Assembly of DNA Networks on the Cell Membrane Enables Confined Detection of MMP-9 Secretion and Evaluation of Drug Efficacy

Proteases secreted by tumor cells have been employed as potential biomarkers for the early assessment of tumor metastasis risk. However, in situ monitoring of their secretion at the single-cell level remains a challenge due to their low abundance and diffusion into complex environments. Despite significant advancements in utilizing DNA as a versatile scaffold for constructing dynamic sensing systems due to its programmability and design flexibility, the spatial control of molecular sensing and imaging functions within living cells is particularly challenging. Herein, we developed a DNA network (termed cAME) by introducing acrylamide copolymers combined with aptamer technology to enable a confined detection of single-cell matrix metalloproteinase-9 (MMP-9) secretion for the assessment of tumor cell invasiveness and drug efficacy evaluation. The cAME was established from a combination of a detecting DNA copolymer (MDDC) containing an MMP-9 responsive aptamer sensor, an encapsulating DNA copolymer (EDC), and a cell membrane anchoring module (chol-A). By monitoring changes in fluorescence signals, real-time detection of MMP-9 activity in individual cells could be achieved. Using various cancer cell lines as models, the feasibility of the proposed confined detection strategy in assessing the heterogeneity of MMP-9 secretion among different subtypes of breast cancer cells was validated. Furthermore, by employing two drugs as models, the application value of the proposed confined detection strategy in rapid and sensitive evaluation of drug efficacy was demonstrated. This strategy contributed to the assessment of tumor metastasis risk while also providing new insights for evaluating the efficacy of antitumor drugs.