Our research focuses on developing advanced nanoparticles for cancer treatment, with a focus on reducing side effects and improving efficacy. We have developed nanoparticles that can not only... Show moreOur research focuses on developing advanced nanoparticles for cancer treatment, with a focus on reducing side effects and improving efficacy. We have developed nanoparticles that can not only target cancer cells but also serve as imaging agents to monitor treatment. In addition, we have investigated how lipids and immune cells in the tumor environment can be manipulated to improve cancer therapy.Our findings show that the developed nanoparticles can improve drug delivery to tumors while reducing side effects to the rest of the body. Moreover, we have shown that lipids can be used to reprogramme immune cells in the tumor environment, causing them to exhibit anti-tumor activity. In addition, the nanoparticles show excellent delivery capabilities in 3D models of skin cancer, allowing them to kill skin tumor cells more effectively.The societal relevance of our research lies in improving cancer treatment options, particularly by reducing side effects and increasing the effectiveness of therapies. These findings provide new insights and approaches for the development of cancer therapies, which can contribute to a better quality of life for patients.We have achieved these results through a multidisciplinary approach using nanotechnology, chemotherapeutic drugs and knowledge of the tumor environment. Our findings are special because they offer promising new strategies for cancer treatment using cutting-edge technologies and insights into tumor biology. Show less
Utilizing the polymeric platform of polypept(o)ides, this thesis describes synthesis and investigation of novel triblock copolymers to obtain carrier systems with multiple compartments for... Show moreUtilizing the polymeric platform of polypept(o)ides, this thesis describes synthesis and investigation of novel triblock copolymers to obtain carrier systems with multiple compartments for efficient siRNA delivery. Although the individual microstructure of nanoparticles differs depending on the polymeric building blocks, desired application and cargo, the final nanoparticles always combine a polysarcosine (pSar) shell with a polypeptide core, providing the ability of siRNA complexation by a polycationic segment. In addition, a third block enabled either covalent cross-linking, hydrophobic / π- π-stacking mediated stabilization or co-encapsulation of small hydrophobic drugs. Broadening the structural variety of such polypept(o)ides, a novel synthetic procedure was introduced to access AA'B- and ABC-type miktoarm star polymers.Investigations have been dedicated to the design of novel polymeric structures based on polypept(o)ides, to improve the delivery of siRNA by Polyion Complex Micelles (PICMs), provide access to different polymeric architectures, and to establish novel synthetic methods for the synthesis of these materials. Covering aspects from the synthesis of novel polymeric species up to advanced drug delivery strategies for siRNA in vivo, developments throughout this thesis extent the accessibility of the polypept(o)ide platform for nucleic acid delivery, highlight their potential in nanomedicine and further elaborate delivery strategies for next-generation nanomedical applications. Show less
The intradermal administration route is attractive for vaccine delivery, because of the highly populated antigen-presenting cells in the skin. However, the skin’s main physical barrier, the stratum... Show moreThe intradermal administration route is attractive for vaccine delivery, because of the highly populated antigen-presenting cells in the skin. However, the skin’s main physical barrier, the stratum corneum, prevents vaccines from entering the skin. To overcome this barrier, dissolving microneedle arrays (dMNAs) have been developed, miniaturized needles that enter the skin without reaching the blood vessels and pain receptors. After entering, they dissolve and leave the vaccine in the skin, leaving no sharps waste behind. The dissolution is achieved effectively because dMNAs are composed of sugar or biodegradable polymers. For vaccination purposes, such microneedles have another huge potential, as antigens are stored in a dry form that improves thermal stability. This includes that dMNAs ensure antigen stability at room- and elevated temperatures, thereby circumventing the cold chain requirement for storage. Furthermore, since dMNAs don't require qualified medical personnel for application, self-administration is feasible.Currently, the most frequently used fabrication approach (centrifugation method) results in major antigen waste. As antigens are generally very expensive, the current method is not suitable for the cost-effective fabrication of dMNAs. Therefore, a novel dMNA production technology that lowers antigen loss is required. In the thesis, the engineering of a novel dMNA production system based on a dispensing approach that minimizes antigen loss was presented. Furthermore, a comparison was made between the two production approaches (centrifugation and dispensing methods) for vaccine delivery via dMNAs. In additional studies, this novel method has been used to deliver influenza vaccine into the skin leading successfully to a high and protective immune response. In conclusion, a dispensing system for dMNAs production, as opposed to the centrifugation process, is a promising method for the cost-effective fabrication of dMNAs. Show less
