Team:NU Kazakhstan/Modeling

Cancer development is a complex process requiring the coordinated interactions of numerous proteins, signal pathways and cell types. Its detection infers that certain characteristics of the tumor are different from corresponding normal tissue and can be seen and measured as biomarkers of tumorigenesis. Detection of disease is not the same as its diagnosis. Identification of disease does not require recognition of symptoms but also detection of specific features that would indicate the presence of certain disease. Diagnosis based on symptoms is not acceptable for cancer because symptoms usually appear when tumor is large enough to be able to detect. Therefore, finding appropriate cancer biomarkers in minimum amount of biological fluids such as serum, urine and exhaled breath and detecting them in patients would improve diagnosis and prove the presence of cancerous cells. Biomarker is a characteristic that is objectively measured and evaluated as an indicator of normal or pathogenic processes. Appropriate biomarkers may be able to define risks and identify early stages of tumor development, assist in tumor detection and diagnosis, verify stratification of patients for treatment, predict outcomes of the disease and help surveillance for disease recurrence. Improved and timely diagnosis of cancer will positively affect treatment outcomes and increase cancer survival rates. Detecting cancer as early as possible will help to reduce the cancer burden (Etzioni et al., 2003).

Screening tools for biomarkers are needed that exhibit the combined features of high sensitivity and high specificity for early stages of cancers, and which are widely accepted, affordable, and safe to use. There are various types of cancer biomarkers are known such as protein biomarkers, peptide biomarkers, volatile biomarkers, cell based biomarkers, and circulating free DNA biomarkers. Careful selection of these biomarkers in cancer detection is important in diagnosis. Protein biomarkers such as carcinoembryonic antigen (CEA) has been implicated in various types of human cancer and, therefore, will be used as targets of the proposed study. Their combination in one simple, easy and portable test detection assay or test system would generate a fast diagnostic methodology. CEA is a 201 kDa glycoprotein that is involved in a cell adhesion. Its specialized sialofucosylated glycoforms serve as functional L-selectin and E-selectin ligands which are critical to the metastatic dissemination for cancer cells. Serum CEA is currently one of the most relevant and widespread tumor markers of colorectal cancer (Christenson et al., 2011). It is used for prognosis and monitoring in colorectal cancer patients (Sturgeon et al., 2008). The use of CEA for screening and early diagnosis is limited due to a lack of diagnostic sensitivity and specificity (Smith et al., 2002).

Biosensors and immunoassays as more recent methods for the rapid detection of single-tumor markers have been well progressed for cancer diagnosis. According to the International Union of Applied and Pure Chemistry, a biosensor is a sensor composed of biological recognition elements (e.g., antibodies, enzymes, or aptamers) whose interaction with their analytes is detected with a transducer. The transducer is a device that converts the chemical or physical signals into something measurable such as an electrical signal (Thevenot et al., 1999). Biosensors consisting of various types of transducers, such as optical, electrochemical, mass-based or calorimetric were designed for detection of either single or multiple tumor markers. Biosensors for the detection of CEA, IL-6, and autoantibodies to ECPKA are reviewed in Rusling et al. (2010), Tothill et al. (2009), Arya and Bhansali (2011), and Tan et al. (2009). Most of such biosensors use antibodies as biorecognition element. As for DNA aptamers, Wang et al. (2007) selected aptamers against CEA, although the quantitative data on aptamer affinity towards its target were not presented.

Application of tumor markers for cancer diagnosis is facing great challenges, because most markers are not specific to a particular tumor and no single marker can be used for accurately predicting disease in all of its stages. Panels of cancer biomarkers can improve their diagnostic value in complex biological samples. Therefore, the development of highly sensitive and selective sensors capable of simultaneous detection of multiple analytes has attracted much attention. Compared with the traditional single-analyte immunoassay, the simultaneous multiplexed immunoassay is more efficient in clinical application since it can quantitatively detect a panel of biomarkers in a single run with improved diagnostic specificity (Tian et al., 2012). Moreover, the multiplexed immunoassay can shorten analytical time, enhance detection throughput, and decrease sampling volume and detection costs. The approach proposed in this work combines the advantage of using aptamers for recognition of specific cancer biomarkers, magnetic nanoparticles for separation and quantum dots for detection, resulting in a novel, portable, and rapid competitive tool for sensitive and selective multiplexed detection of cancer biomarkers. Since biosensors for biomarker detection involve a biological recognition element, it is necessary to develop an ideal candidate possessing advantages over traditional antibodies, and aptamers can be a good example of it. Aptamers are short single stranded DNAs (ssDNAs) or RNAs that have an ability to bind to various targets with high affinity and specificity and can be developed by way of repetitive cycles of affinity selection and PCR amplification. Being an emerging group of recognition elements, aptamers hold significant advantages over antibodies such as they do not require a host animal for production since in vitro combinatorial biochemistry is applied in this process, exhibit high binding affinities for their targets, and are resistant to biodegradation and denaturation. To use aptamers versus antibodies in our experiments will be vital since aptamers have been shown to distinguish intimately related substances from their targets on the basis of minor structural changes, such as a methyl group, a hydroxyl group, and a urea vs. a guanidine group. Moreover, there is one more big advantage of aptamer application in biosensors. The capability to regenerate the function of immobilized aptamers would be the most attractive characteristic of aptamers. Being nucleic acids, aptamers could be exposed to repeated cycles of denaturation and renaturation. Heat, salt concentration, pH of the medium, and chelating agents could work as aptamer regeneration methods (Jayasena, 1999). According to experience of Bruno et al. (2009), the DNA aptamer–MB and aptamet–QD sandwich assay components remain adherent to the inner surface of the polystyrene cuvette for days to weeks and they have much stronger adherence in comparison to antibody–MBs, which fall away from the collection site when the magnetic insert is removed, but a very thin brown film sometimes remain. They explain it with the fact that antibody coated–MBs adhere to polystyrene cuvettes with lesser affinity at neutral pH and most proteins adhere optimally to polystyrene microtiter plates only at elevated pH. Given that aptamer based optical biosensors represent an unexplored field, we expect many exciting opportunities for aptamer based bioelectronic devices.