CPE Peptide-Based Nanoparticles for the Diagnosis and Therapy of Chemotherapy Res

摘要 卵巢癌仍是妇科肿瘤中死亡率最高的癌症。虽然 许多卵巢癌患者对手术和化疗的标准组合完全有反应， 90%的人后来发展为复发性耐化疗癌症，并不可避免地死于疾病。因此，在本发明中， 开发针对复发性/化疗耐药性卵巢癌的创新有效疗法 仍然是改善公共卫生的一个高度优先事项。使用高通量技术分析基因 卵巢癌的指纹，我们最近发现了极高的表达基因编码 蛋白质密蛋白-3和密蛋白-4。因为claudin-3和claudin-4是梭菌的上皮受体 产气荚膜杆菌肠毒素（CPE），并足以介导CPE结合，我们假设，使用 可生物降解的纳米颗粒如聚（乳酸-共-乙醇酸，即，PLGA-NP）封装 与CPE的结合结构域复合以靶向卵巢癌细胞的治疗剂，是一种新的， 这是一种潜在的高效治疗方法，用于治疗化疗耐药性卵巢癌。 与这一观点一致，初步的体外数据清楚地表明，荧光素（FITC）标记的-C-CPE 有效地结合并迅速内化在化疗耐药的原代OSPC细胞系中，这表明C- CPE能够结合药物，也能够跨细胞肿瘤膜转运药物。更重要的是， 初步的体内实验结果表明，CPE-PLGA-NP选择性地包封荧光染料（香豆素）， 在化疗耐药的卵巢肿瘤中体内蓄积， RES器官和仅在各种其他器官的正常细胞中的背景结合。因此，本提案 有三个相关的具体目的：1）表征荧光PLGA-NP包封模型和治疗 与不同长度的CPE肽复合的药物（即，30 aa、17 aa和9 aa），并评估结合活性 以及这种NP在针对原发性化疗抗性卵巢癌的多个体外测定中的治疗功效 2）检测包封香豆素的PLGA-NP的分布和药代动力学 与125 I标记的C-CPE肽在临床相关的主要化疗的动物模型中体内复合 耐药卵巢癌和3）检查包裹质粒DNA的PLGA-NP的治疗潜力 编码白喉毒素A自杀蛋白的基因的启动子序列高度控制， 优先在卵巢癌中有活性（即，间皮素和HE 4/WFDC 2），在体内与CPE肽复合。 原发性化疗耐药卵巢癌的临床相关动物模型。完成后 在这个项目中，我们将定位于快速将这项研究转化为一种新颖，高效的治疗方法。 对化疗耐药疾病患者的治疗。

Abstract Ovarian carcinoma remains the cancer with the highest mortality rate among gynecological tumors. Although many ovarian cancer patients fully respond to the standard combination of surgery and chemotherapy, nearly 90% later develop recurrent chemotherapy-resistant cancer and inevitably succumb to their disease. Thus, development of innovative, effective therapies against recurrent/chemotherapy-resistant ovarian cancer remains a high priority for improving public health. Using high-throughput technologies to analyze genetic fingerprints of ovarian cancer, we have recently discovered extremely high expression of the genes encoding the proteins claudin-3 and claudin-4. Because claudin-3 and -4 are the epithelial receptors for Clostridium perfringens enterotoxin (CPE), and are sufficient to mediate CPE binding, we hypothesize that using biodegradable nanoparticles such as poly(lactic-co-glycolic acid, i.e., PLGA-NP) encapsulating therapeutic agents complexed to the binding domain of CPE, to target ovarian cancer cells, is a novel, potentially highly effective therapeutic approach to treat chemotherapy-resistant ovarian cancer. Consistent with this view, preliminary in vitro data clearly showed that fluorescein(FITC)-labeled-C-CPE effectively binds and rapidly internalizes in chemotherapy-resistant primary OSPC cell lines, suggesting that C- CPE is capable to bind and also of translocating drugs across cell tumor membrane. More importantly, preliminary in vivo results showed that CPE-PLGA-NP encapsulating a fluorescent dye (coumarin) selectively accumulate in vivo in chemotherapy resistant ovarian tumors with minimal nonspecific accumulation in RES organs and only background binding in normal cells of various other organs. Accordingly, this proposal has three related specific aims: 1) Characterize fluorescent PLGA-NP encapsulating model and therapeutic drugs, complexed to CPE peptides of different lengths (i.e., 30aa, 17aa and 9 aa), and evaluate binding activity and therapeutic efficacy of such NP in multiple in vitro assays against primary chemotherapy resistant ovarian carcinoma cell lines, 2) Examine the distribution and pharmacokinetics of PLGA-NP encapsulating coumarin complexed to 125I labeled C-CPE peptide in vivo in clinically relevant animal models of primary chemotherapy resistant ovarian carcinoma and 3) Examine the therapeutic potential of PLGA-NP encapsulating plasmid DNA encoding Diptheria Toxin A suicide protein under the control of promoter sequences of genes highly and preferentially active in ovarian cancer (i.e., mesothelin and HE4/WFDC2), complexed to CPE peptide in vivo in clinically relevant animal models of primary chemotherapy resistant ovarian carcinoma. Upon completion of this project, we will be positioned to quickly translate this research into a novel, highly effective therapeutic treatment for patients with chemotherapy-resistant disease.