2013  •   Health Sciences

Activation of HIF-1? in 3D Ovarian Tumor Spheroids and Evaluation of 2-Methoxyestradiol Therapy using Targeted Polymeric Nanoparticle Formulation

Lead Presenter: Ankita Raikar

Purpose: To evaluate activation of HIF-1?, a major determinant of tumor resistance and metastasis, using 3D cellular spheroid models of human ovarian adenocarcinoma (SKOV3) and to investigate whether targeted polymeric nanoparticles encapsulating 2-methoxyestradiol (2-ME), an inhibitor of HIF-1?, can be used for more effective delivery and therapy. æExperimental Methods: SKOV3 cellular spheroids were grown in culture for 3 and 5 days using the hanging drop method. HIF-1? activation was evaluated by RT-PCR and ELISA in both 2D culture (under normoxia and hypoxia) and 3D spheroids. Control PLGA-PEG-modified and Lyp-1 peptide-targeted PLGA-PEG/PCL blend nanoparticles were formulated by solvent evaporation technique for 2ME encapsulation and delivery. Using rhodamine 123-encapsulated nanoparticles, permeability through spheroids was assessed by Z-stack analysis with fluorescence confocal microscopy. ææResults: SKOV3 cellular spheroids were roughly circular in shape and 200-400um in diameter. RT-PCR results confirmed higher HIF-1? mRNA levels and ELISA confirmed higher protein levels in 3D spheroids compared to 2D SKOV3 cell cultures grown under both normoxic (21% O2) and hypoxic (0.5% O2) conditions for 3 and 5 days The hydrodynamic diameter of nanoparticles was around 250nm and surface charge around -45mV. Fluorescence confocal microscopy studies showed that the nanoparticle formulation has greater permeability efficiency to the core of the spheroids as compared to the free dye. æConclusions: The preliminary studies showed that spheroids exhibited higher HIF-1? levels compared to 2D cell cultures and served as better models to perform in vitro studies. Also, PEG-modified PLGA/PCL nanoparticles assisted in better delivery to the core of the spheroid.