Liposome-based Nanoprobe With Antibody Conjugation Combined With MRI Contrast Encapsulation as a Potential Diagnostic and Therapeutic Agent for Human Head-and-Neck Cancer Cells

Abstract

Purpose/Objective(s): To develop a tumor-targeting nanoprobe that selectively binds to epidermal growth factor receptor (EGFR) overexpressing tumor cells by encapsulating a magnetic resonance (MR) imaging contrast agent gadolinium (Gd) and conjugating an EGFR-specific monoclonal antibody within liposome particles. Materials/Methods: Encapsulation of Gd within liposome particles was optimized by combining different concentrations of Dipalmitoylphosphatidylcholine (DPPC), cholesterol, 1,2-dipalmitoyl-sn-glycero-3phosphoethanolamine-N-(biotinyl) (Biotinyl PE), and Diethylenetriaminepentaacetic acid-bis(stearylamide)-Gd (DTPA-BSA-Gd). Avidin was included in the liposomal membrane to conjugate with biotinmodulated anti-EGFR antibody. The resultant probes were characterized for their particle size, surface potential and structural integrity with dynamic light scattering (DLS), Zetasizer, and transmission electronic microscopy (TEM), respectively. In-vitro targeting specificity of the probe was evaluated with fluorescence microscopy by comparing the binding capacity of a fluorescence-labeled probe in a human head and neck cancer cell line, 15B (high EGFR overexpression) and a human embryonic kidney cell line, HEK293 (low EGFR control). The subsequent cytotoxicity toward these two cell lines was quantified by MTT assay. Finally, the in vitro applicability of MRI contrast Gd within the probe was demonstrated by detecting cellbound particles with MR imaging. Results: Using TEM scanning, our probe was observed to be spherical in shape and identical in particle size, approximately 100nm. The surface potential was +5.13mV. The EGFR-specific targeting assay with fluorescence-labeled probes showed significantly high occupancy of the probe on the surface of 15B cells in comparison with HEK293 cells. Furthermore, the T1 relaxation time of this EGFR-specific probe attached to15B cells was measured to be 379.3 ms, shorter than that of control vector without anti-EGFR conjugation (516.2 ms). Conclusion: We have demonstrate targeting specificity of a multi-functional nanoprobe in discriminating high and low EGFR-expressing cells. The incorporation of MRI contrast Gd successfully revealed the cell-targeted probe under MR imaging. These results suggest that this probe can function as a targeting tracer for tumor cells, potentiating it in improving treatment efficacy through MR imaging. Author Disclosure: Y. Kuo: None. C. Hung: None. S.R. Raghavan: None. W.D. D’Souza: None.