCa1 demonstrated a broad-spectrum resistance to 17 tested crop pathogenic fungi and exhibited a high biocontrol efficiency against mango anthracnose and banana fusarium wilt. To uncover the critical bioactive secondary metabolites basis, genome assembly and annotation, metabolomic analysis, and a semipreparative HPLC-based activity-guide method were employed. Finally, geldanamycin and ectoine involved in codifferential secondary metabolites were also found to be related to biosynthetic gene clusters in the genome of HNBCa1. Reblastatin and geldanamycin were uncovered in response to broad-spectrum resistance to the 17 crop pathogenic fungi. Our results suggested that reblastatin and geldanamycin were critical to maintaining the broad-spectrum resistance property and highly efficient antifungal activity of HNBCa1, which could be further developed as a biological control agent to control crop fungal diseases.
Assuntos
Fusarium , Lactamas Macrocíclicas , Doenças das Plantas , Metabolismo Secundário , Streptomyces , Streptomyces/genética , Streptomyces/metabolismo , Streptomyces/química , Doenças das Plantas/microbiologia , Lactamas Macrocíclicas/farmacologia , Lactamas Macrocíclicas/metabolismo , Lactamas Macrocíclicas/química , Fusarium/efeitos dos fármacos , Benzoquinonas/farmacologia , Benzoquinonas/metabolismo , Benzoquinonas/química , Fungos/genética , Antifúngicos/farmacologia , Antifúngicos/metabolismo , Antifúngicos/químicaRESUMO
A new polycyclic tetramate macrolactam designated allostreptamide (1), together with four known congeners, were isolated from the culture extract of Allostreptomyces RD068384. The planar structure of the new compound was elucidated through interpretation of NMR and MS data. The absolute configuration was determined through ROESY and ECD analyses. The isolated compounds revealed antifungal potential against fourteen Candida albicans isolates with minimum inhibitory concentrations (MICs) ranging from 64 to 2048 µg ml-1. Compound 3 showed antibiofilm action and considerably reduced the viability of five isolates (36%) in the formed biofilm. The qRT-PCR revealed that 3 downregulated the BCR1, PLB2, ALS1, and SAP5 biofilm related gene expression. Therefore, 3 could be a promising antifungal therapy for C. albicans infections.
Assuntos
Antifúngicos , Biofilmes , Candida albicans , Antifúngicos/farmacologia , Antifúngicos/química , Antifúngicos/isolamento & purificação , Biofilmes/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Lactamas Macrocíclicas/farmacologia , Lactamas Macrocíclicas/química , Lactamas Macrocíclicas/isolamento & purificação , Espectroscopia de Ressonância Magnética , Testes de Sensibilidade Microbiana , Estrutura Molecular , EstereoisomerismoRESUMO
Polyene macrolactams are a special group of natural products with great diversity, unique structural features, and a wide range of biological activities. Herein, a cryptic gene cluster for the biosynthesis of putative macrolactams was disclosed from a sponge-associated bacterium, Streptomyces sp. DSS69, by genome mining. Cloning and heterologous expression of the whole biosynthetic gene cluster led to the discovery of weddellamycin, a polyene macrolactam bearing a 23/5/6 ring skeleton. A negative regulator, WdlO, and two positive regulators, WdlA and WdlB, involved in the regulation of weddellamycin production were unraveled. The fermentation titer of weddellamycin was significantly improved by overexpression of wdlA and wdlB and deletion of wdlO. Notably, weddellamycin showed remarkable antibacterial activity against various Gram-positive bacteria including MRSA, with MIC values of 0.10-0.83 µg/mL, and antifungal activity against Candida albicans, with an MIC value of 3.33 µg/mL. Weddellamycin also displayed cytotoxicity against several cancer cell lines, with IC50 values ranging from 2.07 to 11.50 µM.
