RESUMO
Furmonertinib (AST2818) is a novel third-generation irreversible EGFR TKI and recently has been approved in China for the treatment of non-small cell lung cancer (NSCLC) with EGFR-sensitizing and T790M resistance mutations. In the current study, we developed a semi-mechanistic population pharmacokinetic model to characterize the nonstationary pharmacokinetics (PK) of the furmonertinib and its active metabolite AST5902 simultaneously. The PK data of furmonertinib and AST5902 were obtained from 38 NSCLC patients and 16 healthy volunteers receiving 20-240 mg furmonertinib in three clinical trials. A nonlinear mixed-effects modeling approach was used to describe the PK data. The absorption process of furmonertinib was described by a transit compartment model. The disposition of both furmonertinib and AST5902 was described by a two-compartment model. An indirect response model accounted for the autoinduction of furmonertinib metabolism mediated by CYP3A4. The model-based simulation suggested that furmonertinib clearance was increased in one cycle of treatment (orally once daily for 21 days) compared to baseline, ranging from 1.1 to 1.8 fold corresponding to the dose range of 20-240 mg. The concentration of furmonertinib was decreased over time whereas that of AST5902 was increased. Interestingly, the concentration of the total active compounds (furmonertinib and AST5902) appeared to be stable. The food intake, serum alkaline phosphatase and body weight were identified as statistically significant covariates. The mechanism of food effect on PK was investigated, where the food intake might increase the bioavailability of furmonertinib via increasing the splanchnic blood flow. Overall, a population PK model was successfully developed to characterize the nonstationary PK of furmonertinib and AST5902 simultaneously. The concentrations of total active compounds were less affected by the autoinduction of furmonertinib metabolism.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Receptores ErbB , Alimentos , Humanos , Modelos Biológicos , Mutação , Inibidores de Proteínas QuinasesRESUMO
During our search for novel prenyltransferases, a putative gene ATEG_04218 from Aspergillus terreus raised our attention and was therefore amplified from strain DSM 1958 and expressed in Escherichia coli. Biochemical investigations with the purified recombinant protein and different aromatic substrates in the presence of dimethylallyl diphosphate revealed the acceptance of all the tested tryptophan-containing cyclic dipeptides. Structure elucidation of the main enzyme products by NMR and MS analyses confirmed the attachment of the prenyl moiety to C-7 of the indole ring, proving the identification of a cyclic dipeptide C7-prenyltransferase (CdpC7PT). For some substrates, reversely C3- or N1-prenylated derivatives were identified as minor products. In comparison to the known tryptophan-containing cyclic dipeptide C7-prenyltransferase CTrpPT from Aspergillus oryzae, CdpC7PT showed a much higher substrate flexibility. It also accepted cyclo-L-Tyr-L-Tyr as substrate and catalyzed an O-prenylation at the tyrosyl residue, providing the first example from the dimethylallyltryptophan synthase (DMATS) superfamily with an O-prenyltransferase activity towards dipeptides. Furthermore, products with both C7-prenyl at tryptophanyl and O-prenyl at tyrosyl residue were detected in the reaction mixture of cyclo-L-Trp-L-Tyr. Determination of the kinetic parameters proved that (S)-benzodiazepinedione consisting of a tryptophanyl and an anthranilyl moiety was accepted as the best substrate with a K M value of 204.1 µM and a turnover number of 0.125 s(-1). Cyclo-L-Tyr-L-Tyr was accepted with a K M value of 1,411.3 µM and a turnover number of 0.012 s(-1).
