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Potent synthetic drugs, as well as biomolecules extracted from plants, have been investigated for their selectivity toward cancer cells. The main limitation in cancer treatment is the ability to bring such molecules within each single cancer cell, which requires accumulation in the peritumoral region followed by homogeneous spreading within the entire tissue. In the last decades, nanotechnology has emerged as a powerful tool due to its ability to protect the drug during blood circulation and allow enhanced accumulation around the leaky regions of the tumor vasculature. However, the ideal size for accumulation of around 100 nm is too large for effective penetration into the dense collagen matrix. Therefore, we propose a multistage system based on graphene oxide nanosheet-based quantum dots (GOQDs) with dimensions that are 12 nm, functionalized with hyaluronic acid (GOQDs-HA), and deposited using the layer-by-layer technique onto an oil-in-water nanoemulsion (O/W NE) template that is around 100 nm in size, previously stabilized by a biodegradable polymer, chitosan. The choice of a biodegradable core for the nanocarrier is to degrade once inside the tumor, thus promoting the release of smaller compounds, GOQDs-HA, carrying the adsorbed anticancer compound, which in this work is represented by curcumin as a model bioactive anticancer molecule. Additionally, modification with HA aims to promote active targeting of stromal and cancer cells. Cell uptake experiments and preliminary penetration experiments in three-dimensional microtissues were performed to assess the proposed multistage nanocarrier.

Current 3D cancer models (in vitro) fail to reproduce complex cancer cell extracellular matrices (ECMs) and the interrelationships occurring (in vivo) in the tumor microenvironment (TME). Herein, we propose 3D in vitro colorectal cancer microtissues (3D CRC μTs), which reproduce the TME more faithfully in vitro. Normal human fibroblasts were seeded onto porous biodegradable gelatin microbeads (GPMs) and were continuously induced to synthesize and assemble their own ECMs (3D Stroma μTs) in a spinner flask bioreactor. Then, human colon cancer cells were dynamically seeded onto the 3D Stroma μTs to achieve the 3D CRC μTs. Morphological characterization of the 3D CRC μTs was performed to assess the presence of different complex macromolecular components that feature in vivo in the ECM. The results showed the 3D CRC μTs recapitulated the TME in terms of ECM remodeling, cell growth, and the activation of normal fibroblasts toward an activated phenotype. Then, the microtissues were assessed as a drug screening platform by evaluating the effect of 5-Fluorouracil (5-FU), curcumin-loaded nanoemulsions (CT-NE-Curc), and the combination of the two. When taken together, the results showed that our microtissues are promising in that they can help clarify complex cancer-ECM interactions and evaluate the efficacy of therapies. Moreover, they may be combined with tissue-on-chip technologies aimed at addressing further studies in cancer progression and drug discovery.

Cystic fibrosis is characterized by lung dysfunction involving mucus hypersecretion, bacterial infections, and inflammatory response. Inflammation triggers pro-fibrotic signals that compromise lung structure and function. At present, several in vitro cystic fibrosis models have been developed to study epithelial dysfunction but none of these focuses on stromal alterations. Here we show a new cystic fibrosis 3D stromal lung model made up of primary fibroblasts embedded in their own extracellular matrix and investigate its morphological and transcriptomic features. Cystic fibrosis fibroblasts showed a high proliferation rate and produced an abundant and chaotic matrix with increased protein content and elastic modulus. More interesting, they had enhanced pro-fibrotic markers and genes involved in epithelial function and inflammatory response. In conclusion, our study reveals that cystic fibrosis fibroblasts maintain in vitro an activated pro-fibrotic state. This abnormality may play in vivo a role in the modulation of epithelial and inflammatory cell behavior and lung remodeling. We argue that the proposed bioengineered model may provide new insights on epithelial/stromal/inflammatory cells crosstalk in cystic fibrosis, paving the way for novel therapeutic strategies.

Despite the well-known role of chronic human papillomavirus (HPV) infections in causing tumors (i.e., all cervical cancers and other human malignancies from the mucosal squamous epithelia, including anogenital and oropharyngeal cavity), its persistence is not sufficient for cancer development. Other co-factors contribute to the carcinogenesis process. Recently, the critical role of the underlying stroma during the HPV life cycle and HPV-induced disease have been investigated. The tumor stroma is a key component of the tumor microenvironment (TME), which is a specialized entity. The TME is dynamic, interactive, and constantly changing-able to trigger, support, and drive tumor initiation, progression, and metastasis. In previous years, in vitro organotypic raft cultures and in vivo genetically engineered mouse models have provided researchers with important information on the interactions between HPVs and the epithelium. Further development for an in-depth understanding of the interaction between HPV-infected tissue and the surrounding microenvironment is strongly required. In this review, we critically describe the HPV-related cancers modeled in vitro from the simplified 'raft culture' to complex three-dimensional (3D) organotypic models, focusing on HPV-associated cervical cancer disease platforms. In addition, we review the latest knowledge in the field of in vitro culture systems of HPV-associated malignancies of other mucosal squamous epithelia (anogenital and oropharynx), as well as rare cutaneous non-melanoma associated cancer.