Assessing chemotherapeutic effectiveness using a paper-based tumor model

Abstract

In vitro models for screening new cancer chemotherapeutics often rely on two-dimensional cultures to predict therapeutic potential. Unfortunately, the predictive power of these models is limited, as they fail to recapitulate the complex three-dimensional environments in tumors that promote the development of a chemoresistant phenotype. In this study, we describe the preparation and characterization of paper-based cultures (PBC) engineered to assess chemotherapeutic effectiveness in three dimensional, diffusion-limited environments. Similar environments are found within poorly vascularized tumors. These cultures, which are assembled by stacking together cell-laden paper scaffolds to yield thick tissue-like structures, generate monotonic gradients vertically through the culture and provide distinct chemical environments for each scaffold. After prolonged incubation, the scaffolds can simply be peeled apart and analyzed to relate cellular responses to the chemical environment experienced at that scaffold. Through fluorescence imaging, viable and proliferative cell populations were mapped within a stacked culture and found to be most abundant in scaffolds close to the nutrient-rich medium. By adjusting the cell density, we modulated the spatiotemporal evolution of oxygen gradients across the cultures and correlated these environmental changes with cellular sensitivity to SN-38 exposure. From these results, we showed that differences in the oxygen gradients produced cellular populations with significantly different chemosensitivities. Through this work, we highlight PBCs ability to serve as an analytical model capable of determining chemotherapeutic effectiveness under a range of chemical environments.

The Current Landscape of 3D In Vitro Tumor Models: What Cancer Hallmarks Are Accessible for Drug Discovery?

“…A number of paper‐supported multilayer culture platforms have emerged over the last 10 years . These involve encapsulating cells within a biopolymer, typically a natural hydrogel such as collagen or Matrigel, infiltrated into a thin porous paper scaffold.…”

“…As a follow‐up, a 96 well plate format was developed by first patterning wax circles into the paper sheet to generate “wells” that defined the wicking boundaries of the cell suspensions in the paper which is necessary for controlled deposition of the cells (Figure B). Since then, a number of variations of paper‐based tumor models have been published (Figure D–F) …”

“…Chemotherapeutic response was assessed and related to the oxygen tension throughout the stack. Reproduced (Adapted) with permission . Copyright 2017, Royal Society of Chemistry.…”

“…Major limitations of these in vitro approaches include the need to perform rapid histological sectioning to prevent redistribution of drug gradients and to enable accurate examination of drug diffusion profiles, and the use of fluorescently or radiolabeled drugs for detection and quantification within the tissue slices; however, HPLC and mass spectrometry can be used to assess drug diffusion kinetics through MCCs by assessing changes in reservoir medium concentration on either side of the tissue membrane. More recently, we and others have proposed the use of stacked paper models for drug penetration analysis to address many of these challenges. Throughput, however, may limit the translatability of these approaches for understanding drug penetration in the context of preclinical drug discovery.…”

Tracking the invasion of breast cancer cells in paper-based 3D cultures by OCT motility analysis

“…To quantify cellular movement in these assays, we relied on confocal fluorescence imaging and microscopy [12,16,17]. The opaque and highly scattering nature of the cellulose fibers in the paper scaffolds limits the imaging depth, requiring the culture to be disassembled prior to analysis.…”