1615 JPP 2003, 55: 1615–1622 ß 2003 The Authors Received May 1, 2003 Accepted July 22, 2003 DOI 10.1211/0022357022232 ISSN 0022-3573 Lipid Metabolism Laboratory, the Heart Institute (InCor) of the Medical School Hospital, Sa ˜o Paulo, Brazil Claudete J. Valduga, Denise C. Fernandes, Debora G. Rodrigues, Raul C. Maranha ˜o Faculty of Pharmaceutical Sciences, University of Sa ˜ o Paulo, Sa ˜ o Paulo, Brazil Ana C. Lo Prete, Carolina H. M. Azevedo, Debora G. Rodrigues, Raul C. Maranha ˜o Correspondence: R. C. Maranha ˜ o, Instituto do Corac ¸a ˜ o (InCor) do Hospital das Clõ ´nicas da FMUSP, Laborato ´ rio de Metabolismo de Lõ ´pides, Av. Dr. Ene ´ as de Carvalho Aguiar, 44, Anexo II, 1 SS. Sa ˜ o Paulo–SP, 05403-000, Brazil. E-mail: ramarans@usp.br Acknowledgement and funding: The authors are grateful to Debora F. Deus, Douglas M. Miyamoto and Ricardo D. Couto for help with the experiments. This study was supported by FAPESP (Grant no. 99/01229-2). Use of a cholesterol-rich microemulsion that binds to low-density lipoprotein receptors as vehicle for etoposide Claudete J. Valduga, Denise C. Fernandes, Ana C. Lo Prete, Carolina H. M. Azevedo, Debora G. Rodrigues and Raul C. Maranha ˜o Abstract A cholesterol-rich microemulsion (LDE) that binds to low-density lipoprotein (LDL) receptors is selec- tively taken up by malignant cells that overexpress those receptors and may be used as vehicle for antineoplastic agents. This study aimed to develop the association of etoposide with LDE. It was firstly observed that etoposide poorly associates with the microemulsion, therefore the experiments were performed with a lipophilic fatty acid derivative of the drug. The association of etoposide oleate with LDE was almost 100% and was tested for physical and chemical stability, as well as for cellular uptake, toxicity in mice and cytotoxic activity against a neoplastic cell line (NCI-H292). Uptake and cytotoxic activity of LDE-etoposide oleate by NCI-H292 cells was mediated by LDL receptors. The anti-prolifera- tive activity of LDE-etoposide oleate against the neoplastic cells was smaller than that of etoposide oleate (IC50 (drug concentration required to inhibit 50% of the cell growth) ˆ 0.48 and 0.19 mM, respectively). This difference, however, can be ascribed to the activity of the commercially used vehicle and not the drug itself because when this vehicle was added to the cultures with LDE-etoposide oleate, the IC50 decreased. On the other hand, the tolerability of LDE-etoposide oleate to mice was remark- able, such that its lethal dose (LD50) was about five-fold that of the commercial formulation (LD50 ˆ 315 and 58 mg kg ¡1 , respectively). In conclusion, LDE-etoposide oleate association is stable and the cytostatic activity of the drug is preserved while its toxicity to animals is small. By diminishing the side effects and directing etoposide to neoplastic tissues, LDE may be regardedas an advance in chemotherapy with this drug. Introduction In previous studies (MaranhaÄ o etal 1994), we showed that an artificially made micro- emulsion, termed LDE, had the ability to concentrate in neoplastic cells after injection into the bloodstream. LDE is composed of quasispherical nanoparticles basically formed by a monolayer of phosphatidylcholine surrounding a core of cholesteryl esters. Small amounts of unesterified cholesterol and triglycerides are also present. LDE mimics the lipidic portion of low-density lipoprotein (LDL) and in contact with plasma it acquires apolipoproteins from the circulating native lipoproteins. LDE is then removed from the circulation into the body tissues by LDL receptors present on the cell membrane. One of the proteins acquired by the microemulsion, apolipoprotein E (apo E), is recognised by the LDL receptors and enables the LDE particles to bind to the receptors. LDE is then internalised into the cytoplasm via the LDL-receptor- mediated endocytosis. Because most cancer cells show LDL receptor upregulation, LDE may target those cells and eventually may specifically deliver to them chemother- apeutic agents loaded into the microemulsion particles (MaranhaÄ o etal 1993). In recent studies, direct evidence was provided of the selective LDE uptake by neoplastic tissues in patients with ovarian and breast carcinoma (Ades et al 2001; Graziani et al 2002). Previously, it had been suggested that in patients with acute myelocytic leukaemia those leukaemia cells also take up the microemulsion. It was also shown in patients with advanced cancers that the association of carmustine with LDE markedly diminishes the