Search

The multidrug resistance-associated protein transporter ABCC1 (MRP1) is an integral plasma membrane protein involved in the multidrug resistance phenotype. It actively expels a number of cytotoxic molecules from cells. To gain insight into the modulation of the functional properties of this integral membrane protein by cholesterol, a main component of the lipid bilayer, we used multidrug-resistant GLC4/ADR cells, which overexpress MRP1. Upon altering the plasma membrane cholesterol content of these cells, membrane localization and the activity of MRP1 were analyzed. A detergent-free methodology was used to separate ‘light’ and ‘heavy’ plasma membrane fractions. Our data show that MRP1 was exclusively found in ‘light’ fractions known as L0 phase membrane microdomains, together with ∼ 23% of gangliosides GM1 and 40% of caveolin-1. Depletion of the membrane cholesterol level to 40% by treatment with the cholesterol-chelating agent methyl-β-cyclodextrin did not modify MRP1 activity, as evidenced either by the rate of efflux of pirarubicin or that of glutathione. Further cholesterol depletion below 40% yielded both a partial shift of MRP1 to the high-density fraction and a decrease of its functionality. Taken together, these data suggest that MRP1 funtionality depends on its localization in cholesterol-rich membrane microdomains. [ABSTRACT FROM AUTHOR]

Multidrug resistance (MDR) in model systems is known to be conferred by two different integral proteins, the 170-kDa P-glycoprotein (P-gp) and the 190-kDa multidrug resistance-associated protein (MRP1), both of which pump drugs out of MDR cells. The presence of a nitrogen atom, charged at physiological pH, has frequently been considered to be a hallmark of P-gp substrates and inhibitors. The present study was aimed at investigating the role of nitrogen in the ability of the pump to recognise substrate. We measured the kinetics of active efflux of seven new anthracycline derivatives in P-gp-expressing K562/ADR cells and in MRP1-expressing GLC4/ADR cells. Six of these compounds represent analogues of daunorubicin in which the amino sugar nitrogen is bound to an amino- or a nitro-substituted benzyl moiety, the seventh is a doxorubicin derivative in which benzyl group is bound with 4′-oxygen. We found that the compounds with a nitro group on the benzyl ring were poor substrates for P-gp despite the presence of a secondary amine that can be protonated. In contrast, compounds that have a free amino group were very good substrates even though this amine is not protonated in the pH range studied (pK∼3). These results show that the nitrogen atom does not interact with P-gp in a charged form but rather as an electron donating group. [Copyright &y& Elsevier]

Abstract In this study, a new series of N,N - bis(alkanol)amine aryl ester heterodimers was synthesized and studied. The new compounds were designed based on the structures of our previous arylamine ester derivatives endowed with high P-gp-dependent multidrug resistance reversing activity on a multidrug-resistant leukemia cell line. All new compounds were active in the pirarubicin uptake assay on the doxorubicin–resistant erythroleukemia K562 cells (K562/DOX). Compounds bearing a linker made up of 10 methylenes showed unprecedented high reversal activities regardless of the combination of aromatic moieties. Docking results obtained by an in silico study supported the data obtained by the biological tests and a study devoted to establish the chemical stability in phosphate buffer solution (PBS) and human plasma showed that only a few compounds exhibited a significant degradation in the human plasma matrix. Ten selected non-hydrolysable derivatives were able to inhibit the P-gp-mediated rhodamine-123 efflux on K562/DOX cells, and the evaluation of their apparent permeability and ATP consumption on other cell lines suggested that the compounds can behave as unambiguous or not transported substrates. The activity of these the compounds on the transport proteins breast cancer resistance protein (BCRP) and multidrug resistance associated protein 1 (MRP1) was also analyzed. All tested derivatives displayed a moderate potency on the BCRP overexpressing cells; while only four molecules showed to be effective on MRP1 overexpressing cells, highlighting a clear structural requirement for selectivity. In conclusion, we have identified a new very powerful series of compounds which represent interesting leads for the development of new potent and efficacious P-gp-dependent MDR modulators. Graphical abstract Image 1 Highlights • A new series of N,N -bis(alkanol)amine aryl esters was designed and synthesized. • The compounds were tested on K562/DOX cells in the pirarubicin uptake assay. • In silico studies and stability tests in PBS and human plasma were also performed. • Further experiments confirmed that they are P-gp-dependent MDR reversers. • A different selectivity vs sister proteins MRP1 and BCRP was highlighted. [ABSTRACT FROM AUTHOR]

In this study, a new series of heterodimers was synthesized. These derivatives are N , N - bis(alkanol)amine aryl esters or N , N - bis(ethoxyethanol)amine aryl esters carrying a methoxylated aryl residue combined with a flavone or chromone moiety. The new compounds were studied to evaluate their P-gp modulating activity on a multidrug-resistant leukemia cell line. Some of the new compounds show a good MDR reversing activity; interestingly this new series of compounds does not comply with the structure-activity relationships (SAR) outlined by previously synthesized analogs carrying different aromatic moieties. In the case of the compounds described in this paper, activity is linked to different features, in particular the characteristics of the spacer, which seem to be critical for the interaction with the pump. This fact indicates that the presence of a flavone or chromone residue influences the SAR of these series of products, and that flexible molecules can find different productive binding modes with the P-gp recognition site. These results support the synthesis of new compounds that might be useful leads for the development of drugs to control P-gp-dependent MDR. [ABSTRACT FROM AUTHOR]