Assuntos
Antibacterianos , Lactamas Macrocíclicas , Testes de Sensibilidade Microbiana , Família Multigênica , Streptomyces , Streptomyces/genética , Streptomyces/metabolismo , Antibacterianos/farmacologia , Antibacterianos/biossíntese , Antibacterianos/química , Humanos , Lactamas Macrocíclicas/farmacologia , Lactamas Macrocíclicas/química , Lactamas Macrocíclicas/isolamento & purificação , Polienos/farmacologia , Polienos/isolamento & purificação , Polienos/química , Candida albicans/efeitos dos fármacos , Linhagem Celular Tumoral , Regiões Antárticas , Animais , Poríferos/microbiologia , Antifúngicos/farmacologia , Antifúngicos/química , Antifúngicos/isolamento & purificaçãoRESUMO
During our screening for anti-mycobacterial agents against Mycobacterium avium complex (MAC), two new polycyclic tetramate macrolactams (PTMs), named hydroxycapsimycin (1) and brokamycin (2), were isolated along with the known PTM, ikarugamycin (3), from the culture broth of marine-derived Streptomyces sp. KKMA-0239. The relative structures of 1 and 2 were elucidated by spectroscopic data analyses, including 1D and 2D NMR. Furthermore, the absolute configuration of 1 was confirmed by a single-crystal X-ray diffraction analysis. Compounds 2 and 3 exhibited moderate antimycobacterial activities against MAC, including clinically isolated drug-resistant M. avium.
Assuntos
Antibacterianos , Lactamas , Testes de Sensibilidade Microbiana , Streptomyces , Streptomyces/metabolismo , Streptomyces/química , Antibacterianos/farmacologia , Antibacterianos/química , Antibacterianos/isolamento & purificação , Complexo Mycobacterium avium/efeitos dos fármacos , Espectroscopia de Ressonância Magnética , Lactamas Macrocíclicas/farmacologia , Lactamas Macrocíclicas/química , Lactamas Macrocíclicas/isolamento & purificação , Cristalografia por Raios X , Antituberculosos/farmacologia , Antituberculosos/química , Antituberculosos/isolamento & purificação , Compostos Policíclicos/farmacologia , Compostos Policíclicos/isolamento & purificação , Compostos Policíclicos/química , Estrutura MolecularRESUMO
Paritaprevir is an orally bioavailable, macrocyclic drug used for treating chronic Hepatitis C virus (HCV) infection. Its structures have been elusive to the public until recently when one of the crystal forms is solved by microcrystal electron diffraction (MicroED). In this work, the MicroED structures of two distinct polymorphic crystal forms of paritaprevir are reported from the same experiment. The different polymorphs show conformational changes in the macrocyclic core, as well as the cyclopropyl sulfonamide and methyl pyrazinamide substituents. Molecular docking shows that one of the conformations fits well into the active site pocket of the HCV non-structural 3/4A (NS3/4A) serine protease target, and can interact with the pocket and catalytic triad via hydrophobic interactions and hydrogen bonds. These results can provide further insight for optimization of the binding of acyl sulfonamide inhibitors to the HCV NS3/4A serine protease. In addition, this also demonstrates the opportunity to derive different polymorphs and distinct macrocycle conformations from the same experiments using MicroED.
Assuntos
Ciclopropanos , Lactamas Macrocíclicas , Simulação de Acoplamento Molecular , Prolina , Sulfonamidas , Sulfonamidas/química , Sulfonamidas/farmacologia , Ciclopropanos/química , Ciclopropanos/farmacologia , Lactamas Macrocíclicas/química , Lactamas Macrocíclicas/farmacologia , Prolina/análogos & derivados , Prolina/química , Compostos Macrocíclicos/química , Compostos Macrocíclicos/farmacologia , Antivirais/química , Antivirais/farmacologia , Hepacivirus/efeitos dos fármacos , Hepacivirus/enzimologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Proteínas não Estruturais Virais/química , Proteínas não Estruturais Virais/metabolismoRESUMO
Twelve new polyenic macrolactams, cyclamenols G-R (1 and 3-13), together with the known analogue cyclamenol A (2), were isolated from Streptomyces sp. OUCMDZ-4348. Their structures were elucidated by spectroscopic analysis, quantum chemical calculations, chemical derivatizations, and Mosher's methods. The sequenced genome of OUCMDZ-4348 revealed the putative biosynthetic gene cluster of cyclamenols. It was proposed that the polycyclic natural products, cyclamenols H-R, might be formed from cyclamenols A and G through nonenzymatic intramolecular cycloadditions and oxidative cyclizations.
