Toxicogenomic Profiling of 28 Nanomaterials in Mouse Airways.

Toxicogenomics opens novel opportunities for hazard assessment by utilizing computational methods to map molecular events and biological processes. In this study, the transcriptomic and immunopathological changes associated with airway exposure to a total of 28 engineered nanomaterials (ENM) are investigated. The ENM are selected to have different core (Ag, Au, TiO2, CuO, nanodiamond, and multiwalled carbon nanotubes) and surface chemistries (COOH, NH2, or polyethylene glycosylation (PEG)). Additionally, ENM with variations in either size (Au) or shape (TiO2) are included. Mice are exposed to 10 µg of ENM by oropharyngeal aspiration for 4 consecutive days, followed by extensive histological/cytological analyses and transcriptomic characterization of lung tissue. The results demonstrate that transcriptomic alterations are correlated with the inflammatory cell infiltrate in the lungs. Surface modification has varying effects on the airways with amination rendering the strongest inflammatory response, while PEGylation suppresses toxicity. However, toxicological responses are also dependent on ENM core chemistry. In addition to ENM-specific transcriptional changes, a subset of 50 shared differentially expressed genes is also highlighted that cluster these ENM according to their toxicity. This study provides the largest in vivo data set currently available and as such provides valuable information to be utilized in developing predictive models for ENM toxicity.

Nano-SAR Modeling for Predicting the Cytotoxicity of Metal Oxide Nanoparticles to PaCa2.

Nowadays, the impact of engineered nanoparticles (NPs) on human health and environment has aroused widespread attention. It is essential to assess and predict the biological activity, toxicity, and physicochemical properties of NPs. Computation-based methods have been developed to be efficient alternatives for understanding the negative effects of nanoparticles on the environment and human health. Here, a classification-based structure-activity relationship model for nanoparticles (nano-SAR) was developed to predict the cellular uptake of 109 functionalized magneto-fluorescent nanoparticles to pancreatic cancer cells (PaCa2). The norm index descriptors were employed for describing the structure characteristics of the involved nanoparticles. The Random forest algorithm (RF), combining with the Recursive Feature Elimination (RFE) was employed to develop the nano-SAR model. The resulted model showed satisfactory statistical performance, with the accuracy (ACC) of the test set and the training set of 0.950 and 0.966, respectively, demonstrating that the model had satisfactory classification effect. The model was rigorously verified and further extensively compared with models in the literature. The proposed model could be reasonably expected to predict the cellular uptakes of nanoparticles and provide some guidance for the design and manufacture of safer nanomaterials.

Rationale and decision rules behind the ECETOC NanoApp to support registration of sets of similar nanoforms within REACH.

New registration requirements for nanomaterials under REACH consider the possibility to form 'sets of similar nanoforms' for a joined human health and environmental hazard, exposure and risk assessment. We developed a tool to create and justify sets of similar nanoforms and to ensure that each of the nanoforms is sufficiently similar to all other nanoforms. The decision logic is following the ECHA guidance in a transparent and evidence-based manner. For each two nanoforms the properties under consideration are compared and corresponding thresholds for maximal differences are proposed. In tier1, similarity is assessed based on intrinsic properties that mostly correspond to those required for nanoform identification under REACH: composition, impurities/additives, size, crystallinity, shape and surface treatment. Moreover, potential differences in the agglomeration/aggregation state resulting from different production processes are considered. If nanoforms were not sufficiently similar based on tier1 criteria, additional data from functional assays are required in tier2. In rare cases, additional short-term in vivo rodent data could be required in a third tier. Data required by tier 2 are triggered by the intrinsic properties in the first tier that did not match the similarity criteria. Most often this will be data on dissolution and surface reactivity followed by in vitro toxicity, dispersion stability, dustiness. Out of several nanoforms given by the user, the tool concludes which nanoforms could be justified to be in the same set and which nanoforms are outside. It defines the boundaries of sets of similar nanoforms and generates a justification for the REACH registration.

When Would Immunologists Consider a Nanomaterial to be Safe? Recommendations for Planning Studies on Nanosafety.

