Browsing by Title

Patient safety is a part of quality and safety of care. Patient safety is defined as freedom for a patient from unnecessary harm or potential harm associated with healthcare. Patient safety covers safety of care, medication safety and safety of devices. Different authorities have promoted patient safety in Finland. The Ministry of Social Affairs and Health set up the Steering Group for the Promotion of Patient Safety for a term extending from 1 November 2006 to 31 October 2009 to coordinate the work for promoting patient safety and to evaluate related development needs at the national level. It has published a national Patient Safety Strategy for the years 2009-2013. Patient safety is also included in the Health Care Act. The National Institute for Welfare and Health (THL) has a Finnish national programme on patient safety: Patient Safety with Skills. Other authorities promoting patient safety in Finland are Finnish Medicines Agency (Fimea) and National Supervisory Authority for Welfare and Health (Valvira). Many studies are related to Patient Safety. In the Seminar of Patient Safety Research 2011 studies were separated to following categories: Patient Safety Culture as a Challenge for Organisations, Medication Safety, Safeguard of Care, Preparation for Patient Safety, Learning of Patient Safety, Control of Patient Documents and Financing of Patient Safety. The aim of this study is to explore Finnish patient safety studies. This study was conducted by using an electronic survey. The survey was sent to members of Finnish Patient Safety Society and a mailing list of Patient Safety Network. The survey was also sent to attendees of the Seminar of Patient Safety Research 2011. Altogether 81 responses were obtained. A patient safety research had been done in 60 per cent of organizations. A patient safety research will be done in 62 per cent of organizations. 10 per cent of the researches were meant for the internal use of the organization but were also published in Finland and abroad. 21 per cent of the researches were published in Finland and abroad. 18 per cent of the researches were published only in Finland and 12 per cent only abroad. 25 per cent of the researches were meant only for the internal use of organization. 14 per cent of the respondents left this question unanswered. A personal grant from a foundation was the most common way of financing for patient safety research. Many different kinds of sponsors were also mentioned. There was co-operation between organizations in 58 per cent of researches. 86 per cent of respondents were interested in a network of patient safety researchers. Using of Reporting System for Safety Incidents in Health Care Organizations (HaiPro) was asked as a detail of this study. HaiPro was used in 65 per cent of organizations. 89 per cent of respondents said that their organization takes advantage of HaiPro but the level of use varied between respondents.

Physiologically based pharmacokinetic modelling (PBPK) can be used to predict pharmacokinetic behaviour of new drug molecules in human. PBPK model represents the body anatomically and physiologically with compartments connected to each other and combines those to drug specific parameters. PBPK modelling can be used to predict the absorption, disposition, and time-concentration profiles of drug molecules. The purpose of the study was to build a PBPK model for new drug molecule under research (compound A) and predict pharmacokinetics in human, to support the selection of dosing interval, formulation, and sampling time points for the first clinical trial. In this work it is described the building of the model in the ”bottom-up”-approach using in vitro parameters in GastroPlusTM-software. The modelling was done also for preclinical species (mouse, rat, dog) comparing the simulations to the observed in vivo data, which gave the confidence to the methods used in the modelling also for human. The model was first built for systemic kinetics and thereafter it was used for predicting pharmacokinetics after oral dosing. Parameters of systemic kinetics were compared also to the predictions from allometric scaling. Based on the preclinical species the most predictive method for the volume of distribution of compound A was the method by Lukacova, which predicted the volume of distribution to be moderate in human (1.7 l/kg). From the in vitro-to-in vivo -extrapolation methods the most predictive method to predict the clearance was the method by Poulin, which predicted low clearance in human (8.1-14.3 l/h). Empirical scaling factors based on the preclinical data were not needed, as the models predicted well the observed in vivo data. Allometric methods predicted the systemic kinetic parameters to be in the similar range. Advanced compartmental absorption transit -model (ACAT) integrated to GastroPlusTM-software predicted the absorption after oral dosing well in the preclinical species (predicted/observed ratio 0.8-1.3 for systemic exposure) despite the low solubility of the compound A. The model predicted the absorption in human to be sensitive to particle size and absorption rate to be clearly affected by the particle size. The feeding status was also predicted to affect on the absorption with larger particle sizes. The gut metabolism was not predicted to limit the oral exposure notably, whereas moderate bioavailability was predicted to be achievable. Compound A could be given in a capsule if the target particle size distribution could be achieved. The built PBPK-model can be used in the future to predict the first clinical doses by comparing the predicted plasma concentrations to in vitro pharmacodynamic parameters and to the plasma concentrations needed for efficacy in the pharmacodynamic models. The model can also be used to predict the drug-drug interactions.

