Toxicity assessment of tamoxifen by means of a bacterial model

Abstract A strain of Bacillus stearothermophilus was used as a model to study physical perturbations induced in the membrane by the cytostatic tamoxifen (TAM). This study was carried out using two lines of criteria: (1) bacterial growth, and temperature growth range, with determination of growth parameters as a function of TAM concentration; and (2) biophysical studies by differential scanning calorimetry (DSC) and by means of two fluorescent probes to evaluate perturbations promoted by the drug on the structural order of bacterial lipid membranes. The inhibition of growth induced by TAM, the structural bilayer disordering, and the shift in the phase transition temperature to a lower range were also determined in the presence of Ca2+, i.e., a natural membrane stabilizer, to elucidate further perturbing effects of TAM on membranes with putative implications in cell toxicity. Growth inhibition promoted by TAM is potentiated by an increase in growth temperature above the optimal range, but attenuated or relieved by the addition of 2.5 mM Ca2+ to the culture medium. Consistently, fluorescence polarization and DSC studies showed that Ca2+ ions (2.5 mM) effectively compensated for the destabilizing effects promoted by TAM in bacterial lipid membranes

Bacillus stearothermophilus: modelo para o estudo de interacções membranares do tamoxifeno.

O tamoxifeno (TAM) é um fármaco muito utilizado no tratamento e prevenção do cancro da mama. Apesar do tamoxifeno ser vulgarmente classificado como um antagonista de estrogénios, os múltiplos efeitos celulares deste fármaco não podem ser totalmente explicados com base na ligação ao receptor de estrogénio (RE). O TAM é um composto lipofílico que perturba as propriedades físicas de membranas nativas e de lipossomas preparados com lípidos puros, e modifica a ordem membranar em células humanas de cancro da mama. Estes efeitos em biomembranas podem estar envolvidos, tanto na sua acção farmacológica como nos seus efeitos citotóxicos. Com o objectivo de esclarecer os mecanismos de acção do TAM, a nível membranar, utilizámos, como modelo, uma bactéria. Bacillus stearothermophilus foi a bactéria utilizada neste trabalho, devido à grande experiência antecedente com este microrganismo, conhecendo-se bem a sua composição lipídica e o modo como varia ao longo do intervalo de temperaturas de crescimento. Por outro lado, esta bactéria é muito sensível a alterações ambientais, promovendo modificações da sua composição lipídica. Este apurado mecanismo de adaptação poderá também actuar em resposta à acção de moléculas estranhas, nomeadamente fármacos lipofílicos. A adição de tamoxifeno ao meio de crescimento inibe o crescimento de Bacillus stearothermophilus e induz alterações ultrastruturais características de lesão membranar. Estudos biofíscos realizados por polarização de fluorescência e por calorimetria diferencial de varrimento mostraram que o tamoxifeno perturba a organização estrutural de lipossomas preparados com os lípidos extraídos de Bacillus stearothermophilus. Adicionalmente verificou-se que Bacillus stearothermophilus modifica a composição lipídica da membrana em resposta à presença de fármaco

Differences in Hepatitis C Virus (HCV)-Specific CD8 T-Cell Phenotype during Pegylated Alpha Interferon and Ribavirin Treatment Are Related to Response to Antiviral Therapy in Patients Chronically Infected with HCV▿

CD8 T cells play a major role in antiviral immune responses. Their importance for progression to chronic hepatitis C and response to treatment are still unclear. To address these issues, hepatitis C virus (HCV)-specific CD8 T-cell responses were monitored, at the single-cell level, using HLA class I pentamers specific for HCV core and HCV NS3 epitopes, in 23 chronically infected patients during treatment with pegylated alpha interferon and ribavirin. Patients who presented a sustained-response to therapy had stronger HCV-specific CD8 T-cell responses at all time points studied. Moreover, there were clear differences in the phenotypes of these cells during therapy: in responder patients, terminally differentiated effector cells increased more rapidly, and their frequency was always higher than in nonresponder patients. Sustained-responder patients also showed a higher frequency of HCV-specific CD8 T cells producing cytotoxic factors. Overall, a late and inefficient differentiation process of HCV-specific CD8 T cells might be associated with lack of response to treatment. A better knowledge of the mechanisms underlying this impairment may be important for the development of new therapeutic strategies to maintain, restore, or increase CD8 T-cell effectiveness in chronic HCV infection

