Seek and destroy
9 Dec 2014 by Evoluted New Media
The use of nanoparticles in cancer research is considered to be a promising approach in the fight against tumour cells. Yet the method often fails because the human immune system recognises the particles as foreign objects and rejects them before they can fulfil their function. Could nanoparticles prove to be the ‘magic bullet’ needed for efficient detection and treatment of tumour cells?
Researchers at the Helmholtz-Zentrum Dresden-Rossendorf (HZDR) and University College Dublin have, along with other partners, developed nanoparticles that find their way to the diseased cells. This procedure uses fragments from a particular type of antibody that only occurs in camels and llamas. The small particles were even successful under conditions which are very similar to the situation within potential patients’ bodies.
With 14.1 million new cancer cases, 8.2 million cancer deaths and 32.6 million people living with cancer in 2012 worldwide1, this disease represents one of the most serious health problems of our society. However, the chances for successful treatment are substantially increased if tumours and metastatic cells are detected at an early stage. Among various diagnostic tests, nuclear medicine imaging techniques, such as positron emission tomography (PET), allow functional analyses of processes in the body and quantitative evaluation of cancerous tissues in a non-invasive manner. These diagnostic techniques require the intravenous injection of radiopharmaceutical tracers into the patient’s body in very small amounts, which become subsequently concentrated in diseased tissues.
In clinical oncology, such radiotracers consist of a short-lived radioisotope for detection and a “transporting” biological vector molecule that guides the radiolabel with high selectivity to the malignant tissue. This concept takes advantage of typical characteristics of cancer cells and their environment such as increased glucose consumption, low oxygen levels and low pH values2. Furthermore, malignant cells often overexpress some membrane receptors at their surface, which enable them to grow and multiply uncontrollably. Specific radiolabelled peptides or antibodies directed against these so-called tumour markers can be used to accumulate the radiotracers at the tumour site allowing their tomographic detection (Figure 1). However, this active targeting strategy presupposes that they only enrich at the target and that the radiotracers are excreted rapidly once they accomplished their tracking task3.
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Figure 1: Positron emission tomography is a nuclear medicine imaging technique that produces three-dimensional images of functional processes in the body.[/caption]
Both conditions – rapid blood clearance as well as high target specificity – are fulfilled by a novel type of antibody fragments originating from camels and llamas. In contrast to conventional antibodies, which consist of two light and two heavy protein chains, these are less complex and are made up of only two heavy chains4. Furthermore, only a small fragment of these camelid antibodies, a so-called single-domain antibody-fragment (sdAbs), is required to achieve high target specificity. Within the last ten years scientists have shown that these fragments offer several advantages for radiotracer applications. In fact, radiolabelled sdAbs generally show rapid and specific tumour uptake and fast clearance from the body permitting a diagnostic scan within the first hours after radiotracer injection5.
Due to their simplified structure, sdAbs are more facile to synthesise than normal antibodies6. Scientists from the Institute of Radiopharmaceutical Cancer Research located at the HZDR recently described an economic method to produce substantial amounts of soluble and functional sdAbs. They manipulated a genetically engineered strain of Escherichia coli in a way that it synthesises up to 200 mg of the camelid antibody fragment per litre of culture. “For us, E. coli is truly the workhorse of protein expression, producing more than enough antibody fragments to perform a multitude of experiments,” explains Dr Kristof Zarschler, who is responsible for sdAb production and purification. For their study, the Dresden researchers have chosen sdAbs which specifically bind to the epidermal growth factor receptor (EGFR). This cell-surface molecule is abnormally expressed and aberrantly activated in various types of tumours, resulting in unregulated growth stimulation and promotion of tumour processes including angiogenesis and metastasis. Using different human cancer cell lines, the German scientists confirmed the high affinity and specificity of the expressed sdAbs towards its molecular target EGFR7.
Once these antibody fragments were identified as suitable candidates for tracking certain tumour cells the research was taken a major step further: together with a collaborating group from the Centre For BioNano Interactions (CBNI) at the University College Dublin (UCD) they equipped fluorescent nanoparticles with EGFR-specific sdAbs.
“The strategy of using functionalised nanoparticles as non-invasive diagnostic tools has several advantages,” says Dr Holger Stephan, who is spokesperson of the Helmholtz Virtual Institute “NanoTracking” dealing with the development of novel multimodal nanoparticle-based probes for early diagnosis of cancer. “Most importantly, one may fine-tune the properties of the materials in a way that the nanoparticles distribute in the body favourably.”
To achieve this, the collaborating scientists from Australia, Ireland and Germany have to face three challenges, says Stephan. “First, we need to establish the smallest possible nanoparticles. We then need to modify their surface in a way that the proteins in the human bodies do not envelop them, which would thus render them ineffective. In order to ensure, that the particles do their job, we must also somehow program them to find the diseased cells.”
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Figure 2: Nanoparticles functionalised with camelid single-domain antibodies bind specifically to certain tumour markers located on the surface of malignant cells.[/caption]
For tackling the latter issue, the researchers use the camelid antibody fragment, which they grafted onto the surface of the nanoparticles8. So they could demonstrate that nanoparticles that have been combined with sdAbs can more firmly bind to the cancer cells (Figure 2). They even obtained the same results in experiments involving human blood serum – a biologically relevant environment the scientists point out: “This means that we carried out the tests under conditions that are very similar to the reality of the human body,” explains Dr Eugene Mahon from the CBNI in Dublin. “The problem with many current studies is that artificial conditions are chosen where no disruptive factors exist. While this provides good results, it is ultimately useless because the nanoparticles fail finally in experiments conducted under more complex conditions9. In our case, we could at least reduce this error source.”
However, more time is required before the nanoparticles can be utilised in diagnosing human tumours. “The successful tests have brought us one step further,” says Stephan. “The road, however, to its clinical use is long.” The next aim is to reduce the size of the nanoparticles, which are now approximately 50 nanometres in diameter, to less than ten nanometres; “that would be optimal,” according to Zarschler. “Then they would only remain in the human body for a short period – just long enough to detect the tumour.”
Acknowledgements
Financial support by the Helmholtz Virtual Institute NanoTracking (Agreement Number VH-VI-421) is gratefully acknowledged. This study is part of a research initiative “Technologie und Medizin – Multimodale Bildgebung zur Aufklärung des in vivo Verhaltens von polymeren Biomaterialien” of the Helmholtz-Portfoliothema. Financial support through Science Foundation Ireland and the Irish Research Council (IRC) are gratefully acknowledged. The work performed was supported by the IRC through an Enterprise Partnership Postdoctoral Fellowship with Intel (Ireland) (Ref no: EPSPD/2012/443) and the IRCSET EMPOWER Postdoctoral Fellowship Scheme. Experimental method development supported through the QualityNano research infrastructure is acknowledged.
References
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