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Published on: May 24, 2019
Simultaneous quantification of exosomal MMP14 expression and proteolysis activity on a spherical dual-probe-based
Shuo Yin1, Aipeng Chen2, Xiaoni Fang2
1Institute of Molecular Medicine, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200127, China. cyyang@xmu.edu.cn.
Abstract:
Among various exosomal proteins, matrix metalloproteinases (MMPs) are a family of membrane associated endopeptidases and have been considered as potential biomarkers in liquid biopsy owing to their multiple roles in pathological processes. However, the potential of MMP14 expression (MMP14-E) and MMP14 proteolytic activity (MMP14-A) in clinical diagnosis is still not clear due to the lack of sensitive and simultaneous detection techniques. Herein, we propose a fluorescent nanosensor for the simultaneous detection of MMP14-E and MMP14-A using a spherical aptamer/peptide dual-probe strategy. The aptamer and peptide probes were sequentially immobilized on Fe3O4 magnetic nanoparticles coated with gold nanoparticles (m-AuNPs) using a disulfide linker. MMP14 can be specifically recognized by the aptamer, while the proteolytic-active MMP14 can cleave the peptide probe. While achieving simultaneous detection, the proposed sensor provides better analytical performances than traditional MMP14 sensors owing to the m-AuNP-based spherical dual-probe strategy. This sensor has been successfully applied for the detection of exosomal MMP14 from cell culture media and real serum samples. Levels of both MMP14-E and MMP14-A increase in serum from cancer patients, indicating their potential applications as biomarkers in liquid biopsy for disease diagnosis and real-time surveillance.
Insights
This study introduces a novel fluorescent nanosensor for simultaneously detecting matrix metalloproteinase-14 (MMP14) expression and activity in exosomes. This advancement offers potential for improved liquid biopsy diagnostics in cancer patients.
Area of Science:
- Biomedical Engineering
- Nanotechnology
- Biochemistry
Background:
- Exosomal matrix metalloproteinases (MMPs), particularly MMP14, are implicated in pathological processes and show potential as liquid biopsy biomarkers.
- Current detection methods lack the sensitivity and simultaneous capability for MMP14 expression (MMP14-E) and activity (MMP14-A), hindering clinical application.
Purpose of the Study:
- To develop a sensitive fluorescent nanosensor for simultaneous detection of exosomal MMP14-E and MMP14-A.
- To evaluate the clinical diagnostic potential of MMP14-E and MMP14-A as biomarkers in liquid biopsy.
Main Methods:
- A spherical aptamer/peptide dual-probe strategy was employed, immobilizing probes on magnetic gold nanoparticles (m-AuNPs) via disulfide linkers.
- The nanosensor utilizes aptamer recognition for MMP14 binding and peptide cleavage for proteolytic activity detection.
- Simultaneous detection was achieved using a fluorescent readout on the m-AuNP platform.
Main Results:
- The developed nanosensor demonstrated enhanced analytical performance for simultaneous MMP14-E and MMP14-A detection compared to traditional sensors.
- The sensor successfully detected exosomal MMP14 from cell culture media and human serum samples.
- Elevated levels of both MMP14-E and MMP14-A were observed in serum from cancer patients.
Conclusions:
- The fluorescent nanosensor enables sensitive and simultaneous detection of exosomal MMP14 expression and activity.
- MMP14-E and MMP14-A show promise as valuable biomarkers for liquid biopsy in cancer diagnosis and surveillance.
- This dual-probe strategy offers a significant advancement in exosomal biomarker detection for clinical applications.
