Model Organisms in Drug Discovery

54 C. ELEGANS FUNCTIONAL GENOMICS IN DRUG DISCOVERY
an enhancer/suppressor screen but, instead of using a mutant background, a
compound is employed to screen for mutants that are either resistant or
hypersensitive to the effect of the compound. The use of chemical ligands in
target identification and validation programs also allows for the concurrent
analysis of a target’s role in a disease process in parallel with an assessment of
the target’s druggability. In this reverse chemical genetics approach, small,
bioavailable and target-specific compounds are used to study biological
questions and to expand the pathways around validated drug targets. For
example, the acetylcholinesterase inhibitor aldicarb has been used extensively
in genetic screens to identify genes involved in synaptic vesicle exocytosis, such
as unc-18/nSec-1 (Hosono and Kamiya, 1991; Hosono et al., 1992). Mutations
in the gene unc-18 are resistant to the paralyzing effect of aldicarb.
Caenorhabditis elegans-based studies of this gene provided the first evidence
for the role of unc-18 in synaptic vesicle fusion (Gengyo-Ando et al., 1993;
Garcia et al., 1994). Aldicarb has been used to identify presynaptic genes. The
acetylcholine receptor agonist levamisol has been used to find postsynaptic
targets (Lewis et al., 1980a,b; Kim et al., 2001a).
Genetic screens also can be readily configured as mode-of-action (MOA)
assays to identify the molecular targets of drug candidates. This forward
chemical genetics approach is extremely useful for natural product molecules
or lead compounds arising from in vivo screens. Identification of the target
allows for the development of assays to enable lead optimization or the
identification of further chemical hits and leads. Yet hits and leads generated
from ‘on-target’ screens may, nevertheless, induce clinically relevant effects
through interactions with additional targets. Such effects require further MOA
analysis. An example is the antidepressant fluoxetine, which inhibits the
serotonin reuptake transporter and potentially interacts with other targets.
The effect of fluoxetine in C. elegans resembles the effect of an SSRI in that it
enhances drinking or a particular movement behavior called the ‘slow-down
response’. Further evidence for fluoxetine action on the C. elegans serotonin
reuptake transporter, MOD-5, is that mod-5 mutants are resistant to
fluoxetine (Ranganathan et al., 2001). Fluoxetine also induces a ‘nose
contraction’ phenotype in C. elegans, suggesting that fluoxetine acts on
additional targets. A genetic screen for fluoxetine-resistant mutants identified
two novel genes, nrf-6 and ndg-4, that define a novel gene family of multipass
transmembrane proteins (Choy and Thomas, 1999). The role of these genes in
serotonergic signaling and in depression is currently under investigation.
The suitability of the MOA studies described above depends largely on the
conservation of the binding site of the test compound. Although the
conservation of genes and pathways between humans and C. elegans is
remarkably high, a compound’s action often depends upon interaction of the
compound with only a few amino acid residues. A striking example of the
conservation of a compound’s binding site is given by the thapsigargin-