Giorgia Pastorin

Cellular uptake of functionalized carbon nanotubes is independent of functional group and cell type

The development of nanomaterials for biomedical and biotechnological applications is an area of research that holds great promise and intense interest1, and carbon-based nanostructures in particular, such as carbon nanotubes (CNTs), are attracting an increasing level of attention2,3. One of the key advantages that CNTs offer is the possibility of effectively crossing biological barriers, which would allow their use in the delivery of therapeutically active molecules. Our laboratories have been investigating the use of CNTs in biomedical applications, and in particular as nanovectors for therapeutic agent delivery4,5,6,7,8. The interaction between cells and CNTs is a critical issue that will determine any future biological application of such structures. Here we show that various types of functionalized carbon nanotubes (f-CNTs) exhibit a capacity to be taken up by a wide range of cells and can intracellularly traffic through different cellular barriers.

Carbon nanotubes for delivery of small molecule drugs.

In the realm of drug delivery, carbon nanotubes (CNTs) have gained tremendous attention as promising nanocarriers, owing to their distinct characteristics, such as high surface area, enhanced cellular uptake and the possibility to be easily conjugated with many therapeutics, including both small molecules and biologics, displaying superior efficacy, enhanced specificity and diminished side effects. While most CNT-based drug delivery system (DDS) had been engineered to combat cancers, there are also emerging reports that employ CNTs as either the main carrier or adjunct material for the delivery of various non-anticancer drugs. In this review, the delivery of small molecule drugs is expounded, with special attention paid to the current progress of in vitro and in vivo research involving CNT-based DDSs, before finally concluding with some consideration on inevitable complications that hamper successful disease intervention with CNTs.

Translocation mechanisms of chemically functionalised carbon nanotubes across plasma membranes

Understanding the mechanisms responsible for carbon nanotube (CNT) internalisation into live cells is considered critical both from a fundamental point of view and for further engineering of CNT-based delivery systems to intracellular targets. While several studies are focused on the development of such CNT-based delivery systems, attempts to systematically elucidate the cellular uptake mechanisms of CNTs are still rather limited. The aim of the present study is to evaluate the cellular internalisation of chemically functionalised multi-walled carbon nanotubes (f-MWCNTs) in the presence of different well-known cellular uptake inhibitors. Our data reveal how f-MWCNTs are able to translocate across cell membranes of both phagocytic and non-phagocytic cell lines. We have evidenced that at least 30-50% of f-MWCNTs are taken up by cells through an energy-independent mechanism. This characteristic makes nanotubes loaded with therapeutic or diagnostic cargos extremely interesting as the release of active molecules directly into the cytoplasm increase their biological activity and therapeutic efficacy.

Tissue histology and physiology following intravenous administration of different types of functionalized multiwalled carbon nanotubes.

BACKGROUND Carbon nanotubes (CNTs) constitute one of the most important types of nanomaterials, increasingly gaining interest as tools for nanomedicine applications, such as sensors, implants or delivery systems. Our groups have reported previously that chemical functionalization of CNTs can lead to their almost complete elimination from the body of animals through the urinary excretion route. The administration of CNTs may, however, impact the physiological function of organs through which CNTs traverse or accumulate. AIM The present study addresses the short-term impact (first 24 h) of intravenous administration of various types of multiwalled nanotubes (MWNTs) on the physiology of healthy mice. MATERIALS & METHODS Nonfunctionalized, purified MWNTs (pMWNTs) and different types of water-dispersible, functionalized MWNTs (f-MWNTs) were tail-vein injected. Histological examination of tissues (kidney, liver, spleen and lung) harvested 24 h post-administration indicated that organ accumulation depended on the degree of ammonium (NH(3)(+)) functionalization at the f-MWNT surface. RESULTS The higher the degree of functionalization of MWNT-NH(3)(+), the less their accumulation in tissues. pMWNTs coated with autologous serum proteins prior to injection accumulated almost entirely in the lung and liver in large dark clusters. Moreover, various indicators of serum and urine analyses also confirmed that MWNT-NH(3)(+) injections did not induce any physiological abnormality in all major organs within the first 24 h post-injection. Interestingly, no abnormalities were observed either for f-MWNTs highly functionalized with carboxylate groups (diethylentriaminepentaacetic-functionalized MWNTs) or by upscaling to the highest doses ever injected so far in vivo (20 mg/kg). CONCLUSION The high degree of f-MWNT functionalization responsible for adequate individualization of nanotubes and not the nature of the functional groups was the critical factor leading to less tissue accumulation and normal tissue physiology at least within the first 24 h post-administration, even at the highest carbon nanotube doses ever administered in any study today.