Antimicrobial peptides (AMPs) are promising alternatives to antibiotics for treatment of antimicrobial resistant (AMR) bacterial infections. However, their narrow therapeutic window due to in vivo... Show moreAntimicrobial peptides (AMPs) are promising alternatives to antibiotics for treatment of antimicrobial resistant (AMR) bacterial infections. However, their narrow therapeutic window due to in vivo toxicity and limited stability hampers their clinical use. Here, we evaluated encapsulation of two amphiphilic AMPs, SAAP-148 and snake cathelicidin Ab-Cath, into oleyl-modified hyaluronic acid (OL-HA) nanogels to improve their selectivity index. The AMP-loaded OL-HA nanogels ranged 181–206 nm in size with a PDI of 0.2, highly negative surface charge (−47 to −48 mV) and moderate encapsulation efficiency (53–63%). The AMP-loaded OL-HA nanogels displayed similar activity in vitro as AMP solutions against AMR Staphylococcus aureus and Acinetobacter baumannii, with a dose-dependent effect over time. Importantly, the AMP-loaded OL-HA nanogels showed decreased cytotoxicity towards human erythrocytes and primary skin fibroblast, thereby improving the selectivity index of SAAP-148 and Ab-Cath by 2- and 16.8-fold, respectively. Particularly, the selectivity of Ab-Cath-loaded OL-HA nanogels has great clinical potential, with an index that reached ≥ 300 for S. aureus and ≥ 3000 for A. baumannii. These findings indicate that OL-HA nanogels are a promising drug delivery system to reduce the cytotoxicity of AMPs without substantially affecting their antimicrobial activity, thereby increasing their selectivity index and potential as therapeutics to combat AMR bacterial infections. Show less
Gent, M.E. van; Kiodsinska, S.N.; Severin, M.; Ali, M.; Doodewaerd, B.R. van; Bos, E.; ... ; Nibbering, P.H. 2023
The antimicrobial peptide Ab-Cath, is a promising candidate for development as treatment for antimicrobial resistant (AMR) bacterial infections. Future clinical use is hampered by Ab-Cath's... Show moreThe antimicrobial peptide Ab-Cath, is a promising candidate for development as treatment for antimicrobial resistant (AMR) bacterial infections. Future clinical use is hampered by Ab-Cath's cationic peptidic nature and limited therapeutic window. Here, we evaluated hyaluronic acid-based nanogels for encapsulation of Ab-Cath to circumvent these limitations. Using microfluidics, monodispersed anionic nanogels of 156–232 nm encapsulating >99 % Ab-Cath were prepared. Unprecedented, lyophilization using polyvinyl alcohol and dextran-40 provided Ab-Cath nanogel protection and allowed easy dose adjustment. Lyophilized and redispersed Ab-Cath nanogels were as effective as Ab-Cath solution in killing AMR Staphylococcus aureus, Acinetobacter baumannii and Escherichia coli in biological fluids, and in reducing S. aureus and A. baumanniibiofilms. Importantly, encapsulation of Ab-Cath in nanogels reduced Ab-Cath's cytotoxic effects on human fibroblasts by ≥10-fold. Moreover, cutaneous application of Ab-Cath nanogels eliminated bacteria colonizing 3D human skin. These findings affirm the use of nanogels to increase the selectivity index of antimicrobial peptides. Show less
Bacterial infections are becoming harder-to-treat with current antibiotics, due to antimicrobial resistance and biofilm-formation. Therefore, there is an urgent need for novel antibacterial agents... Show moreBacterial infections are becoming harder-to-treat with current antibiotics, due to antimicrobial resistance and biofilm-formation. Therefore, there is an urgent need for novel antibacterial agents and antimicrobial peptides (AMPs) may fulfill this role. Herein, three strategies were explored for optimization of our lead AMP SAAP-148 to combat bacterial infections: i) chemical lead-optimization, ii) combination therapy with other antimicrobial agents and iii) innovative AMP delivery systems. The latter strategy was also applied to another promising AMP, the snake cathelicidin Ab-Cath. First, we demonstrated that conjugation of short polyethylene glycol chains to SAAP-148 reduced the peptide’s cytotoxicity and remarkably improved its ability to modulate the immune system to a more pro-inflammatory subset. Second, it was shown that combinations of SAAP-148 and novel antibiotic halicin were more effective than single agent treatment against planktonic bacteria of specific resistant bacterial strains, also in clinically relevant cell models. Third, we demonstrated that hyaluronic acid-based nanogels allow for efficient encapsulation of SAAP-148 and Ab-Cath, thereby improving the selectivity index of both peptides by maintaining antimicrobial activities against resistant bacteria and reducing cytotoxic activities against mammalian cells. Thus, the findings described in this thesis contribute to the development of SAAP-148 and Ab-Cath as therapeutics to combat bacterial infections. Show less
