An overview for the effects of lactitol, gelucire 44/14 and copovidone on lysozyme biological activity

Lysozyme, a model protein, was used to study the effects of excipients (lactitol, copovidone and gelucire 44/14) on the protein biological activity. Three different concentrations (0.1, 0.5 and 10 %w/v) of the excipients were used with a relatively high (10%w/v) lysozyme concentration. Analytical methods such as UV spectroscopy and fluorescence technique were used to study the effect of excipients on protein biological stability and integrity. The data obtained from analysis showed that gelucire 44/14 had a disadvantage effect on lysozyme stability as a single excipient. However, by combining gelucire 44/14 with lactitol, the protein activity was preserved. Also, the combination of lactitol and copovidone preserved the integrity and activity of 6% lysozyme to a great extent. Different excipients stabilised proteins to different degree. As conclusion, there is no excipient that works well for even one type of proteins. The most suitable excipient for each particular protein is found by extensive research studying the activity and stability of the protein in the presence of excipients. High efficacy and accuracy analytic methods should be used in order to give reliable information on protein stability and integrity. Keywords: lysozyme, lactitol, copovidone, gelucire 44/14, stability, biological activity

Effect of Spray-Drying and Electrospraying as Drying Techniques on Lysozyme Characterisation

The production of biopharmaceutical formulation incorporates several difficulties embracing their physical and chemical instabilities. In this study, two drying techniques, namely, spray-drying and electrospraying, were used to assess their application on lysozyme (as a model protein) without and with the use of betacyclodextrin. Samples were prepared in the ratio of 1:1 w/w (protein/betacyclodextrin), and several characterisation methods were applied to study the percentage (%) yield, morphology of the produced partials, thermal stability and biological activity of the protein. The results show the two drying methods led to different particle morphology as spherical-like shape was produced by spray-drying, while rodlike shape was generated by electrospraying with larger particle size. Lysozyme formulations produced by electrospraying were stable just directly after preparation, but after few weeks, those formulations showed visible aggregates. The biological activity of lysozyme was preserved by both drying techniques. In conclusion, both drying methods have different effects on the protein integrity and biological activity in which spray-drying shows more promising results

Liposomal drug delivery systems and anticancer drugs

Cancer is a life-threatening disease contributing to ~3.4 million deaths worldwide. There are various causes of cancer, such as smoking, being overweight or obese, intake of processed meat, radiation, family history, stress, environmental factors, and chance. The first-line treatment of cancer is the surgical removal of solid tumours, radiation therapy, and chemotherapy. The systemic administration of the free drug is considered to be the main clinical failure of chemotherapy in cancer treatment, as limited drug concentration reaches the tumour site. Most of the active pharmaceutical ingredients (APIs) used in chemotherapy are highly cytotoxic to both cancer and normal cells. Accordingly, targeting the tumour vasculatures is essential for tumour treatment. In this context, encapsulation of anti-cancer drugs within the liposomal system offers secure platforms for the targeted delivery of anti-cancer drugs for the treatment of cancer. This, in turn, can be helpful for reducing the cytotoxic side effects of anti-cancer drugs on normal cells. This short-review focuses on the use of liposomes in anti-cancer drug delivery

Electrically atomised formulations of timolol maleate for direct and on-demand ocular lens coatings

The file attached to this record is the author's final peer reviewed version. The Publisher's final version can be found by following the DOI link

The prognostic significance of lysosomal protective protein (Cathepsin A) in breast ductal carcinoma in situ

Background: Cathepsin A (CTSA) is a key regulatory enzyme for galactoside metabolism. Additionally, it has a distinct proteolytic activity and plays a role in tumour progression. CTSA is differentially expressed at the mRNA level between breast ductal carcinoma in situ (DCIS) and invasive breast carcinoma (IBC). In this study, we aimed to characterise CTSA protein expression in DCIS and evaluate its prognostic significance. Methods: A large cohort of DCIS (n=776 for pure DCIS and n=239 for DCIS associated with IBC (DCIS/IBC)) prepared as tissue microarray was immunohistochemically stained for CTSA. Results: High CTSA expression was observed in 48% of pure DCIS. High expression was associated with features of poor DCIS prognosis including younger age at diagnosis (less than 50 years), higher nuclear grade, hormone receptor negativity, HER2 positivity, high proliferative index and high hypoxia inducible factor 1 alpha expression. High CTSA expression was associated with shorter recurrence free interval (RFI) (p=0.0001). In multivariate survival analysis for patients treated with breast conserving surgery, CTSA was an independent predictor of shorter RFI (p=0.015). DCIS associated with IBC showed higher CTSA expression than pure DCIS (p=0.04). In the DCIS/IBC cohort, CTSA expression was higher in the invasive component than DCIS component (p less than 0.0001). Conclusion: CTSA is not only associated with aggressive behaviour and poor outcome in DCIS but also a potential marker to predict co-existing invasion in DCIS

