Generally applicable circulating tumor cell enrichment and identification through a membrane glycoprotein-targeting strategy combining magnetic isolation and biological orthogonality labeling.

Although the importance of circulating tumor cells (CTCs) has been widely recognized, it is still a challenge to realize high-efficiency and accurate enrichment and identification of highly heterogeneous CTCs derived from various types of tumors in complex cancer processes. Currently, the most widely used methods follow the general idea of sequential immunoaffinitive capture and immunostaining to achieve the abovementioned goal. However, different organ/tissue origins as well as the inherent heterogeneity of CTCs would lead to the missed detection of certain CTC subtypes using such methods. Further, immunocytochemistry (ICC) immunostaining disrupts the physiological structure of cells, severely limiting the detection and application scenarios that require the participation of live cells. To address these limitations, we have developed a generally applicable strategy for the isolation and labeling of CTCs. This strategy focuses on targeting the universal characteristics of all tumor cells, specifically the abnormally expressed cell membrane glycoproteins, such as the transferrin receptor and sialic acid. Strategically, transferrin-functionalized magnetic beads (TMBs) were applied to enrich CTCs, and azide-based bioorthogonal chemistry was employed to label target CTCs. Accordingly, the membrane glycoprotein-targeting strategy achieved unbiased enrichment and labeling of broad-spectrum CTCs that were both epithelial and non-epithelial phenotypic populations with varied organ/tissue origins (MCF-7, HepG2, A549, Jurkat, and B16), with a capture efficiency of >95% and a detection limit as low as 5 cells per mL in artificial blood. In particular, our developed strategy displayed excellent specificity, and the CTCs under capture and fluorescence labelling remained with good viability and could be further cultivated and analyzed. Finally, the membrane glycoprotein-targeting strategy successfully detected and identified 33-223 CTCs from 1 mL patient blood samples.

Pro-endothelialization of nitinol alloy cardiovascular stents enhanced by the programmed assembly of exosomes and endothelial affinity peptide.

Stent implantation is one of the most effective methods for the treatment of atherosclerosis. Nitinol stent is a type of stent with good biocompatibility and relatively mature development; however, it cannot effectively achieve long-term anticoagulation and early endothelialization. In this study, nitinol surfaces with the programmed assembly of heparin, exosomes from endothelial cells, and endothelial affinity peptide (REDV) were fabricated through layer-by-layer assembly technology and click-chemistry, and then exosomes/REDV-modified nitinol interface (ACC-Exo-REDV) was prepared. ACC-Exo-REDV could promote the rapid proliferation and adhesion of endothelial cells and achieve anticoagulant function in the blood. Besides, ACC-Exo-REDV had excellent anti-inflammatory properties and played a positive role in the transformation of macrophage from the pro-inflammatory to anti-inflammatory phenotype. Ex vivo and in vivo experiments demonstrated the effectiveness of ACC-Exo-REDV in preventing thrombosis and hyperplasia formation. Hence, the programmed assembly of exosome interface could contribute to endothelialization and have potential application on the cardiovascular surface modification to prevent stent thrombosis and restenosis.

In vitro detection of circulating tumor cells using the nicking endonuclease-assisted lanthanide metal luminescence amplification strategy.

The in vitro detection of circulating tumor cells (CTCs) has been proven as a vital method for early diagnosis and evaluation of cancer metastasis, since the existence and number fluctuation of CTCs have shown close correlation with clinical outcomes. However, it remains difficult and technically challenging to realize accurate CTCs detection, due to the rarity of CTCs in the blood samples with complex components. Herein, we reported a CTCs in vitro detection strategy, utilizing a loop amplification strategy based on DNA tetrahedron and nicking endonuclease reaction, as well as the anti-background interference based on lanthanide metal luminescence strategy. In this work, a detection system (ATDN-MLLPs) composed of an aptamer-functionalized tetrahedral DNA nanostructure (ATDN) and magnetic lanthanide luminescent particles (MLLPs) was developed. ATDN targeted the tumor cells via aptamer-antigen recognition and extended three hybridizable target DNA segments from the apex of a DNA tetrahedron to pair with probe DNA on MLLPs. Then, the nicking endonuclease (Nt.BbvCI) recognized the formed double-strand DNA and nicked the probe DNA to release the target DNA for recycling, and the released TbNps served as a high signal-to-noise ratio fluorescence signal source for CTCs detection. With a detection limit of 5 cells/mL, CTCs were selectively screened throughout a linear response range of low orders of magnitude. In addition, the ATDN-MLLPs system was attempted to detect possible existence of CTCs in biological samples in vitro.

