Essay: Gold Nanostructures for Cancer Treatment

With cancer currently one of the most fatal diseases in the world, it is important to develop effective methods of detection and treatment, which would allow for quick diagnosis and efficient treatment that is of little discomfort to patients. Gold nanostructures with near infrared absorption may be the most promising solution, due to their ability to selectively destroy tumour tissue when coupled with laser light, and their potential to be applied in imaging as a contrast agent. This review will discuss the methods of synthesizing various gold nanostructures and the studies in which they have been applied in potentially treating cancer.
Cancer can be defined as the “abnormal growth of cells”,1 a result of changes in the cells’ genetic information so that they can no longer carry out the appropriate functions. When cancer cells grow in a mass together, it is referred to as a tumour and this can be either benign or malignant – benign tumours being those which do not affect nearby cells, and malignant tumours being those which will affect other cells, by a process known as “metastasis”. A tumour is defined as being benign or malignant based on the results of a biopsy, a method of examining the tumour by removing a small piece of it. Benign tumours are easier to treat through surgically removing them, due to the fact they are “self-contained”; malignant tumours, however, can affect the cells and tissue around them and so are far more difficult to treat.2 Currently, cancer is second only to heart disease as the most fatal disease in the US.3 Early detection is important to ensure that the treatment is as effective as possible – however, traditional methods need in excess of one million cells to detect the cancer, which does not allow for the early diagnosis that is desirable.4
Photothermal therapy using nanoparticles is a method for the treatment of cancer and, in particular, tumours. Compared to surgical methods, photothermal therapy has the ability to penetrate tumours in otherwise difficult to reach areas, and is non-invasive, so of less discomfort to the patient.5 The use of heat in the treatment of tumours is not a new concept (it has been used as early as 1700BC); however it is not without its disadvantages. Although tumours have a lower heat tolerance than healthy tissue and so can be selectively damaged by controlling the temperature range, it is difficult to have this level of control with traditional heat sources, and this can lead to healthy cells being damaged alongside the tumour.6 This is where laser light is especially suited, due to the light being in a small, in-phase beam – although even laser light presents difficulties, as it will destroy anything in its path.6
A potential application to solve the issues of non-selectivity is to couple the laser light with gold nanostructures which are near-IR absorbing: that is, they absorb in the near-infrared region of the electromagnetic spectrum, 700-2500nm.7 Examples of the gold nanostructures with near-IR absorption include gold-gold sulfide nanoparticles8, gold-chitosan nanocomposites9, and Fe3O4 polymer nanoparticles with a gold shell10, to name a few, and these will be discussed in further detail later in this review. When nanostructures which absorb in the NIR region are treated with such light, they convert the energy of the light into heat – enough to destroy a tumour, and thus can be used to selectively destroy tumours by being injected into them and irradiated with NIR laser light.11 NIR light is attractive due to the fact that it does not harm the tissue itself; it is only when coupled with the gold nanostructures that any damage is caused.12
An alternate method to photothermal therapy but still utilising the NIR absorption of gold nanostructures is to use NIR light to initiate the release of an anti-cancer drug, and use the gold nanostructures as a carrier for such drugs. An example of this is gold/gold-sulfide nanoparticles, and their synthesis and detailed application will be discussed further.8
This literature review will discuss the history and basic chemistry of gold nanoparticles, various methods of synthesising gold nanostructures with NIR absorption, and the ways in which they have been applied (or have the potential to be applied) in studies in order to detect and treat cancer and tumours, as well as exploring the ethical considerations of this branch of science and suggesting areas in which further research should be undertaken.
