Despite the large number of polymeric nanodelivery systems that have been recently developed, there is still room for improvement in terms of therapeutic efficiency. Most reported nanodevices for controlled release are based on drug encapsulation, which can lead to undesired drug leakage with a consequent reduction in efficacy and an increase in systemic toxicity. Herein, we present a strategy for covalent drug conjugation to the nanodevice to overcome this drawback. In particular, we characterize and evaluate an effective therapeutic polymeric PEGylated nanosystem for controlled pH-sensitive drug release on a breast cancer (MDA-MB-231) and two lung cancer (A549 and H520) cell lines. A significant reduction in the required drug dose to reach its half maximal inhibitory concentration (IC50 value) was achieved by conjugation of the drug to the nanoparticles, which leads to an improvement in the therapeutic index by increasing the efficiency. The genotoxic effect of this nanodevice in cancer cells was confirmed by nucleus histone H2AX specific immunostaining. In summary, we successfully characterized and validated a pH responsive therapeutic polymeric nanodevice in vitro for controlled anticancer drug release.

Barcoding and pooling cells for processing as a composite sample are critical to minimize technical variability in multiplex technologies. Fluorescent cell barcoding has been established as a standard method for multiplexing in flow cytometry analysis. In parallel, mass-tag barcoding is routinely used to label cells for mass cytometry. Barcode reagents currently used label intracellular proteins in fixed and permeabilized cells and, therefore, are not suitable for studies with live cells in long-term culture prior to analysis. In this study, we report the development of fluorescent palladium-based hybrid-tag nanotrackers to barcode live cells for flow and mass cytometry dual-modal readout. We describe the preparation, physicochemical characterization, efficiency of cell internalization, and durability of these nanotrackers in live cells cultured over time. In addition, we demonstrate their compatibility with standardized cytometry reagents and protocols. Finally, we validated these nanotrackers for drug response assays during a long-term coculture experiment with two barcoded cell lines. This method represents a new and widely applicable advance for fluorescent and mass-tag barcoding that is independent of protein expression levels and can be used to label cells before long-term drug studies.

MicroRNAs (miRNAs) are considered potential biomarkers due to their specific expression patterns in tissues and their changes in expression under pathological conditions. MicroRNA-122 (miR-122) is a vertebrate-specific miRNA known for its high expression in the liver, where it plays a role in hepatic metabolism, liver development and cellular differentiation. Dysregulation of miR-122 has been found in pathological conditions such as hepatocellular carcinoma or drug-induced liver injury (DILI). Currently, miRNA detection faces challenges such as improving point-of-care efficiency, obtaining direct results and accurately identifying miRNA isoforms. Overcoming these challenges is essential to improve the clinical efficacy of miR-122 detection as a diagnostic and prognostic biomarker in liver-related diseases. In this study, we present a click chemistry driven dual nanosystem designed for single-base specific miR-122 detection using a flow cytometer as the reading platform. Using the dual nanosystem, we analysed miR-122 expression in various tumour cell lines with specificity and robustness, showing a high correlation with results obtained from a TaqMan assay. We also detected miR-122 using a serum matrix and identified four single nucleotide variations at a specific position within the miR-122 sequence. Our approach involves a fully bioorthogonal strategy and includes signal amplification using two types of different nanoparticles together with the high affinity of peptide nucleic acid probes. The chemistry used in this dual nanosystem offers remarkable versatility and provides a promising opportunity to pioneer enzyme-free nanoparticle-based strategies based on click chemistry and bioorthogonality miRNA detection and isomiR studies, thus advancing the field of molecular diagnostics.