EFFICACY AND ANTITUMOR MECHANISMS OF EGFR ANTISENSE GENE THERAPY

EGFR反义基因治疗的疗效及抗肿瘤机制

• 批准号: 6144203
• 负责人: Jennifer Rubin Grandis
• 金额: $ 34.34万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-08-01 至 2000-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6144203
• 关键词: antisense nucleic acid; apoptosis; carcinogenesis inhibitor; clinical research; clinical trial phase I; dosage; epidermal growth factor; gene therapy; growth factor receptors; human subject; human therapy evaluation; laboratory mouse; liposomes; neoplasm /cancer genetics; neoplasm /cancer transplantation; neoplastic growth; nonhuman therapy evaluation; oral pharyngeal neoplasm; protein isoforms; squamous cell carcinoma; transcription factor

We have demonstrated that EGFR up-regulation in OSCC is due to activated gene transcription and that down-modulation of EGFR results in decreased proliferation of OSCC but not normal cells. Further investigation in our laboratory revealed inhibition of tumor growth in vivo following intratumoral inoculation of an EGFR antisense expression construct based on the U6 small nuclear RNA promoter in complexed with DC-chol liposomes. This anti-tumor effect was accompanied by decreased EGFR protein expression in the tumors and increased apoptosis. Preliminary results suggest that EGFR signaling in OSCC involves constitutive activation of Stat3alpha-beta and that down-modulation of Stat3 using antisense oligonucleotides or dominant negative mutants result in inhibition of OSCC proliferation. The importance of this autocrine pathway is underscore by our finding that protein expression levels of EGFR in the primary OSCC tumor is a significant and independent predictor of decreased survival. Therefore, we propose to test the hypothesis that the loss of growth control in OSCC is mediated through acquisition of an EGFR autocrine signaling pathway, which can be targeted using an antisense gene therapy approach. In specific aim 1 we propose to characterize the effects of EGFR antisense therapy in OSCC in vitro and in murine xenograft models by determining: a) the association between EGFR expression levels and anti-tumor activity; and b) the dose, schedule, and time-dependent parameters for optimal effect. In specific aim 2 we propose to determine the mechanism of the anti-tumor effects of EGFR antisense gene therapy in vitro and in murine xenograft models by: a) characterizing the impact of treatment on expression and activation of specific STAT protein isoforms; and b) examination the consequences of therapy on apoptosis. In specific aim 3 we propose to determine in the phase I setting, the maximally tolerated dose (MTD) and biologic effects of intratumoral liposome-mediated EGFR antisense gene therapy in OSCC patients.

我们已经证明，OSCC 中 EGFR 的上调是由于基因转录激活所致，而 EGFR 的下调会导致 OSCC 增殖减少，但不会导致正常细胞增殖减少。我们实验室的进一步研究表明，在肿瘤内接种基于 U6 小核 RNA 启动子与 DC-chol 脂质体复合的 EGFR 反义表达构建体后，体内肿瘤生长受到抑制。这种抗肿瘤作用伴随着肿瘤中 EGFR 蛋白表达的降低和细胞凋亡的增加。初步结果表明，OSCC 中的 EGFR 信号传导涉及 Stat3α-β 的组成型激活，并且使用反义寡核苷酸或显性失活突变体下调 Stat3 会导致 OSCC 增殖的抑制。我们的发现强调了这种自分泌途径的重要性，即原发性 OSCC 肿瘤中 EGFR 的蛋白表达水平是生存率下降的重要且独立的预测因子。因此，我们建议检验以下假设：OSCC 生长控制的丧失是通过获得 EGFR 自分泌信号通路介导的，可以使用反义基因治疗方法来靶向该信号通路。在具体目标 1 中，我们建议通过确定以下因素来表征 EGFR 反义疗法在体外 OSCC 和小鼠异种移植模型中的效果： a) EGFR 表达水平与抗肿瘤活性之间的关联； b) 最佳效果的剂量、时间表和时间相关参数。在具体目标 2 中，我们建议通过以下方式确定 EGFR 反义基因治疗在体外和小鼠异种移植模型中的抗肿瘤作用机制： a) 表征治疗对特定 STAT 蛋白亚型表达和激活的影响； b) 检查治疗对细胞凋亡的影响。在具体目标 3 中，我们建议在 I 期试验中确定 OSCC 患者瘤内脂质体介导的 EGFR 反义基因治疗的最大耐受剂量 (MTD) 和生物学效应。