UHN UT University of Toronto University Health Network

 

Aptamers

 

Lab Members: Aaron Prodeus, Nicholas Fischer, Mays Alwash, Marzena Cydzik

 

What is an Aptamer?

“Aptamers” are single stranded nucleic acids (RNA or ssDNA) which bind to molecular targets of interest with high affinity and high specificity. Such sequences, which are generally 15-90 bases in length, are derived to a selected target through an iterative in vitro selection process known as SELEX (Figure 1). These aptamers bind to target molecules such as proteins with comparable affinity (KD) to monoclonal antibodies. In contrast to monoclonal antibodies however, aptamers have distinct advantages, including 1) low immunogenicity, 2) inexpensive chemical synthesis, and 3) long shelf life. Furthermore, SELEX can be tailored to evolve aptamers with selected characteristics, such as fast association rates (Ka) or slow dissociation rates (Kd) in selected environmental conditions (temperature, pH, ion concentrations, etc). Lastly, chemical moieties can be readily conjugated to defined sites on aptamers, enabling the use of aptamers in a multitude of applications including drug targeting, imaging, biosensors, bioanalysis, immunohistochemistry, and in vivo drug targeting.

Functional Aptamers as Therapeutic Agents

Our lab has recently been interested in the development of functional DNA aptamers - sequences which exhibit antagonistic or agonistic effects on cellular signalling pathways. Antagonistic aptamers function by sterically blocking the engagement of selected cell surface receptors to their cognate ligands, thereby inhibiting a signalling pathway. As many diseases, including cancer, depend on deregulated signalling pathways, such functional aptamers can represent novel treatments. Notably, our laboratory has developed antagonistic or blocking aptamers towards carcinoembryonic antigen (CEA) and tumor necrosis factor alpha (TNFα). Current efforts involve the continued development of both agonistic and antagonistic aptamers to reagent targets implicated in cancers or other diseases.

 

Figure 1. SELEX: Systematic Evolution of Ligands by Exponential Enrichment is used to evolve aptamers towards targets of therapeutic relevance.

 

Equipment and Techniques

The “Molecular Targeting and Therapeutics lab” on the 7th floor of the M-Wing in Sunnybrook Hospital is home to a number of important pieces of equipment for rapid aptamer development and characterization. Briefly, our laboratory hosts an on-site DNA synthesizer for aptamer synthesis, high performance liquid chromatraphy (HPLC) for purification of aptamers and conjugates, as well as a Biacore T200 Surface Plasmon Resonance for accurate characterization of aptamer binding affinity and kinetics. (See “Equipment” section for further details)

 

 

Selected References

1) Prodeus A, A Abdul-Wahid, NW Fischer, EH-B Huang, M Cydzik, Gariépy J. (2015) Targeting the PD-1/PD-L1 immune evasion axis with DNA aptamers as a novel therapeutic strategy for the treatment of disseminated cancers. Mol Ther Nucleic Acids. 4:e237 aaron-2

2) Prodeus A, Cydzik M, Abdul-Wahid A, Huang EH, Khatri I, Gorczynski R, Gariépy J. (2014) Agonistic CD200R1 DNA aptamers are potent immunosuppressants that prolong allogeneic skin graft survival. Mol Ther Nucleic Acids. 3:e190 Aaron1

3) Orava EW, Abdul-Wahid A, Huang EH, Mallick AI, Gariépy J. (2013) Blocking the attachment of cancer cells in vivo with DNA aptamers displaying anti-adhesive properties against the carcinoembryonic antigen. Mol Oncol. 7(4):799-811.ORAVA1

4) Orava EW, Jarvik N, Shek YL, Sidhu SS, Gariépy J. (2013) A short DNA aptamer that recognizes TNFα and blocks its activity in vitro. ACS Chem Biol. 18:8(1):170-178. Orava2

5) Orava EW, Cicmil N, and Gariépy J. (2010) Delivering Cargoes into cancer cells using DNA aptamers targeting internalized surface portals. Biochim Biophys Acta 1798:2190-2200. ORAVA3

 

 

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