# Hypoxia-induced drug-resistance bias 3D cancer spheroid drug screens

## TL;DR

This study shows that hypoxia in 3D cancer spheroids causes drug resistance, and offers a framework to select effective drugs for all cell types in spheroids.

## Contribution

The study identifies hypoxia-induced quiescence as a key barrier to drug efficacy in 3D spheroids and proposes a method to overcome it.

## Key findings

- Oxygen decay in spheroids causes hypoxia beyond 20 μm from the surface.
- Spheroids larger than 150 μm cannot achieve IC50 for drugs targeting proliferating cells.
- Targeting viability barriers enables effective drug selection for all cell populations in spheroids.

## Abstract

Cellular 3D cancer spheroid technologies are novel tools that facilitate large-scale drug screening to bridge the in vitro–in vivo gap, without the cross-species effects of animal models. However, many spheroid studies fail to achieve
IC50 (dosage for 50% inhibition) even for unreasonably high applied drug concentrations (up to 1000× 2D
IC50). By mapping oxygen transport in patient-derived pancreatic cancer spheroids, this limiting viability is attributed to a near-universal oxygen decay gradient that renders cells deeper than 20 μm from the spheroid surface hypoxically quiescent and resistant to many chemotherapeutic drugs. The dose-independent viability barrier prevents
IC50 from being achieved for spheroids larger than 150 μm in diameter if the applied drug is dependent on the proliferating cell behavior. By examining three cancer cell types and five chemotherapeutic drugs, targeting this limiting viability barrier allows the selection of drugs and adjuvants that are effective in treating all cell populations within a spheroid. The reported analysis provides a framework for the accurate assessment of drug efficacy to target both well-oxygenated proliferating cells and hypoxically quiescent cells in biologically relevant and realistic 3D spheroid systems.