Table 2 - uploaded by Ian Magrath
Content may be subject to copyright.
Whole-cellDPH fluorescence polarization P value of P388 and P388/ADRcells For experimental details, see "Materials and Methods."
Source publication
We have studied the structural order of the lipid phase of plasma membranes from P388 murine leukemia cells and from a Doxorubicin-resistant subline, P388/ADR, using electron spin resonance spectroscopy and fluorescence depolarization measurements. Measurements of the order parameter, S, following incubation of cells from both lines with the N-oxyl...
Citations
... Therefore, the transfer of this drug through lipid membranes is a wellstudied topic [37,[42][43][44]. Doxorubicin-membrane interactions were also proposed as an important factor contributing to drug resistance mechanisms [45][46][47]. Doxorubicin has a strong affinity towards cell membranes [48]. This affinity is especially pronounced for membranes containing anionic lipids, as can be expected for the protonated form of the drug. ...
Doxorubicin (DOX) is a commonly used chemotherapeutic drug, from the anthracycline class, which is genotoxic to neoplastic cells via a DNA intercalation mechanism. It is effective and universal; however, it also causes numerous side effects. The most serious of them are cardiotoxicity and a decrease in the number of myeloid cells. For this reason, targeted DOX delivery systems are desirable, since they would allow lowering the drug dose and therefore limiting systemic side effects. Recently, synthetic dyes, in particular Congo red (CR), have been proposed as possible DOX carriers. CR is a planar molecule, built of a central biphenyl moiety and two substituted naphthalene rings, connected with diazo bonds. In water, it forms elongated ribbon-shaped supramolecular structures, which are able to selectively interact with immune complexes. In our previous studies, we have shown that CR aggregates can intercalate DOX molecules. In this way, they preclude DOX precipitation in water solutions and increase its uptake by MCF7 breast cancer cells. In the present work, we further explore the interactions between DOX, CR, and their aggregates (CR/DOX) with phospholipid membranes. In addition to neutral molecules, the protonated doxorubicin form, DXP, is also studied. Molecular dynamics simulations are employed to study the transfer of CR, DOX, DXP, and their aggregates through POPC bilayers. Interactions of CR, DOX, and CR/DOX with model monolayers are studied with Langmuir trough measurements. This study shows that CR may support the transfer of doxorubicin molecules into the bilayer. Both electrostatic and van der Waals interactions with lipids are important in this respect. The former promote the initial stages of the insertion process, the latter keep guest molecules inside the bilayer.
... Modulators, such as verapamil and cyclosporin A, have been identified as chemosensitizers that can reverse drug resistance in MDR cells in vitro. In comparison with mouse leukemia cells (P388), drug-resistant cells (P388/ADR) had reduced PC content and increased SM content, and electron spin resonance spectroscopy showed that the lipid structure of drug-resistant cells was more ordered (Xiang et al., 2020;Ramu et al., 1984;Ramu et al., 1983). Kok et al., 2000 found that in HT29 cells overexpressing MRP1, glucosylceramide and galactosylceramide were increased in the composition ratio of sphingolipids. ...
There have been significant advances in our understanding of how changes in the fluidity and permeability of the cell membrane can affect drug resistance in cancer. Research has shown that cancer cells often have changes in the fluidity and permeability of their cell membrane that contribute to their resistance to drugs used to treat cancer. These changes may be due to changes in the composition and organization of the lipid bilayer that makes up the membrane, as well as changes in the expression or localization of proteins and other molecules embedded in the membrane. The lipid composition in the tumor cell membrane changes with drug resistance, which can affect the fluidity and permeability of the cell membrane. Reversal of drug resistance can be achieved by altering cell membrane fluidity and permeability. In recent years, there have been numerous studies aimed at understanding the mechanisms underlying these changes and identifying strategies to overcome drug resistance in cancer. This research has led to the development of new drugs and drug delivery systems that are designed to target specific changes in the cell membrane of cancer cells and improve the effectiveness of chemotherapy. Overall, the advances in our understanding of the role of cell membrane fluidity and permeability in drug resistance in cancer have led to the development of new approaches to treat cancer and improve patient outcomes and further research is needed to continue to improve the understanding of these mechanisms and to identify new strategies to overcome drug resistance in cancer. This article highlights the research status and detection methods of cell membrane fluidity and permeability affecting tumor drug resistance.
