Ficin-Cyclodextrin-Based Docking Nanoarchitectonics of Self-Propelled Nanomotors for Bacterial Biofilm Eradication.

Development of bioinspired nanomotors showing effective propulsion and cargo delivery capabilities has attracted much attention in the last few years due to their potential use in biomedical applications. However, implementation of this technology in realistic settings is still a barely explored field. Herein, we report the design and application of a multifunctional gated Janus platinum-mesoporous silica nanomotor constituted of a propelling element (platinum nanodendrites) and a drug-loaded nanocontainer (mesoporous silica nanoparticle) capped with ficin enzyme modified with ß-cyclodextrins (ß-CD). The engineered nanomotor is designed to effectively disrupt bacterial biofilms via H2O2-induced self-propelled motion, ficin hydrolysis of the extracellular polymeric matrix (EPS) of the biofilm, and controlled pH-triggered cargo (vancomycin) delivery. The effective synergic antimicrobial activity of the nanomotor is demonstrated in the elimination of Staphylococcus aureus biofilms. The nanomotor achieves 82% of EPS biomass disruption and a 96% reduction in cell viability, which contrasts with a remarkably lower reduction in biofilm elimination when the components of the nanomotors are used separately at the same concentrations. Such a large reduction in biofilm biomass in S. aureus has never been achieved previously by any conventional therapy. The strategy proposed suggests that engineered nanomotors have great potential for the elimination of biofilms.

Nanodevices for the Efficient Codelivery of CRISPR-Cas9 Editing Machinery and an Entrapped Cargo: A Proposal for Dual Anti-Inflammatory Therapy.

In this article, we report one of the few examples of nanoparticles capable of simultaneously delivering CRISPR-Cas9 gene-editing machinery and releasing drugs for one-shot treatments. Considering the complexity of inflammation in diseases, the synergistic effect of nanoparticles for gene-editing/drug therapy is evaluated in an in vitro inflammatory model as proof of concept. Mesoporous silica nanoparticles (MSNs), able to deliver the CRISPR/Cas9 machinery to edit gasdermin D (GSDMD), a key protein involved in inflammatory cell death, and the anti-inflammatory drug VX-765 (GSDMD45CRISPR-VX-MSNs), were prepared. Nanoparticles allow high cargo loading and CRISPR-Cas9 plasmid protection and, thus, achieve the controlled codelivery of CRISPR-Cas9 and the drug in cells. Nanoparticles exhibit GSDMD gene editing by downregulating inflammatory cell death and achieving a combined effect on decreasing the inflammatory response by the codelivery of VX-765. Taken together, our results show the potential of MSNs as a versatile platform by allowing multiple combinations for gene editing and drug therapy to prepare advanced nanodevices to meet possible biomedical needs.

Nanoprogrammed Cross-Kingdom Communication Between Living Microorganisms.

The engineering of chemical communication at the micro/nanoscale is a key emergent topic in micro/nanotechnology, synthetic biology, and related areas. However, the field is still in its infancy; previous advances, although scarce, have mainly focused on communication between abiotic micro/nanosystems or between microvesicles and living cells. Here, we have implemented a nanoprogrammed cross-kingdom communication involving two different microorganisms and tailor-made nanodevices acting as "nanotranslators". Information flows from the sender cells (bacteria) to the nanodevice and from the nanodevice to receiver cells (yeasts) in a hierarchical way, allowing communication between two microorganisms that otherwise would not interact.

Nanoparticle-cell-nanoparticle communication by stigmergy to enhance poly(I:C) induced apoptosis in cancer cells.

Nanoparticle-cell-nanoparticle communication by stigmergy was demonstrated using two capped nanodevices. The first community of nanoparticles (i.e.S(RA)IFN) is loaded with 9-cis-retinoic acid and capped with interferon-γ, whereas the second community of nanoparticles (i.e.S(sulf)PIC) is loaded with sulforhodamine B and capped with poly(I:C). The uptake of S(RA)IFN by SK-BR-3 breast cancer cells enhanced the expression of TLR3 receptor facilitating the subsequent uptake of S(sulf)PIC and cell killing.

An Interactive Model of Communication between Abiotic Nanodevices and Microorganisms.

The construction of communication models at the micro-/nanoscale involving abiotic nanodevices and living organisms has the potential to open a wide range of applications in biomedical and communication technologies. However, this area remains almost unexplored. Herein, we report, as a proof of concept, a stimuli-responsive interactive paradigm of communication between yeasts (as a model microorganism) and enzyme-controlled Janus Au-mesoporous silica nanoparticles. In the presence of the stimulus, the information flows from the microorganism to the nanodevice, and then returns from the nanodevice to the microorganism as a feedback.

