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Updated: Jan 6, 2026
A Flow Cytometry-Based Cell Surface Protein Binding Assay for Assessing Selectivity and Specificity of an Anticancer Aptamer
Published on: September 13, 2022
Hypoxia-Activated PEGylated Conditional Aptamer/Antibody for Cancer Imaging with Improved Specificity
Fang Zhou1, Ting Fu1, Qin Huang1
1Molecular Science and Biomedicine Laboratory (MBL), State Key Laboratory for Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, and Aptamer Engineering Center of Hunan Province , Hunan University , Changsha 410082 , P. R. China.
Abstract:
Aptamers and antibodies, as molecular recognition probes, play critical roles in cancer diagnosis and therapy. However, their recognition ability is based on target overexpression in disease cells, not target exclusivity, which can cause on-target off-tumor effects. To address the limitation, we herein report a novel strategy to develop a conditional aptamer conjugate which recognizes its cell surface target, but only after selective activation, as determined by characteristics of the disease microenvironment, which, in our model, involve tumor hypoxia. This conditional aptamer is the result of conjugating the aptamer with PEG5000-azobenzene-NHS, which is responsive to hypoxia, here acting as a caging moiety of conditional recognition. More specifically, the caging moiety is unresponsive in the intact conjugate and prevents target recognition. However, in the presence of sodium dithionite or hypoxia (<0.1% O2) or in the tumor microenvironment, the caging moiety responds by allowing conditional recognition of the cell-surface target, thereby reducing the chance of on-target off-tumor effects. It is also confirmed that the strategy can be used for developing a conditional antibody. Therefore, this study demonstrates an efficient strategy by which to develop aptamer/antibody-based diagnostic probes and therapeutic drugs for cancers with a unique hypoxic microenvironment.
Insights
This study introduces a conditional aptamer conjugate that activates only in hypoxic tumor environments, reducing off-tumor effects for cancer diagnosis and therapy. This hypoxia-activated probe enhances targeted cancer treatment strategies.
Area of Science:
- Bioconjugation Chemistry
- Molecular Oncology
- Biomedical Engineering
Background:
- Aptamers and antibodies are crucial for cancer detection and treatment but can cause off-tumor effects due to non-specific binding.
- Target recognition relies on overexpression, not exclusivity, leading to limitations in current diagnostic and therapeutic probes.
Purpose of the Study:
- To develop a novel conditional aptamer conjugate for targeted cancer therapy and diagnosis.
- To engineer a probe that selectively activates in the tumor microenvironment, specifically under hypoxic conditions.
Main Methods:
- Conjugating aptamers with PEG5000-azobenzene-NHS to create a hypoxia-responsive caging moiety.
- Investigating the activation mechanism of the caging moiety in response to hypoxia (<0.1% O2) and sodium dithionite.
- Evaluating the conditional recognition of cell surface targets and potential for antibody conjugation.
Main Results:
- The conditional aptamer conjugate remained inactive until exposed to hypoxia or reducing agents.
- Hypoxia triggered the release of the caging moiety, enabling target recognition.
- The strategy proved effective for developing conditional antibodies, demonstrating broad applicability.
Conclusions:
- A novel strategy for conditional aptamer/antibody probes was successfully developed.
- Hypoxia-activated targeting significantly reduces on-target, off-tumor effects in cancer therapy.
- This approach offers a promising method for developing targeted cancer therapeutics and diagnostics for hypoxic tumors.
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