Influence of breast cancer resistance protein (Abcg2) and p-glycoprotein (Abcb1a) on the transport of imatinib mesylate (Gleevec) across the mouse blood-brain barrier

Imatinib, a protein tyrosine kinase inhibitor, may prevent the growth of glioblastoma cells. Unfortunately, its brain distribution is restricted by p-glycoprotein (p-gp or multidrug resistance protein Mdr1a), and probably by breast cancer resistance protein (Bcrp1), two efflux pumps expressed at the blood-brain barrier (BBB). We have used in situ brain perfusion to investigate the mechanisms of imatinib transport across the mouse BBB. The brain uptake of imatinib in wild-type mice was limited by saturable efflux processes. The inhibition of p-gp, by valspodar and zosuquidar, increased imatinib uptake (2.5-fold), as did the deficiency of p-gp in Mdr1a/1b(-/-) mice (5.5-fold). Perfusing imatinib with the p-gp/Bcrp1 inhibitor, elacridar, enhanced the brain uptake of imatinib in wild-type (4.1-fold) and Mdr1a/1b(-/-) mice (1.2-fold). However, the brain uptake of imatinib was similar in wild-type and Bcrp1(-/-) mice when it was perfused at a non-saturating concentration. The brain uptake of CGP74588, an active metabolite of imatinib, was low. It was increased by perfusion with elacridar (twofold), but not with valspodar and zosuquidar. CGP74588 uptake was 1.5 times greater in Bcrp1(-/-) mice than in wild-type mice. These data suggest that imatinib transport at the mouse BBB is limited by p-gp and probably by Bcrp1, and that CGP74588 transport is restricted by Bcrp1.     

Discovery of tetrahydroindazoles as a novel class of potent and in vivo efficacious gamma secretase modulators

The identification and optimization of a novel series of centrally efficacious gamma secretase modulators (GSMs) offering an alternative to the privileged aryl imidazole motif is described. Chiral bicyclic tetrahydroindazolyl amine substituted triazolopyridines were identified as structurally distinct novel series of GSMs. Representative compound BI-1408 ((R)-42) was demonstrated to be centrally efficacious in rats at a 30?mg/kg oral dose.     

The multi-xenobiotic resistance phenotype as a tool to biomonitor the environment

Organisms use a variety of cellular mechanisms to avoid the effects of toxins. These strategies include de-toxification of putative toxins, sequestration of the toxins or the utilization of transport mechanisms to actually prevent the entry and accumulation of toxins in the cells. These toxin avoidance mechanisms, which presumably evolved in response to natural toxins, can also be used to counter the effects of anthropogenic compounds introduced into the environment by the activities of our modern society. In this article we discuss (1) the use of transport mechanism strategies to protect against toxins and (2) the possible use of these mechanisms as biomarkers indicative of exposure to man-made toxins. We will first review the characteristics of these transport mechanisms, including their biology, genetics and molecular properties and then discuss their use as biomarkers.     

SP analysis may be used to identify cancer stem cell populations

Side populations (SP), as defined by Hoechst exclusion in flow cytometry, have been described a few years ago. While they represent only a small fraction of the whole cell population, their properties confer an important place in several investigations. SP cells express high levels of various members of ABC transporters family, such as MDR1 and BCRP, which are responsible for drug resistance. Targeting SP could improve cancer therapy by blocking these transporters. In addition, SP appear to be enriched in stem cells, cells that play a pivotal role in normal development and cancer biology. Thus, they could provide a useful tool and a readily accessible source for stem cell studies in both the normal and cancerous settings. However, these cells are poorly defined and pose challenges in their identification and isolation, particularly since they are few in number. Thus, better characterization of SP will advance our understanding of stem cells and will provide us an accessible target for drug resistance in cancer therapy.     

Minocycline and riluzole brain disposition: interactions with p-glycoprotein at the blood-brain barrier

Amyotrophic lateral sclerosis is a neurodegenerative fatal disease. The only drug recognized to increase the survival time is riluzole(RLZ). In animal models, minocycline (MNC) delayed the onset of the disease and increased the survival time (in combination with RLZ). The objective of our work was to study the interactions between RLZ, MNC and the efflux pump p-glycoprotein (p-gp) at the blood–brain barrier. We investigated these two drugs as: (i) p-gp substrates by comparing their brain uptake in CF1 mdr1a (−/−) and mdr1a (+/+) mice, (ii) p-gp modulators by studying their effect on the cerebral uptake of digoxin. mdr1a (−/−) mice showed higher brain uptake of MNC and RLZ than mdr1a (+/+) (in a 1.6- and 1.4-fold, respectively); and in mdr1a (+/+) mice pre-treated with repeated doses of MNC, brain uptake of digoxin was increased. When both drugs were administrated to mdr1a (+/+) mice, MNC increased the brain uptake of RLZ in a 2.1-fold. In conclusion, MNC and RLZ are both p-gp substrates. MNC is also a p-gp inhibitor and increases the brain diffusion of RLZ. In vitro experiments with the GPNT cell line confirmed these results. These interactions should be taken into account in the design of future clinical trials.     