Traditional 2D cell cultivating vessels and experimental models cannot often simulate natural chemical and physical environment of different cell types. For example, availability of oxygen, chemical gradients, messaging molecules, fluid pressure, flow and surface topography are factors that may affect significantly in cell differentiation, growth, cellular structure, and metabolism. Modular bioreactors like Quasi-Vivo® -system can be used to simulate these factors. Liposomes are particles of phospholipid bilayer with aqueous space enclosed within. They can be modified in numerous ways, like loading them with hydrophobic and hydrophilic molecules, changing their transition temperature or coating them according to different needs. Doxorubicin is effective and widely used cytostatic agent, but when administered as a free drug it has often severe side-effects, like cardiotoxicity. Goal of this thesis is to determine appropriate manufacturing parameters and verify adequate shelf-life of ICG-Doxorubicin liposomes, that they are applicable for future in vitro experiments. Then survival of HepG2 cell line under flow in Quasi-Vivo®-equipment is determined, after which A549 and HepG2 will be then combined into one two-cell model. Finally, a simple illumination experiment in this cell model with previously made liposomes is conducted, and the effect in whole system is examined. Using protocol presented in this thesis it is possible to produce successfully and repeatedly liposomes with both ICG and doxorubicin encapsulation over 70%. Their shelf-life was at least 14 days when stored in 4°C protected from light. This was determined to be sufficient for in vitro testing. Cultivating A549 and HepG2 cell lines combined in the same system with shared media and fluid flow conditions was successful. Neither of the cell lines show significant difference in viability when compared to static control. When light-activating liposomes are administered to the system and then illuminated, from preliminary results we can see significant difference in drug effect. Both illuminated chambers and off-target chambers connected via Quasi-Vivo® show increased suppression, which shows promise that this in vitro model would be useful for future experiments.

Prolyl oligopeptidase (PREP) is a serine protease which is extensively present in the mammalian system and especially abundant in the brain. Despite the long research history of PREP its physiological function has remained unclear. PREP has been suggested to regulate the functions of many bioactive peptides by hydrolysis and on the other hand to participate in several intracellular processes probably via direct protein-protein interactions. One of the functions suggested for PREP is the regulation of the brain neurotransmitter systems and based on, for instance, the location in the brain PREP has been connected to both excitatory and inhibitory neurotransmitter systems. The literature review of this thesis first describe the brain neurotransmitter systems associated to PREP in general with some examples of diseases related to their malfunctions. In addition the structure of PREP and its location in the brain, both subcellular and cellular levels, and in distinct neurotransmitter systems, are presented, after which the different proposed functions for PREP are reviewed. The aim of the experimental part of this thesis was to investigate the effects of PREP on the brain neurotransmitter concentrations in the mouse nigrostriatal pathway and also to mouse motor behavior. The main research methods were microdialysis, tissue assays and cylinder test. The study was composed of two sections with five week duration each. The first section was performed with wild-type mice expressing naturally PREP and the second section with PREP-knockout (ko) mice and their wild-type littermates. The mice were injected unilaterally above the substantia nigra with adeno associated (AAV1) hPREP viral vector or with AAV1-eGFP (green fluorescent protein) viral vector as a control treatment. The cylinder test was carried out before the injection, and two and four weeks afterwards. Microdialysis was used to study the actions of PREP on the extracellular concentrations of dopamine (DA) and its metabolites dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), gamma-aminobutyric acid (GABA) and 5-hydroxyindoleacetic acid (5-HIAA), the major metabolite of serotonin (5-HT). In addition to the baseline assay the concentrations were measured after two amphetamine treatments (10 and 30 µM) administered via the microdialysis probe. The probe guide cannulas were inserted to mice striatums 1-2 weeks before the microdialysis measurement. In the end of the experiment the tissue concentrations of DA, DOPAC, HVA, 5-HT and 5-HIAA were measured from striatum and substantia nigra. Both the microdialysis and tissue sample concentrations were quantified with high performance liquid chromatography. In the first study section the PREP enzyme activity was also determined from striatum. Neither the complete deprivation nor over-expression of PREP in the nigrostriatal pathway had clear or consistent effects on the levels of neurotransmitters studied when compared to naturally occurring PREP expression. When comparing the differences between control treated groups of PREP-ko and littermate mice, a greater amphetamine stimulated DA-levels was seen in the former group proposing negative regulatory influence of PREP. In both study sections the tissue assay results were difficult to interpret due to observed responses also with AAV1-eGFP control treatment in comparison with untreated side of the brain. This was seen as a lower DA- and DOPAC-levels in substantia nigra and thus the meaning of the changes caused by PREP treatment is hard to comprehend. The results of the cylinder test may implicate some protective effect of the PREP-ko-genotype against viral vector injections in general. Then again the existence of compensatory mechanisms is possible when using knockout animals and thus the genotype differences are hardly ever unequivocal. The results of this thesis do not suggest outright regulatory effects of PREP on the neurotransmitter functions in the mouse nigrostriatal pathway although the confirmation of this requires further studies, especially in regard to GABAergic and glutamatergic systems. Studies should include a scale of different behavioral tests of motor activity and repeated microdialysis experiment with some defining method changes. The possible function and mechanisms of PREP as a regulator of neurotransmitter intake or release is rationale to study at molecular level with applicable methods.

