RESUMEN
Targeting phospholipid biosynthesis, specifically phosphatidylcholine (PC), which is enhanced in tumor cells, has been proven a suitable antitumor strategy. In fact, the overexpression of the choline kinase α1 (ChoKα1) isoform has been found in malignant cells and tumors, thus becoming an excellent antitumor target. ChoKα1 inhibitors are being synthesized at the present that show a large inhibitory activity. Two of them have been chosen in this study as representatives of different structural families: a biscationic biphenyl derivative of thieno[3,2-d]pyrimidinium substituted with a cyclic amine (here referred to as Fa22) and a biscationic biphenyl thioethano derivative of 7-chloro-quinolinium substituted with a pyrrolidinic moiety (here referred to as PL48). However, the potential use of these types of compounds in systemic treatments is hampered because of their low specificity. In fact, to enter the cell and reach their target, these inhibitors use choline transporters and inhibit choline uptake, being that one of the causes of their toxicity. One way to solve this problem could be allowing their entrance into the cells by alternative ways. With this goal, MamC-mediated magnetic nanoparticles (BMNPs), already proven effective drug nanocarriers, have been used to immobilize Fa22 and PL48. The idea is to let BMNPs enter the cell (they enter the cell by endocytosis) carrying these molecules, and, therefore, offering another way in for these compounds. In the present study, we demonstrate that the coupling of Fa22 and PL48 to BMNPs allows these molecules to enter the tumoral cell without completely inhibiting choline uptake, so, therefore, the use of Fa22 and PL48 in these nanoformulations reduces the toxicity compared to that of the soluble drugs. Moreover, the nanoassemblies Fa22-BMNPs and PL48-BMNPs allow the combination of chemotherapy and local hyperthermia therapies for a enhanced cytotoxic effect on the tumoral HepG2 cell line. The consistency of the results, independently of the drug structure, may indicate that this behavior could be extended to other ChoKα1 inhibitors, opening up a possibility for their potential use in clinics.
RESUMEN
Purpose: The lack of specificity of conventional chemotherapy is one of the main difficulties to be solved in cancer therapy. Biomimetic magnetoliposomes are successful chemotherapy controlled-release systems, hyperthermia, and active targeting agents by functionalization of their surface with monoclonal antibodies. The membrane receptor Leucine-rich repeat-containing G-protein coupled receptor 5 (LGR5) stands out as colorectal cancer (CRC) biomarker and appears to be related to treatment resistance and the development of metastasis. The aim of this study was to assess the effectiveness and safety of LGR5-targeted biomimetic magnetoliposomes loaded with oxaliplatin (OXA) or 5-fluorouracil (5-FU) in the selective treatment of CRC and their possible application in hyperthermia. Methods: Synthesis, characterization and determination of heating capacity of magnetoliposomes transporting OXA or 5-FU (with and without LGR5 functionalization) were conducted. In vitro antitumoral activity was assayed in multiple colorectal cell lines at different times of exposition. In addition to this, cell internalization was studied by Prussian Blue staining, flow cytometry and fluorescence microscopy. In vivo acute toxicity of magnetoliposomes was performed to evaluate iron-related toxicity. Results: OXA and 5-FU loaded magnetoliposomes functionalized with LGR5 antibody showed higher cellular uptake than non-targeted nanoformulation with a reduction of the percentage of proliferation in colon cancer cell lines up to 3.2-fold of the IC50 value compared to that of free drug. The differences between non-targeted and targeted nanoformulations were more evident after short exposure times (4 and 8 hours). Interestingly, assays in the MC38 transduced cells with reduced LGR5 expression (MC38-L(-)), showed lower cell internalization of LGR5-targeted magnetoliposomes compared to non-transduced MC38 cell line. In addition, magnetoliposomes showed an in vitro favorable heating response under magnetic excitation and great iron-related biocompatibility data in vivo. Conclusion: Drug-loaded magnetoliposomes functionalized with anti-LGR5 antibodies could be a promising CRC treatment strategy for LGR5+ targeted chemotherapy, magnetic hyperthermia, and both in combination.
