Design and Synthesis of Selenazole‐Containing Peptides for Cocrystallization with P‐Glycoprotein

Tao, Houchao; Weng, Yue; Zhuo, Ren Xi; Chang, Geoffrey; Urbatsch, Ina L.; Zhang, Qinghai

doi:10.1002/cbic.201100048

Cited by 23 publications

(23 citation statements)

References 29 publications

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“…The QZ59‐ RRR and QZ59‐ SSS cyclic hexapeptides were synthesized as described by Tao et al . . The compounds were dissolved in 100% dimethylsulfoxide at 20 m m stock concentration and stored at −20 °C.…”

Section: Methodssupporting

confidence: 83%

Understanding polyspecificity within the substrate‐binding cavity of the human multidrug resistance P‐glycoprotein

Martinez

Arnaud

Hénin³

et al. 2014

The FEBS Journal

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Human P-glycoprotein (P-gp) controls drugs bioavailability by pumping out of the cells many structurally-unrelated drugs. The x-ray structure of the mouse P-gp ortholog was solved with two SSS- and one RRR-enantiomers of the selenohexapeptide inhibitor QZ59, found within the putative drug-binding pocket of the membrane domain outer leaflet. This offered the first opportunity to localize the well-known H- and R- drug-substrate sites in light of QZ59 inhibition mechanisms that were characterized here in cellulo and modelled towards Hoechst 33342 and daunorubicin transport. We found that QZ59-SSS competes efficiently with both substrates, displaying KI,app values of 0.15 and 0.3 μM, respectively 13 and 2 times lower than corresponding Km,app. In contrast, QZ59-RRR non-competitively inhibited daunorubicin transport with moderate efficacy (KI,app = 1.9 μM) and displayed a mixed-type inhibition towards Hoechst 33342 transport, resulting from a mainly non-competitive (Ki2,app = 1.6 μM) and a poor but significant competitive tendency (Ki1,app = 5 μM). These results suppose a positional overlap of QZ59 – drug-transport sites, total for the SSS enantiomer and partial for the RRR one. Crystal structures analysis suggests that the H site overlaps both QZ59-SSS locations while the R-site overlaps the most embedded one.

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Section: Methodssupporting

confidence: 83%

Understanding polyspecificity within the substrate‐binding cavity of the human multidrug resistance P‐glycoprotein

Martinez

Arnaud

Hénin³

et al. 2014

The FEBS Journal

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“…While apo forms of P-gp from mouse and C. elegans were crystallized in the open inward conformation, as was the GltPh aspartate transporter, the latter was shown by EPR spectroscopy to be equally distributed between outward-and inward-facing states (37,38). This result, together with the absence of X-ray structures for P-gp with nucleotides or CsA bound, in an outward-facing form (39), imply that nucleotide or drug binding alone is not sufficient to stabilize an outward-facing conformer (9,37,38). Concomitant binding of nucleotides, lipids, and drugs did, however, show an increase in the population of a smaller conformer.…”

Section: Discussionmentioning

confidence: 99%

Mass spectrometry reveals synergistic effects of nucleotides, lipids, and drugs binding to a multidrug resistance efflux pump

