Deciphering pharmacokinetics and pharmacodynamics of fosfomycin.

Fosfomycin, a low molecular weight and hydrophilic drug with negligible protein binding, is eliminated almost exclusively by glomerular filtration, whose clearance is subject to patient renal function. The volume of distribution approximates to the extracellular body water (about 0.3 L/Kg) in healthy volunteers, but it is increased in critically ill patients with bacterial infections. Fosfomycin presents a high ability to distribute into many tissues, including inflamed tissues and abscess fluids. Based on PK/PD analysis and Monte Carlo simulations, we have evaluated different fosfomycin dosing regimen to optimize the treatment of septic patients due to Enterobacterales and Pseudomonas aeruginosa. As PK/PD targets, we selected %T>MIC > 70% for all pathogens, and AUC24/MIC > 24 and AUC24/MIC > 15 for net stasis of Enterobacterales and P. aeruginosa, respectively. Pharmacokinetic parameters in critically ill patients were obtained from the literature. Several dosing regimens were studied in patients with normal renal function: fosfomycin 2-8 g given every 6-12 hours, infused over 30 minutes- 24 hours. At the susceptibility EUCAST breakpoint for Enterobacterales and Staphylococcus spp. (MIC ≤ 32 mg/L), fosfomycin 4 g/8h or higher infused over 30 minutes achieved a probability of target attainment (PTA) > 90%, based in both %T>MIC and AUC24/MIC. For MIC of 64 mg/L, fosfomycin 6 g/6h in 30-minute infusion and 8 g/ 8h in 30-minute and 6 hours infusions also achieved PTA values higher than 90%. No fosfomycin monotherapy regimen was able to achieve PK/PD targets related to antimicrobial efficacy for P. aeruginosa with MICs of 256-512 mg/L.

Application of pharmacokinetic/pharmacodynamic analysis to evaluate the adequacy of antimicrobial therapy for pediatric acute otitis media in Spain before and after the introduction of the PCV7 vaccine.

OBJECTIVE: To evaluate, by applying pharmacokinetic/pharmacodynamic (PK/PD) analysis, if the change in antibiotic susceptibility after the introduction of the 7-valent pneumococcal conjugate vaccine (PCV7) in Spain had any influence on the usefulness of the antimicrobials more frequently used as empirical treatment of pediatric acute otitis media (AOM). METHODS: PK parameters and susceptibility of Streptococcus pneumoniae and Haemophilus influenzae were obtained from bibliography. Monte Carlo simulation was used to estimate the cumulative fraction of response (CFR), understood as the expected probability of therapy success. For amoxicillin and amoxicillin/clavulanate, the target was free antibiotic concentration remaining above the minimum inhibitory concentration (MIC) for ≥50% of the dosing interval (fT>MIC≥50%), whereas for cefuroxime axetil and cefotaxime, the target was fT>MIC≥60%. CFR values ≥90% were considered successful. RESULTS: When all serotypes of S. pneumoniae are considered, amoxicillin and cefotaxime turned out to reach a high probability of success, and difference before and after vaccination was scarce. For H. influenzae, CFR values were higher with amoxicillin/clavulanate than with amoxicillin. For both microorganisms, cefuroxime axetil resulted in low probability of success in the two periods of study. CONCLUSIONS: We have shown that the introduction of the PCV7 vaccination did not lead to changes in the probability of success of the current empiric treatments of the AOM. Integrated PK/PD analysis has demonstrated to be a useful tool to identify changes in antimicrobial activity after the implantation of a vaccination program, providing complementary information to the simple assessment of MIC values.

Interleukin (IL) -1 ß, IL-6 and tumor necrosis factor in patients with seasonal flu.

