Pathologically proven regional lymph node metastasis affects the prognosis in early stage oral cancer. Therefore we investigated invasive tumor patterns predicting nodal involvement and survival in patients with clinically node-negative T1 and T2 oral squamous cell carcinoma (cT1,2N0M0 OSCC). Ninety-one cases of cT1,2N0M0 OSCC treated with transoral resection of the primary tumor were assessed based on 3 types of invasive tumor patterns on histopathologic and pancytokeratin-stained immunohistological sections: the mode of invasion, worst pattern of invasion (WPOI), and tumor budding. The correlations among invasive tumor patterns, regional metastasis, and disease-free survival were analyzed. Of the 91 cases, 22 (24%) had pathologically proven regional metastasis. The mode of invasion (p<0.01) and tumor budding (p<0.01) were associated with regional metastasis as well as lymphovascular invasion (p = 0.04) in univariate analysis. In logistic regression analysis, however, tumor budding was the only independent predictor of regional metastasis (hazard ratio (HR) = 3.05, 95% confidence interval (CI) = 0.29–5.30, p<0.01). All three invasive patterns, the mode of invasion, WPOI, and tumor budding, were found to be significant predictors of 5-year disease-free survival (p<0.01, p = 0.03, and p<0.01, respectively) as well as lymphovascular invasion (p = 0.02) and perineural invasion (p = 0.02). A final model for Cox multivariate analysis identified the prognostic advantage of the intensity of tumor budding (HR = 2.19, 95% CI = 1.51–3.18, p<0.01) compared with the mode of invasion and WPOI in disease-free survival. Our results indicate that the intensity of tumor budding may be a novel diagnostic biomarker, as well as a therapeutic tool, for regional metastasis in patients with cT1,2N0M0 OSCC. If the pancytokeratin-based immunohistochemical features of more than five buds, and a grade 4C or 4D mode of invasion are identified, careful wait-and-see follow-up in a short period with the use of imaging modalities is desirable. If there are more than ten buds, a grade 4D mode of invasion, or WPOI-5 in the same section, wide resection of the primary tumor with elective neck dissection should be recommended.

Gene targeting using short interfering RNA (siRNA) has become a common strategy to explore gene function because of its prominent efficacy and specificity. For the application of siRNA technology to gene therapy, however, still more efficient transduction of siRNA into target cells is needed. In this study, we developed an adenoviral vector harboring a tandem-type siRNA expression unit, in which sense and antisense strands composing the siRNA duplex were separately transcribed by two human U6 promoters. Targeting survivin, an antiapoptotic molecule widely overexpressed in malignancies but not detected in terminally differentiated adult tissues, this type of adenoviral vector (Adv-siSurv) successfully exerted a gene knockdown effect and induced apoptosis in HeLa, U251, and MCF-7 cells. These cancer cells, once infected with Adv-siSurv, displayed remarkably attenuated growth potential, both in vitro and in vivo. Moreover, intratumoral injection of Adv-siSurv significantly suppressed tumor growth in a xenograft model using U251 glioma cells. This novel modality may be a promising tool for cancer therapy.

Strategies using mesenchymal stem cell (MSC)-mediated gene therapy have been developed to improve bone healing. However, transduction efficiency into MSCs by each vector is not always high. To overcome this problem, we used a modified adenoviral vector (Adv-F/RGD) with an RGD-containing peptide in the HI loop of the fiber knob domain of adenovirus type 5 (Ad5). Transduction efficiency into bone marrow-derived MSCs with Adv-F/RGD increased 12-fold compared with a vector containing the wild-type fiber (Adv-F/wt) by beta-galactosidase chemiluminescent assay. As a next step, we constructed AxCAhBMP2-F/RGD and AxCAhBMP2-F/wt carrying human bone morphogenetic protein 2 (BMP2). At the same multiplicity of infection, MSCs infected with AxCAhBMP2-F/RGD produced higher amounts of BMP2 than cells infected with AxCAhBMP2-F/wt, and also differentiated towards the osteogenic lineage more efficiently in vitro. Furthermore, using ex vivo gene transduction, we evaluated the potential for ectopic bone formation by the transduced MSCs in vivo. Transduction with AxCAhBMP2-F/RGD exhibited greatly enhanced new bone formation. These data suggest that Adv-F/RGD is useful for introducing foreign genes into MSCs and that it will be a powerful gene therapy tool for bone regeneration and other tissue engineering.

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In acute myocardial infarction (AMI), prognosis and mortality rate are closely related to the infarct size and the progression of postinfarction cardiac failure. Angiogenic gene therapy has presented a new approach for the treatment of AMI. Angiopoietin-1 (Ang1) is a critical angiogenic factor for vascular maturation and enhances vascular endothelial growth factor (VEGF)-induced angiogenesis in a complementary manner. We hypothesized that gene therapy using Ang1 for AMI might promote angiogenesis cooperatively with intrinsic VEGF, since high concentrations of circulating VEGF have been reported in AMI. To evaluate our hypothesis, we employed a rat AMI model and adenoviral Ang1 (HGMW-approved gene symbol ANGPT1) gene transfer to the heart. A significant increase in capillary density and reduction in infarct sizes were noted in the infarcted hearts with adenoviral Ang1 gene treatment compared with control infarcted hearts treated with saline or adenoviral vector containing the beta-galactosidase gene. Furthermore, the Ang1 group showed significantly higher cardiac performance in echocardiography (55.0% of ejection fraction, P < 0.05 vs control) than the saline or adenoviral controls (36.0 or 40.5%, respectively) 4 weeks after myocardial infarction. The adenoviral delivery of Ang1 during the acute phase of myocardial infarction would be feasible to attenuate the progression of cardiac dysfunction in the rat model.