The immune system is professional in recognizing and responding to non-self, including nanomaterials. Immune responses by professional and nonprofessional immune cells are thus nearly inevitable upon exposure of cells and organisms to such materials. The state of research into taking the immune system into account in nanosafety studies is reviewed and three aspects in which further improvements are desirable are identified: 1) Due to technical limitations, more stringent testing for endotoxin contamination should be made. 2) Since under overdose conditions immunity shows unphysiological responses, all doses used should be justified by being equivalent to tissue-delivered doses. 3) When markers of acute inflammation or cell stress are observed, functional assays are necessary to distinguish between homeostatic fluctuation and genuine defensive or tolerogenic responses. Since immune activation can also indicate that the immune system considers a stimulus to be harmless and induces tolerance, activation markers by themselves do not necessarily imply a danger to the body. Guidelines such as these are necessary to approach the point where specific nanomaterials are classified as safe based on reliable testing strategies.

Nanopharmaceuticals: A focus on their clinical translatability.

Nanoscale materials have contributed changing the way biomedical researchers address the biological and therapeutic limitations of conventional drugs and diagnostic agents. Thanks to a plethora of different materials and synthetic routes, particulate carriers can be designed to modify the half-life of compounds, alter their biodistribution and control the drug release profile, eventually providing an overall clinical benefit to patients. While around 50 nanoformulations (excluding biologics) are already on the market, several challenges still withhold them from unlocking their full translational potential. This review discusses the advantages and current hurdles in the use of nanopharmaceuticals, and describes the most important nanotechnological approaches which have been investigated so far. A focus is given on the record of clinical success and failures and current clinical trends. In an effort to identify opportunities and problems associated with each specific nanosystem, this manuscript underlines the need of a more product-oriented research, that can foster the progress of nanomedicines to the clinic.

An in-depth multi-omics analysis in RLE-6TN rat alveolar epithelial cells allows for nanomaterial categorization.

BACKGROUND: Nanomaterials (NMs) can be fine-tuned in their properties resulting in a high number of variants, each requiring a thorough safety assessment. Grouping and categorization approaches that would reduce the amount of testing are in principle existing for NMs but are still mostly conceptual. One drawback is the limited mechanistic understanding of NM toxicity. Thus, we conducted a multi-omics in vitro study in RLE-6TN rat alveolar epithelial cells involving 12 NMs covering different materials and including a systematic variation of particle size, surface charge and hydrophobicity for SiO2 NMs. Cellular responses were analyzed by global proteomics, targeted metabolomics and SH2 profiling. Results were integrated using Weighted Gene Correlation Network Analysis (WGCNA). RESULTS: Cluster analyses involving all data sets separated Graphene Oxide, TiO2_NM105, SiO2_40 and Phthalocyanine Blue from the other NMs as their cellular responses showed a high degree of similarities, although apical in vivo results may differ. SiO2_7 behaved differently but still induced significant changes. In contrast, the remaining NMs were more similar to untreated controls. WGCNA revealed correlations of specific physico-chemical properties such as agglomerate size and redox potential to cellular responses. A key driver analysis could identify biomolecules being highly correlated to the observed effects, which might be representative biomarker candidates. Key drivers in our study were mainly related to oxidative stress responses and apoptosis. CONCLUSIONS: Our multi-omics approach involving proteomics, metabolomics and SH2 profiling proved useful to obtain insights into NMs Mode of Actions. Integrating results allowed for a more robust NM categorization. Moreover, key physico-chemical properties strongly correlating with NM toxicity were identified. Finally, we suggest several key drivers of toxicity that bear the potential to improve future testing and assessment approaches.

Catalytic Nanoassemblies Formed by Short Peptides Promote Highly Enantioselective Transfer Hydrogenation.

Self-assembly enables formation of incredibly diverse supramolecular structures with practically important functions from simple and inexpensive building blocks. Here, we show how a semirational, bottom-up approach to create emerging properties can be extended to a design of highly enantioselective catalytic nanoassemblies. The designed peptides comprising as few as two amino acid residues spontaneously self-assemble in the presence of metal ions to form supramolecular, vesicle-like nanoassemblies that promote transfer hydrogenation of ketones in an aqueous phase with excellent conversion rates and enantioselectivities (>90% ee).

