Targeting TF/CD44v6 for In Vivo Nano-generated alpha-therapy of Ovarian Cancer

描述(申请人提供)：这项研究的目标是使用放射性标记的工程化多肽支架，利用体内匹配的~(203)铅/~(212)铅粒子发生器，开发敏感的卵巢癌诊断和治疗(治疗)试剂。A粒子的高线性能量转移显著高于b粒子发射的治疗常用的放射性核素。虽然早期卵巢癌的存活率很高，但晚期疾病往往是致命的。卵巢癌预后差的原因是诊断复杂，这是由于基本上没有症状的疾病发展，存在侵袭性转移细胞，以及缺乏疾病分期的生物标志物。我们建议靶向肿瘤特异性Thomsen-Friedenreich(TF)糖抗原和卵巢癌生物标记物CD44v6，这是透明质酸(HA)受体CD44的肿瘤特异性剪接变体。CD44v6糖基化导致HA结合和内化增加，导致一系列信号事件。Tf二糖显示在CD44上，可能位于肿瘤特异性V6区域，与侵袭和转移有关。假设靶向这个tf/CD44v6糖表位将提供a粒子辐射的特异性定位。 放射对肿瘤的成像和治疗。所使用的靶向载体将是25个氨基酸的海普西丁的工程衍生物，它与TF/CD44v6结合，并显示出与抗体或小肽相比的最佳体内特性，包括高体内稳定性和快速通过肾脏，从而增加肿瘤滞留和减少非靶器官辐射。延长生物利用度和最大限度地减少对健康组织的暴露可能通过传递HA来进一步实现，以确保TF/CD44v6多肽的内化。这项研究的具体目的是：1)筛选和鉴定Tf/CD44v6-Avid噬菌体展示衍生肽；2)将Tf/CD44v6-Avid多肽(S)生物工程到海普西丁支架中，并研究其生物素化和1，4，7，10-tetraazacyclododecane-1，4，7，10-tetraacetic酸(DOTA)结合物的体外结合卵巢癌细胞的特性；以及3)测定212Pb和203Pb标记的多肽结合物在人卵巢癌移植瘤小鼠体内的药代动力学、生物分布和肿瘤靶向性。HA处理对放射性标记的海普西丁多肽支架的肿瘤滞留的影响将通过生物分布研究来确定。为了可视化肿瘤归巢，将用203Pb标记多肽构建体，用于单光子发射计算机断层成像。所提出的a-颗粒标记多肽支架体内靶向治疗卵巢癌具有广泛的适用性，这一点很重要，因为早期卵巢癌通常是无症状的，而晚期卵巢癌通常是致命的。此外，所提出的技术可以应用于靶向和杀死其他癌细胞，特别是因为转铁蛋白存在于80%的腺癌上。

DESCRIPTION (provided by applicant): The goal of this research is to use radiolabeled engineered peptide scaffolds for development of sensitive ovarian carcinoma diagnostic and therapeutic (theranostic) agents using a matched pair 203Pb/212Pb in vivo ¿-particle generator. The high linear energy transfer of a-particle is significantly higher than that of b-particle emittng radionuclides commonly used for therapy. Although the survival rates of early stage ovarian cancer are high, late stage disease is often fatal. The poor prognosis of ovarian cancer results from complicated diagnosis due to a largely asymptomatic disease development, the presence of aggressive metastatic cells, as well as a lack of biomarkers for disease stages. We propose to target the carcinoma-specific Thomsen- Friedenreich (TF) glycoantigen and the ovarian carcinoma biomarker CD44v6, a tumor-specific splice variant of the hyaluronic acid (HA) receptor CD44. CD44v6 glycosylation results in increased HA binding and internalization, leading to a multitude of signaling events. TF disaccharide is displayed on CD44, likely in the cancer-specific v6 region and is associated with invasion and metastasis. It is hypothesized that targeting this TF/CD44v6 glycoepitope will afford the specific localization of a-particle radiation radioactivity to tumors for both imaging and therapy. The targeting vehicle employed will be an engineered derivative of 25 amino acid hepcidin that binds TF/CD44v6 and exhibits optimum in vivo properties compared to antibodies or small peptides, including high in vivo stability and rapid clearance through the kidney, thereby increasing tumor retention and reducing non-target organ radiation. Prolonged bioavailability and minimal exposure to healthy tissues may be further accomplished by delivery of HA to ensure internalization of the TF/CD44v6 peptide. The specific aims of the proposed research are to 1): select and characterize TF/CD44v6-avid phage display derived peptides; 2) bioengineer TF/CD44v6-avid peptide(s) into hepcidin scaffolds and characterize biotinylated and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-conjugated forms for their in vitro ovarian cancer cell binding properties; and 3) determine the pharmacokinetics, biodistribution and tumor-targeting ability of both 212Pb and 203Pb-labeled peptide conjugates in vivo in human ovarian carcinoma xenografted mice. The effect of HA treatment on the tumor retention of the radiolabeled hepcidin peptide scaffold will be determined by biodistribution studies. In order to visualize tumor homing, the peptide construct will be labeled with 203Pb for single photon emission computed tomography imaging. The proposed in vivo targeting of ovarian cancers with a-particle labeled peptide scaffolds has wide applicability for the imaging and treatment of ovarian cancer, which is important because early ovarian cancer is often asymptomatic and late stage ovarian cancer is usually fatal. Further, the technology proposed can be applied to target and kill other cancer cells, especially since TF is present on >80% of adenocarcinomas.