OBJECTIVE: The role of tumor necrosis factor (TNF), interleukin (IL)-1ß and IL-6 in the pathogenicity of seasonal flu is unknown. METHODS: We analyzed the profiles of these cytokines in 77 flu patients and 17 controls with non-flu respiratory infection, using molecular biology techniques (real-time polymerase chain reaction). RESULTS: Flu patients had lower monocyte counts (p=0.029) and a slightly lower median level of IL-6 (P=0.05) than the control group. Twenty-four flu patients (31.2%) had pneumonia; this group had higher C-reactive proteins (p=0.01) and monocyte levels (p=0.009). Pro-inflammatory cytokines levels did not rise in patients with pneumonia complicating seasonal influenza. CONCLUSIONS: IL-6 levels were lower in adults with influenza.

Population pharmacokinetics of daptomycin in critically ill patients.

Daptomycin has shown activity against a wide range of Gram-positive bacteria; however, the approved dosages usually seem insufficient for critically ill patients. The aim of this study was to develop a population pharmacokinetic model for daptomycin in critically ill patients and to estimate the success of the therapy by applying pharmacokinetic/pharmacodynamic (PK/PD) criteria. Sixteen intensive care unit patients were included, four of whom underwent continuous renal replacement therapies (CRRT). Blood and, when necessary, effluent samples were drawn after daptomycin administration at previously defined time points. A population approach using NONMEM 7.3 was performed to analyse data. Monte Carlo simulations were executed to evaluate the suitability of different dosage regimens. The probabilities of achieving the PK/PD target value associated with treatment success (ratio of the area under the plasma concentration-time curve over 24 h divided by the minimum inhibitory concentration (AUC24/MIC ≥ 666)) and to reach daptomycin concentrations linked to toxicity (minimum concentration at steady-state (Cminss) ≥ 24.3 mg/L) were calculated. The pharmacokinetics of daptomycin was best described by a one-compartment model. Elimination was conditioned by the creatinine clearance (Clcr) and also by the extra-corporeal clearance when patients were subjected to continuous renal replacement therapy (CRRT). The PK/PD analysis confirmed that 280- and 420-mg/d dosages would not be enough to achieve high probabilities of target attainment for MIC values ≥ 1 mg/L in patients with Clcr ≥ 60 mL/min or in subjects with lower Clcrs but receiving CRRT. In these patients, higher dosages (560-840 mg/d) should be needed. When treating infections due to MIC values ≥ 4 mg/L, even the highest dose would be insufficient.

Sperm preparation after freezing improves motile sperm count, motility, and viability in frozen-thawed sperm compared with sperm preparation before freezing-thawing process.

PURPOSE: The aim of this study is to evaluate which cryopreservation protocol, freezing before or after swim-up, optimizes cryopreservation outcomes in terms of motile sperm count, motility, morphology, and viability, and also to establish whether sperm viability could be assessed based on sperm motility. METHODS: Fifty-three fresh and 53 swim-up prepared samples were considered for the first experiment. In parallel, total motility evaluation by CASA system (computer-assisted sperm analyzer) and hypoosmotic swelling test (HOS-test) was performed in each sample to compare the viability results of both methods. In the second experiment, 21 normozoospermic semen samples and 20 semen samples from male factor patients were included. After fresh ejaculate evaluation, the semen sample of each patient was divided into two aliquots, one of them was frozen before swim-up and the other was frozen after swim-up. Motility, sperm count, morphology, and viability were evaluated after thawing. RESULTS: A linear regression model allows prediction of HOS-test viability results based on total motility: HOS = 1.38 + 0.97 · TM (R 2 = 99.10, residual mean squares = 9.51). Freezing before sperm selection leads to higher total and progressive motility, total motile sperm count, and viability rates than when sperm selection is performed before freezing (P < 0.005 in all cases). In fact, sperm selection prior to freezing reaches critical values when subfertile patients are considered. CONCLUSIONS: To conclude, total motility evaluation can predict HOS-test viability results, resulting in a more objective and less time-consuming method to assess viability. In addition, sperm freezing prior to swim-up selection must be considered in order to achieve better outcomes after thawing, especially in patients presenting poor sperm baseline.