Crucial functionalizations of carbon nanotubes for improved drug delivery: a valuable option?

Amidst the myriad of Drug Delivery Systems able to enhance delivery, absorption and intracellular uptake of a bioactive molecule while protecting it from deactivation, Carbon Nanotubes (CNTs) have emerged as a recent and promising option especially in cancer therapy. This is mainly due to their unique properties, which render them extremely versatile through the incorporation of several functional groups and targeting molecules at the same time, while their natural shape allows them to selectively penetrate across biological barriers in a non-invasive way. In this expert review we aim to evaluate whether this innovative material, once chemically-modified with suitable functionalizations, can be considered as a valuable system in comparison to the already existing nanodevices. This will include the estimation of the most recent advances in the field of nanotechnology, together with a cautious evaluation of potential risks and hazards associated with the extensive use of this fascinating, but still unknown, nanomaterial.

Nanomaterial scaffolds for stem cell proliferation and differentiation in tissue engineering.

Tissue engineering is a clinically driven field and has emerged as a potential alternative to organ transplantation. The cornerstone of successful tissue engineering rests upon two essential elements: cells and scaffolds. Recently, it was found that stem cells have unique capabilities of self-renewal and multilineage differentiation to serve as a versatile cell source, while nanomaterials have lately emerged as promising candidates in producing scaffolds able to better mimic the nanostructure in natural extracellular matrix and to efficiently replace defective tissues. This article, therefore, reviews the key developments in tissue engineering, where the combination of stem cells and nanomaterial scaffolds has been utilized over the past several years. We consider the high potential, as well as the main issues related to the application of stem cells and nanomaterial scaffolds for a range of tissues including bone, cartilage, nerve, liver, eye etc. Promising in vitro results such as efficient attachment, proliferation and differentiation of stem cells have been compiled in a series of examples involving different nanomaterials. Furthermore, the merits of the marriage of stem cells and nanomaterial scaffolds are also demonstrated in vivo, providing early successes to support subsequent clinical investigations. This progress simultaneously drives mechanistic research into the mechanotransduction process responsible for the observations in order to optimize the process further. Current understanding is chiefly reported to involve the interaction of stem cells and the anchoring nanomaterial scaffolds by activating various signaling pathways. Substrate surface characteristics and scaffold bulk properties are also reported to influence not only short term stem cell adhesion, spreading and proliferation, but also longer term lineage differentiation, functionalization and viability. It is expected that the combination of stem cells and nanomaterials will develop into an important tool in tissue engineering for the innovative treatment of many diseases.

Mitochondria-targeted antioxidants and metabolic modulators as pharmacological interventions to slow ageing

Populations in many nations today are rapidly ageing. This unprecedented demographic change represents one of the main challenges of our time. A defining property of the ageing process is a marked increase in the risk of mortality and morbidity with age. The incidence of cancer, cardiovascular and neurodegenerative diseases increases non-linearly, sometimes exponentially with age. One of the most important tasks in biogerontology is to develop interventions leading to an increase in healthy lifespan (health span), and a better understanding of basic mechanisms underlying the ageing process itself may lead to interventions able to delay or prevent many or even all age-dependent conditions. One of the putative basic mechanisms of ageing is age-dependent mitochondrial deterioration, closely associated with damage mediated by reactive oxygen species (ROS). Given the central role that mitochondria and mitochondrial dysfunction play not only in ageing but also in apoptosis, cancer, neurodegeneration and other age-related diseases there is great interest in approaches to protect mitochondria from ROS-mediated damage. In this review, we explore strategies of targeting mitochondria to reduce mitochondrial oxidative damage with the aim of preventing or delaying age-dependent decline in mitochondrial function and some of the resulting pathologies. We discuss mitochondria-targeted and -localized antioxidants (e.g.: MitoQ, SkQ, ergothioneine), mitochondrial metabolic modulators (e.g. dichloroacetic acid), and uncouplers (e.g.: uncoupling proteins, dinitrophenol) as well as some alternative future approaches for targeting compounds to the mitochondria, including advances from nanotechnology.