Accurate and noninvasive diagnosis of epithelial cancers through AND gate photoluminescence on tumor-derived small extracellular vesicles.

Noninvasive detection of small extracellular vesicles (sEVs) has become one of the most promising liquid biopsy methodologies for effective and timely cancer diagnosis and prognostic monitoring. Currently, accurate and sensitive detection of tumor-derived sEVs is compromised by their heterogeneous nature, and the tissue origin and parent cell cycle change may significantly affect the tumor-associated information (e.g., phenotypic proteins) of sEVs. Accordingly, many of the single-marker dependent detections on sEVs may not provide comprehensive information about the tumor, and their reliability and clinical applicability cannot be guaranteed. Herein, a strategy for constructing AND gate photoluminescence on tumor-derived sEVs is proposed. Briefly, only after co-recognition of the two epithelial phenotypic proteins (EpCAM and MUC1) on tumor-derived sEVs simultaneously, can our designed lanthanide luminescence probe precursors then assemble to form the AND gate for photoluminescence detection. Consequently, the generated AND gate photoluminescence provided time-resolved luminescence for a wide cancerous sEV linear detection range of 6.0 × 104-6.0 × 109 particles per mL, with a calculated detection limitation of 1.42 × 102 particles per mL. Furthermore, the AND gate photoluminescence can significantly distinguish epithelial cancer patients from healthy controls, displaying its great potential for accurate and noninvasive cancer diagnosis.

Floating Immunomagnetic Microspheres for Highly Efficient Circulating Tumor Cell Isolation under Facile Magnetic Manipulation.

Among circulating tumor cell enrichment strategies, immunomagnetic beads (IMBs) have received great attention due to their excellent performance. However, traditional strategies using IMBs normally require an additional mechanical stirring device to fully mix the IMBs and specimens, and this step may cause mechanical cellular damage. In this study, by changing the architecture and motion trajectory control strategy of the IMBs, floating immunomagnetic microspheres (FIMMs) and their matching rotary magnetic manipulation device were proposed to achieve highly efficient CTC capture under a cell-friendly condition. Generally, the FIMMs were prepared through layer-by-layer assembly of the individual functional components, and their stress state governed by either buoyancy or magnetic force was tuned by the rotary magnetic manipulation device. Consequently, recognition of FIMMs and target cells as well as CTC recovery can be simply realized through external magnetic manipulation. Accordingly, satisfactory enrichment efficiencies for CTCs with varied epithelial expression levels were obtained as 92.93 ± 3.23% for MCF-7, 79.93 ± 3.31% for A549, and 92.57 ± 5.22% for HepG2. Besides, an extremely low detection limitation of 5 cells mL-1 can be achieved from complex sample conditions, even the whole blood. In addition, FIMMs successfully enriched 23-56 CTCs from 1.5 mL of blood samples from cancer patients.

Self-assembled supramolecular immunomagnetic nanoparticles through π-π stacking strategy for the enrichment of circulating tumor cells.

Owing to their high-specific binding toward targets as well as fast and convenient separation operations, immunomagnetic beads (IMBs) are widely used in the capture and detection of circulating tumor cells (CTCs). To construct the IMBs, surface modifications are generally performed to functionalize the magnetic cores (e.g. Fe3O4 nanoparticles), and the employed surface modification strategies normally influence the structure and functions of the prepared IMBs in return. Different from the existing work, we proposed the use of supramolecular layer-by-layer (LBL) self-assembly strategy to construct the IMBs. In general, owing to the π-π stacking interactions, the polydopamine, graphene oxide and 'molecular glue' γ-oxo-1-pyrenebutyric acid were self-assembled on Fe3O4 nanoparticles sequentially, thereby accomplishing the integration of different functional components onto magnetic cores to prepare the self-assembled supramolecular immunomagnetic beads (ASIMBs). The ASIMBs showed high sensitivity, specificity and good biocompatibility to the model CTCs and low nonspecific adsorption to the negative cells (∼93% for MCF-7 cells and 17% for Jurkat cells). Meanwhile, ASIMBs possessed a remarkable potential to screen the rare MCF-7 cells out of large amounts of interfering Jurkat cells with the capture efficiency of 75-100% or out of mouse whole blood with the capture efficiency of 20-90%. The captured cells can be further recultured directly without any more treatment, which showed huge applicability of the ASIMBs for in vitro detection in clinical practices.

