title: Short-chain glycoceramides promote intracellular mitoxantrone delivery from novel nanoliposomes into breast cancer cells.
abstract: OBJECTIVE
  To improve therapeutic activity of mitoxantrone (MTO)-based chemotherapy by reducing toxicity through encapsulation in nanoliposomes and enhancing intracellular drug delivery using short-chain sphingolipid (SCS) mediated tumor cell membrane permeabilization.
  
  
  METHODS
  Standard (MTOL) and nanoliposomes enriched with the SCS, C8-Glucosylceramide or C8-Galactosylceramide (SCS-MTOL) were loaded by a transmembrane ammonium sulphate gradient and characterized by DLS and cryo-TEM. Intracellular MTO delivery was measured by flow cytometry and imaged by fluorescence microscopy. In vitro cytotoxicity was studied in breast carcinoma cell lines. Additionally, live cell confocal microscopy addressed the drug delivery mechanism by following the intracellular fate of the nanoliposomes, the SCS and MTO. Intratumoral MTO localization in relation to CD31-positive tumor vessels and CD11b positive cells was studied in an orthotopic MCF-7 breast cancer xenograft.
  
  
  RESULTS
  Stable SCS-MTOL were developed increasing MTO delivery and cytotoxicity to tumor cells compared to standard MTOL. This effect was much less pronounced in normal cells. The drug delivery mechanism involved a transfer of SCS to the cell membrane, independently of drug transfer and not involving nanoliposome internalization. MTO was detected intratumorally upon MTOL and SCS-MTOL treatment, but not after free MTO, suggesting an important improvement in tumor drug delivery by nanoliposomal formulation. Nanoliposomal MTO delivery and cellular uptake was heterogeneous throughout the tumor and clearly correlated with CD31-positive tumor vessels. Yet, MTO uptake by CD11b positive cells in tumor stroma was minor.
  
  
  CONCLUSIONS
  Nanoliposomal encapsulation improves intratumoral MTO delivery over free drug. Liposome bilayer-incorporated SCS preferentially permeabilize tumor cell membranes enhancing intracellular MTO delivery.
authors.affiliation: Section Surgical Oncology, Department of Surgery, Laboratory Experimental Surgical Oncology, Room Ee151b, Erasmus MC, 3000CA, Rotterdam, PO Box 2040, The Netherlands.
authors.forename: Lília R Cordeiro
authors.initials: LR
authors.lastname: Pedrosa
authors.forename: Timo L M
authors.initials: TL
authors.lastname: Ten Hagen
authors.forename: Regine
authors.initials: R
authors.lastname: Süss
authors.forename: Albert
authors.initials: A
authors.lastname: van Hell
authors.forename: Alexander M M
authors.initials: AM
authors.lastname: Eggermont
authors.forename: Marcel
authors.initials: M
authors.lastname: Verheij
authors.forename: Gerben A
authors.initials: GA
authors.lastname: Koning
mesh_terms: D000970:Antineoplastic Agents / Q000008:administration & dosage* / Q000493:pharmacokinetics / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D004355:Drug Stability; D004356:Drug Storage; D005699:Galactosylceramides / Q000737:chemistry*; D005963:Glucosylceramides / Q000737:chemistry*; D006801:Humans; D008081:Liposomes; D061986:MCF-7 Cells; D008942:Mitoxantrone / Q000008:administration & dosage* / Q000493:pharmacokinetics / Q000494:pharmacology; D053758:Nanoparticles / Q000737:chemistry*
chemical_list: D000970:Antineoplastic Agents; D005699:Galactosylceramides; D005963:Glucosylceramides; D008081:Liposomes; D008942:Mitoxantrone