Polymer-grafted gold nanoparticles for cancer treatment: synthesis and evaluation of their radiosensitizing properties

International audienceToday, even though treatments have much improved, cancer is still a leading cause of death in the world, being responsible for 1 death out of 6. Radiotherapy is widely used for tumor treatment, but suffers from side effects due to the irradiation of healthy surrounding tissues. Another issue is the radioresistance developed by some tumor cells, which implies to increase the involved doses. The challenge remains to deliver curative doses to tumor tissues while sparing sound ones. Hence the use of tumor-located radiosensitizers is a promising way to improve the efficacy of radiotherapy. High-Z materials have been known for several decades to amplify the damaging effects of both photon and ion radiations. Various nanoparticles have already been developed to take advantage of this property: gold, platinum and gadolinium are amongst the most investigated elements. A well-controlled synthesis is key to obtain stable and scalable nano-objects. Here, various polymers were grafted onto metallic nanoparticles to improve stability and biocompatibility and to facilitate subsequent functionalization. Advanced methods of characterization attested both robustness and reproducibility of the synthesis procedure. Moreover, promising results were obtained regarding the radioenhancing properties of these hybrid nanocompounds. Therefore, special attention has been given to the underlying mechanisms of the assessed radiosensitization, since they are not fully understood yet. Synthesis of polymer-grafted gold nanoparticles was performed through an in situ method, via the reduction of gold salts in the presence of polymeric ligands mainly prepared using controlled radical polymerization. The resulting nano-objects were fully characterized by thermogravimetric analysis, inductively coupled plasma mass spectrometry (ICP-MS), transmission electronic microscopy and small-angle x-ray and neutron scattering. Interactions between our nanocompounds and biological systems were studied in order to better understand the mechanisms at play. At the cellular scale, three aspects were examined for each type of nanoparticles: cellular uptake, cytotoxicity and radiosensitizing properties, through ICP-MS measurements, cell proliferation assays and clonogenic assays respectively. All irradiations were performed while keeping the delivered doses to low values (under 30 Gy) that are typical of clinic reality. Different types of radiations were tested, in order to compare their effects and their synergy with the nanocompounds. The synthesized nano-objects have shown great potential to enhance radiation cancer treatment. Their stability and controlled surface chemistry have allowed to develop multiple strategies in order to optimize their radiosensitizing effect and in vitro behavior

Deep tissue penetration of bottle-brush polymers via cell capture evasion and fast diffusion

Drug nanocarriers (NCs) capable of crossing the vascular endothelium and deeply penetrating into dense tissues of the CNS could potentially transform the management of neurological diseases. In the present study, we investigated the interaction of bottle-brush (BB) polymers with different biological barriers in vitro and in vivo and compared it to nanospheres of similar composition. In vitro internalization and permeability assays revealed that BB polymers are not internalized by brain-associated cell lines and translocate much faster across a blood–brain barrier model compared to nanospheres of similar hydrodynamic diameter. These observations performed under static, no-flow conditions were complemented by dynamic assays performed in microvessel arrays on chip and confirmed that BB polymers can escape the vasculature compartment via a paracellular route. BB polymers injected in mice and zebrafish larvae exhibit higher penetration in brain tissues and faster extravasation of microvessels located in the brain compared to nanospheres of similar sizes. The superior diffusivity of BBs in extracellular matrix-like gels combined with their ability to efficiently cross endothelial barriers via a paracellular route position them as promising drug carriers to translocate across the blood–brain barrier and penetrate dense tissue such as the brain, two unmet challenges and ultimate frontiers in nanomedicine

Nanoparticules d'or à couronne polymère modulable : synthèse, interactions avec les systèmes biologiques et propriétés de radiosensibilisation