A comprehensive perspective of food nanomaterials.

Nanotechnology is a rapidly developing toolbox that provides solutions to numerous challenges in the food industry and meet public demands for healthier and safer food products. The diversity of nanostructures and their vast, tunable functionality drives their inclusion in food products and packaging materials to improve their nutritional quality through bioactive fortification and probiotics encapsulation, enhance their safety due to their antimicrobial and sensing capabilities and confer novel sensorial properties. In this food nanotechnology state-of-the-art communication, matrix materials with particular focus on food-grade components, existing and novel production techniques, and current and potential applications in the fields of food quality, safety and preservation, nutrient bioaccessibility and digestibility will be detailed. Additionally, a thorough analysis of potential strategies to assess the safety of these novel nanostructures is presented.

Legal and practical challenges in classifying nanomaterials according to regulatory definitions.

The European Union (EU) has adopted nano-specific provisions for cosmetics, food and biocides, among others, which include binding definitions of the term "nanomaterial". Here we take an interdisciplinary approach to analyse the respective definitions from a legal and practical perspective. Our assessment reveals that the definitions contain several ill-defined terms such as "insoluble" or "characteristic properties" and/or are missing thresholds. Furthermore, the definitions pose major and so far unsolved analytical challenges that, in practice, make it nearly impossible to classify nanomaterials according to EU regulatory requirements. An important purpose of the regulations, the protection of human health and the environment, may remain unfulfilled and the development of innovative applications of nanomaterials may be facing a path full of (legal) uncertainties. Based on our findings, we provide five recommendations for a more coherent and practical approach towards the regulation of nanomaterials.

Material-specific properties applied to an environmental risk assessment of engineered nanomaterials - implications on grouping and read-across concepts.

Engineered nanomaterials (ENMs) are intentionally designed in different nano-forms of the same parent material in order to meet application requirements. Different grouping and read-across concepts are proposed to streamline risk assessments by pooling nano-forms in one category. Environmental grouping concepts still are in their infancy and mainly focus on grouping by hazard categories. Complete risk assessments require data on environmental release and exposure not only for ENMs but also for their nano-forms. The key requirement is to identify and to distinguish the production volumes of the ENMs regarding nano-form-specific applications. The aim of our work was to evaluate whether such a grouping is possible with the available data and which influence it has on the environmental risk assessment of ENMs. A functionality-driven approach was applied to match the material-specific property (i.e. crystal form/morphology) with the functions employed in the applications. We demonstrate that for nano-TiO2, carbon nanotubes (CNTs), and nano-Al2O3 the total production volume can be allocated to specific nano-forms based on their functionalities. The differentiated assessments result in a variation of the predicted environmental concentrations for anatase vs. rutile nano-TiO2, single-wall vs. multi-wall CNTs and α- vs. γ-nano-Al2O3 by a factor of 2 to 13. Additionally, the nano-form-specific predicted no-effect concentrations for these ENMs were derived. The risk quotients for all nano-forms indicated no immediate risk in freshwaters. Our results suggest that grouping and read-across concepts should include both a nano-form release potential for estimating the environmental exposure and separately consider the nano-forms in environmental risk assessments.

Insights into possibilities for grouping and read-across for nanomaterials in EU chemicals legislation.

This paper presents a comprehensive review of European Union (EU) legislation addressing the safety of chemical substances, and possibilities within each piece of legislation for applying grouping and read-across approaches for the assessment of nanomaterials (NMs). Hence, this review considers both the overarching regulation of chemical substances under REACH (Regulation (EC) No 1907/2006 on registration, evaluation, authorization, and restriction of chemicals) and CLP (Regulation (EC) No 1272/2008 on classification, labeling and packaging of substances and mixtures) and the sector-specific pieces of legislation for cosmetic, plant protection and biocidal products, and legislation addressing food, novel food, and food contact materials. The relevant supporting documents (e.g. guidance documents) regarding each piece of legislation were identified and reviewed, considering the relevant technical and scientific literature. Prospective regulatory needs for implementing grouping in the assessment of NMs were identified, and the question whether each particular piece of legislation permits the use of grouping and read-across to address information gaps was answered.