Diseases and injuries related to bone and cartilage severely affect the function of the musculoskeletal system. They affect the life quality of patients and make them unable to move. The... Show moreDiseases and injuries related to bone and cartilage severely affect the function of the musculoskeletal system. They affect the life quality of patients and make them unable to move. The peculiarities of bone and cartilage structures mean that the therapeutic and surgical treatments that have been developed till date have a lot of disadvantages and limitations. Consequently, the development of new strategies for administering drugs that favor localized delivery is very important to achieve increasingly better treatments.The design and preparation of platforms for this purpose is a constant challenge, and for this reason three types of matrices were designed and prepared in the thesis. The drugs encapsulated in these matrices were cefazolin sodium, betamethasone, dexamethasone sodium phosphate, and Etanercept (anti-TNF-α drug). We were able to show that our matrices modified the release of the incorporated molecules and the composition of the prepared materials played an important role in the kinetics of the release process. Physicochemical characterizations were carried out on all the materials, obtaining that in each case the morphology varied once modifications in the matrix composition had been introduced. Biological studies in vitro and in vivo showed the cytocompatibility of the materials and also the ability to help repair damaged cartilage. Show less
Background Extracellular vesicles (EVs) have shown great potential for targeted therapy, as they have a natural ability to pass through biological barriers and, depending on their origin, can... Show moreBackground Extracellular vesicles (EVs) have shown great potential for targeted therapy, as they have a natural ability to pass through biological barriers and, depending on their origin, can preferentially accumulate at defined sites, including tumors. Analyzing the potential of EVs to target specific cells remains challenging, considering the unspecific binding of lipophilic tracers to other proteins, the limitations of fluorescence for deep tissue imaging and the effect of external labeling strategies on their natural tropism. In this work, we determined the cell-type specific tropism of B16F10-EVs towards cancer cell and metastatic tumors by using fluorescence analysis and quantitative gold labeling measurements. Surface functionalization of plasmonic gold nanoparticles was used to promote indirect labeling of EVs without affecting size distribution, polydispersity, surface charge, protein markers, cell uptake or in vivo biodistribution. Double-labeled EVs with gold and fluorescent dyes were injected into animals developing metastatic lung nodules and analyzed by fluorescence/computer tomography imaging, quantitative neutron activation analysis and gold-enhanced optical microscopy. Results We determined that B16F10 cells preferentially take up their own EVs, when compared with colon adenocarcinoma, macrophage and kidney cell-derived EVs. In addition, we were able to detect the preferential accumulation of B16F10 EVs in small metastatic tumors located in lungs when compared with the rest of the organs, as well as their precise distribution between tumor vessels, alveolus and tumor nodules by histological analysis. Finally, we observed that tumor EVs can be used as effective vectors to increase gold nanoparticle delivery towards metastatic nodules. Conclusions Our findings provide a valuable tool to study the distribution and interaction of EVs in mice and a novel strategy to improve the targeting of gold nanoparticles to cancer cells and metastatic nodules by using the natural properties of malignant EVs. Show less
This thesis describes the design, characterization and application of dextran based crosslinked network. Dextran, a neutral and hydrophilic polysaccharide, was functionalized with maleimide or... Show moreThis thesis describes the design, characterization and application of dextran based crosslinked network. Dextran, a neutral and hydrophilic polysaccharide, was functionalized with maleimide or vinyl sulfone groups which are able to form covalent bonds with nucleophilic thiols via a thiol-Michael addition reaction. The functionalized dextran polymers are liable to be crosslinked to generate 3D networks. Novel drug delivery platforms, including dextran-albumin hydrogels and dextran-poly(ethylene glycol)-albumin hydrogels, were established in which human serum albumin acts simultaneously as an affinity-based drug carrier and crosslinker. The obtained drug delivery platforms showed high drug loading efficiency and demonstrated sustained release of drug molecules. This thesis also explores emerging applications of hydrogels. New method to form giant unilamellar vesicles was established by assisting the hydration process of a lipid film with a dextran-poly(ethylene glycol) hydrogel film. Using this approach, polymer and additive-free GUVs can be prepared rapidly in high yield under physiological ionic strength conditions. Show less