These past twenty years, there has been a great increase in the number of studies concerning the use of nanoparticles for medical applications, especially for cancer treatment. In particular, radiosensitizing effects of metal nanoparticles have been studied a lot in radiotherapy, in order to reduce the side effects created by the irradiation of healthy tissues surrounding the tumor. In this work, we focused on polymer-grafted gold nanoparticles. The polymer corona both ensured a great stability of the objects and allowed to change their physico-chemical properties, in order to study their impact on the nanoparticles behavior. We used polymethacrylates which were obtained through controlled radical polymerization and acted as ligands during the gold nanoparticles synthesis. A library of nano-objects with different properties was established by varying both molar mass and chemical nature of (co)polymers. Thorough characterization, including by small-angle radiation scattering, revealed similar structures for all synthesized objects. Grafting of one chemotherapy agent, doxorubicin, and two proteins was also performed on polymer ligands. Interactions between these various nanoparticles and biological systems were studied in detail. Special attention was given to the impact of polymer corona properties. Four aspects were examined: colloidal stability in biological media, ability to diffuse inside the extracellular matrix, cellular uptake, and cytotoxicity. All studied ligands ensured a great stability. Regarding the other aspects, systematic comparison of the results obtained for the whole library highlighted a strong impact of the ligands nature, especially the presence of positive charges or hydrophobic segments. We have also shown that grafted doxorubicin and protein kept their toxic and targeting properties respectively. Lastly, the prospect of using these nanoparticles for radiosensitization led us to study their behavior under radiations. When irradiated, their structure was found stable. Combining them with radioiodine (internal radiotherapy) showed a great radiosensitizing effect, both in vitro and in vivo, but experiments with protontherapy (external radiotherapy) revealed different behaviors depending on the type of radiations. We also investigated the use of particle-induced X-ray emission to detect nanoparticles in situ, during protontherapy treatment.La recherche concernant l’utilisation de nanoparticules dans le domaine médical a connu un essor considérable ces vingt dernières années, notamment dans le cadre du traitement du cancer. En particulier, l’effet radiosensibilisant des nanoparticules métalliques a été beaucoup étudié en radiothérapie, dans une perspective de réduction des effets secondaires générés par l’irradiation des tissus sains autour de la tumeur. Dans ce travail, nous nous sommes intéressés à des nanoparticules d’or greffées d’une couronne polymère. Celle-ci assure une grande stabilité des objets, tout en permettant de faire varier leurs propriétés physico-chimiques et d’en étudier l’impact sur leur comportement. Les polymères utilisés sont des polyméthacrylates, obtenus par polymérisation radicalaire contrôlée, qui jouent le rôle de ligands lors de la synthèse des nanoparticules d’or. Nous avons pu faire varier deux paramètres, la masse molaire et la nature chimique des (co)polymères, afin de constituer une gamme de nano-objets aux propriétés différentes. Une caractérisation approfondie, notamment par diffusion de rayonnements aux petits angles, a mis en évidence une structure similaire pour l’ensemble des objets synthétisés. Un agent de chimiothérapie, la doxorubicine, et deux protéines ont en outre pu être greffés sur les ligands polymères. Les interactions de ces différentes nanoparticules avec les systèmes biologiques ont été étudiées de manière détaillée, en particulier l’impact des propriétés de la couronne polymère. Quatre aspects ont été examinés : la stabilité colloïdale en milieux biologiques, la capacité à diffuser dans la matrice extracellulaire, la captation cellulaire et la cytotoxicité. Tous les ligands étudiés assurent une bonne stabilité. Concernant les autres aspects, la comparaison systématique des résultats obtenus pour la gamme d’objets a permis de mettre en évidence un fort impact de la nature de la couronne polymère, notamment de la présence de charges positives ou de segments hydrophobes. Nous avons également montré que les propriétés de toxicité de la doxorubicine, et de captation accrue d’une protéine étaient bien conservées après greffage. Dans une perspective d’utilisation en radiosensibilisation, nous avons enfin étudié le comportement de nos nanoparticules sous irradiation. Une bonne stabilité de leur structure a été observée sous rayonnement, et leur association à l’iode 131 (radiothérapie interne) a montré un fort effet radiosensibilisant, à la fois in vitro et in vivo. Les résultats des essais menés en protonthérapie (radiothérapie externe) mettent cependant en évidence des différences de comportements selon le type d’irradiation. Nous avons également exploré la possibilité de détecter les nanoparticules in situ, lors d’une irradiation en protonthérapie, grâce à l’analyse des rayons X émis

Gold nanoparticles grafted with a versatile polymer corona : synthesis, interactions with biological systems and radiosensitizing properties

