Despite existing chemotherapy and surgical resection strategies, salivary gland adenocarcinoma(AdCa NOS) is one of the major causes of mortality among malignant salivary gland tumors. New therapeutic measure are needed to improve the outcome for patients with AdCa. Overexpression of urokinase-type plasminogen activator receptor/urokinase-type plasminogen activator(uPAR-uPA) has been implicated in progression and metastasis of oral cancer. RNA interference(RNAi) which has emerged as an effective method to target specific genes for silencing has provided new opportunities for cancer therapy. But there has been rarely reported using RNAi-uPAR/uPA transfection in salivary gland AdCa. The purpose of this study were to examine the specific inhibition of uPAR/uPA mRNA and protein expression by RNAi transfection of uPAR/uPA through RT-PCR and Immunoslot blot, and to study tumor cell proliferation activity, adhesion, invasion and migration of SGT cell line in vitro compared to the controls. In adhesion assay, cells transfected with RNAi-uPAR/uPA inhibited markedly adhesion to vitronectin compared to parental cells. Angiogenic assays revealed a significant decrease in the angiogenic potential of SGT cells downregulated by both uPAR and uPA. In migration assay, suppressing uPAR and uPA inhibited the capacity of the cells to migrate compared to parental cells. In invasion assay, cells transfected with RNAi-uPAR/uPA showed the maximum decrease in invasion when compared to all other treatment conditions. RNAi expressing plasmids efficiently downregulated mRNA and protein expression of uPAR and uPA. Cell cycle analysis showed that the simultaneous downregulation of uPAR and uPA caused the accumulation of cells in the sub-G0/G1 phase in SGT cells. Immunoslot blot analysis revealed that downregulation of uPAR and uPA caused the prominent activation of caspase 8. It suggested that the RNAi targeting of the uPAR/uPA system could have a therapeutic potentiality for malignant salivary gland tumors.

New therapeutic measure are needed to improve the outcome for patients with oral squamous cell carcinoma(OSCC) because OSCC continues to portend a relatively unfavorable prognosis. Recently RNA interference(RNAi) has emerged as an effective method to target specific genes for silencing. Although overexpression of urokinase-type plasminogen activator receptor(uPAR) has been implicated in progression and metastasis of OSCC, the transfection effect of RNAi- uPAR on OSCC has been rarely reported. The purpose of this study were to examine the efficient and specific inhibition of uPAR mRNA and protein expression by siRNA targeting of uPAR through RT-PCR and immunoslot blotting, and to study cell proliferation activity, adhesion, invasion and migration in vitro compared to the controls. In MTT assay, siRNA-uPAR transfected cells showed about 70-80% cell proliferation compared to OSCC cell lines after 2 days. In adhesion assay, siRNA-uPAR transfected cells showed about 20-30% adhesion activity compared to OSCC cell lines, but similar features to those of BSA coated wells. In migration assay, siRNA-uPAR transfected cells showed about 60% migration activity compared to OSCC cell lines, but higher 3.5 folds to those of BSA coated wells. In invasion assay, siRNA-uPAR transfected cells showed about 55% invasive activity compared to parental cell lines. mRNA expression of siRNA-uPAR transfected cells showed about 10-15 % compared to parental cell lines by RT-PCR. Protein expression of siRNA-uPAR transfected cells showed about 25% compared to parental cell lines by ELISA assay. It suggested that RNAi-uPAR tranfection might be used as a potent and specific therapeutic tool for the treatment of oral squamous cell carcinoma, especially in inhibiting invasion and metastasis.