Assuntos
Aspergillus/enzimologia , Dimetilaliltranstransferase/metabolismo , Dipeptídeos/metabolismo , Triptofano/metabolismo , Tirosina/metabolismo , Aspergillus/genética , Clonagem Molecular , Escherichia coli/genética , Expressão Gênica , Cinética , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Prenilação , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Especificidade por SubstratoRESUMO
Copper (Cu) is an essential micronutrient for algal growth and development; however, it is also generally considered to be one of the most toxic metals when present at higher levels. Seaweeds are often exposed to low concentrations of metals, including Cu, for long time periods. In cases of ocean outfall, they may even be abruptly exposed to high levels of metals. The physiological processes that are active under Cu stress are largely unknown. In this study, the brown macroalga Sargassum fusiforme was cultured in fresh seawater at final Cu concentrations of 0, 4, 8, 24 and 47 µM. The Cu(2+) concentration and chlorophyll autofluorescence were measured to establish the toxic effects of Cu on this economically important seaweed. The accumulation of Cu by S. fusiforme was also dependent upon the external Cu concentration. Algal growth displayed a general decline with increasing media Cu concentrations, indicating that S. fusiforme was able to tolerate Cu stress at low concentrations, while it was negatively impacted at high concentrations. The term "acute stress" was employed to indicate exposure to high Cu concentrations for 1 day in this study. On the other hand, "chronic stress" was defined as exposure to lower sub-lethal Cu concentrations for 7 days. Proteins were extracted from control and Cu-treated S. fusiforme samples and separated by two-dimensional gel electrophoresis. Distinct patterns of protein expression in the acute and chronic stress conditions were observed. Proteins related to energy metabolism and photosynthesis were reduced significantly, whereas those related to carbohydrate metabolism, protein destination, RNA degradation and signaling regulation were induced in S. fusiforme in response to acute copper stress. Energy metabolism-related proteins were significantly induced by chronic Cu stress. Proteins from other functional groups, such as those related to membranes and transport, were present in minor quantities. These results suggest that S. fusiforme is sensitive to excess Cu, regardless of the presence of acute or chronic stress. We discuss the possible function of these identified proteins, taking into consideration the information available from other plant models.
Assuntos
Cobre/toxicidade , Intoxicação por Metais Pesados , Intoxicação/metabolismo , Proteômica , Sargassum/efeitos dos fármacos , Sargassum/metabolismo , Clorofila/metabolismo , Eletroforese em Gel Bidimensional , Metabolismo Energético/efeitos dos fármacos , Monitoramento Ambiental/métodos , Metais Pesados/metabolismo , Fotossíntese/efeitos dos fármacos , Proteoma/análise , Proteoma/efeitos dos fármacos , Proteoma/metabolismo , Água do Mar/análise , Alga Marinha , Poluentes Químicos da Água/toxicidadeRESUMO
Pathogen attacks can cause significant damage to plants, posing a threaten to global food production. Plants have developed exquisite methods to rapidly store a key defensive hormone jasmonate (JA), which stimulates their entire evolutionary adaptive response to pathogen attack. However, understanding how plants initiate JA biosynthesis in response to pathogen attacks has remained elusive. In this review, we discuss the newly discovered JAV1-JAZ8-WRKY51 (JJW) complex, which plays a crucial role in regulating JA production to deter insect attacks. The JJW complex inhibits JA production in plants, maintaining a low baseline level of JA that promotes optimal plant development. However, when plants are attacked by insects, a rapid influx of calcium stimulates the JAV1 calcium-dependent protein phosphate, leading to the breakdown of the JJW complex and the activation of JA production. This surge in JA levels, initiates plant defense mechanisms against the invading insects. These findings shed light on the intricate defense system that plants have evolved to combat diseases.
RESUMO
Siderophores have great application potential in metal pollutant remediation because of their effective cost and friendly impact on the environment. However, the practical use of siderophores in the remediation of specific metals is rather limited because of the weak nonspecific interactions between the siderophores and different metals. Thus, screening for a siderophore with optimal interaction with a specific metal would be necessary. In this study, the interaction between metal ions and moieties that donate the oxygen ligands for the coordination of four types of siderophore (hydroxamates, catecholates, phenolates, and carboxylates) was modeled and analyzed. As revealed by DFT-based analysis, the four types of siderophore generally exhibited selection preference for different metal ions in the order Ga3+ > Al3+ > Fe3+ > Cr3+ > Ni2+ > Cu2+ > Zn2+ > Co2+ > Mn2+ > Hg2+ > Pb2+ > Cd2+, which was determined mainly by the electronegativity of the siderophore functional groups, the electronegativity of the metals, and the ionic radius of the metals, as well as the interaction between the siderophores and the metals. Moreover, the effect of linear or nonlinear (cyclic) structure on the affinity of each siderophore for different metal ions was evaluated. In most situations, metal-bound cyclic siderophores were found to be more stable than their linear counterparts. Thus, proper siderophores for the remediation of metal pollution may be rapidly screened using this model.