Seasonal influenza in octogenarians and nonagenarians admitted to a general hospital: epidemiology, clinical presentation and prognostic factors.

OBJECTIVE: Seasonal influenza is responsible for high annual morbidity and mortality worldwide, especially in elderly patients. The aim of the study was to analyse the epidemiological, clinical and prognostic features of influenza in octogenarians and nonagenarians admitted to a general hospital, as well as risk factors associated with mortality. METHODS: Retrospective, cross-sectional, descriptive study in patients admitted and diagnosed with influenza by molecular biology in the General University Hospital of Alicante from 1 January to 31 April 2015. RESULTS: A total of 219 patients were diagnosed with influenza in the study period: 55 (25.1%) were ≤64 years-old; 77 (35.2%) were aged 65-79; 67 (30.6%) were aged 80-89 years; and 20 (9.1%) were aged ≥90 years. Most flu episodes were caused by influenza A (n=181, 82.6%). Patients aged 80 years or older had lower glomerular filtration rate (mean: 49.7 mL/min vs. 62.2 mL/min; p=0.006), a greater need for non-invasive mechanical ventilation (22% vs 9.3%; p=0.02), greater co-morbidity due to cardiac insufficiency (40.5% vs. 16.4%; p<0.001) and chronic renal disease (32.9 vs. 20%, p=0.03), and greater mortality (19% vs. 2.9%; p<0.001). In a multivariate analysis, mortality was higher in those aged 80 or over (adjusted odds ratio [ORa] 9.2, 95% confidence interval [CI] 1.65-51.1), those who had acquired the flu in a long-term care facility (ORa 11.9, 95% CI 1.06-134), and those with hyperlactataemia (ORa 1.89, 95% CI 1.20-3.00). CONCLUSIONS: Seasonal influenza is a serious problem leading to elevated mortality in octogenarian and nonagenarian patients admitted to a general hospital.

Structural recovery of the retina in a retinoschisin-deficient mouse after gene replacement therapy by solid lipid nanoparticles.

X-linked juvenile retinoschisis (XLRS) is a retinal degenerative disorder caused by mutations in the RS1 gene encoding a protein termed retinoschisin. The disease is an excellent candidate for gene replacement therapy as the majority of mutations have been shown to lead to a complete deficiency of the secreted protein in the retinal structures. In this work, we have studied the ability of non-viral vectors based on solid lipid nanoparticles (SLN) to induce the expression of retinoschisin in photoreceptors (PR) after intravitreal administration to Rs1h-deficient mice. We designed two vectors prepared with SLN, protamine, and dextran (DX) or hyaluronic acid (HA), bearing a plasmid containing the human RS1 gene under the control of the murin opsin promoter (mOPS). In vitro, the nanocarriers were able to induce the expression of retinoschisin in a PR cell line. After injection into the murine vitreous, the formulation prepared with HA induced a higher transfection level in PR than the formulation prepared with DX. Moreover, the level of retinoschisin in the inner nuclear layer (INL), where bipolar cells are located, was also higher. Two weeks after vitreal administration into Rs1h-deficient mice, both formulations showed significant improvement of the retinal structure by inducing a decrease of cavities and PR loss, and an increase of retinal and outer nuclear layer (ONL) thickness. HA-SLN resulted in a significant higher increase in the thickness of both retina and ONL, which can be explained by the higher transfection level of PR. In conclusion, we have shown the structural improvement of the retina of Rs1h-deficient mice with PR specific expression of the RS1 gene driven by the specific promoter mOPS, after successful delivery via SLN-based non-viral vectors.

Meropenem dosing requirements against Enterobacteriaceae in critically ill patients: influence of renal function, geographical area and presence of extended-spectrum ß-lactamases.

