Background: While Thioredoxin Reductase (TrxR) plays an important role in regulation of the
intracellular redox balance and various signalling pathways, Glutathione S-Transferase (GSTs) enzymes belong to
the detoxification family that catalyse the conjugation of glutathione with various endogenous and xenobiotic
electrophiles. Since TrxR and GSTs are overexpressed in many cancer cells, they have been identified as potential
targets to develop chemotherapeutic strategies.
Method: The mitochondrial TrxR (TrxR2) enzyme and the cytosolic GST enzyme was purified from rat liver via
affinity chromatography. After the purification, the in vitro inhibition effects of some anticancer drugs (cisplatin,
calcium folinate, carboplatin, epirubicin hydrochloride, doxorubicin hydrochloride, paclitaxel, etoposide,
fluorouracil, and methotrexate) were investigated on both enzymes. Since only methotrexate inhibits both enzymes
among all the anticancer drugs, a molecular docking study was performed to determine the binding site and the
binding affinity of methotrexate to the enzymes.
Results: Firstly, TrxR2 and GST were found to have a specific activity of 0.436, 1765 EU/mg proteins with a yield
of 39.20%, 31.28% and 207.6, 3516.6 of purification fold, respectively. While TrxR2 was strongly inhibited by all of
the anticancer drugs, GST was not inhibited by any of the anticancer drugs except methotrexate.
Conclusion: Both enzymes were inhibited by only methotrexate in rat liver, and methotrexate was well placed in the
active sites of both proteins. Therefore, it may be argued that methotrexate may be a more effective anticancer drug
than all other drugs used in this study against the multi drug resistance that will occur during chemotherapy.