Synthesis and Anticancer Cytotoxicity of Azaaurones Overcoming Multidrug Resistance.

The resistance of tumors against anticancer drugs is a major impediment for chemotherapy. Tumors often develop multidrug resistance as a result of the cellular efflux of chemotherapeutic agents by ABC transporters such as P-glycoprotein (ABCB1/P-gp), Multidrug Resistance Protein 1 (ABCC1/MRP1), or Breast Cancer Resistance Protein (ABCG2/BCRP). By screening a chemolibrary comprising 140 compounds, we identified a set of naturally occurring aurones inducing higher cytotoxicity against P-gp-overexpressing multidrug-resistant (MDR) cells versus sensitive (parental, non-P-gp-overexpressing) cells. Follow-up studies conducted with the P-gp inhibitor tariquidar indicated that the MDR-selective toxicity of azaaurones is not mediated by P-gp. Azaaurone analogs possessing pronounced effects were then designed and synthesized. The knowledge gained from structure-activity relationships will pave the way for the design of a new class of anticancer drugs selectively targeting multidrug-resistant cancer cells.

Lathyrol and epoxylathyrol derivatives: Modulation of Cdr1p and Mdr1p drug-efflux transporters of Candida albicans in Saccharomyces cerevisiae model.

Macrocyclic diterpenes were previously found to be able to modulate the efflux pump activity of Candida albicans multidrug transporters. Most of these compounds were jatrophanes, but only a few number of lathyrane-type diterpenes was evaluated. Therefore, the aim of this study was to evaluate the ability of nineteen structurally-related lathyrane diterpenes (1-19) to overcome the drug-efflux activity of Cdr1p and Mdr1p transporters of C. albicans, and get some insights on their structure-activity relationships. The transport assay was performed by monitoring Nile Red (NR) efflux in a Saccharomyces cerevisiae strain overexpressing the referred efflux pumps from C. albicans. Moreover, a chemosensitization assay was performed in order to evaluate the type of interaction between the inhibitory compounds and the antifungal drug fluconazole. Compounds 1-13 were previously isolated from Euphorbia boetica or obtained by derivatization, and compounds 14-19 were prepared by chemical transformations of compound 4. In the transport assays, compounds 14-19 revealed the strongest inhibitory activity of the Cdr1p efflux pump, ranging from 65 to 85%. Concerning Mdr1p efflux pump, the most active compounds were 1, 3, 6, 8, and 12 (75-85%). When used in combination with fluconazole, epoxyboetirane K (2) and euphoboetirane N (18) revealed synergistic effects in the AD-CDR1 yeast strain, overexpressing the Cdr1p transporter, through their ability to reduce the effective concentration of the antifungal drug by 23- and 52-fold, respectively.

Evaluation of Jatrophane Esters from Euphorbia spp. as Modulators of Candida albicans Multidrug Transporters.

Twenty-nine jatrophane esters (1-10, 12-30) and one lathyrane (11) diterpenoid ester isolated from Euphorbia species were evaluated for their capacity to inhibit drug-efflux activities of the primary ABC transporter CaCdr1p and the secondary MFS transporter CaMdr1p of Candida albicans, in yeast strains overexpressing the corresponding transporter. These diterpenoid esters were obtained from Euphorbia semiperfoliata (1-10), E. insularis (11), and E. dendroides (12-30) and included five new compounds, euphodendroidins P-T (26-30). The jatrophane esters 12 and 23 were found to inhibit the efflux of Nile Red (NR) mediated by the two multidrug transporters, at 85-64% for CaCdr1p and 79-65% for CaMdr1p. In contrast, compound 21 was selective for CaCdr1p and induced a strong inhibition (92%), whereas compound 8 was selective for CaMdr1p, with a 74% inhibition. It was demonstrated further that potency and selectivity are sensitive to the substitution pattern on the jatrophane skeleton. However, these compounds were not transported and showed no synergism with fluconazole cytotoxicity.

Pharmacokinetic interactions in mice between irinotecan and MBL-II-141, an ABCG2 inhibitor.