The aim of this study was the evaluation of the influence of the susceptibility patterns of Escherichia coli and Klebsiella pneumoniae isolates, specifically the presence of extended-spectrum ß-lactamases and the geographical area (Europe and USA), on the meropenem dosing requirements in critically ill patients with different degrees of renal function by estimation of the probability of pharmacokinetic/pharmacodynamic (PK/PD) target attainment. Additionally, estimation of the PK/PD breakpoints according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) approach was also an objective. Six dosing regimens were evaluated: 0.5 g, 1 g and 2 g every 8 h given as 0.5-h or 3-h infusions. Pharmacokinetic data were obtained from the literature, and susceptibility data to meropenem for E. coli and K. pneumoniae were collected from the Tigecycline Evaluation and Surveillance Trial (T.E.S.T.) surveillance study. For the same dose level, the 3-h infusion provided a probability of target attainment (PTA) ≥90 % for minimum inhibitory concentration (MIC) values up to two-fold dilution higher than those obtained with the 0.5-h infusion. For E. coli, the cumulative fraction of response (CFR) was 100 % in most cases, and neither the dose nor the infusion length nor the geographical area significantly affected the probability to reach the target. With regards to K. pneumoniae, the CFR increased when increasing the dose and decreasing the creatinine clearance (CLCR). The CFR for Spanish and USA strains was higher than that calculated for European strains. Meropenem PK/PD breakpoints are dependent on the dose, infusion length and CLCR, ranging from 2 to 32 mg/L. Based on our results, meropenem administered as a extended infusion is the best option to treat infections due to E. coli and K. pneumoniae.

Solid lipid nanoparticle-based vectors intended for the treatment of X-linked juvenile retinoschisis by gene therapy: In vivo approaches in Rs1h-deficient mouse model.

X-linked juvenile retinoschisis (XLRS), which results from mutations in the gene RS1 that encodes the protein retinoschisin, is a retinal degenerative disease affecting between 1/5000 and 1/25,000 people worldwide. Currently, there is no cure for this disease and the treatment is based on the application of low-vision aids. The aim of the present work was the in vitro and in vivo evaluation of two different non-viral vectors based on solid lipid nanoparticles (SLNs), protamine and two anionic polysaccharides, hyaluronic acid (HA) or dextran (DX), for the treatment of XLRS. First, the vectors containing a plasmid which encodes both the reporter green fluorescent protein (GFP) and the therapeutic protein retinoschisin, under the control of CMV promoters, were characterized in vitro. Then, the vectors were subretinally or intravitreally administrated to C57BL/6 wild type mice. One week later, GFP was detected in all treated mice and in all retinal layers except in the Outer Nuclear Layer (ONL) and the Inner Nuclear Layer (INL), regardless of the administration route and the vector employed. Finally, two weeks after subretinal or intravitreal injection to Rs1h-deficient mice, GFP and retinoschisin expression was detected in all retinal layers, except in the ONL, which was maintained for at least two months after subretinal administration. The structural analysis of the treated Rs1h-deficient eyes showed a partial recovery of the retina related to the production of retinoschisin. This work shows for the first time a successful RS1 gene transfer to Rs1h-deficient animals using non-viral nanocarriers, with promising results that point to non-viral gene therapy as a feasible future therapeutic tool for retinal disorders.

Solid Lipid Nanoparticles as Non-Viral Vectors for Gene Transfection in a Cell Model of Fabry Disease.

