(1227-D) Automated Screening Workflows with Animal-free Nanofibrillar Cellulose 3D hydrogels

Abstract: In precision medicine, it is essential to develop biologically relevant and physiologically tissue mimicking, but cost-effective and experiment-reproducible cell models. 3D spheroid/organoid culture methods with hydrogels have become a powerful tool for examination of better in vivo relevancy compared to well-established 2D monolayer models. However, due to the complexity of 3D models, usually these are challenging to scale-up for automated high-throughput workflows. The selection of suitable hydrogel with ease-of-use and repeatability is essential towards developing automation friendly 3D models.

GrowDex hydrogels are made from plant-based nanofibrillar cellulose and ultrapure water and have shown to provide biocompatible ECM-like support matrix for development of clinically relevant healthy and cancerous cell models for drug discovery and development. The hydrogels are well-defined and animal-free, shear thinning and temperature stable which makes them ideal for various pipetting robots and dispensers for automated and scalable 3D models, with a possibility of direct biochemical and image-based cell assays.

Recently, Feodoroff et al. (2023) established a robust 3D drug sensitivity and resistance testing (3D-DSRT) cell model pipeline with GrowDex in a HT mode. First, they successfully developed the workflow with a hepatocyte cell line (HepG2) with two methods: pre-culturing and pre-drugging method with 35 compounds in 5 different concentrations. In both, lab automation included: dispensing of GrowDex hydrogel with BioMek FX liquid handler, addition of compounds with Echo 550 Acoustic Dispenser directly on the plate and at the end of culture, CellTiter-Glo® 2.0 was directly added with Multidrop and cell viability was read with the PHERAstar FS plate reader directly from the samples, independent of the method. After establishing the pipeline, they were able to translate the workflow to ovarian cancer PDCs of two patients for functional precision medicine study with 52 oncological compounds in 5 clinical concentrations in 96 and 384 well formats.

Ideally, any 3D culture modality and method that are being developed for screening, translatability to other cell types and automatable workflows are the desired factors for scalable and reproducible assays. Stirnimann & Booij (2022) were able to increase their screening throughput from 96 to 1536 well format, by easy automated dispensing of GrowDex hydrogels with CertusFlex contactless dispenser and were able to see high batch-to-batch reproducibility of their assays. Additionally, to maximise readouts retrieved from each cell or well, we have developed a multiplexing assay inhouse, in collaboration with Promega, specifically for liver toxicity assays. This includes well-based readouts of cell viability (CellTiterGlo2.0), cell death (CellTox Green), liver cell transcription, translation, processing and export functionality by measuring albumin secretion in the media, and finally liver enzymatic capacity and induction (P450-Glo). GrowDex can be used to develop animal-free, scalable, automated workflows for their screening purposes, with maximized readouts, further increased screening value and output.