Assuntos
Streptomyces , Lactamas Macrocíclicas/química , Streptomyces/genética , Streptomyces/química , Regiões Antárticas , Estrutura MolecularRESUMO
Chemical investigation of Streptomyces sp. NA07423 led to the discovery of two unreported macrolactams, nagimycins A (1) and B (2). Their structures were elucidated by NMR, HRESIMS, X-ray crystallography, and comparison of experimental and theoretical ECD spectra. The nagimycins have a unique butenolide moiety rarely found in ansamycin antibiotics. Genome analysis revealed the putative biosynthetic gene cluster for nagimycins, and a likely biosynthetic pathway was proposed. Notably, compounds 1 and 2 exhibited potent antibacterial activity against two pathogenic Xanthomonas bacteria.
Assuntos
Rifabutina , Streptomyces , Lactamas Macrocíclicas/química , Rifabutina/química , Streptomyces/química , Antibacterianos/química , Espectroscopia de Ressonância MagnéticaRESUMO
The logical and effective discovery of macrolactams, structurally unique natural molecules with diverse biological activities, has been limited by a lack of targeted search methods. Herein, a targeted discovery method for natural macrolactams was devised by coupling genomic signature-based PCR screening of a bacterial DNA library with spectroscopic signature-based early identification of macrolactams. DNA library screening facilitated the efficient selection of 43 potential macrolactam-producing strains (3.6% of 1,188 strains screened). The PCR amplicons of the amine-deprotecting enzyme-coding genes were analyzed to predict the macrolactam type (α-methyl, α-alkyl, or ß-methyl) produced by the hit strains. 1H-15N HSQC-TOCSY NMR analysis of 15N-labeled culture extracts enabled macrolactam detection and structural type assignment without any purification steps. This method identified a high-titer Micromonospora strain producing salinilactam (1), a previously reported α-methyl macrolactam, and two Streptomyces strains producing new α-alkyl and ß-methyl macrolactams. Subsequent purification and spectroscopic analysis led to the structural revision of 1 and the discovery of muanlactam (2), an α-alkyl macrolactam with diene amide and tetraene chromophores, and concolactam (3), a ß-methyl macrolactam with a [16,6,6]-tricyclic skeleton. Detailed genomic analysis of the strains producing 1-3 identified putative biosynthetic gene clusters and pathways. Compound 2 displayed significant cytotoxicity against various cancer cell lines (IC50 = 1.58 µM against HCT116), whereas 3 showed inhibitory activity against Staphylococcus aureus sortase A. This genomic and spectroscopic signature-based method provides an efficient search strategy for new natural macrolactams and will be generally applicable for the discovery of nitrogen-bearing natural products.
Assuntos
Streptomyces , Estrutura Molecular , Lactamas Macrocíclicas/farmacologia , Lactamas Macrocíclicas/química , Streptomyces/metabolismo , Genômica , Reação em Cadeia da Polimerase , Família MultigênicaRESUMO
Covering: 2011 to 2021Structural division of natural naphthalenoid ansamycins, regarding the type of the core and length of the ansa chain, and their biosynthetic pathways in microorganisms are discussed. The great biosynthetic plasticity of natural naphthalenoid ansamycins is reflected in their structural variety due to the alterations within ansa bridge or naphthalenoid core portions. A comparison between the biological potency of natural and semisynthetic naphthalenoid ansamycins was performed and discussed in relation to the molecular targets in cells. The antibacterial potency of naphthalenoid ansamycins seems to be dependent on the ansa chain length and conformational flexibility - the higher flexibility of the ansa chain the better biological outcome is noted.
