ABSTRACT
Liposomes carrying chemotherapeutic drugs can accumulate passively in solid tumors at high levels. However, additional targeting of the liposomes towards e.g. receptors expressed on cancer cells may improve their interaction and therapeutic properties. In this study, we designed a liposomal delivery system, which utilizes the intrinsic characteristics of HER2-positive tumors to ensure efficient delivery of oxaliplatin to the cancer cells. On the liposome surface, trastuzumab, an antibody specific to the HER2 receptor, was shown to facilitate internalization by the cancer cells. A polyethylene glycol (PEG) layer on the liposome surface provides protection from mononuclear phagocyte system uptake. To optimize the interaction between liposomes and cancer cells, a protease-sensitive cleavable peptide linker was inserted at the base of each PEG. The PEG layer is then cleaved off by intra- and extracellular matrix metalloproteinases (MMPs) upon accumulation in the tumor. Our data demonstrate that the removal of PEG significantly destabilizes the liposomes and leads to substantial oxaliplatin release. The proposed beneficial effect of combining antibody-mediated internalization with MMP sensitivity was confirmed in a series of in vivo studies using ovarian cancer xenograft models. The results demonstrated that HER2-targeted MMP-sensitive liposomes have superior anticancer activity compared to non-targeted and non-cleavable liposomes.
Subject(s)
Antineoplastic Agents , Liposomes , Ovarian Neoplasms , Oxaliplatin , Polyethylene Glycols , Receptor, ErbB-2 , Trastuzumab , Female , Ovarian Neoplasms/drug therapy , Ovarian Neoplasms/pathology , Animals , Humans , Receptor, ErbB-2/metabolism , Receptor, ErbB-2/immunology , Oxaliplatin/administration & dosage , Cell Line, Tumor , Polyethylene Glycols/chemistry , Polyethylene Glycols/administration & dosage , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/therapeutic use , Antineoplastic Agents/pharmacology , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/chemistry , Trastuzumab/administration & dosage , Trastuzumab/chemistry , Mice, Nude , Drug Delivery Systems , Organoplatinum Compounds/administration & dosage , Organoplatinum Compounds/chemistry , Organoplatinum Compounds/pharmacology , Xenograft Model Antitumor Assays , Matrix Metalloproteinases/metabolism , Mice , Mice, Inbred BALB CABSTRACT
Members of the TMEM16 (Anoctamin) family of membrane proteins have been shown to be essential constituents of the Ca(2+)-activated Cl(-) channel (CaCC) in many cell types. In this study, we have investigated the electrophysiological properties of mouse TMEM16F. Heterologous expression of TMEM16F in HEK293 cells resulted in plasma membrane localization and an outwardly rectifying ICl,Ca that was activated with a delay of several minutes. Furthermore, a significant Na(+) current was activated, and the two permeabilities were correlated according to PNa = 0.3 PCl. The current showed an EC50 of 100 µM intracellular free Ca(2+) concentration and an Eisenman type 1 anion selectivity sequence of PSCN > PI > PBr > PCl > PAsp. The mTMEM16F-associated ICl,Ca was abolished in one mutant of the putative pore region (R592E) but retained in two other mutants (K616E and R636E). The mutant K616E had a lower relative permeability to iodide, and the mutant R636E had an altered anion selectivity sequence (PSCN = PI = PBr = PCl > PAsp). Our data provide evidence that TMEM16F constitutes a Ca(2+)-activated anion channel or a pore-forming subunit of an anion channel with properties distinct from TMEM16A.
