Multifunctional siRNA/antibody nanocarriers to treat metastatic triple-negative breast cancer

项目概要/摘要 三阴性乳腺癌（TNBC）是一种侵袭性疾病，生长迅速，早期转移。 对于转移性TNBC没有有效的治疗方法，因此需要新的策略。我们将满足这一需求 使用我们开发的纳米颗粒（NPs）干扰Wnt信号传导，这是TNBC背后的关键驱动力 生长和转移。Wnt信号传导的主要效应物是β-连环蛋白，并且当Wnt信号传导时，β-连环蛋白在TNBC细胞中被激活。 wnt配体结合相对于正常细胞在TNBC细胞上过表达的frizzled 7（FZD 7）受体。 活性β-连环蛋白通过扩增支持TNBC生长和转移的基因的表达来促进TNBC生长和转移。 细胞存活、增殖、干细胞样行为、迁移和侵袭。我们的核心假设是， 在两个水平上抑制Wnt信号传导将降低TNBC细胞的转移潜力，因此我们创造了Wnt 用FZD 7抗体和β-连环蛋白小干扰RNA（siRNA）两者包被的抑制性NP， 这个任务。FZD 7抗体使得能够结合TNBC细胞并防止Wnt信号传导被TNBC细胞激活。 阻断wnt配体结合FZD 7。siRNA还通过直接抑制β-淀粉样蛋白而抑制Wnt信号传导。 通过RNA干扰。我们的初步研究表明，这些纳米粒子可以进入现有的TNBC， 肺转移的小鼠模型，以抑制其生长。在拟议的工作中，我们将阐明 通过研究这些NP与体外TNBC细胞的相互作用，更详细地研究了它们的作用机制（Aim 1），验证它们可以穿透小鼠中现有的转移瘤以阻止它们的生长（目的2），并表明它们 可以安全地预防免疫活性荷瘤小鼠中新转移的形成（目的3）。所有 Aims将比较共同递送FZD 7抗体和β-连环蛋白siRNA的NP与递送任一药剂的NP 以揭示它们的组合效应是相加的还是协同的。除了研究 除了NPs对TNBC细胞的影响外，我们还将评估它们对来自乳腺、肺和乳腺癌的非癌细胞的影响。 骨组织，以确认它们具有可忽略的脱靶效应。我们会在目的3中进一步研究它们的安全性 通过监测动物体重、血液化学和主要器官组织病理学。我们期望发现这些 NP可以抑制TNBC细胞中的Wnt信号传导，以降低其转移潜力，而不影响非肿瘤细胞。 癌细胞，使它们能够被用作治疗现有转移或预防癌细胞形成的工具。 新的转移我们还期望共递送FZD 7抗体和β-连环蛋白siRNA的NP能够更有效地递送FZD 7抗体和β-连环蛋白siRNA。 比单独递送任一药剂的NP有效。总之，本项目将评估Wnt抑制剂 NP作为治疗和/或预防TNBC转移的工具。与目前的疗法不同， 由于结果不充分，我们的NP将直接靶向TNBC细胞，并通过以下方式降低其生长和转移的能力： 抑制Wnt信号传导。如果成功，这种疗法将提高患者的生存率和生活质量。 转移性TNBC，并且它最终可能通过以下方式扩展到具有不同分子驱动的其他癌症： 改变与NP一起递送的抗体和siRNA。

PROJECT SUMMARY/ABSTRACT Triple-negative breast cancer (TNBC) is an aggressive disease that grows quickly and metastasizes early. There are no effective therapies for metastatic TNBC, so new strategies are needed. We will meet this need using nanoparticles (NPs) we developed that interfere with Wnt signaling, the key driving force behind TNBC growth and metastasis. The main effector of Wnt signaling is β-catenin, and it is activated in TNBC cells when wnt ligands bind frizzled7 (FZD7) receptors that are overexpressed on TNBC cells relative to normal cells. Active β-catenin promotes TNBC growth and metastasis by amplifying the expression of genes that support cell survival, proliferation, stem-like behavior, migration, and invasion. Our central hypothesis is that suppressing Wnt signaling at two levels will reduce TNBC cells’ metastatic potential, so we have created Wnt inhibitory NPs coated with both FZD7 antibodies and β-catenin small interfering RNAs (siRNAs) to accomplish this task. The FZD7 antibodies enable TNBC cell binding and prevent Wnt signaling from being activated by blocking wnt ligands from binding FZD7. The siRNAs further suppress Wnt signaling by directly inhibiting β- catenin through RNA interference. Our preliminary studies indicate that these NPs can enter existing TNBC lung metastases in mouse models to inhibit their growth. In the proposed work we will elucidate the mechanism of action of these NPs in greater detail by studying their interaction with TNBC cells in vitro (Aim 1), validate that they can penetrate existing metastases in mice to halt their growth (Aim 2), and show that they can safely prevent the formation of new metastases in immune competent tumor-bearing mice (Aim 3). All aims will compare NPs that co-deliver FZD7 antibodies and β-catenin siRNAs to NPs that deliver either agent individually in order to reveal if their combined effects are additive or synergistic. In addition to studying the NPs’ impact on TNBC cells, we will also evaluate their effect on non-cancerous cells from breast, lung, and bone tissue in order to confirm they have negligible off-target effects. We will further study their safety in Aim 3 by monitoring animal weight, blood chemistry, and major organ histopathology. We expect to find that these NPs can inhibit Wnt signaling in TNBC cells to reduce their metastatic potential without impacting non- cancerous cells, enabling them to be used as tools to either treat existing metastases or prevent formation of new metastases. We also expect for NPs that co-deliver FZD7 antibodies and β-catenin siRNAs to be more effective than NPs that deliver either agent individually. In summary, this project will evaluate Wnt inhibitory NPs as tools to treat and/or prevent TNBC metastasis. Unlike current therapies, which have unpredictable and insufficient results, our NPs will directly target TNBC cells and reduce their ability to grow and metastasize by suppressing Wnt signaling. If successful, this therapy will improve survival and quality of life for patients with metastatic TNBC, and it may ultimately be expanded to other cancers with different molecular drivers by altering the antibodies and siRNAs that are delivered with the NPs.