P-glycoprotein transporter in drug development

Proceedings of the National Academy of Sciences, India Section B: Biological Sciences, 2014

Permeability or P-glycoprotein (P-gp), present in both prokaryotes and eukaryotes is an essential protein belonging to the adenosine triphosphate (ATP) binding cassette transporter family. It functions as a protective barrier by extrusion of wide range of substrates including toxins, xenobiotic compounds and drugs from the cell. These are ubiquitous integral membrane proteins that have ATPase activity for substrate transport across the lipid membrane. The potential role of P-gp in efflux mechanism is to avoid drug accumulation and to provide intrinsic resistance to a broad range of anticancer compounds against sarcoma, breast cancer and certain types of leukemia. This review recapitulates the structural and functional aspects of core domains of P-gp. Furthermore, various proposed mechanisms of translocation of substrates across the membrane which explain the conformational changes in different domains upon ATP binding and hydrolysis. Another critical aspect focuses on different modulators and their mode of binding on P-gp which facilitates the inhibition of efflux mechanism. Ten models of drug efflux mechanism have been illustrated and a comparative account of their applicability and limitations given, paving way for further improvements. In the present study it has been observed that contrary to the earlier reported models where transmembrane domain is the preferred binding site of ligands resulting in their efflux through ATP hydrolysis at nucleotide binding domain (NBD) site. The more hydrophilic NBD appears to be the appropriate binding site for majority of the hydrophobic third and fourth generation modulators thus inhibiting the binding and hydrolysis of ATP resulting in inhibition of efflux.

Pharmacological Research, 2003

Drug efflux pumps like P-glycoprotein (P-gp) and multidrug resistance (MDR) proteins were recognized to posses functional role in determining the pharmacokinetics of drugs administered by peroral as well as parenteral route. Advancements in molecular biology, to some extent, had revealed the structure, localization and functional role of P-glycoprotein and its mechanism of drug efflux. Broad substrate recognization by this protein and clinical implications of its inhibition has revolutionized cancer chemotherapy leading to design and development of novel P-glycoprotein inhibitors. In the recent times, the application of these inhibitors in improving peroral drug delivery has gained special interest. Inhibition of P-glycoprotein improves intestinal absorption and tissue distribution while reducing the substrate metabolism and its elimination. Eventually, various screening methodologies have been developed for determining the activity of P-glycoprotein, kinetics of drug transport and identification of substrates and inhibitors. In the present review, techniques used for screening P-glycoprotein inhibitors and the scope of these inhibitors in optimizing peroral drug absorption and pharmacokinetics are discussed along with a brief introduction to P-glycoprotein, its physiological function and active role in extrusion of drugs.

1999

Modeling of P-glycoprotein-involved epithelial drug transport in MDCK cells. Am. J. Physiol. 277 (Renal Physiol. 46): F84-F96, 1999.-P-glycoprotein (P-gp) on the apical membranes of epithelial cells is known as a drug efflux pump. However, unclear is its integral quantitative role in the overall epithelial drug transfer, which also involves distinct diffusion processes in parallel and sequence. We used a simple threecompartment model to obtain kinetic parameters of each drug transfer mechanism, which can quantitatively describe the transport time courses of P-gp substrates, digoxin and vinblastine, across P-gp-expressing MDCK cell monolayers grown on permeable filters. Our results show that the model, which assumes a functionally single drug efflux pump in the apical membrane with diffusion across two membranes and intercellular junctions, is the least complex model with which to quantitatively reproduce the characteristics of the data. Interestingly, the model predicts that the MDCK apical membranes are less diffusion permeable than the basolateral membrane for both drugs and that the distribution volume of vinblastine is 10-fold higher than that of digoxin. Additional experiments verified these model predictions. The modeling approach is feasible to quantitatively describe overall kinetic picture of epithelial drug transport. Further model refinement is necessary to incorporate other modes of drug transport such as transcytosis. Also, whether P-gp solely accounts for the pump function in this model awaits more studies. P-glycoprotein; multidrug resistance; diffusion; itraconazole 0363-6127/99 $5.00

ABC ATP-binding cassette (transport protein) MDR Multidrug resistance Pgp P-Glycoprotein (MDR1) (Q)SAR (Quantitative) structure activity relationship TM Transmembrane (domain) H-bonding Hydrogen bonding Symbols A Strong H-bond acceptor (or strong electron donor) a Weak H-bond acceptor (or weak electron donor) ∆G aw Free Energy of partitioning into the air-water interface ∆G lw Free Energy of drug partitioning into the lipid membrane ∆G t Free Energy of drug binding to the transporter Pgp from within the lipid phase ∆G app Apparent Free Energy of binding to Pgp from the aqueous phase

General Pharmacology: The Vascular System, 1996

1. P-glycoprotein (P-gp) is a transmembrane protein involved in ATP-dependent efflux of various structurally unrelated anticancer drugs. Its overexpression in cancer cells decreases intracellular drug concentrations and, thus, confers a multidrug resistance phenotype.