Marcoux¹,

Wang²,

Politis³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Multidrug resistance is a serious barrier to successful treatment of many human diseases, including cancer, wherein chemotherapeutics are exported from target cells by membrane-embedded pumps. The most prevalent of these pumps, the ATP-Binding Cassette transporter P-glycoprotein (P-gp), consists of two homologous halves each comprising one nucleotide-binding domain and six transmembrane helices. The transmembrane region encapsulates a hydrophobic cavity, accessed by portals in the membrane, that binds cytotoxic compounds as well as lipids and peptides. Here we use mass spectrometry (MS) to probe the intact P-gp small molecule-bound complex in a detergent micelle. Activation in the gas phase leads to formation of ions, largely devoid of detergent, yet retaining drug molecules as well as charged or zwitterionic lipids. Measuring the rates of lipid binding and calculating apparent K D values shows that up to six negatively charged diacylglycerides bind more favorably than zwitterionic lipids. Similar experiments confirm binding of cardiolipins and show that prior binding of the immunosuppressant and antifungal antibiotic cyclosporin A enhances subsequent binding of cardiolipin. Ion mobility MS reveals that P-gp exists in an equilibrium between different states, readily interconverted by ligand binding. Overall these MS results show how concerted small molecule binding leads to synergistic effects on binding affinities and conformations of a multidrug efflux pump.mass spectrometry from native state | real time substrate monitoring P -glycoprotein (P-gp) is an ATP-driven low-specificity efflux pump that plays an important role in the clearance of xenotoxins (1, 2). P-gp is also a member of the ATP-Binding Cassette (ABC) family of transporters and exports hydrophobic cytotoxic compounds as well as natural products, cyclic, and linear peptides (1, 3-5). Overexpression of P-gp in tumor cells impairs targeted drug delivery and is a major pitfall for chemotherapies. Small molecule substrates partition in the plasma membrane (6, 7), before binding in the internal hydrophobic cavity formed in the inward conformation of the pump (8). Export is then thought to proceed in an ATP-dependent way through conformational changes from the inward to the outward facing forms, evidenced by FRET spectroscopy (9). Recent highresolution structures of eukaryotic P-gp from mouse (8) and Caenorhabditis elegans (10) were obtained in inward conformations. Two prokaryotic homologs of P-gp [Sav1866 (11) and MsbA (12)] were captured crystallographically in outward-facing conformers, reflecting ATP-bound states, as well as two different inward states for MsbA. From these X-ray structures it is possible to build up a picture of P-gp, alternating between inward-and outward-facing conformations.Despite decades of careful biochemical studies (13), and recent insights from crystallography, many questions remain, however. Specifically it has not yet been possible to trap P-gp in an outwardfacing state or to show how substrate binding activates ATPase act...

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“…Therefore, in our design strategy described below, we chose to maintain S stereochemistry at all chiral centers. 3) Comparison of inhibitory activity of QZ59S-SSS for Chinese hamster or mouse P-gp (IC 50 = 2.7 mm) [15] with that of potent inhibitor valspodar [16] suggests a role of macrocyclic peptidic nature of valspodar for its high potency as compared to QZ59Se-SSS ( Figure 1). During ongoing phase III clinical trials, valspodar has shown very restricted oral bioavailability which, along with its efficacy and safety concerns, further suggested the need for optimization.…”

mentioning

confidence: 99%

Design, Synthesis, and Biological Evaluation of (S)‐Valine Thiazole‐Derived Cyclic and Noncyclic Peptidomimetic Oligomers as Modulators of Human P‐Glycoprotein (ABCB1)

et al. 2013

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Multidrug resistance (MDR) caused by ATP-binding cassette (ABC) transporter P-glycoprotein (P-gp) through extrusion of anticancer drugs from the cells is a major cause of failure to cancer chemotherapy. Previously, selenazole containing cyclic peptides were reported as P-gp inhibitors and these were also used for co-crystallization with mouse P-gp, which has 87% homology to human P-gp. It has been reported that human P-gp, can simultaneously accommodate 2-3 moderate size molecules at the drug binding pocket. Our in-silico analysis based on the homology model of human P-gp spurred our efforts to investigate the optimal size of (S)-valine-derived thiazole units that can be accommodated at drug-binding pocket. Towards this goal, we synthesized varying lengths of linear and cyclic derivatives of (S)-valine-derived thiazole units to investigate the optimal size, lipophilicity and the structural form (linear and cyclic) of valine-derived thiazole peptides that can accommodate well in the P-gp binding pocket and affects its activity, previously an unexplored concept. Among these oligomers, lipophilic linear- (13) and cyclic-trimer (17) derivatives of QZ59S-SSS were found to be the most and equally potent inhibitors of human P-gp (IC50 = 1.5 μM). Cyclic trimer and linear trimer being equipotent, future studies can be focused on non-cyclic counterparts of cyclic peptides maintaining linear trimer length. Binding model of the linear trimer (13) within the drug-binding site on the homology model of human P-gp represents an opportunity for future optimization, specifically replacing valine and thiazole groups in the non-cyclic form.

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Design and Synthesis of Selenazole‐Containing Peptides for Cocrystallization with P‐Glycoprotein

Cited by 23 publications

References 29 publications

Understanding polyspecificity within the substrate‐binding cavity of the human multidrug resistance P‐glycoprotein

Understanding polyspecificity within the substrate‐binding cavity of the human multidrug resistance P‐glycoprotein

Mass spectrometry reveals synergistic effects of nucleotides, lipids, and drugs binding to a multidrug resistance efflux pump

Design, Synthesis, and Biological Evaluation of (S)‐Valine Thiazole‐Derived Cyclic and Noncyclic Peptidomimetic Oligomers as Modulators of Human P‐Glycoprotein (ABCB1)

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