PURPOSE: The chromone derivative MBL-II-141, specifically designed to inhibit ABCG2, was previously demonstrated to combine strong inhibition potency, low toxicity and good efficiency in reversing resistance to irinotecan in a xenografted mouse model. Here, the pharmacokinetic interactions in mice between irinotecan, its active metabolite SN-38 and MBL-II-141 were characterized quantitatively in the blood and in the brain. METHODS: Compartmental models were used to fit the data. Goodness-of-fit was assessed by simulation-based diagnostic tools. RESULTS: Irinotecan increased the MBL-II-141 apparent clearance and Vss 1.5-fold, probably by increasing the MBL-II-141 unbound fraction. MBL-II-141 decreased the total apparent clearance of irinotecan by 23%, by decreasing its biliary clearance. MBL-II-141 increased 3-fold the brain accumulation of irinotecan, as a result of the rise of systemic exposure combined with the inhibition of ABCG2-mediated efflux at the blood-brain barrier. Finally, SN-38 exposure was increased by 1.16-fold under treatment with MBL-II-141, owing to the higher irinotecan exposure with increased metabolism towards the formation of SN-38. CONCLUSIONS: These results may help to anticipate the pharmacokinetic interactions between MBL-II-141 and other ABCG2 substrates. The irinotecan-MBL-II-141 interaction is also expected to occur in humans. Copyright © 2017 John Wiley & Sons, Ltd.

Overcoming Multidrug Resistance in Candida albicans: Macrocyclic Diterpenes from Euphorbia Species as Potent Inhibitors of Drug Efflux Pumps.

Thirteen macrocyclic diterpenes (1-13) of the jatrophane and lathyrane types, either isolated from Euphorbia species or obtained by chemical derivatization, were evaluated for their ability to inhibit the drug efflux activity of Candida albicans CaCdr1p and CaMdr1p multidrug transporters overexpressed in a Saccharomyces cerevisiae strain. Their inhibitory potential was assessed through a functional assay of Nile Red accumulation monitored by flow cytometry. A chemosensitization assay, using the checkerboard method, was also performed with the active compounds in order to evaluate their type of interaction with fluconazole.In the transport assay, most compounds were found to inhibit both transporters, most likely as non-substrates, as shown by relative resistance indices close to unity. In contrast, the jatrophanes euphopubescenol (10) and euphomelliferene A (11) were selective for CaMdr1p and CaCdr1p, respectively. Moreover, when used in combination with fluconazole, compounds 12 and 13 displayed strong synergistic interactions (FICI = 0.071) against the yeast strain overexpressing CaMdr1p, decreasing the MIC80 of the antifungal agent 13-fold. Both compounds were also able to reduce the effective concentration of this antifungal agent by 4- to 8-fold against an azole-resistant clinical isolate of C. albicans (F5).

Dual properties of hispidulin: antiproliferative effects on HepG2 cancer cells and selective inhibition of ABCG2 transport activity.

Hepatocellular carcinoma is the third most common cause of cancer-related deaths worldwide. Furthermore, the existing pharmacological-based treatments are insufficiently effective and generate many side effects. Hispidulin (6-methoxy-5,7,4'-trihydroxyflavone) is a flavonoid found in various medicinal herbs that present antineoplastic properties. Here we evaluated how modulation of reactive oxygen species (ROS) and alterations of antioxidant defenses could be associated to the antiproliferative effects of hispidulin in HepG2 cells. In addition, we studied the inhibitory activity of hispidulin on the efflux of drugs mediated by ABC transporters involved in multidrug resistance. In order to understand the increase of intracellular ROS promoted by hispidulin, we investigated the mRNA expression levels and activities of antioxidant enzymes, and the GSH/GSSG ratio. We showed that hispidulin significantly down-regulated the transcription levels of catalase, leading to reduction of enzyme activity and decrease of the GSH content. We also observed that, in the presence of N-acetylcysteine or exogenous catalase, the proliferation was lowered back to the control levels. These data clearly indicate a strong involvement of intracellular ROS levels for triggering the antiproliferative effects. We also demonstrated that the inhibition produced by hispidulin on drug efflux was specific for ABCG2, since no effects were observed with ABCB1 and ABCC1. Furthermore, HepG2 cells were more sensitive to hispidulin-mediated cell death than immortalized L929 fibroblasts, suggesting a differential toxicity of this compound between tumor and non-tumor cell lines. Our results suggest that hispidulin constitutes a promising candidate to sensitize chemoresistant cancer cells overexpressing ABCG2.

Biologically active carbazole derivatives: focus on oxazinocarbazoles and related compounds.