Spray drying is one way to dry protein medicines and it has many advantages compared to other drying methods, for example it is a fast process. In spray drying high temperature and mechanical stress can inactivate the protein. Disaccharides are generally used as protective agents of protein in spray drying because they have an ability to protect the structure of the protein during drying and storage. Aim of this research was to study the stability of the protein during spray drying and storage by using β-galactosidace as a model protein. Aim was also to characterize the physical properties of trehalose and melibiose and to study how well they protect the protein. Some of the central matters to be examined were the glass transition temperature, crystallinity, water activity, yield of the spray dried powder and protein activity. Especially studying the properties of melibiose in spray drying was important because it has not been used before. The study also included the optimization of the process parameters to be suitable for the product. Trehalose and melibiose transformed to an amorphous form during spray drying. Both XRPD and DSC showed an amorfous form. Trehalose and melibiose proved to be good protective agents for the protein during spray drying and storatge probably because they remained their amorphous structure. β-galactosidase remained activity very well. Optimizing of the process parameters was successful because protein remained its activity and still the powder was quite dry and yield was good. The changes in the structure of the protein were studied with FT-IR but the amount of the protein was too small. Problems caused by the spray drier may have an effect to the results, but on the other hand the spray dryer was made to work optimally.

Lead molecule search is the first part of drug design. This process can be done using computerized docking of ligands into target proteins. Usually this requires expensive software and powerful computer systems specifically made for the process. There are however some programs that are available for free and can be run on home computers. The purpose of this Master's Thesis was to see how these free software can be used for the task of docking and also to create a method or a guideline for such work. Protein kinase C (PKC), a popular target for drug design, was chosen as a target of inhibitor design. PKC is part of a larger family of serine/threonine kinases and formed of 10 isoforms all with different effects on cellular functions. The large amount of related kinases and the similarities in their sequences make finding selective inhibitors a difficult process. Homology models of all PKC isoforms in three known conformations solved by x-ray diffraction (pdb: 1XJD, 2I0E and 3A8W) were created using Modeller. Into these models a set of possible ligands from the free database of molecules ZINC was docked using Autodock Vina utilizing a script created for docking multiple ligands into multiple targets. The dockings resulted in some interesting results. Six molecules were recognized as possible lead molecules for further research. None of these molecules had any patents or previous results of protein kinase inhibition connected to them. The most interesting result was the finding that coluracetam, a nootropic drug of the racetam family, might be a protein kinase inhibitor. Racetams are usually considered drugs that lead to PKC activation. It has been proposed that inhibitors may prolong the lifetime of kinases in the cells leading to increased activity in the long term. In our opinion coluracetam might prove to be a good tool for studying the complex way kinase activity is modulated. The methods and scripts used in this work will be released for free use.