Assuntos
Antibacterianos , Antibacterianos/farmacologia , Lactamas Macrocíclicas/química , Conformação MolecularRESUMO
Third generation Hepatitis C virus (HCV) NS3/4A protease inhibitors (PIs), glecaprevir and voxilaprevir, are highly effective across genotypes and against many resistant variants. Unlike earlier PIs, these compounds have fluorine substitutions on the P2-P4 macrocycle and P1 moieties. Fluorination has long been used in medicinal chemistry as a strategy to improve physicochemical properties and potency. However, the molecular basis by which fluorination improves potency and resistance profile of HCV NS3/4A PIs is not well understood. To systematically analyze the contribution of fluorine substitutions to inhibitor potency and resistance profile, we used a multi-disciplinary approach involving inhibitor design and synthesis, enzyme inhibition assays, co-crystallography, and structural analysis. A panel of inhibitors in matched pairs were designed with and without P4 cap fluorination, tested against WT protease and the D168A resistant variant, and a total of 22 high-resolution co-crystal structures were determined. While fluorination did not significantly improve potency against the WT protease, PIs with fluorinated P4 caps retained much better potency against the D168A protease variant. Detailed analysis of the co-crystal structures revealed that PIs with fluorinated P4 caps can sample alternate binding conformations that enable adapting to structural changes induced by the D168A substitution. Our results elucidate molecular mechanisms of fluorine-specific inhibitor interactions that can be leveraged in avoiding drug resistance.
Assuntos
Ácidos Aminoisobutíricos , Ciclopropanos , Desenho de Fármacos , Farmacorresistência Viral , Inibidores da HCV NS3-4A Protease , Lactamas Macrocíclicas , Leucina/análogos & derivados , Prolina/análogos & derivados , Quinoxalinas , Sulfonamidas , Proteases Virais , Ácidos Aminoisobutíricos/química , Ácidos Aminoisobutíricos/farmacologia , Ciclopropanos/química , Ciclopropanos/farmacologia , Farmacorresistência Viral/genética , Flúor/química , Inibidores da HCV NS3-4A Protease/química , Inibidores da HCV NS3-4A Protease/farmacologia , Halogenação , Hepacivirus/efeitos dos fármacos , Hepacivirus/enzimologia , Hepacivirus/genética , Humanos , Lactamas Macrocíclicas/química , Lactamas Macrocíclicas/farmacologia , Leucina/química , Leucina/genética , Leucina/farmacologia , Prolina/química , Prolina/genética , Prolina/farmacologia , Quinoxalinas/química , Quinoxalinas/farmacologia , Sulfonamidas/química , Sulfonamidas/farmacologia , Proteases Virais/química , Proteases Virais/genéticaRESUMO
The controlled release of medications using nanoparticle-based drug delivery carriers is a promising method to increase the efficacy of pharmacotherapy and gene therapy. One critical issue that needs to be overcome with these drug delivery carriers is their target specificity. We focused on the cell tropism of a virus to solve this issue, i.e., we attempted to apply hepatitis B virus-like particle (HBV-VLP) as a novel hepatic cell-selective carrier for medication and DNA. To prepare HBV-VLP, 293T cells were transfected with expression plasmids carrying HBV envelope surface proteins, large envelope protein (L), and small envelope protein (S). After 72 h post-transfection, VLP-containing culture supernatants were harvested, and HBV-VLP was labeled with red fluorescent dye (DiI) and was purified by sucrose gradient ultracentrifugation. An anticancer drugs (geldanamycin or doxorubicin) and GFP-expressing plasmid DNA were incorporated into HBV-VLP, and medication- and plasmid DNA-loaded VLPs were prepared. We evaluated their delivery capabilities into hepatocytes, other organ-derived cells, and hepatocytes expressing sodium taurocholate cotransporting polypeptide (NTCP), which functions as the cellular receptor for HBV by binding to HBV L protein. HBV-VLP selectively delivered both anticancer drugs and plasmid DNA not into HepG2, Huh7, and other organ cells but into HepG2 cells expressing NTCP. In summary, we developed a novel delivery nanocarrier using HBV-VLP that could be used as a hepatitis selective drug- and DNA-carrier for cancer treatment and gene therapy.
