Iontophoresis-Enhanced Buccal Delivery of Cisplatin-Encapsulated Chitosan Nanoparticles for Treating Oral Cancer in a Mouse Model

Yi-Wen Chen,1,2 Ai-Chia He,3 Tzu-Yun Huang,1,3 De-Hao Lai,3 Yi-Ping Wang,1 Wei-Wen Liu,1,3 Wei-Ting Kuo,1,3 Hsin-Han Hou,1,3 Shih-Jung Cheng,1,3 Chen-Yi Lee,1 Wei-Chun Chuang,1 Che-Chen Chang,4 Bor-Shiunn Lee1,3 1Department of Dentistry, National Taiwan University Hospital, Taipei, 100229, Taiwan; 2Graduate Institute of Clinical Dentistry, School of Dentistry, National Taiwan University, Taipei, 100229, Taiwan; 3Graduate Institute of Oral Biology, School of Dentistry, National Taiwan University, Taipei, 100229, Taiwan; 4Department of Chemistry, National Taiwan University, Taipei, 10617, TaiwanCorrespondence: Bor-Shiunn Lee, Graduate Institute of Oral Biology, School of Dentistry, National Taiwan University, No. 1, Changde Street, Taipei, 100229, Taiwan, Tel +886-2-23123456 ext 66846, Fax +886-2-2383-1346, Email [email protected]: Cisplatin is one of the most effective chemotherapeutic drugs used in oral cancer treatment, but systemic administration has side effects. The purpose of this study was to evaluate the effect of iontophoresis on the enhancement of cisplatin release from cisplatin-encapsulated chitosan nanoparticles.Methods: The effect of different mass ratios of chitosan to tripolyphosphate (TPP) (5:1, 10:1, 15:1, 20:1) on the encapsulation efficiency of cisplatin was investigated. Uptake of cisplatin-encapsulated chitosan by cells was observed using a confocal laser scanning microscope. The cell viability at different cisplatin concentrations was examined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Three iontophoresis methods, namely constant-current chronopotentiometry (CCCP), cyclic chronopotentiometry (CCP), and differential pulse voltammetry (DPV), were used to enhance cisplatin release from cisplatin-encapsulated chitosan nanoparticles. In addition, mouse oral squamous cell carcinoma cell lines were implanted into the mouse oral mucosa to induce oral cancer. The effects of enhanced cisplatin release by CCCP, CCP, and DPV on tumor suppression in mice were evaluated. Tumors and lymph nodes were isolated for hematoxylin-eosin staining and immunohistochemistry staining including Ki-67 and pan CK after sacrifice. Inductively coupled plasma mass spectrometry was conducted to quantify the platinum content within the tumors.Results: The results showed that nanoparticles with a mass ratio of 15:1 exhibited the highest cisplatin encapsulation efficiency (approximately 15.6%) and longest continued release (up to 35 days) in phosphate buffered saline with a release rate of 100%. Cellular uptake results suggested that chitosan nanoparticles were delivered to the cytoplasm via endocytosis. The results of the MTT assay revealed that the survival rate of cells decreased as the cisplatin concentration increased. The CCP (1 mA, on:off = 1 s: 1 s) and DPV (0– 0.06 V) groups were the most effective in inhibiting tumor growth, and both groups exhibited the lowest percentage of Ki-67 positive and pan CK positive.Conclusion: This study is the first to investigate and determine the efficacy of DPV in enhancing in vivo drug release from nanoparticles for the treatment of cancer in animals. The results suggest that the CCP and DPV methods have the potential to be combined with surgery for oral cancer treatment. Keywords: oral cancer, cisplatin, chitosan, nanoparticle, iontophoresis