The earliest known use of gold nanoparticles (and perhaps the most famous) is the Lycurgus cup, a Roman cup from roughly the 4th Century which appears green when illuminated from the front, and red when illuminated from the inside. In 1980, the cup was confirmed to contain nanoparticles of silver and gold, with diameters ranging from 50nm to 100nm. The green colour is due to the diffusion of light from the outside, and the red colour is due to the silver-gold alloy that is present and absorbs at 515nm.13
Although gold nanoparticles are a new branch of medicine, gold itself has been used for medicinal applications as early as 2500BC in Ancient Egypt, and records show gold being used to treat fevers in the 17th Century and later syphilis in the 19th Century
The simplest and most common method of gold nanoparticle synthesis is the Turkevich method, involving the reduction of gold. This reaction is carried out at 100°C, and chloroauric acid is reduced by sodium citrate with constant stirring. It is possible to have variation in the diameters of the gold nanoparticles by varying the concentration of citrate used.17 This method was first proposed in 1951 by Turkevich, and in the 1970s was improved by Frens. Other methods which include the reduction of gold are the Brust-Schriffin method, using sodium borohydride as the reducing agent; the Murphy method, which uses ascorbic acid as the reducing agent; the Perrault method, where hydroquinone is the reducing agent, and the Polyol process, with various diols reducing the gold. Each of these methods will produce “colloidal gold”, i.e. gold nanoparticles in the liquid phase.13
This section will discuss the various methods proposed of synthesising gold nanostructures which are biocompatible and have NIR absorption, ideal for application in cancer diagnosis and therapy. The cost of materials will also be evaluated for each method, as well as the time consumed and ease of preparing the nanostructures.
Gao et al (2014) proposed a method for the synthesis of multilayered gold nanoshells, which consist of a gold nanoparticle core, a silicon coating, and a gold outer shell.18 In this method, gold nanoparticles are first synthesised according to methods suggested by Bastus et al, 2011, and then an organosilica layer added. Bastus’ method involves preparing gold seeds, by heating 150mL of 2.2mM sodium citrate solution with stirring, before the addition of 1mL of 25mM chloroauric acid. Methods of growth for nanoparticles of both 30nm and 180nm diameters were suggested, and given that Gao’s method uses 50nm gold nanoparticles, it can be assumed the latter was chosen. In order to grow the seeds to diameters of up to 180nm, the reaction mixture was cooled prior to the addition of a further two 1mL aliquots of 25mM chloroauric acid. 55mL of the solution was removed and replaced with 53mL of water and 2mL of 60mM sodium citrate, in order to dilute the solution.19
In order to form the organosilica layer, 16mL of 50nm colloidal gold was mixed with 100μl of a 100mM mercaptopropyltriethoxysilane (MPTES) solution, and left to stir for 10 minutes, before adding 150μL of 150mM PEG solution, and leaving this to stir for 15 minutes. After being centrifuged to produce a pellet, the pellet was dissolved in water and added to 25μL of MPTES, along with enough ammonia that it would be 25% of the final concentration. This was left overnight in order to produce the organosilica layer on the gold nanoparticles.
The gold-organosilicas were purified against water for 4 hours and then their pH adjusted to 9 using 0.5mM chloroauric acid and 0.01M sodium hydroxide. 4mL of 0.1M sodium borohydride was added in order to reduce the gold before centrifugation to produce gold-gold-organosilicas. To give the final layer of gold, 5mg of potassium carbonate, 750μL of 0.01M chloroauric acid and 20mL of water was mixed for 10 minutes and then added to the gold-gold-organosilicas, along with 250μL of 40mM ascorbic acid, in order to produce multi-layered gold nanoshells.20
The table above details the materials used and their respective cost; in cases where several purities were available, it was assumed that the highest purity would be required due to the potential biological applications of these nanoshells, where impurities could be problematic. The pricing shows that this is a particularly costly method – in particular, chloroauric acid, ammonia, sodium borohydride, polyethylene glycol and l-ascorbic acid, although these prices were based on the assumption that the highest purity would be required. However, materials such as potassium carbonate, sodium citrate and sodium hydroxide, it could be argued, are comparatively cheap and would likely be found in a typical laboratory, thus diminishing the disadvantage of the costlier reagents. This method also requires several steps and to allow the formation of the organosilica layer, the reaction mixture must be left overnight so overall this is a very time-consuming way of synthesising gold nanostructures.21
Another structure that has been proposed is gold chitosan nanocomposites (Zhang et al, 2012), in particular for application in photothermal therapy thanks to their absorption in the NIR region of the spectrum. The synthesis of these structures is detailed below.
