Exhaustive Sampling of Docking Poses Reveals Binding Hypotheses for Propafenone Type Inhibitors of P-Glycoprotein

Abstract

Overexpression of the xenotoxin transporter P-glycoprotein (P-gp) represents one major reason for the development of multidrug resistance (MDR), leading to the failure of antibiotic and cancer therapies. Inhibitors of P-gp have thus been advocated as promising candidates for overcoming the problem of MDR. However, due to lack of a high-resolution structure the concrete mode of interaction of both substrates and inhibitors is still not known. Therefore, structure-based design studies have to rely on protein homology models. In order to identify binding hypotheses for propafenone-type P-gp inhibitors, five different propafenone derivatives with known structure-activity relationship (SAR) pattern were docked into homology models of the apo and the nucleotide-bound conformation of the transporter. To circumvent the uncertainty of scoring functions, we exhaustively sampled the pose space and analyzed the poses by combining information retrieved from SAR studies with common scaffold clustering. The results suggest propafenone binding at the transmembrane helices 5, 6, 7 and 8 in both models, with the amino acid residue Y307 playing a crucial role. The identified binding site in the non-energized state is overlapping with, but not identical to, known binding areas of cyclic P-gp inhibitors and verapamil. These findings support the idea of several small binding sites forming one large binding cavity. Furthermore, the binding hypotheses for both catalytic states were analyzed and showed only small differences in their protein-ligand interaction fingerprints, which indicates only small movements of the ligand during the catalytic cycle. A major reason for the failure of cancer, antibiotic and antiviral therapies is the development of multidrug resistance (MDR). P-glycoprotein (P-gp), an ATP-dependent transport protein located in the membrane of epithelial cells of the kidney, liver, pancreas, colon and the blood-brain barrier, has been linked to the export of a broad variety of xenotoxins. Overexpression of P-gp leads to extrusion of therapeutic drugs and therefore triggers MDR. Thus, identification of potential P-gp inhibitors represents a promising concept for treatment of multiresistant tumours. However, due to lack of high resolution structural information and the polyspecific ligand recognition pattern only very limited information is available on the molecular basis of ligand/transporter interaction. Within this study we characterized the propafenone binding site of P-gp by docking a set of derivatives with known SAR into homology models of P-gp which represent both the apo and the nucleotide-bound state. Poses retrieved are in accordance with results from previous photoaffinity labeling studies and thus pave the way for structure-based in silico screening approaches.