Assuntos
Mercúrio , Metais Pesados , Sideróforos , Ácidos Carboxílicos , ÍonsRESUMO
Recently, the prenyltransferase SirD was found to be responsible for the O-prenylation of tyrosine in the biosynthesis of sirodesmin PL in Leptosphaeria maculans. In this study, the behavior of SirD towards phenylalanine/tyrosine and tryptophan derivatives was investigated. Product formation has been observed with 12 of 19 phenylalanine/tyrosine derivatives. It was shown that the alanine structure attached to the benzene ring and an electron donor, e.g., OH or NH2, at its para-position are essential for the enzyme activity. Modifications were possible both at the side chain and the benzene ring. Enzyme products from seven phenylalanine/tyrosine derivatives were isolated and characterized by MS and NMR analyses including HSQC and HMBC and proven to be O- or N-prenylated derivatives at position C4 of the benzene rings. K ( M ) values of six selected derivatives were found in the range of 0.10-0.68 mM. Catalytic efficiencies (K(cat)/K(M)) were determined in the range of 430-1,110 s⻹·M⻹ with L-tyrosine as the best substrate. In addition, 7 of 14 tested tryptophan analogs were also accepted by SirD and converted to C7-prenylated derivatives, which was confirmed by comparison with products obtained from enzyme assays using a 7-dimethylallyltryptophan synthase 7-DMATS from Aspergillus fumigatus.
Assuntos
Aspergillus fumigatus/enzimologia , Dimetilaliltranstransferase/metabolismo , Prenilação , Tirosina/metabolismo , Aspergillus fumigatus/metabolismo , Cinética , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Fenilalanina/metabolismo , Triptofano/metabolismoRESUMO
A type III polyketide synthase (SfuPKS1) from the edible seaweed Sargassum fusiforme was molecularly cloned and biochemically characterized. The recombinant SfuPKS1 catalyzed the condensation of fatty acyl-CoA with two or three malonyl-CoA using lactone-type intramolecular cyclization to produce tri- and/or tetraketides. Moreover, it can also utilize phenylpropanoyl-CoA to synthesize phloroglucinol derivatives through Claisen-type cyclization, exhibiting broad substrate and catalysis specificity. Furthermore, the catalytic efficiency (kcat/KM) for acetyl-CoA was 11.8-fold higher than that for 4-coumaroyl-CoA. A pathway for the synthesis of naringenin involving SfuPKS1 was also constructed in Escherichia coli by recombinant means, resulting in 4.9 mg of naringenin per liter.
Assuntos
Sargassum , Alga Marinha , Aciltransferases , Catálise , Cinética , Especificidade por SubstratoRESUMO
A putative prenyltransferase gene cTrpPT was amplified from Aspergillus oryzae DSM1147, cloned into pQE70 and overexpressed in Escherichia coli. The overproduced His(6)-CTrpPT was purified to near homogeneity and incubated with L-tryptophan or tryptophan-containing cyclic dipeptides in the presence of dimethylallyl diphosphate. The formation of the enzyme products was monitored with HPLC. It was shown that CTrpPT differed clearly from other known indole prenyltransferases in several aspects. This enzyme showed higher substrate specificity towards aromatic substrates, but lower regioselectivity regarding the prenylation position than other indole prenyltransferases. Cyclo-L-Trp-L-Trp was much better accepted than other cyclic dipeptides tested in this study. In comparison to other indole prenyltransferases with one dominant enzyme product, at least two product peaks were detected in the reaction mixtures of CTrpPT. (1)H- and (13)C-NMR analyses, including long-range (1)H-(13)C connectivities in Heteronuclear Multiple-Bond Correlation (HMBC) and Nuclear Overhauser Effect Spectroscopy (NOESY), proved the structures of the enzyme products as C7- and N1-prenylated derivatives with a ratio of 1:1.2 using cyclo-L-Trp-L-Trp as substrate. The K(M) values were determined at about 2.5 mM for dimethylallyl diphosphate and 0.3 mM for cyclo-L-Trp-L-Trp with a turnover number of 0.33 s(-1).