A versatile drug delivery system targeting senescent cells.

Senescent cells accumulate in multiple aging-associated diseases, and eliminating these cells has recently emerged as a promising therapeutic approach. Here, we take advantage of the high lysosomal ß-galactosidase activity of senescent cells to design a drug delivery system based on the encapsulation of drugs with galacto-oligosaccharides. We show that gal-encapsulated fluorophores are preferentially released within senescent cells in mice. In a model of chemotherapy-induced senescence, gal-encapsulated cytotoxic drugs target senescent tumor cells and improve tumor xenograft regression in combination with palbociclib. Moreover, in a model of pulmonary fibrosis in mice, gal-encapsulated cytotoxics target senescent cells, reducing collagen deposition and restoring pulmonary function. Finally, gal-encapsulation reduces the toxic side effects of the cytotoxic drugs. Drug delivery into senescent cells opens new diagnostic and therapeutic applications for senescence-associated disorders.

ϵ-Polylysine-Capped Mesoporous Silica Nanoparticles as Carrier of the C9h Peptide to Induce Apoptosis in Cancer Cells.

Apoptotic signaling pathways are altered in numerous pathologies such as cancer. In this scenario, caspase-9/PP2Acα interaction constitutes a key target with pharmacological interest to re-establish apoptosis in tumor cells. Very recently, a short peptide (C9h) known to disrupt caspase-9/PP2Acα interaction with subsequent apoptosis induction was described. Here, we prepared two sets of mesoporous silica nanoparticles loaded with safranin O (S2) or with C9h peptide (S4) and functionalized with ϵ-polylysine as capping unit. Aqueous suspensions of both nanoparticles showed negligible cargo release whereas in the presence of pronase, a marked delivery of safranin O or C9h was observed. Confocal microscopy studies carried out with HeLa cells indicated that both materials were internalized and were able to release their entrapped cargos. Besides, a marked decrease in HeLa cell viability (ca. 50 %) was observed when treated with C9h-loaded S4 nanoparticles. Moreover, S4 provides peptide protection from degradation additionally allowing for a dose reduction to observe an apoptotic effect when compared with C9h alone or in combination with a cell-penetrating peptide (i.e., Mut3DPT-C9h). Flow cytometry studies, by means of Annexin V-FITC staining, showed the activation of apoptotic pathways in HeLa as a consequence of S4 internalization, release of C9h peptide and disruption of caspase-9/PP2Acα interaction.

Targeting inflammasome by the inhibition of caspase-1 activity using capped mesoporous silica nanoparticles.

Acute inflammation is a protective response of the body to harmful stimuli, such as pathogens or damaged cells. However, dysregulated inflammation can cause secondary damage and could thus contribute to the pathophysiology of many diseases. Inflammasomes, the macromolecular complexes responsible for caspase-1 activation, have emerged as key regulators of immune and inflammatory responses. Therefore, modulation of inflammasome activity has become an important therapeutic approach. Here we describe the design of a smart nanodevice that takes advantage of the passive targeting of nanoparticles to macrophages and enhances the therapeutic effect of caspase-1 inhibitor VX-765 in vivo. The functional hybrid systems consisted of MCM-41-based nanoparticles loaded with anti-inflammatory drug VX-765 (S2-P) and capped with poly-L-lysine, which acts as a molecular gate. S2-P activity has been evaluated in cellular and in vivo models of inflammation. The results indicated the potential advantage of using nanodevices to treat inflammatory diseases.

Broadening the antibacterial spectrum of histidine kinase autophosphorylation inhibitors via the use of ε-poly-L-lysine capped mesoporous silica-based nanoparticles.

Two-component systems (TCS) regulate diverse processes such as virulence, stress responses, metabolism and antibiotic resistance in bacteria but are absent in humans, making them promising targets for novel antibacterials. By incorporating recently described TCS histidine kinase autophosphorylation inhibitors (HKAIs) into ε-poly-L-lysine capped nanoparticles (NPs) we could overcome the Gram negative (Gr-) permeability barrier for the HKAIs. The observed bactericidal activity against Gr- bacteria was shown to be due to the enhanced delivery and internalization of the HKAIs and not an inhibitory or synergistic effect of the NPs. The NPs had no adverse effects on mammalian cell viability or the immune function of macrophages in vitro and showed no signs of toxicity to zebrafish larvae in vivo. These results show that HKAIs are promising antibacterials for both Gr- and Gr+pathogens and that NPs are a safe drug delivery technology that can enhance the selectivity and efficacy of HKAIs against bacteria.

