Horizon eBook - 14

Innovations in Cell-Based Screening * More Predictive Cell-Based Assays

spatial oxygen gradients like those near tumor
blood vessels. Employing this device to mimic in
vivo oxygen profiles in vitro could facilitate the
development of a screening methodology that better
predicts how a given drug will act in the body."
Identifying JAK Inhibitors
Atli Thorarensen, Ph.D., medicinal chemistry project
lead at Pfizer, is developing JAK signaling pathway
inhibitors. He chose his assay to employ primary
cells for HTS and overcome limitations associated
with cell-line model systems.
"I think the industry needs to move away from
utilizing cell lines as surrogates of pharmacology,"
asserts Dr. Thorarensen. "Our learnings have taught
us that primary cells are much more valuable as predictors of pharmacology then stable expressed cells,
which have forced target expression and may have
lost many pathway signaling components."
Historically, results with JAK inhibitors show
a disconcerting disconnect between enzyme and
cellular function. For example, in the cell-line model
Dr. Thorarensen used there was a "very poor

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correlation between JAK3 enzyme inhibition with
ATP competitive inhibitors and the observed cellular
response (measurement of STAT phosphorylation).
This indicated that there are artifacts in the cell lines
not allowing us to accurately measure inhibition of
JAK3 function.
"The cell assay we then developed is a measurement of STAT phosphorylation in primary cells,
[which were] PBMCs expanded from a donor. The
results showed we could obtain a cellular signal that
accurately represented inhibition of JAK3 enzyme
activity with ATP competitive inhibitors."
The sensitivity of the primary-cell-based model
system is enabling Dr. Thorarensen to move to the
next step and screen for allosteric inhibitors that
bind outside of the conserved ATP-binding sites of
the JAK enzymes. According to Dr. Thorarensen,
the system "should allow for the identification of
inhibitors that selectively inhibit specific family
members-JAK1, JAK2, JAK3, or TYK2."
Integrating Live Imaging
Imagen Biotech uses high-content screening of

patient samples to add a personal touch to cancer
drug discovery.
"We choose the Thermo Scientific ArrayScan, a
high-content fluorescence-based screening machine,
for our project," explains CSO Garath Griffiths,
Ph.D. "It automatically captures images and analyzes
them with different algorithms; you can teach the
machine to extract the information that you want
from the assay."
The drug testing platform employs live cancer
cells with live imaging, and it uses the fluorescence
of cell death as a readout assay for screening
compounds. The red fluorescent is a cell-permeable
probe, a death marker, that enters dying but not
healthy cells. Fresh ovarian cancer cells or stem
cell lines generated from glioblastoma cells, taken
directly from patient brain tumors, were used for
the assays.
Automation enables rapid analysis of a huge
amount of cell data from dose responses from a
large number of drugs. "For every hundred hours
of work it takes to pull in the samples to analyze,
with the software algorithms, [it is possible to]


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