Model Organisms in Drug Discovery

DROSOPHILA AS A MODEL ORGANISM FOR BIOMEDICAL SCIENCE 97
organism-based screen the delivery of compounds will not be specific to a
particular tissue or cell type, nor would it be uniform for all cells. Thus, the
effect of a specific chemical inhibitor is expected to mimic that of a LOF
mutation but with varied effectiveness in different tissue and cell types due to
local concentrations of the compound. By controlling compound dosage, one
can adjust the degree of functional loss in the target – a situation analogous to
the creation of an allelic series of mutations in a gene. Another unique feature
is that hit compounds are selected not only by their potency against the targets
but also by pharmacological properties such as absorption, membrane
permeability and cellular/organism stability. Thus, the hit rate might be low
but the value of hits is high.
Technical hurdles for carrying out compound screens in Drosophila
There are a number of technical challenges in using organism-based assays for
screening large chemical diversity. First, even though we have achieved in-house
success in sorting and dispensing embryos and larvae into 96-well plates using
instruments available in the market (Li et al., 2001), growing flies in a 96-well
format has not yet been optimized. In particular, the culture medium needs to
be modified to be compatible for automation, compound addition, larval
growth and adult viability. Second, machine-readable assay phenotypes need to
be developed. Traditional morphological phenotypes such as bristle number,
roughness of eyes or behavior are difficult to adapt to a high-throughput
compound screen. For example, the classical readout for circadian rhythm is the
rhythmic behavior of local motor activity, which is not very easy to scale up. By
using the rhythmic activity of the promoter of the period gene to drive the
expression of the firefly luciferase, the throughput of the circadian rhythm
readout is dramatically increased (Plautz et al., 1997). Third, most compound
libraries formated for in vitro screens and cell-based screens may not be useful
for compound screens in Drosophila. This results from the fly’s tolerance limit
for dimethylsulfoxide (DMSO), combined with a requirement for higher
compound concentrations in fly growth medium relative to cell culture media.
For example, 40 mM rapamycin and 2 mM cycloheximide have been used to
delay Drosophila larval development (Britton and Edgar, 1998; Oldham et al.,
2000) and 40 mM G418 is typically used for selecting flies expressing a G418
resistance gene (Xu and Rubin, 1993). If a screen is to be done at 100 mMin1%
DMSO, a 10 mM library in 100% DMSO would be needed to keep the DMSO
concentration at or below 1%, above which there is toxicity. As a comparison,
the compound library provided by the National Cancer Institute contains
compounds at a concentration of 1 mM in 100% DMSO.
Using sensitized assays may help to reduce the demand on compound
concentration. For example, the effective rapamycin concentration for