La recherche concernant l’utilisation de nanoparticules dans le domaine médical a connu un essor considérable ces vingt dernières années, notamment dans le cadre du traitement du cancer. En particulier, l’effet radiosensibilisant des nanoparticules métalliques a été beaucoup étudié en radiothérapie, dans une perspective de réduction des effets secondaires générés par l’irradiation des tissus sains autour de la tumeur. Dans ce travail, nous nous sommes intéressés à des nanoparticules d’or greffées d’une couronne polymère. Celle-ci assure une grande stabilité des objets, tout en permettant de faire varier leurs propriétés physico-chimiques et d’en étudier l’impact sur leur comportement. Les polymères utilisés sont des polyméthacrylates, obtenus par polymérisation radicalaire contrôlée, qui jouent le rôle de ligands lors de la synthèse des nanoparticules d’or. Nous avons pu faire varier deux paramètres, la masse molaire et la nature chimique des (co)polymères, afin de constituer une gamme de nano-objets aux propriétés différentes. Une caractérisation approfondie, notamment par diffusion de rayonnements aux petits angles, a mis en évidence une structure similaire pour l’ensemble des objets synthétisés. Un agent de chimiothérapie, la doxorubicine, et deux protéines ont en outre pu être greffés sur les ligands polymères. Les interactions de ces différentes nanoparticules avec les systèmes biologiques ont été étudiées de manière détaillée, en particulier l’impact des propriétés de la couronne polymère. Quatre aspects ont été examinés : la stabilité colloïdale en milieux biologiques, la capacité à diffuser dans la matrice extracellulaire, la captation cellulaire et la cytotoxicité. Tous les ligands étudiés assurent une bonne stabilité. Concernant les autres aspects, la comparaison systématique des résultats obtenus pour la gamme d’objets a permis de mettre en évidence un fort impact de la nature de la couronne polymère, notamment de la présence de charges positives ou de segments hydrophobes. Nous avons également montré que les propriétés de toxicité de la doxorubicine, et de captation accrue d’une protéine étaient bien conservées après greffage. Dans une perspective d’utilisation en radiosensibilisation, nous avons enfin étudié le comportement de nos nanoparticules sous irradiation. Une bonne stabilité de leur structure a été observée sous rayonnement, et leur association à l’iode 131 (radiothérapie interne) a montré un fort effet radiosensibilisant, à la fois in vitro et in vivo. Les résultats des essais menés en protonthérapie (radiothérapie externe) mettent cependant en évidence des différences de comportements selon le type d’irradiation. Nous avons également exploré la possibilité de détecter les nanoparticules in situ, lors d’une irradiation en protonthérapie, grâce à l’analyse des rayons X émis.These past twenty years, there has been a great increase in the number of studies concerning the use of nanoparticles for medical applications, especially for cancer treatment. In particular, radiosensitizing effects of metal nanoparticles have been studied a lot in radiotherapy, in order to reduce the side effects created by the irradiation of healthy tissues surrounding the tumor. In this work, we focused on polymer-grafted gold nanoparticles. The polymer corona both ensured a great stability of the objects and allowed to change their physico-chemical properties, in order to study their impact on the nanoparticles behavior. We used polymethacrylates which were obtained through controlled radical polymerization and acted as ligands during the gold nanoparticles synthesis. A library of nano-objects with different properties was established by varying both molar mass and chemical nature of (co)polymers. Thorough characterization, including by small-angle radiation scattering, revealed similar structures for all synthesized objects. Grafting of one chemotherapy agent, doxorubicin, and two proteins was also performed on polymer ligands. Interactions between these various nanoparticles and biological systems were studied in detail. Special attention was given to the impact of polymer corona properties. Four aspects were examined: colloidal stability in biological media, ability to diffuse inside the extracellular matrix, cellular uptake, and cytotoxicity. All studied ligands ensured a great stability. Regarding the other aspects, systematic comparison of the results obtained for the whole library highlighted a strong impact of the ligands nature, especially the presence of positive charges or hydrophobic segments. We have also shown that grafted doxorubicin and protein kept their toxic and targeting properties respectively. Lastly, the prospect of using these nanoparticles for radiosensitization led us to study their behavior under radiations. When irradiated, their structure was found stable. Combining them with radioiodine (internal radiotherapy) showed a great radiosensitizing effect, both in vitro and in vivo, but experiments with protontherapy (external radiotherapy) revealed different behaviors depending on the type of radiations. We also investigated the use of particle-induced X-ray emission to detect nanoparticles in situ, during protontherapy treatment