Grouping of nanomaterials to read-across hazard endpoints: a review.

The use of non-testing strategies like read-across in the hazard assessment of chemicals and nanomaterials (NMs) is deemed essential to perform the safety assessment of all NMs in due time and at lower costs. The identification of physicochemical (PC) properties affecting the hazard potential of NMs is crucial, as it could enable to predict impacts from similar NMs and outcomes of similar assays, reducing the need for experimental (and in particular animal) testing. This manuscript presents a review of approaches and available case studies on the grouping of NMs to read-across hazard endpoints. We include in this review grouping frameworks aimed at identifying hazard classes depending on PC properties, hazard classification modules in control banding (CB) approaches, and computational methods that can be used for grouping for read-across. The existing frameworks and case studies are systematically reported. Relevant nanospecific PC properties taken into account in the reviewed frameworks to support grouping are shape and surface properties (surface chemistry or reactivity) and hazard classes are identified on the basis of biopersistence, morphology, reactivity, and solubility.

Grouping of nanomaterials to read-across hazard endpoints: from data collection to assessment of the grouping hypothesis by application of chemoinformatic techniques.

BACKGROUND: An increasing number of manufactured nanomaterials (NMs) are being used in industrial products and need to be registered under the REACH legislation. The hazard characterisation of all these forms is not only technically challenging but resource and time demanding. The use of non-testing strategies like read-across is deemed essential to assure the assessment of all NMs in due time and at lower cost. The fact that read-across is based on the structural similarity of substances represents an additional difficulty for NMs as in general their structure is not unequivocally defined. In such a scenario, the identification of physicochemical properties affecting the hazard potential of NMs is crucial to define a grouping hypothesis and predict the toxicological hazards of similar NMs. In order to promote the read-across of NMs, ECHA has recently published "Recommendations for nanomaterials applicable to the guidance on QSARs and Grouping", but no practical examples were provided in the document. Due to the lack of publicly available data and the inherent difficulties of reading-across NMs, only a few examples of read-across of NMs can be found in the literature. This manuscript presents the first case study of the practical process of grouping and read-across of NMs following the workflow proposed by ECHA. METHODS: The workflow proposed by ECHA was used and slightly modified to present the read-across case study. The Read-Across Assessment Framework (RAAF) was used to evaluate the uncertainties of a read-across within NMs. Chemoinformatic techniques were used to support the grouping hypothesis and identify key physicochemical properties. RESULTS: A dataset of 6 nanoforms of TiO2 with more than 100 physicochemical properties each was collected. In vitro comet assay result was selected as the endpoint to read-across due to data availability. A correlation between the presence of coating or large amounts of impurities and negative comet assay results was observed. CONCLUSION: The workflow proposed by ECHA to read-across NMs was applied successfully. Chemoinformatic techniques were shown to provide key evidence for the assessment of the grouping hypothesis and the definition of similar NMs. The RAAF was found to be applicable to NMs.

Development of a systematic method to assess similarity between nanomaterials for human hazard evaluation purposes - lessons learnt.

Within the EU FP-7 GUIDEnano project, a methodology was developed to systematically quantify the similarity between a nanomaterial (NM) that has been tested in toxicity studies and the NM for which risk needs to be evaluated, for the purpose of extrapolating toxicity data between the two materials. The methodology is a first attempt to use current knowledge on NM property-hazard relationships to develop a series of pragmatic and systematic rules for assessing NM similarity. Moreover, the methodology takes into account the practical feasibility, in that it is based on generally available NM characterization information. In addition to presenting this methodology, the lessons learnt and the challenges faced during its development are reported here. We conclude that there is a large gap between the information that is ideally needed and its application to real cases. The current database on property-hazard relationships is still very limited, which hinders the agreement on the key NM properties constituting the basis of the similarity assessment and the development of associated science-based and unequivocal rules. Currently, one of the most challenging NM properties to systematically assess in terms of similarity between two NMs is surface coating and functionalization, which lacks standardized parameters for description and characterization methodology. Standardization of characterization methods that lead to quantitative, unambiguous, and measurable parameters describing NM properties are necessary in order to build a sufficiently robust property-hazard database that allows for evidence-based refinement of our methodology, or any other attempt to systematically assess the similarity of NMs.

