Design and efficacy of bioactive drugs is restricted by their (in)ability to traverse cellular membranes. Therapy resistance, a major cause of ineffective cancer treatment, is frequently due to suboptimal intracellular accumulation of the drug. We report a molecular mechanism that promotes trans-membrane movement of a stereotypical, widely used anti-cancer agent to counteract resistance. Well-defined lipid analogues adapt to the amphiphilic drug doxorubicin, when co-inserted into the cell membrane, and assemble a transient channel that rapidly facilitates the translocation of the drug onto the intracellular membrane leaflet. Molecular dynamic simulations unveiled the structure and dynamics of membrane channel assembly. We demonstrate that this principle successfully addresses multi-drug resistance of genetically engineered mouse breast cancer models. Our results illuminate the role of the plasma membrane in restricting the efficacy of established therapies and drug resistance - and provide a mechanism to overcome ineffectiveness of existing and candidate drugs.
Poor accumulation of anti-cancer drugs in tumor cells is a major limitation in clinical cancer therapy. The main barrier for a drug to traverse through the body and reach its intracellular target is the plasma membrane. A typical (candidate) drug is therefore small, amphiphilic and of limited charge. Still, membrane traversal remains a highly inefficient and impeding process. In clinical oncology in particular there is an urgent need for new ways to improve drug efficacy and reduce toxicity. We have elucidated a mechanism that effectively accelerates membrane translocation of the stereotypical and widely used amphiphilic drug, doxorubicin. Well-defined short-chain sphingolipids, when co-inserted into the membrane, diminish the barrier for doxorubicin translocation. In vicinity of doxorubicin, the lipid analogues rapidly self-assemble at nanosecond timescale, and form a small, transient membrane channel. As a result, the doxorubicin drug readily translocates the membrane, thereby reducing the energetic barrier significantly by two-fold. Monte-Carlo based full-atom simulations revealed the structure and dynamics of channel formation at molecular detail. By means of a high-throughput screening approach of classical and targeted anti-cancer agent libraries, we identified various anti-cancer drugs in addition to doxorubicin, which share critical molecular characteristics, like defined amphiphilicity. Short-chain sphingolipids enhance cellular accumulation of these anti-cancer compounds similar to doxorubicin. Guided by these mechanistic insights, we applied the concept of facilitated doxorubicin traversal in genetically engineered WAPcre;EcadF/F;p53F/F mouse models for breast cancer. These tumors are multi-drug resistant to conventional and targeted anti-cancer agents. Co-administration of the sphingolipid analogue GC with liposomal doxorubicin effectively overcame drug resistance. While toxicity and normal tissue exposure reduced, GC caused elevated levels of intracellular doxorubicin in the tumor, improved tumor growth inhibition and significantly prolonged survival. Thus, a strategy of GC-mediated doxorubicin translocation was the only therapeutic approach that generated a sustained doxorubicin anti-tumor response. Notably, enhanced doxorubicin translocation was strongest over membranes of the tumor, a spectacular observation confirmed in vitro. We demonstrate that composition and local organization of the plasma membrane determine the efficiency of membrane channel formation. In conclusion, transient membrane channels target the tumor cell membrane to overcome multi-drug resistance. Our results illuminate a critical role of the (tumor) plasma membrane in restricting the efficacy of anti-cancer drugs and its contribution to multi-drug resistance. Moreover, our findings present a mechanism to address these limitations of (candidate) drugs in a clinically applicable way. Citation Format: Albert J. van Hell, Manuel Melo, Wim van Blitterswijk, Tim Dijkema, Lilia Pedrosa, Gerben Koning, Siewert Jan Marrink, Jos Jonkers, Marcel Verheij. Formation of transient membrane channels targets doxorubicin resistance. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3382. doi:10.1158/1538-7445.AM2013-3382
Edelfosine (ALP, Et-18-OCH3) is a prototype alkyl-lysophospholipid that has effective antitumoral activity. It inserts in the membrane and accumulates in lipid rafts leading to apoptosis in S49 mouse T cell lymphoma cells. A variant cell line, made resistant to ALP (S49AR) shows in impaired uptake of ALP. S49AR cells are not only resistant to a variety of ALP analogues, but also to DNA damage and Fas/CD95 death receptor induced apoptosis, suggesting another cause for resistance in addition to impaired ALP uptake. Here we studied the ALP resistance of S49AR. We observe an upregulation of phosho-PKB/Akt and phospho-ERK1/2 which are key proteins in the canonical survival signaling pathways. However inhibition of ERK and/or PKB/Akt using pharmacological inhibitors has no effect on the resistance of the S49AR cells to apoptotic stimuli. PKB/Akt and ERK1/2 activation is regulated by PI(3,4,5)P3 formation. The levels of phosphoinositides in the various cell lines were measured with HPLC. The cell lines resistant to apoptotic inducing agents show decreased levels of the diverse phosphopoinositides. SHIP-1 is a SH2 domain containing inositol phosphatase that removes the 5′ phosphate of PI(3,4,5)P3 and is mainly expressed in hemotopoietic cells. It decreases the pool of PI(3,4,5)P3 thereby negatively regulating the activation of PKB/Akt. We see down regulation of SHIP-1 both on gene expression and protein level in the S49AR compared to S49 cells. Other components of the phosphoinositide metabolism, like PI3Kinase and SHIP2, did not show any changes. Resensitization of the S49AR cells to ALP leads to restoration of SHIP-1 expression. Knocking down SHIP-1 using siRNA causes resistance to various ALPs, DNA damage and Fas/CD95 death receptor induced apoptosis. Microarray analysis show that the S49AR and S49siSHIP cells show remarkably similar expression profile which are distinct from the expression profiles shown by S49 and S49mock cells. Concluding, SHIP-1 is downregulated in ALP resistant T cell lymphoma cells concurrent with a general downregulation of phosphoinositide levels in the cells which is associated with the resistance to ALP induced apoptosis. How ALP resistance is caused is yet unknown and is the scope of current research. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4025.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.