Hybrid Extracellular Vesicles-Liposomes Camouflaged Magnetic Vesicles Cooperating with Bioorthogonal Click Chemistry for High-Efficient Melanoma Circulating Tumor Cells Enrichment.

The capture of melanoma circulating tumor cells (melanoma CTCs, MelCTCs) is of great significance for the early diagnosis and personalized treatment of melanoma. The rarity and heterogeneity of MelCTCs have greatly limited the development of MelCTCs capture methods, especially those based on immune/aptamer-affinity. Herein, an extracellular vesicles-camouflaged strategy is designed to functionalize the magnetic nanoparticles (Fe3 O4 ) and to generate magnetic vesicles (Fe3 O4 @lip/ev) with excellent antifouling and active tumor cell targeting properties. Combined with the bioorthogonal click chemistry, the engineered magnetic vesicles with dibenzocyclooctyne can be widely used to target and separate all the metabolically labeled CTCs with varied phenotypes, organ origin, and even the biological species. The capture efficiency exceeded 80% with an extremely low detection limitation of ten cells. Most importantly, the strategy proposed can be directly applied to enrich MelCTCs from 0.5 mL blood samples of melanoma-bearing mice, with a greatly minimized residue of white blood cells (only 21-568) while ignoring the fluctuations of MelCTC phenotype.

Hedgehog-inspired immunomagnetic beads for high-efficient capture and release of exosomes.

Exosomes are small extracellular vesicles secreted by cells. They play an important regulatory role in the physiological and pathological processes of the body, and participate in the occurrence and development of many diseases. Although tumor-derived exosomes have been used as biomarkers for cancer detection, it is still a huge challenge to efficiently capture and release functionally complete exosomes. In our research, inspired by the structure of hedgehog burrs, we proposed immunomagnetic hedgehog particles (IMHPs) to efficiently capture and release exosomes. In general, after the assembly of one-dimensional nanostructural TiO2 bundles into hedgehog TiO2 particles with 356.12 ± 38.32 nm spikes, magnetic responsive nanoparticles (Fe3O4, ∼20 nm), an antifouling polyethylene glycol (PEG) component containing a redox responsive disulfide linkage and anti-CD63 antibody were introduced stepwise to functionalize hedgehog particles and generate IMHPs (1.23 ± 0.18 µm). Due to their unique topological structures, exosomes were positively selected with an exosomal marker (CD63) and negatively selected by depleting environmental pollutants (protein precipitates, cell debris) with the nano-spikes. These prepared IMHPs were successfully applied to capture exosomes from MCF-7 cells, with a capture efficiency of 91.70%. Then, tris (2-carboxyethyl) phosphine hydrochloride (TCEP) was used to reduce the disulfide bond to release exosomes, and the release efficiency was up to 82.45%. The exosomes that experienced successive immunomagnetic separation and release well maintained their structural integrity and good bioactivity to promote MCF-7 cell migration, as compared with those exosomes separated by the classic ultracentrifugation approach. These results also indicated that IMHPs would have broad prospects in biomedicine and clinical applications, where highly efficient and non-destructive separation of bio-substances (cells, extracellular vesicles, etc.) is critically required.

Controllable Environment Protein Corona-Disguised Immunomagnetic Beads for High-Performance Circulating Tumor Cell Enrichment.

