Gold nanoparticle-modified black phosphorus nanosheets with improved stability for detection of circulating tumor cells.

Gold nanoparticle (AuNP)-anchored BP nanosheets were synthesized through in situ growth of AuNPs onto BP. Due to the strong chelating ability of P or phosphorus oxides with AuNPs, the stability of BP is improved. As proof-of-concept demonstration of the functionalized BP, electrochemical detection of circulating tumor cells (CTCs) based on BP@AuNPs@aptamer as a probe combined with immunomagnetic separation is reported. The aptamer can specifically bind with CTCs, while the phosphorus oxides including phosphite ion and phosphate ion (PxOy species) on BP and aptamer can react with molybdate to generate an electrochemical current, leading to dual signal amplification. The biosensor is applied to MCF-7 cell detection and displays good analytical performance with a detection limit of 2 cell mL-1. Furthermore, the practicality of this biosensor was validated through sensitive determination of MCF-7 cells in human blood. Therefore, the reported biosensor could be applied to detect other biomarkers, offering an ultrasensitive strategy for clinical diagnostics. Graphical abstract Electrochemical detection of circulating tumor cells based on gold nanoparticle-modified black phosphorus nanosheets is reported.

Targeted removal of leukemia cells from the circulating system by whole-body magnetic hyperthermia in mice.

Until now, magnetic hyperthermia was used to remove solid tumors by targeting magnetic nanoparticles (MNPs) to tumor sites. In this study, leukemia cells in the bloodstream were directly removed by whole-body hyperthermia, using leukemia cell-specific MNPs. An epithelial cellular adhesion molecule (EpCAM) antibody was immobilized on the surface of MNPs (EpCAM-MNPs) to introduce the specificity of MNPs to leukemia cells. The viability of THP1 cells (human monocytic leukemia cells) was decreased to 40.8% of that in control samples by hyperthermia using EpCAM-MNPs. In AKR mice, an animal model of lymphoblastic leukemia, the number of leukemia cells was measured following the intravenous injection of EpCAM-MNPs and subsequent whole-body hyperthermia treatment. The result showed that the leukemia cell number was also decreased to 43.8% of that without the treatment of hyperthermia, determined by Leishman staining of leukemia cells. To support the results, simulation analysis of heat transfer from MNPs to leukemia cells was performed using COMSOL Multiphysics simulation software. The surface temperature of leukemia cells adhered to EpCAM-MNPs was predicted to be increased to 82 °C, whereas the temperature of free cells without adhered MNPs was predicted to be 38 °C. Taken together, leukemia cells were selectively removed by magnetic hyperthermia from the bloodstream, because EpCAM-modified magnetic particles were specifically attached to leukemia cell surfaces. This approach has the potential to remove metastatic cancer cells, and pathogenic bacteria and viruses floating in the bloodstream.

Preclinical Evaluation of Chimeric Antigen Receptor-Modified T Cells Specific to Epithelial Cell Adhesion Molecule for Treating Colorectal Cancer.

Chimeric antigen receptor-modified T cells (CAR-T cells) have emerged as a promising cancer immunotherapy for solid tumors. Epithelial cell adhesion molecule (EpCAM) is overexpressed in a variety of tumors and is recognized as a biomarker for circulating tumor cells and cancer stem cells, representing an attractive target for adoptive T-cell immunotherapy. This study generated third-generation CAR-T cells with redirected specificity to EpCAM (EpCAM CAR-T) by lentiviral vector. The study demonstrated that EpCAM CAR-T cells can elicit lytic cytotoxicity to target cells in an EpCAM-dependent manner and secrete cytotoxic cytokines, including interferon gamma and tumor necrosis factor alpha. Furthermore, adoptive transfer of EpCAM CAR-T cells significantly delayed tumor growth and formation in xenograft models. In addition, the safety evaluation showed that CAR-T cells have no systemic toxicity in mice. The data confirmed the antitumor ability and safety of CAR-T cells targeting EpCAM and may provide a new target for CAR-T cell therapies in treating solid tumors.