Quantum dots and carbon nanotubes in oncology: a review on emerging theranostic applications in nanomedicine

Cancer is one of the main causes of death in the world, and according to the WHO it is projected to continue rising. Current diagnostic modalities for the detection of cancer include the use of x-rays, magnetic resonance imaging and positron emission tomography, among others. The treatment of cancer often involves the use (or combination) of chemotherapeutic drugs, radiotherapy and interventional surgery (for solid and operable tumors). The application of nanotechnology in biology and medicine is advancing rapidly. Recent evidence suggests that quantum dots (QDs) can be used to image cancer cells as they display superior fluorescent properties compared with conventional chromophores and contrast agents. In addition, carbon nanotubes (CNTs) have emerged as viable candidates for novel chemotherapeutic drug delivery-platforms. The unique photothermal properties of CNTs also allow them to be used in conjunction with near infrared radiation and lasers to thermally ablate cancer cells. Furthermore, mounting evidence indicates that it is possible to conjugate QDs to CNTs, making it possible to exploit their novel attributes in the realm of cancer theranostics (diagnostics and therapy). Here we review the current literature pertaining to the applications of QDs and CNTs in oncology, and also discuss the relevance and implications of nanomedicine in a clinical setting.

The influence of carbon nanotubes on enzyme activity and structure: investigation of different immobilization procedures through enzyme kinetics and circular dichroism studies

In the last decade, many environmental organizations have devoted their efforts to identifying renewable biosystems, which could provide sustainable fuels and thus enhance energy security. Amidst the myriad of possibilities, some biofuels make use of different types of waste biomasses, and enzymes are often employed to hydrolyze these biomasses and produce sugars that will be subsequently converted into ethanol. In this project, we aimed to bridge nanotechnology and biofuel production: here we report on the activity and structure of the enzyme amyloglucosidase (AMG), physically adsorbed or covalently immobilized onto single-walled carbon nanotubes (SWNTs) and multi-walled carbon nanotubes (MWNTs). In fact, carbon nanotubes (CNTs) present several properties that render them ideal support systems, without the diffusion limitations displayed by porous material and with the advantage of being further functionalizable at their surface. Chemical ligation was achieved both on oxidized nanotubes (via carbodiimide chemistry), as well as on amino-functionalized nanotubes (via periodate-oxidized AMG). Results showed that AMG retained a certain percentage of its specific activity for all enzyme-carbon nanotubes complexes prepared, with the physically adsorbed samples displaying better catalytic efficiency than the covalently immobilized samples. Analysis of the enzymes structure through circular dichroism (CD) spectroscopy revealed significant structural changes in all samples, the degree of change being consistent with the activity profiles. This study proves that AMG interacts differently with carbon nanotubes depending on the method employed. Due to the higher activity reported by the enzyme physically adsorbed onto CNTs, these samples demonstrated a vast potential for further development. At the same time, the possibility of inducing magnetic properties into CNTs offers the opportunity to easily separate them from the original solution. Hence, substances to which they have been attached can be separated from a reaction medium, or directed by an external magnetic field to achieve efficient biofuel production. This paves the way for future design of efficient CNT-enzyme nanostructure bioreactors.

Discovery and Investigation of Anticancer Ruthenium–Arene Schiff-Base Complexes via Water-Promoted Combinatorial Three-Component Assembly

The structural diversity of metal scaffolds makes them a viable alternative to traditional organic scaffolds for drug design. Combinatorial chemistry and multicomponent reactions, coupled with high-throughput screening, are useful techniques in drug discovery, but they are rarely used in metal-based drug design. We report the optimization and validation of a new combinatorial, metal-based, three-component assembly reaction for the synthesis of a library of 442 Ru-arene Schiff-base (RAS) complexes. These RAS complexes were synthesized in a one-pot, on-a-plate format using commercially available starting materials under aqueous conditions. The library was screened for their anticancer activity, and several cytotoxic lead compounds were identified. In particular, [(η6-1,3,5-triisopropylbenzene)RuCl(4-methoxy-N-(2-quinolinylmethylene)aniline)]Cl (4) displayed low micromolar IC50 values in ovarian cancers (A2780, A2780cisR), breast cancer (MCF7), and colorectal cancer (HCT116, SW480). The absence of p53 activation or changes in IC50 value between p53+/+ and p53-/- cells suggests that 4 and possibly the other lead compounds may act independently of the p53 tumor suppressor gene frequently mutated in cancer.