In the United States pharmacists have prescribed medicines and managed patient's drug therapy since the 1970s, and in the United Kingdom pharmacists have been authorization to prescribe medications since 2003. The discussion about the right of Masters of Science in Pharmacy will be renewed prescriptions during the last decade in Finland but few Finnish studies have been published from the subject. In the document Medicines Policy 2020 published by Ministry of Social Affairs and Health states that by prescribing should be used cost-effective modes of operation. The knowledge about pharmacist prescribing benefits and costs, and also prescribing practice in Finland, is needed to evaluate the cost-effectiveness of pharmacists' authorization to prescribe and to support the decision-making concerning pharmacist prescribing. The aim of this master's thesis is to gather all existing knowledge about the economic and other effects of pharmacist prescribing using a systematic literature review method. The aim of theoretical part of this master's thesis is to explain the Finnish prescribing, the participation of pharmacists in drug therapy management in Finland and internationally pharmacist prescribing. The empirical part of this master's thesis is also to assess the quality of the studies of pharmacist prescribing benefits and costs using quality assessment checklists. In addition, this thesis describes the principles of the cost and benefit analyses, economic evaluations and systematic literature reviews. As a result of the literature search were found 1825 references. Based on the inclusion and exclusion criteria, 17 studies were selected to include in the systematic review. Of these studies three were economic assessments, 8 randomized controlled trials and 6 observational studies. The quality of these studies was assessed using four quality assessment checklists. On the basis of a systematic literature review pharmacist prescribing has been studied in the treatment of type 2 diabetes, hypertension, dyslipidemia, anticoagulation, chronic pain, emergency contraception and minor ailments and renewal of long-term medicines. Pharmacists reduced blood pressure by providing follow-up care with prescribing compared with the usual care, but not compared with the case management, which does not include prescribing. In addition, the follow-up care was to improve the treatment results of type II diabetes. The results obtained in the care of dyslipidemia were partly unclear. In the clinic follow-up care with prescribing could be reduced LDL-cholesterol, but not the risk of cardiovascular disease compared with the control group. In the pharmacy follow-up care had no effect on the treatment of patients with LDL-cholesterol compared with the control group. In addition, pharmacist prescribing improved how well patients stayed within INR target range. Pharmacist medication review with pharmacist prescribing achieved in the care of chronic pain patients differed few from the results of pharmacist medication review with feedback for a general practitioner. Pharmacist prescribing could reduce errors in inpatient medication compared with usual care. Much uncertainty is connected to the results of the study. The limited amount of studies, heterogeneity of the studies and methodological quality make the evaluation of real effects more difficult. The included studies of pharmacist prescribing were so heterogeneous. In addition treated disease, assessed benefits and scope of working environment were varied in included studies. Pharmacist prescribing was often studied as part of other care or pharmaceutical service, such as chronic disease management or medication review. The quality assessment of the included studies revealed several sources of bias. The available research information is the insufficient reliable evaluation of economic and other effects of pharmacist prescribing and the need for the further research is big.

There are dozens of bromotyrosine alkaloids extracted from the marine sponge Pseudoceratina purpurea. Bromotyrosines have for example antibacterial, antiviral and antitumor activity in vitro and therefore bromotyrosines are potential drug lead molecules. By far, bromotyrosines have been extracted from different marine sponges of the Verongida order for the use in biological activity research. Collecting marine sponges by scuba diving is neither fast nor ecological and therefore finding an effective synthesis strategy is vital so that the research could continue. One new bromotyrosine, purpurealidin I, was found from the marine sponge Pseudoceratina purpurea in the four year (2010-2014) Marex-project. The aim of this thesis was to synthesize the compound purpurealidin I and its derivatives. Synthesized compounds were tested against hepatitis C and chikungunya viruses. It is important to find new potent drug molecules, because approximately 350 000-500 000 people die from hepatitis C and there is no curative medication for the chikungunya. Purpurealidin I is synthesized from tyrosine and tyramine parts, which will be put together to form an amide bond in the final step of the synthesis. The synthesis of purpurealidin I was not completed during the Master's thesis. However there were two purpurealidin I derivatives and four purpurealidin I tyramine part derivatives that were successfully synthesized. One of the compounds is purpurealidin E, which can be extracted from the sponge Pseudoceratina purpurea. The t-Boc derivative of purpurealidin E was examined against hepatitis C and chikungunya and the compound showed moderate activity against hepatitis C virus, but it wasn't active against chikungunya virus. The original plan to synthesize purpurealidin I is possible, although some reactions and purification of crude products need to be optimized in order to get better yields. For the future research derivatives of the t-Boc derivative of purpurealidin E should be synthesized and studied against hepatitis C virus.

