Biomedical Engineering Reference
In-Depth Information
pose of obtaining information about the antibiotic's mechanism of action, poten-
tial toxicity, and possible mechanisms of bacterial resistance. An application is
described by Evers and Gray (9), who identified the novel antimicrobial com-
pound Ro-64-1874. The effects of this compound on protein expression in the
bacterium
Haemophilus influenzae
was compared to the effects of a series of
well-characterized other antibiotics that affect cellular processes such as tran-
scription, translation, or tRNA synthesis. It was found that Ro-64-1874 causes a
protein expression profile (obtained through 2D electrophoresis) very distinct
from most characterized antibiotics but similar to that of the antibiotic
trimethoprim, which is an inhibitor of the enzyme dihydrofolate reductase
(DHFR). This assay thus suggests that Ro-64-1874 may also inhibit DHFR ac-
tivity.
4.2. Drug Target Validation
The ideal inhibitory drug is highly specific, i.e., it inhibits a single protein
or other gene product so completely as if the encoding gene itself was absent.
Marton and collaborators (21) developed a test for drug specificity that is based
on this observation. For their proof-of-principle experiments, they chose the
drug target calcineurin in the budding yeast
Saccharomyces cerevisiae
. Cal-
cineurin is a protein phosphatase implicated in T-cell activation, apoptosis, and
cardiac hypertrophy in higher organisms. In yeast, it is involved in the regulation
of the cell cycle and in osmotic homeostasis. Marton and collaborators tested
two drugs—FK506 and Cyclosporin A—both immunosupressants that can in-
hibit calcineurin activity in higher organisms and in yeast. How specific is the
action of these drugs? They asked this question by comparing gene expression
profiles between yeast mutants from which the calcineurin gene had been de-
leted, and strains where calcineurin had been pharmacologically inactivated by
application of either drug. If the drugs act with high specificity, one would ex-
pect that both the genetic and pharmacological inactivation yield very similar
effects on gene expression. This is what the authors observed. In contrast, dele-
tions in genes unrelated to the calcineurin pathway yielded effects on gene ex-
pression uncorrelated with those of pharmacological inhibition. This approach is
also suited to identify the exact nature of a drug's off-target effects, by studying
how gene expression changes in pharmacologically versus genetically manipu-
lated organisms (21).
4.3. Toxicology
Cyclosporin A is an immunosuppressant with a number of side effects, most
importantly among them nephrotoxicity. What causes this side effect? Evers and