The enrichment performance of immunomagnetic beads (IMBs) in blood samples is usually challenging due to the ungoverned, in situ-formed protein corona, as it generally leads to negative effects, such as impeded targeting capacity and unwanted nonspecific absorption. On the contrary, a controlled protein premodification of IMBs with diverse functional environment (blood) proteins endows the composites with a new biological identity and may improve the anti-nonspecific ability, resulting in promising isolation benefits for circulating tumor cell (CTC) enrichment and downstream analyses. Specifically, fetal bovine serum and the four most abundant blood proteins, including human serum albumin, fibrinogen, immunoglobulin, and transferrin, were separately applied in this work. Conclusively, the biological properties of the applied protein corona camouflage have a great influence on the capture performance of IMBs, and certain proteins can enhance the enrichment performance to a large extent. Promisingly, human serum albumin-camouflaged IMBs (HSA-PIMBs) achieved a capture efficiency of 84.0-90.0% and significantly minimized nonspecific absorbed leukocytes to 164-264 in blood samples (0.5 mL, 25-55 model CTCs). Furthermore, HSA-PIMBs isolated 62-505 CTCs and 13-31 leukocytes from the blood samples of five cancer patients. The novel environment camouflage strategy provides a new insight into protein corona utilization and may improve the performance of targeted nanomaterials in a complex biological environment.

Artificial cell membrane camouflaged immunomagnetic nanoparticles for enhanced circulating tumor cell isolation.

Precise and specific circulating tumor cell (CTC) isolation is heavily interfered with by blood cells and proteins. Although satisfactory results have been achieved by some cell membrane-derived platforms, the following limitations have seriously limited the commercialization potential: complex membrane composition, difficult batch difference control, inconvenient source cell expansion, etc. To overcome these limitations, artificial cell membrane camouflage made from commercialized lipids and proteins was proposed in this work. Specifically, a biotinylated phospholipid which can serve as a lipid component and provide active sites (biotin) for antibody modification, and human serum albumin (HSA), which can effectively reduce certain blood protein adsorption, were applied simultaneously to endow our immunomagnetic platform with a new biological identity. Besides, making full use of the robust lipid and protein absorption ability of graphene nanosheets (GNs), the biomimetic cell membrane can be easily integrated into the magnetic core through simple lipid and protein solution incubation. Surprisingly, the resulting artificial cell membrane camouflaged immunomagnetic nanoparticles (AIMNPs) achieved high specificity (average capture efficiency 87.0%), good sensitivity (7 model CTCs per 0.5 mL) and an enhanced anti-nonspecific absorption ability (15-105 white blood cells per 1.5 mL) in both mimic and clinical blood samples.

Immuno-affinitive supramolecular magnetic nanoparticles incorporating cucurbit[8]uril-mediated ternary host-guest complexation structures for high-efficient small extracellular vesicle enrichment.

Enriching small extracellular vesicles (sEVs) with undamaged structure and function is a pivotal step for further applications in biological and clinical fields. It has prompted researchers to explore a carrier material that can efficiently capture sEVs while also gently release the captured sEVs. Here, 1-adamantylamine (1-ADA) responsive immuno-affinitive supramolecular magnetic nanoparticles (ISM-NPs) incorporating ternary host-guest complexation structures mediated by CB[8] were proposed to achieved the goal. In particular, the ternary host-guest complexation was constructed by the host molecule (cucurbit[8]uril, CB[8]) mediated assembly of two guest molecules (naphthol and bipyridine), and served as a cleavable bridge to connect the magnetic core and peripheral antibody. These constructed ISM-NPs performed well in the applications of capturing sEVs with a high capture efficiency of 85.5%. Further, the CB[8]-mediated ternary host-guest complexation structures can be disassembled with addition of the 1-ADA. Thus, the sEVs recognized by the anti-CD63 were released competitively, with a decent release efficiency more than 82%. The released sEVs kept intact morphology and exhibited appropriate size distribution and concentration. This supramolecular magnetic system, with 1-ADA responsive ternary host-guest complexation structures, may contribute to efficient enrichment of any other biomarkers, likely cells, proteins, peptides, etc.

Correction: A light-up fluorescence resonance energy transfer magnetic aptamer-sensor for ultra-sensitive lung cancer exosome detection.

Correction for 'A light-up fluorescence resonance energy transfer magnetic aptamer-sensor for ultra-sensitive lung cancer exosome detection' by Nanhang Zhu et al., J. Mater. Chem. B, 2021, 9, 2483-2493, DOI: 10.1039/D1TB00046B.

