Zwitterionic nanoparticles for thermally activated drug delivery in hyperthermia cancer treatment.

Hyperthermia is considered a promising strategy to boost the curative outcome of traditional chemotherapeutic treatments. However, this thermally mediated drug delivery is still affected by important limitations. First, the poor accumulation of the conventional anticancer formulations in the target site limits the bioavailability of the active ingredient and induces off-site effects. In addition, some tumoral scenarios, such as ovarian carcinoma, are characterized by cell thermotolerance, which induces tumoral cells to activate self-protecting mechanisms against high temperatures. To overcome these constraints, we developed thermoresponsive nanoparticles (NPs) with an upper critical solution temperature (UCST) to intracellularly deliver a therapeutic payload and release it on demand through hyperthermia stimulation. These NPs were synthesized via reversible addition-fragmentation chain transfer (RAFT) emulsion polymerization and combine polyzwitterionic stabilizing segments and an oligoester-based biodegradable core. By leveraging the pseudo-living nature of RAFT polymerization, important physicochemical properties of the NPs were controlled and optimized, including their cloud point (Tcp) and size. We have tuned the Tcp of NPs to match the therapeutic needs of hyperthermia treatments at 43 °C and tested the nanocarriers in the controlled delivery of paclitaxel, a common anticancer drug. The NPs released almost entirely the encapsulated drug only following 1 h incubation at 43 °C, whereas they retained more than 95% of the payload in the physiological environment (37 °C), thus demonstrating their efficacy as on-demand drug delivery systems. The administration of drug-loaded NPs to ovarian cancer cells led to therapeutic effects outperforming the conventional administration of non-encapsulated paclitaxel, which highlights the potential of the zwitterionic UCST-type NPs as an innovative hyperthermia-responsive drug delivery system.

The Impact of 3D Nichoids and Matrix Stiffness on Primary Malignant Mesothelioma Cells.

Malignant mesothelioma is a type of cancer that affects the mesothelium. It is an aggressive and deadly form of cancer that is often caused by exposure to asbestos. At the molecular level, it is characterized by a low number of genetic mutations and high heterogeneity among patients. In this work, we analyzed the plasticity of gene expression of primary mesothelial cancer cells by comparing their properties on 2D versus 3D surfaces. First, we derived from primary human samples four independent primary cancer cells. Then, we used Nichoids, which are micro-engineered 3D substrates, as three-dimensional structures. Nichoids limit the dimension of adhering cells during expansion by counteracting cell migration between adjacent units of a substrate with their microarchitecture. Tumor cells grow effectively on Nichoids, where they show enhanced proliferation. We performed RNAseq analyses on all the samples and compared the gene expression pattern of Nichoid-grown tumor cells to that of cells grown in a 2D culture. The PCA analysis showed that 3D samples were more transcriptionally similar compared to the 2D ones. The 3D Nichoids induced a transcriptional remodeling that affected mainly genes involved in extracellular matrix assembly. Among these genes responsible for collagen formation, COL1A1 and COL5A1 exhibited elevated expression, suggesting changes in matrix stiffness. Overall, our data show that primary mesothelioma cells can be effectively expanded in Nichoids and that 3D growth affects the cells' tensegrity or the mechanical stability of their structure.

In vivo label-free tissue histology through a microstructured imaging window.

Tissue histopathology, based on hematoxylin and eosin (H&E) staining of thin tissue slices, is the gold standard for the evaluation of the immune reaction to the implant of a biomaterial. It is based on lengthy and costly procedures that do not allow longitudinal studies. The use of non-linear excitation microscopy in vivo, largely label-free, has the potential to overcome these limitations. With this purpose, we develop and validate an implantable microstructured device for the non-linear excitation microscopy assessment of the immune reaction to an implanted biomaterial label-free. The microstructured device, shaped as a matrix of regular 3D lattices, is obtained by two-photon laser polymerization. It is subsequently implanted in the chorioallantoic membrane (CAM) of embryonated chicken eggs for 7 days to act as an intrinsic 3D reference frame for cell counting and identification. The histological analysis based on H&E images of the tissue sections sampled around the implanted microstructures is compared to non-linear excitation and confocal images to build a cell atlas that correlates the histological observations to the label-free images. In this way, we can quantify the number of cells recruited in the tissue reconstituted in the microstructures and identify granulocytes on label-free images within and outside the microstructures. Collagen and microvessels are also identified by means of second-harmonic generation and autofluorescence imaging. The analysis indicates that the tissue reaction to implanted microstructures is like the one typical of CAM healing after injury, without a massive foreign body reaction. This opens the path to the use of similar microstructures coupled to a biomaterial, to image in vivo the regenerating interface between a tissue and a biomaterial with label-free non-linear excitation microscopy. This promises to be a transformative approach, alternative to conventional histopathology, for the bioengineering and the validation of biomaterials in in vivo longitudinal studies.