Free Radical Formation from the Antineoplastic Agent VP 16-213

Free radical formation from VP 16-213 was studied by ESR spectroscopy. Incubation of VP 16-213 with the one-electron oxidators persulphate-ferrous, myeloperoxidase (MPO)/hydrogen peroxide and horseradish peroxidase (HRP)/hydrogen peroxide readily led to the formation of a free radical. The ESR spectra obtained in the last two cases, were in perfect accord with that of a product obtained by electrochemical oxidation of VP 16-213 at +550 mV. The half-life of the free radical in 1 mM Tris (pH 7.4), 0.1 MNaClat 20°C, was 257 ± 4 s. The signal recorded on incubation with HRP/H₂O₂ or MPO/H₂O₂ did not disappear on addition of 0.3 - 1.2 mg/ml microsomal protein. From incubations with rat liver microsomes in the presence of NADPH, no ESR signals were obtained.     

Flicker spectroscopy of erythrocytes

Frequency analysis of thermally excited surface undulations of erythrocytes leading to the flicker phenomenon is applied to determine biochemically and physically induced modulations of the membrane curvature elasticity. Flicker spectra of individual cells fixed to the window of a flow chamber by polylysine are taken by phase contrast microscopy, enabling investigations of the reversibility of the structural modifications. The spectra may be approximated by Lorentzian lines in most cases. By measuring the amplitude (at zero frequency) and the line width, effects of the structural changes on the curvature elastic constant, K _c , and the wavelength distribution of the undulations may be studied separately. Effect of physically induced modifications: The temperature dependence of the flicker spectra are taken from 10°C to 37°C. Above 20°C, K _c decreases with increasing temperature whereas the reverse holds below this limit. The latter anomalous behaviour is explained in terms of a conformational change associated with protein and lipid lateral phase separation. The bending stiffness increases when the cells swell osmotically, owing to surface tension effects. The dependence of the flicker spectra on the viscosity of the suspension medium agrees with the theoretical prediction. Biochemically and drug induced modifications: 5 vol of ethanol leads to a pronounced and reversible suppression of the long wavelength undulations without altering the discoid cell shape and without affecting the bending stiffness appreciably. Adsorption of dextran to the glycocalix increases K _c by a factor of 1.6 at saturation. The bending stiffness is increased by a factor of 1.3 after cross-linking the proteins with the SH-oxidizing agent diamid. Injection of Ca⁺⁺ into the cell via ionophores evokes (within 10 min) the formation of fine — probably spectrin free — spicules. This leads to an increase in K _c by a factor of 1.3 which is explained in terms of a lateral condensation of the spectrin/actin network. The spicule formation and K _c change is completely reversible (within 2 min) after perfusion with Ca⁺⁺-free buffer. Cholesterol depletion leads first to a continuous increase in K _c without change of the cell shape whereas a sudden discocyte- to echinocyte transformation sets in below a critical steroid content. The latter transition is also observed in cell suspensions and is reminiscent of a phase transition. The anti-tumor drug actinomycin D evokes an increase in the bending stiffness K _c by a factor of two, suggesting that its effect is at least partially due to a modulation of the membrane structure. The -receptor agonist leads to a remarkable increase in K _c (by about 25%) at 10^-4 M but the effect is not reversed by the -antagonist prazosin, suggesting that the agonist exerts a non-specific effect.A new technique, dynamic reflection interference contrast microscopy, is introduced by which absolute values of the amplitudes of the surface undulations and therefore K _c can be determined. The value obtained: K _c =5·10^-13 erg is about a factor of two larger then the bending stiffness of pure lipid bilayers. We suggest that the surface undulations may also be determined by lateral fluctuations of the quasi-two-dimensional spectrin/actin network.     

