| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2004: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2003: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2002: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
Objective We report the in viitmestablishment of a highly stable green fluorescent protein (GFP) -expressing transfectant of highly-invasive human tongue squamous cell carcinoma cells, SAS-Hi. The fluorescent cells permitted the visualization of tumor growth, local invasion, micrmetastasis, and eeevical lymph node metastasis aver submucosal injection into the tongue of nude mice. This model should be useful for studying the metastatic process and for evaluating anti-metastatic agents in pre-clinesl trials. Using GFP-tagged human tongue squamous cell carcinoma cells, we examined the LY294002 reduce local invasion and cervical lymph node metastasis in vivo. Additionally, using this model, eflect ofanti invasion and -metastasis by antimic mbial peptide hCAP18, and MEK inhibitorwas studied. Methods High invasion human tongue squamous cell carcinoma cell line, as SAS-H1 cells were transfected with the pEGFP-N1 plasmid (CLONTECH) using Nucleofector4device. Exponentially growing GFP-tagged tu
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mor cells suspension (2X10^6) was injected into the tongue of 12 KSN strained male athymic nude mice (4-6-week-old). Day 1 post-injection, mice were treated with LY294002 (0, 25, 50; or 100 mg/kg) i.p. for 4 weeks. At week 4, tumor-bearing mice were sacrificed and autopsied under light and fluorescent multi-angle digital microscope (KEYENCE). Furthermore, mice were treated with hCAP18 peptide (0, 50, 100 μg/ml) in local injection twice a week for 4weeks.
Results At day 1 post-injection, all mice expressed fluorescent tongue tumors. At week 4, disseminated GFP-tagged cells around the primary tumor could be observed, and all tumors metastasized to the cervical lymph nodes in sacrificed mice. GFP-tagged tumor cells detected in fresh cervical lymph nodes. LY294002-treated group was reduced dose-dependently local invasion and lymph node metastasis than control group at 4 weeks. 100 mg/kg of LY294002 treated mice were being completely free from cervical metastasis. On one hand. hCAP 18-treated group was reduced dose dependently tumor volume in primary site, but not inhibited cervical lymph node metastasis.
Conclusion These results that LY294002 inhibits local invasion, micrometastasis and cervical lymph node metastasis in viva, suggesting an important role of P13-K inhibitors as a potentially useful treatment for oral squamous cell carcinoma. Our data suggest that a combination of a P13K inhibitor and antimicrobial peptide, hCAP 18 may provide an effective approach to inhibiting tumor metastasis and tumor growth in oral squamous cell carcinoma. Less
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