First, gold nanoparticles were synthesised using a one-step reaction, where 3mM sodium thiosulfate was quickly added to 1.71mM chloroauric acid, then vortexed for 20 seconds.22 Next, carboxymethylated chitosan to coat the gold nanoparticles was prepared according to the method reported by Chen et al (2003)23, by combining 10g of chitosan, 13.5g of sodium hydroxide and 100mL of solvent (20mL of deionised water and 80mL of isopropanol) in a 500mL flask for one hour. 15g of monochloroacetic acid was then dissolved using 20mL isopropanol, added slowly to the mixture in the flask over a period of 30 minutes, then left to react at ~60°C for four hours. Ethyl alcohol (80% purity) was used to bring the reaction to a halt and remove additional salt and water, before the product was vacuum-dried. A solution of carboxymethylated chitosan was prepared by dissolving 1g of the product in 100mL of deionised water. A stock solution was also prepared of chitosan, by dissolving 1g of chitosan with 100mL of water.
The final step is to coat the gold nanoparticles with the chitosan and carboxymethylated chitosan, and purify them. This is achieved by centrifuging he chitosan and carboxymethylated chitosan solutions for ten minutes and dialysing for 48 hours. The three types of chitosan coated gold nanoparticles prepared were as follows: chitosan coated gold nanoparticles; carboxymethylated chitosan coated gold nanoparticles; chitosan/carboxymethylated chitosan coated gold nanoparticles. The appropriate chitosan solution was added to one of three solutions of gold nanoparticles, left to mix for a full day, and then centrifuged to remove impurities, unwanted components, and any excess chitosan. After centrifugation, the precipitate was suspended in deionised water.9
Compared to the previous method proposed by Gao et al, gold-chitosan nanocomposites are comparatively inexpensive to make – whilst the chloroauric acid is costly, the other materials required are of low cost and likely to be found in a typical laboratory; in particular chitosan is easy to come by and is widely used as a dietary supplement24. A low molecular weight of chitosan is also required, which contributes to a lower cost (as before, all other materials were priced assuming the highest purity would be required).
The bare gold nanoparticles synthesised for the initial stage of the reaction were prepared using a one step reaction, which adds to the desirability of this method. The three types of gold-chitosan nanocomposites (chitosan coated, carboxymethylated chitosan coated and chitosan/carboxymethylated chitosan coated) could also be prepared simultaneously, thus saving on the time consumed in synthesising them. However, there is a great deal of waiting involved in this method – one hour following the addition of sodium hydroxide and solvent to chitosan, four hours following the addition of isopropanol to monochloroacetic acid, 48 hours for the dialysis of the chitosan solutions following centrifugation, and a full day after the addition of the three chitosan solutions to the gold nanoparticles.
The third method that this review shall examine is those of gold/gold-sulfide nanoparticles (Ren and Chow, 2003), which are designed to release an anti-cancer drug upon irradiation with NIR light, rather than be used in the application of photothermal therapy as with previous structures. The procedure for synthesising these nanoparticles is detailed below.
First, three 20mL aliquots of 2mM chloroauric acid were mixed with 16mL, 20mL and 40mL of 1mM sodium sulfide. The reaction mixture was stored at room temperature for 24 hours and then centrifuged. The precipitate was dispersed in a solution of 100mM 11-mercaptoundecanoic acid in ethanol, and stored for 72 hours at 40°C. The resulting mixture was centrifuged a further three times and the precipitate suspended in water, before the addition of 10mg of cisplatin, an anti-cancer drug
In comparison to previous structures, the synthesis of gold/gold-sulfide nanoparticles is far simpler, both in terms of the number of steps required and the number of materials used. Previous methods have required the synthesis of “bare” gold nanoparticles before adding additional reagents to alter their shape or add a coating, etc. This method does not require any “bare” nanoparticles to be created prior to starting. However, although there are fewer steps, there are two points in this method which involve storing the reaction mixture for extended periods of time – first, for 24 hours after combining the chloroauric acid and sodium sulfide, and for another 72 hours after dispersing the solution in 11-mercaptoundecanoic acid.
