Distribution of fluorescent nanoparicles in mouse and breast cancer cell model

B77

Purpose: Over the years, despite progress in understanding the molecular mechanism of cancer, targeted delivery of anticancer drugs remains a challenge. Attempts have been made to overcome this challenge by developing biodegradable nanoparticles which have been conjugated with breast cancer cell specific antibodies. In order to facilitate the study of cancer cell specific uptake, such particles are also labeled with fluorescent markers. This report describes our efforts in developing fluorescent tagged biodegradable nanoparticles of PLGA and to study: i) their uptake by cancer cells, and ii) their biodistribution in mouse following systemic delivery. Methods: Particles of PLGA 50:50 containing Coumarin-6 as a fluorescent marker were fabricated using a double emulsion-solvent evaporation technique. The fractionation of larger and smaller particles was achieved by two-step differential centrifugation. The morphology of nanoparticles was determined by scanning electron microscopy and zeta potential (surface charge) was measured by a Zetasizer. Qualitative and quantitative analysis of cellular uptake was done by fluorescent microscopy and flow cytometry using a murine breast cancer cell line (4T1). Tissue distribution of the nanoparticles after systemic delivery was studied in BALB/C female mice by injecting the particles via tail vein. Results: Solvent evaporation method yielded fluorescent PLGA particles with heterogeneous sizes. The median size of the particles was between 500 nm and 7 µm. Scanning electron microscope pictures showed that the particles were smooth and spherical. The average zeta potential of the particles was approximately -6.2 mV. Quantitative FACS analysis showed that smaller size nanoparticles were highly efficient for intracellular delivery to breast cancer cells compared to larger particles. Image analysis of the frozen tissue sections of BALB/C mice showed that fluorescent particles were distributed preferentially into the lungs followed by the liver, spleen, kidney and heart. Conclusion: The results demonstrate that the nano-particles of PLGA were efficiently taken up by the mammary breast cancer cells and that they were distributed preferentially to the sites frequently seen as sites of cancer metastasis in the BALB/C mice following the tail vein injection of breast cancer cells. Moreover, both the cellular uptake and the biodistribution were found to be more efficient with smaller sized fraction of nanoparticles. Acknowledgement: This work was funded in part by the NIH/NIGMS Grant # GM08008-32, NASA Grant # NCC3-946-S4, Louisiana Board of Regents RC/EEP (2006-16), NIH Grant# 5P20CA118768-02, and Louisiana Cancer Research Consortium.