Lipid composition changes induced by tamoxifen in a bacterial model system

A putative relationship between growth impairment of Bacillus stearothermophilus by tamoxifen (TAM) and TAM-induced perturbation of the physical properties of bacterial membrane lipids has been observed. The supplementation of the growth medium with Ca2+ (a membrane stabilizer) partially relieves growth inhibition by TAM, allowing growth at TAM concentrations that fully impair growth in the basal medium. B. stearothermophilus modifies the membrane lipid composition in response to the addition of TAM to the growth medium and the response is sensitive to Ca2+. Changes in lipid composition are observed in the acyl chains and in the polar head groups of phospholipids. The physical effects of alteration in these lipids was studied by fluorescence polarization of DPH and DPH-PA. Polar lipid dispersions from TAM-adapted cells grown in a Ca2+ medium show a shift of Tm to higher temperatures and a significant increase of the structural order as compared to lipids from control cells, suggesting that TAM-induced lipid composition changes compensate for the destabilizing effects of the cytostatic on membrane organization. The polar lipids from cells grown in the basal medium containing tamoxifen are also altered, but these alterations do not promote order increase of the bilayer in spite of a deviation of Tm to higher temperatures as detected by DPH. Data indicate that B. stearothermophilus controls the membrane lipid composition in response to tamoxifen, to compensate for TAM-promoted disordering in membranes and to provide an appropriate packing of phospholipid molecules in a stable bilayer, putatively disturbed by TAM incorporation.http://www.sciencedirect.com/science/article/B6T1T-3Y6PVPV-9/1/5fd659e10f4fc0e7d0a423e4e48becd

Triple Therapy with High-Dose Proton-Pump Inhibitor, Amoxicillin, and Doxycycline Is Useless for Helicobacter pylori Eradication: A Proof-of-Concept Study

Abstract Introduction: Helicobacter pylori resistance to antibiotics is steadily increasing and multidrug-resistant strains are common and difficult to eliminate, mainly in countries where bismuth, tetracycline, furazolidone, and rifabutin are unavailable. Aim: To evaluate the efficacy and safety of a triple therapy with protonpump inhibitor (PPI), amoxicillin, and doxycycline in patients with multidrug-resistant H. pylori. Patients and Methods: This prospective study involved 16 patients (13 females; mean age -50 AE 11.3 years) infected by H. pylori with known resistance to clarithromycin, metronidazole, and levofloxacin, but susceptibility to amoxicillin and tetracycline. All patients were previously submitted to upper endoscopy with gastric biopsies for H. pylori culture and susceptibility testing by Etest. Mutations in 23S rRNA and gyrA genes were determined by real-time PCR. A 10-day eradication regimen with PPI (double-standard dose b.i.d.), amoxicillin (1000 mg b.i.d.), and doxycycline (100 mg b.i.d.) was prescribed after pretreatment with PPI during 3 days. Eradication success was assessed by 13 C-urea breath test 6-10 weeks after treatment

Beyond Maastricht IV: are standard empiric triple therapies for Helicobacter pylori

BACKGROUND: Empiric triple treatments for Helicobacter pylori (H. pylori) are increasingly unsuccessful. We evaluated factors associated with failure of these treatments in the central region of Portugal. METHODS: This single-center, prospective study included 154 patients with positive (13)C-urea breath test (UBT). Patients with no previous H. pylori treatments (Group A, n = 103) received pantoprazole 40 mg 2×/day, amoxicillin 1000 mg 12/12 h and clarithromycin (CLARI) 500 mg 12/12 h, for 14 days. Patients with previous failed treatments (Group B, n = 51) and no history of levofloxacin (LVX) consumption were prescribed pantoprazole 40 mg 2×/day, amoxicillin 1000 mg 12/12 h and LVX 250 mg 12/12 h, for 10 days. H. pylori eradication was assessed by UBT 6-10 weeks after treatment. Compliance and adverse events were assessed by verbal and written questionnaires. Risk factors for eradication failure were determined by multivariate analysis. RESULTS: Intention-to-treat and per-protocol eradication rates were Group A: 68.9% (95% CI: 59.4-77.1%) and 68.8% (95% CI: 58.9-77.2%); Group B: 52.9% (95% CI: 39.5-66%) and 55.1% (95% CI: 41.3-68.2%), with 43.7% of Group A and 31.4% of Group B reporting adverse events. Main risk factors for failure were H. pylori resistance to CLARI and LVX in Groups A and B, respectively. Another independent risk factor in Group A was history of frequent infections (OR = 4.24; 95% CI 1.04-17.24). For patients with no H. pylori resistance to CLARI, a history of frequent infections (OR = 4.76; 95% CI 1.24-18.27) and active tobacco consumption (OR = 5.25; 95% CI 1.22-22.69) were also associated with eradication failure. CONCLUSIONS: Empiric first and second-line triple treatments have unacceptable eradication rates in the central region of Portugal and cannot be used, according to Maastricht recommendations. Even for cases with no H. pylori resistance to the used antibiotics, results were unacceptable and, at least for CLARI, are influenced by history of frequent infections and tobacco consumption.info:eu-repo/semantics/publishedVersio