Emerging applications of nanotechnology for diagnosis and therapy of disease: a review

Nanotechnology is of increasing interest in the fields of medicine and physiology over recent years. Its application could considerably improve disease detection and therapy, and although the potential is considerable, there are still many challenges, which need to be addressed before it is accepted in routine clinical use. This review focuses on emerging applications that nanotechnology could enhance or provide new approaches in diagnoses and therapy. The main focus of recent research centres on targeted therapies and enhancing imaging; however, the introduction of nanomaterial into the human body must be controlled, as there are many issues with possible toxicity and long-term effects. Despite these issues, the potential for nanotechnology to provide new methods of combating cancer and other disease conditions is considerable. There are still key challenges for researchers in this field, including the means of delivery and targetting in the body to provide effective treatment for specific disease conditions. Nanoparticles are difficult to measure due to the size and physical properties; hence there is still a great need to improve physiological measurements method in the field to ascertain how effective their use is in the human subject. This review is a brief snapshot into the fast changing research field of measurement and physiological links to nanoparticle use and its potential in the future

Exploiting the efficacy of Tyro3 and folate receptors to enhance the delivery of gold nanoparticles into colorectal cancer cells in vitro

Colorectal cancer (CRC) is the fourth most common cancer in the world. Due to its asymptomatic nature, CRC is diagnosed at an advanced stage where the survival rate is <5%. Besides, CRC treatment using chemotherapy, radiotherapy and surgery often causes undesirable side-effects. As such, gold nanoparticles (GNPs) are envisaged in the field for the diagnosis and treatment of CRC. GNPs have unique physical, chemical and electrical properties at the nanoscale which make them suitable for application in biomedicine. However, for GNPs to become clinically effective, their internalisation efficiency in cancer cells must be enhanced. Folate receptor-α (FR) is overexpressed in CRC cells wherein FR helps in the uptake of folic acid within the cells. Tyro3, a novel tyrosine kinase receptor, drives cell proliferation and its overexpression is correlated with poor prognosis in CRC. Their upregulated expression in CRC cells relative to normal cells makes them an ideal target for GNPs using active targeting. Therefore, in this study receptors FR and Tyro3 were simultaneously targeted using specific antibody-coated GNPs in order to enhance the uptake and internalisation of GNPs in CRC cells in vitro. Four different types of coated-GNPs were synthesised GNPs-PEG, GNPs-anti-FR, GNPs-anti-Tyro3 and GNPs-anti-(FR + Tyro3) and incubated (0–50 ng) with three CRC cell lines namely CRL1790, CRL2159 and HCT116. Simultaneous targeting of these receptors by GNPs-anti-(FR + Tyro3) was found to be the most effective in internalisation in CRC cells compared with GNPs targeted singly to FR or Tyro3 (p <0.05). Besides this, results show that Tyro3 mediated similar internalisation efficacy to FR (p <0.05) in CRC cells using ICP-OES

Inhibition of HIV virus by neutralizing Vhh attached to dual functional liposomes encapsulating dapivirine

Although highly active antiretroviral therapy (HAART) has greatly improved the life expectancy of HIV/AIDS patients, the treatment is not curative. It is a global challenge which fosters an urgent need to develop an effective drug or neutralizing antibody delivery approach for the prevention and treatment of this disease. Due to the low density of envelope spikes with restricted mobility present on the surface of HIV virus, which limit the antibody potency and allow virus mutation and escape from the immune system, it is important for a neutralizing antibody to form bivalent or multivalent bonds with the virus. Liposome constructs could fulfil this need due to the flexible mobility of the membrane with its attached antibodies and the capacity for drug encapsulation. In this study, we evaluated the neutralization activity of a range of liposome formulations in different sizes coated with anti-gp120 llama antibody fragments (Vhhs) conjugated via either non-covalent metal chelation or a covalent linkage. The non-covalent construct demonstrated identical binding affinity to HIV-1 envelope glycoprotein gp120 and neutralizing ability for HIV virus as free Vhh. Although covalently linked Vhh showed significant binding affinity to gp120, it unexpectedly had a lower neutralization potency. This may be due to the comparability in size of the viral and liposome particles restricting the number which can be bound to the liposome surface so involving only a fraction of the antibodies, whereas non-covalently attached antibodies dissociate from the surface after acting with gp120 and free the remainder to bind further viruses. Covalently conjugated Vhh might also trigger the cellular uptake of a liposome-virion complex. To explore the possible ability of the antibody-coated liposomes to have a further function, we encapsulated the hydrophobic antiviral drug dapivirine into both of the non-covalently and covalently conjugated liposome formulations, both of which revealed high efficacy in reducing viral replication in vitro. Thus, dual function liposomes may lead to a novel strategy for the prophylaxis of HIV/AIDS by combining the neutralizing activity of Vhh with antiviral effects of high drug concentrations

Detection of the tau protein in human serum by a sensitive four-electrode electrochemical biosensor

