Abstract

There is a need for technologies which enable high-throughput
screening of 3D microtissue models. While immunostaining can  be effective for imaging microtissues in whole
mount preparations high-throughput analysis at this level is challenging.

To overcome this challenge we have developed a spheroid
tissue microarray (microTMA) technology which facilitates quantitative
histology and integrated genomics analysis of microtissues in 96 well format. Multiplex
immunofluorescent staining can be performed using existing automated staining
technology, and bespoke image analysis scripts applied to quantify toxicity and
efficacy end points.

Cellular organisation was visualised in a glioblastoma 3D
model containing normal neurons/astrocytes/oligodendrocytes in a "mini-brain"
model (gBS). The glioblastoma cells aggregate together to form a tumour-like
structures within the microtissues. Anti-cancer drug responses affecting cell
proliferation and apoptosis were measured in a cell-type specific fashion.

To test
whether liver 3D microtissues (LiMTs) 
recapitulate CAR- mediated procarcinogenic key events in response to
phenobarbital (PB) we performed hepatocyte proliferation (LI%) analysis in rat
and human LiMTs integrated with transcriptomics analysis.Rat and
human LiMTs were treated with PB (500 uM - 2000 uM) for 24 hr - 96 hr. There
was a dose-dependent induction of LI% in rat LiMTs, however there was little or
no effect of PB on LI% in human LiMTs. There was a dose- and
time-dependent PB-mediated induction of CAR regulated genes in human and rat
LiMTs.

Human
and rat LiMTs gave expected responses in PB-induced hepatocyte proliferation
and enzyme induction with rat LiMTs showing significant dose-dependent effects
while human LiMTs showed no hepatocyte proliferation.

In conclusion our novel microTMA technology facilitates understanding
of the cellular organisation and quantification of molecular mechanisms of drug
action that can support drug development/risk assessment.