European Journal of Biochemistry, 1997

In this paper, we show that P-glycoprotein contains two distinct sites for drug binding and transport, and that, unexpectedly, these sites interact in a positively cooperative manner. The kinetics of transport of rhodamine 123 and Hoechst 33342 in isolated P-glycoprotein-rich plasma membrane vesicles from Chinese hamster ovary CH'B30 cells were followed by continuous fluorescence monitoring. Each substrate stimulated P-glycoprotein-mediated transport of the other. Colchicine and quercetin stimulated rhodamine 123 transport and inhibited Hoechst 33342 transport. In contrast, anthracyclines such as daunorubicin and doxorubicin stimulated Hoechst 33342 transport and inhibited rhodamine 123 transport. Vinblastine, actinomycin D, and etoposide inhibited transport of both dyes. The results are consistent with a functional model of P-glycoprotein containing at least two positively cooperative sites (H site and R site) for drug binding and transport. This model is consistent with earlier observations of competitive and noncompetitive effects of P-glycoprotein substrates and chemosensitizers. Such a two-site model may be fundamental to multidrug transport by P-glycoprotein, and it may be a feature common to other ATPdependent transporters belonging to the ATP-binding cassette superfamily.

Molecular Pharmaceutics, 2006

Multidrug resistance mediated by ATP binding cassette (ABC) transporters such as P-glycoprotein (P-gp) represents a serious problem for the development of effective anticancer drugs. In addition, P-gp has been shown to reduce oral absorption, modulate hepatic, renal, or intestinal elimination, and restrict blood-brain barrier penetration of several drugs. Consequently, there is a great interest in anticipating whether drug candidates are P-gp substrates or inhibitors. In this respect, two different computational models have been developed. A method for discriminating P-gp substrates and nonsubstrates has been set up based on calculated molecular descriptors and multivariate analysis using a training set of 53 diverse drugs. These compounds were previously classified as P-gp substrates or nonsubstrates on the basis of the efflux ratio from Caco-2 permeability measurements. The program Volsurf was used to compute the compounds' molecular descriptors. The descriptors were correlated to the experimental classes using partial least squares discriminant analysis (PLSD). The model was able to predict correctly the behavior of 72% of an external set of 272 proprietary compounds. Thirty of the 53 previously mentioned drugs were also evaluated for P-gp inhibition using a calcein-AM (CAM) assay. On the basis of these additional P-gp functional data, a PLSD analysis using GRIND-pharmacophorebased descriptors was performed to model P-gp substrates having poor or no inhibitory activity versus inhibitors having no evidence of significant transport. The model was able to discriminate between 69 substrates and 56 inhibitors taken from the literature with an average accuracy of 82%. The model allowed also the identification of some key molecular features that differentiate a substrate from an inhibitor, which should be taken into consideration in the design of new candidate drugs. These two models can be implemented in a virtual screening funnel.

Perspectives in Drug Discovery and Design, 2000

We have investigated the relationship between drug retention in immobilized liposome partitioning chromatography and liposome partitioning and found a strong linear correlation. Separate linear relationships were found depending on the charge of the compound when liposome chromatographic measurements were related to the octanol/water partition coefficients. We have also investigated the importance of the water/octanol partition coefficient in quantitative structure–property relationships related to drug transport properties. The studies show that the inclusion of a parameter related to lipophilicity causes only, at best, a marginal increase in internal predictivity and, at worst, a decrease in external predictivity. The studies also show that parameters related to hydrogen bonding, polarizability and size are important properties that need to be included in quantitative models for drug transport processes. We believe that the use of multivariate characterizations of compounds based on non-composite parameters may result in better and more predictive models compared with models based on parameters of a more composite nature when investigating the possibilities to establish quantitative structure–property relationships.

Computational and Structural Biotechnology Journal, 2019

Organic anion and cation transporting proteins (OATs, OATPs, and OCTs), as well as the Multidrug and Toxin Extrusion (MATE) transporters of the Solute Carrier (SLC) family are playing a pivotal role in the discovery and development of new drugs due to their involvement in drug disposition, drug-drug interactions, adverse drug effects and related toxicity. Computational methods to understand and predict clinically relevant transporter interactions can provide useful guidance at early stages in drug discovery and design, especially if they include contemporary data science approaches. In this review, we summarize the current state-of-the-art of computational approaches for exploring ligand interactions and selectivity for these drug (uptake) transporters. The computational methods discussed here by highlighting interesting examples from the current literature are ranging from semiautomatic data mining and integration, to ligand-based methods (such as quantitative structureactivity relationships, and combinatorial pharmacophore modeling), and finally structure-based methods (such as comparative modeling, molecular docking, and molecular dynamics simulations). We are focusing on promising computational techniques such as fold-recognition methods, proteochemometric modeling or techniques for enhanced sampling of protein conformations used in the context of these ADMET-relevant SLC transporters with a special focus on methods useful for studying ligand selectivity.

Proceedings of the National Academy of Sciences, 2020

P-glycoprotein (P-gp), also known as ABCB1, is a cell membrane transporter that mediates the efflux of chemically dissimilar amphipathic drugs and confers resistance to chemotherapy in most cancers. Homologous transmembrane helices (TMHs) 6 and 12 of human P-gp connect the transmembrane domains with its nucleotide-binding domains, and several residues in these TMHs contribute to the drug-binding pocket. To investigate the role of these helices in the transport function of P-gp, we substituted a group of 14 conserved residues (seven in both TMHs 6 and 12) with alanine and generated a mutant termed 14A. Although the 14A mutant lost the ability to pump most of the substrates tested out of cancer cells, surprisingly, it acquired a new function. It was able to import four substrates, including rhodamine 123 (Rh123) and the taxol derivative flutax-1. Similar to the efflux function of wild-type P-gp, we found that uptake by the 14A mutant is ATP hydrolysis-, substrate concentration-, and t...