Developing aptamer probes for acute myelogenous leukemia detection and surface protein biomarker discovery

Abstract

BackgroundThe majority of patients with acute myelogenous leukemia (AML) still die of their disease. In order to improve survival rates in AML patients, new strategies are necessary to discover biomarkers for the detection and targeted therapy of AML. One of the advantages of the aptamer-based technology is the unique cell-based selection process, which allows us to efficiently select for cell-specific aptamers without knowing which target molecules are present on the cell surface.MethodsThe NB4 AML cell line was used as the target cell population for selecting single stranded DNA aptamers. After determining the affinity of selected aptamers to leukocytes, the aptamers were used to phenotype human bone marrow leukocytes and AML cells in clinical specimens. Then a biotin-labelled aptamer was used to enrich and identify its target surface protein.ResultsThree new aptamers were characterized from the selected aptamer pools (JH6, JH19, and K19). All of them can selectively recognize myeloid cells with Kd in the low nanomole range (2.77 to 12.37 nM). The target of the biotin-labelled K19 aptamer probe was identified as Siglec-5, a surface membrane protein in low abundance whose expression can serve as a biomarker of granulocytic maturation and be used to phenotype AML. More importantly, Siglec-5 expression can be used to detect low concentrations of AML cells in human bone marrow specimens, and functions as a potential target for leukemic therapy.ConclusionsWe have demonstrated a pipeline approach for developing single stranded DNA aptamer probes, phenotyping AML cells in clinical specimens, and then identifying the aptamer-recognized target protein. The developed aptamer probes and identified Siglec-5 protein may potentially be used for leukemic cell detection and therapy in our future clinical practice.

Selection and characterization of novel DNA aptamers specifically recognized by Singapore grouper iridovirus-infected fish cells

“…In fish, with purified virus particles used as targets, RNA aptamers with therapeutic potential were generated against viral hemorrhagic septicemia virus and Hirame rhabdovirus (Hwang et al, 2012;Punnarak et al, 2012). In a previous study, we selected a panel of DNA aptamers against purified SGIV particles, and demonstrated their inhibitory effects on viral infection in vitro and in vivo (Li et al, 2014 Verdaguer et al, 2014). Based on these modifications, or other markers of virus-infected cells, highly specific molecular probes could be designed.…”

“…However, the number of known SGIV target molecules in cell membranes is low, thereby limiting the development of biomarker probes via traditional antibody-based techniques. Aptamers have been used to recognize targets on and within cell membranes via Cell-SELEX technology (Shangguan et al, 2008a, b;Simaeys et al, 2014;Yang et al, 2014). Shangguan et al (2008a) applied Cell-SELEX technology to leukaemia research and found that protein tyrosine kinase 7 was highly expressed in a series of leukaemia cell lines.…”

“…Aptamers also have been used to recognize targets on and within cell membranes via Cell-SELEX technology (Shangguan et al, 2008a, b;Simaeys et al, 2014;Yang et al, 2014). Yang et al (2014) applied aptamers in acute myelogenous leukaemia detection and determined the target protein of aptamer K19 was sialic acid-binding Ig-like lectin 5 (Siglec-5), which could then be used as a potential biomarker for acute myelogenous leukaemia. Shangguan et al (2008a) applied Cell-SELEX technology to leukaemia research and found that protein tyrosine kinase 7 was highly expressed in a series of leukaemia cell lines.…”

Aptamer for imaging and therapeutic targeting of brain tumor glioblastoma

“…Aptamers have been developed for flow cytometric identification of various tumor cells, such as small cell lung cancer, breast and gastric cancers, acute myelogenous leukemia and glioblastoma among further examples (21,26,(47)(48)(49)(50)(51).…”

Facile Discovery of Cell-Surface Protein Targets of Cancer Cell Aptamers

“…Thus, the identification of target protein(s) for aptamers is a challenging task. Through extraction and affinity purification of proteins of cancer cells with the use of cellrecognition aptamers, protein tyrosine kinase 7 and Siglec-5 were identified as protein targets for aptamers that can bind to T-lineage acute lymphoblastic leukemia cells (8) and acute myelogenous leukemia cells (9), respectively. In addition, an aptamer-facilitated biomarker discovery method was developed for the identification of biomarkers of immature and mature dendritic cells (10).…”