A physiologically-based pharmacokinetic modeling approach for dosing amiodarone in children on ECMO.

Extracorporeal membrane oxygenation (ECMO) is a cardiopulmonary bypass device commonly used to treat cardiac arrest in children. The American Heart Association guidelines for cardiopulmonary resuscitation (CPR) and emergency cardiovascular care recommend using amiodarone as a first-line agent to treat ventricular arrhythmias in children with cardiac arrest. However, there are no dosing recommendations for amiodarone to treat ventricular arrhythmias in pediatric patients on ECMO. Amiodarone has a high propensity for adsorption to the ECMO components due to its physicochemical properties leading to altered pharmacokinetics (PK) in ECMO patients. The change in amiodarone PK due to interaction with ECMO components may result in a difference in optimal dosing in patients on ECMO when compared with non-ECMO patients. To address this clinical knowledge gap, a physiologically-based pharmacokinetic model of amiodarone was developed in adults and scaled to children, followed by the addition of an ECMO compartment. The pediatric model included ontogeny functions of cytochrome P450 (CYP450) enzyme maturation across various age groups. The ECMO compartment was parameterized using the adsorption data of amiodarone obtained from ex vivo studies. Model predictions captured observed concentrations of amiodarone in pediatric patients with ECMO well with an average fold error between 0.5 and 2. Model simulations support an amiodarone intravenous (i.v) bolus dose of 22 mg/kg (neonates), 13 mg/kg (infants), 8 mg/kg (children), and 6 mg/kg (adolescents). This PBPK modeling approach can be applied to explore the dosing of other drugs used in children on ECMO.

Assessing the predictive capabilities of design heuristics and coarse-grain simulation toward understanding and optimizing site-specific covalent immobilization of ß-lactamase.

For industrial applications, covalent immobilization of enzymes provides minimum leakage, recoverability, reusability, and high stability. Yet, the suitability of a given site on the enzyme for immobilization remains a trial-and-error procedure. Here, we investigate the reliability of design heuristics and a coarse-grain molecular simulation in predicting the optimum sites for covalent immobilization of TEM-1 ß-lactamase. We utilized Escherichia coli-lysate-based cell-free protein synthesis (CFPS) to produce variants containing a site-specific incorporated unnatural amino acid with a unique moiety to facilitate site directed covalent immobilization. To constrain the number of potential immobilization sites, we investigated the predictive capability of several design heuristics. The suitability of immobilization sites was determined by analyzing expression yields, specific activity, immobilization efficiency, and stability of variants. These experimental findings are compared with coarse-grain simulation of TEM-1 domain stability and thermal stability and analyzed for a priori predictive capabilities. This work demonstrates that the design heuristics successfully identify a subset of locations for experimental validation. Specifically, the nucleotide following amber stop codon and domain stability correlate well with the expression yield and specific activity of the variants, respectively. Our approach highlights the advantages of combining coarse-grain simulation and high-throughput experimentation using CFPS to identify optimal enzyme immobilization sites.

Mechanistic discoveries and simulation-guided assay optimization of portable hormone biosensors with cell-free protein synthesis.

Nuclear receptors (NRs) influence nearly every system of the body and our lives depend on correct NR signaling. Thus, a key environmental and pharmaceutical quest is to identify and detect chemicals which interact with nuclear hormone receptors, including endocrine disrupting chemicals (EDCs), therapeutic receptor modulators, and natural hormones. Previously reported biosensors of nuclear hormone receptor ligands facilitated rapid detection of NR ligands using cell-free protein synthesis (CFPS). In this work, the advantages of CFPS are further leveraged and combined with kinetic analysis, autoradiography, and western blot to elucidate the molecular mechanism of this biosensor. Additionally, mathematical simulations of enzyme kinetics are used to optimize the biosensor assay, ultimately lengthening its readable window by five-fold and improving sensor signal strength by two-fold. This approach enabled the creation of an on-demand thyroid hormone biosensor with an observable color-change readout. This mathematical and experimental approach provides insight for engineering rapid and field-deployable CFPS biosensors and promises to improve methods for detecting natural hormones, therapeutic receptor modulators, and EDCs.

The use of medicines and standard treatment guidelines in rural Timor-Leste / 国際保健医療

<b>Background</b><br> Under resource-limited circumstances, standard clinical practice for prioritized illnesses and conditions were introduced to nurses and midwives in primary health care (PHC) facilities in Timor-Leste. This research aims to asses the use of medicines and standard treatment guidelines (STGs) in community health centers (CHCs) in Timor-Leste and to analyze factors that influence adherence to STGs.<br><b>Methods</b><br> Randomly sampled 20 CHCs without beds were visited from February to August, 2006. In each CHC, 100 retrospective samples from patient registration books and 30 prospective observations were collected and then quantitatively analyzed. Open-ended interviews to three members of health personnel per CHC were qualitatively analyzed.<br><b>Results</b><br> Use of injections in Timor-Leste was extremely low when compared to results from other countries that used the same international indicators. The percentage of encounters with an antibiotic prescribed was significantly lower for prescribers with clinical nurse training than those without the training. A significantly higher level of prescribing adherence was observed among clinical nurse prescribers. None of the facility characteristics investigated was associated with the CHC's overall prescribing adherence to STGs. Open-ended interviews to CHC health personnel revealed that changes brought about by the introduction of STGs were positively perceived by respondents, especially clinical nurses.<br><b>Discussion</b><br> Unlike previous studies on physician adherence to STGs in western countries, changes brought about by the introduction of STGs were positively perceived by PHC health personnel in Timor-Leste. STGs were developed and introduced in a policy framework that reflected local needs and reality and related with the Basic Package of Health Services policy and other policies and programs, such as human resource development, medicines policy and resource allocation plans. That fact was considered to have produced positive results in this study. Timor-Leste's experience implies a potential of STGs for non-physician health personnel working at PHC level in other resource-limited areas.