Thrombin-Responsive Gated Silica Mesoporous Nanoparticles As Coagulation Regulators.

The possibility of achieving sophisticated actions in complex biological environments using gated nanoparticles is an exciting prospect with much potential. We herein describe new gated mesoporous silica nanoparticles (MSN) loaded with an anticoagulant drug and capped with a peptide containing a thrombin-specific cleavage site. When the coagulation cascade was triggered, active thrombin degraded the capping peptidic sequence and induced the release of anticoagulant drugs to delay the clotting process. The thrombin-dependent response was assessed and a significant increase in coagulation time in plasma from 2.6 min to 5 min was found. This work broadens the application of gated silica nanoparticles and demonstrates their ability to act as controllers in a complex scenario such as hemostasis.

Gated Materials for On-Command Release of Guest Molecules.

Multidisciplinary research at the forefront of the field of hybrid materials has paved the way to the development of endless examples of smart devices. One appealing concept in this fertile field is related to the design of gated materials. These are constructed for finely tuning the delivery of chemical or biochemical species from voids of porous supports to a solution in response to predefined stimuli. Such gated materials are composed mainly of two subunits: (i) a porous inorganic support in which a cargo is loaded and (ii) certain molecular or supramolecular entities, generally grafted onto the external surface, which can control mass transport from pores. On the basis of this concept, a large number of imaginative examples have been developed. This review intends to be a comprehensive analysis of papers published until 2014 on hybrid mesoporous gated materials. The molecules used as gates, the opening mechanisms, and controlled release behavior are detailed. We hope this review will not only help researchers who work in this field but also may open the minds of related ones to develop new advances in this fertile research area.

Targeting Innate Immunity with dsRNA-Conjugated Mesoporous Silica Nanoparticles Promotes Antitumor Effects on Breast Cancer Cells.

We describe herein a Toll-like receptor 3 (TLR3) targeting delivery system based on mesoporous silica nanoparticles capped with the synthetic double stranded RNA polyinosinic-polycytidylic acid (poly(I:C)) for controlled cargo delivery in SK-BR-3 breast carcinoma cells. Our results show that poly(I:C)-conjugated nanoparticles efficiently targeted breast cancer cells due to dsRNA-TLR3 interaction. Such interaction also triggered apoptotic pathways in SK-BR-3, significantly decreasing cells viability. Poly(I:C) cytotoxic effect in breast carcinoma cells was enhanced by loading nanoparticles' mesopores with the anthracyclinic antibiotic doxorubicin, a commonly used chemotherapeutic agent.

Poly(N-isopropylacrylamide)-gated Fe3O4/SiO2 core shell nanoparticles with expanded mesoporous structures for the temperature triggered release of lysozyme.

Core-shell nanoparticles comprised of Fe3O4 cores and a mesoporous silica shell with an average expanded pore size of 6.07 nm and coated with a poly(N-isopropylacrylamide) (PNIPAM) layer (CS-MSNs-EP-PNIPAM) were prepared and characterized. The nanoparticles was loaded with (Ru(bipy)3(2+)) dye or an antibacterial enzyme, lysozyme, to obtain CS-MSNs-EP-PNIPAM-Ru(bipy)3(2+) and CS-MSNs-EP-PNIPAM-Lys, respectively. The lysozyme loading was determined to be 160 mg/g of nanoparticle. It was seen that Ru(bipy)3(2+) and lysozyme release was minimal at a room temperature of 25 °C while at physiological temperature (37 °C), abrupt release was observed. The applicability of the CS-MSNs-EP-PNIPAM-Lys was further tested with two Gram-positive bacteria samples, Bacillus cereus and Micrococcus luteus. At physiological temperature, the nanoparticles were shown to reduce bacterial growth, indicating a successful release of lysozyme from the nanoparticles. This nanoparticle system shows potential as a nanocarrier for the loading of similarly sized proteins or other species as a drug delivery platform.

Synthesis and In†Vitro Evaluation of a Photosensitizer-BODIPY Derivative for Potential Photodynamic Therapy Applications.