... Increase in cholesterol concentration results in increased hydrogen bonding between headgroups of lipids and cholesterol [86] and subsequently to a tighter and thicker membrane with a reduced number of empty spaces, thus creating a greater distance and a greater barrier in the membrane for the influx of drugs [12,26,85]. MDR breast [65], leukemic [4] and ovarian [66] cancers have all been found to contain significantly greater cholesterol content, and higher membrane lipid order [87] than their drug-sensitive counterparts. Peetla et al. suggested that this cholesterol-driven increase in membrane rigidity results in the chemotherapy drug being trapped in the membrane [65] (Fig. 1) and Rivel et al. even provided evidence that membrane permeability to cisplatin was proportional to cholesterol concentration [88,89]. ...
The lipid bilayer is a functional component of cells, forming a stable platform for the initiation of key biological processes, including cell signalling. There are distinct changes in the lipid composition of cell membranes during oncogenic transformation resulting in aberrant activation and inactivation of signalling transduction pathways. Studying the role of the cell membrane in cell signalling is challenging, since techniques are often limited to by timescale, resolution, sensitivity, and averaging. To overcome these limitations, combining ‘computational’, ‘wet-lab’ and ‘semi-dry’ approaches offers the best opportunity to resolving complex biological processes involved in membrane organisation. In this review, we highlight analytical tools that have been applied for the study of cell signalling initiation from the cancer cell membranes through computational microscopy, biological assays, and membrane biophysics. The cancer therapeutic potential of extracellular membrane-modulating agents, such as cholesterol-reducing agents is also discussed, as is the need for future collaborative inter-disciplinary research for studying the role of the cell membrane and its components in cancer therapy.
... Considerable efforts have been directed to elucidate the differences in doxorubicin delivery to resistant/sensitive cancer cells. Since the early 1980s, these studies have received great attention in the context of multidrug resistance problem [4][5][6][7]. As demonstrated in [4,5], cell's susceptibility to doxorubicin depends strongly on biophysical properties of membrane (lipid packing, membrane fluidity, electric charge) and, ultimately, on doxorubicinlipid interaction. ...
The effect of anticancer antibiotic doxorubicin on structural organization of anionic lipid monolayers has been studied. X-ray reflectivity and grazing incidence diffraction techniques were applied to monitor the changes in 2D structure and electron density distribution of Langmuir monolayer composed of negatively charged dipalmitoylphosphatidylglycerol (DPPG) and dioleoylphosphatidylserine (DOPS). For comparison, monolayer of zwitterionic dipalmitoylphosphatidylethanolamine (DPPE) also was investigated. The presented experimental results suggest that doxorubicin interaction with anionic lipid monolayers (DPPG and DOPS) proceeds preferentially via electrostatic attraction—positively charged amino groups of doxorubicin bind to negatively charged head groups of phospholipid molecules. Based on the obtained data, the penetration of doxorubicin into the hydrophobic part of anionic lipid monolayers does not occur. X-ray measurements on DPPE monolayer indicated that doxorubicin did not cause any significant alterations of molecular packing in condensed monolayer of zwitterionic DPPE molecules.
... Cells that are resistant to the drug doxorubicin have been reported to have a higher degree of structural order in the PM [118] and to have higher levels of SM and CHOL [119] than the corresponding doxorubicin-sensitive cells. Urothelial cancer cells contained a CHOL level that correlated with the cancerous transformation, and it was proposed that the CHOL/SM-rich membrane domains in these cancer cells should constitute a selective therapeutic target for elimination of these cells [120]. ...
Several studies have demonstrated interactions between the two leaflets in membrane bilayers and the importance of specific lipid species for such interaction and membrane function. We here discuss these investigations with a focus on the sphingolipid and cholesterol-rich lipid membrane domains called lipid rafts, including the small flask-shaped invaginations called caveolae, and the importance of such membrane structures in cell biology and cancer. We discuss the possible interactions between the very long-chain sphingolipids in the outer leaflet of the plasma membrane and the phosphatidylserine species PS 18:0/18:1 in the inner leaflet and the importance of cholesterol for such interactions. We challenge the view that lipid rafts contain a large fraction of lipids with two saturated fatty acyl groups and argue that it is important in future studies of membrane models to use asymmetric membrane bilayers with lipid species commonly found in cellular membranes. We also discuss the need for more quantitative lipidomic studies in order to understand membrane function and structure in general, and the importance of lipid rafts in biological systems. Finally, we discuss cancer-related changes in lipid rafts and lipid composition, with a special focus on changes in glycosphingolipids and the possibility of using lipid therapy for cancer treatment.
