Abstract
Mammaglobin B (MGB2) and mammaglobin A (MGB1) are proteins expressed in metastatic breast cancers. The early detection of circulating tumor cells (CTCs) in breast cancer patients is crucial to decrease mortality rate. Herein, novel aptamers were successfully selected and characterized against MGB2 and MGB1 proteins using a hybrid SELEX approach. The potential use of the selected aptamers in breast CTC detection was studied using spiked breast cancer cells in whole blood lysate. The results obtained from this study showed that the selected aptamers (MAMB1 and MAMA2) bind to their target breast cancer cell lines with high affinity (low nanomolar Kd values) and specificity. They also bind to their free recombinant target proteins and show minimal non-specific binding to normal and other cancer cell lines. Additionally, they were able to distinguish a low number of breast cancer cells spiked in whole blood lysate containing normal blood cells. The results obtained in this study indicate the great potential for the use of aptamers to detect MGB1 and MGB2 protein biomarkers, expressed on the surface of breast CTCs.
Highlights
Cancer is a complex disease that originates as a result of multiple genomic mutations leading to a disruption of normal cellular homeostasis[1]
The percentage recovery of the ssDNA libraries for both recombinant glutathione S-transferase (GST)tagged MGB2 and MGB1 was monitored by measuring fluorescein emission (520 nm) from aptamer fractions eluted from glutathione modified beads (Fig. 2A and B)
Our results confirmed that the MGB2 and MGB1 proteins are exclusively expressed on the surface of MCF7 and MDA-MB-415 cells respectively, compared to other cell lines used in the Systematic Evolution of Ligands by EXponential enrichment (SELEX) (Supplementary Fig. S1) which confirms the results found in previous studies[13,14]
Summary
Cancer is a complex disease that originates as a result of multiple genomic mutations leading to a disruption of normal cellular homeostasis[1]. Each round of aptamer selection in SELEX is performed by binding and eluting aptamers from target molecules or cells, leading to the selection of aptamers from the library with high affinity and specificity for their targets[32,33]. Compared to their broadly-used antibody counterparts, aptamers possess unique properties in that they can be synthesized and have the ability to be chemically modified, which makes aptamers more convenient to use as molecular probes for various applications[34,35,36]
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