Here, we demonstrate the ability of solid lipid nanoparticle-based non-viral vectors to increase the α-galactosidase A levels of the IMFE1 cell line, an in vitro model for target cells in Fabry disease. For this purpose, vectors containing the pR-M10-αGal A plasmid, which encodes the α-galactosidase A enzyme, were prepared; the in vitro transfection efficacy was studied in IMFE1 cells, and the results were confirmed by RT-PCR. The cellular uptake of the vectors, intracellular disposition of the plasmid, and probable endocytosis pathways of the nanoparticles were also analyzed. The vectors used for the studies carried protamine (P-DNA-SLN), dextran and protamine (D-P-DNA-SLN), or hyaluronic acid of two different molecular weights and protamine (HA150-P-DNA-SLN or HA500-P-DNA-SLN). The new formulations, which presented a particle size in the range of nanometers (from 218 nm to 348 nm) and a positive superficial charge, were able to increase α-galactosidase A activity up to 4-fold in comparison to non treated IMFE1 cells. The most efficient vectors were those that included HA, and no differences due to changes in the molecular weight of HA were detected. The observed lack of colocalization with each of the four different Nile Red-labeled vectors and transferrin or cholera toxin appears to indicate that clathrin- and caveolae-independent pathways may be involved in their cellular uptake. Additionally, colocalization with LysoTracker indicated that the formulations were exposed to lysosomal activity, which may be responsible for the release of the plasmid from the vector. In conclusion, we reveal the potential of SLN-based vectors to efficiently transfect an immortalized Fabry patient cell line.

Surgery improves survival in elderly with breast cancer. A study of 465 patients in a single institution.

INTRODUCTION: Breast cancer treatment in elderly patients is controversial. This single-centre study was conducted to review the treatment and outcomes for octogenarian women treated for breast cancer. METHODS: Data from all patients aged 80 years or more with primary breast cancer treated at our institution between 1995 and 2012 were included. Patients with carcinoma in-situ (stage 0) and advanced breast cancer (stage IV) were excluded. RESULTS: The study population consisted of 369 patients (median age 84 years). A total of 277 (75%) patients underwent surgical treatment (PST) and 92 (25%) received primary endocrine treatment (PET). Prognostic factors (HER-2, tumour grade, lymphovascular invasion and subsequent adjuvant therapy) were homogeneously distributed in both groups. PST and PET were stratified according to stage: 273 (66%) patients with early stage disease (I, IIA, IIB) and 96 (34%) with locally advanced disease (IIIA, IIIB, IIIC). Patients were followed-up for a median of 63 months. In patients with early stage disease, the mean breast cancer-specific survival (BCSS) was 109 months (95% CI = 101-115) in PST patients, and 50 months (95% CI = 40-60) in PET patients (P < 0.01). Conversely, for patients with locally advanced breast cancer, there was no significant difference in BCSS between the surgical and non-surgical groups. In the PST group, BCSS and disease-free survival were significantly better among patients who underwent standard surgical treatment than among those who received suboptimal treatment. There were no differences in the Charlson comorbidity index scores between the PST and PET groups. CONCLUSION: In women ≥80 years with early-stage breast cancer, standard surgical treatment was associated with a better BCSS when compared with PET.

Pharmacokinetic/pharmacodynamic analysis to evaluate ceftaroline fosamil dosing regimens for the treatment of community-acquired bacterial pneumonia and complicated skin and skin-structure infections in patients with normal and impaired renal function.

In this study, the probability of pharmacokinetic/pharmacodynamic target attainment (PTA) of ceftaroline against clinical isolates causing community-acquired bacterial pneumonia (CABP) and complicated skin and skin-structure infection (cSSSI) in Europe was evaluated. Three dosing regimens were assessed: 600 mg every 12 h (q12 h) as a 1-h infusion (standard dose) or 600 mg every 8 h (q8 h) as a 2-h infusion in virtual patients with normal renal function; and 400 mg q12 h as a 1-h infusion in patients with moderate renal impairment. Pharmacokinetic and microbiological data were obtained from the literature. The PTA and the cumulative fraction of response (CFR) were calculated by Monte Carlo simulation. In patients with normal renal function, the ceftaroline standard dose (600 mg q12 h as a 1-h infusion) can be sufficient to treat CABP due to ceftazidime-susceptible (CAZ-S) Escherichia coli, CAZ-S Klebsiella pneumoniae, meticillin-susceptible Staphylococcus aureus, Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis (CFR>90%). However, against meticillin-resistant S. aureus (MRSA), the CFR was 72%. In cSSSI, the CFR was also <80% for MRSA. Administration of ceftaroline 600 mg q8 h as a 2-h infusion or 400 mg q12 h as a 1-h infusion in patients with moderate renal insufficiency provided a high probability of treatment success (CFR ca. 100%) for most micro-organisms causing CABP and cSSSI, including MRSA and penicillin-non-susceptible S. pneumoniae. These results suggest that in patients with normal renal function, ceftaroline 600 mg q8 h as a 2-h infusion may be a better option than the standard dose, especially if the MRSA rate is high.

