Endothelial Cell-Targeted Amatoxin Conjugates for Effective Therapy of Breast Cancer

总结 拟议项目的总体目标是利用新发现的活性跨内皮细胞 运输途径，小窝泵系统，提供了一个有效的解决方案的交付和毒性 转移性乳腺癌治疗中的化疗问题。我们打算开发新的抗体- 利用我们新发现的内皮细胞（EC）小窝靶向系统的药物缀合物（ADC）， 以改善现代化疗药物的递送和靶向。我们已经确定， 在临床前模型中的小窝可以快速和特异性地泵出膜联蛋白A1（AnnA 1）抗体， 货物穿过血管内皮屏障直接进入实体瘤。基于这一发现，我们建议 设计新型EC caveolae靶向ADC，可能会彻底改变转移性乳腺癌的治疗方法。我们 通过用鹅膏毒素武装hAnnA 1，创造了一个强大的传递平台，鹅膏毒素是一类超强效毒素， 能有效杀死快速分裂和休眠的肿瘤细胞。其中包括α-鹅膏毒肽， 从“死亡帽”蘑菇和Chaetocin，一种抗增殖，广谱毒素来自 丝状真菌Chaetomiun sp.我们的主要假设是，靶向小窝的抗体-药物缀合物 （CTA），将大大增加特异性进入肿瘤的超强效毒素的递送，从而需要大量的药物。 更低的剂量和显著提高癌症治疗的功效。这一假设将由 以下具体目标：在目标1中，我们将设计和表征新型hAnnA 1毒素CTA。我们将优化 它们的物理化学性质、稳定性、在肿瘤组织中的释放动力学以及对 人膜联蛋白A1蛋白。在目标2中，我们将评估治疗效果、药物递送和脱靶 使用活体显微术（IVM）的候选CTA在乳腺癌的人肿瘤模型中的作用。在目标3中， 我们将在转移性肿瘤模型和患者源性肿瘤模型中检查所选CTA的治疗效果。 异种移植物（PDX）模型。我们还将为临床试验奠定基础，成功的靶向 治疗药物将被选为未来1期安全性试验的候选药物。肿瘤靶向、递送和 将使用X射线/CT成像、IVM和组织学在啮齿动物中评估这些新疗法的累积。 长期目标是将我们的关键基础发现转化为独特的创新交付平台， 极大地改善了对实体癌的治疗。

SUMMARY The overall objective of the proposed project is to harness a newly discovered, active transendothelial transport pathway, the caveolae pumping system, to provide an effective solution to the delivery and toxicity problem of chemotherapeutics in metastatic breast cancer treatment. We propose to develop novel antibody- drug conjugates (ADCs) that exploit our newly discovered endothelial cell (EC) caveolae targeting system in order to improve the delivery and targeting of modern chemotherapeutics. We have established that EC caveolae in preclinical models can rapidly and specifically pump Annexin A1 (AnnA1) antibodies and attached cargo across the vascular endothelial barrier directly into solid tumors. Based on this discovery, we propose to design novel EC caveolae-targeted ADCs that may revolutionize treatments for metastatic breast cancer. We have created a robust delivery platform by arming hAnnA1 with amatoxins, a class of ultra-potent toxins that can effectively kill both rapidly dividing and dormant tumor cells. These include α-Amanitin, a toxin derived from the “death cap” mushroom and Chaetocin, an anti-proliferative, broad-spectrum toxin derived from the filamentous fungus Chaetomiun sp. Our main hypothesis is that caveolae-targeted antibody-drug conjugates (CTAs), will dramatically increase delivery of ultra-potent toxins specifically into tumors, thereby requiring much lower dosages and dramatically enhancing efficacy of cancer treatment. This hypothesis will be tested by the following specific aims: In Aim 1, we will design and characterize novel hAnnA1-toxin CTAs. We will optimize their physicochemical properties, stability, release kinetics in the tumor interstitium, and binding affinity towards the human Annexin A1 protein. In Aim 2, we will evaluate therapeutic efficacy, drug delivery and off-target effects of candidate CTAs in human tumor models of breast cancer using intravital microscopy (IVM). In Aim 3, we will examine the therapeutic efficacy of the select CTAs in metastatic tumor models and in patient-derived xenograft (PDX) models. We will also prepare a foundation for clinical testing, where successful targeted therapeutics will be selected as candidates for future Phase 1 safety trial. Tumor targeting, delivery and accumulation of these novel therapies will be assessed in rodents using x-ray/CT imaging, IVM, and histology. The long-term goal is to translate our key basic discoveries into a unique, innovative delivery platform that may vastly improve treatments for solid cancers.