9 Aptamers for the Diagnosis of Malign Tumors
241
However, the identification and detection of protein biomarkers still face challenges.
Because there are nonspecific overlaps between the malign tumors and healthy
tissues, intra-individual mutations in the proteome, as well as the complexity and low
abundance of biological fluid components [15]. Therefore, the analysis methods for
protein biomarkers with high sensitivity, selectivity, reproducibility, and repeatability
are urgently needed.
Several classical techniques have been used for protein biomarkers discovery, such
as mass spectrometry and its related system, and affinity-based proteomic assays
[16]. However, these methods suffer from several disadvantages like poor reproducibility, high costs, and the inherent defect of antibodies. Recently, aptamer-based
bio-affinity proteomic assays have been developed, such as enzyme-linked aptamer
assay (ELAA). The ELAA method depends on a sandwich-type assay containing
aptamers as specific binding elements, and exhibit superior sensitivity and selectivity comparable to enzyme-linked immunosorbent processes [17]. Also, several
excellent aptamer-based methods have appeared for cancer biomarkers discovery.
SELEX is a useful technique for cancer biomarkers discovery. For classical
SELEX, it is necessary to take an oligomers library for the identification of potential targets, and the target entities could be molecules, cells, and even tissues. In
protein-SELEX, the known targets should be collected from cancer secretomes [18],
or recombinant protein through purified processes
16] . After negative select steps, the
unbound oligomers incubate with the designed targets, and the binding aptamers are
finally harvested. Taking this strategy, several cancer biomarkers have been identified,
like cyclophilin B [18]. However, this method suffers from some drawbacks. The ‘of
target’ proteins should be the known ones, rendering the potential biomarkers loss.
Besides, the purification step might change the conformation of the isolated protein,
especially the physiologically binding domains, resulting in nonspecific adsorption.
As an improvement method, cell-SELEX could harvest aptamers binding on
unknown protein targets. In this procedure, aptamers could be selected from the
cancer cell surface biomarkers, without losing their native structures [19]. In this
case, the targets remain better representation so that they could provide more details
on disease states. Moreover, the prior knowledge of the structure or expression
levels of cell membrane protein is not necessary anymore, which facilitates the
discovery of new biomarkers [20]. The cancer cells and normal ones could be
used as targets for positive and negative selection, respectively, and the aptamers
and their bound molecules could be identified at the same time (Fig. 9.1). Several
cancer biomarkers have been discovered successful, then used in cancer diagnosis
and therapy. Using cell-SELEX, Larry Gold group identified a glioblastoma-derived
cell line U251-targeting DNA aptamer GBI-10, and then further obtained protein
biomarker tenascin-C [21]. Aptamer sgc8 binds to the protein tyrosine kinase 7
(PTK7) on the cell surface of T-cell acute lymphoblastic leukemia [22], and aptamer
TD05 identifies Burkitt’s lymphoma through binding immunoglobulin heavy μ chain
molecules [23]. Also, aptamer AS1411, HCHO7, and Apt-32 isolated using the cellSELEX are confirmed to target the biomarkers on the surface of specific cancer cells
[24]. These aptamers have already been further used in cancer therapy, as drugs or
siRNA delivery. However, it is not always successful to isolate cancer biomarkers
241
However, the identification and detection of protein biomarkers still face challenges.
Because there are nonspecific overlaps between the malign tumors and healthy
tissues, intra-individual mutations in the proteome, as well as the complexity and low
abundance of biological fluid components [15]. Therefore, the analysis methods for
protein biomarkers with high sensitivity, selectivity, reproducibility, and repeatability
are urgently needed.
Several classical techniques have been used for protein biomarkers discovery, such
as mass spectrometry and its related system, and affinity-based proteomic assays
[16]. However, these methods suffer from several disadvantages like poor reproducibility, high costs, and the inherent defect of antibodies. Recently, aptamer-based
bio-affinity proteomic assays have been developed, such as enzyme-linked aptamer
assay (ELAA). The ELAA method depends on a sandwich-type assay containing
aptamers as specific binding elements, and exhibit superior sensitivity and selectivity comparable to enzyme-linked immunosorbent processes [17]. Also, several
excellent aptamer-based methods have appeared for cancer biomarkers discovery.
SELEX is a useful technique for cancer biomarkers discovery. For classical
SELEX, it is necessary to take an oligomers library for the identification of potential targets, and the target entities could be molecules, cells, and even tissues. In
protein-SELEX, the known targets should be collected from cancer secretomes [18],
or recombinant protein through purified processes
16] . After negative select steps, the
unbound oligomers incubate with the designed targets, and the binding aptamers are
finally harvested. Taking this strategy, several cancer biomarkers have been identified,
like cyclophilin B [18]. However, this method suffers from some drawbacks. The ‘of
target’ proteins should be the known ones, rendering the potential biomarkers loss.
Besides, the purification step might change the conformation of the isolated protein,
especially the physiologically binding domains, resulting in nonspecific adsorption.
As an improvement method, cell-SELEX could harvest aptamers binding on
unknown protein targets. In this procedure, aptamers could be selected from the
cancer cell surface biomarkers, without losing their native structures [19]. In this
case, the targets remain better representation so that they could provide more details
on disease states. Moreover, the prior knowledge of the structure or expression
levels of cell membrane protein is not necessary anymore, which facilitates the
discovery of new biomarkers [20]. The cancer cells and normal ones could be
used as targets for positive and negative selection, respectively, and the aptamers
and their bound molecules could be identified at the same time (Fig. 9.1). Several
cancer biomarkers have been discovered successful, then used in cancer diagnosis
and therapy. Using cell-SELEX, Larry Gold group identified a glioblastoma-derived
cell line U251-targeting DNA aptamer GBI-10, and then further obtained protein
biomarker tenascin-C [21]. Aptamer sgc8 binds to the protein tyrosine kinase 7
(PTK7) on the cell surface of T-cell acute lymphoblastic leukemia [22], and aptamer
TD05 identifies Burkitt’s lymphoma through binding immunoglobulin heavy μ chain
molecules [23]. Also, aptamer AS1411, HCHO7, and Apt-32 isolated using the cellSELEX are confirmed to target the biomarkers on the surface of specific cancer cells
[24]. These aptamers have already been further used in cancer therapy, as drugs or
siRNA delivery. However, it is not always successful to isolate cancer biomarkers