Biodistribution of Nanostructured Lipid Carriers (NLCs) after intravenous administration to rats: influence of technological factors.

Nanoparticles for medical applications are frequently administered via parenteral administration. In this study, the tissue distribution of three lipid formulations based on Nanostructured Lipid Carriers (NLCs) after intravenous administration to rats was evaluated. NLCs were prepared by a high pressure homogenization method and varied in terms of particle size, surface charge, and surfactant content. The (99m)Tc radiolabeled NLCs were intravenously administered to rats, and radioactivity levels in blood and tissues were measured. Cmax, AUC0-24, and MRT0-24 were obtained from the radioactivity level versus time profiles. The radiolabeled nanocarriers exhibited a long circulation time since radioactivity was detected in blood even 24 h post-injection. No differences on the MRT values in blood among the NLCs were observed, in spite of the different particle size and surface charge. The highest radioactivity levels were measured in the kidney, followed by the bone marrow, the liver, and the spleen. In the kidney, there was a higher accumulation of the positive nanoparticles, and in the liver, uptake of negative nanoparticles was higher than positive ones. NLCs with the largest particle size showed a higher uptake in the lung and lower accumulation in liver and bone marrow, in comparison with the smaller ones.

Hábitos de sueño de la población infantil del Grau de Gandía. Un estudio descriptivo / Sleep habits of the children in the Grau de Gandia. A descriptive study

El insomnio es es trastorno de sueño infantil más frecuente y puede aparecer en la edad lactante o en la etapa preescolar, siendo de utilidad para la detección precoz el uso de cuestionarios de sueño, como la versión española del Pediatric Sleep Questionnaire (PSQ). Nos planteamos conocer la prevalencia de los trastornos de sueño en una población de 3 a 6 años del Grau de Gandía, utilizando para ello la versión española del PSQ mediante una encuesta autocumplimentada por los padres de los alumnos de los diferentes colegios. Se ha realizado un estudio descriptivo de corte transversal, recogiéndose 144 encuestas válidas, realizándose la entrada de los datos en una base de datos EXCEL para su posterior descripción. Un 23,4% de los niños presentan insomnio en función de las respuestas dadas, siendo importante la presencia de otras parasomnias como somniloquias (70%), ronquidos (7.2%), bruxismo (22.3%), trastornos por movimientos rítmicos (7.1%) y un 4.1% de los niños presentan síntomas desatencionales independientemente de que se presente o no síntomas de hiperactividad, un 2.08% de las encuestas respondían a un TDHA combinado y un 6.25% sintomatología de hiperactividad-impulsividad. Nuestro estudio proporcióna datos similares a otros estudios poblacionales realizados en el medio escolar, tanto en este rango de edad como en superiores, respecto al insomnio, parasomnias, hábitos de sueño y los TDHA, siendo la etiología más frecuente la falta de educación del hábito de sueño, por lo que el papel de la enfermera es fundamental en la orientación y formación a los padres (AU)