There are far fewer reactants required for this method in comparison to the two previously mentioned, however with the exception of ethanol, the three main reactants (chloroauric acid, sodium sulfide and 11-mercaptoundecanoic acid) are extremely costly when the highest purity is chosen, and no data was available for cisplatin in order to comment on the total cost
In a variation from the relatively simple nanostructures discussed above, the following method proposed by Wang et al 2008 is one for preparing Fe3O4-polymer-gold shell nanoparticles, which not
only absorb in the near-infrared region, but also display magnetism, which would allow them to be used in MRI, and also to be controlled via a magnetic field to the area of a malignant tumour.10
First, “bare” gold nanoparticles were synthesised from the reduction of gold. This was achieved by mixing 100mL of deionised water with 2mL of 1% by weight sodium citrate solution, and adding this to 1mL of 1% by weight chloroauric acid with stirring. 1mL of 0.1% by weight sodium borohydride solution was then added and the mixture stirred for ten minutes, which allowed the sodium borohydride to reduce the gold. The resulting gold nanoparticles were stored at 4C.25
Following the preparation of gold nanoparticles, magnetite nanoparticles were synthesised for the coating of the gold nanoparticles, according to the method proposed by Li et al, 2005. 1.35 g of 0.005M iron (III) chloride hexahydrate, 40mL of ethylene glycol, 3.6g of sodium acetate and 1g of polyethylene glycol were combined and stirred for half an hour. The solution was then autoclaved at 200C for eight hours before being washed and dried, in preparation for the addition of a polymer coating.26
The magnetite nanoparticles were diluted to 10mg mL-1 with ethanol, before addition to 30mg methacrylic acid, 30mg acrylamide, 20mg ethylene glycol dimethacrylate, 25mg azobisisobutyronitrile and 40mL of acetonitrile. This mixture was sonicated then boiled for two hours before removing impurities by the use of a magnetic field, and washing with ethanol and water. The polymer-coated nanoparticles were then suspended in water.
Before the addition of the gold nanoparticles, a layer of poly (allylamine hydrochloride) was added to the polymer-coated nanoparticles. 1mL of the nanoparticles were diluted with 20mL of water, and the pH of this solution adjusted to 8.5 by ammonia, of 30% by weight. 20l of 10mg mL-1 poly (allylamine hydrochloride) was added to the mixture before purification involving a magnetic field and water, as previously described. This resulted in poly (allylamine hydrochloride) coated nanoparticles, which were suspended in 20mL of water.
The penultimate step was the addition of a coating of gold seeds to the poly (allylamine hydrochloride) coated nanoparticles, by combining 20mL of the “bare” gold nanoparticles from the first step with 20mL of the poly (allylamine hydrochloride) coated-nanoparticles in an ice bath. Again, purification was achieved with a magnetic field and water. The nanoparticles were then suspended in 20mL of water with 5mg of thiolated polyethylene glycol, and again washed with water before being suspended in 40mL of water.
The final stage was the formation of the gold shell, where 50L of 1.0% by weight chloroauric acid was added to the thiolated PEG-modified nanoparticles, followed by 100L of 80mM hydroxylamine. Again, a magnetic field and water were used to separate the nanoparticles before suspending them in 20mL of water with a further 5mg of polyethylene glycol, and another purification step involving a magnetic field and water was carried out. The steps involving the formation of the gold shell were repeated three times in order to achieve a full coverage of the nanoparticle.10
Of all the methods discussed until this point, this is by far the most complex in terms of the number of steps, and the variety of materials used. This is due to the number of layers involved in the nanoparticles – magnetite nanoparticles, a poly (allylamine hydrochloride) coating, a thiolated PEG coating, and a gold shell. This is also a very costly procedure, not just due to the number of reagents but because several reagents are individually expensive – chloroauric acid, polyethylene glycol, ammonia, poly (allylamine hydrochloride), thiolated PEG, and hydroxylamine in particular. It was again assumed that the highest purity of each reagent would be desired given the potential biological application.
