Simultaneous targeting of tumor cells and tumor associated macrophages by RNA nanovector

项目摘要/摘要 与其他类型的癌症相比，在治疗方面取得了相当大的进步，治疗性 由于对常规治疗的抵抗，胶质母细胞瘤(GBM)患者的结果仍然令人沮丧。 这种不良反应通常与肿瘤细胞和肿瘤中基因表达的异常失调有关。 支持性肿瘤微环境(TME)，由肿瘤细胞通过免疫编辑进行教育， 这表明由于不利的基因表达，肿瘤还没有为治疗做好准备。为了 启动肿瘤细胞和免疫抑制剂TME以改善GBM患者的临床结局 目前的治疗选择，甚至是新的治疗方法，包括肿瘤细胞和肿瘤相关的巨噬细胞 (TAMS)需要通过逆转异常的基因表达来重新编程 信号通路。肿瘤抑制microRNAs(MiRNAs)已被认为是最合适的 发挥肿瘤重编程作用的治疗靶点。我们最近发现一种可能的肿瘤 抑制性miRNA，miR-138，具有通过下调主要基因表达而重新编程GBM肿瘤细胞的潜能。 通过PD-L1调控细胞增殖和免疫逃避的致癌途径CD44和EGFR 和细胞因子的表达。然而，基于miRNA的治疗进展甚微，这主要是由于 缺乏有效和安全的靶向跨血脑肿瘤屏障(BBTB)进入肿瘤细胞的方法。 此外，肿瘤细胞和TAMs的双重靶向，以传递这种治疗有用的肿瘤 抑制性miRNAs从未被尝试过。此前，我们已经证明了RNA纳米载体(RNV) DNA包装中基于3WJ基序的包装RNA(3WJ-PRNA)平台 噬菌体phi29结合叶酸(FA)的马达(FA-RNV)具有选择性靶向的潜力 叶酸受体(FR)阳性的GBM细胞，并递送具有良好生物分布和 小鼠模型中的药代动力学特征。我们假设靶向递送肿瘤抑制因子miR-138 通过我们的配体连接的RNV进入GBM肿瘤细胞和TAMs将重新编程GBM肿瘤和肿瘤- 相关先天免疫通过直接杀伤肿瘤细胞和敏感性导致更好的肿瘤消退 传统的免疫疗法。在这里，我们建议优化从三向结导出的RNV (3WJ)Motif通过测试不同的细胞特异性配体或RNA适配子来实现这一具有挑战性的任务(目标1)。 此外，我们将测试同时靶向将miR-138导入肿瘤细胞和 细胞特异性RNV的TAMS(目标2)。据我们所知，这是第一个试图 同时调节肿瘤细胞和天然免疫，通过使用 用于GBM治疗的RNV递送系统。我们的研究结果可以迅速转化为人体临床试验 与目前的免疫疗法相结合。

PROJECT SUMMARY/ABSTRACT Compared to other types of cancer that have benefited from considerable advances in therapy, therapeutic outcomes in glioblastoma (GBM) patients still remains dismal due to its resistance to conventional therapies. Such poor response is often related to aberrant dysregulation of gene expressions in tumor cell and tumor- supportive tumor microenvironment (TME) that is educated by the tumor cell through immune editing, indicating that the tumor is not primed for the treatment due to the unfavorable gene expressions. In order to prime the tumor cells and the immunosuppressive TME to improve the clinical outcomes in GBM patients from current treatment options or even new therapeutics, both tumor cells and tumor-associated macrophages (TAMs) need to be reprogrammed by reversing the aberrantly dysregulated gene expressions across various signaling pathways. Tumor suppressive microRNAs (miRNAs) have been proposed as the most suitable therapeutic target to perform the tumor reprogramming role. We recently found that a putative tumor suppressive miRNA, miR-138, has a potential to reprogram GBM tumor cells through down-regulation of major oncogenic pathways CD44 and EGFR, which governs cell proliferation and immune evasion through PD-L1 and cytokine expressions. However, little progress has been made in miRNA-based therapy mainly due to the lack of efficient and safe targeted delivery method into tumor cells across the blood brain tumor barrier (BBTB). In addition, dual targeting of both tumor cells and TAMs to delivery such therapeutically useful tumor suppressive miRNAs has never been attempted. Previously, we have shown that RNA nanovector (RNV) platform based on the Three-Way-Junction (3WJ) motif of packaging RNA (3WJ-pRNA) in the DNA packaging motor of bacteriophage phi29 with conjugation of folate (FA) (FA-RNV) has a potential to selectively target folate receptor (FR) positive GBM cells and deliver a small RNA with favorable bio-distribution and pharmacokinetic profiles in mice model. We hypothesize that targeted delivery of tumor suppressive miR-138 into GBM tumor cells and TAMs by our ligand-conjugated RNV will reprogram both GBM tumor and tumor- associated innate immunity to result better tumor regression through direct tumor cell killing and susceptibility to conventional immunotherapy. Here, we propose to optimze the RNV derived from the three-way junction (3WJ) motif to achieve this challenging task by testing different cell-specific ligand or RNA aptamers (Aim 1). In addition, we will test the preclinical impact of simultaneous targeted delivery of miR-138 into tumor cells and TAMs by cell-specific RNVs (Aim 2). To our knowledge, this is the first preclinical study attempting to simultaneously modulate tumor cells and innate immunity by delivering a tumor suppressive miRNA using the RNV delivery system for GBM treatment. Our study results can be rapidly translated into human clinical trials in a combination with current immunotherapies.