Cell-Released Magnetic Vesicles Capturing Metabolic Labeled Rare Circulating Tumor Cells Based on Bioorthogonal Chemistry.

Capture of circulating tumor cells (CTCs) with high efficiency and high purity holds great value for potential clinical applications. Besides the existing problems of contamination from blood cells and plasma proteins, unknown/down-regulated expression of targeting markers (e.g., antigen, receptor, etc.) of CTCs have questioned the reliability and general applicability of current CTCs capture methodologies based on immune/aptamer-affinity. Herein, a cell-engineered strategy is designed to break down such barriers by employing the cell metabolism as the leading force to solve key problems. Generally, through an extracellular vesicle generation way, the cell-released magnetic vesicles inherited parent cellular membrane characteristics are produced, and then functionalized with dibenzoazacyclooctyne to target and isolate the metabolic labeled rare CTCs. This strategy offers good reliability and broader possibilities to capture different types of tumor cells, as proven by the capture efficiency above 84% and 82% for A549 and HepG2 cell lines as well as an extremely low detection limitation of 5 cells. Moreover, it enabled high purity enrichment of CTCs from 1 mL blood samples of tumor-bearing mice, only ≈5-757 white blood cells are non-specific caught, ignoring the potential phenotypic fluctuation associated with the cancer progression.

A magnetic surface-enhanced Raman scattering platform for performing successive breast cancer exosome isolation and analysis.

The rapid development of exosome research provides new insights into the physiological role of exosomes and their significant correlation with human health. Although the exosomes derived from tumor sources have been proven to be promising biomarkers for cancer detection and disease progression due to their inherited biological contents from the parent cancer cells and unique roles in tumor metastasis and invasion, it is still a challenging task to perform rapid and effective isolation from complex biological samples and conduct high-precision real-time analysis. Herein, we propose a magnetic surface-enhanced Raman scattering (SERS) platform to integrate successive breast cancer exosome isolation and Raman signal enhancement into one system to achieve the goal. In addition, principal component analysis (PCA) was conducted to investigate major patterns of the samples. According to the results, the magnetic SERS platform can be applied to distinguish exosomes derived from MCF-7 and MDA-MB-231 cells with 100% sensitivity and 100% specificity for the 95% confidence interval. More importantly, this platform can fully identify breast cancer patients and healthy people with 91.67% sensitivity and 100% specificity. These studies revealed that our magnetic SERS platform would serve as a great potential system for highly efficient real-time liquid biopsy by using the exosomes as cancerous markers, while exempting from pre-treatment of clinical samples or the extra introduction of elements for SERS signal enhancement.

A light-up fluorescence resonance energy transfer magnetic aptamer-sensor for ultra-sensitive lung cancer exosome detection.

In vitro liquid biopsy based on exosomes offers promising opportunities for fast and reliable detection of lung cancers. In this work, we present a fluorescence resonance energy transfer (FRET) magnetic aptamer-sensor for magnetic enrichment of exosomes with aptamers and detection of cancerous-surface proteins based on a light-up FRET strategy. Fluorescent quantum dots (QDs) and aptamers were introduced onto magnetic nanoparticles and the fluorescence emission turned down when the aptamers were paired with their complementary DNA on the surface of Au nanoparticles. Later, competitive binding of exosomes with the aptamers expelled the Au nanoparticles resulting in an exosome concentration-dependent linear increase of QD fluorescence intensity in a broad exosome concentration range (5 × 102-5 × 109 particles per mL). As found in our work, this system behaved ultra-sensitively and the calculated detection limit of this FRET magnetic aptamer-sensor was as low as 13 particles per mL. Furthermore, taking epithelial cancer-specific antigen (epithelial cell adhesion molecule, EpCAM) screening as a typical example, our built FRET magnetic aptamer-sensor allowed a rapid and efficient distinction of all the epithelial cancer cases (7 lung cancers and 5 other cancers) from health volunteers with 100% accuracy.

Bottlebrush-like highly efficient antibacterial coating constructed using α-helical peptide dendritic polymers on the poly(styrene-b-(ethylene-co-butylene)-b-styrene) surface.