Global, regional, and national burden of neurological disorders, 1990–2016 : a systematic analysis for the Global Burden of Disease Study 2016

Background: Neurological disorders are increasingly recognised as major causes of death and disability worldwide. The aim of this analysis from the Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2016 is to provide the most comprehensive and up-to-date estimates of the global, regional, and national burden from neurological disorders. Methods: We estimated prevalence, incidence, deaths, and disability-adjusted life-years (DALYs; the sum of years of life lost [YLLs] and years lived with disability [YLDs]) by age and sex for 15 neurological disorder categories (tetanus, meningitis, encephalitis, stroke, brain and other CNS cancers, traumatic brain injury, spinal cord injury, Alzheimer's disease and other dementias, Parkinson's disease, multiple sclerosis, motor neuron diseases, idiopathic epilepsy, migraine, tension-type headache, and a residual category for other less common neurological disorders) in 195 countries from 1990 to 2016. DisMod-MR 2.1, a Bayesian meta-regression tool, was the main method of estimation of prevalence and incidence, and the Cause of Death Ensemble model (CODEm) was used for mortality estimation. We quantified the contribution of 84 risks and combinations of risk to the disease estimates for the 15 neurological disorder categories using the GBD comparative risk assessment approach. Findings: Globally, in 2016, neurological disorders were the leading cause of DALYs (276 million [95% UI 247–308]) and second leading cause of deaths (9·0 million [8·8–9·4]). The absolute number of deaths and DALYs from all neurological disorders combined increased (deaths by 39% [34–44] and DALYs by 15% [9–21]) whereas their age-standardised rates decreased (deaths by 28% [26–30] and DALYs by 27% [24–31]) between 1990 and 2016. The only neurological disorders that had a decrease in rates and absolute numbers of deaths and DALYs were tetanus, meningitis, and encephalitis. The four largest contributors of neurological DALYs were stroke (42·2% [38·6–46·1]), migraine (16·3% [11·7–20·8]), Alzheimer's and other dementias (10·4% [9·0–12·1]), and meningitis (7·9% [6·6–10·4]). For the combined neurological disorders, age-standardised DALY rates were significantly higher in males than in females (male-to-female ratio 1·12 [1·05–1·20]), but migraine, multiple sclerosis, and tension-type headache were more common and caused more burden in females, with male-to-female ratios of less than 0·7. The 84 risks quantified in GBD explain less than 10% of neurological disorder DALY burdens, except stroke, for which 88·8% (86·5–90·9) of DALYs are attributable to risk factors, and to a lesser extent Alzheimer's disease and other dementias (22·3% [11·8–35·1] of DALYs are risk attributable) and idiopathic epilepsy (14·1% [10·8–17·5] of DALYs are risk attributable). Interpretation: Globally, the burden of neurological disorders, as measured by the absolute number of DALYs, continues to increase. As populations are growing and ageing, and the prevalence of major disabling neurological disorders steeply increases with age, governments will face increasing demand for treatment, rehabilitation, and support services for neurological disorders. The scarcity of established modifiable risks for most of the neurological burden demonstrates that new knowledge is required to develop effective prevention and treatment strategies. Funding: Bill & Melinda Gates Foundation

Effect of processing techniques and excipients on integrity and stability of protein formulations

