Design, synthesis and evaluation of advanced polymeric taxane formulations

Hypersensitivity reaction upon intravenous administration of taxane medicines still severely affects the quality of life of patients in treatment with Taxol or Taxotere. Abraxane and Genexol-PM have been able to significantly lower or even exclude allergic reaction caused by surfactants such as Cremophor EL and polysorbate 80, used in Taxol and Taxotere, respectively. However, the manifested intrinsic systemic toxicity of chemotherapeutics including taxanes remains a major issue in conventional chemotherapy and shows that further exploration towards alternative formulations is required for pursuing more efficient and safer administration. Polymer-based drug formulations are attractive systems due to the chemical versatility and low-cost of polymeric carrier material compared to biotechnological carriers such as solubilizing proteins and monoclonal antibodies. As physical entrapment and chemical conjugation have already shown great promise in the development of second-generation polymeric chemotherapeutics, the overall aim of this thesis was to develop advanced polymeric taxane formulations based on these formulation strategies

Acrylamides with hydrolytically labile carbonate ester side chains as versatile building blocks for well-defined block copolymer micelles via RAFT polymerization

En route towards improved delivery systems for targeted chemotherapy, we propose a straightforward approach for the hydrophobic modification of the acrylamide N-(2-Hydroxyethyl) acrylamide (HEAm). An ethyl or benzyl group was introduced via a hydrolytically sensitive carbonate ester yielding HEAm-EC and HEAm-BC, respectively. Block copolymers of HEAm, respectively PEG and HEAm-EC or HEAm-BC were successfully synthesized by reversible addition-fragmentation chain transfer (RAFT) polymerization, obtaining a library of well-defined block copolymers with different degrees of polymerization (DP). To further explore the versatility of our approach in terms of polymer synthesis, self-assembly, drug solubilization and in vitro cell interaction, polyethylene glycol (PEG) and polyHEAm as hydrophilic polymer blocks were compared. The block copolymers formed micellar nanoparticles (10-100 nm) in PBS and could efficiently solubilize hydrophobic dyes and anti-cancer drugs. Benzyl carbonate ester side chains increased micellar stability and drug loading capacity. Moreover, PEG as hydrophilic block showed in comparison to HEAm more promising results concerning both colloidal stability and drug loading capacity. Confocal microscopy showed that the micelles could efficiently deliver a hydrophobic dye inside the cells. Finally, we also demonstrated efficient formulation of the anti-cancer drug paclitaxel with an in vitro cancer cell killing performance comparable or even better than the two commercial PTX nano-formulations Abraxane and Genexol-PM at equal drug dose. In conclusion, modification of HEAm through carbonate linkages offers a versatile platform for the design of degradable polymers with potential for biomedical applications

pH-Degradable mannosylated nanogels for dendritic cell targeting

We report on the design of glycosylated nanogels via core-cross linking of amphiphilic non-water-soluble block copolymers composed of an acetylated glycosylated block and a pentafluorophenyl (PFP) activated ester block prepared by reversible addition fragmentation (RAFT) polymerization. Self-assembly, pH-sensitive core-cross-linking, and removal of remaining PFP esters and protecting groups are achieved in one pot and yield fully hydrated sub-100 nm nanogels. Using cell subsets that exhibit high and low expression of the mannose receptor (MR) under conditions that suppress active endocytosis, we show that mannosylated but not galactosylated nanogels can efficiently target the MR that is expressed on the cell surface of primary dendritic cells (DCs). These nanogels hold promise for immunological applications involving DCs and macrophage subsets

Sequence-selective DNA recognition and enhanced cellular up-take by peptide–steroid conjugates

Several GCN4 bZIP TF models have previously been designed and synthesized. However, the synthetic routes towards these constructs are typically tedious and difficult. We here describe the substitution of the Leucine zipper domain of the protein by a deoxycholic acid derivative appending the two GCN4 binding region peptides through an optimized double azide–alkyne cycloaddition click reaction. In addition to achieving sequence specific dsDNA binding, we have investigated the potential of these compounds to enter cells. Confocal microscopy and flow cytometry show the beneficial influence of the steroid on cell uptake. This unique synthetic model of the bZIP TF thus combines sequence specific dsDNA binding properties with enhanced cell-uptake. Given the unique properties of deoxycholic acid and the convergent nature of the synthesis, we believe this work represents a key achievement in the field of TF mimicry