Insomnia is the most common sleep disorder in chilhood and it may occur to unweaned babies or during the infant period. In order to be able to detect as early as possible this disease, it is useful to make sleep questionnaires, such as the Spanish Version of the Pediatric Sleep Questionnaire (PSQ). We considered to know the prevalence of sleep disorders in a population from 3 to 6 years old from the "Grau de Gandia", using de Spanish Version of PSQ by means of an autofilled survey answered by the parents of the students from the different schools. We performed a cross sectional study, collecting 144 valid surveys, performing data entry in a EXCEL database for further description. A 23.4% of the children have insomnia based on the responses and it is significant the presence of other parasomnias such as somniloquy (70%), snoring (7.2%), bruxism (22.3%), rhythmic movement disorders (7.1% ) and a 4.1% of the children have symptoms of attention deficit regardless of havingr or not symptoms of hyperactivity. A 2.08% of the survey responded to a combined ADHD and a 6.25% of the children had symptoms of hyperactivity-impulsivity. Our study provides similar data to other population studies carried out at schools at this age range and also in higher, with regard to insomnia, parasomnias, sleep habits and ADHD, being the lack of education of the habit of sleep the most common etiology, so the role of the nurse is fundamental in the orientation and training for parents (AU)

Pharmacokinetic-pharmacodynamic evaluation of daptomycin, tigecycline, and linezolid versus vancomycin for the treatment of MRSA infections in four western European countries.

PURPOSE: To evaluate the usefulness of daptomycin, tigecycline, and linezolid for the treatment of MRSA infection compared with vancomycin in Belgium, the United Kingdom/Ireland, and Spain. METHODS: The methodology included the following steps: acquisition of microbiological and pharmacokinetic data, Monte Carlo simulation, estimation of the probability of target attainment (PTA), and calculation of the cumulative fraction of response (CFR). RESULTS: We showed that differences in the susceptibility of MRSA strains among countries may justify differences in the antibiotic dose selection. Two, 3, and 4 g daily of vancomycin seem be adequate in Belgium, Spain, and United Kingdom/Ireland respectively. The CFR obtained with 50 mg tigecycline every 12 h was higher in Spain than in Belgium and the United Kingdom/Ireland, but with the highest dose (100 mg q12h) the CFR was always 100%. At least 8 mg/kg daptomycin is necessary in United Kingdom/Ireland, but 4 mg/kg may be sufficient in Spain, and probably in Belgium. Six hundred mg q12h linezolid may be adequate in the four countries. CONCLUSION: Our study reinforces the idea that the local MIC distribution must be considered in order to increase the probability of success of empirical treatment and must be periodically updated.

Novel extended-release formulation of lovastatin by one-step melt granulation: in vitro and in vivo evaluation.

The objective of this study was to apply a one-step melt granulation method to develop an extended-release formulation of lovastatin (LOV-ER). We prepared a formulation using PEG 6000 as binder agent in a laboratory scale high-shear mixer. In vitro dissolution studies showed that the release of the drug from the new formulation followed a zero-order kinetic with no differences in the release profile with either the pH media or the agitation rate. The pharmacokinetic of lovastatin and its metabolite lovastatin acid was evaluated after the administration of the new formulation to Beagle dogs in fasted conditions and after a high-fat meal, and compared to the marketed formulation Altoprev®. After the administration of LOV-ER, extended plasma profiles of lovastatin and its active metabolite were achieved in both fasted conditions and after the high-fat meal. Plasma levels of lovastatin and lovastatin acid were always higher when the LOV-ER formulation was administered with the high-fat meal. A high variability in plasma levels and pharmacokinetic parameters was obtained, being this variability higher when the formulation was administered under fasting conditions. Our results suggest that there is an increase in lovastatin bioavailability when the formulation is administered after the high-fat meal. When we compare LOV-ER and Altoprev®, both administered after the high-fat meal, we found significant differences (p<0.05) in C(max) of lovastatin and in AUC(0-∞) and MRT of lovastatin acid. No differences were detected between both formulations in fasting conditions. In this regard, the high-fat meal seems to increase the absorption extent of lovastatin from LOV-ER formulation and to delay the absorption rate of the drug from Altoprev®. In conclusion, we developed a lovastatin formulation that provided extended plasma levels that confirm that one-step melt granulation in high-shear mixer could be an easy and cost-effective technique for extended-release formulation development.

