Nisin inducible production of listeriolysin O in Lactococcus lactis NZ9000

<p>Abstract</p> <p>Background</p> <p><it>Listeria monocytogenes </it>is a well-characterized food-borne pathogen that infects pregnant women and immunocompromised individuals. Listeriolysin O (LLO) is the major virulence factor of the pathogen and is often used as a diagnostic marker for detection of <it>L. monocytogenes</it>. In addition, LLO represents a potent antigen driving T cell-mediated immunity during infection. In the present work, <it>Lactococcus lactis </it>NZ9000 was used as an expression host to hyper-produce LLO under inducible conditions using the NICE (NIsin Controlled Expression) system. We created a modified pNZ8048 vector encoding a six-His-tagged LLO downstream of the strong inducible PnisA promoter.</p> <p>Results</p> <p>The constructed vector (pNZPnisA:CYTO-LLO) was expressed in <it>L. lactis </it>NZ9000 and was best induced at mid-log phase with 0.2% v/v nisin for 4 h statically at 30°C. Purification of the His-tagged LLO was accomplished by Ni-NTA affinity chromatography and functionality was confirmed through haemolytic assays. Total LLO yield (measured as total protein content) was 4.43–5.9 mg per litre culture and the haemolytic activity was still detectable after 8 months of storage at 4°C.</p> <p>Conclusion</p> <p>The LLO production method described in this work provides an approach to efficient LLO production in the Gram-positive <it>Lactococcus </it>bacterium to yield a significant source of the protein for research and diagnostic applications. Expression of LLO in <it>L. lactis </it>has a number of benefits over <it>E. coli </it>which may facilitate both <it>in vivo </it>and <it>in vitro </it>applications of this system.</p

The impact of the gut microbiota on drug metabolism and clinical outcome

The significance of the gut microbiota as a determinant of drug pharmacokinetics and accordingly therapeutic response is of increasing importance with the advent of modern medicines characterised by low solubility and/or permeability, or modified-release. These physicochemical properties and release kinetics prolong drug residence times within the gastrointestinal tract, wherein biotransformation by commensal microbes can occur. As the evidence base in support of this supplementary metabolic “organ” expands, novel opportunities to engineer the microbiota for clinical benefit have emerged. This review provides an overview of microbe-mediated alteration of drug pharmacokinetics, with particular emphasis on studies demonstrating proof of concept in vivo. Additionally, recent advances in modulating the microbiota to improve clinical response to therapeutics are explored

An analysis of the pharmacist workforce capacity in Ireland over the past 15 years

Background: The FIP 2018 global pharmacy workforce report identified that the pharmacy workforce is facing increasing capacity challenges and predicted that globally, the pharmacy workforce would grow by 40% over the next 15 years. The purpose of this report was to specifically examine the pharmacist workforce in Ireland over the last 15 years with reference to a number of developments that have both directly and indirectly affected the capacity of the pharmacy sector. A second objective was to benchmark the pharmacy workforce capacity in Ireland against other countries of similar demographic and economic standing in order to assess how Ireland compares. Methods: Data was collected from PSI Annual Reports, Eurostat, OECD, PHARMINE Reports, UCAS, CSO Census Reports, WHO and FIP Global Pharmacy Workforce Reports regarding the pharmacy workforce in Ireland and selected comparison countries. Comparison countries were chosen based on population, GDP/capita, healthcare expenditure and HDI values. The data was analysed and presented using graphs and tables. Results: The number of pharmacists in Ireland has increased by 90% over the last 15 years. Despite two new Schools of Pharmacy opening in 2002 (RCSI) & 2003 (UCC), 57% of new registrants to the PSI over the last 15 years qualified via the EU route, predominately from the UK. Since the first graduates from RCSI and UCC qualified, PSI registrants via the national route range between 27-56% of total additions annually. Ireland’s output of pharmacy graduates per population is 40% lower than the UK and the number of pharmacy graduates per school of pharmacy in the UK is over twice that of Ireland. Ireland has the second highest number of pharmacies per 100,000 population out of 10 comparator countries. Ireland also has the joint highest number of pharmacists per 100,000, based on the total number of pharmacists registered with the PSI. This includes 5.5 % of PSI registrants who are in non-patient facing roles, 2.5% who are not-practicing/other and 21.5% of registrants who do not state their area of practice, with only 70.5% of registrants declaring as ‘patient-facing’. Conclusions: The pharmacy workforce in Ireland is highly dependent on new registrants applying via the EU mutual recognition route, predominantly from the UK. Any interruption to mutual recognition of pharmacists between the EU and UK, as a result of Brexit, would significantly affect the capacity of pharmacy services in Ireland. Compared to similar EU countries, pharmacists/pharmacies per head of population is relatively high. However, estimates of pharmacy workforce in Ireland based on all pharmacists registered with the PSI may overestimate capacity in Ireland. In order to meet global trends of the increasing number of patients needing access to pharmacy related services and diversification of pharmacist roles, ongoing review of capacity in pharmacy is essential