Assuntos
Partículas Artificiais Semelhantes a Vírus/metabolismo , Portadores de Fármacos , Técnicas de Transferência de Genes , Vírus da Hepatite B/química , Proteínas do Envelope Viral/genética , Antineoplásicos/química , Antineoplásicos/farmacologia , Partículas Artificiais Semelhantes a Vírus/química , Benzoquinonas/química , Benzoquinonas/farmacologia , Carbocianinas/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/química , Doxorrubicina/farmacologia , Composição de Medicamentos/métodos , Corantes Fluorescentes/química , Expressão Gênica , Células HEK293 , Células HeLa , Células Hep G2 , Vírus da Hepatite B/metabolismo , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Hepatócitos/patologia , Humanos , Lactamas Macrocíclicas/química , Lactamas Macrocíclicas/farmacologia , Transportadores de Ânions Orgânicos Dependentes de Sódio/genética , Transportadores de Ânions Orgânicos Dependentes de Sódio/metabolismo , Plasmídeos/química , Plasmídeos/metabolismo , Receptores Virais/genética , Receptores Virais/metabolismo , Coloração e Rotulagem/métodos , Simportadores/genética , Simportadores/metabolismo , Proteínas do Envelope Viral/metabolismoRESUMO
Geldanamycin (GDM) has been modified by different type neutral/acidic/basic substituents (1-7) and by quinuclidine motif (8), transformed into ammonium salts (9-13) at C(17). These compounds have been characterised by spectroscopic and x-ray methods. Derivative 8 shows better potency than GDM in MCF-7, MDA-MB-231, A549 and HeLa (IC50s = 0.09-1.06 µM). Transformation of 8 into salts 9-13 reduces toxicity (by 11-fold) at attractive potency, e.g. MCF-7 cell line (IC50â¼2 µM). Our studies show that higher water solubility contributes to lower toxicity of salts than GDM in healthy CCD39Lu and HDF cells. The use of 13 mixtures with potentiators PEI and DOX enhanced anticancer effects from IC50â¼2 µM to IC50â¼0.5 µM in SKBR-3, SKOV-3, and PC-3 cancer cells, relative to 13. Docking studies showed that complexes between quinuclidine-bearing 8-13 and Hsp90 are stabilised by extra hydrophobic interactions between the C(17)-arms and K58 or Y61 of Hsp90.
Assuntos
Antineoplásicos/farmacologia , Benzoquinonas/farmacologia , Lactamas Macrocíclicas/farmacologia , Compostos de Amônio Quaternário/química , Antineoplásicos/química , Benzoquinonas/química , Linhagem Celular , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Sinergismo Farmacológico , Humanos , Lactamas Macrocíclicas/química , Estrutura Molecular , Sais/química , Análise Espectral/métodosRESUMO
OBJECTIVES: Pancreatic ductal adenocarcinoma is one of the deadliest cancers for which few curative therapies are available to date. Heat shock protein 90 (Hsp90) inhibitors have shown activity against numerous cancers in vitro; therefore, we tested whether they could be used to target pancreatic ductal adenocarcinoma. METHODS: Inhibitors of Hsp90 ATPase activity were applied on low-passage pancreatic cell line cultures (Panc10.05, Panc215, A6L) in a dose-response manner, and the inhibitor in vitro effect on cell growth was evaluated. Seven of novel Hsp90 inhibitors based on resorcinol fragment and 5 commercially available Hsp90 inhibitors (17-AAG, AT-13387, AUY-922, ganetespib, and rifabutin) as well as control compound triptolide were tested yielding IC50 values in 2- and 3-dimensional assays. RESULTS: The novel Hsp90 inhibitors exhibited strong effects on all 3 tested pancreatic cell line cultures (Panc10.05, Panc215, A6L) reaching the IC50 of 300 to 600 nM in 2- and 3-dimensional assays. CONCLUSIONS: Novel Hsp90 inhibitors can be developed as antipancreatic cancer agents. Their chemical structures are simpler, and they are likely to exhibit lower side effects than the much more complex inhibitors used as controls.
