Accurate detection and imaging of tumor-related mRNA in living cells hold great promise for early cancer detection. However, currently, most probes designed to image intracellular mRNA confront intrinsic interferences arising from complex biological matrices and resulting in inevitable false-positive signals. To circumvent this problem, an intracellular DNA nanoprobe, termed DNA tetrahedron nanotweezer (DTNT), was developed to reliably image tumor-related mRNA in living cells based on the FRET (fluorescence resonance energy transfer) “off” to “on” signal readout mode. DTNT was self-assembled from four single-stranded DNAs. In the absence of target mRNA, the respectively labeled donor and acceptor fluorophores are separated, thus inducing low FRET efficiency. However, in the presence of target mRNA, DTNT alters its structure from the open to closed state, thus bringing the dual fluorophores into close proximity for high FRET efficiency. The DTNT exhibited high cellular permeability, fast response and excellent biocompatibility. Moreover, intracellular imaging experiments showed that DTNT could effectively distinguish cancer cells from normal cells and, moreover, distinguish among changes of mRNA expression levels in living cells. The DTNT nanoprobe also exhibits minimal effect of probe concentration, distribution and laser power as other ratiometric probe. More importantly, as a result of the FRET “off” to “on” signal readout mode, the DTNT nanoprobe almost entirely avoids false-positive signals due to intrinsic interferences, such as nuclease digestion, protein binding and thermodynamic fluctuations in complex biological matrices. This design blueprint can be applied to the development of powerful DNA nanomachines for biomedical research and clinical early diagnosis.

Biomimetic giant membrane vesicles, with size and lipid compositions comparable to cells, have been recognized as an attractive experimental alternative to living systems. Due to the similarity of their membrane structure to that of body cells, cell-derived giant plasma membrane vesicles have been used as a membrane model for studying lipid/protein behavior of plasma membranes. However, further application of biomimetic giant membrane vesicles has been hampered by the side-effects of chemical vesiculants and the utilization of osmotic buffer. We herein develop a facile strategy to derive giant membrane vesicles (GMVs) from mammalian cells in biofriendly medium with high yields. These GMVs preserve membrane properties and adaptability for surface modification and encapsulation of exogenous molecules, which would facilitate their potential biological applications. Moreover, by loading GMVs with therapeutic drugs, GMVs could be employed for drug transport to tumor cells, which represents another step forward in the biomedical application of giant membrane vesicles. This study highlights biocompatible GMVs with biomimicking membrane surface properties and adaptability as an ideal platform for drug delivery strategies with potential clinical applications.

In vivo noninvasive molecular imaging requires precise recognition and in situ, real-time imaging of specific cellular and molecular signatures at the site of interest. However, this is often hindered by issues of current imaging probes relating to either the lack of active recognition or the overall nonspecific mechanism of action. Here, we present an aptamer-signal base conjugate (ApSC) concept to engineer AND-gate molecular tools for tumor-targeted molecular imaging. Superior to conventional synthetic methods for imaging probes, our design enables programmable and precise conjugation between recognition and signaling units in a modular synthesis manner with high fidelity for both the conjugating chemistry and binding affinity to the molecular target. Moreover, this design is endowed with simultaneous multivariate activation that readily adapts to tumor microenvironments for signal output, thus providing improved imaging specificity and sensitivity. Such a concept has been successfully shown in magnetic resonance imaging (MRI), the modality of choice for in vivo noninvasive molecular imaging. The engineered ApSC can produce amplified MR signals only after activation by the unique metabolism and dysregulation of redox balance in cancer. In mouse models of xenograft and metastatic breast cancer, the AND-gate molecular MRI probe elicits high imaging contrast in primary tumors and micrometastases. This study promises to provide synthetically accessible scaffolds that can be extended to a large library of advanced molecular imaging tools with varied imaging modalities and mechanisms of action for preventative, predictive, and personalized medicine.

Membrane vesicles derived from live cells show great potential in biological applications due to their preserved cell membrane properties. Here, we demonstrate that cell-derived giant membrane vesicles can be used as vectors to deliver multiple therapeutic drugs and carry out combinational phototherapy for targeted cancer treatment. We show that therapeutic drugs can be efficiently encapsulated into giant membrane vesicles and delivered to target cells by membrane fusion, resulting in synergistic photodynamic/photothermal therapy under light irradiation. This study highlights biomimetic giant membrane vesicles for drug delivery with potential biomedical application in cancer therapeutics.

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