In a continuing search for potent P-gp-dependent multidrug-resistant (MDR) reversers we synthesized and studied a new series of N -alkanol- N -cyclohexanol amine aryl esters characterized by the presence of two linkers with different flexibility: a polymethylene chain of variable length and a cyclohexylic scaffold, that gave origin to two geometrical isomers ( cis and trans ). The reversal activity of the new compounds was evaluated on the K562/DOX cell line by three tests: pirarubicin uptake modulation, doxorubicin cytotoxicity enhancement (reversal fold, RF) and inhibition of P-gp-mediated rhodamine-123 (Rhd 123) efflux tests. The chemical stability of their ester function was evaluated in the experimental conditions utilized (phosphate buffer solution (PBS), bovine serum and in the presence of K562/DOX cells) and in human plasma. The new series of molecules showed very interesting MDR reversing properties; in particular compound 5b (ELF26B), characterized by trans stereochemistry and a 5-methylene chain, presented the best pharmacological profile and is stable in each tested medium. Compound 5b could be an interesting lead for the development of new potent and efficacious P-gp-dependent MDR modulators. [ABSTRACT FROM AUTHOR]

As a continuation of our research on potent and efficacious P-gp-dependent multidrug resistance (MDR) reversers, several new N , N -bis(alkanol)amine aryl esters were designed and synthesized, varying the aromatic moieties or the length of the methylenic chain. The new compounds were tested on doxorubicin-resistant erythroleukemia K562 cells (K562/DOX) in the pirarubicin uptake assay, where most of the new compounds were shown to be active. In particular the asymmetrical compounds, characterized by two linkers of different length, generally showed fairly high activities as MDR reversers. Some selected compounds (isomers 15 – 17 ) were further studied by evaluating their doxorubicin cytotoxicity enhancement (reversal fold, RF) on the K562/DOX cell line. The results of both pharmacological assays indicate that compounds 16 (GDE6) and 17 (GDE19) could be interesting leads for the development of new P-gp dependent MDR modulators. [ABSTRACT FROM AUTHOR]

Abstract: Conformational modulation of the aryl portion of a set of N,N-bis(cyclohexanol)amine aryl esters (1a–d) that are potent Pgp-dependent MDR inhibitors has been performed. Toward this end the trans-3-(3,4,5-trimethoxyphenyl)acrylic acid present in set 1 was substituted with 3-(3,4,5-trimethoxyphenyl)propanoic and 3-(3,4,5-trimethoxyphenyl)propiolic moieties to give sets 2 and 3, respectively. While the introduction of 3-(3,4,5-trimethoxyphenyl)propanoic moiety resulted in a definite drop in potency and efficacy, esterification with 3-(3,4,5-trimethoxyphenyl)propiolic acid gave four isomers (3a–d) that maintain high potency and possess optimal efficacy. These results are discussed in terms of conformational flexibility of the different sets of compounds. [Copyright &y& Elsevier]

Stereo- and regioisomers of a series of N,N -bis(alkanol)amine aryl ester derivatives have been prepared and studied as multidrug resistance (MDR) modulators. The new compounds contain a 2-(methyl)propyl chain combined with a 3-, 5- or 7-methylenes long chain and carry different aromatic ester portions. Thus, these compounds have a methyl group on the 3-methylenes chain and represent branched homologues of previously studied derivatives. The introduction of the methyl group gives origin to a stereogenic center and consequently to (R) and (S) enantiomers. In the pirarubicin uptake assay on K562/DOX cell line these compounds showed good activity and efficacy and in many cases enantioselectivity was observed. Docking studies confirmed the influence of the stereocenter on the interaction in the P-gp pocket. The P-gp interaction mechanism and selectivity towards MRP1 and BCRP were also evaluated on MDCK transfected cells overexpressing the three transporters. Almost all these compounds inhibited both P-gp and BCRP, but only derivatives with specific structural characteristics showed MRP1 activity. Moreover, two compounds, (S)- 3 and (R)- 7 , showed the ability to induce collateral sensitivity (CS) against MDR cells. Therefore, these two CS-promoting agents could be considered interesting leads for the development of selective cytotoxic agents for drug-resistant cells. •Stereo- and regioisomers of amino aryl esters showed different MDR reversing activity profiles.•Compounds (S)- 3 (Ar = b, Ar1 = a, n = 3) and (R)- 7 (Ar = b, Ar1 = a, n = 5) induced collateral sensitivity (CS) against MDR cells. Image 1 • Stereo- and regioisomers of N,N -bis(alkanol)amine aryl esters were described. • The biological behavior of these compounds was studied by various tests. • They act as multidrug resistance (MDR) reversers. • Two compounds were identified as MDR1 or MRP1-overexpressing cells sensitizers. • Docking studies and stability tests in human plasma were performed. [ABSTRACT FROM AUTHOR]