Criteria for grouping of manufactured nanomaterials to facilitate hazard and risk assessment, a systematic review of expert opinions.

With the emergence of nanotechnology the number of manufactured nanomaterials (MNM) in production and use is constantly increasing. Exposure of workers to MNM is of concern, because still much is unknown about health effects. MNM may have different properties, testing of each material is time consuming and costly. Experts have proposed various approaches to categorize MNM to facilitate risk assessment of human health effects based on shared properties of various materials. A systematic literature survey was undertaken to identify expert opinions on grouping of MNM published between the years 2000 and 2015. We summarized and synthesized the opinions according to a systematic review of text and opinion. We identified 22 articles that fulfilled our inclusion criteria reporting 17 proposals with three proposals for groups and 14 proposals for criteria for grouping. Five proposals suggested one or more of the following groups of concern: fibrous, biopersistent, high solubility with high toxicity, chemically active. Criteria proposed in multiple studies were: viable testing options, mode of action, physicochemical properties predicting toxicity. We conclude that a limited number of groups have been proposed to categorize MNM according to human health concern. Further research should be conducted to underpin the proposed groups with empirical evidence.

Decision tree models to classify nanomaterials according to the DF4nanoGrouping scheme.

To keep pace with its rapid development an efficient approach for the risk assessment of nanomaterials is needed. Grouping concepts as developed for chemicals are now being explored for its applicability to nanomaterials. One of the recently proposed grouping systems is DF4nanoGrouping scheme. In this study, we have developed three structure-activity relationship classification tree models to be used for supporting this system by identifying structural features of nanomaterials mainly responsible for the surface activity. We used data from 19 nanomaterials that were synthesized and characterized extensively in previous studies. Subsets of these materials have been used in other studies (short-term inhalation, protein carbonylation, and intrinsic oxidative potential), resulting in a unique data set for modeling. Out of a large set of 285 possible descriptors, we have demonstrated that only three descriptors (size, specific surface area, and the quantum-mechanical calculated property 'lowest unoccupied molecular orbital') need to be used to predict the endpoints investigated. The maximum number of descriptors that were finally selected by the classification trees (CT) was very low- one for intrinsic oxidative potential, two for protein carbonylation, and three for NOAEC. This suggests that the models were well-constructed and not over-fitted. The outcome of various statistical measures and the applicability domains of our models further indicate their robustness. Therefore, we conclude that CT can be a useful tool within the DF4nanoGrouping scheme that has been proposed before.

Desenvolvimento e caracterização de polimerossomos para veiculação de L-asparaginase / Development and characterization of polymersomes for the release of L-asparaginase