Impact of acute otitis media pathogen shifts on the clinical efficacy of several antibiotics: a therapeutic outcomes model.

The aim of this study was to predict the clinical efficacy of different antimicrobials in the treatment of patients with acute otitis media (AOM), before and after the change in the proportion of middle ear pathogens observed after the introduction of the new conjugated heptavalent penumococcal vaccine (pPCV-7). The therapeutic Outcomes model was used to predict the likelihood of clinical success. According to this mathematical model the obtained rank order of predicted clinical efficacy was similar in the pre-PVC7 period and the post-PVC period. The results suggest that ceftriaxone and amoxicillin/clavulanate are the antibiotics with the highest predicted clinical efficacy, whereas cefaclor, azithromycin, erythromycin and clarithromycin are those with the lowest predicted clinical efficacy. The differences between antibiotics with good and those with low antibacterial activity were greater when only cases of bacterial AOM were considered. Antibiotics for which the highest clinical efficacy was predicted should maximize the likelihood of cure in outpatient antibiotic treatment of AOM.

Short- and long-term stability study of lyophilized solid lipid nanoparticles for gene therapy.

Most studies in gene therapy are focused on developing more efficient non-viral vectors, ignoring their stability, even though physically and chemically stable vectors are necessary to achieve large easily shipped and stored batches. In the present work, the effect of lyophilization on the morphological characteristics and transfection capacity of solid lipid nanoparticles (LyoSLN) and SLN-DNA vectors (Lyo(SLN-DNA)) has been evaluated. The lyophilized preparations were stored under three different sets of temperature and humidity ICH conditions: 25 degrees C/60%RH, 30 degrees C/65%RH and 40 degrees C/75%RH. After lyophilization we found an increase in particle size which did not imply a reduction of "in vitro" transfection capacity. Stability studies of formulations lyophilized with trehalose showed that SLNs were physically stable during 9 months at 25 degrees C/60%RH and 6 months at 30 degrees C/65%RH. This stability was lost when harder conditions were employed (40 degrees C/75%RH). LyoSLNs maintained or increased the transfection efficacy (from 19% to approximately 40% EGFP positive cells) over time only at 25 degrees C/60%RH and 30 degrees C/65%RH. Lyo(SLN-DNA) resulted in almost no transfection under all conditions. LyoSLNs showed less DNA condensation capacity, whereas in Lyo(SLN-DNA) the plasmid became strongly bound, hampering the transfection. Furthermore, the storage of lyophilized lipoplexes stabilized with the disaccharide trehalose did not affect cell viability.

A proline-rich peptide improves cell transfection of solid lipid nanoparticle-based non-viral vectors.

The aim of this work was to improve the transfection efficacy of solid lipid nanoparticle (SLN)-based non-viral vectors into ARPE-19 cells through the addition of Sweet Arrow Peptide (SAP). First, we prepared SAP-DNA complexes at ratios of at least 50:1, and then incorporated them into the SLNs. All formulations were able to protect DNA, and the peptide favoured the most bioactive form (supercoiled) of open circular DNA turns. In vitro transfection studies of the vectors containing the pCMS-EGFP plasmid in HEK293 and ARPE-19 cell lines revealed that incorporation of SAP led to greater transfection in both cell lines, although via different mechanisms. The presence of SAP in the formulations did not affect the viability of HEK293 or ARPE-19 cells. In HEK293 cells, SAP enabled greater uptake of the vectors, and an SAP to DNA ratio of 50:1 was sufficient for enhancing transfection. In contrast, in ARPE-19 cells, SAP induced a change in the dominant entrance mechanism, from clathrin endocytosis to caveolae/raft-dependent endocytosis, thereby decreasing use of the lysosomal pathway and consequently, reducing vector degradation. The extent to which SAP uses one mechanism or the other largely depends on its concentration in the formulation.