Block of single cardiac Na+ channels by antiarrhythmic drugs: The effect of amiodarone,propafenone and diprafenone

Summary Cell-attached patch-clamp experiments were performed in cultured cardicyytes of neonatal rats at 19°C to analyze elementary currents through single Na⁺ channels under control conditions and in the presence of the class 1 antiarrhythmic drugs amiodarone, propafenone, and diprafenone. As observed in a cell-attached patch with only one functioning Na⁺ channel, repetitive stepping of the membrane at 0.4 Hz triggered periodically channel openings except during a silent period of about 1.5 min. The latter began and ceased abruptly and did not fit the monoexponential distribution of the run length of sweeps without activity (blank sweeps). Treating the cardiocytes with amiodarone, propafenone or diprafenone (10 to 20 mol/liter) led rapidly to a blockage and reduced the likelihood that membrane depolarization triggers the opening of Na⁺ channels. The number of blank sweeps increased at the expense of the number of sweeps with activity. The fraction of activity sweeps with superpositions, indicating the simultaneous activation of two or more Na⁺ channels, also declined. As tested with amiodarone, the run length of blank sweeps is voltage- and time-dependent, analogous to the intensity of the block of macroscopic Na⁺ currents. Open time, open-time distribution, unitary current size and the tendency to reopen did not differ in unblocked cardiac Na⁺ channels (i.e. that channel fraction capable of opening in the presence of amiodarone or propafenone) from the respective control values obtained before superfusing the cardiocytes with these drugs. Apart from its blocking action, the propafenone derivative diprafenone exerted additionally a modifying effect and reduced mean open time by up to 45%. In contrast to the block, this reduction in conducting state proved insensitive to changes in holding potential, at least between –130 and –150 mV, the range tested. This means that block was attenuated on hyperpolarization whereas the reduction in open time persisted. It is concluded that, in the presence of these drugs, unblocked cardiac Na⁺ channels share a number of properties with normal Na⁺ channels in the absence of these drugs. Shortening of channel lifetime by diprafenone may be indicative of a channel modification brought about possibli by a receptor-mediated facilitation of the transition from the open to the inactivated state.     

Synthesis and anticonvulsant activities of novel 2-(cyclopentylmethylene)hydrazinyl-1,3-thiazoles in mouse models of seizures

Synthesis, characterization and investigation of in vivo anticonvulsant activities of 13 novel cyclopentanecarbaldehyde-based 2,4-disubstituted 1,3-thiazoles are presented. Their structures were determined using ¹H and ¹³C NMR, FAB(+)-MS, HRMS and elemental analyses. The results of anticonvulsant screening reveal that seven intraperitoneally administered compounds: 3a, 3b, 3d, 3e, 3f, 3k and 3m containing F-, Cl-, Br-, CF₃-, CH₃- and adamantyl substituents demonstrated significant anticonvulsant activity in the pentylenetetrazole model with median effective doses (ED₅₀)?≤?20?mg/kg, respectively, which was approximately seven-fold lower than that reported for the reference drug, ethosuximide. Noteworthy, none of these compounds impaired animals’ motor skills in the rotarod test.     

Investigations on the interactions between naphthalimide‐based anti‐tumor drugs and human serum albumin by spectroscopic and molecular modeling methods

The interactions between the three kinds of naphthalimide‐based anti‐tumor drugs (NADA, NADB, NADC) and human serum albumin (HSA) under simulated physiological conditions were investigated by fluorescence spectroscopy, circular dichroism spectroscopy and molecular modeling. The results of the fluorescence quenching spectroscopy showed that the quenching mechanisms for different drugs were static and their affinity was in a descending order of NADA > NADB > NADC. The relative thermodynamic parameters indicated that hydrophobic force was the predominant intermolecular force in the binding of NAD to HSA, while van der Waals interactions and hydrogen bonds could not be ignored. The results of site marker competitive experiment confirmed that the binding site of HSA primarily took place in site I. Furthermore, the molecular modeling study was consistent with these results. The study of circular dichroism spectra demonstrated that the presence of NADs decreased the α‐helical content of HSA and induced the change of the secondary structure of HSA. Copyright © 2015 John Wiley & Sons, Ltd.     