O-Nitrobenzyl groepen worden veel gebruikt als moleculaire beschermgroepen die je met licht kunt verwijderen. Deze zogenaamde photocages worden vaak toegepast in zowel de organische chemie als in... Show moreO-Nitrobenzyl groepen worden veel gebruikt als moleculaire beschermgroepen die je met licht kunt verwijderen. Deze zogenaamde photocages worden vaak toegepast in zowel de organische chemie als in de biologie. Photocaging blokkeert de functionaliteit van een molecuul en deze blokkade kan verwijderd worden door middel van licht met een specifieke golflengte. Dit proefschrift laat zien dat licht gebruikt kan worden om precieze controle te verkrijgen over waar en wanneer geneesmiddelenafgifte plaatsvindt. Show less
Silva, C.G. da; Peters, G.J.; Ossendorp, F.; Cruz, L.J. 2017
Proteins play a crucial role in life, taking part in all vital process in the body, and are therefore used as therapeutic agents in a diverse range of biomedical applications. When... Show more Proteins play a crucial role in life, taking part in all vital process in the body, and are therefore used as therapeutic agents in a diverse range of biomedical applications. When administrated into bodily fluids, most native proteins are prone to degradation or inactivation process. The challenges of protein delivery are overcoming poor stability, low permeability toward cell membrane. Among all existing materials for protein delivery, mesoporous silica nanoparticles (MSNs) are one of the most promising intracellular nanocarriers due to its key properties: biocompatible, straightforward synthesis, and surface modification. For various biomedical applications, monodisperse MSNs with a particle size in the 50-200 nm range,3 controllable surface chemistry,4 and a large pore size (> 5 nm) are desired. This thesis presents a new method to synthesize large disc-like pore (10 ± 1 nm) containing MSNs with an elongated cuboidal-like geometry (90 × 43 nm), which effectively encapsulate and release proteins. Show less
This thesis presents another approach for direct cytosolic delivery via membrane fusion. This approach is based on a complementary pair of coiled-coil forming peptides, K (KIAALKE)4 and E (EIAALEK... Show moreThis thesis presents another approach for direct cytosolic delivery via membrane fusion. This approach is based on a complementary pair of coiled-coil forming peptides, K (KIAALKE)4 and E (EIAALEK)4 and is mimicking the action of the SNARE-complex. The SNARE-complex is responsible for fusion between vesicles and membranes of cells in the synapse and is also composed of coiled-coil forming peptides. In our system, the peptides are conjugated to a cholesterol anchor via a polyethylene glycol (PEG) spacer, yielding lipopeptides CP4K4 and CP4E4. Membrane fusion between liposomes and the plasma membrane of the cells is triggered by these two lipopeptides when each embedded within the lipid bilayer of the liposomes or the plasma membrane of the cell. Show less
Ruthenium complexes are promising prodrugs in photoactivated chemotherapy (PACT): to prevent systemic therapeutic side-effects, a non-toxic version of the drug is introduced in the body and is... Show moreRuthenium complexes are promising prodrugs in photoactivated chemotherapy (PACT): to prevent systemic therapeutic side-effects, a non-toxic version of the drug is introduced in the body and is only activated at the place of the tumor by means of visible light irradiation. However, most of these PACT compounds are only sensitive for UV or blue light, while this light does not permeate the body very well, in contrast to red or near-infrared light. To circumvent this problem, the principle of light-upconversion can be used to "upgrade" red light to blue light in a drug carrier such as a nanovesicle: the tumor is irradiated with red light, after which blue light is generated locally and used to activate the prodrug. Among the various methods of light-upconversion, triplet-triplet annihilation upconversion (TTA-UC) was selected as the most promising. In this thesis it is described that green-to-blue and red-to-blue upconverting nanovesicles were prepared. The red-to-blue upconverted light was successfully used to activate a ruthenium polypyridyl complex that was anchored to the same vesicle. Finally, the inherent oxygen-sensitivity of TTA-UC was greatly mitigated by the addition of water-soluble and biocompatible anti-oxidants. We expect that the results of this thesis will lead to exciting applications in PACT. Show less
Bunschoten, A.; Chin, P.T.K.; Buckle, T.; Linden, M. van der; Barendregt, A.; Verheijen, M.A.; Leeuwen, F.W.B. van 2016
Inorganic nanoparticles are attractive materials due to their unique properties and prominent role in the fields of material science, nanotechnology and nanomedicine. Modern therapies aim to... Show moreInorganic nanoparticles are attractive materials due to their unique properties and prominent role in the fields of material science, nanotechnology and nanomedicine. Modern therapies aim to deliver drugs specifically to defective cells and mesoporous silica nanoparticles (MSNs) are considered to be promising candidates for this goal. In this thesis the synthesis, characterization and bio-applications of silica nanoparticles will be discussed. Moreover, the potential application of mesoporous silica nanoparticles as drug delivery systems will be discussed using two animal models: the Xenopus laevis and the Danio rerio. Show less