... Also, in general a decrease in drug influx correlates to the membrane biophysical properties. Doxorubicin-resistant P338 cell line had a decreased PC/SM ratio and an increase in the membrane order [1,43]; Vinblastine-resistant leukemia T cells showed an increase of up to 60% in protein/lipid content compared to drug-sensitive cells [44]. Peetla et al. reported that lipids isolated from doxorubicin-resistant MCF-7 breast cancer cells (MCF-7/ADR) had higher concentrations of SM, PI, and cholesterol than sensitive cells and that membrane lipids are more rigid than sensitive cells, showing also that doxorubicin had strong hydrophobic interactions with resistant cell membrane lipids when compared to lower and ionic interactions that established with membrane lipids from sensitive cells [45]. ...
Cancer is a multi-process disease where different mechanisms exist in parallel to ensure cell survival and constant adaptation to the extracellular environment. To adapt rapidly, cancer cells re-arrange their plasma membranes to sustain proliferation, avoid apoptosis and resist anticancer drugs. In this review, we discuss novel approaches based on the modifications and manipulations that new classes of molecules can exert in the plasma membrane lateral organization and order of cancer cells, affecting growth factor signaling, invasiveness, and drug resistance. Furthermore, we present azurin, an anticancer protein from bacterial origin, as a new approach in the development of therapeutic strategies that target the cell membrane to improve the existing standard therapies.
... Interestingly, there seems to be a consensus that the membranes of resistant cells show an abundance of sphingolipids, Chol and Chol esters. This has been reported in case of doxorubicin-resistant (MCF-7/ADR) breast cancer cells [9], doxorubicin-resistant P388/ADR murine leukemia cells [10], vinblastine resistant leukemic T-lymphoblasts [11] and others [12]. In all these cases, drug resistance was in part attributed to a decrease in membrane fluidity and hence a lower ability of the lipophilic drugs to diffuse across the membrane [12]. ...
... On the contrary, MDR reversal by the nonionic surfactants tocopheryl polyethylene glycol succinate (TPGS) and alpha-Tocopherol was attributed to an increased membrane rigidity [116,117]. The MDR reversing potentials of nonionic surfactants is a function of their hydrophilic-lipophilic balance (HLB), and an intermediate HLB value (10)(11)(12)(13)(14)(15)(16)(17) has been proposed for their optimal MDR reversing effect [118]. ...
Multidrug resistance (MDR) is associated with a wide range of pathological changes at different cellular and intracellular levels. Nanoparticles (NPs) have been extensively exploited as the carriers of MDR reversing payloads to resistant tumor cells. However, when properly formulated in terms of chemical composition and physicochemical properties, NPs can serve as beyond delivery systems and help overcome MDR even without carrying a load of chemosensitizers or MDR reversing molecular cargos. Whether serving as drug carriers or beyond, a wise design of the nanoparticulate systems to overcome the cellular and intracellular alterations underlying the resistance is imperative. Within the current review, we will initially discuss the cellular changes occurring in resistant cells and how such changes lead to chemotherapy failure and cancer cell survival. We will then focus on different mechanisms through which nanosystems with appropriate chemical composition and physicochemical properties can serve as MDR reversing units at different cellular and intracellular levels according to the changes that underlie the resistance. Finally, we will conclude by discussing logical grounds for a wise and rational design of MDR reversing nanoparticulate systems to improve the cancer therapeutic approaches.
... Changes in lipid composition and the biophysical properties of membrane lipids have been implicated in drug resistance in several cell lines. [8][9][10][11][12] We recently reviewed the role of membrane lipids in cancer progression and drug resistance. 13 In our previous studies, we demonstrated that composition and biophysical characteristics of membrane lipids of drug-resistant breast cancer cells (MCF-7/ADR) are significantly different than that of drug-sensitive cells (MCF-7). ...