Supersaturated lipid-based formulations to enhance the oral bioavailability of venetoclax

Increasing numbers of beyond Rule-of-Five drugs are emerging from discovery pipelines, generating a need for bio-enabling formulation approaches, such as lipid-based formulations (LBF), to ensure maximal in vivo exposure. However, many drug candidates display insufficient lipid solubility, leading to dose-loading limitations in LBFs. The aim of this study was to explore the potential of supersaturated LBFs (sLBF) for the beyond Rule-of-Five drug venetoclax. Temperature-induced sLBFs of venetoclax were obtained in olive oil, Captex® 1000, Peceol® and Capmul MCM®, respectively. A Peceol®-based sLBF displayed the highest drug loading and was therefore evaluated further. In vitro lipolysis demonstrated that the Peceol®-based sLBF was able to generate higher venetoclax concentrations in the aqueous phase compared to a Peceol®-based suspension and an aqueous suspension. A subsequent bioavailability study in pigs demonstrated for sLBF a 3.8-fold and 2.1-fold higher bioavailability compared to the drug powder and Peceol®-based suspension, respectively. In conclusion, sLBF is a promising bio-enabling formulation approach to enhance in vivo exposure of beyond Rule-of-Five drugs, such as venetoclax. The in vitro lipolysis results correctly predicted a higher exposure of the sLBF in vivo. The findings of this study are of particular relevance to pre-clinical drug development, where maximum exposure is required

Pharmacotherapy for neonatal seizures: current knowledge and future perspectives

Seizures are the most common neurological emergencies in the neonatal period and are associated with poor neurodevelopmental outcomes. Seizures affect up to five per 1000 term births and population-based studies suggest that they occur even more frequently in premature infants. Seizures are a sign of an underlying cerebral pathology, the most common of which is hypoxic-ischaemic encephalopathy in term infants. Due to a growing body of evidence that seizures exacerbate cerebral injury, effective diagnosis and treatment of neonatal seizures is of paramount importance to reduce long-term adverse outcomes. Electroencephalography is essential for the diagnosis of seizures in neonates due to their subtle clinical expression, non-specific neurological presentation and a high frequency of electro-clinical uncoupling in the neonatal period. Hypoxic-ischaemic encephalopathy may require neuroprotective therapeutic hypothermia, accompanying sedation with opioids, anticonvulsant drugs or a combination of all of these. The efficacy, safety, tolerability and pharmacokinetics of seven anticonvulsant drugs (phenobarbital, phenytoin, levetiracetam, lidocaine, midazolam, topiramate and bumetanide) are reviewed. This review is focused only on studies reporting electrographically confirmed seizures and highlights the knowledge gaps that exist in optimal treatment regimens for neonatal seizures. Randomised controlled trials are needed to establish a safe and effective treatment protocol for neonatal seizures

Assessing awareness and attitudes of healthcare professionals on the use of biosimilar medicines: A survey of physicians and pharmacists in Ireland