The multi-xenobiotic resistance phenotype as a tool to biomonitor the environment

Organisms use a variety of cellular mechanisms to avoid the effects of toxins. These strategies include de-toxification of putative toxins, sequestration of the toxins or the utilization of transport mechanisms to actually prevent the entry and accumulation of toxins in the cells. These toxin avoidance mechanisms, which presumably evolved in response to natural toxins, can also be used to counter the effects of anthropogenic compounds introduced into the environment by the activities of our modern society. In this article we discuss (1) the use of transport mechanism strategies to protect against toxins and (2) the possible use of these mechanisms as biomarkers indicative of exposure to man-made toxins. We will first review the characteristics of these transport mechanisms, including their biology, genetics and molecular properties and then discuss their use as biomarkers.     

Modulation of p-glycoprotein function by caveolin-1 phosphorylation

p-glycoprotein (p-gp) is an ATP-binding cassette transporter and its overexpression is responsible for the acquisition of the multidrug resistance phenotype in human tumors. p-gp is localized at the blood-brain barrier and is involved in brain cytoprotection. Our previous work used immunoprecipitation to show that caveolin-1 can interact with p-gp. In this study, we provide evidence that caveolin-1 regulates p-gp transport activity in a rat brain endothelial cell line (RBE4). Down-regulation of caveolin-1 by siRNA reduced the interaction between p-gp and caveolin-1, followed by a decrease in [3H]-Taxol and [3H]-Vinblastine accumulation in RBE4 cells. The latter result showed that down-regulation of caveolin-1 enhanced p-gp transport activity. RBE4 cells were also transfected with Sarcoma in order to modulate caveolin-1 phosphorylation. Overexpression of Sarcoma, a protein tyrosine kinase, stimulated caveolin-1 phosphorylation and increased both [3H]-Taxol and [3H]-Vinblastine accumulation as well as Hoechst 33342 accumulation. Transfection of caveolin-1 inhibits p-gp transport activity. Conversely, transfection of the mutant cavY14F decreased the p-gp/caveolin-1 interaction and reduced accumulation of the two p-gp substrates. Thus, our data show that caveolin-1 regulates p-gp function through the phosphorylation state of caveolin-1 in endothelial cells from the blood-brain barrier.     

Oral administration of a potent and selective non-peptidic BACE-1 inhibitor decreases beta-cleavage of amyloid precursor protein and amyloid-beta production in vivo

Generation and deposition of the amyloid beta (Abeta) peptide following proteolytic processing of the amyloid precursor protein (APP) by BACE-1 and gamma-secretase is central to the aetiology of Alzheimer's disease. Consequently, inhibition of BACE-1, a rate-limiting enzyme in the production of Abeta, is an attractive therapeutic approach for the treatment of Alzheimer's disease. We have designed a selective non-peptidic BACE-1 inhibitor, GSK188909, that potently inhibits beta-cleavage of APP and reduces levels of secreted and intracellular Abeta in SHSY5Y cells expressing APP. In addition, we demonstrate that this compound can effectively lower brain Abeta in vivo. In APP transgenic mice, acute oral administration of GSK188909 in the presence of a p-glycoprotein inhibitor to markedly enhance the exposure of GSK188909 in the brain decreases beta-cleavage of APP and results in a significant reduction in the level of Abeta40 and Abeta42 in the brain. Encouragingly, subchronic dosing of GSK188909 in the absence of a p-glycoprotein inhibitor also lowers brain Abeta. This pivotal first report of central Abeta lowering, following oral administration of a BACE-1 inhibitor, supports the development of BACE-1 inhibitors for the treatment of Alzheimer's disease.     

Roxithromycin potentiates the effects of chloroquine and mefloquine on multidrug-resistant Plasmodium falciparum in vitro

Chloroquine (CQ) and mefloquine (MQ) are no longer potent antimalarial drugs due to the emergence of resistant Plasmodium falciparum. Combination therapy has become the standard for many regimes in overcoming drug resistance. Roxithromycin (ROM), a known p-glycoprotein inhibitor, is reported to have antimalarial activity and it is hoped it will potentiate the effects of both CQ/MQ and reverse CQ/MQ-resistance. We assayed the effects of CQ and MQ individually and in combination with ROM on synchronized P. falciparum (Dd2 strain) cultures. The IC(50) values of CQ and MQ were 60.0+/-5.0 and 16.0+/-3.0 ng/ml; these were decreased substantially when combined with ROM. Isobolograms indicate that CQ-ROM combinations were relatively more synergistic (mean FICI 0.70) than MQ-ROM (mean FICI 0.85) with their synergistic effect at par with CQ-verapamil (VRP) (mean FICI 0.64) and MQ-VRP (mean FICI 0.60) combinations. We conclude that ROM potentiates the CQ/MQ response on multidrug-resistant P. falciparum.