FNA fluorescence reporting within silica-zirconia porous thin films

International audienceFunctional nucleic acids (FNAs) are promising molecular recognition elements, either as specific ligands (aptamers) or as catalysts (deoxyribo-or ribozymes). Indeed, FNAs may theoretically be selected to match any given target, regardless of its nature (ions, small molecules, proteins, peptides, or even cells). They are thus widely used for analysis applications, through different sensing technologies. Porous materials, when adequately tailored, offer the possibility to entrap various biomolecules without altering their conformation or function. The sol-gel process enables the formation of porous glasses or ceramics through polymerization at room temperature of liquid precursors (mainly metal or metalloid alkoxides). Upon fine-tuning, the resulting materials may display interesting properties such as optical transparency, tailored porosity, thermal or mechanical stability. In addition, such materials can be produced in a great variety of shapes (thin films, powders, monoliths, fibers, etc.). The sol-gel process is thus ideally versatile to prepare biohybrid materials, and has been applied to the encapsulation of all kinds of biological species, from proteins to cells, usually in order to develop biosensors. Performing sol-gel encapsulation of FNAs is a promising way to develop new medical sensors, especially through fluorescence transduction. However, literature examples of such biomaterials are still few, and focus exclusively on silica materials [1][2]. Considering the poor stability of silica at alkaline pH and the fact that most biological conditions require a pH higher than 7, the encapsulation of FNAs in silica-zirconia mixed oxides thin films was investigated. In order to spare precious biological samples, small volumes of solutions containing silica/zirconia precursors and fluorescently-labelled oligonucleotides were spin-coated onto quartz substrates. Uniform films with optical transparency, adjustable porosity and controlled thickness were obtained. Films stability was assessed in alkaline solutions up to pH 11 and resistance to basic conditions was established. Films were further characterized through front-face fluorescence spectroscopy by carrying out leaching tests and FNAs assays. FNA encapsulation was successfully performed in the silica-zirconia materials, establishing the possibility to create fluorescence-based sensors with these biomaterials

FNA Encapsulation witihn silica-zirconia porous thin films for fluorescence reporting

International audiencePorous materials, when adequately tailored, offer the possibility to entrap various biomolecules without altering their conformation or function. The sol-gel process, ideally versatile to prepare such biohybrid materials, has been applied to the encapsulation of all kinds of biological species, from proteins to whole cells, usually in order to develop biosensors. Simultaneously, functional nucleic acids (FNAs) were reported as new promising molecular recognition elements, whether as specific ligands (aptamers) or catalysts (deoxyribo-or ribozymes). Indeed, FNAs may theoretically be selected to match any given target, regardless of its nature (ions, small molecules, proteins, peptides, or even cells). Performing sol-gel encapsulation of FNAs is a promising way to develop new medical sensors, especially through fluorescence transduction. However, literature examples of such biomaterials are still few, and focus exclusively on silica materials [1][2]. Considering the poor stability of silica at alkaline pH and the fact that most biological conditions require a pH higher than 7, the encapsulation of FNAs in silica-zirconia mixed oxides thin films was investigated. Special attention was paid to the diffusion of small molecules into the immersed resulting materials, as this aspect is essential to obtain such a functional biomaterial. In order to spare precious biological samples, small volumes of sols containing silica/zirconia precursors and fluorescently-labelled oligonucleotides were spin-coated onto quartz substrates and uniform films with controlled thickness were obtained. Films stability was assessed in alkaline solutions up to pH 11 and resistance to basic conditions was established. Films were further characterized through front face fluorescence spectroscopy by carrying out leaching tests and FNAs assays. FNA encapsulation was successfully performed in the silica-zirconia materials, establishing the possibility to create fluorescence-based sensors with these biomaterials. Diffusion experiments of molecules of various sizes and natures (fluorophores, FNA's target, FNA) in the films were performed to better understand the impact of the porous network on such functional materials