Assuntos
Adenosina Trifosfatases/antagonistas & inibidores , Antineoplásicos/farmacologia , Carcinoma Ductal Pancreático/metabolismo , Proliferação de Células/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Neoplasias Pancreáticas/metabolismo , Adenosina Trifosfatases/metabolismo , Antineoplásicos/química , Benzamidas/química , Benzamidas/farmacologia , Benzoquinonas/química , Benzoquinonas/farmacologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/patologia , Técnicas de Cultura de Células em Três Dimensões/métodos , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Isoindóis/química , Isoindóis/farmacologia , Isoxazóis/química , Isoxazóis/farmacologia , Lactamas Macrocíclicas/química , Lactamas Macrocíclicas/farmacologia , Estrutura Molecular , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Resorcinóis/química , Resorcinóis/farmacologia , Rifabutina/química , Rifabutina/farmacologia , Triazóis/química , Triazóis/farmacologiaRESUMO
This study demonstrates the rational fabrication of a magnetic composite nanofiber mesh that can achieve mutual synergy of hyperthermia, chemotherapy, and thermo-molecularly targeted therapy for highly potent therapeutic effects. The nanofiber is composed of biodegradable poly(ε-caprolactone) with doxorubicin, magnetic nanoparticles, and 17-allylamino-17-demethoxygeldanamycin. The nanofiber exhibits distinct hyperthermia, owing to the presence of magnetic nanoparticles upon exposure of the mesh to an alternating magnetic field, which causes heat-induced cell killing as well as enhanced chemotherapeutic efficiency of doxorubicin. The effectiveness of hyperthermia is further enhanced through the inhibition of heat shock protein activity after hyperthermia by releasing the inhibitor 17-allylamino-17-demethoxygeldanamycin. These findings represent a smart nanofiber system for potent cancer therapy and may provide a new approach for the development of localized medication delivery.
Assuntos
Benzoquinonas/farmacologia , Preparações de Ação Retardada/farmacologia , Doxorrubicina/farmacologia , Hipertermia/tratamento farmacológico , Lactamas Macrocíclicas/farmacologia , Nanofibras/química , Neoplasias/tratamento farmacológico , Benzoquinonas/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Preparações de Ação Retardada/química , Doxorrubicina/química , Liberação Controlada de Fármacos , Sinergismo Farmacológico , Compostos Férricos/química , Humanos , Lactamas Macrocíclicas/química , Células MCF-7 , Magnetismo/métodos , Nanopartículas de Magnetita/químicaRESUMO
Macrocyclic natural products are plentiful in the bacteria, archaea, and eukaryote domains of life. For the significant advantages that they provide to the producing organisms, evolution has learned how to implement various types of macrocyclization reactions into the different biosynthetic pathways and how to effect them with remarkable ease. Mankind greatly benefits from nature's pool, not least because naturally occurring macrocycles or derivatives thereof serve as important drugs for the treatment of many serious ailments.In stark contrast, macrocyclization reactions are usually perceived as difficult to accomplish by purely chemical means. While it is true that ring closure necessarily entails an entropic loss and may result in the buildup of (considerable) ring strain that must be compensated for in one way or the other, it is also fair to note tremendous methodological advances during the last decades that greatly alleviated this traditional "macrocycle challenge". It is therefore increasingly possible to explore the advantages provided by large as well as medium-size ring systems in a more systematic manner. This venture also holds the promise of increasing the "chemical space" amenable to drug development to a considerable extent.In consideration of this and other important long-term perspectives, it is appropriate to revisit the current state of the art. To this end, a number of vignettes are presented, each of which summarizes a total synthesis project targeting macrocyclic natural products of greatly different chemotypes using a variety of transformations to reach these goals. Although we were occasionally facing "dead ends", which are also delineated for the sake of a complete picture, these case studies illustrate the notion that the formation of a certain macrocyclic perimeter is (usually) no longer seriously limiting. In addition to substantial progress in the "classical" repertoire (macrolactonization and macrolactamization (pateamine A, spirastrellolide, and