ABCB1 limits the cytotoxic activity of TAK-243, an inhibitor of the ubiquitin-activating enzyme UBA1

Abstract

Background: One of the major concerns of cancer therapy is the emergence of multidrug resistance (MDR). The MDR-associated ATPbinding cassette sub-family B member 1 (ABCB1) transporter is established to mediate resistance against numerous anticancer drugs. In this study, we demonstrated that the Ubiquitin-like modifier activating enzyme 1 (UBA1) inhibitor TAK-243 is transported by the ABCB1. Methods: MTT assay was performed to evaluate the cytotoxicity of TAK-243. Western blot was carried out to investigate if TAK-243 affect to ABCB1 protein expression in cancer cells. High Performance Liquid Chromatography (HPLC) and ATPase assay were carried out to confirm TAK-243 as an ABCB1 substrate. [ 3 H]-paclitaxel accumulation assay was used to determine the MDR reversal effect of TAK-243. Computational docking analysis was performed to investigate the drug-transporter binding position. Results: The cytotoxicity profile showed that TAK-243 was less effective in ABCB1-overexpressing cells than in the parental cells, but pharmacological inhibition or knockout the gene of ABCB1 was able to reverse TAK-243 resistance. Furthermore, TAK-243 potently stimulated ABCB1 ATPase activity and the HPLC analysis revealed that TAK-243 accumulation was significantly reduced in ABCB1-overexpressing cells. Finally, the computational docking analysis indicates a high binding affinity between TAK-243 and human ABCB1 transporter. Conclusions: Our in vitro data characterized TAK-243 as a substrate of ABCB1, which may predict limited anticancer effect of this compound in drug resistant tumors.

Activity of the Ubiquitin-activating Enzyme Inhibitor TAK-243 in Adrenocortical Carcinoma Cell Lines, Patient-derived Organoids, and Murine Xenografts

“…Because TAK-243 has been reported to be a substrate for the drug efflux P-glycoprotein pumps ( 29 ), we tested the expression of the ATP-binding cassette transporters MDR-1 ( ABCB1 ) and BCRP ( ABCG2 ). Consistent with the report that genetically engineered ABCB1-overexpressing colon SW620, kidney HEK, and mouth epithelial KB cells are highly resistant to TAK-243, we found that the NCI-H295R and CU-ACC1 cell line overexpress MDR1 and are the most resistant to TAK-243 ( Fig.…”

“…Expression of MDR1 and SLFN11 are potential predictors of TAK-243 response. Increased expression of the ABC transporter MDR1 has been associated with decreased TAK-243 sensitivity ( 29 ). Accordingly, we found that the ACC cell lines, NCI-H295R and CU-ACC1, which express ABCB1 , the gene encoding MDR1/PgP, are less sensitive to TAK-243 than the CU-ACC2 cells ( Fig.…”

Progress in the studies on the molecular mechanisms associated with multidrug resistance in cancers

“…Till now, many ABC members were found to be overexpressed in the cancer cells, responsible for transporting certain anti-tumor drug(s) out from the cells, voiding overaccumulation of intracellular drug or blockage of the pathways necessary for cell survival 13 , 14 . The two most famous ABCB1/P-glycoprotein (P-gp) and ABCG2/Breast cancer resistance protein (BCRP) transporters, which were confirmed to have transmembrane domain(s) [TMD(s)] and cytoplasmic nucleotide-binding domain(s) [NBD(s)] 15 , 16 , 17 , exhibited strong ability to keep pumping various substrate anti-tumor drugs (including inhibitors) out, such as KRAS-G12C inhibitor ARS-1620, Aurora kinase inhibitor GSK1070916, paclitaxel, doxorubicin, and TAK-243, an inhibitor of the ubiquitin-activating enzyme UBA1 for ABCB1 and mitoxantrone, ubiquitin activating enzyme (UAE) inhibitor MLN7243 for ABCG2 4 , 11 , 18 , 19 , 20 , 21 , 22 . ABCC1/MRP1, which mediates MDR in several types of cancer, including lung, childhood neuroblastoma, ovary, prostate, and breast cancers, can transport a variety of substrates including drugs, toxic molecules and important physiological substrates like lipid peroxidation products, for example, S -(6-(7- 11 C-methylpurinyl))glutathione [ 11 C-MPG], lipid peroxidation products, GSSG, and reduced glutathione (GSH) 23 , 24 , 25 , 26 .…”

Utilization of Photoaffinity Labeling to Investigate Binding of Microtubule Stabilizing Agents to P-Glycoprotein and β-Tubulin

“…The binding could compete with unlabeled taxol. [ 3 H]­Taxol continues to be employed in studies of taxane-site ligand interactions …”