FNA fluorescence reporting within silica-zirconia porous thin films

International audienceFunctional nucleic acids (FNAs) are promising molecular recognition elements, either as specific ligands (aptamers) or as catalysts (deoxyribo-or ribozymes). Indeed, FNAs may theoretically be selected to match any given target, regardless of its nature (ions, small molecules, proteins, peptides, or even cells). They are thus widely used for analysis applications, through different sensing technologies. Porous materials, when adequately tailored, offer the possibility to entrap various biomolecules without altering their conformation or function. The sol-gel process enables the formation of porous glasses or ceramics through polymerization at room temperature of liquid precursors (mainly metal or metalloid alkoxides). Upon fine-tuning, the resulting materials may display interesting properties such as optical transparency, tailored porosity, thermal or mechanical stability. In addition, such materials can be produced in a great variety of shapes (thin films, powders, monoliths, fibers, etc.). The sol-gel process is thus ideally versatile to prepare biohybrid materials, and has been applied to the encapsulation of all kinds of biological species, from proteins to cells, usually in order to develop biosensors. Performing sol-gel encapsulation of FNAs is a promising way to develop new medical sensors, especially through fluorescence transduction. However, literature examples of such biomaterials are still few, and focus exclusively on silica materials [1][2]. Considering the poor stability of silica at alkaline pH and the fact that most biological conditions require a pH higher than 7, the encapsulation of FNAs in silica-zirconia mixed oxides thin films was investigated. In order to spare precious biological samples, small volumes of solutions containing silica/zirconia precursors and fluorescently-labelled oligonucleotides were spin-coated onto quartz substrates. Uniform films with optical transparency, adjustable porosity and controlled thickness were obtained. Films stability was assessed in alkaline solutions up to pH 11 and resistance to basic conditions was established. Films were further characterized through front-face fluorescence spectroscopy by carrying out leaching tests and FNAs assays. FNA encapsulation was successfully performed in the silica-zirconia materials, establishing the possibility to create fluorescence-based sensors with these biomaterials

Hybrid metal-polymer nanoparticles as promising radiosensitizers for cancer treatment

International audienceNanotechnologies are being widely studied for medical applications, both diagnosis and treatment. They have already shown great promise, especially to treat cancer through various strategies such as chemotherapy, photothermal therapy or radiation therapies. High-Z elements nanoparticles are of particular interest for the latter, considering their ability to amplify the damaging effects of both photon and ion radiations: gold, platinum and gadolinium are amongst the most investigated elements. A well-controlled synthesis is key to obtain stable and scalable nano-objects. Here, various polymers were grafted onto metallic nanoparticles to improve stability and biocompatibility and to facilitate subsequent functionalization. Advanced methods of characterization attested to the robustness and reproducibility of the synthesis procedure. Moreover, promising results were obtained regarding the radioenhancing properties of these hybrid nanocompounds. Polymers mainly synthesized via controlled radical polymerization were grafted onto gold and platinum nanoparticles by a "grafting to" or "grafting from" method. Subsequent grafting of a chemotherapy drug onto the polymer corona was also successfully carried out. The resulting nano-objects were fully characterized by thermogravimetric analysis, transmission electronic microscopy and small-angle x-ray scattering. Small-angle neutron scattering was also performed, taking advantage of possible contrast matching. The impact of various radiation doses on the nanoparticles structure was studied. Finally, radiosensitizing effects were investigated through in vitro tests. Under irradiation, uncoupling and cleavage of polymer chains were demonstrated, leading to an overall size reduction of the hybrid nano-objects. The location of target sites during irradiation was determined and helped to better understand the underlying mechanism of the radiosensitization assessed by the in vitro results. The synthesized nano-objects have therefore shown great potential to enhance radiation cancer treatment. Their stability and controlled surface chemistry will allow to develop multiple strategies to further improve their radiosensitizing effect and in vitro behavior. In vivo tests are currently under study, as well as experiments regarding radioenhancement for proton therapy

How do surface properties of nanoparticles influence their diffusion in the extracellular matrix? A model study in Matrigel using polymer-grafted nanoparticles

International audienceDiffusion of nanomedicines inside the extracellular matrix (ECM) has been identified as a key factor to achieve homogeneous distribution and therefore therapeutic efficacy. Here, we sought to determine the impact of nanoparticles surface properties on their ability to diffuse in the ECM. As model nano-objects, we used a library of gold nanoparticles grafted with a versatile polymethacrylate corona which enabled to modify the surface properties. To accurately recreate the features of native ECM, diffusion studies were carried out in a tumor-derived gel (Matrigel®). We developed two methods to evaluate the diffusion ability of NPs inside this model gel: an easy to implement one based on optical monitoring and another one using small-angle X-ray scattering (SAXS) measurements. Both enabled to determine the diffusion coefficients of NPs and compare the influence of their various surface properties, while the SAXS technique also allowed to monitor the NPs structure as they diffused inside the gel. Positive charges and hydrophobicity were found to particularly hinder diffusion, and the different results suggested on the whole the presence of NPs-matrix interactions, therefore underlying the importance of the ECM model. The accuracy of the tumor-derived gels used in this study was evidenced by in vivo experiments involving intratumoral injections of NPs on mice, which showed that diffusion patterns in the peripheral tumor tissues were quite similar to the ones obtained within the chosen ECM model