Four series of carbazole derivatives, including N-substituted-hydroxycarbazoles, oxazinocarbazoles, isoxazolocarbazolequinones, and pyridocarbazolequinones, were studied using diverse biological test methods such as a CE-based assay for CK2 activity measurement, a cytotoxicity assay with IPC-81 cell line, determination of MIC of carbazole derivatives as antibacterial agents, a Plasmodium falciparum susceptibility assay, and an ABCG2-mediated mitoxantrone assay. Two oxazinocarbazoles Ib and Ig showed CK2 inhibition with IC50 = 8.7 and 14.0 µM, respectively. Further chemical syntheses were realized and the 7-isopropyl oxazinocarbazole derivative 2 displayed a stronger activity against CK2 (IC50 = 1.40 µM). Oxazinocarbazoles Ib, Ig, and 2 were then tested against IPC-81 leukemia cells and showed the ability to induce leukemia cell death with IC50 values between 57 and 62 µM. Further investigations were also reported on antibacterial and antiplasmodial activities. No significant inhibitory activity on ABCG2 efflux pump was detected.

Identification of specific reversal agents for Leishmania ABCI4-mediated antimony resistance by flavonoid and trolox derivative screening.

OBJECTIVES: To identify reversal agents for the Leishmania ABCI4 transporter that confers resistance to antimony. METHODS: Selective ABCI4 inhibitors among a series of 15 flavonoid and trolox derivatives or analogues were investigated by evaluating their ability to reverse antimony resistance in Leishmania parasites overexpressing ABCI4. Among the compounds screened, N-ethyltrolox carboxamide (compound D2) produced the highest reversal activity. In order to optimize the activity of D2, we synthesized a series of 10 derivatives by condensation of various amines with trolox. RESULTS: Analysis of antimony resistance reversal activity showed that N-propyltrolox carboxamide (compound D4) was the most potent ABCI4 inhibitor, with reversal activity being maintained in the intracellular amastigote stage. In addition, trolox derivatives significantly reverted the resistance to zinc protoporphyrin. The mechanism of action of these active derivatives was found to be related to significant reversion of Sb(III) and zinc protoporphyrin accumulation and to a decrease in drug efflux. CONCLUSIONS: Our findings suggest that trolox derivatives D2 and D4 could be considered to be specific reversal agents targeting the Leishmania ABCI4 transporter. The structure-activity relationship obtained in the present study highlights the importance of the size and length of the alkyl substituent linked to trolox. Furthermore, the structural data obtained provide valuable information for the further development of new, even more specific and potent Leishmania ABCI4 reversal agents.

Jatrophanes from Euphorbia squamosa as potent inhibitors of Candida albicans multidrug transporters.

A series of structurally related jatrophane diterpenoids (1-6), including the new euphosquamosins A-C (4-6), was purified from the Iranian spurge Euphorbia squamosa and evaluated for its capacity to inhibit drug efflux by multidrug transporters of Candida albicans. Three of these compounds showed an interesting profile of activity. In particular, deacetylserrulatin B (2) and euphosquamosin C (6) strongly inhibited the drug-efflux activity of the primary ABC-transporter CaCdr1p, an effect that translated, in a yeast strain overexpressing this transporter, into an increased sensitivity to fluconazole. These compounds were transported by CaCdr1p, as shown by the observation of an 11-14-fold cross-resistance of yeast growth, and could also inhibit the secondary MFS-transporter CaMdr1p. In contrast, euphosquamosin A (4) was selective for CaCdr1p, possibly as a result of a different binding mode. Taken together, these observations suggest jatrophane diterpenes to be a new class of potent inhibitors of multidrug transporters critical for drug resistance in pathogenic yeasts.

Localization of putative binding sites for cyclic guanosine monophosphate and the anti-cancer drug 5-fluoro-2'-deoxyuridine-5'-monophosphate on ABCC11 in silico models.