Assuntos
Aspergillus oryzae/enzimologia , Biocatálise , Carbono/química , Dimetilaliltranstransferase/metabolismo , Nitrogênio/química , Peptídeos Cíclicos/química , Prenilação , Clonagem Molecular , Dimetilaliltranstransferase/biossíntese , Dimetilaliltranstransferase/genética , Indóis/química , Peptídeos Cíclicos/metabolismo , Especificidade por SubstratoRESUMO
To better understand the mechanism of inherent salt resistance in Jerusalem artichoke (Helianthus tuberosus L.), physiological and metabolic responses of tubers at the initiation stage of sprouting under different salt stress levels were evaluated in the present study. As a result, 28 metabolites were identified using proton nuclear magnetic resonance (1H-NMR) spectroscopy. Jerusalem artichoke tubers showed minor changes in metabolic response under moderate salt stress when they had not yet sprouted, where metabolism was downregulated at the start of sprouting and then upregulated significantly after plants became autotrophic. However, mild and severe salt stress levels caused different metabolic response patterns. In addition, the accumulation of fructose and sucrose was enhanced by moderate salt stress, while glucose was highly consumed. Aspartate and asparagine showed accelerated accumulation in sprouting Jerusalem artichoke tubers that became autotrophic, suggesting the enhancement of photosynthesis by moderate salt stress.
Assuntos
Frutose/metabolismo , Glucose/metabolismo , Helianthus/metabolismo , Tubérculos/metabolismo , Estresse Salino , Sacarose/metabolismo , Helianthus/crescimento & desenvolvimento , Fotossíntese , Tubérculos/crescimento & desenvolvimentoRESUMO
A fundamental investigation of the biosorption of Cd2+ from aqueous solution by the edible seaweed Sargassum fusiforme was performed under batch conditions. The influences of experimental parameters, such as the initial pH, sorption time, temperature, and initial Cd2+ concentration, on Cd2+ uptake by S. fusiforme were evaluated. The results indicated that the biosorption of Cd2+ depended on the initial Cd2+ concentration, as well as the pH. The uptake of Cd2+ could be described by the Langmuir isotherm model, and both the Langmuir biosorption equilibrium constant and the maximum biosorption capacity of the monolayer decreased with increasing temperature, thereby confirming the exothermic character of the sorption process. The biosorption kinetics follows the pseudo-second-order kinetic model, and intraparticle diffusion is the sole rate-limiting step for the entire biosorption period. These fundamental equilibrium and kinetic results can support further studies to the removal of cadmium from S. fusiforme harvested from cadmium-polluted waters.
Assuntos
Cádmio/metabolismo , Sargassum/metabolismo , Poluentes Químicos da Água/metabolismo , Absorção Fisico-Química , Transporte Biológico , Concentração de Íons de Hidrogênio , Cinética , TemperaturaRESUMO
Aquatic agriculture in heavy-metal-polluted coastal areas faces major problems due to heavy metal transfer into aquatic organisms, leading to various unexpected changes in nutrition and primary and/or secondary metabolism. In the present study, the dual role of heavy metal copper (Cu) played in the metabolism of photosynthetic organism, the edible seaweed Sargassum fusiforme, was evaluated by characterization of biochemical and metabolic responses using both 1H NMR and GC-MS techniques under acute (47 µM, 1 day) and chronic stress (8 µM, 7 days). Consequently, photosynthesis may be seriously inhibited by acute Cu exposure, resulting in decreasing levels of carbohydrates, e.g., mannitol, the main products of photosynthesis. Ascorbate may play important roles in the antioxidant system, whose content was much more seriously decreased under acute than that under chronic Cu stress. Overall, these results showed differential toxicological responses on metabolite profiles of S. fusiforme subjected to acute and chronic Cu exposures that allowed assessment of impact of Cu on marine organisms.