Pyrazoles are five-membered nitrogen containing heterocycles, whose derivatives can be widely used in medicinal chemistry. One of the most common ways to produce pyrazoles is 1,3-dipolar cycloaddition, where the dipole containing heteroatom is reacting with a dipolarophile, and forming a cycloadduct. Among others, mesoionic sydnones have been used as dipoles in 1,3-dipolar cycloadditions of pyrazoles. During the last decades solid phase methods, where either a dipole or dipolariphile is being temporarily bound to solid support, have been exploited in 1,3-dipolar synthesis. With the aid of solid phase methods, the synthesis can be controlled chemically by protecting groups that react easily. Also the isolation and purification can be made easier by using these techniques. The 1,3-dipolar solid phase methods can be combined with microwave techniques, to make the synthesis shorter and more effective. The goal of this work was to synthesize new N-unsubstituted pyrazoles, starting from sydnones bound to solid support and alkynes, with use of 1,3-dipolar cycloaddition reaction and to purify and analyze prepared compounds. The aim was also to develop a new 1,3-dipolar solid phase method so, that the end products could be cleaved easily in a traceless manner, and that there would be minor need for purification. There was also an aim to make the last step of the synthesis faster and easier with microwave reactor, and by using this method to standardize the conditions used in the cycloaddition-dehydration reaction step. The AMEBA-resin was chosen to be the solid support in this synthesis. Its traceless cleavage by trifluoroaceitic acid made possible not only the formation of the desired structure, but also effortless purification of the end product. The amino group of pyrazole was protected by the solid support during the N-nitrosation, so that after the traceless cleavage of the resin, N-unsubstituted pyrazoles were obtained. By changing the amino acids used in this synthesis, it was possible to alter the structure of the synthesized pyrazoles, and to create four structurally new pyrazoles. Microwave method used during the last step of synthesis for heating shortened the reaction step significantly, and also the yields of end products were better compared to conventional heating. During this work a functioning and developable 1,3-dipolar solid phase method was created, that can be utilized to synthesize pyrazoles and other compound of similar nature also the in future.

Pharmaceutical industry is supervised by several competent authorities. These authorities all over the world inspect manufacturers in order to make sure they comply with the Good Manufacturing Practice (GMP) guidelines and produce quality products. If non-compliance with the guidelines is detected, the authorities can revoke manufacturing licenses and deny access of the products. Recent trend in pharmaceutical industry is that the Active Pharmaceutical Ingredient (API) manufacturing is concentrated in few factories. If this kind of manufacturer is declared non-compliant and is therefore unable to supply an API, it can lead to drug shortages. This research aimed to find out what kind of quality problems occur in API manufacturing. Because of the concentration trend, it is important to understand what kind of problems the manufacturers do struggle with to prevent any risk for shortages. This research aimed also to determine how much the quality problems in API manufacturing can impact on drug shortages. Also, the number and location of these non-compliance cases were investigated. The chosen time frame was 2016-2018. Several databases were used as information sources in this research. These databases are maintained by the authorities in the U.S. and Europe and they contain information about the inspections and the GMP deficiencies they have found during these inspections. With the information collected from the databases, an inductive content analysis was conducted to determine the reasons for non-compliance with GMP in API manufacturing. Other information (e.g. locations, names of APIs) was also collected from the databases and analysed to answer the rest of the research questions. Results show that the biggest problem areas in API manufacturing were data integrity and analytical testing. Other problems relating to documentation occurred also. The amount of these cases was quite stable, and the relative proportion declined during the time period. Comparison between the list of APIs and drug shortage databases showed that even over 30% of the non-compliant APIs were later in shortage. The effect was greater in Finland than in the U.S. Therefore, it was concluded that the most significant GMP deficiencies in API manufacturing were poor data integrity and inappropriate analytical testing procedures. Secondly, the number of non-compliance cases in API manufacturing has not increased during this time, but these problems may have had an impact on drug availability problems.