BACKGROUND: The Multidrug Resistance Protein ABCC11/MRP8 is expressed in physiological barriers and tumor breast tissues in which it secretes various substrates including cGMP (cyclic guanosine monophosphate) and 5FdUMP (5-fluoro-2'-deoxyuridine-5'-monophosphate), the active metabolite of the anticancer drug 5-FluoroUracil (frequently included to anticancer therapy).Previously, we described that ABCC11 high levels are associated to the estrogen receptor (ER) expression level in breast tumors and in cell lines resistant to tamoxifen. Consequently, by lowering the intracellular concentration of anticancer drugs, ABCC11 likely promotes a multidrug resistance (MDR) phenotype and decreases efficiency of anticancer therapy of 5FdUMP. Since no experimental data about binding sites of ABCC11 substrate are available, we decided to in silico localize putative substrate interaction sites of the nucleotide derivatives. Taking advantage of molecular dynamics simulation, we also analysed their evolution under computational physiological conditions and during the time. RESULTS: Since ABCC11 crystal structure is not resolved yet, we used the X-ray structures of the mouse mdr3 (homologous to human ABCB1) and of the bacterial homolog Sav1866 to generate two independent ABCC11 homology models in inward- and outward-facing conformations. Based on docking analyses, two putative binding pockets, for cGMP and 5FdUMP, were localized in both inward- and outward-facing conformations. Furthermore, based on our 3D models, and available biochemical data from homologous transporters, we identified several residues, potentially critical in ABCC11 transport function. Additionally, molecular dynamics simulation on our inward-facing model revealed for the first time conformation changes assumed to occur during transport process. CONCLUSIONS: ABCC11 would present two binding sites for cGMP and for 5FdUMP. Substrates likely first bind at the intracellular side of the transmembrane segment while ABCC11 is open forward the cytoplasm (inward-facing conformation). Then, along with conformational changes, it would pass through ABCC11 and fix the second site (close to the extracellular side), until the protein open itself to the extracellular space and allow substrate release.

ABCG2 transports and transfers heme to albumin through its large extracellular loop.

ABCG2 is an ATP-binding cassette (ABC) transporter preferentially expressed by immature human hematopoietic progenitors. Due to its role in drug resistance, its expression has been correlated with a protection role against protoporhyrin IX (PPIX) accumulation in stem cells under hypoxic conditions. We show here that zinc mesoporphyrin, a validated fluorescent heme analog, is transported by ABCG2. We also show that the ABCG2 large extracellular loop ECL3 constitutes a porphyrin-binding domain, which strongly interacts with heme, hemin, PPIX, ZnPPIX, CoPPIX, and much less efficiently with pheophorbide a, but not with vitamin B12. K(d) values are in the range 0.5-3.5 µm, with heme displaying the highest affinity. Nonporphyrin substrates of ABCG2, such as mitoxantrone, doxo/daunorubicin, and riboflavin, do not bind to ECL3. Single-point mutations H583A and C603A inside ECL3 prevent the binding of hemin but hardly affect that of iron-free PPIX. The extracellular location of ECL3 downstream from the transport sites suggests that, after membrane translocation, hemin is transferred to ECL3, which is strategically positioned to release the bound porphyrin to extracellular partners. We show here that human serum albumin could be one of these possible partners as it removes hemin bound to ECL3 and interacts with ABCG2, with a K(d) of about 3 µm.

The multidrug resistance half-transporter ABCG2 is purified as a tetramer upon selective extraction from membranes.

ABCG2 is a human membrane ATP-binding cassette half-transporter that hydrolyzes ATP to efflux a large number of chemotherapeutic agents. Several oligomeric states of ABCG2 from homodimers to dodecamers have been reported depending on the overexpression systems and/or the protocols used for purification. Here, we compared the oligomeric state of His(6)-ABCG2 expressed in Sf9 insect cells and in human Flp-In-293/ABCG2 cells after solubilization in mild detergents. His(6)-ABCG2 was purified through a new approach involving its specific recognition onto a functionalized lipid layer containing a Ni-NTA lipid. This approach allowed the purification of His-ABCG2 in presence of all solubilized membrane components that might be involved in the stabilisation of native oligomers and without requiring any additional washing or concentration passages. ABCG2 purified onto the NiNTA lipid surfaces were directly analyzed by electron microscopy and by biochemical assays. Altogether, our data are consistent with a tetrameric organization of ABCG2 when expressed in either heterologous Sf9 insect cells or in human homologous cells.

Uncompetitive nanomolar dimeric indenoindole inhibitors of the human breast cancer resistance pump ABCG2.