belizentrin)), various metal-catalyzed reactions have arguably led to the greatest leaps forward. Among them, palladium-catalyzed C-C bond formation (roseophilin and nominal xestocyclamine A) and, in particular, alkene and alkyne metathesis stand out (iejimalide, spirastrellolide, enigmazole, ingenamine, and sinulariadiolide). In some cases, different methods were pursued in parallel, thus allowing for a critical assessment and comparison.To the extent that the macrocyclic challenge is vanishing, the opportunity arises to focus attention on the postmacrocyclization phase. One may stipulate that a well-designed cyclization precursor does not only ensure efficient ring closure but also fosters and streamlines the steps that come after the event. One way to do so is dual (multiple) use in that the functional groups serving the actual cyclization reaction also find productive applications downstream from it rather than being subject to simple defunctionalization. In this context, better insight into the conformational peculiarities of large rings and the growing confidence in their accessibility in a stereochemically well defined format rejuvenate the implementation of transannular reactions or reaction cascades that can lead to rapid and substantial increases in molecular complexity. The examples summarized herein showcase such possibilities, with special emphasis on tranannular gold catalysis and the emerging ruthenium-catalyzed trans-hydrometalation chemistry for the selective functionalization of alkynes.
Assuntos
Produtos Biológicos/síntese química , Alcenos/química , Alcinos/química , Produtos Biológicos/química , Catálise , Ciclização , Compostos Heterocíclicos com 3 Anéis/síntese química , Compostos Heterocíclicos com 3 Anéis/química , Lactamas Macrocíclicas/síntese química , Lactamas Macrocíclicas/química , Macrolídeos/síntese química , Macrolídeos/química , Metais/química , Pirróis/síntese química , Pirróis/química , Compostos de Espiro/síntese química , Compostos de Espiro/químicaRESUMO
ß-lactam antibiotics are among the most important and widely used antimicrobials worldwide and are comprised of a large family of compounds, obtained by chemical modifications of the common scaffolds. Usually these modifications include the addition of active groups, but less frequently, molecules were synthesized in which either two ß-lactam rings were joined to create a single bifunctional compound, or the azetidinone ring was joined to another antibiotic scaffold or another molecule with a different activity, in order to create a molecule bearing two different pharmacophoric functions. In this review, we report some examples of these derivatives, highlighting their biological properties and discussing how this strategy can lead to the development of innovative antibiotics that can represent either novel weapons against the rampant increase of antimicrobial resistance, or molecules with a broader spectrum of action.
Assuntos
Antibacterianos/química , Azetidinas/química , Lactamas Macrocíclicas/química , beta-Lactamas/química , Animais , Antibacterianos/farmacologia , Azetidinas/farmacologia , Bactérias/efeitos dos fármacos , Infecções Bacterianas/tratamento farmacológico , Calixarenos/química , Calixarenos/farmacologia , Descoberta de Drogas , Humanos , Lactamas Macrocíclicas/farmacologia , beta-Lactamas/farmacologiaRESUMO
The natural triterpene celastrol (CE) is here used as lead compound for the design and synthesis of a panel of eleven CE carboxamides that were tested in vitro for their growth inhibitory activity against Leishmania infantum and L.tropica parasites. Among them, in vitro screening identified four basic CE carboxamides endowed with nanomolar leishmanicidal activity, against both the promastigotes and the intramacrophage Leishmania amastigotes forms. These compounds also showed low toxicity toward two human (HMEC-1 and THP-1) and one murine (BMDM) cell lines. Interestingly, the most selective CE analogue (compound 3) was also endowed with the ability to inhibit the ATPase activity of the Leishmania protein chaperone Hsp90 as demonstrated by the in vitro assay conducted on a purified, full-length recombinant protein. Preliminary investigations by comparing it with the naturally occurring Hsp90 active site inhibitor Geldanamycin (GA) in two different in vitro experiments were performed. These promising results set the basis for a future biochemical investigation of the mode of interaction of celastrol and CE-inspired compounds with Leishmania Hsp90.