| dc.description.abstract |
Numerous studies have demonstrated that membrane transporters are critical
determinants for drug disposition, drug absorption, therapeutic efficacy, and
adverse drug reactions. In the human genome, more than 400 membrane
transporters have been annotated and been divided into two superfamilies: ATPbinding cassette (ABC) family and solute carrier (SLC) family. Many ABC and
SLC transporters not only contribute to the influx of metabolites and drugs but are
also involved in their efflux. In vitro and in vivo, various animal models including
mice, rats, rabbits, monkeys, and different cell lines were used to study the
relationship between clinical pharmacokinetic drug-drug interaction (DDI) and
human transporters. However, these studies are usually expensive and timeconsuming. Therefore, we planned to use the fruit fly Drosophila melanogaster
as a rapid testing and low-cost animal model to study the function of human
membrane transporters.
At first, I sought to study the function of the organic anion transporters hOATP1B1
(encoded by SLCO1B1), hOATP2B1 (SLCO2B1) and hOATP1B3 (SLCO1B3),
which are clinically relevant uptake transporters in the sinusoidal membrane of
human hepatocytes. For this purpose, I generated flies expressing these human
uptake transporters in the salivary glands. They did not localize to the target
tissue, but were instead found in excretory cells suggesting that they were
eliminated because of potential cell toxic properties. Arguing that undetectable
levels of hOATP1B1, hOATP2B1 and hOATP1B3 may nevertheless be localized
correctly, I performed tracer transport assays by confocal microscopy. My results
suggested that fluorescent substrates of hOATP1B1, hOATP2B1 and hOATP1B3
(Mercury dibromofluorescein disodium salt, MDBF; 4,5- Dibromofluorescein, DBF;
Cholyl-Lysyl-Fluorescein, CLF) were transported to the lumen of the salivary
glands of embryos independently of these human transporters. Further results
failed to demonstrate that fly organic anion-transporting polypeptides (Oatps)
may assist fluorescent substrate transfer across the cell membrane into the
lumen of the salivary glands. Rather, my data suggested that dyes were
transported by transcytosis to the lumen of this organ. Thus, contrary to our5
expectation, the fruit fly is not a suitable model for pharmacological studies of
OATPs.
In a second project, I established a test system for DDI involving hOCT1, a further
clinically relevant uptake transporter in human hepatocytes. hOCT1 was
expressed in various organs and tissues in embryos produced by females that
were fed with the cytotoxin and hOCT1 substrate cisplatin. My results suggested
that cisplatin was transported into embryos by hOCT1 and was embryonic lethal.
Interestingly, cimetidine, another substrate of hOCT1, reduced the toxicity of
cisplatin in embryos that expressed hOCT1 in the nervous system. Thus, this
setup allows studying successfully DDI in D. melanogaster and may ultimately
serve to identify and evaluate new hOCT1 interacting molecules in
pharmaceutical experiments. |
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