Multidrug resistance membrane pumps reduce the efficacy of chemotherapies by exporting a wide panel of structurally-divergent drugs. Here, to take advantage of the polyspecificity of the human Breast Cancer Resistance Protein (BCRP/ABCG2) and the dimeric nature of this pump, new dimeric indenoindole-based inhibitors from the monomeric α,ß-unsaturated ketone 4b and phenolic derivative 5a were designed. A library of 18 homo/hetero-dimers was synthesised. Homo-dimerization shifted the inhibition efficacy from sub-micromolar to nanomolar range, correlated with the presence of 5a, linked by a 2-6 methylene-long linker. Non-toxic, the best dimers displayed a therapeutic ratio as high as 70,000. It has been found that the high potency of the best compound 7b that displays a KI of 17 nM is due to an uncompetitive behavior toward mitoxantrone efflux and specific for that drug, compared to Hoechst 33342 efflux. Such property may be useful to target such anticancer drug efflux mediated by ABCG2. Finally, at a molecular level, an uncompetitive mechanism by which substrate promotes inhibitor binding implies that at least 2 ligands should bind simultaneously to the drug-binding pocket of ABCG2.

Iodination of verapamil for a stronger induction of death, through GSH efflux, of cancer cells overexpressing MRP1.

The multidrug resistance protein 1 (MRP1), involved in multidrug resistance (MDR) of cancer cells, was found to be modulated by verapamil, through stimulation of GSH transport, leading to apoptosis of MRP1-overexpressing cells. In this study, various iodinated derivatives of verapamil were synthesized, including iodination on the B ring, known to be involved in verapamil cardiotoxicity, and assayed for the stimulation of GSH efflux by MRP1. The iodination, for nearly all compounds, led to a higher stimulation of GSH efflux. However, determination of concomitant cytotoxicity is also important for selecting the best compound, which was found to be 10-fold more potent than verapamil. This will then allow us to design original anti-cancer compounds which could specifically kill the resistant cancer cells.

Synthesis of new piperazinyl-pyrrolo[1,2-a]quinoxaline derivatives as inhibitors of Candida albicans multidrug transporters by a Buchwald-Hartwig cross-coupling reaction.

Two series of piperazinyl-pyrrolo[1,2-a]quinoxaline derivatives were prepared via a Buchwald-Hartwig cross-coupling reaction and then evaluated for their ability to inhibit the drug efflux activity of CaCdr1p and CaMdr1p transporters of Candida albicans overexpressed in a Saccharomyces cerevisiae strain. In the initial screening of twenty-nine piperazinyl-pyrrolo[1,2-a]quinoxaline derivatives, twenty-three compounds behaved as dual inhibitors of CaCdr1p and CaMdr1p. Only four compounds showed exclusive inhibition of CaCdr1p or CaMdr1p. Further biological investigations were developed and for example, their antifungal potential was evaluated by measuring the growth of control yeast cells (AD1-8u-) and efflux pump-overexpressing cells (AD-CDR1 and AD-MDR1) after exposition to variable concentrations of the tested compounds. The MIC80 values of nineteen compounds ranging from 100 to 901 µM for AD-CDR1 demonstrated that relative resistance index (RI) values were between 8 and 274. In comparison, only seven compounds had RI values superior to 4 in cells overexpressing Mdr1p. These results indicated substrate behavior for nineteen compounds for CaCdr1p and seven compounds for CaMdr1p, as these compounds were transported via MDR transporter overexpressing cells and not by the AD1-8u- cells. Finally, in a combination assay with fluconazole, two compounds (1d and 1f) have shown a synergistic effect (fractional inhibitory concentration index (FICI) values ≤ 0.5) at micromolar concentrations in the AD-MDR1 yeast strain overexpressing CaMdr1p-protein, indicating an excellent potency toward chemosensitization.

Overexpression of homogeneous and active ABCG2 in insect cells.

The multidrug transporter ABCG2, a membrane protein with six transmembrane segments, can be overexpressed with the baculovirus/insect cell system. However, ABCG2 is produced as two species with different migration behavior via SDS-PAGE. Evidences suggest that this is due to the accumulation of an immature ABCG2 species, since: (i) the upper species, with higher apparent molecular weight, was favored by treatments reducing the rate of protein synthesis; (ii) the lower species was accumulated in presence of an endoplasmic reticulum stress inducer, and could be converted into the upper species during electrophoresis with 9 M urea; (iii) each species was differently solubilized by detergents: the upper species was partially solubilized by non-ionic and zwitterionic detergents, whereas the lower one required stronger surfactants; (iv) membrane ATPase activity from infected insect cells was essentially associated to the upper species. Altogether, these results suggest that although the insect cell/baculovirus system is not ideally adapted to overexpress human ABCG2, it is able to produce appreciable amounts of purified protein and the addition of agents reducing the rate of protein synthesis improves the homogeneity, making it a suitable heterologous expression system.