However, despite the disadvantages in the synthesis of these nanoparticles, the end product stands out from those discussed in previous methods, due to the magnetic nanoparticles at the core. The resulting magnetism introduces a level of controllability that is not present in, for example, the gold-chitosan nanocomposites or the multi-layered gold nanoshells, both discussed above.
The nanostructures discussed so far have mainly been spherical in geometry – Huang et al (2005) proposed a method of preparing gold nanorods with near-infrared absorption which could be applied in both the detection and treatment of malignant cells.
The first stage in synthesising the gold nanorods was the preparation of a “seed solution”, from the reduction of 0.0005M chloroauric acid in 0.2M cetyltrimethylammonium bromide (CTAB), by 0.01M sodium borohydride. Next, a “growth solution” was prepared. Chloroauric acid (0.001M) was reduced by 0.2M cetyltrimethylammonium bromide, 0.15M benzyldimethylhexadecylammonium chloride, 0.004M silver salt, and 0.0788M ascorbic acid. Gold nanorods were then achieved by adding 8L of the seed solution into the prepared growth solution and storing until nanorods had formed.
Following the formation of gold nanorods, they were conjugated to anti-epidermal growth factor receptor. First, any excess cetyltrimethylammonium bromide was removed by centrifugation, and then the precipitate suspended in HEPES (N-(2-hydroxyethyl) piperazine-N’-2-ethanesulfonic acid) solution of pH 7.4, then added to an antibody solution and allowed to mix for 20 minutes. This solution was then centrifuged, and the precipitate suspended in phosphate buffered saline of pH 7.4.27
Chloroauric acid, CTAB, sodium borohydride and ascorbic acid are all extremely costly, and although BAC, HEPES and phosphate buffered saline solution are relatively inexpensive, they are outweighed by these four expensive materials (however, these costs were based on the assumption that the highest purity was required). As the method did not specify which silver salt was used, no price could accurately be given for this material. This makes it difficult to comment on the total cost incurred in synthesising these nanorods and conjugating them to anti-EGFR. Compared to other methods discussed, this is simple with few steps, and no time is consumed in leaving the reaction mixture for hours at a time.
Silica nanoparticles were first prepared using the Stober process, where tetraethyl orthosilicate (TEOS) was reduced by ethanol, with ammonia being used as a catalyst.28 (3-aminopropryl) triethoxysilane (APTES) was added to the silica nanoparticles to allow the gold colloid to adsorb to the nanoparticle surface in the next step.29
Gold colloid was synthesised according to a method detailed by Duff et al, 1993. In this method, 1.5mL of 0.2M sodium hydroxide was added to 45.5mL of water, along with 1mL of a M solution of tetrakis(hydroxymethyl)phosphonium chloride and 2mL of 25mM chloroauric acid.30 Following preparation of the gold colloid, a rotary evaporator was used to concentrate the nanoparticles before mixing them with the silica nanoparticles. Formaldehyde was then used to grow the gold shell, by acting as a reducing agent for the gold in the chloroauric acid. Finally, 20L of 5M thiolated polyethylene glycol was used to increase the biocompatibility of the nanoshells by preventing the body from having an “immune response”. Filtration and centrifugation were used to purify the nanoshells before suspending them in phosphate buffer saline.31
Chloroauric acid, thiolated PEG and APTES all contribute to this being a particularly costly method, however materials such as formaldehyde, ethanol, ammonia and sodium hydroxide would most likely be found in a typical laboratory, which reduces the impact of the higher cost materials. However, as before, it was assumed that the highest purity was required which would obviously increase the cost. The method of preparing these gold nanoshells is also far simpler than others, for example to Fe3O4-polymer-gold nanoshells, and no time is consumed in leaving the reaction mixture for hours at a time.