Cell-membrane lipid composition can greatly influence biophysical properties of cell membranes, affecting various cellular functions. We previously showed that lipid synthesis becomes altered in the membranes of resistant breast cancer cells (MCF-7/ADR); they form a more rigid, hydrophobic lipid monolayer than do sensitive cell membranes (MCF-7). These changes in membrane lipids of resistant cells, attributed to epigenetic aberration, significantly affected drug transport and endocytic function, thus impacting the efficacy of anticancer drugs. The present study's objective was to determine the effects of the epigenetic drug 5-aza-2'-deoxycytidine (DAC), delivered in sustained-release nanogels (DAC-NGs), on the composition and biophysical properties of membrane lipids of resistant cells. Resistant and sensitive cells were treated with DAC in solution (DAC-sol) or DAC-NGs, and cell-membrane lipids were isolated and analyzed for lipid composition and biophysical properties. In resistant cells, we found increased formation of Cholesterol-Sphingomyelin (CHOL-SM) rafts with culturing time, whereas DAC treatment reduced their formation. In general, the effect of DAC-NGs was greater in changing the lipid composition than with DAC-sol. DAC treatment also caused a rise in levels of certain phospholipids and neutral lipids known to increase membrane fluidity while reducing the levels of certain lipids known to increase membrane rigidity. Isotherm data showed increased lipid membrane fluidity following DAC treatment, attributed to decrease levels of CHOL-SM rafts (lamellar beta [Lβ] structures or ordered gel) and a corresponding increase in lipids that form lamellar alpha structures (Lα, liquid crystalline phase). Sensitive cells showed marginal or insignificant changes in lipid profile following DAC-treatment, suggesting that epigenetic changes affecting lipid biosynthesis are more specific to resistant cells. Since membrane fluidity plays a major role in drug transport and endocytic function, treatment of resistant cells with epigenetic drugs with altered lipid profile could facilitate anticancer drug transport to overcome acquired drug resistance in a combination therapy.
... Several studies have reported lower fluidity and higher heterogeneity of plasma membrane in MDR cells compared to sensitive cells. 86,87 Drug-resistant cells also contain smaller amounts of unsaturated fatty acids and have higher content of esterified cholesterol and triglycerides. 88,89 Using liposomes of different lipid composition and viscosity it was demonstrated that the L61 effects on lipid flip-flop and membrane permeability toward Dox increase as the membrane viscosity increases. ...
Multidrug resistance (MDR) remains one of the biggest obstacles for effective cancer therapy. Currently there is only few methods that are available clinically that are used to bypass MDR with very limited success. In this review we describe how MDR can be overcome by simple yet effective approach of using amphiphilic block copolymers. Tri-block copolymers of poly(ethylene oxide) (PEO) and poly(propylene oxide) (PPO), arranged in a tri-block structure PEO-PPO-PEO, Pluronics or "poloxamers", raised a considerable interest in drug delivery field. Previous studies demonstrated that Pluronics sensitize MDR cancer cells resulting in increased cytotoxic activity of Dox, paclitaxel, and other drugs by 2-3 orders of magnitude. Pluronics can also prevent the development of MDR in vitro and in vivo. Additionally, promising results of clinical studies of Dox/Pluronic formulation reinforced the need to ascertain a thorough understanding of Pluronic effects in tumors. These effects are extremely comprehensive and appear on the level of plasma membranes, mitochondria, and regulation of gene expression selectively in MDR cancer cells. Moreover, it has been demonstrated recently that Pluronics can effectively deplete tumorigenic intrinsically drug resistant cancer stem cells (CSC). Interestingly, sensitization of MDR and inhibition of drug efflux transporters is not specific or selective to Pluronics. Other amphiphilic polymers have shown similar activities in various experimental models. This review summarizes recent advances of understanding the Pluronic effects in sensitization and prevention of MDR.
... Interestingly the two most efficient extractions were from drug selected cancer cell lines (H69AR and MCF7-FLV) which overexpress transporters due to development of drug resistance. It is well known that such drug resistant cell lines display altered membrane constituents and properties [36][37][38], and it seems that this improves the ability of SMA to solubilise these transporters. ...
ABC (ATP Binding Cassette) transporters carry out many vital functions and are involved in numerous diseases, but study of the structure and function of these proteins is often hampered by their large size and membrane location. Membrane protein purification usually utilises detergents to solubilise the protein from the membrane, effectively removing it from its native lipid environment. Subsequently lipids have to be added back and detergent removed to reconstitute the protein into a lipid bilayer. We present here the application of a new methodology for the extraction and purification of ABC transporters without the use of detergent, instead using a styrene maleic acid co-polymer (SMA). SMA inserts in a bilayer and assembles into discrete particles, essentially solubilising the membrane into small discs of bilayer encircled by polymer, termed SMA lipid particles (SMALPs). We show that this polymer can extract several eukaryotic ABC transporters; P-glycoprotein (ABCB1), MRP1 (ABCC1), MRP4 (ABCC4), ABCG2 and CFTR (ABCC7), from a range of different expression systems. The SMALP encapsulated ABC transporters can be purified by affinity chromatography, and are able to bind ligands comparably to those in native membranes or detergent micelles. A greater degree of purity and enhanced stability is seen compared to detergent solubilisation. This study demonstrates that eukaryotic ABC transporters can be extracted and purified without ever being removed from their lipid bilayer environment, opening up a wide range of possibilities for the future study of their structure and function.
