Personalized Medicine Using Patient-Derived Explants
Above: Pancreatic Ductal AdenoCarcinoma (PDAC) microtissue cultured in perfusion for 14 days.
Microtissues Preserve Sample Complexity
The history of cell culture is resplendent in efforts to extend the long-term culture of primary cells and tissues. Many existing models and techniques for tissue culture (e.g., organoids and patient-derived explants) aim to recapitulate an aspect of in vivo biology to a level of detail not possible with cells grown in a monolayer. The high rate of failure in phase 1 clinical trials is often attributed to ambiguous drug response in cell lines cultured in a 2D liquid-cover monolayer, a lack of autologous immune cells in models, and the inaccurate representation of inter-individual heterogeneity in over-simplified in vitro models. Relevant in vivo conditions have been more appropriately recapitulated in 3D cell models such as cell-cell interaction, migration, differentiation, and drug sensitivity. Patient-derived explants, or microtissues, are mechanically disaggregated biopsies on the order of 100 micrometers in size that are cultured without enzymatic digestion preparation to better retain native heterogeneity and complexity. In other words, cellular diversity, architecture, fibrotic stroma, resident immune cells, and extracellular matrix proteins are retained by via minimal processing to the tissue before culture.
Left: Pancreatic Ductal AdenoCarcinoma (PDAC) microtissue cultured in perfusion for 14 days.
Above: Pancreatic Adenocarcinoma (PDAC) cells co-cultured with Cancer-Associated Fibroblasts (CAFs) in a BME droplet in perfusion for 21 days.
The Limitations of Basement Membrane Extract (BME) and Dome Culture
Unfortunately, the culture of explants or complex organoids is inherently difficult using most traditional culture systems. One of the most common methodologies for 3D culture revolves around the use of basement membrane extracts, or commercially, Matrigel. By encapsulating organoids or explants in domes of highly nutrient and growth factor rich gel, these cultures can be kept relatively healthy for extended durations. However, due to limitations of diffusive transport, the size and number of samples in each dome is often limited. Furthermore, growth may be slow due to a need for cells to digest the gel around them in order to have space. Moreover, unless paying a steep price for chemically defined gels, the results from studies using this method are prone to unpredictability due to the unknown concentrations of certain factors and batch to batch variability which is common for Matrigel and other basement membrane extract gels.
Right: Pancreatic Adenocarcinoma (PDAC) cells co-cultured with Cancer-Associated Fibroblasts (CAFs) in a BME droplet in perfusion for 21 days.
Above: Human pancreatic slice cultured in perfusion for 10 days. Functional Islets of Langerhans, the endocrine structures of the pancreas responsible for the metabolism of glucose, are shown in white (Zinquin).
Preserving Microtissue Viability Using Perfusion
Aurita has developed a 3D culture platform for ex-vivo samples, as well as organoids, spheroids, and tumoroids, which supports their viability while providing a conducive environment for the associated motile cells, like immune cells, to function as normal. The interstitial spaces between LLS™ particles are tuned to emulate capillary beds in both their physical dimensions and functional surfaces as well as their action as fluid channels for controlled media perfusion in Darcy Plates™ bringing in nutrients and removing waste. By placing microtissues into 3D culture in LLS™, they have been put into an in vitro capillary bed surrogate. This platform has enabled the successful culture of a diverse array of microtissues and organoids including both normal and cancerous, human and otherwise. With minimal processing and no enzyme digestion, preparation of samples is fast and can be done immediately following a surgery or biopsy collection and quickly put into culture in order to avoid as much degradation of viability as possible. As the field of personalized medicine grows, functional microtissue models will play a critical role in clinical research.
Left: Human pancreatic slice cultured in perfusion for 10 days. Functional Islets of Langerhans, the endocrine structures of the pancreas responsible for the metabolism of glucose, are shown in white (Zinquin).