Infectious diseases induced by pathogenic bacteria are the major causes for the failure of medical implants. Meanwhile, the drug-resistance is steadily developed because of the large and even inappropriate use of antibiotics. Therefore, the development of antibacterial coating with non-antibiotic-based agents on the surfaces of medical implants and devices has been an urgent need. Herein, we propose a bottlebrush-like antibacterial coating on a poly(styrene-b-(ethylene-co-butylene)-b-styrene) (SEBS) triblock copolymer surface by UV-induced graft polymerization of poly(ethylene glycol) (PEG) acrylate terminated poly(lysine dendrimer). This PEG-conjugated antibacterial polymer possessed a substructure of α-helical backbone and cation dendrimer side chains stretching in the radial directions of the helix. The introduction of lysine peptide dendrimers endowed the prepared antibacterial polymer with precisely controlled characteristics of its local cation density, amphipathic composition as well as three-dimensional (3D) conformation to improve interactions with bacterial membranes. The antimicrobial assay and biocompatibility assay results showed that 96.83% of S. aureus and 99.99% of E. coli were killed after being in contact with the antibacterial coating, while no toxicity to mammalian cells or no hemolysis was detected. This antimicrobial activity was further confirmed by the molecular dynamics simulation results, which demonstrated that the employment of lysine peptide dendrimers enhanced the electrostatic interaction and hydrogen bonding between the brush and bacterial membranes remarkably. Such bottlebrush-like antibacterial coating constructed using α-helical peptide dendritic polymers may become an effective strategy for manufacturing antibacterial products for biomedical uses.

Leukocyte-Repelling Biomimetic Immunomagnetic Nanoplatform for High-Performance Circulating Tumor Cells Isolation.

Downstream studies of circulating tumor cells (CTCs), which may provide indicative evaluation information for therapeutic efficacy, cancer metastases, and cancer prognosis, are seriously hindered by the poor purity of enriched CTCs as large amounts of interfering leukocytes still nonspecifically bind to the isolation platform. In this work, biomimetic immunomagnetic nanoparticles (BIMNs) with the following features are designed: i) the leukocyte membrane camouflage, which could greatly reduce homologous leukocyte interaction and actualize high-purity CTCs isolation, is easily extracted by graphene nanosheets; ii) facile antibody conjugation can be achieved through the "insertion" of biotinylated lipid molecules into leukocyte-membrane-coated nanoparticles and streptavidin conjunction; iii) layer-by-layer assembly techniques could integrate high-magnetization Fe3 O4 nanoparticles and graphene nanosheets efficiently. Consequently, the resulting BIMNs achieve a capture efficiency above 85.0% and CTCs purity higher than 94.4% from 1 mL blood with 20-200 CTCs after 2 min incubation. Besides, 98.0% of the isolated CTCs remain viable and can be directly cultured in vitro. Moreover, application of the BIMNs to cancer patients' peripheral blood shows good reproducibility (mean relative standard deviation 8.7 ± 5.6%). All results above suggest that the novel biomimetic nanoplatform may serve as a promising tool for CTCs enrichment and detection from clinical samples.

Multi-targeting magnetic hyaluronan capsules efficiently capturing circulating tumor cells.

Accurate detection and isolation of circulating tumor cells (CTCs) hold high value for urgent clinical applications in cancer prognosis and treatment monitoring. Even though the immuno-magnetic isolation strategy has been an important milestone for enrichment of precious CTCs from complex clinical samples, the origin and cell cycle change of CTCs would influence greatly on the performance of antibody-mediated capture process. Herein, we proposed the multi-targeting magnetic capsules (TMCs), which combine the natural immune recognition and tumor-specific ligand targeting, for the purpose of rare CTCs capture. Generally, the fluorescence-visible magnetic capsules (MCs) were constructed via an ultra-sonication assembly procedure using fluorescently-labeled thiolated hyaluronan (RhB-HA-SH) and hydrophobic Fe3O4 nanoparticles, during which superparamagnetic Fe3O4 were blocked by RhB-HA-SH cross-linking via oxidation of the thiol groups. Functionalized with folic acid (FA) and anti-epithelial-cell-adhesion-molecule (anti-EpCAM) antibody, the obtained fluorescent-visible multi-targeting hyaluronan magnetic capsules could capture and isolate more than 88% of rare MCF-7 cells in mimic biological samples in 15 min of incubation. Moreover, these isolated cells still showed good proliferation and migration abilities, facilitating further CTCs-related studies. Furthermore, these TMCs were successfully employed to the capture and detection of few CTCs in 1.5 mL cancer patient peripheral blood samples, indicating that they could be a promising candidate for CTCs capture and detection through simple and fast magnetic manipulation.

