Mapping discontinuous epitopes for MRK-16, UIC2 and 4E3 antibodies to extracellular loops 1 and 4 of human P-glycoprotein

Scientific Reports
Shahrooz VahediS V Ambudkar

Abstract

P-glycoprotein (P-gp), an ATP-dependent efflux pump, is associated with the development of multidrug resistance in cancer cells. Antibody-mediated blockade of human P-gp activity has been shown to overcome drug resistance by re-sensitizing resistant cancer cells to anticancer drugs. Despite the potential clinical application of this finding, the epitopes of the three human P-gp-specific monoclonal antibodies MRK-16, UIC2 and 4E3, which bind to the extracellular loops (ECLs) have not yet been mapped. By generating human-mouse P-gp chimeras, we mapped the epitopes of these antibodies to ECLs 1 and 4. We then identified key amino acids in these regions by replacing mouse residues with homologous human P-gp residues to recover binding of antibodies to the mouse P-gp. We found that changing a total of ten residues, five each in ECL1 and ECL4, was sufficient to recover binding of both MRK-16 and 4E3 antibodies, suggesting a common epitope. However, recovery of the conformation-sensitive UIC2 epitope required replacement of thirteen residues in ECL1 and the same five residues replaced in the ECL4 for MRK-16 and 4E3 binding. These results demonstrate that discontinuous epitopes for MRK-16, UIC2 and 4E3 are located in the same regions o...Continue Reading

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Citations

Aug 3, 2019·Drug Metabolism and Disposition : the Biological Fate of Chemicals·Andaleeb SajidSuresh V Ambudkar
Oct 7, 2020·Proceedings of the National Academy of Sciences of the United States of America·Kamil NosolKaspar P Locher
Nov 11, 2020·Proceedings of the National Academy of Sciences of the United States of America·Andaleeb SajidSuresh V Ambudkar
Dec 17, 2019·Trends in Biochemical Sciences·Robert C FordJohn Schuetz
Jul 7, 2021·Molecular Pharmaceutics·Lara García-VarelaGert Luurtsema

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Methods Mentioned

BETA
flow cytometry
glycosylation

Software Mentioned

FlowJo
GraphPad Prism

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