This section will discuss the ways in which the NIR-absorbing nanostructures detailed in the above section can be applied in cancer diagnosis and treatment, from photothermal therapy, to contrast agents in imaging, to drug delivery systems
After synthesis of the multi-layered gold nanoshells, proposed by Gao et al and discussed above, their applicability in photothermal therapy was studied both in vitro (outside of the body) and in vivo (inside of the body). Breast cancer tumour cells were chosen for this study. First, the effectiveness of the nanoshells was tested in vitro by treating four groups of breast cancer cells under a microscope, the results of which are shown in Figure 11 below. It can be seen clearly that neither the gold nanoshells or 808nm laser light alone (C2 and C3) caused a significant effect, however when combined, as in C4, only 10% remained capable of surviving.
For the in vivo study, mice with breast cancer tumours were injected with either saline (as a control) or the multi-layered gold nanoshells, and exposed to laser radiation of 808nm.
As shown in the image above, the tumour regions in mice treated with multi-layered gold nanoshells were significantly hotter after laser irradiation than in those which had been treated with saline – the tumour regions treated with multi-layered gold nanoshells reached temperatures of 58C, whilst those treated with saline only reached temperatures of 38C.
The study carried out by Gao et al is particularly extensive as it studies the effects of the nanoshells in response to NIR light both in and outside of the body, giving it an advantage over the majority of studies which will only choose one option – studying the effects in both environments is necessary as nanoparticles can react very differently with in vitro cells compared to how they react once inside a living body. However, there is a further step which could improve the thoroughness of this study, and that is to test the gold nanoshells on humans – different animals will metabolise and interact with such species in different ways, and so a successful test on mice is no indication that there will be the same success in humans.
Despite the lack of testing in humans, this study has still demonstrated the promise of gold nanoshells in photothermal therapy, with only 10% of cancerous cells surviving after treatment with nanoshells and NIR light. The study has also proven that gold nanoshells and NIR light are only capable of producing a photothermal effect when coupled with one another, and alone neither will produce heat great enough to destroy a tumour.
As with the multi-layered gold nanoshells, the gold chitosan nanocomposites proposed by were studied in the application of photothermal therapy. First, the optical stability of the nanocomposites was determined by measuring their NIR absorbance against time, using “bare” gold nanoparticles as a control. This test found that bare gold nanoparticles had the lowest optical stability.
Following this, both HDF (human dermal fibroblast) and HepG2 (human liver cancer) cells were treated with laser light of 817nm and one of the three gold-chitosan nanocomposites, shown in the images below.
From the images above, it can be seen clearly that laser light and gold-chitosan nanocomposites (d) were most effective at causing cell death in both cases, however it is the gold-chitosan-carboxymethylated chitosan nanocomposites (f) that give the most desirable effect, as they destroyed almost all of the liver cancer cells but less than one third of the healthy HDF cells. Laser light alone (b) and “bare” gold nanoparticles (c) had little to no effect on either cell. However, although promising results, the entirety of this study was conducted in vitro, and so further testing in vitro would be desirable in order to fully predict how these nanocomposites will react in photothermal therapy when inside the body – for example, in the treatment of mice injected with tumours.9
Thus far, most of the suggested gold nanostructures have been applied in the treatment of tumours via photothermal therapy. The gold/gold sulfide nanoparticles prepared in the method described by Ren and Chow were studied as a method of drug delivery, for the anti-cancer drug cisplatin
Traditionally, drugs would be delivered via chemotherapy, however this method is least convenient to the patient. There is no selectivity as with photothermal therapy, and so healthy cells can also be damaged, although because malignant cells have “abnormal” properties, they are more likely to be affected by the drug.2 Loading the drug onto a carrier presents a level of selectivity, so that the drug will only be released once in the region of the target cells, and thus only the malignant cells are destroyed.