Formulation of nanoparticles ribosome inactivating proteins from Mirabilis jalapa L. (RIP MJ) conjugated AntiEpCAM antibody using low chain chitosan-pectin and cytotoxic activity against breast cancer cell line.

Ribosome Inactivating Proteins (RIPs) isolated from Mirabilis jalapa L. (MJ protein) leaves showed high cytotoxic effect on malignant. Chitosan nanoparticles have frequently been used in protein delivery applications. The aim of this study was to develop targeted drug delivery system of RIP MJ for breast cancer therapy with chitosan nanoparticles conjugated antiEpCAM antibody. RIP MJ nanoparticles were prepared using low viscous chitosan and pectin using polyelectrolit complex method, followed by conjugation process with antiEpCAM antibody. Characterization of this formula was then carried out for its entrapment efficiency, particles size, zeta potential, morphology using transmission electron microscope (TEM) and cytotoxic assay against T47D and Vero cell line. The optimal concentration of MJ protein; low viscous chitosan; pectin for preparing AntiEpCAM conjugated of RIP MJ nanoparticles was 0.1%; 0.01%;1% (m/v) respectively and showed satisfactory formula with the average particle size of 376.8±105.2nm, polydispersity index (PI) 0.401, zeta potential 43,71 mV, high entrapment efficiency 98,97±0,12%. Transmission electron microscope (TEM) imaging showed a spherical and homogenous structure for nanoparticles. The in vitro cytotoxicity analysis showed that RIP MJ nanoparticle had more cytotoxic effect compared to unformulated RIP against T47D cell-lines. AntiEpCAM conjugated RIP MJ nanoparticles however, increased cytotoxic effect of RIPs on Vero cell-lines not for T47D cell-lines. Chitosan-Pectin nanoparticles suitable for delivering protein to target cancer cells.

Formulation and Cytotoxicity of Ribosome-Inactivating Protein Mirabilis Jalapa L. Nanoparticles Using Alginate-Low Viscosity Chitosan Conjugated with Anti-Epcam Antibodies in the T47D Breast Cancer Cell Line.

Ribosome-inactivating protein (RIP) from Mirabilis jalapa L. leaves has cytotoxic effects on breast cancer cell lines but is less toxic towards normal cells. However, it can easily be degraded after administration so it needs to be formulated into nanoparticles to increase its resistance to enzymatic degradation. The objectives of this study were to develop a protein extract of M. jalapa L. leaves (RIP-MJ) incorporated into nanoparticles conjugated with Anti-EpCAM antibodies, and to determine its cytotoxicity and selectivity in the T47D breast cancer cell line. RIP-MJ was extracted from red-flowered M. jalapa L. leaves. Nanoparticles were formulated based on polyelectrolyte complexation using low viscosity chitosan and alginate, then chemically conjugated with anti-EpCAM antibody using EDAC based on carbodiimide reaction. RIP-MJ nanoparticles were characterised for the particle size, polydispersity index, zeta potential, particle morphology, and entrapment efficiency. The cytotoxicity of RIP-MJ nanoparticles against T47D and Vero cells was then determined with MTT assay. The optimal formula of RIP-MJ nanoparticles was obtained at the concentration of RIP-MJ, low viscosity chitosan and alginate respectively 0.05%, 1%, and 0.4% (m/v). RIP-MJ nanoparticles are hexagonal with high entrapment efficiency of 98.6%, average size of 130.7 nm, polydispersity index of 0.380 and zeta potential +26.33 mV. The IC50 values of both anti-EpCAM-conjugated and non-conjugated RIP-MJ nanoparticles for T47D cells (13.3 and 14.9 µg/mL) were lower than for Vero cells (27.8 and 33.6 µg/mL). The IC50 values of conjugated and non- conjugated RIP-MJ for both cells were much lower than IC50 values of non-formulated RIP-MJ (>500 µg/mL).