P-Glycoprotein 1 Affects Chemoactivities of Resveratrol against Human Colorectal Cancer Cells.

Resveratrol has been proposed to prevent tumor growth and the different steps of carcinogenesis; nevertheless, these biological effects are sometimes discordant between different cell types. Several hypotheses and works have suggested that the metabolism of resveratrol could be at the origin of a different cellular response. We show here, using colorectal tumor cell lines, that the biological effects of RSV result mainly from its carriage by carriers of the superfamily of ABC transporter, i.e., P-gP, MRP, or BCRP. Using cell lines overexpressing these different transporters, we have been able to highlight the importance of P-gP in the response of cells to RSV. These results were confirmed by invalidating the gene coding for P-gP, which restored the sensitivity of colorectal cells resistant to the polyphenol. Subsequently, the status of P-glycoprotein expression is an important element to be taken into consideration in the cytotoxic activity of resveratrol in colorectal cancer cells.

Dexamethasone down-regulates ABCG2 expression levels in breast cancer cells.

The breast cancer resistance protein ABCG2 effluxes a variety of drugs and is believed to play an important role in multidrug resistance to chemotherapy. We show here for the first time that dexamethasone (DEX) and progesterone (PROG) are able to strongly inhibit ABCG2 expression in progesterone receptor (PR)-positive MCF7 and PR-negative MDA-MB-231 breast cells. In contrast, in the latter cells stably-transfected with progesterone receptor isoforms A and B, ABCG2 expression was strongly up-regulated by DEX and PROG. In addition, two other ligands of Pregnane X Receptor (PXR) and/or Glucocorticoid Receptor (GR) were also able to down-regulate ABCG2 expression in PXR- and GR-positive MCF7 cells. ABCG2 expression regulation by DEX likely resulted from the activation of PR-, PXR-, and/or GR-signaling pathways. ABCG2 expression inhibition by DEX was associated with increased sensitivity to mitoxantrone, a known ABCG2 substrate. The findings suggest that DEX may be useful in improving drug efficacy under certain conditions.

Cytotoxicity of η6-areneruthenium-based molecules to glioblastoma cells and their recognition by multidrug ABC transporters.

A new series of amphiphilic η6-areneruthenium(II) compounds containing phenylazo ligands (group I: compounds 1a, 1b, 2a and 2b) and phenyloxadiazole ligands (group II: compounds 3a, 3b, 4a and 4b) were synthesized and characterized for their anti-glioblastoma activity. The effects of the amphiphilic η6-areneruthenium(II) complexes on the viability of three human glioblastoma cell lines, U251, U87MG and T98G, were evaluated. The azo-derivative ruthenium complexes (group I) showed high cytotoxicity to all cell lines, whilst most oxadiazole-derivative complexes (group II) were less cytotoxic, except for compound 4a. The cationic complexes 2a, 2b and 4b were more cytotoxic than the neutral complexes. Compounds 2a and 2b caused a significant reduction in the percentage of cells in the G0/G1 phase, with concomitant increases in the G2/M phase and fragmented DNA in the T98G cell line. The η6-areneruthenium(II) compounds were also tested in cell lines that overexpress the multidrug ABC transporters P-gp, MRP1 and ABCG2. Compounds 2b and 4a were substrates for the P-gp protein, with resistance indexes of 8.6 and 1.9, respectively. Compound 2b was also a substrate for ABCG2 and MRP1 proteins, with lower resistance indexes (1.8 and 1.6, respectively). The contribution of multidrug ABC transporters to the cytotoxicity of compound 2b in T98G cells was evidenced, since verapamil (a characteristic inhibitor of MRP1) increased the cytotoxicity of compound 2b at concentrations up to 20 µmol L-1, whilst GF120918 and Ko143 (specific inhibitors of P-gp and ABCG2, respectively) had no significant effect. In addition, we showed that compound 2b interacts with glutathione (GSH), which could explain its cellular efflux by MRP1. Our results showed that the amphiphilic η6-areneruthenium(II) complexes are promising anti-glioblastoma compounds, especially compound 2b, which was cytotoxic for all three cell lines, although it is transported by the three main multidrug ABC transporters.