Increasing numbers of biosimilar medicines are becoming available. The objective of this survey was to assess awareness of and attitudes to biosimilars amongst physicians (medical specialists and General Practitioners (GPs)) and community pharmacists in Ireland. Physicians were invited to complete an online questionnaire during April and May 2016. Community pharmacists received a postal questionnaire in August 2015. Responses from 102 medical specialists, 253 GPs and 125 community pharmacists were analysed. The majority of medical specialists (85%) and pharmacists (77%) claimed to be either very familiar or familiar with the term biosimilar, whereas many GPs (60%) were unable to define or had never heard of the term. One in five (21%) healthcare professionals responded that biosimilars were the same as generic medicines. The majority of medical specialists opposed pharmacist-led substitution of biological medicines but some thought it could be appropriate if agreed with the clinician in advance. Medical specialists who prescribe biosimilars (n = 43) were more likely to do so on treatment initiation (67%), than switch a patient from an originator medicine to a biosimilar (28%). The findings will aid the design of educational initiatives for healthcare professionals and highlight attitudes of healthcare professionals to biosimilars, so informing regulators, policy makers and industry

Machine learning methods for prediction of food effects on bioavailability: A comparison of Support Vector Machines and Artificial Neural Networks

Despite countless advances in recent decades across various in vitro, in vivo and in silico tools, anticipation of whether a drug will show a human food effect (FE) remains challenging. One means to predict potential FE involves probing any dependence between FE and drug properties. Accordingly, this study explored the potential for two machine learning (ML) algorithms to predict likely FE. Using a collated database of drugs licensed from 2016-2020, drugs were classified into three groups; positive, negative or no FE. Greater than 250 drug properties were predicted for each drug which were used to train predictive models using Support Vector Machine (SVM) and Artificial Neural Network (ANN) algorithms. When compared, ANN outperformed SVM for FE classification upon training (82%, 72%) and testing (72%, 69%). Both models demonstrated higher FE prediction accuracy than the Biopharmaceutics Classification System (BCS) (46%). This exploratory work provided new insights into the connection between FE and drug properties as the Octanol Water Partition Coefficient (S+logP), Number of Hydrogen Bond Donors (HBD), Topological Polar Surface Area (T_PSA) and Dose (mg) were all significant for prediction. Overall, this study demonstrated the utility of ML to facilitate early anticipation of likely FE in pre-clinical development using four well-known drug properties

Perspectives of pharmacists on facilitating experiential learning placements for pharmacy students in non-patient facing settings

Full text available via ShareLink until August 07 2020: https://authors.elsevier.com/a/1bFq96gprIBGQ%7EIntroduction: Recently, the model of pharmacy education in Ireland changed to a five-year pharmacy degree, with three distinct blocks of experimental placements dispersed throughout the degree. The United Kingdom is also considering the introduction of a similar five-year pharmacy degree, while the United States is looking to further expand non-clinical experiential learning opportunities. This study was carried out to ascertain the perspectives of pharmacists working in non-patient facing roles on the barriers to and facilitators of placements to aid in identifying placement recruitment strategies for non-patient facing placements. Methods: A questionnaire was distributed to pharmacists employed in non-patient facing settings, including in pharmaceutical industry, education, and regulation. Quantitative responses were analyzed using descriptive statistics, while qualitative questions were analyzed thematically. Results: Regardless of experience in the practice setting or supervision, the majority expressed a preference for offering paid placements of six months' duration. There was divided opinion regarding whether students should be given study leave, whether the student's supervisor should be a pharmacist, and whether students should undertake specialized postgraduate training. The main barriers to placements were time, the placement structure, availability of suitable projects or supervisors, and awareness of placement opportunities. Prior experience in the practice area, developing the talent pipeline, and personal interests were all placement facilitators. Conclusions: Given the increasing roles for pharmacists in non-patient facing practice settings, this study highlights the importance of stakeholder involvement during the implementation of a new model of education to ensure that placements in all settings are feasible

Regulatory Science Ireland: bridging the information gap on biosimilar medicines

Regulatory Science Ireland (RSI) is a voluntary network of interested parties from academia, the Health Products Regulatory Authority (HPRA), pharmaceutical and medical device industries and government agencies. RSI is conducting a research project, the objective of which is to enhance understanding of biosimilar medicines amongst stakeholders and encourage best practice in the use of these medicines