This study presents a novel approach based on a four-electrode electrochemical biosensor for the detection of tau protein – one of the possible markers for the prediction of Alzheimer's disease (AD). The biosensor is based on the formation of stable antibody–antigen complexes on gold microband electrodes covered with a layer of a self-assembled monolayer and protein G. Antibodies were immobilized on the gold electrode surface in an optimal orientation by protein G interaction. Electrochemical impedance spectroscopy was used to analyze impedance change, which revealed a linear response with increasing tau concentrations. The assay is fast (<1 h for incubation and measurement) and very sensitive. The limit of quantification for the full-length 2N4R tau protein is 0.03 pM, a value unaltered when the assay was processed in bovine serum albumin or human serum. This technology could be adapted for the detection of other biomarkers to provide a multiple assay to identify AD progression in a point of care setting

Glucose-coated gold nanoparticles transfer across human brain endothelium and enter astrocytes in vitro

The blood-brain barrier prevents the entry of many therapeutic agents into the brain. Various nanocarriers have been developed to help agents to cross this barrier, but they all have limitations, with regard to tissue-selectivity and their ability to cross the endothelium. This study investigated the potential for 4 nm coated gold nanoparticles to act as selective carriers across human brain endothelium and subsequently to enter astrocytes. The transfer rate of glucose-coated gold nanoparticles across primary human brain endothelium was at least three times faster than across non-brain endothelia. Movement of these nanoparticles occurred across the apical and basal plasma membranes via the cytosol with relatively little vesicular or paracellular migration; antibiotics that interfere with vesicular transport did not block migration. The transfer rate was also dependent on the surface coating of the nanoparticle and incubation temperature. Using a novel 3-dimensional co-culture system, which includes primary human astrocytes and a brain endothelial cell line hCMEC/D3, we demonstrated that the glucose-coated nanoparticles traverse the endothelium, move through the extracellular matrix and localize in astrocytes. The movement of the nanoparticles through the matrix was >10 µm/hour and they appeared in the nuclei of the astrocytes in considerable numbers. These nanoparticles have the correct properties for efficient and selective carriers of therapeutic agents across the blood-brain barrier

Nanoparticle electrical impedance tomography

We have developed a new approach to imaging with electrical impedance tomography (EIT) using gold nanoparticles (AuNPs) to enhance impedance changes at targeted tissue sites. This is achieved using radio frequency (RF) to heat nanoparticles while applying EIT imaging. The initial results using 5-nm citrate coated AuNPs show that heating can enhance the impedance in a solution containing AuNPs due to the application of an RF field at 2.60 GHz

Investigation of factors influencing the immunogenicity of hCG as a potential cancer vaccine

Human hCG and its β‐subunit (hCGβ) are tumour autocrine growth factors whose presence in the serum of cancer patients has been linked to poorer prognosis. Previous studies have shown that vaccines, which target these molecules and/or the 37 amino acid C‐terminal hCGβ peptide (hCGβCTP), induce antibody responses in a majority of human recipients. Here we explored whether the immunogenicity of vaccines containing an hCGβ mutant (hCGβR68E, designed to eliminate cross‐reactivity with luteinizing hormone) or hCGβCTP could be enhanced by coupling the immunogen to different carriers (KLH or Hsp70) using different cross‐linkers (EDC or GAD) and formulated with different adjuvants (RIBI or Montanide ISA720). While there was little to choose between KLH and Hsp70 as carriers, their influence on the effectiveness of a vaccine containing the BAChCGβR68E mutant was less marked, presumably because being a foreign species, this mutant protein itself might provide T‐helper epitopes. The mutant provided a significantly better vaccine than the hCGβCTP peptide irrespective of the carrier used, how it was cross‐linked to the carrier or which adjuvant was used when hCG was the target. Nonetheless, for use in humans where hCG is a tolerated self‐protein, the need for a carrier is of fundamental importance. Highest antibody titres were obtained by linking the BAChCGβR68E to Hsp70 as a carrier by GAD and using RIBI as the adjuvant, which also resulted in antibodies with significantly higher affinity than those elicited by hCGβCTP peptide vaccine. This makes this mutant vaccine a promising candidate for therapeutic studies in hCGβ‐positive cancer patients

Locating functionalized gold nanoparticles using electrical impedance tomography

Objective: An imaging device to locate functionalised nanoparticles, whereby therapeutic agents are transported from the site of administration specifically to diseased tissues, remains a challenge for pharmaceutical research. Here, we show a new method based on electrical impedance tomography (EIT) to provide images of the location of gold nanoparticles (GNPs) and the excitation of GNPs with radio frequencies (RF) to change impedance permitting an estimation of their location in cell models Methods: We have created an imaging system using quantum cluster GNPs as contrast agent, activated with RF fields to heat the functionalized GNPs, which causes a change in impedance in the surrounding region. This change is then identified with EIT. Results: Images of impedance changes of around 80±4% are obtained for a sample of citrate stabilized GNPs in a solution of phosphate-buffered saline. A second quantification was carried out using colorectal cancer cells incubated with culture media, and the internalization of GNPs into the colorectal cancer cells was undertaken to compare them with the EIT images. When the cells were incubated with functionalised GNPs, the change was more apparent, approximately 40±2%. This change was reflected in the EIT image as the cell area was more clearly identifiable from the rest of the area. Significance: EIT can be used as a new method to locate functionalized GNPs in human cells and help in the development of GNP-based drugs in humans to improve their efficacy in the future