Multifunctional luminescent immuno-magnetic nanoparticles: toward fast, efficient, cell-friendly capture and recovery of circulating tumor cells.

Highly efficient isolation and recovery of viable circulating tumor cells (CTCs) from the blood of patients is an important precondition to address the current dilemma of insufficient CTC studies, and can promote the development of individualized antitumor therapies. Herein, a cell-friendly CTC isolation and recovery nanoplatform with luminescent labelling was established using a layer-by-layer (LbL) assembly technique and a stimulated cellular-release strategy. In particular, the anti-epithelial cell adhesion molecule (anti-EpCAM) antibody was introduced with a disulfide bond-containing linker for further bio-friendly recovery of the CTCs. Quantum dots (QDs) were deposited onto fast magnet-responsive Fe3O4 nanoparticles through a facile LbL assembly method to monitor the capture and recovery process in real time. The obtained PEGlyated immuno-magnetic nanospheres (PIMNs) can all be magnetically collected within 2 min. Capture efficiencies above 90% can be achieved from blood samples with 5-200 CTCs per mL after only 1-2 min incubation. Nearly all PIMNs on the surface of the CTCs were detached after 15 min of glutathione (GSH) treatment with the disappearance of QD signals. Recovered CTCs could be directly used for culture (cell viability, ∼98%), and their invasiveness and migration characteristics remained unchanged. Furthermore, the PIMNs were successfully applied to isolate CTCs in cancer patients' peripheral blood samples, and an average of 8.6 ± 5.8 CTCs per mL was detected. The results above suggested that PIMNs may serve as a powerful nanoplatform for CTC screening, isolation and recovery.

Detachable Polyzwitterion-Coated Ternary Nanoparticles Based on Peptide Dendritic Carbon Dots for Efficient Drug Delivery in Cancer Therapy.

In this work, we presented ternary nanoparticles [poly(carboxybetaine methacrylate) (pCBMA)(peptide dendrimer-modified carbon dots (CD-D)/doxorubicin (DOX))] based on peptide dendritic carbon dots (CDs) to realize tumor-specific drug delivery and highly efficient cancer therapy. The versatile nanoparticles could achieve "stealth" delivery in blood due to the antifouling zwitterion coating. Meanwhile, charge changes of the zwitterions could be moderated during their transportation toward/inside tumor cells, where subtle environmental pH variations acted as potent stimuli to actualize desired functions. In particular, the detachment of the zwitterionic "coat" at the tumor site resulted in the exposure of abundant peripheral guanidine groups on peptide dendritic carbon dots (CD-D/DOX) owing to the extracellular pH environment (pH 6.8)-induced charge conversion. Consequently, the positively charged CD-D/DOX (+7.02 mV) interacted with the negatively charged cancer cell membrane to enhance cellular uptake. After endocytosis, tumor intracellular microenvironments (acidic conditions and high glutathione (GSH) levels) could lead to effective disintegration of the CD-D/DOX entities due to acid-induced protonation of guanidine groups and glutathione-induced cleavage of peptide dendritic components on CDs, and then effective endosomal escape and fast doxorubicin hydrochloride (DOX·HCl) release (73.2% accumulative release within 4 h) were achieved successively. This strategy enabled a 9.19-fold drug release rate at tumor sites in comparison with the one in the physiological environment. Moreover, the excellent fluorescence properties of CDs endowed the pCBMA(CD-D/DOX) with fluorescence bioimaging function. In view of the above-mentioned advantages, pCBMA(CD-D/DOX) exhibited outstanding antitumor activities both in vitro and in vivo, demonstrating much higher antitumor efficacy and less side effects than the free DOX·HCl.