Strengthening Gastric Cancer Therapy by Trastuzumab-Conjugated Nanoparticles with Simultaneous Encapsulation of Anti-MiR-21 and 5-Fluorouridine.

ABSTRACT

BACKGROUND/AIMS: MicroRNA-21 is an oncogenic miR (oncomiR) frequently elevated in gastric cancer (GC). Overexpression of miR-21 decreases the sensitivity of GC cells to 5-fluorouridine (5-Fu) and trastuzumab, a humanized monoclonal antibody targeting human epidermal growth factor receptor 2 (HER2). Receptor-mediated endocytosis plays a crucial role in the delivery of biotherapeutics including anti-miRNA oligonucleotides (AMOs). This study is a continuation of earlier findings involving poly(ε-caprolactone) (PCL)-poly (ethylene glycol) (PEG) nanoparticles (PEG-PCL NPs), which were coated with trastuzumab to target GC with HER2 receptor over-expression using anti-miRNA-21 (AMO-21) and 5-Fu. METHODS: HER-PEG-PCL NPs were prepared by one-step carbodiimide coupling using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDAc) and Sulfo-NHS in aqueous phase. Covalent coupling of amino groups at the surface of PEG-PCL with the carboxyl groups of trastuzumab was analyzed by X-ray photoelectron spectroscopy (XPS). AMO-21/5-Fu NPs were formulated by a double-emulsion solvent evaporation technique. The cell line specificity, cellular uptake and AMO-21 delivery were investigated through the rhodamine-B-labeled 6-carboxyfluorescein (FAM)-AMO-21-PEG-PCL NPs coated with or without the antibody in both Her2-positive (NUGC4) and negative GC cells (SGC7901) visualized by fluorescence microscopy. The cytotoxicity of the HER-PEG-PCL NPs encapsulating AMO-21 was evaluated by MTT and apoptosis. Real-time reverse-transcription polymerase chain reaction (RT-PCR) was used to examine miR-21 and phosphatase and tensin homolog (PTEN) and Sprouty2 expression in GC cell lines. The antitumor effects of AMO-21/5-Fu NPs were compared with other groups in xenograft gastric cancer mice. RESULTS: The antibody conjugates significantly enhanced the cellular uptake of NPs. The AMO-21/5-Fu NPs effectively suppressed the target miRNA expression in GC cells, which further up-regulated PTEN and Sprouty2. As a result, the sensitivity of HER2-expressing gastric cancer to trastuzumab and 5-Fu were enhanced both in vitro and in vivo. The approach enhanced the targeting by trastuzumab as well as antibody-dependent cellular cytotoxicity (ADCC) of immune effector cells Conclusions: Taken together, the results provide insight into the biological and clinical potential of targeted AMO-21 and 5-Fu co-delivery using modified trastuzumab for GC treatment.