A new photosensitizer (1) based on the 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY) scaffold has been synthesized. 1 is water soluble and showed an intense absorption band at 490 nm (ɛ=77,600 cm(-1) m(-1)) and an emission at 514 nm. In vitro toxicity of 1 in the presence of light and in darkness has been studied with HeLa, HaCaT, MCF-7, and SCC-13 cell lines. Moreover, internalization studies of 1 in these cell lines were also performed. These results suggested that 1 is more toxic for SCC-13 and HeLa carcinoma cells than for the HaCaT non-cancerous immortal human keratinocytes. Toxicity upon light irradiation was due to the formation of singlet oxygen and reactive oxygen species (ROS). Cellular co-localization experiments revealed preferential localization of the dye in the endoplasmic reticulum.

Gated mesoporous silica nanoparticles for the controlled delivery of drugs in cancer cells.

In recent years, mesoporous silica nanoparticles (MSNs) have been used as effective supports for the development of controlled-release nanodevices that are able to act as multifunctional delivery platforms for the encapsulation of therapeutic agents, enhancing their bioavailability and overcoming common issues such as poor water solubility and poor stability of some drugs. In particular, redox-responsive delivery systems have attracted the attention of scientists because of the intracellular reductive environment related to a high concentration of glutathione (GSH). In this context, we describe herein the development of a GSH-responsive delivery system based on poly(ethylene glycol)- (PEG-) capped MSNs that are able to deliver safranin O and doxorubicin in a controlled manner. The results showed that the PEG-capped systems designed in this work can be maintained closed at low GSH concentrations, yet the cargo can be delivered when the concentration of GSH is increased. Moreover, the efficacy of the PEG-capped system in delivering the cytotoxic agent doxorubicin in cells was also demonstrated.

eIF2 kinases mediate ß-lapachone toxicity in yeast and human cancer cells.

ß-Lapachone (ß-lap) is a novel anticancer agent that selectively induces cell death in human cancer cells, by activation of the NQO1 NAD(P)H dehydrogenase and radical oxygen species (ROS) generation. We characterized the gene expression profile of budding yeast cells treated with ß-lap using cDNA microarrays. Genes involved in tolerance to oxidative stress were differentially expressed in ß-lap treated cells. ß-lap treatment generated reactive oxygen species (ROS), which were efficiently blocked by dicoumarol, an inhibitor of NADH dehydrogenases. A yeast mutant in the mitochondrial NADH dehydrogenase Nde2p was found to be resistant to ß-lap treatment, despite inducing ROS production in a WT manner. Most interestingly, DNA damage responses triggered by ß-lap were abolished in the nde2Δ mutant. Amino acid biosynthesis genes were also induced in ß-lap treated cells, suggesting that ß-lap exposure somehow triggered the General Control of Nutrients (GCN) pathway. Accordingly, ß-lap treatment increased phosphorylation of eIF2α subunit in a manner dependent on the Gcn2p kinase. eIF2α phosphorylation required Gcn1p, Gcn20p and Nde2p. Gcn2p was also required for cell survival upon exposure to ß-lap and to elicit checkpoint responses. Remarkably, ß-lap treatment increased phosphorylation of eIF2α in breast tumor cells, in a manner dependent on the Nde2p ortholog AIF, and the eIF2 kinase PERK. These findings uncover a new target pathway of ß-lap in yeast and human cells and highlight a previously unknown functional connection between Nde2p, Gcn2p and DNA damage responses.

Oligonucleotide-capped mesoporous silica nanoparticles as DNA-responsive dye delivery systems for genomic DNA detection.

New hybrid oligonucleotide-capped mesoporous silica nanoparticles able to detect genomic DNA were designed.

Toward the design of smart delivery systems controlled by integrated enzyme-based biocomputing ensembles.

We report herein the design of a smart delivery system in which cargo delivery from capped mesoporous silica (MS) nanoparticles is controlled by an integrated enzyme-based "control unit". The system consists of Janus-type nanoparticles having opposing Au and MS faces, functionalized with a pH-responsive ß-cyclodextrin-based supramolecular nanovalve on the MS surface and two effectors, glucose oxidase and esterase, immobilized on the Au face. The nanodevice behaves as an enzymatic logical OR operator which is selectively fueled by the presence of D-glucose and ethyl butyrate.

Enhanced antifungal efficacy of tebuconazole using gated pH-driven mesoporous nanoparticles.

pH-sensitive gated mesoporous silica nanoparticles have been synthesized. Increased extracellular pH and internalization into living yeast cells triggered molecular gate aperture and cargo release. Proper performance of the system was demonstrated with nanodevices loaded with fluorescein or with the antifungal agent tebuconazole. Interestingly, nanodevices loaded with tebuconazole significantly enhanced tebuconazole cytotoxicity. As alterations of acidic external pH are a key parameter in the onset of fungal vaginitis, this nanodevice could improve the treatment for vaginal mycoses.