A enzima L-Asparaginase (ASNase) é um biofámaco utilizado no tratamento da leucemia linfoblástica aguda, no entanto, a evolução da produção da ASNase como um medicamento desde o final da década de 1970 resultou em apenas quatro alternativas disponíveis no mercado farmacêutico, com relatos de graves reações imunogênicas e toxicidade. Desse modo, a nanotecnologia é uma plataforma que pode ser explorada para administração dessa enzima diminuindo a exposição da mesma a proteases e aumentando a sua meia-vida aparente. Os polimerossomos (PL) são opções que pela nanoestrutura vesicular poderiam encapsular a ASNase em seu core aquoso e pela presença de uma membrana polimérica, são mais robustos que os lipossomos. Assim, neste trabalho objetivou-se desenvolver PL para encapsulação da ASNase como uma alternativa às formulações deste biofármaco existentes. Foram desenvolvidos PL de PEG-PLA, PMPC-PDPA, PEG-PDPA e Pluronic® L-21. Foram estudados fatores relacionados à composição dos copolímeros (fração hidrofílica, responsividade a fatores externos tais como pH e temperatura) e métodos de elaboração (hidratação do filme polimérico, troca de pH e temperatura) bem como foi feita a caracterização dos PL obtidos (tamanho, índice de polidispersão, espessura de membrana, formação de excessivo bulk polimérico, obtenção de micelas). Também foi feito um planejamento racional para encapsulação da ASNase (hidratação direta do filme polimérico e encapsulação por eletroporação, autoagregação com encapsulação por troca de pH ou de temperatura). Para os PL preparados com PEG-PLA, a extrusão resultou em distribuição de tamanhos mais estreitos correspondentes aos valores de PDI de 0,345, 0,144 e 0,081 para PEG45-PLA69, PEG114-PLA153 e PEG114-PLA180, respectivamente. Foi demonstrado que copolímeros com menor fração hidrofóbica resultam em maior eficiência de encapsulação para proteínas, já que possuem volumes aquosos maiores. Com o PMPC25-PDPA72 foi possível encapsular em média três unidades de ASNase por vesículas através da eletroporação ou troca de pH, sendo que no primeiro método houve formação de túbulos e no último método as micelas não foram completamente removidas. Para PEG100-PDPA80, grandes agregados permaneceram após a purificação levando a um PDI alto, mas não foi observada a formação de túbulos, já a troca de pH para este copolímero resultou em maior perda de copolímeros como bulk polimérico precipitado. Para o copolimero tribloco Pluronic® L-121, foi observado que as vesículas eram estáveis durante uma semana à temperatura ambiente, contrariando o que era descrito na literatura. Nesses sistemas, quando preparados por hidratação do filme, a encapsulação da ASNase foi realizada por eletroporação mas a proteína não foi detectada dentro das vesículas. Atribuímos a não-encapsulação à organização da bicamada Pluronic® L-121 sem conformação definida das cadeias poliméricas, dificultando a reorganização do bloco hidrofílico na porção interna do poro durante eletroporação. Por troca de temperatura, cerca de 5 % de ASNase foi encapsulada e o método resultou em total recuperação da atividade da enzima. Desse modo foram obtidos diferentes PL com diferentes características nanoestruturais de acordo com os copolímeros utilizados para carreamento da ASNase

The enzyme L-Asparaginase (ASNase) is a biopharmaceutical used in the treatment of acute lymphoblastic leukemia, still the industrial production of ASNase as a marketable drug since the late 1970s has resulted in only four alternatives available in the pharmaceutical market, with reports of severe immunogenic reactions and toxicity. In this sense, nanotechnology is a platform that can be exploited to administer this enzyme by decreasing its exposure to proteases and increasing its apparent half-life. Polymerosomes (PL) are interesting routes which by its intrinsically vesicular nanostructure could encapsulate the ASNase in its aqueous core and by the presence of a polymeric membrane, being more robust than the liposomes. Thus, in this work it was intended to develop PL for ASNase encapsulation as an alternative to existing formulations of this biopharmaceutical. PL of PEG-PLA, PMPC-PDPA, PEG-PDPA and Pluronic® L-21 were developed. It was studied the copolymers composition (i.e. hydrophilic fraction, responsiveness to external factors such as pH and temperature), PL design (i.e. polymer film hydration, pH change and temperature) and PL characterization (i.e. size, polydispersity index - PDI, membrane thickness, formation of excessive polymer bulk, micelles production). A suitable experimental planning for ASNase encapsulation (i.e. direct hydration of the polymeric film and encapsulation by electroporation, self-aggregation with encapsulation by pH or temperature change) was also performed. For the PL prepared with PEG-PLA, the extrusion resulted in narrower size distribution corresponding to the PDI values of 0.345, 0.144 and 0.081 for PEG45-PLA69, PEG114-PLA153 and PEG114-PLA180, respectively. It has been shown that copolymers with lower hydrophobic fraction result in higher encapsulation efficiency for proteins, since they have larger aqueous volumes. With PMPC25-PDPA72 PL, it was possible to encapsulate three units of ASNase per vesicles through electroporation or pH change. In the first method, tubules were formed and in the latter one the micelles were not completely removed. For PEO100-PDPA80 PL, large aggregates remained after purification leading to a high PDI value, nevertheless no tubule formation was observed, since the pH change for this copolymer resulted in greater loss of copolymers as a precipitated polymer bulk. For the Pluronic® L-121 triblock copolymer PL, it was observed that the vesicles were stable for one week at room temperature, contrary to what was described in the literature. These PLs were prepared by film hydration method and ASNase encapsulation was performed by electroporation, nonetheless the protein was not detected within the vesicles. It is attributed the non-encapsulation to the organization of the Pluronic® L-121 bilayer without defined conformation of the polymer chains, making it difficult to reorganize the hydrophilic block in the internal portion of the pore during electroporation. By temperature change, about 5% of ASNase was encapsulated and the method resulted in complete recovery of enzyme activity. In conclusion, several PLs with a vast range of differential nanostructural characteristics were obtained according to the copolymers used for ASNase loading