The release of a drug upon heating can be defined as “photodynamic therapy”. In order to determine how effective the gold/gold-sulfide nanoparticles were at releasing cisplatin, the nanoparticles were suspended in 2mL of water and irradiated with light from a 1064nm Nd:YAG laser for one hour. Throughout this hour, samples of the gold/gold-sulfide nanoparticle solution were taken and analysed by HPLC in order to analyse how much cisplatin had been released, in addition to samples which had not been irradiated with the laser.
It can be seen from the figure above that the nanoparticles treated with laser radiation were the most effective at releasing cis-platin, with approximately 90% of cisplatin released within the first 10 minutes. The nanoparticles which had been heated to 40C demonstrated some photodynamic effect, with approximately 40% of cisplatin released over the course of an hour. Those which had not been treated with laser light or heat (control) showed virtually no release of cis-platin.
This study has shown the potential for gold/gold-sulfide nanoparticles as drug delivery systems, due to the effective release of cis-platin under NIR light, however this study was conducted entirely in vitro and so further testing in vivo would be suggested in order to fully predict how effectively cisplatin can be delivered this way, inside the body.
The Fe3O4-polymer-gold shell nanoparticles proposed by Wang et al have the potential to be applied in both the diagnosis and treatment of cancer tumours, however no physical studies were carried out involving such an application, and so there is a disadvantage that any application suggested by this literature is entirely speculative.
MRI is a method of diagnostic imaging, with the pictures produced by the process being either T1 weighted, T2 weighted, or proton density weighted, and tumours are visualised as dark areas in T1 weighted images.33 Metal oxide nanoparticles, such as the Fe3O4-polymer-gold shell nanoparticles, can act as a contrasting agent in MRI, thus enhancing the ability to see areas of abnormality in the images produced.34
The fact that the nanoparticles possess both magnetism and NIR absorption makes them especially versatile, as their magnetism would allow application in imaging techniques in MRI and to be moved to the specific region of a tumour via the control of a magnetic field, whilst their NIR absorption would allow application in photothermal therapy of tumours, much like the nanostructures discussed previously.10
The nanoparticles’ theoretical ability to be used simultaneously in both diagnostics and treatment presents a number of advantages, such as quicker, efficient treatment at almost the same time as the diagnosis, and less wastage as it is the same material used in both methods. However, it is important to remember that the potential of these nanoparticles has not been tested, either in vivo or in vitro. It is impossible to predict exactly how a nanoparticle will react once inside the body due to the way in which different environments can affect their activity and toxicity. Further studies of these nanoparticles would be suggested, both in vivo and in vitro, in order to confirm how these nanoparticles will react when in contact with human cells and tissue.
The gold nanorods, conjugated to anti-EGFR and suggested by Huang et al, are particularly promising due to their potential application in both cancer diagnosis and treatment – all nanostructure applications discussed so far have been in cancer therapy only. To test the potential application in cancer diagnosis, three types of cell were incubated with anti-EGFR/gold nanorods and anti-EGFR/gold nanospheres – healthy HaCaT cells (keratinocyte cells found in the epidermis), cancerous HSC cells (hematopoietic stem cells) and cancerous HOC cells (human ovarian carcinoma cells). These cells, incubated with the anti-EGFR/gold nanostructures, were subjected to dark field microscopy in order to scatter light and produce the following images.
Figure 16: Dark field microscopy images of HaCaT, HSC and HOC cells after incubation with anti-EGFR/gold nanospheres (A) and anti-EGFR/gold nanorods. The gold nanospheres strongly scatter green and yellow light, and the gold nanorods strongly scatter red and orange light.
The images above show clearly that both the nanospheres and nanorods bind more strongly to the cancerous cells than the healthy HaCaT cells, and that gold nanorods are more effective at this than gold nanospheres. The clear visual difference in binding means that this is a promising application in the diagnosis of cancer.