Production of protein pharmaceutical formulations has been challenging due to the complexity of protein synthesis, purification and physical and chemical instabilities of the proteins. Accordingly, the aim of this research is to assess the effects of spray drying, freeze drying and crystallisation using different concentrations of four excipients (pluronic®F127, cremophor®EL, β-cyclodextrin and inulin) on protein integrity, thermal stability and biological activity. Lysozyme and Bovine serum albumin were chosen as model proteins. The oral administration of protein pharmaceuticals to the systemic circulation has numerous barriers, including, sharp pH gradients, photolytic enzymes and low epithelial permeability. Trying to overcome these barriers, proteins can be encapsulated in niosomes that are known to protect proteins against the surrounding environment. In this study, Pluronic® F-127 and cremophor® EL were used as co-surfactants for niosome preparations. Niosomes of Span™ 65: cholesterol: co-surfactant were prepared for encapsulation of proteins by film hydration method. Lysozyme and insulin were chosen as model proteins for noisome formulations. Unprocessed, spray-dried, freeze-dried and crystallised proteins were characterised by the following methods: thermal analysis using Differential Scanning Calorimetry (DSC), microscopic examination of the protein particles using Scanning Electron Microscopy and spectroscopic analysis employing Fourier Transform Infra-Red (FT-IR). Moisture contents were determined by Karl Fisher titration. Enzymatic assay were used to measure biological activity. The percentage yield of proteins and protein content were also determnined. Morphology and vesicular sizes of the prepared niosomes were investigated by Transmission Electron Microscopy (TEM) and Dynamic Light Scattering (ZetaPlus), respectively. The entrapment efficiency of protein in niosomes was determined by complete vesicle disruption using 50:50 isopropanol:buffer, followed by analysis of the resulting solutions by HPLC. Thermal behaviour of the niosomes was investigated using Differential Scanning Calometry (DSC). Protection of proteins against stimulated gastric fluid (SGF) stimulated intestinal fluid (SIF) were also assessed. Spray-dried and freeze-dried forms of proteins with pluronic® F127 better maintained the native conformation, as indicated by enzymatic assay, of lysozyme. The storage stability of unprocessed, spray-dried and freeze-dried lysozyme was studied over a period of 20 weeks at different temperatures. The results suggested that protein samples prepared with pluronic® F127 were more stable after storage. Crystallising lysozyme with (0.05%) of: pluronic®F-127, cremophor®EL and inulin significantly (P<0.05) maintained higher biological activity compared to a high concentration (0.2%) of the same excipients. However, using (0.2%) of ~cyclodextrin in crystallisation medium led to higher (P<0.05) protein activity compared to (0 .05%) β-cyclodextrin. The storage stability of unprocessed and crystallised protein samples, with and without excipients, showed that crystallisation of lysozyme with 0.05% (w/v) of pluronic• F-127 and 0.2% (w/v) of β-cyclodextrin better maintained the protein activity when compared to unprocessed and crystallised protein with and without excipients. The protein niosomes that prepared with different molar ratios of Span n.1 65, cholesterol and co-surfactants showed different encapsulation efficiency. For Span™ 65, cholesterol and cremophor® EL niosomes, they were not able to encapsulate any protein. Whilst, Span™ 65, cholesterol and pluron ic® F-127 formulations successfully produced insulin and lysozyme niosomes. For insulin containing niosomes, the ratio of 64.7 (Span™ 65): 32.3 (cholesterol): 3.0 (pluronic@ F-127) produced the highest protein encapsulation efficiency and the smallest vesicle size of O.74j..1m . For lysozyme containing niosomes, the maximum protein encapsulation was found in 72.75/24.25/3 .00% molar ratio of Span™ 651 cholesterol/pluronic®F-127 niosomes with vesicle size of 648.0IJm. The release study of proteins from the niosomal preparations in simulated gastric fluid (SGF) and simulated intestinal fiuid (SIF) revealed that insulin and lysozyme efflux from the niosomes is a biphasic process. The release of both proteins (insulin and lysozyme) from the niosomal formulations was found to be dependent on vesicle size, vesicle lamilarity, protein encapsulation, the position of the protein within the niosomal lamilarity and niosomal formulation component ratio . In conclusion , protein formulations using pluronic@F-127 increased protein stabilisation. Pluronic® F-127 is a good candidate to be used for protein formulations employing different processing techniques. The overall results show promise for protein drug delivery systems.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Route of monoclonal antibodies administration

Monoclonal antibodies are biotherapeutics and one of the distinguished class of medicines which are being extensively used for the treatment of life-threatening diseases such as cancer and various infectious diseases. However, their formulations are challenging due to the complex structure of those protein-type therapeutics. mAbs are administered via injections either intravenous (IV), infusion route (need hospitalization) for high bioavailability and rapid action, or subcutaneous (SC) route (self-administration by patients using specialized SC injection pens), or intravitreally for very few mAbs. The injection route is not convenient for patients. Therefore, pharmaceutical industries and scientists are trying to find an alternative for this painful injectable route via noninvasive routes like oral, transdermal, and inhalation routes. Consequently, novel delivery systems have been explored to optimize the final products of mAbs—to be adaptable by these novel routes—through either formulations' technology or fabrication of the mAb delivery systems. Therefore, this chapter will discuss various novel approaches (e.g., drying technologies and particulate systems) to solve challenges in achieving effective mAb delivery via noninvasive routes (more specifically oral, transdermal, and inhalation routes of drug administrations)