How semantic searching can improve ROI of pharmaceutical projects

As part of our Master in Financial Management program at Vlerick Business School (Belgium) we - Dr. Benoit Louage, Trevor Hougen and Tarquin Blancquaert - had the opportunity to perform an In-Company Project on behalf of ONTOFORCE. ONTOFORCE is a young company developing a data analytics platform for application in the biotechnology and pharmaceutical sectors, named DISQOVER. By way of its semantic search technology, the database/platform seeks to maintain all individual forms of research-relevant data under one scalable cloud architecture. Overall, this report highlights the disruptive potential of 'Big Data' and semantic search technology for the pharmaceutical industry. As the latter is currently still experiencing a decline in R&D productivity, several strategies should be investigated in order to improve the drug development processing. Our main objective was to quantify the impact of (linked) data on R&D processes of Pharmaceutical and Biotech companies. More precisely we had to quantify the potential of ONTOFORCE's semantic search platform DISQOVER within R&D processes. Eventually we came up with a valuation model, which calculates the ROI (return on investment) and 'Wins per Day' of the DISQOVER platform towards its users. After conducting extensive research and interviewing many experts within the field, we believe that the model we created is well-thought-out and the assumptions are substantiated by expert opinions and academic literature. In addition, the model is highly flexible, providing the opportunity for users to fill in their own input regarding R&D, financial and market assumptions. In conclusion, the results of this project were positive, quantifiable and derivable, which was especially important considering the original objectives of the project. Considering these results, it is safe to note that the outcomes of quantifying ROI and 'Wins per Day' were attained with supportable assumptions. Our results are clearly reflecting the high potential of the platform. It should be noted that these results are still preliminary outcomes. Further integration of this tool by pharmaceutical companies' R&D projects will allow rendering the model outcomes more accurate as the latter will increasingly provide real company-originated input data (e.g. cycle time reductions). It is noteworthy that during our research we encountered two major future opportunities that would further lift the DISQOVER platform to a higher level. Firstly, there was a clear demand for a platform that starts to learn the user's movements and begins to recommend searches and bring relevant information to the reader through understanding of the searching behaviour. The truly valuable databases such as DISQOVER are those that can, besides allowing the user to search information, also send relevant information to the user based on its searching pattern and interests. In this way the user stays up-to-date at any time and can immediately anticipate on progresses made in the field. Secondly, further insights lent to us are that the ability of the platform to connect institutional knowledge based on clinical trial population would also be key, in order to pre-determine relevant populations for clinical trials, which is very time consuming. Finally, we would like to mention that throughout our project we could at any time rely on the best assistance from Hans Constandt (CEO and founder of ONTOFORCE) and Dr. Filip Pattyn (Scientific Lead at ONTOFORCE), and the knowledge of Walter Van Dyck (Associate Professor and Partner at Vlerick Business School). For this, we are very grateful towards them. Despite some difficulties we came across, we look back on the project as being very challenging and instructive, resulting in a very satisfying final result

Poly(glycerol sebacate) nanoparticles for encapsulation of hydrophobic anti-cancer drugs

Physical encapsulation of hydrophobic compounds into nanocarriers that are stable in aqueous medium is of high interest as it can increase solubilization of the drug, lower its toxicity, control its pharmacokinetic profile and thus overall improve the therapeutic efficacy. To increase solubilization of a drug in aqueous medium, the carrier should contain hydrophobic domains that can form non-covalent interactions with hydrophobic drug molecules. Apart from liposomes, polymers have been widely acknowledged as promising nanocarriers. In this paper, we report the design of poly(glycerol sebacate) (PGS), an inexpensive, water insoluble but biodegradable and biocompatible polymer, into nanocarriers for hydrophobic drugs. Mixing of alcoholic PGS solutions with water (i.e. solvent displacement) produced a fine and highly stable dispersion with a size that can be controlled by the PGS concentration and solvent to water ratio. These dispersions were used for the encapsulation of hydrophobic compounds such as a fluorescent dye and two drugs known for their anti-mitotic activity (i.e. paclitaxel (PTX) and flubendazole (FLU)). These formulations were then evaluated in vitro with cancer cells

Dual pH- and temperature-responsive RAFT-based block co-polymer micelles and polymer-protein conjugates with transient solubility

Via a smart combination of temperature-responsive and acid labile acetal monomers, copolymers are obtained with a la carte lower critical solution temperature behavior. RAFT copolymerization of these monomers using, respectively, a PEG-functionalized or amine-reactive NHS-functionalized chain transfer agent allows designing of micelles and polymer-protein conjugates with transient solubility properties within a physiologically relevant window

Stapling monomeric GCN4 peptides allows for DNA binding and enhanced cellular uptake

The basic DNA recognition region of the GCN4 protein comprising 23 amino acids has been modified to contain two optimally positioned cysteines which have been linked and stapled using crosslinkers of suitable lengths. This results in stapled peptides with a stabilized alpha-helical conformation which allows for DNA binding and concurrent enhancement of cellular uptake