The use of a novel taxane-based P-glycoprotein inhibitor to identify mutations that alter the interaction of the protein with paclitaxel
Autor: | Donald J. Gruol, Alba E. Phippard, Iwao Ojima, Ralph J. Bernacki, Josh Bernd |
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Rok vydání: | 2001 |
Předmět: |
ATP Binding Cassette Transporter
Subfamily B Paclitaxel Drug resistance Pharmacology Biology medicine.disease_cause chemistry.chemical_compound Mice medicine Tumor Cells Cultured Animals Humans P-glycoprotein Inhibitor Mutation Mice Inbred BALB C Taxane Membrane Proteins Molecular biology Antineoplastic Agents Phytogenic Transmembrane domain chemistry Puromycin Cytoplasm Molecular Medicine Female Taxoids Cell Division |
Zdroj: | Molecular pharmacology. 60(1) |
ISSN: | 0026-895X |
Popis: | Murine thymoma cell lines expressing mutated forms of the mdr1b P-glycoprotein were isolated using a novel taxane-based P-glycoprotein inhibitor tRA-96023 (SB-RA-31012). The selection strategy required resistance to a combination of tRA-96023 and colchicine. Five mutations were identified (N350I, I862F, L865F, L868W, and A933T) that reduce the capacity of tRA-96023 to inhibit P-glycoprotein-dependent drug resistance. These mutations also result in a loss of paclitaxel resistance ranging from 47 to 100%. Four mutations are located in the second half of the protein, within or near the proposed transmembrane segment (TMS) 10--11 regions. The fifth mutation (N350I) is within the first half of the protein, proximal (cytoplasmic) to TMS 6. The variant cell line expressing the L868W mutation was subjected to a second round of selection involving tRA-96023 and the toxic drug puromycin. This resulted in the isolation of a cell line expressing a P-glycoprotein with a double mutation. The additional mutation (N988D) is located within TMS 12 and conveys further decreases in resistance to paclitaxel and the capacity of tRA-96023 to inhibit drug resistance. Taken together, the results indicate a significant contribution by the TMS 10--12 portion of the protein to the recognition and transport of taxanes and give evidence that the cytoplasmic region proximal to TMS 6 also plays a role in taxane interactions with P-glycoproteins. Interestingly, mutations within TMS 6 and 12 were found to cause a partial loss of PSC-833 inhibitor activity, suggesting that these regions participate in the interactions with cyclosporin and its derivatives. |
Databáze: | OpenAIRE |
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