High-throughput Optimization of Polymeric Nanoparticles for Small RNA Delivery to Treat Glioblastoma

美国每年新诊断出近5万例胶质母细胞瘤(GBM)，其症状令人沮丧 中位生存期14.6个月。目前可用的疗法在很大程度上是无效的，因为遗传， 表观遗传学和基底膜肿瘤内信号的异质性。非编码小RNA，如Short 干扰RNA和微小RNA正在成为细胞命运和肿瘤发生的强有力的表观遗传调节因子 代表了一种有前途的针对肿瘤细胞异质性的量身定制的治疗策略。然而，临床上 小RNA的翻译一直受到关于其安全性和有效性的重大知识差距的限制 向GBM细胞递送。这项研究的总体目标是使用高通量筛选方法来 优化聚β-氨基酯(PBAE)聚合物纳米粒用于治疗性小RNA输送治疗 GBM。我们的初步数据显示，第一代PBAE材料使小RNA能够传递到 体外抑制GBM增殖表型及显著延缓小鼠GBM移植瘤生长 活着。然而，这些纳米颗粒需要在传递效率、生物材料介导的肿瘤方面进行优化。 靶向性，纳米颗粒的长期胶体稳定性，并渗透到整个肿瘤体积，以进一步 临床可译性。为了开发优化的第二代PBAE纳米颗粒配方，拟议的工作 将利用新的高通量方法来产生聚合物结构多样性，并筛选数百 独特的聚合物结构并行，以确定提高效力和癌症靶向的输送材料。在……里面 目标1，创新的体外检测纳米颗粒在克服关键细胞内的性能方面的性能 给药障碍，如纳米颗粒摄取和内体逃逸，将在原发患者- 衍生的GBM细胞模型可以更好地预测纳米颗粒在体内的性能。此外，这些化验结果将 在如何改变生物材料结构以控制其结构方面产生重要的结构-功能关系 以癌症选择性的方式与细胞相互作用。在目标2中，纳米颗粒表面工程技术将 用于增强纳米粒子的稳定性和肿瘤穿透能力。原位肾小球基底膜瘤 将用优化的纳米颗粒配方治疗荷瘤小鼠，以表征纳米颗粒的扩散 在整个肿块中。这对于实现均匀的纳米颗粒递送以及在达到 肿瘤周围的浸润性GBM细胞，它们是肿瘤术后复发的主要原因 治疗。最后，在目标3中，携带两个GBM抑制微RNA的纳米颗粒将被评估其 能够减少GBM的增殖和自我更新。最先进的人类GBM细胞模型将是 用于评估纳米颗粒在体外诱导的表型变化，纳米颗粒也将被注入 荷瘤小鼠评估体内3D肿瘤环境中的治疗递送。这些发现将 对开发可扩展、可生物降解的小RNA递送系统具有实质性的积极影响 脑癌。

Nearly 50,000 new cases of glioblastoma (GBM) are diagnosed in the United States each year, with a dismal median survival of 14.6 months. Currently available therapeutics are largely ineffective due to the genetic, epigenetic, and signaling heterogeneity within the GBM tumor. Non-coding small RNAs such as short interfering RNA and micro-RNA are emerging as potent epigenetic regulators of cell fate and oncogenesis, and represent a promising tailored therapeutic strategy to counter tumor cell heterogeneity. However, clinical translation of small RNAs has been limited by significant knowledge gaps regarding their safe and effective delivery to GBM cells. The overall objective of this study is to use high-throughput screening approaches to optimize poly(beta-amino ester) (PBAE) polymeric nanoparticles for therapeutic small RNA delivery to treat GBM. Our preliminary data have shown that 1st generation PBAE materials enabled small RNA delivery to inhibit GBM proliferative phenotype in vitro and significantly slowed GBM tumor growth in GBM xenografts in vivo. However, these nanoparticles need to be optimized in delivery efficiency, biomaterial-mediated tumor targeting, long-term nanoparticle colloidal stability, and permeation throughout the tumor bulk to further their clinical translatability. To develop optimized 2nd generation PBAE nanoparticle formulations, the proposed work will utilize novel high-throughput approaches to generate polymer structural diversity and screen hundreds of unique polymer structures in parallel to identify delivery materials of improved potency and cancer targeting. In Aim 1, innovative in vitro assays examining nanoparticle performance in overcoming critical intracellular delivery barriers such as nanoparticle uptake and endosomal escape will be performed in primary patient- derived GBM cell models to better predict nanoparticle performance in vivo. Furthermore, these assays will yield important structure-functional relationships on how biomaterial structures can be altered to control their interactions with cells in a cancer-selective manner. In Aim 2, nanoparticle surface engineering techniques will be employed to enhance nanoparticle stability and tumor penetration capabilities. Orthotopic GBM tumor bearing mice will be treated with optimized nanoparticle formulations to characterize nanoparticle diffusion throughout the tumor bulk. This is critical in achieving uniform nanoparticle delivery as well as in reaching infiltrative GBM cells at the tumor periphery, which are primarily responsible for tumor recurrence after treatment. Finally, in Aim 3, nanoparticles carrying two GBM-inhibiting micro-RNAs will be evaluated for their ability to reduce GBM proliferation and self-renewal. State of the art primary human GBM cell models will be used to assess nanoparticle-induced phenotypic changes in vitro, and nanoparticles will also be infused into tumor-bearing mice to assess therapeutic delivery in the 3D tumor environment in vivo. These findings will have substantial positive impact on developing a scalable, bio-degradable small RNA delivery system to treat brain cancer.