Cytotoxicity screening and cytokine profiling of nineteen nanomaterials enables hazard ranking and grouping based on inflammogenic potential.

Engineered nanomaterials (ENMs) are being produced for an increasing number of applications. Therefore, it is important to assess and categorize ENMs on the basis of their hazard potential. The immune system is the foremost defence against foreign bodies. Here we performed cytokine profiling of a panel of nineteen representative ENMs procured from the Joint Research Centre (JRC) and commercial sources. Physicochemical characterization was performed using dynamic light scattering. The ENMs were all shown to be endotoxin content free. The human macrophage-differentiated THP.1 cell line was employed for cytotoxicity screening and based on the calculated IC50 values, the multi-walled carbon nanotubes (MWCNTs), ZnO, Ag and SiO2 NMs were found to be the most cytotoxic while single-walled carbon nanotubes (SWCNTs), TiO2, BaSO4 and CeO2 NMs, as well as the nanocellulose materials, were non-cytotoxic (at doses up to 100 µg/mL). Multiplex profiling of cytokine and chemokine secretion indicated that the TiO2, SiO2, BaSO4, CeO2 and nanocellulose materials induced potent inflammatory responses at sub-cytotoxic doses. Hierarchical clustering of cytokine responses coupled with pathway analysis demonstrated that the panel of ENMs could be segregated into two distinct groups characterized by activation and deactivation, respectively, of PPAR (peroxisome proliferator-activated receptor)/LXR (liver X receptor/retinoid X receptor) nuclear receptor pathways (NRPs). Furthermore, using rosiglitazone, a selective PPAR-γ agonist, we could show that PPAR-γ played an important role in the activation of inflammatory responses in cells exposed to TiO2 and SiO2 NMs. These studies show that ENMs of diverse chemical compositions can be grouped according to their inflammatory potential.

Smart materials on the way to theranostic nanorobots: Molecular machines and nanomotors, advanced biosensors, and intelligent vehicles for drug delivery.

BACKGROUND: Theranostics, a fusion of two key parts of modern medicine - diagnostics and therapy of the organism's disorders, promises to bring the efficacy of medical treatment to a fundamentally new level and to become the basis of personalized medicine. Extrapolating today's progress in the field of smart materials to the long-run prospect, we can imagine future intelligent agents capable of performing complex analysis of different physiological factors inside the living organism and implementing a built-in program thereby triggering a series of therapeutic actions. These agents, by analogy with their macroscopic counterparts, can be called nanorobots. It is quite obscure what these devices are going to look like but they will be more or less based on today's achievements in nanobiotechnology. SCOPE OF REVIEW: The present Review is an attempt to systematize highly diverse nanomaterials, which may potentially serve as modules for theranostic nanorobotics, e.g., nanomotors, sensing units, and payload carriers. MAJOR CONCLUSIONS: Biocomputing-based sensing, externally actuated or chemically "fueled" autonomous movement, swarm inter-agent communication behavior are just a few inspiring examples that nanobiotechnology can offer today for construction of truly intelligent drug delivery systems. GENERAL SIGNIFICANCE: The progress of smart nanomaterials toward fully autonomous drug delivery nanorobots is an exciting prospect for disease treatment. Synergistic combination of the available approaches and their further development may produce intelligent drugs of unmatched functionality.