In the figure above, it can be seen that at a laser power of 80mW, both HSC and HOC cells incubated with anti-EGFR/gold nanorods begin to suffer damage, and the HaCat cells incubated with anti-EGFR/gold nanorods do not suffer any damage until treatment with a laser of power 120mW. These results are promising as it shows the selectivity of anti-EGFR/gold nanorods in only harming cancerous cells and not normal, healthy tissue. As with several studies previously discussed, however, these results come from an in vitro experiment and should not be taken as confirmation that the same selectivity would be shown inside the human body, or that the nanorods would be as effective in destroying cancerous cells. However, the potential application in cancer diagnosis may not require further in vivo testing as it could be used in the context of a biopsy, which would be carried out in vitro.
Nanotechnology is still a relatively new branch of science that is constantly developing, and as such there are issues that must be considered, such as the impact on the environment and human health. As with any new technology, there is the possibility of individuals abusing it for their own gain and so thought must also be given to how use can be regulated
Before any new product is available for use, especially in the case of nanomedicine, a great deal of testing must be carried out to assess the safety. However, nanoparticles are different from other molecules in that their properties are dependent on size and shape – testing the toxicity of, for example, gold nanoparticles, at a particular diameter or geometry may show that they are not toxic to humans, while a smaller diameter and geometry may be toxic. The environment that nanoparticles are in can also cause changes in diameter thanks to aggregation, so they may behave differently inside a petri dish or even inside an animal, compared to how they would inside a human body – several studies have been undertaken using mice, which, although give an idea of how nanoparticles will react in vivo, cannot be assumed to be fully representative of their reactivity inside humans. This has to be taken into consideration whilst testing for clinical applications.35 However, whilst it is important to move towards human testing, the mental capacity of a test subject must be taken into account – those with learning difficulties or not made fully aware of potential risks could not be considered.36
Thus far, this review has discussed the use of nanomedicine as a diagnostic and therapeutic tool; however, it must be acknowledged that there are rising concerns of nanotechnology being used as a tool for “human enhancement”. The use of medicine to enhance the abilities of a perfectly healthy individual is not new – anabolic steroids, for example, whilst intended for the healing of injuries have been used since the 1970s to enhance performance in sports.37 Although concerns of nanomedicine being used for human enhancement may have been spurred on by science fiction films and books, it is an unfortunate fact that where there is scientific development, there is a possibility of an individual or group of individuals using the advances for their own personal gain. However unrealistic the concerns of nanotechnology for human enhancement are, it is important that the use is still controlled in order to prevent circumstances which would allow such abuse.
In addition to the effect of nanoparticles and nanomedicine on humans, there is also an effect on the environment. Whilst the nanoparticles themselves do not create a great deal of wastage, the volume of materials used in synthesizing them can produce upwards of thousands of tons of wastage each year.
It has already been discussed that the toxicology of nanoparticles can vary immensely depending on their environment, even between animals (for example mice and humans). The study of nanoparticle toxicology with regards to their disposal is something that has not been researched to its full potential and must be explored further before any assumptions can be made about the safety of disposing of them. 36
As with any developing branch of science, careful consideration must also be given to the environmental implications and effect on human health as a result of using gold nanostructures in biomedical applications, as well as ensuring that usage is controlled and available for medical use only, and not for personal advancement.
An area which is in need of further research is that of gold nanostructures in the detection of cancer and tumours. Currently, the majority of studies have focussed on using such structures in the treatment, whether it is photothermal therapy or the release of anti-cancer drugs upon NIR irradiation. Those few studies which have explored the use of nanostructures in diagnosis have simultaneously investigated their use in photothermal therapy. This is now known to be a particularly effective method of treatment, and so it could be suggested that the focus now needs to shift towards developing nanostructures in the diagnosis of cancer and tumours, and applying existing structures to such methods.