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Three-dimensional (3D) in vitro models of tumors are gaining interest as versatile platforms for treatment screening. In this context, heterocellular cultures in which various cell types are co-cultured are being explored to investigate whether partner cells can influence the treatment efficacies. However, when the cells are co-cultured, it is challenging to distinguish them and it becomes impossible to identify whether the treatment affects each cell line in a similar way or if there is a certain selectivity. Here, we propose a protocol in which different cell types are pre-labeled with fluorescent reporters prior to 3D culture initiation. Subsequently, the internal architecture of the 3D cancer models can be longitudinally monitored for model characterization, and to potentially detect architectural and treatment selectivity in response to therapy. This protocol employs quantum dots as non-photobleaching dyes and two-photon excited microscopy as a widely accessible imaging modality. In combination with an appropriate image analysis workflow, this protocol will help to investigate the architectural development of heterotypic microtumor/spheroid/organoid models and possibly identify treatment efficacies on individual cell populations represented within the models., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

Ultra-small gold nanoclusters (AuNCs) are increasingly investigated for cancer imaging and radiotherapy enhancement. While fine-tuning the AuNC surface chemistry can optimize their pharmacokinetics, its effects on radiotherapy enhancement remain largely unexplored. This study demonstrates that optimizing the surface chemistry of AuNCs for increased tumor uptake can significantly affect its potential to augment radiotherapy outcomes.

Bioengineered three-dimensional (3D) tumor models that incorporate heterotypic cellular communication are gaining interest as they can recapitulate key features regarding the intrinsic heterogeneity of cancer tissues. However, the architectural complexity and heterogeneous contents associated with these models pose a challenge for toxicological assays to accurately report treatment outcomes. To address this issue, we describe a comprehensive image analysis procedure for structurally complex organotypic cultures (CALYPSO) applied to fluorescence-based assays to extract multiparametric readouts of treatment effects for heterotypic tumor cultures that enables advanced analyses. The capacity of this approach is exemplified on various 3D models including adherent/suspension, mono-/heterocellular cultures and several disease types. The subsequent analysis revealed specific morphological effects of oxaliplatin chemotherapy, radiotherapy, and photodynamic therapy. The procedure can be readily implemented in most laboratories to facilitate high-throughput toxicological screening of pharmaceutical agents and treatment regimens on organotypic cultures of human disease to expedite drug and therapy development.

As clinical nanomedicine has emerged over the past two decades, phototherapeutic advancements using nanotechnology have also evolved and impacted disease management. Because of unique features attributable to the light activation process of molecules, photonanomedicine (PNM) holds significant promise as a personalized, image-guided therapeutic approach for cancer and non-cancer pathologies. The convergence of advanced photochemical therapies such as photodynamic therapy (PDT) and imaging modalities with sophisticated nanotechnologies is enabling the ongoing evolution of fundamental PNM formulations, such as Visudyne®, into progressive forward-looking platforms that integrate theranostics (therapeutics and diagnostics), molecular selectivity, the spatiotemporally controlled release of synergistic therapeutics, along with regulated, sustained drug dosing. Considering that the envisioned goal of these integrated platforms is proving to be realistic, this review will discuss how PNM has evolved over the years as a preclinical and clinical amalgamation of nanotechnology with PDT. The encouraging investigations that emphasize the potent synergy between photochemistry and nanotherapeutics, in addition to the growing realization of the value of these multi-faceted theranostic nanoplatforms, will assist in driving PNM formulations into mainstream oncological clinical practice as a necessary tool in the medical armamentarium.