戈舍瑞林联合内分泌药物对绝经前晚期乳腺癌患者疗效评估

目的:研究戈舍瑞林联合内分泌药物对绝经前晚期乳腺癌患者疾病进展的作用效果及生存率的影响作用。方法:回顾性分析绝经前晚期乳腺癌患者157例,其中78例住院期间服用戈舍瑞林作为对照组,79例采用戈舍瑞林联合内分泌药物治疗作为治疗组;对比两组患者住院期间的疾病进展状况及生存率情况。结果:两组患者标准治疗过程中数据资料比较不存在差异性(P0.05);治疗后,治疗组患者疾病进展情况明显高于对照组,在不同时间段内组间比较差异有统计学意义(P0.05);随访一年结果显示患者复发、生活质量及死亡例数情况组间对比,其中复发、生活质量为优及死亡例数情况组间比较差异均有统计学意义(P0.01)。结论:戈舍瑞林结合内分泌药物对于绝经前晚期乳腺癌患者的疾病进展具有良好的控制作用。     

A novel nucleoside hydrolase from Lactobacillus buchneri LBK78 catalyzing hydrolysis of 2′-O-methylribonucleosides

2′-O-Methylribonucleosides (2′-OMe-NRs) are promising raw materials for nucleic acid drugs because of their high thermal stability and nuclease tolerance. In the course of microbial screening for metabolic activity toward 2′-OMe-NRs, Lactobacillus buchneri LBK78 was found to decompose 2′-O-methyluridine (2′-OMe-UR). The enzyme responsible was partially purified from L. buchneri LBK78 cells by a four-step purification procedure, and identified as a novel nucleoside hydrolase. This enzyme, LbNH, belongs to the nucleoside hydrolase superfamily, and formed a homotetrameric structure composed of subunits with a molecular mass around 34 kDa. LbNH hydrolyzed 2′-OMe-UR to 2′-O-methylribose and uracil, and the kinetic constants were K_m of 0.040 mM, k_cat of 0.49 s^?1, and k_cat/K_m of 12 mM^?1 s^?1. In a substrate specificity analysis, LbNH preferred ribonucleosides and 2′-OMe-NRs as its hydrolytic substrates, but reacted weakly with 2′-deoxyribonucleosides. In a phylogenetic analysis, LbNH showed a close relationship with purine-specific nucleoside hydrolases from trypanosomes.     

In vitro interactions between anidulafungin and nonsteroidal anti-inflammatory drugs on biofilms of Candida spp.

Candida spp. are responsible for many biomaterial-related infections; they give rise to infective pathologies typically associated with biofilm formation. We recently reported that the echinocandin anidulafungin (ANF) showed a strong in vitro activity against both planktonic and biofilms cells. Herein, we report the antifungal activities of ANF alone and in association with some non-steroidal anti-inflammatory drugs (NSAIDs) against nine Candida strain biofilms: four Candida albicans, two Candida glabrata and three Candida guilliermondii. The activity of ANF was assessed using an in vitro microbiological model relevant for clinical practice. ANF proved oneself to be active against biofilms cells, and a clear-cut synergism was found against Candida species biofilms when ANF was used in combination with three NSAIDs: aspirin, diclofenac, ibuprofen. The positive synergism against Candida spp. of ANF in association with aspirin or the other NSAIDs proved to be a very effective antifungal treatment (FICI <0.5). These results may provide the starting point for new combination therapies of ANF with NSAIDs against Candida biofilm pathologies.     

Design and comparative anticonvulsant activity assessment of CNS-active alkyl-carbamoyl imidazole derivatives

A novel series of carbamoyl derivatives of alkylimidazole has been designed and their anticonvulsant activity was comparatively evaluated in the mice- and rats-maximal-electroshock (MES), subcutaneous-metrazol (scMet) seizure tests and the mice-6 Hz psychomotor (6 Hz) models. The ten new designed molecules contain in their chemical structure imidazole, alkyl side-chain and carbamate as three potential active moieties.In spite of the close structural features of the carbamoyl imidazole derivatives only compounds 7, 8, 13 and 16 were active at the MES test with ED₅₀ values ranging from 12 to 20 mg/kg coupled with high protective index (PI = TD₅₀/ED₅₀) values of 4.1–7.3 after ip administration to rats. A similar phenomenon was observed in mice where compounds 7, 8, 9, 12 had MES-ED₅₀ values of 14–26 mg/kg. Compounds 7 and 13 also demonstrated anticonvulsant activity in the 6 Hz model with ED₅₀ values of 32 and 44 mg/kg, respectively. As the most active entities, compounds 7, 8 followed by 13 and 16, thus offer an optimal efficacy–safety profile and consequently, might be promising candidates for development as new antiepileptics.